Sulfate determines the glucosinolate concentration of horseradish in vitro plants (Armoracia rusticana Gaertn., Mey. & Scherb.).

PubMed

Alnsour, Mohammad; Kleinwächter, Maik; Böhme, Julia; Selmar, Dirk

2013-03-15

Horseradish plants (Armoracia rusticana) contain high concentrations of glucosinolates. Former studies have revealed that Armoracia plants cultivated in vitro have markedly lower glucosinolate concentrations than those grown in soils. Yet, these studies neglected that the sulfate concentration in the growth medium may have had a strong impact on glucosinolate metabolism. Accordingly, in this study horseradish in vitro plants were cultivated with differing sulfate concentrations and the glucosinolate concentrations were quantified by ion pair HPLC. Cultivation in 1.7 mmol L(-1) sulfate (as used in the prior studies) resulted in the accumulation of 16.2 µmol g(-1) DW glucosinolates, while the glucosinolate concentration increased to more than 23 µmol g(-1) DW when 23.5 mmol L(-1) sulfate was used in the medium. Correspondingly, the glucosinolate concentration decreased to 1.6 µmol g(-1) DW when sulfate concentration was lowered to 0.2 mmol L(-1). Since the glucosinolate accumulation in relation to the sulfate concentration follows a typical saturation curve, we deduce that the availability of sulfate determines the glucosinolate concentration in horseradish in vitro plants. © 2012 Society of Chemical Industry.

Investigation of the multifunctional gene AOP3 expands the regulatory network fine-tuning glucosinolate production in Arabidopsis

PubMed Central

Jensen, Lea M.; Kliebenstein, Daniel J.; Burow, Meike

2015-01-01

Quantitative trait loci (QTL) mapping studies enable identification of loci that are part of regulatory networks controlling various phenotypes. Detailed investigations of genes within these loci are required to ultimately understand the function of individual genes and how they interact with other players in the network. In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds. Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation. Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation. PMID:26442075

Identification and expression pattern analysis of BoMYB51 involved in indolic glucosinolate biosynthesis from broccoli (Brassica oleracea var. italica).

PubMed

Yu, Qingyue; Hao, Guodong; Zhou, Jianxin; Wang, Jingying; Evivie, Ejiroghene Ruona; Li, Jing

2018-06-22

Glucosinolates are a class of amino acid-derived specialized metabolites characteristic of the Brassicales order. Trp derived indolic glucosinolates are essential for the effective plant defense responses to a wide range of pathogens and herbivores. In Arabidopsis, MYB51 is the key transcription factor positively regulates indolic glucosinolate production by activating certain biosynthetic genes. In this study, we report the isolation and identification of a MYB51 from broccoli designated as BoMYB51. Overexpression of BoMYB51 in Arabidopsis increased indolic glucosinolate production by upregulating biosynthetic genes and resulted in enhanced flagellin22 (Flg22) induced callose deposition. The spatial expression pattern and responsive expression of BoMYB51 to several hormones and stress treatments were investigated by expressing the β-glucuronidase (GUS) reporter gene driven by BoMYB51 promotor in Arabidopsis and quantitative real-time PCR analysis in broccoli. Our study provides information on molecular characteristics of BoMYB51 and possible physiological process BoMYB51 may involve. Copyright © 2018 Elsevier Inc. All rights reserved.

Identification and quantification of glucosinolate and flavonol compounds in rocket salad (Eruca sativa, Eruca vesicaria and Diplotaxis tenuifolia) by LC-MS: highlighting the potential for improving nutritional value of rocket crops.

PubMed

Bell, Luke; Oruna-Concha, Maria Jose; Wagstaff, Carol

2015-04-01

Liquid chromatography mass spectrometry (LC-MS) was used to obtain glucosinolate and flavonol content for 35 rocket accessions and commercial varieties. 13 glucosinolates and 11 flavonol compounds were identified. Semi-quantitative methods were used to estimate concentrations of both groups of compounds. Minor glucosinolate composition was found to be different between accessions; concentrations varied significantly. Flavonols showed differentiation between genera, with Diplotaxis accumulating quercetin glucosides and Eruca accumulating kaempferol glucosides. Several compounds were detected in each genus that have only previously been reported in the other. We highlight how knowledge of phytochemical content and concentration can be used to breed new, nutritionally superior varieties. We also demonstrate the effects of controlled environment conditions on the accumulations of glucosinolates and flavonols and explore the reasons for differences with previous studies. We stress the importance of consistent experimental design between research groups to effectively compare and contrast results. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Involvement of a glucosinolate (sinigrin) in the regulation of water transport in Brassica oleracea grown under salt stress.

PubMed

Martínez-Ballesta, Maria del Carmen; Muries, Beatriz; Moreno, Diego Ángel; Dominguez-Perles, Raúl; García-Viguera, Cristina; Carvajal, Micaela

2014-02-01

Members of the Brassicaceae are known for their contents of nutrients and health-promoting phytochemicals, including glucosinolates. The concentrations of these chemopreventive compounds (glucosinolate-degradation products, the bioactive isothiocyanates) may be modified under salinity. In this work, the effect of the aliphatic glucosinolate sinigrin (2-propenyl-glucosinolate) on plant water balance, involving aquaporins, was explored under salt stress. For this purpose, water uptake and its transport through the plasma membrane were determined in plants after NaCl addition, when sinigrin was also supplied. We found higher hydraulic conductance (L0 ) and water permeability (Pf ) and increased abundance of PIP2 aquaporins after the direct administration of sinigrin, showing the ability of the roots to promote cellular water transport across the plasma membrane in spite of the stress conditions imposed. The higher content of the allyl-isothiocyanate and the absence of sinigrin in the plant tissues suggest that the isothiocyanate is related to water balance; in fact, a direct effect of this nitro-sulphate compound on water uptake is proposed. This work provides the first evidence that the addition of a glucosinolate can regulate aquaporins and water transport: this effect and the mechanism(s) involved merit further investigation. © 2013 Scandinavian Plant Physiology Society.

Feeding behaviour of generalist pests on Brassica juncea: implication for manipulation of glucosinolate biosynthesis pathway for enhanced resistance.

PubMed

Kumar, Pawan; Augustine, Rehna; Singh, Amarjeet Kumar; Bisht, Naveen C

2017-10-01

Differential accumulation of plant defence metabolites has been suggested to have important ecological consequence in the context of plant-insect interactions. Feeding of generalist pests on Brassica juncea showed a distinct pattern with selective exclusion of leaf margins which are high in glucosinolates. Molecular basis of this differential accumulation of glucosinolates could be explained based on differential expression profile of BjuMYB28 homologues, the major biosynthetic regulators of aliphatic glucosinolates, as evident from quantitative real-time PCR and promoter:GUS fusion studies in allotetraploid B. juncea. Constitutive overexpression of selected BjuMYB28 homologues enhanced accumulation of aliphatic glucosinolates in B. juncea. Performance of two generalist pests, Helicoverpa armigera and Spodoptera litura larvae, on transgenic B. juncea plants were poor compared to wild-type plants in a no-choice experiment. Correlation coefficient analysis suggested that weight gain of H. armigera larvae was negatively correlated with gluconapin (GNA) and glucobrassicanapin (GBN), whereas that of S. litura larvae was negatively correlated with GNA, GBN and sinigrin (SIN). Our study explains the significance and possible molecular basis of differential distribution of glucosinolates in B. juncea leaves and shows the potential of overexpressing BjuMYB28 for enhanced resistance of Brassica crops against the tested generalist pests. © 2017 John Wiley & Sons Ltd.

Plant defence responses in oilseed rape MINELESS plants after attack by the cabbage moth Mamestra brassicae.

PubMed

Ahuja, Ishita; van Dam, Nicole Marie; Winge, Per; Trælnes, Marianne; Heydarova, Aysel; Rohloff, Jens; Langaas, Mette; Bones, Atle Magnar

2015-02-01

The Brassicaceae family is characterized by a unique defence mechanism known as the 'glucosinolate-myrosinase' system. When insect herbivores attack plant tissues, glucosinolates are hydrolysed by the enzyme myrosinase (EC 3.2.1.147) into a variety of degradation products, which can deter further herbivory. This process has been described as 'the mustard oil bomb'. Additionally, insect damage induces the production of glucosinolates, myrosinase, and other defences. Brassica napus seeds have been genetically modified to remove myrosinase-containing myrosin cells. These plants are termed MINELESS because they lack myrosin cells, the so-called toxic mustard oil mines. Here, we examined the interaction between B. napus wild-type and MINELESS plants and the larvae of the cabbage moth Mamestra brassicae. No-choice feeding experiments showed that M. brassicae larvae gained less weight and showed stunted growth when feeding on MINELESS plants compared to feeding on wild-type plants. M. brassicae feeding didn't affect myrosinase activity in MINELESS plants, but did reduce it in wild-type seedlings. M. brassicae feeding increased the levels of indol-3-yl-methyl, 1-methoxy-indol-3-yl-methyl, and total glucosinolates in both wild-type and MINELESS seedlings. M. brassicae feeding affected the levels of glucosinolate hydrolysis products in both wild-type and MINELESS plants. Transcriptional analysis showed that 494 and 159 genes were differentially regulated after M. brassicae feeding on wild-type and MINELESS seedlings, respectively. Taken together, the outcomes are very interesting in terms of analysing the role of myrosin cells and the glucosinolate-myrosinase defence system in response to a generalist cabbage moth, suggesting that similar studies with other generalist or specialist insect herbivores, including above- and below-ground herbivores, would be useful. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Correlation of glucosinolate content to myrosinase activity in horseradish (Armoracia rusticana).

PubMed

Li, Xian; Kushad, Mosbah M

2004-11-17

Fully developed horseradish (Armoracia rusticana Gaertn., Mey., & Scherb.) roots from 27 accessions and leaves from a subset of 9 accessions were evaluated for glucosinolates and myrosinase enzyme activity. Eight different glucosinolates were detected (based on HPLC retention times as desulfoglucosinolates) in both root and leaf tissues. The sum of these glucosinolates, referred to as total, ranged from 2 to 296 micromol g(-1) of dry weight (DW) in both tissues. Four glucosinolates (sinigrin, glucobrassicin, neoglucobrassicin, and gluconasturtiin) were detected in major quantities. In fully developed roots, sinigrin concentration represented approximately 83%, gluconasturtiin approximately 11%, and glucobrassicin approximately 1% of the total glucosinolates. Approximately the same proportions of individual glucosinolates appeared in fully developed leaves, except that glucobrassicin was substituted by neoglucobrassicin and gluconasturtiin concentration was significantly lower (<1%). At least four other glucosinolates were detected in very small quantities (<1%) in both roots and leaves. Myrosinase (beta-thioglucoside glucohydrolase, EC 3.2.3.1) is the enzyme responsible for the hydrolysis of the parent glucosinolates into biologically active products. Very little is known about myrosinase activity and the correlation of its activity to total and individual glucosinolates in plant tissues. Significant differences in myrosinase activity were detected between the roots and leaves, ranging from 1.2 to 57.1 units g(-1) of DW. Data showed no correlation between myrosinase activity and total and/or individual glucosinolates in the roots. However, in the leaves, significant correlations were found between myrosinase activity and total glucosinolates (0.78 at P = 0.01) and between myrosinase activity and sinigrin (0.80 at P = 0.01). Glucosinolates content and myrosinase activity were also correlated in young and fully developed roots and leaves and during tissue crushing. Glucobrassicin concentration in the roots and neoglucobrassicin concentration in the leaves were significantly higher in young than in fully developed tissue. Crushing of the tissue resulted in rapid hydrolysis of sinigrin and glucobrassicin, as expected, from the presence of myrosinase. Likewise, myrosinase activity declined rapidly after crushing, perhaps due to inactivation by the reaction products and/or the depletion of its substrates.

Intraspecific chemical diversity among neighbouring plants correlates positively with plant size and herbivore load but negatively with herbivore damage.

PubMed

Bustos-Segura, Carlos; Poelman, Erik H; Reichelt, Michael; Gershenzon, Jonathan; Gols, Rieta

2017-01-01

Intraspecific plant diversity can modify the properties of associated arthropod communities and plant fitness. However, it is not well understood which plant traits determine these ecological effects. We explored the effect of intraspecific chemical diversity among neighbouring plants on the associated invertebrate community and plant traits. In a common garden experiment, intraspecific diversity among neighbouring plants was manipulated using three plant populations of wild cabbage that differ in foliar glucosinolates. Plants were larger, harboured more herbivores, but were less damaged when plant diversity was increased. Glucosinolate concentration differentially correlated with generalist and specialist herbivore abundance. Glucosinolate composition correlated with plant damage, while in polycultures, variation in glucosinolate concentrations among neighbouring plants correlated positively with herbivore diversity and negatively with plant damage levels. The results suggest that intraspecific variation in secondary chemistry among neighbouring plants is important in determining the structure of the associated insect community and positively affects plant performance. © 2016 The Authors. Ecology Letters published by CNRS and John Wiley & Sons Ltd.

Development of a reliable extraction and quantification method for glucosinolates in Moringa oleifera.

PubMed

Förster, Nadja; Ulrichs, Christian; Schreiner, Monika; Müller, Carsten T; Mewis, Inga

2015-01-01

Glucosinolates are the characteristic secondary metabolites of plants in the order Brassicales. To date the common DIN extraction 'desulfo glucosinolates' method remains the common procedure for determination and quantification of glucosinolates. However, the desulfation step in the extraction of glucosinolates from Moringa oleifera leaves resulted in complete conversion and degradation of the naturally occurring glucosinolates in this plant. Therefore, a method for extraction of intact Moringa glucosinolates was developed and no conversion and degradation of the different rhamnopyranosyloxy-benzyl glucosinolates was found. Buffered eluents (0.1 M ammonium acetate) were necessary to stabilize 4-α-rhamnopyranosyloxy-benzyl glucosinolate (Rhamno-Benzyl-GS) and acetyl-4-α-rhamnopyranosyloxy-benzyl glucosinolate isomers (Ac-Isomers-GS) during HPLC analysis. Due to the instability of intact Moringa glucosinolates at room temperature and during the purification process of single glucosinolates, influences of different storage (room temperature, frozen, thawing and refreezing) and buffer conditions on glucosinolate conversion were analysed. Conversion and degradations processes were especially determined for the Ac-Isomers-GS III. Copyright © 2014 Elsevier Ltd. All rights reserved.

Integration of Biosynthesis and Long-Distance Transport Establish Organ-Specific Glucosinolate Profiles in Vegetative Arabidopsis[W

PubMed Central

Andersen, Tonni Grube; Nour-Eldin, Hussam Hassan; Fuller, Victoria Louise; Olsen, Carl Erik; Burow, Meike; Halkier, Barbara Ann

2013-01-01

Although it is essential for plant survival to synthesize and transport defense compounds, little is known about the coordination of these processes. Here, we investigate the above- and belowground source-sink relationship of the defense compounds glucosinolates in vegetative Arabidopsis thaliana. In vivo feeding experiments demonstrate that the glucosinolate transporters1 and 2 (GTR1 and GTR2), which are essential for accumulation of glucosinolates in seeds, are likely to also be involved in bidirectional distribution of glucosinolates between the roots and rosettes, indicating phloem and xylem as their transport pathways. Grafting of wild-type, biosynthetic, and transport mutants show that both the rosette and roots are able to synthesize aliphatic and indole glucosinolates. While rosettes constitute the major source and storage site for short-chained aliphatic glucosinolates, long-chained aliphatic glucosinolates are synthesized both in roots and rosettes with roots as the major storage site. Our grafting experiments thus indicate that in vegetative Arabidopsis, GTR1 and GTR2 are involved in bidirectional long-distance transport of aliphatic but not indole glucosinolates. Our data further suggest that the distinct rosette and root glucosinolate profiles in Arabidopsis are shaped by long-distance transport and spatially separated biosynthesis, suggesting that integration of these processes is critical for plant fitness in complex natural environments. PMID:23995084

Glucosinolates from Host Plants Influence Growth of the Parasitic Plant Cuscuta gronovii and Its Susceptibility to Aphid Feeding1[OPEN

PubMed Central

2016-01-01

Parasitic plants acquire diverse secondary metabolites from their hosts, including defense compounds that target insect herbivores. However, the ecological implications of this phenomenon, including the potential enhancement of parasite defenses, remain largely unexplored. We studied the translocation of glucosinolates from the brassicaceous host plant Arabidopsis (Arabidopsis thaliana) into parasitic dodder vines (Convolvulaceae; Cuscuta gronovii) and its effects on the parasite itself and on dodder-aphid interactions. Aliphatic and indole glucosinolates reached concentrations in parasite tissues higher than those observed in corresponding host tissues. Dodder growth was enhanced on cyp79B2 cyp79B3 hosts (without indole glucosinolates) but inhibited on atr1D hosts (with elevated indole glucosinolates) relative to wild-type hosts, which responded to parasitism with localized elevation of indole and aliphatic glucosinolates. These findings implicate indole glucosinolates in defense against parasitic plants. Rates of settling and survival on dodder vines by pea aphids (Acyrthosiphon pisum) were reduced significantly when dodder parasitized glucosinolate-producing hosts (wild type and atr1D) compared with glucosinolate-free hosts (cyp79B2 cyp79B3 myb28 myb29). However, settling and survival of green peach aphids (Myzus persicae) were not affected. M. persicae population growth was actually reduced on dodder parasitizing glucosinolate-free hosts compared with wild-type or atr1D hosts, even though stems of the former contain less glucosinolates and more amino acids. Strikingly, this effect was reversed when the aphids fed directly upon Arabidopsis, which indicates an interactive effect of parasite and host genotype on M. persicae that stems from host effects on dodder. Thus, our findings indicate that glucosinolates may have both direct and indirect effects on dodder-feeding herbivores. PMID:27482077

CYP79F1 and CYP79F2 have distinct functions in the biosynthesis of aliphatic glucosinolates in Arabidopsis.

PubMed

Chen, Sixue; Glawischnig, Erich; Jørgensen, Kirsten; Naur, Peter; Jørgensen, Bodil; Olsen, Carl-Erik; Hansen, Carsten H; Rasmussen, Hasse; Pickett, John A; Halkier, Barbara A

2003-03-01

Cytochromes P450 of the CYP79 family catalyze the conversion of amino acids to oximes in the biosynthesis of glucosinolates, a group of natural plant products known to be involved in plant defense and as a source of flavor compounds, cancer-preventing agents and bioherbicides. We report a detailed biochemical analysis of the substrate specificity and kinetics of CYP79F1 and CYP79F2, two cytochromes P450 involved in the biosynthesis of aliphatic glucosinolates in Arabidopsis thaliana. Using recombinant CYP79F1 and CYP79F2 expressed in Escherichia coli and Saccharomyces cerevisiae, respectively, we show that CYP79F1 metabolizes mono- to hexahomomethionine, resulting in both short- and long-chain aliphatic glucosinolates. In contrast, CYP79F2 exclusively metabolizes long-chain elongated penta- and hexahomomethionines. CYP79F1 and CYP79F2 are spatially and developmentally regulated, with different gene expression patterns. CYP79F2 is highly expressed in hypocotyl and roots, whereas CYP79F1 is strongly expressed in cotyledons, rosette leaves, stems, and siliques. A transposon-tagged CYP79F1 knockout mutant completely lacks short-chain aliphatic glucosinolates, but has an increased level of long-chain aliphatic glucosinolates, especially in leaves and seeds. The level of long-chain aliphatic glucosinolates in a transposon-tagged CYP79F2 knockout mutant is substantially reduced, whereas the level of short-chain aliphatic glucosinolates is not affected. Biochemical characterization of CYP79F1 and CYP79F2, and gene expression analysis, combined with glucosinolate profiling of knockout mutants demonstrate the functional role of these enzymes. This provides valuable insights into the metabolic network leading to the biosynthesis of aliphatic glucosinolates, and into metabolic engineering of altered aliphatic glucosinolate profiles to improve nutritional value and pest resistance.

Glucosinolates from Host Plants Influence Growth of the Parasitic Plant Cuscuta gronovii and Its Susceptibility to Aphid Feeding.

PubMed

Smith, Jason D; Woldemariam, Melkamu G; Mescher, Mark C; Jander, Georg; De Moraes, Consuelo M

2016-09-01

Parasitic plants acquire diverse secondary metabolites from their hosts, including defense compounds that target insect herbivores. However, the ecological implications of this phenomenon, including the potential enhancement of parasite defenses, remain largely unexplored. We studied the translocation of glucosinolates from the brassicaceous host plant Arabidopsis (Arabidopsis thaliana) into parasitic dodder vines (Convolvulaceae; Cuscuta gronovii) and its effects on the parasite itself and on dodder-aphid interactions. Aliphatic and indole glucosinolates reached concentrations in parasite tissues higher than those observed in corresponding host tissues. Dodder growth was enhanced on cyp79B2 cyp79B3 hosts (without indole glucosinolates) but inhibited on atr1D hosts (with elevated indole glucosinolates) relative to wild-type hosts, which responded to parasitism with localized elevation of indole and aliphatic glucosinolates. These findings implicate indole glucosinolates in defense against parasitic plants. Rates of settling and survival on dodder vines by pea aphids (Acyrthosiphon pisum) were reduced significantly when dodder parasitized glucosinolate-producing hosts (wild type and atr1D) compared with glucosinolate-free hosts (cyp79B2 cyp79B3 myb28 myb29). However, settling and survival of green peach aphids (Myzus persicae) were not affected. M. persicae population growth was actually reduced on dodder parasitizing glucosinolate-free hosts compared with wild-type or atr1D hosts, even though stems of the former contain less glucosinolates and more amino acids. Strikingly, this effect was reversed when the aphids fed directly upon Arabidopsis, which indicates an interactive effect of parasite and host genotype on M. persicae that stems from host effects on dodder. Thus, our findings indicate that glucosinolates may have both direct and indirect effects on dodder-feeding herbivores. © 2016 American Society of Plant Biologists. All rights reserved.

Extraction and analysis of intact glucosinolates--a validated pressurized liquid extraction/liquid chromatography-mass spectrometry protocol for Isatis tinctoria, and qualitative analysis of other cruciferous plants.

PubMed

Mohn, Tobias; Cutting, Brian; Ernst, Beat; Hamburger, Matthias

2007-09-28

Glucosinolates have attracted significant interest due to the chemopreventive properties of some of their transformation products. Numerous protocols for the extraction and analysis of glucosinolates have been published, but limited effort has been devoted to optimize and validate crucial extraction parameters and sample preparation steps. We carried out a systematic optimization and validation of a quantitative assay for the direct analysis of intact glucosinolates in Isatis tinctoria leaves (woad, Brassicaceae). Various parameters such as solvent composition, particle size, temperature, and number of required extraction steps were optimized using pressurized liquid extraction (PLE). We observed thermal degradation of glucosinolates at temperatures above 50 degrees C, and loss of >60% within 10min at 100 degrees C, but no enzymatic degradation in the leaf samples at ambient temperature. Excellent peak shape and resolution was obtained by reversed-phase chromatography on a Phenomenex Aqua column using 10mM ammonium formate as ion-pair reagent. Detection was carried out by electrospray ionisation mass spectrometry in the negative ion mode. Analysis of cruciferous vegetables and spices such as broccoli (Brassica oleracea L. var. italica), garden cress (Lepidium sativum L.) and black mustard (Sinapis nigra L.) demonstrated the general applicability of the method.

Glucosinolate biosynthesis in hairy root cultures of broccoli (Brassica oleracea var. italica).

PubMed

Kim, Sun-Ju; Park, Woo Tae; Uddin, Md Romij; Kim, Yeon Bok; Nam, Sang-Yong; Jho, Kwang Hyun; Park, Sang Un

2013-02-01

Here we present previously unreported glucosinolate production by hairy root cultures of broccoli (B. oleracea var. italica). Growth media greatly influenced the growth and glucosinolate content of hairy root cultures of broccoli. Seven glucosinolates, glucoraphanin, gluconapin, glucoerucin, glucobrassicin, 4-methoxyglucobrassicin, gluconasturtiin, and neoglucobrassicin, were identified by analysis of the broccoli hairy root cultures. Both half and full strength B5 and SH media enabled the highest accumulation of glucosinolates. In most cases, the levels of glucosinolates were higher in SH and BS media. Among the 7 glucosinolates, the accumulation of neoglucobrassicin was very high, irrespective of growth medium. The neoglucobrassicin content was 7.4-fold higher in SH medium than 1/2 MS, in which its level was the lowest. The 1/2 B5 medium supported the production of the highest amounts of glucobrassicin and 4-methoxyglucobrassicin, the levels for which were 36.2- and 7.9- fold higher, respectively, than their lowest content in 1/2 MS medium. The 1/2 SH medium enabled the highest accumulation of glucoraphanin and gluconapin in the broccoli hairy root cultures, whose levels were 1.8- and 4.6-fold higher, respectively, than their lowest content in 1/2 MS medium. Our results suggest that hairy root cultures of broccoli could be a valuable alternative approach for the production of glucosinolate compounds.

Effects of different cooking methods on health-promoting compounds of broccoli*

PubMed Central

Yuan, Gao-feng; Sun, Bo; Yuan, Jing; Wang, Qiao-mei

2009-01-01

The effects of five domestic cooking methods, including steaming, microwaving, boiling, stir-frying, and stir-frying followed by boiling (stir-frying/boiling), on the nutrients and health-promoting compounds of broccoli were investigated. The results show that all cooking treatments, except steaming, caused significant losses of chlorophyll and vitamin C and significant decreases of total soluble proteins and soluble sugars. Total aliphatic and indole glucosinolates were significantly modified by all cooking treatments but not by steaming. In general, the steaming led to the lowest loss of total glucosinolates, while stir-frying and stir-frying/boiling presented the highest loss. Stir-frying and stir-frying/boiling, the two most popular methods for most homemade dishes in China, cause great losses of chlorophyll, soluble protein, soluble sugar, vitamin C, and glucosinolates, but the steaming method appears the best in retention of the nutrients in cooking broccoli. PMID:19650196

Metabolomic assessment reveals an elevated level of glucosinolate content in CaCl₂ treated broccoli microgreens.

PubMed

Sun, Jianghao; Kou, Liping; Geng, Ping; Huang, Huilian; Yang, Tianbao; Luo, Yaguang; Chen, Pei

2015-02-18

Preharvest calcium application has been shown to increase broccoli microgreen yield and extend shelf life. In this study, we investigated the effect of calcium application on its metabolome using ultra-high-performance liquid chromatography with mass spectrometry. The data collected were analyzed using principal component analysis and orthogonal projection to latent structural discriminate analysis. Chemical composition comparison shows that glucosinolates, a very important group of phytochemicals, are the major compounds enhanced by preharvest treatment with 10 mM calcium chloride (CaCl2). Aliphatic glucosinolates (glucoerucin, glucoiberin, glucoiberverin, glucoraphanin, pentyl glucosinolate, and hexyl glucosinolate) and indolic glucosinolates (glucobrassicin, neoglucobrassicin, and 4-hydroxyglucobrassicin) were increased significantly in the CaCl2 treated microgreens using metabolomic approaches. Targeted glucosinolate analysis using the ISO 9167-1 method was further employed to confirm the findings. Results indicate that glucosinolates can be considered as a class of compounds that are responsible for the difference between two groups and a higher glucosinolate level was found in CaCl2 treated groups at each time point after harvest in comparison with the control group.

Plant defence responses in oilseed rape MINELESS plants after attack by the cabbage moth Mamestra brassicae

PubMed Central

Ahuja, Ishita; van Dam, Nicole Marie; Winge, Per; Trælnes, Marianne; Heydarova, Aysel; Rohloff, Jens; Langaas, Mette; Bones, Atle Magnar

2015-01-01

The Brassicaceae family is characterized by a unique defence mechanism known as the ‘glucosinolate–myrosinase’ system. When insect herbivores attack plant tissues, glucosinolates are hydrolysed by the enzyme myrosinase (EC 3.2.1.147) into a variety of degradation products, which can deter further herbivory. This process has been described as ‘the mustard oil bomb’. Additionally, insect damage induces the production of glucosinolates, myrosinase, and other defences. Brassica napus seeds have been genetically modified to remove myrosinase-containing myrosin cells. These plants are termed MINELESS because they lack myrosin cells, the so-called toxic mustard oil mines. Here, we examined the interaction between B. napus wild-type and MINELESS plants and the larvae of the cabbage moth Mamestra brassicae. No-choice feeding experiments showed that M. brassicae larvae gained less weight and showed stunted growth when feeding on MINELESS plants compared to feeding on wild-type plants. M. brassicae feeding didn’t affect myrosinase activity in MINELESS plants, but did reduce it in wild-type seedlings. M. brassicae feeding increased the levels of indol-3-yl-methyl, 1-methoxy-indol-3-yl-methyl, and total glucosinolates in both wild-type and MINELESS seedlings. M. brassicae feeding affected the levels of glucosinolate hydrolysis products in both wild-type and MINELESS plants. Transcriptional analysis showed that 494 and 159 genes were differentially regulated after M. brassicae feeding on wild-type and MINELESS seedlings, respectively. Taken together, the outcomes are very interesting in terms of analysing the role of myrosin cells and the glucosinolate–myrosinase defence system in response to a generalist cabbage moth, suggesting that similar studies with other generalist or specialist insect herbivores, including above- and below-ground herbivores, would be useful. PMID:25563968

De novo Transcriptome Assembly of Chinese Kale and Global Expression Analysis of Genes Involved in Glucosinolate Metabolism in Multiple Tissues

PubMed Central

Wu, Shuanghua; Lei, Jianjun; Chen, Guoju; Chen, Hancai; Cao, Bihao; Chen, Changming

2017-01-01

Chinese kale, a vegetable of the cruciferous family, is a popular crop in southern China and Southeast Asia due to its high glucosinolate content and nutritional qualities. However, there is little research on the molecular genetics and genes involved in glucosinolate metabolism and its regulation in Chinese kale. In this study, we sequenced and characterized the transcriptomes and expression profiles of genes expressed in 11 tissues of Chinese kale. A total of 216 million 150-bp clean reads were generated using RNA-sequencing technology. From the sequences, 98,180 unigenes were assembled for the whole plant, and 49,582~98,423 unigenes were assembled for each tissue. Blast analysis indicated that a total of 80,688 (82.18%) unigenes exhibited similarity to known proteins. The functional annotation and classification tools used in this study suggested that genes principally expressed in Chinese kale, were mostly involved in fundamental processes, such as cellular and molecular functions, the signal transduction, and biosynthesis of secondary metabolites. The expression levels of all unigenes were analyzed in various tissues of Chinese kale. A large number of candidate genes involved in glucosinolate metabolism and its regulation were identified, and the expression patterns of these genes were analyzed. We found that most of the genes involved in glucosinolate biosynthesis were highly expressed in the root, petiole, and in senescent leaves. The expression patterns of ten glucosinolate biosynthetic genes from RNA-seq were validated by quantitative RT-PCR in different tissues. These results provided an initial and global overview of Chinese kale gene functions and expression activities in different tissues. PMID:28228764

Variations in the most abundant types of glucosinolates found in the leaves of baby leaf rocket under typical commercial conditions.

PubMed

Hall, Matthew K D; Jobling, Jenny J; Rogers, Gordon S

2015-02-01

Changes in the concentration of the three most abundant glucosinolates were measured in the leaves of perennial wall rocket [Diplotaxis tenuifolia (L.) DC.], and annual garden rocket (Eruca sativa Mill.). HPLC-MS was used to identify glucoraphanin, 4-hydroxyglucobrassin and glucoerucin from perennial wall rocket, and glucoraphanin, glucobrassicin and 4-methoxyglucobrassicin from annual garden rocket. In separate experiments the responses of glucosinolates to harvest number, seasonal conditions, nitrogen supply and post-harvest storage conditions were measured. For perennial wall rocket, season influenced the concentration of glucoraphanin, which were highest for the spring [379 µg kg(-1) fresh weight (FW)] and summer (317 µg kg(-1) FW) plantings. The concentration of 4-hydroxyglucobrassin was higher in the leaves of first harvest crops. This response was due to this glucosinolate not being detected in the leaves of second harvest crops. Thus, the parent glucosinolate was altered between the first and second harvests in response to the abiotic stresses caused by harvesting. For annual garden rocket, there was an interaction between the harvest number and season for all glucosinolates measured. However, no clear response was observed between these factors. Higher concentrations of glucobrassicin and 4-methoxyglucobrassicin were measured for first harvest leaves when compared to the second harvest. This was due to the absence of detection of these glucosinolates in the leaves of second harvested plants; consequently higher total glucosinolate concentrations were measured for the first harvest winter (1224 µg kg(-1) FW) and summer (864 µg kg(-1) FW) crops. The concentrations of individual glucosinolates vary greatly over typical pre- and post-harvest commercial conditions. The absence of 4-hydroxyglucobrassin for perennial wall rocket, and glucobrassicin and 4-methoxyglucobrassicin for annual garden rocket between harvests, illustrates that abiotic stress from harvesting has the capacity to alter the types of glucosinolates in leaves. Concentrations do not generally decline during a typical storage period, indicating that the potential benefits of these compounds are not lost during post-harvest storage. © 2014 Society of Chemical Industry.

HPLC-Based Method to Evaluate Kinetics of Glucosinolate Hydrolysis by Sinapis alba Myrosinase1

PubMed Central

Vastenhout, Kayla J.; Tornberg, Ruthellen H.; Johnson, Amanda L.; Amolins, Michael W.; Mays, Jared R.

2014-01-01

Isothiocyanates (ITCs) are one of several hydrolysis products of glucosinolates, plant secondary metabolites which are substrates for the thioglucohydrolase myrosinase. Recent pursuits toward the development of synthetic, non-natural ITCs have consequently led to an exploration of generating these compounds from non-natural glucosinolate precursors. Evaluation of the myrosinase-dependent conversion of select non-natural glucosinolates to non-natural ITCs cannot be accomplished using established UV-Vis spectroscopic methods. To overcome this limitation, an alternative HPLC-based analytical approach was developed where initial reaction velocities were generated from non-linear reaction progress curves. Validation of this HPLC method was accomplished through parallel evaluation of three glucosinolates with UV-Vis methodology. The results of this study demonstrate that kinetic data is consistent between both analytical methods and that the tested glucosinolates respond similarly to both Michaelis–Menten and specific activity analyses. Consequently, this work resulted in the complete kinetic characterization of three glucosinolates with Sinapis alba myrosinase, with results that were consistent with previous reports. PMID:25068719

NSP-Dependent Simple Nitrile Formation Dominates upon Breakdown of Major Aliphatic Glucosinolates in Roots, Seeds, and Seedlings of Arabidopsis thaliana Columbia-0.

PubMed

Wittstock, Ute; Meier, Kathrin; Dörr, Friederike; Ravindran, Beena M

2016-01-01

One of the best-studied plant defense systems, the glucosinolate-myrosinase system of the Brassicales, is composed of thioglucosides known as glucosinolates and their hydrolytic enzymes, the myrosinases. Tissue disruption brings these components together, and bioactive products are formed as a consequence of myrosinase-catalyzed glucosinolate hydrolysis. Among these products, isothiocyanates have attracted most interest as chemical plant defenses against herbivores and pathogens and health-promoting compounds in the human diet. Previous research has identified specifier proteins whose presence results in the formation of alternative product types, e.g., nitriles, at the expense of isothiocyanates. The biological roles of specifier proteins and alternative breakdown products are poorly understood. Here, we assessed glucosinolate breakdown product profiles obtained upon maceration of roots, seedlings and seeds of Arabidopsis thaliana Columbia-0. We identified simple nitriles as the predominant breakdown products of the major endogenous aliphatic glucosinolates in root, seed, and seedling homogenates. In agreement with this finding, genes encoding nitrile-specifier proteins (NSPs) are expressed in roots, seeds, and seedlings. Analysis of glucosinolate breakdown in mutants with T-DNA insertions in any of the five NSP genes demonstrated, that simple nitrile formation upon tissue disruption depended almost entirely on NSP2 in seeds and mainly on NSP1 in seedlings. In roots, about 70-80% of the nitrile-forming activity was due to NSP1 and NSP3 . Thus, glucosinolate breakdown product profiles are organ-specifically regulated in A. thaliana Col-0, and high proportions of simple nitriles are formed in some parts of the plant. This should be considered in future studies on biological roles of the glucosinolate-myrosinase system.

NSP-Dependent Simple Nitrile Formation Dominates upon Breakdown of Major Aliphatic Glucosinolates in Roots, Seeds, and Seedlings of Arabidopsis thaliana Columbia-0

PubMed Central

Wittstock, Ute; Meier, Kathrin; Dörr, Friederike; Ravindran, Beena M.

2016-01-01

One of the best-studied plant defense systems, the glucosinolate-myrosinase system of the Brassicales, is composed of thioglucosides known as glucosinolates and their hydrolytic enzymes, the myrosinases. Tissue disruption brings these components together, and bioactive products are formed as a consequence of myrosinase-catalyzed glucosinolate hydrolysis. Among these products, isothiocyanates have attracted most interest as chemical plant defenses against herbivores and pathogens and health-promoting compounds in the human diet. Previous research has identified specifier proteins whose presence results in the formation of alternative product types, e.g., nitriles, at the expense of isothiocyanates. The biological roles of specifier proteins and alternative breakdown products are poorly understood. Here, we assessed glucosinolate breakdown product profiles obtained upon maceration of roots, seedlings and seeds of Arabidopsis thaliana Columbia-0. We identified simple nitriles as the predominant breakdown products of the major endogenous aliphatic glucosinolates in root, seed, and seedling homogenates. In agreement with this finding, genes encoding nitrile-specifier proteins (NSPs) are expressed in roots, seeds, and seedlings. Analysis of glucosinolate breakdown in mutants with T-DNA insertions in any of the five NSP genes demonstrated, that simple nitrile formation upon tissue disruption depended almost entirely on NSP2 in seeds and mainly on NSP1 in seedlings. In roots, about 70–80% of the nitrile-forming activity was due to NSP1 and NSP3. Thus, glucosinolate breakdown product profiles are organ-specifically regulated in A. thaliana Col-0, and high proportions of simple nitriles are formed in some parts of the plant. This should be considered in future studies on biological roles of the glucosinolate-myrosinase system. PMID:27990154

[Determination of benzyl glucosinolate in Lepidium meyenii from different regions by HPLC].

PubMed

Tang, Lin; Yin, Hong-jun; Si, Cong-cong; Hu, Xiao-yan; Long, Zheng-hai

2015-12-01

The content of benzyl isothiocyanate (BITC) which as the enzymatic hydrolysis product of benzyl glucosinolate through thioglucosidase was determined by HPLC. The content of benzyl isothiocyanate (BITC) which as the enzymatic hydrolysis product of benzyl glucosinolate through thioglucosidase was determined by HPLC. The chromatography condition was as follows: Kaseisorb LC ODS 2000 (4.6 mm x 150 mm, 5 min) column with the mobile phase of acetonitrile(A)-water( B) under gradient elution (0-5 min, 3%-8% A; 5-9 min, 8%-48% A; 9-23 min, 48%-62% A; 23-28 min, 62%-99% A); the flow rate was 1.0 mL x min(-1) with 10 microL injection volume; detection wavelength was 246 nm and temperature of column was 40 degrees C. The content of benzyl glucosinolate was in the range of 10.76-17.91 g x L(-1). The method is simple, accurate and good reproducibility which can be used for the determination of benzyl glucosinolate in Lepidium meyenii, effectively.

Molecular Characterization of MYB28 Involved in Aliphatic Glucosinolate Biosynthesis in Chinese Kale (Brassica oleracea var. alboglabra Bailey).

PubMed

Yin, Ling; Chen, Hancai; Cao, Bihao; Lei, Jianjun; Chen, Guoju

2017-01-01

Glucosinolates are Brassicaceae-specific secondary metabolites that act as crop protectants, flavor precursors, and cancer-prevention agents, which shows strong evidences of anticarcinogentic, antioxidant, and antimicrobial activities. MYB28 , the R2R3-MYB28 transcription factor, directly activates genes involved in aliphatic glucosinolate biosynthesis. In this study, the MYB28 homology ( BoaMYB28 ) was identified in Chinese kale ( Brassica oleracea var. alboglabra Bailey). Analysis of the nucleotide sequence indicated that the cDNA of BoaMYB28 was 1257 bp with an ORF of 1020 bp. The deduced BoaMYB28 protein was a polypeptide of 339 amino acid with a putative molecular mass of 38 kDa and a pI of 6.87. Sequence homology and phylogenetic analysis showed that BoaMYB28 was most closely related to MYB28 homologs from the Brassicaceae family. The expression levels of BoaMYB28 varies across the tissues and developmental stages. BoaMYB28 transcript levels were higher in leaves and stems compared with those in cotyledons, flowers, and siliques. BoaMYB28 was expressed across all developmental leaf stages, with higher transcript accumulation in mature and inflorescence leaves. Over-expression and RNAi studies showed that BoaMYB28 retains the basic MYB28 gene function as a major transcriptional regulator of aliphatic glucosinolate pathway. The results indicated that over-expression and RNAi lines showed no visible difference on plant morphology. The contents of aliphatic glucosinolates and transcript levels of aliphatic glucosinolate biosynthesis genes increased in over-expression lines and decreased in RNAi lines. In over-expression lines, aliphatic glucosinolate contents were 1.5- to 3-fold higher than those in the wild-type, while expression levels of aliphatic glucosinolate biosynthesis genes were 1.5- to 4-fold higher than those in the wild-type. In contrast, the contents of aliphatic glucosinolates and transcript levels of aliphatic glucosinolate biosynthesis genes in RNAi lines were considerably lower than those in the wild-type. The results suggest that BoaMYB28 has the potential to alter the aliphatic glucosinolates contents in Chinese kale at the genetic level.

Molecular Characterization of MYB28 Involved in Aliphatic Glucosinolate Biosynthesis in Chinese Kale (Brassica oleracea var. alboglabra Bailey)

PubMed Central

Yin, Ling; Chen, Hancai; Cao, Bihao; Lei, Jianjun; Chen, Guoju

2017-01-01

Glucosinolates are Brassicaceae-specific secondary metabolites that act as crop protectants, flavor precursors, and cancer-prevention agents, which shows strong evidences of anticarcinogentic, antioxidant, and antimicrobial activities. MYB28, the R2R3-MYB28 transcription factor, directly activates genes involved in aliphatic glucosinolate biosynthesis. In this study, the MYB28 homology (BoaMYB28) was identified in Chinese kale (Brassica oleracea var. alboglabra Bailey). Analysis of the nucleotide sequence indicated that the cDNA of BoaMYB28 was 1257 bp with an ORF of 1020 bp. The deduced BoaMYB28 protein was a polypeptide of 339 amino acid with a putative molecular mass of 38 kDa and a pI of 6.87. Sequence homology and phylogenetic analysis showed that BoaMYB28 was most closely related to MYB28 homologs from the Brassicaceae family. The expression levels of BoaMYB28 varies across the tissues and developmental stages. BoaMYB28 transcript levels were higher in leaves and stems compared with those in cotyledons, flowers, and siliques. BoaMYB28 was expressed across all developmental leaf stages, with higher transcript accumulation in mature and inflorescence leaves. Over-expression and RNAi studies showed that BoaMYB28 retains the basic MYB28 gene function as a major transcriptional regulator of aliphatic glucosinolate pathway. The results indicated that over-expression and RNAi lines showed no visible difference on plant morphology. The contents of aliphatic glucosinolates and transcript levels of aliphatic glucosinolate biosynthesis genes increased in over-expression lines and decreased in RNAi lines. In over-expression lines, aliphatic glucosinolate contents were 1.5- to 3-fold higher than those in the wild-type, while expression levels of aliphatic glucosinolate biosynthesis genes were 1.5- to 4-fold higher than those in the wild-type. In contrast, the contents of aliphatic glucosinolates and transcript levels of aliphatic glucosinolate biosynthesis genes in RNAi lines were considerably lower than those in the wild-type. The results suggest that BoaMYB28 has the potential to alter the aliphatic glucosinolates contents in Chinese kale at the genetic level. PMID:28680435

Phyllotreta striolata flea beetles use host plant defense compounds to create their own glucosinolate-myrosinase system

PubMed Central

Beran, Franziska; Pauchet, Yannick; Kunert, Grit; Reichelt, Michael; Wielsch, Natalie; Vogel, Heiko; Reinecke, Andreas; Svatoš, Aleš; Mewis, Inga; Schmid, Daniela; Ramasamy, Srinivasan; Ulrichs, Christian; Hansson, Bill S.; Gershenzon, Jonathan; Heckel, David G.

2014-01-01

The ability of a specialized herbivore to overcome the chemical defense of a particular plant taxon not only makes it accessible as a food source but may also provide metabolites to be exploited for communication or chemical defense. Phyllotreta flea beetles are adapted to crucifer plants (Brassicales) that are defended by the glucosinolate-myrosinase system, the so-called “mustard-oil bomb.” Tissue damage caused by insect feeding brings glucosinolates into contact with the plant enzyme myrosinase, which hydrolyzes them to form toxic compounds, such as isothiocyanates. However, we previously observed that Phyllotreta striolata beetles themselves produce volatile glucosinolate hydrolysis products. Here, we show that P. striolata adults selectively accumulate glucosinolates from their food plants to up to 1.75% of their body weight and express their own myrosinase. By combining proteomics and transcriptomics, a gene responsible for myrosinase activity in P. striolata was identified. The major substrates of the heterologously expressed myrosinase were aliphatic glucosinolates, which were hydrolyzed with at least fourfold higher efficiency than aromatic and indolic glucosinolates, and β-O-glucosides. The identified beetle myrosinase belongs to the glycoside hydrolase family 1 and has up to 76% sequence similarity to other β-glucosidases. Phylogenetic analyses suggest species-specific diversification of this gene family in insects and an independent evolution of the beetle myrosinase from other insect β-glucosidases. PMID:24799680

Phyllotreta striolata flea beetles use host plant defense compounds to create their own glucosinolate-myrosinase system.

PubMed

Beran, Franziska; Pauchet, Yannick; Kunert, Grit; Reichelt, Michael; Wielsch, Natalie; Vogel, Heiko; Reinecke, Andreas; Svatoš, Aleš; Mewis, Inga; Schmid, Daniela; Ramasamy, Srinivasan; Ulrichs, Christian; Hansson, Bill S; Gershenzon, Jonathan; Heckel, David G

2014-05-20

The ability of a specialized herbivore to overcome the chemical defense of a particular plant taxon not only makes it accessible as a food source but may also provide metabolites to be exploited for communication or chemical defense. Phyllotreta flea beetles are adapted to crucifer plants (Brassicales) that are defended by the glucosinolate-myrosinase system, the so-called "mustard-oil bomb." Tissue damage caused by insect feeding brings glucosinolates into contact with the plant enzyme myrosinase, which hydrolyzes them to form toxic compounds, such as isothiocyanates. However, we previously observed that Phyllotreta striolata beetles themselves produce volatile glucosinolate hydrolysis products. Here, we show that P. striolata adults selectively accumulate glucosinolates from their food plants to up to 1.75% of their body weight and express their own myrosinase. By combining proteomics and transcriptomics, a gene responsible for myrosinase activity in P. striolata was identified. The major substrates of the heterologously expressed myrosinase were aliphatic glucosinolates, which were hydrolyzed with at least fourfold higher efficiency than aromatic and indolic glucosinolates, and β-O-glucosides. The identified beetle myrosinase belongs to the glycoside hydrolase family 1 and has up to 76% sequence similarity to other β-glucosidases. Phylogenetic analyses suggest species-specific diversification of this gene family in insects and an independent evolution of the beetle myrosinase from other insect β-glucosidases.

Health-promoting compounds of broccoli (Brassica oleracea L. var. italica) plants as affected by nitrogen fertilisation in projected future climatic change environments.

PubMed

Zaghdoud, Chokri; Carvajal, Micaela; Moreno, Diego A; Ferchichi, Ali; Del Carmen Martínez-Ballesta, María

2016-01-30

The complex interactions between CO2 increase and salinity were investigated in relation to decreased N supply, in order to determine the nutritional quality of broccoli (Brassica oleracea L. var. italica) plants under these conditions. Three different decreased N fertilisation regimes (NO3(-)/NH4(+) ratios of 100:0, 50:50 and 0:100 respectively) were combined with ambient (380 ppm) and elevated (800 ppm) [CO2 ] under non-saline (0 mmol L(-1) NaCl) and saline (80 mmol L(-1) NaCl) conditions. Nutrients (minerals, soluble protein and total amino acids) and natural antioxidants (glucosinolates, phenolic acids, flavonoids and vitamin C) were determined. In NH4(+) -fed broccoli plants, a marked growth reduction was shown and a redistribution of amino acids to cope with NH4(+) toxicity resulted in higher levels of indolic glucosinolate and total phenolic compounds. However, the positive effect of the higher [CO2] - ameliorating adverse effects of salinity--was only observed when N was supplied as NO3(-). Under reduced N fertilisation, the total glucosinolates were increased by a decreased NO3(-)/NH4 (+) ratio and elevated [CO2] but were unaffected by salinity. Under future climatic challenges, such as increased salinity and elevated [CO2], a clear genotypic dependence of S metabolism was observed in broccoli plants. In addition, an influence of the form in which N was supplied on plant nutritional quality was observed; a combined NO3(-)/NH4(+) (50:50) supply allowed broccoli plants not only to deal with NH4(+) toxicity but also to modify their glucosinolate content and profile. Thus, for different modes of N fertilisation, the interaction with climatic factors must be considered in the search for an optimal balance between yield and nutritional quality. © 2015 Society of Chemical Industry.

Exogenous Methyl Jasmonate and Salicylic Acid Induce Subspecies-Specific Patterns of Glucosinolate Accumulation and Gene Expression in Brassica oleracea L.

PubMed

Yi, Go-Eun; Robin, Arif Hasan Khan; Yang, Kiwoung; Park, Jong-In; Hwang, Byung Ho; Nou, Ill-Sup

2016-10-24

Glucosinolates have anti-carcinogenic properties. In the recent decades, the genetics of glucosinolate biosynthesis has been widely studied, however, the expression of specific genes involved in glucosinolate biosynthesis under exogenous phytohormone treatment has not been explored at the subspecies level in Brassica oleracea . Such data are vital for strategies aimed at selective exploitation of glucosinolate profiles. This study quantified the expression of 38 glucosinolate biosynthesis-related genes in three B. oleracea subspecies, namely cabbage, broccoli and kale, and catalogued associations between gene expression and increased contents of individual glucosinolates under methyl jasmonate (MeJA) and salicylic acid (SA) treatments. Glucosinolate accumulation and gene expression in response to phytohormone elicitation was subspecies specific. For instance, cabbage leaves showed enhanced accumulation of the aliphatic glucoiberin, progoitrin, sinigrin and indolic neoglucobrassicin under both MeJA and SA treatment. MeJA treatment induced strikingly higher accumulation of glucobrassicin (GBS) in cabbage and kale and of neoglucobrassicin (NGBS) in broccoli compared to controls. Notably higher expression of ST5a (Bol026200), CYP81F1 (Bol028913, Bol028914) and CYP81F4 genes was associated with significantly higher GBS accumulation under MeJA treatment compared to controls in all three subspecies. CYP81F4 genes, trans-activated by MYB34 genes, were expressed at remarkably high levels in all three subspecies under MeJA treatment, which also induced in higher indolic NGBS accumulation in all three subspecies. Remarkably higher expression of MYB28 (Bol036286), ST5b , ST5c , AOP2 , FMOGS-OX5 (Bol031350) and GSL-OH (Bol033373) was associated with much higher contents of aliphatic glucosinolates in kale leaves compared to the other two subspecies. The genes expressed highly could be utilized in strategies to selectively increase glucosinolate compounds in B. oleracea subspecies. These results promote efforts to develop genotypes of B. oleracea and other species with enhanced levels of desired glucosinolates.

Effects of intact glucosinolates and products produced from glucosinolates in myrosinase-catalyzed hydrolysis on the potato cyst nematode (Globodera rostochiensis Cv. Woll).

PubMed

Buskov, S; Serra, B; Rosa, E; Sørensen, H; Sørensen, J C

2002-02-13

The potato cyst nematode (Globodera rostochiensis cv. Woll) is responsible for large yield losses in the potato crop, and opportunities for reducing the attack of these plant nematode species are, therefore, important. This study has been devoted to the testing of the in vitro effects on the potato cyst nematode of eight glucosinolates [prop-2-enyl-, but-3-enyl-, (R)-4-methylsulfinylbut-3-enyl-, benzyl-, phenethyl-, 4-hydroxybenzyl-, (2S)-2-hydroxybut-3-enyl-, and (2R)-2-hydroxy-2-phenylethylglucosinolate] as well as the effects of the products of this myrosinase-catalyzed hydrolysis. The glucosinolates were used at three concentrations, 0.05, 0.3, and 1.0 mg/mL, in the presence or absence of the enzyme myrosinase. The effects of the compounds on the mortality were monitored every 8 h for a 72 h period. No effects were found for any of the intact glucosinolates. However, when active myrosinase was included with 1 mg/mL phenethylglucosinolate at pH 6.5, 100% mortality was observed within just 16 h. A similar effect was achieved at the same concentration of benzyl- and prop-2-enylglucosinolates in the myrosinase-containing solutions, although longer exposures were required (24 and 40 h, respectively). The main aglucone products released from the glucosinolates with pronounced effects on the nematodes were shown to be the corresponding isothiocyanates. The results suggest that mixtures of these specific glucosinolates and active myrosinase or autolysis of plant materials containing these enzymes and glucosinolates might be used to control the potato cyst nematode in the soil.

Metabolic Engineering of Valine- and Isoleucine-Derived Glucosinolates in Arabidopsis Expressing CYP79D2 from Cassava

PubMed Central

Mikkelsen, Michael Dalgaard; Halkier, Barbara Ann

2003-01-01

Glucosinolates are amino acid-derived natural products that, upon hydrolysis, typically release isothiocyanates with a wide range of biological activities. Glucosinolates play a role in plant defense as attractants and deterrents against herbivores and pathogens. A key step in glucosinolate biosynthesis is the conversion of amino acids to the corresponding aldoximes, which is catalyzed by cytochromes P450 belonging to the CYP79 family. Expression of CYP79D2 from cassava (Manihot esculenta Crantz.) in Arabidopsis resulted in the production of valine (Val)- and isoleucine-derived glucosinolates not normally found in this ecotype. The transgenic lines showed no morphological phenotype, and the level of endogenous glucosinolates was not affected. The novel glucosinolates were shown to constitute up to 35% of the total glucosinolate content in mature rosette leaves and up to 48% in old leaves. Furthermore, at increased concentrations of these glucosinolates, the proportion of Val-derived glucosinolates decreased. As the isothiocyanates produced from the Val- and isoleucine-derived glucosinolates are volatile, metabolically engineered plants producing these glucosinolates have acquired novel properties with great potential for improvement of resistance to herbivorous insects and for biofumigation. PMID:12586901

Behavior of glucosinolates in pickling cruciferous vegetables.

PubMed

Suzuki, Chise; Ohnishi-Kameyama, Mayumi; Sasaki, Keisuke; Murata, Takashi; Yoshida, Mitsuru

2006-12-13

Crucifer species, which include widely consumed vegetables, contain glucosinolates as secondary metabolites. Cruciferous vegetables are consumed in Japan in salt-preserved or pickled form as well as cooked and raw fresh vegetables. In this study, changes in contents of glucosinolates during the pickling process were investigated. 4-Methylthio-3-butenyl glucosinolate, a major glucosinolate in the root of Japanese radish, daikon (Raphanus sativus L.), was detected in pickled products with a short maturation period but not in those with a long maturation period. As a model pickling experiment, fresh watercress (Nasturtium officinale) and blanched watercress were soaked in 3% NaCl solution for 7 days. The results showed that the ratio of indole glucosinolates to total glucosinolates increased during the pickling process, whereas total glucosinolates decreased. Myrosinase digestion of glucosinolates in nozawana (Brassica rapa L.) indicated that indole glucosinolates, especially 4-methoxyglucobrassicin, were relatively resistant to the enzyme. The effect of pickling on glucosinolate content and the possible mechanism are discussed in view of degradation by myrosinase and synthetic reaction in response to salt stress or compression during the pickling process.

Isothiocyanates, Nitriles, and Epithionitriles from Glucosinolates Are Affected by Genotype and Developmental Stage in Brassica oleracea Varieties.

PubMed

Hanschen, Franziska S; Schreiner, Monika

2017-01-01

Vegetables of the Brassica oleracea group, such as broccoli, cauliflower, and cabbage, play an important role for glucosinolate consumption in the human diet. Upon maceration of the vegetable tissue, glucosinolates are degraded enzymatically to form volatile isothiocyanates, nitriles, and epithionitriles. However, only the uptake of isothiocyanates is linked to the cancer-preventive effects. Thus, it is of great interest to evaluate especially the isothiocyanate formation. Here, we studied the formation of glucosinolates and their respective hydrolysis products in sprouts and fully developed vegetable heads of different genotypes of the five B. oleracea varieties: broccoli, cauliflower as well as white, red, and savoy cabbages. Further, the effect of ontogeny (developmental stages) during the head development on the formation of glucosinolates and their respective hydrolysis products was evaluated at three different developmental stages (mini, fully developed, and over-mature head). Broccoli and red cabbage were mainly rich in 4-(methylsulfinyl)butyl glucosinolate (glucoraphanin), whereas cauliflower, savoy cabbage and white cabbage contained mainly 2-propenyl (sinigrin) and 3-(methylsulfinyl)propyl glucosinolate (glucoiberin). Upon hydrolysis, epithionitriles or nitriles were often observed to be the main hydrolysis products, with 1-cyano-2,3-epithiopropane being most abundant with up to 5.7 μmol/g fresh weight in white cabbage sprouts. Notably, sprouts often contained more than 10 times more glucosinolates or their hydrolysis products compared to fully developed vegetables. Moreover, during head development, both glucosinolate concentrations as well as hydrolysis product concentrations changed and mini heads contained the highest isothiocyanate concentrations. Thus, from a cancer-preventive point of view, consumption of mini heads of the B. oleracea varieties is recommended.

Isothiocyanates, Nitriles, and Epithionitriles from Glucosinolates Are Affected by Genotype and Developmental Stage in Brassica oleracea Varieties

PubMed Central

Hanschen, Franziska S.; Schreiner, Monika

2017-01-01

Vegetables of the Brassica oleracea group, such as broccoli, cauliflower, and cabbage, play an important role for glucosinolate consumption in the human diet. Upon maceration of the vegetable tissue, glucosinolates are degraded enzymatically to form volatile isothiocyanates, nitriles, and epithionitriles. However, only the uptake of isothiocyanates is linked to the cancer-preventive effects. Thus, it is of great interest to evaluate especially the isothiocyanate formation. Here, we studied the formation of glucosinolates and their respective hydrolysis products in sprouts and fully developed vegetable heads of different genotypes of the five B. oleracea varieties: broccoli, cauliflower as well as white, red, and savoy cabbages. Further, the effect of ontogeny (developmental stages) during the head development on the formation of glucosinolates and their respective hydrolysis products was evaluated at three different developmental stages (mini, fully developed, and over-mature head). Broccoli and red cabbage were mainly rich in 4-(methylsulfinyl)butyl glucosinolate (glucoraphanin), whereas cauliflower, savoy cabbage and white cabbage contained mainly 2-propenyl (sinigrin) and 3-(methylsulfinyl)propyl glucosinolate (glucoiberin). Upon hydrolysis, epithionitriles or nitriles were often observed to be the main hydrolysis products, with 1-cyano-2,3-epithiopropane being most abundant with up to 5.7 μmol/g fresh weight in white cabbage sprouts. Notably, sprouts often contained more than 10 times more glucosinolates or their hydrolysis products compared to fully developed vegetables. Moreover, during head development, both glucosinolate concentrations as well as hydrolysis product concentrations changed and mini heads contained the highest isothiocyanate concentrations. Thus, from a cancer-preventive point of view, consumption of mini heads of the B. oleracea varieties is recommended. PMID:28690627

Extraction and identification of isothiocyanates from broccolini seeds.

PubMed

Zhang, Bochao; Wang, Xiaoqin; Yang, Yanjing; Zhang, Xuewu

2011-01-01

Broccolini (Brassica oleracea Italica x Alboglabra) is a cross between broccoli and kai-lan (Chinese broccoli), which contains abundant glucosinolates. The intact glucosinolates are believed to be inactive, while their hydrolysis products, such as isothiocyanates (ITCs), are found to have bacteriocidal and anticarcinogenic activities. So far, no report is available about generation of ITCs during the process of glucosinolate hydrolysis in broccolini. In this study, the hydrolysis of broccolini seed glucosinolates was investigated under controlled conditions of pH, time and temperature, and the ITCs produced were determined. The results showed that an optimum hydrolysis of glucosinolates could be achieved at a temperature of 250C, at pH 7.0, and a reaction time of eight hours. Furthermore, GC-MS analysis indicated that the extracted ITCs primarily were: 3-BITC (3-benzyl-ITC) (10.8%), 4-methylpentyl-ITC (0.5%), 1-isothiocyanato-butane (26.8%), PEITC (phenethyl-ITC) (22.6%) and SFN (sulforaphane) (19.2%).

Transcriptome and Metabolome Analyses of Glucosinolates in Two Broccoli Cultivars Following Jasmonate Treatment for the Induction of Glucosinolate Defense to Trichoplusia ni (Hübner).

PubMed

Ku, Kang-Mo; Becker, Talon M; Juvik, John A

2016-07-15

Lepidopteran larvae growth is influenced by host plant glucosinolate (GS) concentrations, which are, in turn, influenced by the phytohormone jasmonate (JA). In order to elucidate insect resistance biomarkers to lepidopteran pests, transcriptome and metabolome analyses following JA treatments were conducted with two broccoli cultivars, Green Magic and VI-158, which have differentially induced indole GSs, neoglucobrassicin and glucobrassicin, respectively. To test these two inducible GSs on growth of cabbage looper (Trichoplusia ni), eight neonate cabbage looper larvae were placed onto each of three plants per JA treatments (0, 100, 200, 400 µM) three days after treatment. After five days of feeding, weight of larvae and their survival rate was found to decrease with increasing JA concentrations in both broccoli cultivars. JA-inducible GSs were measured by high performance liquid chromatography. Neoglucobrassicin in Green Magic and glucobrassicin in VI-158 leaves were increased in a dose-dependent manner. One or both of these glucosinolates and/or their hydrolysis products showed significant inverse correlations with larval weight and survival (five days after treatment) while being positively correlated with the number of days to pupation. This implies that these two JA-inducible glucosinolates can influence the growth and survival of cabbage looper larvae. Transcriptome profiling supported the observed changes in glucosinolate and their hydrolysis product concentrations following JA treatments. Several genes related to GS metabolism differentiate the two broccoli cultivars in their pattern of transcriptional response to JA treatments. Indicative of the corresponding change in indole GS concentrations, transcripts of the transcription factor MYB122, core structure biosynthesis genes (CYP79B2, UGT74B1, SUR1, SOT16, SOT17, and SOT18), an indole glucosinolate side chain modification gene (IGMT1), and several glucosinolate hydrolysis genes (TGG1, TGG2, and ESM1) were significantly increased in Green Magic (statistically significant in most cases at 400 µM) while UGT74B1 and MYB122 were significantly increased in VI-158. Therefore, these metabolite and transcript biomarker results indicate that transcriptome profiling can identify genes associated with the formation of two different indole GS and their hydrolysis products. Therefore, these metabolite and transcript biomarkers could be useful in an effective marker-assisted breeding strategy for resistance to generalist lepidopteran pests in broccoli and potentially other Brassica vegetables.

Transcriptome and Metabolome Analyses of Glucosinolates in Two Broccoli Cultivars Following Jasmonate Treatment for the Induction of Glucosinolate Defense to Trichoplusia ni (Hübner)

PubMed Central

Ku, Kang-Mo; Becker, Talon M.; Juvik, John A.

2016-01-01

Lepidopteran larvae growth is influenced by host plant glucosinolate (GS) concentrations, which are, in turn, influenced by the phytohormone jasmonate (JA). In order to elucidate insect resistance biomarkers to lepidopteran pests, transcriptome and metabolome analyses following JA treatments were conducted with two broccoli cultivars, Green Magic and VI-158, which have differentially induced indole GSs, neoglucobrassicin and glucobrassicin, respectively. To test these two inducible GSs on growth of cabbage looper (Trichoplusia ni), eight neonate cabbage looper larvae were placed onto each of three plants per JA treatments (0, 100, 200, 400 µM) three days after treatment. After five days of feeding, weight of larvae and their survival rate was found to decrease with increasing JA concentrations in both broccoli cultivars. JA-inducible GSs were measured by high performance liquid chromatography. Neoglucobrassicin in Green Magic and glucobrassicin in VI-158 leaves were increased in a dose-dependent manner. One or both of these glucosinolates and/or their hydrolysis products showed significant inverse correlations with larval weight and survival (five days after treatment) while being positively correlated with the number of days to pupation. This implies that these two JA-inducible glucosinolates can influence the growth and survival of cabbage looper larvae. Transcriptome profiling supported the observed changes in glucosinolate and their hydrolysis product concentrations following JA treatments. Several genes related to GS metabolism differentiate the two broccoli cultivars in their pattern of transcriptional response to JA treatments. Indicative of the corresponding change in indole GS concentrations, transcripts of the transcription factor MYB122, core structure biosynthesis genes (CYP79B2, UGT74B1, SUR1, SOT16, SOT17, and SOT18), an indole glucosinolate side chain modification gene (IGMT1), and several glucosinolate hydrolysis genes (TGG1, TGG2, and ESM1) were significantly increased in Green Magic (statistically significant in most cases at 400 µM) while UGT74B1 and MYB122 were significantly increased in VI-158. Therefore, these metabolite and transcript biomarker results indicate that transcriptome profiling can identify genes associated with the formation of two different indole GS and their hydrolysis products. Therefore, these metabolite and transcript biomarkers could be useful in an effective marker-assisted breeding strategy for resistance to generalist lepidopteran pests in broccoli and potentially other Brassica vegetables. PMID:27428958

How Glucosinolates Affect Generalist Lepidopteran Larvae: Growth, Development and Glucosinolate Metabolism

PubMed Central

Jeschke, Verena; Kearney, Emily E.; Schramm, Katharina; Kunert, Grit; Shekhov, Anton; Gershenzon, Jonathan; Vassão, Daniel G.

2017-01-01

Multiple lepidopteran larvae feed successfully on plants containing glucosinolates despite the diverse array of toxic and deterrent breakdown products, such as isothiocyanates (ITCs), formed upon plant damage. While much is known about how specialist lepidopterans metabolize and tolerate glucosinolates, there is little information about the metabolic fate of these plant defense compounds in specialized herbivores. Employing 13C- and 14C-labeled 4-methylsulfinylbutyl glucosinolate (glucoraphanin), we identified and quantified the major detoxification products of glucosinolates and ITCs in selected specialized and generalist larvae. While specialists prevented glucosinolate hydrolysis or diverted hydrolysis to form nitriles, hydrolysis in generalists proceeded to toxic ITCs, of which a portion were conjugated to glutathione. However, a large amount of ITCs remained unmodified, which may have led to the observed negative effects on growth and development. The performance of two generalist-feeding caterpillars, Spodoptera littoralis (African cotton leafworm) and Mamestra brassicae (cabbage moth) on Arabidopsis thaliana Col-0 and various glucosinolate-deficient mutants was investigated from hatching until pupation. We found that glucosinolates negatively affected larval growth and development, but not survival, with aliphatic glucosinolates having stronger effects than indolic glucosinolates, and the combination of the two glucosinolate types being even more detrimental to growth and development. Curiously, last instar larvae grew better on wild type than on non-glucosinolate-containing plant lines, but this could not be attributed to a change in detoxification rate or feeding behavior. Glucosinolates thus appear to be effective defenses against generalist lepidopteran herbivores at least during most stages of larval development. Nevertheless, the reversal of negative effects in the oldest instar is intriguing, and further investigation of this phenomenon may shed light on how generalists adjust their physiology to feed on diets with many different types of plant defense compounds. PMID:29209354

Two novel bioactive glucosinolates from Broccoli (Brassica oleracea L. var. italica) florets.

PubMed

Survay, Nazneen Shaik; Kumar, Brajesh; Jang, Mi; Yoon, Do-Young; Jung, Yi-Sook; Yang, Deok-Chun; Park, Se Won

2012-09-01

Two novel glucosinolates along with one known glucosinolate were isolated from Broccoli (Brassica oleracea L. var. italica) florets. Their structures were established mainly by 1D ((1)H and (13)C NMR), 2D NMR ((1)H-(1)H COSY, DEPT 135°, HSQC and HMBC), and Tandem MS-MS spectrometric data as 2-mercaptomethyl sulfinyl glucosinolate [(Z)-4-(methylsulfinyl)-N-(sulfooxy)-2-((2'S,3'R,4'S,5'S,6'R)-3',4',5'-trihydroxy-6'(hydroxylmethyl)-2'-mercapto tetrahydro-2H-pyran-2-yl) butane amide] 1, (Z)-1-((2S,5S)-5-hydroxytetra-hydro-2H-pyran-2-ylthio)-2-(1H-indol-3-yl) ethylidene amino sulfate 2 and a known cinnamoyl [6'-O-trans-(4″-hydroxy cinnamoyl)4-(methylsulphinyl)butyl glucosinolate] 3. Compound 1 exhibited scavenging activity against DPPH with an inhibitory concentration IC(50) of 20 mM, whereas compound 3 was a weak antioxidant when compared to the standard quercetin (5 mM) as a positive control. Both the compounds showed a significant and similar antimicrobial activity against Staphylococcus aureus with an IC(50) of <625 μg/mL when compared to antibiotic duricef. Against Salmonella typhimurium the IC(50) of 1 and 3 was determined as <625 μg/mL and <1250 μg/mL, respectively, when compared to ampicillin (IC(50) ≤ 39 μg/mL) as a positive control. Copyright © 2012 Elsevier Ltd. All rights reserved.

Engineering of benzylglucosinolate in tobacco provides proof-of-concept for dead-end trap crops genetically modified to attract Plutella xylostella (diamondback moth).

PubMed

Møldrup, Morten E; Geu-Flores, Fernando; de Vos, Martin; Olsen, Carl E; Sun, Joel; Jander, Georg; Halkier, Barbara A

2012-05-01

Glucosinolates are biologically active natural products characteristic of crucifers, including oilseed rape, cabbage vegetables and the model plant Arabidopsis thaliana. Crucifer-specialist insect herbivores, like the economically important pest Plutella xylostella (diamondback moth), frequently use glucosinolates as oviposition stimuli. This suggests that the transfer of a glucosinolate biosynthetic pathway to a non-crucifer would stimulate oviposition on an otherwise non-attractive plant. Here, we demonstrate that stable genetic transfer of the six-step benzylglucosinolate pathway from A. thaliana to Nicotiana tabacum (tobacco) results in the production of benzylglucosinolate without causing morphological alterations. Benzylglucosinolate-producing tobacco plants were more attractive for oviposition by female P. xylostella moths than wild-type tobacco plants. As newly hatched P. xylostella larvae were unable to survive on tobacco, these results represent a proof-of-concept strategy for rendering non-host plants attractive for oviposition by specialist herbivores with the long-term goal of generating efficient dead-end trap crops for agriculturally important pests. © 2012 The Authors. Plant Biotechnology Journal © 2012 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

Profiling of Glucosinolates and Flavonoids in Rorippa indica (Linn.) Hiern. (Cruciferae) by UHPLC-PDA-ESI/HRMSn

PubMed Central

2015-01-01

An UHPLC-PDA-ESI/HRMSn profiling method was used to identify the glucosinolates and flavonoids of Rorippa indica (Cruciferae), a wild vegetable and Chinese herb used to treat cough, diarrhea, and rheumatoid arthritis. Thirty-three glucosinolates, more than 40 flavonol glycosides, and 18 other phenolic and common organic compounds were identified. The glucosinolates and polyphenols were separated by UHPLC. High-resolution deprotonated molecules provided high accuracy mass values that were used to determine formulas and provide putative identification of the glucosinolates and flavonoids. The fragments from multistage mass spectrometry were used to elucidate the structures. The concentrations of the main components were based on UV peak areas and molar relative response factors with a single calibration standard. This study found this plant to be a rich source for glucosinolates, containing 24 new glucosinolates, including 14 glucosylated glucosinolates that were previously unidentified. PMID:24893216

Effect of microwave treatment on the efficacy of expeller pressing of Brassica napus rapeseed and Brassica juncea mustard seeds.

PubMed

Niu, Yanxing; Rogiewicz, Anna; Wan, Chuyun; Guo, Mian; Huang, Fenghong; Slominski, Bogdan A

2015-04-01

A study was conducted to evaluate the effect of microwave heating on the efficacy of expeller pressing of rapeseed and mustard seed and the composition of expeller meals in two types of Brassica napus rapeseed (intermediate- and low-glucosinolate) and in Brassica juncea mustard (high-glucosinolate). Following microwave treatment, the microstructure of rapeseed using transmission electron microscopy showed a significant disappearance of oil bodies and myrosin cells. After 6 min of microwave heating (400 g, 800 W), the oil content of rapeseed expeller meal decreased from 44.9 to 13.5% for intermediate-glucosinolate B. napus rapeseed, from 42.6 to 11.3% for low-glucosinolate B. napus rapeseed, and from 44.4 to 14.1% for B. juncea mustard. The latter values were much lower than the oil contents of the corresponding expeller meals derived from the unheated seeds (i.e., 26.6, 22.6, and 29.8%, respectively). Neutral detergent fiber (NDF) contents showed no differences except for the expeller meal from the intermediate-glucosinolate B. napus rapeseed, which increased from 22.7 to 29.2% after 6 min of microwave heating. Microwave treatment for 4 and 5 min effectively inactivated myrosinase enzyme of intermediate-glucosinolate B. napus rapeseed and B. juncea mustard seed, respectively. In low-glucosinolate B. napus rapeseed the enzyme appeared to be more heat stable, with some activity being present after 6 min of microwave heating. Myrosinase enzyme inactivation had a profound effect on the glucosinolate content of expeller meals and prevented their hydrolysis to toxic breakdown products during the expelling process. It appeared evident from this study that microwave heating for 6 min was an effective method of producing expeller meal without toxic glucosinolate breakdown products while at the same time facilitating high yield of oil during the expelling process.

Effect of storage, processing and cooking on glucosinolate content of Brassica vegetables.

PubMed

Song, Lijiang; Thornalley, Paul J

2007-02-01

Epidemiological studies have shown that consumption of Brassica vegetables decrease the risk of cancer. These associations are linked to dietary intake of glucosinolates and their metabolism to cancer preventive isothiocyanates. Bioavailability of glucosinolates and related isothiocyanates are influenced by storage and culinary processing of Brassica vegetables. In this work, the content of the 7 major glucosinolates in broccoli, Brussels sprouts, cauliflower and green cabbage and their stability under different storage and cooking conditions is examined. Glucosinolates and isothiocyanates were quantified by liquid chromatography with tandem mass spectrometric detection (LC-MS/MS). Isothiocyanates were detected with high sensitivity as the corresponding thiourea derivatives. Storage at ambient temperature and in a domestic refrigerator showed no significant difference and a minor loss (9-26%) of glucosinolate levels over 7 days. Vegetables shredded finely showed a marked decline of glucosinolate level with post-shredding dwell time - up to 75% over 6h. Glucosinolate losses were detected partly as isothiocyanates. Cooking by steaming, microwaving and stir-fry did not produce significant loss of glucosinolates whereas boiling showed significant losses by leaching into cooking water. Most of the loss of the glucosinolates (approximately 90%) was detected in the cooking water. Increased bioavailability of dietary isothiocyanates may be achieved by avoiding boiling of vegetables.

Cloning of genes related to aliphatic glucosinolate metabolism and the mechanism of sulforaphane accumulation in broccoli sprouts under jasmonic acid treatment.

PubMed

Guo, Liping; Yang, Runqiang; Gu, Zhenxin

2016-10-01

Cytochrome P450 79F1 (CYP79F1), cytochrome P450 83A1 (CYP83A1), UDP-glucosyltransferase 74B1 (UGT74B1), sulfotransferase 18 (ST5b) and flavin-containing monooxygenase GS-OX1 (FMOGS - OX1 ) are important enzymes in aliphatic glucosinolate biosynthesis. In this study, their full-length cDNA in broccoli was firstly cloned, then the mechanism of sulforaphane accumulation under jasmonic acid (JA) treatment was investigated. The full-length cDNA of CYP79F1, CYP83A1, UGT74B1, ST5b and FMOGS - OX1 comprised 1980, 1652, 1592, 1378 and 1623 bp respectively. The increase in aliphatic glucosinolate accumulation in broccoli sprouts treated with JA was associated with elevated expression of genes in the aliphatic glucosinolate biosynthetic pathway. Application of 100 µmol L(-1) JA increased myrosinase (MYR) activity but did not affect epithiospecifier protein (ESP) activity in broccoli sprouts, which was supported by the expression of MYR and ESP. Sulforaphane formation in 7-day-old sprouts treated with 100 µmol L(-1) JA was 3.36 and 1.30 times that in the control and 300 µmol L(-1) JA treatment respectively. JA enhanced the accumulation of aliphatic glucosinolates in broccoli sprouts via up-regulation of related gene expression. Broccoli sprouts treated with 100 µmol L(-1) JA showed higher sulforphane formation than those treated with 300 µmol L(-1) JA owing to the higher glucoraphanin content and myrosinase activity under 100 µmol L(-1) JA treatment. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Identification and expression analysis of glucosinolate biosynthetic genes and estimation of glucosinolate contents in edible organs of Brassica oleracea subspecies.

PubMed

Yi, Go-Eun; Robin, Arif Hasan Khan; Yang, Kiwoung; Park, Jong-In; Kang, Jong-Goo; Yang, Tae-Jin; Nou, Ill-Sup

2015-07-20

Glucosinolates are anti-carcinogenic, anti-oxidative biochemical compounds that defend plants from insect and microbial attack. Glucosinolates are abundant in all cruciferous crops, including all vegetable and oilseed Brassica species. Here, we studied the expression of glucosinolate biosynthesis genes and determined glucosinolate contents in the edible organs of a total of 12 genotypes of Brassica oleracea: three genotypes each from cabbage, kale, kohlrabi and cauliflower subspecies. Among the 81 genes analyzed by RT-PCR, 19 are transcription factor-related, two different sets of 25 genes are involved in aliphatic and indolic biosynthesis pathways and the rest are breakdown-related. The expression of glucosinolate-related genes in the stems of kohlrabi was remarkably different compared to leaves of cabbage and kale and florets of cauliflower as only eight genes out of 81 were expressed in the stem tissues of kohlrabi. In the stem tissue of kohlrabi, only one aliphatic transcription factor-related gene, Bol036286 (MYB28) and one indolic transcription factor-related gene, Bol030761 (MYB51), were expressed. The results indicated the expression of all genes is not essential for glucosinolate biosynthesis. Using HPLC analysis, a total of 16 different types of glucosinolates were identified in four subspecies, nine of them were aliphatic, four of them were indolic and one was aromatic. Cauliflower florets measured the highest number of 14 glucosinolates. Among the aliphatic glucosinolates, only gluconapin was found in the florets of cauliflower. Glucoiberverin and glucobrassicanapin contents were the highest in the stems of kohlrabi. The indolic methoxyglucobrassicin and aromatic gluconasturtiin accounted for the highest content in the florets of cauliflower. A further detailed investigation and analyses is required to discern the precise roles of each of the genes for aliphatic and indolic glucosinolate biosynthesis in the edible organs.

Optimization of pulsed electric field pre-treatments to enhance health-promoting glucosinolates in broccoli flowers and stalk.

PubMed

Aguiló-Aguayo, Ingrid; Suarez, Manuel; Plaza, Lucia; Hossain, Mohammad B; Brunton, Nigel; Lyng, James G; Rai, Dilip K

2015-07-01

The effect of pulsed electric field (PEF) treatment variables (electric field strength and treatment time) on the glucosinolate content of broccoli flowers and stalks was evaluated. Samples were subjected to electric field strengths from 1 to 4 kV cm(-1) and treatment times from 50 to 1000 µs at 5 Hz. Data fitted significantly (P < 0.0014) the proposed second-order response functions. The results showed that PEF combined treatment conditions of 4 kV cm(-1) for 525 and 1000 µs were optimal to maximize glucosinolate levels in broccoli flowers (ranging from 187.1 to 212.5%) and stalks (ranging from 110.6 to 203.0%) respectively. The predicted values from the developed quadratic polynomial equation were in close agreement with the actual experimental values, with low average mean deviations (E%) ranging from 0.59 to 8.80%. The use of PEF processing at moderate conditions could be a suitable method to stimulate production of broccoli with high health-promoting glucosinolate content. © 2014 Society of Chemical Industry.

Impact of selenium supply on Se-methylselenocysteine and glucosinolate accumulation in selenium-biofortified Brassica sprouts.

PubMed

Avila, Fabricio William; Yang, Yong; Faquin, Valdemar; Ramos, Silvio Junio; Guilherme, Luiz Roberto G; Thannhauser, Theodore W; Li, Li

2014-12-15

Brassica sprouts are widely marketed as functional foods. Here we examined the effects of Se treatment on the accumulation of anticancer compound Se-methylselenocysteine (SeMSCys) and glucosinolates in Brassica sprouts. Cultivars from the six most extensively consumed Brassica vegetables (broccoli, cauliflower, green cabbage, Chinese cabbage, kale, and Brussels sprouts) were used. We found that Se-biofortified Brassica sprouts all were able to synthesize significant amounts of SeMSCys. Analysis of glucosinolate profiles revealed that each Brassica crop accumulated different types and amounts of glucosinolates. Cauliflower sprouts had high total glucosinolate content. Broccoli sprouts contained high levels of glucoraphanin, a precursor for potent anticancer compound. Although studies have reported an inverse relationship between accumulation of Se and glucosinolates in mature Brassica plants, Se supply generally did not affect glucosinolate accumulation in Brassica sprouts. Thus, Brassica vegetable sprouts can be biofortified with Se for the accumulation of SeMSCys without negative effects on chemopreventive glucosinolate contents. Published by Elsevier Ltd.

Glucosinolate metabolism, functionality and breeding for the improvement of Brassicaceae vegetables

PubMed Central

Ishida, Masahiko; Hara, Masakazu; Fukino, Nobuko; Kakizaki, Tomohiro; Morimitsu, Yasujiro

2014-01-01

Unique secondary metabolites, glucosinolates (S-glucopyranosyl thiohydroximates), are naturally occurring S-linked glucosides found mainly in Brassicaceae plants. They are enzymatically hydrolyzed to produce sulfate ions, D-glucose, and characteristic degradation products such as isothiocyanates. The functions of glucosinolates in the plants remain unclear, but isothiocyanates possessing a pungent or irritating taste and odor might be associated with plant defense from microbes. Isothiocyanates have been studied extensively in experimental in vitro and in vivo carcinogenesis models for their cancer chemopreventive properties. The beneficial isothiocyanates, glucosinolates that are functional for supporting human health, have received attention from many scientists studying plant breeding, plant physiology, plant genetics, and food functionality. This review presents a summary of recent topics related with glucosinolates in the Brassica family, along with a summary of the chemicals, metabolism, and genes of glucosinolates in Brassicaceae. The bioavailabilities of isothiocyanates from certain functional glucosinolates and the importance of breeding will be described with emphasis on glucosinolates. PMID:24987290

Brassicaceae tissues as inhibitors of nitrification in soil.

PubMed

Brown, Paul D; Morra, Matthew J

2009-09-09

Brassicaceae crops often produce an unexplained increase in plant-available soil N possibly related to bioactive compounds produced from glucosinolates present in the tissues. Our objective was to determine if glucosinolate-containing tissues inhibit nitrification, thereby potentially explaining this observation. Ammonium, NO(2)(-), and NO(3)(-) N were measured in soils amended with Brassicaceae ( Isatis tinctoria L., Brassica napus L., Brassica juncea L., and Sinapis alba L.) tissues containing different glucosinolate types and concentrations or Kentucky bluegrass ( Poa pratensis L.) residues with equivalent C/N ratios as the Brassicaceae samples. There was greater accumulation of NH(4)(+) N in soils amended with tissues containing high glucosinolate concentrations as compared to soils amended with tissues containing no or low glucosinolate concentrations. Nitrite N was detected only in soils amended with Brassicaceae tissues having the highest glucosinolate concentrations. The positive correlation of both NH(4)(+) and NO(2)(-) N accumulation with the glucosinolate concentration indicates the participation of glucosinolate hydrolysis products in nitrification inhibition.

Chemopreventive glucosinolate accumulation in various broccoli and collard tissues: Microfluidic-based targeted transcriptomics for by-product valorization

PubMed Central

Becker, Talon M.; Juvik, John A.

2017-01-01

Floret, leaf, and root tissues were harvested from broccoli and collard cultivars and extracted to determine their glucosinolate and hydrolysis product profiles using high performance liquid chromatography and gas chromotography. Quinone reductase inducing bioactivity, an estimate of anti-cancer chemopreventive potential, of the extracts was measured using a hepa1c1c7 murine cell line. Extracts from root tissues were significantly different from other tissues and contained high levels of gluconasturtiin and glucoerucin. Targeted gene expression analysis on glucosinolate biosynthesis revealed that broccoli root tissue has elevated gene expression of AOP2 and low expression of FMOGS-OX homologs, essentially the opposite of what was observed in broccoli florets, which accumulated high levels of glucoraphanin. Broccoli floret tissue has significantly higher nitrile formation (%) and epithionitrile specifier protein gene expression than other tissues. This study provides basic information of the glucosinolate metabolome and transcriptome for various tissues of Brassica oleracea that maybe utilized as potential byproducts for the nutraceutical market. PMID:28945821

Chemopreventive glucosinolate accumulation in various broccoli and collard tissues: Microfluidic-based targeted transcriptomics for by-product valorization.

PubMed

Lee, Young-Sang; Ku, Kang-Mo; Becker, Talon M; Juvik, John A

2017-01-01

Floret, leaf, and root tissues were harvested from broccoli and collard cultivars and extracted to determine their glucosinolate and hydrolysis product profiles using high performance liquid chromatography and gas chromotography. Quinone reductase inducing bioactivity, an estimate of anti-cancer chemopreventive potential, of the extracts was measured using a hepa1c1c7 murine cell line. Extracts from root tissues were significantly different from other tissues and contained high levels of gluconasturtiin and glucoerucin. Targeted gene expression analysis on glucosinolate biosynthesis revealed that broccoli root tissue has elevated gene expression of AOP2 and low expression of FMOGS-OX homologs, essentially the opposite of what was observed in broccoli florets, which accumulated high levels of glucoraphanin. Broccoli floret tissue has significantly higher nitrile formation (%) and epithionitrile specifier protein gene expression than other tissues. This study provides basic information of the glucosinolate metabolome and transcriptome for various tissues of Brassica oleracea that maybe utilized as potential byproducts for the nutraceutical market.

Secondary Metabolism in Brassica Rapa Under Hypergravity

NASA Astrophysics Data System (ADS)

Levine, Lanfang; Darnell, Rebecca; Allen, Joan; Musgrave, Mary; Bisbee, Patricia

Effect of altered gravity on secondary metabolism is of critical importance not only from the viewpoint of plant evolution, but also of productivity (carbon partition between edible and non-edible parts), plant fitness, as well as culinary and nutraceutical values to human diet. Previous work found that lignin content decreases in microgravity as the need for mechanical support decreases, while the response of other small molecular secondary metabolites to microgravity varies. Our recent ISS experiment showed that 3-butenyl glucosinolate (a predominant glucosinolate in Brassica rapa) increased in stems of B. rapa grown in the microgravity conditions. To further elucidate the role of gravity in plant secondary metabolism, a series of hypergravity (the other end of gravity spectrum) experiments were carried out using the 24-ft centrifuge at Ames Research Center. Thirteen-day-old B. rapa L. (cv. Astroplants) were transferred to the Plant Growth Facility attached to the centrifuge following previous experimental conditions, and subsequently grown for 16 days. Plants were harvested, immediately frozen in liquid nitrogen, and lyophilized prior to analysis for glucosinolates and lignin. In general, glucosinolate concentration was the highest in stems, followed by leaves, then roots. Glucosinolate concentration was significantly lower in stems of the 2-g and 4-g plants - averaging 4.6 and 2.5 ng/g DW, respectively - compared with the stationary control plants, which averaged 7.9 ng/g DW. Similarly, there was a 2.2-fold and 7.5-fold decrease in 3-butenyl glucosinolate in roots of the 2-g and 4-g plants, respectively, compared with the control (2.6 ng/g DW). There was a significant decrease in 3-butenyl glucosinolate concentration in leaves of the 4-g compared to leaves of the control plants (2.6 and 4.5 ng/g DW, respectively); however, there was no effect of 2-g on leaf glucosinolate concentration. Increasing gravity from 1-g to 2-g to 4-g generally resulted in further decreases in glucosinolate accumulation. Combining these results with the findings from ISS confirmed a negative correlation between glucosinolate and gravity. Lignin in the same plant materials is currently under investigation. A positive relationship between lignin and gravity is expected. This project was supported by NASA grant NAG10-329 and NNX07AT77G.

Rocket salad (Diplotaxis and Eruca spp.) sensory analysis and relation with glucosinolate and phenolic content.

PubMed

Pasini, Federica; Verardo, Vito; Cerretani, Lorenzo; Caboni, Maria Fiorenza; D'Antuono, Luigi Filippo

2011-12-01

Salad crops of the Brassicaceae family, such as Diplotaxis tenuifolia and Eruca vesicaria, commonly referred to as 'rocket salads', have attracted considerable interest as culinary vegetables because of their strong flavour and their content of putative health-promoting compounds. Among such compounds, glucosinolates and phenolics are well-known phytochemicals with an important role also in determining the characteristic flavour of these species. In this study, to identify potentially high-value rocket salads, 37 cultivated types were examined for sensory characters and their relations with glucosinolate and phenolic contents, which ranged from 0.76 to 3.03 g kg(-1) dry weight (DW) and from 4.68 to 31.39 g kg(-1) DW, respectively. The perception of bitter taste was significantly affected by specific glucosinolates, namely progoitrin/epiprogoitrin and dimeric glucosativin. Aroma intensity was negatively related to glucoalyssin content, whereas pungency was significantly related to total glucosinolate content. Kaempferol-3-(2-sinapoyl-glucoside)-4'-glucoside was positively and significantly related to all flavour trait perceptions. Aroma intensity, pungency, crunchiness and juiciness were positively related to typical rocket salad flavour perception through a prominent direct effect. Aroma intensity, pungency, crunchiness and juiciness were strong determinants of overall rocket salad flavour perception. Visual traits also characterised sensory components. Bitterness, usually considered a negative flavour trait, was moderately perceived in the examined material, without negatively affecting typical flavour perception. In the range of the examined material, glucosinolate content did not contrast with typical flavour, demonstrating that good taste and putative health-promoting properties may coexist. Copyright © 2011 Society of Chemical Industry.

Reduction of antinutritional glucosinolates in Brassica oilseeds by mutation of genes encoding transporters.

PubMed

Nour-Eldin, Hussam Hassan; Madsen, Svend Roesen; Engelen, Steven; Jørgensen, Morten Egevang; Olsen, Carl Erik; Andersen, Jonathan Sonne; Seynnaeve, David; Verhoye, Thalia; Fulawka, Rudy; Denolf, Peter; Halkier, Barbara Ann

2017-04-01

The nutritional value of Brassica seed meals is reduced by the presence of glucosinolates, which are toxic compounds involved in plant defense. Mutation of the genes encoding two glucosinolate transporters (GTRs) eliminated glucosinolates from Arabidopsis thaliana seeds, but translation of loss-of-function phenotypes into Brassica crops is challenging because Brassica is polyploid. We mutated one of seven and four of 12 GTR orthologs and reduced glucosinolate levels in seeds by 60-70% in two different Brassica species (Brassica rapa and Brassica juncea). Reduction in seed glucosinolates was stably inherited over multiple generations and maintained in field trials of two mutant populations at three locations. Successful translation of the gtr loss-of-function phenotype from model plant to two Brassica crops suggests that our transport engineering approach could be broadly applied to reduce seed glucosinolate content in other oilseed crops, such as Camelina sativa or Crambe abyssinica.

UVA, UVB Light Doses and Harvesting Time Differentially Tailor Glucosinolate and Phenolic Profiles in Broccoli Sprouts.

PubMed

Moreira-Rodríguez, Melissa; Nair, Vimal; Benavides, Jorge; Cisneros-Zevallos, Luis; Jacobo-Velázquez, Daniel A

2017-06-26

Broccoli sprouts contain health-promoting glucosinolate and phenolic compounds that can be enhanced by applying ultraviolet light (UV). Here, the effect of UVA or UVB radiation on glucosinolate and phenolic profiles was assessed in broccoli sprouts. Sprouts were exposed for 120 min to low intensity and high intensity UVA (UVA L , UVA H ) or UVB (UVB L , UVB H ) with UV intensity values of 3.16, 4.05, 2.28 and 3.34 W/m², respectively. Harvest occurred 2 or 24 h post-treatment; and methanol/water or ethanol/water (70%, v / v ) extracts were prepared. Seven glucosinolates and 22 phenolics were identified. Ethanol extracts showed higher levels of certain glucosinolates such as glucoraphanin, whereas methanol extracts showed slight higher levels of phenolics. The highest glucosinolate accumulation occurred 24 h after UVB H treatment, increasing 4-methoxy-glucobrassicin, glucobrassicin and glucoraphanin by ~170, 78 and 73%, respectively. Furthermore, UVA L radiation and harvest 2 h afterwards accumulated gallic acid hexoside I (~14%), 4- O -caffeoylquinic acid (~42%), gallic acid derivative (~48%) and 1-sinapoyl-2,2-diferulolyl-gentiobiose (~61%). Increases in sinapoyl malate (~12%), gallotannic acid (~48%) and 5-sinapoyl-quinic acid (~121%) were observed with UVB H Results indicate that UV-irradiated broccoli sprouts could be exploited as a functional food for fresh consumption or as a source of bioactive phytochemicals with potential industrial applications.

Evaluating the impact of sprouting conditions on the glucosinolate content of Brassica oleracea sprouts.

PubMed

Vale, A P; Santos, J; Brito, N V; Fernandes, D; Rosa, E; Oliveira, M Beatriz P P

2015-07-01

The glucosinolates content of brassica plants is a distinctive characteristic, representing a healthy advantage as many of these compounds are associated to antioxidant and anti-carcinogenic properties. Brassica sprouts are still an underutilized source of these bioactive compounds. In this work, four varieties of brassica sprouts (red cabbage, broccoli, Galega kale and Penca cabbage), including two local varieties from the North of Portugal, were grown to evaluate the glucosinolate profile and myrosinase activity during the sprouting. Also the influence of light/darkness exposure during sprouting on the glucosinolate content was assessed. Glucosinolate content and myrosinase activity of the sprouts was evaluated by HPLC methods. All sprouts revealed a higher content of aliphatic glucosinolates than of indole glucosinolates, contrary to the profile described for most of brassica mature plants. Galega kale sprouts had the highest glucosinolate content, mainly sinigrin and glucoiberin, which are recognized for their beneficial health effects. Penca cabbage sprouts were particularly richer in glucoraphanin, who was also one of the major compounds in broccoli sprouts. Red cabbage showed a higher content of progoitrin. Regarding myrosinase activity, Galega kale sprouts showed the highest values, revealing that the use of light/dark cycles and a sprouting phase of 7-9 days could be beneficial to preserve the glucosinolate content of this variety. Copyright © 2015 Elsevier Ltd. All rights reserved.

Interaction of cruciferous phytoanticipins with plant fungal pathogens: indole glucosinolates are not metabolized but the corresponding desulfo-derivatives and nitriles are.

PubMed

Pedras, M Soledade C; Hossain, Sajjad

2011-12-01

Glucosinolates represent a large group of plant natural products long known for diverse and fascinating physiological functions and activities. Despite the relevance and huge interest on the roles of indole glucosinolates in plant defense, little is known about their direct interaction with microbial plant pathogens. Toward this end, the metabolism of indolyl glucosinolates, their corresponding desulfo-derivatives, and derived metabolites, by three fungal species pathogenic on crucifers was investigated. While glucobrassicin, 1-methoxyglucobrassicin, 4-methoxyglucobrassicin were not metabolized by the pathogenic fungi Alternaria brassicicola, Rhizoctonia solani and Sclerotinia sclerotiorum, the corresponding desulfo-derivatives were metabolized to indolyl-3-acetonitrile, caulilexin C (1-methoxyindolyl-3-acetonitrile) and arvelexin (4-methoxyindolyl-3-acetonitrile) by R. solani and S. sclerotiorum, but not by A. brassicicola. That is, desulfo-glucosinolates were metabolized by two non-host-selective pathogens, but not by a host-selective. Indolyl-3-acetonitrile, caulilexin C and arvelexin were metabolized to the corresponding indole-3-carboxylic acids. Indolyl-3-acetonitriles displayed higher inhibitory activity than indole desulfo-glucosinolates. Indolyl-3-methanol displayed antifungal activity and was metabolized by A. brassicicola and R. solani to the less antifungal compounds indole-3-carboxaldehyde and indole-3-carboxylic acid. Diindolyl-3-methane was strongly antifungal and stable in fungal cultures, but ascorbigen was not stable in solution and displayed low antifungal activity; neither compound appeared to be metabolized by any of the three fungal species. The cell-free extracts of mycelia of A. brassicicola displayed low myrosinase activity using glucobrassicin as substrate, but myrosinase activity was not detectable in mycelia of either R. solani or S. sclerotiorum. Copyright © 2011 Elsevier Ltd. All rights reserved.

Degradation of Biofumigant Isothiocyanates and Allyl Glucosinolate in Soil and Their Effects on the Microbial Community Composition

PubMed Central

Hanschen, Franziska S.; Yim, Bunlong; Winkelmann, Traud; Smalla, Kornelia; Schreiner, Monika

2015-01-01

Brassicales species rich in glucosinolates are used for biofumigation, a process based on releasing enzymatically toxic isothiocyanates into the soil. These hydrolysis products are volatile and often reactive compounds. Moreover, glucosinolates can be degraded also without the presence of the hydrolytic enzyme myrosinase which might contribute to bioactive effects. Thus, in the present study the stability of Brassicaceae plant-derived and pure glucosinolates hydrolysis products was studied using three different soils (model biofumigation). In addition, the degradation of pure 2-propenyl glucosinolate was investigated with special regard to the formation of volatile breakdown products. Finally, the influence of pure glucosinolate degradation on the bacterial community composition was evaluated using denaturing gradient gel electrophoresis of 16S rRNA gene amplified from total community DNA. The model biofumigation study revealed that the structure of the hydrolysis products had a significant impact on their stability in the soil but not the soil type. Following the degradation of pure 2-propenyl glucosinolate in the soils, the nitrile as well as the isothiocyanate can be the main degradation products, depending on the soil type. Furthermore, the degradation was shown to be both chemically as well as biologically mediated as autoclaving reduced degradation. The nitrile was the major product of the chemical degradation and its formation increased with iron content of the soil. Additionally, the bacterial community composition was significantly affected by adding pure 2-propenyl glucosinolate, the effect being more pronounced than in treatments with myrosinase added to the glucosinolate. Therefore, glucosinolates can have a greater effect on soil bacterial community composition than their hydrolysis products. PMID:26186695

Purification of glucosinolates from Camelina sativa seeds

USDA-ARS?s Scientific Manuscript database

Camelina sativa L. Crantz defatted seed press cake contains a number of phytochemicals, including the flavonoid rutin (quercetin 3-O-rutinoside), an acylated quercetin glycoside, and three glucosinolates: glucoarabin (9-(methylsulfinyl)nonyl-glucosinolate) glucocamelinin (10-(methylsulfinyl)decyl-gl...

Diversity of Kale (Brassica oleracea var. sabellica): Glucosinolate Content and Phylogenetic Relationships.

PubMed

Hahn, Christoph; Müller, Anja; Kuhnert, Nikolai; Albach, Dirk

2016-04-27

Recently, kale has become popular due to nutritive components beneficial for human health. It is an important source of phytochemicals such as glucosinolates that trigger associated cancer-preventive activity. However, nutritional value varies among glucosinolates and among cultivars. Here, we start a systematic determination of the content of five glucosinolates in 25 kale varieties and 11 non-kale Brassica oleracea cultivars by HPLC-DAD-ESI-MS(n) and compare the profiles with results from the analysis of SNPs derived from a KASP genotyping assay. Our results demonstrate that the glucosinolate levels differ markedly among varieties of different origin. Comparison of the phytochemical data with phylogenetic relationships revealed that the common name kale refers to at least three different groups. German, American, and Italian kales differ morphologically and phytochemically. Landraces do not show outstanding glucosinolate levels. Our results demonstrate the diversity of kale and the importance of preserving a broad genepool for future breeding purposes.

Bioavailability of Glucosinolates and Their Breakdown Products: Impact of Processing

PubMed Central

Barba, Francisco J.; Nikmaram, Nooshin; Roohinejad, Shahin; Khelfa, Anissa; Zhu, Zhenzhou; Koubaa, Mohamed

2016-01-01

Glucosinolates are a large group of plant secondary metabolites with nutritional effects, and are mainly found in cruciferous plants. After ingestion, glucosinolates could be partially absorbed in their intact form through the gastrointestinal mucosa. However, the largest fraction is metabolized in the gut lumen. When cruciferous are consumed without processing, myrosinase enzyme present in these plants hydrolyzes the glucosinolates in the proximal part of the gastrointestinal tract to various metabolites, such as isothiocyanates, nitriles, oxazolidine-2-thiones, and indole-3-carbinols. When cruciferous are cooked before consumption, myrosinase is inactivated and glucosinolates transit to the colon where they are hydrolyzed by the intestinal microbiota. Numerous factors, such as storage time, temperature, and atmosphere packaging, along with inactivation processes of myrosinase are influencing the bioavailability of glucosinolates and their breakdown products. This review paper summarizes the assimilation, absorption, and elimination of these molecules, as well as the impact of processing on their bioavailability. PMID:27579302

Insect Attraction versus Plant Defense: Young Leaves High in Glucosinolates Stimulate Oviposition by a Specialist Herbivore despite Poor Larval Survival due to High Saponin Content

PubMed Central

Badenes-Perez, Francisco R.; Gershenzon, Jonathan; Heckel, David G.

2014-01-01

Glucosinolates are plant secondary metabolites used in plant defense. For insects specialized on Brassicaceae, such as the diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), glucosinolates act as “fingerprints” that are essential in host plant recognition. Some plants in the genus Barbarea (Brassicaceae) contain, besides glucosinolates, saponins that act as feeding deterrents for P. xylostella larvae, preventing their survival on the plant. Two-choice oviposition tests were conducted to study the preference of P. xylostella among Barbarea leaves of different size within the same plant. P. xylostella laid more eggs per leaf area on younger leaves compared to older ones. Higher concentrations of glucosinolates and saponins were found in younger leaves than in older ones. In 4-week-old plants, saponins were present in true leaves, while cotyledons contained little or no saponins. When analyzing the whole foliage of the plant, the content of glucosinolates and saponins also varied significantly in comparisons among plants that were 4, 8, and 12 weeks old. In Barbarea plants and leaves of different ages, there was a positive correlation between glucosinolate and saponin levels. This research shows that, in Barbarea plants, ontogenetical changes in glucosinolate and saponin content affect both attraction and resistance to P. xylostella. Co-occurrence of a high content of glucosinolates and saponins in the Barbarea leaves that are most valuable for the plant, but are also the most attractive to P. xylostella, provides protection against this specialist herbivore, which oviposition behavior on Barbarea seems to be an evolutionary mistake. PMID:24752069

Insect attraction versus plant defense: young leaves high in glucosinolates stimulate oviposition by a specialist herbivore despite poor larval survival due to high saponin content.

PubMed

Badenes-Perez, Francisco R; Gershenzon, Jonathan; Heckel, David G

2014-01-01

Glucosinolates are plant secondary metabolites used in plant defense. For insects specialized on Brassicaceae, such as the diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), glucosinolates act as "fingerprints" that are essential in host plant recognition. Some plants in the genus Barbarea (Brassicaceae) contain, besides glucosinolates, saponins that act as feeding deterrents for P. xylostella larvae, preventing their survival on the plant. Two-choice oviposition tests were conducted to study the preference of P. xylostella among Barbarea leaves of different size within the same plant. P. xylostella laid more eggs per leaf area on younger leaves compared to older ones. Higher concentrations of glucosinolates and saponins were found in younger leaves than in older ones. In 4-week-old plants, saponins were present in true leaves, while cotyledons contained little or no saponins. When analyzing the whole foliage of the plant, the content of glucosinolates and saponins also varied significantly in comparisons among plants that were 4, 8, and 12 weeks old. In Barbarea plants and leaves of different ages, there was a positive correlation between glucosinolate and saponin levels. This research shows that, in Barbarea plants, ontogenetical changes in glucosinolate and saponin content affect both attraction and resistance to P. xylostella. Co-occurrence of a high content of glucosinolates and saponins in the Barbarea leaves that are most valuable for the plant, but are also the most attractive to P. xylostella, provides protection against this specialist herbivore, which oviposition behavior on Barbarea seems to be an evolutionary mistake.

Biofumigation for control of pale potato cyst nematodes: activity of brassica leaf extracts and green manures on Globodera pallida in vitro and in soil.

PubMed

Lord, James S; Lazzeri, Luca; Atkinson, Howard J; Urwin, Peter E

2011-07-27

The effects of brassica green manures on Globodera pallida were assessed in vitro and in soil microcosms. Twelve of 22 brassica accessions significantly inhibited the motility of G. pallida infective juveniles in vitro. Green manures of selected brassicas were then incorporated into soil containing encysted eggs of G. pallida. Their effect on egg viability was estimated by quantifying nematode actin 1 mRNA by RT-qPCR. The leaf glucosinolate profiles of the plants were determined by high-performance liquid chromatography. Three Brassica juncea lines (Nemfix, Fumus, and ISCI99) containing high concentrations of 2-propenyl glucosinolate were the most effective, causing over 95% mortality of encysted eggs of G. pallida in polyethylene-covered soil. The toxic effects of green manures were greater in polyethylene-covered than in open soil. Toxicity in soil correlated with the concentration of isothiocyanate-producing glucosinolate but not total glucosinolate in green manures.

A Straightforward Method for Glucosinolate Extraction and Analysis with High-pressure Liquid Chromatography (HPLC).

PubMed

Grosser, Katharina; van Dam, Nicole M

2017-03-15

Glucosinolates are a well-studied and highly diverse class of natural plant compounds. They play important roles in plant resistance, rapeseed oil quality, food flavoring, and human health. The biological activity of glucosinolates is released upon tissue damage, when they are mixed with the enzyme myrosinase. This results in the formation of pungent and toxic breakdown products, such as isothiocyanates and nitriles. Currently, more than 130 structurally different glucosinolates have been identified. The chemical structure of the glucosinolate is an important determinant of the product that is formed, which in turn determines its biological activity. The latter may range from detrimental (e.g., progoitrin) to beneficial (e.g., glucoraphanin). Each glucosinolate-containing plant species has its own specific glucosinolate profile. For this reason, it is important to correctly identify and reliably quantify the different glucosinolates present in brassicaceous leaf, seed, and root crops or, for ecological studies, in their wild relatives. Here, we present a well-validated, targeted, and robust method to analyze glucosinolate profiles in a wide range of plant species and plant organs. Intact glucosinolates are extracted from ground plant materials with a methanol-water mixture at high temperatures to disable myrosinase activity. Thereafter, the resulting extract is brought onto an ion-exchange column for purification. After sulfatase treatment, the desulfoglucosinolates are eluted with water and the eluate is freeze-dried. The residue is taken up in an exact volume of water, which is analyzed by high-pressure liquid chromatography (HPLC) with a photodiode array (PDA) or ultraviolet (UV) detector. Detection and quantification are achieved by conducting comparisons of the retention times and UV spectra of commercial reference standards. The concentrations are calculated based on a sinigrin reference curve and well-established response factors. The advantages and disadvantages of this straightforward method, when compared to faster and more technologically advanced methods, are discussed here.

A Straightforward Method for Glucosinolate Extraction and Analysis with High-pressure Liquid Chromatography (HPLC)

PubMed Central

Grosser, Katharina; van Dam, Nicole M.

2017-01-01

Glucosinolates are a well-studied and highly diverse class of natural plant compounds. They play important roles in plant resistance, rapeseed oil quality, food flavoring, and human health. The biological activity of glucosinolates is released upon tissue damage, when they are mixed with the enzyme myrosinase. This results in the formation of pungent and toxic breakdown products, such as isothiocyanates and nitriles. Currently, more than 130 structurally different glucosinolates have been identified. The chemical structure of the glucosinolate is an important determinant of the product that is formed, which in turn determines its biological activity. The latter may range from detrimental (e.g., progoitrin) to beneficial (e.g., glucoraphanin). Each glucosinolate-containing plant species has its own specific glucosinolate profile. For this reason, it is important to correctly identify and reliably quantify the different glucosinolates present in brassicaceous leaf, seed, and root crops or, for ecological studies, in their wild relatives. Here, we present a well-validated, targeted, and robust method to analyze glucosinolate profiles in a wide range of plant species and plant organs. Intact glucosinolates are extracted from ground plant materials with a methanol-water mixture at high temperatures to disable myrosinase activity. Thereafter, the resulting extract is brought onto an ion-exchange column for purification. After sulfatase treatment, the desulfoglucosinolates are eluted with water and the eluate is freeze-dried. The residue is taken up in an exact volume of water, which is analyzed by high-pressure liquid chromatography (HPLC) with a photodiode array (PDA) or ultraviolet (UV) detector. Detection and quantification are achieved by conducting comparisons of the retention times and UV spectra of commercial reference standards. The concentrations are calculated based on a sinigrin reference curve and well-established response factors. The advantages and disadvantages of this straightforward method, when compared to faster and more technologically advanced methods, are discussed here. PMID:28362416

Evolution of specifier proteins in glucosinolate-containing plants

PubMed Central

2012-01-01

Background The glucosinolate-myrosinase system is an activated chemical defense system found in plants of the Brassicales order. Glucosinolates are stored separately from their hydrolytic enzymes, the myrosinases, in plant tissues. Upon tissue damage, e.g. by herbivory, glucosinolates and myrosinases get mixed and glucosinolates are broken down to an array of biologically active compounds of which isothiocyanates are toxic to a wide range of organisms. Specifier proteins occur in some, but not all glucosinolate-containing plants and promote the formation of biologically active non-isothiocyanate products upon myrosinase-catalyzed glucosinolate breakdown. Results Based on a phytochemical screening among representatives of the Brassicales order, we selected candidate species for identification of specifier protein cDNAs. We identified ten specifier proteins from a range of species of the Brassicaceae and assigned each of them to one of the three specifier protein types (NSP, nitrile-specifier protein, ESP, epithiospecifier protein, TFP, thiocyanate-forming protein) after heterologous expression in Escherichia coli. Together with nine known specifier proteins and three putative specifier proteins found in databases, we subjected the newly identified specifier proteins to phylogenetic analyses. Specifier proteins formed three major clusters, named AtNSP5-cluster, AtNSP1-cluster, and ESP/TFP cluster. Within the ESP/TFP cluster, specifier proteins grouped according to the Brassicaceae lineage they were identified from. Non-synonymous vs. synonymous substitution rate ratios suggested purifying selection to act on specifier protein genes. Conclusions Among specifier proteins, NSPs represent the ancestral activity. The data support a monophyletic origin of ESPs from NSPs. The split between NSPs and ESPs/TFPs happened before the radiation of the core Brassicaceae. Future analyses have to show if TFP activity evolved from ESPs at least twice independently in different Brassicaceae lineages as suggested by the phylogeny. The ability to form non-isothiocyanate products by specifier protein activity may provide plants with a selective advantage. The evolution of specifier proteins in the Brassicaceae demonstrates the plasticity of secondary metabolism within an activated plant defense system. PMID:22839361

Rapid glucosinolate detection and identification using accurate mass MS-MS

USDA-ARS?s Scientific Manuscript database

Currently, there is a demand for accurate evaluation of brassica plat species for their glucosinolate content. An optimized method has been developed for detecting and identifying glucosinolates in plant extracts using MS-MS fragmentation with ion trap collision induced dissociation (CID) and higher...

Plant plasma membrane aquaporins in natural vesicles as potential stabilizers and carriers of glucosinolates.

PubMed

Martínez-Ballesta, Maria Del Carmen; Pérez-Sánchez, Horacio; Moreno, Diego A; Carvajal, Micaela

2016-07-01

Their biodegradable nature and ability to target cells make biological vesicles potential nanocarriers for bioactives delivery. In this work, the interaction between proteoliposomes enriched in aquaporins derived from broccoli plants and the glucosinolates was evaluated. The vesicles were stored at different temperatures and their integrity was studied. Determination of glucosinolates, showed that indolic glucosinolates were more sensitive to degradation in aqueous solution than aliphatic glucosinolates. Glucoraphanin was stabilized by leaf and root proteoliposomes at 25°C through their interaction with aquaporins. An extensive hydrogen bond network, including different aquaporin residues, and hydrophobic interactions, as a consequence of the interaction between the linear alkane chain of glucoraphanin and Glu31 and Leu34 protein residues, were established as the main stabilizing elements. Combined our results showed that plasma membrane vesicles from leaf and root tissues of broccoli plants may be considered as suitable carriers for glucosinolate which stabilization can be potentially attributed to aquaporins. Copyright © 2016 Elsevier B.V. All rights reserved.

Identification of Metabolic QTLs and Candidate Genes for Glucosinolate Synthesis in Brassica oleracea Leaves, Seeds and Flower Buds

PubMed Central

Sotelo, Tamara; Soengas, Pilar; Velasco, Pablo; Rodríguez, Víctor M.; Cartea, María Elena

2014-01-01

Glucosinolates are major secondary metabolites found in the Brassicaceae family. These compounds play an essential role in plant defense against biotic and abiotic stresses, but more interestingly they have beneficial effects on human health. We performed a genetic analysis in order to identify the genome regions regulating glucosinolates biosynthesis in a DH mapping population of Brassica oleracea. In order to obtain a general overview of regulation in the whole plant, analyses were performed in the three major organs where glucosinolates are synthesized (leaves, seeds and flower buds). Eighty two significant QTLs were detected, which explained a broad range of variability in terms of individual and total glucosinolate (GSL) content. A meta-analysis rendered eighteen consensus QTLs. Thirteen of them regulated more than one glucosinolate and its content. In spite of the considerable variability of glucosinolate content and profiles across the organ, some of these consensus QTLs were identified in more than one tissue. Consensus QTLs control the GSL content by interacting epistatically in complex networks. Based on in silico analysis within the B. oleracea genome along with synteny with Arabidopsis, we propose seven major candidate loci that regulate GSL biosynthesis in the Brassicaceae family. Three of these loci control the content of aliphatic GSL and four of them control the content of indolic glucosinolates. GSL-ALK plays a central role in determining aliphatic GSL variation directly and by interacting epistatically with other loci, thus suggesting its regulatory effect. PMID:24614913

Effect of Brassicaceae seed meals with different glucosinolate profiles on Rhizoctonia root rot of wheat

USDA-ARS?s Scientific Manuscript database

Tissues of plants in the family Brassicaceae contain glucosinolates, compounds whose hydrolysis results in the release of various bioactive products including isothiocyanates. The broad spectrum of biological activity of these glucosinolate hydrolysis products has led to the promotion of brassicace...

Effects of glucosinolates and their hydrolysis products on biochemical and performance parameters in broiler chicken diets

USDA-ARS?s Scientific Manuscript database

Glucosinolates are important bioactive molecules and widely found in Brassicaceae species (cress, brussels sprouts, mustard, broccoli, kale, etc.). Depending on the amount of these vegetables consumed, both positive and negative metabolic effects from glucosinolate metabolites may occur. The aim of ...

Effects of glucosinolates and their hydrolysis products on biochemical and performance parameters in broiler chicken diets

USDA-ARS?s Scientific Manuscript database

Glucosinolates are important bioactive molecules and widely found in Brassicaceae species (cress, brussels sprouts, mustard, broccoli, kale etc.). Depending on the amount of these vegetables consumed, both positive and negative metabolic effects from glucosinolate metabolites may occur. The aims of ...

Macroevolutionary patterns of glucosinolate defense and tests of defense-escalation and resource availability hypotheses.

PubMed

Cacho, N Ivalú; Kliebenstein, Daniel J; Strauss, Sharon Y

2015-11-01

We explored macroevolutionary patterns of plant chemical defense in Streptanthus (Brassicaceae), tested for evolutionary escalation of defense, as predicted by Ehrlich and Raven's plant-herbivore coevolutionary arms-race hypothesis, and tested whether species inhabiting low-resource or harsh environments invest more in defense, as predicted by the resource availability hypothesis (RAH). We conducted phylogenetically explicit analyses using glucosinolate profiles, soil nutrient analyses, and microhabitat bareness estimates across 30 species of Streptanthus inhabiting varied environments and soils. We found weak to moderate phylogenetic signal in glucosinolate classes and no signal in total glucosinolate production; a trend toward evolutionary de-escalation in the numbers and diversity of glucosinolates, accompanied by an evolutionary increase in the proportion of aliphatic glucosinolates; some support for the RAH relative to soil macronutrients, but not relative to serpentine soil use; and that the number of glucosinolates increases with microhabitat bareness, which is associated with increased herbivory and drought. Weak phylogenetic signal in chemical defense has been observed in other plant systems. A more holistic approach incorporating other forms of defense might be necessary to confidently reject escalation of defense. That defense increases with microhabitat bareness supports the hypothesis that habitat bareness is an underappreciated selective force on plants in harsh environments. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

Profiling of Glucosinolates and Flavonoids in Rorippa indica (Linn.) Hiern. (Cruciferae) by UHPLC-PDA-ESI/HRMSn.

USDA-ARS?s Scientific Manuscript database

An UHPLC-DAD-ESI/HRMSn profiling method was used to identify the glucosinolates and flavonoids of Rorippa montana (Cruciferae), a Chinese herb used to treat cough, diarrhea and rheumatoid arthritis. Thirty three glucosinolates, over 40 flavonol glycosides, and more than 20 other phenolic and common ...

Glucosinolate diversity within a phylogenetic framework of the tribe Cardamineae (Brassicaceae) unraveled with HPLC-MS/MS and NMR-based analytical distinction of 70 desulfoglucosinolates.

PubMed

Olsen, Carl Erik; Huang, Xiao-Chen; Hansen, Cecilie I C; Cipollini, Don; Ørgaard, Marian; Matthes, Annemarie; Geu-Flores, Fernando; Koch, Marcus A; Agerbirk, Niels

2016-12-01

As a basis for future investigations of evolutionary trajectories and biosynthetic mechanisms underlying variations in glucosinolate structures, we screened members of the crucifer tribe Cardamineae by HPLC-MS/MS, isolated and identified glucosinolates by NMR, searched the literature for previous data for the tribe, and collected HPLC-MS/MS data for nearly all glucosinolates known from the tribe as well as some related structures (70 in total). This is a considerable proportion of the approximately 142 currently documented natural glucosinolates. Calibration with authentic references allowed distinction (or elucidation) of isomers in many cases, such as distinction of β-hydroxyls, methylthios, methylsulfinyls and methylsulfonyls. A mechanism for fragmentation of secondary β-hydroxyls in MS was elucidated, and two novel glucosinolates were discovered: 2-hydroxy-3-methylpentylglucosinolate in roots of Cardamine pratensis and 2-hydroxy-8-(methylsulfinyl)octylglucosinolate in seeds of Rorippa amphibia. A large number of glucosinolates (ca. 54 with high structural certainty and a further 28 or more suggested from tandem MS), representing a wide structural variation, is documented from the tribe. This included glucosinolates apparently derived from Met, Phe, Trp, Val/Leu, Ile and higher homologues. Normal side chain elongation and side chain decoration by oxidation or methylation was observed, as well as rare abnormal side chain decoration (hydroxylation of aliphatics at the δ rather than β-position). Some species had diverse profiles, e.g. R. amphibia and C. pratensis (19 and 16 individual glucosinolates, respectively), comparable to total diversity in literature reports of Armoracia rusticana (17?), Barbarea vulgaris (20-24), and Rorippa indica (>20?). The ancestor or the tribe would appear to have used Trp, Met, and homoPhe as glucosinolate precursor amino acids, and to exhibit oxidation of thio to sulfinyl, formation of alkenyls, β-hydroxylation of aliphatic chains and hydroxylation and methylation of indole glucosinolates. Two hotspots of apparent biochemical innovation and loss were identified: C. pratensis and the genus Barbarea. Diversity in other species mainly included structures also known from other crucifers. In addition to a role of gene duplication, two contrasting genetic/biochemical mechanisms for evolution of such combined diversity and redundancy are discussed: (i) involvement of widespread genes with expression varying during evolution, and (ii) mutational changes in substrate specificities of CYP79F and GS-OH enzymes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Biosynthesis of Mustard Oil Glucosides: Sodium Phenylacetothiohydroximate and Desulfobenzylglucosinolate, Precursors of Benzylglucosinolate in Tropaeolum majus1

PubMed Central

Underhill, L. E. W.; Wetter, L. R.

1969-01-01

The biosynthesis of the mustard oil glucoside, benzylglucosinolate, was studied in Tropaeolum majus L. A number of labeled compounds were administered to plant shoots and the incorporation of tracer into benzylglucosinolate, isolated as the crystalline tetramethyl-ammonium salt, was measured. In order of decreasing efficiency of conversion into benzyl-glucosinolate the compounds fed were S-(β-d-glucopyranosyl)phenylacetothiohydroximic acid (desulfobenzylglucosinolate), sodium phenylacetothiohydroximate, dl-phenylalanine, d-glucose, and sodium-d-1-glucopyranosyl mercaptide (1-thioglucose). The results are consistent with the hypothesis that the thioglucosyl group of benzylglucosinolate is derived by glucosylation of phenylacetothiohydroximate and not from 1-thioglucose. The results also suggest that benzylglucosinolate is formed by sulfation of desulfobenzylglucosinolate as the final step in its biosynthesis. A method for the isolation of a number of glucosinolates (mustard oil glucosides) is described which utilizes anion exchange chromatography on diethylaminoethyl (DEAE) cellulose. Potassium allylglucosinolate, tetramethylammonium benzylglucosinolate, potassium 2-hydroxy-2-phenylethylglucosinolate and potassium 2-phenylethylglucosinolate were obtained on recrystallization of the glucosinolate fraction eluted from the column. PMID:16657104

Mutation of the Glucosinolate Biosynthesis Enzyme Cytochrome P450 83A1 Monooxygenase Increases Camalexin Accumulation and Powdery Mildew Resistance.

PubMed

Liu, Simu; Bartnikas, Lisa M; Volko, Sigrid M; Ausubel, Frederick M; Tang, Dingzhong

2016-01-01

Small secondary metabolites, including glucosinolates and the major phytoalexin camalexin, play important roles in immunity in Arabidopsis thaliana. We isolated an Arabidopsis mutant with increased resistance to the powdery mildew fungus Golovinomyces cichoracearum and identified a mutation in the gene encoding cytochrome P450 83A1 monooxygenase (CYP83A1), which functions in glucosinolate biosynthesis. The cyp83a1-3 mutant exhibited enhanced defense responses to G. cichoracearum and double mutant analysis showed that this enhanced resistance requires NPR1, EDS1, and PAD4, but not SID2 or EDS5. In cyp83a1-3 mutants, the expression of genes related to camalexin synthesis increased upon G. cichoracearum infection. Significantly, the cyp83a1-3 mutant also accumulated higher levels of camalexin. Decreasing camalexin levels by mutation of the camalexin synthetase gene PAD3 or the camalexin synthesis regulator AtWRKY33 compromised the powdery mildew resistance in these mutants. Consistent with these observations, overexpression of PAD3 increased camalexin levels and enhanced resistance to G. cichoracearum. Taken together, our data indicate that accumulation of higher levels of camalexin contributes to increased resistance to powdery mildew.

Mutation of the Glucosinolate Biosynthesis Enzyme Cytochrome P450 83A1 Monooxygenase Increases Camalexin Accumulation and Powdery Mildew Resistance

PubMed Central

Liu, Simu; Bartnikas, Lisa M.; Volko, Sigrid M.; Ausubel, Frederick M.; Tang, Dingzhong

2016-01-01

Small secondary metabolites, including glucosinolates and the major phytoalexin camalexin, play important roles in immunity in Arabidopsis thaliana. We isolated an Arabidopsis mutant with increased resistance to the powdery mildew fungus Golovinomyces cichoracearum and identified a mutation in the gene encoding cytochrome P450 83A1 monooxygenase (CYP83A1), which functions in glucosinolate biosynthesis. The cyp83a1-3 mutant exhibited enhanced defense responses to G. cichoracearum and double mutant analysis showed that this enhanced resistance requires NPR1, EDS1, and PAD4, but not SID2 or EDS5. In cyp83a1-3 mutants, the expression of genes related to camalexin synthesis increased upon G. cichoracearum infection. Significantly, the cyp83a1-3 mutant also accumulated higher levels of camalexin. Decreasing camalexin levels by mutation of the camalexin synthetase gene PAD3 or the camalexin synthesis regulator AtWRKY33 compromised the powdery mildew resistance in these mutants. Consistent with these observations, overexpression of PAD3 increased camalexin levels and enhanced resistance to G. cichoracearum. Taken together, our data indicate that accumulation of higher levels of camalexin contributes to increased resistance to powdery mildew. PMID:26973671

Biofumigation with Brassica juncea, Raphanus sativus and Eruca sativa for the management of field populations of the potato cyst nematode Globodera pallida.

PubMed

Ngala, Bruno M; Haydock, Patrick P J; Woods, Simon; Back, Matthew A

2015-05-01

The viability of potato cyst nematode (PCN) populations (Globodera pallida) was evaluated in three field experiments using Brassica juncea, Raphanus sativus and Eruca sativa amendments. These species were summer cultivated and autumn incorporated in experiment 1; in experiment 2, overwintered brassicaceous cover crops were spring incorporated. Experiment 3 involved determination of effects of metconazole application on biomass/glucosinolate production by B. juncea and R. sativus and on PCN pre- and post-incorporation. Glucosinolate contents were determined before incorporation. Following cover crop incorporation, field plots were planted with susceptible potatoes to evaluate the biofumigation effects on PCN reproduction. In experiment 1, PCN population post-potato harvest was reduced (P = 0.03) in B. juncea-treated plots, while R. sativus prevented further multiplication, but in experiment 2 there were no significant effects on PCN reproduction. In experiment 3, B. juncea or R. sativus either untreated or treated with metconazole reduced PCN populations. Glucosinolate concentrations varied significantly between different plant regions and cultivation seasons. Metconazole application increased the sinigrin concentration in B. juncea tissues. Glucosinolate concentrations correlated positively with PCN mortality for summer-cultivated brassicaceous plants. The results demonstrated that B. juncea and R. sativus green manures can play an important role in PCN management, particularly if included in an integrated pest management scheme. © 2014 Society of Chemical Industry.

Effects of Organic and Waste-Derived Fertilizers on Yield, Nitrogen and Glucosinolate Contents, and Sensory Quality of Broccoli (Brassica oleracea L. var. italica).

PubMed

Øvsthus, Ingunn; Breland, Tor Arvid; Hagen, Sidsel Fiskaa; Brandt, Kirsten; Wold, Anne-Berit; Bengtsson, Gunnar B; Seljåsen, Randi

2015-12-23

Organic vegetable production attempts to pursue multiple goals concerning influence on environment, production resources, and human health. In areas with limited availability of animal manure, there is a need for considering various off-farm nutrient resources for such production. Different organic and waste-derived fertilizer materials were used for broccoli production at two latitudes (58° and 67°) in Norway during two years. The fertilizer materials were applied at two rates of total N (80 and 170 kg ha(-1)) and compared with mineral fertilizer (170 kg ha(-1)) and no fertilizer. Broccoli yield was strongly influenced by fertilizer materials (algae meal < unfertilized control < sheep manure < extruded shrimp shell < anaerobically digested food waste < mineral fertilizer). Yield, but not glucosinolate content, was linearly correlated with estimated potentially plant-available N. However, extruded shrimp shell and mineral NPK fertilizer gave higher glucosinolate contents than sheep manure and no fertilizer. Sensory attributes were less affected by fertilizer material and plant-available N.

Cytotoxic and genotoxic potential of food-borne nitriles in a liver in vitro model

PubMed Central

Kupke, Franziska; Herz, Corinna; Hanschen, Franziska S.; Platz, Stefanie; Odongo, Grace A.; Helmig, Simone; Bartolomé Rodríguez, María M.; Schreiner, Monika; Rohn, Sascha; Lamy, Evelyn

2016-01-01

Isothiocyanates are the most intensively studied breakdown products of glucosinolates from Brassica plants and well recognized for their pleiotropic effects against cancer but also for their genotoxic potential. However, knowledge about the bioactivity of glucosinolate-borne nitriles in foods is very poor. As determined by GC-MS, broccoli glucosinolates mainly degrade to nitriles as breakdown products. The cytotoxicity of nitriles in human HepG2 cells and primary murine hepatocytes was marginal as compared to isothiocyanates. Toxicity of nitriles was not enhanced in CYP2E1-overexpressing HepG2 cells. In contrast, the genotoxic potential of nitriles was found to be comparable to isothiocyanates. DNA damage was persistent over a certain time period and CYP2E1-overexpression further increased the genotoxic potential of the nitriles. Based on actual in vitro data, no indications are given that food-borne nitriles could be relevant for cancer prevention, but could pose a certain genotoxic risk under conditions relevant for food consumption. PMID:27883018

Arabidopsis myrosinases link the glucosinolate-myrosinase system and the cuticle

PubMed Central

Ahuja, Ishita; de Vos, Ric C. H.; Rohloff, Jens; Stoopen, Geert M.; Halle, Kari K.; Ahmad, Samina Jam Nazeer; Hoang, Linh; Hall, Robert D.; Bones, Atle M.

2016-01-01

Both physical barriers and reactive phytochemicals represent two important components of a plant’s defence system against environmental stress. However, these two defence systems have generally been studied independently. Here, we have taken an exclusive opportunity to investigate the connection between a chemical-based plant defence system, represented by the glucosinolate-myrosinase system, and a physical barrier, represented by the cuticle, using Arabidopsis myrosinase (thioglucosidase; TGG) mutants. The tgg1, single and tgg1 tgg2 double mutants showed morphological changes compared to wild-type plants visible as changes in pavement cells, stomatal cells and the ultrastructure of the cuticle. Extensive metabolite analyses of leaves from tgg mutants and wild-type Arabidopsis plants showed altered levels of cuticular fatty acids, fatty acid phytyl esters, glucosinolates, and indole compounds in tgg single and double mutants as compared to wild-type plants. These results point to a close and novel association between chemical defence systems and physical defence barriers. PMID:27976683

Taste detection of the non-volatile isothiocyanate moringin results in deterrence to glucosinolate-adapted insect larvae.

PubMed

Müller, Caroline; van Loon, Joop; Ruschioni, Sara; De Nicola, Gina Rosalinda; Olsen, Carl Erik; Iori, Renato; Agerbirk, Niels

2015-10-01

Isothiocyanates (ITCs), released from Brassicales plants after hydrolysis of glucosinolates, are known for their negative effects on herbivores but mechanisms have been elusive. The ITCs are initially present in dissolved form at the site of herbivore feeding, but volatile ITCs may subsequently enter the gas phase and all ITCs may react with matrix components. Deterrence to herbivores resulting from topically applied volatile ITCs in artificial feeding assays may hence lead to ambiguous conclusions. In the present study, the non-volatile ITC moringin (4-(α-L-rhamnopyranosyloxy)benzyl ITC) and its glucosinolate precursor glucomoringin were examined for effects on behaviour and taste physiology of specialist insect herbivores of Brassicales. In feeding bioassays, glucomoringin was not deterrent to larvae of Pieris napi (Lepidoptera: Pieridae) and Athalia rosae (Hymenoptera: Tenthredinidae), which are adapted to glucosinolates. Glucomoringin stimulated feeding of larvae of the related Pieris brassicae (Lepidoptera: Pieridae) and also elicited electrophysiological activity from a glucosinolate-sensitive gustatory neuron in the lateral maxillary taste sensilla. In contrast, the ITC moringin was deterrent to P. napi and P. brassicae at high levels and to A. rosae at both high and low levels when topically applied to cabbage leaf discs (either 12, 120 or 1200 nmol moringin per leaf disc of 1cm diameter). Survival of A. rosae was also significantly reduced when larvae were kept on leaves treated with moringin for several days. Furthermore, moringin elicited electrophysiological activity in a deterrent-sensitive neuron in the medial maxillary taste sensillum of P. brassicae, providing a sensory mechanism for the deterrence and the first known ITC taste response of an insect. In simulated feeding assays, recovery of moringin was high, in accordance with its non-volatile nature. Our results demonstrate taste-mediated deterrence of a non-volatile, natural ITC to glucosinolate-adapted insects. Copyright © 2015 Elsevier Ltd. All rights reserved.

Determination of glucosinolates in rapeseed and Thlaspi caerulescens plants by liquid chromatography-atmospheric pressure chemical ionization mass spectrometry.

PubMed

Tolrà, R P; Alonso, R; Poschenrieder, C; Barceló, D; Barceló, J

2000-08-11

Liquid chromatography-atmospheric pressure chemical ionization mass spectrometry was used to identify glucosinolates in plant extracts. Optimization of the analytical conditions and the determination of the method detection limit was performed using commercial 2-propenylglucosinolate (sinigrin). Optimal values for the following parameters were determined: nebulization pressure, gas temperature, flux of drying gas, capillar voltage, corona current and fragmentor conditions. The method detection limit for sinigrin was 2.85 ng. For validation of the method the glucosinolates in reference material (rapeseed) from the Community Bureau of Reference Materials (BCR) were analyzed. The method was applied for the determination of glucosinolates in Thlaspi caerulescens plants.

CB5C affects the glucosinolate profile in Arabidopsis thaliana

PubMed Central

Vik, Daniel; Crocoll, Christoph; Andersen, Tonni Grube; Burow, Meike; Halkier, Barbara Ann

2016-01-01

ABSTRACT Cytochrome b5 (CB5) proteins are small heme-binding proteins, that influence cytochrome P450 activity. While only one CB5 isoform is found in mammals, higher plants have several isoforms of these proteins. The roles of the many CB5 isoforms in plants remain unknown. We hypothesized that CB5 proteins support the cytochrome P450 enzymes of plant specialized metabolism and found CB5C from Arabidopsis thaliana to co-express with glucosinolate biosynthetic genes. We characterized the glucosinolate profiles of 2 T-DNA insertion mutants of CB5C, and found that long-chained aliphatic glucosinolates were reduced in one of the mutant lines – a phenotype that was exaggerated upon methyl-jasmonate treatment. These results support the hypothesis, that CB5C influences glucosinolate biosynthesis, however, the mode of action remains unknown. Furthermore, the mutants differed in their biomass response to methyl jasmonate treatment. Thereby, our results highlight the varying effects of T-DNA insertion sites, as the 2 analyzed alleles show different phenotypes. PMID:27454255

Flavor, glucosinolates, and isothiocyanates of nau (Cook's scurvy grass, Lepidium oleraceum) and other rare New Zealand Lepidium species.

PubMed

Sansom, Catherine E; Jones, Veronika S; Joyce, Nigel I; Smallfield, Bruce M; Perry, Nigel B; van Klink, John W

2015-02-18

The traditionally consumed New Zealand native plant nau, Cook's scurvy grass, Lepidium oleraceum, has a pungent wasabi-like taste, with potential for development as a flavor ingredient. The main glucosinolate in this Brassicaceae was identified by LC-MS and NMR spectroscopy as 3-butenyl glucosinolate (gluconapin, 7-22 mg/g DM in leaves). The leaves were treated to mimic chewing, and the headspace was analyzed by solid-phase microextraction and GC-MS. This showed that 3-butenyl isothiocyanate, with a wasabi-like flavor, was produced by the endogenous myrosinase. Different postharvest treatments were used to create leaf powders as potential flavor products, which were tasted and analyzed for gluconapin and release of 3-butenyl isothiocyanate. A high drying temperature (75 °C) did not give major glucosinolate degradation, but did largely inactivate the myrosinase, resulting in no wasabi-like flavor release. Drying at 45 °C produced more pungent flavor than freeze-drying. Seven other Lepidium species endemic to New Zealand were also analyzed to determine their flavor potential and also whether glucosinolates were taxonomic markers. Six contained mostly gluconapin, but the critically endangered Lepidium banksii had a distinct composition including isopropyl glucosinolate, not detected in the other species.

Glucosinolates in broccoli sprouts (Brassica oleracea var. italica) as conditioned by sulphate supply during germination.

PubMed

Pérez-Balibrea, Santiago; Moreno, Diego A; García-Viguera, Cristina

2010-10-01

Sulphur (S) fertilization is essential for primary and secondary metabolism in cruciferous foods. Deficient, suboptimal, or excessive S affects the growth and biosynthesis of secondary metabolites in adult plants. Nevertheless, there is little information regarding the influence of S fertilization on sprouts and seedlings. An experiment was set up to evaluate the effect of S fertilization, supplied as K(2)SO(4) at 0, 15, 30, and 60 mg/L, on the glucosinolate content of broccoli sprouts during the germination course of 3, 6, 9, and 12 d after sowing. Glucosinolate concentration was strongly influenced by germination, causing a rapid increase during the first 3 d after sowing, and decreasing afterwards. The S supply increased aliphatic and total glucosinolate content at the end of the monitored sprouting period. S-treated sprouts, with S(15), S(30), and S(60) at 9 and 12 d after sowing presented enhanced glucosinolate content. Overall, both germination time and S fertilization were key factors in maximizing the bioactive health-promoting phytochemicals of broccoli. Practical Application: Germination with sulphate is a simple and inexpensive way to obtain sprouts that contain much higher levels of glucosinolates (health promoting compounds), than the corresponding florets from the same seeds.

Two Young MicroRNAs Originating from Target Duplication Mediate Nitrogen Starvation Adaptation via Regulation of Glucosinolate Synthesis in Arabidopsis thaliana1[W

PubMed Central

He, Hua; Liang, Gang; Li, Yang; Wang, Fang; Yu, Diqiu

2014-01-01

Nitrogen is an essential macronutrient required for plant growth and development. A number of genes respond to nitrogen starvation conditions. However, the functions of most of these nitrogen starvation-responsive genes are unclear. Our recent survey suggested that many microRNAs (miRNAs) are responsive to nitrogen starvation in Arabidopsis thaliana. Here, we identified a new miRNA (miR5090) from the complementary transcript of the MIR826 gene. Further investigation uncovered that both miRNA genes recently evolved from the inverse duplication of their common target gene, ALKENYL HYDROXALKYL PRODUCING2 (AOP2). Similar to miR826, miR5090 is induced by nitrogen starvation. By contrast, the AOP2 transcript level was negatively correlated with miR826 and miR5090 under nitrogen starvation. GUS-fused AOP2 expression suggested that AOP2 was posttranscriptionally suppressed by miR826 and miR5090. miRNA transgenic plants with significantly low AOP2 expression accumulated fewer Met-derived glucosinolates, phenocopying the aop2 mutants. Most glucosinolate synthesis-associated genes were repressed under nitrogen starvation conditions. Furthermore, miRNA transgenic plants with less glucosinolate displayed enhanced tolerance to nitrogen starvation, including high biomass, more lateral roots, increased chlorophyll, and decreased anthocyanin. Meanwhile, nitrogen starvation-responsive genes were up-regulated in transgenic plants, implying improved nitrogen uptake activity. Our study reveals a mechanism by which Arabidopsis thaliana regulates the synthesis of glucosinolates to adapt to environmental changes in nitrogen availability. PMID:24367020

Plants as Biofactories: Postharvest Stress-Induced Accumulation of Phenolic Compounds and Glucosinolates in Broccoli Subjected to Wounding Stress and Exogenous Phytohormones

PubMed Central

Villarreal-García, Daniel; Nair, Vimal; Cisneros-Zevallos, Luis; Jacobo-Velázquez, Daniel A.

2016-01-01

Broccoli contains high levels of bioactive molecules and is considered a functional food. In this study, postharvest treatments to enhance the concentration of glucosinolates and phenolic compounds were evaluated. Broccoli whole heads were wounded to obtain florets and wounded florets (florets cut into four even pieces) and stored for 24 h at 20 °C with or without exogenous ethylene (ET, 1000 ppm) or methyl jasmonate (MeJA, 250 ppm). Whole heads were used as a control for wounding treatments. Regarding glucosinolate accumulation, ET selectively induced the 4-hydroxylation of glucobrassicin in whole heads, resulting in ∼223% higher 4-hydroxyglucobrassicin than time 0 h samples. Additionally, glucoraphanin was increased by ∼53% in whole heads treated with ET, while neoglucobrassicin was greatly accumulated in wounded florets treated with ET or MeJA, showing increases of ∼193 and ∼286%, respectively. On the other hand, although only whole heads stored without phytohormones showed higher concentrations of phenolic compounds, which was reflected in ∼33, ∼30, and ∼46% higher levels of 1,2,2-trisinapoylgentiobose, 1,2-diferulolylgentiobiose, and 1,2-disinapoyl-2-ferulolylgentiobiose, respectively; broccoli florets stored under air control conditions showed enhanced concentrations of 3-O-caffeoylquinic acid, 1,2-disinapoylgentiobiose, and 1,2-disinapoyl-2-ferulolylgentiobiose (∼22, ∼185, and ∼65% more, respectively). Furthermore, exogenous ET and MeJA impeded individual phenolics accumulation. Results allowed the elucidation of simple and effective postharvest treatment to enhance the content of individual glucosinolates and phenolic compounds in broccoli. The stressed-broccoli tissue could be subjected to downstream processing in order to extract and purify bioactive molecules with applications in the dietary supplements, agrochemical and cosmetics markets. PMID:26904036

Jasmonate-Dependent Induction of Indole Glucosinolates in Arabidopsis by Culture Filtrates of the Nonspecific Pathogen Erwinia carotovora1

PubMed Central

Brader, Günter; Tas, Éva; Palva, E. Tapio

2001-01-01

Elicitors from the plant pathogen Erwinia carotovora trigger coordinate induction of the tryptophan (Trp) biosynthesis pathway and Trp oxidizing genes in Arabidopsis. To elucidate the biological role of such pathogen-induced activation we characterized the production of secondary defense metabolites such as camalexin and indole glucosinolates derived from precursors of this pathway. Elicitor induction was followed by a specific increase in 3-indolylmethylglucosinolate (IGS) content, but only a barely detectable accumulation of the indole-derived phytoalexin camalexin. The response is mediated by jasmonic acid as shown by lack of IGS induction in the jasmonate-insensitive mutant coi1-1. In accordance with this, methyl jasmonate was able to trigger IGS accumulation in Arabidopsis. In contrast, ethylene and salicylic acid seem to play a minor role in the response. They did not trigger alterations in IGS levels, and methyl jasmonate- or elicitor-induced IGS accumulation in NahG and ethylene-insensitive ein2-1 mutant plants was similar as in the wild type. The breakdown products of IGS and other glucosinolates were able to inhibit growth of E. carotovora. The results suggest that IGS is of importance in the defense against bacterial pathogens. PMID:11402212

Exploring genotypic variations for improved oil content and healthy fatty acids composition in rapeseed (Brassica napus L.).

PubMed

Ishaq, Muhammad; Razi, Raziuddin; Khan, Sabaz Ali

2017-04-01

Development of new genotypes having high oil content and desirable levels of fatty acid compositions is a major objective of rapeseed breeding programmes. In the current study combining ability was determined for oil, protein, glucosinolates and various fatty acids content using 8 × 8 full diallel in rapeseed (Brassica napus). Highly significant genotypic differences were observed for oil, protein, glucosinolates, oleic acid, linolenic acid and erucic acid content. Mean squares due to general combining ability (GCA), specific combining ability (SCA) and reciprocal combining ability (RCA) were highly significant (P ≤ 0.01) for biochemical traits. Parental line AUP-17 for high oil content and low glucosinolates, genotype AUP-2 for high protein and oleic acids, and AUP-18 for low lenolenic and erucic acid were best general combiners. Based on desirable SCA effects, F 1 hybrids AUP-17 × AUP-20; AUP-2 × AUP-8; AUP-7 × AUP-14; AUP-2 × AUP-9; AUP-7 × AUP-14 and AUP-2 × AUP-9 were found superior involving at least one best general combiner. F 1 hybrids AUP-17 × AUP-20 (for oil content); AUP-2 × AUP-8 (for protein content); AUP-7 × AUP-14 (for glucosinolates); AUP-2 × AUP-9 (for oleic acid); AUP-7 × AUP-14 (for linolenic acid) and AUP-2 × AUP-9 (for erucic acid) were found superior involving at least one best general combiner. As reciprocal crosses of AUP-14 with AUP-7 and AUP-8 were superior had low × low and low × high GCA effects for glucosinolates and oleic acid, respectively therefore, these could be exploited in future rapeseed breeding programmes to develop new lines with good quality. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

UVA, UVB Light, and Methyl Jasmonate, Alone or Combined, Redirect the Biosynthesis of Glucosinolates, Phenolics, Carotenoids, and Chlorophylls in Broccoli Sprouts

PubMed Central

Moreira-Rodríguez, Melissa; Benavides, Jorge

2017-01-01

Broccoli sprouts contain health-promoting phytochemicals that can be enhanced by applying ultraviolet light (UV) or phytohormones. The separate and combined effects of methyl jasmonate (MJ), UVA, or UVB lights on glucosinolate, phenolic, carotenoid, and chlorophyll profiles were assessed in broccoli sprouts. Seven-day-old broccoli sprouts were exposed to UVA (9.47 W/m2) or UVB (7.16 W/m2) radiation for 120 min alone or in combination with a 25 µM MJ solution, also applied to sprouts without UV supplementation. UVA + MJ and UVB + MJ treatments increased the total glucosinolate content by ~154% and ~148%, respectively. MJ induced the biosynthesis of indole glucosinolates, especially neoglucobrassicin (~538%), showing a synergistic effect with UVA stress. UVB increased the content of aliphatic and indole glucosinolates, such as glucoraphanin (~78%) and 4-methoxy-glucobrassicin (~177%). UVA increased several phenolics such as gallic acid (~57%) and a kaempferol glucoside (~25.4%). MJ treatment decreased most phenolic levels but greatly induced accumulation of 5-sinapoylquinic acid (~239%). MJ treatments also reduced carotenoid and chlorophyll content, while UVA increased lutein (~23%), chlorophyll b (~31%), neoxanthin (~34%), and chlorophyll a (~67%). Results indicated that UV- and/or MJ-treated broccoli sprouts redirect the carbon flux to the biosynthesis of specific glucosinolates, phenolics, carotenoids, and chlorophylls depending on the type of stress applied. PMID:29113068

UVA, UVB Light, and Methyl Jasmonate, Alone or Combined, Redirect the Biosynthesis of Glucosinolates, Phenolics, Carotenoids, and Chlorophylls in Broccoli Sprouts.

PubMed

Moreira-Rodríguez, Melissa; Nair, Vimal; Benavides, Jorge; Cisneros-Zevallos, Luis; Jacobo-Velázquez, Daniel A

2017-11-04

Broccoli sprouts contain health-promoting phytochemicals that can be enhanced by applying ultraviolet light (UV) or phytohormones. The separate and combined effects of methyl jasmonate (MJ), UVA, or UVB lights on glucosinolate, phenolic, carotenoid, and chlorophyll profiles were assessed in broccoli sprouts. Seven-day-old broccoli sprouts were exposed to UVA (9.47 W/m²) or UVB (7.16 W/m²) radiation for 120 min alone or in combination with a 25 µM MJ solution, also applied to sprouts without UV supplementation. UVA + MJ and UVB + MJ treatments increased the total glucosinolate content by ~154% and ~148%, respectively. MJ induced the biosynthesis of indole glucosinolates, especially neoglucobrassicin (~538%), showing a synergistic effect with UVA stress. UVB increased the content of aliphatic and indole glucosinolates, such as glucoraphanin (~78%) and 4-methoxy-glucobrassicin (~177%). UVA increased several phenolics such as gallic acid (~57%) and a kaempferol glucoside (~25.4%). MJ treatment decreased most phenolic levels but greatly induced accumulation of 5-sinapoylquinic acid (~239%). MJ treatments also reduced carotenoid and chlorophyll content, while UVA increased lutein (~23%), chlorophyll b (~31%), neoxanthin (~34%), and chlorophyll a (~67%). Results indicated that UV- and/or MJ-treated broccoli sprouts redirect the carbon flux to the biosynthesis of specific glucosinolates, phenolics, carotenoids, and chlorophylls depending on the type of stress applied.

Arbuscular mycorrhizal fungi affect glucosinolate and mineral element composition in leaves of Moringa oleifera.

PubMed

Cosme, Marco; Franken, Philipp; Mewis, Inga; Baldermann, Susanne; Wurst, Susanne

2014-10-01

Moringa is a mycorrhizal crop cultivated in the tropics and subtropics and appreciated for its nutritive and health-promoting value. As well as improving plant mineral nutrition, arbuscular mycorrhizal fungi (AMF) can affect plant synthesis of compounds bioactive against chronic diseases in humans. Rhizophagus intraradices and Funneliformis mosseae were used in a full factorial experiment to investigate the impact of AMF on the accumulation of glucosinolates, flavonoids, phenolic acids, carotenoids, and mineral elements in moringa leaves. Levels of glucosinolates were enhanced, flavonoids and phenolic acids were not affected, levels of carotenoids (including provitamin A) were species-specifically reduced, and mineral elements were affected differently, with only Cu and Zn being increased by the AMF. This study presents novel results on AMF effects on glucosinolates in leaves and supports conclusions that the impacts of these fungi on microelement concentrations in edible plants are species dependent. The nonspecific positive effects on glucosinolates and the species-specific negative effects on carotenoids encourage research on other AMF species to achieve general benefits on bioactive compounds in moringa.

High pressure effects on myrosinase activity and glucosinolate preservation in seedlings of Brussels sprouts.

PubMed

Wang, Jia; Barba, Francisco J; Sørensen, Jens C; Frandsen, Heidi B; Sørensen, Susanne; Olsen, Karsten; Orlien, Vibeke

2018-04-15

Combinations of pressure, temperature and time (100-600 MPa, 30-60 °C, 3-10 min) influence enzyme activity of the myrosinase-glucosinolate system. Seedlings of Brussels sprouts were used as a model, which constitutes a well-defined and homogenous sample matrix with simple cell structures. A response surface methodology approach was used to determine the combined effect of pressure level, temperature and time on glucosinolate concentration and myrosinase activity in Brussels sprouts seedlings. The effects on residual myrosinase activity and intact glucosinolate concentration differed according to combinations of pressure, time and temperature. The results showed that maximum inactivation of myrosinase and preservation of glucosinolate (85% of the untreated level) was obtained after HP treatment at 600 MPa, 60 °C, 10 min. The highest preservation of myrosinase activity compared to untreated seedlings was after HP at 100 MPa, 30 °C, 3 min and 10 min with low degree of cell permeabilization. Copyright © 2017 Elsevier Ltd. All rights reserved.

Egg Laying of Cabbage White Butterfly (Pieris brassicae) on Arabidopsis thaliana Affects Subsequent Performance of the Larvae

PubMed Central

Blenn, Beatrice; Drechsler, Navina; Gershenzon, Jonathan; Kunze, Reinhard; Hilker, Monika

2013-01-01

Plant resistance to the feeding by herbivorous insects has recently been found to be positively or negatively influenced by prior egg deposition. Here we show how crucial it is to conduct experiments on plant responses to herbivory under conditions that simulate natural insect behaviour. We used a well-studied plant – herbivore system, Arabidopsis thaliana and the cabbage white butterfly Pieris brassicae, testing the effects of naturally laid eggs (rather than egg extracts) and allowing larvae to feed gregariously as they do naturally (rather than placing single larvae on plants). Under natural conditions, newly hatched larvae start feeding on their egg shells before they consume leaf tissue, but access to egg shells had no effect on subsequent larval performance in our experiments. However, young larvae feeding gregariously on leaves previously laden with eggs caused less feeding damage, gained less weight during the first 2 days, and suffered twice as high a mortality until pupation compared to larvae feeding on plants that had never had eggs. The concentration of the major anti-herbivore defences of A. thaliana, the glucosinolates, was not significantly increased by oviposition, but the amount of the most abundant member of this class, 4-methylsulfinylbutyl glucosinolate was 1.8-fold lower in larval-damaged leaves with prior egg deposition compared to damaged leaves that had never had eggs. There were also few significant changes in the transcript levels of glucosinolate metabolic genes, except that egg deposition suppressed the feeding-induced up-regulation of FMOGS-OX2, a gene encoding a flavin monooxygenase involved in the last step of 4-methylsulfinylbutyl glucosinolate biosynthesis. Hence, our study demonstrates that oviposition does increase A. thaliana resistance to feeding by subsequently hatching larvae, but this cannot be attributed simply to changes in glucosinolate content. PMID:23527243

Phenological and phytochemical changes correlate with differential interactions of Verticillium dahliae with broccoli and cauliflower.

PubMed

Njoroge, S M C; Vallad, G E; Park, S-Y; Kang, S; Koike, S T; Bolda, M; Burman, P; Polonik, W; Subbarao, K V

2011-05-01

Cauliflower (Brassica oleracea var. botrytis subvar. cauliflora) is susceptible to wilt caused by Verticillium dahliae but broccoli (B. oleracea var. italica subvar. cyamosa) is not. Infection of broccoli and cauliflower by a green fluorescent protein-expressing isolate of V. dahliae was examined using epifluorescence and confocal laser-scanning microscopy to follow infection and colonization in relation to plant phenology. Plant glucosinolate, phenolic, and lignin contents were also assayed at 0, 4, 14, and 28 days postinoculation. V. dahliae consistently infected and colonized the vascular tissues of all cauliflower plants regardless of age at inoculation, with the pathogen ultimately appearing in the developing seed; however, colonization decreased with plant age. In broccoli, V. dahliae infected and colonized root and stem xylem tissues of plants inoculated at 1, 2, or 3 weeks postemergence. However, V. dahliae colonized only the root xylem and the epidermal and cortical tissues of broccoli plants inoculated at 4, 5, and 6 weeks postemergence. The frequency of reisolation of V. dahliae from the stems (4 to 22%) and roots (10 to 40%) of mature broccoli plants was lower than for cauliflower stems (25 to 64%) and roots (31 to 71%). The mean level of aliphatic glucosinolates in broccoli roots was 6.18 times higher than in the shoots and did not vary with age, whereas it was 3.65 times higher in cauliflower shoots than in the roots and there was a proportional increase with age. Indole glucosinolate content was identical in both cauliflower and broccoli, and both indole and aromatic glucosinolates did not vary with plant age in either crop. Qualitative differences in characterized glucosinolates were observed between broccoli and cauliflower but no differences were observed between inoculated and noninoculated plants for either broccoli or cauliflower. However, the phenolic and lignin contents were significantly higher in broccoli following inoculation than in noninoculated broccoli or inoculated cauliflower plants. The increased resistance of broccoli to V. dahliae infection was related to the increase in phenolic and lignin contents. Significant differential accumulation of glucosinolates associated with plant phenology may also contribute to the resistant and susceptible reactions of broccoli and cauliflower, respectively, against V. dahliae.

The major glucosinolate hydrolysis product in rocket (Eruca sativa L.), sativin, is 1,3-thiazepane-2-thione: Elucidation of structure, bioactivity, and stability compared to other rocket isothiocyanates.

PubMed

Fechner, Jana; Kaufmann, Martin; Herz, Corinna; Eisenschmidt, Daniela; Lamy, Evelyn; Kroh, Lothar W; Hanschen, Franziska S

2018-09-30

Rocket is rich in glucosinolates and valued for its hot and spicy taste. Here we report the structure elucidation, bioactivity, and stability of the mainly formed glucosinolate hydrolysis product, namely sativin, which was formerly thought to be 4-mercaptobutyl isothiocyanate. However, by NMR characterization we revealed that sativin is in fact 1,3-thiazepane-2-thione, a tautomer of 4-mercaptobutyl isothiocyanate with 7-membered ring structure and so far unknown. This finding was further substantiated by conformation sampling using molecular modeling and total enthalpy calculation with density functional theory. During aqueous heat treatment sativin in general was quite stable, while the isothiocyanates erucin and sulforaphane were labile, having half-lives of 132 min and 56 min (pH 5, 100 °C), respectively. Moreover, using a WST-1 assay, we found that sativin did not reduce cell viability of HepG2 cells in a range of 0.3-30 µM, and, therefore, exhibited no cytotoxic effects in this cell line. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Effects of seed priming, salinity and methyl jasmonate treatment on bioactive composition of Brassica oleracea var. capitata (white and red varieties) sprouts.

PubMed

Hassini, Ismahen; Baenas, Nieves; Moreno, Diego A; Carvajal, Micaela; Boughanmi, Neziha; Martinez Ballesta, Maria Del Carmen

2017-06-01

Brassica spp. sprouts are rich in nutrients and bioactive compounds, especially glucosinolates and phenolic acid derivatives, and the composition of these young germinating seeds can be altered by several external factors. In this study two cabbage varieties (Brassica oleracea var. capitata, red and white) were studied using seed priming (KCl 50 mmol L -1 ; NaCl 150 mmol L -1 ) and MeJA spraying (25 µmol L -1 ) to elicit the phytochemical content of edible sprouts. The red variety was richer in glucosinolates and phenolic compounds than the white one but not in mineral nutrients. Seed priming enhanced the potassium (K) content and flavonols in both varieties, while the total content of glucosinolates was reduced after seed priming only in the red variety. The white variety responded better than the red one to KCl seed priming, increasing the flavonols (89%). Salinity did not induce any change in the phytochemical content of these two varieties. Elicitation with sprayed MeJA was effective in significantly increasing the content of indolic glucosinolates glucobrassicin (5.7-fold) and neoglucobrassicin (9.7-fold) in the red cultivar. In the white variety, in addition to glucobrassicin (19.4-fold) and neoglucobrassicin (9.4-fold), 4-hydroxyglucobrassicin (2.3-fold) was also enhanced. MeJA also elicited significant amounts of anthocyanins (41%) and chlorogenic acid derivatives (329%) in the white variety. KCl seed priming and MeJA elicitation promoted the phytochemical composition of the cabbage varieties, especially in the white variety. The application of NaCl resulted in less efficient elicitation. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

An Intermediate in the Synthesis of Glucobrassicins from 3-Indoleacetaldoxime by Woad Leaves 1

PubMed Central

Mahadevan, S.; Stowe, Bruce B.

1972-01-01

Leaves of woad (Isatis tinctoria L.) were found to incorporate efficiently tritiated indoleacetaldoxime and 35S from 35S-l-cystine into glucobrassicin and sulfoglucobrassicin. Time course of incorporation of 35S from 35S-cystine into the glucosinolates indicated that glucobrassicin was formed first and then sulfoglucobrassicin. Simultaneous administration of tritiated indoleacetaldoxime and 35S-cystine gave doubly labeled glucobrassicin and sulfoglucobrassicin. About twice as much 35S was present in sulfoglucobrassicin as compared to glucobrassicin per unit of 3H incorporated, indicating that a second, probably oxidized, atom of 35S was later introduced into sulfoglucobrassicin. However, the 35S incorporated from cystine into both glucosinolates during the first 8 hours of metabolism was almost exclusively in the divalent sulfur moiety. The incorporation patterns of 35S and titritated indoleacetaldoxime into the glucosinolates suggested a fast turnover of glucobrassicin in the metabolizing leaves. A new indolic, sulfur-containing neutral compound X was found to accumulate in woad leaves when administered 3H-3-indoleacetaldoxime and cold cystine or 35S-cystine and cold 3-indoleacetaldoxime. This accumulation was enhanced about 2- to 2.5-fold by the simultaneous administration of postassium selenate, an inhibitor of biological sulfation processes. Selenate also appeared to inhibit the conversion of glucobrassicin to 1-sulfoglucobrassicin. Partially purified compound X was efficiently converted (56-60%) to glucobrassicin and 1-sulfoglucobrassicin on readministration to woad leaves, indicating it to be a precursor of the glucosinolates. Compound X, on treatment with myrosinase, slowly yielded a less polar, indolic, sulfur containing compound Y and glucose. Compound Y decomposed with time into indoleacetonitrile suggesting that it may be indoleacetothiohydroximate. Compound X has been tentatively assigned the structure of desthioglucobrassicin, the nonsulfated form of glucobrassicin. PMID:16658130

Effect of NaCl treatments on glucosinolate metabolism in broccoli sprouts*

PubMed Central

Guo, Rong-fang; Yuan, Gao-feng; Wang, Qiao-mei

2013-01-01

To understand the regulation mechanism of NaCl on glucosinolate metabolism in broccoli sprouts, the germination rate, fresh weight, contents of glucosinolates and sulforaphane, as well as myrosinase activity of broccoli sprouts germinated under 0, 20, 40, 60, 80, and 100 mmol/L of NaCl were investigated in our experiment. The results showed that glucoerucin, glucobrassicin, and 4-hydroxy glucobrassicin in 7-d-old broccoli sprouts were significantly enhanced and the activity of myrosinase was inhibited by 100 mmol/L of NaCl. However, the total glucosinolate content in 7-d-old broccoli sprouts was markedly decreased although the fresh weight was significantly increased after treatment with NaCl at relatively low concentrations (20, 40, and 60 mmol/L). NaCl treatment at the concentration of 60 mmol/L for 5 d maintained higher biomass and comparatively higher content of glucosinolates in sprouts of broccoli with decreased myrosinase activity. A relatively high level of NaCl treatment (100 mmol/L) significantly increased the content of sulforaphane in 7-d-old broccoli sprouts compared with the control. These results indicate that broccoli sprouts grown under a suitable concentration of NaCl could be desirable for human nutrition. PMID:23365011

Herbicidal Activity of Glucosinolate Degradation Products in Fermented Meadowfoam (Limnanthes alba) Seed Meal

PubMed Central

STEVENS, JAN F.; REED, RALPH L.; ALBER, SUSAN; PRITCHETT, LARRY; MACHADO, STEPHEN

2009-01-01

Meadowfoam (Limnanthes alba) is an oilseed crop grown in western Oregon. After extraction of the oil from the seeds, the remaining seed meal contains 2-4% of the glucosinolate, glucolimnanthin. We investigated the effect of fermentation of seed meal on its chemical composition and the effect of the altered composition on downy brome (Bromus tectorum) coleoptile emergence. Incubation of enzyme-inactive seed meal with enzyme-active seeds (1% by weight) resulted in complete degradation of glucolimnanthin and formation of 3-methoxybenzyl isothiocyanate in 28% yield. Fermentation in the presence of an aqueous solution of FeSO4 (10 mM) resulted in the formation of 3-methoxyphenylacetonitrile and 2-(3-methoxyphenyl)ethanethioamide, a novel natural product. The formation of the isothiocyanate, the nitrile and the thioamide, as a total, correlated with an increase of herbicidal potency of seed meal (r2 = 0.96). The results of this study open new possibilities for the refinement of glucosinolate-containing seed meals for use as bioherbicides. PMID:19170637

Herbicidal activity of glucosinolate degradation products in fermented meadowfoam ( Limnanthes alba ) seed meal.

PubMed

Stevens, Jan F; Reed, Ralph L; Alber, Susan; Pritchett, Larry; Machado, Stephen

2009-03-11

Meadowfoam ( Limnanthes alba ) is an oilseed crop grown in western Oregon. After extraction of the oil from the seeds, the remaining seed meal contains 2-4% of the glucosinolate glucolimnanthin. This study investigated the effect of fermentation of seed meal on its chemical composition and the effect of the altered composition on downy brome ( Bromus tectorum ) coleoptile emergence. Incubation of enzyme-inactive seed meal with enzyme-active seeds (1% by weight) resulted in complete degradation of glucolimnanthin and formation of 3-methoxybenzyl isothiocyanate in 28% yield. Fermentation in the presence of an aqueous solution of FeSO(4) (10 mM) resulted in the formation of 3-methoxyphenylacetonitrile and 2-(3-methoxyphenyl)ethanethioamide, a novel natural product. The formation of the isothiocyanate, the nitrile, and the thioamide, as a total, correlated with an increase of herbicidal potency of the seed meal (r(2) = 0.96). The results of this study open new possibilities for the refinement of glucosinolate-containing seed meals for use as bioherbicides.

Distribution and Variation of Indole Glucosinolates in Woad (Isatis tinctoria L.) 1

PubMed Central

Elliott, Malcolm C.; Stowe, Bruce B.

1971-01-01

The exceptionally high levels in woad (Isatis tinctoria L.) of three indolic goitrogens, namely glucobrassicin, neoglucobrassicin, and glucobrassicin-1-sulfonate, permit the facile study of their distribution in the plant and their changes during its development. Woad seeds contain as much as 0.23% fresh weight of glucobrassicin but no other indole glucosinolate, while 1-week-old seedlings also contain substantial amounts of neoglucobrassicin and glucobrassicin-1-sulfonate in their shoots whether grown in the light or dark. The sulfonate is not found in roots, and light depresses neoglucobrassicin levels in shoots. Sterile root cultures synthesize glucobrassicin and neoglucobrassicin, and significant quantities of these were even found to be excreted by the roots of intact sterile seedlings in culture. This may explain the long known deleterious effect of woad and other cruciferous crops on subsequent plantings and the observation could be of ecological importance. Long term changes in levels of all three substances in the plant are similar and are compatible with earlier suggestions that the compounds could be auxin precursors at the time of flower stem elongation. Since sterile seedlings readily incorporate 35SO42− into indole glucosinolates and relative specific radioactivities suggest that glucobrassicin is the precursor of the other two compounds, pathways of goitrogen biosynthesis should be relatively easily determined in this material. PMID:16657825

Effects of MeJA on Arabidopsis metabolome under endogenous JA deficiency

NASA Astrophysics Data System (ADS)

Cao, Jingjing; Li, Mengya; Chen, Jian; Liu, Pei; Li, Zhen

2016-11-01

Jasmonates (JAs) play important roles in plant growth, development and defense. Comprehensive metabolomics profiling of plants under JA treatment provides insights into the interaction and regulation network of plant hormones. Here we applied high resolution mass spectrometry based metabolomics approach on Arabidopsis wild type and JA synthesis deficiency mutant opr3. The effects of exogenous MeJA treatment on the metabolites of opr3 were investigated. More than 10000 ion signals were detected and more than 2000 signals showed significant variation in different genotypes and treatment groups. Multivariate statistic analyses (PCA and PLS-DA) were performed and a differential compound library containing 174 metabolites with high resolution precursor ion-product ions pairs was obtained. Classification and pathway analysis of 109 identified compounds in this library showed that glucosinolates and tryptophan metabolism, amino acids and small peptides metabolism, lipid metabolism, especially fatty acyls metabolism, were impacted by endogenous JA deficiency and exogenous MeJA treatment. These results were further verified by quantitative reverse transcription PCR (RT-qPCR) analysis of 21 related genes involved in the metabolism of glucosinolates, tryptophan and α-linolenic acid pathways. The results would greatly enhance our understanding of the biological functions of JA.

Molecular characterization of BrMYB28 and BrMYB29 paralogous transcription factors involved in the regulation of aliphatic glucosinolate profiles in Brassica rapa ssp. pekinensis.

PubMed

Baskar, Venkidasamy; Park, Se Won

2015-07-01

Glucosinolates (GSL) are one of the major secondary metabolites of the Brassicaceae family. In the present study, we aim at characterizing the multiple paralogs of aliphatic GSL regulators, such as BrMYB28 and BrMYB29 genes in Brassica rapa ssp. pekinensis, by quantitative real-time PCR (qRT-PCR) analysis in different tissues and at various developmental stages. An overlapping gene expression pattern between the BrMYBs as well as their downstream genes (DSGs) was found at different developmental stages. Among the BrMYB28 and BrMYB29 paralogous genes, the BrMYB28.3 and BrMYB29.1 genes were dominantly expressed in most of the developmental stages, compared to the other paralogs of the BrMYB genes. Furthermore, the differential expression pattern of the BrMYBs was observed under various stress treatments. Interestingly, BrMYB28.2 showed the least expression in most developmental stages, while its expression was remarkably high in different stress conditions. More specifically, the BrMYB28.2, BrMYB28.3, and BrMYB29.1 genes were highly responsive to various abiotic and biotic stresses, further indicating their possible role in stress tolerance. Moreover, the in silico cis motif analysis in the upstream regulatory regions of BrMYBs showed the presence of various putative stress-specific motifs, which further indicated their responsiveness to biotic and abiotic stresses. These observations suggest that the dominantly expressed BrMYBs, both in different developmental stages and under various stress treatments (BrMYB28.3 and BrMYB29.1), may be potential candidate genes for altering the GSL level through genetic modification studies in B. rapa ssp. pekinensis. Copyright © 2015. Published by Elsevier SAS.

Reduced Arogenate Dehydratase Expression: Ramifications for Photosynthesis and Metabolism1[OPEN

PubMed Central

Höhner, Ricarda; Ito, Tetsuro; Amakura, Yoshiaki; Weitz, Karl

2018-01-01

Arogenate dehydratase (ADT) catalyzes the final step of phenylalanine (Phe) biosynthesis. Previous work showed that ADT-deficient Arabidopsis (Arabidopsis thaliana) mutants had significantly reduced lignin contents, with stronger reductions in lines that had deficiencies in more ADT isoforms. Here, by analyzing Arabidopsis ADT mutants using our phenomics facility and ultra-performance liquid chromatography-mass spectrometry-based metabolomics, we describe the effects of the modulation of ADT on photosynthetic parameters and secondary metabolism. Our data indicate that a reduced carbon flux into Phe biosynthesis in ADT mutants impairs the consumption of photosynthetically produced ATP, leading to an increased ATP/ADP ratio, the overaccumulation of transitory starch, and lower electron transport rates. The effect on electron transport rates is caused by an increase in proton motive force across the thylakoid membrane that down-regulates photosystem II activity by the high-energy quenching mechanism. Furthermore, quantitation of secondary metabolites in ADT mutants revealed reduced flavonoid, phenylpropanoid, lignan, and glucosinolate contents, including glucosinolates that are not derived from aromatic amino acids, and significantly increased contents of putative galactolipids and apocarotenoids. Additionally, we used real-time atmospheric monitoring mass spectrometry to compare respiration and carbon fixation rates between the wild type and adt3/4/5/6, our most extreme ADT knockout mutant, which revealed no significant difference in both night- and day-adapted plants. Overall, these data reveal the profound effects of altered ADT activity and Phe metabolism on secondary metabolites and photosynthesis with implications for plant improvement. PMID:29523714

Effects of industrial pre-freezing processing and freezing handling on glucosinolates and antioxidant attributes in broccoli florets.

PubMed

Cai, Congxi; Miao, Huiying; Qian, Hongmei; Yao, Leishuan; Wang, Bingliang; Wang, Qiaomei

2016-11-01

The effects of industrial pre-freezing processing and freezing handling on the contents of glucosinolates and antioxidants (vitamin C, polyphenols, carotenoid and chlorophyll), as well as the antioxidant capacity in broccoli (Brassica oleracea L. var. italica) florets were investigated in the present study. Our results showed that the glucosinolate accumulations were significantly decreased after pre-freezing processing, whereas elevated levels of phenols, carotenoids, chlorophyll, and also antioxidant capacity were observed in frozen broccoli florets. The contents of vitamin C remained constant during above mentioned processing. In conclusion, the current industrial freezing processing method is a good practice for the preservation of main antioxidant nutrients in broccoli florets, although some improvements in pre-freezing processing, such as steam blanching and ice-water cooling, are needed to attenuate the decrease in glucosinolate content. Copyright © 2016 Elsevier Ltd. All rights reserved.

Optimized analysis and quantification of glucosinolates from Camelina sativa seeds by reverse-phase liquid chromatography

USDA-ARS?s Scientific Manuscript database

Gold-of-pleasure or false flax (Camelina sativa L. Crantz) presscake contains three relatively unique glucosinolates: glucoarabin (9-(methylsulfinyl) nonylglucosinolate)glucocamelinin (10-(methylsulfinyl)decylglucosinolate), and 11-(methylsulfinyl)undecylglucosinolate. Using defatted seed material...

Concentrations of thiocyanate and goitrin in human plasma, their precursor concentrations in brassica vegetables, and associated potential risk for hypothyroidism

PubMed Central

Bunch, Ronald; Leung, Angela M.

2016-01-01

Brassica vegetables are common components of the diet and have beneficial as well as potentially adverse health effects. Following enzymatic breakdown, some glucosinolates in brassica vegetables produce sulforaphane, phenethyl, and indolylic isothiocyanates that possess anticarcinogenic activity. In contrast, progoitrin and indolylic glucosinolates degrade to goitrin and thiocyanate, respectively, and may decrease thyroid hormone production. Radioiodine uptake to the thyroid is inhibited by 194 μmol of goitrin, but not by 77 μmol of goitrin. Collards, Brussels sprouts, and some Russian kale (Brassica napus) contain sufficient goitrin to potentially decrease iodine uptake by the thyroid. However, turnip tops, commercial broccoli, broccoli rabe, and kale belonging to Brassica oleracae contain less than 10 μmol of goitrin per 100-g serving and can be considered of minimal risk. Using sulforaphane plasma levels following glucoraphanin ingestion as a surrogate for thiocyanate plasma concentrations after indole glucosinolate ingestion, the maximum thiocyanate contribution from indole glucosinolate degradation is estimated to be 10 μM, which is significantly lower than background plasma thiocyanate concentrations (40–69 μM). Thiocyanate generated from consumption of indole glucosinolate can be assumed to have minimal adverse risks for thyroid health. PMID:26946249

Distribution and Variation of Indole Glucosinolates in Woad (Isatis tinctoria L.).

PubMed

Elliott, M C; Stowe, B B

1971-10-01

The exceptionally high levels in woad (Isatis tinctoria L.) of three indolic goitrogens, namely glucobrassicin, neoglucobrassicin, and glucobrassicin-1-sulfonate, permit the facile study of their distribution in the plant and their changes during its development. Woad seeds contain as much as 0.23% fresh weight of glucobrassicin but no other indole glucosinolate, while 1-week-old seedlings also contain substantial amounts of neoglucobrassicin and glucobrassicin-1-sulfonate in their shoots whether grown in the light or dark. The sulfonate is not found in roots, and light depresses neoglucobrassicin levels in shoots. Sterile root cultures synthesize glucobrassicin and neoglucobrassicin, and significant quantities of these were even found to be excreted by the roots of intact sterile seedlings in culture. This may explain the long known deleterious effect of woad and other cruciferous crops on subsequent plantings and the observation could be of ecological importance. Long term changes in levels of all three substances in the plant are similar and are compatible with earlier suggestions that the compounds could be auxin precursors at the time of flower stem elongation. Since sterile seedlings readily incorporate (35)SO(4) (2-) into indole glucosinolates and relative specific radioactivities suggest that glucobrassicin is the precursor of the other two compounds, pathways of goitrogen biosynthesis should be relatively easily determined in this material.

Functional identification of genes responsible for the biosynthesis of 1-methoxy-indol-3-ylmethyl-glucosinolate in Brassica rapa ssp. chinensis

PubMed Central

2014-01-01

Background Brassica vegetables contain a class of secondary metabolites, the glucosinolates (GS), whose specific degradation products determine the characteristic flavor and smell. While some of the respective degradation products of particular GS are recognized as health promoting substances for humans, recent studies also show evidence that namely the 1-methoxy-indol-3-ylmethyl GS might be deleterious by forming characteristic DNA adducts. Therefore, a deeper knowledge of aspects involved in the biosynthesis of indole GS is crucial to design vegetables with an improved secondary metabolite profile. Results Initially the leafy Brassica vegetable pak choi (Brassica rapa ssp. chinensis) was established as suitable tool to elicit very high concentrations of 1-methoxy-indol-3-ylmethyl GS by application of methyl jasmonate. Differentially expressed candidate genes were discovered in a comparative microarray analysis using the 2 × 104 K format Brassica Array and compared to available gene expression data from the Arabidopsis AtGenExpress effort. Arabidopsis knock out mutants of the respective candidate gene homologs were subjected to a comprehensive examination of their GS profiles and confirmed the exclusive involvement of polypeptide 4 of the cytochrome P450 monooxygenase subfamily CYP81F in 1-methoxy-indol-3-ylmethyl GS biosynthesis. Functional characterization of the two identified isoforms coding for CYP81F4 in the Brassica rapa genome was performed using expression analysis and heterologous complementation of the respective Arabidopsis mutant. Conclusions Specific differences discovered in a comparative microarray and glucosinolate profiling analysis enables the functional attribution of Brassica rapa ssp. chinensis genes coding for polypeptide 4 of the cytochrome P450 monooxygenase subfamily CYP81F to their metabolic role in indole glucosinolate biosynthesis. These new identified Brassica genes will enable the development of genetic tools for breeding vegetables with improved GS composition in the near future. PMID:24886080

Allelopathic effects of glucosinolate breakdown products in Hanza [Boscia senegalensis (Pers.) Lam.] processing waste water

PubMed Central

Rivera-Vega, Loren J.; Krosse, Sebastian; de Graaf, Rob M.; Garvi, Josef; Garvi-Bode, Renate D.; van Dam, Nicole M.

2015-01-01

Boscia senegalensis is a drought resistant shrub whose seeds are used in West Africa as food. However, the seeds, or hanza, taste bitter which can be cured by soaking them in water for 4–7 days. The waste water resulting from the processing takes up the bitter taste, which makes it unsuitable for consumption. When used for irrigation, allelopathic effects were observed. Glucosinolates and their breakdown products are the potential causes for both the bitter taste and the allelopathic effects. The objectives of this study are to identify and quantify the glucosinolates present in processed and unprocessed hanza as well as different organs of B. senegalensis, to analyze the chemical composition of the processing water, and to pinpoint the causal agent for the allelopathic properties of the waste water. Hanza (seeds without testa), leaves, branches, unripe, and ripe fruits were collected in three populations and subjected to glucosinolate analyses. Methylglucosinolates (MeGSL) were identified in all plant parts and populations, with the highest concentrations being found in the hanza. The levels of MeGSLs in the hanza reduced significantly during the soaking process. Waste water was collected for 6 days and contained large amounts of macro- and micronutrients, MeGSL as well as methylisothiocyanate (MeITC), resulting from the conversion of glucosinolates. Waste water from days 1–3 (High) and 4–6 (Low) was pooled and used to water seeds from 11 different crops to weeds. The High treatment significantly delayed or reduced germination of all the plant species tested. Using similar levels of MeITC as detected in the waste water, we found that germination of a subset of the plant species was inhibited equally to the waste water treatments. This confirmed that the levels of methylisiothiocyanate in the waste water were sufficient to cause the allelopathic effect. This leads to the possibility of using hanza waste water in weed control programs. PMID:26236325

An intermediate in the synthesis of glucobrassicins from 3-indoleacetaldoxime by woad leaves.

PubMed

Mahadevan, S; Stowe, B B

1972-07-01

Leaves of woad (Isatis tinctoria L.) were found to incorporate efficiently tritiated indoleacetaldoxime and (35)S from (35)S-l-cystine into glucobrassicin and sulfoglucobrassicin. Time course of incorporation of (35)S from (35)S-cystine into the glucosinolates indicated that glucobrassicin was formed first and then sulfoglucobrassicin. Simultaneous administration of tritiated indoleacetaldoxime and (35)S-cystine gave doubly labeled glucobrassicin and sulfoglucobrassicin. About twice as much (35)S was present in sulfoglucobrassicin as compared to glucobrassicin per unit of (3)H incorporated, indicating that a second, probably oxidized, atom of (35)S was later introduced into sulfoglucobrassicin. However, the (35)S incorporated from cystine into both glucosinolates during the first 8 hours of metabolism was almost exclusively in the divalent sulfur moiety. The incorporation patterns of (35)S and titritated indoleacetaldoxime into the glucosinolates suggested a fast turnover of glucobrassicin in the metabolizing leaves.A new indolic, sulfur-containing neutral compound X was found to accumulate in woad leaves when administered (3)H-3-indoleacetaldoxime and cold cystine or (35)S-cystine and cold 3-indoleacetaldoxime. This accumulation was enhanced about 2- to 2.5-fold by the simultaneous administration of postassium selenate, an inhibitor of biological sulfation processes. Selenate also appeared to inhibit the conversion of glucobrassicin to 1-sulfoglucobrassicin. Partially purified compound X was efficiently converted (56-60%) to glucobrassicin and 1-sulfoglucobrassicin on readministration to woad leaves, indicating it to be a precursor of the glucosinolates. Compound X, on treatment with myrosinase, slowly yielded a less polar, indolic, sulfur containing compound Y and glucose. Compound Y decomposed with time into indoleacetonitrile suggesting that it may be indoleacetothiohydroximate. Compound X has been tentatively assigned the structure of desthioglucobrassicin, the nonsulfated form of glucobrassicin.

Glucosinolate Profiles in Cabbage Genotypes Influence the Preferential Feeding of Diamondback Moth (Plutella xylostella)

PubMed Central

Robin, Arif Hasan Khan; Hossain, Mohammad Rashed; Park, Jong-In; Kim, Hye R.; Nou, Ill-Sup

2017-01-01

Diamondback moth (DBM), Plutella xylostella L., is a devastating pest of cabbage worldwide whose feeding attributes are influenced by glucosinolate profiles of the plant. Identifying the specific glucosinolates associated with plants’ resistance mechanism can provide cues to novel points of intervention in developing resistant cultivars. We studied the DBM larval feeding preference and extent of damage on cabbage leaves via controlled glass-house and in vitro multiple- and two-choice feeding tests. These feeding attributes were associated with the individual glucosinolate profiles, analyzed by HPLC, of each of the eight cabbage genotypes using multivariate analytical approach to identify the glucosinolates that may have roles in resistance. Both the glass-house and in vitro multiple-choice feeding tests identified the genotype BN4303, BN4059, and BN4072 as the least preferred (resistant) and Rubra, YR Gold and BN3383 as most preferred (susceptible) genotypes by DBM larvae. The principal component analysis separated the genotypes based on lower feeding scores in association with higher contents of glucobrassicin, glucoiberin, glucoiberverin in one direction and 4-hydroxyglucobrassicin, glucoerucin, glucoraphanin, and progoitrin in opposite direction in a way to explain the major variation in resistant versus susceptible genotypes based on their extent of preference and leaf area damage. The simultaneous presence (or higher contents) of glucobrassicin, glucoiberin, and glucoiberverin and the absence (or lower contents) of 4-hydroxyglucobrassicin, glucoerucin, glucoraphanin, and progoitrin in the least preferred genotypes and vice-versa in most preferred genotypes indicated their apparent role as putative repellents and attractants of DBM larvae in cabbage genotypes, respectively. These novel findings add to the current knowledgebase on the roles of glucosinolates in plant–herbivore interactions and will be helpful in setting breeding priorities for improving the resistance against DBM in cabbage using conventional and biotechnological approaches. PMID:28769953

Enhancement of broccoli indole glucosinolates by methyl jasmonate treatment and effects on prostate carcinogenesis.

PubMed

Liu, Ann G; Juvik, John A; Jeffery, Elizabeth H; Berman-Booty, Lisa D; Clinton, Steven K; Erdman, John W

2014-11-01

Broccoli is rich in bioactive components, such as sulforaphane and indole-3-carbinol, which may impact cancer risk. The glucosinolate profile of broccoli can be manipulated through treatment with the plant stress hormone methyl jasmonate (MeJA). Our objective was to produce broccoli with enhanced levels of indole glucosinolates and determine its impact on prostate carcinogenesis. Brassica oleracea var. Green Magic was treated with a 250 μM MeJA solution 4 days prior to harvest. MeJA-treated broccoli had significantly increased levels of glucobrassicin, neoglucobrassicin, and gluconasturtiin (P < .05). Male transgenic adenocarcinoma of mouse prostate (TRAMP) mice (n = 99) were randomized into three diet groups at 5-7 weeks of age: AIN-93G control, 10% standard broccoli powder, or 10% MeJA broccoli powder. Diets were fed throughout the study until termination at 20 weeks of age. Hepatic CYP1A was induced with MeJA broccoli powder feeding, indicating biological activity of the indole glucosinolates. Following ∼ 15 weeks on diets, neither of the broccoli treatments significantly altered genitourinary tract weight, pathologic score, or metastasis incidence, indicating that broccoli powder at 10% of the diet was ineffective at reducing prostate carcinogenesis in the TRAMP model. Whereas broccoli powder feeding had no effect in this model of prostate cancer, our work demonstrates the feasibility of employing plant stress hormones exogenously to stimulate changes in phytochemical profiles, an approach that may be useful for optimizing bioactive component patterns in foods for chronic-disease-prevention studies.

Near-infrared reflectance spectroscopy calibrations for assessment of oil, phenols, glucosinolates and fatty acid content in the intact seeds of oilseed Brassica species.

PubMed

Sen, Rahul; Sharma, Sanjula; Kaur, Gurpreet; Banga, Surinder S

2018-01-31

Very few near-infrared reflectance spectroscopy (NIRS) calibration models are available for non-destructive estimation of seed quality traits in Brassica juncea. Those that are available also fail to adequately discern variation for oleic acid (C 18:1 ) , linolenic (C 18:3 ) fatty acids, meal glucosinolates and phenols. We report the development of a new NIRS calibration equation that is expected to fill the gaps in the existing NIRS equations. Calibrations were based on the reference values of important quality traits estimated from a purposely selected germplasm set comprising 240 genotypes of B. juncea and 193 of B. napus. We were able to develop optimal NIRS-based calibration models for oil, phenols, glucosinolates, oleic acid, linoleic acid and erucic acid for B. juncea and B. napus. Correlation coefficients (RSQ) of the external validations appeared greater than 0.7 for the majority of traits, such as oil (0.766, 0.865), phenols (0.821, 0.915), glucosinolates (0.951, 0.986), oleic acid (0.814. 0.810), linoleic acid (0.974, 0.781) and erucic acid (0.963, 0.943) for B. juncea and B. napus, respectively. The results demonstrate the robust predictive power of the developed calibration models for rapid estimation of many quality traits in intact rapeseed-mustard seeds which will assist plant breeders in effective screening and selection of lines in quality improvement breeding programmes. © 2018 Society of Chemical Industry. © 2018 Society of Chemical Industry.

Effects of photoperiod, growth temperature and cold acclimatisation on glucosinolates, sugars and fatty acids in kale.

PubMed

Steindal, Anne Linn Hykkerud; Rødven, Rolf; Hansen, Espen; Mølmann, Jørgen

2015-05-01

Curly kale is a robust, cold tolerant plant with a high content of health-promoting compounds, grown at a range of latitudes. To assess the effects of temperature, photoperiod and cold acclimatisation on levels of glucosinolates, fatty acids and soluble sugars in kale, an experiment was set up under controlled conditions. Treatments consisted of combinations of the temperatures 15/9 or 21/15 °C, and photoperiods of 12 or 24h, followed by a cold acclimatisation period. Levels of glucosinolates and fatty acid types in leaves were affected by growth conditions and cold acclimatisation, being generally highest before acclimatisation. The effects of growth temperature and photoperiod on freezing tolerance were most pronounced in plants grown without cold acclimatisation. The results indicate that cold acclimatisation can increase the content of soluble sugar and can thereby improve the taste, whilst the content of unsaturated fatty and glucosinolates acids may decrease. Copyright © 2014 Elsevier Ltd. All rights reserved.

Variation of glucosinolates and quinone reductase activity among different varieties of Chinese kale and improvement of glucoraphanin by metabolic engineering.

PubMed

Qian, Hongmei; Sun, Bo; Miao, Huiying; Cai, Congxi; Xu, Chaojiong; Wang, Qiaomei

2015-02-01

The variation of glucosinolates and quinone reductase (QR) activity in fourteen varieties of Chinese kale (Brassica oleracea var. alboglabra Bailey) was investigated in the present study. Results showed that gluconapin (GNA), instead of glucoraphanin (GRA), was the most predominant glucosinolate in all varieties, and QR activity was remarkably positively correlated with the glucoraphanin level. AOP2, a tandem 2-oxoglutarate-dependent dioxygenase, catalyzes the conversion of glucoraphanin to gluconapin in glucosinolate biosynthesis. Here, antisense AOP2 was transformed into Gailan-04, the variety with the highest gluconapin content and ratio of GNA/GRA. The glucoraphanin content and corresponding QR activity were notably increased in transgenic plants, while no significant difference at the level of other main nutritional compounds (total phenolics, vitamin C, carotenoids and chlorophyll) was observed between the transgenic lines and the wide-type plants. Taken together, metabolic engineering is a good practice for improvement of glucoraphanin in Chinese kale. Copyright © 2014 Elsevier Ltd. All rights reserved.

A young root-specific gene (ArMY2) from horseradish encoding a MYR II myrosinase with kinetic preference for the root-specific glucosinolate gluconasturtiin.

PubMed

Loebers, Andreas; Müller-Uri, Frieder; Kreis, Wolfgang

2014-03-01

The pungent taste of horseradish is caused by isothiocyanates which are released from glucosinolates by myrosinases. These enzymes are encoded by genes belonging to one of two subfamilies, termed MYR I and MYR II, respectively. A MYR II-type myrosinase gene was identified for the first time in horseradish. The gene termed ArMY2 was only expressed in young roots. A full-length cDNA encoding a myrosinase termed ArMy2 was isolated and heterologously expressed in Pichia pastoris. The recombinant His-tagged enzyme was characterized biochemically. Substrate affinity was 5 times higher towards gluconasturtiin than towards sinigrin. Gluconasturtiin was found to be the most abundant glucosinolate in young horseradish roots while sinigrin dominated in storage roots and leaves. This indicates that a specialized glucosinolate-myrosinase defense system might be active in young roots. Copyright © 2013 Elsevier Ltd. All rights reserved.

Dietary Glucosinolates Sulforaphane, Phenethyl Isothiocyanate, Indole-3-Carbinol/3,3'-Diindolylmethane: Anti-Oxidative Stress/Inflammation, Nrf2, Epigenetics/Epigenomics and In Vivo Cancer Chemopreventive Efficacy.

PubMed

Fuentes, Francisco; Paredes-Gonzalez, Ximena; Kong, Ah-Ng Tony

2015-05-01

Glucosinolates are a group of sulfur-containing glycosides found in many plant species, including cruciferous vegetables such as broccoli, cabbage, brussels sprouts, and cauliflower. Accumulating evidence increasingly supports the beneficial effects of dietary glucosinolates on overall health, including as potential anti-cancer agents, because of their role in the prevention of the initiation of carcinogenesis via the induction of cellular defense detoxifying/antioxidant enzymes and their epigenetic mechanisms, including modification of the CpG methylation of cancer-related genes, histone modification regulation and changes in the expression of miRNAs. In this context, the defense mechanism mediated by Nrf2-antioxidative stress and anti-inflammatory signaling pathways can contribute to cellular protection against oxidative stress and reactive metabolites of carcinogens. In this review, we summarize the cancer chemopreventive role of naturally occurring glucosinolate derivatives as inhibitors of carcinogenesis, with particular emphasis on specific molecular targets and epigenetic alterations in in vitro and in vivo human cancer animal models.

Proposed Method for Estimating Health-Promoting Glucosinolates and Hydrolysis Products in Broccoli (Brassica oleracea var. italica) Using Relative Transcript Abundance.

PubMed

Becker, Talon M; Jeffery, Elizabeth H; Juvik, John A

2017-01-18

Due to the importance of glucosinolates and their hydrolysis products in human nutrition and plant defense, optimizing the content of these compounds is a frequent breeding objective for Brassica crops. Toward this goal, we investigated the feasibility of using models built from relative transcript abundance data for the prediction of glucosinolate and hydrolysis product concentrations in broccoli. We report that predictive models explaining at least 50% of the variation for a number of glucosinolates and their hydrolysis products can be built for prediction within the same season, but prediction accuracy decreased when using models built from one season's data for prediction of an opposing season. This method of phytochemical profile prediction could potentially allow for lower phytochemical phenotyping costs and larger breeding populations. This, in turn, could improve selection efficiency for phase II induction potential, a type of chemopreventive bioactivity, by allowing for the quick and relatively cheap content estimation of phytochemicals known to influence the trait.

Characterization of phenolics, glucosinolates and antioxidant activity of beverages based on apple juice with addition of frozen and freeze-dried curly kale leaves (Brassica oleracea L. var. acephala L.).

PubMed

Biegańska-Marecik, Róża; Radziejewska-Kubzdela, Elżbieta; Marecik, Roman

2017-09-01

The aim of this study was to determine the polyphenols, glucosinolates and ascorbic acid content as well as antioxidant activity of beverages on the base of apple juice with addition of frozen and freeze-dried curly kale leaves. Upon enrichment with frozen (13%) and freeze-dried curly kale (3%), the naturally cloudy apple juice was characterized by an increase in phenolic compounds by 2.7 and 3.3-times, accordingly. The antioxidant activity of beverages with the addition of curly kale ranged from 6.6 to 9.4μmol Trolox/mL. The obtained beverages were characterized glucosinolates content at 117.6-167.6mg/L and ascorbic acid content at 4,1-31,9mg/L. The results of sensory evaluation of colour, taste and consistency of apple juice and beverages with the addition of kale did not differ significantly prior to pasteurization (P≤0.05), whereas after the pasteurization the evaluated factors decreased significantly. Copyright © 2017 Elsevier Ltd. All rights reserved.

The mechanism of deterioration of the glucosinolate-myrosynase system in radish roots during cold storage after harvest.

PubMed

Lee, Jeong Gu; Lim, Sooyeon; Kim, Jongkee; Lee, Eun Jin

2017-10-15

The hydrolysis of glucosinolates (GSLs) by myrosinase yields varieties of degradation products including isothiocyanates (ITCs). This process is controlled by the glucosinolate-myrosinase (G-M) system. The major ITCs in radish roots are raphasatin and sulforaphene (SFE), and the levels of these compounds decrease during storage after harvest. We investigated the G-M system to understand the mechanism behind the decrease in the ITCs in radish roots. Six varieties of radish roots were stored for 8weeks at 0-1.5°C. The concentrations of GSLs (glucoraphasatin and glucoraphenin) were maintained at harvest levels without significant changes during the storage period. However, SFE concentration and myrosinase activity remarkably decreased for 8weeks. Pearson correlation analysis between ITCs, GSLs, and myrosinase activity showed that a decrease of SFE during storage had a positive correlation with a decrease in myrosinase activity, which resulted from a decrease of ascorbic acid but also a decrease of myrosinase activity-related gene expressions. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Myrosinase-dependent and –independent formation and control of isothiocyanate products of glucosinolate hydrolysis

USDA-ARS?s Scientific Manuscript database

Brassicales contain a myrosinase enzyme that hydrolyzes glucosinolates to form toxic isothiocyanates, as a defense against bacteria, fungi, insects and herbivores including man. Low levels of isothiocyanates trigger a host defense system in mammals that protects them against chronic diseases. Becaus...

Fighting Cancer with Good Taste (and a Good Diet).

ERIC Educational Resources Information Center

Hoffmann, Franz

1997-01-01

Highlights glucosinolates-a family of secondary metabolites from plants of high culinary and nutritional value. Describes experiments that demonstrate the enzymatic degradation of glucosinolates. Teaches students about the distribution, variation, chemistry, and function of a family of natural products important in plant protection, food spicing,…

Challenges of developing a valid Dietary Glucosinolate database

USDA-ARS?s Scientific Manuscript database

Glucosinolates are a group of important sulfur-containing compounds found in cruciferous vegetables. They have been suggested to be the major cancer chemo-preventive agents in these vegetables. To estimate the dietary intake of this group of compounds and evaluate their health impact, there is a g...

Camelina sativa defatted seed meal contains both alkyl sulfinyl glucosinolates and quercetin that synergize bioactivity

USDA-ARS?s Scientific Manuscript database

Camelina sativa L. Crantz is under development as a novel oil-seed crop, yet bioefficacy of camelina phytochemicals is unknown. Defatted camelina seed meal contains two major aliphatic glucosinolates (GSL), glucoarabin (9-(methylsulfinyl)nonylglucosinolate; GSL 9) and glucocamelinin (10-(methylsulfi...

How specialized volatiles respond to chronic and short-term physiological and shock heat stress in Brassica nigra.

PubMed

Kask, Kaia; Kännaste, Astrid; Talts, Eero; Copolovici, Lucian; Niinemets, Ülo

2016-09-01

Brassicales release volatile glucosinolate breakdown products upon tissue mechanical damage, but it is unclear how the release of glucosinolate volatiles responds to abiotic stresses such as heat stress. We used three different heat treatments, simulating different dynamic temperature conditions in the field to gain insight into stress-dependent changes in volatile blends and photosynthetic characteristics in the annual herb Brassica nigra (L.) Koch. Heat stress was applied by either heating leaves through temperature response curve measurements from 20 to 40 °C (mild stress), exposing plants for 4 h to temperatures 25-44 °C (long-term stress) or shock-heating leaves to 45-50 °C. Photosynthetic reduction through temperature response curves was associated with decreased stomatal conductance, while the reduction due to long-term stress and collapse of photosynthetic activity after heat shock stress were associated with non-stomatal processes. Mild stress decreased constitutive monoterpene emissions, while long-term stress and shock stress resulted in emissions of the lipoxygenase pathway and glucosinolate volatiles. Glucosinolate volatile release was more strongly elicited by long-term stress and lipoxygenase product released by heat shock. These results demonstrate that glucosinolate volatiles constitute a major part of emission blend in heat-stressed B. nigra plants, especially upon chronic stress that leads to induction responses. © 2016 John Wiley & Sons Ltd.

Augmenting Sulfur Metabolism and Herbivore Defense in Arabidopsis by Bacterial Volatile Signaling.

PubMed

Aziz, Mina; Nadipalli, Ranjith K; Xie, Xitao; Sun, Yan; Surowiec, Kazimierz; Zhang, Jin-Lin; Paré, Paul W

2016-01-01

Sulfur is an element necessary for the life cycle of higher plants. Its assimilation and reduction into essential biomolecules are pivotal factors determining a plant's growth and vigor as well as resistance to environmental stress. While certain soil microbes can enhance ion solubility via chelating agents or oxidation, microbial regulation of plant-sulfur assimilation has not been reported. With an increasing understanding that soil microbes can activate growth and stress tolerance in plants via chemical signaling, the question arises as to whether such beneficial bacteria also regulate sulfur assimilation. Here we report a previously unidentified mechanism by which the growth-promoting rhizobacterium Bacillus amyloliquefaciens (GB03) transcriptionally activates genes responsible for sulfur assimilation, increasing sulfur uptake and accumulation in Arabidopsis. Transcripts encoding for sulfur-rich aliphatic and indolic glucosinolates are also GB03 induced. As a result, GB03-exposed plants with elevated glucosinolates exhibit greater protection against the generalist herbivore, Spodoptera exigua (beet armyworm, BAW). In contrast, a previously characterized glucosinolate mutant compromised in the production of both aliphatic and indolic glucosinolates is also compromised in terms of GB03-induced protection against insect herbivory. As with in vitro studies, soil-grown plants show enhanced glucosinolate accumulation and protection against BAW feeding with GB03 exposure. These results demonstrate the potential of microbes to enhance plant sulfur assimilation and emphasize the sophisticated integration of microbial signaling in plant defense.

Concentrations of thiocyanate and goitrin in human plasma, their precursor concentrations in brassica vegetables, and associated potential risk for hypothyroidism.

PubMed

Felker, Peter; Bunch, Ronald; Leung, Angela M

2016-04-01

Brassica vegetables are common components of the diet and have beneficial as well as potentially adverse health effects. Following enzymatic breakdown, some glucosinolates in brassica vegetables produce sulforaphane, phenethyl, and indolylic isothiocyanates that possess anticarcinogenic activity. In contrast, progoitrin and indolylic glucosinolates degrade to goitrin and thiocyanate, respectively, and may decrease thyroid hormone production. Radioiodine uptake to the thyroid is inhibited by 194 μmol of goitrin, but not by 77 μmol of goitrin. Collards, Brussels sprouts, and some Russian kale (Brassica napus) contain sufficient goitrin to potentially decrease iodine uptake by the thyroid. However, turnip tops, commercial broccoli, broccoli rabe, and kale belonging to Brassica oleracae contain less than 10 μmol of goitrin per 100-g serving and can be considered of minimal risk. Using sulforaphane plasma levels following glucoraphanin ingestion as a surrogate for thiocyanate plasma concentrations after indole glucosinolate ingestion, the maximum thiocyanate contribution from indole glucosinolate degradation is estimated to be 10 μM, which is significantly lower than background plasma thiocyanate concentrations (40-69 μM). Thiocyanate generated from consumption of indole glucosinolate can be assumed to have minimal adverse risks for thyroid health. © The Author(s) 2016. Published by Oxford University Press on behalf of the International Life Sciences Institute. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Genotype influences sulfur metabolism in broccoli (Brassica oleracea L.) under elevated CO2 and NaCl stress.

PubMed

Rodríguez-Hernández, María del Carmen; Moreno, Diego A; Carvajal, Micaela; Martínez-Ballesta, María del Carmen

2014-12-01

Climatic change predicts elevated salinity in soils as well as increased carbon dioxide dioxide [CO2] in the atmosphere. The present study aims to determine the effect of combined salinity and elevated [CO2] on sulfur (S) metabolism and S-derived phytochemicals in green and purple broccoli (cv. Naxos and cv. Viola, respectively). Elevated [CO2] involved the amelioration of salt stress, especially in cv. Viola, where a lower biomass reduction by salinity was accompanied by higher sodium (Na(+)) and chloride (Cl(-)) compartmentation in the vacuole. Moreover, salinity and elevated [CO2] affected the mineral and glucosinolate contents and the activity of biosynthetic enzymes of S-derived compounds and the degradative enzyme of glucosinolate metabolism, myrosinase, as well as the related amino acids and the antioxidant glutathione (GSH). In cv. Naxos, elevated [CO2] may trigger the antioxidant response to saline stress by means of increased GSH concentration. Also, in cv. Naxos, indolic glucosinolates were more influenced by the NaCl×CO2 interaction whereas in cv. Viola the aliphatic glucosinolates were significantly increased by these conditions. Salinity and elevated [CO2] enhanced the S cellular partitioning and metabolism affecting the myrosinase-glucosinolate system. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Impact of selenium supply on se-methylselenocysteine and glucosinolates accumulation in selenium-biofortified brassica sprouts

USDA-ARS?s Scientific Manuscript database

Brassica sprouts are widely marketed as functional foods. Here we examined the effects of Se treatment on the accumulation of anticancer compound Se-methylselenocysteine (SeMSCys) and glucosinolates in Brassica sprouts. Cultivars from the six most extensively consumed Brassica vegetables (broccoli, ...

The effect of post-harvest and packaging treatments on glucoraphanin concentration in broccoli (Brassica oleracea var. italica).

PubMed

Rangkadilok, Nuchanart; Tomkins, Bruce; Nicolas, Marc E; Premier, Robert R; Bennett, Richard N; Eagling, David R; Taylor, Paul W J

2002-12-04

The effects of post-harvest and packaging treatments on glucoraphanin (4-methylsulfinylbutyl glucosinolate), the glucosinolate precursor of anticancer isothiocyanate sulforaphane [4-methylsulfinylbutyl isothiocyanate], were examined in broccoli (Brassica oleracea var. italica) during storage times. The results showed that at 20 degrees C, 55% loss of glucoraphanin concentration occurred in broccoli stored in open boxes during the first 3 days of the treatment and 56% loss was found in broccoli stored in plastic bags by day 7. Under both air and controlled atmosphere (CA) storage, glucoraphanin concentration appeared to fluctuate slightly during 25 days of storage and the concentrations under CA was significantly higher than those stored under air treatment. In modified atmosphere packaging (MAP) treatments, glucoraphanin concentration in air control packaging decreased significantly whereas there were no significant changes in glucoraphanin concentration in MAP with no holes at 4 degrees C and two microholes at 20 degrees C for up to 10 days. Decreases in glucoraphanin concentration occurred when the broccoli heads deteriorated. In the present study, the best method for preserving glucoraphanin concentration in broccoli heads after harvest was storage of broccoli in MAP and refrigeration at 4 degrees C. This condition maintained the glucoraphanin concentration for at least 10 days and also maintained the visual quality of the broccoli heads.

Hierarchical nuclear and cytoplasmic genetic architectures for plant growth and defense within Arabidopsis.

PubMed

Joseph, Bindu; Corwin, Jason A; Züst, Tobias; Li, Baohua; Iravani, Majid; Schaepman-Strub, Gabriela; Turnbull, Lindsay A; Kliebenstein, Daniel J

2013-06-01

To understand how genetic architecture translates between phenotypic levels, we mapped the genetic architecture of growth and defense within the Arabidopsis thaliana Kas × Tsu recombinant inbred line population. We measured plant growth using traditional size measurements and size-corrected growth rates. This population contains genetic variation in both the nuclear and cytoplasmic genomes, allowing us to separate their contributions. The cytoplasmic genome regulated a significant variance in growth but not defense, which was due to cytonuclear epistasis. Furthermore, growth adhered to an infinitesimal model of genetic architecture, while defense metabolism was more of a moderate-effect model. We found a lack of concordance between quantitative trait loci (QTL) regulating defense and those regulating growth. Given the published evidence proving the link between glucosinolates and growth, this is likely a false negative result caused by the limited population size. This size limitation creates an inability to test the entire potential genetic landscape possible between these two parents. We uncovered a significant effect of glucosinolates on growth once we accounted for allelic differences in growth QTLs. Therefore, other growth QTLs can mask the effects of defense upon growth. Investigating direct links across phenotypic hierarchies is fraught with difficulty; we identify issues complicating this analysis.

Distribution of glucosinolates and sulphur-rich cells in roots of field-grown canola (Brassica napus).

PubMed

McCully, Margaret E; Miller, Celia; Sprague, Susan J; Huang, Cheng X; Kirkegaard, John A

2008-01-01

To investigate the role played by the distribution pattern of glucosinolates (GSLs) in root systems in the release of biocides to the rhizosphere, GSLs have been localized, for the first time, to specific regions and cells in field-grown roots. GSL concentrations in separated tissues of canola (Brassica napus) were determined by chemical analysis, and cell-specific concentrations by extrapolation from sulphur concentrations obtained by quantitative cryo-analytical scanning electron microscopy (SEM). In roots with secondary growth, GSL concentrations in the outer secondary tissues were up to 5x those of the inner core. The highest GSL concentrations (from sulphur measurements) were in two cell layers just under the outermost periderm layer, with up to 100x published concentrations for whole roots. Primary tissues had negligible GSL. Release and renewal of the peripheral GSLs is probably a normal developmental process as secondary thickening continues and surface cells senesce, accounting for published observations that intact roots release GSLs and their biocide hydrolosates to the rhizosphere. Absence of myrosin idioblasts close to the root surface suggests that GSLs released developmentally are hydrolysed by myrosinase in the rhizosphere, ensuring a continuous localized source of biotoxic hydrolysates which can deter soil-borne pests, and influence microbial populations associated with long-lived components of the root system.

Hierarchical Nuclear and Cytoplasmic Genetic Architectures for Plant Growth and Defense within Arabidopsis[C][W

PubMed Central

Joseph, Bindu; Corwin, Jason A.; Züst, Tobias; Li, Baohua; Iravani, Majid; Schaepman-Strub, Gabriela; Turnbull, Lindsay A.; Kliebenstein, Daniel J.

2013-01-01

To understand how genetic architecture translates between phenotypic levels, we mapped the genetic architecture of growth and defense within the Arabidopsis thaliana Kas × Tsu recombinant inbred line population. We measured plant growth using traditional size measurements and size-corrected growth rates. This population contains genetic variation in both the nuclear and cytoplasmic genomes, allowing us to separate their contributions. The cytoplasmic genome regulated a significant variance in growth but not defense, which was due to cytonuclear epistasis. Furthermore, growth adhered to an infinitesimal model of genetic architecture, while defense metabolism was more of a moderate-effect model. We found a lack of concordance between quantitative trait loci (QTL) regulating defense and those regulating growth. Given the published evidence proving the link between glucosinolates and growth, this is likely a false negative result caused by the limited population size. This size limitation creates an inability to test the entire potential genetic landscape possible between these two parents. We uncovered a significant effect of glucosinolates on growth once we accounted for allelic differences in growth QTLs. Therefore, other growth QTLs can mask the effects of defense upon growth. Investigating direct links across phenotypic hierarchies is fraught with difficulty; we identify issues complicating this analysis. PMID:23749847

An approach to the phytochemical profiling of rocket [Eruca sativa (Mill.) Thell].

PubMed

Villatoro-Pulido, Myriam; Priego-Capote, Feliciano; Álvarez-Sánchez, Beatriz; Saha, Shikha; Philo, Mark; Obregón-Cano, Sara; De Haro-Bailón, Antonio; Font, Rafael; Del Río-Celestino, Mercedes

2013-12-01

Eruca sativa (rocket) contains a wide range of compounds with nutraceutical and organoleptical properties. This research aimed to characterise the nutraceutical interest of four rocket accessions by analysis of glucosinolates, isothiocyanates, phenolics, carotenoids and carbohydrates. Different methods based on chromatographic separation with ultraviolet absorbance or mass spectrometry detection were used. The total content of glucosinolates ranged from 14.02 to 28.24 µmol g(-1) of dry weight. Glucoraphanin represented up to 52% of the total glucosinolates in leaves of one accession. Accessions showed differences in the hydrolysis of glucoraphanin to the isothiocyanate sulforaphane. No correlation between these compounds was observed, which insisted differences in the myrosinase activity within accessions. Rocket leaves had variable phenolic profiles represented by quercetin-3-glucoside, rutin, myricetin, quercetin and ferulic and p-coumaric acids. A high variability was observed for the total carotenoids ranged from 16.2 to 275 µg g(-1) with lutein as the main carotenoid. Glucose was the predominant sugar, representing >70% of the total soluble carbohydrates. Some accessions could be candidates for future breeding programmes because of their pattern of beneficial compounds for human health. However, further research is essential to evaluate the biological activity of these accessions before designing functional food. © 2013 Society of Chemical Industry.

Influence of cultivar and fertilizer approach on curly kale (Brassica oleracea L. var. sabellica). 1. Genetic diversity reflected in agronomic characteristics and phytochemical concentration.

PubMed

Groenbaek, Marie; Jensen, Sidsel; Neugart, Susanne; Schreiner, Monika; Kidmose, Ulla; Kristensen, Hanne Lakkenborg

2014-11-26

The objectives were to investigate if genetic diversity among field-grown traditional and F1 hybrid kale cultivars was reflected in different agronomic characteristics and consequently glucosinolate (GLS) and flavonoid glycoside concentration. This study evaluated how nitrogen and sulfur supply and biomass allocation modified phytochemicals in two experiments with combinations of three cultivars and four N and two S application levels. Results showed less growth, and higher N concentration in the traditional cultivar 'Tiara' was associated with increased indole and total GLSs compared to traditional 'Høj Amager Toftø' and F1 hybrid 'Reflex' cultivars, which exhibited higher yield, lower N concentration, and different biomass allocation. S application increased total GLS concentration, whereas aliphatic GLS percentage decreased when N application increased. Decrease of six 'Reflex' GLSs besides quercetin glycosides and total flavonoid glycosides with increased N indicated higher N responsiveness for 'Reflex'. In conclusion, differences in agronomic characteristics were reflected in diverse phytochemical composition.

Glucosinolate-derived compounds as a green manure for controlling Escherichia coli O157:H7 and Salmonella in soil

USDA-ARS?s Scientific Manuscript database

Plants from the Brassica family contain glucosinolate-derived compounds (GDC) which may act as natural antimicrobials in soil. Consequently, Brassica cover crops planted after harvest of the primary crop in the fall, and/or ntercropped during the growing season, could provide benefits derived from...

Reduction of total glucosinolates in canola meal via thermal treatment and fungal bioprocessing

USDA-ARS?s Scientific Manuscript database

On a worldwide basis, canola (Brassica napus) meal is second only to soybean meal as a protein source for livestock. A general limitation of Brassica spp. meals is the presence of glucosinolates (GLS). GLS and the enzyme myrosinase are compartmentally stored separately in the plant. Upon disruption ...

Assessing the anticancer compounds Se-methylselenocysteine and glucosinolates in Se-biofortified broccoli (brassica oleracea L. var. italica) sprouts and florets

USDA-ARS?s Scientific Manuscript database

Broccoli (Brassica oleracea L. var. italica) is a rich source of chemopreventive compounds. Here, we evaluated and compared the effect of selenium (Se) treatment on the accumulation of anticancer compound Se-methylselenocysteine (SeMSCys) and glucosinolates in broccoli sprouts and florets. Total Se ...

Changes in the glucosinolate-myrosinase defense system in Brassica juncea cotyledons during seedling development.

PubMed

Wallace, S K; Eigenbrode, Sanford D

2002-02-01

Optimal defense theory (ODT) predicts that plant defenses will be allocated to plant organs and tissues in proportion to their relative fitness values and susceptibilities to attack. This study was designed to test ODT predictions on the myrosinase-glucosinolate defense system in Brassica juncea by examining the relationships between the fitness value of B. juncea cotyledons and the levels and effectiveness of cotyledon defenses. Specifically, we estimated fitness value of cotyledons during plant development by measuring plant growth and seed production after cotyledon damage or removal at successive seedling ages. Cotyledon removal within five days of emergence had a significant impact on growth and seed production, but cotyledon removal at later stages did not. Consistent with ODT, glucosinolate and myrosinase levels in cotyledons also declined with seedling age, as did relative defenses against a generalist herbivore, Spodoptera eridania, as estimated by bioassay. Declines in glucosinolates were as predicted by a passive, allometric dilution model based on cotyledon expansion. Declines in myrosinase activity were significantly more gradual than predicted by allometric dilution, suggesting active retention of myrosinase activity as young cotyledons expand.

Assessment of the anticancer compounds Se-methylselenocysteine and glucosinolates in Se-biofortified broccoli (Brassica oleracea L. var. italica) sprouts and florets.

PubMed

Ávila, Fabricio William; Faquin, Valdemar; Yang, Yong; Ramos, Silvio Junio; Guilherme, Luiz Roberto G; Thannhauser, Theodore W; Li, Li

2013-07-03

Broccoli (Brassica oleracea L. var. italica) is a rich source of chemopreventive compounds. Here, we evaluated and compared the effect of selenium (Se) treatment on the accumulation of anticancer compounds Se-methylselenocysteine (SeMSCys) and glucosinolates in broccoli sprouts and florets. Total Se and SeMSCys content in sprouts increased concomitantly with increasing Se doses. Selenate was superior to selenite in inducing total Se accumulation, but selenite is equally effective as selenate in promoting SeMSCys synthesis in sprouts. Increasing sulfur doses reduced total Se and SeMSCys content in sprouts treated with selenate, but not in those with selenite. Examination of five broccoli cultivars reveals that sprouts generally have better fractional ability than florets to convert inorganic Se into SeMSCys. Distinctive glucosinolate profiles between sprouts and florets were observed, and sprouts contained approximately 6-fold more glucoraphanin than florets. In contrast to florets, glucosinolate content was not affected by Se treatment in sprouts. Thus, Se-enriched broccoli sprouts are excellent for simultaneous accumulation of chemopreventive compounds SeMSCys and glucoraphanin.

Characterization of industrial broccoli discards (Brassica oleracea var. italica) for their glucosinolate, polyphenol and flavonoid contents using UPLC MS/MS and spectrophotometric methods.

PubMed

Thomas, Minty; Badr, Ashraf; Desjardins, Yves; Gosselin, Andre; Angers, Paul

2018-04-15

The agrifood industry produces tons of waste and substandard products that are discarded at great expense. Valorization of industrial residues curbs issues related to food security and environmental problems. Broccoli (Brassica oleracea var. italica) is associated with varied beneficial health effects, but its production yields greater than 25% rejects. We aimed to characterize and quantify industrial broccoli by-products for their glucosinolate and polyphenol contents as a first step towards industrial bio-refining. Broccoli segments and rejected lots of 10 seed cultivars were analyzed using UPLC MS/MS. Variability in the contents of bioactive molecules was observed within and between the cultivars. Broccoli by-products were rich in glucosinolates (0.2-2% dry weight sample), predominantly glucoraphanin (32-64% of the total glucosinolates), whereas the polyphenolic content was less than 0.02% dry weight sample. Valorization of industrial residues facilitates the production of high value functional food ingredients along with socio-economic sustainability. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mid-infrared spectral characteristics of lipid molecular structures in Brassica carinata seeds: relationship to oil content, fatty acid and glucosinolate profiles, polyphenols, and condensed tannins.

PubMed

Xin, Hangshu; Khan, Nazir A; Falk, Kevin C; Yu, Peiqiang

2014-08-13

The objectives of this study were to quantify lipid-related inherent molecular structures using a Fourier transform infrared spectroscopy (FT-IR) technique and determine their relationship to oil content, fatty acid and glucosinolate profile, total polyphenols, and condensed tannins in seeds from newly developed yellow-seeded and brown-seeded Brassica carinata lines. Canola seeds were used as a reference. The lipid-related molecular spectral band intensities were strongly correlated to the contents of oil, fatty acids, glucosinolates, and polyphenols. The regression equations gave relatively high predictive power for the estimation of oil (R² = 0.99); all measured fatty acids (R² > 0.80), except C14:0, C20:3n-3, C22:2n-9, and C22:2n-6; 3-butenyl, 2-OH-3-butenyl, 4-OH-3-CH3-indolyl, and total glucosinolates (R² > 0.686); and total polyphenols (R² = 0.935). However, further study is required to obtain predictive equations based on large numbers of samples from diverse sources to illustrate the general applicability of these regression equations.

The SAL-PAP Chloroplast Retrograde Pathway Contributes to Plant Immunity by Regulating Glucosinolate Pathway and Phytohormone Signaling.

PubMed

Ishiga, Yasuhiro; Watanabe, Mutsumi; Ishiga, Takako; Tohge, Takayuki; Matsuura, Takakazu; Ikeda, Yoko; Hoefgen, Rainer; Fernie, Alisdair R; Mysore, Kirankumar S

2017-10-01

Chloroplasts have a crucial role in plant immunity against pathogens. Increasing evidence suggests that phytopathogens target chloroplast homeostasis as a pathogenicity mechanism. In order to regulate the performance of chloroplasts under stress conditions, chloroplasts produce retrograde signals to alter nuclear gene expression. Many signals for the chloroplast retrograde pathway have been identified, including chlorophyll intermediates, reactive oxygen species, and metabolic retrograde signals. Although there is a reasonably good understanding of chloroplast retrograde signaling in plant immunity, some signals are not well-understood. In order to understand the role of chloroplast retrograde signaling in plant immunity, we investigated Arabidopsis chloroplast retrograde signaling mutants in response to pathogen inoculation. sal1 mutants (fry1-2 and alx8) responsible for the SAL1-PAP retrograde signaling pathway showed enhanced disease symptoms not only to the hemibiotrophic pathogen Pseudomonas syringae pv. tomato DC3000 but, also, to the necrotrophic pathogen Pectobacterium carotovorum subsp. carotovorum EC1. Glucosinolate profiles demonstrated the reduced accumulation of aliphatic glucosinolates in the fry1-2 and alx8 mutants compared with the wild-type Col-0 in response to DC3000 infection. In addition, quantification of multiple phytohormones and analyses of their gene expression profiles revealed that both the salicylic acid (SA)- and jasmonic acid (JA)-mediated signaling pathways were down-regulated in the fry1-2 and alx8 mutants. These results suggest that the SAL1-PAP chloroplast retrograde pathway is involved in plant immunity by regulating the SA- and JA-mediated signaling pathways.

Exogenous glucosinolate produced by Arabidopsis thaliana has an impact on microbes in the rhizosphere and plant roots.

PubMed

Bressan, Mélanie; Roncato, Marie-Anne; Bellvert, Floriant; Comte, Gilles; Haichar, Feth Zahar; Achouak, Wafa; Berge, Odile

2009-11-01

A specificity of Brassicaceous plants is the production of sulphur secondary metabolites called glucosinolates that can be hydrolysed into glucose and biocidal products. Among them, isothiocyanates are toxic to a wide range of microorganisms and particularly soil-borne pathogens. The aim of this study was to investigate the role of glucosinolates and their breakdown products as a factor of selection on rhizosphere microbial community associated with living Brassicaceae. We used a DNA-stable isotope probing approach to focus on the active microbial populations involved in root exudates degradation in rhizosphere. A transgenic Arabidopsis thaliana line producing an exogenous glucosinolate and the associated wild-type plant associated were grown under an enriched (13)CO(2) atmosphere in natural soil. DNA from the rhizospheric soil was separated by density gradient centrifugation. Bacterial (Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria and Acidobacteria), Archaea and fungal community structures were analysed by DGGE fingerprints of amplified 16S and 18S rRNA gene sequences. Specific populations were characterized by sequencing DGGE fragments. Roots of the transgenic plant line presented an altered profile of glucosinolates and other minor additional modifications. These modifications significantly influenced microbial community on roots and active populations in the rhizosphere. Alphaproteobacteria, particularly Rhizobiaceae, and fungal communities were mainly impacted by these Brassicaceous metabolites, in both structure and composition. Our results showed that even a minor modification in plant root could have important repercussions for soil microbial communities.

Development of a generalist predator, Podisus maculiventris, on glucosinolate sequestering and nonsequestering prey

NASA Astrophysics Data System (ADS)

van Geem, Moniek; Harvey, Jeffrey A.; Gols, Rieta

2014-09-01

Insect herbivores exhibit various strategies to counter the toxic effects of plant chemical defenses. These strategies include the detoxification, excretion, and sequestration of plant secondary metabolites. The latter strategy is often considered to provide an additional benefit in that it provides herbivores with protection against natural enemies such as predators. Profiles of sequestered chemicals are influenced by the food plants from which these chemicals are derived. We compared the effects of sequestration and nonsequestration of plant secondary metabolites in two specialist herbivores on the development of a generalist predator, Podisus maculiventris. Profiles of glucosinolates, secondary metabolites characteristic for the Brassicaceae, are known to differ considerably both inter- and intraspecifically. Throughout their immature (=nymphal) development, the predator was fed on larval stages of either sequestering (turnip sawfly, Athalia rosae) or nonsequestering (small cabbage white butterfly, Pieris rapae) prey that in turn had been feeding on plants originating from three wild cabbage ( Brassica oleracea) populations that have previously been shown to differ in their glucosinolate profiles. We compared survival, development time, and adult body mass as parameters for bug performance. Our results show that sequestration of glucosinolates by A. rosae only marginally affected the development of P. maculiventris. The effects of plant population on predator performance were variable. We suggest that sequestration of glucosinolates by A. rosae functions not only as a defensive mechanism against some predators, but may also be an alternative way of harmlessly dealing with plant allelochemicals.

Dynamics of glucosinolate-myrosinase system during Plutella xylostella interaction to a novel host Lepidium latifolium L.

PubMed

Kaur, Tarandeep; Bhat, Rohini; Khajuria, Manu; Vyas, Ruchika; Kumari, Anika; Nadda, Gireesh; Vishwakarma, Ram; Vyas, Dhiraj

2016-09-01

Plutella xylostella L. is a notorious pest of cruciferous crops causing worldwide losses of $4-5 billion per year. Developing classical biological control to this pest include an introduction of host plants that act as natural enemies showing deviation from the preference-performance regimen in the evolutionary ecology of plant-insect interactions. The present study was designed to understand the role of glucosinolate-myrosinase system during P. xylostella interactions with a novel host. Adult moth preference and larval performance study were conducted on a novel host Lepidium latifolium L. (LL) that has high sinigrin content and was compared with its laboratory host Arabidopsis thaliana (AT). The glucosinolate-myrosinase system was studied in a time course experiment during larval feeding in choice and no-choice experiments. Adult moths visit and prefers LL over AT for oviposition. Conversely, LL leaves were not preferred and proved detrimental for P. xylostella larvae. Aliphatic and indolic glucosinolates were found to decrease significantly (p≤0.05) in AT during initial 12h of P. xylostella challenge, whereas, they were not affected in LL. Also, MYB transcription factor expression and myrosinase activity in LL do not suggest a typical host response to a specialist insect. This preference-performance mismatch of P. xylostella on LL mediated by glucosinolate pattern suggests that this novel plant could be utilized in P. xylostella management. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

RNA-Seq Analysis of Transcriptome and Glucosinolate Metabolism in Seeds and Sprouts of Broccoli (Brassica oleracea var. italic)

PubMed Central

Gao, Jinjun; Yu, Xinxin; Ma, Fengming; Li, Jing

2014-01-01

Background Broccoli (Brassica oleracea var. italica), a member of Cruciferae, is an important vegetable containing high concentration of various nutritive and functional molecules especially the anticarcinogenic glucosinolates. The sprouts of broccoli contain 10–100 times higher level of glucoraphanin, the main contributor of the anticarcinogenesis, than the edible florets. Despite the broccoli sprouts’ functional importance, currently available genetic and genomic tools for their studies are very limited, which greatly restricts the development of this functionally important vegetable. Results A total of ∼85 million 251 bp reads were obtained. After de novo assembly and searching the assembled transcripts against the Arabidopsis thaliana and NCBI nr databases, 19,441 top-hit transcripts were clustered as unigenes with an average length of 2,133 bp. These unigenes were classified according to their putative functional categories. Cluster analysis of total unigenes with similar expression patterns and differentially expressed unigenes among different tissues, as well as transcription factor analysis were performed. We identified 25 putative glucosinolate metabolism genes sharing 62.04–89.72% nucleotide sequence identity with the Arabidopsis orthologs. This established a broccoli glucosinolate metabolic pathway with high colinearity to Arabidopsis. Many of the biosynthetic and degradation genes showed higher expression after germination than in seeds; especially the expression of the myrosinase TGG2 was 20–130 times higher. These results along with the previous reports about these genes’ studies in Arabidopsis and the glucosinolate concentration in broccoli sprouts indicate the breakdown products of glucosinolates may play important roles in the stage of broccoli seed germination and sprout development. Conclusion Our study provides the largest genetic resource of broccoli to date. These data will pave the way for further studies and genetic engineering of broccoli sprouts and will also provide new insight into the genomic research of this species and its relatives. PMID:24586398

RNA-seq analysis of transcriptome and glucosinolate metabolism in seeds and sprouts of broccoli (Brassica oleracea var. italic).

PubMed

Gao, Jinjun; Yu, Xinxin; Ma, Fengming; Li, Jing

2014-01-01

Broccoli (Brassica oleracea var. italica), a member of Cruciferae, is an important vegetable containing high concentration of various nutritive and functional molecules especially the anticarcinogenic glucosinolates. The sprouts of broccoli contain 10-100 times higher level of glucoraphanin, the main contributor of the anticarcinogenesis, than the edible florets. Despite the broccoli sprouts' functional importance, currently available genetic and genomic tools for their studies are very limited, which greatly restricts the development of this functionally important vegetable. A total of ∼85 million 251 bp reads were obtained. After de novo assembly and searching the assembled transcripts against the Arabidopsis thaliana and NCBI nr databases, 19,441 top-hit transcripts were clustered as unigenes with an average length of 2,133 bp. These unigenes were classified according to their putative functional categories. Cluster analysis of total unigenes with similar expression patterns and differentially expressed unigenes among different tissues, as well as transcription factor analysis were performed. We identified 25 putative glucosinolate metabolism genes sharing 62.04-89.72% nucleotide sequence identity with the Arabidopsis orthologs. This established a broccoli glucosinolate metabolic pathway with high colinearity to Arabidopsis. Many of the biosynthetic and degradation genes showed higher expression after germination than in seeds; especially the expression of the myrosinase TGG2 was 20-130 times higher. These results along with the previous reports about these genes' studies in Arabidopsis and the glucosinolate concentration in broccoli sprouts indicate the breakdown products of glucosinolates may play important roles in the stage of broccoli seed germination and sprout development. Our study provides the largest genetic resource of broccoli to date. These data will pave the way for further studies and genetic engineering of broccoli sprouts and will also provide new insight into the genomic research of this species and its relatives.

Herbivore Diet Breadth and Host Plant Defense Mediate the Tri-Trophic Effects of Plant Toxins on Multiple Coccinellid Predators.

PubMed

Katsanis, Angelos; Rasmann, Sergio; Mooney, Kailen A

2016-01-01

Host plant defenses are known to cascade up food chains to influence herbivores and their natural enemies, but how herbivore and predator traits and identity mediate such tri-trophic dynamics is largely unknown. We assessed the influence of plant defense on aphid and coccinellid performance in laboratory trials with low- vs. high-glucosinolate varieties of Brassica napus, a dietary specialist (Brevicoryne brassicae) and generalist (Myzus persicae) aphid, and five species of aphidophagous coccinellids. The performance of the specialist and generalist aphids was similar and unaffected by variation in plant defense. Aphid glucosinolate concentration and resistance to predators differed by aphid species and host plant defense, and these effects acted independently. With respect to aphid species, the dietary generalist aphid (vs. specialist) had 14% lower glucosinolate concentration and coccinellid predators ate three-fold more aphids. With respect to host plant variety, the high-glucosinolate plants (vs. low) increased aphid glucosinolate concentration by 21%, but had relatively weak effects on predation by coccinellids and these effects varied among coccinellid species. In turn, coccinellid performance was influenced by the interactive effects of plant defense and aphid species, as the cascading, indirect effect of plant defense was greater when feeding upon the specialist than generalist aphid. When feeding upon specialist aphids, low- (vs. high-) glucosinolate plants increased coccinellid mass gain by 78% and accelerated development by 14%. In contrast, when feeding upon generalist aphids, low- (vs. high-) glucosinolate plants increased coccinellid mass gain by only 11% and had no detectable effect on development time. These interactive effects of plant defense and aphid diet breadth on predator performance also varied among coccinellid species; the indirect negative effects of plant defenses on predator performance was consistent among the five predators when transmitted via the dietary specialist aphid, but these effects varied substantially among predators-in both the magnitude and direction-when transmitted via the dietary generalist aphid. Accordingly, the cascading effect of plant defense on predators was stronger in magnitude and more consistent among predator taxa when transmitted by the specialist than generalist herbivore. Overall, these findings support a central role of herbivore diet breadth in mediating both the strength and contingency of tri-trophic interactions.

Proliferation and glucosinolates accumulation of broccoli adventitious roots in liquid medium

NASA Astrophysics Data System (ADS)

Nhut, Nguyen Minh; Tien, Le Thi Thuy

2017-09-01

Cotyledons from 7-day-old in vitro broccoli seedling were used as explant source in adventitious root induction on MS medium supplemented with 30 g/l sucrose, 1.6 mg/l IBA and 7 g/l agar. Adventitious roots from cotyledons were transferred to liquid medium containing the same components as rooting medium for two weeks, then subcultured to MS medium with diferent sugar, macrominerals and casein hydrolysate concentrations. The best adventitious root growth was observed in half-strength MS medium supplemented with 40 g/l sucrose, 600 mg/l casein hydrolysate and 1.6 mg/l IBA (growth index of 4.00 in about 14 culture days with inoculum density of 1.0 g fresh weight / 30 ml of culture medium). The culturing process can be stopped on the 28th day for root biomass and on the 35th day for glucosinolates.

The Brassica epithionitrile 1-cyano-2,3-epithiopropane triggers cell death in human liver cancer cells in vitro.

PubMed

Hanschen, Franziska S; Herz, Corinna; Schlotz, Nina; Kupke, Franziska; Bartolomé Rodríguez, María M; Schreiner, Monika; Rohn, Sascha; Lamy, Evelyn

2015-11-01

Glucosinolates are secondary metabolites present in Brassica vegetables. Alkenyl glucosinolates are enzymatically degraded forming nitriles or isothiocyanates, but in the presence of epithiospecifier protein, epithionitriles are released. However, studies on the occurrence of epithionitriles in Brassica food and knowledge about their biological effects are scarce. Epithionitrile formation from glucosinolates of seven Brassica vegetables was analyzed using GC-MS and HPLC-DAD. Bioactivity of synthetic and plant-derived 1-cyano-2,3-epithiopropane (CETP) - the predominant epithionitrile in Brassica vegetables - in three human hepatocellular carcinoma (HCC) cell lines and primary murine hepatocytes was also evaluated. The majority of the Brassica vegetables were producers of nitriles or epithionitriles as hydrolysis products and not of isothiocyanates. For example, Brussels sprouts and savoy cabbage contained up to 0.8 μmol CETP/g vegetable. Using formazan dye assays, concentrations of 380-1500 nM CETP were observed to inhibit the mitochondrial dehydrogenase activity of human HCC cells without impairment of cell growth. At 100-fold higher CETP concentrations, cell death was observed. Presence of plant matrix increased CETP-based toxicity. These in vitro data provide no indication that epithionitriles will severely affect human health by Brassica consumption. In contrast to isothiocyanates, no evidence of selective toxicity against HCC cells was found. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Quality characteristics of the radish grown under reduced atmospheric pressure

NASA Astrophysics Data System (ADS)

Levine, Lanfang H.; Bisbee, Patricia A.; Richards, Jeffrey T.; Birmele, Michele N.; Prior, Ronald L.; Perchonok, Michele; Dixon, Mike; Yorio, Neil C.; Stutte, Gary W.; Wheeler, Raymond M.

This study addresses whether reduced atmospheric pressure (hypobaria) affects the quality traits of radish grown under such environments. Radish (Raphanus sativus L. cv. Cherry Bomb Hybrid II) plants were grown hydroponically in specially designed hypobaric plant growth chambers at three atmospheric pressures; 33, 66, and 96 kPa (control). Oxygen and carbon dioxide partial pressures were maintained constant at 21 and 0.12 kPa, respectively. Plants were harvested at 21 days after planting, with aerial shoots and swollen hypocotyls (edible portion of the radish referred to as the “root” hereafter) separated immediately upon removal from the chambers. Samples were subsequently evaluated for their sensory characteristics (color, taste, overall appearance, and texture), taste-determining factors (glucosinolate and soluble carbohydrate content and myrosinase activity), proximate nutrients (protein, dietary fiber, and carbohydrate) and potential health benefit attributes (antioxidant capacity). In roots of control plants, concentrations of glucosinolate, total soluble sugar, and nitrate, as well as myrosinase activity and total antioxidant capacity (measured as ORACFL), were 2.9, 20, 5.1, 9.4, and 1.9 times greater than the amount in leaves, respectively. There was no significant difference in total antioxidant capacity, sensory characteristics, carbohydrate composition, or proximate nutrient content among the three pressure treatments. However, glucosinolate content in the root and nitrate concentration in the leaf declined as the atmospheric pressure decreased, suggesting perturbation to some nitrogen-related metabolism.

Characterisation of taste-active extracts from raw Brassica oleracea vegetables.

PubMed

Zabaras, Dimitrios; Roohani, Mahshid; Krishnamurthy, Raju; Cochet, Maeva; Delahunty, Conor M

2013-04-25

Chemical and sensory characterisation of whole and fractionated myrosinase-free extracts from selected Australian-grown, raw Brassica vegetables (broccoli, cauliflower, Brussels sprouts and red cabbage) was carried out to determine the contribution of key phytochemicals (i.e. glucosinolates, free sugars, phenolics) to the taste profiles of these vegetables. Glucosinolate (GS) and phenolic profiles were determined by liquid chromatography coupled with photodiode array detection and mass spectrometry. Ten glucosinolates (GS) were quantified across the vegetables investigated. Brussels sprouts (186.3 μg g(-1) FW) followed by broccoli (164.1 μg g(-1) FW) were found to contain the most GS. The phenolic profiles of all samples were dominated by hydroxycinnamic acid derivatives. As expected, red cabbage was the only vegetable with a significant anthocyanin signal (574.0 μg g(-1) FW). Red cabbage (26.7 mg g(-1) FW) and cauliflower (18.7 mg g(-1) FW) were found to contain a higher concentration of free sugars than Brussels sprouts (12.6 mg g(-1) FW) and broccoli (10.2 mg g(-1) FW). Descriptive sensory analysis of the whole extracts found sweetness (cauliflower and red cabbage sweeter than broccoli and Brussels sprouts) and bitterness (Brussels sprouts more bitter than others) as the most discriminating attributes. A hydrophilic fraction with sweetness, umami and saltiness as the main attributes was the most taste active fraction across all Brassica whole extracts. Sub-fractionation showed that this fraction was also bitter but the presence of sugars counteracted bitterness. Several components within each extract were found to contribute to the bitterness of whole Brassica extracts. The total and individual GS content alone could not explain the perceived bitterness of these extracts. Phenolics and/or other components are likely to be contributing to the bitterness associated with these vegetables.

Peruvian Maca (Lepidium peruvianum): (I) Phytochemical and Genetic Differences in Three Maca Phenotypes.

PubMed

Meissner, Henry O; Mscisz, Alina; Mrozikiewicz, Mieczyslaw; Baraniak, Marek; Mielcarek, Sebastian; Kedzia, Bogdan; Piatkowska, Ewa; Jólkowska, Justyna; Pisulewski, Pawel

2015-09-01

Glucosinolates were previously reported as physiologically-important constituents present in Peruvian Maca (Lepidium peruvianum Chacon) and linked to various therapeutic functions of differently-colored Peruvian Maca hypocotyls. In two separate Trials, three colours of Maca hypocotyls "Black", "Red" and "Yellow" (termed "Maca phenotypes"), were selected from mixed crops of Peruvian Maca for laboratory studies as fresh and after being dried. Individual Maca phenotypes were cultivated in the highlands of the Peruvian Andes at 4,200m a.s.l. (Junin and Ninacaca). Glucosinolate levels, chromatographic HPLC profiles and DNA variability in the investigated Maca phenotypes are presented. Genotypic profiles were determined by the ISSR-PCR and RAPD techniques. Compared to the Black and Red phenotypes, the Yellow phenotype contained much lower Glucosinolate levels measured against Glucotropaeolin and m-methoxy-glucotropaeolin standards, and exhibited different RAPD and ISSR-PCR reactions. The Red Maca phenotype showed the highest concentrations of Glucosinolates as compared to the Black and Yellow Maca. It appears that the traditional system used by natives of the Peruvian Andean highlands in preparing Maca as a vegetable dish (boiling dried Maca after soaking in water), to supplement their daily meals, is as effective as laboratory methods - for extracting Glucosinolates, which are considered to be one of the key bioactive constituents responsible for therapeutic functions of Peruvian Maca phenotypes. It is reasonable to assume that the HPLC and DNA techniques combined, or separately, may assist in determining ID and "Fingerprints" identifying individual Peruvian Maca phenotypes, hence confirming the authenticity of marketable Maca products. The above assumptions warrant further laboratory testing.

Peruvian Maca (Lepidium peruvianum): (I) Phytochemical and Genetic Differences in Three Maca Phenotypes

PubMed Central

Meissner, Henry O.; Mscisz, Alina; Mrozikiewicz, Mieczyslaw; Baraniak, Marek; Mielcarek, Sebastian; Kedzia, Bogdan; Piatkowska, Ewa; Jólkowska, Justyna; Pisulewski, Pawel

2015-01-01

Glucosinolates were previously reported as physiologically-important constituents present in Peruvian Maca (Lepidium peruvianum Chacon) and linked to various therapeutic functions of differently-colored Peruvian Maca hypocotyls. In two separate Trials, three colours of Maca hypocotyls “Black”, “Red” and “Yellow” (termed “Maca phenotypes”), were selected from mixed crops of Peruvian Maca for laboratory studies as fresh and after being dried. Individual Maca phenotypes were cultivated in the highlands of the Peruvian Andes at 4,200m a.s.l. (Junin and Ninacaca). Glucosinolate levels, chromatographic HPLC profiles and DNA variability in the investigated Maca phenotypes are presented. Genotypic profiles were determined by the ISSR-PCR and RAPD techniques. Compared to the Black and Red phenotypes, the Yellow phenotype contained much lower Glucosinolate levels measured against Glucotropaeolin and m-methoxy-glucotropaeolin standards, and exhibited different RAPD and ISSR-PCR reactions. The Red Maca phenotype showed the highest concentrations of Glucosinolates as compared to the Black and Yellow Maca. It appears that the traditional system used by natives of the Peruvian Andean highlands in preparing Maca as a vegetable dish (boiling dried Maca after soaking in water), to supplement their daily meals, is as effective as laboratory methods - for extracting Glucosinolates, which are considered to be one of the key bioactive constituents responsible for therapeutic functions of Peruvian Maca phenotypes. It is reasonable to assume that the HPLC and DNA techniques combined, or separately, may assist in determining ID and “Fingerprints” identifying individual Peruvian Maca phenotypes, hence confirming the authenticity of marketable Maca products. The above assumptions warrant further laboratory testing. PMID:26508907

Evaluation of different cooking conditions on broccoli (Brassica oleracea var. italica) to improve the nutritional value and consumer acceptance.

PubMed

Bongoni, Radhika; Verkerk, Ruud; Steenbekkers, Bea; Dekker, Matthijs; Stieger, Markus

2014-09-01

The objective of this study was to gain insights into the effect of the cooking method on the liking as well as the retention of glucosinolates in broccoli. With this knowledge it can be concluded whether the health aspects of broccoli be improved by the cooking method without deteriorating sensory perception. For this, broccoli was cooked by methods commonly applied by consumers: boiling with a cold (water) start; boiling with a hot (water) start; and steaming. Firmness, greenness and amount of total glucosinolates in cooked broccoli were instrumentally determined. Sensory evaluation by untrained consumers (n = 99) for liking and sensory attributes intensity rating were performed on broccoli cooked by steaming and boiling-cold start at three time points, which resulted in 'high', 'medium', 'low' firm broccoli samples. At the end of cooking, steaming showed an increase in the amount of total glucosinolates (+17%). Boiling-hot start (-41%) and boiling-cold start (-50%) showed a decrease in amount of total glucosinolates. Sensory evaluation did not show statistically significant differences between steaming and boiling-cold start in liking at 'high' and 'medium' firmness; and in the attribute intensity ratings (except for juiciness at 'medium' firmness, and flavour at 'medium' and 'low' firmness). This study demonstrates that medium firm broccoli showed optimum liking and that steaming compared to boiled-cold start showed higher amount of glucosinolates. It is concluded that the health aspects of broccoli can be improved without reducing the sensory aspects by optimising the cooking method.

The microRNA miR393 re-directs secondary metabolite biosynthesis away from camalexin and towards glucosinolates.

PubMed

Robert-Seilaniantz, Alexandre; MacLean, Dan; Jikumaru, Yusuke; Hill, Lionel; Yamaguchi, Shinjiro; Kamiya, Yuji; Jones, Jonathan D G

2011-07-01

flg22 treatment increases levels of miR393, a microRNA that targets auxin receptors. Over-expression of miR393 renders plants more resistant to biotroph pathogens and more susceptible to necrotroph pathogens. In contrast, over-expression of AFB1, an auxin receptor whose mRNA is partially resistant to miR393 degradation, renders the plant more susceptible to biotroph pathogens. Here we investigate the mechanism by which auxin signalling and miR393 influence plant defence. We show that auxin signalling represses SA levels and signalling. We also show that miR393 represses auxin signalling, preventing it from antagonizing SA signalling. In addition, over-expression of miR393 increases glucosinolate levels and decreases the levels of camalexin. Further studies on pathogen interactions in auxin signalling mutants revealed that ARF1 and ARF9 negatively regulate glucosinolate accumulation, and that ARF9 positively regulates camalexin accumulation. We propose that the action of miR393 on auxin signalling triggers two complementary responses. First, it prevents suppression of SA levels by auxin. Second, it stabilizes ARF1 and ARF9 in inactive complexes. As a result, the plant is able to mount a full SA response and to re-direct metabolic flow toward the most effective anti-microbial compounds for biotroph resistance. We propose that miR393 levels can fine-tune plant defences and prioritize resources. © 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.

Comparative Phytonutrient Analysis of Broccoli By-Products: The Potentials for Broccoli By-Product Utilization.

PubMed

Liu, Mengpei; Zhang, Lihua; Ser, Suk Lan; Cumming, Jonathan R; Ku, Kang-Mo

2018-04-13

The phytonutrient concentrations of broccoli ( Brassica oleracea var. italica) florets, stems, and leaves were compared to evaluate the value of stem and leaf by-products as a source of valuable nutrients. Primary metabolites, including amino acids, organic acids, and sugars, as well as glucosinolates, carotenoids, chlorophylls, vitamins E and K, essential mineral elements, total phenolic content, antioxidant activity, and expression of glucosinolate biosynthesis and hydrolysis genes were quantified from the different broccoli tissues. Broccoli florets had higher concentrations of amino acids, glucoraphanin, and neoglucobrassicin compared to other tissues, whereas leaves were higher in carotenoids, chlorophylls, vitamins E and K, total phenolic content, and antioxidant activity. Leaves were also good sources of calcium and manganese compared to other tissues. Stems had the lowest nitrile formation from glucosinolate. Each tissue exhibited specific core gene expression profiles supporting glucosinolate metabolism, with different gene homologs expressed in florets, stems, and leaves, which suggests that tissue-specific pathways function to support primary and secondary metabolic pathways in broccoli. This comprehensive nutrient and bioactive compound profile represents a useful resource for the evaluation of broccoli by-product utilization in the human diet, and as feedstocks for bioactive compounds for industry.

Identification of Glucosinolates in Seeds of Three Brassicaceae Species Known to Hyperaccumulate Heavy Metals.

PubMed

Montaut, Sabine; Guido, Benjamin S; Grison, Claude; Rollin, Patrick

2017-03-01

Plants from the Brassicaceae family are known to contain secondary metabolites called glucosinolates. Our goal was to establish by LC/MS the glucosinolate profile of seeds of three Brassicaceae species known to hyperaccumulate heavy metals. We investigated Alyssum fallacinum auct. non Hausskn., Iberis intermedia Guers., and Noccaea caerulescens (J. Presl & C. Presl) F. K. Mey. Our results indicate that A. fallacinum seeds contain glucoiberin and glucoibervirin, which had not been previously identified in this plant. Furthermore, we report for the first time the presence of glucoiberin, glucoibervirin, glucotropaeolin, and sinigrin in I. intermedia. We have detected for the first time glucoconringiin in N. caerulescens. In addition, glucosinalbin, 4-hydroxyglucobrassicin, and glucomoringin were also detected. © 2017 Wiley-VHCA AG, Zurich, Switzerland.

Genes affecting novel seed constituents in Limnanthes alba Benth: transcriptome analysis of developing embryos and a new genetic map of meadowfoam

PubMed Central

Cooper, Laurel D.; Kishore, Venkata K.; Knapp, Steven J.; Kling, Jennifer G.

2015-01-01

The seed oil of meadowfoam, a new crop in the Limnanthaceae family, is highly enriched in very long chain fatty acids that are desaturated at the Δ5 position. The unusual oil is desirable for cosmetics and innovative industrial applications and the seed meal remaining after oil extraction contains glucolimnanthin, a methoxylated benzylglucosinolate whose degradation products are herbicidal and anti-microbial. Here we describe EST analysis of the developing seed transcriptome that identified major genes involved in biosynthesis and assembly of the seed oil and in glucosinolate metabolic pathways. mRNAs encoding acyl-CoA Δ5 desaturase were notably abundant. The library was searched for simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs). Fifty-four new SSR markers and eight candidate gene markers were developed and combined with previously developed SSRs to construct a new genetic map for Limnanthes alba. Mapped genes in the lipid biosynthetic pathway encode 3-ketoacyl-CoA synthase (KCS), Δ5 desaturase (Δ5DS), lysophosphatidylacyl-acyl transferase (LPAT), and acyl-CoA diacylglycerol acyl transferase (DGAT). Mapped genes in glucosinolate biosynthetic and degradation pathways encode CYP79A, myrosinase (TGG), and epithiospecifier modifier protein (ESM). The resources developed in this study will further the domestication and improvement of meadowfoam as an oilseed crop. PMID:26038713

Peruvian Maca (Lepidium peruvianum): (II) Phytochemical Profiles of Four Prime Maca Phenotypes Grown in Two Geographically-Distant Locations

PubMed Central

O. Meissner, Henry; Mscisz, Alina; Piatkowska, Ewa; Baraniak, Marek; Mielcarek, Sebastian; Kedzia, Bogdan; Holderna-Kedzia, Elzbieta; Pisulewski, Pawel

2016-01-01

Peruvian Maca crops (Lepidium peruvianum), grown in two geographically-distant cultivation sites located at similar altitudes in the highlands of the Peruvian Andes (Junin at 4,200 m a.s.l. and Ancash 4,150 m a.s.l.), were used in the study. Four prime Maca phenotypes, distinguished by hypocotyl colours labelled as “Yellow”, “Purple”, “Red” and “Black” were selected to determine distribution in levels and corresponding ratios between individual Glucosinolates (Glucotropaeolin and m-methylglucotropaeolin) in an attempt to identify four Peruvian Maca phenotypes from analyses of powdered hypocotyls. There were highly significant differences (P<0.01) in hypocotyl weight/size of four Maca phenotypes harvested in two locations. The Junin crop represented a mostly “large” class (13.3 g) with “small” size hypocotyls (7.2 g), while a “small” class was predominant in Ancash (3.5 g). Powdered Yellow Maca showed significantly higher (P<0.001) microbial contamination than the other three, with Black Maca being the least infected. Only minor, statistically-confirmed differences were detected in nutritive characteristics between the four Maca phenotypes grown in Junin, however highly significant differences (P<0.01) in Glucosinolates existed between the Red and Black Maca grown in Junin and Ancash. Irrespective of the cultivation location, Red phenotypes showed the highest content of Total Glucosinolates, followed by Black and Purple, with the Yellow phenotype showing consistently lower levels. Highly significant P<0.01) differences determined in ratios of individual Glucosinolates between four Maca phenotypes grown in two locations, confirms an earlier assumption that sums of individual Glucosinolates, their ratios and profiles, may be feasible to explore in analytically identifying individual Maca phenotypes in pulverised marketed Maca products. PMID:27127450

Peruvian Maca (Lepidium peruvianum): (II) Phytochemical Profiles of Four Prime Maca Phenotypes Grown in Two Geographically-Distant Locations.

PubMed

O Meissner, Henry; Mscisz, Alina; Piatkowska, Ewa; Baraniak, Marek; Mielcarek, Sebastian; Kedzia, Bogdan; Holderna-Kedzia, Elzbieta; Pisulewski, Pawel

2016-03-01

Peruvian Maca crops (Lepidium peruvianum), grown in two geographically-distant cultivation sites located at similar altitudes in the highlands of the Peruvian Andes (Junin at 4,200 m a.s.l. and Ancash 4,150 m a.s.l.), were used in the study. Four prime Maca phenotypes, distinguished by hypocotyl colours labelled as "Yellow", "Purple", "Red" and "Black" were selected to determine distribution in levels and corresponding ratios between individual Glucosinolates (Glucotropaeolin and m-methylglucotropaeolin) in an attempt to identify four Peruvian Maca phenotypes from analyses of powdered hypocotyls. There were highly significant differences (P<0.01) in hypocotyl weight/size of four Maca phenotypes harvested in two locations. The Junin crop represented a mostly "large" class (13.3 g) with "small" size hypocotyls (7.2 g), while a "small" class was predominant in Ancash (3.5 g). Powdered Yellow Maca showed significantly higher (P<0.001) microbial contamination than the other three, with Black Maca being the least infected. Only minor, statistically-confirmed differences were detected in nutritive characteristics between the four Maca phenotypes grown in Junin, however highly significant differences (P<0.01) in Glucosinolates existed between the Red and Black Maca grown in Junin and Ancash. Irrespective of the cultivation location, Red phenotypes showed the highest content of Total Glucosinolates, followed by Black and Purple, with the Yellow phenotype showing consistently lower levels. Highly significant P<0.01) differences determined in ratios of individual Glucosinolates between four Maca phenotypes grown in two locations, confirms an earlier assumption that sums of individual Glucosinolates, their ratios and profiles, may be feasible to explore in analytically identifying individual Maca phenotypes in pulverised marketed Maca products.

Piriformospora indica promotes growth, seed yield and quality of Brassica napus L.

PubMed

Su, Zhen-Zhu; Wang, Ting; Shrivastava, Neeraj; Chen, You-Yuan; Liu, Xiaoxi; Sun, Chao; Yin, Yufeng; Gao, Qi-Kang; Lou, Bing-Gan

2017-06-01

In current scenario, crop productivity is being challenged by decreasing soil fertility. To cope up with this problem, different beneficial microbes are explored to increase the crop productivity with value additions. In this study, Brassica napus L., an important agricultural economic oilseed crop with rich source of nutritive qualities, was interacted with Piriformospora indica, a unique root colonizing fungus with wide host range and multifunctional aspects. The fungus-treated plants showed a significant increase in agronomic parameters with plant biomass, lodging-resistance, early bolting and flowering, oil yield and quality. Nutritional analysis revealed that plants treated by P. indica had reduced erucic acid and glucosinolates contents, and increased the accumulation of N, Ca, Mg, P, K, S, B, Fe and Zn elements. Low erucic acid and glucosinolates contents are important parameters for high quality oil, because oils high in erucic acid and glucosinolates are considered undesirable for human nutrition. Furthermore, the expression profiles of two encoding enzyme genes, Bn-FAE1 and BnECR, which are responsible for regulating erucic acid biosynthesis, were down-regulated at mid- and late- life stages during seeds development in colonized plants. These results demonstrated that P. indica played an important role in enhancing plant growth, rapeseed yield and quality improvement of B. napus. Copyright © 2017 Elsevier GmbH. All rights reserved.

Piriformospora indica Stimulates Root Metabolism of Arabidopsis thaliana.

PubMed

Strehmel, Nadine; Mönchgesang, Susann; Herklotz, Siska; Krüger, Sylvia; Ziegler, Jörg; Scheel, Dierk

2016-07-08

Piriformospora indica is a root-colonizing fungus, which interacts with a variety of plants including Arabidopsis thaliana. This interaction has been considered as mutualistic leading to growth promotion of the host. So far, only indolic glucosinolates and phytohormones have been identified as key players. In a comprehensive non-targeted metabolite profiling study, we analyzed Arabidopsis thaliana's roots, root exudates, and leaves of inoculated and non-inoculated plants by ultra performance liquid chromatography/electrospray ionization quadrupole-time-of-flight mass spectrometry (UPLC/(ESI)-QTOFMS) and gas chromatography/electron ionization quadrupole mass spectrometry (GC/EI-QMS), and identified further biomarkers. Among them, the concentration of nucleosides, dipeptides, oligolignols, and glucosinolate degradation products was affected in the exudates. In the root profiles, nearly all metabolite levels increased upon co-cultivation, like carbohydrates, organic acids, amino acids, glucosinolates, oligolignols, and flavonoids. In the leaf profiles, we detected by far less significant changes. We only observed an increased concentration of organic acids, carbohydrates, ascorbate, glucosinolates and hydroxycinnamic acids, and a decreased concentration of nitrogen-rich amino acids in inoculated plants. These findings contribute to the understanding of symbiotic interactions between plant roots and fungi of the order of Sebacinales and are a valid source for follow-up mechanistic studies, because these symbioses are particular and clearly different from interactions of roots with mycorrhizal fungi or dark septate endophytes.

Evaluating the Antibacterial Properties of Polyacetylene and Glucosinolate Compounds with Further Identification of Their Presence within Various Carrot (Daucus carota) and Broccoli (Brassica oleracea) Cultivars Using High-Performance Liquid Chromatography with a Diode Array Detector and Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry Analyses.

PubMed

Hinds, L; Kenny, O; Hossain, M B; Walsh, D; Sheehy, E; Evans, P; Gaffney, M; Rai, D K

2017-08-23

Ongoing consumer concerns over using synthetic additives in foods has strongly influenced efforts worldwide to source suitable natural alternatives. In this study, the antibacterial efficacy of polyacetylene and glucosinolate compounds was evaluated against both Gram positive and Gram negative bacterial strains. Falcarinol [minimum inhibitory concentration (MIC) = 18.8-37.6 μg/mL] demonstrated the best overall antibacterial activity, while sinigrin (MIC = 46.9-62.5 μg/mL) was the most active glucosinolate compound. High-performance liquid chromatography with a diode array detector analysis showed falcarinol [85.13-244.85 μg/g of dry weight (DW)] to be the most abundant polyacetylene within six of the eight carrot (Daucus carota) cultivars investigated. Meanwhile, sinigrin (100.2-244.3 μg/g of DW) was the most abundant glucosinolate present within the majority of broccoli (Brassica oleracea) cultivars investigated using ultra performance liquid chromatography-tandem mass spectrometry analysis. The high abundance of both falcarinol and sinigrin within these respective species suggests that they could serve as potential sources of natural antibacterial agents for use as such in food products.

Characterization of mustard seeds and paste by DART ionization with time-of-flight mass spectrometry.

PubMed

Prchalová, Jana; Kovařík, František; Ševčík, Rudolf; Čížková, Helena; Rajchl, Aleš

2014-09-01

Direct analysis in real time (DART) is a novel technique with great potential for rapid screening analysis. The DART ionization method coupled with high-resolution time-of-flight mass spectrometry (TOF-MS) has been used for characterization of mustard seeds and table mustard. The possibility to use DART to analyse glucosinolates was confirmed on determination of sinalbin (4-hydroxybenzyl glucosinolate). The DART-TOF-MS method was optimized and validated. A set of samples of mustard seeds and mustard products was analyzed. High-performance liquid chromatography and DART-TOF-MS were used to determine glucosinolates in mustard seeds and compared. The correlation equation between these methods was DART = 0.797*HPLC + 6.987, R(2)  = 0.972. The DART technique seems to be a suitable method for evaluation of the quality of mustard seeds and mustard products. Copyright © 2014 John Wiley & Sons, Ltd.

Glucosinolates, myrosinase hydrolysis products, and flavonols found in rocket (Eruca sativa and Diplotaxis tenuifolia).

PubMed

Bell, Luke; Wagstaff, Carol

2014-05-21

Rocket species have been shown to have very high concentrations of glucosinolates and flavonols, which have numerous positive health benefits with regular consumption. This review highlights how breeders and processors of rocket species can utilize genomic and phytochemical research to improve varieties and enhance the nutritive benefits to consumers. Plant breeders are increasingly looking to new technologies such as HPLC, UPLC, LC-MS, and GC-MS to screen populations for their phytochemical content to inform plant selections. This paper collates the research that has been conducted to date in rocket and summarizes all glucosinolate and flavonol compounds identified in the species. The paper emphasizes the importance of the broad screening of populations for phytochemicals and myrosinase degradation products, as well as unique traits that may be found in underutilized gene bank resources. This review also stresses that collaboration with industrial partners is becoming essential for long-term plant breeding goals through research.

Quantitative proteomics reveals a role of JAZ7 in plant defense response to Pseudomonas syringae DC3000.

PubMed

Zhang, Tong; Meng, Li; Kong, Wenwen; Yin, Zepeng; Wang, Yang; Schneider, Jacqueline D; Chen, Sixue

2018-03-20

Jasmonate ZIM-domain (JAZ) proteins are key transcriptional repressors regulating various biological processes. Although many studies have studied JAZ proteins by genetic and biochemical analyses, little is known about JAZ7-associated global protein networks and how JAZ7 contributes to bacterial pathogen defense. In this study, we aim to fill this knowledge gap by conducting unbiased large-scale quantitative proteomics using tandem mass tags (TMT). We compared the proteomes of a JAZ7 knock-out line, a JAZ7 overexpression line, as well as the wild type Arabidopsis plants in the presence and absence of Pseudomonas syringae DC3000 infection. Both pairwise comparison and multi-factor analysis of variance reveal that differential proteins are enriched in biological processes such as primary and secondary metabolism, redox regulation, and response to stress. The differential regulation in these pathways may account for the alterations in plant size, redox homeostasis and accumulation of glucosinolates. In addition, possible interplay between genotype and environment is suggested as the abundance of seven proteins is influenced by the interaction of the two factors. Collectively, we demonstrate a role of JAZ7 in pathogen defense and provide a list of proteins that are uniquely responsive to genetic disruption, pathogen infection, or the interaction between genotypes and environmental factors. We report proteomic changes as a result of genetic perturbation of JAZ7, and the contribution of JAZ7 in plant immunity. Specifically, the similarity between the proteomes of a JAZ7 knockout mutant and the wild type plants confirmed the functional redundancy of JAZs. In contrast, JAZ7 overexpression plants were much different, and proteomic analysis of the JAZ7 overexpression plants under Pst DC3000 infection revealed that JAZ7 may regulate plant immunity via ROS modulation, energy balance and glucosinolate biosynthesis. Multiple variate analysis for this two-factor proteomics experiment suggests that protein abundance is determined by genotype, environment and the interaction between them. Copyright © 2018 Elsevier B.V. All rights reserved.

Endophytic fungi from the roots of horseradish (Armoracia rusticana) and their interactions with the defensive metabolites of the glucosinolate - myrosinase - isothiocyanate system.

PubMed

Szűcs, Zsolt; Plaszkó, Tamás; Cziáky, Zoltán; Kiss-Szikszai, Attila; Emri, Tamás; Bertóti, Regina; Sinka, László Tamás; Vasas, Gábor; Gonda, Sándor

2018-05-09

The health of plants is heavily influenced by the intensively researched plant microbiome. The microbiome has to cope with the plant's defensive secondary metabolites to survive and develop, but studies that describe this interaction are rare. In the current study, we describe interactions of endophytic fungi with a widely researched chemical defense system, the glucosinolate - myrosinase - isothiocyanate system. The antifungal isothiocyanates are also of special interest because of their beneficial effects on human consumers. Seven endophytic fungi were isolated from horseradish roots (Armoracia rusticana), from the genera Fusarium, Macrophomina, Setophoma, Paraphoma and Oidiodendron. LC-ESI-MS analysis of the horseradish extract incubated with these fungi showed that six of seven strains could decompose different classes of glucosinolates. Aliphatic, aromatic, thiomethylalkyl and indolic glucosinolates were decomposed by different strains at different rates. SPME-GC-MS measurements showed that two strains released significant amounts of allyl isothiocyanate into the surrounding air, but allyl nitrile was not detected. The LC-ESI-MS analysis of many strains' media showed the presence of allyl isothiocyanate - glutathione conjugate during the decomposition of sinigrin. Four endophytic strains also accepted sinigrin as the sole carbon source. Isothiocyanates inhibited the growth of fungi at various concentrations, phenylethyl isothiocyanate was more potent than allyl isothiocyanate (mean IC 50 was 2.30-fold lower). As a control group, ten soil fungi from the same soil were used. They decomposed glucosinolates with lower overall efficiency: six of ten strains had insignificant or weak activities and only three could use sinigrin as a carbon source. The soil fungi also showed lower AITC tolerance in the growth inhibition assay: the median IC 50 values were 0.1925 mM for endophytes and 0.0899 mM for soil fungi. The host's glucosinolates can be used by the tested endophytic fungi as nutrients or to gain competitive advantage over less tolerant species. These activities were much less apparent among the soil fungi. This suggests that the endophytes show adaptation to the host plant's secondary metabolites and that host metabolite specific activities are enriched in the root microbiome. The results present background mechanisms enabling an understanding of how plants shape their microbiome.

Red maca (Lepidium meyenii) reduced prostate size in rats

PubMed Central

Gonzales, Gustavo F; Miranda, Sara; Nieto, Jessica; Fernández, Gilma; Yucra, Sandra; Rubio, Julio; Yi, Pedro; Gasco, Manuel

2005-01-01

Background Epidemiological studies have found that consumption of cruciferous vegetables is associated with a reduced risk of prostate cancer. This effect seems to be due to aromatic glucosinolate content. Glucosinolates are known for have both antiproliferative and proapoptotic actions. Maca is a cruciferous cultivated in the highlands of Peru. The absolute content of glucosinolates in Maca hypocotyls is relatively higher than that reported in other cruciferous crops. Therefore, Maca may have proapoptotic and anti-proliferative effects in the prostate. Methods Male rats treated with or without aqueous extracts of three ecotypes of Maca (Yellow, Black and Red) were analyzed to determine the effect on ventral prostate weight, epithelial height and duct luminal area. Effects on serum testosterone (T) and estradiol (E2) levels were also assessed. Besides, the effect of Red Maca on prostate was analyzed in rats treated with testosterone enanthate (TE). Results Red Maca but neither Yellow nor Black Maca reduced significantly ventral prostate size in rats. Serum T or E2 levels were not affected by any of the ecotypes of Maca assessed. Red Maca also prevented the prostate weight increase induced by TE treatment. Red Maca administered for 42 days reduced ventral prostatic epithelial height. TE increased ventral prostatic epithelial height and duct luminal area. These increases by TE were reduced after treatment with Red Maca for 42 days. Histology pictures in rats treated with Red Maca plus TE were similar to controls. Phytochemical screening showed that aqueous extract of Red Maca has alkaloids, steroids, tannins, saponins, and cardiotonic glycosides. The IR spectra of the three ecotypes of Maca in 3800-650 cm (-1) region had 7 peaks representing 7 functional chemical groups. Highest peak values were observed for Red Maca, intermediate values for Yellow Maca and low values for Black Maca. These functional groups correspond among others to benzyl glucosinolate. Conclusions Red Maca, a cruciferous plant from the highland of Peru, reduced ventral prostate size in normal and TE treated rats. PMID:15661081

Entry Mode–Dependent Function of an Indole Glucosinolate Pathway in Arabidopsis for Nonhost Resistance against Anthracnose Pathogens[W

PubMed Central

Hiruma, Kei; Onozawa-Komori, Mariko; Takahashi, Fumika; Asakura, Makoto; Bednarek, Paweł; Okuno, Tetsuro; Schulze-Lefert, Paul; Takano, Yoshitaka

2010-01-01

When faced with nonadapted fungal pathogens, Arabidopsis thaliana mounts nonhost resistance responses, which typically result in the termination of early pathogenesis steps. We report that nonadapted anthracnose fungi engage two alternative entry modes during pathogenesis on leaves: turgor-mediated invasion beneath melanized appressoria, and a previously undiscovered hyphal tip–based entry (HTE) that is independent of appressorium formation. The frequency of HTE is positively regulated by carbohydrate nutrients and appears to be subject to constitutive inhibition by the fungal mitogen-activated protein kinase (MAPK) cascade of MAPK ESSENTIAL FOR APPRESSORIUM FORMATION1. The same MAPK cascade is essential for appressorium formation. Unexpectedly, the Arabidopsis indole glucosinolate pathway restricts entry of the nonadapted anthracnose fungi only when these pathogens employ HTE. Arabidopsis mutants defective in indole glucosinolate biosynthesis or metabolism support the initiation of postinvasion growth of nonadapted Colletotrichum gloeosporioides and Colletotrichum orbiculare. However, genetic disruption of Colletotrichum appressorium formation does not permit HTE on host plants. Thus, Colletotrichum appressoria play a critical role in the suppression of preinvasion plant defenses, in addition to their previously described role in turgor-mediated plant cell invasion. We also show that HTE is the predominant morphogenetic response of Colletotrichum at wound sites. This implies the existence of a fungal sensing system to trigger appropriate morphogenetic responses during pathogenesis at wound sites and on intact leaf tissue. PMID:20605856

Effects of Combined Low Glutathione with Mild Oxidative and Low Phosphorus Stress on the Metabolism of Arabidopsis thaliana

PubMed Central

Fukushima, Atsushi; Iwasa, Mami; Nakabayashi, Ryo; Kobayashi, Makoto; Nishizawa, Tomoko; Okazaki, Yozo; Saito, Kazuki; Kusano, Miyako

2017-01-01

Plants possess highly sensitive mechanisms that monitor environmental stress levels for a dose-dependent fine-tuning of their growth and development. Differences in plant responses to severe and mild abiotic stresses have been recognized. Although many studies have revealed that glutathione can contribute to plant tolerance to various environmental stresses, little is known about the relationship between glutathione and mild abiotic stress, especially the effect of stress-induced altered glutathione levels on the metabolism. Here, we applied a systems biology approach to identify key pathways involved in the gene-to-metabolite networks perturbed by low glutathione content under mild abiotic stress in Arabidopsis thaliana. We used glutathione synthesis mutants (cad2-1 and pad2-1) and plants overexpressing the gene encoding γ-glutamylcysteine synthetase, the first enzyme of the glutathione biosynthetic pathway. The plants were exposed to two mild stress conditions—oxidative stress elicited by methyl viologen and stress induced by the limited availability of phosphate. We observed that the mutants and transgenic plants showed similar shoot growth as that of the wild-type plants under mild abiotic stress. We then selected the synthesis mutants and performed multi-platform metabolomics and microarray experiments to evaluate the possible effects on the overall metabolome and the transcriptome. As a common oxidative stress response, several flavonoids that we assessed showed overaccumulation, whereas the mild phosphate stress resulted in increased levels of specific kaempferol- and quercetin-glycosides. Remarkably, in addition to a significant increased level of sugar, osmolytes, and lipids as mild oxidative stress-responsive metabolites, short-chain aliphatic glucosinolates over-accumulated in the mutants, whereas the level of long-chain aliphatic glucosinolates and specific lipids decreased. Coordinated gene expressions related to glucosinolate and flavonoid biosynthesis also supported the metabolite responses in the pad2-1 mutant. Our results suggest that glutathione synthesis mutants accelerate transcriptional regulatory networks to control the biosynthetic pathways involved in glutathione-independent scavenging metabolites, and that they might reconfigure the metabolic networks in primary and secondary metabolism, including lipids, glucosinolates, and flavonoids. This work provides a basis for the elucidation of the molecular mechanisms involved in the metabolic and transcriptional regulatory networks in response to combined low glutathione content with mild oxidative and nutrient stress in A. thaliana. PMID:28894456

Metabolic Changes during Storage of Brassica napus Seeds under Moist Conditions and the Consequences for the Sensory Quality of the Resulting Virgin Oil.

PubMed

Bonte, Anja; Schweiger, Rabea; Pons, Caroline; Wagner, Claudia; Brühl, Ludger; Matthäus, Bertrand; Müller, Caroline

2017-12-20

Virgin rapeseed (Brassica napus) oil is a valuable niche product, if delivered with a high quality. In this study, the effects of moist storage of B. napus seeds for 1 to 4 days on the seed metabolome and the chemo-sensory properties of the produced oils were determined. The concentrations of several primary metabolites, including monosaccharides and amino acids, rapidly increased in the seeds, probably indicating the breakdown of storage compounds to support seed germination. Seed concentrations of indole glucosinolates increased with a slight time offset suggesting that amino acids may be used to modify secondary metabolism. The volatile profiles of the oils were pronouncedly influenced by moist seed storage, with the sensory quality of the oils decreasing. This study provides a direct time-resolved link between seed metabolism under moist conditions and the quality of the resulting oils, thereby emphasizing the crucial role of dry seed storage in ensuring high oil quality.

21 CFR 573.310 - Crambe meal, heat toasted.

Code of Federal Regulations, 2012 CFR

2012-04-01

...; oil, not more than 4 percent; crude protein, not less than 24 percent; crude fiber, not more than 26 percent; glucosinolate calculated as epi-progoitrin, not more than 4 percent; goitrin, not more than 0.1 percent; nitrile calculated as 1-cyano-2-hydroxy-3-butene, not more than 1.4 percent. At least 50 percent...

21 CFR 573.310 - Crambe meal, heat toasted.

Code of Federal Regulations, 2010 CFR

2010-04-01

...; oil, not more than 4 percent; crude protein, not less than 24 percent; crude fiber, not more than 26 percent; glucosinolate calculated as epi-progoitrin, not more than 4 percent; goitrin, not more than 0.1 percent; nitrile calculated as 1-cyano-2-hydroxy-3-butene, not more than 1.4 percent. At least 50 percent...

21 CFR 573.310 - Crambe meal, heat toasted.

Code of Federal Regulations, 2013 CFR

2013-04-01

...; oil, not more than 4 percent; crude protein, not less than 24 percent; crude fiber, not more than 26 percent; glucosinolate calculated as epi-progoitrin, not more than 4 percent; goitrin, not more than 0.1 percent; nitrile calculated as 1-cyano-2-hydroxy-3-butene, not more than 1.4 percent. At least 50 percent...

21 CFR 573.310 - Crambe meal, heat toasted.

Code of Federal Regulations, 2014 CFR

2014-04-01

...; oil, not more than 4 percent; crude protein, not less than 24 percent; crude fiber, not more than 26 percent; glucosinolate calculated as epi-progoitrin, not more than 4 percent; goitrin, not more than 0.1 percent; nitrile calculated as 1-cyano-2-hydroxy-3-butene, not more than 1.4 percent. At least 50 percent...

21 CFR 573.310 - Crambe meal, heat toasted.

Code of Federal Regulations, 2011 CFR

2011-04-01

...; oil, not more than 4 percent; crude protein, not less than 24 percent; crude fiber, not more than 26 percent; glucosinolate calculated as epi-progoitrin, not more than 4 percent; goitrin, not more than 0.1 percent; nitrile calculated as 1-cyano-2-hydroxy-3-butene, not more than 1.4 percent. At least 50 percent...

Broccoli sprouts: An exceptionally rich source of inducers of enzymes that protect against chemical carcinogens

PubMed Central

Fahey, Jed W.; Zhang, Yuesheng; Talalay, Paul

1997-01-01

Induction of phase 2 detoxication enzymes [e.g., glutathione transferases, epoxide hydrolase, NAD(P)H: quinone reductase, and glucuronosyltransferases] is a powerful strategy for achieving protection against carcinogenesis, mutagenesis, and other forms of toxicity of electrophiles and reactive forms of oxygen. Since consumption of large quantities of fruit and vegetables is associated with a striking reduction in the risk of developing a variety of malignancies, it is of interest that a number of edible plants contain substantial quantities of compounds that regulate mammalian enzymes of xenobiotic metabolism. Thus, edible plants belonging to the family Cruciferae and genus Brassica (e.g., broccoli and cauliflower) contain substantial quantities of isothiocyanates (mostly in the form of their glucosinolate precursors) some of which (e.g., sulforaphane or 4-methylsulfinylbutyl isothiocyanate) are very potent inducers of phase 2 enzymes. Unexpectedly, 3-day-old sprouts of cultivars of certain crucifers including broccoli and cauliflower contain 10–100 times higher levels of glucoraphanin (the glucosinolate of sulforaphane) than do the corresponding mature plants. Glucosinolates and isothiocyanates can be efficiently extracted from plants, without hydrolysis of glucosinolates by myrosinase, by homogenization in a mixture of equal volumes of dimethyl sulfoxide, dimethylformamide, and acetonitrile at −50°C. Extracts of 3-day-old broccoli sprouts (containing either glucoraphanin or sulforaphane as the principal enzyme inducer) were highly effective in reducing the incidence, multiplicity, and rate of development of mammary tumors in dimethylbenz(a)anthracene-treated rats. Notably, sprouts of many broccoli cultivars contain negligible quantities of indole glucosinolates, which predominate in the mature vegetable and may give rise to degradation products (e.g., indole-3-carbinol) that can enhance tumorigenesis. Hence, small quantities of crucifer sprouts may protect against the risk of cancer as effectively as much larger quantities of mature vegetables of the same variety. PMID:9294217

Comparison of Glucosinolate Profiles in Different Tissues of Nine Brassica Crops.

PubMed

Bhandari, Shiva Ram; Jo, Jung Su; Lee, Jun Gu

2015-08-31

Glucosinolate (GSL) profiles and concentrations in various tissues (seeds, sprouts, mature root, and shoot) were determined and compared across nine Brassica species, including cauliflower, cabbage, broccoli, radish, baemuchae, pakchoi, Chinese cabbage, leaf mustard, and kale. The compositions and concentrations of individual GSLs varied among crops, tissues, and growth stages. Seeds had highest total GSL concentrations in most of crops, whereas shoots had the lowest GSL concentrations. Aliphatic GSL concentrations were the highest in seeds, followed by that in sprouts, shoots, and roots. Indole GSL concentration was the highest in the root or shoot tissues in most of the crops. In contrast, aromatic GSL concentrations were highest in roots. Of the nine crops examined, broccoli exhibited the highest total GSL concentration in seeds (110.76 µmol·g(-1)) and sprouts (162.19 µmol·g(-1)), whereas leaf mustard exhibited the highest total GSL concentration in shoots (61.76 µmol·g(-1)) and roots (73.61 µmol·g(-1)). The lowest GSL concentrations were observed in radish across all tissues examined.

[The role of glucosinolates in the prevention of cancer--mechanisms of actions].

PubMed

Kwiatkowska, Edyta; Bawa, Sa'eed

2007-01-01

Foods of plant origin, despite plenty of nutrients contain many non-nutrition compounds, which may prevent many diet-related non-communicable diseases, such as cancer. Plants produce thousands of phenolic compounds as secondary metabolites, such as nitrous compounds. Glucosinolates are responsible for the secretion of detoxifying enzymes that remove carcinogens for the organism. Furthermore, they activate proteins and II phase detoxifying enzymes. The compounds are very important that is why scientists are still investigating their beneficial note in cancer prevention and management.

Transcriptome analysis and metabolic profiling of green and red kale (Brassica oleracea var. acephala) seedlings.

PubMed

Jeon, Jin; Kim, Jae Kwang; Kim, HyeRan; Kim, Yeon Jeong; Park, Yun Ji; Kim, Sun Ju; Kim, Changsoo; Park, Sang Un

2018-02-15

Kale (Brassica oleracea var. acephala) is a rich source of numerous health-benefiting compounds, including vitamins, glucosinolates, phenolic compounds, and carotenoids. However, the genetic resources for exploiting the phyto-nutritional traits of kales are limited. To acquire precise information on secondary metabolites in kales, we performed a comprehensive analysis of the transcriptome and metabolome of green and red kale seedlings. Kale transcriptome datasets revealed 37,149 annotated genes and several secondary metabolite biosynthetic genes. HPLC analysis revealed 14 glucosinolates, 20 anthocyanins, 3 phenylpropanoids, and 6 carotenoids in the kale seedlings that were examined. Red kale contained more glucosinolates, anthocyanins, and phenylpropanoids than green kale, whereas the carotenoid contents were much higher in green kale than in red kale. Ultimately, our data will be a valuable resource for future research on kale bio-engineering and will provide basic information to define gene-to-metabolite networks in kale. Copyright © 2017 Elsevier Ltd. All rights reserved.

Distribution of selenoglucosinolates and their metabolites in Brassica treated with sodium selenate.

PubMed

Matich, Adam J; McKenzie, Marian J; Lill, Ross E; McGhie, Tony K; Chen, Ronan K-Y; Rowan, Daryl D

2015-02-25

In Brassica species, hydrolysis of (methylthio)glucosinolates produces sulfur-containing aglycons which have demonstrated anticancer benefits. Selenized Brassicaceae contain (methylseleno)glucosinolates and their selenium-containing aglycons. As a prelude to biological testing, broccoli, cauliflower, and forage rape plants were treated with sodium selenate and their tap roots, stems, leaves, and florets analyzed for selenoglucosinolates and their Se aglycons. Two new selenoglucosinolates were identified: glucoselenoraphanin in broccoli florets and glucoselenonasturtiin in forage rape roots. A new aglycon, selenoberteroin nitrile, was identified in forage rape. The major selenoglucosinolates were glucoselenoerucin in broccoli, glucoselenoiberverin in cauliflower, and glucoselenoerucin and glucoselenoberteroin in forage rape roots. In broccoli florets, the concentrations of selenglucosinolates exceeded those of their sulfur analogues. Fertilization with selenium slightly reduced (methylthio)glucosinolates and aglycons in the roots, but increased them in the florets, the leaves, and sometimes the stems. These discoveries provide a new avenue for investigating how consumption of Brassica vegetables and their organoselenides may promote human health.

Protein-accumulating cells and dilated cisternae of the endoplasmic reticulum in three glucosinolate-containing genera: Armoracia, Capparis, Drypetes.

PubMed

Jørgensen, L B; Behnke, H D; Mabry, T J

1977-01-01

Three glucosinolate-containing species, Armoracia rusticana Gaertner, Meyer et Scherbius (Brassicaceae), Capparis cynophallophora L. (Capparaceae) and Drypetes roxburghii (Wall.) Hurusawa (Euphorbiaceae), are shown by both light and electron microscopy to contain protein-accumulating cells (PAC). The PAC of Armoracia and Copparis (former "myrosin cells") occur as idioblasts. The PAC of Drypetes are usual members among axial phloem parenchyma cells rather than idioblasts. In Drypetes the vacuoles of the PAC are shown ultrastructurally to contain finely fibrillar material and to originate from local dilatations of the endoplasmic reticulum. The vacuoles in PAC of Armoracia and Capparis seem to originate in the same way; but ultrastructurally, their content is finely granular. In addition, Armoracia and Capparis are shown by both light and electron microscopy to contain dilated cisternae (DC) of the endoplasmic reticulum in normal parenchyma cells, in accord with previous findings for several species within Brassicaceae. The relationship of PAC and DC to glucosinolates and the enzyme myrosinase is discussed.

Analysis of seven salad rocket (Eruca sativa) accessions: The relationships between sensory attributes and volatile and non-volatile compounds.

PubMed

Bell, Luke; Methven, Lisa; Signore, Angelo; Oruna-Concha, Maria Jose; Wagstaff, Carol

2017-03-01

Sensory and chemical analyses were performed on accessions of rocket (Eruca sativa) to determine phytochemical influences on sensory attributes. A trained panel was used to evaluate leaves, and chemical data were obtained for polyatomic ions, amino acids, sugars and organic acids. These chemical data (and data of glucosinolates, flavonols and headspace volatiles previously reported) were used in Principal Component Analysis (PCA) to determine variables statistically important to sensory traits. Significant differences were observed between samples for polyatomic ion and amino acid concentrations. PCA revealed strong, positive correlations between glucosinolates, isothiocyanates and sulfur compounds with bitterness, mustard, peppery, warming and initial heat mouthfeel traits. The ratio between glucosinolates and sugars inferred reduced perception of bitter aftereffects. We highlight the diversity of E. sativa accessions from a sensory and phytochemical standpoint, and the potential for breeders to create varieties that are nutritionally and sensorially superior to existing ones. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Influence of day length and temperature on the content of health-related compounds in broccoli (Brassica oleracea L. var. italica).

PubMed

Steindal, Anne Linn Hykkerud; Mølmann, Jørgen; Bengtsson, Gunnar B; Johansen, Tor J

2013-11-13

Vegetables grown at different latitudes are exposed to various temperatures and day lengths, which can affect the content of health- and sensory-related compounds in broccoli florets. A 2 × 2 factorial experiment was conducted under controlled growth conditions, with contrasting temperatures (15/9 and 21/15 °C) and day lengths (12 and 24 h), to investigate the effect on glucosinolates, vitamin C, flavonols, and soluble sugars. Aliphatic glucosinolates, quercetin, and kaempferol were at their highest levels at high temperatures combined with a 12 h day. Levels of total glucosinolates, d-glucose, and d-fructose were elevated by high temperatures. Conversely, the content of vitamin C was highest with a 12 h day length combined with 15/9 °C. Our results indicate that temperature and day length influence the contents of health-related compounds in broccoli florets in a complex way, suggesting no general superiority of any of the contrasting growth conditions.

Selenium Biofortification in Radish Enhances Nutritional Quality via Accumulation of Methyl-Selenocysteine and Promotion of Transcripts and Metabolites Related to Glucosinolates, Phenolics, and Amino Acids.

PubMed

Schiavon, Michela; Berto, Chiara; Malagoli, Mario; Trentin, Annarita; Sambo, Paolo; Dall'Acqua, Stefano; Pilon-Smits, Elizabeth A H

2016-01-01

Two selenium (Se) fertilization methods were tested for their effects on levels of anticarcinogenic selenocompounds in radish (Raphanus sativus), as well as other nutraceuticals. First, radish was grown on soil and foliar selenate applied 7 days before harvest at 0, 5, 10, and 20 mg Se per plant. Selenium levels were up to 1200 mg Se/kg DW in leaves and 120 mg Se/kg DW in roots. The thiols cysteine and glutathione were present at 2-3-fold higher levels in roots of Se treated plants, and total glucosinolate levels were 35% higher, due to increases in glucoraphanin. The only seleno-aminoacid detected in Se treated plants was Se-methyl-SeCys (100 mg/kg FW in leaves, 33 mg/kg FW in roots). The levels of phenolic aminoacids increased with selenate treatment, as did root total nitrogen and protein content, while the level of several polyphenols decreased. Second, radish was grown in hydroponics and supplied with 0, 5, 10, 20, or 40 μM selenate for 1 week. Selenate treatment led to a 20-30% increase in biomass. Selenium concentration was 242 mg Se/kg DW in leaves and 85 mg Se/kg DW in roots. Cysteine levels decreased with Se in leaves but increased in roots; glutatione levels decreased in both. Total glucosinolate levels in leaves decreased with Se treatment due to repression of genes involved in glucosinolates metabolism. Se-methyl-SeCys concentration ranged from 7-15 mg/kg FW. Aminoacid concentration increased with Se treatment in leaves but decreased in roots. Roots of Se treated plants contained elevated transcript levels of sulfate transporters (Sultr) and ATP sulfurylase, a key enzyme of S/Se assimilation. No effects on polyphenols were observed. In conclusion, Se biofortification of radish roots may be achieved via foliar spray or hydroponic supply. One to ten radishes could fulfill the daily human requirement (70 μg) after a single foliar spray of 5 mg selenate per plant or 1 week of 5-10 μM selenate supply in hydroponics. The radishes metabolized selenate to the anticarcinogenic compound Se-methyl-selenocysteine. Selenate treatment enhanced levels of other nutraceuticals in radish roots, including glucoraphanin. Therefore, Se biofortification can produce plants with superior health benefits.

Selenium Biofortification in Radish Enhances Nutritional Quality via Accumulation of Methyl-Selenocysteine and Promotion of Transcripts and Metabolites Related to Glucosinolates, Phenolics, and Amino Acids

PubMed Central

Schiavon, Michela; Berto, Chiara; Malagoli, Mario; Trentin, Annarita; Sambo, Paolo; Dall'Acqua, Stefano; Pilon-Smits, Elizabeth A. H.

2016-01-01

Two selenium (Se) fertilization methods were tested for their effects on levels of anticarcinogenic selenocompounds in radish (Raphanus sativus), as well as other nutraceuticals. First, radish was grown on soil and foliar selenate applied 7 days before harvest at 0, 5, 10, and 20 mg Se per plant. Selenium levels were up to 1200 mg Se/kg DW in leaves and 120 mg Se/kg DW in roots. The thiols cysteine and glutathione were present at 2–3-fold higher levels in roots of Se treated plants, and total glucosinolate levels were 35% higher, due to increases in glucoraphanin. The only seleno-aminoacid detected in Se treated plants was Se-methyl-SeCys (100 mg/kg FW in leaves, 33 mg/kg FW in roots). The levels of phenolic aminoacids increased with selenate treatment, as did root total nitrogen and protein content, while the level of several polyphenols decreased. Second, radish was grown in hydroponics and supplied with 0, 5, 10, 20, or 40 μM selenate for 1 week. Selenate treatment led to a 20–30% increase in biomass. Selenium concentration was 242 mg Se/kg DW in leaves and 85 mg Se/kg DW in roots. Cysteine levels decreased with Se in leaves but increased in roots; glutatione levels decreased in both. Total glucosinolate levels in leaves decreased with Se treatment due to repression of genes involved in glucosinolates metabolism. Se-methyl-SeCys concentration ranged from 7–15 mg/kg FW. Aminoacid concentration increased with Se treatment in leaves but decreased in roots. Roots of Se treated plants contained elevated transcript levels of sulfate transporters (Sultr) and ATP sulfurylase, a key enzyme of S/Se assimilation. No effects on polyphenols were observed. In conclusion, Se biofortification of radish roots may be achieved via foliar spray or hydroponic supply. One to ten radishes could fulfill the daily human requirement (70 μg) after a single foliar spray of 5 mg selenate per plant or 1 week of 5–10 μM selenate supply in hydroponics. The radishes metabolized selenate to the anticarcinogenic compound Se-methyl-selenocysteine. Selenate treatment enhanced levels of other nutraceuticals in radish roots, including glucoraphanin. Therefore, Se biofortification can produce plants with superior health benefits. PMID:27683583

Real-time quantitative polymerase chain reaction methods for four genetically modified maize varieties and maize DNA content in food.

PubMed

Brodmann, Peter D; Ilg, Evelyn C; Berthoud, Hélène; Herrmann, Andre

2002-01-01

Quantitative detection methods are needed for enforcement of the recently introduced labeling threshold for genetically modified organisms (GMOs) in food ingredients. This labeling threshold, which is set to 1% in the European Union and Switzerland, must be applied to all approved GMOs. Four different varieties of maize are approved in the European Union: the insect-resistant Bt176 maize (Maximizer), Btl 1 maize, Mon810 (YieldGard) maize, and the herbicide-tolerant T25 (Liberty Link) maize. Because the labeling must be considered individually for each ingredient, a quantitation system for the endogenous maize content is needed in addition to the GMO-specific detection systems. Quantitative real-time polymerase chain reaction detection methods were developed for the 4 approved genetically modified maize varieties and for an endogenous maize (invertase) gene system.

How Different Genetically Manipulated Brassica Genotypes Affect Life Table Parameters of Plutella xylostella (Lepidoptera: Plutellidae).

PubMed

Nikooei, Mehrnoosh; Fathipour, Yaghoub; Jalali Javaran, Mokhtar; Soufbaf, Mahmoud

2015-04-01

The fitness of Plutella xylostella L. on different genetically manipulated Brassica plants, including canola's progenitor (Brassica rapa L.), two cultivated canola cultivars (Opera and RGS003), one hybrid (Hyula401), one gamma-ray mutant-RGS003, and one transgenic (PF) genotype was compared using two-sex and female-based life table parameters. All experiments were conducted in a growth chamber at 25±1°C, 65±5% relative humidity, and a photoperiod of 16:8 (L:D) h. There were significant differences in duration of different life stages of P. xylostella on different plant genotypes. The shortest (13.92 d) and longest (24.61 d) total developmental time were on Opera and PF, respectively. The intrinsic rate of increase of P. xylostella ranged between 0.236 (Opera) and 0.071 day(-1) (PF). The highest (60.79 offspring) and lowest (7.88 offspring) net reproductive rates were observed on Opera and PF, respectively. Comparison of intrinsic rate of increase, net reproductive rates, finite rate of increase, mean generation time, fecundity, and survivorship of P. xylostella on the plant genotypes suggested that this pest performed well on cultivars (RGS003 and Opera) and performed poorly on the other manipulated genotypes especially on mutant-RGS003 and PF. Glucosinolate levels were significantly higher in damaged plants than undamaged ones and the lowest and highest concentrations of glucosinolates were found in transgenic genotype and canola's progenitor, respectively. Interestingly, our results showed that performance and fitness of this pest was better on canola's progenitor and cultivated plants, which had high levels of glucosinolate. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Water Stress and Aphid Feeding Differentially Influence Metabolite Composition in Arabidopsis thaliana (L.)

PubMed Central

Mewis, Inga; Khan, Mohammed A. M.; Glawischnig, Erich; Schreiner, Monika; Ulrichs, Christian

2012-01-01

Little is known about how drought stress influences plant secondary metabolite accumulation and how this affects plant defense against different aphids. We therefore cultivated Arabidopsis thaliana (L.) plants under well-watered, drought, and water-logged conditions. Two aphid species were selected for this study: the generalist Myzus persicae (Sulzer) and the crucifer specialist Brevicoryne brassicae (L.). Metabolite concentrations in the phloem sap, which influence aphid growth, changed particularly under drought stress. Levels of sucrose and several amino acids, such as glutamic acid, proline, isoleucine, and lysine increased, while concentrations of 4-methoxyindol-3-ylmethyl glucosinolate decreased. M. persicae population growth was highest on plants under drought stress conditions. However, B. brassicae did not profit from improved phloem sap quality under drought stress and performed equally in all water treatments. Water stress and aphids generally had an opposite effect on the accumulation of secondary metabolites in the plant rosettes. Drought stress and water-logging led to increased aliphatic glucosinolate and flavonoid levels. Conversely, aphid feeding, especially of M. persicae, reduced levels of flavonoids and glucosinolates in the plants. Correspondingly, transcript levels of aliphatic biosynthetic genes decreased after feeding of both aphid species. Contrary to M. persicae, drought stress did not promote population growth of B. brassicae on these plants. The specialist aphid induced expression of CYP79B2, CYP79B3, and PAD3 with corresponding accumulation of indolyl glucosinolates and camalexin. This was distinct from M. persicae, which did not elicit similarly strong camalexin accumulation, which led to the hypothesis of a specific defense adaptations against the specialist aphid. PMID:23144921

The Defense Metabolite, Allyl Glucosinolate, Modulates Arabidopsis thaliana Biomass Dependent upon the Endogenous Glucosinolate Pathway

PubMed Central

Francisco, Marta; Joseph, Bindu; Caligagan, Hart; Li, Baohua; Corwin, Jason A.; Lin, Catherine; Kerwin, Rachel; Burow, Meike; Kliebenstein, Daniel J.

2016-01-01

Glucosinolates (GSLs) play an important role in plants as direct mediators of biotic and abiotic stress responses. Recent work is beginning to show that the GSLs can also inducing complex defense and growth networks. However, the physiological significance of these GSL-induced responses and the molecular mechanisms by which GSLs are sensed and/or modulate these responses are not understood. To identify these potential mechanisms within the plant and how they may relate to the endogenous GSLs, we tested the regulatory effect of exogenous allyl GSL application on growth and defense metabolism across sample of Arabidopsis thaliana accessions. We found that application of exogenous allyl GSL had the ability to initiate changes in plant biomass and accumulation of defense metabolites that genetically varied across accessions. This growth effect was related to the allyl GSL side-chain structure. Utilizing this natural variation and mutants in genes within the GSL pathway we could show that the link between allyl GSL and altered growth responses are dependent upon the function of known genes controlling the aliphatic GSL pathway. PMID:27313596

The Defense Metabolite, Allyl Glucosinolate, Modulates Arabidopsis thaliana Biomass Dependent upon the Endogenous Glucosinolate Pathway.

PubMed

Francisco, Marta; Joseph, Bindu; Caligagan, Hart; Li, Baohua; Corwin, Jason A; Lin, Catherine; Kerwin, Rachel; Burow, Meike; Kliebenstein, Daniel J

2016-01-01

Glucosinolates (GSLs) play an important role in plants as direct mediators of biotic and abiotic stress responses. Recent work is beginning to show that the GSLs can also inducing complex defense and growth networks. However, the physiological significance of these GSL-induced responses and the molecular mechanisms by which GSLs are sensed and/or modulate these responses are not understood. To identify these potential mechanisms within the plant and how they may relate to the endogenous GSLs, we tested the regulatory effect of exogenous allyl GSL application on growth and defense metabolism across sample of Arabidopsis thaliana accessions. We found that application of exogenous allyl GSL had the ability to initiate changes in plant biomass and accumulation of defense metabolites that genetically varied across accessions. This growth effect was related to the allyl GSL side-chain structure. Utilizing this natural variation and mutants in genes within the GSL pathway we could show that the link between allyl GSL and altered growth responses are dependent upon the function of known genes controlling the aliphatic GSL pathway.

LC-MS profiling of glucosinolates in the seeds of Brassica elongata Ehrh., and of the two stenoendemic B. botteri Vis and B. cazzae Ginzb. & Teyber.

PubMed

Montaut, Sabine; Blažević, Ivica; Ruščić, Mirko; Rollin, Patrick

2017-01-01

The glucosinolates (GLs) present in seed extracts of Brassica elongata Ehrh., B. botteri Vis and B. cazzae Ginzb. & Teyber from Croatia were identified by LC-MS. 4-Hydroxyindol-3-ylmethyl GL (3) was the major GL in the seeds of B. elongata, along with the four minor GLs 2-(R)-hydroxy-3-butenyl- (1), 3-butenyl- (2), 4-pentenyl- (4) and indol-3-ylmethyl (5). The seeds of B. botteri (Vis island) and B. cazzae (Sušac island) contained 2 as the major GL as well as 1, 3, 5 and 4-methoxyindol-3-ylmethyl GL (6). However, the GLs in B. botteri (Palagruža island) differed from other varieties having 2-propenyl GL (7) as the major GL in the seeds, and the four minor GLs 2, 3, 5 and 6. This first report of the GL content in the seeds of B. elongata, B. botteri and B. cazzae indicates that the unique GL profiles could be specific to the geographical origin of the plant.

Membrane Proteomics of Arabidopsis Glucosinolate Mutants cyp79B2/B3 and myb28/29

PubMed Central

Mostafa, Islam; Yoo, Mi-Jeong; Zhu, Ning; Geng, Sisi; Dufresne, Craig; Abou-Hashem, Maged; El-Domiaty, Maher; Chen, Sixue

2017-01-01

Glucosinolates (Gls) constitute a major group of natural metabolites represented by three major classes (aliphatic, indolic and aromatic) of more than 120 chemical structures. In our previous work, soluble proteins and metabolites in Arabidopsis mutants deficient of aliphatic (myb28/29) and indolic Gls (cyp79B2B3) were analyzed. Here we focus on investigating the changes at the level of membrane proteins in these mutants. Our LC/MS-MS analyses of tandem mass tag (TMT) labeled peptides derived from the cyp79B2/B3 and myb28/29 relative to wild type resulted in the identification of 4,673 proteins, from which 2,171 are membrane proteins. Fold changes and statistical analysis showed 64 increased and 74 decreased in cyp79B2/B3, while 28 increased and 17 decreased in myb28/29. As to the shared protein changes between the mutants, one protein was increased and eight were decreased. Bioinformatics analysis of the changed proteins led to the discovery of three cytochromes in glucosinolate molecular network (GMN): cytochrome P450 86A7 (At1g63710), cytochrome P450 71B26 (At3g26290), and probable cytochrome c (At1g22840). CYP86A7 and CYP71B26 may play a role in hydroxyl-indolic Gls production. In addition, flavone 3′-O-methyltransferase 1 represents an interesting finding as it is likely to participate in the methylation process of the hydroxyl-indolic Gls to form methoxy-indolic Gls. The analysis also revealed additional new nodes in the GMN related to stress and defense activity, transport, photosynthesis, and translation processes. Gene expression and protein levels were found to be correlated in the cyp79B2/B3, but not in the myb28/29. PMID:28443122

Photoprotective Properties of Isothiocyanate and Nitrile Glucosinolate Derivatives From Meadowfoam (Limnanthes alba) Against UVB Irradiation in Human Skin Equivalent

PubMed Central

Carpenter, Evan L.; Le, Mai N.; Miranda, Cristobal L.; Reed, Ralph L.; Stevens, Jan F.; Indra, Arup K.; Ganguli-Indra, Gitali

2018-01-01

Exposure to ultraviolet B (UVB) irradiation of the skin leads to numerous dermatological concerns including skin cancer and accelerated aging. Natural product glucosinolate derivatives, like sulforaphane, have been shown to exhibit chemopreventive and photoprotective properties. In this study, we examined meadowfoam (Limnanthes alba) glucosinolate derivatives, 3-methoxybenzyl isothiocyanate (MBITC) and 3-methoxyphenyl acetonitrile (MPACN), for their activity in protecting against the consequences of UV exposure. To that end, we have exposed human primary epidermal keratinocytes (HPEKs) and 3D human skin reconstructed in vitro (EpiDermTM FT-400) to UVB insult and investigated whether MBITC and MPACN treatment ameliorated the harmful effects of UVB damage. Activity was determined by the compounds’ efficacy in counteracting UVB-induced DNA damage, matrix-metalloproteinase (MMP) expression, and proliferation. We found that in monolayer cultures of HPEK, MBITC and MPACN did not protect against a UVB-induced loss in proliferation and MBITC itself inhibited cell proliferation. However, in human reconstructed skin-equivalents, MBITC and MPACN decrease epidermal cyclobutane pyrimidine dimers (CPDs) and significantly reduce total phosphorylated γH2A.X levels. Both MBITC and MPACN inhibit UVB-induced MMP-1 and MMP-3 expression indicating their role to prevent photoaging. Both compounds, and MPACN in particular, showed activity against UVB-induced proliferation as indicated by fewer epidermal PCNA+ cells and prevented UVB-induced hyperplasia as determined by a reduction in reconstructed skin epidermal thickness (ET). These data demonstrate that MBITC and MPACN exhibit promising anti-photocarcinogenic and anti-photoaging properties in the skin microenvironment and could be used for therapeutic interventions. PMID:29867483

Photoprotective Properties of Isothiocyanate and Nitrile Glucosinolate Derivatives From Meadowfoam (Limnanthes alba) Against UVB Irradiation in Human Skin Equivalent.

PubMed

Carpenter, Evan L; Le, Mai N; Miranda, Cristobal L; Reed, Ralph L; Stevens, Jan F; Indra, Arup K; Ganguli-Indra, Gitali

2018-01-01

Exposure to ultraviolet B (UVB) irradiation of the skin leads to numerous dermatological concerns including skin cancer and accelerated aging. Natural product glucosinolate derivatives, like sulforaphane, have been shown to exhibit chemopreventive and photoprotective properties. In this study, we examined meadowfoam ( Limnanthes alba ) glucosinolate derivatives, 3-methoxybenzyl isothiocyanate (MBITC) and 3-methoxyphenyl acetonitrile (MPACN), for their activity in protecting against the consequences of UV exposure. To that end, we have exposed human primary epidermal keratinocytes (HPEKs) and 3D human skin reconstructed in vitro (EpiDerm TM FT-400) to UVB insult and investigated whether MBITC and MPACN treatment ameliorated the harmful effects of UVB damage. Activity was determined by the compounds' efficacy in counteracting UVB-induced DNA damage, matrix-metalloproteinase (MMP) expression, and proliferation. We found that in monolayer cultures of HPEK, MBITC and MPACN did not protect against a UVB-induced loss in proliferation and MBITC itself inhibited cell proliferation. However, in human reconstructed skin-equivalents, MBITC and MPACN decrease epidermal cyclobutane pyrimidine dimers (CPDs) and significantly reduce total phosphorylated γH2A.X levels. Both MBITC and MPACN inhibit UVB-induced MMP-1 and MMP-3 expression indicating their role to prevent photoaging. Both compounds, and MPACN in particular, showed activity against UVB-induced proliferation as indicated by fewer epidermal PCNA+ cells and prevented UVB-induced hyperplasia as determined by a reduction in reconstructed skin epidermal thickness (ET). These data demonstrate that MBITC and MPACN exhibit promising anti-photocarcinogenic and anti-photoaging properties in the skin microenvironment and could be used for therapeutic interventions.

Evaluation of the Nutritional Quality of Chinese Kale (Brassica alboglabra Bailey) Using UHPLC-Quadrupole-Orbitrap MS/MS-Based Metabolomics.

PubMed

Wang, Ya-Qin; Hu, Li-Ping; Liu, Guang-Min; Zhang, De-Shuang; He, Hong-Ju

2017-07-27

Chinese kale ( Brassica alboglabra Bailey) is a widely consumed vegetable which is rich in antioxidants and anticarcinogenic compounds. Herein, we used an untargeted ultra-high-performance liquid chromatography (UHPLC)-Quadrupole-Orbitrap MS/MS-based metabolomics strategy to study the nutrient profiles of Chinese kale. Seven Chinese kale cultivars and three different edible parts were evaluated, and amino acids, sugars, organic acids, glucosinolates and phenolic compounds were analysed simultaneously. We found that two cultivars, a purple-stem cultivar W1 and a yellow-flower cultivar Y1, had more health-promoting compounds than others. The multivariate statistical analysis results showed that gluconapin was the most important contributor for discriminating both cultivars and edible parts. The purple-stem cultivar W1 had higher levels of some phenolic acids and flavonoids than the green stem cultivars. Compared to stems and leaves, the inflorescences contained more amino acids, glucosinolates and most of the phenolic acids. Meanwhile, the stems had the least amounts of phenolic compounds among the organs tested. Metabolomics is a powerful approach for the comprehensive understanding of vegetable nutritional quality. The results provide the basis for future metabolomics-guided breeding and nutritional quality improvement.

Peruvian Maca (Lepidium peruvianum) - III: The Effects of Cultivation Altitude on Phytochemical and Genetic Differences in the Four Prime Maca Phenotypes.

PubMed

Meissner, Henry O; Mscisz, Alina; Baraniak, Marek; Piatkowska, Ewa; Pisulewski, Pawel; Mrozikiewicz, Mieczyslaw; Bobkiewicz-Kozlowska, Teresa

2017-06-01

In two trials, dietary and Glucosinolates' characteristics in four Maca phenotypes have been examined with an extension into the determination of DNA sequences. Hypocotyls of the four prime phenotypes of Peruvian Maca - Lepidium peruvianum Chacon, labelled as "Yellow", "Black", "Red" and "Purple" were separated from mixed Maca crops cultivated in four geographically-distant locations in the Peruvian Andes at altitudes between 2,800m and 4,300 m a.s.l. It was found that at higher altitudes where Red and Purple Maca phenotypes were grown, the significantly higher ( P <0.05) Glucosinolates' concentrations, adopted as the marker of Maca physiological activity, were observed with the Purple phenotype showing the highest Glucosinolates' content at 4,300m a.s.l., followed by the Red-coloured hypocotyls. Black Maca showed a reversal, but also a significant ( P <0.05) trend, while the Yellow phenotype showed no visible altitude-inflicted response ( P >0.05) and has consistently the lowest Glucosinolates content. Thus, it is reasonable to assume that the altitude at which Red, Purple and Black phenotypes of L. peruvianum are grown, may be responsible for the variation in physiologic functionalities, leading to different than expected specific therapeutic and health benefits induced by Maca phenotypes grown at diverse altitudes. Although promising, insufficiently precise differences in DNA sequences failed to distinguish, without any reasonable doubt, four Maca phenotypes cultivated either in the same or geographically-distant locations, and harvested at different altitudes a.s.l. Further research on DNA sequences is needed, with more primers and larger number of Maca phenotypes, considering biosynthesis of secondary metabolites and adaptation pathways induced by harsh environment at altitudes where Maca is cultivated.

Differential roles of glucosinolates and camalexin at different stages of Agrobacterium-mediated transformation.

PubMed

Shih, Po-Yuan; Chou, Shu-Jen; Müller, Caroline; Halkier, Barbara Ann; Deeken, Rosalia; Lai, Erh-Min

2018-03-02

Agrobacterium tumefaciens is the causal agent of crown gall disease in a wide range of plants via a unique interkingdom DNA transfer from bacterial cells into the plant genome. Agrobacterium tumefaciens is capable of transferring its T-DNA into different plant parts at different developmental stages for transient and stable transformation. However, the plant genes and mechanisms involved in these transformation processes are not well understood. We used Arabidopsis thaliana Col-0 seedlings to reveal the gene expression profiles at early time points during Agrobacterium infection. Common and differentially expressed genes were found in shoots and roots. A gene ontology analysis showed that the glucosinolate (GS) biosynthesis pathway was an enriched common response. Strikingly, several genes involved in indole glucosinolate (iGS) modification and the camalexin biosynthesis pathway were up-regulated, whereas genes in aliphatic glucosinolate (aGS) biosynthesis were generally down-regulated, on Agrobacterium infection. Thus, we evaluated the impacts of GSs and camalexin during different stages of Agrobacterium-mediated transformation combining Arabidopsis mutant studies, metabolite profiling and exogenous applications of various GS hydrolysis products or camalexin. The results suggest that the iGS hydrolysis pathway plays an inhibitory role on transformation efficiency in Arabidopsis seedlings at the early infection stage. Later in the Agrobacterium infection process, the accumulation of camalexin is a key factor inhibiting tumour development on Arabidopsis inflorescence stalks. In conclusion, this study reveals the differential roles of GSs and camalexin at different stages of Agrobacterium-mediated transformation and provides new insights into crown gall disease control and improvement of plant transformation. © 2018 THE AUTHORS. MOLECULAR PLANT PATHOLOGY PUBLISHED BY BRITISH SOCIETY FOR PLANT PATHOLOGY AND JOHN WILEY & SONS LTD.

Cruciferous Vegetables and Human Cancer Risk: Epidemiologic Evidence and Mechanistic Basis

PubMed Central

Higdon, Jane V.; Delage, Barbara; Williams, David E.; Dashwood, Roderick H.

2009-01-01

Cruciferous vegetables are a rich source of glucosinolates and their hydrolysis products, including indoles and isothiocyanates, and high intake of cruciferous vegetables has been associated with lower risk of lung and colorectal cancer in some epidemiological studies. Glucosinolate hydrolysis products alter the metabolism or activity of sex hormones in ways that could inhibit the development of hormone-sensitive cancers, but evidence of an inverse association between cruciferous vegetable intake and breast or prostate cancer in humans is limited and inconsistent. Organizations such as the National Cancer Institute recommend the consumption of 5–9 servings of fruits and vegetables daily, but separate recommendations for cruciferous vegetables have not been established. Isothiocyanates and indoles derived from the hydrolysis of glucosinolates, such as sulforaphane and indole-3-carbinol (I3C), have been implicated in a variety of anticarcinogenic mechanisms, but deleterious effects also have been reported in some experimental protocols, including tumor promotion over prolonged periods of exposure. Epidemiological studies indicate that human exposure to isothiocyanates and indoles through cruciferous vegetable consumption may decrease cancer risk, but the protective effects may be influenced by individual genetic variation (polymorphisms) in the metabolism and elimination of isothiocyanates from the body. Cooking procedures also affect the bioavailability and intake of glucosinolates and their derivatives. Supplementation with I3C or the related dimer 3,3′-diindolylmethane (DIM) alters urinary estrogen metabolite profiles in women, but the effects of I3C and DIM on breast cancer risk are not known. Small preliminary trials in humans suggest that I3C supplementation may be beneficial in treating conditions related to human papilloma virus infection, such as cervical intraepithelial neoplasia and recurrent respiratory papillomatosis, but larger randomized controlled trials are needed. PMID:17317210

Genes Involved in the Evolution of Herbivory by a Leaf-Mining, Drosophilid Fly

PubMed Central

Whiteman, Noah K.; Gloss, Andrew D.; Sackton, Timothy B.; Groen, Simon C.; Humphrey, Parris T.; Lapoint, Richard T.; Sønderby, Ida E.; Halkier, Barbara A.; Kocks, Christine; Ausubel, Frederick M.; Pierce, Naomi E.

2012-01-01

Herbivorous insects are among the most successful radiations of life. However, we know little about the processes underpinning the evolution of herbivory. We examined the evolution of herbivory in the fly, Scaptomyza flava, whose larvae are leaf miners on species of Brassicaceae, including the widely studied reference plant, Arabidopsis thaliana (Arabidopsis). Scaptomyza flava is phylogenetically nested within the paraphyletic genus Drosophila, and the whole genome sequences available for 12 species of Drosophila facilitated phylogenetic analysis and assembly of a transcriptome for S. flava. A time-calibrated phylogeny indicated that leaf mining in Scaptomyza evolved between 6 and 16 million years ago. Feeding assays showed that biosynthesis of glucosinolates, the major class of antiherbivore chemical defense compounds in mustard leaves, was upregulated by S. flava larval feeding. The presence of glucosinolates in wild-type (WT) Arabidopsis plants reduced S. flava larval weight gain and increased egg–adult development time relative to flies reared in glucosinolate knockout (GKO) plants. An analysis of gene expression differences in 5-day-old larvae reared on WT versus GKO plants showed a total of 341 transcripts that were differentially regulated by glucosinolate uptake in larval S. flava. Of these, approximately a third corresponded to homologs of Drosophila melanogaster genes associated with starvation, dietary toxin-, heat-, oxidation-, and aging-related stress. The upregulated transcripts exhibited elevated rates of protein evolution compared with unregulated transcripts. The remaining differentially regulated transcripts also contained a higher proportion of novel genes than the unregulated transcripts. Thus, the transition to herbivory in Scaptomyza appears to be coupled with the evolution of novel genes and the co-option of conserved stress-related genes. PMID:22813779

Comparative study between extraction techniques and column separation for the quantification of sinigrin and total isothiocyanates in mustard seed.

PubMed

Cools, Katherine; Terry, Leon A

2012-07-15

Glucosinolates are β-thioglycosides which are found naturally in Cruciferae including the genus Brassica. When enzymatically hydrolysed, glucosinolates yield isothiocyanates and give a pungent taste. Both glucosinolates and isothiocyanates have been linked with anticancer activity as well as antifungal and antibacterial properties and therefore the quantification of these compounds is scientifically important. A wide range of literature exists on glucosinolates, however the extraction and quantification procedures differ greatly resulting in discrepancies between studies. The aim of this study was therefore to compare the most popular extraction procedures to identify the most efficacious method and whether each extraction can also be used for the quantification of total isothiocyanates. Four extraction techniques were compared for the quantification of sinigrin from mustard cv. Centennial (Brassica juncea L.) seed; boiling water, boiling 50% (v/v) aqueous acetonitrile, boiling 100% methanol and 70% (v/v) aqueous methanol at 70 °C. Prior to injection into the HPLC, the extractions which involved solvents (acetonitrile or methanol) were freeze-dried and resuspended in water. To identify whether the same extract could be used to measure total isothiocyanates, a dichloromethane extraction was carried out on the sinigrin extracts. For the quantification of sinigrin alone, boiling 50% (v/v) acetonitrile was found to be the most efficacious extraction solvent of the four tested yielding 15% more sinigrin than the water extraction. However, the removal of the acetonitrile by freeze-drying had a negative impact on the isothiocyanate content. Quantification of both sinigrin and total isothiocyanates was possible when the sinigrin was extracted using boiling water. Two columns were compared for the quantification of sinigrin revealing the Zorbax Eclipse to be the best column using this particular method. Copyright © 2012 Elsevier B.V. All rights reserved.

Caecal fermentation characteristics, blood composition and growth of rabbits on substitution of soya-bean meal by unconventional high-glucosinolate mustard ( Brassica juncea) meal as protein supplement.

PubMed

Tripathi, M K; Mishra, A S; Mondal, D; Misra, A K; Prasad, R; Jakhmola, R C

2008-02-01

Effect of graded levels of high-glucosinolate mustard (Brassica juncea) meal as substitute of soya-bean meal (SBM) in broiler rabbit diets was studied. Forty weaning rabbits of Soviet Chinchilla and White Giant breed were randomly allocated to one of four experimental diets containing mustard meal (MM) 0, 80, 160 and 245 g/kg. The experiment lasted for 8 weeks. MM had 54.8 mg total glucosinolates (TGLSs) per g dry matter (DM). Diets had TGLS 3.8, 8.4 and 11.98 mg/g DM in 80, 160 and 245 g MM diets, respectively. MM-incorporated diets had higher digestible and linearly (P < 0.01) higher metabolisable energy (ME) content. However, the effect on total tract apparent digestibility of DM, and crude protein was quadratic. Average daily gain (ADG) reduced (P < 0.05) linearly with increasing MM levels in diet, still 80 and 160 g MM diets had similar ADG compared to that of SBM diet. Caecum weight reduced linearly (P < 0.05) with increasing MM levels in diet. The pH of caecal content ranged between 5.85 and 6.19, total N between 1.19 and 1.48 (g per 100 g) and total volatile fatty acids between 4.7 and 5.8 mmol per 100 g, and they were not statistically different. NH3-N ranged between 31.2 and 39.0 mg per 100 ml, and reduced linearly (P < 0.05) while trichloroacetic acid-precipitable nitrogen increased linearly (P < 0.01, ranged between 114 and 247 mg per 100 ml) with increasing MM levels in diet. Blood haemoglobin, packed cell volume and lymphocytes were higher (quadratic effects, P < 0.05) on 245 MM diet, whereas white blood cell count reduced linearly (P < 0.01). Serum aspartate aminotransferase increased linearly (P < 0.01) while alanine aminotransferase and alkaline phosphatase activity, protein, erythrocytes sedimentation rate and red blood cell counts were not affected by MM. Serum Cu, Na and K content increased linearly (P < 0.05) with increasing MM levels. Liver Cu concentration showed quadratic (P < 0.05) increase. Rabbits tolerated 8.4 mg TGLS per g diet (160 g MM per kg) during active growth without any apparent effect on health and growth. It is concluded that MM can replace up to 66% SBM protein in rabbit feeding, whereas complete replacement of SBM with MM reduced feed intake and ADG by 23% and 13%, respectively. Further studies are required to confirm these inclusion levels and glucosinolate tolerance of rabbits.

Event-specific qualitative and quantitative detection of five genetically modified rice events using a single standard reference molecule.

PubMed

Kim, Jae-Hwan; Park, Saet-Byul; Roh, Hyo-Jeong; Shin, Min-Ki; Moon, Gui-Im; Hong, Jin-Hwan; Kim, Hae-Yeong

2017-07-01

One novel standard reference plasmid, namely pUC-RICE5, was constructed as a positive control and calibrator for event-specific qualitative and quantitative detection of genetically modified (GM) rice (Bt63, Kemingdao1, Kefeng6, Kefeng8, and LLRice62). pUC-RICE5 contained fragments of a rice-specific endogenous reference gene (sucrose phosphate synthase) as well as the five GM rice events. An existing qualitative PCR assay approach was modified using pUC-RICE5 to create a quantitative method with limits of detection correlating to approximately 1-10 copies of rice haploid genomes. In this quantitative PCR assay, the square regression coefficients ranged from 0.993 to 1.000. The standard deviation and relative standard deviation values for repeatability ranged from 0.02 to 0.22 and 0.10% to 0.67%, respectively. The Ministry of Food and Drug Safety (Korea) validated the method and the results suggest it could be used routinely to identify five GM rice events. Copyright © 2017 Elsevier Ltd. All rights reserved.

In Vitro Activity of Glucosinolates and Their Degradation Products against Brassica-Pathogenic Bacteria and Fungi

PubMed Central

Sotelo, T.; Lema, M.; Soengas, P.; Cartea, M. E.

2014-01-01

Glucosinolates (GSLs) are secondary metabolites found in Brassica vegetables that confer on them resistance against pests and diseases. Both GSLs and glucosinolate hydrolysis products (GHPs) have shown positive effects in reducing soil pathogens. Information about their in vitro biocide effects is scarce, but previous studies have shown sinigrin GSLs and their associated allyl isothiocyanate (AITC) to be soil biocides. The objective of this work was to evaluate the biocide effects of 17 GSLs and GHPs and of leaf methanolic extracts of different GSL-enriched Brassica crops on suppressing in vitro growth of two bacterial (Xanthomonas campestris pv. campestris and Pseudomonas syringae pv. maculicola) and two fungal (Alternaria brassicae and Sclerotinia scletoriorum) Brassica pathogens. GSLs, GHPs, and methanolic leaf extracts inhibited the development of the pathogens tested compared to the control, and the effect was dose dependent. Furthermore, the biocide effects of the different compounds studied were dependent on the species and race of the pathogen. These results indicate that GSLs and their GHPs, as well as extracts of different Brassica species, have potential to inhibit pathogen growth and offer new opportunities to study the use of Brassica crops in biofumigation for the control of multiple diseases. PMID:25362058

Variation in bioactive content in broccoli (Brassica oleracea var. italica) grown under conventional and organic production systems.

PubMed

Valverde, Juan; Reilly, Kim; Villacreces, Salvador; Gaffney, Michael; Grant, James; Brunton, Nigel

2015-04-01

Broccoli and other cruciferous vegetables contain a number of bioactive compounds, in particular glucosinolates and polyphenols, which are proposed to confer health benefits to the consumer. Demand for organic crops is at least partly based on a perception that organic crops may contain higher levels of bioactive compounds; however, insufficient research has been carried out to either support or refute such claims. In this study we examined the effect of conventional, organic, and mixed cultivation practices on the content of total phenolics, total flavonoids, and total and individual glucosinolates in two varieties of broccoli grown over 2 years in a split-plot factorial systems comparison trial. Levels of total phenolics and total flavonoids showed a significant year-on-year variation but were not significantly different between organic and conventional production systems. In contrast, levels of the indolyl glucosinolates glucobrassicin and neoglucobrassicin were significantly higher (P < 0.05) under fully organic compared to fully conventional management. Organic cultivation practices resulted in significantly higher levels of glucobrassicin and neoglucobrassicin in broccoli florets; however, other investigated compounds were unaffected by production practices. © 2014 Society of Chemical Industry.

Selenium treatment differentially affects sulfur metabolism in high and low glucosinolate producing cultivars of broccoli (Brassica oleracea L.).

PubMed

McKenzie, Marian J; Chen, Ronan K Y; Leung, Susanna; Joshi, Srishti; Rippon, Paula E; Joyce, Nigel I; McManus, Michael T

2017-12-01

The effect of selenium (Se) application on the sulfur (S)-rich glucosinolate (GSL)-containing plant, broccoli (Brassica oleracea L. var. italica) was examined with a view to producing germplasm with increased Se and GSL content for human health, and to understanding the influence of Se on the regulation of GSL production. Two cultivars differing in GSL content were compared. Increased Se application resulted in an increase in Se uptake in planta, but no significant change in total S or total GSL content in either cultivar. Also no significant change was observed in the activity of ATP sulfurylase (ATPS, EC 2.7.7.4) or O-acetylserine(thiol) lyase (OASTL, EC 2.5.1.47) with increased Se application. However, in the first investigation of APS kinase (APSK, EC 2.7.1.25) expression in response to Se fertilisation, an increase in transcript abundance of one variant of APS kinase 1 (BoAPSK1A) was observed in both cultivars, and an increase in BoAPSK2 transcript abundance was observed in the low GSL producing cultivar. A mechanism by which increased APSK transcription may provide a means of controlling the content of S-containing compounds, including GSLs, following Se uptake is proposed. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Development of an innovative immunoassay for CP4EPSPS and Cry1AB genetically modified protein detection and quantification.

PubMed

Ermolli, M; Prospero, A; Balla, B; Querci, M; Mazzeo, A; Van Den Eede, G

2006-09-01

An innovative immunoassay, called enzyme-linked immunoabsorbant assay (ELISA) Reverse, based on a new conformation of the solid phase, was developed. The solid support was expressly designed to be immersed directly in liquid samples to detect the presence of protein targets. Its application is proposed in those cases where a large number of samples have to be screened simultaneously or when the simultaneous detection of different proteins is required. As a first application, a quantitative immunoassay for Cry1AB protein in genetically modified maize was optimized. The method was tested using genetically modified organism concentrations from 0.1 to 2.0%. The limit of detection and limit of quantitation of the method were determined as 0.0056 and 0.0168 (expressed as the percentage of genetically modified organisms content), respectively. A qualitative multiplex assay to assess the presence of two genetically modified proteins simultaneously was also established for the case of the Cry1AB and the CP4EPSPS (5-enolpyruvylshikimate-3-phosphate synthase) present in genetically modified maize and soy, respectively.

Development of an event-specific hydrolysis probe quantitative real-time polymerase chain reaction assay for Embrapa 5.1 genetically modified common bean (Phaseolus vulgaris).

PubMed

Treml, Diana; Venturelli, Gustavo L; Brod, Fábio C A; Faria, Josias C; Arisi, Ana C M

2014-12-10

A genetically modified (GM) common bean event, namely Embrapa 5.1, resistant to the bean golden mosaic virus (BGMV), was approved for commercialization in Brazil. Brazilian regulation for genetically modified organism (GMO) labeling requires that any food containing more than 1% GMO be labeled. The event-specific polymerase chain reaction (PCR) method has been the primary trend for GMO identification and quantitation because of its high specificity based on the flanking sequence. This work reports the development of an event-specific assay, named FGM, for Embrapa 5.1 detection and quantitation by use of SYBR Green or hydrolysis probe. The FGM assay specificity was tested for Embrapa 2.3 event (a noncommercial GM common bean also resistant to BGMV), 46 non-GM common bean varieties, and other crop species including maize, GM maize, soybean, and GM soybean. The FGM assay showed high specificity to detect the Embrapa 5.1 event. Standard curves for the FGM assay presented a mean efficiency of 95% and a limit of detection (LOD) of 100 genome copies in the presence of background DNA. The primers and probe developed are suitable for the detection and quantitation of Embrapa 5.1.

Quantification of glucosinolates, anthocyanins, free amino acids, and vitamin C in inbred lines of cabbage (Brassica oleracea L.).

PubMed

Park, Suhyoung; Valan Arasu, Mariadhas; Lee, Min-Ki; Chun, Jin-Hyuk; Seo, Jeong Min; Lee, Sang-Won; Al-Dhabi, Naif Abdullah; Kim, Sun-Ju

2014-02-15

We profiled and quantified glucosinolates (GSLs), anthocyanins, free amino acids, and vitamin C metabolites in forty-five lines of green and red cabbages. Analysis of these distinct cabbages revealed the presence of 11 GSLs, 13 anthocyanins, 22 free amino acids, and vitamin C. GSL contents were varied amongst the different lines of cabbage. The total GSL content was mean 10.6 μmol/g DW, and sinigrin was the predominant GSL accounted mean 4.0 μmol/g DW (37.7% of the total) followed by glucoraphanin (1.9) and glucobrassicin (2.4). Amongst the 13 anthocyanins, cyanidin 3-(sinapoyl) diglucoside-5-glucoside levels were the highest. The amounts of total free amino acids in green cabbage lines ranged 365.9 mg/100g fresh weight (FW) to 1089.1mg/100g FW. Vitamin C levels were much higher in red cabbage line (129.9 mg/100g FW). Thus, the amounts of GSLs, anthocyanins, free amino acids, and vitamin C varied widely, and the variations in these compounds between the lines of cabbage were significant. Copyright © 2013 Elsevier Ltd. All rights reserved.

Modification of oil and glucosinolate content in canola seeds with altered expression of Brassica napus LEAFY COTYLEDON1.

PubMed

Elahi, Nosheen; Duncan, Robert W; Stasolla, Claudio

2016-03-01

Over the last few decades, research focusing on canola (Brassica napus L.) seed oil content and composition has expanded. Oil production and accumulation are influenced by genes participating in embryo and seed development. The Arabidopsis LEAFY COTYLEDON1 (LEC1) is a well characterized regulator of embryo development that also enhances the expression of genes involved in fatty acid (FA) synthesis. B. napus lines over-expressing or down-regulating BnLEC1 were successfully generated by Agrobacterium-mediated transformation. The constitutive expression of BnLEC1 in B. napus var. Polo, increased seed oil content by 7-16%, while the down-regulation of BnLEC1 in B. napus var. Topas reduced oil content by 9-12%. Experimental manipulation of BnLEC1 caused transcriptional changes in enzymes participating in sucrose metabolism, glycolysis, and FA biosynthesis, suggesting an enhanced carbon flux towards FA biosynthesis in tissues over-expressing BnLEC1. The increase in oil content induced by BnLEC1 was not accompanied by alterations in FA composition, oil nutritional value or glucosinolate (GLS) levels. Suppression of BnLEC1 reduced seed oil accumulation and elevated the level of GLS possibly through the transcriptional regulation of BnST5a (Sulphotransferase5a), the last GLS biosynthetic enzyme. Collectively, these findings demonstrate that experimental alterations of BnLEC1 expression can be used to influence oil production and quality in B. napus. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Degradation of sinigrin by Lactobacillus agilis strain R16.

PubMed

Llanos Palop, M; Smiths, J P; Brink, B T

1995-07-01

Forty-two lactobacilli were screened for their potential to degrade glucosinolate sinigrin. One of them, strain R16, demonstrated a high level of sinigrin degradation; it was identified as Lactobacillus agilis. The sinigrin degrading activity of L. agilis R16 could only be demonstrated when intact cells were used. The products of sinigrin degradation are allyl-isothiocyanate (AITC) and glucose (which is further fermented to DL-lactic acid), suggesting that myrosinase activity is involved. The activity was induced by the presence of sinigrin. Glucose inhibited the myrosinase activity, even in induced cells. Lactobacillus agilis R16 was able to grow on an extract of brown mustard seed and caused glucosinolate degradation.

Cogeneration of biodiesel and nontoxic rapeseed meal from rapeseed through in-situ alkaline transesterification.

PubMed

Qian, Junfeng; Yang, Qiuhui; Sun, Fuan; He, Mingyang; Chen, Qun; Yun, Zhi; Qin, Lizhen

2013-01-01

In-situ alkaline transesterification of rapeseed oil with methanol for the production of biodiesel and nontoxic rapeseed meal was carried out. Water removal from milled rapeseed by methanol washing was more effective than vacuum drying. The conversion rate of rapeseed oil into FAME was 92%, FAME mass was 8.81 g, glucosinolates content in remaining rapeseed meal was 0.12% by methanol washing, while by vacuum drying the values were 46%, 4.44 g, 0.58%, respectively. In the presence of 0.10 mol/L NaOH in methanol, with methanol/oil molar ratio of 180:1 and a 3h reaction at 40 °C, a conversion rate of 98% was achieved, and the glucosinolates content was reduce to 0.07%, a value which below the GB/T 22514-2008 standard in China. Thus the rapeseed meal can be used as a source of protein in animal feed. The FAME prepared through in-situ alkaline transesterification met the ASTM specifications for biodiesel. Copyright © 2012 Elsevier Ltd. All rights reserved.

Influence of colour type and previous cultivation on secondary metabolites in hypocotyls and leaves of maca (Lepidium meyenii Walpers).

PubMed

Clément, Céline; Diaz Grados, Diego A; Avula, Bharathi; Khan, Ihklas A; Mayer, Andrea C; Ponce Aguirre, Dante D; Manrique, Ivan; Kreuzer, Michael

2010-04-15

Maca is an Andean crop of the Brassicaceae family which is mainly known for its fertility-enhancing properties following consumption. The hypocotyls display various colours ranging from white to black. Each colour has different biological effects. The aim of this study was to analyse the concentrations of major secondary metabolites in hypocotyls and leaves of maca in a controlled planting experiment in the Peruvian Andes at 4130 m above sea level. The effects of colour type and of previous cultivation of the field were examined. In the hypocotyls, the colour type effect was significant for most secondary metabolites; exceptions were beta-sitosterol and campesterol. The lead-coloured, yellow and violet maca hypocotyls were rich in glucosinolates, macaene and macamides, respectively. Previous cultivation affected macaene, campesterol and indole glucosinolate concentrations. Effects on metabolite concentrations in the leaves were minor. Hypocotyls were richer in macaene, macamides and glucosinolates than were leaves, and were poorer in beta-sitosterol and total phenols. Colour type has to be considered in maca production, as colour associates with variations in concentrations of distinct bioactive metabolites. Leaves may be interesting for animal nutrition purposes as they contain essentially the same secondary metabolites as the hypocotyls but in clearly lower concentrations. (c) 2010 Society of Chemical Industry.

The genetic basis of a plant–insect coevolutionary key innovation

PubMed Central

Wheat, Christopher W.; Vogel, Heiko; Wittstock, Ute; Braby, Michael F.; Underwood, Dessie; Mitchell-Olds, Thomas

2007-01-01

Ehrlich and Raven formally introduced the concept of stepwise coevolution using butterfly and angiosperm interactions in an attempt to account for the impressive biological diversity of these groups. However, many biologists currently envision butterflies evolving 50 to 30 million years (Myr) after the major angiosperm radiation and thus reject coevolutionary origins of butterfly biodiversity. The unresolved central tenet of Ehrlich and Raven's theory is that evolution of plant chemical defenses is followed closely by biochemical adaptation in insect herbivores, and that newly evolved detoxification mechanisms result in adaptive radiation of herbivore lineages. Using one of their original butterfly-host plant systems, the Pieridae, we identify a pierid glucosinolate detoxification mechanism, nitrile-specifier protein (NSP), as a key innovation. Larval NSP activity matches the distribution of glucosinolate in their host plants. Moreover, by using five different temporal estimates, NSP seems to have evolved shortly after the evolution of the host plant group (Brassicales) (≈10 Myr). An adaptive radiation of these glucosinolate-feeding Pierinae followed, resulting in significantly elevated species numbers compared with related clades. Mechanistic understanding in its proper historical context documents more ancient and dynamic plant–insect interactions than previously envisioned. Moreover, these mechanistic insights provide the tools for detailed molecular studies of coevolution from both the plant and insect perspectives. PMID:18077380

Optimizing isothiocyanate formation during enzymatic glucosinolate breakdown by adjusting pH value, temperature and dilution in Brassica vegetables and Arabidopsis thaliana

NASA Astrophysics Data System (ADS)

Hanschen, Franziska S.; Klopsch, Rebecca; Oliviero, Teresa; Schreiner, Monika; Verkerk, Ruud; Dekker, Matthijs

2017-01-01

Consumption of glucosinolate-rich Brassicales vegetables is associated with a decreased risk of cancer with enzymatic hydrolysis of glucosinolates playing a key role. However, formation of health-promoting isothiocyanates is inhibited by the epithiospecifier protein in favour of nitriles and epithionitriles. Domestic processing conditions, such as changes in pH value, temperature or dilution, might also affect isothiocyanate formation. Therefore, the influences of these three factors were evaluated in accessions of Brassica rapa, Brassica oleracea, and Arabidopsis thaliana. Mathematical modelling was performed to determine optimal isothiocyanate formation conditions and to obtain knowledge on the kinetics of the reactions. At 22 °C and endogenous plant pH, nearly all investigated plants formed nitriles and epithionitriles instead of health-promoting isothiocyanates. Response surface models, however, clearly demonstrated that upon change in pH to domestic acidic (pH 4) or basic pH values (pH 8), isothiocyanate formation considerably increases. While temperature also affects this process, the pH value has the greatest impact. Further, a kinetic model showed that isothiocyanate formation strongly increases due to dilution. Finally, the results show that isothiocyanate intake can be strongly increased by optimizing the conditions of preparation of Brassicales vegetables.

In vitro activity of glucosinolates and their degradation products against brassica-pathogenic bacteria and fungi.

PubMed

Sotelo, T; Lema, M; Soengas, P; Cartea, M E; Velasco, P

2015-01-01

Glucosinolates (GSLs) are secondary metabolites found in Brassica vegetables that confer on them resistance against pests and diseases. Both GSLs and glucosinolate hydrolysis products (GHPs) have shown positive effects in reducing soil pathogens. Information about their in vitro biocide effects is scarce, but previous studies have shown sinigrin GSLs and their associated allyl isothiocyanate (AITC) to be soil biocides. The objective of this work was to evaluate the biocide effects of 17 GSLs and GHPs and of leaf methanolic extracts of different GSL-enriched Brassica crops on suppressing in vitro growth of two bacterial (Xanthomonas campestris pv. campestris and Pseudomonas syringae pv. maculicola) and two fungal (Alternaria brassicae and Sclerotinia scletoriorum) Brassica pathogens. GSLs, GHPs, and methanolic leaf extracts inhibited the development of the pathogens tested compared to the control, and the effect was dose dependent. Furthermore, the biocide effects of the different compounds studied were dependent on the species and race of the pathogen. These results indicate that GSLs and their GHPs, as well as extracts of different Brassica species, have potential to inhibit pathogen growth and offer new opportunities to study the use of Brassica crops in biofumigation for the control of multiple diseases. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Changes in SeMSC, glucosinolates and sulforaphane levels, and in proteome profile in broccoli (Brassica oleracea var. Italica) fertilized with sodium selenate.

PubMed

Sepúlveda, Ignacio; Barrientos, Herna; Mahn, Andrea; Moenne, Alejandra

2013-05-07

The aim of this work was to analyze the effect of sodium selenate fortification on the content of selenomethyl selenocysteine (SeMSC), total glucosinolates and sulforaphane, as well as the changes in protein profile of the inflorescences of broccoli (Brassica oleracea var. Italica). Two experimental groups were considered: plants treated with 100 μmol/L sodium selenate (final concentration in the pot) and control plants treated with water. Fortification began 2 weeks after transplantation and was repeated once a week during 10 weeks. Broccoli florets were harvested when they reached appropriate size. SeMSC content in broccoli florets increased significantly with sodium selenate fortification; but total glucosinolates and sulforaphane content as well as myrosinase activity were not affected. The protein profile of broccoli florets changed due to fortification with sodium selenate. Some proteins involved in general stress-responses were up-regulated, whereas down-regulated proteins were identified as proteins involved in protection against pathogens. This is the first attempt to evaluate the physiological effect of fortification with sodium selenate on broccoli at protein level. The results of this work will contribute to better understanding the metabolic processes related with selenium uptake and accumulation in broccoli.

Cytotoxic and genotoxic effects of metal(oid)s bioactivated in rocket leaves (Eruca vesicaria subsp. sativa Miller).

PubMed

Villatoro-Pulido, Myriam; Font, Rafael; Obregón-Cano, Sara; Moreno-Rojas, Rafael; Amaro-López, Manuel Ángel; Anter, Jauoad; Muñoz-Serrano, Andrés; De Haro Bailón, Antonio; Alonso-Moraga, Angeles; Del Río-Celestino, Mercedes

2013-11-01

Rocket is an important source of essential elements. However, it may also accumulate toxic elements such as metal(oids). The objectives of the present work were (i) to study the uptake of arsenic, lead, cadmium and zinc in rocket grown in contaminated soils, (ii) to establish the genotoxic and cytotoxic activities of this vegetable material, and (iii) to study the modulator role of the glucosinolate and metal contents in the genotoxic/cytotoxic activities. Lead, cadmium and zinc leaf concentrations in our study were over the concentrations allowed by the statutory limit set for metal(oid) contents in vegetables. The accessions were non genotoxic at the different concentrations studied, although one of the accessions showed the highest mutation rates doubling those of negative control. The cytotoxicity assays with HL60 human leukaemia cells showed that the tumouricide activities of rocket leaves decreased with the increasing of metal(oid) concentrations and also with the decreasing of glucosinolate concentrations in their tissues. An interaction between metal(oid)s and glucosinolate degradation products contained in rocket leaves is suggested as the main modulator agents of the biological activity of the plants grown in metal-contaminated soils. Copyright © 2013. Published by Elsevier Ltd.

Genome-Wide Association Study Dissects the Genetic Architecture of Seed Weight and Seed Quality in Rapeseed (Brassica napus L.)

PubMed Central

Li, Feng; Chen, Biyun; Xu, Kun; Wu, Jinfeng; Song, Weilin; Bancroft, Ian; Harper, Andrea L.; Trick, Martin; Liu, Shengyi; Gao, Guizhen; Wang, Nian; Yan, Guixin; Qiao, Jiangwei; Li, Jun; Li, Hao; Xiao, Xin; Zhang, Tianyao; Wu, Xiaoming

2014-01-01

Association mapping can quickly and efficiently dissect complex agronomic traits. Rapeseed is one of the most economically important polyploid oil crops, although its genome sequence is not yet published. In this study, a recently developed 60K Brassica Infinium® SNP array was used to analyse an association panel with 472 accessions. The single-nucleotide polymorphisms (SNPs) of the array were in silico mapped using ‘pseudomolecules’ representative of the genome of rapeseed to establish their hypothetical order and to perform association mapping of seed weight and seed quality. As a result, two significant associations on A8 and C3 of Brassica napus were detected for erucic acid content, and the peak SNPs were found to be only 233 and 128 kb away from the key genes BnaA.FAE1 and BnaC.FAE1. BnaA.FAE1 was also identified to be significantly associated with the oil content. Orthologues of Arabidopsis thaliana HAG1 were identified close to four clusters of SNPs associated with glucosinolate content on A9, C2, C7 and C9. For seed weight, we detected two association signals on A7 and A9, which were consistent with previous studies of quantitative trait loci mapping. The results indicate that our association mapping approach is suitable for fine mapping of the complex traits in rapeseed. PMID:24510440

Nitrates and Glucosinolates as Strong Determinants of the Nutritional Quality in Rocket Leafy Salads

PubMed Central

Cavaiuolo, Marina; Ferrante, Antonio

2014-01-01

Rocket is an important leafy vegetable crop and a good source of antioxidants and anticancer molecules such as glucosinolates and other sulfur compounds. Rocket is also a hyper-accumulator of nitrates which have been considered for long time the main factors that cause gastro-intestinal cancer. In this review, the content of these compounds in rocket tissues and their levels at harvest and during storage are discussed. Moreover, the effect of these compounds in preventing or inducing human diseases is also highlighted. This review provides an update to all the most recent studies carried out on rocket encouraging the consumption of this leafy vegetable to reduce the risk of contracting cancer and other cardiovascular diseases. PMID:24736897

Nitrates and glucosinolates as strong determinants of the nutritional quality in rocket leafy salads.

PubMed

Cavaiuolo, Marina; Ferrante, Antonio

2014-04-14

Rocket is an important leafy vegetable crop and a good source of antioxidants and anticancer molecules such as glucosinolates and other sulfur compounds. Rocket is also a hyper-accumulator of nitrates which have been considered for long time the main factors that cause gastro-intestinal cancer. In this review, the content of these compounds in rocket tissues and their levels at harvest and during storage are discussed. Moreover, the effect of these compounds in preventing or inducing human diseases is also highlighted. This review provides an update to all the most recent studies carried out on rocket encouraging the consumption of this leafy vegetable to reduce the risk of contracting cancer and other cardiovascular diseases.

Effect of different proportion of sulphur treatments on the contents of glucosinolate in kale (Brassica oleracea var. acephala) commonly consumed in Republic of Korea.

PubMed

Park, Ye-Jin; Lee, Hye-Min; Shin, MinJung; Arasu, Mariadhas Valan; Chung, Doug Young; Al-Dhabi, Naif Abdullah; Kim, Sun-Ju

2018-02-01

Kale ( Brassica oleracea L. Acephala Group) is the rich source of medicinal value sulphur compounds, glucosinolates (GLSs). The aim of this study was to investigate the effect of different proportion of sulphur (S) supplementation levels on the accumulation of GLSs in the leaves of the kale cultivar ('TBC'). High performance liquid chromatography (HPLC) separation method guided to identify and quantify six GSLs including three aliphatic (progoitrin, sinigrin and gluconapin) and three indolyl (glucobrassicin, 4-methoxyglucobrassicin and neoglucobrasscin) respectively. Analysis of these distinct levels of S supplementation revealed that the accumulation of individual and total GLSs was directly proportional to the S concentration. The maximum levels of total GLSs (26.8 µmol/g DW) and glucobrassicin (9.98 µmol/g DW) were found in lower and upper parts of the leaves supplemented with 1 mM and 2 mM S, respectively. Interestingly, aliphatic GSLs were noted predominant in all the parts (50.1, 59.3 and 56% of total GSLs). Among the aliphatic and indolyl GSLs, sinigrin and glucobrassicin account 35.3 and 30.88% of the total GSLs. From this study, it is concluded that supply of S enhance the GSLs accumulation in kale.

The Deposition and Elimination of Glucosinolate Metabolites Derived from Rapeseed Meal in Eggs of Laying Hens.

PubMed

Zhu, L P; Wang, J P; Ding, X M; Bai, S P; Zeng, Q F; Su, Z W; Xuan, Y; Zhang, K Y

2018-02-14

This study was to investigate the deposition and elimination of glucosinolate metabolites including 5-vinyl-1,3-oxazolidine-2-thione (5-VOT) and thiocyanate ion (SCN - ) derived from rapeseed meal (RSM) in hen eggs. During 12 weeks accumulation phase, the serum triiodothyronine, thyronine, blood urea nitrogen, kidney index, and thyroid index linearly increased with the RSM at week 12 (P < 0.05). The thyroid histopathology revealed a sign of hyperplastic goiter in hens fed with 17.64-29.40% RSM. The 5-VOT content of eggs (Y, ng/g) was correlated with 5-VOT intake (X 2 , μg/d·bird) and 5-VOT feeding time (X 1 , week): Y = 54.94X 1 + 0.51X 2 - 430.34 (P < 0.01, R 2 = 0.80). The SCN - content of eggs (Y, mg/kg) was correlated with RSM intake (X 2 , μg/d·bird) and RSM feeding time (X 1 , week): Y = 0.095X 1 + 0.302X 2 - 0.4211 (P < 0.01, R 2 = 0.70). After a 4-week withdrawal of RSM, the 5-VOT and SCN - did not show in eggs. Taken together, 5.88% RSM with dietary supplements of 23.55 mg/kg 5-VOT and 10.76 mg/kg SCN - had no effects on hens with regard to serum parameters, organ index, and thyroid histopathology, and more than 4 weeks withdrawal should be considered for human and hen health.

The kinetic of key phytochemical compounds of non-heading and heading leafy Brassica oleracea landraces as affected by traditional cooking methods.

PubMed

Giambanelli, Elisa; Verkerk, Ruud; D'Antuono, L Filippo; Oliviero, Teresa

2016-11-01

Kales are often a key ingredient of traditional foods, containing high amounts of indolic glucosinolates (precursors of indole-3-carbinol and ascorbigen), carotenoids and phenolics. The present trend to associate traditional foods crops with health-promoting properties suggested to investigate the degradation kinetic of three Brassica oleracea landraces' phytochemicals subjected to boiling, steaming and stir-frying. Boiling led to substantial losses due to leaching. Glucosinolates followed a second-order degradation kinetic (20% of their initial values after 10 min in Nero di Toscana). Phenolic content in leaves + cooking water remained unchanged, whereas their antioxidant capacity was reduced. Carotenoid content increased during the first minutes of boiling. Steaming showed the highest retention of phytochemicals, with often zero-order degradation kinetic, having however a strong effect on colour. Stir-frying produced high losses for all measured compounds; also, β-carotene reduced its content to 10-23% independently of variety. Conversion values for indole-derived compounds ranged from non-detectable to 23.5%. Variety strongly affected observed degradation rates because of a different glucosinolate composition and leaf structure. With this research, more information has been gained on the degradation kinetic of B. oleracea landraces' phytochemical compounds upon cooking, highlighting the possibility of improving bioactive component retention. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Only a minority of broad-range detoxification genes respond to a variety of phytotoxins in generalist Bemisia tabaci species

PubMed Central

Halon, Eyal; Eakteiman, Galit; Moshitzky, Pnina; Elbaz, Moshe; Alon, Michal; Pavlidi, Nena; Vontas, John; Morin, Shai

2015-01-01

Generalist insect can utilize two different modes for regulating their detoxification genes, the constitutive mode and the induced mode. Here, we used the Bemisia tabaci sibling species MEAM1 and MED, as a model system for studying constitutive and induced detoxification resistance and their associated tradeoffs. B. tabaci adults were allowed to feed through membranes for 24 h on diet containing only sucrose or sucrose with various phytotoxins. Quantitative real-time PCR analyses of 18 detoxification genes, indicated that relatively few transcripts were changed in both the MEAM1 and MED species, in response to the addition of phytotoxins to the diet. Induced transcription of detoxification genes only in the MED species, in response to the presence of indole-3-carbinol in the insect’s diet, was correlated with maintenance of reproductive performance in comparison to significant reduction in performance of the MEAM1 species. Three genes, COE2, CYP6-like 5 and BtGST2, responded to more than one compound and were highly transcribed in the insect gut. Furthermore, functional assays showed that the BtGST2 gene encodes a protein capable of interacting with both flavonoids and glucosinolates. In conclusion, several detoxification genes were identified that could potentially be involved in the adaptation of B. tabaci to its host plants. PMID:26655836

Mechanisms of Action of Isothiocyanates in Cancer Chemoprevention: An Update

PubMed Central

Navarro, Sandi L.; Li, Fei; Lampe, Johanna W.

2011-01-01

Isothiocyanates, derived from glucosinolates, are thought to be responsible for the chemoprotective actions conferred by higher cruciferous vegetable intake. Evidence suggests that isothiocyanates exert their effects through a variety of distinct but interconnected signaling pathways important for inhibiting carcinogenesis, including those involved in detoxification, inflammation, apoptosis, and cell cycle and epigenetic regulation, among others. This article provides an update on the latest research on isothiocyanates and these mechanisms, and points out remaining gaps in our understanding of these events. Given the variety of ITC produced from glucosinolates, and the diverse pathways on which these compounds act, a systems biology approach, in vivo, may help to better characterize their integrated role in cancer prevention. In addition, the effects of dose, duration of exposure, and specificity of different ITC should be considered. PMID:21935537

Effect of atmospheric carbon dioxide levels and nitrate fertilization on glucosinolate biosynthesis in mechanically damaged Arabidopsis plants.

PubMed

Paudel, Jamuna Risal; Amirizian, Alexandre; Krosse, Sebastian; Giddings, Jessica; Ismail, Shoieb Akaram Arief; Xia, Jianguo; Gloer, James B; van Dam, Nicole M; Bede, Jacqueline C

2016-03-22

Increased atmospheric carbon dioxide (CO2) levels predicted to occur before the end of the century will impact plant metabolism. In addition, nitrate availability will affect metabolism and levels of nitrogen-containing defense compounds, such as glucosinolates (GSLs). We compared Arabidopsis foliar metabolic profile in plants grown under two CO2 regimes (440 vs 880 ppm), nitrate fertilization (1 mM vs 10 mM) and in response to mechanical damage of rosette leaves. Constitutive foliar metabolites in nitrate-limited plants show distinct global patterns depending on atmospheric CO2 levels; in contrast, plants grown under higher nitrate fertilization under elevated atmospheric CO2 conditions have a unique metabolite signature. Nitrate fertilization dampens the jasmonate burst in response to wounding in plants grown at elevated CO2 levels. Leaf GSL profile mirrors the jasmonate burst; in particular, indole GSLs increase in response to damage in plants grown at ambient CO2 but only in nitrate-limited plants grown under elevated CO2 conditions. This may reflect a reduced capacity of C3 plants grown under enriched CO2 and nitrate levels to signal changes in oxidative stress and has implications for future agricultural management practices.

A modifier of Huntington's disease onset at the MLH1 locus.

PubMed

Lee, Jong-Min; Chao, Michael J; Harold, Denise; Abu Elneel, Kawther; Gillis, Tammy; Holmans, Peter; Jones, Lesley; Orth, Michael; Myers, Richard H; Kwak, Seung; Wheeler, Vanessa C; MacDonald, Marcy E; Gusella, James F

2017-10-01

Huntington's disease (HD) is a dominantly inherited neurodegenerative disease caused by an expanded CAG repeat in HTT. Many clinical characteristics of HD such as age at motor onset are determined largely by the size of HTT CAG repeat. However, emerging evidence strongly supports a role for other genetic factors in modifying the disease pathogenesis driven by mutant huntingtin. A recent genome-wide association analysis to discover genetic modifiers of HD onset age provided initial evidence for modifier loci on chromosomes 8 and 15 and suggestive evidence for a locus on chromosome 3. Here, genotyping of candidate single nucleotide polymorphisms in a cohort of 3,314 additional HD subjects yields independent confirmation of the former two loci and moves the third to genome-wide significance at MLH1, a locus whose mouse orthologue modifies CAG length-dependent phenotypes in a Htt-knock-in mouse model of HD. Both quantitative and dichotomous association analyses implicate a functional variant on ∼32% of chromosomes with the beneficial modifier effect that delays HD motor onset by 0.7 years/allele. Genomic DNA capture and sequencing of a modifier haplotype localize the functional variation to a 78 kb region spanning the 3'end of MLH1 and the 5'end of the neighboring LRRFIP2, and marked by an isoleucine-valine missense variant in MLH1. Analysis of expression Quantitative Trait Loci (eQTLs) provides modest support for altered regulation of MLH1 and LRRFIP2, raising the possibility that the modifier affects regulation of both genes. Finally, polygenic modification score and heritability analyses suggest the existence of additional genetic modifiers, supporting expanded, comprehensive genetic analysis of larger HD datasets. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

A 2-Oxoglutarate-Dependent Dioxygenase Mediates the Biosynthesis of Glucoraphasatin in Radish1[OPEN

PubMed Central

Kitashiba, Hiroyasu; Li, Feng; Fukino, Nobuko; Ohara, Takayoshi; Nishio, Takeshi; Ishida, Masahiko

2017-01-01

Glucosinolates (GSLs) are secondary metabolites whose degradation products confer intrinsic flavors and aromas to Brassicaceae vegetables. Several structures of GSLs are known in the Brassicaceae, and the biosynthetic pathway and regulatory networks have been elucidated in Arabidopsis (Arabidopsis thaliana). GSLs are precursors of chemical defense substances against herbivorous pests. Specific GSLs can act as feeding blockers or stimulants, depending on the pest species. Natural selection has led to diversity in the GSL composition even within individual species. However, in radish (Raphanus sativus), glucoraphasatin (4-methylthio-3-butenyl glucosinolate) accounts for more than 90% of the total GSLs, and little compositional variation is observed. Because glucoraphasatin is not contained in other members of the Brassicaceae, like Arabidopsis and cabbage (Brassica oleracea), the biosynthetic pathways for glucoraphasatin remain unclear. In this report, we identified and characterized a gene encoding GLUCORAPHASATIN SYNTHASE 1 (GRS1) by genetic mapping using a mutant that genetically lacks glucoraphasatin. Transgenic Arabidopsis, which overexpressed GRS1 cDNA, accumulated glucoraphasatin in the leaves. GRS1 encodes a 2-oxoglutarate-dependent dioxygenase, and it is abundantly expressed in the leaf. To further investigate the biosynthesis and transportation of GSLs in radish, we grafted a grs1 plant onto a wild-type plant. The grafting experiment revealed a leaf-to-root long-distance glucoraphasatin transport system in radish and showed that the composition of GSLs differed among the organs. Based on these observations, we propose a characteristic biosynthesis pathway for glucoraphasatin in radish. Our results should be useful in metabolite engineering for breeding of high-value vegetables. PMID:28100450

Analysis of the extracts of Isatis tinctoria by new analytical approaches of HPLC, MS and NMR.

PubMed

Zhou, Jue; Qu, Fan

2011-01-01

The methods of extraction, separation and analysis of alkaloids and indole glucosinolates (GLs) ofIsatis tinctoria were reviewed. Different analytical approaches such as High-pressure Liquid Chromatography (HPLC), Liquid Chromatography with Electrospray Ionization Mass Spectrometry (LC/ESI/MS), Electrospray Ionization Time-Of-Flight Mass Spectrometry (ESI-TOF-MS), and Nuclear Magnetic Resonance (NMR) were used to validate and identity of these constituents. These methods provide rapid separation, identification and quantitative measurements of alkaloids and GLs of Isatis tinctoria. By connection with different detectors to HPLC such as PDA, ELSD, ESI- and APCI-MS in positive and negative ion modes, complicated compounds could be detected with at least two independent detection modes. The molecular formula can be derived in a second step of ESI-TOF-MS data. But for some constituents, UV and MS cannot provide sufficient structure identification. After peak purification, NMR by semi-preparative HPLC can be used as a complementary method.

Internal-Modified Dithiol DNA–Directed Au Nanoassemblies: Geometrically Controlled Self–Assembly and Quantitative Surface–Enhanced Raman Scattering Properties

PubMed Central

Yan, Yuan; Shan, Hangyong; Li, Min; Chen, Shu; Liu, Jianyu; Cheng, Yanfang; Ye, Cui; Yang, Zhilin; Lai, Xuandi; Hu, Jianqiang

2015-01-01

In this work, a hierarchical DNA–directed self–assembly strategy to construct structure–controlled Au nanoassemblies (NAs) has been demonstrated by conjugating Au nanoparticles (NPs) with internal–modified dithiol single-strand DNA (ssDNA) (Au–B–A or A–B–Au–B–A). It is found that the dithiol–ssDNA–modified Au NPs and molecule quantity of thiol–modified ssDNA grafted to Au NPs play critical roles in the assembly of geometrically controlled Au NAs. Through matching Au–DNA self–assembly units, geometrical structures of the Au NAs can be tailored from one–dimensional (1D) to quasi–2D and 2D. Au–B–A conjugates readily give 1D and quasi–2D Au NAs while 2D Au NAs can be formed by A–B–Au–B–A building blocks. Surface-enhanced Raman scattering (SERS) measurements and 3D finite–difference time domain (3D-FDTD) calculation results indicate that the geometrically controllable Au NAs have regular and linearly “hot spots”–number–depended SERS properties. For a certain number of NPs, the number of “hot spots” and accordingly enhancement factor of Au NAs can be quantitatively evaluated, which open a new avenue for quantitative analysis based on SERS technique. PMID:26581251

Internal-Modified Dithiol DNA-Directed Au Nanoassemblies: Geometrically Controlled Self-Assembly and Quantitative Surface-Enhanced Raman Scattering Properties

NASA Astrophysics Data System (ADS)

Yan, Yuan; Shan, Hangyong; Li, Min; Chen, Shu; Liu, Jianyu; Cheng, Yanfang; Ye, Cui; Yang, Zhilin; Lai, Xuandi; Hu, Jianqiang

2015-11-01

In this work, a hierarchical DNA-directed self-assembly strategy to construct structure-controlled Au nanoassemblies (NAs) has been demonstrated by conjugating Au nanoparticles (NPs) with internal-modified dithiol single-strand DNA (ssDNA) (Au-B-A or A-B-Au-B-A). It is found that the dithiol-ssDNA-modified Au NPs and molecule quantity of thiol-modified ssDNA grafted to Au NPs play critical roles in the assembly of geometrically controlled Au NAs. Through matching Au-DNA self-assembly units, geometrical structures of the Au NAs can be tailored from one-dimensional (1D) to quasi-2D and 2D. Au-B-A conjugates readily give 1D and quasi-2D Au NAs while 2D Au NAs can be formed by A-B-Au-B-A building blocks. Surface-enhanced Raman scattering (SERS) measurements and 3D finite-difference time domain (3D-FDTD) calculation results indicate that the geometrically controllable Au NAs have regular and linearly “hot spots”-number-depended SERS properties. For a certain number of NPs, the number of “hot spots” and accordingly enhancement factor of Au NAs can be quantitatively evaluated, which open a new avenue for quantitative analysis based on SERS technique.

Quantitation of exposure to benzo[a]pyrene with monoclonal antibodies.

PubMed Central

Santella, R M; Hsieh, L L; Lin, C D; Viet, S; Weinstein, I B

1985-01-01

It is now possible to quantitate carcinogen adducts on DNA by highly sensitive immunoassays. These techniques are particularly useful for screening human populations for exposure to potential environmental carcinogens. We have developed a panel of monoclonal antibodies that react with benzo(a)pyrene (BP) modified DNA to be used in an enzyme linked immunoassay (ELISA) to quantitate adduct levels of both human and animal samples. BALBc/Cr mice were immunized with either DNA modified by 7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9, 10-tetrahydrobenzo(a)pyrene (BPDE-I-DNA) complexed electrostatically to methylated bovine serum albumin or with BPDE-I-modified guanosine conjugated with bovine serum albumin (BPDE-I-G-BSA). Four stable clones were produced from the spleen cells of animals immunized with BPDE-I-DNA and one from BPDE-I-G-BSA immunized animals. All antibodies were shown to be highly specific for BPDE-I-DNA and did not crossreact with nonmodified DNA or with N-2-acetylaminofluorene or 1-aminopyrene modified DNA. The antibodies differed in their sensitivity to BPDE-II-DNA, BPDE-I-poly G, BPDE-I-tetraols and BPDE-I-dG. In general, all the antibodies showed the greatest affinity for their original antigen. Those generated against modified DNA showed highest reactivity against modified DNA while the one antibody generated against the monoadduct showed highest reactivity with the monoadduct. These antibodies are currently being used in a highly sensitive competitive ELISA to quantitate levels of BP-DNA adducts in various animal and human tissue samples. PMID:4085452

Effect of industrial freezing on the stability of chemopreventive compounds in broccoli.

PubMed

Alanís-Garza, Pedro A; Becerra-Moreno, Alejandro; Mora-Nieves, José Luis; Mora-Mora, Juan Pablo; Jacobo-Velázquez, Daniel A

2015-05-01

Broccoli (Brassica oleracea L. var. Italica) is largely consumed all over the world and has a high economic importance. Likewise, broccoli contains high levels of glucosinolates, carotenoids and total phenols, which are related with the prevention of chronic diseases. The present project's objective was to evaluate the effect of industrial freezing on the stability of bioactive molecules in seven commercial broccoli cultivars (Tlaloc®, Endurance®, Florapack®, Domador®, Steel®, Iron Man® and Avenger®). In general, industrial freezing increased the extractability of total glucosinolates, whereas total phenols remained constant in most broccoli cultivars. Likewise, broccoli subjected to industrial freezing showed higher levels of total carotenoids (∼60-300% higher) as compared with fresh broccoli. Results suggest that bioactive compounds in frozen broccoli would be more bioavailable than in raw.

Phytochemicals of Brassicaceae in plant protection and human health--influences of climate, environment and agronomic practice.

PubMed

Björkman, Maria; Klingen, Ingeborg; Birch, Andrew N E; Bones, Atle M; Bruce, Toby J A; Johansen, Tor J; Meadow, Richard; Mølmann, Jørgen; Seljåsen, Randi; Smart, Lesley E; Stewart, Derek

2011-05-01

In this review, we provide an overview of the role of glucosinolates and other phytochemical compounds present in the Brassicaceae in relation to plant protection and human health. Current knowledge of the factors that influence phytochemical content and profile in the Brassicaceae is also summarized and multi-factorial approaches are briefly discussed. Variation in agronomic conditions (plant species, cultivar, developmental stage, plant organ, plant competition, fertilization, pH), season, climatic factors, water availability, light (intensity, quality, duration) and CO(2) are known to significantly affect content and profile of phytochemicals. Phytochemicals such as the glucosinolates and leaf surface waxes play an important role in interactions with pests and pathogens. Factors that affect production of phytochemicals are important when designing plant protection strategies that exploit these compounds to minimize crop damage caused by plant pests and pathogens. Brassicaceous plants are consumed increasingly for possible health benefits, for example, glucosinolate-derived effects on degenerative diseases such as cancer, cardiovascular and neurodegenerative diseases. Thus, factors influencing phytochemical content and profile in the production of brassicaceous plants are worth considering both for plant and human health. Even though it is known that factors that influence phytochemical content and profile may interact, studies of plant compounds were, until recently, restricted by methods allowing only a reductionistic approach. It is now possible to design multi-factorial experiments that simulate their combined effects. This will provide important information to ecologists, plant breeders and agronomists. Copyright © 2011 Elsevier Ltd. All rights reserved.

Protective Effects of Lepidium meyenii (Maca) Aqueous Extract and Lycopene on Testosterone Propionate-Induced Prostatic Hyperplasia in Mice.

PubMed

Zou, Ying; Aboshora, Waleed; Li, Jing; Xiao, Tiancun; Zhang, Lianfu

2017-08-01

The inhibitory effect of maca extractant, lycopene, and their combination was evaluated in benign prostatic hyperplasia (BPH) mice induced by testosterone propionate. Mice were divided into a saline group, solvent control group and testosterone propionate-induced BPH mice [BPH model group, solvent BPH model group, benzyl glucosinolate group (1.44 mg/kg), maca group (60 mg/kg), lycopene treated (15, 5, and 2.5 mg/kg), maca (30 mg/kg) combine lycopene treated (7.5, 2.5, and 1.25 mg/kg), and finasteride treated]. Benzyl glucosinolate was used in order to evaluate its pharmacological activity on BPH to find out whether it is the major active component of maca aqueous extract. Finasteride was used as positive control. The compounds were administered once for 30 successive days. Compared with solvent BPH model group, BPH mice fed with maca (30 mg/kg) and lycopene (7.5 mg/kg) combination exhibited significant reductions in the prostatic index, prostatic acid phospatase, estradiol, testosterone, and dihydrotestosterone levels in serum. They also had similar histological compared with those aspects observed in the mice in the solvent control group. The results indicated that combination of maca and lycopene synergistically inhibits BPH in mice. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Glucosinolates from pak choi and broccoli induce enzymes and inhibit inflammation and colon cancer differently.

PubMed

Lippmann, Doris; Lehmann, Carsten; Florian, Simone; Barknowitz, Gitte; Haack, Michael; Mewis, Inga; Wiesner, Melanie; Schreiner, Monika; Glatt, Hansruedi; Brigelius-Flohé, Regina; Kipp, Anna P

2014-06-01

High consumption of Brassica vegetables is considered to prevent especially colon carcinogenesis. The content and pattern of glucosinolates (GSLs) can highly vary among different Brassica vegetables and may, thus, affect the outcome of Brassica intervention studies. Therefore, we aimed to feed mice with diets containing plant materials of the Brassica vegetables broccoli and pak choi. Further enrichment of the diets by adding GSL extracts allowed us to analyze the impact of different amounts (GSL-poor versus GSL-rich) and different patterns (broccoli versus pak choi) of GSLs on inflammation and tumor development in a model of inflammation-triggered colon carcinogenesis (AOM/DSS model). Serum albumin adducts were analyzed to confirm the up-take and bioactivation of GSLs after feeding the Brassica diets for four weeks. In agreement with their high glucoraphanin content, broccoli diets induced the formation of sulforaphane-lysine adducts. Levels of 1-methoxyindolyl-3-methyl-histidine adducts derived from neoglucobrassicin were the highest in the GSL-rich pak choi group. In the colon, the GSL-rich broccoli and the GSL-rich pak choi diet up-regulated the expression of different sets of typical Nrf2 target genes like Nqo1, Gstm1, Srxn1, and GPx2. GSL-rich pak choi induced the AhR target gene Cyp1a1 but did not affect Ugt1a1 expression. Both colitis and tumor number were drastically reduced after feeding the GSL-rich pak choi diet while the other three diets had no effect. GSLs can act anti-inflammatory and anti-carcinogenic but both effects depend on the specific amount and pattern of GSLs within a vegetable. Thus, a high Brassica consumption cannot be generally considered to be cancer-preventive.

Nutritional characteristics of camelina meal for 3-week-old broiler chickens.

PubMed

Pekel, A Y; Kim, J I; Chapple, C; Adeola, O

2015-03-01

Limited information on nutritional characteristics on camelina meal for broiler chickens limits its use in diets of broiler chickens. The objectives of this study were to determine the ileal digestible energy (IDE), ME, and MEn contents of 2 different camelina meal (CM1 and CM2) samples for 3-wk-old broiler chickens using the regression method and to determine glucosinolate compounds in the camelina meal samples. The CM1 and CM2 were incorporated into a corn-soybean meal-based reference diet at 3 levels (0, 100, or 200 g/kg) by replacing the energy-yielding ingredients. These 5 diets (reference diet, and 100 and 200 g/kg camelina meal from each of CM1 and CM2) were fed to 320 male Ross 708 broilers from d 21 to 28 post hatching with 8 birds per cage and 8 replicates per treatment in a randomized complete block design. Excreta were collected twice daily from d 25 to 28, and jejunal digesta and ileal digesta from the Meckel's diverticulum to approximately 2 cm proximal to the ileocecal junction were collected on d 28. The total glucosinolate content for CM1 and CM2 were 24.2 and 22.7 nmol/mg, respectively. Jejunal digesta viscosity was linearly increased (P<0.001) from 2.2 to 4.1 cP with increasing dietary camelina meal levels. There were linear effects (P<0.001) of CM1 and CM2 substitution on final weight, weight gain, feed intake, and G:F. The inclusion of CM1 and CM2 linearly decreased (P<0.001) ileal digestibility of DM, energy, and IDE. The supplementation of CM1 and CM2 linearly decreased (P<0.001) the retention of DM, nitrogen, and energy; ME, and MEn. By regressing the CM1 and CM2-associated IDE intake in kilocalories against kilograms of CM1 and CM2 intake, the IDE regression equation was Y=-10+1,429×CM1+2,125×CM2, r2=0.55, which indicates that IDE values were 1,429 kcal/kg of DM for CM1 and 2,125 kcal/kg of DM for CM2. The ME regression was Y=5+882×CM1+925×CM2, r2=0.54, which implies ME values of 882 kcal/kg of DM for CM1 and 925 kcal/kg of DM for CM2. MEn regression was Y=2+795×CM1+844×CM2, r2=0.52, which implies MEn values of 795 kcal/kg of DM for CM1 and 844 kcal/kg of DM for CM2. Based on these results, utilization of energy and nitrogen in camelina meal by broiler chickens is low and the high viscosity observed in jejunal digesta as well as the total glucosinolate in camelina meal may have contributed to the poor energy and nitrogen utilization. © 2015 Poultry Science Association Inc.

7 CFR 810.301 - Definition of canola.

Code of Federal Regulations, 2013 CFR

2013-01-01

... genus Brassica from which the oil shall contain less than 2 percent erucic acid in its fatty acid... glucosinolate, per gram of air-dried, oil free solid. Before the removal of dockage, the seed shall contain not...

7 CFR 810.301 - Definition of canola.

Code of Federal Regulations, 2012 CFR

2012-01-01

... genus Brassica from which the oil shall contain less than 2 percent erucic acid in its fatty acid... glucosinolate, per gram of air-dried, oil free solid. Before the removal of dockage, the seed shall contain not...

7 CFR 810.301 - Definition of canola.

Code of Federal Regulations, 2014 CFR

2014-01-01

... genus Brassica from which the oil shall contain less than 2 percent erucic acid in its fatty acid... glucosinolate, per gram of air-dried, oil free solid. Before the removal of dockage, the seed shall contain not...

Oil Crop Potential for Biodiesel Production: Summary of Three Years of Spring Mustard Research -- Methodologies, Results, and Recommendations; 2000-2003

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brown, J.

2005-07-01

This report summarizes a project whose goal was to support R&D to develop an oil-seed crop that has the potential to reduce the feedstock cost of biodiesel to between 7 and 8 cents per pound of oil and expand supplies of biodiesel as demand for biodiesel grows. The key to this goal is that the non-oil fraction of the oil crop (the seed meal) must have a high value outside of the animal feed markets and produce oil that is not suitable for human consumption. To that end, a spring breeding program was developed to increase diversity of glucosinolate andmore » the concentration of glucosinolates in the meal and to optimize the oil composition for biodiesel fuels. This report presents the research on the spring planted hybrids.« less

Nutritional and Flavor Components of Brassica Xapa L. Grown on ISS

NASA Technical Reports Server (NTRS)

Musgrave, M. E.; Kuang, A.; Blasiak, J.; Tuominen, L. K.; Levine, L. H.; Morrow, R. C.

2005-01-01

Brassica rapa L. cv. 'Astroplants' were grown on the International Space Station during April - June 2002 in the Biomass Production System. Plants were manually pollinated and were maturing seeds when they were harvested for preservation in flight by fixation or freezing. Overall growth and development were comparable between flight and ground control plants. Chlorophyll and carbohydrate content of the leaves were the same in the two treatments. Although comparable numbers of seeds were produced inside the seed pods, the developing seeds from the spaceflight treatment had only half of the dry weight of the ground controls and had altered storage components. Glucosinolate content of the stem tissue was also determined. The concentration of 3-butenyl-glucosinolate was on average 75% greater in the spaceflight samples than in the ground control. The results demonstrate how the spaceflight environment influences nutritional and flavor characteristics of a potential crop for use in a Biological Life Support System.

Myrosin Idioblast Cell Fate and Development Are Regulated by the Arabidopsis Transcription Factor FAMA, the Auxin Pathway, and Vesicular Trafficking[W

PubMed Central

Li, Meng; Sack, Fred D.

2014-01-01

Crucifer shoots harbor a glucosinolate-myrosinase system that defends against insect predation. Arabidopsis thaliana myrosinase (thioglucoside glucohydrolase [TGG]) accumulates in stomata and in myrosin idioblasts (MIs). This work reports that the basic helix-loop-helix transcription factor FAMA that is key to stomatal development is also expressed in MIs. The loss of FAMA function abolishes MI fate as well as the expression of the myrosinase genes TGG1 and TGG2. MI cells have previously been reported to be located in the phloem. Instead, we found that MIs arise from the ground meristem rather than provascular tissues and thus are not homologous with phloem. Moreover, MI patterning and morphogenesis are abnormal when the function of the ARF-GEF gene GNOM is lost as well as when auxin efflux and vesicular trafficking are chemically disrupted. Stomata and MI cells constitute part of a wider system that reduces plant predation, the so-called “mustard oil bomb,” in which vacuole breakage in cells harboring myrosinase and glucosinolate yields a brew toxic to many animals, especially insects. This identification of the gene that confers the fate of MIs, as well as stomata, might facilitate the development of strategies for engineering crops to mitigate predation. PMID:25304201

Simplex and duplex event-specific analytical methods for functional biotech maize.

PubMed

Lee, Seong-Hun; Kim, Su-Jeong; Yi, Bu-Young

2009-08-26

Analytical methods are very important in the control of genetically modified organism (GMO) labeling systems or living modified organism (LMO) management for biotech crops. Event-specific primers and probes were developed for qualitative and quantitative analysis for biotech maize event 3272 and LY 038 on the basis of the 3' flanking regions, respectively. The qualitative primers confirmed the specificity by a single PCR product and sensitivity to 0.05% as a limit of detection (LOD). Simplex and duplex quantitative methods were also developed using TaqMan real-time PCR. One synthetic plasmid was constructed from two taxon-specific DNA sequences of maize and two event-specific 3' flanking DNA sequences of event 3272 and LY 038 as reference molecules. In-house validation of the quantitative methods was performed using six levels of mixing samples, from 0.1 to 10.0%. As a result, the biases from the true value and the relative deviations were all within the range of +/-30%. Limits of quantitation (LOQs) of the quantitative methods were all 0.1% for simplex real-time PCRs of event 3272 and LY 038 and 0.5% for duplex real-time PCR of LY 038. This study reports that event-specific analytical methods were applicable for qualitative and quantitative analysis for biotech maize event 3272 and LY 038.

Can narrow-bandwidth light from UV-A to green alter secondary plant metabolism and increase Brassica plant defenses against aphids?

PubMed

Rechner, Ole; Neugart, Susanne; Schreiner, Monika; Wu, Sasa; Poehling, Hans-Michael

2017-01-01

Light of different wavelengths is essential for plant growth and development. Short-wavelength radiation such as UV can shift the composition of flavonoids, glucosinolates, and other plant metabolites responsible for enhanced defense against certain herbivorous insects. The intensity of light-induced, metabolite-based resistance is plant- and insect species-specific and depends on herbivore feeding guild and specialization. The increasing use of light-emitting diodes (LEDs) in horticultural plant production systems in protected environments enables the creation of tailor-made light scenarios for improved plant cultivation and induced defense against herbivorous insects. In this study, broccoli (Brassica oleracea var. italica) plants were grown in a climate chamber under broad spectra photosynthetic active radiation (PAR) and were additionally treated with the following narrow-bandwidth light generated with LEDs: UV-A (365 nm), violet (420 nm), blue (470 nm), or green (515 nm). We determined the influence of narrow-bandwidth light on broccoli plant growth, secondary plant metabolism (flavonol glycosides and glucosinolates), and plant-mediated light effects on the performance and behavior of the specialized cabbage aphid Brevicoryne brassicae. Green light increased plant height more than UV-A, violet, or blue LED treatments. Among flavonol glycosides, specific quercetin and kaempferol glycosides were increased under violet light. The concentration of 3-indolylmethyl glucosinolate in plants was increased by UV-A treatment. B. brassicae performance was not influenced by the different light qualities, but in host-choice tests, B. brassicae preferred previously blue-illuminated plants (but not UV-A-, violet-, or green-illuminated plants) over control plants.

Leaf Colour as a Signal of Chemical Defence to Insect Herbivores in Wild Cabbage (Brassica oleracea).

PubMed

Green, Jonathan P; Foster, Rosie; Wilkins, Lucas; Osorio, Daniel; Hartley, Susan E

2015-01-01

Leaf colour has been proposed to signal levels of host defence to insect herbivores, but we lack data on herbivory, leaf colour and levels of defence for wild host populations necessary to test this hypothesis. Such a test requires measurements of leaf spectra as they would be sensed by herbivore visual systems, as well as simultaneous measurements of chemical defences and herbivore responses to leaf colour in natural host-herbivore populations. In a large-scale field survey of wild cabbage (Brassica oleracea) populations, we show that variation in leaf colour and brightness, measured according to herbivore spectral sensitivities, predicts both levels of chemical defences (glucosinolates) and abundance of specialist lepidopteran (Pieris rapae) and hemipteran (Brevicoryne brassicae) herbivores. In subsequent experiments, P. rapae larvae achieved faster growth and greater pupal mass when feeding on plants with bluer leaves, which contained lower levels of aliphatic glucosinolates. Glucosinolate-mediated effects on larval performance may thus contribute to the association between P. rapae herbivory and leaf colour observed in the field. However, preference tests found no evidence that adult butterflies selected host plants based on leaf coloration. In the field, B. brassicae abundance varied with leaf brightness but greenhouse experiments were unable to identify any effects of brightness on aphid preference or performance. Our findings suggest that although leaf colour reflects both levels of host defences and herbivore abundance in the field, the ability of herbivores to respond to colour signals may be limited, even in species where performance is correlated with leaf colour.

Can narrow-bandwidth light from UV-A to green alter secondary plant metabolism and increase Brassica plant defenses against aphids?

PubMed Central

Neugart, Susanne; Schreiner, Monika; Wu, Sasa; Poehling, Hans-Michael

2017-01-01

Light of different wavelengths is essential for plant growth and development. Short-wavelength radiation such as UV can shift the composition of flavonoids, glucosinolates, and other plant metabolites responsible for enhanced defense against certain herbivorous insects. The intensity of light-induced, metabolite-based resistance is plant- and insect species-specific and depends on herbivore feeding guild and specialization. The increasing use of light-emitting diodes (LEDs) in horticultural plant production systems in protected environments enables the creation of tailor-made light scenarios for improved plant cultivation and induced defense against herbivorous insects. In this study, broccoli (Brassica oleracea var. italica) plants were grown in a climate chamber under broad spectra photosynthetic active radiation (PAR) and were additionally treated with the following narrow-bandwidth light generated with LEDs: UV-A (365 nm), violet (420 nm), blue (470 nm), or green (515 nm). We determined the influence of narrow-bandwidth light on broccoli plant growth, secondary plant metabolism (flavonol glycosides and glucosinolates), and plant-mediated light effects on the performance and behavior of the specialized cabbage aphid Brevicoryne brassicae. Green light increased plant height more than UV-A, violet, or blue LED treatments. Among flavonol glycosides, specific quercetin and kaempferol glycosides were increased under violet light. The concentration of 3-indolylmethyl glucosinolate in plants was increased by UV-A treatment. B. brassicae performance was not influenced by the different light qualities, but in host-choice tests, B. brassicae preferred previously blue-illuminated plants (but not UV-A-, violet-, or green-illuminated plants) over control plants. PMID:29190278

Leaf Colour as a Signal of Chemical Defence to Insect Herbivores in Wild Cabbage (Brassica oleracea)

PubMed Central

Wilkins, Lucas; Osorio, Daniel; Hartley, Susan E.

2015-01-01

Leaf colour has been proposed to signal levels of host defence to insect herbivores, but we lack data on herbivory, leaf colour and levels of defence for wild host populations necessary to test this hypothesis. Such a test requires measurements of leaf spectra as they would be sensed by herbivore visual systems, as well as simultaneous measurements of chemical defences and herbivore responses to leaf colour in natural host-herbivore populations. In a large-scale field survey of wild cabbage (Brassica oleracea) populations, we show that variation in leaf colour and brightness, measured according to herbivore spectral sensitivities, predicts both levels of chemical defences (glucosinolates) and abundance of specialist lepidopteran (Pieris rapae) and hemipteran (Brevicoryne brassicae) herbivores. In subsequent experiments, P. rapae larvae achieved faster growth and greater pupal mass when feeding on plants with bluer leaves, which contained lower levels of aliphatic glucosinolates. Glucosinolate-mediated effects on larval performance may thus contribute to the association between P. rapae herbivory and leaf colour observed in the field. However, preference tests found no evidence that adult butterflies selected host plants based on leaf coloration. In the field, B. brassicae abundance varied with leaf brightness but greenhouse experiments were unable to identify any effects of brightness on aphid preference or performance. Our findings suggest that although leaf colour reflects both levels of host defences and herbivore abundance in the field, the ability of herbivores to respond to colour signals may be limited, even in species where performance is correlated with leaf colour. PMID:26353086

Hybrid quantitative MRI using chemical shift displacement and recovery-based simultaneous water and lipid imaging: A preliminary study.

PubMed

Ohno, Naoki; Miyati, Tosiaki; Suzuki, Shuto; Kan, Hirohito; Aoki, Toshitaka; Nakamura, Yoshitaka; Hiramatsu, Yuki; Kobayashi, Satoshi; Gabata, Toshifumi

2018-07-01

To suppress olefinic signals and enable simultaneous and quantitative estimation of multiple functional parameters associated with water and lipid, we investigated a modified method using chemical shift displacement and recovery-based separation of lipid tissue (SPLIT) involving acquisitions with different inversion times (TIs), echo times (TEs), and b-values. Single-shot diffusion echo-planar imaging (SSD-EPI) with multiple b-values (0-3000 s/mm 2 ) was performed without fat suppression to separate water and lipid images using the chemical shift displacement of lipid signals in the phase-encoding direction. An inversion pulse (TI = 292 ms) was applied to SSD-EPI to remove olefinic signals. Consecutively, SSD-EPI (b = 0 s/mm 2 ) was performed with TI = 0 ms and TE = 31.8 ms for T 1 and T 2 measurements, respectively. Under these conditions, transverse water and lipid images at the maximum diameter of the right calf were obtained in six healthy subjects. T 1 , T 2 , and the apparent diffusion coefficients (ADC) were then calculated for the tibialis anterior (TA), gastrocnemius (GM), and soleus (SL) muscles, tibialis bone marrow (TB), and subcutaneous fat (SF). Perfusion-related (D*) and restricted diffusion coefficients (D) were calculated for the muscles. Lastly, the lipid fractions (LF) of the muscles were determined after T 1 and T 2 corrections. The modified SPLIT method facilitated sufficient separation of water and lipid images of the calf, and the inversion pulse with TI of 292 ms effectively suppressed olefinic signals. All quantitative parameters obtained with the modified SPLIT method were found to be in general agreement with those previously reported in the literature. The modified SPLIT technique enabled sufficient suppression of olefinic signals and simultaneous acquisition of quantitative parameters including diffusion, perfusion, T 1 and T 2 relaxation times, and LF. Copyright © 2018. Published by Elsevier Inc.

Understanding Gas Phase Modifier Interactions in Rapid Analysis by Differential Mobility-Tandem Mass Spectrometry

NASA Astrophysics Data System (ADS)

Kafle, Amol; Coy, Stephen L.; Wong, Bryan M.; Fornace, Albert J.; Glick, James J.; Vouros, Paul

2014-07-01

A systematic study involving the use and optimization of gas-phase modifiers in quantitative differential mobility-mass spectrometry (DMS-MS) analysis is presented using nucleoside-adduct biomarkers of DNA damage as an important reference point for analysis in complex matrices. Commonly used polar protic and polar aprotic modifiers have been screened for use against two deoxyguanosine adducts of DNA: N-(deoxyguanosin-8-yl)-4-aminobiphenyl (dG-C8-4-ABP) and N-(deoxyguanosin-8-y1)-2-amino-l-methyl-6-phenylimidazo[4,5-b]pyridine (dG-C8-PhIP). Particular attention was paid to compensation voltage (CoV) shifts, peak shapes, and product ion signal intensities while optimizing the DMS-MS conditions. The optimized parameters were then applied to rapid quantitation of the DNA adducts in calf thymus DNA. After a protein precipitation step, adduct levels corresponding to less than one modification in 106 normal DNA bases were detected using the DMS-MS platform. Based on DMS fundamentals and ab initio thermochemical results, we interpret the complexity of DMS modifier responses in terms of thermal activation and the development of solvent shells. At very high bulk gas temperature, modifier dipole moment may be the most important factor in cluster formation and cluster geometry, but at lower temperatures, multi-neutral clusters are important and less predictable. This work provides a useful protocol for targeted DNA adduct quantitation and a basis for future work on DMS modifier effects.

Understanding gas phase modifier interactions in rapid analysis by Differential Mobility-Tandem Mass Spectrometry

PubMed Central

Kafle, Amol; Coy, Stephen L.; Wong, Bryan M.; Fornace, Albert J.; Glick, James J.; Vouros, Paul

2014-01-01

A systematic study involving the use and optimization of gas phase modifiers in quantitative differential mobility- mass spectrometry (DMS-MS) analysis is presented using mucleoside-adduct biomarkers of DNA damage as an important reference point for analysis in complex matrices. Commonly used polar protic and polar aprotic modifiers have been screened for use against two deoxyguanosine adducts of DNA: N-(deoxyguanosin-8-yl)-4-aminobiphenyl (dG-C8-4-ABP) and N-(deoxyguanosin-8-y1)-2-amino-l-methyl-6-phenylimidazo[4,5-b]pyridine (dG-C8-PhIP). Particular attention was paid to compensation voltage (CoV) shifts, peak shapes and product ion signal intensities while optimizing the DMS-MS conditions. The optimized parameters were then applied to rapid quantitation of the DNA adducts in calf thymus DNA. After a protein precipitation step, adduct levels corresponding to less than one modification in 106 normal DNA bases were detected using the DMS-MS platform. Based on DMS fundamentals and ab-initio thermochemical results we interpret the complexity of DMS modifier responses in terms of thermal activation and the development of solvent shells. At very high bulk gas temperature, modifier dipole moment may be the most important factor in cluster formation and cluster geometry in mobility differences, but at lower temperatures multi-neutral clusters are important and less predictable. This work provides a useful protocol for targeted DNA adduct quantitation and a basis for future work on DMS modifier effects. PMID:24452298

Partial deficiency of isoleucine impairs root development and alters transcript levels of the genes involved in branched-chain amino acid and glucosinolate metabolism in Arabidopsis

PubMed Central

Liu, Dong

2013-01-01

Isoleucine is one of the branched-chain amino acids (BCAAs) that are essential substrates for protein synthesis in all organisms. Although the metabolic pathway for isoleucine has been well characterized in higher plants, it is not known whether it plays a specific role in plant development. In this study, an Arabidopsis mutant, lib (low isoleucine biosynthesis), that has defects in both cell proliferation and cell expansion processes during root development, was characterized. The lib mutant carries a T-DNA insertion in the last exon of the OMR1 gene that encodes a threonine deaminase/dehydratase (TD). TD catalyses the deamination and dehydration of threonine, which is the first and also the committed step in the biosynthesis of isoleucine. This T-DNA insertion results in a partial deficiency of isoleucine in lib root tissues but it does not affect its total protein content. Application of exogenous isoleucine or introduction of a wild-type OMR1 gene into the lib mutant can completely rescue the mutant phenotypes. These results reveal an important role for isoleucine in plant development. In addition, microarray analysis indicated that the partial deficiency of isoleucine in the lib mutant triggers a decrease in transcript levels of the genes encoding the major enzymes involved in the BCAA degradation pathway; the analysis also indicated that many genes involved in the biosynthesis of methionine-derived glucosinolates are up-regulated. PMID:23230023

Endogenous Reference Genes and Their Quantitative Real-Time PCR Assays for Genetically Modified Bread Wheat (Triticum aestivum L.) Detection.

PubMed

Yang, Litao; Quan, Sheng; Zhang, Dabing

2017-01-01

Endogenous reference genes (ERG) and their derivate analytical methods are standard requirements for analysis of genetically modified organisms (GMOs). Development and validation of suitable ERGs is the primary step for establishing assays that monitoring the genetically modified (GM) contents in food/feed samples. Herein, we give a review of the ERGs currently used for GM wheat analysis, such as ACC1, PKABA1, ALMT1, and Waxy-D1, as well as their performances in GM wheat analysis. Also, we discussed one model for developing and validating one ideal RG for one plant species based on our previous research work.

Methyl Jasmonate and 1-Methylcyclopropene Treatment Effects on Quinone Reductase Inducing Activity and Post-Harvest Quality of Broccoli

PubMed Central

Ku, Kang Mo; Choi, Jeong Hee; Kim, Hyoung Seok; Kushad, Mosbah M.; Jeffery, Elizabeth H.; Juvik, John A.

2013-01-01

Effect of pre-harvest methyl jasmonate (MeJA) and post-harvest 1-methylcyclopropene (1-MCP) treatments on broccoli floret glucosinolate (GS) concentrations and quinone reductase (QR, an in vitro anti-cancer biomarker) inducing activity were evaluated two days prior to harvest, at harvest and at 10, 20, and 30 days of post-harvest storage at 4 °C. MeJA treatments four days prior to harvest of broccoli heads was observed to significantly increase floret ethylene biosynthesis resulting in chlorophyll catabolism during post-harvest storage and reduced product quality. Post-harvest treatment with 1-methylcyclopropene (1-MCP), which competitively binds to protein ethylene receptors, maintained post-harvest floret chlorophyll concentrations and product visual quality in both control and MeJA-treated broccoli. Transcript abundance of BoPPH, a gene which is responsible for the synthesis of pheophytinase, the primary enzyme associated with chlorophyll catabolism in broccoli, was reduced by 1-MCP treatment and showed a significant, negative correlation with floret chlorophyll concentrations. The GS, glucobrassicin, neoglucobrassicin, and gluconasturtiin were significantly increased by MeJA treatments. The products of some of the GS from endogenous myrosinase hydrolysis [sulforaphane (SF), neoascorbigen (NeoASG), N-methoxyindole-3-carbinol (NI3C), and phenethyl isothiocyanate (PEITC)] were also quantified and found to be significantly correlated with QR. Sulforaphane, the isothiocyanate hydrolysis product of the GS glucoraphanin, was found to be the most potent QR induction agent. Increased sulforaphane formation from the hydrolysis of glucoraphanin was associated with up-regulated gene expression of myrosinase (BoMyo) and the myrosinase enzyme co-factor gene, epithiospecifier modifier1 (BoESM1). This study demonstrates the combined treatment of MeJA and 1-MCP increased QR activity without post-harvest quality loss. PMID:24146962

Development of a screening method for genetically modified soybean by plasmid-based quantitative competitive polymerase chain reaction.

PubMed

Shimizu, Eri; Kato, Hisashi; Nakagawa, Yuki; Kodama, Takashi; Futo, Satoshi; Minegishi, Yasutaka; Watanabe, Takahiro; Akiyama, Hiroshi; Teshima, Reiko; Furui, Satoshi; Hino, Akihiro; Kitta, Kazumi

2008-07-23

A novel type of quantitative competitive polymerase chain reaction (QC-PCR) system for the detection and quantification of the Roundup Ready soybean (RRS) was developed. This system was designed based on the advantage of a fully validated real-time PCR method used for the quantification of RRS in Japan. A plasmid was constructed as a competitor plasmid for the detection and quantification of genetically modified soy, RRS. The plasmid contained the construct-specific sequence of RRS and the taxon-specific sequence of lectin1 (Le1), and both had 21 bp oligonucleotide insertion in the sequences. The plasmid DNA was used as a reference molecule instead of ground seeds, which enabled us to precisely and stably adjust the copy number of targets. The present study demonstrated that the novel plasmid-based QC-PCR method could be a simple and feasible alternative to the real-time PCR method used for the quantification of genetically modified organism contents.

Quantitative analysis by UV-Vis absorption spectroscopy of amino groups attached to the surface of carbon-based nanoparticles

NASA Astrophysics Data System (ADS)

Saraswati, T. E.; Astuti, A. R.; Rismana, N.

2018-03-01

Carbon-based nanoparticles must be modified due to their wide array of applications, especially when they are used as biomaterials. After modifying, quantitative analysis of the functional group is essential to evaluate a number of the available functional groups applied for further functionalization. In this study, we modified the carbon-based nanoparticles by amino group using submerged arc discharge in different liquids. The attached amino groups were then characterised and quantified by UV-Vis spectroscopy. This amino group functionalization was also confirmed by Fourier transform infrared (FTIR) spectra. The FTIR spectra of amine-modified nanoparticles show the definitive absorption peaks of N—H amine, C—H, C=O, C—N and Fe—O at 3418.97; 3000–2850 1700–1600 1400–1100 and 480-550 cm-1, respectively. The amine groups have different performance signals between the amine-modified and unmodified nanoparticles. The FTIR spectra results were correlated with the UV-Vis absorption spectroscopy method using acidic methyl orange. The UV-Vis absorption spectroscopy shows that the absorbance of methyl orange represented to amino groups number was 1.3 times higher when the pH of the solution was increased. The absorbance intensity was then used to estimate the quantity of amine groups attached.

Effect of substitution of groundnut with high glucosinolate mustard (Brassica juncea) meal on nutrient utilization, growth, vital organ weight and blood composition of lambs.

PubMed

Tripathi, M K.; Mishra, A S.; Misra, A K.; Mondal, D; Karim, S A.

2001-03-01

Twenty-four 14-day-old weaner Avivastra (Russian MerinoxNali) male lambs were maintained for 180 days on ad libidum Cenchrus (Cenchrus ciliaris) hay and concentrate mixture (CM) contained groundnut meal (control) and mustard meal (MM group) as major protein source. The two CMs were isonitrogenous (21% CP) and isocaloric (2.78McalMEkg(-1) DM), while, CM fed to MM group contained 24.6mg glucosinolatesg(-1) DM. Digestibility of nutrients was similar (P>0.05) in the two groups except for CP and hemicellulose, which was higher (P<0.05) in control. Urinary N loss was higher (P<0.01) in control than in MM group, whereas N retention (% of N intake and absorbed) was higher (P<0.01) in MM group compared to control. Dry matter (gkg(-1)BW) and glucosinolate intakes were higher in MM fed group, whereas DCP and ME intakes were similar (P>0.05) in the two groups. Average daily gain (ADG) was, however, 22% higher (P<0.01) in control than in MM group. Hemoglobin and albumin contents were lower (P<0.01) in MM group than in control. Serum thiocyanate content was 26.7µgg(-1) in MM fed group, while it was not detected in control group. Thyroid weight was higher (P<0.01) while liver and kidney weights were lower (P<0.01) in MM group. Meat from dissected carcass of control group contained more protein and less fat, whereas the reverse was noticed in MM group. It is concluded that feeding mustard meal as protein supplement reduced growth rate and induced iodine deficiency. Carcass of lambs fed mustard meal had more fat and less protein.

Altered seed oil and glucosinolate levels in transgenic plants overexpressing the Brassica napus SHOOTMERISTEMLESS gene.

PubMed

Elhiti, Mohamed; Yang, Cunchun; Chan, Ainsley; Durnin, Douglas C; Belmonte, Mark F; Ayele, Belay T; Tahir, Muhammad; Stasolla, Claudio

2012-07-01

SHOOTMERISTEMLESS (STM) is a homeobox gene conserved among plant species which is required for the formation and maintenance of the shoot meristem by suppressing differentiation and maintaining an undetermined cell fate within the apical pole. To assess further the role of this gene during seed storage accumulation, transgenic Brassica napus (Bn) plants overexpressing or down-regulating BnSTM under the control of the 35S promoter were generated. Overexpression of BnSTM increased seed oil content without affecting the protein and sucrose level. These changes were accompanied by the induction of genes encoding several transcription factors promoting fatty acid (FA) synthesis: LEAFY COTYLEDON1 (BnLEC1), BnLEC2, and WRINKLE1 (BnWRI1). In addition, expression of key representative enzymes involved in sucrose metabolism, glycolysis, and FA biosynthesis was up-regulated in developing seeds ectopically expressing BnSTM. These distinctive expression patterns support the view of an increased carbon flux to the FA biosynthetic pathway in developing transformed seeds. The overexpression of BnSTM also resulted in a desirable reduction of seed glucosinolate (GLS) levels ascribed to a transcriptional repression of key enzymes participating in the GLS biosynthetic pathway, and possibly to the differential utilization of common precursors for GLS and indole-3-acetic acid synthesis. No changes in oil and GLS levels were observed in lines down-regulating BnSTM. Taken together, these findings provide evidence for a novel function for BnSTM in promoting desirable changes in seed oil and GLS levels when overexpressed in B. napus plants, and demonstrate that this gene can be used as a target for genetic improvement of oilseed species.

CYP79D enzymes contribute to jasmonic acid-induced formation of aldoximes and other nitrogenous volatiles in two Erythroxylum species.

PubMed

Luck, Katrin; Jirschitzka, Jan; Irmisch, Sandra; Huber, Meret; Gershenzon, Jonathan; Köllner, Tobias G

2016-10-04

Amino acid-derived aldoximes and nitriles play important roles in plant defence. They are well-known as precursors for constitutive defence compounds such as cyanogenic glucosides and glucosinolates, but are also released as volatiles after insect feeding. Cytochrome P450 monooxygenases (CYP) of the CYP79 family catalyze the formation of aldoximes from the corresponding amino acids. However, the majority of CYP79s characterized so far are involved in cyanogenic glucoside or glucosinolate biosynthesis and only a few have been reported to be responsible for nitrogenous volatile production. In this study we analysed and compared the jasmonic acid-induced volatile blends of two Erythroxylum species, the cultivated South American crop species E. coca and the African wild species E. fischeri. Both species produced different nitrogenous compounds including aliphatic aldoximes and an aromatic nitrile. Four isolated CYP79 genes (two from each species) were heterologously expressed in yeast and biochemically characterized. CYP79D62 from E. coca and CYP79D61 and CYP79D60 from E. fischeri showed broad substrate specificity in vitro and converted L-phenylalanine, L-isoleucine, L-leucine, L-tryptophan, and L-tyrosine into the respective aldoximes. In contrast, recombinant CYP79D63 from E. coca exclusively accepted L-tryptophan as substrate. Quantitative real-time PCR revealed that CYP79D60, CYP79D61, and CYP79D62 were significantly upregulated in jasmonic acid-treated Erythroxylum leaves. The kinetic parameters of the enzymes expressed in vitro coupled with the expression patterns of the corresponding genes and the accumulation and emission of (E/Z)-phenylacetaldoxime, (E/Z)-indole-3-acetaldoxime, (E/Z)-3-methylbutyraldoxime, and (E/Z)-2-methylbutyraldoxime in jasmonic acid-treated leaves suggest that CYP79D60, CYP79D61, and CYP79D62 accept L-phenylalanine, L-leucine, L-isoleucine, and L-tryptophan as substrates in vivo and contribute to the production of volatile and semi-volatile nitrogenous defence compounds in E. coca and E. fischeri.

Oilseed rape seeds with ablated defence cells of the glucosinolate–myrosinase system. Production and characteristics of double haploid MINELESS plants of Brassica napus L.

PubMed Central

Ahuja, Ishita; Borgen, Birgit Hafeld; Hansen, Magnor; Honne, Bjørn Ivar; Müller, Caroline; Rohloff, Jens; Rossiter, John Trevor; Bones, Atle Magnar

2011-01-01

Oilseed rape and other crop plants of the family Brassicaceae contain a unique defence system known as the glucosinolate–myrosinase system or the ‘mustard oil bomb’. The ‘mustard oil bomb’ which includes myrosinase and glucosinolates is triggered by abiotic and biotic stress, resulting in the formation of toxic products such as nitriles and isothiocyanates. Myrosinase is present in specialist cells known as ‘myrosin cells’ and can also be known as toxic mines. The myrosin cell idioblasts of Brassica napus were genetically reprogrammed to undergo controlled cell death (ablation) during seed development. These myrosin cell-free plants have been named MINELESS as they lack toxic mines. This has led to the production of oilseed rape with a significant reduction both in myrosinase levels and in the hydrolysis of glucosinolates. Even though the myrosinase activity in MINELESS was very low compared with the wild type, variation was observed. This variability was overcome by producing homozygous seeds. A microspore culture technique involving non-fertile haploid MINELESS plants was developed and these plants were treated with colchicine to produce double haploid MINELESS plants with full fertility. Double haploid MINELESS plants had significantly reduced myrosinase levels and glucosinolate hydrolysis products. Wild-type and MINELESS plants exhibited significant differences in growth parameters such as plant height, leaf traits, matter accumulation, and yield parameters. The growth and developmental pattern of MINELESS plants was relatively slow compared with the wild type. The characteristics of the pure double haploid MINELESS plant are described and its importance for future biochemical, agricultural, dietary, functional genomics, and plant defence studies is discussed. PMID:21778185

Both heavy metal-amendment of soil and aphid-infestation increase Cd and Zn concentrations in phloem exudates of a metal-hyperaccumulating plant.

PubMed

Stolpe, Clemens; Giehren, Franziska; Krämer, Ute; Müller, Caroline

2017-07-01

Plants that are able to hyperaccumulate heavy metals show increased concentrations of these metals in their leaf tissue. However, little is known about the concentrations of heavy metals and of organic defence metabolites in the phloem sap of these plants in response to either heavy metal-amendment of the soil or biotic challenges such as aphid-infestation. In this study, we investigated the effects of heavy metal-exposure and of aphid-infestation on phloem exudate composition of the metal hyperaccumulator species Arabidopsis halleri L. O'Kane & Al-Shehbaz (Brassicaceae). The concentrations of elements and of organic defence compounds, namely glucosinolates, were measured in phloem exudates of young and old (mature) leaves of plants challenged either by amendment of the soil with cadmium and zinc and/or by an infestation with the generalist aphid Myzus persicae. Metal-amendment of the soil led to increased concentrations of Cd and Zn, but also of two other elements and one indole glucosinolate, in phloem exudates. This enhanced defence in the phloem sap of heavy metal-hyperaccumulating plants can thus potentially act as effective protection against aphids, as predicted by the elemental defence hypothesis. Aphid-infestation also caused enhanced Cd and Zn concentrations in phloem exudates. This result provides first evidence that metal-hyperaccumulating plants can increase heavy metal concentrations tissue-specifically in response to an attack by phloem-sucking herbivores. Overall, the concentrations of most elements, including the heavy metals, and glucosinolates were higher in phloem exudates of young leaves than in those of old leaves. This defence distribution highlights that the optimal defence theory, which predicts more valuable tissue to be better defended, is applicable for both inorganic and organic defences. Copyright © 2017 Elsevier Ltd. All rights reserved.

Epigenetics of drought-induced trans-generational plasticity: consequences for range limit development

PubMed Central

Alsdurf, Jacob; Anderson, Cynthia; Siemens, David H.

2016-01-01

Genetic variation gives plants the potential to adapt to stressful environments that often exist beyond their geographic range limits. However, various genetic, physiological or developmental constraints might prevent the process of adaptation. Alternatively, environmentally induced epigenetic changes might sustain populations for several generations in stressful areas across range boundaries, but previous work on Boechera stricta, an upland mustard closely related to Arabidopsis, documented a drought-induced trans-generational plastic trade-off that could contribute to range limit development. Offspring of parents who were drought treated had higher drought tolerance, but lower levels of glucosinolate toxins. Both drought tolerance and defence are thought to be needed to expand the range to lower elevations. Here, we used methylation-sensitive amplified fragment length polymorphisms to determine whether environmentally induced DNA methylation and thus epigenetics could be a mechanism involved in the observed trans-generational plastic trade-off. We compared 110 offspring from the same self-fertilizing lineages whose parents were exposed to experimental drought stress treatments in the laboratory. Using three primer combinations, 643 polymorphic epi-loci were detected. Discriminant function analysis (DFA) on the amount of methylation detected resulted in significant combinations of epi-loci that distinguished the parent drought treatments in the offspring. Principal component (PC) and univariate association analyses also detected the significant differences, even after controlling for lineage, planting flat, developmental differences and multiple testing. Univariate tests also indicated significant associations between the amount of methylation and drought tolerance or glucosinolate toxin concentration. One epi-locus that was implicated in DFA, PC and univariate association analysis may be directly involved in the trade-off because increased methylation at this site on the genome decreased drought tolerance, but increased glucosinolate concentration. PMID:26685218

Potential of cultivar and crop management to affect phytochemical content in winter-grown sprouting broccoli (Brassica oleracea L. var. italica).

PubMed

Reilly, Kim; Valverde, Juan; Finn, Leo; Rai, Dilip K; Brunton, Nigel; Sorensen, Jens C; Sorensen, Hilmer; Gaffney, Michael

2014-01-30

Variety and crop management strategies affect the content of bioactive compounds (phenolics, flavonoids and glucosinolates) in green broccoli (calabrese) types, which are cultivated during summer and autumn in temperate European climates. Sprouting broccoli types are morphologically distinct and are grown over the winter season and harvested until early spring. Thus they show considerable potential for development as an import substitution crop for growers and consumers during the 'hungry gap' of early spring. The present study investigated the effect of variety and management practices on phytochemical content in a range of sprouting broccoli varieties. Yields were significantly higher in white sprouting broccoli varieties. Levels of phenolics and flavonoids were in the range 81.64-297.65 and 16.95-104.80 mg 100 g⁻¹ fresh weight, respectively, depending on year and cultivar, and were highest in variety 'TZ 5052' in both years. In-row spacing did not affect flavonoid content. Phenolic and flavonoid content generally increased with increasing floret maturity and levels were high in edible portions of the crop. Crop wastes (leaf and flower) contained 145.9-239.3 and 21.5-116.6 mg 100 g⁻¹ fresh weight total phenolics and flavonoids, respectively, depending on cultivar, tissue and year. Climatic factors had a significant effect on phenolic and flavonoid content. Levels of total and some individual glucosinolates were higher in sprouting broccoli than in the green broccoli variety 'Ironman'. Levels of total phenolics, flavonoids and glucosinolates are higher in sprouting than green broccoli types. Sprouting broccoli represents an excellent source of dietary bioactive compounds. © 2013 Society of Chemical Industry.

Dose-response effect of Red Maca (Lepidium meyenii) on benign prostatic hyperplasia induced by testosterone enanthate.

PubMed

Gasco, M; Villegas, L; Yucra, S; Rubio, J; Gonzales, G F

2007-08-01

The main goal of this study was to determine the effect of a freeze-dried aqueous extract of the red variety of Lepidium meyenii (Red Maca) on testosterone-induced benign prostatic hyperplasia (BPH) in adult rats of the Holtzman strain. Rats were treated with freeze-dried aqueous extract of Red Maca at doses of 0, 0.01, 0.05, 0.1, and 0.5 g/kg body wt. A positive control group received Finasteride (0.6 mg/kg body wt.). After treatment, the animals were sacrificed, and the ventral prostate was extracted, and weighed. HPLC was used to determine the presence of glucosinolates in Red Maca. The prostate weight diminished in a dose-dependent fashion in rats treated with Red Maca. The effect of Red Maca was better than that observed with Finasteride. Finasteride, but not Red Maca, reduced seminal vesicles weight. Analysis of the HPLC indicated the presence of benzyl glucosinolate (Glucotropaeolin) with a content of 0.639%. Serum testosterone levels were not affected by Red Maca. Moreover, serum testosterone levels were not related to prostate or seminal vesicles weight in rats treated with vehicle and Red Maca. In conclusion, Red Maca administered orally in rats seems to exert an inhibitory effect at a level post DHT conversion, on the BPH-induced experimentally, although a direct measure of reductase action would still be required.

Impact of thermal processing on sulforaphane yield from broccoli (Brassica oleracea L. var. italica)

USDA-ARS?s Scientific Manuscript database

In broccoli, sulforaphane forms when the glucosinolate glucoraphanin is hydrolyzed by the endogenous plant thiohydrolase myrosinase. A myrosinase cofactor directs hydrolysis away from formation of bioactive sulforaphane and toward an inactive product, sulforaphane nitrile. The cofactor is more hea...

Bioherbicidal activity of Sinapis alba seed meal extracts

USDA-ARS?s Scientific Manuscript database

Although seed meal from yellow mustard (Sinapis alba L.) is a potential tool for controlling weeds as a consequence of contained glucosinolate substrates that are enzymatically hydrolyzed to produce phytotoxic products, use is limited by batch-to-batch variability and logistical constraints. Our obj...

Screening and identification of major phytochemical compounds in seeds, sprouts and leaves of Tuscan black kale Brassica oleracea (L.) ssp acephala (DC) var. sabellica L.

PubMed

Giorgetti, Lucia; Giorgi, Gianluca; Cherubini, Edoardo; Gervasi, Pier Giovanni; Della Croce, Clara Maria; Longo, Vincenzo; Bellani, Lorenza

2018-07-01

We report the spectrophotometric determination of total polyphenols, flavonoids, glucosinolates and antioxidant activity in seeds, seedlings and leaves of Tuscan black kale. The highest content of phytochemicals was observed in 10 days sprouts and antioxidant activity was maximum in 2, 4 days seedlings. Identification and characterisation of phytochemicals were performed by mass spectrometry (MS), high resolution and tandem MS with electrospray ionisation mode. Low-molecular-weight metabolites were evidenced in seeds while metabolites at high m/z range were detected in cotyledons and leaves. MS spectra evidenced different phenolic compounds (flavonoid caffeoyl glucose, hydroxycinnamic acid sinapine) and glucosinolates (glucoerucin, glucobrassicin and glucoraphanin) in function of developmental stage; galactolipids ω3 and ω6 were observed in leaves. Identification of stages with the highest phytochemicals content encourages the consumption of black kale sprouts and young leaves. Our research can support food and pharmaceutical industries for production of health promoting products from black kale.

An evolutionarily young defense metabolite influences the root growth of plants via the ancient TOR signaling pathway.

PubMed

Malinovsky, Frederikke Gro; Thomsen, Marie-Louise F; Nintemann, Sebastian J; Jagd, Lea Møller; Bourgine, Baptiste; Burow, Meike; Kliebenstein, Daniel J

2017-12-12

To optimize fitness a plant should monitor its metabolism to appropriately control growth and defense. Primary metabolism can be measured by the universally conserved TOR (Target of Rapamycin) pathway to balance growth and development with the available energy and nutrients. Recent work suggests that plants may measure defense metabolites to potentially provide a strategy ensuring fast reallocation of resources to coordinate plant growth and defense. There is little understanding of mechanisms enabling defense metabolite signaling. To identify mechanisms of defense metabolite signaling, we used glucosinolates, an important class of plant defense metabolites. We report novel signaling properties specific to one distinct glucosinolate, 3-hydroxypropylglucosinolate across plants and fungi. This defense metabolite, or derived compounds, reversibly inhibits root growth and development. 3-hydroxypropylglucosinolate signaling functions via genes in the ancient TOR pathway. If this event is not unique, this raises the possibility that other evolutionarily new plant metabolites may link to ancient signaling pathways.

An evolutionarily young defense metabolite influences the root growth of plants via the ancient TOR signaling pathway

PubMed Central

Malinovsky, Frederikke Gro; Thomsen, Marie-Louise F; Nintemann, Sebastian J; Jagd, Lea Møller; Bourgine, Baptiste; Burow, Meike

2017-01-01

To optimize fitness a plant should monitor its metabolism to appropriately control growth and defense. Primary metabolism can be measured by the universally conserved TOR (Target of Rapamycin) pathway to balance growth and development with the available energy and nutrients. Recent work suggests that plants may measure defense metabolites to potentially provide a strategy ensuring fast reallocation of resources to coordinate plant growth and defense. There is little understanding of mechanisms enabling defense metabolite signaling. To identify mechanisms of defense metabolite signaling, we used glucosinolates, an important class of plant defense metabolites. We report novel signaling properties specific to one distinct glucosinolate, 3-hydroxypropylglucosinolate across plants and fungi. This defense metabolite, or derived compounds, reversibly inhibits root growth and development. 3-hydroxypropylglucosinolate signaling functions via genes in the ancient TOR pathway. If this event is not unique, this raises the possibility that other evolutionarily new plant metabolites may link to ancient signaling pathways. PMID:29231169

Attenuation of Carcinogenesis and the Mechanism Underlying by the Influence of Indole-3-carbinol and Its Metabolite 3,3'-Diindolylmethane: A Therapeutic Marvel.

PubMed

Maruthanila, V L; Poornima, J; Mirunalini, S

2014-01-01

Rising evidence provides credible support towards the potential role of bioactive products derived from cruciferous vegetables such as broccoli, cauliflower, kale, cabbage, brussels sprouts, turnips, kohlrabi, bok choy, and radishes. Many epidemiological studies point out that Brassica vegetable protects humans against cancer since they are rich sources of glucosinolates in addition to possessing a high content of flavonoids, vitamins, and mineral nutrients. Indole-3-carbinol (I3C) belongs to the class of compounds called indole glucosinolate, obtained from cruciferous vegetables, and is well-known for tits anticancer properties. In particular, I3C and its dimeric product, 3,3'-diindolylmethane (DIM), have been generally investigated for their value against a number of human cancers in vitro as well as in vivo. This paper reviews an in-depth study of the anticancer activity and the miscellaneous mechanisms underlying the anticarcinogenicity thereby broadening its therapeutic marvel.

Protective Effect of Glucosinolates Hydrolytic Products in Neurodegenerative Diseases (NDDs).

PubMed

Jaafaru, Mohammed Sani; Abd Karim, Nurul Ashikin; Enas, Mohamad Eliaser; Rollin, Patrick; Mazzon, Emanuela; Abdull Razis, Ahmad Faizal

2018-05-08

Crucifer vegetables, Brassicaceae and other species of the order Brassicales, e.g., Moringaceae that are commonly consumed as spice and food, have been reported to have potential benefits for the treatment and prevention of several health disorders. Though epidemiologically inconclusive, investigations have shown that consumption of those vegetables may result in reducing and preventing the risks associated with neurodegenerative disease development and may also exert other biological protections in humans. The neuroprotective effects of these vegetables have been ascribed to their secondary metabolites, glucosinolates (GLs), and their related hydrolytic products, isothiocyanates (ITCs) that are largely investigated for their various medicinal effects. Extensive pre-clinical studies have revealed more than a few molecular mechanisms of action elucidating multiple biological effects of GLs hydrolytic products. This review summarizes the most significant and up-to-date in vitro and in vivo neuroprotective actions of sulforaphane (SFN), moringin (MG), phenethyl isothiocyanate (PEITC), 6-(methylsulfinyl) hexyl isothiocyanate (6-MSITC) and erucin (ER) in neurodegenerative diseases.

High-resolution metabolic mapping of cell types in plant roots

PubMed Central

Moussaieff, Arieh; Rogachev, Ilana; Brodsky, Leonid; Malitsky, Sergey; Toal, Ted W.; Belcher, Heather; Yativ, Merav; Brady, Siobhan M.; Benfey, Philip N.; Aharoni, Asaph

2013-01-01

Metabolite composition offers a powerful tool for understanding gene function and regulatory processes. However, metabolomics studies on multicellular organisms have thus far been performed primarily on whole organisms, organs, or cell lines, losing information about individual cell types within a tissue. With the goal of profiling metabolite content in different cell populations within an organ, we used FACS to dissect GFP-marked cells from Arabidopsis roots for metabolomics analysis. Here, we present the metabolic profiles obtained from five GFP-tagged lines representing core cell types in the root. Fifty metabolites were putatively identified, with the most prominent groups being glucosinolates, phenylpropanoids, and dipeptides, the latter of which is not yet explored in roots. The mRNA expression of enzymes or regulators in the corresponding biosynthetic pathways was compared with the relative metabolite abundance. Positive correlations suggest that the rate-limiting steps in biosynthesis of glucosinolates in the root are oxidative modifications of side chains. The current study presents a work flow for metabolomics analyses of cell-type populations. PMID:23476065

Enhancement Of Glucosinolate and Isothiocyanate Profiles in Brassicaceae Crops: Addressing Challenges in Breeding for Cultivation, Storage, and Consumer-Related Traits.

PubMed

Bell, Luke; Wagstaff, Carol

2017-11-01

Glucosinolates (GSLs) and isothiocyanates (ITCs) produced by Brassicaceae plants are popular targets for analysis due to the health benefits associated with them. Breeders aim to increase the concentrations in commercial varieties; however, there are few examples of this. The most well-known is Beneforté broccoli, which has increased glucoraphanin/sulforaphane concentrations compared to those of conventional varieties. It was developed through traditional breeding methods with considerations for processing, consumption, and health made throughout this process. Many studies presented in the literature do not take a holistic approach, and key points about breeding, cultivation methods, postharvest storage, sensory attributes, and consumer preferences are not properly taken into account. In this review, we draw together data for multiple species and address how such factors can influence GSL profiles. We encourage researchers and institutions to engage with industry and consumers to produce research that can be utilized in the improvement of Brassicaceae crops.

Ecotype variability in growth and secondary metabolite profile in Moringa oleifera: impact of sulfur and water availability.

PubMed

Förster, Nadja; Ulrichs, Christian; Schreiner, Monika; Arndt, Nick; Schmidt, Reinhard; Mewis, Inga

2015-03-25

Moringa oleifera is widely cultivated in plantations in the tropics and subtropics. Previous cultivation studies with M. oleifera focused primarily only on leaf yield. In the present study, the content of potentially health-promoting secondary metabolites (glucosinolates, phenolic acids, and flavonoids) were also investigated. Six different ecotypes were grown under similar environmental conditions to identify phenotypic differences that can be traced back to the genotype. The ecotypes TOT4880 (origin USA) and TOT7267 (origin India) were identified as having the best growth performance and highest secondary metabolite production, making them an ideal health-promoting food crop. Furthermore, optimal cultivation conditions-exemplarily on sulfur fertilization and water availability-for achieving high leaf and secondary metabolite yields were investigated for M. oleifera. In general, plant biomass and height decreased under water deficiency compared to normal cultivation conditions, whereas the glucosinolate content increased. The effects depended to a great extent on the ecotype.

The effect of the germination temperature on the phytochemical content of broccoli and rocket sprouts.

PubMed

Ragusa, Lucia; Picchi, Valentina; Tribulato, Alessandro; Cavallaro, Chiara; Lo Scalzo, Roberto; Branca, Ferdinando

2017-06-01

This study investigates the effect of different germination temperatures (10, 20 and 30 °C) on the phytochemical content as well as reducing and antioxidant capacity of broccoli and rocket sprouts. In both seeds and sprouts, the total glucosinolates and ascorbic acid contents did not differ between vegetables, while broccoli exhibited exceptionally higher polyphenols and greater reducing and antioxidant capacity compared to rocket. In both species, an increase in germination temperature positively affected the glucosinolate content. Ascorbic acid increased during germination without a difference among the three tested temperatures. The phenol content in broccoli sprouts increased when they were grown at 30 °C, but the amount decreased at the highest temperatures in rocket. The reducing and antioxidant capacities increased with germination, and higher indexes were detected at 10 °C, particularly in rocket. Different germination temperatures differentiate the health-promoting phytochemical content and antioxidant properties in broccoli and rocket sprouts.

Leaf Metabolic, Genetic, and Morphophysiological Profiles of Cultivated and Wild Rocket Salad (Eruca and Diplotaxis Spp.).

PubMed

Taranto, Francesca; Francese, Gianluca; Di Dato, Francesco; D'Alessandro, Antonietta; Greco, Barbara; Onofaro Sanajà, Vincenzo; Pentangelo, Alfonso; Mennella, Giuseppe; Tripodi, Pasquale

2016-07-27

Rocket salad (Diplotaxis spp., Eruca spp.) is a leafy vegetable rich in health-promoting compounds and widely consumed. In the present study, metabolic profiles of 40 rocket accessions mainly retrieved from gene banks were assessed. Seven glucosinolates (GLSs) and 15 flavonol compounds were detected across genotypes. Dimeric 4-mercaptobutyl-GLS and 4-(β-d-glucopyranosyldisulfanyl)butyl-GLS were the major components of the total glucosinolate content. Flavonols were different between genera, with the exception of isorhamnetin 3,4'-diglucoside. Morphoagronomic traits and color coordinates were also scored. Results showed a negative correlation between color and GLSs, indicating these last as responsible for the increase of the intensity of green and yellow pigments as well as for the darkness of the leaf, whereas agronomic traits showed positive correlation with GLSs. Genetic diversity was assessed using inter simple sequence repeat (ISSR) markers, allowing separation of the accessions on the basis of the species and elucidating the observations made by means of phenotypic data.

Detection of vitamin b1 (thiamine) using modified carbon paste electrodes with polypyrrole

NASA Astrophysics Data System (ADS)

Muppariqoh, N. M.; Wahyuni, W. T.; Putra, B. R.

2017-03-01

Vitamin B1 (thiamine) is oxidized in alkaline medium and can be detected by cyclic voltammetry technique using carbon paste electrode (CPE) as a working electrode. polypyrrole-modified CPE were used in this study to increase sensitivity and selectivity measurement of thiamine. Molecularly imprinted polymers (MIP) of the modified CPE was prepared through electrodeposition of pyrrole. Measurement of thiamine performed in KCl 0.05 M (pH 10, tris buffer) using CPE and the modified CPE gave an optimum condition anodic current of thiamine at 0.3 V, potential range (-1.6_1 V), and scan rate of 100 mV/s. Measurement of thiamine using polypyrrole modified CPE (CPE-MIPpy) showed better result than CPE itself with detection limit of 6.9×10-5 M and quantitation limit 2.1×10-4 M. CPE-MIPpy is selective to vita min B1. In conclusion, CPE-MIPpy as a working electrode showed better performance of thiamine measurement than that of CPE.

Glucoraphanin and other glucosinolates in heads of broccoli cultivars

USDA-ARS?s Scientific Manuscript database

Broccoli (Brassica oleracea L. var. italica) emerged as an increasingly popular vegetable of North American consumers during the second half of the 20th Century, with per captita consumption increasing nearly eight fold during this period. Likewise, production and consumption of broccoli has also i...

Development and Evaluation of Event-Specific Quantitative PCR Method for Genetically Modified Soybean MON87701.

PubMed

Tsukahara, Keita; Takabatake, Reona; Masubuchi, Tomoko; Futo, Satoshi; Minegishi, Yasutaka; Noguchi, Akio; Kondo, Kazunari; Nishimaki-Mogami, Tomoko; Kurashima, Takeyo; Mano, Junichi; Kitta, Kazumi

2016-01-01

A real-time PCR-based analytical method was developed for the event-specific quantification of a genetically modified (GM) soybean event, MON87701. First, a standard plasmid for MON87701 quantification was constructed. The conversion factor (C f ) required to calculate the amount of genetically modified organism (GMO) was experimentally determined for a real-time PCR instrument. The determined C f for the real-time PCR instrument was 1.24. For the evaluation of the developed method, a blind test was carried out in an inter-laboratory trial. The trueness and precision were evaluated as the bias and reproducibility of relative standard deviation (RSDr), respectively. The determined biases and the RSDr values were less than 30 and 13%, respectively, at all evaluated concentrations. The limit of quantitation of the method was 0.5%, and the developed method would thus be applicable for practical analyses for the detection and quantification of MON87701.

Connecting qualitative observation and quantitative measurement for enhancing quantitative literacy in plant anatomy course

NASA Astrophysics Data System (ADS)

Nuraeni, E.; Rahmat, A.

2018-05-01

Forming of cognitive schemes of plant anatomy concepts is performed by processing of qualitative and quantitative data obtained from microscopic observations. To enhancing student’s quantitative literacy, strategy of plant anatomy course was modified by adding the task to analyze quantitative data produced by quantitative measurement of plant anatomy guided by material course. Participant in this study was 24 biology students and 35 biology education students. Quantitative Literacy test, complex thinking in plant anatomy test and questioner used to evaluate the course. Quantitative literacy capability data was collected by quantitative literacy test with the rubric from the Association of American Colleges and Universities, Complex thinking in plant anatomy by test according to Marzano and questioner. Quantitative literacy data are categorized according to modified Rhodes and Finley categories. The results showed that quantitative literacy of biology education students is better than biology students.

Leaves play a central role in the adaptation of nitrogen and sulfur metabolism to ammonium nutrition in oilseed rape (Brassica napus).

PubMed

Coleto, Inmaculada; de la Peña, Marlon; Rodríguez-Escalante, Jon; Bejarano, Iraide; Glauser, Gaëtan; Aparicio-Tejo, Pedro M; González-Moro, M Begoña; Marino, Daniel

2017-09-20

The coordination between nitrogen (N) and sulfur (S) assimilation is required to suitably provide plants with organic compounds essential for their development and growth. The N source induces the adaptation of many metabolic processes in plants; however, there is scarce information about the influence that it may exert on the functioning of S metabolism. The aim of this work was to provide an overview of N and S metabolism in oilseed rape (Brassica napus) when exposed to different N sources. To do so, plants were grown in hydroponic conditions with nitrate or ammonium as N source at two concentrations (0.5 and 1 mM). Metabolic changes mainly occurred in leaves, where ammonium caused the up-regulation of enzymes involved in the primary assimilation of N and a general increase in the concentration of N-compounds (NH 4 + , amino acids and proteins). Similarly, the activity of key enzymes of primary S assimilation and the content of S-compounds (glutathione and glucosinolates) were also higher in leaves of ammonium-fed plants. Interestingly, sulfate level was lower in leaves of ammonium-fed plants, which was accompanied by the down-regulation of SULTR1 transporters gene expression. The results highlight the impact of the N source on different steps of N and S metabolism in oilseed rape, notably inducing N and S assimilation in leaves, and put forward the potential of N source management to modulate the synthesis of compounds with biotechnological interest, such as glucosinolates.

Promising effect of Magliasa, a traditional Iranian formula, on experimental colitis on the basis of biochemical and cellular findings

PubMed Central

Rahimi, Roja; Baghaei, Amir; Baeeri, Maryam; Amin, Gholamreza; Shams-Ardekani, Mohammad Reza; Khanavi, Mahnaz; Abdollahi, Mohammad

2013-01-01

AIM: To investigate the efficacy of Magliasa, a traditional Iranian formula, on experimental colitis. METHODS: After botanical authentication of herbal ingredients, formulation of Magliasa, quantitative determination of total glucosinolates and total phenolic content, and analysis of the thin layer chromatography profile were performed. Colitis was then induced in male rats by instillation of 2,4,6-trinitrobenzenesulfonic acid (TNBS) in all groups, aside from the Sham group. The experimental groups consisted of: the Sham group that received only normal saline; the Mag-50, Mag-100 and Mag-200 groups, which received 50, 100 and 200 mg/kg per day of Magliasa, respectively; the control group, which received vehicle water orally; the infliximab group, which received infliximab (5 mg/kg per day, subcutaneously); and the Dexa group, which received dexamethasone (1 mg/kg per day, orally). After completing the treatment period (2 wk), the rats were sacrificed, the colon was removed, its macroscopic and microscopic changes were recorded, and tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), total antioxidant capacity, myeloperoxidase (MPO), and lipid peroxidation (LPO) were assessed in colon homogenate. RESULTS: The mean value of total glucosinolates in one gram of Magliasa was 19 ± 1 μmol. The mean value of the total phenolic content was 293.8 ± 17.6 mg gallic acid equivalents per 100 gram of Magliasa. Macroscopic scores were significantly decreased in Mag-100 (1.80 ± 0.58, P = 0.019) and Mag-200 (1.20 ± 0.20, P = 0.001) compared to the control group (3.40 ± 0.24), although some inflammation and hyperemia were evident. Treatment of rats by dexamethasone (0.33 ± 0.21, P < 0.001) and infliximab (0.83 ± 0.31, P < 0.001) remarkably attenuated scores where mild hyperemia was observed macroscopically. In comparison to the control group (4.00 ± 0.32), only Mag-200 (1.60 ± 0.40) showed a significant decrease in colonic histopathological scores (P = 0.005). Minimal mucosal inflammation was observed in the Dexa group (0.67 ± 0.21, P < 0.001). The levels of TNF-α, IL-1β and MPO were significantly lower in all groups compared to the controls (P < 0.05). A significant decrease in LPO was seen in the Mag-200 (3.27 ± 0.77, P = 0.01) and Dexa (3.44 ± 0.22, P = 0.011) groups in comparison to the control group (6.43 ± 0.61). Only dexamethasone caused a significant increase in antioxidant power in comparison to the control group (346.73 ± 9.9 vs 228.33 ± 2.75, P < 0.001). Infliximab and different doses of Magliasa did not show any remarkable increase in antioxidant capacity (P > 0.05). The effect of Magliasa in all of mentioned parameters, except antioxidant capacity, was dose dependent. CONCLUSION: The effects of Magliasa in TNBS-induced colitis are encouraging and warrant clinical trials for further confirmation. PMID:23569335

First application of a microsphere-based immunoassay to the detection of genetically modified organisms (GMOs): quantification of Cry1Ab protein in genetically modified maize.

PubMed

Fantozzi, Anna; Ermolli, Monica; Marini, Massimiliano; Scotti, Domenico; Balla, Branko; Querci, Maddalena; Langrell, Stephen R H; Van den Eede, Guy

2007-02-21

An innovative covalent microsphere immunoassay, based on the usage of fluorescent beads coupled to a specific antibody, was developed for the quantification of the endotoxin Cry1Ab present in MON810 and Bt11 genetically modified (GM) maize lines. In particular, a specific protocol was developed to assess the presence of Cry1Ab in a very broad range of GM maize concentrations, from 0.1 to 100% [weight of genetically modified organism (GMO)/weight]. Test linearity was achieved in the range of values from 0.1 to 3%, whereas fluorescence signal increased following a nonlinear model, reaching a plateau at 25%. The limits of detection and quantification were equal to 0.018 and 0.054%, respectively. The present study describes the first application of quantitative high-throughput immunoassays in GMO analysis.

Utilizing pretreatment and fungal incubation to enhance the nutritional value of canola meal

USDA-ARS?s Scientific Manuscript database

The objective of this study was to determine the optimal pretreatment and fungal strain to reduce glucosinolates (GLS), fiber, and residual sugars while increasing the nutritional value of canola meal. Submerged incubation conditions were used to evaluate four pretreatment methods (extrusion, hot wa...

Activity of meadowfoam (Limnanthes alba) seed meal glucolimnanthin degradation products against soilborne pathogens

USDA-ARS?s Scientific Manuscript database

Meadowfoam (Limnanthes alba L.) is a herbaceous winter-spring annual grown as a commercial oilseed crop. The meal remaining after oil extraction from the seed contains up to 4% of the glucosinolate glucolimnanthin. Degradation of glucolimnanthin yields toxic breakdown products, and therefore the mea...

Brassica vegetables as a green manure to control Escherichia coli O157:H12 in soil

USDA-ARS?s Scientific Manuscript database

Plant remnants tilled over in soil after harvest of Brassica crops may possess antimicrobial from exudates secreted in soil following residual incorporation. We investigated the role of broccoli remnants tilled over after harvest for reducing enteric pathogens in soil. The glucosinolate-hydrolyzed c...

Metabolomic assessment reveals a stimulatory effect of calcium treatment on glucosinolates contents in broccoli microgreen

USDA-ARS?s Scientific Manuscript database

Preharvest calcium application has been shown to increase broccoli microgreen yield and extend shelf life. Here we investigated the effect of calcium application on its metabolome using ultra high-performance liquid chromatography (UHPLC) tandem with mass spectrometry (HRMS). The data collected were...

Brassica vegetables as a green manure to control Escherichia coli O157:H12 in soil

USDA-ARS?s Scientific Manuscript database

Plant remnants tilled over in soil after harvest of Brassica crops may possess antimicrobial from exudates secreted in soil following residual incorporation (till over). We investigated the role of broccoli remnants tilled over after harvest for reducing enteric pathogens in soil. The glucosinolate-...

Determination of Mycotoxin Production of Fusarium Species in Genetically Modified Maize Varieties by Quantitative Flow Immunocytometry.

PubMed

Bánáti, Hajnalka; Darvas, Béla; Fehér-Tóth, Szilvia; Czéh, Árpád; Székács, András

2017-02-22

Levels of mycotoxins produced by Fusarium species in genetically modified (GM) and near-isogenic maize, were determined using multi-analyte, microbead-based flow immunocytometry with fluorescence detection, for the parallel quantitative determination of fumonisin B1, deoxynivalenol, zearalenone, T-2, ochratoxin A, and aflatoxin B1. Maize varieties included the genetic events MON 810 and DAS-59122-7 , and their isogenic counterparts. Cobs were artificially infested by F. verticillioides and F. proliferatum conidia, and contained F. graminearum and F. sporotrichoides natural infestation. The production of fumonisin B1 and deoxynivalenol was substantially affected in GM maize lines: F. verticillioides , with the addition of F. graminearum and F. sporotrichoides , produced significantly lower levels of fumonisin B1 (~300 mg·kg -1 ) in DAS-59122-7 than in its isogenic line (~580 mg·kg -1 ), while F. proliferatum , in addition to F. graminearum and F. sporotrichoides , produced significantly higher levels of deoxynivalenol (~18 mg·kg -1 ) in MON 810 than in its isogenic line (~5 mg·kg -1 ). Fusarium verticillioides , with F. graminearum and F. sporotrichoides , produced lower amounts of deoxynivalenol and zearalenone than F. proliferatum , with F. graminearum and F. sporotrichoides . T-2 toxin production remained unchanged when considering the maize variety. The results demonstrate the utility of the Fungi-Plex™ quantitative flow immunocytometry method, applied for the high throughput parallel determination of the target mycotoxins.

Taste and physiological responses to glucosinolates: seed predator versus seed disperser.

PubMed

Samuni-Blank, Michal; Izhaki, Ido; Gerchman, Yoram; Dearing, M Denise; Karasov, William H; Trabelcy, Beny; Edwards, Thea M; Arad, Zeev

2014-01-01

In contrast to most other plant tissues, fleshy fruits are meant to be eaten in order to facilitate seed dispersal. Although fleshy fruits attract consumers, they may also contain toxic secondary metabolites. However, studies that link the effect of fruit toxins with seed dispersal and predation are scarce. Glucosinolates (GLSs) are a family of bitter-tasting compounds. The fleshy fruit pulp of Ochradenus baccatus was previously found to harbor high concentrations of GLSs, whereas the myrosinase enzyme, which breaks down GLSs to produce foul tasting chemicals, was found only in the seeds. Here we show the differential behavioral and physiological responses of three rodent species to high dose (80%) Ochradenus' fruits diets. Acomys russatus, a predator of Ochradenus' seeds, was the least sensitive to the taste of the fruit and the only rodent to exhibit taste-related physiological adaptations to deal with the fruits' toxins. In contrast, Acomys cahirinus, an Ochradenus seed disperser, was more sensitive to a diet containing the hydrolyzed products of the GLSs. A third rodent (Mus musculus) was deterred from Ochradenus fruits consumption by the GLSs and their hydrolyzed products. We were able to alter M. musculus avoidance of whole fruit consumption by soaking Ochradenus fruits in a water solution containing 1% adenosine monophosphate, which blocks the bitter taste receptor in mice. The observed differential responses of these three rodent species may be due to evolutionary pressures that have enhanced or reduced their sensitivity to the taste of GLSs.

Chemical composition, traditional and professional use in medicine, application in environmental protection, position in food and cosmetics industries, and biotechnological studies of Nasturtium officinale (watercress) - a review.

PubMed

Klimek-Szczykutowicz, Marta; Szopa, Agnieszka; Ekiert, Halina

2018-05-28

The herb of Nasturtium officinale is a raw material that has long been used in the traditional medicine of Iran, Azerbaijan, Morocco and Mauritius. Nowadays, this raw material is the object of numerous professional pharmacological studies that have demonstrated its antioxidant, anticancer, antibacterial, anti-inflammatory and cardioprotective properties. These therapeutic effects are caused by glucosinolates present in the plant, isothiocyanates, polyphenols (flavonoids, phenolic acids, proanthocyanidins), terpenes (including carotenoids), vitamins (B1, B2, B3, B6, E, C) and bioelements. The article presents the current state of phytochemical research on the generative and vegetative organs of aboveground parts. A special spotlight is put on the main N. officinale secondary metabolites - glucosinolates. Attention is drawn to the important position of N. officinale in the production of healthy foods and in the production of cosmetics. A large part of the article is devoted to the importance of this species in phytoremediation processes used in the protection of soil environments and water reservoirs. The biotechnological research on this species has also been reviewed. Those studies are of particular importance not only due to the attractiveness of this species in phytotherapy and cosmetology, but also due to the deteriorating natural state of this species and the threat of extinction. The aim of this review is to promote N. officinale as a very valuable species, not yet fully discovered by global medicine. Copyright © 2017. Published by Elsevier B.V.

PCR-free quantitative detection of genetically modified organism from raw materials – A novel electrochemiluminescence-based bio-barcode method

PubMed Central

Zhu, Debin; Tang, Yabing; Xing, Da; Chen, Wei R.

2018-01-01

Bio-barcode assay based on oligonucleotide-modified gold nanoparticles (Au-NPs) provides a PCR-free method for quantitative detection of nucleic acid targets. However, the current bio-barcode assay requires lengthy experimental procedures including the preparation and release of barcode DNA probes from the target-nanoparticle complex, and immobilization and hybridization of the probes for quantification. Herein, we report a novel PCR-free electrochemiluminescence (ECL)-based bio-barcode assay for the quantitative detection of genetically modified organism (GMO) from raw materials. It consists of tris-(2’2’-bipyridyl) ruthenium (TBR)-labele barcode DNA, nucleic acid hybridization using Au-NPs and biotin-labeled probes, and selective capture of the hybridization complex by streptavidin-coated paramagnetic beads. The detection of target DNA is realized by direct measurement of ECL emission of TBR. It can quantitatively detect target nucleic acids with high speed and sensitivity. This method can be used to quantitatively detect GMO fragments from real GMO products. PMID:18386909

PCR-free quantitative detection of genetically modified organism from raw materials. An electrochemiluminescence-based bio bar code method.

PubMed

Zhu, Debin; Tang, Yabing; Xing, Da; Chen, Wei R

2008-05-15

A bio bar code assay based on oligonucleotide-modified gold nanoparticles (Au-NPs) provides a PCR-free method for quantitative detection of nucleic acid targets. However, the current bio bar code assay requires lengthy experimental procedures including the preparation and release of bar code DNA probes from the target-nanoparticle complex and immobilization and hybridization of the probes for quantification. Herein, we report a novel PCR-free electrochemiluminescence (ECL)-based bio bar code assay for the quantitative detection of genetically modified organism (GMO) from raw materials. It consists of tris-(2,2'-bipyridyl) ruthenium (TBR)-labeled bar code DNA, nucleic acid hybridization using Au-NPs and biotin-labeled probes, and selective capture of the hybridization complex by streptavidin-coated paramagnetic beads. The detection of target DNA is realized by direct measurement of ECL emission of TBR. It can quantitatively detect target nucleic acids with high speed and sensitivity. This method can be used to quantitatively detect GMO fragments from real GMO products.

Vegetative anatomy and relationships of Setchellanthus caeruleus (Setchellanthaceae)

Treesearch

Sherwin Carlquist; Regis B. Miller

1999-05-01

On account of its distinctive features, Setchellanthus cannot be included within any of the families of glucosinolate-producing plants. Features unknown in any of these families include abundant vasicentric tracheids, abaxial axial parenchyma, and only short uniseriate rays (composed of upright cells) in wood: and the presence of wide-helix tracheary elements in leaves...

Conversion of canola meal into a high-protein feed additive via solid-state fungal incubation process

USDA-ARS?s Scientific Manuscript database

The study goal was to determine the optimal fungal culture to reduce glucosinolates (GLS), fiber, and residual sugars while increasing the protein content and nutritional value of canola meal. Solid-state incubation conditions were used to enhance filamentous growth of the fungi. Flask trials were p...

Could nitrile derivatives of turnip (Brassica rapa) glucosinolates be Hepato-and/or cholangiotoxic in cattle?

USDA-ARS?s Scientific Manuscript database

Turnip (Brassica rapa ssp. rapa) and rape (Brassica napus ssp. biennis) and other brassica forage crops are generally regarded as “safe” feed for cattle during late summer and fall in New Zealand. However, when Pithomyces chartarum spore counts are high there are epidemics of sporidesmin toxicity (...

7 CFR 810.301 - Definition of canola.

Code of Federal Regulations, 2010 CFR

2010-01-01

... glucosinolate, per gram of air-dried, oil free solid. Before the removal of dockage, the seed shall contain not... GRAIN United States Standards for Canola-Terms Defined § 810.301 Definition of canola. Seeds of the genus Brassica from which the oil shall contain less than 2 percent erucic acid in its fatty acid...

7 CFR 810.301 - Definition of canola.

Code of Federal Regulations, 2011 CFR

2011-01-01

... glucosinolate, per gram of air-dried, oil free solid. Before the removal of dockage, the seed shall contain not... GRAIN United States Standards for Canola-Terms Defined § 810.301 Definition of canola. Seeds of the genus Brassica from which the oil shall contain less than 2 percent erucic acid in its fatty acid...

Potential of forage brassicas for use in pasture-based livestock systems

USDA-ARS?s Scientific Manuscript database

Brassicas (Brassica spp.) are gaining popularity as high-quality forage for pasture-based livestock producers due to their use to extend the fall grazing season and during the summer forage slump. However, inclusion of brassicas in the diet can be limited by the presence of glucosinolates (a class o...

Extraction and Purification of Glucoraphanin by Preparative High-Performance Liquid Chromatography (HPLC)

ERIC Educational Resources Information Center

Lee, Iris; Boyce, Mary C.

2011-01-01

A student activity that focuses on the isolation of glucoraphanin from broccoli using preparative high-performance liquid chromatography (HPLC) is presented here. Glucoraphanin is a glucosinolate, whose byproducts are known to possess anticancer properties. It is present naturally at high levels in broccoli and other "Brassica" vegetables. This…

Detection of susceptibility genes as modifiers due to subgroup differences in complex disease.

PubMed

Bergen, Sarah E; Maher, Brion S; Fanous, Ayman H; Kendler, Kenneth S

2010-08-01

Complex diseases invariably involve multiple genes and often exhibit variable symptom profiles. The extent to which disease symptoms, course, and severity differ between affected individuals may result from underlying genetic heterogeneity. Genes with modifier effects may or may not also influence disease susceptibility. In this study, we have simulated data in which a subset of cases differ by some effect size (ES) on a quantitative trait and are also enriched for a risk allele. Power to detect this 'pseudo-modifier' gene in case-only and case-control designs was explored blind to case substructure. Simulations involved 1000 iterations and calculations for 80% power at P<0.01 while varying the risk allele frequency (RAF), sample size (SS), ES, odds ratio (OR), and proportions of the case subgroups. With realistic values for the RAF (0.20), SS (3000) and ES (1), an OR of 1.7 is necessary to detect a pseudo-modifier gene. Unequal numbers of subjects in the case groups result in little decrement in power until the group enriched for the risk allele is <30% or >70% of the total case population. In practice, greater numbers of subjects and selection of a quantitative trait with a large range will provide researchers with greater power to detect a pseudo-modifier gene. However, even under ideal conditions, studies involving alleles with low frequencies or low ORs are usually underpowered for detection of a modifier or susceptibility gene. This may explain some of the inconsistent association results for many candidate gene studies of complex diseases.

Investigation of glucosinolate profile and qualitative aspects in sprouts and roots of horseradish (Armoracia rusticana) using LC-ESI-hybrid linear ion trap with Fourier transform ion cyclotron resonance mass spectrometry and infrared multiphoton dissociation.

PubMed

Agneta, Rosa; Rivelli, Anna Rita; Ventrella, Emanuela; Lelario, Filomena; Sarli, Giulio; Bufo, Sabino Aurelio

2012-08-01

Within the family of Brassicaceae, an important source of glucosinolates (GLSs) is represented by horseradish ( Armoracia rusticana P. Gaertner, B. Meyer & Scherbius), cultivated for its roots, which are grated fresh or processed into a sauce and used as a condiment. The characteristic pungent flavor of the root depends on the abundance of the bioactive GLS molecules. In crude plant extracts (sprouts and roots) of an accession of horseradish largely diffused in the Basilicata region (southern Italy), which develops many sprouts and produces white, fiery, and sharp-flavored marketable roots, we characterized the GLS profile by LC-ESI-LTQ-FTICR-MS and IRMPD. In sprouts and roots we identified 16 and 11 GLSs, respectively. We confirmed the presence of sinigrin, 4-hydroxyglucobrassicin, glucobrassicin, gluconasturtin, and 4-methoxyglucobrassicin and identified glucoiberin, gluconapin, glucocochlearin, glucoconringianin, glucosativin, glucoibarin, 5-hydroxyglucobrassicin, glucocapparilinearisin or glucobrassicanapin, glucotropaeolin, and glucoarabishirsutain, not previously characterized in horseradish. Of particular note was the presence of the putative 2-methylsulfonyl-oxo-ethyl-GLS.

Spasmolytic, antimicrobial and cytotoxic activities of 5-phenylpentyl isothiocyanate, a new glucosinolate autolysis product from horseradish (Armoracia rusticana P. Gaertn., B. Mey. & Scherb., Brassicaceae).

PubMed

Dekić, Milan S; Radulović, Niko S; Stojanović, Nikola M; Randjelović, Pavle J; Stojanović-Radić, Zorica Z; Najman, Stevo; Stojanović, Sanja

2017-10-01

Detailed analyses of horseradish autolysates led to the identification of a new natural product, 5-phenylpentyl isothiocyanate (PhPeITC). The structural assignment was corroborated by synthesis, and the identity unequivocally established by spectral means. The occurrence of PhPeITC is the first direct proof of the existence of a 5-phenylpentyl glucosinolate in the aerial parts of this species as one of the possible "mustard oil" precursors. To verify its possible contribution to the horseradish functional food status, horseradish above- and underground autolysates, together with five ω-phenylalkyl isothiocyanates were tested for their spasmolytic, cytotoxic and antimicrobial activities. Specifically, the cytotoxic effect on Caco-2, HeLa (cancer) and MDCK (non-cancer) cell lines was established. Additionally, the five tested ITCs exerted significant spasmolytic activity (on rat distal colon), with PhPeITC being almost 100 times more potent than papaverine. A non-selective antimicrobial activity of all ITCs was revealed in the case of 6 bacterial and 2 fungal strains. Copyright © 2017 Elsevier Ltd. All rights reserved.

Structural elucidation of 4-(cystein-S-yl)butyl glucosinolate from the leaves of Eruca sativa.

PubMed

Kim, Sun-Ju; Kawaharada, Chiami; Jin, Shigeki; Hashimoto, Makoto; Ishii, Gensho; Yamauchi, Hiroaki

2007-01-01

The structurally unique glucosinolate (GSL), 4-(cystein-S-yl)butyl GSL, was identified in the leaves of hydroponically-grown rocket salad (Eruca sativa Mill.). Its electrospray ionization mass spectrometry (ESI-MS)/MS spectrum indicated that this unusual GSL had a molecular weight of 414 as a desulfo (DS)-GSL, and a molecular formula of C(14)H(25)N(2)O(8)S(2) based on its negative ion matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) spectrum. For further confirmation, the 4-(cystein-S-yl)butyl DS-GSL was prepared with authentic L-Ser and purified dimeric 4-mercaptobutyl DS-GSL, and its chemical structure then confirmed by ESI-MS/MS data. It is named "glucorucolamine" as a trivial name from its ammonia sensitivity. This unique GSL was found to the greatest extent when rocket salad was grown in a 100% NH4+-N nutrient solution. Despite it clearly seems to reduce the detoxification of excess NH4+ in the leaves of rocket salad, present knowledge about the unique GSL is still far from being sufficient.

Effect of water content and temperature on inactivation kinetics of myrosinase in broccoli (Brassica oleracea var. italica).

PubMed

Oliviero, T; Verkerk, R; Van Boekel, M A J S; Dekker, M

2014-11-15

Broccoli belongs to the Brassicaceae plant family consisting of widely eaten vegetables containing high concentrations of glucosinolates. Enzymatic hydrolysis of glucosinolates by endogenous myrosinase (MYR) can form isothiocyanates with health-promoting activities. The effect of water content (WC) and temperature on MYR inactivation in broccoli was investigated. Broccoli was freeze dried obtaining batches with WC between 10% and 90% (aw from 0.10 to 0.96). These samples were incubated for various times at different temperatures (40-70°C) and MYR activity was measured. The initial MYR inactivation rates were estimated by the first-order reaction kinetic model. MYR inactivation rate constants were lower in the driest samples (10% WC) at all studied temperatures. Samples with 67% and 90% WC showed initial inactivation rate constants all in the same order of magnitude. Samples with 31% WC showed intermediate initial inactivation rate constants. These results are useful to optimise the conditions of drying processes to produce dried broccoli with optimal MYR retention for human health. Copyright © 2014 Elsevier Ltd. All rights reserved.

Effect of different cooking methods on color, phytochemical concentration, and antioxidant capacity of raw and frozen brassica vegetables.

PubMed

Pellegrini, Nicoletta; Chiavaro, Emma; Gardana, Claudio; Mazzeo, Teresa; Contino, Daniele; Gallo, Monica; Riso, Patrizia; Fogliano, Vincenzo; Porrini, Marisa

2010-04-14

This study evaluated the effect of common cooking practices (i.e., boiling, microwaving, and basket and oven steaming) on the phytochemical content (carotenoids, chlorophylls, glucosinolates, polyphenols, and ascorbic acid), total antioxidant capacity (TAC), and color changes of three generally consumed Brassica vegetables analyzed fresh and frozen. Among cooking procedures, boiling determined an increase of fresh broccoli carotenoids and fresh Brussels sprout polyphenols, whereas a decrease of almost all other phytochemicals in fresh and frozen samples was observed. Steaming procedures determined a release of polyphenols in both fresh and frozen samples. Microwaving was the best cooking method for maintaining the color of both fresh and frozen vegetables and obtaining a good retention of glucosinolates. During all cooking procedures, ascorbic acid was lost in great amount from all vegetables. Chlorophylls were more stable in frozen samples than in fresh ones, even though steaming methods were able to better preserve these compounds in fresh samples than others cooking methods applied. The overall results of this study demonstrate that fresh Brassica vegetables retain phytochemicals and TAC better than frozen samples.

Identification and quantification of three genetically modified insect resistant cotton lines using conventional and TaqMan real-time polymerase chain reaction methods.

PubMed

Yang, Litao; Pan, Aihu; Zhang, Kewei; Guo, Jinchao; Yin, Changsong; Chen, Jianxiu; Huang, Cheng; Zhang, Dabing

2005-08-10

As the genetically modified organisms (GMOs) labeling policies are issued in many countries, qualitative and quantitative polymerase chain reaction (PCR) techniques are increasingly used for the detection of genetically modified (GM) crops in foods. Qualitative PCR and TaqMan real-time quantitative PCR methods to detect and identify three varieties of insect resistant cotton, i.e., Mon531 cotton (Monsanto Co.) and GK19 and SGK321 cottons (Chinese Academy of Agricultural Sciences), which were approved for commercialization in China, were developed in this paper. Primer pairs specific to inserted DNAs, such as Cowpea trypsin inhibitor (CpTI) gene of SGK321 cotton and the specific junction DNA sequences containing partial Cry1A(c) gene and NOS terminator of Mon531, GK19, and SGK321 cotton varieties were designed to conduct the identified PCR assays. In conventional specific identified PCR assays, the limit of detection (LOD) was 0.05% for Mon531, GK19, or SGK321 in 100 ng of cotton genomic DNA for one reaction. Also, the multiplex PCR method for screening the three GM cottons was also established, which could save time and cost in practical detection. Furthermore, a real-time quantitative PCR assay based on TaqMan chemistry for detection of insect resistant gene, Cry1A(c), was developed. This assay also featured the use of a standard plasmid as a reference molecule, which contained both a specific region of the transgene Cry1A(c) and an endogenous stearoyl-acyl carrier protein desaturase (Sad1) gene of the cotton. In quantitative PCR assay, the quantification range was from 0.01 to 100% in 100 ng of the genome DNA template, and in the detection of 1.0, 3.0, and 5.0% levels of three insect resistant cotton lines, respectively, all of the relative standard deviations (RSDs) were less than 8.2% except for the GM cotton samples with 1.0% Mon531 or GK19, which meant that our real-time PCR assays involving the use of reference molecule were reliable and practical for GM insect resistant cottons quantification. All of these results indicated that our established conventional and TaqMan real-time PCR assays were applicable to detect the three insect resistant cottons qualitatively and quantitatively.

Event specific qualitative and quantitative polymerase chain reaction detection of genetically modified MON863 maize based on the 5'-transgene integration sequence.

PubMed

Yang, Litao; Xu, Songci; Pan, Aihu; Yin, Changsong; Zhang, Kewei; Wang, Zhenying; Zhou, Zhigang; Zhang, Dabing

2005-11-30

Because of the genetically modified organisms (GMOs) labeling policies issued in many countries and areas, polymerase chain reaction (PCR) methods were developed for the execution of GMO labeling policies, such as screening, gene specific, construct specific, and event specific PCR detection methods, which have become a mainstay of GMOs detection. The event specific PCR detection method is the primary trend in GMOs detection because of its high specificity based on the flanking sequence of the exogenous integrant. This genetically modified maize, MON863, contains a Cry3Bb1 coding sequence that produces a protein with enhanced insecticidal activity against the coleopteran pest, corn rootworm. In this study, the 5'-integration junction sequence between the host plant DNA and the integrated gene construct of the genetically modified maize MON863 was revealed by means of thermal asymmetric interlaced-PCR, and the specific PCR primers and TaqMan probe were designed based upon the revealed 5'-integration junction sequence; the conventional qualitative PCR and quantitative TaqMan real-time PCR detection methods employing these primers and probes were successfully developed. In conventional qualitative PCR assay, the limit of detection (LOD) was 0.1% for MON863 in 100 ng of maize genomic DNA for one reaction. In the quantitative TaqMan real-time PCR assay, the LOD and the limit of quantification were eight and 80 haploid genome copies, respectively. In addition, three mixed maize samples with known MON863 contents were detected using the established real-time PCR systems, and the ideal results indicated that the established event specific real-time PCR detection systems were reliable, sensitive, and accurate.

Modified ion source triple quadrupole mass spectrometer gas chromatograph for polycyclic aromatic hydrocarbon analyses

PubMed Central

Anderson, Kim A.; Szelewski, Michael J.; Wilson, Glenn; Quimby, Bruce D.; Hoffman, Peter D.

2015-01-01

We describe modified gas chromatography electron-impact/triple-quadrupole mass spectrometry (GC–EI/MS/MS) utilizing a newly developed hydrogen-injected self-cleaning ion source and modified 9 mm extractor lens. This instrument, with optimized parameters, achieves quantitative separation of 62 polycyclic aromatic hydrocarbons (PAHs). Existing methods historically limited rigorous identification and quantification to a small subset, such as the 16 PAHs the US EPA has defined as priority pollutants. Without the critical source and extractor lens modifications, the off-the-shelf GC–EI/MS/MS system was unsuitable for complex PAH analysis. Separations were enhanced by increased gas flow, a complex GC temperature profile incorporating multiple isothermal periods, specific ramp rates, and a PAH-optimized column. Typical determinations with our refined GC–EI/MS/MS have a large linear range of 1–10,000 pg μl−1 and detection limits of <2 pg μl−1. Included in the 62 PAHs, multiple-reaction-monitoring (MRM) mode enabled GC-EI/MS/MS identification and quantitation of several constituents of the MW 302 PAHs isomers. Using calibration standards, values determined were within 5% of true values over many months. Standard curve r2 values were typically >0.998, exceptional for compounds which are archetypally difficult. With this method benzo[a]fluorene, benzo[b]fluorene, benzo[c]fluorene were fully separated as was benzo[b]fluoranthene, benzo[k]fluoranthene, and benzo[j]fluoranthene. Chrysene and triphenylene, were sufficiently separated to allow accurate quantitation. Mean limits of detection (LODs) across all PAHs were 1.02 ± 0.84 pg μl−1 with indeno[1,2,3-c,d] pyrene having the lowest LOD at 0.26 pg μl−1 and only two analytes above 2.0 pg μl−1; acenaphthalene (2.33 pg μl−1) and dibenzo[a,e]pyrene (6.44 pg μl−1). PMID:26454790

Advanced glycation end product (AGE) modified proteins in tears of diabetic patients.

PubMed

Zhao, Zhenjun; Liu, Jingfang; Shi, Bingyin; He, Shuixiang; Yao, Xiaoli; Willcox, Mark D P

2010-08-11

High glucose level in diabetic patients may lead to advanced glycation end product (AGE) modified proteins. This study investigated AGE modified proteins in tears and compared their levels in diabetic patients (DM) with non-diabetic controls (CTL). Basal tears were collected from DM with (DR) or without (DNR) retinopathy and CTL. Total AGE modified proteins were detected quantitatively by a dot immunobinding assay. The AGE modified proteins were separated in 1D- and 2D-SDS gels and detected by western-blotting. The individual AGE modified proteins were also compared between groups using densitometry. Compared with the CTL group, tear concentrations of AGE modified proteins were significantly elevated in DR and DNR groups. The concentration of AGE modified proteins in diabetic tears were positively correlated with AGE modified hemoglobin (HbA1c) and postprandial blood glucose level (PBG). Western blotting of AGE modified proteins from 1D-SDS gels showed several bands, the major one at around 60 kDa. The intensities of AGE modified protein bands were higher in DM tears than in CTL tears. Western blotting from 2D-SDS gels showed a strongly stained horizontal strip, which corresponded to the major band in 1D-SDS gels. Most of the other AGE modified protein species were within molecular weight of 30-60 kDa, PI 5.2-7.0. Densitometry analysis demonstrated several AGE modified proteins were elevated in DR or DNR tears. Total and some individual AGE modified proteins were elevated in DM tears. AGE modified proteins in tears may be used as biomarkers to diagnose diabetes and/or diabetic retinopathy.

Exogenous Methyl Jasmonate Treatment Increases Glucosinolate Biosynthesis and Quinone Reductase Activity in Kale Leaf Tissue

PubMed Central

Ku, Kang-Mo; Jeffery, Elizabeth H.; Juvik, John A.

2014-01-01

Methyl jasmonate (MeJA) spray treatments were applied to the kale varieties ‘Dwarf Blue Curled Vates’ and ‘Red Winter’ in replicated field plantings in 2010 and 2011 to investigate alteration of glucosinolate (GS) composition in harvested leaf tissue. Aqueous solutions of 250 µM MeJA were sprayed to saturation on aerial plant tissues four days prior to harvest at commercial maturity. The MeJA treatment significantly increased gluconasturtiin (56%), glucobrassicin (98%), and neoglucobrassicin (150%) concentrations in the apical leaf tissue of these genotypes over two seasons. Induction of quinone reductase (QR) activity, a biomarker for anti-carcinogenesis, was significantly increased by the extracts from the leaf tissue of these two cultivars. Extracts of apical leaf tissues had greater MeJA mediated increases in phenolics, glucosinolate concentrations, GS hydrolysis products, and QR activity than extracts from basal leaf tissue samples. The concentration of the hydrolysis product of glucoraphanin, sulforphane was significantly increased in apical leaf tissue of the cultivar ‘Red Winter’ in both 2010 and 2011. There was interaction between exogenous MeJA treatment and environmental conditions to induce endogenous JA. Correlation analysis revealed that indole-3-carbanol (I3C) generated from the hydrolysis of glucobrassicin significantly correlated with QR activity (r = 0.800, P<0.001). Concentrations required to double the specific QR activity (CD values) of I3C was calculated at 230 µM, which is considerably weaker at induction than other isothiocyanates like sulforphane. To confirm relationships between GS hydrolysis products and QR activity, a range of concentrations of MeJA sprays were applied to kale leaf tissues of both cultivars in 2011. Correlation analysis of these results indicated that sulforaphane, NI3C, neoascorbigen, I3C, and diindolylmethane were all significantly correlated with QR activity. Thus, increased QR activity may be due to combined increases in phenolics (quercetin and kaempferol) and GS hydrolysis product concentrations rather than by individual products alone. PMID:25084454

Consumer acceptability and sensory profile of cooked broccoli with mustard seeds added to improve chemoprotective properties.

PubMed

Ghawi, Sameer Khalil; Shen, Yuchi; Niranjan, Keshavan; Methven, Lisa

2014-09-01

Broccoli, a rich source of glucosinolates, is a commonly consumed vegetable of the Brassica family. Hydrolysis products of glucosinolates, isothiocyanates, have been associated with health benefits and contribute to the flavor of Brassica. However, boiling broccoli causes the myrosinase enzyme needed for hydrolysis to denature. In order to ensure hydrolysis, broccoli must either be mildly cooked or active sources of myrosinase, such as mustard seed powder, can be added postcooking. In this study, samples of broccoli were prepared in 6 different ways; standard boiling, standard boiling followed by the addition of mustard seeds, sous vide cooking at low temperature (70 °C) and sous vide cooking at higher temperature (100 °C) and sous vide cooking at higher temperature followed by the addition of mustard seeds at 2 different concentrations. The majority of consumers disliked the mildly cooked broccoli samples (70 °C, 12 min, sous vide) which had a hard and stringy texture. The highest mean consumer liking was for standard boiled samples (100 °C, 7 min). Addition of 1% mustard seed powder developed sensory attributes, such as pungency, burning sensation, mustard odor, and flavor. One cluster of consumers (32%) found mustard seeds to be a good complement to cooked broccoli; however, the majority disliked the mustard-derived sensory attributes. Where the mustard seeds were partially processed, doubling the addition to 2% led to only the same level of mustard and pungent flavors as 1% unprocessed seeds, and mean consumer liking remained unaltered. This suggests that optimization of the addition level of partially processed mustard seeds may be a route to enhance bioactivity of cooked broccoli without compromising consumer acceptability. © 2014 Institute of Food Technologists®

Detection methods for biotech cotton MON 15985 and MON 88913 by PCR.

PubMed

Lee, Seong-Hun; Kim, Jin-Kug; Yi, Bu-Young

2007-05-02

Plants derived through agricultural biotechnology, or genetically modified organisms (GMOs), may affect human health and ecological environment. A living GMO is also called a living modified organism (LMO). Biotech cotton is a GMO in food or feed and also an LMO in the environment. Recently, two varieties of biotech cotton, MON 15985 and MON 88913, were developed by Monsanto Co. The detection method is an essential element for the GMO labeling system or LMO management of biotech plants. In this paper, two primer pairs and probes were designed for specific amplification of 116 and 120 bp PCR products from MON 15985 and MON 88913, respectively, with no amplification from any other biotech cotton. Limits of detection of the qualitative method were all 0.05% for MON 15985 and MON 88913. The quantitative method was developed using a TaqMan real-time PCR. A synthetic plasmid, as a reference molecule, was constructed from a taxon-specific DNA sequence of cotton and two construct-specific DNA sequences of MON 15985 and MON 88913. The quantitative method was validated using six samples that contained levels of biotech cotton mixed with conventional cotton ranging from 0.1 to 10.0%. As a result, the biases from the true value and the relative deviations were all within the range of +/-20%. Limits of quantitation of the quantitative method were all 0.1%. Consequently, it is reported that the proposed detection methods were applicable for qualitative and quantitative analyses for biotech cotton MON 15985 and MON 88913.

Surface-enhanced Raman spectroscopy detection of polybrominated diphenylethers using a portable Raman spectrometer.

PubMed

Jiang, Xiaohong; Lai, Yongchao; Wang, Wei; Jiang, Wei; Zhan, Jinhua

2013-11-15

Polybrominated diphenylethers (PBDEs), one of the most common brominated flame retardants, are toxic and persistent, generally detected by the chromatographic method. In this work, qualitative and quantitative detection of PBDEs were explored based on surface-enhanced Raman spectroscopy (SERS) technique using a portable Raman spectrometer. Alkanethiol modified silver nanoparticle aggregates were used as the substrate and PBDEs could be pre-concentrated close to the substrate surface through their hydrophobic interactions with alkanethiol. The effect of alkanethiols with different chain length on the SERS detection of PBDEs was evaluated. It was shown that 1-hexanethiol (HT) modified substrate has higher sensitivity, good stability and reusability. Qualitative and quantitative SERS detection of PBDEs in real sea water was accomplished, with the measured detection limits at 1.2×10(2) μg L(-1). These results illustrate SERS could be used as an effective method for the detection of PBDEs. Copyright © 2013 Elsevier B.V. All rights reserved.

Quantitative determination of Escherichia coli based on the electrochemical measurement of bacterial catalase activity using H2O2-selective organic/inorganic-hybrid sol-gel film-modified Pt electrode.

PubMed

Hasebe, Yasushi; Fukuzawa, Michiru; Matsuhisa, Hironori

2009-01-01

Quantitative determination of Escherichia coli (E. coli) concentration was achieved by measuring the intrinsic catalase activity of E. coli using novel H2O2-selective organic/inorganic-hybrid sol-gel film-modified platinum (Pt) wire electrode. This hybrid sol-gel film is composed of three kinds of organosilanes and two biopolymers (i.e., chitosan and bovine serum albumin), and exhibited an excellent permselectivity toward H2O2 based on a size-exclusive mechanism. The steady-state anodic current for 100 [xmol/L H2O2 at +0.6 V (vs. Ag/AgCl) in 0.1 mol/L phosphate buffer (pH 6.5) solution was apparently diminished by the addition of E. coli samples, due to the decomposition of H2O2 by intrinsic catalase activity of E. coli. The time-dependent decrease in current (-AI/At) was significantly dependent on the E. coli concentration. The -AI/At was enhanced by the permeabilization pretreatment of E. coli samples with the mixed solution of polymyxin B and lysozyme. This H2O2-selective organic/inorganic-hybrid sol-gel film-modified platinum (Pt) wire electrode allowed quantitative determination of E. coli concentration ranging from 10(6) to 10(9) CFU/mL within 30 min. This method required no label and complicated procedure, and allowed rapid, simple and cost-effective quantitative electrochemical determination of catalase-positive bacteria.

Combinatorial modification of human histone H4 quantitated by two-dimensional liquid chromatography coupled with top down mass spectrometry.

PubMed

Pesavento, James J; Bullock, Courtney R; LeDuc, Richard D; Mizzen, Craig A; Kelleher, Neil L

2008-05-30

Quantitative proteomics has focused heavily on correlating protein abundances, ratios, and dynamics by developing methods that are protein expression-centric (e.g. isotope coded affinity tag, isobaric tag for relative and absolute quantification, etc.). These methods effectively detect changes in protein abundance but fail to provide a comprehensive perspective of the diversity of proteins such as histones, which are regulated by post-translational modifications. Here, we report the characterization of modified forms of HeLa cell histone H4 with a dynamic range >10(4) using a strictly Top Down mass spectrometric approach coupled with two dimensions of liquid chromatography. This enhanced dynamic range enabled the precise characterization and quantitation of 42 forms uniquely modified by combinations of methylation and acetylation, including those with trimethylated Lys-20, monomethylated Arg-3, and the novel dimethylated Arg-3 (each <1% of all H4 forms). Quantitative analyses revealed distinct trends in acetylation site occupancy depending on Lys-20 methylation state. Because both modifications are dynamically regulated through the cell cycle, we simultaneously investigated acetylation and methylation kinetics through three cell cycle phases and used these data to statistically assess the robustness of our quantitative analysis. This work represents the most comprehensive analysis of histone H4 forms present in human cells reported to date.

Development and in-house validation of the event-specific qualitative and quantitative PCR detection methods for genetically modified cotton MON15985.

PubMed

Jiang, Lingxi; Yang, Litao; Rao, Jun; Guo, Jinchao; Wang, Shu; Liu, Jia; Lee, Seonghun; Zhang, Dabing

2010-02-01

To implement genetically modified organism (GMO) labeling regulations, an event-specific analysis method based on the junction sequence between exogenous integration and host genomic DNA has become the preferential approach for GMO identification and quantification. In this study, specific primers and TaqMan probes based on the revealed 5'-end junction sequence of GM cotton MON15985 were designed, and qualitative and quantitative polymerase chain reaction (PCR) assays were established employing the designed primers and probes. In the qualitative PCR assay, the limit of detection (LOD) was 0.5 g kg(-1) in 100 ng total cotton genomic DNA, corresponding to about 17 copies of haploid cotton genomic DNA, and the LOD and limit of quantification (LOQ) for quantitative PCR assay were 10 and 17 copies of haploid cotton genomic DNA, respectively. Furthermore, the developed quantitative PCR assays were validated in-house by five different researchers. Also, five practical samples with known GM contents were quantified using the developed PCR assay in in-house validation, and the bias between the true and quantification values ranged from 2.06% to 12.59%. This study shows that the developed qualitative and quantitative PCR methods are applicable for the identification and quantification of GM cotton MON15985 and its derivates.

Estimating the fates of organic contaminants in an aquifer using QSAR.

PubMed

Lim, Seung Joo; Fox, Peter

2013-01-01

The quantitative structure activity relationship (QSAR) model, BIOWIN, was modified to more accurately estimate the fates of organic contaminants in an aquifer. The predictions from BIOWIN were modified to include oxidation and sorption effects. The predictive model therefore included the effects of sorption, biodegradation, and oxidation. A total of 35 organic compounds were used to validate the predictive model. The majority of the ratios of predicted half-life to measured half-life were within a factor of 2 and no ratio values were greater than a factor of 5. In addition, the accuracy of estimating the persistence of organic compounds in the sub-surface was superior when modified by the relative fraction adsorbed to the solid phase, 1/Rf, to that when modified by the remaining fraction of a given compound adsorbed to a solid, 1 - fs.

Separation and simultaneous quantitation of PGF2α and its epimer 8-iso-PGF2α using modifier-assisted differential mobility spectrometry tandem mass spectrometry.

PubMed

Liang, Chunsu; Sun, Hui; Meng, Xiangjun; Yin, Lei; Fawcett, J Paul; Yu, Huaidong; Liu, Ting; Gu, Jingkai

2018-03-01

Because many therapeutic agents are contaminated by epimeric impurities or form epimers as a result of metabolism, analytical tools capable of determining epimers are increasingly in demand. This article is a proof-of-principle report of a novel DMS-MS/MS method to separate and simultaneously quantify epimers, taking PGF2 α and its 8-epimer, 8- iso -PGF2 α , as an example. Good accuracy and precision were achieved in the range of 10-500 ng/mL with a run time of only 1.5 min. Isopropanol as organic modifier facilitated a good combination of sensitivity and separation. The method is the first example of the quantitation of epimers without chromatographic separation.

Pseudomonas sax genes overcome aliphatic isothiocyanate-mediated non-host resistance in Arabidopsis

Treesearch

Jun Fan; Casey Crooks; Gary Creissen; Lionel Hill; Shirley Fairhurst; Peter Doerner; Chris Lamb

2011-01-01

Most plant-microbe interactions do not result in disease; natural products restrict non-host pathogens. We found that sulforaphane (4-methylsulfinylbutyl isothiocyanate), a natural product derived from aliphatic glucosinolates, inhibits growth in Arabidopsis of non-host Pseudomonas bacteria in planta. Multiple sax genes (saxCAB/F/D/G) were identified in Pseudomonas...

Comparison of DNA extraction kits and modification of DNA elution procedure for the quantitation of subdominant bacteria from piggery effluents with real-time PCR

PubMed Central

Desneux, Jérémy; Pourcher, Anne-Marie

2014-01-01

Four commercial DNA extraction kits and a minor modification in the DNA elution procedure were evaluated for the quantitation of bacteria in pig manure samples. The PowerSoil®, PowerFecal®, NucleoSpin® Soil kits and QIAamp® DNA Stool Mini kit were tested on raw manure samples and on lagoon effluents for their ability to quantify total bacteria and a subdominant bacteria specific of pig manure contamination: Lactobacillus amylovorus. The NucleoSpin® Soil kit (NS kit), and to a lesser extent the PowerFecal® kit were the most efficient methods. Regardless of the kit utilized, the modified elution procedure increased DNA yield in the lagoon effluent by a factor of 1.4 to 1.8. When tested on 10 piggery effluent samples, compared to the QIAamp kit, the NS kit combined with the modified elution step, increased by a factor up to 1.7 log10 the values of the concentration of L. amylovorus. Regardless of the type of manure, the best DNA quality and the highest concentrations of bacteria were obtained using the NS kit combined with the modification of the elution procedure. The method recommended here significantly improved quantitation of subdominant bacteria in manure. PMID:24838631

Determination of Acid Herbicides Using Modified QuEChERS with Fast Switching ESI(+)/ESI(-) LC-MS/MS.

PubMed

Sack, Chris; Vonderbrink, John; Smoker, Michael; Smith, Robert E

2015-11-04

A method for the determination of 35 acid herbicides in food matrices was developed, validated, and implemented. It utilizes a modified QuEChERS extraction procedure coupled with quantitation by liquid chromatography tandem mass spectrometry (LC-MS/MS). The acid herbicides analyzed are all organic carboxylic acids, including the older chlorophenoxy acid herbicides such as 2,4-dichlorophenoxyacetic acid (2,4-D), dicamba, 4-chlorophenoxyacetic acid (4-CPA), quinclorac, and many of the newer imidazolinone herbicides such as imazethapyr and imazaquin. In the procedure, 10 mL of water is added to 5 g of sample and then extracted with 1% formic acid in acetonitrile for 1 min. The acetonitrile phase is salted out of the extract by adding sodium chloride and magnesium sulfate, followed by centrifugation. The acetonitrile is diluted 1:1 with water to enable quantitation by LC-MS/MS using fast switching between positive and negative electrospray ionization modes. The average recoveries for all the compounds except aminocyclopyrachlor were 95% with a precision of 8%. The method detection limits for all residues were less than 10 ng/g, and the correlation coefficients for the calibration curves was greater than 0.99 for all but two compounds tested. The method was used successfully for the quantitation of acid herbicides in the FDA's total diet study. The procedure proved to be accurate, precise, linear, sensitive, and rugged.

Flow cytometric quantitation of phagocytosis in heparinized complete blood with latex particles and Candida albicans.

PubMed

Egido, J M; Viñuelas, J

1997-01-01

We report a rapid method for the flow cytometric quantitation of phagocytosis in heparinized complete peripheral blood (HCPB), using commercially available phycoerythrin-conjugated latex particles of 1 micron diameter. The method is faster and shows greater reproducibility than Bjerknes' (1984) standard technique using propidium iodide-stained Candida albicans, conventionally applied to the leukocytic layer of peripheral blood but here modified for HCPB. We also report a modification of Bjerknes' Intracellular Killing Test to allow its application to HCPB.

Dose-dependent responses of I3C and DIM on T-cell activation in the human T lymphocyte Jurkat cell line

USDA-ARS?s Scientific Manuscript database

Indole-3-carbinol (I3C) and its dimer diindolylmethane (DIM) are bioactive metabolites of a glucosinolate glucobrassicin found in cruciferous vegetables. Both I3C and DIM have been reported to possess anti-apoptotic, anti-proliferative and anti-carcinogenic properties via the modulation of immune p...

Glucosinolates in Brassica vegetables: the influence of the food supply chain on intake, bioavailability and human health.

PubMed

Verkerk, Ruud; Schreiner, Monika; Krumbein, Angelika; Ciska, Ewa; Holst, Birgit; Rowland, Ian; De Schrijver, Remi; Hansen, Magnor; Gerhäuser, Clarissa; Mithen, Richard; Dekker, Matthijs

2009-09-01

Glucosinolates (GLSs) are found in Brassica vegetables. Examples of these sources include cabbage, Brussels sprouts, broccoli, cauliflower and various root vegetables (e.g. radish and turnip). A number of epidemiological studies have identified an inverse association between consumption of these vegetables and the risk of colon and rectal cancer. Animal studies have shown changes in enzyme activities and DNA damage resulting from consumption of Brassica vegetables or isothiocyanates, the breakdown products (BDP) of GLSs in the body. Mechanistic studies have begun to identify the ways in which the compounds may exert their protective action but the relevance of these studies to protective effects in the human alimentary tract is as yet unproven. In vitro studies with a number of specific isothiocyanates have suggested mechanisms that might be the basis of their chemoprotective effects. The concentration and composition of the GLSs in different plants, but also within a plant (e.g. in the seeds, roots or leaves), can vary greatly and also changes during plant development. Furthermore, the effects of various factors in the supply chain of Brassica vegetables including breeding, cultivation, storage and processing on intake and bioavailability of GLSs are extensively discussed in this paper.

The biosynthetic gene cluster for the cyanogenic glucoside dhurrin in Sorghum bicolor contains its co-expressed vacuolar MATE transporter

PubMed Central

Darbani, Behrooz; Motawia, Mohammed Saddik; Olsen, Carl Erik; Nour-Eldin, Hussam H.; Møller, Birger Lindberg; Rook, Fred

2016-01-01

Genomic gene clusters for the biosynthesis of chemical defence compounds are increasingly identified in plant genomes. We previously reported the independent evolution of biosynthetic gene clusters for cyanogenic glucoside biosynthesis in three plant lineages. Here we report that the gene cluster for the cyanogenic glucoside dhurrin in Sorghum bicolor additionally contains a gene, SbMATE2, encoding a transporter of the multidrug and toxic compound extrusion (MATE) family, which is co-expressed with the biosynthetic genes. The predicted localisation of SbMATE2 to the vacuolar membrane was demonstrated experimentally by transient expression of a SbMATE2-YFP fusion protein and confocal microscopy. Transport studies in Xenopus laevis oocytes demonstrate that SbMATE2 is able to transport dhurrin. In addition, SbMATE2 was able to transport non-endogenous cyanogenic glucosides, but not the anthocyanin cyanidin 3-O-glucoside or the glucosinolate indol-3-yl-methyl glucosinolate. The genomic co-localisation of a transporter gene with the biosynthetic genes producing the transported compound is discussed in relation to the role self-toxicity of chemical defence compounds may play in the formation of gene clusters. PMID:27841372

Thermally induced degradation of sulfur-containing aliphatic glucosinolates in broccoli sprouts (Brassica oleracea var. italica) and model systems.

PubMed

Hanschen, Franziska S; Platz, Stefanie; Mewis, Inga; Schreiner, Monika; Rohn, Sascha; Kroh, Lothar W

2012-03-07

Processing reduces the glucosinolate (GSL) content of plant food, among other aspects due to thermally induced degradation. Since there is little information about the thermal stability of GSL and formation of corresponding breakdown products, the thermally induced degradation of sulfur-containing aliphatic GSL was studied in broccoli sprouts and with isolated GSL in dry medium at different temperatures as well as in aqueous medium at different pH values. Desulfo-GSL have been analyzed with HPLC-DAD, while breakdown products were estimated using GC-FID. Whereas in the broccoli sprouts structural differences of the GSL with regard to thermal stability exist, the various isolated sulfur-containing aliphatic GSL degraded nearly equally and were in general more stable. In broccoli sprouts, methylsulfanylalkyl GSL were more susceptible to degradation at high temperatures, whereas methylsulfinylalkyl GSL were revealed to be more affected in aqueous medium under alkaline conditions. Besides small amounts of isothiocyanates, the main thermally induced breakdown products of sulfur-containing aliphatic GSL were nitriles. Although they were most rapidly formed at comparatively high temperatures under dry heat conditions, their highest concentrations were found after cooking in acidic medium, conditions being typical for domestic processing.

Evaluation of genotypic variation of broccoli (Brassica oleracea var. italic) in response to selenium treatment.

PubMed

Ramos, Silvio J; Yuan, Youxi; Faquin, Valdemar; Guilherme, Luiz Roberto G; Li, Li

2011-04-27

Broccoli (Brassica oleracea var. italic) fortified with selenium (Se) has been promoted as a functional food. Here, we evaluated 38 broccoli accessions for their capacity to accumulate Se and for their responses to selenate treatment in terms of nutritional qualities and sulfur gene expresion. We found that the total Se content varied with over 2-fold difference among the leaf tissues of broccoli accessions when the plants were treated with 20 μM Na(2)SeO(4). Approximately half of total Se accumulated in leaves was Se-methylselenocysteine and selenomethionine. Transcriptional regulation of adenosine 5'-phosphosulfate sulfurylase and selenocysteine Se-methyltransferase gene expression might contribute to the different levels of Se accumulation in broccoli. Total glucosinolate contents were not affected by the concentration of selenate application for the majority of broccoli accessions. Essential micronutrients (i.e., Fe, Zn, Cu, and Mn) remained unchanged among half of the germplasm. Moreover, the total antioxidant capacity was greatly stimulated by selenate in over half of the accessions. The diverse genotypic variation in Se, glucosinolate, and antioxidant contents among accessions provides the opportunity to breed broccoli cultivars that simultaneously accumulate Se and other health benefit compounds.

Development and evaluation of event-specific quantitative PCR method for genetically modified soybean A2704-12.

PubMed

Takabatake, Reona; Akiyama, Hiroshi; Sakata, Kozue; Onishi, Mari; Koiwa, Tomohiro; Futo, Satoshi; Minegishi, Yasutaka; Teshima, Reiko; Mano, Junichi; Furui, Satoshi; Kitta, Kazumi

2011-01-01

A novel real-time PCR-based analytical method was developed for the event-specific quantification of a genetically modified (GM) soybean event; A2704-12. During the plant transformation, DNA fragments derived from pUC19 plasmid were integrated in A2704-12, and the region was found to be A2704-12 specific. The pUC19-derived DNA sequences were used as primers for the specific detection of A2704-12. We first tried to construct a standard plasmid for A2704-12 quantification using pUC19. However, non-specific signals appeared with both qualitative and quantitative PCR analyses using the specific primers with pUC19 as a template, and we then constructed a plasmid using pBR322. The conversion factor (C(f)), which is required to calculate the amount of the genetically modified organism (GMO), was experimentally determined with two real-time PCR instruments, the Applied Biosystems 7900HT and the Applied Biosystems 7500. The determined C(f) values were both 0.98. The quantitative method was evaluated by means of blind tests in multi-laboratory trials using the two real-time PCR instruments. The limit of quantitation for the method was estimated to be 0.1%. The trueness and precision were evaluated as the bias and reproducibility of relative standard deviation (RSD(R)), and the determined bias and RSD(R) values for the method were each less than 20%. These results suggest that the developed method would be suitable for practical analyses for the detection and quantification of A2704-12.

Real-Time PCR-Based Quantitation Method for the Genetically Modified Soybean Line GTS 40-3-2.

PubMed

Kitta, Kazumi; Takabatake, Reona; Mano, Junichi

2016-01-01

This chapter describes a real-time PCR-based method for quantitation of the relative amount of genetically modified (GM) soybean line GTS 40-3-2 [Roundup Ready(®) soybean (RRS)] contained in a batch. The method targets a taxon-specific soybean gene (lectin gene, Le1) and the specific DNA construct junction region between the Petunia hybrida chloroplast transit peptide sequence and the Agrobacterium 5-enolpyruvylshikimate-3-phosphate synthase gene (epsps) sequence present in GTS 40-3-2. The method employs plasmid pMulSL2 as a reference material in order to quantify the relative amount of GTS 40-3-2 in soybean samples using a conversion factor (Cf) equal to the ratio of the RRS-specific DNA to the taxon-specific DNA in representative genuine GTS 40-3-2 seeds.

QUANTITATION OF CYP1A1 AND 1B1 MRNA IN POLYCYCLIC AROMATIC HYDROCARBON-TREATED HUMAN T-47D AND HEPG2 CELLS BY A MODIFIED BDNA ASSAY USING FLUORESCENCE DETECTION. (R827180)

EPA Science Inventory

The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

Collaborative ring trial of the papaya endogenous reference gene and its polymerase chain reaction assays for genetically modified organism analysis.

PubMed

Wei, Jiaojun; Li, Feiwu; Guo, Jinchao; Li, Xiang; Xu, Junfeng; Wu, Gang; Zhang, Dabing; Yang, Litao

2013-11-27

The papaya (Carica papaya L.) Chymopapain (CHY) gene has been reported as a suitable endogenous reference gene for genetically modified (GM) papaya detection in previous studies. Herein, we further validated the use of the CHY gene and its qualitative and quantitative polymerase chain reaction (PCR) assays through an interlaboratory collaborative ring trial. A total of 12 laboratories working on detection of genetically modified organisms participated in the ring trial and returned test results. Statistical analysis of the returned results confirmed the species specificity, low heterogeneity, and single-copy number of the CHY gene among different papaya varieties. The limit of detection of the CHY qualitative PCR assay was 0.1%, while the limit of quantification of the quantitative PCR assay was ∼25 copies of haploid papaya genome with acceptable PCR efficiency and linearity. The differences between the tested and true values of papaya content in 10 blind samples ranged from 0.84 to 6.58%. These results indicated that the CHY gene was suitable as an endogenous reference gene for the identification and quantification of GM papaya.

Comparison of intradermal dilutional testing, skin prick testing, and modified quantitative testing for common allergens.

PubMed

Peltier, Jacques; Ryan, Matthew W

2007-08-01

To compare and correlate wheal size using the Multi-Test II applicator with the endpoint obtained by intradermal dilutional testing (IDT) for 5 common allergens. To examine the safety of modified quantitative testing (MQT) for determining immunotherapy starting doses. Prospective comparative clinical study. A total of 134 subjects were simultaneously skin tested for immediate hypersensitivity using the Multi-Test II device and IDT. There was a 77% concordance between results from IDT and results from MQT. When there was a difference, MQT predicted a safer endpoint for starting immunotherapy in all but 2 cases. Wheal size by SPT is predictive of endpoint by IDT. MQT is nearly as effective as formal IDT in determining endpoint. Modified quantitative testing appears to be a safe alternative to IDT for determining starting doses for immunotherapy.

A simplified and accurate detection of the genetically modified wheat MON71800 with one calibrator plasmid.

PubMed

Kim, Jae-Hwan; Park, Saet-Byul; Roh, Hyo-Jeong; Park, Sunghoon; Shin, Min-Ki; Moon, Gui Im; Hong, Jin-Hwan; Kim, Hae-Yeong

2015-06-01

With the increasing number of genetically modified (GM) events, unauthorized GMO releases into the food market have increased dramatically, and many countries have developed detection tools for them. This study described the qualitative and quantitative detection methods of unauthorized the GM wheat MON71800 with a reference plasmid (pGEM-M71800). The wheat acetyl-CoA carboxylase (acc) gene was used as the endogenous gene. The plasmid pGEM-M71800, which contains both the acc gene and the event-specific target MON71800, was constructed as a positive control for the qualitative and quantitative analyses. The limit of detection in the qualitative PCR assay was approximately 10 copies. In the quantitative PCR assay, the standard deviation and relative standard deviation repeatability values ranged from 0.06 to 0.25 and from 0.23% to 1.12%, respectively. This study supplies a powerful and very simple but accurate detection strategy for unauthorized GM wheat MON71800 that utilizes a single calibrator plasmid. Copyright © 2014 Elsevier Ltd. All rights reserved.

Modified telomeric repeat amplification protocol: a quantitative radioactive assay for telomerase without using electrophoresis.

PubMed

Szatmari, I; Tókés, S; Dunn, C B; Bardos, T J; Aradi, J

2000-06-15

A polymerase chain reaction (PCR)-based radioactive telomerase assay was developed in our laboratory which is quantitative and does not require electrophoretic evaluation (designated as TP-TRAP; it utilizes two reverse primers). The main steps of the assay include (1) extension of a 20-mer oligonucleotide substrate (MTS) by telomerase, (2) amplification of the telomerase products in the presence of [(3)H]dTTP using the substrate oligonucleotide and two reverse primers (RPC3, 38 mer; RP, 20 mer), (3) isolation of the amplified radioactive dsDNA by precipitation and filtration, (4) determination of the radioactivity of the acid-insoluble DNA. The length of the telomerase products does not increase on amplification. This valuable feature of the assay is achieved by utilization of the two reverse primers and a highly specific PCR protocol. The assay is linear, accurate, and suitable for cell-biological studies where slight quantitative differences in telomerase activity must be detected. The assay is also suitable for screening and characterization of telomerase inhibitors, as shown with a chemically modified oligonucleotide reverse transcriptase inhibitor [(s(4)dU)(35)]. Copyright 2000 Academic Press.

International collaborative study of the endogenous reference gene, sucrose phosphate synthase (SPS), used for qualitative and quantitative analysis of genetically modified rice.

PubMed

Jiang, Lingxi; Yang, Litao; Zhang, Haibo; Guo, Jinchao; Mazzara, Marco; Van den Eede, Guy; Zhang, Dabing

2009-05-13

One rice ( Oryza sativa ) gene, sucrose phosphate synthase (SPS), has been proven to be a suitable endogenous reference gene for genetically modified (GM) rice detection in a previous study. Herein are the reported results of an international collaborative ring trial for validation of the SPS gene as an endogenous reference gene and its optimized qualitative and quantitative polymerase chain reaction (PCR) systems. A total of 12 genetically modified organism (GMO) detection laboratories from seven countries participated in the ring trial and returned their results. The validated results confirmed the species specificity of the method through testing 10 plant genomic DNAs, low heterogeneity, and a stable single-copy number of the rice SPS gene among 7 indica varieties and 5 japonica varieties. The SPS qualitative PCR assay was validated with a limit of detection (LOD) of 0.1%, which corresponded to about 230 copies of haploid rice genomic DNA, while the limit of quantification (LOQ) for the quantitative PCR system was about 23 copies of haploid rice genomic DNA, with acceptable PCR efficiency and linearity. Furthermore, the bias between the test and true values of eight blind samples ranged from 5.22 to 26.53%. Thus, we believe that the SPS gene is suitable for use as an endogenous reference gene for the identification and quantification of GM rice and its derivates.

Synthesis of Dendronized Poly(l-Glutamate) via Azide-Alkyne Click Chemistry

PubMed Central

Perdih, Peter; Kržan, Andrej; Žagar, Ema

2016-01-01

Poly(l-glutamate) (PGlu) was modified with a second-generation dendron to obtain the dendronized polyglutamate, P(Glu-D). Synthesized P(Glu-D) exhibited a degree of polymerization (DPn) of 46 and a 43% degree of dendronization. Perfect agreement was found between the P(Glu-D) expected structure and the results of nuclear magnetic resonance spectroscopy (NMR) and size-exclusion chromatography coupled to a multi-angle light-scattering detector (SEC-MALS) analysis. The PGlu precursor was modified by coupling with a bifunctional building block (N3-Pr-NH2) in the presence of 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM) coupling reagent. The second-generation polyamide dendron was prepared by a stepwise procedure involving the coupling of propargylamine to the l-lysine carboxyl group, followed by attaching the protected 2,2-bis(methylol)propionic acid (bis-MPA) building block to the l-lysine amino groups. The hydroxyl groups of the resulting second-generation dendron were quantitatively deprotected under mild acidic conditions. The deprotected dendron with an acetylene focal group was coupled to the pendant azide groups of the modified linear copolypeptide, P(Glu-N3), in a Cu(I) catalyzed azide-alkyne cycloaddition reaction to form a 1,4-disubstituted triazole. The dendronization reaction proceeded quantitatively in 48 hours in aqueous medium as confirmed by 1H NMR and Fourier transform infrared spectroscopy (FT-IR) spectroscopy. PMID:28773369

Quantitative detection method for Roundup Ready soybean in food using duplex real-time PCR MGB chemistry.

PubMed

Samson, Maria Cristina; Gullì, Mariolina; Marmiroli, Nelson

2010-07-01

Methodologies that enable the detection of genetically modified organisms (GMOs) (authorized and non-authorized) in food and feed strongly influence the potential for adequate updating and implementation of legislation together with labeling requirements. Quantitative polymerase chain reaction (qPCR) systems were designed to boost the sensitivity and specificity on the identification of GMOs in highly degraded DNA samples; however, such testing will become economically difficult to cope with due to increasing numbers of approved genetically modified (GM) lines. Multiplexing approaches are therefore in development to provide cost-efficient solution. Construct-specific primers and probe were developed for quantitative analysis of Roundup Ready soybean (RRS) event glyphosate-tolerant soybean (GTS) 40-3-2. The lectin gene (Le1) was used as a reference gene, and its specificity was verified. RRS- and Le1-specific quantitative real-time PCR (qRTPCR) were optimized in a duplex platform that has been validated with respect to limit of detection (LOD) and limit of quantification (LOQ), as well as accuracy. The analysis of model processed food samples showed that the degradation of DNA has no adverse or little effects on the performance of quantification assay. In this study, a duplex qRTPCR using TaqMan minor groove binder-non-fluorescent quencher (MGB-NFQ) chemistry was developed for specific detection and quantification of RRS event GTS 40-3-2 that can be used for practical monitoring in processed food products.

Versatile quantitative phase imaging system applied to high-speed, low noise and multimodal imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

Federici, Antoine; Aknoun, Sherazade; Savatier, Julien; Wattellier, Benoit F.

2017-02-01

Quadriwave lateral shearing interferometry (QWLSI) is a well-established quantitative phase imaging (QPI) technique based on the analysis of interference patterns of four diffraction orders by an optical grating set in front of an array detector [1]. As a QPI modality, this is a non-invasive imaging technique which allow to measure the optical path difference (OPD) of semi-transparent samples. We present a system enabling QWLSI with high-performance sCMOS cameras [2] and apply it to perform high-speed imaging, low noise as well as multimodal imaging. This modified QWLSI system contains a versatile optomechanical device which images the optical grating near the detector plane. Such a device is coupled with any kind of camera by varying its magnification. In this paper, we study the use of a sCMOS Zyla5.5 camera from Andor along with our modified QWLSI system. We will present high-speed live cell imaging, up to 200Hz frame rate, in order to follow intracellular fast motions while measuring the quantitative phase information. The structural and density information extracted from the OPD signal is complementary to the specific and localized fluorescence signal [2]. In addition, QPI detects cells even when the fluorophore is not expressed. This is very useful to follow a protein expression with time. The 10 µm spatial pixel resolution of our modified QWLSI associated to the high sensitivity of the Zyla5.5 enabling to perform high quality fluorescence imaging, we have carried out multimodal imaging revealing fine structures cells, like actin filaments, merged with the morphological information of the phase. References [1]. P. Bon, G. Maucort, B. Wattellier, and S. Monneret, "Quadriwave lateral shearing interferometry for quantitative phase microscopy of living cells," Opt. Express, vol. 17, pp. 13080-13094, 2009. [2] P. Bon, S. Lécart, E. Fort and S. Lévêque-Fort, "Fast label-free cytoskeletal network imaging in living mammalian cells," Biophysical journal, 106(8), pp. 1588-1595, 2014

Indole Compounds Related to Auxins and Goitrogens of Woad (Isatis tinctoria L.) 1

PubMed Central

Elliott, Malcolm C.; Stowe, Bruce B.

1971-01-01

Five conspicuous indole derivatives are present in leaves and other tissues of woad (Isatis tinctoria L.). They were identified as tryptophan, isatan B, glucobrassicin, neoglucobrassicin, and glucobrassicin-1-sulfonate. The latter three indole glucosinolates are present at levels of at least 260, 69, and 200 milligrams per kilogram fresh weight and were isolated as crystalline salts. Comparison of physical and chemical properties, particularly NMR spectral analysis, confirms that the 1-methoxyglucobrassicin structure suggested for neoglucobrassicin is correct, whereas further evidence for the even more unusual sulfonation of the ring nitrogen in glucobrassicin-1-sulfonate was obtained. Glucobrassicin-1-sulfonate has an enzymic degradation pattern identical to that of glucobrassicin. As it too releases thiocyanate, it must be added to the list of known plant goitrogens. These studies and the techniques described establish woad as exceptionally suitable higher plant material for metabolic studies of indoles related to goitrogens and auxins. PMID:16657624

Absolute quantitation of NAPQI-modified rat serum albumin by LC-MS/MS: monitoring acetaminophen covalent binding in vivo.

PubMed

LeBlanc, André; Shiao, Tze Chieh; Roy, René; Sleno, Lekha

2014-09-15

Acetaminophen is known to cause hepatoxicity via the formation of a reactive metabolite, N-acetyl p-benzoquinone imine (NAPQI), as a result of covalent binding to liver proteins. Serum albumin (SA) is known to be covalently modified by NAPQI and is present at high concentrations in the bloodstream and is therefore a potential biomarker to assess the levels of protein modification by NAPQI. A newly developed method for the absolute quantitation of serum albumin containing NAPQI covalently bound to its active site cysteine (Cys34) is described. This optimized assay represents the first absolute quantitation of a modified protein, with very low stoichiometric abundance, using a protein-level standard combined with isotope dilution. The LC-MS/MS assay is based on a protein standard modified with a custom-designed reagent, yielding a surrogate peptide (following digestion) that is a positional isomer to the target peptide modified by NAPQI. To illustrate the potential of this approach, the method was applied to quantify NAPQI-modified SA in plasma from rats dosed with acetaminophen. The resulting method is highly sensitive (capable of quantifying down to 0.0006% of total RSA in its NAPQI-modified form) and yields excellent precision and accuracy statistics. A time-course pharmacokinetic study was performed to test the usefulness of this method for following acetaminophen-induced covalent binding at four dosing levels (75-600 mg/kg IP), showing the viability of this approach to directly monitor in vivo samples. This approach can reliably quantify NAPQI-modified albumin, allowing direct monitoring of acetaminophen-related covalent binding.

Preservation of bioactive compounds of a green vegetable smoothie using short time-high temperature mild thermal treatment.

PubMed

Castillejo, Noelia; Martínez-Hernández, Ginés Benito; Monaco, Kamila; Gómez, Perla A; Aguayo, Encarna; Artés, Francisco; Artés-Hernández, Francisco

2017-01-01

Smoothies represent an excellent and convenient alternative to promote the daily consumption of fruit and vegetables in order to obtain their health-promoting benefits. Accordingly, a green fresh vegetables smoothie (77.2% cucumber, 12% broccoli and 6% spinach) rich in health-promoting compounds was developed. Soluble solids content, pH and titratable acidity of the smoothie were 4.3 ± 0.4°Bx, 4.49 ± 0.01 and 0.22 ± 0.02 mg citric acid 100 -1 g fw, respectively. Two thermal treatments to reduce microbial loads and preserve quality were assayed: T1 (3 min at 80 ℃) and T2 (45 s at 90 ℃). Fresh blended unheated samples were used as control (CTRL). The smoothie presented a viscoelastic behaviour. T1 and T2 treatments reduced initial microbial loads by 1.3-2.4 and 1.4-3.1 log units, respectively. Samples were stored in darkness at 5 and 15 ℃. Colour and physicochemical changes were reduced in thermal-treated samples throughout storage, which were better preserved at 5 ℃ rather than at 15 ℃. Vitamin C changes during storage were fitted with a Weibullian distribution. Total vitamin C losses of T1 and T2 samples during storage at 15 ℃ were greatly reduced when they were stored at 5 ℃. Initial total phenolic content (151.1 ± 4.04 mg kg -1 fw) was 44 and 36% increased after T1 and T2 treatments, respectively. The 3-p-coumaroyl quinic and chlorogenic acids accounted the 84.7 and 7.1% relative abundance, respectively. Total antioxidant capacity (234.2 ± 20.3 mg Trolox equivalent kg -1 fw) remained constant after the thermal treatments and was better maintained during storage in thermal-treated samples. Glucobrassicin accounted the 81% of the initial total glucosinolates content (117.8 ± 22.2 mg kg -1 fw) of the smoothie. No glucosinolates losses were observed after T2 treatment being better preserved in thermal-treated samples. Conclusively, a short time-high temperature mild thermal treatment (T2) showed better quality and bioactive compounds retention in a green fresh vegetable smoothie during low temperature storage. © The Author(s) 2016.

A new separation and preconcentration method for selenium in some foods using modified silica gel with 2,6-diamino-4-phenil-1,3,5-triazine.

PubMed

Mendil, Durali; Demirci, Zafer; Uluozlu, Ozgur Dogan; Tuzen, Mustafa; Soylak, Mustafa

2017-04-15

A novel and simple solid phase extraction method was improved and recommended for selenium. Silica gel was modified with 2,6-diamino-4-phenil-1,3,5-triazine and characterized by FTIR, SEM and elemental analysis and used adsorbent for column solid phase extraction of selenium ions. The experimental parameters (pH, flow rates, amounts of the modified silica gel, concentration and type of eluent, volume of sample, etc.) on the recoveries of selenium were optimized. Standard reference materials were analyzed for validation of method. The present method was successfully applied to the detection of total selenium in water and microwave digested some food samples with quantitative recoveries (> 95%). The relative standard deviations were<8%. Matrix influences were not observed. The adsorption capacity of modified silica gel was 5.90mgg -1 . The LOD was 0.015μgL -1 . Enrichment factor was obtained as 50 for the introduced method. Copyright © 2016 Elsevier Ltd. All rights reserved.

Notes on Experiments.

ERIC Educational Resources Information Center

Physics Education, 1979

1979-01-01

Describes the following: use and construction of a lens-pinhole spatial filter assembly to produce expanded beams; how to modify a unilab V. L. F. oscillator to give variable frequencies between .1 Hz and 10 Hz; to use Crookes radiometer quantitatively; and an externally located, movable probe for plasma physics experiments, using conventional…

Quantitative analysis of modified proteins and their positional isomers by tandem mass spectrometry: human histone H4.

PubMed

Pesavento, James J; Mizzen, Craig A; Kelleher, Neil L

2006-07-01

Here we show that fragment ion abundances from dissociation of ions created from mixtures of multiply modified histone H4 (11 kDa) or of N-terminal synthetic peptides (2 kDa) correspond to their respective intact ion abundances measured by Fourier transform mass spectrometry. Isomeric mixtures of modified forms of the same protein are resolved and quantitated with a precision of

Effect of storage time on metabolite profile and alpha-glucosidase inhibitory activity of Cosmos caudatus leaves - GCMS based metabolomics approach.

PubMed

Javadi, Neda; Abas, Faridah; Mediani, Ahmed; Abd Hamid, Azizah; Khatib, Alfi; Simoh, Sanimah; Shaari, Khozirah

2015-09-01

Cosmos caudatus, which is a commonly consumed vegetable in Malaysia, is locally known as "Ulam Raja". It is a local Malaysian herb traditionally used as a food and medicinal herb to treat several maladies. Its bioactive or nutritional constituents consist of a wide range of metabolites, including glucosinolates, phenolics, amino acids, organic acids, and sugars. However, many of these metabolites are not stable and easily degraded or modified during storage. In order to investigate the metabolomics changes occurring during post-harvest storage, C. caudatus samples were subjected to seven different storage times (0 hours, 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, and 12 hours) at room temperature. As the model experiment, the metabolites identified by gas chromatography-mass spectrometry (GC-MS) were correlated with α-glucosidase inhibitory activity analyzed with multivariate data analysis (MVDA) to find out the variation among samples and metabolites contributing to the activity. Orthogonal partial least squares (OPLS) analysis was applied to investigate the metabolomics changes. A profound chemical alteration, both in primary and secondary metabolites, was observed. The α-tocopherol, catechin, cyclohexen-1-carboxylic acid, benzoic acid, myo-inositol, stigmasterol, and lycopene compounds were found to be the discriminating metabolites at early storage; however, sugars such as sucrose, α-d-galactopyranose, and turanose were detected, which was attributed to the discriminating metabolites for late storage. The result shows that the MVDA method is a promising technique to identify biomarker compounds relative to storage at different times. Copyright © 2015. Published by Elsevier B.V.

Chemical Diversity and Defence Metabolism: How Plants Cope with Pathogens and Ozone Pollution

PubMed Central

Iriti, Marcello; Faoro, Franco

2009-01-01

Chemical defences represent a main trait of the plant innate immune system. Besides regulating the relationship between plants and their ecosystems, phytochemicals are involved both in resistance against pathogens and in tolerance towards abiotic stresses, such as atmospheric pollution. Plant defence metabolites arise from the main secondary metabolic routes, the phenylpropanoid, the isoprenoid and the alkaloid pathways. In plants, antibiotic compounds can be both preformed (phytoanticipins) and inducible (phytoalexins), the former including saponins, cyanogenic glycosides and glucosinolates. Chronic exposure to tropospheric ozone (O3) stimulates the carbon fluxes from the primary to the secondary metabolic pathways to a great extent, inducing a shift of the available resources in favour of the synthesis of secondary products. In some cases, the plant defence responses against pathogens and environmental pollutants may overlap, leading to the unspecific synthesis of similar molecules, such as phenylpropanoids. Exposure to ozone can also modify the pattern of biogenic volatile organic compounds (BVOC), emitted from plant in response to herbivore feeding, thus altering the tritrophic interaction among plant, phytophagy and their natural enemies. Finally, the synthesis of ethylene and polyamines can be regulated by ozone at level of S-adenosylmethionine (SAM), the biosynthetic precursor of both classes of hormones, which can, therefore, mutually inhibit their own biosynthesis with consequence on plant phenotype. PMID:20111684

Rates of in vivo (arterial) and in vitro biocorrosion for pure magnesium.

PubMed

Bowen, Patrick K; Drelich, Adam; Drelich, Jaroslaw; Goldman, Jeremy

2015-01-01

The development of magnesium-based materials for bioabsorbable stents relies heavily on corrosion testing by immersion in pseudophysiological solutions, where magnesium degrades faster than it does in vivo. The quantitative difference in corrosion kinetics in vitro and in vivo is largely unknown, but, if determined, would help reduce dependence on animal models. In order to create a quantitative in vitro-in vivo correlation based on an accepted measure of corrosion (penetration rate), commercially pure magnesium wires were corroded in vivo in the abdominal aortas of rats for 5-32 days, and in vitro for up to 14 days using Dulbecco's modified eagle medium. Cross-sectioning, scanning electron microscopy, image analysis, a modified penetration rate tailored to degraded wires, and empirical modeling were used to analyze the corroded specimens. In vitro penetration rates were consistently higher than comparable in vivo rates by a factor of 1.2-1.9× (±0.2×). For a sample <20% corroded, an approximate in vitro-in vivo multiplier of 1.3 ± 0.2× was applied, whereas a multiplier of 1.8 ± 0.2× became appropriate when the magnesium specimen was 25-35% degraded. © 2014 Wiley Periodicals, Inc.

Integration of genetic, genomic and transcriptomic information identifies putative regulators of adventitious root formation in Populus

DOE PAGES

Ribeiro, Cintia L.; Silva, Cynthia M.; Drost, Derek R.; ...

2016-03-16

In this study, adventitious roots (AR) develop from tissues other than the primary root, in a process physiologically regulated by phytohormones. Adventitious roots provide structural support and contribute to water and nutrient absorption, and are critical for commercial vegetative propagation of several crops. Here we quantified the number of AR, root architectural traits and root biomass in cuttings from a pseudo-backcross population of Populus deltoides and Populus trichocarpa. Quantitative trait loci (QTL) mapping and whole-transcriptome analysis of individuals with alternative QTL alleles for AR number were used to identify putative regulators of AR development. As a result, parental individuals andmore » progeny showed extensive segregation for AR developmental traits. Quantitative trait loci for number of AR mapped consistently in the same interval of linkage group (LG) II and LG XIV, explaining 7–10 % of the phenotypic variation. A time series transcriptome analysis identified 26,121 genes differentially expressed during AR development, particularly during the first 24 h after cuttings were harvested. Of those, 1929 genes were differentially regulated between individuals carrying alternative alleles for the two QTL for number of AR, in one or more time point. Eighty-one of these genes were physically located within the QTL intervals for number of AR, including putative homologs of the Arabidopsis genes SUPERROOT2 (SUR2) and TRYPTOPHAN SYNTHASE ALPHA CHAIN (TSA1), both of which are involved in the auxin indole-3-acetic acid (IAA) biosynthesis pathway. In conclusion, this study suggests the involvement of two genes of the tryptophan-dependent auxin biosynthesis pathway, SUR2 and TSA1, in the regulation of a critical trait for the clonal propagation of woody species. A possible model for this regulation is that poplar individuals that have poor AR formation synthesize auxin indole-3-acetic acid (IAA) primarily through the tryptophan (Trp) pathway. Much of the Trp pathway flux appears to be directed to the synthesis of indole glucosinolates (IG), as suggested by the over-expression of SUR2. Individuals that are efficient in AR formation may utilize alternative (non-Trp) pathways to synthesize IAA, based on the observation that they down-regulate the expression of TSA1, one of the critical steps in the synthesis of tryptophan.« less

Indole Compounds Related to Auxins and Goitrogens of Woad (Isatis tinctoria L.).

PubMed

Elliott, M C; Stowe, B B

1971-03-01

Five conspicuous indole derivatives are present in leaves and other tissues of woad (Isatis tinctoria L.). They were identified as tryptophan, isatan B, glucobrassicin, neoglucobrassicin, and glucobrassicin-1-sulfonate. The latter three indole glucosinolates are present at levels of at least 260, 69, and 200 milligrams per kilogram fresh weight and were isolated as crystalline salts. Comparison of physical and chemical properties, particularly NMR spectral analysis, confirms that the 1-methoxyglucobrassicin structure suggested for neoglucobrassicin is correct, whereas further evidence for the even more unusual sulfonation of the ring nitrogen in glucobrassicin-1-sulfonate was obtained. Glucobrassicin-1-sulfonate has an enzymic degradation pattern identical to that of glucobrassicin. As it too releases thiocyanate, it must be added to the list of known plant goitrogens. These studies and the techniques described establish woad as exceptionally suitable higher plant material for metabolic studies of indoles related to goitrogens and auxins.

Macrophage phenotype in the epigallocatechin-3-gallate (EGCG)-modified collagen determines foreign body reaction.

PubMed

Chu, Chenyu; Liu, Li; Wang, Yufei; Wei, Shimin; Wang, Yuanjing; Man, Yi; Qu, Yili

2018-04-28

Collagen has been widely used in guided bone regeneration, and the implantation of collagen membranes will elicit the foreign body reaction (FBR). The imbalance of FBR often leads to failure of dental implants. Therefore, modulation of the FBR after implantation of collagen membranes becomes increasingly important. Macrophages, pivotal in FBR, have been distinguished into pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes. Epigallocatechin-3-gallate (EGCG)-modified collagen membranes have been previously shown to regulate secretion of inflammatory factors. In this study, immunohistochemistry of CD31 showed that areas of blood vessels were significantly enlarged after implantation of EGCG-modified collagen membranes compared with those treated with pure collagen membranes. Besides, haematoxylin-eosin staining and immunofluorescence showed an increased number of M2 macrophages after implantation of EGCG-modified collagen membranes. In addition, quantitative real-time polymerase chain reaction showed that after implantation of EGCG-modified collagen membranes, expression of CXCL1 (predominant chemoattractants to neutrophils and inflammation promotors) was significantly downregulated, whereas expressions of STAB1, CCR2, CCR3, CCL2, and CCL3 (related to M2 macrophages) were significantly upregulated. From these findings, we conclude that EGCG-modified collagen membranes were able to regulate the recruitment and polarization of macrophages, so that ameliorate FBR. Copyright © 2018 John Wiley & Sons, Ltd.

Glutathione Transferase U13 Functions in Pathogen-Triggered Glucosinolate Metabolism.

PubMed

Piślewska-Bednarek, Mariola; Nakano, Ryohei Thomas; Hiruma, Kei; Pastorczyk, Marta; Sanchez-Vallet, Andrea; Singkaravanit-Ogawa, Suthitar; Ciesiołka, Danuta; Takano, Yoshitaka; Molina, Antonio; Schulze-Lefert, Paul; Bednarek, Paweł

2018-01-01

Glutathione (GSH) and indole glucosinolates (IGs) exert key functions in the immune system of the model plant Arabidopsis ( Arabidopsis thaliana ). Appropriate GSH levels are important for execution of both pre- and postinvasive disease resistance mechanisms to invasive pathogens, whereas an intact PENETRATION2 (PEN2)-pathway for IG metabolism is essential for preinvasive resistance in this species. Earlier indirect evidence suggested that the latter pathway involves conjugation of GSH with unstable products of IG metabolism and further processing of the resulting adducts to biologically active molecules. Here we describe the identification of Glutathione- S -Transferase class-tau member 13 (GSTU13) as an indispensable component of the PEN2 immune pathway for IG metabolism. gstu13 mutant plants are defective in the pathogen-triggered biosynthesis of end products of the PEN2 pathway, including 4-O-β-d-glucosyl-indol-3-yl formamide, indole-3-ylmethyl amine, and raphanusamic acid. In line with this metabolic defect, lack of functional GSTU13 results in enhanced disease susceptibility toward several fungal pathogens including Erysiphe pisi , Colletotrichum gloeosporioides , and Plectosphaerella cucumerina Seedlings of gstu13 plants fail also to deposit the (1,3)-β-glucan cell wall polymer, callose, after recognition of the bacterial flg22 epitope. We show that GSTU13 mediates specifically the role of GSH in IG metabolism without noticeable impact on other immune functions of this tripeptide. We postulate that GSTU13 connects GSH with the pathogen-triggered PEN2 pathway for IG metabolism to deliver metabolites that may have numerous functions in the innate immune system of Arabidopsis. © 2018 American Society of Plant Biologists. All Rights Reserved.

A modified ion-selective electrode method for measurement of chloride in sweat.

PubMed

Finley, P R; Dye, J A; Lichti, D A; Byers, J M; Williams, R J

1978-06-01

A modified method of analysis of sweat chloride concentration with an ion-selective electrode is presented. The original method of sweat chloride analysis proposed by the Orion Research Corporation (Cambridge, Massachusetts 02139) is inadequate because it produces erratic and misleading results. The modified method was compared with the reference quantitative method of Gibson and Cooke. In the modified method, individual electrode pads are cut and placed in the electrodes rather than using the pads supplied by the company; pilocarpine nitrate (2,000 mg/l) is used in place of pilocarpine HCl (640 mg/l); sodium bicarbonate as the weak electrolyte is used instead of K2SO4. A 10-minute period for sweat accumulation is employed rather than a zero-time collection as in the original Orion method. The modification has been studied for reproducibility in individuals, reproducibility between right and left arm in individuals; it has been compared extensively with the quantitative method of Gibson and Cooke, both in normal individuals and in patients with cystic fibrosis. There is excellent agreement between the modified method and the quantitative reference method. There appears to be a slight bias toward higher concentrations of chloride from the right arm compared with the left arm, but this difference is not medically significant.

Bovine viral diarrhea virus fetal persistent infection after immunization with a contaminated modified-live virus vaccine.

PubMed

Palomares, Roberto A; Marley, Shonda M; Givens, M Daniel; Gallardo, Rodrigo A; Brock, Kenny V

2013-05-01

The objective was to determine whether a multivalent modified-live virus vaccine containing noncytopathic bovine viral diarrhea virus (BVDV) administered off-label to pregnant cattle can result in persistently infected fetuses and to assess whether vaccinal strains can be shed to unvaccinated pregnant cattle commingling with vaccinates. Nineteen BVDV-naïve pregnant heifers were randomly assigned to two groups: cattle vaccinated near Day 77 of gestation with modified-live virus vaccine containing BVDV-1a (WRL strain), bovine herpes virus-1, parainfluenza 3, and bovine respiratory syncytial virus (Vx group; N = 10) or control unvaccinated cattle (N = 9). During the course of the study a voluntary stop-sale/recall was conducted by the manufacturer because of the presence of a BVDV contaminant in the vaccine. At Day 175 of gestation, fetuses were removed by Cesarean section and fetal tissues were submitted for virus isolation, and quantitative reverse transcription polymerase chain reaction using BVDV-1- and BVDV-2-specific probes. Nucleotide sequencing of viral RNA was performed for quantitative reverse transcription polymerase chain reaction-positive samples. Two vaccinated and two control heifers aborted their pregnancies, but their fetuses were unavailable for BVDV testing. Virus was isolated from all eight fetuses in the Vx group heifers and from 2 of 7 fetuses in the control unvaccinated heifers. Only BVDV-2 was detected in fetuses from the Vx group, and only BVDV-1 was detected in the two fetuses from the control group. Both BVDV-1 and BVDV-2 were detected in the vaccine. In conclusion, vaccination of pregnant heifers with a contaminated modified-live BVDV vaccine resulted in development of BVDV-2 persistently infected fetuses in all tested vaccinated animals. Furthermore, BVDV was apparently shed to unvaccinated heifers causing fetal infections from which only BVDV-1 was detected. Published by Elsevier Inc.

Improved quantitative recovery of Listeria monocytogenes from stainless steel surfaces using a one-ply composite tissue.

PubMed

Vorst, Keith L; Todd, Ewen C D; Rysert, Elliot T

2004-10-01

Four sampling devices, a sterile environmental sponge (ES), a sterile cotton-tipped swab (CS), a sterile calcium alginate fiber-tipped swab (CAS), and a one-ply composite tissue (CT), were evaluated for quantitative recovery of Listeria monocytogenes from a food-grade stainless steel surface. Sterile 304-grade stainless steel plates (6 by 6 cm) were inoculated with approximately 106 CFU/cm2 L. monocytogenes strain Scott A and dried for 1 h. The ES and CT sampling devices were rehydrated in phosphate buffer solution. After plate swabbing, ES and CT were placed in 40 ml of phosphate buffer solution, stomached for 1 min and hand massaged for 30 s. Each CS and CAS device was rehydrated in 0.1% peptone before swabbing. After swabbing, CS and CAS were vortexed in 0.1% peptone for 1 min. Samples were spiral plated on modified Oxford agar with modified Oxford agar Rodac Contact plates used to recover any remaining cells from the stainless steel surface. Potential inhibition from CT was examined in both phosphate buffer solution and in a modified disc-diffusion assay. Recovery was 2.70, 1.34, and 0.62 log greater using CT compared with ES, CS, and CAS, respectively, with these differences statistically significant (P < 0.001) for ES and CT and for CAS, CS, and CT (P < 0.05). Rodac plates were typically overgrown following ES, positive after CS and CAS, and negative after CT sampling. CT was noninhibitory in both phosphate buffer solution and the modified disc-diffusion assay. Using scanning electron microscopy, Listeria cells were observed on stainless steel plates sampled with each sampling device except CT. The CT device, which is inexpensive and easy to use, represents a major improvement over other methods in quantifying L. monocytogenes on stainless steel surfaces and is likely applicable to enrichment of environmental samples.

Epistasis × environment interactions among Arabidopsis thaliana glucosinolate genes impact complex traits and fitness in the field.

PubMed

Kerwin, Rachel E; Feusier, Julie; Muok, Alise; Lin, Catherine; Larson, Brandon; Copeland, Daniel; Corwin, Jason A; Rubin, Matthew J; Francisco, Marta; Li, Baohua; Joseph, Bindu; Weinig, Cynthia; Kliebenstein, Daniel J

2017-08-01

Despite the growing number of studies showing that genotype × environment and epistatic interactions control fitness, the influences of epistasis × environment interactions on adaptive trait evolution remain largely uncharacterized. Across three field trials, we quantified aliphatic glucosinolate (GSL) defense chemistry, leaf damage, and relative fitness using mutant lines of Arabidopsis thaliana varying at pairs of causal aliphatic GSL defense genes to test the impact of epistatic and epistasis × environment interactions on adaptive trait variation. We found that aliphatic GSL accumulation was primarily influenced by additive and epistatic genetic variation, leaf damage was primarily influenced by environmental variation and relative fitness was primarily influenced by epistasis and epistasis × environment interactions. Epistasis × environment interactions accounted for up to 48% of the relative fitness variation in the field. At a single field site, the impact of epistasis on relative fitness varied significantly over 2 yr, showing that epistasis × environment interactions within a location can be temporally dynamic. These results suggest that the environmental dependency of epistasis can profoundly influence the response to selection, shaping the adaptive trajectories of natural populations in complex ways, and deserves further consideration in future evolutionary studies. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Nutrition versus defense: Why Myzus persicae (green peach aphid) prefers and performs better on young leaves of cabbage

PubMed Central

Cao, He-He; Zhang, Zhan-Feng; Wang, Xiao-Feng

2018-01-01

Plant leaves of different ages differ in nutrients and toxic metabolites and thus exhibit various resistance levels against insect herbivores. However, little is known about the influence of leaf ontogeny on plant resistance to phloem-feeding insects. In this study, we found that the green peach aphid, Myzus persicae, preferred to settle on young cabbage leaves compared with mature or old leaves, although young leaves contained the highest concentration of glucosinolates. Furthermore, aphids feeding on young leaves had higher levels of glucosinolates in their body, but aphids performed better on young leaves in terms of body weight and population growth. Phloem sap of young leaves had higher amino acid:sugar molar ratio than mature leaves, and aphids feeding on young leaves showed two times longer phloem feeding time and five times more honeydew excretion than on other leaves. These results indicate that aphids acquired the highest amount of nutrients and defensive metabolites when feeding on young cabbage leaves that are strong natural plant sinks. Accordingly, we propose that aphids generally prefer to obtain more nutrition rather than avoiding host plant defense, and total amount of nutrition that aphids could obtain is significantly influenced by leaf ontogeny or source-sink status of feeding sites. PMID:29684073

The green peach aphid Myzus persicae perform better on pre-infested Chinese cabbage Brassica pekinensis by enhancing host plant nutritional quality

PubMed Central

Cao, He-He; Liu, Hui-Ru; Zhang, Zhan-Feng; Liu, Tong-Xian

2016-01-01

The green peach aphid, Myzus persicae Sulzer, is a notorious pest on vegetables, which often aggregates in high densities on crop leaves. In this study, we investigated whether M. persicae could suppress the resistance level of Chinese cabbage Brassica pekinensis. M. persicae performed better in terms of weight gain (~33% increase) and population growth (~110% increase) when feeding on previously infested (pre-infested) Chinese cabbage compared with those on non-infested plants. However, when given a choice, 64% of the aphids preferred to settle on non-infested leaves, while 29% of aphids chose pre-infested leaves that had a 2.9 times higher concentration of glucosinolates. Aphid feeding significantly enhanced the amino acid:sugar ratio of phloem sap and the absolute amino acid concentration in plant leaves. Aphid infestation significantly increased the expression levels of salicylic acid (SA) marker genes, while it had marginal effects on the expression of jasmonate marker genes. Exogenously applied SA or methyl jasmonate had no significant effects on M. persicae performance, although these chemicals increased glucosinolates concentration in plant leaves. M. persicae infestation increase amino acid:sugar ratio and activate plant defenses, but aphid performed better on pre-infested plants, suggesting that both nutrition and toxics should be considered in insect-plant interaction. PMID:26905564

Metabolic profiling of glucosinolates and their hydrolysis products in a germplasm collection of Brassica rapa turnips.

PubMed

Klopsch, Rebecca; Witzel, Katja; Börner, Andreas; Schreiner, Monika; Hanschen, Franziska S

2017-10-01

About 10% of the world's vegetable production is generated from Brassicaceae, wherein Brassica rapa is a dominating species. There is growing evidence that glucosinolates (GLSs), main plant secondary metabolites in Brassicales, play an important role in promoting human health. Natural genetic diversity of B. rapa can be explored for vegetable improvement. We analyzed leaves and tubers of 16 B. rapa turnips for their GLS composition by UHPLC-DAD and the corresponding hydrolysis products by GC-MS. Thirteen GLSs were identified, 8 aliphatic, 4 indolic and one aromatic. 3-Butenyl GLS was prevailing in both plant organs while in tubers 2-hydroxy-3-butenyl GLS and 2-phenylethyl GLS occurred in high amounts. A total of 24 GLS breakdown products were detected in tubers and 16 in leaves. Epithionitriles were the main hydrolysis products in both plant organs with 4,5-epithiopentanenitrile and 3-hydroxy-4,5-epithiopentanenitrile being the main compounds. When comparing leaves and tubers, an accumulation of GLSs and their breakdown products was observed in tubers compared to leaves. Our analysis achieved the comprehensive profiling of all GLS metabolites in a collection of B. rapa turnips, underlining the natural variation not only of intact GLS, but also of their breakdown products. Copyright © 2017 Elsevier Ltd. All rights reserved.

SUBCRITICAL (HOT) WATER/ETHANOL EXTRACTION OF NONYLPHENOL POLYETHOXY CARBOXYLATES IN INDUSTRIAL AND MUNICIPAL SLUDGES. (R821195)

EPA Science Inventory

Subcritical (hot) water with ethanol as modifier was used
to extract nonylphenol polyethoxy carboxylates (NPECs)
with 1-4 ethoxy groups from sludge samples. Quantitative
recovery of native NPECs from sludge was accomplished
by extracting 0.25 g samples for 20 min w...

Distance-Based Tear Lactoferrin Assay on Microfluidic Paper Device Using Interfacial Interactions on Surface-Modified Cellulose.

PubMed

Yamada, Kentaro; Henares, Terence G; Suzuki, Koji; Citterio, Daniel

2015-11-11

"Distance-based" detection motifs on microfluidic paper-based analytical devices (μPADs) allow quantitative analysis without using signal readout instruments in a similar manner to classical analogue thermometers. To realize a cost-effective and calibration-free distance-based assay of lactoferrin in human tear fluid on a μPAD not relying on antibodies or enzymes, we investigated the fluidic mobilities of the target protein and Tb(3+) cations used as the fluorescent detection reagent on surface-modified cellulosic filter papers. Chromatographic elution experiments in a tear-like sample matrix containing electrolytes and proteins revealed a collapse of attractive electrostatic interactions between lactoferrin or Tb(3+) and the cellulosic substrate, which was overcome by the modification of the paper surface with the sulfated polysaccharide ι-carrageenan. The resulting μPAD based on the fluorescence emission distance successfully analyzed 0-4 mg mL(-1) of lactoferrin in complex human tear matrix with a lower limit of detection of 0.1 mg mL(-1) by simple visual inspection. Assay results of 18 human tear samples including ocular disease patients and healthy volunteers showed good correlation to the reference ELISA method with a slope of 0.997 and a regression coefficient of 0.948. The distance-based quantitative signal and the good batch-to-batch fabrication reproducibility relying on printing methods enable quantitative analysis by simply reading out "concentration scale marks" printed on the μPAD without performing any calibration and using any signal readout instrument.

Joint analysis of binary and quantitative traits with data sharing and outcome-dependent sampling.

PubMed

Zheng, Gang; Wu, Colin O; Kwak, Minjung; Jiang, Wenhua; Joo, Jungnam; Lima, Joao A C

2012-04-01

We study the analysis of a joint association between a genetic marker with both binary (case-control) and quantitative (continuous) traits, where the quantitative trait values are only available for the cases due to data sharing and outcome-dependent sampling. Data sharing becomes common in genetic association studies, and the outcome-dependent sampling is the consequence of data sharing, under which a phenotype of interest is not measured for some subgroup. The trend test (or Pearson's test) and F-test are often, respectively, used to analyze the binary and quantitative traits. Because of the outcome-dependent sampling, the usual F-test can be applied using the subgroup with the observed quantitative traits. We propose a modified F-test by also incorporating the genotype frequencies of the subgroup whose traits are not observed. Further, a combination of this modified F-test and Pearson's test is proposed by Fisher's combination of their P-values as a joint analysis. Because of the correlation of the two analyses, we propose to use a Gamma (scaled chi-squared) distribution to fit the asymptotic null distribution for the joint analysis. The proposed modified F-test and the joint analysis can also be applied to test single trait association (either binary or quantitative trait). Through simulations, we identify the situations under which the proposed tests are more powerful than the existing ones. Application to a real dataset of rheumatoid arthritis is presented. © 2012 Wiley Periodicals, Inc.

Selective solid-phase extraction using oxidized activated carbon modified with triethylenetetramine for preconcentration of metal ions

NASA Astrophysics Data System (ADS)

Zhang, Li; Chang, Xijun; Li, Zhenhua; He, Qun

2010-02-01

A new selective solid-phase extractant using activated carbon as matrix which was purified, oxidized and modified by triethylenetetramine (AC-TETA) was prepared and characterized by FT-IR spectroscopy. At pH 4, quantitative extraction of trace Cr(III), Fe(III) and Pb(II) was obtained and determined by inductively coupled plasma optical emission spectrometry (ICP-OES). Complete elution of the adsorbed metal ions from the sorbent surface was carried out using 0.5 mol L -1 HCl. The maximum static adsorption capacity of sorbent for Cr(III), Fe(III) and Pb(II) was 34.6, 36.5 and 51.9 mg g -1, respectively. The time of quantitative adsorption was less than 2 min. The detection limits of the method was found to be 0.71, 0.35 and 0.45 ng mL -1 for Cr(III), Fe(III) and Pb(II), and the relative standard deviation (RSD) was 3.7%, 2.2% and 2.5%, respectively. Moreover, the method was free from interference with common coexiting ions. The method was also successfully applied to the preconcentration of trace Cr(III), Fe(III) and Pb(II) in synthetic samples and a real sample with satisfactory results.

Characterization of the exogenous insert and development of event-specific PCR detection methods for genetically modified Huanong No. 1 papaya.

PubMed

Guo, Jinchao; Yang, Litao; Liu, Xin; Guan, Xiaoyan; Jiang, Lingxi; Zhang, Dabing

2009-08-26

Genetically modified (GM) papaya (Carica papaya L.), Huanong No. 1, was approved for commercialization in Guangdong province, China in 2006, and the development of the Huanong No. 1 papaya detection method is necessary for implementing genetically modified organism (GMO) labeling regulations. In this study, we reported the characterization of the exogenous integration of GM Huanong No. 1 papaya by means of conventional polymerase chain reaction (PCR) and thermal asymmetric interlaced (TAIL)-PCR strategies. The results suggested that one intact copy of the initial construction was integrated in the papaya genome and which probably resulted in one deletion (38 bp in size) of the host genomic DNA. Also, one unintended insertion of a 92 bp truncated NptII fragment was observed at the 5' end of the exogenous insert. Furthermore, we revealed its 5' and 3' flanking sequences between the insert DNA and the papaya genomic DNA, and developed the event-specific qualitative and quantitative PCR assays for GM Huanong No. 1 papaya based on the 5' integration flanking sequence. The relative limit of detection (LOD) of the qualitative PCR assay was about 0.01% in 100 ng of total papaya genomic DNA, corresponding to about 25 copies of papaya haploid genome. In the quantitative PCR, the limits of detection and quantification (LOD and LOQ) were as low as 12.5 and 25 copies of papaya haploid genome, respectively. In practical sample quantification, the quantified biases between the test and true values of three samples ranged from 0.44% to 4.41%. Collectively, we proposed that all of these results are useful for the identification and quantification of Huanong No. 1 papaya and its derivates.

The quantitative surface analysis of an antioxidant additive in a lubricant oil matrix by desorption electrospray ionization mass spectrometry

PubMed Central

Da Costa, Caitlyn; Reynolds, James C; Whitmarsh, Samuel; Lynch, Tom; Creaser, Colin S

2013-01-01

RATIONALE Chemical additives are incorporated into commercial lubricant oils to modify the physical and chemical properties of the lubricant. The quantitative analysis of additives in oil-based lubricants deposited on a surface without extraction of the sample from the surface presents a challenge. The potential of desorption electrospray ionization mass spectrometry (DESI-MS) for the quantitative surface analysis of an oil additive in a complex oil lubricant matrix without sample extraction has been evaluated. METHODS The quantitative surface analysis of the antioxidant additive octyl (4-hydroxy-3,5-di-tert-butylphenyl)propionate in an oil lubricant matrix was carried out by DESI-MS in the presence of 2-(pentyloxy)ethyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate as an internal standard. A quadrupole/time-of-flight mass spectrometer fitted with an in-house modified ion source enabling non-proximal DESI-MS was used for the analyses. RESULTS An eight-point calibration curve ranging from 1 to 80 µg/spot of octyl (4-hydroxy-3,5-di-tert-butylphenyl)propionate in an oil lubricant matrix and in the presence of the internal standard was used to determine the quantitative response of the DESI-MS method. The sensitivity and repeatability of the technique were assessed by conducting replicate analyses at each concentration. The limit of detection was determined to be 11 ng/mm2 additive on spot with relative standard deviations in the range 3–14%. CONCLUSIONS The application of DESI-MS to the direct, quantitative surface analysis of a commercial lubricant additive in a native oil lubricant matrix is demonstrated. © 2013 The Authors. Rapid Communications in Mass Spectrometry published by John Wiley & Sons, Ltd. PMID:24097398

Quantitation of CYP1A1 and 1B1 mRNA in polycyclic aromatic hydrocarbon-treated human T-47D and HepG2 cells by a modified bDNA assay using fluorescence detection.

PubMed

Wu, Susan J; Spink, David C; Spink, Barbara C; Kaminsky, Laurence S

2003-01-15

The quantitation of mRNA, essential for assessing mechanisms of enzyme regulation, is normally carried out using reverse transcriptase-polymerase chain reaction (RT-PCR). An alternative method uses a signal-amplification nucleic acid probe assay, which measures RNA directly by the QuantiGene Expression Kit and incorporates branched DNA technology from Bayer and luminometer-based readings of a chemilumigenic alkaline phosphatase substrate. To broaden the utility of this assay, we investigated substitution of a fluorescent substrate, 2'-(2-benzothiazol)-6'-hydroxybenzothiazole phosphate and a fluorometer, and applied the method to quantitation of CYP1A1 and 1B1 mRNA in human T-47D and HepG2 cells following induction by benzo[a]pyrene (B[a]P) and dibenzo[a,h]anthracene (DB[a,h]A). The fluorescence response increased linearly for 200 min without photobleaching and increased linearly (r2=0.997) up to at least 0.2 microg total RNA. The data revealed that at 0.5 and 1.0 microM inducing agent, the induction of CYP1A1 mRNA in HepG2 cells by DB[a,h]A exceeded that by B[a]P by 18- and 6-fold, respectively. In T-47D cells B[a]P induced CYP1A1 mRNA by 23-fold and CYP1B1 mRNA by 3.9-fold. A B[a]P cocontaminant in the environment, arsenite, did not affect B[a]P-induced levels of CYP1A1 or 1B1 mRNA in these cells. The modified analytical system provides a rapid-throughput, reproducible, and less labor-intensive method than RT-PCR for quantifying cellular mRNA levels.

Brassica vegetables as sources of epithionitriles: Novel secondary products formed during cooking.

PubMed

Hanschen, Franziska S; Kaufmann, Martin; Kupke, Franziska; Hackl, Thomas; Kroh, Lothar W; Rohn, Sascha; Schreiner, Monika

2018-04-15

The epithionitriles, 1-cyano-2,3-epithiopropane, in particular, and 1-cyano-3,4-epithiobutane, are important, but yet underestimated glucosinolate hydrolysis products that are released instead of the cancer preventative isothiocyanates in Brassica vegetables, such as cabbage, broccoli, or pak choi. Here, we characterized the reactivity of 1-cyano-2,3-epithiopropane under aqueous heat treatment conditions and compared our findings to those of the related epithionitriles 1-cyano-3,4-epithiobutane and 1-cyano-4,5-epithiopentane. In contrast to the other epithionitriles, 1-cyano-2,3-epithiopropane is highly reactive. As a result, 2-aminothiophene and dimeric 1,4-dithiane-2,5-diacetonitrile were identified as main products and a reaction mechanism is proposed. Formation of 2-aminothiophene was also observed in cooked white cabbage samples. Moreover, three novel compounds were identified as derivatives of the related epithionitriles. The results imply that apart from isothiocyanates, process-derived compounds should be considered in regards to cancer preventative Brassica vegetable related bioactivity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mild hypothermia for treatment of diffuse axonal injury: a quantitative analysis of diffusion tensor imaging

PubMed Central

Jing, Guojie; Yao, Xiaoteng; Li, Yiyi; Xie, Yituan; Li, Wang#x2019;an; Liu, Kejun; Jing, Yingchao; Li, Baisheng; Lv, Yifan; Ma, Baoxin

2014-01-01

Fractional anisotropy values in diffusion tensor imaging can quantitatively reflect the consistency of nerve fibers after brain damage, where higher values generally indicate less damage to nerve fibers. Therefore, we hypothesized that diffusion tensor imaging could be used to evaluate the effect of mild hypothermia on diffuse axonal injury. A total of 102 patients with diffuse axonal injury were randomly divided into two groups: normothermic and mild hypothermic treatment groups. Patient's modified Rankin scale scores 2 months after mild hypothermia were significantly lower than those for the normothermia group. The difference in average fractional anisotropy value for each region of interest before and after mild hypothermia was 1.32-1.36 times higher than the value in the normothermia group. Quantitative assessment of diffusion tensor imaging indicates that mild hypothermia therapy may be beneficial for patients with diffuse axonal injury. PMID:25206800

Semi-quantitative methods yield greater inter- and intraobserver agreement than subjective methods for interpreting 99m technetium-hydroxymethylene-diphosphonate uptake in equine thoracic processi spinosi.

PubMed

van Zadelhoff, Claudia; Ehrle, Anna; Merle, Roswitha; Jahn, Werner; Lischer, Christoph

2018-05-09

Scintigraphy is a standard diagnostic method for evaluating horses with back pain due to suspected thoracic processus spinosus pathology. Lesion detection is based on subjective or semi-quantitative assessments of increased uptake. This retrospective, analytical study is aimed to compare semi-quantitative and subjective methods in the evaluation of scintigraphic images of the processi spinosi in the equine thoracic spine. Scintigraphic images of 20 Warmblood horses, presented for assessment of orthopedic conditions between 2014 and 2016, were included in the study. Randomized, blinded image evaluation was performed by 11 veterinarians using subjective and semi-quantitative methods. Subjective grading was performed for the analysis of red-green-blue and grayscale scintigraphic images, which were presented in full-size or as masked images. For the semi-quantitative assessment, observers placed regions of interest over each processus spinosus. The uptake ratio of each processus spinosus in comparison to a reference region of interest was determined. Subsequently, a modified semi-quantitative calculation was developed whereby only the highest counts-per-pixel for a specified number of pixels was processed. Inter- and intraobserver agreement was calculated using intraclass correlation coefficients. Inter- and intraobserver intraclass correlation coefficients were 41.65% and 71.39%, respectively, for the subjective image assessment. Additionally, a correlation between intraobserver agreement, experience, and grayscale images was identified. The inter- and intraobserver agreement was significantly increased when using semi-quantitative analysis (97.35% and 98.36%, respectively) or the modified semi-quantitative calculation (98.61% and 98.82%, respectively). The proposed modified semi-quantitative technique showed a higher inter- and intraobserver agreement when compared to other methods, which makes it a useful tool for the analysis of scintigraphic images. The association of the findings from this study with clinical and radiological examinations requires further investigation. © 2018 American College of Veterinary Radiology.

A sensitive electrochemical immunosensor based on poly(2-aminobenzylamine) film modified screen-printed carbon electrode for label-free detection of human immunoglobulin G.

PubMed

Putnin, Thitirat; Jumpathong, Watthanachai; Laocharoensuk, Rawiwan; Jakmunee, Jaroon; Ounnunkad, Kontad

2018-08-01

This work focuses on fabricating poly(2-aminobenzylamine)-modified screen-printed carbon electrode as an electrochemical immunosensor for the label-free detection of human immunoglobulin G. To selectively detect immunoglobulin G, the anti-immunoglobulin G antibody with high affinity to immunoglobulin G was covalently linked with the amine group of poly(2-aminobenzylamine) film-deposited screen-printed carbon electrode. The selectivity for immunoglobulin G was subsequently assured by being challenged with redox-active interferences and adventitious adsorption did not significantly interfere the analyte signal. To obviate the use of costly secondary antibody, the [Fe(CN) 6 ] 4-/3- redox probe was instead applied to measure the number of human immunoglobulin G through the immunocomplex formation that is quantitatively related to the level of the differential pulse voltammetric current. The resulting immunosensor exhibited good sensitivity with the detection limit of 0.15 ng mL -1 , limit of quantitation of 0.50 ng mL -1 and the linear range from 1.0 to 50 ng mL -1 . Given those striking analytical performances and the affordability arising from using cheap screen-printed carbon electrode with label-free detection, the immunosensor serves as a promising model for the next-step development of a diagnostic tool.

Selective determination of four arsenic species in rice and water samples by modified graphite electrode-based electrolytic hydride generation coupled with atomic fluorescence spectrometry.

PubMed

Yang, Xin-An; Lu, Xiao-Ping; Liu, Lin; Chi, Miao-Bin; Hu, Hui-Hui; Zhang, Wang-Bing

2016-10-01

This work describes a novel non-chromatographic approach for the accurate and selective determining As species by modified graphite electrode-based electrolytic hydride generation (EHG) for sample introduction coupled with atomic fluorescence spectrometry (AFS) detection. Two kinds of sulfydryl-containing modifiers, l-cysteine (Cys) and glutathione (GSH), are used to modify cathode. The EHG performance of As has been changed greatly at the modified cathode, which has never been reported. Arsenite [As(III)] on the GSH modified graphite electrode (GSH/GE)-based EHG can be selectively and quantitatively converted to AsH3 at applied current of 0.4A. As(III) and arsenate [As(V)] on the Cys modified graphite electrode (Cys/GE) EHG can be selectively and efficiently converted to arsine at applied current of 0.6A, whereas monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) do not form any or only less volatile hydrides under this condition. By changing the analytical conditions, we also have achieved the analysis of total As (tAs) and DMA. Under the optimal condition, the detection limits (3s) of As(III), iAs and tAs in aqueous solutions are 0.25μgL(-1), 0.22μgL(-1) and 0.10μgL(-1), respectively. The accuracy of the method is verified through the analysis of standard reference materials (SRM 1568a). Copyright © 2016 Elsevier B.V. All rights reserved.

Glucosinolate profile and distribution among plant tissues and phenological stages of field-grown horseradish.

PubMed

Agneta, Rosa; Lelario, Filomena; De Maria, Susanna; Möllers, Christian; Bufo, Sabino Aurelio; Rivelli, Anna Rita

2014-10-01

Profile and distribution of glucosinolates (GLS) were detected in plant tissues of horseradish at different developmental stages: beginning of vegetative re-growth, flowering and silique formation. The GLS profile varied widely in the different tissues: we identified 17 GLS in roots and sprouts, one of which was not previously characterized in horseradish, i.e. the 2(S)-hydroxy-2-phenylethyl-GLS (glucobarbarin) and/or 2(R)-hydroxy-2-phenylethyl-GLS (epiglucobarbarin), 11 already found in the roots, including the putative 2-methylsulfonyl-oxo-ethyl-GLS, and 5 previously recognized only in the sprouts. Fifteen of those GLS were also identified in young and cauline leaves, 12 in the mature leaves and 13 in the inflorescences. No difference in GLS profile was observed in plant among the phenological stages. Differences in concentrations of GLS, quantified as desulfated, were found in plant. At the beginning of vegetative re-growth, sprouts while showing the same profile of the roots were much richer in GLS having the highest total GLS concentrations (117.5 and 7.7μmolg(-1) dry weight in sprouts and roots, respectively). During flowering and silique forming stages, the roots still maintained lower amount of total GLS (7.4μmolg(-1) of dry weight, on average) with respect to the epigeous tissues, in which mature and young leaves showed the highest total concentrations (70.5 and 73.8μmolg(-1) of dry weight on average, respectively). Regardless of the phenological stages, the aliphatic GLS were always predominant in all tissues (95%) followed by indolic (2.6%) and benzenic (2.4%) GLS. Sinigrin contributed more than 90% of the total GLS concentration. Aliphatic GLS concentrations were much higher in the epigeous tissues, particularly in the mature and young leaves, while benzenic and indolic GLS concentrations were higher in the roots. Through the phenological stages, GLS concentration increased in young and mature leaves and decreased in cauline leaves and inflorescences, while it remained constant over time in roots. Copyright © 2014 Elsevier Ltd. All rights reserved.

Adsorption and desorption of carbaryl on hexadecyl trimethyl ammonium bromide modified zeolite NaY using RGB portable photometer

NASA Astrophysics Data System (ADS)

Patdhanagul, Nopbhasinthu; Chanpaka, Saiphon; Intharaksa, Orapan; Sirival, Rujikarn; Thanomsith, Kannikar; Wongkwanklom, Sarayuth

2018-04-01

The carbaryl adsorption-desorption isotherms of zeolite NaY and hexadecyl trimethyl ammonium bromide (HTAB) modified zeolite NaY were investigated. Zeolite NaY was synthesized and modified by HTAB in the concentration range 0.1 - 10.0 mM. The adsorption isotherms indicated that zeolite modified with HTAB could significantly enhance the carbaryl adsorption capacity. Zeolite NaY modified with 5.0 mM HTAB gave great carbaryl adsorption because of hydrophilic surface. The 5.0 mM HTAB could adsorb up to 145.75 ppm g-1 of carbaryl which was equivalent to a 36.7% increase. The Surface area characterization showed the remaining of pore volume and pore size diameter and external surface area whereas the BET surface area and micropore surface area of modified zeolite slightly decreased. The XRD results indicate that modification of zeolite NaY with HTAB does not change the crystallinity of the starting zeolite. The elemental analysis indicated that the Si/Al ratio of synthesized zeolite NaY was close to 2.43. Desorption of carbaryl was tested by organic solvents such as methanol, ethanol, tetrahydrofuran, hexane and Deionized water. The results demonstrated that the percentage desorption of methanol is the highest. Carbaryl was quantitatively desorbed with percentage desorption of 82-100 %. It indicated sorption mechanism of carbaryl on the modified sorbent which was principally driven by hydrophobic forces.

Quantitative targeted proteomic analysis of potential markers of tyrosine kinase inhibitor (TKI) sensitivity in EGFR mutated lung adenocarcinoma.

PubMed

Awasthi, Shivangi; Maity, Tapan; Oyler, Benjamin L; Qi, Yue; Zhang, Xu; Goodlett, David R; Guha, Udayan

2018-04-13

Lung cancer causes the highest mortality among all cancers. Patients harboring kinase domain mutations in the epidermal growth factor receptor (EGFR) respond to EGFR tyrosine kinase inhibitors (TKIs), however, acquired resistance always develops. Moreover, 30-40% of patients with EGFR mutations exhibit primary resistance. Hence, there is an unmet need for additional biomarkers of TKI sensitivity that complement EGFR mutation testing and predict treatment response. We previously identified phosphopeptides whose phosphorylation is inhibited upon treatment with EGFR TKIs, erlotinib and afatinib in TKI sensitive cells, but not in resistant cells. These phosphosites are potential biomarkers of TKI sensitivity. Here, we sought to develop modified immuno-multiple reaction monitoring (immuno-MRM)-based quantitation assays for select phosphosites including EGFR-pY1197, pY1172, pY998, AHNAK-pY160, pY715, DAPP1-pY139, CAV1-pY14, INPPL1-pY1135, NEDD9-pY164, NF1-pY2579, and STAT5A-pY694. These sites were significantly hypophosphorylated by erlotinib and a 3rd generation EGFR TKI, osimertinib, in TKI-sensitive H3255 cells, which harbor the TKI-sensitizing EGFR L858R mutation. However, in H1975 cells, which harbor the TKI-resistant EGFR L858R/T790M mutant, osimertinib, but not erlotinib, could significantly inhibit phosphorylation of EGFR-pY-1197, STAT5A-pY694 and CAV1-pY14, suggesting these sites also predict response in TKI-resistant cells. We could further validate EGFR-pY-1197 as a biomarker of TKI sensitivity by developing a calibration curve-based modified immuno-MRM assay. In this report, we have shown the development and optimization of MRM assays coupled with global phosphotyrosine enrichment (modified immuno-MRM) for a list of 11 phosphotyrosine peptides. Our optimized assays identified the targets reproducibly in biological samples with good selectivity. We also developed and characterized quantitation methods to determine endogenous abundance of these targets and correlated the results of the relative quantification with amounts estimated from the calibration curves. This approach represents a way to validate and verify biomarker candidates discovered from large-scale global phospho-proteomics analysis. The application of these modified immuno-MRM assays in lung adenocarcinoma cells provides proof-of concept for the feasibility of clinical applications. These assays may be used in prospective clinical studies of EGFR TKI treatment of EGFR mutant lung cancer to correlate treatment response and other clinical endpoints. Copyright © 2018. Published by Elsevier B.V.

Biodistribution and Pharmacokinetics of EGFR-Targeted Thiolated Gelatin Nanoparticles Following Systemic Administration in Pancreatic Tumor-Bearing Mice

PubMed Central

Xu, Jing; Gattacceca, Florence; Amiji, Mansoor

2013-01-01

The objective of this study was to evaluate qualitative and quantitative biodistribution of epidermal growth factor receptor (EGFR)-targeted thiolated type B gelatin nanoparticles in vivo in a subcutaneous human pancreatic adenocarcinoma (Panc-1) bearing female SCID Beige mice. EGFR-targeted nanoparticles showed preferential and sustained accumulation in the tumor mass, especially at early time points. Higher blood concentrations and higher tumor accumulations were observed with PEG-modified and EGFR-targeted nanoparticles during the study (AUClast: 17.38 and 19.56 %ID/mL*h in blood, 187 and 322 %ID/g*h in tumor for PEG-modified and EGFR-targeted nanoparticles, respectively), as compared to control, unmodified particles (AUClast: 10.71 %ID/mL*h in blood and 138 %ID/g*h in tumor). EGFR-targeted nanoparticles displayed almost twice tumor targeting efficiency than either PEG-modified or the unmodified nanoparticles, highlighting the efficacy of the active targeting strategy. In conclusion, this study shows that EGFR-targeted and PEG-modified nanoparticles were suitable vehicles for specific systemic delivery in subcutaneous Panc-1 tumor xenograft models. PMID:23544877

Biodistribution and pharmacokinetics of EGFR-targeted thiolated gelatin nanoparticles following systemic administration in pancreatic tumor-bearing mice.

PubMed

Xu, Jing; Gattacceca, Florence; Amiji, Mansoor

2013-05-06

The objective of this study was to evaluate qualitative and quantitative biodistribution of epidermal growth factor receptor (EGFR)-targeted thiolated type B gelatin nanoparticles in vivo in subcutaneous human pancreatic adenocarcinoma (Panc-1) bearing female SCID Beige mice. EGFR-targeted nanoparticles showed preferential and sustained accumulation in the tumor mass, especially at early time points. Higher blood concentrations and higher tumor accumulations were observed with PEG-modified and EGFR-targeted nanoparticles during the study (AUClast: 17.38 and 19.56%ID/mL·h in blood, 187 and 322%ID/g·h in tumor for PEG-modified and EGFR-targeted nanoparticles, respectively), as compared to control, unmodified particles (AUClast: 10.71%ID/mL·h in blood and 138%ID/g·h in tumor). EGFR-targeted nanoparticles displayed almost twice tumor targeting efficiency than either PEG-modified or the unmodified nanoparticles, highlighting the efficacy of the active targeting strategy. In conclusion, this study shows that EGFR-targeted and PEG-modified nanoparticles were suitable vehicles for specific systemic delivery in subcutaneous Panc-1 tumor xenograft models.

Screening and prevention of neonatal glucose 6-phosphate dehydrogenase deficiency in Guangzhou, China.

PubMed

Jiang, J; Li, B; Cao, W; Jiang, X; Jia, X; Chen, Q; Wu, J

2014-06-09

We aimed to summarize the results of screening protocol and prevention of neonatal glucose 6-phosphate dehydrogenase (G6PD) deficiency during a 22-year-long period to provide a basis of reference for the screening of this disease. About 1,705,569 newborn subjects in Guangzhou City were screened for this deficiency. Specimens were collected according to the conventional method of specimen acquisition for "newborn dried bloodspot screening", preserved, and inspected. The specimens were studied with fluorescent spot test and quantitative fluorescence assay. Diagnosis was performed using the modified NBTG6PD/6PGD ratio method. Bloodspot filter paper specimens were sent to the laboratory within 24 h via EMS Express, and the G6PD test was performed on the same day. The G6PD deficiency-positive rate was 4.2% in the samples screened using the fluorescent spot test, while it was 5% in case of the quantitative fluorescence assay. Neonatal screening for G6PD deficiency for 11,437 cases (6117 boys and 5320 girls) showed positive results in 481 cases. About 420 cases (318 boys and 102 girls) of G6PD deficiency were confirmed with the modified Duchenne NBT ratio method. The total detection rate was 3.7:5.2% for boys and 1.9% for girls. Quantitative fluorescence assay improved the sensitivity and detection rate. Accelerating the speed of sample delivery by using Internet network systems and ensuring online availability of screening results can aid the screening and diagnosis of this deficiency within 1 week of birth.

Genome Wide Association Mapping in Arabidopsis thaliana Identifies Novel Genes Involved in Linking Allyl Glucosinolate to Altered Biomass and Defense.

PubMed

Francisco, Marta; Joseph, Bindu; Caligagan, Hart; Li, Baohua; Corwin, Jason A; Lin, Catherine; Kerwin, Rachel E; Burow, Meike; Kliebenstein, Daniel J

2016-01-01

A key limitation in modern biology is the ability to rapidly identify genes underlying newly identified complex phenotypes. Genome wide association studies (GWAS) have become an increasingly important approach for dissecting natural variation by associating phenotypes with genotypes at a genome wide level. Recent work is showing that the Arabidopsis thaliana defense metabolite, allyl glucosinolate (GSL), may provide direct feedback regulation, linking defense metabolism outputs to the growth, and defense responses of the plant. However, there is still a need to identify genes that underlie this process. To start developing a deeper understanding of the mechanism(s) that modulate the ability of exogenous allyl GSL to alter growth and defense, we measured changes in plant biomass and defense metabolites in a collection of natural 96 A. thaliana accessions fed with 50 μM of allyl GSL. Exogenous allyl GSL was introduced exclusively to the roots and the compound transported to the leaf leading to a wide range of heritable effects upon plant biomass and endogenous GSL accumulation. Using natural variation we conducted GWAS to identify a number of new genes which potentially control allyl responses in various plant processes. This is one of the first instances in which this approach has been successfully utilized to begin dissecting a novel phenotype to the underlying molecular/polygenic basis.

Production of glucosinolates, phenolic compounds and associated gene expression profiles of hairy root cultures in turnip (Brassica rapa ssp. rapa).

PubMed

Chung, Ill-Min; Rekha, Kaliyaperumal; Rajakumar, Govindasamy; Thiruvengadam, Muthu

2016-12-01

Turnip (Brassica rapa ssp. rapa) is an important vegetable crop producing glucosinolates (GSLs) and phenolic compounds. The GSLs, phenolic compound contents and transcript levels in hairy root cultures, as well as their antioxidant, antimicrobial and anticancer activity were studied in turnip. Transgenic hairy root lines were confirmed by polymerase chain reaction (PCR) and reverse transcription-PCR. GSLs levels (glucoallysin, glucobrassicanapin, gluconasturtiin, glucobrassicin, 4-methoxyglucobrassicin, neoglucobrassicin and 4-hydroxyglucobrassicin) and their gene expression levels (BrMYB28, BrMYB29, BrMYB34, BrMYB51, BrMYB122, CYP79 and CYP83) significantly increased in hairy roots compared with that in non-transformed roots. Furthermore, hairy roots efficiently produced several important individual phenolic compounds (flavonols, hydroxybenzoic and hydroxycinnamic acids). Colorimetric analysis revealed that the highest levels of total phenol, flavonoid contents, and their gene expression levels (PAL, CHI and FLS) in hairy roots than non-transformed roots. Our study provides beneficial information on the molecular and physiological active processes that are associated with the phytochemical content and biosynthetic gene expression in turnip. Moreover, antioxidant activity, as measured by DPPH scavenging activity, reducing potential, phosphomolybdenum and ferrous ion chelating ability assays was significantly higher in hairy roots. Hairy root extracts exhibited higher antimicrobial activity against bacterial and fungal species. The extract of hairy roots showed inhibition of human breast and colon cancer cell lines.

Modification of two capripoxvirus quantitative real-time PCR assays to improve diagnostic sensitivity and include beta-actin as an internal positive control.

PubMed

Das, Amaresh; Deng, Ming Y; Babiuk, Shawn; McIntosh, Michael T

2017-05-01

Capripoxviruses (CaPVs), consisting of Sheeppox virus (SPV), Goatpox virus (GPV), and Lumpy skin disease virus (LSDV) species, cause economically significant diseases in sheep, goats, and cattle, respectively. Quantitative real-time polymerase chain reaction (qPCR) assays are routinely used for rapid detection of CaPVs in surveillance and outbreak management programs. We further modified and optimized 2 previously published CaPV qPCR assays, referred to as the Balinsky and Bowden assays, by changing commercial PCR reagents used in the tests. The modified assays displayed 100% analytical specificity and showed no apparent changes in analytical sensitivities for detection of CaPVs compared with the original assays. Diagnostic sensitivities, assessed using 50 clinical reference samples from experimentally infected sheep, goats, and cattle, improved from 82% to 92% for the modified Balinsky assay and from 58% to 82% for the modified Bowden assay. The modified qPCR assays were multiplexed for detection of beta-actin as an indicator for potential false-negative results. The multiplex modified qPCR assays exhibited the same diagnostic sensitivities as the singleplex assays suggesting their utility in the detection of CaPVs.

Development of a colloidal gold immunochromatographic strip assay for simple and fast detection of human α-lactalbumin in genetically modified cow milk.

PubMed

Tao, Chenyu; Zhang, Qingde; Feng, Na; Shi, Deshi; Liu, Bang

2016-03-01

The qualitative and quantitative declaration of food ingredients is important to consumers, especially for genetically modified food as it experiences a rapid increase in sales. In this study, we designed an accurate and rapid detection system using colloidal gold immunochromatographic strip assay (GICA) methods to detect genetically modified cow milk. First, we prepared 2 monoclonal antibodies for human α-lactalbumin (α-LA) and measured their antibody titers; the one with the higher titer was used for further experiments. Then, we found the optimal pH value and protein amount of GICA for detection of pure milk samples. The developed strips successfully detected genetically modified cow milk and non-modified cow milk. To determine the sensitivity of GICA, a quantitative ELISA system was used to determine the exact amount of α-LA, and then genetically modified milk was diluted at different rates to test the sensitivity of GICA; the sensitivity was 10 μg/mL. Our results demonstrated that the applied method was effective to detect human α-LA in cow milk. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kroupa, Daniel M.; Anderson, Nicholas C.; Castaneda, Chloe V.

Here, we employed quantitative NMR spectroscopy and spectrophotometric absorbance titration to study a quantum dot X-type ligand exchange reaction. We find that the exchange is highly cooperative, where at low extents of exchange the change in free energy of the reaction, Δ G XC, is ~11 kJ mol –1 while at higher extents of exchange Δ G XC saturates to ~–4 kJ mol –1. A modified Fowler binding isotherm is developed to describe the reaction.

Cytotoxicity assessment of modified bioactive glasses with MLO-A5 osteogenic cells in vitro.

PubMed

Modglin, Vernon C; Brown, Roger F; Jung, Steven B; Day, Delbert E

2013-05-01

The primary objective of this study was to evaluate in vitro responses of MLO-A5 osteogenic cells to two modifications of the bioactive glass 13-93. The modified glasses, which were designed for use as cell support scaffolds and contained added boron to form the glasses 13-93 B1 and 13-93 B3, were made to accelerate formation of a bioactive hydroxyapatite surface layer and possibly enhance tissue growth. Quantitative MTT cytotoxicity tests revealed no inhibition of growth of MLO-A5 cells incubated with 13-93 glass extracts up to 10 mg/ml, moderate inhibition of growth with 13-93 B1 glass extracts, and noticeable inhibition of growth with 13-93 B3 glass extracts. A morphology-based biocompatibility test was also performed and yielded qualitative assessments of the relative biocompatibilities of glass extracts that agree with those obtained by the quantitative MTT test. However, as a proof of concept experiment, when MLO-A5 cells were seeded onto 13-93 B3 scaffolds in a dynamic in vitro environment, cell proliferation occurred as evidenced by qualitative and quantitative MTT labeling of scaffolds. Together these results demonstrate the in vitro toxicity of released borate ion in static experiments; however borate ion release can be mitigated in a dynamic environment similar to the human body where microvasculature is present. Here we argue that despite toxicity in static environments, boron-containing 13-93 compositions may warrant further study for use in tissue engineering applications.

Anti-inflammatory activity of horseradish (Armoracia rusticana) root extracts in LPS-stimulated macrophages.

PubMed

Marzocco, Stefania; Calabrone, Luana; Adesso, Simona; Larocca, Marilena; Franceschelli, Silvia; Autore, Giuseppina; Martelli, Giuseppe; Rossano, Rocco

2015-12-01

Horseradish (Armoracia rusticana) is a perennial crop belonging to the Brassicaceae family. Horseradish root is used as a condiment due to its extremely pungent flavour, deriving from the high content of glucosinolates and their breakdown products such as isothiocyanates and other sulfur compounds. Horseradish also has a long history in ethnomedicine. In this study the anti-inflammatory potential of three accessions of Armoracia rusticana on lipopolysaccharide from E. coli treated J774A.1 murine macrophages was evaluated. Our results demonstrate that Armoracia rusticana reduced nitric oxide, tumor necrosis factor-α and interleukin-6 release and nitric oxide synthase and cyclooxygenase-2 expression in macrophages, acting on nuclear transcription factor NF-κB p65 activation. Moreover Armoracia rusticana reduced reactive oxygen species release and increased heme-oxygenase-1 expression, thus contributing to the cytoprotective cellular effect during inflammation.

Effect of shear stress on platelet adhesion to expanded polytetrafluoroethylene, a silicone sheet, and an endothelial cell monolayer.

PubMed

Furukawa, K S; Ushida, T; Sugano, H; Tamaki, T; Ohshima, N; Tateishi, T

2000-01-01

We visualized in real-time platelets adhering to the surface of three representative biomaterials, by using an apparatus consisting of a modified cone and plate rheometer combined with an upright epifluorescence microscope under two shear flows (0.1 and 5.0 dyne/cm2). The materials were expanded polytetrafluoroethylene (ePTFE), silicone sheet, and a monolayer of bovine endothelial cells (ECs) formed on glass, all of which are opaque materials used for artificial blood vessels and medical devices. According to quantitative analysis, the monolayer of ECs formed on glass had better blood compatibility than did either the ePTFE or the silicone sheet under shear flow conditions. Under a shear flow condition of 0.1 dyne/cm2, platelet adhesion was silicone sheet > ePTFE. In contrast, under a shear flow condition of 5.0 dyne/cm2, ePTFE > silicone sheet. These results indicate that the intensity of shear stress could modify the order of hemocompatibility of the materials. Therefore, direct observation of platelet adhesion under shear flow conditions is indispensable for testing and screening biomaterials and for providing a precise quantitative evaluation of platelet adhesion.

Science meets regulation.

PubMed

Bilia, Anna Rita

2014-12-02

The European Pharmacopoeia (Ph. Eur.) is a standard reference for both European and non-European countries and defines requirements for the qualitative and quantitative composition of medicines. Herbal drug (HD) monographs state which aspects have to be considered for quality assurance through the relevant chapters "Definition", "Characters", "Identification", "Tests", and "Assay". Identification of botanical material is achieved by macroscopic and microscopic morphology, generally examined by a trained expert. Content or assay is the most difficult area of quality control to perform, since in most herbal drugs the active constituents are unknown and markers should be used which cannot be really related to the quality. The other critical points are represented by the purity tests, in particular some tests such as heavy metals, aflatoxins and pesticides are laborious and time intensive, requiring a significant investment in equipment, materials, and maintenance. A literature survey concerning alternative and/or complementary tools for quality control of botanicals has been performed by searching the scientific databases Pubmed, SciFinder, Scopus and Web of Science. Diverse analytical methods including DNA fingerprinting, Nuclear Magnetic Resonance (NMR), Near Infra Red (NIR) and (bio)sensors have been reported in the literature to evaluate the quality of botanical products. Identification of plants at the species level can be successfully based on genome-based methods, using DNA barcodes, the nucleotide sequence of a short DNA fragment. NMR can provide direct NMR fingerprint determination (complete assignment of the signals by 1D and 2D experiments), quantitative NMR and chemometric analysis (the metabolite fingerprint is based on the distribution of intensity in the NMR spectrum to provide sample classification). NIR spectroscopy is a fast qualitative and quantitative analytical method, getting knowledge about plant species and/or its geographic origin. Finally, the development of chemical and biological sensors is currently one of the most active areas of analytical research. Immobilization of specific enzymes led to recognize definite class of compounds such as cysteine sulfoxides, glucosinolates, cyanogenic glycosides, and polyphenols. Other recognition elements are nucleic acids to evaluate the ability of different molecules to bind DNA. Sensors have also been developed for the detection of heavy metals in botanicals. Moreover, the analysis of mycotoxins and pesticides, could represent another field of possible application. These alternative/complementary analytical methods represent tools which appear to be an analyst's dream: they are able to give rapid analysis responses; to operate directly on complex matrices, in many cases; to be selective and sensitive enough for the required application; to be portable and sometimes also disposable; and to have fast analysis times. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Quantitative Evaluation of Heavy Duty Machine Tools Remanufacturing Based on Modified Catastrophe Progression Method

NASA Astrophysics Data System (ADS)

shunhe, Li; jianhua, Rao; lin, Gui; weimin, Zhang; degang, Liu

2017-11-01

The result of remanufacturing evaluation is the basis for judging whether the heavy duty machine tool can remanufacture in the EOL stage of the machine tool lifecycle management.The objectivity and accuracy of evaluation is the key to the evaluation method.In this paper, the catastrophe progression method is introduced into the quantitative evaluation of heavy duty machine tools’ remanufacturing,and the results are modified by the comprehensive adjustment method,which makes the evaluation results accord with the standard of human conventional thinking.Using the catastrophe progression method to establish the heavy duty machine tools’ quantitative evaluation model,to evaluate the retired TK6916 type CNC floor milling-boring machine’s remanufacturing.The evaluation process is simple,high quantification,the result is objective.

Asymmetric arginine dimethylation of heterogeneous nuclear ribonucleoprotein K by protein-arginine methyltransferase 1 inhibits its interaction with c-Src.

PubMed

Ostareck-Lederer, Antje; Ostareck, Dirk H; Rucknagel, Karl P; Schierhorn, Angelika; Moritz, Bodo; Huttelmaier, Stefan; Flach, Nadine; Handoko, Lusy; Wahle, Elmar

2006-04-21

Arginine methylation is a post-translational modification found in many RNA-binding proteins. Heterogeneous nuclear ribonucleoprotein K (hnRNP K) from HeLa cells was shown, by mass spectrometry and Edman degradation, to contain asymmetric N(G),N(G)-dimethylarginine at five positions in its amino acid sequence (Arg256, Arg258, Arg268, Arg296, and Arg299). Whereas these five residues were quantitatively modified, Arg303 was asymmetrically dimethylated in <33% of hnRNP K and Arg287 was monomethylated in <10% of the protein. All other arginine residues were unmethylated. Protein-arginine methyltransferase 1 was identified as the only enzyme methylating hnRNP K in vitro and in vivo. An hnRNP K variant in which the five quantitatively modified arginine residues had been substituted was not methylated. Methylation of arginine residues by protein-arginine methyltransferase 1 did not influence the RNA-binding activity, the translation inhibitory function, or the cellular localization of hnRNP K but reduced the interaction of hnRNP K with the tyrosine kinase c-Src. This led to an inhibition of c-Src activation and hnRNP K phosphorylation. These findings support the role of arginine methylation in the regulation of protein-protein interactions.

Locating a modifier gene of Ovum mutant through crosses between DDK and C57BL/6J inbred strains in mice.

PubMed

Tan, Jing; Song, Gen Di; Song, Jia Sheng; Ren, Shi Hao; Li, Chun Li; Zheng, Zhen Yu; Zhao, Wei Dong

2016-06-01

A striking infertile phenotype has been discovered in the DDK strain of mouse. The DDK females are usually infertile when crossed with males of other inbred strains, whereas DDK males exhibit normal fertility in reciprocal crosses. This phenomenon is caused by mutation in the ovum (Om) locus on chromosome 11 and known as the DDK syndrome. Previously, some research groups reported that the embryonic mortality deviated from the semilethal rate in backcrosses between heterozygous (Om/+) females and males of other strains. This embryonic mortality exhibited an aggravated trend with increasing background genes of other strains. These results indicated that some modifier genes of Om were present in other strains. In the present study, a population of N₂2 (Om/+) females from the backcrosses between C57BL/6J (B6) and F₁ (B6♀ × DDK♂) was used to map potential modifier genes of Om. Quantitative trait locus showed that a major locus, namely Amom1 (aggravate modifier gene of Om 1), was located at the middle part of chromosome 9 in mice. The Amom1 could increase the expressivity of Om gene, thereby aggravating embryonic lethality when heterozygous (Om/+) females mated with males of B6 strain. Further, the 1.5 LOD-drop analysis indicated that the confidence interval was between 37.54 and 44.46 cM, ~6.92 cM. Amom1 is the first modifier gene of Om in the B6 background.

Correlation of quinone reductase activity and allyl isothiocyanate formation among different genotypes and grades of horseradish roots.

PubMed

Ku, Kang-Mo; Jeffery, Elizabeth H; Juvik, John A; Kushad, Mosbah M

2015-03-25

Horseradish (Armoracia rusticana) is a perennial crop and its ground root tissue is used in condiments because of the pungency of the glucosinolate (GS)-hydrolysis products allyl isothiocyanate (AITC) and phenethyl isothiocyanate (PEITC) derived from sinigrin and gluconasturtiin, respectively. Horseradish roots are sold in three grades: U.S. Fancy, U.S. No. 1, and U.S. No. 2 according to the USDA standards. These grading standards are primarily based on root diameter and length. There is little information on whether root grades vary in their phytochemical content or potential health promoting properties. This study measured GS, GS-hydrolysis products, potential anticancer activity (as quinone reductase inducing activity), total phenolic content, and antioxidant activities from different grades of horseradish accessions. U.S. Fancy showed significantly higher sinigrin and AITC concentrations than U.S. No. 1 ,whereas U.S. No. 1 showed significantly higher concentrations of 1-cyano 2,3-epithiopropane, the epithionitrile hydrolysis product of sinigrin, and significantly higher total phenolic concentrations than U.S. Fancy.

[Detection of recombinant-DNA in foods from stacked genetically modified plants].

PubMed

Sorokina, E Iu; Chernyshova, O N

2012-01-01

A quantitative real-time multiplex polymerase chain reaction method was applied to the detection and quantification of MON863 and MON810 in stacked genetically modified maize MON 810xMON 863. The limit of detection was approximately 0,1%. The accuracy of the quantification, measured as bias from the accepted value and the relative repeatability standard deviation, which measures the intra-laboratory variability, were within 25% at each GM-level. A method verification has demonstrated that the MON 863 and the MON810 methods can be equally applied in quantification of the respective events in stacked MON810xMON 863.

S-Nitrosothiol measurements in biological systems⋄

PubMed Central

Gow, Andrew; Doctor, Allan; Mannick, Joan; Gaston, Benjamin

2007-01-01

S-Nitrosothiol (SNO) cysteine modifications are regulated signaling reactions that dramatically affect, and are affected by, protein conformation. The lability of the S-NO bond can make SNO-modified proteins cumbersome to measure accurately. Here, we review methodologies for detecting SNO modifications in biology. There are three caveats. 1) Many assays for biological SNOs are used near the limit of detection: standard curves must be in the biologically relevant concentration range. 2) The assays that are most reliable are those that modify SNO protein or peptide chemistry the least. 3) Each result should be quantitatively validated using more than one assay. Improved assays are needed and are in development. PMID:17379583

Quantitative Correlation of in Vivo Properties with in Vitro Assay Results: The in Vitro Binding of a Biotin–DNA Analogue Modifier with Streptavidin Predicts the in Vivo Avidin-Induced Clearability of the Analogue-Modified Antibody

PubMed Central

Dou, Shuping; Virostko, John; Greiner, Dale L.; Powers, Alvin C.; Liu, Guozheng

2016-01-01

Quantitative prediction of in vivo behavior using an in vitro assay would dramatically accelerate pharmaceutical development. However, studies quantitatively correlating in vivo properties with in vitro assay results are rare because of the difficulty in quantitatively understanding the in vivo behavior of an agent. We now demonstrate such a correlation as a case study based on our quantitative understanding of the in vivo chemistry. In an ongoing pretargeting project, we designed a trifunctional antibody (Ab) that concomitantly carried a biotin and a DNA analogue (hereafter termed MORF). The biotin and the MORF were fused into one structure prior to conjugation to the Ab for the concomitant attachment. Because it was known that avidin-bound Ab molecules leave the circulation rapidly, this design would theoretically allow complete clearance by avidin. The clearability of the trifunctional Ab was determined by calculating the blood MORF concentration ratio of avidin-treated Ab to non-avidin-treated Ab using mice injected with these compounds. In theory, any compromised clearability should be due to the presence of impurities. In vitro, we measured the biotinylated percentage of the Ab-reacting (MORF-biotin)⊃-NH2 modifier, by addition of streptavidin to the radiolabeled (MORF-biotin)⊃-NH2 samples and subsequent high-performance liquid chromatography (HPLC) analysis. On the basis of our previous quantitative understanding, we predicted that the clearability of the Ab would be equal to the biotinylation percentage measured via HPLC. We validated this prediction within a 3% difference. In addition to the high avidin-induced clearability of the trifunctional Ab (up to ~95%) achieved by the design, we were able to predict the required quality of the (MORF-biotin)⊃-NH2 modifier for any given in vivo clearability. This approach may greatly reduce the steps and time currently required in pharmaceutical development in the process of synthesis, chemical analysis, in vitro cell study, and in vivo validation. PMID:26103429

Size-exclusive Nanosensor for Quantitative Analysis of Fullerene C60: A Concept Paper

EPA Science Inventory

This paper presents the first development of a mass-sensitive nanosensor for the isolation and quantitative analyses of engineered fullerene (C₆₀) nanoparticles, while excluding mixtures of structurally similar fullerenes. Amino-modified beta cyclodextrin (β-CD-NH

A mass spectrometry-based method for comprehensive quantitative determination of post-transcriptional RNA modifications: the complete chemical structure of Schizosaccharomyces pombe ribosomal RNAs

PubMed Central

Taoka, Masato; Nobe, Yuko; Hori, Masayuki; Takeuchi, Aiko; Masaki, Shunpei; Yamauchi, Yoshio; Nakayama, Hiroshi; Takahashi, Nobuhiro; Isobe, Toshiaki

2015-01-01

We present a liquid chromatography–mass spectrometry (LC-MS)-based method for comprehensive quantitative identification of post-transcriptional modifications (PTMs) of RNA. We incorporated an in vitro-transcribed, heavy isotope-labeled reference RNA into a sample RNA solution, digested the mixture with a number of RNases and detected the post-transcriptionally modified oligonucleotides quantitatively based on shifts in retention time and the MS signal in subsequent LC-MS. This allowed the determination and quantitation of all PTMs in Schizosaccharomyces pombe ribosomal (r)RNAs and generated the first complete PTM maps of eukaryotic rRNAs at single-nucleotide resolution. There were 122 modified sites, most of which appear to locate at the interface of ribosomal subunits where translation takes place. We also identified PTMs at specific locations in rRNAs that were altered in response to growth conditions of yeast cells, suggesting that the cells coordinately regulate the modification levels of RNA. PMID:26013808

Determination of kojic acid based on the interface enhancement effects of carbon nanotube/alizarin red S modified electrode.

PubMed

Liu, Jieshu; Zhou, Dazhai; Liu, Xiaopeng; Wu, Kangbing; Wan, Chidan

2009-04-01

Based on non-covalent interactions such as pi-pi stacking, van der Waals interactions and strong adsorption, alizarin red S (ARS) interacts with multi-walled carbon nanotubes (MWNT), improving the solubility of MWNT in water and resulting in a stable MWNT/ARS solution. By successive cyclic sweeps between 0.0 and 2.2V in the MWNT/ARS solution, a MWNT/ARS composite film was fabricated on an electrode surface. The electrochemical behaviors of kojic acid at the bare electrode, the ARS film-modified electrode and the MWNT/ARS film-modified electrode were investigated. It was found that the oxidation signal of kojic acid significantly increased at the MWNT/ARS film-modified electrode, which was attributed to the unique properties of MWNT such as large surface area, strong adsorptive ability and subtle electronic character. The effects of pH and cyclic number of electropolymerization were examined. A rapid, sensitive and simple electrochemical method was then developed for the determination of kojic acid. This method exhibits good linearity over the range from 4.0 x 10(-7) to 6.0 x 10(-5)mol L(-1), and the limit of detection is as low as 1.0 x 10(-7)mol L(-1). In order to validate feasibility, the MWNT/ARS film-modified electrode was used for quantitative analysis of kojic acid in food samples.

Effect of Structural Heat Conduction on the Performance of Micro-combustors and Micro-thrusters

DTIC Science & Technology

2005-01-01

numerical models lends credence to the principles underlying the modified Mallard- Le Chatelier theory, the quantitative discrepancy between the analytical...employs reasoning similar to that used by Mallard and Le Chatelier to predict the velocity of a freely propagating flame but extends it to include the...equation (2-1). 19 2.2.1. Adiabatic theory The thermal theory of Mallard and Le Chatelier applies to freely propagating flame that does not

Composition and Quantitation of Microalgal Lipids by ERETIC 1H NMR Method

PubMed Central

Nuzzo, Genoveffa; Gallo, Carmela; d’Ippolito, Giuliana; Cutignano, Adele; Sardo, Angela; Fontana, Angelo

2013-01-01

Accurate characterization of biomass constituents is a crucial aspect of research in the biotechnological application of natural products. Here we report an efficient, fast and reproducible method for the identification and quantitation of fatty acids and complex lipids (triacylglycerols, glycolipids, phospholipids) in microalgae under investigation for the development of functional health products (probiotics, food ingredients, drugs, etc.) or third generation biofuels. The procedure consists of extraction of the biological matrix by modified Folch method and direct analysis of the resulting material by proton nuclear magnetic resonance (1H NMR). The protocol uses a reference electronic signal as external standard (ERETIC method) and allows assessment of total lipid content, saturation degree and class distribution in both high throughput screening of algal collection and metabolic analysis during genetic or culturing studies. As proof of concept, the methodology was applied to the analysis of three microalgal species (Thalassiosira weissflogii, Cyclotella cryptica and Nannochloropsis salina) which drastically differ for the qualitative and quantitative composition of their fatty acid-based lipids. PMID:24084790

A new method for rapid determination of indole-3-carbinol and its condensation products in nutraceuticals using core-shell column chromatography method.

PubMed

Fibigr, Jakub; Šatínský, Dalibor; Havlíková, Lucie; Solich, Petr

2016-02-20

Indole-3-carbinol is a natural glucosinolate known for prevention of human breast, prostate and other types of cancer and it started to be used in commercial preparations, as food supplements. However no analytical method has been proposed for quality control of nutraceuticals with this substance yet. In this paper a new high-performance liquid chromatography (HPLC) method using core-shell column for separation of indole-3-carbinol and its condensation/degradation products was developed and used for the quantitative determination of indole-3-carbinol in nutraceuticals. Separation of indole-3-carbinol, its condensation/degradation products and internal standard ethylparaben was performed on the core-shell column Kinetex 5μ XB-C18 100A (100×4.6mm), particle size 5.0μm, with mobile phase acetonitrile/water according to the gradient program at a flow rate of 1.25mLmin(-1) and at temperature 50°C. The detection wavelength was set at 270nm. Under the optimal chromatographic conditions good linearity of determination was achieved. Available commercial samples of nutraceuticals were extracted with 100% methanol using ultrasound bath. A 5-μL sample volume of the supernatant was directly injected into the HPLC system. The developed method provided rapid and accurate tool for quality control of nutraceuticals based on cruciferous vegetable extracts with indole-3-carbinol content. The presented study showed that the declared content of indole-3-carbinol significantly varied in the different nutraceuticals available on the market. Two analyzed preparations showed the presence of condensation/degradation products of indole-3-carbinol which were not officially declared by the manufacturer. Moreover, further two analyzed nutraceutical preparations showed absolutely no content of declared amount of indole-3-carbinol. Copyright © 2015 Elsevier B.V. All rights reserved.

Determination of nickel in water, food, and biological samples by electrothermal atomic absorption spectrometry after preconcentration on modified carbon nanotubes.

PubMed

Taher, Mohammad Ali; Mazaheri, Lida; Ashkenani, Hamid; Mohadesi, Alireza; Afzali, Daryoush

2014-01-01

A new and sensitive SPE method using modified carbon nanotubes for extraction and preconcentration, and electrothermal atomic absorption spectrometric determination of nickel (Ni) in real samples at ng/L levels was investigated. First, multiwalled carbon nanotubes were oxidized with concentrated HNO3, then modified with 2-(5-bormo-2-pyridylazo)-5-diethylaminophenol reagent. The adsorption was achieved quantitatively on a modified carbon nanotubes column in a pH range of 6.5 to 8.5; the adsorbed Ni(II) ions were then desorbed by passing 5.0 mL of 1 M HNO3. The effects of analytical parameters, including pH of the solution, eluent type and volume, sample volume, flow rate of the eluent, and matrix ions, were investigated for optimization of the presented procedure. The enrichment factor was 180, and the LOD for Ni was 4.9 ng/L. The method was applied to the determination of Ni in water, food, and biological samples, and reproducible results were obtained.

Mapping Nuclear Fallout Using the Weather Research & Forecasting (WRF) Model

DTIC Science & Technology

2012-09-01

relevant modules, originally designed to predict the settling of volcanic ash, such that a stabilized cloud of nuclear particulate is initialized...within the model. This modified code is then executed for various atmospheric test explosions and the results are qualitatively and quantitatively...HYSPLIT Simulation ....................................... 44  Figure 7. WRF Fallout Prediction for Test Shot George, 0.8 R/h at H+1

Simultaneous trace multielement determination by ICP-OES after solid phase extraction with modified octadecyl silica gel.

PubMed

Karbasi, Mohamad-Hadi; Jahanparast, Babak; Shamsipur, Mojtaba; Hassan, Jalal

2009-10-15

Multielement simultaneous determination of 35 trace elements in environmental samples was carried out by inductively coupled plasma emission spectrometry (ICP-OES) after preconcentration with octadecyl silicagel, modified with aurin tricarboxylic acid (Aluminon). Optimal experimental conditions including pH of sample solution, sample volume, sample and eluent flow rate, type, concentration and volume of eluent and foreign ions effect were investigated and established. Trace element ions in aqueous solution were quantitatively adsorbed onto octadecyl silicagel modified with aurin tricarboxylic acid at pH 8.0 with a flow rate of 11.0 mL min(-1). The adsorbed element ions were eluted with 3-5 mL of 0.5 mol L(-1) HNO(3) at a flow rate of 10.0 mL min(-1) and analyzed by ICP-OES simultaneously. The proposed method has at least preconcentration factor of 100 in water samples, which results high sensitive detection of ultra-trace and trace analysis. The present methodology gave recoveries better than 70% and RSD less than 16%.

Application of microchip CGE for the analysis of PEG-modified recombinant human granulocyte-colony stimulating factors.

PubMed

Park, Eun Ji; Lee, Kyung Soo; Lee, Kang Choon; Na, Dong Hee

2010-11-01

The purpose of this study was to evaluate the microchip CGE (MCGE) for the analysis of PEG-modified granulocyte-colony stimulating factor (PEG-G-CSF) prepared with PEG-aldehydes. The unmodified and PEG-modified G-CSFs were analyzed by Protein 80 and 230 Labchips on the Agilent 2100 Bioanalyzer. The MCGE allowed size-based separation and quantitation of PEG-G-CSF. The Protein 80 Labchip was useful for PEG-5K-G-CSF, while the Protein 230 Labchip was more suitable for PEG-20K-G-CSF. The MCGE was also used to monitor a search for optimal PEG-modification (PEGylation) conditions to produce mono-PEG-G-CSF. This study demonstrates the usefulness of MCGE for monitoring and optimizing the PEGylation of G-CSF with the advantages of speed, minimal sample consumption, and automatic quantitation.

Genomic Prediction of Testcross Performance in Canola (Brassica napus)

PubMed Central

Jan, Habib U.; Abbadi, Amine; Lücke, Sophie; Nichols, Richard A.; Snowdon, Rod J.

2016-01-01

Genomic selection (GS) is a modern breeding approach where genome-wide single-nucleotide polymorphism (SNP) marker profiles are simultaneously used to estimate performance of untested genotypes. In this study, the potential of genomic selection methods to predict testcross performance for hybrid canola breeding was applied for various agronomic traits based on genome-wide marker profiles. A total of 475 genetically diverse spring-type canola pollinator lines were genotyped at 24,403 single-copy, genome-wide SNP loci. In parallel, the 950 F1 testcross combinations between the pollinators and two representative testers were evaluated for a number of important agronomic traits including seedling emergence, days to flowering, lodging, oil yield and seed yield along with essential seed quality characters including seed oil content and seed glucosinolate content. A ridge-regression best linear unbiased prediction (RR-BLUP) model was applied in combination with 500 cross-validations for each trait to predict testcross performance, both across the whole population as well as within individual subpopulations or clusters, based solely on SNP profiles. Subpopulations were determined using multidimensional scaling and K-means clustering. Genomic prediction accuracy across the whole population was highest for seed oil content (0.81) followed by oil yield (0.75) and lowest for seedling emergence (0.29). For seed yieId, seed glucosinolate, lodging resistance and days to onset of flowering (DTF), prediction accuracies were 0.45, 0.61, 0.39 and 0.56, respectively. Prediction accuracies could be increased for some traits by treating subpopulations separately; a strategy which only led to moderate improvements for some traits with low heritability, like seedling emergence. No useful or consistent increase in accuracy was obtained by inclusion of a population substructure covariate in the model. Testcross performance prediction using genome-wide SNP markers shows considerable potential for pre-selection of promising hybrid combinations prior to resource-intensive field testing over multiple locations and years. PMID:26824924

21 CFR 101.13 - Nutrient content claims-general principles.

Code of Federal Regulations, 2014 CFR

2014-04-01

... must also bear: (A) Clear and concise quantitative information comparing the amount of the subject... is on the information panel, the quantitative information may be located elsewhere on the information...., “Modified fat cheesecake”). This statement of identity must be immediately followed by the comparative...

21 CFR 101.13 - Nutrient content claims-general principles.

Code of Federal Regulations, 2011 CFR

2011-04-01

... must also bear: (A) Clear and concise quantitative information comparing the amount of the subject... is on the information panel, the quantitative information may be located elsewhere on the information...., “Modified fat cheesecake”). This statement of identity must be immediately followed by the comparative...

21 CFR 101.13 - Nutrient content claims-general principles.

Code of Federal Regulations, 2010 CFR

2010-04-01

... must also bear: (A) Clear and concise quantitative information comparing the amount of the subject... is on the information panel, the quantitative information may be located elsewhere on the information...., “Modified fat cheesecake”). This statement of identity must be immediately followed by the comparative...

21 CFR 101.13 - Nutrient content claims-general principles.

Code of Federal Regulations, 2012 CFR

2012-04-01

... must also bear: (A) Clear and concise quantitative information comparing the amount of the subject... is on the information panel, the quantitative information may be located elsewhere on the information...., “Modified fat cheesecake”). This statement of identity must be immediately followed by the comparative...

21 CFR 101.13 - Nutrient content claims-general principles.

Code of Federal Regulations, 2013 CFR

2013-04-01

... must also bear: (A) Clear and concise quantitative information comparing the amount of the subject... is on the information panel, the quantitative information may be located elsewhere on the information...., “Modified fat cheesecake”). This statement of identity must be immediately followed by the comparative...

Simultaneous extraction and preconcentration of uranium and thorium in aqueous samples by new modified mesoporous silica prior to inductively coupled plasma optical emission spectrometry determination.

PubMed

Yousefi, Seyed Reza; Ahmadi, Seyed Javad; Shemirani, Farzaneh; Jamali, Mohammad Reza; Salavati-Niasari, Masoud

2009-11-15

A new synthesized modified mesoporous silica (MCM-41) using 5-nitro-2-furaldehyde (fural) was applied as an effective sorbent for the solid phase extraction of uranium(VI) and thorium(IV) ions from aqueous solution for the measurement by inductively coupled plasma optical emission spectrometry (ICP OES). The influences of some analytical parameters on the quantitative recoveries of the analyte ions were investigated in batch method. Under optimal conditions, the analyte ions were sorbed by the sorbent at pH 5.5 and then eluted with 1.0 mL of 1.0 mol L(-1) HNO(3). The preconcentration factor was 100 for a 100mL sample volume. The limits of detection (LOD) obtained for uranium(VI) and thorium(IV) were 0.3 microg L(-1). The maximum sorption capacity of the modified MCM-41 was found to be 47 and 49 mg g(-1) for uranium(VI) and thorium(IV), respectively. The sorbent exhibited good stability, reusability, high adsorption capacity and fast rate of equilibrium for sorption/desorption of uranium and thorium ions. The applicability of the synthesized sorbent was examined using CRM and real water samples.

Applicability of the chymopapain gene used as endogenous reference gene for transgenic huanong no. 1 papaya detection.

PubMed

Guo, Jinchao; Yang, Litao; Liu, Xin; Zhang, Haibo; Qian, Bingjun; Zhang, Dabing

2009-08-12

The virus-resistant papaya (Carica papaya L.), Huanong no. 1, was the genetically modified (GM) fruit approved for growing in China in 2006. To implement the labeling regulation of GM papaya and its derivates, the development of papaya endogenous reference gene is very necessary for GM papaya detection. Herein, we reported one papaya specific gene, Chymopapain (CHY), as one suitable endogenous reference gene, used for GM papaya identification. Thereafter, we established the conventional and real-time quantitative PCR assays of the CHY gene. In the CHY conventional PCR assay, the limit of detection (LOD) was 25 copies of haploid papaya genome. In the CHY real-time quantitative PCR assay, both the LOD and the limit of quantification (LOQ) were as low as 12.5 copies of haploid papaya genome. Furthermore, we revealed the construct-specific sequence of Chinese GM papaya Huanong no. 1 and developed its conventional and quantitative PCR systems employing the CHY gene as endogenous reference gene. This work is useful for papaya specific identification and GM papaya detection.

Development and application of absolute quantitative detection by duplex chamber-based digital PCR of genetically modified maize events without pretreatment steps.

PubMed

Zhu, Pengyu; Fu, Wei; Wang, Chenguang; Du, Zhixin; Huang, Kunlun; Zhu, Shuifang; Xu, Wentao

2016-04-15

The possibility of the absolute quantitation of GMO events by digital PCR was recently reported. However, most absolute quantitation methods based on the digital PCR required pretreatment steps. Meanwhile, singleplex detection could not meet the demand of the absolute quantitation of GMO events that is based on the ratio of foreign fragments and reference genes. Thus, to promote the absolute quantitative detection of different GMO events by digital PCR, we developed a quantitative detection method based on duplex digital PCR without pretreatment. Moreover, we tested 7 GMO events in our study to evaluate the fitness of our method. The optimized combination of foreign and reference primers, limit of quantitation (LOQ), limit of detection (LOD) and specificity were validated. The results showed that the LOQ of our method for different GMO events was 0.5%, while the LOD is 0.1%. Additionally, we found that duplex digital PCR could achieve the detection results with lower RSD compared with singleplex digital PCR. In summary, the duplex digital PCR detection system is a simple and stable way to achieve the absolute quantitation of different GMO events. Moreover, the LOQ and LOD indicated that this method is suitable for the daily detection and quantitation of GMO events. Copyright © 2016 Elsevier B.V. All rights reserved.

Validated UPLC-MS/MS Methods To Quantitate Free and Conjugated Alternaria Toxins in Commercially Available Tomato Products and Fruit and Vegetable Juices in Belgium.

PubMed

Walravens, Jeroen; Mikula, Hannes; Rychlik, Michael; Asam, Stefan; Devos, Tom; Njumbe Ediage, Emmanuel; Diana Di Mavungu, José; Jacxsens, Liesbeth; Van Landschoot, Anita; Vanhaecke, Lynn; De Saeger, Sarah

2016-06-22

Ultraperformance liquid chromatography tandem mass spectrometry and Quick, Easy, Cheap, Effective, Rugged, and Safe based analytical methodologies to quantitate both free (alternariol (1), alternariol monomethyl ether (2), tenuazonic acid (3), tentoxin (4), altenuene (5), altertoxin-I (6)) and conjugated (sulfates and glucosides of 1 and 2) Alternaria toxins in fruit and vegetable juices and tomato products were developed and validated. Acceptable limits of quantitation (0.7-5.7 μg/kg), repeatability (RSDr < 15.7%), reproducibility (RSDR < 17.9%), and apparent recovery (87.0-110.6%) were obtained for all analytes in all matrices investigated. 129 commercial foodstuffs were analyzed, and 3 was detected in 100% of tomato product samples (

Impact of thermal processing on sulforaphane yield from broccoli ( Brassica oleracea L. ssp. italica).

PubMed

Wang, Grace C; Farnham, Mark; Jeffery, Elizabeth H

2012-07-11

In broccoli, sulforaphane forms when the glucosinolate glucoraphanin is hydrolyzed by the endogenous plant thiohydrolase myrosinase. A myrosinase cofactor directs hydrolysis away from the formation of bioactive sulforaphane and toward an inactive product, sulforaphane nitrile. The cofactor is more heat sensitive than myrosinase, presenting an opportunity to preferentially direct hydrolysis toward sulforaphane formation through regulation of thermal processing. Four broccoli cultivars were microwave heated, boiled, or steamed for various lengths of time. Production of nitrile during hydrolysis of unheated broccoli varied among cultivars from 91 to 52% of hydrolysis products (Pinnacle > Marathon > Patriot > Brigadier). Boiling and microwave heating caused an initial loss of nitrile, with a concomitant increase in sulforaphane, followed by loss of sulforaphane, all within 1 min. In contrast, steaming enhanced sulforaphane yield between 1.0 and 3.0 min in all but Brigadier. These data are proof of concept that steaming for 1.0-3.0 min provides less nitrile and more sulforaphane yield from a broccoli meal.

Long-term sulphur starvation of Arabidopsis thaliana modifies mitochondrial ultrastructure and activity and changes tissue energy and redox status.

PubMed

Ostaszewska, Monika; Juszczuk, Izabela M; Kołodziejek, Izabella; Rychter, Anna M

2014-04-15

Sulphur, as a constituent of amino acids (cysteine and methionine), iron-sulphur clusters, proteins, membrane sulpholipids, glutathione, glucosinolates, coenzymes, and auxin precursors, is essential for plant growth and development. Absence or low sulphur concentration in the soil results in severe growth retardation. Arabidopsis thaliana plants grown hydroponically for nine weeks on Knop nutrient medium without sulphur showed morphological symptoms of sulphur deficiency. The purpose of our study was to investigate changes that mitochondria undergo and the role of the highly branched respiratory chain in survival during sulphur deficiency stress. Ultrastructure analysis of leaf mesophyll cells of sulphur-deficient Arabidopsis showed heterogeneity of mitochondria; some of them were not altered, but the majority had swollen morphology. Dilated mitochondria displayed a lower matrix density and fewer cristae compared to control mitochondria. Disintegration of the inner and outer membranes of some mitochondria from the leaves of sulphur-deficient plants was observed. On the contrary, chloroplast ultrastructure was not affected. Sulphur deficiency changed the respiratory activity of tissues and isolated mitochondria; Complex I and IV capacities and phosphorylation rates were lower, but external NAD(P)H dehydrogenase activity increased. Higher external NAD(P)H dehydrogenase activity corresponded to increased cell redox level with doubled NADH/NAD ratio in the leaf and root tissues. Sulphur deficiency modified energy status in the tissues of Arabidopsis plants. The total concentration of adenylates (expressed as ATP+ADP), measured in the light, was lower in the leaves and roots of sulphur-deficient plants than in the controls, which was mainly due to the severely decreased ATP levels. We show that the changes in mitochondrial ultrastructure are compensated by the modifications in respiratory chain activity. Although mitochondria of Arabidopsis tissues are affected by sulphur deficiency, their metabolic and structural features, which readily reach new homeostasis, make these organelles crucial for adaptation of plants to survive sulphur deficiency. Copyright © 2013 Elsevier GmbH. All rights reserved.

Enantioselective determination of metconazole in multi matrices by high-performance liquid chromatography.

PubMed

He, Rujian; Fan, Jun; Tan, Qi; Lai, Yecai; Chen, Xiaodong; Wang, Tai; Jiang, Ying; Zhang, Yaomou; Zhang, Weiguang

2018-02-01

A reliable and effective HPLC analytical method has been developed to stereoselectively quantify metconazole in soil and flour matrices. Effects of polysaccharide chiral stationary phase, type and content of alcoholic modifier on separation of racemic metconazole have been discussed in detail. Resolution and quantitative determination of metconazole stereoisomers were performed by using an Enantiopak OD column, with the n-hexane-ethanol mixture (97:3, v/v) at the flow rate of 1.0mL/min. Then, extraction and cleanup procedures followed by the modified QuEChERS (quick, easy, cheap, effective, rugged and safe) method were used for metconazole racemate in soil and flour matrices. The residual analysis method was validated. Good linearity (R 2 ≥ 0.9997) and recoveries (94.98-104.89%, RSD ≤ 2.0%) for four metconazole stereoisomers were obtained. In brief, this proposed method showed good accuracy and precision, which might be applied in enantioselective determination, residual quantitative analysis, and degradation of metconazole in food and environmental matrices. Copyright © 2017 Elsevier B.V. All rights reserved.

Effects of root herbivory by nematodes on the performance and preference of a leaf-infesting generalist aphid depend on nitrate fertilization.

PubMed

Kutyniok, Magdalene; Persicke, Marcus; Müller, Caroline

2014-02-01

The performance and behavior of herbivores is strongly affected by the quality of their host plants, which is determined by various environmental conditions. We investigated the performance and preference of the polyphagous shoot-infesting aphid Myzus persicae on the host-plant Arabidopsis thaliana in a two-factorial design in which nitrate fertilization was varied by 33 %, and the root-infesting cyst-nematode Heterodera schachtii was present or absent. Aphid performance was influenced by these abiotic and biotic factors in an interactive way. Nematode presence decreased aphid performance when nitrate levels were low, whereas nematode infestation did not influence aphid performance under higher nitrate fertilization. Aphids followed the "mother knows best" principle when given a choice, settling preferentially on those plants on which they performed best. Hence, they preferred nematode-free over nematode-infested plants in the low fertilization treatment but host choice was not affected by nematodes under higher nitrate fertilization. The amino acid composition of the phloem exudates was significantly influenced by fertilization but also by the interaction of the two treatments. Various glucosinolates in the leaves, which provide an estimate of phloem glucosinolates, were not affected by the individual treatments but by the combination of fertilization and herbivory. These changes in primary and secondary metabolites may be decisive for the herbivore responses. Our data demonstrate that abiotic and biotic factors can interactively affect herbivores, adding a layer of complexity to plant-mediated herbivore interactions.

Crossfit analysis: a novel method to characterize the dynamics of induced plant responses.

PubMed

Jansen, Jeroen J; van Dam, Nicole M; Hoefsloot, Huub C J; Smilde, Age K

2009-12-16

Many plant species show induced responses that protect them against exogenous attacks. These responses involve the production of many different bioactive compounds. Plant species belonging to the Brassicaceae family produce defensive glucosinolates, which may greatly influence their favorable nutritional properties for humans. Each responding compound may have its own dynamic profile and metabolic relationships with other compounds. The chemical background of the induced response is therefore highly complex and may therefore not reveal all the properties of the response in any single model. This study therefore aims to describe the dynamics of the glucosinolate response, measured at three time points after induction in a feral Brassica, by a three-faceted approach, based on Principal Component Analysis. First the large-scale aspects of the response are described in a 'global model' and then each time-point in the experiment is individually described in 'local models' that focus on phenomena that occur at specific moments in time. Although each local model describes the variation among the plants at one time-point as well as possible, the response dynamics are lost. Therefore a novel method called the 'Crossfit' is described that links the local models of different time-points to each other. Each element of the described analysis approach reveals different aspects of the response. The crossfit shows that smaller dynamic changes may occur in the response that are overlooked by global models, as illustrated by the analysis of a metabolic profiling dataset of the same samples.

Crossfit analysis: a novel method to characterize the dynamics of induced plant responses

PubMed Central

2009-01-01

Background Many plant species show induced responses that protect them against exogenous attacks. These responses involve the production of many different bioactive compounds. Plant species belonging to the Brassicaceae family produce defensive glucosinolates, which may greatly influence their favorable nutritional properties for humans. Each responding compound may have its own dynamic profile and metabolic relationships with other compounds. The chemical background of the induced response is therefore highly complex and may therefore not reveal all the properties of the response in any single model. Results This study therefore aims to describe the dynamics of the glucosinolate response, measured at three time points after induction in a feral Brassica, by a three-faceted approach, based on Principal Component Analysis. First the large-scale aspects of the response are described in a 'global model' and then each time-point in the experiment is individually described in 'local models' that focus on phenomena that occur at specific moments in time. Although each local model describes the variation among the plants at one time-point as well as possible, the response dynamics are lost. Therefore a novel method called the 'Crossfit' is described that links the local models of different time-points to each other. Conclusions Each element of the described analysis approach reveals different aspects of the response. The crossfit shows that smaller dynamic changes may occur in the response that are overlooked by global models, as illustrated by the analysis of a metabolic profiling dataset of the same samples. PMID:20015363

Ultraviolet-B radiation modifies the quantitative and qualitative profile of flavonoids and amino acids in grape berries.

PubMed

Martínez-Lüscher, J; Torres, N; Hilbert, G; Richard, T; Sánchez-Díaz, M; Delrot, S; Aguirreolea, J; Pascual, I; Gomès, E

2014-06-01

Grapevine cv. Tempranillo fruit-bearing cuttings were exposed to supplemental ultraviolet-B (UV-B) radiation under controlled conditions, in order to study its effect on grape traits, ripening, amino acids and flavonoid profile. The plants were exposed to two doses of UV-B biologically effective (5.98 and 9.66kJm(-2)d(-1)), applied either from fruit set to ripeness or from the onset of veraison to ripeness. A 0kJm(-2)d(-1) treatment was included as a control. UV-B did not significantly modify grape berry size, but increased the relative mass of berry skin. Time to reach ripeness was not affected by UV-B, which may explain the lack of changes in technological maturity. The concentration of must extractable anthocyanins, colour density and skin flavonols were enhanced by UV-B, especially in plants exposed from fruit set. The quantitative and qualitative profile of grape skin flavonols were modified by UV-B radiation. Monosubstituted flavonols relative abundance increased proportionally to the accumulated UV-B doses. Furthermore, trisubstituted forms, which where predominant in non-exposed berries, were less abundant as UV-B exposure increased. Although total free amino acid content remained unaffected by the treatments, the increased levels of gamma-aminobutyric acid (GABA), as well as the decrease in threonine, isoleucine, methionine, serine and glycine, revealed a potential influence of UV-B on the GABA-mediated signalling and amino acid metabolism. UV-B had an overall positive impact on grape berry composition. Copyright © 2014 Elsevier Ltd. All rights reserved.

Lateral flow nucleic acid biosensor for sensitive detection of microRNAs based on the dual amplification strategy of duplex-specific nuclease and hybridization chain reaction.

PubMed

Ying, Na; Ju, Chuanjing; Sun, Xiuwei; Li, Letian; Chang, Hongbiao; Song, Guangping; Li, Zhongyi; Wan, Jiayu; Dai, Enyong

2017-01-01

MicroRNAs (miRNAs) constitute novel biomarkers for various diseases. Accurate and quantitative analysis of miRNA expression is critical for biomedical research and clinical theranostics. In this study, a method was developed for sensitive and specific detection of miRNAs via dual signal amplification based on duplex specific nuclease (DSN) and hybridization chain reaction (HCR). A reporter probe (RP), comprising recognition sequence (3' end modified with biotin) for a target miRNA of miR-21 and capture sequence (5' end modified with Fam) for HCR product, was designed and synthesized. HCR was initiated by partial sequence of initiator probe (IP), the other part of which can hybridize with capture sequence of RP, and was assembled by hairpin probes modified with biotin (H1-bio and H2-bio). A miR-21 triggered cyclical DSN cleavage of RP, which was immobilized to a streptavidin (SA) coated magnetic bead (MB). The released Fam labeled capture sequence then hybridized with the HCR product to generate a detectable dsDNA. This polymer was then dropped on lateral flow strip and positive result was observed. The proposed method allowed quantitative sequence-specific detection of miR-21 (with a detection limit of 2.1 fM, S/N = 3) in a dynamic range from 100 fM to 100 pM, with an excellent ability to discriminate differences in miRNAs. The method showed acceptable testing recoveries for the determination of miRNAs in serum.

Adaptable Detection Strategies in Membrane-Based Immunoassays: Calibration-Free Quantitation with Surface-Enhanced Raman Scattering Readout.

PubMed

Skuratovsky, Aleksander; Soto, Robert J; Porter, Marc D

2018-06-19

This paper presents a method for immunometric biomarker quantitation that uses standard flow-through assay reagents and obviates the need for constructing a calibration curve. The approach relies on a nitrocellulose immunoassay substrate with multiple physical addresses for analyte capture, each modified with different amounts of an analyte-specific capture antibody. As such, each address generates a distinctly different readout signal that is proportional to the analyte concentration in the sample. To establish the feasibility of this concept, equations derived from antibody-antigen binding equilibrium were first applied in modeling experiments. Next, nitrocellulose membranes with multiple capture antibody addresses were fabricated for detection of a model analyte, human Immunoglobulin G (hIgG), by a heterogeneous sandwich immunoassay using antibody-modified gold nanoparticles (AuNPs) as the immunolabel. Counting the number of colored capture addresses visible to the unassisted eye enabled semiquantitative hIgG determination. We then demonstrated that, by leveraging the localized surface plasmon resonance of the AuNPs, surface-enhanced Raman spectroscopy (SERS) can be used for quantitative readout. By comparing the SERS signal intensities from each capture address with values predicted using immunoassay equilibrium theory, the concentration of hIgG can be determined (∼30% average absolute deviation) without reference to a calibration curve. This work also demonstrates the ability to manipulate the dynamic range of the assay over ∼4 orders of magnitude (from 2 ng mL -1 to 10 μg mL -1 ). The potential prospects in applying this concept to point-of-need diagnostics are also discussed.

The Role of Introductory Geosciences in Students' Quantitative Literacy

NASA Astrophysics Data System (ADS)

Wenner, J. M.; Manduca, C.; Baer, E. M.

2006-12-01

Quantitative literacy is more than mathematics; it is about reasoning with data. Colleges and universities have begun to recognize the distinction between mathematics and quantitative literacy, modifying curricula to reflect the need for numerate citizens. Although students may view geology as 'rocks for jocks', the geosciences are truthfully rife with data, making introductory geoscience topics excellent context for developing the quantitative literacy of students with diverse backgrounds. In addition, many news items that deal with quantitative skills, such as the global warming phenomenon, have their basis in the Earth sciences and can serve as timely examples of the importance of quantitative literacy for all students in introductory geology classrooms. Participants at a workshop held in 2006, 'Infusing Quantitative Literacy into Introductory Geoscience Courses,' discussed and explored the challenges and opportunities associated with the inclusion of quantitative material and brainstormed about effective practices for imparting quantitative literacy to students with diverse backgrounds. The tangible results of this workshop add to the growing collection of quantitative materials available through the DLESE- and NSF-supported Teaching Quantitative Skills in the Geosciences website, housed at SERC. There, faculty can find a collection of pages devoted to the successful incorporation of quantitative literacy in introductory geoscience. The resources on the website are designed to help faculty to increase their comfort with presenting quantitative ideas to students with diverse mathematical abilities. A methods section on "Teaching Quantitative Literacy" (http://serc.carleton.edu/quantskills/methods/quantlit/index.html) focuses on connecting quantitative concepts with geoscience context and provides tips, trouble-shooting advice and examples of quantitative activities. The goal in this section is to provide faculty with material that can be readily incorporated into existing introductory geoscience courses. In addition, participants at the workshop (http://serc.carleton.edu/quantskills/workshop06/index.html) submitted and modified more than 20 activities and model courses (with syllabi) designed to use best practices for helping introductory geoscience students to become quantitatively literate. We present insights from the workshop and other sources for a framework that can aid in increasing quantitative literacy of students from a variety of backgrounds in the introductory geoscience classroom.

Quantitative Peptidomics with Five-plex Reductive Methylation labels

NASA Astrophysics Data System (ADS)

Tashima, Alexandre K.; Fricker, Lloyd D.

2017-12-01

Quantitative peptidomics and proteomics often use chemical tags to covalently modify peptides with reagents that differ in the number of stable isotopes, allowing for quantitation of the relative peptide levels in the original sample based on the peak height of each isotopic form. Different chemical reagents have been used as tags for quantitative peptidomics and proteomics, and all have strengths and weaknesses. One of the simplest approaches uses formaldehyde and sodium cyanoborohydride to methylate amines, converting primary and secondary amines into tertiary amines. Up to five different isotopic forms can be generated, depending on the isotopic forms of formaldehyde and cyanoborohydride reagents, allowing for five-plex quantitation. However, the mass difference between each of these forms is only 1 Da per methyl group incorporated into the peptide, and for many peptides there is substantial overlap from the natural abundance of 13C and other isotopes. In this study, we calculated the contribution from the natural isotopes for 26 native peptides and derived equations to correct the peak intensities. These equations were applied to data from a study using human embryonic kidney HEK293T cells in which five replicates were treated with 100 nM vinblastine for 3 h and compared with five replicates of cells treated with control medium. The correction equations brought the replicates to the expected 1:1 ratios and revealed significant decreases in levels of 21 peptides upon vinblastine treatment. These equations enable accurate quantitation of small changes in peptide levels using the reductive methylation labeling approach. [Figure not available: see fulltext.

Quantitative Peptidomics with Five-plex Reductive Methylation labels

NASA Astrophysics Data System (ADS)

Tashima, Alexandre K.; Fricker, Lloyd D.

2018-05-01

Quantitative peptidomics and proteomics often use chemical tags to covalently modify peptides with reagents that differ in the number of stable isotopes, allowing for quantitation of the relative peptide levels in the original sample based on the peak height of each isotopic form. Different chemical reagents have been used as tags for quantitative peptidomics and proteomics, and all have strengths and weaknesses. One of the simplest approaches uses formaldehyde and sodium cyanoborohydride to methylate amines, converting primary and secondary amines into tertiary amines. Up to five different isotopic forms can be generated, depending on the isotopic forms of formaldehyde and cyanoborohydride reagents, allowing for five-plex quantitation. However, the mass difference between each of these forms is only 1 Da per methyl group incorporated into the peptide, and for many peptides there is substantial overlap from the natural abundance of 13C and other isotopes. In this study, we calculated the contribution from the natural isotopes for 26 native peptides and derived equations to correct the peak intensities. These equations were applied to data from a study using human embryonic kidney HEK293T cells in which five replicates were treated with 100 nM vinblastine for 3 h and compared with five replicates of cells treated with control medium. The correction equations brought the replicates to the expected 1:1 ratios and revealed significant decreases in levels of 21 peptides upon vinblastine treatment. These equations enable accurate quantitation of small changes in peptide levels using the reductive methylation labeling approach. [Figure not available: see fulltext.

Development and interlaboratory validation of quantitative polymerase chain reaction method for screening analysis of genetically modified soybeans.

PubMed

Takabatake, Reona; Onishi, Mari; Koiwa, Tomohiro; Futo, Satoshi; Minegishi, Yasutaka; Akiyama, Hiroshi; Teshima, Reiko; Kurashima, Takeyo; Mano, Junichi; Furui, Satoshi; Kitta, Kazumi

2013-01-01

A novel real-time polymerase chain reaction (PCR)-based quantitative screening method was developed for three genetically modified soybeans: RRS, A2704-12, and MON89788. The 35S promoter (P35S) of cauliflower mosaic virus is introduced into RRS and A2704-12 but not MON89788. We then designed a screening method comprised of the combination of the quantification of P35S and the event-specific quantification of MON89788. The conversion factor (Cf) required to convert the amount of a genetically modified organism (GMO) from a copy number ratio to a weight ratio was determined experimentally. The trueness and precision were evaluated as the bias and reproducibility of relative standard deviation (RSDR), respectively. The determined RSDR values for the method were less than 25% for both targets. We consider that the developed method would be suitable for the simple detection and approximate quantification of GMO.

Rapid Quantitative Detection of Brucella melitensis by a Label-Free Impedance Immunosensor Based on a Gold Nanoparticle-Modified Screen-Printed Carbon Electrode

PubMed Central

Wu, Haiyun; Zuo, Yueming; Cui, Chuanjin; Yang, Wei; Ma, Haili; Wang, Xiaowen

2013-01-01

A rapid and simple method for quantitative monitoring of Brucella melitensis using electrochemical impedance spectroscopy (EIS) is reported for the first time. The label-free immunosensors were fabricated by immobilizing Brucella melitensis antibody on the surface of gold nanoparticle-modified screen-printed carbon electrodes (GNP-SPCEs). Cyclic voltammetry (CV) and EIS were used to characterize the Brucella melitensis antigen interaction on the surface of GNP-SPCEs with antibody. A general electronic equivalent model of an electrochemical cell was introduced for interpretation of the impedance components of the system. The results showed that the change in electron-transfer resistance (Rct) was significantly different due to the binding of Brucella melitensis cells. A linear relationship between the Rct variation and logarithmic value of the cell concentration was found from 4 × 104 to 4 × 106 CFU/mL in pure culture. The label-free impedance biosensor was able to detect as low as 1 × 104 and 4 × 105 CFU/mL of Brucella melitensis in pure culture and milk samples, respectively, in less than 1.5 h. Moreover, a good selectivity versus Escherichia coli O157:H7 and Staphylococcus aureus cells was obtained for our developed immunosensor demonstrating its specificity towards only Brucella melitensis. PMID:23881126

Construction of measurement uncertainty profiles for quantitative analysis of genetically modified organisms based on interlaboratory validation data.

PubMed

Macarthur, Roy; Feinberg, Max; Bertheau, Yves

2010-01-01

A method is presented for estimating the size of uncertainty associated with the measurement of products derived from genetically modified organisms (GMOs). The method is based on the uncertainty profile, which is an extension, for the estimation of uncertainty, of a recent graphical statistical tool called an accuracy profile that was developed for the validation of quantitative analytical methods. The application of uncertainty profiles as an aid to decision making and assessment of fitness for purpose is also presented. Results of the measurement of the quantity of GMOs in flour by PCR-based methods collected through a number of interlaboratory studies followed the log-normal distribution. Uncertainty profiles built using the results generally give an expected range for measurement results of 50-200% of reference concentrations for materials that contain at least 1% GMO. This range is consistent with European Network of GM Laboratories and the European Union (EU) Community Reference Laboratory validation criteria and can be used as a fitness for purpose criterion for measurement methods. The effect on the enforcement of EU labeling regulations is that, in general, an individual analytical result needs to be < 0.45% to demonstrate compliance, and > 1.8% to demonstrate noncompliance with a labeling threshold of 0.9%.

Quantitative Förster resonance energy transfer analysis for kinetic determinations of SUMO-specific protease.

PubMed

Liu, Yan; Song, Yang; Madahar, Vipul; Liao, Jiayu

2012-03-01

Förster resonance energy transfer (FRET) technology has been widely used in biological and biomedical research, and it is a very powerful tool for elucidating protein interactions in either dynamic or steady state. SUMOylation (the process of SUMO [small ubiquitin-like modifier] conjugation to substrates) is an important posttranslational protein modification with critical roles in multiple biological processes. Conjugating SUMO to substrates requires an enzymatic cascade. Sentrin/SUMO-specific proteases (SENPs) act as an endopeptidase to process the pre-SUMO or as an isopeptidase to deconjugate SUMO from its substrate. To fully understand the roles of SENPs in the SUMOylation cycle, it is critical to understand their kinetics. Here, we report a novel development of a quantitative FRET-based protease assay for SENP1 kinetic parameter determination. The assay is based on the quantitative analysis of the FRET signal from the total fluorescent signal at acceptor emission wavelength, which consists of three components: donor (CyPet-SUMO1) emission, acceptor (YPet) emission, and FRET signal during the digestion process. Subsequently, we developed novel theoretical and experimental procedures to determine the kinetic parameters, k(cat), K(M), and catalytic efficiency (k(cat)/K(M)) of catalytic domain SENP1 toward pre-SUMO1. Importantly, the general principles of this quantitative FRET-based protease kinetic determination can be applied to other proteases. Copyright © 2011 Elsevier Inc. All rights reserved.

Stable, water extractable isothiocyanates from Moringa oleifera leaves attenuate inflammation in vitro

PubMed Central

Waterman, Carrie; Cheng, Diana M.; Rojas-Silva, Patricio; Poulev, Alexander; Dreifus, Julia; Ann Lila, Mary; Raskin, Ilya

2014-01-01

Moringa (Moringa oleifera Lam.) is an edible plant used as food and medicine throughout the tropics. A moringa concentrate (MC) made by extracting fresh leaves with water utilized naturally occurring myrosinase to convert four moringa glucosinolates (1–4) into moringa isothiocyanates (5–8). Optimum conditions maximizing MC yield, compound 5 (4-[(α-L-rhamnosyloxy)benzyl]isothiocyanate), and compound 8 (4-[(4’-O-acetyl-α-L-rhamnosyloxy)benzyl]isothiocyanate) content were established (1:5 fresh leaf weight to water ratio at room temperature). The optimized MC contained 1.66% isothiocyanates and 3.82% total polyphenols. Compound 8 exhibited 80% stability at 37 °C for 30 days. MC, 5, and 8 significantly decreased gene expression and production of inflammatory markers in RAW macrophages. Specifically, 5 and 8 attenuated expression of iNOS and IL-1β and production of nitric oxide and TNFβ at 1 and 5 µM. Our results suggest a potential for stable and concentrated moringa isothiocyanates (5–8), delivered in MC as a food-grade product, to alleviate low-grade inflammation associated with chronic diseases. PMID:24731259

Quantitative 13C NMR characterization of fast pyrolysis oils

DOE PAGES

Happs, Renee M.; Lisa, Kristina; Ferrell, III, Jack R.

2016-10-20

Quantitative 13C NMR analysis of model catalytic fast pyrolysis (CFP) oils following literature procedures showed poor agreement for aromatic hydrocarbons between NMR measured concentrations and actual composition. Furthermore, modifying integration regions based on DEPT analysis for aromatic carbons resulted in better agreement. Solvent effects were also investigated for hydrotreated CFP oil.

Quantitative 13C NMR characterization of fast pyrolysis oils

DOE Office of Scientific and Technical Information (OSTI.GOV)

Happs, Renee M.; Lisa, Kristina; Ferrell, III, Jack R.

Quantitative 13C NMR analysis of model catalytic fast pyrolysis (CFP) oils following literature procedures showed poor agreement for aromatic hydrocarbons between NMR measured concentrations and actual composition. Furthermore, modifying integration regions based on DEPT analysis for aromatic carbons resulted in better agreement. Solvent effects were also investigated for hydrotreated CFP oil.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Meisner, Ludmila, E-mail: llm@ispms.tsc.ru; Meisner, Stanislav, E-mail: msn@ispms.tsc.ru; Mironov, Yurii, E-mail: myp@ispms.tsc.ru

The paper considers the effects arising on X-ray diffraction patterns taken in different diffraction geometries and how these effects can be interpreted to judge structural states in NiTi near-surface regions after electron and ion beam treatment. It is shown that qualitative and quantitative analysis of phase composition, lattice parameters of main phases, elastic stress states, and their in-depth variation requires X-ray diffraction patterns in both symmetric Bragg–Brentano and asymmetric Lambot–Vassamilleta geometries with variation in X-ray wavelengths and imaging conditions (with and with no β-filter). These techniques of structural phase analysis are more efficient when the thickness of modified NiTi surfacemore » layers is 1–10 μm (after electron beam treatment) and requires special imaging conditions when the thickness of modified NiTi surface layers is no greater than 1 μm (after ion beam treatment)« less

A Sensitive Branched DNA HIV-1 Signal Amplification Viral Load Assay with Single Day Turnaround

PubMed Central

Baumeister, Mark A.; Zhang, Nan; Beas, Hilda; Brooks, Jesse R.; Canchola, Jesse A.; Cosenza, Carlo; Kleshik, Felix; Rampersad, Vinod; Surtihadi, Johan; Battersby, Thomas R.

2012-01-01

Branched DNA (bDNA) is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA) (“Versant Assay”) currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16–18 h to 2.5 h, composition of only the “Lysis Diluent” solution was modified. Nucleic acid probes in the assay were unchanged. Performance of the modified assay (assay in development; not commercially available) was evaluated and compared to the Versant Assay. Dilution series replicates (>950 results) were used to demonstrate that analytical sensitivity, linearity, accuracy, and precision for the shorter modified assay are comparable to the Versant Assay. HIV RNA-positive clinical specimens (n = 135) showed no significant difference in quantification between the modified assay and the Versant Assay. Equivalent relative quantification of samples of eight genotypes was demonstrated for the two assays. Elevated levels of several potentially interfering endogenous substances had no effect on quantification or specificity of the modified assay. The modified assay with drastically improved turnaround time demonstrates the viability of signal-amplifying technology, such as bDNA, as an alternative to the PCR-based assays dominating viral load monitoring in clinical laboratories. Highly sensitive bDNA assays with a single day turnaround may be ideal for laboratories with especially stringent cost, contamination, or reliability requirements. PMID:22479381

A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

PubMed

Baumeister, Mark A; Zhang, Nan; Beas, Hilda; Brooks, Jesse R; Canchola, Jesse A; Cosenza, Carlo; Kleshik, Felix; Rampersad, Vinod; Surtihadi, Johan; Battersby, Thomas R

2012-01-01

Branched DNA (bDNA) is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA) ("Versant Assay") currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16-18 h to 2.5 h, composition of only the "Lysis Diluent" solution was modified. Nucleic acid probes in the assay were unchanged. Performance of the modified assay (assay in development; not commercially available) was evaluated and compared to the Versant Assay. Dilution series replicates (>950 results) were used to demonstrate that analytical sensitivity, linearity, accuracy, and precision for the shorter modified assay are comparable to the Versant Assay. HIV RNA-positive clinical specimens (n = 135) showed no significant difference in quantification between the modified assay and the Versant Assay. Equivalent relative quantification of samples of eight genotypes was demonstrated for the two assays. Elevated levels of several potentially interfering endogenous substances had no effect on quantification or specificity of the modified assay. The modified assay with drastically improved turnaround time demonstrates the viability of signal-amplifying technology, such as bDNA, as an alternative to the PCR-based assays dominating viral load monitoring in clinical laboratories. Highly sensitive bDNA assays with a single day turnaround may be ideal for laboratories with especially stringent cost, contamination, or reliability requirements.

Functionalized mesoporous silica materials for molsidomine adsorption: Thermodynamic study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alyoshina, Nonna A.; Parfenyuk, Elena V., E-mail: evp@iscras.ru

2013-09-15

A series of unmodified and organically modified mesoporous silica materials was prepared. The unmodified mesoporous silica was synthesized via sol–gel synthesis in the presence of D-glucose as pore-forming agent. The functionalized by phenyl, aminopropyl and mercaptopropyl groups silica materials were prepared via grafting. The fabricated adsorbent materials were characterized by Fourier transform infrared spectroscopy (FTIR) analysis, N{sub 2} adsorption/desorption and elemental analysis methods. Then their adsorption properties for mesoionic dug molsidomine were investigated at 290–313 K and physiological pH value. Thermodynamic parameters of molsidomine adsorption on the synthesized materials have been calculated. The obtained results showed that the adsorption processmore » of molsidomine on the phenyl modified silica is the most quantitatively and energetically favorable. The unmodified and mercaptopropyl modified silica materials exhibit significantly higher adsorption capacities and energies for molsidomine than the aminopropyl modified sample. The effects are discussed from the viewpoint of nature of specific interactions responsible for the adsorption. - Graphical abstract: Comparative analysis of the thermodynamic characteristics of molsidomine adsorption showed that the adsorption process on mesoporous silica materials is controlled by chemical nature of surface functional groups. Molsidomine adsorption on the phenyl modified silica is the most quantitatively and energetically favorable. Taking into account ambiguous nature of mesoionic compounds, it was found that molsidomine is rather aromatic than dipolar. Display Omitted - Highlights: • Unmodified and organically modified mesoporous silica materials were prepared. • Molsidomine adsorption on the silica materials was studied. • Phenyl modified silica shows the highest adsorption capacity and favorable energy. • Molsidomine exhibits the lowest affinity to aminopropyl modified silica.« less

Association between floods and infectious diarrhea and their effect modifiers in Hunan province, China: A two-stage model.

PubMed

Liu, Zhidong; Zhang, Feifei; Zhang, Ying; Li, Jing; Liu, Xuena; Ding, Guoyong; Zhang, Caixia; Liu, Qiyong; Jiang, Baofa

2018-06-01

Understanding the potential links between floods and infectious diarrhea is important under the context of climate change. However, little is known about the risk of infectious diarrhea after floods and what factors could modify these effects in China. This study aims to quantitatively examine the relationship between floods and infectious diarrhea and their effect modifiers. Weekly number of infectious diarrhea cases from 2004 to 2011 during flood season in Hunan province were supplied by the National Notifiable Disease Surveillance System. Flood and meteorological data over the same period were obtained. A two-stage model was used to estimate a provincial average association and their effect modifiers between floods and infectious diarrhea, accounting for other confounders. A total of 134,571 cases of infectious diarrhea were notified from 2004 to 2011. After controlling for seasonality, long-term trends, and meteorological factors, floods were significantly associated with infectious diarrhea in the provincial level with a cumulative RR of 1.22 (95% CI: 1.05, 1.43) with a lagged effect of 0-1 week. Geographic locations and economic levels were identified as effect modifiers, with a higher impact of floods on infectious diarrhea in the western and regions with a low economic level of Hunan. Our study provides strong evidence of a positive association between floods and infectious diarrhea in the study area. Local control strategies for public health should be taken in time to prevent and reduce the risk of infectious diarrhea after floods, especially for the vulnerable regions identified. Copyright © 2018 Elsevier B.V. All rights reserved.

LC-MS/MS Based Quantitation of ABC and SLC Transporter Proteins in Plasma Membranes of Cultured Primary Human Retinal Pigment Epithelium Cells and Immortalized ARPE19 Cell Line.

PubMed

Pelkonen, Laura; Sato, Kazuki; Reinisalo, Mika; Kidron, Heidi; Tachikawa, Masanori; Watanabe, Michitoshi; Uchida, Yasuo; Urtti, Arto; Terasaki, Tetsuya

2017-03-06

The retinal pigment epithelium (RPE) forms the outer blood-retinal barrier between neural retina and choroid. The RPE has several important vision supporting functions, such as transport mechanisms that may also modify pharmacokinetics in the posterior eye segment. Expression of plasma membrane transporters in the RPE cells has not been quantitated. The aim of this study was to characterize and compare transporter protein expression in the ARPE19 cell line and hfRPE (human fetal RPE) cells by using quantitative targeted absolute proteomics (QTAP). Among 41 studied transporters, 16 proteins were expressed in hfRPE and 13 in ARPE19 cells. MRP1, MRP5, GLUT1, 4F2hc, TAUT, CAT1, LAT1, and MATE1 proteins were detected in both cell lines within 4-fold differences. MPR7, OAT2 and RFC1 were detected in the hfRPE cells, but their expression levels were below the limit of quantification in ARPE19 cells. PCFT was detected in both studied cell lines, but the expression was over 4-fold higher in hfRPE cells. MCT1, MCT4, MRP4, and Na + /K + ATPase were upregulated in the ARPE19 cell line showing over 4-fold differences in the quantitative expression values. Expression levels of 25 transporters were below the limit of quantification in both cell models. In conclusion, we present the first systematic and quantitative study on transporter protein expression in the plasma membranes of ARPE19 and hfRPE cells. Overall, transporter expression in the ARPE19 and hfRPE cells correlated well and the absolute expression levels were similar, but not identical. The presented quantitative expression levels could be a useful basis for further studies on drug permeation in the outer blood-retinal barrier.

Compositional and proteomic analyses of genetically modified broccoli (Brassica oleracea var. italica) harboring an agrobacterial gene.

PubMed

Liu, Mao-Sen; Ko, Miau-Hwa; Li, Hui-Chun; Tsai, Shwu-Jene; Lai, Ying-Mi; Chang, You-Ming; Wu, Min-Tze; Chen, Long-Fang O

2014-08-28

Previously, we showed improved shelf life for agrobacterial isopentenyltransferase (ipt) transgenic broccoli (Brassica oleracea var. italica), with yield comparable to commercial varieties, because of the protection mechanism offered by molecular chaperones and stress-related proteins. Here, we used proximate analysis to examine macronutrients, chemical and mineral constituents as well as anti-nutrient and protein changes of ipt-transgenic broccoli and corresponding controls. We also preliminarily assessed safety in mice. Most aspects were comparable between ipt-transgenic broccoli and controls, except for a significant increase in carbohydrate level and a decrease in magnesium content in ipt-transgenic lines 101, 102 and 103, as compared with non-transgenic controls. In addition, the anti-nutrient glucosinolate content was increased and crude fat content decreased in inbred control 104 and transgenic lines as compared with the parental control, "Green King". Gel-based proteomics detected more than 50 protein spots specifically found in ipt-transgenic broccoli at harvest and after cooking; one-third of these proteins showed homology to potential allergens that also play an important role in plant defense against stresses and senescence. Mice fed levels of ipt-transgenic broccoli mimicking the 120 g/day of broccoli eaten by a 60-kg human adult showed normal growth and immune function. In conclusion, the compositional and proteomic changes attributed to the transgenic ipt gene did not affect the growth and immune response of mice under the feeding regimes examined.

Compositional and Proteomic Analyses of Genetically Modified Broccoli (Brassica oleracea var. italica) Harboring an Agrobacterial Gene

PubMed Central

Liu, Mao-Sen; Ko, Miau-Hwa; Li, Hui-Chun; Tsai, Shwu-Jene; Lai, Ying-Mi; Chang, You-Ming; Wu, Min-Tze; Chen, Long-Fang O.

2014-01-01

Previously, we showed improved shelf life for agrobacterial isopentenyltransferase (ipt) transgenic broccoli (Brassica oleracea var. italica), with yield comparable to commercial varieties, because of the protection mechanism offered by molecular chaperones and stress-related proteins. Here, we used proximate analysis to examine macronutrients, chemical and mineral constituents as well as anti-nutrient and protein changes of ipt-transgenic broccoli and corresponding controls. We also preliminarily assessed safety in mice. Most aspects were comparable between ipt-transgenic broccoli and controls, except for a significant increase in carbohydrate level and a decrease in magnesium content in ipt-transgenic lines 101, 102 and 103, as compared with non-transgenic controls. In addition, the anti-nutrient glucosinolate content was increased and crude fat content decreased in inbred control 104 and transgenic lines as compared with the parental control, “Green King”. Gel-based proteomics detected more than 50 protein spots specifically found in ipt-transgenic broccoli at harvest and after cooking; one-third of these proteins showed homology to potential allergens that also play an important role in plant defense against stresses and senescence. Mice fed levels of ipt-transgenic broccoli mimicking the 120 g/day of broccoli eaten by a 60-kg human adult showed normal growth and immune function. In conclusion, the compositional and proteomic changes attributed to the transgenic ipt gene did not affect the growth and immune response of mice under the feeding regimes examined. PMID:25170807

Validation of PCR methods for quantitation of genetically modified plants in food.

PubMed

Hübner, P; Waiblinger, H U; Pietsch, K; Brodmann, P

2001-01-01

For enforcement of the recently introduced labeling threshold for genetically modified organisms (GMOs) in food ingredients, quantitative detection methods such as quantitative competitive (QC-PCR) and real-time PCR are applied by official food control laboratories. The experiences of 3 European food control laboratories in validating such methods were compared to describe realistic performance characteristics of quantitative PCR detection methods. The limit of quantitation (LOQ) of GMO-specific, real-time PCR was experimentally determined to reach 30-50 target molecules, which is close to theoretical prediction. Starting PCR with 200 ng genomic plant DNA, the LOQ depends primarily on the genome size of the target plant and ranges from 0.02% for rice to 0.7% for wheat. The precision of quantitative PCR detection methods, expressed as relative standard deviation (RSD), varied from 10 to 30%. Using Bt176 corn containing test samples and applying Bt176 specific QC-PCR, mean values deviated from true values by -7to 18%, with an average of 2+/-10%. Ruggedness of real-time PCR detection methods was assessed in an interlaboratory study analyzing commercial, homogeneous food samples. Roundup Ready soybean DNA contents were determined in the range of 0.3 to 36%, relative to soybean DNA, with RSDs of about 25%. Taking the precision of quantitative PCR detection methods into account, suitable sample plans and sample sizes for GMO analysis are suggested. Because quantitative GMO detection methods measure GMO contents of samples in relation to reference material (calibrants), high priority must be given to international agreements and standardization on certified reference materials.

Glucosinolate-Containing Seed Meal as a Soil Amendment to Control Plant Pests: 2000-2002

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brown, J.; Morra, M. J.

Plants may produce compounds that directly or indirectly affect their biological environment. These compounds fall within a broad category of compounds called allelochemicals, and are exclusive of food that influences growth, health, or behavior of other organisms (Whittaker and Feeney 1971). One reason for interest in allelochemicals is their potential for use in alternative pest management systems. Using plant-produced allelochemicals in agricultural and horticultural practices could minimize synthetic pesticide use, reduce the associated potential for environmental contamination, and contribute to a more sustainable agricultural system.

In vitro adherence of radioactively labeled Escherichia coli in normal and cystitis-prone females

DOE Office of Scientific and Technical Information (OSTI.GOV)

Parsons, C.L.; Anwar, H.; Stauffer, C.

Numerous investigators report data obtained using an in vitro quantitative assay for measuring bacterial adherence to epithelial cells. In the modified assay described here, we eliminated the need for visual counting of bacteria by incorporating the use of radioactively labeled Escherichia coli. This allowed quantitation of bacterial adherence to as many as 50,000 vaginal cells, whereas the visual counting system limits the determination to perhaps 50 cells. Using the modified method, we found no statistically significant differences among values for adherence of E. coli type 04 to the vaginal cells of control and cystitis-prone women at either pH 6.4 ormore » 4.0.« less

Stable, water extractable isothiocyanates from Moringa oleifera leaves attenuate inflammation in vitro.

PubMed

Waterman, Carrie; Cheng, Diana M; Rojas-Silva, Patricio; Poulev, Alexander; Dreifus, Julia; Lila, Mary Ann; Raskin, Ilya

2014-07-01

Moringa (Moringa oleifera Lam.) is an edible plant used as both a food and medicine throughout the tropics. A moringa concentrate (MC), made by extracting fresh leaves with water, utilized naturally occurring myrosinase to convert four moringa glucosinolates into moringa isothiocyanates. Optimum conditions maximizing MC yield, 4-[(α-L-rhamnosyloxy)benzyl]isothiocyanate, and 4-[(4'-O-acetyl-α-L-rhamnosyloxy)benzyl]isothiocyanate content were established (1:5 fresh leaf weight to water ratio at room temperature). The optimized MC contained 1.66% isothiocyanates and 3.82% total polyphenols. 4-[(4'-O-acetyl-α-L-rhamnosyloxy)benzyl]isothiocyanate exhibited 80% stability at 37°C for 30 days. MC, and both of the isothiocyanates described above significantly decreased gene expression and production of inflammatory markers in RAW macrophages. Specifically, both attenuated expression of iNOS and IL-1β and production of nitric oxide and TNFα at 1 and 5 μM. These results suggest a potential for stable and concentrated moringa isothiocyanates, delivered in MC as a food-grade product, to alleviate low-grade inflammation associated with chronic diseases. Copyright © 2014 Elsevier Ltd. All rights reserved.

One Novel Multiple-Target Plasmid Reference Molecule Targeting Eight Genetically Modified Canola Events for Genetically Modified Canola Detection.

PubMed

Li, Zhuqing; Li, Xiang; Wang, Canhua; Song, Guiwen; Pi, Liqun; Zheng, Lan; Zhang, Dabing; Yang, Litao

2017-09-27

Multiple-target plasmid DNA reference materials have been generated and utilized as good substitutes of matrix-based reference materials in the analysis of genetically modified organisms (GMOs). Herein, we report the construction of one multiple-target plasmid reference molecule, pCAN, which harbors eight GM canola event-specific sequences (RF1, RF2, MS1, MS8, Topas 19/2, Oxy235, RT73, and T45) and a partial sequence of the canola endogenous reference gene PEP. The applicability of this plasmid reference material in qualitative and quantitative PCR assays of the eight GM canola events was evaluated, including the analysis of specificity, limit of detection (LOD), limit of quantification (LOQ), and performance of pCAN in the analysis of various canola samples, etc. The LODs are 15 copies for RF2, MS1, and RT73 assays using pCAN as the calibrator and 10 genome copies for the other events. The LOQ in each event-specific real-time PCR assay is 20 copies. In quantitative real-time PCR analysis, the PCR efficiencies of all event-specific and PEP assays are between 91% and 97%, and the squared regression coefficients (R 2 ) are all higher than 0.99. The quantification bias values varied from 0.47% to 20.68% with relative standard deviation (RSD) from 1.06% to 24.61% in the quantification of simulated samples. Furthermore, 10 practical canola samples sampled from imported shipments in the port of Shanghai, China, were analyzed employing pCAN as the calibrator, and the results were comparable with those assays using commercial certified materials as the calibrator. Concluding from these results, we believe that this newly developed pCAN plasmid is one good candidate for being a plasmid DNA reference material in the detection and quantification of the eight GM canola events in routine analysis.

Quantitation of phosphatidic acid and lysophosphatidic acid molecular species using hydrophilic interaction liquid chromatography coupled to electrospray ionization high resolution mass spectrometry.

PubMed

Triebl, Alexander; Trötzmüller, Martin; Eberl, Anita; Hanel, Pia; Hartler, Jürgen; Köfeler, Harald C

2014-06-20

A method for a highly selective and sensitive identification and quantitation of lysophosphatidic acid (LPA) and phosphatidic acid (PA) molecular species was developed using hydrophilic interaction liquid chromatography (HILIC) followed by negative-ion electrospray ionization high resolution mass spectrometry. Different extraction methods for the polar LPA and PA species were compared and a modified Bligh & Dyer extraction by addition of 0.1M hydrochloric acid resulted in a ≈1.2-fold increase of recovery for the 7 PA and a more than 15-fold increase for the 6 LPA molecular species of a commercially available natural mix compared to conventional Bligh & Dyer extraction. This modified Bligh & Dyer extraction did not show any artifacts resulting from hydrolysis of natural abundant phospholipids. The developed HILIC method is able to separate all PA and LPA species from major polar membrane lipid classes which might have suppressive effects on the minor abundant lipid classes of interest. The elemental compositions of intact lipid species are provided by the high mass resolution of 100,000 and high mass accuracy below 3ppm of the Orbitrap instrument. Additionally, tandem mass spectra were generated in a parallel data dependent acquisition mode in the linear ion trap to provide structural information at molecular level. Limits of quantitation were identified at 45fmol on column and the dynamic range reaches 20pmol on column, covering the range of natural abundance well. By applying the developed method to mouse brain it can be shown that phosphatidic acid contains less unsaturated fatty acids with PA 34:1 and PA 36:1 as the major species. In contrast, for LPA species a high content of polyunsaturated fatty acids (LPA 20:4 and LPA 22:6) was quantified. Copyright © 2014 Elsevier B.V. All rights reserved.

Absolute quantitation of isoforms of post-translationally modified proteins in transgenic organism.

PubMed

Li, Yaojun; Shu, Yiwei; Peng, Changchao; Zhu, Lin; Guo, Guangyu; Li, Ning

2012-08-01

Post-translational modification isoforms of a protein are known to play versatile biological functions in diverse cellular processes. To measure the molar amount of each post-translational modification isoform (P(isf)) of a target protein present in the total protein extract using mass spectrometry, a quantitative proteomic protocol, absolute quantitation of isoforms of post-translationally modified proteins (AQUIP), was developed. A recombinant ERF110 gene overexpression transgenic Arabidopsis plant was used as the model organism for demonstration of the proof of concept. Both Ser-62-independent (14)N-coded synthetic peptide standards and (15)N-coded ERF110 protein standard isolated from the heavy nitrogen-labeled transgenic plants were employed simultaneously to determine the concentration of all isoforms (T(isf)) of ERF110 in the whole plant cell lysate, whereas a pair of Ser-62-dependent synthetic peptide standards were used to quantitate the Ser-62 phosphosite occupancy (R(aqu)). The P(isf) was finally determined by integrating the two empirically measured variables using the following equation: P(isf) = T(isf) · R(aqu). The absolute amount of Ser-62-phosphorylated isoform of ERF110 determined using AQUIP was substantiated with a stable isotope labeling in Arabidopsis-based relative and accurate quantitative proteomic approach. The biological role of the Ser-62-phosphorylated isoform was demonstrated in transgenic plants.

Quantitative description of respiration processes in meso-eutrophic and eutrophic freshwater environments.

PubMed

Kiersztyn, Bartosz; Kauppinen, Elsi S; Kaliński, Tomasz; Chróst, Ryszard; Siuda, Waldemar

2018-06-01

We propose a modification of measurement methodology allowing the overall respiration rate (V Resp ) close to the in situ conditions; size of the labile, respirable organic matter pool (OM Resp ); and its turnover time (Tt) to be calculated. In addition to the respiration of dissolved substrates by free-living bacteria, the respiration of attached bacteria and other planktonic organisms is also taken into account. In case study we evaluated the modified, quantitative description of respiration processes in surface waters of lakes of different trophic status: mezzo-eutrophic and eutrophic. In both types of studied environments, V Resp oscillated between 1.0 μmol C l -1  h -1 and 3.0 μmol C l -1  h -1 , and the size of the OM Resp pool varied from 39.3 μM C to 828.7 μM C. Despite of higher OM Resp concentrations in eutrophic lakes, we found a lower susceptibility of OM to respiration processes in eutrophic than in meso-eutrophic lakes but similar V Resp in both types of lakes. We conclude that the proposed method allows a fast quantitative description of labile organic matter utilization by aerobic aquatic microorganisms. Copyright © 2018 Elsevier B.V. All rights reserved.

Utility of Von Pechman synthesis of coumarin reaction for development of spectrofluorimetric method for quantitation of salmeterol xinafoate in pharmaceutical preparations and human plasma.

PubMed

Awad, Mohamed; Hammad, Mohamed A; Abdel-Megied, Ahmed M; Omar, Mahmoud A

2018-04-30

Simple, precise and selective spectrofluorimetric technique was evolved for quantitation of selective β 2 agonist drug namely salmeterol xinafoate (SAL). Utilizing its phenolic nature, a method was described based on the reaction of the studied drug with ethyl acetoacetate (EAA) to yield extremely fluorescent coumarin product which can be detected at 480 nm (λ ex  = 420 nm). The procedure obeys Beer's law with a correlation coefficient of r = 0.9999 in the concentration range between 500 and 5000 ng ml -1 with and 177 ng ml -1 for limit of detection (LOD) and limit of quantification (LOQ), respectively. Diverse reaction variables influencing the firmness and formation of the coumarin product were accurately examined and modified to ensure greatest sensitivity of the procedure. The proposed technique was performed and examined according to the US Food and Drug Administration (FDA) guidelines for bio-analytical methods and was efficiently applied for quantitation of SAL in both pharmaceutical preparations (% recovery = 100.06 ± 1.07) and spiked human plasma (% recovery = 96.64-97.14 ± 1.01-1.52). Copyright © 2018 John Wiley & Sons, Ltd.

A sensitive method for determination of COL-3, a chemically modified tetracycline, in human plasma using high-performance liquid chromatography and ultraviolet detection.

PubMed

Rudek, Michelle A; Hartke, Carol; Zabelina, Yelena; Zhao, Ming; New, Pamela; Baker, Sharyn D

2005-04-01

COL-3, 6-deoxy-6-desmethyl-4-desdimethylamino-tetracycline, is a matrix metalloproteinase inhibitor currently in clinical development. A HPLC-UV method to quantitate COL-3 in human plasma was developed. COL-3 was extracted from plasma using solid-phase extraction cartridges. COL-3 is separated on a Waters Symmetry Shield RP8 (3.9 mm x150 mm, 5 microm) column with EDTA (0.001 M) in sodium acetate (0.01 M, pH 3.5)-acetonitrile mobile phase using a gradient profile at a flow rate of 1 ml/min for 22 min. Carryover was eliminated by using an extended needle wash of methanol:acetonitrile:dichloromethane (1:1:1, v/v/v). Detection of COL-3 and the internal standard, chrysin, was observed at 350 nm. COL-3 and chrysin elute at 8.9 and 9.9 min, respectively. The lower limit of quantitation in human plasma of COL-3 was 75 ng/ml, linearity was observed from 75 to 10,000 ng/ml. A 30,000 ng/ml sample that was diluted 1:50 with plasma was accurately quantitated. This method is rapid, widely applicable, and suitable for quantifying COL-3 in patient samples enabling further clinical pharmacology characterization of COL-3.

Pre-harvest methyl jasmonate treatment enhances cauliflower chemoprotective attributes without a loss in postharvest quality.

PubMed

Ku, Kang Mo; Choi, Jeong-Hee; Kushad, Mosbah M; Jeffery, Elizabeth H; Juvik, John A

2013-06-01

Methyl jasmonate (MeJA) treatment can significantly increase glucosinolate (GS) concentrations in Brassica vegetables and potentially enhance anticancer bioactivity. Although MeJA treatment may promote ethylene biosynthesis, which can be detrimental to postharvest quality, there are no previous reports of its effect on cauliflower postharvest quality. To address this, cauliflower curds in field plots were sprayed with either 0.1 % Triton X-100 (control) or 500 μM MeJA solutions four days prior to harvest, then stored at 4 °C. Tissue subsamples were collected after 0, 10, 20, and 30 days of postharvest storage and assayed for visual color change, ethylene production, GS concentrations, and extract quinone reductase inductive activity. MeJA treatment increased curd GS concentrations of glucoraphanin, glucobrassicin, and neoglucobrassicin by 1.5, 2.4, and 4.6-fold over controls, respectively. MeJA treated cauliflower showed significantly higher quinone reductase activity, a biomarker for anticancer bioactivity, without reducing visual color and postharvest quality for 10 days at 4 °C storage.

Quantitative Microbial Risk Assessment Tutorial - SDMProjectBuilder: Import Local Data Files to Identify and Modify Contamination Sources and Input ParametersUpdated 2017

EPA Science Inventory

Twelve example local data support files are automatically downloaded when the SDMProjectBuilder is installed on a computer. They allow the user to modify values to parameters that impact the release, migration, fate, and transport of microbes within a watershed, and control delin...

Quantitative Microbial Risk Assessment Tutorial – SDMProjectBuilder: Import Local Data Files to Identify and Modify Contamination Sources and Input Parameters

EPA Science Inventory

Twelve example local data support files are automatically downloaded when the SDMProjectBuilder is installed on a computer. They allow the user to modify values to parameters that impact the release, migration, fate, and transport of microbes within a watershed, and control delin...

SU-E-I-62: Reduction of Susceptibility Artifacts by Increasing the Bandwidth (BW) and Echo Train Length (ETL)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mavroidis, P; Boci, N; Kostopoulos, S

2015-06-15

Purpose: The aim of this present study is to increase bandwidth (BW) and echo train length (ETL) in Proton Density Turbo Spin Echo (PD TSE) sequences with and without fat saturation (FS) as well as in Turbo Inversion Recovery Magnitude sequences (TIRM) in order to assess whether these sequences are capable of reducing susceptibility artifacts. Methods: We compared 1) TIRM coronal (COR) with the same sequence with increased both BW and ETL 2) Conventional PD TSE sagittal (SAG) with FS with an increased BW 3) Conventional PD TSE SAG without FS with an increased BW 4) Conventional PD TSE SAGmore » without FS with increased both BW and ETL. A quantitative analysis was performed to measure the extent of the susceptibility artifacts. Furthermore, a qualitative analysis was performed by two radiologists in order to evaluate the susceptibility artifacts, image distortion and fat suppression. The depiction of cartilage, menisci, muscles, tendons and bone marrow were also qualitatively analyzed. Results: The quantitative analysis found that the modified TIRM sequence is significantly superior to the conventional one regarding the extent of the susceptibility artifacts. In the qualitative analysis, the modified TIRM sequence was superior to the corresponding conventional one in eight characteristics out of ten that were analyzed. The modified PD TSE with FS was superior to the corresponding conventional one regarding the susceptibility artifacts, image distortion and depiction of bone marrow and cartilage while achieving effective fat saturation. The modified PD TSE sequence without FS with a high (H) BW was found to be superior corresponding to the conventional one in the case of cartilage. Conclusion: Consequently, TIRM sequence with an increased BW and ETL is proposed for producing images of high quality and modified PD TSE with H BW for smaller metals, especially when FS is used.« less

Qualitative PCR method for Roundup Ready soybean: interlaboratory study.

PubMed

Kodama, Takashi; Kasahara, Masaki; Minegishi, Yasutaka; Futo, Satoshi; Sawada, Chihiro; Watai, Masatoshi; Akiyama, Hiroshi; Teshima, Reiko; Kurosawa, Yasunori; Furui, Satoshi; Hino, Akihiro; Kitta, Kazumi

2011-01-01

Quantitative and qualitative methods based on PCR have been developed for genetically modified organisms (GMO). Interlaboratory studies were previously conducted for GMO quantitative methods; in this study, an interlaboratory study was conducted for a qualitative method for a GM soybean, Roundup Ready soy (RR soy), with primer pairs designed for the quantitative method of RR soy studied previously. Fourteen laboratories in Japan participated. Each participant extracted DNA from 1.0 g each of the soy samples containing 0, 0.05, and 0.10% of RR soy, and performed PCR with primer pairs for an internal control gene (Le1) and RR soy followed by agarose gel electrophoresis. The PCR product amplified in this PCR system for Le1 was detected from all samples. The sensitivity, specificity, and false-negative and false-positive rates of the method were obtained from the results of RR soy detection. False-negative rates at the level of 0.05 and 0.10% of the RR soy samples were 6.0 and 2.3%, respectively, revealing that the LOD of the method was somewhat below 0.10%. The current study demonstrated that the qualitative method would be practical for monitoring the labeling system of GM soy in kernel lots.

Method for the routine quantitative gas chromatographic analysis of major free fatty acids in butter and cream.

PubMed

Woo, A H; Lindsay, R C

1980-07-01

A rapid quantiative method was developed for routine analysis of the major, even carbon-numbered free fatty acids in butter and cream. Free fatty acids were isolated directly from intact samples by a modified silicic acid-potassium hydroxide arrestant column and were separated by gas chromatography with a 1.8 m x 2 mm inner diameter glass column packed with 10% neopentyl glycol adipate on 80/100 Chromosorb W. Purified, formic acid-saturated carrier gas was required for minimal peak tailing and extended column life. The accuracy and reproducibility of the mmethod was established through quantitative recovery studies of free fatty acid mixtures, free fatty acids added to butter, and replicate analysis of butter and cream samples.

[Development and validation of event-specific quantitative PCR method for genetically modified maize LY038].

PubMed

Mano, Junichi; Masubuchi, Tomoko; Hatano, Shuko; Futo, Satoshi; Koiwa, Tomohiro; Minegishi, Yasutaka; Noguchi, Akio; Kondo, Kazunari; Akiyama, Hiroshi; Teshima, Reiko; Kurashima, Takeyo; Takabatake, Reona; Kitta, Kazumi

2013-01-01

In this article, we report a novel real-time PCR-based analytical method for quantitation of the GM maize event LY038. We designed LY038-specific and maize endogenous reference DNA-specific PCR amplifications. After confirming the specificity and linearity of the LY038-specific PCR amplification, we determined the conversion factor required to calculate the weight-based content of GM organism (GMO) in a multilaboratory evaluation. Finally, in order to validate the developed method, an interlaboratory collaborative trial according to the internationally harmonized guidelines was performed with blind DNA samples containing LY038 at the mixing levels of 0, 0.5, 1.0, 5.0 and 10.0%. The precision of the method was evaluated as the RSD of reproducibility (RSDR), and the values obtained were all less than 25%. The limit of quantitation of the method was judged to be 0.5% based on the definition of ISO 24276 guideline. The results from the collaborative trial suggested that the developed quantitative method would be suitable for practical testing of LY038 maize.

Mixing of Exciton and Charge-Transfer States in Photosystem II Reaction Centers: Modeling of Stark Spectra with Modified Redfield Theory

PubMed Central

Novoderezhkin, Vladimir I.; Dekker, Jan P.; van Grondelle, Rienk

2007-01-01

We propose an exciton model for the Photosystem II reaction center (RC) based on a quantitative simultaneous fit of the absorption, linear dichroism, circular dichroism, steady-state fluorescence, triplet-minus-singlet, and Stark spectra together with the spectra of pheophytin-modified RCs, and so-called RC5 complexes that lack one of the peripheral chlorophylls. In this model, the excited state manifold includes a primary charge-transfer (CT) state that is supposed to be strongly mixed with the pure exciton states. We generalize the exciton theory of Stark spectra by 1), taking into account the coupling to a CT state (whose static dipole cannot be treated as a small parameter in contrast to usual excited states); and 2), expressing the line shape functions in terms of the modified Redfield approach (the same as used for modeling of the linear responses). This allows a consistent modeling of the whole set of experimental data using a unified physical picture. We show that the fluorescence and Stark spectra are extremely sensitive to the assignment of the primary CT state, its energy, and coupling to the excited states. The best fit of the data is obtained supposing that the initial charge separation occurs within the special-pair PD1PD2. Additionally, the scheme with primary electron transfer from the accessory chlorophyll to pheophytin gave a reasonable quantitative fit. We show that the effectiveness of these two pathways is strongly dependent on the realization of the energetic disorder. Supposing a mixed scheme of primary charge separation with a disorder-controlled competition of the two channels, we can explain the coexistence of fast sub-ps and slow ps components of the Phe-anion formation as revealed by different ultrafast spectroscopic techniques. PMID:17526589

Distribution network connection pricing framework and methodology: identification of areas of improvement for Sarawak Energy Berhad Connection Charges Guidelines through modified delphi method

NASA Astrophysics Data System (ADS)

Tan, J. K.; Abas, N.

2017-07-01

Complaints on issues and matters related to connection charges have been very common for electricity supply utility companies around the world including Sarawak Energy Berhad. In order to identify the areas that can be improved, a mixed method of exploratory research involving qualitative and quantitative methods have been designed and undertaken rather than a single method of survey. This will ensure a more comprehensive and detailed understanding of the issues from various target groups. The method is designed under three phases, employing Modified Delphi Technique for phase 1 through a series of stake holder engagements, online and offline survey questionnaires to be filled by internal wiring contractors for phase 2 whilst under phase 3, case studies shall be carried out on the issues identified from phase 1 and phase 2 of the study. This paper presented the findings from the Modified Delphi Technique. The findings revealed that there are areas of improvement for Sarawak Energy Berhad connection guidelines in term of differentiation of dedicated and shared assets which leads to unfairness to the connecting customers, inconsistency and non-transparent in charging. The findings of Modified Delphi Technique shall be used for implementation of phase 2 and phase 3 of the study.

Genome-wide association study to identify potential genetic modifiers in a canine model for Duchenne muscular dystrophy.

PubMed

Brinkmeyer-Langford, Candice; Balog-Alvarez, Cynthia; Cai, James J; Davis, Brian W; Kornegay, Joe N

2016-08-22

Duchenne muscular dystrophy (DMD) causes progressive muscle degeneration, cardiomyopathy and respiratory failure in approximately 1/5,000 boys. Golden Retriever muscular dystrophy (GRMD) resembles DMD both clinically and pathologically. Like DMD, GRMD exhibits remarkable phenotypic variation among affected dogs, suggesting the influence of modifiers. Understanding the role(s) of genetic modifiers of GRMD may identify genes and pathways that also modify phenotypes in DMD and reveal novel therapies. Therefore, our objective in this study was to identify genetic modifiers that affect discrete GRMD phenotypes. We performed a linear mixed-model (LMM) analysis using 16 variably-affected dogs from our GRMD colony (8 dystrophic, 8 non-dystrophic). All of these dogs were either full or half-siblings, and phenotyped for 19 objective, quantitative biomarkers at ages 6 and 12 months. Each biomarker was individually assessed. Gene expression profiles of 59 possible candidate genes were generated for two muscle types: the cranial tibialis and medial head of the gastrocnemius. SNPs significantly associated with GRMD biomarkers were identified on multiple chromosomes (including the X chromosome). Gene expression levels for candidate genes located near these SNPs correlated with biomarker values, suggesting possible roles as GRMD modifiers. The results of this study enhance our understanding of GRMD pathology and represent a first step toward the characterization of GRMD modifiers that may be relevant to DMD pathology. Such modifiers are likely to be useful for DMD treatment development based on their relationships to GRMD phenotypes.

Cooperativity and specificity of association of a designed transmembrane peptide.

PubMed Central

Gratkowski, Holly; Dai, Qing-Hong; Wand, A Joshua; DeGrado, William F; Lear, James D

2002-01-01

Thermodynamics studies aimed at quantitatively characterizing free energy effects of amino acid substitutions are not restricted to two state systems, but do require knowing the number of states involved in the equilibrium under consideration. Using analytical ultracentrifugation and NMR methods, we show here that a membrane-soluble peptide, MS1, designed by modifying the sequence of the water-soluble coiled-coil GCN4-P1, exhibits a reversible monomer-dimer-trimer association in detergent micelles with a greater degree of cooperativity in C14-betaine than in dodecyl phosphocholine detergents. PMID:12202385

In situ spectroscopic characterization of a solution-phase X-type ligand exchange at colloidal lead sulphide quantum dot surfaces

DOE PAGES

Kroupa, Daniel M.; Anderson, Nicholas C.; Castaneda, Chloe V.; ...

2016-11-07

Here, we employed quantitative NMR spectroscopy and spectrophotometric absorbance titration to study a quantum dot X-type ligand exchange reaction. We find that the exchange is highly cooperative, where at low extents of exchange the change in free energy of the reaction, Δ G XC, is ~11 kJ mol –1 while at higher extents of exchange Δ G XC saturates to ~–4 kJ mol –1. A modified Fowler binding isotherm is developed to describe the reaction.

Promoter isolation and characterization of GhAO-like1, a Gossypium hirsutum gene similar to multicopper oxidases that is highly expressed in reproductive organs.

PubMed

Lambret-Frotté, Julia; Artico, Sinara; Muniz Nardeli, Sarah; Fonseca, Fernando; Brilhante Oliveira-Neto, Osmundo; Grossi-de-Sá, Maria Fatima; Alves-Ferreira, Marcio

2016-01-01

Cotton is one of the most economically important cultivated crops. It is the major source of natural fiber for the textile industry and an important target for genetic modification for both biotic stress and herbicide tolerance. Therefore, the characterization of genes and regulatory regions that might be useful for genetic transformation is indispensable. The isolation and characterization of new regulatory regions is of great importance to drive transgene expression in genetically modified crops. One of the major drawbacks in cotton production is pest damage; therefore, the most promising, cost-effective, and sustainable method for pest control is the development of genetically resistant cotton lines. Considering this scenario, our group isolated and characterized the promoter region of a MCO (multicopper oxidase) from Gossypium hirsutum, named GhAO-like1 (ascorbate oxidase-like1). The quantitative expression, together with the in vivo characterization of the promoter region reveals that GhAO-like1 has a flower- and fruit-specific expression pattern. The GUS activity is mainly observed in stamens, as expected considering that the GhAO-like1 regulatory sequence is enriched in cis elements, which have been characterized as a target of reproductive tissue specific transcription factors. Both histological and quantitative analyses in Arabidopsis thaliana have confirmed flower (mainly in stamens) and fruit expression of GhAO-like1. In the present paper, we isolated and characterized both in silico and in vivo the promoter region of the GhAO-like1 gene. The regulatory region of GhAO-like1 might be useful to confer tissue-specific expression in genetically modified plants.

The dose-dependent influence of zinc and cadmium contamination of soil on their uptake and glucosinolate content in white cabbage (Brassica oleracea var. capitata f. alba).

PubMed

Kusznierewicz, Barbara; Bączek-Kwinta, Renata; Bartoszek, Agnieszka; Piekarska, Anna; Huk, Anna; Manikowska, Anna; Antonkiewicz, Jacek; Namieśnik, Jacek; Konieczka, Piotr

2012-11-01

The relationship between the ability to accumulate heavy metals (represented by Cd and Zn) and to synthesize bioactive compounds (represented by glucosinolates [GLS]) was investigated in two cabbage cultivars. Plants were grown in the greenhouse of a phytotron under controlled conditions in soils spiked with two different Zn or Cd concentrations. The measurements of Cd and Zn contents in soil and cabbage (leaf) samples were performed by atomic absorption spectroscopy, whereas GLS levels in cabbage were determined by high-performance liquid chromatography. The ranges of metal contents in soil were 80 to 450 mg/kg dry weight for Zn and 0.3 to 30 mg/kg dry weight for Cd, whereas the levels of accumulated Zn and Cd in cabbage amounted to 15 to 130 and 0.02 to 3 mg/kg dry weight, respectively. After initial symptoms of toxicity, during a later stage of growth, the plants exhibited very good tolerance to both metals. Enhanced biosynthesis of GLS was observed in a dose-dependent manner following exposure to the heavy metals. The GLS content in Zn-exposed cabbage rose from 3.2 to 12 µmol/g dry weight, and the corresponding values for Cd-treated plants were 3.5 to 10 µmol/g dry weight. Thus, the increased soil contamination by metals caused greater accumulation in cabbage, as well as stimulation of GLS biosynthesis. The results obtained point to the high phytoremediation and biofumigation potential of white cabbage. Copyright © 2012 SETAC.

Food as Pharma? The Case of Glucosinolates.

PubMed

Capuano, Edoardo; Dekker, Matthijs; Verkerk, Ruud; Oliviero, Teresa

2017-01-01

Glucosinolates (GLSs) are dietary plant secondary metabolites occurring in the order Brassicales with potential health effects, in particular as anti-carcinogenic compounds. GLSs are converted into a variety of breakdown products (BPs) upon plant tissue damage and by the gut microbiota. GLS biological activity is related to BPs rather than to GLSs themselves. we have reviewed the most recent scientific literature on the metabolic fate and the biological effect of GLSs with particular emphasis on the epidemiological evidence for health effect and evidence from clinical trials. An overview of potential molecular mechanisms underlying GLS biological effect is provided. The potential toxic or anti-nutritional effect has also been discussed. Epidemiological and human in vivo evidence point towards a potential anti-cancer effect for sulforaphane, indole-3-carbinol and 3,3-diindolylmethane. A number of new human clinical trials are on-going and will likely shed further light on GLS protective effect towards cancer as well as other diseases. BPs biological effect is the results of a plurality of molecular mechanisms acting simultaneously which include modulation of xenobiotic metabolism, modulation of inflammation, regulation of apoptosis, cell cycle arrest, angiogenesis and metastasis and regulation of epigenetic events. BPs have been extensively investigated for their protective effect towards cancer but in recent years the interest also includes other diseases. It appears that certain BPs may protect against and may even represent a therapeutic strategy against several forms of cancer. Whether this latter effect can be achieved through diet or supplements should be investigated more thoroughly. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Combined effect of Nitrogen, Phosphorus and Potassium fertilizers on the contents of glucosinolates in rocket salad (Eruca sativa Mill.).

PubMed

Chun, Jin-Hyuk; Kim, Silbia; Arasu, Mariadhas Valan; Al-Dhabi, Naif Abdullah; Chung, Doug Young; Kim, Sun-Ju

2017-02-01

Nitrogen (N), phosphorous (P) and potassium (K) are the most limiting factors in crop production. N often affects the amino acid composition of protein and in turn its nutritional quality. In Brassica plants, abundant supply of N fertilizer decreases the relative proportion of glucosinolates (GSLs), thus reducing the biological and medical values of the vegetables. Hence effort was made to evaluate the influence of different proportions of nutrient solutions containing N-P-K on the GSL profiles of rocket salad ( Eruca sativa Mill.). Fifteen desulpho-(DS) GSLs were isolated and identified using liquid chromatography-mass spectrometry (LC/MS) analysis. Rocket salad plants supplied with lesser amount of N, P or higher concentrations of K showed a typical improvement in total GSL contents. In contrast, total GSL levels were less at higher N supply. Furthermore, with N concentrations above 5 mM and K concentrations less than 2.5 mM, the GSL amounts were on average 13.51 and 13.75 μmol/g dry weight (DW), respectively. Aliphatic GSLs predominated in all concentrations of NPK while indolyl GSLs made up marginally less amount of the total compositions. Five and 2 mM N and P possessed much higher levels of several types of aliphatic GSLs than other concentrations, including glucoerucin, glucoraphanin and dimeric 4-mercaptobutyl GSL. From this perspective, it is contended that supply of less N results in enhancing the metabolic pathway for the synthesis of GSLs in rocket salad.

Disintegration of microtubules in Arabidopsis thaliana and bladder cancer cells by isothiocyanates

PubMed Central

Øverby, Anders; Bævre, Mette S.; Thangstad, Ole P.; Bones, Atle M.

2015-01-01

Isothiocyanates (ITCs) from biodegradation of glucosinolates comprise a group of electrophiles associated with growth-inhibitory effects in plant- and mammalian cells. The underlying modes of action of this feature are not fully understood. Clarifying this has involved mammalian cancer cells due to ITCs' chemopreventive potential. The binding of ITCs to tubulins has been reported as a mechanism by which ITCs induce cell cycle arrest and apoptosis. In the present study we demonstrate that ITCs disrupt microtubules in Arabidopsis thaliana contributing to the observed inhibited growth phenotype. We also confirmed this in rat bladder cancer cells (AY-27) suggesting that cells from plant and animals share mechanisms by which ITCs affect growth. Exposure of A. thaliana to vapor-phase of allyl ITC (AITC) inhibited growth and induced a concurrent bleaching of leaves in a dose-dependent manner. Transcriptional analysis was used to show an upregulation of heat shock-genes upon AITC-treatment. Transgenic A. thaliana expressing GFP-marked α-tubulin was employed to show a time- and dose-dependent disintegration of microtubules by AITC. Treatment of AY-27 with ITCs resulted in a time- and dose-dependent decrease of cell proliferation and G2/M-arrest. AY-27 transiently transfected to express GFP-tagged α-tubulin were treated with ITCs resulting in a loss of microtubular filaments and the subsequent formation of apoptotic bodies. In conclusion, our data demonstrate an ITC-induced mechanism leading to growth inhibition in A. thaliana and rat bladder cancer cells, and expose clues to the mechanisms underlying the physiological role of glucosinolates in vivo. PMID:25657654

Whole Genome and Tandem Duplicate Retention Facilitated Glucosinolate Pathway Diversification in the Mustard Family

PubMed Central

Hofberger, Johannes A.; Lyons, Eric; Edger, Patrick P.; Chris Pires, J.; Eric Schranz, M.

2013-01-01

Plants share a common history of successive whole-genome duplication (WGD) events retaining genomic patterns of duplicate gene copies (ohnologs) organized in conserved syntenic blocks. Duplication was often proposed to affect the origin of novel traits during evolution. However, genetic evidence linking WGD to pathway diversification is scarce. We show that WGD and tandem duplication (TD) accelerated genetic versatility of plant secondary metabolism, exemplified with the glucosinolate (GS) pathway in the mustard family. GS biosynthesis is a well-studied trait, employing at least 52 biosynthetic and regulatory genes in the model plant Arabidopsis. In a phylogenomics approach, we identified 67 GS loci in Aethionema arabicum of the tribe Aethionemae, sister group to all mustard family members. All but one of the Arabidopsis GS gene families evolved orthologs in Aethionema and all but one of the orthologous sequence pairs exhibit synteny. The 45% fraction of duplicates among all protein-coding genes in Arabidopsis was increased to 95% and 97% for Arabidopsis and Aethionema GS pathway inventory, respectively. Compared with the 22% average for all protein-coding genes in Arabidopsis, 52% and 56% of Aethionema and Arabidopsis GS loci align to ohnolog copies dating back to the last common WGD event. Although 15% of all Arabidopsis genes are organized in tandem arrays, 45% and 48% of GS loci in Arabidopsis and Aethionema descend from TD, respectively. We describe a sequential combination of TD and WGD events driving gene family extension, thereby expanding the evolutionary playground for functional diversification and thus potential novelty and success. PMID:24171911

Isothiocyanates: cholinesterase inhibiting, antioxidant, and anti-inflammatory activity.

PubMed

Burčul, Franko; Generalić Mekinić, Ivana; Radan, Mila; Rollin, Patrick; Blažević, Ivica

2018-12-01

Finding a new type of cholinesterase inhibitor that would overcome the brain availability and pharmacokinetic parameters or hepatotoxic liability has been a focus of investigations dealing with the treatment of Alzheimer's disease. Isothiocyanates have not been previously investigated as potential cholinesterase inhibitors. These compounds can be naturally produced from their glucosinolate precursors, secondary metabolites widely distributed in our daily Brassica vegetables. Among 11 tested compounds, phenyl isothiocyanate and its derivatives showed the most promising inhibitory activity. 2-Methoxyphenyl ITC showed best inhibition on acetylcholinesterase with IC 50 of 0.57 mM, while 3-methoxyphenyl ITC showed the best inhibition on butyrylcholinesterase having 49.2% at 1.14 mM. Assessment of the antioxidant efficacy using different methods led to a similar conclusion. The anti-inflammatory activity was also tested using human COX-2 enzyme, ranking phenyl isothiocyanate, and 3-methoxyphenyl isothiocyanate as most active, with ∼99% inhibition at 50 μM.

Pendulum and modified pendulum appliances for maxillary molar distalization in Class II malocclusion - a systematic review.

PubMed

Al-Thomali, Yousef; Basha, Sakeenabi; Mohamed, Roshan Noor

2017-08-01

The main purpose of the present systematic review was to evaluate the quantitative effects of the pendulum appliance and modified pendulum appliances for maxillary molar distalization in Class II malocclusion. Our systematic search included MEDLINE, EMBASE, CINAHL, PsychINFO, Scopus and key journals and review articles; the date of the last search was 30 January 2017. We graded the methodological quality of the studies by means of the Quality Assessment Tool for Quantitative Studies, developed for the Effective Public Health Practice Project (EPHPP). In total, 203 studies were identified for screening, and 25 studies were eligible. The quality assessment rated four (16%) of the study as being of strong quality and 21 (84%) of these studies as being of moderate quality. The pendulum appliances showed mean molar distalization of 2-6.4 mm, distal tipping of molars from 6.67° to 14.50° and anchorage loss with mean premolar and incisor mesial movement of 1.63-3.6 mm and 0.9-6.5 mm, respectively. The bone anchored pendulum appliances (BAPAs) showed mean molar distalization of 4.8-6.4 mm, distal tipping of molars from 9° to 11.3° and mean premolar distalization of 2.7-5.4 mm. Pendulum and modified pendulum appliances are effective in molar distalization. Pendulum appliance with K-loop modification, implant supported pendulum appliance and BAPA significantly reduced anchorage loss of the anterior teeth and distal tipping of the molar teeth.