Multi-laboratory survey of qPCR enterococci analysis method performance

EPA Pesticide Factsheets

Quantitative polymerase chain reaction (qPCR) has become a frequently used technique for quantifying enterococci in recreational surface waters, but there are several methodological options. Here we evaluated how three method permutations, type of mastermix, sample extract dilution and use of controls in results calculation, affect method reliability among multiple laboratories with respect to sample interference. Multiple samples from each of 22 sites representing an array of habitat types were analyzed using EPA Method 1611 and 1609 reagents with full strength and five-fold diluted extracts. The presence of interference was assessed three ways: using sample processing and PCR amplifications controls; consistency of results across extract dilutions; and relative recovery of target genes from spiked enterococci in water sample compared to control matrices with acceptable recovery defined as 50 to 200%. Method 1609, which is based on an environmental mastermix, was found to be superior to Method 1611, which is based on a universal mastermix. Method 1611 had over a 40% control assay failure rate with undiluted extracts and a 6% failure rate with diluted extracts. Method 1609 failed in only 11% and 3% of undiluted and diluted extracts analyses. Use of sample processing control assay results in the delta-delta Ct method for calculating relative target gene recoveries increased the number of acceptable recovery results. Delta-delta tended to bias recoveries fr

An improved method for field extraction and laboratory analysis of large, intact soil cores

USGS Publications Warehouse

Tindall, J.A.; Hemmen, K.; Dowd, J.F.

1992-01-01

Various methods have been proposed for the extraction of large, undisturbed soil cores and for subsequent analysis of fluid movement within the cores. The major problems associated with these methods are expense, cumbersome field extraction, and inadequate simulation of unsaturated flow conditions. A field and laboratory procedure is presented that is economical, convenient, and simulates unsaturated and saturated flow without interface flow problems and can be used on a variety of soil types. In the field, a stainless steel core barrel is hydraulically pressed into the soil (30-cm diam. and 38 cm high), the barrel and core are extracted from the soil, and after the barrel is removed from the core, the core is then wrapped securely with flexible sheet metal and a stainless mesh screen is attached to the bottom of the core for support. In the laboratory the soil core is set atop a porous ceramic plate over which a soil-diatomaceous earth slurry has been poured to assure good contact between plate and core. A cardboard cylinder (mold) is fastened around the core and the empty space filled with paraffin wax. Soil cores were tested under saturated and unsaturated conditions using a hanging water column for potentials ???0. Breakthrough curves indicated that no interface flow occurred along the edge of the core. This procedure proved to be reliable for field extraction of large, intact soil cores and for laboratory analysis of solute transport.

Field and laboratory determination of a poly(vinyl/vinylidene chloride) additive in brick mortar.

PubMed

Law, S L; Newman, J H; Ptak, F L

1990-02-01

A polymerized vinyl/vinylidene chloride additive, used in brick mortar during the 60s and 70s, is detected at the building site by the field method, which employs a commercially available chloride test strip. The field test results can then be verified by the laboratory methods. In one method, total chlorine in the mortar is determined by an oxygen-bomb method and the additive chloride is determined by difference after water-soluble chlorides have been determined on a separate sample. In the second method, the polymerized additive is extracted directly from the mortar with tetrahydrofuran (THF). The difference in weight before and after extraction of the additive gives the weight of additive in the mortar. Evaporation of the THF from the extract leaves a thin film of the polymer, which gives an infrared "fingerprint" spectrum characteristic of the additive polymer.

Evaluation of micro-colorimetric lipid determination method with samples prepared using sonication and accelerated solvent extraction methods

EPA Science Inventory

Two common laboratory extraction techniques were evaluated for routine use with the micro-colorimetric lipid determination method developed by Van Handel (1985) [E. Van Handel, J. Am. Mosq. Control Assoc. 1(1985) 302] and recently validated for small samples by Inouye and Lotufo ...

Comparison of methods of extracting messenger Ribonucleic Acid from ejaculated Porcine (Sus Scrofa) Spermatozoa

USDA-ARS?s Scientific Manuscript database

H. D. Guthrie, G.R. Welch, and L. A. Blomberg. Comparison of Methods of Extracting Messenger Ribonucleic Acid from Ejaculated Porcine (Sus Scrofa) Spermatozoa. Biotechnology and Germplasm Laboratory, Agricultural Research Service U. S. Department of Agriculture, Beltsville, MD 20705 The purpos...

Validation of a PCR-based method for the detection of various rendered materials in feedstuffs using a forensic DNA extraction kit.

PubMed

Myers, Michael J; Yancy, Haile F; Araneta, Michael; Armour, Jennifer; Derr, Janice; Hoostelaere, Lawrence A D; Farmer, Doris; Jackson, Falana; Kiessling, William M; Koch, Henry; Lin, Huahua; Liu, Yan; Mowlds, Gabrielle; Pinero, David; Riter, Ken L; Sedwick, John; Shen, Yuelian; Wetherington, June; Younkins, Ronsha

2006-01-01

A method trial was initiated to validate the use of a commercial DNA forensic kit to extract DNA from animal feed as part of a PCR-based method. Four different PCR primer pairs (one bovine pair, one porcine pair, one ovine primer pair, and one multispecies pair) were also evaluated. Each laboratory was required to analyze a total of 120 dairy feed samples either not fortified (control, true negative) or fortified with bovine meat and bone meal, porcine meat and bone meal (PMBM), or lamb meal. Feeds were fortified with the animal meals at a concentration of 0.1% (wt/wt). Ten laboratories participated in this trial, and each laboratory was required to evaluate two different primer pairs, i.e., each PCR primer pair was evaluated by five different laboratories. The method was considered to be validated for a given animal source when three or more laboratories achieved at least 97% accuracy (29 correct of 30 samples for 96.7% accuracy, rounded up to 97%) in detecting the fortified samples for that source. Using this criterion, the method was validated for the bovine primer because three laboratories met the criterion, with an average accuracy of 98.9%. The average false-positive rate was 3.0% in these laboratories. A fourth laboratory was 80% accurate in identifying the samples fortified with bovine meat and bone meal. A fifth laboratory was not able to consistently extract the DNA from the feed samples and did not achieve the criterion for accuracy for either the bovine or multispecies PCR primers. For the porcine primers, the method was validated, with four laboratories meeting the criterion for accuracy with an average accuracy of 99.2%. The fifth laboratory had a 93.3% accuracy outcome for the porcine primer. Collectively, these five laboratories had a 1.3% false-positive rate for the porcine primer. No laboratory was able to meet the criterion for accuracy with the ovine primers, most likely because of problems with the synthesis of the primer pair; none of the positive control DNA samples could be detected with the ovine primers. The multispecies primer pair was validated in three laboratories for use with bovine meat and bone meal and lamb meal but not with PMBM. The three laboratories had an average accuracy of 98.9% for bovine meat and bone meal, 97.8% for lamb meal, and 63.3% for PMBM. When examined on an individual laboratory basis, one of these four laboratories could not identify a single feed sample containing PMBM by using the multispecies primer, whereas the other laboratory identified only one PMBM-fortified sample, suggesting that the limit of detection for PMBM with this primer pair is around 0.1% (wt/wt). The results of this study demonstrated that the DNA forensic kit can be used to extract DNA from animal feed, which can then be used for PCR analysis to detect animal-derived protein present in the feed sample.

Determination of emamectin benzoate in medicated fish feed: a multisite study.

PubMed

Farer, Leslie J

2005-01-01

A new method was developed for the quantitation of emamectin benzoate in medicated fish feed at levels between 1 and 30 ppm. The new procedure, based on a previously reported assay, consists of a wet methanolic extraction of ground feed, followed by solid-phase extraction and injection onto a gradient liquid chromatographic system. A multisite study involving 3 laboratories (the developing laboratory and 2 independent laboratories) was performed to evaluate precision, recovery, linearity, and sensitivity. Mean recove;ries for triplicate analyses at 3 levels, performed by 2 analysts per laboratory, were between 89 and 97%, with coefficients of variation ranging from 1.6 to 8.6%. Coefficients of determination (r2) obtained from the plotted data were > or =0.993. The precision of the method, determined from 6 replicate preparations from the same batch of medicated feed assayed in 3 separate trials per laboratory, was between 0.6 and 5.8%. The quantitation limit was established at 0.5 ppm. Specificity and robustness studies were performed by the developing laboratory.

Methods of Analysis by the U.S. Geological Survey National Water Quality Laboratory - Determination of Moderate-Use Pesticides and Selected Degradates in Water by C-18 Solid-Phase Extraction and Gas Chromatography/Mass Spectrometry

USGS Publications Warehouse

Sandstrom, Mark W.; Stroppel, Max E.; Foreman, William T.; Schroeder, Michael P.

2001-01-01

A method for the isolation and analysis of 21 parent pesticides and 20 pesticide degradates in natural-water samples is described. Water samples are filtered to remove suspended particulate matter and then are pumped through disposable solid-phase-extraction columns that contain octadecyl-bonded porous silica to extract the analytes. The columns are dried by using nitrogen gas, and adsorbed analytes are eluted with ethyl acetate. Extracted analytes are determined by capillary-column gas chromatography/mass spectrometry with selected-ion monitoring of three characteristic ions. The upper concentration limit is 2 micrograms per liter (?g/L) for most analytes. Single-operator method detection limits in reagent-water samples range from 0.00 1 to 0.057 ?g/L. Validation data also are presented for 14 parent pesticides and 20 degradates that were determined to have greater bias or variability, or shorter holding times than the other compounds. The estimated maximum holding time for analytes in pesticide-grade water before extraction was 4 days. The estimated maximum holding time for analytes after extraction on the dry solid-phase-extraction columns was 7 days. An optional on-site extraction procedure allows for samples to be collected and processed at remote sites where it is difficult to ship samples to the laboratory within the recommended pre-extraction holding time. The method complements existing U.S. Geological Survey Method O-1126-95 (NWQL Schedules 2001 and 2010) by using identical sample preparation and comparable instrument analytical conditions so that sample extracts can be analyzed by either method to expand the range of analytes determined from one water sample.

Single Laboratory Validated Method for Determination of Microcystins and Nodularin in Ambient Freshwaters by Solid Phase Extraction and Liquid Chromatography/ Tandem Mass Spectrometry (LC/MS/MS)

EPA Pesticide Factsheets

This document is a standardized, single laboratory validated liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for the detection of cyanotoxins—microsystins and nodularin (combined intracellular and extracellular)—in ambient freshwaters.

Inter-laboratory variation in the chemical analysis of acidic forest soil reference samples from eastern North America

USGS Publications Warehouse

Ross, Donald S.; Bailiey, Scott W; Briggs, Russell D; Curry, Johanna; Fernandez, Ivan J.; Fredriksen, Guinevere; Goodale, Christine L.; Hazlett, Paul W.; Heine, Paul R; Johnson, Chris E.; Larson, John T; Lawrence, Gregory B.; Kolka, Randy K; Ouimet, Rock; Pare, D; Richter, Daniel D.; Shirmer, Charles D; Warby, Richard A.F.

2015-01-01

Long-term forest soil monitoring and research often requires a comparison of laboratory data generated at different times and in different laboratories. Quantifying the uncertainty associated with these analyses is necessary to assess temporal changes in soil properties. Forest soil chemical properties, and methods to measure these properties, often differ from agronomic and horticultural soils. Soil proficiency programs do not generally include forest soil samples that are highly acidic, high in extractable Al, low in extractable Ca and often high in carbon. To determine the uncertainty associated with specific analytical methods for forest soils, we collected and distributed samples from two soil horizons (Oa and Bs) to 15 laboratories in the eastern United States and Canada. Soil properties measured included total organic carbon and nitrogen, pH and exchangeable cations. Overall, results were consistent despite some differences in methodology. We calculated the median absolute deviation (MAD) for each measurement and considered the acceptable range to be the median 6 2.5 3 MAD. Variability among laboratories was usually as low as the typical variability within a laboratory. A few areas of concern include a lack of consistency in the measurement and expression of results on a dry weight basis, relatively high variability in the C/N ratio in the Bs horizon, challenges associated with determining exchangeable cations at concentrations near the lower reporting range of some laboratories and the operationally defined nature of aluminum extractability. Recommendations include a continuation of reference forest soil exchange programs to quantify the uncertainty associated with these analyses in conjunction with ongoing efforts to review and standardize laboratory methods.

Filtration Isolation of Nucleic Acids: A Simple and Rapid DNA Extraction Method.

PubMed

McFall, Sally M; Neto, Mário F; Reed, Jennifer L; Wagner, Robin L

2016-08-06

FINA, filtration isolation of nucleic acids, is a novel extraction method which utilizes vertical filtration via a separation membrane and absorbent pad to extract cellular DNA from whole blood in less than 2 min. The blood specimen is treated with detergent, mixed briefly and applied by pipet to the separation membrane. The lysate wicks into the blotting pad due to capillary action, capturing the genomic DNA on the surface of the separation membrane. The extracted DNA is retained on the membrane during a simple wash step wherein PCR inhibitors are wicked into the absorbent blotting pad. The membrane containing the entrapped DNA is then added to the PCR reaction without further purification. This simple method does not require laboratory equipment and can be easily implemented with inexpensive laboratory supplies. Here we describe a protocol for highly sensitive detection and quantitation of HIV-1 proviral DNA from 100 µl whole blood as a model for early infant diagnosis of HIV that could readily be adapted to other genetic targets.

Rapid method for direct identification of bacteria in urine and blood culture samples by matrix-assisted laser desorption ionization time-of-flight mass spectrometry: intact cell vs. extraction method.

PubMed

Ferreira, L; Sánchez-Juanes, F; Muñoz-Bellido, J L; González-Buitrago, J M

2011-07-01

Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a fast and reliable technology for the identification of microorganisms with proteomics approaches. Here, we compare an intact cell method and a protein extraction method before application on the MALDI plate for the direct identification of microorganisms in both urine and blood culture samples from clinical microbiology laboratories. The results show that the intact cell method provides excellent results for urine and is a good initial method for blood cultures. The extraction method complements the intact cell method, improving microorganism identification from blood culture. Thus, we consider that MALDI-TOF MS performed directly on urine and blood culture samples, with the protocols that we propose, is a suitable technique for microorganism identification, as compared with the routine methods used in the clinical microbiology laboratory. © 2010 The Authors. Clinical Microbiology and Infection © 2010 European Society of Clinical Microbiology and Infectious Diseases.

Extraction of Maltol from Fraser Fir: A Comparison of Microwave-Assisted Extraction and Conventional Heating Protocols for the Organic Chemistry Laboratory

ERIC Educational Resources Information Center

Koch, Andrew S.; Chimento, Clio A.; Berg, Allison N.; Mughal, Farah D.; Spencer, Jean-Paul; Hovland, Douglas E.; Mbadugha, Bessie; Hovland, Allan K.; Eller, Leah R.

2015-01-01

Two methods for the extraction of maltol from Fraser fir needles are performed and compared in this two-week experiment. A traditional benchtop extraction using dichloromethane is compared to a microwave-assisted extraction using aqueous ethanol. Students perform both procedures and weigh the merits of each technique. In doing so, students see a…

Flotation of Mineral and Dyes: A Laboratory Experiment for Separation Method Molecular Hitchhikers

ERIC Educational Resources Information Center

Rappon, Tim; Sylvestre, Jarrett A.; Rappon, Manit

2016-01-01

Flotation as a method of separation is widely researched and is applied in many industries. It has been used to address a wide range of environmental issues including treatment of wastewater, recovery of heavy metals for recycling, extraction of minerals in mining, and so forth. This laboratory attempts to show how such a simple method can be used…

Natural colorants: Pigment stability and extraction yield enhancement via utilization of appropriate pretreatment and extraction methods.

PubMed

Ngamwonglumlert, Luxsika; Devahastin, Sakamon; Chiewchan, Naphaporn

2017-10-13

Natural colorants from plant-based materials have gained increasing popularity due to health consciousness of consumers. Among the many steps involved in the production of natural colorants, pigment extraction is one of the most important. Soxhlet extraction, maceration, and hydrodistillation are conventional methods that have been widely used in industry and laboratory for such a purpose. Recently, various non-conventional methods, such as supercritical fluid extraction, pressurized liquid extraction, microwave-assisted extraction, ultrasound-assisted extraction, pulsed-electric field extraction, and enzyme-assisted extraction have emerged as alternatives to conventional methods due to the advantages of the former in terms of smaller solvent consumption, shorter extraction time, and more environment-friendliness. Prior to the extraction step, pretreatment of plant materials to enhance the stability of natural pigments is another important step that must be carefully taken care of. In this paper, a comprehensive review of appropriate pretreatment and extraction methods for chlorophylls, carotenoids, betalains, and anthocyanins, which are major classes of plant pigments, is provided by using pigment stability and extraction yield as assessment criteria.

An Improved Method for Extraction and Separation of Photosynthetic Pigments

ERIC Educational Resources Information Center

Katayama, Nobuyasu; Kanaizuka, Yasuhiro; Sudarmi, Rini; Yokohama, Yasutsugu

2003-01-01

The method for extracting and separating hydrophobic photosynthetic pigments proposed by Katayama "et al." ("Japanese Journal of Phycology," 42, 71-77, 1994) has been improved to introduce it to student laboratories at the senior high school level. Silica gel powder was used for removing water from fresh materials prior to…

Alternative method of removing otoliths from sturgeon

USGS Publications Warehouse

Chalupnicki, Marc A.; Dittman, Dawn E.

2016-01-01

Extracting the otoliths (ear bones) from fish that have very thick skulls can be difficult and very time consuming. The common practice of making a transverse vertical incision on the top of the skull with a hand or electrical saw may damage the otolith if not performed correctly. Sturgeons (Acipenseridae) are one family in particular that have a very large and thick skull. A new laboratory method entering the brain cavity from the ventral side of the fish to expose the otoliths was easier than other otolith extraction methods found in the literature. Methods reviewed in the literature are designed for the field and are more efficient at processing large quantities of fish quickly. However, this new technique was designed to be more suited for a laboratory setting when time is not pressing and successful extraction from each specimen is critical. The success of finding and removing otoliths using this technique is very high and does not compromise the structure in any manner. This alternative technique is applicable to other similar fish species for extracting the otoliths.

Application of solid/liquid extraction for the gravimetric determination of lipids in royal jelly.

PubMed

Antinelli, Jean-François; Davico, Renée; Rognone, Catherine; Faucon, Jean-Paul; Lizzani-Cuvelier, Louisette

2002-04-10

Gravimetric lipid determination is a major parameter for the characterization and the authentication of royal jelly quality. A solid/liquid extraction was compared to the reference method, which is based on liquid/liquid extraction. The amount of royal jelly and the time of the extraction were optimized in comparison to the reference method. Boiling/rinsing ratio and spread of royal jelly onto the extraction thimble were identified as critical parameters, resulting in good accuracy and precision for the alternative method. Comparison of reproducibility and repeatability of both methods associated with gas chromatographic analysis of the composition of the extracted lipids showed no differences between the two methods. As the intra-laboratory validation tests were comparable to the reference method, while offering rapidity and a decrease in amount of solvent used, it was concluded that the proposed method should be used with no modification of quality criteria and norms established for royal jelly characterization.

Measurement of bromate in bread by liquid chromatography with post-column flow reactor detection.

PubMed

Himata, K; Noda, M; Ando, S; Yamada, Y

2000-01-01

This method is suitable for the determination of bromate residues in a variety of baked goods. The peer-verified method trial was performed on white bread, multigrain bread, and coffee cake spiked with known levels of potassium bromate. The analytical portion is extracted with deionized water to remove bromate from the bulk of the baked product. The aqueous extract is carried through a series of steps to remove co-extractives that would interfere with the liquid chromatography (LC) in the determinative step or hasten the deterioration of the LC column. The extract is filtered before passing it through a reversed-phase solid-phase extraction (SPE) column and a cation-exchange column in the silver form to remove lipids and chloride, respectively. Ultrafiltration is then used to remove proteins with molecular weights of >30,000 daltons. Finally, a cation-exchange column in the sodium form is used to remove silver ions from the extract. The determinative step uses LC with a reversed-phase column and an ion-pairing agent in the mobile phase. Detection is based on the post-column reaction of bromate with o-dianisidine to form an oxidation product that is quantitated spectrophotometrically at 450 nm. Overall agreement between the submitting and peer laboratories was quite good. For bromate levels of 10-52 ppb, overall mean recoveries were 76.9 and 78.8% for the submitting and peer laboratories, respectively. The standard deviations were higher for the results of the peer laboratory, probably because of the generally higher level of baseline noise present in the chromatograms. The results demonstrate that the method provides adequate accuracy with low-fat as well as high-fat foods. Bromate at levels as low as 5 ppb (ng/g) can be detected with the method.

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

Campbell, J.A.; Clauss, S.A.; Grant, K.E.

The objectives of this task are to develop and document extraction and analysis methods for organics in waste tanks, and to extend these methods to the analysis of actual core samples to support the Waste Tank organic Safety Program. This report documents progress at Pacific Northwest Laboratory (a) during FY 1994 on methods development, the analysis of waste from Tank 241-C-103 (Tank C-103) and T-111, and the transfer of documented, developed analytical methods to personnel in the Analytical Chemistry Laboratory (ACL) and 222-S laboratory. This report is intended as an annual report, not a completed work.

Determination of catechins and caffeine in camillia sinensis raw materials, extracts, and dietary supplements by HPLC-uv: single-laboratory validation.

PubMed

Roman, Mark C

2013-01-01

A rapid method has been developed to quantify seven catechins and caffeine in green tea (Camillia sinensis) raw material and powdered extract, and dietary supplements containing green tea extract. The method utilizes RP HPLC with a phenyl-based stationary phase and gradient elution. Detection is by UV absorbance. The total run time, including column re-equilibration, is 13 min. Single-laboratory validation (SLV) has been performed on the method to determine the repeatability, accuracy, selectivity, LOD, LOQ, ruggedness, and linearity for (+)-catechin, (-)-epicatechin, (-)-epicatechin gallate, (-)-epigallocatechin, (-)-gallocatechin gallate, (-)-epigallocatechin gallate, and (+)-gallocatechin, as well as caffeine. Repeatability precision and recovery results met AOAC guidelines for SLV studies for all catechins and caffeine down to a level of approximately 20 mg/g. Finished products containing high concentrations of minerals require the use of EDTA to prevent decomposition of the catechins.

Rice hull smoke extract inactivates Salmonella Typhimurium in laboratory media and protects infected mice against mortality

USDA-ARS?s Scientific Manuscript database

A recently discovered and characterized rice hull liquid smoke extract was tested for bactericidal activity against Salmonella Typhimurium using the disc-agar method. The Minimum Inhibitory Concentration (MIC) value of rice hull smoke extract was found to be 0.822% (v/v). The in vivo antibacterial a...

METHOD 544. DETERMINATION OF MICROCYSTINS AND ...

EPA Pesticide Factsheets

Method 544 is an accurate and precise analytical method to determine six microcystins (including MC-LR) and nodularin in drinking water using solid phase extraction and liquid chromatography tandem mass spectrometry (SPE-LC/MS/MS). The advantage of this SPE-LC/MS/MS is its sensitivity and ability to speciate the microcystins. This method development task establishes sample preservation techniques, sample concentration and analytical procedures, aqueous and extract holding time criteria and quality control procedures. Draft Method 544 undergone a multi-laboratory verification to ensure other laboratories can implement the method and achieve the quality control measures specified in the method. It is anticipated that Method 544 may be used in UCMR 4 to collect nationwide occurrence data for selected microcystins in drinking water. The purpose of this research project is to develop an accurate and precise analytical method to concentrate and determine selected MCs and nodularin in drinking water.

Viability testing of material derived from Mycobacterium tuberculosis prior to removal from a Containment Level-III Laboratory as part of a Laboratory Risk Assessment Program

PubMed Central

Blackwood, Kym S; Burdz, Tamara V; Turenne, Christine Y; Sharma, Meenu K; Kabani, Amin M; Wolfe, Joyce N

2005-01-01

Background In the field of clinical mycobacteriology, Mycobacterium tuberculosis (MTB) can be a difficult organism to manipulate due to the restrictive environment of a containment level 3 (CL3) laboratory. Tests for rapid diagnostic work involving smears and molecular methods do not require CL3 practices after the organism has been rendered non-viable. While it has been assumed that after organism deactivation these techniques can be performed outside of a CL3, no conclusive study has consistently confirmed that the organisms are noninfectious after the theoretical 'deactivation' steps. Previous studies have shown that initial steps (such as heating /chemical fixation) may not consistently kill MTB organisms. Methods An inclusive viability study (n = 226) was undertaken to determine at which point handling of culture extraction materials does not necessitate a CL3 environment. Four different laboratory protocols tested for viability included: standard DNA extractions for IS6110 fingerprinting, crude DNA preparations for PCR by boiling and mechanical lysis, protein extractions, and smear preparations. For each protocol, laboratory staff planted a proportion of the resulting material to Bactec 12B medium that was observed for growth for 8 weeks. Results Of the 208 isolates initially tested, 21 samples grew within the 8-week period. Sixteen (7.7%) of these yielded positive results for MTB that included samples of: deactivated culture resuspensions exposed to 80°C for 20 minutes, smear preparations and protein extractions. Test procedures were consequently modified and tested again (n = 18), resulting in 0% viability. Conclusions This study demonstrates that it cannot be assumed that conventional practices (i.e. smear preparation) or extraction techniques render the organism non-viable. All methodologies, new and existing, should be examined by individual laboratories to validate the safe removal of material derived from MTB to the outside of a CL3 laboratory. This process is vital to establish in house biosafety-validated practices with the aim of protecting laboratory workers conducting these procedures. PMID:15667662

Validation of a standardized extraction method for formalin-fixed paraffin-embedded tissue samples.

PubMed

Lagheden, Camilla; Eklund, Carina; Kleppe, Sara Nordqvist; Unger, Elizabeth R; Dillner, Joakim; Sundström, Karin

2016-07-01

Formalin-fixed paraffin-embedded (FFPE) samples can be DNA-extracted and used for human papillomavirus (HPV) genotyping. The xylene-based gold standard for extracting FFPE samples is laborious, suboptimal and involves health hazards for the personnel involved. To compare extraction with the standard xylene method to a xylene-free method used in an HPV LabNet Global Reference Laboratory at the Centers for Disease Control (CDC); based on a commercial method with an extra heating step. Fifty FFPE samples were randomly selected from a national audit of all cervical cancer cases diagnosed in Sweden during 10 years. For each case-block, a blank-block was sectioned, as a control for contamination. For xylene extraction, the standard WHO Laboratory Manual protocol was used. For the CDC method, the manufacturers' protocol was followed except for an extra heating step, 120°C for 20min. Samples were extracted and tested in parallel with β-globin real-time PCR, HPV16 real-time PCR and HPV typing using modified general primers (MGP)-PCR and Luminex assays. For a valid result the blank-block had to be betaglobin-negative in all tests and the case-block positive for beta-globin. Overall, detection was improved with the heating method and the amount of HPV-positive samples increased from 70% to 86% (p=0.039). For all samples where HPV type concordance could be evaluated, there was 100% type concordance. A xylene-free and robust extraction method for HPV-DNA typing in FFPE material is currently in great demand. Our proposed standardized protocol appears to be generally useful. Copyright © 2016. Published by Elsevier B.V.

Laboratory Production of Lemon Liqueur (Limoncello) by Conventional Maceration and a Two-Syringe System to Illustrate Rapid Solid-Liquid Dynamic Extraction

ERIC Educational Resources Information Center

Naviglio, Daniele; Montesano, Domenico; Gallo, Monica

2015-01-01

Two experimental techniques of solid-liquid extraction are compared relating to the lab-scale production of lemon liqueur, most commonly named "limoncello"; the first is the official method of maceration for the solid-liquid extraction of analytes and is widely used to extract active ingredients from a great variety of natural products;…

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of pesticides in water by Carbopak-B solid-phase extraction and high-preformance liquid chromatography

USGS Publications Warehouse

Werner, Stephen L.; Burkhardt, Mark R.; DeRusseau, Sabrina N.

1996-01-01

In accordance with the needs of the National Water-Quality Assessment Program (NAWQA), the U.S. Geological Survey has developed and implemented a graphitized carbon-based solid-phase extraction and high-performance liquid chromatographic analytical method. The method is used to determine 41 pesticides and pesticide metabolites that are not readily amenable to gas chromatography or other high-temperature analytical techniques. Pesticides are extracted from filtered environmental water samples using a 0.5-gram graphitized carbon-based solid-phase cartridge, eluted from the cartridge into two analytical fractions, and analyzed using high-performance liquid chromatography with photodiode-array detection. The upper concentration limit is 1.6 micrograms per liter (=B5g/L) for most compounds. Single-operator method detection limits in organic-free water samples ranged from 0.006 to 0.032 =B5g/L= Recoveries in organic-free water samples ranged from 37 to 88 percent. Recoveries in ground- and surface-water samples ranged from 29 to 94 percent. An optional on-site extraction procedure allows for samples to be collected and processed at remote sites where it is difficult to ship samples to the laboratory within the recommended pre-extraction holding time of 7 days.

Accelerated solvent extraction (ASE) for purification and extraction of silicone passive samplers used for the monitoring of organic pollutants.

PubMed

Brockmeyer, Berit; Kraus, Uta R; Theobald, Norbert

2015-12-01

Silicone passive samplers have gained an increasing attention as single-phased, practical and robust samplers for monitoring of organic contaminants in the aquatic environment in recent years. However, analytical challenges arise in routine application during the extraction of analytes as silicone oligomers are co-extracted and interfere severely during chemical analyses (e.g. gas chromatographic techniques). In this study, we present a fast, practical pre-cleaning method for silicone passive samplers applying accelerated solvent extraction (ASE) for the removal of silicone oligomers prior to the water deployment (hexane/dichloromethane, 100 °C, 70 min). ASE was also shown to be a very fast (10 min) and efficient extraction method for non-polar contaminants (non-exposed PRC recoveries 66-101 %) sampled by the silicone membrane. For both applications, temperature, extraction time and the solvent used for ASE have been optimized. Purification of the ASE extract was carried out by silica gel and high-pressure liquid size exclusion chromatography (HPLC-SEC). The silicone oligomer content was checked by total reflection X-ray fluorescence spectroscopy (TXRF) in order to confirm the absence of the silicone oligomers prior to analysis of passive sampler extracts. The established method was applied on real silicone samplers from the North- and Baltic Sea and showed no matrix effects during analysis of organic pollutants. Internal laboratory standard recoveries were in the same range for laboratory, transport and exposed samplers (85-126 %).

Use of FTA gene guard filter paper for the storage and transportation of tumor cells for molecular testing.

PubMed

Dobbs, Larry J; Madigan, Merle N; Carter, Alexis B; Earls, Lori

2002-01-01

Efficient methods of storing tumor specimens for molecular testing are needed in the modern surgical pathology laboratory. The FTA Gene Guard system is a novel method for the collection and room temperature storage of blood samples for DNA testing. The method uses index card-sized filter papers that provide an ideal medium on which to store tumor specimens for DNA testing. To determine whether FTA filter paper can be used in the surgical pathology laboratory to store tumor cells for DNA testing. Cell suspensions were prepared from 60 surgical specimens, and DNA was extracted either immediately or after storage on FTA paper. The DNA extracted by each method was tested by polymerase chain reaction (PCR) for the beta-globin and interferon gamma genes, and the results were compared. Fifteen lymph node specimens stored on FTA paper were then tested for immunoglobulin heavy chain (IgH) gene rearrangement by PCR, and these results were compared with those obtained for immediately extracted DNA. University medical center. The DNA extracted from cells stored on FTA paper performed as well in the PCR as the freshly extracted DNA in nearly all cases (>95%). The results of tests for IgH gene rearrangements showed 100% concordance between the 2 methods of DNA extraction.Conclusion.-Cells from surgical specimens can be stored on FTA paper for extended lengths of time, and DNA can be extracted from these cells for PCR-based testing. FTA filter paper is a reliable medium for the storage and/or transport of tumor cells for PCR-based DNA analysis.

Direct PCR amplification of forensic touch and other challenging DNA samples: A review.

PubMed

Cavanaugh, Sarah E; Bathrick, Abigail S

2018-01-01

DNA evidence sample processing typically involves DNA extraction, quantification, and STR amplification; however, DNA loss can occur at both the DNA extraction and quantification steps, which is not ideal for forensic evidence containing low levels of DNA. Direct PCR amplification of forensic unknown samples has been suggested as a means to circumvent extraction and quantification, thereby retaining the DNA typically lost during those procedures. Direct PCR amplification is a method in which a sample is added directly to an amplification reaction without being subjected to prior DNA extraction, purification, or quantification. It allows for maximum quantities of DNA to be targeted, minimizes opportunities for error and contamination, and reduces the time and monetary resources required to process samples, although data analysis may take longer as the increased DNA detection sensitivity of direct PCR may lead to more instances of complex mixtures. ISO 17025 accredited laboratories have successfully implemented direct PCR for limited purposes (e.g., high-throughput databanking analysis), and recent studies indicate that direct PCR can be an effective method for processing low-yield evidence samples. Despite its benefits, direct PCR has yet to be widely implemented across laboratories for the processing of evidentiary items. While forensic DNA laboratories are always interested in new methods that will maximize the quantity and quality of genetic information obtained from evidentiary items, there is often a lag between the advent of useful methodologies and their integration into laboratories. Delayed implementation of direct PCR of evidentiary items can be attributed to a variety of factors, including regulatory guidelines that prevent laboratories from omitting the quantification step when processing forensic unknown samples, as is the case in the United States, and, more broadly, a reluctance to validate a technique that is not widely used for evidence samples. The advantages of direct PCR of forensic evidentiary samples justify a re-examination of the factors that have delayed widespread implementation of this method and of the evidence supporting its use. In this review, the current and potential future uses of direct PCR in forensic DNA laboratories are summarized. Copyright © 2017 Elsevier B.V. All rights reserved.

Swab Protocol for Rapid Laboratory Diagnosis of Cutaneous Anthrax

PubMed Central

Marston, Chung K.; Bhullar, Vinod; Baker, Daniel; Rahman, Mahmudur; Hossain, M. Jahangir; Chakraborty, Apurba; Khan, Salah Uddin; Hoffmaster, Alex R.

2012-01-01

The clinical laboratory diagnosis of cutaneous anthrax is generally established by conventional microbiological methods, such as culture and directly straining smears of clinical specimens. However, these methods rely on recovery of viable Bacillus anthracis cells from swabs of cutaneous lesions and often yield negative results. This study developed a rapid protocol for detection of B. anthracis on clinical swabs. Three types of swabs, flocked-nylon, rayon, and polyester, were evaluated by 3 extraction methods, the swab extraction tube system (SETS), sonication, and vortex. Swabs were spiked with virulent B. anthracis cells, and the methods were compared for their efficiency over time by culture and real-time PCR. Viability testing indicated that the SETS yielded greater recovery of B. anthracis from 1-day-old swabs; however, reduced viability was consistent for the 3 extraction methods after 7 days and nonviability was consistent by 28 days. Real-time PCR analysis showed that the PCR amplification was not impacted by time for any swab extraction method and that the SETS method provided the lowest limit of detection. When evaluated using lesion swabs from cutaneous anthrax outbreaks, the SETS yielded culture-negative, PCR-positive results. This study demonstrated that swab extraction methods differ in their efficiency of recovery of viable B. anthracis cells. Furthermore, the results indicated that culture is not reliable for isolation of B. anthracis from swabs at ≥7 days. Thus, we recommend the use of the SETS method with subsequent testing by culture and real-time PCR for diagnosis of cutaneous anthrax from clinical swabs of cutaneous lesions. PMID:23035192

Proficiency program for real-time PCR diagnosis of Bordetella pertussis infections in French hospital laboratories and at the French National Reference Center for Whooping Cough and other Bordetelloses.

PubMed

Caro, Valérie; Guiso, Nicole; Alberti, Corinne; Liguori, Sandrine; Burucoa, Christophe; Couetdic, Gérard; Doucet-Populaire, Florence; Ferroni, Agnès; Papin-Gibaud, Sophie; Grattard, Florence; Réglier-Poupet, Hélène; Raymond, Josette; Soler, Catherine; Bouchet, Sylvie; Charreau, Sandrine; Couzon, Brigitte; Leymarie, Isabelle; Tavares, Nicole; Choux, Mathilde; Bingen, Edouard; Bonacorsi, Stéphane

2009-10-01

With the support of a ministerial program for innovative and expensive technologies, dedicated to the economic evaluation of laboratory diagnosis of pertussis by real-time PCR, external quality assessment for real-time IS481 PCR was carried out. Coordinated by the National Centre of Reference of Pertussis and other Bordetelloses (NCR), this study aimed to harmonize and to assess the performances of eight participating microbiology hospital laboratories throughout the French territory. Between January 2006 and February 2007, 10 proficiency panels were sent by the NCR (ascending proficiency program), representing a total of 49 samples and including eight panels to analyze and evaluate the global sensitivity and specificity of real-time PCR, one to assess the limit of detection, and one to evaluate nucleic acid extraction methods. As part of the descending proficiency program, extracted DNA from clinical samples was sent by the eight participating laboratories in different panels and analyzed by the NCR. In the ascending proficiency analysis, the sensitivity and specificity of the real-time PCR methods were 92.2% and 94.3%, respectively. The limit of detection of the different methods ranged between 0.1 and 1 fg/microl (0.2 to 2 CFU/microl). The nucleic acid extraction methods showed similar performances. During the descending proficiency analysis, performed with 126 samples, the result of the NCR for 15 samples (11.9%) was discordant with the result obtained by the source laboratory. Despite several initial differences, harmonization was easy and performances were homogeneous. However, the risk of false-positive results remains quite high, and we strongly recommend establishment of uniform quality control procedures performed regularly.

Evaluation of methods for the extraction of DNA from drinking water distribution system biofilms.

PubMed

Hwang, Chiachi; Ling, Fangqiong; Andersen, Gary L; LeChevallier, Mark W; Liu, Wen-Tso

2012-01-01

While drinking water biofilms have been characterized in various drinking water distribution systems (DWDS), little is known about the impact of different DNA extraction methods on the subsequent analysis of microbial communities in drinking water biofilms. Since different DNA extraction methods have been shown to affect the outcome of microbial community analysis in other environments, it is necessary to select a DNA extraction method prior to the application of molecular tools to characterize the complex microbial ecology of the DWDS. This study compared the quantity and quality of DNA yields from selected DWDS bacteria with different cell wall properties using five widely used DNA extraction methods. These were further selected and evaluated for their efficiency and reproducibility of DNA extraction from DWDS samples. Terminal restriction fragment length analysis and the 454 pyrosequencing technique were used to interpret the differences in microbial community structure and composition, respectively, from extracted DNA. Such assessments serve as a concrete step towards the determination of an optimal DNA extraction method for drinking water biofilms, which can then provide a reliable comparison of the meta-analysis results obtained in different laboratories.

OPTIMIZED DETERMINATION OF TRACE JET FUEL VOLATILE ORGANIC COMPOUNDS IN HUMAN BLOOD USING IN-FIELD LIQUID-LIQUID EXTRACTION WITH SUBSEQUENT LABORATORY GAS CHROMATOGRAPHIC-MASS SPECTROMETRIC ANALYSIS AND ON-COLUMN LARGE VOLUME INJECTION

EPA Science Inventory

A practical and sensitive method to assess volatile organic compounds (VOCs) from JP-8 jet fuel in human whole blood was developed by modifying previously established liquid-liquid extraction procedures, optimizing extraction times, solvent volume, specific sample processing te...

Application of the BioMek 2000 Laboratory Automation Workstation and the DNA IQ System to the extraction of forensic casework samples.

PubMed

Greenspoon, Susan A; Ban, Jeffrey D; Sykes, Karen; Ballard, Elizabeth J; Edler, Shelley S; Baisden, Melissa; Covington, Brian L

2004-01-01

Robotic systems are commonly utilized for the extraction of database samples. However, the application of robotic extraction to forensic casework samples is a more daunting task. Such a system must be versatile enough to accommodate a wide range of samples that may contain greatly varying amounts of DNA, but it must also pose no more risk of contamination than the manual DNA extraction methods. This study demonstrates that the BioMek 2000 Laboratory Automation Workstation, used in combination with the DNA IQ System, is versatile enough to accommodate the wide range of samples typically encountered by a crime laboratory. The use of a silica coated paramagnetic resin, as with the DNA IQ System, facilitates the adaptation of an open well, hands off, robotic system to the extraction of casework samples since no filtration or centrifugation steps are needed. Moreover, the DNA remains tightly coupled to the silica coated paramagnetic resin for the entire process until the elution step. A short pre-extraction incubation step is necessary prior to loading samples onto the robot and it is at this step that most modifications are made to accommodate the different sample types and substrates commonly encountered with forensic evidentiary samples. Sexual assault (mixed stain) samples, cigarette butts, blood stains, buccal swabs, and various tissue samples were successfully extracted with the BioMek 2000 Laboratory Automation Workstation and the DNA IQ System, with no evidence of contamination throughout the extensive validation studies reported here.

Evaluation of mericon E. coli O157 Screen Plus and mericon E. coli STEC O-Type Pathogen Detection Assays in Select Foods: Collaborative Study, First Action 2017.05.

PubMed

Bird, Patrick; Benzinger, M Joseph; Bastin, Benjamin; Crowley, Erin; Agin, James; Goins, David; Armstrong, Marcia

2018-05-01

QIAGEN mericon Escherichia coli O157 Screen Plus and mericon E. coli Shiga toxin-producing E. coli (STEC) O-Type Pathogen Detection Assays use Real-Time PCR technology for the rapid, accurate detection of E. coli O157 and the "big six" (O26, O45, O103, O111, O121, O145) (non-O157 STEC) in select food types. Using a paired study design, the assays were compared with the U.S. Department of Agriculture, Food Safety Inspection Service Microbiology Laboratory Guidebook Chapter 5.09 reference method for the detection of E. coli O157:H7 in raw ground beef. Both mericon assays were evaluated using the manual and an automated DNA extraction method. Thirteen technicians from five laboratories located within the continental United States participated in the collaborative study. Three levels of contamination were evaluated. Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low-inoculum level test portions produced a difference between laboratories POD (dLPOD) value with a 95% confidence interval of 0.00 (-0.12, 0.12) for the mericon E. coli O157 Screen Plus with manual and automated extraction and mericon E. coli STEC O-Type with manual extraction and -0.01 (-0.13, 0.10) for the mericon E. coli STEC O-Type with automated extraction. The dLPOD results indicate equivalence between the candidate methods and the reference method.

Laboratory studies on the removal of radon-born lead from KamLAND's organic liquid scintillator

NASA Astrophysics Data System (ADS)

Keefer, G.; Grant, C.; Piepke, A.; Ebihara, T.; Ikeda, H.; Kishimoto, Y.; Kibe, Y.; Koseki, Y.; Ogawa, M.; Shirai, J.; Takeuchi, S.; Mauger, C.; Zhang, C.; Schweitzer, G.; Berger, B. E.; Dazeley, S.; Decowski, M. P.; Detwiler, J. A.; Djurcic, Z.; Dwyer, D. A.; Efremenko, Y.; Enomoto, S.; Freedman, S. J.; Fujikawa, B. K.; Furuno, K.; Gando, A.; Gando, Y.; Gratta, G.; Hatakeyama, S.; Heeger, K. M.; Hsu, L.; Ichimura, K.; Inoue, K.; Iwamoto, T.; Kamyshkov, Y.; Karwowski, H. J.; Koga, M.; Kozlov, A.; Lane, C. E.; Learned, J. G.; Maricic, J.; Markoff, D. M.; Matsuno, S.; McKee, D.; McKeown, R. D.; Miletic, T.; Mitsui, T.; Motoki, M.; Nakajima, Kyo; Nakajima, Kyohei; Nakamura, K.; O`Donnell, T.; Ogawa, H.; Piquemal, F.; Ricol, J.-S.; Shimizu, I.; Suekane, F.; Suzuki, A.; Svoboda, R.; Tajima, O.; Takemoto, Y.; Tamae, K.; Tolich, K.; Tornow, W.; Watanabe, Hideki; Watanabe, Hiroko; Winslow, L. A.; Yoshida, S.

2015-01-01

The removal of radioactivity from liquid scintillator has been studied in preparation of a low background phase of KamLAND. This paper describes the methods and techniques developed to measure and efficiently extract radon decay products from liquid scintillator. We report the radio-isotope reduction factors obtained when applying various extraction methods. During this study, distillation was identified as the most efficient method for removing radon-born lead from liquid scintillator.

Laboratory studies on the removal of radon-born lead from KamLAND׳s organic liquid scintillator

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

Keefer, G.; Grant, C.; Piepke, A.

We studied the removal of radioactivity from liquid scintillator in preparation of a low background phase of KamLAND. We describe the methods and techniques developed to measure and efficiently extract radon decay products from liquid scintillator. Lastly, we report the radio-isotope reduction factors obtained when applying various extraction methods. During this study, distillation was identified as the most efficient method for removing radon daughters from liquid scintillator.

Laboratory studies on the removal of radon-born lead from KamLAND׳s organic liquid scintillator

DOE PAGES

Keefer, G.; Grant, C.; Piepke, A.; ...

2014-09-28

We studied the removal of radioactivity from liquid scintillator in preparation of a low background phase of KamLAND. We describe the methods and techniques developed to measure and efficiently extract radon decay products from liquid scintillator. Lastly, we report the radio-isotope reduction factors obtained when applying various extraction methods. During this study, distillation was identified as the most efficient method for removing radon daughters from liquid scintillator.

The current role of on-line extraction approaches in clinical and forensic toxicology.

PubMed

Mueller, Daniel M

2014-08-01

In today's clinical and forensic toxicological laboratories, automation is of interest because of its ability to optimize processes, to reduce manual workload and handling errors and to minimize exposition to potentially infectious samples. Extraction is usually the most time-consuming step; therefore, automation of this step is reasonable. Currently, from the field of clinical and forensic toxicology, methods using the following on-line extraction techniques have been published: on-line solid-phase extraction, turbulent flow chromatography, solid-phase microextraction, microextraction by packed sorbent, single-drop microextraction and on-line desorption of dried blood spots. Most of these published methods are either single-analyte or multicomponent procedures; methods intended for systematic toxicological analysis are relatively scarce. However, the use of on-line extraction will certainly increase in the near future.

Pressure-Assisted Chelating Extraction as a Teaching Tool in Instrumental Analysis

ERIC Educational Resources Information Center

Sadik, Omowunmi A.; Wanekaya, Adam K.; Yevgeny, Gelfand

2004-01-01

A novel instrumental-digestion technique using pressure-assisted chelating extraction (PACE), for undergraduate laboratory is reported. This procedure is used for exposing students to safe sample-preparation techniques, for correlating wet-chemical methods with modern instrumental analysis and comparing the performance of PACE with conventional…

Analytical methods for characterization of explosives-contaminated sites on U.S. Army installations

NASA Astrophysics Data System (ADS)

Jenkins, Thomas F.; Walsh, Marianne E.; Thorne, Philip G.

1995-10-01

The U.S. Army manufactures munitions at facilities throughout the United States. Many of these facilities are contaminated with residues of explosives from production, disposal of off- specification, and out-of-data munitions. The first step in remediating these sites is careful characterization. Currently sites are being characterized using a combination of on-site field screening and off-site laboratory analysis. Most of the contamination is associated with TNT (2,4,6-trinitrotoluene) and RDX (hexahydro-1,3,5-tri-nitro-1,3,5-triazine) and their manufacturing impurities and environmental transformation products. Both colorimetric and enzyme immunoassay-based field screening methods have been used successfully for on-site characterization. These methods have similar detection capabilities but differ in their selectivity. Although field screening is very cost-effective, laboratory analysis is still required to fully characterize a site. Laboratory analysis for explosives residues in the United States is generally conducted using high-performance liquid chromatography equipped with a UV detector. Air-dried soils are extracted with acetonitrile in an ultrasonic bath. Water is analyzed directly if detection limits in the range of 10 - 20 (mu) g/L are acceptable, or preconcentrated using either salting-out solvent extraction with acetonitrile or solid phase extraction.

Fly Diversity Revealed by PCR-RFLP of Mitochondrial DNA

ERIC Educational Resources Information Center

Asraoui, Jimmy F.; Sayar, Nancy P.; Knio, Khouzama M.; Smith, Colin A.

2008-01-01

In this article, we describe an inexpensive, two-session undergraduate laboratory activity that introduces important molecular biology methods in the context of biodiversity. In the first session, students bring tentatively identified flies (order Diptera, true flies) to the laboratory, extract DNA, and amplify a region of the mitochondrial gene…

Guaiacol Peroxidase Zymography for the Undergraduate Laboratory

ERIC Educational Resources Information Center

Wilkesman, Jeff; Castro, Diana; Contreras, Lellys M.; Kurz, Liliana

2014-01-01

This laboratory exercise presents a novel way to introduce undergraduate students to the specific detection of enzymatic activity by electrophoresis. First, students prepare a crude peroxidase extract and then analyze the homogenate via electrophoresis. Zymography, that is, a SDS-PAGE method to detect enzyme activity, is used to specifically…

Research of facial feature extraction based on MMC

NASA Astrophysics Data System (ADS)

Xue, Donglin; Zhao, Jiufen; Tang, Qinhong; Shi, Shaokun

2017-07-01

Based on the maximum margin criterion (MMC), a new algorithm of statistically uncorrelated optimal discriminant vectors and a new algorithm of orthogonal optimal discriminant vectors for feature extraction were proposed. The purpose of the maximum margin criterion is to maximize the inter-class scatter while simultaneously minimizing the intra-class scatter after the projection. Compared with original MMC method and principal component analysis (PCA) method, the proposed methods are better in terms of reducing or eliminating the statistically correlation between features and improving recognition rate. The experiment results on Olivetti Research Laboratory (ORL) face database shows that the new feature extraction method of statistically uncorrelated maximum margin criterion (SUMMC) are better in terms of recognition rate and stability. Besides, the relations between maximum margin criterion and Fisher criterion for feature extraction were revealed.

An efficient field and laboratory workflow for plant phylotranscriptomic projects1

PubMed Central

Yang, Ya; Moore, Michael J.; Brockington, Samuel F.; Timoneda, Alfonso; Feng, Tao; Marx, Hannah E.; Walker, Joseph F.; Smith, Stephen A.

2017-01-01

Premise of the study: We describe a field and laboratory workflow developed for plant phylotranscriptomic projects that involves cryogenic tissue collection in the field, RNA extraction and quality control, and library preparation. We also make recommendations for sample curation. Methods and Results: A total of 216 frozen tissue samples of Caryophyllales and other angiosperm taxa were collected from the field or botanical gardens. RNA was extracted, stranded mRNA libraries were prepared, and libraries were sequenced on Illumina HiSeq platforms. These included difficult mucilaginous tissues such as those of Cactaceae and Droseraceae. Conclusions: Our workflow is not only cost effective (ca. $270 per sample, as of August 2016, from tissue to reads) and time efficient (less than 50 h for 10–12 samples including all laboratory work and sample curation), but also has proven robust for extraction of difficult samples such as tissues containing high levels of secondary compounds. PMID:28337391

Influence of extractives on bonding properties of white and southern red oak

Treesearch

Mon-Lin Kuo; Daniel DiCarlo; Chung-Yun Hse

1984-01-01

White and southern red oak veneer were subjected to four methods of drying followed by five surface treatments. The four drying methods were mill drying at 350°F, laboratory drying at 350 and 212°F, and air drying. The five surface treatments were no treatment, surface scraping, soaking and dipping in 1% NaOH aqueous solution, and water extraction. Plywood panels were...

Comparison of marine sampling methods for organic contaminants: Passive samplers, water extractions, and live oyster deployment.

PubMed

Raub, Kristin B; Vlahos, Penny; Whitney, Michael

2015-08-01

Laboratory and field trials evaluated the efficacy of three methods of detecting aquatic pesticide concentrations. Currently used pesticides: atrazine, metolachlor, and diazinon and legacy pesticide dieldrin were targeted. Pesticides were extracted using solid-phase extraction (SPE) of water samples, titanium plate passive samplers coated in ethylene vinyl acetate (EVA) and eastern oysters (Crassostrea viginica) as biosamplers. A laboratory study assessed the extraction efficiencies and precision of each method. Passive samplers yielded the highest precision of the three methods (RSD: 3-14% EVA plates; 19-60% oysters; and 25-56% water samples). Equilibrium partition coefficients were derived. A significant relationship was found between the concentration in oyster tissue and the ambient aquatic concentration. In the field (Housatonic River, CT (U.S.)) water sampling (n = 5) detected atrazine at 1.61-7.31 μg L(-1), oyster sampling (n = 2×15) detected dieldrin at n.d.-0.096 μg L(-1) SW and the passive samplers (n = 5×3) detected atrazine at 0.97-3.78 μg L(-1) SW and dieldrin at n.d.-0.68 μg L(-1) SW. Copyright © 2015 Elsevier Ltd. All rights reserved.

Rapid detection of potyviruses from crude plant extracts.

PubMed

Silva, Gonçalo; Oyekanmi, Joshua; Nkere, Chukwuemeka K; Bömer, Moritz; Kumar, P Lava; Seal, Susan E

2018-04-01

Potyviruses (genus Potyvirus; family Potyviridae) are widely distributed and represent one of the most economically important genera of plant viruses. Therefore, their accurate detection is a key factor in developing efficient control strategies. However, this can sometimes be problematic particularly in plant species containing high amounts of polysaccharides and polyphenols such as yam (Dioscorea spp.). Here, we report the development of a reliable, rapid and cost-effective detection method for the two most important potyviruses infecting yam based on reverse transcription-recombinase polymerase amplification (RT-RPA). The developed method, named 'Direct RT-RPA', detects each target virus directly from plant leaf extracts prepared with a simple and inexpensive extraction method avoiding laborious extraction of high-quality RNA. Direct RT-RPA enables the detection of virus-positive samples in under 30 min at a single low operation temperature (37 °C) without the need for any expensive instrumentation. The Direct RT-RPA tests constitute robust, accurate, sensitive and quick methods for detection of potyviruses from recalcitrant plant species. The minimal sample preparation requirements and the possibility of storing RPA reagents without cold chain storage, allow Direct RT-RPA to be adopted in minimally equipped laboratories and with potential use in plant clinic laboratories and seed certification facilities worldwide. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Evaluation of Methods for the Extraction of DNA from Drinking Water Distribution System Biofilms

PubMed Central

Hwang, Chiachi; Ling, Fangqiong; Andersen, Gary L.; LeChevallier, Mark W.; Liu, Wen-Tso

2012-01-01

While drinking water biofilms have been characterized in various drinking water distribution systems (DWDS), little is known about the impact of different DNA extraction methods on the subsequent analysis of microbial communities in drinking water biofilms. Since different DNA extraction methods have been shown to affect the outcome of microbial community analysis in other environments, it is necessary to select a DNA extraction method prior to the application of molecular tools to characterize the complex microbial ecology of the DWDS. This study compared the quantity and quality of DNA yields from selected DWDS bacteria with different cell wall properties using five widely used DNA extraction methods. These were further selected and evaluated for their efficiency and reproducibility of DNA extraction from DWDS samples. Terminal restriction fragment length analysis and the 454 pyrosequencing technique were used to interpret the differences in microbial community structure and composition, respectively, from extracted DNA. Such assessments serve as a concrete step towards the determination of an optimal DNA extraction method for drinking water biofilms, which can then provide a reliable comparison of the meta-analysis results obtained in different laboratories. PMID:22075624

Analysis of DDT and its metabolites in soil and water samples obtained in the vicinity of a closed-down factory in Bangladesh using various extraction methods.

PubMed

Al Mahmud, M N U; Khalil, Farzana; Rahman, Md Musfiqur; Mamun, M I R; Shoeb, Mohammad; Abd El-Aty, A M; Park, Jong-Hyouk; Shin, Ho-Chul; Nahar, Nilufar; Shim, Jae-Han

2015-12-01

This study was conducted to monitor the spread of dichlorodiphenyltrichloroethane (DDT) and its metabolites (dichlorodiphenyldichloroethylene (DDE), dichlorodiphenyldichloroethane (DDD)) in soil and water to regions surrounding a closed DDT factory in Bangladesh. This fulfillment was accomplished using inter-method and inter-laboratory validation studies. DDTs (DDT and its metabolites) from soil samples were extracted using microwave-assisted extraction (MAE), supercritical fluid extraction (SFE), and solvent extraction (SE). Inter-laboratory calibration was assessed by SE, and all methods were validated by intra- and inter-day accuracy (expressed as recovery %) and precision (expressed as relative standard deviation (RSD)) in the same laboratory, at three fortified concentrations (n = 4). DDTs extracted from water samples by liquid-liquid partitioning and all samples were analyzed by gas chromatography (GC)-electron capture detector (ECD) and confirmed by GC/mass spectrometry (GC/MS). Linearities expressed as determination coefficients (R (2)) were ≥0.995 for matrix-matched calibrations. The recovery rate was in the range of 72-120 and 83-110%, with <15% RSD in soil and water, respectively. The limit of quantification (LOQ) was 0.0165 mg kg(-1) in soil and 0.132 μg L(-1) in water. Greater quantities of DDTs were extracted from soil using the MAE and SE techniques than with the SFE method. Higher amounts of DDTs were discovered in the southern (2.2-936 × 10(2) mg kg(-1)) or southwestern (86.3-2067 × 10(2) mg kg(-1)) direction from the factory than in the eastern direction (1.0-48.6 × 10(2) mg kg(-1)). An exception was the soil sample collected 50 ft (15.24 m) east (2904 × 10(2) mg kg(-1)) of the factory. The spread of DDTs in the water bodies (0.59-3.01 μg L(-1)) was approximately equal in all directions. We concluded that DDTs might have been dumped randomly around the warehouse after the closing of the factory.

High Throughput Determination of Tetramine in Drinking ...

EPA Pesticide Factsheets

Report The sampling and analytical procedure (SAP) presented herein, describes a method for the high throughput determination of tetramethylene disulfotetramine in drinking water by solid phase extraction and isotope dilution gas chromatography/mass spectrometry. This method, which will be included in the SAM, is expected to provide the Water Laboratory Alliance, as part of EPA’s Environmental Response Laboratory Network, with a more reliable and faster means of analyte collection and measurement.

Simultaneous Microwave Extraction and Separation of Volatile and Non-Volatile Organic Compounds of Boldo Leaves. From Lab to Industrial Scale

PubMed Central

Petigny, Loïc; Périno, Sandrine; Minuti, Matteo; Visinoni, Francesco; Wajsman, Joël; Chemat, Farid

2014-01-01

Microwave extraction and separation has been used to increase the concentration of the extract compared to the conventional method with the same solid/liquid ratio, reducing extraction time and separate at the same time Volatile Organic Compounds (VOC) from non-Volatile Organic Compounds (NVOC) of boldo leaves. As preliminary study, a response surface method has been used to optimize the extraction of soluble material and the separation of VOC from the plant in laboratory scale. The results from the statistical analysis revealed that the optimized conditions were: microwave power 200 W, extraction time 56 min and solid liquid ratio of 7.5% of plants in water. Lab scale optimized microwave method is compared to conventional distillation, and requires a power/mass ratio of 0.4 W/g of water engaged. This power/mass ratio is kept in order to upscale from lab to pilot plant. PMID:24776762

Noncontact methods for measuring water-surface elevations and velocities in rivers: Implications for depth and discharge extraction

USGS Publications Warehouse

Nelson, Jonathan M.; Kinzel, Paul J.; McDonald, Richard R.; Schmeeckle, Mark

2016-01-01

Recently developed optical and videographic methods for measuring water-surface properties in a noninvasive manner hold great promise for extracting river hydraulic and bathymetric information. This paper describes such a technique, concentrating on the method of infrared videog- raphy for measuring surface velocities and both acoustic (laboratory-based) and laser-scanning (field-based) techniques for measuring water-surface elevations. In ideal laboratory situations with simple flows, appropriate spatial and temporal averaging results in accurate water-surface elevations and water-surface velocities. In test cases, this accuracy is sufficient to allow direct inversion of the governing equations of motion to produce estimates of depth and discharge. Unlike other optical techniques for determining local depth that rely on transmissivity of the water column (bathymetric lidar, multi/hyperspectral correlation), this method uses only water-surface information, so even deep and/or turbid flows can be investigated. However, significant errors arise in areas of nonhydrostatic spatial accelerations, such as those associated with flow over bedforms or other relatively steep obstacles. Using laboratory measurements for test cases, the cause of these errors is examined and both a simple semi-empirical method and computational results are presented that can potentially reduce bathymetric inversion errors.

Viability testing of material derived from Mycobacterium tuberculosis prior to removal from a containment level-III laboratory as part of a Laboratory Risk Assessment Program.

PubMed

Blackwood, Kym S; Burdz, Tamara V; Turenne, Christine Y; Sharma, Meenu K; Kabani, Amin M; Wolfe, Joyce N

2005-01-24

In the field of clinical mycobacteriology, Mycobacterium tuberculosis (MTB) can be a difficult organism to manipulate due to the restrictive environment of a containment level 3 (CL3) laboratory. Tests for rapid diagnostic work involving smears and molecular methods do not require CL3 practices after the organism has been rendered non-viable. While it has been assumed that after organism deactivation these techniques can be performed outside of a CL3, no conclusive study has consistently confirmed that the organisms are noninfectious after the theoretical 'deactivation' steps. Previous studies have shown that initial steps (such as heating/chemical fixation) may not consistently kill MTB organisms. An inclusive viability study (n = 226) was undertaken to determine at which point handling of culture extraction materials does not necessitate a CL3 environment. Four different laboratory protocols tested for viability included: standard DNA extractions for IS6110 fingerprinting, crude DNA preparations for PCR by boiling and mechanical lysis, protein extractions, and smear preparations. For each protocol, laboratory staff planted a proportion of the resulting material to Bactec 12B medium that was observed for growth for 8 weeks. Of the 208 isolates initially tested, 21 samples grew within the 8-week period. Sixteen (7.7%) of these yielded positive results for MTB that included samples of: deactivated culture resuspensions exposed to 80 degrees C for 20 minutes, smear preparations and protein extractions. Test procedures were consequently modified and tested again (n = 18), resulting in 0% viability. This study demonstrates that it cannot be assumed that conventional practices (i.e. smear preparation) or extraction techniques render the organism non-viable. All methodologies, new and existing, should be examined by individual laboratories to validate the safe removal of material derived from MTB to the outside of a CL3 laboratory. This process is vital to establish in house biosafety-validated practices with the aim of protecting laboratory workers conducting these procedures.

Optimizing techniques to capture and extract environmental DNA for detection and quantification of fish.

PubMed

Eichmiller, Jessica J; Miller, Loren M; Sorensen, Peter W

2016-01-01

Few studies have examined capture and extraction methods for environmental DNA (eDNA) to identify techniques optimal for detection and quantification. In this study, precipitation, centrifugation and filtration eDNA capture methods and six commercially available DNA extraction kits were evaluated for their ability to detect and quantify common carp (Cyprinus carpio) mitochondrial DNA using quantitative PCR in a series of laboratory experiments. Filtration methods yielded the most carp eDNA, and a glass fibre (GF) filter performed better than a similar pore size polycarbonate (PC) filter. Smaller pore sized filters had higher regression slopes of biomass to eDNA, indicating that they were potentially more sensitive to changes in biomass. Comparison of DNA extraction kits showed that the MP Biomedicals FastDNA SPIN Kit yielded the most carp eDNA and was the most sensitive for detection purposes, despite minor inhibition. The MoBio PowerSoil DNA Isolation Kit had the lowest coefficient of variation in extraction efficiency between lake and well water and had no detectable inhibition, making it most suitable for comparisons across aquatic environments. Of the methods tested, we recommend using a 1.5 μm GF filter, followed by extraction with the MP Biomedicals FastDNA SPIN Kit for detection. For quantification of eDNA, filtration through a 0.2-0.6 μm pore size PC filter, followed by extraction with MoBio PowerSoil DNA Isolation Kit was optimal. These results are broadly applicable for laboratory studies on carps and potentially other cyprinids. The recommendations can also be used to inform choice of methodology for field studies. © 2015 John Wiley & Sons Ltd.

Development of Solvent Extraction Approach to Recycle Enriched Molybdenum Material

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

Tkac, Peter; Brown, M. Alex; Sen, Sujat

2016-06-01

Argonne National Laboratory, in cooperation with Oak Ridge National Laboratory and NorthStar Medical Technologies, LLC, is developing a recycling process for a solution containing valuable Mo-100 or Mo-98 enriched material. Previously, Argonne had developed a recycle process using a precipitation technique. However, this process is labor intensive and can lead to production of large volumes of highly corrosive waste. This report discusses an alternative process to recover enriched Mo in the form of ammonium heptamolybdate by using solvent extraction. Small-scale experiments determined the optimal conditions for effective extraction of high Mo concentrations. Methods were developed for removal of ammonium chloridemore » from the molybdenum product of the solvent extraction process. In large-scale experiments, very good purification from potassium and other elements was observed with very high recovery yields (~98%).« less

[The research and application of pretreatment method for matrix-assisted laser desorption ionization-time of flight mass spectrometry identification of filamentous fungi].

PubMed

Huang, Y F; Chang, Z; Bai, J; Zhu, M; Zhang, M X; Wang, M; Zhang, G; Li, X Y; Tong, Y G; Wang, J L; Lu, X X

2017-08-08

Objective: To establish and evaluate the feasibility of a pretreatment method for matrix-assisted laser desorption ionization-time of flight mass spectrometry identification of filamentous fungi developed by the laboratory. Methods: Three hundred and eighty strains of filamentous fungi from January 2014 to December 2016 were recovered and cultured on sabouraud dextrose agar (SDA) plate at 28 ℃ to mature state. Meanwhile, the fungi were cultured in liquid sabouraud medium with a vertical rotation method recommended by Bruker and a horizontal vibration method developed by the laboratory until adequate amount of colonies were observed. For the strains cultured with the three methods, protein was extracted with modified magnetic bead-based extraction method for mass spectrum identification. Results: For 380 fungi strains, it took 3-10 d to culture with SDA culture method, and the ratio of identification of the species and genus was 47% and 81%, respectively; it took 5-7 d to culture with vertical rotation method, and the ratio of identification of the species and genus was 76% and 94%, respectively; it took 1-2 d to culture with horizontal vibration method, and the ratio of identification of the species and genus was 96% and 99%, respectively. For the comparison between horizontal vibration method and SDA culture method comparison, the difference was statistically significant (χ(2)=39.026, P <0.01); for the comparison between horizontal vibration method and vertical rotation method recommended by Bruker, the difference was statistically significant(χ(2)=11.310, P <0.01). Conclusion: The horizontal vibration method and modified magnetic bead-based extraction method developed by the laboratory is superior to the method recommended by Bruker and SDA culture method in terms of the identification capacity for filamentous fungi, which can be applied in clinic.

Simultaneous extraction and clean-up of polychlorinated biphenyls and their metabolites from small tissue samples using pressurized liquid extraction

PubMed Central

Kania-Korwel, Izabela; Zhao, Hongxia; Norstrom, Karin; Li, Xueshu; Hornbuckle, Keri C.; Lehmler, Hans-Joachim

2008-01-01

A pressurized liquid extraction-based method for the simultaneous extraction and in situ clean-up of polychlorinated biphenyls (PCBs), hydroxylated (OH)-PCBs and methylsulfonyl (MeSO2)-PCBs from small (< 0.5 gram) tissue samples was developed and validated. Extraction of a laboratory reference material with hexane:dichloromethane:methanol (48:43:9, v/v) and Florisil as fat retainer allowed an efficient recovery of PCBs (78–112%; RSD: 13–37%), OH-PCBs (46±2%; RSD: 4%) and MeSO2-PCBs (89±21%; RSD: 24%). Comparable results were obtained with an established analysis method for PCBs, OH-PCBs and MeSO2-PCBs. PMID:19019378

Determination of Total Biotin by Liquid Chromatography Coupled with Immunoaffinity Column Cleanup Extraction: Multilaboratory Testing, Final Action 2016.02.

PubMed

Joseph, George; Devi, Ranjani; Marley, Elaine C; Leeman, David

2018-05-01

Single- and multilaboratory testing data have provided systematic scientific evidence that a simple, selective, accurate, and precise method can be used as a potential candidate reference method for dispute resolution in determining total biotin in all forms of infant, adult, and/or pediatric formula. Using LC coupled with immunoaffinity column cleanup extraction, the method fully meets the intended purpose and applicability statement in AOAC Standard Method Performance Requirement 2014.005. The method was applied to a cross-section of infant formula and adult nutritional matrixes, and acceptable precision and accuracy were established. The analytical platform is inexpensive, and the method can be used in almost any laboratory worldwide with basic facilities. The immunoaffinity column cleanup extraction is the key step to successful analysis.

Rapid and reliable high-throughput methods of DNA extraction for use in barcoding and molecular systematics of mushrooms.

PubMed

Dentinger, Bryn T M; Margaritescu, Simona; Moncalvo, Jean-Marc

2010-07-01

We present two methods for DNA extraction from fresh and dried mushrooms that are adaptable to high-throughput sequencing initiatives, such as DNA barcoding. Our results show that these protocols yield ∼85% sequencing success from recently collected materials. Tests with both recent (<2 year) and older (>100 years) specimens reveal that older collections have low success rates and may be an inefficient resource for populating a barcode database. However, our method of extracting DNA from herbarium samples using small amount of tissue is reliable and could be used for important historical specimens. The application of these protocols greatly reduces time, and therefore cost, of generating DNA sequences from mushrooms and other fungi vs. traditional extraction methods. The efficiency of these methods illustrates that standardization and streamlining of sample processing should be shifted from the laboratory to the field. © 2009 Blackwell Publishing Ltd.

Introducing automation to the molecular diagnosis of Trypanosoma cruzi infection: A comparative study of sample treatments, DNA extraction methods and real-time PCR assays.

PubMed

Abras, Alba; Ballart, Cristina; Llovet, Teresa; Roig, Carme; Gutiérrez, Cristina; Tebar, Silvia; Berenguer, Pere; Pinazo, María-Jesús; Posada, Elizabeth; Gascón, Joaquim; Schijman, Alejandro G; Gállego, Montserrat; Muñoz, Carmen

2018-01-01

Polymerase chain reaction (PCR) has become a useful tool for the diagnosis of Trypanosoma cruzi infection. The development of automated DNA extraction methodologies and PCR systems is an important step toward the standardization of protocols in routine diagnosis. To date, there are only two commercially available Real-Time PCR assays for the routine laboratory detection of T. cruzi DNA in clinical samples: TCRUZIDNA.CE (Diagnostic Bioprobes Srl) and RealCycler CHAG (Progenie Molecular). Our aim was to evaluate the RealCycler CHAG assay taking into account the whole process. We assessed the usefulness of an automated DNA extraction system based on magnetic particles (EZ1 Virus Mini Kit v2.0, Qiagen) combined with a commercially available Real-Time PCR assay targeting satellite DNA (SatDNA) of T. cruzi (RealCycler CHAG), a methodology used for routine diagnosis in our hospital. It was compared with a well-known strategy combining a commercial DNA isolation kit based on silica columns (High Pure PCR Template Preparation Kit, Roche Diagnostics) with an in-house Real-Time PCR targeting SatDNA. The results of the two methodologies were in almost perfect agreement, indicating they can be used interchangeably. However, when variations in protocol factors were applied (sample treatment, extraction method and Real-Time PCR), the results were less convincing. A comprehensive fine-tuning of the whole procedure is the key to successful results. Guanidine EDTA-blood (GEB) samples are not suitable for DNA extraction based on magnetic particles due to inhibition, at least when samples are not processed immediately. This is the first study to evaluate the RealCycler CHAG assay taking into account the overall process, including three variables (sample treatment, extraction method and Real-Time PCR). Our findings may contribute to the harmonization of protocols between laboratories and to a wider application of Real-Time PCR in molecular diagnostic laboratories associated with health centers.

Development and validation of a magnetic solid-phase extraction with high-performance liquid chromatography method for the simultaneous determination of amphetamine and methadone in urine.

PubMed

Taghvimi, Arezou; Hamishehkar, Hamed; Ebrahimi, Mahmoud

2016-06-01

The simultaneous determination of amphetamine and methadone was carried out by magnetic graphene oxide nanoparticles, a magnetic solid-phase extraction adsorbent, as a new sample treatment technique. The main factors (the amounts of sample volume, amount of adsorbent, type and amount of extraction organic solvent, time of extraction and desorption, pH, the ionic strength of extraction medium, and agitation rate) influencing the extraction efficiency were investigated and optimized. Under the optimized conditions, good linearity was observed in the range of 100-1500 ng/mL for amphetamine and 100-1000 ng/mL for methadone. The method was evaluated for determination of AM and methadone in positive urine samples, satisfactory results were obtained, therefore magnetic solid-phase extraction can be applied as a novel method for the determination of drugs of abuse in forensic laboratories. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The impact of different DNA extraction kits and laboratories upon the assessment of human gut microbiota composition by 16S rRNA gene sequencing.

PubMed

Kennedy, Nicholas A; Walker, Alan W; Berry, Susan H; Duncan, Sylvia H; Farquarson, Freda M; Louis, Petra; Thomson, John M; Satsangi, Jack; Flint, Harry J; Parkhill, Julian; Lees, Charlie W; Hold, Georgina L

2014-01-01

Determining bacterial community structure in fecal samples through DNA sequencing is an important facet of intestinal health research. The impact of different commercially available DNA extraction kits upon bacterial community structures has received relatively little attention. The aim of this study was to analyze bacterial communities in volunteer and inflammatory bowel disease (IBD) patient fecal samples extracted using widely used DNA extraction kits in established gastrointestinal research laboratories. Fecal samples from two healthy volunteers (H3 and H4) and two relapsing IBD patients (I1 and I2) were investigated. DNA extraction was undertaken using MoBio Powersoil and MP Biomedicals FastDNA SPIN Kit for Soil DNA extraction kits. PCR amplification for pyrosequencing of bacterial 16S rRNA genes was performed in both laboratories on all samples. Hierarchical clustering of sequencing data was done using the Yue and Clayton similarity coefficient. DNA extracted using the FastDNA kit and the MoBio kit gave median DNA concentrations of 475 (interquartile range 228-561) and 22 (IQR 9-36) ng/µL respectively (p<0.0001). Hierarchical clustering of sequence data by Yue and Clayton coefficient revealed four clusters. Samples from individuals H3 and I2 clustered by patient; however, samples from patient I1 extracted with the MoBio kit clustered with samples from patient H4 rather than the other I1 samples. Linear modelling on relative abundance of common bacterial families revealed significant differences between kits; samples extracted with MoBio Powersoil showed significantly increased Bacteroidaceae, Ruminococcaceae and Porphyromonadaceae, and lower Enterobacteriaceae, Lachnospiraceae, Clostridiaceae, and Erysipelotrichaceae (p<0.05). This study demonstrates significant differences in DNA yield and bacterial DNA composition when comparing DNA extracted from the same fecal sample with different extraction kits. This highlights the importance of ensuring that samples in a study are prepared with the same method, and the need for caution when cross-comparing studies that use different methods.

Inorganic arsenic in seafood: does the extraction method matter?

PubMed

Pétursdóttir, Ásta H; Gunnlaugsdóttir, Helga; Krupp, Eva M; Feldmann, Jörg

2014-05-01

Nine different extraction methods were evaluated for three seafood samples to test whether the concentration of inorganic arsenic (iAs) determined in seafood is dependent on the extraction method. Certified reference materials (CRM) DOLT-4 (Dogfish Liver) and TORT-2 (Lobster Hepatopancreas), and a commercial herring fish meal were evaluated. All experimental work described here was carried out by the same operator using the same instrumentation, thus eliminating possible differences in results caused by laboratory related factors. Low concentrations of iAs were found in CRM DOLT-4 (0.012±0.003mgkg(-1)) and the herring fish meal sample (0.007±0.002mgkg(-1)) for all extraction methods. When comparing the concentration of iAs in CRM TORT-2 found in this study and in the literature dilute acids, HNO3 and HCl, showed the highest extracted iAs wheras dilute NaOH (in 50% ethanol) showed significantly lower extracted iAs. However, most other extraction solvents were not statistically different from one another. Copyright © 2013 Elsevier Ltd. All rights reserved.

Evolution of Toxoplasma-PCR methods and practices: a French national survey and proposal for technical guidelines.

PubMed

Roux, Guillaume; Varlet-Marie, Emmanuelle; Bastien, Patrick; Sterkers, Yvon

2018-06-08

The molecular diagnosis of toxoplasmosis lacks standardisation due to the use of numerous methods with variable performance. This diversity of methods also impairs robust performance comparisons between laboratories. The harmonisation of practices by diffusion of technical guidelines is a useful way to improve these performances. The knowledge of methods and practices used for this molecular diagnosis is an essential step to provide guidelines for Toxoplasma-PCR. In the present study, we aimed (i) to describe the methods and practices of Toxoplasma-PCR used by clinical microbiology laboratories in France and (ii) to propose technical guidelines to improve molecular diagnosis of toxoplasmosis. To do so, a yearly self-administered questionnaire-based survey was undertaken in proficient French laboratories from 2008 to 2015, and guidelines were proposed based on the results of those as well as previously published work. This period saw the progressive abandonment of conventional PCR methods, of Toxoplasma-PCR targeting the B1 gene and of the use of two concomitant molecular methods for this diagnosis. The diversity of practices persisted during the study, in spite of the increasing use of commercial kits such as PCR kits, DNA extraction controls and PCR inhibition controls. We also observed a tendency towards the automation of DNA extraction. The evolution of practices did not always go together with an improvement in those, as reported notably by the declining use of Uracil-DNA Glycosylase to avoid carry-over contamination. We here propose technical recommendations which correspond to items explored during the survey, with respect to DNA extraction, Toxoplasma-PCR and good PCR practices. Copyright © 2018 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

Rapid extraction from and direct identification in clinical samples of methicillin-resistant staphylococci using the PCR.

PubMed

Jaffe, R I; Lane, J D; Albury, S V; Niemeyer, D M

2000-09-01

Methicillin-resistant staphylococci (MRS) are one of the most common causes of nosocomial infections and bacteremia. Standard bacterial identification and susceptibility testing frequently require as long as 72 h to report results, and there may be difficulty in rapidly and accurately identifying methicillin resistance. The use of the PCR is a rapid and simple process for the amplification of target DNA sequences, which can be used to identify and test bacteria for antimicrobial resistance. However, many sample preparation methods are unsuitable for PCR utilization in the clinical laboratory because they either are not cost-effective, take too long to perform, or do not provide a satisfactory DNA template for PCR. Our goal was to provide same-day results to facilitate rapid diagnosis and therapy. In this report, we describe a rapid method for extraction of bacterial DNA directly from blood culture bottles that gave quality DNA for PCR in as little as 20 min. We compared this extraction method to the standard QIAGEN method for turnaround time (TAT), cost, purity, and use of template in PCR. Specific identification of MRS was determined using intragenic primer sets for bacterial and Staphylococcus 16S rRNA and mecA gene sequences. The PCR primer sets were validated with 416 isolates of staphylococci, including methicillin-resistant Staphylococcus aureus (n = 106), methicillin-sensitive S. aureus (n = 134), and coagulase-negative Staphylococcus (n = 176). The total supply cost of our extraction method and PCR was $2.15 per sample with a result TAT of less than 4 h. The methods described herein represent a rapid and accurate DNA extraction and PCR-based identification system, which makes the system an ideal candidate for use under austere field conditions and one that may have utility in the clinical laboratory.

Non-structural carbohydrates in woody plants compared among laboratories.

PubMed

Quentin, Audrey G; Pinkard, Elizabeth A; Ryan, Michael G; Tissue, David T; Baggett, L Scott; Adams, Henry D; Maillard, Pascale; Marchand, Jacqueline; Landhäusser, Simon M; Lacointe, André; Gibon, Yves; Anderegg, William R L; Asao, Shinichi; Atkin, Owen K; Bonhomme, Marc; Claye, Caroline; Chow, Pak S; Clément-Vidal, Anne; Davies, Noel W; Dickman, L Turin; Dumbur, Rita; Ellsworth, David S; Falk, Kristen; Galiano, Lucía; Grünzweig, José M; Hartmann, Henrik; Hoch, Günter; Hood, Sharon; Jones, Joanna E; Koike, Takayoshi; Kuhlmann, Iris; Lloret, Francisco; Maestro, Melchor; Mansfield, Shawn D; Martínez-Vilalta, Jordi; Maucourt, Mickael; McDowell, Nathan G; Moing, Annick; Muller, Bertrand; Nebauer, Sergio G; Niinemets, Ülo; Palacio, Sara; Piper, Frida; Raveh, Eran; Richter, Andreas; Rolland, Gaëlle; Rosas, Teresa; Saint Joanis, Brigitte; Sala, Anna; Smith, Renee A; Sterck, Frank; Stinziano, Joseph R; Tobias, Mari; Unda, Faride; Watanabe, Makoto; Way, Danielle A; Weerasinghe, Lasantha K; Wild, Birgit; Wiley, Erin; Woodruff, David R

2015-11-01

Non-structural carbohydrates (NSC) in plant tissue are frequently quantified to make inferences about plant responses to environmental conditions. Laboratories publishing estimates of NSC of woody plants use many different methods to evaluate NSC. We asked whether NSC estimates in the recent literature could be quantitatively compared among studies. We also asked whether any differences among laboratories were related to the extraction and quantification methods used to determine starch and sugar concentrations. These questions were addressed by sending sub-samples collected from five woody plant tissues, which varied in NSC content and chemical composition, to 29 laboratories. Each laboratory analyzed the samples with their laboratory-specific protocols, based on recent publications, to determine concentrations of soluble sugars, starch and their sum, total NSC. Laboratory estimates differed substantially for all samples. For example, estimates for Eucalyptus globulus leaves (EGL) varied from 23 to 116 (mean = 56) mg g(-1) for soluble sugars, 6-533 (mean = 94) mg g(-1) for starch and 53-649 (mean = 153) mg g(-1) for total NSC. Mixed model analysis of variance showed that much of the variability among laboratories was unrelated to the categories we used for extraction and quantification methods (method category R(2) = 0.05-0.12 for soluble sugars, 0.10-0.33 for starch and 0.01-0.09 for total NSC). For EGL, the difference between the highest and lowest least squares means for categories in the mixed model analysis was 33 mg g(-1) for total NSC, compared with the range of laboratory estimates of 596 mg g(-1). Laboratories were reasonably consistent in their ranks of estimates among tissues for starch (r = 0.41-0.91), but less so for total NSC (r = 0.45-0.84) and soluble sugars (r = 0.11-0.83). Our results show that NSC estimates for woody plant tissues cannot be compared among laboratories. The relative changes in NSC between treatments measured within a laboratory may be comparable within and between laboratories, especially for starch. To obtain comparable NSC estimates, we suggest that users can either adopt the reference method given in this publication, or report estimates for a portion of samples using the reference method, and report estimates for a standard reference material. Researchers interested in NSC estimates should work to identify and adopt standard methods. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Evaluation of a solid-phase extraction method for benzoylecgonine urine analysis in a high-throughput forensic urine drug-testing laboratory.

PubMed

Stout, Peter R; Gehlhausen, Jay M; Horn, Carl K; Klette, Kevin L

2002-10-01

A novel extraction and derivatization procedure for the cocaine metabolite benzoylecgonine (BZE) was developed and evaluated for use in a high-volume forensic urine analysis laboratory. Extractions utilized a Speedisk 48 positive pressure extraction manifold and polymer-based cation-exchange extraction columns. Samples were derivatized by the addition of pentafluoropropionic anhydride and pentafluoropropanol. All analyses were performed in selected ion monitoring mode; ions included m/z 421, 300, 272, 429, and 303 with m/z 421 to 429 ratio used for quantitation. The average extraction efficiency was 80%. Seventy-five common over-the-counter products, including prescription drugs, drug metabolites, and other drugs of abuse, demonstrated no significant interference with respect to chromatography or quantitation. The limit of detection and limit of quantitation were calculated at 12.5 ng/mL, and the assay was linear from 12.5 to 20,000 ng/mL with an r2 of 0.99932. A series of 20 precision samples (100 ng/mL) produced an average response of 97.8 ng/mL and a percent coefficient of variation of 4.1%. A set of 79 archived human urine samples that had previously been found to contain BZE were analyzed by 3 separate laboratories. The results did not differ significantly from prior quantitation or between laboratories. The Speedisk has proven viable for a high-volume production facility reducing overall cost of analysis by decreasing analysis time and minimizing waste production while meeting strict forensic requirements.

Isolation of Mitochondrial DNA from Single, Short Hairs without Roots Using Pressure Cycling Technology.

PubMed

Harper, Kathryn A; Meiklejohn, Kelly A; Merritt, Richard T; Walker, Jessica; Fisher, Constance L; Robertson, James M

2018-02-01

Hairs are commonly submitted as evidence to forensic laboratories, but standard nuclear DNA analysis is not always possible. Mitochondria (mt) provide another source of genetic material; however, manual isolation is laborious. In a proof-of-concept study, we assessed pressure cycling technology (PCT; an automated approach that subjects samples to varying cycles of high and low pressure) for extracting mtDNA from single, short hairs without roots. Using three microscopically similar donors, we determined the ideal PCT conditions and compared those yields to those obtained using the traditional manual micro-tissue grinder method. Higher yields were recovered from grinder extracts, but yields from PCT extracts exceeded the requirements for forensic analysis, with the DNA quality confirmed through sequencing. Automated extraction of mtDNA from hairs without roots using PCT could be useful for forensic laboratories processing numerous samples.

Screening of 23 β-lactams in foodstuffs by LC-MS/MS using an alkaline QuEChERS-like extraction.

PubMed

Bessaire, Thomas; Mujahid, Claudia; Beck, Andrea; Tarres, Adrienne; Savoy, Marie-Claude; Woo, Pei-Mun; Mottier, Pascal; Desmarchelier, Aurélien

2018-04-01

A fast and robust high performance LC-MS/MS screening method was developed for the analysis of β-lactam antibiotics in foods of animal origin: eggs, raw milk, processed dairy ingredients, infant formula, and meat- and fish-based products including baby foods. QuEChERS extraction with some adaptations enabled 23 drugs to be simultaneously monitored. Screening target concentrations were set at levels adequate to ensure compliance with current European, Chinese, US and Canadian regulations. The method was fully validated according to the European Community Reference Laboratories Residues Guidelines using 93 food samples of different composition. False-negative and false-positive rates were below 5% for all analytes. The method is adequate for use in high-routine laboratories. A 1-year study was additionally conducted to assess the stability of the 23 analytes in the working standard solution.

Solvent-free microwave extraction of essential oil from aromatic herbs: from laboratory to pilot and industrial scale.

PubMed

Filly, Aurore; Fernandez, Xavier; Minuti, Matteo; Visinoni, Francesco; Cravotto, Giancarlo; Chemat, Farid

2014-05-01

Solvent-free microwave extraction (SFME) has been proposed as a green method for the extraction of essential oil from aromatic herbs that are extensively used in the food industry. This technique is a combination of microwave heating and dry distillation performed at atmospheric pressure without any added solvent or water. The isolation and concentration of volatile compounds is performed in a single stage. In this work, SFME and a conventional technique, hydro-distillation HD (Clevenger apparatus), are used for the extraction of essential oil from rosemary (Rosmarinus officinalis L.) and are compared. This preliminary laboratory study shows that essential oils extracted by SFME in 30min were quantitatively (yield and kinetics profile) and qualitatively (aromatic profile) similar to those obtained using conventional hydro-distillation in 2h. Experiments performed in a 75L pilot microwave reactor prove the feasibility of SFME up scaling and potential industrial applications. Copyright © 2013 Elsevier Ltd. All rights reserved.

Sensitive and specific identification by polymerase chain reaction of Eimeria tenella and Eimeria maxima, important protozoan pathogens in laboratory avian facilities.

PubMed

Lee, Hyun-A; Hong, Sunhwa; Chung, Yungho; Kim, Okjin

2011-09-01

Eimeria tenella and Eimeria maxima are important pathogens causing intracellular protozoa infections in laboratory avian animals and are known to affect experimental results obtained from contaminated animals. This study aimed to find a fast, sensitive, and efficient protocol for the molecular identification of E. tenella and E. maxima in experimental samples using chickens as laboratory avian animals. DNA was extracted from fecal samples collected from chickens and polymerase chain reaction (PCR) analysis was employed to detect E. tenella and E. maxima from the extracted DNA. The target nucleic acid fragments were specifically amplified by PCR. Feces secreting E. tenella and E. maxima were detected by a positive PCR reaction. In this study, we were able to successfully detect E. tenella and E. maxima using the molecular diagnostic method of PCR. As such, we recommended PCR for monitoring E. tenella and E. maxima in laboratory avian facilities.

Extraction and analysis of cortisol from human and monkey hair.

PubMed

Meyer, Jerrold; Novak, Melinda; Hamel, Amanda; Rosenberg, Kendra

2014-01-24

The stress hormone cortisol (CORT) is slowly incorporated into the growing hair shaft of humans, nonhuman primates, and other mammals. We developed and validated a method for CORT extraction and analysis from rhesus monkey hair and subsequently adapted this method for use with human scalp hair. In contrast to CORT "point samples" obtained from plasma or saliva, hair CORT provides an integrated measure of hypothalamic-pituitary-adrenocortical (HPA) system activity, and thus physiological stress, during the period of hormone incorporation. Because human scalp hair grows at an average rate of 1 cm/month, CORT levels obtained from hair segments several cm in length can potentially serve as a biomarker of stress experienced over a number of months. In our method, each hair sample is first washed twice in isopropanol to remove any CORT from the outside of the hair shaft that has been deposited from sweat or sebum. After drying, the sample is ground to a fine powder to break up the hair's protein matrix and increase the surface area for extraction. CORT from the interior of the hair shaft is extracted into methanol, the methanol is evaporated, and the extract is reconstituted in assay buffer. Extracted CORT, along with standards and quality controls, is then analyzed by means of a sensitive and specific commercially available enzyme immunoassay (EIA) kit. Readout from the EIA is converted to pg CORT per mg powdered hair weight. This method has been used in our laboratory to analyze hair CORT in humans, several species of macaque monkeys, marmosets, dogs, and polar bears. Many studies both from our lab and from other research groups have demonstrated the broad applicability of hair CORT for assessing chronic stress exposure in natural as well as laboratory settings.

Applicability of the DPPH assay for evaluating the antioxidant capacity of food additives - inter-laboratory evaluation study -.

PubMed

Shimamura, Tomoko; Sumikura, Yoshihiro; Yamazaki, Takeshi; Tada, Atsuko; Kashiwagi, Takehiro; Ishikawa, Hiroya; Matsui, Toshiro; Sugimoto, Naoki; Akiyama, Hiroshi; Ukeda, Hiroyuki

2014-01-01

An inter-laboratory evaluation study was conducted in order to evaluate the antioxidant capacity of food additives by using a 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. Four antioxidants used as existing food additives (i.e., tea extract, grape seed extract, enju extract, and d-α-tocopherol) and 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) were used as analytical samples, and 14 laboratories participated in this study. The repeatability relative standard deviation (RSD(r)) of the IC50 of Trolox, four antioxidants, and the Trolox equivalent antioxidant capacity (TEAC) were 1.8-2.2%, 2.2-2.9%, and 2.1-2.5%, respectively. Thus, the proposed DPPH assay showed good performance within the same laboratory. The reproducibility relative standard deviation (RSD(R)) of IC50 of Trolox, four antioxidants, and TEAC were 4.0-7.9%, 6.0-11%, and 3.7-9.3%, respectively. The RSD(R)/RSD(r) values of TEAC were lower than, or nearly equal to, those of IC50 of the four antioxidants, suggesting that the use of TEAC was effective for reducing the variance among the laboratories. These results showed that the proposed DPPH assay could be used as a standard method to evaluate the antioxidant capacity of food additives.

The Effect of Storage and Extraction Methods on Amplification of Plasmodium falciparum DNA from Dried Blood Spots.

PubMed

Schwartz, Alanna; Baidjoe, Amrish; Rosenthal, Philip J; Dorsey, Grant; Bousema, Teun; Greenhouse, Bryan

2015-05-01

Extraction and amplification of DNA from dried blood spots (DBS) collected in field studies is commonly used for detection of Plasmodium falciparum. However, there have been few systematic efforts to determine the effects of storage and extraction methods on the sensitivity of DNA amplification. We investigated the effects of storage conditions, length of storage, and DNA extraction methods on amplification via three PCR-based assays using field samples and laboratory controls. Samples stored as DBS for 2 or more years at ambient temperature showed a significant loss of sensitivity that increased with time; after 10 years only 10% samples with parasite densities > 1,000 parasites/μL were detectable by nested polymerase chain reaction (PCR). Conversely, DBS and extracted DNA stored at -20°C showed no loss of sensitivity with time. Samples with low parasite densities amplified more successfully with saponin/Chelex compared with spin-column-based extraction, though the latter method performed better on samples with higher parasite densities stored for 2 years at ambient temperature. DNA extracted via both methods was stable after 20 freeze-thaw cycles. Our results suggest that DBS should be stored at -20°C or extracted immediately, especially if anticipating 2 or more years of storage. © The American Society of Tropical Medicine and Hygiene.

The Effect of Storage and Extraction Methods on Amplification of Plasmodium falciparum DNA from Dried Blood Spots

PubMed Central

Schwartz, Alanna; Baidjoe, Amrish; Rosenthal, Philip J.; Dorsey, Grant; Bousema, Teun; Greenhouse, Bryan

2015-01-01

Extraction and amplification of DNA from dried blood spots (DBS) collected in field studies is commonly used for detection of Plasmodium falciparum. However, there have been few systematic efforts to determine the effects of storage and extraction methods on the sensitivity of DNA amplification. We investigated the effects of storage conditions, length of storage, and DNA extraction methods on amplification via three PCR-based assays using field samples and laboratory controls. Samples stored as DBS for 2 or more years at ambient temperature showed a significant loss of sensitivity that increased with time; after 10 years only 10% samples with parasite densities > 1,000 parasites/μL were detectable by nested polymerase chain reaction (PCR). Conversely, DBS and extracted DNA stored at −20°C showed no loss of sensitivity with time. Samples with low parasite densities amplified more successfully with saponin/Chelex compared with spin-column-based extraction, though the latter method performed better on samples with higher parasite densities stored for 2 years at ambient temperature. DNA extracted via both methods was stable after 20 freeze-thaw cycles. Our results suggest that DBS should be stored at −20°C or extracted immediately, especially if anticipating 2 or more years of storage. PMID:25758652

DNA Everywhere. A Guide for Simplified Environmental Genomic DNA Extraction Suitable for Use in Remote Areas

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

Gabrielle N. Pecora; Francine C. Reid; Lauren M. Tom

2016-05-01

Collecting field samples from remote or geographically distant areas can be a financially and logistically challenging. With participation of a local organization where the samples are originated from, gDNA samples can be extracted from the field and shipped to a research institution for further processing and analysis. The ability to set up gDNA extraction capabilities in the field can drastically reduce cost and time when running long-term microbial studies with a large sample set. The method outlined here has developed a compact and affordable method for setting up a “laboratory” and extracting and shipping gDNA samples from anywhere in themore » world. This white paper explains the process of setting up the “laboratory”, choosing and training individuals with no prior scientific experience how to perform gDNA extractions and safe methods for shipping extracts to any research institution. All methods have been validated by the Andersen group at Lawrence Berkeley National Laboratory using the Berkeley Lab PhyloChip.« less

A Novel Extraction Method Combining Plasma with a Whole-Blood Fraction Shows Excellent Sensitivity and Reproducibility for Patients at High Risk for Invasive Aspergillosis

PubMed Central

Springer, Jan; Schloßnagel, Hannes; Heinz, Werner; Doedt, Thomas; Soeller, Rainer; Einsele, Hermann

2012-01-01

Diagnosis of invasive aspergillosis (IA) is still a major problem in routine clinical practice. Early diagnosis is essential for a good patient prognosis. PCR is a highly sensitive method for the detection of nucleic acids and could play an important role in improving the diagnosis of fungal infections. Therefore, a novel DNA extraction method, ultraclean production (UCP), was developed allowing purification of both cellular and cell-free circulating fungal DNA. In this prospective study we evaluated the commercially available UCP extraction system and compared it to an in-house system. Sixty-three patients at high risk for IA were screened twice weekly, and DNA extracted by both methods was cross-analyzed, in triplicate, by two different real-time PCR assays. The negative predictive values were high for all methods (94.3 to 100%), qualifying them as screening methods, but the sensitivity and diagnostic odds ratios were higher using the UCP extraction method. Sensitivity ranged from 33.3 to 66.7% using the in-house extracts to 100% using the UCP extraction method. Most of the unclassified patients showed no positive PCR results; however, single-positive PCR replicates were observed in some cases. These can bear clinical relevance but should be interpreted with additional clinical and laboratory data. The PCR assays from the UCP extracts showed greater reproducibility than the in-house method for probable IA patients. The standardized UCP extraction method yielded superior results, with regard to sensitivity and reproducibility, than the in-house method. This was independent of the PCR assay used to detect fungal DNA in the sample extracts. PMID:22593600

Molecular detection of Borrelia burgdorferi sensu lato – An analytical comparison of real-time PCR protocols from five different Scandinavian laboratories

PubMed Central

Faller, Maximilian; Wilhelmsson, Peter; Kjelland, Vivian; Andreassen, Åshild; Dargis, Rimtas; Quarsten, Hanne; Dessau, Ram; Fingerle, Volker; Margos, Gabriele; Noraas, Sølvi; Ornstein, Katharina; Petersson, Ann-Cathrine; Matussek, Andreas; Lindgren, Per-Eric; Henningsson, Anna J.

2017-01-01

Introduction Lyme borreliosis (LB) is the most common tick transmitted disease in Europe. The diagnosis of LB today is based on the patient´s medical history, clinical presentation and laboratory findings. The laboratory diagnostics are mainly based on antibody detection, but in certain conditions molecular detection by polymerase chain reaction (PCR) may serve as a complement. Aim The purpose of this study was to evaluate the analytical sensitivity, analytical specificity and concordance of eight different real-time PCR methods at five laboratories in Sweden, Norway and Denmark. Method Each participating laboratory was asked to analyse three different sets of samples (reference panels; all blinded) i) cDNA extracted and transcribed from water spiked with cultured Borrelia strains, ii) cerebrospinal fluid spiked with cultured Borrelia strains, and iii) DNA dilution series extracted from cultured Borrelia and relapsing fever strains. The results and the method descriptions of each laboratory were systematically evaluated. Results and conclusions The analytical sensitivities and the concordance between the eight protocols were in general high. The concordance was especially high between the protocols using 16S rRNA as the target gene, however, this concordance was mainly related to cDNA as the type of template. When comparing cDNA and DNA as the type of template the analytical sensitivity was in general higher for the protocols using DNA as template regardless of the use of target gene. The analytical specificity for all eight protocols was high. However, some protocols were not able to detect Borrelia spielmanii, Borrelia lusitaniae or Borrelia japonica. PMID:28937997

Supercritical Fluid Extraction versus Traditional Solvent Extraction of Caffeine from Tea Leaves: A Laboratory-Based Case Study for an Organic Chemistry Course

ERIC Educational Resources Information Center

Schaber, Peter M.; Larkin, Judith E.; Pines, Harvey A.; Berchou, Kelly; Wierchowski, Elizabeth; Marconi, Andrew; Suriani, Allison

2012-01-01

In this case-based laboratory, an instrument sales person attempts to convince an analysis laboratory of the virtues of supercritical fluid extraction (SFE). The sales person deals directly with the laboratory technicians who will make the decision. Arrangements are made to have SFE instrumentation brought into the laboratory for a comparative…

An Undergraduate Laboratory Experiment that Utilizes a Glass Fiber Filter Assay to Determine the Steroid Specificity and Equilibrium Binding Properties of Glucocorticoid Receptors.

ERIC Educational Resources Information Center

John, Nancy J.; Firestone, Gary L.

1987-01-01

Describes two complementary laboratory exercises that use the glass fiber assay to assess receptor specificity and hormone binding affinity in rat liver cytoplasmic extracts. Details the methods, materials and protocol of the experiments. Discusses the basic concepts illustrated and the feasibility of using the experiments at the undergraduate…

Solid-phase extraction and GC-MS analysis of THC-COOH method optimized for a high-throughput forensic drug-testing laboratory.

PubMed

Stout, P R; Horn, C K; Klette, K L

2001-10-01

In order to facilitate the confirmation analysis of large numbers of urine samples previously screened positive for delta9-tetrahydrocannabinol (THC), an extraction, derivitization, and GC-MS analysis method was developed. This method utilized a positive pressure manifold anion-exchange polymer-based solid-phase extraction followed by elution directly into the automated liquid sampling (ALS) vials. Rapid derivitization was accomplished using pentafluoropropionic anhydride/pentafluoropropanol (PFPA/PFPOH). Recoveries averaged 95% with a limit of detection of 0.875 ng/mL with a 3-mL sample volume. Performance of 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH)-d3 and THC-COOH-d9 internal standards were evaluated. The method was linear to 900 ng/mL THC-COOH using THC-COOH-d9 with negligible contribution from the internal standard to very weak samples. Excellent agreement was seen with previous quantitations of human urine samples. More than 1000 human urine samples were analyzed using the method with 300 samples analyzed using an alternate qualifier ion (m/z 622) after some interference was observed with a qualifier ion (m/z 489). The 622 ion did not exhibit any interference even in samples with interfering peaks present in the 489 ion. The method resulted in dramatic reductions in processing time, waste production, and exposure hazards to laboratory personnel.

DNA extraction from formalin-fixed, paraffin-embedded tissues: protein digestion as a limiting step for retrieval of high-quality DNA.

PubMed

Díaz-Cano, S J; Brady, S P

1997-12-01

Several DNA extraction methods have been used for formalin-fixed, paraffin-embedded tissues, with variable results being reported regarding the suitability of DNA obtained from such sources to serve as template in polymerase chain reaction (PCR)-based genetic analyses. We present a method routinely used for archival material in our laboratory that reliably yields DNA of sufficient quality for PCR studies. This method is based on extended proteinase K digestion (250 micrograms/ml in an EDTA-free calcium-containing buffer supplemented with mussel glycogen) followed by phenol-chloroform extraction. Agarose gel electrophoresis of both digestion buffer aliquots and PCR amplification of the beta-globin gene tested the suitability of the retrieved DNA for PCR amplification.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of pesticides in water by C-18 solid-phase extraction and capillary-column gas chromatography/mass spectrometry with selected-ion monitoring

USGS Publications Warehouse

Zaugg, Steven D.; Sandstrom, Mark W.; Smith, Steven G.; Fehlberg, Kevin M.

1995-01-01

A method for the isolation of 41 pesticides and pesticide metabolites in natural-water samples using C-18 solid-phase extraction and determination by capillary-column gas chromatography/mass spectrometry with selected-ion monitoring is described. Water samples are filtered to remove suspended particulate matter and then are pumped through disposable solid-phase extraction columns containing octadecyl-bonded porous silica to extract the pesticides. The columns are dried using carbon dioxide or nitrogen gas, and adsorbed pesticides are removed from the columns by elution with 3.0 milliliters of hexane-isopropanol (3:1). Extracted pesticides are determined by capillary- column gas chromatography/mass spectrometry with selected-ion monitoring of three characteristic ions. The upper concentration limit is 4 micrograms per liter (g/L) for most pesticides, with the exception of widely used corn herbicides--atrazine, alachlor, cyanazine, and metolachlor--which have upper concentration limits of 20 g/L. Single- operator method detection limits in reagent-water samples range from 0.001 to 0.018 g/L. Average short-term single-operator precision in reagent- water samples is 7 percent at the 0.1- and 1.0-g/L levels and 8 percent at the 0.01-g/L level. Mean recoveries in reagent-water samples are 73 percent at the 0.1- and 1.0-g/L levels and 83 percent at the 0.01-g/L level. The estimated holding time for pesticides after extraction on the solid-phase extraction columns was 7 days. An optional on-site extraction procedure allows for samples to be collected and processed at remote sites where it is difficult to ship samples to the laboratory within the recommended pre-extraction holding time.

COMPARISON OF FIVE EXTRACTION METHODS FOR DETERMINING INCURRED AND FORTIFIED PESTICIDES IN DIETARY COMPOSITES

EPA Science Inventory

U.S. EPA's National Exposure Research Laboratory conducts research to measure exposure of individuals to chemical pollutants through the diet. In support of this research, methods are being evaluated for determination of pesticides in dietary composite samples. In the present s...

Application of micro-solid-phase extraction for the on-site extraction of heterocyclic aromatic amines in seawater.

PubMed

Basheer, Chanbasha

2018-04-01

An efficient on-site extraction technique to determine carcinogenic heterocyclic aromatic amines in seawater has been reported. A micro-solid-phase extraction device placed inside a portable battery-operated pump was used for the on-site extraction of seawater samples. Before on-site applications, parameters that influence the extraction efficiency (extraction time, type of sorbent materials, suitable desorption solvent, desorption time, and sample volume) were investigated and optimized in the laboratory. The developed method was then used for the on-site sampling of heterocyclic aromatic amines determination in seawater samples close to distillation plant. Once the on-site extraction completed, the small extraction device with the analytes was brought back to the laboratory for analysis using high-performance liquid chromatography with fluorescence detection. Based on the optimized conditions, the calibration curves were linear over the concentration range of 0.05-20 μg/L with correlation coefficients up to 0.996. The limits of detection were 0.004-0.026 μg/L, and the reproducibility values were between 1.3 and 7.5%. To evaluate the extraction efficiency, a comparison was made with conventional solid-phase extraction and it was applied to various fortified real seawater samples. The average relative recoveries obtained from the spiked seawater samples varied in the range 79.9-95.2%. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Determination of Major Phenolic Compounds in Echinacea spp. Raw Materials and Finished Products by High-Performance Liquid Chromatography with Ultraviolet Detection: Single-Laboratory Validation Matrix Extension

PubMed Central

Brown, Paula N.; Chan, Michael; Paley, Lori; Betz, Joseph M.

2013-01-01

A method previously validated to determine caftaric acid, chlorogenic acid, cynarin, echinacoside, and cichoric acid in echinacea raw materials has been successfully applied to dry extract and liquid tincture products in response to North American consumer needs. Single-laboratory validation was used to assess the repeatability, accuracy, selectivity, LOD, LOQ, analyte stability (ruggedness), and linearity of the method, with emphasis on finished products. Repeatability precision for each phenolic compound was between 1.04 and 5.65% RSD, with HorRat values between 0.30 and 1.39 for raw and dry extract finished products. HorRat values for tinctures were between 0.09 and 1.10. Accuracy of the method was determined through spike recovery studies. Recovery of each compound from raw material negative control (ginseng) was between 90 and 114%, while recovery from the finished product negative control (maltodextrin and magnesium stearate) was between 97 and 103%. A study was conducted to determine if cichoric acid, a major phenolic component of Echinacea purpurea (L.) Moench and E. angustifolia DC, degrades during sample preparation (extraction) and HPLC analysis. No significant degradation was observed over an extended testing period using the validated method. PMID:22165004

Determination of major phenolic compounds in Echinacea spp. raw materials and finished products by high-performance liquid chromatography with ultraviolet detection: single-laboratory validation matrix extension.

PubMed

Brown, Paula N; Chan, Michael; Paley, Lori; Betz, Joseph M

2011-01-01

A method previously validated to determine caftaric acid, chlorogenic acid, cynarin, echinacoside, and cichoric acid in echinacea raw materials has been successfully applied to dry extract and liquid tincture products in response to North American consumer needs. Single-laboratory validation was used to assess the repeatability, accuracy, selectivity, LOD, LOQ, analyte stability (ruggedness), and linearity of the method, with emphasis on finished products. Repeatability precision for each phenolic compound was between 1.04 and 5.65% RSD, with HorRat values between 0.30 and 1.39 for raw and dry extract finished products. HorRat values for tinctures were between 0.09 and 1.10. Accuracy of the method was determined through spike recovery studies. Recovery of each compound from raw material negative control (ginseng) was between 90 and 114%, while recovery from the finished product negative control (maltodextrin and magnesium stearate) was between 97 and 103%. A study was conducted to determine if cichoric acid, a major phenolic component of Echinacea purpurea (L.) Moench and E. angustifolia DC, degrades during sample preparation (extraction) and HPLC analysis. No significant degradation was observed over an extended testing period using the validated method.

The utility of an automated electronic system to monitor and audit transfusion practice.

PubMed

Grey, D E; Smith, V; Villanueva, G; Richards, B; Augustson, B; Erber, W N

2006-05-01

Transfusion laboratories with transfusion committees have a responsibility to monitor transfusion practice and generate improvements in clinical decision-making and red cell usage. However, this can be problematic and expensive because data cannot be readily extracted from most laboratory information systems. To overcome this problem, we developed and introduced a system to electronically extract and collate extensive amounts of data from two laboratory information systems and to link it with ICD10 clinical codes in a new database using standard information technology. Three data files were generated from two laboratory information systems, ULTRA (version 3.2) and TM, using standard information technology scripts. These were patient pre- and post-transfusion haemoglobin, blood group and antibody screen, and cross match and transfusion data. These data together with ICD10 codes for surgical cases were imported into an MS ACCESS database and linked by means of a unique laboratory number. Queries were then run to extract the relevant information and processed in Microsoft Excel for graphical presentation. We assessed the utility of this data extraction system to audit transfusion practice in a 600-bed adult tertiary hospital over an 18-month period. A total of 52 MB of data were extracted from the two laboratory information systems for the 18-month period and together with 2.0 MB theatre ICD10 data enabled case-specific transfusion information to be generated. The audit evaluated 15,992 blood group and antibody screens, 25,344 cross-matched red cell units and 15,455 transfused red cell units. Data evaluated included cross-matched to transfusion ratios and pre- and post-transfusion haemoglobin levels for a range of clinical diagnoses. Data showed significant differences between clinical units and by ICD10 code. This method to electronically extract large amounts of data and linkage with clinical databases has provided a powerful and sustainable tool for monitoring transfusion practice. It has been successfully used to identify areas requiring education, training and clinical guidance and allows for comparison with national haemoglobin-based transfusion guidelines.

Evaluation and comparison of FTA card and CTAB DNA extraction methods for non-agricultural taxa.

PubMed

Siegel, Chloe S; Stevenson, Florence O; Zimmer, Elizabeth A

2017-02-01

An efficient, effective DNA extraction method is necessary for comprehensive analysis of plant genomes. This study analyzed the quality of DNA obtained using paper FTA cards prepared directly in the field when compared to the more traditional cetyltrimethylammonium bromide (CTAB)-based extraction methods from silica-dried samples. DNA was extracted using FTA cards according to the manufacturer's protocol. In parallel, CTAB-based extractions were done using the automated AutoGen DNA isolation system. DNA quality for both methods was determined for 15 non-agricultural species collected in situ, by gel separation, spectrophotometry, fluorometry, and successful amplification and sequencing of nuclear and chloroplast gene markers. The FTA card extraction method yielded less concentrated, but also less fragmented samples than the CTAB-based technique. The card-extracted samples provided DNA that could be successfully amplified and sequenced. The FTA cards are also useful because the collected samples do not require refrigeration, extensive laboratory expertise, or as many hazardous chemicals as extractions using the CTAB-based technique. The relative success of the FTA card method in our study suggested that this method could be a valuable tool for studies in plant population genetics and conservation biology that may involve screening of hundreds of individual plants. The FTA cards, like the silica gel samples, do not contain plant material capable of propagation, and therefore do not require permits from the U.S. Department of Agriculture (USDA) Animal and Plant Health Inspection Service (APHIS) for transportation.

Separation of the Components of a Commercial Analgesic Tablet: A Two-Week Sequence Comparing Purification by Two-Base Extraction and Column Chromatography

ERIC Educational Resources Information Center

Revell, Kevin D.

2011-01-01

A new laboratory experiment is described in which students compare two benchtop separation methods to isolate the three active components of the commercial analgesic Excedrin. In the two-week sequence, aspirin, acetaminophen, and caffeine are separated using either a two-base liquid-liquid extraction or silica column chromatography. Students then…

Detection of lysergic acid diethylamide (LSD) in urine by gas chromatography-ion trap tandem mass spectrometry.

PubMed

Sklerov, J H; Kalasinsky, K S; Ehorn, C A

1999-10-01

A confirmatory method for the detection and quantitation of lysergic acid diethylamide (LSD) is presented. The method employs gas chromatography-tandem mass spectrometry (GC-MS-MS) using an internal ionization ion trap detector for sensitive MS-MS-in-time measurements of LSD extracted from urine. Following a single-step solid-phase extraction of 5 mL of urine, underivatized LSD can be measured with limits of quantitation and detection of 80 and 20 pg/mL, respectively. Temperature-programmed on-column injections of urine extracts were linear over the concentration range 20-2000 pg/mL (r2 = 0.999). Intraday and interday coefficients of variation were < 6% and < 13%, respectively. This procedure has been applied to quality-control specimens and LSD-positive samples in this laboratory. Comparisons with alternate GC-MS methods and extraction procedures are discussed.

Longitudinal Analysis of New Information Types in Clinical Notes

PubMed Central

Zhang, Rui; Pakhomov, Serguei; Melton, Genevieve B.

2014-01-01

It is increasingly recognized that redundant information in clinical notes within electronic health record (EHR) systems is ubiquitous, significant, and may negatively impact the secondary use of these notes for research and patient care. We investigated several automated methods to identify redundant versus relevant new information in clinical reports. These methods may provide a valuable approach to extract clinically pertinent information and further improve the accuracy of clinical information extraction systems. In this study, we used UMLS semantic types to extract several types of new information, including problems, medications, and laboratory information. Automatically identified new information highly correlated with manual reference standard annotations. Methods to identify different types of new information can potentially help to build up more robust information extraction systems for clinical researchers as well as aid clinicians and researchers in navigating clinical notes more effectively and quickly identify information pertaining to changes in health states. PMID:25717418

Development and Multi-laboratory Verification of U.S. EPA Method 540 for the Analysis of Drinking Water Contaminants by Solid Phase Extraction-LC/MS/MS

EPA Science Inventory

A drinking water method for 12 chemicals, predominately pesticides, is presented that addresses the occurrence monitoring needs of the U.S. Environmental Protection Agency (EPA) for a future Unregulated Contaminant Monitoring Regulation (UCMR). The method employs solid phase ext...

Method Development for the Analysis of 1,4-Dioxane in Drinking Water Using Solid Phase Extraction and Gas Chromatography/Mass Spectrometry

EPA Science Inventory

1,4-Dioxane has been identified as a probable human carcinogen and an emerging contaminant in drinking water. The National Exposure Research Laboratory (NERL) has developed a method for the analysis of 1,4-dioxane in drinking water at ng/L concentrations. The method consists of...

Development and Multi-laboratory Verification of US EPA Method 543 for the Analysis of Drinking Water Contaminants by Online Solid Phase Extraction-LC–MS-MS

EPA Science Inventory

A drinking water method for seven pesticides and pesticide degradates is presented that addresses the occurrence monitoring needs of the US Environmental Protection Agency (EPA) for a future Unregulated Contaminant Monitoring Regulation (UCMR). The method employs online solid pha...

Multiclass pesticide determination in olives and their processing factors in olive oil: comparison of different olive oil extraction systems.

PubMed

Amvrazi, Elpiniki G; Albanis, Triantafyllos A

2008-07-23

The processing factors (pesticide concentration found in olive oil/pesticide concentration found in olives) of azinphos methyl, chlorpyrifos, lambda-cyhalothrin, deltamethrin, diazinon, dimethoate, endosulfan, and fenthion were determined in olive oil production process in various laboratory-scale olive oil extractions based on three- or two-phase centrifugation systems in comparison with samples collected during olive oil extractions in conventional olive mills located at different olive oil production areas in Greece. Pesticide analyses were performed using a multiresidue method developed in our laboratory for the determination of different insecticides and herbicides in olive oil by solid-phase extraction techniques coupled to gas chromatography detection (electron capture detection and nitrogen phosphorus detection), optimized, and validated for olive fruits sample preparation. Processing factors were found to vary among the different pesticides studied. Water addition in the oil extraction procedure (as in a three-phase centrifugation system) was found to decrease the processing factors of dimethoate, alpha-endosulfan, diazinon, and chlorpyrifos, whereas those of fenthion, azinphos methyl, beta-endosulfan, lambda-cyhalothrin, and deltamethrin residues were not affected. The water content of olives processed was found to proportionally affect pesticide processing factors. Fenthion sulfoxide and endosulfan sulfate were the major metabolites of fenthion and endosulfan, respectively, that were detected in laboratory-produced olive oils, but only the concentration of fenthion sulfoxide was found to increase with the increase of water addition in the olive oil extraction process.

Development and validation of a method for determination of corticosteroids in pig fat using liquid chromatography-tandem mass spectrometry.

PubMed

Tölgyesi, Adám; Sharma, Virender K; Fekete, Jeno

2011-02-15

A new method was developed to determine five corticosteroids (prednisolone, methylprednisone, flumethasone, dexamethasone, and methylprednisolone) in pig fat samples by liquid chromatography-tandem mass spectrometry (LC-MS/MS) utilizing an optimized liquid-liquid extraction (LLE) and subsequent solid-phase extraction (SPE) for sample clean-up. In the sample preparation, a pig fat sample was dissolved in n-hexane and then extracted into the methanol-water (50/50, v/v) mixture that enabled extraction of only medium polar corticosteroids and not the non-polar components of matrices. This extract was cleaned-up and concentrated on polymeric Oasis HLB SPE cartridge. Separation involved isocratic solvent (methanol-acetate buffer, pH 5.4) and Ascentis Express Fused-Core type HLPC column; reduced the analysis time to 7.5 min, which is at least two times lower than time required for separation using conventional techniques. Other advantage of the developed method is the minimized ion suppression of LC-MS/MS analysis, which allowed detection of corticosteroids in sub μg/kg. Method was validated according to European Union (EU) Commission Decision 2002/657/EC. Measured parameters such as selectivity, linearity, recovery, within-laboratory reproducibility, decision limit, and detection capability satisfied the EU Directive. Ranges of mean recoveries and within-laboratory reproducibility were 81-100% and 8.0-20.5%, respectively. Decision limits were calculated in the range from 4.5 to 11.9 μg/kg for MRL compounds and varied from 0.1 to 0.2 μg/kg for banned substances. Limit of detections (LODs), calculated as three time signal-to-noise ratio, were in the range of 0.1-0.3 μg/kg. Copyright © 2010 Elsevier B.V. All rights reserved.

Development of an Efficient Entire-Capsid-Coding-Region Amplification Method for Direct Detection of Poliovirus from Stool Extracts

PubMed Central

Kilpatrick, David R.; Nakamura, Tomofumi; Burns, Cara C.; Bukbuk, David; Oderinde, Soji B.; Oberste, M. Steven; Kew, Olen M.; Pallansch, Mark A.; Shimizu, Hiroyuki

2014-01-01

Laboratory diagnosis has played a critical role in the Global Polio Eradication Initiative since 1988, by isolating and identifying poliovirus (PV) from stool specimens by using cell culture as a highly sensitive system to detect PV. In the present study, we aimed to develop a molecular method to detect PV directly from stool extracts, with a high efficiency comparable to that of cell culture. We developed a method to efficiently amplify the entire capsid coding region of human enteroviruses (EVs) including PV. cDNAs of the entire capsid coding region (3.9 kb) were obtained from as few as 50 copies of PV genomes. PV was detected from the cDNAs with an improved PV-specific real-time reverse transcription-PCR system and nucleotide sequence analysis of the VP1 coding region. For assay validation, we analyzed 84 stool extracts that were positive for PV in cell culture and detected PV genomes from 100% of the extracts (84/84 samples) with this method in combination with a PV-specific extraction method. PV could be detected in 2/4 stool extract samples that were negative for PV in cell culture. In PV-positive samples, EV species C viruses were also detected with high frequency (27% [23/86 samples]). This method would be useful for direct detection of PV from stool extracts without using cell culture. PMID:25339406

Evaluating the efficacy of DNA differential extraction methods for sexual assault evidence.

PubMed

Klein, Sonja B; Buoncristiani, Martin R

2017-07-01

Analysis of sexual assault evidence, often a mixture of spermatozoa and victim epithelial cells, represents a significant portion of a forensic DNA laboratory's case load. Successful genotyping of sperm DNA from these mixed cell samples, particularly with low amounts of sperm, depends on maximizing sperm DNA recovery and minimizing non-sperm DNA carryover. For evaluating the efficacy of the differential extraction, we present a method which uses a Separation Potential Ratio (SPRED) to consider both sperm DNA recovery and non-sperm DNA removal as variables for determining separation efficiency. In addition, we describe how the ratio of male-to-female DNA in the sperm fraction may be estimated by using the SPRED of the differential extraction method in conjunction with the estimated ratio of male-to-female DNA initially present on the mixed swab. This approach may be useful for evaluating or modifying differential extraction methods, as we demonstrate by comparing experimental results obtained from the traditional differential extraction and the Erase Sperm Isolation Kit (PTC © ) procedures. Copyright © 2017 Elsevier B.V. All rights reserved.

Determination of Nitrogen, Phosphorus, and Potassium Release Rates of Slow- and Controlled-Release Fertilizers: Single-Laboratory Validation, First Action 2015.15.

PubMed

Thiex, Nancy

2016-01-01

A previously validated method for the determination of nitrogen release patterns of slow- and controlled-release fertilizers (SRFs and CRFs, respectively) was submitted to the Expert Review Panel (ERP) for Fertilizers for consideration of First Action Official Method(SM) status. The ERP evaluated the single-laboratory validation results and recommended the method for First Action Official Method status and provided recommendations for achieving Final Action. The 180 day soil incubation-column leaching technique was demonstrated to be a robust and reliable method for characterizing N release patterns from SRFs and CRFs. The method was reproducible, and the results were only slightly affected by variations in environmental factors such as microbial activity, soil moisture, temperature, and texture. The release of P and K were also studied, but at fewer replications than for N. Optimization experiments on the accelerated 74 h extraction method indicated that temperature was the only factor found to substantially influence nutrient-release rates from the materials studied, and an optimized extraction profile was established as follows: 2 h at 25°C, 2 h at 50°C, 20 h at 55°C, and 50 h at 60°C.

Poly(methyloctylsiloxane) immobilized on silica as a sorbent for solid-phase extraction of some pesticides.

PubMed

Vigna, Camila R M; Morais, Lais S R; Collins, Carol H; Jardim, Isabel C S F

2006-05-12

A laboratory-made sorbent for solid-phase extraction (SPE) was obtained by thermal immobilization of poly(methyloctylsiloxane) (PMOS) onto silica. Cartridges packed with the new sorbent were used for the simultaneous determination of imazethapyr, nicosulfuron, diuron, linuron and chlorimuron-ethyl in water. These pesticides were separated and quantified using high-performance liquid chromatography with diode array detection (HPLC-DAD). The recoveries achieved with the laboratory-made PMOS cartridges were compared with those of some commercially available silica-based and polymer-based cartridges having C18, C8 and NH(2) pendant groups. Method validation using the laboratory-made sorbent was performed for the five pesticides at three fortifications levels (1x, 2x and 10x the limit of quantification of each pesticide). The laboratory-made PMOS cartridge has low cost preparation and showed good recoveries (72-111%) for all pesticides. Repeatability and intermediate precision were lower than 15%. Its performance was similar or even better, in some cases, than those of the commercial cartridges.

GPR-Based Water Leak Models in Water Distribution Systems

PubMed Central

Ayala-Cabrera, David; Herrera, Manuel; Izquierdo, Joaquín; Ocaña-Levario, Silvia J.; Pérez-García, Rafael

2013-01-01

This paper addresses the problem of leakage in water distribution systems through the use of ground penetrating radar (GPR) as a nondestructive method. Laboratory tests are performed to extract features of water leakage from the obtained GPR images. Moreover, a test in a real-world urban system under real conditions is performed. Feature extraction is performed by interpreting GPR images with the support of a pre-processing methodology based on an appropriate combination of statistical methods and multi-agent systems. The results of these tests are presented, interpreted, analyzed and discussed in this paper.

Direct immersion single drop micro-extraction method for multi-class pesticides analysis in mango using GC-MS.

PubMed

Pano-Farias, Norma S; Ceballos-Magaña, Silvia G; Muñiz-Valencia, Roberto; Jurado, Jose M; Alcázar, Ángela; Aguayo-Villarreal, Ismael A

2017-12-15

Due the negative effects of pesticides on environment and human health, more efficient and environmentally friendly methods are needed. In this sense, a simple, fast, free from memory effects and economical direct-immersion single drop micro-extraction (SDME) method and GC-MS for multi-class pesticides determination in mango samples was developed. Sample pre-treatment using ultrasound-assisted solvent extraction and factors affecting the SDME procedure (extractant solvent, drop volume, stirring rate, ionic strength, time, pH and temperature) were optimized using factorial experimental design. This method presented high sensitive (LOD: 0.14-169.20μgkg -1 ), acceptable precision (RSD: 0.7-19.1%), satisfactory recovery (69-119%) and high enrichment factors (20-722). Several obtained LOQs are below the MRLs established by the European Commission; therefore, the method could be applied for pesticides determination in routing analysis and custom laboratories. Moreover, this method has shown to be suitable for determination of some of the studied pesticides in lime, melon, papaya, banana, tomato, and lettuce. Copyright © 2017 Elsevier Ltd. All rights reserved.

Method optimization for non-equilibrium solid phase microextraction sampling of HAPs for GC/MS analysis

NASA Astrophysics Data System (ADS)

Zawadowicz, M. A.; Del Negro, L. A.

2010-12-01

Hazardous air pollutants (HAPs) are usually present in the atmosphere at pptv-level, requiring measurements with high sensitivity and minimal contamination. Commonly used evacuated canister methods require an overhead in space, money and time that often is prohibitive to primarily-undergraduate institutions. This study optimized an analytical method based on solid-phase microextraction (SPME) of ambient gaseous matrix, which is a cost-effective technique of selective VOC extraction, accessible to an unskilled undergraduate. Several approaches to SPME extraction and sample analysis were characterized and several extraction parameters optimized. Extraction time, temperature and laminar air flow velocity around the fiber were optimized to give highest signal and efficiency. Direct, dynamic extraction of benzene from a moving air stream produced better precision (±10%) than sampling of stagnant air collected in a polymeric bag (±24%). Using a low-polarity chromatographic column in place of a standard (5%-Phenyl)-methylpolysiloxane phase decreased the benzene detection limit from 2 ppbv to 100 pptv. The developed method is simple and fast, requiring 15-20 minutes per extraction and analysis. It will be field-validated and used as a field laboratory component of various undergraduate Chemistry and Environmental Studies courses.

A modified method for determining tannin-protein precipitation capacity using accelerated solvent extraction (ASE) and microplate gel filtration.

PubMed

McArt, Scott H; Spalinger, Donald E; Kennish, John M; Collins, William B

2006-06-01

The protein precipitation assay used by Robbins et al., (1987) Ecology 68:98-107 has been shown to predict successfully the reduction in protein availability to some ruminants due to tannins. The procedure, however, is expensive and laborious, which limits its utility, especially for quantitative ecological or nutritional applications where large numbers of assays may be required. We have modified the method to decrease its cost and increase laboratory efficiency by: (1) automating the extraction by using Accelerated Solvent Extraction (ASE); and (2) by scaling and automating the precipitation reaction, chromatography, and spectrometry with microplate gel filtration and an automated UV-VIS microplate spectrometer. ASE extraction is shown to be as effective at extracting tannins as the hot methanol technique. Additionally, the microplate assay is sensitive and precise. We show that the results from the new technique correspond in a nearly 1:1 relationship to the results of the previous technique. Hence, this method could reliably replace the older method with no loss in relevance to herbivore protein digestion. Moreover, the ASE extraction technique should be applicable to other tannin-protein precipitation assays and possibly other phenolic assays.

Application of supercritical fluid carbon dioxide to the extraction and analysis of lipids.

PubMed

Lee, Jae Won; Fukusaki, Eiichiro; Bamba, Takeshi

2012-10-01

Supercritical carbon dioxide (SCCO(2)) is an ecofriendly supercritical fluid that is chemically inert, nontoxic, noninflammable and nonpolluting. As a green material, SCCO(2) has desirable properties such as high density, low viscosity and high diffusivity that make it suitable for use as a solvent in supercritical fluid extraction, an effective and environment-friendly analytical method, and as a mobile phase for supercritical fluid chromatography, which facilitates high-throughput, high-resolution analysis. Furthermore, the low polarity of SCCO(2) is suitable for the extraction and analysis of hydrophobic compounds. The growing concern surrounding environmental pollution has triggered the development of green analysis methods based on the use of SCCO(2) in various laboratories and industries. SCCO(2) is becoming an effective alternative to conventional organic solvents. In this review, the usefulness of SCCO(2) in supercritical fluid extraction and supercritical fluid chromatography for the extraction and analysis of lipids is described.

Multi-Laboratory Study of Five Methods for the Determination of Brevetoxins in Shellfish Tissue Extracts.

PubMed

Dickey, Robert W; Plakas, Steven M; Jester, Edward L E; El Said, Kathleen R; Johannessen, Jan N; Flewelling, Leanne J; Scott, Paula; Hammond, Dan G; Van Dolah, Frances M; Leighfield, Tod A; Bottein Dachraoui, Marie-Yasmine; Ramsdell, John S; Pierce, Richard H; Henry, Mike S; Poli, Mark A; Walker, Calvin; Kurtz, Jan; Naar, Jerome; Baden, Daniel G; Musser, Steve M; White, Kevin D; Truman, Penelope; Miller, Aaron; Hawryluk, Timothy P; Wekell, Marleen M; Stirling, David; Quilliam, Michael A; Lee, Jung K

A thirteen-laboratory comparative study tested the performance of four methods as alternatives to mouse bioassay for the determination of brevetoxins in shellfish. The methods were N2a neuroblastoma cell assay, two variations of the sodium channel receptor binding assay, competitive ELISA, and LC/MS. Three to five laboratories independently performed each method using centrally prepared spiked and naturally incurred test samples. Competitive ELISA and receptor binding (96-well format) compared most favorably with mouse bioassay. Between-laboratory relative standard deviations (RSDR) ranged from 10 to 20% for ELISA and 14 to 31% for receptor binding. Within-laboratory (RSDr) ranged from 6 to 15% for ELISA, and 5 to 31% for receptor binding. Cell assay was extremely sensitive but data variation rendered it unsuitable for statistical treatment. LC/MS performed as well as ELISA on spiked test samples but was inordinately affected by lack of toxin-metabolite standards, uniform instrumental parameters, or both, on incurred test samples. The ELISA and receptor binding assay are good alternatives to mouse bioassay for the determination of brevetoxins in shellfish.

Evaluation and comparison of FTA card and CTAB DNA extraction methods for non-agricultural taxa1

PubMed Central

Siegel, Chloe S.; Stevenson, Florence O.; Zimmer, Elizabeth A.

2017-01-01

Premise of the study: An efficient, effective DNA extraction method is necessary for comprehensive analysis of plant genomes. This study analyzed the quality of DNA obtained using paper FTA cards prepared directly in the field when compared to the more traditional cetyltrimethylammonium bromide (CTAB)–based extraction methods from silica-dried samples. Methods: DNA was extracted using FTA cards according to the manufacturer’s protocol. In parallel, CTAB-based extractions were done using the automated AutoGen DNA isolation system. DNA quality for both methods was determined for 15 non-agricultural species collected in situ, by gel separation, spectrophotometry, fluorometry, and successful amplification and sequencing of nuclear and chloroplast gene markers. Results: The FTA card extraction method yielded less concentrated, but also less fragmented samples than the CTAB-based technique. The card-extracted samples provided DNA that could be successfully amplified and sequenced. The FTA cards are also useful because the collected samples do not require refrigeration, extensive laboratory expertise, or as many hazardous chemicals as extractions using the CTAB-based technique. Discussion: The relative success of the FTA card method in our study suggested that this method could be a valuable tool for studies in plant population genetics and conservation biology that may involve screening of hundreds of individual plants. The FTA cards, like the silica gel samples, do not contain plant material capable of propagation, and therefore do not require permits from the U.S. Department of Agriculture (USDA) Animal and Plant Health Inspection Service (APHIS) for transportation. PMID:28224056

Comparison of solvent/derivatization agent systems for determination of extractable toluene diisocyanate from flexible polyurethane foam.

PubMed

Vangronsveld, Erik; Berckmans, Steven; Spence, Mark

2013-06-01

Flexible polyurethane foam (FPF) is produced from the reaction of toluene diisocyanate (TDI) and polyols. Limited and conflicting results exist in the literature concerning the presence of unreacted TDI remaining in FPF as determined by various solvent extraction and analysis techniques. This study reports investigations into the effect of several solvent/derivatization agent combinations on extractable TDI results and suggests a preferred method. The suggested preferred method employs a syringe-based multiple extraction of foam samples with a toluene solution of 1-(2-methoxyphenyl)-piperazine. Extracts are analyzed by liquid chromatography using an ion trap mass spectrometry detection technique. Detection limits of the method are ~10ng TDI g(-1) foam (10 ppb, w/w) for each TDI isomer (i.e. 2,4-TDI and 2,6-TDI). The method was evaluated by a three-laboratory interlaboratory comparison using two representative foam samples. The total extractable TDI results found by the three labs for the two foams were in good agreement (relative standard deviation of the mean of 30-40%). The method has utility as a basis for comparing FPFs, but the interpretation of extractable TDI results using any solvent as the true value for 'free' or 'unreacted' TDI in the foam is problematic, as demonstrated by the difference in the extracted TDI results from the different extraction systems studied. Further, a consideration of polyurethane foam chemistry raises the possibility that extractable TDI may result from decomposition of parts of the foam structure (e.g. dimers, biurets, and allophanates) by the extraction system.

Biology Notes.

ERIC Educational Resources Information Center

School Science Review, 1982

1982-01-01

Describes laboratory procedures, demonstrations, and classroom materials, including "diet poker" (nutrition game); an experiment on enzyme characteristics; demonstrations of yeast anaerobic respiration and color preference in Calliphora larvae; method to extract eugenol from clove oil to show antibiotic properties; and Benedict's test.…

Optimization and single-laboratory validation of a method for the determination of flavonolignans in milk thistle seeds by high-performance liquid chromatography with ultraviolet detection.

PubMed

Mudge, Elizabeth; Paley, Lori; Schieber, Andreas; Brown, Paula N

2015-10-01

Seeds of milk thistle, Silybum marianum (L.) Gaertn., are used for treatment and prevention of liver disorders and were identified as a high priority ingredient requiring a validated analytical method. An AOAC International expert panel reviewed existing methods and made recommendations concerning method optimization prior to validation. A series of extraction and separation studies were undertaken on the selected method for determining flavonolignans from milk thistle seeds and finished products to address the review panel recommendations. Once optimized, a single-laboratory validation study was conducted. The method was assessed for repeatability, accuracy, selectivity, LOD, LOQ, analyte stability, and linearity. Flavonolignan content ranged from 1.40 to 52.86% in raw materials and dry finished products and ranged from 36.16 to 1570.7 μg/mL in liquid tinctures. Repeatability for the individual flavonolignans in raw materials and finished products ranged from 1.03 to 9.88% RSDr, with HorRat values between 0.21 and 1.55. Calibration curves for all flavonolignan concentrations had correlation coefficients of >99.8%. The LODs for the flavonolignans ranged from 0.20 to 0.48 μg/mL at 288 nm. Based on the results of this single-laboratory validation, this method is suitable for the quantitation of the six major flavonolignans in milk thistle raw materials and finished products, as well as multicomponent products containing dandelion, schizandra berry, and artichoke extracts. It is recommended that this method be adopted as First Action Official Method status by AOAC International.

Extraction fatty acid as a source to produce biofuel in microalgae Chlorella sp. and Spirulina sp. using supercritical carbon dioxide

NASA Astrophysics Data System (ADS)

Tai, Do Chiem; Hai, Dam Thi Thanh; Vinh, Nguyen Hanh; Phung, Le Thi Kim

2016-06-01

In this research, the fatty acids of isolated microalgae were extracted by some technologies such as maceration, Soxhlet, ultrasonic-assisted extraction and supercritical fluid extraction; and analyzed for biodiesel production using GC-MS. This work deals with the extraction of microalgae oil from dry biomass by using supercritical fluid extraction method. A complete study at laboratory of the influence of some parameters on the extraction kinetics and yields and on the composition of the oil in terms of lipid classes and profiles is proposed. Two types of microalgae were studied: Chlorella sp. and Spirulina sp. For the extraction of oil from microalgae, supercritical CO2 (SC-CO2) is regarded with interest, being safer than n-hexane and offering a negligible environmental impact, a short extraction time and a high-quality final product. Whilst some experimental papers are available on the supercritical fluid extraction (SFE) of oil from microalgae, only limited information exists on the kinetics of the process. These results demonstrate that supercritical CO2 extraction is an efficient method for the complete recovery of the neutral lipid phase.

MALDI-TOF identification of Gram-negative bacteria directly from blood culture bottles containing charcoal: Sepsityper® kits versus centrifugation-filtration method.

PubMed

Riederer, Kathleen; Cruz, Kristian; Shemes, Stephen; Szpunar, Susan; Fishbain, Joel T

2015-06-01

Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry has dramatically altered the way microbiology laboratories identify clinical isolates. Direct blood culture (BC) detection may be hampered, however, by the presence of charcoal in BC bottles currently in clinical use. This study evaluates an in-house process for extraction and MALDI-TOF identification of Gram-negative bacteria directly from BC bottles containing charcoal. Three hundred BC aliquots were extracted by a centrifugation-filtration method developed in our research laboratory with the first 96 samples processed in parallel using Sepsityper® kits. Controls were colonies from solid media with standard phenotypic and MALDI-TOF identification. The identification of Gram-negative bacteria was successful more often via the in-house method compared to Sepsityper® kits (94.7% versus 78.1%, P≤0.0001). Our in-house centrifugation-filtration method was further validated for isolation and identification of Gram-negative bacteria (95%; n=300) directly from BC bottles containing charcoal. Copyright © 2015 Elsevier Inc. All rights reserved.

Simultaneous determination of diaveridine, trimethoprim and ormetoprim in feed using high performance liquid chromatography tandem mass spectrometry.

PubMed

Yang, Ya-Jun; Liu, Xi-Wang; Li, Bing; Li, Shi-Hong; Kong, Xiao-Jun; Qin, Zhe; Li, Jian-Yong

2016-12-01

This study developed and validated a simple and reliable method for detecting and quantifying DVD, TMP and OMP in feed using dichloromethane extraction followed by HPLC-MS/MS. A matrix effect evaluation was performed using the post-extraction spiking method, and levels were less than ±15% in all three feeds with their corresponding concentrations. LOD and LOQ, CCα and CCβ were 20μgkg(-1) and 40μgkg(-1), 8.68-15.55μgkg(-1) and 10.61-18.92μgkg(-1) for all analytes, respectively. Calibration curves were linear for DVD, TMP and OMP with R(2)⩾0.990 and r⩾0.995, respectively. Recoveries of low, medium and high concentrations using the proposed method ranged from 74.4 to 105.2%. Repeatability and within-laboratory reproducibility were <7.4% (RSD). The chosen seven factors had no a significant influence on robustness. The method showed good performance when it was applied to analyze other laboratory-prepared or actual feed samples. Copyright © 2016 Elsevier Ltd. All rights reserved.

Quantitative determination of methamphetamine in oral fluid by liquid-liquid extraction and gas chromatography/mass spectrometry.

PubMed

Bahmanabadi, L; Akhgari, M; Jokar, F; Sadeghi, H B

2017-02-01

Methamphetamine abuse is one of the most medical and social problems many countries face. In spite of the ban on the use of methamphetamine, it is widely available in Iran's drug black market. There are many analytical methods for the detection of methamphetamine in biological specimen. Oral fluid has become a popular specimen to test for the presence of methamphetamine. The purpose of the present study was to develop a method for the extraction and detection of methamphetamine in oral fluid samples using liquid-liquid extraction (LLE) and gas chromatography/mass spectrometry (GC/MS) methods. An analytical study was designed in that blank and 50 authentic oral fluid samples were collected to be first extracted by LLE and subsequently analysed by GC/MS. The method was fully validated and showed an excellent intra- and inter-assay precision (reflex sympathetic dystrophy ˂ 10%) for external quality control samples. Recovery with LLE methods was 96%. Limit of detection and limit of quantitation were 5 and 15 ng/mL, respectively. The method showed high selectivity, no additional peak due to interfering substances in samples was observed. The introduced method was sensitive, accurate and precise enough for the extraction of methamphetamine from oral fluid samples in forensic toxicology laboratories.

Application of selected methods of remote sensing for detecting carbonaceous water pollution

NASA Technical Reports Server (NTRS)

Davis, E. M.; Fosbury, W. J.

1973-01-01

A reach of the Houston Ship Channel was investigated during three separate overflights correlated with ground truth sampling on the Channel. Samples were analyzed for such conventional parameters as biochemical oxygen demand, chemical oxygen demand, total organic carbon, total inorganic carbon, turbidity, chlorophyll, pH, temperature, dissolved oxygen, and light penetration. Infrared analyses conducted on each sample included reflectance ATR analysis, carbon tetrachloride extraction of organics and subsequent scanning, and KBr evaporate analysis of CCl4 extract concentrate. Imagery which was correlated with field and laboratory data developed from ground truth sampling included that obtained from aerial KA62 hardware, RC-8 metric camera systems, and the RS-14 infrared scanner. The images were subjected to analysis by three film density gradient interpretation units. Data were then analyzed for correlations between imagery interpretation as derived from the three instruments and laboratory infrared signatures and other pertinent field and laboratory analyses.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of chlorinated pesticides in aquatic tissue by capillary-column gas chromatography with electron-capture detection

USGS Publications Warehouse

Leiker, Thomas J.; Madsen, J.E.; Deacon, J.R.; Foreman, W.T.

1995-01-01

A method for the determination of chlorinated organic compounds in aquatic tissue by dual capillary-column gas chromatography with electron-capture detection is described. Whole-body-fish or corbicula tissue is homogenized, Soxhlet extracted, lipid removed by gel permeation chromatography, and fractionated using alumina/silica adsorption chromatography. The extracts are analyzed by dissimilar capillary-column gas chromatography with electron-capture detection. The method reporting limits are 5 micrograms per kilogram (μg/kg) for chlorinated compounds, 50 μg/kg for polychlorinated biphenyls, and 200 μg/kg for toxaphene.

Comparision of ICP-OES and MP-AES in determing soil nutrients by Mechlich3 method

NASA Astrophysics Data System (ADS)

Tonutare, Tonu; Penu, Priit; Krebstein, Kadri; Rodima, Ako; Kolli, Raimo; Shanskiy, Merrit

2014-05-01

Accurate, routine testing of nutrients in soil samples is critical to understanding soil potential fertility. There are different factors which must be taken into account selecting the best analytical technique for soil laboratory analysis. Several techniques can provide adequate detection range for same analytical subject. In similar cases the choise of technique will depend on factors such as sample throughput, required infrastructure, ease of use, used chemicals and need for gas supply and operating costs. Mehlich 3 extraction method is widely used for the determination of the plant available nutrient elements contents in agricultural soils. For determination of Ca, K, and Mg from soil extract depending of laboratory ICP and AAS techniques are used, also flame photometry for K in some laboratories. For the determination of extracted P is used ICP or Vis spectrometry. The excellent sensitivity and wide working range for all extracted elements make ICP a nearly ideal method, so long as the sample throughput is big enough to justify the initial capital outlay. Other advantage of ICP techniques is the multiplex character (simultaneous acquisition of all wavelengths). Depending on element the detection limits are in range 0.1 - 1000 μg/L. For smaller laboratories with low sample throughput requirements the use of AAS is more common. Flame AAS is a fast, relatively cheap and easy technique for analysis of elements. The disadvantages of the method is single element analysis and use of flammable gas, like C2H2 and oxidation gas N2O for some elements. Detection limits of elements for AAS lays from 1 to 1000 μg/L. MP-AES offers a unique alternative to both, AAS and ICP-OES techniques with its detection power, speed of analysis. MP-AES is quite new, simple and relatively inexpensive multielemental technique, which is use self-sustained atmospheric pressure microwave plasma (MP) using nitrogen gas generated by nitrogen generator. Therefore not needs for argon and flammable (C2H2) gases, cylinder handling and the running costs of equipment are low. Detection limits of elements for MP-AES lays between the AAS and ICP ones. The objective of this study was to compare the results of soil analysis using two multielemental analytical methods - ICP-OES and MP-AES. In the experiment, different soil types with various texture, content of organic matter and pH were used. For the study soil samples of Albeluvisols, Leptosols, Cambisols, Regosols and Histosols were used . The plant available nutrients were estimated by Mehlich 3 extraction. The ICP-OES analysis were provided in the Estonian Agricultural Research Centre and MP-AES analysis in department of Soil Science and Agrochemistry at Estonian University of Life Sciences. The detection limits and limits of quantification of Ca, K, Mg and P in extracts are calculated and reported.

Interlaboratory comparison of extraction efficiency of pesticides from surface and laboratory water using solid-phase extraction disks.

PubMed

Senseman, Scott A; Mueller, Thomas C; Riley, Melissa B; Wauchope, R Don; Clegg, Chris; Young, Roddy W; Southwick, Lloyd M; Moye, H Anson; Dumas, Jose A; Mersie, Wondi; Mattice, John D; Leidy, Ross B

2003-06-18

A continuation of an earlier interlaboratory comparison was conducted (1) to assess solid-phase extraction (SPE) using Empore disks to extract atrazine, bromacil, metolachlor, and chlorpyrifos from various water sources accompanied by different sample shipping and quantitative techniques and (2) to compare quantitative results of individual laboratories with results of one common laboratory. Three replicates of a composite surface water (SW) sample were fortified with the analytes along with three replicates of deionized water (DW). A nonfortified DW sample and a nonfortified SW sample were also extracted. All samples were extracted using Empore C(18) disks. After extraction, part of the samples were eluted and analyzed in-house. Duplicate samples were evaporated in a 2-mL vial, shipped dry to a central laboratory (SDC), redissolved, and analyzed. Overall, samples analyzed in-house had higher recoveries than SDC samples. Laboratory x analysis type and laboratory x water source interactions were significant for all four compounds. Seven laboratories participated in this interlaboratory comparison program. No differences in atrazine recoveries were observed from in-house samples analyzed by laboratories A, B, D, and G compared with the recovery of SDC samples. In-house atrazine recoveries from laboratories C and F were higher when compared with recovery from SDC samples. However, laboratory E had lower recoveries from in-house samples compared with SDC samples. For each laboratory, lower recoveries were observed for chlorpyrifos from the SDC samples compared with samples analyzed in-house. Bromacil recovery was <65% at two of the seven laboratories in the study. Bromacil recoveries for the remaining laboratories were >75%. Three laboratories showed no differences in metolachlor recovery; two laboratories had higher recoveries for samples analyzed in-house, and two other laboratories showed higher metolachlor recovery for SDC samples. Laboratory G had a higher recovery in SW for all four compounds compared with DW. Other laboratories that had significant differences in pesticide recovery between the two water sources showed higher recovery in DW than in the SW regardless of the compound. In comparison to earlier work, recovery of these compounds using SPE disks as a temporary storage matrix may be more effective than shipping dried samples in a vial. Problems with analytes such as chlorpyrifos are unavoidable, and it should not be assumed that an extraction procedure using SPE disks will be adequate for all compounds and transferrable across all chromatographic conditions.

An Efficient, Simple, and Noninvasive Procedure for Genotyping Aquatic and Nonaquatic Laboratory Animals.

PubMed

Okada, Morihiro; Miller, Thomas C; Roediger, Julia; Shi, Yun-Bo; Schech, Joseph Mat

2017-09-01

Various animal models are indispensible in biomedical research. Increasing awareness and regulations have prompted the adaptation of more humane approaches in the use of laboratory animals. With the development of easier and faster methodologies to generate genetically altered animals, convenient and humane methods to genotype these animals are important for research involving such animals. Here, we report skin swabbing as a simple and noninvasive method for extracting genomic DNA from mice and frogs for genotyping. We show that this method is highly reliable and suitable for both immature and adult animals. Our approach allows a simpler and more humane approach for genotyping vertebrate animals.

Method development for the analysis of N-nitrosodimethylamine and other N-nitrosamines in drinking water at low nanogram/liter concentrations using solid-phase extraction and gas chromatography with chemical ionization tandem mass spectrometry.

PubMed

Munch, Jean W; Bassett, Margarita V

2006-01-01

N-nitrosodimethylamine (NDMA) is a probable human carcinogen of concern that has been identified as a drinking water contaminant. U.S. Environmental Protection Agency Method 521 has been developed for the analysis of NDMA and 6 additional N-nitrosamines in drinking water at low ng/L concentrations. The method uses solid-phase extraction with coconut charcoal as the sorbent and dichloromethane as the eluent to concentrate 0.50 L water samples to 1 mL. The extracts are analyzed by gas chromatography-chemical ionization tandem mass spectrometry using large-volume injection. Method performance was evaluated in 2 laboratories. Typical analyte recoveries of 87-104% were demonstrated for fortified reagent water samples, and recoveries of 77-106% were demonstrated for fortified drinking water samples. All relative standard deviations on replicate analyses were < 11%.

Determination of five tetracyclines and their epimers by LC-MS/MS based on a liquid-liquid extraction with low temperature partitioning.

PubMed

Desmarchelier, Aurélien; Anizan, Sébastien; Minh Tien, Mai; Savoy, Marie-Claude; Bion, Cindy

2018-04-01

An LC-MS/MS method is presented for screening five tetracyclines and their epimers in a broad range of food products. The scope of matrices includes meat-, fish-, seafood-based products, various dairy ingredients, infant formulae and fats. The method principle is based on a liquid-liquid extraction with aqueous ethylenediaminetetraacetic acid (EDTA) and acetonitrile followed by a freezing step to promote phase separation at low temperature. After defatting with hexane, sample extracts were evaporated and reconstituted before injection onto the LC-MS/MS system. The addition of oxalic acid in the aqueous mobile phase was mandatory to maintain good peak shape and sensitivity over the run. The screening is based upon a double preparation of each sample, one 'as such' and a second one with the analytes spiked in the sample, in order to mitigate the risk of false negative response. The method was validated according to the European Community Reference Laboratories Residues Guidelines. A total of 93 samples were included in the validation by two independent laboratories giving both false-negative and false-positive rates at 0% for all compounds. Over the last two years, 2600 samples were analysed routinely and only one chicken sample was found above the regulatory limit.

Robust determination of surface relaxivity from nuclear magnetic resonance DT2 measurements

NASA Astrophysics Data System (ADS)

Luo, Zhi-Xiang; Paulsen, Jeffrey; Song, Yi-Qiao

2015-10-01

Nuclear magnetic resonance (NMR) is a powerful tool to probe into geological materials such as hydrocarbon reservoir rocks and groundwater aquifers. It is unique in its ability to obtain in situ the fluid type and the pore size distributions (PSD). The T1 and T2 relaxation times are closely related to the pore geometry through the parameter called surface relaxivity. This parameter is critical for converting the relaxation time distribution into the PSD and so is key to accurately predicting permeability. The conventional way to determine the surface relaxivity ρ2 had required independent laboratory measurements of the pore size. Recently Zielinski et al. proposed a restricted diffusion model to extract the surface relaxivity from the NMR diffusion-T2 relaxation (DT2) measurement. Although this method significantly improved the ability to directly extract surface relaxivity from a pure NMR measurement, there are inconsistencies with their model and it relies on a number of preset parameters. Here we propose an improved signal model to incorporate a scalable LT and extend their method to extract the surface relaxivity based on analyzing multiple DT2 maps with varied diffusion observation time. With multiple diffusion observation times, the apparent diffusion coefficient correctly describes the restricted diffusion behavior in samples with wide PSDs, and the new method does not require predetermined parameters, such as the bulk diffusion coefficient and tortuosity. Laboratory experiments on glass beads packs with the beads diameter ranging from 50 μm to 500 μm are used to validate the new method. The extracted diffusion parameters are consistent with their known values and the determined surface relaxivity ρ2 agrees with the expected value within ±7%. This method is further successfully applied on a Berea sandstone core and yields surface relaxivity ρ2 consistent with the literature.

"In situ" extraction of essential oils by use of Dean-Stark glassware and a Vigreux column inside a microwave oven: a procedure for teaching green analytical chemistry.

PubMed

Chemat, Farid; Perino-Issartier, Sandrine; Petitcolas, Emmanuel; Fernandez, Xavier

2012-08-01

One of the principal objectives of sustainable and green processing development remains the dissemination and teaching of green chemistry in colleges, high schools, and academic laboratories. This paper describes simple glassware that illustrates the phenomenon of extraction in a conventional microwave oven as energy source and a process for green analytical chemistry. Simple glassware comprising a Dean-Stark apparatus (for extraction of aromatic plant material and recovery of essential oils and distilled water) and a Vigreux column (as an air-cooled condenser inside the microwave oven) was designed as an in-situ extraction vessel inside a microwave oven. The efficiency of this experiment was validated for extraction of essential oils from 30 g fresh orange peel, a by-product in the production of orange juice. Every laboratory throughout the world can use this equipment. The microwave power is 100 W and the irradiation time 15 min. The method is performed at atmospheric pressure without added solvent or water and furnishes essential oils similar to those obtained by conventional hydro or steam distillation. By use of GC-MS, 22 compounds in orange peel were separated and identified; the main compounds were limonene (72.1%), β-pinene (8.4%), and γ-terpinene (6.9%). This procedure is appropriate for the teaching laboratory, does not require any special microwave equipment, and enables the students to learn the skills of extraction, and chromatographic and spectroscopic analysis. They are also exposed to a dramatic visual example of rapid, sustainable, and green extraction of an essential oil, and are introduced to successful sustainable and green analytical chemistry.

Evaluation of Brazilian Sugarcane Bagasse Characterization: An Interlaboratory Comparison Study

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

Sluiter, Justin B.; Chum, Helena; Gomes, Absai C.

2016-05-01

This paper describes a study of the variability of measured composition for a single bulk sugarcane bagasse conducted across eight laboratories using similar analytical methods, with the purpose of determining the expected variation for compositional analysis performed by different laboratories. The results show good agreement of measured composition within a single laboratory, but greater variability when results are compared among laboratories. These interlaboratory variabilities do not seem to be associated with a specific method or technique or any single piece of instrumentation. The summary censored statistics provide mean values and pooled standard deviations as follows: total extractives 6.7% (0.6%), wholemore » ash 1.5% (0.2%), glucan 42.3% (1.2%), xylan 22.3% (0.5%), total lignin 21.3% (0.4%), and total mass closure 99.4% (2.9%).« less

Development of an efficient entire-capsid-coding-region amplification method for direct detection of poliovirus from stool extracts.

PubMed

Arita, Minetaro; Kilpatrick, David R; Nakamura, Tomofumi; Burns, Cara C; Bukbuk, David; Oderinde, Soji B; Oberste, M Steven; Kew, Olen M; Pallansch, Mark A; Shimizu, Hiroyuki

2015-01-01

Laboratory diagnosis has played a critical role in the Global Polio Eradication Initiative since 1988, by isolating and identifying poliovirus (PV) from stool specimens by using cell culture as a highly sensitive system to detect PV. In the present study, we aimed to develop a molecular method to detect PV directly from stool extracts, with a high efficiency comparable to that of cell culture. We developed a method to efficiently amplify the entire capsid coding region of human enteroviruses (EVs) including PV. cDNAs of the entire capsid coding region (3.9 kb) were obtained from as few as 50 copies of PV genomes. PV was detected from the cDNAs with an improved PV-specific real-time reverse transcription-PCR system and nucleotide sequence analysis of the VP1 coding region. For assay validation, we analyzed 84 stool extracts that were positive for PV in cell culture and detected PV genomes from 100% of the extracts (84/84 samples) with this method in combination with a PV-specific extraction method. PV could be detected in 2/4 stool extract samples that were negative for PV in cell culture. In PV-positive samples, EV species C viruses were also detected with high frequency (27% [23/86 samples]). This method would be useful for direct detection of PV from stool extracts without using cell culture. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Application of PLE for the determination of essential oil components from Thymus vulgaris L.

PubMed

Dawidowicz, Andrzej L; Rado, Ewelina; Wianowska, Dorota; Mardarowicz, Marek; Gawdzik, Jan

2008-08-15

Essential plants, due to their long presence in human history, their status in culinary arts, their use in medicine and perfume manufacture, belong to frequently examined stock materials in scientific and industrial laboratories. Because of a large number of freshly cut, dried or frozen plant samples requiring the determination of essential oil amount and composition, a fast, safe, simple, efficient and highly automatic sample preparation method is needed. Five sample preparation methods (steam distillation, extraction in the Soxhlet apparatus, supercritical fluid extraction, solid phase microextraction and pressurized liquid extraction) used for the isolation of aroma-active components from Thymus vulgaris L. are compared in the paper. The methods are mainly discussed with regard to the recovery of components which typically exist in essential oil isolated by steam distillation. According to the obtained data, PLE is the most efficient sample preparation method in determining the essential oil from the thyme herb. Although co-extraction of non-volatile ingredients is the main drawback of this method, it is characterized by the highest yield of essential oil components and the shortest extraction time required. Moreover, the relative peak amounts of essential components revealed by PLE are comparable with those obtained by steam distillation, which is recognized as standard sample preparation method for the analysis of essential oils in aromatic plants.

Change Analysis in Structural Laser Scanning Point Clouds: The Baseline Method

PubMed Central

Shen, Yueqian; Lindenbergh, Roderik; Wang, Jinhu

2016-01-01

A method is introduced for detecting changes from point clouds that avoids registration. For many applications, changes are detected between two scans of the same scene obtained at different times. Traditionally, these scans are aligned to a common coordinate system having the disadvantage that this registration step introduces additional errors. In addition, registration requires stable targets or features. To avoid these issues, we propose a change detection method based on so-called baselines. Baselines connect feature points within one scan. To analyze changes, baselines connecting corresponding points in two scans are compared. As feature points either targets or virtual points corresponding to some reconstructable feature in the scene are used. The new method is implemented on two scans sampling a masonry laboratory building before and after seismic testing, that resulted in damages in the order of several centimeters. The centres of the bricks of the laboratory building are automatically extracted to serve as virtual points. Baselines connecting virtual points and/or target points are extracted and compared with respect to a suitable structural coordinate system. Changes detected from the baseline analysis are compared to a traditional cloud to cloud change analysis demonstrating the potential of the new method for structural analysis. PMID:28029121

Change Analysis in Structural Laser Scanning Point Clouds: The Baseline Method.

PubMed

Shen, Yueqian; Lindenbergh, Roderik; Wang, Jinhu

2016-12-24

A method is introduced for detecting changes from point clouds that avoids registration. For many applications, changes are detected between two scans of the same scene obtained at different times. Traditionally, these scans are aligned to a common coordinate system having the disadvantage that this registration step introduces additional errors. In addition, registration requires stable targets or features. To avoid these issues, we propose a change detection method based on so-called baselines. Baselines connect feature points within one scan. To analyze changes, baselines connecting corresponding points in two scans are compared. As feature points either targets or virtual points corresponding to some reconstructable feature in the scene are used. The new method is implemented on two scans sampling a masonry laboratory building before and after seismic testing, that resulted in damages in the order of several centimeters. The centres of the bricks of the laboratory building are automatically extracted to serve as virtual points. Baselines connecting virtual points and/or target points are extracted and compared with respect to a suitable structural coordinate system. Changes detected from the baseline analysis are compared to a traditional cloud to cloud change analysis demonstrating the potential of the new method for structural analysis.

Materials and Methods for Streamlined Laboratory Analysis of Environmental Samples, FY 2016 Report

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

Addleman, Raymond S.; Naes, Benjamin E.; McNamara, Bruce K.

The International Atomic Energy Agency (IAEA) relies upon laboratory analysis of environmental samples (typically referred to as “swipes”) collected during on-site inspections of safeguarded facilities to support the detection and deterrence of undeclared activities. Unfortunately, chemical processing and assay of the samples is slow and expensive. A rapid, effective, and simple extraction process and analysis method is needed to provide certified results with improved timeliness at reduced costs (principally in the form of reduced labor), while maintaining or improving sensitivity and efficacy. To address these safeguard needs the Pacific Northwest National Laboratory (PNNL) explored and demonstrated improved methods for environmentalmore » sample (ES) analysis. Improvements for both bulk and particle analysis were explored. To facilitate continuity and adoption, the new sampling materials and processing methods will be compatible with existing IAEA protocols for ES analysis. PNNL collaborated with Oak Ridge National Laboratory (ORNL), which performed independent validation of the new bulk analysis methods and compared performance to traditional IAEA’s Network of Analytical Laboratories (NWAL) protocol. ORNL efforts are reported separately. This report describes PNNL’s FY 2016 progress, which was focused on analytical application supporting environmental monitoring of uranium enrichment plants and nuclear fuel processing. In the future the technology could be applied to other safeguard applications and analytes related to fuel manufacturing, reprocessing, etc. PNNL’s FY 2016 efforts were broken into two tasks and a summary of progress, accomplishments and highlights are provided below. Principal progress and accomplishments on Task 1, Optimize Materials and Methods for ICP-MS Environmental Sample Analysis, are listed below. • Completed initial procedure for rapid uranium extraction from ES swipes based upon carbonate-peroxide chemistry (delivered to ORNL for evaluation). • Explored improvements to carbonate-peroxide rapid uranium extraction chemistry. • Evaluated new sampling materials and methods (in collaboration with ORNL). • Demonstrated successful ES extractions from standard and novel swipes for a wide range uranium compounds of interest including UO 2F 2 and UO 2(NO 3) 2, U 3O 8 and uranium ore concentrate. • Completed initial discussions with commercial suppliers of PTFE swipe materials. • Submitted one manuscript for publication. Two additional drafts are being prepared. Principal progress and accomplishments on Task 2, Optimize Materials and Methods for Direct SIMS Environmental Sample Analysis, are listed below. • Designed a SIMS swipe sample holder that retrofits into existing equipment and provides simple, effective, and rapid mounting of ES samples for direct assay while enabling automation and laboratory integration. • Identified preferred conductive sampling materials with better performance characteristics. • Ran samples on the new PNNL NWAL equivalent Cameca 1280 SIMS system. • Obtained excellent agreement between isotopic ratios for certified materials and direct SIMS assay of very low levels of LEU and HEU UO 2F 2 particles on carbon fiber sampling material. Sample activities range from 1 to 500 CPM (uranium mass on sample is dependent upon specific isotope ratio but is frequently in the subnanogram range). • Found that the presence of the UF molecular ions, as measured by SIMS, provides chemical information about the particle that is separate from the uranium isotopics and strongly suggests that those particles originated from an UF6 enrichment activity. • Submitted one manuscript for publication. Another manuscript is in preparation.« less

Method 522 - Determination of 1,4-Dioxane in Drinking Water by Solid Phase Extraction (SPE) and Gas Chromatography Mass Spectrometry (GC/MS) with Selected Ion Monitoring (SIM)

EPA Science Inventory

1,4-Dioxane has been identified as a probable human carcinogen and an emerging contaminant in drinking water. The National Exposure Research Laboratory (NERL) has developed a method for the analysis of 1,4-dioxane in drinking water at ng/L concentrations. The method consists of...

A RAPID METHOD FOR THE EXTRACTION OF FUNGAL DNA FROM ENVIRONMENTAL SAMPLES: EVALUATION IN THE QUANTITATIVE ANALYSIS OF MEMNONIELLA ECHINATA CONIDIA USING REAL TIME DETECTION OF PCR PRODUCTS

EPA Science Inventory

New technologies are creating the potential for using nucleic acid sequence detection to perform routine microbiological analyses of environmental samples. Our laboratory has recently reported on the development of a method for the quantitative detection of Stachybotrys chartarum...

Extraction study on uranyl nitrate for energy applications

NASA Astrophysics Data System (ADS)

Giri, R.; Nath, G.

2017-07-01

Due to the ever-growing demand of energy nuclear reactor materials and the nuclear energy are now considered to be the most critical materials and source of energy for future era. Deposition of nuclear wastes in different industry, nuclear power sector are very much toxic in open environment which are hazardous to living being. There are different methods for extraction and reprocessing of these materials which are cost effective and tedious process. Ultrasonic assisted solvent extraction process is a most efficient and economical way for extraction of such type materials. The presence of third phase in mixing of extractants-diluent pair with aqueous phase imposes the problems in extraction of nuclear reactor materials. The appropriate solvent mixture in proper concentration is an important step in the solvent extraction process. Study of thermo-physical properties helps in selecting an optimum blend for extraction process. In the present work, the extraction of uranium with the binary mixture of Methyl Ethyl Ketone (MEK) and Kerosene was investigated and discussed with the variation of ultrasonic frequency for different temperatures. The result shows that the low frequency and low temperature is suitable environment for extraction. The extraction of uranium by this method is found to be a better result for extraction study in laboratory scale as well as industrial sector.

Deep liquid-chromatographic purification of uranium extract from technetium

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

Volk, V.; Dvoeglazov, K; Podrezova, L.

The recycling of uranium in the nuclear fuel cycle requires the removal of a number of radioactive and stable impurities like {sup 99}Tc from spent fuels. In order to improve the grade of uranium extract purification from technetium the method of liquid chromatography and the apparatus for its performance have been developed. Process of technetium extraction and concentrating in aqueous solution containing reducing agent has been studied on simulated solutions (U-Tc-HNO{sub 3}-30% TBP-isoparM). The dynamic tests of the method have been carried out on the laboratory unit. Solution of diformyl-hydrazine in nitric acid was used as a stationary phase. Silicamore » gel with specific surface of 186 m{sup 2}/g was used as a carrier of the stationary phase. It is shown that the volume of purified extract increases as the solution temperature increases, concentration of reducing agent increases and extract flow rate decreases. It is established that the technetium content in uranium by this method could achieve a value below 0.3 ppm. Some variants of overload and composition of the stationary phase containing the extracted technetium have been offered and tested. It is defined that the method provides reduction of processing medium-active wastes by more than 10 times during finish refining process. (authors)« less

Determination of Free and Total Choline and Carnitine in Infant Formula and Adult/Pediatric Nutritional Formula by Liquid Chromatography/Tandem Mass Spectrometry (LC/MS/MS): Single-Laboratory Validation, First Action 2015.10.

PubMed

Ellingson, David J; Shippar, Jeffrey J; Gilmore, Justin M

2016-01-01

Analytical methods for the analysis of both L-carnitine and choline are needed for reliable and accurate determination in infant formula and adult/pediatric nutritional formula. These compounds are different in how they are utilized by the human body, but are structurally similar. L-carnitine and choline are quaternary ammonium compounds, enabling both to be retained under acidic conditions with strong cation exchange (SCX) chromatography. This method analyzes both compounds simultaneously as either the free forms or as a total amount that includes bound sources such as phosphatidylcholine or acetylcarnitine. The free analysis consists of water extraction and analysis by LC/MS/MS, while the total analysis consists of extraction by acid assisted microwave hydrolysis and analysis by LC/MS/MS. Calibration standards used for calculations are extracted with all samples in the batch. A single laboratory validation (SLV) was performed following the guidelines of the AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN) utilizing the kit of materials provided. The results achieved meet the requirements of SMPR 2012.010 and 2012.013 for L-carnitine and total choline, respectively.

Determination of Flavonol Aglycones in Ginkgo biloba Dietary Supplement Crude Materials and Finished Products by High-Performance Liquid Chromatography: Single Laboratory Validation

PubMed Central

Gray, Dean; LeVanseler, Kerri; Pan, Meide

2008-01-01

A single laboratory validation (SLV) was completed for a method to determine the flavonol aglycones quercetin, kaempferol, and isorhamnetin in Ginkgo biloba products. The method calculates total glycosides based on these aglycones formed following acid hydrolysis. Nine matrixes were chosen for the study, including crude leaf material, standardized dry powder extract, single and multiple entity finished products, and ethanol and glycerol tinctures. For the 9 matrixes evaluated as part of this SLV, the method appeared to be selective and specific, with no observed interferences. The simplified 60 min oven heating hydrolysis procedure was effective for each of the matrixes studied, with no apparent or consistent differences between 60, 75, and 90 min at 90°C. A Youden ruggedness trial testing 7 factors with the potential to affect quantitative results showed that 2 factors (volume hydrolyzed and test sample extraction/hydrolysis weight) were the most important parameters for control during sample preparation. The method performed well in terms of precision, with 4 matrixes tested in triplicate over a 3-day period showing an overall repeatability (relative standard deviation, RSD) of 2.3%. Analysis of variance testing at α = 0.05 showed no significant differences among the within- or between-group sources of variation, although comparisons of within-day (Sw), between-day (Sb), and total (St) precision showed that a majority of the standard deviation came from within-day determinations for all matrixes. Accuracy testing at 2 levels (approximately 30 and 90% of the determined concentrations in standardized dry powder extract) from 2 complex negative control matrixes showed an overall 96% recovery and RSD of 1.0% for the high spike, and 94% recovery and RSD of 2.5% for the low spike. HorRat scores were within the limits for performance acceptability, ranging from 0.4 to 1.3. Based on the performance results presented herein, it is recommended that this method progress to the collaborative laboratory trial. PMID:16001841

ON-SITE SOLID-PHASE EXTRACTION AND LABORATORY ...

EPA Pesticide Factsheets

Fragrance materials such as synthetic musks in aqueous samples, are normally determined by gas chromatography/mass spectrometry in the selected ion monitoring (SIM) mode to provide maximum sensitivity after liquid-liquid extraction of I -L samples. Full-scan mass spectra are required to verify that a target analyte has been found by comparison with the mass spectra of fragrance compounds in the NIST mass spectral library. A I -L sample usually provides insufficient analyte for full scan data acquisition. This paper describes an on-site extraction method developed at the U.S. Environmental Protection Agency (USEPA)- Las Vegas Nevada - for synthetic musks from 60 L of wastewater effluent. Such a large sample volume permits high-quality, full-scan mass spectra to be obtained for a wide array of synthetic musks. Quantification of these compounds was achieved from the full-scan data directly, without the need to acquire SIM data. The detection limits obtained with this method are an order of magnitude lower than those obtained from liquid-liquid and other solid phase extraction methods. This method is highly reproducible, and recoveries ranged from 80 to 97% in spiked sewage treatment plant effluent. The high rate of sorbent-sample mass transfer eliminated the need for a methanolic activation step, which reduced extraction time, labor, and solvent use, More samples could be extracted in the field at lower cost. After swnple extraction, the light- weight cartridges ar

Liquid-Liquid Extraction of Insecticides from Juice: An Analytical Chemistry Laboratory Experiment

ERIC Educational Resources Information Center

Radford, Samantha A.; Hunter, Ronald E., Jr.; Barr, Dana Boyd; Ryan, P. Barry

2013-01-01

A laboratory experiment was developed to target analytical chemistry students and to teach them about insecticides in food, sample extraction, and cleanup. Micro concentrations (sub-microgram/mL levels) of 12 insecticides spiked into apple juice samples are extracted using liquid-liquid extraction and cleaned up using either a primary-secondary…

Distinguishing Vaccinium Species by Chemical Fingerprinting Based on NMR Spectra, Validated with Spectra Collected in Different Laboratories

PubMed Central

Markus, Michelle A.; Ferrier, Jonathan; Luchsinger, Sarah M.; Yuk, Jimmy; Cuerrier, Alain; Balick, Michael J.; Hicks, Joshua M.; Killday, K. Brian; Kirby, Christopher W.; Berrue, Fabrice; Kerr, Russell G.; Knagge, Kevin; Gödecke, Tanja; Ramirez, Benjamin E.; Lankin, David C.; Pauli, Guido F.; Burton, Ian; Karakach, Tobias K.; Arnason, John T.; Colson, Kimberly L.

2014-01-01

A method was developed to distinguish Vaccinium species based on leaf extracts using nuclear magnetic resonance spectroscopy. Reference spectra were measured on leaf extracts from several species, including lowbush blueberry (Vaccinium angustifolium), oval leaf huckleberry (Vaccinium ovalifolium), and cranberry (Vaccinium macrocarpon). Using principal component analysis, these leaf extracts were resolved in the scores plot. Analysis of variance statistical tests demonstrated that the three groups differ significantly on PC2, establishing that the three species can be distinguished by nuclear magnetic resonance. Soft independent modeling of class analogies models for each species also showed discrimination between species. To demonstrate the robustness of nuclear magnetic resonance spectroscopy for botanical identification, spectra of a sample of lowbush blueberry leaf extract were measured at five different sites, with different field strengths (600 versus 700 MHz), different probe types (cryogenic versus room temperature probes), different sample diameters (1.7 mm versus 5 mm), and different consoles (Avance I versus Avance III). Each laboratory independently demonstrated the linearity of their NMR measurements by acquiring a standard curve for chlorogenic acid (R2 = 0.9782 to 0.9998). Spectra acquired on different spectrometers at different sites classifed into the expected group for the Vaccinium spp., confirming the utility of the method to distinguish Vaccinium species and demonstrating nuclear magnetic resonance fingerprinting for material validation of a natural health product. PMID:24963620

A Simple and Rapid Protocol for Producing Yeast Extract from Saccharomyces cerevisiae Suitable for Preparing Bacterial Culture Media

PubMed Central

Zarei, Omid; Dastmalchi, Siavoush; Hamzeh-Mivehroud, Maryam

2016-01-01

Yeasts, especially Saccharomyces cerevisiae, are one of the oldest organisms with broad spectrum of applications, owing to their unique genetics and physiology. Yeast extract, i.e. the product of yeast cells, is extensively used as nutritional resource in bacterial culture media. The aim of this study was to develop a simple, rapid and cost benefit process to produce the yeast extract. In this procedure mechanical methods such as high temperature and pressure were utilized to produce the yeast extract. The growth of the bacteria feed with the produced yeast extract was monitored in order to assess the quality of the product. The results showed that the quality of the produced yeast extract was very promising concluded from the growth pattern of bacterial cells in media prepared from this product and was comparable with that of the three commercial yeast extracts in terms of bacterial growth properties. One of the main advantages of the current method was that no chemicals and enzymes were used, leading to the reduced production cost. The method is very simple and cost effective, and can be performed in a reasonable time making it suitable for being adopted by research laboratories. Furthermore, it can be scaled up to produce large quantities for industrial applications. PMID:28243289

Methods of Analysis by the U.S. Geological Survey National Water Quality Laboratory - Determination of Wastewater Compounds by Polystyrene-Divinylbenzene Solid-Phase Extraction and Capillary-Column Gas Chromatography/Mass Spectrometry

USGS Publications Warehouse

Zaugg, Steven D.; Smith, Steven G.; Schroeder, Michael P.; Barber, Larry B.; Burkhardt, Mark R.

2002-01-01

A method for the determination of 67 compounds typically found in domestic and industrial wastewater is described. The method was developed in response to increasing concern over the impact of endocrine-disrupting chemicals in wastewater on aquatic organisms. This method also may be useful for evaluating the impact of combined sanitary and storm-sewer overflow on the water quality of urban streams. The method focuses on the determination of compounds that are an indicator of wastewater or that have been chosen on the basis of their endocrine-disrupting potential or toxicity. These compounds include the alkylphenol ethoxylate nonionic surfactants and their degradates, food additives, fragrances, antioxidants, flame retardants, plasticizers, industrial solvents, disinfectants, fecal sterols, polycyclicaromatic hydrocarbons, and high-use domestic pesticides. Water samples are filtered to remove suspended particulate matter and then are extracted by vacuum through disposable solid-phase cartridges that contain polystyrene-divinylbenzene resin. Cartridges are dried with nitrogen gas, and then sorbed compounds are eluted with dichloromethane-diethyl ether (4:1) and determined by capillary-column gas chromatography/mass spectrometry. Recoveries in reagent-water samples fortified at 4 micrograms per liter averaged 74 percent ? 7 percent relative standard deviation for all method compounds. Initial method detection limits for single-component compounds (excluding hormones and sterols) averaged 0.15 microgram per liter. Samples are preserved by filtration, the addition of 60 grams NaCl, and storage at 4 degrees Celsius. The laboratory has established a sample-holding time (prior to sample extraction) of 14 days from the date of sample collection until a statistically accepted method can be used to determine the effectiveness of these sample-preservation procedures.

Portable microwave assisted extraction: An original concept for green analytical chemistry.

PubMed

Perino, Sandrine; Petitcolas, Emmanuel; de la Guardia, Miguel; Chemat, Farid

2013-11-08

This paper describes a portable microwave assisted extraction apparatus (PMAE) for extraction of bioactive compounds especially essential oils and aromas directly in a crop or in a forest. The developed procedure, based on the concept of green analytical chemistry, is appropriate to obtain direct in-field information about the level of essential oils in natural samples and to illustrate green chemical lesson and research. The efficiency of this experiment was validated for the extraction of essential oil of rosemary directly in a crop and allows obtaining a quantitative information on the content of essential oil, which was similar to that obtained by conventional methods in the laboratory. Copyright © 2013 Elsevier B.V. All rights reserved.

Variability of bioaccessibility results using seventeen different methods on a standard reference material, NIST 2710.

PubMed

Koch, Iris; Reimer, Kenneth J; Bakker, Martine I; Basta, Nicholas T; Cave, Mark R; Denys, Sébastien; Dodd, Matt; Hale, Beverly A; Irwin, Rob; Lowney, Yvette W; Moore, Margo M; Paquin, Viviane; Rasmussen, Pat E; Repaso-Subang, Theresa; Stephenson, Gladys L; Siciliano, Steven D; Wragg, Joanna; Zagury, Gerald J

2013-01-01

Bioaccessibility is a measurement of a substance's solubility in the human gastro-intestinal system, and is often used in the risk assessment of soils. The present study was designed to determine the variability among laboratories using different methods to measure the bioaccessibility of 24 inorganic contaminants in one standardized soil sample, the standard reference material NIST 2710. Fourteen laboratories used a total of 17 bioaccessibility extraction methods. The variability between methods was assessed by calculating the reproducibility relative standard deviations (RSDs), where reproducibility is the sum of within-laboratory and between-laboratory variability. Whereas within-laboratory repeatability was usually better than (<) 15% for most elements, reproducibility RSDs were much higher, indicating more variability, although for many elements they were comparable to typical uncertainties (e.g., 30% in commercial laboratories). For five trace elements of interest, reproducibility RSDs were: arsenic (As), 22-44%; cadmium (Cd), 11-41%; Cu, 15-30%; lead (Pb), 45-83%; and Zn, 18-56%. Only one method variable, pH, was found to correlate significantly with bioaccessibility for aluminum (Al), Cd, copper (Cu), manganese (Mn), Pb and zinc (Zn) but other method variables could not be examined systematically because of the study design. When bioaccessibility results were directly compared with bioavailability results for As (swine and mouse) and Pb (swine), four methods returned results within uncertainty ranges for both elements: two that were defined as simpler (gastric phase only, limited chemicals) and two were more complex (gastric + intestinal phases, with a mixture of chemicals).

Importance of tissue sampling, laboratory methods, and patient characteristics for detection of Pneumocystis in autopsied lungs of non-immunosuppressed individuals.

PubMed

Vargas, S L; Ponce, C; Bustamante, R; Calderón, E; Nevez, G; De Armas, Y; Matos, O; Miller, R F; Gallo, M J

2017-10-01

To understand the epidemiological significance of Pneumocystis detection in a lung tissue sample of non-immunosuppressed individuals, we examined sampling procedures, laboratory methodology, and patient characteristics of autopsy series reported in the literature. Number of tissue specimens, DNA-extraction procedures, age and underlying diagnosis highly influence yield and are critical to understand yield differences of Pneumocystis among reports of pulmonary colonization in immunocompetent individuals.

Validated Method for the Characterization and Quantification of Extractable and Nonextractable Ellagitannins after Acid Hydrolysis in Pomegranate Fruits, Juices, and Extracts.

PubMed

García-Villalba, Rocío; Espín, Juan Carlos; Aaby, Kjersti; Alasalvar, Cesarettin; Heinonen, Marina; Jacobs, Griet; Voorspoels, Stefan; Koivumäki, Tuuli; Kroon, Paul A; Pelvan, Ebru; Saha, Shikha; Tomás-Barberán, Francisco A

2015-07-29

Pomegranates are one of the main highly valuable sources of ellagitannins. Despite the potential health benefits of these compounds, reliable data on their content in pomegranates and derived extracts and food products is lacking, as it is usually underestimated due to their complexity, diversity, and lack of commercially available standards. This study describes a new method for the analysis of the extractable and nonextractable ellagitannins based on the quantification of the acid hydrolysis products that include ellagic acid, gallic acid, sanguisorbic acid dilactone, valoneic acid dilactone, and gallagic acid dilactone in pomegranate samples. The study also shows the occurrence of ellagitannin C-glycosides in pomegranates. The method was optimized using a pomegranate peel extract. To quantify nonextractable ellagitannins, freeze-dried pomegranate fruit samples were directly hydrolyzed with 4 M HCl in water at 90 °C for 24 h followed by extraction of the pellet with dimethyl sulfoxide/methanol (50:50, v/v). The method was validated and reproducibility was assessed by means of an interlaboratory trial, showing high reproducibility across six laboratories with relative standard deviations below 15%. Their applicability was demonstrated in several pomegranate extracts, different parts of pomegranate fruit (husk, peels, and mesocarp), and commercial juices. A large variability has been found in the ellagitannin content (150-750 mg of hydrolysis products/g) and type (gallagic acid/ellagic acid ratios between 4 and 0.15) of the 11 pomegranate extracts studied.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; in-bottle acid digestion of whole-water samples

USGS Publications Warehouse

Hoffman, G.L.; Fishman, M. J.; Garbarino, J.R.

1996-01-01

Water samples for trace-metal determinations routinely have been prepared in open laboratories. For example, the U.S. Geological Survey method I-3485-85 (Extraction Procedure, for Water- Suspended Sediment) is performed in a laboratory hood on a laboratory bench without any special precautions to control airborne contamination. This method tends to be contamination prone for several trace metals primarily because the samples are transferred, acidified, digested, and filtered in an open laboratory environment. To reduce trace-metal contamination of digested water samples, procedures were established that rely on minimizing sample-transfer steps and using a class-100 clean bench during sample filtration. This new procedure involves the following steps: 1. The sample is acidified with HCl directly in the original water-sample bottle. 2. The water-sample bottle with the cap secured is heated in a laboratory oven. 3. The digestate is filtered in a class-100 laminar-flow clean bench. The exact conditions used (that is, oven temperature, time of heating, and filtration methods) for this digestion procedure are described. Comparisons between the previous U.S Geological Survey open-beaker method I-3485-85 and the new in-bottle procedure for synthetic and field-collected water samples are given. When the new procedure is used, blank concentrations for most trace metals determined are reduced significantly.

A rapid monitoring method for inorganic arsenic in rice flour using reversed phase-high performance liquid chromatography-inductively coupled plasma mass spectrometry.

PubMed

Narukawa, Tomohiro; Chiba, Koichi; Sinaviwat, Savarin; Feldmann, Jörg

2017-01-06

A new rapid monitoring method by means of high performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS) following the heat-assisted extraction was developed for measurement of total inorganic arsenic species in rice flour. As(III) and As(V) eluted at the same retention time and completely separated from organoarsenic species by an isocratic elution program on a reversed phase column. Therefore, neither ambiguous oxidation of arsenite to arsenate nor the integration of two peaks were necessary to determine directly the target analyte inorganic arsenic. Rapid injection allowed measuring 3 replicates within 6min and this combined with a quantitative extraction of all arsenic species from rice flour by a 15min HNO 3 -H 2 O 2 extraction makes this the fastest laboratory based method for inorganic arsenic in rice flour. Copyright © 2016 Elsevier B.V. All rights reserved.

Single-laboratory validation of the microplate receptor binding assay for paralytic shellfish toxins in shellfish.

PubMed

Van Dolah, Frances M; Leighfield, Tod A; Doucette, Gregory J; Bean, Laurie; Niedzwiadek, Barbara; Rawn, Dorothea F K

2009-01-01

A single-laboratory validation (SLV) study was conducted for the microplate receptor binding assay (RBA) for paralytic shellfish poisoning (PSP) toxins in shellfish. The basis of the assay is the competition between [3H]saxitoxin (STX) and STX in a standard or sample for binding to the voltage dependent sodium channel. A calibration curve is generated by the addition of 0.01-1000 nM STX, which results in the concentration dependent decrease in [3H]STX-receptor complexes formed and serves to quantify STX in unknown samples. This study established the LOQ, linearity, recovery, accuracy, and precision of the assay for determining PSP toxicity in shellfish extracts, as performed by a single analyst on multiple days. The standard curve obtained on 5 independent days resulted in a half-maximal inhibition (IC50) of 2.3 nM STX +/- 0.3 (RSD = 10.8%) with a slope of 0.96 +/- 0.06 (RSD = 6.3%) and a dynamic range of 1.2-10.0 nM. The LOQ was 5.3 microg STX equivalents/100 g shellfish. Linearity, established by quantification of three levels of purified STX (1.5, 3, and 6 nM), yielded an r2 of 0.97. Recovery from mussels spiked with three levels (40, 80, and 120 microg STX/100 g) averaged 121%. Repeatability (RSD(r)), determined on six naturally contaminated shellfish samples on 5 independent days, was 17.7%. A method comparison with the AOAC mouse bioassay yielded r2 = 0.98 (slope = 1.29) in the SLV study. The effects of the extraction method on RBA-based toxicity values were assessed on shellfish extracted for PSP toxins using the AOAC mouse bioassay method (0.1 M HCI) compared to that for the precolumn oxidation HPLC method (0.1% acetic acid). The two extraction methods showed linear correlation (r2 = 0.99), with the HCl extraction method yielding slightly higher toxicity values (slope = 1.23). A similar relationship was observed between HPLC quantification of the HCI- and acetic acid-extracted samples (r2 = 0.98, slope 1.19). The RBA also had excellent linear correlation with HPLC analyses (r2 = 0.98 for HCl, r2 = 0.99 for acetic acid), but gave somewhat higher values than HPLC using either extraction method (slope = 1.39 for HCl extracts, slope = 1.32 for acetic acid). Overall, the excellent linear correlations with the both mouse bioassay and HPLC method and sufficient interassay repeatability suggest that the RBA can be effective as a high throughput screen for estimating PSP toxicity in shellfish.

On-Site Molecular Detection of Soil-Borne Phytopathogens Using a Portable Real-Time PCR System

PubMed Central

DeShields, Joseph B.; Bomberger, Rachel A.; Woodhall, James W.; Wheeler, David L.; Moroz, Natalia; Johnson, Dennis A.; Tanaka, Kiwamu

2018-01-01

On-site diagnosis of plant diseases can be a useful tool for growers for timely decisions enabling the earlier implementation of disease management strategies that reduce the impact of the disease. Presently in many diagnostic laboratories, the polymerase chain reaction (PCR), particularly real-time PCR, is considered the most sensitive and accurate method for plant pathogen detection. However, laboratory-based PCRs typically require expensive laboratory equipment and skilled personnel. In this study, soil-borne pathogens of potato are used to demonstrate the potential for on-site molecular detection. This was achieved using a rapid and simple protocol comprising of magnetic bead-based nucleic acid extraction, portable real-time PCR (fluorogenic probe-based assay). The portable real-time PCR approach compared favorably with a laboratory-based system, detecting as few as 100 copies of DNA from Spongospora subterranea. The portable real-time PCR method developed here can serve as an alternative to laboratory-based approaches and a useful on-site tool for pathogen diagnosis. PMID:29553557

What Is in Your Wallet? Quantitation of Drugs of Abuse on Paper Currency with a Rapid LC-MS/MS Method

ERIC Educational Resources Information Center

Parker, Patrick D.; Beers, Brandon; Vergne, Matthew J.

2017-01-01

Laboratory experiments were developed to introduce students to the quantitation of drugs of abuse by high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). Undergraduate students were introduced to internal standard quantitation and the LC-MS/MS method optimization for cocaine. Cocaine extracted from paper currency was…

Analysis of food taints and off-flavours: a review.

PubMed

Ridgway, K; Lalljie, S P D; Smith, R M

2010-02-01

Taints and off-flavours in foods are a major concern to the food industry. Identification of the compound(s) causing a taint or off-flavour in food and accurate quantification are critical in assessing the potential safety risks of a product or ingredient. Even when the tainting compound(s) are not at a level that would cause a safety concern, taints and off-flavours can have a significant impact on the quality and consumers' acceptability of products. The analysis of taints and off-flavour compounds presents an analytical challenge especially in an industrial laboratory environment because of the low levels, often complex matrices and potential for contamination from external laboratory sources. This review gives an outline of the origins of chemical taints and off-flavours and looks at the methods used for analysis and the merits and drawbacks of each technique. Extraction methods and instrumentation are covered along with possible future developments. Generic screening methods currently lack the sensitivity required to detect the low levels required for some tainting compounds and a more targeted approach is often required. This review highlights the need for a rapid but sensitive universal method of extraction for the unequivocal determination of tainting compounds in food.

A simplified method for extracting microplastics in laboratory-cultured coral and water samples

EPA Science Inventory

The deleterious effects of microplastic ingestion by marine organisms and the impacts across trophic levels are widely unknown. Due to the extensive ecological and socioeconomic benefits provided by coral reefs, investigating the impacts of microplastics on reef ecosystems is of ...

MUTAGENIC PROPERTIES OF RIVER WATERS FLOWING THROUGH BIG CITY AREAS IN NORTH AMERICA

EPA Science Inventory

The hanging technique using Blue rayon, which specifically adsorbs mutagens with multicyclic planer structures, has advantages over the conventional method of bringing-large volumes of water back to the laboratory for extraction. Therefore, the analysis of many samples from remot...

New targets for expedient detection of viruses within shellfish

USDA-ARS?s Scientific Manuscript database

Previously our laboratory developed an expedient method for extraction of viral RNA from food-borne virus contaminated bivalve shellfish, termed the GPTT protocol. This protocol utilizes either whole shellfish or dissected digestive diverticula. This four step protocol utilizes a high pH glycine or...

Evaluation of extraction methods for hexavalent chromium determination in dusts, ashes, and soils

USGS Publications Warehouse

Wolf, Ruth E.; Wilson, Stephen A.

2010-01-01

One of the difficulties in performing speciation analyses on solid samples is finding a suitable extraction method. Traditional methods for extraction of hexavalent chromium, Cr(VI), in soils, such as SW846 Method 3060A, can be tedious and are not always compatible with some determination methods. For example, the phosphate and high levels of carbonate and magnesium present in the U.S. Environmental Protection Agency (USEPA) Method 3060A digestion for Cr(VI) were found to be incompatible with the High Performance Liquid Chromatography-Inductively Coupled Plasma-Mass Spectrometry (HPLC-ICP-MS) detection method used by our laboratory. Modification of Method 3060A by eliminating the use of the phosphate buffer provided improved performance with the detection method, however dilutions are still necessary to achieve good chromatographic separation and detection of Cr(VI). An ultrasonic extraction method using a 1 mM Na2CO3 - 9 mM NaHCO3 buffer solution, adapted from Occupational Safety and Health Administration (OSHA) Method ID215, has been used with good results for the determination of Cr(VI) in air filters. The average recovery obtained for BCR-545 - Welding Dust Loaded on Filter (IRMM, Belgium) using this method was 99 percent (1.2 percent relative standard deviation) with no conversion of Cr(VI) to Cr(III) during the extraction process. This ultrasonic method has the potential for use with other sample matrices, such as ashes and soils. Preliminary investigations using NIST 2701 (Hexavalent Chromium in Contaminated Soil) loaded onto quartz filters showed promising results with approximately 90 percent recovery of the certified Cr(VI) value. Additional testing has been done using NIST 2701 and NIST 2700 using different presentation methods. Extraction efficiency of bulk presentation, where small portions of the sample are added to the bottom of the extraction vessel, will be compared with supported presentation, where small portions of the sample are loaded onto a quartz filter prior to extraction. In addition, results obtained from the standard grinding preparation of NIST 2701 and NIST 2700 will be compared with micronizing to reduce particle size before extraction.

Evaluation of the antioxidants activities of four Slovene medicinal plant species by traditional and novel biosensory assays.

PubMed

Kintzios, Spiridon; Papageorgiou, Katerina; Yiakoumettis, Iakovos; Baricevic, Dea; Kusar, Anita

2010-11-02

We investigated the antioxidant activity of methanolic and water extracts of Slovene accessions of four medicinal plant species (Salvia officinalis, Achillea millefolium, Origanum vulgare subsp. vulgare and Gentiana lutea). Their free radical-scavenging activity against the DPPH. free radical was studied with a spectrophotometric assay, while their biological activity with the help of a laboratory-made biosensor based on immobilized fibroblast cells (assay duration: 3 min). The observed antioxidant activity of the extracts from the four investigated medicinal plant species was dependent on both the solvent used for extraction and the assay method (conventional or biosensor-based). Independently from the assay method and the solvent used for extraction, the lowest scavenging activity was observed in root extracts of G. lutea. Treatment of the immobilized cells with the plant extracts resulted in an increase of the cell membrane potential (membrane hyperpolarization), possibly due to the reduction of membrane damage due to oxidation. The novel cell biosensor could be utilized as a rapid, high throughput tool for screening the antioxidant properties of plant-derived compounds. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Development of a modified cortisol extraction procedure for intermediately sized fish not amenable to whole-body or plasma extraction methods.

PubMed

Guest, Taylor W; Blaylock, Reginald B; Evans, Andrew N

2016-02-01

The corticosteroid hormone cortisol is the central mediator of the teleost stress response. Therefore, the accurate quantification of cortisol in teleost fishes is a vital tool for addressing fundamental questions about an animal's physiological response to environmental stressors. Conventional steroid extraction methods using plasma or whole-body homogenates, however, are inefficient within an intermediate size range of fish that are too small for phlebotomy and too large for whole-body steroid extractions. To assess the potential effects of hatchery-induced stress on survival of fingerling hatchery-reared Spotted Seatrout (Cynoscion nebulosus), we developed a novel extraction procedure for measuring cortisol in intermediately sized fish (50-100 mm in length) that are not amenable to standard cortisol extraction methods. By excising a standardized portion of the caudal peduncle, this tissue extraction procedure allows for a small portion of a larger fish to be sampled for cortisol, while minimizing the potential interference from lipids that may be extracted using whole-body homogenization procedures. Assay precision was comparable to published plasma and whole-body extraction procedures, and cortisol quantification over a wide range of sample dilutions displayed parallelism versus assay standards. Intra-assay %CV was 8.54%, and average recovery of spiked samples was 102%. Also, tissue cortisol levels quantified using this method increase 30 min after handling stress and are significantly correlated with blood values. We conclude that this modified cortisol extraction procedure provides an excellent alternative to plasma and whole-body extraction procedures for intermediately sized fish, and will facilitate the efficient assessment of cortisol in a variety of situations ranging from basic laboratory research to industrial and field-based environmental health applications.

Microbial Abundances in Salt Marsh Soils: A Molecular Approach for Small Spatial Scales

NASA Astrophysics Data System (ADS)

Granse, Dirk; Mueller, Peter; Weingartner, Magdalena; Hoth, Stefan; Jensen, Kai

2016-04-01

The rate of biological decomposition greatly determines the carbon sequestration capacity of salt marshes. Microorganisms are involved in the decomposition of biomass and the rate of decomposition is supposed to be related to microbial abundance. Recent studies quantified microbial abundance by means of quantitative polymerase chain reaction (QPCR), a method that also allows determining the microbial community structure by applying specific primers. The main microbial community structure can be determined by using primers specific for 16S rRNA (Bacteria) and 18S rRNA (Fungi) of the microbial DNA. However, the investigation of microbial abundance pattern at small spatial scales, such as locally varying abiotic conditions within a salt-marsh system, requires high accuracy in DNA extraction and QPCR methods. Furthermore, there is evidence that a single extraction may not be sufficient to reliably quantify rRNA gene copies. The aim of this study was to establish a suitable DNA extraction method and stable QPCR conditions for the measurement of microbial abundances in semi-terrestrial environments. DNA was extracted from two soil samples (top WE{5}{cm}) by using the PowerSoil DNA Extraction Kit (Mo Bio Laboratories, Inc., Carlsbad, CA) and applying a modified extraction protocol. The DNA extraction was conducted in four consecutive DNA extraction loops from three biological replicates per soil sample by reusing the PowerSoil bead tube. The number of Fungi and Bacteria rRNA gene copies of each DNA extraction loop and a pooled DNA solution (extraction loop 1 - 4) was measured by using the QPCR method with taxa specific primer pairs (Bacteria: B341F, B805R; Fungi: FR1, FF390). The DNA yield of the replicates varied at DNA extraction loop 1 between WE{25 and 85}{ng

Anti-ulcer and ulcer healing potentials of Musa sapientum peel extract in the laboratory rodents

PubMed Central

Onasanwo, Samuel Adetunji; Emikpe, Benjamin Obukowho; Ajah, Austin Azubuike; Elufioye, Taiwo Olayemi

2013-01-01

Background: This study investigated the anti-ulcer and ulcer healing potentials of the methanol extract of Musa sapientum peel in the laboratory rats. Materials and Methods: Methanol extract of the peels on Musa sapientum (MEMS) was evaluated for its anti-ulcer using alcohol-induced, aspirin-induced, and pyloric ligation-induced models, and for its ulcer healing employing acetic acid-induced ulcer models in rats. Results: The findings from this experiment showed that MEMS (50, 100 and 200 mg/kg, b.w.) anti-ulcer and ulcer healing activity (P ≤ 0.05) is dose-dependent. Also, MEMS exhibited healing of the ulcer base in all the treated groups when compared with the control group. Conclusion: The outcomes of this experiment revealed that the anti-ulcer effect of MEMS may be due to its anti-secretory and cyto-protective activity. The healing of the ulcer base might not be unconnected with basic fibroblast growth factors responsible for epithelial regeneration. PMID:23900937

Piscicidal activity of leaf and bark extract of Thevetia peruviana plant and their biochemical stress response on fish metabolism.

PubMed

Singh, S K; Yadav, R P; Singh, A

2010-11-01

The leaf and bark of Thevetia peruviana (Family: Apocynaceae) plant was administered for 24 h to the freshwater fish Catla catla (Hamilton) to evaluate their piscicidal activity in laboratory and cemented pond condition. The LC0 values of lef and bark extracts of different solvents (i.e., acetone, diethyl ether, ethyl alcohol, chloroform and carbon tetrachloride) of this plant to fish Catla catla were determined. The LC50 values of acetone leaf extract of Thevetia peruviana plant is 88.80 mg/L (24h) in laboratory condition and 529.38 mg/L (24h) in cemented pond condition; acetone bark extract of this plant is 99.43 mg/L (24h) in laboratory condition and 591.78 mg/L (24h) in cemented pond condition against freshwater fish Catla catla. Similar trend was also observed in case of other solvent (i.e., diethyl ether, ethyl alcohol, chloroform and carbon tetrachloride) of leaf and bark extracts of Thevetia peruviana plant against freshwater fish Catla catla in laboratory and cemented pond conditions. The acetone leaf and bark extract of this plant was very effective in comparison to other solvent extract in both the conditions. So, the biochemical analysis is taken only acetone leaf and bark extract of Thevetia peruviana plant in laboratory condition. Exposure of sub-lethal doses (40% and 80% of LC,) of acetone leaf and bark extract of this plant over 24 h caused significant (P < 0.05) alterations in total protein, free amino acids, DNA & RNA, protease and acid and alkaline phosphatase activity in muscle, liver and gonadal tissues of fish Catla catla in laboratory condition.

Molecular identification of polymers and anthropogenic particles extracted from oceanic water and fish stomach - A Raman micro-spectroscopy study.

PubMed

Ghosal, Sutapa; Chen, Michael; Wagner, Jeff; Wang, Zhong-Min; Wall, Stephen

2018-02-01

Pacific Ocean trawl samples, stomach contents of laboratory-raised fish as well as fish from the subtropical gyres were analyzed by Raman micro-spectroscopy (RMS) to identify polymer residues and any detectable persistent organic pollutants (POP). The goal was to access specific molecular information at the individual particle level in order to identify polymer debris in the natural environment. The identification process was aided by a laboratory generated automated fluorescence removal algorithm. Pacific Ocean trawl samples of plastic debris associated with fish collection sites were analyzed to determine the types of polymers commonly present. Subsequently, stomach contents of fish from these locations were analyzed for ingested polymer debris. Extraction of polymer debris from fish stomach using KOH versus ultrapure water were evaluated to determine the optimal method of extraction. Pulsed ultrasonic extraction in ultrapure water was determined to be the method of choice for extraction with minimal chemical intrusion. The Pacific Ocean trawl samples yielded primarily polyethylene (PE) and polypropylene (PP) particles >1 mm, PE being the most prevalent type. Additional microplastic residues (1 mm - 10 μm) extracted by filtration, included a polystyrene (PS) particle in addition to PE and PP. Flame retardant, deca-BDE was tentatively identified on some of the PP trawl particles. Polymer residues were also extracted from the stomachs of Atlantic and Pacific Ocean fish. Two types of polymer related debris were identified in the Atlantic Ocean fish: (1) polymer fragments and (2) fragments with combined polymer and fatty acid signatures. In terms of polymer fragments, only PE and PP were detected in the fish stomachs from both locations. A variety of particles were extracted from oceanic fish as potential plastic pieces based on optical examination. However, subsequent RMS examination identified them as various non-plastic fragments, highlighting the importance of chemical analysis in distinguishing between polymer and non-polymer residues. Copyright © 2017 Elsevier Ltd. All rights reserved.

Processing Protocol for Soil Samples Potentially ...

EPA Pesticide Factsheets

Method Operating Procedures This protocol describes the processing steps for 45 g and 9 g soil samples potentially contaminated with Bacillus anthracis spores. The protocol is designed to separate and concentrate the spores from bulk soil down to a pellet that can be used for further analysis. Soil extraction solution and mechanical shaking are used to disrupt soil particle aggregates and to aid in the separation of spores from soil particles. Soil samples are washed twice with soil extraction solution to maximize recovery. Differential centrifugation is used to separate spores from the majority of the soil material. The 45 g protocol has been demonstrated by two laboratories using both loamy and sandy soil types. There were no significant differences overall between the two laboratories for either soil type, suggesting that the processing protocol would be robust enough to use at multiple laboratories while achieving comparable recoveries. The 45 g protocol has demonstrated a matrix limit of detection at 14 spores/gram of soil for loamy and sandy soils.

Modified procedure to determine acid-insoluble lignin in wood and pulp

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

Effland, M.J.

1977-10-01

If wood is treated with strong acid, carbohydrates are hydrolyzed and solubilized. The insoluble residue is by definition lignin and can be measured gravimetrically. The standard method of analysis requires samples of 1 or 2 g of wood or pulp. In research at this laboratory these amounts of sample are often not available for analytical determinations. Thus we developed a modification of the standard procedure suitable for much smaller sample amounts. The modification is based on the procedure of Saeman. Wood samples require extraction prior to lignin analysis to remove acid-insoluble extractives that will be measured as lignin. Usually thismore » involves only a standard extraction with ethanol--benzene. However, woods high in tannin must also be subjected to extraction with alcohol. Pulps seldom require extraction.« less

Pomegranate extract exhibits in vitro activity against Clostridium difficile.

PubMed

Finegold, Sydney M; Summanen, Paula H; Corbett, Karen; Downes, Julia; Henning, Susanne M; Li, Zhaoping

2014-10-01

To determine the possible utility of pomegranate extract in the management or prevention of Clostridium difficile infections or colonization. The activity of pomegranate was tested against 29 clinical C. difficile isolates using the Clinical and Laboratory Standards Institute-approved agar dilution technique. Total phenolics content of the pomegranate extract was determined by Folin-Ciocalteau colorimetric method and final concentrations of 6.25 to 400 μg/mL gallic acid equivalent were achieved in the agar. All strains had MICs at 12.5 to 25 mg/mL gallic acid equivalent range. Our results suggest antimicrobial in vitro activity for pomegranate extract against toxigenic C. difficile. Pomegranate extract may be a useful contributor to the management and prevention of C. difficile disease or colonization. Copyright © 2014 Elsevier Inc. All rights reserved.

Preparation of bone samples in the Gliwice Radiocarbon Laboratory for AMS radiocarbon dating.

PubMed

Piotrowska, N; Goslar, T

2002-12-01

In the Gliwice Radiocarbon Laboratory, a system for preparation of samples for AMS dating has been built. At first it was used to produce graphite targets from plant macrofossils and sediments. In this study we extended its capabilities with the preparation of bones. We dealt with 3 methods; the first was the classical Longin method of collagen extraction, the second one included additional treatment of powdered bone in alkali solution, while in the third one carboxyl carbon was separated from amino acids obtained after hydrolysis of protein. The suitability of the methods was tested on 2 bone samples. Most of our samples gave ages > 40 kyr BP, suggesting good performance of the adapted methods, except for one sample prepared with simple Longin method. For routine preparation of bones we chose the Longin method with additional alkali treatment.

Development of analytical methodologies to assess recalcitrant pesticide bioremediation in biobeds at laboratory scale.

PubMed

Rivero, Anisleidy; Niell, Silvina; Cerdeiras, M Pía; Heinzen, Horacio; Cesio, María Verónica

2016-06-01

To assess recalcitrant pesticide bioremediation it is necessary to gradually increase the complexity of the biological system used in order to design an effective biobed assembly. Each step towards this effective biobed design needs a suitable, validated analytical methodology that allows a correct evaluation of the dissipation and bioconvertion. Low recovery yielding methods could give a false idea of a successful biodegradation process. To address this situation, different methods were developed and validated for the simultaneous determination of endosulfan, its main three metabolites, and chlorpyrifos in increasingly complex matrices where the bioconvertor basidiomycete Abortiporus biennis could grow. The matrices were culture media, bran, and finally a laboratory biomix composed of bran, peat and soil. The methodology for the analysis of the first evaluated matrix has already been reported. The methodologies developed for the other two systems are presented in this work. The targeted analytes were extracted from fungi growing over bran in semisolid media YNB (Yeast Nitrogen Based) with acetonitrile using shaker assisted extraction, The salting-out step was performed with MgSO4 and NaCl, and the extracts analyzed by GC-ECD. The best methodology was fully validated for all the evaluated analytes at 1 and 25mgkg(-1) yielding recoveries between 72% and 109% and RSDs <11% in all cases. The application of this methodology proved that A. biennis is able to dissipate 94% of endosulfan and 87% of chlorpyrifos after 90 days. Having assessed that A. biennis growing over bran can metabolize the studied pesticides, the next step faced was the development and validation of an analytical procedure to evaluate the analytes in a laboratory scale biobed composed of 50% of bran, 25% of peat and 25% of soil together with fungal micelium. From the different procedures assayed, only ultrasound assisted extraction with ethyl acetate allowed recoveries between 80% and 110% with RSDs <18%. Linearity, recovery, precision, matrix effect and LODs/LOQs of each method were studied for all the analytes: endosulfan isomers (α & β) and its metabolites (endosulfan sulfate, ether and diol) as well as for chlorpyrifos. In the first laboratory evaluation of these biobeds endosulfan was bioconverted up to 87% and chlorpyrifos more than 79% after 27 days. Copyright © 2016 Elsevier B.V. All rights reserved.

First external quality assurance program of the Italian HLA-B*57:01 Network assessing the performance of clinical virology laboratories in HLA-B*57:01 testing.

PubMed

Meini, Genny; Dello Russo, Cinzia; Allice, Tiziano; Barresi, Renata; D'Arrigo, Roberta; Falasca, Francesca; Lipsi, Maria Rosaria; Paolucci, Stefania; Zanussi, Stefania; Antonetti, Raffaele; Baldanti, Fausto; Basaglia, Giancarlo; Bruzzone, Bianca; Polilli, Ennio; Ghisetti, Valeria; Pucillo, Leopoldo Paolo; Turriziani, Ombretta; Pirazzoli, Antonella; Navarra, Pierluigi; Zazzi, Maurizio

2016-05-01

Since the HLA-B*57:01 allele is strongly associated with abacavir hypersensitivity reaction, testing for the presence of HLA-B*57:01 is mandatory before administration of abacavir. While HLA-B*57:01 testing is usually provided by pharmacogenetics, genetics or blood transfusion services, clinical virology laboratories can be an optimal opportunity for HLA-B*57:01 testing since they receive blood samples for routine HIV monitoring and have the expertise for convenient and less expensive PCR-based point mutation assays. The Italian HLA-B*57:01 Network gathers accredited clinical virology laboratories offering HLA-B*57:01 testing in Italy with the aim to share protocols, test new methods, develop and maintain external quality assurance (EQA) programs. A panel of 9HLA-B*57:01-positive and 16HLA-B*57:01-negative frozen blood samples were blindly distributed to 10 units including 9 clinical virology laboratories and one reference pharmacology laboratory. Each laboratory was free to use its own routine method for DNA extraction and HLA-B*57:01 testing. DNA was extracted by automated workstations in 6 units and by manual spin columns in 4. Eight units used the Duplicα Real Time HLA-B*57:01 kit by Euroclone and two units used two different PCR homemade protocols. All the 10 units correctly identified all the 25 samples. The first HLA-B*57:01 EQA program run in Italy showed that clinical virology units are equipped and proficient for providing HLA-B*57:01 testing by inexpensive assays easy to integrate into their routine. Copyright © 2016 Elsevier B.V. All rights reserved.

An evaluation of condition indices for birds

USGS Publications Warehouse

Johnson, D.H.; Krapu, G.L.; Reinecke, K.J.; Jorde, Dennis G.

1985-01-01

A Lipid Index, the ratio of fat to fat-free dry weight, is proposed as a measure of fat stores in birds. The estimation of the index from field measurements of live birds is illustrated with data on the sandhill crane (Grus canadensis) and greater white-fronted goose (Anser albifrons). Of the various methods of assessing fat stores, lipid extraction is the most accurate but also the most involved. Water extraction is a simpler laboratory method that provides a good index to fat and can be calibrated to serve as an estimator. Body weight itself is often inadequate as a condition index, but scaling by morphological measurements can markedly improve its value.

Ring trial 2016 for Bluetongue virus detection by real-time RT-PCR in France.

PubMed

Sailleau, Corinne; Viarouge, Cyril; Breard, Emmanuel; Vitour, Damien; Zientara, Stephan

2017-05-01

Since the unexpected emergence of BTV-8 in Northern Europe and the incursion of BTV-8 and 1 in France in 2006-2007, molecular diagnosis has considerably evolved. Several real-time RT-PCR (rtRT-PCR) methods have been developed and published, and are currently being used in many countries across Europe for BTV detection and typing. In France, the national reference laboratory (NRL) for orbiviruses develops and validates 'ready-to-use' kits with private companies for viral RNA detection. The regional laboratories network that was set up to deal with a heavy demand for analyses has used these available kits. From 2007, ring tests were organized to monitor the performance of the French laboratories. This study presents the results of 63 regional laboratories in the ring trial organized in 2016. Blood samples were sent to the laboratories. Participants were asked to use the rtRT-PCR methods in place in their laboratory, for detection of all BTV serotypes and specifically BTV-8. The French regional laboratories are able to detect and genotype BTV in affected animals. Despite the use of several methods (i.e. RNA extraction and different commercial rtRT-PCRs), the network is homogeneous. The ring trial demonstrated that the French regional veterinary laboratories have reliable and robust BTV diagnostic tools for BTV genome detection.

Quick, easy, cheap, effective, rugged, and safe sample preparation approach for pesticide residue analysis using traditional detectors in chromatography: A review.

PubMed

Rahman, Md Musfiqur; Abd El-Aty, A M; Kim, Sung-Woo; Shin, Sung Chul; Shin, Ho-Chul; Shim, Jae-Han

2017-01-01

In pesticide residue analysis, relatively low-sensitivity traditional detectors, such as UV, diode array, electron-capture, flame photometric, and nitrogen-phosphorus detectors, have been used following classical sample preparation (liquid-liquid extraction and open glass column cleanup); however, the extraction method is laborious, time-consuming, and requires large volumes of toxic organic solvents. A quick, easy, cheap, effective, rugged, and safe method was introduced in 2003 and coupled with selective and sensitive mass detectors to overcome the aforementioned drawbacks. Compared to traditional detectors, mass spectrometers are still far more expensive and not available in most modestly equipped laboratories, owing to maintenance and cost-related issues. Even available, traditional detectors are still being used for analysis of residues in agricultural commodities. It is widely known that the quick, easy, cheap, effective, rugged, and safe method is incompatible with conventional detectors owing to matrix complexity and low sensitivity. Therefore, modifications using column/cartridge-based solid-phase extraction instead of dispersive solid-phase extraction for cleanup have been applied in most cases to compensate and enable the adaptation of the extraction method to conventional detectors. In gas chromatography, the matrix enhancement effect of some analytes has been observed, which lowers the limit of detection and, therefore, enables gas chromatography to be compatible with the quick, easy, cheap, effective, rugged, and safe extraction method. For liquid chromatography with a UV detector, a combination of column/cartridge-based solid-phase extraction and dispersive solid-phase extraction was found to reduce the matrix interference and increase the sensitivity. A suitable double-layer column/cartridge-based solid-phase extraction might be the perfect solution, instead of a time-consuming combination of column/cartridge-based solid-phase extraction and dispersive solid-phase extraction. Therefore, replacing dispersive solid-phase extraction with column/cartridge-based solid-phase extraction in the cleanup step can make the quick, easy, cheap, effective, rugged, and safe extraction method compatible with traditional detectors for more sensitive, effective, and green analysis. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

COMPARISON OF FIVE EXTRACTION METHODS ON INCURRED AND FORTIFIED PESTICIDES IN COMPOSITE DIETS: BLENDER, SOXHLET, ASE, MICROWAVE AND SFE

EPA Science Inventory

The USEPA National Exposure Research Laboratory studies dietary exposure to a diverse group of semi-volatile pesticides by analyzing 24 hour duplicate composite diets. The pesticides of interest include organochlorines, organophosphates, anilines, and triazines. Currently, there ...

EVALUATION OF DISPOSABLE DIAPERS FOR QUANTATIVE MEASUREMENTS OF PESTICIDE METABOLITES AND CREATININE IN URINE SAMPLES

EPA Science Inventory

This project consisted of a laboratory study to evaluate an extraction and analysis method for quantifying biomarkers of pesticide exposure and creatinine in urine samples collected with commercially-available disposable diapers. For large exposure studies, such as the National ...

Compression and Associated Properties of Boron Carbide

DTIC Science & Technology

2008-12-01

of Brookhaven National Laboratory ( BNL ) with the light extracted from the x-ray source in a method previously described (Liu et al., 2002).. A...are sharper and more intense relative to as received material. ACKNOWLEDGMENTS Thanks are due to Dr. Jingzhu Hu and Dr. Zhenxian Liu of BNL

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of methylene blue active substances by spectrophotometry

USGS Publications Warehouse

Burkhardt, Mark R.; Cinotto, Pete J.; Frahm, Galen W.; Woodworth, Mark T.; Pritt, Jeffrey W.

1995-01-01

A method for the determination of methylene blue active substances in whole-water samples by liquid-liquid extraction and spectrophotometric detection is described. Sulfate and sulfonate-based surfectants are reacted with methylene blue to form a blue-colored complex. The complex is extracted into chloroform, back-washed with an acidified phosphate-based buffer solution, and measured against external standards with a probe spectrophotometer. The method detection limt for routine analysis is 0.02 milligram per liter. The precision is plus/minus 10 percent relative standard deviation. The positive bias from nitrate and chloride and U.S. Geological Survey method O-3111-83 for methylene blue active substances is minized by adding a back-washing step.

Optimization of Polycyclic Aromatic Hydrocarbon (PAH) Extraction Efficiency Parameters for Sub- and Supercritical Water Extraction (SCWE) Instrument

NASA Technical Reports Server (NTRS)

Okada, Asahi A.

2005-01-01

Polycyclic aromatic hydrocarbons are a class of molecules composed of multiple, bonded benzene rings. As PAHS are believed to be present on Mars, positive confirmation of their presence on Mars is highly desirable. To extract PAHS, which have low volatility, a fluid extraction method is ideal, and one that does not utilize organic solvents is especially ideal for in situ instrumental analysis. The use of water as a solvent, which at subcritical pressures and temperatures is relatively non-Polar, has significant potential. As SCWE instruments have not yet been commercialized, all instruments are individually-built research prototypes: thus, initial efforts were intended to determine if extraction efficiencies on the JPL-built laboratory-scale SCWE instrument are comparable to differing designs built elsewhere. Samples of soil with certified reference concentrations of PAHs were extracted using SCWE as well as conventional Soxhlet extraction. Continuation of the work would involve extractions on JPL'S newer, portable SCWE instrument prototype to determine its efficiency in extracting PAHs.

Preparation and use of Xenopus egg extracts to study DNA replication and chromatin associated proteins

PubMed Central

Gillespie, Peter J.; Gambus, Agnieszka; Blow, J. Julian

2012-01-01

The use of cell-free extracts prepared from eggs of the South African clawed toad, Xenopus laevis, has led to many important discoveries in cell cycle research. These egg extracts recapitulate the key nuclear transitions of the eukaryotic cell cycle in vitro under apparently the same controls that exist in vivo. DNA added to the extract is first assembled into a nucleus and is then efficiently replicated. Progression of the extract into mitosis then allows the separation of paired sister chromatids. The Xenopus cell-free system is therefore uniquely suited to the study of the mechanisms, dynamics and integration of cell cycle regulated processes at a biochemical level. In this article we describe methods currently in use in our laboratory for the preparation of Xenopus egg extracts and demembranated sperm nuclei for the study of DNA replication in vitro. We also detail how DNA replication can be quantified in this system. In addition, we describe methods for isolating chromatin and chromatin-bound protein complexes from egg extracts. These recently developed and revised techniques provide a practical starting point for investigating the function of proteins involved in DNA replication. PMID:22521908

Quantitative 31P NMR for Simultaneous Trace Analysis of Organophosphorus Pesticides in Aqueous Media Using the Stir Bar Sorptive Extraction Method

NASA Astrophysics Data System (ADS)

Ansari, S.; Talebpour, Z.; Molaabasi, F.; Bijanzadeh, H. R.; Khazaeli, S.

2016-09-01

The analysis of pesticides in water samples is of primary concern for quality control laboratories due to the toxicity of these compounds and their associated public health risk. A novel analytical method based on stir bar sorptive extraction (SBSE), followed by 31P quantitative nuclear magnetic resonance (31P QNMR), has been developed for simultaneously monitoring and determining four organophosphorus pesticides (OPPs) in aqueous media. The effects of factors on the extraction efficiency of OPPs were investigated using a Draper-Lin small composite design. An optimal sample volume of 4.2 mL, extraction time of 96 min, extraction temperature of 42°C, and desorption time of 11 min were obtained. The results showed reasonable linearity ranges for all pesticides with correlation coefficients greater than 0.9920. The limit of quantification (LOQ) ranged from 0.1 to 2.60 mg/L, and the recoveries of spiked river water samples were from 82 to 94% with relative standard deviation (RSD) values less than 4%. The results show that this method is simple, selective, rapid, and can be applied to other sample matrices.

Method of analysis and quality-assurance practices for determination of pesticides in water by solid-phase extraction and capillary-column gas chromatography/mass spectrometry at the U.S. Geological Survey California District Organic Chemistry Laboratory, 1996-99

USGS Publications Warehouse

Crepeau, Kathryn L.; Baker, Lucian M.; Kuivila, Kathryn

2000-01-01

A method of analysis and quality-assurance practices were developed to study the fate and transport of pesticides in the San Francisco Bay-Estuary by the U.S. Geological Survey. Water samples were filtered to remove suspended-particulate matter and pumped through C-8 solid-phase extraction cartridges to extract the pesticides. The cartridges were dried with carbon dioxide and the pesticides were eluted with three cartridge volumes of hexane:diethyl ether (1:1) solution. The eluants were analyzed using capillary-column gas chromatography/mass spectrometry in full-scan mode. Method detection limits for pesticides ranged from 0.002 to 0.025 microgram per liter for 1-liter samples. Recoveries ranged from 44 to 140 percent for 25 pesticides in samples of organic-free reagent water and Sacramento-San Joaquin Delta and Suisun Bay water fortified at 0.05 and 0.50 microgram per liter. The estimated holding time for pesticides after extraction on C-8 solid-phase extraction cartridges ranged from 10 to 257 days.

Rapid extraction and assay of uranium from environmental surface samples

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

Barrett, Christopher A.; Chouyyok, Wilaiwan; Speakman, Robert J.

Extraction methods enabling faster removal and concentration of uranium compounds for improved trace and low-level assay are demonstrated for standard surface sampling material in support of nuclear safeguards efforts, health monitoring, and other nuclear analysis applications. A key problem with the existing surface sampling swipes is the requirement for complete digestion of sample and sampling matrix. This is a time-consuming and labour-intensive process that limits laboratory throughput, elevates costs, and increases background levels. Various extraction methods are explored for their potential to quickly and efficiently remove different chemical forms of uranium from standard surface sampling material. A combination of carbonatemore » and peroxide solutions is shown to give the most rapid and complete form of uranyl compound extraction and dissolution. This rapid extraction process is demonstrated to be compatible with standard inductive coupled plasma mass spectrometry methods for uranium isotopic assay as well as screening techniques such as x-ray fluorescence. The general approach described has application beyond uranium to other analytes of nuclear forensic interest (e.g., rare earth elements and plutonium) as well as heavy metals for environmental and industrial hygiene monitoring.« less

Impact of pH on the stability and the cross-reactivity of ochratoxin A and citrinin.

PubMed

Bazin, Ingrid; Faucet-Marquis, Virginie; Monje, Marie-Carmen; El Khoury, Micheline; Marty, Jean-Louis; Pfohl-Leszkowicz, Annie

2013-11-28

Mycotoxins are secondary metabolites produced by several fungi contaminating crops. In several countries, the maximum permitted levels of mycotoxins are found in foodstuffs and feedstuffs. The common strategy of mycotoxin analysis involves extraction, clean-up and quantification by chromatography. In this paper, we analyzed the reasons of underestimation of ochratoxin A (OTA) content in wine, and overestimation of OTA in wheat, depending on the pH of the clean-up step and the simultaneous presence of citrinin (CIT). We demonstrated that the increase of pH by adding polyethylene glycol (PEG) to wine led to an underestimation of OTA by conversion of OTA into open ring ochratoxin A OP-OA. In comparing three methods of extraction and clean-up for the determination of OTA and CIT in wheat--(i) an inter-laboratory validated method for OTA in cereals using immunoaffinity column clean-up (IAC) and extraction by acetonitrile/water; (ii) a validated method using IAC and extraction with 1% bicarbonate Na; and (iii) an in-house validated method based on acid liquid/liquid extraction--we observed an overestimation of OTA after immunoaffinity clean-up when CIT is also present in the sample, whereas an underestimation was observed when OTA was alone. Under neutral and alkaline conditions, CIT was partially recognized by OTA antibodies.

Qualitative and quantitative assessment of single fingerprints in forensic DNA analysis.

PubMed

Ostojic, Lana; Klempner, Stacey A; Patel, Rosni A; Mitchell, Adele A; Axler-DiPerte, Grace L; Wurmbach, Elisa

2014-11-01

Fingerprints and touched items are important sources of DNA for STR profiling, since this evidence can be recovered in a wide variety of criminal offenses. However, there are some fundamental difficulties in working with these samples, including variability in quantity and quality of extracted DNA. In this study, we collected and analyzed over 700 fingerprints. We compared a commercially available extraction protocol (Zygem) to two methods developed in our laboratory, a simple one-tube protocol and a high sensitivity protocol (HighSens) that includes additional steps to concentrate and purify the DNA. The amplification protocols tested were AmpFLSTR® Identifiler® using either 28 or 31 amplification cycles, and Identifiler® Plus using 32 amplification cycles. We found that the HighSens and Zygem extraction methods were significantly better in their DNA yields than the one-tube method. Identifiler® Plus increased the quality of the STR profiles for the one-tube extraction significantly. However, this effect could not be verified for the other extraction methods. Furthermore, microscopic analysis of single fingerprints revealed that some individuals tended to shed more material than others onto glass slides. However, a dense deposition of skin flakes did not strongly correlate with a high quality STR profile. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparison of methods for nutrient measurement in calcareous soils: Ion-exchange resin bag, capsule, membrane, and chemical extractions

USGS Publications Warehouse

Sherrod, S.K.; Belnap, J.; Miller, M.E.

2002-01-01

Four methods for measuring quantities of 12 plant-available nutrients were compared using three sandy soils in a series of three experiments. Three of the methods use different ion-exchange resin forms—bags, capsules, and membranes—and the fourth was conventional chemical extraction. The first experiment compared nutrient extraction data from a medium of sand saturated with a nutrient solution. The second and third experiments used Nakai and Sheppard series soils from Canyonlands National Park, which are relatively high in soil carbonates. The second experiment compared nutrient extraction data provided by the four methods from soils equilibrated at two temperatures, “warm” and “cold.” The third experiment extracted nutrients from the same soils in a field equilibration. Our results show that the four extraction techniques are not comparable. This conclusion is due to differences among the methods in the net quantities of nutrients extracted from equivalent soil volumes, in the proportional representation of nutrients within similar soils and treatments, in the measurement of nutrients that were added in known quantities, and even in the order of nutrients ranked by net abundance. We attribute the disparities in nutrient measurement among the different resin forms to interacting effects of the inherent differences in resin exchange capacity, differences among nutrients in their resin affinities, and possibly the relatively short equilibration time for laboratory trials. One constraint for measuring carbonate-related nutrients in high-carbonate soils is the conventional ammonium acetate extraction method, which we suspect of dissolving fine CaCO3 particles that are more abundant in Nakai series soils, resulting in erroneously high Ca2+ estimates. For study of plant-available nutrients, it is important to identify the nutrients of foremost interest and understand differences in their resin sorption dynamics to determine the most appropriate extraction method.

Establishing a novel automated magnetic bead-based method for the extraction of DNA from a variety of forensic samples.

PubMed

Witt, Sebastian; Neumann, Jan; Zierdt, Holger; Gébel, Gabriella; Röscheisen, Christiane

2012-09-01

Automated systems have been increasingly utilized for DNA extraction by many forensic laboratories to handle growing numbers of forensic casework samples while minimizing the risk of human errors and assuring high reproducibility. The step towards automation however is not easy: The automated extraction method has to be very versatile to reliably prepare high yields of pure genomic DNA from a broad variety of sample types on different carrier materials. To prevent possible cross-contamination of samples or the loss of DNA, the components of the kit have to be designed in a way that allows for the automated handling of the samples with no manual intervention necessary. DNA extraction using paramagnetic particles coated with a DNA-binding surface is predestined for an automated approach. For this study, we tested different DNA extraction kits using DNA-binding paramagnetic particles with regard to DNA yield and handling by a Freedom EVO(®)150 extraction robot (Tecan) equipped with a Te-MagS magnetic separator. Among others, the extraction kits tested were the ChargeSwitch(®)Forensic DNA Purification Kit (Invitrogen), the PrepFiler™Automated Forensic DNA Extraction Kit (Applied Biosystems) and NucleoMag™96 Trace (Macherey-Nagel). After an extensive test phase, we established a novel magnetic bead extraction method based upon the NucleoMag™ extraction kit (Macherey-Nagel). The new method is readily automatable and produces high yields of DNA from different sample types (blood, saliva, sperm, contact stains) on various substrates (filter paper, swabs, cigarette butts) with no evidence of a loss of magnetic beads or sample cross-contamination. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

[Validation Study for Analytical Method of Diarrhetic Shellfish Poisons in 9 Kinds of Shellfish].

PubMed

Yamaguchi, Mizuka; Yamaguchi, Takahiro; Kakimoto, Kensaku; Nagayoshi, Haruna; Okihashi, Masahiro; Kajimura, Keiji

2016-01-01

A method was developed for the simultaneous determination of okadaic acid, dinophysistoxin-1 and dinophysistoxin-2 in shellfish using ultra performance liquid chromatography with tandem mass spectrometry. Shellfish poisons in spiked samples were extracted with methanol and 90% methanol, and were hydrolyzed with 2.5 mol/L sodium hydroxide solution. Purification was done on an HLB solid-phase extraction column. This method was validated in accordance with the notification of Ministry of Health, Labour and Welfare of Japan. As a result of the validation study in nine kinds of shellfish, the trueness, repeatability and within-laboratory reproducibility were 79-101%, less than 12 and 16%, respectively. The trueness and precision met the target values of notification.

Collection and processing of whole blood for transformation of peripheral blood mononuclear cells and extraction of DNA: the Type 1 Diabetes Genetics Consortium

PubMed Central

Rosinger, Silke; Nutland, Sarah; Mickelson, Eric; Varney, Michael D; Boehm, Bernard O; Olsem, Gary J; Hansen, John A; Nicholson, Ian; Hilner, Joan E; Perdue, Letitia H; Pierce, June J; Akolkar, Beena; Nierras, Concepcion; Steffes, Michael W

2010-01-01

Background and Purpose To yield large amounts of DNA for many genotype analyses and to provide a renewable source of DNA, the Type 1 Diabetes Genetics Consortium (T1DGC) harvested DNA and peripheral blood mononuclear cells (PBMCs) from individuals with type 1 diabetes and their family members in several regions of the world. Methods DNA repositories were established in Asia-Pacific, Europe, North America, and the United Kingdom. To address region-specific needs, different methods and sample processing techniques were used among the laboratories to extract and to quantify DNA and to establish Epstein-Barr virus transformed cell lines. Results More than 98% of the samples of PBMCs were successfully transformed. Approximately 20–25 µg of DNA were extracted per mL of whole blood. Extraction of DNA from the cell pack ranged from 92 to 165 µg per cell pack. In addition, the extracted DNA from whole blood or transformed cells was successfully utilized in each regional human leukocyte antigen genotyping laboratory and by several additional laboratories performing consortium-wide genotyping projects. Limitations Although the isolation of PBMCs was consistent among sites, the measurement of DNA was difficult to harmonize. Conclusions DNA repositories can be established in different regions of the world and produce similar amounts of high-quality DNA for a variety of high-throughput genotyping techniques. Furthermore, even with the distances and time necessary for transportation, highly efficient transformation of PBMCs is possible. For future studies/trials involving several laboratories in different locations, the T1DGC experience includes examples of protocols that may be applicable. In summary, T1DGC has developed protocols that would be of interest to any scientific organization attempting to overcome the logistical problems associated with studies/trials spanning multiple research facilities, located in different regions of the world. PMID:20595244

Extraction and quantitative analysis of iodine in solid and solution matrixes.

PubMed

Brown, Christopher F; Geiszler, Keith N; Vickerman, Tanya S

2005-11-01

129I is a contaminant of interest in the vadose zone and groundwater at numerous federal and privately owned facilities. Several techniques have been utilized to extract iodine from solid matrixes; however, all of them rely on two fundamental approaches: liquid extraction or chemical/heat-facilitated volatilization. While these methods are typically chosen for their ease of implementation, they do not totally dissolve the solid. We defined a method that produces complete solid dissolution and conducted laboratory tests to assess its efficacy to extract iodine from solid matrixes. Testing consisted of potassium nitrate/potassium hydroxide fusion of the sample, followed by sample dissolution in a mixture of sulfuric acid and sodium bisulfite. The fusion extraction method resulted in complete sample dissolution of all solid matrixes tested. Quantitative analysis of 127I and 129I via inductively coupled plasma mass spectrometry showed better than +/-10% accuracy for certified reference standards, with the linear operating range extending more than 3 orders of magnitude (0.005-5 microg/L). Extraction and analysis of four replicates of standard reference material containing 5 microg/g 127I resulted in an average recovery of 98% with a relative deviation of 6%. This simple and cost-effective technique can be applied to solid samples of varying matrixes with little or no adaptation.

Determination of the Effect of Various Modes of Cooking on the Vitamin C Content of a Common Food, Green Pepper: An Introductory Biochemistry Experiment.

ERIC Educational Resources Information Center

Johnson, Eric R.

1988-01-01

Describes a laboratory experiment that measures the amount of ascorbic acid destroyed by food preparation methods (boiling and steaming). Points out that aqueous extracts of cooked green pepper samples can be analyzed for ascorbic acid by a relatively simple redox titration. Lists experimental procedure for four methods of preparation. (MVL)

Improvement of the lipophilic-oxygen radical absorbance capacity (L-ORAC) method and single-laboratory validation.

PubMed

Watanabe, Jun; Oki, Tomoyuki; Takebayashi, Jun; Yamasaki, Koji; Takano-Ishikawa, Yuko; Hino, Akihiro; Yasui, Akemi

2013-01-01

We improved the procedure for lipophilic-oxygen radical absorbance capacity (L-ORAC) measurement for better repeatability and intermediate precision. A sealing film was placed on the assay plate, and glass vials and microdispensers equipped with glass capillaries were used. The antioxidant capacities of food extracts can be evaluated by this method with nearly the same precision as antioxidant solutions.

A simple, rapid atmospheric pressure chemical ionization liquid chromatography tandem mass spectrometry method for the determination of 25-hydroxyvitamin D2 and D3.

PubMed

Garg, Uttam; Munar, Ada; Frazee, Clinton; Scott, David

2012-09-01

Vitamin D plays a vital role not only in bone health but also in pathophysiology of many other body functions. In recent years, there has been significant increase in testing of 25-hydroxyvitamin D (25-OH vitamin D), a marker of vitamin D deficiency. The most commonly used methods for the measurement of 25-OH vitamin D are immunoassays and liquid chromatography tandem mass spectrometry (LC-MS-MS). Since immunoassays suffer from inaccuracies and interferences, LC-MS-MS is a preferred method. In LC-MS-MS methods, 25-OH vitamin D is extracted from serum or plasma by solid-phase or liquid-phase extraction. Because these extraction methods are time consuming, we developed an easy method that uses simple protein precipitation followed by injection of the supernatant to LC-MS-MS. Several mass-to-charge (m/z) ratio transitions, including commonly used transitions based on water loss, were evaluated and several tube types were tested. The optimal transitions for 25-OH vitamin D2 and D3 were 395.5 > 269.5 and 383.4 > 257.3, respectively. The reportable range of the method was 1-100 ng/mL, and repeatability (within-run) and within-laboratory imprecision were <4% and <6%, respectively. The method agreed well with the solid-phase extraction methods. © 2012 Wiley Periodicals, Inc.

A validated UHPLC-tandem mass spectrometry method for quantitative analysis of flavonolignans in milk thistle (Silybum marianum) extracts.

PubMed

Graf, Tyler N; Cech, Nadja B; Polyak, Stephen J; Oberlies, Nicholas H

2016-07-15

Validated methods are needed for the analysis of natural product secondary metabolites. These methods are particularly important to translate in vitro observations to in vivo studies. Herein, a method is reported for the analysis of the key secondary metabolites, a series of flavonolignans and a flavonoid, from an extract prepared from the seeds of milk thistle [Silybum marianum (L.) Gaertn. (Asteraceae)]. This report represents the first UHPLC MS-MS method validated for quantitative analysis of these compounds. The method takes advantage of the excellent resolution achievable with UHPLC to provide a complete analysis in less than 7min. The method is validated using both UV and MS detectors, making it applicable in laboratories with different types of analytical instrumentation available. Lower limits of quantitation achieved with this method range from 0.0400μM to 0.160μM with UV and from 0.0800μM to 0.160μM with MS. The new method is employed to evaluate variability in constituent composition in various commercial S. marianum extracts, and to show that storage of the milk thistle compounds in DMSO leads to degradation. Copyright © 2016 Elsevier B.V. All rights reserved.

A thesis on the Development of an Automated SWIFT Edge Detection Algorithm

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

Trujillo, Christopher J.

Throughout the world, scientists and engineers such as those at Los Alamos National Laboratory, perform research and testing unique only to applications aimed towards advancing technology, and understanding the nature of materials. With this testing, comes a need for advanced methods of data acquisition and most importantly, a means of analyzing and extracting the necessary information from such acquired data. In this thesis, I aim to produce an automated method implementing advanced image processing techniques and tools to analyze SWIFT image datasets for Detonator Technology at Los Alamos National Laboratory. Such an effective method for edge detection and point extractionmore » can prove to be advantageous in analyzing such unique datasets and provide for consistency in producing results.« less

Direct PCR Offers a Fast and Reliable Alternative to Conventional DNA Isolation Methods for Gut Microbiomes.

PubMed

Videvall, Elin; Strandh, Maria; Engelbrecht, Anel; Cloete, Schalk; Cornwallis, Charlie K

2017-01-01

The gut microbiome of animals is emerging as an important factor influencing ecological and evolutionary processes. A major bottleneck in obtaining microbiome data from large numbers of samples is the time-consuming laboratory procedures required, specifically the isolation of DNA and generation of amplicon libraries. Recently, direct PCR kits have been developed that circumvent conventional DNA extraction steps, thereby streamlining the laboratory process by reducing preparation time and costs. However, the reliability and efficacy of direct PCR for measuring host microbiomes have not yet been investigated other than in humans with 454 sequencing. Here, we conduct a comprehensive evaluation of the microbial communities obtained with direct PCR and the widely used Mo Bio PowerSoil DNA extraction kit in five distinct gut sample types (ileum, cecum, colon, feces, and cloaca) from 20 juvenile ostriches, using 16S rRNA Illumina MiSeq sequencing. We found that direct PCR was highly comparable over a range of measures to the DNA extraction method in cecal, colon, and fecal samples. However, the two methods significantly differed in samples with comparably low bacterial biomass: cloacal and especially ileal samples. We also sequenced 100 replicate sample pairs to evaluate repeatability during both extraction and PCR stages and found that both methods were highly consistent for cecal, colon, and fecal samples ( r s > 0.7) but had low repeatability for cloacal ( r s = 0.39) and ileal ( r s = -0.24) samples. This study indicates that direct PCR provides a fast, cheap, and reliable alternative to conventional DNA extraction methods for retrieving 16S rRNA data, which can aid future gut microbiome studies. IMPORTANCE The microbial communities of animals can have large impacts on their hosts, and the number of studies using high-throughput sequencing to measure gut microbiomes is rapidly increasing. However, the library preparation procedure in microbiome research is both costly and time-consuming, especially for large numbers of samples. We investigated a cheaper and faster direct PCR method designed to bypass the DNA isolation steps during 16S rRNA library preparation and compared it with a standard DNA extraction method. We used both techniques on five different gut sample types collected from 20 juvenile ostriches and sequenced samples with Illumina MiSeq. The methods were highly comparable and highly repeatable in three sample types with high microbial biomass (cecum, colon, and feces), but larger differences and low repeatability were found in the microbiomes obtained from the ileum and cloaca. These results will help microbiome researchers assess library preparation procedures and plan their studies accordingly.

Phytochemical Comparison of the Water and Ethanol Leaf Extracts of the Cree medicinal plant, Sarracenia purpurea L. (Sarraceniaceae).

PubMed

Cieniak, Carolina; Walshe-Roussel, Brendan; Liu, Rui; Muhammad, Asim; Saleem, Ammar; Haddad, Pierre S; Cuerrier, Alain; Foster, Brian C; Arnason, John T

2015-01-01

The Cree of Eeyou Istchee in Northern Quebec identified Sarracenia purpurea L. as an important plant for the treatment of Type 2 diabetes. Traditionally the plant is used as a decoction (boiling water extract) of the leaf, however, in order to study the extract in a laboratory setting, an 80% ethanol extract was used. In this study, the phytochemistry of both extracts of the leaves was compared and quantified. Two S. purpurea leaf extracts were prepared, one a traditional hot water extract and the other an 80% ethanol extract. Using UPLC-ESI-MS, the extracts were phytochemically compared for 2 triterpenes, betulinic acid and ursolic acid, using one gradient method and for 10 additional substances, including the actives quercetin-3-O-galactoside and morroniside, using a different method. The concentrations of the nine phenolic substances present, as well as an active principle, the iridoid glycoside morroniside, were very similar between the two extracts, with generally slightly higher concentrations of phenolics in the ethanol extract as expected. However, two triterpenes, betulinic acid and ursolic acid, were 107 and 93 times more concentrated, respectively, in the ethanol extract compared to the water extract. The main phytochemical markers and most importantly the antidiabetic active principles, quercetin-3-O-galactoside and morroniside, were present in similar amounts in the two extracts, which predicts similar bioactivity.This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Ovicidal and repellent activities of botanical extracts against Culex quinquefasciatus, Aedes aegypti and Anopheles stephensi (Diptera: Culicidae)

PubMed Central

Govindarajan, M; Mathivanan, T; Elumalai, K; Krishnappa, K; Anandan, A

2011-01-01

Objective To determine the ovicidal and repellent activities of methanol leaf extract of Ervatamia coronaria (E. coronaria) and Caesalpinia pulcherrima (C. pulcherrima) against Culex quinquefasciatus (Cx. quinquefasciatus), Aedes aegypti (Ae. aegypti) and Anopheles stephensi (An. stephensi). Methods The ovicidal activity was determined against three mosquito species at various concentrations ranging from 50-450 ppm under the laboratory conditions. The hatch rates were assessed 48 h after treatment. The repellent efficacy was determined against three mosquito species at three concentrations viz., 1.0, 2.5 and 5.0 mg/cm2 under the laboratory conditions. Results The crude extract of E. coronaria exerted zero hatchability (100% mortality) at 250, 200 and 150 ppm for Cx. quinquefasciatus, Ae. aegypti and An. stephensi, respectively. The crude extract of C. pulcherrima exerted zero hatchability (100% mortality) at 375, 300 and 225 ppm for Cx. quinquefasciatus, Ae. aegypti and An. Stephensi, respectively. The methanol extract of E. coronaria found to be more repellenct than C. pulcherrima extract. A higher concentration of 5.0 mg/cm2 provided 100% protection up to 150, 180 and 210 min against Cx. quinquefasciatus, Ae. aegypti and An. stephensi, respectively. The results clearly showed that repellent activity was dose dependent. Conclusions From the results it can be concluded the crude extracts of E. coronaria and C. pulcherrima are an excellent potential for controlling Cx. quinquefasciatus, Ae. aegypti and An. stephensi mosquitoes. PMID:23569723

Sulfur geochemistry of hydrothermal waters in Yellowstone National Park, Wyoming, USA. III. An anion-exchange resin technique for sampling and preservation of sulfoxyanions in natural waters

USGS Publications Warehouse

Druschel, G.K.; Schoonen, M.A.A.; Nordstorm, D.K.; Ball, J.W.; Xu, Y.; Cohn, C.A.

2003-01-01

A sampling protocol for the retention, extraction, and analysis of sulfoxyanions in hydrothermal waters has been developed in the laboratory and tested at Yellowstone National Park and Green Lake, NY. Initial laboratory testing of the anion-exchange resin Bio-Rad??? AG1-X8 indicated that the resin was well suited for the sampling, preservation, and extraction of sulfate and thiosulfate. Synthetic solutions containing sulfate and thiosulfate were passed through AG1-X8 resin columns and eluted with 1 and 3 M KCl, respectively. Recovery ranged from 89 to 100%. Comparison of results for water samples collected from five pools in Yellowstone National Park between on-site IC analysis (U.S. Geological Survey mobile lab) and IC analysis of resin-stored sample at SUNY-Stony Brook indicates 96 to 100% agreement for three pools (Cinder, Cistern, and an unnamed pool near Cistern) and 76 and 63% agreement for two pools (Sulfur Dust and Frying Pan). Attempts to extract polythionates from the AG1-X8 resin were made using HCl solutions, but were unsuccessful. Bio-Rad??? AG2-X8, an anion-exchange resin with weaker binding sites than the AG1-X8 resin, is better suited for polythionate extraction. Sulfate and thiosulfate extraction with this resin has been accomplished with KCl solutions of 0.1 and 0.5 M, respectively. Trithionate and tetrathionate can be extracted with 4 M KCl. Higher polythionates can be extracted with 9 M hydrochloric acid. Polythionate concentrations can then be determined directly using ion chromatographic methods, and laboratory results indicate recovery of up to 90% for synthetic polythionate solutions using AG2-X8 resin columns. ?? The Royal Society of Chemistry and the Division of Geochemistry of the American Chemical Society 2003.

A Guided Inquiry Liquid/Liquid Extractions Laboratory for Introductory Organic Chemistry

ERIC Educational Resources Information Center

Raydo, Margaret L.; Church, Megan S.; Taylor, Zane W.; Taylor, Christopher E.; Danowitz, Amy M.

2015-01-01

A guided inquiry laboratory experiment for teaching liquid/liquid extractions to first semester undergraduate organic chemistry students is described. This laboratory is particularly useful for introductory students as the analytes that are separated are highly colored dye molecules. This allows students to track into which phase each analyte…

Comprehensive Urine Drug Screen by Gas Chromatography/Mass Spectrometry (GC/MS).

PubMed

Ramoo, Bheemraj; Funke, Melissa; Frazee, Clint; Garg, Uttam

2016-01-01

Drug screening is an essential component of clinical toxicology laboratory service. Some laboratories use only automated chemistry analyzers for limited screening of drugs of abuse and few other drugs. Other laboratories use a combination of various techniques such as immunoassays, colorimetric tests, and mass spectrometry to provide more detailed comprehensive drug screening. Mass spectrometry, gas or liquid, can screen for hundreds of drugs and is often considered the gold standard for comprehensive drug screening. We describe an efficient and rapid gas chromatography/mass spectrometry (GC/MS) method for comprehensive drug screening in urine which utilizes a liquid-liquid extraction, sample concentration, and analysis by GC/MS.

Determination of flavanol and procyanidin (by degree of polymerization 1-10) content of chocolate, cocoa liquors, powder(s), and cocoa flavanol extracts by normal phase high-performance liquid chromatography: collaborative study.

PubMed

Robbins, Rebecca J; Leonczak, Jadwiga; Li, Julia; Johnson, J Christopher; Collins, Tom; Kwik-Uribe, Catherine; Schmitz, Harold H

2012-01-01

An international collaborative study was conducted on an HPLC method with fluorescent detection (FLD) for the determination of flavanols and procyanidins in materials containing chocolate and cocoa. The sum of the oligomeric fractions with degree of polymerization 1-10 was the determined content value. Sample materials included dark and milk chocolates, cocoa powder, cocoa liquors, and cocoa extracts. The content ranged from approximately 2 to 500 mg/g (defatted basis). Thirteen laboratories representing commercial, industrial, and academic institutions in six countries participated in the study. Fourteen samples were sent as blind duplicates to the collaborators. Results from 12 laboratories yielded repeatability relative standard deviation (RSDr) values that were below 10% for all materials analyzed, ranging from 4.17 to 9.61%. The reproducibility relative standard deviation (RSD(R)) values ranged from 5.03 to 12.9% for samples containing 8.07 to 484.7 mg/g. In one sample containing a low content of flavanols and procyanidins (approximately 2 mg/g), the RSD(R) was 17.68%. Based on these results, the method is recommended for Official First Action for the determination of flavanols and procyanidins in chocolate, cocoa liquors, powder(s), and cocoa extracts.

Arsenic in coal of the Thar coalfield, Pakistan, and its behavior during combustion.

PubMed

Ali, Jamshed; Kazi, Tasneem G; Baig, Jameel A; Afridi, Hassan I; Arain, Mariam S; Brahman, Kapil D; Naeemullah; Panhwar, Abdul H

2015-06-01

The aim of the current study is to evaluate the occurrence of arsenic in coal collected from Thar coalfield, Pakistan, and its behavior during the combustion. Fractionation of arsenic (As) in coal samples was carried out by Community Bureau of Reference sequential extraction scheme (BCR-SES) and single-step-based BCR method (BCR-SS). These methods are validated using the certified reference material of sediment BCR 701 and standard addition method. The stepwise fractions of As in laboratory-made ash (LMA) have been also investigated. The extractable As content associated with different phases in coal and LMA samples were analyzed by electrothermal atomic absorption spectrophotometer. The extraction efficiency of As by BCR-SS was slightly higher than BCR-SES, while the difference was not significant (p < 0.05). The BCR-SS method is a time-saving method because it can reduce the extraction time from 51 to 22 h. The As contents in LMA revealed that during combustion of the coal, >85 % of As may be released into atmosphere. The relative mobility of As in the coal samples was found in increasing order as follows: oxidizable fraction < reducible fraction < acid soluble fraction. The total and extractable As obtained by BCR-SES and BCR-SS were higher in coal samples of block III as compared to block V (p > 0.05).

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory : determination of polycyclic aromatic hydrocarbon compounds in sediment by gas chromatography/mass spectrometry

USGS Publications Warehouse

Olson, Mary C.; Iverson, Jana L.; Furlong, Edward T.; Schroeder, Michael P.

2004-01-01

A method for the determination of 28 polycyclic aromatic hydrocarbons (PAHs) and 25 alkylated PAH homolog groups in sediment samples is described. The compounds are extracted from sediment by solvent extraction, followed by partial isolation using high-performance gel permeation chromatography. The compounds are identified and uantitated using capillary-column gas chromatography/mass spectrometry. The report presents performance data for full-scan ion monitoring. Method detection limits in laboratory reagent matrix samples range from 1.3 to 5.1 micrograms per kilogram for the 28 PAHs. The 25 groups of alkylated PAHs are homologs of five groups of isomeric parent PAHs. Because of the lack of authentic standards, these homologs are reported semiquantitatively using a response factor from a parent PAH or a specific alkylated PAH. Precision data for the alkylated PAH homologs are presented using two different standard reference manuals produced by the National Institute of Standards and Technology: SRM 1941b and SRM 1944. The percent relative standard deviations for identified alkylated PAH homolog groups ranged from 1.55 to 6.98 for SRM 1941b and from 6.11 to 12.0 for SRM 1944. Homolog group concentrations reported under this method include the concentrations of individually identified compounds that are members of the group. Organochlorine (OC) pesticides--including toxaphene, polychlorinated biphenyls (PCBs), and organophosphate (OP) pesticides--can be isolated simultaneously using this method. In brief, sediment samples are centrifuged to remove excess water and extracted overnight with dichloromethan (95 percent) and methanol (5 percent). The extract is concentrated and then filtered through a 0.2-micrometer polytetrafluoroethylene syringe filter. The PAH fraction is isolated by quantitatively injecting an aliquot of sample onto two polystyrene-divinylbenzene gel-permeation chromatographic columns connected in series. The compounds are eluted with dichloromethane, a PAH fraction is collected, and a portion of the coextracted interferences, including elemental sulfur, is separated and discarded. The extract is solvent exchanged, the volume is reduced, and internal standard is added. Sample analysis is completed using a gas chromatograph/mass spectrometer and full-scan acquisition.

Multicenter Comparative Evaluation of Five Commercial Methods for Toxoplasma DNA Extraction from Amniotic Fluid▿

PubMed Central

Yera, H.; Filisetti, D.; Bastien, P.; Ancelle, T.; Thulliez, P.; Delhaes, L.

2009-01-01

Over the past few years, a number of new nucleic acid extraction methods and extraction platforms using chemistry combined with magnetic or silica particles have been developed, in combination with instruments to facilitate the extraction procedure. The objective of the present study was to investigate the suitability of these automated methods for the isolation of Toxoplasma gondii DNA from amniotic fluid (AF). Therefore, three automated procedures were compared to two commercialized manual extraction methods. The MagNA Pure Compact (Roche), BioRobot EZ1 (Qiagen), and easyMAG (bioMérieux) automated procedures were compared to two manual DNA extraction kits, the QIAamp DNA minikit (Qiagen) and the High Pure PCR template preparation kit (Roche). Evaluation was carried out with two specific Toxoplasma PCRs (targeting the 529-bp repeat element), inhibitor search PCRs, and human beta-globin PCRs. The samples each consisted of 4 ml of AF with or without a calibrated Toxoplasma gondii RH strain suspension (0, 1, 2.5, 5, and 25 tachyzoites/ml). All PCR assays were laboratory-developed real-time PCR assays, using either TaqMan or fluorescent resonance energy transfer probes. A total of 1,178 PCRs were performed, including 978 Toxoplasma PCRs. The automated and manual methods were similar in sensitivity for DNA extraction from T. gondii at the highest concentration (25 Toxoplasma gondii cells/ml). However, our results showed that the DNA extraction procedures led to variable efficacy in isolating low concentrations of tachyzoites in AF samples (<5 Toxoplasma gondii cells/ml), a difference that might have repercussions since low parasite concentrations in AF exist and can lead to congenital toxoplasmosis. PMID:19846633

One simple DNA extraction device and its combination with modified visual loop-mediated isothermal amplification for rapid on-field detection of genetically modified organisms.

PubMed

Zhang, Miao; Liu, Yinan; Chen, Lili; Quan, Sheng; Jiang, Shimeng; Zhang, Dabing; Yang, Litao

2013-01-02

Quickness, simplicity, and effectiveness are the three major criteria for establishing a good molecular diagnosis method in many fields. Herein we report a novel detection system for genetically modified organisms (GMOs), which can be utilized to perform both on-field quick screening and routine laboratory diagnosis. In this system, a newly designed inexpensive DNA extraction device was used in combination with a modified visual loop-mediated isothermal amplification (vLAMP) assay. The main parts of the DNA extraction device included a silica gel membrane filtration column and a modified syringe. The DNA extraction device could be easily operated without using other laboratory instruments, making it applicable to an on-field GMO test. High-quality genomic DNA (gDNA) suitable for polymerase chain reaction (PCR) and isothermal amplification could be quickly isolated from plant tissues using this device within 15 min. In the modified vLAMP assay, a microcrystalline wax encapsulated detection bead containing SYBR green fluorescent dye was introduced to avoid dye inhibition and cross-contaminations from post-LAMP operation. The system was successfully applied and validated in screening and identification of GM rice, soybean, and maize samples collected from both field testing and the Grain Inspection, Packers, and Stockyards Administration (GIPSA) proficiency test program, which demonstrated that it was well-adapted to both on-field testing and/or routine laboratory analysis of GMOs.

Mosquito larvicidal properties of Orthisiphon thymiflorus (Roth) Sleesen. (Family: Labiatae) against mosquito vectors, Anopheles stephensi, Culex quinquefasciatus and Aedes aegypti (Diptera: Culicidae)

USDA-ARS?s Scientific Manuscript database

Objective: To determine the larvicidal activity of hexane, chloroform, ethyl acetate, acetone, and methanol extracts of Orthosiphon thymiflorus leaves against Anopheles stephensi, Culex quinquefasciatus and Aedes aegypti. Methods: Larvicidal activity was determined in laboratory bioassays using var...

AUTOMATED PRESSURIZED FLUID EXTRACTION WITH IN-LINE CLEAN-UP FOR DETERMINATION OF PESTICIDES IN COMPOSITE DIETS.

EPA Science Inventory

USEPA's National Exposure Research Laboratory conducts research to measure the exposure of individuals to chemical pollutants through the diet, as well as other media. In support of this research, methods are being evaluated for determination of organophosphate pesticides in co...

DETERMINATION OF PESTICIDES IN COMOSITE DIETS USING LARGE VOLUME PRESSURIZED FLUID EXTRACTION WITH IN-LINE SAMPLE PREPARATION

EPA Science Inventory

USEPA's National Exposure Research Laboratory conducts research to measure the exposure of individuals to chemical pollutants through the diet, as well as other media. In support of this research, methods are being evaluated for determination of various classes of pesticides in ...

Generic sample preparation combined with high-resolution liquid chromatography-time-of-flight mass spectrometry for unification of urine screening in doping-control laboratories.

PubMed

Peters, R J B; Oosterink, J E; Stolker, A A M; Georgakopoulos, C; Nielen, M W F

2010-04-01

A unification of doping-control screening procedures of prohibited small molecule substances--including stimulants, narcotics, steroids, beta2-agonists and diuretics--is highly urgent in order to free resources for new classes such as banned proteins. Conceptually this may be achieved by the use of a combination of one gas chromatography-time-of-flight mass spectrometry method and one liquid chromatography-time-of-flight mass spectrometry method. In this work a quantitative screening method using high-resolution liquid chromatography in combination with accurate-mass time-of-flight mass spectrometry was developed and validated for determination of glucocorticosteroids, beta2-agonists, thiazide diuretics, and narcotics and stimulants in urine. To enable the simultaneous isolation of all the compounds of interest and the necessary purification of the resulting extracts, a generic extraction and hydrolysis procedure was combined with a solid-phase extraction modified for these groups of compounds. All 56 compounds are determined using positive electrospray ionisation with the exception of the thiazide diuretics for which the best sensitivity was obtained by using negative electrospray ionisation. The results show that, with the exception of clenhexyl, procaterol, and reproterol, all compounds can be detected below the respective minimum required performance level and the results for linearity, repeatability, within-lab reproducibility, and accuracy show that the method can be used for quantitative screening. If qualitative screening is sufficient the instrumental analysis may be limited to positive ionisation, because all analytes including the thiazides can be detected at the respective minimum required levels in the positive mode. The results show that the application of accurate-mass time-of-flight mass spectrometry in combination with generic extraction and purification procedures is suitable for unification and expansion of the window of screening methods of doping laboratories. Moreover, the full-scan accurate-mass data sets obtained still allow retrospective examination for emerging doping agents, without re-analyzing the samples.

The room temperature preservation of filtered environmental DNA samples and assimilation into a phenol–chloroform–isoamyl alcohol DNA extraction

PubMed Central

Renshaw, Mark A; Olds, Brett P; Jerde, Christopher L; McVeigh, Margaret M; Lodge, David M

2015-01-01

Current research targeting filtered macrobial environmental DNA (eDNA) often relies upon cold ambient temperatures at various stages, including the transport of water samples from the field to the laboratory and the storage of water and/or filtered samples in the laboratory. This poses practical limitations for field collections in locations where refrigeration and frozen storage is difficult or where samples must be transported long distances for further processing and screening. This study demonstrates the successful preservation of eDNA at room temperature (20 °C) in two lysis buffers, CTAB and Longmire's, over a 2-week period of time. Moreover, the preserved eDNA samples were seamlessly integrated into a phenol–chloroform–isoamyl alcohol (PCI) DNA extraction protocol. The successful application of the eDNA extraction to multiple filter membrane types suggests the methods evaluated here may be broadly applied in future eDNA research. Our results also suggest that for many kinds of studies recently reported on macrobial eDNA, detection probabilities could have been increased, and at a lower cost, by utilizing the Longmire's preservation buffer with a PCI DNA extraction. PMID:24834966

Analysis of laboratory compaction methods of roller compacted concrete

NASA Astrophysics Data System (ADS)

Trtík, Tomáš; Chylík, Roman; Bílý, Petr; Fládr, Josef

2017-09-01

Roller-Compacted Concrete (RCC) is an ordinary concrete poured and compacted with machines typically used for laying of asphalt road layers. One of the problems connected with this technology is preparation of representative samples in the laboratory. The aim of this work was to analyse two methods of preparation of RCC laboratory samples with bulk density as the comparative parameter. The first method used dynamic compaction by pneumatic hammer. The second method of compaction had a static character. The specimens were loaded by precisely defined force in laboratory loading machine to create the same conditions as during static rolling (in the Czech Republic, only static rolling is commonly used). Bulk densities obtained by the two compaction methods were compared with core drills extracted from real RCC structure. The results have shown that the samples produced by pneumatic hammer tend to overestimate the bulk density of the material. For both compaction methods, immediate bearing index test was performed to verify the quality of compaction. A fundamental difference between static and dynamic compaction was identified. In static compaction, initial resistance to penetration of the mandrel was higher, after exceeding certain limit the resistance was constant. This means that the samples were well compacted just on the surface. Specimens made by pneumatic hammer actively resisted throughout the test, the whole volume was uniformly compacted.

Methods of analysis by the U. S. Geological Survey National Water Quality Laboratory - determination of organonitrogen herbicides in water by solid-phase extraction and capillary-column gas chromatography/mass spectrometry with selected-ion monitoring

USGS Publications Warehouse

Sandstrom, Mark W.; Wydoski, Duane S.; Schroeder, Michael P.; Zamboni, Jana L.; Foreman, William T.

1992-01-01

A method for the isolation of organonitrogen herbicides from natural water samples using solid-phase extraction and analysis by capillary-column gas chromatography/mass spectrometry with selected-ion monitoring is described. Water samples are filtered to remove suspended particulate matter and then are pumped through disposable solid-phase extraction cartridges containing octadecyl-bonded porous silica to remove the herbicides. The cartridges are dried using carbon dioxide, and adsorbed herbicides are removed from the cartridges by elution with 1.8 milliliters of hexaneisopropanol (3:1). Extracts of the eluants are analyzed by capillary-column gas chromatography/mass spectrometry with selected-ion monitoring of at least three characteristic ions. The method detection limits are dependent on sample matrix and each particular herbicide. The method detection limits, based on a 100-milliliter sample size, range from 0.02 to 0.25 microgram per liter. Recoveries averaged 80 to 115 percent for the 23 herbicides and 2 metabolites in 1 reagent-water and 2 natural-water samples fortified at levels of 0.2 and 2.0 micrograms per liter.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of pesticides in water by graphitized carbon-based solid-phase extraction and high-performance liquid chromatography/mass spectrometry

USGS Publications Warehouse

Furlong, Edward T.; Anderson, Bruce D.; Werner, Stephen L.; Soliven, Paul P.; Coffey, Laura J.; Burkhardt, Mark R.

2001-01-01

In 1996, the U.S. Geological Survey National Water Quality Laboratory (NWQL) developed and implemented a graphitized carbon-based solid-phase extraction and high-performance liquid chromatographic analytical method to determine polar pesticide concentrations in surface- and ground-water samples. Subsequently, the NWQL developed a complementary analysis that uses high-performance liquid chromatography/mass spectrometry to detect, identify, and quantify polar pesticides and pesticide metabolites in filtered water at concentrations as low as 10 nanograms per liter. This new method was designed to improve sensitivity and selectivity over the prior method, and to reduce known interferences from natural organic matter.In this new method, pesticides are extracted from filtered water samples by useing a 0.5-gram graphitized carbon-based solid-phase extraction cartridge, eluted from the cartridge, and concentrations determined by using high-performance liquid chromatography with electrospray ionization-mass spectrometry. The upper concentration limit is 1.000 microgram per liter (μg/L) for most compounds. Single-operator method detection limits in organic-free water samples fortified with pesticides at a concentration of 0.025 μg/L ranged from 0.0019 to 0.022 μg/L for all compounds in the method. The grand mean (mean of mean recoveries for individual compounds) recoveries in organic-free water samples ranged from 72 to 89 percent, fortified with pesticides at three concentrations between 0.025 and 0.5 μg/L. Grand mean recoveries in ground- and surface-water samples ranged from 46 to 119 percent, also fortified with pesticides at three concentrations between 0.025 and 0.5 μg/L. Long-term recoveries from reagent water spikes were used to demonstrate that 38 of 65 compounds can be reported without qualification of the quantitative result across the analytical range of the method. The remaining 27 are reported with qualified estimates of concentration because of greater variability of recovery.

The advent of a new pseudoephedrine product to combat methamphetamine abuse

PubMed Central

Leech, Ronald; Stark, Jeffrey G.

2013-01-01

Background: The personal and societal effects of methamphetamine abuse are well documented. The ease of accessibility to methamphetamine and the quality of the “high” it produces makes the drug highly desired by its abusers. Over time, many methamphetamine users will also become methamphetamine cooks, where pseudoephedrine in over-the-counter cold products is converted to methamphetamine through a simple, albeit extremely dangerous, process. New laws limiting access to these products have had limited success. No existing commercial pseudoephedrine products offer significant impediments to slow or limit the extraction and conversion of pseudoephedrine in clandestine methamphetamine laboratories. Objective and Methods: A new pseudoephedrine 30 mg tablet product using Impede technology (Nexafed®) to deter methamphetamine production has recently been introduced into the marketplace. Using methods designed to mimic clandestine laboratory processes, the ability of this product to disrupt extraction and conversion of pseudoephedrine to methamphetamine yet provide therapeutic effectiveness was evaluated. Results: Impede™ technology tablets limited the extraction and/or conversion of pseudoephedrine to methamphetamine when compared to a commercially marketed pseudoephedrine product (Sudafed®). Nexafed® tablets were also shown to be bioequivalent to the same control product, thus ensuring therapeutic equivalence. Conclusions: With the advent of new pseudoephedrine products in the marketplace with features to limit the extraction and conversion of pseudoephedrine to methamphetamine, new tools are now available to minimize the clandestine manufacture of the drug and potentially limit its social impact. PMID:23968171

Distinguishing Vaccinium species by chemical fingerprinting based on NMR spectra, validated with spectra collected in different laboratories.

PubMed

Markus, Michelle A; Ferrier, Jonathan; Luchsinger, Sarah M; Yuk, Jimmy; Cuerrier, Alain; Balick, Michael J; Hicks, Joshua M; Killday, K Brian; Kirby, Christopher W; Berrue, Fabrice; Kerr, Russell G; Knagge, Kevin; Gödecke, Tanja; Ramirez, Benjamin E; Lankin, David C; Pauli, Guido F; Burton, Ian; Karakach, Tobias K; Arnason, John T; Colson, Kimberly L

2014-06-01

A method was developed to distinguish Vaccinium species based on leaf extracts using nuclear magnetic resonance spectroscopy. Reference spectra were measured on leaf extracts from several species, including lowbush blueberry (Vaccinium angustifolium), oval leaf huckleberry (Vaccinium ovalifolium), and cranberry (Vaccinium macrocarpon). Using principal component analysis, these leaf extracts were resolved in the scores plot. Analysis of variance statistical tests demonstrated that the three groups differ significantly on PC2, establishing that the three species can be distinguished by nuclear magnetic resonance. Soft independent modeling of class analogies models for each species also showed discrimination between species. To demonstrate the robustness of nuclear magnetic resonance spectroscopy for botanical identification, spectra of a sample of lowbush blueberry leaf extract were measured at five different sites, with different field strengths (600 versus 700 MHz), different probe types (cryogenic versus room temperature probes), different sample diameters (1.7 mm versus 5 mm), and different consoles (Avance I versus Avance III). Each laboratory independently demonstrated the linearity of their NMR measurements by acquiring a standard curve for chlorogenic acid (R(2) = 0.9782 to 0.9998). Spectra acquired on different spectrometers at different sites classifed into the expected group for the Vaccinium spp., confirming the utility of the method to distinguish Vaccinium species and demonstrating nuclear magnetic resonance fingerprinting for material validation of a natural health product. Georg Thieme Verlag KG Stuttgart · New York.

Preliminary Tests For Development Of A Non-Pertechnetate Analysis Method

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

Diprete, D.; McCabe, D.

2016-09-28

The objective of this task was to develop a non-pertechnetate analysis method that 222-S lab could easily implement. The initial scope involved working with 222-S laboratory personnel to adapt the existing Tc analytical method to fractionate the non-pertechnetate and pertechnetate. SRNL then developed and tested a method using commercial sorbents containing Aliquat ® 336 to extract the pertechnetate (thereby separating it from non-pertechnetate), followed by oxidation, extraction, and stripping steps, and finally analysis by beta counting and Mass Spectroscopy. Several additional items were partially investigated, including impacts of a 137Cs removal step. The method was initially tested on SRS tankmore » waste samples to determine its viability. Although SRS tank waste does not contain non-pertechnetate, testing with it was useful to investigate the compatibility, separation efficiency, interference removal efficacy, and method sensitivity.« less

Pungency Quantitation of Hot Pepper Sauces Using HPLC

NASA Astrophysics Data System (ADS)

Betts, Thomas A.

1999-02-01

A class of compounds known as capsaicinoids are responsible for the "heat" of hot peppers. To determine the pungency of a particular pepper or pepper product, one may quantify the capsaicinoids and relate those concentrations to the perceived heat. The format of the laboratory described here allows students to collectively develop an HPLC method for the quantitation of the two predominant capsaicinoids (capsaicin and dihydrocapsaicin) in hot-pepper products. Each small group of students investigated one of the following aspects of the method: detector wavelength, mobile-phase composition, extraction of capsaicinoids, calibration, and quantitation. The format of the lab forced students to communicate and cooperate to develop this method. The resulting HPLC method involves extraction with acetonitrile followed by solid-phase extraction clean-up, an isocratic 80:20 methanol-water mobile phase, a 4.6 mm by 25 cm C-18 column, and UV absorbance detection at 284 nm. The method developed by the students was then applied to the quantitation of capsaicinoids in a variety of hot pepper sauces. Editor's Note on Hazards in our April 2000 issue addresses the above.

The difference between laboratory and in-situ pixel-averaged emissivity: The effects on temperature-emissivity separation

NASA Technical Reports Server (NTRS)

Matsunaga, Tsuneo

1993-01-01

Advanced Spaceborne Thermal Emission and Reflection Radiometer (ASTER) is a Japanese future imaging sensor which has five channels in thermal infrared (TIR) region. To extract spectral emissivity information from ASTER and/or TIMS data, various temperature-emissivity (T-E) separation methods have been developed to date. Most of them require assumptions on surface emissivity, in which emissivity measured in a laboratory is often used instead of in-situ pixel-averaged emissivity. But if these two emissivities are different, accuracies of separated emissivity and surface temperature are reduced. In this study, the difference between laboratory and in-situ pixel-averaged emissivity and its effect on T-E separation are discussed. TIMS data of an area containing both rocks and vegetation were also processed to retrieve emissivity spectra using two T-E separation methods.

Carbon coated magnetic nanoparticles as a novel magnetic solid phase extraction adsorbent for simultaneous extraction of methamphetamine and ephedrine from urine samples.

PubMed

Taghvimi, Arezou; Hamishehkar, Hamed

2017-01-15

This paper develops a highly selective, specific and efficient method for simultaneous determination of ephedrine and methamphetamine by a new carbon coated magnetic nanoparticles (C/MNPs) as a magnetic solid phase extraction (MSPE) adsorbent in biological urine medium. The characterization of synthesized magnetic nano adsorbent was completely carried out by various characterization techniques like Fourier transform infrared (FT-IR) spectroscopy, powder x-ray diffraction (XRD), scanning electron microscopy (SEM) and vibrating sample magnetometer (VSM). Nine important parameters influencing extraction efficiency including amount of adsorbent, amounts of sample volume, pH, type and amount of extraction organic solvent, time of extraction and desorption, agitation rate and ionic strength of extraction medium, were studied and optimized. Under optimized extraction conditions, a good linearity was observed in the concentration range of 100-2000ng/mL for ephedrine and 100-2500ng/mL for methamphetamine. Analysis of positive urine samples was carried out by proposed method with the recovery of 98.71 and 97.87% for ephedrine and methamphetamine, respectively. The results indicated that carbon coated magnetic nanoparticles could be applied in clinical and forensic laboratories for simultaneous determination of abused drugs in urine media. Copyright © 2016 Elsevier B.V. All rights reserved.

Antibacterial Activity of Ethanolic Extract of Cinnamon Bark, Honey, and Their Combination Effects against Acne-Causing Bacteria

PubMed Central

Julianti, Elin; Rajah, Kasturi K.; Fidrianny, Irda

2017-01-01

Propionibacterium acnes and Staphylococcus epidermidis are the major skin bacteria that cause the formation of acne. The present study was conducted to investigate antibacterial activity of ethanolic extract of cinnamon bark, honey, and their combination against acne bacteria. The antibacterial activity of extract of cinnamon bark and honey were investigated against P. acnes and S. epidermidis using disc diffusion. Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) were attained using Clinical and Laboratory Standard Institute (CLSI) methods. The interaction between cinnamon bark extract and honey was determined using a checkerboards method. The results showed that the MICs of cinnamon bark extract and honey against P. acne were 256 µg/mL and 50% v/v, respectively, while those against S. epidermidis were 1024 µg/mL and 50% v/v, respectively. The MBC of cinnamon bark extract against P. acnes and S. epidermidis were more than 2048 µg/mL, whereas the MBC for honey against P. acnes and S. epidermidis were 100%. The combination of cinnamon bark extract and honey against P. acnes and S. epidermidis showed additive activity with a fractional inhibitory concentration index (FICI) value of 0.625. Therefore, the combination of cinnamon bark extract and honey has potential activity against acne-causing bacteria. PMID:28398231

Color and polyphenolic stability in extracts produced from muscadine grape (Vitis rotundifolia) pomace.

PubMed

Cardona, Jorge A; Lee, Joon-Hee; Talcott, Stephen T

2009-09-23

The muscadine grape ( Vitis rotundifolia ) industry of the southern United States is largely devoid of value-added processes that capture the phytochemical content of wine and juice byproducts. Methods to recover and stabilize polyphenolics from muscadine grape pomace following juice manufacture were evaluated in laboratory-scale and pilot-scale trials. In laboratory-scale trials using osmotic equilibration, water-based extracts from juice pomace initially extracted 31-42% of total polyphenolics, 26-32% of total ellagic acid, and 36-62% of total anthocyanins. When adsorbed onto Amberlite XAD-4 resin to concentrate polyphenolics, these extracts lost 10.5% of their total ellagic acid from inefficient adsorption to the solid phase support. Subsequent pilot-scale trials were evaluated using hot water extracts from grape juice pomace followed by aerobic yeast fermentation to remove sugars and comparison to reversed phase C(18) and Amberlite XAD-4. Extracts were also concentrated using spray-drying and vacuum evaporation. Fermentation had a minor impact on the retention of most polyphenolic compounds evaluated, yet resulted in a 16.3% decrease in antioxidant capacity. Spray-drying resulted in a 30.3% loss in total anthocyanins, a 21.5% loss in total phenolics, and a 23.3% decrease in antioxidant activity, whereas vacuum evaporation had no deleterious impact on these parameters. The physiology of the muscadine grape and its unique phytochemical composition has limited utilization of pomace from wine and juice manufacture. However, these studies demonstrated the potential to extract and concentrate polyphenolic-rich extracts for use in value-added applications.

Ag nanoparticles agargel nanocomposites for SERS detection of cultural heritage interest pigments

NASA Astrophysics Data System (ADS)

Amato, F.; Micciche', C.; Cannas, M.; Gelardi, F. M.; Pignataro, B.; Li Vigni, M.; Agnello, S.

2018-02-01

Agarose gel (agargel) composites with commercial and laboratory made silver nanoparticles were prepared by a wet solution method at room temperature. The gel composites were used for pigment extraction and detection by Raman spectroscopy. Red (alizarin) and violet (crystal violet) pigments deposited on paper were extracted by the composites and were investigated by micro-Raman spectroscopy. Evaluation was carried out of the surface-enhanced Raman spectroscopy (SERS) effect induced by the silver nanoparticles embedded in the gel. A kinetic approach as a function of time was used to determine the efficiency of pigments extraction by composites deposition. A non-invasive extraction process of few minutes is demonstrated. This process induces active SERS for both used pigments. The reported results show the full exploitability of agargel silver nanoparticle composites for the extraction of pigments from paper based artworks.

Development and validation of an automated liquid-liquid extraction GC/MS method for the determination of THC, 11-OH-THC, and free THC-carboxylic acid (THC-COOH) from blood serum.

PubMed

Purschke, Kirsten; Heinl, Sonja; Lerch, Oliver; Erdmann, Freidoon; Veit, Florian

2016-06-01

The analysis of Δ(9)-tetrahydrocannabinol (THC) and its metabolites 11-hydroxy-Δ(9)-tetrahydrocannabinol (11-OH-THC), and 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol (THC-COOH) from blood serum is a routine task in forensic toxicology laboratories. For examination of consumption habits, the concentration of the phase I metabolite THC-COOH is used. Recommendations for interpretation of analysis values in medical-psychological assessments (regranting of driver's licenses, Germany) include threshold values for the free, unconjugated THC-COOH. Using a fully automated two-step liquid-liquid extraction, THC, 11-OH-THC, and free, unconjugated THC-COOH were extracted from blood serum, silylated with N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA), and analyzed by GC/MS. The automation was carried out by an x-y-z sample robot equipped with modules for shaking, centrifugation, and solvent evaporation. This method was based on a previously developed manual sample preparation method. Validation guidelines of the Society of Toxicological and Forensic Chemistry (GTFCh) were fulfilled for both methods, at which the focus of this article is the automated one. Limits of detection and quantification for THC were 0.3 and 0.6 μg/L, for 11-OH-THC were 0.1 and 0.8 μg/L, and for THC-COOH were 0.3 and 1.1 μg/L, when extracting only 0.5 mL of blood serum. Therefore, the required limit of quantification for THC of 1 μg/L in driving under the influence of cannabis cases in Germany (and other countries) can be reached and the method can be employed in that context. Real and external control samples were analyzed, and a round robin test was passed successfully. To date, the method is employed in the Institute of Legal Medicine in Giessen, Germany, in daily routine. Automation helps in avoiding errors during sample preparation and reduces the workload of the laboratory personnel. Due to its flexibility, the analysis system can be employed for other liquid-liquid extractions as well. To the best of our knowledge, this is the first publication on a comprehensively automated classical liquid-liquid extraction workflow in the field of forensic toxicological analysis. Graphical abstract GC/MS with MPS Dual Head at the Institute of Legal Medicine, Giessen, Germany. Modules from left to right: (quick) Mix (for LLE), wash station, tray 1 (vials for extracts), solvent reservoir, (m) VAP (for extract evaporation), Solvent Filling Station (solvent supply), cooled tray 2 (vials for serum samples), and centrifuge (for phase separation).

Rapid fusion method for the determination of refractory thorium and uranium isotopes in soil samples

DOE PAGES

Maxwell, Sherrod L.; Hutchison, Jay B.; McAlister, Daniel R.

2015-02-14

Recently, approximately 80% of participating laboratories failed to accurately determine uranium isotopes in soil samples in the U.S Department of Energy Mixed Analyte Performance Evaluation Program (MAPEP) Session 30, due to incomplete dissolution of refractory particles in the samples. Failing laboratories employed acid dissolution methods, including hydrofluoric acid, to recover uranium from the soil matrix. The failures illustrate the importance of rugged soil dissolution methods for the accurate measurement of analytes in the sample matrix. A new rapid fusion method has been developed by the Savannah River National Laboratory (SRNL) to prepare 1-2 g soil sample aliquots very quickly, withmore » total dissolution of refractory particles. Soil samples are fused with sodium hydroxide at 600 ºC in zirconium crucibles to enable complete dissolution of the sample. Uranium and thorium are separated on stacked TEVA and TRU extraction chromatographic resin cartridges, prior to isotopic measurements by alpha spectrometry on cerium fluoride microprecipitation sources. Plutonium can also be separated and measured using this method. Batches of 12 samples can be prepared for measurement in <5 hours.« less

Microchip-based cell lysis and DNA extraction from sperm cells for application to forensic analysis.

PubMed

Bienvenue, Joan M; Duncalf, Natalie; Marchiarullo, Daniel; Ferrance, Jerome P; Landers, James P

2006-03-01

The current backlog of casework is among the most significant challenges facing crime laboratories at this time. While the development of next-generation microchip-based technology for expedited forensic casework analysis offers one solution to this problem, this will require the adaptation of manual, large-volume, benchtop chemistry to small volume microfluidic devices. Analysis of evidentiary materials from rape kits where semen or sperm cells are commonly found represents a unique set of challenges for on-chip cell lysis and DNA extraction that must be addressed for successful application. The work presented here details the development of a microdevice capable of DNA extraction directly from sperm cells for application to the analysis of sexual assault evidence. A variety of chemical lysing agents are assessed for inclusion in the extraction protocol and a method for DNA purification from sperm cells is described. Suitability of the extracted DNA for short tandem repeat (STR) analysis is assessed and genetic profiles shown. Finally, on-chip cell lysis methods are evaluated, with results from fluorescence visualization of cell rupture and DNA extraction from an integrated cell lysis and purification with subsequent STR amplification presented. A method for on-chip cell lysis and DNA purification is described, with considerations toward inclusion in an integrated microdevice capable of both differential cell sorting and DNA extraction. The results of this work demonstrate the feasibility of incorporating microchip-based cell lysis and DNA extraction into forensic casework analysis.

Extracting metalworking fluid aerosol samples in cassettes by provisional ASTM and NIOSH methods.

PubMed

Harper, Martin

2002-01-01

Recent provisional methods for the determination of metalworking fluid aerosol in workplace air involve a solvent extraction procedure to separate the nonvolatile fraction of the fluid from insoluble material such as metal turnings and dirt. The procedure calls for preweighing a filter (W1) and assembling it into a cassette and taking a sample. In the laboratory the filter is removed from the cassette, desiccated to remove any collected water or other volatile substances, and weighed again (W2). The filter is then extracted in an organic solvent blend, allowed to dry, and weighed a final time (W3). The total weight collected by the filter is given by (W2-W1), and the weight of (nonvolatile) metalworking fluid collected is given by (W2-W3). The extraction step can take place within a cassette housing if it is relatively inert to the solvent blend used. The extraction of four metalworking fluids (straight oil, soluble oil, synthetic and semisynthetic) within disposable polypropylene cassettes was investigated using the same protocol used to evaluate the original method. For all fluids the extraction efficiency was greater than 95% with a precision better than 5%. The mean blank contribution to the extraction step was 16 micrograms. Blanks were also evaluated after storage, and after transport and storage. A small additional blank contribution could be removed by desiccation. The limits of detection and quantitation of the extraction step were calculated to be 28 and 94 micrograms, respectively.

Development and validation of LC-MS/MS method for the determination of Ochratoxin A and its metabolite Ochratoxin α in poultry tissues and eggs.

PubMed

Paoloni, Angela; Solfrizzo, Michele; Bibi, Rita; Pecorelli, Ivan

2018-05-04

The objective of this study was to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the determination of Ochratoxin A (OTA) and Ochratoxin α (OTα) in poultry tissues and eggs. The two toxins were extracted by a mixture of acetonitrile/water, purified with a reversed phase C18 solid phase extraction column (SPE) and determined by LC-MS/MS. The LC-MS/MS method performances were evaluated in terms of linearity in solvent and in matrix (ranged from 0.5 to 15.10 µg L -1 for OTA and from 0.60 to 17.85 µg L -1 for OTα), limit of detection (LOD), limit of quantitation (LOQ), specificity, accuracy and precision in repeatability conditions. Recovery experiments were performed by spiking poultry liver, kidney, muscle and eggs around 1 µg kg -1 and 10 µg kg -1 . LODs were 0.27 and 0.26 µg kg -1 while LOQs were fixed at 1.0 and 1.2 µg kg -1 for OTA and OTα, respectively. Main recoveries for OTA ranged from 82 to 109% and for OTα ranged from 55 to 89%. The values of within-laboratory relative standard deviation (RSD r ) were equal to or below 20%. Considering the results obtained and that all analytical performance criteria were fulfilled, the new extraction and purification method developed for OTA and OTα determination in animal tissues and eggs was found appropriate for control laboratories and research activities designed to ensure food safety.

Quantifying MMA by SLE LC-MS/MS: Unexpected challenges in assay development.

PubMed

Lo, Sheng-Ying; Gordon, Cindy; Sadilkova, Katerina; Jack, Rhona M; Dickerson, Jane A

2016-09-01

Analysis of serum/plasma methylmalonic acid (MMA) is important for the diagnosis and management of methylmalonic acidemia in pediatric populations. This work focuses on developing and validating a liquid chromatography tandem mass spectrometry (LC-MS/MS) method to monitor methylmalonic acidemia using a simple method preparation. MMA and stable isotope labeled d3-MMA was extracted using supported liquid extraction (SLE). Assay imprecision, bias, linearity, recovery and carryover were determined. The relationship between MMA and propionyl acylcarnitine (C3-acylcarnitine) was also evaluated using historical paired results from 51 unique individuals. Baseline separation between MMA and succinic acid was completed in 7min. The assay was linear from 0.1 to 500μM. The intra-day and inter-day imprecision CV ranged from 4.1 to 13.2% (0.3 to 526μM) and 5.0 to 15.7% (0.3 to 233μM), respectively. Recovery ranged from 93 to 125%. The correlation with a national reference laboratory LC-MS/MS assay showed a Deming regression of 1.026 and intercept of -1.335. Carryover was determined to be <0.04%. Patient-specific correlation was observed between MMA and C3-acylcarnitine. This report describes the first LC-MS/MS method using SLE for MMA extraction. In addition, we illustrate the challenges encountered during this method development that should be assessed and resolved by any laboratory implementing a SLE LC-MS/MS assay designed to quantify analytes across several orders of magnitude. Copyright © 2016 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

A novel method for room temperature distribution and conservation of RNA and DNA reference materials for guaranteeing performance of molecular diagnostics in onco-hematology: A GBMHM study.

PubMed

Cayuela, Jean-Michel; Mauté, Carole; Fabre, Anne-Lise; Nibourel, Olivier; Dulucq, Stéphanie; Delabesse, Eric; Villarèse, Patrick; Hayette, Sandrine; Mozziconacci, Marie-Joelle; Macintyre, Elizabeth

2015-10-01

Performance of methods used for molecular diagnostics must be closely controlled by regular analysis of internal quality controls. However, conditioning, shipping and long lasting storage of nucleic acid controls remain problematic. Therefore, we evaluated the minicapsule-based innovative process developed by Imagene (Evry, France) for implementing DNA and RNA controls designed for clonality assessment of lymphoproliferations and BCR-ABL1 mRNA quantification, respectively. DNA samples were extracted from 12 cell lines selected for giving specific amplifications with most BIOMED-2 PCR tubes. RNA samples were extracted from 8 cell line mixtures expressing various BCR-ABL1 transcript levels. DNA and RNA were encapsulated by Imagene and shipped at room temperature to participating laboratories. Biologists were asked to report quality data of recovered nucleic acids as well as PCR results. Encapsulated nucleic acids samples were easily and efficiently recovered from minicapsules. The expected rearrangements at immunoglobulin, T-cell receptor and BCL2 loci were detected in DNA samples by all laboratories. Quality of RNA was consistent between laboratories and met the criteria requested for quantification of BCR-ABL1 transcripts. Expression levels measured by the 5 laboratories were within ±2 fold interval from the corresponding pre-encapsulation reference value. Moreover aging studies of encapsulated RNA simulating up to 100 years storage at room temperature show no bias in quantitative outcome. Therefore, Imagene minicapsules are suitable for storage and distribution at room temperature of genetic material designed for proficiency control of molecular diagnostic methods based on end point or real-time quantitative PCR. Copyright © 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Optimization of the extraction and purification of the compatible solute ectoine from Halomonas elongate in the laboratory experiment of a commercial production project.

PubMed

Chen, Ruifeng; Zhu, Lijun; Lv, Lihuo; Yao, Su; Li, Bin; Qian, Junqing

2017-06-01

Optimization of compatible solutes (ectoine) extraction and purification from Halomonas elongata cell fermentation had been investigated in the laboratory tests of a large scale commercial production project. After culturing H. elongata cells in developed medium at 28 °C for 23-30 h, we obtained an average yield and biomass of ectoine for 15.9 g/L and 92.9 (OD 600 ), respectively. Cell lysis was performed with acid treatment at moderate high temperature (60-70 °C). The downstream processing operations were designed to be as follows: filtration, desalination, cation exchange, extraction of crude product and three times of refining. Among which the cation exchange and extraction of crude product acquired a high average recovery rate of 95 and 96%; whereas a great loss rate of 19 and 15% was observed during the filtration and desalination, respectively. Combined with the recovering of ectoine from the mother liquor of the three times refining, the average of overall yield (referring to the amount of ectoine synthesized in cells) and purity of final product obtained were 43% and over 98%, respectively. However, key factors that affected the production efficiency were not yields but the time used in the extraction of crude product, involving the crystallization step from water, which spended 24-72 h according to the production scale. Although regarding to the productivity and simplicity on laboratory scale, the method described here can not compete with other investigations, in this study we acquired higher purity of ectoine and provided downstream processes that are capable of operating on industrial scale.

Multilaboratory evaluation of methods for detecting enteric viruses in soils.

PubMed Central

Hurst, C J; Schaub, S A; Sobsey, M D; Farrah, S R; Gerba, C P; Rose, J B; Goyal, S M; Larkin, E P; Sullivan, R; Tierney, J T

1991-01-01

Two candidate methods for the recovery and detection of viruses in soil were subjected to round robin comparative testing by members of the American Society for Testing and Materials D19:24:04:04 Subcommittee Task Group. Selection of the methods, designated "Berg" and "Goyal," was based on results of an initial screening which indicated that both met basic criteria considered essential by the task group. Both methods utilized beef extract solutions to achieve desorption and recovery of viruses from representative soils: a fine sand soil, an organic muck soil, a sandy loam soil, and a clay loam soil. One of the two methods, Goyal, also used a secondary concentration of resulting soil eluants via low-pH organic flocculation to achieve a smaller final assay volume. Evaluation of the two methods was simultaneously performed in replicate by nine different laboratories. Each of the produced samples was divided into portions, and these were respectively subjected to quantitative viral plaque assay by both the individual, termed independent, laboratory which had done the soil processing and a single common reference laboratory, using a single cell line and passage level. The Berg method seemed to produce slightly higher virus recovery values; however, the differences in virus assay titers for samples produced by the two methods were not statistically significant (P less than or equal to 0.05) for any one of the four soils. Despite this lack of a method effect, there was a statistically significant laboratory effect exhibited by assay titers from the independent versus reference laboratories for two of the soils, sandy loam and clay loam. PMID:1849712

Multilaboratory evaluation of methods for detecting enteric viruses in soils.

PubMed

Hurst, C J; Schaub, S A; Sobsey, M D; Farrah, S R; Gerba, C P; Rose, J B; Goyal, S M; Larkin, E P; Sullivan, R; Tierney, J T

1991-02-01

Two candidate methods for the recovery and detection of viruses in soil were subjected to round robin comparative testing by members of the American Society for Testing and Materials D19:24:04:04 Subcommittee Task Group. Selection of the methods, designated "Berg" and "Goyal," was based on results of an initial screening which indicated that both met basic criteria considered essential by the task group. Both methods utilized beef extract solutions to achieve desorption and recovery of viruses from representative soils: a fine sand soil, an organic muck soil, a sandy loam soil, and a clay loam soil. One of the two methods, Goyal, also used a secondary concentration of resulting soil eluants via low-pH organic flocculation to achieve a smaller final assay volume. Evaluation of the two methods was simultaneously performed in replicate by nine different laboratories. Each of the produced samples was divided into portions, and these were respectively subjected to quantitative viral plaque assay by both the individual, termed independent, laboratory which had done the soil processing and a single common reference laboratory, using a single cell line and passage level. The Berg method seemed to produce slightly higher virus recovery values; however, the differences in virus assay titers for samples produced by the two methods were not statistically significant (P less than or equal to 0.05) for any one of the four soils. Despite this lack of a method effect, there was a statistically significant laboratory effect exhibited by assay titers from the independent versus reference laboratories for two of the soils, sandy loam and clay loam.

Determination of Aristolochic Acid in Botanicals and Dietary Supplements by Liquid Chromatography with Ultraviolet Detection and by Liquid Chromatography/Mass Spectrometry: Single Laboratory Validation Confirmation

PubMed Central

Trujillo, William A.; Sorenson, Wendy R.; La Luzerne, Paul; Austad, John W.; Sullivan, Darryl

2008-01-01

The presence of aristolochic acid in some dietary supplements is a concern to regulators and consumers. A method has been developed, by initially using a reference method as a guide, during single laboratory validation (SLV) for the determination of aristolochic acid I, also known as aristolochic acid A, in botanical species and dietary supplements at concentrations of approximately 2 to 32 μg/g. Higher levels were determined by dilution to fit the standard curve. Through the SLV, the method was optimized for quantification by liquid Chromatography with ultraviolet detection (LC-UV) and LC/mass Spectrometry (MS) confirmation. The test samples were extracted with organic solvent and water, then injected on a reverse phase LC column. Quantification was achieved with linear regression using a laboratory automation system. The SLV study included systematically optimizing the LC-UV method with regard to test sample size, fine grinding of solids, and solvent extraction efficiency. These parameters were varied in increments (and in separate optimization studies), in order to ensure that each parameter was individually studied; the test results include corresponding tables of parameter variations. In addition, the chromatographic conditions were optimized with respect to injection volume and detection wavelength. Precision studies produced overall relative standard deviation values from 2.44 up to 8.26% for aristolochic acid I. Mean recoveries were between 100 and 103% at the 2 μg/g level, between 102 and 103% at the 10 μg/g level, and 104% at the 30 μg/g level. PMID:16915829

Determination of aristolochic acid in botanicals and dietary supplements by liquid chromatography with ultraviolet detection and by liquid chromatography/mass spectrometry: single laboratory validation confirmation.

PubMed

Trujillo, William A; Sorenson, Wendy R; La Luzerne, Paul; Austad, John W; Sullivan, Darryl

2006-01-01

The presence of aristolochic acid in some dietary supplements is a concern to regulators and consumers. A method has been developed, by initially using a reference method as a guide, during single laboratory validation (SLV) for the determination of aristolochic acid I, also known as aristolochic acid A, in botanical species and dietary supplements at concentrations of approximately 2 to 32 microg/g. Higher levels were determined by dilution to fit the standard curve. Through the SLV, the method was optimized for quantification by liquid chromatography with ultraviolet detection (LC-UV) and LC/mass spectrometry (MS) confirmation. The test samples were extracted with organic solvent and water, then injected on a reverse phase LC column. Quantification was achieved with linear regression using a laboratory automation system. The SLV study included systematically optimizing the LC-UV method with regard to test sample size, fine grinding of solids, and solvent extraction efficiency. These parameters were varied in increments (and in separate optimization studies), in order to ensure that each parameter was individually studied; the test results include corresponding tables of parameter variations. In addition, the chromatographic conditions were optimized with respect to injection volume and detection wavelength. Precision studies produced overall relative standard deviation values from 2.44 up to 8.26% for aristolochic acid I. Mean recoveries were between 100 and 103% at the 2 microg/g level, between 102 and 103% at the 10 microg/g level, and 104% at the 30 microg/g level.

A comparison of residual smear layer and erosion following different endodontic irrigation protocols tested under clinical and laboratory conditions.

PubMed

Cehreli, Zafer C; Uyanik, M Ozgur; Nagas, Emre; Tuncel, Behram; Er, Nuray; Comert, Fugen Dagli

2013-09-01

To compare the smear layer removal efficacy and erosive effects of different irrigation protocols under clinical and laboratory conditions. Mandibular third molars (n = 32) of 30-45 year-old patients were instrumented with rotary files and were randomly assigned to one of the following groups for final irrigation: (1) 5.25% NaOCl; (2) 17% EDTA; and (3) BioPure MTAD. Thereafter, the teeth were immediately extracted and processed for micromorphological investigation. In vitro specimen pairs were prepared by repeating the clinical experiments on freshly-extracted mandibular third molars. To compare open and closed systems, laboratory experiments were repeated on 32 additional teeth with enlarged apical foramen. The cleanliness of the root canals and the extent of erosion were assessed by environmental scanning electron microscopy. Specimens prepared under clinical and laboratory conditions had similar cleanliness and erosion scores (p > 0.05). Under both conditions, the tested solutions were more effective in removing the smear layer in the coronal and middle regions than in the apical one. Comparison of closed and open systems showed similar levels of cleanliness and erosion in all regions (p > 0.05), with the exception of 17% EDTA showing significantly higher levels of cleanliness and erosion in the apical third of open-end specimens. Based on clinical correlates of in vitro root canal cleanliness and erosion, laboratory testing of root canal irrigants on extracted teeth with closed apices can serve as a reliable method to simulate the clinical condition. EDTA was the most effective final irrigation solution in removing the smear layer at the expense of yielding the greatest erosive effect.

Laboratory Evaluation of Australian Ration Packs.

DTIC Science & Technology

1982-09-01

Monitoring of Freeze Dried Meals , Potato & Onion Powder .......................... 40 Appendix 6 Emergency Flying Ration ............................ 41...Moulds were enumerated using the pour plate method as described in AS 1766 Part 2.1.2 1976. On occasions malt extract agar was used in place of...potato dextrose agar . Coliforms & E. coli were enumerated using the pour plate method as described in AS 1766 Part 2.1.3.7 1976. Violet red bile dextrose

Method for the Determination of Aconitum Alkaloids in Dietary Supplements and Raw Materials by Reversed-Phase Liquid Chromatography with Ultraviolet Detection and Confirmation by Tandem Mass Spectrometry: Single-Laboratory Validation

PubMed Central

Tang, Wai-Tong; Wong, Sui-Kay; Law, Tin-Yau; Pang, Kwok-Chu; Sin, Della; Tam, Yin-King

2008-01-01

A study of single-laboratory validation (SLV) of a reversed-phase liquid Chromatography (RP-LC) method was conducted for the determination of diester-diterpene Aconitum alkaloids, viz., aconitine, mesaconitine, and hypaconitine, in a variety of dietary supplements, including single-arid multiple-ingredient dry powder extracts, pills, capsules, and raw materials. The Aconitum alkaloids in the samples were extracted by diethyl ether in the presence of ammonia. After cleanup with solid-phase extraction to remove the matrix interferences, the alkaloids were determined by RP-LC with UV detection at 235 nm, and the results were confirmed by tandem mass Spectrometry. The linear responses for aconitine, mesaconitine, and hypaconitine based on the present LC system ranged from 0.5 to 200 μg/mL. Relative standard deviations of 2.0 to 6.9% were obtained from duplicate analysis of 6 test materials of different matrixes for the 3 Aconitum alkaloids performed by 2 analysts on 5 different days. The recoveries determined for supplements and raw materials spiked with 3 Aconitum alkaloids at levels of 2.5–10 μg/g were in the range of 86–99%. In view of the attainment of satisfactory results for accuracy, precision, and recovery in the SLV study, it is recommended that the method validation process proceed to a collaborative study. PMID:17225594

Identification and quantification of ethylene oxide in sterilized medical devices using multiple headspace GC/MS measurement.

PubMed

Gimeno, Pascal; Auguste, Marie-Laure; Handlos, Vagn; Nielsen, Anne Mette; Schmidt, Stephan; Lassu, Nelly; Vogel, Martin; Fischer, Antonius; Brenier, Charlotte; Duperray, Françoise

2018-05-22

This manuscript, based on the ISO 10993-7 approach, describes a multiple HS-GC measurement of residual EO present in sterilized plastic samples. The quantification of EO is done, according to the ISO standard, by addition of EO amounts extracted for each repeated extraction. During the method development, the specificity of the detection of EO regarding acetaldehyde (structural isomer of EO) which may be formed from EO has been ensured and different tests were performed to check a possible influence of the sample preparation. Assays to maximize EO extraction were performed for different materials (Cyclo-olefine Copolymer (COC), Cyclo-olefine Polymer (COP), Silicon, Polyurethane (PUR)) changing extraction temperatures and times for the headspace and the pre-thermal treatment. Results highlight that depending on the material, EO can be more or less retained and thus thermal extraction conditions to maximize the amount of extractible EO from plastics may change accordingly. For COC syringes a validation according to ICH guidelines and an inter-laboratories study were performed. The method has been used for a market survey of EO sterilized medical devices, results obtained are reported in this manuscript. Copyright © 2018 Elsevier B.V. All rights reserved.

Effect of Harsh or Mild Extraction of Soil on Pesticide Leaching to Groundwater.

PubMed

Boesten, Jos J T I

2016-07-01

Assessment of leaching to groundwater is an important aspect of pesticide risk assessment. The first leaching tier usually consists of simulations with leaching scenarios based on pesticide-soil properties derived from laboratory studies. Because the extractability of pesticide residues in such studies decreases with time, the harshness of the extraction method influences these pesticide-soil properties. This study investigates the effect of using a mild or harsh extraction method on simulated leaching to groundwater with consideration of substances with a range of half-lives and organic matter sorption coefficient values for selected leaching scenarios. The model for linking the concentrations of the mild and the harsh systems was based on laboratory studies with two pesticides and a Dutch sandy soil and was tested against Canadian field studies with atrazine (6-chloro-2-ethyl-4-isopropyl-1,3,5-triazine-2,4-diamine). The degradation rate and the aged-sorption parameters of each "mild" soil-substance system were derived from a hypothetical laboratory incubation study using prescribed parameter values for the corresponding "harsh" soil-substance system. Simulations were performed for three European leaching scenarios (United Kingdom, France, Portugal). For the best-guess parameter set, the leaching concentrations of the harsh system were approximately equal to those of the mild system at leaching concentrations greater than 1 μg L and were at most approximately a factor of two higher than those of the mild systems at mild leaching concentrations between 0.01 and 0.1 μg L. However, an extreme parameter set led to harsh leaching concentrations that were at most approximately 10 times higher than the mild leaching concentrations at levels between 0.01 and 0.1 μg L. Copyright © by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America, Inc.

Round robin investigation of methods for the recovery of poliovirus from drinking water.

PubMed Central

Melnick, J L; Safferman, R; Rao, V C; Goyal, S; Berg, G; Dahling, D R; Wright, B A; Akin, E; Stetler, R; Sorber, C

1984-01-01

Six laboratories actively involved in water virology research participated in a methods evaluation study, conducted under the auspices of the American Society for Testing and Materials Committee on Viruses in the Aquatic Environment, Task Force on Drinking Water. Each participant was asked to examine the Viradel (virus adsorption-elution) method with cartridge-type Filterite filters for virus adsorption and organic flocculation and aluminum hydroxide-hydroextraction for reconcentration. Virus was adsorbed to filter media at pH 3.5 and eluted with either glycine buffer (pH 10.5) or beef extract-glycine (pHG 9.0). Considerable variation was noted in the quantity of virus recovered from four 100-liter samples of dechlorinated tapwater seeded with low (350 to 860 PFU) and high (1,837 to 4,689 PFU) doses of poliovirus type 1. To have a more uniform standard of comparison, all the test samples were reassayed in one laboratory, where titers were also determined for the virus seed. Test results of the Viradel-organic flocculation method indicated that the average percentage of virus recovery for low-input experiments was 66%, with a range of 8 to 20% in two laboratories, 49 to 63% in three laboratories, and 198% in one laboratory. For the high-input experiments, two laboratories reported recoveries of 6 to 12%, and four laboratories reported recoveries of 26 to 46%. For the Viradel aluminum hydroxide-hydroextraction procedure, two laboratories recovered 9 to 11%, whereas four obtained 17 to 34% for low-input experiments. For the high-input tests, two laboratories reported a recovery of 3 to 5%, and four recovered 11 to 18% of the seeded virus.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6320720

Determination of Aflatoxins and Ochratoxin A in Traditional Turkish Concentrated Fruit Juice Products by Multi-Immunoaffinity Column Cleanup and LC Fluorescence Detection: Single-Laboratory Validation.

PubMed

Kaymak, Tugrul; Türker, Levent; Tulay, Hüseyin; Stroka, Joerg

2018-04-27

Background : Pekmez and pestil are traditional Turkish foods made from concentrated grapejuice, which can be contaminated with mycotoxins such as aflatoxins and ochratoxin A (OTA). Objective : To carry out a single-laboratory validation of a method to simultaneously determine aflatoxins B 1 , B₂, G 1 , and G₂ and ochratoxin A in pekmez and pestil. Methods : The homogenized sample is extracted with methanol-water (80 + 20) using a high-speed blender. The (sample) extract is filtered, diluted with phosphate-buffered saline solution, and applied to a multi-immunoaffinity column (AFLAOCHRA PREP®). Aflatoxins and ochratoxin A are removed with (neat) methanol and then directly analyzed by reversed-phase LC with fluorescence detection using post-column bromination (Kobra cell®). Results : Test portions of blank pekmez and pestil were spiked with a mixture of aflatoxins and ochratoxin A to give levels ranging from 2.6 to 10.4 μg/kg and 1.0-4.0 μg/kg, respectively. Recoveries for total aflatoxins and ochratoxin A ranged from 84 to 106% and 80-97%, respectively, for spiked samples. Based on results for spiked pekmez and pestil (30 replicates each at three levels), the repeatability RSD ranged from 1.6 to 12% and 2.7-11% for total aflatoxins and ochratoxin A, respectively. Conclusions : The method performance in terms of recovery, repeatability, and detection limits has been demonstrated to be suitable for use as an Official Method. Highlights : First immunoaffinity column method validated for simultaneous analysis of aflatoxins and ochratoxin A in pekmez and pestil. Suitability for use for official purposes in Turkey, demonstrated by single-laboratory validation. Co-occurrence of aflatoxins and OTA in mulberry and carob pekmez reported for the first time.

New geothermal heat extraction process to deliver clean power generation

ScienceCinema

McGrail, Pete

2017-12-27

A new method for capturing significantly more heat from low-temperature geothermal resources holds promise for generating virtually pollution-free electrical energy. Scientists at the Department of Energys Pacific Northwest National Laboratory will determine if their innovative approach can safely and economically extract and convert heat from vast untapped geothermal resources. The goal is to enable power generation from low-temperature geothermal resources at an economical cost. In addition to being a clean energy source without any greenhouse gas emissions, geothermal is also a steady and dependable source of power.

Evaluation of a low-cost commercially available extraction device for assessing lead bioaccessibility in contaminated soils.

PubMed

Nelson, Clay M; Gilmore, Thomas M; Harrington, M; Scheckel, Kirk G; Miller, Bradley W; Bradham, Karen D

2013-03-01

The U.S. EPA's in vitro bioaccessibility (IVBA) method 9200.1-86 defines a validated analytical procedure for the determination of lead bioaccessibility in contaminated soils. The method requires the use of a custom-fabricated extraction device that uses a heated water bath for sample incubation. In an effort to improve ease of use, increase sample throughput, and reduce equipment acquisition and maintenance costs, an alternative low-cost, commercially available extraction device capable of sample incubation via heated air and end-over-end rotation was evaluated. An intra-laboratory study was conducted to compare lead bioaccessibility values derived using the two extraction devices. IVBA values were not statistically different (α = 0.05) between the two extraction devices for any of the soils (n = 6) evaluated in this study, with an average difference in mean lead IVBA of 0.8% (s.d. = 0.5%). The commercially available extraction device was able to generate accurate lead IVBA data as compared to the U.S. EPA's expected value for a National Institute of Standards and Technology standard reference material soil. The relative percent differences between high and low IVBA values for each soil, a measure of instrument precision, were also not statistically different (α = 0.05) between the two extraction devices. The statistical agreement of lead IVBA values observed using the two extraction devices supports the use of a low-cost, commercially available extraction device as a reliable alternative to a custom-fabricated device as required by EPA method 9200.1-86.

International Study to Evaluate PCR Methods for Detection of Trypanosoma cruzi DNA in Blood Samples from Chagas Disease Patients

PubMed Central

Schijman, Alejandro G.; Bisio, Margarita; Orellana, Liliana; Sued, Mariela; Duffy, Tomás; Mejia Jaramillo, Ana M.; Cura, Carolina; Auter, Frederic; Veron, Vincent; Qvarnstrom, Yvonne; Deborggraeve, Stijn; Hijar, Gisely; Zulantay, Inés; Lucero, Raúl Horacio; Velazquez, Elsa; Tellez, Tatiana; Sanchez Leon, Zunilda; Galvão, Lucia; Nolder, Debbie; Monje Rumi, María; Levi, José E.; Ramirez, Juan D.; Zorrilla, Pilar; Flores, María; Jercic, Maria I.; Crisante, Gladys; Añez, Néstor; De Castro, Ana M.; Gonzalez, Clara I.; Acosta Viana, Karla; Yachelini, Pedro; Torrico, Faustino; Robello, Carlos; Diosque, Patricio; Triana Chavez, Omar; Aznar, Christine; Russomando, Graciela; Büscher, Philippe; Assal, Azzedine; Guhl, Felipe; Sosa Estani, Sergio; DaSilva, Alexandre; Britto, Constança; Luquetti, Alejandro; Ladzins, Janis

2011-01-01

Background A century after its discovery, Chagas disease still represents a major neglected tropical threat. Accurate diagnostics tools as well as surrogate markers of parasitological response to treatment are research priorities in the field. The purpose of this study was to evaluate the performance of PCR methods in detection of Trypanosoma cruzi DNA by an external quality evaluation. Methodology/Findings An international collaborative study was launched by expert PCR laboratories from 16 countries. Currently used strategies were challenged against serial dilutions of purified DNA from stocks representing T. cruzi discrete typing units (DTU) I, IV and VI (set A), human blood spiked with parasite cells (set B) and Guanidine Hidrochloride-EDTA blood samples from 32 seropositive and 10 seronegative patients from Southern Cone countries (set C). Forty eight PCR tests were reported for set A and 44 for sets B and C; 28 targeted minicircle DNA (kDNA), 13 satellite DNA (Sat-DNA) and the remainder low copy number sequences. In set A, commercial master mixes and Sat-DNA Real Time PCR showed better specificity, but kDNA-PCR was more sensitive to detect DTU I DNA. In set B, commercial DNA extraction kits presented better specificity than solvent extraction protocols. Sat-DNA PCR tests had higher specificity, with sensitivities of 0.05–0.5 parasites/mL whereas specific kDNA tests detected 5.10−3 par/mL. Sixteen specific and coherent methods had a Good Performance in both sets A and B (10 fg/µl of DNA from all stocks, 5 par/mL spiked blood). The median values of sensitivities, specificities and accuracies obtained in testing the Set C samples with the 16 tests determined to be good performing by analyzing Sets A and B samples varied considerably. Out of them, four methods depicted the best performing parameters in all three sets of samples, detecting at least 10 fg/µl for each DNA stock, 0.5 par/mL and a sensitivity between 83.3–94.4%, specificity of 85–95%, accuracy of 86.8–89.5% and kappa index of 0.7–0.8 compared to consensus PCR reports of the 16 good performing tests and 63–69%, 100%, 71.4–76.2% and 0.4–0.5, respectively compared to serodiagnosis. Method LbD2 used solvent extraction followed by Sybr-Green based Real time PCR targeted to Sat-DNA; method LbD3 used solvent DNA extraction followed by conventional PCR targeted to Sat-DNA. The third method (LbF1) used glass fiber column based DNA extraction followed by TaqMan Real Time PCR targeted to Sat-DNA (cruzi 1/cruzi 2 and cruzi 3 TaqMan probe) and the fourth method (LbQ) used solvent DNA extraction followed by conventional hot-start PCR targeted to kDNA (primer pairs 121/122). These four methods were further evaluated at the coordinating laboratory in a subset of human blood samples, confirming the performance obtained by the participating laboratories. Conclusion/Significance This study represents a first crucial step towards international validation of PCR procedures for detection of T. cruzi in human blood samples. PMID:21264349

Linking Laboratory Experiences to the Real World: The Extraction of Octylphenoxyacetic Acid from Water

ERIC Educational Resources Information Center

Loyo-Rosales, Jorge E.; Torrents, Alba; Rosales-Rivera, Georgina C.; Rice, Clifford C.

2006-01-01

Several chemical concepts to the extraction of a water pollutant OPC (octylphenoxyacetic acid) is presented. As an introduction to the laboratory experiment, a discussion on endocrine disrupters is conducted to familiarize the student with the background of the experiment and to explain the need for the extraction and quantitation of the OPC which…

Texture Analysis and Cartographic Feature Extraction.

DTIC Science & Technology

1985-01-01

Investigations into using various image descriptors as well as developing interactive feature extraction software on the Digital Image Analysis Laboratory...system. Originator-supplied keywords: Ad-Hoc image descriptor; Bayes classifier; Bhattachryya distance; Clustering; Digital Image Analysis Laboratory

Laboratory and quality assurance protocols for the analysis of herbicides in ground water from the Management Systems Evaluation Area, Princeton, Minnesota

USGS Publications Warehouse

Larson, S.J.; Capel, P.D.; VanderLoop, A.G.

1996-01-01

Laboratory and quality assurance procedures for the analysis of ground-water samples for herbicides at the Management Systems Evaluation Area near Princeton, Minnesota are described. The target herbicides include atrazine, de-ethylatrazine, de-isopropylatrazine, metribuzin, alachlor, 2,6-diethylaniline, and metolachlor. The analytical techniques used are solid-phase extraction, and analysis by gas chromatography with mass-selective detection. Descriptions of cleaning procedures, preparation of standard solutions, isolation of analytes from water, sample transfer methods, instrumental analysis, and data analysis are included.

Scaled-up remediation of CCA-treated wood

Treesearch

Carol A. Clausen; William R. Kenealy

2004-01-01

Bioremediation is a novel approach to recycling waste wood treated with chromated copper arsenate (CCA). Remediating CCA-treated waste wood diverts this fiber source from our landfills and provides tangible secondary products from the cleaned fiber. On a laboratory scale, this method, which utilizes oxalic acid extraction and bioleaching with a metal- tolerant...

Utilization of the wastes of vital activity

NASA Technical Reports Server (NTRS)

Gusarov, B. G.; Drigo, Y. A.; Novikov, V. M.; Samsonov, N. M.; Farafonov, N. S.; Chizhov, S. V.; Yazdovskiy, V. I.

1979-01-01

The recycling of wastes from the biological complex for use in life-support systems is discussed. Topics include laboratory equipment, heat treatment of waste materials, mineralization of waste products, methods for production of ammonium hydroxide and nitric acid, the extraction of sodium chloride from mineralized products, and the recovery of nutrient substances for plants from urine.

Easy Extraction of Roundworms from Earthworm Hosts.

ERIC Educational Resources Information Center

Eyster, Linda S.; Fried, Bernard

2000-01-01

Describes the inexpensive and safe method of using roundworms in the classroom or laboratories. Because parasitic infections are so common, students should learn about worms. Provides statistics on just how many people have a worm infection in the world. Explains how to study living nematodes, and obtain and use earthworms. (Contains 13…

Optimization of extractants, purifying packings, and eluents for analytical extraction of organochlorine pesticides in Hydragric Acrisols.

PubMed

Zhao, Zhenhua; Xia, Liling; Wang, Fang; Jiang, Xin; Gao, Yanzheng

2012-08-01

In this study, we screened for an economic, rapid, and efficient hypotoxic pretreatment method for organochlorine pesticides in soil samples for gas chromatography (GC) analysis. The analytical extraction efficiencies of 11 different extractants, nine types of solid-phase purification (SPP) cartridge packings, and three types of eluents for 13 organochlorine pesticides (OCPs) in spiked and natural Chinese red soil (Hydragric Acrisols) were evaluated using an ultrasonic extraction and solid-phase purification method. High percent recoveries (85-106%) were obtained for the 13 organochlorine pesticides in soil when petroleum ether/acetone/water (10:5:2, v/v) was used an extractant. They were purified using celite SPP cartridge packing and eluted with 9 mL of dichloromethane/petroleum ether (1:9, v/v). The OCPs purification pretreatment of Hydragric Acrisols, using the above method, meets the GC analysis requirements. Compared with other traditional pretreatment methods for OCPs in soil samples, this method has several advantages, such as a short extraction time, reducing the amount of solvent, having no emulsion phenomenon, and hypotoxicity to the laboratory technicians. The concentrations of 1,1,1,-trichloro-2(p-chlorophenyl)-2-(o-chlorophenyl) ethane (DDTs; 3.42-8.08 ng g(-1)) in field soils were higher than the hexachlorocyclohexane concentration (2.94-6.12 ng g(-1)). The 1,1-dichloro-2,2-bis(p-chlorophenyl) ethylene (p,p'-DDE) + 1,1-dichloro-2,2-bis(p-chlorophenyl)-ethane (p,p'-DDD)/p,p'-DDT ratio in this field soil was approximately 2.7, suggesting that no new DDT pollution source was introduced into the sampling site.

Occupational allergy to Spagulax®(Plantago ovata seed).

PubMed

Viñas, M; Pineda, F; Izquierdo-Domínguez, A; Castillo, M; Castillo, M J; Hernández, N; Ibero, M

2017-11-01

We report the case of a 36-year-old male pharmaceutical laboratory worker. On handling Spagulax ® sachets whose content is a laxative called Plantago ovata, he immediately presented rhinoconjunctivitis. Methods. Specific allergy study included SDS-PAGE with Western Blot and specific nasal challenge to Plantago ovata extract. Results. Prick by prick for Spagulax ® was negative. Total IgE: 126.5 U/mL. Western Blot recognized two proteins of 15 and 20 kDa in the extract of Plantago ovata and three proteins of 15, 18 and 50 kDa in the extract of Plantago lanceolata. Conclusions. We present a case of occupational allergy due to inhalation of and/or contact with Plantago ovata seeds.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; a method supplement for the determination of Fipronil and degradates in water by gas chromatography/mass spectrometry

USGS Publications Warehouse

Madsen, James F.; Sandstrom, Mark W.; Zaugg, Steven D.

2002-01-01

A method for the isolation and detemrination of fipronil and four of its degradates has been developed. This method adapts an analytical method created by the U.S. Geological Survey National Water Quality Laboratory in 1995 for the determination of a broad range of high-use pesticides typically found in filtered natural-water samples. In 2000, fipronil and four of its degradates were extracted, analyzed, and validated using this method. The recoveries for these five compounds in reagent-water samples fortified at 1 microgram per liter (ug/L) avereraged 98 percent. Initial method detection limits averaged 0.0029 ug/L. The performance of these five new compounds is consistent with the performance of the compounds in the initial method, making it possible to include them in addition to the other 41 pesticides and pesticide degradates in the original method.

Increasing productivity for the analysis of trace contaminants in food by gas chromatography-mass spectrometry using automated liner exchange, backflushing and heart-cutting.

PubMed

David, Frank; Tienpont, Bart; Devos, Christophe; Lerch, Oliver; Sandra, Pat

2013-10-25

Laboratories focusing on residue analysis in food are continuously seeking to increase sample throughput by minimizing sample preparation. Generic sample extraction methods such as QuEChERS lack selectivity and consequently extracts are not free from non-volatile material that contaminates the analytical system. Co-extracted matrix constituents interfere with target analytes, even if highly sensitive and selective GC-MS/MS is used. A number of GC approaches are described that can be used to increase laboratory productivity. These techniques include automated inlet liner exchange and column backflushing for preservation of the performance of the analytical system and heart-cutting two-dimensional GC for increasing sensitivity and selectivity. The application of these tools is illustrated by the analysis of pesticides in vegetables and fruits, PCBs in milk powder and coplanar PCBs in fish. It is demonstrated that considerable increase in productivity can be achieved by decreasing instrument down-time, while analytical performance is equal or better compared to conventional trace contaminant analysis. Copyright © 2013 Elsevier B.V. All rights reserved.

Application of solid-phase microextraction for in vivo laboratory and field sampling of pharmaceuticals in fish.

PubMed

Zhou, Simon Ningsun; Oakes, Ken D; Servos, Mark R; Pawliszyn, Janusz

2008-08-15

Previous field studies utilizing solid-phase microextraction (SPME) predominantly focused on volatile and semivolatile compounds in air or water. Earlier in vivo sampling studies utilizing SPME were limited to the liquid matrix (blood). The present study has expanded the SPME technique to semisolid tissues under laboratory and field conditions through the investigation of both theoretical and applied experimental approaches. Pre-equilibrium extraction and desorption were performed in vivo in two separate animals. Excellent linearity was found between the amounts extracted by SPME from the muscle of living fish and the waterborne concentrations of pharmaceuticals. A simple SPME method is also described to simultaneously determine free and total analyte concentrations in living tissue. The utility of in vivo SPME sampling was evaluated in wild fish collected from a number of different river locations under varying degrees of influence from municipal wastewater effluents. Diphenhydramine and diltiazem were detected in the muscle of fish downstream of a local wastewater treatment plant. Based on this study, SPME demonstrated several important advantages such as simplicity, sensitivity, and robustness under laboratory and in vivo field sampling conditions.

A reproducible and scalable procedure for preparing bacterial extracts for cell-free protein synthesis.

PubMed

Katsura, Kazushige; Matsuda, Takayoshi; Tomabechi, Yuri; Yonemochi, Mayumi; Hanada, Kazuharu; Ohsawa, Noboru; Sakamoto, Kensaku; Takemoto, Chie; Shirouzu, Mikako

2017-11-01

Cell-free protein synthesis is a useful method for preparing proteins for functional or structural analyses. However, batch-to-batch variability with regard to protein synthesis activity remains a problem for large-scale production of cell extract in the laboratory. To address this issue, we have developed a novel procedure for large-scale preparation of bacterial cell extract with high protein synthesis activity. The developed procedure comprises cell cultivation using a fermentor, harvesting and washing of cells by tangential flow filtration, cell disruption with high-pressure homogenizer and continuous diafiltration. By optimizing and combining these methods, ∼100 ml of the cell extract was prepared from 150 g of Escherichia coli cells. The protein synthesis activities, defined as the yield of protein per unit of absorbance at 260 nm of the cell extract, were shown to be reproducible, and the average activity of several batches was twice that obtained using a previously reported method. In addition, combinatorial use of the high-pressure homogenizer and diafiltration increased the scalability, indicating that the cell concentration at disruption varies from 0.04 to 1 g/ml. Furthermore, addition of Gam protein and examinations of the N-terminal sequence rendered the extract prepared here useful for rapid screening with linear DNA templates. © The Authors 2017. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Acidified pressurized hot water for the continuous extraction of cadmium and lead from plant materials prior to ETAAS

NASA Astrophysics Data System (ADS)

Morales-Muñoz, S.; Luque-García, J. L.; Luque de Castro, M. D.

2003-01-01

Acidified and pressurized hot water is proposed for the continuous leaching of Cd and Pb from plants prior to determination by electrothermal atomic absorption spectrometry. Beech leaves (a certified reference material—CRM 100—where the analytes were not certified) were used for optimizing the method by a multivariate approach. The samples (0.5 g) were subjected to dynamic extraction with water modified with 1% v/v HNO 3 at 250 °C as leachant. A kinetics study was performed in order to know the pattern of the extraction process. The method was validated with a CRM (olive leaves, 062 from the BCR) where the analytes had been certified. The agreement between the certified values and those found using the proposed method demonstrates its usefulness. The repeatability and within-laboratory reproducibility were 3.7 and 2.3% for Cd and 1.04% and 6.3% for Pb, respectively. The precision of the method, together with its efficiency, rapidity, and environmental acceptability, makes it a good alternative for the determination of trace metals in plant material.

A novel optimised and validated method for analysis of multi-residues of pesticides in fruits and vegetables by microwave-assisted extraction (MAE)-dispersive solid-phase extraction (d-SPE)-retention time locked (RTL)-gas chromatography-mass spectrometry with Deconvolution reporting software (DRS).

PubMed

Satpathy, Gouri; Tyagi, Yogesh Kumar; Gupta, Rajinder Kumar

2011-08-01

A rapid, effective and ecofriendly method for sensitive screening and quantification of 72 pesticides residue in fruits and vegetables, by microwave-assisted extraction (MAE) followed by dispersive solid-phase extraction (d-SPE), retention time locked (RTL) capillary gas-chromatographic separation in trace ion mode mass spectrometric determination has been validated as per ISO/IEC: 17025:2005. Identification and reporting with total and extracted ion chromatograms were facilitated to a great extent by Deconvolution reporting software (DRS). For all compounds LOD were 0.002-0.02mg/kg and LOQ were 0.025-0.100mg/kg. Correlation coefficients of the calibration curves in the range of 0.025-0.50mg/kg were >0.993. To validate matrix effects repeatability, reproducibility, recovery and overall uncertainty were calculated for the 35 matrices at 0.025, 0.050 and 0.100mg/kg. Recovery ranged between 72% and 114% with RSD of <20% for repeatability and intermediate precision. The reproducibility of the method was evaluated by an inter laboratory participation and Z score obtained within ±2. Copyright © 2011 Elsevier Ltd. All rights reserved.

Environmentally friendly microwave-assisted sequential extraction method followed by ICP-OES and ion-chromatographic analysis for rapid determination of sulphur forms in coal samples.

PubMed

Mketo, Nomvano; Nomngongo, Philiswa N; Ngila, J Catherine

2018-05-15

A rapid three-step sequential extraction method was developed under microwave radiation followed by inductively coupled plasma-optical emission spectroscopic (ICP-OES) and ion-chromatographic (IC) analysis for the determination of sulphur forms in coal samples. The experimental conditions of the proposed microwave-assisted sequential extraction (MW-ASE) procedure were optimized by using multivariate mathematical tools. Pareto charts generated from 2 3 full factorial design showed that, extraction time has insignificant effect on the extraction of sulphur species, therefore, all the sequential extraction steps were performed for 5 min. The optimum values according to the central composite designs and counter plots of the response surface methodology were 200 °C (microwave temperature) and 0.1 g (coal amount) for all the investigated extracting reagents (H 2 O, HCl and HNO 3 ). When the optimum conditions of the proposed MW-ASE procedure were applied in coal CRMs, SARM 18 showed more organic sulphur (72%) and the other two coal CRMs (SARMs 19 and 20) were dominated by sulphide sulphur species (52-58%). The sum of the sulphur forms from the sequential extraction steps have shown consistent agreement (95-96%) with certified total sulphur values on the coal CRM certificates. This correlation, in addition to the good precision (1.7%) achieved by the proposed procedure, suggests that the sequential extraction method is reliable, accurate and reproducible. To safe-guard the destruction of pyritic and organic sulphur forms in extraction step 1, water was used instead of HCl. Additionally, the notorious acidic mixture (HCl/HNO 3 /HF) was replaced by greener reagent (H 2 O 2 ) in the last extraction step. Therefore, the proposed MW-ASE method can be applied in routine laboratories for the determination of sulphur forms in coal and coal related matrices. Copyright © 2018 Elsevier B.V. All rights reserved.

Comparing Standard and Selective Degradation DNA Extraction Methods: Results from a Field Experiment with Sexual Assault Kits^.

PubMed

Campbell, Rebecca; Pierce, Steven J; Sharma, Dhruv B; Shaw, Jessica; Feeney, Hannah; Nye, Jeffrey; Schelling, Kristin; Fehler-Cabral, Giannina

2017-01-01

A growing number of U.S. cities have large numbers of untested sexual assault kits (SAKs) in police property facilities. Testing older kits and maintaining current case work will be challenging for forensic laboratories, creating a need for more efficient testing methods. We evaluated selective degradation methods for DNA extraction using actual case work from a sample of previously unsubmitted SAKs in Detroit, Michigan. We randomly assigned 350 kits to either standard or selective degradation testing methods and then compared DNA testing rates and CODIS entry rates between the two groups. Continuation-ratio modeling showed no significant differences, indicating that the selective degradation method had no decrement in performance relative to customary methods. Follow-up equivalence tests indicated that CODIS entry rates for the two methods could differ by more than ±5%. Selective degradation methods required less personnel time for testing and scientific review than standard testing. © 2016 American Academy of Forensic Sciences.

Manganese speciation of laboratory-generated welding fumes

PubMed Central

Andrews, Ronnee N.; Keane, Michael; Hanley, Kevin W.; Feng, H. Amy; Ashley, Kevin

2015-01-01

The objective of this laboratory study was to identify and measure manganese (Mn) fractions in chamber-generated welding fumes (WF) and to evaluate and compare the results from a sequential extraction procedure for Mn fractions with that of an acid digestion procedure for measurement of total, elemental Mn. To prepare Mn-containing particulate matter from representative welding processes, a welding system was operated in short circuit gas metal arc welding (GMAW) mode using both stainless steel (SS) and mild carbon steel (MCS) and also with flux cored arc welding (FCAW) and shielded metal arc welding (SMAW) using MCS. Generated WF samples were collected onto polycarbonate filters before homogenization, weighing and storage in scintillation vials. The extraction procedure consisted of four sequential steps to measure various Mn fractions based upon selective solubility: (1) soluble Mn dissolved in 0.01 M ammonium acetate; (2) Mn (0,II) dissolved in 25 % (v/v) acetic acid; (3) Mn (III,IV) dissolved in 0.5% (w/v) hydroxylamine hydrochloride in 25% (v/v) acetic acid; and (4) insoluble Mn extracted with concentrated hydrochloric and nitric acids. After sample treatment, the four fractions were analyzed for Mn by inductively coupled plasma-atomic emission spectroscopy (ICP-AES). WF from GMAW and FCAW showed similar distributions of Mn species, with the largest concentrations of Mn detected in the Mn (0,II) and insoluble Mn fractions. On the other hand, the majority of the Mn content of SMAW fume was detected as Mn (III,IV). Although the concentration of Mn measured from summation of the four sequential steps was statistically significantly different from that measured from the hot block dissolution method for total Mn, the difference is small enough to be of no practical importance for industrial hygiene air samples, and either method may be used for Mn measurement. The sequential extraction method provides valuable information about the oxidation state of Mn in samples and allows for comparison to results from previous work and from total Mn dissolution methods. PMID:26345630

Manganese speciation of laboratory-generated welding fumes.

PubMed

Andrews, Ronnee N; Keane, Michael; Hanley, Kevin W; Feng, H Amy; Ashley, Kevin

The objective of this laboratory study was to identify and measure manganese (Mn) fractions in chamber-generated welding fumes (WF) and to evaluate and compare the results from a sequential extraction procedure for Mn fractions with that of an acid digestion procedure for measurement of total, elemental Mn. To prepare Mn-containing particulate matter from representative welding processes, a welding system was operated in short circuit gas metal arc welding (GMAW) mode using both stainless steel (SS) and mild carbon steel (MCS) and also with flux cored arc welding (FCAW) and shielded metal arc welding (SMAW) using MCS. Generated WF samples were collected onto polycarbonate filters before homogenization, weighing and storage in scintillation vials. The extraction procedure consisted of four sequential steps to measure various Mn fractions based upon selective solubility: (1) soluble Mn dissolved in 0.01 M ammonium acetate; (2) Mn (0,II) dissolved in 25 % (v/v) acetic acid; (3) Mn (III,IV) dissolved in 0.5% (w/v) hydroxylamine hydrochloride in 25% (v/v) acetic acid; and (4) insoluble Mn extracted with concentrated hydrochloric and nitric acids. After sample treatment, the four fractions were analyzed for Mn by inductively coupled plasma-atomic emission spectroscopy (ICP-AES). WF from GMAW and FCAW showed similar distributions of Mn species, with the largest concentrations of Mn detected in the Mn (0,II) and insoluble Mn fractions. On the other hand, the majority of the Mn content of SMAW fume was detected as Mn (III,IV). Although the concentration of Mn measured from summation of the four sequential steps was statistically significantly different from that measured from the hot block dissolution method for total Mn, the difference is small enough to be of no practical importance for industrial hygiene air samples, and either method may be used for Mn measurement. The sequential extraction method provides valuable information about the oxidation state of Mn in samples and allows for comparison to results from previous work and from total Mn dissolution methods.

[Method validation according to ISO 15189 and SH GTA 04: application for the extraction of DNA and its quantitative evaluation by a spectrophotometric assay].

PubMed

Harlé, Alexandre; Lion, Maëva; Husson, Marie; Dubois, Cindy; Merlin, Jean-Louis

2013-01-01

According to the French legislation on medical biology (January 16th, 2010), all biological laboratories must be accredited according to ISO 15189 for at least 50% of their activities before the end of 2016. The extraction of DNA from a sample of interest, whether solid or liquid is one of the critical steps in molecular biology and specifically in somatic or constitutional genetic. The extracted DNA must meet a number of criteria such quality and also be in sufficient concentration to allow molecular biology assays such as the detection of somatic mutations. This paper describes the validation of the extraction and purification of DNA using chromatographic column extraction and quantitative determination by spectrophotometric assay, according to ISO 15189 and the accreditation technical guide in Human Health SH-GTA-04.

An optimized work-flow to reduce time-to-detection of carbapenemase-producing Enterobacteriaceae (CPE) using direct testing from rectal swabs.

PubMed

O'Connor, C; Kiernan, M G; Finnegan, C; O'Hara, M; Power, L; O'Connell, N H; Dunne, C P

2017-05-04

Rapid detection of patients with carbapenemase-producing Enterobacteriaceae (CPE) is essential for the prevention of nosocomial cross-transmission, allocation of isolation facilities and to protect patient safety. Here, we aimed to design a new laboratory work-flow, utilizing existing laboratory resources, in order to reduce time-to-diagnosis of CPE. A review of the current CPE testing processes and of the literature was performed to identify a real-time commercial polymerase chain reaction (PCR) assay that could facilitate batch testing of CPE clinical specimens, with adequate CPE gene coverage. Stool specimens (210) were collected; CPE-positive inpatients (n = 10) and anonymized community stool specimens (n = 200). Rectal swabs (eSwab™) were inoculated from collected stool specimens and a manual DNA extraction method (QIAamp® DNA Stool Mini Kit) was employed. Extracted DNA was then processed on the Check-Direct CPE® assay. The three step process of making the eSwab™, extracting DNA manually and running the Check-Direct CPE® assay, took <5 min, 1 h 30 min and 1 h 50 min, respectively. It was time efficient with a result available in under 4 h, comparing favourably with the existing method of CPE screening; average time-to-diagnosis of 48/72 h. Utilizing this CPE work-flow would allow a 'same-day' result. Antimicrobial susceptibility testing results, as is current practice, would remain a 'next-day' result. In conclusion, the Check-Direct CPE® assay was easily integrated into a local laboratory work-flow and could facilitate a large volume of CPE screening specimens in a single batch, making it cost-effective and convenient for daily CPE testing.

Determination of paralytic shellfish toxins in shellfish by receptor binding assay: collaborative study.

PubMed

Van Dolah, Frances M; Fire, Spencer E; Leighfield, Tod A; Mikulski, Christina M; Doucette, Gregory J

2012-01-01

A collaborative study was conducted on a microplate format receptor binding assay (RBA) for paralytic e shellfish toxins (PST). The assay quantifies the composite PST toxicity in shellfish samples based on the ability of sample extracts to compete with (3)H saxitoxin (STX) diHCl for binding to voltage-gated sodium channels in a rat brain membrane preparation. Quantification of binding can be carried out using either a microplate or traditional scintillation counter; both end points were included in this study. Nine laboratories from six countries completed the study. One laboratory analyzed the samples using the precolumn oxidation HPLC method (AOAC Method 2005.06) to determine the STX congener composition. Three laboratories performed the mouse bioassay (AOAC Method 959.08). The study focused on the ability of the assay to measure the PST toxicity of samples below, near, or slightly above the regulatory limit of 800 (microg STX diHCl equiv./kg). A total of 21 shellfish homogenates were extracted in 0.1 M HCl, and the extracts were analyzed by RBA in three assays on separate days. Samples included naturally contaminated shellfish samples of different species collected from several geographic regions, which contained varying STX congener profiles due to their exposure to different PST-producing dinoflagellate species or differences in toxin metabolism: blue mussel (Mytilus edulis) from the U.S. east and west coasts, California mussel (Mytilus californianus) from the U.S. west coast, chorito mussel (Mytilus chiliensis) from Chile, green mussel (Perna canaliculus) from New Zealand, Atlantic surf clam (Spisula solidissima) from the U.S. east coast, butter clam (Saxidomus gigantea) from the west coast of the United States, almeja clam (Venus antiqua) from Chile, and Atlantic sea scallop (Plactopecten magellanicus) from the U.S. east coast. All samples were provided as whole animal homogenates, except Atlantic sea scallop and green mussel, from which only the hepatopancreas was homogenized. Among the naturally contaminated samples, five were blind duplicates used for calculation of RSDr. The interlaboratory RSDR of the assay for 21 samples tested in nine laboratories was 33.1%, yielding a HorRat value of 2.0. Removal of results for one laboratory that reported systematically low values resulted in an average RSDR of 28.7% and average HorRat value of 1.8. Intralaboratory RSDr based on five blind duplicate samples tested in separate assays, was 25.1%. RSDr obtained by individual laboratories ranged from 11.8 to 34.9%. Laboratories that are routine users of the assay performed better than nonroutine users, with an average RSDr of 17.1%. Recovery of STX from spiked shellfish homogenates was 88.1-93.3%. Correlation with the mouse bioassay yielded a slope of 1.64 and correlation coefficient (r(2)) of 0.84, while correlation with the precolumn oxidation HPLC method yielded a slope of 1.20 and an r(2) of 0.92. When samples were sorted according to increasing toxin concentration (microg STX diHCl equiv./kg) as assessed by the mouse bioassay, the RBA returned no false negatives relative to the 800 microg STX diHCl equiv./kg regulatory limit for shellfish. Currently, no validated methods other than the mouse bioassay directly measure a composite toxic potency for PST in shellfish. The results of this interlaboratory study demonstrate that the RBA is suitable for the routine determination of PST in shellfish in appropriately equipped laboratories.

SafePort Proposal - Henry Laboratory 2010

DTIC Science & Technology

2013-11-14

with the high bonding tempature of glass and the melting point of the metals we used . As a result, we focused more on studies done comparing PDMS, PMMA...strength samples. A method was developed for removing the majority of the interfering high concentration species using solid phase extraction. Using the...These polymers were chosen because they are the most common polymers used in microfluidics and can be manufactured via a wide range of methods

Laboratory procedures to generate viral metagenomes.

PubMed

Thurber, Rebecca V; Haynes, Matthew; Breitbart, Mya; Wegley, Linda; Rohwer, Forest

2009-01-01

This collection of laboratory protocols describes the steps to collect viruses from various samples with the specific aim of generating viral metagenome sequence libraries (viromes). Viral metagenomics, the study of uncultured viral nucleic acid sequences from different biomes, relies on several concentration, purification, extraction, sequencing and heuristic bioinformatic methods. No single technique can provide an all-inclusive approach, and therefore the protocols presented here will be discussed in terms of hypothetical projects. However, care must be taken to individualize each step depending on the source and type of viral-particles. This protocol is a description of the processes we have successfully used to: (i) concentrate viral particles from various types of samples, (ii) eliminate contaminating cells and free nucleic acids and (iii) extract, amplify and purify viral nucleic acids. Overall, a sample can be processed to isolate viral nucleic acids suitable for high-throughput sequencing in approximately 1 week.

Evaluation of a manual DNA extraction protocol and an isothermal amplification assay for detecting HIV-1 DNA from dried blood spots for use in resource-limited settings.

PubMed

Jordan, Jeanne A; Ibe, Christine O; Moore, Miranda S; Host, Christel; Simon, Gary L

2012-05-01

In resource-limited settings (RLS) dried blood spots (DBS) are collected on infants and transported through provincial laboratories to a central facility where HIV-1 DNA PCR testing is performed using specialized equipment. Implementing a simpler approach not requiring such equipment or skilled personnel could allow the more numerous provincial laboratories to offer testing, improving turn-around-time to identify and treat infected infants sooner. Assess performances of a manual DNA extraction method and helicase-dependent amplification (HDA) assay for detecting HIV-1 DNA from DBS. 60 HIV-1 infected adults were enrolled, blood samples taken and DBS made. DBS extracts were assessed for DNA concentration and beta globin amplification using PCR and melt-curve analysis. These same extracts were then tested for HIV-1 DNA using HDA and compared to results generated by PCR and pyrosequencing. Finally, HDA limit of detection (LOD) studies were performed using DBS extracts prepared with known numbers of 8E5 cells. The manual extraction protocol consistently yielded high concentrations of amplifiable DNA from DBS. LOD assessment demonstrated HDA detected ∼470 copies/ml of HIV-1 DNA extracts in 4/4 replicates. No statistical difference was found using the McNemar's test when comparing HDA to PCR for detecting HIV-1 DNA from DBS. Using just a magnet, heat block and pipettes, the manual extraction protocol and HDA assay detected HIV-1 DNA from DBS at levels that would be useful for early infant diagnosis. Next steps will include assessing HDA for non-B HIV-1 subtypes recognition and comparison to Roche HIV-1 DNA v1.5 PCR assay. Copyright © 2012 Elsevier B.V. All rights reserved.

Allied Health Chemistry Laboratory: Amino Acids, Insulin, Proteins, and Skin

ERIC Educational Resources Information Center

Dever, David F.

1975-01-01

Presents a laboratory experiment specifically designed for allied health students. The students construct molecular models of amino acids, extract amino acids from their skin with hot water, and chromatographically analyze the skin extract and hydrolyzed insulin. (MLH)

Determination of Ethyl Carbamate in Chinese Yellow Rice Wine by Diatomaceous Earth Extraction and GC/MS Method.

PubMed

Wu, Pinggu; Zhang, Liqun; Shen, Xianghong; Wang, Liyuan; Zou, Yan; Zhang, Jing; Tan, Ying; Tang, Jun; Ma, Bingjie; Pan, Xiaodong; Jiang, Wei

2015-01-01

A sensitive and rapid analytical method based on alkaline diatomaceous earth extraction followed by GC/MS was developed for the quantitative determination of the toxic contaminant ethyl carbamate (EC) in yellow rice wines. The optimal extraction conditions were investigated. With the application of diatomaceous earth extraction, the damage of organic acids to the capillary column was greatly reduced. By using d5-EC as an internal standard for quantitative analysis of EC, the linearity of the calibration curves was good between 10 and 1000 ng/mL. The LOD and LOQ were 1.7 and 5.0 μg/kg, respectively. The spiked level of EC was 5.0-300 μg/kg, and the average recovery of the spikes was between 78.4 and 98.2%, with an RSD between 4.3 and 8.3%. Upon validation by five laboratories when spiked with 50, 100, and 300 μg/kg, the average respective recoveries were 102.9, 102.2, and 98.7% with a RSD between 0.7 and 8.1%. The validation results demonstrated that the method is fast, simple, selective, and suitable for the determination of EC in yellow rice wines.

Determination of steroid hormones and related compounds in filtered and unfiltered water by solid-phase extraction, derivatization, and gas chromatography with tandem mass spectrometry

USGS Publications Warehouse

Foreman, William T.; Gray, James L.; ReVello, Rhiannon C.; Lindley, Chris E.; Losche, Scott A.; Barber, Larry B.

2012-01-01

A new analytical method has been developed and implemented at the U.S. Geological Survey National Water Quality Laboratory that determines a suite of 20 steroid hormones and related compounds in filtered water (using laboratory schedule 2434) and in unfiltered water (using laboratory schedule 4434). This report documents the procedures and initial performance data for the method and provides guidance on application of the method and considerations of data quality in relation to data interpretation. The analytical method determines 6 natural and 3 synthetic estrogen compounds, 6 natural androgens, 1 natural and 1 synthetic progestin compound, and 2 sterols: cholesterol and 3--coprostanol. These two sterols have limited biological activity but typically are abundant in wastewater effluents and serve as useful tracers. Bisphenol A, an industrial chemical used primarily to produce polycarbonate plastic and epoxy resins and that has been shown to have estrogenic activity, also is determined by the method. A technique referred to as isotope-dilution quantification is used to improve quantitative accuracy by accounting for sample-specific procedural losses in the determined analyte concentration. Briefly, deuterium- or carbon-13-labeled isotope-dilution standards (IDSs), all of which are direct or chemically similar isotopic analogs of the method analytes, are added to all environmental and quality-control and quality-assurance samples before extraction. Method analytes and IDS compounds are isolated from filtered or unfiltered water by solid-phase extraction onto an octadecylsilyl disk, overlain with a graded glass-fiber filter to facilitate extraction of unfiltered sample matrices. The disks are eluted with methanol, and the extract is evaporated to dryness, reconstituted in solvent, passed through a Florisil solid-phase extraction column to remove polar organic interferences, and again evaporated to dryness in a reaction vial. The method compounds are reacted with activated -methyl--trimethylsilyl trifluoroacetamide at 65 degrees Celsius for 1 hour to form trimethylsilyl or trimethylsilyl-enol ether derivatives that are more amenable to gas chromatographic separation than the underivatized compounds. Analysis is carried out by gas chromatography with tandem mass spectrometry using calibration standards that are derivatized concurrently with the sample extracts. Analyte concentrations are quantified relative to specific IDS compounds in the sample, which directly compensate for procedural losses (incomplete recovery) in the determined and reported analyte concentrations. Thus, reported analyte concentrations (or analyte recoveries for spiked samples) are corrected based on recovery of the corresponding IDS compound during the quantification process. Recovery for each IDS compound is reported for each sample and represents an absolute recovery in a manner comparable to surrogate recoveries for other organic methods used by the National Water Quality Laboratory. Thus, IDS recoveries provide a useful tool for evaluating sample-specific analytical performance from an absolute mass recovery standpoint. IDS absolute recovery will differ and typically be lower than the corresponding analyte’s method recovery in spiked samples. However, additional correction of reported analyte concentrations is unnecessary and inappropriate because the analyte concentration (or recovery) already is compensated for by the isotope-dilution quantification procedure. Method analytes were spiked at 10 and 100 nanograms per liter (ng/L) for most analytes (10 times greater spike levels were used for bisphenol A and 100 times greater spike levels were used for 3--coprostanol and cholesterol) into the following validation-sample matrices: reagent water, wastewater-affected surface water, a secondary-treated wastewater effluent, and a primary (no biological treatment) wastewater effluent. Overall method recovery for all analytes in these matrices averaged 100 percent, with overall relative standard deviation of 28 percent. Mean recoveries of the 20 individual analytes for spiked reagent-water samples prepared along with field samples and analyzed in 2009–2010 ranged from 84–104 percent, with relative standard deviations of 6–36 percent. Concentrations for two analytes, equilin and progesterone, are reported as estimated because these analytes had excessive bias or variability, or both. Additional database coding is applied to other reported analyte data as needed, based on sample-specific IDS recovery performance. Detection levels were derived statistically by fortifying reagent water at six different levels (0.1 to 4 ng/L) and range from about 0.4 to 4 ng/L for 16 analytes. Interim reporting levels applied to analytes in this report range from 0.8 to 8 ng/L. Bisphenol A and the sterols (cholesterol and 3-beta-coprostanol) were consistently detected in laboratory and field blanks. The minimum reporting levels were set at 100 ng/L for bisphenol A and at 200 ng/L for the two sterols to prevent any bias associated with the presence of these compounds in the blanks. A minimum reporting level of 2 ng/L was set for 11-ketotestosterone to minimize false positive risk from an interfering siloxane compound emanating as chromatographic-column bleed, from vial septum material, or from other sources at no more than 1 ng/L.

One-day rate measurements for estimating net nitrification potential in humid forest soils

USGS Publications Warehouse

Ross, D.S.; Fredriksen, G.; Jamison, A.E.; Wemple, B.C.; Bailey, S.W.; Shanley, J.B.; Lawrence, G.B.

2006-01-01

Measurements of net nitrification rates in forest soils have usually been performed by extended sample incubation (2-8 weeks), either in the field or in the lab. Because of disturbance effects, these measurements are only estimates of nitrification potential and shorter incubations may suffice. In three separate studies of northeastern USA forest soil surface horizons, we found that laboratory nitrification rates measured over 1 day related well to those measured over 4 weeks. Soil samples of Oa or A horizons were mixed by hand and the initial extraction of subsamples, using 2 mol L-1 KCl, occurred in the field as soon as feasible after sampling. Soils were kept near field temperature and subsampled again the following day in the laboratory. Rates measured by this method were about three times higher than the 4-week rates. Variability in measured rates was similar over either incubation period. Because NO3- concentrations were usually quite low in the field, average rates from 10 research watersheds could be estimated with only a single, 1-day extraction. Methodological studies showed that the concentration of NH4+ increased slowly during contact time with the KCl extractant and, thus, this contact time should be kept similar during the procedure. This method allows a large number of samples to be rapidly assessed. ?? 2006 Elsevier B.V. All rights reserved.

A New Method and Mass-Spectrometric Instrument for Extraterrestrial Microbial Life Detection Using the Elemental Composition Analyses of Martian Regolith and Permafrost/Ice

NASA Astrophysics Data System (ADS)

Managadze, G. G.; Safronova, A. A.; Luchnikov, K. A.; Vorobyova, E. A.; Duxbury, N. S.; Wurz, P.; Managadze, N. G.; Chumikov, A. E.; Khamizov, R. Kh.

2017-05-01

We propose a new technique for the detection of microorganisms by elemental composition analyses of a sample extracted from regolith, permafrost, and ice of extraterrestrial bodies. We also describe the design of the ABIMAS instrument, which consists of the onboard time-of-flight laser mass-reflectron (TOF LMR) and the sample preparation unit (SPU) for biomass extraction. This instrument was initially approved to fly on board the ExoMars 2020 lander mission. The instrument can be used to analyze the elemental composition of possible extraterrestrial microbial communities and compare it to that of terrestrial microorganisms. We have conducted numerous laboratory studies to confirm the possibility of biomass identification via the following biomarkers: P/S and Ca/K ratios, and C and N abundances. We underline that only the combination of these factors will allow one to discriminate microbial samples from geological ones. Our technique has been tested experimentally in numerous laboratory trials on cultures of microorganisms and polar permafrost samples as terrestrial analogues for martian polar soils. We discuss various methods of extracting microorganisms and sample preparation. The developed technique can be used to search for and identify microorganisms in different martian samples and in the subsurface of other planets, satellites, comets, and asteroids—in particular, Europa, Ganymede, and Enceladus.

Impact of pH on the Stability and the Cross-Reactivity of Ochratoxin A and Citrinin

PubMed Central

Bazin, Ingrid; Faucet-Marquis, Virginie; Monje, Marie-Carmen; El Khoury, Micheline; Marty, Jean-Louis; Pfohl-Leszkowicz, Annie

2013-01-01

Mycotoxins are secondary metabolites produced by several fungi contaminating crops. In several countries, the maximum permitted levels of mycotoxins are found in foodstuffs and feedstuffs. The common strategy of mycotoxin analysis involves extraction, clean-up and quantification by chromatography. In this paper, we analyzed the reasons of underestimation of ochratoxin A (OTA) content in wine, and overestimation of OTA in wheat, depending on the pH of the clean-up step and the simultaneous presence of citrinin (CIT). We demonstrated that the increase of pH by adding polyethylene glycol (PEG) to wine led to an underestimation of OTA by conversion of OTA into open ring ochratoxin A OP-OA. In comparing three methods of extraction and clean-up for the determination of OTA and CIT in wheat—(i) an inter-laboratory validated method for OTA in cereals using immunoaffinity column clean-up (IAC) and extraction by acetonitrile/water; (ii) a validated method using IAC and extraction with 1% bicarbonate Na; and (iii) an in-house validated method based on acid liquid/liquid extraction—we observed an overestimation of OTA after immunoaffinity clean-up when CIT is also present in the sample, whereas an underestimation was observed when OTA was alone. Under neutral and alkaline conditions, CIT was partially recognized by OTA antibodies. PMID:24287570

The room temperature preservation of filtered environmental DNA samples and assimilation into a phenol-chloroform-isoamyl alcohol DNA extraction.

PubMed

Renshaw, Mark A; Olds, Brett P; Jerde, Christopher L; McVeigh, Margaret M; Lodge, David M

2015-01-01

Current research targeting filtered macrobial environmental DNA (eDNA) often relies upon cold ambient temperatures at various stages, including the transport of water samples from the field to the laboratory and the storage of water and/or filtered samples in the laboratory. This poses practical limitations for field collections in locations where refrigeration and frozen storage is difficult or where samples must be transported long distances for further processing and screening. This study demonstrates the successful preservation of eDNA at room temperature (20 °C) in two lysis buffers, CTAB and Longmire's, over a 2-week period of time. Moreover, the preserved eDNA samples were seamlessly integrated into a phenol-chloroform-isoamyl alcohol (PCI) DNA extraction protocol. The successful application of the eDNA extraction to multiple filter membrane types suggests the methods evaluated here may be broadly applied in future eDNA research. Our results also suggest that for many kinds of studies recently reported on macrobial eDNA, detection probabilities could have been increased, and at a lower cost, by utilizing the Longmire's preservation buffer with a PCI DNA extraction. © 2014 The Authors. Molecular Ecology Resources Published by John Wiley & Sons Ltd.

TRANSURANIC STUDIES STATUS AND PROBLEM STATEMENT

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

Leuze, R E

1959-04-29

The purpose of the Transuranics Program is to develop separation processes for the transuranic elements, primarily those produced by long-term neutron irradiation of Pu/sup 239/. The program includes laboratory process development, pilot-plant process testing, processing of 10 kg of Pu/sup 239/ irradiated to greater than 99% burn-up for plutonium and americium-curium recovery, and processing the reirradiated plutonium and americium-curium fractions. The proposed method for processing highly irradiated plutonium is: (1) plutonium-aluminum alloy dissolution in HNO/sub 3/; (2) plutonium recovery by TBP extraction; (3) americium, curium, and rare-earth extraction by TBP from neutral nitrate solution; (4) partial rare-earth removal (primarily lanthanum)more » by americium-curium extraction into 100% TBP from 15M HNO/sub 3/; (5) additional rare-earth removal by extraction in 0.48M mono-2-ethylhexylphosphoric acid from 12M HCl; and (6) americium-curium purification by chloride anion exchange. Processing through the 100% TBP, 15M HNO/sub 3/ cycle can be carried out in the Power Reactor Fuel Reprocessing Pilot Plant. New facilities are proposed 15M HNO/ sub 3/ cycle can be carried out in the Power Reactor Fuel Reprocessing Pilot Plant. New facilities are proposed for laboratory process development studies and the final processing of the transplutonic elements. (auth)« less

A sensitive method for the determination of Sulfonamides in seawater samples by Solid Phase Extraction and UV-Visible spectrophotometry

NASA Astrophysics Data System (ADS)

Errayess, Sophia Ait; Lahcen, Abdellatif Ait; Idrissi, Laila; Marcoaldi, Caterina; Chiavarini, Salvatore; Amine, Aziz

2017-06-01

The authors have developed a sensitive spectrophotometric method for determination of sulfonamide derivatives such as sulfanilamide (SAA), sulfadiazine (SDZ), sulfacetamide (SCT) sulfamethoxazole (SMX), sulfamerazine (SMR), sulfadimethoxine (SDX), sulfamethiazole (SMT) and Sulfathiazole (STZ). This method is based on the Bratton-Marshall reaction, which involves the diazotization of sulfonamides with sodium nitrite under acidic conditions, followed by coupling with N-(1-naphtyl) ethylenediamine dihydrochloride (NED) to form a pink colored compound. Therefore, the Bratton-Marshall method was modified by optimizing the reaction conditions, which allows us to determine a low concentration range of sulfonamides compared to the reported methods. The limits of detection and quantification obtained were 0.019-0.05 and 0.06-0.16 μg mL- 1, respectively. In comparison with other reported methods using different coupling agents, the proposed method was found to be the most simple and sensitive for sulfonamides determination. In this paper, the modified method was successfully employed for the determination of sulfonamides in drinking water, seawater and pharmaceutical and veterinary formulations. The purpose of this work is to optimize and develop a simple method for extraction and concentration of sulfonamides present as residues in seawater and their quantification with the recommended spectrophotometric method. Solid phase extraction (SPE) of sulfonamides from seawater samples was evaluated using Oasis HLB cartridges (3 mL, 540 mg). The recovery efficiency was investigated in the sulfonamides concentration range comprised between 0.19 and 126 ng mL- 1. The ease of use of this extraction method makes it very useful for routine laboratory work.

European validation of a real-time PCR-based method for detection of Listeria monocytogenes in soft cheese.

PubMed

Gianfranceschi, Monica Virginia; Rodriguez-Lazaro, David; Hernandez, Marta; González-García, Patricia; Comin, Damiano; Gattuso, Antonietta; Delibato, Elisabetta; Sonnessa, Michele; Pasquali, Frederique; Prencipe, Vincenza; Sreter-Lancz, Zuzsanna; Saiz-Abajo, María-José; Pérez-De-Juan, Javier; Butrón, Javier; Kozačinski, Lidija; Tomic, Danijela Horvatek; Zdolec, Nevijo; Johannessen, Gro S; Jakočiūnė, Džiuginta; Olsen, John Elmerdahl; De Santis, Paola; Lovari, Sarah; Bertasi, Barbara; Pavoni, Enrico; Paiusco, Antonella; De Cesare, Alessandra; Manfreda, Gerardo; De Medici, Dario

2014-08-01

The classical microbiological method for detection of Listeria monocytogenes requires around 7 days for final confirmation, and due to perishable nature of RTE food products, there is a clear need for an alternative methodology for detection of this pathogen. This study presents an international (at European level) ISO 16140-based validation trial of a non-proprietary real-time PCR-based methodology that can generate final results in the following day of the analysis. This methodology is based on an ISO compatible enrichment coupled to a bacterial DNA extraction and a consolidated real-time PCR assay. Twelve laboratories from six European countries participated in this trial, and soft cheese was selected as food model since it can represent a difficult matrix for the bacterial DNA extraction and real-time PCR amplification. The limit of detection observed was down to 10 CFU per 25 of sample, showing excellent concordance and accordance values between samples and laboratories (>75%). In addition, excellent values were obtained for relative accuracy, specificity and sensitivity (82.75%, 96.70% and 97.62%, respectively) when the results obtained for the real-time PCR-based methods were compared to those of the ISO 11290-1 standard method. An interesting observation was that the L. monocytogenes detection by the real-time PCR method was less affected in the presence of Listeria innocua in the contaminated samples, proving therefore to be more reliable than the reference method. The results of this international trial demonstrate that the evaluated real-time PCR-based method represents an excellent alterative to the ISO standard since it shows a higher performance as well as reduce the extent of the analytical process, and can be easily implemented routinely by the competent authorities and food industry laboratories. Copyright © 2014 Elsevier B.V. All rights reserved.

Evaluation of DNA extraction methods and their clinical application for direct detection of causative bacteria in continuous ambulatory peritoneal dialysis culture fluids from patients with peritonitis by using broad-range PCR.

PubMed

Kim, Si Hyun; Jeong, Haeng Soon; Kim, Yeong Hoon; Song, Sae Am; Lee, Ja Young; Oh, Seung Hwan; Kim, Hye Ran; Lee, Jeong Nyeo; Kho, Weon-Gyu; Shin, Jeong Hwan

2012-03-01

The aims of this study were to compare several DNA extraction methods and 16S rDNA primers and to evaluate the clinical utility of broad-range PCR in continuous ambulatory peritoneal dialysis (CAPD) culture fluids. Six type strains were used as model organisms in dilutions from 10(8) to 10(0) colony-forming units (CFU)/mL for the evaluation of 5 DNA extraction methods and 5 PCR primer pairs. Broad-range PCR was applied to 100 CAPD culture fluids, and the results were compared with conventional culture results. There were some differences between the various DNA extraction methods and primer sets with regard to the detection limits. The InstaGene Matrix (Bio-Rad Laboratories, USA) and Exgene Clinic SV kits (GeneAll Biotechnology Co. Ltd, Korea) seem to have higher sensitivities than the others. The results of broad-range PCR were concordant with the results from culture in 97% of all cases (97/100). Two culture-positive cases that were broad-range PCR-negative were identified as Candida albicans, and 1 PCR-positive but culture-negative sample was identified as Bacillus circulans by sequencing. Two samples among 54 broad-range PCR-positive products could not be sequenced. There were differences in the analytical sensitivity of various DNA extraction methods and primers for broad-range PCR. The broad-range PCR assay can be used to detect bacterial pathogens in CAPD culture fluid as a supplement to culture methods.

HPLC Quantification of astaxanthin and canthaxanthin in Salmonidae eggs.

PubMed

Tzanova, Milena; Argirova, Mariana; Atanasov, Vasil

2017-04-01

Astaxanthin and canthaxanthin are naturally occurring antioxidants referred to as xanthophylls. They are used as food additives in fish farms to improve the organoleptic qualities of salmonid products and to prevent reproductive diseases. This study reports the development and single-laboratory validation of a rapid method for quantification of astaxanthin and canthaxanthin in eggs of rainbow trout (Oncorhynchus mykiss) and brook trout (Salvelinus fontinalis М.). An advantage of the proposed method is the perfect combination of selective extraction of the xanthophylls and analysis of the extract by high-performance liquid chromatography and photodiode array detection. The method validation was carried out in terms of linearity, accuracy, precision, recovery and limits of detection and quantification. The method was applied for simultaneous quantification of the two xanthophylls in eggs of rainbow trout and brook trout after their selective extraction. The results show that astaxanthin accumulations in salmonid fish eggs are larger than those of canthaxanthin. As the levels of these two xanthophylls affect fish fertility, this method can be used to improve the nutritional quality and to minimize the occurrence of the M74 syndrome in fish populations. Copyright © 2016 John Wiley & Sons, Ltd.

The numerical simulation on swelling factor and extraction rate of a tight crude oil and SC-CO2 system

NASA Astrophysics Data System (ADS)

Zou, Hongjun; Gong, Houjian; Li, Yajun; Dong, Mingzhe

2018-03-01

A method was established to study swelling and extraction between CO2 and crude oil, and the influences of pressure, temperature and molecular weight were investigated. Firstly, laboratory analysis was conducted to determine the pseudo-component and other parameters of the crude oil. Then swelling and extraction of the crude oil and SC-CO2 system were calculated by computer simulation. The results show that the pressure and temperature have little influence on the swelling and extraction between CO2 and crude oil when the mole fraction of CO2 is lower. A higher pressure and temperature is more beneficial to the interaction of CO2 and crude oil, while the swelling and extraction will not be obvious when the system is miscible. And the smaller the molecular weight of the oil is, the larger the maximum value of the swelling factor of CO2 and crude oil changes. The study of swelling and extraction plays an important role in the oilfield stimulation.

Experiment design for pilot identification in compensatory tracking tasks

NASA Technical Reports Server (NTRS)

Wells, W. R.

1976-01-01

A design criterion for input functions in laboratory tracking tasks resulting in efficient parameter estimation is formulated. The criterion is that the statistical correlations between pairs of parameters be reduced in order to minimize the problem of nonuniqueness in the extraction process. The effectiveness of the method is demonstrated for a lower order dynamic system.

Mold Species in Dust from the International Space Station Identified and Quantified by Mold Specific Quantitative PCR - MCEARD

EPA Science Inventory

Dust was collected over a period of several weeks in 2007 from HEPA filters in the U.S. Laboratory Module of the International Space Station (ISS). The dust was returned on the Space Shuttle Atlantis, mixed, sieved, and the DNA was extracted. Using a DNA-based method called mo...

Mold Species in Dust from the International Space Station Identified and Quantified by Mold Specific Quantitative PCR

EPA Science Inventory

Dust was collected over a period of several weeks in 2007 from HEPA filters in the U.S. Laboratory Module of the International Space Station (ISS). The dust was returned on the Space Shuttle Atlantis, mixed, sieved, and the DNA was extracted. Using a DNA-based method called mol...

Mendel Meets CSI: Forensic Genotyping as a Method to Teach Genetics & DNA Science

ERIC Educational Resources Information Center

Kurowski, Scotia; Reiss, Rebecca

2007-01-01

This article describes a forensic DNA science laboratory exercise for advanced high school and introductory college level biology courses. Students use a commercial genotyping kit and genetic analyzer or gene sequencer to analyze DNA recovered from a fictitious crime scene. DNA profiling and STR genotyping are outlined. DNA extraction, PCR, and…

Occupational exposure to complex mixtures of volatile organic compounds in ambient air: desorption from activated charcoal using accelerated solvent extraction can replace carbon disulfide?

PubMed

Fabrizi, Giovanni; Fioretti, Marzia; Rocca, Lucia Mainero

2013-01-01

A desorption study of 57 volatile organic compounds (VOCs) has been conducted by use of accelerated solvent extraction (ASE) and gas chromatography-mass spectrometry. Different solvents were tested to extract activated charcoal tubes with the objective of replacing carbon disulfide, used in official methods, because of its highly toxic health and environmental effects. Extraction conditions, for example temperature and number of cycles, were investigated and optimized. The definitive extraction procedure selected was use of acetone at 150 °C and two consecutive extraction cycles at a pressure of 1,500 psi. Considering a sample volume of 0.005 Nm(3), corresponding to a sampling time of 8 h at a flow rate of 0.01 L min(-1), the method was validated over the concentration range 65-26,300 μg Nm(-3). The lowest limit of quantification was 6 μg Nm(-3), and recovery for the 93 % of analytes ranged from 65 to 102 %. For most of the compounds, relative standard deviations were less than 15 % for inter and intra-day precision. Uncertainty of measurement was also determined: the relative expanded uncertainty was always below 29.6 %, except for dichlorodifluoromethane. This work shows that use of friendlier solvent, for example acetone, coupled with use of ASE, can replace use of CS(2) for chemical removal of VOCs from activated charcoal. ASE has several advantages over traditional solvent-extraction methods, including shorter extraction time, minimum sample manipulation, high reproducibility, and less extraction discrimination. No loss of sensitivity occurs and there is also a salutary effect on bench workers' health and on the smell of laboratory air.

Analysis of acrylamide in coffee and cocoa by isotope dilution liquid chromatography-tandem mass spectrometry.

PubMed

Aguas, Patricia C; Fitzhenry, Matthew J; Giannikopoulos, Georgina; Varelis, Peter

2006-08-01

An accurate and precise method for the quantification of acrylamide using stable isotope dilution liquid chromatography-tandem mass spectrometry was developed and used to measure acrylamide in coffee and cocoa samples. The sample preparation involved extraction of the analyte and its internal standard, 13C3-acrylamide, into water and subsequent defatting of the aqueous extract with dichloromethane. An aliquot of the resulting aqueous extract was then azeotropically dried under reduced pressure and subsequently purified using an aminopropyl-bonded silica cartridge. The purified extracts were then chromatographed on a 5-microm 2.1 x 150 mm Hypercarb column, the effluent of which was monitored for the analyte and its internal standard using positive-ion APCI-selected reaction monitoring. The intra-laboratory reproducibility of the method, expressed as a relative coefficient of variation (%, n=5), was determined at four levels of concentration (12.3, 42.3, 139.3 and 464.8 microg kg(-1)) and was found to vary between 0.6-2.5%. The accuracy of the method was assessed using a reference sample of coffee. The average result obtained using our method differed from the assigned value of the reference material by less than 1%. An analysis of a cocoa sample revealed that the method is capable of precisely estimating acrylamide in challenging matrices down to a level of at least 12.3 microg kg(-1).

Impact of Processing Method on Recovery of Bacteria from Wipes Used in Biological Surface Sampling

PubMed Central

Olson, Nathan D.; Filliben, James J.; Morrow, Jayne B.

2012-01-01

Environmental sampling for microbiological contaminants is a key component of hygiene monitoring and risk characterization practices utilized across diverse fields of application. However, confidence in surface sampling results, both in the field and in controlled laboratory studies, has been undermined by large variation in sampling performance results. Sources of variation include controlled parameters, such as sampling materials and processing methods, which often differ among studies, as well as random and systematic errors; however, the relative contributions of these factors remain unclear. The objective of this study was to determine the relative impacts of sample processing methods, including extraction solution and physical dissociation method (vortexing and sonication), on recovery of Gram-positive (Bacillus cereus) and Gram-negative (Burkholderia thailandensis and Escherichia coli) bacteria from directly inoculated wipes. This work showed that target organism had the largest impact on extraction efficiency and recovery precision, as measured by traditional colony counts. The physical dissociation method (PDM) had negligible impact, while the effect of the extraction solution was organism dependent. Overall, however, extraction of organisms from wipes using phosphate-buffered saline with 0.04% Tween 80 (PBST) resulted in the highest mean recovery across all three organisms. The results from this study contribute to a better understanding of the factors that influence sampling performance, which is critical to the development of efficient and reliable sampling methodologies relevant to public health and biodefense. PMID:22706055

Conversion and Extraction of Insoluble Organic Materials in Meteorites

NASA Technical Reports Server (NTRS)

Locke, Darren R.; Burton, Aaron S.; Niles, Paul B.

2016-01-01

We endeavor to develop and implement methods in our laboratory to convert and extract insoluble organic materials (IOM) from low car-bon bearing meteorites (such as ordinary chondrites) and Precambrian terrestrial rocks for the purpose of determining IOM structure and prebiotic chemistries preserved in these types of samples. The general scheme of converting and extracting IOM in samples is summarized in Figure 1. First, powdered samples are solvent extracted in a micro-Soxhlet apparatus multiple times using solvents ranging from non-polar to polar (hexane - non-polar, dichloromethane - non-polar to polar, methanol - polar protic, and acetonitrile - polar aprotic). Second, solid residue from solvent extractions is processed using strong acids, hydrochloric and hydrofluoric, to dissolve minerals and isolate IOM. Third, the isolated IOM is subjected to both thermal (pyrolysis) and chemical (oxidation) degradation to release compounds from the macromolecular material. Finally, products from oxidation and pyrolysis are analyzed by gas chromatography - mass spectrometry (GCMS). We are working toward an integrated method and analysis scheme that will allow us to determine prebiotic chemistries in ordinary chondrites and Precambrian terrestrial rocks. Powerful techniques that we are including are stepwise, flash, and gradual pyrolysis and ruthenium tetroxide oxidation. More details of the integrated scheme will be presented.

Effect of leaf extract buasbuas (Premna pubescens Blume) for against of bacteria growth Staphylococcus aureus and Salmonella sp in vitro

NASA Astrophysics Data System (ADS)

Restuati, Martina; Pratiwi, Nanda; Widiyastuti, Gita

2017-08-01

Buasbuas (Premna pubescens Blume) is the one of the medicinal plants in Indonesia. This plant contains flavonoids potential as an antibacterial. Staphylococcus aureus and Salmonella sp is the one of the bacteria that can cause poisoning. This research aims to find out of inhibitory the extract of buasbuas leaves the growth of Staphylococcus aureus and Salmonella sp. This research was conducted in Medan Health Laboratory used hole sinks methods. Buasbuas leaves which has been condensed ethanol extract was diluted using aquadest with a concentration of 0%, 50%, 60%, 70%, 80%, 90% and dropped into MSA (Mannitol Salt Agar) media inoculated bacteria and wich is a hole sinks. Them incubated for 1 × 24 hours at a temperature of 37°C. The results of this research that the buasbuas extract with the different concentrations can inhibit the growth of Staphylococcus aureus, then the result were analyzed used of Non Factorial Experimental Method (ANOVA) where Fcount (337.54)> Ftable 0,05 (2,77); Ftabel 0.01 (4.25). While the maximum concentration buasbuas extract were more effective the growh of bacteria is 90% with average inhibition zone diameter of 11 mm.

Harvesting (67)Cu from the Collection of a Secondary Beam Cocktail at the National Superconducting Cyclotron Laboratory.

PubMed

Mastren, Tara; Pen, Aranh; Loveless, Shaun; Marquez, Bernadette V; Bollinger, Elizabeth; Marois, Boone; Hubley, Nicholas; Brown, Kyle; Morrissey, David J; Peaslee, Graham F; Lapi, Suzanne E

2015-10-20

Isotope harvesting is a promising new method to obtain isotopes for which there is no reliable continuous supply at present. To determine the possibility of obtaining radiochemically pure radioisotopes from an aqueous beam dump at a heavy-ion fragmentation facility, preliminary experiments were performed to chemically extract a copper isotope from a large mixture of projectile fragmentation products in an aqueous medium. In this work a 93 MeV/u secondary beam cocktail was collected in an aqueous beam stop at the National Superconducting Cyclotron Laboratory (NSCL) located on the Michigan State University (MSU) campus. The beam cocktail consisted of ∼2.9% (67)Cu in a large mixture of co-produced isotopes ranging in atomic number from ∼19 to 34. The chemical extraction of (67)Cu was achieved via a two-step process: primary extraction using a divalent metal chelation disk followed by anion-exchange chromatography. A significant fraction (74 ± 4%) of the (67)Cu collected in the aqueous beam stop was recovered with >99% radiochemical purity. To illustrate the utility of this product, the purified (67)Cu material was then used to radiolabel an anti-EGFR antibody, Panitumumab, and injected into mice bearing colon cancer xenografts. The tumor uptake at 5 days postinjection was found to be 12.5 ± 0.7% which was in very good agreement with previously reported studies with this radiolabeled antibody. The present results demonstrate that harvesting isotopes from a heavy-ion fragmentation facility could be a promising new method for obtaining high-quality isotopes that are not currently available by traditional methods.

The detection of NBOMe designer drugs on blotter paper by high resolution time-of-flight mass spectrometry (TOFMS) with and without chromatography.

PubMed

Botch-Jones, Sabra; Foss, Jamie; Barajas, David; Kero, Frank; Young, Craig; Weisenseel, Jason

2016-10-01

New psychoactive substances (NPS) have been associated with fatalities and severe injuries in a number of cases in the United States and have led investigators to rethink traditional drug monitoring protocols. Of particular interest are the variable phenethylamine chemical structures known as 'NBOMes', which pose an emerging threat to public health with incidence steadily growing over the past decade. In this study, direct sample analysis (DSA)-time of flight mass spectrometry was employed to leverage rapid and sensitive ambient ionization mass spectrometry without chromatographic separation as verified with an authentic case sample. Samples for method development were prepared at Boston University School of Medicine's Biomedical Forensic Sciences program (Boston, MA) and analyzed at the State of Maine Health and Environmental Testing Laboratory's Forensic Chemistry Section (Augusta, ME). Preliminary method development work was performed at the University of Central Florida (Orlando, FL). DSA without any extraction step in addition to the evaluation of methanol, dichloromethane and hexane extractions were conducted. Methanol was found to not be a suitable extraction solvent for DSA analysis of these compounds. For the screening of NBOMe designer drug variables on blotter paper, DSA-TOFMS was successful at reducing analysis time to ∼15s per sample, for qualitative identification for the selected analytes of interest. The analysis of an authentic forensic case sample by DSA-TOFMS using the method development parameters demonstrates its utility in forensic laboratories. 25C-NBOMe was identified with an exact mass accuracy of 0.60ppm. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Rapid simultaneous determination of amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine, and 3,4-methylenedioxyethylamphetamine in urine by solid-phase extraction and GC-MS: a method optimized for high-volume laboratories.

PubMed

Stout, Peter R; Horn, Carl K; Klette, Kevin L

2002-01-01

To facilitate analysis of high sample volumes, an extraction, derivatization and gas chromatographic-mass spectrometric analysis method was developed to simultaneously determine amphetamine (AMP), methamphetamine (MAMP), 3,4-methylenedioxyamphetamine (MDA) 3,4-methylenedioxymethamphetamine (MDMA), and 3,4-methylenedioxyethylamphetamine (MDEA) in urine. This method utilized a positive-pressure manifold cation-exchange polymer-based solid-phase extraction followed by elution directly into automated liquid sampler (ALS) vials. Rapid derivatization was accomplished using heptafluorobutyric anhydride (HFBA). Recoveries averaged 90% or greater for each of the compounds. Limits of detection were 62.5 ng/mL (AMP and MDEA), 15.6 ng/mL (MAMP), and 31.3 ng/mL (MDA and MDMA) using a 2-mL sample volume. The method was linear to 5000 ng/mL for all compounds using MDMA-d5 and MAMP-d14 as internal standards. Over 200 human urine samples previously determined to contain the target analytes were analyzed using the method. Excellent agreement was seen with previous quantitations. The method was challenged with 75 potentially interfering compounds and no interferences were seen. These interfering compounds included ephedrine, pseudoephedrine, phenylpropanolamine, and phenethylamine. The method resulted in dramatic reductions in processing time and waste production.

SANDIA NATIONAL LABORATORIES IN SITU ELECTROKINETIC EXTRACTION TECHNOLOGY; INNOVATIVE TECHNOLOGY EVALUATION REPORT

EPA Science Inventory

As a part of the Superfund Innovative Technology Evaluation (SITE) Program, the U.S. Environmental Protection Agency evaluated the In-Situ Electrokinetic Extraction (ISEE) system at Sandia National Laboratories, Albuquerque, New Mexico.

The SITE demonstration results show ...

Modified salting-out method: high-yield, high-quality genomic DNA extraction from whole blood using laundry detergent.

PubMed

Nasiri, H; Forouzandeh, M; Rasaee, M J; Rahbarizadeh, F

2005-01-01

Different approaches have been used to extract DNA from whole blood. In most of these methods enzymes (such as proteinase K and RNAse A) or toxic organic solvents (such as phenol or guanidine isothiocyanate) are used. Since these enzymes are expensive, and most of the materials that are used routinely are toxic, it is desirable to apply an efficient DNA extraction procedure that does not require the use of such materials. In this study, genomic DNA was extracted by the salting-out method, but instead of using an analytical-grade enzyme and chemical detergents, as normally used for DNA isolation, a common laundry powder was used. Different concentrations of the powder were tested, and proteins were precipitated by NaCl-saturated distilled water. Finally, DNA precipitation was performed with the use of 96% ethanol. From the results, we conclude that the optimum concentration of laundry powder for the highest yield and purity of isolated DNA is 30 mg/mL. The procedure was optimized, and a final protocol is suggested. Following the same protocol, DNA was extracted from 100 blood samples, and their amounts were found to be >50 microg/mL of whole blood. The integrity of the DNA fragments was confirmed by agarose gel electrophoresis. Furthermore, the extracted DNA was used as a template for PCR reaction. The results obtained from PCR showed that the final solutions of extracted DNA did not contain any inhibitory material for the enzyme used in the PCR reaction, and indicated that the isolated DNA was of good quality. These results show that this method is simple, fast, safe, and cost-effective, and can be used in medical laboratories and research centers. Copyright 2005 Wiley-Liss, Inc.

Validation of a high-throughput real-time polymerase chain reaction assay for the detection of capripoxviral DNA.

PubMed

Stubbs, Samuel; Oura, Chris A L; Henstock, Mark; Bowden, Timothy R; King, Donald P; Tuppurainen, Eeva S M

2012-02-01

Capripoxviruses, which are endemic in much of Africa and Asia, are the aetiological agents of economically devastating poxviral diseases in cattle, sheep and goats. The aim of this study was to validate a high-throughput real-time PCR assay for routine diagnostic use in a capripoxvirus reference laboratory. The performance of two previously published real-time PCR methods were compared using commercially available reagents including the amplification kits recommended in the original publication. Furthermore, both manual and robotic extraction methods used to prepare template nucleic acid were evaluated using samples collected from experimentally infected animals. The optimised assay had an analytical sensitivity of at least 63 target DNA copies per reaction, displayed a greater diagnostic sensitivity compared to conventional gel-based PCR, detected capripoxviruses isolated from outbreaks around the world and did not amplify DNA from related viruses in the genera Orthopoxvirus or Parapoxvirus. The high-throughput robotic DNA extraction procedure did not adversely affect the sensitivity of the assay compared to manual preparation of PCR templates. This laboratory-based assay provides a rapid and robust method to detect capripoxviruses following suspicion of disease in endemic or disease-free countries. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

High-throughput DNA extraction of forensic adhesive tapes.

PubMed

Forsberg, Christina; Jansson, Linda; Ansell, Ricky; Hedman, Johannes

2016-09-01

Tape-lifting has since its introduction in the early 2000's become a well-established sampling method in forensic DNA analysis. Sampling is quick and straightforward while the following DNA extraction is more challenging due to the "stickiness", rigidity and size of the tape. We have developed, validated and implemented a simple and efficient direct lysis DNA extraction protocol for adhesive tapes that requires limited manual labour. The method uses Chelex beads and is applied with SceneSafe FAST tape. This direct lysis protocol provided higher mean DNA yields than PrepFiler Express BTA on Automate Express, although the differences were not significant when using clothes worn in a controlled fashion as reference material (p=0.13 and p=0.34 for T-shirts and button-down shirts, respectively). Through in-house validation we show that the method is fit-for-purpose for application in casework, as it provides high DNA yields and amplifiability, as well as good reproducibility and DNA extract stability. After implementation in casework, the proportion of extracts with DNA concentrations above 0.01ng/μL increased from 71% to 76%. Apart from providing higher DNA yields compared with the previous method, the introduction of the developed direct lysis protocol also reduced the amount of manual labour by half and doubled the potential throughput for tapes at the laboratory. Generally, simplified manual protocols can serve as a cost-effective alternative to sophisticated automation solutions when the aim is to enable high-throughput DNA extraction of complex crime scene samples. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Determination of Ochratoxin A in Black and White Pepper, Nutmeg, Spice Mix, Cocoa, and Drinking Chocolate by High-Performance Liquid Chromatography Coupled with Fluorescence Detection: Collaborative Study.

PubMed

Cubero-Leon, Elena; Bouten, Katrien; Senyuva, Hamide; Stroka, Joerg

2017-09-01

A method validation study for the determination of ochratoxin A in black and white pepper (Piper spp.), nutmeg (Myristica fragrans), spice mix (blend of ginger, turmeric, pepper, nutmeg, and chili), cocoa powder, and drinking chocolate was conducted according to the International Harmonized Protocol of the International Union of Pure and Applied Chemistry. The method is based on the extraction of samples with aqueous methanol, followed by a cleanup of the extract with an immunoaffinity column. The determination is carried out by reversed-phase LC coupled with a fluorescence detector. The study involved 25 participants representing a cross-section of research, private, and official control laboratories from 12 European Union (EU) Member States, together with Turkey and Macedonia. Mean recoveries ranged from 71 to 85% for spices and from 85 to 88% for cocoa and drinking chocolate. The RSDr values ranged from 5.6 to 16.7% for spices and from 4.5 to 18.7% for cocoa and drinking chocolate. The RSDR values ranged from 9.5 to 22.6% for spices and from 13.7 to 30.7% for cocoa and drinking chocolate. The resulting Horwitz ratios ranged from 0.4 to 1 for spices and from 0.6 to 1.4 for cocoa and drinking chocolate according to the Horwitz function modified by Thompson. The method showed acceptable within-laboratory and between-laboratory precision for each matrix, and it conforms to requirements set by current EU legislation.

MALDI-TOF MS for the identification of veterinary non-C. neoformans-C. gattii Cryptococcus spp. isolates from Italy.

PubMed

Danesi, Patrizia; Drigo, Ilenia; Iatta, Roberta; Firacative, Carolina; Capelli, Gioia; Cafarchia, Claudia; Meyer, Wieland

2014-08-01

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) offers an effective alternative to phenotypic and molecular methods for the rapid identification of microorganisms. Our aim in this study was to create an in-house library for a set of strains of nine uncommonly reported human and animal cryptococcal species, including Cryptococcus adeliensis, C. albidosimilis, C. albidus, C. aureus, C. carnescens, C. laurentii, C. magnus, C. victoriae and C. uniguttulatus, and to use this library to make timely and correct identifications using MALDI-TOF MS for use in routine laboratory diagnostics. Protein extracts obtained via the formic acid extraction method of 62 veterinary non-C. neoformans-C. gattii cryptococcal isolates were studied. The obtained mass spectra correctly grouped all 62 studied isolates according to species identification previously obtained by internal transcribe spacer sequence analysis. The in-house database was than exported and successfully uploaded to the Microflex LT (Maldi Biotyper; Bruker Daltonics) instrument at a different diagnostic laboratory in Italy. Scores >2.7 obtained from isolates reanalyzed in the latter laboratory supported the high reproducibility of the method. The possibility of creating and transferring an in-house library adds to the usefulness MALDI-TOF MS an important tool for the rapid and inexpensive identification of pathogenic and saprophytic fungi as required for differential diagnosis of human and animal mycoses. © The Author 2014. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Feasibility of Isotope Harvesting at a Projectile Fragmentation Facility: 67Cu

PubMed Central

Mastren, Tara; Pen, Aranh; Peaslee, Graham F.; Wozniak, Nick; Loveless, Shaun; Essenmacher, Scott; Sobotka, Lee G.; Morrissey, David J.; Lapi, Suzanne E.

2014-01-01

The work presented here describes a proof-of-principle experiment for the chemical extraction of 67Cu from an aqueous beam stop at the National Superconducting Cyclotron Laboratory (NSCL). A 76 MeV/A 67Cu beam was stopped in water, successfully isolated from the aqueous solution through a series of chemical separations involving a chelating disk and anion exchange chromatography, then bound to NOTA-conjugated Herceptin antibodies, and the bound activity was validated using instant thin-layer chromatography (ITLC). The chemical extraction efficiency was found to be 88 ± 3% and the radiochemical yield was ≥95%. These results show that extraction of radioisotopes from an aqueous projectile-fragment beam dump is a feasible method for obtaining radiochemically pure isotopes. PMID:25330839

A rapid and efficient DNA extraction protocol from fresh and frozen human blood samples.

PubMed

Guha, Pokhraj; Das, Avishek; Dutta, Somit; Chaudhuri, Tapas Kumar

2018-01-01

Different methods available for extraction of human genomic DNA suffer from one or more drawbacks including low yield, compromised quality, cost, time consumption, use of toxic organic solvents, and many more. Herein, we aimed to develop a method to extract DNA from 500 μL of fresh or frozen human blood. Five hundred microliters of fresh and frozen human blood samples were used for standardization of the extraction procedure. Absorbance at 260 and 280 nm, respectively, (A 260 /A 280 ) were estimated to check the quality and quantity of the extracted DNA sample. Qualitative assessment of the extracted DNA was checked by Polymerase Chain reaction and double digestion of the DNA sample. Our protocol resulted in average yield of 22±2.97 μg and 20.5±3.97 μg from 500 μL of fresh and frozen blood, respectively, which were comparable to many reference protocols and kits. Besides yielding bulk amount of DNA, our protocol is rapid, economical, and avoids toxic organic solvents such as Phenol. Due to unaffected quality, the DNA is suitable for downstream applications. The protocol may also be useful for pursuing basic molecular researches in laboratories having limited funds. © 2017 Wiley Periodicals, Inc.

Larvicidal & ovicidal efficacy of Pithecellobium dulce (Roxb.) Benth. (Fabaceae) against Anopheles stephensi Liston & Aedes aegypti Linn. (Diptera: Culicidae)

PubMed Central

Govindarajan, M.; Rajeswary, M.; Sivakumar, R.

2013-01-01

Background & objectives: In view of the recently increased interest in developing plant origin insecticides as an alternative to chemical insecticide, this study was undertaken to assess the larvicidal and ovicidal potential of the crude hexane, benzene, chloroform, ethyl acetate and methanol solvent extracts from the medicinal plant Pithecellobium dulce against the mosquito vectors, Anopheles stephensi and Aedes aegypti (Diptera: Culicidae). Methods: Larvicidal activity of P. dulce plant extracts was studied in the range of 60 to 450 mg/l against early third instar larvae of An. stephensi and Ae. aegypti in the laboratory. The larval mortality was observed after 24 h of exposure. The ovicidal activity was determined against An. stephensi and Ae. aegypti to various concentrations ranging from 100 to 750 mg/l under the laboratory conditions. Mean per cent hatchability of the eggs were observed after 48 h post treatment. Results: All leaf and seed extracts showed moderate larvicidal and ovicidal effects; however, the highest larval mortality was found in methanol extract of leaf of P. dulce against the larvae of An. stephensi and Ae. aegypti with the LC50 and LC90 values 145.43, 155.78 mg/l and 251.23, 279.73 mg/l, respectively. The per cent hatchability was inversely proportional to the concentration of extract and directly proportional to the eggs. Zero hatchability was observed at 400 mg/l for leaf methanol extract and 625 mg/l for seed methanol extract of P. dulce against An. stephensi and Ae. aegypti, respectively. Compared to leaf extracts, seed extracts have low potency against the two mosquitoes. Interpretation & conclusions: The present results suggest that the leaf and seed extracts of P. dulce have the potential to be used as an ideal eco-friendly approach for the control of mosquitoes. PMID:24056567

Development of Singlet Oxygen Absorption Capacity (SOAC) Assay Method Using a Microplate Reader.

PubMed

Takahashi, Shingo; Iwasaki-Kino, Yuko; Aizawa, Koichi; Terao, Junji; Mukai, Kazuo

2016-01-01

Recently, a new assay method that can quantify the singlet oxygen absorption capacity (SOAC) of natural antioxidants and food extracts was developed. The SOAC values were measured in ethanol-chloroform-D2O (50 + 50 + 1, v/v/v) solution at 35°C using a UV-Vis spectrophotometer equipped with a six-channel cell positioner and an electron-temperature control unit. In the present study, measurement of the SOAC values was performed for eight representative carotenoids and three vegetable extracts (tomato, carrot, and red paprika) using a versatile instrument, the microplate reader. A 24-well glass microplate was used for measurements because a plastic microplate, commonly used in the laboratory, dissolves in the ethanol-chloroform-D2O solution. The SOAC values of eight carotenoids and three vegetable extracts measured using a microplate reader were in good agreement with the corresponding values measured using a UV-Vis spectrophotometer, suggesting that the microplate reader is an applicable instrument for the measurement of reliable SOAC values for general antioxidants and food extracts in solution.

Method of analysis at the U.S. Geological Survey California Water Science Center, Sacramento Laboratory - determination of haloacetic acid formation potential, method validation, and quality-control practices

USGS Publications Warehouse

Zazzi, Barbara C.; Crepeau, Kathryn L.; Fram, Miranda S.; Bergamaschi, Brian A.

2005-01-01

An analytical method for the determination of haloacetic acid formation potential of water samples has been developed by the U.S. Geological Survey California Water Science Center Sacramento Laboratory. The haloacetic acid formation potential is measured by dosing water samples with chlorine under specified conditions of pH, temperature, incubation time, darkness, and residual-free chlorine. The haloacetic acids formed are bromochloroacetic acid, bromodichloroacetic acid, dibromochloroacetic acid, dibromoacetic acid, dichloroacetic acid, monobromoacetic acid, monochloroacetic acid, tribromoacetic acid, and trichloroacetic acid. They are extracted, methylated, and then analyzed using a gas chromatograph equipped with an electron capture detector. Method validation experiments were performed to determine the method accuracy, precision, and detection limit for each of the compounds. Method detection limits for these nine haloacetic acids ranged from 0.11 to 0.45 microgram per liter. Quality-control practices include the use of blanks, quality-control samples, calibration verification standards, surrogate recovery, internal standard, matrix spikes, and duplicates.

Extraction process of palm kernel cake as a source of mannan for feed additive on poultry diet

NASA Astrophysics Data System (ADS)

Tafsin, M.; Hanafi, N. D.; Yusraini, E.

2017-05-01

Palm Kernel Cake (PKC) is a by-product of palm kernel oil extraction and found in large quantity in Indonesia. The inclusion of PKC on poultry diet are limited due to some nutritional problems such as anti-nutritional properties (mannan). On the other hand, mannan containing polysaccharides play in various biological functions particularly in enhancing the immune response and to control pathogen in poultry. The research objective to find out the extraction process of PKC and conducted at animal nutrition and Feed Science Laboratory, Agricultural Faculty, University of Sumatera Utara. Various extraction methode were used in this experiment, including fraction analysis used 7 number sieves, and followed by water and acetic acid extraction. The result indicated that PKC had different particle size according to sieve size and dominated by particle size 850 um. The analysis of sugar content indicated that each particle size had different characteristic on treatment by hot water extraction. The particle size 180—850 um had higher sugar content than coarse PKC (2000—3000 um). The total sugar content were recovered vary between 0.9—3,2% from PKC were extracted. Treatment grinding method followed by hot water extraction (100—120 °C, 1 h) increased total sugar content than previous treatments and reach 8% from PKC were extracted. Utilisation acetic acid decreased the total amount of total sugar from PKC were extracted. It is concluded that treatment by hot temperature (110—120 °C) for 1 h show highest yield to extract sugar from PKC.

A fast and sensitive method for the simultaneous analysis of a wide range of per- and polyfluoroalkyl substances in indoor dust using on-line solid phase extraction-ultrahigh performance liquid chromatography-time-of-flight-mass spectrometry.

PubMed

Padilla-Sánchez, Juan Antonio; Haug, Line Småstuen

2016-05-06

A fast and sensitive method for simultaneous determination of 18 traditional and 6 alternative per- and polyfluoroalkyl substances (PFASs) using solid-liquid extraction (SLE), off-line clean-up using activated carbon and on-line solid phase extraction-ultrahigh performance liquid chromatography-time-of-flight-mass spectrometry (on-line SPE-UHPLC-TOF-MS) was developed. The extraction efficiency was studied and recoveries in range the 58-114% were obtained. Extraction and injection volumes were also optimized to 2mL and 400μL, respectively. The method was validated by spiking dust from a vacuum cleaner bag that had been found to contain low levels of the PFASs in focus. Low method detection limits (MDLs) and method quantification limits (MQLs) in the range 0.008-0.846ngg(-1) and 0.027-2.820ngg(-1) were obtained, respectively. For most of the PFASs, the accuracies were between 70 and 125% in the range from 2 to100ngg(-1) dust. Intra-day and inter-day precisions were in general well below 30%. Analysis of a Standard Reference Material (SRM 2585) showed high accordance with results obtained by other laboratories. Finally, the method was applied to seven indoor dust samples, and PFAS concentrations in the range 0.02-132ngg(-1) were found. The highest median concentrations were observed for some of the alternative PFASs, such as 6:2-diPAP (25ngg(-1)), 8:2-diPAP (49ngg(-1)), and PFOPA (23ngg(-1)), illustrating the importance of inclusion of new PFASs in the analytical methods. Copyright © 2016 Elsevier B.V. All rights reserved.

Control of key pecan insect pests using biorational pesticides.

PubMed

Shapiro-Ilan, David I; Cottrell, Ted E; Jackson, Mark A; Wood, Bruce W

2013-02-01

Key pecan insect pests include the black pecan aphid, Melanocallis caryaefoliae (Davis), pecan weevil, Curculio caryae (Horn), and stink bugs (Hemiptera: Pentatomidae). Alternative control tactics are needed for management of these pests in organic and conventional systems. Our objective was to evaluate the potential utility of several alternative insecticides including three plant extract formulations, eucalyptus extract, citrus extract-8.92%, and citrus extract-19.4%, and two microbial insecticides, Chromobacterium subtsugae (Martin et al.) and Isaria fumosorosea (Wize). In the laboratory, eucalyptus extract, citrus extract-8.92%, citrus extract-19.4%, and C. subtsugae caused M. caryaefoliae mortality (mortality was reached approximately 78, 83, and 96%, respectively). In field tests, combined applications of I. fumosorosea with eucalyptus extract were synergistic and caused up to 82% mortality in M. caryaefoliae. In laboratory assays focusing on C. caryae suppression, C. subtsugae reduced feeding and oviposition damage, eucalyptus extract and citrus extract-19.4% were ineffective, and antagonism was observed when citrus extract-19.4% was combined with the entomopathogenic nematode, Steinernema carpocapsae (Weiser). In field tests, C. subtsugae reduced C. caryae damage by 55% within the first 3d, and caused 74.5% corrected mortality within 7 d posttreatment. In the laboratory, C. subtsugae and eucalyptus extract did not cause mortality in the brown stink bug, Euschistus servus (Say). Applications of C. subtsugae for suppression of C. caryae, and eucalyptus extract plus I. fumosorosea for control of M. caryaefoliae show promise as alternative insecticides and should be evaluated further.

COMPARISON OF FIELD AEROBIC BIODEGRADATION RATES TO LABORATORY

EPA Science Inventory

It is common to use bioventing as a polishing step for soil vapor extraction. It was originally planned to use soil vapor extraction and bioventing at a former landfill site in Delaware but laboratory scale biodegradation studies indicated that most of the volatile organic compou...

Automated PCR setup for forensic casework samples using the Normalization Wizard and PCR Setup robotic methods.

PubMed

Greenspoon, S A; Sykes, K L V; Ban, J D; Pollard, A; Baisden, M; Farr, M; Graham, N; Collins, B L; Green, M M; Christenson, C C

2006-12-20

Human genome, pharmaceutical and research laboratories have long enjoyed the application of robotics to performing repetitive laboratory tasks. However, the utilization of robotics in forensic laboratories for processing casework samples is relatively new and poses particular challenges. Since the quantity and quality (a mixture versus a single source sample, the level of degradation, the presence of PCR inhibitors) of the DNA contained within a casework sample is unknown, particular attention must be paid to procedural susceptibility to contamination, as well as DNA yield, especially as it pertains to samples with little biological material. The Virginia Department of Forensic Science (VDFS) has successfully automated forensic casework DNA extraction utilizing the DNA IQ(trade mark) System in conjunction with the Biomek 2000 Automation Workstation. Human DNA quantitation is also performed in a near complete automated fashion utilizing the AluQuant Human DNA Quantitation System and the Biomek 2000 Automation Workstation. Recently, the PCR setup for casework samples has been automated, employing the Biomek 2000 Automation Workstation and Normalization Wizard, Genetic Identity version, which utilizes the quantitation data, imported into the software, to create a customized automated method for DNA dilution, unique to that plate of DNA samples. The PCR Setup software method, used in conjunction with the Normalization Wizard method and written for the Biomek 2000, functions to mix the diluted DNA samples, transfer the PCR master mix, and transfer the diluted DNA samples to PCR amplification tubes. Once the process is complete, the DNA extracts, still on the deck of the robot in PCR amplification strip tubes, are transferred to pre-labeled 1.5 mL tubes for long-term storage using an automated method. The automation of these steps in the process of forensic DNA casework analysis has been accomplished by performing extensive optimization, validation and testing of the software methods.

Cadmium and zinc in soil solution extracts following the application of phosphate fertilizers.

PubMed

Lambert, Raphaël; Grant, Cynthia; Sauvé, Sébastien

2007-06-01

This study investigated the solubility of cadmium and zinc in soils after the application of phosphate fertilizers containing those two metals. The solubility of cadmium and zinc was assessed by measuring their concentration in soil water extracts. Three monoammonium phosphate fertilizers containing various amounts of metals were applied on cultivated fields for 3 years at three different rates. In order to investigate the effects of long-term applications of fertilizers on the solubility of Cd and Zn, a similar design was used to apply contaminated fertilizers to soils in a laboratory experiment using a single fertilizer addition equivalent to 15 years of application. Phosphate fertilizers increased the concentration of Cd in soil extracts compared to control in 87% and 80% of the treatments in field and laboratory experiments respectively. Both increasing the rate of application and using fertilizer containing more Cd lead to higher Cd concentrations in extracts for the field and the laboratory experiments. The addition of the equivalent of 15 years of fertilizer application in the laboratory results in higher Cd concentration in extracts compared to the field experiment. For Zn, the fertilizer treatments enhanced the metal solution concentration in 83% of field treatments, but no significant correlations could be found between Zn inputs and its concentration in solution. In the laboratory, fertilizer additions increase the Zn concentrations in 53% of the treatments and decrease it in most of the other treatments. The decrease in Zn concentrations in the laboratory trial is attributed to the higher phosphate concentrations in the soil solution; which is presumed to have contributed to the precipitation of Zn-phosphates. For both trials, the metal concentrations in soil extracts cannot be related to the Zn concentration in the fertilizer or the rate of application. The high Zn to Cd ratio is presumably responsible for the Cd increase in the soil extracts due to competitive displacement by Zn. Finally, the observed acidification of soils with fertilizer application will also contribute to metal solubilisation.

Conditions of High Resolution Melting Analysis on the Cobas z480 Instrument for the Genotyping of VKORC1 in the Clinical Routine Laboratory.

PubMed

Paar, Christian; Hammerl, Verena; Blessberger, Hermann; Stekel, Herbert; Steinwender, Clemens; Berg, Jörg

2016-12-01

High resolution melting (HRM) of amplicons is a simple method for genotyping of single nucleotide polymorphisms (SNPs). Albeit many applications reported, HRM seems to be rarely used in clinical laboratories. The suitability of HRM-PCR for the clinical laboratory was investigated for genotyping of SNPs of the vitamin K epoxide reductase complex unit 1 gene. About 100 DNA samples were analyzed by two different HRM-PCRs on the Cobas z480 instrument and compared with a PCR with fluorescently labeled probes (HybProbe-PCR) on the LightCycler 2.0 instrument as reference. Reliable genotyping with 100% matching results was obtained, when the amplicon size was small (63 bp) and DNA input was limited by e.g., sample dilution with salt-free water. DNA extracted by differing methods may be used for genotyping by HRM-PCR. Compared with HybProbe-PCR, HRM-PCR on the Cobas z480 instrument allows for higher through-put, however, at the cost of a higher degree of laboratory standardization and a slower turnaround.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory : determination of organochlorine pesticides and polychlorinated biphenyls in bottom and suspended sediment by gas chromatography with electron-capture detection

USGS Publications Warehouse

Noriega, Mary C.; Wydoski, Duane S.; Foreman, William T.

2004-01-01

A method applicable for the determination of 19 organochlorine (OC) pesticides, including total toxaphene as a complex mixture, and 3 polychlorinated biphenyl (PCB) mixtures as Aroclor equivalents--Aroclor 1016/1242, 1254, and 1260--in soil, aquatic bottom sediment, and suspended sediment is described. Method performance data are presented. The solvent system is designed to extract simultaneously selected OC pesticides and PCBs from the same sample matrix. The compounds are extracted by conventional Soxhlet extraction with dichloromethane, followed by partial isolation using gel permeation chromatography (GPC) to remove inorganic sulfur and large naturally present molecules from the sediment extract. The aliquot of extract collected from the GPC for OCs (OC pesticides and PCBs) is split into two sample fractions by alumina/silica combined-column chromatography, followed by Florisil adsorption chromatography to remove interfering compounds in the second fraction. The OC fractions are analyzed by dual capillary-column gas chromatography with electron-capture detection (GC/ECD). This report is limited to the determination of selected OC pesticides and PCBs by GC/ECD using this method. Interim reporting levels (IRLs) have been set at 0.400 to 3.12 micrograms per kilogram for 18 individual OC pesticides, 200 micrograms per kilogram for toxaphene, and 4.04 to 4.68 micrograms per kilogram for the PCBs, based on a sample size of 25-gram equivalent dry weight. These reporting levels may change following additional determinations of method detection limits.

Efficacy of Some Botanical Extracts against Trogoderma granarium in Wheat Grains with Toxicity Evaluation

PubMed Central

Derbalah, Aly S.

2012-01-01

In an attempt to find alternative control methods for stored products insects, extracts of seven plant species (Cassia senna, Caesalpinia gilliesii, Thespesia populnea var. acutiloba, Chrysanthemum frutescens, Euonymus japonicus, Bauhinia purpurea, and Cassia fistula) were evaluated under laboratory conditions for their ability to protect wheat (Triticum spp.) grains against Trogoderma granarium insect. Moreover, gas chromatography-mass spectrometry (GC-MS) analysis was carried to identify the chemical components of the most effective plant extract against T. granarium. Furthermore, the safety of the most effective plant extract was evaluated with respect to biochemical and histological changes in treated rats relative to control. The results revealed that, the tested botanical extracts showed high efficiency against T. granarium with respect to mortality and progeny of the adults. C. senna was the most effective botanical extract against T. granarium. The GC-MS analysis of the most effective plant extract showed the presence of different bioactive compounds that is known by its insecticidal activity. The most effective plant extract showed no toxicity on treated rats relative to control with respect to biochemical and histological changes. The results suggest the ability of using these plant extracts for wheat grains protection as a safe alternative to insecticides. PMID:22606054

Qualitative identification of permitted and non-permitted colour additives in food products.

PubMed

Harp, Bhakti Petigara; Miranda-Bermudez, Enio; Baron, Carolina I; Richard, Gerald I

2012-01-01

Colour additives are dyes, pigments or other substances that can impart colour when added or applied to food, drugs, cosmetics, medical devices, or the human body. The substances must be pre-approved by the US Food and Drug Administration (USFDA) and listed in Title 21 of the US Code of Federal Regulations before they may be used in products marketed in the United States. Some also are required to be batch certified by the USFDA prior to their use. Both domestic and imported products sold in interstate commerce fall under USFDA jurisdiction, and the USFDA's district laboratories use a combination of analytical methods for identifying or confirming the presence of potentially violative colour additives. We have developed a qualitative method for identifying 17 certifiable, certification exempt, and non-permitted colour additives in various food products. The method involves extracting the colour additives from a product and isolating them from non-coloured components with a C(18) Sep-Pak cartridge. The colour additives are then separated and identified by liquid chromatography (LC) with photodiode array detection, using an Xterra RP18 column and gradient elution with aqueous ammonium acetate and methanol. Limits of detection (LODs) ranged from 0.02 to 1.49 mg/l. This qualititative LC method supplements the visible spectrophotometric and thin-layer chromatography methods currently used by the USFDA's district laboratories and is less time-consuming and requires less solvent compared to the other methods. The extraction step in the new LC method is a simple and an efficient process that can be used for most food types.

Forensic validation of the SNPforID 52-plex assay.

PubMed

Musgrave-Brown, Esther; Ballard, David; Balogh, Kinga; Bender, Klaus; Berger, Burkhard; Bogus, Magdalena; Børsting, Claus; Brion, María; Fondevila, Manuel; Harrison, Cheryl; Oguzturun, Ceylan; Parson, Walther; Phillips, Chris; Proff, Carsten; Ramos-Luis, Eva; Sanchez, Juan J; Sánchez Diz, Paula; Sobrino Rey, Bea; Stradmann-Bellinghausen, Beate; Thacker, Catherine; Carracedo, Angel; Morling, Niels; Scheithauer, Richard; Schneider, Peter M; Syndercombe Court, Denise

2007-06-01

The advantages of single nucleotide polymorphism (SNP) typing in forensic genetics are well known and include a wider choice of high-throughput typing platforms, lower mutation rates, and improved analysis of degraded samples. However, if SNPs are to become a realistic supplement to current short tandem repeat (STR) typing methods, they must be shown to successfully and reliably analyse the challenging samples commonly encountered in casework situations. The European SNPforID consortium, supported by the EU GROWTH programme, has developed a multiplex of 52 SNPs for forensic analysis, with the amplification of all 52 loci in a single reaction followed by two single base extension (SBE) reactions which are detected with capillary electrophoresis. In order to validate this assay, a variety of DNA extracts were chosen to represent problems such as low copy number and degradation that are commonly seen in forensic casework. A total of 40 extracts were used in the study, each of which was sent to two of the five participating laboratories for typing in duplicate or triplicate. Laboratories were instructed to carry out their analyses as if they were dealing with normal casework samples. Results were reported back to the coordinating laboratory and compared with those obtained from traditional STR typing of the same extracts using Powerplex 16 (Promega). These results indicate that, although the ability to successfully type good quality, low copy number extracts is lower, the 52-plex SNP assay performed better than STR typing on degraded samples, and also on samples that were both degraded and of limited quantity, suggesting that SNP analysis can provide advantages over STR analysis in forensically relevant circumstances. However, there were also additional problems arising from contamination and primer quality issues and these are discussed.

Technologies for Extracting Valuable Metals and Compounds from Geothermal Fluids

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

Harrison, Stephen

2014-04-30

Executive Summary Simbol Materials studied various methods of extracting valuable minerals from geothermal brines in the Imperial Valley of California, focusing on the extraction of lithium, manganese, zinc and potassium. New methods were explored for managing the potential impact of silica fouling on mineral extraction equipment, and for converting silica management by-products into commercial products.` Studies at the laboratory and bench scale focused on manganese, zinc and potassium extraction and the conversion of silica management by-products into valuable commercial products. The processes for extracting lithium and producing lithium carbonate and lithium hydroxide products were developed at the laboratory scale andmore » scaled up to pilot-scale. Several sorbents designed to extract lithium as lithium chloride from geothermal brine were developed at the laboratory scale and subsequently scaled-up for testing in the lithium extraction pilot plant. Lithium The results of the lithium studies generated the confidence for Simbol to scale its process to commercial operation. The key steps of the process were demonstrated during its development at pilot scale: 1. Silica management. 2. Lithium extraction. 3. Purification. 4. Concentration. 5. Conversion into lithium hydroxide and lithium carbonate products. Results show that greater than 95% of the lithium can be extracted from geothermal brine as lithium chloride, and that the chemical yield in converting lithium chloride to lithium hydroxide and lithium carbonate products is greater than 90%. The product purity produced from the process is consistent with battery grade lithium carbonate and lithium hydroxide. Manganese and zinc Processes for the extraction of zinc and manganese from geothermal brine were developed. It was shown that they could be converted into zinc metal and electrolytic manganese dioxide after purification. These processes were evaluated for their economic potential, and at the present time Simbol Materials is evaluating other products with greater commercial value. Potassium Silicotitanates, zeolites and other sorbents were evaluated as potential reagents for the extraction of potassium from geothermal brines and production of potassium chloride (potash). It was found that zeolites were effective at removing potassium but the capacity of the zeolites and the form that the potassium is in does not have economic potential. Iron-silica by-product The conversion of iron-silica by-product produced during silica management operations into more valuable materials was studied at the laboratory scale. Results indicate that it is technically feasible to convert the iron-silica by-product into ferric chloride and ferric sulfate solutions which are precursors to a ferric phosphate product. However, additional work to purify the solutions is required to determine the commercial viability of this process. Conclusion Simbol Materials is in the process of designing its first commercial plant based on the technology developed to the pilot scale during this project. The investment in the commercial plant is hundreds of millions of dollars, and construction of the commercial plant will generate hundreds of jobs. Plant construction will be completed in 2016 and the first lithium products will be shipped in 2017. The plant will have a lithium carbonate equivalent production capacity of 15,000 tonnes per year. The gross revenues from the project are expected to be approximately $ 80 to 100 million annually. During this development program Simbol grew from a company of about 10 people to over 60 people today. Simbol is expected to employ more than 100 people once the plant is constructed. Simbol Materials’ business is scalable in the Imperial Valley region because there are eleven geothermal power plants already in operation, which allows Simbol to expand its business from one plant to multiple plants. Additionally, the scope of the resource is vast in terms of potential products such as lithium, manganese and zinc and potentially potassium.« less

Identification of fish species after cooking by SDS-PAGE and urea IEF: a collaborative study.

PubMed

Etienne, M; Jérôme, M; Fleurence, J; Rehbein, H; Kündiger, R; Mendes, R; Costa, H; Pérez-Martín, R; Piñeiro-González, C; Craig, A; Mackie, I; Malmheden Yman, I; Ferm, M; Martínez, I; Jessen, F; Smelt, A; Luten, J

2000-07-01

A collaborative study, to validate the use of SDS-PAGE and urea IEF, for the identification of fish species after cooking has been performed by nine laboratories. By following optimized standard operation procedures, 10 commercially important species (Atlantic salmon, sea trout, rainbow trout, turbot, Alaska pollock, pollack, pink salmon, Arctic char, chum salmon, and New Zealand hake) had to be identified by comparison with 22 reference samples. Some differences in the recoveries of proteins from cooked fish flesh were noted between the urea and the SDS extraction procedures used. Generally, the urea extraction procedure appears to be less efficient than the SDS extraction for protein solubilization. Except for some species belonging to the Salmonidae family (Salmo, Oncorhynchus), both of the analytical techniques tested (urea IEF, SDS-PAGE) enabled identification of the species of the samples to be established. With urea IEF, two laboratories could not differentiate Salmo salar from Salmo trutta. The same difficulties were noted for differentiation between Oncorhynchus gorbuscha and Oncorhynchus keta samples. With SDS-PAGE, three laboratories had some difficulties in identifying the S. trutta samples. However, in the contrast with the previous technique, SDS-PAGE allows the characterization of most of the Oncorhynchus species tested. Only Oncorhynchus mykiss was not clearly recognized by one laboratory. Therefore, SDS-PAGE (Excel gel homogeneous 15%) appears to be better for the identification, after cooking, of fish such as the tuna and salmon species which are characterized by neutral and basic protein bands, and urea IEF (CleanGel) is better for the gadoid species, which are characterized by acid protein bands (parvalbumins). Nevertheless, in contentious cases it is preferable to use both analytical methods.

Verification of Eulerian-Eulerian and Eulerian-Lagrangian simulations for fluid-particle flows

NASA Astrophysics Data System (ADS)

Kong, Bo; Patel, Ravi G.; Capecelatro, Jesse; Desjardins, Olivier; Fox, Rodney O.

2017-11-01

In this work, we study the performance of three simulation techniques for fluid-particle flows: (1) a volume-filtered Euler-Lagrange approach (EL), (2) a quadrature-based moment method using the anisotropic Gaussian closure (AG), and (3) a traditional two-fluid model. By simulating two problems: particles in frozen homogeneous isotropic turbulence (HIT), and cluster-induced turbulence (CIT), the convergence of the methods under grid refinement is found to depend on the simulation method and the specific problem, with CIT simulations facing fewer difficulties than HIT. Although EL converges under refinement for both HIT and CIT, its statistical results exhibit dependence on the techniques used to extract statistics for the particle phase. For HIT, converging both EE methods (TFM and AG) poses challenges, while for CIT, AG and EL produce similar results. Overall, all three methods face challenges when trying to extract converged, parameter-independent statistics due to the presence of shocks in the particle phase. National Science Foundation and National Energy Technology Laboratory.

Antifungal Effect of Malaysian Aloe vera Leaf Extract on Selected Fungal Species of Pathogenic Otomycosis Species in In Vitro Culture Medium.

PubMed

Saniasiaya, Jeyasakthy; Salim, Rosdan; Mohamad, Irfan; Harun, Azian

2017-01-01

Aloe barbadensis miller or Aloe vera has been used for therapeutic purposes since ancient times with antifungal activity known to be amongst its medicinal properties. We conducted a pilot study to determine the antifungal properties of Malaysian Aloe vera leaf extract on otomycosis species including Aspergillus niger and Candida albicans. This laboratory-controlled prospective study was conducted at the Universiti Sains Malaysia. Extracts of Malaysian Aloe vera leaf was prepared in ethanol and solutions via the Soxhlet extraction method. Sabouraud dextrose agar cultured with the two fungal isolates were inoculated with the five different concentrations of each extract (50 g/mL, 25 g/mL, 12.5 g/mL, 6.25 g/mL, and 3.125 g/mL) using the well-diffusion method. Zone of inhibition was measured followed by minimum inhibitory concentration (MIC). For A. niger, a zone of inhibition for alcohol and aqueous extract was seen for all concentrations except 3.125 g/mL. There was no zone of inhibition for both alcohol and aqueous extracts of Aloe vera leaf for C. albicans . The MIC values of aqueous and alcohol extracts were 5.1 g/mL and 4.4 g/mL for A. niger and since no zone of inhibition was obtained for C. albicans the MIC was not determined. The antifungal effect of alcohol extracts of Malaysian Aloe vera leaf is better than the aqueous extract for A. niger ( p < 0.001). Malaysian Aloe vera has a significant antifungal effect towards A. niger.

Systematic optimization of ethyl glucuronide extraction conditions from scalp hair by design of experiments and its potential effect on cut-off values appraisal.

PubMed

Alladio, Eugenio; Biosa, Giulia; Seganti, Fabrizio; Di Corcia, Daniele; Salomone, Alberto; Vincenti, Marco; Baumgartner, Markus R

2018-05-11

The quantitative determination of ethyl glucuronide (EtG) in hair samples is consistently used throughout the world to assess chronic excessive alcohol consumption. For administrative and legal purposes, the analytical results are compared with cut-off values recognized by regulatory authorities and scientific societies. However, it has been recently recognized that the analytical results depend on the hair sample pretreatment procedures, including the crumbling and extraction conditions. A systematic evaluation of the EtG extraction conditions from pulverized scalp hair was conducted by design of experiments (DoE) considering the extraction time, temperature, pH, and solvent composition as potential influencing factors. It was concluded that an overnight extraction at 60°C with pure water at neutral pH represents the most effective conditions to achieve high extraction yields. The absence of differential degradation of the internal standard (isotopically-labeled EtG) under such conditions was confirmed and the overall analytical method was validated according to SGWTOX and ISO17025 criteria. Twenty real hair samples with different EtG content were analyzed with three commonly accepted procedures: (a) hair manually cut in snippets and extracted at room temperature; (b) pulverized hair extracted at room temperature; (c) hair treated with the optimized method. Average increments of EtG concentration around 69% (from a to c) and 29% (from b to c) were recorded. In light of these results, the authors urge the scientific community to undertake an inter-laboratory study with the aim of defining more in detail the optimal hair EtG detection method and verifying the corresponding cut-off level for legal enforcements. This article is protected by copyright. All rights reserved.

Fusariotoxicosis from barley in British Columbia. II. Analysis and toxicity of syspected barley.

PubMed Central

Puls, R; Greenway, J A

1976-01-01

Fusariotoxin T-2, a trichothecene, was tentatively identified in barley samples which caused field outbreaks of mycotoxicosis in British Columbia. Geese died when fed the contaminated barley experimentally but mice were little affected after long term feeding. The methods used in the laboratory for trichothecene extraction and identification of T-2 toxin are described. PMID:1000373

Organic Chemistry and the Native Plants of the Sonoran Desert: Conversion of Jojoba Oil to Biodiesel

ERIC Educational Resources Information Center

Daconta, Lisa V.; Minger, Timothy; Nedelkova, Valentina; Zikopoulos, John N.

2015-01-01

A new, general approach to the organic chemistry laboratory is introduced that is based on learning about organic chemistry techniques and research methods by exploring the natural products found in local native plants. As an example of this approach for the Sonoran desert region, the extraction of jojoba oil and its transesterification to…

The Zymogen-Enteropeptidase System: A Practical Approach to Study the Regulation of Enzyme Activity by Proteolytic Cleavage

ERIC Educational Resources Information Center

Pizauro, Joao M., Jr.; Ferro, Jesus A.; de Lima, Andrea C. F.; Routman, Karina S.; Portella, Maria Celia

2004-01-01

The present research describes an efficient procedure to obtain high levels of trypsinogen and chymotrypsinogen by using a simple, rapid, and easily reproducible method. The extraction process and the time-course of activation of zymogens can be carried out in a single laboratory period, without sophisticated equipment. The main objective was to…

Rapid Methods for the Laboratory Identification of Pathogenic Microorganisms.

DTIC Science & Technology

1981-09-01

Preliminary results provide strong evidence to show that the fungi, Candida and Cryptococcus , can be raoidly differentiated by a lectin test. SFor Oro...SUMMATION LECTIN-YEAST INTERACTIONS Objective: To find a lectin that selectively agglutinates Cryptococcus neoformans (the etiologic agent of...peanut), Conavalia ensiformis (Con A) and mango extract may potentially be utilized to differentiate Cryptococcus from the other yeasts most commonly

A Simple and Rapid Method for Preparing a Cell-Free Bacterial Lysate for Protein Synthesis

PubMed Central

Kaduri, Maya; Shainsky-Roitman, Janna; Goldfeder, Mor; Ivanir, Eran; Benhar, Itai; Shoham, Yuval; Schroeder, Avi

2016-01-01

Cell-free protein synthesis (CFPS) systems are important laboratory tools that are used for various synthetic biology applications. Here, we present a simple and inexpensive laboratory-scale method for preparing a CFPS system from E. coli. The procedure uses basic lab equipment, a minimal set of reagents, and requires less than one hour to process the bacterial cell mass into a functional S30-T7 extract. BL21(DE3) and MRE600 E. coli strains were used to prepare the S30-T7 extract. The CFPS system was used to produce a set of fluorescent and therapeutic proteins of different molecular weights (up to 66 kDa). This system was able to produce 40–150 μg-protein/ml, with variations depending on the plasmid type, expressed protein and E. coli strain. Interestingly, the BL21-based CFPS exhibited stability and increased activity at 40 and 45°C. To the best of our knowledge, this is the most rapid and affordable lab-scale protocol for preparing a cell-free protein synthesis system, with high thermal stability and efficacy in producing therapeutic proteins. PMID:27768741

Microvolume trace environmental analysis using peak-focusing online solid-phase extraction-nano-liquid chromatography-high-resolution mass spectrometry.

PubMed

Stravs, Michael A; Mechelke, Jonas; Ferguson, P Lee; Singer, Heinz; Hollender, Juliane

2016-03-01

Online solid-phase extraction was combined with nano-liquid chromatography coupled to high-resolution mass spectrometry (HRMS) for the analysis of micropollutants in environmental samples from small volumes. The method was validated in surface water, Microcystis aeruginosa cell lysate, and spent Microcystis growth medium. For 41 analytes, quantification limits of 0.1-28 ng/L (surface water) and 0.1-32 ng/L (growth medium) were obtained from only 88 μL of sample. In cell lysate, quantification limits ranged from 0.1-143 ng/L or 0.33-476 ng/g dry weight from a sample of 88 μL, or 26 μg dry weight, respectively. The method matches the sensitivity of established online and offline solid-phase extraction-liquid chromatography-mass spectrometry methods but requires only a fraction of the sample used by those techniques, and is among the first applications of nano-LC-MS for environmental analysis. The method was applied to the determination of bioconcentration in Microcystis aeruginosa in a laboratory experiment, and the benefit of coupling to HRMS was demonstrated in a transformation product screening.

Analysis of polycyclic aromatic hydrocarbons extracted from air particulate matter using a temperature programmable injector coupled to GC-C-IRMS.

PubMed

Mikolajczuk, Agnieszka; Przyk, Elzbieta Perez; Geypens, Benny; Berglund, Michael; Taylor, Philip

2010-03-01

Compound specific isotopic analysis (CSIA) can provide information about the origin of analysed compounds - in this case, polycyclic aromatic hydrocarbons (PAHs). In the study, PAHs were extracted from three dust samples: winter and summer filter dust and tunnel dust. The measurement was performed using the method validated in our laboratory using pure, solid compounds and EPA 610 reference assortment. CSIA required an appropriate clean-up method to avoid an unresolved complex in the gas chromatographic analysis usually found in the chromatography of PAHs. Extensive sample clean-up for this particular matrix was found to be necessary to obtain good gas chromatography-combustion-isotope ratio mass spectrometry analysis results. The sample purification method included two steps in which the sample is cleaned up and the aliphatic and aromatic hydrocarbons are separated. The concentration of PAHs in the measured samples was low; so a large volume injection technique (100 microl) was applied. The delta(VPDB)(13)C was measured with a final uncertainty smaller than 1 per thousand. Comparison of the delta(VPDB)(13)C signatures of PAHs extracted from different dust samples was feasible with this method and, doing so, significant differences were observed.

Microburger Biochemistry: Extraction and Spectral Characterization of Myoglyobin from Hamburger

NASA Astrophysics Data System (ADS)

Bylkas, Sheri A.; Andersson, Laura A.

1997-04-01

This experiment provides a demonstration of useful biochemical methods at a Basic or Advanced Level, depending upon the available spectrophotometric equipment. The protocol combines protein extraction, ox-i-dation and reduction, and simple spectroscopic analysis, as well as gel filtration chromatography and generation/analysis of spectral scans. Mammalian myoglobin (Mb) is a monomeric O2-binding protein that functions in muscle to store oxygen. The single iron protoporphyrin IX (heme) group is bound to protein by the amino acid Histidine93. The common, stable forms, Met-Mb and Oxy-Mb are studied because in a non-living system, red Oxy-Mb is converted to brown Met-Mb as bound O2 molecule is released. Mb is easily extracted from steak, to illustrate and address why fresh meat is red and aged meat is brown; the protein has unique spectral properties that are diagnostic for characterization of sample identity. After application of heme redox chemical methods, the MetMb or OxyMb samples can be studied spectroscopically. The color change between Oxy-Mb and Met-Mb is dramatic (illustrating bright red fresh meat vs. brown older meat), and method(s) used in this laboratory are simple, inexpensive, and non-harmful to the student.

Development and Multi-laboratory Verification of US EPA ...

EPA Pesticide Factsheets

A drinking water method for seven pesticides and pesticide degradates is presented that addresses the occurrence monitoring needs of the US Environmental Protection Agency (EPA) for a future Unregulated Contaminant Monitoring Regulation (UCMR). The method employs online solid phase extraction-liquid chromatography–tandem mass spectrometry (SPE-LC–MS-MS). Online SPE-LC–MS-MS has the potential to offer cost-effective, faster, more sensitive and more rugged methods than the traditional offline SPE approach due to complete automation of the SPE process, as well as seamless integration with the LC–MS-MS system. The method uses 2-chloroacetamide, ascorbic acid and Trizma to preserve the drinking water samples for up to 28 days. The mean recoveries in drinking water (from a surface water source) fortified with method analytes are 87.1–112% with relative standard deviations of <14%. Single laboratory lowest concentration minimum reporting levels of 0.27–1.7 ng/L are demonstrated with this methodology. Multi-laboratory data are presented that demonstrate method ruggedness and transferability. The final method meets all of the EPA's UCMR survey requirements for sample collection and storage, precision, accuracy, and sensitivity. The journal article describes the development of drinking water Method 543 for analysis of selected CCL 3 chemicals. It is anticipated this method may be used in a future Unregulated Contaminant Monitoring Regulation to gather nationw

Evaluation of Antimicrobial Activity of the Methanol Extracts from 8 Traditional Medicinal Plants

PubMed Central

Kang, Chang-Geun; Hah, Dae-Sik; Kim, Chung-Hui; Kim, Young-Hwan; Kim, Euikyung

2011-01-01

The methanol extract of 12 medicinal plants were evaluated for its antibacterial activity against Gram-positive (5 strains) and Gram-negative bacteria (10 strains) by assay for minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) . The antibacterial activity was determined by an agar dilution method (according to the guidelines of Clinical and Laboratory Standard Institute) . All the compounds (12 extracts) of the 8 medicinal plants (leaf or root) were active against both Gram-negative and Gram-positive bacteria. Gram-negative showed a more potent action than Gram positive bacteria. The MIC concentrations were various ranged from 0.6 μg/ml to 5000 μg/ml. The lowest MIC (0.6 μg/ml) and MBC (1.22 μg/ml) values were obtained with extract on 4 and 3 of the 15 microorganisms tested, respectively. PMID:24278548

Detection and Quantification of Cannabinoids in Extracts of Cannabis sativa Roots Using LC-MS/MS.

PubMed

Gul, Waseem; Gul, Shahbaz W; Chandra, Suman; Lata, Hemant; Ibrahim, Elsayed A; ElSohly, Mahmoud A

2018-03-01

A liquid chromatography-tandem mass spectrometry single-laboratory validation was performed for the detection and quantification of the 10 major cannabinoids of cannabis, namely, (-)- trans -Δ 9 -tetrahydrocannabinol, cannabidiol, cannabigerol, cannabichromene, tetrahydrocannabivarian, cannabinol, (-)- trans -Δ 8 -tetrahydrocannabinol, cannabidiolic acid, cannabigerolic acid, and Δ 9 -tetrahydrocannabinolic acid-A, in the root extract of Cannabis sativa . Acetonitrile : methanol (80 : 20, v/v) was used for extraction; d 3 -cannabidiol and d 3 - tetrahydrocannabinol were used as the internal standards. All 10 cannabinoids showed a good regression relationship with r 2  > 0.99. The validated method is simple, sensitive, and reproducible and is therefore suitable for the detection and quantification of these cannabinoids in extracts of cannabis roots. To our knowledge, this is the first report for the quantification of cannabinoids in cannabis roots. Georg Thieme Verlag KG Stuttgart · New York.

Determination of ephedrine alkaloids in botanicals and dietary supplements by HPLC-UV: collaborative study.

PubMed

Roman, Mark C

2004-01-01

An international collaborative study was conducted of a high-performance liquid chromatography (HPLC)-UV method for the determination of the major (ephedrine [EP] and pseudoephedrine [PS]) and minor (norephedrine [NE], norpseudoephedrine [NP], methylephedrine [ME], and methylpseudoephedrine [MP]) alkaloids in selected dietary supplements representative of the commercially available products. Ten collaborating laboratories determined the ephedrine-type alkaloid content in 8 blind replicate samples. Five products contained ephedra ground herb or ephedra extract. These 5 products included ground botanical raw material of Ephedra sinica, a common powdered extract of Ephedra sinica, a finished product containing only Ephedra sinica ground botanical raw material, a complex multicomponent dietary supplement containing Ma Huang, and a high-protein chocolate flavored drink mix containing Ma Huang extract. In addition, collaborating laboratories received a negative control and negative control spiked with ephedrine alkaloids at high and low levels for recovery studies. Test extracts were treated to solid-phase extraction using a strong-cation exchange column to help remove interferences. The HPLC analyses were performed on a polar-embedded phenyl column using UV detection at 210 nm. Repeatability relative standard deviations (RSDr) ranged from 0.64-3.0% for EP and 2.0-6.6% for PS, excluding the high protein drink mix. Reproducibility relative standard deviations (RSDR) ranged from 2.1-6.6% for EP and 9.0-11.4% for PS, excluding the high protein drink mix. Recoveries ranged from 84.7-87.2% for EP and 84.6-98.2% for PS. The data developed for the minor alkaloids are more variable with generally unsatisfactory HORRATS (i.e., >2). However, since these alkaloids generally add little to the total alkaloid content of the products, the method gives satisfactory results in measuring total alkaloid content (RSDr 0.85-3.13%; RSDR 2.03-10.97%, HORRAT 0.69-3.23, exclusive of the results from the high protein drink). On the basis of these results, the method is recommended for Official First Action for determination of EP and PS in dietary supplements exclusive of the high protein drinks.

Teaching Experimental Design Using a GC MS Analysis of Cocaine on Money: A Cross-Disciplinary Laboratory 1254 Christopher A. Heimbuck and Nathan W. Bower Quantitative Determination of Nicotine and Cotinine in Urine and Sputum Using a Combined SPME-GC/MS Method

NASA Astrophysics Data System (ADS)

Witter, A. E.; Klinger, D. M.; Fan, X.; Lam, M.; Mathers, D. T.; Mabury, S. A.

2002-10-01

The forensic analysis of cocaine on currencies was optimized using a fractional, two-level experimental design that compared methanol and HCl extraction, SPE versus heptane pre-concentration, and extracted versus total ion chromatography. Subsequent student-initiated questions about levels of cocaine on U.S. and world currencies helped make connections to societal issues while teaching method optimization and chromatography. A significant correlation was found between the levels of cocaine and the age of the bills. Levels of cocaine on various world currencies followed expected drug-trafficking patterns with the highest levels found in the most developed countries.

Safety and Toxicity of Saw palmetto in the Complementary and Alternative Medicine for Urological Symptoms (CAMUS) Trial

PubMed Central

Avins, Andrew L.; Lee, Jeannette Y.; Meyers, Catherine M.; Barry, Michael J.

2013-01-01

Purpose Extracts of the saw palmetto berry are used by many men in the U.S. as self-treatment for lower urinary tract symptoms due to benign prostatic hyperplasia. While the most recent data from double-blind clinical trials do not support efficacy superior to that of placebo, there are few data on the toxicity of saw palmetto. Materials and Methods 369 patients were randomized in the Complementary and Alternative Medicine for Urological Symptoms (CAMUS) trial; 357 participants are included in this modified intention-to-treat analysis. Participants were randomized to 320mg, 640mg, and 960mg daily of an ethanolic saw palmetto extract or an identical-appearing placebo, in an escalating manner at 6-month intervals, for a total of 18 months follow-up. Adverse-event assessments, vital signs, and blood and urine laboratory tests were obtained at regular intervals. Results There were no statistically significant differences between groups in rates of serious or non-serious adverse events, changes in vital signs, digital prostate exam findings, or study withdrawal rates. Overall, there were no significant inter-group differences in the occurrence of laboratory-test abnormalities; differences in individual laboratory tests were uncommon and small in magnitude. No evidence of significant dose-response phenomena were identified. Conclusions The saw palmetto extract used in the CAMUS trial showed no evidence of toxicity at doses up to three times the usual clinical dose over a period of 18 months. PMID:23063633

An image-processing methodology for extracting bloodstain pattern features.

PubMed

Arthur, Ravishka M; Humburg, Philomena J; Hoogenboom, Jerry; Baiker, Martin; Taylor, Michael C; de Bruin, Karla G

2017-08-01

There is a growing trend in forensic science to develop methods to make forensic pattern comparison tasks more objective. This has generally involved the application of suitable image-processing methods to provide numerical data for identification or comparison. This paper outlines a unique image-processing methodology that can be utilised by analysts to generate reliable pattern data that will assist them in forming objective conclusions about a pattern. A range of features were defined and extracted from a laboratory-generated impact spatter pattern. These features were based in part on bloodstain properties commonly used in the analysis of spatter bloodstain patterns. The values of these features were consistent with properties reported qualitatively for such patterns. The image-processing method developed shows considerable promise as a way to establish measurable discriminating pattern criteria that are lacking in current bloodstain pattern taxonomies. Copyright © 2017 Elsevier B.V. All rights reserved.

Syringe-cartridge solid-phase extraction method for patulin in apple juice.

PubMed

Eisele, Thomas A; Gibson, Midori Z

2003-01-01

A syringe-cartridge solid-phase extraction (SPE) method was developed for determination of patulin in apple juice. A 2.5 mL portion of test sample was passed through a conditioned macroporous SPE cartridge and washed with 2 mL 1% sodium bicarbonate followed by 2 mL 1% acetic acid. Patulin was eluted with 1 mL 10% ethyl acetate in ethyl ether and determined by reversed-phase liquid chromatography using a mobile phase consisting of 81% acetonitrile, 9% water, and 10% 0.05M potassium phosphate buffer, pH 2.4. Recoveries averaged 92% and the relative standard deviation was 8.0% in test samples spiked with 50 ng/mL patulin. The method appears to be applicable for monitoring apple juice samples to meet the U.S. Food and Drug Administration compliance action level of 50 microg/kg in an industrial quality assurance laboratory environment.

Using SPME fibers and Tenax to predict the bioavailability of pyrethroids and chlorpyrifos in field sediments.

PubMed

Harwood, Amanda D; Landrum, Peter F; Weston, Donald P; Lydy, Michael J

2013-02-01

The presence of pyrethroids in both urban and agricultural sediments at levels lethal to invertebrates has been well documented. However, variations in bioavailability among sediments make accurate predictions of toxicity based on whole sediment concentrations difficult. A proposed solution to this problem is the use of bioavailability-based estimates, such as solid phase microextraction (SPME) fibers and Tenax beads. This study compared three methods to assess the bioavailability and ultimately toxicity of pyrethroid pesticides including field-deployed SPME fibers, laboratory-exposed SPME fibers, and a 24-h Tenax extraction. The objective of the current study was to compare the ability of these methods to quantify the bioavailable fraction of pyrethroids in contaminated field sediments that were toxic to benthic invertebrates. In general, Tenax proved a more sensitive method than SPME fibers and a correlation between Tenax extractable concentrations and mortality was observed. Copyright © 2012 Elsevier Ltd. All rights reserved.

Simultaneous determination of the HIV nucleoside analogue reverse transcriptase inhibitors lamivudine, didanosine, stavudine, zidovudine and abacavir in human plasma by reversed phase high performance liquid chromatography.

PubMed

Verweij-van Wissen, C P W G M; Aarnoutse, R E; Burger, D M

2005-02-25

A reversed phase high performance liquid chromatography method was developed for the simultaneous quantitative determination of the nucleoside reverse transcriptase inhibitors (NRTIs) lamivudine, didanosine, stavudine, zidovudine and abacavir in plasma. The method involved solid-phase extraction with Oasis MAX cartridges from plasma, followed by high performance liquid chromatography with a SymmetryShield RP 18 column and ultraviolet detection set at a wavelength of 260 nm. The assay was validated over the concentration range of 0.015-5 mg/l for all five NRTIs. The average accuracies for the assay were 92-102%, inter- and intra-day coefficients of variation (CV) were <2.5% and extraction recoveries were higher than 97%. This method proved to be simple, accurate and precise, and is currently in use in our laboratory for the quantitative analysis of NRTIs in plasma.

A Microcontroller Operated Device for the Generation of Liquid Extracts from Conventional Cigarette Smoke and Electronic Cigarette Aerosol.

PubMed

Anderson, Chastain A; Bokota, Rachael E; Majeste, Andrew E; Murfee, Walter L; Wang, Shusheng

2018-01-18

Electronic cigarettes are the most popular tobacco product among middle and high schoolers and are the most popular alternative tobacco product among adults. High quality, reproducible research on the consequences of electronic cigarette use is essential for understanding emerging public health concerns and crafting evidence based regulatory policy. While a growing number of papers discuss electronic cigarettes, there is little consistency in methods across groups and very little consensus on results. Here, we describe a programmable laboratory device that can be used to create extracts of conventional cigarette smoke and electronic cigarette aerosol. This protocol details instructions for the assembly and operation of said device, and demonstrates the use of the generated extract in two sample applications: an in vitro cell viability assay and gas-chromatography mass-spectrometry. This method provides a tool for making direct comparisons between conventional cigarettes and electronic cigarettes, and is an accessible entry point into electronic cigarette research.

Clonazepam quantification in human plasma by high-performance liquid chromatography coupled with electrospray tandem mass spectrometry in a bioequivalence study.

PubMed

Cavedal, Luiz E; Mendes, Fabiana D; Domingues, Claudia C; Patni, Anil K; Monif, Tausif; Reyar, Simrit; Pereira, Alberto Dos S; Mendes, Gustavo D; De Nucci, Gilberto

2007-01-01

A rapid, sensitive and specific method for quantifying clonazepam in human plasma using diazepam as the internal standard (IS) is described. The analyte and the IS were extracted from plasma by liquid-liquid extraction using a hexane/diethylether (20 : 80, v/v) solution. The extracts were analysed by high-performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC-MS-MS). Chromatography was performed on a Jones Genesis C8 4 microm analytical column (100 x 2.1 mm i.d.). The method had a chromatographic run time of 3.0 min and a linear calibration curve over the range 0.5-50 ng/ml (r2 > 0.9965). The limit of quantification was 0.5 ng/ml. This HPLC/MS/MS procedure was used to assess the bioequivalence of two clonazepam 2 mg tablet formulations (clonazepam test formulation from Ranbaxy Laboratories Ltd and Rivotril from Roche Laboratórios Ltda as standard reference formulation). Copyright 2006 John Wiley & Sons, Ltd.

Determination of phosphate in soil extracts in the field: A green chemistry enzymatic method.

PubMed

Campbell, Ellen R; Warsko, Kayla; Davidson, Anna-Marie; Bill Campbell, Wilbur H

2015-01-01

Measurement of ortho-phosphate in soil extracts usually involves sending dried samples of soil to a laboratory for analysis and waiting several weeks for the results. Phosphate determination methods often involve use of strong acids, heavy metals, and organic dyes. To overcome limitations of this approach, we have developed a phosphate determination method which can be carried out in the field to obtain results on the spot. This new method uses: •Small volumes.•An enzymatic reaction.•Green chemistry. First, the soil sample is extracted with deionized water and filtered. Next, an aliquot of the soil extract (0.5 mL) is transferred to a disposable cuvette, containing 0.5 mL of reaction mixture [200 mM HEPES, pH 7.6, 20 mM MgCl2, with 80 nmol 2-amino-6-mercapto-7-methylpurine ribonucleoside (MESG) and 1 unit of recombinant purine nucleoside phosphorylase (PNP; EC 2.4.2.1)], mixed, and incubated for 10 min at field temperature. Absorbance of the completed reaction is measured at 360 nm in open-source, portable photometer linked by bluetooth to a smartphone. The phosphate and phosphorus content of the soil is determined by comparison of its absorbance at 360 nm to a previously prepared standard phosphate curve, which is stored in the smartphone app.

Determination of phosphate in soil extracts in the field: A green chemistry enzymatic method

PubMed Central

Campbell, Ellen R.; Warsko, Kayla; Davidson, Anna-Marie; (Bill) Campbell, Wilbur H.

2015-01-01

Measurement of ortho-phosphate in soil extracts usually involves sending dried samples of soil to a laboratory for analysis and waiting several weeks for the results. Phosphate determination methods often involve use of strong acids, heavy metals, and organic dyes. To overcome limitations of this approach, we have developed a phosphate determination method which can be carried out in the field to obtain results on the spot. This new method uses: • Small volumes. • An enzymatic reaction. • Green chemistry. First, the soil sample is extracted with deionized water and filtered. Next, an aliquot of the soil extract (0.5 mL) is transferred to a disposable cuvette, containing 0.5 mL of reaction mixture [200 mM HEPES, pH 7.6, 20 mM MgCl2, with 80 nmol 2-amino-6-mercapto-7-methylpurine ribonucleoside (MESG) and 1 unit of recombinant purine nucleoside phosphorylase (PNP; EC 2.4.2.1)], mixed, and incubated for 10 min at field temperature. Absorbance of the completed reaction is measured at 360 nm in open-source, portable photometer linked by bluetooth to a smartphone. The phosphate and phosphorus content of the soil is determined by comparison of its absorbance at 360 nm to a previously prepared standard phosphate curve, which is stored in the smartphone app. PMID:26150991

Simultaneous analysis of Alternaria toxins and citrinin in tomato: an optimised method using liquid chromatography-tandem mass spectrometry

PubMed Central

Tölgyesi, Ádám; Stroka, Joerg; Tamosiunas, Vytautas; Zwickel, Theresa

2015-01-01

Alternaria toxins and citrinin are mycotoxins produced by fungi growing on different raw materials and agricultural commodities. Maximum levels of these toxins in foods are currently under consideration by the European Commission as a risk management measure. In this study, a new quantitative method is described for the determination of five Alternaria toxins and citrinin in tomato and tomato juice samples based on LC-MS/MS detection. Samples were extracted with pure methanol, followed by a derivatisation step with 2,4-dinitrophenylhydrazine to improve the determination of tenuazonic acid and to decrease the wide polarity difference between the compounds of interest. Samples were purified on hydrophilic-modified styrene polymer solid-phase extraction cartridges. High-performance liquid chromatographic columns packed with different core–shell materials were tested for the separation of toxins and a C-18 phase was in the final method applied to achieve sufficient separation of all relevant analytes. A key element of this approach was to prove successful transferability of the method to three different triple quadrupole mass spectrometers. A full single laboratory method validation was performed on two LC-MS/MS systems and performance characteristics met the predefined requirements. Moreover, the method was used in an international proficiency test and the satisfactory z-scores obtained (−0.1 to 0.8 in tomato juice samples) demonstrated the reliability of the approach described. The method will be validated in an inter-laboratory collaborative study and if the criteria for method precision are met, the method will be proposed as a new Work Item to the European Committee for Standardisation. PMID:26212568

Determination of Yohimbine in Yohimbe Bark and Related Dietary Supplements Using UHPLC-UV/MS: Single-Laboratory Validation.

PubMed

Chen, Pei; Bryden, Noella

2015-01-01

A single-laboratory validation was performed on a practical ultra-HPLC (UHPLC)-diode array detector (DAD)/tandem MS method for determination of yohimbine in yohimbe barks and related dietary supplements. Good separation was achieved using a Waters Acquity ethylene bridged hybrid C18 column with gradient elution using 0.1% (v/v) aqueous ammonium hydroxide and 0.1% ammonium hydroxide in methanol as the mobile phases. The method can separate corynanthine from yohimbine in yohimbe bark extract, which is critical for accurate quantitation of yohimbine in yohimbe bark and related dietary supplements. Accuracy of the method was demonstrated using standard addition methods. Both intraday and interday precisions of the method were good. The method can be used without MS since yohimbine concentration in yohimbe barks and related dietary supplements are usually high enough for DAD detection, which can make it an easy and economical method for routine analysis of yohimbe barks and related dietary supplements. On the other hand, the method can be used with MS if desired for more challenging work such as biological and/or clinical studies.

Determination of Antimycin-A in water by liquid chromatographic/mass spectrometry: single-laboratory validation

USGS Publications Warehouse

Bernardy, Jeffry A.; Hubert, Terrance D.; Ogorek, Jacob M.; Schmidt, Larry J.

2013-01-01

An LC/MS method was developed and validated for the quantitative determination and confirmation of antimycin-A (ANT-A) in water from lakes or streams. Three different water sample volumes (25, 50, and 250 mL) were evaluated. ANT-A was stabilized in the field by immediately extracting it from water into anhydrous acetone using SPE. The stabilized concentrated samples were then transported to a laboratory and analyzed by LC/MS using negative electrospray ionization. The method was determined to have adequate accuracy (78 to 113% recovery), precision (0.77 to 7.5% RSD with samples ≥500 ng/L and 4.8 to 17% RSD with samples ≤100 ng/L), linearity, and robustness over an LOQ range from 8 to 51 600 ng/L.

Determination of antimycin-A in water by liquid chromatographic/mass spectrometry: single-laboratory validation.

PubMed

Bernardy, Jeffry A; Hubert, Terrance D; Ogorek, Jacob M; Schmidt, Larry J

2013-01-01

An LC/MS method was developed and validated for the quantitative determination and confirmation of antimycin-A (ANT-A) in water from lakes or streams. Three different water sample volumes (25, 50, and 250 mL) were evaluated. ANT-A was stabilized in the field by immediately extracting it from water into anhydrous acetone using SPE. The stabilized concentrated samples were then transported to a laboratory and analyzed by LC/MS using negative electrospray ionization. The method was determined to have adequate accuracy (78 to 113% recovery), precision (0.77 to 7.5% RSD with samples > or = 500 ng/L and 4.8 to 17% RSD with samples < or = 100 ng/L), linearity, and robustness over an LOQ range from 8 to 51 600 ng/L.

Various extraction and analytical techniques for isolation and identification of secondary metabolites from Nigella sativa seeds.

PubMed

Liu, X; Abd El-Aty, A M; Shim, J-H

2011-10-01

Nigella sativa L. (black cumin), commonly known as black seed, is a member of the Ranunculaceae family. This seed is used as a natural remedy in many Middle Eastern and Far Eastern countries. Extracts prepared from N. sativa have, for centuries, been used for medical purposes. Thus far, the organic compounds in N. sativa, including alkaloids, steroids, carbohydrates, flavonoids, fatty acids, etc. have been fairly well characterized. Herein, we summarize some new extraction techniques, including microwave assisted extraction (MAE) and supercritical extraction techniques (SFE), in addition to the classical method of hydrodistillation (HD), which have been employed for isolation and various analytical techniques used for the identification of secondary metabolites in black seed. We believe that some compounds contained in N. sativa remain to be identified, and that high-throughput screening could help to identify new compounds. A study addressing environmentally-friendly techniques that have minimal or no environmental effects is currently underway in our laboratory.

Preparation of a sewage sludge laboratory quality control material for butyltin compounds and their determination by isotope-dilution mass spectrometry.

PubMed

Zuliani, Tea; Milačič, Radmila; Ščančar, Janez

2012-05-01

The characterisation of a laboratory quality control material (QCM) for dibutyltin (DBT) and tributyltin (TBT) in sewage sludge is described. The reference values were determined by the use of two different types of isotope-dilution mass spectrometry: gas chromatography-mass spectrometry and gas chromatography-inductively coupled plasma mass spectrometry. To avoid possible analytical errors such as non-quantitative extraction and species degradation during sample preparation, different extraction methods were tested (microwave- and ultrasound-assisted extraction and mechanical stirring). The reference values were based on the unweighted means of results from the homogenisation and characterisation studies. The reference values obtained were 1,553 ± 87 and 534 ± 38 ng Sn g(-1) for DBT and TBT, respectively. In the uncertainty budget estimation, the sample inhomogeneity and between-method imprecision were taken into account. The concentrations of DBT and TBT in QCM are similar to those in the harbour sediment certified reference material PACS-2. Likewise, the levels of DBT and TBT are in the range of these compounds normally present in sewage sludge worldwide. In the future, the QCM will be used for an intercomparison study on DBT and TBT in sewage sludge, and as a day-to-day QCM during studies concerning the application of sewage sludge as an additive to artificial soil or as a raw material in civil engineering construction.

A New Method and Mass-Spectrometric Instrument for Extraterrestrial Microbial Life Detection Using the Elemental Composition Analyses of Martian Regolith and Permafrost/Ice.

PubMed

Managadze, G G; Safronova, A A; Luchnikov, K A; Vorobyova, E A; Duxbury, N S; Wurz, P; Managadze, N G; Chumikov, A E; Khamizov, R Kh

2017-05-01

We propose a new technique for the detection of microorganisms by elemental composition analyses of a sample extracted from regolith, permafrost, and ice of extraterrestrial bodies. We also describe the design of the ABIMAS instrument, which consists of the onboard time-of-flight laser mass-reflectron (TOF LMR) and the sample preparation unit (SPU) for biomass extraction. This instrument was initially approved to fly on board the ExoMars 2020 lander mission. The instrument can be used to analyze the elemental composition of possible extraterrestrial microbial communities and compare it to that of terrestrial microorganisms. We have conducted numerous laboratory studies to confirm the possibility of biomass identification via the following biomarkers: P/S and Ca/K ratios, and C and N abundances. We underline that only the combination of these factors will allow one to discriminate microbial samples from geological ones. Our technique has been tested experimentally in numerous laboratory trials on cultures of microorganisms and polar permafrost samples as terrestrial analogues for martian polar soils. We discuss various methods of extracting microorganisms and sample preparation. The developed technique can be used to search for and identify microorganisms in different martian samples and in the subsurface of other planets, satellites, comets, and asteroids-in particular, Europa, Ganymede, and Enceladus. Key Words: Mass spectrometry-Life-detection instruments-Biomarkers-Earth Mars-Biomass spectra. Astrobiology 17, 448-458.

A new carbon-based magnetic material for the dispersive solid-phase extraction of UV filters from water samples before liquid chromatography-tandem mass spectrometry analysis.

PubMed

Piovesana, Susy; Capriotti, Anna Laura; Cavaliere, Chiara; La Barbera, Giorgia; Samperi, Roberto; Zenezini Chiozzi, Riccardo; Laganà, Aldo

2017-07-01

Magnetic solid-phase extraction is one of the most promising new extraction methods for liquid samples before ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analysis. Several types of materials, including carbonaceous ones, have been prepared for this purpose. In this paper, for the first time, the preparation, characterization, and sorption capability of Fe 3 O 4 -graphitized carbon black (mGCB) composite toward some compounds of environmental interest were investigated. The synthesized mGCB consisted of micrometric GCB particles with 55 m 2  g -1 surface area bearing some carbonyl and hydroxyl functionalities and the surface partially decorated by Fe 3 O 4 microparticles. The prepared mGCB was firstly tested as an adsorbent for the extraction from surface water of 50 pollutants, including estrogens, perfluoroalkyl compounds, UV filters, and quinolones. The material showed good affinity to many of the tested compounds, except carboxylates and glucoronates; however, some compounds were difficult to desorb. Ten UV filters belonging to the chemical classes of benzophenones and p-aminobenzoates were selected, and parameters were optimized for the extraction of these compounds from surface water before UHPLC-MS/MS determination. Then, the method was validated in terms of linearity, trueness, intra-laboratory precision, and detection and quantification limits. In summary, the method performance (trueness, expressed as analytical recovery, 85-114%; RSD 5-15%) appears suitable for the determination of the selected compounds at the level of 10-100 ng L -1 , with detection limits in the range of 1-5 ng L -1 . Finally, the new method was compared with a published one, based on conventional solid-phase extraction with GCB, showing similar performance in real sample analysis. Graphical Abstract Workflow of the analytical method based on magnetic solid-phase extraction followed by LC-MS/MS determination.

Identification and measurement of beta-lactam antibiotic residues in milk: integration of screening kits with liquid chromatography.

PubMed

Harik-Khan, R; Moats, W A

1995-01-01

A procedure for identifying and quantitating violative beta-lactams in milk is described. This procedure integrates beta-lactam residue detection kits with the multiresidue automated liquid chromatographic (LC) cleanup method developed in our laboratory. Spiked milk was deproteinized, extracted, and subjected to reversed-phase LC using a gradient program that concentrated the beta-lactams. Amoxicillin, ampicillin, cephapirin, ceftiofur, cloxacillin, and penicillin G were, thus, separated into 5 fractions that were subsequently tested for activity by using 4 kits. beta-lactams in the positive fractions were quantitated by analytical LC methods developed in our laboratory. The LC cleanup method separated beta-lactam antibiotics from each other and from interferences in the matrix and also concentrated the antibiotics, thus increasing the sensitivity of the kits to the beta-lactam antibiotics. The procedure facilitated the task of identifying and measuring the beta-lactam antibiotics that may be present in milk samples.

A simple method for the construction of small format tissue arrays

PubMed Central

Hidalgo, A; Piña, P; Guerrero, G; Lazos, M; Salcedo, M

2003-01-01

Tissue arrays can evaluate molecular targets in high numbers of samples in parallel. Array construction presents technical difficulties and tissue arrayers are expensive, particularly for small and medium sized laboratories. This report describes a method for the construction of 36 sample arrays using widely available materials. A blunted 16 gauge needle for bone marrow aspiration was used to extract paraffin wax cylinders and manually define a 6 × 6 matrix on a blank paraffin wax block. Tissue cores from 36 paraffin wax embedded premalignant lesions and invasive cervical carcinomas were injected into the matrix using a 14 gauge needle. This tissue array was sectioned using a standard microtome and used for the immunodetection of CD44 variant 9 and interleukin 18 with satisfactory results. This method can be applied in any laboratory, without the need of specialised equipment, offering a good alternative for the wider application of tissue arrays. PMID:12560397

The advent of a new pseudoephedrine product to combat methamphetamine abuse.

PubMed

Brzeczko, Albert W; Leech, Ronald; Stark, Jeffrey G

2013-09-01

The personal and societal effects of methamphetamine abuse are well documented. The ease of accessibility to methamphetamine and the quality of the "high" it produces makes the drug highly desired by its abusers. Over time, many methamphetamine users will also become methamphetamine cooks, where pseudoephedrine in over-the-counter cold products is converted to methamphetamine through a simple, albeit extremely dangerous, process. New laws limiting access to these products have had limited success. No existing commercial pseudoephedrine products offer significant impediments to slow or limit the extraction and conversion of pseudoephedrine in clandestine methamphetamine laboratories. A new pseudoephedrine 30 mg tablet product using Impede technology (Nexafed®) to deter methamphetamine production has recently been introduced into the marketplace. Using methods designed to mimic clandestine laboratory processes, the ability of this product to disrupt extraction and conversion of pseudoephedrine to methamphetamine yet provide therapeutic effectiveness was evaluated. Impede™ technology tablets limited the extraction and/or conversion of pseudoephedrine to methamphetamine when compared to a commercially marketed pseudoephedrine product (Sudafed®). Nexafed® tablets were also shown to be bioequivalent to the same control product, thus ensuring therapeutic equivalence. With the advent of new pseudoephedrine products in the marketplace with features to limit the extraction and conversion of pseudoephedrine to methamphetamine, new tools are now available to minimize the clandestine manufacture of the drug and potentially limit its social impact.

Comparison of pore water samplers and cryogenic distillation under laboratory and field conditions for soil water stable isotope analysis.

PubMed

Thoma, Michael; Frentress, Jay; Tagliavini, Massimo; Scandellari, Francesca

2018-02-15

We used pore water samplers (PWS) to sample for isotope analysis (1) only water, (2) soil under laboratory conditions, and (3) soil in the field comparing the results with cryogenic extraction (CE). In (1) and (2), no significant differences between source and water extracted with PWS were detected with a mean absolute difference (MAD) always lower than 2 ‰ for δ 2 H and 1 ‰ for δ 18 O. In (2), CE water was more enriched than PWS-extracted water, with a MAD respect to source water of roughly 8 ‰ for δ 2 H and 4 ‰ for δ 18 O. In (3), PWS water was enriched relative to CE water by 3 ‰ for δ 2 H and 0.9 ‰ for δ 18 O. The latter result may be due to the distinct water portions sampled by the two methods. Large pores, easily sampled by PWS, likely retain recent, and enriched, summer precipitation while small pores, only sampled by CE, possibly retain isotopically depleted water from previous winter precipitation or irrigation inputs. Accuracy and precision were greater for PWS relative to CE. PWS is therefore suggested as viable tool to extract soil water for stable isotope analysis, particularly for soils used in this study (sandy and silty loams).

Response to comment on “Persistence of pharmaceutical compounds and other organic wastewater contaminants in a conventional drinking-water-treatment plant”

USGS Publications Warehouse

Stackelberg, Paul E.; Furlong, Edward T.; Meyer, Michael T.; Zaugg, Steven D.; Henderson, Alden K.; Reissman, Dori B.

2006-01-01

The U.S. Geological Survey (USGS) and the Centers for Disease Control thank Dr. Till for her comments concerning our research (Till, 2005) and welcome the opportunity to respond. The primary objective of our study was to evaluate the potential for organic wastewater-related contaminants (OWCs), including pharmaceuticals, to survive a conventional drinking-water-treatment process and persist in potable-water supplies (Stackelberg et al., 2004). Our study was supported by two USGS laboratories: the National Water Quality Laboratory (NWQL), which provided the HPLC/ESI-MS and CLLE GC/MS data and the Ocala Water Quality and Research Laboratory (OWQRL), which provided the LC/MS data (Stackelberg et al., 2004). Although discussed as distinct techniques by Dr. Till and indicated by differing acronyms to distinguish the laboratories producing the data, as described in our paper, the two LC/MS methods are very similar; they consist of a solid-phase extraction method with analysis of the extract produced using high-performance liquid chromatography coupled to an electrospray ionization mass spectrometer operated in the positive mode. The NWQL and OWQRL report ‘trace’ and ‘ultratrace’ determinations of analytes that provide significant benefit for describing the presence and fate of low-level contaminants. For mass spectral methods, an analyte is qualitatively identified by its retention time on the chromatographic column as well as the presence of two or more confirming ions with area ratios that match that of the reference standard compounds. Because of a recognized increased risk of false positives, these qualitative identification criteria are used in conjunction with abundant quality-control samples (detailed below) to confirm detection prior to making an estimate of the concentration. These qualitative identification criteria must be met before a compound is considered present (or detected) in a sample (Oblinger Childress et al., 1999). When a compound has been qualitatively identified in an environmental sample (whether above or below its reporting level [RL]), it is assessed in context with associated field and laboratory blanks, field and laboratory replicates, and other data, such as appropriate laboratory reagent spikes. An environmental concentration is calculated only after determining that field and laboratory procedures did not contaminate the samples. The concentrations are then calculated from 5- to 8-point calibration curves using internal standard quantitation. Our lowest calibration standard is intentionally much lower than the RL, typically 10 times lower. The most abundant molecular or fragment ion is used for quantitation, and, for the two LC/MS methods, at least one, and where possible two, qualifier ions are used for confirmation. For the GC/MS method, with its greater degree of fragmentation, one quantitation and two qualifier ions are used. When any of the abovementioned qualitative identification criteria are not met, the analyte is considered not present and is reported as “less than” the RL.

Improved Methods for Capture, Extraction, and Quantitative Assay of Environmental DNA from Asian Bigheaded Carp (Hypophthalmichthys spp.)

PubMed Central

Turner, Cameron R.; Miller, Derryl J.; Coyne, Kathryn J.; Corush, Joel

2014-01-01

Indirect, non-invasive detection of rare aquatic macrofauna using aqueous environmental DNA (eDNA) is a relatively new approach to population and biodiversity monitoring. As such, the sensitivity of monitoring results to different methods of eDNA capture, extraction, and detection is being investigated in many ecosystems and species. One of the first and largest conservation programs with eDNA-based monitoring as a central instrument focuses on Asian bigheaded carp (Hypophthalmichthys spp.), an invasive fish spreading toward the Laurentian Great Lakes. However, the standard eDNA methods of this program have not advanced since their development in 2010. We developed new, quantitative, and more cost-effective methods and tested them against the standard protocols. In laboratory testing, our new quantitative PCR (qPCR) assay for bigheaded carp eDNA was one to two orders of magnitude more sensitive than the existing endpoint PCR assays. When applied to eDNA samples from an experimental pond containing bigheaded carp, the qPCR assay produced a detection probability of 94.8% compared to 4.2% for the endpoint PCR assays. Also, the eDNA capture and extraction method we adapted from aquatic microbiology yielded five times more bigheaded carp eDNA from the experimental pond than the standard method, at a per sample cost over forty times lower. Our new, more sensitive assay provides a quantitative tool for eDNA-based monitoring of bigheaded carp, and the higher-yielding eDNA capture and extraction method we describe can be used for eDNA-based monitoring of any aquatic species. PMID:25474207

Improved methods for capture, extraction, and quantitative assay of environmental DNA from Asian bigheaded carp (Hypophthalmichthys spp.).

PubMed

Turner, Cameron R; Miller, Derryl J; Coyne, Kathryn J; Corush, Joel

2014-01-01

Indirect, non-invasive detection of rare aquatic macrofauna using aqueous environmental DNA (eDNA) is a relatively new approach to population and biodiversity monitoring. As such, the sensitivity of monitoring results to different methods of eDNA capture, extraction, and detection is being investigated in many ecosystems and species. One of the first and largest conservation programs with eDNA-based monitoring as a central instrument focuses on Asian bigheaded carp (Hypophthalmichthys spp.), an invasive fish spreading toward the Laurentian Great Lakes. However, the standard eDNA methods of this program have not advanced since their development in 2010. We developed new, quantitative, and more cost-effective methods and tested them against the standard protocols. In laboratory testing, our new quantitative PCR (qPCR) assay for bigheaded carp eDNA was one to two orders of magnitude more sensitive than the existing endpoint PCR assays. When applied to eDNA samples from an experimental pond containing bigheaded carp, the qPCR assay produced a detection probability of 94.8% compared to 4.2% for the endpoint PCR assays. Also, the eDNA capture and extraction method we adapted from aquatic microbiology yielded five times more bigheaded carp eDNA from the experimental pond than the standard method, at a per sample cost over forty times lower. Our new, more sensitive assay provides a quantitative tool for eDNA-based monitoring of bigheaded carp, and the higher-yielding eDNA capture and extraction method we describe can be used for eDNA-based monitoring of any aquatic species.

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

Watrous, Matthew George; Adamic, Mary Louise; Olson, John Eric

The goal of the project, New Paradigms for Isotope Ratio Mass Spectrometry: Raising the Scientific Profile and Improved Performance for Accelerator Mass Spectrometry (AMS) and Thermal Ionization Mass Spectrometry (TIMS), is to ensure that the ongoing isotope ratio determination capability within the U.S. Department of Energy complex is the world’s best for application to nonproliferation. This report spells out the progress of Task 4, Transition of TIMS to AMS for Iodine Analysis, of the larger project. The subtasks under Task 4 and the accomplishments throughout the three year project life cycle are presented in this report. Progress was made inmore » optimization of chemical extraction, determination of a detection limit for 127Iodine, production of standard materials for AMS analysis quality assurance, facilitation of knowledge exchange with respect to analyzing iodine on an AMS, cross comparison with a world-leading AMS laboratory, supercritical fluid extraction of iodine for AMS analysis and electrodeposition of seawater as a direct method of preparation for iodine analysis by AMS--all with the goal of minimizing the time required to stand up an AMS capability for iodine analysis of exposed air filters at INL. An effective extraction method has been developed and demonstrated for iodine analysis of exposed air filters. Innovative techniques to accomplish the cathode preparation for AMS analysis were developed and demonstrated and published. The known gap of a lack of available materials for reference standards in the analysis of iodine by AMS was filled by the preparation of homogenous materials that were calibrated against NIST materials. A minimum limit on the amount of abundant isotope in a sample was determined for AMS analysis. The knowledge exchange occurred with fantastic success. Scientists engaged the international AMS community at conferences, as well as in their laboratories for collaborative work. The supercritical fluid extraction work has positive data, but is not a replacement for leaching yet. The added seawater work has led to a good method and a possible publication. The focus of this project was to minimize the time to stand up the AMS capability, by having all the preparation and supporting functions worked out ahead of the instrument arrival. Due to all the preparatory work and its success, the instrument was delivered and turned over to the INL in February 2015. Since then, INL scientists have been successfully vetting the capabilities and accomplishing their own measurements that agree well with the leading laboratories of the world for iodine analysis by AMS. Initially, all AMS data was gathered on other laboratories instruments, but during the last six months data for this project has come from INL’s AMS.« less

Feasibility of Isotope Harvesting at a Projectile Fragmentation Facility: 67Cu

DOE PAGES

Mastren, Tara; Pen, Aranh; Peaslee, Graham F.; ...

2014-10-21

The work presented here describes a proof-of-principle experiment for the chemical extraction of 67Cu from an aqueous beam stop at the National Superconducting Cyclotron Laboratory (NSCL). A 76 MeV/A 67Cu beam was stopped in water, successfully isolated from the aqueous solution through a series of chemical separations involving a chelating disk and anion exchange chromatography, then bound to NOTA-conjugated Herceptin antibodies, and the bound activity was validated using instant thin-layer chromatography (ITLC). The chemical extraction efficiency was found to be 88 ± 3% and the radiochemical yield was ≥95%. These results show that extraction of radioisotopes from an aqueous projectile-fragmentmore » beam dump is a feasible method for obtaining radiochemically pure isotopes.« less

The utility of measles and rubella IgM serology in an elimination setting, Ontario, Canada, 2009-2014.

PubMed

Bolotin, Shelly; Lim, Gillian; Dang, Vica; Crowcroft, Natasha; Gubbay, Jonathan; Mazzulli, Tony; Schabas, Richard

2017-01-01

In Canada, measles was eliminated in 1998 and rubella in 2000. Effective measles and rubella surveillance is vital in elimination settings, hinging on reliable laboratory methods. However, low-prevalence settings affect the predictive value of laboratory tests. We conducted an analysis to determine the performance of measles and rubella IgM testing in a jurisdiction where both infections are eliminated. 21,299 test results were extracted from the Public Health Ontario Laboratories database and 1,239 reports were extracted from the Ontario Integrated Public Health Information System (iPHIS) from 2008 and 2010 for measles and rubella, respectively, to 2014. Deterministic linkage resulted in 658 linked measles records (2009-2014) and 189 linked rubella records (2010-2014). Sixty-six iPHIS measles entries were classified as confirmed cases, of which 53 linked to laboratory data. Five iPHIS rubella entries were classified as confirmed, all linked to IgM results. The positive predictive value was 17.4% for measles and 3.6% for rubella. Sensitivity was 79.2% for measles and 100.0% for rubella. Specificity was 65.7% for measles and 25.8% for rubella. Our study confirms that a positive IgM alone does not confirm a measles case in elimination settings. This has important implications for countries that are working towards measles and rubella elimination.

Segmentation-less Digital Rock Physics

NASA Astrophysics Data System (ADS)

Tisato, N.; Ikeda, K.; Goldfarb, E. J.; Spikes, K. T.

2017-12-01

In the last decade, Digital Rock Physics (DRP) has become an avenue to investigate physical and mechanical properties of geomaterials. DRP offers the advantage of simulating laboratory experiments on numerical samples that are obtained from analytical methods. Potentially, DRP could allow sparing part of the time and resources that are allocated to perform complicated laboratory tests. Like classic laboratory tests, the goal of DRP is to estimate accurately physical properties of rocks like hydraulic permeability or elastic moduli. Nevertheless, the physical properties of samples imaged using micro-computed tomography (μCT) are estimated through segmentation of the μCT dataset. Segmentation proves to be a challenging and arbitrary procedure that typically leads to inaccurate estimates of physical properties. Here we present a novel technique to extract physical properties from a μCT dataset without the use of segmentation. We show examples in which we use segmentation-less method to simulate elastic wave propagation and pressure wave diffusion to estimate elastic properties and permeability, respectively. The proposed method takes advantage of effective medium theories and uses the density and the porosity that are measured in the laboratory to constrain the results. We discuss the results and highlight that segmentation-less DRP is more accurate than segmentation based DRP approaches and theoretical modeling for the studied rock. In conclusion, the segmentation-less approach here presented seems to be a promising method to improve accuracy and to ease the overall workflow of DRP.

Insights from a Modular Interdisciplinary Laboratory Course

NASA Astrophysics Data System (ADS)

Bouvier-Brown, N. C.; Carmona, V.

2016-12-01

A laboratory curriculum is naturally oriented towards "hands on" and problem-based learning. An earth science course, like chemical ecology, has the additional advantage of interdisciplinary education. Our Chemical Ecology course at Loyola Marymount University was structured as a small modular workshop. Students first gained hands-on experience with each analytical technique in the laboratory. The class then discussed these first datasets, delved into lecture topics, and tweaked their experimental procedures. Lastly, students were given time to design and execute their own experiments and present their findings. Three-to-four class periods were allotted for each laboratory topic. Detailed information and student reflections will be presented from the Spectroscopy module. Students used the Folin-Ciocalteau reagent method to extract the phenolic content of vegetation and/or soils. Because phenols are produced by plants for defense, this spectroscopic laboratory activity provided insight on allelopathy using analytical chemistry. Students were extremely engaged and learned not only the lab theory and technique, but also its application to our local ecology.

A comparison of DNA extraction protocols from blood spotted on FTA cards for the detection of tick-borne pathogens by Reverse Line Blot hybridization.

PubMed

Hailemariam, Zerihun; Ahmed, Jabbar Sabir; Clausen, Peter-Henning; Nijhof, Ard Menzo

2017-01-01

An essential step in the molecular detection of tick-borne pathogens (TBPs) in blood is the extraction of DNA. When cooled storage of blood under field conditions prior to DNA extraction in a dedicated laboratory is not possible, the storage of blood on filter paper forms a promising alternative. We evaluated six DNA extraction methods from blood spotted on FTA Classic ® cards (FTA cards), to determine the optimal protocol for the subsequent molecular detection of TBPs by PCR and the Reverse Line Blot hybridization assay (RLB). Ten-fold serial dilutions of bovine blood infected with Babesia bovis, Theileria mutans, Anaplasma marginale or Ehrlichia ruminantium were made by dilution with uninfected blood and spotted on FTA cards. Subsequently, DNA was extracted from FTA cards using six different DNA extraction protocols. DNA was also isolated from whole blood dilutions using a commercial kit. PCR/RLB results showed that washing of 3mm discs punched from FTA cards with FTA purification reagent followed by DNA extraction using Chelex ® resin was the most sensitive procedure. The detection limit could be improved when more discs were used as starting material for the DNA extraction, whereby the use of sixteen 3mm discs proved to be most practical. The presented best practice method for the extraction of DNA from blood spotted on FTA cards will facilitate epidemiological studies on TBPs. It may be particularly useful for field studies where a cold chain is absent. Copyright © 2016 Elsevier GmbH. All rights reserved.

EVALUATION OF ANTIBACTERIAL, ANTITUMOR, ANTIOXIDANT ACTIVITIES AND PHENOLIC CONSTITUENTS OF FIELD-GROWN AND IN VITRO-GROWN LYSIMACHIA VULGARIS L

PubMed Central

Yildirim, Arzu Birinci; Guner, Birgul; Karakas, Fatma Pehlivan; Turker, Arzu Ucar

2017-01-01

Background: Lysimachia vulgaris L. (Yellow loosestrife) is a medicinal plant in the family Myrsinaceae. It has been used in the treatment of fever, ulcer, diarrhea and wounds in folk medicine. It has also analgesic, expectorant, astringent and anti-inflammatory activities. Two different sources of the plant (field-grown and in vitro-grown) were used to evaluate the biological activities (antibacterial, antitumor and antioxidant) of L. vulgaris. In vitro-grown plant materials were collected from L. vulgaris plants that were previously regenerated in our laboratory. Materials and Methods: Plant materials were extracted with water, ethanol and acetone. For antibacterial test, disc diffusion method and 10 different pathogenic bacteria were used. Antioxidant activity was indicated by using DPPH method. The total phenol amount by using Folin-Ciocaltaeu method and the total flavonoid amount by using aluminum chloride (AlCl3) colorimetric method were determined. Results: Generally, yellow loosestrife extracts demonstrated antibacterial activity against Gram-positive bacteria (Staphylococcus aureus, S. epidermidis and Streptococcus pyogenes). Strong antitumor activity of yellow loosestrife was observed via potato disc diffusion bioassay. Nine different phenolics were also determined and compared by using High-Performance Liquid Chromatography (HPLC). Conclusion: Future investigations should be focused on fractionation of the extracts to identify active components for biological activity. PMID:28573234

Comprehensive automation of the solid phase extraction gas chromatographic mass spectrometric analysis (SPE-GC/MS) of opioids, cocaine, and metabolites from serum and other matrices.

PubMed

Lerch, Oliver; Temme, Oliver; Daldrup, Thomas

2014-07-01

The analysis of opioids, cocaine, and metabolites from blood serum is a routine task in forensic laboratories. Commonly, the employed methods include many manual or partly automated steps like protein precipitation, dilution, solid phase extraction, evaporation, and derivatization preceding a gas chromatography (GC)/mass spectrometry (MS) or liquid chromatography (LC)/MS analysis. In this study, a comprehensively automated method was developed from a validated, partly automated routine method. This was possible by replicating method parameters on the automated system. Only marginal optimization of parameters was necessary. The automation relying on an x-y-z robot after manual protein precipitation includes the solid phase extraction, evaporation of the eluate, derivatization (silylation with N-methyl-N-trimethylsilyltrifluoroacetamide, MSTFA), and injection into a GC/MS. A quantitative analysis of almost 170 authentic serum samples and more than 50 authentic samples of other matrices like urine, different tissues, and heart blood on cocaine, benzoylecgonine, methadone, morphine, codeine, 6-monoacetylmorphine, dihydrocodeine, and 7-aminoflunitrazepam was conducted with both methods proving that the analytical results are equivalent even near the limits of quantification (low ng/ml range). To our best knowledge, this application is the first one reported in the literature employing this sample preparation system.

Inter-laboratory validation of an inexpensive streamlined method to measure inorganic arsenic in rice grain.

PubMed

Chaney, Rufus L; Green, Carrie E; Lehotay, Steven J

2018-05-04

With the establishment by CODEX of a 200 ng/g limit of inorganic arsenic (iAs) in polished rice grain, more analyses of iAs will be necessary to ensure compliance in regulatory and trade applications, to assess quality control in commercial rice production, and to conduct research involving iAs in rice crops. Although analytical methods using high-performance liquid chromatography-inductively coupled plasma-mass spectrometry (HPLC-ICP-MS) have been demonstrated for full speciation of As, this expensive and time-consuming approach is excessive when regulations are based only on iAs. We report a streamlined sample preparation and analysis of iAs in powdered rice based on heated extraction with 0.28 M HNO 3 followed by hydride generation (HG) under control of acidity and other simple conditions. Analysis of iAs is then conducted using flow-injection HG and inexpensive ICP-atomic emission spectroscopy (AES) or other detection means. A key innovation compared with previous methods was to increase the acidity of the reagent solution with 4 M HCl (prior to reduction of As 5+ to As 3+ ), which minimized interferences from dimethylarsinic acid. An inter-laboratory method validation was conducted among 12 laboratories worldwide in the analysis of six shared blind duplicates and a NIST Standard Reference Material involving different types of rice and iAs levels. Also, four laboratories used the standard HPLC-ICP-MS method to analyze the samples. The results between the methods were not significantly different, and the Horwitz ratio averaged 0.52 for the new method, which meets official method validation criteria. Thus, the simpler, more versatile, and less expensive method may be used by laboratories for several purposes to accurately determine iAs in rice grain. Graphical abstract Comparison of iAs results from new and FDA methods.

Flavanol and procyanidin content (by degree of polymerization 1-10) of chocolate, cocoa liquors, cocoa powders, and cocoa extracts: first action 2012.24.

PubMed

Robbins, Rebecca J; Leonczak, Jadwiga; Li, Julia; Johnson, J Christopher; Collins, Tom; Kwik-Uribe, Catherine; Schmitz, Harold H

2013-01-01

An international collaborative study was conducted on an HPLC method with fluorescent detection for the determination of flavanols and procyanidins in chocolate and cocoa-containing materials. The sum of the oligomeric fractions with degree of polymerization 1-10 was the determined content value. Sample materials included dark and milk chocolates, cocoa powder, cocoa liquors, and cocoa extracts. The content ranged from approximately 2 to 500 mg/g (defatted basis). Thirteen laboratories--representing commercial, industrial, and academic institutions in six countries--participated in this interlaboratory study. Fourteen samples were sent as blind duplicates to the collaborators. Results for 12 laboratories yielded repeatability RSD (RSDr) values below 10% for all materials analyzed, ranging from 4.17 to 9.61%. Reproducibility RSD (RSDR) values ranged from 5.03 to 12.9% for samples containing 8.07 to 484.7 mg/g material analyzed. In one sample containing a low content of flavanols and procyanidins (approximately 2 mg/g), the RSDR was 17.68%.

On the problem of nonsense correlations in allergological tests after routine extraction.

PubMed

Rijckaert, G

1981-01-01

The influence of extraction procedures and culturing methods of material used for the preparation of allergenic extracts on correlation patterns found in allergological testing (skin test and RAST) was investigated. In our laboratory a short extraction procedure performed at O degrees C was used for Aspergillus repens. A. penicilloides, Wallemia sebi, their rearing media and non-inoculated medium. For the commercially available extracts from house dust, house-dust mite, pollen of Dactylus glomerata and A. penicilloides a longer procedure (several days) performed at room temperature was used. Statistical analysis showed a separation of all test results into two clusters, each cluster being composed of correlations between extracts from only one the manufacturers did not show any correlation. The correlations found between the short time incubated extracts of the xerophilic fungi and their rearing media could be explained by genetical and biochemical relationships between these fungi depending on ecological conditions. However, while the correlation found between house dust and house-dust mite is understandable, correlations found between long time incubated extracts from house-dust mite and D. glomerata or A. penicilloides may be nonsense correlations, that do not adequately describe the in vivo situation. The similarity of these extracts is presumably artificially created during extraction.

Remediation and recycling of WBP-treated lumber for use as flakeboard

Treesearch

Ronald Sabo; Jerrold E. Winandy; Carol A. Clausen; Altaf Basta

2008-01-01

Laboratory-scale experiments were conducted in which preservative metals (As, Cr, & Cu) were thermochemically extracted from CCA-treated spruce (Picea engelmannii) using oxalic acid and sodium hydroxide. The effects of extraction time, temperature, and pH were examined and laboratory scale optimization was achieved. Two series of experiments were carried out. In...

Clinical Performance of a Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Method for Detection of Certain blaKPC-Containing Plasmids

PubMed Central

Youn, Jung-Ho; Drake, Steven K.; Weingarten, Rebecca A.; Frank, Karen M.; Dekker, John P.

2015-01-01

Rapid detection of blaKPC-containing organisms can significantly impact infection control and clinical practices, as well as therapeutic choices. Current molecular and phenotypic methods to detect these organisms, however, require additional testing beyond routine organism identification. In this study, we evaluated the clinical performance of matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) to detect pKpQIL_p019 (p019)—an ∼11,109-Da protein associated with certain blaKPC-containing plasmids that was previously shown to successfully track a clonal outbreak of blaKPC-pKpQIL-Klebsiella pneumoniae in a proof-of-principle study (A. F. Lau, H. Wang, R. A. Weingarten, S. K. Drake, A. F. Suffredini, M. K. Garfield, Y. Chen, M. Gucek, J. H. Youn, F. Stock, H. Tso, J. DeLeo, J. J. Cimino, K. M. Frank, and J. P. Dekker, J Clin Microbiol 52:2804–2812, 2014, http://dx.doi.org/10.1128/JCM.00694-14). PCR for the p019 gene was used as the reference method. Here, blind analysis of 140 characterized Enterobacteriaceae isolates using two protein extraction methods (plate extraction and tube extraction) and two peak detection methods (manual and automated) showed sensitivities and specificities ranging from 96% to 100% and from 95% to 100%, respectively (2,520 spectra analyzed). Feasible laboratory implementation methods (plate extraction and automated analysis) demonstrated 96% sensitivity and 99% specificity. All p019-positive isolates (n = 26) contained blaKPC and were carbapenem resistant. Retrospective analysis of an additional 720 clinical Enterobacteriaceae spectra found an ∼11,109-Da signal in nine spectra (1.3%), including seven from p019-containing, carbapenem-resistant isolates (positive predictive value [PPV], 78%). Instrument tuning had a significant effect on assay sensitivity, highlighting important factors that must be considered as MALDI-TOF MS moves into applications beyond microbial identification. Using a large blind clinical data set, we have shown that spectra acquired for routine organism identification can also be analyzed automatically in real time at high throughput, at no additional expense to the laboratory, to enable rapid detection of potentially blaKPC-containing carbapenem-resistant isolates, providing early and clinically actionable results. PMID:26338858

Toxicity of some essential oils and plant extracts against Sitophilus oryzea, Aconthocelides obtectus and Rhizoperta dominica

NASA Astrophysics Data System (ADS)

Şimşek, Şeyda; Yaman, Cennet; Yarımoǧlu, Berkan; Yılmaz, Ayşegül

2017-04-01

In the present study, experiments were conducted to investigate fumigant toxicity of the essential oil from Myrtus sp plants for adult Aconthocelides obtectus, Sitophilus oryzea and Rhizoperta dominica in vitro conditions. The essential oils were isolated with the water distillation method by Neo-Clevenger apparatus. During the study 10% (v/v) doses of oils in 20 cc of compressed rubber-capped glass tubes were used. After 24 hours mortality rates of the essential oil were compared. Myrtus sp essential oil showed the highest fumigant toxicity on A. obtectus (46.66%). The lowest fumigant toxicity on S. oryzea (8.88%). The contact toxicity plant extracts (Prangos ferulacea, Alkanna orientalis, Myrtus communis) were tested against S. oryzea under laboratory conditions. Single dose contact toxicity effects of plant extracts were tested on S. oryzea adults via applying 1 µl extract suspension (10% w/v) to individual insect. The greatest contact toxicity to S. oryzea adults was observed with M. communis (43.33%) and A. orientalis (41.11%) extracts. P. ferulacea (34.44%) extracts produced moderate toxicity to S. oryzea adults.

Determination of Wastewater Compounds in Sediment and Soil by Pressurized Solvent Extraction, Solid-Phase Extraction, and Capillary-Column Gas Chromatography/Mass Spectrometry

USGS Publications Warehouse

Burkhardt, Mark R.; Zaugg, Steven D.; Smith, Steven G.; ReVello, Rhiannon C.

2006-01-01

A method for the determination of 61 compounds in environmental sediment and soil samples is described. The method was developed in response to increasing concern over the effects of endocrine-disrupting chemicals in wastewater and wastewater-impacted sediment on aquatic organisms. This method also may be used to evaluate the effects of combined sanitary and storm-sewer overflow on the water and sediment quality of urban streams. Method development focused on the determination of compounds that were chosen on the basis of their endocrine-disrupting potential or toxicity. These compounds include the alkylphenol ethoxylate nonionic surfactants and their degradates, food additives, fragrances, antioxidants, flame retardants, plasticizers, industrial solvents, disinfectants, fecal sterols, polycyclic aromatic hydrocarbons, and high-use domestic pesticides. Sediment and soil samples are extracted using a pressurized solvent extraction system. The compounds of interest are extracted from interfering matrix components by high-pressure water/isopropyl alcohol extraction. The compounds were isolated using disposable solid-phase extraction (SPE) cartridges containing chemically modified polystyrene-divinylbenzene resin. The cartridges were dried with nitrogen gas, and then sorbed compounds were eluted with methylene chloride (80 percent)-diethyl ether (20 percent) through Florisil/sodium sulfate SPE cartridge, and then determined by capillary-column gas chromatography/mass spectrometry. Recoveries in reagent-sand samples fortified at 4 to 72 micrograms averaged 76 percent ?13 percent relative standard deviation for all method compounds. Initial method reporting levels for single-component compounds ranged from 50 to 500 micrograms per kilogram. The concentrations of 20 out of 61 compounds initially will be reported as estimated with the 'E' remark code for one of three reasons: (1) unacceptably low-biased recovery (less than 60 percent) or highly variable method performance (greater than 25 percent relative standard deviation), (2) reference standards prepared from technical mixtures, or (3) potential blank contamination. Samples were preserved by freezing to -20 degrees Celsius. The U.S. Geological Survey National Water Quality Laboratory has established a 1-year sample-holding time limit (prior to sample extraction) from the date of sample collection (if the sample is kept at -20?C) until a statistically accepted method can be used to determine the effectiveness of the sample-freezing procedure.

Toxic reagents and expensive equipment: are they really necessary for the extraction of good quality fungal DNA?

PubMed

Rodrigues, P; Venâncio, A; Lima, N

2018-01-01

The aim of this work was to evaluate a fungal DNA extraction procedure with the lowest inputs in terms of time as well as of expensive and toxic chemicals, but able to consistently produce genomic DNA of good quality for PCR purposes. Two types of fungal biological material were tested - mycelium and conidia - combined with two protocols for DNA extraction using Sodium Dodecyl Sulphate (SDS) and Cetyl Trimethyl Ammonium Bromide as extraction buffers and glass beads for mechanical disruption of cell walls. Our results showed that conidia and SDS buffer was the combination that lead to the best DNA quality and yield, with the lowest variation between samples. This study clearly demonstrates that it is possible to obtain high yield and pure DNA from pigmented conidia without the use of strong cell disrupting procedures and of toxic reagents. There are numerous methods for DNA extraction from fungi. Some rely on expensive commercial kits and/or equipments, unavailable for many laboratories, or make use of toxic chemicals such as chloroform, phenol and mercaptoethanol. This study clearly demonstrates that it is possible to obtain high yields of pure DNA from pigmented conidia without the use of strong and expensive cell disrupting procedures and of toxic reagents. The method herein described is simultaneously inexpensive and adequate to DNA extraction from several different types of fungi. © 2017 The Society for Applied Microbiology.

Charged derivatization and on-line solid phase extraction to measure extremely low cortisol and cortisone levels in human saliva with liquid chromatography-tandem mass spectrometry.

PubMed

Magda, Balázs; Dobi, Zoltán; Mészáros, Katalin; Szabó, Éva; Márta, Zoltán; Imre, Tímea; Szabó, Pál T

2017-06-05

The aim of this study was to develop a sensitive, reliable and high-throughput liquid chromatography - electrospray ionization - mass spectrometric (LC-ESI-MS/MS) method for the simultaneous quantitation of cortisol and cortisone in human saliva. Derivatization with 2-hydrazino-1-methylpyridine (HMP) was one of the most challenging aspects of the method development. The reagent was reacting with cortisol and cortisone at 60°C within 1h, giving mono- and bis-hydrazone derivatives. Investigation of derivatization reaction and sample preparation was detailed and discussed. Improvement of method sensitivity was achieved with charged derivatization and use of on-line solid phase extraction (on-line SPE). The lower limit of quantitation (LLOQ) was 5 and 10pg/ml for cortisol and cortisone, respectively. The developed method was subsequently applied to clinical laboratory measurement of cortisol and cortisone in human saliva. Copyright © 2017 Elsevier B.V. All rights reserved.

Application of infrared reflection and Raman spectroscopy for quantitative determination of fat in potato chips

NASA Astrophysics Data System (ADS)

Mazurek, Sylwester; Szostak, Roman; Kita, Agnieszka

2016-12-01

Potato chips are important products in the snack industry. The most significant parameter monitored during their quality control process is fat content. The Soxhlet method, which is applied for this purpose, is time consuming and expensive. We demonstrate that both infrared and Raman spectroscopy can effectively replace the extraction method. Raman, mid-infrared (MIR) and near-infrared (NIR) spectra of the homogenised laboratory-prepared chips were recorded. On the basis of obtained spectra, partial least squares (PLS) calibration models were constructed. They were characterised by the values of relative standard errors of prediction (RSEP) in the 1.0-1.9% range for both calibration and validation data sets. Using the developed models, six commercial products were successfully quantified with recovery in the 98.5-102.3% range against the AOAC extraction method. The proposed method for fat quantification in potato chips based on Raman spectroscopy can be easily adopted for on-line product analysis.

Measurement Tools for the Immersive Visualization Environment: Steps Toward the Virtual Laboratory.

PubMed

Hagedorn, John G; Dunkers, Joy P; Satterfield, Steven G; Peskin, Adele P; Kelso, John T; Terrill, Judith E

2007-01-01

This paper describes a set of tools for performing measurements of objects in a virtual reality based immersive visualization environment. These tools enable the use of the immersive environment as an instrument for extracting quantitative information from data representations that hitherto had be used solely for qualitative examination. We provide, within the virtual environment, ways for the user to analyze and interact with the quantitative data generated. We describe results generated by these methods to obtain dimensional descriptors of tissue engineered medical products. We regard this toolbox as our first step in the implementation of a virtual measurement laboratory within an immersive visualization environment.

[Extraction of artemisinin and synthesis of its derivates artesunate and artemether].

PubMed

Chekem, L; Wierucki, S

2006-12-01

Artemisinin is extracted from Artemisia annua, a shrub also known as sweet wormwood that was used in traditional medicine in Asia for more than 1500 years. Recent studies in numerous malarious zones have demonstrated the effectiveness of artemisinin and have reported no evidence of the resistance now associated with almost all other antimalarials on the market. Despite its remarkable activity, artermisinin is not accessible to many patients due to high cost. This situation confronts all players in the fight against malaria with the urgent need to develop a simple process to produce massive supplies of artemisinin and its derivative at an affordable price. The purpose of the study described here was to develop a simple, cost-effective method that could be used by all professionals to extract artemisinin and transform it into artesunate or artemether. Artemisinin was extracted with dichloromethane and purified on the basis of variations in polarity and in the hydrophile/lipophile balance of solvents. Transformation into artesunate was a two-step process involving reduction to dihydroartemisinin using diisobutylaluminium hydride (DIBAL) followed by esterification using succinic anhydride. Artemether was obtained from dihydroartemisinin using boron trifluoride. Extraction using dichloromethane presents several advantages. Since dichloromethane is not explosive it can be safely transported and used for extraction on farms where Artemisia annua is grown. Evaporation and recovery of dichloromethane is relatively easy so that it can be re-used. These advantages result in a significant decrease in purchasing and shipping costs. Extraction on the farm eliminates the expense and facilities that would otherwise be required to transport and store leaves at the laboratory (250 kg of leaves yield 4 to 5 kg of raw artemisinin extract that yields approximately 1 kg of pure artemisinin). The low-cost process described here is feasible for any pharmaceutical laboratory including those in developing countries.

Role of Aqueous Extract of Morinda Citrifolia (Indian Noni) Ripe Fruits in Inhibiting Dental Caries-Causing Streptococcus Mutans and Streptococcus Mitis

PubMed Central

Kumarasamy, Barani; Manipal, Sunayana; Duraisamy, Prabu; Ahmed, Adil; Mohanaganesh, SP; Jeevika, C

2014-01-01

Objectives: Use of alternative medicine to control oral streptococci is a new topic worthy of further investigation. This study aimed to elucidate the dose-dependent anti-bacterial activity of crude aqueous extract of ripe Morinda citrifolia L. (Family: Rubiaceae) fruits against oral streptococci i.e. Streptococcus mutans and Streptococcus mitis, that cause dental caries in humans. Methods: Fresh ripe M. citrifolia fruits (750g) were ground in an electronic blender with sterile water (500ml). The crude aqueous extract was lyophilized to yield a brown colored powder. Various concentrations (1000-100μg/ ml) of the extract were tested for its antibacterial activity (Kirby and Bauer method) against whole cells of S. mutans and S. mitis. Minimum Inhibitory Concentration (MIC) was determined by micro-dilution method, using serially diluted (2 folds) fruit extract, according to the National Committee for Clinical Laboratory Standards (NCCLS). Results: Crude aqueous extract (1000μg/ ml) of ripe M. citrifolia fruits effectively inhibited the growth of S. mutans (19±0.5 mm) and S. mitis (18.6±0.3 mm) compared to the streptomycin control (21.6±0.3 mm). The growth inhibition was clearly evident with “nil” bacteriostasis, even after 48 hours of incubation at 37°C. The MIC of the extract for S. mutans and S. mitis was 125 μg and 62.5 μg, respectively. Conclusion: Our results suggest that phytochemicals naturally synthesized by M. citrifolia have an inhibitory effect on oral streptococci. Furthermore, purification and molecular characterization of the “bioactive principle” would enable us to formulate a sustainable oral hygiene product. PMID:25628701

Determination of antioxidant activity in foods and beverages by reaction with 2,2'-diphenyl-1-picrylhydrazyl (DPPH): collaborative study First Action 2012.04.

PubMed

Plank, David W; Szpylka, John; Sapirstein, Harry; Woollard, David; Zapf, Charles M; Lee, Vong; Chen, C Y Oliver; Liu, Rui Hai; Tsao, Rong; Düsterloh, André; Baugh, Steve

2012-01-01

A colorimetric method for the determination of total antioxidant activity in a variety of foods and beverages was validated in both a single-laboratory validation and a collaborative laboratory validation study. The procedure involved extraction of the antioxidants directly into a methanol-water solution containing a known amount of 2,2'-diphenyl-1-picrylhydrazyl (DPPH), thus promoting the rapid reaction of extracted materials with DPPH. The reaction was monitored by spectrophotometric measurement of the absorbance loss at 517 nm. Antioxidant activity was quantified relative to a dilution series of vitamin E analog standards (Trolox), which were analyzed in parallel simultaneously with the food and beverage samples. The antioxidant activities of the samples ranged from 131 to 131 000 micromole Trolox equivalents/100 g. Statistical analysis of the results showed that nine of the 11 matrixes gave acceptable HorRat values, indicating that the method performed well in these cases. The acceptable matrixes include pomegranate juice, blueberry juice, carrot juice, green tea, wine, rosemary spice, ready-to-eat cereal, and yogurt. Two samples failed the HorRat test: the first was an almond milk that had an antioxidant level below the practical LOQ for the method; the second was a sample of canola oil with added omega-3 fatty acid that was immiscible in the reaction medium.

Antimycoplasmal activity of some plant species from northern Nigeria compared to the currently used therapeutic agent.

PubMed

Muraina, I A; Adaudi, A O; Mamman, M; Kazeem, H M; Picard, J; McGaw, L J; Eloff, J N

2010-10-01

Mycoplasma spp. are obligate parasites of humans and animals. But due to the special requirements needed to culture Mycoplasma in the laboratory, little or no research has been done to evaluate the efficacy of medicinal plants on the organism. To screen medicinal plants traditionally used to treat infections for possible antimycoplasmal and cytotoxic activities. Acetone extracts of 21 Nigerian medicinal plants were analyzed for antimycoplasmal and cytotoxicity activities using the metabolic inhibition and colorimetric methods, respectively. The extract with the best antimycoplasmal activities was also analyzed for its phytochemical constituents using the desktop method. Calotropis procera (Aiton) R.Br (Asclepiadaceae) extract had the best antimycoplasmal effect with a minimum inhibitory concentration (MIC) of 80 µg/mL and minimum mycoplasmacidal concentration (MMC) of 160 µg/mL. This extract contained saponins, tannins, cardiac glycosides, alkaloids, and flavonoids. The extract of Vernonia amygdalina Delile (Compositae) was the most cytotoxic with median lethal concentration (LC(50)) of approximately 17 µg/mL, and that of Anacardium occidentale L. (Anacardiaceae) was the least cytotoxic with an LC(50) of approximately 1919 µg/mL. Calotropis procera is a promising plant for an alternative antimycoplasmal agent because the crude acetone extract had a higher mycoplasmacidal activity than the conventional drug tylosin, which is currently used in treatment of the disease in Nigeria. The crude extract of Calotropis procera is worth investigating for the development of a potent agent against cattle Mycoplasma, which has long defied solution by conventional chemotherapy.

Efficient screening method for determination of polycyclic aromatic hydrocarbons (PAHs) in airborne particles. Application in real samples of Santiago-Chile metropolitan urban area

NASA Astrophysics Data System (ADS)

Romero, Rodrigo; Sienra, Rosario; Richter, Pablo

A rapid analytical approach for determination of polycyclic aromatic hydrocarbons (PAHs) present in real samples of particulate matter (PM10 filters) was investigated, based on the use of water under sub critical conditions, and the subsequent determination by GC-MS (SIM). The method avoids the use of large volumes of organic solvents as dichloromethane, toluene or other unhealthy liquid organic mixtures which are normally used in time-consuming conventional sample preparation methods. By using leaching times <1 h, the method allows determination of PAHs in the range of ng/m 3 (detection limits between 0.05 and 0.2 ng/m 3 for 1458 m 3 of sampled air) with a precision expressed as RSD between 5.6% and 11.2%. The main idea behind this approach is to raise the temperature and pressure of water inside a miniaturized laboratory-made extraction unit and to decrease its dielectric constant from 80 to nearly 20. This effect allows an increase in the solubility of low polarity hydrocarbons such as PAHs. In this way, an extraction step of a few minutes can be sufficient for a quantitative extraction of airborne particles collected in high volume PM10 samplers. Parameters such as: extraction flow, static or dynamic extraction times and water volume were optimized by using a standard reference material. Technical details are given and a comparison using real samples is made between the conventional Soxhlet extraction method and the proposed approach. The proposed approach can be used as a quantitative method to characterize low molecular PAHs and simultaneously as a screening method for high molecular weight PAHs, because the recoveries are not quantitative for molecular weights over 202. In the specific case of the Santiago metropolitan area, due to the frequent occurrence of particulate matter during high pollution episodes, this approach was applied as an efficient short-time screening method for urban PAHs. Application of this screening method is recommended especially during the winter, when periods of clear detriment of the atmospheric and meteorological conditions occur in the area.

Results of a collaborative study on DNA identification of aged bone samples

PubMed Central

Vanek, Daniel; Budowle, Bruce; Dubska-Votrubova, Jitka; Ambers, Angie; Frolik, Jan; Pospisek, Martin; Al Afeefi, Ahmed Anwar; Al Hosani, Khalid Ismaeil; Allen, Marie; Al Naimi, Khudooma Saeed; Al Salafi, Dina; Al Tayyari, Wafa Ali Rashid; Arguetaa, Wendy; Bottinelli, Michel; Bus, Magdalena M.; Cemper-Kiesslich, Jan; Cepil, Olivier; De Cock, Greet; Desmyter, Stijn; El Amri, Hamid; El Ossmani, Hicham; Galdies, Ruth; Grün, Sebastian; Guidet, Francois; Hoefges, Anna; Iancu, Cristian Bogdan; Lotz, Petra; Maresca, Alessandro; Nagy, Marion; Novotny, Jindrich; Rachid, Hajar; Rothe, Jessica; Stenersen, Marguerethe; Stephenson, Mishel; Stevanovitch, Alain; Strien, Juliane; Sumita, Denilce R.; Vella, Joanna; Zander, Judith

2017-01-01

Aim A collaborative exercise with several institutes was organized by the Forensic DNA Service (FDNAS) and the Institute of the Legal Medicine, 2nd Faculty of Medicine, Charles University in Prague, Czech Republic, with the aim to test performance of different laboratories carrying out DNA analysis of relatively old bone samples. Methods Eighteen laboratories participating in the collaborative exercise were asked to perform DNA typing of two samples of bone powder. Two bone samples provided by the National Museum and the Institute of Archaelogy in Prague, Czech Republic, came from archeological excavations and were estimated to be approximately 150 and 400 years old. The methods of genetic characterization including autosomal, gonosomal, and mitochondrial markers was selected solely at the discretion of the participating laboratory. Results Although the participating laboratories used different extraction and amplification strategies, concordant results were obtained from the relatively intact 150 years old bone sample. Typing was more problematic with the analysis of the 400 years old bone sample due to poorer quality. Conclusion The laboratories performing identification DNA analysis of bone and teeth samples should regularly test their ability to correctly perform DNA-based identification on bone samples containing degraded DNA and potential inhibitors and demonstrate that risk of contamination is minimized. PMID:28613037

Critical points of DNA quantification by real-time PCR – effects of DNA extraction method and sample matrix on quantification of genetically modified organisms

PubMed Central

Cankar, Katarina; Štebih, Dejan; Dreo, Tanja; Žel, Jana; Gruden, Kristina

2006-01-01

Background Real-time PCR is the technique of choice for nucleic acid quantification. In the field of detection of genetically modified organisms (GMOs) quantification of biotech products may be required to fulfil legislative requirements. However, successful quantification depends crucially on the quality of the sample DNA analyzed. Methods for GMO detection are generally validated on certified reference materials that are in the form of powdered grain material, while detection in routine laboratories must be performed on a wide variety of sample matrixes. Due to food processing, the DNA in sample matrixes can be present in low amounts and also degraded. In addition, molecules of plant origin or from other sources that affect PCR amplification of samples will influence the reliability of the quantification. Further, the wide variety of sample matrixes presents a challenge for detection laboratories. The extraction method must ensure high yield and quality of the DNA obtained and must be carefully selected, since even components of DNA extraction solutions can influence PCR reactions. GMO quantification is based on a standard curve, therefore similarity of PCR efficiency for the sample and standard reference material is a prerequisite for exact quantification. Little information on the performance of real-time PCR on samples of different matrixes is available. Results Five commonly used DNA extraction techniques were compared and their suitability for quantitative analysis was assessed. The effect of sample matrix on nucleic acid quantification was assessed by comparing 4 maize and 4 soybean matrixes. In addition 205 maize and soybean samples from routine analysis were analyzed for PCR efficiency to assess variability of PCR performance within each sample matrix. Together with the amount of DNA needed for reliable quantification, PCR efficiency is the crucial parameter determining the reliability of quantitative results, therefore it was chosen as the primary criterion by which to evaluate the quality and performance on different matrixes and extraction techniques. The effect of PCR efficiency on the resulting GMO content is demonstrated. Conclusion The crucial influence of extraction technique and sample matrix properties on the results of GMO quantification is demonstrated. Appropriate extraction techniques for each matrix need to be determined to achieve accurate DNA quantification. Nevertheless, as it is shown that in the area of food and feed testing matrix with certain specificities is impossible to define strict quality controls need to be introduced to monitor PCR. The results of our study are also applicable to other fields of quantitative testing by real-time PCR. PMID:16907967

Characterization and Testing of Amidoxime-Based Adsorbent Materials to Extract Uranium from Natural Seawater

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

Kuo, Li-Jung; Janke, Christopher J.; Wood, Jordana R.

Extraction of uranium (U) from seawater for use as a nuclear fuel is a significant challenge due to the low concentration of U in seawater (~3.3 ppb) and difficulties to selectively extract U from the background of major and trace elements in seawater. The Pacific Northwest National Laboratory (PNNL)’s Marine Sciences Laboratory (MSL) has been serving as a marine test site for determining performance characteristics (adsorption capacity, adsorption kinetics, and selectivity) of novel amidoxime-based polymeric adsorbents developed at Oak Ridge National Laboratory (ORNL) under natural seawater exposure conditions. This manuscript describes the performance of three formulations (38H, AF1, AI8) ofmore » amidoxime-based polymeric adsorbents produced at ORNL in MSL’s ambient seawater testing facility. The adsorbents were produced in two forms, fibrous material (40-100 mg samples) and braided material (5-10 g samples), and exposed to natural seawater using flow-through columns and recirculating flumes. All three formulations demonstrated high 56 day uranium adsorption capacity (>3 g U/kg adsorbent). The AF1 formulation had the best uranium adsorption performance, with a 56 day capacity of 3.9 g U/ kg adsorbent, a saturation capacity of 5.4 g U/kg adsorbent, and ~25 days half-saturation time. The two exposure methods, flowthrough columns and flumes, were demonstrated to produce similar performance results, providing confidence that the test methods were reliable, that scaling up from 10’s of mg quantities of exposure in flow-through columns to gram quantities in flumes produced similar results, and confirm that the manufacturing process produces a homogeneous adsorbent. Adsorption kinetics appear to be element specific, with half-saturation times ranging from minutes for the major cations in seawater, to 8-10 weeks for V and Fe. Reducing the exposure time provides a potential pathway to improve the adsorption capacity of U by reducing the V/U ratio on the adsorbent.« less

Characterization and testing of amidoxime-based adsorbent materials to extract uranium from natural seawater

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

Kuo, Li-Jung; Janke, Christopher James; Wood, Jordana

Extraction of uranium (U) from seawater for use as a nuclear fuel is a significant challenge due to the low concentration of U in seawater (~3.3 ppb) and difficulties to selectively extract U from the background of major and trace elements in seawater. The Pacific Northwest National Laboratory (PNNL) s Marine Sciences Laboratory (MSL) has been serving as a marine test site for determining performance characteristics (adsorption capacity, adsorption kinetics, and selectivity) of novel amidoxime-based polymeric adsorbents developed at Oak Ridge National Laboratory (ORNL) under natural seawater exposure conditions. This report describes the performance of three formulations (38H, AF1, AI8)more » of amidoxime-based polymeric adsorbent produced at ORNL in MSL s ambient seawater testing facility. The adsorbents were produced in two forms, fibrous material (40-100 mg samples) and braided material (5-10 g samples), exposed to natural seawater using flow-through columns and recirculating flumes. All three formulations demonstrated high 56 day uranium adsorption capacity (>3 gU/kg adsorbent). The AF1 formulation had the best uranium adsorption performance, with 56-day capacity of 3.9 g U/kg adsorbent, saturation capacity of 5.4 g U/kg adsorbent, and ~25 days half-saturation time. The two exposure methods, flow-through columns and flumes were demonstrated to produce similar performance results, providing confidence that the test methods were reliable, that scaling up from 10 s of mg quantities of exposure in flow-through columns to gram quantities in flumes produced similar results, and that the manufacturing process produces a homogenous adsorbent. Adsorption kinetics appear to be element specific, with half-saturation times ranging from minutes for the major cations in seawater to 8-10weeks for V and Fe. Reducing the exposure time provides a potential pathway to improve the adsorption capacity of U by reducing the V/U ratio on the adsorbent.« less

Solid-phase extraction combined with high-performance liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry analysis of pesticides in water: method performance and application in a reconnaissance survey of residues in drinking water in Greater Cairo, Egypt.

PubMed

Potter, Thomas L; Mohamed, Mahmoud A; Ali, Hannah

2007-01-24

Monitoring of water resources for pesticide residues is often needed to ensure that pesticide use does not adversely impact the quality of public water supplies or the environment. In many rural areas and throughout much of the developing world, monitoring is often constrained by lack of testing facilities; thus, collection of samples and shipment to centralized laboratories for analysis is required. The portability, ease of use, and potential to enhance analyte stability make solid-phase extraction (SPE) an attractive technique for handling water samples prior to their shipment. We describe performance of an SPE method targeting a structurally diverse mixture of 25 current-use pesticides and two common degradates in samples of raw and filtered drinking water collected in Greater Cairo, Egypt. SPE was completed in a field laboratory in Egypt, and cartridges were shipped to the United States for elution and high-performance liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry analysis. Quantitative and reproducible recovery of 23 of 27 compounds (average = 96%; percent relative standard deviation = 21%) from matrix spikes (1 microg L-1 per component) prepared in the field and from deionized water fortified similarly in the analytical laboratory was obtained. Concurrent analysis of unspiked samples identified four parent compounds and one degradate in drinking water samples. No significant differences were observed between raw and filtered samples. Residue levels in all cases were below drinking water and "harm to aquatic-life" thresholds, indicating that human and ecological risks of pesticide contamination were relatively small; however, the study was limited in scale and scope. Further monitoring is needed to define spatial and temporal variation in residue concentrations. The study has demonstrated the feasibility of performing studies of this type using SPE to extract and preserve samples in the field. The approach should be broadly applicable in many settings.

Characterization and testing of amidoxime-based adsorbent materials to extract uranium from natural seawater

DOE PAGES

Kuo, Li-Jung; Janke, Christopher James; Wood, Jordana; ...

2015-11-19

Extraction of uranium (U) from seawater for use as a nuclear fuel is a significant challenge due to the low concentration of U in seawater (~3.3 ppb) and difficulties to selectively extract U from the background of major and trace elements in seawater. The Pacific Northwest National Laboratory (PNNL) s Marine Sciences Laboratory (MSL) has been serving as a marine test site for determining performance characteristics (adsorption capacity, adsorption kinetics, and selectivity) of novel amidoxime-based polymeric adsorbents developed at Oak Ridge National Laboratory (ORNL) under natural seawater exposure conditions. This report describes the performance of three formulations (38H, AF1, AI8)more » of amidoxime-based polymeric adsorbent produced at ORNL in MSL s ambient seawater testing facility. The adsorbents were produced in two forms, fibrous material (40-100 mg samples) and braided material (5-10 g samples), exposed to natural seawater using flow-through columns and recirculating flumes. All three formulations demonstrated high 56 day uranium adsorption capacity (>3 gU/kg adsorbent). The AF1 formulation had the best uranium adsorption performance, with 56-day capacity of 3.9 g U/kg adsorbent, saturation capacity of 5.4 g U/kg adsorbent, and ~25 days half-saturation time. The two exposure methods, flow-through columns and flumes were demonstrated to produce similar performance results, providing confidence that the test methods were reliable, that scaling up from 10 s of mg quantities of exposure in flow-through columns to gram quantities in flumes produced similar results, and that the manufacturing process produces a homogenous adsorbent. Adsorption kinetics appear to be element specific, with half-saturation times ranging from minutes for the major cations in seawater to 8-10weeks for V and Fe. Reducing the exposure time provides a potential pathway to improve the adsorption capacity of U by reducing the V/U ratio on the adsorbent.« less

Improved sample preparation to determine acrylamide in difficult matrixes such as chocolate powder, cocoa, and coffee by liquid chromatography tandem mass spectroscopy.

PubMed

Delatour, Thierry; Périsset, Adrienne; Goldmann, Till; Riediker, Sonja; Stadler, Richard H

2004-07-28

An improved sample preparation (extraction and cleanup) is presented that enables the quantification of low levels of acrylamide in difficult matrixes, including soluble chocolate powder, cocoa, coffee, and coffee surrogate. Final analysis is done by isotope-dilution liquid chromatography-electrospray ionization tandem mass spectrometry (LC-MS/MS) using d3-acrylamide as internal standard. Sample pretreatment essentially encompasses (a) protein precipitation with Carrez I and II solutions, (b) extraction of the analyte into ethyl acetate, and (c) solid-phase extraction on a Multimode cartridge. The stability of acrylamide in final extracts and in certain commercial foods and beverages is also reported. This approach provided good performance in terms of linearity, accuracy and precision. Full validation was conducted in soluble chocolate powder, achieving a decision limit (CCalpha) and detection capability (CCbeta) of 9.2 and 12.5 microg/kg, respectively. The method was extended to the analysis of acrylamide in various foodstuffs such as mashed potatoes, crisp bread, and butter biscuit and cookies. Furthermore, the accuracy of the method is demonstrated by the results obtained in three inter-laboratory proficiency tests. Copyright 2004 American Chemical Society

Distribution of naphthenic acids in tissues of laboratory-exposed fish and in wild fishes from near the Athabasca oil sands in Alberta, Canada.

PubMed

Young, Rozlyn F; Michel, Lorelei Martínez; Fedorak, Phillip M

2011-05-01

Naphthenic acids, which have a variety of commercial applications, occur naturally in conventional crude oil and in highly biodegraded petroleum such as that found in the Athabasca oil sands in Alberta, Canada. Oil sands extraction is done using a caustic aqueous extraction process. The alkaline pH releases the naphthenic acids from the oil sands and dissolves them into water as their soluble naphthenate forms, which are anionic surfactants. These aqueous extracts contain concentrations of naphthenates that are acutely lethal to fishes and other aquatic organisms. Previous research has shown that naphthenic acids can be taken up by fish, but the distribution of these acids in various tissues of the fish has not been determined. In this study, rainbow trout (Oncorhynchus mykiss) were exposed to commercial (Merichem) naphthenic acids in the laboratory. After a 10-d exposure to approximately 3mg naphthenic acids/L, the fish were dissected and samples of gills, heart, liver, kidney, muscle, and eggs were extracted and analyzed for free (unconjugated) naphthenic acids by a gas chromatography-mass spectrometry method. Each of the tissues contained naphthenic acids and non-parametric statistical analyses showed that gills and livers contained higher concentrations than the muscles and that the livers had higher concentrations than the hearts. Four different species of fish (two fish of each species) were collected from the Athabasca River near two oil sands mining and extraction operations. No free naphthenic acids were detected in the muscle or liver of these fish. Copyright © 2010 Elsevier Inc. All rights reserved.

A single method for detecting 11 organophosphate pesticides in human plasma and breastmilk using GC-FPD

PubMed Central

Naksen, Warangkana; Prapamontol, Tippawan; Mangklabruks, Ampica; Chantara, Somporn; Thavornyutikarn, Prasak; Robson, Mark G.; Ryan, P. Barry; Barr, Dana Boyd; Panuwet, Parinya

2016-01-01

Organophosphate (OP) pesticides are widely used for crop protection in many countries including Thailand. Aside from causing environmental contamination, they affect human health especially by over-stimulating of the neurotransmission system. OP pesticides, as with other non-persistent pesticides, degrade quickly in the environment as well as are metabolized quite rapidly in humans. Assessing human exposures to these compounds requires analytical methods that are sensitive, robust, and most importantly, suitable for specific laboratory settings. The aim of this study was to develop and validate an analytical method for measuring 11 OP pesticide residues in human plasma and breast milk. Analytes in both plasma and breast milk samples were extracted with acetone and methylene chloride, cleaned-up using aminopropyl solid phase extraction cartridges, and analyzed by gas chromatography with flame photometric detection. The optimized method exhibited good linearity, with the coefficients of determination of 0.996–0.999 and <7% error about the slope. Extraction recoveries from spiked plasma and breast milk samples at low and medium concentrations (0.8–5.0 and 1.6–10 ng mL−1, respectively) ranged from 59.4 % (ethion) to 94.0 % (chlorpyrifos). Intra-batch and inter-batch precisions ranged from 2.3–18.9% and 5.8–19.5%, respectively. Method detection limits of plasma and breast milk ranged from 0.18–1.36 and 0.09–2.66 ng mL−1, respectively. We analyzed 63 plasma and 30 breastmilk samples collected from farmworkers in Chiang Mai Province to determine the suitability of this method for occupational exposure assessment. Of the 11 pesticides measured, seven were detected in plasma samples and five were detected in breast milk samples. Mass spectrometry was used to confirm results. Overall, this method is rapid and reliable. It offers the laboratories with limited access to mass spectrometry a capacity to investigate levels OP pesticides in plasma and breastmilk in those occupationally exposed for health risk assessment. PMID:27232054

[The isolation of organic compounds from hydrosulfuric mineral waters with the use of the extractive freezing-out technique with centrifugation].

PubMed

Bekhterev, V N; Kabina, E A

The mineral waters, enriched with organic substances find extensive application in balneotherapy. The fast and efficient methods for the identification and quantitative measurement of organic compounds (in the first place, organic acids) in such waters need to be developed for the estimation of their quality and biological activity. The objective of the present study was to elaborate a gas chromatographic method for the determination of monobasic carbonic acids in sulfide-containing mineral waters by means of extractive freezing-out in combination with the application of the centrifugal forces for the elucidation of the metrological characteristics of the compounds of interest. The secondary objective was to estimate the prospects for the application of the method of interest for determining the dissolved organic compounds in mineral waters. The following carbonic acids were used for the purposes of the study: acetic acid (analytical grade), Russia; propionic grade (extra pure), Ferak, Germany; butyric acid (pure), Russia; valeric acid (pure), Russia; caproic acid (pure), Russia; oenanthic acid (pure), Russia; and caprylic acid (pure), Russia). Acetonitrile («O» grade), Russia, was used as the extracting agent. The LV-210 analytical balance (Russia) was used to prepare the model and standard solutions of the organic compounds and to determine their mass. The extracts and standard mixture were investigated by the gas chromatographic technique with the use of the Kristallyuks apparatus («Meta-Khrom», Russia) equipped with the flame ionization detector and the capillary column. Extractive freezing-out in the combination with centrifugation was performed with the laboratory installation for this purpose. Under the model conditions, a single extractive freezing-out procedure with the centrifugation of the sample made it possible to reach the 22-37-fold concentration of C2-C8 monobasic carbonic acids during their transfer from water into acetonitrile. The metrological parameters of the proposed procedure for gas-chromatographic determination of the aforementioned acids in hydrosulfuric mineral waters were estimated. The limit of detection of acetic and propionic acids has been 0,2 mg/l, that for butyric acid 0,1 mg/l, for valeric, caproic, oenanthic and caprylic acids 0,05 mg/l. We failed to identify C2-C8 carbonic acids in hydrosulfuric water from the 6T well of the field Matsesta fields. The investigation of the extracts obtained by the proposed extraction procedure with the use of the newly developed variant of high performance liquid chromatography (HPLC) with diode-matrix detection revealed the presence of various amounts of unknown organic substances. The UV-spectrum of one of those components was obtained which made it possible to conjecture its molecular structure. The procedure for the sample preparation and the following determination of organic acids in water is simple, carried out in a single stage, and does not impose special requirements to the qualification of the researcher. It allows to describe the newly developed technique as a rapid test method. Along with it, realization of extraction in the negative temperature mode reduces the risk of decomposition of the thermally unstable chemical compounds being extracted and reduces volatility of the organic solvents being used. The technology of the method does not demand a special laboratory glassware and expendables.

Fitting It All In: Adapting a Green Chemistry Extraction Experiment for Inclusion in an Undergraduate Analytical Laboratory

ERIC Educational Resources Information Center

Buckley, Heather L.; Beck, Annelise R.; Mulvihill, Martin J.; Douskey, Michelle C.

2013-01-01

Several principles of green chemistry are introduced through this experiment designed for use in the undergraduate analytical chemistry laboratory. An established experiment of liquid CO2 extraction of D-limonene has been adapted to include a quantitative analysis by gas chromatography. This facilitates drop-in incorporation of an exciting…

Single-laboratory validation of a high-performance liquid chromatographic-diode array detector-fluorescence detector/mass spectrometric method for simultaneous determination of water-soluble vitamins in multivitamin dietary tablets.

PubMed

Chen, Pei; Atkinson, Renata; Wolf, Wayne R

2009-01-01

The purpose of this study was to develop a single-laboratory validated (SLV) method using high-performance liquid chromatography with different detectors [diode array detector (DAD); fluorescence detector (FLD); and mass spectrometry (MS)] for determination of 7 B-complex vitamins (B1-thiamin, B2-riboflavin, B3-nicotinamide, B6-pyridoxine, B9-folic acid, pantothenic acid, and biotin) and vitamin C in multivitamin/multimineral dietary supplements. The method involves the use of a reversed-phase octadecylsilyl column (4 microm, 250 x 2.0 mm id) and a gradient mobile phase profile. Gradient elution was performed at a flow rate of 0.25 mL/min. After a 5 min isocratic elution at 100% A (0.1% formic acid in water), a linear gradient to 50% A and 50% B (0.1% formic acid in acetonitrile) at 15 min was employed. Detection was performed with a DAD as well as either an FLD or a triple-quadrupole MS detector in the multiple reaction monitoring mode. SLV was performed using Standard Reference Material (SRM) 3280 Multivitamin/Multimineral Tablets, being developed by the National Institute of Standards and Technology, with support by the Office of Dietary Supplements of the National Institutes of Health. Phosphate buffer (10 mM, pH 2.0) extracts of the NIST SRM 3280 were analyzed by the liquid chromatographic (LC)-DAD-FLDIMS method. Following extraction, the method does not require any sample cleanup/preconcentration steps except centrifugation and filtration.

Single-Laboratory Validation of a High-Performance Liquid Chromatographic-Diode Array Detector-Fluorescence Detector/Mass Spectrometric Method for Simultaneous Determination of Water-Soluble Vitamins in Multivitamin Dietary Tablets

PubMed Central

Chen, Pei; Atkinson, Renata; Wolf, Wayne R.

2014-01-01

The purpose of this study was to develop a single-laboratory validated (SLV) method using high-performance liquid chromatography with different detectors [diode array detector (DAD); fluorescence detector (FLD); and mass spectrometry (MS)] for determination of 7 B-complex vitamins (B1-thiamin, B2-riboflavin, B3-nicotinamide, B6-pyridoxine, B9-folic acid, pantothenic acid, and biotin) and vitamin C in multivitamin/multimineral dietary supplements. The method involves the use of a reversed-phase octadecylsilyl column (4 µm, 250 × 2.0 mm id) and a gradient mobile phase profile. Gradient elution was performed at a flow rate of 0.25 mL/min. After a 5 min isocratic elution at 100% A (0.1% formic acid in water), a linear gradient to 50% A and 50% B (0.1% formic acid in acetonitrile) at 15 min was employed. Detection was performed with a DAD as well as either an FLD or a triple-quadrupole MS detector in the multiple reaction monitoring mode. SLV was performed using Standard Reference Material (SRM) 3280 Multivitamin/Multimineral Tablets, being developed by the National Institute of Standards and Technology, with support by the Office of Dietary Supplements of the National Institutes of Health. Phosphate buffer (10 mM, pH 2.0) extracts of the NIST SRM 3280 were analyzed by the liquid chromatographic (LC)-DAD-FLD/MS method. Following extraction, the method does not require any sample cleanup/preconcentration steps except centrifugation and filtration. PMID:19485230

An exploratory clustering approach for extracting stride parameters from tracking collars on free-ranging wild animals.

PubMed

Dewhirst, Oliver P; Roskilly, Kyle; Hubel, Tatjana Y; Jordan, Neil R; Golabek, Krystyna A; McNutt, J Weldon; Wilson, Alan M

2017-02-01

Changes in stride frequency and length with speed are key parameters in animal locomotion research. They are commonly measured in a laboratory on a treadmill or by filming trained captive animals. Here, we show that a clustering approach can be used to extract these variables from data collected by a tracking collar containing a GPS module and tri-axis accelerometers and gyroscopes. The method enables stride parameters to be measured during free-ranging locomotion in natural habitats. As it does not require labelled data, it is particularly suitable for use with difficult to observe animals. The method was tested on large data sets collected from collars on free-ranging lions and African wild dogs and validated using a domestic dog. © 2017. Published by The Company of Biologists Ltd.

Repellent properties of Cardiospermum halicacabum Linn. (Family: Sapindaceae) plant leaf extracts against three important vector mosquitoes

PubMed Central

Govindarajan, M; Sivakumar, R

2012-01-01

Objective To determine repellent activity of hexane, ethyl acetate, benzene, chloroform and methanol extract of Cardiospermum halicacabum (C. halicacabum) against Culex quinquefasciatus (Cx. quinquefasciatus), Aedes aegypti (Ae. aegypti) and Anopheles stephensi (An. stephensi). Methods Evaluation was carried out in a net cage (45 cm×30 cm×25 cm) containing 100 blood starved female mosquitoes of three mosquito species and were assayed in the laboratory condition by using the protocol of WHO 2005; The plant leaf crude extracts of C. halicacabum was applied at 1.0, 2.5, and 5.0 mg/cm2 separately in the exposed area of the fore arm. Only ethanol served as control. Results In this observation, the plant crude extracts gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity was dependent on the strength of the plant extracts. The tested plant crude extracts had exerted promising repellent against all the three mosquitoes. Conclusions From the results it can be concluded the crude extract of C. halicacabum was potential for controlling Cx. quinquefasciatus, Ae. aegypti and An. stephensi mosquitoes. PMID:23569979

Analytical methods for determination of mycotoxins: a review.

PubMed

Turner, Nicholas W; Subrahmanyam, Sreenath; Piletsky, Sergey A

2009-01-26

Mycotoxins are small (MW approximately 700), toxic chemical products formed as secondary metabolites by a few fungal species that readily colonise crops and contaminate them with toxins in the field or after harvest. Ochratoxins and Aflatoxins are mycotoxins of major significance and hence there has been significant research on broad range of analytical and detection techniques that could be useful and practical. Due to the variety of structures of these toxins, it is impossible to use one standard technique for analysis and/or detection. Practical requirements for high-sensitivity analysis and the need for a specialist laboratory setting create challenges for routine analysis. Several existing analytical techniques, which offer flexible and broad-based methods of analysis and in some cases detection, have been discussed in this manuscript. There are a number of methods used, of which many are lab-based, but to our knowledge there seems to be no single technique that stands out above the rest, although analytical liquid chromatography, commonly linked with mass spectroscopy is likely to be popular. This review manuscript discusses (a) sample pre-treatment methods such as liquid-liquid extraction (LLE), supercritical fluid extraction (SFE), solid phase extraction (SPE), (b) separation methods such as (TLC), high performance liquid chromatography (HPLC), gas chromatography (GC), and capillary electrophoresis (CE) and (c) others such as ELISA. Further currents trends, advantages and disadvantages and future prospects of these methods have been discussed.

Guidance for laboratories performing molecular pathology for cancer patients

PubMed Central

Cree, Ian A; Deans, Zandra; Ligtenberg, Marjolijn J L; Normanno, Nicola; Edsjö, Anders; Rouleau, Etienne; Solé, Francesc; Thunnissen, Erik; Timens, Wim; Schuuring, Ed; Dequeker, Elisabeth; Murray, Samuel; Dietel, Manfred; Groenen, Patricia; Van Krieken, J Han

2014-01-01

Molecular testing is becoming an important part of the diagnosis of any patient with cancer. The challenge to laboratories is to meet this need, using reliable methods and processes to ensure that patients receive a timely and accurate report on which their treatment will be based. The aim of this paper is to provide minimum requirements for the management of molecular pathology laboratories. This general guidance should be augmented by the specific guidance available for different tumour types and tests. Preanalytical considerations are important, and careful consideration of the way in which specimens are obtained and reach the laboratory is necessary. Sample receipt and handling follow standard operating procedures, but some alterations may be necessary if molecular testing is to be performed, for instance to control tissue fixation. DNA and RNA extraction can be standardised and should be checked for quality and quantity of output on a regular basis. The choice of analytical method(s) depends on clinical requirements, desired turnaround time, and expertise available. Internal quality control, regular internal audit of the whole testing process, laboratory accreditation, and continual participation in external quality assessment schemes are prerequisites for delivery of a reliable service. A molecular pathology report should accurately convey the information the clinician needs to treat the patient with sufficient information to allow for correct interpretation of the result. Molecular pathology is developing rapidly, and further detailed evidence-based recommendations are required for many of the topics covered here. PMID:25012948

Molluscicidal activity of Physalis angulata L. extracts and fractions on Biomphalaria tenagophila (d'Orbigny, 1835) under laboratory conditions.

PubMed

dos Santos, José Augusto A; Tomassini, Therezinha Coelho B; Xavier, Deise Cristina Drummond; Ribeiro, Ivone Maria; da Silva, Melissa Teixeira G; de Morais Filho, Zenildo Buarque

2003-04-01

The main objective of this research is to evaluate the molluscicide activity of Physalis angulata L. Biomphalaria tenagophila specimens under laboratory conditions. Extracts and fractions were supplied by the Laborat rio de Qu mica de Produtos Naturais, Farmanguinhos-Fiocruz. Experiments were performed according to the methodology described by the World Health Organization for molluscicide tests using the concentrations from 0.1 to 500 mg/l of the extracts, fractions and of a pool of physalins modified steroids present in this species. The results show that ethyl acetate and acetone extracts from the whole plant, the ethanolic extracts of the roots and the physalins pool from stems and leaves were active. Only the whole plant extracts were available in sufficient quantity for the determination of LD50 and LD90 values.

Determination of lipophilic toxins by LC/MS/MS: single-laboratory validation.

PubMed

Villar-González, Adriano; Rodríguez-Velasco, María Luisa; Gago-Martínez, Ana

2011-01-01

An LC/MS/MS method has been developed, assessed, and intralaboratory-validated for the analysis of the lipophilic toxins currently regulated by European Union legislation: okadaic acid (OA) and dinophysistoxins 1 and 2, including their ester forms; azaspiracids 1, 2, and 3; pectenotoxins 1 and 2; yessotoxin (YTX), and the analogs 45 OH-YTX, Homo YTX, and 45 OH-Homo YTX; as well as for the analysis of 13-desmetil-spirolide C. The method consists of duplicate sample extraction with methanol and direct analysis of the crude extract without further cleanup or concentration. Ester forms of OA and dinophysistoxins are detected as the parent ions after alkaline hydrolysis of the extract. The validation process of this method was performed using both fortified and naturally contaminated samples, and experiments were designed according to International Organization for Standardization, International Union of Pure and Applied Chemistry, and AOAC guidelines. With the exception of YTX in fortified samples, RSDr below 15% and RSDR were below 25%. Recovery values were between 77 and 95%, and LOQs were below 60 microg/kg. These data together with validation experiments for recovery, selectivity, robustness, traceability, and linearity, as well as uncertainty calculations, are presented in this paper.

Comparison of manual methods of extracting genomic DNA from dried blood spots collected on different cards: implications for clinical practice.

PubMed

Molteni, C G; Terranova, L; Zampiero, A; Galeone, C; Principi, N; Esposito, S

2013-01-01

Isolating genomic DNA from blood samples is essential when studying the associations between genetic variants and susceptibility to a given clinical condition, or its severity. This study of three extraction techniques and two types of commercially available cards involved 219 children attending our outpatient pediatric clinic for follow-up laboratory tests after they had been hospitalised. An aliquot of venous blood was drawn into plastic tubes without additives and, after several inversions, 80 microL were put on circles of common paper cards and Whatman FTA-treated cards. Three extraction methods were compared: the Qiagen Investigator, Gensolve, and Masterpure. The best method in terms of final DNA yield was Masterpure, which led to a significantly higher yield regardless of the type of card (p less than 0.001), followed by Qiagen Investigator and Gensolve. Masterpure was also the best in terms of price, seemed to be simple and reliable, and required less hands-on time than other techniques. These conclusions support the use of Masterpure in studies that evaluate the associations between genetic variants and the severity or prevalence of infectious diseases.

An alternative method for analysis of food taints using stir bar sorptive extraction.

PubMed

Ridgway, Kathy; Lalljie, Sam P D; Smith, Roger M

2010-09-10

The determination of taints in food products currently can involve the use of several sample extraction techniques, including direct headspace (DHS), steam distillation extraction (SDE) and more recently solid phase microextraction (SPME). Each of these techniques has disadvantages, such as the use of large volumes of solvents (SDE), or limitations in sensitivity (DHS), or have only been applied to date for determination of individual or specific groups of tainting compounds (SPME). The use of stir bar sorptive extraction (SBSE) has been evaluated as a quantitative screening method for unknown tainting compounds in foods. A range of commonly investigated problem compounds, with a range of physical and chemical properties, were examined. The method was optimised to give the best response for the majority of compounds and the performance was evaluated by examining the accuracy, precision, linearity, limits of detection and quantitation and uncertainties for each analyte. For most compounds SBSE gave the lowest limits of detection compared to steam distillation extraction or direct headspace analysis and in general was better than these established techniques. However, for methyl methacrylate and hexanal no response was observed following stir bar extraction under the optimised conditions. The assays were carried out using a single quadrupole GC-MS in scan mode. A comparison of acquisition modes and instrumentation was performed using standards to illustrate the increase in sensitivity possible using more targeted ion monitoring or a more sensitive high resolution mass spectrometer. This comparison illustrated the usefulness of this approach as an alternative to specialised glassware or expensive instrumentation. SBSE in particular offers a 'greener' extraction method by a large reduction in the use of organic solvents and also minimises the potential for contamination from external laboratory sources, which is of particular concern for taint analysis. Copyright © 2010 Elsevier B.V. All rights reserved.

Evaluation of coral pathogen growth rates after exposure to atmospheric African dust samples

USGS Publications Warehouse

Lisle, John T.; Garrison, Virginia H.; Gray, Michael A.

2014-01-01

Laboratory experiments were conducted to assess if exposure to atmospheric African dust stimulates or inhibits the growth of four putative bacterial coral pathogens. Atmospheric dust was collected from a dust-source region (Mali, West Africa) and from Saharan Air Layer masses over downwind sites in the Caribbean [Trinidad and Tobago and St. Croix, U.S. Virgin Islands (USVI)]. Extracts of dust samples were used to dose laboratory-grown cultures of four putative coral pathogens: Aurantimonas coralicida (white plague type II), Serratia marcescens (white pox), Vibrio coralliilyticus, and V. shiloi (bacteria-induced bleaching). Growth of A. coralicida and V. shiloi was slightly stimulated by dust extracts from Mali and USVI, respectively, but unaffected by extracts from the other dust sources. Lag time to the start of log-growth phase was significantly shortened for A. coralicida when dosed with dust extracts from Mali and USVI. Growth of S. marcescens and V. coralliilyticus was neither stimulated nor inhibited by any of the dust extracts. This study demonstrates that constituents from atmospheric dust can alter growth of recognized coral disease pathogens under laboratory conditions.

Development of a subsurface gas flow probe

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

Cutler, R.P.; Ballard, S.; Barker, G.T.

1997-04-01

This report describes a project to develop a flow probe to monitor gas movement in the vadose zone due to passive venting or active remediation efforts such as soil vapor extraction. 3-D and 1-D probes were designed, fabricated, tested in known flow fields under laboratory conditions, and field tested. The 3-D pores were based on technology developed for ground water flow monitoring. The probes gave excellent agreement with measured air velocities in the laboratory tests. Data processing software developed for ground water flow probes was modified for use with air flow, and to accommodate various probe designs. Modifications were mademore » to decrease the cost of the probes, including developing a downhole multiplexer. Modeling indicated problems with flow channeling due to the mode of deployment. Additional testing was conducted and modifications were made to the probe and to the deployment methods. The probes were deployed at three test sites: a large outdoor test tank, a brief vapor extraction test at the Chemical Waste landfill, and at an active remediation site at a local gas station. The data from the field tests varied markedly from the laboratory test data. All of the major events such as vapor extraction system turn on and turn off, as well as changes in the flow rate, could be seen in the data. However, there were long term trends in the data which were much larger than the velocity signals, which made it difficult to determine accurate air velocities. These long term trends may be due to changes in soil moisture content and seasonal ground temperature variations.« less

EVALUATION OF ANTIBACTERIAL, ANTITUMOR, ANTIOXIDANT ACTIVITIES AND PHENOLIC CONSTITUENTS OF FIELD-GROWN AND IN VITRO-GROWN LYSIMACHIA VULGARIS L.

PubMed

Yildirim, Arzu Birinci; Guner, Birgul; Karakas, Fatma Pehlivan; Turker, Arzu Ucar

2017-01-01

Lysimachia vulgaris L. (Yellow loosestrife) is a medicinal plant in the family Myrsinaceae. It has been used in the treatment of fever, ulcer, diarrhea and wounds in folk medicine. It has also analgesic, expectorant, astringent and anti-inflammatory activities. Two different sources of the plant (field-grown and in vitro -grown) were used to evaluate the biological activities (antibacterial, antitumor and antioxidant) of L. vulgaris. In vitro-grown plant materials were collected from L. vulgaris plants that were previously regenerated in our laboratory. Plant materials were extracted with water, ethanol and acetone. For antibacterial test, disc diffusion method and 10 different pathogenic bacteria were used. Antioxidant activity was indicated by using DPPH method. The total phenol amount by using Folin-Ciocaltaeu method and the total flavonoid amount by using aluminum chloride (AlCl 3 ) colorimetric method were determined. Generally, yellow loosestrife extracts demonstrated antibacterial activity against Gram-positive bacteria (Staphylococcus aureus, S. epidermidis and Streptococcus pyogenes) . Strong antitumor activity of yellow loosestrife was observed via potato disc diffusion bioassay. Nine different phenolics were also determined and compared by using High-Performance Liquid Chromatography (HPLC). Future investigations should be focused on fractionation of the extracts to identify active components for biological activity.

Determination of trace metals in drinking water using solid-phase extraction disks and X-ray fluorescence spectrometry.

PubMed

Hou, Xiandeng; Peters, Heather L; Yang, Zheng; Wagner, Karl A; Batchelor, James D; Daniel, Meredith M; Jones, Bradley T

2003-03-01

A convenient method is described for monitoring Cd, Ni, Cu, and Pb at trace levels in drinking water samples. These metals are preconcentrated on a chelating solid-phase extraction disk and then determined by X-ray fluorescence spectrometry. The method tolerates a wide pH range (pH 6-14) and a large amount of alkaline and alkaline earth elements. The preconcentration factor is well over 1600, assuming a 1 L water sample volume. The limits of detection for Cd, Ni, Cu, and Pb are 3.8, 0.6, 0.4, and 0.3 ng/mL, respectively. These are well below the federal maximum contaminant level values, which are 5, 100, 1300, and 15 ng/mL, respectively. The proposed method has many advantages including ease of operation, multielement capability, nondestructiveness, high sensitivity, and relative cost efficiency. The solid-phase extraction step can be conducted in the field and then the disks can be mailed to a laboratory for the analysis, eliminating the cost of transporting large volumes of water samples. Furthermore, the color of the used extraction disk provides an initial estimate of the degree of contamination for some transition metals (for example, Ni and Cu). Thus, the overall cost for analysis of metals in drinking water can be minimized by implementing the method, and small water supply companies with limited budgets will be better able to comply with the Safe Drinking Water Act.

Foucault's method for measuring the speed of light with modern apparatus

NASA Astrophysics Data System (ADS)

Vörös, Zoltán; Weihs, Gregor

2015-05-01

In this paper, we introduce two simple and inexpensive, yet quite accurate versions of the well-known Foucault method for measuring the speed of light. In a compact footprint of just 20 cm by 270 cm with readily available laboratory items, a rotating mirror taken from a laser printer, and a webcam, we obtained c=296\\720+/- 3000 km s-1, and c=302\\295+/- 3000 km s-1, respectively, both within less than a per cent of the defined value. The experiment also prepares students for extracting data through image processing.

Identification of Curie temperature distributions in magnetic particulate systems

NASA Astrophysics Data System (ADS)

Waters, J.; Berger, A.; Kramer, D.; Fangohr, H.; Hovorka, O.

2017-09-01

This paper develops a methodology for extracting the Curie temperature distribution from magnetisation versus temperature measurements which are realizable by standard laboratory magnetometry. The method is integral in nature, robust against various sources of measurement noise, and can be adopted to a wide range of granular magnetic materials and magnetic particle systems. The validity and practicality of the method is demonstrated using large-scale Monte-Carlo simulations of an Ising-like model as a proof of concept, and general conclusions are drawn about its applicability to different classes of systems and experimental conditions.

HORMONAL PROPERTIES OF ETHANOLIC EXTRACT OF JUNIPERUS COMMUNIS LINN.

PubMed Central

Pathak, Sandhya; Tewari, R. K.; Prakash, A.O.

1990-01-01

To evaluate antifertility mode of action of extracts of Juniperus communis Linn., its various extracts were investigated for estrogenic, antiestrogenic, progestagenic and antiprogestagenic properties in laboratory animals. Investigations reveal that the extract possessed only antiprogestational activity which accounts for its antifertility effect. PMID:22556519

A simple method for the small scale synthesis and solid-phase extraction purification of steroid sulfates.

PubMed

Waller, Christopher C; McLeod, Malcolm D

2014-12-01

Steroid sulfates are a major class of steroid metabolite that are of growing importance in fields such as anti-doping analysis, the detection of residues in agricultural produce or medicine. Despite this, many steroid sulfate reference materials may have limited or no availability hampering the development of analytical methods. We report simple protocols for the rapid synthesis and purification of steroid sulfates that are suitable for adoption by analytical laboratories. Central to this approach is the use of solid-phase extraction (SPE) for purification, a technique routinely used for sample preparation in analytical laboratories around the world. The sulfate conjugates of sixteen steroid compounds encompassing a wide range of steroid substitution patterns and configurations are prepared, including the previously unreported sulfate conjugates of the designer steroids furazadrol (17β-hydroxyandrostan[2,3-d]isoxazole), isofurazadrol (17β-hydroxyandrostan[3,2-c]isoxazole) and trenazone (17β-hydroxyestra-4,9-dien-3-one). Structural characterization data, together with NMR and mass spectra are reported for all steroid sulfates, often for the first time. The scope of this approach for small scale synthesis is highlighted by the sulfation of 1μg of testosterone (17β-hydroxyandrost-4-en-3-one) as monitored by liquid chromatography-mass spectrometry (LCMS). Copyright © 2014 Elsevier Inc. All rights reserved.

Development and Validation of a Reliable and Robust Method for the Analysis of Cannabinoids and Terpenes in Cannabis.

PubMed

Giese, Matthew W; Lewis, Mark A; Giese, Laura; Smith, Kevin M

2015-01-01

The requirements for an acceptable cannabis assay have changed dramatically over the years resulting in a large number of laboratories using a diverse array of analytical methodologies that have not been properly validated. Due to the lack of sufficiently validated methods, we conducted a single- laboratory validation study for the determination of cannabinoids and terpenes in a variety of commonly occurring cultivars. The procedure involves high- throughput homogenization to prepare sample extract, which is then profiled for cannabinoids and terpenes by HPLC-diode array detector and GC-flame ionization detector, respectively. Spike recovery studies for terpenes in the range of 0.03-1.5% were carried out with analytical standards, while recovery studies for Δ9-tetrahydrocannabinolic acid, cannabidiolic acid, Δ9-tetrahydrocannabivarinic acid, and cannabigerolic acid and their neutral counterparts in the range of 0.3-35% were carried out using cannabis extracts. In general, accuracy at all levels was within 5%, and RSDs were less than 3%. The interday and intraday repeatabilities of the procedure were evaluated with five different cultivars of varying chemotype, again resulting in acceptable RSDs. As an example of the application of this assay, it was used to illustrate the variability seen in cannabis coming from very advanced indoor cultivation operations.

Nuclear deformation in the laboratory frame

NASA Astrophysics Data System (ADS)

Gilbreth, C. N.; Alhassid, Y.; Bertsch, G. F.

2018-01-01

We develop a formalism for calculating the distribution of the axial quadrupole operator in the laboratory frame within the rotationally invariant framework of the configuration-interaction shell model. The calculation is carried out using a finite-temperature auxiliary-field quantum Monte Carlo method. We apply this formalism to isotope chains of even-mass samarium and neodymium nuclei and show that the quadrupole distribution provides a model-independent signature of nuclear deformation. Two technical advances are described that greatly facilitate the calculations. The first is to exploit the rotational invariance of the underlying Hamiltonian to reduce the statistical fluctuations in the Monte Carlo calculations. The second is to determine quadruple invariants from the distribution of the axial quadrupole operator in the laboratory frame. This allows us to extract effective values of the intrinsic quadrupole shape parameters without invoking an intrinsic frame or a mean-field approximation.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of organochlorine pesticides and polychlorinated biphenyls in bottom sediment by dual capillary-column gas chromatography with electron-capture detection

USGS Publications Warehouse

Foreman, William T.; Connor, Brooke F.; Furlong, Edward T.; Vaught, Deborah G.; Merten, Leslie M.

1995-01-01

A method for the determination of 30 individual organochlorine pesticides, total toxaphene, and total polychlorinated biphenyls (PCBs) in bottom sediment is described. The method isolates the pesticides and PCBs by solvent extraction with dichlorobenzene, removes inorganic sulfur, large naturally occurring molecules, and other unwanted interferences by gel permeation chromatography, and further cleans up and class fractionates the extract using adsorption chromatography. The com- pounds then are instrumentally determined using dual capillary-column gas chromatography with electron-capture detection. Reporting limits range from 1 to 5 micrograms per kilogram for 30 individual pesticides, 50 micrograms per kilogram for total PCBs, and 200 micrograms per kilogram for total toxaphene. The method also is designed to allow the simultaneous isolation of 79 other semivolatile organic compounds from the sediment, which are separately quantified using gas chromatography with mass spectrometric detection. The method was developed in support of the U.S. Geological Survey's National Water-Quality Assessment program.

An atomic-absorption method for the determination of gold in large samples of geologic materials

USGS Publications Warehouse

VanSickle, Gordon H.; Lakin, Hubert William

1968-01-01

A laboratory method for the determination of gold in large (100-gram) samples has been developed for use in the study of the gold content of placer deposits and of trace amounts of gold in other geologic materials. In this method the sample is digested with bromine and ethyl ether, the gold is extracted into methyl isobutyl ketone, and the determination is made by atomicabsorption spectrophotometry. The lower limit of detection is 0.005 part per million in the sample. The few data obtained so far by this method agree favorably with those obtained by assay and by other atomic-absorption methods. About 25 determinations can be made per man-day.

Extraction and [superscript 1]H NMR Analysis of Fats from Convenience Foods: A Laboratory Experiment for Organic Chemistry

ERIC Educational Resources Information Center

Hartel, Aaron M.; Moore, Amy C.

2014-01-01

The extraction and analysis of fats from convenience foods (crackers, cookies, chips, candies) has been developed as an experiment for a second-year undergraduate organic chemistry laboratory course. Students gravimetrically determine the fat content per serving and then perform a [superscript 1]H NMR analysis of the recovered fat to determine the…

Evaluation of a multiresidue method for measuring fourteen chemical groups of pesticides in water by use of LC-MS-MS.

PubMed

Carvalho, J J; Jerónimo, P C A; Gonçalves, C; Alpendurada, M F

2008-11-01

European Council Directive 98/83/EC on the quality of water intended for human consumption brought a new challenge for water-quality control routine laboratories, mainly on pesticides analysis. Under the guidelines of ISO/IEC 17025:2005, a multiresidue method was developed, validated, implemented in routine, and studied with real samples during a one-year period. The proposed method enables routine laboratories to handle a large number of samples, since 28 pesticides of 14 different chemical groups can be quantitated in a single procedure. The method comprises a solid-phase extraction step and subsequent analysis by liquid chromatography-mass spectrometry (LC-MS-MS). The accuracy was established on the basis of participation in interlaboratory proficiency tests, with encouraging results (majority |z-score| <2), and the precision was consistently analysed over one year. The limits of quantitation (below 0.050 microg L(-1)) are in agreement with the enforced threshold value for pesticides of 0.10 microg L(-1). Overall method performance is suitable for routine use according to accreditation rules, taking into account the data collected over one year.

Evaluation of FTA ® paper for storage of oral meta-genomic DNA.

PubMed

Foitzik, Magdalena; Stumpp, Sascha N; Grischke, Jasmin; Eberhard, Jörg; Stiesch, Meike

2014-10-01

The purpose of the present study was to evaluate the short-term storage of meta-genomic DNA from native oral biofilms on FTA(®) paper. Thirteen volunteers of both sexes received an acrylic splint for intraoral biofilm formation over a period of 48 hours. The biofilms were collected, resuspended in phosphate-buffered saline, and either stored on FTA(®) paper or directly processed by standard laboratory DNA extraction. The nucleic acid extraction efficiencies were evaluated by 16S rDNA targeted SSCP fingerprinting. The acquired banding pattern of FTA-derived meta-genomic DNA was compared to a standard DNA preparation protocol. Sensitivity and positive predictive values were calculated. The volunteers showed inter-individual differences in their bacterial species composition. A total of 200 bands were found for both methods and 85% of the banding patterns were equal, representing a sensitivity of 0.941 and a false-negative predictive value of 0.059. Meta-genomic DNA sampling, extraction, and adhesion using FTA(®) paper is a reliable method for storage of microbial DNA for a short period of time.

Disposable pipette extraction for gas chromatographic determination of codeine, morphine, and 6-monoacetylmorphine in vitreous humor.

PubMed

Kovatsi, Leda; Rentifis, Konstantinos; Giannakis, Dimitrios; Njau, Samuel; Samanidou, Victoria

2011-07-01

The availability of a sensitive and rapid analytical method for the determination of opiates, and other substances of forensic interest, in a variety of biological specimens is of utmost importance to forensic laboratories. Solid-phase extraction is very popular in the pre-treatment of forensic samples. Nevertheless, a new approach, disposable pipette extraction (DPX), is gaining increasing interest in sample preparation. DPX has already been applied to the analysis of drugs of abuse in common biological matrices, such as urine and blood, but has not yet been evaluated on alternative biological samples, such as vitreous humor. The objective of this study was to evaluate the applicability of DPX on the analysis of opiates in vitreous humor. The currently developed method is fast, reliable, and easy to perform. The sensitivity, precision, and accuracy are satisfactory. Recoveries obtained are within the range of 72-91%, whereas the sample volume of vitreous humor required is only 100 μL. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Application of dispersive solid phase extraction for trace analysis of toxic chemicals in foods.

PubMed

Neely, Sarah; Martin, Jordan; da Cruz, Natalia Ferreira; Piester, Gavin; Robinson, Morgan; Okoniewski, Richard; Tran, Buu N

2018-05-29

The objectives of this study were to develop and validate a method for the identification of toxic organic chemicals, including groups of controlled substances, alkaloids and pesticides that are highly toxic and considered threats to public health. This project aims to ensure our laboratory's readiness to respond to emergencies involving our food supply in cooperation with the Food Emergency Response Network (FERN) program. The food matrices were homogenized in a blender or food processor prior to extraction with an acetonitrile-water mixture using a QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) procedure. The extracts were then analyzed by either gas chromatography-mass spectrometry (GC-MS) or liquid chromatography-electrospray tandem mass spectrometry (LC-ESI/MS/MS). Method validation was performed on a variety of food matrices including lettuce, grapes, milk, chicken, pork and beef. MDLs for the toxic compounds ranged from 0.01 to 0.66 mg/kg (ppm). The findings in this study will provide a valuable resource for the determination of toxic chemicals in food matrices for emergency response situations. Copyright © 2018. Published by Elsevier B.V.

The Roche Immunoturbidimetric Albumin Method on Cobas c 501 Gives Higher Values Than the Abbott and Roche BCP Methods When Analyzing Patient Plasma Samples.

PubMed

Helmersson-Karlqvist, Johanna; Flodin, Mats; Havelka, Aleksandra Mandic; Xu, Xiao Yan; Larsson, Anders

2016-09-01

Serum/plasma albumin is an important and widely used laboratory marker and it is important that we measure albumin correctly without bias. We had indications that the immunoturbidimetric method on Cobas c 501 and the bromocresol purple (BCP) method on Architect 16000 differed, so we decided to study these methods more closely. A total of 1,951 patient requests with albumin measured with both the Architect BCP and Cobas immunoturbidimetric methods were extracted from the laboratory system. A comparison with fresh plasma samples was also performed that included immunoturbidimetric and BCP methods on Cobas c 501 and analysis of the international protein calibrator ERM-DA470k/IFCC. The median difference between the Abbott BCP and Roche immunoturbidimetric methods was 3.3 g/l and the Roche method overestimated ERM-DA470k/IFCC by 2.2 g/l. The Roche immunoturbidimetric method gave higher values than the Roche BCP method: y = 1.111x - 0.739, R² = 0.971. The Roche immunoturbidimetric albumin method gives clearly higher values than the Abbott and Roche BCP methods when analyzing fresh patient samples. The differences between the two methods were similar at normal and low albumin levels. © 2016 Wiley Periodicals, Inc.

Laboratory assessment of indigenous plant extracts for anti-juvenile hormone activity in Culex quinquefasciatus.

PubMed

Saxena, R C; Dixit, O P; Sukumaran, P

1992-07-01

Of 15 plants tested, five plant extracts showed anti-juvenile hormone-like activity against laboratory colonised late fourth instar larvae and adult female mosquitoes. Petroleum ether extract of Eichhornia crassipes and acetone extracts of Ageratum conyzoides, Cleome icosandra, Tagetes erectes and Tridax procumbens showed growth inhibitory (P less than 0.001) and juvenile hormone mimicing activity to the treated larvae of C. quinquefasciatus.. Larval pupal intermediates, demalanised pupae, defective egg rafts and adult with deformed flight muscles were few noticeable changes. Biting behaviour was observed to be affected only in Ageratum, Cleome and Tridax extracts (P less than 0.001). Loss of fecundity was observed in the treated mosquitoes but no sterilant effects could be seen. Adults, obtained from larvae exposed to the plant extracts produced significantly shorter egg-rafts (P less than 0.005) than in control.

Laboratory evaluation of methanolic extract of Atlantia monophylla (Family: Rutaceae) against immature stages of mosquitoes and non-target organisms.

PubMed

Sivagnaname, N; Kalyanasundaram, M

2004-02-01

Methanolic extracts of the leaves of Atlantia monophylla (Rutaceae) were evaluated for mosquitocidal activity against immature stages of three mosquito species, Culex quinquefasciatus, Anopheles stephensi, and Aedes aegypti in the laboratory. Larvae of Cx. quinquefasciatus and pupae of An. stephensi were found more susceptible, with LC50 values of 0.14 mg/l and 0.05 mg/l, respectively. Insect growth regulating activity of this extract was more pronounced against Ae. aegypti, with EI50 value 0.002 mg/l. The extract was found safe to aquatic mosquito predators Gambusia affinis, Poecilia reticulata, and Diplonychus indicus, with the respective LC50 values of 23.4, 21.3, and 5.7 mg/l. The results indicate that the mosquitocidal effects of the extract of this plant were comparable to neem extract and certain synthetic chemical larvicides like fenthion, methoprene, etc.

Development of the front end test stand and vessel for extraction and source plasma analyses negative hydrogen ion sources at the Rutherford Appleton Laboratory.

PubMed

Lawrie, S R; Faircloth, D C; Letchford, A P; Perkins, M; Whitehead, M O; Wood, T; Gabor, C; Back, J

2014-02-01

The ISIS pulsed spallation neutron and muon facility at the Rutherford Appleton Laboratory (RAL) in the UK uses a Penning surface plasma negative hydrogen ion source. Upgrade options for the ISIS accelerator system demand a higher current, lower emittance beam with longer pulse lengths from the injector. The Front End Test Stand is being constructed at RAL to meet the upgrade requirements using a modified ISIS ion source. A new 10% duty cycle 25 kV pulsed extraction power supply has been commissioned and the first meter of 3 MeV radio frequency quadrupole has been delivered. Simultaneously, a Vessel for Extraction and Source Plasma Analyses is under construction in a new laboratory at RAL. The detailed measurements of the plasma and extracted beam characteristics will allow a radical overhaul of the transport optics, potentially yielding a simpler source configuration with greater output and lifetime.

Methods of analysis by the U.S. Geological Survey National Water Quality Laboratory; determination of semivolatile organic compounds in bottom sediment by solvent extraction, gel permeation chromatographic fractionation, and capillary-column gas chromatography/mass spectrometry

USGS Publications Warehouse

Furlong, E.T.; Vaught, D.G.; Merten, L.M.; Foreman, W.T.; Gates, Paul M.

1996-01-01

A method for the determination of 79 semivolatile organic compounds (SOCs) and 4 surrogate compounds in soils and bottom sediment is described. The SOCs are extracted from bottom sediment by solvent extraction, followed by partial isolation using high-performance gel permeation chromatography (GPC). The SOCs then are qualitatively identified and quantitative concentrations determined by capillary-column gas chromatography/mass spectrometry (GC/MS). This method also is designed for an optional simultaneous isolation of polychlorinated biphenyls (PCBs) and organochlorine (OC) insecticides, including toxaphene. When OCs and PCBs are determined, an additional alumina- over-silica column chromatography step follows GPC cleanup, and quantitation is by dual capillary- column gas chromatography with electron-capture detection (GC/ECD). Bottom-sediment samples are centrifuged to remove excess water and extracted overnight with dichloromethane. The extract is concentrated, centrifuged, and then filtered through a 0.2-micrometer polytetrafluoro-ethylene syringe filter. Two aliquots of the sample extract then are quantitatively injected onto two polystyrene- divinylbenzene GPC columns connected in series. The SOCs are eluted with dichloromethane, a fraction containing the SOCs is collected, and some coextracted interferences, including elemental sulfur, are separated and discarded. The SOC-containing GPC fraction then is analyzed by GC/MS. When desired, a second aliquot from GPC is further processed for OCs and PCBs by combined alumina-over-silica column chromatography. The two fractions produced in this cleanup then are analyzed by GC/ECD. This report fully describes and is limited to the determination of SOCs by GC/MS.

Guaiacol peroxidase zymography for the undergraduate laboratory.

PubMed

Wilkesman, Jeff; Castro, Diana; Contreras, Lellys M; Kurz, Liliana

2014-01-01

This laboratory exercise presents a novel way to introduce undergraduate students to the specific detection of enzymatic activity by electrophoresis. First, students prepare a crude peroxidase extract and then analyze the homogenate via electrophoresis. Zymography, that is, a SDS-PAGE method to detect enzyme activity, is used to specifically detect peroxidase activity and furthermore, to analyze the total protein profile. After the assay, students may estimate the apparent molecular mass of the enzyme and discuss its structure. After the 4-h experiment, students gain knowledge concerning biological sample preparation, gel preparation, electrophoresis, and the importance of specific staining procedures for the detection of enzymatic activity. Copyright © 2014 The International Union of Biochemistry and Molecular Biology.

The LabTube - a novel microfluidic platform for assay automation in laboratory centrifuges.

PubMed

Kloke, A; Fiebach, A R; Zhang, S; Drechsel, L; Niekrawietz, S; Hoehl, M M; Kneusel, R; Panthel, K; Steigert, J; von Stetten, F; Zengerle, R; Paust, N

2014-05-07

Assay automation is the key for successful transformation of modern biotechnology into routine workflows. Yet, it requires considerable investment in processing devices and auxiliary infrastructure, which is not cost-efficient for laboratories with low or medium sample throughput or point-of-care testing. To close this gap, we present the LabTube platform, which is based on assay specific disposable cartridges for processing in laboratory centrifuges. LabTube cartridges comprise interfaces for sample loading and downstream applications and fluidic unit operations for release of prestored reagents, mixing, and solid phase extraction. Process control is achieved by a centrifugally-actuated ballpen mechanism. To demonstrate the workflow and functionality of the LabTube platform, we show two LabTube automated sample preparation assays from laboratory routines: DNA extractions from whole blood and purification of His-tagged proteins. Equal DNA and protein yields were observed compared to manual reference runs, while LabTube automation could significantly reduce the hands-on-time to one minute per extraction.

DNA extraction protocols cause differences in 16S rRNA amplicon sequencing efficiency but not in community profile composition or structure

DOE PAGES

None

2014-12-01

The recent development of methods applying next-generation sequencing to microbial community characterization has led to the proliferation of these studies in a wide variety of sample types. Yet, variation in the physical properties of environmental samples demands that optimal DNA extraction techniques be explored for each new environment. The microbiota associated with many species of insects offer an extraction challenge as they are frequently surrounded by an armored exoskeleton, inhibiting disruption of the tissues within. In this study, we examine the efficacy of several commonly used protocols for extracting bacterial DNA from ants. While bacterial community composition recovered using Illuminamore » 16S rRNA amplicon sequencing was not detectably biased by any method, the quantity of bacterial DNA varied drastically, reducing the number of samples that could be amplified and sequenced. These results indicate that the concentration necessary for dependable sequencing is around 10,000 copies of target DNA per microliter. Exoskeletal pulverization and tissue digestion increased the reliability of extractions, suggesting that these steps should be included in any study of insect-associated microorganisms that relies on obtaining microbial DNA from intact body segments. Although laboratory and analysis techniques should be standardized across diverse sample types as much as possible, minimal modifications such as these will increase the number of environments in which bacterial communities can be successfully studied.« less

Simple procedures to obtain exogenous internal controls for use in RT-PCR detection of bovine pestiviruses.

PubMed

Golemba, Marcelo D; Parreño, Viviana; Jones, Leandro R

2008-06-01

Pestiviruses are ubiquitous pathogens of cattle and frequent adventitious viruses in biologicals. Furthermore, it has been suggested that these agents might be related to infantile gastroenteritis and microencephaly. Since the virus is highly prevalent in fetal bovine serum, the risk of contamination is high in most laboratories. Thus, the implementation of detection methods in all laboratories is of worth. Despite continuous surveillance, these agents have been detected in cell lines, fetal bovine serum, live and inactivated animal and human vaccines and interferon for human use. In this report, DNA and RNA internal controls (ICs) which can be implemented in laboratories with minimal equipment are described. The developed standards can be added before RNA purification, allowing to monitor all steps of the protocol (viral RNA extraction, reverse transcription and cDNA amplification). It is shown that inhibitory effects that could lead to decreased sensitivity can be minimized by controlling the amount of mimic molecules added to the samples. A method to avoid the problem of DNA traces present in in vitro transcribed RNA preparations is provided.

Otolith Trace Element Chemistry of Juvenile Black Rockfish

NASA Astrophysics Data System (ADS)

Hardin, W.; Bobko, S. J.; Jones, C. M.

2002-12-01

In the summer of 1997 we collected young-of -the-year (YOY) black rockfish, Sebastes melanops, from floating docks and seagrass beds in Newport and Coos Bay, Oregon. Otoliths were extracted from randomly selected fish, sectioned and polished under general laboratory conditions, and cleaned in a class 100 clean room. We used Laser Ablation - Inductively Coupled Mass Spectrometry (LA-ICPMS) to analyze elemental composition of the estuarine phase of the otoliths. While we observed differences in Mn/Ca ratios between the two estuaries, there was no statistical difference in otolith trace element chemistry ratios between estuaries using MANOVA. To determine if laboratory processing of otoliths might have impeded us from detecting differences in otolith chemistry, we conducted a second experiment. Right and left otoliths from 10 additional Coos Bay fish were randomly allocated to two processing methods. The first method was identical to our initial otolith processing, sectioning and polishing under normal laboratory conditions. In the second method, polishing was done in the clean room. For both methods otoliths went through a final cleaning in the clean room and analyzed with LA-ICPMS. While we did not detect statistical differences in element ratios between the two methods, otoliths polished outside the clean room had much higher variances. This increased variance might have lowered our ability to detect differences in otolith chemistry between estuaries. Based on our results, we recommend polishing otoliths under clean room conditions to reduce contamination.

Blood meal analysis of tabanid fly after it biting the rare Sumatran rhinoceros

PubMed Central

Rovie-Ryan, Jeffrine Japning; Zainuddin, Zainal Zahari; Marni, Wahap; Ahmad, Abdul Hamid; Ambu, Laurentius N.; Payne, Junaidi

2013-01-01

Objective To demonstrate a noninvasive large mammalian genetic sampling method using blood meal obtained from a tabanid fly. Methods Blood meal was recovered from the abdomen of an engorged tabanid fly (Haematopota sp.) which was captured immediately after biting a Sumatran rhino in captivity. The blood was applied on to a Whatman FTA® blood card. Subsequent laboratory work was conducted to extract, amplify and sequence the DNA from the sample. Validation was done by sampling the hair follicles and blood samples from the rhinoceros and subjecting it to the same laboratory process. Results BLAST search and constructed phylogenetic trees confirmed the blood meal samples were indeed from the rhino. Conclusions This method could be used in the field application to noninvasively collect genetic samples. Collection of tabanids and other haematophagous arthropods (e.g. mosquitoes and ticks) and other blood-sucking parasites (e.g. leeches and worms) could also provide information on vector-borne diseases. PMID:23593586

Inter-laboratory comparison study on measuring semi-volatile organic chemicals in standards and air samples.

PubMed

Su, Yushan; Hung, Hayley

2010-11-01

Measurements of semi-volatile organic chemicals (SVOCs) were compared among 21 laboratories from 7 countries through the analysis of standards, a blind sample, an air extract, and an atmospheric dust sample. Measurement accuracy strongly depended on analytes, laboratories, and types of standards and samples. Intra-laboratory precision was generally good with relative standard deviations (RSDs) of triplicate injections <10% and with median differences of duplicate samples between 2.1 and 22%. Inter-laboratory variability, measured by RSDs of all measurements, was in the range of 2.8-58% in analyzing standards, and 6.9-190% in analyzing blind sample and air extract. Inter-laboratory precision was poorer when samples were subject to cleanup processes, or when SVOCs were quantified at low concentrations. In general, inter-laboratory differences up to a factor of 2 can be expected to analyze atmospheric SVOCs. When comparing air measurements from different laboratories, caution should be exercised if the data variability is less than the inter-laboratory differences. 2010. Published by Elsevier Ltd. All rights reserved.

A small-scale, portable method for extracting microplastics from marine sediments.

PubMed

Coppock, Rachel L; Cole, Matthew; Lindeque, Penelope K; Queirós, Ana M; Galloway, Tamara S

2017-11-01

Microplastics (plastic particles, 0.1 μm-5 mm in size) are widespread marine pollutants, accumulating in benthic sediments and shorelines the world over. To gain a clearer understanding of microplastic availability to marine life, and the risks they pose to the health of benthic communities, ecological processes and food security, it is important to obtain accurate measures of microplastic abundance in marine sediments. To date, methods for extracting microplastics from marine sediments have been disadvantaged by complexity, expense, low extraction efficiencies and incompatibility with very fine sediments. Here we present a new, portable method to separate microplastics from sediments of differing types, using the principle of density floatation. The Sediment-Microplastic Isolation (SMI) unit is a custom-built apparatus which consistently extracted microplastics from sediments in a single step, with a mean efficiency of 95.8% (±SE 1.6%; min 70%, max 100%). Zinc chloride, at a density of 1.5 g cm -3 , was deemed an effective and relatively inexpensive floatation media, allowing fine sediment to settle whilst simultaneously enabling floatation of dense polymers. The method was validated by artificially spiking sediment with low and high density microplastics, and its environmental relevance was further tested by extracting plastics present in natural sediment samples from sites ranging in sediment type; fine silt/clay (mean size 10.25 ± SD 3.02 μm) to coarse sand (mean size 149.3 ± SD 49.9 μm). The method presented here is cheap, reproducible and is easily portable, lending itself for use in the laboratory and in the field, eg. on board research vessels. By employing this method, accurate estimates of microplastic type, distribution and abundance in natural sediments can be achieved, with the potential to further our understanding of the availability of microplastics to benthic organisms. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Determination of submicrogram-per-liter concentrations of caffeine in surface water and groundwater samples by solid-phase extraction and liquid chromatography

USGS Publications Warehouse

Burkhardt, M.R.; Soliven, P.P.; Werner, S.L.; Vaught, D.G.

1999-01-01

A method for determining submicrogram-per-liter concentrations of caffeine in surface water and groundwater samples has been developed. Caffeine is extracted from a 1 L water sample with a 0.5 g graphitized carbon-based solid-phase cartridge, eluted with methylene chloride-methanol (80 + 20, v/v), and analyzed by liquid chromatography with photodiode-array detection. The single-operator method detection limit for organic-free water samples was 0.02 ??g/L. Mean recoveries and relative standard deviations were 93 ?? 13% for organicfree water samples fortified at 0.04 ??g/L and 84 ?? 4% for laboratory reagent spikes fortified at 0.5 ??g/L. Environmental concentrations of caffeine ranged from 0.003 to 1.44 ??g/L in surface water samples and from 0.01 to 0.08 ??g/L in groundwater samples.

Determination of the acid value of instant noodles: interlaboratory study.

PubMed

Hakoda, Akiko; Sakaida, Kenichi; Suzuki, Tadanao; Yasui, Akemi

2006-01-01

An interlaboratory study was performed to evaluate the method for determining the acid value of instant noodles, based on the Japanese Agricultural Standard (JAS), with extraction of lipid using petroleum ether at a volume of 100 mL to the test portion of 25 g. Thirteen laboratories participated and analyzed 5 test samples as blind duplicates. Statistical treatment revealed that the repeatability (RSDr) of acid value was <6.5%, and the reproducibility (RSDR) of acid value was <9.6%. The HorRat values (RSDR/predicted RSDR) were 1.2-1.8, where the RSDR and the predicted RSDR were obtained in terms of free fatty acids in the noodles per unit weight, using the equation [acid value = percent free fatty acids (as oleic) x 1.99] and the extracted lipid contents. This method was shown to have acceptable precision by the present study.

Report to DHS on Summer Internship 2006

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

Beckwith, R H

2006-07-26

This summer I worked at Lawrence Livermore National Laboratory in a bioforensics collection and extraction research group under David Camp. The group is involved with researching efficiencies of various methods for collecting bioforensic evidence from crime scenes. The different methods under examination are a wipe, swab, HVAC filter and a vacuum. The vacuum is something that has particularly gone uncharacterized. My time was spent mostly on modeling and calculations work, but at the end of the summer I completed my internship with a few experiments to supplement my calculations. I had two major projects this summer. My first major projectmore » this summer involved fluid mechanics modeling of collection and extraction situations. This work examines different fluid dynamic models for the case of a micron spore attached to a fiber. The second project I was involved with was a statistical analysis of the different sampling techniques.« less

A novel procedure for the extraction of protein deposits from soft hydrophilic contact lenses for analysis.

PubMed

Keith, D; Hong, B; Christensen, M

1997-05-01

A quick, simple, and efficient extraction technique was developed for the removal of protein from soft hydrophilic contact lenses. An extraction solvent consisting of a 50:50 mix of 0.2% trifluoroacetic acid and acetonitrile was used to remove protein from in vitro laboratory-deposited and human-worn contact lenses. The protein removed was analyzed using HPLC, bicinchoninic acid (BCA) analysis, and SDS-PAGE gel electreophoresis. Extraction efficiency for lysozyme from laboratory-deposited Group IV lenses was determined to be approximately 100%. Group IV human-worn contact lenses were extracted and analyzed for lysozyme by HPLC and total protein by bicinchoninic acid (BCA) analysis. Groups I, II, III, and IV contact lenses deposited with an artificial tear protein solution and human-worn lenses were extracted and analyzed by SDS-PAGE gel electreophoresis and micro-BCA. The ACN/TFA procedure offers a simple, quick, and efficient extraction technique for removal of protein from contact lenses for subsequent analysis.

Use of a Latent Topic Model for Characteristic Extraction from Health Checkup Questionnaire Data.

PubMed

Hatakeyama, Y; Miyano, I; Kataoka, H; Nakajima, N; Watabe, T; Yasuda, N; Okuhara, Y

2015-01-01

When patients complete questionnaires during health checkups, many of their responses are subjective, making topic extraction difficult. Therefore, the purpose of this study was to develop a model capable of extracting appropriate topics from subjective data in questionnaires conducted during health checkups. We employed a latent topic model to group the lifestyle habits of the study participants and represented their responses to items on health checkup questionnaires as a probability model. For the probability model, we used latent Dirichlet allocation to extract 30 topics from the questionnaires. According to the model parameters, a total of 4381 study participants were then divided into groups based on these topics. Results from laboratory tests, including blood glucose level, triglycerides, and estimated glomerular filtration rate, were compared between each group, and these results were then compared with those obtained by hierarchical clustering. If a significant (p < 0.05) difference was observed in any of the laboratory measurements between groups, it was considered to indicate a questionnaire response pattern corresponding to the value of the test result. A comparison between the latent topic model and hierarchical clustering grouping revealed that, in the latent topic model method, a small group of participants who reported having subjective signs of urinary disorder were allocated to a single group. The latent topic model is useful for extracting characteristics from a small number of groups from questionnaires with a large number of items. These results show that, in addition to chief complaints and history of past illness, questionnaire data obtained during medical checkups can serve as useful judgment criteria for assessing the conditions of patients.

Analysis of Organic Molecules Extracted from Mars Analogues and Influence of Their Mineralogy Using N-Methyl-N-(tert-butyldimethylsilyl)Trifluoroacetamide Derivatization Coupled with Gas Chromatography Mass Spectrometry in Preparation for the Sample Analysis at Mars Derivatization Experiment on the Mars Science Laboratory Mission

NASA Technical Reports Server (NTRS)

Stalport, F.; Glavin, D. P.; Eigenbrode, J. L.; Bish, D.; Blake, D.; Coll, P.; Szopa, C.; Buch, A.; McAdam, A.; Dworkin, J. P.;

Laboratory Evaluations of the Fractions Efficacy of Annona senegalensis (Annonaceae) Leaf Extract on Immature Stage Development of Malarial and Filarial Mosquito Vectors

PubMed Central

Lame, Younoussa; Nukenine, Elias Nchiwan; Pierre, Danga Yinyang Simon; Elijah, Ajaegbu Eze; Esimone, Charles Okechukwu

2015-01-01

Background: Within the framework to control mosquitoes, ovicidal, larvicidal and pupicidal activity of Annona senegalensis leaf extract and its 4 fractions against Anopheles gambiae and Culex quinquefasciatus were evaluated in the laboratory conditions. Methods: Ovicidal test was performed by submitting at least 100 eggs of mosquitoes to 125, 250, 500, 1000 and 2000 ppm concentrations, while larvicidal and pupicidal effects were assessed by submitting 25 larvae or pupae to the concentrations of 2500, 1250, 625 and 312.5 ppm of plant extract or fractions of A. senegalensis. Results: The eggs of An. gambiae were most affected by N-hexane (0.00% hatchability) and chloroform (03.67% hatchability) fractions compared to Cx. quinquefasciatus where at least 25 % hatchability were recorded at 2000 ppm. For larvicidal test, N-hexane (LC50= 298.8 ppm) and chloroform (LC50= 418.3 ppm) fractions were more effective than other fractions on An. gambiae larvae while, a moderate effectiveness was also observed with N-hexane (LC50= 2087.6 ppm), chloroform (LC50= 9010.1 ppm) fractions on Cx. quinquefasciatus larvae. The highest mortality percent of the pupae were also recorded with N-hexane and chloroform fractions on An. gambiae at 2500 ppm. As for Cx. quinquefasciatus only 50 % and 36 % mortality were recorded with N-hexane and chloroform fractions respectively. Conclusion: The extract of A. senegalensis was toxic on immature stage of mosquito species tested. By splitting methanolic crude extract, only N-hexane and chloroform fractions were revealed to possess a mosquitocidal effects and could be considered and utilized for future immature mosquito vectors control. PMID:26623434

Antibacterial and antioxidant properties of the methanol extracts of the leaves and stems of Calpurnia aurea

PubMed Central

Adedapo, Adeolu A; Jimoh, Florence O; Koduru, Srinivas; Afolayan, Anthony J; Masika, Patrick J

2008-01-01

Background In South Africa, Calpurnia aurea (Ait.) Benth is used to destroy lice and to relieve itches, to destroy maggots and to treat allergic rashes, particularly those caused by caterpillars. Antioxidants play an important role protecting against damage by reactive oxygen species. Plants containing flavonoids have been reported to possess strong antioxidant properties. Methods The antibacterial, antioxidant activities and phenolic contents of the methanol extracts of the leaves and stems of Calpurnia aurea were evaluated using in vitro standard methods. Spectrophotometry was the basis for the determinations of total phenol, total flavonoids, flavonols, and proanthocyanidins. Tannins, quercetin and catechin equivalents were used for these parameters. The antioxidant activities of the stem extract of Calpurnia aurea were determined by ABTS, DPPH, and ferrous reducing antioxidant property (FRAP) methods. Laboratory isolates of 10 bacteria species which included five Gram-positive and five Gram-negative strains were used to assay for antibacterial activity of this plant. Results The results from this study showed that the antioxidant activities of the stem extract of Calpurnia aurea as determined by the total phenol, flavonoids, and FRAP methods were higher than that of the leaves. On the other hand, the leaf extract of the plant has higher level of total flavonols and proanthocyanidins. The leaf extract also has higher radical scavenging activity as shown in 1, 1-Diphenyl-2-picrylhydrazyl (DPPH), and 2,2¿-azinobis-3- ethylbenzothiazoline-6-sulfonic acid (ABTS) assay. The leaf extract showed activity against seven of the bacterial organisms. Conclusion The results from this study indicate that the leaves and stem extracts of Calpurnia aurea possess antioxidant properties and could serve as free radical inhibitors or scavenger or, acting possibly as primary antioxidants. Although, the antibacterial properties of Calpurnia aurea are not as effective as the standard drugs- Chloramphenicol and Streptomycin, they still possess some activity against bacterial strains used in this study. Calpurnia aurea may therefore be a good candidate for functional foods as well as pharmaceutical plant-based products. PMID:18803865

Novel synthesis of nanocomposite for the extraction of Sildenafil Citrate (Viagra) from water and urine samples: Process screening and optimization.

PubMed

Asfaram, Arash; Ghaedi, Mehrorang; Purkait, Mihir Kumar

2017-09-01

A sensitive analytical method is investigated to concentrate and determine trace level of Sildenafil Citrate (SLC) present in water and urine samples. The method is based on a sample treatment using dispersive solid-phase micro-extraction (DSPME) with laboratory-made Mn@ CuS/ZnS nanocomposite loaded on activated carbon (Mn@ CuS/ZnS-NCs-AC) as a sorbent for the target analyte. The efficiency was enhanced by ultrasound-assisted (UA) with dispersive nanocomposite solid-phase micro-extraction (UA-DNSPME). Four significant variables affecting SLC recovery like; pH, eluent volume, sonication time and adsorbent mass were selected by the Plackett-Burman design (PBD) experiments. These selected factors were optimized by the central composite design (CCD) to maximize extraction of SLC. The results exhibited that the optimum conditions for maximizing extraction of SLC were 6.0 pH, 300μL eluent (acetonitrile) volume, 10mg of adsorbent and 6min sonication time. Under optimized conditions, virtuous linearity of SLC was ranged from 30 to 4000ngmL -1 with R 2 of 0.99. The limit of detection (LOD) was 2.50ngmL -1 and the recoveries at two spiked levels were ranged from 97.37 to 103.21% with the relative standard deviation (RSD) less than 4.50% (n=15). The enhancement factor (EF) was 81.91. The results show that the combination UAE with DNSPME is a suitable method for the determination of SLC in water and urine samples. Copyright © 2017 Elsevier B.V. All rights reserved.

Structural system identification based on variational mode decomposition

NASA Astrophysics Data System (ADS)

Bagheri, Abdollah; Ozbulut, Osman E.; Harris, Devin K.

2018-03-01

In this paper, a new structural identification method is proposed to identify the modal properties of engineering structures based on dynamic response decomposition using the variational mode decomposition (VMD). The VMD approach is a decomposition algorithm that has been developed as a means to overcome some of the drawbacks and limitations of the empirical mode decomposition method. The VMD-based modal identification algorithm decomposes the acceleration signal into a series of distinct modal responses and their respective center frequencies, such that when combined their cumulative modal responses reproduce the original acceleration response. The decaying amplitude of the extracted modal responses is then used to identify the modal damping ratios using a linear fitting function on modal response data. Finally, after extracting modal responses from available sensors, the mode shape vector for each of the decomposed modes in the system is identified from all obtained modal response data. To demonstrate the efficiency of the algorithm, a series of numerical, laboratory, and field case studies were evaluated. The laboratory case study utilized the vibration response of a three-story shear frame, whereas the field study leveraged the ambient vibration response of a pedestrian bridge to characterize the modal properties of the structure. The modal properties of the shear frame were computed using analytical approach for a comparison with the experimental modal frequencies. Results from these case studies demonstrated that the proposed method is efficient and accurate in identifying modal data of the structures.

Analysis of the stability of urea in dried blood spots collected and stored on filter paper.

PubMed

Quraishi, Rizwana; Lakshmy, Ramakrishnan; Mukhopadhyay, Ashok Kumar; Jailkhani, Bansi Lal

2013-05-01

The ability to use dry blood spots (DBSs) on filter paper for the analysis of urea levels could be an important diagnostic tool for areas that have limited access to laboratory facilities. We developed a method for the extraction and quantification of urea from DBSs that were stored on 3M Whatman filter paper and investigated the effect of long-term storage on the level of urea in DBSs. DBSs of 4.5 mm in diameter were used for our assay, and we determined the urea levels in blood using a commercially available enzymatic kit (UV GLDH-method; Randox laboratories Ltd., UK). The DBSs on filter discs were stored at 4℃ or at 37℃ for 120 days. The mean intra- and inter-assay coefficient of variance for our method of urea extraction from dried blood was 4.2% and 6.3%, respectively. We collected 75 fresh blood samples and compared the urea content of each fresh sample with the urea content of DBSs taken from corresponding fresh blood samples. Regression analysis reported a regression coefficient (r) value of 0.97 and a recovery of urea from dried spots was 102.2%. Urea concentrations in DBSs were stable for up to 120 and 90 days when stored at 4℃ and 37℃, respectively. Our results show that urea can be stored and quantitatively recovered from small volumes of blood that was collected on filter paper.

Blood meal analysis of tabanid fly after it biting the rare Sumatran rhinoceros.

PubMed

Rovie-Ryan, Jeffrine Japning; Zainuddin, Zainal Zahari; Marni, Wahap; Ahmad, Abdul Hamid; Ambu, Laurentius N; Payne, Junaidi

2013-02-01

To demonstrate a noninvasive large mammalian genetic sampling method using blood meal obtained from a tabanid fly. Blood meal was recovered from the abdomen of an engorged tabanid fly (Haematopota sp.) which was captured immediately after biting a Sumatran rhino in captivity. The blood was applied on to a Whatman FTA(®) blood card. Subsequent laboratory work was conducted to extract, amplify and sequence the DNA from the sample. Validation was done by sampling the hair follicles and blood samples from the rhinoceros and subjecting it to the same laboratory process. BLAST search and constructed phylogenetic trees confirmed the blood meal samples were indeed from the rhino. This method could be used in the field application to noninvasively collect genetic samples. Collection of tabanids and other haematophagous arthropods (e.g. mosquitoes and ticks) and other blood-sucking parasites (e.g. leeches and worms) could also provide information on vector-borne diseases.

Simultaneous analysis of aminoglycosides with many other classes of drug residues in bovine tissues by ultrahigh-performance liquid chromatography-tandem mass spectrometry using an ion-pairing reagent added to final extracts

USDA-ARS?s Scientific Manuscript database

The way to maximize scope of analysis, sample throughput, and laboratory efficiency in the monitoring of veterinary drug residues in food animals is to determine as many analytes as possible as fast as possible in as few methods as possible. Capital and overhead expenses are also reduced by using f...

Round-robin comparison of methods for the detection of human enteric viruses in lettuce.

PubMed

Le Guyader, Françoise S; Schultz, Anna-Charlotte; Haugarreau, Larissa; Croci, Luciana; Maunula, Leena; Duizer, Erwin; Lodder-Verschoor, Froukje; von Bonsdorff, Carl-Henrik; Suffredini, Elizabetha; van der Poel, Wim M M; Reymundo, Rosanna; Koopmans, Marion

2004-10-01

Five methods that detect human enteric virus contamination in lettuce were compared. To mimic multiple contaminations as observed after sewage contamination, artificial contamination was with human calicivirus and poliovirus and animal calicivirus strains at different concentrations. Nucleic acid extractions were done at the same time in the same laboratory to reduce assay-to-assay variability. Results showed that the two critical steps are the washing step and removal of inhibitors. The more reliable methods (sensitivity, simplicity, low cost) included an elution/concentration step and a commercial kit. Such development of sensitive methods for viral detection in foods other than shellfish is important to improve food safety.

Cytotoxicity of Brazilian plant extracts against oral microorganisms of interest to dentistry.

PubMed

de Oliveira, Jonatas Rafael; de Castro, Vinicius Carlos; das Graças Figueiredo Vilela, Polyana; Camargo, Samira Esteves Afonso; Carvalho, Cláudio Antonio Talge; Jorge, Antonio Olavo Cardoso; de Oliveira, Luciane Dias

2013-08-15

With the emergence of strains resistant to conventional antibiotics, it is important to carry studies using alternative methods to control these microorganisms causing important infections, such as the use of products of plant origin that has demonstrated effective antimicrobial activity besides biocompatibility. Therefore, this study aimed to evaluate the antimicrobial activity of plant extracts of Equisetum arvense L., Glycyrrhiza glabra L., Punica granatum L. and Stryphnodendron barbatimam Mart. against Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus mutans, Candida albicans, Candida tropicalis, and Candida glabrata, and to analyze the cytotoxicity of these extracts in cultured murine macrophages (RAW 264.7). Antimicrobial activity of plant extracts was evaluated by microdilution method based on Clinical and Laboratory Standards Institute (CLSI), M7-A6 and M27-A2 standards. The cytotoxicity of concentrations that eliminated the microorganisms was evaluated by MTT colorimetric method and by quantification of proinflammatory cytokines (IL-1β and TNF-α) using ELISA. In determining the minimum microbicidal concentration, E. arvense L., P. granatum L., and S. barbatimam Mart. extracts at a concentration of 50 mg/mL and G. glabra L. extract at a concentration of 100 mg/mL, were effective against all microorganisms tested. Regarding cell viability, values were 48% for E. arvense L., 76% for P. granatum L, 86% for S. barbatimam Mart. and 79% for G. glabra L. at the same concentrations. About cytokine production after stimulation with the most effective concentrations of the extracts, there was a significant increase of IL-1β in macrophage cultures treated with S. barbatimam Mart. (3.98 pg/mL) and P. granatum L. (7.72 pg/mL) compared to control (2.20 pg/mL) and a significant decrease of TNF-α was observed in cultures treated with G. glabra L. (4.92 pg/mL), S. barbatimam Mart. (0.85 pg/mL), E. arvense L. (0.83 pg/mL), and P. granatum L. (0.00 pg/mL) when compared to control (41.96 pg/mL). All plant extracts were effective against the microorganisms tested. The G. glabra L. extract exhibited least cytotoxicity and the E. arvense L. extract was the most cytotoxic.

Single-Laboratory Validation for the Determination of Flavonoids in Hawthorn Leaves and Finished Products by LC-UV.

PubMed

Mudge, Elizabeth M; Liu, Ying; Lund, Jensen A; Brown, Paula N

2016-11-01

Suitably validated analytical methods that can be used to quantify medicinally active phytochemicals in natural health products are required by regulators, manufacturers, and consumers. Hawthorn ( Crataegus ) is a botanical ingredient in natural health products used for the treatment of cardiovascular disorders. A method for the quantitation of vitexin-2″- O - rhamnoside, vitexin, isovitexin, rutin, and hyperoside in hawthorn leaf and flower raw materials and finished products was optimized and validated according to AOAC International guidelines. A two-level partial factorial study was used to guide the optimization of the sample preparation. The optimal conditions were found to be a 60-minute extraction using 50 : 48 : 2 methanol : water : acetic acid followed by a 25-minute separation using a reversed-phased liquid chromatography column with ultraviolet absorbance detection. The single-laboratory validation study evaluated method selectivity, accuracy, repeatability, linearity, limit of quantitation, and limit of detection. Individual flavonoid content ranged from 0.05 mg/g to 17.5 mg/g in solid dosage forms and raw materials. Repeatability ranged from 0.7 to 11.7 % relative standard deviation corresponding to HorRat ranges from 0.2 to 1.6. Calibration curves for each flavonoid were linear within the analytical ranges with correlation coefficients greater than 99.9 %. Herein is the first report of a validated method that is fit for the purpose of quantifying five major phytochemical marker compounds in both raw materials and finished products made from North American ( Crataegus douglasii ) and European ( Crataegus monogyna and Crataegus laevigata) hawthorn species. The method includes optimized extraction of samples without a prolonged drying process and reduced liquid chromatography separation time. Georg Thieme Verlag KG Stuttgart · New York.

Direct total and free testosterone measurement by liquid chromatography tandem mass spectrometry across two different platforms.

PubMed

Rhea, Jeanne M; French, Deborah; Molinaro, Ross J

2013-05-01

To develop and validate liquid chromatography tandem mass spectrometry (LC-MS/MS) methods for the direct measurement of total and free testosterone in patient samples on two different analytical systems. An API 4000 and 5000 triple quadropoles were used and compared; the former is reported to be 3-5 times less sensitive, as was used to set the quantitation limits. Free testosterone was separated from the protein-bound fraction by equilibrium dialysis followed by derivatization. Either free or total testosterone, and a deuterated internal standard (d3-testosterone) were extracted by liquid-liquid extraction. The validation results were compared to two different clinical laboratories. The use of d2-testosterone was found to be unacceptable for our method. The total testosterone LC-MS/MS methods on both systems were linear over a wide concentration range of 1.5-2000ng/dL. Free testosterone was measured directly using equilibrium dialysis coupled LC-MS/MS and linear over the concentration range of 2.5-2500pg/mL. Good correlation (total testosterone, R(2)=0.96; free testosterone, R(2)=0.98) was observed between our LC-MS/MS systems and comparator laboratory. However, differences in absolute values for both free and total testosterone measurements were observed while a comparison to a second published LC-MS/MS method showed excellent correlation. Free and total testosterone measurements correlated well with clinical observations. To our knowledge, this is the first published validation of free and total testosterone methods across two analytical systems of different analytical sensitivities. A less sensitive system does not sacrifice analytical or clinical sensitivity to directly measure free and total testosterone in patient samples. Copyright © 2013 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Nucleic acid purification from plants, animals and microbes in under 30 seconds

PubMed Central

Zou, Yiping; Wang, Yuling; Wee, Eugene; Turni, Conny; Blackall, Patrick J.; Trau, Matt; Botella, Jose Ramon

2017-01-01

Nucleic acid amplification is a powerful molecular biology tool, although its use outside the modern laboratory environment is limited due to the relatively cumbersome methods required to extract nucleic acids from biological samples. To address this issue, we investigated a variety of materials for their suitability for nucleic acid capture and purification. We report here that untreated cellulose-based paper can rapidly capture nucleic acids within seconds and retain them during a single washing step, while contaminants present in complex biological samples are quickly removed. Building on this knowledge, we have successfully created an equipment-free nucleic acid extraction dipstick methodology that can obtain amplification-ready DNA and RNA from plants, animals, and microbes from difficult biological samples such as blood and leaves from adult trees in less than 30 seconds. The simplicity and speed of this method as well as the low cost and availability of suitable materials (e.g., common paper towelling), means that nucleic acid extraction is now more accessible and affordable for researchers and the broader community. Furthermore, when combined with recent advancements in isothermal amplification and naked eye DNA visualization techniques, the dipstick extraction technology makes performing molecular diagnostic assays achievable in limited resource settings including university and high school classrooms, field-based environments, and developing countries. PMID:29161268

Laboratory Information Bulletin: Quantitation of Aflatoxin M1 in Bovine Milk by Liquid Chromatography with Fluorescence Detection.

PubMed

Vega, Victor A; Young, Michelle; Todd, Sarah

2016-01-01

An extraction for aflatoxin M1 from bovine milk samples is described. The samples were extracted by adding 10 mL acetonitrile to 10 g of sample. The extract was salted out with sodium chloride and magnesium sulfate to separate the water and acetonitrile. The organic layer was dried down and reconstituted in water before being subjected to an immunoaffinity column for cleanup. Once the analyte was isolated, quantitation was obtained by LC with fluorescence detection. LC/fluorescence parameters were optimized with an Agilent Poroshell 120 C18 LC column resulting in a 4 min run time. To test the procedure's robustness, three different kinds of matrixes were fortified at three different levels each. Whole milk, reduced fat milk, and skim milk samples were fortified at approximately 0.25, 0.5, and 1.0 μg/kg. Recoveries from all samples ranged from 70 to 100%. Confirmation was accomplished by injecting the samples in an ion trap mass spectrometer. The method presented here entails an extraction step followed by an immunoaffinity column clean-up that leads to fast analysis time and consistent recoveries with an uncertainty measurement of 10.5% and method detection limit of less than 0.011 μg/kg.

Determination of Wastewater Compounds in Whole Water by Continuous Liquid-Liquid Extraction and Capillary-Column Gas Chromatography/Mass Spectrometry

USGS Publications Warehouse

Zaugg, Steven D.; Smith, Steven G.; Schroeder, Michael P.

2006-01-01

A method for the determination of 69 compounds typically found in domestic and industrial wastewater is described. The method was developed in response to increasing concern over the impact of endocrine-disrupting chemicals on aquatic organisms in wastewater. This method also is useful for evaluating the effects of combined sanitary and storm-sewer overflow on the water quality of urban streams. The method focuses on the determination of compounds that are indicators of wastewater or have endocrine-disrupting potential. These compounds include the alkylphenol ethoxylate nonionic surfactants, food additives, fragrances, antioxidants, flame retardants, plasticizers, industrial solvents, disinfectants, fecal sterols, polycyclic aromatic hydrocarbons, and high-use domestic pesticides. Wastewater compounds in whole-water samples were extracted using continuous liquid-liquid extractors and methylene chloride solvent, and then determined by capillary-column gas chromatography/mass spectrometry. Recoveries in reagent-water samples fortified at 0.5 microgram per liter averaged 72 percent ? 8 percent relative standard deviation. The concentration of 21 compounds is always reported as estimated because method recovery was less than 60 percent, variability was greater than 25 percent relative standard deviation, or standard reference compounds were prepared from technical mixtures. Initial method detection limits averaged 0.18 microgram per liter. Samples were preserved by adding 60 grams of sodium chloride and stored at 4 degrees Celsius. The laboratory established a sample holding-time limit prior to sample extraction of 14 days from the date of collection.

Guidance for laboratories performing molecular pathology for cancer patients.

PubMed

Cree, Ian A; Deans, Zandra; Ligtenberg, Marjolijn J L; Normanno, Nicola; Edsjö, Anders; Rouleau, Etienne; Solé, Francesc; Thunnissen, Erik; Timens, Wim; Schuuring, Ed; Dequeker, Elisabeth; Murray, Samuel; Dietel, Manfred; Groenen, Patricia; Van Krieken, J Han

2014-11-01

Molecular testing is becoming an important part of the diagnosis of any patient with cancer. The challenge to laboratories is to meet this need, using reliable methods and processes to ensure that patients receive a timely and accurate report on which their treatment will be based. The aim of this paper is to provide minimum requirements for the management of molecular pathology laboratories. This general guidance should be augmented by the specific guidance available for different tumour types and tests. Preanalytical considerations are important, and careful consideration of the way in which specimens are obtained and reach the laboratory is necessary. Sample receipt and handling follow standard operating procedures, but some alterations may be necessary if molecular testing is to be performed, for instance to control tissue fixation. DNA and RNA extraction can be standardised and should be checked for quality and quantity of output on a regular basis. The choice of analytical method(s) depends on clinical requirements, desired turnaround time, and expertise available. Internal quality control, regular internal audit of the whole testing process, laboratory accreditation, and continual participation in external quality assessment schemes are prerequisites for delivery of a reliable service. A molecular pathology report should accurately convey the information the clinician needs to treat the patient with sufficient information to allow for correct interpretation of the result. Molecular pathology is developing rapidly, and further detailed evidence-based recommendations are required for many of the topics covered here. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Mean centering of ratio spectra and concentration augmented classical least squares in a comparative approach for quantitation of spectrally overlapped bands of antihypertensives in formulations

NASA Astrophysics Data System (ADS)

Hegazy, Maha Abdel Monem; Fayez, Yasmin Mohammed

2015-04-01

Two different methods manipulating spectrophotometric data have been developed, validated and compared. One is capable of removing the signal of any interfering components at the selected wavelength of the component of interest (univariate). The other includes more variables and extracts maximum information to determine the component of interest in the presence of other components (multivariate). The applied methods are smart, simple, accurate, sensitive, precise and capable of determination of spectrally overlapped antihypertensives; hydrochlorothiazide (HCT), irbesartan (IRB) and candesartan (CAN). Mean centering of ratio spectra (MCR) and concentration residual augmented classical least-squares method (CRACLS) were developed and their efficiency was compared. CRACLS is a simple method that is capable of extracting the pure spectral profiles of each component in a mixture. Correlation was calculated between the estimated and pure spectra and was found to be 0.9998, 0.9987 and 0.9992 for HCT, IRB and CAN, respectively. The methods were successfully determined the three components in bulk powder, laboratory-prepared mixtures, and combined dosage forms. The results obtained were compared statistically with each other and to those of the official methods.

Intra-laboratory validation of chronic bee paralysis virus quantitation using an accredited standardised real-time quantitative RT-PCR method.

PubMed

Blanchard, Philippe; Regnault, Julie; Schurr, Frank; Dubois, Eric; Ribière, Magali

2012-03-01

Chronic bee paralysis virus (CBPV) is responsible for chronic bee paralysis, an infectious and contagious disease in adult honey bees (Apis mellifera L.). A real-time RT-PCR assay to quantitate the CBPV load is now available. To propose this assay as a reference method, it was characterised further in an intra-laboratory study during which the reliability and the repeatability of results and the performance of the assay were confirmed. The qPCR assay alone and the whole quantitation method (from sample RNA extraction to analysis) were both assessed following the ISO/IEC 17025 standard and the recent XP U47-600 standard issued by the French Standards Institute. The performance of the qPCR assay and of the overall CBPV quantitation method were validated over a 6 log range from 10(2) to 10(8) with a detection limit of 50 and 100 CBPV RNA copies, respectively, and the protocol of the real-time RT-qPCR assay for CBPV quantitation was approved by the French Accreditation Committee. Copyright © 2011 Elsevier B.V. All rights reserved.

A Laboratory Exercise to Understand the Importance of Enzyme Technology in the Fruit-Processing Industry: Viscosity Decrease and Phenols Release from Apple Mash

ERIC Educational Resources Information Center

Pinelo, Manuel; Nielsen, Michael K.; Meyer, Anne S.

2011-01-01

In a 4-h laboratory exercise, students accomplish a series of enzymatic macerations of apple mash, assess the viscosity of the mash during the maceration, extract the juice by centrifugation, and measure the levels of antioxidant phenols extracted into the juice after different enzyme treatments. The exercise shows the impact of enzyme-catalyzed…

Rapid determination of fumonisins in corn-based products by liquid chromatography/tandem mass spectrometry.

PubMed

Li, Wei; Herrman, Timothy J; Dai, Susie Y

2010-01-01

A simple, fast, and robust method was developed for the determination of fumonisin B1 (FB1), fumonisin B2 (FB2), and fumonisin B3 (FB3) in corn-based human food and animal feed (cornmeal). The method involves a single extraction step followed by centrifugation and filtration before analysis by ultra-performance liquid chromatographylelectrospray ionization (UPLC/ESI)-MS/MS. The LC/MS/MS method developed here represents the fastest and simplest procedure (<30 min) among both conventional HPLC methods and other LC/MS methods using SPE cleanup. The potential for high throughput analysis makes the method particularly beneficial for regulatory agencies and analytical laboratories with a high sample volume. A single-laboratory validation was conducted by testing three different spiking levels (200, 500, and 1000 ng/g for FB1 and FB2; 100, 250, and 500 ng/g for FB3) for accuracy and precision. Recoveries of FB1 ranged from 93 to 98% with RSD values of 3-8%. Recoveries of FB2 ranged from 104 to 108%, with RSD values of 2-6%. Recoveries of FB3 ranged from 94 to 108%, with RSD values of 2-5%.

Simultaneous Determination of Total Vitamins B1, B2, B3, and B6 in Infant Formula and Related Nutritionals by Enzymatic Digestion and LC-MS/MS: Single-Laboratory Validation, First Action 2015.14.

PubMed

Salvati, Louis M; McClure, Sean C; Reddy, Todime M; Cellar, Nicholas A

2016-05-01

This method provides simultaneous determination of total vitamins B1, B2, B3, and B6 in infant formula and related nutritionals (adult and infant). The method was given First Action for vitamins B1, B2, and B6, but not B3, during the AOAC Annual Meeting in September 2015. The method uses acid phosphatase to dephosphorylate the phosphorylated vitamin forms. It then measures thiamine (vitamin B1); riboflavin (vitamin B2); nicotinamide and nicotinic acid (vitamin B3); and pyridoxine, pyridoxal, and pyridoxamine (vitamin B6) from digested sample extract by liquid chromatography-tandem mass spectrometry. A single-laboratory validation was performed on 14 matrixes provided by the AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN) to demonstrate method effectiveness. The method met requirements of the AOAC SPIFAN Standard Method Performance Requirement for each of the three vitamins, including average over-spike recovery of 99.6 ± 3.5%, average repeatability of 1.5 ± 0.8% relative standard deviation, and average intermediate precision of 3.9 ± 1.3% relative standard deviation.

European validation of Real-Time PCR method for detection of Salmonella spp. in pork meat.

PubMed

Delibato, Elisabetta; Rodriguez-Lazaro, David; Gianfranceschi, Monica; De Cesare, Alessandra; Comin, Damiano; Gattuso, Antonietta; Hernandez, Marta; Sonnessa, Michele; Pasquali, Frédérique; Sreter-Lancz, Zuzsanna; Saiz-Abajo, María-José; Pérez-De-Juan, Javier; Butrón, Javier; Prukner-Radovcic, Estella; Horvatek Tomic, Danijela; Johannessen, Gro S; Jakočiūnė, Džiuginta; Olsen, John E; Chemaly, Marianne; Le Gall, Francoise; González-García, Patricia; Lettini, Antonia Anna; Lukac, Maja; Quesne, Segolénè; Zampieron, Claudia; De Santis, Paola; Lovari, Sarah; Bertasi, Barbara; Pavoni, Enrico; Proroga, Yolande T R; Capuano, Federico; Manfreda, Gerardo; De Medici, Dario

2014-08-01

The classical microbiological method for detection of Salmonella spp. requires more than five days for final confirmation, and consequently there is a need for an alternative methodology for detection of this pathogen particularly in those food categories with a short shelf-life. This study presents an international (at European level) ISO 16140-based validation study of a non-proprietary Real-Time PCR-based method that can generate final results the day following sample analysis. It is based on an ISO compatible enrichment coupled to an easy and inexpensive DNA extraction and a consolidated Real-Time PCR assay. Thirteen laboratories from seven European Countries participated to this trial, and pork meat was selected as food model. The limit of detection observed was down to 10 CFU per 25 g of sample, showing excellent concordance and accordance values between samples and laboratories (100%). In addition, excellent values were obtained for relative accuracy, specificity and sensitivity (100%) when the results obtained for the Real-Time PCR-based methods were compared to those of the ISO 6579:2002 standard method. The results of this international trial demonstrate that the evaluated Real-Time PCR-based method represents an excellent alternative to the ISO standard. In fact, it shows an equal and solid performance as well as it reduces dramatically the extent of the analytical process, and can be easily implemented routinely by the Competent Authorities and Food Industry laboratories. Copyright © 2014 Elsevier B.V. All rights reserved.

Comparison of four DNA extraction methods for the detection of Mycobacterium leprae from Ziehl-Neelsen-stained microscopic slides.

PubMed

Ruiz-Fuentes, Jenny Laura; Díaz, Alexis; Entenza, Anayma Elena; Frión, Yahima; Suárez, Odelaisy; Torres, Pedro; de Armas, Yaxsier; Acosta, Lucrecia

2015-12-01

The diagnosis of leprosy has been a challenge due to the low sensibility of the conventional methods and the impossibility of culturing the causative organism. In this study, four methods for Mycobacterium leprae nucleic-acid extraction from Ziehl-Neelsen-stained slides (ZNS slides) were compared: Phenol/chloroform, Chelex 100 resin, and two commercial kits (Wizard Genomic DNA Purification Kit and QIAamp DNA Mini Kit). DNA was extracted from four groups of slides: a high-codification-slide group (bacteriological index [BI]⩾4), a low-codification-slide group (BI=1), a negative-slide group (BI=0), and a negative-control-slide group (BI=0). Quality DNA was evidenced by the amplification of specific repetitive element present in M. leprae genomic DNA (RLEP) using a nested polymerase chain reaction. This is the first report comparing four different extraction methods for obtaining M. leprae DNA from ZNS slides in Cuban patients, and applied in molecular diagnosis. Good-quality DNA and positive amplification were detected in the high-codification-slide group with the four methods, while from the low-codification-slide group only the QIAGEN and phenol-chloroform methods obtained amplification of M. leprae. In the negative-slide group, only the QIAGEN method was able to obtain DNA with sufficient quality for positive amplification of the RLEP region. No amplification was observed in the negative-control-slide group by any method. Patients with ZNS negative slides can still transmit the infection, and molecular methods can help identify and treat them, interrupting the chain of transmission and preventing the onset of disabilities. The ZNS slides can be sent easily to reference laboratories for later molecular analysis that can be useful not only to improve the diagnosis, but also for the application of other molecular techniques. Copyright © 2015 Asian-African Society for Mycobacteriology. Published by Elsevier Ltd. All rights reserved.

Rapid method for the simultaneous detection of boar taint compounds by means of solid phase microextraction coupled to gas chromatography/mass spectrometry.

PubMed

Verplanken, Kaat; Wauters, Jella; Van Durme, Jim; Claus, Dirk; Vercammen, Joeri; De Saeger, Sarah; Vanhaecke, Lynn

2016-09-02

Because of animal welfare issues, the voluntary ban on surgical castration of male piglets, starting January 2018 was announced in a European Treaty. One viable alternative is the fattening of entire male pigs. However, this can cause negative consumer reactions due to the occurrence of boar taint and possibly lead to severe economic losses in pig husbandry. In this study, headspace solid phase microextraction (HS-SPME) coupled to GC-MS was used in the development and optimization of a candidate method for fast and accurate detection of the boar taint compounds. Remarkably fast extraction (45s) of the boar taint compounds from adipose tissue was achieved by singeing the fat with a soldering iron while released volatiles were extracted in-situ using HS-SPME. The obtained method showed good performance characteristics after validation according to CD 2002/657/EC and ISO/IEC 17025 guidelines. Moreover, cross-validation with an in-house UHPLC-HR-Orbitrap-MS method showed good agreement between an in-laboratory method and the new candidate method for the fast extraction and detection of skatole and androstenone, which emphasizes the accuracy of this new SPME-GC-MS method. Threshold detection of the boar taint compounds on a portable GC-MS could not be achieved. However, despite the lack of sensitivity obtained on the latter instrument, a very fast method with run-to-run time of 3.5min for the detection of the boar taint compounds was developed. Copyright © 2016 Elsevier B.V. All rights reserved.

Screening and confirmation of steroids and nitroimidazoles in urine, blood, and food matrices: Sample preparation methods and liquid chromatography tandem mass spectrometric separations.

PubMed

Tölgyesi, Ádám; Barta, Enikő; Simon, Andrea; McDonald, Thomas J; Sharma, Virender K

2017-10-25

Veterinary drugs containing synthetic anabolic steroid and nitroimidazole active agents are not allowed for their applications in livestock of the European Union (EU). This paper presents analyses of twelve selected steroids and six nitroimidazole antibiotics at low levels (1.56μg/L-4.95μg/L and 0.17μg/kg-2.14μg/kg, respectively) in body fluids and egg incurred samples. Analyses involved clean-up procedures, high performance liquid chromatography (HPLC) separation, and tandem mass spectrometric screening and confirmatory methods. Target steroids and nitroimidazoles in samples were cleaned by two independent supported liquid extraction and solid phase extraction procedures. Separation of the selected compounds was conducted on Kinetex XB C-18 HPLC column using gradient elution. The screening methods utilised supported liquid extraction that enabled fast and cost effective clean-up. The confirmatory methods were improved by extending the number of matrices and compounds, and by introducing an isotope dilution mass spectrometry for nitroimidazoles. The new methods were validated according to the recommendation of the European Union Reference Laboratories and the performance characteristics evaluated met fully the criteria. The methods were applied to incurred samples in the proficiency tests. The obtained results of Z-scores demonstrated the applicability of developed protocols of the methods to real samples. The confirmatory methods were applied to the national monitoring program and natural contamination of prednisolone could be detected in urine at low concentration in few samples. Copyright © 2017 Elsevier B.V. All rights reserved.

Automated solid-phase extraction-liquid chromatography-tandem mass spectrometry analysis of 6-acetylmorphine in human urine specimens: application for a high-throughput urine analysis laboratory.

PubMed

Robandt, P V; Bui, H M; Scancella, J M; Klette, K L

2010-10-01

An automated solid-phase extraction-liquid chromatography- tandem mass spectrometry (SPE-LC-MS-MS) method using the Spark Holland Symbiosis Pharma SPE-LC coupled to a Waters Quattro Micro MS-MS was developed for the analysis of 6-acetylmorphine (6-AM) in human urine specimens. The method was linear (R² = 0.9983) to 100 ng/mL, with no carryover at 200 ng/mL. Limits of quantification and detection were found to be 2 ng/mL. Interrun precision calculated as percent coefficient of variation (%CV) and evaluated by analyzing five specimens at 10 ng/mL over nine batches (n = 45) was 3.6%. Intrarun precision evaluated from 0 to 100 ng/mL ranged from 1.0 to 4.4%CV. Other opioids (codeine, morphine, oxycodone, oxymorphone, hydromorphone, hydrocodone, and norcodeine) did not interfere in the detection, quantification, or chromatography of 6-AM or the deuterated internal standard. The quantified values for 41 authentic human urine specimens previously found to contain 6-AM by a validated gas chromatography (GC)-MS method were compared to those obtained by the SPE-LC-MS-MS method. The SPE-LC-MS-MS procedure eliminates the human factors of specimen handling, extraction, and derivatization, thereby reducing labor costs and rework resulting from human error or technique issues. The time required for extraction and analysis was reduced by approximately 50% when compared to a validated 6-AM procedure using manual SPE and GC-MS analysis.

Measurement of water-soluble B vitamins in infant formula by liquid chromatography/tandem mass spectrometry.

PubMed

Huang, Min; Winters, Doug; Crowley, Richard; Sullivan, Darryl

2009-01-01

A method has been developed for the simultaneous measurement of multiple B vitamins (i.e., B1, B2, B3, B5, and B6) in infant formulas by LC-MSIMS. The vitamins were extracted with acidic solvent, followed by protein precipitation at a pH range of 4.5 to 5.5, and filtered. This simplified procedure eliminates many of the potential sources of laboratory error and facilitates rapid and efficient analysis. As is common in most cases, isotope internal standards were added to account for variations in sample preparation, as well as changes in MS measurement. In this method, isotope-labeled internal standards of B1, B3, B5, and B6 were used. The factors affecting analytical performance were investigated and optimized. In addition, the stability of these vitamins in the extraction solution was investigated. An acidic condition (5 mM HCl) was applied to successfully stabilize B1, which had shown a decrease in signal when other solvents were used. The quantitative extraction and good stability allowed isotope standards to be added to the filtered sample solution, instead of to the extraction solvent. The addition of the isotope to the small portion of the filtered sample solution significantly reduces cost. A comprehensive evaluation of the analysis of the standard reference material and good spike recovery of the vitamins (100 +/- 6%) demonstrates the accuracy of the method. The results for commercially available infant formula samples were also compared with those obtained using the current microbiological method.

Differences among heat-treated, raw, and commercial peanut extracts by skin testing and immunoblotting.

PubMed

Maleki, Soheila J; Casillas, Adrian M; Kaza, Ujwala; Wilson, Brian A; Nesbit, Jacqueline B; Reimoneqnue, Chantrel; Cheng, Hsiaopo; Bahna, Sami L

2010-12-01

Peanut allergenicity has been reported to be influenced by heat treatment, yet the commonly available extracts for skin prick testing (SPT) are derived from raw extracts. To assess the effect of heat treatment on the SPT reactivity and specific IgE binding to peanut. Three commercial extracts and 3 laboratory-prepared extracts, including raw, roasted, and boiled, were used for SPT in 19 patients with suspected peanut allergy and in 4 individuals who eat peanut without any symptoms. Serum samples were obtained to measure total IgE in addition to specific IgE binding to the study extracts by immunoblotting. Peanut allergy was confirmed with challenge test unless the individual had a convincing history of a severe reaction. Eleven study participants were considered peanut allergic based on a strong history or positive challenge test result. SPT with the prepared and commercial reagents showed that the boiled extract had the highest specificity (67% vs 42%-63% for the other extracts). The prepared extracts showed similar SPT sensitivity (81%). Three patients with a history of severe reaction and elevated specific IgE levels to peanut to the 3 study extracts had variable SPT reactivity to 1 or more of the commercial extracts. IgE binding to Ara h 2 was found in nearly all patients, regardless of their clinical reactivity. None of the extracts tested showed optimal diagnostic reliability regarding both sensitivity and specificity. Perhaps testing should be performed with multiple individual extracts prepared by different methods. Copyright © 2010 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Standard Operating Procedures and Field Methods Used for Conducting Ecological Risk Assessment Case Studies. Naval Construction Battalion Center, Davisville, Rhode Island, and Naval Shipyard, Portsmouth, Kittery, Maine.

DTIC Science & Technology

1992-05-01

Studies by Craig et al. 20 have chloride extraction/Grignard derivatization method. shown that significant evaporative losses of TBT + Methylene...and Craig . P1J Anahiw. 1987. 112: 1781 available from IEEE Service Center. 445 Hoes tLane. 21/ Aue. W A and Flinn. C G J Cltroinato-er.. 1977, 142 145...Environmental Research Laboratory Narragansett, Narragansett, RI. Keith, L.H., W. Crummett, J. Deegan , Jr, R.A. Libby, J.K. Taylor, and B. Wentler, 1983. Annal

Cytotoxic and aryl hydrocarbon hydroxylase-inducing effects of laboratory rodent diets. A cell culture study

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

Toerroenen, R.; Pelkonen, K.; Kaerenlampi, S.

1991-01-01

Extracts of several rodent diets were studied for their cytotoxic and aryl hydrocarbon hydroxylase-inducing properties by an in vitro method. The cell culture system based on a mouse hepatoma cell line (Hepa-1) was shown to be convenient and sensitive method for screening of diets for these parameters implying the presence of compounds potentially harmful in vivo. Considerable differences among diets and batches were detected. Smallest effects were observed with a semipurified diet and with the unrefined diet which - contrary to other four unrefined diets - contained no fish.

Thermal oil recovery method using self-contained windelectric sets

NASA Astrophysics Data System (ADS)

Belsky, A. A.; Korolyov, I. A.

2018-05-01

The paper reviews challenges associated with questions of efficiency of thermal methods of impact on productive oil strata. The concept of using electrothermal complexes with WEG power supply for the indicated purposes was proposed and justified, their operating principles, main advantages and disadvantages, as well as a schematechnical solution for the implementation of the intensification of oil extraction, were considered. A mathematical model for finding the operating characteristics of WEG is presented and its main energy parameters are determined. The adequacy of the mathematical model is confirmed by laboratory simulation stand tests with nominal parameters.

Doping control analysis of seven bioactive peptides in horse plasma by liquid chromatography-mass spectrometry.

PubMed

Kwok, Wai Him; Ho, Emmie N M; Lau, Ming Yip; Leung, Gary N W; Wong, April S Y; Wan, Terence S M

2013-03-01

In recent years, there has been an ongoing focus for both human and equine doping control laboratories on developing detection methods to control the misuse of peptide therapeutics. Immunoaffinity purification is a common extraction method to isolate peptides from biological matrices and obtain sufficient detectability in subsequent instrumental analysis. However, monoclonal or polyclonal antibodies for immunoaffinity purification may not be commercially available, and even if available, such antibodies are usually very costly. In our study, a simple mixed-mode anion exchange solid-phase extraction cartridge was employed for the extraction of seven target peptides (GHRP-1, GHRP-2, GHRP-6, ipamorelin, hexarelin, CJC-1295, and N-acetylated LKKTETQ (active ingredient of TB-500)) and their in vitro metabolites from horse plasma. The final extract was subject to ultra-high-performance liquid chromatographic separation and analysed with a hybrid high-resolution mass spectrometer. The limits of detection for all seven peptides were estimated to be less than 50 pg/mL. Method validation was performed with respect to specificity, precision, and recovery. The applicability of this multi-analyte method was demonstrated by the detection of N-acetylated LKKTETQ and its metabolite N-acetylated LK from plasma samples obtained after subcutaneous administration of TB-500 (10 mg N-acetylated LKKTETQ) to two thoroughbred geldings. This method could easily be modified to cover more bioactive peptides, such as dermorphin, β-casomorphin, and desmopressin. With the use of high-resolution mass spectrometry, the full-scan data acquired can also be re-processed retrospectively to search for peptides and their metabolites that have not been targeted at the time of analysis. To our knowledge, this is the first identification of in vitro metabolites of all the studied peptides other than TB-500 in horses.

Application of supercritical fluid chromatography coupled to mass spectrometry to the determination of fat-soluble vitamins in selected food products.

PubMed

Oberson, Jean-Marie; Campos-Giménez, Esther; Rivière, Johann; Martin, Frédéric

2018-06-01

In the present manuscript, we describe a fully optimized and validated method suitable to analyse nine compounds (retinyl acetate, retinyl palmitate, retinol, α-tocopherol, α-tocopheryl acetate, cholecalciferol, ergocalciferol, phylloquinone, menaquinone-4) representing the major contributors to the fat-soluble vitamin activity of selected food products (infant formulas, adult nutritionals, infant cereals and mixed meals). Sample preparation involves direct solvent extraction using enzyme-assisted matrix disintegration and methanolic protein precipitation. Direct injection of the extract allows quantification of vitamins A, E and K in only 7 min, while vitamin D is determined after fast derivatization of the extract. Separation is achieved by supercritical fluid chromatography and detection performed by tandem mass spectrometry in positive Atmospheric Pressure Chemical Ionization mode. Results on a Standard Reference Material (SRM 1849a Infant/Adult Nutritional) were not statistically different from reference values. Full validation of the method showed excellent overall performance. Average recovery rate was between 90 and 110% for all vitamins and matrixes. The methodology shows enhanced safety and reduced cost as compared with previously published methods, together with potential for application to more complex matrixes. The full procedure can be easily applied in control laboratories dramatically increasing sample throughput and reducing solvent consumption. Copyright © 2018 Elsevier B.V. All rights reserved.

Field Application of a Rapid Spectrophotometric Method for Determination of Persulfate in Soil

PubMed Central

Cunningham, Colin J.; Pitschi, Vanessa; Anderson, Peter; Barry, D. A.; Patterson, Colin; Peshkur, Tanya A.

2013-01-01

Remediation of hydrocarbon contaminated soils can be performed both in situ and ex situ using chemical oxidants such as sodium persulfate. Standard methods for quantifying persulfate require either centrifugation or prolonged settling times. An optimized soil extraction procedure was developed for persulfate involving simple water extraction using a modified disposable syringe. This allows considerable saving of time and removes the need for centrifugation. The extraction time was reduced to only 5 min compared to 15 min for the standard approach. A comparison of the two approaches demonstrated that each provides comparable results. Comparisons were made using high (93 g kg−1 soil) and low (9.3 g kg−1 soil) additions of sodium persulfate to a petroleum hydrocarbon-contaminated soil, as well as sand spiked with diesel. Recoveries of 95±1% and 96±10% were observed with the higher application rate in the contaminated soil and spiked sand, respectively. Corresponding recoveries of 86±5% and 117±19% were measured for the lower application rate. Results were obtained in only 25 min and the method is well suited to batch analyses. In addition, it is suitable for application in a small field laboratory or even a mobile, vehicle-based system, as it requires minimal equipment and reagents. PMID:23776446

Screening of crude extracts of six medicinal plants used in South-West Nigerian unorthodox medicine for anti-methicillin resistant Staphylococcus aureus activity

PubMed Central

Akinyemi, Kabir O; Oladapo, Olukayode; Okwara, Chidi E; Ibe, Christopher C; Fasure, Kehinde A

2005-01-01

Background Six Nigerian medicinal plants Terminalia avicennioides, Phylantus discoideus, Bridella ferruginea, Ageratum conyzoides, Ocimum gratissimum and Acalypha wilkesiana used by traditional medical practitioners for the treatment of several ailments of microbial and non-microbial origins were investigated for in vitro anti-methicillin Resistant Staphylococcus aureus (MRSA) activity. Methods Fresh plant materials were collected from the users. Water and ethanol extracts of the shredded plants were obtained by standard methods. The Bacterial cultures used were strains of MRSA isolated from patients. MRSA was determined by the reference broth microdilution methods using the established National Committee for Clinical Laboratory Standards break points. Staphylococcus aureus NCIB 8588 was used as a standard strain. Susceptibility testing and phytochemical screening of the plant extracts were performed by standard procedures. Controls were maintained for each test batch. Results Both water and ethanol extracts of T. avicennioides, P. discoideus, O. gratissimum, and A. wilkesiana were effective on MRSA. The Minimum Inhibition Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of the ethanol extracts of these plants range from 18.2 to 24.0 mcg/ml and 30.4 to 37.0 mcg/ml respectively. In contrast, MIC range of 30.6 to 43.0 mcg/ml and 55.4 to 71.0 mcg/ml were recorded for ethanol and water extracts of B. ferruginea, and A. conyzoides respectively. Higher MBC values were obtained for the two plants. These concentrations were too high to be considered active in this study. All the four active plants contained at least trace amount of anthraquinones. Conclusion Our results offer a scientific basis for the traditional use of water and ethanol extracts of A. wilkesiana, O. gratissimum, T. avicennioides and P. discoideus against MRSA-associated diseases. However, B. ferruginea and A. conyzoides were ineffective in vitro in this study; we therefore suggest the immediate stoppage of their traditional use against MRSA-associated diseases in Lagos, Nigeria. PMID:15762997

Development and Single-Laboratory Validation of a Liquid Chromatography Tandem Mass Spectrometry Method for Quantitation of Tetrodotoxin in Mussels and Oysters.

PubMed

Turner, Andrew D; Boundy, Michael J; Rapkova, Monika Dhanji

2017-09-01

In recent years, evidence has grown for the presence of tetrodotoxin (TTX) in bivalve mollusks, leading to the potential for consumers of contaminated products to be affected by Tetrodotoxin Shellfish Poisoning (TSP). A single-laboratory validation was conducted for the hydrophilic interaction LC (HILIC) tandem MS (MS/MS) analysis of TTX in common mussels and Pacific oysters-the bivalve species that have been found to contain TTXs in the United Kingdom in recent years. The method consists of a single-step dispersive extraction in 1% acetic acid, followed by a carbon SPE cleanup step before dilution and instrumental analysis. The full method was developed as a rapid tool for the quantitation of TTX, as well as for the associated analogs 4-epi-TTX; 5,6,11-trideoxy TTX; 11-nor TTX-6-ol; 5-deoxy TTX; and 4,9-anhydro TTX. The method can also be run as the acquisition of TTX together with paralytic shellfish toxins. Results demonstrated acceptable method performance characteristics for specificity, linearity, recovery, ruggedness, repeatability, matrix variability, and within-laboratory reproducibility for the analysis of TTX. The LOD and LOQ were fit-for-purpose in comparison to the current action limit for TTX enforced in The Netherlands. In addition, aspects of method performance (LOD, LOQ, and within-laboratory reproducibility) were found to be satisfactory for three other TTX analogs (11-nor TTX-6-ol, 5-deoxy TTX, and 4,9-anhydro TTX). The method was found to be practical and suitable for use in regulatory testing, providing rapid turnaround of sample analysis. Plans currently underway on a full collaborative study to validate a HILIC-MS/MS method for paralytic shellfish poisoning toxins will be extended to include TTX in order to generate international acceptance, ultimately for use as an alternative official control testing method should regulatory controls be adopted.

Composite Magnetic Nanoparticles (CuFe₂O₄) as a New Microsorbent for Extraction of Rhodamine B from Water Samples.

PubMed

Roostaie, Ali; Allahnoori, Farzad; Ehteshami, Shokooh

2017-09-01

In this work, novel composite magnetic nanoparticles (CuFe2O4) were synthesized based on sol-gel combustion in the laboratory. Next, a simple production method was optimized for the preparation of the copper nanoferrites (CuFe2O4), which are stable in water, magnetically active, and have a high specific area used as sorbent material for organic dye extraction in water solution. CuFe2O4 nanopowders were characterized by field-emission scanning electron microscopy (SEM), FTIR spectroscopy, and energy dispersive X-ray spectroscopy. The size range of the nanoparticles obtained in such conditions was estimated by SEM images to be 35-45 nm. The parameters influencing the extraction of CuFe2O4 nanoparticles, such as desorption solvent, amount of sorbent, desorption time, sample pH, ionic strength, and extraction time, were investigated and optimized. Under the optimum conditions, a linear calibration curve in the range of 0.75-5.00 μg/L with R2 = 0.9996 was obtained. The LOQ (10Sb) and LOD (3Sb) of the method were 0.75 and 0.25 μg/L (n = 3), respectively. The RSD for a water sample spiked with 1 μg/L rhodamine B was 3% (n = 5). The method was applied for the determination of rhodamine B in tap water, dishwashing foam, dishwashing liquid, and shampoo samples. The relative recovery percentages for these samples were in the range of 95-99%.

Rapid filtration separation-based sample preparation method for Bacillus spores in powdery and environmental matrices.

PubMed

Isabel, Sandra; Boissinot, Maurice; Charlebois, Isabelle; Fauvel, Chantal M; Shi, Lu-E; Lévesque, Julie-Christine; Paquin, Amélie T; Bastien, Martine; Stewart, Gale; Leblanc, Eric; Sato, Sachiko; Bergeron, Michel G

2012-03-01

Authorities frequently need to analyze suspicious powders and other samples for biothreat agents in order to assess environmental safety. Numerous nucleic acid detection technologies have been developed to detect and identify biowarfare agents in a timely fashion. The extraction of microbial nucleic acids from a wide variety of powdery and environmental samples to obtain a quality level adequate for these technologies still remains a technical challenge. We aimed to develop a rapid and versatile method of separating bacteria from these samples and then extracting their microbial DNA. Bacillus atrophaeus subsp. globigii was used as a simulant of Bacillus anthracis. We studied the effects of a broad variety of powdery and environmental samples on PCR detection and the steps required to alleviate their interference. With a benchmark DNA extraction procedure, 17 of the 23 samples investigated interfered with bacterial lysis and/or PCR-based detection. Therefore, we developed the dual-filter method for applied recovery of microbial particles from environmental and powdery samples (DARE). The DARE procedure allows the separation of bacteria from contaminating matrices that interfere with PCR detection. This procedure required only 2 min, while the DNA extraction process lasted 7 min, for a total of <10 min. This sample preparation procedure allowed the recovery of cleaned bacterial spores and relieved detection interference caused by a wide variety of samples. Our procedure was easily completed in a laboratory facility and is amenable to field application and automation.

Rapid Filtration Separation-Based Sample Preparation Method for Bacillus Spores in Powdery and Environmental Matrices

PubMed Central

Isabel, Sandra; Boissinot, Maurice; Charlebois, Isabelle; Fauvel, Chantal M.; Shi, Lu-E; Lévesque, Julie-Christine; Paquin, Amélie T.; Bastien, Martine; Stewart, Gale; Leblanc, Éric; Sato, Sachiko

2012-01-01

Authorities frequently need to analyze suspicious powders and other samples for biothreat agents in order to assess environmental safety. Numerous nucleic acid detection technologies have been developed to detect and identify biowarfare agents in a timely fashion. The extraction of microbial nucleic acids from a wide variety of powdery and environmental samples to obtain a quality level adequate for these technologies still remains a technical challenge. We aimed to develop a rapid and versatile method of separating bacteria from these samples and then extracting their microbial DNA. Bacillus atrophaeus subsp. globigii was used as a simulant of Bacillus anthracis. We studied the effects of a broad variety of powdery and environmental samples on PCR detection and the steps required to alleviate their interference. With a benchmark DNA extraction procedure, 17 of the 23 samples investigated interfered with bacterial lysis and/or PCR-based detection. Therefore, we developed the dual-filter method for applied recovery of microbial particles from environmental and powdery samples (DARE). The DARE procedure allows the separation of bacteria from contaminating matrices that interfere with PCR detection. This procedure required only 2 min, while the DNA extraction process lasted 7 min, for a total of <10 min. This sample preparation procedure allowed the recovery of cleaned bacterial spores and relieved detection interference caused by a wide variety of samples. Our procedure was easily completed in a laboratory facility and is amenable to field application and automation. PMID:22210204

Amino acid distribution in meteorites: diagenesis, extraction methods, and standard metrics in the search for extraterrestrial biosignatures.

PubMed

McDonald, Gene D; Storrie-Lombardi, Michael C

2006-02-01

The relative abundance of the protein amino acids has been previously investigated as a potential marker for biogenicity in meteoritic samples. However, these investigations were executed without a quantitative metric to evaluate distribution variations, and they did not account for the possibility of interdisciplinary systematic error arising from inter-laboratory differences in extraction and detection techniques. Principal component analysis (PCA), hierarchical cluster analysis (HCA), and stochastic probabilistic artificial neural networks (ANNs) were used to compare the distributions for nine protein amino acids previously reported for the Murchison carbonaceous chondrite, Mars meteorites (ALH84001, Nakhla, and EETA79001), prebiotic synthesis experiments, and terrestrial biota and sediments. These techniques allowed us (1) to identify a shift in terrestrial amino acid distributions secondary to diagenesis; (2) to detect differences in terrestrial distributions that may be systematic differences between extraction and analysis techniques in biological and geological laboratories; and (3) to determine that distributions in meteoritic samples appear more similar to prebiotic chemistry samples than they do to the terrestrial unaltered or diagenetic samples. Both diagenesis and putative interdisciplinary differences in analysis complicate interpretation of meteoritic amino acid distributions. We propose that the analysis of future samples from such diverse sources as meteoritic influx, sample return missions, and in situ exploration of Mars would be less ambiguous with adoption of standardized assay techniques, systematic inclusion of assay standards, and the use of a quantitative, probabilistic metric. We present here one such metric determined by sequential feature extraction and normalization (PCA), information-driven automated exploration of classification possibilities (HCA), and prediction of classification accuracy (ANNs).

Separated kaon electroproduction cross section and the kaon form factor from 6 GeV JLab data

DOE PAGES

Carmignotto, M.; Ali, S.; Aniol, K.; ...

2018-02-28

The 1H(e,e 'K +)Λ reaction was studied as a function of the Mandelstam variable -t using data from the E01-004 (FPI-2) and E93-018 experiments that were carried out in Hall C at the 6 GeV Jefferson Laboratory. The cross section was fully separated into longitudinal and transverse components, and two interference terms at four-momentum transfers Q 2 of 1.00, 1.36, and 2.07 GeV 2. The kaon form factor was extracted from the longitudinal cross section using the Regge model by Vanderhaeghen et al. [Phys. Rev. C 57, 1454 (1998)]. Here, the results establish the method, previously used successfully for pionmore » analyses, for extracting the kaon form factor. Data from 12 GeV Jefferson Laboratory experiments are expected to have sufficient precision to distinguish between theoretical predictions, for example, recent perturbative QCD calculations with modern parton distribution amplitudes. The leading-twist behavior for light mesons is predicted to set in for values of Q 2 between 5 and 10 GeV 2, which makes data in the few-GeV regime particularly interesting. Finally, the Q 2 dependence at fixed x and -t of the longitudinal cross section that we extracted seems consistent with the QCD factorization prediction within the experimental uncertainty.« less

Separated kaon electroproduction cross section and the kaon form factor from 6 GeV JLab data

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

Carmignotto, M.; Ali, S.; Aniol, K.

The 1H(e,e 'K +)Λ reaction was studied as a function of the Mandelstam variable -t using data from the E01-004 (FPI-2) and E93-018 experiments that were carried out in Hall C at the 6 GeV Jefferson Laboratory. The cross section was fully separated into longitudinal and transverse components, and two interference terms at four-momentum transfers Q 2 of 1.00, 1.36, and 2.07 GeV 2. The kaon form factor was extracted from the longitudinal cross section using the Regge model by Vanderhaeghen et al. [Phys. Rev. C 57, 1454 (1998)]. Here, the results establish the method, previously used successfully for pionmore » analyses, for extracting the kaon form factor. Data from 12 GeV Jefferson Laboratory experiments are expected to have sufficient precision to distinguish between theoretical predictions, for example, recent perturbative QCD calculations with modern parton distribution amplitudes. The leading-twist behavior for light mesons is predicted to set in for values of Q 2 between 5 and 10 GeV 2, which makes data in the few-GeV regime particularly interesting. Finally, the Q 2 dependence at fixed x and -t of the longitudinal cross section that we extracted seems consistent with the QCD factorization prediction within the experimental uncertainty.« less

Separated kaon electroproduction cross section and the kaon form factor from 6 GeV JLab data

NASA Astrophysics Data System (ADS)

Carmignotto, M.; Ali, S.; Aniol, K.; Arrington, J.; Barrett, B.; Beise, E. J.; Blok, H. P.; Boeglin, W.; Brash, E. J.; Breuer, H.; Chang, C. C.; Christy, M. E.; Dittmann, A.; Ent, R.; Fenker, H.; Gaskell, D.; Gibson, E.; Holt, R. J.; Horn, T.; Huber, G. M.; Jin, S.; Jones, M. K.; Keppel, C. E.; Kim, W.; King, P. M.; Kovaltchouk, V.; Liu, J.; Lolos, G. J.; Mack, D. J.; Margaziotis, D. J.; Markowitz, P.; Matsumura, A.; Meekins, D.; Miyoshi, T.; Mkrtchyan, H.; Niculescu, G.; Niculescu, I.; Okayasu, Y.; Pegg, I. L.; Pentchev, L.; Perdrisat, C.; Potterveld, D.; Punjabi, V.; Reimer, P. E.; Reinhold, J.; Roche, J.; Sarty, A.; Smith, G. R.; Tadevosyan, V.; Tang, L. G.; Trotta, R.; Tvaskis, V.; Vargas, A.; Vidakovic, S.; Volmer, J.; Vulcan, W.; Warren, G.; Wood, S. A.; Xu, C.; Zheng, X.; JLAB FPI-2; E93-018 Collaboration

2018-02-01

The 1H(e ,e'K+ )Λ reaction was studied as a function of the Mandelstam variable -t using data from the E01-004 (FPI-2) and E93-018 experiments that were carried out in Hall C at the 6 GeV Jefferson Laboratory. The cross section was fully separated into longitudinal and transverse components, and two interference terms at four-momentum transfers Q2 of 1.00, 1.36, and 2.07 GeV2. The kaon form factor was extracted from the longitudinal cross section using the Regge model by Vanderhaeghen et al. [Phys. Rev. C 57, 1454 (1998), 10.1103/PhysRevC.57.1454]. The results establish the method, previously used successfully for pion analyses, for extracting the kaon form factor. Data from 12 GeV Jefferson Laboratory experiments are expected to have sufficient precision to distinguish between theoretical predictions, for example, recent perturbative QCD calculations with modern parton distribution amplitudes. The leading-twist behavior for light mesons is predicted to set in for values of Q2 between 5 and 10 GeV2, which makes data in the few-GeV regime particularly interesting. The Q2 dependence at fixed x and -t of the longitudinal cross section that we extracted seems consistent with the QCD factorization prediction within the experimental uncertainty.

Analysis of tacrolimus and creatinine from a single dried blood spot using liquid chromatography tandem mass spectrometry.

PubMed

Koop, Dennis R; Bleyle, Lisa A; Munar, Myrna; Cherala, Ganesh; Al-Uzri, Amira

2013-05-01

Long term therapeutic drug monitoring and assessment of renal function are required in renal transplant recipients on immunosuppressant therapy such as tacrolimus. Dry blood spots (DBS) have been used successfully in the clinic for many years and offers a convenient, simple and non-invasive method for repeated blood tests. We developed and performed a preliminary validation of a method for the analysis of tacrolimus and creatinine from a single DBS using liquid chromatography-tandem mass spectrometric (LC-MS/MS). Tacrolimus and creatinine were extracted from a 6mm punch with a mixture of methanol/acetonitrile containing ascomycin and deuterated creatinine as internal standards. A 10 μl aliquot of the extract was analyzed directly after dilution for creatinine with normal phase high performance liquid chromatography and multiple reaction monitoring. The remainder of the extract was processed and analyzed for tacrolimus. The lower limit of quantification for tacrolimus was 1 ng/ml with accuracy of 0.34% bias and precision (CV) of 11.1%. The precision ranged from 1.33% to 7.68% and accuracy from -4.44% to 11.6% bias for the intra- and inter-day analysis. The lower limit of quantification of creatinine was 0.01 mg/dL with precision of 7.94%. Accuracy was based on recovery of additional creatinine spiked into whole blood samples and ranged from -2.45% bias at 5 mg/dL to 3.75% bias at 0.5 mg/dL. Intra- and inter-day precision was from 3.48 to 4.11%. The assay was further validated with DBS prepared from pediatric renal transplant recipients. There was excellent correlation between the levels of tacrolimus and creatinine obtained from the clinical laboratory and the DBS method developed. After additional validation, this assay may have a significant impact on compliance with medication intake as well as potentially lowering the cost associated with intravenous blood draws in clinical laboratories. Copyright © 2013 Elsevier B.V. All rights reserved.