Biogenic amine formation and bacterial contribution in Natto products.

PubMed

Kim, Bitna; Byun, Bo Young; Mah, Jae-Hyung

2012-12-01

Twenty-one Natto products currently distributed in Korea were analysed for biogenic amine contents and tested to determine physicochemical and bacterial contributions to biogenic amine formation. Among them, nine products (about 43%) had β-phenylethylamine or tyramine contents greater than the toxic dose (30mg/kg and 100mg/kg, respectively) of each amine, although no products showed total amounts of biogenic amines above the harmful level (1000mg/kg), which indicates that the amounts of biogenic amines in some Natto products are not within the safe level for human health. From four different Natto products, that contained noticeable levels of β-phenylethylamine and tyramine, 80 bacterial strains were isolated. All the strains were identified to be Bacillus subtilis and highly capable of producing β-phenylethylamine and tyramine. Therefore, it seems likely that the remarkable contents of β-phenylethylamine and tyramine in Natto predominantly resulted from the strains highly capable of producing those amines present in the food. Copyright © 2012 Elsevier Ltd. All rights reserved.

Dietary soy and natto intake and cardiovascular disease mortality in Japanese adults: the Takayama study.

PubMed

Nagata, Chisato; Wada, Keiko; Tamura, Takashi; Konishi, Kie; Goto, Yuko; Koda, Sachi; Kawachi, Toshiyuki; Tsuji, Michiko; Nakamura, Kozue

2017-02-01

Whether soy intake is associated with a decreased risk of cardiovascular disease (CVD) remains unclear. A traditional Japanese soy food, natto, contains a potent fibrinolytic enzyme. However, its relation to CVD has not been studied. We aimed to examine the association of CVD mortality with the intake of natto, soy protein, and soy isoflavones in a population-based cohort study in Japan. The study included 13,355 male and 15,724 female Takayama Study participants aged ≥35 y. At recruitment in 1992, each subject was administered a validated semiquantitative food-frequency questionnaire. Deaths from CVD were ascertained over 16 y. A total of 1678 deaths from CVD including 677 stroke and 308 ischemic heart disease occurred during follow-up. The highest quartile of natto intake compared with the lowest intake was significantly associated with a decreased risk of mortality from total CVD after control for covariates: the HR was 0.75 (95% CI: 0.64, 0.88, P-trend = 0.0004). There were no significant associations between the risk of mortality from total CVD and intakes of total soy protein, total soy isoflavone, and soy protein or soy isoflavone from soy foods other than natto. The highest quartiles of total soy protein and natto intakes were significantly associated with a decreased risk of mortality from total stroke (HR = 0.75, 95% CI: 0.57, 0.99, P-trend = 0.03 and HR = 0.68, 95% CI: 0.52, 0.88, P-trend = 0.0004, respectively). The highest quartile of natto intake was also significantly associated with a decreased risk of mortality from ischemic stroke (HR = 0.67, 95% CI:0.47, 0.95, P-trend = 0.03). Data suggest that natto intake may contribute to the reduction of CVD mortality. © 2017 American Society for Nutrition.

Identification of a Key Gene Involved in Branched-Chain Short Fatty Acids Formation in Natto by Transcriptional Analysis and Enzymatic Characterization in Bacillus subtilis.

PubMed

Hong, Chenlu; Chen, Yangyang; Li, Lu; Chen, Shouwen; Wei, Xuetuan

2017-03-01

Natto as a fermented soybean product has many health benefits for human due to its rich nutritional and functional components. However, the unpleasant odor of natto, caused by the formation of branched-chain short fatty acids (BCFAs), prohibits the wide acceptance of natto products. This work is to identify the key gene of BCFAs formation and develop the guidance to reduce natto odor. Transcriptional analysis of BCFAs synthesis pathway genes was conducted in two Bacillus subtilis strains with obvious different BCFAs synthesis abilities. The transcriptional levels of bcd, bkdAA, and ptb in B. subtilis H-9 were 2.7-fold, 0.7-fold, and 8.9-fold higher than that of B. subtilis H-4, respectively. Therefore, the ptb gene with the highest transcriptional change was considered as the key gene in BCFAs synthesis. The ptb encoded enzyme Ptb was further characterized by inducible expression in Escherichia coli. The recombinant Ptb protein (about 32 kDa) was verified by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis analysis. The catalysis functions of Ptb were confirmed on substrates of isovaleryl-CoA and isobutyryl-CoA, and the higher catalysis efficiency of Ptb on isovaleryl-CoA explained the higher level of isovaleric acid in natto. The optimal activities of Ptb were observed at 50 °C and pH 8.0, and the enzymatic activity was inhibited by Ca 2+ , Zn 2+ , Ba 2+ , Mn 2+ , Cu 2+ , SDS, and EDTA. Collectively, this study reports a key gene responsible for BCFAs formation in natto fermentation and provides potential strategies to solve the odor problem.

Genetic and Physiological Studies of Bacillus anthracis Related to Development of an Improved Vaccine

DTIC Science & Technology

1987-07-01

nontransformable Bacillus species such as B. anthracis. Our results suggest that plasmid pLS20 of Bacillus subtilis ( natto ), which promotes transfer of the...mobilizing pBC16, pLS20 mediates transfer of the B. subtills ( natto ) plasmid pLS19 and the Staphylococcus aureus plasmid pUB110. To facilitate direct...and (v) transformation of B. cereus and B. anthracis with plasmid DNA. The 55-kb plasmid, pLS20, of Bacillus subtilis ( natto ) 3335 promotes tr msfer

Genetic and Physiological Studies of Bacillus Anthracis Related to Development of An Improved Vaccine

DTIC Science & Technology

1988-07-01

to replicate at all in B. anthracis. Such mutants al currently being tested as vehicles for transposon mutagenesis in B. anthracis. B. subtilis ( natto ...subtilis ( natto ) 3335 harbors a plasmid, pLS20, which encodes functions responsible for conjugal transfer of plasmids among genetically re- lated and...Cultures were grown for 16 hours in BHI broth supplemented with 0.1% glycerol, o- in LG broth for B. subtilis natto . After cell pellets were suspended in

Surfactins in natto: the surfactin production capacity of the starter strains and the actual surfactin contents in the products.

PubMed

Juola, Mari; Kinnunen, Kristiina; Nielsen, Kristian Fog; von Wright, Atte

2014-12-01

Surfactin-type lipopeptides are suspected of being implicated in the rare food poisonings caused by Bacillus species outside the Bacillus cereus cluster. In order to get information on surfactin levels in actual human foods, bacilli from three commercial samples of a Japanese traditional bean product, natto, were isolated in order to clarify their potential to produce the suspect lipopeptides. The isolated bacilli were characterized as Bacillus subtilis. They were β-hemolytic and gave a positive signal in the PCR screen for genes associated with surfactin production, and their culture extracts were cytotoxic to boar sperm cells. Organic extracts of both Bacillus cultures and the natto samples were analyzed for their surfactin content using ultrahigh-performance liquid chromatography with high-resolution mass spectrometry. All the strains proved to be surfactin producers (15 to 39 μg/ml culture medium); the natto samples contained as much as 2.2 mg g(-1) of surfactins. This means that consumers can ingest at least approximately 80 to 100 mg of surfactins per single 50-g natto serving apparently without suffering any ill effects, indicating a very low human toxicity.

Purification and characterization of nattokinase from Bacillus subtilis natto B-12.

PubMed

Wang, Cong; Du, Ming; Zheng, Dongmei; Kong, Fandong; Zu, Guoren; Feng, Yibing

2009-10-28

Bacillus subtilis natto B-12 was isolated from natto, a traditional fermented soybean food in Japan. A fibrinolytic enzyme (B-12 nattokinase) was purified from the supernatant of B. subtilis natto B-12 culture broth and showed strong fibrinolytic activity. The enzyme was homogenously purified to 56.1-fold, with a recovery of 43.2% of the initial activity. B-12 nattokinase was demonstrated to be homogeneous by SDS-PAGE and was identified as a monomer of 29000 +/- 300 Da in its native state by SDS-PAGE and size exclusion methods. The optimal pH value and temperature were 8.0 and 40 degrees C, respectively. Purified nattokinase showed high thermostability at temperatures from 30 to 50 degrees C and alkaline stability within the range of pH 6.0-9.0. The enzyme activity was activated by Zn(2+) and obviously inhibited by Fe(3+) and Al(3+). This study provides some important information for the effect factors of fibrinolytic activity, the purification methods, and characterization of nattokinase from B. subtilis natto B-12, which enriches the theoretical information of nattokinase for the research and development of nattokinase as a functional additive of food.

Genetic and Physiological Studies of Bacillus anthracis Related to Development of an Improved Vaccine

DTIC Science & Technology

1986-07-01

demonstrated that plasmid pLS20 of B. subtilis ( natto ) is capa le of promoting the transfer of pBC16 from B. subtilis to a variety of Bacillus s ecies...anthracis. Hofetver, results of recent experiments demonstrate that pL320, a 34-megadalton plasmid of B. subtilis ( natto ), is capable of promoting the...plasmid in Bacillus subtilis ( natto ).20 IV. Determination of the size of pX02 by restriction analysis-....... 24 V. Transfer of pXO1 by the B

Genetic and Physiological Studies of Bacillus Anthracis Related to Development of an Improved Vaccine

DTIC Science & Technology

1990-07-01

charartorization of the conjugative pl&a.dd, PLS2O, of Baci~llu sbtllis ( natto ) amd its ability to tranisfer pla&iuids v" iitrains of B. inthraci~s, B...2-r-ich atms;hwre. A smll amount of effort wa spent on further’ characterization of the Bacillus subtilis ( natto ) fertility plasmid. pLS2I, which...previously cured of pX02. All transcipients that inherited the pXO2::Tn9l7 derivatives exhibited the donor phenotype. The 64.2-kb Bacillus subtilis ( natto

Whole Genome Complete Resequencing of Bacillus subtilis Natto by Combining Long Reads with High-Quality Short Reads

PubMed Central

Kamada, Mayumi; Hase, Sumitaka; Sato, Kengo; Toyoda, Atsushi; Fujiyama, Asao; Sakakibara, Yasubumi

2014-01-01

De novo microbial genome sequencing reached a turning point with third-generation sequencing (TGS) platforms, and several microbial genomes have been improved by TGS long reads. Bacillus subtilis natto is closely related to the laboratory standard strain B. subtilis Marburg 168, and it has a function in the production of the traditional Japanese fermented food “natto.” The B. subtilis natto BEST195 genome was previously sequenced with short reads, but it included some incomplete regions. We resequenced the BEST195 genome using a PacBio RS sequencer, and we successfully obtained a complete genome sequence from one scaffold without any gaps, and we also applied Illumina MiSeq short reads to enhance quality. Compared with the previous BEST195 draft genome and Marburg 168 genome, we found that incomplete regions in the previous genome sequence were attributed to GC-bias and repetitive sequences, and we also identified some novel genes that are found only in the new genome. PMID:25329997

Phylogenetic analysis of Bacillus subtilis strains applicable to natto (fermented soybean) production

USDA-ARS?s Scientific Manuscript database

Spore-forming Bacillus strains that produce extracellular poly-'-glutamic acid were screened for their application to natto (fermented soybean food) fermentation. Among the 365 strains, including B. subtilis and B. amyloliquefaciens, which we isolated from rice straw, 59 were capable of fermenting n...

Inhibition of angiotensin I converting enzyme by subtilisin NAT (nattokinase) in natto, a Japanese traditional fermented food.

PubMed

Murakami, Keiko; Yamanaka, Naoki; Ohnishi, Katsunori; Fukayama, Minoru; Yoshino, Masataka

2012-06-01

Angiotensin I converting enzyme (ACE) was inhibited by the culture medium of Bacillus subtilis subsp. natto, which ferments boiled soy beans to natto, a Japanese traditional food. Subtilisin NAT (nattokinase) produced by B. subtilis also inhibited ACE, and the inhibition was markedly stimulated by heat treatment of subtilisin at 120 °C for 15 min. Inhibition of ACE by subtilisin was of a mixed type: the decrease in V(max) and the increase in K(m) value. SDS-polyacrylamide gel electrophoresis showed that heat treatment of subtilisin caused inactivation with fragmentation of the enzyme protein into small peptides. The inhibitory action of subtilisin was not due to an enzymatic action of protease, but may be ascribed to the potent ACE-inhibitory peptides such as LY and FY, amino acid sequences in subtilisin. HPLC-MS analysis of heat-inactivated subtilisin confirmed that LY and FY were liberated by fragmentation of the enzyme. Inhibition of ACE by subtilisin and its degradation peptides such as LY and FY may participate in the suppression of blood pressure by ingestion of natto.

Chemical changes associated with lotus and water lily natto production

NASA Astrophysics Data System (ADS)

Lestari, S. D.; Fatimah, N.; Nopianti, R.

2017-04-01

Natto is a traditional Japanese food made by fermenting whole soybean seeds with pure culture of Bacillus subtilis subsp. natto. The purpose of this study was to investigate the suitability of lotus (Nelumbo nucifera) and water lily (Nymphaea stellata) seeds as the raw materials for natto production. Chemical (proximate, amino acids and minerals) changes were observed on raw, steamed and fermented seeds. Proximate compositions of all samples were calculated in both wet basis and dry basis. In wet basis calculation, steaming and fermentation tended to lower the carbohydrates, ashes, fats and protein content which were attributed to the increase of moisture. The total amino acid, iron and magnesium contents of raw lotus seeds were 24.29%, 5.08 mg 100g-1 and 174.23 mg 100g-1 dry matter, respectively. After a 24h-fermentation at 40°C, the total amino acids decreased while iron and magnesium contents increased significantly reaching, in respective order, 9.9 mg 100g-1 and 411.36 mg 100g-1 dry matter. Changes in chemical composition after fermentation were more pronounced in lotus seeds than water lily seeds indicating that their nutrient composition were more suitable to support Bacillus subtilis growth.

Distribution of radioactive cesium ((134)Cs Plus(137)Cs) in a contaminated Japanese soybean cultivar during the preparation of tofu, natto, and nimame (Boiled Soybean).

PubMed

Hachinohe, Mayumi; Kimura, Keitarou; Kubo, Yuji; Tanji, Katsuo; Hamamatsu, Shioka; Hagiwara, Shoji; Nei, Daisuke; Kameya, Hiromi; Nakagawa, Rikio; Matsukura, Ushio; Todoriki, Setsuko; Kawamoto, Shinichi

2013-06-01

We investigated the fate of radioactive cesium ((134)Cs plus (137)Cs) during the production of tofu, natto, and nimame (boiled soybean) from a contaminated Japanese soybean cultivar harvested in FY2011. Tofu, natto, and nimame were made from soybean grains containing radioactive cesium (240 to 340 Bq/kg [dry weight]), and the radioactive cesium in the processed soybean foods and in by-product fractions such as okara, broth, and waste water was measured with a germanium semiconductor detector. The processing factor is the ratio of radioactive cesium concentration of a product before and after processing. For tofu, natto, nimame, and for the by-product okara, processing factors were 0.12, 0.40, 0.20, and 0.18, respectively; this suggested that these three soybean foods and okara, used mainly as an animal feed, can be considered safe for human and animal consumption according to the standard limit for radioactive cesium of soybean grains. Furthermore, the ratio of radioactive cesium concentrations in the cotyledon, hypocotyl, and seed coat portions of the soybean grain was found to be approximately 1:1:0.4.

Natto and viscous vegetables in a Japanese-style breakfast improved insulin sensitivity, lipid metabolism and oxidative stress in overweight subjects with impaired glucose tolerance.

PubMed

Taniguchi-Fukatsu, Akiko; Yamanaka-Okumura, Hisami; Naniwa-Kuroki, Yuko; Nishida, Yuka; Yamamoto, Hironori; Taketani, Yutaka; Takeda, Eiji

2012-04-01

We previously suggested that the consumption of natto and viscous vegetables as part of a Japanese-style meal based on white rice (WR) reduced postprandial glucose and insulin levels in healthy subjects. The aim of the present study was to assess whether a single breakfast of natto and viscous vegetables or the same breakfast consumed for 2 weeks could improve glucose control, insulin sensitivity, lipid metabolism and oxidative stress in overweight subjects with impaired glucose tolerance (IGT). A total of eleven free-living subjects with IGT followed a randomised, crossover breakfast intervention for 2 weeks. The test meal included boiled WR with natto (viscous fermented soyabeans), Japanese yam and okra. The control meal included WR with non-viscous boiled soyabeans, potatoes and broccoli. Both meals contained comparable amounts of carbohydrate, fat, protein and fibre. The test meal reduced acute glucose and insulin responses compared to the control meal in the study participants. Insulin sensitivity was assessed using the composite insulin sensitivity index (CISI) after both the test and control meal periods. The test meal resulted in improvements in CISI compared to the baseline, whereas no significant changes were observed after the control meal period. Serum levels of both total and LDL-cholesterol were assessed before and after the test meal period and found to decrease significantly. There was also a tendency towards reduced serum malondialdehyde-modified LDL and N(ɛ)-carboxymethyllysine. No differences were observed in the measures of chronic glycaemic control. Thus, we conclude that a breakfast of natto and viscous vegetables consumed for 2 weeks improves insulin sensitivity, serum lipid and oxidative stress.

Isolation of antifungal bacteria from Japanese fermented soybeans, natto.

PubMed

Murata, Daichi; Sawano, Sayaka; Ohike, Tatsuya; Okanami, Masahiro; Ano, Takashi

2013-12-01

An inhibitory effect of a traditional Japanese fermented food, natto, was found against plant pathogens such as Rhizoctonia solani and Fusarium oxysporum, and the bacteria which showed inhibition were isolated from the natto. Among isolated bacteria, BC-1 and GAc exhibited a strong antagonistic effect in vitro against plant pathogens on an agar medium. The supernatant of bacterial culture also showed strong activity against R. solani, which meant the antimicrobial substances were produced and secreted into the medium. Both of the bacteria were estimated as Bacillus amyloliquefaciens from a partial sequence of the 16s rRNA gene. High performance liquid chromatography analysis clearly showed the production of the lipopeptide antibiotic iturin A by BC-1 and GAc. Copyright © 2013 The Research Centre for Eco-Environmental Sciences, Chinese Academy of Sciences. Published by Elsevier B.V. All rights reserved.

Spores of two probiotic Bacillus species enhance cellular immunity in BALB/C mice.

PubMed

Gong, Li; Huang, Qin; Fu, Aikun; Wu, YanPing; Li, Yali; Xu, Xiaogang; Huang, Yi; Yu, Dongyou; Li, Weifen

2018-01-01

Previous studies found that Bacillus subtilis BS02 and B. subtilis subsp. natto BS04 isolated in our laboratory could activate the immune response of murine macrophages in vitro. This study aims to investigate the effects of dietary supplementation with Bacillus species spores on the systemic cellular immune response in BALB/C mice. Results showed that both B. subtilis BS02 and B. subtilis natto BS04 enhanced the phagocytic function of the mononuclear phagocyte system (MPS) and the cytotoxicity of natural killer (NK) cells. In addition, B. subtilis BS02 could increase the respiratory burst activity of blood phagocytes. Furthermore, B. subtilis BS02 and B. subtilis natto BS04 increased the percentage of gamma-interferon-producing CD4 + cells and CD8 + T-cells, but only BS04 increased the percentage of CD3 + cells and CD3 +  CD4 + cells in splenocytes. However, there were no effects on other subsets of splenic lymphocytes and mitogen-induced splenic lymphocyte proliferation. All data suggested that oral administration of B. subtilis BS02 or B. subtilis natto BS04 could significantly enhance cellular immunity in BALB/C mice by increasing phagocytic activity of MPS and cytotoxic activity of NK cells in a strain-specific manner.

Inhibition of P-glycoprotein in Caco-2 cells: effects of herbal remedies frequently used by cancer patients.

PubMed

Engdal, S; Nilsen, O G

2008-06-01

1. The herbal products Natto K2, Agaricus, mistletoe, noni juice, green tea and garlic were investigated for in vitro inhibitory potential on P-glycoprotein (P-gp)-mediated transport of digoxin (30 nM) in differentiated and polarized Caco-2 cells. 2. Satisfactory cell functionality was demonstrated through measurements of assay linearity, transepithelial electric resistance (TEER), cytotoxicity, mannitol permeability, and inclusion of the positive inhibition control verapamil. 3. The most potent inhibitors of the net digoxin flux (IC(50)) were mistletoe > Natto K2 > Agaricus > green tea (0.022, 0.62, 3.81, >4.5 mg ml(-1), respectively). Mistletoe also showed the lowest IC(25) value, close to that obtained by verapamil (1.0 and 0.5 microg ml(-1), respectively). The IC(50)/IC(25) ratio was found to be a good parameter for the determination of inhibition profiles. Garlic and noni juice were classified as non-inhibitors. 4. This study shows that mistletoe, Natto K2, Agaricus and green tea inhibit P-gp in vitro. Special attention should be paid to mistletoe due to very low IC(50) and IC(25) values and to Natto K2 due to a low IC(50) value and a low IC(50)/IC(25) ratio.

Effects of Bacillus subtilis var. natto products on symptoms caused by blood flow disturbance in female patients with lifestyle diseases

PubMed Central

Hitosugi, Masahito; Hamada, Katsuo; Misaka, Kazutaka

2015-01-01

The fermented soybean product natto is a popular traditional food in Japan and is considered a health supplement. NKCP®, a natto-derived dietary food supplement whose main component is bacillopeptidase F, has antithrombotic, fibrinolytic, and blood pressure-lowering effects. We examined whether daily intake of NKCP® effectively improves subjective symptoms in patients with lifestyle diseases in this cross-over, double-blind study. Fermented soya extract with subtilisin NAT (nattokinase) as the main component was used as an active placebo. A 4-week course of NKCP® significantly decreased the visual analog scale (VAS) score for shoulder stiffness from 42.3 to 32.4 (P=0.009), the VAS score for low back pain from 25.5 to 18.8 (P=0.02), and the VAS score for coldness of the extremities from 33.1 to 25.7 (P=0.002). However, no significant difference was found in the VAS score for headache. After a 4-week course of active placebo, no significant changes in the VAS score were found for any symptoms. The significant improvement in the symptoms secondary to blood flow disturbance was caused by the improvement in blood flow by NKCP®. The use of dietary supplements based on the Japanese traditional food natto helps to relieve subjective symptoms for patients with lifestyle diseases receiving medical care. PMID:25653551

Purification and characterization of a strong fibrinolytic enzyme (nattokinase) in the vegetable cheese natto, a popular soybean fermented food in Japan.

PubMed

Fujita, M; Nomura, K; Hong, K; Ito, Y; Asada, A; Nishimuro, S

1993-12-30

A strong fibrinolytic enzyme (nattokinase) was purified from the vegetable cheese natto. Nattokinase was extracted from natto with saline and isolated by sequential use of hydrophobic chromatography on Butyl-Toyopearl, ion-exchange chromatography on CM-Toyopearl, and gel-filtration on Sephadex G-50. The isolated protein gave a single sharp band on SDS-PAGE either before or after reduction. The sequence, as determined by automated Edman degradation of the uncleaved molecule and its enzymatically derived peptide, consisted of a total 275 amino acid residues (M.W = 27,728) and exhibited a high homology with the subtilisins. The purified nattokinase digested not only fibrin but also several synthetic substrates. Among the synthetic substrates, the most sensitive substrate was Suc-Ala-Ala-Pro-Phe-pNA for subtilisin. PMSF inhibited both the fibrinolytic activity and the amidolytic activity. The results indicate that nattokinase is a subtilisin-like serine protease.

Maternal intake of Natto, a Japan's traditional fermented soybean food, during pregnancy and the risk of eczema in Japanese babies.

PubMed

Ozawa, Naoko; Shimojo, Naoki; Suzuki, Yoichi; Ochiai, Shingo; Nakano, Taiji; Morita, Yoshinori; Inoue, Yuzaburo; Arima, Takayasu; Suzuki, Shuichi; Kohno, Yoichi

2014-06-01

There are reports that the maternal diet during pregnancy may affect development of babies' eczema. We sought to investigate the association between the maternal diet during pregnancy and the risk of eczema in infancy in Japan. A birth cohort was set up at 2 hospitals in Chiba city. Dietary habits concerning fish, butter, margarine, yogurt and natto during pregnancy was obtained from mothers just after delivery. The intake frequencies of these foods were classified into four groups: 1) daily, 2) 2-3 times a week, 3) once a week and 4) once a month or less. Diagnosis of eczema at 6 months of age was made by the presence of an itchy rash that persisted more than two months. Valid data on 650 mother-baby pairs were obtained. No relationship between frequencies of the maternal intake of fish, margarine and yogurt during pregnancy and the onset rate of the babies' eczema were observed. For butter consumption, the incidence of babies' eczema was significantly higher in the group with daily intake than in those with an intake 2-3 times a week or less (p = 0.044). For natto, incidence of babies' eczema was significantly lower in the group with everyday intake than those eating it 2-3 times a week or less (p = 0.020). High frequency intake of natto during pregnancy possibly reduces the incidence of eczema in children at 6 months of age.

Natto and viscous vegetables in a Japanese style meal suppress postprandial glucose and insulin responses.

PubMed

Taniguchi, Akiko; Yamanaka-Okumura, Hisami; Nishida, Yuka; Yamamoto, Hironori; Taketani, Yutaka; Takeda, Eiji

2008-01-01

Naturally viscous vegetables and natto, made by fermenting soybeans, are very palatable and considered to be healthy foods in Japan. The objective was to assess whether the consumption of natto and viscous vegetables as part of a traditional Japanese breakfast based on high-glycemic index white rice affects glycemic, insulinemic, lipidemic and satiety responses in healthy subjects. Eleven healthy subjects consumed the reference, control and test meals in a randomized cross-over design. The test meal, comprising 200 g of boiled white rice with viscous meal (50 g natto, 60 g Japanese yams and 40 g okras), and the control meal, comprising 200 g of white rice with non-viscous boiled soybeans, potatoes and broccoli, contained comparable amounts of carbohydrate, fat, protein and fiber. In addition, whiter rice was used as a reference meal. Blood samples over 180 min were analyzed for glucose, insulin, non-esterified free fatty acid and triacylglyceride. Peak glucose and insulin concentrations after the test meal (6.0 mmol/L and 262 pmol/L) were significantly lower than after the control meal (6.8 mmol/L and 360 pmol/L). The incremental areas under the curve for glucose and insulin over 0-120 min after the test meal were also significantly reduced as compared with the control meal (28 and 27%). The consumption of naturally viscous vegetables with white rice reduced acute glycemia and insulinemia. This practical dietary combination would ensure compliance and favorably alter the risk for diabetes and cardiovascular diseases.

Genomic analysis of Bacillus subtilis lytic bacteriophage ϕNIT1 capable of obstructing natto fermentation carrying genes for the capsule-lytic soluble enzymes poly-γ-glutamate hydrolase and levanase.

PubMed

Ozaki, Tatsuro; Abe, Naoki; Kimura, Keitarou; Suzuki, Atsuto; Kaneko, Jun

2017-01-01

Bacillus subtilis strains including the fermented soybean (natto) starter produce capsular polymers consisting of poly-γ-glutamate and levan. Capsular polymers may protect the cells from phage infection. However, bacteriophage ϕNIT1 carries a γ-PGA hydrolase gene (pghP) that help it to counteract the host cell's protection strategy. ϕNIT had a linear double stranded DNA genome of 155,631-bp with a terminal redundancy of 5,103-bp, containing a gene encoding an active levan hydrolase. These capsule-lytic enzyme genes were located in the possible foreign gene cluster regions between central core and terminal redundant regions, and were expressed at the late phase of the phage lytic cycle. All tested natto origin Spounavirinae phages carried both genes for capsule degrading enzymes similar to ϕNIT1. A comparative genomic analysis revealed the diversity among ϕNIT1 and Bacillus phages carrying pghP-like and levan-hydrolase genes, and provides novel understanding on the acquisition mechanism of these enzymatic genes.

[Nutrition and bone health. Soybean and soy foods, and bone health].

PubMed

Kubota, Megumi; Shimizu, Hirotoshi

2009-10-01

Soybean and various types of soy products, such as natto, tofu, miso, and soy sauce, have long been consumed in Japan. Soybean, a rich source of plant proteins, contains a relatively high amount of calcium as well as being an important source of isoflavones, a group of substances whose chemical structure is similar to that of estrogen. Natto, fermented soybeans, contains vitamin K, which is involved in the activation of osteocalcin. For bone health and osteoporosis prevention in Japanese, it is thus beneficial to consume adequate amounts of soybean and soy products on a daily basis.

Occurrence of culturable soil fungi in a tropical moist deciduous forest Similipal Biosphere Reserve, Odisha, India.

PubMed

Jena, Santanu K; Tayung, Kumanand; Rath, Chandi C; Parida, Debraj

2015-03-01

Similipal Biosphere Reserve (SBR) is a tropical moist deciduous forest dominated by the species Shorea robusta . To the best of our knowledge their rich biodiversity has not been explored in term of its microbial wealth. In the present investigation, soil samples were collected from ten selected sites inside SBR and studied for their physicochemical parameters and culturable soil fungal diversity. The soil samples were found to be acidic in nature with a pH ranging from of 5.1-6.0. Highest percentage of organic carbon and moisture content were observed in the samples collected from the sites, Chahala-1 and Chahala-2. The plate count revealed that fungal population ranged from 3.6 × 10 (4) -2.1 × 10 (5) and 5.1 × 10 (4) -4.7 × 10 (5) cfu/gm of soil in summer and winter seasons respectively. The soil fungus, Aspergillus niger was found to be the most dominant species and Species Important Values Index (SIVI) was 43.4 and 28.6 in summer and winter seasons respectively. Among the sites studied, highest fungal diversity indices were observed during summer in the sites, Natto-2 and Natto-1. The Shannon-Wiener and Simpson indices in these two sites were found to be 3.12 and 3.022 and 0.9425 and 0.9373 respectively. However, the highest Fisher's alpha was observed during winter in the sites Joranda, Natto-2, Chahala-1 and Natto-1 and the values were 3.780, 3.683, 3.575 and 3.418 respectively. Our investigation revealed that, fungal population was dependent on moisture and organic carbon (%) of the soil but its diversity was found to be regulated by sporulating species like Aspergillus and Penicillium.

Occurrence of culturable soil fungi in a tropical moist deciduous forest Similipal Biosphere Reserve, Odisha, India

PubMed Central

Jena, Santanu K.; Tayung, Kumanand; Rath, Chandi C.; Parida, Debraj

2015-01-01

Similipal Biosphere Reserve (SBR) is a tropical moist deciduous forest dominated by the species Shorea robusta . To the best of our knowledge their rich biodiversity has not been explored in term of its microbial wealth. In the present investigation, soil samples were collected from ten selected sites inside SBR and studied for their physicochemical parameters and culturable soil fungal diversity. The soil samples were found to be acidic in nature with a pH ranging from of 5.1–6.0. Highest percentage of organic carbon and moisture content were observed in the samples collected from the sites, Chahala-1 and Chahala-2. The plate count revealed that fungal population ranged from 3.6 × 10 4 –2.1 × 10 5 and 5.1 × 10 4 –4.7 × 10 5 cfu/gm of soil in summer and winter seasons respectively. The soil fungus, Aspergillus niger was found to be the most dominant species and Species Important Values Index (SIVI) was 43.4 and 28.6 in summer and winter seasons respectively. Among the sites studied, highest fungal diversity indices were observed during summer in the sites, Natto-2 and Natto-1. The Shannon-Wiener and Simpson indices in these two sites were found to be 3.12 and 3.022 and 0.9425 and 0.9373 respectively. However, the highest Fisher’s alpha was observed during winter in the sites Joranda, Natto-2, Chahala-1 and Natto-1 and the values were 3.780, 3.683, 3.575 and 3.418 respectively. Our investigation revealed that, fungal population was dependent on moisture and organic carbon (%) of the soil but its diversity was found to be regulated by sporulating species like Aspergillus and Penicillium . PMID:26221092

Characterization of Bacillus phage-K2 isolated from chungkookjang, a fermented soybean foodstuff.

PubMed

Kim, Eun Ju; Hong, Jeong Won; Yun, Na-Rae; Lee, Young Nam

2011-01-01

An investigation of a virulent Bacillus phage-K2 (named Bp-K2) isolated from chungkookjang (a fermented soybean foodstuff) was made. Bp-K2 differed in infectivity against a number of Bacillus subtilis strains including starter strains of chungkookjang and natto, being more infectious to Bacillus strains isolated from the chungkookjang, but much less active against a natto strain. Bp-K2 is a small DNA phage whose genome size is about 21 kb. Bp-K2 is a tailed bacteriophage with an isometric icosahedral head (50 nm long on the lateral side, 80 nm wide), a long contractile sheath (85-90 nm × 28 nm), a thin tail fiber (80-85 nm long, 10 nm wide), and a basal plate (29 nm long, 47 nm wide) with a number of spikes, but no collar. The details of the structures of Bp-K2 differ from natto phage ϕBN100 as well as other known Bacillus phages such as SPO1-like or ϕ 29-like viruses. These data suggest that Bp-K2 would be a new member of the Myoviridae family of Bacillus bacteriophages.

Calcium regulates glutamate dehydrogenase and poly-γ-glutamic acid synthesis in Bacillus natto.

PubMed

Meng, Yonghong; Dong, Guiru; Zhang, Chen; Ren, Yuanyuan; Qu, Yuling; Chen, Weifeng

2016-04-01

To study the effect of Ca(2+) on glutamate dehydrogenase (GDH) and its role in poly-γ-glutamic acid (γ-PGA) synthesis in Bacillus natto HSF 1410. When the concentration of Ca(2+) varied from 0 to 0.1 g/l in the growth medium of B. natto HSF 1410, γ-PGA production increased from 6.8 to 9.7 g/l, while GDH specific activity and NH4Cl consumption improved from 183 to 295 U/mg and from 0.65 to 0.77 g/l, respectively. GDH with α-ketoglutarate as substrate primarily used NADPH as coenzyme with a K m of 0.08 mM. GDH was responsible for the synthesis of endogenous glutamate. The specific activity of GDH remained essentially unchanged in the presence of CaCl2 (0.05-0.2 g/l) in vitro. However, the specific activity of GDH and its expression was significantly increased by CaCl2 in vivo. Therefore, the regulation of GDH and PGA synthesis by Ca(2+) is an intracellular process. Calcium regulation may be an effective approach for producing γ-PGA on an industrial scale.

[Transformation of baicalin and wogonoside through liquid fermentation with Bacillus natto].

PubMed

Long, Hou-ning; Zhang, Shuo; Yao, Lei; Zhang, Min; Wang, Peng-jiao; Meng, Xiao-xia; Gao, Xiu; Zhang, Rong-ping

2015-12-01

This experiment aimed to explore and research the process of preparing baicalein and wogonin through liquid fermentation with Bacillus natto. Active enzymes of produced by B. natto was used for the biological transformation of baclin and wogonoside, in order to increase the content of the haicalein and wogonin in the scutellaria. With the content of the baicalein and wogonin as evaluating indexes, the effects of carbon source, nitrogen source, the types and suitable concentration of inorganic salt, medium pH, granularities of medical materials, liquid volume in flask, shaking speed, liquid-to-solid ratio, fermentation time on the fermentation process were studied. The optimal process conditions for liquid fermentation of scutellaria were 1.0% of peptone, 0.05% of NaCl, pH at 6, the granularities of medical materials of the scutellaria screened through 40-mesh sifter, 33% of liquid, shaker incubator speed at 200 r x min(-1), liquid-to-solid ratio of 5:1, temperature at 37 degrees C, fermentation for 6 days, baclin's conversion rate at 97.6% and wogonoside's conversion rate at 97% in the scutellaria. According to the verification test, the process was stable and feasible, and could provide data reference for the industrial production.

X-ray structure determination and deuteration of nattokinase.

PubMed

Yanagisawa, Yasuhide; Chatake, Toshiyuki; Naito, Sawa; Ohsugi, Tadanori; Yatagai, Chieko; Sumi, Hiroyuki; Kawaguchi, Akio; Chiba-Kamosida, Kaori; Ogawa, Megumi; Adachi, Tatsumi; Morimoto, Yukio

2013-11-01

Nattokinase (NK) is a strong fibrinolytic enzyme, which is produced in abundance by Bacillus subtilis natto. Although NK is a member of the subtilisin family, it displays different substrate specificity when compared with other subtilisins. The results of molecular simulations predict that hydrogen arrangements around Ser221 at the active site probably account for the substrate specificity of NK. Therefore, neutron crystallographic analysis should provide valuable information that reveals the enzymatic mechanism of NK. In this report, the X-ray structure of the non-hydrogen form of undeuterated NK was determined, and the preparation of deuterated NK was successfully achieved. The non-hydrogen NK structure was determined at 1.74 Å resolution. The three-dimensional structures of NK and subtilisin E from Bacillus subtilis DB104 are near identical. Deuteration of NK was carried out by cultivating Bacillus subtilis natto in deuterated medium. The D2O resistant strain of Bacillus subtilis natto was obtained by successive cultivation rounds, in which the concentration of D2O in the medium was gradually increased. NK was purified from the culture medium and its activity was confirmed by the fibrin plate method. The results lay the framework for neutron protein crystallography analysis.

X-ray structure determination and deuteration of nattokinase

PubMed Central

Yanagisawa, Yasuhide; Chatake, Toshiyuki; Naito, Sawa; Ohsugi, Tadanori; Yatagai, Chieko; Sumi, Hiroyuki; Kawaguchi, Akio; Chiba-Kamosida, Kaori; Ogawa, Megumi; Adachi, Tatsumi; Morimoto, Yukio

2013-01-01

Nattokinase (NK) is a strong fibrinolytic enzyme, which is produced in abundance by Bacillus subtilis natto. Although NK is a member of the subtilisin family, it displays different substrate specificity when compared with other subtilisins. The results of molecular simulations predict that hydrogen arrangements around Ser221 at the active site probably account for the substrate specificity of NK. Therefore, neutron crystallographic analysis should provide valuable information that reveals the enzymatic mechanism of NK. In this report, the X-ray structure of the non-hydrogen form of undeuterated NK was determined, and the preparation of deuterated NK was successfully achieved. The non-hydrogen NK structure was determined at 1.74 Å resolution. The three-dimensional structures of NK and subtilisin E from Bacillus subtilis DB104 are near identical. Deuteration of NK was carried out by cultivating Bacillus subtilis natto in deuterated medium. The D2O resistant strain of Bacillus subtilis natto was obtained by successive cultivation rounds, in which the concentration of D2O in the medium was gradually increased. NK was purified from the culture medium and its activity was confirmed by the fibrin plate method. The results lay the framework for neutron protein crystallography analysis. PMID:24121331

Directed evolution improves the fibrinolytic activity of nattokinase from Bacillus natto.

PubMed

Yongjun, Cai; Wei, Bao; Shujun, Jiang; Meizhi, Weng; Yan, Jia; Yan, Yin; Zhongliang, Zheng; Goulin, Zou

2011-12-01

Nattokinase (subtilisin NAT, NK) is a relatively effective microbial fibrinolytic enzyme that has been identified and characterized from Bacillus natto. In the current report, DNA family shuffling was used to improve the fibrinolytic activity of nattokinase. Three homologous genes from B. natto AS 1.107, Bacillus amyloliquefaciens CICC 20164 and Bacillus licheniformis CICC 10092 were shuffled to generate a mutant library. A plate-based method was used to screen the mutant libraries for improved activity. After three rounds of DNA shuffling, one desirable mutant with 16 amino acid substitutions was obtained. The mutant enzyme was purified and characterized. The kinetic measurements showed that the catalytic efficiency of the mutant NK was approximately 2.3 times higher than that of the wild-type nattokinase. In addition, the molecular modeling analysis suggested that the mutations affect the enzymatic function by changing the surface conformation of the substrate-binding pocket. The current study shows that the evolution of nattokinase with improved fibrinolytic activity by DNA family shuffling is feasible and provides useful references to facilitate the application of nattokinase in thrombolytic therapy. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Co-production of Fructooligosaccharides and Levan by Levansucrase from Bacillus subtilis natto with Potential Application in the Food Industry.

PubMed

Bersaneti, Gabrielly Terassi; Pan, Nicole Caldas; Baldo, Cristiani; Celligoi, Maria Antonia Pedrine Colabone

2018-03-01

Fructooligosaccharides and levan have a wide range of applications in the food industry due to their physiological and functional properties. The enzymatic synthesis of these molecules exhibits great advantages when compared with microbial fermentation. In this study, the production of levansucrase from Bacillus subtilis natto and its utilization in fructooligosaccharides and levan syntheses using different reaction conditions were described. The best condition for levansucrase production was 420.7 g L -1 of sucrose at pH 7.0, which reached 23.9 U ml -1 of transfructosylation activity. In a bioreactor, the highest production of fructooligosaccharides was 41.3 g L -1 using a medium containing 350 g L -1 sucrose at 35 °C for 36 h. The enzymatic synthesis of levan resulted in 86.9 g L -1 when conditions similar to those used for fructooligosaccharides synthesis were applied. These results indicate that the levansucrase from B. subtilis natto could be applied for the co-production of fructooligosaccharides and levan, which are biomolecules that have health benefits and are used successfully in the food industry.

Enhancement of the fibrinolytic activity in plasma by oral administration of nattokinase.

PubMed

Sumi, H; Hamada, H; Nakanishi, K; Hiratani, H

1990-01-01

The existence of a potent fibrinolytic enzyme (nattokinase, NK) in the traditional fermented food called 'natto', was reported by us previously. It was confirmed that oral administration of NK (or natto) produced a mild and frequent enhancement of the fibrinolytic activity in the plasma, as indicated by the fibrinolytic parameters, and the production of tissue plasminogen activator. NK capsules were also administered orally to dogs with experimentally induced thrombosis, and lysis of the thrombi was observed by angiography. The results obtained suggest that NK represents a possible drug for use not only in the treatment of embolism but also in the prevention of the disease, since NK has a proven safety and can be massproduced.

A novel fibrinolytic enzyme (nattokinase) in the vegetable cheese Natto; a typical and popular soybean food in the Japanese diet.

PubMed

Sumi, H; Hamada, H; Tsushima, H; Mihara, H; Muraki, H

1987-10-15

A strong fibrinolytic activity was demonstrated in the vegetable cheese Natto, which is a typical soybean food eaten in Japan. The average activity was calculated at about 40 CU (plasmin units)/g wet weight. This novel fibrinolytic enzyme, named nattokinase, was easily extracted with saline. The mol. wt and pI were about 20,000 and 8.6, respectively. Nattokinase not only digested fibrin but also the plasmin substrate H-D-Val-Leu-Lys-pNA (S-2251), which was more sensitive to the enzyme than other substrates tried. Diisopropyl fluorophosphate and 2,2,2-trichloro-1-hydroxyethyl-o,o-dimethylphosphate strongly inhibited this fibrinolytic enzyme.

Identification of quantitative trait loci associated with boiled seed hardness in soybean

PubMed Central

Hirata, Kaori; Masuda, Ryoichi; Tsubokura, Yasutaka; Yasui, Takeshi; Yamada, Tetsuya; Takahashi, Koji; Nagaya, Taiko; Sayama, Takashi; Ishimoto, Masao; Hajika, Makita

2014-01-01

Boiled seed hardness is an important factor in the processing of soybean food products such as nimame and natto. Little information is available on the genetic basis for boiled seed hardness, despite the wide variation in this trait. DNA markers linked to the gene controlling this trait should be useful in soybean breeding programs because of the difficulty of its evaluation. In this report, quantitative trait locus (QTL) analysis was performed to reveal the genetic factors associated with boiled seed hardness using a recombinant inbred line population developed from a cross between two Japanese cultivars, ‘Natto-shoryu’ and ‘Hyoukei-kuro 3’, which differ largely in boiled seed hardness, which in ‘Natto-shoryu’ is about twice that of ‘Hyoukei-kuro 3’. Two significantly stable QTLs, qHbs3-1 and qHbs6-1, were identified on chromosomes 3 and 6, for which the ‘Hyoukei-kuro 3’ alleles contribute to decrease boiled seed hardness for both QTLs. qHbs3-1 also showed significant effects in progeny of a residual heterozygous line and in a different segregating population. Given its substantial effect on boiled seed hardness, SSR markers closely linked to qHbs3-1, such as BARCSOYSSR_03_0165 and BARCSOYSSR_03_0185, could be useful for marker-assisted selection in soybean breeding. PMID:25914591

Whole-Genome Sequencing and Comparative Genome Analysis of Bacillus subtilis Strains Isolated from Non-Salted Fermented Soybean Foods.

PubMed

Kamada, Mayumi; Hase, Sumitaka; Fujii, Kazushi; Miyake, Masato; Sato, Kengo; Kimura, Keitarou; Sakakibara, Yasubumi

2015-01-01

Bacillus subtilis is the main component in the fermentation of soybeans. To investigate the genetics of the soybean-fermenting B. subtilis strains and its relationship with the productivity of extracellular poly-γ-glutamic acid (γPGA), we sequenced the whole genome of eight B. subtilis stains isolated from non-salted fermented soybean foods in Southeast Asia. Assembled nucleotide sequences were compared with those of a natto (fermented soybean food) starter strain B. subtilis BEST195 and the laboratory standard strain B. subtilis 168 that is incapable of γPGA production. Detected variants were investigated in terms of insertion sequences, biotin synthesis, production of subtilisin NAT, and regulatory genes for γPGA synthesis, which were related to fermentation process. Comparing genome sequences, we found that the strains that produce γPGA have a deletion in a protein that constitutes the flagellar basal body, and this deletion was not found in the non-producing strains. We further identified diversity in variants of the bio operon, which is responsible for the biotin auxotrophism of the natto starter strains. Phylogenetic analysis using multilocus sequencing typing revealed that the B. subtilis strains isolated from the non-salted fermented soybeans were not clustered together, while the natto-fermenting strains were tightly clustered; this analysis also suggested that the strain isolated from "Tua Nao" of Thailand traces a different evolutionary process from other strains.

Soy Allergy

MedlinePlus

... any of these ingredients: Cold-pressed, expelled or extruded soy oil* Edamame Miso Natto Shoyu Soy (soy ... soy lecithin. But avoid cold-pressed, expelled or extruded soy oil—sometimes called gourmet oils. These ingredients ...

A simple and cost-saving approach to optimize the production of subtilisin NAT by submerged cultivation of Bacillus subtilis natto.

PubMed

Ku, Ting-Wei; Tsai, Ruei-Lan; Pan, Tzu-Ming

2009-01-14

Subtilisin NAT, formerly designated nattokinase or subtilisin BSP, is a potent cardiovascular drug because of its strong fibrinolytic activity and safety. In this study, one Bacillus subtilis natto strain with high fibrinolytic activity was isolated. We further studied the optimal conditions for subtilisin NAT production by submerged cultivation and three variables/three levels of response surface methodology (RSM) using various inoculum densities, glucose concentrations, and defatted soybean concentrations as the three variables. According to the RSM analysis, while culturing by 2.93% defatted soybean, 1.75% glucose, and 4.00% inoculum density, we obtained an activity of 13.78 SU/mL. Processing the batch fermentation with this optimal condition, the activity reached 13.69 SU/mL, which is equal to 99.3% of the predicted value.

Toxicological assessment of nattokinase derived from Bacillus subtilis var. natto.

PubMed

Lampe, Bradley J; English, J Caroline

2016-02-01

Subtilisin NAT, commonly known as "nattokinase," is a fibrinolytic enzyme produced by the bacterial strain B. subtilis var. natto, which plays a central role in the fermentation of soybeans into the popular Japanese food natto. Recent studies have reported on the potential anticoagulatory and antihypertensive effects of nattokinase administration in humans, with no indication of adverse effects. To evaluate the safety of nattokinase in a more comprehensive manner, several GLP-compliant studies in rodents and human volunteers have been conducted with the enzyme product, NSK-SD (Japan Bio Science Laboratory Co., Ltd., Japan). Nattokinase was non-mutagenic and non-clastogenic in vitro, and no adverse effects were observed in 28-day and 90-day subchronic toxicity studies conducted in Sprague-Dawley rats at doses up to 167 mg/kg-day and 1000 mg/kg-day, respectively. Mice inoculated with 7.55 × 10(8) CFU of the enzyme-producing bacterial strain showed no signs of toxicity or residual tissue concentrations of viable bacteria. Additionally consumption of 10 mg/kg-day nattokinase for 4 weeks was well tolerated in healthy human volunteers. These findings suggest that the oral consumption of nattokinase is of low toxicological concern. The 90-day oral subchronic NOAEL for nattokinase in male and female Sprague-Dawley rats is 1000 mg/kg-day, the highest dose tested. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Whole-Genome Sequencing and Comparative Genome Analysis of Bacillus subtilis Strains Isolated from Non-Salted Fermented Soybean Foods

PubMed Central

Kamada, Mayumi; Hase, Sumitaka; Fujii, Kazushi; Miyake, Masato; Sato, Kengo; Kimura, Keitarou; Sakakibara, Yasubumi

2015-01-01

Bacillus subtilis is the main component in the fermentation of soybeans. To investigate the genetics of the soybean-fermenting B. subtilis strains and its relationship with the productivity of extracellular poly-γ-glutamic acid (γPGA), we sequenced the whole genome of eight B. subtilis stains isolated from non-salted fermented soybean foods in Southeast Asia. Assembled nucleotide sequences were compared with those of a natto (fermented soybean food) starter strain B. subtilis BEST195 and the laboratory standard strain B. subtilis 168 that is incapable of γPGA production. Detected variants were investigated in terms of insertion sequences, biotin synthesis, production of subtilisin NAT, and regulatory genes for γPGA synthesis, which were related to fermentation process. Comparing genome sequences, we found that the strains that produce γPGA have a deletion in a protein that constitutes the flagellar basal body, and this deletion was not found in the non-producing strains. We further identified diversity in variants of the bio operon, which is responsible for the biotin auxotrophism of the natto starter strains. Phylogenetic analysis using multilocus sequencing typing revealed that the B. subtilis strains isolated from the non-salted fermented soybeans were not clustered together, while the natto-fermenting strains were tightly clustered; this analysis also suggested that the strain isolated from “Tua Nao” of Thailand traces a different evolutionary process from other strains. PMID:26505996

Characterization of Bacillus subtilis strains in Thua nao, a traditional fermented soybean food in northern Thailand.

PubMed

Inatsu, Y; Nakamura, N; Yuriko, Y; Fushimi, T; Watanasiritum, L; Kawamoto, S

2006-09-01

To clarify the diversity of Bacillus subtilis strains in Thua nao that produce high concentrations of products useful in food manufacturing and in health-promoting compounds. Production of amylase, protease, subtilisin NAT (nattokinase), and gamma-polyglutamic acid (PGA) by the Bacillus subtilis strains in Thua nao was measured. Productivity of protease NAT by these strains tended to be higher than by Japanese commercial natto-producing strains. Molecular diversity of isolated strains was analysed via randomly amplified polymorphic DNA-PCR fingerprinting. The strains were divided into 19 types, including a type with the same pattern as a Japanese natto-producing strain. B. subtilis strains that could be a resource for effective production of protease, amylase, subtilisin NAT, or PGA were evident in Thua nao produced in various regions in northern Thailand. This study clearly demonstrated the value of Thua nao as a potential resource of food-processing enzymes and health-promoting compounds.

Purification, crystallization and preliminary X-ray diffraction experiment of nattokinase from Bacillus subtilis natto.

PubMed

Yanagisawa, Yasuhide; Chatake, Toshiyuki; Chiba-Kamoshida, Kaori; Naito, Sawa; Ohsugi, Tadanori; Sumi, Hiroyuki; Yasuda, Ichiro; Morimoto, Yukio

2010-12-01

Nattokinase is a single polypeptide chain composed of 275 amino acids (molecular weight 27,724) which displays strong fibrinolytic activity. Moreover, it can activate other fibrinolytic enzymes such as pro-urokinase and tissue plasminogen activator. In the present study, native nattokinase from Bacillus subtilis natto was purified using gel-filtration chromatography and crystallized to give needle-like crystals which could be used for X-ray diffraction experiments. The crystals belonged to space group C2, with unit-cell parameters a=74.3, b=49.9, c=56.3 Å, β=95.2°. Diffraction images were processed to a resolution of 1.74 Å with an Rmerge of 5.2% (15.3% in the highest resolution shell) and a completeness of 69.8% (30.0% in the highest resolution shell). This study reports the first X-ray diffraction analysis of nattokinase.

Purification, crystallization and preliminary X-ray diffraction experiment of nattokinase from Bacillus subtilis natto

PubMed Central

Yanagisawa, Yasuhide; Chatake, Toshiyuki; Chiba-Kamoshida, Kaori; Naito, Sawa; Ohsugi, Tadanori; Sumi, Hiroyuki; Yasuda, Ichiro; Morimoto, Yukio

2010-01-01

Nattokinase is a single polypeptide chain composed of 275 amino acids (molecular weight 27 724) which displays strong fibrinolytic activity. Moreover, it can activate other fibrinolytic enzymes such as pro-urokinase and tissue plasminogen activator. In the present study, native nattokinase from Bacillus subtilis natto was purified using gel-filtration chromatography and crystallized to give needle-like crystals which could be used for X-ray diffraction experiments. The crystals belonged to space group C2, with unit-cell parameters a = 74.3, b = 49.9, c = 56.3 Å, β = 95.2°. Diffraction images were processed to a resolution of 1.74 Å with an R merge of 5.2% (15.3% in the highest resolution shell) and a completeness of 69.8% (30.0% in the highest resolution shell). This study reports the first X-ray diffraction analysis of nattokinase. PMID:21139221

Nattokinase: An Oral Antithrombotic Agent for the Prevention of Cardiovascular Disease.

PubMed

Weng, Yunqi; Yao, Jian; Sparks, Sawyer; Wang, Kevin Yueju

2017-02-28

Natto, a fermented soybean product, has been consumed as a traditional food in Japan for thousands of years. Nattokinase (NK), a potent blood-clot dissolving protein used for the treatment of cardiovascular diseases, is produced by the bacterium Bacillus subtilis during the fermentation of soybeans to produce Natto. NK has been extensively studied in Japan, Korea, and China. Recently, the fibrinolytic (anti-clotting) capacity of NK has been recognized by Western medicine. The National Science Foundation in the United States has investigated and evaluated the safety of NK. NK is currently undergoing a clinical trial study (Phase II) in the USA for atherothrombotic prevention. Multiple NK genes have been cloned, characterized, and produced in various expression system studies. Recombinant technology represents a promising approach for the production of NK with high purity for its use in antithrombotic applications. This review covers the history, benefit, safety, and production of NK. Opportunities for utilizing plant systems for the large-scale production of NK, or for the production of edible plants that can be used to provide oral delivery of NK without extraction and purification are also discussed.

Construction of a 3D model of nattokinase, a novel fibrinolytic enzyme from Bacillus natto. A novel nucleophilic catalytic mechanism for nattokinase.

PubMed

Zheng, Zhong-liang; Zuo, Zhen-yu; Liu, Zhi-gang; Tsai, Keng-chang; Liu, Ai-fu; Zou, Guo-lin

2005-01-01

A three-dimensional structural model of nattokinase (NK) from Bacillus natto was constructed by homology modeling. High-resolution X-ray structures of Subtilisin BPN' (SB), Subtilisin Carlsberg (SC), Subtilisin E (SE) and Subtilisin Savinase (SS), four proteins with sequential, structural and functional homology were used as templates. Initial models of NK were built by MODELLER and analyzed by the PROCHECK programs. The best quality model was chosen for further refinement by constrained molecular dynamics simulations. The overall quality of the refined model was evaluated. The refined model NKC1 was analyzed by different protein analysis programs including PROCHECK for the evaluation of Ramachandran plot quality, PROSA for testing interaction energies and WHATIF for the calculation of packing quality. This structure was found to be satisfactory and also stable at room temperature as demonstrated by a 300ps long unconstrained molecular dynamics (MD) simulation. Further docking analysis promoted the coming of a new nucleophilic catalytic mechanism for NK, which is induced by attacking of hydroxyl rich in catalytic environment and locating of S221.

Nattokinase: An Oral Antithrombotic Agent for the Prevention of Cardiovascular Disease

PubMed Central

Weng, Yunqi; Yao, Jian; Sparks, Sawyer; Wang, Kevin Yueju

2017-01-01

Natto, a fermented soybean product, has been consumed as a traditional food in Japan for thousands of years. Nattokinase (NK), a potent blood-clot dissolving protein used for the treatment of cardiovascular diseases, is produced by the bacterium Bacillus subtilis during the fermentation of soybeans to produce Natto. NK has been extensively studied in Japan, Korea, and China. Recently, the fibrinolytic (anti-clotting) capacity of NK has been recognized by Western medicine. The National Science Foundation in the United States has investigated and evaluated the safety of NK. NK is currently undergoing a clinical trial study (Phase II) in the USA for atherothrombotic prevention. Multiple NK genes have been cloned, characterized, and produced in various expression system studies. Recombinant technology represents a promising approach for the production of NK with high purity for its use in antithrombotic applications. This review covers the history, benefit, safety, and production of NK. Opportunities for utilizing plant systems for the large-scale production of NK, or for the production of edible plants that can be used to provide oral delivery of NK without extraction and purification are also discussed. PMID:28264497

Inhibition of Lipopolysaccharide-Induced Interleukin 8 in Human Adenocarcinoma Cell Line HT-29 by Spore Probiotics: B. coagulans and B. subtilis (natto).

PubMed

Azimirad, Masoumeh; Alebouyeh, Masoud; Naji, Tahereh

2017-03-01

Probiotics are used as a treatment for different intestinal disorders. They confer health benefits by different ways. This study was aimed to investigate immunomodulatory effect of Bacillus probiotic spores on the production of lipopolysaccharide (LPS)-induced interleukin 8 (IL-8) in HT-29 intestinal epithelial cells. Differentiated intestinal epithelial cell line was used as a model for the study of colonization of purified spores (Bacillus subtilis (natto) and B. coagulans) and their anti-inflammatory effects. MTT assay and trypan blue staining were used for the detection of optimal concentration of the purified spores and LPS. Pre-treatment assay was done by treatment of the cells with the purified spores for 2 h, followed by challenges with LPS for 3 and 18 h. Post-treatment assay was done by initial treatment of the cells with LPS for 18 h, followed by the spores for 3 and 6 h. Levels of IL-8 secretion and its mRNA expression were measured by ELISA and relative Q real-time PCR. Our results showed similar rates of adherence to intestinal epithelial cells by the spore probiotics, while displaying no cytotoxic effect. In the pre-treatment assay, a significant decrease in IL-8, at both protein and mRNA levels, was measured for B. coagulans spores after the addition of LPS, which was higher than those observed for Bacillus subtilis (natto) spores. In the post-treatment assay, while Bacillus subtilis (but not B. coagulans) diminished the LPS-stimulated IL-8 levels after 3 h of incubation, the inhibitory effect was not constant. In conclusion, ability of Bacillus spore probiotics for adherence to intestinal epithelial cell and their anti-inflammatory effects, through interference with LPS/IL-8 signaling, was shown in this study. Further studies are needed to characterize responsible bacterial compounds associated with these effects.

Jet-Boundary and Plan-Form Corrections for Partial-Span Models with Reflection Plane, End Plate, or No End Plate in a Closed Circular Wind Tunnel

DTIC Science & Technology

1946-06-01

complete-span models. Such models are used to--best- advantage to determine the aerodynamic characteristics of wings, flaps, lateral-control devices, and...d~mensfons w? Indes> . . ... ,. ., I . .’ . .,. . , 84295 #aOO . ~- 4 g— --% ii E~ [039 E. miiw%“” NATto )w. AWIWRY mwl—nEmkMEwrK5 . faKw+- WIW m&J. I

Japan Report - Science and Technology

DTIC Science & Technology

1986-10-17

Stock of Bacillus Subtilis Bacillus subtilis is a relative of bacillus natto that has been in use in Japan from the ancient times. It is safe and...addition to the advantages of this combination of personal styles, the two had great confidence in each other. Chairman Kaneo is the so-called "fair...sixth year of his presidency. That has been just the right age for a president, and given these advantages Sunagane has a good chance of becoming

Effects of nattokinase, a pro-fibrinolytic enzyme, on red blood cell aggregation and whole blood viscosity.

PubMed

Pais, Eszter; Alexy, Tamas; Holsworth, Ralph E; Meiselman, Herbert J

2006-01-01

The vegetable cheese-like food, natto, is extremely popular in Japan with a history extending back over 1000 years. A fibrinolytic enzyme, termed nattokinase, can be extracted from natto; the enzyme is a subtilisin-like serine protease composed of 275 amino acid residues and has a molecular weight of 27.7 kDa. In vitro and in vivo studies have consistently demonstrated the potent pro-fibrinolytic effect of the enzyme. However, no studies to date have evaluated the effects of nattokinase on various hemorheological parameters and thus we have begun to assess the effects of the enzyme on RBC aggregation and blood viscosity. Blood samples were incubated with nattokinase (final activities of 0, 15.6, 31.3, 62.5 and 125 units/ml) for 30 minutes at 37 degrees C. RBC aggregation was measured using a Myrenne MA-1 aggregometer and blood viscosity assessed over 1-1000 s(-1) with a computer controlled scanning capillary rheometer (Rheolog). Our in vitro results showed a significant, dose-dependent decrease of RBC aggregation and low-shear viscosity, with these beneficial effects evident at concentrations similar to those achieved in previous in vivo animal trials. Our preliminary data thus indicate positive in vitro hemorheological effects of nattokinase, and suggest its potential value as a therapeutic agent and the need for additional studies and clinical trials.

Type II Forward Storage Site Facilities. POMCUS System. Volume 1

DTIC Science & Technology

1980-09-01

GENERAL ILLUM! NATTO . jl 37 900 ’ M23218 -DICAL EQUIPMENT SET S,.TTALI1.; a 115 1,355 N54691 LHARGER BATTERY 2 i8 200 N82364 PERISCOPE BATTER’ COMMA!-D I 3...the Pact has improved its capability to exploit these advantages , the United States has responded by improving its reinforcement capability. In...areas, but they are also large enough and positioned in such a way as to cp-italize on their basic advantages . Hence it is possible * that the Pact

Visual Approach and Landing Aids for Aircraft. A Theoretical Analysis of Some Fundamental Aspects of the Problem by Means of Perspective Diagrams

DTIC Science & Technology

1947-01-01

mm one) THAN TNI «umom») HCU<H.»OM OWA—U rowappw or TM» oocmpff. iv ~»^^»K.iMouA»iMiMAf.TOor.TMPii»tMN»iiA«« «it> Aamt.» A natto EIAAUCTE urrm...noted that the pilot is not tied to an ideal approach path fixed in space. 6.5 An important advantage of this pattern is that it resolves the

In vitro inhibition of CYP3A4 by herbal remedies frequently used by cancer patients.

PubMed

Engdal, Silje; Nilsen, Odd Georg

2009-07-01

The herbal remedies Natto K2, Agaricus, mistletoe, noni juice, green tea and garlic, frequently used by cancer patients, were investigated for their in vitro inhibition potential of cytochrome P-450 3A4 (CYP3A4) metabolism. To our knowledge, only garlic and green tea had available data on the possible inhibition of CYP3A4 metabolism. Metabolic studies were performed with human c-DNA baculovirus expressed CYP3A4. Testosterone was used as a substrate and ketoconazole as a positive quantitative inhibition control. The formation of 6-beta-OH-testosterone was quantified by a validated HPLC methodology. Green tea was the most potent inhibitor of CYP3A4 metabolism (IC(50): 73 microg/mL), followed by Agaricus, mistletoe and noni juice (1324, 3594, >10 000 microg/mL, respectively). All IC(50) values were high compared with those determined for crude extracts of other herbal remedies. The IC(50)/IC(25) ratios for the inhibiting herbal remedies ranged from 2.15 to 2.67, indicating similar inhibition profiles of the herbal inhibitors of CYP3A4. Garlic and Natto K2 were classified as non-inhibitors. Although Agaricus, noni juice, mistletoe and green tea inhibited CYP3A4 metabolism in vitro, clinically relevant systemic or intestinal interactions with CYP3A4 were considered unlikely, except for a probable inhibition of intestinal CYP3A4 by the green tea product. Copyright 2009 John Wiley & Sons, Ltd.

[Recommended soy and soy products intake to prevent bone fracture and osteoporosis].

PubMed

Uenishi, Kazuhiro

2005-08-01

Soy contains isoflavones, which are phytoestrogens, and its intake may help to prevent some diseases including menopausal disorder, osteoporosis, and breast cancer. Natto, a fermented soy product, is rich in vitamin K, which also contributes to bone health. In this report, we overviewed peer-reviewed papers showing relationship between soy product intake and risks of bone fracture and osteoporosis. It is suggested that that intake of soy products is not strongly enough to conclude but possible to be efficient in prevention of bone fracture and osteoporosis.

Association between Dietary Intake and Bone Mineral Density in Japanese Postmenopausal Women: The Yokogoshi Cohort Study.

PubMed

Hirata, Harumi; Kitamura, Kaori; Saito, Toshiko; Kobayashi, Ryosaku; Iwasaki, Masanori; Yoshihara, Akihiro; Watanabe, Yumi; Oshiki, Rieko; Nishiwaki, Tomoko; Nakamura, Kazutoshi

2016-06-01

Diet and food intake play an important role in the development of osteoporosis. However, apart from calcium and vitamin D, how nutrients affect bone status is not fully understood. The purpose of this study was to determine cross-sectional and longitudinal associations between dietary intake and bone mineral density (BMD) in Japanese postmenopausal women. This 5-year cohort study included 600 community-dwelling women aged 55-74 years at baseline in 2005. Information on demographics, nutrition, and lifestyle was obtained through interviews, and nutritional and dietary intake was assessed using a validated food frequency questionnaire. BMD measurements were performed by dual energy X-ray absorptiometry. In 2010, 498 women underwent follow-up BMD examinations. Multiple linear regression analysis was performed to determine associations of predictor variables with BMD, adjusting for confounders. In cross-sectional analyses, coffee or black tea consumption was positively associated with lumbar spine (P = 0.004) and total hip (P = 0.003) BMD, and alcohol intake was positively associated with femoral neck (P = 0.005) and total hip (P = 0.001) BMD. In longitudinal analyses, vitamin K (P = 0.028) and natto (fermented soybeans) (P = 0.023) were positively associated with lumbar spine BMD, and meat or meat product consumption was inversely associated with total hip (P = 0.047) BMD. In conclusion, dietary factors other than calcium and vitamin D intake are predictors of bone mass and bone loss in Japanese postmenopausal women. In particular, natto intake is recommended for preventing postmenopausal bone loss on the basis of current evidence.

Expression of nattokinase in Escherichia coli and renaturation of its inclusion body.

PubMed

Ni, He; Guo, Peng-Cheng; Jiang, Wei-Ling; Fan, Xiao-Min; Luo, Xiang-Yu; Li, Hai-Hang

2016-08-10

Nattokinase is an important fibrinolytic enzyme with therapeutic applications for cardiovascular diseases. The full-length and mature nattokinase genes were cloned from Bacillus subtilis var. natto and expressed in pQE30 vector in Escherichia coli. The full-length gene expressed low nattokinase activity in the intracellular soluble and the medium fractions. The mature gene expressed low soluble nattokinase activity and large amount insoluble protein in inclusion bodies without enzyme activity. Large amount of refolding solutions (RSs) at different pH values were screening and RS-10 and RS-11 at pH 9 were selected to refold nattokinase inclusion bodies. The recombinant cells were lysed with 0.1mg/mL lysozyme and ultrasonic treatment. After centrifugation, the pellete was washed twice with 20mM Tris-HCl buffer (pH 7.5) containing 1% Triton X-100 to purify the inclusion bodies. The inclusion bodies were dissolved in water at pH 12.0 and refolded with RS-10. The refolded proteins showed 42.8IU/mg and 79.3IU/mg fibrinolytic activity by the traditional dilution method (20-fold dilution into RS-10) and the directly mixing the protein solution with equal volume RS-10, respectively, compared to the 52.0IU/mg of total water-soluble proteins from B. subtilis var. natto. This work demonstrated that the inclusion body of recombinant nattokinase expressed in E. coli could be simply refolded to the natural enzyme activity level by directly mixing the protein solution with equal volume refolding solution. Copyright © 2016 Elsevier B.V. All rights reserved.

Conjoint analysis on the purchase intent for traditional fermented soy product (natto) among Japanese housewives.

PubMed

Kimura, Atsushi; Kuwazawa, Shigetaka; Wada, Yuji; Kyutoku, Yasushi; Okamoto, Masako; Yamaguchi, Yui; Masuda, Tomohiro; Dan, Ippeita

2011-04-01

The effect of sensory and extrinsic attributes on consumer intentions to purchase the Japanese traditional fermented soybean product natto was evaluated using conjoint analysis. Six attributes with 2 levels each were chosen and manipulated: price (high compared with low), the country of origin of the soybeans (domestic compared with imported), stickiness (strong compared with moderate), smell (rich compared with moderate), attached seasonings (attached compared with no attached seasonings), and the environmental friendliness of the packaging (high compared with low). A fractional factorial design was applied and 8 hypothetical product labels were produced. A sample of 479 Japanese housewives ranked these product labels based on their purchase intentions. Overall purchase intention was affected by country of origin, attached seasonings, and price; those attributes accounted for 81.0%, while the sensory attributes of the product accounted for 19.0% of purchase intents. In order to estimate market segments for the natto products based on consumer preference, a cluster analysis was performed. It identified 4 segments of consumers: 1 oriented to attached seasonings, another conscious of the price, and the other 2 oriented to origins. The behavioral and demographic characteristics of the respondents had a limited influence on segment membership.   This research was conducted to understand how consumers valuate various sensory and nonsensory product attributes based on their assessment of the overall product in the case of Japanese fermented soy product (natto). The data of this research would be of great importance both in understanding consumer behavior and in designing strategies for product development.

Mosquito larvicidal effectiveness of EcoBio-Block S: a novel integrated water-purifying concrete block formulation containing insect growth regulator pyriproxyfen.

PubMed

Kawada, Hitoshi; Saita, Susumu; Shimabukuro, Kozue; Hirano, Masachika; Koga, Masayuki; Iwashita, Toshiaki; Takagi, Masahiro

2006-09-01

EcoBio-Block S, a novel controlled release system (CRS) for the insect growth regulator pyriproxyfen, uses a water-purifying concrete block system (EcoBio-Block) composed of a porous volcanic rock and cement, and it incorporates the aerobic bacterial groups of Bacillus subtilis natto. EcoBio-Block S showed high inhibitory activity against mosquito emergence as well as a water-purifying effect. Chemical analysis and bioassay showed that EcoBio-Block S provides a high-performance CRS that controls the release of pyriproxyfen at low levels according to "zero order kinetics".

Consequence of patient substitution of nattokinase for warfarin after aortic valve replacement with a mechanical prosthesis.

PubMed

Elahi, Maqsood M; Choi, Charles H; Konda, Subbareddy; Shake, Jay G

2015-01-01

This report describes a patient's self-substitution of nattokinase for the vitamin K antagonist warfarin after aortic valve replacement with a mechanical prosthesis. Nattokinase is an enzyme derived from a popular fermented soybean preparation in Japan (natto), which has fibrinolytic properties and is gaining popularity in nontraditional health journals and nonmedical health websites as an over-the-counter thrombolytic. After nearly a year of use of nattokinase without warfarin, the patient developed thrombus on the mechanical valve and underwent successful repeat valve replacement. We believe this is the first documented case of nattokinase being used as a substitute for warfarin after valve replacement, and we strongly discourage its use for this purpose.

[Molecular cloning and expression of Nattokinase gene in Bacillus subtilis].

PubMed

Liu, B Y; Song, H Y

2002-05-01

In order to characterize biochemically the nattokinase,the nucleotide sequence of the nattokinase gene was amplified from the chromosomal DNA of B.subtilis (natto) by PCR. The expression plasmid pBL NK was constructed and was used to transform Bacillus subtilis containing a chromosomal deletion in its subtilisin gene. The supernatant of the culture was collected after 15 h culture. The target proteins were identified by SDS-PAGE. Nattokinase was purified by a method including ultrafiltration, Sephacryl S-100 gel filtration and S-Sepharose ion-exchange chromatography, and 100 mg of purified nattokinase was obtained from one liter of culture. The purity of the protein and the specific activity were 95% and 12 000 u/mg (compared to tPA), respectively.

Amyloid-degrading ability of nattokinase from Bacillus subtilis natto.

PubMed

Hsu, Ruei-Lin; Lee, Kung-Ta; Wang, Jung-Hao; Lee, Lily Y-L; Chen, Rita P-Y

2009-01-28

More than 20 unrelated proteins can form amyloid fibrils in vivo which are related to various diseases, such as Alzheimer's disease, prion disease, and systematic amyloidosis. Amyloid fibrils are an ordered protein aggregate with a lamellar cross-beta structure. Enhancing amyloid clearance is one of the targets of the therapy of these amyloid-related diseases. Although there is debate on whether the toxicity is due to amyloids or their precursors, research on the degradation of amyloids may help prevent or alleviate these diseases. In this study, we explored the amyloid-degrading ability of nattokinase, a fibrinolytic subtilisin-like serine protease, and determined the optimal conditions for amyloid hydrolysis. This ability is shared by proteinase K and subtilisin Carlsberg, but not by trypsin or plasmin.

Consequence of patient substitution of nattokinase for warfarin after aortic valve replacement with a mechanical prosthesis

PubMed Central

Elahi, Maqsood M.; Choi, Charles H.; Konda, Subbareddy

2015-01-01

This report describes a patient's self-substitution of nattokinase for the vitamin K antagonist warfarin after aortic valve replacement with a mechanical prosthesis. Nattokinase is an enzyme derived from a popular fermented soybean preparation in Japan (natto), which has fibrinolytic properties and is gaining popularity in nontraditional health journals and nonmedical health websites as an over-the-counter thrombolytic. After nearly a year of use of nattokinase without warfarin, the patient developed thrombus on the mechanical valve and underwent successful repeat valve replacement. We believe this is the first documented case of nattokinase being used as a substitute for warfarin after valve replacement, and we strongly discourage its use for this purpose. PMID:25552810

Nattokinase: production and application.

PubMed

Dabbagh, Fatemeh; Negahdaripour, Manica; Berenjian, Aydin; Behfar, Abdolazim; Mohammadi, Fatemeh; Zamani, Mozhdeh; Irajie, Cambyz; Ghasemi, Younes

2014-11-01

Nattokinase (NK, also known as subtilisin NAT) (EC 3.4.21.62) is one of the most considerable extracellular enzymes produced by Bacillus subtilis natto. The main interest about this enzyme is due to its direct fibrinolytic activity. Being stable enough in the gastrointestinal tract makes this enzyme a useful agent for the oral thrombolytic therapy. Thus, NK is regarded as a valuable dietary supplement or nutraceutical. Proven safety and ease of mass production are other advantages of this enzyme. In addition to these valuable advantages, there are other applications attributed to NK including treatment of hypertension, Alzheimer's disease, and vitreoretinal disorders. This review tends to bring a brief description about this valuable enzyme and summarizes the various biotechnological approaches used in its production, recovery, and purification. Some of the most important applications of NK, as well as its future prospects, are also discussed.

Production, optimization and characterization of fibrinolytic enzyme by Bacillus subtilis RJAS19.

PubMed

Kumar, D J Mukesh; Rakshitha, R; Vidhya, M Annu; Jennifer, P Sharon; Prasad, Sandip; Kumar, M Ravi; Kalaichelvan, P T

2014-04-01

The present study aimed at the production, purification and characterization of fibrinolytic nattokinase enzyme from the bacteria isolated from natto food. For the purpose, a fibrinolytic bacterium was isolated and identified as Bacillus subtilis based on 16S rDNA sequence analysis. The strain was employed for the production and optimization of fibrinolytic enzyme. The strain showed better enzyme production during 72nd h of incubation time with 50 degrees C at the pH 9. The lactose and peptone were found to be increasing the enzyme production rate. The enzyme produced was purified and also characterized with the help of SDS-PAGE analysis. The activity and stability profile of the purified enzyme was tested against different temperature and pH. The observations suggesting that the potential of fibrinolytic enzyme produced by Bacillus subtilis RJAS 19 for its applications in preventive medicines.

Nattokinase: an updated critical review on challenges and perspectives.

PubMed

Selvarajan, Ethiraj; Bhatnagar, Niharika

2017-12-07

Natto, a fermented soybean food, has been consumed by oriental people for more than 1000 years. Nattokinase, formerly called subtilisin NAT is a well studied protease of microbial origin that possesses fibrinolytic (anti-clotting) activities. Due to its strong fibrinolytic and thrombolytic activity, Nattokinase is regarded as a precious dietary supplement or nutraceutical for the oral thrombolytic therapy. Nattokinase is witnessed to be a useful enzyme for the com-plete removal of the vitreous and associated proliferative tissues in proliferative vitreo retinal disorders. This review focuses on the native and recombinant Nattokinase from bacteria and other sources, their production, purification, immobilization and nano-immobilization studies, which aid in ameliorating their properties to suit the targeted industrial applications. Recent development in these fields are presented and discussed, and prospective developments are suggested. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Determination of 15 isoflavone isomers in soy foods and supplements by high-performance liquid chromatography.

PubMed

Yanaka, Kaoru; Takebayashi, Jun; Matsumoto, Teruki; Ishimi, Yoshiko

2012-04-25

Soy isoflavone is the generic name for the isoflavones found in soy. We determined the concentrations of 15 soy isoflavone species, including 3 succinyl glucosides, in 22 soy foods and isoflavone supplements by high-performance liquid chromatography (HPLC). The total isoflavone contents in 14 soy foods and 8 supplements ranged from 45 to 735 μg/g and from 1,304 to 90,224 μg/g, respectively. Higher amounts of succinyl glucosides were detected in natto, a typical fermented soy product in Japan; these ranged from 30 to 80 μg/g and comprised 4.1-10.9% of the total isoflavone content. In soy powder, 59 μg/g of succinyl glucosides were detected, equivalent to 4.6% of the total isoflavone content. These data suggest that the total isoflavone contents may be underestimated in the previous studies that have not included succinyl glucosides, especially for Bacillus subtilis -fermented soy food products.

A pilot study on the serum pharmacokinetics of nattokinase in humans following a single, oral, daily dose.

PubMed

Ero, Michael Penfield; Ng, Connie M; Mihailovski, Tamara; Harvey, Nathaniel R; Lewis, Brad Howard

2013-01-01

Nattokinase is a serine protease and is derived from natto, a traditional Japanese, fermented, soybean food meal. Multiple authors have described the significant fibrinolytic, antithrombotic, and antihypertensive effects of natto. Nattokinase has been growing in popularity for use as a dietary supplement for the benefit of cardiovascular health. Little is known regarding the pharmacokinetic and pharmacodynamic properties of this enzyme, and the bioavailability of nattokinase is currently unknown. This study intended to (1) detect nattokinase directly and immunologically, (2) show that nattokinase and/or its metabolites were detectable in human blood following ingestion of a commercial preparation, and (3) chart a pharmacokinetic dosing effect for nattokinase. The research team designed the pilot study as an in vivo, human clinical trial. Healthy human subjects responded to an advertisement and were screened. Subjects who satisfied both inclusion and exclusion criteria were enrolled into the study. Subjects were then instructed to orally ingest a single capsule containing a known concentration of nattokinase immediately following a baseline blood draw. Subsequent blood draws occurred over a 24-h period. This study was conducted in Oakland, California, at a clinical reference laboratory and was performed with the approval of an institutional review board (IRB) to ensure that appropriate ethical standards were met. Eleven healthy participants (five male, six female, ages 21-65), who met eligibility criteria, were enrolled. Administration of nattokinase occurred orally with the ingestion of a single daily dose (2000 FU) of nattokinase. Capsules, each containing approximately 100 mg of nattokinase, in softgel form (NSK-SD, Japan Bio Science Laboratory, Osaka, Japan), were used in the study. Baseline blood samples were collected, and participants were observed swallowing a single capsule of the nattokinase supplement before returning at 2, 4, 8, 12, 24, and 48 h post ingestion for subsequent blood draws. The presence of nattokinase in serum was measured by an enzyme-linked immunosorbent assay (ELISA), using a rabbit, polyclonal, antinattokinase-capture antibody. A pharmacokinetic pattern was observed for nattokinase between baseline and 48 h postdose. Peak serum levels of nattokinase were observed at approximately 13.3 h ± 2.5 h (mean ± standard error) postdose. Statistically significant increases in binding were detectable from baseline when comparing averaged data at time points t = 2 h-t = 24 h. These results provided the first evidence that nattokinase can be measured directly in the blood of humans. The results further suggest that a larger, more extensive, pharmacokinetic study of nattokinase is warranted. Additionally, looking for intact enzyme and bioactive nattokinase peptides should be a consideration for future studies investigating the pharmacokinetic profile of nattokinase.

Antioxidation, angiotensin converting enzyme inhibition activity, nattokinase, and antihypertension of Bacillus subtilis (natto)-fermented pigeon pea.

PubMed

Lee, Bao-Hong; Lai, Yi-Syuan; Wu, She-Ching

2015-12-01

Because of the high incidence of cardiovascular diseases in Asian countries, traditional fermented foods from Asia have been increasingly investigated for antiatherosclerotic effects. This study investigated the production of nattokinase, a serine fibrinolytic enzyme, in pigeon pea by Bacillus subtilis fermentation. B. subtilis 14714, B. subtilis 14715, B. subtilis 14716, and B. subtilis 14718 were employed to produce nattokinase. The highest nattokinase activity in pigeon pea was obtained using B. subtilis 14715 fermentation for 32 hours. In addition, the levels of antioxidants (phenolics and flavonoids) and angiotensin converting enzyme inhibitory activity were increased in B. subtilis 14715-fermented pigeon pea, compared with those in nonfermented pigeon pea. In an animal model, we found that both water extracts of pigeon pea (100 mg/kg body weight) and water extracts of B. subtilis-fermented pigeon pea (100 mg/kg body weight) significantly improved systolic blood pressure (21 mmHg) and diastolic blood pressure (30 mmHg) in spontaneously hypertensive rats. These results suggest that Bacillus-fermented pigeon pea has benefits for cardiovascular health and can be developed as a new dietary supplement or functional food that prevents hypertension. Copyright © 2015. Published by Elsevier B.V.

Microbial production and chemical transformation of poly-γ-glutamate.

PubMed

Ashiuchi, Makoto

2013-11-01

Poly-γ-glutamate (PGA), a novel polyamide material with industrial applications, possesses a nylon-like backbone, is structurally similar to polyacrylic acid, is biodegradable and is safe for human consumption. PGA is frequently found in the mucilage of natto, a Japanese traditional fermented food. To date, three different types of PGA, namely a homo polymer of D-glutamate (D-PGA), a homo polymer of L-glutamate (L-PGA), and a random copolymer consisting of D- and L-glutamate (DL-PGA), are known. This review will detail the occurrence and physiology of PGA. The proposed reaction mechanism of PGA synthesis including its localization and the structure of the involved enzyme, PGA synthetase, are described. The occurrence of multiple carboxyl residues in PGA likely plays a role in its relative unsuitability for the development of bio-nylon plastics and thus, establishment of an efficient PGA-reforming strategy is of great importance. Aside from the potential applications of PGA proposed to date, a new technique for chemical transformation of PGA is also discussed. Finally, some techniques for PGA and its derivatives in advanced material technology are presented. © 2013 The Author. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Relationship between Mutations of the Pectin Methylesterase Gene in Soybean and the Hardness of Cooked Beans.

PubMed

Toda, Kyoko; Hirata, Kaori; Masuda, Ryoichi; Yasui, Takeshi; Yamada, Tetsuya; Takahashi, Koji; Nagaya, Taiko; Hajika, Makita

2015-10-14

Hardness of cooked soybeans [Glycine max (L). Merr.] is an important attribute in food processing. We found one candidate gene, Glyma03g03360, to be associated with the hardness of cotyledons of cooked soybeans, based on a quantitative trait locus and fine-scale mapping analyses using a recombinant inbred line population developed from a cross between two Japanese cultivars, "Natto-shoryu" and "Hyoukei-kuro 3". Analysis of the DNA sequence of Glyma03g03360, a pectin methylesterase gene homologue, revealed three patterns of mutations, two of which result in truncated proteins and one of which results in an amino acid substitution. The truncated proteins are presumed to lack the enzymatic activity of Glyma03g03360. We classified 24 cultivars into four groups based on the sequence of Glyma03g03360. The texture analysis using the 22 cultivars grown in different locations indicated that protein truncation of Glyma03g03360 resulted in softer cotyledons of cooked soybeans, which was further confirmed by texture analysis performed using F2 populations of a cross between "Enrei" and "LD00-3309", and between "Satonohohoemi" and "Sakukei 98". A positive correlation between hardness and calcium content implies the possible effect of calcium binding to pectins on the hardness of cooked soybean cotyledons.

Direct high-pressure NMR observation of dipicolinic acid leaking from bacterial spore: A crucial step for thermal inactivation.

PubMed

Akasaka, Kazuyuki; Maeno, Akihiro; Yamazaki, Akira

2017-12-01

A bacterial spore protects itself with an unusually high concentration (~10% in dry weight of spore) of dipicolinic acid (DPA), the release of which is considered the crucial step for inactivating it under mild pressure and temperature conditions. However, the process of how the spore releases DPA in response to pressure remains obscure. Here we apply 1 H high-resolution high-pressure NMR spectroscopy, for the first time, to the spore suspension of Bacillus subtilis natto and monitor directly and in real-time the leaking process of DPA in response to pressure of 200MPa at 20°C. We find that about one third of the total DPA leaks immediately upon applying pressure, but that the rest leaks slowly in hrs upon decreasing the pressure. Once DPA is fully released from the spore, the proteins of the spore become easily denatured at a mild temperature, e.g., 80°C, much below the temperature commonly used to inactivate spores (121°C). The success of the present experiment opens a new avenue for studying bacterial spores and cells at the molecular level in response to pressure, temperature and other perturbations. Copyright © 2017 Elsevier B.V. All rights reserved.

[Effect of nattokinase on restenosis after percutaneous transluminal angioplasty of the abdominal artery in rabbits].

PubMed

Gong, Min; Lin, Huan-bing; Wang, Qian; Xu, Jiang-ping

2008-08-01

To investigate the effect of nattokinase on intimal hyperplasia in rabbit abdominal artery after balloon injury and explore a novel strategy for the preventing restenosis after percutaneous transluminal angioplasty. Fifty-six New Zealand rabbits were randomly divided into 7 groups, namely the solvent control group, model group, natto extract lavage group, refined nattokinse lavage group, intravenous refined nattokinse injection group, clopidogrel group and clopidogrel-aspirin group. Balloon injury was induced by inserting the catheter through the femoral artery into the thoracic aorta of the rabbits. The platelet counts were notad and platelet aggregation was observed, and the abdominal artery was taken for pathological analysis. The expressions of MMP-2 and -9 in the abdominal artery were detected immunohistochemically. There was no significant difference in the platelet counts, platelet aggregation rate or MMP-2 and -9 expression between the model group and the nattokinse-treated groups (P>0.05). The stenosis index in each nattokinse-treated group was significantly greater and the neointimal proliferation index smaller than that of the model group (P<0.01 or 0.05). Nattokinse can inhibit restenosis of rabbit abdominal artery after percutaneous transluminal angioplasty, which is independent of its actions on the platelet or MMP-2 and -9 expressions.

Improving the activity of the subtilisin nattokinase by site-directed mutagenesis and molecular dynamics simulation.

PubMed

Weng, Meizhi; Deng, Xiongwei; Bao, Wei; Zhu, Li; Wu, Jieyuan; Cai, Yongjun; Jia, Yan; Zheng, Zhongliang; Zou, Guolin

2015-09-25

Nattokinase (NK), a bacterial serine protease from Bacillus subtilis var. natto, is a potential cardiovascular drug exhibiting strong fibrinolytic activity. To broaden its commercial and medical applications, we constructed a single-mutant (I31L) and two double-mutants (M222A/I31L and T220S/I31L) by site-directed mutagenesis. Active enzymes were expressed in Escherichia coli with periplasmic secretion and were purified to homogeneity. The kinetic parameters of enzymes were examined by spectroscopy assay and isothermal titration calorimetry (ITC), and their fibrinolytic activities were determined by fibrin plate method. The substitution of Leu(31) for Ile(31) resulted in about 2-fold enhancement of catalytic efficiency (Kcat/KM) compared with wild-type NK. The specific activities of both double-mutants (M222A/I31L and T220S/I31L) were significantly increased when compared with the single-mutants (M222A and T220S) and the oxidative stability of M222A/I31L mutant was enhanced with respect to wild-type NK. This study demonstrates the feasibility of improving activity of NK by site-directed mutagenesis and shows successful protein engineering cases to improve the activity of NK as a potent therapeutic agent. Copyright © 2015 Elsevier Inc. All rights reserved.

Thrombolytic effects in vivo of nattokinase in a carrageenan-induced rat model of thrombosis.

PubMed

Xu, Jianping; Du, Ming; Yang, Xiulin; Chen, Qingquan; Chen, Hong; Lin, Dong-Hong

2014-01-01

Nattokinase is a serine protease produced by Bacillus subtilis during the fermentation of the soybean product natto. The fibrinolytic activity and thrombolytic effects of nattokinase have been observed in vitro, but the effect in vivo has still to be researched. The objective of this study was to demonstrate the activity of nattokinase in vivo. To establish a rat model of thrombosis, κ-carrageenan was injected subcutaneously into the toes of Sprague-Dawley (SD) rats. Histological examination confirmed thrombosis. The rats were then treated with varying doses of nattokinase and the resulting thrombolysis was histologically assessed. ELISA was used to determine the levels of the fibrin/fibrinogen degradation products (FDPs) and D-dimer, which are sensitive indices of fibrinolytic activity. Vermis kinase, a known thrombolytic agent, was used as a positive control. Biopsy results revealed partial thrombolysis in the tail vessels of the rats treated with nattokinase or vermis kinase. FDP and D-dimer levels were higher in rats treated with high-dose nattokinase than in those treated with saline. No difference in FDP or D-dimer levels was observed between rats treated with high-dose nattokinase and those treated with vermis kinase. Both the histological and physiological evidence from this study indicate that nattokinase exerts thrombolytic effects in vivo.

Soy food consumption and risk of prostate cancer: a meta-analysis of observational studies.

PubMed

Hwang, Ye Won; Kim, Soo Young; Jee, Sun Ha; Kim, Youn Nam; Nam, Chung Mo

2009-01-01

Soybean products have been suggested to have a chemo preventive effect against prostate cancer. The aim of this study was to provide a comprehensive meta-analysis on the extent of the possible association between soy-based food consumption and the risk of prostate cancer. Five cohort studies and 8 case-control studies were identified using MEDLINE, EMBASE, CINAHL, Korea Medical Database, KoreaMed, Korean studies Information Service System, Japana Centra Revuo Medicina, China National Knowledge Infrastructure, and a manual search. Summary odds ratios (ORs) comparing high versus low categories of soybean consumptions were calculated on the basis of the random effect model. We analyzed the associations based on the different types of soybean consumptions. The summary ORs (95% CI) for total soy foods were 0.69 (CI = 0.57-0.84) and 0.75 (CI = 0.62-0.89) for nonfermented soy foods. Among individual soy foods, only tofu yielded a significant value of 0.73 (CI = 0.57-0.92). Consumption of soybean milk, miso, or natto did not significantly reduce the risk of prostate cancer. Genistein and daidzein were associated with a lower risk of prostate cancer. This systematic review suggests that soy food consumption could lower the risk of prostate cancer. This conclusion, however, should be interpreted with caution because various biases can affect the results of a meta-analysis.

Interactions between nattokinase and heparin/GAGs

PubMed Central

Zhang, Fuming

2015-01-01

Nattokinase (NK) is a serine protease extracted from a traditional Japanese food called natto. Due to its strong fibrinolytic and thrombolytic activity, NK is regarded as a valuable dietary supplement or nutraceutical for the oral thrombolytic therapy. In addition, NK has been investigated for some other medical applications including treatment of hypertension, Alzheimer’s disease, and vitreoretinal disorders. The most widely used clinical anticoagulants are heparin and low molecular weight heparins. The interactions between heparin and proteins modulate a diverse patho-physiological processes and heparin modifies the activity of serine proteases. Indeed, heparin plays important roles in almost all of NK’s potential therapeutically applications. The current report relies on surface plasmon resonance spectroscopy to examine NK interacting with heparin as well as other glycosaminoglycans (GAGs). These studies showed that NK is a heparin binding protein with an affinity of ~250 nM. Examination with differently sized heparin oligosaccharides indicated that the interaction between NK and heparin is chain-length dependent and the minimum size for heparin binding is a hexasaccharide. Studies using chemically modified heparin showed the 6-O-sulfo as well as the N-sulfo groups but not the 2-O-sulfo groups within heparin, are essential for heparin’s interaction with NK. Other GAGs (including HS, DS, and CSE) displayed modest binding affinity to NK. NK also interfered with other heparin-protein interactions, including heparin’s interaction with antithrombin and fibroblast growth factors. PMID:26412225

Interactions between nattokinase and heparin/GAGs.

PubMed

Zhang, Fuming; Zhang, Jianhua; Linhardt, Robert J

2015-12-01

Nattokinase (NK) is a serine protease extracted from a traditional Japanese food called natto. Due to its strong fibrinolytic and thrombolytic activity, NK is regarded as a valuable dietary supplement or nutraceutical for the oral thrombolytic therapy. In addition, NK has been investigated for some other medical applications including treatment of hypertension, Alzheimer's disease, and vitreoretinal disorders. The most widely used clinical anticoagulants are heparin and low molecular weight heparins. The interactions between heparin and proteins modulate diverse patho-physiological processes and heparin modifies the activity of serine proteases. Indeed, heparin plays important roles in almost all of NK's potential therapeutically applications. The current report relies on surface plasmon resonance spectroscopy to examine NK interacting with heparin as well as other glycosaminoglycans (GAGs). These studies showed that NK is a heparin binding protein with an affinity of ~250 nM. Examination with differently sized heparin oligosaccharides indicated that the interaction between NK and heparin is chain-length dependent and the minimum size for heparin binding is a hexasaccharide. Studies using chemically modified heparin showed the 6-O-sulfo as well as the N-sulfo groups but not the 2-O-sulfo groups within heparin, are essential for heparin's interaction with NK. Other GAGs (including HS, DS, and CSE) displayed modest binding affinity to NK. NK also interfered with other heparin-protein interactions, including heparin's interaction with antithrombin and fibroblast growth factors.

Nattokinase-promoted tissue plasminogen activator release from human cells.

PubMed

Yatagai, Chieko; Maruyama, Masugi; Kawahara, Tomoko; Sumi, Hiroyuki

2008-01-01

When heated to a temperature of 70 degrees C or higher, the strong fibrinolytic activity of nattokinase in a solution was deactivated. Similar results were observed in the case of using Suc-Ala-Ala-Pro-Phe-pNA and H-D-Val-Leu-Lys-pNA, which are synthetic substrates of nattokinase. In the current study, tests were conducted on the indirect fibrinolytic effects of the substances containing nattokinase that had been deactivated through heating at 121 degrees C for 15 min. Bacillus subtilis natto culture solutions made from three types of bacteria strain were heat-treated and deactivated, and it was found that these culture solutions had the ability to generate tissue plasminogen activators (tPA) from vascular endothelial cells and HeLa cells at certain concentration levels. For example, it was found that the addition of heat-treated culture solution of the Naruse strain (undiluted solution) raises the tPA activity of HeLa cells to about 20 times that of the control. Under the same conditions, tPA activity was raised to a level about 5 times higher for human vascular endothelial cells (HUVEC), and to a level about 24 times higher for nattokinase sold on the market. No change in cell count was observed for HeLa cells and HUVEC in the culture solution at these concentrations, and the level of activity was found to vary with concentration. Copyright 2009 S. Karger AG, Basel.

Development of a soy food frequency questionnaire to estimate isoflavone consumption in US adults.

PubMed

Kirk, P; Patterson, R E; Lampe, J

1999-05-01

To develop a dietary assessment instrument to measure soy food consumption and isoflavone intake and test it for reliability. A soy food frequency questionnaire, designed for use in this study, was administered twice to participants, separated by a 2-week interval. The questionnaire was tested for reproducibility of estimates of soy food consumption and isoflavone (genistein and daidzein) intake. A convenience sample of 51 faculty, staff, and students was recruited from a naturopathic university. Intraclass correlation coefficients were used to assess reproducibility of estimates of soy food intake and genistein and daidzein intake. Correlation coefficients comparing mean soy food servings per month between the 2 administrations of the questionnaire ranged from 0.50 for soy yogurt to 0.89 for tempeh. Correlation coefficients for genistein and daidzein intake estimated by the 2 administrations of the questionnaire were the same: 0.89. Mean intake (+/- standard deviation) of genistein and daidzein was 7 +/- 10 and 4 +/- 6 mg/day, respectively. Fifteen soy foods contributed 95% of the total genistein and daidzein intake: tofu, soy yogurt, tempeh, soy milk, low-fat tofu, soy flour, miso, soy protein isolate, low-fat soy milk, veggie soy burger, textured vegetable protein, miso soup, cooked soybeans, soy hot dogs, and natto (fermented soy beans). The soy food frequency questionnaire developed in this study provided highly reproducible estimates of soy food consumption and isoflavone intake. This instrument may be a useful tool in studies of the associations between isoflavone exposure and risk for chronic disease.

Posterior vitreous detachment induced by nattokinase (subtilisin NAT): a novel enzyme for pharmacologic vitreolysis.

PubMed

Takano, Akiomi; Hirata, Akira; Ogasawara, Kazuya; Sagara, Nina; Inomata, Yasuya; Kawaji, Takahiro; Tanihara, Hidenobu

2006-05-01

To investigate the effects of intravitreal injection of nattokinase (subtilisin NAT), a serine protease that is produced by Bacillus subtilis (natto), for induction of posterior vitreous detachment (PVD). Different doses of nattokinase (1, 0.1, or 0.01 fibrin-degradation units [FU]) or physiologic saline as a control were injected into the vitreous cavity of rabbit eyes. Scanning electron microscopy was used to observe the retinal surfaces of four rabbit eyes per concentration. Histologic alterations were assessed by light microscopy, using four eyes from each group. Electroretinography (ERG) was performed to observe retinal function, ranging from 1 hour to 1 week after the nattokinase (1 or 0.1 FU) or saline solution administration, using four eyes from each group at each time point. Also, findings in all rabbits were monitored by slit lamp examination and by indirect ophthalmoscopy with a 20-D lens. Scanning electron microscopy showed smooth retinal surfaces, indicating the occurrence of PVD at 30 minutes after intervention in all the experimental eyes injected with 0.1 or 1.0 FU nattokinase, but none of the control eyes. Light microscopy and ERG analysis showed no critical change even after the use of 0.1 FU nattokinase, an amount sufficient to induce PVD. However, toxicity in the forms of preretinal hemorrhage and ERG changes was noted with the higher dose (1 FU) of nattokinase. The results suggested that nattokinase is a useful enzyme for pharmacologic vitreolysis because of its efficacy in inducing PVD.

Suppression of initiation defects of chromosome replication in Bacillus subtilis dnaA and oriC-deleted mutants by integration of a plasmid replicon into the chromosomes.

PubMed

Hassan, A K; Moriya, S; Ogura, M; Tanaka, T; Kawamura, F; Ogasawara, N

1997-04-01

We constructed Bacillus subtilis strains in which chromosome replication initiates from the minimal replicon of a plasmid isolated from Bacillus natto, independently of oriC. Integration of the replicon in either orientation at the proA locus (115 degrees on the genetic map) suppressed the temperature-sensitive phenotype caused by a mutation in dnaA, a gene required for initiation of replication from oriC. In addition, in a strain with the plasmid replicon integrated into the chromosome, we were able to delete sequences required for oriC function. These strains were viable but had a slower growth rate than the oriC+ strains. Marker frequency analysis revealed that both pyrD and metD, genes close to proA, showed the highest values among the markers (genes) measured, and those of other markers decreased symmetrically with distance from the site of the integration (proA). These results indicated that the integrated plasmid replicon operated as a new and sole origin of chromosome replication in these strains and that the mode of replication was bidirectional. Interestingly, these mutants produced anucleate cells at a high frequency (about 40% in exponential culture), and the distribution of chromosomes in the cells was irregular. A change in the site and mechanism (from oriC to a plasmid system) of initiation appears to have resulted in a drastic alteration in coordination between chromosome replication and chromosome partition or cell division.

An updated review of dietary isoflavones: Nutrition, processing, bioavailability and impacts on human health.

PubMed

Zaheer, Khalid; Humayoun Akhtar, M

2017-04-13

Isoflavones (genistein, daidzein, and glycitein) are bioactive compounds with mildly estrogenic properties and often referred to as phytoestrogen. These are present in significant quantities (up to 4-5 mg·g -1 on dry basis) in legumes mainly soybeans, green beans, mung beans. In grains (raw materials) they are present mostly as glycosides, which are poorly absorbed on consumption. Thus, soybeans are processed into various food products for digestibility, taste and bioavailability of nutrients and bioactives. Main processing steps include steaming, cooking, roasting, microbial fermentation that destroy protease inhibitors and also cleaves the glycoside bond to yield absorbable aglycone in the processed soy products, such as miso, natto, soy milk, tofu; and increase shelf lives. Processed soy food products have been an integral part of regular diets in many Asia-Pacific countries for centuries, e.g. China, Japan and Korea. However, in the last two decades, there have been concerted efforts to introduce soy products in western diets for their health benefits with some success. Isoflavones were hailed as magical natural component that attribute to prevent some major prevailing health concerns. Consumption of soy products have been linked to reduction in incidence or severity of chronic diseases such as cardiovascular, breast and prostate cancers, menopausal symptoms, bone loss, etc. Overall, consuming moderate amounts of traditionally prepared and minimally processed soy foods may offer modest health benefits while minimizing potential for any adverse health effects.

Compound-specific nitrogen isotope analysis of D-alanine, L-alanine, and valine: application of diastereomer separation to delta15N and microbial peptidoglycan studies.

PubMed

Takano, Yoshinori; Chikaraishi, Yoshito; Ogawa, Nanako O; Kitazato, Hiroshi; Ohkouchi, Naohiko

2009-01-01

We have developed an analytical method to determine the compound-specific nitrogen isotope compositions of individual amino acid enantiomers using gas chromatography/combustion/isotope ratio mass spectrometry. A novel derivatization of amino acid diastereomers by optically active (R)-(-)-2-butanol or (S)-(+)-2-butanol offers two advantages for nitrogen isotope analysis. First, chromatographic chiral separation can be achieved without the use of chiral stationary-phase columns. Second, the elution order of these compounds on the chromatogram can be switched by a designated esterification reaction. We applied the method to the compound-specific nitrogen isotope analysis of D- and L-alanine in a peptidoglycan derived from the cell walls of cultured bacteria (Firmicutes and Actinobacteria; Enterococcus faecalis, Staphylococcus aureus, Staphylococcus staphylolyticus, Lactobacillus acidophilus, Bacillus subtilis, Micrococcus luteus, and Streptomyces sp.), natural whole bacterial cells (Bacillus subtilis var. natto), (pseudo)-peptidoglycan from archaea (Methanobacterium sp.), and cell wall from eukaryota (Saccharomyces cerevisiae). We observed statistically significant differences in nitrogen isotopic compositions; e.g., delta15N ( per thousand vs air) in Staphylococcus staphylolyticus for d-alanine (19.2 +/- 0.5 per thousand, n = 4) and L-alanine (21.3 +/- 0.8 per thousand, n = 4) and in Bacillus subtilis for D-alanine (6.2 +/- 0.2 per thousand, n = 3) and L-alanine (8.2 +/- 0.4 per thousand, n = 3). These results suggest that enzymatic reaction pathways, including the alanine racemase reaction, produce a nitrogen isotopic difference in amino acid enantiomers, resulting in 15N-depleted D-alanine. This method is expected to facilitate compound-specific nitrogen isotope studies of amino acid stereoisomers.

Impact of Interactions Between Self-Reported Psychological Stress and Habitual Exercise on the Dietary Intake of Japanese Men and Women: a Large-Scale Cross-Sectional Study.

PubMed

Endoh, Kaori; Kuriki, Kiyonori; Kasezawa, Nobuhiko; Tohyama, Kazushige; Goda, Toshinao

2016-01-01

Modifying lifestyle factors such as diet and exercise can reduce the risk of cancer. Psychological stress (PS) might be indirectly associated with cancer because it alters lifestyle factors. However, the relationship among these variables has not been fully investigated. Thus, we examined interactions between self-reported PS (SRPS) and habitual exercise on diet. In all, 5,587 men and 2,718 women were divided into "exerciser" and "non-exerciser" groups, based on whether they exercised regularly, and classified into three SRPS levels: low, moderate and high. Diet was estimated using a validated food frequency questionnaire. Using a general linear model, food and nutrient consumption was estimated for each SRPS level in the 2 exercise groups, and the interactions between SRPS levels and exercise were calculated. In women, the intake of pork and beef, low fat milk and yogurt, natto (fermented soybean), carrots and squash, other root vegetables, mushrooms, seaweeds, and wine, along with the nutrients vegetable protein, soluble, insoluble and total dietary fiber, daidzein, genistein, carotene, retinol equivalents, vitamin B2, pantothenic acid, potassium, calcium, magnesium, phosphorus and iron demonstrated significant interaction with SRPS and habitual exercise (p for interaction <0.05). In men, raw and green leafy vegetables and fruit and vegetable juice significantly interacted with SRPS and habitual exercise (p for interaction <0.05). We suggest that certain foods and nutrients, which are thought to have a protective effect against cancer, interact with SRPS and habitual exercise, especially in women. This information is valuable for understanding and improving interventions for cancer prevention.

Nattokinase decreases plasma levels of fibrinogen, factor VII, and factor VIII in human subjects.

PubMed

Hsia, Chien-Hsun; Shen, Ming-Ching; Lin, Jen-Shiou; Wen, Yao-Ke; Hwang, Kai-Lin; Cham, Thau-Ming; Yang, Nae-Cherng

2009-03-01

Nattokinase, a serine proteinase from Bacillus subtilis, is considered to be one of the most active functional ingredients found in natto. In this study, we hypothesized that nattokinase could reduce certain factors of blood clotting and lipids that are associated with an increase risk for cardiovascular disease (CVD). Thus, an open-label, self-controlled clinical trial was conducted on subjects of the following groups: healthy volunteers (Healthy Group), patients with cardiovascular risk factors (Cardiovascular Group), and patients undergoing dialysis (Dialysis Group). All subjects ingested 2 capsules of nattokinase (2000 fibrinolysis units per capsule) daily orally for 2 months. The laboratory measurements were performed on the screening visit and, subsequently, regularly after the initiation of the study. The intent-to-treat analysis was performed on all 45 enrolled subjects. By use of mixed model analysis, a significant time effect, but not group effect, was observed in the change from baseline of fibrinogen (P = .003), factor VII (P < .001), and factor VIII (P < .001), suggesting that the plasma levels of the 3 coagulation factors continuously declined during intake; also, the extents of decrease were similar between groups. After 2 months of administration, fibrinogen, factor VII, and factor VIII decreased 9%, 14%, and 17%, respectively, for the Healthy Group; 7%, 13%, and 19%, respectively, for the Cardiovascular Group; and 10%, 7%, and 19%, respectively, for the Dialysis Group, whereas blood lipids were unaffected by nattokinase. No significant changes of uric acid or notable adverse events were observed in any of the subjects. In summary, this study showed that oral administration of nattokinase could be considered as a CVD nutraceutical by decreasing plasma levels of fibrinogen, factor VII, and factor VIII.

Mediators of the effects of rice intake on health in individuals consuming a traditional Japanese diet centered on rice

PubMed Central

Toyomaki, Atsuhito; Miyazaki, Akane; Nakai, Yukiei; Yamaguchi, Atsuko; Kubo, Chizuru; Suzuki, Junko; Ohkubo, Iwao; Shimizu, Mari; Musashi, Manabu; Kiso, Yoshinobu; Kusumi, Ichiro

2017-01-01

Although the Japanese diet is believed to be balanced and healthy, its benefits have been poorly investigated, especially in terms of effects on mental health. We investigated dietary patterns and physical and mental health in the Japanese population using an epidemiological survey to determine the health benefits of the traditional Japanese diet. Questionnaires to assess dietary habits, quality of life, sleep quality, impulsivity, and depression severity were distributed to 550 randomly selected middle-aged and elderly individuals. Participants with any physical or mental disease were excluded. Two-hundred and seventy-eight participants were selected for the final statistical analysis. We determined rice to be one of the most traditional foods in Japanese cuisine. Scores for each questionnaire were computed, and the correlations between rice intake and health indices were assessed. When analyzing the direct correlations between rice intake and health indices, we found only two correlations, namely those with quality of life (vitality) and sleep quality. Path analysis using structural equation modeling was performed to investigate the association between rice intake and health, with indirect effects included in the model. Additional associations between rice intake and health were explained using this model when compared to those using direct correlation analysis. Path analysis was used to identify mediators of the rice-health association. These mediators were miso (soybean paste) soup, green tea, and natto (fermented soybean) intake. Interestingly, these mediators have been major components of the Japanese diet since 1975, which has been considered one of the healthiest diets since the 1960s. Our results indicate that the combination of rice with other healthy foods, which is representative of the traditional Japanese diet, may contribute to improvements in physical and mental health. PMID:28968452

Cryo-EM reconstruction of AlfA from Bacillus subtilis reveals the structure of a simplified actin-like filament at 3.4-Å resolution.

PubMed

Szewczak-Harris, Andrzej; Löwe, Jan

2018-03-27

Low copy-number plasmid pLS32 of Bacillus subtilis subsp. natto contains a partitioning system that ensures segregation of plasmid copies during cell division. The partitioning locus comprises actin-like protein AlfA, adaptor protein AlfB, and the centromeric sequence parN Similar to the ParMRC partitioning system from Escherichia coli plasmid R1, AlfA filaments form actin-like double helical filaments that arrange into an antiparallel bipolar spindle, which attaches its growing ends to sister plasmids through interactions with AlfB and parN Because, compared with ParM and other actin-like proteins, AlfA is highly diverged in sequence, we determined the atomic structure of nonbundling AlfA filaments to 3.4-Å resolution by cryo-EM. The structure reveals how the deletion of subdomain IIB of the canonical actin fold has been accommodated by unique longitudinal and lateral contacts, while still enabling formation of left-handed, double helical, polar and staggered filaments that are architecturally similar to ParM. Through cryo-EM reconstruction of bundling AlfA filaments, we obtained a pseudoatomic model of AlfA doublets: the assembly of two filaments. The filaments are antiparallel, as required by the segregation mechanism, and exactly antiphasic with near eightfold helical symmetry, to enable efficient doublet formation. The structure of AlfA filaments and doublets shows, in atomic detail, how deletion of an entire domain of the actin fold is compensated by changes to all interfaces so that the required properties of polymerization, nucleotide hydrolysis, and antiparallel doublet formation are retained to fulfill the system's biological raison d'être.

Fermented Soy Product Intake Is Inversely Associated with the Development of High Blood Pressure: The Japan Public Health Center-Based Prospective Study.

PubMed

Nozue, Miho; Shimazu, Taichi; Sasazuki, Shizuka; Charvat, Hadrien; Mori, Nagisa; Mutoh, Michihiro; Sawada, Norie; Iwasaki, Motoki; Yamaji, Taiki; Inoue, Manami; Kokubo, Yoshihiro; Yamagishi, Kazumasa; Iso, Hiroyasu; Tsugane, Shoichiro

2017-09-01

Background: Randomized controlled studies have investigated the short-term effect of soy product intake on blood pressure (BP) in normotensive people. To our knowledge, no prospective studies exist on the effect of habitual intake of fermented soy products, separate from total soy products, on BP in the general population. Objective: We examined the association between the habitual intake of soy products, including fermented soy products, and the development of high BP during a 5-y period among participants in a population-based prospective cohort study in Japan. Methods: The study included normotensive participants aged 40-69 y at baseline (926 men and 3239 women) who completed 2 questionnaires and whose BP was measured at the baseline survey between 1993 and 1994 and the 5-y follow-up in the Japan Public Health Center-Based Prospective Study Cohort II. The intake of soy products was assessed with a food-frequency questionnaire. High BP was defined as systolic blood pressure ≥130 mm Hg, diastolic blood pressure ≥85 mm Hg, or antihypertensive medication use. ORs and 95% CIs of high BP by frequency of soy products (miso, natto, and tofu) consumption, intake of total and fermented soy products, and intake of isoflavones from total and fermented soy products were estimated with the use of multiple logistic regression analysis. Results: Multivariable-adjusted ORs of high BP for the highest compared with the lowest tertile of total and fermented soy product intake were 1.03 (95% CI: 0.84, 1.25; P -trend = 0.786) and 0.72 (95% CI: 0.56, 0.92; P -trend = 0.009), respectively. The frequency of nonfermented soy product (tofu) intake was not associated with the development of high BP ( P -trend = 0.597). Conclusions: The intake of fermented soy products, but not total or nonfermented soy products, was inversely associated with developing high BP in men and women with normal BP. © 2017 American Society for Nutrition.

Stimulating effects of Bacillus subtilis natto-fermented Radix astragali on hyaluronic acid production in human skin cells.

PubMed

Hsu, Mei-Fang; Chiang, Been-Huang

2009-09-25

Radix astragali, a well-known Chinese herb, which has been traditionally used for skincare, and microbial fermentation is one of the conventional methods for processing Chinese herbs. This research studied the effects of non-fermented (HQNB) and fermented preparations (HQB) of Radix astragali on hyaluronic acid (HA) production in primary human skin cells. HQB and HQNB were prepared and added to the cultures of primary human skin cells. Hyaluronic acid content was determined using ELISA. Real-time RT-PCR was used to evaluate hyaluronan synthase gene expression. The bioactive compounds were analyzed by HPLC. The growth-stimulating effect of HQNB on both of keratinocytes and fibroblasts were significantly higher than that of HQB. Conversely, HQB, but not HQNB significantly stimulated HA production in both cultured primary human epidermal keratinocytes and human dermal fibroblasts in dose-dependent manners. In addition, HQB markedly and dose-dependently increased the expression of hyaluronan synthase 3 and hyaluronan synthase 2 mRNA in HaCaT cells and human fibroblasts, respectively. Therefore, HQB might be a promising candidate for preventing the age-dependent loss of HA content in aged human skin, and its effect on the enhancement of HA synthesis in skin cells is highly related to its effect on the expression of hyaluronan synthase genes. The three major active isoflavonoids in Radix astragali were identified as ononin, calycosin, and formononetin. After fermentation, all of these three compounds in HQB were significantly reduced. However, HQB still had significantly higher enhancement effect on the production of HA than HQNB. It appeared that isoflavonoid aglycones or other metabolites, converted from their primary isoflavones during fermentation, might be responsible for the skincare functions found in this study. This study demonstrated the low toxicity and the stimulating effects of HQB on HA synthesis, and suggests that HQB may play a promising role in anti-aging cosmetic applications.

Inoculating poultry manure with companion bacteria influences growth and development of black soldier fly (Diptera: Stratiomyidae) larvae.

PubMed

Yu, Guohui; Cheng, Ping; Chen, Yanhong; Li, Yongjian; Yang, Zihong; Chen, Yuanfeng; Tomberlin, Jeffery K

2011-02-01

The growth and development of black soldier fly, Hermetia illucens (L.), larvae fed chicken manure inoculated with bacteria isolated from black soldier fly larvae and associated larval feed was evaluated. Four strains of Bacillus subtilis were evaluated. B. subtilis strains S15, S16, S19, were isolated from the gut of black soldier fly larvae. B. natto strain D1 was isolated from the diet fed to black soldier fly larvae. These bacteria were added individually into nonsterile 200 g fresh hen manure at 10(6) cfu/g and homogenized. Treated manure was then inoculated with 4-d old black soldier fly larvae. Prepupal weight ranged from 0.0606 g in the control to 0.0946 g in manure treated with the S15 strain. Larval survivorship to the prepupal stage in all treatments ranged from 98.00 ± 2.65% to 99.33 ± 1.15%. Prepupal survivorship to the pupal stage ranged from 91.92 ± 1.87% to 97.95 ± 1.03%. Adult emergence from the pupal stage did not significantly (P < 0.05) differ across treatments and ranged from 98.95 ± 1.82% to 100.00 ± 0.00%. Adult body length resulting from the larvae in each of the treatments was significantly greater than those from the control. Longevity of adults did not differ significantly between treatments. Time from hatching to the development of the first pupa did not differ significantly across treatments; however, development time from hatching to 90% reaching the prepupual stage was significantly different between treatments and ranged from 29.00 ± 1.00 d to 34.33 ± 3.51 d. Development time from hatching to 90% reaching the adult stages was significantly different between treatments. Our results demonstrate that inoculating poultry manure with bacteria from black soldier fly larvae influences the growth and development of conspecific larvae feeding on the manure. © 2011 Entomological Society of America

Screening lactic acid bacteria to manufacture two-stage fermented feed and pelleting to investigate the feeding effect on broilers.

PubMed

Yeh, Ruei Han; Hsieh, Chia Wen; Chen, Kuo Lung

2018-01-01

Bacillus subtilis var. natto N21 (BS) and different lactic acid bacteria were applied to produce two-stage fermented feeds. Broilers were fed these feeds to select the best fermented feed. The selected fermented feed was pelleted and investigated for its effects on growth performance, carcass traits, intestinal microflora, serum biochemical constituents, and apparent ileal nutrient digestibility. Trial 1 involved three hundred thirty-six 1-d-old broilers with equal numbers of each sex, randomly assigned into control, BS + Bacillus coagulans L12 (BBC), BS + Lactobacillus casei (BLC), BS + Lactobacillus acidophilus (BLA), BS + Lactobacillus acidophilus L15 (BLA15), BS + Lactobacillus delbruekckii (BLD), and BS + Lactobacillus reuteri P24 (BLR24) groups with 3 replicates per group. Trial 2 involved two hundred forty 1-d-old broilers with equal numbers of each sex, randomly assigned into control, BBC, and pelleted BS + Bacillus coagulans L12 fermented feed (PBBC) groups with 4 replicates per group. Trial 3 involved sixteen 21-d-old male broilers randomly assigned into control and PBBC groups with 4 replicates per group for a nutrient digestibility trial. The feed conversion ratio (FCR) in the BBC group was better than the control (P < 0.05), and the production efficiency factor (PEF) was the best. However, weight gain (WG), feed intake (FI), and PEF were the lowest in the BLD group (P < 0.05). The WG during 0 to 21 d and 0 to 35 d in the PBBC groups were higher than the control (P < 0.05). The relative weight of the proventriculus + gizzard in the BBC and PBBC groups were higher than the control (P < 0.05). The digestible amino acid content in the PBBC group increased significantly (P < 0.05). Bacillus coagulans L12 is the best lactic acid bacteria for second stage fermentation. PBBC improved broiler growth performance, which may be due to the higher digestible amino acid content, it has the potential to become a commercial feed. © The Author 2017. Published by Oxford University Press on behalf of Poultry Science Association.

Screening lactic acid bacteria to manufacture two-stage fermented feed and pelleting to investigate the feeding effect on broilers

PubMed Central

Yeh, Ruei Han; Hsieh, Chia Wen; Chen, Kuo Lung

2018-01-01

Abstract Bacillus subtilis var. natto N21 (BS) and different lactic acid bacteria were applied to produce two-stage fermented feeds. Broilers were fed these feeds to select the best fermented feed. The selected fermented feed was pelleted and investigated for its effects on growth performance, carcass traits, intestinal microflora, serum biochemical constituents, and apparent ileal nutrient digestibility. Trial 1 involved three hundred thirty-six 1-d-old broilers with equal numbers of each sex, randomly assigned into control, BS + Bacillus coagulans L12 (BBC), BS + Lactobacillus casei (BLC), BS + Lactobacillus acidophilus (BLA), BS + Lactobacillus acidophilus L15 (BLA15), BS + Lactobacillus delbruekckii (BLD), and BS + Lactobacillus reuteri P24 (BLR24) groups with 3 replicates per group. Trial 2 involved two hundred forty 1-d-old broilers with equal numbers of each sex, randomly assigned into control, BBC, and pelleted BS + Bacillus coagulans L12 fermented feed (PBBC) groups with 4 replicates per group. Trial 3 involved sixteen 21-d-old male broilers randomly assigned into control and PBBC groups with 4 replicates per group for a nutrient digestibility trial. The feed conversion ratio (FCR) in the BBC group was better than the control (P < 0.05), and the production efficiency factor (PEF) was the best. However, weight gain (WG), feed intake (FI), and PEF were the lowest in the BLD group (P < 0.05). The WG during 0 to 21 d and 0 to 35 d in the PBBC groups were higher than the control (P < 0.05). The relative weight of the proventriculus + gizzard in the BBC and PBBC groups were higher than the control (P < 0.05). The digestible amino acid content in the PBBC group increased significantly (P < 0.05). Bacillus coagulans L12 is the best lactic acid bacteria for second stage fermentation. PBBC improved broiler growth performance, which may be due to the higher digestible amino acid content, it has the potential to become a commercial feed. PMID:29126320

Effects of the probiotic, Bacillus subtilis E20, on the survival, development, stress tolerance, and immune status of white shrimp, Litopenaeus vannamei larvae.

PubMed

Liu, Kuan-Fu; Chiu, Chiu-Hsia; Shiu, Ya-Li; Cheng, Winton; Liu, Chun-Hung

2010-01-01

In this study, the probiotic, Bacillus subtilis E20, isolated from the human health food, natto, was used for white shrimp, Litopenaeus vannamei, larvae breeding to improve the larval survival rate and development by adding probiotic to the rearing water at (control), 10(8), and 10(9) cfu L(-1) salt water once every 3 days during the 14 days of breeding experiment. Thereafter, stress tolerance and immune status of postlarvae were evaluated. Shrimp larval development was significantly accelerated after adding the probiotic to the larval rearing water at a level of 10(9) cfu L(-1). The survival rate of larvae was significantly higher in the treatment with 10(9) cfu L(-1) compared to the control and the treatment with 10(8) cfu L(-1) after all larvae had metamorphosed to postlarvae. Adding the probiotic to the shrimp larvae rearing water produced a weak inhibition of bacterial growth by an analysis of the total bacterial count and presumptive Vibrio count. For stress tests, no postlarvae died when they were reared in water in which the temperature was decreased from 30 to 2 degrees C at a rate of 0.1 degrees C min(-1). Postlarvae had significantly lower cumulate mortality in the treatments with 10(8) and 10(9) cfu L(-1) compared to the control when they were suddenly exposed to fresh water and 60 per thousand salt water. A significant decrease in the cumulative mortality of postlarvae treated with the probiotic at a level of 10(9) cfu L(-1) was recorded after the sudden transfer to 300 mg L(-1) nitrite-N compared to the control and treatment with 10(8) cfu L(-1). The analysis of immune-related gene expressions showed that the gene expression of prophenoloxidase I, prophenoloxidase II, and lysozyme of larvae were significantly increased after being reared in probiotic-containing water at the levels of 10(8) and 10(9) cfu L(-1). However, no significant difference in serine proteinase or glutathione peroxidase gene expressions was recorded in this study. It is therefore suggested that 10(9) cfu L(-1) of probiotic, B. subtilis E20 adding to rearing water for shrimp larva breeding. 2010 Elsevier Ltd. All rights reserved.

Cardiovascular Disease Death Before Age 65 in 168 Countries Correlated Statistically with Biometrics, Socioeconomic Status, Tobacco, Gender, Exercise, Macronutrients, and Vitamin K

PubMed Central

Agutter, Paul S

2016-01-01

Background Nutrition researchers recently recognized that deficiency of vitamin K2 (menaquinone: MK-4–MK-13) is widespread and contributes to cardiovascular disease (CVD). The deficiency of vitamin K2 or vitamin K inhibition with warfarin leads to calcium deposition in the arterial blood vessels. Methods Using publicly available sources, we collected food commodity availability data and derived nutrient profiles including vitamin K2 for people from 168 countries. We also collected female and male cohort data on early death from CVD (ages 15–64 years), insufficient physical activity, tobacco, biometric CVD risk markers, socioeconomic risk factors for CVD, and gender. The outcome measures included (1) univariate correlations of early death from CVD with each risk factor, (2) a multiple regression-derived formula relating early death from CVD (dependent variable) to macronutrient profile, vitamin K1 and K2 and other risk factors (independent variables), (3) for each risk factor appearing in the multiple regression formula, the portion of CVD risk attributable to that factor, and (4) similar univariate and multivariate analyses of body mass index (BMI), fasting blood sugar (FBS) (simulated from diabetes prevalence), systolic blood pressure (SBP), and cholesterol/ HDL-C ratio (simulated from serum cholesterol) (dependent variables) and dietary and other risk factors (independent variables). Results Female and male cohorts in countries that have vitamin K2 < 5µg per 2000 kcal/day per capita (n = 70) had about 2.2 times the rate of early CVD deaths as people in countries with > 24 µg/day of vitamin K2 per 2000 kcal/day (n = 72). A multiple regression-derived formula relating early death from CVD to dietary nutrients and other risk factors accounted for about 50% of the variance between cohorts in early CVD death. The attributable risks of the variables in the CVD early death formula were: too much alcohol (0.38%), too little vitamin K2 (6.95%), tobacco (6.87%), high blood pressure (9.01%), air pollution (9.15%), early childhood death (3.64%), poverty (7.66%), and male gender (6.13%). Conclusions Worldwide dietary vitamin K2 data derived from food commodities add much understanding to the analysis of CVD risk factors and the etiology of CVD. Vitamin K2 in food products should be systematically quantified. Public health programs should be considered to increase the intake of vitamin K2-containing fermented plant foods such as sauerkraut, miso, and natto. PMID:27688985

Cardiovascular Disease Death Before Age 65 in 168 Countries Correlated Statistically with Biometrics, Socioeconomic Status, Tobacco, Gender, Exercise, Macronutrients, and Vitamin K.

PubMed

Cundiff, David K; Agutter, Paul S

2016-08-24

Nutrition researchers recently recognized that deficiency of vitamin K2 (menaquinone: MK-4-MK-13) is widespread and contributes to cardiovascular disease (CVD). The deficiency of vitamin K2 or vitamin K inhibition with warfarin leads to calcium deposition in the arterial blood vessels. Using publicly available sources, we collected food commodity availability data and derived nutrient profiles including vitamin K2 for people from 168 countries. We also collected female and male cohort data on early death from CVD (ages 15-64 years), insufficient physical activity, tobacco, biometric CVD risk markers, socioeconomic risk factors for CVD, and gender. The outcome measures included (1) univariate correlations of early death from CVD with each risk factor, (2) a multiple regression-derived formula relating early death from CVD (dependent variable) to macronutrient profile, vitamin K1 and K2 and other risk factors (independent variables), (3) for each risk factor appearing in the multiple regression formula, the portion of CVD risk attributable to that factor, and (4) similar univariate and multivariate analyses of body mass index (BMI), fasting blood sugar (FBS) (simulated from diabetes prevalence), systolic blood pressure (SBP), and cholesterol/ HDL-C ratio (simulated from serum cholesterol) (dependent variables) and dietary and other risk factors (independent variables). Female and male cohorts in countries that have vitamin K2 < 5µg per 2000 kcal/day per capita (n = 70) had about 2.2 times the rate of early CVD deaths as people in countries with > 24 µg/day of vitamin K2 per 2000 kcal/day (n = 72). A multiple regression-derived formula relating early death from CVD to dietary nutrients and other risk factors accounted for about 50% of the variance between cohorts in early CVD death. The attributable risks of the variables in the CVD early death formula were: too much alcohol (0.38%), too little vitamin K2 (6.95%), tobacco (6.87%), high blood pressure (9.01%), air pollution (9.15%), early childhood death (3.64%), poverty (7.66%), and male gender (6.13%). Worldwide dietary vitamin K2 data derived from food commodities add much understanding to the analysis of CVD risk factors and the etiology of CVD. Vitamin K2 in food products should be systematically quantified. Public health programs should be considered to increase the intake of vitamin K2-containing fermented plant foods such as sauerkraut, miso, and natto.