Seong-Il Lim

Microbial community analysis of Korean soybean pastes by next-generation sequencing

Soybean pastes, doenjang, have long been consumed as a fortified protein source in Korea. The quality of doenjang is determined by fermentation and Bacillus subtilis is suspected to be the main microorganism responsible. In the current culture-independent analysis, 17,675 bacterial sequences were derived from nine local and two commercial brands of doenjang samples by a barcoded pyrosequencing method targeting the hyper-variable regions V1/V2 of the 16S rRNA gene. In contrast to what has previously been found using plating or conventional molecular biology based methods, doenjang contains a diversity of bacterial species (total 208 species) and each doenjang reflects a region-specific bacterial community. While the Bacillus species was thought to be dominant in soybean pastes, we found that they were in high abundance (58.3-91.6%) only in samples from the central region of Korea, whereas lactic acid bacteria (LAB) (39.8-77.7%) were the dominant bacterial members of other doenjang samples. Compared to local brands of doenjang, commercial brands contain simple microbial communities dominated by Tetragenococcus and Staphylococcus that resemble the microbial communities of Japanese miso; this suggests that artificial inoculation was used for the quality control and standardization of doenjang. In this study, a massive sequencing approach was applied for the first time to analyze the microbial communities of different doenjang samples. Thus, we have determined that massive sequencing is a valid approach for assessing the overall microbial community of Korean fermented soybean pastes.

Hypolipidemic and Antioxidant Effects of Dandelion (Taraxacum officinale) Root and Leaf on Cholesterol-Fed Rabbits

Dandelion (Taraxacum officinale), an oriental herbal medicine, has been shown to favorably affect choleretic, antirheumatic and diuretin properties. Recent reports have indicated that excessive oxidative stress contributes to the development of atherosclerosis-linked metabolic syndrome. The objective of this current study was to investigate the possible hypolipidemic and antioxidative effects of dandelion root and leaf in rabbits fed with a high-cholesterol diet. A group of twenty eight male rabbits was divided into four subgroups; a normal diet group, a high-cholesterol diet group, a high-cholesterol diet with 1% (w/w) dandelion leaf group, and a high-cholesterol diet with 1% (w/w) dandelion root group. After the treatment period, the plasma antioxidant enzymes and lipid profiles were determined. Our results show that treatment with dandelion root and leaf positively changed plasma antioxidant enzyme activities and lipid profiles in cholesterol-fed rabbits, and thus may have potential hypolipidemic and antioxidant effects. Dandelion root and leaf could protect against oxidative stress linked atherosclerosis and decrease the atherogenic index.

Microbial Composition of the Korean Traditional Food “kochujang” Analyzed by a Massive Sequencing Technique

Kochujang is a traditional Korean fermented food that is made with red pepper, glutinous rice, salt, and soybean. Kochujang is fermented by naturally occurring microorganisms through which it obtains various health-promoting properties. In this study, the bacterial diversities of 9 local and 2 commercial brands of kochujang were analyzed with a barcoded pyrosequencing technique targeting the hyper-variable regions V1/V2 of the 16S rRNA gene. Through the analysis of 13524 bacterial pyrosequences, 223 bacterial species were identified, most of which converged on the phylum Firmicutes (average 93.1%). All of the kochujang samples were largely populated (>90.9% of abundance) by 12 bacterial families, and Bacillaceae showed the highest abundance in all but one sample. Bacillus subtilis and B. licheniformis were the most dominant bacterial species and were broadly distributed among the kochujang samples. Each sample contained a high abundance of region-specific bacterial species, such as B. sonorensis, B. pumilus, Weissella salipiscis, and diverse unidentified Bacillus species. Phylotype- and phylogeny-based community comparison analysis showed that the microbial communities of the two commercial brands were different from those of the local brands. Moreover, each local brand kochujang sample had region-specific microbial community reflecting the manufacturing environment.

Expression and Characterization of a Glutamate Decarboxylase from Lactobacillus brevis 877G Producing γ-Aminobutyric Acid

The glutamate decarboxylase of γ-aminobutyric acid-producing Lactobacillus brevis 877G (LbGAD) was expressed in Escherichia coli. The optimal pH and temperature for the purified LbGAD activity were respectively determined to be pH 5.2 and 45 °C. CaCl2 was shown to be a potent activator of this LbGAD activity. The kinetic parameters for LbGAD were a Km value of 3.6 mmol/L and a Vmax value of 0.06 mmol/L/min for L-monosodium glutamate.

Antidiabetic Effect of Morinda citrifolia (Noni) Fermented by Cheonggukjang in KK-Ay Diabetic Mice

Antidiabetic effects of Morinda citrifolia (aka Noni) fermented by Cheonggukjang (fast-fermented soybean paste) were evaluated using a T2DM (type 2 diabetes mellitus) murine model. Six-week-old KK-Ay/TaJcl mice were randomly divided into four groups: (1) the diabetic control (DC) group, provided with a normal mouse diet; (2) the positive control (PC) group, provided with a functional health food diet; (3) the M. citrifolia (MC) group, provided with an MC-based diet; (4) the fermented M. citrifolia (FMC) group, provided with an FMC-based diet. Over a testing period of 90 days, food and water intake decreased significantly in the FMC and PC groups compared with the DC group. Blood glucose levels in the FMC group were 211.60–252.20 mg/dL after 90 days, while those in the control group were over 400 mg/dL after 20 days. In addition, FMC supplementation reduced glycosylated hemoglobin (HbA1c) levels, enhanced insulin sensitivity, and significantly decreased serum triglycerides and low-density lipoprotein (LDL) cholesterol. Furthermore, a fermented M. citrifolia 70% ethanolic extract (FMCE) activated peroxisome proliferator-activated receptor-(PPAR-) γ and stimulated glucose uptake via stimulation of AMP-activated protein kinase (AMPK) in cultured C2C12 cells. These results suggest that FMC can be employed as a functional health food for T2DM management.

Optimization of Antibacterial Activity of Perilla frutescens var. acuta Leaf against Pseudomonas aeruginosa Using the Evolutionary Operation-Factorial Design Technique

This study was undertaken to optimize the conditions for the extraction of antibacterial activity of Perilla frutescens var. acuta leaf against Pseudomonas aeruginosa KCTC 2004 using the evolutionary operation-factorial (EVOP) design technique. Increased antibacterial activity was achieved at higher extraction temperatures and with a longer extraction time. Antibacterial activity was not affected by differing ethanol concentration in the extraction solvent. The maximum antibacterial activity of ethanolic extract of P. frutescens var. acuta leaf against P. aeruginosa, determined by the EVOP factorial technique, was obtained at an extraction temperature of 80 °C (R = −0.800**), 26 h (R = −0.731**) extraction time, and 50% (R = −0.075) ethanol concentration. The population of P. aeruginosa also decreased from 6.660 log CFU/mL in the initial set to 4.060 log CFU/mL in the third set. Also, scanning electron microscopy study of the ethanolic extract of P. frutescens var. acuta revealed potential detrimental effects on the morphology of P. aeruginosa.

Bioconversion of Capsaicin by Aspergillus oryzae

This study identified metabolites of capsaicin bioconverted by Aspergillus oryzae, which is generally used for mass production of gochujang prepared by fermenting red pepper powder in Korea. A. oryzae was incubated with capsaicin in potato dextrose broth. Capsaicin decreased depending on the incubation period, but new metabolites increased. Five capsaicin metabolites purified from the ethyl acetate fraction of the capsaicin culture were identified as N-vanillylcarbamoylbutyric acid, N-vanillyl-9-hydroxy-8-methyloctanamide, ω-hydroxycapsaicin, 8-methyl-N-vanillylcarbamoyl-6(E)-octenoic acid, and 2-methyl-N-vanillylcarbamoyl-6(Z)-octenoic acid by nuclear magnetic resonance (NMR) and mass spectrometry (MS). The capsaicin metabolites in gochujang were confirmed and quantitated by selective multiple reaction monitoring detection after liquid chromatography electrospray ionization MS using the isolated compounds as external standards. On the basis of the structures of the capsaicin metabolites, it is proposed that capsaicin metabolites were converted by A. oryzae by ω-hydroxylation, alcohol oxidation, hydrogenation, isomerization, and α- and/or β-oxidation.

Isolation of the Putative Biosynthetic Gene Cluster of 1-Deoxynojirimycin by Bacillus amyloliquefaciens 140N, Its Production and Application to the Fermentation of Soybean Paste

The 1-deoxynojirimycin (DNJ) biosynthetic gene cluster of Bacillus amyloliquefaciens 140N isolated from traditional Korean fermented food was isolated by PCR screening. It showed 78.9% inhibitory activity against α-glucosidase and produced 0.8 g/L of DNJ in an optimized medium containing 2% soluble starch, 1% tryptone, 0.05% KH(2)PO(4), and 0.05% (NH(2))(4)SO(4). Soybean paste fermented with B. amyloliquefaciens 140N produced DNJ with 84.4% inhibitory activity.

Anti-diabetic Effects of Fermented Green Tea in KK-A y Diabetic Mice

The anti-diabetic effect of green tea fermented by cheonggukjang was evaluated using KK-A y mice, an animal model of type 2 diabetes mellitus. Over a 90 day testing period, food and water intake decreased significantly in the group fed fermented green tea (FGT) and a group fed commercially available health functional food (PC), when compared with a diabetic control group (DC). The blood glucose levels of FGT mice were lower than in DC mice throughout the test period and were similar to the levels in PC after 60 days. Levels of Hemoglobin A1c (HbA1c) levels and insulin resistance were lower in mice of the FGT group than in mice of the DC group. DNA microarray analysis showed that administration of FGT increased the abundance of 12 mRNA transcripts related to diabetes. Whereas FGT increased hexokinase transcripts related to glycolysis more than 37 fold, levels of Pdx1 (pancreatic and duodenal homeobox1) and Cacna1e (calcium channel) transcripts increased more than 1.8 fold.

Genome Sequence of Lysinibacillus boronitolerans F1182, Isolated from a Traditional Korean Fermented Soybean Product

Lysinibacillus is a Gram-positive, rod-shaped, and round-spore-forming bacterial genus of the family Bacillaceae. We analyzed the genome sequence of Lysinibacillus boronitolerans F1182, isolated from a traditional Korean fermented soybean product. The genome sequence contained 4.46 Mbp with a G+C content of 37.5%. This is the first report of an L. boronitolerans genome.