• 한국미생물·생명공학회지
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• 제23권5호
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• pp.513-518
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• 1995

To evaluate bile salt-tolerance of lactic acid bacteria (LAB, Lactobacillus acidophilus ATCC 4356, Lactobacillus casei IFO 3533, Streptococcus thermnophilus KCTC 2185, Lactobacillus lactis ATCC 4797, and Lactobacillus bulgaricus ATCC 11842), We investigated the survivals, acid production and $\beta $-galactosidase activity of LAB under anaerobic broth system. Cellular permeability of LAB and their cellular retention of $\beta $-galactosidase were also examined in the same system. Although the growth of LAB was slightly suppressed by 0.3% bile salt, they showed normal growth curve. Streptococcus thermophilus KCTC 2185 was significantly more resistant to bile salt than the others. The $\beta $-galactosidase activity from Streptococcus thermophilus KCTC 2185 and Lactobacillus bulgaricus ATCC 11842 and their cellular retention of $\beta $-galactosidase decreased by 0.3% bile salt. The cellular permeability of LAB in the presence of bile salt increased significantly.

• Journal of Microbiology and Biotechnology
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• 제25권12호
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• pp.2100-2105
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• 2015

Probiotic bacteria must have not only tolerance against bile salt but also no genes for antibiotic resistance. Leuconostoc citreum is a dominant lactic acid bacterium in various fermented foods, but it is not regarded as a probiotic because it lacks bile salt resistance. Therefore, we aimed to construct a bile salt-resistant L. citreum strain by transforming it with a bile salt hydrolase gene (bsh). We obtained the 1,001 bp bsh gene from the chromosomal DNA of Lactobacillus plantarum and subcloned it into the pCB4170 vector under a constitutive P710 promoter. The resulting vector, pCB4170BSH was transformed into L. citreum CB2567 by electroporation, and bile salt-resistant transformants were selected. Upon incubation with glycodeoxycholic acid sodium salt (GDCA), the L. citreum transformants grew and formed colonies, successfully transcribed the bsh gene, and expressed the BSH enzyme. The recombinant strain grew in up to 0.3% (w/v) GDCA, conditions unsuitable for the host strain. In in vitro digestion conditions of 10 mM bile salt, the transformant was over 67.6% viable, whereas only 0.8% of the host strain survived.

• 한국가금학회지
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• 제27권3호
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• pp.227-233
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• 2000

Preset study was carried out to evaluate characteristics of lactic acid producing bacteria(LAB) in hen's cecum as probiotics value. Distribution of LAB in intestinal tracts was investigated using 5∼25 weeks - old hens. So, 12 strains to LAB with different morphology were isolated purely. Acid tolerance of LAB tested at pH 1, 2, 3, and 4, and bile resistant also tested at 0, 0.3% and 0.5% bile salt concentration. Growth pattern of LAB observed to 60h. All strains of cecal LAB couldn't survive at pH 1, and decreased linearly survival colony after incubation at pH 2 although some strains could survive for 2h. Most of LAB maintained constant number at pH 3 and 4. The bacterial action could increase linearly at 0% bile salt concentration in all of tested strains. However, only one strain could multiply at 0.3% bile salt, others were influenced by bile salt. That tendency was similar at 0.5% bile salt. Growth was peaked at 12 to 18 h after innoculation. After peak, the decreasing pattern of colony was different to strains which some strains decreased rapidly or maintained for long time. The LAB of hen's cecum was similar to intolerance acidity, but different to resistant to bile salt and growth pattern by strain. So, we choose three strains which have probiocs value, and identified as Lactobacillus amylovorus LLA7, Lactobacillus crispatus LLA9 and Lactobacillus vaginalis LLA11.

• Journal of Microbiology and Biotechnology
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• 제31권6호
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• pp.840-846
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• 2021

To improve the bile salt and acid tolerance of probiotics against gastrointestinal stresses, we investigated the effects of soybean lecithin and whey protein concentrate (WPC) 80 on the bile salt tolerance of Lacticaseibacillus paracasei L9 using a single-factor methodology, which was optimized using response surface methodology (RSM). The survival rate of L. paracasei L9 treated with 0.3% (w/v) bile salt for 2.5 h, and combined with soybean lecithin or WPC 80, was lower than 1%. After optimization, the survival rate of L. paracasei L9 incubated in 0.3% bile salt for 2.5 h reached 52.5% at a ratio of 0.74% soybean lecithin and 2.54% WPC 80. Moreover, this optimized method improved the survival rate of L. paracasei L9 in low pH condition and can be applied to other lactic acid bacteria (LAB) strains. Conclusively, the combination of soybean lecithin and WPC 80 significantly improved the bile salt and acid tolerance of LAB. Our study provides a novel approach for enhancing the gastrointestinal tolerance of LAB by combining food-derived components that have different properties.

• Asian-Australasian Journal of Animal Sciences
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• 제18권10호
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• pp.1505-1512
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• 2005

Bile salt deconjugation is the most biologically significant reaction among the bacterial alterations of bile acids in the gastrointestinal tract of human and animal. The responsible enzyme, bile salt hydrolase (BSH), catalyzes the hydrolysis of glycineand/or taurine-conjugated bile salts into amino acid residues and deconjugated bile acids. Herein we review current knowledge on the distribution of BSH activity among various microorganisms with respect to their biochemical and molecular characteristics. The proposed physiological impact of BSH activity on the host animal as well as on the BSH-producing bacterial cells is discussed. BSH activity of the probiotic strains is examined on the basis of BSH hypothesis, which was proposed to explain cholesterol-lowering effects of probiotics. Finally, the potential applications of BSH research are briefly discussed.

• Journal of Microbiology and Biotechnology
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• 제21권8호
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• pp.838-845
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• 2011

The present work describes the identification, purification, and characterization of bile salt hydrolase (BSH) from Bifidobacterium animalis subsp. lactis. The enzyme was purified to electrophoretic homogeneity by hydrophobic chromatography, ion-exchange chromatography and ultrafiltration. SDS-PAGE analysis of putative BSH and gel filtration revealed that the analyzed protein is presumably a tetramer composed of four monomers each of about 35 kDa. The purified enzyme was analyzed by liquid chromatography coupled to LTQ FT ICR mass spectrometry and unambiguously identified as a bile salt hydrolase from B. animalis. The isoelectric point of the studied protein was estimated to be around pH 4.9. The pH optimum of the purified BSH is between 4.7 to 6.5, and the temperature optimum is around 50oC. The BSH of B. animalis could deconjugate all tested bile salts, with clear preference for glycine-conjugated bile salts over taurine-conjugated forms. Genetic analysis of the bsh showed high similarity to the previously sequenced bsh gene from B. animalis and confirmed the usefulness of bile salt hydrolase as a genetic marker for B. animalis identification.

• Journal of Dairy Science and Biotechnology
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• 제29권1호
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• pp.49-54
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• 2011

담즙산 분해효소(Bile salt hydrolase, EC 3.5.1.24) 활성은 담즙산의 카르복실기와 결합되어 있는 아미노기(glycine or taurine)와의 amide 결합을 끊는 작용을 하며, 이 효소 활성은 사람이나 동물의 장내 미생물들에서 널리 분포하고 있다. 노인 분변으로부터 분리한 여러 균주의 Enterococcus faecalis중에서 BSH activity가 가장 높은 CU30-2를 선발하였다. BSH 유전자를 pET22b expression vector에 클로닝하여 Escherichia coli BL21(DE3) Gold를 이용하여 단백질을 발현하였다. 6x His-tag이 있는 BSH 효소를 $Ni^+$-NTA agarose column을 이용하여 분리 정제하였고, 6가지의 다른 담즙산염을 이용하여 기질 특이성을 비교하였다. E. faecalis CU30-2의 BSH 효소는 glycine이 결합된 담즙산염에 대한 효소활성이 taurine이 결합된 것에 대한 활성보다 약 50배 정도 높게 나타났다. 이 효소의 최적 pH와 온도는 각각 7.0과 40$^{\circ}C$로 확인되었다.

• 대한약리학회지
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• 제18권1호
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• pp.43-54
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• 1982

Bile formation is a complex process comprised of three separate physiologic mechanism operating at two anatomical sites. At present time, it was known that at least two processes are responsible for total canalicular secretion at the bile canaliculus. One of the processes is bile salt-dependent secretion (BSDS) hypothesis that the active transport of bile salts from plasma to bile provided a primary stimulus for bile formation: the osmotic effect of actively transported bile acid was responsible for the movement of water and ions into bile. The other process is bile salt-independent secretion (ESIS), which is unrelated to bile salt secretion at the canaliculus and which may involve the active transport of sodium. The third process for bile formation involves the biliary ductal epithelium. Secretin-stimulated bile characteristically contained bicarbonate in high concentration. Therefor, it was suggested that secretin stimulated water and bicarbonate secretion from the biliary ductules. One the other hand, it was found that a large amounts of cAMP was present in canine bile but no apparent relationship between bile salt secretion and cAMP content in dog bile. However, bile flow studies in human have demonstrated that secretin and glucagon increase bile cAMP secretion as does secretin in baboons. Secretin increases baboon bile duct mucosal cAMP levels in addition to bile CAMP levels suggesting that in that species secretin-stimulated bile flow may be cAMP mediated. It has been postulated that glucagon and theophylline which increase the bile salt-independent secretion in dogs might act through an increased in liver cAMP content. In a few studies, the possible role of cAMP on bile formation has teen tested by administration of an exogenous derivative of cAMP, dibutyryl cAMP. In the rat, DB cAMP did not modify bile flow, but injection of DB cAMP in the dog promoted an increase in the bile salt-independent secretion. Because of these contradictory results, this study was carried out to examine the relationship between cyclic nucleotides and bile flow due to various bile salts as well as secretin or theophylline. Experiments were performed in rabbits with anesthesia produced by the injection of seconal(30 mg/kg). Rabbits had the cystic duct ligated and the proximal end of the divided common duct cannulated with an appropriately sized polyethylene catheter. A similar catheter was placed into the inferior vena cava for administration of drugs. Bile was collected for determination of cyclic nucleotides and total cholate in 15 min. intervals for a few hours. The results are summerized as followings. 1) Administrations of taurocholic acid or chenodeoxycholic acid increased significantly the concentrations of cAMP and cGMP in bile of rabbits. 2) Concentration of cAMP in bile during the continuous infusion of ursodeoxycholic acid, was remarkedly increased in accordance with the increase of bile flow, while on the contrary concentration of cGMP in bile was decreased significantly. 3) Dehydrocholic acid and deoxycholic acid significantly increased bile flow, total cholate output and cyclic nucleotides in bile. 4) Only cAMP concentration in bile was significantly increased from control value by secretin, while theophylline increased cAMP as well as cGMP in rabbit bile. 5) In addition, the administration of secretin to taurocholic acid-stimulated bile flow increased cAMP while theophylline produced the increases of cAMP and cGMP in bile. 6) The administration of insulin to taurocholic acid-stimulated bile flow decreased cAMP concentration, while on the contrary cGMP was remarkedly increased in rabbit bile.

• 한국미생물·생명공학회지
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• 제38권2호
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• pp.222-226
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• 2010

This study was carried out to isolate and characterize the Lactobacillus plantarum with bile salt deconjugation activity that was isolated from Kimchi. Some isolates were selected and identified as L. plantarum by 16S rRNA gene sequence and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of whole cell protein patterns. They were assayed to determine their capacities to express bile salt hydrolase (BSH) activity. Among the identified strains, L. plantarum CIB 001 showed the highest level of BSH activity. Then, resistance to gastric acidity and bile condition were analyzed for further characterization. This strain was able to maintain viability for 1h at pH 2.0 and to survive in a MRS (deMan, Rogosa, and Sharpe) broth with 1.0% of bile acids. L. plantarum CIB 001 would potentially be useful in the food industry as probiotics.

• 한국미생물·생명공학회지
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• 제50권1호
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• pp.102-109
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• 2022

Bile acids are essential molecules produced by vertebrates that are involved in several physiological roles, including the uptake of nutrients. Bacterial isolates capable of producing bile acids de novo have been identified and characterized. Such isolates may provide access to novel biochemical pathways suitable for the design of microbial cell factories. Here, we further characterized the ability of Maribacter dokdonensis, Dokdonia donghaensis, and Myroides pelagicus to produce bile acids. Contrary to previous reports, we did not observe de novo production of bile acids by these isolates. Instead, we found that these isolates deconjugated the amino acid moiety of bile acids present in the growth medium used in previous reports. Through genomic analysis, we identified putative bile salt hydrolases, which could be responsible for the different bile acid modifications observed. Our results challenge the hypothesis of de novo microbial bile acid production, while further demonstrating the diverse capacity of bacteria to modify bile acids.