Effect of free and conjugated bile salts on alpha-amylase activity.

The effect of individual bile salts on alpha-amylase hydrolysis of Cibachron Blue starch was studied at pH 6.0. With sodium cholate, taurocholate and taurodeoxycholate, enzyme activity was increased to 150-160 percent of the control value, at a concentration of similar to 1 mmol/l bile salt. The increased activity extended up to 4 mmol/l. The bile salts sodium deoxycholate and taurochenodeoxycholate exerted activation and inhibition depending on the concentration. With deoxycholate (0.75 mmol/l), activation (150 percent) was evident, while inhibition was apparent above 2.5 mmol/l. With taurochenodeoxycholate maximum activity (135 percent) was observed at 0.25 mmol/l, while inhibition was evident above 1.5 mmol/l. Chenodeoxycholate and lithocholate exerted marked inhibition at concentrations as low as 0.5 mmol/l. Inhibition of alpha-amylase by chenodeoxycholate was competitive with both soluble and insoluble starch substrates. Since the pH of the jejunum is in the region of 6.0 the phenomenon of activation and inhibition of alpha-amylase by bile salts at this pH could be of physiological significance.     

Transbuccal delivery of 5-Aza-2′-deoxycytidine: Effects of drug concentration,buffer solution,and bile salts on permeation

Delivery of 5-aza-2′-deoxycytidine (decitabine) across porcine buccal mucosa was evaluated as an alternative to the complex intravenous infusion regimen currently used to administer the drug. A reproducible high-performance liquid chromatography method was developed and optimized for the quantitative determination of this drug. Decitabine showed a concentration-dependent passive diffusion process across porcine buccal mucosa. An increase in the ionic strength of the phosphate buffer from 100 to 400 mM decreased the flux from 3.57±0.65 to 1.89±0.61 μg/h/cm². Trihydroxy bile salts significantly enhanced the flux of decitabine at a 100 mM concentration (P>.05). The steady-state flux of decitabine in the presence of 100 mM of sodium taurocholate and sodium glycocholate was 52.65±9.48 and 85.22±7.61 μg/cm²/h, respectively. Two dihydroxy bile salts, sodium deoxytaurocholate and sodium deoxyglycocholate, showed better enhancement effect than did trihydroxy bile salts. A 38-fold enhancement in flux was achieved with 10 mM of sodium deoxyglycocholate. Published: July 13, 2007     

Bile salt dependent bile flow in the rabbit: evidence for the importance of an amiloride inhibitable pathway

Bile salt dependent flow and electrolyte secretion in response to two bile salts were studied in awake rabbits. It was found that sodium glycodeoxycholate had a much greater choleretic and cholioneretic efficiency than sodium taurocholate. The effect of the bile salts on flow and electrolyte secretion was not linear across the range of bile salt secretion rates studied. When amiloride was administered significant decreases in choleretic and cholioneretic efficiencies occurred, but furosemide had no effect. It is concluded that bile salts stimulate electrolyte transport via amiloride inhibitable cellular processes, and that this electrolyte transport is in part responsible for bile salt dependent bile flow.     

Characterization and selection of <Emphasis Type="Italic">Lactobacillus</Emphasis> strains for their effect on bile tolerance,taurocholate deconjugation and cholesterol removal

Fourteen Lactobacillus strains of six species were investigated with their characteristics of bile salt tolerance, deconjugation of sodium taurocholate and cholesterol removal in the spent broth. Meanwhile, a co-precipitation curve of cholesterol with cholic acid at concentrations ranged 0.0–6.0 μM/ml was involved in the evaluation of cholesterol removal. Results demonstrated that both co-precipitation and assimilation effects contributed to cholesterol removal during the incubation of these Lactobacillus strains. It was also indicated that the supplementation of bile salts influenced the cholesterol removal, not only as an essential factor related to co-precipitation but also a critical condition for cholesterol assimilation. Out of all strains tested, four L. plantarum strains LS12, LS31, Lp501 and Lp529 exhibited a high ability of cholesterol assimilation (maximum 20.76 μg/ml), deconjugation of sodium taurocholate (maximum 5.00 μM/ml) and bile tolerance. They could be further studied and used as potential probiotics strains to reduce serum cholesterol in humans     

The effect of bile salts on survival and morphology of a potential probiotic strain Lactobacillus acidophilus M92

Bile tolerance is an important criterion in the selection of microbial strains for probiotic use. The survival and morphological changes of a potential probiotic strain, Lactobacillus acidophilus M92, in the presence of bile salts were examined. Lactobacillus acidophilus M92 has shown a satisfactory degree of tolerance against oxgall and individual bile salts tested, especially to taurocholate. The higher resistance of L. acidophilus M92 against taurine-conjugated bile salts relative to deconjugated and glycine-conjugated bile salts was attributed to its reaction to the stronger acidity of the former. Furthermore, bile salt hydrolase (BSH) was active when L. acidophilus M92 was grown in the presence of sodium taurocholate. The rate of BSH activity was highest at the exponential growth phase. It was hypothesised that BSH activity may be important for the bile salt resistance of this strain. The colonial and cellular morphology may also be a valuable parameter in the selection of bile salt-resistant Lactobacillus strains for probiotic use. Smooth (S) and rough (R) colonies, appeared in the original L. acidophilus M92 bacterial culture and demonstrated a different degree of bile tolerance. Rough colonies were more sensitive to bile salts than smooth ones. The R colony cells assumed a round form, probably induced by gaps in the cell wall caused by the cytotoxicity of glycodeoxycholate. This revised version was published online in July 2006 with corrections to the Cover Date.     

Dictyocaulus viviparus: The role of pepsin in the exsheathment of infective larvae

Following the report of Silverman and Podger (1964) that pepsin formed an association with larval receptor sites on D. viviparus and that exsheathment had an absolute requirement for pepsin, the role of pepsin was studied in greater detail. A range of enzyme incubation, pepsin labeling, histochemical and electron microscopical techniques were used. Pepsin did cause exsheathment of D. viviparus but, it was not an absolute requirement. Exsheathment occurred in a range of proteolytic enzymes each at its optimum pH. Findings suggest that the area of weakness around the anterior end of the larvae is digested by external protease and that, in vivo, exsheathment is caused by the gut enzymes of the host.     

Transbuccal delivery of 5-aza-2 -deoxycytidine: effects of drug concentration, buffer solution, and bile salts on permeation

Delivery of 5-aza-2 -deoxycytidine (decitabine) across porcine buccal mucosa was evaluated as an alternative to the complex intravenous infusion regimen currently used to administer the drug. A reproducible high-performance liquid chromatography method was developed and optimized for the quantitative determination of this drug. Decitabine showed a concentration-dependent passive diffusion process across porcine buccal mucosa. An increase in the ionic strength of the phosphate buffer from 100 to 400 mM decreased the flux from 3.57 +/- 0.65 to 1.89 +/- 0.61 microg/h/cm2. Trihydroxy bile salts significantly enhanced the flux of decitabine at a 100 mM concentration (P > .05). The steady-state flux of decitabine in the presence of 100 mM of sodium taurocholate and sodium glycocholate was 52.65 +/- 9.48 and 85.22 +/- 7.61 microg/cm2/h, respectively. Two dihydroxy bile salts, sodium deoxytaurocholate and sodium deoxyglycocholate, showed better enhancement effect than did trihydroxy bile salts. A 38-fold enhancement in flux was achieved with 10 mM of sodium deoxyglycocholate.     

Phospholipid fatty acid and lipopolysaccharide fatty acid signature lipids in methane-utilizing bacteria

The effect of human bile juice and bile salts (sodium cholate, sodium taurocholate, sodium glycochenodeoxycholate and sodium chenodeoxycholate) on growth, sporulation and enterotoxin production by enterotoxin-positive and enterotoxin-negative strains of Clostridium perfringens was determined. Each bile salt inhibited growth to a different degree. A mixture of bile salts completely inhibited the growth of enterotoxin-positive strains of this organism. Human bile juice completely inhibited the growth of all the strains at a dilution of 1:320. A distinct stimulatory effect of the bile salts on sporulation was observed in the case of C. perfringens strains NCTC 8239 and NCTC 8679. The salts also increased enterotoxin concentrations in the cell extracts of the enterotoxin-positive strains tested. No effect on enterotoxin production was detected when an enterotoxin-negative strain was examined.     

乳杆菌耐胆汁、降解结合胆盐和同化胆固醇能力的研究

对8株植物乳杆菌的胆汁耐受力、降解结合胆盐能力以及同化胆固醇能力进行了研究。不同的菌株在添加了牛胆汁的MRS中生长速度具有明显差异,同化胆固醇能力也明显不同,而降解结合胆盐的能力没有明显区别。分析发现,菌株的胆汁耐受力和降解结合胆盐能力,胆汁耐受力和同化胆固醇能力,以及降解结合胆盐能力和同化胆固醇能力之间都没有明显的相关性。     

Effects of bile salts on the adsorption of a hydrophobic mutagen to dietary fiber

The effect of the bile salts, sodium cholate, deoxycholate, glycocholate and taurocholate, on the solubility in aqueous solution of the hydrophobic, environmental mutagen, 1,8-dinitropyrene (DNP), was examined. In the absence of bile salts, the DNP appeared to precipitate out of solution, whereas bile salts at a concentration of greater than or equal to 4 mM maintained the DNP in solution. In the presence of the model dietary fiber, alpha-cellulose, the DNP absorbed to this preferentially. Bile salts reduced this adsorption at low alpha-cellulose levels, but had little effect at high alpha-cellulose levels. The implication of these results is that bile salts have solubilising properties that could affect the distribution of hydrophobic molecules, including mutagens, in the digestive tract.     

Bile salt related secretion of acid phosphatase in rat bile

The biliary excretion of bile salts, lysosomal acid phosphatase, and total proteins were studied in rats under different experimental conditions: during bile salt loss through a bile fistula and after loading with exogenous sodium taurocholate. The experimental models were suitable to demonstrate that variations in the excretion of bile salts were associated with those of acid phosphatase output. During bile salt depletion, acid phosphatase output showed a decrease parallel to that of bile salts. Following a single i.v. injection of sodium taurocholate and during its i.v. infusion, a rapid increase of acid phosphatase excretion in bile was seen. The patterns of enzyme outputs observed after administration of sodium taurocholate suggested a bulk discharge in bile of lysosomal contents. The profiles of protein output were similar to those of acid phosphatase suggesting an association between the secretory mechanism of these bile constituents. In contrast to sodium taurocholate, 4-methylumbelliferone, which also increases canalicular bile flow, did not produce changes in the excretory patterns of the bile components studied. Therefore, the results suggested a bile salt related secretion of acid phosphatase in the rat, which may involve protein secretion in bile.     

Effect of sodium taurocholate and sodium cholate on short-circuit current on amphibian membranes

The effects of the bile salts, sodium taurocholate (NaTc) and sodium cholate (NaCh), and toad bile gallbladder (bile) on short-circuit current (SCC) across isolated skin, and sodium taurocholate (NaTc) on isolated bladder of Bufo arenarum toads were tested. Sodium taurocholate (NaTc), sodium cholate (NaCh) and toad bile gallbladder (bile) promoted an increase in SCC, when added to the external side. The stimulatory effect was reversible after rinsing the preparation for 60 min. Implications on in vivo renal function of these results are discussed.     

Effects of divalent cations and sodium taurocholate on pancreatic lipase activity with gum arabic-emulsified tributyrylglycerol substrates.

The effects of Ca2+ and/or sodium taurocholate on lipase activity with gum arabic-emulsified tributyrylglycerol substrates were investigated. Calcium was found to slightly increase lipase activity while bile salts showed marked inhibition except at very low concentrations. Calcium eliminated inhibition seen with low concentrations of bile salts and reduced the inhibition seen at higher bile shift of the enzyme from the alkaline region in the absence of bile salt to the slightly acidic region in the presence of bile salt. Calcium was shown to eliminate the time lag periods between enzyme addition and maximum rate of hydrolysis seen at low substrate concentrations and the time lag noted when bile salts were included with normal (substrate concentration not limiting) assay concentrations of substrate. Zeta potential measurements indicated that Ca2+ reduced the negative charge on the gum arabic-emulsified particle while bile salts did not increase the negative charge. Commercial preparations of gum arabic were found to have significant concentrations of Ca2+ and Mg2+.     

Substrate specificity of human alpha-L-fucosidase.

Human α-l-fucosidase is a soluble lysosomal enzyme which hydrolyzes α-l-fucose residues linked to the 2 position of galactose or the 3, 4, or 6 position ofN-acetylglucosamine. Demonstration of activity towards natural oligosaccharide or glycosphingolipid substrates was achieved by measuring liberated l-fucose by coupling to fucose dehydrogenase and NAD and measuring NADH production spectrophotometrically. Activity of purified human spleen, brain, and cultured skin fibroblast or crude cell extracts towards 4-methylumbelliferyl-α-l-fucoside had a pH optimum of 4.5 to 5.5 and was unaffected by the presence of neutral detergents such as Triton X-100. However, the addition of sodium taurocholate or other bile salts to the incubation mixture caused a marked inhibition at pH 5 and a shift in pH optimum to the pH 6–7 region. Sodium taurocholate effected a threefold reduction in the apparent K_m for α-l-fucosidase at pH 6.0, but studies on fucosidosis tissue (α-fucosidase deficiency) or subcellular fractions derived from rat liver failed to indicate the existence of a membrane-bound α-l-fucosidase. The response of other lysosomal hydrolases to the presence of bile salts was investigated and was found to be variable, perhaps depending upon the hydrophilic or hydrophobic nature of the natural substrate and/or the state of association of the active enzyme.     

Enzymatic oxidation of unconjugated bilirubin to assess its interactions with taurocholate

The rate of peroxidation of unconjugated bilirubin (UCB), catalyzed by horseradish peroxidase (HRP), has been employed by Jacobsen (1969. FEBS Lett. 5: 112-114) to assess the fraction of unbound UCB in the presence of serum albumin. We used this method to examine the interactions of UCB with taurocholate (TC) at pH 8.2, assuming solubilization of UCB by TC is due to pigment binding and/or to partitioning into the micelle, thus rendering UCB unavailable for peroxidation. Inhibition of UCB peroxidation conformed with predictions based on these assumptions and demonstrated significant interaction of UCB with both monomeric and micellar TC. Although significant inhibition of UCB peroxidation was seen with TC monomer, inhibition was even greater with TC micelles. In contrast, pyrogallol, another substrate of HRP, acted very differently in the presence of TC. Inhibition of pyrogallol peroxidation by TC was much less than with UCB and occurred primarily with monomeric TC, with little further inhibition in the micellar range. The results of this study suggest that at taurocholate concentrations above 50 mM, similar to the physiologic bile salt concentrations in human bile, at least 99% of UCB is bound to bile salt, dramatically decreasing the concentration of unbound UCB. Since bile salts also bind Ca2+, they play a dual role in protection against the precipitation of calcium bilirubinate from bile. Therefore, bile salts are a major factor in the prevention of the formation and growth of pigment gallstones.     

Phospholipid-deacylating enzymes of rat stomach mucosa

1. Rat stomach mucosa exhibited three distinguishable phospholipid-deacylating enzyme activities: lysophospholipase, phospholipase A1 and phospholipase A2. 2. The lysophospholipase hydrolyzed 1-palmitoyl lysophosphatidylcholine to free fatty acid and glycerophosphorylcholine. This enzyme had an optimum pH of 8.0, was heat labile, did not require Ca2+ for maximum activity and was not inhibited by bile salts or buffers of high ionic strength. 3. Phospholipase A2 and phospholipase A1 deacylated dipalmitoyl phophatidylcholine to the corresponding lyso compound and free fatty acid. The specific activity of phospholipase A2 was 2--4-fold higher than that of phospholipase A1 under all the conditions tested. Both activities were enhanced 4--7.5-fold in the presence of bile salts at alkaline pH and 11-18-fold at acidic pH. 4. In the absence of bile salts, phospholipase A1 exhibited pH optima at 6.5 and 9.5 and phospholipase A2 at pH 6.5, 8.0 and 9.5. The pH optima for phospholipase A1 were shifted to pH 3.0, 6.0 and 9.0 in presence of sodium taurocholate; the activity was detected only at a single pH of 9.5 in the presence of sodium deoxycholate and at pH 10.0 in the presence of sodium glycocholate. Phospholipase A2 optimum activity was displayed at pH 3.0, 6.0 and 8.0 in presence of taurocholage, pH 7.5 and 9.0, in presence of glycocholate and only at pH 9.0 in presence of deoxycholate. 5. Ca2+ was essential for optimum activity of phospholipases A1 and A2. But phospholipase A1 lost complete activity in presence of 0.5 mM ethyleneglycolbis-(beta-aminoethylether)-N,N'-tetraacetic acid (EGTA) at pH 6.0, whereas phospholipase A2 lost only 50%. 6. Phospholipases A1 and A2 retained about 50% of their activities by heating at 75 degrees for 10 min. At 100 degrees, phospholipase A1 retained 22% of its activity, whereas phospholipase A2 retained only 7%.     

An active type IV secretion system encoded by the F plasmid sensitizes Escherichia coli to bile salts

F(+) strains of Escherichia coli infected with donor-specific bacteriophage such as M13 are sensitive to bile salts. We show here that this sensitivity has two components. The first derives from secretion of bacteriophage particles through the cell envelope, but the second can be attributed to expression of the F genes required for the formation of conjugative (F) pili. The latter component was manifested as reduced or no growth of an F(+) strain in liquid medium containing bile salts at concentrations that had little or no effect on the isogenic F(-) strain or as a reduced plating efficiency of the F(+) strain on solid media; at 2% bile salts, plating efficiency was reduced 10(4)-fold. Strains with F or F-like R factors were consistently more sensitive to bile salts than isogenic, plasmid-free strains, but the quantitative effect of bile salts depended on both the plasmid and the strain. Sensitivity also depended on the bile salt, with conjugated bile salts (glycocholate and taurocholate) being less active than unconjugated bile salts (deoxycholate and cholate). F(+) cells were also more sensitive to sodium dodecyl sulfate than otherwise isogenic F(-) cells, suggesting a selectivity for amphipathic anions. A mutation in any but one F tra gene required for the assembly of F pili, including the traA gene encoding F pilin, substantially restored bile salt resistance, suggesting that bile salt sensitivity requires an active system for F pilin secretion. The exception was traW. A traW mutant was 100-fold more sensitive to cholate than the tra(+) strain but only marginally more sensitive to taurocholate or glycocholate. Bile salt sensitivity could not be attributed to a generalized change in the surface permeability of F(+) cells, as judged by the effects of hydrophilic and hydrophobic antibiotics and by leakage of periplasmic beta-lactamase into the medium.     

Acid dissociation constants of bilirubin and related carboxylic acid compounds in bile salt solutions

Bilirubin, the yellow-orange tetrapyrrole pigment of jaundice, is essentially insoluble in pure water, but is much more soluble in solutions of bile salts such as sodium taurocholate. The biophysical chemistry of bilirubin in bile salt solutions is affected by changes in the pH of the solution in the range 5-9, suggesting that interactions with bile salt molecules and micelles may alter the acidity of the pigment. We have examined this possibility by determining the apparent pKa values for a series of carboxyl 13C-enriched model compounds, including the bilirubin analog mesobilirubin XIIIalpha, in solutions of sodium taurocholate and sodium taurodeoxycholate. Apparent pKa values were determined by 13C NMR titrations in dimethyl sulfoxide-water mixtures. The results show that the acidity of all compounds is decreased, or pKa increased, in micellar bile salt solution relative to pure water and that the effect is greatest for the larger, less water-soluble compounds. We have proposed a model to explain these results and discussed the implications of these findings for the biophysical chemistry of bilirubin in bile.     

Estimation of the contribution of halobacterial to the bacterial biomass and activity in solar salterns by the use of bile salts

Abstract Bile salts (deoxycholate, taurocholate) were used to estimate the contribution of bacteria of the Halobacterium group to bacterial community size and activity at different salinities as found in a multi-pond saltern. Low concentrations of bile salts cause lysis of halophilic archaebacteria of the Halobacterium group, while halophilic eubacteria and halococci remain microscopically intact. Upon addition of bile salts, total bacterial numbers (as estimated microscopically) in saltern ponds at salinities below 250 g/l did not decrease, while above this salinity bacterial numbers decreased by 30–50%. To estimate the contribution of halobacteria to overall heterotrophic activity, the effect of bile salt addition was tested on the incorporation of labelled amino acids. In saltern ponds of a salinity below 250 g/l activity was not greatly inhibited by taurocholate, while at salinity above 300 g/l taurocholate completely abolished incorporation of amino acids.     

Taurocholate is more potent than cholate in suppression of bile salt synthesis in the rat

Synthesis of bile salts is regulated through negative feedback inhibition by bile salts returning to the liver. Individual bile salts have not been distinguished with regard to inhibitory potential. We assessed inhibition of bile salt synthesis by either cholate or its taurine conjugate in bile fistula rats. After allowing synthesis to maximize, baseline synthesis was determined by measuring bile salt output in four consecutive 6-hr periods. Next, sodium cholate (+[(14)C]cholate) or taurocholate (+[(14)C]taurocholate) was infused into the jugular vein for 36 hr and bile was collected in 6-hr aliquots. Hepatic flux of exogenous bile salt was determined by measuring output of radioactivity in bile divided by specific activity of the infusate. Synthesis was determined during the last four 6-hr periods of infusion by subtracting exogenous bile salt secretion from the total bile salt output. Thirteen studies using cholate and 13 using taurocholate were performed. Hepatic flux of infused bile salt varied from 1 to 12 micro mol/100 g per rat per hr. Percent suppression of synthesis varied directly with hepatic flux of exogenous bile salt for both cholate and taurocholate in a linear fashion (r = 0.66, P < 0.01 and r = 0.87, P < 0.0005, respectively). Slope of the taurocholate line was 7.82 (% suppression/ micro mol per 100 g per hr), while slope of the cholate line was 3.66 (P < 0.05), indicating that taurocholate was approximately twice as potent as cholate in suppression of synthesis. At fluxes of 10-12 micro mol/100 g per hr, taurocholate suppressed synthesis 84 +/- 8 (SEM) % while cholate suppressed synthesis only 42 +/- 12% (P < 0.02). The x-intercept of the taurocholate line was 0.65 ( micro mol/100 g per hr), while that of the cholate line was -1.01 (NS) suggesting that the threshold for initial suppression of synthesis did not differ for these two bile salts. We conclude that taurocholate is a more effective inhibitor of hepatic bile salt synthesis than cholate, and that intestinal deconjugation of bile salts may play a role in the regulation of synthesis.-Pries, J. M., A. Gustafson, D. Wiegand, and W. C. Duane. Taurocholate is more potent than cholate in suppression of bile salt synthesis in the rat.