Presence Of Sodium Taurocholate Research Articles

Impact of Swelling of Spray Dried Dispersions in Dissolution Media on their Dissolution: An Investigation Based on UV Imaging

Impact of SDD-dissolution medium interactions on the swelling and dissolution of spray dried dispersions (SDDs) was investigated using UV imaging by monitoring SDD swelling in situ, along with correlating of the swelling with the micro-dissolution and intrinsic dissolution of SDDs. SDDs of ketoconazole or indomethacin with three polymers: polyvinylpyrrolidone (PVP), hydroxypropyl cellulose (HPC), and hydroxypropyl methylcellulose acetate succinate (HPMC-AS) were prepared for the study. Dissolution media employed for assessing swelling and dissolution include water, acetate buffer, phosphate buffer, fasted state simulated intestinal fluid (FaSSIF), and fed state simulated intestinal fluid (FeSSIF), in which influence of polymers and drugs together with the physical-chemical properties of dissolution media (pH, and the presence of sodium taurocholate and lecithin) on SDD swelling and dissolution was evaluated. It appears that hydrophilic and hydrophobic properties of polymers can significantly impact SDD swelling and thus the dissolution. Furthermore, properties of dissolution media such as pH as well as presence of bile salts and lecithin seems to affect SDD swelling and dissolution as well. Throughout the text, thermodynamic swelling of polymers was used to interpret SDD dissolution behavior. Finally, practical implication of polymer swelling on dissolution was discussed.

In vitro inhibition of Clostridium difficile by commercial probiotics: A microcalorimetric study

The aim of the study was to investigate the influence of some commercial probiotics on the growth of Clostridium difficile using isothermal microcalorimetry, a technique which can monitor the real time growth of bacteria. Commercial probiotic strains and products, Lactobacillus acidophilus LA-5®, Bifidobacterium lactis BB-12®, Probio 7® and Symprove™ were co-cultured with C. difficile in Brain Heart Infusion (BHI) broth supplemented with 0.1% (w/v) l-cysteine hydrochloride and 0.1% (w/v) sodium taurocholate and monitored in the microcalorimeter. Pseudomonas aeruginosa NCIMB 8628 was also co-cultured with C. difficile and studied. The results indicated inhibition of C. difficile by the probiotics. The inhibition of C. difficile was shown to be pH-dependent using neutralized and unmodified cell free supernatant (CFS) produced by the probiotic strains. However, concentrated CFS of the probiotics also inhibited the intestinal pathogen in a non pH-dependent manner, likely due to specific antimicrobial substances produced. The results also indicated that C. difficile growth was greatly influenced by the presence of sodium taurocholate and by the pH of the medium. A medium pH of between 6.45 and 6.9 demonstrated maximum growth of the organism in the microcalorimeter.

Biochemical Characterization and Comparison of Pancreatic Lipases from the Pacific Bluefin Tuna, Thunnus orientalis; Totoaba, Totoaba macdonaldi; and Striped Bass, Morone saxatilis

AbstractLipolytic activity from pancreatic crude extracts from the Pacific bluefin tuna (PBT), Thunnus orientalis; totoaba, Totoaba macdonaldi; and striped bass, Morone saxatilis, were biochemically characterized to detect possible differences in the digestive capacity of each fish. The maximum activity was at temperatures ranging from 35 to 45 C and at pH 8.0; the activity decreased significantly at a pH lower than 7.0. Lipases from PBT and totoaba showed an increase in activity with the presence of sodium taurocholate (NaTC; 6 mM). However, no increase in activity could be observed when a mixture of bile salts or natural bile extracts was used in the reaction mixture, which was observed in the presence of NaTC. In contrast, the lipase activity from striped bass increased accordingly when NaCl was added at different levels (0–1.5 M). Calcium was not required in the reaction mixture for any of the fish species tested. We concluded that all three lipases have different physicochemical properties and ionic strength and emulsifier requirements to obtain their maximum activity. Striped bass was far too sensitive to NaCl, while totoaba and PBT were sensitive to NaTC. The differences in activity shown here suggest the possibility of studying the use of different additives on feed formulations to maximize lipase efficiency.

Characterization, phylogenetic affiliation and probiotic properties of high cell density Lactobacillus strains recovered from silage

The aim of the present study was to isolate high cell density Lactobacillus (LAB) from different forages and select the best strains for production of silage with improved the lactic acid production. Twenty heterofermentative LAB strains were selected and their probiotic properties were analyzed by evaluating their tolerance to low pH, bile salts, biogenic amine production, enzyme activity, antibiotic susceptibility pattern and antifungal activity. The 16S rRNA gene-based phylogenetic affiliation indicated that 16 strains were Lactobacillus plantarum and others were L. bobalius, L. zymae, L. crustorum and L. diolivorans. Shake-flask cultivation of these strains under aerobic conditions showed comparatively higher growth and organic acid production than that achieved using the well-studied LAB strains. In addition, all the strains were highly sensitive towards ox gall (0.3%), but grew well in the presence of sodium taurocholate (0.3%). Antimicrobial susceptibility pattern is an intrinsic feature of these LAB strains; thus consumption does not represent a health risk to humans. Lactobacillus plantarum strains exhibited considerable antifungal activity against food pathogens. The present finding raises the possibility that high cell density LAB strains with potential probiotic properties could be used to prepare quality silages for animals.

Improved oral delivery of resveratrol using proliposomal formulation: investigation of various factors contributing to prolonged absorption of unmetabolized resveratrol

Objectives: The objective of this study was to design lipid-based formulation to enhance the absorption of unmetabolized resveratrol (RSV) over adequate time and investigate various factors that contribute to prolonged absorption of RSV.Methods: Proliposomal formulations containing distearoyl phosphatidyl choline (DSPC) with or without cholesterol were prepared and evaluated. The liposomes obtained from hydration of proliposomal mixture were evaluated for size, zeta, physical appearance and entrapment. The integrity of liposomes in bile salt solution and solubility of RSV in sodium taurocholate solution in the presence of various concentrations of DSPC were evaluated to assess the stability and in varied gastrointestinal conditions. Finally, oral pharmacokinetic studies of liposomal dispersions in comparison with RSV solution were evaluated.Results: Results revealed that spontaneous formation of liposomes did not occur upon hydration of RSV: DSPC proliposomes rather showed tendency to form loose cotton-like aggregates. Cholesterol aided in the formation of stable liposomes with large negative zeta potential. Release of RSV from liposomes in the presence of taurocholate was dependent on the amount and type of total lipid. Liposomes without cholesterol showed faster release, and release increased as the amount of DSPC in the formulation increased. Solubility studies indicated that DSPC increases the solubility of RSV in the presence of sodium taurocholate, and corroborates that bilayer assembly is disrupted because of interaction between RSV and DSPC. Mixture of RSV:DSPC:Chol at 1:0.25:0.25 formed stable colloidal dispersion with zeta potential −22 and released only 20 – 23% of entrapped RSV when incubated with 20 mM sodium taurocholate. Pharmacokinetic profile revealed that AUC and Cmax were twofold higher than plain RSV.Conclusion: The proliposomal formulation optimized by considering various physicochemical factors and simulated in vitro testing result in significant improvement rate and extent of absorption of unmetabolized RSV.

Variations in bile tolerance among Lactococcus lactis strains derived from different sources

Lactococcus lactis subsp. lactis has been isolated from the intestines of marine fish and is a candidate probiotic for aquaculture. In order to use the bacterium as a probiotic, properties such as bile tolerance need to be assessed. Here, we compared bile tolerance in L. lactis strains derived from different sources. Three L. lactis subsp. lactis strains from marine fish (MFL), freshwater fish (FFL), and cheese starter (CSL) were used along with an Lactococcus lactis subsp. cremoris strain from cheese starter (CSC). The four strains were grown under various culture conditions: deMan-Rogosa-Sharpe (MRS) broth containing bile salts/acids, MRS agar containing oxgall, and phosphate-buffered saline (PBS) containing fish bile. Survival/growth of the strains in the presence of sodium cholate and sodium deoxycholate varied in the order MFL, CSL > CSC > FFL; in the presence of sodium taurocholate, the order was MFL > CSL > CSC > FFL. In liquid media containing various concentrations of oxgall, survival of the strains was observed in the order MFL > CSL > FFL and CSC. The survival of MFL was not affected by bile collected from the goldfish (Carassius auratus subsp. auratus) or the puffer fish (Takifugu niphobles), although the other strains showed significant inhibition of growth. It is a novel and beneficial finding that MFL has the highest resistance to bile acid.

Determining probiotic potential of exopolysaccharide producing lactic acid bacteria isolated from vegetables and traditional Indian fermented food products

Abstract Exopolysaccharide producing lactic acid bacterial (LAB) were tested in vitro to select a candidate probiotic strain by testing their tolerance to low pH and bile, bile salt hydrolase (BSH) activity, antibiotics susceptibility pattern and antimicrobial activity. Results indicated that LAB isolates were highly sensitive against oxbile (0.3%) but could grow in the presence of sodium taurocholate (0.3%). Seven out of 9 isolates were found to be BSH positive. Two of the isolates, Lactobacillus plantarum 86 and Weissella cibaria 92 showed considerable antimicrobial activity against Gram-positive and Gram-negative pathogens. The study reveals that traditional fermented products of India could be an alternative and readily available resource for LAB starter cultures with interesting functional characteristics.

Isolation and characterization of antifungal compound from Lactobacillus plantarum KCC-10 from forage silage with potential beneficial properties

The purpose of this study was to isolate, identify and characterize an antifungal compound from Lactobacillus plantarum KCC-10 from forage silage with potential beneficial properties. The 16S rRNA gene-based phylogenetic affiliation was determined using bioinformatic tools and identified as Lactobacillus sp. KCC-10 with 100% sequence similarity to L. plantarum. The antifungal substances were extracted with ethyl acetate from spent medium in which Lactobacillus sp. KCC-10 was cultivated. Antifungal activity was assessed using the broth microdilution technique. The compounds were obtained by eluting the crude extract with various concentrations of solvents followed by chromatographic purification. Based on the infrared, (13) C nuclear magnetic resonance (NMR) and (1) H NMR spectral data, the compound was identified as a phenolic-related antibiotic. The minimum inhibitory concentration of the compound against Aspergillus clavatus, A. oryzae, Botrytis elliptica and Scytalidium vaccinii was 2.5 mg ml(-1) and that against A. fumigatus, A. niger and S. fusca was 5.0 mg ml(-1) , respectively. In addition, Lactobacillus sp. KCC-10 was highly sensitive towards oxgall (0.3%) but grew well in the presence of sodium taurocholate (0.3%). An antimicrobial susceptibility pattern was an intrinsic feature of this strain; thus, consumption does not represent a health risk to humans or animals. Novel L. plantarum KCC-10 with antifungal and potential probiotic properties was characterized for use in animal food. This study revealed that L. plantarum KCC-10 exhibited good antifungal activity similar to that of probiotic Lactobacillus strains.

Sodium taurocholate-dependent lipid efflux by ABCA1: effects of W590S mutation on lipid translocation and apolipoprotein A-I dissociation

ABCA1 plays a major role in HDL metabolism. Cholesterol secretion by ABCA1 is dependent on the presence of extracellular acceptors, such as lipid-free apolipoprotein A-I (apoA-I). However, the importance of the direct interaction between apoA-I and ABCA1 in HDL formation remains unclear. In contrast, ABCB4 mediates the secretion of phospholipids and cholesterol in the presence of sodium taurocholate (NaTC) but not in the presence of apoA-I. In this study, we analyzed apoA-I binding and NaTC-dependent lipid efflux by ABCA1. ABCA1 mediated the efflux of cholesterol and phospholipids in the presence of NaTC as well as in the presence of apoA-I in an ATP-dependent manner. The Tangier disease mutation W590S, which resides in the extracellular domain and impairs apoA-I-dependent lipid efflux, greatly decreased NaTC-dependent cholesterol and phospholipid efflux. However, the W590S mutation did not impair apoA-I binding and, conversely, retarded the dissociation of apoA-I from ABCA1. These results suggest that the W590S mutation impairs ATP-dependent lipid translocation and that lipid translocation or possibly lipid loading, facilitates apoA-I dissociation from ABCA1. NaTC is a good tool for analyzing ABCA1-mediated lipid efflux and allows dissection of the steps of HDL formation by ABCA1.

Propriétés des formes moléculaires de la β-glucosidase et de la β-glucocéréhrosidase de rate humaine normale et de maladie de Gaucher

Properties of the Molecular Forms of β-Glucosidase and β-Glucocerebrosidase from Normal Human and Gaucher Disease Spleen Comparative studies of β-glucosidases and β-glucocerebrosidases from normal human and Gaucher disease spleen have allowed to identify three enzymatic groups on the basis of their subcellular localization (soluble or membrane-bound), enzymatic and genetic properties. β-Glucocerebrosidase cleaves both the natural substrate, β-glucocerebroside and the artificial substrate, methylumbelliferyl(MeUmb)-β-glucoside. A minor part (25%) is soluble in water whereas 75% of the total activity is membrane-bound and can be solubilized by 0.25% Triton X-100. Using preparative electrofocusing, the soluble β-glucocerebrosidase shows two molecular forms (pI 5.2 ± 0.2 and 6.4 ± 0.2). Exactly the same forms are found for the membrane-bound β-glucocerebrosidase solubilized by Triton X-100. All these molecular forms display the same enzymatic properties: optimum pH of 5.4, heat-stability at 42° C, Km values of 3.7 ± 0.5 mM for the artificial substrate in the presence of sodium taurocholate and 0.06 ± 0.01 mM for the natural substrate. In addition they are activated at the same extent (600%) by sodium taurocholate. Furthermore, these two molecular forms are coded by the same gene since both are deficient in type I non neuropathic Gaucher disease. The soluble non specific MeUmb-fl-glucosidase catalyses the hydrolysis of the artificial substrate but not of the natural substrate and is focused in one single peak (pI 4.7 ± 0.2) different from the two forms of β-glucocerebrosidase discussed above. It is strongly inhibited by sodium taurocholate, weakly by Triton X-100 and is heat-labile at 42° C. No deficiency is detected in the case of type I Gaucher disease reported here. it is thus suggested that this enzyme corresponds to a gene different from that coding for β-glucocerebrosidase. The membrane-bound non specific MeUmb-β-glucosidase also hydrolyses the artificial substrate but not the natural substrate. This enzyme differs from the soluble one with regard to its heat-stability at 42° C and its. strong inhibition by Triton X-100. On the other hand, the membrane-bound non specific MeUmb-β-glucosidase differs from the membrane-bound β-glucocerebrosidase in its heat-stability at 50° C, its substrate specificity and the sodium-taurocholate effect. This enzyme activity is about normal in Gaucher disease and is thus coded by a gene different from the β-glucocerebrosidase one.

Transfection with fluorinated lipoplexes based on new fluorinated cationic lipids and in the presence of a bile salt surfactant.

The synthesis of two fluorinated cationic lipids, which are analogues of frequently used synthetic gene carrier agents (including the cationic 2,3-dioleoyloxy-N-[2-(spermine-carboxamido)ethyl]-N,N-dimethyl-1-propanaminium (DOSPA) component of the commercially available liposomal Lipofectamine), and the disintegration and DNA accessibility (evaluated by the ethidium bromide (BET) intercalation assay) as well as the in vitro transfection efficacy of cationic lipoplexes formulated with these new lipids in conjunction with conventional or fluorinated helper lipids, in the absence or presence of sodium taurocholate (STC), a powerful anionic bile salt detergent, is reported. A higher stability, with respect to the STC lytic activity and DNA accessibility, of the fluorinated cationic lipoplexes as compared with their respective lipofectamine-based ones was demonstrated. Indeed, while the Lipofectamine lipoplexes were fully disintegrated at a [STC]/[lipid] molar ratio of 2000, only 40-60% of the DNA intercalation sites of the lipoplexes based on the fluorinated analogue of DOSPA were accessible to ethidium bromide. A higher transfection potential in the presence of STC was further found for the lipoplexes formulated with the fluorinated analogue of DOSPA as compared with the Lipofectamine preparation. For a STC concentration of 7.5 mM, lipofection mediated with these fluorinated lipoplexes was significantly higher (nearly 30- to 50-fold, p < 0.05) than with the Lipofectamine ones. These results confirm the remarkable transfection potential of fluorinated lipoplexes.

Alternate Substrate Inhibition of Cholesterol Esterase by Thieno[2,3-d][1,3]oxazin-4-ones

In a kinetic study, the interaction of bovine pancreatic cholesterol esterase (CEase) with fused 1,3-oxazin-4-ones and 1,3-thiazin-4-ones was investigated, and the compounds were characterized as alternate substrate inhibitors. Inhibition assays were performed in the presence of sodium taurocholate with p-nitrophenyl butyrate as chromogenic substrate. Strong active site-directed inhibition was detected for 2-diethylaminothieno[2,3-d][1,3]oxazin-4-ones with a cycloaliphatic chain at positions 5,6. The most potent inhibitors, compounds 3 and 4, exhibited K(i) values of 0.58 and 1.86 microm, respectively. An exchange of the ring oxygen by sulfur and the removal of the cycloaliphatic moiety as well as the replacement of the thiophene ring by benzene led to a loss of inhibitory potency. CEase has the capability to catalyze the hydrolysis of representatives of fused 1,3-oxazin-4-ones as well as the highly stable 1,3-thiazin-4-ones by using an acylation-deacylation mechanism. Hydrolyses were performed in the presence of a high enzyme concentration, and products were identified spectrophotometrically and by means of high performance liquid chromatography. The kinetic parameters V(max)I and V(max)I/K(m)(I) for the CEase-catalyzed turnover were determined. The compounds whose enzyme-catalyzed hydrolysis followed first-order kinetics (K(m)(I) > 25 microm) failed to inhibit CEase. On the other hand, inhibitors 3 (initial concentration of 25 microm) and 4 (20 microm) were hydrolyzed by CEase under steady-state conditions in the first phase of the reaction. Rate-limiting deacylation was demonstrated in nucleophilic competition experiments with ethanol as acyl acceptor wherein the conversion of compound 3 was accelerated up to an ethanol concentration of 1.5 m. The characterization of these compounds (i.e. 3 and 4) as alternate substrate inhibitors is not only based on the verification of the CEase-catalyzed hydrolysis. It also rests upon the concurrence of corresponding K(i) values obtained in the inhibition assay compared with separately determined K(m)(I) values of their enzyme-catalyzed consumption, as could be predicted from the kinetic model used in this study.

In vitro taurocholate-induced segmentation and clustering of Mesocestoides vogae (syn. corti) tetrathyridia (Cestoda)--inhibition byh cestocidal drugs.

Mesocestoides vogae (syn. M. corti) tetrathyridia were cultured in the presence of sodium taurocholate, for the purpose of exploring the suitability of this organism for the in vitro assay of cestocidal drugs. Parasite clustering and segmentation were observed as taurocholate-dependent effects in biphasic and monophasic media, respectively. Interestingly, representative members of two major classes of known cestocidal agents (namely, albendazole and praziquantel) blocked these effects. Furthermore, it was possible to determine a specific concentration of the drugs that inhibited clustering and segmentation (minimum inhibitory concentration). In contrast, no inhibition was obtained in the presence of anthelmintics without cestocidal activity. These observations open the way for further studies focused at understanding how the activity of the drugs is involved in the suppression of the taurocholate-induced effects.

The induction of SOS function in Escherichia coli K-12/PQ37 by 4-nitroquinoline oxide (4-NQO) and fecapentaenes-12 and -14 is bile salt sensitive: implications for colon carcinogenesis

The response of Escherichia coli to genotoxic agents involves the triggering of a complex system of genes known as the SOS response. In E. coli PQ37, a test organism used for the assessment of genotoxicity, lacZ, the β-galactosidase gene is placed under the control of sfiA, one of the SOS genes through an operon fusion. The induction of β-galactosidase activity, when the organism is exposed to genotoxic agents, is an indirect measure of the genotoxic activity of the test compound. Incubation of E. coli PQ37 with either 4-nitroquinoline oxide (4-NQO) or one of the fecal mutagens, fecapentaene-12 or -14 (F-12 or F-14) in the presence of sodium taurocholate or sodium deoxycholate resulted in a significant enhancement of induction of β-galactosidase activity. The molecular mechanisms of 4-NQO-induced mutagenesis in E. coli are similar to those of the effects of UV light in which both replication-dependent and repair-dependent pathways of mutagenesis exist. Since E. coli PQ37 is excision-repair-deficient, alternate pathways are involved in this system. Bile salts by themselves do not trigger the SOS response, and hence their role in enhancing the SOS-inducing potency of mutagens may involve the potentiation of the cleavage-inactivation of lexA (repressor of SOS) by the protein product of the SOS-controlled gene, recA. The potentiating effect of bile salts on the fecal mutagens, F-12 and F-14, has implications in their suspected role in colon carcinogenesis associated with high-fat, low-fiber diets.

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.

The uptake and esterification of radiolabelled fatty acids by enterocytes isolated from rainbow trout (Oncorhynchus mykiss)

To study the intestinal fatty acid absorption in fish in vitro, enterocytes were isolated from the intestine of rainbow trout and incubated with an equimolar mixture of seven fatty acids [16:0, 18:1n-9, 18:2n-6, 18:3n-3, 20:4n-6, 20:5n-3 and 22:6n-3] in which the component carrying a radioactive label was varied. The fatty acid mixture was presented in the form of micelles formed by sonication with sodium taurocholate. Control studies showed that the presence of sodium taurocholate in the incubation medium caused an immediate 23% increase in the mortality of the cells but the remaining cells were viable. The effect of the bile salt on cellular permeability was evident at longer exposure periods. The proportions of 14C-labelled 20:5n-3 and 22:6n-3 present as monomeric fatty acids in the micellar solutions were higher than those of 16:0, 18:1n-9, 18:2n-6, 18:3n-3 and 20:4n-6. The rates of uptake of 14C-labelled 20:4n-6, 20:5n-3 and 22:6n-3 by enterocytes were significantly lower than those of the other fatty acid substrates over the first minute of incubation but no significant differences in uptake rate between fatty acids were obvious over a 15 min incubation period. Notable differences were observed between substrates in their distribution pattern in enterocyte lipid classes. Although most of the radioactivity from all radiolabelled substrates was recovered in triacylglycerols, the amounts of 14C- labelled 16:0, 20:4n-6, 20:5n-3 and 22:6n-3 recovered in the polar lipid fraction were higher than those of 14C-labelled C18 substrates, particularly after 15 min. Conversely, the initial esterification rates of 18:1n-9, 18:2n-6 and 18:3n-3 into triacylglycerols were significantly higher than those of 20:4n-6, 20:5n-3 and 22:6n-3. It is concluded that isolated enterocytes can be used for the study of the mechanism of intestinal fatty acid absorption in fish.

The enzymatic hydrolysis of 6-acylamino-4-methylumbelliferyl-β- d-glucosides: identification of a novel human acid β-glucosidase

Fluorogenic 6-acylamino-4-methylumbelliferyl-β- d-glucosides were found to be poor substrates for the three known human β-gluco-sidases, i.e., lysosomal and non-lysosomal glucocerebrosidases and cytosolic broad-specificity β-glucosidase. However, homogenates of human tissues and human cell types showed significant enzymatic hydrolysis of 6-ethanoylamino-4-methylumbelliferyl-β- d-glucoside (EMGlc) due to the activity of a hitherto undescribed β-glucosidase, called here EMGlc-ase. It was shown that the isozyme is hardly active towards 4-methylumbelliferyl-β- d-glucoside or glucosylceramide. EMGlc-ase exhibits maximal activity at 4.5 and 5.0 in the absence and presence of sodium taurocholate respectively. It is a soluble lysosomal enzyme with a discrete isoelectric point of about 5.0. EMGlc-ase is not inhibited by conduritol 13-epoxide, is activated by sodium taurocholate and binds strongly to Concanavalin A. This enzyme is not deficient in relation to Gaucher disease.

Estimation of the increase in solubility of drugs as a function of bile salt concentration.

The objective of this study was to develop a model to predict the extent to which bile salts can enhance the solubility of a drug, based on the physicochemical properties of the compound. The ability to predict bile salt solubilization of poorly soluble drugs would be a key component in determining which drugs will exhibit fed vs. fasted differences in drug absorption. A correlation between the logarithm of the octanol/water partition coefficient [log P] of six steroidal compounds and their solubilities in the presence of various concentrations of sodium taurocholate at 37 degrees C, log [SR] = 2.234 + 0.606 log [P] (r2 = 0.987) where SR is the ratio of the stabilization capacity of the bile salt to the solubilization capacity of water for the drug, was used to predict the solubility of the compounds in presence of sodium taurocholate were then measured. The predicted solubilities were within 10% of the experimentally observed solubilities for griseofulvin, cyclosporin A and pentazocine. The model overpredicted the solubility of phenytoin and diazepam in 15 mM sodium taurocholate solution by a factor of 1.33 and 1.62 respectively. The expected increase in solubility as a function of bile salt concentration can be estimated on the basis of the partition coefficient and aqueous solubility of the compound.

Lamb Pregastric Esterase Catalyzed Hydrolysis of 4-Nitrophenyl-Acetate and -Dodecanoate: pH, Temperature and Bile Salt Effects

Two fractions, F2 and F3, eluted by ion exchange chromatography of the commercial extract of lamb pregastric enzyme have esterase activity against 4-nitrophenylacetate, PNPA. Preheat treatment of fraction F3 at pH 7.2, 50°C for 15 min effectively removes its lipase activity against tributyrin while leaving the esterase component relatively unaffected. The esterase components in fractions F2 and F3 and the lipase component in fraction F3 have K m values against PNPA equal to 0.96, 2.1 and 2.7 mM, respectively. When 4-nitrophenyldecanoate, PNPDe, was used as substrate, the maximum activity was reached at its critical micelle concentration, 1.6 μM, for catalysis by all three enzymes. The reactivity is dependent upon pH and p K values of 6.7 and 8.4, 6.7 and 7.5, and 7.3 were determined from the lipase and esterase components in fraction F3, and the fraction F2 esterase, respectively. The dependence upon temperature of the activities of the esterase components against PNPA were determined within the range 25-47.5°C and Arrhenius parameters have been calculated. The presence of sodium taurocholate affected the activity of each enzyme to a small and differing extent.

Isolation and Characterization of Purified Bile Salt Stimulated Human Milk Lipase

A method has been developed for the isolation and purification of bile-salt-stimulated human milk lipase. This method yields up to six times more enzyme than other reported methods and the specific activity is compara ble. The concentration of BSSL recovered from the whole milk was 0.65 percent of the original protein content. The molecular weight of the isolated protein was 120 kDa. During the course of the purification, both protein content and specific activity were monitored and the esterase and lipase activities of the isolated product were characterized in the presence of sodium taurocholate. Five separate isolations were carried out with the introduction of minor varia tions in the procedure, but the catalytic properties of the product remain unchanged.