Metabolism of dextran sulfate sodium by intestinal bacteria in rat cecum is related to induction of colitis.

Ulcerative colitis induced by dextran sulfate sodium (DSS) is one of the most widely used experimental animal models. However, the mechanism responsible for the pathogenesis of the colitis is still unclear. The aim of the present study was to clarify the events occurring after administration of DSS to rats focusing on the relationship between the intestinal bacterial metabolism of DSS and the intestinal mucosal lesions in the model. Within 2 d after DSS administration, severe injury of the cecal mucosa was evident, together with bloody feces and blood in the cecum. However, these lesions were repressed by administration of antibiotics. On the other hand, DSS was found to be metabolized under anaerobic conditions upon incubation with cecal content in vitro, first being desulfated and then undergoing carbohydrate moiety degradation. However, no such metabolic process occurred when cecal content from rats that had been administered antibiotics was employed. These results indicate that the initial step of DSS-induced ulcerative colitis is lesioning of the cecal mucosa, which is related to metabolism of DSS by intestinal bacteria.

Da-Chaihu-Tang alters the pharmacokinetics of nifedipine in rats and a treatment regimen to avoid this.

OBJECTIVES: To investigate the influence of co-administrated Da-Chaihu-Tang (DCT; a traditional Chinese formulation) on the pharmacokinetics of nifedipine, as well as the safe optimal dosing interval to avoid the adverse interactions. METHODS: A single dose of DCT was administered with nifedipine simultaneously, 2 h before, 30 min before or 30 min after nifedipine administration. Pharmacokinetics of nifedipine with or without DCT were compared. The influences of DCT on nifedipine intestinal mucosal and hepatic metabolism were studied by using rat in-vitro everted jejunal sac model and hepatic microsomes. KEY FINDINGS: A simultaneous co-administration of DCT significantly increased the area under concentration-time curve from time zero to infinity (AUC0-inf ) of nifedipine. In-vitro mechanism investigations revealed that DCT inhibited both the intestinal and the hepatic metabolism of nifedipine. Further study on the optimal dosing interval for nifedipine and DCT revealed that administration of DCT 30 min before or after nifedipine did not significantly change the AUC of nifedipine. CONCLUSIONS: The bioavailability of nifedipine is significantly increased by a simultaneous oral co-administration of DCT. This increase is caused by the inhibitory effect of DCT on both the intestinal mucosal and the hepatic metabolism of nifedipine. The dose interval between DCT and nifedipine needs to be set for over 30 min to avoid such drug-drug interactions.

Baicalein 6-O-ß-D-glucopyranuronoside is a main metabolite in the plasma after oral administration of baicalin, a flavone glucuronide of scutellariae radix, to rats.

Baicalin (BG) and its aglycone, baicalein (B) are strong antioxidants that exert various pharmacological actions and show unique metabolic fates in the rat. The aim of the present study was to identify major metabolite(s) besides BG in rat plasma after oral administration of BG or B. The main metabolite was detected by HPLC equipped with an electrochemical detector at a potential of +500 mV and identified as baicalein 6-O-ß-D-glucopyranuronoside (B6G) by HPLC/MS/MS. When BG at a dose of 20 mg/kg was administered orally to Wistar rats, the level of B6G in plasma was higher than that of BG. Cmax and the area under the concentration-curve from 0 to 24 h (AUC0-24 h) values of the plasma B6G were 1.66 ± 0.34 µM and 19.8 3.9 ± µM · h, respectively, whereas those of BG were 0.853 ± 0.065 µM and 10.0 ± 3.1 µM · h, respectively. When B was administered, similar results were also obtained. B6G-producing activities from B were found in microsomes of both rat jejunum and liver, in spite of the low activity. Rat everted jejunal sacs formed B6G after application of B, but only in a small amount that was excreted into the mucosal side, and not the serosal side, indicating little contribution to the appearance of B6G in plasma. On the other hand, when B was injected into the rat portal vein, B6G was detected at a higher level than BG in the systemic circulation, demonstrating the hepatic contribution to the appearance of plasma B6G.

Further evidence on the intramolecular lactonization in rat liver microsomal metabolism of 12-O-acetylated retronecine-type pyrrolizidine alkaloids.

We have previously found evidence of intramolecular lactonization in rat liver microsomal metabolism of isoline, a 12-O-acetylated pyrrolizidine alkaloid. In this study, the metabolism of another 12-O-acetylated pyrrolizidine alkaloid, acetylduciformine, by the proposed transformation pathway was investigated under the same incubation conditions. Two deacetylated metabolites from acetylduciformine were isolated and purified by chromatographic methods, and further characterized based on their physical properties and spectral data. One metabolite (lankongensisine A) was the lactone of another one (duciformine). Both compounds were first obtained as hydrolyzed metabolites from acetylduciformine by rat liver microsomes. More importantly, the present study provided further evidence for the intramolecular lactonization in the microsomal metabolism of 12-O-acetylated retronecine-type PAs.

Hepatic contribution to a marked increase in the plasma concentration of baicalin after oral administration of its aglycone, baicalein, in multidrug resistance-associated protein 2-deficient rat.

Baicalin (BG) and its aglycone, baicalein (B), are strong antioxidants and have various pharmacological actions and show unique metabolic fates in the rat. The aim of the present study was to evaluate the contribution of multidrug resistance-associated protein 2 (Mrp2, Abcc2) to the disposition of BG after oral administration of B using the Eisai hyperbilirubinemic rat (EHBR, Mrp2-deficient), in comparison with the Sprague-Dawley rat (SD, the wild-type form of EHBR). When B at a dose of 12.1 mg/kg was administered orally to EHBRs, the area under the concentration curve from time 0 to 24 h (AUC(0-24 h)) and C(max) values of plasma BG, 27.6 + or - 3.5 microM x h and 11.4 + or - 3.9 microM, respectively, were significantly higher at 5-fold and 8-fold, respectively, than those (AUC(0-24 h) value of 5.39 + or - 1.37 microM x h and C(max) value of 1.51 + or - 0.38 microM) in SD rats. In addition, when B at a dose of 1.2 mg/kg was injected into the portal vein of EHBRs, a marked reduction in the biliary excretion of BG (6.59 + or - 5.64 nmol) and a marked increase of BG in the systemic circulation (AUC(0-120 min) of 75.4 + or - 34.5 microM x min) were observed, in comparison with those (biliary excretion of 29.7 + or - 9.3 nmol and AUC(0-120 min) value of 2.42 + or - 1.44 microM x min) in SD rats. Thus, it was clarified that after oral administration of B, the markedly higher concentration of plasma BG in EHBRs than in SD rats was caused mainly by a drastic reduction in the biliary excretion of BG and marked enhancement of its sinusoidal efflux from the liver.

Involvement of the CYP1A subfamily in stereoselective metabolism of carvedilol in beta-naphthoflavone-treated Caco-2 cells.

We have previously reported that the metabolism of S-carvedilol in beta-naphthoflavone (beta-NF)-treated Caco-2 cells is faster than that of R-carvedilol. The aim of the present study was to identify the enzyme responsible for the stereoselective metabolism of carvedilol in the cells. The expression of cytochrome P450 (CYP) 1A1 and CYP1A2 mRNA, but not CYP2D6, CYP3A4, and CYP2C9 mRNA, was increased in beta-NF-treated Caco-2 cells, as compared with non-treated cells. Furafylline, an inhibitor of the CYP1A subfamily, decreased the metabolism of S-carvedilol in Caco-2 cells cultured on plastic dishes. In addition, the glucuronidation of carvedilol was not significant in microsomes of beta-NF-treated Caco-2 cells. On the other hand, the oxidation of S-carvedilol in microsomes of beta-NF-treated Caco-2 cells was faster than that of R-carvedilol, and furafylline decreased the oxidative activity of S-carvedilol. These findings suggested that the CYP1A subfamily was responsible for the stereoselective metabolism of carvedilol in beta-NF-treated Caco-2 cells.

Enantioselective dehydroxylation of enterodiol and enterolactone precursors by human intestinal bacteria.

During the course of experiments on the transformation of lignans to phytoestrogenic substances, such as enterodiol (END) and enterolactone (ENL), a previously isolated bacterium, Eubacterium (E.) sp. strain SDG-2, capable of phenolic p-dehydroxylation in the biotransformation of secoisolariciresinol diglucoside to END and ENL, was concluded to be Eggerthella (Eg.) lenta (Eg. sp. SDG-2) on the basis of 16S rRNA gene sequence analysis. The bacterium could transform (+)-dihydroxyenterodiol (DHEND, 3a) to (+)-END (1a), but not for (-)-DHEND (3b) to (-)-END (1b) under anaerobic conditions. By incubation of a mixture of (+)- and (-)-dihydroxyenterolactone (DHENL, 4a and 4b) with Eg. sp. SDG-2, only (-)-DHENL (4b) was converted to (-)-ENL (2b), selectively. On the other hand, we isolated a different bacterium, strain ARC-1, capable of dehydroxylating (-)-DHEND (3b) to (-)-END (1b) from human feces. Strain ARC-1 could transform not only (-)-DHEND (3b) to (-)-END (1b), but also (+)-DHENL (4a) to (+)-ENL (2b). However, the bacterium could not transform (+)-DHEND (3a) and (-)-DHENL (4b). Both bacterial strains demonstrated different enantioselective dehydroxylation.

Antispasmodic activity of licochalcone A, a species-specific ingredient of Glycyrrhiza inflata roots.

Licochalcone A, a species-specific and characteristic retrochalcone ingredient of Glycyrrhiza inflata root, has been shown to possess multiple bioactive properties. However, its muscle relaxant activity has not been reported previously. Licochalcone A showed a concentration-dependent relaxant effect on the contraction induced by carbachol (50% effective concentration (EC50) = 5.64 +/- 1.61 microM), KCl (EC50 5.12 +/- 1.68 microM), BaCl2 (EC50 1.97 +/- 0.48 microM) and A23187 (EC50 2.63 +/- 2.05 microM). Pretreatment with licochalcone A enhanced the relaxant effect of forskolin, an adenylyl cyclase activator, on the contraction in a similar manner to 3-isobutyl-1-methylxanthine (IBMX), a phosphodiesterase (PDE) inhibitor. Furthermore, the IC50 (22.1 +/- 10.9 microM) of licochalcone A against cAMP PDE was similar to that of IBMX (26.2 +/- 7.4 microM). These results indicated that licochalcone A may have been responsible for the relaxant activity of G. inflata root and acted through the inhibition of cAMP PDE.

In vitro metabolism of isoline, a pyrrolizidine alkaloid from Ligularia duciformis, by rodent liver microsomal esterase and enhanced hepatotoxicity by esterase inhibitors.

Isoline, a major retronecine-type pyrrolizidine alkaloid (PA) from the Chinese medicinal herb Ligularia duciformis, was suggested to be the most toxic known PA. Its in vitro metabolism was thus examined in rat and mouse liver microsomes, and its toxicity was compared with that of clivorine and monocrotaline after i.p. injection in mice. Isoline was more rapidly metabolized by both microsomes than clivorine and monocrotaline and converted to two polar metabolites M1 and M2, which were spectroscopically determined to be bisline (a deacetylated metabolite of isoline) and bisline lactone, respectively. Both metabolites were formed in the presence or absence of an NADPH-generating system with liver microsomes but not cytosol. Their formation was completely inhibited by the esterase inhibitors, triorthocresyl phosphate (TOCP) and phenylmethylsulfonyl fluoride, but not at all or partially by cytochrome P450 (P450) inhibitors, alpha-naphthoflavone and proadifen (SKF 525A), respectively. These results demonstrated that both metabolites were produced by microsomal esterase(s) but not P450 isozymes. The esterase(s) involved showed not only quite different activities but also responses to different inhibitors in rat and mouse liver microsomes, suggesting that different key isozyme(s) or combinations might be responsible for the deacetylation of isoline. Isoline injected i.p. into mice induced liver-specific toxicity that was much greater than that with either clivorine or monocrotaline, as judged by histopathology as well as serum alanine aminotransferase and aspartate aminotransferase levels. Isoline-induced hepatotoxicity was remarkably enhanced by the esterase inhibitor TOCP but was reduced by the P450 inhibitor SKF 525A, indicating that rodent hepatic esterase(s) played a principal role in the detoxification of isoline via rapid deacetylation in vivo.

Isolation and characterization of a human intestinal bacterium, Eubacterium sp. ARC-2, capable of demethylating arctigenin, in the essential metabolic process to enterolactone.

Plant lignans, such as pinoresinol diglucoside, secoisolariciresinol diglucoside and arctiin, are metabolized to mammalian lignans, enterolactone or enterodiol, by human intestinal bacteria. Their metabolic processes include deglucosylation, ring cleavage, demethylation, dehydroxylation and oxidation. Here we isolated an intestinal bacterium capable of demethylating arctigenin, an aglycone of arctiin, to 2,3-bis(3,4-dihydroxybenzyl)butyrolactone (1) from human feces, and identified as an Eubacterium species (E. sp. ARC-2), which is similar to Eubacterium limosum on the basis of morphological and biochemical properties and 16S rRNA gene sequencing. By incubating with E. sp. ARC-2, arctigenin was converted to 1 through stepwise demethylation. Demethylation of arctigenin by E. sp. ARC-2 was tetrahydrofolate- and ATP-dependent, indicating that the reaction was catalyzed by methyltransferase. Moreover, E. sp. ARC-2 transformed secoisolariciresinol to 2,3-bis(3,4-dihydroxybenzyl)-1,4-butanediol by demethylation.

Isoliquiritigenin, one of the antispasmodic principles of Glycyrrhiza ularensis roots, acts in the lower part of intestine.

Glycyrrhizae radix is used to treat abdominal pain as a component of shakuyakukanzoto (shaoyao-gancao-tang), a traditional Chinese medicine formulation. Previously, we have reported the isolation of glycycoumarin as a potent antispasmodic with an IC50 value of 2.93+/-0.94 microM for carbamylcholine (CCh)-induced contraction of mouse jejunum from an aqueous extract of Glycyrrhizae radix (licorice), and clarified that its mechanism of action involves inhibition of phosphodiesterase 3. The purpose of the present study was to examine an antispasmodic principle of licorice other than glycycoumarin. Isoliquiritigenin was isolated from an aqueous extract of licorice as a potent relaxant, which inhibited the contraction induced by various types of stimulants, such as CCh, KCl, and BaCl2 with IC50 values of 4.96+/-1.97 microM, 4.03+/-1.34 microM and 3.70+/-0.58 microM, respectively, which are close to those of papaverine. However, the amount of isoliquiritigenin in the aqueous extract of licorice was very small. When the aqueous licorice extract was treated with naringinase, the amounts of glycosides such as isoliquiritin, which were abundant but had much less potent relaxant activity, were decreased while isoliquiritigenin was increased. At the time, the relaxant activity of the treated sample was increased significantly, shifting the IC50 from 358+/-104 to 150+/-38 microg/ml for CCh-induced contraction. Isoliquiritigenin also showed the most potent inhibition of mouse rectal contraction induced by CCh with an IC50 value of 1.70+/-0.07 microM. These results suggest that isoliquiritigenin acts as a potent relaxant in the lower part of the intestine by transformation from its glycosides.

Efflux of baicalin, a flavone glucuronide of Scutellariae Radix, on Caco-2 cells through multidrug resistance-associated protein 2.

Baicalin and its aglycone, baicalein, being are strong antioxidants and have various pharmacological actions. Baicalein has shown a unique metabolic fate in rat intestine, being excreted into the intestinal lumen from mucosal (epithelial) cells following glucuronidation of baicalein absorbed after oral administration. The purpose of this study was to examine the absorption and excretion of baicalin and baicalein in a Caco-2 cell monolayer model to evaluate the disposition of baicalin and baicalein in the human intestine. When baicalein at 5 microM was loaded on the apical side of the Caco-2 cell monolayer, baicalein was not transferred to the basolateral side, but more baicalin was excreted onto the apical side than was being absorbed onto the basolateral side. The amount of baicalin recovered on both sides accounted for more than 90% of the baicalein absorbed from the apical surface. This was supported by the fact that Caco-2 cell microsomes showed UDP-glucuronate glucuronosyltransferase activity towards baicalein to form baicalin. On the other hand, when baicalein was loaded at higher concentrations, baicalin excretion became saturated, and then baicalein was transferred to the basolateral side. Furthermore, baicalin efflux was not inhibited by MDR1/P-glycoprotein substrates such as ciclosporin and vinblastine, but significantly inhibited by multidrug resistance-associated protein 2 (MRP2, ABCC2) substrates such as probenecid and genistein. MRP2 was also detected in Caco-2 cells by Western blotting using specific antibodies. In addition, baicalin, but not baicalein, enhanced dose-dependently the vanadate-sensitive ATPase activity of human MRP2. These results indicated that, in Caco-2 cells, any baicalein absorbed after loading at low concentrations of baicalein was not transferred to the basolateral side, but was first transformed into baicalin in the cells and excreted through the action of MRP2, mainly to the apical side.

Interaction between Shaoyao-Gancao-Tang and a laxative with respect to alteration of paeoniflorin metabolism by intestinal bacteria in rats.

Shaoyao-Gancao-Tang (SGT), a traditional Chinese herbal medicine (Kampo formulation) containing Shaoyao (Paeoniae Radix) and Gancao (Glycyrrhizae Radix), is co-administered with laxative sodium picosulfate as a premedication for relieving the pain accompanying colonoscopy. Paeoniflorin (PF), an active glycoside of SGT, is metabolized into the antispasmodic agent paeonimetabolin-I (PM-I) by intestinal bacteria after oral administration. The objective of the present study was to investigate whether the co-administered laxative (sodium picosulfate) influences the metabolism of PF to PM-I by intestinal bacteria. We found that the PF-metabolizing activity of intestinal bacteria in rat feces was significantly reduced to approximately 34% of initial levels by a single sodium picosulfate pretreatment and took approximately 6 days to recover. Repeated administration of SGT after the sodium picosulfate pretreatment significantly shortened the recovery period to around 2 days. Similar results were also observed for plasma PM-I concentration. Since PM-I has muscle relaxant activity, the present results suggest that repetitive administration of SGT after sodium picosulfate pretreatment might be useful to relieve the pain associated with colonoscopy.

Pharmaceutical evaluation of cultivated Glycyrrhiza uralensis roots in comparison of their antispasmodic activity and glycycoumarin contents with those of licorice.

In China, the collection of wild Glycyrrhiza uralensis, one of the raw materials of Chinese licorice, has been restricted to prevent desertification. To compensate for the reduced supply of wild Glycyrrhiza plants, cultivation programs of G. uralensis have been initiated in eastern Inner Mongolia. The goal of the present study was to compare the chemical and pharmacological properties of cultivated G. uralensis roots to those of licorice prepared from wild Glycyrrhiza plants. The antispasmodic effect of boiled water extract of 4-year-old cultivated G. uralensis roots and licorice on carbachol-induced contraction in mice jejunum was similar (ED(50): 134+/-21 microg/ml vs. 134+/-16 microg/ml). In addition, glycycoumarin content, which is an antispasmodic and species-specific ingredient of G. uralensis, was similar when comparing the boiled water extracts of 4-year-old cultivated roots and licorice (0.10+/-0.02% vs. 0.10+/-0.06%). These data suggest that cultivated G. uralensis roots may be an adequate replacement for the generation of licorice in the context of the restriction of wild Glycyrrhiza plant collection.

Pharmacokinetics of berberine and its main metabolites in conventional and pseudo germ-free rats determined by liquid chromatography/ion trap mass spectrometry.

Berberine (Ber) and its main metabolites were identified and quantified using liquid chromatography/electrospray ionization/ion trap mass spectrometry. Rat plasma contained the main metabolites, berberrubine, thalifendine, demethyleneberberine, and jatrorrhizine, as free and glucuronide conjugates after p.o. Ber administration. Moreover, the original drug, the four main metabolites, and their glucuronide conjugates were all detected in liver tissues after 0.5 h and in bile samples 1 h after p.o. Ber administration. Therefore, the metabolic site seemed to be the liver, and the metabolites and conjugates were evidently excreted into the duodenum as bile. The pharmacokinetics of Ber and the four metabolites were determined in conventional and pseudo germ-free rats (treated with antibiotics) after p.o. administration with 40 mg/kg Ber. The AUC0-limt and mean transit time values of the metabolites significantly differed between conventional and pseudo germ-free rats. The amounts of metabolites were remarkably reduced in the pseudo germ-free rats, whereas levels of Ber did not obviously differ between the two groups. The intestinal flora did not exert significant metabolic activity against Ber and its metabolites, but it played a significant role in the enterohepatic circulation of metabolites. In this sense, the liver and intestinal bacteria participate in the metabolism and disposition of Ber in vivo.

Glycycoumarin from Glycyrrhizae Radix acts as a potent antispasmodic through inhibition of phosphodiesterase 3.

Glycyrrhizae Radix is used to treat abdominal pain as a component of Shakuyaku-kanzo-to, a traditional Chinese medicine formulation. We aim at clarifying the antispasmodic principles of Glycyrrhizae Radix, and consequently isolated glycycoumarin as a potent relaxant on the carbamylcholine (CCh)-induced contraction of mouse jejunum. In this paper we investigated the effects and the action mechanism of glycycoumarin on the contraction of mouse jejunum. Glycycoumarin inhibited the contraction induced by various types of stimulants, such as CCh, KCl, BaCl(2), and A23187 (calcium ionophore III) with IC(50) values of 2.93+/-0.94 micromol/l (1.08+/-0.35 microg/ml), 2.59+/-0.58 micromol/l (0.95+/-0.29 microg/ml), 4.09+/-1.82 micromol/l (1.51+/-0.67 microg/ml) and 7.39+/-5.19 micromol/l (2.72+/-1.91 microg/ml), respectively, with a potency similar to that of papaverine (a representative antispasmodic for smooth muscle). Furthermore, pretreatment with glycycoumarin enhanced the relaxation induced by forskolin on CCh-evoked contraction, similar to that by pretreatment with IBMX, a non-specific inhibitor of phosphodiesterases (PDEs). Pretreatment with glycycoumarin also enhanced the relaxation effect of rolipram, a specific inhibitor of PDE isozyme 4, as pretreatment with milrinone, a specific inhibitor of isozyme 3, did. Moreover, the effect of glycycoumarin was associated with dose-dependent accumulation of cAMP, but not cGMP, in mouse jejunum. These results indicate that glycycoumarin has an inhibitory effect on smooth muscle contraction induced by various types of stimulants through the inhibition of PDEs, especially isozyme 3, followed by the accumulation of intracellular cAMP.

Two proteins, Mn2+, and low molecular cofactor are required for C-glucosyl-cleavage of mangiferin.

C-Glucosides, in which sugars are attached to the aglycone by carbon-carbon bonds, are generally resistant to acid and enzyme hydrolysis. The C-glucosyl bond of mangiferin, a xanthone C-glucoside, was cleaved by anaerobic incubation with a human intestinal bacterium, Bacteroides sp. MANG, to give norathyriol. A cell-free extract obtained by sonication of B. sp. MANG demonstrated cleaving activity for mangiferin to norathyriol by adding NADH, diaphorase, and dithiothreitol. Both high molecular weight (>10 k) and low molecular weight (<10 k) fractions obtained from the cell-free extract were required for the activity. MnCl2 was necessary for the activity, but other metal ions were not. By purification of the high molecular weight fraction using DEAE-cellulose and Phenyl Sepharose column chromatography, two fractions, designated as proteins A and B, were separated and required for the activity. Neither protein A nor protein B alone showed any activity. This is the first report describing a C-glucosyl-cleaving enzyme from human intestinal bacterium that seems to involve a novel enzyme mechanism.

Bioavailability of glycyrrhizin from Shaoyao-Gancao-Tang in laxative-treated rats.

Shaoyao-Gancao-Tang (SGT), a traditional Chinese formulation composed of Shaoyao (Paeoniae Radix) and Gancao (Glycyrrhizae Radix), is frequently used in conjunction with laxatives such as sodium picosulfate in colonoscopy to relieve abdominal pains. We have investigated the alterations of the bioavailability of glycyrrhizin when SGT was co-administered with sodium picosulfate and we tried to identify a regimen that might minimize the alterations. Glycyrrhizin is one of the active glycosides in Gancao and SGT and is hydrolysed into the bioactive metabolite, 18 beta-glycyrrhetic acid (GA) by intestinal bacteria following oral administration. We found that the maximum plasma concentration (C(max)) and the area under the mean concentration vs time curve from zero to 24 h (AUC(0-24 h)) of GA from a single dose of SGT administered 5 h after a single pretreatment with sodium picosulfate were significantly reduced to 15% and 20% of the control level in rats, respectively. These reductions were still significant four days after sodium picosulfate pretreatment, but were restored by repetitive administration of SGT following sodium picosulfate pretreatment. Similar reductions and recovery were observed for the glycyrrhizin-metabolizing activity of intestinal bacteria in rat faeces. The results warrant clinical studies for co-administration of laxatives such as sodium picosulfate and SGT.

Isolation of a human intestinal bacterium that transforms mangiferin to norathyriol and inducibility of the enzyme that cleaves a C-glucosyl bond.

The C-glucosyl bond of C-glucosides generally tolerates acid and enzymatic hydrolysis. Many C-glucosides are cleaved by human intestinal bacteria. We isolated the specific bacterium involved in the metabolism of mangiferin (2-beta-D-glucopyranosyl-1,3,6,7-tetrahydroxyxanthone), C-glucosyl xanthone, from a mixture of human fecal bacteria. The anaerobic Bacteroides species named MANG, transformed mangiferin to the aglycone, norathyriol, suggesting cleavage of a C-glucosyl bond. However, B. sp. MANG cleaved C-glucosyl in a dose- and time-dependent manner only when cultivated in the presence of mangiferin. Cleavage was abolished by inhibitors of RNA and protein syntheses, such as rifampicin and chloramphenicol, respectively, indicating that the enzyme that cleaves C-glucosyl is induced by mangiferin. In contrast, mangiferin did not affect bacterial alpha- and beta-glucosidase activities under any conditions. The C-glucosyl-cleavage in cell-free extracts was not altered by potent glucosidase inhibitors such as 1-deoxynojirimycin and gluconolactone. Therefore, the C-glucosyl-cleaving enzyme substantially differs from known glucosidases that cleave O-glucosides. This is the first description of a specific intestinal bacterium that is involved in the metabolism of mangiferin and which produces a novel and inducible C-glucosyl-cleaving enzyme.

Enteric excretion of baicalein, a flavone of Scutellariae Radix, via glucuronidation in rat: involvement of multidrug resistance-associated protein 2.

PURPOSE: Baicalin (BG) and its aglycone, baicalein (B), are strong antioxidants and have various pharmacological actions. The purpose of this study was to evaluate efflux of BG from rat intestinal mucosal cell following glucuronidation of B absorbed after oral administration of B. METHODS: The absorption and excretion of BG and B were evaluated in rats using the in situ jejunal loop technique and in vitro jejunal everted sac experiments. BG and B levels were determined by high-performance liquid chromatography with electro-chemical detection to ensure selectivity and high sensitivity. RESULTS: A large amount (30.4% recovery) of BG, but no B, was detected in the intestinal lumens of germ-free rats 4 h after oral administration of B (12.1 mg/kg), in comparison with a substantial recovery (55.1%) of unabsorbed BG 4 h after its administration. During the in situ rat jejunal loop absorption experiment, B disappeared rapidly, and 8% of the lost B was excreted into the loop as BG 20 min after infusing 0.1 mM B. In an in vitro absorption experiment using everted rat jejunal sac, BG also appeared outside the sac, accompanied by the disappearance of B from the outer (mucosal) side. However, very little of B was transferred to the inner (serosal) side of the sac, and only a trace of BG was detected inside the sac. Thus, in both the loop and the everted sac systems, the efflux of BG from the mucosal surface was saturated with the concentration of B added. Moreover, the efflux rate of BG in the everted jejunal sac from Eisai hyperbilirubinemic rat (EHBR) was significantly lower by 56.4% than that from Sprague-Dawley rat. CONCLUSIONS: These results indicate that, in rat, a large proportion of any B absorbed is retained, transformed into BG within the intestinal mucosal cells, and coordinately excreted through multidrug resistance-associated protein 2 (MRP2) into the intestinal lumen.