Rat Bile Research Articles

Metabolite profiling and identification of triptolide in rats

The purpose of the current study was to investigate the metabolite profile of [(3)H]triptolide in rats. The separation and characterisation techniques used to identify the major metabolites were high-performance liquid chromatography-online radiodetector, ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry, and nuclear magnetic resonance. In all, 33 major metabolites were detected. The major components found in the rat plasma included the parent drug and its monohydroxy- and dihydroxy-metabolites. Reference standards for the monohydroxy-metabolites were obtained either by the incubation of the parent drug with rat liver microsomes or by microbial transformation with Cunninghamella blakesleana. The metabolites' structures were identified as 17-hydroxytriptolide, 16-hydroxytriptolide, tripdiolide, and 15-hydroxytriptolide. The major metabolites found in male rat urine included the monohydroxy-, dihydroxy-, and trihydroxy-metabolites. The major metabolites in female rat urine were the monohydroxy- and dihydroxy-metabolites, as well as sulphates of the monohydroxy-metabolites. A glutathione adduct, multiple hydroxy-metabolites, and a number of unidentified metabolites were detected in the bile and faeces of male rats. Sulphates of monohydroxy-metabolites were detected in the bile and faeces of female rats.

A validated HPLC‐MS/MS method for determination of genipin‐1‐o‐glucuronic acid in rat plasma after administration of genipin and its application to a pharmacokinetic study

A sensitive and specific method was developed and validated for the quantitation of one major metabolite of genipin in rats plasma. The major metabolite was isolated from rat bile via semi-preparative HPLC technology and its chemical structure was identified as genipin-1-o-glucuronic acid (GNP-GLU), which was for the first time used as a standard compound for quantitative analysis in rat plasma after administration of genipin. The application of high-performance liquid chromatography-tandem mass spectrometry in negative mode in multiple reaction monitoring mode was investigated. Chromatographic separation was achieved on an Eclipse XDB-C18 column using a mobile phase consisting of water with 0.1% formic acid (A)-acetonitrile (B). The limit of detecation was 0.214 ng/mL and the lower limit of quantification was 0.706 ng/mL. The calibration curve was linear from 1.27 to 3810 ng/mL for plasma samples, with a correlation coefficient of 0.9924. The intra- and inter-day precisions and accuracy were all within 15%. The recoveries of GNP-GLU and puerarin were above 90.0 and 76.2%, respectively. The highly sensitive method was successfully applied to estimate pharmacokinetic parameters of GNP-GLU following oral and intravenous administration of genipin to rats.

The identification and pharmacokinetic studies of metabolites of salvianolic acid B after intravenous administration in rats

AimTo identify and quantify the major metabolites of salvianolic acid B (SAB) after intravenous injection in rats. MethodsLC-IT/TOF-MS was used to identify the metabolites in rat bile, plasma, and urine; LC-MS/MS was used to quantify the two major metabolites. ResultsIn rat bile, plasma, and urine, nine metabolites were identified, including methylated metabolites of SAB, lithospermic acid (LSA), the decarboxylation and methylation metabolites of LSA, salvianolic acid S (SAS), and dehydrated-SAS. The t1/2 of monomethyl-SAB and LSA were both very short, and monomethyl-SAB had a larger AUC than LSA in rats. ConclusionNine metabolites were found, the metabolic pathway was described, and the pharmacokinetic profiles of LSA and monomethyl-SAB were studied, thereby clarifying that methylation was the dominant metabolic pathway for SAB in rats.

Development of an HPLC fluorescence method for determination of boldine in plasma, bile and urine of rats and identification of its major metabolites by LC–MS/MS

Boldine belongs to the group of aporphine alkaloids isolated from Boldo tree. In contrast with numerous reports on the pharmacological effects of boldine, the data about its pharmacokinetics and biotransformation are scarce. No validated bioanalytical method of sufficient sensitivity has so far been described in the literature which could be used for quantification of boldine in various body fluids collected in pharmacokinetic studies. This work presents, for the first time, the assay for boldine in the plasma, bile and urine of rats. It includes liquid-liquid extraction/back-extraction of boldine, its chromatographic separation and sensitive fluorescence detection. Separation was carried out on a pentafluorophenyl core-shell column (Kinetex PFP, 150×3mm, 2.6μm) in gradient elution mode with solvent system consisting of an acetonitrile-ammonium formate buffer (5mM, pH=3.8). Fluorimetric detection (λEX=320nm, λEM=370nm) was used for quantitative work. Validation according to the EMEA guideline proved the assay LLOQ (0.1μmolL(-1)), linearity over a broad range of 0.1-50μmolL(-1), precision (intra- and inter-day CVs less than 4.5% and 6.1%, respectively) and accuracy (relative errors between -5.8% and 4.8%). In a pilot pharmacokinetic experiment, the concentration-time profiles were described for boldine (single i.v. bolus 50mgkg(-1)) in plasma and bile and cumulative excretion in urine was investigated. The major metabolites identified by means of LC-MS(n) were boldine-O-glucuronide, boldine-O-sulphate and disulphate, boldine-O-glucuronide-O-sulphate and N-demethyl-boldine-O-sulphate.

Identification of metabolites of Radix Paeoniae Alba extract in rat bile, plasma and urine by ultra-performance liquid chromatography–quadrupole time-of-flight mass spectrometry

Ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UPLC–Q-TOF/MS) was developed to identify the absorbed parent components and metabolites in rat bile, plasma and urine after oral administration of Radix Paeoniae Alba extract (RPAE). A total of 65 compounds were detected in rat bile, plasma and urine samples, including 11 parent compounds and 54 metabolites. The results indicated that glucuronidation, hydroxylation and methylation were the major metabolic pathways of the components of RPAE. Furthermore, the results of this work demonstrated that UPLC–Q-TOF/MS combined with MetaboLynx™ software and mass defect filtering (MDF) could provide unique high throughput capabilities for drug metabolism study, with excellent MS mass accuracy and enhanced MSE data acquisition. With the MSE technique, both precursor and fragment mass spectra can be simultaneously acquired by alternating between high and low collision energy during a single chromatographic run.

Simultaneous and sensitive determination of eight coumarins in rat bile and urine after oral administration ofRadix Angelicae Dahuricaeextract by liquid chromatography-electrospray ionization-mass spectrometry

A selective and sensitive liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) method was developed and validated for analysis of xanthotoxol (1), xanthotoxin (2), isoimpinellin (3), bergapten (4), oxypeucedanin (5), imperatorin (6), cnidilin (7), and isoimperatorin (8) in rat bile and urine using pimpinellin as an internal standard (IS). An Agilent 1200 liquid chromatography system (Agilent Technologies, USA) equipped with a quaternary pump, an autosampler, and a column compartment was used for all analyses. Chromatographic separations were performed on a Sapphire C18 column (150 mm × 4.6 mm, 5 µm), and the column temperature was maintained at 30°C; the sample injection volume was 10 µL. The specificity, linearity, accuracy, precision, recovery, matrix effect, and several stabilities were validated for all analytes in the rat bile and urine samples. The method was successfully applied in monitoring the concentrations of eight coumarins in rat bile and urine after a single oral administration of Radix Angelicae Dahuricae extract with a dosage of 8.0 mL/kg. In the bile samples, the eight coumarins excreted completely in twenty-four hours. The average percentages of coumarins (1-8) excreted were 0.045%, 0.019%, 0.177%, 0.105%, 0.337%, 0.023%, 0.024%, 0.021%. In the urine samples, the eight coumarins excreted completely in seventy-two hours. The average percentages of coumarins (1-8) excreted were 1.78%, 0.095%, 0.130%, 0.292%, 0.082%, 0.008%, 0.005%, 0.004%. The method is robust and specific and it can successfully complete the requirements of the excretion study of the eight coumarins in Radix Angelicae Dahuricae.

Structural identification of the metabolites of ganoderic acid B from Ganoderma lucidum in rats based on liquid chromatography coupled with electrospray ionization hybrid ion trap and time‐of‐flight mass spectrometry

Ganoderic acid B (GAB), a representative triterpenoid in Ganoderma lucidum, possesses various pharmaceutical effects and has been used as a chemical marker in quality control of G. lucidum and related products. The metabolites of GAB in vivo after its oral administration to rats were investigated by liquid chromatography coupled with electrospray ionization hybrid ion trap and time-of-flight mass spectrometry. A total of 14 metabolites of GAB in rat plasma, bile and various organs were detected and identified by direct comparison with the authentic compounds and their characteristic mass fragmentation patterns. The results showed that oxidization and hydroxylation were the common metabolic pathways for GAB in rats. Moreover, some reduction metabolites of GAB were detected in rat kidney and stomach and glucuronidation only appeared in rat bile. This is the first report on the metabolites of GAB in vivo.

Identification of senkyunolide I metabolites in rats using ultra performance liquid chromatography/quadrupole-time-of-flight tandem mass spectrometry

Ligusticum chuanxiong Hort. (Umbelliferae) has been widely prescribed to treat cardiovascular disease in China for centuries. Senkyunolide I is one of the major bioactive components in L. chuanxiong, which shows pharmacological activities against migraines and oxidative damage. In this paper, ultra performance liquid chromatography/quadrupole-time-of-flight tandem mass spectrometry (UPLC/Q-TOF-MS) was applied for the rapid analysis of senkyunolide I metabolites in rats after its intravenous administration. The non-metabolized parent compound and eighteen metabolites from drug-treated samples in rat plasma, urine and bile were identified. Our analysis indicated that methylation, hydration, epoxidation, glucuronidation and glutathione conjugation were the major pathways of senkyunolide I metabolism in vivo. This study provides important information regarding the metabolism of senkyunolide I, which will be helpful for understanding its mechanism of action. Furthermore, this work demonstrates the potential of using UPLC/Q-TOF-MS for the rapid and reliable characterization of the metabolites of natural products.

Propitious role of bone marrow-derived mononuclear cells in an experimental bile duct ligation model: potential clinical implications in obstructive cholangiopathy

To evaluate the role of bone marrow-derived mononuclear cells (BMC) in rat bile duct ligation (BDL) model. Wistar rats were categorized into four Groups A-D. Normal liver biopsy was taken from Group A. BDL model was created in Groups B and C (15 each). Normal saline and BMC were injected through portal vein (PV) in Groups B and C, respectively. In Group D (healthy rat), only BMC were infused through PV. Groups B and C were compared for body weight, liver functions, survival, and histopathological changes. Serum bilirubin was lower in Group C at day 6 (p=0.0010). Median survival time was 5 (4, 6) and 13 (9, 17) days in Groups B and C (p=0.0147), respectively. Portal edema (p=0.013) and portal inflammation (p=0.025) were less in Group C vs Group B. On post hoc subgroup analysis of rats surviving 8-26days, portal inflammation (p=0.004), bile duct proliferation (p=0.016) and portal fibrosis (p=0.038) were less in Group C vs Group B. Hepatocyte regeneration was found in four rats in Group C. CD34-positive cells were prominent in sinusoids and portal tracts in the BDL rat model. BMC have shown to delay fibrosis, facilitate hepatocyte regeneration and improve survival in an experimental BDL model, with potential clinical implication in obstructive cholangiopathy.

An integrated strategy for in vivo metabolite profiling using high-resolution mass spectrometry based data processing techniques

An ongoing challenge of drug metabolite profiling is to detect and identify unknown or low-level metabolites in complex biological matrices. Here we present a generic strategy for metabolite detection using multiple accurate-mass-based data processing tools via the analysis of rat samples of two model drug candidates, AZD6280 and AZ12488024. First, the function of isotopic pattern recognition was proved to be highly effective in the detection of metabolites derived from [14C]-AZD6280 that possesses a distinct isotopic pattern. The metabolites revealed using this approach were in excellent qualitative correlation to those observed in radiochromatograms. Second, the effectiveness of accurate mass based untargeted data mining tools such as background subtraction, mass defect filtering, or a data mining package (MZmine) used for metabolomic analysis in detection of metabolites of [14C]-AZ12488024 in rat urine, feces, bile and plasma samples was examined and a total of 33 metabolites of AZ12488024 were detected. Among them, at least 16 metabolites were only detected by the aid of the data mining packages and not via radiochromatograms. New metabolic pathways such as S-oxidation and thiomethylation reactions occurring on the thiazole ring were proposed based on the processed data. The results of these experiments also demonstrated that accurate mass-based mass defect filtering (MDF) and data mining techniques used in metabolomics are complementary and can be valuable tools for delineating low-level metabolites in complex matrices. Furthermore, the application of distinct multiple data-mining algorithms in parallel, or in tandem, can be effective for rapidly profiling in vivo drug metabolites.

Metabolite Profiles of Epimedin B in Rats by Ultraperformance Liquid Chromatography/Quadrupole-Time-of-Flight Mass Spectrometry

In this work, the metabolite profiles of epimedin B in rat feces, bile, urine, and plasma were qualitatively investigated, and the possible metabolic pathways of epimedin B were subsequently proposed. After oral administration of epimedin B at a single dose of 80 mg/kg, rat biological samples were collected and pretreated by protein precipitation. Then, these pretreated samples were injected into an Acquity ultraperformance liquid chromatography BEH C₁₈ column with mobile phase consisting of 0.1% formic acid-water and 0.1% formic acid-acetonitrile and detected by ultraperformance liquid chromatography/quadrupole-time-of-flight mass spectrometry. In all, 43 metabolites were identified in the biosamples. Of these, 13, including F5, F7, F16-F18, D5-D7, D9, N5, N7, M1, and M3, were to our knowledge reported for the first time. The results indicated that epimedin B was metabolized via desugarization, dehydrogenation, hydrogenation, hydroxylation, demethylation, glucuronidation, and glycosylation pathways in vivo. Specific hydrolysis of 7-O-glucosides in the gut lumen and glucuronic acid conjugation in the liver were considered as the main physiologic processes of epimedin B. This study revealed the possible metabolite profiles of epimedin B in rats.

Omeprazole inhibits biliary proliferation during extrahepatic cholestasis

Omeprazole (OME) has been shown to reduce liver regeneration and enhance gastrin serum levels. Since gastrin inhibits biliary growth, we tested the hypothesis that omeprazole inhibits cholangiocyte proliferation in BDL rats by increasing gastrin serum levels.MethodsIn vivo, male Fischer 344 rats (immediately BDL) were treated with vehicle or OME (40 mg/Kg BW daily) for 1 wk. We measured changes in: liver histology, and intrahepatic bile duct mass in liver sections; H3 histone and PCNA and secretin receptor (SR) gene expression; basal and secretin‐stimulated cAMP levels in cholangiocytes bile and bicarbonate secretion in bile fistula rats. Changes in SR and secretin‐stimulated secretion are key functional tools for evaluating the degree of biliary hyperplasia.ResultsChronic in vivo administration of OME to BDL rats increased gastrin serum levels and reduced biliary proliferation and hyperplasia and decreased SR expression and significantly inhibited the stimulatory effects of secretin on cAMP levels and ductal secretion compared to vehicle treated controls.ConclusionThe usage of OME may be important for the management of chronic cholestatic liver diseases.

A strategy to reduce biliary clearance in early drug discovery

IntroductionBiliary excretion can modulate the pharmacokinetic profile of drug candidates, and may represent a liability for drug-drug interactions. This study proposes a strategy to reduce biliary clearance using the efflux ratio in Caco-2 cells in parallel to an abbreviated pharmacokinetic study in bile duct-cannulated rats (BDC). MethodsApical to basolateral (A to B) and basolateral to apical (B to A) permeability of 20 new chemical entities (NCEs) were determined in a 24-well permeability assay. In parallel, biliary excretion was determined in an abbreviated format in BDC rats. Test compounds were administered via an intravenous dose of 1mg/kg and the percentage (%) of parent compound excreted in the bile in the first 3hours after dosing was determined by LC-MS/MS analysis. ResultsA reasonably good correlation (r2=0.635) between the in vitro efflux ratio from the Caco-2 assay and in vivo biliary excretion of parent compound in BDC rats was observed. All seven compounds with an efflux ratio of <5 had less than 25% of the parent excreted in rat bile. In contrast, 3 out of the 13 compounds with an efflux ratio >5 had less than 25% of the dose excreted in rat bile. DiscussionThis suggests that a compound with an efflux ratio of <5 is at lower risk of having significant biliary clearance and that Caco-2 efflux ratio obtained from a high throughput screening assay may be used as an early indicator of biliary excretion. Although, we propose to reduce the occurrence of false positive prediction for biliary clearance (23%) by performing abbreviated PK in BDC rats for compounds with high efflux ratio.

Detection and characterization of ticlopidine conjugates in rat bile using high‐resolution mass spectrometry: applications of various data acquisition and processing tools

Ticlopidine, an antiplatelet drug, undergoes extensive oxidative metabolism to form S-oxide, N-oxide, hydroxylated and dealkylated metabolites. However, metabolism of ticlopidine via conjugation has not been thoroughly investigated. In this study, multiple data acquisition and processing tools were applied to the detection and characterization of ticlopidine conjugates in rat bile. Accurate full-scan mass spectrometry (MS) and collision-induced dissociation (CID) MS/MS data sets were recorded using isotope pattern-dependent acquisition on an LTQ/Orbitrap system. In addition, mass spectral data from online H/D exchanging and high collision energy dissociation (HCD) were recorded. Data processes were carried out using extracted ion chromatography (EIC), mass defect filter (MDF) and isotope pattern filter (IPF). The total ion chromatogram displayed a few major conjugated metabolites and many endogenous components. Profiles from EIC and IPF processes exhibited multiple conjugates with no or minimal false positives. However, ticlopidine conjugates that were not predictable or lost a chorine atom were not found by EIC or IPF, respectively. MDF was able to detect almost all of ticlopidine conjugates although it led to a few more false positives. In addition to CID spectra, data from HCD, H/D exchanging experiments and isotope pattern simulation facilitated structural characterization of unknown conjugates. Consequently, 20 significant ticlopidine conjugates, including glucuronide, glutathione, cysteinylglycine, cysteine and N-acetylcysteine conjugates, were identified in rat bile, a majority of which are associated with bioactivation and not previously reported. This study demonstrates the utility and limitation of various high-resolution MS-based data acquisition and processing techniques in detection and characterization of conjugated metabolites.

QUANTITATIVE ANALYSIS OF TEN DITERPENOIDS IN RAT BILE AFTER ORAL ADMINISTRATION OF Isodon rubescens EXTRACT BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY-ELECTROSPRAY IONIZATION TANDEM MASS SPECTROMETRY

Diterpenoids are the primary bioactive ingredients in Isodon rubescens named Dong Ling Cao in China which possess many pharmacological activities, including anti-tumor, anti-microbial, anti-inflammatory, anti-catastrophe, anti-oxidation, and anti-inflammatory effects. We employed a sensitive and selective high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry (HPLC–ESI–MS/MS) method for the quantification of ten diterpenoids in rat bile. The ten diterpenoids are effusanin A (1), lasiodonin (2), oridonin (3), epinodosin (4), nodosin (5), ponicidin (6), rabdoternin A (7), enmenol (8), lasiokaurin (9), and lasiokaurinol (10). The sulfamethoxazole was used as internal standard (IS). The bile samples were pretreated by liquid-liquid extraction with ethyl acetate (EtOAc). Chromatographic separation was performed on a C18 column with linear gradient elution using water and methanol at a flow rate of 0.8 mL/min. The precursor and product ions of analytes were monitored on a hybrid quadrupole linear ion trap mass spectrometer equipped with a turbo ion spray interface in positive and negative mode, respectively, using multiple–reaction monitoring (MRM). The specificity, linearity, accuracy, precision, recovery, matrix effect, and several of stabilities have been validated for diterpenoids in rat bile samples. The results showed that this method is robust, specific, and sensitive and it can successfully fulfill the requirement of excretion study of the ten diterpenoids in Isodon rubescens.

Identification of in vitro and in vivo metabolites of isoimperatorin using liquid chromatography/mass spectrometry

The objective of the present study was to develop a practical strategy for the identification of metabolites following the in vivo metabolism and in vitro microbial biotransformation of isoimperatorin using liquid chromatography hybrid triple quadrupole-linear ion trap mass spectrometry (LC/QTRAP–MS) and liquid chromatography time of flight mass spectrometry (LC/TOF–MS). As a result, 19 metabolites were characterised in rat urine, plasma, bile and faeces after the oral administration of isoimperatorin and 13 products were identified in the sample from the transformation. Four metabolites were prepared by in vitro microbial biotransformation, and one was confirmed to be a novel compound. The side chain of isoimperatorin was found to be the primary metabolic site that underwent oxidation metabolism both in vivo and in vitro, and the metabolism of isoimperatorin in vivo and in vitro has good correlation. This is the first study of the metabolism of isoimperatorin in vivo.

Pharmacokinetics of 2,3,5,4′‐tetrahydroxystilbene‐2‐O‐β‐D‐glucoside in rat using ultra‐performance LC‐quadrupole TOF‐MS

2,3,5,4'-Tetrahydroxystilbene-2-O-β-D-glucoside (THSG) from Polygoni multiflori has been demonstrated to possess a variety of pharmacological activities, including antioxidant, anti-inflammatory and hepatoprotective activities. Ultra-performance LC-quadrupole TOF-MS with MS Elevated Energy data collection technique and rapid resolution LC with diode array detection and ESI multistage MS(n) methods were developed for the pharmacokinetics, tissue distribution, metabolism, and excretion studies of THSG in rats following a single intravenous or oral dose. The three metabolites were identified by rapid resolution LC-MS(n). The concentrations of the THSG in rat plasma, bile, urine, feces, or tissue samples were determined by ultra-performance LC-MS. The results showed that THSG was rapidly distributed and eliminated from rat plasma. After the intravenous administration, THSG was mainly distributing in the liver, heart, and lung. For the rat, the major distribution tissues after oral administration were heart, kidney, liver, and lung. There was no long-term storage of THSG in rat tissues. Total recoveries of THSG within 24 h were low (0.1% in bile, 0.007% in urine, and 0.063% in feces) and THSG was excreted mainly in the forms of metabolites, which may resulted from biotransformation in the liver.

PHARMACOKINETICS, BIOAVAILABILITY, AND METABOLISM OF 2,3,5,4′-TETRAHYDROXYSTILBENE-2-O-β-D-GLUCOSIDE IN RATS BY ULTRA-PERFORMANCE LIQUID CHROMATOGRAPHY–QUADRUPOLE TIME-OF-FLIGHT MASS SPECTROMETRY AND HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY-ULTRAVIOLET DETECTION

2,3,5,4′-tetrahydroxystilbene-2-O-β-D-glucoside (THSG), a natural product purified from the Chinese medical herb Polygoni multiflori Radix, has been demonstrated to possess many pharmacological activities, including antioxidant, anti-inflammatory, hepatoprotective, and antitumor properties. Ultra-performance liquid chromatography–time-of-flight mass spectrometry (UPLC-Q-TOF/MS) and high-performance liquid chromatography-ultraviolet detection (HPLC-UV) methods were developed for the pharmacokinetics, bioavailability, absorption, and metabolism studies of THSG in rats following a single intravenous or oral administration. The metabolites (M1 and M2) were identified in plasma by UPLC-Q-TOF/MS. The concentrations of the THSG in rat plasma, and bile, urine, or feces samples were determined by HPLC-UV. The results show THSG was rapidly absorbed within 30 min in plasma. Absolute bioavailability of THSG was 40%. Total recovery of unchanged THSG within 24 hr were low (0.041% in bile, 0.06% in feces), whereas the amount of unchanged THSG excreted in the urine within 24 hr was lower than LLOQ.

Simple and rapid quantitation of 21 bile acids in rat serum and liver by UPLC-MS-MS: effect of high fat diet on glycine conjugates of rat bile acids.

In this report, we present a simple and rapid method for analysis of 21 kinds of bile acids and the conjugates in rat serum and liver samples by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) in the negative ionization mode, using cholic-2, 2, 4, 4-d4 acid as internal standard. After liquid-liguid extraction from serum and liver samples, specimens were analyzed by UPLC equipped with an Acquity TQD tandem quadrupole mass spectrometer. All of the 21 bile acids were sufficiently separated within 5 min. For most bile acids, calibration curves showed good linearities in the range of 0.25 to 5000 ng/mL for serum samples, 2.5 ng/g to 50 microg/g for liver samples. The limits of detection (LOD) were estimated to be less than 0.25 to 7.5 ng/mL in serum, less than 2.5 to 10 ng/g in liver samples. The present method was validated with respect to repeatability; the coefficient of variation (CV) values were less than 26.7% in the serum and 25.9% in the liver. In the animal study, we compared 21 bile acids in the serum and liver samples of the stroke-prone spontaneously hypertensive (SHRSP) rats fed with control (SP) diet or high-fat and high-cholesterol-containing (HFC) diet. By feeding with HFC diet, the glycine conjugates of some bile acids significantly increased and the taurine conjugate of ulsodeoxicolate (TUDC) decreased in serum and liver samples. Our results suggest that the change of bile acid profiles could be applied for the diagnosis of non-alcoholic fatty liver disease (NAFLD).

Pharmacokinetics, tissue distribution and excretion of manganese (III) meso-tetra [3-(2-(2-methoxy)-ethoxy) ethoxy] phenyl porphyrin chloride, a novel superoxide dismutase mimic, in Wistar rats

Manganese (III) 5, 10, 15, 20-tetrakis [3-(2-(2-methoxy)-ethoxy) ethoxy] phenyl porphyrin chloride, designated HSJ-0017, is a novel superoxide dismutase mimic. It exhibits strong free-radical scavenging activities in vitro and in vivo. The aim of the present study was to investigate the pharmacokinetics, tissue distribution and excretion of HSJ-0017 in Wistar rats following a single intravenous administration. Wistar rats were given different doses of HSJ-0017 by single intravenous injection. Biological samples of rats were collected and were assayed by the HPLC method. The pharmacokinetics, tissue distribution and excretion of HSJ-0017 were investigated. The pharmacokinetic data of HSJ-0017 in rats following intravenous injection was best-fit by a two-compartment model. T max of HSJ-0017 in plasma following intravenous injection was 0.083 h. AUC and plasma drug concentration were found to increase in a dose-related fashion. The highest concentrations of HSJ-0017 were detected in the liver (82.25 ± 13.99 μg/g) of rats, followed by the kidney, small intestine, lung, plasma, heart, spleen, and stomach within 2 h postdose. No HSJ-0017 was detected in the uterus, parorchis or brain of rats during the 24-h period of examination. The total cumulative excretion of HSJ-0017 in rat bile and urine were found to be 78.85 and 67.58 %, respectively. Our study has led to the view that the HSJ-0017 can be rapidly distributed to tissues after intravenous administration, but cannot diffuse through the blood-brain barrier. The faecal and biliary excretion of unchanged HSJ-0017 are the major routes of HSJ-0017 elimination.