•A highly sensitive and effective LC–MS/MS method was developed and validated.•Simultaneous quantification of a novel adenosine analog and its two major metabolites in hamster blood was accomplished.•The method was successfully applied to the pharmacokinetic study of the compound on hamster.2′,3′,5′-Tri-O-acetyl-N6-(3-hydroxylaniline)adenosine (IMM-H007, once called WS070117) is being developed as a novel anti-hyperlipidemia agent for its high efficacy and low toxicity. In this study, a sensitive and specific liquid chromatography–tandem mass spectrometric (LC–MS/MS) method was established for the simultaneous quantification of IMM-H007 and its two major metabolites (3S,4R,5R)-2-(hydroxymethyl)-5-(6-((3-hydroxyphenyl)amino)-9H-purin-9-yl)tetrahdrofuran-3,4-diol (M1) and ((2R,3S,4R,5R)-3,4-dihydroxy-5-(6-((3-hydroxyphenyl)amino)-9H-purin-9-yl)tetrahydrofuran-2-yl)methyl dihydrogen phosphate (MP) in hamster blood. An analogue of IMM-H007, WS070119 was used as the internal standard. Blood samples were prepared by a simple protein precipitation with acetonitrile. The chromatographic separation was performed on a ReproSil-Pur 120C18 column (3 μm, 2 mm × 100 mm) with a gradient mobile phase of methanol/water containing 0.1% formic acid (v/v) in a flow rate of 0.2 mL/min. Detection was carried out on a triple quadrupole tandem mass spectrometer equipped with electrospray ionization (ESI) in positive ion selective reaction monitoring (SRM) mode. The monitored transitions were 486.2 → 228.1 for IMM-H007, 360.0 → 228.0 for M1, 440.0 → 228.0 for MP and 374.1 → 242.0 for the internal standard, respectively. Satisfactory linearity was obtained for the analytes over the range of 1–500 ng/mL for IMM-H007, 2–1000 ng/mL for M1 and 10–5000 ng/mL for MP. The lower limits of the quantification (LLOQs) were 1 ng/mL for IMM-H007, 2 ng/mL for M1 and 10 ng/mL for MP. The intra-day and inter-day precisions (RSD, %) of the analytes were within 14.2%, and the accuracy (RE, %) ranged from −9.4% to 10.7%. The average recoveries of the analytes were more than 80.0%. The analytes were proved to be stable during given storage, preparation, and analytic procedures. The method was successfully applied to the pharmacokinetic study in hamsters after oral administration of IMM-H007.

•Simultaneous determination of DNJ, fagomine and DAB in rat plasma was done.•A simple, selective and sensitive LC–MS/MS method was developed and validated.•The method was applied to a pharmacokinetic study of mulberry twig in rats.Fagomine, 1-deoxynojirimycin (DNJ) and 1,4-dideoxy-1,4-imino-d-arabinitol (DAB) are the major bioactive constituents in the active fraction of alkaloids from the traditional Chinese medicine mulberry twig (Ramulus Mori, Chinese name Sang Zhi), which has a strong activity on α-glucosidase in vitro and in vivo. A rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of DNJ, fagomine and DAB in rat plasma. Plasma samples were prepared using a simple protein precipitation by the addition of 1% volume of Tris and two volumes of methanol–acetonitrile. The analytes and internal standard (IS, miglitol) were chromatographed in an XBridge™ amide column with a gradient mobile phase of acetonitrile–water (0.1% ammonium hydroxide) at a flow rate of 0.7 mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer equipped with electrospray ionization (ESI) source in positive ion mode by multiple reaction monitoring (MRM) mode. Linear detection responses were obtained for DNJ ranging from 5.00 to 5000.00 ng/mL, 10.00 to 2500.00 ng/mL for fagomine and DAB. The lower limits of quantification (LLOQs) were 5.00, 10.00, 10.00 ng/mL for DNJ, fagomine and DAB, respectively. Intra-day and inter-day precisions (R.S.D.%) were within 10% for three analytes with accuracies (R.E.%) less than 12%. The mean recoveries of analytes were greater than 85%. All analytes were proved to be stable during the sample storage, preparation and analytic procedures. The method was successfully applied to the pharmacokinetic study of the three alkaloids in rats after oral administration of the active fraction of alkaloids from mulberry twig.

•MTC-220, a conjugate of paclitaxel and muramyl dipeptide analog, has been synthesized as a new anti-tumor agent.•MTC-220 cannot only retain paclitaxel's anti-tumor ability but also significantly prevent tumor metastasis in vivo.•Simultaneous determination method of MTC-220 and its metabolites (paclitaxel and MDA-linker) in beagle dog plasma was developed.•This method was useful for further characterizations in pharmacokinetics of MTC-220 and its metabolites.A sensitive LC–ESI-MS/MS method for simultaneous determination of MTC-220 and its metabolites (paclitaxel and MDA-linker) in dog plasma has been developed and validated. After addition of docetaxel (internal standard), plasma samples containing MTC-220, paclitaxel and MDA-linker were prepared based on a simple protein precipitation by adding two volumes of acetonitrile. The separation was performed on a ZorbaxSB-C18 column (3.5 μm, 2.1 mm × 100 mm) at a flow rate of 0.2 ml/min, using acetonitrile/water containing 0.1% formic acid (v/v) as mobile phase. The detection was performed on a triple quadrupole tandem mass spectrometer equipped with electrospray ionization (ESI) by selected reaction monitoring (SRM). The MS/MS ion transit ions monitored were 1444.4 → 623.8 for MTC-220, 876.4 → 307.9 for paclitaxel, 631.2 → 531.2 for MDA-linker and 830.2 → 549.1 for the internal standard. Linear detection responses were obtained for MTC-220, paclitaxel and MDA-linker ranging from 10 to 5000, 5 to 2500 and 5 to 500 ng/ml, respectively. The lower limits of quantitation (LLOQs) for MTC-220, paclitaxel and MDA-linker were 10, 5 and 5 ng/ml, respectively. The intra-day and inter-day precisions (RSD, %) of the three analytes do not exceed 10.9% except for LLOQs (≤17.50), and the accuracy (RE, %) were within ±17.5% for LLOQs and ±12.6% for the others. The average recoveries of three compounds were greater than 85.0%. The analytes were proved to be stable during all sample storage, preparation and analytic procedures. The validated method was successfully applied to pharmacokinetic studies of MTC-220 and its metabolites in beagle dogs after intravenous infusion of MTC-220 at 2.5 mg/kg.

•Lx2-32C, a novel taxane derivative, had significant antitumor activity.•Lx2-32c had potent antiproliferative effect on several drug-resistant tumor cells.•Quantitative determination method of Lx2-32C in rat plasma was developed.•This method was useful for further characterizations of pharmacokinetics of Lx2-32c.A sensitive and reliable LC–MS/MS method for the determination of Lx2-32c, a novel taxane derived from cephalomannine, has been developed and validated. Plasma samples containing Lx2-32c and paclitaxel (internal standard) were prepared based on a simple protein precipitation by the addition of two volumes of acetonitrile. The analyte and internal standard were separated on a Zorbax SB-C18 column (3.5 μm, 2.1 mm × 100 mm) with the mobile phase of acetonitrile/water containing 0.1% formic acid (v/v) with gradient elution at a flow rate of 0.2 ml/min. The detection was performed on a triple quadrupole tandem mass spectrometer equipped with atmospheric pressure chemical ionization (APCI) by multiple reactions monitoring (MRM) of the transitions at m/z 887.5 → 264.3 for Lx2-32c and 854.5 → 286.2 for IS. Linear detection responses were obtained for Lx2-32c ranging from 1 to 1000 ng/ml. Inter- and intra-day precision (R.S.D.%) were all within 15% and the accuracy (R.E.%) was equal or lower than 8%. The lower limit of quantitation (LLOQ) was 1 ng/ml and the average recovery was greater than 91.5%. The method was successfully applied to the pharmacokinetic study of Lx2-32c in rat plasma.

A rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for the determination of TBI-166 in rat plasma was developed and validated. Propranolol was used as the internal standard (IS). Plasma samples containing TBI-166 and IS were prepared based on a simple protein precipitation by the addition of ten volumes of acetonitrile. Thermo Scientific TSQ Quantum triple quadrupole system with multiple reaction monitoring (MRM) positive scanning mode was applied. The monitored transitions were 590 → 478 for TBI-166 and 260 → 183 for IS. The separation was performed on a Symmetry C8 column (3.5 μm, 2.1 × 50 mm) with the mobile phase of acetonitrile/water containing 0.1 % formic acid (v/v) with gradient elution at a flow rate of 0.2 mL/min. Linear detection responses were obtained for TBI-166 ranging from 5 to 2000 ng/mL and the lower limit of quantitation (LLOQ) was 5 ng/mL. The intra- and inter-day precisions (RSD) were all within 7.2 %, while the deviation of assay accuracies was equal to or lower than ±5.5 %. The average recoveries of analyte were greater than 95.2 %. The analyte was proved to be stable during all sample storage, preparation and analytic procedures. The optimized method was successfully applied to the pharmacokinetic studies of TBI-166 in rats.

•SYL-1119 is a S1P1 modulator for the treatment of autoimmune disease with better selectivity.•A rapid LC–MS/MS method for the simultaneous determination of SYL-1119 and SYL-1119-P in rat plasma is developed.•The method was successfully applied to the pharmacokinetic study of SYL-1119 and SYL-1119-P in rats.•The pharmacokinetic parameters of SYL-1119 and SYL-1119-P in this paper are reported for the first time.SYL-1119 is a sphingosine-1-phosphate receptor 1 modulator for the treatment of autoimmune disease with better selectivity, while SYL-1119-P is its active phosphate. A sensitive and specific liquid chromatography–tandem mass spectrometry method was developed and validated for the simultaneous determination of SYL-1119 and SYL-1119-P in rat plasma. SYL-1110, an analogue of SYL-1119, was used as the internal standard. Plasma samples were prepared by protein precipitation using acetonitrile. The analytes and internal standard were separated on a Zorbax SB-C18 column (3.5 μm, 100mm × 2.1 mm) with a gradient mobile phase consisting of methanol and water containing 0.1% formic acid at a flow rate of 0.2 ml/min with an operating temperature of 20 °C. The detection was performed on a triple quadrupole tandem mass spectrometer with positive electrospray ionization in multiple reaction monitoring mode of the transitions at m/z 364 → 259 for SYL-1119, m/z 444 → 259 for SYL-1119-P, and m/z 378 → 273 for the IS. Calibration curves were linear in the range of 0.2–50 ng/ml for SYL-1119 and 10–1000 ng/ml for SYL-1119-P. The lower limit of quantification (LLOQ) was 0.2 ng/ml for SYL-1119 and 10 ng/ml for SYL-1119-P. The intra- and inter-day precisions were 5.4–12.8% for two analytes with accuracies within ±10%. The recoveries for two compounds were 91.3–104.5%. The analytes were proved to be stable during all sample storage, preparation, and analytic procedures. The method was successfully applied to the pharmacokinetic study of SYL-1119 and SYL-1119-P in rats after oral administration of SYL-1119.

•YZG-331 is a promising candidate with new mechanism for the treatment of insomnia.•A rapid and sensitive LC–MS/MS method for analysis of YZG-331 in mouse plasma is developed for the first time.•The method was fully validated and successfully applied to a pharmacokinetic study of YZG-331 in mice.•The pharmacokinetic parameters of YZG-331 in this paper are reported for the first time.A rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for the determination of YZG-331 in mouse plasma was developed. Plasma samples containing YZG-331 and YZG-441 (internal standard, IS) were prepared using a simple protein precipitation by the addition of acetonitrile. Thermo Scientific TSQ Quantum triple quadrupole system with multiple reaction monitoring (MRM) positive scanning mode was applied. The separation was performed on a ZorbaxSB-C18 column (3.5 μm, 2.1 mm × 100 mm) at a flow rate of 0.2 mL/min, using acetonitrile/water containing 0.1% formic acid (v/v) as mobile phase. The MS/MS ion transit ions monitored were 386→254 for YZG-331 and 400→268 for IS. Linear detection responses were obtained for YZG-331 ranging from 25 to 5000 ng/mL and the lower limits of quantitation (LLOQ) for the compound was 25 ng/mL. The intra- and inter-day precisions (R.S.D.%) were within 12.6% for all analytes, while the deviation of assay accuracies was within ±6.9%. The average recoveries of analytes were greater than 94.3%. The analytes were proved to be stable during all sample storage, preparation and analytic procedures. The method was successfully applied to the pharmacokinetic studies of YZG-331 in mice.

MTC-220, a conjugate of paclitaxel and muramyl dipeptide analogue, was reported to exhibit anti-tumor ability and anti-metastatic effect. The aim of present study was to investigate the elimination of MTC-220 and the related mechanisms in rats.The excretion of MTC-220 and its metabolites in bile and urine were determined in rats after intravenous administration at 4 mg/kg. Caco-2 cell monolayer, in situ liver perfusion model and in vivo pharmacokinetics with selected inhibitors in rats were used to confirm the involvement of hepatic transporters in the elimination of MTC-220. The metabolic stability of MTC-220 was assessed by the incubation with rat liver microsomes and plasma.Approximately 72 % of MTC-220 was excreted into bile and less than 0.02 % into urine after administration in rats. The Caco-2 cell monolayer was impermeable to MTC-220. In in situ liver perfusion model, the hepatic extraction ratio of MTC-220 was reduced to 40 % of control in the presence of rifampicin, an Oatps inhibitor, and the cumulative biliary excretion rates of MTC-220 were reduced to 52.9, 71.5 and 62.9 % of control when concomitant perfusion with probenecid, novobiocin and verapamil, the inhibitors of Mrp2, Bcrp and P-gp, respectively. Co-administration of rifampicin, probenecid, novobiocin and verapamil with MTC-220 increased the AUC0–t and decreased the CL of MTC-220 in certain extents in rats. MTC-220 remained metabolically intact in rat liver microsomes, but less stable in plasma incubation.In summary, the elimination of MTC-220 was mainly through the biliary excretion in unchanged form in rats. Liver transporters including Oatps, Mrp2, Bcrp and P-gp might be all involved in the hepatic elimination of MTC-220. MTC-220 exhibited the high metabolic stability in liver microsomes, but less stable in plasma. The esterases might involve in the metabolism of MTC-220 in plasma.

A rapid, simple and sensitive HPLC–ESI–MS/MS method was developed for the simultaneous determination of capsaicin and dihydrocapsaicin in rat plasma. Plasma samples containing capsaicin, dihydrocapsaicin and phenacetin (internal standard) were prepared based on a simple protein precipitation by the addition of two volumes of acetonitrile. The analytes and internal standard were separated on a Zorbax SB-C18 column (3.5 μm, 2.1 mm × 100 mm) with mobile phase of acetonitrile/water (55:45, v/v) containing 0.1% formic acid (v/v) at a flow rate of 0.2 mL/min with an operating temperature of 25 °C. Quantification was performed on a triple quadrupole mass spectrometer equipped with electrospray ionization (ESI) source by selected reaction monitoring (SRM) of the transitions at m/z 306–137 for capsaicin, m/z 308–137 for dihydrocapsaicin and m/z 180–110 for the IS. Linear detection responses were obtained for capsaicin and dihydrocapsaicin ranging from 1 to 500 ng/mL and the lower limits of quantitation (LLOQs) for the two compounds were 1 ng/mL. The intra- and inter-day precisions (R.S.D.%) were within 9.79% for the two analytes, while the deviations of assay accuracies were within ±10.63%. The average recoveries of the analytes were greater than 89.88%. The analytes were proved to be stable during all sample storage, preparation and analytic procedures. The method was successfully applied to the pharmacokinetic studies of capsaicin and dihydrocapsaicin in rats after subcutaneous administration of capsaicin (natural, containing 65% capsaicin and 35% dihydrocapsaicin).

A sensitive and accurate method for determination of bicyclol in dog plasma was developed. Thermo Scientific TSQ Quantum triple quadrupole system with multiple ion monitoring (MIM) positive scanning mode was applied. Bicyclol and DDB (IS) sodium adduct molecular ions were monitored at m/z 413 and m/z 441 in both Q1 and Q3, respectively. The collision energy in Q2 was set to 15 eV. Precipitation method was employed in the extraction of bicyclol and DDB from the biological matrix. The method was validated over 1–500 ng/mL for bicyclol. The recovery was 96.5–109.5%, and the limit of quantitation (LOQ) detection was 1 ng/mL for bicyclol. The intra- and inter-day precision of the method at three concentrations was 3.3–14.3% with accuracy of 99.9–109.0%. The method was successfully applied to bioequivalence studies of bicyclol controlled-release formulation to obtain the pharmacokinetic parameters.

This study investigated the protective effects of bicyclol against cisplatin-induced nephrotoxicity and the possible mechanisms in mice. Bicyclol (250 mg/kg, p.o., 5 days) showed significant protection as evidenced by the decrease of elevated serum creatine and blood urea nitrogen, and improvement of histopathological injury induced by cisplatin. The formation of kidney malondialdehyde with a concomitant reduction of reduced glutathione were also inhibited by bicyclol, while the activities of kidney superoxide dismutase, catalase and glutathione peroxidase were all increased, respectively. Bicyclol also inhibited the increase of kidney and serum nitric oxide induced by cisplatin. In addition, induction of induced nitric oxide synthase and nitrotyrosine were suppressed by bicyclol. Bicyclol suppressed cisplatin-induced extracelluar signal regulated kinases 1/2 and p38 mitogen-activated protein kinase activation in the kidney of mice. Results obtained demonstrate that bicyclol pre-administration can prevent the nephrotoxicity induced by cisplatin.

Bicyclol is a synthetic antihepatitis drug with antioxidative property. The present study was performed to investigate the effect of bicyclol on liver regeneration after partial hepatectomy in rats. Bicyclol (300 mg/kg) was given to rats subjected to 70% hepatectomy three times before operation. At 6, 24, and 48 h after resection, samples were collected for the measurement of serum alanine aminotransferase (ALT), total bilirubin (TBil), hepatic glycogen, malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH). Moreover, liver regeneration rate, proliferating cell nuclear antigen (PCNA) labeling, proliferation index, and histopathological examination were evaluated at 48 h after hepatectomy. As a result, bicyclol significantly increased regeneration rate, mitotic index (MI), PCNA labeling index, and proliferation index in PH rats. Additionally, bicyclol remarkably inhibited the elevation of serum ALT and TBil levels, alleviated the formation of liver MDA, restored impaired antioxidant SOD and GSH, increased hepatic glycogen content, and also attenuated hepatic vacuolar degeneration. These results suggested that bicyclol had a beneficial effect on liver regenerative capacity of the remnant liver tissue after hepatectomy, probably due to its antioxidative property.

A rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of H002 and its phosphorylated metabolite, H002-P and hydroxylated metabolite H002-M, in rat blood. H001, an analogue of H002, was used as the internal standard. Blood samples were prepared by simple protein precipitation. The analytes and internal standard were separated on a Zorbax SB-C18 column with a gradient mobile phase consisting of methanol and water containing 0.1% formic acid at a flow rate of 0.2 mL/min with an operating temperature of 20 °C. The detection was performed on a triple quadrupole tandem mass spectrometer with positive electrospray ionization in multiple-reaction monitoring mode. Linear detection responses were obtained from 0.2–100 ng/mL for H002 and H002-M, while 0.5–100 ng/mL for H002-P. The intra- and inter-day precision (RSD%) was within 11.76%, with the accuracy (RE%) ranging from –9.84% to 9.12%. The analytes were shown to be stable during sample storage, preparation and analytic procedures. The method was applied to determine the pharmacokinetics of H002 in rats, and a preliminary study showed that the pharmacokinetics of H002 correlated with its biological effect on peripheral blood lymphocytes.A rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of H002 and its phosphorylated metabolites, H002-P and hydroxylated metabolite H002-M, in rat blood. The pharmacokinetics of H002 correlated with its biological effect on peripheral blood lymphocytes.Download full-size image

Syl930 is a novel sphingosine-1-phosphate receptor subtype 1 (S1PR1) agonist for the treatment of autoimmune encephalitis with promising receptor selectivity and little risk of bradycardia. Syl930 could be reversibly converted to its phosphorylated metabolite, acting as the active form to provide therapeutic effects, but eliminated principally in the form of oxidative metabolites. The aim of the present study was to identify the cytochrome P450 isoforms (CYPs) responsible for the oxidative metabolism of Syl930. Considerable production of hydroxylated metabolite (Syl930-M1) was found in both rat blood and tissue homogenates in vivo and in vitro. Moreover, another hydroxylated metabolite, Syl930-M2, was detected in human, beagle dog and cynomolgus monkey liver microsomes with significant differences in the Km, Vmax and CLint of the metabolites among species. CYP1A1, CYP2J2, CYP4F2 and CYP3A4 were identified to be the major CYPs mediated in the hydroxylation of Syl930 by using 14 recombinant human CYPs, selective chemical inhibitors and monoclonal antibodies against CYPs. The multiple CYPs mediated oxidation was believed to be one of the reasons for the relatively short elimination half-life of Syl930.

F18 (10-chloromethyl-11-demethyl-12-oxo-calanolide), an analog of (+)-Calanolide A, is a novel small-molecule nonnucleoside reverse transcriptase inhibitor for the therapy of human immunodeficiency virus (HIV) infection. M3, the most abundant primary metabolite of F18 in human liver microsomes (HLMs) and rat liver microsomes (RLMs), is mainly excreted in bile as a glucuronide conjugate in rats after oral administration. The aim of this study was to identify the UDP glucuronosyltransferase (UGT) isoforms involved in the glucuronidation of M3 by HLMs and recombinant human UGTs and investigate the metabolic interactions of M3 with the substrates of UGTs in HLMs. As a result, UGT1A1 was the major isozyme responsible for the glucuronidation of M3, followed by UGT1A4, UGT1A9 and UGT2B7. M3 exhibited significant inhibition against UGT1A9 and UGT2B7 in both HLMs and recombinant human UGTs. In addition, M3 inhibited UGT1A9 catalyzed mycophenolic acid (MPA) glucuronidation with Ki of 0.39 μM, and M3 also inhibited the glucuronidation of 3′-azido-3′-deoxythymidine (AZT) by a “mixed-type” mechanism with Ki of 16.8 μM. The results suggest that UGT1A1 provides the major contribution to M3 glucuronidation in vitro and M3 has the potential to interact with xenobiotics and endogenous chemicals that are UGT1A9 and UGT 2B7 substrates.Download full-size image

Ethnopharmacological relevanceSchisandra chinensis (SC), officially listed as a sedative and tonic in the Chinese Pharmacopoeia, has been used as a common component in various prescriptions in Traditional Chinese Medicine (TCM) and more recently in western medicine for its antihepatotoxic effect. To assess the possible herb–drug interaction, effects of SC extracts on hepatic cytochrome P450 (P450, CYP) enzymes were studied.Material and methodsEffects of SC extracts on rat hepatic CYP450 enzymes in vitro and in vivo were investigated by probe substrates method, real-time RT-PCR assay and Western blotting analysis. Furthermore, the effects of SC alcoholic extract on the PK of four SC lignans and the drugs possibly co-administrated in vivo were studied in male Sprague-Dawley rat.ResultsSC aqueous extract and alcoholic extract showed significant inhibitory effect on the activities of rat liver microsomal CYP1A2, 2C6, 2C11, 2D2, 2E1 and 3A1/2 in vitro. Multiple administrations of SC aqueous extract (1.5 g/kg, qd×7d) and alcoholic extract (1.5 g/kg, qd×7d) increased the activities, mRNA and protein expressions of CYP2E1 and CYP3A1/2, and meanwhile, inhibited the activities and mRNA expression of CYP2D2 in vivo. The in vivo metabolism of four SC lignans, such as schisandrin, schisantherin A, deoxyshisandrin and γ-schisandrin, and chlorzoxazone was significantly accelerated, exhibited by the reduced AUC and increased CLz/F, by 7-day pretreatment with SC alcoholic extract. However, both single and multiple dosing treatments of SC alcoholic extract remarkably decreased the in vivo metabolism of tacrolimus indicated by the enhanced AUC (7–12 fold) and elevated Cmax (10 fold).ConclusionThese results revealed that the SC extracts exhibited multifaceted effects on rat hepatic CYP450 enzymes. Herb–drug interaction should be paid intense attention between SC components and drugs metabolized by different CYP450 enzymes.Download high-res image (152KB)Download full-size image

FB2 is a promising Abl/Src dual tyrosine kinase inhibitor which is designed to overcome imatinib resistance. The present study aims to investigate the role of P-glycoprotein (P-gp) in intestinal absorption of FB2 with an in vitro Caco-2 and MDCK-MDR1 cell model, single-pass intestinal perfusion model and in vivo pharmacokinetics with a selective inhibitor in rats. The results from Caco-2 cells indicated that PappB-A of FB2 and its metabolites (FB7 and FB10) were much higher than PappA-B, and the efflux ratio (PappB-A/PappA-B) of FB2, FB7 and FB10 were decreased with P-gp inhibitor LSN335984; FB2 was further confirmed to be the substrate of P-gp in MDCK-MDR1 cells. In addition, Pblood of FB2 and the cumulative amount of metabolites in mesenteric blood were elevated in a concentration-dependent manner in rat intestinal perfusion, while both of them were remarkably increased when P-gp inhibitor was added. The Foral of FB2 was increased to 24.52% when orally coadministrated with verapamil (25 mg/kg), which was significantly higher than that (5.7%) by FB2 (18 mg/kg) alone in rats. The AUC and Cmax of FB2 metabolites (FB7 and FB10) were also increased in the presence of verapamil. In conclusion, the low bioavailability of FB2 is believed to be partially due to the P-gp mediated active efflux and first-pass metabolism in the rat intestine.

Peroxisome proliferators-activated receptor α (PPARα) and oxidative stress are two important pathological factors in non-alcoholic fatty liver disease (NAFLD). Tetracycline-induced fatty liver was partly due to the disturbance of mitochondrial fatty acids β-oxidation regulated by PPARα. Bicyclol was found to protect against high fat diet-induced fatty liver through modulating PPARα and clearing reactive oxygen species (ROS). The present study was performed to further investigate the effect of bicyclol on tetracycline-induced fatty liver and related mechanism in mice. Bicyclol (75, 150, 300 mg/kg) was given orally three times in two consecutive days. Tetracycline (200 mg/kg) was injected intraperitoneally 1 h after the last administration of bicyclol. Oxidative stress, mitochondrial function, PPARα and its target genes were evaluated by biochemical and RT-PCR analysis. The activity of CYP4A was assessed by liquid chromatography/mass spectrometry (LC/MS) method. Bicyclol significantly protected against tetracycline-induced fatty liver by reducing the accumulation of hepatic lipids and elevation of serum aminotransferase. In addition, bicyclol remarkably alleviated the over-production of thiobarbituric acid-reactive substance. The reduced activity of mitochondrial respiratory chain (MRC) complexes I and IV and mitochondrial permeability transition (MPT) were also improved by bicyclol. Furthermore, bicyclol inhibited the decrease of PPARα expression and its target genes, including long-chain acyl CoA dehydrogenase (LCAD), acetyl CoA oxidase (AOX) and CYP4A at mRNA and enzyme activity level. Bicyclol protected against tetracycline-induced fatty liver mainly through modulating the disturbance of PPARα pathway and ameliorating mitochondrial function.