Identification of human cytochrome P450 and UGT enzymes involved in the metabolism of ferulic acid, a major bioactive component in traditional Chinese medicines.

Ferulic acid (FA) is an active component of herbal medicines. One of the best documented activities of FA is its antioxidant property. Moreover, FA exerts antiallergic, anti-inflammatory, and hepatoprotective effects. However, the metabolic pathways of FA in humans remain unclear. To identify whether human CYP or UGT enzymes are involved in the metabolism of FA, reaction phenotyping of FA was conducted using major CYP-selective chemical inhibitors together with individual CYP and UGT Supersomes. The CYP- and/or UGT-mediated metabolism kinetics were examined simultaneously or individually. Relative activity factor and total normalized rate approaches were used to assess the relative contributions of each major human CYPs towards the FA metabolism. Incubations of FA with human liver microsomes (HLM) displayed NADPH- and UDPGA-dependent metabolism with multiple CYP and UGT isoforms involved. CYPs and UGTs contributed equally to the metabolism of FA in HLM. Although CYP1A2 and CYP3A4 appeared to be the major contributors in the CYP-mediated clearance, their contributions to the overall clearance are still minor (< 25%). As a constitute of many food and herbs, FA poses low drug-drug interaction risk when co-administrated with other herbs or conventional medicines because multiple phase I and phase II enzymes are involved in its metabolism.

Simultaneous determination of triptolide and its prodrug MC002 in dog blood by LC-MS/MS and its application in pharmacokinetic studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Tripterygium wilfordii HOOK F (TWHF) is a traditional Chinese medicine used in the treatment of various autoimmune diseases and inflammatory disorders including rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and skin diseases. Triptolide (TP) is one of the main active ingredients of this traditional Chinese medicine. MC002 is a novel semi-synthetic derivate of TP which is highly water soluble, acts as a prodrug and is converted to TP in vivo. AIM OF THIS STUDY: A sensitive, rapid method for the simultaneous determination of TP and its chemo-unstable prodrug MC002 in dog blood was developed and validated using electrospray ionization (ESI) liquid chromatography-tandem mass spectrometry (LC-MS/MS). Using this method, a pharmacokinetic study of MC002 and TP following an intravenous drip infusion of 0.2mg/kg MC002 in dogs was performed. MATERIALS AND METHODS: Chemo-degradation of the prodrug in blood samples was inhibited by the addition of a small amount of sodium fluoride solution before using liquid-liquid extraction with ethyl acetate. The concentrations of MC002 and TP in dog blood were determined using the LC-MS/MS method. RESULTS: The quantitative method showed good precision and stability and is suitable for the assay of biological samples. The pharmacokinetic study showed that the elimination of MC002 was faster than that of TP, and the concentrations and AUC0-t values of TP were higher than MC002. MC002 can rapidly convert to TP in vivo. CONCLUSIONS: This validated method was successfully applied in a pharmacokinetic study of MC002 following an intravenous drip infusion in dogs. With the development of this new prodrug of TP as a promising anti-cancer drug, this method is suitable for its further analysis in clinical studies.

Identification and characterization of psoralen and isopsoralen as potent CYP1A2 reversible and time-dependent inhibitors in human and rat preclinical studies.

Naturally occurring furanocoumarin compounds psoralen (PRN) and isopsoralen (IPRN) are bioactive constituents found in herbaceous plants. They are widely used as active ingredients in several Chinese herbal medicines. In this study, the CYP1A2 inhibitory potential of PRN and IPRN was investigated in rats in vitro and in vivo as well as in human liver microsomes. Both compounds exhibited reversible and time-dependent inhibition toward rat microsomal cyp1a2. The IC(50), k(inact), and K(I) values were 10.4 ± 1.4 µM, 0.060 ± 0.002 min(-1), and 1.13 ± 0.12 µM for PRN, and 7.1 ± 0.6 µM, 0.10 ± 0.01 min(-1), and 1.95 ± 0.31 µM for IPRN, respectively. In human liver microsomal incubations, potent reversible CYP1A2 inhibition was observed for both compounds, with IC(50) values of 0.26 ± 0.01 µM and 0.22 ± 0.03 µM for PRN and IPRN, respectively. However, time-dependent inhibition was only observed for IPRN, with kinact and KI values of 0.050 ± 0.002 min(-1) and 0.40 ± 0.06 µM, respectively. Coadministration with PRN or IPRN significantly inhibited cyp1a2 activity in rats, with the area under the curve (AUC) of phenacetin increasing more than 5-fold. Simcyp simulation predicted that PRN would cause 1.71- and 2.12-fold increases in the phenacetin AUC in healthy volunteers and smokers, respectively. IPRN, on the other hand, would result in 3.24- and 5.01-fold increases in phenacetin AUCs in healthy volunteers and smokers, respectively. These findings represent the first detailed report comparing the potential drug-drug interactions of PRN and IPRN, and provide useful information for balancing safe and efficacious doses of PRN and IPRN.

Development and preclinical studies of broad-spectrum anti-HIV agent (3'R,4'R)-3-cyanomethyl-4-methyl-3',4'-di-O-(S)-camphanoyl-(+)-cis-khellactone (3-cyanomethyl-4-methyl-DCK).

In prior investigation, we discovered that (3'R,4'R)-3-cyanomethyl-4-methyl-3',4'-di-O-(S)-camphanoyl-(+)-cis-khellactone (4, 3-cyanomethyl-4-methyl-DCK) showed promising anti-HIV activity. In these current studies, we developed and optimized successfully a practical 10-step synthesis for scale-up preparation to increase the overall yield of 4 from 7.8% to 32%. Furthermore, compound 4 exhibited broad-spectrum anti-HIV activity against wild-type and drug-resistant viral infection of CD4+ T cell lines as well as peripheral blood mononuclear cells by both laboratory-adapted and primary HIV-1 isolates with distinct subtypes and tropisms. Compound 4 was further subjected to in vitro and in vivo pharmacokinetic studies. These studies indicated that 4 has moderate cell permeability, moderate oral bioavailability, and low systemic clearance. These results suggest that 4 should be developed as a promising anti-HIV agent for development as a clinical trial candidate.