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.

Rapid screening of drug-protein binding using high-performance affinity chromatography with columns containing immobilized human serum albumin.

For drug candidates, a plasma protein binding (PPB) more than 90% is more meaningful and deserves further investigation in development. In the study, a high-performance liquid chromatography method employing column containing immobilized human serum albumin (HSA) to screen in vitro PPB of leading compounds was established and successfully applied to tested compounds. Good correlation (a coefficient correlation of 0.96) was attained between the reciprocal values (X) of experimentally obtained retention time of reference compounds eluted through HSA column and the reported PPB values (Y) with a correlation equation of Y = 92.03 - 97.01X. The method was successfully applied to six test compounds, and the result was confirmed by the conventional ultrafiltration technique, and both yielded equal results. However, due to the particular protein immobilized to column, the method cannot be applied for all compounds and should be exploited judiciously based on the value of the logarithmic measure of the acid dissociation constant (pKa) as per the requirement. If α1-acid glycoprotein and other plasma proteins could be immobilized like HSA with their actual ratio in plasma to column simultaneously, the result attained using immobilized column may be more accurate, and the method could be applied to more compounds without pKa limitation.