Development and validation of HPLC-MS/MS procedure for determination of 3,4,4',5-tetra-methoxystilbene (DMU-212) and its metabolites in ovarian cancer cells and culture medium.

The synthetic resveratrol analogue DMU-212 (3,4,4',5-tetramethoxystilbene) has been shown to possess stronger anticancer activity than resveratrol in a variety of tumour cells. To date, there has been no appropriate procedure that would ensure a reliable data about levels of metabolic products of DMU-212 in cancer cell lines. The purpose of this study was to develop a new procedure for determination of DMU-212 and its three metabolites (DMU-214, DMU-281, DMU-291) in cell lines. Analyses were performed using an HPLC system coupled with a triple quadrupole mass spectrometer operating in multiple reaction monitoring mode. Separation was conducted using a C18 column at a flow rate 800μL/min with a mobile phase consisting of 5mM ammonium acetate with 0.1% formic acid (solvent A) and acetonitrile (solvent B). The new methodology is fast, simple and has excellent specificity. Moreover, it showed good linearity in two matrices - cell lysates and culture media. Accuracy values for analytes evaluated at different concentration levels ranged from 0.43 to 18% (%bias). The intra-day and inter-day precision, expressed as CV, was in a range 0.49-5.5% and 0.83-13%, respectively. The validated procedure was successfully applied to quantify the resveratrol analogues in the human ovarian cancer cell line SKOV3.