DEVELOPMENT OF A VALIDATED HIGH PERFORMANCE LIQUID CHROMATOGRAPHY ASSAY FOR THE DETECTION AND QUANTIFICATION OF METHOXYCHLOR AND ITS MONO- AND BIS-HYDROXY METABOLITES FROM OVARIAN FOLLICLE CULTURE MEDIA

The chlorinated organic pesticide methoxychlor is a proven endocrine disrupting chemical. Methoxychlor is readily metabolized in the body to form polar metabolites. The major metabolic pathway is demethylation of methoxychlor to mono-OH methoxychlor and bis-OH methoxychlor (HPTE). There is no validated analytical method in the literature for the detection and quantification of methoxychlor and its two major metabolites in cell culture medium. Therefore, in the present study, we developed and validated a simple high performance liquid chromatography (HPLC) method with a one-step liquid/liquid extraction procedure to detect and quantify methoxychlor and its metabolites (mono-OH methoxychlor and HPTE) from alpha-minimum essential media that is used for ovarian follicle cell culture studies. The HPLC diode array detection method reported here readily detects methoxychlor and its metabolites (mono-OH methoxychlor and HPTE), with a limit of detection of methoxychlor, mono-OH methoxychlor, and HPTE in culture medium of about 7.8 ng/120 mu L, 4.8 ng/120 mu L,and 10.5 ng/120 mu L,respectively. The mean extraction efficiency values for methoxychlor, mono-OH methoxychlor, and HPTE were 82%, 88%, and 88%, respectively.