Determination of endogenous testosterone in rat tissues following fetal alcohol exposure using HPLC with UV detection

A novel method for quantitation of brain neurosteroid levels using HPLC with UV detection is described. In this simple and reliable method, testosterone from the brain and whole blood, and the internal standard, 17α-methyl testosterone, were extracted in 20% acetonitrile–phosphate buffer (pH 2.8), followed by solid phase extraction (SPE). The calibration curve was linear in concentration ranges from 0.1 to 10 ng from 0.2 g of tissue. We successfully applied this method to the analysis of endogenous testosterone in the male offspring of rats exposed to alcohol in utero. The concentration of testosterone at 21 post delivery in fetal alcohol exposure (FAE) group was significantly greater than the concentrations in either pair-fed or the ad libitum controls. These results support the usefulness of this method as a means of quantitating neurosteroids, and illustrate its applicability to fetal alcohol exposure.