Multiple internal reflection spectroscopy has been applied to the study and identification of microgram quantities of pure steroids. The methodology described is simple and requires a minimum of sample preparation. The steroid spectra which have been obtained on 20 to 100 μg of sample are comparable to those obtained with KBr pellets. The steroids chosen to illustrate the method are of particular interest in assisting with the identification of the products of intermediary metabolism as it pertains to steroid hormones. The anomalous behavior of one of these steroids is discussed.

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