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Abstract
5α-pregnane-3α, 7α-diol-20-one, the main progesterone derivative in marmoset monkeys, shows a very low absorption in UV-light. Labelling of this metabolite prior to HPLC and subsequent monitoring by an UV-detector is essential. We used dansyl-hydrazine for the formation of a fluorescent derivative. The optimum conditions for the reaction of dansyl-hydrazone with hydroxypregnanolone were controlled by HPTLC, chromatographic separation is carried out by HPLC and HPTLC and is described in this paper. When the quantitation of the dansylhydrazones is carried out by HPTLC, the fluorescence intensity of the derivative can be increased by a factor up to 5 by dipping the HPTLC plate into a mixture of paraffin/n-hexane or after treatment with triethanolamin/isopropanol (1:4; by vol). Hydroxypregnanolone from marmoset urine is detected for the first time by liquid chromatography. The derivative is of practical use to determine ovulation and pregnancy in the marmoset monkey. Quantitation by HPLC and thin-layer chromatography is possible in the range of 10 to 1000 ng. The concentrations in marmoset urine during the luteal phase is in the range of 50 to 400 ng/mg creatinine.