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Precise ultramicro determination of steroid hormones by combined TLC and GLC analysis
Author(s) -
Vandenheuvel F. A.,
Hinderks G. J.,
Nixon J. C.
Publication year - 1965
Publication title -
journal of the american oil chemists' society
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.512
H-Index - 117
eISSN - 1558-9331
pISSN - 0003-021X
DOI - 10.1007/bf02540130
Subject(s) - steroid , elution , chemistry , trimethylsilyl , chromatography , reagent , ether , quantitative analysis (chemistry) , steroid hormone , hormone , organic chemistry , biochemistry
The migration of steroid hormones on TLC plates is predictable on the basis of the number of double bonds and carbonyl and hydroxyl groups they contain. Developed plates can therefore be divided into zones or bands corresponding to well‐defined classes of steroids. Steroids can be eluted from the adsorbent quantitatively and without contamination. The eluate from each zone contains a mixture which is much simpler than the original one. The conversion of OH‐containing steroids to trimethylsilyl ether (TMS) derivatives is quantitative; it is complete after 3 hr by the described technic. All TMS derivatives are very stable in CS 2 solution. Whether or not they contain OH groups, steroids other than corticosteroids are stable under the present GLC conditions after treatment with TMS reagents. With amts of injected sample increasing from 0 to 0.2 Ɓg, a small apparent increase in specific response is observed with some steroids. This effect is reproducible. The use of calibration curves obtained with an internal standard (5ॅ‐pregnane) leads to accurate analyses of mixtures with injections containing 25 nanograms (0.025 Ɓg) of individual steroid. Values obtained from single trials fall 95% of the time within 2% of the mean. Peak elution time of each compound is accurately predictable on the basis of number, position, and stereoconfiguration of functional groups. An integrated TLC‐GLC method of analysis based on these findings is outlined.