Gas Phase Chromatography of Steroids by Kristen B. Eik-Nes, Evan Charles Horning

By Kristen B. Eik-Nes, Evan Charles Horning

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Temperature programmed separation of Sa- and SfJ-cholestane with on OV-17 column; see Fig. 1 for an OV-1 separation. Conditions: 12 ft 10/o OV-17 (on 100-120 mesh Gas Chrom P); temperature programmed at 1°/min from 230°; injector, 260°; detector, 300°; nitrogen, 26 psi; flame detector. The separation is based upon a difference in molecular shape, but the MU value difference is slightly greater for OV-17 than for OV-1 (see Table 2). ffect which is discussed in a later section 25 With Selective Phases phase.

9 28 Gas Phase Analytical Methods for the Study of Steroid Hormones The chromatographic relationships between androsterone and pregnanediol, both as free steroids and as TMSi derivatives, are MO-TMSi 1% OV-17 TP 1"/min Test Epitest 20 30 Time(minl Fig. 9. Separation of methoxime isomers for the MO-TMSi derivatives of testosterone (TEST) and epitestosterone (EPITEST) with on OV-17 column. Conditions: 12 ft 10fo OV-17 (on 100-120 mesh Gas Chrom P) column; temperature programmed at 1°/min from 200°; injector, 250°; detector, 300°; nitrogen, 26 psi; flame detector.

The appearance of two peaks, or a shoulder, is sufficient to indicate unequivocally the existence of a mixture. VI. GLC Characterization of Steroids 1. Relative Retention Time The most popular method of GLC characterization at present is through calculation of relative retention time. The most attractive aspect of this method is that the numerical value can be determined with high precision. Unfortunately, these values, which are based upon a comparison with the behavior of a single reference substance, are temperature dependent and for this reason they are not highly useful Relative Retention Time 31 in interlaboratory comparisons of data.

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