Kinetic evaluation of 3β-hydroxycholest-5-en-7-one (7-ketocholesterol) stability during saponification

Article abstract of DOI:10.1007/BF02518118

To clarify conflicting information regarding 7-ketocholesterol (7-KC) recovery from saponification, we evaluated the stability of 7-KC in methanolic alkaline medium. We added 1 N alcoholic KOH to 7-KC or lard spiked with 7-KC and held the mixtures at 45, 55, 65, and 75°C for different times to simulate various saponification conditions. Gas-chromatographic determination of residual 7-KC with 5α-cholestane as the internal standard (IS) showed that the higher the saponification temperature, the greater the 7-KC degradation. Hot saponification at 75°C for 30 min caused extensive destruction and left only 31% 7-KC. 7-KC loss during saponification could be described by pseudo first-order kinetics, and the dependence of degradation rate on temperature (T, K) by k (h^-1) = (2.6 × 10¹⁷) exp (-1.36 × 10⁴/T). As predicted by the kinetic equation, 7-KC loss during room-temperature saponification (21°C) was practically negligible; following the exposure of 7-KC or lard spiked with 7-KC to 1 N alcoholic KOH for 18 h, about 97% 7-KC was recovered. 6-Ketocholestanol, when used as an IS, should be looked at carefully because of potential tautomerization, leading to the formation of two enol isomers when in extended contact with trimethylsilyl derivatization reagents.