25876-54-4Relevant academic research and scientific papers
Analysis of derivatised steroids by matrix-assisted laser desorption/ionisation and post-source decay mass spectrometry
Khan, Muhammad Atif,Wang, Yuqin,Heidelberger, Sibylle,Alvelius, Gunvor,Liu, Suya,Sjoevall, Jan,Griffiths, William J.
, p. 42 - 53 (2006)
Neutral steroids are difficult to analyse using desorption ionisation methods coupled with mass spectrometry (MS). However, steroids with an unhindered ketone group can readily be derivatised with the Girard P (GP) reagent to give GP hydrazones. Steroid GP hydrazones contain a quaternary nitrogen atom and are readily desorbed in the matrix-assisted laser desorption/ionisation (MALDI) process, giving an improvement in sensitivity of two orders of magnitude. Steroids without a ketone group, but with a 3β-hydroxy-Δ5 function, can be readily converted to 3-oxo-Δ4 steroids and subsequently derivatised to GP hydrazones for MALDI analysis. In addition to giving strong [M]+ ions upon MALDI, steroid GP hydrazones give informative post-source decay (PSD) spectra. By using the accurate mass of the precursor-ion measured by MALDI-MS, in combination with the structural information encoded in its PSD spectrum, steroid structures can readily be determined.
Identification of unusual oxysterols and bile acids with 7-oxo or 3,5,6-trihydroxy functions in human plasma by charge-tagging mass spectrometry with multistage fragmentation
Griffiths, William J.,Gilmore, Ian,Yutuc, Eylan,Abdel-Khalik, Jonas,Crick, Peter J.,Hearn, Thomas,Dickson, Alison,Bigger, Brian W.,Wu, Teresa Hoi-Yee,Goenka, Anu,Ghosh, Arunabha,Jones, Simon A.,Wang, Yuqin
, p. 1058 - 1070 (2018)
7-Oxocholesterol (7-OC), 5,6-epoxycholesterol (5,6-EC), and its hydrolysis product cholestane-3,5,6-triol (3,5,6-triol) are normally minor oxysterols in human samples; however, in disease, their levels may be greatly elevated. This is the case in plasma from patients suffering from some lysosomal storage disorders, e.g., Niemann-Pick disease type C, or the inborn errors of sterol metabolism, e.g., Smith-Lemli-Opitz syndrome and cerebrotendinous xanthomatosis. A complication in the analysis of 7-OC and 5,6-EC is that they can also be formed ex vivo from cholesterol during sample handling in air, causing confusion with molecules formed in vivo. When formed endogenously, 7-OC, 5,6-EC, and 3,5,6-triol can be converted to bile acids. Here, we describe methodology based on chemical derivatization and LC/MS with multistage fragmentation (MSn) to identify the necessary intermediates in the conversion of 7-OC to 3-hydroxy-7-oxochol-5-enoic acid and 5,6-EC and 3,5,6-triol to 3,5,6-trihydroxycholanoic acid. Identification of intermediate metabolites is facilitated by their unusual MSn fragmentation patterns. Semiquantitative measurements are possible, but absolute values await the synthesis of isotope-labeled standards.—Griffiths, W. J., I. Gilmore, E. Yutuc, J. Abdel-Khalik, P. J. Crick, T. Hearn, A. Dickson, B. W. Bigger, T. H-Y. Wu, A. Goenka, A. Ghosh, S. A. Jones, and Y. Wang. Identification of unusual oxysterols and bile acids with 7-oxo or 3,5,6-trihydroxy functions in human plasma by charge-tagging mass spectrometry with multistage fragmentation.
A convenient synthesis of 7α-hydroxycholest-4-en-3-one by the hydroxypropyl-β-cyclodextrin-facilitated cholesterol oxidase oxidation of 3β,7α-cholest-5-ene-3,7-diol
Alexander, David L.,Fisher, Jed F.
, p. 290 - 294 (2007/10/02)
The initial biosynthetic conversions of cholesterol to the bile acids involve sequential 7α-hydroxylation (catalyzed by cholesterol 7α-hydroxylase) followed by C-3 oxidation and concomittant double bond migration (to a Δ4-configuration, catalyzed by 3β-Δ5-C27-steroid oxidoreductase) to provide 7α-hydroxycholest-4-en-3-one.A straightforward, and economical, preparation (on a 0.1 g scale) of this pivotal biosynthetic intermediate has been devised.Reduction of 3β-(benzoyloxy)-cholest-5-en-7-one with LiB(sec-butyl)3H provided a 4:1 mixture, respectively, of the 7α- and 7β-hydroxy diastereomers, which were separated chromatographically.Solvolytic removal of the C-3 benzoyl group gave 3β,7α-cholest-5-ene-3,7-diol.A suspension of the 1:1 (v/v) complex (formed by mutual dissolution in MeOH, followed by evaporation of the solvent) of this diol with hydroxypropyl-β-cyclodextrin, at a concentration of 1 mg mL-1 (in neutral phosphate buffer), was converted by Brevibacterium sp cholesterol oxidase (0.25 U mg-1 of substrate) and catalase (70 U mg-1 of substrate, to recover O2 from the H2O2 produced by the enzymatic oxidation) to a suspension of 7α-hydroxycholest-4-en-3-one and the hydroxypropyl-β-cyclodextrin.The yield for the enzymatic conversion was in excess of 90percent.A much poorer and less reproducible yield ( 20percent) was seen in the absence of the hydroxypropyl-β-cyclodextrin.Routine extraction of this aqueous suspension, and chromatographic purification (85:15 CHCl3/acetone v/v on silica) of the residue, gave pure 7α-hydroxycholest-4-en-3-one in 68percent isolated yield.This route is a significant improvement, in terms of reaction scale and convenience, over the previous procedures for the preparation of this steroid. - Keywords: (7α)-hydroxycholesterol; hydroxypropyl-β-cyclodextrin; cholesterol oxidase; (7α)-hydroxycholest-4-en-3-one; bile acid biosynthesis; cholesterol 7α-hydroxylase
CHOLESTEROL CONVERSION TO Δ4-CHOLESTENONE BY CHOLESTEROL OXIDASE IN POLYPHASIC SYSTEMS : EXTENSION TO THE SELECTIVE OXIDATION OF 7Β-HYDROXYCHOLESTEROL
Lee, Kang Min,Biellmann, Jean-Francois
, p. 1135 - 1140 (2007/10/02)
The preparative use of cholesterol oxidase has been extended to polyphasic systems.The enzyme is active in microemulsions with the organic phase composed of mixtures of cyclohexane and chloroform.The kinetic data for oxidation of 7α- and 7β-hydroxycholesterol in microemulsion with the enzyme from Streptomyces are similar to those of cholesterol.The cholesterol oxidase is active in the two phase system aqueous buffer - butyl acetate and preparative enzymic conversion of 7β-hydroxycholesterol to Δ4-7β-hydroxycholestenone was performed in this medium.The enzymic conversion of cholesterol to Δ4-cholestenone was also performed in two liquis-solid systems, in buffer with cholesterol adsorbed on silica gel and in organic medium with cholesterol oxidase and catalase entrapped in Chromosorb.
