Article: HPLC analysis of desmosterol, 7-dehydrocholesterol, and cholesterol

A simple and sensitive method to analyze mixtures of desmosterol, 7-dehydrocholesterol and cholesterol is described. The method involves the oxidative conversion of the sterols with cholesterol oxidase, followed by high performance liquid chromatographic (HPLC) analysis. A C18 reversed phase column (3 microns, 75 X 4.6 mm) and a mixture of methanol and acetonitrile (1:1, v/v) at a rate of 1 ml/min are used to separate the sterols. The eluted sterols are quantified by measuring UV absorption at 240 nm. As little as 10 pmoles of sterol can be quantified under these conditions.

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References

1.

Ogishima T, Okuda K . An improved method for assay of cholesterol 7 alpha-hydroxylase activity. Anal Biochem. 1986; 158(1):228-32. DOI: 10.1016/0003-2697(86)90613-5. View

2.

Smith A, Brooks C . Cholesterol oxidases: properties and applications. J Steroid Biochem. 1976; 7(9):705-13. DOI: 10.1016/0022-4731(76)90071-6. View

3.

Goh E . Transfer of exogenous cholesterol to microsomes of hepatocytes investigated with [3H]desmosterol tracer. Biochem J. 1983; 216(1):137-42. PMC: 1152480. DOI: 10.1042/bj2160137. View

4.

Takada K, Takashima A, Kobayashi T, Shimoi Y . Application of high-performance liquid chromatography to the study of esterified 7-dehydrocholesterol in rat skin. Biochim Biophys Acta. 1981; 666(3):307-12. DOI: 10.1016/0005-2760(81)90288-5. View

5.

Smith A, Brooks C . Application of cholesterol oxidase in the analysis of steroids. J Chromatogr. 1974; 101(2):373-8. DOI: 10.1016/s0021-9673(00)82854-x. View

6.

Carroll R, Rudel L . Evaluation of a high-performance liquid chromatography method for isolation and quantitation of cholesterol and cholesteryl esters. J Lipid Res. 1981; 22(2):359-63. View

7.

Bricker L, Weis H, SIPERSTEIN M . In vivo demonstration of the cholesterol feedback system by means of a desmosterol suppression technique. J Clin Invest. 1972; 51(2):197-205. PMC: 302117. DOI: 10.1172/JCI106804. View

8.

Horton B, Horton J, SABINE J . Repeated estimation of liver cholesterol synthesis in vivo, using the inhibitor AY-9944. Biochim Biophys Acta. 1971; 239(3):475-81. DOI: 10.1016/0005-2760(71)90039-7. View

9.

Kraml M, Dubuc J, Dvornik D . Agents affecting lipid metabolism. XXVI. Specificity of some inhibitors of the late stages of cholesterol biosynthesis. Lipids. 1967; 2(1):5-7. DOI: 10.1007/BF02531992. View

10.

Slotte J, Bierman E . Movement of plasma-membrane sterols to the endoplasmic reticulum in cultured cells. Biochem J. 1987; 248(1):237-42. PMC: 1148524. DOI: 10.1042/bj2480237. View

11.

Bates S, Rothblat G . Regulation of cellular sterol flux and synthesis by human serum lipoproteins. Biochim Biophys Acta. 1974; 360(1):38-55. DOI: 10.1016/0005-2760(74)90178-7. View

HPLC Analysis of Desmosterol, 7-dehydrocholesterol, and Cholesterol