A validated LC-MS/MS assay for quantification of 24(S)-hydroxycholesterol in plasma and cerebrospinal fluid.

Methods Development

Abstract

24(S)-hydroxycholesterol [24(S)-HC] is a cholesterol metabolite that is formed almost exclusively in the brain. The concentrations of 24(S)-HC in cerebrospinal fluid (CSF) and/or plasma might be a sensitive marker of altered cholesterol metabolism in the CNS. A highly sensitive 2D-LC-MS/MS assay was developed for the quantification of 24(S)-HC in human plasma and CSF. In the development of an assay for 24(S)-HC in CSF, significant nonspecific binding of 24(S)-HC was observed and resolved with the addition of 2.5% 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) into CSF samples. The sample preparation consists of liquid-liquid extraction with methyl-tert-butyl ether and derivatization with nicotinic acid. Good linearity was observed in a range from 1 to 200 ng/ml and from 0.025 to 5 ng/ml, for plasma and CSF, respectively. Acceptable precision and accuracy were obtained for concentrations over the calibration curve ranges. Stability of 24(S)-HC was reported under a variety of storage conditions. This method has been successfully applied to support a National Institutes of Health-sponsored clinical trial of HP-β-CD in Niemann-Pick type C1 patients, in which 24(S)-HC is used as a pharmacodynamic biomarker.

Authors

Sidhu, Rohini; Jiang, Hui; Farhat, Nicole Y; Carrillo-Carrasco, Nuria; Woolery, Myra; Ottinger, Elizabeth; Porter, Forbes D; Schaffer, Jean E; Ory, Daniel S; Jiang, Xuntian;

Keywords

  • Central Nervous System/ metabolism
  • Cholesterol/ metabolism
  • Chromatography, Liquid
  • Humans
  • Hydroxycholesterols/ blood
  • Hydroxycholesterols/ cerebrospinal fluid
  • Niemann-Pick Disease, Type C/ blood
  • Niemann-Pick Disease, Type C/ cerebrospinal fluid
  • Tandem Mass Spectrometry
  • United States
  • beta-Cyclodextrins/ pharmacology

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