Investigation of the Cholesterol Biosynthesis by Heart-Cut Liquid Chromatography and Mass Spectrometric Detection

J Chromatogr A. 2024 Dec 6:1738:465475. doi: 10.1016/j.chroma.2024.465475. Epub 2024 Oct 26.

Abstract

The biosynthesis and homeostasis of cholesterol are essential for cellular function. Cholesterol is a major lipid with multiple roles in membrane stability, signaling, or as a precursor for other molecules. Because of the structural similarity of the sterols involved in the biosynthesis, their accurate identification and quantification is still challenging. Moreover, the huge difference in the concentration of cholesterol and its precursors can cause interferences during the detection. To overcome these problems, a heart-cut liquid chromatographic method was developed by evaluating 38 different columns to achieve optimal separation. The method efficiently separates all sterol biosynthesis intermediates, with detection limits in the low nmol/L-range and an upper limit of quantification of 9 mmol/L for cholesterol by using triple quadrupole mass spectrometric detection. Investigation of lung carcinoma cells treated with statins demonstrated the capability to detect a biological response, showing inhibition of sterol synthesis. This technique offers a robust tool for studying cholesterol biosynthesis and its role in disease.

Keywords: 2D-LC; Cancer; Lipidomics; Statins; Sterols.

MeSH terms

  • Cell Line, Tumor
  • Cholesterol* / analysis
  • Cholesterol* / metabolism
  • Chromatography, High Pressure Liquid / methods
  • Chromatography, Liquid / methods
  • Humans
  • Hydroxymethylglutaryl-CoA Reductase Inhibitors / analysis
  • Hydroxymethylglutaryl-CoA Reductase Inhibitors / chemistry
  • Limit of Detection
  • Mass Spectrometry / methods

Substances

  • Cholesterol
  • Hydroxymethylglutaryl-CoA Reductase Inhibitors