title: Evaluation of novel derivatisation reagents for the analysis of oxysterols.
abstract: Oxysterols are oxidised forms of cholesterol that are intermediates in the synthesis of bile acids and steroid hormones. They are also ligands to nuclear and G protein-coupled receptors. Analysis of oxysterols in biological systems is challenging due to their low abundance coupled with their lack of a strong chromophore and poor ionisation characteristics in mass spectrometry (MS). We have previously used enzyme-assisted derivatisation for sterol analysis (EADSA) to identify and quantitate oxysterols in biological samples. This technique relies on tagging sterols with the Girard P reagent to introduce a charged quaternary ammonium group. Here, we have compared several modified Girard-like reagents and show that the permanent charge is vital for efficient MS(n) fragmentation. However, we find that the reagent can be extended to include sites for potential stable isotope labels without a loss of performance.
authors: Institute of Mass Spectrometry, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK. Electronic address: p.j.crick@swansea.ac.uk.
authors: Peter J
authors: PJ
authors: Crick
authors: Institute of Mass Spectrometry, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors: Jennifer
authors: J
authors: Aponte
authors: Institute of Mass Spectrometry, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors: T William
authors: TW
authors: Bentley
authors: College of Engineering, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors: Ian
authors: I
authors: Matthews
authors: Institute of Mass Spectrometry, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors: Yuqin
authors: Y
authors: Wang
authors: Institute of Mass Spectrometry, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK. Electronic address: w.j.griffiths@swansea.ac.uk.
authors: William J
authors: WJ
authors: Griffiths
mesh_terms: D001622:Betaine / Q000031:analogs & derivatives* / Q000737:chemistry; D002789:Cholesterol Oxidase / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D006888:Hydroxycholesterols / Q000097:blood*; D007202:Indicators and Reagents / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013261:Sterols
chemical_list: D006888:Hydroxycholesterols; D007202:Indicators and Reagents; D013261:Sterols; D001622:Betaine; D002789:Cholesterol Oxidase; C012332:Girard's reagent T
keywords: Derivatisation; Girard P reagent; Liquid chromatography; Mass spectrometry; Sterols