Product Spotlight: OxysterolSPLASH™ LIPIDOMIX® Quantitative Mass Spec Internal Standard Mixture

Posted on November 01, 2019

Oxysterols are generated enzymatically by members of the cytochrome P450 family, hydoxysteroid dehydrogenases and non-catalytically via auto-oxidation reactions. They exist in two different states in vivo: as free oxysterols or as sterol esters that contain a fatty acyl moiety at the 3-carbon of cholesterol.

Due to the biological actions of this class of molecules, and its potential involvement in the pathophysiology of some human diseases, the structural identification and quantitation of oxysterol molecular species is a key area of efforts in this field. A significant challenge to these efforts has been the limited availability of pure primary reference and labeled internal standards.


The importance of high quality standards for absolute quantitation has been discussed in a previous blog entry (Key Considerations for Absolute Quantitation). Briefly, the LLOQ, ULOQ and the dynamic range of your quantitative assay are dependent on the design and isotopic purity of your labeled internal standard. The accuracy of your assay depends on the purity of your primary reference standards. We are proud to introduce OxysterolSPLASH™ LIPIDOMIX Quantitative mass spectrometry standards that have the requisite purity and coverage to ensure accurate, absolute quantitation of oxysterols in biological samples. Deuterated standards are currently available for purchase (OxysterolSPLASH™); primary reference standards will be available at the end of Q4, 2019.


The methods used to analyze oxysterols are as varied as there are oxysterol molecular species. Most of the initial characterization of oxysterols was performed using TLC or Gas chromatography. More recently, HPLC ESI-MS/MS has emerged as an attractive means to analyze oxysterols, because there is no need to derivatize/protect the compounds prior to analysis as is needed for GC. Derivatization of oxysterols by the Girard reagent (1-(2-hydrazinyl-2-oxoethyl)pyridin-1-ium bromide) can improve the sensitivity and specificity by HPLC ESI-MS/MS analysis (Girard Reagent, Girard Reagent-d5). (For a description of the method, please see BBRC (2014) 446(3), 756-761.) But, it is also possible to measure oxysterol molecular species without any derivatization, depending on the sensitivity of your instrument. Below is an example chromatogram of OxysterolSPLASH analysis done here at Avanti.


If you are interested in having Avanti measure the oxysterol content in your samples, please visit the Avanti website and fill out a sample submission form. Alternatively, e-mail us at if you have any additional questions about this service or other Avanti products and services.

Contributed By: Paul RS Baker, PhD, Avanti Polar Lipids