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ISOLUTE® FL Used in Tuberculosis Research

25 February 2015

A proof of principle study presents a method aiming to improve the analysis of methyl mycocerosate tuberculosis biomarkers from sputum using ISOLUTE® FL sample preparation columns. 

 

The purification procedures involving solid phase extraction (SPE) based methods with both commercial Isolute-Florisil cartridges, and purpose designed molecularly imprinted polymeric materials (MIPs), resulted in cleaner chromatograms, while the mycocerosates are still present. The clean-up performed on solutions of phthiocerol dimycocerosates (PDIMs) and cholesterol standards in petroleum ether show recovery of PDIMs between 64–70%, whilst most of the cholesterol is removed from the system. When applied to petroleum ether extracts from representative sputum samples, the clean-up procedures resulted in recoveries of 36 - 68% for PDIMs, allowing some superior detection of the target analytes.

Methyl mycocerosates are generated from the transesterification of PDIMs, extracted in petroleum ether from sputum of tuberculosis suspect patients. When a high matrix background is present in the sputum extracts, the identification of the chromatographic peaks corresponding to the methyl derivatives of PDIMs analytes may be hindered by the closely eluting methyl ether of cholesterol, usually an abundant matrix constituent frequently present in sputum samples.  

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Simona C. Nicoaraa, Nicholas W. Turnerb, David E. Minnikinc, Oona Y-C. Leec, Denise M. O'Sullivand, Ruth McNerneyd, Reggie Mutetwae, Liz E. Corbette, Geraint H. Morgana. 2015. Development of Sample Clean Up Methods for the Analysis of Mycobacterium tuberculosis Methyl Mycocerosate Biomarkers in Sputum Extracts by Gas Chromatography–Mass Spectrometry. Journal of Chromatography B