An Ultrasonication-Assisted Extraction and Derivatization Protocol for GC/TOFMS-Based Metabolite Profiling By: Yumin Liu, Tianlu Chen, Yunpin Qiu, Yu Cheng, Yu Cao, Aihua Zhao, and Wei Jia Liu, Y.M., Chen, T.L., Qiu, Y.P., Cheng, Y., Cao, Y., Zhao, A.H., Jia, W. (2011). An ultrasonication-assisted extraction and deriatization protocol for GC/TOFMS-based metabolite profiling, Analytical and Bioanalytical Chemistry, 400(5), 1405-1417. ***Note: This version of the document is not the copy of record. Made available courtesy of Springer Verlag. The original publication is available at www.springerlink.com. Link to Article: http://www.springerlink.com/content/1q8353g821421380/ Abstract: Conventional chemical derivatization of metabolites in biological specimens is time-consuming, which limits the throughput and efficiency of metabolite profiling using a gas chromatography/time-of-flight mass spectrometry (GC/TOFMS) platform. We report an ultrasonication-assisted protocol which reduces the derivatization time from hours to about 30 min and significantly enhances the derivatization efficiency prior to a GC/TOFMS analysis. The protocol was evaluated using 40 compounds representing different classes of human metabolites, and demonstrated good analytical precision and accuracy. In comparison with the conventional method, the new protocol was able to increase the intensity of most of the identified peaks (71.0%) in the GC/TOFMS chromatograms of human serum samples. The detected compounds with increased intensity include most amino acids, keto-containing organic acids, carbonyl-containing carbohydrates, and unsaturated fatty acids. We applied this protocol in a metabolomic study of human serum samples obtained from 34 patients diagnosed with hypertension and 29 age- and gender-matched healthy subjects. Metabolite markers associated with hypertension, including glucosamine, D-sorbitol, 1-stearoylglycerol, and homocysteine, were identified and validated by statistical methods and use of reference standards. Our work highlights the potential of this novel approach for the large-scale metabolite profiling of samples generated from plant, animal, and clinical and epidemiological studies. Article: INTRODUCTION Metabonomics or metabolomics emerged following genomics, transcriptomics, and proteomics as an approach for capturing global biochemical alterations associated with pathophysiological changes [1–3]. To date, various analytical approaches, such as proton nuclear magnetic resonance, liquid chromatography/mass spectrometry (LC/MS), and gas chromatography/mass spectrometry (GC/MS), are employed to detect the variations of low molecular mass metabolites (less than 1,000 Da) in biological samples (e.g., urine, blood, tissue, cell, organs) [4–6]. LC/MS can analyze a wide range of chemical species that are not volatile and do not need chemical derivatization. As a result, LC/MS is best suited for the measurement of targeted metabolites that are not readily amenable to GC/MS analysis because of volatility issues. As one of the analytical