Identification of Major Histocompatibility Complex-Regulated Body Odorants by Statistical Analysis of a Comparative Gas Chromatography/Mass Spectrometry Experiment

Abstract

This paper examines the application of gas chromatography/mass spectrometry (GC/MS) in a comparative experiment to identify volatile compounds from urine that differ in concentration between two groups of inbred mice. A complex mixture might comprise several hundred or even thousands of volatile compounds. Because their number and location in a chromatogram are generally unknown, and because components overlap in populous chromatograms, the statistical problems offer significant challenges beyond traditional two-group screening procedures. We describe a statistical procedure to compare two-dimensional GC/MS profiles between groups, which entails (1) signal processing, baseline correction, and peak detection in single ion chromatograms; (2) aligning chromatograms in time; (3) normalizing differences in overall signal intensities; and (4) detecting chromatographic regions that differ between groups. In an application to chemosignaling, we detect differences in GC/MS chromatograms of ether-extracted urine collected from two inbred groups of mice that differ only in genes of the major histocompatibility complex (MHC). Several dozen MHC-regulated compounds are found, including two known mouse pheromones, 2,5-dimethylpyrazine and 2-sec-butyl-4,5-dihydrothiazole.

In search of the chemical basis for MHC odourtypes

“…For example, phenylacetic acid was reported to be MHC-regulated in one study (Singer et al 1997), but not in a second study employing the same extraction method (Willse et al 2005). These results were not necessarily contradictory since, as Willse et al…”

“…Some of these fractionation techniques were modified and two separate chemical analyses of the urines from B6-H2 b and B6-H2 k mice were conducted in subsequent studies by our group (Singer et al 1997;Willse et al 2005). The extraction method was the same, but the extracted material in the earlier study was analysed by gas chromatography equipped with a flame ionization detector (GC/FID), whereas the later study analysed the extracted material by gas chromatography/mass spectrometry (GC/MS).…”

“…The Willse et al (2005) study employed a comprehensive statistical method to examine all detectable components in the GC/MS data. This 'metabolomic' approach permitted unbiased analyses of a broad spectrum of metabolites in the search for MHC-regulated compounds.…”

“…For example, Willse et al (2005Willse et al ( , 2006 and Kwak et al (2008) reported that the concentration of 2-sec-butyl-4,5-dihydrothiazole was higher in urines of B6-H2 k compared with B6 mice. However, Novotny et al (2007) reported the opposite pattern.…”

A Novel Approach for Nontargeted Data Analysis for Metabolomics. Large-Scale Profiling of Tomato Fruit Volatiles

“…The entire strategy of data analysis is universal for many kinds of mass spectral data and exceeds approaches of unbiased metabolomic data analysis in terms of resolution and comprehensiveness (Nielsen et al, 1998;Fraga et al, 2001;Johnson et al, 2003;Jonsson et al, 2004;Wiener et al, 2004;Willse et al, 2005). Also, it uses widely available software tools and simple basic statistical procedures, both of which make it useful for a wide range of studies in the fields of biochemistry, physiology, functional genomics, and systems biology.…”