The portable GC-MS system distinguished the treatments based on their detected volatile profiles. Additional statistical analysis identified five possible biomarker volatiles for the treatments, among them cyclosativene and copaene, which indicated damaged flower heads.
A simple method was developed for detection of Bacillus anthracis (BA) endospores and for differentiation of them from other species in the Bacillus cereus group. Chemical profiles that include lipids (i.e., fatty acids), carbohydrates (i.e., sugars), and the spore-specific biomarker, dipicolinic acid, were generated by one-step thermochemolysis (TCM) at 140 °C in 5 min to provide specific biomarker signatures. Anthrose, which is a biomarker characteristic of the B. cereus group of bacteria, was determined from a fragment produced by TCM. Surprisingly, several virulent BA strains contained very low levels of anthrose, which confounded their detection. A statistical discrimination algorithm was constructed using a combination of biomarkers, which was robust against different growth conditions (medium and temperature). Fifteen endospore-forming Bacillus species were confirmed in a statistically designed test (~90%) using the algorithm, including six BA strains (four virulent isolates), five B. thuringiensis (BT) isolates, and one isolate each for B. cereus (BC), B. mycoides (BM), B. atrophaeus (BG), and B. subtilis (BS). The detection limit for B. anthracis was found to be 50,000 endospores, on the basis of the GC/MS detection limits for 3-methyl-2-butenoic acid methyl ester, which is the biomarker derived from TCM of anthrose.
A simple method to detect and differentiate Bacillus anthracis (BA), Bacillus thuringiensis (BT), Bacillus atrophaeus (BG), and Bacillus cereus (BC) endospores using biomarker compounds, including dipicolinic acid methyl ester (DPAME) and fatty acid methyl esters (FAMEs), has been developed. The method is based on thermochemolysis methylation (TCM) of the endospores and gas chromatographymass spectrometry (GC-MS) of the reaction products. A suspension of the sample mixed with sulfuric acid (H 2 SO 4 ) and tetramethylammonium hydroxide (TMAH) in methanol (MeOH) at room temperature is sampled using a coiled wire filament (CWF) device, which consists of a tiny platinum helical wire coil attached to a retractable plunger that moves the coil in and out of a syringe needle housing. Sampling is accomplished by dipping the CWF in an endospore sample suspension, evaporating the suspension liquid, and then introducing the CWF into the injection port. While DPAME can be used for the general detection of endospores, specific saturated and unsaturated C 15 , C 16 , and C 17 fatty acid methyl esters provide additional information for differentiating various Bacillus species grown at different temperatures and in different media. DPAME could be detected in samples containing as few as 6000 endospores, and the GC-MS peak area percent reproducibility for FAMEs varied from 3 to 13% (RSD). Better than 97% correct predictability of Bacillus species identity was obtained from a blind experiment consisting of 145 samples.
A simple approach for preparing standard mixtures of volatile and semi-volatile organic compounds is reported. When placed in a closed container, standard mixture components partition between a polymeric material such as poly(dimethylsiloxane) (PDMS) and headspace to provide constant vapor concentrations. The granular form of heat-conditioned PDMS provides rapid equilibration with the headspace vapor and serves as a standard reservoir. Solid phase micro extraction (SPME) or gas-tight syringe can be used to deliver sample from the headspace to the analytical instrument. Quantitative calibration can be achieved with either active temperature control or by using a previously constructed look-up table. The effects of PDMS form and temperature on equilibrium distribution, initial equilibrium time, and re-equilibrium time after sampling were investigated. With respect to long term use and stability, analytes introduced onto 2.0 g of PDMS in a 7.4 mL vial were sampled more than 114 times during a test period of 43 days, giving chromatographic peak area %RSD values below 4.5% for all compounds. This device was designed to be solventless, quantitative, reproducible, environmentally friendly, and robust for routine evaluation and calibration of gas chromatography-mass spectrometry (GC-MS) systems.
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