Thermal desorption comprehensive two-dimensional gas chromatography coupled to variable-energy electron ionization time-of-flight mass spectrometry for monitoring subtle changes in volatile organic compound profiles of human blood

Blood is a matrix with high potential for forensic investigations and human rescue. Its volatile signature can be used in search exercises to locate injured or deceased individuals. Little is known, however, about the volatile organic compound (VOC) profile of blood, except that it is complex and varies while blood ages. In the present study, we used thermal desorption (TD) and comprehensive two-dimensional gas chromatography (GCxGC) coupled to variable-energy electron ionization time-of-flight mass spectrometry (TOFMS) to monitor VOC signatures of human blood. A highly complex reference standard (Century Mix) containing 108 compounds of various chemical functionalities and several homologue series of compounds was used for the purpose of transposing our previously developed cryogenically modulated GCxGC-TOFMS methods into the use of a reverse fill/flush (RFF) flow modulator. The average peak width at half height was 340 ms and the average tailing factor was 1.16. Light VOCs (down to C4) were effectively flow modulated and exhibited minimal breakthrough over a large dynamic range spanning four orders of magnitude. Mass spectrometric detection was performed using electron impact ionization (EI) carried out at 70 eV and lower energies (12, 14, and 16 eV). The use of variable-energy (ve) EI allowed mass spectra to be produced with less fragmentation and an increased presence of structurally significant ions and the molecular ion. This provided additional confidence in peak assignments, especially for closely eluting isomers often observed in the profiling of the headspace of blood. Variable-energy EI TD-GCxGC-TOFMS blood data sets were statistically processed using principal component analyses (PCA) and hierarchical cluster analyses (HCA). These techniques demonstrated that the effect of aging was greater than the inter-individual variation on the blood VOC profile. The combination of retention indices, low and high EI MS spectra served as a strong basis to gain more confidence in analytical identification by excluding identities proposed by mass spectral databases (70 eV) for compounds contributing to the separation of blood of different ages.