Background: Noninvasive detection of innate immune function such as the accumulation of neutrophils remains a challenge in many areas of clinical medicine. We hypothesized that granulocytes could generate volatile organic compounds.

Background The gaseous headspace above naïve Escherichia Coli (E. coli) cultures and whole human blood inoculated with E. coli were collected and analyzed for the presence of trace gases that may have the potential to be used as novel, non‐invasive markers of infectious disease. Methods The naïve E. coli culture, LB broth, and human whole blood or E. coli inoculated whole blood were incubated in hermetically sealable glass bioreactors at 37°C for 24 hrs. LB broth and whole human blood were used as controls for background volatile organic compounds (VOCs). The headspace gases were collected after incubation and analyzed using a gas chromatographic system with multiple column/detector combinations. Results Six VOCs were observed to be produced by E. coli‐infected whole blood while there existed nearly zero to relatively negligible amounts of these gases in the whole blood alone, LB broth, or E. coli‐ inoculated LB broth. These VOCs included dimethyl sulfide (DMS), carbon disulfide (CS2), ethanol, acetaldehyde, methyl butanoate, and an unidentified gas S. In contrast, there were several VOCs significantly elevated in the headspace above the E. coli in LB broth, but not present in the E. coli/blood mixture. These VOCs included dimethyl disulfide (DMDS), dimethyl trisulfide (DMTS), methyl propanoate, 1‐propanol, methylcyclohexane, and unidentified gases R2 and Q. Conclusions This study demonstrates 1) that cultivated E. coli in LB broth produce distinct gas profiles, 2) for the first time, the ability to modify E. coli‐specific gas profiles by the addition of whole human blood, and 3) that E. coli‐human whole blood interactions present different gas emission profiles that have the potential to be used as non‐invasive volatile biomarkers of E. coli infection.

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Rationale. The presence of leukocytes in airways indicates substantial inflammation. We wondered whether volatile organic compounds produced by these cells could be detected and ultimately used as noninvasive biomarkers. We recently demonstrated that distinct gases were produced by transformed promyelocitic cells (J. transl Med, 2009 7:31). We hypothesized that neutrophils and peripheral blood mononuclear cells (PBMCs) could generate cell-type specific gases which could be detected even in the parts-per-billion and trillion (ppbv, pptv) range using appropriate technologies.Methods. This study was approved by the UCI IRB. Neutrophils and PBMCs were isolated from the peripheral blood of healthy donors (n=10, 18-65 yrs old). The cells were resuspended in RPMI 1640 supplemented with 10% FCS. The vials containing cell suspensions (30 x 10 cells/10mL) were placed inside bioreactors (that we specially designed for this purpose) in an incubator at 37°C for 24 h. The headspace 6 gas was then collected and analyzed using a GC-MS system built originally to detect trace volatile organic compounds in the atmosphere. We performed 10 separate experiments and compared target gases between media alone and cell culture conditions using paired t-tests.Results. In the cultured neutrophils, acetaldehyde in the headspace was significantly elevated (220±140 (SD) ppbv) compared with media (90±20 ppbv, p=0.024). In contrast, the level of acetaldehyde from PBMCs was 30±20 ppbv, significantly below media alone (p<0.001) In . cultured PBMCs, we found 2 unidentified gases (A and B, 260±110 pptv and 80±70 pptv) greater than media alone (p<0.002). For both neutrophils and PBMCs hexanaldehyde (1±1 ppbv, 0.6±0.2 ppbv, respectively) was below medium alone (4±2 ppbv).Conclusions. Human primary immune cells produce volatile gases that can be measured in trace amounts. This study demonstrates in vitro for the first time the ability to detect basal levels of acetaldehyde produced by unperturbed cultured neutrophils. Moreover, the data suggest that different leukocyte subtypes have different gas profiles (e.g., no acetaldehyde was found in PBMCs). Intriguingly, the data suggest that some gases such as acetaldehyde in PBMCs and hexanaldehyde in both neutrophils and PBMCs appear to be metabolized by the cultured cells. Finally, we measured several as yet unidentified gases produced by leukocytes. Detection of key gases in human breath may eventually reveal the extent and type of pulmonary leukocyte infiltration. This abstract is funded by: National Institutes of Health (R01-HL-080947 to D.M.C); and the Physical Sciences Dean's Innovation fund (D.R. B.) Am J Respir Crit Care Med

Background: Noninvasive detection of innate immune function such as the accumulation of neutrophils remains a challenge in many areas of clinical medicine. We hypothesized that granulocytes could generate volatile organic compounds. Methods:To begin to test this, we developed a bioreactor and analytical GC-MS system to accurately identify and quantify gases in trace concentrations (parts per billion) emitted solely from cell/media culture. A human promyelocytic leukemia cell line, HL60, frequently used to assess neutrophil function, was grown in serum-free medium.Results: HL60 cells released acetaldehyde and hexanaldehyde in a time-dependent manner. The mean ± SD concentration of acetaldehyde in the headspace above the cultured cells following 4-, 24-and 48-h incubation was 157 ± 13 ppbv, 490 ± 99 ppbv, 698 ± 87 ppbv. For hexanaldehyde these values were 1 ± 0.3 ppbv, 8 ± 2 ppbv, and 11 ± 2 ppbv. In addition, our experimental system permitted us to identify confounding trace gas contaminants such as styrene. Conclusion:This study demonstrates that human immune cells known to mimic the function of innate immune cells, like neutrophils, produce volatile gases that can be measured in vitro in trace amounts.