Real-time monitoring of exhaled volatiles using atmospheric pressure chemical ionization on a compact mass spectrometer

© 2016 Future Science Ltd.Aim: Breath analyses have potential to detect early signs of disease onset. Ambient ionization allows direct combination of breath gases with MS for fast, on-line analysis. Portable MS systems would facilitate field/clinic-based breath analyses. Results & methodology: Volunteers ingested peppermint oil capsules and exhaled volatile compounds were monitored over 10 h using a compact mass spectrometer. A rise and fall in exhaled menthone was observed, peaking at 60-120 min. Real-time analysis showed a gradual rise in exhaled menthone postingestion. Sensitivity was comparable to established methods, with detection in the parts per trillion range. Conclusion: Breath volatiles were readily analyzed on a portable mass spectrometer through a simple inlet modification. Induced changes in exhaled profiles were detectable with high sensitivity and measurable in real-time

Saliva's unrevealed stories: characterisation of the human saliva volatilome in remote clinical settings by Thermal Desorption Gas Chromatography Mass Spectrometry

Saliva is a complex, rich matrix containing significant biochemical information, which can potentially be used to monitor an individual’s health, well-being and toxicological state. Saliva samples can be taken non-invasively and at low cost under challenging conditions such as heavily contaminated environments, for example from casualties in a Hot Zone of a CBRN incident, in primary care settings with no clinical infrastructure, from unconscious patients, as well as anxious patients or children. However, little is known about the volatile organic compound (VOC) biochemistry of human saliva and use of VOC saliva analysis in clinical settings has yet to be described. This research set up saliva sampling in two clinical centres for analysis with thermal desorption gas chromatography mass spectrometry to evaluate the saliva volatilome. Analytical workflows for saliva VOC analysis were developed and tested for clinical use. Recruitment included 16 participants (6 female and 10 male) in the age range 22 to 29 years who participated in a “Peppermint Test” that enabled preliminary comparisons between breath and saliva volatilome. The peppermint test also showed a well-defined eucalyptol washout, indicative of reliable and reproducible sampling and analysis. Clinical research assistants were trained to take VOC saliva samples following an end-to-end quality control and quality assurance protocol that included a proof of concept of digital global tracing system of samples. Forty-seven adults receiving treatment for acute pesticide poisoning (5 female and 42 male) in the age range 16 to 78 years were recruited at the Peradeniya Teaching Hospital Sri Lanka. Four samples were collected from each participant post treatment at 0, 1, 3 and 6 hr after recruitment. Samples were shipped to the Centre for Analytical Science Loughborough University Chemistry Department for analysis and subsequent data processing. A total of 131 samples were obtained over a period of 23 months. 38257 VOC features were isolated and were then classified into 1937 separate clusters out of which 1619 were present in only less than 20% of the samples. They were statistically sorted by VOCCluster based on similarity in retention index values and the intensities of the five most abundant fragment ions. 318 features were commonly found in a saliva matrix. 13 known siloxane products were detected from which 7 are by-products originating from the polydimethylsiloxane sampling media used for sample collection and 6 siloxane products were identified as ingredients of herbal medicine and cooking oil. The comparison of breath and saliva volatilome demonstrated similarities in biochemistry mechanisms between the two matrices from the behavioural trends of commonly endogenous VOCs. The replication of exhaled peppermint oil volatiles elimination behaviour in data from saliva samples supports the proposition that exposure to, and elimination of VOC insults may be monitored using saliva samples and that the Peppermint Test may be used to determine the efficacy of a saliva volatilome workflow. Targeted analysis from post-hoc samples confirmed that detectable levels of OP pesticides and their breakdown products were not recovered from the saliva samples as opposed to formulation solvents xylene and ethylbenzene that were detected and found to reliably indicate pesticide ingestion. A non-targeted (compound agnostic) multi-variate analysis workflow based on SIMCA-P consisting of data reduction with orthogonal partial least squares followed by principal component analysis and the proposal of a market score identified four candidate markers of pesticide ingestion, two of which were solvents identified through the targeted approach. Statistical scoring at 95% confidence intervals gave an AUROC of 0.959 with sensitivity of 0.714 and specificity of 1 suggesting a statistical viable model for identifying individuals exposed to pesticides. This research demonstrated the feasibility for saliva to be used as a diagnostic tool in primary care settings. To explicitly unfold the richness of the saliva matrix, further studies are required with the use of more sensitive mass analysers such as time of flight mass spectrometers to accurately identify features of interest. Saliva VOC sampling is a platform which can further be used for the investigation of ethanol and radiation toxicology and its development and validation may allow rapid triage in acute care settings.</p

Breath markers for therapeutic radiation

Radiation dose is important in radiotherapy. Too little, and the treatment is not effective, too much causes radiation toxicity. A biochemical measurement of the effect of radiotherapy would be useful in personalisation of this treatment. This study evaluated changes in exhaled breath volatile organic compounds (VOC) associated with radiotherapy with thermal desorption gas chromatography mass-spectrometry followed by data processing and multivariate statistical analysis. Further the feasibility of adopting gas chromatography ion mobility spectrometry for radiotherapy point-of-care breath was assessed. A total of 62 participants provided 240 end-tidal 1 dm3 breath samples before radiotherapy and at 1, 3, and 6 h post-exposure, that were analysed by thermal-desorption/gas-chromatography/quadrupole mass-spectrometry. Data were registered by retention-index and mass-spectra before multivariate statistical analyses identified candidate markers.A panel of sulfur containing compounds (thio-VOC) were observed to increase in concentration over the 6 h following irradiation. 3-methylthiophene (80 ng.m−3 to 790 ng.m−3) had the lowest abundance while 2-thiophenecarbaldehyde(380 ng.m−3 to 3.85 μg.m−3) the highest; note, exhaled 2-thiophenecarbaldehyde has not been observed previously. The putative tumour metabolite 2,4-dimethyl-1-heptene concentration reduced by an average of 73% over the same time. Statistical scoring based on the signal intensities thio-VOC and 3-methylthiophene appears to reflect individuals' responses to radiation exposure from radiotherapy. The thio-VOC are hypothesised to derive from glutathione and Maillard-based reactions and these are of interest as they are associated with radio-sensitivity. Further studies with continuous monitoring are needed to define the development of the breath biochemistry response to irradiation and to determine the optimum time to monitor breath for radiotherapy markers. Consequently, a single 0.5 cm3 breath-sample gas chromatography-ion mobility approach was evaluated. The calibrated limit of detection for 3-methylthiophene was 10 μg.m−3 with a lower limit of the detector's response estimated to be 210 fg.s−1; the potential for a point-of-care radiation exposure study exists.</div

Real-time monitoring of exhaled volatiles using atmospheric pressure chemical ionization on a compact mass spectrometer

AIM: Breath analyses have potential to detect early signs of disease onset. Ambient ionization allows direct combination of breath gases with MS for fast, on-line analysis. Portable MS systems would facilitate field/clinic-based breath analyses. Results & methodology: Volunteers ingested peppermint oil capsules and exhaled volatile compounds were monitored over 10 h using a compact mass spectrometer. A rise and fall in exhaled menthone was observed, peaking at 60-120 min. Real-time analysis showed a gradual rise in exhaled menthone postingestion. Sensitivity was comparable to established methods, with detection in the parts per trillion range. CONCLUSION: Breath volatiles were readily analyzed on a portable mass spectrometer through a simple inlet modification. Induced changes in exhaled profiles were detectable with high sensitivity and measurable in real-time

THE VARIABILITY OF VOLATILE ORGANIC COMPOUNDS IN CLINICAL ENVIRONMENTS

The development of clinical breath-analysis is confounded by the variability of background volatile organic compounds (VOC). Interpretation of clinical breath-data at individual, and cohort levels requires characterisation of clinical-VOC levels and exposures. Active-sampling with thermal-desorption/gas chromatography-mass spectrometry recorded and evaluated VOC concentrations in 245 samples of indoor air from three sites in a large NHS provider trust in the UK over 27 months. 7344 clinical VOC were isolated and 328 VOC and 68 were observed in more than 5% and 30% of samples respectively; associated with exogenous and endogenous sources. 17 VOC were seasonal differentiators. Metabolites from the anaesthetic sevoflurane, and putative-disease biomarkers in room air indicated that exhaled VOC were a source of background-pollution in clinical breath-tests. Apart from solvents, and PPE-waxes, exhaled VOC concentrations above 3 µgm-3 are unlikely to arise from room air contamination. This level could be applied as a threshold for inclusion in studies

The variability of volatile organic compounds in the indoor air of clinical environments

The development of clinical breath-analysis is confounded by the variability of background volatile organic compounds (VOC). Reliable interpretation of clinical breath-analysis at individual, and cohort levels requires characterisation of clinical-VOC levels and exposures. Active-sampling with thermal-desorption/gas chromatography-mass spectrometry recorded and evaluated VOC concentrations in 245 samples of indoor air from three sites in a large NHS provider trust in the UK over 27 months. Data deconvolution, alignment and clustering isolated 7344 features attributable to VOC and described the variability (composition and concentration) of respirable clinical VOC. 328 VOC were observed in more than 5% of the samples and 68 VOC appeared in more than 30% of samples. Common VOC were associated with exogenous and endogenous sources and 17 VOC were identified as seasonal differentiators. The presence of metabolites from the anaesthetic sevoflurane, and putative-disease biomarkers in room air, indicated that exhaled VOC were a source of background-pollution in clinical breath-testing activity. With the exception of solvents, and PPE waxes, exhaled VOC concentrations above 3 µg m-3 are unlikely to arise from room air contamination, and in the absence of extensive survey-data, this level could be applied as a threshold for inclusion in studies, removing a potential environmental confounding-factor in developing breath-based diagnostics

A benchmarking protocol for breath analysis: The peppermint experiment

International audienceSampling of volatile organic compounds (VOCs) has shown promise for detection of a range of diseases but results have proved hard to replicate due to a lack of standardization. In this work we introduce the 'Peppermint Initiative'. The initiative seeks to disseminate a standardized experiment that allows comparison of breath sampling and data analysis methods. Further, it seeks to share a set of benchmark values for the measurement of VOCs in breath. Pilot data are presented to illustrate the standardized approach to the interpretation of results obtained from the Peppermint experiment. This pilot study was conducted to determine the washout profile of peppermint compounds in breath, identify appropriate sampling time points, and formalise the data analysis. Five and ten participants were recruited to undertake a standardized intervention by ingesting a peppermint oil capsule that engenders a predictable and controlled change in the VOC profile in exhaled breath. After collecting a pre-ingestion breath sample, five further samples are taken at 2, 4, 6, 8, and 10 h after ingestion. Samples were analysed using ion mobility spectrometry coupled to multi-capillary column and thermal desorption gas chromatography mass spectrometry. A regression analysis of the washout data was used to determine sampling times for the final peppermint protocol, and the time for the compound measurement to return to baseline levels was selected as a benchmark value. A measure of the quality of the data generated from a given technique is proposed by comparing data fidelity. This study protocol has been used for all subsequent measurements by the Peppermint Consortium (16 partners from seven countries). So far 1200 breath samples from 200 participants using a range of sampling and analytical techniques have been collected. The data from the consortium will be disseminated in subsequent technical notes focussing on results from individual platforms