Physiological processes within the body may change emitted volatile organic compound (VOC) compos... more Physiological processes within the body may change emitted volatile organic compound (VOC) composition, and may therefore cause confounding biological background variability in breath gas analyses. To evaluate the effect of food intake on VOC concentration patterns in exhaled breath, this study assessed the variability of VOC concentrations due to food intake in a standardized caprine animal model. VOCs in (i) alveolar breath gas samples of nine clinically healthy goats and (ii) room air samples were collected and pre-concentrated before morning feeding and repeatedly after (+60 min, +150 min, +240 min) using needle trap microextraction (NTME). Analysis of VOCs was performed by gas chromatography and mass spectrometry (GC-MS). Only VOCs with significantly higher concentrations in breath gas samples compared to room air samples were taken into consideration. Six VOCs that belonged to the chemical classes of hydrocarbons and alcohols were identified presenting significantly different concentrations before and after feeding. Selected hydrocarbons showed a concentration pattern that was characterized by an initial increase 60 min after food intake, and a subsequent gradual decrease. Results emphasize consideration of physiological effects on exhaled VOC concentrations due to food intake with respect to standardized protocols of sample collection and critical evaluation of results.
Physiological effects may change volatile organic compound (VOC) concentrations and may therefore... more Physiological effects may change volatile organic compound (VOC) concentrations and may therefore act as confounding factors in the definition of VOCs as disease biomarkers. To evaluate the extent of physiological background variability, this study assessed the effects of feed composition and somatic growth on VOC patterns in a standardized large animal model. Fifteen clinically healthy goats were followed during their first year of life. VOCs present in the headspace over faeces, exhaled breath and ambient air inside the stable were repeatedly assessed in parallel with the concentrations of glucose, protein, and albumin in venous blood. VOCs were collected and analysed using solid-phase or needle-trap microextraction and gas chromatograpy together with mass spectroscopy. The concentrations of VOCs in exhaled breath and above faeces varied significantly with increasing age of the animals. The largest variations in volatiles detected in the headspace over faeces occurred with the cha...
Concentrations of exhaled volatile organic compounds (VOCs) may depend not only on biochemical or... more Concentrations of exhaled volatile organic compounds (VOCs) may depend not only on biochemical or pathologic processes but also on physiological parameters. As breath sampling may be done in different body positions, effects of the sampling position on exhaled VOC concentrations were investigated by means of real-time mass spectrometry.Breaths from 15 healthy volunteers were analyzed in real-time by PTR-ToF-MS-8000 during paced breathing (12/min) in a continuous side-stream mode. We applied two series of body positions (setup 1: sitting, standing, supine, and sitting; setup 2: supine, left lateral, right lateral, prone, and supine). Each position was held for 2 min. Breath VOCs were quantified in inspired and alveolar air by means of a custom-made algorithm. Parallel monitoring of hemodynamics and capnometry was performed noninvasively.In setup 1, when compared to the initial sitting position, normalized mean concentrations of isoprene, furan, and acetonitrile decreased by 24%, 26%,...
ABSTRACT Analysis of volatile organic compounds (VOCs) in breath holds great promise as a novel n... more ABSTRACT Analysis of volatile organic compounds (VOCs) in breath holds great promise as a novel non-invasive diagnostic tool. Clinical application, however, is hampered by problems arising from actual or recent uptake of exogenous substances, especially in the clinical environment, and from quickly and abruptly changing VOC profiles. Real time measurements can help to address these issues. Therefore, a PTR-ToF-MS was adapted for clinical breath analysis. For reasons of medical safety, the PTR-ToF-MS instrument could not be positioned at the bedside. Hence, a 6m long heated silcosteel transfer line connected to a sterile mouthpiece was used for breath sampling. Breath from mechanically ventilated patients was measured continuously for one hour directly after cardiac surgery. Room air and exhaled air from 32 volunteers was analyzed in the post anesthetic care unit (PACU). In addition, ambient air in teh PACU was analyzed continuously over 7 days. Breath resolved measurements in mechanically ventilated patients and spontaneously breathing volunteers were possible using a time resolution of 200 ms. Detection limits were in the high pptV / low ppbV range (e.g. 0.4 ppbV for sevoflurane). Changes of ventilation-perfusion-ratio, cardiac output and application of intravenous drugs were mirrored by VOC concentrations (e.g. isoprene) in the breath of mechanically ventilated patients. Exhaled concentrations of sevoflurane in the PACU strongly depended on background concentrations and a good correlation with room air concentrations was observed (R2=0.94). Endogenous acetone in contrast did not show such a dependency. PTR-ToF-MS adapted to clinical conditions is a powerful tool to address fundamental as well as practical questions in clinical breath analysis.
Chemical analysis of VOCs in human breath requires sensitivities on the pptV – ppbV level to dete... more Chemical analysis of VOCs in human breath requires sensitivities on the pptV – ppbV level to detect relevant biomarkers. For unequivocal identification of volatile substances mass spectrometric methods are necessary. Analytical procedures like GC-MS, however, require complex sampling and sample preparation procedures to provide sufficient sensitivity. Existing techniques such as SPE or SPME face limitations regarding sampling time and sensitivity. To overcome these limitations a sensitive, clinically applicable and universal sampling- and preconcentration method is desirable. Needle Trap Devices (NTDs) have the potential to bridge this gap and serve as an interface between the patient and highly sophisticated analytical techniques, such as GC-MS or GC-GC-TOF-MS, as well as between the patient and point-of-care applicable methods like sensor systems. Since preconcentration and sampling is carried out in one concerted step, sample preparation is simplified and fewer potential error so...
NTME is a promising new tool for sample preparation. Needle-Trap-Devices (NTDs) have shown many a... more NTME is a promising new tool for sample preparation. Needle-Trap-Devices (NTDs) have shown many advantages such as improved detection limits, reduced sampling time and volume, improved stability and reproducibility if compared to other techniques used in breath analysis such as SPE and SPME. To fully explore the potential NTME effects of sampling flow (2-40 ml/ min), volume (5-60 ml) and humidity in dry gas standards and (humid bzw water saturated) breath samples was investigated for NTDs with (single phase) polymer packing and (triple phase) combinations of DVB/Carbopack X/Carboxen. An automated CO2 controlled sampling device for direct alveolar sampling at the point-of-care was developed. Triple phase NTDs showed only small dependencies on sampling flow rate and low breakthrough from 1-5 % for the investigated substances, including hydrocarbons, aldehydes and aromatics, when sample volumes up to 60 mL were applied. RSDs were between 1 (o-Xylene) and 17 % (Acetone) for DVB triple b...
A crucial issue in breath research is the impact of environmental compounds on breath profiles. A... more A crucial issue in breath research is the impact of environmental compounds on breath profiles. Among more than 1000 different compounds detected in human breath only few are known to be originating from endogenous sources. As most studies are performed in a clinical environment typical contaminations such as disinfectants, anesthetics or emission from plastic materials have to be taken into account. The aim of this study was to investigate the effect of exogenous contaminantsfromtheclinicalenvironmentontobreathprofilesofhospitalstaff. 15 physicians, 10 PACU nurses and 7 staff members not working in the OR/PACU. Exhaled breathofallparticipantswas analyzedbymeansofPTR-TOFinthecentralPACUoftheRostock University Medical Center. Samples were collected before and after working shifts of the volunteers.Inparallel,inspiredandroomairconcentrationsweredetermined. Breath profiles from physicians and nurses showed distinct concentrations of anesthetics and disinfectants after the end of their ...
Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of a chronic enteric disea... more Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of a chronic enteric disease of ruminants. Available diagnostic tests are complex and slow. In vitro, volatile organic compound (VOC) patterns emitted from MAP cultures mirrored bacterial growth and enabled distinction of different strains. This study was intended to determine VOCs in vivo in the controlled setting of an animal model. VOCs were pre-concentrated from breath and feces of 42 goats (16 controls and 26 MAP-inoculated animals) by means of needle trap microextraction (breath) and solid phase microextraction (feces) and analyzed by gas chromatography/ mass spectrometry. Analyses were performed 18, 29, 33, 41 and 48 weeks after inoculation. MAP-specific antibodies and MAP-specific interferon-γ-response were determined from blood. Identities of all marker-VOCs were confirmed through analysis of pure reference substances. Based on detection limits in the high pptV and linear ranges of two orders of magnitud...
Bulky and hyphenated laboratory-based analytical instrumentation such as gas chromatography/mass ... more Bulky and hyphenated laboratory-based analytical instrumentation such as gas chromatography/mass spectrometry is still required to trace breath biomarkers in the low ppbV level. Innovative sensor-based technologies could provide on-site and point-of-care (POC) detection of volatile biomarkers such as breath aldehydes related to oxidative stress and cancer.An electrochemical sensor system was developed for direct detection of the total abundance of aldehydes in exhaled breath in the ppbV level and for simultaneous determination of the airway inflammation markers carbon monoxide (CO) and nitric oxide (NO). The sensor system was tested in vitro with gaseous standard mixtures and in vivo in spontaneously breathing patients and under mechanical ventilation in an animal model.The sensor system provided in vitro and in vivo detection of trace levels of aldehydes, CO and NO. Inertness of the tubing system was important for reliable results. Sensitivity of the aldehyde sensor increased with ...
Rapid concentration changes due to physiological or pathophysiological effects rather than appear... more Rapid concentration changes due to physiological or pathophysiological effects rather than appearance of unique disease biomarkers are important for clinical application of breath research. Simple maneuvers such as breath holding may significantly affect breath biomarker concentrations. In this study, exhaled volatile organic compound (VOC) concentrations were assessed in real time before and after different breath holding maneuvers. Continuous breath-resolved measurements (PTR-ToF-MS-8000) were performed in 31 healthy human subjects in a side-stream sampling mode. After 1 min of tidal breathing participants held their breath for 10, 20, 40, 60 s and as long as possible. Afterwards they continued to breathe normally for another minute. VOC profiles could be monitored in real time by assigning online PTR-ToF-MS data to alveolar or inspired phases of breath. Sudden and profound changes of exhaled VOC concentrations were recorded after different breath holding maneuvers. VOC concentrations returned to base line levels 10–20 s after breath holding. Breath holding induced concentration changes depended on physico-chemical properties of the substances. When substance concentrations were normalized onto end-tidal CO2 content, variation of acetone concentrations decreased, whereas variations of isoprene concentrations were not affected. As the effects of breathing patterns on exhaled substance concentrations depend on individual substance properties, sampling procedures have to be validated for each compound by means of appropriate real-time analysis. Normalization of exhaled concentrations onto exhaled CO2 is only valid for substances having similar physico-chemical properties as CO2.
Physiological processes within the body may change emitted volatile organic compound (VOC) compos... more Physiological processes within the body may change emitted volatile organic compound (VOC) composition, and may therefore cause confounding biological background variability in breath gas analyses. To evaluate the effect of food intake on VOC concentration patterns in exhaled breath, this study assessed the variability of VOC concentrations due to food intake in a standardized caprine animal model. VOCs in (i) alveolar breath gas samples of nine clinically healthy goats and (ii) room air samples were collected and pre-concentrated before morning feeding and repeatedly after (+60 min, +150 min, +240 min) using needle trap microextraction (NTME). Analysis of VOCs was performed by gas chromatography and mass spectrometry (GC-MS). Only VOCs with significantly higher concentrations in breath gas samples compared to room air samples were taken into consideration. Six VOCs that belonged to the chemical classes of hydrocarbons and alcohols were identified presenting significantly different concentrations before and after feeding. Selected hydrocarbons showed a concentration pattern that was characterized by an initial increase 60 min after food intake, and a subsequent gradual decrease. Results emphasize consideration of physiological effects on exhaled VOC concentrations due to food intake with respect to standardized protocols of sample collection and critical evaluation of results.
Physiological effects may change volatile organic compound (VOC) concentrations and may therefore... more Physiological effects may change volatile organic compound (VOC) concentrations and may therefore act as confounding factors in the definition of VOCs as disease biomarkers. To evaluate the extent of physiological background variability, this study assessed the effects of feed composition and somatic growth on VOC patterns in a standardized large animal model. Fifteen clinically healthy goats were followed during their first year of life. VOCs present in the headspace over faeces, exhaled breath and ambient air inside the stable were repeatedly assessed in parallel with the concentrations of glucose, protein, and albumin in venous blood. VOCs were collected and analysed using solid-phase or needle-trap microextraction and gas chromatograpy together with mass spectroscopy. The concentrations of VOCs in exhaled breath and above faeces varied significantly with increasing age of the animals. The largest variations in volatiles detected in the headspace over faeces occurred with the cha...
Concentrations of exhaled volatile organic compounds (VOCs) may depend not only on biochemical or... more Concentrations of exhaled volatile organic compounds (VOCs) may depend not only on biochemical or pathologic processes but also on physiological parameters. As breath sampling may be done in different body positions, effects of the sampling position on exhaled VOC concentrations were investigated by means of real-time mass spectrometry.Breaths from 15 healthy volunteers were analyzed in real-time by PTR-ToF-MS-8000 during paced breathing (12/min) in a continuous side-stream mode. We applied two series of body positions (setup 1: sitting, standing, supine, and sitting; setup 2: supine, left lateral, right lateral, prone, and supine). Each position was held for 2 min. Breath VOCs were quantified in inspired and alveolar air by means of a custom-made algorithm. Parallel monitoring of hemodynamics and capnometry was performed noninvasively.In setup 1, when compared to the initial sitting position, normalized mean concentrations of isoprene, furan, and acetonitrile decreased by 24%, 26%,...
ABSTRACT Analysis of volatile organic compounds (VOCs) in breath holds great promise as a novel n... more ABSTRACT Analysis of volatile organic compounds (VOCs) in breath holds great promise as a novel non-invasive diagnostic tool. Clinical application, however, is hampered by problems arising from actual or recent uptake of exogenous substances, especially in the clinical environment, and from quickly and abruptly changing VOC profiles. Real time measurements can help to address these issues. Therefore, a PTR-ToF-MS was adapted for clinical breath analysis. For reasons of medical safety, the PTR-ToF-MS instrument could not be positioned at the bedside. Hence, a 6m long heated silcosteel transfer line connected to a sterile mouthpiece was used for breath sampling. Breath from mechanically ventilated patients was measured continuously for one hour directly after cardiac surgery. Room air and exhaled air from 32 volunteers was analyzed in the post anesthetic care unit (PACU). In addition, ambient air in teh PACU was analyzed continuously over 7 days. Breath resolved measurements in mechanically ventilated patients and spontaneously breathing volunteers were possible using a time resolution of 200 ms. Detection limits were in the high pptV / low ppbV range (e.g. 0.4 ppbV for sevoflurane). Changes of ventilation-perfusion-ratio, cardiac output and application of intravenous drugs were mirrored by VOC concentrations (e.g. isoprene) in the breath of mechanically ventilated patients. Exhaled concentrations of sevoflurane in the PACU strongly depended on background concentrations and a good correlation with room air concentrations was observed (R2=0.94). Endogenous acetone in contrast did not show such a dependency. PTR-ToF-MS adapted to clinical conditions is a powerful tool to address fundamental as well as practical questions in clinical breath analysis.
Chemical analysis of VOCs in human breath requires sensitivities on the pptV – ppbV level to dete... more Chemical analysis of VOCs in human breath requires sensitivities on the pptV – ppbV level to detect relevant biomarkers. For unequivocal identification of volatile substances mass spectrometric methods are necessary. Analytical procedures like GC-MS, however, require complex sampling and sample preparation procedures to provide sufficient sensitivity. Existing techniques such as SPE or SPME face limitations regarding sampling time and sensitivity. To overcome these limitations a sensitive, clinically applicable and universal sampling- and preconcentration method is desirable. Needle Trap Devices (NTDs) have the potential to bridge this gap and serve as an interface between the patient and highly sophisticated analytical techniques, such as GC-MS or GC-GC-TOF-MS, as well as between the patient and point-of-care applicable methods like sensor systems. Since preconcentration and sampling is carried out in one concerted step, sample preparation is simplified and fewer potential error so...
NTME is a promising new tool for sample preparation. Needle-Trap-Devices (NTDs) have shown many a... more NTME is a promising new tool for sample preparation. Needle-Trap-Devices (NTDs) have shown many advantages such as improved detection limits, reduced sampling time and volume, improved stability and reproducibility if compared to other techniques used in breath analysis such as SPE and SPME. To fully explore the potential NTME effects of sampling flow (2-40 ml/ min), volume (5-60 ml) and humidity in dry gas standards and (humid bzw water saturated) breath samples was investigated for NTDs with (single phase) polymer packing and (triple phase) combinations of DVB/Carbopack X/Carboxen. An automated CO2 controlled sampling device for direct alveolar sampling at the point-of-care was developed. Triple phase NTDs showed only small dependencies on sampling flow rate and low breakthrough from 1-5 % for the investigated substances, including hydrocarbons, aldehydes and aromatics, when sample volumes up to 60 mL were applied. RSDs were between 1 (o-Xylene) and 17 % (Acetone) for DVB triple b...
A crucial issue in breath research is the impact of environmental compounds on breath profiles. A... more A crucial issue in breath research is the impact of environmental compounds on breath profiles. Among more than 1000 different compounds detected in human breath only few are known to be originating from endogenous sources. As most studies are performed in a clinical environment typical contaminations such as disinfectants, anesthetics or emission from plastic materials have to be taken into account. The aim of this study was to investigate the effect of exogenous contaminantsfromtheclinicalenvironmentontobreathprofilesofhospitalstaff. 15 physicians, 10 PACU nurses and 7 staff members not working in the OR/PACU. Exhaled breathofallparticipantswas analyzedbymeansofPTR-TOFinthecentralPACUoftheRostock University Medical Center. Samples were collected before and after working shifts of the volunteers.Inparallel,inspiredandroomairconcentrationsweredetermined. Breath profiles from physicians and nurses showed distinct concentrations of anesthetics and disinfectants after the end of their ...
Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of a chronic enteric disea... more Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of a chronic enteric disease of ruminants. Available diagnostic tests are complex and slow. In vitro, volatile organic compound (VOC) patterns emitted from MAP cultures mirrored bacterial growth and enabled distinction of different strains. This study was intended to determine VOCs in vivo in the controlled setting of an animal model. VOCs were pre-concentrated from breath and feces of 42 goats (16 controls and 26 MAP-inoculated animals) by means of needle trap microextraction (breath) and solid phase microextraction (feces) and analyzed by gas chromatography/ mass spectrometry. Analyses were performed 18, 29, 33, 41 and 48 weeks after inoculation. MAP-specific antibodies and MAP-specific interferon-γ-response were determined from blood. Identities of all marker-VOCs were confirmed through analysis of pure reference substances. Based on detection limits in the high pptV and linear ranges of two orders of magnitud...
Bulky and hyphenated laboratory-based analytical instrumentation such as gas chromatography/mass ... more Bulky and hyphenated laboratory-based analytical instrumentation such as gas chromatography/mass spectrometry is still required to trace breath biomarkers in the low ppbV level. Innovative sensor-based technologies could provide on-site and point-of-care (POC) detection of volatile biomarkers such as breath aldehydes related to oxidative stress and cancer.An electrochemical sensor system was developed for direct detection of the total abundance of aldehydes in exhaled breath in the ppbV level and for simultaneous determination of the airway inflammation markers carbon monoxide (CO) and nitric oxide (NO). The sensor system was tested in vitro with gaseous standard mixtures and in vivo in spontaneously breathing patients and under mechanical ventilation in an animal model.The sensor system provided in vitro and in vivo detection of trace levels of aldehydes, CO and NO. Inertness of the tubing system was important for reliable results. Sensitivity of the aldehyde sensor increased with ...
Rapid concentration changes due to physiological or pathophysiological effects rather than appear... more Rapid concentration changes due to physiological or pathophysiological effects rather than appearance of unique disease biomarkers are important for clinical application of breath research. Simple maneuvers such as breath holding may significantly affect breath biomarker concentrations. In this study, exhaled volatile organic compound (VOC) concentrations were assessed in real time before and after different breath holding maneuvers. Continuous breath-resolved measurements (PTR-ToF-MS-8000) were performed in 31 healthy human subjects in a side-stream sampling mode. After 1 min of tidal breathing participants held their breath for 10, 20, 40, 60 s and as long as possible. Afterwards they continued to breathe normally for another minute. VOC profiles could be monitored in real time by assigning online PTR-ToF-MS data to alveolar or inspired phases of breath. Sudden and profound changes of exhaled VOC concentrations were recorded after different breath holding maneuvers. VOC concentrations returned to base line levels 10–20 s after breath holding. Breath holding induced concentration changes depended on physico-chemical properties of the substances. When substance concentrations were normalized onto end-tidal CO2 content, variation of acetone concentrations decreased, whereas variations of isoprene concentrations were not affected. As the effects of breathing patterns on exhaled substance concentrations depend on individual substance properties, sampling procedures have to be validated for each compound by means of appropriate real-time analysis. Normalization of exhaled concentrations onto exhaled CO2 is only valid for substances having similar physico-chemical properties as CO2.
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