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Physics Department PHYSICS DEPARTMENT Investigations on the use of breath gas analysis with Proton Transfer Reaction Mass Spectrometry (PTR-MS) for a non-invasive method of early lung cancer detection Dissertation of Bhushan Thekedar TECHNISCHE UNIVERSITÄT MÜNCHEN Technische Universität München Physik-Department Investigations on the use of breath gas analysis with Proton Transfer Reaction Mass Spectrometry (PTR-MS) for a non-invasive method of early lung cancer detection Bhushan Thekedar Vollständiger Abdruck der von der Fakultät für Physik der Technischen Universität München zur Erlangung des akademischen Grades eines Doktors der Naturwissenschaften genehmigten Dissertation. Vorsitzender: Univ.-Prof. Dr. R. Netz Prüfer der Dissertation: 1. Hon.-Prof. Dr. H.G. Paretzke 2. Univ.-Prof. Dr. T. Hugel Die Dissertation wurde am 16.11.09 bei der Technischen Universität München eingereicht und durch die Fakultät für Physik am 14.12.09 angenommen. Author contact information: Bhushan Thekedar Institute of Radiation Protection Helmholtz Zentrum München German Research Center for Environmental Health (GmbH) Ingolstaedter Landstrasse 1 85764 Neuherberg Germany Email: [email protected] Contents 1 Introduction.................................................................................................................................... 1 2 Theoretical background .................................................................................................................. 6 2.1 Volatile organic compounds (VOCs)...................................................................................... 6 2.2 Proton transfer reaction mass spectrometry (PTR-MS) ........................................................... 6 2.2.1 Ion-molecule reactions ....................................................................................................... 7 2.2.2 Mass separation and ion detection .....................................................................................16 2.2.3 Equation of concentration of VOCs measured by PTR-MS................................................22 2.3 Measurement statistics ..........................................................................................................24 2.4 Receiver-operator characteristics (ROC) curves.....................................................................28 3 Material and Methods....................................................................................................................31 3.1 Sample analysis and data evaluation......................................................................................31 3.1.1 Breath gas sampling..........................................................................................................31 3.1.2 Measurement of samples with PTR-MS ............................................................................31 3.1.3 Normalization of counts rates and transmission measurement............................................33 3.1.4 Tentative determination of the chemical identity of the VOCs ...........................................39 3.1.5 Identification of endogenous VOCs...................................................................................40 3.2 Variability in measurements ..................................................................................................43 3.2.1 Variability due to the PTR-MS instrument.........................................................................43 3.2.2 Variability due to intra individual differences of the volunteers .........................................47 3.2.3 Variability due to inter individual differences of the volunteers .........................................52 3.2.4 Variability due to room air influences................................................................................53 3.3 Influences of sampling specific parameters on VOC concentrations.......................................54 3.3.1 Temperature and humidity of surrounding air....................................................................54 3.3.2 Exhalation with breath holding..........................................................................................54 3.3.3 Velocity of exhalation.......................................................................................................54 3.3.4 Volume of exhalation........................................................................................................54 3.3.5 Multiple exhalations..........................................................................................................55 3.3.6 Volume of inhalation ........................................................................................................55 3.3.7 Breath sampling with isothermal re-breathing....................................................................55 3.3.8 Statistical evaluation .........................................................................................................58 3.4 Influences of volunteer specific parameters on VOC concentrations ......................................59 3.5 Investigations on breath gas VOCs for detection of lung cancer.............................................60 3.5.1 Classification of human subjects .......................................................................................60 3.5.2 Monitoring study of breath gas VOCs of lung cancer patients during therapy ....................63 3.5.3 Statistical evaluation .........................................................................................................63 4 Results and Discussions.................................................................................................................66 4.1 Variability in measurements ..................................................................................................66 4.1.1 Variability due to the PTR-MS instrument.........................................................................66 4.1.2 Variability due to intra individual differences of the volunteers .........................................67 4.1.3 Variability due to inter individual differences of the volunteers .........................................73 4.1.4 Variability due to room air influences................................................................................74 4.2 Influences of sampling specific parameters on VOC concentrations in breath gas ..................77 4.2.1 Temperature and humidity of the environmental air...........................................................77 4.2.2 Exhalation with breath holding..........................................................................................79 4.2.3 Velocity of exhalation.......................................................................................................80 4.2.4 Volume of exhalation........................................................................................................81 4.2.5 Multiple exhalations..........................................................................................................83 4.2.6 Volume of inhalation ........................................................................................................84 4.2.7 Comparison of isothermal rebreath with single exhalation .................................................84 4.3 Influences of volunteer specific parameters on VOC concentrations in breath gas..................86 4.3.1 Gender..............................................................................................................................86 4.3.2 Fasting and feeding ...........................................................................................................87 4.3.3 Age and Body mass index (BMI) ......................................................................................89 4.4 Investigations on breath gas VOCs for detection of lung cancer.............................................92 4.4.1 General comparison of single VOCs from patients, controls and surrounding air ...............92 4.4.2 Stage-wise comparison of single VOCs between patients and controls.............................105 4.4.3 Monitoring study of lung cancer biomarkers during therapy ............................................107 4.4.4 Multivariate statistical analysis........................................................................................109 5 Summary.....................................................................................................................................118 5.1 Variability in measurement of breath gas analysis................................................................118 5.2 Influences of sampling specific parameters on VOC concentrations.....................................118 5.3 Influences on volunteer specific parameters on VOC concentrations....................................119 5.4 On the identification of lung cancer biomarkers...................................................................120 6 Conclusions.................................................................................................................................122 7 Acknowledgements .....................................................................................................................124 8 References...................................................................................................................................126 Appendix. Proton affinities and reaction rate constants of common VOCs………………...……….138 Abbreviations VOCs = Volatile organic compounds KEcm = Centre-of-mass
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