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					Oral Presentation                                                                                                    Room Europa 2 - 08:30-10:30

                                                  M ONDAY, O CTOBER 6 TH 2008

                                                                                  189. New frontiers in noninvasive
                                                                                       assessment of the lungs

                                                                        Metabonomic analysis of EBC in adults by nuclear magnetic resonance
                                                                        Guglielmo de Laurentiis 1,2 , Debora Paris 3 , Dominique Melck 3 ,
                                                                        Mauro Maniscalco 1 , Serafino Marsico 2 , Gaetano Corso 4 , Andrea Motta 3 ,
                                                                        Matteo Sofia 1 . 1 Respiratory Diseases, University “Federico II”, Naples, Italy;
                                                                          Respiratory Diseases, Second University of Naples, Naples, Italy; 3 Institute of
                                                                        Biomolecular Chemistry, Consiglio Nazionale delle Ricerche, Pozzuoli, Naples,
                                                                        Italy; 4 Biomedical Sciences, University of Foggia, Foggia, Italy

                                                                        Exhaled breath condensate (EBC) is a non-invasive method to study airway lining
                                                                        fluid. NuclearMagneticResonance (NMR) based metabonomic analysis provides
                                                                        biochemical fingerprint in biological fluids.
                                                                        Aim: to evaluate EBC in healthy adults (HS) and in COPD patients using NMR
                                                                        spectroscopy examining the role of preanalytical variables such as saliva or
                                                                        Material and Methods: EBC and saliva samples from 11 HS and 11 COPD were
                                                                        analyzed by means of 1 H-NMR spectroscopy (Bruker DRX-600MHz).The effect
                                                                        on EBC of disinfectant utilized to sterilize reusable parts was assessed.
                                                                        Results: COPD and HS NMR saliva spectra were strikingly different from those
                                                                        of corresponding EBC.

                                                                        The disinfectant perturbation on EBC spectra was almost completely removed
                                                                        using ethanol.COPD spectra differed from HS by five signals, which appear to be
                                                                        able to clusterize the data in two different groups (94%).
                                                                        Conclusions: EBC metabonomics in adults discriminate the potential perturbation
                                                                        by preanalytical variables such saliva or disinfectants.The NMR-metabonomic ap-

                    Abstract printing supported by Nonin Medical, Inc. Visit Nonin Medical, Inc. at Hall 3.2 Stand 3.57
Oral Presentation                                                                                                                         Room Europa 2 - 08:30-10:30

                                                                M ONDAY, O CTOBER 6 TH 2008

                                                                                          the mechanisms responsible for this change and its precise role requires further
                                                                                          The study was supported by OTKA (68808).

                                                                                          Alpha-1-antitrypsin is detectable and can be quantified in exhaled breath
                                                                                          Sarah Noeske, Robert Bals, Christian Herr, Claus Vogelmeier, Andreas
                                                                                          Rembert Koczulla. Department of Internal Medicine, Division for Pulmonary
                                                                                          Diseases, Philipps-University Marburg, Marburg/Hesse, Germany

                                                                                          Rationale: Alpha-1-antitrypsin deficiency (AATD) is a genetic disorder that pre-
                                                                                          disposes to chronic obstructive pulmonary disease (COPD). The main function
                                                                                          of alpha-1-antitrypsin (AAT) is to protect the lungs against neutrophil elastase.
proach could disclose the metabolic fingerprint of EBC giving a clearcut separation        Until now a non invasive method to detect AAT reliably in the epithelial lining
between HS and COPD patients.                                                             fluid (ELF) is missing. Measurement of exhaled breath condensate (EBC) is a non
                                                                                          invasive method to investigate parameters of the ELF of the lung.
                                                                                          Aim: It was the aim of the study to show that AAT is measurable with the non
1564                                                                                      invasive method of EBC. A Western Blot and ELISA should be validated and
Breath analysis in renal failure: effect of hemodialysis                                  standardized to detect very small amounts of AAT in EBC.
Enrico Heffler 1 , Nicoletta Ferrero 1 , Luisa Bommarito 1 , Massimiliano Bugiani 2 ,      Methods: 1ml of EBC from each patient was lyophilized. Afterwards Western Blot
Roberta Bergia 1 , Giuseppe Guida 1 , Stefano Pizzimenti 1 , Michele Bruno 3 ,            and ELISA were established. Design: Measurement was performed in 10 healthy
Michele Petrarulo 3 , Giovanni Rolla 1 . 1 Department of Allergy and Clinical             controls (HC), 10 patients with AATD and 10 smokers without lung disease. EBC
Immunology, University of Torino–Azienda Sanitaria Ospedale Mauriziano                    from all individuals was collected with the RTube™ device.
“Umberto I”, Torino, Italy; 2 Department of Respiratory Medicine,                         Results: AAT in EBC was detectable by Western Blot and ELISA in each in-
CPA–ASL-TO2, Torino, Italy; 3 Department of Nephrology, Azienda Sanitaria                 dividual. We could detect AAT in concentrations down to 0.2ng/ml. We found
Ospedale Mauriziano “Umberto I”, Torino, Italy                                            differences regarding the AAT levels between HCs, patients with AATD and
Background: Impaired nitric oxide (NO) signaling is thought to play a major               Conclusion: We were able to detect and quantify AAT in EBC. The highly sen-
role in endothelial dysfunction of patients with renal failure (RF). A simple way         sitive developed assay can be used for measuring levels of AAT from the lung of
to assess NO production is to measure its concentration in exhaled air (FENO).            humans non-invasively. This could be of value for therapy monitoring.
Airway pH may influence FENO and in patients in hemodialysis (HD) it may have
significant shift, with possible influence on FENO. Ammonia (NH3) is involved
in airway pH homeostasis and it is present in great concentration in the breath of        1567
RF patients, and it is largely removed by HD                                              An electronic nose distinguishes the exhaled breath condensates of AATD
Methods: Airway and arterial blood pH, metabolites of NO (NOx/NO2) and NH3                patients and healthy controls
in Exhaled Breath Condensate (EBC), and FENO were measured in 12 patients                 Rembert Koczulla, Akira Hattesohl, Sarah Noeske, Claus Vogelmeier,
with RF in mantainance HD 30 minutes before and after HD, and in 14 healthy               Robert Bals. Department of Medicine, Division of Pulmonology,
controls (HC)                                                                             Philipps-Universität Marburg, Marburg, Hessen, Germany
                                                                                          Background: Exhaled respiratory air contains countless volatile organic com-
                          RF before HD      RF after HD   HC         p value† p value‡    pounds (VOCs). The composition of those VOCs varies dependent of different
                                                                                          pulmonary diseases. With the customizable electronic nose Cyranose 320 (C-320)
FENO, ppb (IC95%) 14.5 (12.2-21.0) 11.5 (7.2-19.0) 13.0 (10.7-21.0) >0.05      0.012      it is possible to detect VOCs and distinguish between different VOC patterns. It
EBC-pH (IC95%)     7.16 (6.8-7.4)    6.4 (5.6-6.8)    6.6 (6.4-6.8)  0.004     0.002
                                                                                          was the aim of this study to analyze and distinguish the VOCs in exhaled breath
Arterial blood pH
                                                                                          condensates (EBC) of patients with α1 -antitrypsin deficiency (AATD) and healthy
   (IC95%)        7.35 (7.33-7.35) 7.46 (7.44-7.46) 7.41 (7.39-7.42) 0.03      0.002
                                                                                          controls (HC).
NOx, μM (IC95%)   12.3 (11.1-41.9)  9.9 (5.0-16.4)   9.4 (4.6-10.9)  0.007      0.01
NO2, μM (IC95%)     4.7 (1.2-8.2)    0.8 (0.6-1.2)    0.9 (0.7-1.2)  0.004     0.008
                                                                                          Material and Methods: EBC was collected from ten HCs and ten AATD patients
NH3, μM (IC95%) 2340 (1325-3922) 527 (318-895) 660 (406-872) <0.001            0.002      with the RTube by ten minutes of tidal breathing. 200 μl of EBC were heated
                                                                                          up 37° C and argon was passed through with a constant flow of 350 ml /min for
    before RD vs HC; ‡ before RD vs after HD.                                             two minutes. Subsequently, the samples were measured by holding the snout of
                                                                                          the C-320 above the surface of the samples and drawing gas for ten seconds. Ten
Conclusion: FENO is normal in RF, while EBC-NOx/NO2 are increased, suggest-               samples of ambient lab air were taken as loading controls. A linear discriminant
ing oxidative stress of respiratory system. The huge amount of EBC-NH3 and its            analysis (LDA) has been performed on the data.
dramatic decrease after HD may explain the higher than normal levels of EBC-pH            Results: The analyses showed that AATD patients, HCs and lab air are clearly sep-
in RF before HD and its decrease after HD.                                                arable. The predictive positive value (PPV) was calculated with a cross-validation
                                                                                          using ten percent of the training data as test data (see table).

1565                                                                                      Mahalanobis distances (and PPVs) between the groups
Effect of exercise on exhaled breath condensate pH in young healthy subjects                                        HC                          lab air
Andras Bikov 1 , Balazs Szili 1 , Balazs Antus 2 , Mark Kollai 1 , Ildiko Horvath 1 .
  Human Physiology and Clinical Experimental Research, Semmelweis University,             AATD                 2.390 (63.0%)                2.393 (93.5%)
Budapest, Hungary; 2 Pathophysiology, National Korányi Institute for TB and               HC                                                2.402 (79.0%)
Pulmonology, Budapest, Hungary
                                                                                          Conclusion: Using the C-320, we could distinguish between EBCs from AATD
Introduction: Asthma-like symptoms are frequent in healthy people performing              patients and HCs. This approach also shows EBC contains enough VOCs to
regular exercise in cold temperature. However the mechanism responsible for this          separate two different patient groups. An advantage of using the EBC is the lower
link is not known. Based on current knowledge changes may involve an increase in          contamination risk by not measuring for example infectious patients directly with
oxidative stress and in association with it a decrease in pH that may be represented      the C-320 and the easiness to sample EBC.
in exhaled biomarker concentrations.
Objective: We aimed to follow changes in exhaled breath condensate (EBC) pH
and 8-isoprostane, a biomarker of oxidative stress, together with lung function in        1568
young healthy subjects performing exercise challenge at cold temperature.                 Repeated eNose assessment in the monitoring of O2 -induced changes in
Methods: Ten healthy subjects (21.4±1.1 years) participated in the study. At rest,        exhaled breath metabolites in COPD
immediately and 30 minutes after exercise challenge at 5±1 °C FEV1 , FVC was              Marten de Vries 1 , Jan van der Maten 1 , Peter J. Sterk 2 , Anneke Ten Brinke 1 .
measured and EBC was collected to determinate the changes in airway pH and                  Department of Pulmonary Diseases, Medical Centre Leeuwarden, Leeuwarden,
8-isoprostane levels. EBC pH was measured as previously described (Kullmann et            Netherlands; 2 Department of Pulmonary Diseases, Academic Medical Center,
al, Eur Resp J, 2007; 29:185-7) and 8-isoprostane was determined by commercially          Amsterdam, Netherlands
available ELISA. Data are expressed as mean±SD.
Results: Exercise caused a small decrease in FEV1 , while FVC did not change.             Background: COPD is a complex disease in which oxidative stress-induced in-
There was a significant increase in EBC pH (from 5.9±0.13 to 6.07±0.12,                    flammation may play a pathophysiological role. Oxygen (O2 ) administration has
p=0.002), but there was no significant change in 8-isoprostane concentration.              been shown to increase markers of oxidative stress in exhaled breath condensate
Conclusion: The increased airway pH suggested by changes in EBC pH could                  (Carpagnano, Thorax 2004). Electronic nose (eNose) technology allows online
play a role in airway regulation during and after exercise. However to understand         pattern recognition of volatile organic compounds (VOCs). We hypothesized that

                             Abstract printing supported by Nonin Medical, Inc. Visit Nonin Medical, Inc. at Hall 3.2 Stand 3.57
Oral Presentation                                                                                                                      Room Europa 2 - 08:30-10:30

                                                                   M ONDAY, O CTOBER 6 TH 2008

the O2 -induced changes in exhaled metabolites in COPD can be detected by                 observed in healthy smokers even without having airways obstruction detected
changes in VOC profiles (smellprints).                                                     with PFTs. Similar characteristics of dynamic lung images have been recorded in
Aim: We investigated in 10 ex-smoking COPD patients (GOLD stage II/III: 7/3;              COPD patients but with higher intensity and proportion. Spirometric values and
smoking: 30 (10-45) pck yrs) whether smellprints before and repeatedly after short        VRI characteristics are shown. VRI seems to be a valuable system applicable in
term O2 supply can be discriminated by eNose.                                             airways obstructive diseases and might be an additional tool for screening smokers.
Methods: After baseline sampling (t0 ), O2 (3L/min, VOC-free) was supplied for 40
min through a nasal canula. Thereafter exhaled breath was collected at 0(t1 ), 10(t2 ),
30(t3 ), 50(t4 ), 70(t5 ) and 100(t6 ) min, by expiratory VC according to Dragonieri
(JACI 2007), and sampled by eNose (Cyranose 320). Smellprints were analysed
by linear discriminant analysis on principal component reduction. Cross-validation
values (CVV) and Mahalanobis distance (M-dist) were calculated.
Results: After 40 min of O2 supply, pO2 changed from 9.1±1.3 to 13.0±2.5 kPa
(p<0.001). Smellprints at all time points after O2 administration could be well
discriminated from baseline (M-dist 2.69-4.27; CVV ≥90%), except for t3 (M-dist
1.70; CVV 75%).
Conclusion: eNose technology is able to discriminate smellprints before and after
short term O2 supply in COPD patients. This suggests that oxidative stress can be
readily assessed from VOC patterns in exhaled air, which provides a monitoring
tool in the clinical research of COPD.

Breath analysis by ion mobility spectrometry in sarcoidosis – results of a
feasibility study
Michael Westhoff 1 , Patric Litterst 1 , Barbara Obertrifter 1 ,
Alexander Bunkowski 2 , Joerg Ingo Baumbach 2 . 1 Pneumology and Sleep
Medicine, Lung Clinic Hemer, Hemer, Germany; 2 Metabolomics Department,
ISAS–Institute for Analytical Sciences, Dortmund, Germany

Purpose: The development of non-invasive diagnostic tools for the differentiation
of pulmonary disorders is desirable. We examined if ion mobility spectrometry can
detect volatile organic compounds (VOC) in human exhaled air that characterize
patients with sarcoidosis.
Methods: An ion mobility spectrometer (IMS) coupled to a multi-capillary-column
(MCC) was used to identify and quantify volatile metabolites occurring in human
breath down to the ng/L- and pg/L-range of analytes within less than 500 s and
without any pre-concentration. The IMS investigations are based on different drift
times of swarms of ions of metabolites formed directly in air at ambient pressure.
The exhaled breath of 20 patients with mediastinal lymph node enlargement (11
patients with sarcoidosis and 9 with exclusion of sarcoidosis) was investigated.
Results: A procedure related to a single peak in the IMS-chromatogram delivered
a differentiation into two groups of mediastinal lymph node enlargement (con-
firmed and excluded sarcoidosis) with a sensitivity of 0.91 and a specifity of 0.89.
The potential biomarker for sarcoidosis is characterised by the parameters inverse
mobility (1/K0 ) 0.53±0.01 Vs/cm2 and retention time 22±5 s.
Conclusion: The data show that ion mobility spectrometry may provide a typical
chromatogram in patients with sarcoidosis. This might allow differentiation of
mediastinal lymph node enlargement related to sarcoidosis from other diseases.
These first data need further confirmation by prospective studies.
Clinical Implications: Ion mobility spectrometry might gain an important role in
the non-invasive diagnosis of sarcoidosis.

Is vibration response imaging (VRI) valuable in early detection of airways
obstruction in smokers?
Angeliki Florou, Sofia Vakali, Dimitrios Michailidis, Charis Roussos,
Christina Gratziou. Asthma and Allergy Center, Pulmonary and Critical Care
Department, Medical School-University of Athens-Evgenidion Hospital, Athens,

Vibration Response Imaging (VRI) is a new system that provides dynamic image
of the lung by a detection of vibrations produced by airflow during the respiratory
cycle.This study was undertaken in order to validate if VRI can be valuable for
detection of airways obstruction in smokers and in COPD patients and to investi-
gate if correlates with PFTs.25 “healthy”smokers (mean age 45,6±9,6),23 COPD
patients (mean age 59,6±8,) and 10 healthy non-smokers were included in the
study.Healthy smokers were definned those with no spirometric evidence of airways
obstruction.64% of this group reported one or more respiratory symptoms.The VRI
recordings were blindly evaluated characterized as normal and abnormal accord-
ing to dynamic image and maximal energy frame.Abnormal images have been

Spirometric values and VRI characteristics

                                          Non-smokers N=10   Smokers N=25   COPD N=23
FEV1%pred mean (SD)                           96,8 (7,6)      98,8 (17,9)   60,6 (20,3)
FEV1/FVC mean (SD)                            81,5 (8,8)       78,7 (4,7)   58,4 (10,5)
MMEF75/25%pred mean (SD)                      72,7 (18)       71,3 (26,7)   26,3 (12,6)
VRI characteristics                            %cases           %cases       %cases
Dynamic disturbance                               0               64           95,7
Uneven distribution of vibration energy           0               28           82,6
Low vibration energy                              0               24           73,9
Abnormal progression of the image                 0               20           43,5
Wheezing                                          0                8           21,7
Prologation of expiration                         0                8           30,4

                           Abstract printing supported by Nonin Medical, Inc. Visit Nonin Medical, Inc. at Hall 3.2 Stand 3.57

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