Human Exhaled Breath Condensate Research Articles

Biosensor for Rapid Measurement of Lactate in Exhaled Breath Condensate

Lactate concentration plays an important role in disease diagnostics. Higher lactate concentrations are associated with sepsis and septic shock [2], trauma [3], tissue hypoxia due to acute lung injury [4], and respiratory diseases [5]. Lactate level is also widely used in monitoring athletes’ training and fitness as long-term exhausting exercise leads to a rapid increase in lactate level and result in muscle weakening and fatigue [6]. Compared with the traditional blood sample media which is invasive and capillary blood monitoring is not suitable for routine clinical use, the exhaled breath condensate (EBC) is simple and non-invasive method with no potential for adverse effects for lactate analysis.In this presentation, a point-of-care electrochemical biosensor is introduced for the EBC lactate analysis. This disposable and low-cost sensor is aimed to be inserted in an EBC collection device developed by our team that enables real-time monitoring and rapid measurements of EBC lactate level. The sensor measurement is done directly after EBC sample collection, that minimizes the risk of sample degradation and is cheaper than the traditional laboratory techniques such as fluorometry or mass spectrometry.The sensor is modified with poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS), Prussian blue (PB) and lactate oxidase (LOD) enzyme. Lactate concentration is measured from amperometric current response generated by the redox reactions on sensor surface. The enzyme gel formulation is optimized based on rotating disk electrode (RDE) experiments and kinetics analysis, investigating the effect of LOD concentration and gel thickness, and was then applied to disposable sensors.Finally, human EBC analysis is conducted on healthy subjects at rest and after 30 min of intense aerobic cycling exercise. Sensor’s results in real EBC lactate measurements demonstrate a good correlation with fluorometry and mass spectrometry calibrations and exhibit high stability and sensitivity in both short-term and long-term use. Mass production of the sensor is done by robotic gel dispensing using a Biodot aspirate dispensing machine.Note: This presentation is based on the corresponding author's recent publication ‘Rapid Measurement of Lactate in the Exhaled Breath Condensate: Biosensor Optimization and In-Human Proof of Concept’ in ACS Sensors (doi.10.1021/acssensors.2c01739) [1]

A clinical proteomics study of exhaled breath condensate and biomarkers for pulmonary embolism

Pulmonary embolism (PE) can be a diagnostic challenge. Current diagnostic markers for PE are unspecific and new diagnostic tools are needed. The air we exhale is a possible new source for biomarkers which can be tapped into by analysing the exhaled breath condensate (EBC). We analysed the EBC from patients with PE and controls to investigate if the EBC is a useful source for new diagnostic biomarkers of PE. We collected and analysed EBC samples from patients with suspected PE and controls matched on age and sex. Patients in whom PE was ruled out after diagnostic work-up were included in the control group to increase the sensitivity and generalizability of the identified markers. EBC samples were collected using an RTube™. The protein composition of the EBCs were analysed using data dependent label-free quantitative nano liquid chromatography–tandem mass spectrometry. EBC samples from 28 patients with confirmed PE, and 49 controls were analysed. A total of 928 EBC proteins were identified in the 77 EBC samples. As expected, a low protein concentration was determined which resulted in many proteins with unmeasurable levels in several samples. The levels of HSPA5, PEBP1 and SFTPA2 were higher and levels of POF1B, EPPK1, PSMA4, ALDOA, and CFL1 were lower in PE compared with controls. In conclusion, the human EBC contained a variety of endogenous proteins and may be a source for new diagnostic markers of PE and other diseases.

Low fouling aptasensing of rivaroxaban in real samples using poly (toluidine blue) decorated by silver nanoparticle: A new platform for the cardiovascular disease analysis

A novel ultrasensitive electrochemical aptasensor was developed for the detection of rivaroxaban (RIV). For this purpose, poly toluidine blue (P(TB)), silver nanoparticles (AgNPs), aptamer conjugated with polyethylene glycol (PEG), and ITO-PET (indium tin oxide coated polyethylene terephthalate film) electrode was utilized. In order to fabrication of the low fouling aptasensor, the surface of ITO-PET was efficiently modified by P(TB-AgNPs) to provide a suitable substrate for the stable immobilization of aptamer sequence, 5′ SH-TAG GGA AGA GAA GGA CAT ATG ATG ACT CAC AAC TGG ACG AAC GTA CTT ATC CCC CCC AAT CAC TAG TGA ATT-3′). Using ITO PET-OH-P (TB-Ag)-PEG as substrate and aptamer as bioreceptor, RIV were measured in human plasma and exhaled breath condensate samples by chronoamperometry technique with limit of detections of 6.65 and 4.13 nM, respectively. Importantly, using PEG conjugated aptamer, a low fouling aptasensor was developed for the sensitive and selective determination of RIV in spiked samples. For the first time, a novel aptasensor for determination of RIV in human plasma and exhaled breath condensate (EBC) samples was developed which is able to use for therapeutic drug monitoring (TDM) studies.

Construction of a hydrophobic-hydrophilic open-droplet microfluidic chemosensor towards colorimetric/spectrophotometric recognition of quetiapine fumarate: a cost-benefit method for biomedical analysis using a smartphone.

Quetiapine fumarate (QF) is used to treat a number of mental/emotional diseases, including schizophrenia, bipolar disorder, and abrupt bouts of mania or depression linked to bipolar disorder. This antipsychotic medicine can be deadly if an overdose is given to a person. Therefore, the sensitive identification of QF in bodily fluids is very important. In this study, an innovative low-cost colorimetric chemosensor based on silver nanoprism transfiguration in a phosphate-buffered saline (PBS)/Cl- matrix was developed and successfully tested for the recognition of QF in human-exhaled breath condensate. Using this non-invasive colorimetric chemosensor, a broad linearity range of 0.001-1000 μM and a low limit of quantification of 0.001 μM for QF were attained. Notably, the proposed optical chemosensor is capable of detecting QF from a minimum amount of sample [500 μM in PBS and 0.001 μM in exhaled breath condensate] in the first few seconds of reaction by the naked eye. So, a rapid colorimetric assay for the on-site analysis of QF was developed and validated. Moreover, for the first time, a semi-analytical method was introduced that can provide a rough estimation of the QF concentration. This colorimetric system was, for the first time, integrated in an optimized microfluidic paper-based colorimetric device (μPCD), promising the development of an engineered colorimetric opto-sensor toward real-time and therapeutic drug monitoring (TDM) assay of drugs in real-world samples.

Rapid Measurement of Lactate in the Exhaled Breath Condensate: Biosensor Optimization and In-Human Proof of Concept.

Lactate concentration is of increasing interest as a diagnostic for sepsis, septic shock, and trauma. Compared with the traditional blood sample media, the exhaled breath condensate (EBC) has the advantages of non-invasiveness and higher user acceptance. An amperometric biosensor was developed and its application in EBC lactate detection was investigated in this paper. The sensor was modified with PEDOT:PSS-PB, and two different lactate oxidases (LODs). A rotating disk electrode and Koutecky-Levich analysis were applied for the kinetics analysis and gel optimization. The optimized gel formulation was then tested on disposable screen-printed sensors. The disposable sensors exhibited good performance and presented a high stability for both LOD modifications. Finally, human EBC analysis was conducted from a healthy subject at rest and after 30 min of intense aerobic cycling exercise. The sensor coulometric measurements showed good agreement with fluorometric and triple quadrupole liquid chromatography mass spectrometry reference methods. The EBC lactate concentration increased from 22.5 μM (at rest) to 28.0 μM (after 30 min of cycling) and dropped back to 5.3 μM after 60 min of rest.

A non-invasive method for the detection of glucose in human exhaled breath by condensation collection coupled with ion chromatography

An analytical method for the detection of glucose in human exhaled breath by non-invasive condensation collection coupled with ion chromatography was developed. A self-designed exhaled breath condensation device was constructed, through which human exhaled breath was condensed and collected. And the glucose in collected human exhaled breath condensate (EBC) was analyzed by ion chromatography with a pulsed amperometric instrument. The standard EBC collection method was established, and the key factors such as cooling temperature and sampling flow rate during condensation collection were investigated deeply, and a good linear correlation between blood glucose levels and exhaled breath glucose levels was obtained. The intra-day precision was 3.37%, and the inter-day precision was 3.83%. Furthermore, EBC from healthy people and diabetic patients was collected in fasting state and after meal. We found the breath glucose level in healthy volunteers was 0.11–2.34 ng/L and that in diabetic patients was 0.23–135.92 ng/L. In after meal samples, the breath glucose level is 10–100 times higher than that of healthy subjects, which offering the prospect of a non-invasive approach to the monitoring of diabetes.

Selection of Specific Aptamer against Rivaroxaban and Utilization for Label-Free Electrochemical Aptasensing Using Gold Nanoparticles: First Announcement and Application for Clinical Sample Analysis.

For the first time, a novel aptamer was designed and utilized for the selective detection of rivaroxaban (RIV) using the integration of bioinformatics with biosensing technology. The selected aptamer with the sequence 5′-TAG GGA AGA GAA GGA CAT ATG ATG ACT CAC AAC TGG ACG AAC GTA CTT ATC CCC CCC AAT CAC TAG TGA ATT-3′ displayed a high binding affinity to RIV and had an efficient ability to discriminate RIV from similar molecular structures. A novel label-free electrochemical aptasensor was designed and fabricated through the conjugation of a thiolated aptamer with Au nanoparticles (Au-NPs). Then, the aptasensor was successfully applied for the quantitative determination of RIV in human plasma and exhaled breath condensate (EBC) samples with limits of detection (LODs) of 14.08 and 6.03 nM, respectively. These valuable results provide ample evidence of the green electrogeneration of AuNPs on the surface of electrodes and their interaction with loaded aptamers (based on Au-S binding) towards the sensitive and selective monitoring of RIV in human plasma and EBC samples. This bio-assay is an alternative approach for the clinical analysis of RIV and has improved specificity and affinity. As far as we know, this is the first time that an electrochemical aptasensor has been verified for the recognition of RIV and that allows for the easy, fast, and precise screening of RIV in biological samples.

EBC-SURE (exhaled breath condensate- scanning using rapid electro analytics): A non-faradaic and non-invasive electrochemical assay to screen for pro-inflammatory biomarkers in human breath condensate

The innovation of this work lies in the trace detection of inflammatory biomarkers (IL-6, hs-CRP) in human exhaled breath condensate on the developed EBC-SURE platform as a point-of-care aid for respiratory disorder diagnosis. The unique design of the EBC-SURE leverages non-faradaic electrochemical impedance spectroscopy to capture target-specific biomolecular interactions for highly sensitive biomarker detection. For sensor calibration, EBC-SURE's performance is assessed to measure the response of the sensor to a known concentration by spike and recovery analysis with a recovery error of <20% and an extended dynamic range over 3-log orders. The lowest detection limits for IL-6 and hs-CRP detection in EBC were found to be 3.2 pg/mL and 4 pg/mL respectively. The intra-assay and inter-assay efficacy of EBC-SURE for its usage as a diagnostic device was established through repeatability and reproducibility (over 48 h s) performance testing. The percentage variations (<20%) met the Clinical and Laboratory Standards Institute standards (CLSI) indicating a highly stable performance for robust biomarker detection. EBC-SURE generated highly selective IL-6 and hs-CRP responses in the presence of other non-specific cytokines. Statistical validation methods- Correlation and Bland Altman analysis established the one-to-one agreement between EBC-SURE and the reference method. Correlation analysis generated a Pearson's R value of 0.99 for IL-6 and hs-CRP. Bland-Altman analysis indicated a good agreement between both the methods with all data points confined within the ±2SD limits. We have demonstrated EBC-SURE's ability in detecting inflammatory biomarkers in human breath condensate towards developing a non-invasive technology that can quantify biomarker levels associated with healthy and acute inflammatory conditions.

Optical biosensing of markers of mucosal inflammation

We report the design and adaptation of iron/iron oxide nanoparticle-based optical nanobiosensors for enzymes or cytokine/chemokines that are established biomarkers of lung diseases. These biomarkers comprise ADAM33, granzyme B, MMP-8, neutrophil elastase, arginase, chemokine (C-C motif) ligand 20 and interleukin-6. The synthesis of nanobiosensors for these seven biomarkers, their calibration with commercially available enzymes and cytokines/chemokines, as well as their validation using bronchoalveolar lavage (BAL) obtained from a mouse model of TLR3-mediated inflammation are discussed here. Exhaled Breath Condensate (EBC) is a minimally invasive approach for sampling airway fluid in the diagnosis and management of various lung diseases in humans (e.g., asthma, COPD and viral infections). We report the proof-of-concept of using human EBC in conjunction with nanobiosensors for diagnosis/monitoring airway inflammation. These findings suggest that, with nanosensor technology, human EBC can be utilized as a liquid biopsy to monitor inflammation/remodeling in lung disease.

Metabolomics profiling of human exhaled breath condensate by SPME/GC × GC-ToFMS: Exploratory study on the use of face masks at the level of lipid peroxidation volatile markers

Metabolomics encompasses the acquisition of large-scale datasets from a single sample to identify and/or elucidate physiological or pathological mechanisms, or even to assess environmental exposure, being sample preparation and instrumental analysis two fundamental steps in the construction of a metabolomics workflow. The study of the human exhaled breath condensate (EBC) has gained tremendous interest recently owing to the advances in analytical techniques. Thus, this study aims to develop a methodology for EBC metabolites profiling based on headspace solid phase microextraction (SPME) combined with comprehensive two-dimensional gas chromatography (GC × GC). Metabolite’s extraction efficiency was improved by the optimization of four SPME parameters using a real EBC matrix: fibre coating, sample pH and volume, and extraction temperature. EBC analysis was performed according to the optimized parameters, using aliquots of 500 μL of EBC, at its physiological pH, with the DVB/CAR/PDMS fibre coating, at 40.0 ± 0.1 °C, being able to detect analytes at pg/mL, with acceptable reproducibility. EBC volatile composition was provided, with around 400 chemical features detected, in which 130 were putatively identified as belonging to hydrocarbons, aldehydes, alcohols, ketones, esters, monoterpenic and sesquiterpenic compounds, ethers, norisoprenoids and furans, revealing therefore its high complexity. This research represents the most detailed study on the volatile composition of EBC and allows the assessment of a broad range of endogenous and exogenous metabolites, with further relevance on clinical or environmental exposure studies. To test the applicability of this methodology, an exploratory study was carried out to assess the impact of the use of surgical facemasks on young researcher’s lipid peroxidation volatile markers during a normal working day in a laboratory. In young healthy adults (age ≤ 32), the use of surgical facemasks seems not significantly affect the levels of the volatile markers of lipid peroxidation (aliphatic alkanes and aldehydes), neither the O2 saturation level - SpO2. This methodology shows promise as a convenient way to follow specific physiological or pathological conditions based on EBC metabolome and may be paired as part of an aggregate analytical structure.

A novel electrochemical sensor based on ion imprinted polymer and gold nanomaterials for nitrite ion analysis in exhaled breath condensate

Human Exhaled Breath Condensate (EBC) contains markers of several inflammatory diseases. Its analysis is of interest to a number of researchers. Nitrite ions (NO2-), which are widely used in our daily lives, are nevertheless among these indicators. In this study, a simple, fast, portable, non-invasive and cheap electrochemical sensor is developed for the analysis of the nitrite profile in EBC. In this regard, sodium nitrite (NaNO2) was first immobilized on self-assembled 2-aminothiophenol (2-ATP) on a screen-printed gold electrode (Au-SPE). Then, a polymer matrix composed of polyvinyl alcohol (PVA) crosslinked with glutaraldehyde (GA) was combined with gold nanoparticles (Au-NPs) to cover the modified Au-SPE and complete the fabrication of the Ion Imprinted Polymer (IIP) sensor. The electrochemical behaviour of the sensor was monitored using Cyclic Voltammetry (CV), Electrochemical Impedance Spectroscopy (EIS) and Differential Pulse Voltammetry (DPV) methods, while the morphology and chemical composition of its layers were observed by infrared Fourier transform (FTIR), Atomic Force Microscopy (AFM) and Scanning Electron Microscopy coupled with energy dispersion X-Ray spectroscopy (SEM-EDS) techniques. In addition, after a successful control test using a Non-Imprinted Ion Polymer (NIIP) sensor, the obtained results demonstrated satisfactory sensitivity and selectivity to nitrite compared to co-existing interfering substances in EBC, such as nitrate, acetate and ammonium nitrate. Under improved experimental conditions, the nitrite IIP sensor exhibits responses proportional to nitrite concentrations (R2 = 0.96) over a concentration range of 0.5–50 μg mL−1 with a detection limit (LOD) of 4 μmol L−1 (signal-to-noise ratio S/N = 3). The proposed approach was well applied for the nitrite determination in EBC samples with a relative standard deviation (RSD = 4%) and could open clinical applications in respiratory medicine.

A Facile Profiling Method of Short Chain Fatty Acids Using Liquid Chromatography-Mass Spectrometry

Short chain fatty acids (SCFAs) are the main products of dietary fibers that are not digested by the human body, and they have been shown to affect human metabolism and inflammation. The amount of SCFAs in the body is related to many human diseases, and studies have focused on elucidating their roles and target molecules in both metabolic and immune responses. Thus, the quantitation of SCFAs in biological samples becomes crucial in understanding their important roles in the human body. Herein, a facile profiling method of SCFAs using liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed and then applied to biological samples. C2-C6 SCFAs were derivatized while using 4-acetamido-7-mercapto-2,1,3-benzoxadiazole for 5 min. at room temperature prior to LC-MS/MS analysis, and characteristic fragmentation patterns and increased hydrophobicity after chemical derivatization enabled specific discrimination among 12 SCFAs. Derivatization was fast and reliable, and the reaction products were stable for a week at 4 °C. The developed method was applied to measure SCFAs in mouse feces, plasma, and human exhaled breath condensates. This fast and simple method can save labor and effort to profile SCFAs from various biological samples.

Electrochemical determination of malondialdehyde biomarker in exhaled breath condensate using poly arginine functionalized by graphene quantum dots and chitosan

Background: Lipid peroxidation is a prominent manifestation of free radical activity in biological systems, and it is involved in the development of different diseases such as cancer or cardiovascular and liver diseases. Malondialdehyde (MDA) is globally recognized as a scientifically accepted biomarker of oxidative stress. Due to the significance of MDA as a marker of lipid peroxidation and because of its elevations in various diseases, the assessment of MDA with new and reliable assays, the development of a fast, sensitive, and selective method to detect MDA with simple instruments is necessary. Methods: This study used the electro polymerization of a low toxic and biocompatible polymer with entitle poly arginine-graphene quantum dots-chitosan (PARG-GQDs-CS) as a novel strategy for surface modification of glassy carbon (GC) surface and preparation a new interface for measurement of MDA in human exhaled breath condensate (EBC). Nanocomposite PARG/CS/GQDs the method of precipitation out of place (ex situ) on the electrode surface was prepared. To verify sequestration nanobiopolymer synthesized Scanning electron microscope images (SEM) were recorded. To identify and study the behavior of malondialdehyde electrochemical methods such as cyclic voltammetry and differential pulse voltammetry were used. Results: The scanning electron microscopy (SEM) images indicated immobilization of uniformly CS and GQDs onto PARG film. The modified electrode appeared as an effective electro activity for detection of MDA. Under the optimized analysis conditions and physiological pH, the prepared sensor showed one oxidation processes for MDA at potentials about 1 V vs. Ag/AgCl with a low limit of quantification (LLOQ) 5.94 nM. Conclusion: The Glassy carbon electrod modified by poly-arginine/chitosan/graphene quantum dots, has good electrical activity for Electro-oxidation malondialdehyde that could be due to synergistic effect poly-arginine, graphene quantum dots and chitosan to create many active sites for this purpose.

Determination of Verapamil in Exhaled Breath Condensate by Using Microextraction and Liquid Chromatography

Background:Developing a simple analysis method for quantification of drug concentration is one of the essential issues in pharmacokinetic and therapeutic drug monitoring studies.Objective:A fast and reliable dispersive liquid-liquid microextraction procedure was employed for preconcentration of verapamil in exhaled breath condensate (EBC) samples and this was followed by the determination with high-performance liquid chromatography-ultraviolet detection.Methods:A reverse-phase high-performance liquid chromatography (RP-HPLC) combined with a dispersive liquid-liquid microextraction method (DLLME) was applied for quantification of verapamil in the EBC samples. The developed method was validated according to FDA guidelines.Results:Under the optimum conditions, the method provided a linear range between 0.07 and 0.8 µg.mL-1 with a coefficient of determination of 0.998. The intra- and inter-day relative standard deviation and relative error values of the method were below 15%, which indicated good precision and accuracy. The proposed method was successfully applied for the analysis of verapamil in two real samples with concentrations of 0.07 and 0.09 µg.mL-1.Conclusion:The established HPLC-UV-DLLME method could be applied for the analysis of verapamil in human EBC samples.

Electrophoretic analysis of polyamines in exhaled breath condensate based on gold-nanoparticles microextraction coupled with field-amplified sample stacking

In this work, citrate-capped gold-nanoparticles (citrate-AuNPs) have been firstly used for selective extraction of trace polyamines, putrescine (Put) and cadaverine (Cad), followed by field-amplified sample stacking (FASS) coupled with capillary electrophoresis and capacitively coupled contactless conductivity detection (FASS-CE-C4D). Put and Cad were extracted by electrostatic attractions between the amine group of the polyamines and the citrate ligands adsorbed on the surfaces of AuNPs. AuNPs microextraction (AuNPs-ME) effectively shortened preparation time (50 min) by introducing ultrasound, and the required sample extraction volume was only 1 mL. Furthermore, a synergistic enrichment strategy based on off-line AuNPs-ME and on-line FASS significantly improved the detection sensitivity, making the enrichment factors up to 1726–1887 times. Under the optimum conditions, Put and Cad could be well separated from the potential coexisting substances and then directly determined by CE-C4D without derivatization. Due to its low sample consumption, high sensitivity (LODs: 0.070–0.17 ng mL−1), and acceptable recoveries (90–105%), this AuNPs-ME/FASS-CE-C4D method provides a rapid, economical and eco-friendly approach for direct determination of polyamines in human exhaled breath condensate, and has potential application prospects in preliminary noninvasive diagnosis of oral and respiratory inflammation.

Non-invasive quantification of malondialdehyde biomarker in human exhaled breath condensate using self-assembled organic-inorganic nanohybrid: A new platform for early diagnosis of lung disease

A novel electrochemical sensor was developed towards recognition of malondialdehyde (MDA) with modifying the glassy carbon electrode (GCE) by self-assembled riboflavin-taurine (RFPT) as organic substrate and silver nanoparticle (Ag NPs) as inorganic nano-catalyst. Therefore, a novel biocompatible and anti-bacterial substrate were prepared for the construction of electrode. In proposed sensor, the RF-PT polymer film led to the emergence of electroactive and suitable substrate for the immobilization of Ag NPs. The use of the proposed nano-hybrid significantly amplified the electrochemical signals the redox behavior was investigated using various electrochemical methods including cyclic voltammetry (CV), differential pulse voltammetry (DPV), square wave voltammetry (SWV), and linear sweep voltammetry (LSV). It is the adaptation of the established RF-PT-Ag NPs organic-inorganic hybrid for a selective, robust, and generalizable sensing system that is the emphasis of this work. The results show that the proposed platform has a good sensitivity in the detecting of MDA with a low limit of quantification (LLOQ) of 0.59 ± 0.05 μM. In addition, the modified electrode reveals promising efficiency against MDA oxidation and its side-products destructive effects. For the first time, the engineered sensor was used to non-invasive determination MDA in exhaled breath condensate (EBC) samples without the need to extra sample preparation steps. The results show RF-PT-AgNPs as a novel biocompatible interface can provide appropriate, reliable, in-expensive, fast, and user-friendly diagnostic tools in the detection of MDA in human real samples.

Metabolomics of tracheal wash samples and exhaled breath condensates in healthy horses and horses affected by equine asthma

The present work characterized the metabolomic profile of tracheal wash (TW) and exhaled breath condensate (EBC) in healthy horses and horses with respiratory disease. Six asthma-affected horses (group A) and six healthy controls (group H) underwent clinical, endoscopic and cytologic examinations of upper airways to confirm the active phase of asthma. TW and EBC samples were collected from each animal and investigated by proton nuclear magnetic resonance (1H-NMR) metabolomic analysis. A total of ten out of 38 metabolites found in the TW were significantly different between the groups (p < 0.05). Higher concentrations of histamine and oxidant agents, such as glutamate, valine, leucine and isoleucine, as well as lower levels of ascorbate, methylamine, dimethylamine and O-phosphocholine, were found in group A compared to group H. Eight metabolites were found in equine EBC, namely methanol, ethanol, formate, trimethylamine, acetone, acetate, lactate and butanone, previously observed also in human EBC. Despite the fact that this was a pilot study, the results showed that the metabolomic analysis of TW and EBC has the potentiality to serve as a basis for diagnostic tools in horses with asthma.

The use of chitosan as a bioactive polysaccharide in non-invasive detection of malondialdehyde biomarker in human exhaled breath condensate: A new platform towards diagnosis of some lung disease

Herein, a reusable and time-saving strategy for the electrochemical polymerization of dopamine (EPD) is reported. In this work, biocompatible polymer chitosan (CS) was electrodeposited on the surface of PDA modified glassy carbon electrode. Owing to the abundant catechol and amine groups in the PDA layer, uniform silver nanoparticles (Ag NPs) are deposited onto the POLY(DA-CS) and can effectively prevent the recombination of electron-hole pairs generated from photo-electrocatalysis and transfer the captured electrons to participate in the photo-electrocatalytic reaction process. Compared with pristine POLY(DA-CS), the as-prepared POLY(DA-CS)-AgNPs organic-inorganic hybrid exhibit electrochemical sensitivity for detecting MDA and display excellent durability. Furthermore, the mussel-inspired electropolymerization strategy and the fast EPD-Ag nanoparticle decorating process presented herein can be readily extended to various biomedical analysis and lung cancer detection. It is the adaptation of the established POLY(DA-CS)-Ag NPs organic-inorganic hybrid for a selective, robust, and generalizable sensing system that is the emphasis of this work.

Proteomic characterization of human exhaled breath condensate

To improve biomedical knowledge and to support biomarker discovery studies, it is essential to establish comprehensive proteome maps for human tissues and biofluids, and to make them publicly accessible. In this study, we performed an in-depth proteomics characterization of exhaled breath condensate (EBC), a sample obtained non-invasively by condensation of exhaled air that contains submicron droplets of airway lining fluid. Two pooled samples of EBC, each obtained from 10 healthy donors, were processed using a straightforward protocol based on sample lyophilization, in-gel digestion and liquid chromatography tandem-mass spectrometry analysis. Two ‘technical’ control samples were processed in parallel to the pooled samples to correct for exogenous protein contamination. A total of 229 unique proteins were identified in EBC among which 153 proteins were detected in both EBC pooled samples. A detailed bioinformatics analysis of these 153 proteins showed that most of the proteins identified corresponded to proteins secreted in the respiratory tract (lung, bronchi). Eight proteins were salivary proteins. Our dataset is described and has been made accessible through the ProteomeXchange database (dataset identifier: PXD007591) and is expected to be useful for future MS-based biomarker studies using EBC as the diagnostic specimen.

Bioanalysis of underivatized amino acids in non-invasive exhaled breath condensate samples using liquid chromatography coupled with tandem mass spectrometry

Exhaled breath condensate (EBC) is receiving increased attention as a novel, entirely non-invasive technique for collecting biomarker samples. This increased attention is due to the fact that EBC is simple, effort independent, rapid, can be repeated frequently, and can be performed on young children and patients suffering from a variety of diseases. By having a subject breathe tidally through a cooling system for 15–20 min, a sufficient amount of condensate is collected for analysis of biomarkers in clinical studies. However, bioanalysis of EBC involves an unavoidable sample preparation step due to the low concentration of its components. Thus, there is a need for a new and more sensitive analytical method of assessing EBC samples. While researchers have considered analyses of single and small quantities of amino acids – for example, those connected with leukemia – no one has previously attempted to simultaneously analyze a panel of 23 amino acids. Moreover, the present study is well-justified, as prior studies focusing on single amino acids and leukemia at the moment of diagnosis and during chemotherapy (33 days of treatment) are inconsistent. In the present study, amino acids were separated using an XBridge Amide column (3 mm × 100 mm, 3.5 μm). The mobile phase consisted of 10 mM of ammonium buffer in water with a pH of 3 (Phase A) and 10 mM ammonium buffer in acetonitrile (Phase B) under gradient program elution. The analytes were detected in electrospray positive ionization mode. Under optimal conditions, the proposed method exhibited limits of quantification (LOQ) in the range of 0.05–0.5 ng/mL, and good linearity, with the determination coefficient (R2) falling between 0.9904 and 0.9998. The accuracy in human exhaled breath condensate samples ranged between 93.3–113.3% for the 23 studied amino acids, with intra- and inter-day coefficient of variation (CVs) of 0.13–9.92% and 0.17–10.53%, respectively. To demonstrate the liquid chromatography with hydrophilic interaction with electrospray source coupled to tandem mass spectrometry (LC-HILIC-ESI–MS/MS) method’s applicability for biomedical investigations, it was verified and applied to determine amino acids in pediatric patients with leukemia. These tests confirmed that glutamine, arginine, homoarginine, asparagine, histidine, methionine, proline, hydroxyproline, threonine, tyrosine, and valine were present in significantly higher levels in pediatric leukemia patients than in the healthy control group. The developed assay is an attractive alternative to standard analytical methods, because it allows for the non-invasive, fast, sensitive, and reliable analysis of amino acids without derivatization in EBC.