Search

Infrared spectroscopy is a rapid, nondestructive analytical technique that can be applied to the authentication and characterization of food samples in high throughput. In particular, near infrared spectroscopy is commonly utilized in the food quality control industry to monitor the physical attributes of numerous cereal grains for protein, carbohydrate, and lipid content. IR-based methods require little sample preparation, labor, or technical competence if multivariate data mining techniques are implemented; however, they do require extensive calibration. Economically important crops are infected by fungi that can severely reduce crop yields and quality and, in addition, produce mycotoxins. Owing to the health risks associated with mycotoxins in the food chain, regulatory limits have been set by both national and international institutions for specific mycotoxins and mycotoxin classes. This article discusses the progress and potential of IR-based methods as an alternative to existing chemical methods for the determination of fungal contamination in crops, as well as emerging spectroscopic methods.

An advanced software-controlled focused ion beam (FIB) patterning process for the fabrication of combined atomic force-scanning electrochemical microscopy (AFM-SECM) probes is reported. FIB milling is a standard process in scanning probe microscopy (SPM) for specialized SPM probe fabrication. For AFM-SECM, milling of bifunctional probes usually requires several milling steps. Milling such complex multi-layer/multi-material structures using a single milling routine leads to significantly reduced fabrication times and costs. Based on an advanced patterning routine, a semi-automated FIB milling routine for fabricating combined AFM-SECM probes with high reproducibility is presented with future potential for processing at a wafer level. The fabricated bifunctional probes were electrochemically characterized using cyclic voltammetry, and their performance for AFM-SECM imaging experiments was tested. Different insulation materials (Parylene-C and SixNy) have been evaluated with respect to facilitating the overall milling process, the influence on the electrochemical behavior and the long-term stability of the obtained probes. Furthermore, the influence of material composition and layer sequence to the overall shape and properties of the combined probes were evaluated., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Perfluorocarbon (PFC) compounds have been used as chemical tracer molecules to understand the movement of supercritical carbon dioxide for geosequestration monitoring and verification purposes. A commonly used method for detecting PFCs involves the collection of a sample from either soil-gas or the atmosphere via carbon-based sorbents which are then analyzed in a laboratory. However, PFC analysis in aquatic environments is neglected and this is an issue that needs to be considered since the PFC is likely to undergo permeation through the overlying water formations. This paper presents for the first time an innovative analytical method for the trace level in situ detection of PFCs in water. It reports on the development of a sensor based on mid-infrared attenuated total reflection (MIR-ATR) spectroscopy for determining the concentration of perfluoromethylcyclohexane (PMCH) and perfluoro-1,3-dimethylcyclohexane (PDCH) in aquatic systems. The sensor comprises a zinc selenide waveguide with the surface modified by a thin polymer film. The sensitivity of this device was investigated as a function of polymer type, coating thickness, and solution flow rates. The limit of detection (LOD) was determined to be 23 ppb and 79 ppb for PMCH and PDCH, respectively when using a 5 μm thick polyisobutylene (PIB) coated waveguide. This study has shown that the MIR-ATR sensor can be used to directly quantify PFC-based chemical tracer compounds in water over the 20-400 ppb concentration range., (Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.)

The direct qualitative and quantitative determination of mineral components in shale rocks is a problem that has not been satisfactorily resolved to date. Infrared spectroscopy (IR) is a non-destructive method frequently used in mineral identification, yet challenging due to the similarity of spectral features resulting from quartz, clay, and feldspar minerals. This study reports on a significant improvement of this methodology by combining infrared attenuated total reflection spectroscopy (IR-ATR) with partial least squares (PLS) regression techniques for classifying and quantifying various mineral components present in a number of different shale rocks. The developed multivariate classification model was calibrated using pure component mixtures of the most common shale minerals (i.e., kaolinite, illite, montmorillonite, calcite, and quartz). Using this model, the IR spectra of 11 real-world shale samples were analyzed and evaluated. Finally, the performance of the developed IR-ATR method was compared with results obtained via X-ray diffraction (XRD) analysis.

An infrared attenuated total reflection (IR-ATR) method for detecting, differentiating, and quantifying hydrocarbons dissolved in water relevant for oil spills by evaluating the "fingerprint" of the volatile organic compounds (VOCs) associated with individual oil types in the mid-infrared spectral range (i.e., 800-600 cm(-1)) is presented. In this spectral regime, these hydrocarbons provide distinctive absorption features, which may be used to identify specific hydrocarbon patterns that are characteristic for different crude and refined oils. For analyzing the "VOC fingerprint" resulting from various oil samples, aqueous solutions containing the dissolved hydrocarbons from different crude oils (i.e., types "Barrow", "Goodwyn", and "Saladin") and refined oils (i.e., "Petrol" and "Diesel") were analyzed using a ZnSe ATR waveguide as the optical sensing element. To minimize interferences from the surrounding water matrix and for amplifying the VOC signatures by enrichment, a thin layer of poly(ethylene-co-propylene) was coated onto the ATR waveguide surface, thereby enabling the establishment of suitable calibration functions for the quantification of characteristic concentration patterns of the detected VOCs. Multivariate data analysis was then used for a prelininary classification of various oil-types via their VOC patterns.

The first combination of mid-infrared (MIR) tunable quantum cascade lasers (tQCLs) with thin-film diamond strip waveguides (DSWGs) suitable for advanced chemical sensing/biosensing is demonstrated. The sensing system is composed of thin diamond films grown on surface-passivated Si wafers via chemical vapor deposition (CVD) and microstructured using inductively coupled plasma (ICP) etching, serving as photonic waveguides for radiation emitted by a broadly tunable quantum cascade laser (tQCL) in the spectral regime of 5.78-6.35 μm (1570-1730 cm(-1)). The characterization of the free-standing diamond waveguides reveals excellent transmission properties across a broad MIR band. As a proof of concept, the detection of acetone in D2O via evanescent field absorption is demonstrated achieving a limit of detection (LOD) as low as 200 pL, which indicates a significant sensitivity improvement compared to conventional MIR slab/strip waveguides reported to date. Providing characteristic absorption features within the tuning range of the tQCL, studies using anisaldehyde as an analyte further corroborate the potential of tQCL-DSWG-based chemical sensors/biosensors.

A miniaturised gas analyser is described and evaluated based on the use of a substrate-integrated hollow waveguide (iHWG) coupled to a microsized near-infrared spectrophotometer comprising a linear variable filter and an array of InGaAs detectors. This gas sensing system was applied to analyse surrogate samples of natural fuel gas containing methane, ethane, propane and butane, quantified by using multivariate regression models based on partial least square (PLS) algorithms and Savitzky-Golay 1(st) derivative data preprocessing. The external validation of the obtained models reveals root mean square errors of prediction of 0.37, 0.36, 0.67 and 0.37% (v/v), for methane, ethane, propane and butane, respectively. The developed sensing system provides particularly rapid response times upon composition changes of the gaseous sample (approximately 2 s) due the minute volume of the iHWG-based measurement cell. The sensing system developed in this study is fully portable with a hand-held sized analyser footprint, and thus ideally suited for field analysis. Last but not least, the obtained results corroborate the potential of NIR-iHWG analysers for monitoring the quality of natural gas and petrochemical gaseous products.

Selected volatile organic compounds (VOCs) in breath may be considered biomarkers if they are indicative of distinct diseases or disease states. Given the inherent molecular selectivity of vibrational spectroscopy, infrared sensing technologies appear ideally suitable for the determination of endogenous VOCs in breath. The aim of this study was to determine that mid-infrared (MIR; 3-20 µm) gas phase sensing is capable of determining isoprene in exhaled breath as an exemplary medically relevant VOC by hyphenating novel substrate-integrated hollow waveguides (iHWG) with a likewise miniaturized preconcentration system. A compact preconcentrator column for sampling isoprene from exhaled breath was coupled to an iHWG serving simultaneously as highly miniaturized gas cell and light conduit in combination with a compact Fourier transform infrared spectrometer. A gas mixing system enabled extensive system calibration using isoprene standards. After system optimization, a calibration function obtaining a limit of quantification of 106 ppb was achieved. According to the literature, the obtained sensitivity is sufficient for quantifying middle to high isoprene concentrations occurring in exhaled breath. Finally, a volunteer breath sample was analysed proving comparable values of isoprene in a real-world scenario. Despite its fundamental utility, the proposed methodology contains some limitations in terms of sensitivity and temporal resolution in comparison with the readily available measurement techniques that should be addressed during future optimization of the system. Nonetheless, this study presents the first determination of endogenous VOCs in breath via advanced hollow waveguide MIR sensor technology, clearly demonstrating its potential for the analysis of volatile biomarkers in exhaled breath.

Proper apical airway surface hydration is essential to maintain lung function. This hydration depends on well-balanced water resorption and secretion. The mechanisms involved in resorption are still a matter of debate, especially as the measurement of transepithelial water transport remains challenging. In this study, we combined classical short circuit current (I SC) measurements with a novel D2O dilution method to correlate ion and water transport in order to reveal basic transport mechanisms in lung epithelia. D2O dilution method enabled precise analysis of water resorption with an unprecedented resolution. NCI-H441 cells cultured at an air-liquid interface resorbed water at a rate of 1.5 ± 0.4 μL/(h cm(2)). Water resorption and I SC were reduced by almost 80% in the presence of the bulk Cl(-) channel inhibitor 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) or amiloride, a specific inhibitor of epithelial sodium channel (ENaC). However, water resorption and I SC were only moderately affected by forskolin or cystic fibrosis transmembrane regulator (CFTR) channel inhibitors (CFTRinh-172 and glybenclamide). In line with previous studies, we demonstrate that water resorption depends on ENaC, and CFTR channels have only a minor but probably modulating effect on water resorption. However, the major ENaC-mediated water resorption depends on an apical non-CFTR Cl(-) conductance.

In situ synthesis of bare gold nanoparticles mediated by stainless steel as reducing agent was monitored via infrared attenuated total reflection (IR-ATR) spectroscopy. Gold nanoparticles were directly synthesized within the liquid cell of the ATR unit taking immediate advantage of the stainless steel walls of the ATR cell. As nanoparticles were formed, a layer of particles was deposited at the SiO2 ATR waveguide surface. Incidentally, the absorption bands of water increased resulting from surface-enhanced infrared absorption (SEIRA) effects arising from the presence of the gold nanoparticles within the evanescent field. Next to the influence of the Au(III) precursor concentration and the temperature, the suitability of IR-ATR spectroscopy as an innovative tool for investigating changes of nanoparticles in solution, including their aggregation promoted by an increase of the ionic strength or via a pH decrease, and for detailing the sedimentation process of gold nanoparticles was confirmed.

Hydrogen sulfide is a highly corrosive, harmful, and toxic gas produced under anaerobic conditions within industrial processes or in natural environments, and plays an important role in the sulfur cycle. According to the U.S. Occupational Safety and Health Administration (OSHA), the permissible exposure limit (during 8 hours) is 10 ppm. Concentrations of 20 ppm are the threshold for critical health issues. In workplace environments with human subjects frequently exposed to H2S, e.g., during petroleum extraction and refining, real-time monitoring of exposure levels is mandatory. Sensors based on electrochemical measurement principles, semiconducting metal-oxides, taking advantage of their optical properties, have been described for H2S monitoring. However, extended response times, limited selectivity, and bulkiness of the instrumentation are common disadvantages of the sensing techniques reported to date. Here, we describe for the first time usage of a new generation of compact gas cells, i.e., so-called substrate-integrated hollow waveguides (iHWGs), combined with a compact Fourier transform infrared (FTIR) spectrometer for advanced gas sensing of H2S. The principle of detection is based on the immediate UV-assisted conversion of the rather weak IR-absorber H2S into much more pronounced and distinctively responding SO2. A calibration was established in the range of 10-100 ppm with a limit of detection (LOD) at 3 ppm, which is suitable for occupational health monitoring purposes. The developed sensing scheme provides an analytical response time of less than 60 seconds. Considering the substantial potential for miniaturization using e.g., a dedicated quantum cascade laser (QCL) in lieu of the FTIR spectrometer, the developed sensing approach may be evolved into a hand-held instrument, which may be tailored to a variety of applications ranging from environmental monitoring to workplace safety surveillance, process analysis and clinical diagnostics, e.g., breath analysis.

A simple, environmentally friendly, one-pot method to synthesize highly stable bare gold nanoparticles (AuNPs) has been developed. AuNPs have been synthesized from tetrachloroauric acid solution using steel or stainless steel as solid reducing agent, which can be reused. The proposed method yields bare gold nanoparticles at atmospheric pressure and room temperature for potentially producing large quantities. The obtained AuNPs have been characterized by SEM, TEM and AFM finding an average diameter of around 20 nm, polygonal yet nearly spherical shape and a narrow size distribution. The mechanism of reaction has been investigated by UV-vis spectroscopy, ICP-OES and EDX analysis. The obtained dispersed gold nanoparticles proved to be stable if stored a 4 °C for over four months without the addition of a stabilizing agent. Their analytical potential as SERS substrate has been demonstrated and their performance compared with that showed by citrate-coated gold nanoparticles. Thanks to their unique properties, their use as analytical tools provides analytical processes with enhanced selectivity and precision., (© 2013 Elsevier B.V. All rights reserved.)

A new generation of hollow waveguide (HWG) gas cells of unprecedented compact dimensions facilitating low sample volumes suitable for broad- and narrow-band mid-infrared (MIR; 2.5-20 μm) sensing applications is reported: the substrate-integrated hollow waveguide (iHWG). iHWGs are layered structures providing light guiding channels integrated into a solid-state substrate material, which are competitive if not superior in performance to conventional leaky-mode fiber optic silica HWGs having similar optical pathlengths. In particular, the provided flexibility in device and optical design and the wide variety of manufacturing strategies, substrate materials, access to the optical channel, and optical coating options highlight the advantages of iHWGs in terms of robustness, compactness, and cost-effectiveness. Finally, the unmatched modularity of this novel waveguide approach facilitates tailoring iHWGs to almost any kind of gas sensor technology providing adaptability to the specific demands of a wide range of sensing scenarios. Device fabrication is demonstrated for the example of a yin-yang-shaped gold-coated iHWG fabricated within an aluminum substrate with a footprint of only 75 mm × 50 mm × 12 mm (L × W × H), yet providing a nominal optical absorption path length of more than 22 cm. The analytical utility of this device for advanced MIR gas sensing applications is demonstrated for the gaseous constituents butane, carbon dioxide, cyclopropane, isobutylene, and methane.

Despite providing the opportunity for directly sensing molecular constituents with inherent fingerprint specificity in the 2.5-20 μm spectral regime, mid-infrared optical sensing technologies have not yet achieved the same penetration in waveguide-based chem/bio sensing compared to related sensing schemes operating at visible and near-infrared frequencies. In this review, current advances in mid-infrared chem/bio sensor technology will be highlighted and contrasted with the prevalent bottlenecks that have to date limited a more widespread adoption of mid-infrared sensing devices. However, with the increasing availability of advanced light sources such as quantum cascade lasers and the advent of on-chip semiconductor waveguide technologies, a prosperous future of this sensing concept for label-free detection in environmental analysis, process monitoring, and bioanalytics is perceived.

A novel analytical platform combining infrared attenuated total reflection (IR-ATR) spectroelectrochemistry (SE) with atomic force microscopy (AFM) using a boron-doped diamond (BDD)-modified ATR crystal is presented. The utility of this combination is demonstrated investigating the electrodeposition of a polymer film via IR spectroscopy, while the surface modification is simultaneously imaged by AFM. The ATR waveguide consists of a single-crystal intrinsic diamond overgrown with a homoepitaxial BDD layer (thickness: ∼100 nm, boron content: ∼5 × 10(20) cm(-3)) to provide electric conductivity. The diamond ATR crystal is shaped in the form of a hemisphere with a beveled top and an octahedronal surface area of approximately 400 μm(2). To demonstrate combined IR-ATR-SE-AFM measurements, the electro-polymerization of 3,4-ethylenedioxothiophene (EDOT) was selected as a model system. Depositions were obtained from aqueous solutions, while changes in IR signature, topography, and electrochemical behavior were recorded in situ and simultaneously during the polymerization process.

We report the first planar waveguides made from mercury-cadmium-telluride (MCT)-a material to date exclusively used for mid-infrared (MIR) detector elements-serving as on-chip MIR evanescent field transducers in combination with tunable quantum cascade lasers (tQCLs) emitting in the spectral regime of 5.78-6.35 μm. This novel MIR sensing approach utilizes structured MCT chips fabricated via molecular beam epitaxy (MBE) as waveguide enabling sensing via evanescent field absorption spectroscopy, as demonstrated by the detection of 1 nL of acetone. Complementary finite difference time domain (FDTD) simulations fit well with the experimentally obtained data and predict an improvement of the limit of detection by at least 2 orders of magnitude upon implementation of thinner MCT waveguides. With the first demonstration of chemical sensing using on-chip MCT waveguides, monolithically fabricated IR sensing systems directly interfacing the waveguide with the MCT detector element may be envisaged.

Ozone is a strong oxidant that is globally used as disinfection agent for many purposes including indoor building air cleaning, during food preparation procedures, and for control and killing of bacteria such as E. coli and S. aureus. However, it has been shown that effective ozone concentrations for controlling e.g., microbial growth need to be higher than 5 ppm, thereby exceeding the recommended U.S. EPA threshold more than 10 times. Consequently, real-time monitoring of such ozone concentration levels is essential. Here, we describe the first online gas sensing system combining a compact Fourier transform infrared (FTIR) spectrometer with a new generation of gas cells, a so-called substrate-integrated hollow waveguide (iHWG). The sensor was calibrated using an UV lamp for the controlled generation of ozone in synthetic air. A calibration function was established in the concentration range of 0.3-5.4 mmol m⁻³ enabling a calculated limit of detection (LOD) at 0.14 mmol m⁻³ (3.5 ppm) of ozone. Given the adaptability of the developed IR sensing device toward a series of relevant air pollutants, and considering the potential for miniaturization e.g., in combination with tunable quantum cascade lasers in lieu of the FTIR spectrometer, a wide range of sensing and monitoring applications of beyond ozone analysis are anticipated.

Surface imprinted polymers allow accessibility of the selective binding sites to large molecules such as proteins. In this work, small polymer particles offering a large surface area were prepared via miniemulsion polymerization in the presence of pepsin serving as a template molecule. The influence of four different functional monomers and of the amount of the template on the imprinting effect of pepsin was investigated. After the miniemulsion polymerization and a washing step, stable polymer suspensions with an average particle diameter of 400-600 nm and a specific surface area of 30-65 m 2 g -1 were obtained. The results of detailed rebinding experiments revealed that the highest imprinting effect was achieved with (3-acrylamidopropyl)trimethylammonium chloride as a functional monomer and a high amount of the template. These polymer particles also showed selectivity for pepsin against various proteins. This approach provides a fundamental step towards the development of synthetic protein receptors and protein scavenger materials useful in biomimetic assays and for clean-up in biotechnology.

The use of chemometrics in order to improve the molecular selectivity of infrared (IR) spectra has been evaluated using classic least squares (CLS), partial least squares (PLS), science-based calibration (SBC), and multivariate curve resolution-alternate least squares (MCR-ALS) techniques for improving the discriminatory and quantitative performance of infrared hollow waveguide gas sensors. Spectra of mixtures of isobutylene, methane, carbon dioxide, butane, and cyclopropane were recorded, analyzed, and validated for optimizing the prediction of associated concentrations. PLS, CLS, and SBC provided equivalent results in the absence of interferences. After addition of the spectral characteristics of water by humidifying the sample mixtures, CLS and SBC results were similar to those obtained by PLS only if the water spectrum was included in the calibration model. In the presence of an unknown interferant, CLS revealed errors up to six times higher than those obtained by PLS. However, SBC provided similar results compared to PLS by adding a measured noise matrix to the model. Using MCR-ALS provided an excellent estimation of the spectra of the unknown interference. Furthermore, this method also provided a qualitative and quantitative estimation of the components of an unknown set of samples. In summary, using the most suitable chemometrics approach could improve the selectivity and quality of the calibration model derived for a sensor system, and may avoid the need to analyze expensive calibration data sets. The results obtained in the present study demonstrated that (1) if all sample components of the system are known, CLS provides a sufficiently accurate solution; (2) the selection between PLS and SBC methods depends on whether it is easier to measure a calibration data set or a noise matrix; and (3) MCR-ALS appears to be the most suitable method for detecting interferences within a sample. However, the latter approach requires the most extensive calculations and may thus result in limited temporal resolution, if the concentration of a component should be continuously monitored.

The phytopathogen Xylella fastidiosa grows as a biofilm causing vascular occlusion and consequently nutrient and water stress in different plant hosts by adhesion on xylem vessel surfaces composed of cellulose, hemicellulose, pectin and proteins. Understanding the factors which influence bacterial adhesion and biofilm development is a key issue in identifying mechanisms for preventing biofilm formation in infected plants. In this study, we show that X. fastidiosa biofilm development and architecture correlate well with physicochemical surface properties after interaction with the culture medium. Different biotic and abiotic substrates such as silicon (Si) and derivatized cellulose films were studied. Both biofilms and substrates were characterized at the micro- and nanoscale, which corresponds to the actual bacterial cell and membrane/ protein length scales, respectively. Our experimental results clearly indicate that the presence of surfaces with different chemical composition affect X. fastidiosa behavior from the point of view of gene expression and adhesion functionality. Bacterial adhesion is facilitated on more hydrophilic surfaces with higher surface potentials; XadA1 adhesin reveals different strengths of interaction on these surfaces. Nonetheless, despite different architectural biofilm geometries and rates of development, the colonization process occurs on all investigated surfaces. Our results univocally support the hypothesis that different adhesion mechanisms are active along the biofilm life cycle representing an adaptation mechanism for variations on the specific xylem vessel composition, which the bacterium encounters within the infected plant.

Grating couplers for planar silver halide waveguides were designed and fabricated by using focused ion beam (FIB) milling technology, facilitating coupling of mid-infrared radiation from quantum cascade lasers into thin-film waveguide structures. An optimized rectangular grating structure for an emitted wavelength of 10.4 μm, with a grating constant of 16.4 μm was integrated into a silver halide waveguide substrate via an optimized FIB fabrication procedure. Efficient incoupling and radiation propagation through the waveguide was confirmed by analyzing droplets of acetic acid at different concentrations, deposited at the waveguide surface via evanescent field absorption spectroscopy.

The paper proposes a simple and portable approach for the surface enhanced Raman scattering (SERS) spectroscopy in situ determination of carboxylated single walled carbon nanotubes (SWNTs) in river water samples. The method is based on the subsequent microfiltration of a bare gold nanoparticles solution and the water sample containing soluble carbon nanotubes by using a home-made filtration device with a small filtration diameter. An acetate cellulose membrane with a pore size of 0.2 μm first traps gold nanoparticles to form the SERS-active substrate and then concentrates the carbon nanotubes. The measured SERS intensity data were closely fit with a Langmuir isotherm. A portable Raman spectrometer was employed to measure SERS spectra, which enables in situ determination of SWNTs in river waters. The limit of detection was 10 μg L(-1). The precision, for a 10 mg L(-1) concentration of carbon nanotubes, is 1.19% intra-membrane and 10.5% inter-membrane., (Copyright © 2013. Published by Elsevier B.V.)

This paper describes the determination of bisphenol A (BPA) in milk samples, using a novel molecularly imprinted polymer. The imprinted polymer was developed using a rational design approach, and pre-polymerization interactions were investigated using molecular dynamics simulations and X-ray crystallography. A hydroquinone-imprinted polymer was used for solid phase extraction (SPE) clean-up of samples. BPA was quantified by high performance liquid chromatography (HPLC) and fluorescence (FLD) detection. Following validation, the method described was capable of determining bisphenol A in milk down to a limit of detection of 1.32μgkg(-1). The method was applied to a survey (n=27) of commercial milk products; BPA was detected in one of the samples, at a level of 176μgkg(-1). Test results were confirmed by a parallel UHPLC-MS/MS analytical method. This demonstrates the utility of the hydroquinone-imprinted polymer for application to selective sample clean-up and analysis of bisphenol A in milk, avoiding possible detrimental affects associated with template bleeding and without the need for expensive or difficult-to-obtain template., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The mechanically induced release of adenosine-5'-triphosphate (ATP) from osteoblastic cells (MC3T3-E1) was measured in real time. A stretching device integrated into scanning electrochemical microscopy was developed to apply controlled mechanical strain to MC3T3-E1 cells. For ATP secretion, a stepwise yet uniform mechanical stress was imposed onto MC3T3-E1 cells. The ATP biosensors were positioned at a distance of approximately 30-40 μm above the cell surface. Calibration functions were recorded prior to the cell measurements and revealed a linear response up to 40 μM with a sensitivity of 1-5pA/μM ATP. Stretching MC3T3-E1 cells up to 21% resulted in a concentration of 30.57±4.82 μM of extracellular ATP (N=12) detected above the cell surface. As a control experiment, nifedipine, a L-type voltage sensitive calcium channel (L-VSCC) inhibitor was applied, which blocks Ca(2+)entry from the outer medium into the cell. Inhibition resulted in a significantly smaller amount of released ATP, i.e., 7.08±1.93 μM ATP (N=10). Further control experiments with glucose microbiosensors did not yield significant changes of the baseline current (N=8)., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Lung epithelia regulate the water flux between gas filled airways and the interstitial compartment in order to maintain organ function. Current methodology to assess transepithelial water transport is limited. We present a D2O dilution method to quantify submicroliter volumes of aqueous solutions on epithelial cell layers. Evaluating D2O/H2O mixtures using mid-infrared (2-25 μm) attenuated total reflection (ATR) spectroscopy, with a resolution of 0.06% vol/vol change, corresponding to 24 nL, was achieved. Using this method, we demonstrate that water transport across NCI-H441 lung epithelial cell layers and apical surface liquid (ASL) volumes are coupled to dexamethasone dependent amiloride-sensitive ion transport. However, contrary to current dogma, electrogenic transport is not rate-limiting for water transport. This clearly indicates the need to directly assess net water rather than ion transport across epithelial cell layers. The presented D2O dilution method enables such direct and quick quantification of transepithelial water transport with high resolution.

The (12)CO2/(13)CO2 isotope ratio is a well-known marker in breath for a variety of biochemical processes and enables monitoring, e.g., of the glucose metabolism during sepsis. Using animal models-here, at a mouse intensive care unit-the simultaneous determination of (12)CO2 and (13)CO2 within small volumes of mouse breath was enabled by coupling a novel low-volume hollow waveguide gas cell to a compact Fourier transform infrared spectrometer combined with multivariate data evaluation based on partial least squares regression along with optimized data preprocessing routines.

We report the design, fabrication, and first functional verification of mid-infrared (MIR; 3-12 μm) Mach-Zehnder interferometers (MZIs). The developed MIR-MZIs are entirely chip-integrated solid-state devices based on GaAs/AlGaAs technology waveguide fabricated via conventional optical lithography and reactive ion etching (RIE). Thus, fabricated MIR-MZIs were combined with a broadly tunable quantum cascade laser (tQCL) providing a wavelength coverage of 5.78-6.35 μm. MIR-MZIs have been designed with a waveguide width of 5 μm to ensure single mode behavior, avoiding optically undefined interference patterns. Several structures with different opening angles of the Y-junction were fabricated and tested for maximizing IR radiation throughput. This study demonstrates the feasibility of the very first chip-integrated mid-infrared Mach-Zehnder structures via interference patterns produced by minute amounts of water deposited at different positions of the MIR-MZI structure.

With the availability of broadly tunable external cavity quantum cascade lasers (EC-QCLs), particularly bright mid-infrared (MIR; 3-20 μm) light sources are available offering high spectral brightness along with an analytically relevant spectral tuning range of >2 μm. Accurate isotope ratio determination of (12)CO2 and (13)CO2 in exhaled breath is of critical importance in the field of breath analysis, which may be addressed via measurements in the MIR spectral regime. Here, we combine for the first time an EC-QCL tunable across the (12)CO2/(13)CO2 spectral band with a miniaturized hollow waveguide gas cell for quantitatively determining the (12)CO2/(13)CO2 ratio within the exhaled breath of mice. Due to partially overlapping spectral features, these studies are augmented by appropriate multivariate data evaluation and calibration techniques based on partial least-squares regression along with optimized data preprocessing. Highly accurate determinations of the isotope ratio within breath samples collected from a mouse intensive care unit validated via hyphenated gas chromatography-mass spectrometry confirm the viability of IR-HWG-EC-QCL sensing techniques for isotope-selective exhaled breath analysis.

Asthma and chronic obstructive pulmonary disease (COPD) are distinct but clinically overlapping airway disorders which often create diagnostic and therapeutic dilemmas. Current strategies to discriminate these diseases are limited by insensitivity and poor performance due to biologic variability. We tested the hypothesis that a gas chromatograph/differential mobility spectrometer (GC/DMS) sensor could distinguish between clinically well-defined groups with airway disorders based on the volatile organic compounds (VOCs) obtained from exhaled breath. After comparing VOC profiles obtained from 13 asthma, 5 COPD and 13 healthy control subjects, we found that VOC profiles distinguished asthma from healthy controls and also a subgroup of asthmatics taking the drug omalizumab from healthy controls. The VOC profiles could not distinguish between COPD and any of the other groups. Our results show a potential application of the GC/DMS for non-invasive and bedside diagnostics of asthma and asthma therapy monitoring. Future studies will focus on larger sample sizes and patient cohorts.

The structural integrity and protection of bacterial biofilms are intrinsically associated with a matrix of extracellular polymeric substances (EPS) produced by the bacteria cells. However, the role of these substances during biofilm adhesion to a surface remains largely unclear. In this study, the influence of EPS on Xylella fastidiosa biofilm formation was investigated. This bacterium is associated with economically important plant diseases; it presents a slow growth rate and thus allows us to pinpoint more precisely the early stages of cell-surface adhesion. Scanning electron microscopy and atomic force microscopy show evidence of EPS production in such early stages and around individual bacteria cells attached to the substrate surface even a few hours after inoculation. In addition, EPS formation was investigated via attenuated total reflectance (ATR) Fourier transform infrared spectroscopy (FTIR). To this end, X. fastidiosa cells were inoculated within an ATR liquid cell assembly. IR-ATR spectra clearly reveal EPS formation already during the early stages of X. fastidiosa biofilm formation, thereby providing supporting evidence for the hypothesis of the relevance of the EPS contribution to the adhesion process., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Chlorinated aliphatic hydrocarbons and chlorinated aromatic hydrocarbons (CHCs) are toxic and carcinogenic contaminants commonly found in environmental samples, and efficient online detection of these contaminants is still challenging at the present stage. Here, we report an advanced Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR) sensor for in-situ and simultaneous detection of multiple CHCs, including monochlorobenzene, 1,2-dichlorobenzene, 1,3-dichlorobenzene, trichloroethylene, perchloroethylene, and chloroform. The polycrystalline silver halide sensor fiber had a unique integrated planar-cylindric geometry, and was coated with an ethylene/propylene copolymer membrane to act as a solid phase extractor, which greatly amplified the analytical signal and contributed to a higher detection sensitivity compared to the previously reported sensors. This system exhibited a high detection sensitivity towards the CHCs mixture at a wide concentration range of 5~700 ppb. The FTIR-ATR sensor described in this study has a high potential to be utilized as a trace-sensitive on-line device for water contamination monitoring.

Focused ion beam/scanning electron microscopy (FIB/SEM) tomography is a novel powerful approach for three-dimensional (3D) imaging of biological samples. Thereby, a sample is repeatedly milled with the focused ion beam (FIB) and each newly produced block face is imaged with the scanning electron microscope (SEM). This process can be repeated ad libitum in arbitrarily small increments allowing 3D analysis of relatively large volumes such as eukaryotic cells. High-pressure freezing and freeze substitution, on the other hand, are the gold standards for electron microscopic preparation of whole cells. In this work, we combined these methods and substantially improved resolution by using the secondary electron signal for image formation. With this imaging mode, contrast is formed in a very small, well-defined area close to the newly produced surface. By using this approach, small features, so far only visible in transmission electron microscope (TEM) (e.g., the two leaflets of the membrane bi-layer, clathrin coats and cytoskeletal elements), can be resolved directly in the FIB/SEM in the 3D context of whole cells.

In molecular imprinting the porogen plays a decisive role, as it not only affects the physical properties of the resulting polymer including its porosity, the specific surface area, and the swelling behavior, but also governs the stability of the prepolymerization complex, which in turn decisively determines the recognition properties of the resulting molecularly imprinted polymer (MIP). In this study, the influence of the porogen on the selectivity of MIPs was investigated. Therefore, bulk MIPs against 4-nitrophenol using 4-vinylpyridine (4-VP) as functional monomer and ethylene glycol dimethacrylate (EDMA) as crosslinker were prepared in acetonitrile and chloroform. The recognition properties of both MIPs were evaluated during chromatographic studies using the respective porogenic solvents as mobile phase for both MIPs. Along with the characterization of the morphology of the obtained polymers via SEM and BET analysis, the beneficial nature of chloroform as porogen for imprinting 4-NP was experimentally demonstrated and verified by findings obtained from complementary molecular dynamics simulations. Moreover, the application of chloroform as mobile phase for the MIP prepared in acetonitrile and vice versa clearly demonstrated the dependence of the resulting recognition properties on the selection of the mobile phase., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Hybrid atomic force microscopy (AFM)-fluorescence microscopy (FM) investigation of exocytosis in lung epithelial cells (ATII cells) allows the detection of individual exocytic events by FM, which can be simultaneously correlated to structural changes in individual cells by AFM. Exocytosis of lamellar bodies (LBs) represents a slow form of exocytosis found in many non-neuronal cells. Exocytosis of LBs, following stimulation with adenosine-5'-triphosphate (ATP) and phorbol 12-myristate 13-acetate (PMA), results in a cation influx via P2X(4) receptors at the site of LB fusion with the plasma membrane (PM), which should induce a temporary increase in cell height/volume. AFM measurements were performed in single-line scans across the cell surface. Five minutes after stimulation, ATII cells revealed a cell height and volume increase of 13.7% ± 4.1% and 15.9 ± 4.8% (N = 9), respectively. These transient changes depend on exocytic LB-PM fusion. Nonstimulated cells and cells lacking LB fusions did not show a significant change in cell height/volume (N = 8). In addition, a cell height decrease was observed in ATII cells stimulated by uridine-5'-triphosphate (UTP) and PMA, agonists inducing LB fusion with the PM, but not activation of P2X(4) receptors. The cell height and volume decreased by -8.6 ± 3.6% and -11.2 ± 3.9% (N = 5), respectively. Additionally, low force contact and dynamic mode AFM imaging of cell areas around the nucleus after stimulation with ATP/PMA was performed. Fused LBs are more pronounced in AFM topography images compared to nonfused LBs, concluding that different "dynamic states" of LBs or locations from the PM are captured during imaging.

We demonstrate ultra-sensitive chemical sensing in the mid-infrared spectral regime with a combination of quantum cascade lasers (QCLs) with GaAs/Al(0.2)Ga(0.8)As strip waveguides fabricated via metal-organic vapor-phase epitaxy (MOVPE) and reactive ion etching (RIE) using evanescent field absorption spectroscopy. These strip waveguides have been designed with a width of 200 μm, thereby facilitating 2-D confinement and mode-matched propagation of mid-infrared radiation emitted from a distributed feedback (DFB) QCL at a wavelength of 10.3 μm. Acetic anhydride was detected with a limit of detection (LOD) of 18 pL (19.4 ng) deposited at the waveguide surface by overlapping of the vibrational absorption of the methyl group with the emission frequency of the QCL. The obtained results indicate a remarkable enhancement in sensitivity by three orders of magnitude compared to previously reported multimode planar silver halide waveguides. Further reduction of the waveguide strip width to 50 μm resulted in an additional sensitivity enhancement yielding a calculated LOD of 0.05 pL for the exemplary analyte acetic anhydride, which is among the most sensitive evanescent field absorption measurements with a miniaturized mid-infrared sensor system reported to date.

For facilitating the identification of appropriate functionalities that may serve as a binding motif of functional monomers, a selection strategy based on high-throughput screening of the binding properties of readily available sorbent materials has been developed. Thereby, the affinity of such ligands to the protein of interest may be rapidly determined. From these studies, it is anticipated that ligand functionalities will be derived, which may lead to advanced selection and design of dedicated functional monomers suitable for decorating the surface of a scavenger material. Thus, specific binding of the target protein of interest should be enabled even in complex solutions such as e.g., biotechnologically relevant cell lysates. In the present contribution, an automated screening method for studying ligand interactions of selected sorbent materials with pepsin - a protein of the protease family - was developed. Aqueous buffer solutions containing pepsin at known constant concentration were pipetted through an array of miniaturized chromatographic solid phase extraction (SPE) columns containing a variety of sorbent materials, and the eluted solutions were analyzed by UV/vis spectroscopy. The established screening protocol was validated against resin materials of known interaction with pepsin. Finally, the developed screening strategy was adapted for a robot system enabling high-throughput screening for a wide variety of sorbent materials and ligand functionalities in a fully automated approach. The obtained results clearly indicate that the established screening routine provides valuable data for characterizing resin-immobilized ligands, and their affinity toward pepsin., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The determination of trace amounts of oil in water facilitates the forensic analysis on the presence and origin of oil in the aqueous environment. To this end, the present study focuses on direct sensing schemes for quantifying trace amounts of oil in water using mid-infrared (MIR) evanescent field absorption spectroscopy via fiberoptic chemical sensors. MIR transparent silver halide fibers were utilized as optical transducer for interrogating oil-in-water emulsions via the evanescent field emanating from the waveguide surface, and penetrating the surrounding aqueous environment by a couple of micrometers. Unmodified fibers and fibers surface-modified with grafted epoxidized polybutadiene layers enabled the direct detection of crude oil in a deionized water matrix at the ppm level to ppb concentration level, respectively. Thus, direct chemical sensing of crude oil IR signatures without any sample preparation as low as 46 ppb was achieved with a response time of a few seconds., (© 2011 American Chemical Society)

Water is a common contaminant in a variety of industrial oils and petroleum products. Thus, the detection of water in these products is of substantial relevance. Hence, this study focuses on quantifying trace amounts of water in hydrocarbons using hexane as a model system for industrial oils and petroleum matrices via mid-infrared (MIR) evanescent field absorption spectroscopy. A silver halide fiberoptic waveguide was used to interrogate in situ water-in-hexane emulsions. Either unmodified fibers or waveguides surface-modified with polyacrylic acid layers were used. The limits of detection (LOD) and limits of quantification (LOQ) of water in hexane utilizing tin-crosslinked polyacrylic acid modified fibers were 76 and 170 ppm, respectively. Consequently, the IR absorption signature of water in hexane is detectable at concentrations as low as 10 ppm. The proposed fiberoptic sensing strategy requires a single measurement only, requires no sample preparation, and thus has potential for the direct in situ detection and monitoring of water in industrial oils and petroleum products.

Gas hydrates are crystalline structures comprising a guest molecule surrounded by a water cage, and are particularly relevant due to their natural occurrence in the deep sea and in permafrost areas. Low molecular weight molecules such as methane and carbon dioxide can be sequestered into that cage at suitable temperatures and pressures, facilitating the transition to the solid phase. While the composition and structure of gas hydrates appear to be well understood, their formation and dissociation mechanisms, along with the dynamics and kinetics associated with those processes, remain ambiguous. In order to take advantage of gas hydrates as an energy resource (e.g., methane hydrate), as a sequestration matrix in (for example) CO(2) storage, or for chemical energy conservation/storage, a more detailed molecular level understanding of their formation and dissociation processes, as well as the chemical, physical, and biological parameters that affect these processes, is required. Spectroscopic techniques appear to be most suitable for analyzing the structures of gas hydrates (sometimes in situ), thus providing access to such information across the electromagnetic spectrum. A variety of spectroscopic methods are currently used in gas hydrate research to determine the composition, structure, cage occupancy, guest molecule position, and binding/formation/dissociation mechanisms of the hydrate. To date, the most commonly applied techniques are Raman spectroscopy and solid-state nuclear magnetic resonance (NMR) spectroscopy. Diffraction methods such as neutron and X-ray diffraction are used to determine gas hydrate structures, and to study lattice expansions. Furthermore, UV-vis spectroscopic techniques and scanning electron microscopy (SEM) have assisted in structural studies of gas hydrates. Most recently, waveguide-coupled mid-infrared spectroscopy in the 3-20 μm spectral range has demonstrated its value for in situ studies on the formation and dissociation of gas hydrates. This comprehensive review summarizes the importance of spectroscopic analytical techniques to our understanding of the structure and dynamics of gas hydrate systems, and highlights selected examples that illustrate the utility of these individual methods.

Mouse sepsis models are used to gain insight into the complex processes involved with patients suffering from glucose metabolism disorders. Measuring the expiratory release of (13)CO(2) after administering stable labeled (13)C(6)-glucose enables assessment of the in vivo integrity and functionality of key metabolic processes. In the present study, we demonstrate that Fourier transform infrared spectroscopy operating in the mid-infrared spectral regime (2-20 μm) combined with hollow waveguide gas sensing modules simultaneously serving as a miniaturized gas cell and as a waveguide are capable of quantitatively monitoring (13)CO(2) enrichment levels in low volume mouse breath samples.

This communication presents a novel label-free biosensing method to monitor DNA hybridization via infrared attenuated total reflection (IR-ATR) spectroscopy using surface-modified ZnSe waveguides. Well-defined carboxyl-terminated monolayers were formed at H-terminated ZnSe by direct photochemical activation. Chemical activation of the acidic function was obtained by using succinimide/carbodiimide linkers. The sequential surface modification reactions were characterized by XPS and IR-ATR spectroscopy. Finally, a single stranded DNA probe with a C6-NH(2) 5' modifier was coupled to the ester-terminated surface via peptide bonding, and the hybridization of the immobilized DNA sequence with its complementary strand was directly evaluated by IR-ATR spectroscopy in the mid-infrared (MIR) spectral regime (3-20 μm) without requiring an additional label. A shift of the vibrational modes corresponding to the phosphodiester and deoxyribose structures of the DNA backbone was observed. Hence, this approach substantiates a novel strategy for label-free DNA detection utilizing mid-infrared spectroscopy as the optical sensing platform.

The development of a hollow core waveguide (HWG) gas sensor in combination with a fast and compact near-infrared (NIR) spectrometer is presented. The spectrometer operates in the spectral range of 1200-1400 nm and may thus be applied for the detection of gas-phase analytes providing NIR absorptions in that spectral window such as, e.g., methane. Since mid-infrared spectroscopy in combination with HWGs has already been successfully demonstrated for probing hydrocarbons in the gas phase, the present study investigates the achievable sensitivity in the NIR spectral regime. Methane has been selected as an exemplary analyte due to the fact that it shows strong absorption features in the mid-infrared (mid-IR) fingerprint area, but also overtone bands in the NIR. Since the HWG simultaneously serves as a miniaturized absorption gas cell and as an optical waveguide for NIR radiation, a compact yet optical and cost-efficient sensor device was established providing an interesting alternative in target sensing for mid-IR devices. The achieved limit of detection (LOD) was 5.7% (vol./vol.) methane for a 9.5 cm long HWG, 1.6% (vol./vol.) methane for a 39.1 cm long HWG, and 1.3% (vol./vol.) methane for a setup using a 77.4 cm long HWG, which provides the most practical HWG dimensions among the three investigated setups. Limit of quantitation (LOQ) values were calculated at 20.1% (vol./vol.) methane, 8.7% (vol./vol.) methane, and 5.6% (vol./vol.) methane, respectively.

Simultaneous and molecularly selective parts-per-billion detection of benzene, toluene, and xylenes (BTX) using a thermal desorption (TD)-FTIR hollow waveguide (HWG) trace gas sensor is demonstrated here for the first time combining laboratory calibration with real-world sample analysis in field. A calibration range of 100-1000 ppb analyte/N(2) was developed and applied for predicting the concentration of blinded environmental air samples within the same concentration range, and demonstrate close agreement with the validation method used here, GC-FID. The analyte concentration prediction capability of the TD-FTIR-HWG trace gas sensor also compares well with the industrial standard and other experimental techniques including GC-PID, ultrafast GC-FID, and GC-DMS, which were simultaneously operated in the field. With the advent of a quantum cascade laser with emission frequencies specifically tailored to efficiently overlap benzene absorption as the most relevant analyte, the overall sensor footprint could be considerably reduced to ultimately yield hand-held trace gas sensors facilitating direct and real-time detection of BTX in air down to low ppb levels.

Biofilms are complex microbial communities with important biological functions including enhanced resistance against external factors like antimicrobial agents. The formation of a biofilm is known to be strongly dependent on substrate properties including hydrophobicity/hydrophilicity, structure, and roughness. The adsorption of (macro)molecules on the substrate, also known as conditioning film, changes the physicochemical properties of the surface and affects the bacterial adhesion. In this study, we investigate the physicochemical changes caused by Periwinkle wilt (PW) culture medium conditioning film formation on different surfaces (glass and silicon) and their effect on X. fastidiosa biofilm formation. Contact angle measurements have shown that the film formation decreases the surface hydrophilicity degree of both glass and silicon after few hours. Atomic force microscopy (AFM) images show the glass surface roughness is drastically reduced with conditioning film formation. First-layer X. fastidiosa biofilm on glass was observed in the AFM liquid cell after a period of time similar to that determined for the hydrophilicity changes. In addition, attenuation total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy supports the AFM observation, since the PW absorption spectra increases with time showing a stronger contribution from the phosphate groups. Although hydrophobic and rough surfaces are commonly considered to increase bacteria cell attachment, our results suggest that these properties are not as important as the surface functional groups resulting from PW conditioning film formation for X. fastidiosa adhesion and biofilm development., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Breath analysis has received attention as a noninvasive diagnostic tool with increasing research into its potential usefulness. We are investigating the utility of the analysis of breath volatile organic compounds (VOCs) as an effective modality for breast cancer (BC) detection and monitoring by collecting breath samples with a simple portable device to determine whether BC patients have breath VOCs distinct from those in healthy volunteers. We prospectively enrolled 20 healthy volunteers and 20 newly diagnosed stage II-IV BC patients. The study subjects deeply exhaled into a commercially available Teflon/valved breath sampler equipped with a rapid passive diffusive sampler five times at 5-minute intervals trapping alveolar breath VOCs. The exhaled breath samples were analyzed by thermal desorption/gas chromatography/mass spectrometry monitoring 383 VOCs in the breath of both populations. Our results indicate that aggregate low-dimensional summaries and compound quantities result in specific patterns that can confirm BC. We found a definite clustering of the presence of BC from cancer-free points. Overall sensitivity was 72 per cent and specificity was 64 per cent resulting in a correct classification rate of approximately 77 per cent. Our data show promising evidence that BC patients can be differentiated from healthy volunteers through distinct breath VOCs.

Interactions of ozone with organic precursors during water treatment may generate carcinogenic N-nitrosodimethylamine (NDMA) byproduct. This study investigates the reaction mechanisms responsible for NDMA formation from ozonation of the commonly used poly(diallyldimethylammonium chloride) (polyDADMAC) coagulant. Upon ozonation, polyDADMAC yields the highest amount of NDMA among several water treatment polymers, including polyamines and cationic polyacrylamides. Ozonation transforms polyDADMAC to dimethylamine (DMA) and NDMA formation is correlated to polyDADMAC degradation and DMA release. Hydroxyl radicals generated from ozone play an important role in the degradation of polyDADMAC's quaternary ammonium ring groups and subsequent release of secondary amine. Although nitrite and formaldehyde are detected as ozonation products of DMA and polyDADMAC, contribution of formaldehyde-enhanced nitrosation pathway is determined to be insignificant in NDMA formation. In contrast, reaction of hydroxylamine, another ozonation product of DMA, with DMA in the presence of ozone is deemed critical in the formation of NDMA during ozonation. The study results show that that contact of polyDADMAC with ozone will lead to release of the more potent NDMA precursor DMA but may not generate a significant amount of NDMA under typical drinking water treatment conditions due to low yield. The mechanistic understanding from this study can help develop source control strategies for minimization of NDMA formation risk at water and wastewater utilities.

This contribution describes the development of nitrogen-doped diamond-like carbon (N-DLC) thin films for multi-reflection mid-infrared (MIR) attenuated total reflectance (IR-ATR) spectroelectrochemistry. N-DLC coatings were deposited using pulsed laser deposition (PLD) involving the ablation of a high purity graphite target. The DLC matrix was further modified by ablating the target in the presence of nitrogen gas. This technique offers the advantage of depositing thin films at room temperature, thereby enabling coating of temperature-sensitive substrates including e.g., MIR waveguides. The resulting films were analyzed with X-ray photoelectron spectroscopy (XPS), and determined to be composed of carbon, nitrogen, and adventitious oxygen. Raman spectroscopic studies indicate that the addition of nitrogen induces further clustering and ordering of the sp(2)-hybridized carbon phase. The electrochemical activity of PLD fabricated N-DLC films was verified using the Ru(NH(3))(3+/2+) redox couple, and was determined to be comparable with that of other carbon-based electrodes. In situ spectroelectrochemical studies involving N-DLC coated zinc selenide (ZnSe) MIR waveguides provided evidence concerning the oxidation of N-DLC at anodic potentials in 1 M HClO(4) solutions. Finally, the electropolymerization of polyaniline (PAni) was performed at N-DLC-modified waveguide surfaces, which enabled spectroscopic monitoring of the electropolymerization, as well as in situ studying the structural conversion of PAni at different potentials.