Biomedical subjects

B Lendl

Publications and source records attributed to B Lendl.

14 recordsLinked to original sources

PubMed2006

Alternatives for coupling sequential injection systems to commercial capillary electrophoresis-mass spectrometry equipment.

On-line coupling of an automated flow system with a commercially available capillary electrophoresis (CE) system with an electrospray interface (ESI) for mass spectroscopic (MS) detection is described. The peculiarities of CE-ESI-MS interfaces, in which a high electrical field must be applied to the capillary end where the sample is provided by the flow system, introduce significant difficulties for the appropriate work of the entire arrangement. Experimental strategies are proposed for achieving stable conditions for on-line sample pre-treatment, conditioning of the separation capillary, sample injection, as the proper separation. The versatility and robustness of the proposed arrangement is discussed, taken as example the separation of a variety of amines. Connection of the CE system's pressure to the automated flow system enables hydrodynamic introduction of sample with high precision. The developed hyphenated system is of practical relevance as it opens an avenue for the simplification and automation of the whole analytical process required when using powerful CE-ESI-MS equipments.

Butylamines↗

PubMed2006

Towards biochemical reaction monitoring using FT-IR synchrotron radiation.

A lab-on-a-chip device made of CaF2 windows and SU-8 polymer was used for fluid lamination to achieve rapid mixing of two streamlines with a cross section of 300 x 5 microm each. Time resolved measurements of the induced chemical reaction was achieved by applying constant feeding low flow rates and by on-chip measurement at defined distances after the mixing point. Synchrotron IR microscopic detection was employed for direct and label-free monitoring of (bio)chemical reactions. Furthermore, using synchrotron IR microscopy the measurement spot could be reduced to the diffraction limit, thus maximizing time resolution in the experimental set-up under study. Based on computational fluid dynamic simulations the principle of the set-up is discussed. Experimental results on the basic hydrolysis of methyl chloroacetate proved the working principle of the experimental set-up. First results on the interaction between the antibiotic vancomycin and a tripeptide (Ac2KAA) involved in the build up of the membrane proteins of gram-positive bacteria are presented.

Animals↗

PubMed2006

Time-resolved Fourier transform infrared spectroscopy of chemical reactions in solution using a focal plane array detector.

A Fourier transform infrared (FT-IR) microscope equipped with a single as well as a 64 x 64 element focal plane array MCT detector was used to measure chemical reaction taking place in a microstructured flow cell designed for time-resolved FT-IR spectroscopy. The flow cell allows transmission measurements through aqueous solutions and incorporates a microstructured mixing unit. This unit achieves lamination of the two input streams with a cross-section of 300 x 5 microm each, resulting in fast diffusion-controlled mixing of the two input streams. Microscopic measurement at defined positions along the outlet channel allows time-resolved information of the reaction taking place in the flow cell to be obtained. In this paper we show experimental results on the model reaction between formaldehyde and sulfite. Using the single-point MCT detector, high-quality FT-IR spectra could be obtained from a spot size of 80 x 200 microm whereas the FPA detector allowed recording light from an area of 260 x 260 microm focused on its 64 x 64 detector elements. Therefore, more closely spaced features could be discerned at the expense of a significantly lower signal-to-noise (S/N) ratio per spectrum. Multivariate curve resolution-alternating least squares was used to extract concentration profiles of the reacting species along the outlet channel axis.

Journal Article↗

PubMed2005

Terahertz pulsed spectroscopy as a new tool for measuring the structuring effect of solutes on water.

Absorption spectra of aqueous solution of ''chaotropes'' (structure maker) and ''kosmotropes'' (structure breaker) have been recorded in the mid-infrared (MIR) and terahertz (THz) spectral region. A different impact of the two groups of solutes on the absorption spectrum of water was found in the recorded THz spectra. A concentration-dependent increased absorption across the investigated THz spectral region (0.04-2 THz, 1.3-66 cm(-1), respectively) has been recorded for all studied chaotropic solutions, whereas the opposite has been obtained for kosmotrope containing solutions. In the case of ionic solutes a further increase in absorption towards higher frequencies was measured. The distinction between chaotrope and kosmotrope solutes was, as expected, also possible in the MIR spectral region. Depending on the structure-forming effect of the solute the OH stretch vibration of the water (around 3400 cm(-1)) was slightly shifted. A red shift has been observed for solution of kosmotropes, whereas a blue shift was observed in the case of solutions containing chaotropes. Compared to the MIR spectral region the structure influencing effect of solutes can be more efficiently studied in the THz spectral region, which provides information from interactions between neighboring water molecules.

Complex Mixtures↗

PubMed2004

Mid-IR synchrotron radiation for molecular specific detection in microchip-based analysis systems.

Microstructures constructed from SU-8 polymer and produced on CaF(2) base plates have been developed for microchip-based analysis systems used to perform FTIR spectroscopic detection using mid-IR synchrotron radiation. The high brilliance of the synchrotron source enables measurements at spot sizes at the diffraction limit of mid-IR radiation. This corresponds to a spatial resolution of a few micrometers (5-20 microm). These small measurement spots are useful for lab-on-a-chip devices, since their sizes are comparable to those of the structures usually used in these devices. Two different types of microchips are introduced here. The first chip was designed for time-resolved FTIR investigations of chemical reactions in solution. The second chip was designed for chip-based electrophoresis with IR detection on-chip. The results obtained prove the operational functionality of these chips, and indicate the potential of these new devices for further applications in (bio)analytical chemistry.

Journal Article↗

PubMed2004

Direct determination of carbon dioxide in aqueous solution using mid-infrared quantum cascade lasers.

A method for the direct determination of carbon dioxide in aqueous solutions using a room-temperature mid-infrared (MIR) quantum cascade laser at 2330 cm(-1) is reported. The absorption values of different carbon dioxide concentrations were measured in a 119 microm CaF2 flow-through cell. An optical system made of parabolic mirrors was used to probe the flow cell and to focus the laser beam on the mercury cadmium telluride (MCT) detector. Aqueous carbon dioxide standards were prepared by feeding different mixtures of gaseous N2 and CO2 through wash bottles at controlled temperature. The concentration of the dissolved CO2 was calculated according to Henry's law, taking into account the temperature and the partial pressure of CO2. The carbon dioxide standards were connected via a selection valve to a peristaltic pump for subsequent, automated measurement in the flow-through cell. A calibration curve was obtained in the range of 0.338 to 1.350 g/L CO2 with a standard deviation of the method sxo equal to 19.4 mg/L CO2. The limit of detection was calculated as three times the baseline noise over time and was determined to be 39 mg/L.

Beverages↗

PubMed2002

A mid-IR flow-through sensor for direct monitoring of enzyme catalysed reactions. Case study: Measurement of carbohydrates in beer.

A novel mid-IR flow-through sensor for in situ monitoring of the enzymatic reaction of amyloglucosidase with carbohydrates was developed. Amyloglucosidase was immobilised on agarose beads with N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) and directly placed in a conventional IR flow-through cell. The carbohydrate content of various beer samples was then determined by following the enzymatic hydrolytic cleavage of carbohydrates to glucose with Fourier-transform infrared (FTIR) spectroscopy. The whole procedure was done in an automated way operating in the stopped flow mode by incorporating the flow-through sensor in a sequential injection (SI) manifold. As the immobilised enzyme was directly probed by the IR beam, an in situ study of the enzymatic reaction was possible enabling determination of the Michaelis-Menten constant of the immobilised enzyme. A linear calibration curve was recorded using maltose standards in the range between 0.86 and 7.13 g L(-1). The proposed method was successfully applied to the determination of the carbohydrate content of four different beer samples by the standard addition method. Moreover experiments showed the possibility of monitoring in situ the immobilisation of the enzyme as well as a small organic acid (malic acid) on the agarose beads using EDC.

Journal Article↗

PubMed2001

Design, simulation and application of a new micromixing device for time resolved infrared spectroscopy of chemical reactions in solution.

We present a novel micromachined fast diffusion based mixing unit for the study of rapid chemical reactions in solution with stopped-flow time resolved Fourier transform infrared spectroscopy (TR-FTIR). The presented approach is based on a chip for achieving lamination of two liquid sheets of 10 microm thickness and approximately 1 mm width on top of each other and operation in the stopped-flow mode. The microstructure is made on infrared transmitting calcium fluoride discs and built up with two epoxy negative photoresist layers and one silver layer in between. Due to the highly laminar flow conditions and the short residence time in the mixer there is hardly any mixing when the two liquid streamlines pass through the mixing unit, which allows one to record a mid-IR transmission spectrum of the analytes prior to reaction. When the flow is stopped, the reactant streams are arrested in the flow-cell and rapidly mixed by diffusion due to the reduced interstream distances and the reaction can be directly followed with hardly any dead time. On the basis of two model reactions-neutralisation of acetic acid with sodium hydroxide as well as saponification of methyl monochloroacetate-the performance of the mixing device was tested revealing proper functioning of the device with a time for complete mixing of less than 100 ms. The experimental results were supported by numerical simulations using computational fluid dynamics (CFD), which allowed a reliable, quantitative analysis of concentration, pressure and flow profiles in the course of the mixing process.

Journal Article↗

PubMed2001

Rapid method for the discrimination of red wine cultivars based on mid-infrared spectroscopy of phenolic wine extracts.

Mid-infrared spectroscopy and UV-vis spectroscopy combined with multivariate data analysis have been applied for the discrimination of Austrian red wines, including the cultivars Cabernet Sauvignon, Merlot, Pinot Noir, Blaufränkisch (Lemberger), St. Laurent, and Zweigelt. Both authentic wines and their phenolic extracts were investigated by attenuated total reflectance (ATR)-mid-infrared spectroscopy. Phenolic extracts were also investigated by UV-vis spectroscopy. The wine extracts were obtained by solid-phase extraction with C-18 columns and elution by methanol containing 0.01% hydrochloric acid. Hierarchical cluster analysis was performed with mid-infrared spectra of both wines and extracts, as well as with UV-vis spectra of the phenolic extracts. Data processing involved vector normalization and derivation of the spectra. Due to varying concentrations of main components including sugar and organic acids, satisfactory classification of untreated wines was not achieved. However, when using mid-infrared spectra of the phenolic extracts, almost complete discrimination of all cultivars investigated was achieved. The use of UV-vis spectroscopy for cultivar discrimination was found to be limited to the authentication of the Burgundy species Pinot Noir. In addition, soft independent modeling of class analogy was applied to the mid-infrared spectra of the extracts. It was possible to establish class models for five different wine cultivars and to classify test samples correctly.

Anthocyanins↗

PubMed2001

A rapid method for peroxide value determination in edible oils based on flow analysis with Fourier transform infrared spectroscopic detection.

The development of an automated, rapid and highly precise method for determination of the peroxide value in edible oils based on a continuous flow system and Fourier transform infrared (FTIR) spectroscopic detection is described. The sample stream was mixed with a solvent mixture consisting of 25% (v/v) toluene in hexanol which contained triphenylphosphine (TPP). The hydroperoxides present in the sample reacted stoichiometrically with TPP to give triphenylphosphine oxide (TPPO) which has a characteristic and intense absorption band at 542 cm-1. A 10% (m/v) TPP solution in the solvent mixture and a 100 cm reaction coil were necessary for complete reaction. FTIR transmission spectra were recorded using a flow cell equipped with CsI windows having an optical pathlength of 100 microns. By using tert-butyl hydroperoxide spiked oil standards and evaluation of the band formed at 542 cm-1 a linear calibration graph covering the range 1-100 PV (peroxide value; mequiv O2 kg-1 oil) was obtained. The relative standard deviation was 0.23% (n = 11) and the throughput 24 samples h-1. The developed system was also applied to the determination of PV in olive, sunflower and corn oils, showing good agreement with the official reference method of the European Community which is based on titration using organic solvents. The results obtained clearly show that the developed method is superior to the standard wet chemical method, hence suggesting its application in routine analysis and quality control.

Flow Injection Analysis↗

PubMed2000

Multidimensional information on the chemical composition of single bacterial cells by confocal Raman microspectroscopy.

In many biotechnological processes, living microorganisms are used as biocatalysts. Biochemical engineering science is becoming more aware that individual cells of an organism in a process can be fairly inhomogeneous regarding their properties and physiological status. Raman microspectroscopy is a novel approach to characterize such differentiated populations. Cells of the anaerobic bacterium Clostridium beijerinckii were dried on transparent support surfaces. The laser beam of a confocal Raman microscope was focused on individual cells viewed through the objective. Single bacterial cells in size approximately 1 microm and sample mass approximately 1 pg could be analyzed within a few minutes, when placed on a calcium fluoride support and using excitation at 632.8 nm. Spectral features could be attributed to all major cell components. Cells from a morphologically differentiated culture sample showed different compositions, indicating the presence of subpopulations. As a reference, the storage polymer granulose was detected. The multidimensional information in Raman spectra gives a global view on all major components of the cell at once, complementing other more specific information-rich methods for single-cell analysis. The method can be used, for example, to study heterogeneities in a microbial population.

Clostridium↗

PubMed2000

Mid-infrared quantum cascade lasers for flow injection analysis

A Fabry-Perot quantum cascade laser (QCL) was used as a powerful light source for mid infrared (MIR) detection in flow injection analysis. The QCL lased at several wavelengths close to each other within a few wavenumbers (990-1010 cm-1), hence fitting well to the broad absorption bands of molecules in liquid phase. As compared with that of a state-of-the-art Fourier transform spectrometer, the signal-to-noise ratio could be improved by a factor of 50. Additionally, by using a QCL as the light source, optical path lengths of more than 100 microns could be used even in aqueous matrixes which reduced the danger of cell clogging. In the example shown here phosphate was determined in Diet Coke samples. The flow injection system used allowed the measurement of the sample at two pH values (5 and 13) at which the analyte was present as H2PO4- and PO4(3-), respectively. As the analytical readout the difference in IR absorption of H2PO4- and PO4(3-) at the laser wavelengths was taken. The FIA-QCL measurements were corroborated by ion chromatography which was used as a reference method.

Journal Article↗

PubMed1997

Hyphenation of ion exchange high-performance liquid chromatography with Fourier transform infrared detection for the determination of sugars in nonalcoholic beverages.

Fourier transform infrared spectroscopy (FT-IR) is presented here as a molecular-specific detection system for high-performance liquid chromatography (HPLC) in an aqueous phase, focusing on the chromatographic separation of sugars in beverages. The separation was achieved with an isocratic HPLC setup using an ion exchange column (counterion, Ca2+). The FT-IR detection of the C-O bands in the mid-IR between 1000 and 1200 cm-1 was performed in real time with a 25 microns flow cell without elimination of the solvent. Characteristic FT-IR spectra of the common sugars sucrose, glucose, and fructose in concentrations of 1 mg/mL could be recorded during the separation. The calibration of these compounds in the 5-100 mg/mL range resulted in a linear correlation with a standard deviation of the method (Sx0) of 0.11, 0.07, and 0.11 mg/mL for sucrose, glucose, and fructose, respectively. The method was, furthermore, applied to the analysis of nine soft drinks and fruit juices containing between 6 and 97 mg/mL of each carbohydrate. The accuracy of the method was confirmed by standard ion exchange HPLC with refractive index detection. The average deviation from the reference method was in the range of 0.5-0.9 mg/mL. Furthermore, the method was found to be suitable to identify and quantify also minor components in beverages, such as taurine (4 mg/mL) and ethanol (0.4 mg/mL).

Beverages↗

PubMed1996

Determination of alpha-amylase activity using Fourier transform infrared spectroscopy.

A new method for the determination of alpha-amylase activity in aqueous solutions and human serum with FTIR-spectroscopy is proposed. The chemical reaction catalyzed by the enzyme under study can be followed directly when applying FTIR-spectroscopic detection also in the case, where no colored or electrochemical active species are generated or consumed during the course of the reaction of alpha-amylase with simple starch. Therefore the determination of the alpha-amylase activity could successfully be performed by recording two FTIR-spectra, one immediately after mixing the sample and a substrate (starch-) solution and the other after a 20 min reaction time. From these two FTIR-spectra a difference spectrum was calculated hereby eliminating an unspecific absorption of the matrix. The intensities of the resulting difference spectra corresponded to the extent of the reaction which took place during the investigated time interval and hence could be related to the activity of the enzyme in the sample. The developed method is linear from 80 to 1400 U/l (r.s.d.=5% for 700 U/l) in aqueous solutions and was also successfully applied to the determination of alpha-amylase activity in human serum where a linear working range from 100 to 800 U/l (r.s.d.=11% for 150 U/l) was achieved.

Journal Article↗