Uwe Karst

Simultaneous HPLC Determination of Peroxyacetic Acid and Hydrogen Peroxide

The first instrumental method for simultaneous determination of peroxyacetic acid (PAA) and hydrogen peroxide has been developed. The successive quantitative reaction of PAA with methyl p-tolyl sulfide (MTS) and hydrogen peroxide with triphenylphosphine (TPP) yields the corresponding sulfoxide MTSO and phosphine oxide TPPO. The reagents and their oxides are separated by HPLC on reversed-phase columns with acetonitrile/water gradient elution within 5 min. External calibration with the solid standards of MTSO and TPPO leads to a very accurate and reliable method. Samples are stable and can be stored after derivatization for several days prior to analysis. Real samples from brewery disinfection were analyzed in comparison to titration with excellent correlation.

Recent developments in optical detection methods for microchip separations

This paper summarizes the features and performances of optical detection systems currently applied in order to monitor separations on microchip devices. Fluorescence detection, which delivers very high sensitivity and selectivity, is still the most widely applied method of detection. Instruments utilizing laser-induced fluorescence (LIF) and lamp-based fluorescence along with recent applications of light-emitting diodes (LED) as excitation sources are also covered in this paper. Since chemiluminescence detection can be achieved using extremely simple devices which no longer require light sources and optical components for focusing and collimation, interesting approaches based on this technique are presented, too. Although UV/vis absorbance is a detection method that is commonly used in standard desktop electrophoresis and liquid chromatography instruments, it has not yet reached the same level of popularity for microchip applications. Current applications of UV/vis absorbance detection to microchip separations and innovative approaches that increase sensitivity are described. This article, which contains 85 references, focuses on developments and applications published within the last three years, points out exciting new approaches, and provides future perspectives on this field.

The Chloroplast Calcium Sensor CAS Is Required for Photoacclimation in Chlamydomonas reinhardtii

Reverse genetics approaches were conducted to elucidate the function of the plant-specific calcium binding protein CAS (calcium sensor) that is localized in chloroplast thylakoid membranes. Our findings demonstrate that CAS and Ca2+ are critically involved in the regulation of the high-light response and particularly in the regulation of LHCSR3 expression in green alga Chlamydomonas reinhardtii. The plant-specific calcium binding protein CAS (calcium sensor) has been localized in chloroplast thylakoid membranes of vascular plants and green algae. To elucidate the function of CAS in Chlamydomonas reinhardtii, we generated and analyzed eight independent CAS knockdown C. reinhardtii lines (cas-kd). Upon transfer to high-light (HL) growth conditions, cas-kd lines were unable to properly induce the expression of LHCSR3 protein that is crucial for nonphotochemical quenching. Prolonged exposure to HL revealed a severe light sensitivity of cas-kd lines and caused diminished activity and recovery of photosystem II (PSII). Remarkably, the induction of LHCSR3, the growth of cas-kd lines under HL, and the performance of PSII were fully rescued by increasing the calcium concentration in the growth media. Moreover, perturbing cellular Ca2+ homeostasis by application of the calmodulin antagonist W7 or the G-protein activator mastoparan impaired the induction of LHCSR3 expression in a concentration-dependent manner. Our findings demonstrate that CAS and Ca2+ are critically involved in the regulation of the HL response and particularly in the control of LHCSR3 expression.

Strategies for the liquid chromatographic-mass spectrometric analysis of non-polar compounds

Electrospray ionization and atmospheric pressure chemical ionization (APCI) have evolved recently as very useful tools for the liquid chromatographic-mass spectrometric (LC-MS) analysis of polar substances. Non-polar compounds, however, are difficult to analyze with these atmospheric pressure ionization techniques due to their soft ionization mechanism. Recently, new approaches have been introduced which are likely to overcome this obstacle, at least partly. On-line electrochemical conversion of the analytes to more polar reaction products, atmospheric pressure photoionization, atmospheric pressure electron capture negativeion-MS and coordination ionspray-MS are four techniques which are presented in detail compared and discussed critically with respect to their current status and future perspectives. Particular focus is directed from a chemical viewpoint on the substance groups which are accessible by each of the new approaches.

Ion chromatographic determination of hydrolysis products of hexafluorophosphate salts in aqueous solution.

In this work, hydrolysis of three different hexafluorophosphate salts in purified water was investigated. Aqueous samples of lithium hexafluorophosphate (LiPF(6)), sodium hexafluorophosphate (NaPF(6)) and potassium hexafluorophosphate (KPF(6)) were prepared and stored for different times. Ion chromatography (IC) with UV as well as non-suppressed and suppressed conductivity detection was used for the analysis of the reaction products. For the detection and identification of the formed decomposition products, an IC method using IonPac AS14A 250 mm × 4.0 mm i.d. column and 2.5 mM KHCO(3)-2.5 mM K(2)CO(3) eluent was established. Besides hexafluorophosphate, four other anionic species were detected in fresh and matured aqueous solutions. The hydrolysis products fluoride (F(-)), monofluorophosphate (HPO(3)F(-)), phosphate (HPO(4)(2-)) and difluorophosphate (PO(2)F(2)(-)) were found and were unambiguously identified by means of standards or electrospray ionization mass spectrometry (ESI-MS). It was shown that stability of hexafluorophosphate solutions depends on the nature of the counter ion and decreases in the order potassium>sodium>lithium.

Online electrochemistry/mass spectrometry in drug metabolism studies: principles and applications.

Importance of the field: Conventional metabolism studies in early stages of drug development include in vitro tests, often based on hepatic cells and cell extracts, as well as in vivo tests on the basis of animal models. Complementary to these existing techniques, the electrochemical study of oxidative metabolism reactions is gaining increasing attention. Electrochemistry (EC) allows the fast detection of sites labile towards oxidation in a drug molecule and can mimic the formation of potential oxidative metabolites. Areas covered in this review: The present review summarizes the developments of EC-based methods to study the oxidative metabolism of drugs in the past 10 years. The implementation of different coulometric and amperometric cells is described. Furthermore, online set-ups utilizing the hyphenation of EC, liquid chromatography and electrospray ionization mass spectrometry are discussed regarding their applicability in metabolism studies. Besides mass spectrometric detection, the isolation and advanced characterization of oxidation products are reviewed. This includes structure elucidation based on NMR spectroscopy as well as the evaluation of the reactivity of metabolites towards trapping agents or proteins. What the reader will gain: A major focus of the article is directed to the comparability between electrochemically predicted metabolites and those occurring in vitro and in vivo. This comprises the discussion of typical oxidative metabolism reactions and provides a guideline about how far EC is capable of generating a specific oxidative metabolite. Take home message: The study of drug metabolism reactions in an electrochemical cell enables the prediction as well as the synthesis of oxidative metabolites. In particular, reactive metabolites are directly detected using online EC/electrospray ionization mass spectrometry. Thus, the electrochemical technique is a promising tool that complements existing in vivo and in vitro techniques in drug metabolism studies.

Electrochemistry coupled to (liquid chromatography/) mass spectrometry—Current state and future perspectives

The coupling of electrochemistry (EC) to different mass spectrometric (MS) techniques in off-line and especially in on-line approaches is a quickly growing research field in analytical chemistry with numerous distinct objectives. Depending on the analytical problem, a separation step can be further integrated according to the instrumental set-up and, most frequently, liquid chromatography (LC) is selected for this purpose. In this review, various scientific areas of application for this EC/(LC/)MS hybrid method are presented and discussed in detail. Therefore, one major division is made between those applications which are already successfully used on a large scale (current state), and those which have shown promising results for future utilization (future perspectives). The reader shall be provided with a thorough overview on the capabilities of the combination of EC/(LC/)MS and the drawbacks which result in further optimization and exploration of this technique. The major topics addressed in this review include the role of EC/(LC/)MS for drug metabolism studies, peptide, protein and DNA (deoxyribonucleic acid) research and quantification strategies. Promising future applications that are presented and evaluated comprise the fields of toxicology and forensics, targeted product synthesis and environmental analysis.

Diagnosis of Nephrogenic Systemic Fibrosis by means of Elemental Bioimaging and Speciation Analysis

The combined use of elemental bioimaging and speciation analysis is presented as a novel means for the diagnosis of nephrogenic systemic fibrosis (NSF), a rare disease occurring after administration of gadolinium-based contrast agents (GBCA) for magnetic resonance imaging (MRI), in skin samples of patients suffering from renal insufficiency. As the pathogenesis of NSF is still largely unknown particularly with regard to the distribution and potential retention of gadolinium in the human organism, a skin biopsy sample from a suspected NSF patient was investigated. The combination of inductively coupled plasma mass spectrometry (ICP-MS), laser ablation (LA) ICP-MS for quantitative elemental bioimaging, and hydrophilic interaction liquid chromatography (HILIC) ICP-MS for speciation analysis allowed one to unambiguously diagnose the patient as a case of NSF. By means of ICP-MS, a total gadolinium concentration from 3.02 to 4.58 mg/kg was determined in the biopsy sample, indicating a considerable deposition of gadolinium in the patients skin. LA-ICP-MS revealed a distinctly inhomogeneous distribution of gadolinium as well as concentrations of up to 400 mg/kg in individual sections of the skin biopsy. Furthermore, the correlation between the distributions of phosphorus and gadolinium suggests the presence of GdPO4 deposits in the tissue section. Speciation analysis by means of HILIC-ICP-MS showed the presence of the intact GBCA Gd-HP-DO3A eight years after the administration to the patient. The concentration of the contrast agent in the aqueous extract of the skin biopsy was found to be 1.76 nmol/L. Moreover, evidence for the presence of further highly polar gadolinium species in low concentrations was found.

Determination of gadolinium-based MRI contrast agents in biological and environmental samples: A review

The development of analytical methods and strategies to determine gadolinium and its complexes in biological and environmental matrices is evaluated in this review. Gadolinium (Gd) chelates are employed as contrast agents for magnetic resonance imaging (MRI) since the 1980s. In general they were considered as safe and well-tolerated, when in 2006, the disease nephrogenic systemic fibrosis (NSF) was connected to the administration of MRI contrast agents based on Gd. Pathogenesis and etiology of NSF are yet unclear and called for the development of several analytical methods to obtain elucidation in this field. Determination of Gd complex stability in vitro and in vivo, as well as the quantification of Gd in body fluids like blood and urine was carried out. Separation of the Gd chelates was achieved with high performance liquid chromatography (HPLC) and capillary electrophoresis (CE). For detection, various methods were employed, including UV-vis absorbance and fluorescence spectroscopy, electrospray ionization mass spectrometry (ESI-MS) and inductively coupled plasma mass spectrometry (ICP-MS). A second challenge for analysts was the discovery of high concentrations of anthropogenic Gd in surface waters draining populated areas. The source could soon be determined to be the increasing administration of Gd complexes during MRI examinations. Identification and quantification of the contrast agents was carried out in various surface and groundwater samples to determine the behavior and fate of the Gd chelates in the environment. The improvement of limits of detection (LOD) and limits of quantification (LOQ) was and still is the goal of past and ongoing projects.

Identification of chemical interferences in aldehyde and ketone determination using dual-wavelength detection.

A method for the rapid and convenient identification of chemical interferences in the determination of aldehydes and ketones in air samples using the 2,4-dinitrophenylhydrazine (DNPH) method is described. The ratio of absorptions at 360 and 300 nm is characteristic for groups of related aldehydes and ketones as well as for the main interferents. It has been determined by UV/visible spectroscopy of pure standard compounds and confirmed by HPLC analysis with UV/visible detection of complex hydrazone mixtures. The application of this method has led to the identification of 2,4-dinitrochlorobenzene as another interfering compound after sampling of air with high nitrogen dioxide content when using hydrochloric acid as catalyst. The possibilities and limitations of the dual-wavelength detection with the DNPH method are discussed for mixtures of standards and real samples from car exhaust.