Analysis of free analyte fractions by rapid affinity chromatography

The invention is generally directed toward an analytical method to determine the concentration of the free analyte fraction in a sample. More particularly, the method encompasses applying a sample comprising a free and bound analyte fraction to an affinity column capable of selectively extracting the free fraction in the millisecond time domain. The signal generated by the free fraction is then quantified by standard analytical detection techniques. The concentration of the free fraction may then be determined by comparison of its signal with that of a calibration curve depicting the signal of known concentration of the same analyte

Chromatography of beer

The objectives of the review are the collection, concise description and evaluation of the various chromatographic techniques used for the separation and quantitative determination of macro- and microcomponents present in beers

Occurrence of ochratoxin A and citrinin in Czech cereals and comparison of two HPLC methods for ochratoxin A detection.

The aims of the study were to obtain information about the occurrence of ochratoxin A (OTA) and citrinin (CIT) in cereals harvested in the Czech Republic and to compare two analytical procedures for detecting OTA. A total of 34 cereal samples, including two matrix reference materials (R-Biopharm, Germany), were analysed. The results were compared with the limit for raw cereal grains used as a foodstuff according to Commission Regulation No. 1881/2006, which allows a maximum OTA level of 5 µg kg−1. Compared were two methods based on the high-performance liquid chromatography principle, one using the immunoaffinity columns OchraTest™ (VICAM) and the second based on solvent partition (PART), both followed by fluorescence detection. The highest OTA contents were found in two barley samples. According to the method employed, the results for the first sample (malting barley) were VICAM = 31.43 µg kg−1 and PART = 44.74 µg kg−1. For the second sample (feeding barley) they were VICAM = 48.63 µg kg−1 and PART = 34.40 µg kg−1. Two samples of bread wheat had an OTA content approaching the legal limit (VICAM = 4.71 µg kg−1 and PART = 6.03 µg kg−1; VICAM = 4.12 µg kg−1 and PART = 3.95 µg kg−1). CIT was analysed using the PART method only, and its highest content (93.64 µg kg−1) was found for the malting barley sample with high OTA content (44.74 µg kg−1 as analysed using PART)

Multianalyte LC-MS-based methods in doping control: what are the implications for doping athletes?

Over the last 50 years, the list of doping substances and methods has been progressively expanding, being regularly reviewed by the international antidoping authorities (formerly the Medical Commission of the International Olympic Committee, and afterward, following its constitution in 1999, the World Anti-Doping Agency [WADA]). New substances/classes of substances have been periodically included in the list, keeping the pace with more advanced and sophisticated doping trends. At present, and apart from the prohibited performance enhancing and masking methods (e.g., blood transfusions and tampering strategies), the list comprises several hundreds of biologically active substances, with broad differences in their physicochemical properties (i.e., molecular weight, polarity and acid-basic properties) [1]. As a consequence, the ‘one class – one procedure’ approach, which had been followed by nearly all accredited antidoping laboratories worldwide until the turn of the millennium, is no longer sustainable. The need to minimize the overall number of independent analytical procedures, and, in parallel, to reduce the analytical costs, stimulated the development of multitargeted methods, aimed to increase the overall ratio of ‘target analytes: procedure’ [2–6]. The above evolution has not always been a straight forward process. The need to comply with the WADA technical requirements (both in terms of identification criteria and of minimum required performance limits [7,8]) and with the reduction of the reporting time (a constraint that becomes even more critical during international sport events, where the daily workload also drastically increases) has imposed a thorough re-planning of the analytical procedures. The development of an antidoping analytical method requires the appropriate knowledge not only of the biophysicochemical properties of the target analyte, but also of its PK profile. Historically, immunological methods and GC-based techniques were applied in antidoping science, as preferential screening methods for the detection of prohibited substances, which were originally limited to nonendogenous stimulants and narcotics. In the 1980s, GC–MS became the reference analytical platform for the detection and quantification of the majority of the low molecular weight doping substances [3–6]. In the following two decades, with the inclusion in the Prohibited List of new classes of low molecular weight, hydrophilic, thermolabile, nonvolatile analytes (including, but not limited to, glucocorticoids and designer steroids) and simultaneously of peptide hormones, scientists were obliged to design, develop, validate and apply techniques based on LC–MS/MS

Bidimensional Tandem Mass Spectrometry for Selective Identification of Nitration Sites in Proteins.

Nitration of protein tyrosine residues is very often regarded as a molecular signal of peroxynitrite formation during development, oxidative stress, and aging. However, protein nitration might also have biological functions comparable to protein phosphorylation, mainly in redox signaling and in signal transduction. The major challenge in the proteomic analysis of nitroproteins is the need to discriminate modified proteins, usually occurring at substoichiometric levels from the large amount of nonmodified proteins. Moreover, precise localization of the nitration site is often required to fully describe the biological process. Existing methodologies essentially rely on immunochemical techniques either using 2D-PAGE fractionation in combination with western blot analyses or exploiting immunoaffinity procedures to selectively capture nitrated proteins. Here we report a totally new approach involving dansyl chloride labeling of the nitration sites that rely on the enormous potential of MSn analysis. The tryptic digest from the entire protein mixture is directly analyzed by MS on a linear ion trap mass spectrometer. Discrimination between nitro- and unmodified peptide is based on two selectivity criteria obtained by combining a precursor ion scan and an MS3 analysis. This new procedure was successfully applied to the identification of 3-nitrotyrosine residues in complex protein mixtures

2014 Proficiency Test of the European Union Reference Laboratory for Mycotoxins for the Network of National Reference Laboratories - Determination of Zearalenone in Maize Oil

This report presents the results of the PT of the EURL for Mycotoxins which focused on the determination of zearalenone in maize oil. Forty-eight participants from thirty countries (among them 32 NRLs, 2 Non-EU Reference Laboratories and 13 official food control laboratories) registered for the exercise and 46 sets (Sample A and B) of results were reported. Only z-scores were used for the evaluation whether an individual laboratory underperformed. In total, 87 % of the attributed z scores were below an absolute value of two, which indicates that most of the participants performed satisfactorily.JRC.D.5-Standards for Food Bioscienc