Continuous chromatographic separation of glucose-fructose mixtures using anion-exchange resins

Summary A semi-continuous, counter-current chromatographic refiner (SCCR) was packed with an anion resin in the bisulphite form and used to separate mixtures of sugars from different feedstocks. Products with purities of more than 99% were achieved and solid concentrations of the fructose products of 12% w/v were possible. The deterioration of the anion resin was controlled by simple precautions and thus its useful life was substantially prolonged.     

Theoretical aspects of a preparative continuous chromatograph

Summary Equipment which has been used or suggested for preparative scale chromatography is reviewed. Particular attention is paid to one form of equipment for continuous counter-current chromatography which employs a moving mechanical seal. The theoretical effects of practical operating conditions and the mode of operation of this equipment are discussed. The influence of these effects on polymer fractionation is also considered. A theoretical model, based on the equilibrium stage concept, is proposed for the simulation of continuous gel permeation chromatography of a binary feed on such an apparatus. Examples of the results obtained from the computer simulation are given.     

Protein family classification and functional annotation

With the accelerated accumulation of genomic sequence data, there is a pressing need to develop computational methods and advanced bioinformatics infrastructure for reliable and large-scale protein annotation and biological knowledge discovery. The Protein Information Resource (PIR) provides an integrated public resource of protein informatics to support genomic and proteomic research. PIR produces the Protein Sequence Database of functionally annotated protein sequences. The annotation problems are addressed by a classification-driven and rule-based method with evidence attribution, coupled with an integrated knowledge base system being developed. The approach allows sensitive identification, consistent and rich annotation, and systematic detection of annotation errors, as well as distinction of experimentally verified and computationally predicted features. The knowledge base consists of two new databases, sequence analysis tools, and graphical interfaces. PIR-NREF, a non-redundant reference database, provides a timely and comprehensive collection of all protein sequences, totaling more than 1,000,000 entries. iProClass, an integrated database of protein family, function, and structure information, provides extensive value-added features for about 830,000 proteins with rich links to over 50 molecular databases. This paper describes our approach to protein functional annotation with case studies and examines common identification errors. It also illustrates that data integration in PIR supports exploration of protein relationships and may reveal protein functional associations beyond sequence homology.     

Use of homogeneous competition for OH radicals in electrochemical studies of H abstraction from organic molecules

The influence on the N₂O photocurrent of homogeneous competition for OH radicals between two organic solutes which form (as the result of hydrogen abstraction) radicals, one of which is reduced by the electrode, the other being oxidised, is considered theoretically. Such competition can be employed to investigate the kinetics of hydrogen abstraction in a case in which uncompetitive homogeneous destruction of OH radicals by a solute has no effect on the photocurrent. The influence of incomplete oxidation of alcohol radicals when a light pulse of short duration is employed is discussed, together with complications caused by adsorption of the organic solute. Competition for OH radicals between phenol and methanol points to a rate constant for H abstraction from phenol of ca. 2.8 × 10⁹ l mol^?1 s^?1 and to rapid heterogeneous reduction of most of the C₆H₄OH radicals throughout the accessible potential range.     

Detection of DNA adducts of benzo[a]pyrene using immunoelectrophoresis with laser-induced fluorescence. Analysis of A549 cells

Detection of benzo[a]pyrene diol epoxide (BPDE)-damaged DNA in a human lung carcinoma cell line (A549) has been performed using free zone affinity capillary electrophoresis with laser-induced fluorescence (LIF). Using BPDE as a model carcinogenic compound, the speed, sensitivity and specificity of this technique was demonstrated. Under free zone conditions, an antibody bound adduct was baseline-resolved from an unbound adduct in less than 2 min. The efficiencies of separation were in excess of 6 x 10(5) and 1 x 10(6) plates per meter for the antibody-bound and unbound adducts, respectively. Separation using a low ionic strength buffer permitted the use of a high electric field (830 V/cm) without the loss of resolving power. Using LIF detection, a concentration detection limit of roughly 3 x 10(-10) M was achieved for a 90-mer oligonuleotide containing a single BDPE. The use of formamide in the incubation buffer to enhance denaturing of DNA did not affect the stability of the complex between the antibody and the adducts. Using a fluorescently labeled BPDE-modified DNA adduct probe, a competitive assay was established to determine the levels of BPDE-DNA adducts in A549 cells.     

LC/MS/MS analysis of the endogenous dimethyltryptamine hallucinogens,their precursors,and major metabolites in rat pineal gland microdialysate

We report a qualitative liquid chromatography–tandem mass spectrometry (LC/MS/MS) method for the simultaneous analysis of the three known N,N‐dimethyltryptamine endogenous hallucinogens, their precursors and metabolites, as well as melatonin and its metabolic precursors. The method was characterized using artificial cerebrospinal fluid (aCSF) as the matrix and was subsequently applied to the analysis of rat brain pineal gland‐aCSF microdialysate. The method describes the simultaneous analysis of 23 chemically diverse compounds plus a deuterated internal standard by direct injection, requiring no dilution or extraction of the samples. The results demonstrate that this is a simple, sensitive, specific and direct approach to the qualitative analysis of these compounds in this matrix. The protocol also employs stringent MS confirmatory criteria for the detection and confirmation of the compounds examined, including exact mass measurements. The excellent limits of detection and broad scope make it a valuable research tool for examining the endogenous hallucinogen pathways in the central nervous system. We report here, for the first time, the presence of N,N‐dimethyltryptamine in pineal gland microdialysate obtained from the rat. Copyright © 2013 John Wiley & Sons, Ltd.