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Porphyrias are a group of diseases that are clinically and genetically heterogeneous and originate mostly from inherited dysfunctions of specific enzymes involved in heme biosynthesis. Such dysfunctions result in the excessive production and excretion of the intermediates of the heme biosynthesis pathway in the blood, urine, or feces, and these intermediates are responsible for specific clinical presentations. Porphyrias continue to be underdiagnosed, although laboratory diagnosis based on the measurement of metabolites could be utilized to support clinical suspicion in all symptomatic patients. Moreover, the measurement of enzymatic activities along with a molecular analysis may confirm the diagnosis and are, therefore, crucial for identifying pre-symptomatic carriers. The present review provides an overview of the laboratory assays used most commonly for establishing the diagnosis of porphyria. This would assist the clinicians in prescribing appropriate diagnostic testing and interpreting the testing results.

The high pressure liquid chromatography method for determination of glutathione in free and protein-bound forms was re-established and has successfully been developed to measure glutathione related thiol compounds, i.e. l -cysteine, γ- l -glutamyl- l -cysteine and l -cysteinyl- l -glycine, in both free and protein-bound forms. The natural levels of those compounds in typical strong, weak flours, and flours from 36 wheat varieties grown in the UK were investigated. The total free and protein-bound glutathione compounds found in the 36 UK varieties was 358±51 and 190±17 nmol/g, respectively. Multiple correlation analysis did not show a clear-cut relationship between the natural level of glutathione and any related thiol compound in either free or protein-bound forms and flour quality attributes, including rheological properties, baking performance, protein content and SDS sedimentation test values. Therefore, it can be suggested that glutathione and related thiol compounds at natural levels do not lead to significant differences in the rheological properties of dough and the baking performance of flour.

BRIEF COMMUNICATION Pharmacokinetics of Low Molecular Weight Dermatan Sulphate (Desmin) in Different Cohorts of Patients Sylvie Saivin, Didier Carrie, Jean Escourrou, Patrick Duchene, Villiam Zamboni5, Miriam Barbanti5, Ernesto Palazzini5 and Georges Houin1 1Laboratoire de Pharmacocinetique et Toxicologie Clinique, Hopital Rangueil, Toulouse, France; Service de Cardiologie, Hopital Purpan, Toulouse, France; Service de Gastro-Enterologie et Nutrition, Hopital Rangueil, Toulouse, France; 4ADME Bioanalyses, Mougins, France; 5Alfa Wassermann, Bologna, Italy.

The interactions of DNA duplexes with 3-amino 1,4-dimethyl-5H-pyrido[4,3-b]indole(Trp-P-1), a potent mutacarcinogen isolated from tryptophan pyrolysate, have been studied using CD spectroscopy. The results are that (a) the spectral change of B-form DNA caused by the interaction with Trp-P-1 is biphasic, i.e. the enlargement of CD bands characteristic to the B-DNA conformation in the range of r ([Trp-P-1]/[DNA]) = 0–2.5, followed by the rapid transition to the non-B conformation at r > 2.5; (b) this transition degree of B to non-B conformation of DNA is not necessarily dependent on the G-C content; and (c) the salt-induced Z-DNA is transformed to B-DNA (0 < r < 0.1) and then to non-B-DNA (r > 5), depending on the concentration of Trp-P-1 added. These data indicate that the non-covalent interaction of Trp-P-1 with DNA is mainly dependent on the B-DNA conformation.

Separate pathways exist for the processing of antigens to be presented by MHC class I and class II molecules. We are beginning to determine the subcellular location of certain events in both pathways.

Quantification of artemisinin purity and amount in plant material and extracts to date has been characterized by a considerable inconsistency in values. This is likely to be due to the adoption of varied analytical procedures and use of inappropriate to the specific applications analytical techniques. In this paper we are attempting to further develop artemisinin analysis to the point where a universally acceptable reference method is available to the research and end-users communities. Thus, we have developed and validated an HPLC-RI method and optimized an HPLC-ELSD method. We used the gradient HPLC-UV method recommended by the current artemisinin monograph as a comparison for the method improvements presented herein, and show the limitations for its application scope. The data reported should help to allow more reliable laboratory analysis of artemisinin in both pure samples and in Artemisia annua extracts.

BPH Benign prostatic hyperplasis DCIS Ductal carcinoma in situ FNAB Fine needle aspiration biopsy GBM Glioblastoma multiforme HCA Hierarchical cluster analysis 1H MRS Proton magnetic resonance spectroscopy HPLC High pressure liquid chromatography (HR)MAS (High resolution) Magic angle spinning NAA N-acetyl aspartate PA Polyamines PC Phosphocholine PCA Principal component analysis PTC Phosphatidylcholine SCS Statistical classification strategy SSB Spinning side bands

Cushing syndrome is an insidious illness that warrants an early diagnosis to avoid the effects of prolonged hyper-cortisolism. The variability in the clinical features of the disease and the occasional inconsistencies between different biochemical tests performed to identify it render the diagnosis challenging. In this paper the author discusses the various biochemical tests that are useful for the diagnoses of Cushing syndrome and Cushing disease, with an emphasis on the respective sensitivities and specificities of these tests. The measurement of evening salivary cortisol and the combined low-dose dexamethasone–corticotropin-releasing hormone stimulation test have improved overall sensitivity and specificity in the evaluation of Cushing syndrome and Cushing disease.

International audience; To examine the substrate recognition mechanism of the human MTH1 protein, which hydrolyzes 2-hydroxy-dATP, 8-hydroxy-dATP, and 8-hydroxy-dGTP, ten nucleotide analogs (8-bromo-dATP, 8-bromo-dGTP, deoxyisoinosine triphosphate, 8-hydroxy-dITP, 2-aminopurine-deoxyriboside triphosphate, 2-amino-dATP, deoxyxanthosine triphosphate, deoxyoxanosine triphosphate, dITP, and dUTP) were incubated with the MTH1 protein. Of these, the former five nucleotides were hydrolyzed with various efficiencies. The fact that the syn-oriented brominated nucleotides were hydrolyzed suggests that the MTH1 protein binds to deoxynucleotides adopting the syn-conformation. However, 8-hydroxy-dITP, which lacks the 2-amino group of 8-hydroxy-dGTP, was degraded with tenfold less efficiency as compared with 8-hydroxy-dGTP. In addition, deoxyisoinosine triphosphate, lacking the 6-amino group of 2-hydroxy-dATP, was hydrolyzed as efficiently as 8-hydroxy-dGTP, but less efficiently than 2-hydroxy-dATP. These results clarify the effects of the anti/syn conformation and the functional groups on the 2 and 6 positions of the purine ring on the recognition by the human MTH1 protein.

Development of drugs for the effective treatment of depressive disorders requires elucidation of factors that are critical for clinically antidepressant effects. During the past 4 years, we have studied in situ neurochemical alterations in the brain that may underlie depressive behavior. This was achieved using the genetically-selected Flinders Sensitive Line (FSL) of rats (a unique animal model of depression), before and after chronic antidepressant treatment. This line of rats exhibits behavioral features characteristic of depression, and responds to chronic, but not acute, antidepressant treatments. This review summarizes our findings concerning the local neuro-dynamics in the brain during manifestation of depressive behavior and effective antidepressant treatment in this animal model of depression. Understanding the abnormalities manifested in neurochemical pathways during depressive disorders and the dynamic effects of these abnormalities on the onset of action and efficacy of pharmacological treatments are crucial for the development of effective antidepresssant drugs and therapeutic strategies.

The in vivo incorporation of [3H]phenylalanine into an apparent 15 kDa prosomatostatin was observed in the hypothalamus of rats injected with the labeled amino acid in the third ventricle. Precursor-product relationships were established between this newly synthesized material and both somatostatin-28 and -14.

The application of high-pressure liquid chromatography (HPLC) to proteins has undergone a dramatic development in recent years. Nowadays its many variants expand the repertoire of high-performance analysis methods available to the protein chemist, which, until now, have been dominated by electrophoretic techniques. The advent of gene technology has resulted in a renaissance of protein chemistry. The new analytical and preparative problems that have thereby emerged are often ideally solved by HPLC methods. HPLC has long since ceased to be solely a laboratory technique; HPLC systems are now being developed for the separation of proteins–particularly those of great pharmaceutical interest – on a 100-g scale. The range of applications of analytical and preparative HPLC will be illustrated by two examples of pharmaceutical importance—insulin and interleukin 2.

Nonbacterial systems (e.g., mammalian cells) for expressing therapeutic proteins are generally selected because correct folding or post-translational modifications cannot be achieved in microorganisms such as Escherichia coli . In producing recombinant proteins for therapeutic use a number of ‘key’ issues must be considered during their purification and characterization. These issues include: N- and C-terminal heterogeneity; chemical modification and conformational changes; glycosylation status; purification techniques; proteolytic processing; detection and elimination of contaminants such as cellular DNA, viruses, or endotoxins; potency; and safety. Various scientific achievements and challenges in obtaining recombinant therapeutic proteins are also elaborated in this review.

The three RNA trinucleotides; ApApA, ApApG, and ApUpG, have been synthesized in sufficient quantity to obtain natural abundance 13 C{ 1 H}-NMR spectra at strand concentrations between 4 and 100 mM. Comparisons between 70°C spectra of the three trimers and their consistuent dimers ApA, ApG, ApU, and UpG allow secure assignments to be made for most of the resonances. This paper describes the syntheses and 13 C assignments of the oligomers.

The retention of aflatoxin residues in tissues of turkey poults fed a diet containing aflatoxin B 1 (500 ppb) for 18 days was determined. Free and conjugated aflatoxin metabolites were quantified using high-pressure liquid chromatography. Aflatoxin residue levels were greater in liver than muscle tissues, although all levels were low (range 0.01–1.19 ng/g tissue). Free and conjugated aflatoxins B 1 and M 1 , a metabolite of B 1 , were the principal tissue residues. Other metabolites investigated included aflatoxicol, which was detected in certain samples, and aflatoxin Q 1 which was not detected. Conjugated aflatoxins, hydrolysed and extracted from the aqueous tissue fractions, comprised 55–91% of the total detected aflatoxin residues. All aflatoxin residues were rapidly cleared following discontinuation of the dietary aflatoxin B 1 (half-life 1.4 days for total aflatoxin clearance from liver). These results provide further evidence that tissues from animals maintained on diets containing aflatoxins do not provide a major source of aflatoxins when consumed by man.

A program system for automated HPLC measurements of vitamin A concentration is reported. This system provides for guidance of operators, for control of apparatus and for supervision of quality measurements.

An assay is described based on a high pressure liquid chromatography system for measurement of vitamin D3 and 25-hydroxyvitamin D3 concentration in adipose tissue and muscle. The sensitivity of the assay is less than 1 ng/g of tissue. Neither vitamin D3 nor 25-hydroxyvitamin D3 were found in tissues of vitamin D-deficient rats using this method. 25-Hydroxyvitamin D3 could not be detected in adipose tissue and very little was found in muscle of normal rats. The mean vitamin D3 concentrations in 15 samples of human perirenal adipose tissue was 45.3 ng/g +/- 22.2 (SD) and in 6 samples of axillary tissue 115.6 ng/g +/- 52.4 (SD). Human adipose tissue contains a substantial amount of vitamin D3 and its contribution to the maintenance of vitamin D status is discussed.

Pyrene and 1-methylpyrene in the pyrolysis product of cellulose, which enhances mutagenicity of 2-acetylaminofluorene, must be metabolized to expose enhancing activity. 1,6-Pyrenequinone and 1,8-pyrenequinone, having high enhancement activity, were first isolated and identified in the metabolites of pyrene by rat liver microsomal fraction.