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Summary Gallic acid (3,4,5-trihydroxybenzoic acid, GA) is widely distributed in nature, being a major phenolic pollutant and a commonly used antioxidant and building-block for drug development. We have char- acterized the first complete cluster (gal genes) responsible for growth in GA in a derivative of the model bacterium Pseudomonas putida KT2440. GalT mediates specific GA uptake and chemotaxis, and highlights the critical role of GA transport in bacterial adaptation to GA consumption. The proposed GA degradation via the central intermediate 4-- oxalomesaconic acid (OMA) was revisited and all enzymes involved have been identified. Thus, GalD is the prototype of a new subfamily of isomerases that catalyses a biochemical step that remained unknown, i.e. the tautomerization of the OMAketo generated by the GalA dioxygenase to OMAenol. GalB is the founding member of a new family of zinc-containing hydratases that converts OMAenol into 4-carboxy-4- hydroxy-2-oxoadipic acid (CHA). galC encodes the aldolase catalysing CHA cleavage to pyruvic and oxaloacetic acids. The presence of homologous gal clusters outside the Pseudomonas genus sheds light on the evolution and ecology of the gal genes in GA degraders. The gal genes were used for expanding the metabolic abilities of heterologous hosts towards GA degradation, and for engineering a GA cellular biosensor.

The amperometric biosensing of aromatic amines using a composite glucose oxidase (GOD)-peroxidase (HRP) biosensor in reversed micelles is reported. Rigid composite pellets of graphite and Teflon, in which GOD and HRP were coimmobilized by simple physical inclusion, were employed for the biosensor design. This design allows the in situ generation of the H2O2 needed for the enzyme reaction with the aromatic amines, thus preventing the negative effect that the presence of a high H2O2 concentration in solution has on HRP activity. The H2O2 in situ generation is performed by oxidation of glucose catalyzed by GOD. The effect of the composition of the reversed micelles, i.e., the nature of the organic solvent used as the continuous phase, the nature and concentration of the surfactant used as emulsifying agent, the aqueous 0.05 mol L−1 phosphate buffer percentage used as the dispersed phase, and the glucose concentration in the aqueous phase, on the biosensor response was evaluated. Reversed micelles formed with ethyl acetate, a 5% of phosphate buffer (pH 7.0) containing 3.0×10−3 mol L−1 glucose, and 0.1 mol L−1 AOT (sodium dioctylsulfosuccinate), were selected as working medium. Well-defined and reproducible amperometric signals at 0.00 V were obtained for p-phenylenediamine, 2-aminophenol, o-phenylenediamine, m-phenylenediamine, 1-naphthylamine, o-toluidine and aniline. The useful lifetime of one single biosensor was of 60 days. The trend in sensitivity observed for the aromatic amines is discussed considering the effect of their structure on the stabilization of the radicals formed in the enzyme reaction which are electrochemically reduced. The behavior of the composite bienzyme electrode was also evaluated in a FI (flow injection) system using reversed micelles as the carrier. The suitability of the composite bienzyme electrode for the analysis of real samples was demonstrated by determining aniline in spiked carrots.

A biosensor design, in which glucose oxidase and peroxidase are coimmobilized by simple physical inclusion into the bulk of graphite-Teflon pellets, is reported for the detection of phenolic compounds. This design allows the “in situ” generation of the H2O2 needed for the enzyme reaction with the phenolic compounds, which avoids several problems detected in the performance of single peroxidase biosensors as a consequence of the presence of a high H2O2 concentration. So, a much lower surface fouling was found at the GOD-HRP biosensor in comparison with a graphite-Teflon-HRP electrode, suggesting that the controlled generation of H2O2 makes more difficult the formation of polymers from the enzyme reaction products. The construction of trienzyme biosensors, in which GOD, HRP and tyrosinase were coimmobilized into the graphite-Teflon matrix is also reported, and their performance was compared with that of GOD-HRP bienzyme electrodes. The practical applicability of the composite multienzyme amperometric biosensors was evaluated by the estimation of the phenolic compounds content in waste waters from a refinery, and the results were compared with those obtained by using a colorimetric official method based on the reaction with 4-aminoantipyrine.

Graphite-Teflon-peroxidase composite electrodes, constructed by simple physical inclusion of the enzyme into the bulk of the electrode matrix, have been used for the amperometric detection of 18 phenolic compounds. Monitoring of the enzyme reaction was carried out by the electrode reduction of the generated phenoxy radicals, in the presence of a constant concentration of 5.0×10−4 mol L−1 H2O2, at an applied potential of 0.00 V. Different aspects regarding the stability of the composite bioelectrode showed the robustness of the biosensor design. The kinetic constants of the enzyme reactions for the phenolic compounds tested were calculated. The limits of detection obtained were better than those found in the literature for peroxidase biosensors using other composite matrices, and the differences in sensitivity observed for the different phenolic substrates are discussed in terms of their pKa values and substituents. The analytical signals obtained with the graphite-Teflon-HRP electrodes have been compared with those yielded by a graphite-Teflon-tyrosinase composite electrode constructed in a similar way. With the aim of monitoring the largest possible number of phenolic compounds with the highest possible sensitivity, coimmobilization of peroxidase and tyrosinase into the composite electrode matrix was carried out.

Setting: Prenatal diagnosis unit of the Suleymaniye Maternity Hospital and Dr. Fehmi Yazicioglu’s FMF center Materials and Methods: All women screened with a protocol including nuchal translucency + nasal bone + tricuspid regurgitation + biochemistry during the first trimester and having known outcome data were reevaluted retrospectively by dropping out the TR data and recalculation of the risks using the FMF software. With a risk level of 1/300 as the threshold for invasive testing, the diagnostic efficiency of both models for the diagnosis of aneuploidy was compared in terms of sensitivity, specifity and positive and negative predictive values. Findings: A total of 429 patients with a mean age 28,6 ± 5,1 had complete outcome data after FTS. 8 fetuses had aneuploidy consisting of trisomy 21, 18, 13 and 5pin 4, 2, 1and 1fetuses respectively. 28 in TR and 23 in non TR protocol were screen positive. Both models could identify 7 of 8 aneuploidies. False positive rate was 4.8% with and 3.7% without TR. The sensitivity, specifity, positive and negative predictive values of the FTS with and without TR were 0.87, 0.95, 0.25, 0.99 and 087, 0.96, 0.30, 0.99 respectively. Conclusions: Adding TR to FTS in a low risk population resulted in increased false positivity without affecting the overall sensitivity for aneuploidy