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PARADOXICAL HYPERTENSION AND THE ABDOMINAL PAIN SYNDROME FOLLOWING RESECTION OF COARCTATION OF THE AORTA.

Sixteen children with coarctation of the aorta were studied in respect of the incidence of paradoxical hypertension and the abdominal pain syndrome after repair of the coarctation. These postoperative complications were investigated in relation to vanillylmandelic acid (VMA) excretion and extent of operative repair. The results, compared to those in a control group, indicated no evident causal relationship between these complications and catecholamine production. The severity of coarctation, the degree of correction and the operative increase in luminal surface area, however, appeared to be significant factors.

Abdomen↗

CHEMICAL TESTS FOR PHAEOCHROMOCYTOMA.

Urinary catecholamines, 4-hydroxy-3-methoxymandelic acid (V.M.A.), and metadrenalines have been estimated in 39 cases of proven phaeochromocytoma and in many hypertensive patients without phaeochromocytoma. The variation in excretion of these substances is described and their diagnostic significance discussed. Excretion of free catecholamines and metadrenalines showed a proportionately greater increase than of V.M.A. in patients with a phaeochromocytoma, and estimation of the first two appears to be diagnostically more reliable. Hingerty's method for catecholamines appears to be a satisfactory screening test, provided that 90 mug. is taken as the upper limit of normal, and estimation of metadrenalines is considered to be the best confirmatory test for phaeochromocytoma.

Catecholamines↗

L-mandelate dehydrogenase from Rhodotorula graminis: comparisons with the L-lactate dehydrogenase (flavocytochrome b2) from Saccharomyces cerevisiae.

L-Lactate dehydrogenase (L-LDH) from Saccharomyces cerevisiae and L-mandelate dehydrogenase (L-MDH) from Rhodotorula graminis are both flavocytochromes b2. The kinetic properties of these enzymes have been compared using steady-state kinetic methods. The most striking difference between the two enzymes is found by comparing their substrate specificities. L-LDH and L-MDH have mutually exclusive primary substrates, i.e. the substrate for one enzyme is a potent competitive inhibitor for the other. Molecular-modelling studies on the known three-dimensional structure of S. cerevisiae L-LDH suggest that this enzyme is unable to catalyse the oxidation of L-mandelate because productive binding is impeded by steric interference, particularly between the side chain of Leu-230 and the phenyl ring of mandelate. Another major difference between L-LDH and L-MDH lies in the rate-determining step. For S. cerevisiae L-LDH, the major rate-determining step is proton abstraction at C-2 of lactate, as previously shown by the 2H kinetic-isotope effect. However, in R. graminis L-MDH the kinetic-isotope effect seen with DL-[2-2H]mandelate is only 1.1 +/- 0.1, clearly showing that proton abstraction at C-2 of mandelate is not rate-limiting. The fact that the rate-determining step is different indicates that the transition states in each of these enzymes must also be different.

Alcohol Oxidoreductases↗

Evaluation of bactericidal activity and lag of regrowth (postantibiotic effect) of five antiseptics on nine bacterial pathogens.

Lag of regrowth or postantibiotic effect (PAE) relates to suppression of bacterial regrowth following short exposure to an antimicrobial agent. A delay in regrowth has not yet been studied for antiseptics to any great extent. We therefore examined and compared the lag of regrowth and the bactericidal activity of five antiseptics (chloramine T, chlorhexidine, povidone-iodine, phenoxyethanol and mandelic-lactic acid) against nine bacterial pathogens. Delay in regrowth was determined by application of two concentration-time schedules: a test concentration at the MBC with a contact time of 1 h or using fixed suboptimum concentration of each antiseptic for 2 min (optimum concentrations sterilized the culture, impeding assessment of regrowth) followed by a neutralization-dilution step and subsequent viable counting to follow bacterial regrowth. Each antiseptic displayed a different spectrum of activity in terms of MIC or MBC, bactericidal effect and lag of regrowth. The delay in regrowth varied from 0 to 5.7 h with only a few discrepancies between the two treatment schedules. Mandelic-lactic acid and chloramine T induced a significantly longer lag as compared with the other agents, whereas phenoxyethanol produced the shortest lag values. No significant correlation between the killing rate and the lag of regrowth could be demonstrated. Information on bactericidal activity, as well as lag of regrowth, could be a useful screening method for the efficacy of antiseptics. Moreover, data on lag of regrowth could contribute to the choice of antiseptic and guide in determining the optimum interval between repeated applications of antiseptics.

Anti-Infective Agents, Local↗

Capillary electrophoresis and supercritical chromatography, complementary and alternative techniques for the determination of urinary metabolites of styrene.

Two analytical methods without an extraction step were developed using capillary electrophoresis and supercritical fluid chromatography in order to determine phenylglyoxylic (PGA) and mandelic (MA) acids in urine, with minimum treatment and manipulation of biological samples. The urine was diluted ten-fold in acetonitrile and directly injected into the analytical systems after centrifugation. Analysis was performed by capillary electrophoresis on alkyl bonded phase capillary columns with sodium formiate (4 x 10(-2) M)-isopropanol (9:1, v/v) as a buffer, and by supercritical fluid chromatography on a Diol bonded phase silica column with ethanol-water-methanesulphonic acid (97.5:2.4:0.1, v/v) as coeluent of CO2. Detection of PGA and MA was performed by ultraviolet detection at 255 and 210 nm, respectively. The methods are in agreement, and are easily able to detect 5 mg/g creatinine for PGA, and 15 mg/g creatinine for MA, which are one twentieth of the lowest biological exposure index values.

Air Pollutants, Occupational↗

Characterization of a benzoate permease mutant of Pseudomonas putida.

A spontaneous mutant of Pseudomonas putida (PRS 2017) has been isolated which is incapable of growth on benzoate, does not induce the enzymes of the catechol branch of the beta-ketoadipate pathway when grown in the presence of benzoate, cannot accumulate radioactively labeled benzoate, yet grows well with mandelate as sole source of carbon and energy. This strain apparently lacks a benzoate permease, which in the wild type shows a Km of about 0.1 mM for benzoate, is inducible, and is not under the control of the regulatory system which governs the induction of the enzymes of the catechol branch of the beta-ketoadapate pathway. The lesion in PRS2017 is apparently single site and maps near other genes governing benzoate dissimilation.

Adipates↗

A new method for the analysis of styrene mercapturic acids by liquid chromatography/electrospray tandem mass spectrometry.

A new method based on liquid chromatography/tandem mass spectrometry has been developed for the direct determination of specific urinary mercapturic acids arising from the conjugation of (R)-and (S)-enantiomers of styrene 7,8-oxide with glutathione (GSH), i.e. (R,R)- and (S,R)-N-acetyl-S-(1-phenyl-2-hydroxyethyl)cysteine (R,R-M1 and S,R-M1) and (R,R)- and (S,R)-N-acetyl-S-(2-phenyl-2-hydroxyethyl)-cysteine (R,R-M2 and S,R-M2). The four diastereoisomers were separated on a C18-DB (7.5 cm, 3 microm) column using variable proportions of 20 mM aqueous ammonium formate buffer and methanol at a flow-rate of 0.5 mL/min. The analytes were ionized by electrospray, in negative-ion mode. Operating in selected-reaction monitoring mode, linearity of the MS response versus analyte concentration was established over 4 orders of magnitude, the detection limits being 0.7-1.0 microg/L for all the mercapturates. Precision of the method determined at 50 microg/L (n = 12), expressed as relative standard deviation, was respectively 3.1, 4.8 and 6.9% within the run, intra-day and inter-day. The corresponding figures at 1.0 mg/L (n = 12) were respectively 2.0, 3.6 and 5.5%. The method was applied to the quantitative analysis of conjugated metabolites in urine samples from workers occupationally exposed to styrene. The diastereoisomers R,R-M1 and S,R-M2 accounted respectively for 50 and 40% of total mercapturates, whereas the proportion of R,R-M2 was 7% and only minor amounts of S,R-M1 were detectable. Styrene mercapturates represented a minor fraction of total styrene metabolites, less than 1% on average. The ratio mercapturates/main metabolites (mandelic + phenylglyoxylic acid) showed a bimodal distribution, the medians of the two subgroups being 0.2 and 1%, respectively. Such subgroups are probably characterized by the genetic polymorphisms of the drug-metabolizing enzymes to be identified.

Acetylcysteine↗

On the interpretation of quantitative structure-function activity relationship data for lactate oxidase.

The native flavin, FMN, has been removed from the l-lactate oxidase of Aerococcus viridans, and the apoprotein reconstituted with 12 FMN derivatives with various substituents at the flavin 6- and 8-positions. Impressive linear relationships are exhibited between the sum of the Hammett final sigma(para) and final sigma(ortho) parameters and the redox potentials of the free flavins, and between the redox potentials of the free and enzyme-bound flavins. Rapid reaction kinetics studies of the reconstituted enzymes with the substrates l-lactate and l-mandelate show an increase in the reduction rate constant with increasing redox potential, except that, with lactate, a limiting rate constant of approximately 700 s(-1) is obtained with flavins of high potential. Similar breakpoints are found in plots of the rate constants for flavin N5-sulfite adduct formation and for the reaction of the reduced enzymes with molecular oxygen. These results are interpreted in terms of a two-step equilibrium preceding the chemical reaction step, in which the second equilibrium step provides an upper limit to the rate with which the particular substrate or ligand is positioned with the flavin in the correct fashion for the observed chemical reaction to occur. The relationship of rate constants for flavin reduction and N5-sulfite adduct formation with flavin redox potential below the observed breakpoint indicate development of significant negative charge in the transition states of the reactions. In the case of reduction by substrate, the results are consistent either with a hydride transfer mechanism or with the so called "carbanion" mechanism, in which the substrate alpha-proton is abstracted by an enzyme base protected from exchange with solvent. These conclusions are supported by substrate alpha-deuterium isotope effects and by solvent viscosity effects on sulfite binding.

Apoenzymes↗