Reaction of acetaldehyde with hemoglobin

Acetaldehyde reacted with hemoglobin at neutral pH and 37 degrees C to form adducts that were stable to dialysis and that were not reduced by sodium borohydride. Hemoglobin tetramers having 2, 3, and probably 4 molar eq of bound aldehyde were isolated by cation exchange chromatography. The sites of attachment of the aldehyde were the free amino groups of the N-terminal valine residues of the alpha and beta chains of hemoglobin. Derivatization of the beta chains caused a greater increase in the acidity of the hemoglobin than did derivatization of the alpha chains. Derivatization of the beta chains was also preferred over that of the alpha chains. Acetaldehyde derivatives of the N-terminal octapeptide of hemoglobin S (beta sT-1 peptide), Val-Gly-Gly, and tetraglycine were formed readily, contained 1 M eq of acetaldehyde/mol of peptide, and were not reduced by sodium borohydride. In contrast, Ala-Pro-Gly failed to form a 1:1 adduct with acetaldehyde. 13C NMR analysis of the peptide adducts formed with [1,2-13C]acetaldehyde indicated that tetrahedral diastereomeric derivatives were produced. The 13C chemical shifts of the adducts formed between hemoglobin and [1,2-13C]acetaldehyde were identical to those of the peptide adducts although resonances from the individual diastereomeric adducts at each hemoglobin site could not be resolved. The results cited above as well as other evidence indicate that acetaldehyde reacts with the amino termini of hemoglobin to form stable cyclic imidazolidinone derivatives. An exchange of acetaldehyde residues between peptides was also documented.     

Secretory IgA and secretory component in women affected by recidivant vaginal candidiasis

Local immunity was evaluated in 47 patients affected by recidivant vaginal candidiasis and 33 control women. IgG, IgA, IgM and secretory component (SC) were determined by single radial immunodiffusion in samples of cervicovaginal secretion. IgG in dosable levels was detected in 17/47 samples (36.2%) and IgA in 15/47 patients (31.9%) whereas in the controls, the incidence was 31/33 (93.9%) for IgG and 24/33 (72.7%) for IgA. The difference was significative (P< 0.001) for both immunoglobulins. Significant differences were not obtained for IgM. The SC was detected in 4/47 cervicovaginal secretions of patients affected by candidiasis (8.5%) whereas in the control samples the incidence was 21/33 (63.6%) (P<0.001). In only 2/15 patients with dosable levels of IgA (13%) the secretory nature of this immunoglobulin could be shown by its reaction with anti-SC serum. In the control group, secretory IgA was detected in 19/24 cases (79%) (P< 0.001). Serum immunoglobulins levels were normal. The lack of secretory IgA and SC in the secretion could be related to the adherence capacity of the Candida albicans to epithelial cells.     

Restriction cleavage maps of the DNAs of Streptococcus pneumoniae bacteriophages containing protein covalently bound to their 5'' ends

Summary Several pneumococcal bacteriophages showing a morphology similar to that previously described for Cp-1 (Ronda et al. 1981) have been isolated and purified from throat samples taken from healthy children. Three of these phages (Cp-5, Cp-7 and Cp-9) have been studied in detail and compared to Cp-1. The four phages differed in several respects, e.g. size, structural polypeptides, restriction enzyme cleavage patterns, etc. The DNA of Cp-5, Cp-7 and Cp-9 showed protease-sensitive transfecting activity. This, together with the results obtained by electrophoretic analyses as well as by isotopic labelling of these DNAs with [-³²P] ATP and polynucleotide kinase indicated that all these new phages have a protein covalently linked to the 5 ends of their DNAs as in the case of Cp-1 (García et al. 1983). Restriction enzyme cleavage maps of Cp-1, Cp-5, Cp-7 and Cp-9 have been constructed.     

Multiple forms of lactate dehydrogenase in Staphylococcus aureus

Activities for nicotinamide adenine dinucleotide (NAD)-dependent and NAD-independent forms of lactate dehydrogenase (LDH) were measured in cell-free extracts of Staphylococcus aureus strain PS 6 for the d and l isomers of lactate. Data obtained for the NAD-dependent lactate dehydrogenases indicate that oxidation of both isomers of lactate is due to both an l-lactate-specific LDH and a lactate racemase. After acrylamide gel electrophoresis, two bands exhibiting LDH activity were detected in crude or in partially purified cell-free extracts. The fast band exhibited LDH activity that was not NAD-dependent for both isomers of lactate, whereas, the slow band had very high NAD-dependent LDH activity for the l isomer but just detectable activity or the d isomer. Both bands appeared when d-lactate was used as the substrate, but only the slow band was formed when l-lactate was the substrate. NAD-dependent LDH, in apparent association with a nonspecific tetrazolium-reducing protein, is responsible for the production of the slow band.     

Instrumental assay of microbial lipase at constant pH

A rapid, accurate method with high sensitivity and reproducibility, and having the advantage of a short incubation period under constant pH, has been developed for routine measurement of microbial lipase. Assembled from readily available and economical instrumental components, the apparatus includes a pH meter, a thermoelectric heating and stirring device, a motor-driven burette, and an automatic recorder. The reaction mixture, consisting of 5 ml of a 10% olive oil-gum arabic emulsion, 2 ml of 3 m NaCl, 2 ml of sodium taurocholate (15 mg/ml) of 0.075 m CaCl(2), 5 ml of water, and 1 ml of enzyme solution, was adjusted to pH 8.0 and 37 C. The pH was maintained at a constant value by automatic addition of 0.01 n NaOH during the incubation period, which usually lasted 5 min. A lipase unit, derived from the use of this technique, may be defined as the number of microequivalents of acid liberated per minute under the specified conditions. The method was sensitive to 0.01 units. Various organisms tested produced 0.17 to 1.32 units per ml of the cell filtrate. An Arrhenius plot for staphylococcal lipase yielded 14,500 cal for function A (energy of activation).     

Nephelometric Assay of Bovine Antistaphylocoagulase Serum

A nephelometric assay of staphylococcal coagulase has been utilized to measure coagulase inhibition by bovine anticoagulase serum. Suitably diluted antisera produced maximal inhibition when incubated with purified coagulase at pH 7.3 in phosphate-buffered saline for 15 min at 22 C or 1 hr at 4 C. Neutralization of coagulase activity was measured as the reduction in the clotting rate of a fibrinogen-plasma substrate, and was directly proportional to the concentration of antiserum over a wide range of coagulase activity. A unit of anticoagulase was defined as the amount of inhibitor that neutralized one unit of coagulase. In addition to the heat-stable (56 C, 30 min) antibody contained in the crude gamma-globulin fraction, a heat-labile, nondialyzable coagulase inhibitor was also detected in the sera from 15 of 16 randomly selected dairy cows.     

The Effects of Direct Instillations of a Therapeutic Mixture Into the Tracheobronchial Tree

Twenty-two patients with chronic productive bronchitis or bronchiectasis were treated by direct instillations of normal saline and N-acetylcysteine into the trachea through a percutaneous catheter following a period of conventional routine therapy.The instillation, using either normal saline or varying concentrations of N-acetylcysteine did not produce significant change in alveolar gas exchange as reflected by measurement of arterial PaCO₂ and the alveolar arterial gradient for oxygen during and after the introduction of the medication into the bronchial tree. Studies were carried out after patients had been stabilized breathing pure oxygen on an IPPB machine for 30 minutes.Evaluation of the treatment by means of pulmonary function tests demonstrated significant improvement in overall function following therapy.The results indicate that the technique of tracheobronchial lavage is physiologically benign and that overall improvement in pulmonary function can be obtained by this means in cases of the type described in this study.