Photolysis and ozonolysis of (iso) desmosine-containing crosslinked peptides from porcine aorta elastin

This report describes the use of photolysis and ozonolysis as a means of achieving complete cleavage of the pyridinium ring of (iso)desmosine in crosslinked elastin peptides. Although photolysis leads to the opening of the ring with concomitant formation of lysine, the peptide chains remain attached. Subsequent ozonolysis is able to completely achieve the cleavage of the rest of the ring skeleton, thus leading to the separation of the peptide chains. Formation of new amino acids, i.e. α-aminoadipic and glutamic acids, is emphasized. Localization of these amino acids within the released peptides should be of help in structural investigations on the crosslinking zones involving either isodesmosine or desmosine. However, other amino acids such as tyrosine and phenylalanine are sensitive to this procedure and side reactions occur which are responsible for peptide bond cleavage with the formation of breakdown products.     

Sino-Atrial Dissociation: Evidence by Intracardiac Recordings in Man and by Microelectrode Studies on Isolated Rabbit Atrium

The electrophysiologic mechanisms of sinus dysfunction have recently been determined by direct recordings of the sinus node electrogram. The association of various degrees of abnormalities in the formation of the impulse within the sinus node and of sinoatrial conduction block, represents the pathophysiological substrate of the mechanism of sinus node dysfunction. The purpose of this work is to present clinical and experimental data supporting the concept of sinus node isolation. In our clinical case, the sinus node was probably intact despite aspects of sinus node dysfunction on the surface ECG. Sinus node electrograms were recorded with a sinoatrial conduction time of 100 ms (normal values in our laboratory: 83 ms +/- 38 ms). Atrial mapping demonstrated that the area depolarized by the sinus node involved a 2 cm2 zone surrounding it. This perisinusal activity could not be recorded on the surface ECG. Both exit and entry blocks in the sinus node were demonstrated. Our experimental data showed a total desynchronization between the electrical activity of the sinus node and that of the atrium under hypoxic conditions. Both types of cases demonstrated that an atrial dysrhythmia was coexisting with regular sinus activity. From these data we concluded that a sinus node free from any pathological involvement could be associated with severe symptoms of sinus node dysfunction on the surface ECG.     

Solubility of peptides in trichloroacetic acid (TCA) solutions Hypothesis on the precipitation mechanism

The assessment of proteolysis levels is often achieved by global quantification of the peptides soluble at different TCA concentrations, but little information is available on the features of this precipitation mechanism. Peptic, tryptic and chymotryptic digests of x_sl, β, and κ caseins have been prepared and fractionated by RP-HPLC and each isolated peptide was identified. Each digest was precipitated by adding TCA to different final concentrations (2, 4, 8, and 12%). The soluble fraction was analysed by RP-HPLC. Relationships have been searched between the properties of 75 peptides obtained in this way, and their solubilities in TCA. The best correlation was found with the peptide retention time in RP-HPLC, which can be regarded as the experimental measure of peptide hydrophobicity. We concluded that TCA, by interacting with peptides, induces an increase of the hydrophobicity of peptides which can lead to aggregation through hydrophobic interactions.     

Separation of haemoglobin HbE and HbA2 by the fully automated,high‐pressure liquid chromatography Tosoh HLC‐723 G7 analyzer

High‐pressure liquid chromatography instruments specifically devised for separating haemoglobin (Hb) fractions have been increasingly employed by the hospital laboratories over the recent years since they allow easy and fast screening for several Hb variants. Although such instruments may be proposed as sensitive, specific and reliable alternatives to the classic electrophoretic techniques, a major drawback of this screening strategy is the almost identical retention time of several Hb variants. In particular, at least 18 Hb variants have been reported in the same retention window as HbA₂, including HbE, the second most common β‐chain variant in humans after sickle cell trait. Recently, we evaluated the performance characteristics of an improved buffer formulation originally conceived for Hb variants separation procedures on the fully automated high‐pressure liquid chromatography instrument Tosoh G7. At variance with other fully automated high‐pressure liquid chromatography analyzers, the elution pattern on the G7 in subjects heterozygous for HbE is characterized by the presence of four suggestive peaks (HbF, HbA, HbA₂ and HbE), confirming the effective separation of HbE from HbA₂. Because of its potential value in the diagnosis of the thalassaemia syndromes, the effective separation of HbA₂ from HbE can provide clinical laboratories with a valuable information for the diagnostic reasoning.     

Work hours,sleep patterns and fatigue among merchant marine personnel

SUMMARY A field study of work and sleep patterns among commercial merchant marine personnel is reported. Data collected over a 10-30-d period from 141 subjects aboard eight ships included information concerning work-rest schedules, sleep timing, alertness on the job and critical fatigue. The data indicate that watchstanders on the 4-on, 8-off schedule show considerable disruption in their sleep. The average sleep duration for all mariners is 6.6 h; watchstanders obtain their sleep in fragmented periods that are frequently less than 5 h in duration. Analysis of critical fatigue shows an incidence of 1-24% across personnel and measures. Of particular concern are the watchstanders on the 04.00-08.00 schedule, who sleep less than 4 h per 24-h period 22% of the time. Potential countermeasures, including changes in scheduling and staffing are proposed.     

Assay for Recombinant and Native Human Intraacrosomal Antigen SP-10

PROBLEM: To develop a method to measure a recombinant sperm protein, SP-10, during scale up and purification for a contraceptive vaccine formulation. METHOD: A quantitative assay method for the human intraacrosomal protein SP-10 was developed utilizing the format of indirect capture enzyme-linked immunosorbent assay (ELISA). A SP-10 specific monoclonal antibody mAb, MHS-10, was used as the capture antibody. Two recognition reagents, a rabbit polyclonal anti-SP-10 antisera (pAb) and a biotin-labeled mAb, MHS-10, were used as the recognition antibodies, respectively. A SP-10 recombinant fusion protein consisting of 125 SP-10 amino acids linked to glutathione transferase was used as a working SP-10 standard. The coefficient of variance for the assay system using the rabbit pAb was in the range of 0.099 to 0.157, and for the assay system using the biotinylated mAb MHS-10 was in the range of 0.081 to 0.084. RESULTS: Employing biotinylated MHS-10 as the recognition antibody, it was found that the native SP-10 molecule had more than one MHS-10 epitope. The concentration of SP-10 in a pool of human sperm extracts was found to be approximately 1% of the total proteins, assayed by both of the recognition antibody systems. CONCLUSIONS: The assay system described is useful to monitor the yield of recombinant SP-10 during scale-up production of the SP-10 vaccine.