Determination of aluminum in bone in haemodialyzed patients, using inductively coupled argon plasma emission spectrometry.

We propose a new method for measuring aluminium in bone tissue, using argon plasma emission spectrophotometry. The detection limit in the bone nitric digestion liquid was 0.015 mumol/l (corresponding to 0.0075 mumol/g, i.e. 0.2 microgram/g for a tissue sample with 1.0 g wet weight). Within-run CVs were 4.66% and 1.43% for tissues containing 0.15 and 0.64 mumol/g, respectively. Other bone constituents such as calcium, phosphorus, magnesium, sodium, potassium, do not affect aluminum results. Normal values obtained in bone of subjects not suspected of aluminium intoxication were: mean +/- SD; 0.09 +/- 0.044 mumol/g (n = 24). In dialysis patients we found a mean bone aluminium content of 0.75 mumol/g for concentrations ranging between 0.16 and 3.38 mumol/g.

[Blood concentrations of maprotiline in depressive patients]. / Etude des concentrations sanguines de maprotiline chez des patients déprimés.

Blood levels of Maprotiline were analysed and their relationship to the clinical response was examined in 89 depressed inpatients, according DSM III criteria for Major Depressive Episode, given the drug treatment for 3 weeks. Maprotiline produced marked decreases in mean MADRS and COVI scale scores by the end of treatment. On day 21, no correlation between blood levels of Maprotiline and MADRS or COVI scores were found when all patients were considered. Nevertheless, significant correlations were observed on day 14 (r = .22; p less than .05 for MADRS and r = .23; p less than .05 for COVI scale). In addition, a significant correlation between MADRS or COVI scale scores and Maprotiline blood levels were observed on days 14 and 21 in subgroups of young patients, severe depression (high scores to clinical global investigations), during of at least 3 months, treated without other drug than Maprotiline and good responders.

Rapid determination of iron in urine, in the presence of deferoxamine, by inductively coupled plasma emission spectrometry.

Using a spectrometer with an argon plasma source coupled to a high-frequency magnetic field, we developed a direct method for determining iron in urine of patients being treated with deferoxamine. The detection limit for iron was 75 nmol/L; added iron was satisfactorily recovered; and we observed no interference from deferoxamine at its most commonly used concentrations. Values for between-run and within-run precision (CV) was less than 5%. Correlation of results with those obtained with a colorimetric method involving bathophenanthroline was good (r = 0.96).

Swine liver L-leucine aminopeptidase: improved purification procedure.

An L-leucine aminopeptidase, having a specificity toward the substrate L-leucine amide, was purified 1084-fold from swine liver with a yield of 50.7 per cent. Purification procedure was carried out using successively centrifugation at 105 000 X g, fractionation by ammonium sulfate, DEAE Sephacel chromatography and zonal ultracentrifugation. Enzyme homogeneity and purity studies were carried out by analytical ultracentrifugation and polyacrylamide gel electrophoresis. In SDS-gel polyacrylamide, a single band was observed. It corresponded to a 55 000 molecular weight protein.

[Characterization of L-threonine deaminase activity of guinea pig liver induced by a high protein diet]. / Caractérisation de l'activité L-thréonine désaminasique du foie de cobaye induite par un régime hyperprotidique.

In a foregoing paper, we demonstrated that under equilibrated diet conditions, guinea pig liver L-threonine deaminase activity should be allocated to two distinct enzymes: a specific L-threonine deaminase without activity toward L-serine and a L-serine deaminase having a secondary activity toward L-threonine. In the present work, we observed that a high protidic diet caused an elevation of total threonine deaminase activity. Thus purification of guinea pig liver L-threonine deaminase was attempted, using ultracentrifugation, salt precipitation, heat treatment, ion exchange chromatography on DEAE Sephacel, Sephadex G 200 molecular sieve, 2 amino-2 methyl-1 propanol linked CH 4B Sepharose chromatography. The weak variations of the ratios of specific activities respectively toward L-threonine and L-serine observed at each stage of the purification procedure indicated that both activities are very likely supported by a single enzyme preexisting in the liver of guinea pigs fed an equilibrated diet. No isoenzyme was evidenced by polyacrylamide gel electrophoresis or DEAE Sephacel chromatography. Moreover, our purification procedure demonstrated that not only inducible L-threonine deaminase guinea pig liver activity was due to L-serine deaminase, but also that an initially existing specific L-threonine deaminase activity paradoxically disappeared with a protein rich diet.

Human liver L-leucine aminopeptidase: evidence for two forms compared to pig liver enzyme.

Two forms of L-leucine aminopeptidase (E.C. 3.4.11.1) having a specific activity toward L-leucine amide and L-leucylpeptides substrates but not toward chromogenic substrates: L-leucyl paranitroanilide or L-leucyl beta naphthylamide have been evidenced from human liver. Human liver enzymes have been distinguished from pig liver enzyme by DEAE Sephacel chromatography and analytical electrophoresis on cellulose acetate strips. We compared enzymic properties of L-leucine aminopeptidases from human liver with pig liver enzyme: they were activated by Mg2+ and Mn2+ and inhibited by Zn2+ and Co2+, EDTA and citric acid. The optimum pH's were 10. Both human liver L-leucine aminopeptidases were less sensitive to heat elevation than pig liver enzyme.

Purification and enzymatic properties of an L-leucine aminopeptidase from swine liver.

An L-leucine aminopeptidase (alpha-aminoacyl-peptide hydrolase (cytosol), EC 3.4.11.1), having a specificity toward the substrate L-leucine amide, but not toward L-leucyl beta-naphthylamide or L-leucyl p-nitroanilide, has been purified 332-fold from swine liver, with a yield of 8.6%. This is the first purification of this enzyme from hepatic tissue. The purified enzyme submitted to analytical electrophoresis on cellulose acetate strips or in polyacrylamide gel showed a single band after straining with Ponceau S Red dye or Amido black, respectively. Purified swine liver L-leucine aminopeptidase, a cytosol enzyme, exhibited a molecular weight of 268 000 +/- 50 000 by gel filtration. It hydrolyzed L-leucine amide substrate and L-leucyl peptides. It was activated by Mg2+ and Mn2+ and inhibited by Co2+ and Zn2+. The optimum pH was 10. It was rather sensitive to heat elevation. Swine liver L-leucine aminopeptidase was inhibited by EDTA, citric acid, isocaproic acid, dodecylamine, aliphatic alcohols and p-chloromercuribenzoate but unaffected by monoiodoacetic acid and diisopropyl fluorophosphate.

[Neutral leucinamidasic activity of human serum. Semiautomatic method of analysis]. / L'activité leucinamidasique neutre du sérum humain. Méthode de dosage semi-automatique.

Half automated method for determining a L-leucinamide splitting enzymatic activity in human sera. In normal and pathological human sera, two distinct L-leucinamide splitting enzymatic activities have been demonstrated. One of them has an optimal activity at pH 9 (alcaline leucine amidase activity) and shows the most properties of a classic leucinaminopeptidase (E.C. 3.4.11.1). The other (neutral leucine amidase activity) has an optimal activity at pH : 7,5--7,8 and is not activated by Mg2+ ions. In the present work a semi-automated method permitting the determination of the "neutral leucine amidase activity" is presented. The mean of the reference values for normal human sera are established to 31,54 mUI/ml, and the upper normal limit is 48 mUI/ml. The neutral leucine amidase activity is studied in pathological sera comparatively with two other aminopeptidase activities : "alcaline leucine amidase activity", and "leucine-arylamidase". Our study shows that in pathological sera, the neutral leucine amidase activity" varies often without any correlation with those parameters.

[Guinea pig liver cytosol has an L-threonine deaminase activity distinct from L-serine deaminase]. / Mise en évidence dans le cytosol du foie de cobaye d'une activité L-thréonine désaminasique distinct de la L-sérine désaminase

Using sodium sulfate precipitation, "Sephadex G200" gel filtration and polyacrylamide gel electrophoresis, a L-threonine desaminase was demonstrated in the Guinea-Pig liver cytosol. This enzyme was separated from the guinea pig liver L-serine desaminase possessing an auxiliary activity on L-threonine substrate described by us in a previous work. The optimals for pH (7,1) and temperature (+ 55 degrees C) and the apparent molecular weight (134,000 + 20,000) were established.

Guine pig liver L-asparaginase. Separation, purification, and intracellular localisation of two distinct enzymatic activities.

Two distinct L-asparaginase (EC 3.5.1.1) activities were detected in guinea pig liver: Asparaginase 1 and Asparaginase 2. Asparaginase 1 has been purified 272 fold from the crude homogenate; its molecular weight was evaluated by gel filtration to be about 150 000. The purified preparation was shown to be homogeneous by cellulose acetate strip and polyacrylamide disc-gel electrophoresis. Asparaginase 2 has been purified 63.5 fold from the crude homogenate. Its molecular weight was evaluated by gel filtration to be about 21 500. Cellulose acetate strip electrophoresis demonstrated two bands, one of which corresponded to Asparaginase 1 and the other to Asparaginase 2. Cellular fractionation in the ultracentrifuge, showed Asparaginase 1 to be present only in the cytosol fraction. Asparaginase 2 which was unstable at 105 000 X g seemed mostly localized in the mitochondria and secondarily in the cytoplasmic fraction.

[Determination of blood iron and the total fixation capacity of transferrin by a simple automatic technic without dialysis]. / Détermination de la sidérémie et de la capacité totale de fixation de la transferrine par une technique automatique simple sans dialyse

An automated method without dialysis for determining serum iron is described. It is an adaptation of the Lauber procedure to the Technicon Auto Analyser I: serum iron is split from its protein combination by exposure to an acetate pH 5.8 buffer, in the presence of a detergent (Teepol 710) which prevents any precipitation of proteins. Iron is maintained in ferrous state by addition of sodium dithionite and is quantitated with bathophenanthrolin colorimetry. Serum samples are analysed at a rate of 60/H. It is necessary to run a blank for each serum sample. Between 20 and 500 mug iron/100 ml the color intensity agrees with Beer's law. The reproducibility is good, and the variation coefficient less than 1.2 per cent. Our results are not significatively different of those obtained with the ICSH standard method. Bilirubin does not interfere. Iron added is satisfactorily recovered. Our method can also be used for determining the transferrin iron binding capacity.

[Semiautomatic method of measurement of serum 5'nucleotidase activity]. / Méthode semi-automatique de mesure d'activité de la 5'-nucléotidase sérique

A semi-automated method for determining the 5'-nucleotidase activity in human serum for use with the Technicon Autoanalyzer I is described. Based upon the manual method of Persijn and Van Der Slik, adenosine formed by hydrolysis of 5'-AMP is deamined enzymatically and ammonia determined with the Berthelot reaction. The non specific alkaline phosphatases are inhibited by phenylphosphate. Concentrations of 5'-AMP and phenylphosphate are discussed. Nucleotidase activity is determined without dilution up to a value of 210 UI/liter. The precision is better than other manual or semi-automated methods in current use.