Biological variability in aspartate aminotransferase activity in serum of healthy persons, and effect of in vitro supplemantation with pyridoxal 5-phosphate.

1977 ◽  
Vol 23 (3) ◽  
pp. 551-554 ◽  
Author(s):  
M Horder ◽  
G N Bowers
Keyword(s):  
Aspartate Aminotransferase ◽  
Pyridoxal Phosphate ◽  
Reference Interval ◽  
Individual Variability ◽  
Biological Variability ◽  
Healthy Individuals ◽  
Aminotransferase Activity ◽  
Measured Activity ◽  
Phosphate Supplementation

Abstract Aspartate aminotransferase (EC 2.6.1.1) activities in sera from nine healthy individuals were monitored during two weeks, both with and without first supplementing the serum with pyridoxal phosphate. Pyridoxal phosphate supplementation caused a mean increase of 39% (range, 33-55%) in measured activity. The biological variability during the two-week period was independent of pyridoxal phosphate supplemantation. The intra-individual variability (CV) was 5.3% and 5.1% with and without pyridoxal phosphate supplementation, respectively; the corresponding inter-individual variability was 13.2% and 13.6%. We conclude that the reference interval will be insensitive to intra-individual fluctuations in aspartate aminotransferase activity in serum, whether or not the serum is supplemented with pyridoxal phosphate.

Effects of Buffers on Aspartate Aminotransferase Activity and Association of the Enzyme with Pyridoxal Phosphate

1975 ◽  
Vol 21 (11) ◽  
pp. 1585-1591 ◽  
Author(s):  
Robert Rej ◽  
Raymond E Vanderlinde
Keyword(s):  
Schiff Base ◽  
Aspartate Aminotransferase ◽  
Phosphate Buffer ◽  
Pyridoxal Phosphate ◽  
Human Origin ◽  
Aminotransferase Activity ◽  
Factors Influencing ◽  
Absorbance Spectra ◽  
Stimulation Of

Abstract Using purified enzymes of human origin and patients’ sera, we examined factors influencing the in vitro association of pyridoxal phosphate with aspartate aminotransferase (EC 2.6.1.1). The rate of association was markedly retarded by phosphate buffer in comparison with tris(hydroxymethyl)aminomethane or six other buffers. Pyridoxal phosphate at an incubation concentration of 130 µmol/liter reactivated the entire apoenzyme portion of an apoenzyme/holoenzyme mixture within 5 min in tris(hydroxymethyl)aminomethane; in contrast, less than 20% was associated during 15 min in phosphate. Activity measured in tris(hydroxymethyl)aminomethane-buffer without exogenous pyridoxal phosphate was 4% greater than that in phosphate and was slightly increased by increasing the pH of the assay mixture from 7.5 to 8.0. Aspartate in the incubation medium did not retard the stimulation in tris(hydroxymethyl)aminomethane buffer. While the magnitude of stimulation varied greatly among sera, a consistent mean stimulation of 30% for groups of sera with normal activities was found when asparate at 125 mmol/liter, 2-oxoglutarate at 6.7 mmol/liter and tris(hydroxymethyl)aminomethane at 90 mmol/liter were used, an increase over the 16% with phosphate buffer [Clin. Chem. 19, 92 (1973)]. Absorbance spectra suggest pyridoxal phosphate exists as the Schiff base of tris(hydroxymethyl)aminomethane or aspartate, or both, under conditions of assay incubation (without addition of 2-oxoglutarate). Nonenzymatic catalysis of the reaction by pyridoxal phosphate alone or a formation of a protein/pyridoxal phosphate adduct was discounted with use of D-asparate substrates

Increased Aspartate Aminotransferase Activity of Serum after in Vitro Supplementation with Pyridoxal Phosphate

1973 ◽  
Vol 19 (1) ◽  
pp. 92-98 ◽  
Author(s):  
Robert Rej ◽  
Charles F Fasce ◽  
Raymond E Vanderlinde
Keyword(s):  
Aspartate Aminotransferase ◽  
Pyridoxal Phosphate ◽  
Colorimetric Method ◽  
Small Range ◽  
Aminotransferase Activity ◽  
Kinetic Assay ◽  
Serum Supplementation ◽  
Sample Dilution ◽  
Stimulation Of

Abstract We examined the effect of pyridoxal phosphate supplementation on the apparent aspartate aminotransferase (EC 2.6.1.1.) activity of human serum. Supplementation by 25 µmol/liter effected an average increase of 16% in the results for kinetic assay. The increase was not the result of increased enzymatic or nonenzymatic blanks, and, within a small range, sample dilution had no significant effect. Part of the increase was attributable to the enzyme being protected against the loss of activity that occurs during preincubation with L-aspartate. A similar increase was not demonstrated in a two-point colorimetric method, perhaps because of the short reaction time, without preincubation, and the initial presence of both substrates in the assay. We attempted to Correlate such stimulation of aminotransferase activity and the patient’s diagnosis or treatment. Pyridoxal phosphate should be included in the reaction mixture when aspartate aminotransferase activity is being measured clinically

Aspartate aminotransferase activity in human serum. Factors to be considered in supplementation with pyridoxal 5'-phosphate in vitro.

1976 ◽  
Vol 22 (11) ◽  
pp. 1876-1883 ◽  
Author(s):  
M Horder ◽  
R E Moore ◽  
G N Bowers
Keyword(s):  
Human Serum ◽  
Aspartate Aminotransferase ◽  
Pyridoxal Phosphate ◽  
Normal Activity ◽  
Aminotransferase Activity ◽  
Serum Factors ◽  
Volume Fractions ◽  
Sources Of Error ◽  
Potential Sources

Abstract The pyridoxal phosphate reactivation of the apo form of aspartate aminotransferase (EC 2.6.1.1) in human serum has been studied with "normal" and above-normal activity of this enzyme. The extent of the reactionation did not depend on the presence of the substrates, L-aspartate or 2-oxoglutarate. Reactivation was greatest with 110 mumol of added pyridoxal phsophate present per liter during a preinucation for 7 min in tris(hydroxymethyl)methylamine buffer wit;h serum volume fractions ranging from 0.017 to 0.267. In comparison with measurements prformed with no exogenous pyridoxal phosphate present, we found two potential sources of error when this cofactor was added: (a) reagent and sample blanks in the pyridoxal phosphate-supplemented system were two- to eightfold higher and (b) progress curves were nonlinear when L-aspartate rather than 2-oxoglutarate was used as the startin substrate. Aspartate aminotransferase measurement sith pyridoxal phosphate supplementation was slightly more precise than without.

scholarly journals Selective inhibition of alanine aminotransferase and aspartate aminotransferase in rat hepatocytes

1984 ◽  
Vol 220 (3) ◽  
pp. 707-716 ◽  
Author(s):  
N W Cornell ◽  
P F Zuurendonk ◽  
M J Kerich ◽  
C B Straight
Keyword(s):  
Aspartate Aminotransferase ◽  
Alanine Aminotransferase ◽  
Rat Hepatocytes ◽  
Selective Inhibition ◽  
Enoic Acid ◽  
Urea Synthesis ◽  
Aminotransferase Activity ◽  
Glucose Synthesis ◽  
Alanine Aminotransferase Activity

Experiments were conducted with intact rat hepatocytes to identify inhibitors and incubation conditions that cause selective inhibition of alanine aminotransferase or aspartate aminotransferase. Satisfactory results were obtained by preincubating cells with L-cycloserine or L-2-amino-4-methoxy-trans-but-3-enoic acid in the absence of added substrates. When cells were incubated for 20 min with 50 microM-L-cycloserine, alanine aminotransferase activity was decreased by 90%, whereas aspartate aminotransferase was inhibited by 10% or less. On subsequent incubation, synthesis of glucose and urea from alanine was strongly inhibited, but glucose synthesis from lactate was unaffected. L-2-Amino-4-methoxy-trans-but-3-enoic acid (400 microM) in hepatocyte incubations caused 90-95% inactivation of aspartate aminotransferase, but only 15-30% loss of alanine aminotransferase activity. After preincubation with the inhibitor, glucose synthesis from lactate was almost completely blocked; with alanine as the substrate, gluconeogenesis was unaffected, and urea synthesis was only slightly decreased. By comparison with preincubation with inhibitors, simultaneous addition of substrates (alanine; lactate plus lysine) and inhibitors (cycloserine; aminomethoxybutenoic acid) resulted in smaller decreases in aminotransferase activities and in metabolic rates. Other compounds were less satisfactory as selective inhibitors. Ethylhydrazinoacetate inactivated the two aminotransferases to similar extents. Vinylglycine was almost equally effective in blocking the two enzymes in vitro, but was a very weak inhibitor when used with intact cells. Concentrations of DL-propargylglycine (4 mM) required to cause at least 90% inhibition of alanine aminotransferase in hepatocytes also caused a 16% decrease in aspartate aminotransferase. When tested in vitro, alanine aminotransferase was, as previously reported by others, more sensitive to inhibition by amino-oxyacetate than was aspartate aminotransferase, but in liver cell incubations the latter enzyme was more rapidly inactivated by amino-oxyacetate.

scholarly journals The effect of oral contraceptives on the apparent vitamin B6status in some Sudanese women

1986 ◽  
Vol 56 (2) ◽  
pp. 363-367 ◽  
Author(s):  
Eltayeb Y. Salih ◽  
Asma A. Zein ◽  
Riad A. Bayoumi
Keyword(s):  
Side Effects ◽  
Nutritional Status ◽  
Oral Contraceptives ◽  
Aspartate Aminotransferase ◽  
Low Dose ◽  
Pyridoxal Phosphate ◽  
Vitamin B ◽  
In Vitro Activation ◽  
Dose Oral

1. In vitro activation of erythrocyte aspartate aminotransferase (EC2.6.1.1) activity by pyridoxal phosphate was used to assess vitamin B4nutritional status in forty Sudanese women taking combined, low-dose oral contraceptives (oestrogen-progestogen; OC) and in thirty healthy, non-pregnant women not taking OC.2. Fourteen (35%) out of forty OC users showed apparent vitamin B4deficiency.3. Side-effects associated with OC were more common among the apparently vitamin-B4-deficient OC users than among OC users and non-OC users not deficient in vitamin B4.

scholarly journals Red cell aspartate aminotransferase saturation with oral pyridoxine intake

2005 ◽  
Vol 123 (2) ◽  
pp. 54-57 ◽  
Author(s):  
Marilena Oshiro ◽  
Kimiyo Nonoyama ◽  
Raimundo Antônio Gomes Oliveira ◽  
Orlando Cesar de Oliveira Barretto
Keyword(s):  
Aspartate Aminotransferase ◽  
Vitamin B6 ◽  
Pyridoxal Phosphate ◽  
In Vitro Study ◽  
Vitro Study ◽  
Red Cell ◽  
Sideroblastic Anemia ◽  
Before And After

CONTEXT AND OBJECTIVE: The coenzyme of aspartate aminotransferase is pyridoxal phosphate, generated from fresh vegetables containing pyridoxine. Vitamin B6-responsive sideroblastic anemia, myelofibrosis and Peyronie’s syndrome respond to high pyridoxine doses. The objective was to investigate the oral pyridoxine oral dose that would lead to maximized pyridoxal phosphate saturation of red cell aspartate aminotransferase. DESIGN AND SETTING: Controlled trial, in Hematology Division of Instituto Adolfo Lutz. METHODS: Red cell aspartate aminotransferase activity was assayed (before and after) in normal volunteers who were given oral pyridoxine for 15-18 days (30 mg, 100 mg and 200 mg daily). In vitro study of blood from seven normal volunteers was also performed, with before and after assaying of aspartate aminotransferase activity. RESULTS: The in vivo study showed increasing aspartate aminotransferase saturation with increasing pyridoxine doses. 83% saturation was reached with 30 mg daily, 88% with 100 mg, and 93% with 200 mg after 20 days of oral supplementation. The in vitro study did not reach 100% saturation. CONCLUSIONS: Neither in vivo nor in vitro study demonstrated thorough aspartate aminotransferase saturation with its coenzyme pyridoxal phosphate in red cells, from increasing pyridoxine supplementation. However, the 200-mg dose could be employed safely in vitamin B6-responsive sideroblastic anemia, myelofibrosis and Peyronie’s syndrome treatment. Although maximum saturation in circulating red cells is not achieved, erythroblasts and other nucleated and cytoplasmic organelles containing cells certainly will reach thorough saturation, which possibly explains the results obtained in these diseases.

Design, validation and performance of aspartate aminotransferase- and lactate dehydrogenase-reporting algorithms for haemolysed specimens including correction within quality specifications

2019 ◽  
pp. 000456321987847
Author(s):  
Selcuk Colak ◽  
Onur Tasdemir ◽  
Marianne van der Schaaf ◽  
Frans Opdam ◽  
Vincent van den Noort ◽  
...  
Keyword(s):  
Lactate Dehydrogenase ◽  
Aspartate Aminotransferase ◽  
Total Error ◽  
Reference Interval ◽  
Quality Specifications ◽  
Allowable Error ◽  
And Performance ◽  
Roche Diagnostics ◽  
The Impact

Background In vitro haemolysis is a major operational challenge for medical laboratories. A new experimental design was used to investigate under what conditions algorithms could be designed to report either quantitative or qualitative aspartate aminotransferase and lactate dehydrogenase results outside the manufacturer’s haemolysis specifications. Quantitative corrections were required to meet prespecified quality specifications. Methods Twenty-five patient samples were used to design reporting algorithms and another 41 patient samples were used to validate the algorithms. Aspartate aminotransferase, lactate dehydrogenase and haemolysis index were determined using a Cobas 6000 analyser (Roche diagnostics, Mannheim, Germany). Correction factors were determined, and the accuracy of the correction was investigated. Reporting algorithms were designed based on (i) the manufacturer’s cut-off for the haemolysis index, (ii) corrections within the total allowable error specification and (iii) qualitative reporting based on obtained results. The impact of the reporting algorithms was retrospectively determined by recalculating six months of aspartate aminotransferase and lactate dehydrogenase results. Results No correction for aspartate aminotransferase/lactate dehydrogenase was possible for results below the upper reference interval limit, while results equal to or greater than the upper reference interval limit could, up to mild haemolysis, be corrected within the total error criterion. All samples generated from the validated patient cohort fulfilled the set criteria. The algorithms allowed reporting 88.5% and 85.9% of otherwise unreported aspartate aminotransferase and lactate dehydrogenase results, respectively. Conclusions An approach is presented that allows to generate and validate reporting algorithms for aspartate aminotransferase and lactate dehydrogenase compatible with prespecified quality specifications. The designed algorithms resulted in a significant reduction of otherwise unreported aspartate aminotransferase and lactate dehydrogenase results.

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