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H U Bergmeyer

Publications and source records attributed to H U Bergmeyer.

At least 19 recordsLinked to original sources
PubMed1978

Optimization of methods for aspartate aminotransferase and alanine aminotransferase.

Conditions for accurate measurement of catalytic activity of aspartate aminotransferase and alanine aminotransferase in human serum have been reinvestigated. The basic variables (kind of buffer, buffer concentration, pH, ion effects, and the influence of pyridoxal-5-phosphate) can now be considered optimized. On this basis, the kinetic parameters of both aminotransferases were determined, i.e., Michaelis and inhibitor constants for substrates and reaction products. With a mathematical approach for two-substrate enzyme reactions the substrate concentrations were calculated from the viewpoints "most economical," "most convenient," and "lowest variability." Also the conditions for the indicator reactions have been newly defined with respect to a kinetic model. All calculated data were rechecked experimentally and it can be shown that both approaches fully agree. Furthermore, we show that the mathematical approach allows more precise recommendations for optimized methods. For technical reasons, the catalytic activity of aspartate aminotransferase in human serum can only be measured as a 0.96 fraction of its theoretical maximum velocity, the catalytic activity of alanine aminotransferase as a 0.91 fraction. The assay conditions for a Reference Method are finally described and recommendations are made for optimized routine methods for determination of the catalytic activity of these transferases in human serum.

Alanine Transaminase↗
PubMed1977

Evaluation of optimum conditions of two-substrate enzyme reactions.

A modern approach is described for the evaluation of the optimal conditions for two-substrate enzyme reactions. It chiefly involves the determination of the concentration of substrates for the primary reaction and the catalytic concentration of indicator enzymes. The interrelationship between the concentration of the two substrates (concentration pairs) are described mathematically to be hyperbolic, and, in case of competitively inhibited reactions, to be parabolic. Calculated optimum concentrations have been rechecked experimentally for the reactions of aspartate aminotransferase and alanine aminotransferase. For pyridine coenzyme linked indicator reactions it could be demonstrated that they mostly follow zero order kinetics. One of the products of the primary reaction reacts, in its steady state concentration, as the second substrate. This represents the size of the lag phase of the coupled reaction. The Km of this substance must be known in order to calculate the catalytic concentration of the indicator enzyme in relation to that of the primary enzyme. Its concentration can be fixed arbitrarily within certain limits, depending on whether the calculated result actually can be realized; otherwise a larger lag phase must be tolerated. For practical reasons, it is generally possible to measure only a certain percentage of maximum reaction rate.

Alanine Transaminase↗
PubMed1975

[The Km of malate dehydrogenase from pig heart with oxaloacetate as substrate (author's transl)].

Km is necessary to calculate the conditions for indicator reactions in coupled enzymic assays. When malate dehydrogenase is used as an indicator enzyme for the assay of aspartate aminotransferase activity, its Km in relation to oxaloacetate is needed. Km (oxaloacetate) of commercially available mitochondrial malate dehydrogenase from pig heart was determined as Km equals 1.65 x 10(-5) mol/1 using the measurement conditions for aspartate aminotransferase according to the preliminary recommendations of the IFCC.

Animals↗
PubMed1975

[New values for the molar extinction coefficients of NADH and NADPH for the use in routine laboratories (author's transl)].

Extensive re-investigations with regard to the molar extinction coefficients of NADH and NADPH proved that in future, calculations in routine work can be performed with the following much more accurate epsilon-values: 6.15 x 10(3) 1 x mol-1 x cm-1 at Hg 334 nm (NADH and NADPH), 6.3 X 10(3) 1 X mol-1 x cm-1 at 340 nm (NADH and NADPH), 3.4 X 10(3) 1 X mol-1 X Cm-1 (NADH) and 3.5 x 10(3) 1 x mol-1 x cm-1 (NADPH) at Hg 365 nm, respectively. The safest measurement is performed at Hg 334 nm, because here epsilon is identical for both coenzymes and deviations of the epsilon-value caused by temperature, pH and ionic strength are less than 0.5%.

Colorimetry↗
PubMed1967

Purification and properties of crystalline 3-hydroxybutyrate dehydrogenase from Rhodopseudomonas spheroides.

1. The purification and crystallization of 3-hydroxybutyrate dehydrogenase from extracts of Rhodopseudomonas spheroides is described. 2. The molecular weight was calculated to be 85000 by sedimentation equilibrium. 3. Although the enzyme is stable at 0-4 degrees , dilute solutions are rapidly inactivated at 37 degrees ; NADH(2) or Ca(2+) ions prevent this inactivation. 4. The enzyme is extremely sensitive to mercurials, but can be protected by NADH(2) or Ca(2+) ions. 5. From studies on p-hydroxymercuribenzoate binding it is estimated that the enzyme contains 5-6 moles of rapidly reacting thiol groups/mole. 6. d-Lactate and dl-2-hydroxybutyrate are competitive inhibitors of d-3-hydroxybutyrate oxidation. 7. The properties of the crystalline enzyme are compared with those of 3-hydroxybutyrate dehydrogenase preparations from other sources.

Antimetabolites↗