Age-dependent excretion of alanine aminopeptidase, alkaline phosphatase, gamma-glutamyltransferase and N-acetyl-beta-D-glucosaminidase in human urine.

Urinary excretion of alanine aminopeptidase, alkaline phosphatase, gamma-glutamyltransferase and N-acetyl-beta-D-glucosaminidase was determined in gel-filtered samples of morning random urine specimens of 442 subjects of various ages (5 days to 58 years). Enzyme excretion related to urinary creatinine (enzyme/creatinine ratio; U/mmol creatinine) significantly decreased with increasing age. Sex-related differences of some enzyme excretions were found in age groups over 6 years. From these investigations, we calculated upper reference intervals (97.5 percentiles) for 5 age-dependent groups of children and adolescents and for one group of adults.

Quality control material for activity determinations of urinary enzymes.

We examined the stability of N-acetyl-beta-D-glucosaminidase (EC 3.2.1.30), alanine aminopeptidase (EC 3.4.11.2), alkaline phosphatase (EC 3.1.3.1), gamma-glutamyltransferase (EC 2.3.2.2), and lysozyme (EC 3.2.1.17) in urine prepared by gel filtration and supplemented with albumin, or ethylene glycol, or ethylene glycol plus albumin during storage at -20 degrees C for a period of 12 months. The stability was assessed by linear regression analysis of monthly values versus time. All enzymes except for gamma-glutamyltransferase could be considered stable for about one year in all three control materials provided that maximum change of 10% of the starting enzyme activity is accepted as tolerable. If ethylene glycol is used as stabilizer, its suitability must be tested and its inhibitory effect on enzyme activities must be taken into account in intermethod comparisons, because in some methods, it may be removed in a pretreatment step.

Long-term stability of enzymes in human serum stored in liquid nitrogen.

We analyzed the stability of the enzymes alpha-amylase (EC 3.2.1.1), alkaline phosphatase (EC 3.1.3.1), alanine aminotransferase (EC 2.6.1.2), aspartate aminotransferase (EC 2.6.1.1), creatine kinase (EC 2.7.3.2), glutamate dehydrogenase (EC 1.4.1.3), gamma-glutamyltransferase (EC 2.3.2.2) and lactate dehydrogenase (EC 1.1.1.27) of a human serum pool during storage in liquid nitrogen for a period of 10 months. Except amylase and creatine kinase, all enzymes were stable. Amylase increased in activity, creatine kinase activity decreased. Therefore, human serum stored at -196 degrees C can be used as satisfactory substitute for lyophilized enzyme control serum in internal quality control and stable enzyme material for optimization of methods.

Suitability of commercial control sera for the quality control of activity determination of alkaline phosphatase.

The suitability of thirteen commercially available control sera for measuring alkaline phosphatase (EC 3.1.3.1; orthophosphoric acid monoester phosphohydrolase, ALP) activity in human serum was tested. Apart from differences in ALP activity observed in some reconstituted commercial sera, the behaviour of control materials towards experimental variables such as the nature and concentration of the substrate, pH and type of buffer (or PO4-acceptor) together with the composition of the isoenzymes present in human serum highlights the problems and difficulties if commercial materials are to be used as control sera. The half-saturation constants in control sera were in all cases smaller than those of ALP isoenzymes from bone and liver. The shape of substrate activity curves and the pH optimum in most of control sera differed from that of human serum. The discrepant kinetic data of control materials and human serum may mask or suggest changes relevant to commercial quality control serum but not to samples of human serum.

Comparative determinations of aminotransferase activities in serum with so-called "optimised" methods.

Comparative determinations of aspartate aminotransferase and alanine aminotransferase activities were made with so-called "optimised" methods introduced in the G.D.R., G.F.R. and Scandinavia. By means of the paired t-test significant differences could be established. These differences are partly due to different reaction conditions. For practical clinical aspects these differences should be of little relevance. In comparison with above-mentioned activites determined at 37 degrees C, aspartate aminotransferase activities measured with the IFCC reference method are lower by about 30 percent.

Suitability of commercial enzyme control sera for the quality control of activity determinations of L-aspartate aminotransferase and L-alanine aminotransferase in human serum.

Details of a systematic approach to suitability testing of commercial control sera are given for substrate optimized L-aspartate aminotransferase and L-alanine aminotransferase methods at 37 degrees C. Their acceptability for control purposes of standardized methods depends on: (1) the range of control values in relation to borderline values, (2) stability, (3) aspect, clarity, (4) NADH consumption in preincubation time, (5) blank activities, (6) kinetic data as half saturation constants and saturation curves, (7) influence of effectors, (8) isoenzyme pattern. These evaluation criteria are proposed for suitability testing. The term "representativeness" should be introduced as a special criterion for main characteristics of control materials. The authors want to point out the close connection with standardization of methods.