A new continuous automated assay for the determination of intestinal lactase.
BACKGROUND
A reliable, sensitive and practicable method for the measurement of intestinal lactase activity is needed.


METHODS
The assay is based on the continuous measurement of released glucose by a coupled hexokinase/glucose-6-phosphate dehydrogenase (HK/G6PDH) reagent.


RESULTS
The procedure was first optimized for lactase from rat intestinal mucosa. The optimum pH is 6.5 and apparent Km was 17 mmol/l for lactose. The procedure was also adapted on a Cobas Mira automated analyzer; progress curves of the reaction rate can be continuously monitored. The automated assay correlated strongly with the conventional method of Dahlqvist (r(2) = 0.996). The described method has also been applied to human intestinal mucosa biopsies after determination of the catalytic properties of human enzyme (optimum pH 6.0 and apparent Km 34 mmol/l).


CONCLUSIONS
The HK/G6PDH method is reliable, rapid, sensitive and easy to perform both manually as well as in the automated version. It is optimized for human and rat intestinal lactase.
Sall
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Laboratoire de Biochimie Appliquée (Faculté de Pharmacie), Université Louis-Pasteur de Strasbourg, Illkirch, France. isall@evc.net
Férard
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D000328:Adult; D000818:Animals; D001681:Biological Assay / Q000379:methods*; D019411:Clinical Laboratory Techniques / Q000295:instrumentation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000201:enzymology*; D007421:Intestine, Small / Q000201:enzymology*; D043322:Lactase; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013053:Spectrophotometry / Q000295:instrumentation / Q000379:methods; D013393:Sucrase / Q000032:analysis / Q000378:metabolism; D000520:alpha-Glucosidases / Q000032:analysis / Q000378:metabolism; D001616:beta-Galactosidase / Q000032:analysis* / Q000378:metabolism
D043322:Lactase; D000520:alpha-Glucosidases; D001616:beta-Galactosidase; D013393:Sucrase