Practical Use of Enzymatic Properties of Microbial Cultures
In laboratory practice specific biochemical properties of bacterial cultures are used for their precise identification. In most cases carbohydrate fermentation, protease and oxidoreductase activity of microbes are evaluated.
Carbohydrate fermentation is estimated on panels and plates containing dozens of various chemical substrates for fermentation like in Hiss’ media rows. Accumulation of acid products of fermentation is indicated by dye color change, and gas appearance can be revealed within the special tube (float) or by automated gas analyzer.
Proteolytic activity can be visually assessed in special protein-containing media (e.g., gelatin hydrolysis) or according to monitoring the end-products of protein chemical decay. In the latter case the indicator stripes with the necessary reagent (lachmus for ammonia, oxalate for indole detection, and lead acetate for hydrogen sulphide determination) are placed into the wells or near the outlet of the test tubes with substrate meat peptone broth (MPB). Being cultivated, the bacteria hydrolyze the peptones of broth releasing the end products of protein depolymerization (ammonia, indole or hydrogen sulphide).
Also various synthetic labelled substrates are used either for assessment of carbohydrate hydrolysis or proteolytic activity. After the incubation of substrates with microbial cultures the results are registered by colorimetric or fluorimetric detection.
Catalase activity is usually assessed by a simple test, where the loop of microbial culture is added into the drop of hydrogen peroxide. Bubbles of gas indicate the hydrogen peroxide conversion into O2 and H2O.
Urease microbial activity is observed in urea test hydrolysis. The accumulation of ammonia that released after urea decay eventually rises medium pH, and the change of color of indicator dye (e.g., phenol red) is observed.