Activity and properties of extracted soil enzymes

Active enzymes have been extracted from soils by shaking suspensions for periods ranging from 5 min to 24 h, and at temperatures, sometimes ambient or below, but more usually at 37-40°C (Table 3). Generally conditions are chosen to avoid extensive damage to live cells and to favour extraction of abiontic enzymes. Mostly, soil extracts have been assayed for hydrolases, some have been fractionated and the organic matter associated with enzymic activity has been partly characterized. Kinetic constants, stabilities. 

IM phosphate, pH 7.5, 30min, 30°C 0.2M NaOH, 30min, room temp. 

0 and p -Diphenol oxidases have been extracted aseptically from fresh soils and partially purified using a salmine precipitation technique, followed by Sephadex G-25 gel chromatography which removed some associated humic compounds. The enzyme preparation was heterogeneous acid 

The partially-purified extract oxidised a range of phenolic substrates, and also contained proteinases and amino acid decarboxylases. Preincubation of a toluene-treated soil enzyme preparation for 12h at 37°C did not affect diphenol oxidase activities, ie. the oxidases appeared to be resistant to attack by the coextracted soil proteinases. Addition of hyaluronidase before preincubation also was without effect. Preincubation with the microbial proteinase, Pronase for I8h at 37°C decreased diphenol oxidase activities by 307o, and by 100% when both Pronase and hyaluronidase were added. The results suggested that the polysaccharides associated with the extracted soil oxidases protected the enzymes from proteolysis and may play a role in stabilizing exocellular enzymes in soils. 

As with soil, diphenol oxidases extracted from forest litter also can be fractionated to yield components essentially free of co-extracted humic compounds. A comparison of humic-free laccases (p-diphenol oxidases) purified from soil and litter extracts with those from Polyporus versicolor indicated that the former preparations were strongly electronegative and were only weakly adsorbed to humic colloids. By contrast other enzymically-active fractions from soil extracts were associated with humic compounds in complexes which were not separable by chromatography or electrophoresis.  

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