Five grams of fresh soil were suspended in 5 mL of 250 mM tris-HCl buffer (pH 7.4) containing 3.47 mM glucose. The enzymatic reaction started when 1 mL of 2.5% (w/v) triphenyltetrazolium chloride solution was added to the soil suspension. The reaction took 12 h in the dark at 37 ºC, mixing the suspension occasionally. The reaction was stopped by adding methanol. The methanol suspension was passed through cotton plug, filled up to 100 mL with methanol, and the formazan was measured colorimetrically at 480 nm. The control was prepared by mixing the soil sample with methanol before adding the tris-HCl buffer and the tetrazolium solution. The coloration for the control was a negligible value of less than 3% of the sample value, suggesting the negligible abiotic formation of the formazan