Acid and alkaline phosphatase activities were determined
using a modification of the method of Juma and Tabatabai
(1988). Culture supernatant (1 ml) from the previous experi-
ment was incubated with 1 ml of 25 mM r-nitrophenyl
phosphate and 4 ml modified universal buffer (pH 6.5 or pH
11 for the assay of acid or alkaline phosphatase, respectively)
at 37 8C in the dark. The reaction was terminated after 1 h by
adding 1 ml of 0.5 M CaCl 2 and 4 ml of 0.5 M NaOH. The assay
mixtures were filtered through Whatman No. 1 filter paper
(Whatman, Maidstone, England) and the intensity of the
yellow color was measured at 410 nm. There were eight
independent replicate flaks for each isolate.