Preadapted Inocula for Limiting the Risk of Errors in Biodegradability Tests

Reducing the time for biodegradability tests to 28 days poses a problem when the inoculum contains few biodegraders, as a biodegradable xenobiotic must give a positive result within this time. The influence of initial concentration (X0, number of cells liter−1) on the lag time (hours) ofpara-nitrophenol biodegradability tests was examined using different concentrations of adaptedPseudomonas putidawithpara-nitrophenol as the sole carbon and energy source. Lag time decreased as bacterial density increased according to the expressiony= 63.5 − 5.7(log10X0). The addition of river water to theP. putidaconcentrations shortened the lag time–bacterial density relationship and lag time filled the expressiony= 52.4 − 5.1(log10X0). The addition of river water also increased the rate ofpara-nitrophenol biodegradation from 1.29 mgC (liters × hr)−1to 2.11 mgC (liters × hr)−1. An examination of the effect of the initial adaptedP. putidadensity, expressed as total cell, cultivable bacteria, or biodegraders, was also made on thepara-nitrophenol biodegradability test outcome. The model-related cell density and the probability of test response give very similarkconstants (kT= 0.56 × 10−3liter total cells−1;kV= 0.11 × 10−3liter CFU−1,kMPN= 0.16 × 10−3liter MPN−1). Comparisons with nonadapted natural mixed culture (activated sludge, river water) indicate that the biodegradability test responses were the same as with adapted cells when the nonadapted cell concentrations were at least 5 × 1010total cells liter−1. As this high cell concentration led to carbon contamination, adapting mixed inocula before the test to increase the number of biodegraders appears to be the best solution. Before biodegradability tests, cell density can be adjusted using techniques which are not specific to biodegraders, and only 105total adapted cells liter−1are needed for a 99.9% chance of a positive response inpara-nitrophenol biodegradability tests.