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E}3.� Du Pont HLR 399-07 intru6orctton/Purpose Ground Yater Siorumedistion Serviced, a Du Pont service organization, hos accepted the responsibility for cleaning up the subsurface coil and crater contaminated with gasoline at a Kayo station in Lodi, California. As part of the microbiological support for this effort this project was initiated to determine the appropriate nutrient combinations and concentrations for optimal in situ biodegradation-of the contaminated soil and water. Materie4s/Ptethoda Samples were collected using aseptic techniques and sent for microbic- logical testing within 24 hr. The samples were stored at 40C before and aftcr testing. Sample descriptions are listed in Table 1. The water (M-18780) was divided into two lots and the one lot was sterilized by autoclosving for 30 min. The autoclaved water was subsequently used for soil slurry studies. The other lot of water was kept refrigerated until tested. Nutrient Optimization Study Using a 14C-labelled Substrate A radiolabelled substrate (m--ring- 14 -- C]Xylene from Amersham Laboratories; specific activity of 853 uCi/rag)) was used to represent moderately recalcitrant materials found in gasoline. Sixty milliliter serum bottles were arranged according to Table 2 (1-20). Each treatment consisted s of nine replicates such that triplicate bottles could be tested for three time periods (4, 24, end 72 hr). Nutrients were prepared in 7% stock solutions (the sodium phosphate solution was a 50:50 mixture of the dibasic and monobasic chemicals and ammonium chloride served as the nitrogen source) and diluted to the final concentrations shown in Table 2. A 25% soil slurry i sea prepared with groundwater and 8-1 12 (M-18779) or NW-19 BA (M-16773) soil. Following nutrient addition to the bottles, 10 mL of the appropriate soil slurry was added to the bottles. Controls consisted of formaldehyde treated (10 ppm) and untreated coil slurries. Absy4ute ethanol and unlabeled xylene were used to remove the minute quantity of C-labeled xylene from the distillation tube (supplied by the manufacturer) and reduce the specific activity of the compound. A final concentration of 1 ppm total xylene was achieved by. diapensing the appropriate amount of diluted material into each test bottle. The battles were closed with a rubber stopper containing a r plastic bucket and filter paper (the stopper, bucket, and filter were ethylene oxide sterilized). Triplicate bottles were tested at three time periodet 4, 24, and 72. Phonethylamine (PEA) rias injected (0.2 mL )into the filter-paper,--follow94-by-a-small-quentity-of-sulfuric-acid-into-the - oalurry, to drive the carbon dioride from the liquid and into the.PEA maturated filter. The bottles were gently shaken for 4 hr to-facilitate the • •• 4 y