Aqua, a commercial product manufactured by BiOWiSH® Technologies, was utilized in this research to study its effectiveness to biodegrade methyl tert-butyl ether (MTBE) and tert-butyl alcohol (TBA). Microcosms containing varying concentrations of MTBE and TBA as well as a growth media and mineral salt solution were examined. Analytical instrumentation used in this study included the use of a gas chromatograph-mass spectrometer (GC/MS) to determine concentrations of MTBE and TBA and a spectrophotometer to extrapolate approximate active biomass concentrations in each experiment. Four different environmental conditions were tested for both MTBE and TBA. The environmental conditions tested for each contaminant included: biodegradation under aerobic conditions, biodegradation under anaerobic conditions, biodegradation under denitrifying conditions, and biodegradation under aerobic conditions with glucose present. This study concluded that there is potential for degradation of MTBE and TBA using Aqua under the conditions tested. Maximum MTBE biodegradation was observed under aerobic conditions which yielded a first order rate constant of 0.019/hour and a 99.8 percent decrease in MTBE over 14 days. Maximum TBA biodegradation was observed under aerobic conditions with glucose present which yielded a first rate order constant of 0.009/hour and a 95.03 percent decrease in TBA concentrations over 14 days. It is presumed that under both conditions a monooxygenase enzymatic reaction involving Cytochrome P-450 aids in breaking down both MTBE and TBA. However, the results presented are indicative of biodegradation under lab conditions with little to no interference. Further research is needed to determine the effectiveness of Aqua utilizing groundwater or soil samples from MTBE or TBA contaminated sites in order to truly analyze Aqua’s potential to be used as a bioaugmentation product in real world applications.

Abstract Investigation of a Commercial Product (BiOWiSH­­TM) for Nitrogen Management Eva Lee BiOWiSH–Aqua, which has the capability of treating nitrogen from wastewater through bioaugmentation, is a commercial product consisting of a blend of microorganisms developed by BiOWiSH Technologies. A study of the treatment of nitrogen compounds (i.e. , , and ) using Biowish–Aqua was conducted using small scale experiments (flask experiments) and large scale experiments (column reactor experiments). In this work, column reactors were created to test Biowish–Aqua’s nitrogen treating capabilities by providing enough depth to simulate the dissolved oxygen gradient that can be observed in a pond. The results show that the optimal growth conditions for both ammonia assimilating and denitrifying bacteria are an anoxic environment with a carbon-to-nitrogen ratio of 2:1. Under this optimal growth environment, Biowish–Aqua was able to assimilate ammonia with a zero order k value of 3.06 ppm/day. Also, under the same conditions, Biowish–Aqua was able to eliminate nitrate ( ) and nitrite ( ) at a rate of 9.58 ppm/day and 5.64 ppm/day respectively. The experiments also suggested that with a C:N ratio of 2:1, Biowish–Aqua did not have an effect in slowing the hydrolysis of urea. Overall, this research suggests that the application of Biowish–Aqua is a feasible nitrogen removing strategy for wastewater with initial presence of ammonia and nitrate between 10 to 20 ppm. Keywords: Ammonia assimilation, Bioaugmentation, BiOWiSH, Denitrification

Residential septic systems provide reliable wastewater treatment for over 26 million homes and facilities in the United States, and many more worldwide. When properly maintained, these systems are reliable, low-cost, and long-term treatments for residential wastewater. When neglected, septic systems can fail and lead to health concerns and ecological harm to soil and groundwater contamination through the improperly treated wastewater effluent. This study tested the effect of the bio-augmentation product BiOWiSH® Septic Rescue of BiOWiSH® Technologies International, Inc. (hereafter referred to as BiOWiSH) on the biological treatment of residential septic tanks. BiOWiSH is meant to act as a bio-augmentation product through the addition of a proprietary blend of Bacillus and Lactic Acid producing bacteria. These microbes act as a biocatalyst to enhance and encourage a range of hydrolytic, oxidative, and reductive biochemical reaction and promote digestion of bio solids and ammonification within the septic tanks. To test the effect of BiOWiSH on the treatment of residential septic tanks, four 32-gallon tanks were constructed and filled with water and primary sludge from the primary clarifier at the San Luis Obispo Water Resource Recovery Facility. Two tanks were dosed with the recommended amount of BiOWiSH; one tank had no additive biological treatment and served as the control; one tank was dosed with RID-X® Septic Maintenance, a leading competitive product (hereafter referred to as RID-X). Each tank functioned as a plug-flow reactor. Primary sludge and tap water was added daily and effluent was sampled on a daily or weekly basis, based on the parameters being tested. Effluent water samples were tested for removal of ammonia, nitrates, total suspended solids, and biological oxygen demand. Temperature and pH were also recorded. v These analyses indicated no significant advantage from the addition of BiOWiSH in the reduction of ammonia, total suspended solids, or biological oxygen demand over the control tank or the tank dosed with the RID-X competitive product. Nitrates (in the form of nitrate and nitrite) did not form in any of the tanks. Future studies are needed to validate these results. Additional studies should include an analysis of experimental time frames, sampling frequency, and testing additional products designed to rescue failed or failing septic systems. BiOWiSH should also be tested further in its potential ability to enhance the biological treatment of septic tank effluent once the wastewater has entered aerobic leach fields.