Co-Encapsulation of Slow Release Substrates and Microbial Cultures in Alginate and Gellan Gum Beads to Promote the Co-metabolic Transformation of 1,4-Dioxane and Chlorinate Aliphatic Hydrocarbons
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Methods were developed for the co-encapsulation of slow release compounds (SRC) with viable microbial cells in alginate and gellan gum hydrogel beads, for the in-situ aerobic cometabolic treatment of groundwater contaminated with mixtures of 1,4-dioxane and chlorinated aliphatic hydrocarbons, which will be referred to as contaminants of concern (CoC). The bacteria used was Rhodococcus Rhodochrous ATCC 21198 (ATCC 21198) that is capable of cometabolically transforming mixtures of CoCs when grown on isobutane as the primary substrate. In addition to isobutane inducing cometabolic transformation, research conducted concurrently to this project has shown that the growth of ATCC 21198 on non-gaseous substrates, like the branched alcohol 2-butanol, may also stimulate the short chain alkane monooxygenase (SCAM) enzyme of ATCC 21198 that is responsible for the cometabolic transformation of CoCs. Due to this, investigation was conducted into two model slow release compounds, tetrabutylorthosilicate (TBOS) and tetra-s-butylorthosilicate (T2BOS), which hydrolyze slowly at ester bonds to produce 1- and 2- butanol, respectively, and exist in pure phase as light non-aqueous phase liquids (LNAPL).
LNAPL SRCs (TBOS and T2BOS) were successfully encapsulated in both alginate and gellan gum matrices at high mass loadings, >10% (w/w), and co-encapsulated ATCC 21198 was shown to be able to consume SRC products (1- and 2-butanol) prior to diffusion from the beads. The energy gained from the utilization of 1- and 2-butanol by encapsulated cultures was observed to have increased the survivability, overall activity, and contaminant transformation rate and capacity of initially augmented biomass. For example, in batch systems it was observed that cells co-encapsulated with SRSs were able to maintain cometabolic transformation potential for over 70 days, whereas, similar cellular biomass suspended in media lost the majority of CoC transformation potential after the first 12 days. Also, co-encapsulated cultures were able to transform 2-4 times more contaminants than suspended cultures over the 70 day period, and transformation within co-encapsulated systems is continuing to be observed. The results of this study provide strong evidence that the developed co-encapsulation technology may provide a useable solution to many issues encountered during current in-situ bioremediation treatment schemes.