PEDRO J. ALVAREZ: The most common classes of groundwater contaminants for which natural attenuation is considered are chlorinated solvents and hydrocarbons. So in this lecture, we will consider the mechanisms associated with their biodegradation to enhance our understanding of what environmental conditions and factors enhance or hinder their biodegradation. In lectures 2 and 3, we learned that highly chlorinated compounds, such as perchloroethylene and TCE can be reductively dechlorinated under anaerobic conditions. And this process can be through fortuitous cometabolism, meaning that it is not coupled to energy harvesting or cell growth. Or it could be metabolic, meaning that some bacteria can use dechlorinated compounds as terminal electron acceptors in a respiratory process that yields energy to support their growth. And this is known as halorespiration. And in this case, microbial growth results in more bacteria capable of attacking these compounds, which increases degradation rates. And the rates of halorespiration are consequently orders of magnitude faster than those for cometabolic degradation. Let us consider in greater chemical detail, the dechlorination mechanisms, using here perchloroethylene as an example. In the Lewis chemical structure, each covalent bond line represents two shared electrons, shown here in red. The dechlorination process starts when one electron is donated by a microbial enzyme or electron carrier. Also, the electron donor could also be surface-associated ferrous iron, or even zerovalent iron when the reaction is abiotic. The chlorine atom accepts this electron and leaves the molecule as a chloride ion. And since the first step involves the transfer of one electron, because elements such as iron or cobalt that transfer one electron at a time are involved, the dechlorination propensity of a contaminant depends often on the one electron reduction potential. Now, this dechlorinated intermediate, which is sort of an organic free radical, very short lived, is reduced farther. Specifically, another electron is transferred in a similar manner. And then a water-derived proton is incorporated, resulting in the net replacement of the chlorine atom by a hydrogen atom. And overall, this reaction is known as hydrogenolysis. It requires two electrons to knock off each chlorine atom and replace it with a hydrogen atom, and in this case, trichloroethylene or TCE, is formed. And this process can be repeated so that sequentially more chlorine atoms are replaced one at a time by hydrogen atoms. Now one challenge associated with sequential hydrogenolysis is that cometabolic dechlorinators can produce vinyl chloride, which is more toxic than the original trichloroethylene. In contrast, the halorespiring organisms can perform dechlorination all the way to ethene, which results in complete detoxification. Now the thermodynamic feasibility of aerobic biodegradation of the dechlorinated byproducts increases as the chlorination progresses, as indicated by the more positive Delta E values. So it is quite possible that these compounds can be degraded by sequential anaerobic-aerobic processes. For example, as they dechlorinate in an anaerobic zone, and then the byproducts migrate into the aerobic zone. Now the key points are that reductive dechlorination requires appropriate electron donors to first induce anaerobic conditions and stimulate dechlorination or, that is, the removal of chlorine atoms, which will decrease the toxicity with the noted exception of the formation of vinyl chloride and enhance the solubility of these compounds. Some strains can respire these chlorinated solvents as electron acceptors. And this results in faster and greater potential for complete detoxification. These halorespiring organisms include those of the genus Dehalobacter restrictus, Dehalococcoides ethenogenes, and Desulfomonile tiedje, among many. It's interesting to point out that these bacteria are pretty widespread among solvent-contaminated sites. Although it is not always the case that they will be present. And sometimes they're added in bioaugmentation schemes to stimulate dechlorination. Now in contrast to chlorinated solvents, hydrocarbons are very reduced chemically. So their oxidation is very favorable thermodynamically. And that's why we use them as fuel in combustion engines. Now hydroxylation, which refers to the addition of hydroxyl groups, is often the first step. This is mediated by oxygenase enzymes. This transformation increases the solubility of the compound and makes it more susceptible to subsequent metabolism. Now this reaction requires molecular oxygen, and often it is the diffusion and replenishment of molecular oxygen what limits the rate of aerobic biodegradation. One condition for the ring fission for aromatic compounds is that they have to be di-hydroxylated. That is, they need two hydroxyl groups inserted by these oxygenase enzymes before the ring can be opened up. Then once the ring is opened up, the byproducts can be easily funneled into central metabolic pathways, such as Krebs cycle, where complete mineralization to carbon dioxide occurs. Anaerobic degradation of BTEX compounds is very important as a natural attenuation mechanism, even if it proceeds at much slower rates due to the weaker electron acceptors that are utilized in this process. Benzene, which is the most toxic of the BTEX compounds, is unfortunately relatively recalcitrant under anaerobic conditions, where it degrades very slowly, if at all, usually after the other alkylbenzenes are degraded. And benzoyl-CoA is a common intermediate in these reactions. It's reduced prior to ring fission by hydrolysis, and the oxygen that evolves in that CO2 is usually originating from water. A survey of plume dimensions that was conducted by GSI-- they considered hundreds of sites-- they showed that BTEX plumes are relatively small, compared to chlorinated plumes. And this reflects that BTEX compounds are relatively easy to degrade due perhaps to their natural pyrolytic origin, compared to these synthetic chlorinated solvents that are relatively recalcitrant in nature. So the key points are that chlorinated solvents like TCE and PCE degrade relatively fast under anaerobic conditions, provided that suitable electron donors, such as hydrogen and acetate, are present. Some bacteria, known as "dehalorespirers," can obtain metabolic energy from this process and grow, thus increasing the dechlorination rates and dechlorinate TCE all the way to ethene, resulting in complete detoxification. And hydrocarbons degrade faster under aerobic conditions, and their biodegradation rate are often limited by oxygen replenishment rates. Anaerobic degradation of hydrocarbons, although slower, is also an important mechanism for monitored natural attenuation.