Bioremed HW2 Bioremediation - HW2: Organic Chemistry/Microbiology 1. Name the following compounds. What general structurally-related factors might contribute to the fate and transport, biodegradation/bioavailability of these compounds? (a) 2,4,6-Trinitrotoluene (TNT) the compound may be biodegraded as either a source of cell carbon or nitrogen, in addition to energy source; benzene ring structure tends to be stable. Substituents on benzene ring could cause steric hindrane for interaction with bacterial enzymes. (b) n-Octane. A saturated alkane. Hydrocarbon, naturally found so fairly biodegradable. However, large so will diffuse poorly and may be hard to uptake by bacteria. (c) Anthracene. A polyaromatic hydrocarbon, therefore somewhat difficult to biodegrade. Very hydrophobic so will sorb to solids, low soluble concentration available for bacterial uptake and less migration in groundwater. (d) Tetrachloroethylene, perchloroethylene, PCE. The carbon-chlorine bonds are very stable. Cl substituents tend to make a compound more recalcitrant to biodegradation. The fully substituted ethene cannot be degraded aerobically, likely due to steric hindrane of the chlorines with the oxygenase enzyme active sites. 2. For the compounds listed below, describe which would be most likely to be critical hazards via inhalation, injestion from groundwater flow to drinking water wells, and dermal contact with contaminated soils. Show any information you used to arrive at these conclusions. Compounds: benzo(a)pyrene, pentachlorophenol, phenol, vinyl choride at 20°C for PCP, phenol, VC; 25°C B(a)P Cmpd Mol Wt Boil Pt, C Vapor Pr, Solubility Koc, mL/g Toxicity mm Hg mg/L Info b(a)pyrene 252 5.6E-9 3.8E-3 5,500,000 oral CPF PCP 266 309 1.1E-4 14 53,000 oral RfD/CPF phenol 94 0.02 82000 14 oral RfD VC 62.5 -13.37 2660 4270 57 inhal CPF oral CPF Based on the properties listed in the above table: inhalation hazard: The most volatile of the chemicals is vinyl chloride, which will be a gas under typical environmental conditions (temp and pressure) due to its boiling point, and has by far the highest vapor pressure of the four compounds. Also, of the four compounds, it is the only with a carcinogenic or non-carcinogenic toxicity from inhalation. injestion from drinking water: Since the compound must travel through the aquifer to the drinking water wells, the compound that is most soluble and has the least tendency to volatilize away or sorb to solids would pose the greatest hazard. Phenol clearly has the greatest solubility and low vapor pressure and Koc, and would probably pose the greatest risk. VC also has high solubility, but would likely volatilize out of the water when pumped to the surface. (Also, compounds with oral carcinogenic or non-carcinogenic toxicity values are likely hazards. All of these compounds have oral toxicity values - or pending - so all must be considered potential hazards.) dermal contact with soils: Those compounds with high Koc values will partition to the organics in soil. Benzo(a)pyrene is likely to accumulate on the soil, due to its high Koc (and also a low vapor pressure and solubility would minimize loses from the soil by Bioremed HW2 volatilization and desorption into water). PCP also has a high Koc value, and could also pose a dermal exposure risk. (Once in the body, the adsorbed and oral injested compounds react somewhat similarly, so that oral toxicity information is indicative of dermal toxicity. All compounds have oral toxicity information, and the BaP CPF of 7.3 /mg/kg-d is greater than the PCP CPF of 0.12 /mg/kg-d, with the higher slope factor indicating greater toxicity. Therefore, BaP is a greater hazard than PCP based on toxicity, too.) 3. List the “metabolic descriptors” for the microorganisms listed below, and the energy source, electron donor, and biosynthetic carbon source for each which supports your classification. aerobic BTEX-degrading bacteria = Chemoorganotroph and Heterotroph energy source = carbon compound = “chemo” electron donor = carbon compound = “organo” biosynthetic carbon source = BTEX = “hetero” algae = Photolithotroph or Photoautotroph energy source = light = “photo” electron donor = CO2 (inorganic carbon) = “litho” biosynthetic carbon source = CO2 = “auto” Nitrosomonas = Chemolithotroph energy source = inorganic chemical = “chemo” electron donor = inorganic (NH4) = “litho” biosynthetic carbon source = CO2 = “auto” 4. The initial biodegradation rate of 5 mg/L phenol is 1.0 mg phenol/mg VSS-d at 20°C by an aerobic Pseudomonas bacteria, and the half-saturation coefficient is 0.1 mg/L. (a) Assuming Michaelis-Menten kinetics, what would be the initial biodegradation rates of 0.2 mg/L, 2 mg/L, and 10 mg/L phenol? (b) What would you predict the change in biodegradation rates of phenol to be at 15°C and 40°C? (this can be a qualitative response, but justify your answer) (a) M-M Equation: dS = K * S * X dt Ks + S We are given dS/dt * (1/X) , Ks, and S, and can use to solve for K: 1.0 = __K * 5__ => K = 1.02 mg/mg-d (0.1 + 5) Then use K and Ks for each S to find the biodegradation rates in mg/mg-d: S = 0.2 mg/L, Biodegrad Rate = 0.68 g/g-d S = 2 mg/L, Biodegrad Rate = 0.97 g/g-d S = 10 mg/L, Biodegrad Rate = 1.01 g/g-d (b) At 15°C, the temperature is still in a typical survival range for mesophilic bacteria. The lower temperature would result in slower enzyme reactions at 15°C compared to 20°C, and therefore slower biodegradation rates. Since a typical rule of thumb is that rates increase about 2 times for every 10°C increase in temperature, the biodegradation rate at 15°C is probably 0.5 to 0.9 times the 20°C rate. At 40°C, the temperature may be above the optimum growth rate temperature of the bacteria, and the biodegradation rates may be slower. For example, facultative psychrophiles which are typically isolated from soil and water in temperature climates grow best between 25 to 30°C, with a maximum of about 35°C, above which growth rates Bioremed HW2 decrease. Pseudomonads are typically mesophiles, and those typically isolated from the environment would probably be above the optimum growth temperature at 40°C. Therefore, it is not possible to predict the rate response at the higher temperature, and experimental data would be needed. 5. Given the following graphs from experimental data, how would you describe or model the biokinetics? (a) Zero order over the substrate range where curve is horizontal, and substrate toxicity in the S concentration range where the rate is declining (b) Michaelis-Menton, zero order at “high” substrate concentrations where the curve is linear and first order rates at substrate concentrations where the “rate” or slope of the curve declines (c) Competitive inhibition? or inhibition of A on B. “A” degradation curve appears to follow Michaelis-Menten kinetics. When the concentration of A is very low, the degradation rate of “B” increases. This could occur if “B” also follows M-M model when present alone, but has a much higher Ks than “A” and therefore its degradation is greatly inhibited by competition for the degrading enzyme by compound “A”.