Robert J. Larson

Biodegradation of [S,S], [R,R] and mixed stereoisomers of Ethylene Diamine Disuccinic Acid (EDDS), a transition metal chelator

An in-depth biodegradation test program was executed on the hexadentate ligand Ethylene Diamine Di Succinate (EDDS). The EDDS structure contains two chiral carbon atoms, and has three stereoisomers ([R,R], [R,S]/[S,R], [S,S]). Our research has focused on the isomer mixture (i.e. 25%[S,S]; 25%[R,R]; 50%[S,R]/[R,S], as produced from the reaction of ethylene diamine with maleic anhydride) and on the single [S,S]- and [R,R]-isomers. Biodegradation screening of the 14C-labelled EDDS isomer mixture in a Batch Activated Sludge (BAS) test with various inocula revealed incomplete mineralization, up to ca. 65% after 28 days. N-(2-aminoethyl) aspartic acid (AEAA), probably the d-isomer, was identified as the major portion of the 14C-material remaining in solution. Further testing revealed that the [S,S]-isomer is rapidly and completely mineralized in all test systems. By contrast, [R,R]-EDDS remained undegraded in a Sturm (OECD 301B) test, but was very slowly biotransformed into the recalcitrant metabolite AEAA in a BAS test. The [S,R]/[R,S] form undergoes biotransformation to AEAA in both high and low biomass systems. In a sewage treatment simulation test (OECD 303) the steady state DOC removal of mixture-EDDS in a CAS test was limited to 25-35%, even after extensive pre-acclimation, while the [S,S]-isomer achieved nearly complete removal (96%). This study illustrates the importance stereospecificity may have on the biodegradation and metabolite formation of a chemical. A biodegradation scheme for the different EDDS stereoisomers is proposed.

An improved model for predicting the fate of consumer product chemicals in wastewater treatment plants

Abstract The WW-TREAT model was developed to predict the fate of consumer product chemicals in primary and activated sludge wastewater treatment plants using independently determined values for the distribution coefficient between sludge solids and liquid, Henrys law constant and biodegradation rate constants. The major difference between this model and previous models is that it assumes the total chemical, not just the dissolved chemical, is available for biodegradation and that sorption to sludge solids increases the time that a chemical is available for biodegradation. The model was validated with monitoring data for four chemicals which possess sorption and biodegradation characteristics that span the range typical of the broader class of consumer product chemicals. Sensitivity analysis revealed that the most important parameters were the plant operating parameter, sludge/solids retention time and the chemical specific parameters, distribution coefficient and biodegradation rate constants. The sludge/solids retention time had its greatest effect when it was less than 9 days and the distribution coefficient had its greatest effect when it was between 100 and 3000 l/kg. The model predicted removal in primary and activated sludge wastewater treatment plants within 5%. Thus, the model can be used as a valuable tool for predicting the fate of consumer product chemicals.

Environmental fate and effects of DEEDMAC : a new rapidly biodegradable cationic surfactant for use in fabric softeners

This paper introduces the environmental safety database for a new fabric softening cationic surfactant, the di-(tallow fatty acid) ester of di-2-hydroxyethyl dimethyl ammonium chloride, DEEDMAC >Diethyl Ester Dimethyl Ammonium Chloride). The physiochemical properties of DEEDMAC are similar to those for ditallow dimethyl ammonium chloride (DTDMAC), the major cationic surfactant used in fabric softener formulations world-wide for over thirty years. Importantly, however, DEEDMAC differs structurally from DTDMAC by the inclusion of two weak ester linkages between the ethyl and tallow chains. These ester linkages allow DEEDMAC to be rapidly and completely biodegraded in standard laboratory screening tests and a range of environmental compartments. including raw sewage, activated sludge, anaerobic digestor sludge, sludge amended soil, and river waters. Removal of DEEDMAC during sewage treatment is greater than 99%, as determined by computer model predictions and confirmed by laboratory simulation testing (OECD Continuous Activated Sludge confirmatory test). Using estimated tonnages, per capita waste water flows, incidences of sewage treatment for individual countries, measured removal rates, and validated computer models, maximum river water and soil concentrations of DEEDMAC have been estimated for representative usage scenarios. Based upon these maximum predicted environmental concentrations, acute and chronic toxicity testing offish, invertebrates and algae, predicted aquatic safety factors range from 272 to > 1000. Predicted steady state terrestrial safety factors are > 1000, based on EC50 values to earthworms and plants >50 mg/kg. The environmental safety database developed for DEEDMAC indicates that this cationic surfactant is rapidly and completely biodegraded, will be highly removed during sewage treatment. has an ecotoxicity profile similar to broadly used anionic and nonionic surfactants, and is environmentally safe at intended maximum usage volumes.

Kinetics and practical significance of biodegradation of linear alkylbenzene sulfonate in the environment

This paper reviews the kinetics of biodegradation of linear alkylbenzene sulfonate (LAS) in engineered (wastewater treatment) and natural environment systems, focusing on work conducted in our environmental laboratories over the past 10–15 yr. Biodegradation studies were conducted in laboratory microcosms in which pure-chainlength [14C]-ring-labeled LAS homologs were used to allow complete mineralization to be assessed. In general, biodegradation rates for a series of LAS homologs (C10–C14) were comparable to each other and to values for naturally occurring materials such as sugars and fatty acids. Half-lives for LAS mineralization ranged from 1–2 d in aerobic and anaerobic sewage sludges, river water and sediments, to 1–3 wk in surface and subsurface soils and estuarine environments. The half-life for LAS degradation in different environmental compartments, relative to its residence time in these compartments, makes biodegradation a practically significant removal mechanism in a broad range of aquatic, benthic and terrestrial habitats.

Comparison of biodegradation rates in laboratory screening studies with rates in natural waters

Microbial degradation (biodegradation) of organic chemicals is generally recognized as an important removal mechanism in natural systems. For chemicals which reach the aquatic environment in significant quantities, estimates of biodegradability are key in assessing the overall hazard associated with the use of a particular chemical (Larson 1980). Estimates of biodegradability are often generated in the laboratory via biodegradability screening studies (Larson 1979). In screening studies, compounds are tested as sole carbon and energy sources at relatively high (mg/L) concentrations with a dilute synthetic salts solution as the test medium. Degradation is measured nonspecifically by following the amount of carbon dioxide produced or oxygen consumed during microbial metabolism, and soil or sewage are typically used as the source of degradative microorganisms. These experimental conditions do not accurately simulate natural aquatic environments, where chemical concentrations are low (μg/L) and a variety of nutrient conditions and microbial species exist. As a result, legitimate questions have been raised about the reliability of biodegradability screening tests, and the ability of such tests to predict fate of compounds in the “real world” (Alexander 1981).

A rapid method for determining the toxicity of chemicals to activated sludge

Abstract This paper reports the development of a method to determine the toxicity of chemicals to activated sludge. In the method, activated sludge was exposed to various concentrations of test chemicals and the inhibition of [14C]glucose uptake was measured after 15 min. Data for the decrease in glucose uptake as a function of the log of the test chemical concentration was analyzed by a nonlinear regression model to determine the concentration of chemical inhibiting uptake by 50% (IC50 value). In control experiments, glucose uptake in the absence of test chemicals was rapid, specific and totally dependent on the presence of metabolically-active activated sludge. However, uptake in the presence of inorganic and organic test chemicals showed significant, dose-dependent decreases as the concentration of test material increased. Inhibition of uptake was accurately described by the nonlinear regression model, and calculated IC50 values for two chemicals, mercuric chloride and 3,5-dichlorophenol, agreed well with values reported in the literature to adversely affect wastewater treatment plant operation. Based on the results of our studies, inhibition of glucose uptake proved to be a rapid, accurate and reproducible method of determining the toxicity of chemicals to activated sludge.

Characterization and distribution of esterase activity in activated sludge

The location and activity of esterase enzymes in activated sludge from three municipal wastewater treatment plants were characterized using model substrates and denaturing and non-denaturing polyacrylamide gel electrophoresis (PAGE) of particulate, freeze-thaw (primarily periplasmic enzymes and those associated with outer cell surfaces) and extracellular fractions of activated sludge bacteria. Particulate and freeze-thaw fractions had a similar spectrum of substrate specificity and contained significant levels of protein and esterase activity against model substrates, C2-C18 monoesters of p-nitrophenol and C2-C8 diesters of fluorescein. Esterase activity was highest with substrates that had short alkyl chains (C4) and decreased as the chain lengths increased beyond C8. Extracellular fractions contained very low levels of protein (<0.1 mg/l) and showed no esterase activity against any of the model substrates tested. Multiple bands were observed upon analysis of particulate and freeze-thaw fractions by non-denaturing PAGE in combination with activity staining using various alpha-naphthol ester substrates (C2-C8). Our results indicate that esterase enzymes in activated sludge are fairly diverse from a structural standpoint but exhibit a high level of functional redundancy, with different enzymes catalyzing the same reactions in different sludges. Extracellular esterase activity was totally absent for the substrates we tested and the esterase activity that we observed was closely linked to a particulate floc or cellular material.

Acclimation to and biodegradation of nitrilotriacetate (NTA) at trace concentrations in natural waters

Abstract Acclimation to and biodegradation of nitrilotriacetic acid (NTA), an organic builder used in synthetic laundry detergents as the sodium salt, was studied at trace concentrations (ppb) in several river waters. The river waters tested ranged from those where extensive NTA exposure via detergents had not occurred, to those where NTA exposure had been continuous for several years. In rivers not previously exposed to NTA, acclimation and degradation were observed at the lowest initial concentration tested, 5 μg 1 −1 . Degradation of NTA after acclimation followed apparent first order kinetics, and half lives for NTA removal ranged from 7 to 138 h at initial NTA concentrations of 50 and 5 μg 1 −1 , respectively. Degradation of NTA in water samples where prior NTA exposure had already occurred required no acclimation and was less variable than in unexposed rivers. First order rate constants varied only slightly over a 1000-fold initial concentration range (1–1000 μg 1 −1 ) and NTA half lives ranged from 7 to 17 h. In general, our results indicate that microflora present in natural waters can acclimate to and degrade NTA, even if exposed to only trace levels in laboratory experiments. However, rates of NTA biodegradation are more rapid and less variable in river waters where natural NTA exposure has already occurred.

Biodegradation kinetics of linear alkylbenzene sulfonate in sludge-amended agricultural soils

The kinetics of ultimate biodegradation (mineralization to CO2) of linear alkylbenzene sulfonate (LAS) were studied in sludge-amended agricultural soils for a series of pure chain length LAS homologs containing 10 to 14 carbon atoms in the alkyl chain. Degradation rates were measured by following the production of 14CO2 from uniformly 14C-ring-labeled material. In general, degradation of LAS was rapid in soil over a broad concentration range (0.1 to 10 times the expected environmental concentration) and demonstrated little variation among different homologs. Half-lives for mineralization of the benzene ring ranged from 18 to 26 days and were not significantly different for any homolog over the range of alkyl chain lengths tested. Half-lives measured for LAS degradation in these studies were comparable to values reported in the literature and also to values obtained for naturally occurring materials (stearic acid, cellulose) typically present in soil environments. On the basis of the results of the present studies and those of other investigators, it is concluded that soil environments exposed to LAS in sewage sludges contain microbial communities which can actively metabolize this material. Rates of biodegradation of the benzene ring, the final step in the LAS biodegradation pathway prior to complete mineralization, are also sufficient to prevent LAS from accumulating in soil environments.

Effect of temperature and dissolved oxygen on biodegradation of nitrilotriacetate

Abstract The effect of temperature and dissolved oxygen on the rate of biodegradation of nitrilotriacetate (NTA) was examined in water samples collected from the Rur River. Biodegradation of NTA was first order with respect to NTA concentration over a concentration range of 50–1000 μg l −1 . First order rate constants showed a typical temperature dependency (temperature coefficient, Q 10 = 2) and biodegradation of NTA was observed over a temperature range of 2–24°C. The effect of temperature on the rate of NTA biodegradation was described by the Arrhenius equation, with calculated activation energies in the range reported for ordinary enzyme reactions. Biodegradation of NTA was also observed at low dissolved oxygen concentrations (0.3 mg l −1 ), although at reduced rates compared to high oxygen concentrations (13 mg l −1 ). Biodegradation of NTA was oxygen-dependent, suggesting an obligate oxygen requirement for the initial steps in NTA metabolism by natural microbial communities in surface waters. In general, our results indicate that NTA biodegradation will occur in natural waters under conditions of low temperature and low dissolved oxygen and also at low NTA concentrations.