Predicting Hydrocarbon Primary Biodegradation in Soil and Sediment Systems Using System Parameterization and Machine Learning.

Technical complexity associated with biodegradation testing, particularly for substances of unknown or variable composition, complex reaction products, or biological materials (UVCB), necessitates the advancement of non-testing methods such as quantitative structure-property relationships (QSPRs). Models for describing the biodegradation of petroleum hydrocarbons (HCs) have been previously developed. A critical limitation of available models is their inability to capture the variability in biodegradation rates associated with variable test systems and environmental conditions. Recently, the Hydrocarbon Biodegradation System Integrated Model (HC-BioSIM) was developed to characterize the biodegradation of HCs in aquatic systems with the inclusion of key test system variables. The present study further expands the HC-BioSIM methodology to soil and sediment systems using a database of 2195 half-life (i.e., degradation time [DT]50) entries for HCs in soil and sediment. Relevance and reliability criteria were defined based on similarity to standard testing guidelines for biodegradation testing and applied to all entries in the database. The HC-BioSIM soil and sediment models significantly outperformed the existing biodegradation HC half-life (BioHCWin) and virtual evaluation of chemical properties and toxicities (VEGA) quantitative Mario Negri Institute for Pharmacological Research (IRFMN) models in soil and sediment. Average errors in predicted DT50s were reduced by up to 6.3- and 8.7-fold for soil and sediment, respectively. No significant bias as a function of HC class, carbon number, or test system parameters was observed. Model diagnostics demonstrated low variability in performance and high consistency of parameter usage/importance and rule structure, supporting the generalizability and stability of the models for application to external data sets. The HC-BioSIM provides improved accuracy of Persistence categorization, with correct classification rates of 83.9%, and 90.6% for soil and sediment, respectively, demonstrating a significant improvement over the existing BioHCWin (70.7% and 58.6%) and VEGA (59.5% and 18.5%) models. Environ Toxicol Chem 2024;43:1352-1363. © 2024 Concawe. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Comprehensive Two-Dimensional Gas Chromatography with Peak Tracking for Screening of Constituent Biodegradation in Petroleum UVCB Substances.

Petroleum substances, as archetypical UVCBs (substances of unknown or variable composition, complex reaction products, or biological substances), pose a challenge for chemical risk assessment as they contain hundreds to thousands of individual constituents. It is particularly challenging to determine the biodegradability of petroleum substances since each constituent behaves differently. Testing the whole substance provides an average biodegradation, but it would be effectively impossible to obtain all constituents and test them individually. To overcome this challenge, comprehensive two-dimensional gas chromatography (GC × GC) in combination with advanced data-handling algorithms was applied to track and calculate degradation half-times (DT50s) of individual constituents in two dispersed middle distillate gas oils in seawater. By tracking >1000 peaks (representing ∼53-54% of the total mass across the entire chromatographic area), known biodegradation patterns of oil constituents were confirmed and extended to include many hundreds not currently investigated by traditional one-dimensional GC methods. Approximately 95% of the total tracked peak mass biodegraded after 64 days. By tracking the microbial community evolution, a correlation between the presence of functional microbial communities and the observed progression of DT50s between chemical classes was demonstrated. This approach could be used to screen the persistence of GC × GC-amenable constituents of petroleum substance UVCBs.

The Path to UVCB Ecological Risk Assessment: Grappling with Substance Characterization.

Substances of unknown or variable composition, complex reaction products, and biological materials (UVCBs) pose a unique challenge to regulators and to product registrants, who are required to characterize their fate, exposure, hazard, and potential risks to human health and the environment. To address these challenges and ensure an efficient and fit-for-purpose process, it is proposed that the ecological risks of UVCBs be assessed following a tiered strategy. The development of this approach required exploring how substance composition ties into hazard and exposure information and determining the extent to which a UVCB needs to be characterized to ensure a robust risk assessment. The present study highlights the key aspects of this new method. It presents how a tiered substance characterization approach can be integrated into broader UVCB risk-assessment schemes to encourage an examination of data needs before a full substance characterization is performed. The first tier of the characterization process, Tier 0, is a fundamental step that includes data from basic, lower-resolution compositional analyses. Tier 0 assessments can be used to inform hazard and exposure for any substance of interest. The need for more sophisticated, higher-tier characterization is determined by the level of uncertainty of the risk assessment. The next step will integrate a tiered exposure assessment into the characterization scheme featured in the present study, to create a more complete risk-assessment framework. Environ Toxicol Chem 2022;41:2649-2657. © 2022 Her Majesty the Queen in Right of Canada, Health and Environmental Sciences Institute and The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. Reproduced with the permission of the Minister of Environment and Climate Change Canada.

Improving the Environmental Risk Assessment of Substances of Unknown or Variable Composition, Complex Reaction Products, or Biological Materials.

Substances of unknown or variable composition, complex reaction products, or biological materials (UVCBs) pose unique risk assessment challenges to regulators and to product registrants. These substances can contain many constituents, sometimes partially unknown and/or variable, depending on fluctuations in their source material and/or manufacturing process. International regulatory agencies have highlighted the difficulties in characterizing UVCBs and assessing their toxicity and environmental fate. Several industrial sectors have attempted to address these issues by developing frameworks and characterization methods. Based on the output of a 2016 workshop, this critical review examines current practices for UVCB risk assessment and reveals a need for a multipronged and transparent approach integrating whole-substance and constituent-based information. In silico tools or empirical measurements can provide information on discrete and/or blocks of UVCB constituents with similar hazard properties. Read-across and/or whole-substance toxicity and fate testing using adapted emerging methods can provide whole-substance information. Continued collaboration of stakeholders representing government, industry, and academia will facilitate the development of practical testing strategies and guidelines for addressing regulatory requirements for UVCBs. Environ Toxicol Chem 2020;39:2097-2108. © 2020 Health and Environmental Sciences Institute. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Multi-laboratory Validation of a New Marine Biodegradation Screening Test for Chemical Persistence Assessment.

Current biodegradation screening tests are not specifically designed for persistence assessment of chemicals, often show high inter- and intra-test variability, and often give false negative biodegradation results. Based on previous studies and recommendations, an international ring test involving 13 laboratories validated a new test method for marine biodegradation with a focus on improving the reliability of screening to determine the environmental degradation potential of chemicals. The new method incorporated increased bacterial cell concentrations to better represent the microbial diversity; a chemical is likely to be exposed in the sampled environments and ran beyond 60 days, which is the half-life threshold for chemical persistence in the marine environment. The new test provided a more reliable and less variable characterization of the biodegradation behavior of five reference chemicals (sodium benzoate, triethanolamine, 4-nitrophenol, anionic polyacrylamide, and pentachlorophenol), with respect to REACH and OSPAR persistence thresholds, than the current OECD 306 test. The proposed new method provides a cost-effective screening test for non-persistence that could streamline chemical regulation and reduce the cost and animal welfare implications of further higher tier testing.

Determining the water solubility of difficult-to-test substances: A tutorial review.

Water solubility is one of the most important and frequently used physical-chemical properties of chemicals. It is crucial within several industrial sectors, in research and in the regulatory sector e.g. for the risk and hazard assessment of chemicals. The most recent OECD guideline (Test No. 105) for measuring solubility is from 1995 and limited to mono-constituent, stable and non-volatile substances. This OECD guideline and the described methods are not suited for several groups of difficult-to-test substances, such as highly hydrophobic chemicals, volatile chemicals, surfactants and mixtures. The aim of this paper is to review solubility measurement methods for difficult-to-test substances on a technical, analytical and scientific level. Methods to handle highly hydrophobic chemicals and volatile chemicals, and methods to rapidly saturate water with fast degrading chemicals are reviewed. A decision tree is presented outlining the preferred choice of method for each chemical group. This review also includes measurement methods for critical micelle concentrations that set the upper concentration limit for freely dissolved surfactants. Finally, concepts and strategies to measure solubility parameters for mixtures, including multi-constituent substances and chemical substances of unknown or variable composition, are discussed.

In situ TCE degradation mediated by complex dehalorespiring communities during biostimulation processes.

The bioremediation of chloroethene contaminants in groundwater polluted systems is still a serious environmental challenge. Many previous studies have shown that cooperation of several dechlorinators is crucial for complete dechlorination of trichloroethene to ethene. In the present study, we used an explorative functional DNA microarray (DechloArray) to examine the composition of specific functional genes in groundwater samples in which chloroethene bioremediation was enhanced by delivery of hydrogen-releasing compounds. Our results demonstrate for the first time that complete biodegradation occurs through spatial and temporal variations of a wide diversity of dehalorespiring populations involving both Sulfurospirillum, Dehalobacter, Desulfitobacterium, Geobacter and Dehalococcoides genera. Sulfurospirillum appears to be the most active in the highly contaminated source zone, while Geobacter was only detected in the slightly contaminated downstream zone. The concomitant detection of both bvcA and vcrA genes suggests that at least two different Dehalococcoides species are probably responsible for the dechlorination of dichloroethenes and vinyl chloride to ethene. These species were not detected on sites where cis-dichloroethene accumulation was observed. These results support the notion that monitoring dechlorinators by the presence of specific functional biomarkers using a powerful tool such as DechloArray will be useful for surveying the efficiency of bioremediation strategies.

Integrity and biological activity of DNA after UV exposure.

The field of astrobiology lacks a universal marker with which to indicate the presence of life. This study supports the proposal to use nucleic acids, specifically DNA, as a signature of life (biosignature). In addition to its specificity to living organisms, DNA is a functional molecule that can confer new activities and characteristics to other organisms, following the molecular biology dogma, that is, DNA is transcribed to RNA, which is translated into proteins. Previous criticisms of the use of DNA as a biosignature have asserted that DNA molecules would be destroyed by UV radiation in space. To address this concern, DNA in plasmid form was deposited onto different surfaces and exposed to UVC radiation. The surviving DNA was quantified via the quantitative polymerase chain reaction (qPCR). Results demonstrate increased survivability of DNA attached to surfaces versus non-adsorbed DNA. The DNA was also tested for biological activity via transformation into the bacterium Acinetobacter sp. and assaying for antibiotic resistance conferred by genes encoded by the plasmid. The success of these methods to detect DNA and its gene products after UV exposure (254 nm, 3.5 J/m(2)s) not only supports the use of the DNA molecule as a biosignature on mineral surfaces but also demonstrates that the DNA retained biological activity.

Is resistance futile? Changing external resistance does not improve microbial fuel cell performance.

Microbial fuel cells (MFCs) show promise as an alternative to conventional batteries for point source electricity generation. A better understanding of the relationship between the microbiological and electrical aspects of fuels cells is needed prior to successful MFC application. Here, we observed the effects of external resistance on power production and the anodic biofilm community structure. Large differences in the external resistance affected both power production and microbial community structure. After the establishment of the anodic microbial community, change in external resistance (from low to high and vice versa) changed the anodic microbial community structure, but the resulting community did not resemble the communities established at that same external resistance. Different microbial community structures, established under different external resistances, resulted in similar power production, demonstrating the flexibility of the MFC system.

Comparative photoactivity and antibacterial properties of C60 fullerenes and titanium dioxide nanoparticles.

The production of reactive oxygen species (ROS) by aqueous suspensions of fullerenes and nano-TiO2 (Degussa P25) was measured both in ultrapure water and in minimal Davis (MD) microbial growth medium. Fullerol (hydroxylated C60) produced singlet oxygen (1O2) in ultrapure water and both 1O2 and superoxide (O2-*) in MD medium, but no hydroxyl radicals (OH*) were detected in either case. PVP/C60 (C60 encapsulated with poly(N-vinylpyrrolidone)) was more efficient than fullerol in generating singlet oxygen and superoxide. However, two other aggregates of C60, namely THF/nC60 (prepared with tetrahydofuran as transitional solvent) and aqu/nC60 (prepared by vigorous stirring of C60 powder in water), were not photoactive. Nano-TiO2 (also present as aggregates) primarily produced hydroxyl radicals in pure water and superoxide in MD medium. Bacterial (Escherichia coli) toxicity tests suggest that, unlike nano-TiO2 which was exclusively phototoxic, the antibacterial activity of fullerene suspensions was linked to ROS production. Nano-TiO2 may be more efficient for water treatment involving UV or solar energy, to enhance contaminant oxidation and perhaps for disinfection. However, fullerol and PVP/ C60 may be useful as water treatment agents targeting specific pollutants or microorganisms that are more sensitive to either superoxide or singlet oxygen.

Nanomaterials in the environment: behavior, fate, bioavailability, and effects.

The recent advances in nanotechnology and the corresponding increase in the use of nanomaterials in products in every sector of society have resulted in uncertainties regarding environmental impacts. The objectives of this review are to introduce the key aspects pertaining to nanomaterials in the environment and to discuss what is known concerning their fate, behavior, disposition, and toxicity, with a particular focus on those that make up manufactured nanomaterials. This review critiques existing nanomaterial research in freshwater, marine, and soil environments. It illustrates the paucity of existing research and demonstrates the need for additional research. Environmental scientists are encouraged to base this research on existing studies on colloidal behavior and toxicology. The need for standard reference and testing materials as well as methodology for suspension preparation and testing is also discussed.

Effect of soil sorption and aquatic natural organic matter on the antibacterial activity of a fullerene water suspension.

The present study investigated the association of a C60 water suspension (nC6) with natural organic matter, present as a soil constituent or dissolved in the water column, and its effect on the antibacterial activity of nC60. Sorption of nC60 to soil reduced its bioavailability and antibacterial activity, and the sorption capacity strongly depended on the organic content of the soil. Adsorption of aquatic dissolved humic substances onto nC60 and possible subsequent reactions also were found to eliminate nC60 toxicity at humic acid concentrations as low as 0.05 mg/L. These findings indicate that natural organic matter in the environment can mitigate significantly the potential impacts of nC60 on microbial activities that are important to ecosystem health.

Fullerene water suspension (nC60) exerts antibacterial effects via ROS-independent protein oxidation.

Buckminsterfullerene (C60) can form water suspensions (nC60) that exert toxic effects. While reactive oxygen species (ROS) generation has been implicated as the mechanism for mammalian cytotoxicity, we propose that nC60 exerts ROS-independent oxidative stress in bacteria, with evidence of protein oxidation, changes in cell membrane potential, and interruption of cellular respiration. This mechanism requires direct contact between the nanoparticle and the bacterial cell and differs from previously reported nanomaterial antibacterial mechanisms that involve ROS generation (metal oxides) or leaching of toxic elements (nanosilver).

Antimicrobial nanomaterials for water disinfection and microbial control: potential applications and implications.

The challenge to achieve appropriate disinfection without forming harmful disinfection byproducts by conventional chemical disinfectants, as well as the growing demand for decentralized or point-of-use water treatment and recycling systems calls for new technologies for efficient disinfection and microbial control. Several natural and engineered nanomaterials have demonstrated strong antimicrobial properties through diverse mechanisms including photocatalytic production of reactive oxygen species that damage cell components and viruses (e.g. TiO2, ZnO and fullerol), compromising the bacterial cell envelope (e.g. peptides, chitosan, carboxyfullerene, carbon nanotubes, ZnO and silver nanoparticles (nAg)), interruption of energy transduction (e.g. nAg and aqueous fullerene nanoparticles (nC(60))), and inhibition of enzyme activity and DNA synthesis (e.g. chitosan). Although some nanomaterials have been used as antimicrobial agents in consumer products including home purification systems as antimicrobial agents, their potential for disinfection or microbial control in system level water treatment has not been carefully evaluated. This paper reviews the antimicrobial mechanisms of several nanoparticles, discusses their merits, limitations and applicability for water disinfection and biofouling control, and highlights research needs to utilize novel nanomaterials for water treatment applications.

Antibacterial activity of fullerene water suspensions (nC60) is not due to ROS-mediated damage.

The cytotoxic and antibacterial properties of nC 60, a buckminsterfullerene water suspension, have been attributed to photocatalytically generated reactive oxygen species (ROS). However, in this work, neither ROS production nor ROS-mediated damage is found in nC 60-exposed bacteria. Furthermore, the colorimetric methods used to evaluate ROS production and damage are confounded by interactions between nC 60 and the reagents, yielding false positives. Instead, we propose that nC 60 exerts ROS-independent oxidative stress, thus reconciling conflicting results in the literature.

Effect of a fullerene water suspension on bacterial phospholipids and membrane phase behavior.

Several fullerene-based nanomaterials generate reactive oxygen species that can damage cells. In this study, we investigated the effect of buckminsterfullerene (C60) introduced as colloidal aggregates in water (nC60) on bacterial membrane lipid composition and phase behavior. Pseudomonas putida (Gram-negative) and Bacillus subtilis (Gram-positive) responded to nC60 by altering membrane lipid composition, phase transition temperature, and membrane fluidity. P. putida decreased its levels of unsaturated fatty acids and increased the proportions of cyclopropane fatty acids in the presence of nC60, possibly to protect the bacterial membrane from oxidative stress. Fourier transform infrared spectroscopy measurement of intact bacterial cells showed slightly increased phase transition temperatures (Tm) and increased membrane fluidity for cells grown in the presence of high, growth-inhibiting concentrations (0.5 mg L(-1)) of nC60. B. subtilis responded to a low dose of nC6o (0.01 mg L(-1)) by significantly increasing the levels of iso- and anteiso-branched fatty acids (from 5.8 to 31.5% and 12.9 to 32.3% of total fatty acids, respectively) and to a high, growth-inhibiting concentration of nC60 (0.75 mg L-1) by increasing synthesis of monounsaturated fatty acids. In contrast to P. putida, B. subtilis response was a decrease in Tm and an increase in membrane fluidity. These findings represent the first demonstrated physiological adaptation response of bacteria to a manufactured nanomaterial, and they showthat response inlipid composition and membrane phase behavior depends on both the nC60 concentration and the cell wall morphology.

Comparative eco-toxicity of nanoscale TiO2, SiO2, and ZnO water suspensions.

The potential eco-toxicity of nanosized titanium dioxide (TiO(2)), silicon dioxide (SiO(2)), and zinc oxide (ZnO) water suspensions was investigated using Gram-positive Bacillus subtilis and Gram-negative Escherichia coli as test organisms. These three photosensitive nanomaterials were harmful to varying degrees, with antibacterial activity increasing with particle concentration. Antibacterial activity generally increased from SiO(2) to TiO(2) to ZnO, and B. subtilis was most susceptible to their effects. Advertised nanoparticle size did not correspond to true particle size. Apparently, aggregation produced similarly sized particles that had similar antibacterial activity at a given concentration. The presence of light was a significant factor under most conditions tested, presumably due to its role in promoting generation of reactive oxygen species (ROS). However, bacterial growth inhibition was also observed under dark conditions, indicating that undetermined mechanisms additional to photocatalytic ROS production were responsible for toxicity. These results highlight the need for caution during the use and disposal of such manufactured nanomaterials to prevent unintended environmental impacts, as well as the importance of further research on the mechanisms and factors that increase toxicity to enhance risk management.

Antibacterial activity of fullerene water suspensions: effects of preparation method and particle size.

Fullerene research in biological systems has been hindered by the compound's relative insolubility in water. However, C60 molecules can be made to aggregate, forming stable fullerene water suspensions (FWS) whose properties differ from those of bulk solid C60. There are many different protocols for making FWS. This paper explores four of these methods and establishes the antibacterial activity of each resulting suspension, including a suspension made without intermediary solvents. The aggregates in each polydisperse suspension were separated by size using differential centrifugation and tested for antibacterial activity using Bacillus subtilis as a test organism. All suspensions exhibited relatively strong antibacterial activity. Fractions containing smaller aggregates had greater antibacterial activity, although the increase in toxicity was disproportionately higher than the associated increase in putative surface area. This suggests the need for improved understanding of the behavior of FWS towards organisms and in the environment to determine how C60 can be safely used and disposed.

Bacterial cell association and antimicrobial activity of a C60 water suspension.

Prior to the implementation of any new technology, possible environmental and health repercussions first must be researched. Fullerenes are to be produced soon on an industrial scale, with applications quickly following. To investigate the possible environmental impact of fullerenes, a C60-water suspension (nano-C60) was synthesized and then evaluated for cell-association and toxicity, using the bacteria Escherichia coli and Bacillus subtilis as indicator species. In a defined low-salts medium, nano-C60 associated with both the Gram-negative E. coli and the Gram-positive B. subtilis, albeit more strongly with the former. Nano-C60 also displayed antimicrobial properties against both E. coli and B. subtilis, with minimal inhibitory concentrations of 0.5 to 1 mg/ L and 1.5 to 3.0 mg/L, respectively. Media with higher salt contents result in the nano-C60 particles aggregating and falling out of suspension; thus, higher salt solutions reduced or eliminated the antimicrobial properties of nano-C60.

Flux and product distribution during biological treatment of tetrachloroethene dense non-aqueous-phase liquid.

Flux in non-aqueous-phase liquid (NAPL)-contaminated systems containing active microbial populations (including Dehalococcoides sp.) was investigated using a quantitative mass balance and phase distribution approach. Batch systems containing mixed NAPL with an initial tetrachloroethene (PCE) mole fraction ranging from 0.1 to 0.4 provided a means for comparing systems where mass transfer and aqueous concentration were controlled by the initial NAPL composition. Although the use of mixed NAPL with increasing PCE mole fractions introduced a mass-transfer variable on the abiotic dissolution rate, it was determined that biological systems produced flux rates that were similar to each other regardless of the initial PCE mole fraction. Thus, organisms appeared to be dechlorinating near their maximum conversion rates, and the result was the accumulation of cis-1,2-dichloroethene (cDCE) followed by slow conversion to vinyl chloride (VC). Increases in the initial PCE mole fractions in the NAPL had a negative impact on product distribution due to the presence of a larger concentration of a more favorable electron acceptor. Because the mass converted to cDCE was present largely in the dissolved phase in all systems, the production of this metabolite was a favorable outcome in terms of NAPL dissolution. The pH dropped as low as 4.9 in active systems, indicating that the amount of HCl released during the reductive dechlorination process was large enough to overwhelm the buffering capacity. This pH effect was more pronounced in systems that exhibited extensive dechlorination to VC, further suggesting that rapid dechlorination of PCE NAPL can alter chemical characteristics in source zone regions.