Limitations of conventional approaches to identify photochemically produced reactive intermediates involved in contaminant indirect photodegradation.

Dissolved organic matter (DOM) mediated indirect photodegradation can play an important role in the degradation of aquatic contaminants. Predicting the rate of this process requires knowledge of the photochemically produced reactive intermediates (PPRI) that react with the compound of interest, as well as the ability of individual DOM samples to produce PPRI. Key PPRI are typically identified using quencher studies, yet this approach often leads to results that are difficult to interpret. In this work, we analyze the indirect photodegradation of atorvastatin, carbamazepine, sulfadiazine, and benzotriazole using a diverse set of 48 waters from natural and engineered aquatic systems. We use this large data set to evaluate relationships between PPRI formation and indirect photodegradation rate constants, which are directly compared to results using standard quenching experiments. These data demonstrate that triplet state DOM (3DOM) and singlet oxygen (1O2) are critical PPRI for atorvastatin, carbamazepine, and sulfadiazine, while hydroxyl radical (˙OH) contributes to the indirect photodegradation of benzotriazole. We caution against relying on quenching studies because quenching of 3DOM limits the formation of 1O2 and all studied quenchers react with ˙OH. Furthermore, we show that DOM composition directly influences indirect photodegradation and that low molecular weight, microbial-like DOM is positively correlated with the indirect photodegradation rates of carbamazepine, sulfadiazine, and benzotriazole.

Biodegradation, photolysis, and sorption of antibiotics in aquatic environments: A scoping review.

The presence of antibiotics in surface waters is a potential driver of antibiotic resistance and thus of concern to human and environmental health. Key factors driving the potential impact of antibiotics are their persistence and transport in rivers and lakes. The goal of this study was to describe the peer-reviewed published literature on the photolysis (direct and indirect), sorption, and biodegradation of a selected group of antibiotic compounds following a scoping review methodology. Primary research from 2000 to 2021 was surveyed to compile information on these processes for 25 antibiotics from 6 classes. After compilation and assessment of the available parameters, the results indicate that information is present to predict the rates of direct photolysis and reaction with hydroxyl radical (an indirect photolysis process) for most of the selected antibiotics. There is insufficient or inconsistent information for including other indirect photolysis processes, biodegradation, or removal via sorption to settling particles for most of the targeted antibiotic compounds. Future research should focus on collecting fundamental parameters such as quantum yields, second-order rate constants, normalized biodegradation rates, and organic carbon or surface area normalized sorption coefficients rather than pseudo-first order rate constants or sorption equilibrium constants that apply only to specific conditions/sites.

Dissolved Organic Matter Photoreactivity Is Determined by Its Optical Properties, Redox Activity, and Molecular Composition.

Predicting the formation of photochemically produced reactive intermediates (PPRI) during the irradiation of dissolved organic matter (DOM) has remained challenging given the complex nature of this material and differences in PPRI formation mechanisms. We investigate the role of DOM composition in photoreactivity using 48 samples that span the range of DOM in freshwater systems and wastewater. We relate quantum yields for excited triplet-state organic matter (fTMP), singlet oxygen (Φ1O2), and hydroxylating species (Φ•OH) to DOM composition determined using spectroscopy, Fourier-transform ion cyclotron resonance mass spectrometry, and electron-donating capacity (EDC). fTMP and Φ1O2 follow similar trends and are correlated with bulk properties derived from UV-vis spectra and EDC. In contrast, no individual bulk property can be used to predict Φ•OH. At the molecular level, the subset of DOM that is positively correlated to both Φ•OH and EDC is distinct from DOM formulas related to Φ1O2, demonstrating that •OH and 1O2 are formed from different DOM fractions. Multiple linear regressions are used to relate quantum yields of each PPRI to DOM composition parameters derived from multiple techniques, demonstrating that complementary methods are ideal for characterizing DOM because each technique only samples a subset of DOM.

Identifying the spatiotemporal vulnerability of soils to antimicrobial contamination through land application of animal manure in Minnesota, United States.

Antimicrobials may reach the soil environment from a variety of sources and pathways, including land application of human biosolids and animal manure. Once in soil, antimicrobials can affect the abundance and activity of soil microorganisms and exert selection pressures that enhance the emergence and spread of antimicrobial resistance (AMR). To mitigate the spread of AMR it is important to understand the spatial and temporal interactions between antimicrobials and soil. The goal of this study was to assess the vulnerability of Minnesota (U.S.) soil to contamination with specific antimicrobial compounds at temperatures experienced throughout the year. Soil contamination potential was estimated based upon specific antimicrobial drug binding and permanence, and average monthly temperature. Minnesota soil vulnerability was estimated by incorporating spatially explicit soil contamination potential, land cover type, and livestock density. Assessment of antimicrobials used in livestock production showed that soils are most vulnerable to antimicrobial contamination in southwestern Minnesota, to enrofloxacin, chlortetracycline, and oxytetracycline, and in the months of April and October. While the assessment herein was not based on actual on-farm antimicrobial use data and subsequent excretion of antimicrobial metabolites into the environment, this study provides an overview of the spatial and temporal potential for Minnesota soil to be contaminated by several antimicrobial drugs and demonstrates how specific vulnerability assessments might be conducted for geographic areas with known exposure (e.g., cropland fertilized with livestock manure and/or human biosolids). Such assessments might be used to identify best practices for mitigating antimicrobial exposure to soils and guide additional research to understand the role of environmental antimicrobial contamination in the problem of AMR.

Quantifying and predicting antimicrobials and antimicrobial resistance genes in waterbodies through a holistic approach: a study in Minnesota, United States.

The environment plays a key role in the spread and persistence of antimicrobial resistance (AMR). Antimicrobials and antimicrobial resistance genes (ARG) are released into the environment from sources such as wastewater treatment plants, and animal farms. This study describes an approach guided by spatial mapping to quantify and predict antimicrobials and ARG in Minnesota's waterbodies in water and sediment at two spatial scales: macro, throughout the state, and micro, in specific waterbodies. At the macroscale, the highest concentrations across all antimicrobial classes were found near populated areas. Kernel interpolation provided an approximation of antimicrobial concentrations and ARG abundance at unsampled locations. However, there was high uncertainty in these predictions, due in part to low study power and large distances between sites. At the microscale, wastewater treatment plants had an effect on ARG abundance (sul1 and sul2 in water; blaSHV, intl1, mexB, and sul2 in sediment), but not on antimicrobial concentrations. Results from sediment reflected a long-term history, while water reflected a more transient record of antimicrobials and ARG. This study highlights the value of using spatial analyses, different spatial scales, and sampling matrices, to design an environmental monitoring approach to advance our understanding of AMR persistence and dissemination.

Photolysis of Trenbolone Acetate Metabolites in the Presence of Nucleophiles: Evidence for Metastable Photoaddition Products and Reversible Associations with Dissolved Organic Matter.

Photolysis of trenbolone acetate (TBA) metabolites in the presence of various nitrogen-, sulfur-, or oxygen-containing nucleophiles (e.g., azide, ammonia, or thiosulfate, respectively) results in rapid (half-lives ∼20-60 min), photochemically induced nucleophile incorporation across the parent steroid's trienone moiety. The formation of such nucleophile adducts limits formation of photohydrates, suggesting competition between the nucleophile and water for photochemical addition into the activated steroid structure. Analogous to previously reported photohydration outcomes, LC/MS analyses suggest that such photonucleophilic addition reactions are reversible, with more rapid elimination rates than thermal dehydration of photohydrates, and regenerate parent steroid structures. Beyond photonucleophilic addition pathways, we also found that hydroxylamine and presumed nucleophilic moieties in model dissolved organic matter (DOM; Fluka humic acid) can react via thermal substitution with TBA metabolite photohydrates, although this reaction with model DOM was only observed for photohydrates of trendione. Most nucleophile addition products [i.e., formed via (photo)reaction with thiosulfate, hydroxylamine, and ammonia] are notably more polar relative to the parent metabolite and photohydration products. Thus, if present, both nucleophilic adducts and bound residues in organic matter will facilitate transport and help mask detection of TBA metabolites in surface waters and treatment systems.

The Role of Dissolved Organic Matter Composition in Determining Photochemical Reactivity at the Molecular Level.

Dissolved organic matter (DOM) composition influences its ability to form photochemically produced reactive intermediates (PPRI). While relationships have been established between bulk DOM properties and triplet DOM (3DOM) and singlet oxygen (1O2) quantum yields, contradictory evidence exists for hydroxyl radical (•OH) and hydroxylating species. Furthermore, little is known about these relationships at the molecular level. We evaluated DOM composition and photochemical reactivity of water samples from a wastewater treatment plant and the St. Louis River in Minnesota and Wisconsin, U.S.A. Bulk characterization using ultraviolet-visible spectroscopy demonstrates that color and apparent size of DOM decrease downstream, while molecular composition analysis using Fourier-transform ion cyclotron resonance mass spectrometry reveals that saturation and chemodiversity is highest near Lake Superior. 3DOM quantum yield coefficients and 1O2 quantum yields increase downstream and correlate strongly with saturated formulas. Similar results are observed for carbon-normalized photodegradation rate constants of atorvastatin, carbamazepine, and venlafaxine, which react primarily with 3DOM and 1O2. In contrast, •OH quantum yields are lowest downstream and correlate with less saturated, more oxygenated DOM, suggesting that 3DOM is not its major precursor. Mixed relationships are observed for DEET, which reacts with multiple PPRI. Molecular-level compositional data reveal insights into the differing formation pathways of individual PPRI, but information about specific contaminants is needed to predict their photochemical fate.

Intramolecular [2 + 2] Photocycloaddition of Altrenogest: Confirmation of Product Structure, Theoretical Mechanistic Insight, and Bioactivity Assessment.

While studying the environmental fate of potent endocrine-active steroid hormones, we observed the formation of an intramolecular [2 + 2] photocycloaddition product (2) with a novel hexacyclic ring system following the photolysis of altrenogest (1). The structure and absolute configuration were established by X-ray diffraction analysis. Theoretical computations identified a barrierless two-step cyclization mechanism for the formation of 2 upon photoexcitation. 2 exhibited progesterone, estrogen, androgen, and pregnane X receptor activity, albeit generally with reduced potency relative to 1.

Bioactive Rearrangement Products from Aqueous Photolysis of Pharmaceutical Steroids.

In an ongoing effort to study the environmental fate of endocrine-active steroid hormones, we report the formation of phenolic rearrangement products (3 and 4) with a novel 6,5,8,5-ring system following aqueous photolysis of dienogest (1) and methyldienolone (2). The structures were established by analysis of 2D NMR and HRMS data, and that of 3 was confirmed by X-ray diffraction analysis. These photoproducts exhibit progestogenic and androgenic activity, albeit with less potency than their parent compounds.

Comment on "Photodegradation of sulfathiazole under simulated sunlight: Kinetics, photo-induced structural rearrangement, and antimicrobial activities of photoproducts" by Niu et al. [Water Research 124 2017 576-583].

Recent efforts have employed antimicrobial susceptibility assays to describe the residual antimicrobial activity of antibiotics and their transformation products in a variety of environmental processes. Some authors have evaluated the results of these assays using the minimum inhibitory concentration (MIC); however, this approach has fundamental weaknesses. To highlight best practices, this comment describes the advantages of using dose-response curves to calculate the half maximal inhibitory concentration (IC50) and the potential impacts of growth media on the antimicrobial activity of sulfonamide antibiotics.

Environmental photochemistry of dienogest: phototransformation to estrogenic products and increased environmental persistence via reversible photohydration.

Potent trienone and dienone steroid hormones undergo a coupled photohydration (in light)-thermal dehydration (in dark) cycle that ultimately increases their environmental persistence. Here, we studied the photolysis of dienogest, a dienone progestin prescribed as a next-generation oral contraceptive, and used high resolution mass spectrometry and both 1D and 2D nuclear magnetic resonance spectroscopy to identify its phototransformation products. Dienogest undergoes rapid direct photolysis (t1/2 ∼ 1-10 min), forming complex photoproduct mixtures across the pH range examined (pH 2 to 7). Identified products include three photohydrates that account for ∼80% of the converted mass at pH 7 and revert back to parent dienogest in the absence of light. Notably, we also identified two estrogenic compounds produced via the A-ring aromatization of dienogest, evidence for a photochemically-induced increase in estrogenic activity in product mixtures. These results imply that dienogest will undergo complete and facile photolytic transformation in sunlit surface water, yet exhibit greater environmental persistence than might be anticipated by inspection of kinetic rates. Photoproduct mixtures also include transformation products with different nuclear receptor binding capabilities than the parent compound dienogest. These outcomes reveal a dynamic fate and biological risk profile for dienogest that must also take into account the composition and endocrine activity of its transformation products. Collectively, this study further illustrates the need for more holistic regulatory, risk assessment, and monitoring approaches for high potency synthetic pharmaceuticals and their bioactive transformation products.

Environmental Photochemistry of Altrenogest: Photoisomerization to a Bioactive Product with Increased Environmental Persistence via Reversible Photohydration.

Despite its wide use as a veterinary pharmaceutical, environmental fate data is lacking for altrenogest, a potent synthetic progestin. Here, it is reported that direct photolysis of altrenogest under environmentally relevant conditions was extremely efficient and rapid (half-life ∼25 s). Photolysis rates (observed rate constant kobs = 2.7 ± 0.2 × 10(-2) s(-1)) were unaffected by changes in pH or temperature but were sensitive to oxygen concentrations (N2-saturated kobs = 9.10 ± 0.32 × 10(-2) s(-1); O2-saturated kobs = 1.38 ± 0.11 × 10(-2) s(-1)). The primary photoproduct was identified as an isomer formed via an internal 2 + 2 cycloaddition reaction; the triplet lifetime (8.4 ± 0.2 µs) and rate constant (8 × 10(4) s(-1)) of this reaction were measured using transient absorption spectroscopy. Subsequent characterization determined that this primary cycloaddition photoproduct undergoes photohydration. The resultant photostable secondary photoproducts are subject to thermal dehydration in dark conditions, leading to reversion to the primary cycloaddition photoproduct on a time scale of hours to days, with the photohydration and dehydration repeatable over several light/dark cycles. This dehydration reaction occurs more rapidly at higher temperatures and is also accelerated at both high and low pH values. In vitro androgen receptor (AR)-dependent gene transcriptional activation cell assays and in silico nuclear hormone receptor screening revealed that certain photoproducts retain significant androgenic activity, which has implications for exposure risks associated with the presence and cycling of altrenogest and its photoproducts in the environment.

Direct photochemistry of three fluoroquinolone antibacterials: norfloxacin, ofloxacin, and enrofloxacin.

Fluoroquinolone (FQ) antibacterial compounds are frequently detected in the aquatic environment, and photodegradation is expected to play an important role in FQ fate in some sunlit surface waters. This study investigated the direct aquatic photochemistry of three FQs: norfloxacin, ofloxacin, and enrofloxacin. The direct photolysis rate of each drug exhibited strong pH dependence when exposed to simulated sunlight. For each FQ, direct photolysis rates and total light absorbance were used to calculate quantum yields for each of three environmentally relevant protonation states: a cationic, a zwitterionic, and an anionic form. In each case, quantum yields of the species varied significantly. The quantum yield for the zwitterionic form was 2-3 times higher than that of the anionic form and over an order of magnitude higher than that of the cationic form. Antibacterial activity assays were used to determine whether the loss of parent FQ due to photolysis led to loss of activity. Norfloxacin and ofloxacin photoproducts were found to be inactive, whereas enrofloxacin photoproducts were found to retain significant activity. These results are important for aiding in predictions of the potential impacts of FQs in surface waters.

Tetracycline photolysis in natural waters: loss of antibacterial activity.

Previous work has shown that tetracycline undergoes direct photolysis in the presence of sunlight, with the decomposition rate highly dependent on conditions such as water hardness and pH. The purpose of this study was to examine the potential long-term significance of photoproducts formed when tetracycline undergoes photodegradation under a range of environmentally relevant conditions. Tetracycline was photolyzed in nine different natural and artificial water samples using simulated sunlight. The pH values of the samples ranged from 5 to 9. Total hardness values (combined Ca(2+) and Mg(2+) concentrations) varied from 30 to 450 ppm. Assays based on growth inhibition of two bacterial strains, Escherichia coli DH5α and Vibrio fischeri, were used to determine the antibacterial activity of tetracycline's photoproducts in these water samples. In all tested conditions, it was determined that the photoproducts retain no significant antibacterial activity; all observed growth inhibition was attributable to residual tetracycline. This suggests that tetracycline photoproducts formed under a wide range of pH and water hardness conditions will not contribute to the selection of antibiotic-resistant bacteria in environmental systems.

Oxidation of antibiotics during water treatment with potassium permanganate: reaction pathways and deactivation.

Recent work demonstrates that three widely administered antibiotics (ciprofloxacin, lincomycin, and trimethoprim) are oxidized by potassium permanganate [KMnO(4), Mn(VII)] under conditions relevant to water treatment operations. However, tests show that little to no mineralization occurs during reactions with Mn(VII), so studies were undertaken to characterize the reaction products and pathways and to assess the effects of Mn(VII)-mediated transformations on the antibacterial activity of solutions. Several oxidation products were identified for each antibiotic by liquid chromatography-tandem mass spectrometry (LC-MS/MS). For ciprofloxacin, 12 products were identified, consistent with oxidation of the tertiary aromatic and secondary aliphatic amine groups on the piperazine ring and the cyclopropyl group. For lincomycin, seven products were identified that indicate structural changes to the pyrrolidine ring and thioether group. For trimethoprim, seven products were identified, consistent with Mn(VII) reaction at C═C double bonds on the pyrimidine ring and the bridging methylene group. Oxidation pathways are proposed based on the identified products. Bacterial growth inhibition bioassays (E. coli DH5α) show that the mixture of products resulting from Mn(VII) reactions with the antibiotics collectively retain negligible antibacterial potency in comparison to the parent antibiotics. These results suggest that permanganate can be an effective reagent for eliminating the pharmaceutical activity of selected micropollutants during drinking water treatment.

Environmental photochemistry of tylosin: efficient, reversible photoisomerization to a less-active isomer, followed by photolysis.

The environmental photochemical kinetics of tylosin, a common veterinary macrolide antibiotic and growth promoter, were investigated under simulated sunlight. An efficient, reversible photoisomerization was characterized using kinetic, mass spectrometry, and proton nuclear magnetic resonance data. The photoisomerization was confirmed to occur by a rotation about the distal alkene of the ketodiene functionality. Concurrent forward (quantum yield = 0.39 +/- 0.09) and back (quantum yield = 0.32 +/- 0.08) reactions lead to a photochemical equilibrium near a tylosin/photoisomer ratio of 50:50, completed in less than 2 min under a spectrum equivalent to noontime, summer sunlight. The activity of the isomer for the inhibition of Escherichia coli DH5alpha growth was observed to be less than that of tylosin. On a longer time scale than that of isomerization, the isomer mixture undergoes photolysis with a quantum yield of (1.4 +/- 0.3) x 10(-3). The observed quantum yields and UV-vis absorbance data allow for the prediction of the photochemical behavior of tylosin in most environmental systems. Indirect photosensitization was not a significant loss process in solutions of Suwannee River fulvic acid with concentrations from 1 to 20 mg L(-1).

Changes in antibacterial activity of triclosan and sulfa drugs due to photochemical transformations.

Sulfa drugs and triclosan represent two classes of antibacterials that have been found in natural waters and for which photodegradation is anticipated to be a significant loss process. Parent antibacterial compounds and the products of photolysis reactions were compared for three sulfa drugs and triclosan to determine the extent to which photolysis affects their antibacterial potency on Escherichia coli DH5alpha. Sulfathiazole (median effective concentration [EC50] = 20.0 microM), sulfamethoxazole (EC50 = 12.3 microM), and sulfachloropyridazine (EC50 = 6.9 microM) inhibited bacterial growth but did not affect respiratory activity. Photolysis products of these sulfa drugs did not retain any measurable ability to inhibit growth. Triclosan inhibited both the growth (EC50 = 0.24 microM) and respiratory activity of E. coli DH5alpha. Triclosan photolysis products also exhibited no measurable effect on growth or respiratory activity. These experiments indicate that the products of triclosan and sulfa drug photolysis are unlikely to possess antibacterial activity in natural waters. The rapid screening method used for these two classes of compounds will be useful for helping to identify photolabile antibacterial compounds, for which photoproducts could require further investigation.

A molecular modeling analysis of polycyclic aromatic hydrocarbon biodegradation by naphthalene dioxygenase.

A theoretical analysis was performed to examine the role of naphthalene dioxygenase (NDO) enzymes in determining differences in biodegradability and biodegradation rates of two- to four-ring polycyclic aromatic hydrocarbons (PAHs) via oxygenation and desaturation reactions. Investigation of the thermodynamics of PAH biodegradation reactions catalyzed by NDO revealed that enthalpies of reaction can explain reaction patterns or regioselectivity of the enzyme in limited cases. Molecular modeling analysis of the size and shape constraints of PAH-enzyme interactions suggests that PAHs bigger than approximately four rings and compounds with alpha substituents or other structural features contributing to increased width at the end of the substrate near the active site are expected to have binding difficulties. This explains some regioselectivity observations, in that thermodynamically favorable sites on some PAH molecules cannot be positioned correctly to be oxidized at the active site. The enzyme fit analysis also suggests that slower biodegradation rates are expected for compounds with larger widths because of the unique positioning that is required for reaction to occur. An inverse relationship between a molecular descriptor of compound width and previously obtained biodegradation rates suggests that this descriptor may be valuable for predicting relative biodegradation rates of PAHs with dioxygenases other than NDO.

Polycyclic aromatic hydrocarbon biodegradation rates: a structure-based study.

This study was designed to examine the role of molecular structure in determining the biodegradation rates of polycyclic aromatic hydrocarbons (PAHs). Laboratory experiments were performed in aqueous systems, and data were analyzed in a manner that allowed determination of first-order biodegradation rates independent of bioavailability limitations from physical-chemical processes. An aerobic mixed culture was used, which had been enriched on a broad range of PAHs. The 22 PAHs included in this study ranged in size from two to four rings and included compounds with 5-carbon rings and alkyl substituents. The range of observed biodegradation rates was only 1 order of magnitude, which is much less than that which is typically observed in the field. This supports the findings of other types of studies, which conclude that most of the observed variation in environmental PAH biodegradation rates comes from processes controlling the bioavailability of the compounds and not processes controlling uptake or biotransformation. Rate differences that were observed were attributable either to the presence of a 5-carbon ring or an alkyl substituent in an alpha position. Various molecular descriptors that might be expected to correlate with rate-limiting steps in the biodegradation process were used in an attemptto develop a quantitative structure-activity relationship for the PAH biodegradation rates. No significant correlations were found, but rate limitation from interactions with the relevant enzymes remains a possibility.