Researchers have long theorized that solar irradiation of seawater generates low, steady-state levels of reactive halogen species, or RHS (e.g., Br•, ClBr•-, Br2•-, Br2, and HOBr) at the ocean surface. This is expected to occur primarily through the reactions of bromide with hydroxyl radical (HO•), which may be generated by various processes at steady-state concentrations ranging from 10-18 to 10-17 mol/L. Some studies also suggest that RHS may be formed through interactions of photoexcited triplet-state dissolved organic matter, 3DOM*, with bromide. Furthermore, absorptive uptake of ambient ozone (O3) at the ocean surface has recently been identified as a possible source of iodine-containing RHS (e.g., HOI).

A number of RHS (e.g., Br2, HOBr, HOI) are known to be capable of brominating or iodinating dissolved organic matter (DOM), and could in turn represent an important, but relatively unexplored abiotic source of organohalogens at the ocean surface, which could have important implications for a variety of atmospheric processes (e.g., tropospheric and stratospheric O3 depletion, cloud formation, mercury deposition). In the interest of characterizing the potential contributions of sunlight- and O3-driven RHS production to organohalogen formation, we have developed complementary analytical protocols to assess: (a) non-specific halogenation of bulk DOM, and (b) specific halogenation of a well-defined probe compound, during exposure of seawaters to natural levels of sunlight and/or ozone. 

In protocol (a), non-specific DOM bromination and iodination are quantified by ICP-MS analysis of bulk organohalogen concentrations in DOM extracts after sequential solid phase extraction and halide-selective ion exchange. In protocol (b), a non-volatile, UVA-transparent heterocyclic nitrogen compound which yields a single brominated or iodinated product upon reaction with non-radical brominating or iodinating species (i.e., X2, HOX) is utilized as a selective probe for the detection of RHS. Halogenated product yields in seawater samples are then quantified using LC-MS/MS with multiple reaction monitoring. Through the combination of these approaches, we are quantifying the formation of low (typically nanomolar) levels of bulk organobromine and organoiodine in seawater during exposure to solar radiation or ozone, in order to evaluate the significance of photochemical or ozone-driven halogenation of dissolved organic matter as an abiotic source of organohalogens at the ocean surface. Our work focuses in particular on the role of such processes in estuarine seawaters, which are enriched in photoreactive terrestrial DOM originating from riverine plumes.

Aqueous free available chlorine (typically comprising Cl2, HOCl, and OCl-) remains widely utilized as a disinfectant in water treatment on account of its low cost, ease of use, and generally high efficacy. However, certain microbial agents (e.g., bacterial endospores, Cryptosporidium parvum oocysts, Giardia lamblia cysts, Mycobacterium avium cells) exhibit high levels of chlorine-resistance, often necessitating the use of capital- and energy-intensive alternative approaches to disinfection (e.g., UV irradiation, ozonation) to ensure acceptable water quality. In recent investigations, we have found that the effectiveness of chlorine-based disinfection processes toward such microorganisms can be markedly enhanced simply by utilizing sunlight to generate the potent oxidants hydroxyl radical (•OH) and ozone (O3) in situ through the photolysis of free chlorine.

For example, the chlorine exposure, or CT (in mg Cl2 L-1 min), required for 99% inactivation of highly chlorine-resistant Bacillus subtilis endospores (common surrogates for chlorine-resistant pathogens) can be lowered by two-thirds during simulated solar photolysis of chlorine at pH 8 and 10° C, as a result of co-exposure of spores to O3 and •OH. We have also found that C. parvum oocysts can be inactivated by >99% under similar conditions, even though no oocyst inactivation is achievable in the absence of light.

Our ongoing research on this topic focuses on elucidating the mechanisms responsible for inactivation of microorganisms (particularly with respect to synergism amongst multiple disinfectants), investigating the potential for inactivation of other chlorine-resistant pathogens such as M. avium and Coxsackievirus B5, and quantifying impacts of chlorine photolysis on the formation of regulated inorganic and organic disinfection by-products (e.g., chlorite, bromate, trihalomethanes, and haloacetic acids). One of our primary objectives in this investigation is development of a framework for accurately predicting the inactivation of chlorine-resistant microorganisms during sunlight-driven chlorine photolysis, with the ultimate aim of facilitating practical implementation of this approach for use in small municipal water systems and decentralized, point-of-use treatment applications (e.g., as a means of boosting the efficacy of solar disinfection, or SODIS).

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Free available chlorine is most commonly produced through the electrolysis of HCl or NaCl solutions to generate Cl2(g) or NaOCl. However, energy consumption associated with the electrolytic production of chlorine is extremely high, necessitating either dedicated electrical supplies at larger scales, or battery-powered electrochemical cells for smaller scale, decentralized applications. In the interest of lowering the energy footprint and accessibility of chlorine production, we are investigating a potential alternative means for generating chlorine independent of electrical input and with very limited physical infrastructure. Certain classes of natural and/or synthetic organic photosensitizers (e.g., aromatic ketones) can be utilized to produce Cl2(g) directly via sunlight-mediated oxidation of aqueous HCl or of acidified brines containing high concentrations of Cl- (e.g., seawater or NaCl solutions). The resulting Cl2(g) can be sparged with a continuous stream of external gas into a “trap” vessel containing an alkaline solution (to generate a NaOCl stock) or a raw drinking water (to provide direct disinfection), as depicted in the accompanying figure.

We are characterizing and optimizing the photosensitized oxidation of Cl- to Cl2 by such sensitizer compounds through the use of simulated and natural sunlight. The near-term objective of this investigation is the development of a portable, energy-efficient means of generating aqueous free chlorine that may subsequently be used alone or in combination with other processes for the inactivation of pathogenic microorganisms in compromised drinking water sources – without the need for an external electrical power supply. The broader objectives of the investigation are to: 1) expand opportunities for point-of-use drinking water disinfection through on-site generation of chlorine, 2) adapt the technology for use in improving hygiene and access to safe drinking water in developing communities by utilizing widely-available natural products as raw materials, and 3) facilitate development of a “greener,” more sustainable approach to chlorine generation.

Antibiotic resistance genes (ARGs) – present within extracellular DNA or within intracellular DNA in association with antibiotic resistant bacterial (ARB) cells – have been identified as widespread contaminants of treated drinking waters and wastewaters. We are investigating the effectiveness of common drinking water and wastewater disinfectants (i.e., HOCl, NH2Cl, O3, ClO2, and UV light) in deactivating the biological activities of ARGs associated with both extracellular and intracellular ARB DNA. As disinfectant reactivities toward DNA and other cell constituents vary widely, substantial variation is anticipated in the efficacy with which each disinfectant is able to penetrate to and deactivate ARGs contained within the cytoplasm of a given ARB cell, as depicted in the following Figure (note that UV penetrates relatively uniformly to DNA

Figure. Diffusion of HOCl, NH2Cl, O3, and ClO2 into (a) a generic vegetative prokaryotic cell, and (b) probable variations in oxidant concentrations with increasing diffusion distance into the cell, in accord with oxidant reactivities toward constituents of the cell wall and cytoplasm.

contained within the cytoplasm). As a consequence, one can anticipate substantial differences in the relative extent to which intracellular DNA (and associated ARGs) would be degraded during treatment with each disinfectant.

As a major focus of this research, we are utilizing culture-based bacterial DNA transformation assays and real-time quantitative PCR to quantify the kinetics of ARG deactivation (i.e., loss of transforming activity) and degradation (i.e., loss of measurable gene copies) in various strains of bacteria during disinfection. In addition, the kinetics of intracellular ARG deactivation by each disinfectant are being correlated with measured rates of ARB cell inactivation to evaluate the likelihood that intact, biologically-active ARGs may persist within the remaining ARB cell debris even after cell death, and to determine the feasibility of modeling their degradation and deactivation during various water and wastewater disinfection processes. As an extension of this work, we are also investigating the impacts of exposure to disinfectants commonly used in healthcare practice (e.g., benzalkonium chloride, povidone-iodine, hydrogen peroxide, chlorhexidine) on ARG integrity within clinically-relevant strains of antibiotic resistant pathogenic bacteria. The results of this work are anticipated to facilitate optimization of disinfection for minimizing the dissemination of intact ARGs within natural and engineered aquatic systems, as well as in healthcare settings.