Accepted Manuscript A proposed abiotic reaction scheme for hydroxylamine and monochloramine under chloramination relevant drinking water conditions David G. Wahman , Gerald E. Speitel Jr., Madhav V. Machavaram PII:

S0043-1354(14)00345-5

DOI:

10.1016/j.watres.2014.04.051

Reference:

WR 10654

To appear in:

Water Research

Received Date: 24 February 2014 Revised Date:

24 April 2014

Accepted Date: 29 April 2014

Please cite this article as: Wahman, D.G., Speitel Jr., , G.E., Machavaram, M.V., A proposed abiotic reaction scheme for hydroxylamine and monochloramine under chloramination relevant drinking water conditions, Water Research (2014), doi: 10.1016/j.watres.2014.04.051. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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NH2Cl + H2O

HOCl + NH3

+ NH2OH

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+ NH2OH

NO–

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HNO

+NH2OH

+HNO

N2

N2O

NO2–/NO3–

+O2

NO2–/NO3–

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+O2

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ClNHOH

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Title: A proposed abiotic reaction scheme for hydroxylamine and

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monochloramine under chloramination relevant drinking water conditions

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3 Authors: David G. Wahman1*, Gerald E. Speitel Jr.2, and Madhav V. Machavaram3

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United States Environmental Protection Agency, Office of Research and Development, Cincinnati, OH 45268

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University of Texas at Austin, Department of Civil, Architectural and Environmental Engineering, Austin, TX 78712

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Pegasus Technical Services Inc., 26 W. Martin Luther King Drive, Cincinnati, OH 45268

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Corresponding author, mailing address: USEPA, 26 W. Martin Luther King Dr., Cincinnati, OH 45268. Phone: (513) 569–7733. Fax: (513) 487–2543. E–mail:

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[email protected]

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Abstract Drinking water monochloramine (NH2Cl) use may promote ammonia–oxidizing bacteria (AOB). AOB use (i) ammonia monooxygenase for biological ammonia (NH3) oxidation to

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hydroxylamine (NH2OH) and (ii) hydroxylamine oxidoreductase for NH2OH oxidation to nitrite.

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NH2Cl and NH2OH may react, providing AOB potential benefits and detriments. The

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NH2Cl/NH2OH reaction would benefit AOB by removing the disinfectant (NH2Cl) and releasing

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their growth substrate (NH3), but the NH2Cl/NH2OH reaction would also provide a possible

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additional inactivation mechanism besides direct NH2Cl reaction with cells. Because biological

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NH2OH oxidation supplies the electrons required for biological NH3 oxidation, the

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NH2Cl/NH2OH reaction provides a direct mechanism for NH2Cl to inhibit NH3 oxidation,

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starving the cell of reductant by preventing biological NH2OH oxidation. To investigate possible

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NH2Cl/NH2OH reaction implications on AOB, an understanding of the underlying abiotic

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reaction is first required. The present study conducted a detailed literature review and proposed

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an abiotic NH2Cl/NH2OH reaction scheme (RS) for chloramination relevant drinking water

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conditions (µM concentrations, air saturation, and pH 7–9). Next, RS literature based kinetics

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and end–products were evaluated experimentally between pHs 7.7 and 8.3, representing (i) the

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pH range for future experiments with AOB and (ii) mid-range pHs typically found in

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chloraminated drinking water. In addition, a 15N stable isotope experiment was conducted to

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verify nitrous oxide and nitrogen gas production and their nitrogen source. Finally, the RS was

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slightly refined using the experimental data and an AQUASIM implemented kinetic model. A

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chloraminated drinking water relevant RS is proposed and provides the abiotic reaction

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foundation for future AOB biotic experiments.

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Keywords: hydroxylamine; monochloramine; drinking water; nitrification; 15N; isotopes 2

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1.

Introduction Upon final implementation (i.e., ~2015) of the Stage 2 Disinfectants and Disinfection

Byproducts Rule, monochloramine (NH2Cl) use for secondary disinfection in the United States is

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predicted to increase to 57% of all surface and 7% of all ground water systems (USEPA 2005).

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Monochloramine use may promote nitrifying bacteria [i.e., ammonia–oxidizing bacteria (AOB)

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and nitrite–oxidizing bacteria] because of naturally occurring ammonia; residual ammonia

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remaining from initial NH2Cl formation; and ammonia released from NH2Cl decay, oxidation of

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natural organic matter, corrosion, pipe surface reactions, and nitrite (NO2–) oxidation under

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various conditions in chloraminated water systems (Kirmeyer et al. 2004, Wilczak et al. 1996).

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A very rapid NH2Cl residual loss is often associated with nitrification onset (American Water

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Works Association 2013) that may result in noncompliance with existing regulations (e.g.,

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Surface Water Treatment Rule); therefore, understanding nitrification and its control in drinking

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water distribution systems is of practical importance (Wilczak et al. 1996).

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The most studied AOB is the pure culture Nitrosomonas europaea. Figure 1 depicts N. europaea’s central metabolic pathway (Stein 1998). For biological ammonia oxidation to NO2–,

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N. europaea uses two enzymes in two reaction steps: (i) the membrane–bound ammonia

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monooxygenase (AMO) enzyme catalyzes free ammonia (NH3) oxidation to hydroxylamine

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(NH2OH) and (ii) the periplasmic–residing hydroxylamine oxidoreductase (HAO) enzyme

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catalyzes NH2OH oxidation to NO2– (Arp et al. 2002). For AMO, NH3 is the sole reductant [i.e.,

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electron (e–)] source for N. europaea through the subsequent oxidation of NH2OH (Figure 1).

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Two of the four electrons resulting from NH2OH oxidation are cycled back to AMO for NH3

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oxidation, while the other two electrons are used for other cellular processes (approximately 1.65

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electrons passing to the terminal oxidase for ATP generation and 0.35 passing to NAD+ to form

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NADH for biosynthesis) (Arp et al. 2002, Whittaker et al. 2000). NH2Cl inactivation studies on N. europaea have provided widely different estimates for

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inactivation rates based on the criterion used to define inactivation. For example, Oldenburg et

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al. (2002) reported N. europaea inactivation rates based on culturability were three orders of

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magnitude greater than those based on cell membrane integrity. In studies using NH2Cl

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application to a nitrifying biofilm, Lee et al. (2011) reported that NH2Cl application impacted

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biofilm metabolic activity [based on dissolved oxygen (O2) consumption] within 30 minutes;

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whereas, based on cell membrane integrity, minimal biofilm inactivation was seen at 2 hours. In

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general, inactivation rates increased in the following order: (i) cell membrane integrity

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(Oldenburg et al. 2002, Wahman et al. 2010, Wahman et al. 2009), (ii) culturability (Oldenburg

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et al. 2002), and (iii) metabolic activity (Lee et al. 2011, Pressman et al. 2012). Besides AOB

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inactivation from direct NH2Cl cellular reactions, another possible inactivation mechanism

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impacting metabolic activity would be the direct abiotic reaction of NH2Cl and NH2OH. NH2Cl

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and NH2OH are known to react (Giles 1999). Because biological NH2OH oxidation supplies the

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electrons required for biological NH3 oxidation, NH2OH’s reaction with NH2Cl provides a

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mechanism for NH2Cl to inhibit NH3 oxidation by competing with biological NH2OH oxidation

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and starving the cell of reductant. NH2Cl was recently shown to be biologically transformed by

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N. europaea (Maestre et al. 2013); therefore, it is reasonable to assume that NH2Cl can be

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present in the periplasm, providing an opportunity for the abiotic NH2Cl/NH2OH reaction.

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Alternatively, the NH2Cl/NH2OH reaction may benefit AOB by providing a mechanism of

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NH2Cl loss and NH3 release. To summarize, the abiotic NH2Cl/NH2OH reaction represents both

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a possible benefit, disinfectant removal and growth substrate release, and detriment, reductant

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source removal, to AOB. These competing impacts should be evaluated to understand the

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abiotic NH2Cl/NH2OH reaction’s potential importance on preventing or promoting drinking

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water distribution system nitrification. Before conducting biotic experiments with N. europaea to evaluate the possible relevance

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of the NH2Cl/NH2OH reaction, an abiotic model incorporating relevant NH2Cl and NH2OH

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reactions is required. To date, the abiotic NH2Cl/NH2OH reaction has been investigated by a

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few research groups (Aoki et al. 1989, Ferriol et al. 1986, Giles 1999, Robinson et al. 2005), but

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the conditions of this previous research were unrepresentative of chloraminated drinking water

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experiencing nitrification, including (i) mM versus µM reactant concentrations, (ii) pHs outside

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of 7–9, (iii) deoxygenated water versus air saturated water, or (iv) NH2OH in great excess

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relative to NH2Cl. In addition, two additional competing pathways for NH2OH may be relevant

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under chloramination conditions and should be considered: reaction with (i) hypochlorous acid

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(HOCl) (Giles 1999) released from NH2Cl hydrolysis and (ii) O2 (Anderson 1964, Hughes and

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Nicklin 1971, Kono 1978, Moews Jr and Audrieth 1959, Yagil and Anbar 1964). The HOCl

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reaction can be considered through modeling as was done previously for the reaction of NH2Cl

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and HOCl with NO2– (Wahman and Speitel 2012), and the O2 reaction can be evaluated in

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O2/NH2OH control experiments.

The current study represents a first step in evaluating the abiotic NH2Cl/NH2OH reaction

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importance on AOB by proposing and validating an abiotic NH2Cl/NH2OH reaction scheme

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incorporated into a well-established chloramine chemistry model (Jafvert and Valentine 1992,

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Vikesland et al. 2001, Wahman and Speitel 2012). First, a detailed literature evaluation was

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conducted to propose a relevant reaction scheme for the abiotic NH2Cl/NH2OH reaction under

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chloraminated drinking water relevant conditions, including incorporation of revised nitroxyl 5

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(HNO) chemistry and possible additional end-products. Next, the proposed reaction scheme was

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implemented into a chloramine chemistry model in AQUASIM and evaluated using

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experimental data from abiotic batch kinetic experiments at pHs between 7.7 and 8.3, covering

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the pH range (i) in future N. europaea biotic experiments and (ii) representing typical mid-range

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pHs found in chloraminated drinking water. In addition, a 15N stable isotope experiment was

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conducted to verify nitrous oxide (N2O) and nitrogen gas (N2) production and their nitrogen

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source. Finally, the reaction scheme was slightly refined using the experimental data and the

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AQUASIM implemented kinetic model, providing the abiotic foundation for a future model

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incorporating N. europaea’s biotic reactions.

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2.

Materials and Methods

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2.1.

Reagent preparation

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Solutions were prepared in ultra–pure water (Barnstead NANOpure Diamond). Stock NH2OH solutions were prepared from reagent grade hydroxylamine–hydrochloride. Stock

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chlorine solutions were prepared by diluting 4–6% sodium hypochlorite and were standardized

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periodically by Standard Methods 4500B (American Public Health Association 1998). Stock

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TOTNH3 [sum of ammonia (NH3–N) and ammonium (NH4+–N)] solutions were prepared by

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dissolving ammonium sulfate in ultra–pure water (pH 8.3). Stock NH2Cl solutions were

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prepared by additions of stock TOTNH3 solutions to ultra–pure water and then adding an aliquot

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of the stock chlorine solution to this well–stirred TOTNH3 solution [pH > 8.3, 4:1 chlorine to

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nitrogen (Cl2:N) mass ratio]. The NH2Cl stock solution was allowed to mix for 15 minutes

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before use, and scans of NH2Cl stock solutions were conducted on a Nicolet Evolution 300 UV–

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visible spectrophotometer (Thermo Electron Scientific Instruments) to verify only NH2Cl

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formation.

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2.2.

Batch kinetic experiments Two types of batch kinetic experiments were conducted: (i) O2/NH2OH kinetic control

experiments and (ii) NH2Cl/NH2OH kinetic experiments. In general, batch kinetic experiments

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were conducted at room temperature (22±1 °C) in air saturated 4 mM sodium bicarbonate

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buffered ultra–pure water initially contained in a well-mixed Erlenmeyer flask covered with

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aluminum foil. Subsequently, a spike of the appropriate aliquot of NH2Cl stock solution to

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achieve the desired NH2Cl concentration was added (omitted for O2/NH2OH control

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experiments). Before NH2OH addition, initial samples verified experimental conditions. Next,

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the appropriate amount of NH2OH stock solution was added to start an experiment, samples were

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taken, and this solution was subsequently placed in head–space–free 500–mL, glass, gas–tight

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syringes (VICI Precision Sampling) with magnetic stir bars for mixing. Before each experiment,

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syringes were made chlorine–demand–free by soaking in a 5,000 mg Cl2 L–1 free chlorine

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solution for 24 hours, rinsed with distilled water, and air dried. Syringes contained small

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Teflon–coated stir bars for mixing and were wrapped in aluminum foil. Subsequently, samples

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for NH2Cl, TOTNH3, NH2OH, NO2–, nitrate (NO3–), N2O, temperature, and pH were temporally

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collected by depressing the gas-tight syringe plunger to maintain head-space-free conditions

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The O2/NH2OH reaction was evaluated using five O2/NH2OH control experiments. Three experiments (C–1 through C–3) were conducted at pHs (7.7, 8.2, and 8.3) representative

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of those used in the NH2Cl/NH2OH kinetic experiments. Because the O2/NH2OH reaction rate

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increases with pH, two additional experiments (C–4 and C–5) were conducted at elevated pHs

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(9.0 and 9.1), representing an extreme condition to evaluate the reaction relevance.

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The NH2Cl/NH2OH reaction scheme was evaluated using seven experiments (H–1 through H–7) conducted between pHs 7.7 and 8.3, representing the pH range for (i) future N. 7

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europaea biotic experiments and (ii) mid-range chloraminated drinking water pHs. These

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experiments sought to validate abiotic model implementation and provide experimental data to

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refine literature estimates of poorly or variably defined literature rate constants and verify

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proposed end-product production. The conditions used in these experiments are by no means

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meant to be exhaustive; care should be taken in using the resulting model at conditions (e.g., pH,

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Cl2:N mass ratio, temperature) beyond those used in the current research without further

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validation.

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2.3.

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N stable isotope experiment and analysis

We used 14N and 15N stable isotopes to analyze gaseous end-products of the proposed

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NH2Cl/NH2OH reaction scheme, verifying the production of and nitrogen source in N2O and N2.

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A batch experiment was conducted in triplicate using 14NH2Cl made as previously described in

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2.1 Reagent preparation, except for using 14N ammonium sulfate (δ15N = -968‰) as the

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ammonia source and using a 15N enriched NH2OH (δ15N = +955‰) made with 15N

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hydroxylamine-hydrochloride. For this experiment, aluminum foil wrapped 1-L tedlar bags

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(AKC) were filled with 750-mL ultra–pure water (Barnstead NANOpure Diamond) and purged

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with helium gas to substantially reduce background N2 concentrations and fill the headspace.

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Subsequently, concentrated solutions (36 mL total volume) were added in quick succession and

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the bags shaken vigorously to avoid localized plumes and create initial concentrations of 35.5 mg

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Cl2 L–1 (500 µM) NH2Cl and 500 µM NH2OH in a 4 mM sodium bicarbonate buffer (pH

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8.0±0.3) at room temperature (22±0.5 °C). These additions resulted in a 786 mL liquid volume

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and 214 mL headspace in the tedlar bags. Residual N2 and N2 added from solution injections

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were accounted for in subsequent data analysis. After a two-hour reaction (i.e., reaction

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completion), headspace samples were taken to quantify N2O and N2, and liquid samples were

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taken to measure other final products per the batch kinetic experiments. For N2 and N2O gas measurements, headspace gases were injected to a modified

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Elemental Analyzer (EA) coupled to an Isotope Ratio Mass Spectrometer (IRMS) for measuring

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the δ15N stable isotope values (Machavaram et al. 2013). Briefly, the EA was fitted with a

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device that allowed direct injection of gases into the EA and a modified packed column (3.17

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mm x 2m) that allowed accurate detection and measurement of gas samples in nanomolar

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concentrations. From each tedlar bag, multiple aliquots were injected to determine 15N/14N

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isotope ratios of N2 and then N2O by setting up the EA-IRMS accordingly for each gas species.

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The resulting isotope ratios were expressed in standard δ notation as permil (‰) values.

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2.4.

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TOTNH3 and pH were measured on a Model 250 pH/ISE/conductivity meter with an ammonia and pH electrode (Denver Instrument), respectively. NH2OH (H–1, H–2, H–5, and H–

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6), NH2Cl, and NO2– were measured on a Nicolet Evolution 300 UV–visible spectrophotometer

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at 705 nm (Frear and Burrell 1955), 655 nm (HACH Method 10171), and 507 nm (HACH

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Method 8507), respectively. Possible interferences between the NH2OH and NH2Cl methods

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were investigated and are discussed in Appendix – Supplementary Information (SI), Method

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Development (Figures S1–S5). In select NH2Cl/NH2OH kinetic experiments (H–1, H–2, and H–

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6), N2O was measured using an N2O microelectrode (UNISENSE) calibrated with N2O standards

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(SI, Nitrous oxide microelectrode calibration curve, Figure S6) created from dilutions of N2O

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saturated water (Weiss and Price 1980). At experiment end and in real–time for select

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NH2Cl/NH2OH kinetic experiments (H–2 and H–7), NO2– and NO3– were analyzed by ion

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chromatography using EPA method 300.0. Dissolved oxygen was measured with a WTW Multi

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340i Oxygen Meter and WTW CellOx 325 Oxygen Probe (Weilheim) per manufacturer’s

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instructions.

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2.5.

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2.5.1. Reaction rate expressions and stoichiometry

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Model implementation

The chloramine model presented by Wahman and Speitel (2012) and implemented in the computer program AQUASIM (Reichert 1994) was expanded to include the NH2OH relevant

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reactions and rate constants summarized in Table 1 that were taken from the literature as noted.

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The additional proposed NH2OH relevant reactions are also schematically in Figure 2. A

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detailed description of these added reactions is presented in 3.1 Reaction scheme development

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and model implementation.

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2.5.2. Model parameter estimation

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Because experimental conditions were specifically chosen to represent relevant drinking

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water conditions and the reaction scheme is complex, pseudo–first–order assumptions were not

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valid. To estimate parameters in this nonlinear system, all available experimental data (N=150)

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was simultaneously fit using measured concentrations (NH2Cl, NH2OH, NO2–/NO3–, and N2O),

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and the parameter estimation function in AQUASIM was configured to minimize the weighted

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residual sum of squares (WRSS) between measurements and calculated model results (Equation

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1):

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y , − y  y , − y  (1) WRSS =    =  W y ,

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In Equation 1, ymeas,i is the i–th chemical measurement, W is the weighting factor, and yi

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is the model simulated chemical concentration corresponding to the i–th chemical measurement. 10

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Because chemical measurements were changing over an order of magnitude, ymeas,i was

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implemented for W to prevent higher concentrations from biasing the fitting procedure, resulting

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in a dimensionless WRSS (Draper and Smith 1998, Robinson 1985).

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3.

Results and Discussion

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3.1.

Reaction scheme development and model implementation

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The initial step in the NH2OH related reaction scheme (Figure 2) is either the reaction of NH2Cl or HOCl with NH2OH. For both of these reactions, three possible reaction pathways

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exist (Giles 1999), but under the pHs used in this research (pH 1 in Table 3) . Comparing the experiments conducted as pH increased,

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∆$ %&$

∆ 

, Table 3) increased and N2O (∆  , Table

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with respect to NH2OH of NO2–/NO3– (

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3) decreased as the pH increased. Based on the reaction scheme, this effect is expected with

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increasing pH, resulting in an increased conversion of HNO to NO– (R4, Table 1) and

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subsequent oxidation to NO2–/NO3– (R6, Table 1) versus HNO dimerization (R8, Table 1).

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3.2.3. Stable isotope experiment.



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To verify the gaseous end-products (N2O and N2) and their source, a triplicate stable

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isotope experiment was conducted. Please refer to SI (Stable Isotope Experiment Analysis,

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Tables S3 through S6) for a detailed account of these experimental results. These results

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confirmed (i) N2 and N2O formation, (ii) N2 results from NH2OH, and (iii) the majority of N2O

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(87±0.45%) resulted from NH2OH. A small contribution of N2O and N2 from NH2Cl

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autodecomposition is not unexpected at the elevated NH2Cl concentration (500 µM) used in this

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experiment. Regardless, these results confirm that the major pathways for N2O and N2

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generation are from NH2OH. The N2O result confirms the result from Robinson et al. (2005),

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and the N2 result represents the first confirmation of its source and end-product measurement.

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3.3.

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Model simulations with published rate constants.

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To assess the proposed reaction scheme, the implemented model was used to simulate the seven kinetic experiments (Table 3) with the published rate constants and their reported

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uncertainty (Table 1). For the reactions (R1 and R9, Table 1) with multiple estimates for their

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rate constants (k1 and k9), the values reported by Giles (1999) and Liochev and Fridovich (2003)

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were used, respectively. Because of their speculative nature in the literature, two rate constants

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(k7 and k9) did not have published standard deviations (R7 and R9, Table 1); therefore, 10% of

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their reported value was used as an estimate of their standard deviation. The error bounds on the

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simulations were extremely narrow and are not shown in Figure 4 for clarity. In general, these

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simulations showed that (i) NH2Cl loss was simulated well, (ii) NH2OH loss was underpredicted,

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(iii) NO2–/NO3– generation was overpredicted, and (iv) N2O generation was underpredicted. Based on the uncertainty in the published rate constants, three rate constants (k1, k7, and

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k9) were selected for refinement. First, the Giles (1999) rate constant for R1 (k1=1.1 x 109 M–2 s–

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uncertainty for k1. Second, for N2 and NO2–/NO3– generation and as previously mentioned, k7

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and k9 are not well established in the literature and were also selected for refinement.

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3.4.

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Model parameter estimation and simulations

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) lies in the middle of the range reported (k1=0.46–7.58 x 109 M–2 s–1), detailing the reported

Parameter estimation was conducted for the rate constants of R1, R7, and R9, and initial NH2Cl and NH2OH concentrations using the entire data set. The data and model simulations are

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shown in Figure 5 (best–fit only) and SI, Figure S7 (best–fit and associated error bounds). The

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best–fit rate constants are provided in Table 1 along with their 95% confidence limits. The

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reaction scheme simulated the measured chemical concentrations well for the experimental

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conditions, representing a reasonable representation of the abiotic reactions under the conditions

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evaluated.

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A relatively small range of experimental conditions were examined as part of this research as the focus was on validating an abiotic model for use in future biotic experiments to

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be conducted under similar experimental conditions. As mentioned previously, care should be

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taken in using the resulting abiotic model at conditions greatly beyond those used in the current

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research without further validation.

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4.

Conclusions •

The NH2Cl/NH2OH reaction represents an abiotic reaction with potential competing impacts to AOB. The reaction would benefit AOB by removing the disinfectant

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(NH2Cl) and releasing growth substrate (NH3), but it would be detrimental by

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removing NH2OH without generating reductant to sustain ammonia oxidation. A

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detailed literature evaluation was conducted and a relevant reaction scheme for the

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abiotic NH2Cl/NH2OH reaction under relevant chloraminated drinking water

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conditions (i.e., µM reactant concentrations, air saturation, and pH 7–9) was proposed

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and validated.

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Based on O2/NH2OH control experiments, the direct O2/NH2OH reaction was

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sufficiently slow to be ignored at these experimental conditions (pH, NH2OH and O2

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concentrations, temperature, and experiment duration). •

The proposed abiotic NH2Cl/NH2OH reaction scheme was added to a drinking water

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chloramine chemistry model implemented into AQUASIM, evaluated, and refined

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between pHs 7.7 and 8.3 using experimental data. •

predicted by the model, and based on 15N isotopic mass balance the source of N2O

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All proposed reaction products were measured at concentrations similar to those

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and N2 was verified to be predominantly from NH2OH, providing further support for

the reaction scheme.

•

The proposed abiotic NH2Cl/NH2OH reaction scheme provides the foundation for a

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future model incorporating AOB biotic reactions to evaluate the importance of the

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NH2Cl/NH2OH reaction to AOB and chloramine loss. 16

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Appendix – Supplementary information

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Supplementary information related to this article is provided online.

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Acknowledgment

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The authors thank Keith Kelty, David Griffith, Michael Elovitz, Toby Sanan, and Jacob Botkins. The USEPA collaborated in the research described herein. It has been subjected to the

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Agency’s peer and administrative review and has been approved for external publication. Any

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opinions expressed are those of the authors and do not necessarily reflect the views of the

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Agency; therefore, no official endorsement should be inferred. Any mention of trade names or

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commercial products does not constitute endorsement or recommendation for use.

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References

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American Public Health Association (1998) Standard Methods for the Examination of Water and Wastewater, American Public Health Association, Washington, D.C.

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American Water Works Association (2013) Nitrification Prevention and Control in Drinking Water (AWWA Manual M56), American Water Works Association, Denver, Colorado.

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Anderson, J.H. (1964) The copper-catalysed oxidation of hydroxylamine. Analyst 89(1058), 357-362.

368 369

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Stein, L.Y. (1998) Effects of Ammonia, pH, and Nitrite on the Physiology of Nitrosomonas europaea, an Obligate Ammonia-oxidizing Bacterium. Ph.D. Dissertation, Oregon State University, Corvallis, OR.

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Yagil, G. and Anbar, M. (1964) The formation of peroxynitrite by oxidation of chloramine, hydroxylamine and nitrohydroxamate. Journal of Inorganic and Nuclear Chemistry 26(3), 453460.

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Table 1. Monochloramine and hydroxylamine reaction summary.

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Table 2. Reaction stoichiometry for various nitrogen products.

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Table 3. Summary of hydroxylamine and monochloramine batch kinetic experiment nominal conditions and measured molar delta concentration ratios at experiment end. All experiments conducted at room temperature (22±1 °C) and in air saturated 4 mM carbonate buffer. Monochloramine was made at a 4:1 chlorine to nitrogen mass ratio.

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Figure 1. Central metabolism of N. europaea (Stein 1998, Ward et al. 2011).

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Figure 2. Monochloramine and hydroxylamine reaction scheme. Reaction numbers (R#) correspond to those provided in Table 1. Ammonium, chloride ion, hydrogen ion, hydroxide ion, and water not shown for clarity (refer to Table 1 for complete reactions).

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Figure 3. Hydroxylamine concentrations for the hydroxylamine and dissolved oxygen control experiments. All experiments conducted at room temperature (22±1 °C) and in air saturated 4 mM carbonate buffer.

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Figure 4. Comparison of monochloramine (A), hydroxylamine (B), nitrite plus nitrate (C), and nitrous oxide (D) experimental data and model simulations using published rate constants (Table 1). Initial conditions: 4 mM carbonate buffer, 4:1 Cl2:N mass ratio, and all other conditions detailed in Table 3.

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Figure 5. Comparison of monochloramine (A), hydroxylamine (B), nitrite plus nitrate (C), and nitrous oxide (D) experimental data and model simulations using published (reactions 2, 4, 5, 6, and 8) and revised (reactions 1, 7, and 9) rate constants (Table 1). Initial conditions: 4 mM carbonate buffer, 4:1 Cl2:N mass ratio, and all other conditions detailed in Table 3.

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Rate Constants ii

#

Reaction Stoichiometry

Rate Expression i

Published

Current

Research Estimate

Units

a

RI PT

(22±1°C)

(7.58±0.53)x109; 25°C b



R1 H  + NH Cl + NH OH → NH  + ClNHOH

k H NH ClNH OH

(1.8±0.10)x109 M–2s–1

c

R3

ClNHOH  Cl + HNO + H 

R4

HNO + OH → NO + H O

R5

NO + H O → HNO + OH 

R6

 NO + O → ONOO  NO  /NO

R7

HNO + NH OH → N + H O

R8

2HNO → HONNOH  N O + H O









!



&

R9

k HNOOH 



HNO + O → X 

c

(1.35±0.05)x107; 25°C

N/D

M–1s–1

e

N/D

M–1s–1

1.2x102; 22±2°C

N/D

s–1

k NO O 

(2.7±0.2)x109; 22±2°C

N/D

M–1s–1

k " HNONH OH

 NO  /NO

k % HNO

Assumed Fast

(4.9±0.5)x104; 22±2°C e

k  NO 

EP

$

(0.94±0.05)x109; 25°C

c

TE D



k  HOClNH Cl

(1.1±0.05)x10 ; 25°C

M AN U



HOCl + NH OH → H O + ClNHOH

9

SC

d

R2

0.46 x 109

e

f

4x103; pH 7.4; 23°C e

(8±3)x106; 22±2°C

(1.2±0.27)x104 M–1s–1

N/D

M–1s–1

g

3x103; 37°C

k ( HNOO 

where X is unknown intermediate

h

8x103; 23°C

AC C

e

(3.8±0.62)x102 M–1s–1