Aldehydes to the Optical

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Contribution of quinones and ketones/aldehydes to the optical properties of humic substances (HS) and chromophoric dissolved organic matter (CDOM) Rossana Del Vecchio, Tara Marie Schendorf, and Neil V. Blough Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.7b04172 • Publication Date (Web): 10 Nov 2017 Downloaded from http://pubs.acs.org on November 12, 2017

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Environmental Science & Technology

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Contribution of quinones and ketones/aldehydes to the optical properties of humic

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substances (HS) and chromophoric dissolved organic matter (CDOM).

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Rossana Del Vecchio,1,* Tara Marie Schendorf2 and Neil V. Blough2,*

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20742, United States

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submitted to Environmental Science & Technology

Earth System Science Interdisciplinary Center, University of Maryland, College Park, Maryland

Department of Chemistry and Biochemistry, University of Maryland, College Park, MD, USA

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Corresponding authors: Rossana Del Vecchio and Neil V. Blough

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Rossana Del Vecchio

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email: [email protected], telephone: 001 301-405-0337

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Neil Blough

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email: [email protected], telephone: 001 301-405-0051

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Keywords: Quinones, ketones, aldehydes, HS, CDOM, optical properties, chemical structures, borohydride reduction, dithionite reduction, charge-transfer model.

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Number of pages: 31

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Number of figures: 5

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Number of tables: 0

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Number of words: 5470

ACS Paragon Plus Environment

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Abstract

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The molecular basis of the optical properties of chromophoric dissolved organic matter (CDOM)

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and humic substances (HS) remains poorly understood and yet to be investigated adequately.

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This study evaluates the relative contributions of two broad classes of carbonyl-containing

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compounds, ketones/aldehydes versus quinones, to the absorption and emission properties of a

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representative suite of HS as well as a lignin sample. Selective reduction of quinones to

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hydroquinones by addition of small molar excesses of dithionite to these samples under anoxic

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conditions produced small or negligible changes in their optical properties; however, when

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measurable, these changes were largely reversible upon exposure to air, consistent with the re-

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oxidation of hydroquinones to quinones. With one exception, estimates of quinone content based

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on dithionite consumption by the HS under anoxic conditions were in good agreement with past

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electrochemical measurements. In contrast, reduction of ketones/aldehydes to alcohols

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employing excess sodium borohydride produced pronounced and largely, but not completely,

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irreversible changes in the optical properties. The results demonstrate that (aromatic)

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ketones/aldehydes, as opposed to quinones, play a far more prominent role in the optical

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absorption and emission properties of these HS, consistent with these moieties acting as the

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primary acceptors in charge-transfer transitions within these samples. As a method, anoxic

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dithionite titrations may further allow additional insight into the content and impact of

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quinones/hydroquinones to the optical properties of HS and CDOM.


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Introduction The optical absorption and emission properties of humic substances (HS) and

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chromophoric dissolved organic matter (CDOM) have been studied extensively for more than 50

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years, yet an understanding of the molecular basis of these properties remains incomplete.

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Recent work has provided evidence that the optical properties of HS and CDOM cannot arise

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solely from a simple superposition of numerous independently absorbing and emitting

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chromophores.1-3 Instead, electronic interactions between chromophores within HS and CDOM

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have been proposed to play an important role.2-5 According to this model, the optical properties

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result in part from charge transfer (CT) interactions between electron donor moieties (D) (e.g.,

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phenolic and possibly substituted aromatic carboxylic acid groups)6-8 and electron acceptor

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moieties (A) (e.g., carbonyl-containing groups such as aromatic ketones/aldehydes and quinones)

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formed through the partial oxidation of lignin,2, 3, 5 and possibly other hydroxy-aromatic

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precursors.3, 5 Our working structural model envisions these donor/acceptor pairs to be in close

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contact, helping to stabilize the formation of an ensemble of higher molecular weight

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supramolecular aggregates9, 10 in a fashion very similar to that described by Dreyer et al.11 for

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closely related materials.12 Further, similar to the model suggested by Latch and McNeill,13 outer

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charged groups (carboxylate anions) would be expected to encompass an inner core14, 15 in which

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the contact DA complexes form and are further stabilized. The long-wavelength visible

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absorption and emission would thus arise from CT transitions between these contact DA pairs,

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whereas the ultraviolet absorption and near-visible emission would arise principally from the

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aromatic structures known to be a part of lignin (e.g. local donor and acceptor states).2-5

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The presence of electron donor and acceptor moieties within HS and CDOM is now well established based on extensive electrochemical measurements,16-18 acid/base titrimetric


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measurements,7, 19, 20 as well as combined optical, (electro)chemical, photochemical and

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structural measurements.4, 21-34 These studies have provided evidence of phenolic electron donors

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as well as quinoid and aromatic ketone/aldehyde electron acceptors. These donors and acceptors

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have been linked in part to the optical properties of the HS through changes in absorption and

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emission produced by altering the abundance or form of the electron donors/acceptors through

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chemical, photochemical or electrochemical treatments.4, 5, 22, 26-34 However, while these studies

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have provided strong evidence that HS and CDOM contain ketone/aldehyde and quinone

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acceptors, their relative contribution to the absorption and emission properties of HS as well as

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their importance as possible electron acceptors in CT transitions has not been well studied and

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has, in fact, been the subject of some controversy.35 Although some studies have suggested that

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reduction of quinones in HS causes minor and mostly reversible changes in fluorescence

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emission,34, 36, 37 others have claimed significant changes in the fluorescence emission,38 with

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further claims that the fluorescence emission arises primarily from the quinone/hydroquinone

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moieties38, 39 and can be used to deduce the redox state of natural organic matter.40, 41 However, it

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was later shown that the quantum yields of fluorescence for an extensive series of structurally-

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diverse quinones were exceedingly small or negligible in water4, 35 and further, that changes in

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the fluorescence emission upon reduction and re-oxidation of putative quinones within HS were

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also small or negligible.34, 42, 43 In contrast, it has been repeatedly shown that chemical reduction

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with sodium borohydride produces substantial losses in absorption as well as significantly

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enhanced, blue-shifted fluorescence emission.4, 22, 26, 27, 44-48 These substantial changes in optical

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properties were observed to be largely irreversible upon exposure to air, consistent with the

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reduction of ketones/aldehydes to alcohols4, 26, 46 and not of quinones to hydroquinones.


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The relative importance of quinones and ketones/aldehydes to the absorption and emission

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properties of HS and CDOM can be assessed by selectively eliminating these moieties through

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reduction and monitoring the changes in the optical properties as done previously with sodium

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borohydride.4, 46 Reduction of quinones in HS has been achieved chemically,49, 50

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electrochemically16, 34, 36 and microbially.37, 51 Among the chemical reductants is sodium

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dithionite (Na2S2O4), which, at small stoichiometric excesses under anoxic conditions, rapidly

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reduces quinones in water by electron-transfer.52 Quinone moieties in HS and CDOM can also be

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selectively reduced to their respective hydroquinones by direct electrochemical reduction on

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glassy carbon electrodes polarized at low reduction potentials.16, 17 However, despite being

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highly selective, the electrochemical reductions require access to electrochemical equipment,

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need to be carried out in anoxic glove boxes to eliminate interference from O2, and require hours

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to days to complete.16, 17 Quinones are also thought to be reduced microbially;37, 51 microbial HS

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reduction of quinones by Geobacter metallireducens has been suggested to induce red-shifted

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fluorescence emission in the HS.37 However, these biological reductions are far less easy to

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control than either chemical or electrochemical reductions. Because of these factors, dithionite

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was employed as a selective reductant of quinones/hydroquinones in this study.

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Sodium borohydride can be employed to reduce ketones/aldehydes to form the

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corresponding alcohols.4, 21, 22, 25-27, 44, 46, 53 Borohydride, however, also reduces quinones to

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hydroquinones,46 but most hydroquinones can be reversibly oxidized to quinones by O2 under

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aerobic conditions, unlike the alcohols that are stable to air oxidation.4 Thus, additional

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information on the contribution of quinones to the optical properties of HS and CDOM samples

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can be obtained by examining the reversibility of the optical changes after re-oxidation in air of

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either dithionite or borohydride reduced samples.46


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In this work, the relative contributions of carbonyl-containing compounds to the optical

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properties of HS and CDOM was probed through the selective reduction of quinone and

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ketone/aldehyde moieties in a representative group of HS (including aquatic, microbial and soil

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HS), as well as through an examination of the reversibility of the optical changes following their

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aerobic re-oxidation. This work was complemented by a comparison of the selectivity of

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dithionite and borohydride for the reduction of quinone and aromatic ketone model compounds.4

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Selective reduction of quinones to hydroquinones within the HS employing dithionite produced

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small or negligible changes in absorption and emission relative to those produced by borohydride

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reduction; when measurable, however, these changes were largely reversible upon exposure to

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air consistent with the re-oxidation of hydroquinones to quinones. Conversely, reductions

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employing excess sodium borohydride caused pronounced and largely, but not completely,

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irreversible changes in the optical properties. The results reveal a prominent contribution of

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(aromatic) ketones/aldehydes, as opposed to quinones, to the optical properties of these HS,

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consistent with these moieties acting as the primary acceptors in CT interactions.

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Material and Methods

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Materials. Suwannee River Humic Acid Standard II (SRHA) (lot # 2S101H) and Fulvic

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Acid Standard (SRFA) (lot # 1S101F), Pony Lake Fulvic Acid Reference (PLFA) (lot #

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1R109F), Leonardite Humic Acid Standard (LHA) (lot # 1S104H-5) and Elliot Soil Humic Acid

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Standard (ESHA) (lot # 1S102H) were obtained from the International Humic Substances

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Society (IHSS) and used as received. The properties of these HS can be found at the IHSS web

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site (http://humic-substances.org/). Alkali-extracted and carboxylated lignin (LAC; Lot 19714

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DS) was obtained from Sigma-Aldrich and used as received. Model compounds included

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aromatic ketones and quinones from Sigma Aldrich and Fluka. The quinones included p-


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benzoquinone, 2-methyl-p-benzoquinone, 2,5-dimethyl-p-benzoquinone, 4-methyl-p-

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benzoquinone, 2,5 dimethoxy p-benzoquinone, 2,3-dimethoxy-5-methyl-p-benzoquinone, tetra-

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hydroxy-p-benzoquinone, 4-t-butyl-5-methoxy-1,2-benzoquinone, 3,5-di-t-butyl-1,2-

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benzoquinone, anthraquinone 1,5 disulfonic acid disodium salt, 1,4-naphthoquinone, 2-hydroxy-

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naphthoquinone, 5-hydroxy-naphthoquinone and 5,8-dihydroxy-naphthoquinone. These

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compounds were further purified by sublimation or recrystallization as previously reported.4 The

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aromatic ketones included benzophenone (purity > 99%), acetophenone (> 98%), 3-methoxy

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acetophenone (purity > 97 %), and 2-acetonapthone (99% purity) and were used as received.

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Sodium borohydride (NaBH4) was obtained from Fisher, while disodium dithionite (85 %)

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(Na2S2O4) was from Acros. Water was obtained from a Milli-Q (MQ) Plus purification system

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(Millipore). Sodium hydroxide and hydrochloric acid were obtained from Fluka.

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Apparatus. A Shimadzu UVPC 2401 spectrophotometer was employed to acquire UV-

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visible absorption spectra. An Aminco-Bowman AB-2 luminescence spectrometer was employed

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for the fluorescence measurements. A Thermoscientific pH meter was employed for the pH

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measurements.

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Optical Measurements. Absorption spectra were recorded using 1-cm quartz cuvettes

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over the range from 190 to 820 nm against an air reference; a MQ versus air blank was

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subtracted afterward. The absorption changes upon treatment were defined as follows: the

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fractional absorbance change (Af) upon treatment were defined as Af = A(treated)/A(untreated).

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Fluorescence emission spectra were collected over the 300-600 nm excitation range with

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the absorbance kept at < 0.05-0.1 OD at the excitation wavelength.3 Monochromator excitation

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and emission band-passes were set to 4 nm. A MQ blank was subtracted from the emission


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spectra. The blank-subtracted emission spectra were then corrected for instrument response by

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applying the correction factors provided by the manufacturer.

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Chemical reduction of Model Compounds and HS. Dithionite reduction. A stock solution

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of Na2S2O4 was prepared by sparging a 3 mL solution of 0.025 N NaOH with N2 for 30 min,

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followed by addition of a known mass (~ 5 mg) of solid Na2S2O4 ([Na2S2O4] ≈ 9.5 mM). The

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headspace of the sealed vial containing the dithionite solution was continuously flushed at a slow

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flow rate with N2, which was vented through a fine needle in the gas-tight seal. O2 scrubbers

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were installed along the N2 lines to further minimize any traces of O2. Known concentrations (as

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reported in figures) of SRFA, SRHA, PLFA, LHA, ESHA, and LAC in MQ water at pH 7 or 10

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and model compounds (quinones, ~0.1-5 mM and ketones, ~2-50 µM) in MQ water at neutral

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pH were transferred to a 1-cm cuvette (3 mL) and purged with ultrapure N2 for 30 min prior to

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reduction. All HS and lignin solutions were 0.2 µm filtered prior to measurement and subsequent

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reductions. The headspace of the sealed cuvette was then flushed continuously with N2 at a slow

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flow rate. Using gas-tight syringes, known volumes of the stock dithionite solution were

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transferred anaerobically to the sealed 1-cm cuvettes (3 mL) containing the N2-purged HS or

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model compounds. Dithionite was added to these samples in stoichiometric excess,

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approximately 2- to 5-fold higher in molar concentration than those of model compounds and the

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quinone content of the HS as measured electrochemically16 (see further below). These excesses

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were added to ensure rapid and complete reduction. Samples were reduced for 15 minutes under

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anoxic conditions in the dark. For the pH 10 reductions, the HS were then re-oxidized for 24 h in

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the dark following introduction of air into the solutions. The optical properties of the samples

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were measured prior to and following the reduction and subsequent re-oxidation in air. No


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changes in pH were observed during reduction/re-oxidation. The effect of initial pH on the

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reduction of HS was investigated by reducing samples at both pH 7 and pH 10.

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Quinone content in the HS was determined spectrophotometrically by measuring the

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consumption of dithionite in anoxic HS solutions. Identical known volumes of the dithionite

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stock solution were anaerobically transferred using gas-tight syringes to 1-cm cuvettes (3 mL)

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containing either N2-purged HS (100 mg/L) or N2-purged MQ water (blank). The absorbance at

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316 nm and dithionite molar absorption coefficient at this wavelength (ε316=8000 M-1cm-1)54, 55

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were employed to calculate the amount of dithionite in the blank and that of the unreacted

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dithionite in the HS solution. The concentration of unreacted dithionite in the HS solution was

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subtracted from the dithionite concentration in the blank to provide the concentration of

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dithionite consumed in the HS solution by quinone reduction. No significant losses in absorption

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of the HS or LAC samples were observed over the dithionite absorption band following

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reductions (see Results). Six independent determinations of dithionite consumption were

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performed for each sample examined. The dithionite consumed provides a measure of the

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quinone content in each HS based on a 1:1 stoichiometry (one mole of dithionite (two reducing

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equivalents) converts one mole quinone to one mole hydroquinone).

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Borohydride reduction. A stock solution of NaBH4 was prepared by dissolving ~70 mg of

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solid NaBH4 in 1 mL of MQ water at pH 12 pre-flushed with N2 for 30 min, thus providing a

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stock solution of [NaBH4] ≈ 1.85 M. O2 scrubbers were installed along the N2 lines to further

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minimize traces of O2. Known concentrations (as reported in figures) of SRFA, SRHA, PLFA,

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LHA, ESHA, and LAC in MQ water at pH 10 and model compounds (aromatic ketones, ~2-50

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µM) in MQ water at neutral pH were transferred to a 1cm cuvette (3 mL) and purged with

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ultrapure N2 for 30 min prior to reduction. All HS and the lignin samples were 0.2 µm filtered


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prior to measurement and subsequent reduction. Quinones were shown previously to be reduced

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rapidly in water by borohydride;4 the absorption and emission properties of the

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quinones/hydroquinones are reported in that paper4 and elsewhere.52 Reductions of the HS were

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conducted with a large excess of NaBH4 relative to the HS to ensure a significant degree of

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reduction of ketones/aldehydes46 as well as the quinones. Experiments employing approximately

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stoichiometric amounts of NaBH4 relative to the quinone content of the HS16 were not performed

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because NaBH4 is not completely selective and will reduce both quinones and

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ketones/aldehydes, and further because unknown amounts of the NaBH4 will be consumed by

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reaction with water.56

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Reduction of the ketone model compounds were conducted with minimally a 10-fold molar

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excess of NaBH4 relative to ketone concentration. For the HS reductions, a known volume of the

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NaBH4 stock was anaerobically transferred to the cuvettes containing HS (80 µL of NaBH4 stock

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to 3 mL of HS, 100 mg/L), thus providing a ~20-fold mass excess of NaBH4 relative to HS (~

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12-fold molar excess relative to C). The cuvettes were kept under N2 for a period of 24 to 48 h,

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following the procedures of Ma et al and Schendorf et al.4, 46 The reduced samples were

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subsequently re-oxidized in air for 24 h. Samples were treated in the dark, and the optical

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properties recorded prior to and following reduction, and subsequently, following re-oxidation in

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air. Samples were reduced with NaBH4 at a basic pH ~10 to minimize the reaction with H3O+

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and thus the loss of NaBH4 through this pathway as well as the concomitant increase in solution

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pH during the reduction.4, 46 To compare the effects of borohydride reduction to those of

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dithionite reduction at pH 7, samples at pH 7 were reduced with NaBH4 under anoxic conditions,

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re-oxidized under air, with the solution pH then adjusted to 7 and optical properties acquired.


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Sequential borohydride and dithionite reductions. To test for the presence of carbonyl-

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containing groups in HS that are reversibly reduced by both borohydride and dithionite and to

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examine their possible contribution to the HS optical properties, samples were sequentially

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reduced, first by borohydride and then by dithionite. One set of samples was kept strictly

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anaerobic throughout the entire reduction sequence, thereby preventing possible re-oxidation of

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any reduced carbonyl moieties; this set of samples was opened to air only following the

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dithionite reduction step. The second set of samples was allowed to re-oxidize following

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borohydride reduction and prior to the dithionite reduction, thereby allowing possible re-

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oxidation of any reduced carbonyl moieties. The samples were then reduced with dithionite

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under anoxic conditions and opened to air following the dithionite reduction step. Changes in the

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optical properties were monitored throughout the reduction/re-oxidation steps.

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Results

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Chemical reduction of model compounds. The anaerobic transfer of dithionite from the

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stock solution to de-oxygenated Milli-Q water produced the characteristic absorption spectrum

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of dithionite,54 which exhibits a prominent band at 316 nm (ε = 8000 M-1cm-1)54, 55 (Fig. S1). The

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absorbance of this band was reproducible upon independent additions from the stock to Milli-Q

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water (within 5%), was stable for minimally 15 min, but was eliminated within minutes upon

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aeration due to its rapid reaction with O2 (Fig. SI 1).57, 58 These results demonstrate that our

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protocols maintained anoxic conditions during both the transfer of solutions and within the

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samples themselves. Unlike dithionite, sodium borohydride does not react with O2 (but does

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react with H3O+)56 and exhibits no absorption across the UV/visible spectrum upon addition to

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Milli-Q water under either aerobic or anoxic conditions.4, 46


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To assess the reactivity of these reductants with model quinones and ketones, dithionite

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and sodium borohydride were added to aqueous solutions of these compounds under anoxic

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conditions. As revealed by the change in the absorption spectra, all fourteen quinones

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investigated showed evidence of reduction to their corresponding hydroquinones within minutes

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following addition of an approximately 2-fold molar excess of either dithionite (Fig. S2, red

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lines) or borohydride.4 Following reduction, the absorption spectra of all model quinones showed

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the loss of the long wavelength n π* transition and formation of the shorter wavelength

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absorption of the hydroquinone, except for anthraquinone-1,5-disulfonate, whose hydroquinone

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is known to exhibit a red-shifted absorption relative to the quinone (Fig. S2, red lines).52 Thus,

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the absorption spectra of the quinones following dithionite reduction were consistent with those

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observed previously with borohydride reduction.4

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Unlike the quinones, treatment of the four model aromatic ketones with excess dithionite

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did not alter their absorption spectra (Fig. S3, right panel, red lines). Instead, the spectra were

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only altered by the strong absorption of the unreacted dithionite (anoxic conditions; Fig. S1 and

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Fig. S3). Following addition of air, which eliminated the dithionite absorption owing to its rapid

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reaction with O257 (Fig. S1), the original spectra of the ketones were observed (Fig, S3, right

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panel, green lines), thus showing that these ketones do not react with dithionite to form the

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alcohols. These results are consistent with prior work,59-61 which demonstrated that dithionite

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does not reduce ketones/aldehydes in water except at high temperatures (>85 oC)59 or in mixed

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solvents.60, 61

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In contrast to dithionite, treatment of the aromatic ketones with a molar excess of

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borohydride caused the loss of the strong carbonyl absorption of the ketones while producing the

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weaker absorption of the corresponding alcohols, which persisted upon subsequent exposure of


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the samples to air (Fig. S3, left panel, red and green lines, respectively). These findings are in

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agreement with the irreversible reduction of ketones by borohydride to their corresponding

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alcohols.56 Overall, our results thus indicate that dithionite can selectively reduce quinones to

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their corresponding hydroquinones under our conditions, namely at room temperature, under

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anoxia, and employing low dithionite concentrations (range of ~10 to 100 µM) (this work). In

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contrast, borohydride reduces not only quinones,4 but also ketones (Fig. S3).

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Dithionite reduction of HS and lignin. All of the samples examined showed

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spectrophotometric evidence of dithionite consumption (loss of dithionite absorption at 316 nm)

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when the HS and dithionite were combined under anoxic conditions. This loss of dithionite was

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rapid, occurring within the time between sample addition and spectrophotometric observation

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(minutes); no further loss was observed over the course of 15 min. The extent of consumption

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varied with HS (Fig. 1), with LHA and ESHA showing high levels of consumption, and PLFA,

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LAC and SRFA showing significantly smaller levels. Estimates of quinone content based on

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dithionite consumption were in good agreement with past electrochemical measurements of

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quinone content for ESHA, LHA, and PLFA (stars in Fig. 1),16 although our estimate for SRFA

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was significantly smaller. However, Aeschbacher et al.16 employed SRFA II in their analyses,

283

while we employed SRFA I, possibly explaining the observed discrepancy. These data provide

284

evidence that dithionite is reducing the same pool of electron acceptors previously identified as

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quinones via electrochemical measurements.

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Despite the reaction with dithionite, aquatic HS (SRFA and SRHA) and lignin (LAC)

287

exhibited little or no change in absorption following reduction with dithionite under anoxic

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conditions at pH 10; changes were ≤ 5 % at wavelengths shorter than 450 nm (Figs. S4-S5-S6,

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red lines), while only slightly larger changes were observed at longer wavelengths, < 20 % for λ


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> 450 nm) (Fig. 2; Figs. S4-S5-S6, red lines). Microbial HS (PLFA) exhibited no detectable

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changes. By comparison, ESHA and LHA exhibited only very small absorption losses over the

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500-600 nm wavelength range (< 10 %), but enhanced absorption at wavelengths > 600 nm (Fig

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2, Figs. S4-S5-S6); this increase in absorption was more pronounced for ESHA than LHA. For

294

all samples, these changes in absorption, when measurable, were largely reversible upon

295

exposure of the reduced samples to air, as shown by the comparable absorption spectra observed

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for the original and re-oxidized samples following exposure to air for 24 h (Fig. 2, Figs. S4-S5-

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S6, green lines).

298

Similarly, dithionite reduction of all samples except ESHA produced very little or no

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change in the fluorescence spectra of these materials at excitation wavelengths (λexc) across both

300

the visible (Fig. 2, insets, red lines; Fig. S7, right panel, red lines) and UV (Fig. S7, left panel,

301

red lines). The single exception, ESHA, exhibited a significant increase in fluorescence emission

302

in the visible range at λexc=500 nm following dithionite reduction. This distinct change in ESHA

303

emission was largely reversible upon exposure of the reduced ESHA to air, as evidenced by the

304

similar fluorescence spectra observed for the untreated and re-oxidized ESHA sample (Fig. 2,

305

inset; Fig. S7, right panel). Thus, only in this instance was evidence acquired for significant

306

fluorescence emission arising from the formation of putative hydroquinone(s) following

307

reduction.

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A direct comparison between dithionite and borohydride treatments for reduction and re-

309

oxidation can only be made at high pH due to the reaction of the borohydride with H3O+, which

310

raises the pH following its addition. Thus, the above reductions with dithionite were performed

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at pH 10 as a means to directly compare with the borohydride reductions presented below. To

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assess the possible effects of solution pH on the dithionite-induced optical changes, reductions


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were also performed at pH 7 (Fig. 3, Figs. S8-S9). Compared to the pH 10 treatments, dithionite

314

reduction of the HS at pH 7 resulted in slightly larger absorption losses at λ > 400 nm for SRFA

315

and LAC (~40-50% at ~600 nm) (Fig. 3, Figs. S8-S9), but the total absorption losses across the

316

visible remained far smaller than those following borohydride reduction (see below). The

317

reversibility of these changes upon exposure of the reduced HS to air was somewhat less, ~20%

318

recovery of total loss, undoubtedly due to the much shorter air exposure time for these samples -

319

10 min as compared to 24 hr - and possibly due to the lower pH as well. Changes in fluorescence

320

following reduction were also very small or negligible, but when measurable, were partially

321

reversible (Fig. 3, insets). ESHA again exhibited the largest fluorescence increase following

322

reduction with clear evidence of reversibility under aeration.

323

Borohydride reduction of HS and lignin. Reduction of aquatic (SRFA and SRHA),

324

microbial (PLFA) HS and the lignin sample (LAC) with a 20-fold mass excess of borohydride at

325

pH 10 resulted in substantial losses of absorption across the entire UV and visible wavelength

326

ranges (Fig. 4; Figs. S10-12, red lines). Consistent with prior work,4, 22, 26, 27, 44, 46 these losses

327

were most pronounced in the visible (i.e., > 60 %). Borohydride reduction of the ESHA and

328

LHA also resulted in significant absorption losses across the entire UV/VIS range, but far

329

smaller losses were observed at wavelengths > 600 nm. Exposure of the borohydride-reduced HS

330

to air produced only minor reversibility of the absorption changes (~ 20 % of original losses)

331

exclusively at λ > 450 nm (Fig. 4; Figs. S10-12, green lines), in reasonably agreement with the

332

small magnitudes of the spectral changes observed with dithionite reduction at this pH (Fig. 2).

333

These results demonstrate that the absorption changes produced by borohydride reduction are far

334

greater in magnitude than those produced by dithionite reduction and further, that these changes

335

are largely, but not completely, irreversible under aeration in accord with prior work.46 The very


15


Page 16 of 31

336

small degree of reversibility observed following borohydride reduction is consistent with the

337

results of the dithionite reduction and re-oxidation; dithionite reduction of quinones to

338

hydroquinones does not largely alter the absorption of these HS and when measurable, these

339

small changes are largely reversible upon re-oxidation. For all the HS investigated, absorption at

340

long wavelengths in the visible was not completely eliminated upon borohydride reduction, due

341

at least in part to the incomplete reduction of the HS by this mass excess of borohydride;46-48

342

increasing the mass excess of borohydride causes more absorption to be lost, but this absorption

343

is never completely removed.46

344

Reduction of these samples with borohydride produced substantially-enhanced blue-shifted

345

fluorescence emission, with two- to three-fold intensity increases for all samples, except PLFA,

346

in the UV (data not shown) and visible (Fig. 4, insets, red lines). These results are in accord with

347

the prior work of Schendorf et al.46 on these same samples, and as well with other work

348

conducted on differing sets of HS and CDOM.4, 44, 46-48 Except for LAC and ESHA, the

349

fluorescence intensities of all the HS remained enhanced after re-oxidation relative to the

350

untreated materials (Fig. 4, insets, green lines). The small reversibility of the emission for ESHA

351

and LAC was consistent with the optical changes observed with dithionite reduction (Fig. 2). The

352

optical changes produced by reduction with borohydride were therefore largely irreversible,

353

consistent with the irreversible reduction of ketones/aldehydes to alcohols.

354

To assess a possible pH dependence of the borohydride-induced changes in the optical

355

properties of the samples, separate reductions of HS and LAC with a 20-fold mass excess of

356

borohydride were conducted at initial pH of 7, with the pH of the reduced samples (~10) then re-

357

adjusted to pH 7 following re-oxidation by aeration. Consistent with prior work,46 the optical

358

changes observed were spectrally similar overall, except for the gain in absorbance at > 600 nm


16

Page 17 of 31


359

recorded for terrestrial HS (LHA and ESHA) that was more pronounced at the lower pH

360

(compare Fig. 4 to Fig. 5 and Figs. S11 to S13). The reversibility of borohydride reduction at

361

neutral pH could not be investigated as it was not possible re-adjust the pH to 7 anaerobically

362

using our protocol.

363

As these samples were originally 0.2 µm-filtered, scattering contributions to these optical

364

signals should be minimal. Further, that these optical changes could result from scattering

365

induced by the dithionite or borohydride treatments is not supported by the reversible changes in

366

optical properties that are observed (see above and further below), nor by the losses (or absence

367

of losses) in absorption that are seen in the dithionite and borohydride treated samples; scattering

368

by particle formation would produce an apparent enhancement of absorption across the

369

spectrum. Finally, no evidence for an enhanced scattering signal was observed at wavelengths on

370

the red edge of the excitation wavelengths employed in the fluorescence measurements– an

371

enhancement of this scattering signal just off the excitation maxima can be employed as a

372

sensitive measure of particle formation.

373

Sequential borohydride and dithionite reductions. To test further whether borohydride

374

treatment at pH 10 resulted in the reduction of both ketones/aldehydes and quinones under

375

anoxic conditions, as well as to gain further information on their relative contributions to the

376

optical properties, sequential reduction experiments were conducted. Samples were first reduced

377

under anoxic conditions with a 20-fold mass excess of borohydride to allow reduction of both

378

ketones/aldehyde and quinone moieties. This initial borohydride treatment produced results

379

consistent with those described above (Figs. S15-S16, red lines) and by Schendorf et al.46 Further

380

reduction with dithionite while maintaining anoxic condition produced no additional changes in

381

the optical properties (Figs. S15-S16, green lines), indicating that all dithionite-reducible


17


382

moieties in the HS (ie. quinones) had already been reduced in the preceding borohydride-

383

reduction step.

384

Page 18 of 31

In a second experimental approach, borohydride-reduced samples (Fig. S17, red lines)

385

were re-oxidized in air (Fig. S17, green lines) prior to treatment with dithionite. Re-oxidation of

386

the borohydride treated HS resulted in small absorption recoveries at longer wavelengths (> 450

387

nm), consistent with the results described above (Fig. S17, green lines). The subsequent

388

reduction of these re-oxidized HS with dithionite under anoxic conditions reversed the

389

absorption recovery observed during the preceding re-oxidation step (Fig. S17, yellow lines). Re-

390

oxidation of these dithionite-reduced HS again resulted in absorption recovery at longer

391

wavelengths (> 450 nm), similar to that observed upon re-oxidation of the borohydride-reduced

392

HS (Fig. S17, blue lines). These absorption recoveries were small (< 20 %) and restricted to

393

wavelengths > 450 nm.

394

Changes in fluorescence were substantial and mostly irreversible upon borohydride

395

reduction (Fig. S18), except for LAC and ESHA, which showed some reversibility following

396

dithionite reduction and subsequent re-oxidation. Overall, the change in the optical properties

397

observed upon sequential borohydride and dithionite reduction provide additional evidence that

398

borohydride treatment causes the reduction of both ketones/aldehydes and quinones in these

399

samples. Moreover, the optical changes observed in the sequential reduction and oxidation steps

400

mirror the changes that are observed upon treatment of these samples by each reductant

401

individually; pronounced and largely irreversible changes upon borohydride treatment and much

402

smaller and reversible changes upon dithionite treatment.

403

Discussion and Environmental Implications


18

Page 19 of 31

404


Our results indicate that dithionite can be employed to reduce selectively quinones in the

405

presence of ketone/aldehydes, while borohydride can reduce both of these classes of moieties.

406

Further, based on the consumption of dithionite by the HS and LAC under anoxic conditions,

407

estimates of quinone content in these types of materials may be acquired, while the impact of

408

quinone/hydroquinone moieties to the optical properties can be further investigated. Except for

409

ESHA, which shows a relatively strong increase in emission at excitation wavelengths in the

410

visible following reduction with dithionite (Figs. 2-3), small or negligible changes in absorption

411

and emission were observed for the other HS and LAC (Figs. 2-3) as compared with those

412

observed for borohydride reduction (Figs. 4,5), indicating that quinones/hydroquinones have a

413

much smaller influence on the optical properties of these materials, directly contradicting the

414

conclusions of a number of past studies38-41 and the idea that redox state can be monitored by

415

quinone/hydroquinone “fluorescence”.41, 62 These results are, however, in accord with the past

416

measurements and the conclusions offered by Macalady and Walton-Day32, 33 and Maurer et al.25

417

Most of the small changes in absorption observed following dithionite reduction, occurred in the

418

visible at wavelengths greater than ~450 nm (Figs. 2-3), suggesting either a contribution of long-

419

wavelength absorbing quinones/hydroquinones,4,42 such as may be the case for ESHA, or low

420

energy CT transitions with quinones acting as acceptors.3, 5 In contrast, borohydride reduction

421

leads to substantial and largely irreversible changes in the optical properties, indicating a

422

prominent contribution of ketones/aldehydes to the optical properties particularly at visible

423

wavelengths, consistent with their acting as the primary electron acceptors in CT transitions. Our

424

results suggest that the combined use of dithionite and borohydride reductions can lead to a

425

better understanding of the molecular moieties and interactions contributing to the absorption

426

and emission properties of HS and CDOM.


19


Page 20 of 31

427

Associated content

428 429 430

Supporting information

431

addition of air (Fig. S1); Changes in absorption spectra of model quinones following reduction with

432

Na2S2O4 (Fig. S2); Changes in absorption of model ketones following addition of NaBH4 and

433

Na2S2O4 (Fig. S3); Absorption (250-800 nm), fractional absorbance change, Log of absorption

434

and fluorescence emission spectra, prior to and following reduction with Na2S2O4 at pH 10 (Figs.

435

S4-S5-S6-S7); Fractional absorbance change and Log of absorption prior to and following

436

reduction with Na2S2O4 at pH 7 (Figs. S8-S9); Absorption (250-800 nm), fluorescence emission

437

spectra, fractional absorbance change and Log of absorption prior to and following reduction

438

with NaBH4 at pH 10 (Figs. S10-S12); Fractional absorbance change and Log of absorption

439

following reduction with NaBH4 at pH 7 (Figs. S13-S14); Absorption spectra (400-800 nm) and

440

fluorescence emission spectra prior to and following anoxic sequential reduction with NaBH4

441

and NaS2O4 (Figs. S15-S16); Fractional absorbance changes and fluorescence emission spectra

442

prior to and following reduction and re-oxidation with NaBH4 first and Na2S2O4 next (Figs. S17-

443

S18). This material is available free of charge via the Internet at http://pubs.acs.org.

444

Figure Captions

445

Figure 1. Quinone content based on Na2S2O4 consumption during reduction of HS and LAC.

446

Contents are based on Na2S2O4 molar absorptivity at 316 nm of 8000 M-1cm-1. Stars represent

447

quinone content from Aeschbacher et al. (2010).6 Error bars represent the standard deviation of 6

448

independent measurements. SRHA is not shown because our stock ran out and IHSS has been

449

unable to supply more.

450

Figure 2. Absorption (400-800 nm) and fluorescence emission spectra (insets; at λexc, 500, 540,

451

and 580 nm) of aquatic (SRFA, SRHA), microbial (PLFA), terrestrial (ESHA) and lignite-

Absorption spectra of Milli Q water blank at neutral pH, following addition of Na2S2O4 and after


20

Page 21 of 31


452

derived (LHA) humic substances and of lignin (LAC) prior to and following reduction with

453

sodium dithionite (Na2S2O4). Samples prepared in Milli Q water, adjusted to pH 10, and had the

454

following concentrations for absorption: SRFA, PLFA, LAC: 100 mg/L; SRHA: 50 mg/L; LHA:

455

6 mg/L; ESHA: 20 mg/L. Approximately 40 µL of 1.7 g/L Na2S2O4 were added to 3 mL of HS.

456

Figure 3. Absorption spectra (400-800 nm) and corrected fluorescence emission spectra (insets,

457

λexc 500, 540, and 580 nm) of aquatic (SRFA), microbial (PLFA), terrestrial (ESHA), lignite-

458

derived (LHA) humic substances and lignin (LAC) prior to and following reduction (15 min)

459

with sodium dithionate (Na2S2O4) and following re-oxidation in air (10 min). Samples in

460

phosphate buffer (10 mM), pH 7 with concentrations as follows for absorption and fluorescence

461

spectra, respectively: SRFA (100 and 30 mg/L); PLFA (100 and 50 mg/L); LAC (100 and 50

462

mg/L); LHA (50 and 7.5 mg/L); ESHA (50 and 7.5 mg/L). Approximately 10 µL of 1.13 g/L

463

Na2S2O4 were added to 3 mL of HS. SRHA is not shown because our stock ran out and IHSS has

464

been unable to provide more.

465

Figure 4. Absorption (400-800 nm) and fluorescence emission spectra (insets; λexc 500, 540, and

466

580 nm) of aquatic (SRFA, SRHA), microbial (PLFA), terrestrial (ESHA), lignite-derived

467

(LHA) HS and lignin (LAC) prior to and following borohydride reduction (NaBH4) with 20

468

fold mass excess. Samples prepared in Milli Q water and adjusted to pH ~10 and had the

469

following concentrations: SRFA, PLFA, LAC: 100 mg/L; SRHA: 50 mg/L; LHA and ESHA 10

470

mg/L.

471

Figure 5. Absorption spectra (400-800 nm) and corrected fluorescence emission spectra (insets,

472

λexc 500, 540, and 580 nm) of aquatic (SRFA), microbial (PLFA), terrestrial (ESHA), lignite-

473

derived (LHA) humic substances and lignin (LAC) prior to reduction (black lines) and following

474

the reduction/re-oxidation of the reduced samples for 24 hours in air (green lines). Samples in

475

phosphate buffer (10 mM) at pH 7 with concentrations as follows: SRFA (30 mg/L); PLFA (50

476

mg/L); LAC (50 mg/L); LHA (7.5 mg/L); ESHA (7.5 mg/L). SRHA is not shown because our

477

stock ran out and IHSS has been unable to supply more.

478 479

Author information


21


480

Corresponding authors.

481

Rossana Del Vecchio (email: [email protected], telephone: 001-301-405-0337)

482

Neil V. Blough (email: [email protected], telephone: 001 301-405-0051)

Page 22 of 31

483 484

Acknowledgements.

485

RDV and NB thank Michael Sander and Michael Aeschbacher for their comments and

486

discussions to improve this manuscript. RDV and NVB thank the National Science Foundation

487

(OCE1032223) and (OCE 0648414).

488 489 490

References

491 492 493 494 495 496 497 498 499 500 501 502 503 504 505 506 507 508 509 510 511 512 513 514 515 516 517 518

1. Del Vecchio, R.; Blough, N. V., Photobleaching of chromophoric dissolved organic matter in natural waters: kinetics and modeling. Marine Chemistry 2002, 78, (4), 231-253. 2. Boyle, E. S.; Guerriero, N.; Thiallet, A.; Del Vecchio, R.; Blough, N. V., Optical Properties of Humic Substances and CDOM: Relation to Structure. Environmental Science & Technology 2009, 43, (7), 22622268. 3. Del Vecchio, R.; Blough, N. V., On the origin of the optical properties of humic substances. Environmental Science & Technology 2004, 38, (14), 3885-3891. 4. Ma, J. H.; Del Vecchio, R.; Golanoski, K. S.; Boyle, E. S.; Blough, N. V., Optical Properties of Humic Substances and CDOM: Effects of Borohydride Reduction. Environmental Science & Technology 2010, 44, (14), 5395-5402. 5. Sharpless, C. M.; Blough, N. V., The importance of charge-transfer interactions in determining chromophoric dissolved organic matter (CDOM) optical and photochemical properties. Environmental Science-Processes & Impacts 2014, 16, (4), 654-671. 6. Schendorf, T. M. Effect of Borohydride Reduction and pH on the Optical Properties of Humic Substances. MS, University of Maryland, UMD Theses and Dissertations, 2014. 7. Dryer, D. J.; Korshin, G. V.; Fabbricino, M., In situ examination of the protonation behavior of fulvic acids using differential absorbance spectroscopy. Environmental Science & Technology 2008, 42, (17), 6644-6649. 8. Phillips, S. M.; Bellcross, A. D.; Smith, G. D., Light Absorption by Brown Carbon in the Southeastern United States is pH-dependent. Environmental Science & Technology 2017, 51, (12), 67826790. 9. Piccolo, A., The supramolecular structure of humic substances. Soil Science 2001, 166, (11), 810832. 10. Sutton, R.; Sposito, G., Molecular structure in soil humic substances: The new view. Environmental Science & Technology 2005, 39, (23), 9009-9015. 11. Dreyer, D. R.; Miller, D. J.; Freeman, B. D.; Paul, D. R.; Bielawski, C. W., Elucidating the Structure of Poly(dopamine). Langmuir 2012, 28, (15), 6428-6435.


22

Page 23 of 31

519 520 521 522 523 524 525 526 527 528 529 530 531 532 533 534 535 536 537 538 539 540 541 542 543 544 545 546 547 548 549 550 551 552 553 554 555 556 557 558 559 560 561 562 563 564 565 566


12. Liu, Y. L.; Ai, K. L.; Lu, L. H., Polydopamine and Its Derivative Materials: Synthesis and Promising Applications in Energy, Environmental, and Biomedical Fields. Chemical Reviews 2014, 114, (9), 50575115. 13. Latch, D. E.; McNeill, K., Microheterogeneity of singlet oxygen distributions in irradiated humic acid solutions. Science 2006, 311, (5768), 1743-1747. 14. Blough, N. V., Electron-paramagnetic resonance measurements of photochemical radical production in Humic substances.1. Effects of O-2 and charge on radical scavenging by nitroxides. . Environmental Science & Technology 1988, 22, (1), 77-82. 15. Green, S. A.; Morel, F. M. M.; Blough, N. V., Investigation of the electrostatic properties of Humic substances by fluorescence quenching. Environmental Science & Technology 1992, 26, (2), 294-302. 16. Aeschbacher, M.; Sander, M.; Schwarzenbach, R. P., Novel Electrochemical Approach to Assess the Redox Properties of Humic Substances. Environmental Science & Technology 2010, 44, (1), 87-93. 17. Aeschbacher, M.; Vergari, D.; Schwarzenbach, R. P.; Sander, M., Electrochemical Analysis of Proton and Electron Transfer Equilibria of the Reducible Moieties in Humic Acids. Environmental Science & Technology 2011, 45, (19), 8385-8394. 18. Aeschbacher, M.; Graf, C.; Schwarzenbach, R. P.; Sander, M., Antioxidant Properties of Humic Substances. Environmental Science & Technology 2012, 46, (9), 4916-4925. 19. Korshin, G. V.; Benjamin, M. M.; Li, C. W., Use of differential spectroscopy to evaluate the structure and reactivity of humics. Water Science and Technology 1999, 40, (9), 9-16. 20. Ritchie, J. D.; Perdue, E. M., Proton-binding study of standard and reference fulvic acids, humic acids, and natural organic matter. Geochimica Et Cosmochimica Acta 2003, 67, (1), 85-96. 21. Leenheer, J. A.; Wilson, M. A.; Malcolm, R. L., Presence and potential significance of aromaticketone groups in aquatic humic substances. Organic Geochemistry 1987, 11, (4), 273-280. 22. Baluha, D. R.; Blough, N. V.; Del Vecchio, R., Selective Mass Labeling for Linking the Optical Properties of Chromophoric Dissolved Organic Matter to Structure and Composition via Ultrahigh Resolution Electrospray Ionization Mass Spectrometry. Environmental Science & Technology 2013, 47, (17), 9891-9897. 23. Canonica, S.; Jans, U.; Stemmler, K.; Hoigne, J., Transformation kinetics of phenols in water Photosensitization by dissolved natural organic material and aromatic ketones. Environmental Science & Technology 1995, 29, (7), 1822-1831. 24. Canonica, S.; Hellrung, B.; Wirz, J., Oxidation of phenols by triplet aromatic ketones in aqueous solution. Journal of Physical Chemistry A 2000, 104, (6), 1226-1232. 25. Pos, W. H.; Riemer, D. D.; Zika, R. G., Carbonyl sulfide (OCS) and carbon monoxide (CO) in natural waters: evidence of a coupled production pathway. Marine Chemistry 1998, 62, (1-2), 89-101. 26. Golanoski, K. S.; Fang, S.; Del Vecchio, R.; Blough, N. V., Investigating the Mechanism of Phenol Photooxidation by Humic Substances. Environmental Science & Technology 2012, 46, (7), 3912-3920. 27. Zhang, Y.; Del Vecchio, R.; Blough, N. V., Investigating the Mechanism of Hydrogen Peroxide Photoproduction by Humic Substances. Environmental Science & Technology 2012, 46, (21), 1183611843. 28. Zhang, Y.; Simon, K. A.; Andrew, A. A.; Del Vecchio, R.; Blough, N. V., Enhanced Photoproduction of Hydrogen Peroxide by Humic Substances in the Presence of Phenol Electron Donors. Environmental Science & Technology 2014, 48, (21), 12679-12688. 29. Zhang, Y.; Blough, N. V., Photoproduction of One-Electron Reducing Intermediates by Chromophoric Dissolved Organic Matter (CDOM): Relation to O-2(-) and H2O2 Photoproduction and CDOM Photooxidation. Environmental Science & Technology 2016, 50, (20), 11008-11015. 30. Sharpless, C. M.; Aeschbacher, M.; Page, S. E.; Wenk, J.; Sander, M.; McNeill, K., PhotooxidationInduced Changes in Optical, Electrochemical, and Photochemical Properties of Humic Substances. Environmental Science & Technology 2014, 48, (5), 2688-2696.


23


567 568 569 570 571 572 573 574 575 576 577 578 579 580 581 582 583 584 585 586 587 588 589 590 591 592 593 594 595 596 597 598 599 600 601 602 603 604 605 606 607 608 609 610 611 612 613 614

Page 24 of 31

31. Wenk, J.; Aeschbacher, M.; Salhi, E.; Canonica, S.; von Gunten, U.; Sander, M., Chemical Oxidation of Dissolved Organic Matter by Chlorine Dioxide, Chlorine, And Ozone: Effects on Its Optical and Antioxidant Properties. Environmental Science & Technology 2013, 47, (19), 11147-11156. 32. Wenk, J.; Aeschbacher, M.; Sander, M.; von Gunten, U.; Canonica, S., Photosensitizing and Inhibitory Effects of Ozonated Dissolved Organic Matter on Triplet-Induced Contaminant Transformation. Environmental Science & Technology 2015, 49, (14), 8541-8549. 33. Sharpless, C. M., Lifetimes of Triplet Dissolved Natural Organic Matter (DOM) and the Effect of NaBH4 Reduction on Singlet Oxygen Quantum Yields: Implications for DOM Photophysics. Environmental Science & Technology 2012, 46, (8), 4466-4473. 34. Maurer, F.; Christl, I.; Kretzschmar, R., Reduction and Reoxidation of Humic Acid: Influence on Spectroscopic Properties and Proton Binding. Environmental Science & Technology 2010, 44, (15), 57875792. 35. Blough, N. V.; Del Vecchio, R., Comment on "HILIC-NMR: Toward the Identification of Individual Molecular Components in Dissolved Organic Matter". Environmental Science & Technology 2011, 45, (13), 5908-5909. 36. Fimmen, R. L.; Cory, R. M.; Chin, Y.-P.; Trouts, T. D.; McKnight, D. M., Probing the oxidationreduction properties of terrestrially and microbially derived dissolved organic matter. Geochimica Et Cosmochimica Acta 2007, 71, (12), 3003-3015. 37. Klapper, L.; McKnight, D. M.; Fulton, J. R.; Blunt-Harris, E. L.; Nevin, K. P.; Lovley, D. R.; Hatcher, P. G., Fulvic acid oxidation state detection using fluorescence spectroscopy. Environmental Science & Technology 2002, 36, (14), 3170-3175. 38. Cory, R. M.; McKnight, D. M., Fluorescence spectroscopy reveals ubiquitous presence of oxidized and reduced quinones in dissolved organic matter. Environmental Science & Technology 2005, 39, (21), 8142-8149. 39. Ariese, F.; van Assema, S.; Gooijer, C.; Bruccoleri, A. G.; Langford, C. H., Comparison of Laurentian Fulvic Acid luminescence with that of the hydroquinone/quinone model system: Evidence from low temperature fluorescence studies and EPR spectroscopy. Aquatic Sciences 2004, 66, (1), 86-94. 40. Miller, M. P.; McKnight, D. M.; Cory, R. M.; Williams, M. W.; Runkel, R. L., Hyporheic exchange and fulvic acid redox reactions in an alpine stream/wetland ecosystem, Colorado front range. Environmental Science & Technology 2006, 40, (19), 5943-5949. 41. Mladenov, N.; Zheng, Y.; Miller, M. P.; Nemergut, D. R.; Legg, T.; Simone, B.; Hageman, C.; Rahman, M. M.; Ahmed, K. M.; McKnight, D. M., Dissolved Organic Matter Sources and Consequences for Iron and Arsenic Mobilization in Bangladesh Aquifers. Environmental Science & Technology 2010, 44, (1), 123-128. 42. Macalady, D. L.; Walton-Day, K., New light on a dark subject: On the use of fluorescence data to deduce redox states of natural organic matter (NOM). Aquatic Sciences 2009, 71, (2), 135-143. 43. Macalady, D. L.; Walton-Day, K., Redox Chemistry and Natural Organic Matter (NOM): Geochemists' Dream, Analytical Chemists' Nightmare. Aquatic Redox Chemistry 2011, 1071, 85-+. 44. Andrew, A. A.; Del Vecchio, R.; Subramaniam, A.; Blough, N. V., Chromophoric dissolved organic matter (CDOM) in the Equatorial Atlantic Ocean: Optical properties and their relation to CDOM structure and source. Marine Chemistry 2013, 148, 33-43. 45. Andrew, A. A.; Del Vecchio, R.; Zhang, Y.; Subramaniam, A.; Blough, N. V., Are Extracted Materials Truly Representative of Original Samples? Impact of C18 Extraction on CDOM Optical and Chemical Properties. Frontiers in Chemistry 2016, 4. 46. Schendorf, T. M.; Del Vecchio, R.; Koech, K.; Blough, N. V., A standard protocol for NaBH4 reduction of CDOM and HS. Limnology and Oceanography-Methods 2016, 14, (6), 414-423. 47. Phillips, S. M.; Smith, G. D., Light Absorption by Charge Transfer Complexes in Brown Carbon Aerosols. Environmental Science & Technology Letters 2014, 1, (10), 382-386.


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48. Phillips, S. M.; Smith, G. D., Further Evidence for Charge Transfer Complexes in Brown Carbon Aerosols from Excitation-Emission Matrix Fluorescence Spectroscopy. Journal of Physical Chemistry A 2015, 119, (19), 4545-4551. 49. Tratnyek, P. G.; Macalady, D. L., Abiotic reduction of nitro aromatic pesticides in anaerobic laboratory systems. Journal of Agricultural and Food Chemistry 1989, 37, (1), 248-254. 50. Schwarzenbach, R. P.; Stierli, R.; Lanz, K.; Zeyer, J., Quinones and iron porphyrin mediated reduction of nitroaromatic compounds in homogeneous aqueous-solution. Environmental Science & Technology 1990, 24, (10), 1566-1574. 51. Hernandez-Montoya, V.; Alvarez, L. H.; Montes-Moran, M. A.; Cervantes, F. J., Reduction of quinone and non-quinone redox functional groups in different humic acid samples by Geobacter sulfurreducens. Geoderma 2012, 183, 25-31. 52. Thomson, R. H., In Naturally Occurring Quinones (Second Edition), Press, A., Ed. London and New York, 1971; pp 39-92. 53. Tinnacher, R. M.; Honeyman, B. D., A new method to radiolabel natural organic matter by chemical reduction with tritiated sodium borohydride. Environmental Science & Technology 2007, 41, (19), 6776-6782. 54. Dixon, M., Acceptor specificity of flavins and flavoproteins. 1. Techniques for anaerobic spectrophotometry. Biochimica Et Biophysica Acta 1971, 226, (2), 241-&. 55. Mayhew, S. G., Redox potential of dithionite and SO2- from equilibrium reactions with flavodoxins, methyl viologen and hydrogen plus hydrogenase. European Journal of Biochemistry 1978, 85, (2), 535-547. 56. Rohm; Haas, Sodium Borohydride Digest. In http://www.dow.com/assets/attachments/industry/pharma_medical/chemical_reagents/reducing_age nts/sodium_borohydride_digest.pdf, 2003. 57. Creutz, C.; Sutin, N., Kinetics of reactions of sodium dithionite with dioxygen and hydrogenperoxide. Inorganic Chemistry 1974, 13, (8), 2041-2043. 58. Tao, Z.; Goodisman, J.; Souid, A. K., Oxygen measurement via phosphorescence: Reaction of sodium dithionite with dissolved oxygen. Journal of Physical Chemistry A 2008, 112, (7), 1511-1518. 59. Devries, J. G.; Kellogg, R. M., Reduction of aldehydes and ketones by sodium dithionite. Journal of Organic Chemistry 1980, 45, (21), 4126-4129. 60. Dhillon, R. S.; Singh, R. P.; Kaur, D., Selective 1,4-reduction of conjugated aldehydes and ketones in the presence of unconjugated aldehydes and ketones with sodium dithionite. Tetrahedron Letters 1995, 36, (7), 1107-1108. 61. Chung, S. K., Mechanism of sodium dithionite reduction of aldehydes and ketones. Journal of Organic Chemistry 1981, 46, (26), 5457-5458. 62. Yang, Z.; Kappler, A.; Jiang, J., Reducing capacities and distribution of redox-active functional groups in low molecular weight fractions of humic acids. Environmental Science & Technology 2016, 50, (2), 12105–12113.

653 654


25


655

Figures.

656

Figure 1.

Page 26 of 31

657

658 659


26

Page 27 of 31

660


Figure 2. untreated (pH~10) Na2S204 reduced (15 min) (pH ~ 10)

Corrected fluorescence emission


reoxidized (24 hr) (pH ~10)

0.3

0.2

0.2

0.2 0.1

500 550 600 650 Wavelength (nm)

SRFA

Wavelength (nm)

SRHA 0.3 0.2

0.1

0.10

0.10

0.1

0.0

0.0

500 550 600 650

0.05

0.0



0.2

0.15

PLFA

Wavelength (nm)

LHA 0.6

0.06

0.3

0.4

0.04

0.2

0.02

0.2

0.0

0.00 500 550 600 650

0.05

0.00


0.08

0.10

500 550 600 650 Wavelength (nm)

Wavelength (nm)

0.00

661

0.1

ESHA

LAC

400

0.05

500 550 600 650

Wavelength (nm)

0.00

0.1

500 550 600 650


Absorbance (OD)

0.0

0.0

0.0

0.1

0.2

0.1

500

600

700

400

500

600

700

0.0 800

Wavelength (nm) ACS Paragon Plus Environment

27


Figure 3.

0.2

0.08

0.04

oxidized pH 7 550

600


0.08

650

PLFA

0.06 0.04 0.02 0.00500

550

600

1.0

0.10

LHA

0.08

0.8

0.06 0.04

0.6

0.02 0.00 500

650

Wavelength (nm)

550

600

0.4

650

Wavelength (nm)

0.05

0.00 0.15

0.10

LAC

0.04 0.03 0.02 0.01 0.00 500

550

600


0.2


Absorbance (OD)

0.10

reduced pH 7

Wavelength (nm)

0.1

0.15

original pH 7

0.02 0.00 500

0.0

SRFA

0.06


0.3


662

Page 28 of 31

0.3

0.2

1.0

0.1

0.8

0.0 500

650

Wavelength (nm)

0.05

ESHA

0.0 1.2

0.6 550

600

650

Wavelength (nm)

0.4 0.2

0.00 400 663

500

600

700

800

500

600

700

0.0 800

Wavelength (nm)

664


28

Page 29 of 31

665


Figure 4. untreated (pH ~10) NaBH4 reduced (20x) (24 hr) (pH 10.7)

0.1

0.4

SRFA


0.2


reoxidized (48 hr) (pH 9.8)

0.4

0.0

0.0

0.08


0.0

500 550 600 650 Wavelength (nm)

LAC

0.1

0.0

LHA

0.12

0.2 0.1 0.0 500 550 600 650 Wavelength (nm)

0.6

0.00 ESHA

0.12

0.0 500 550 600 650 Wavelength (nm)

0.00 400 666

500

600

700

400

0.06

0.3

500 550 600 650 Wavelength (nm)

0.04

0.1

0.0 0.3

500 550 600 650 Wavelength (nm)


0.00


0.04

PLFA

0.1


Absorbance (OD)

0.08

0.2

0.2

0.2

0.2

500 550 600 650 Wavelength (nm)

0.0

SRHA

500

600

700

0.06

0.00 800

Wavelength (nm)

667


29


668

Page 30 of 31

Figure 5.

0.10 0.08 0.06

Corrected Fluorescence Emission

669

0.04

SRFA

0.08 0.06 0.04

original pH 7

0.02

reduced/oxidized pH 7

500

550 600 650 Wavelength (nm)

0.04 0.02


0.00 0.06 0.04

PLFA

0.06 0.04 0.02 0.00 500 550 600 650 Wavelength (nm)


0.06

0.08

0.05

LAC

0.04 0.03 0.02 0.01 0.00 500

0.02

0.16

550 600 650 Wavelength (nm)

0.08

0.06

0.04 0.00 500 550 600 650 Wavelength (nm)

670

0.03

0.00 0.3

ESHA 0.10

0.2 0.1 0.0 500 550 600 650 Wavelength (nm)

0.05

0.00

0.00 400

0.09

LHA

0.12


Absorbance (OD)

0.00


0.02

500

600

700

800

500

600

700

800

Wavelength (nm)

671


30

Page 31 of 31

672


TOC

673 674 675


31

Aldehydes to the Optical

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