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46557-AC2
Redox Reactions of Iron and Manganese that Chemically Transform Natural Organic Matter Within Sediments

Alan T. Stone, Johns Hopkins University

            Dihydroxybenzenes (XH₂) and benzoquinones (Z), believed to be the principal redox-active moieties within natural organic matter (NOM), are linked through half-reactions, e.g.:

XH₂/Z interconversion within NOM is usually assumed to be reversible.  Nevertheless, Michael-Type nucleophilic addition by bisulfide ion (HS^-), which renders the reaction irreversible, has been documented:

Do weaker sulfur-, nitrogen-, oxygen-, and halogen-donor atom nucleophiles form Michael-Type adducts?  Our hypothesis is that unactivated benzoquinones, i.e. those with only alkyl substituents, cannot engage in reactions with weak nucleophiles, while activated benzoquinones possessing electron-withdrawing carboxylate substituents can.  Carboxylate substituents are abundant in lignin, an important NOM precursor biomolecule.  A number of physical methods have demonstrated that NOM is rich in carboxylate groups.  Carboxylate groups make dihydroxybenzenes more resistant towards oxidation.  In order for this means of weak nucleophile incorporation to be viable, oxidants strong enough to act upon carboxylate-substituted dihydroxybenzenes must exist in marine environments.

            Enrichments of manganese(III,IV) (hydr)oxides have been found at the oxic/anoxic interface within the water column of the Black Sea, within marine sediments, and in a numerous other relevant environments.  Mn^III,IV (hydr)oxides are strong enough in a thermodynamic sense and reactive enough in a kinetic sense to oxidize carboxylate-substituted dihydroxybenzenes to activated benzoquinones.  We have investigated the oxidation of more than sixteen phenols and dihydroxybenzenes by Mn^IVO₂(s, pyrolusite).  Here are a few examples: catechol (2.1x10^-3 s^-1) > hydroquinone (6.1 x10^-4 s^-1) > 2,5-dihydroxybenzoic acid (3.3x10^-4 s^-1) > 4,4'-biphenol (2.0x10^-4 s^-1) > 2,5-dihydroxyterephthalic acid (1.7x10^-4 s^-1) > 2,2'-biphenol (7.2x10^-8 s^-1).  These values reflect initial rates of Mn^II solubilization divided by the dihydroxybenzene substrate concentration, measured in a 10 mM MES medium (pH 6.5).

            Next, we investigated adducts formed through attack by OH^-/H₂O, by the nitrogen-donor nucleophile 4-methylimidazole, and by two sulfur-donor nucleophiles, thiosulfate and sulfite.  Our efforts focused on HPLC with electrospray ionization (ES) mass spectrometric (MS) detection, because of its' ability to identify specific adducts.  Capillary electrophoresis and HPLC with UV detection yielded a similar number of peaks, but detection limits were lower, and structure assignments were not possible.

            As expected, p-benzoquinone, classified as an unactivated electrophile, formed an adduct with the representative sulfur-donor nucleophile thiosulfate, but not with OH^-/H₂O or with 4-methylimidazole.  p-Benzoquinone is stable enough for a stock solution to be prepared from an authentic standard.  Thiosulfate or 4-methylimidazole was then added.  With ESI-MS, the ionization process is strong enough to generates the molecular ion from ^-O₃S-S-XH₂ (the thiosulfate adduct), but not from the p-benzoquinone substrate.  HPLC with UV detection was used to monitor concentrations of p-benzoquinone.

            2,5-Dihydroxybenzoic acid oxidation by MnO₂(s) yields the activated electrophile carboxy-p-benzoquinone.  In simple aqueous media (beginning pH ~ 5.6), MnO₂(s) oxidation followed by OH^-/H₂O nucleophilic attack generated three addition products, and one product of oxidative coupling, all shown to the left of the line, below.

4-Methylimidazole is not subject to oxidation by MnO₂(s), so it can be added along with the 2,5-dihydroxybenzoic acid substrate.  50, 100, 200, 300, and 400 mM concentrations were employed.  As the 4-methylimidazole concentration was increased, peaks collected in total ion mode for the left four species to decreased, and new peaks corresponding to the 4-methylimidazole adducts (to the right of the line) increased.  MS fragmentation patterns are consistent with the structural assignments.  Sulfite ion and iodide are subject to oxidation by MnO₂(s).  It was therefore necessary to first generate carboxy-p-benzoquinone using MnO₂(s), then remove unreacted MnO₂(s) prior to addition of nucleophile stock solution using microfiltration.  Sulfite ion yielded an adduct discernible by HPLC-ESI-MS; iodide did not.

            When 2,5-dihydroxyterephthalic acid is oxidized by MnO₂(s), a highly activated electrophile is generated, dicarboxy-p-benzoquinone.  With have conducted experiments employing 50, 100, 200, 300, and 400 mM 4-methylimidazole, and discerned the following products using HPLC-ESI-MS:

The rightmost product, a mixed hydroxide ion/4-methylimidazole adduct, is particularly noteworthy.  It's peak area reached a maximum value when 200 mM 4-methylimidazole was employed. 

            We have several important objectives for the second and final year of this project.  We will expand our structural determinations to include oxygen-donor and halogen-donor nucleophiles representative of marine sedimentary environments.  We are also interested in whether other transient species generated by oxidation, e.g. quinone methides, provide additional opportunities for adduct formation.  We will employ reactions performed at higher concentrations (~ 10 mM) to generate enough quantities of each product for isolation and purification to be conducted.  Once we have authentic standard prepared, we can quantify production rates and compare rate constants for OH^-/H₂O adduct formation with rate constants obtained with nucleophile amendments.  We are currently testing "Good's Buffers" such as MES and MOPS to see if they are sufficiently nucleophilic to generate Michael Addition products.  Once we have identified an inert buffer system, we can carefully work out rates of competing adduct formation reactions within a range of pH appropriate for marine sedimentary environments.

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