Antioxidant Properties of Humic Substances

Humic substances (HS) are heterogeneous, redox-active organic macromolecules. While electron transfer to and from HS under reducing conditions is well investigated, comparatively little is known on the electron donating (i.e., antioxidant) properties of HS under oxic conditions. In this work, the electron donating capacities (EDCs) of terrestrial and aquatic HS were quantified by mediated electrochemical oxidation over a wide range of pH values and applied redox potentials (<i>E</i>_h) using 2,2′-azino-bis­(3-ethylbenzthiazoline-6-sulfonic acid) as an electron transfer mediator. Electrochemical oxidation of three model humic acids (HAs) was largely irreversible, and the EDCs of these HAs increased with increasing <i>E</i>_h and pH. These results suggest that HS contain a wide variety of moieties that are oxidized at different potentials and that, upon oxidation, release protons and undergo irreversible follow-up reactions. At a given pH and <i>E</i>_h, the EDCs of the HS correlated well with their titrated phenol contents suggesting phenolic moieties as major electron donating groups in HS. Comparing the EDCs of 15 HS with their electron accepting capacities (EACs), aquatic HS had higher EDCs and lower EACs than terrestrial HS of comparable aromaticities. These results indicate that oxidative transformation of HS in the environment results in a depletion of electron donating phenolic moieties with antioxidant properties relative to the electron accepting quinone moieties

Adsorption of Insecticidal Cry1Ab Protein to Humic Substances. 1. Experimental Approach and Mechanistic Aspects

Adsorption is a key process affecting the fate of insecticidal Cry proteins (<i>Bt</i> toxins), produced by genetically modified <i>Bt</i> crops, in soils. However, the mechanisms of adsorption to soil organic matter (SOM) remain poorly understood. This work assesses the forces driving the adsorption of Cry1Ab to Leonardite humic acid (LHA), used as a model for SOM. We studied the effects of solution pH and ionic strength (<i>I</i>) on adsorption using a quartz crystal microbalance with dissipation monitoring and optical waveguide lightmode spectroscopy. Initial Cry1Ab adsorption rates were close to diffusion-limited and resulted in extensive adsorption, even at pH >6, at which LHA and Cry1Ab carry negative net charges. Adsorption increased with decreasing <i>I</i> at pH >6, indicating Cry1Ab–LHA patch-controlled electrostatic attraction via positively charged domains of Cry1Ab. Upon rinsing, only a fraction of Cry1Ab desorbed, suggesting a range of interaction energies of Cry1Ab with LHA. Different interaction energies likely resulted from nonuniformity in the LHA surface polarity, with higher Cry1Ab affinities to more apolar LHA regions due to the hydrophobic effect. Contributions from the hydrophobic effect were substantiated by comparison of the adsorption of Cry1Ab and the reference proteins albumin and lysozyme to LHA and to apolar and polar model surfaces

Assessing the Effect of Humic Acid Redox State on Organic Pollutant Sorption by Combined Electrochemical Reduction and Sorption Experiments

Natural Organic Matter (NOM) is a major sorbent for organic pollutants in soils and sediments. While sorption under oxic conditions has been well investigated, possible changes in the sorption capacity of a given NOM induced by reduction have not yet been studied. Reduction of quinones to hydroquinones, the major redox active moieties in NOM, increases the number of H-donor moieties and thus may affect sorption. This work compares the sorption of four nonionic organic pollutants of different polarities (naphthalene, acetophenone, quinoline, and 2-naphthol), and of the organocation paraquat to unreduced and electrochemically reduced Leonardite Humic Acid (LHA). The redox states of reduced and unreduced LHA in all sorption experiments were stable, as demonstrated by a spectrophotometric 2,6-dichlorophenol indophenol reduction assay. The sorption isotherms of the nonionic pollutants were highly linear, while paraquat sorption was strongly concentration dependent. LHA reduction did not result in significant changes in the sorption of all tested compounds, not even of the cationic paraquat at pH 7, 9, and 11. This work provides the first evidence that changes in NOM redox state do not largely affect organic pollutant sorption, suggesting that current sorption models are applicable both to unreduced and to reduced soil and sediment NOM

Adsorption of Insecticidal Cry1Ab Protein to Humic Substances. 2. Influence of Humic and Fulvic Acid Charge and Polarity Characteristics

Assessing the fate and potential risks of transgenic Cry proteins in soils requires understanding of Cry protein adsorption to soil particles. The companion paper provided evidence that patch-controlled electrostatic attraction (PCEA) and the hydrophobic effect contributed to Cry1Ab protein adsorption to an apolar humic acid (HA). Here, we further assess the relative importance of these contributions by comparing Cry1Ab adsorption to seven humic substances varying in polarity and charge, at different solution pH and ionic strength, <i>I</i>. Cry1Ab adsorption to relatively apolar HAs at <i>I</i> = 50 mM exhibited rapid initial rates, was extensive, and was only partially reversible at pH 5–8, whereas adsorption to more polar fulvic acids was weak and reversible or absent at pH >6. The decrease in adsorption with increasing HS polarity at all tested pH strongly supports a large contribution from the hydrophobic effect to adsorption, particularly at <i>I</i> = 50 mM when PCEA was effectively screened. Using insect bioassays, we further show that Cry1Ab adsorbed to a selected HA retained full insecticidal activity. Our results highlight the need to consider adsorption to soil organic matter in models that assess the fate of Cry proteins in soils