Physicochemical Characterization and Antioxidant Activity of Humic Acids Isolated from Peat of Various Origins

Although humic acids (HAs) from peat exhibit various therapeutic properties, there is little information available concerning their physicochemical and antioxidant properties. To address this issue, nine different types of peat, including oligotrophic, mesotrophic, and minerotrophic peat samples, were used for isolation of HA fractions by basic (HAb) and pyrophosphate (HAp) extractions. Physical parameters of the HAs were analyzed by UV-Vis, fluorescent, infrared (IR), and electron paramagnetic resonance (EPR) spectroscopy. Average Mr of the fractions ranged from 17.2 to 39.7 kDa, while their humification index (HIX) varied from 0.49 to 1.21. HAp fractions had a higher content of aromatic structures compared to HAb fractions. Moreover, HAp fractions had a significantly higher content of phenolic OH groups (3.6 ± 0.5 mmol/g) versus HAb (3.1 ± 0.5 mmol/g). All HA fractions exhibited antioxidant activity in radical scavenging and electrochemical assays, and their EPR signal had a single line with g = 2.0035, which is consistent with semiquinone type radicals. Furthermore, the HIX was found to be important in determining the number of semiquinone-type free radicals in the HA structures. Overall, these data provide a molecular basis to explain at least part of the beneficial therapeutic properties of peat-derived HAs

Quantitative Structure-Activity Relationship, Ontology-Based Model of the Antioxidant and Cell Protective Activity of Peat Humic Acids

Peat humic acids are well known for their wide range of biological effects which can be attributed to the complex chemical structure of naturally occurring humic substances. One of the promising tools is an ontology-based quantitative analysis of the relationship between physical and chemical parameters describing a chemical structure of peat humic acids and their biological activity. This article demonstrates the feasibility of such an approach to estimate the antioxidant and cell protective properties of the peat humic acids. The structural parameters of the peat humic acids were studied by electronic, fluorescence, infrared, 13C-NMR spectroscopy, titrimetric analysis, elemental C,H,N, and O- analysis, and gel chromatography. Antioxidant and antiradical activities were assessed by physicochemical methods of analysis: electronic paramagnetic resonance, cathodic voltammetry, ABTS•+ scavenging, assay of DPPH radical-scavenging activity, assay of superoxide radical-scavenging activity, iron chelating activity, and scavenging of hydroxyl radicals. Cytoprotective activity was evaluated by the neutral red-based cytotoxicity test in 3T3-L1 cell culture in a wide range of concentrations. Assessment of intracellular ROS production was carried out using a 2,7-dichlorodihydrofluoresceindiacetate (DCFDA) fluorescent probe. Intracellular ROS production was induced using two common prooxidants (tert-butyl hydroperoxide, Fe2+ ions). We suggested an ontology-based model for the antioxidant and cytoprotective activity of humic acids based on experimental data and numerical models. This model establishes the way to further research on the biological effects of humic acids and provides a useful tool for numerical simulation of these effects. Remarkable antioxidant and cell protective activity of humic acids makes them a promising natural source of new pharmaceutical substances that feature a wide range of biological effects

Enhanced Antioxidant Activity and Reduced Cytotoxicity of Silver Nanoparticles Stabilized by Different Humic Materials

The current article describes the biological activity of new biomaterials combining the “green” properties of humic substances (HSs) and silver nanoparticles. The aim is to investigate the antioxidant activity (AOA) of HS matrices (macroligands) and AgNPs stabilized with humic macroligands (HS-AgNPs). The unique chemical feature of HSs makes them very promising ligands (matrices) for AgNP stabilization. HSs have previously been shown to exert many pharmacological effects mediated by their AOA. AgNPs stabilized with HS showed a pronounced ability to bind to reactive oxygen species (ROS) in the test with ABTS. Also, higher AOA was observed for HS-AgNPs as compared to the HS matrices. In vitro cytotoxicity studies have shown that the stabilization of AgNPs with the HS matrices reduces the cytotoxicity of AgNPs. As a result of in vitro experiments with the use of 2,7-dichlorodihydrofluorescein diacetate (DCFDA), it was found that all HS materials tested and the HS-AgNPs did not exhibit prooxidant effects. Moreover, more pronounced AOA was shown for HS-AgNP samples as compared to the original HS matrices. Two putative mechanisms of the pronounced AOA of the tested compositions are proposed: firstly, the pronounced ability of HSs to inactivate ROS and, secondly, the large surface area and surface-to-volume ratio of HS-AgNPs, which facilitate electron transfer and mitigate kinetic barriers to the reduction reaction. As a result, the antioxidant properties of the tested HS-AgNPs might be of particular interest for biomedical applications aimed at inhibiting the growth of bacteria and viruses and the healing of purulent wounds

Coal-Derived Humic Substances: Insight into Chemical Structure Parameters and Biomedical Properties

An investigation was carried out on humic substances (HSs) isolated from the coal of the Kansk-Achinsk basin (Krasnoyarsk Territory, Russia). The coal HSs demonstrate the main parameters of molecular structure inherent to this class of natural compounds. An assessment was performed for the chemical, microbiological, and pharmacological safety parameters, as well as the biological efficacy. The HS sample meets the safety requirements in microbiological purity, toxic metals content (lead, cadmium, mercury, arsenic), and radionuclides. The presence of 11 essential elements was determined. The absence of general, systemic toxicity, cytotoxicity, and allergenic properties was demonstrated. The coal HS sample was classified as a Class V hazard (low danger substances). High antioxidant and antiradical activities and immunotropic and cytoprotective properties were identified. The ability of the HS to inhibit hydroxyl radicals and superoxide anion radicals was revealed. Pronounced actoprotective and nootropic activities were also demonstrated in vivo. Intragastric administration of the HS sample resulted in the improvement of physical parameters in mice as assessed by the “swim exhaustion” test. Furthermore, intragastric administration in mice with cholinergic dysfunction led to a higher ability of animals with scopolamine-induced amnesia to form conditioned reflexes. These findings suggest that the studied HS sample is a safe and effective natural substance, making it suitable for use as a dietary bioactive supplement