Antioxidant and semicarbazide-sensitive amine oxidase inhibitory activities of alginic acid hydroxamates

The commercial polysaccharides of alginic acid (medium (3500 cps, 2% solution) and low (250 cps, 2% solution) viscosities) were esterified with acidic methanol (1 mmol L^-1 HCl) at 4°C with gentle stirring for 5 days to obtain methyl esters of medium-viscosity alginic acid (ME-MVA) and low-viscosity alginic acid (ME-LVA). These ME-MVA and ME-LVA were reacted with alkaline hydroxylamine to obtain medium-viscosity alginic acid hydroxamates (MVA-NHOH) and LVA-NHOH. The percentages of hydroxamic acid content in MVA-NHOH and LVA-NHOH were calculated as 25% and 20%, respectively. The hydroxamate derivatives of alginic acid were used to test the antioxidant and semicarbazide-sensitive amine oxidase (SSAO) inhibitory activities in comparison with original materials (MVA and LVA). The half-inhibition concentrations, IC₅₀, of scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) were 24.5 and 29.8 μg mL^-1 for MVA-NHOH and LVA-NHOH, respectively. However, few scavenging activities of the MVA and LVA were found at the same concentrations. The IC₅₀ of the positive control of butylated hydroxytoluene was 5 μg mL^-1. The scavenging activity of DPPH radical was pH-dependent, and the optimal pH for both of MVA-NHOH and LVA-NHOH was the Tris-HCl buffer (pH 7.9). Using electron spin resonance (ESR) to detect the activity of scavenging hydroxyl radicals, both alginic acid hydroxamates showed dose-dependent scavenging activities, and the IC ₅₀ was 90 and 92 μg mL^-1, respectively, for MVA-NHOH and LVA-NHOH. Both alginic acid hydroxamates also exhibited protection against hydroxyl radical-mediated DNA damage. Both MVA-NHOH and LVA-NHOH showed dose-dependent inhibitory activities against bovine SSAO (2.53 units); the IC₅₀ was 0.16 and 0.09 μg mL^-1, respectively, for MVA-NHOH and LVA-NHOH, compared with 3.8 μg mL^-1 of semicarbazide (positive controls). Amine oxidase activity staining also revealed that both MVA-NHOH and LVA-NHOH exhibited SSAO inhibitory activities. Both MVA-NHOH and LVA-NHOH showed mixed non-competitive inhibition against bovine SSAO. It was found that the V′_max value was reduced and the K′_m value was either increased (added MVA-NHOH, 0.05 μg mL^-1) or reduced (added LVA-NHOH, 0.11 μg mL^-1) in the presence of alginic acid hydroxamate.