Superoxide scavenging and antiglycation activity of rhinacanthins-rich extract obtained from the leaves of Rhinacanthus nasutus

Background: Oxidative stress and nonenzymatic protein glycation lead to serious diabetic complications that increase the risk of mortality. Rhinacanthus nasutus leaf crude extracts are previously reported for their antidiabetic, antiglycation, and antioxidant potential. Objective: The present study was performed to prepare a standardized rhinacanthins-rich extract (RRE) and evaluate its superoxide scavenging and antiglycation effects as compared to its marker compounds, namely, rhinacanthin-C (RC), rhinacanthin-D (RD), and rhinacanthin-N (RN). Materials and Methods: RRE was obtained by microwave-assisted green extraction along with a simple step of fractionation using Amberlite^® column. RC, RD, and RN were isolated from the RRE using silica gel column chromatography. Superoxide scavenging activity was performed by cyclic voltammetry, and fructose-mediated human serum albumin glycation model was used for antiglycation activity. In silico studies were conducted to identify the structure-activity relationships of rhinacanthins. Results: On the basis of kinetic measurements, RRE exhibited the most potent antioxidant activity via E_rC_i mechanism, with a 50% inhibitory concentration (IC₅₀) value of 8.0 μg/mL, antioxidant capacity of 39439 M⁻¹, and binding constant of 45709 M⁻¹. Antiglycation assay showed that RRE exhibited almost equivalent glycation inhibitory effect to that of RC, with IC₅₀values of 39.7 and 37.3 μg/mL, respectively, but higher than that of RD (IC₅₀ of 50.4 μg/mL), RN (IC₅₀ of 89.5 μg/mL), as well as the positive control, rutin (IC₅₀of 41.5 μg/mL). Conclusions: The potent superoxide scavenging and albumin glycation inhibitory effect of RRE rationalized its therapeutic application in various chronic diseases, especially in the complications of diabetes.