Antioxidant and immune-enhancing potentials of leaf extract and active constituents of Millettia aboensis

Antioxidant and immune-enhancing potentials of ethanol leaf extract of Millettia aboensis (Hook F.) Baker (Leguminosae), fractions and isolated compounds were determined using in vitro and in vivo models. In vitro antioxidant activities were assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and hydrogen peroxide scavenging activity tests; while in vivo protection against oxidative damages was assessed by carbon tetrachloride (CCl₄)-induced liver damage and streptozotocin (STZ) induced systemic oxidative stress models. In vivo immune enhancing properties were monitored using primary and secondary immune responses to tetanus toxoid. Bioassay guided separations led to the isolation of compounds 1, 2 and 3. Their structures were elucidated by a combination of 1D and 2D NMR and mass spectrometry. In vitro inhibition of liver microsome lipid peroxidation was used to evaluate the antioxidant activity of compounds 1 and 2 while stimulation of specific T-lymphocytes was used for evaluating immune enhancing activity of compound 3. The extract exhibited both antioxidant and immune-enhancing properties however, antioxidant activity was prominent in the ethyl acetate fraction, while butanol fraction expressed more higher immune-enhancing activity. Structural elucidation revealed compounds as catechin (procyanidine A1) (1), epicatechin (procyanidine A2) (2) and quercetin-3-O-rutinoside (rutin) (3). Compounds 1 and 2 demonstrated strong inhibition of liver microsome lipid peroxidation, with EC₅₀ of 46 and 55 µg/mL respectively against 31 µg/mL of α-tocopherol. Compound 3 showed up-regulation of specific CD4+ lymphocytes with up to 38% stimulatory effect of IFNγ at 6.25 µg/mL compared to the baseline effect in DMSO control group. The extract, fractions and isolated compounds from M. aboensis expressed strong antioxidant and immune-enhancing properties which may be responsible for its ethnopharmacological use for general healing.