Protection of HT-22 Neuronal cells Against Glutamate Toxicity Mediated by Antioxidative Activity of Pueraria Candollei var Mirifica Extract

Abstract

Pueraria candollei var. mirifica (P. mirifica) known locally as Kwao Kruae Kao, has long been used in Thai folk medicine as a rejuvenating drug. Extracts of P. mirifica roots have been shown to contain many estrogenic-like isoflavonoids and presumably antioxidative compounds which may play a part in the anti-aging effects of this plant. In this research, various extracts of P. mirifica root powder prepared from different solvents, including hexane (PMH), ethyl acetate (PME), methanol (PMM) and water (PMW) were screened for their antioxidative activities by using 2,2-diphenyl-l-picrylhydrazyl (DPPH) assay. The extracts of PMM and PMW exhibited good antioxidative activities and were further investigated for their neuroprotective activities against glutamate-induced toxicity in HT-22 cells. PMM at the concentrations of 50 and 100 µg/ml completely inhibited glutamate toxicity similar to Trolox at the concentration of 264 µg/ml. PMW, on the other hand, exhibited no protection against glutamate toxicity. Further investigation on the radical scavenging property of PMM in HT-22 cells was performed by using the Oxygen Radical Absorbance Capacity (ORAC) assay for peroxyl radical and the dichlorofluorescin diacetate assay for hydrogen peroxide. It was found that the antioxidative activity of PMM against peroxyl radical was relatively low, whereas the radical scavenging property of PMM against hydrogen peroxide was very high. Therefore, the neuroprotective mechanism of PMM was proposed to be mediated through the antioxidative activity against hydrogen peroxide or the radical formed by hydrogen peroxide not peroxyl radical. Based on these results, it was concluded that PMM at the concentrations of 50 and 100 µg/ml exhibits significant neuronal cell protection against glutamate toxicity which could be the result of synergistic mechanism of many substances in the PMM extract including daidzein and genistein. The protection mechanism of PMM appears to be mediated via its antioxidative mechanism against hydrogen peroxide or the radical formed by hydrogen peroxide.