PHYTOCHEMICAL AND NEUROPROTECTIVE INVESTIGATIONS OF LEAF EXTRACTS OF SAUSAGE TREE (Kigelia africana LAM, BENTH)

Abstract

Neurodegenerative diseases are permanent conditions resulting from progressive deterioration and/or death of nerve cells; they are major causes of disability and premature death around the globe. They share common pathophysiological features like oxidative damage, mitochondrial dysfunction, and neurotransmitter dysregulation among others. Many orthodox drugs used for management of neurodegenerative diseases are expensive, possess and elicit toxic side effects, thereby necessitating sourcing for alternative treatments. Kigelia africana (Lam) Benth, belongs to the family Bignoniaceae which contain many phytoconstituents, such as flavonoids, saponins among others. Several pharmacological properties have been attributed to the plant but there is no data on its neuroprotective properties, hence the study. The phytochemical screening and in vitro antioxidant activities were evaluated by standard protocols. Sub-chronic toxicological investigation of Kigelia africana leaf extract (KAE) was carried out by oral administration of 100-600 mg/kg bw to male Wistar rats for 14 consecutive days, and biochemical parameters were assayed. The flavonoid rich fraction of KAE (FKAE) was obtained through acid hydrolysis. Thereafter, appropriate doses were selected for the study of protective effect of KAE or FKAE on AlCl3-induced Alzheimer’s disease in vivo. Animals were pretreated with KAE or FKAE for 14 days prior to administration of 17 mg/kg of AlCl3 except the control and positive control (AlCl3) groups, followed by combination of AlCl3 and KAE or FKAE treatments for four successive weeks. During this period, animals were scored for neurological deficits, they were then sacrificed; brain homogenate was used to assess antioxidant status, electrolyte status, and neurochemical alteration. Histopathological investigation was carried out on the second part of the brain tissues obtained. Solvent-fractions of KAE were obtained with vacuum-liquid chromatographic technique i.e. n-hexane (100 %), n-hexane:dichloromethane (1:1), dichloromethane:ethylacetate (1:1) and ethylacetate:methanol (1:1). Each solvent-fraction was tested for bioactivity; fraction with the best activity was assayed for its activities on mitochondrial integrity. This fraction was further subjected to gradient elution column chromatography for purification to obtain pure compounds. The structures of the pure compound(s) isolated were elucidated using 1D and 2D nuclear magnetic resonance (NMR) spectroscopy and other techniques. Data obtained were analyzed using suitable software. The phytochemical constituents in KAE are saponins, tannins, flavonoids, alkaloids, terpenoids, steroids and cardiac glycoside; while FKAE mainly possesses flavonoids and traces of tannins. In-vitro antioxidant assays such as total antioxidant capacity (TAC), reductive potential, DPPH radical scavenging, metal chelation, hydroxyl radical scavenging, nitic oxide radical scavenging capacities; demonstrated significant antioxidant potentials of the extract. HPLC-DAD fingerprinting of KAE and FKAE revealed the presence of flavonoids (catechin, rutin, isoquercitrin, quercetin, kaempferol), phenolics acids (gallic, chlorogenic, caffeic, ellagic, rosmarinic acids), carotenoids (Tocopherol, β-carotene). Sub-chronic toxicity studies showed that KAE was toxic at high concentrations (> 100 mg/kg), but optimally protective at 100 mg/kg bw. Results of the AlCl3–intoxication study revealed that KAE, FKAE and Rivastigmine are comparably protective against AlCl3 neurotoxicity as revealed in the neurobehavioural assessments, cholinesterases assessments, biochemical markers, electrolyte status and histopathological analysis of the brain hippocampal section. DCM:EtOAc (1:1) fraction gave the best activity in the in-vitro biological assays conducted, as well as in the protection of mitochondrial integrity. Three pure compounds namely; quercetin, rutin and luteolin were isolated from DCM:EtOAc (1:1) fraction. These structures were elucidated and characterized using NMR and Mass spectroscopies. This buttressed the potency of FKAE used in in vivo studies to combat neurotoxicity. The results of this study present KAE as a good source of raw material in the development of drugs for the management of neurological disorders.