ANTI-PLASMODIAL AND ANTI-INFLAMMATORY ACTIVITIES OF ETHANOL EXTRACT AND ETHYL ACETATE FRACTION OF Psidium guajava Linn LEAVES IN MICE AND RATS

Abstract

Inflammation during malaria is a determinant for the progression of plasmodial infection that causes health burdens in humans. Emergence of parasite resistance, absence of dual antiplasmodial and anti-inflammatory properties of current antimalarials necessitates search for new therapeutic strategies. Psidium guajava (Pg) leaves is commonly used in ethno-medicine for the treatment of malaria and other ailments, however, there is dearth of information of its antiinflammatory activity in malaria. This study was therefore designed to investigate the antiplasmodial and anti-inflammatory activities of Pg leaves in mice and rats. Fresh Psidium guajava leaves were authenticated at Forest Herbarium Ibadan (FHI. 110758), airdried, pulverized, extracted with 70% ethanol (EEPg), and partitioned sequentially into n-hexane (PgHXF), dichloromethane (PgDCF), ethyl acetate (PgEAF), and residual aqueous fractions (PgRAF). Ninety male mice infected with Plasmodium berghei (NK65) were used for prophylaxis, 4-Day SuppressiveTest (4-DST) and Curative Test (CT), respectively. For each test, thirty mice were divided into five groups (n=6); 1%Tween80 (control), EEPg (100, 200, 400mg/kg) and chloroquine (10mg/kg). Anti-inflammatory activities of EEPg were investigated in air-pouch model using thirty-six male rats allotted into six treatment groups (n=6); Normal saline (10 mL/kg), lipopolysaccharide (100 ηg/kg), EEPg (100, 200, 400mg/kg) and indomethacin (10 mg/kg). Cytokines (TNF-α, IL-1β) and leucocytes differentials were determined in exudates and blood obtained from the rats. The EEPg fractions were evaluated in mice using 4-DST. The most active fraction PgEAF was evaluated using CT with parasitemia and Mean Survival Time (MST) determined. Blood, liver and spleen were harvested for haematology, liver function (AST and ALT), inflammatory profiles (TNF-α, IL-6, MPO, IFN-γ, and IgG) and markers of oxidative stress (GSH, MDA, SOD, and CAT). Histological analysis of liver and spleen were done using standard method. Data were analysed using ANOVA at α0.05. Chemo-suppression of EEPg in Prophylaxis (84.0, 93.5, 97.2%), 4-DST (32.5, 61.7, 77.3%), CT (59.4, 78.6, 81.0%) was dose-dependent relative to untreated control. Pouch exudates showed significant decrease in levels of TNF-α (30.17±2.53, 37.13±5.0, 28.605±0.59 vs 55.56±1.74pg/mg protein) and IL-1β (193.80±5.67, 158.205±0.94 vs 241.30±4.21pg/mg protein) in EEPg compared with control. The EEPg (400mg/kg) compared with control significantly reduced lymphocytes (35.75±4.87 vs 57.75±1.03) and neutrophils (40.0±3.63 vs 60.25±1.32) counts. The chemo-suppression in 4-DST were PgHXF (49.3%), PgDCF (68.0%), PgEAF (89.1%) and PgRAF (79.5%) relative to untreated control. In CT, PgEAF significantly increased MST (20.83±1.92, 21.5±2.01, 24.83±1.6days) compared with control (10.3±0.1days). The PgEAF (200mg/kg) significantly reduced serum AST (113.6±2.39 vs 123.4±1.07U/L), ALT (92.62±3.05 vs 103.7±2.71U/L) and IFN-γ (73.76±1.94 vs 88.51±0.95pg/mL), and hepatic TNF-α (38.96±1.78 vs 67.15±5.04pg/mg protein) and IL-6 (60.37±2.92 vs 94.89±0.25pg/mg protein), respectively. Packed cell volume (25.67±0.24 vs 20.33±0.24%), haemoglobin (9.77±0.09 vs 6.63±0.09g/dL), platelet (9.33±0.47 vs 4.00±0.00 x103/µL) and serum IgG (18.54±1.01 vs 10.45±1.47pg/mL) increased significantly compared with control. PgEAF significantly reduced hepatic MDA but increased GSH, SOD and CAT, respectively. The PgEAF prevented hepatic and splenic injury in infected mice when compared with untreated control. Ethanol extract of Psidium guajava leaves and its ethyl acetate fraction exhibited anti-plasmodial and anti-inflammatory activities by parasites suppression through immune-modulation and potentiated antioxidant activities.