EFFECTS OF METHANOL EXTRATOF plukenstia conophora SEED AND 4H-PYRAN-4-ONE2,3-DIHYDRO-3,5-DIHYDROXY-6-METHYL-ON CADIUM CHLORINE-INDUCED RESPODUCTIVE TOXICITY IN WISTAR RATS

Abstract

Infertility is a current global issue with diverse treatments. However, many infertile people especially in developing countries resort to folkloric treatment due to unyielding results from synthetic medications. Plukenetiaconophora(PC) also called walnut is used in traditional medicine in Nigeria to promote fertility. Due to paucity of scientific basis on its fertility claims, this study sought to investigate the probable effects of PC and its flavonoid fraction; 4H-Pyran-4-one-2,3 dihydro-3,5-dihydroxy-6-methyl (DDMP) on cadmium chloride-induced male reproductive toxicity. The PC seeds were collected from Okeho, Oyo state, and identified at Forest Research Institute of Nigeria, Ibadan (FHI No. 109997). The air-dried seeds were extracted by cold maceration in methanol, and evaporated to dryness with rotary evaporator. Isolation and characterisation of the flavonoid fraction were done using column chromatography, thin layer chromatography,gas chromatography mass spectrometry, Fourier transform infrared and ultraviolet spectrophotometers.Seventy-two male Wistar rats (150-190g) were randomly divided into 12 groups (n=6). Administration was done daily for 54 days except for cadmium chloride (CdCl2) as follows: Group 1= Control (normal saline), Group 2= CdCl2 (2mg/kg, i.p.), Group 3= Vitamin E (200mg/kg, p.o), Group 4 and 5= PC (100 and 200 mg/kg, p.o) alone, Group 6= CdCl2(2mg/kg, i.p.)and vitamin E (200mg/kg, p.o), Group 7 and 8= CdCl2 (2mg/kg, i.p.)with PC (100 and 200 mg/kg, p.o), respectively, Group 9 and 10= DDMP (25 and 50 mg/kg, p.o) respectively, Group 11 and 12= CdCl2 (2mg/kg, i.p.)with DDMP (25 and 50 mg/kg, p.o), respectively. Sperm analyses were done by microscopy. Testicular, epididymal antioxidant enzymes and Na+/K+, Ca2+ and Mg2+ ATPases were measured by spectrophotometry. Testosterone was measured using ELISA. The tissues of testis and seminal vesicle were stained with Haematoxylin and Eosin, and viewed under light microscope. Immunohistochemistry (Bax and Bcl2proteinexpression) was done by Avidin biotin Immunoperoxidase method. Testicular DNA fragmentation was by Agarose gel electrophoresis. Data were subjected to descriptive statistics and analysed using ANOVA at α0.05. Isolation and characterisation of PC yielded 56.8 % DDMP. Increase in sperm count, motility and viability observed in group 8 were (61.00±7.57 106cell/mL; 34.00±7.48 %; 26.00±1.87%) and group 12 were(85.00±5.90 106cell/mL; 68.40±9.80 %; 84.60±5.44 %) compared with group 2 (24.20±9.70 106cell/mL; 14.40±3.08%; 8.60±1.02%), respectively. Increases were observed in antioxidant enzymes and testosterone levels ingroups 8 and 12.There were increases in testicular Na+/K+, Ca2+, Mg2+ATPases of group 8 (38.66±10.20; 98.36±18.53; 71.64±3.94), and group 12 (43.19±5.61; 64.30±7.36; 78.32±17.98) compared with group 2(7.33±1.02; 23.22±3.79; 24.40±6.25 µmole/pi/mg pr-/hr x103), respectively. Histology of testes and seminal vesicle showed degenerated cells in group 2 which were significantly restored in groups 8 and 12. There were significant reduction in the expressions of Bax to Bcl2 ratio in Leydig cells, and reduction in total cellular DNA fragmentation of groups 8 and 12 were observed compared with group 2. Plukenetiaconophora alleviated cadmium chloride-induced testicular toxicity in Wistar rats. The flavonoid isolate of Plukenetiaconophora(4H-Pyran-4-one 2,3 dihydro-3,5-dihydroxy-6-methyl)could be responsible for the observed activity.