Biochemical and Physiological Effects of Carissa Spinarum L. Extracts on Oxidative Stress-induced Infertility in the Testis of Male Wister Rats

Abstract

The exploitation of natural plant products in the management of infertility has tremendously increased, leading to the expansion of research to develop drugs targeting various types of male infertility. Infertility affects human reproduction and expensive medical approaches such as in vitro fertilization (IVF) and Intracytoplasmic sperm injection (ICSI) have been in use for its management. There is a need to develop economically friendly and readily available remedies with minimal side effects from sources such as medicinal plants. This study aimed to evaluate the reversal effects of Carissa spinarum L. aqueous root extracts on oxidative stress-induced infertility in male Wister rats. C. spinarum leaves and roots were first extracted with ethanol, methanol, Hydroethanol (70%), Hydromethanol (70%), n-hexane, acetone, distilled water by cold maceration for 72 hours, and by hot maceration for 2 hours at 60 ℃ for comparative phytochemical analysis. Total phenolic content (TPC), tannins content (TTC), flavonoids content (TFC), and alkaloids content (TAC) were spectrophotometrically determined as well as the in vitro antioxidant activities; Ferric reducing antioxidant power (FRAP) and hydrogen peroxide radical scavenging (HPRS). Thirty (30) male Wister rats (of age 70-90 days, weighing 130 – 180 g) were used in in vivo studies using aqueous root extracts prepared by hot maceration. The animals were randomly divided into six experimental groups where group 1 received distilled water, group 2 received 400mg/kg bw of metronidazole, group 3 received 400 mg/kg/bw metronidazole + 250 mg/kg/bw extract, group 4 received 400 mg/kg/bw metronidazole + 500 mg/kg/bw, group 5 received 400 mg/kg/bw metronidazole + 1000 mg/kg/bw while group 6 received 400 mg/kg/bw Vitamin E + 0.36mg/kg/bw of Sustanon weekly intramuscular injection for 8 weeks, with all other treatment administered following oral route. At the end of the 8th week, animals were sacrificed in a humane manner. The epididymis (for sperm retrieval at the cauda part) and the testis (for biochemical assays and histology) were harvested from each rat. Sperm characteristics were determined using a light microscope while the biochemical assays were assessed spectrophotometrically. The obtained numerical data was subjected to a one-way Analysis of Variance (ANOVA), followed by Turkey’s post Hoc analysis. Methanol, ethanol, acetone, and distilled water (hot) showed high TPC, TTC, TFC, TAC, and high antioxidant activity in both the leaves and roots. The leaf extracts generally showed higher levels of phytochemicals and antioxidant activity than the roots. All three doses of C. spinarum showed a significant increase in percent weight gain of the body, testicular, and epididymal weight paralleled to the negative control (P < 0.05) with treatment groups 3 and 4 showing normal testicular histology comparable to the normal and positive controls. Sperm concentration, morphology, and viability were significantly high among the extract treatment animals, compared to the negative control (P < 0.05). Malondialdehyde content was significantly higher among group 2 compared to the medium dose extract treated (group 4) (P < 0.05). All extract treated groups recorded high total testicular protein concentration (P > 0.05), catalase (P < 0.05) and peroxidase (P > 0.05) enzyme activities, compared to the negative control (group 2). The current findings reveal that C. spinarum contains medicinal compounds (phenolics, flavonoids, tannins, and alkaloids) and strong antioxidant activity in the leaves compared to the roots with suitable solvents of extraction. The compounds were confirmed to exert their pharmacological effects in vivo through reversal of oxidative stress-induced infertility in male Wister rats, thus indicating the potential in the treatment of male infertility arising from oxidative stress-related factors.