Considerable success has been achieved in semen preservation of large animals over the past few decades; however, this is not yet much achieved in small ruminants. The decline, which occurs in semen quality during preservation, has not been solved out. Antecedent investigations have investigated that the inclusion of antioxidants can drastically enhance the semen quality by minimizing the ROS production (Maxwell and Salamon 1993). Earlier, various studies reported that addition of antioxidants in semen diluter improves the semen quality (Gangwar et al 2014, Gangwar et al 2015, Gangwar et al 2018). But now a days people are trying natural antioxidants in semen extenders (Wen et al 2019) and extract from Asparagus racemosus (Shatavari) roots have been used in many investigations as antioxidants (Kamat et al. 2000; Goyal et al., 2003). However, there is no evident research findings with respect to the protective effect of Shatavari aqueous extract on buck sperm as well as inhibition of lipid peroxidation. This is the first research work carried out to examine the semen quality and antioxidant activity in the bucks after supplementation of Shatavari aqueous extract in buck semen diluter.
In the present investigation study, herbal additives significantly improved the progressive motility, sperm viability, normal sperm morphology, acrosomal integrity and plasma membrane integrity. Moreover, we found reduced lipid peroxidation of sperm plasma membrane reflected by MDA level in Shatavari extract supplemented group. Though, this type of investigation has not been done or performed with herbal antioxidant supplementation in buck semen diluter in comparison with our results. Very little amount of data is present with regard to the specific use of herbs in laboratory animals, boar and human beings (Kopalli et al., 2015; Kumar et al. 2018; Ansari and Khan, 2017). Thakur et al. (2009) reported that Asparagus racemosus, Chlorophytum borivilianum, and rhizomes of Curculigo orchioides improved the mating behavior and aphrodisiac activity in male rats. During semen cryopreservation, there is imbalance between antioxidant and pro-oxidant activities, with the excess production of ROS and oxidation of polyunsaturated fatty acids in buck sperm membranes leading to formation of lipid peroxides, and ultimately formation of cytotoxic metabolic products, especially MDA (Zhao et al., 2009). In the current investigation, based on the negative correlation between sperm characteristics and MDA, however it is positively correlated with sperm abnormalities (Table 7), hence it was concluded that Shatavari root extract adequately superoxide anions, and hindered lipid peroxidation.
Similar to our findings, Wen et al. (2019) reported that sperm motility, acrosomal integrity, mitochondrial membrane potential, plasma membrane integrity, total antioxidative activity in the 30 mg/L GSPE (Grape seed procyanidin extract) group was significantly enhanced, whereas MDA content was lower than as compared to the control group (P < 0.05). They also reported that AI results with GPSE supplemented semen showed higher litter size as compared to the control group (P < 0.05). Similarly, Zhao et al. (2009) reported the significant correlation between Rhodiola sacra aqueous extract and concentrations of GSH and MDA in frozen-thawed boar semen. As we know, that oxidative stress is a well-documented inducer of apoptosis and lipid peroxidation can initiate aging and a diminished longevity of the cryopreserved spermatozoa.
The results of the current study indicate that there is overall increase in sperm motility, sperm viability, acrosomal integrity, plasma membrane integrity in treatment groups as compared to the control groups whereas, there is significant increase in number of abnormal spermatozoa and MDA level in control group as compared to treatment groups which shows higher peroxidative damage in control group (Fig. 1). In our previous study, we found that dietary azolla supplementation significantly improves the semen quality and libido in bucks (Gangwar et al., 2019). Earlier study has shown that buck sperm motility was 68.59% after three days of storage at 5℃ (Wen et al., 2019); it reached 78.2% (a significant increase) after three days of liquid preservation with Shatavari supplementation. Hence, Shatavari extract could be able to protect the buck sperm from various types of cryo-damages.
Malo et al. (2011) found that by adding rosemary extract into the extender effectively increased the sperm penetration and oocyte cleavage rate, and also improved the effectiveness of the fertilization system. In addition to this, they also found that supplementation of exogenous rosemary to the freezing medium enhanced post-thawed semen quality in boar. Similarly, Zhao et al. (2009) reported that Rhodiola sacra aqueous extract (RSAE) exhibited strong scavenging activity against superoxide anion radical. They observed significant improvement in progressive motility, HOST response, fertility, and safeguarding the functional integrity of sperm plasma membrane. Antecedent findings indicated that the number of hypo-osmotic swelled spermatozoa is positively correlated with the oocyte penetration rate in human oocytes.
In Congruent with our results, El-Sheshtawy et al. (2016) demonstrated that addition of bull semen extender with 10% and 20% Pomegranate Juice provides adequate chilling, and boosted frozen-thawed semen quality. Similarly, Khan et al. (2017) reported that addition of green tea extract in semen diluter exhibited remarkable or noteworthy response on the post-thawed spermatozoa motility, viability and membrane integrity of Achai bull. Vahedi et al. (2018) found that supplementation of Thymus vulgaris extract at the rate of 4–8 ml/dl of diluter improves the attributes of ram sperm after freezing and thawing. Addition of 5% Tribulus terrestris extract in semen diluter improved the sperm motility in Afshari rams at 5°C (Pour et al., 2015). The findings of previous investigation reported that T. terrestris extract had a noteworthy influence on the sperm motility and viability in human. However, in vitro studies on human reported that T. terrestris extract inclusion in extender had no effect on DNA fragmentation of (Kumar et al., 2018). Similarly, Mehdipour et al. (2016) reported that supplementation of Camellia sinensis extract at level of 10 mg/L can enhance post-thaw ram semen quality, when it is cryopreserved in a soybean lecithin extender.
In the present investigation, the enhancement of semen quality and antioxidant activity could be due to the richness of A. racemosus in advantageous polycarbohydrates, i.e., fructo-oligosaccharide with an average degree of polymerization. A. racemosus is also rich in sterols and steroidal saponins, Shatavarins I-IV (Alok et al., 2013). Shatvarin I is the main glycoside with 3-glucose and rhamnose moieties attached to sarsapogenin, effective in establishing the sufficient availability of hormone to gonads (Kukasawa et al., 1994). Diverse working of the herb can be a primary ground for a remarkable enhancement in the sexual attributes. After analyzing the results of the current investigation, it is concluded that A. racemosus has aphrodisiac property, and has potential to treat sexual dysfunction.