# The Effects of p-Coumaric Acid on the Quality of Cryopreserved Boar Spermatozoa

## TL;DR

Adding p-coumaric acid to boar semen freezing solutions improves sperm quality and fertility after thawing.

## Contribution

Identifies 90 μg/mL p-coumaric acid as the optimal concentration for enhancing boar sperm cryopreservation.

## Key findings

- 90 μg/mL p-coumaric acid significantly improved sperm motility, membrane integrity, and DNA quality after freezing.
- This concentration enhanced antioxidant activity and reduced oxidative stress markers in boar sperm.
- It also increased in vitro fertilization and embryo cleavage rates in porcine oocytes.

## Abstract

Artificial insemination technology is one of the important methods for the genetic improvement of excellent boars, and semen preservation is one of the key links of this technology. However, during the process of semen cryopreservation, the sperm plasma membrane contains a large amount of unsaturated fatty acids, which makes it vulnerable to oxidative stress. This leads to a decrease in the integrity of the sperm plasma membrane, acrosome, and DNA after thawing, thereby affecting the fertilizing ability of sperm. To improve sperm characteristics, protectants that enhance the quality of boar sperm after freeze-thawing are added to the freezing extender. PCA has moderate antioxidant activity and high chelating ability. Therefore, we investigated the effects of adding different concentrations of PCA (0, 30, 60, 90, and 120 μg/mL) to the protectant on the quality of boar sperm after freeze-thawing. We found that, within the tested concentration range, 90 μg/mL PCA exhibited the optimal effect: it significantly increased sperm motility and various movement parameters, as well as acrosome integrity, plasma membrane integrity, mitochondrial activity, and DNA integrity; enhanced total antioxidant capacity and the activities of antioxidant enzymes such as superoxide dismutase; reduced the contents of malondialdehyde and hydrogen peroxide; upregulated the expression of the anti-apoptotic protein BCL-2; downregulated the expression of the pro-apoptotic proteins BAX and Caspase-3; and also improved the in vitro fertilization rate of boar oocytes and the embryo cleavage rate. Our study indicates that adding PCA to the semen extender is beneficial for the cryopreservation of boar sperm, with 90 μg/mL being the optimal concentration, which provides support for the development of boar semen cryopreservation technology.

This research explored the effects of different concentrations of p-coumaric acid (PCA) on the quality of frozen-thawed boar semen. Boar sperm samples were pre-treated with different concentrations of PCA (0, 30, 60, 90, 120 μg/mL) prior to the freezing process. Subsequently, multiple parameters were analyzed post-freeze-thawing, including sperm morphological and kinetic characteristics, acrosome and membrane integrity, mitochondrial function, DNA integrity, antioxidant enzyme activities, the expression levels of the BCL-2, BAX, and Caspase-3 proteins, the in vitro fertilization rate of porcine oocytes, and the embryo cleavage rate. The findings indicated that, compared with the control group, the addition of 90 μg/mL PCA led to significant improvements in several key aspects. Sperm motility, average path velocity, straight-line velocity, curvilinear velocity, and beat cross frequency were all notably enhanced. Moreover, parameters related to sperm quality, such as acrosome integrity, plasma membrane integrity, mitochondrial activity, and DNA integrity, also showed significant increases (all p < 0.05). In terms of antioxidant capacity, the 90 μg/mL PCA treatment significantly elevated the total antioxidant capacity, as well as the activities of superoxide dismutase, glutathione peroxidase, and catalase. Simultaneously, it caused a significant reduction in the contents of malondialdehyde and hydrogen peroxide (p < 0.05). Regarding protein expression, the addition of 90 μg/mL PCA significantly upregulated the expression level of the BCL-2 protein, while downregulating the relative expression levels of BAX and Caspase-3 (p < 0.05). Additionally, this concentration of PCA significantly improved the in vitro fertilization rate of porcine oocytes and the embryo cleavage rate (p < 0.05). In conclusion, incorporating PCA into the semen extender can potentially be advantageous for the cryopreservation of boar sperm, with 90 μg/mL being the optimal concentration.