**10.2 Effect of L-Cysteine on frozen boar semen**

L-cysteine, an amino acid containing a sulphydryl group, is a precursor of intracellular glutathione biosynthesis. L-cysteine plays a role in the intracellular protective mechanism against oxidative stress, membrane stabiliser and capacitation inhibitor (Johnson et al., 2000). Glutathione is the most common non-thiol protein in mammalian cells which protects plasma membrane from LPO, scavenges superoxide and minimized O2 formation. It has been demonstrated that the supplementation of *L-cysteine* in the semen extender prevents the loss of sperm motility by minimizing hydrogen peroxide of FT semen in the bull (Bilodeau et al., 2001). Funahashi and Sano (2005) found that the supplement of L-Cysteine for 5 mM improved the viability and functional status of the boar spermatozoa during chilled storage.

During the past few years, many studies have been carried out by supplementation of various antioxidants (e.g. Vitamin E, Glutathione, Taurine) in the freezing extenders of frozen boar semen in order to minimize the detrimental effect of ROS which occurred during the freezing process (Pena et al., 2003; Roca et al., 2004; Breininger et al. 2005; Gadea et al., 2005). Funahashi and Sano (2005) demonstrated that supplement of L-cysteine (5 mM) could improve the viability and progressive motility in fresh boar semen, and also the same case found in frozen bovine semen (Bilodeau et al., 2001). This L-cysteine is also improve survival time of semen and sperm chromatin structure in fresh chilled boar semen at 15oc

Cryopreservation of Boar Spermatozoa: An Important Role of Antioxidants 155

capacitation, inhibiting and acrosome reaction and binding ability of its protein to the sperm plasma membrane) of seminal plasma in protecting sperm from cryoinjury. It has been demonstrated in rams that seminal plasma protein is able to revert the cold shock damage on the sperm membrane (Barrios et al. 2000). This effect has also been reported in boars, in that holding boar spermatozoa in its seminal plasma before cooling and freezing defends against the cold shock (Pursel et al. 1973). In addition, it has been shown that adding seminal plasma to the post-thawing solution increased the percentage of stallion sperm motility (Alghamdi et al. 2005). In dogs, it has been shown that frozen semen diluted with their prostatic fluid yielded a higher post-thawing motility (Rota et al. 2007). As a result, it seems likely that seminal plasma or prostatic fluid constituents have a positive effect on post-thawing sperm motility. In addition, semen with a high percentage of progressive motility illustrates their plasma membrane integrity and superb metabolism (Johnson et al., 2000) and also reflects their ability for fertilization (Vyt et al. 2004; Estienne et al. 2007). It can be hypothesized that the presence of supernatant (semen plasma plus semen extender)

during the thawing process may improve frozen-thawed boar sperm motility.

ovulation time in sows (Waberski et al.,1995; Kaeoket and Tummaruk, 2002b).

et al., 1996; Kaeoket et al., 2002a, 2005).

**of frozen boar semen** 

Seminal plasma is the liquid constituent of an ejaculate, comprising a combination of fluids secreted by the male accessory glands (i.e. seminal vesicle) during an ejaculation. It is evidence that seminal plasma able to arrest or reverse cryoinjury and perhaps extend the longevity of the sperm by inhibiting or reversing capacitation and acrosome reaction (Suzuki et al., 2002). Besides, seminal plasma appears to play an important role in the female reproductive tract after insemination, e.g. attenuate the post insemination inflammatory response in the uterus of the sow which may influence the chances of conception (Rozeboom et al.,1999) and its component such as hormone estrogen may also resulted in a release of prostaglandins from the pigs endometrium to the utero-ovarian veins and lymphatic vessels which in turn decrease duration time from standing oestrus to ovulation in gilts (Clause et al.,1987,1990; Weitze et al.,1990), Therefore, it seems likely that seminal plasma constituents (both the oestrogen and the protein fraction) have an effect on

Generally, limitations to achieve a high reproductive performance in swine arise from a failure of sows to express estrus, failure to accurately determine onset of estrus for artificial inseminations and failure to determine the ovulation time after standing oestrus. The best predictor for time of ovulation is frequent detection of oestrus, because time of ovulation occurs approximately 38 to 48 h after onset of estrus (Anderson et al., 1993; Weitze et al., 1994; Soede et al., 1995). The effects of the timing of insemination relative to ovulation on fertilization rate has been study by Soede et al.(1995).Targeting insemination within 24 h before ovulation seems optimal to achieve a high farrowing rate and large little size (Nissen et al., 1997). Insemination between 0 and 24 h before ovulation results in high fertilization rates and consequently, a low number of re-breeders and a slightly higher litter size (Kemp

**10.5 Effect of long term versus short term extenders as freezing extender I on quality** 

In pig industry, the boar semen used for artificial insemination is extended with semen extender and kept in cold storage at 18-20C for few days before artificial insemination. It has been recently reported that using of long term extenders (i.e. Androstar®Plus,

(Szczesniak-Fabianczyk et al. 2003). In frozen dog semen, Micheal et al. (2007) reported that supplement of L-cysteine resulted in increased viability and rapid steady forward movement (RSF movement). Recently, Kaeoket et al. (2008b) also found that addition of 5 mM L-cysteine (the same concentration used for fresh boar semen preservation by Funahashi and Sano, 2005) has a tendency (not significant difference) to improve post-thawed semen quality when compare with the addition of glutathione and water-soluble vitamin E.

#### **10.3 Effect of L-Cysteine x DHA on frozen boar semen**

The characteristics of the sperm cryoinjury included the destabilization of lipid bilayer of the sperm plasma membrane, change in permeability of plasma membrane and a reduction of the viability of sperm. It is well documented that the boar sperm are highly susceptible to temperature below 15 °C mainly due to a relatively poor lipid composition and structure of plasma membrane compared to other domestic species. The boar sperm plasma membrane consisted of a high levels of polyunsaturated fatty acids (PUFAs) especially docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) and low level of cholesterol: phospholipids ratio. It has been shown that the level of PUFA content play an important role in the sperm membrane fluidity and cause sperm susceptible to lipid peroxidation (LPO). During frozen-thawed (FT) process, sperm are attacked by reactive oxygen species (ROS) owing to LPO and leading to significantly decrease in the PUFAs content of their plasma membrane. ROS were mainly produced by the defective or dead spermatozoa and result in a reduction of sperm motility, sperm viability and eventually fertilizing ability. In order to minimize the sperm cryoinjury, the supplement of antioxidant compounds and some fatty acid to the semen extender have been reported to minimize ROS formation and enhance the plasma membrane function in many species. *L*-cysteine, a precursor of intracellular glutathione, plays an important role in the protecting sperm from oxidative stress and act as capacitation inhibitor. Earlier studies have demonstrated that *L*-cysteine supplement in the semen extender improve the motility of FT bull semen, prolonged sperm survival time and reduced chromatin damage in FT boar sperm. In addition, the use of n-3 enriched hen egg yolk in the semen extender increased the proportion of DHA content in the boar sperm. Our previous study found that the addition of *L*-cysteine directly into lactose egg yolk (DHA-enriched) base extender significantly improves the sperm motility and intact acrosome of FT boar sperm. In addition, Kaeoket et al. (2010) found that the supplement of DHA (fish oil) improves the sperm motility, viability and acrosome integrity of the FT boar sperm.

#### **10.4 Effect of seminal plasma on post-thawing semen quality and reproductive performance after artificial insemination**

Seminal plasma is the liquid constituent of an ejaculate, comprising a combination of fluids secreted by the male accessory glands (i.e., mainly from the seminal vesicle in boars) during an ejaculation. There is evidence that seminal plasma is able to arrest or reverse cryoinjury and perhaps extend the longevity of the sperm by inhibiting or reversing capacitation and acrosome reactions, and also by its antioxidant activity (Brzezińska-Ślebodzińska et al. 1995; Strzezek et al. 1999; Suzuki et al. 2002; Vadnais and Roberts 2007; Bailey et al. 2008). During the cryopreservation process of boar semen, seminal plasma is normally not required, and discarded by the centrifugation at the beginning of semen preparation, which may result in a lack of a significant contribution (i.e., antioxidant property, inhibiting or reversing

(Szczesniak-Fabianczyk et al. 2003). In frozen dog semen, Micheal et al. (2007) reported that supplement of L-cysteine resulted in increased viability and rapid steady forward movement (RSF movement). Recently, Kaeoket et al. (2008b) also found that addition of 5 mM L-cysteine (the same concentration used for fresh boar semen preservation by Funahashi and Sano, 2005) has a tendency (not significant difference) to improve post-thawed semen quality when

The characteristics of the sperm cryoinjury included the destabilization of lipid bilayer of the sperm plasma membrane, change in permeability of plasma membrane and a reduction of the viability of sperm. It is well documented that the boar sperm are highly susceptible to temperature below 15 °C mainly due to a relatively poor lipid composition and structure of plasma membrane compared to other domestic species. The boar sperm plasma membrane consisted of a high levels of polyunsaturated fatty acids (PUFAs) especially docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) and low level of cholesterol: phospholipids ratio. It has been shown that the level of PUFA content play an important role in the sperm membrane fluidity and cause sperm susceptible to lipid peroxidation (LPO). During frozen-thawed (FT) process, sperm are attacked by reactive oxygen species (ROS) owing to LPO and leading to significantly decrease in the PUFAs content of their plasma membrane. ROS were mainly produced by the defective or dead spermatozoa and result in a reduction of sperm motility, sperm viability and eventually fertilizing ability. In order to minimize the sperm cryoinjury, the supplement of antioxidant compounds and some fatty acid to the semen extender have been reported to minimize ROS formation and enhance the plasma membrane function in many species. *L*-cysteine, a precursor of intracellular glutathione, plays an important role in the protecting sperm from oxidative stress and act as capacitation inhibitor. Earlier studies have demonstrated that *L*-cysteine supplement in the semen extender improve the motility of FT bull semen, prolonged sperm survival time and reduced chromatin damage in FT boar sperm. In addition, the use of n-3 enriched hen egg yolk in the semen extender increased the proportion of DHA content in the boar sperm. Our previous study found that the addition of *L*-cysteine directly into lactose egg yolk (DHA-enriched) base extender significantly improves the sperm motility and intact acrosome of FT boar sperm. In addition, Kaeoket et al. (2010) found that the supplement of DHA (fish oil) improves the sperm motility, viability and acrosome integrity

**10.4 Effect of seminal plasma on post-thawing semen quality and reproductive** 

Seminal plasma is the liquid constituent of an ejaculate, comprising a combination of fluids secreted by the male accessory glands (i.e., mainly from the seminal vesicle in boars) during an ejaculation. There is evidence that seminal plasma is able to arrest or reverse cryoinjury and perhaps extend the longevity of the sperm by inhibiting or reversing capacitation and acrosome reactions, and also by its antioxidant activity (Brzezińska-Ślebodzińska et al. 1995; Strzezek et al. 1999; Suzuki et al. 2002; Vadnais and Roberts 2007; Bailey et al. 2008). During the cryopreservation process of boar semen, seminal plasma is normally not required, and discarded by the centrifugation at the beginning of semen preparation, which may result in a lack of a significant contribution (i.e., antioxidant property, inhibiting or reversing

compare with the addition of glutathione and water-soluble vitamin E.

**10.3 Effect of L-Cysteine x DHA on frozen boar semen** 

of the FT boar sperm.

**performance after artificial insemination** 

capacitation, inhibiting and acrosome reaction and binding ability of its protein to the sperm plasma membrane) of seminal plasma in protecting sperm from cryoinjury. It has been demonstrated in rams that seminal plasma protein is able to revert the cold shock damage on the sperm membrane (Barrios et al. 2000). This effect has also been reported in boars, in that holding boar spermatozoa in its seminal plasma before cooling and freezing defends against the cold shock (Pursel et al. 1973). In addition, it has been shown that adding seminal plasma to the post-thawing solution increased the percentage of stallion sperm motility (Alghamdi et al. 2005). In dogs, it has been shown that frozen semen diluted with their prostatic fluid yielded a higher post-thawing motility (Rota et al. 2007). As a result, it seems likely that seminal plasma or prostatic fluid constituents have a positive effect on post-thawing sperm motility. In addition, semen with a high percentage of progressive motility illustrates their plasma membrane integrity and superb metabolism (Johnson et al., 2000) and also reflects their ability for fertilization (Vyt et al. 2004; Estienne et al. 2007). It can be hypothesized that the presence of supernatant (semen plasma plus semen extender) during the thawing process may improve frozen-thawed boar sperm motility.

Seminal plasma is the liquid constituent of an ejaculate, comprising a combination of fluids secreted by the male accessory glands (i.e. seminal vesicle) during an ejaculation. It is evidence that seminal plasma able to arrest or reverse cryoinjury and perhaps extend the longevity of the sperm by inhibiting or reversing capacitation and acrosome reaction (Suzuki et al., 2002). Besides, seminal plasma appears to play an important role in the female reproductive tract after insemination, e.g. attenuate the post insemination inflammatory response in the uterus of the sow which may influence the chances of conception (Rozeboom et al.,1999) and its component such as hormone estrogen may also resulted in a release of prostaglandins from the pigs endometrium to the utero-ovarian veins and lymphatic vessels which in turn decrease duration time from standing oestrus to ovulation in gilts (Clause et al.,1987,1990; Weitze et al.,1990), Therefore, it seems likely that seminal plasma constituents (both the oestrogen and the protein fraction) have an effect on ovulation time in sows (Waberski et al.,1995; Kaeoket and Tummaruk, 2002b).

Generally, limitations to achieve a high reproductive performance in swine arise from a failure of sows to express estrus, failure to accurately determine onset of estrus for artificial inseminations and failure to determine the ovulation time after standing oestrus. The best predictor for time of ovulation is frequent detection of oestrus, because time of ovulation occurs approximately 38 to 48 h after onset of estrus (Anderson et al., 1993; Weitze et al., 1994; Soede et al., 1995). The effects of the timing of insemination relative to ovulation on fertilization rate has been study by Soede et al.(1995).Targeting insemination within 24 h before ovulation seems optimal to achieve a high farrowing rate and large little size (Nissen et al., 1997). Insemination between 0 and 24 h before ovulation results in high fertilization rates and consequently, a low number of re-breeders and a slightly higher litter size (Kemp et al., 1996; Kaeoket et al., 2002a, 2005).

#### **10.5 Effect of long term versus short term extenders as freezing extender I on quality of frozen boar semen**

In pig industry, the boar semen used for artificial insemination is extended with semen extender and kept in cold storage at 18-20C for few days before artificial insemination. It has been recently reported that using of long term extenders (i.e. Androstar®Plus,

Cryopreservation of Boar Spermatozoa: An Important Role of Antioxidants 157

antioxidant activity, inhibition of lipoperoxidation by its scavenging activity, reduction in LDL cholesterol and induction of HDL cholesterol, inhibition of platelet aggregation [20], its potential implications as a UV-A filter in sunscreen cosmetics, treatment of type 2 diabetes mellitus and allergic reactions. These data suggest that gamma-oryzanol, especially, with its antioxidant and scavenging activities can be useful as an antioxidant and lipid peroxidation inhibitor (i.e., membrane stabilizing) during cryopreservation. Rice bran oil is widely used in salad dressing and cooking oil in Asian countries including China, India, Japan and Thailand. At present, it is becoming to gain acceptance in Western countries as well. However, no scientific information is available on its antioxidant and scavenging activities

in minimizing the detrimental effects of ROS during the cryopreservation of semen.

In pigs, it is well-documented that the optimal insemination time for fresh semen to maximize the good fertilisation rate is within 24 h before ovulation. It has been shown that the fertile life span of the pig oocyte is limited to between 8-12 h after ovulation. At suboptimal times for artificial insemination (AI) leads to inferior FR and litter sizes results. In addition, Kaeoket et al. (2002; 2005) demonstrated that when sows were inseminated after ovulation, fertilised oocytes and developed embryos were observed up to Day 11 but no embryos were found at Day 19. Subsequently, these sows returned to oestrus with a

It has been demonstrated that the duration of oestrus is related to the WOI, i.e., sows with a short WOI (3-4 days) on average have a long oestrus duration, which is associated with a longer time from onset of oestrus to ovulation. On the contrary, sows with a WOI of 5-6 days or longer, have a shorter time from onset of oestrus to ovulation, and therefore should be inseminated (with fresh semen) sooner after the onset of oestrus to ensure that the first insemination occurs before ovulation. This recommendation is in accordance with the observation that the average timing of ovulation varies between 64 and to 72% of the

For frozen boar semen, it is predictable that insemination with frozen-thawed semen will result in lower PR, low FR and litter sizes. During the last decade, most of the experiments with fertility tests (field trial) of frozen-thawed boar semen have been carried out by using deep intrauterine insemination (DIUI, dose ranged from 150 million to 1 billion spermatozoa). Nevertheless, for fresh semen, an intrauterine insemination (doses ranging from 1-3 billion) has been performed with a high fertility results (i.e., high PR, FR and litter sizes). Recently, it has been shown that a satisfactory fertility outcome was accomplished by performing IUI (doses raning from 1.5-3 billion) together with fixed-time insemination (using a correlation of WOI-Oestrus duration-Ovulation time). This strategy may improve fertility of frozen boar semen when one performs insemination in a commercial pig farms (a

Based on above review, the conclusion can be drawn as follows: (I) some antioxidants, such as, Oryzanol, L-cysteine and its combination with DHA from fish oil, Vitamin E, Vitamin C, can be used in order to improve the quality of frozen boar semen" (II) the artificial

**12. Artificial insemination with frozen boar semen** 

prolonged interval.

duration of oestrus.

field trial).

**13. Conclusion** 

ModenaTM, Vitasem LD) to preserve fresh semen for 7 days yield a superior fresh boar semen qualities compare with those using of short term extender (Kaeoket et al., 2010d). In addition, the different in extended fresh semen qualities were also found depending on each type of long term extender used. This indicated that some constituents in each long term extender may assist sperm to overcome cold shock during cold storage. Generally, the difference between the short term and long term extenders are the ingredients contained in the extenders. Long term extenders contain complex buffering agent (i.e. HEPES, Tris, TES and MOPs) and antioxidants (i.e., bovine serum albumin (BSA), beta-carotene, cysteine, taurine, vitamin E and ascorbic acid) (Alvarez and Storey, 1995; Gadea, 2003; Funahashi and Sano, 2005), which can maintain semen qualities during cold storage for a longer period than short term extender.

#### **10.6 Effect of differents sugars in LEY freezing extender on frozen boar semen quality**

During cryopreservation, both physical and chemical factors including the rapid change in temperature or thermal stress, the intracellular ice formation, oxidative stress and osmotic stress led to the sperm plasma membrane damage (Meideros et al., 2000). Generally, the freezing extender consists of cryoprotectant, sugars, buffer, and some antibiotics (Johnson et al., 2000). Glycerol is the most common permeable cryoprotectant used for cryopreservation of boar semen (Holt, 2000). Egg yolk is a common non-permeable cryoprotectant. Different types of sugars, such as, trehalose, lactose, fructose, have been used in the freezing extender of boar semen (Purdy et al., 2006). Sugar is not only a source of energy but also protects the spermatozoa from dehydration and intracellular ice formation during the cryopreservation process (Watson, 2000). In general, lactose is the most common sugar used for the cryopreservation of boar semen (Johnson, 1985; Buranaamnuay et al., 2009; Chanapiwat et al., 2009; Chanapiwat et al., 2010; Kaeoket et al., 2010a; Kaeoket et al., 2010b; Kasettrut and Kaeoket, 2010). The effect of either type or concentration of sugar supplement in the freezing extender on the post-thawed semen qualities has been reported in dog (Yildiz et al., 2000; Yamashiro et al., 2007), ram (Aisen et al., 2002), bovine (Woelders, et al., 1997; Hu et al., 2010) and boar (Roca et al., 2008; Gutiérrez-Pérez et al., 2009; Malo et al., 2010; Mercado et al., 2010). For instance, the supplement of 55 mM glucose improved the motility pattern of the FT boar spermatozoa compared to 0 and 180 mM (Roca et al., 2008). Hu et al. (2009) found that the addition of 100 mM trehalose in the extender improved post-thawed boar sperm motility, viability and acrosome integrity compared to 0, 25, 50 and 200 mM. In addition, Malo et al. (2010) found that the trehalose-based freezing extender enhances the sperm survival rate and the fertilization rate by *in vitro* fertilization (IVF) compared to lactose and glucose based freezing extender. In order to improve the post-thawed boar sperm quality, it is important to investigate the influence of different sugars on FT boar sperm.

#### **11. Phytosterol on frozen boar semen quality**

During the past decade, several studies have focused on supplementation with a variety of antioxidants (e.g. vitamin E, vitamin C, L-cysteine, glutathione, taurine, pyruvate, SOD, catalase) in the freezing extenders of frozen boar semen with an attempt to minimize the detrimental effects of ROS, which occur during the freezing process.

Gamma-oryzanol, a phytosteryl ferulate mixture extracted from rice bran oil, has received a great deal of attention because of its significant various health-promoting functions such as

ModenaTM, Vitasem LD) to preserve fresh semen for 7 days yield a superior fresh boar semen qualities compare with those using of short term extender (Kaeoket et al., 2010d). In addition, the different in extended fresh semen qualities were also found depending on each type of long term extender used. This indicated that some constituents in each long term extender may assist sperm to overcome cold shock during cold storage. Generally, the difference between the short term and long term extenders are the ingredients contained in the extenders. Long term extenders contain complex buffering agent (i.e. HEPES, Tris, TES and MOPs) and antioxidants (i.e., bovine serum albumin (BSA), beta-carotene, cysteine, taurine, vitamin E and ascorbic acid) (Alvarez and Storey, 1995; Gadea, 2003; Funahashi and Sano, 2005), which can maintain semen qualities during cold storage for a longer period

**10.6 Effect of differents sugars in LEY freezing extender on frozen boar semen quality**  During cryopreservation, both physical and chemical factors including the rapid change in temperature or thermal stress, the intracellular ice formation, oxidative stress and osmotic stress led to the sperm plasma membrane damage (Meideros et al., 2000). Generally, the freezing extender consists of cryoprotectant, sugars, buffer, and some antibiotics (Johnson et al., 2000). Glycerol is the most common permeable cryoprotectant used for cryopreservation of boar semen (Holt, 2000). Egg yolk is a common non-permeable cryoprotectant. Different types of sugars, such as, trehalose, lactose, fructose, have been used in the freezing extender of boar semen (Purdy et al., 2006). Sugar is not only a source of energy but also protects the spermatozoa from dehydration and intracellular ice formation during the cryopreservation process (Watson, 2000). In general, lactose is the most common sugar used for the cryopreservation of boar semen (Johnson, 1985; Buranaamnuay et al., 2009; Chanapiwat et al., 2009; Chanapiwat et al., 2010; Kaeoket et al., 2010a; Kaeoket et al., 2010b; Kasettrut and Kaeoket, 2010). The effect of either type or concentration of sugar supplement in the freezing extender on the post-thawed semen qualities has been reported in dog (Yildiz et al., 2000; Yamashiro et al., 2007), ram (Aisen et al., 2002), bovine (Woelders, et al., 1997; Hu et al., 2010) and boar (Roca et al., 2008; Gutiérrez-Pérez et al., 2009; Malo et al., 2010; Mercado et al., 2010). For instance, the supplement of 55 mM glucose improved the motility pattern of the FT boar spermatozoa compared to 0 and 180 mM (Roca et al., 2008). Hu et al. (2009) found that the addition of 100 mM trehalose in the extender improved post-thawed boar sperm motility, viability and acrosome integrity compared to 0, 25, 50 and 200 mM. In addition, Malo et al. (2010) found that the trehalose-based freezing extender enhances the sperm survival rate and the fertilization rate by *in vitro* fertilization (IVF) compared to lactose and glucose based freezing extender. In order to improve the post-thawed boar sperm quality, it is important to

During the past decade, several studies have focused on supplementation with a variety of antioxidants (e.g. vitamin E, vitamin C, L-cysteine, glutathione, taurine, pyruvate, SOD, catalase) in the freezing extenders of frozen boar semen with an attempt to minimize the

Gamma-oryzanol, a phytosteryl ferulate mixture extracted from rice bran oil, has received a great deal of attention because of its significant various health-promoting functions such as

investigate the influence of different sugars on FT boar sperm.

detrimental effects of ROS, which occur during the freezing process.

**11. Phytosterol on frozen boar semen quality** 

than short term extender.

antioxidant activity, inhibition of lipoperoxidation by its scavenging activity, reduction in LDL cholesterol and induction of HDL cholesterol, inhibition of platelet aggregation [20], its potential implications as a UV-A filter in sunscreen cosmetics, treatment of type 2 diabetes mellitus and allergic reactions. These data suggest that gamma-oryzanol, especially, with its antioxidant and scavenging activities can be useful as an antioxidant and lipid peroxidation inhibitor (i.e., membrane stabilizing) during cryopreservation. Rice bran oil is widely used in salad dressing and cooking oil in Asian countries including China, India, Japan and Thailand. At present, it is becoming to gain acceptance in Western countries as well. However, no scientific information is available on its antioxidant and scavenging activities in minimizing the detrimental effects of ROS during the cryopreservation of semen.
