ORIGINAL ARTICLE Antioxidant effect of lemon balm (Melissa officinalis) and mate tea (Ilex paraguensys) on quality, lipid peroxidation and DNA oxidation of cryopreserved boar epididymal spermatozoa V. Lu~ no 1 , L. Gil 1 , M. Olaciregui 1 , R. A. Jerez 1 , I. de Blas 1 & F. Hozbor 2 1 Department of Animal Pathology, Universidad de Zaragoza, Spain; 2 INTA Balcarce, Mar del Plata, Argentine Keywords Antioxidant—boar epididymal spermatozoa— cryopreservation—Ilex paraguensys—Melissa officinalis Correspondence Victoria Lu~ no, Universidad de Zaragoza - Department of Animal Pathology, Obstetrics and Reproduction Area, C/Miguel Servet, 177 Zaragoza 50013, Spain. Tel.: +34 976 844113; Fax: +34 976 761612; E-mail: vicluno@unizar.es Accepted: September 15, 2014 doi: 10.1111/and.12370 Summary In this study, we investigated the protective ability of the addition of two anti- oxidant herb extracts, mate tea and lemon balm, on boar epididymal frozen– thawed spermatozoa quality. Testes from mature boars were collected at local slaughterhouse, and sperm samples from epididymis were recovered by flush- ing. Spermatozoa were cryopreserved in lactose–egg yolk buffer supplemented with various concentrations of lemon balm and mate tea (0, 2.5, 5 and 10 g l 1 ) using the straw-freezing procedure. Motion parameters, acrosome and plasma membrane integrity, lipoperoxidation levels and DNA oxidative damage (8-hydroxy-2 0 -deoxyguanosine base lesion) were evaluated. There were no differences among experimental groups with regard to motility characteris- tics, viability, acrosome and plasma membrane integrity; however, the highest concentration of lemon balm produced significant (P < 0.05) improvement in curvilinear trajectory, straightness and amplitude of lateral head displacement after thawing. The supplementation of freezing extender with mate tea and lemon balm reduced sperm lipid membrane peroxidation, and only mate tea protected DNA against oxidative damage during cryopreservation at 120 min post-thawing (P < 0.05). Mate tea experimental extender at concentration of 10 g l 1 showed the lowest percentage of sperm oxidised DNA and malondial- dehyde generation; thus, mate tea is a potential candidate such as antioxidant compound on boar sperm cryopreservation medium. Introduction Boar sperm cryopreservation is a reliable alternative of long-term storage of important genetic material. The conservation of epididymal spermatozoa is increasing in wild and livestock species (Dom  ınguez-Rebolledo et al., 2010; Malo et al., 2011); thus, boars with excellent geno- type are suitable for the creation of the germplasm banks. Epididymal spermatozoa show important differences in terms of functionality, fertility and cold shock resistance respect to ejaculated spermatozoa (Rath & Niemann, 1997). Cryopreservation conditions and the subsequence spermatozoa quality must be optimised for successful of the freezing process in function of sperm source. Cooling, freezing and thawing process increase the generation of reactive oxygen species (ROS) and produce molecular lesions on sperm surface (Storey, 1997). The high content in polyunsaturated fatty acids (PUFA) of plasma membrane of boar spermatozoa makes the structure very susceptible to oxidative damage. PUFAs decrease dramatically by ROS attack because lipid peroxidation, motility, plasma membrane fluidity and integrity are affected (Storey, 1997; Cerolini et al., 2001). Many researchers determined the role of the addition to sperm freezing extenders different antioxidants to mini- mise the prejudicial effects of ROS (Zhao et al., 2009; Malo et al., 2010). In recent years, several studies have determined the antioxidant properties of different natural herbs. These herbs contain many phytochemicals (carote- noids, carnosic acid, rosmarinic acid or flavonoids) widely used in the cosmetic, pharmaceutical and food industry with beneficial effect (Yanishlieva & Marinova, © 2014 Blackwell Verlag GmbH 1 Andrologia 2014, xx, 1–8