Animal Reproduction (AR)
https://animal-reproduction.org/article/doi/10.1590/1984-3143-AR2023-0143
Animal Reproduction (AR)
ORIGINAL ARTICLE

L-Carnitine enhances porcine sperm quality, longevity, and zona pellucida binding in cooled semen

Monique de Albuquerque Lagares; Nathalia Abreu Amaral; Joyce Junia Braga; Natalia de Castro Alves; Marina Morra Freitas; Rafael Romero Nicolino; Raphael Rocha Wenceslau; Fernanda da Rocha Anselmo; Marina Maria do Carmo Silva Oliveira; Eduardo Damasceno Costa; Fernanda Radicchi Campos Lobato de Almeida; Rubens Stahlberg

http://dx.doi.org/10.1590/1984-3143-AR2023-0143 Anim Reprod, vol.22, n1, e20230143, 2025
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Abstract

Porcine breeding industries typically ensure the viability of boar artificial insemination doses during a 5-day liquid storage period at 17 °C. This study aimed to investigate whether the addition of L-carnitine (LC) to boar semen doses on different days of cooled storage could extend their usability. In experiment 1, LC was added to porcine semen doses on the fifth day (d5) of cooled storage performing five treatments control (no LC), 0.5, 1-, 5- and 10-mM LC. On d6 and d8 of storage, semen samples were evaluated for sperm motility and kinematic parameters, membrane functionality, and hydrogen peroxide and nitrite concentrations. In experiment 2, the number of sperm bound to the zona pellucida (ZP) was determined, as a way to investigate sperm penetration capability from boar insemination doses, with co-incubation with porcine oocytes. LC concentration that produced the most favorable outcomes in Experiment 1 was chosen to experiments 2 and 3, performing two treatments in the absence and with the LC. In Experiment 3, LC was added to cooled porcine semen doses after one day of storage (d1), and the same evaluations of experiment 1 were conducted on days 5, 7, 9, and 12, including sperm membrane integrity. The addition of 10 mM LC on d5 and d1 of storage improved sperm motility, which was extended up to 8 and 12 days of cooled storage, respectively. LC addition on d5 of storage increased sperm membrane functionality, while when added to semen on d1 of storage, it decreased NO2- concentration on d9. On d6 of cooled storage 10 mM LC increased the number of sperm bound to ZP compared to the control. In conclusion, adding 10 mM LC to porcine semen doses at 17 °C improved sperm characteristics and ZP binding, ultimately enhancing sperm viability for up to 12d.

Keywords

spermatozoa, swine, boar, antioxidant, artificial insemination

References

Abd-Elrazek AM, Ahmed-Farid OAH. Protective effect of L-carnitine and L-arginine against busulfan-induced oligospermia in adult rat. Andrologia. 2018;50(1):1-8. PMid:28444774.

Agarwal A, Saleh RA. Role of oxidants in male infertility: rationale, significance, and treatment. Urol Clin North Am. 2002;29(4):817. http://doi.org/10.1016/S0094-0143(02)00081-2. PMid:12516754.

Ahmadi S, Bashiri R, Ghadiri-Anari A, Nadjarzadeh A. Antioxidant supplements and semen parameters: an evidence-based review. Int J Reprod Biomed. 2016;14(12):729-36. http://doi.org/10.29252/ijrm.14.12.729. PMid:28066832.

Awda BJ, Mackenzie-Bell M, Buhr MM. Reactive oxygen species and boar sperm function. Biol Reprod. 2009;81(3):553-61. http://doi.org/10.1095/biolreprod.109.076471. PMid:19357363.

Balercia G, Regoli F, Armeni T, Koverech A, Mantero F, Boscaro M. Placebo-controlled double-blind randomized trial on the use of L-carnitine, L-acetylcarnitine or combined L-carnitine and L-acetylcarnitine in men with idiopathic asthenozoospermia. Fertil Steril. 2005;84(3):662-71. http://doi.org/10.1016/j.fertnstert.2005.03.064. PMid:16169400.

Balogun KB, Lu N, Orlando U, Torborg H, Kleve-Feld M, Denton A, Holstine A, Stewart KR. Effects of dietary carnitine supplementation on semen output and quality of boars. Transl Anim Sci. 2022;6(4):c143. http://doi.org/10.1093/tas/txac143. PMid:36381951.

Bandeira TB, Toniolli LS, Guimarães DB, Freitas EN, Nunes TGP, Toniolli R. Curvas de resfriamento do sêmen do varrão diluído em acp 103 adicionado de gema de ovo em concentração fixa. Ciênc Anim. 2016;17:25-30.

Bansal AK, Bilaspuri GS. Impacts of oxidative stress and antioxidants on semen functions. Vet Med Int. 2010;2010:686137. http://doi.org/10.4061/2011/686137. PMid:20871827.

Berger T, Anderson DL, Penedo MCT. Porcine sperm fertilizing potential in relationship to sperm functional capacities. Anim Reprod Sci. 1996;44(4):231-9. http://doi.org/10.1016/0378-4320(96)01565-5.

Betancur GR, Zapata KV, Vidal PAC, Sánchez Y, Rojano BA. Cooling of porcine semen in an extender supplemented with isoespintanol. Cienc Rural. 2023;53(11):e20220508. http://doi.org/10.1590/0103-8478cr20220508.

Braundmeier AG, Demers JM, Shanks RD, Miller DJ. The relationship of porcine sperm zona-binding ability to fertility. J Anim Sci. 2004;82(2):452-8. http://doi.org/10.2527/2004.822452x. PMid:14974543.

Bremer J. Carnitine-metabolism and functions. Physiol Rev. 1983;63(4):1420-80. http://doi.org/10.1152/physrev.1983.63.4.1420. PMid:6361812.

Cabral REL, Mendes TB, Vendramini V, Miraglia SM. Carnitine partially improves oxidative stress, acrosome integrity, and reproductive competence in doxorubicin-treated rats. Andrology. 2018;6(1):236-46. http://doi.org/10.1111/andr.12426. PMid:28973833.

Cerolini S, Maldjian A, Surai P, Noble R. Viability, susceptibility to peroxidation and fatty acid composition of boar semen during liquid storage. Anim Reprod Sci. 2000;58(1-2):99-111. http://doi.org/10.1016/S0378-4320(99)00035-4. PMid:10700648.

Chang D, Kong F, Jiang W, Li F, Zhang C, Ding H, Kang Y, Li W, Huang C, Zhou X, Zhang X, Jiao H, Kang Y, Shang X, Zhang B. Effects of l-carnitine administration on sperm and sex hormone levels in a male Wistar rat reproductive system injury model in a high-altitude hypobaric hypoxic environment. Reprod Sci. 2023;30(7):2231-47. http://doi.org/10.1007/s43032-022-00948-5. PMid:36633830.

Elokil AA, Bhuiyan AA, Liu H-Z, Hussein MN, Ahmed HI, Azmal SA, Yang L, Li S. The capability of L-Carnitine mediated antioxidant on cock during aging: evidence for the improved semen quality and enhanced testicular expressions of GnRH1, GnRHR, and melatonin receptors MT 1/2. Poult Sci. 2019;98(9):4172-81. http://doi.org/10.3382/ps/pez201. PMid:31001634.

Flesch FM, Gadella BM. Dynamics of the mammalian sperm plasma membrane in the process of fertilization. Biochim Biophys Acta. 2000;1469(3):197-235. http://doi.org/10.1016/S0304-4157(00)00018-6. PMid:11063883.

Flowers WL. Factors affecting the production of quality ejaculates from boars. Anim Reprod Sci. 2022;246:106840. http://doi.org/10.1016/j.anireprosci.2021.106840. PMid:34518030.

Gadea J. Sperm factors related to in vitro and in vivo porcine fertility. Theriogenology. 2005;63(2):431-44. http://doi.org/10.1016/j.theriogenology.2004.09.023. PMid:15626409.

Gibb Z, Aitken RJ. The impact of sperm metabolism during in vitro storage: the stallion as a model. BioMed Res Int. 2016;2016:9380609. http://doi.org/10.1155/2016/9380609. PMid:26881234.

Gibb Z, Lambourne SR, Quadrelli J, Smith ND, Aitken RJ. L-Carnitine and Pyruvate are prosurvival factors during the storage of stallion spermatozoa at room temperature. Biol Reprod. 2015;93(4):104. http://doi.org/10.1095/biolreprod.115.131326. PMid:26316064.

Gomez E, Irvine DS, Aitken RJ. Evaluation of a spectrophotometric assay for the measurement of malondialdehyde and 4-hydroxyalkenals in human spermatozoa: relationships with semen quality and sperm function. Int J Androl. 1998;21(2):81-94. http://doi.org/10.1046/j.1365-2605.1998.00106.x. PMid:9675617.

Green LC, Wagner DA, Glogowski J, Skipper PL, Wishnok JS, Tannenbaum SR. Analysis of nitrate, nitrite, and [15N] nitrate in biological fluids. Anal Biochem. 1982;126(1):131-8. http://doi.org/10.1016/0003-2697(82)90118-X. PMid:7181105.

Gülçin I. Antioxidant and antiradical activities of l-carnitine. Life Sci. 2006;78(8):803-11. http://doi.org/10.1016/j.lfs.2005.05.103. PMid:16253281.

Harrison RAP. Sperm plasma membrane characteristics and boar semen fertility. J Reprod Fertil Suppl. 1997;52:195-211. PMid:9602729.

Henning H, Nguyen QT, Wallner U, Waberski D. Temperature limits for storage of extended boar semen from the perspective of the sperm’s energy status. Front Vet Sci. 2022;9:953021. http://doi.org/10.3389/fvets.2022.953021. PMid:35990258.

Hensel B, Jakop U, Scheinpflug K, Mühldorfer K, Schröter F, Schäfer J, Greber K, Jung M, Schulze M. Low temperature preservation of porcine semen: influence of short antimicrobial lipopeptides on sperm quality and bacterial load. Sci Rep. 2020;10(1):13225. http://doi.org/10.1038/s41598-020-70180-1. PMid:32764567.

Hosen MB, Islam MR, Begum F, Kabir Y, Howlader MZH. Oxidative stress induced sperm DNA damage, a possible reason for male infertility. Iran J Reprod Med. 2015;13(9):525-32. PMid:26568756.

Jeulin C, Lewin LM. Role of free L-carnitine and acetyl-L-carnitine in post-gonadal maturation of mammalian spermatozoa. Hum Reprod Update. 1996;2(2):87-102. http://doi.org/10.1093/humupd/2.2.87. PMid:9079406.

Khaw SC, Wong ZZ, Anderson A, Silva SM. L-carnitine and l-acetylcarnitine supplementation for idiopathic male infertility. Reprod Fertil. 2020;1(1):67-81. http://doi.org/10.1530/RAF-20-0037. PMid:35128424.

Lagares MA, Silva GC, Cortes SF, Moreira FHM, Neves FCD, Alves NDC, Viegas RN, Diniz TF, Lemos VS, Rezende ASC, Freitas MM, Stahlberg R, Nicolino RR. L-carnitine added to post-thawed semen acts as an antioxidant and a stimulator of equine sperm metabolism. Andrologia. 2022;54(3):e14338. http://doi.org/10.1111/and.14338. PMid:34878673.

Lagares MA, Petzoldt R, Sieme H, Klug E. Assessing equine sperm-membrane integrity. Andrologia. 2000;32(3):163-7. http://doi.org/10.1046/j.1439-0272.2000.00351.x. PMid:10863971.

Lagares MA, Petzodt R, Sieme H, Klug E. Preservação do sêmen fresco eqüino: avaliação da integridade da membrana espermática sob condições hiposmóticas. Arq Fac Vet UFRGS. 1998;26:29-42.

Lahimer M, Gherissi O, Salem NB, Mustapha HB, Bach V, Khorsi-Cauet H, Ki H, Ali HB, Khalifa MB, Ajina M. Effect of micronutrients and l-carnitine as antioxidant on sperm parameters, genome integrity, and ICSI outcomes: randomized, double-blind, and placebo-controlled clinical trial. Antioxidants. 2023;12(11):1937-54. http://doi.org/10.3390/antiox12111937. PMid:38001791.

Larsson B, Rodriguez-Martinez H. Can we use in vitro fertilization tests to predict semen fertility? Anim Reprod Sci. 2000;60–61:327-36. http://doi.org/10.1016/S0378-4320(00)00089-0. PMid:10844204.

Li R, Jia Z, Trush MA. Defining ROS in biology and medicine. React Oxyg Species. 2016;1(1):9-21. http://doi.org/10.20455/ros.2016.803. PMid:29707643.

Lima ES, Abdalla DSP. Peroxidação lipídica: mecanismos e avaliação em amostras biológicas. Rev Bras Ciênc Farm. 2001;37:293-303.

Lisboa FL, Hartwig FP, Freitas-Dell’Aqua CP, Hartwig FP, Papa FO, Dell’aqua JA Jr. Improvement of cooled equine semen by addition of Carnitines. J Equine Vet Sci. 2014;34(1):48. http://doi.org/10.1016/j.jevs.2013.10.028.

Lopez Rodriguez A, Van Soom A, Arsenakis L, Maes D. Boar management and semen handling factors affect the quality of boar extended semen. Porcine Health Manag. 2017;3(1):15. http://doi.org/10.1186/s40813-017-0062-5. PMid:28770098.

Ma L, Sun Y. Comparison of L-Carnitine vs. Coq10 and Vitamin E for idiopathic male infertility: a randomized controlled trial. Eur Rev Med Pharmacol Sci. 2022;26(13):4698-704. PMid:35856361.

Manee-In S, Parmornsupornvichit S, Kraiprayoon S, Tharasanit T, Chanapiwat P, Kaeoket K. L-carnitine Supplemented Extender Improves Cryopreserved-thawed Cat Epididymal Sperm Motility. Asian-Australas J Anim Sci. 2014;27(6):791-6. http://doi.org/10.5713/ajas.2013.13565. PMid:25050016.

Martins HS, Martins OA Fo, Araujo MSS, Martins NR, Lagares MA. Evaluation of fertilizing ability of frozen equine sperm using a bovine zona pellucida binding assay. Cryo Letters. 2018;39(5):298-305. PMid:30963161.

Mateus FG, Moreira S, Martins AD, Oliveira PF, Alves MG, Pereira ML. L-carnitine and male fertility: is supplementation beneficial? J Clin Med. 2023;12(18):5796-817. http://doi.org/10.3390/jcm12185796. PMid:37762736.

Menegat MB, Mellagi APG, Bortolin RC, Menezes TA, Vargas AR, Bernardi ML, Wentz I, Gelain DP, Moreira JC, Bortolozzo FP. Sperm quality and oxidative status as affected by homogenization of liquid-stored boar semen diluted in short-and long-term extenders. Anim Reprod Sci. 2017;179:67-79. http://doi.org/10.1016/j.anireprosci.2017.02.003. PMid:28215454.

Nazari L, Salehpour S, Hosseini S, Allameh F, Jahanmardi F, Azizi E, Ghodssi-Ghassemabadi R, Hashemi T. Effect of antioxidant supplementation containing L-carnitine on semen parameters: a prospective interventional study. JBRA Assist Reprod. 2021;25(1):76-80. PMid:32598834.

Nogueira BG, Bitencourt JL, Sampaio BFB, Bender ESC, Costa SEV, Zúccari CESN. Peroxidação lipídica e agentes antioxidantes no sêmen de mamíferos. Rev Electrón Vet. 2014;15:1-15.

Nordberg J, Arnér ESJ. Reactive oxygen species. antioxidants. and the mammalian thioredoxin system. Free Radic Biol Med. 2001;31(11):1287-312. http://doi.org/10.1016/S0891-5849(01)00724-9. PMid:11728801.

Nouroozzadeh J, Tajaddinisarmadi J, Wolff SP. Measurement of plasma hydroperoxide concentrations by the ferrous oxidation-xylenol orange assay in conjunction with triphenilphosphine. Anal Biochem. 1994;220(2):403-9. http://doi.org/10.1006/abio.1994.1357. PMid:7978285.

Partyka A, Rodak O, Bajzert J, Kochan J, Nizanski W. The effect of L-carnitine. hypotaurine and taurine supplementation on the quality of cryopreserved chicken semen. BioMed Res Int. 2017;2017:7279341. http://doi.org/10.1155/2017/7279341. PMid:28523277.

Restrepo G, Zapata K, Colorado P, Rojano B. Cooling of porcine semen in an extender supplemented with carvacrol. Reprod Domest Anim. 2023;58(6):860-6. http://doi.org/10.1111/rda.14359. PMid:37029731.

Sanocka D, Kurpisz M. Reactive oxygen species and sperm cells. Reprod Biol Endocrinol. 2004;2(1):12. http://doi.org/10.1186/1477-7827-2-12. PMid:15038829.

Sariozkan S, Ozdamar S, Turk G, Canturk F, Yay A. In vitro effects of L-carnitine and glutamine on motility, acrosomal abnormality, and plasma membrane integrity of rabbit sperm during liquid-storage. Cryobiology. 2014;68(3):349-53. http://doi.org/10.1016/j.cryobiol.2014.04.006. PMid:24759298.

Severo CK, Pereira GR, Pereira AM, Ilha GF, Oliveira JFC, Soares M, Arruda RP, Gonçalves PBD. Cysteine addition on short-term cooled boar semen preservation and its relationship with swine field fertility. Pesq Vet Bras. 2011;31(Suppl 1):25-32. http://doi.org/10.1590/S0100-736X2011001300005.

Staicu F-D, Parra CM. Nitric oxide: key features in spermatozoa. In: Saravi SSS, editor. Nitric oxide synthase: simple enzyme-complex roles. London: IntechOpen; 2017. p. 137-54.

Storey BT. Biochemical of the induction and prevention of lipoperoxidative damage in human spermatozoa. Mol Hum Reprod. 1997;3(3):203-13. http://doi.org/10.1093/molehr/3.3.203. PMid:9237246.

Stradaioli G, Sylla L, Zelli R, Chiodi P, Monaci M. Effect of L-carnitine administration on the seminal characteristics of oligoasthenospermic stallions. Theriogenology. 2004;62(3-4):761-77. http://doi.org/10.1016/j.theriogenology.2003.11.018. PMid:15226028.

Surai PF. Carnitine Enigma: from antioxidant action to vitagene regulation Part 2. transcription factors and practical applications. J. Veter. Sci. Med. 2015;3(2):1-17. http://doi.org/10.13188/2325-4645.1000018.

Szymanowicz J, Schwarz T, Murawski M, MaBopolska M, Oszczda Z, Tuz R, Nowicki J, Bartlewski PM. Storage of boar semen at 16-18 °C in the long-term commercial extender prepared with deionized water or nanowater. Anim Reprod. 2019;16(4):864-70. http://doi.org/10.21451/1984-3143-AR2019-0023. PMid:32368265.

Szymański M, Wandtke T, Wasilow K, Andryszczyk M, Janicki R, Domaracki P. Comparison of 3- and 6-month outcomes of combined oral L-carnitine fumarate and acetyl-L-carnitine therapy, included in an antioxidant formulation, in patients with idiopathic infertility. Am J Mens Health. 2021;15(5):1-11. http://doi.org/10.1177/15579883211036790. PMid:34515581.

Tabatabaei S, Aghaei A. Effect of L-carnitine on sperm quality during liquid storage of chicken semen. Comp Clin Pathol. 2012;21(5):711-7. http://doi.org/10.1007/s00580-010-1163-6.

Waberski D, Riesenbeck A, Schulze M, Weitze KF, Johnson L. Application of preserved boar semen for artificial insemination: past, present and future challenges. Theriogenology. 2019;137:2-7. http://doi.org/10.1016/j.theriogenology.2019.05.030. PMid:31186127.

Yang K, Wang N, Guo H-T, Wang J-R, Sun H-H, Sun L-Z, Yue SL, Zhou JB. Effect of L-carnitine on sperm quality during liquid storage of boar semen. Asian-Australas J Anim Sci. 2020;33(11):1763-9. http://doi.org/10.5713/ajas.19.0455. PMid:32054191.
 

Submitted date:
11/19/2023

Accepted date:
11/14/2024

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