Influence of BHT inclusion on post-thaw attributes of human semen

Abstract

The aim of this study was to determine the effect of butylated hydroxytoluene (BHT) supplemented cryopreservation medium on sperm parameters during the freeze-thaw process. In addition, sperm lipid peroxidation, DNA damage, and the amount of reactive oxygen species (ROS) were determined. Semen samples were obtained from 75 donors. Fifteen semen samples were used for optimizing BHT concentration and incubation time and 60 samples were used for the final experiments. After the determination of basic parameters, groups of three sample with similar parameters were pooled and processed by Pure Sperm gradient centrifugation. The semen samples were then diluted with normal freezing medium (control) or a medium containing 0.5 mM BHT (test) for 5 minute and stored in liquid nitrogen. Frozen cryovials were thawed individually for 20 seconds in a water bath (37°C) for evaluation. Freezing extenders supplemented with 0.5 mM BHT led to higher sperm motility and viability compared with control samples (p < 0.001). Furthermore, the addition of BHT decreased malondialdehyde (MDA) formation, DNA fragmentation, and ROS content compared with controls (p < 0.001). Our results showed that the addition of BHT to the freezing medium could be of advantage in reducing ROS and preventing the detrimental effect of ROS on the human sperm function.

The efficacy of antioxidants in sperm parameters and production of reactive oxygen species levels during the freeze‐thaw process: A systematic review and meta‐analysis

“…During full‐text screening, no common antioxidants were identified in nineteen articles (Asadmobini, Bakhtiari, Khaleghi, Esmaeili, & Mostafaei, ; Brugnon et al, ; Fontoura et al, ; Isachenko et al, ; Kotdawala et al, ; Lu et al, ; Meamar et al, ; Naeini, Bafrani, & Nikzad, ; Najafi et al, ; Oehninger, Duru, Srisombut, & Morshedi, ; Parameswari et al, ; Pariz & Hallak, ; Saeednia et al, ; Sbracia, Grasso, Sayme, Stronk, & Huszar, ; Sobhani, Eftekhaari, Shahrzad, Natami, & Fallahi, ; Taylor, Roberts, Sanders, & Burton, ; Vickram et al, ; Werner et al, ), twelve articles lacked common sperm parameters (Banihani, Agarwal, Sharma, & Bayachou, ; Branco, Garcez, Pasqualotto, Erdtman, & Salvador, ; Donnelly, McClure, & Lewis, ; Esteves, Spaine, & Cedenho, ; Gadea et al, ; Garcez et al, ; Garcez, Branco, Lara, Pasqualotto, & Salvador, ; Jenkins, Aston, & Carrell, ; Li, Lin, Liu, Xiao, & Liu, ; Minaei et al, ; Nabi et al, ; Shabani Nashtaei et al, ) and one article lacked sufficient data (Karimfar et al, ). Finally, 23 articles (Aliabadi, Jahanshahi, Talaei‐Khozani, & Banaei, ; Askari, Check, Peymer, & Bollendorf, ; Azadi, Tavalaee, Deemeh, Arbabian, & Nasr‐Esfahani, ; Bateni et al, ; Deng et al, ; Duru, Morshedi, Schuffner, & Oehninger, ; Esteves, Sharma, Thomas, & Agarwal, ; Ghorbani et al, ; Kalthur et al, ; Keshtgar, Iravanpour, Gharesi‐Fard, & Kazerooni, ; Martinez‐Soto, Dioshourcade, Gutiérrez‐Adán, Landeras, & Gadea, ; Merino et al, ; Moubasher, Din, Ali, El‐sherif, & Gaber, ; Najafi et al, ; Nekoonam, Nashtaei, Naji, Zangi, & Amidi, ; Rossi, Mazzilli, Delfino, & Dondero, ; Taylor et al, ; Thomson et al, ; Varghese et al, ; Wang, Sikka, Veeraragavan, Bell, & Hellstr...…”

“…BHT is another important scavenger of free radicals in freezing media responsible for reduction in SOD, glutathione peroxidase, and ROS production after cryopreservation and consequently protects sperm cells from DNA damage. Ghorbani et al indicated that damage to human sperm reduced in the BHT group compared to controls (25% and 49%, respectively) (Ghorbani et al, ).…”

ADIÇÃO DE HIDROXITOLUENO BUTILADO (BHT) NO DILUIDOR ACP-106c PARA CONGELAÇÃO DE SÊMEN CANINO

“…the samples were thawed and evaluated by computer analysis, as well as for membrane functionality and sperm morphology. Progressive motility in the 2.0 mM BHT group was significantly higher (P <0.05) than that of the 0 mM BHT group (27.6 ± 11.7% vs. 19.0 ± 9.5%, respectively). Regarding all other parameters evaluated, there was no difference between the groups tested.…”

“…Em relação aos valores encontrados após a descongelação, foi possível perceber uma melhor qualidade da motilidade progressiva dos espermatozoides na concentração de 2,0 mM de BHT (27,6%), quando comparado ao grupo sem o antioxidante (19,0%). Esse resultado difere do encontrado por Ghorbani et al (19) em humanos, pois relataram que concentrações de BHT superiores a 0,5 mM resultaram em menor motilidade e viabilidade espermática provavelmente devido a um aumento de ânions superóxido e H2O2 nos espermatozoides. Khumran et al (26) relataram uma toxicidade causada pelo BHT, quando este é utilizado em concentrações maiores de 1 mM em sêmen de touros, levando a redução da motilidade e da viabilidade espermática.…”

“…Existem muitas variedades de antioxidantes que podem ser utilizados, dentre eles o hidroxitolueno butilado (BHT). O efeito do BHT sobre a qualidade do sêmen já foi testado, obtendo-se excelentes resultados pós-descongelação em varrões (13) , perus (12) , búfalos (14) , touros (15) , bodes (16,17) , carpas (Cyprinus carpio) (18) e homens (19) . Entretanto, nos trabalhos já realizados em cães, é possível verificar resultados contraditórios, no qual Neagu et al (20) descrevem uma melhoria na qualidade seminal em dois diluidores à base de Tris adicionados de BHT, enquanto no trabalho realizado por Sahashi et al (21) , os resultados não mostraram uma proteção eficiente aos espermatozoides que justificasse sua inclusão ao meio diluidor.…”