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It was the aim of this study to analyse the quality of frozen\u2010thawed stallion semen after complete or partial replacement of glycerol in the freezing extender by alternative cryoprotectants. We hypothesized that partial or total replacement of glycerol by cryoprotectants occurring in cold\u2010resistant frog, insect or plant species results in similar or better semen quality after freezing\u2013thawing. As basic medium, the commercial Ghent basic extender was used and either supplemented with glucose and urea, trehalose and proline, or trehalose and betaine. Based on a series of preliminary experiments, semen was frozen in either commercial Ghent cryopreservation extender (Ghent control), Ghent glucose\u2013urea extender or a Ghent combined extender (glucose\u2013urea, trehalose\u2010betaine and trehalose\u2010proline; volume ratio of 2:1:2) in a computer\u2010controlled rate freezer. After freezing\u2013thawing, semen was analysed for motility, membrane integrity, phosphatidylserine translocation, mitochondrial membrane potential and chromatin condensation. No differences between Ghent control and Ghent glucose\u2013urea extender were seen, while all endpoints except <jats:styled-content style=\"fixed-case\">DNA<\/jats:styled-content> integrity were negatively affected in Ghent combined extender (e.g., progressive motility: Ghent 49.2\u00a0\u00b1\u00a03.7, Ghent glucose\u2013urea 46.5\u00a0\u00b1\u00a04.6, Ghent combined 24.4\u00a0\u00b1\u00a02.8%; <jats:italic>p<\/jats:italic>\u00a0&lt;\u00a0.001). In conclusion, glycerol concentration in a commercial freezing extender for equine spermatozoa can be successfully reduced when urea as an additive cryoprotectant is added and the glucose concentration is elevated. However, total glycerol replacement with urea, betaine, proline and trehalose was less successful.<\/jats:p>","DOI":"10.1111\/rda.12970","type":"journal-article","created":{"date-parts":[[2017,3,22]],"date-time":"2017-03-22T20:57:38Z","timestamp":1490216258000},"page":"715-721","source":"Crossref","is-referenced-by-count":10,"title":["Influence of partial or total replacement of glycerol by alternative cryoprotectants in Ghent freezing extender on post\u2010thaw sperm quality in stallions"],"prefix":"10.1111","volume":"52","author":[{"given":"RA","family":"De Oliveira","sequence":"first","affiliation":[{"name":"Artificial Insemination and Embryo Transfer Vetmeduni Vienna  Vienna Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"S","family":"Budik","sequence":"additional","affiliation":[{"name":"Artificial Insemination and Embryo Transfer Vetmeduni Vienna  Vienna Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6077-7362","authenticated-orcid":false,"given":"C","family":"Aurich","sequence":"additional","affiliation":[{"name":"Artificial Insemination and Embryo Transfer Vetmeduni Vienna  Vienna Austria"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2017,3,21]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1071\/RD14449"},{"key":"e_1_2_9_3_1","series-title":"Proceedings, Workshop on Transporting Gametes and Embryos","first-page":"74","author":"Alvarenga M. 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