Zhang Y Z, Zhang S C, Liu X Z, Xu Y J, Hu J H, Xu Y Y, Li J, Chen S L
Department of Marine Biology, Ocean University of China, Qingdao 266003, PR China.
Theriogenology. 2005 Feb;63(3):763-73. doi: 10.1016/j.theriogenology.2004.04.011.
With the purpose of finding an ideal cryoprotectant or combination of cryoprotectants in a suitable concentration for flounder (Paralichthys olivaceus) embryo cryopreservation, we tested the toxicities, at culture temperature (16 degrees C), of five most commonly used cryoprotectants-dimethyl sulfoxide (Me2SO), glycerol, methanol (MeOH), 1,2-propylene glycol (PG) and ethylene glycol (EG). In addition, cryoprotective efficiency to flounder embryos of individual and combined cryoprotectants were tested at -15 degrees C for 60 min. Five different concentrations of each of the five cryoprotectants and 20 different combinations of these cryoprotectants were tested for their protective efficiency. The results showed that the toxicity to flounder embryos of the five cryoprotectants are in the following sequence: PG < MeOH < Me2SO < glycerol < EG (P < 0.05); whereas the protective efficiency of each cryoprotectant, at -15 degrees C for a period of 60 min, are in the following sequence: PG > Me2SO approximately MeOH approximately glycerol > EG (greater symbols mean P < 0.05, and approximate symbols mean P > 0.05). Methanol combined with any one of the other cryoprotectants gave the best protection, while ethylene glycol combined with any one of the other cryoprotectants gave the poorest protection at -15 degrees C. Toxicity effect was concentration dependent with the lowest concentration being the least toxic for all five cryoprotectants at 16 degrees C. For PG, MeOH and glycerol, 20% solutions gave the best protection at -15 degrees C; whereas a 15% solution of Me2SO, and a 10% solution of EG, gave the best protection at -15 degrees C.
为了找到一种理想的冷冻保护剂或合适浓度的冷冻保护剂组合用于牙鲆（Paralichthys olivaceus）胚胎冷冻保存，我们在培养温度（16℃）下测试了五种最常用的冷冻保护剂——二甲基亚砜（Me2SO）、甘油、甲醇（MeOH）、1,2 - 丙二醇（PG）和乙二醇（EG）的毒性。此外，在-15℃下对单个及组合冷冻保护剂对牙鲆胚胎的冷冻保护效率进行了60分钟的测试。测试了五种冷冻保护剂各自的五种不同浓度以及它们的20种不同组合的保护效率。结果表明，五种冷冻保护剂对牙鲆胚胎的毒性顺序为：PG < MeOH < Me2SO < 甘油 < EG（P < 0.05）；而在-15℃下60分钟时，每种冷冻保护剂的保护效率顺序为：PG > Me2SO ≈ MeOH ≈ 甘油 > EG（大于号表示P < 0.05，约等于号表示P > 0.05）。在-15℃时，甲醇与其他任何一种冷冻保护剂组合提供的保护效果最佳，而乙二醇与其他任何一种冷冻保护剂组合提供的保护效果最差。毒性效应呈浓度依赖性，在16℃时，所有五种冷冻保护剂浓度最低时毒性最小。对于PG、MeOH和甘油，20%的溶液在-15℃时提供最佳保护；而15%的Me2SO溶液和10%的EG溶液在-15℃时提供最佳保护。
