• Clinical and Experimental Reproductive Medicine
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• v.17 no.2
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• pp.167-172
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• 1990

This study was carried out to clarify the effects of various kinds of cryoprotectants which were frequently used in freezing embryos of domestic animals on the survival of frozen-thawed mouse embryos. Mouse embryos were collected by hyperstimulation induction of ICR mouse. The samples were slowly cooled ($l^{\circ}C/min$) to temperatures between $-7^{\circ}C$ and $-30^{circ}C$ before direct transfer to liquid nitrogen ($-196^{\circ}C$) and thawed rapidly ($-500^{\circ}C$/min). As cryoprotectants, Glycerol, DMSO, Ethylene glycol and Propylene glycol were used and applied each 2 cell, 8 cell, morula in embryo stage. After normal mouse embryos developed to blastocyst by in vitro culture, we observed recovery rate and developing rate of embryos at thawing. The results obtained in these experiments were as follows : 1. The in vitro development rate from the frozen-thawed 2 cell embryos to the blastocyst were 67.7% in ethylene glycol, 65.7% in Propylene glycol, 55.2% in glycerol and 50.0% in DMSO respectively. 2. The in vitro development rate from the frozen-thawed 8 cell embryos to the blastocyst were 83.6% in DMSO, 75.7% in glycerol, 52.2% in propylene glycol respectively. 3. The in vitro development rate from the frozen-thawed morula to the blastocyst were 84.2% in glycerol, 80.0% in DMSO, 66.6% in propylene glycol and 55.2% in ethylene glycol respectively.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.1
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• pp.59-65
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• 2004

Objectives: The aim of this study was to assess toxicities of cryoprotectants. Methods: Toxicities of two cryoprotectants, dimethyl sulfoxide (DMSO) and 1, 2-propanediol (PROH), were investigated using a murine embryo model. Female F-1 mice were stimulated with gonadotropin, induced ovulation with hCG and mated. Two cell embryos were collected and cultured after exposure to either DMSO or PROH. Embryo development was evaluated up to the blastocyst stage. Blastocysts were stained with bis-benzimide to evaluate the cell count and with terminal deoxynucleotidyl transferase mediated dUTP nick labeling (TUNEL) to assess apoptosis. Results: The total cell count of blastocysts that were treated with DMSO at the 2-cell stage was significantly lower than that were treated with PROH ($75.9{\pm}27.0$) or the control ($99.0{\pm}18.3$) (p<0.001). On comparison of two cryoprotectant treated groups, the DMSO treated group showed a decreased cell count compared with the PROH treated group (p<0.05). Both DMSO ($14.2{\pm}1.5$) and PROH ($11.2{\pm}1.4$) treated groups showed higher apoptosis rates of cells in the blastocyst compared with the control ($6.2{\pm}0.9$, p<0.0001). In addition, the DMSO treated group showed more apoptotic cells than the PROH treated group (p<0.001). Conclusions: The potential toxicity of cryoprotectants was uncovered by prolonged exposure of murine embryos to either DMSO or PROH at room temperature. When comparing two cryoprotective agents, PROH appeared to be less toxic than DMSO at least in a murine embryo model.

• Journal of Dairy Science and Biotechnology
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• v.37 no.3
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• pp.206-212
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• 2019

This study was conducted to investigate the effect of cryoprotectants on the storage stability of Lactobacillus fermentum SK152, which was isolated as a probiotic candidate. Solutions of 10% glucose, trehalose, dextrin, and skim milk powder were used as cryoprotectants. The survival rates of L. fermentum SK152 after freeze-drying were 5.6% (dextrin), 2.2% (skim milk powder), 1.7% (glucose), and 1.5% (trehalose), suggesting that dextrin was most effective at minimizing the cell death of L. fermentum SK152 by lyophilization. The survival rates of L. fermentum SK152 stored at 4℃ ranged from 37% (dextrin)-90% (skim milk powder) after 8 weeks, while those at 20℃ ranged from 4% (dextrin)-12% (skim milk powder) after 7 weeks, indicating that skim milk powder was the best at minimizing the cell death of L. fermentum SK152 during storage, irrespective of storage temperature, among the cryoprotectants used.

• Korean Journal of Food Science and Technology
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• v.21 no.3
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• pp.364-369
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• 1989

Experiments were carried out to investigate on the effect of cryoprotectants to the quality changes of pork and beef muscles during frozen storage . Beef and pork muscles were mixed with various cryoprotectants and stored at $-20^{\circ}C$ in a chest freezer for 12 weeks. Samples were analyzed for pH changes, TBA value, free atty acid contents, water and salt soluble protein extractability. The results obtained are summerized as follows. The pH value in all of cryoprotectants added samples were increased up to 0.25-0.5 as in non-treated samples . The TBA value, free fatty acid contents were increased with storage time as compared with the non-treated sample. Cryoprotectant effect on water soluble protein extractability was greater in pork than in beef muscle during frozen storage, especially in pork muscle treated CP-B, mixture of sorbitol, sucrose and sodium tripolyphosphate, as compared with non-treated sample. Cryoprotectant effect on salt soluble protein extractability during frozen storage was more pronounced in the beef muscle treated with CP-A which was mixture of sorbitol, mono sodium glutamate and sodium tripolyphosphate, and in the pork muscle treated with CP-B, mixture of sorbitol, sucrose and sodium tripolyphosphate than in the non-treated sample.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2004.07a
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• pp.26-26
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• 2004

• Korean Journal of Animal Reproduction
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• v.10 no.2
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• pp.175-181
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• 1986

These experiments were carried out to clarify the effects various kinds of cryoprotectants which were frequently used in freezing embryos of domestic animals on the survival of frozen-thawed mouse embryos. As cryoprotectant, glycerol, DMSO and methanol were used and the procedures of adding them in medium were practiced by one-step or six-step adding method. Morphologically normal mouse embryos developed to blastocyst by in vitro culture after freezing and thawing were transferred to pseudopregnant recipients by surgical procedures. The results obtained in these experiments were summarized as follows: 1. The survival rates of the frozen-thawed 8-cell embryos, morulas and blastocysts following one-step addition of glycerol were 83.6, 80.3 adn 70.3%, respectively, while following six-step addition of glycerol, 69.2, 56.3 and 66.7% respectively. 2. When glycerol, DMSO and methanol were used as cryoprotectant under the same condition of freezing and thawing, the survival rates of frozen-thawed embryos were 74.0, 76.1 and 37.6%, respectively. 3. The implantation rate of embryos transferred to pseudopregnant recipients after freezing and thawing was 49.2%.

• Reproductive and Developmental Biology
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• v.28 no.3
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• pp.155-159
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• 2004

This study was carried out to investigate the general characteristics and viability of sperm after freezing and thawing and the pregnancy rates after artificial insemination with thawed semen. The rates of viable sperm after slow and rapid freezing were 87.4±3.85% and 70.8±4.45%, respectively which were significantly lower than that of fresh semen control (91.7±3.45%). The mean concentration of epididymal sperm after dilution in 1.0 ml saline and. 3.0 ml extender in a various concentrations of cryoprotectants was 124.5±48.3 x 10/sup 6/ (range of 45 x 10/sup 6/ to 280 X 10/sup 6/ /ml). There was a significant difference not in the percentage of acrosome-reacted sperm, but in the percentage of capacitated sperm, between fresh and frozen-thawed epididymal semen. When frozen-thawed after diluting with tris-buffer extender containing glycerol, DMSO and ethylene glycol with concentration of 2 to 6%, the rates of epididymal sperm exposed to different cryoprotectants ranged from 14.4±4.7% to 20.7±5.8%, 17.8±5.2% to 36.5±4.9%, and 14.4±4.6% to 18.5±5.3%, respectively which were lower compare to fresh semen control. The pregnancy rate after artificial insemination with frozen semen was 70.6%, whereas that with fresh semen was 90.0% in dogs with naturally induced estrus.

• ALGAE
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• v.18 no.4
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• pp.321-331
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• 2003

Cryopreservation of sporothalli of the red alga Porphyra (P. seriata, P. yezoensis, P. tenera, P. pseudolinearis and P. dentata) was performed by the two-step cooling method in liquid nitrogen. The algal samples were suspended in various cryoprotective solutions, and slowly cooled to -40$^{\circ}C$ in 4 hours using a programmed freezer. After the first slow cooling the suspensions with cryoprotectants were immediately immersed in liquid nitrogen. The suspension from the programmed freezer was thawed quickly by agitation of the vial in a water bath at 40°C. When ice in the suspension of cryogenic vial was mostly melted, the vial was transferred to an ice bath for complete melting of the residual ice. The cryoprotectants in the vial were washed off by gradual dilution with seawater. The viability of the cell was assessed with neutral red staining. The viability of Porphyra samples ranged 54.6-70.9% when the mixed suspension of 10% dimethylsulfoxide and 0.5 M sorbitol in 50% seawater used as a cryoprotectant.

• Fisheries and Aquatic Sciences
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• v.17 no.3
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• pp.291-298
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• 2014

We investigate the effects of cryoprotectant mixtures on the quality of sand lance surimi (SLS) during storage at $-30^{\circ}C$. We monitored freeze-induced denaturation of myofibrillar protein in SLS and examined the texture profile of SLS gel. Freeze-induced denaturation was assessed by evaluating SLS $Ca^{+2}$-ATPase activity. SLS gels prepared with sorbitol or sucrose and a mixture of both as cryoprotectant. Higher concentrations of cryoprotectants resulted in significantly higher residual SLS $Ca^{+2}$-ATPase activity at the same storage time (P < 0.05). Residual $Ca^{2+}$-ATPase activity of SLS prepared with sorbitol was higher than that of sucrose when cryoprotectant concentration and storage period were same. A blend of sorbitol and sucrose resulted in a stronger cryoprptective effect of SLS myofibrillar protein than did sorbitol or sucrose alone. The presence of a phosphate compound in SOP (3% sorbitol + 0.2% phosphate compound) resulted in higher SLS $Ca^{2+}$-ATPase activity than that of did 5% sorbitol. The hardness, brittleness, and elasticity values and a folding test of the SLS gels were significantly affected by cryoprotectant concentrations and the storage time. Preference scores and acceptance for texture in a sensory evaluation of the SLS gels increased with increasing sorbitol or sucrose concentration.

• Journal of Aquaculture
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• v.21 no.1
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• pp.54-59
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• 2008

The present study aimed to find the best diluent and cryoprotectant for sperm cryopreservation of filefish Thamnaconus modestus. Two kinds of artificial seminal plasma(ASP1, ASP2), 0.3 M glucose and marine fish Ringer's solution(MFRS) were employed as diluent. Dimethyl sulfoxide(DMSO) and methanol as cryoprotectant were selected for sperm cryopreservation. Sperm was diluted at the ratio of 1:3 with diluents containing cryoprotectants and adjusted for final concentration at 5%, 10%, 15% and 20%. Mixed milt was frozen at liquid nitrogen vapor after equilibration for 5 min. The highest motility($40.5{\pm}2.8%$) and swimming speed($81.5{\pm}4.1{\mu}m/s$) of frozen/thawed sperm were observed in ASP1 diluent containing 10% DMSO and in ASP2 containing 15% DMSO, respectively. Results showed that cryopreservation with ASP as diluent and DMSO as cryoprotectant could be adopted for long term storage of filefish sperm.