• 대한수의학회지
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• 제31권4호
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• pp.523-527
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• 1991

The effects of ethylene glycol, DMSO and glycerol as cryoprotectants and the effect of concentrations(0, 0.25, 0.5 and 1.0M) of sucrose in the diluent on the viability of the aggregated morulae frozen rapidly in liquid nitrogen$(LN_2)$ vapour were examined. The morulae were produced by aggregation of ICR and CBA mice embryos at 8-cell stage. Before freezing the embryos were equilibrated in 1.5M cryoprotectants+0.25M sucrose in oae-step or in 3.0M cryoprotectants+0.25M sucrose in two-steps. The embryos were pipetted into the freezing medium fraction of 0.25ml plastic straws. The straws were frozeu by directly transfer into $LN_2$ vapour(about lcm above $LN_2$) for 2 minutes, and then plunged into $LN_2$. After thawing the cryoprotectants were diluted with 0, 0.25, 0.5 or 1.0M sucrose solution. The post-thawed in vitro viability of the aggregated embryos was significantly dependent on the type and concentration of cryoprotectants in the freezing medium and also on the concentration of sucrose in the diluent. When the aggregated embryos were equilibrated in 1.5M cryoprotectants +0.25M sucrose in one-step and diluted with 0.5M sucrose after thawing, the survival rate of the embryo5 was significantly(p<0.05) higher in DMSO(62.5%) or ethylene glycol(52.2%) than in glycerol(33.3 %). In the case that the concentration of the cryoprotectants was raised to 3.0M in two-steps, thc higher survival rate of the embryos was obtained in ethylene glycol or glycerol than in DMSO followed by diluting them with 0.5 or 1.0M sucrose after thawing(p<0.01).

• 한국수정란이식학회지
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• 제13권2호
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• pp.147-157
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• 1998

This study was conducted to investigate the effects of in vitro fertilization, culture and embryo development according to in vitro maturation rate, protectant composition and equilibrium time after frozen /thawing of bovine immature oocytes. This results obtained in studies on the effect of different cryoprotectants on the viability, maturation and development of in vitro bovine oocytes were as follow: 1.The post-thawing of immature oocytes matured to metaphase II during culture time for 0 to 26 h, and those group (62~3%) were low than control group (76.7%). The optimal maturation time of frozen-thawed immature oocytes was at 24 h. 2.The viability of cryopreserved immature oocytes was not affected by sort of cryoprotectants. The developmental competence of frozen4hawed oocytes was not affected by cryoprotectants. These results indicate that an optimal maturation time of frozen /thawed immature oocytes was at 24h. Furthermore the viability of cryopreserved immature oocytes was not affected by sort of cryoprotectants and developmental competence of frozen /thawed oocytes.

• Journal of Pharmaceutical Investigation
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• 제40권1호
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• pp.39-43
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• 2010

Solid lipid nanoparticles (SLNs) were freeze-dried to obtain a stable solid dosage form with the aid of various cryoprotectants such as trehalose, sucrose, glucose, fructose, and glycerol. Tricaprin(TC) and trilaurin(TL) were used as lipid matrices for SLNs and stabilizers were egg phosphatidylcholine and pegylated phospholipid. All cryoprotectants tested did not cause changes in mean particle size of SLNs when mixed with SLNs before freeze-drying. However, the mean particle sizes of reconstituted SLNs after freeze-drying were significantly different from those of the un-lyophilized original SLN dispersions depending on the types and concentration of cryoprotectants. Although the freeze-dried SLNs without any cryoprotectants were easily reconstituted by hand-shaking, the mean particle size drastically increased (> $8\;{\mu}m$ for TC SLNs and around $1\;{\mu}m$ for TL SLNs) compared to that of un-lyophilized original dispersion (97 nm for TC SLNs and 164 nm for TL SLNs). Trehalose and sucrose were the most effective additives to protect the SLNs during lyophilization. The reconstituted SLNs were physically stable for 24 hours when lyophilized with 12.5% trehalose, sucrose, glucose, fructose or glycerol.

• 한국수정란이식학회지
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• 제10권3호
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• pp.193-202
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• 1995

In order to improve the cryopreservatory techniques of livestock embryos, the quick freezing method which is directly plunged in liquid nitrogen via prefreezing procedure without freezing machine was carried out for mouse embryos treated with permeable and nonpermeable cryoprotectants. The viability of frozen-thawed embryos were evaluated by FDA vital dye test. The results obtained was summaried as follows: 1. A total of 720 embryos were recovered from frozen embryos for viability test. Evalution of the fluorescein diacetate(FDA) vital dye test with mice embryos were resulted of 2.3 total mean score - evaluted in orderly higher mean grade of P3 453 (63%), P2 133(18%), P1 51(7%) and P0 83(12%). 2. An all-round evalution of these combination, the highest viability was showed in 3M ethylene glycol + 0. 25M trehalose treated with the copper prefreezing. 3. Effects of permeable and nonpermeable cryoprotectants combination were evaluated by means FDA score. 3M ethylene glycol + 0.25M trehalose showed the highest survival rates of 2.8 mean FDA score. 4. Effects of permeable cryoprotectants were evaluated by mean FDA score but the results were not significantly different each other. 5. In evalution of the nonpermeable cryoprotectants, 0. 25M trehalose obtalned higher mean FDA score than of 0.25M sucrose and it was significantly different(P<0.05). 6. There was no significantly difference between copper and stainless-steel in prefreezing procedures.

• 한국가축번식학회지
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• 제16권3호
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• pp.269-276
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• 1992

Studies were carried out to find the freezing media which gives no ice crystals in single(glycerol, ethylene glycol, dimethyl sulfoxide(DMSO)) and mixture solutions(glycerol＋propylene glycol, glycerol＋ethylene glycol) of permeable cryoprotectants in vitrification solution and to study effects of VS on the survival of vitrified mouse morulae. The results are summarized as follows: 1. In toxicity test of permeable cryoprotectants, 30% glycerol of single solution showed the highest FDA-score(4.1) in mouse morulae frozen compared among other single solutions. The FDA-score(4.1) of 30% glycerol was higher than 30% ethylene glycol(3.6) and DMSO(1.4( (P<0.05). 2. 20, 30 or 40% single solution of permeable cryoprotectants containing m-PBS with 10% sucrose and 20% BSA was not crystallized during cooling, but crystallized during warming. However, the 30% mixture solution of the two permeable cryoprotectants was not crystallized both during cooling and warming.3. When mouse morulae were frozen in 30% mixture solutions of two permeable cryoprotectants(glycerol and propylene glycol, glycerol and ethylene glycol), highest FDA-score(4.5) was obtained in a mixture solution of 20% glycerol and 10% ethylene glycol(20G10E) than other 30% mixture solutions(10G20E, 15G15E, 20G10P, 15G15P, 10G20P) and there was significant difference between 20G10E and 10G20E(P<0.05).

• 한국식품과학회지
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• 제31권4호
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• pp.957-963
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• 1999

두부의 냉동저장중에 일어나는 단백질 변성과 이로 인하여 일어나는 조직감 열화현상을 완화하기 위하여 항냉동제 첨가 두부의 제조 공정을 개발하고 항냉동제 최적 배합비율과 첨가량을 조사하였다. 여러 가지 항냉동제를 첨가하여 비압착 두부를 제조하여 냉동저장한 후 다시 해동한 냉동저장 두부의 조직감 변화를 반응표면분석법(RSM)으로 분석하고 관능검사 결과와 비교하였다. 냉동으로 인한 두부의 조직감 변화를 최소화하는데는 glucose, glycerol, sorbitol, sodium tripolyphosphate가 단일 항냉동제로서 효과가 있었으며, glycerol과 sodium tripolyphosphate 사이에서는 상호작용도 있는 것으로 나타났다. 결론적으로 냉동두부에 항냉동제를 단독으로 첨가하는 것보다는 여러 가지를 혼합하여 첨가하는 것이 두부조직의 열화방지에 더 효과적인 것으로 나타났다. 조직감 변화만을 고려한 절단시험의 RSM을 통하여 계산된 냉동두부의 항냉동제 최적 첨가량은 glucose 2.1%, glycerol 6.9%. sorbitol 2.1%, propylene glycol 0.4%, sodium tripolyphosphate 0.3% 이었으며, 조직감과 함께 맛과 향까지 고려한 전체 기호도 조사에 의한 항냉동제 최적 첨가량은 glucose 1%, glycerol 2%, sor-bitol 1%, propylene glycol 0.2%, sodium tripolyphosphate 0.5% 이었다.

• 한국가축번식학회지
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• 제18권1호
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• pp.15-23
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• 1994

This study was carried out to investigate the effects of concentration, kinds of cryoprotectants, equilibration time, optimum thawing temperature on the survival rate of rapidly frozen porcine embryos. The porcine embryos following dehydration by cryoprotectants containing sucrose were directly plunged into liquid nitrogen and thawed in 30, 35 or 37$^{\circ}C$ water bath, Survival rate was defined as development rate on in vitro culture or FDA-test. The results are summarized as follows : 1. The high survival rate of porcine frozen embryos after rapidly thawed in freezing medium was attained 2.0M DMSO, 2.0M glycerol, 2.0M propanediol, 1.5M ethyleneglycol. 2. The high survival rate of porcine frozen embryos after rapidly thawed in freezing medium was obtained using single cryoprotectant(16.6~40.0%) than mixed cryoprotectants(12.5~33.3%). 3. The eqilibration time on the survival rate of rapidly thawed porcine frozen embryos was attained after short period of time(15.0~33.3%) in the freezing medium higher than long period of time(9.10~30.0%). 4. The thawing temperature on the survival rate of rapidly thawed porcine frozen embryos was attained at 3$0^{\circ}C$ of thawing temperature(33.3~40.6%) in the freezing medium higher than 25 or 37$^{\circ}C$ of thawing temperature.

• Journal of Microbiology and Biotechnology
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• 제19권8호
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• pp.810-817
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• 2009

The effect of cryoprotectants (maltodextrin+glycerol) and cryoprotectants+antioxidant [ascorbic acid and/or butylated hydroxytoluene (BHT)] mixtures on the survival, electrolyte leakage, and lipid degradation of freeze-dried Weissella paramesenteroides LC11 during storage was investigated and compared with that of the control (cells without additives) over a 90-day storage period at 4 or $20^{\circ}C$ in glass tubes with water activity ($a_w$) of 0.23. The survival, electrolyte leakage, and lipid degradation were evaluated through colony counts, electrical conductivity, and thiobarbituric acid reactive substances (TBARS) content, respectively. The fatty acids composition was determined by gas chromatography, in both the total lipid extract and the polar lipid fraction, and compared with that of the control after the 90-day storage period. As the storage proceeded, increases in leakage value and TBARS content, as well as a decrease in viability, were observed. After 90 days of storage, the major fatty acids found in both the total lipid extract and the polar lipid fraction were palmitic (16:0), palmitoleic (16:1), stearic (18:0), oleic (18:1), linoleic (18:2), and linolenic (18:3) acids. The survival, leakage value, TBARS content and 18:2/16:0 or 18:3/16:0 ratio were the greatest for the protected strain held at $4^{\circ}C$. Cells with the cryoprotectants+BHT mixture showed the highest percentage of survival and 18:2/16:0 or 18:3/16:0 ratio in both lipid extracts, as well as the lowest leakage value and TBARS content after the 90-day storage period. Drying cells with the cryoprotectants+BHT mixture considerably slowed down polar lipid degradation and loss of membrane integrity, resulting in improved viability during storage.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제31권3호
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• pp.228-238
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• 2005

Purpose: Several cryoprotectants are in use to help the survival of cells during cryopreservation of bone in maxillofacial region. Among them, $Me_2SO$(dimethyl sulfoxide), EG(ethylene glycol), sucrose were used for experimentally created defects with accompanying cryopreserved bone graft in the rabbit model. The aim of this study is to analyze the effect of above mentioned agents on bone formation using histologic and histomorphometrical methods, thus to provide experimental support for clinical application of these agents. Materials and methods: Nine rabbits were used as experimental animals. Surgical defects were created on the distal femoral heads and mesial tibial heads of each animal using trephine drill(5mm diameter and 5mm length). The harvested bones were cryopreserved in $-80^{\circ}C$ refrigerator for one week. The defects were filled with cryopreserved bone with cryoprotectants as experimental groups and cryopreserved bone without cryoprotectant as control. Then, the animals were sacrificed at 1, 2, and 3 weeks after surgery. With Goldner's modified Masson trichrome staining and semiautomatic image analysis system, we observed the change of the cells and bone formation. Results: After bone graft, bone formation and active remodeling process were examined in all experimental groups and the control. But the intensity of such activities of the control were somewhat weaker than that of the experiments. Especially $Me_2SO$+sucrose group was the best in bone formation and bone remodeling. $Me_2SO$ group was more than that of EG group in bone fomation. Sucrose seems to be helpful in survival of the bone cell. Histologic findings showed superior bony quantity and quality in experimental groups than that in control. Conclusions: The data from this study provides the basis for future studies for evaluating the effect of cryoprotectants in the cryopreservation of bone and clinical study for predictable use of these agents.

• 한국양식학회지
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• 제16권4호
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• pp.262-266
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• 2003

어류 배의 냉동보존 방법을 개발하기 위한 연구의 일환으로 감섬돔, Acanthopagrus schlegeli의 초기 배를 대상으로 동해방지제인 DMSO, ethylene glycol 및 glycerol의 최적 농도와 처리시간을 파악하기 위한 연구가 수행되었다. 동해방지제의 적정 농도를 평가하기 위하여 배의 사망률, 부화율 및 반수치사 농도를 조사하였다. 동해방지제로 처리된 감성돔 배의 사망률은 침지농도나 시간과 밀접히 연관되었으며, 감성돔 배에 대한 독성은 DMSO, ethylene glycol, glycerol의 순서로 낮았다. 배의 사망률, 부화율 및 반수치사농도에 대한 결과로부터 감성돔 배를 냉동보존하기 위한 가장 적합한 동해방지제는 DMSO였으며, 다음이 ethylene glycol이었다. 20분의 처리시간에서 효과적인 DMSO의 침지 농도는 2.0∼2.25 M이었으며, ethylene glycol의 경우 1.0∼1.78 M이었다.