Currently, the storage temperature of the production factory of medicinal herbs (hGMP) is about 5~ 12℃. This temperature is possible to suppress pest but can not kill the pests. For this reason, we need to lower the temperature during lethal time because the pest has often occurred inside the packaging of medicinal herbs in the distribution process in summer. In order to solve this problem, we have investigated the lethal time of the storage insect(Tribolium castaneum Herbst) After we froze medicinal herbs(Cnidium officinale Makino and Angelica gigas N. ) at approx. –70℃ and –15℃ respectively. We then investigated the change of bioactive components and exterior characteristics of medicinal herbs in order to determine whether there is a change of quality. The results were as follows. The lethal time of Tribolium castaneum Herbst is about 2 minutes if processed at approx. –70℃, while the other time is about 16 minutes at approx. –15℃. We investigated the change of quality after the treatment of the two medicinal herbs in the similar way but could not confirm the difference of color and brightness and the bioactive components on statistics. Through this research, it has been verified that the process of quick freezing for pest control can not affect the main quality of the medicinal herbs. Therefore this technology can be introduced in the manufacturing process of medicinal herbs through additional research.

Freezing of the pure water saturating a packed bed of spheres in a rectangular cavity is performed experimentally to investigate the effect of bead diameter on the heat transfer rate and temperature vs. time history for the pure water freezing experiments were compared with the freezing of the binary solution(HO-NaCl). Spherical soda-lime glass beads of 2.85mm and 6mm in average diameter constitute the porous medium. The system is cooled through its bottom surface, and the top is maintained at a temperature above the liquids. There was small effect on the temperature distribution in the system of the porosity difference.

This study was carried out to study the survival rate of thawed Hanwoo embryos frozen by the slow-rate freezing or the cryotop vitrification method. Hanwoo cumulus-oocyte complexes were recovered from ovaries at a slaughter house, matured for 20~22 hours, fertilized with Hanwoo semen for 5~6 hours, and cultured for 7~9 days in 38.5℃, 5% CO2 incubator. For freezing, Day 7∼9 blastocysts were collected. Embryos for the slow-rate freezing were equilibrated in 1.8 M ethylene glycol (EG) with Dulbecco's phosphate-buffered saline (D-PBS). Programmable cell freezer was precooled down to —7℃, and the straw was seeded during 8 minutes-holding time, and was cooled to —35℃ at the cooling rate of 0.3℃/min, and then was plunged and stored in liquid nitrogen. Embryos for the cryotop vitrification were treated in TCM199 with 0.5 M sucrose, 16% EG, 16% dimethylsulfoxide (DMSO). Embryos were then loaded individually onto cryotop and plunged directly into liquid nitrogen. The survival rates of embryos frozen by these two freezing methods were evaluated at 12 to 24h post-thawing. The survival rates of frozen/thawed Hanwoo embryos by the cryotop vitrification method (56.86 ± 26.53%) were slightly higher than those by the slow-rate freezing method (55.07 ± 26.43%) with no significant difference. Using the cryotop vitrification and the slow-rate freezing of Hanwoo blastocysts on Day 7 following in-vitro fertilization (IVF) treatment, the survival rates of frozen/thawed Hanwoo embryos were 72.65 ± 18.3% and 79.06 ± 17.8%, respectively. The survival rates by the cryotop vitrification were higher than those by the slow-rate freezing on both Day 8 and 9 with significantly higher survival rate on Day 9 (p<0.05). Using the cryotop vitrification and the slow-rate freezing of Hanwoo embryos to compare between three different blastocyst stages, the survival rates of the blastocyst stage embryos were 66.22 ± 18.8% and 45.76 ± 12.8%, respectively with higher survival rate by the vitrification method (p<0.05). And the survival rate of expanded blastocysts was higher than those of early blastocysts and blastocysts in two freezing methods with significantly higher survival rate by the slow-rate freezing method (p<0.05).

본 연구는 복숭아 유목기에 생육 촉진을 목적으로 질소를 과다 시용하여 동해발생의 우려가 있을 경우 에틸알콜 토양 살포에 의한 수체 내 전분증진과 증가된 전분이 동해에 미치는 영향을 구명하기 위해 수행하였다. 1. 전분함량은 질소 검정시비량 이상 시용에서는 시비량이 많아짐에 따라 감소하였고, 검정시비량의 75% 시용에서도 감 소하는 경향으로 부적절한 질소시비는 전분축적을 저해하였다. 2. 질소가 과다 시용된 복숭아 유목의 전분함량 증진을 위 해서는 9월 5일에 에틸알콜 50 mL를 토양 투입하는 것이 전 분함량을 가장 많이 증가시키는 효과적인 추비방법이었다. 3. 월동 전에 저온지역인 제천과 평창지역으로 수체를 이동 시켜 청원지역의 수체와 비교한 결과 4월 하순의 전분함량은 평창 >제천 >청원 순으로 높았다. 이는 평창지역은 12월 하 순 ~ 1월 상순, 제천지역은 2월 상순에 동해를 받아 전분이 당 으로 전환되어 세포의 결빙을 억제하는 기작이 원활하지 않았 기 때문이라고 판단되었다. 4. 질소 25% 이상 증비된 처리에 에틸알콜을 토양에 살포 하였을 때 동해경감 효과가 있었고, 월동 전 전분함량과 꽃눈동해와는 고도로 유의성 있는 부의 상관관계가 있었다.

This study was carried out to investigate synthetic extender for semen cryopreservation of Jeju Native Black Bull. The semen was collected using an artificial vagina and transported to the laboratory. The semen was diluted 1:1 by Tris-Egg yolk extender and contrifuged in 1,500 rpm for 15 minutes. The supernatant was removed. The pellect was diluted to final sperm concentration of 2×108/ml by doubling in every 30 minutes at 4℃ cold chamber. The semen was equilibrated for 4 hours at cold chamber and packed to 0.5 ml straw. The semen straws were located above 5 cm for 10 minutes. The height and duration affect the freezing speed by temperature. The frozen straw was plunged to LN2. The presented straws were examined the viability and motility after thawed at 37℃ water bath. Frozen-thawed sperm were evaluated sperm viability, membrane integrity and acrosome integrity. Post-thawed sperm viability has been significantly higher (p<0.05) in fresh sperm (93.27±1.62%) than frozen-thawed sperm (73.34±3.27%). However, there were no significant differences between fresh and frozen-thawed dead cell rate (7.35±2.63 vs, 13.71±2.85). In sperm motility, between Triladyl and AndroMed Extender, there was no significant different (72.86±2.83 vs, 81.47±2.48), similarly, the dead cell rates was similar (18.41±3.42% and 17.26±4.25). The results of our study suggest that AndroMed to the freezing extender showed more positive effect on the frozen-thawed spermatozoa in Jeju Native Black bull semen.

Artificial insemination (AI) has been performed widely in swine industry using fresh liquid sperm instead of frozen type of sperm. However fresh sperm are not able to preserve more than three days with optimal motility and other sperm parameters for the successful fertilization, since in vitro stored sperm has an oxidative stress that resulted increase of abnormality and acrosome reation. To overcome these major problems, novel preservative formulation is needed to neutralize the oxidative stress and to provide suitable physiological environment for sperm in in vitro. In this study, naturally derived substances such as Poncirus trifoliate (Trifoliate orange), Garcinia mangostana (Mangosteen), pig placenta and testis extracts were tested as sperm preservative agents. Placenta extracts (PE), trifoliate orange extracts (TOE), testes extracts (TE) and mangosteen extracts (ME) were applied to analyze specific parameters for sperm motion characteristics individually and combinatorial. Each individual extract treatment can accelerate the sperm motility but noticeably TOE, TE and ME treatments exhibited the considerable and significant preservation of sperm motility. PE, TE and ME showed a significant (p<0.05) increase in ALH after one week. Further we evaluated the five different combinations of these extracts on sperm motility and its motion characteristics. Surprisingly even after one week ME, TOE and TE combination significantly preserved the sperm motility about 75%. It is noteworthy that unlike individual extract treatment, combination of ME, TOE and TE simultaneously protect the sperm motility and its motion characteristics. Taken together these data conclude that addition of ME, TOE and TE can be effective for preservation of pig sperm.

외국에서 도입한 블루베리의 내한성을 평가하고, 목재수분계측기가 월동 중 블루베리 가지의 동해피해를 간편하고 신속하게 진단하는데 활용할 수 있는지를 알아보기 위하여 수행되었다. 월동 중 블루베리의 가지의 고사율은 0~100%로 다양하게 나타나 품종별 내한성의 차이가 컸다. 블루베리 가지의 저온처리에 따른 TTC 검정에서 품종별 OD값은 -40℃ 〈 -21℃ 〈 4℃ 순으로 처리온도가 낮을수록 낮게 나타났다. 블루베리 가지의 저온처리에 따른 가지절단면의 색의 검정은 가지고 사율에 의한 내한성과 다른 결과와 차이가 있었다. 목재수분계측기에 의해 측정된 살아있는 블루베리 가지의 월동 중 최저 수분함량은 약 15%였으며, 월동 중 블루베리 가지의 위치별 수분함량은 나무 아랫부분일수록 높고 가지 끝으로 갈수록 낮았으나, 봄으로 접어들면서 가지 끝의 수분함량이 점점 높아져 20~40% 범위로 측정되었다. 월동 중 동해피해를 받은 가지는 점점 건조되어 수분함량이 5% 이하로 낮아졌다. 동해를 받은 블루베리 가지의 수분함량은 14% 수준 이하일 것으로 추정되며, 목재수분계측기가 블루베리 가지의 동해피해를 현장에서 신속하게 진단하는데 활용될 수 있을 것으로 기대된다.

We conducted this study to investigate the quality characteristics of mashed red pepper(MRP) with different freezing(－20, －30, －40, －50, －60 or －70℃) and thawing(4, 10, 20, 30, 40 or 50℃) temperature. Frozen and thawed MRP was evaluated for ascorbic acid contents(AsA), capsaicinoids contents, free sugar contents, ASTA value, and flavor patterns. The AsA of frozen MRP with freezing temperature showed a range of 67.08～80.35 ㎎/100 g FW, and the mean AsA losses after frozen were 57～64% compared to raw red pepper. Capsaicinoids contents, free sugar contents, and ASTA values of frozen MRP with freezing temperature were no significant difference compared to raw red pepper. The AsA of thawed MRP with thawing temperature showed a range of 70.34～81.59 ㎎/100 g FW, and the mean AsA losses after thawed were 45.12～52.69% compared to raw red pepper. Capsaicinoids contents and free sugar contents of thawed MRP with thawing temperature were no significant difference compared to raw red pepper, whereas the ASTA value decreased according to the increasing of thawing temperature. Flavor patterns of thawed MRP after 20～50℃ thawing were clearly different from the raw red pepper. Overall, it is recommended that the best freezing and thawing temperature for preserving the quality of MRP is freezing at －20℃ and thawing at 10℃.

This study was designed to determine whether low-density lipoproteins (LDL) from egg yolk and taurine, hypotaurine and trehalose as antioxidant in extender improve the freezability and fertility of Korean Jeju Black Bull semen. The semen was cryopreserved with tris egg yolk extenders containing 7% glycerol and treated 4% LDL, 20 mM taurine, hypotaurine and trehalose. Frozen-thawed sperm were evaluated motility, viability, membrane, and acrosome integrity and sperm penetration ability. The results were compared to semen cryopreserved in tris egg yolk extender only as control. Frozen-thawed semen evaluation cleary indicated that the addition of LDL and LDL-antioxidants (taurine, hypotaurine and trehalose) combination were significantly improved (p<0.05) the viability (%; with staining test using eosin-Y) compared to control spermatozoa. Also, in membrane integrity (%; with supravital hypo-osmotic swelling test), not only LDL-antioxiants combination but also LDL were significantly increased (p<0.05) the swelled sperm using HOST compared to control. Sperm acrosome integrity state was classified by CTC (chlortetracycline) staining test. F pattern was significantly increased in LDL-antioxidant combination than control (p<0.05) and B pattern was not significantly differences among all treatments and control. However, AR pattern was significantly decreased in LDL-antioxidants combination than control (p<0.05). Pronucleus formation and sperm penetration index (SFI) were significantly increased in LDL and LDL-antioxidants combination than control (p<0.05). Especially, LDL-taurine significantly improved pronucleus fomation and SFI than LDL (p<0.05). It was concluded that LDL and LDL-antioxidants in extender improved the freezability and fertility of Korean Jeju Black bull spermatozoa.

The main purpose of this study is to estimate the effect of adding Tea-N-Tris to the freezing buffer for miniaturepig sperm. In particular, we attempted to identify the association between the MMPs expression and the survival and viability of sperms. Prior to freezing, sperms in LEY without Tea-N-Tris showed 40.3±2.8% viability and 60.3±1.3% acrosome intact rate at 4℃. After freezing, sperms stored in LEY (lactose+Egg yolk) with Tea-N-Tris (=TLE) showed the highest viability (57.4±1.8%) and acrosome intact rate (65.6±4.6%). In accordance with this, DNA fragmentation was the highest among sperms frozen in LEY while the lowest fragmentation was observed among sperms frozen in TLE. When these sperms were used for in vitro fertilization (IVF), the LEY group showed lower rate of blastocyst development, although the difference was not statistically significant. Meanwhile the rate of blastocyst development appeared similar when sperms from TLE and TFGE (Tea-N-Tris+Fructose+ Glucose+Egg yolk) group were used for IVF. We observed MMPs expression in all sperm groups, with pro-MMP showing lower expression than active MMPs. The expression of MMP-9 and MMP-2 was the highest in sperms frozen in LEY, Meanwhile, sperms from the TFGE and TLE group showed lower level of MMP-9 and MMP-2 expression in the order of TLE being the lowest. Together, these results indicate that adding Tea-N-Tris to the sperm freezing buffer would not only suppress MMPs expression but also minimize DNA fragmentation, providing a mean to improve the success rate in the in vitro manipulation of miniaturepig sperms. * This work was supported by BioGreen 21 Program (No. PJ008029). Rural Development Administation, Republic of Korea.

The objective of this study was to assess the effect on post-thawed sperm motility, viability and acrosome integrity of boar semen frozen in the freezing extender with chicken or duck egg yolks. The Sperm rich fraction of ejaculates from three Duroc boars were collected by a glove-hand technique. Samples with more than 80% motile sperm were used for this experiment. Semen was diluted with freezing extender (LEY) containing 11% (v/v) lactose, 20% (v/v) hen egg yolk with 3.5% (v/v) glycerol, and 0.5% (v/v) Orvus Es Paste(OEP, Nova Chemical Sales Inc., Scituate, MA. USA) to yield a final sperm concentration of 5×108 cells/ml. Following complete dilution, semen samples were loaded in 0.5 ml French medium straws (IMV technologies, France) and transferred to programmable semen freezer (SY-LAB Gerate GmbH, Austria). For freezing the semen samples, each straw was cooled from 5℃ to — 5℃ at 6℃/min, auto-seeding at — 5℃ and held for 60sec, samples were then cooled from — 5 to — 80℃ at 40℃/min, and thereafter from — 80℃ to — 150℃ at 60℃/min. The yolks used were sourced from fresh chicken and duck eggs. To evaluate the post-thaw sperm quality, semen was thawed at 38℃ for 20 sec and sperm motility, viability and acrosome integrity were assessed. Motility was assessed for %motile cell characteristics using computer-assisted semen analysis (CASA; SAIS SI-100, Medical supply, Korea). The percentage of sperm viability was assessed using LIVE/DEAD® sperm viability kit (Molecular probes, Eugene, OR, USA). The acrosome integrity was assessed by FITC-PNA staining. Sperm quality in terms of motility, viability and acrosome integrity showed higher after freezing in medium containing duck yolk than chicken yolk. However, there was no significant difference in sperm quality for the different types of yolk(p>0.05). * The result of this study showed that there was no significant difference between the egg yolk types when considering the sperm motility, viability and acrosome integrity of boar semen frozen in the freezing extender with chicken or duck egg yolks.

현재 도로포장 설계법에 따르면, 동상방지층의 두께는 지역별 온도조건에 따라 결정되는 동결깊이에 의해 결정되며 동상방지층의 지지력은 설계에서 고려되지 않고 있다. 동상방지층을 도로포장체에서 구조층으로 고려할 경우에는 기존 도로포장층의 두께를 감소시킬 수 있으며 보다 경제적인 도로 포장단면을 구성할 수 있다. 본 연구에서는 동상방지층의 지지력을 평가하기 위한 통계적 모형을 개발하였다. 동상방지층의 구조적 역할을 규명하고 동상방지층 구조적 평가 모형 개발을 위하여 2m 이하 저성토부, 절토부 및 절성경계부 등을 구분하여 포장 하부층에서 Falling Weight Deflectormeter(FWD) 시험을 계절별로 수행하였다. 본 시험은 동방방지층의 유무에 따른 지지력 차이를 규명하기 위하여 동방방지층이 있는 구간과 없는 구간으로 구분하여 수행하였다. 본 시험결과, 동상방지층이 설치된 구간에서의 FWD 처짐량이 동상방지층 미설치 구간에 비해 0.4~82.6% 작게 측정되어 동상방지층이 포장체에서 지지력을 검증하였다. 다양한 FWD 처짐지수와 동상방지층 두께와의 상관관계를 조사한 결과, 보조기층 파손지수의 차이값(δBDI)과 동상방지층 두께와의 상관도가 가장 높았다. 본 논문에서는 δBDI값을 선형혼합효과 모형에 적용하여 동상방지층을 구조적으로 평가할 수 있는 모형을 개발하였다.

The purpose of this study was to examine the effect of growing stages of the Korean Native Striped Bull (KNSB) on the freezability and fertility of frozen-thawed semen. First, we investigated the total motility (TM) and progressive motility (PM) according to the diluent used for semen freezing. Second, we examined the effect of the age of KNSB on semen volume, TM and PM of fresh and frozen-thawed semen. Third, we examined the effect of frozen semen from the different age of KNSB on the fertilization rate, and the artificial insemination pregnancy rate. The diluents used in this experiment were Triladyl and Tris-egg yolk extender (EYE). Semen was collected from 5 KNSB in the growing stage (15 months) and 5 adult KNSB (36 months). When Triladyl or Tris-EYE extender was used for semen freezing, there was no difference of the mean TM and the mean PM. However, the mean TM was significantly higher in Bull No. 1885 than Bull No. 4283 ( <0.05). The mean volume of semen collected from the 15-month-old bulls (2.3 ml) was significantly lower ( <0.05) than that from the 36-month-old bulls (5.0 ml). The mean semen concentration was similar for the 15-month-old ( spermatozoa/ml) and 36-month-old ( spermatozoa/ml) bulls. For the 15-month-old and 36-month-old bulls, the mean TM of fresh semen were 93.7% and 88.3%, respectively, and the mean PM were 97.0% and 88.3%, respectively; the 15-month-old bulls showed a particularly high PM ( <0.05). For the 15-month-old and 36-month-old bulls, the mean TM (56.0% and 58.0%, respectively) and the mean PM (64.0% and 70.7%, respectively) of frozen-thawed semen did not differ. The development rates of embryos after fertilization and the pregnancy rate after artificial insemination using frozen-thawed semen did not differ according to the bull's age. In summary, semen volume differed according to the bull's age, but semen concentration and survival rate, the fertilization rate, and the pregnancy rate did not differ according to the stripe bull's age. Accordingly, semen from bulls in the growing stage can be collected and frozen for the preservation and multiplication of rare livestock.

서울 기상 관측소(서울시 종로구 송월동, 37˚34'N, 126˚57'E, 이하 송월)의 일 최저기온과 결빙 자료, 그리고 인근인 한강 관측소(37˚30'N, 126˚57'E, 이하 한강)의 결빙 자료를 사용하여, 1907-2006년(100년) 동안 한국 서울에서의 결빙기후의 장기 변동을 조사하였다. 송월의 결빙은 옥외 노장의 물이 동결하는 현상으로 정의하였고, 이곳에서 6 km 떨어진 한강의 결빙은 한강 대교 남단에서 2번째와 4번째 교각의 상류 100 m 부근이 완전히 얼었을 때로 정의한다. 평균 첫 결빙일은 송월이 10월 28일, 한강이 12월 28일이며, 각각 0.78 days decade-1와 3.47 days decade-1로 점차 늦어지는 추세를 나타냈다. 평균 연 결빙 일수는 송월이 159.06일, 한강이 50.33일이며, 각각 2.01 days decade-1와 5.24 days decade-1로 감소하는 추세를 보였다. 특히, 1950년대 후반을 기점으로 변화율이 급변하였는데, 한강에서 100년 동안 발생한 7번의 무결빙해(1960, 1971, 1972, 1978, 1988, 1991, and 2006)도 모두 1950년대 이후에 발생하였다. 송월과 한강에서 첫 결빙일의 일 최저기온은 평균적으로 각각 0.55˚C와 -12.22˚C이다. 영하의 일 최저기온이 송월은 6.43일, 한강은 8.94일 지속되고 난 뒤 첫 결빙이 발생하였다. 송월과 한강의 연 최저기온에 대해서 첫 결빙일은 양의 상관을 보였고, 결빙 일수는 음의 상관을 보였다. 이러한 결과는 결빙기후의 변화가 기후변화의 일환으로 나타나는 기온변화와 유관함을 보였다. 향후 더 넓은 지역에서 다양한 추가 연구를 실시함으로써, 지속적으로 결빙기후를 주시하고자 한다.

The objective of this study was to evaluate the effect of Tea-N-tris medium on the sperm viability and acrosomal morphology for semen of normal and miniaure pig by type of freezing extender. The present study was to determine of Tea-N-tris (0.02 g/ml) effect to freezing extender LEY(Lactose 11% + Egg yolk 20%) and FGE(Fructose 3%+Glucose 7%+Egg yolk 20%) for the spermatozoa viability, acrosomal morphology and DNA fragmented analysis from normal and miniature pig semen, were evaluated freezing extender TFGE, TLE and LEY during thawing at 37℃ for 45 sec and 75℃ for 5 sec, respectively. Interestingly, the result that sperm after addition of Tea-N-tris extender(TFGE, TLE) during 15 4℃ cooling significantly increased the viability(p<0.05), as compared to than of sperm cooling in LEY extender, but lower the percentage of AR(acrosome reacted spermatozoa) pattern than LEY extender. The sperm viability and AR pattern after freezing was appeared like sperm cooling method pattern. And treatment spermatozoa during freezing after addition of Tea-N-tris extender significantly (p<0.05) increased the viability and AR to miniature pig sperm, than normal pig sperm, but most highly percentage of viability and AR pattern to normal pig sperm during freezing in LEY extender. Chromosomal DNA fragmentation increased from LEY extender to sperm of normal and miniature pig, but decreased from the Tea-N-tris extender. Therefore, suggest that Tea-N-tris freezing extender method for freezing of miniature pig sperm is required for increasing viability. This Study was supported by Technology Development Program for Agriculture and Forestry, Ministry for Food, Agriculture, Forestry and Fisheries, Republic of Korea.

The objective of this study was to examine effect of ethylene glycol for semen cryopreservation in Korean Jeju Black Bull. The semen was cryopreserved with extenders containing cryoprotectants (7% glycerol and 3%, 5%, 7% ethylene glycol) and packed to 0.5 ml straws. The semen straws were located above 3 cm of liquid nitrogen for 5 min, 5 cm for 10 min and 8 cm for 10 min. And then frozen straw was plunged into LN2. Post-thawed sperm motility, viability and membrane integrity were significantly higher in 5% ethylene glycol (72.5±5.00%, 54.88±0.66% and 46.00±2.40%; p<0.05). Motility and viability were similar between 7% glycerol and 5% ethylene glycol. However, the membrane integrity was significantly higher in 5% ethylene glycol (34.69±4.64% vs 46.00±2.40%; p<0.05). The viability and membrane integrity were significantly higher in 5 cm for 10 min and 8 cm for 10 min than 3 cm for 5 min (viability: 55.81±2.94, 55.19±3.34 vs 47.94±3.48%; p<0.05 and membrane integrity: 44.94±3.51, 46.06±2.25 vs 40.38±1.03%; p<0.05). The percentage of capacitated sperm assessed by CTC staining, percentage of F pattern was higher in 7% glycerol, 5% and 7% ethylene glycol, and AR pattern was significantly higher in 3% ethylene glycol. F pattern was significantly increased in 5 cm for 10 min and 8 cm for 10 min (p<0.05), but AR pattern was significantly increased in 3 cm for 5 min (p<0.05).

The aim of this study was to evaluate the effect of α-tocopherol on blastocysts development and subsequent cryosurvival of the vitrification. The α-tocopherol(0, 100, 200, 400 μM) was added in to culture medium for the bovine embryos. The blasocysts from the α-tocopherol and untreated control groups were then frozen-thawed, and their cryosurvival was assessed by in vitro culture for 48 h. There were no differences in the overall cleavage rate(56.14±4.66, 58.18±4.70, 62.97±6.86 and 51.17±7.28) among four treatment groups. However, in blastocyst development and total cell number were significantly higher in α-tocopherol 200 μM(38.60±7.12; 106.33±3.50) to culture medium than other treatment groups(29.30±5.24, 31.60±7.12 and 26.37±4.18; 101.36±5.12, 97.27±2.87, and 91.23±7.52 respectively). Before and after vitrification, the total cell number and blastocyst development of embryo were significantly higher in July to August than September to October. In conclusion, addition of α-tocopherol 200 μM to in vitro bovine embryo culture medium was beneficial for improving embryo quality by decreasing the embryo damage blsstocysts cell number and improving the tolerance of the embryos to cryopreservation.