• Clinical and Experimental Reproductive Medicine
• /
• v.33 no.4
• /
• pp.237-243
• /
• 2006

Objective: The aim of this study was to evaluate the efficacy of frozen-thawed ET in poor prognosis patients such as the old age (38~44 years; OA group) and the patients who did not achieve clinical pregnancy with the first fresh ET cycle (non-pregnant patients; NP group). Methods: Laboratory and clinical data were collected from fresh and frozen-thawed ET cycles of OA and NP group. Controlled ovarian hyperstimulation (COH) and conventional insemination or ICSI, in vitro culture and ET were performed by routine procedures. Supernumerary embryos were frozen by the slow freezing method, and frozen embryos were thawed by the rapid thawing method. Embryo development, pregnancy and implantation rates were statistically analyzed by Student t-test and chi square test Results: Mean ages were similar between fresh ET ($40.0{\pm}1.8$ years, n=206) and frozen-thawed ET ($39.9{\pm}1.9$ years, n=69) cycles in OA group. However, the clinical pregnancy and implantation rate of subsequent frozen-thawed ET significantly higher than those of fresh ET cycles (29.0% and 11.2% vs. 16.5% and 7.0%, p<0.05). In NP group, there was no difference in the mean age between fresh ET ($31.2{\pm}2.3$ years, n=40) and frozen-thawed ET ($31.9{\pm}3.1$ years, n=119) in subsequent cycles. The clinical pregnancy and implantation rates were similar between the subsequent fresh ET (42.5% and 22.6%) and the frozen-thawed ET (40.3% and 18.8%). Conclusion: In old age patients, higher pregnancy rate of frozen-thawed ET compared to fresh ET cycles in this study. It may be related that better uterine environments for implantation in frozen-thawed ET cycles than that of non-physiological hormonal condition in uterus of fresh COH cycles.

• Clinical and Experimental Reproductive Medicine
• /
• v.26 no.2
• /
• pp.231-238
• /
• 1999

The objective of this study was to examine the effect of developmental stage and embryo age of in vitro produced bovine blastocysts after vitrification and thawing. In vitro cultured day 8 blastocysts after IVF were equilibrated 20% ethylene glycol (EG) for 3 min. and were vitrified using EFS40, which is consisted of 40% EG, 18% ficoll, 0.3M sucrose and 10% FBS added in mDPBS for 30 sec. before being plunged into $LN_2$. Also, survival in vitro was assessed by re-expansion and hatching or hatched at 24 hand 48 h postwarming, respectively. The results obtained in these experiments were summarized as follows; 1) When the embryos were cultured for 8 day after IVF, 41.0% of the cleaved embryos developed to the blastocysts (early; 7.6%, expanded; 22.9%, hatching; 4.6% and hatched; 5.9%). 2) When the embryos were exposed or vitrified to the freezing solution, the re-expansion of vitrified embryos (73.3%) was significantly lower than that of control and exposed embryos (100, 97.0%) (p<0.05). But the formation rate of hatching or hatched blastocysts of vitrified embryos (66.7, 46.7%) at 48h after thawing was similar to that of exposed embryos (66.7, 39.4%) but not control (100, 100%) (p<0.01). However, in the total cell numbers of those developed hatched blastocysts, there were not significantly different among the treatment groups. 3) When the embryo survival rates by different developmental stage were examined, the re-expansion was not different among the groups $(64.5{\sim}75.6%)$. After warming 48 h, the hatching and hatched formation of early blastocysts (25.8, 9.7%) was significantly lower than those of expanded (69.7, 39.4%) and hatching blastocysts (53.3, 43.3%) (p<0.05). 4) In addition, when the expanded blastocysts at day 7, 8 and 9 were vitrified, the re-expansion of day 8 and 9 embryos was significantly lower than that of day 7 (day 7; 93.9%, day 8; 75.8% and day 9; 87.5%) (p<0.05). However, the rates of development to hatched blastocysts were no difference among the groups (day 7; 36.4%, day 8; 36.4% and day 9; 31.3%). These results suggested that in vitro produced expanded or hatching blastocysts can be efficiently cryopreserved by the two-step vitrification method using EFS40.

• Parasites, Hosts and Diseases
• /
• v.28 no.4
• /
• pp.241-252
• /
• 1990

In a basic attempt to develop the prophylactic and therapeutic measures on intestinal giantcystic disease of the Israel carp, C), prinks carpio nudum, the effects of physical and chemical factors on viability or survival of the spores of Thelchcnellus kiteuei were checked in vitro by means of extrusion test on the polar filament. When the fresh spores suspended with 0.45% and 0.9% scdium chloride solution and distilled water were laid at $5^{\circ}C$ and $28^{\circ}C$ for short terms, the extrusion rates increased until the 3rd day, meanwhile when son;e of them were suspended with Tyrode's solution at $-70^{\circ}C$ the rates increased gradually until the 8th day. Viabilities of the spores suspended with 0.9% saline and added antibiotics to the suspension at $5^{\circ}C$ for long terms lasted for 997 days and 1, 256 days (presumed values) at maximum, respectively. The spores suspended with distilled water at $28^{\circ}C$ for long terms survived 152.4 days, but the spores suspended with Tyrode's solution at $-70^{\circ}C$ for long terms showed almost the same viable pattern as early freezing stages up to 780 days. The spores suspended with Tyrode's solution, frozen at $-70^{\circ}C$ and thawed at $5^{\circ}C$, showed the highest rate of extrusion of the polar filament. In the case of frozen spores, the extrusion rates during heating tend to become higher in accordance with the increase of frozen period, and the critical points of 180 day-frozen spores to be killed were generally 78.5 hr. at $60^{\circ}C$, 23.4 hr. at $70^{\circ}C$, 189.1 min. at $80^{\circ}C$ or 10.5 min. at $90^{\circ}C$. The longer the spores were frozen, the more time was needed for the death of spores after thawing; 20 days-17.4 days, 100 days-33.2 days, and 400 days-37.8 days. The longer the spores were frozen, the more time was needed for the death of spores at a conventional when they were dried air drying condition, 540 days-23.5 days, 160 days-21.0 days, and 20 days-14.4 days. On the other hand, the longer the spores were frozen, the more spores were dead rapidly when they were irradiated with 10W UV-ray; 100 days-26.0 hr, 300 days-21.9 hr, and 540 days-13.9 hr. The time needed for killing 200 days-frozen spores by various disinfectants at 1, 000 ppd was 5.2 min. by calcium oxide, 10.4 min. by potassium permanganate, 27.8 min. by malachite green and 14.3 hr. by formalin. Transient inhibitory effects of the extrusion of the polar filament were observed by various antiprotozoal and antifungal agents in the descending order of ketoconazole. metronidasole and dapsone. The above results presume that full drying, followed by spraying CaO and maintaining sunny condition for a few days on the concrete bottoms of knish farm may be an effective method for the prevention of intestinal giant.cystic disease.

• Korean Journal of Ecology and Environment
• /
• v.37 no.1 s.106
• /
• pp.36-46
• /
• 2004

We evaluated the effect of limiting nutrients and N/P ratio on the growth of phytoplankton in a small eutrophic reservoir from November 2002 to December 2003. Nutrient limitation was investigated seasonally using nutrient enrichment bioassay (NEB). DIN/DTP and TN/TP ratio (by weight) of the reservoir during the study period ranged 17${\sim}$187 and 13${\sim}$60, respectively. Most of nitrogen in the reservoir account for $NO_3$-N, but sharp increase of ammonia was evident during the spring season. Seasonal variation of dissolved inorganic phosphorus concentration was relatively small. DTP ranged 26.5${\sim}$10.1 ${\mu}g\;P\;L^{-1}$, and the highest and lowest concentration was observed in August and December, respectively. Chlorophyll a concentration ranged 28.8${\sim}$109.7 ${\mu}g\;L^{-1}$, and its temporal variation was similar to that of cell density of phytoplankton. Dominant phytoplankton species were Bacillariphyceae (Melosira varians) and Chlorophyceae (Dictyosphaerium puchellum) in Spring (March${\sim}$April). Cyanophyceae, such as Osillatoria spp., Microcystis spp., Aphanizomenon sp. dominated from May to the freezing time. TN/TP ratio ranged from 46 to 13 (Avg. 27${\pm}$6) from June to December when cyanobacteria (Microcystis spp.) dominated. p limitation for algal growth measured in all NEB experiments (17cases), while N limitation occurred in 8 out of 17 cases. The growth rates of phytoplankton slightly increased with decreasing of DIN/DTP ratio. Evident increase was observed in the N/P ratio of > 30, and it was sustained with DTP increase until 50 ${\mu}g\;P\;L^{-1}$. Under the same N/P mass ratio with the different N concentrations (0.07, 0.7and 3.5 mg N $L^{-1}$), Microcystis spp. showed the highest growth rate in the N/P ratio of< 1 with nitrogen concentration of 3.5 mg N $L^{-1}$). The responses of phytoplankton growth to phosphate addition were clearly greater with increase of N concentration. These results indicate that the higher nitrogen concentration in the water likely induce the stronger P-limitation on the phytoplankton growth, while nitrogen deficiency is not likely the case of nutrient limitation.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.41 no.7
• /
• pp.994-1001
• /
• 2012

Quality changes of pre-processed garlic, peeled and chopped, were analyzed during storage at $-18^{\circ}C$ or $2^{\circ}C$ for 30 days and at $20^{\circ}C$ for 3 days only for chopped garlic. As storage time increased, Hunter L values decreased and a, b values increased, indicating browning regardless of the pre-process type and storage temperature. Decay and sprouting rates of peeled garlic during storage at $2^{\circ}C$ significantly increased while those of peeled garlic were maintained during storage at $-18^{\circ}C$. Weight loss of peeled garlic during storage was greater at $-18^{\circ}C$ than at $2^{\circ}C$. Hardness of peeled garlic rapidly decreased by half from 1.04 kg to 0.58 kg by freezing, and it did not significantly change during the storage period. Viable numbers of total aerobic bacteria of peeled and chopped garlic did not significantly change during the storage period at $2^{\circ}C$ but were reduced at $-18^{\circ}C$. Total aerobic bacterial count of chopped garlic stored at $20^{\circ}C$ slightly increased during the storage period. Pyruvic acid content of chopped garlic was almost 2.5 times higher than that of peeled garlic at the initial stage (463.87 ${\mu}mol/g$ and 190.52 ${\mu}mol/g$, respectively). As storage time increased, pyruvic acid content of peeled garlic increased while that of chopped garlic decreased. These results indicate that pre-process type and storage temperature affected the quality changes of garlic during storage.

• Korean Journal of Animal Reproduction
• /
• v.24 no.1
• /
• pp.97-108
• /
• 2000

The objectives of this study were performed to increase the efficiency of the culture conditions of embryos produced in vitro, and to assess the developmental potential after transfer of those embryos into recipients. The mean number of folliclular oocytes recovered from an ovary was 10.7. The rates of maturation and fertilization in Grade I oocytes were significantly (P＜0.05) higher than Graden and III. Developmental rate into blastocyst in the culture group of TCM-199 with BOEC were significantly higher (P＜0.05) than the groups of TCM-199 and conditioned medium (24.7% vs. 12.4% and 18.2%). The survivability of post-thawed blastocysts equilibrated for 3 min in EFS solution was significantly (P＜0.05) lower than l0 for 1 and 2 min (32.1% vs. 82.9% and 73.3%). Significantly higher (P＜0.05) survival rate in blastocysts was seen after freezing than in morulae stage embryos. Out of all 105 recipients, 49 (46.7%) were confirmed in pregnant. On pregnancy of cattle, 48 calves were born from 40 recipients. The ratio of twin and single calves was 30.5% (32/40 and 7.6% (8/40), respectively. However, the others composed of abnormal, as judging as 6 (12.2%) for abortion and 3 (6.1%) for stillbirth during the pregnant period.

• Korean Journal of Animal Reproduction
• /
• v.25 no.1
• /
• pp.1-7
• /
• 2001

This study was to test whether in vitro matured Hanwoo oocytes can be successfully cryopreserved by a new vitrification procedure using MVC method. For the vitrification, oocytes were pretreated in 10% ethylene glycol (EG10) for 5~10 min, exposed in EG30 for 30 sec, each oocyte was individually put on the inner wall of 0.25 $m\ell$ straw, and then straws were directly plunged into L$N_2$. Thawing was taken by 4-step procedures 〔1.0 M sucrose (MS), 0.5 MS, 0.25 MS, and 0.125 MS〕 at 37$^{\circ}C$. In vitro developmental capacity (survival, cleavage ($\geq$2-cell) and blastocyst rates) in vitrified group was no significant difference compared to that in other treatment groups (exposed; 100.0, 74.4, 32.3% and control; 100.0, 78.3, 36.3%): high mean percentage of oocytes (91.2%) was survived, 69.4% of them were cleaved and 27.9% of cleaved embryos were developed to blastocyst. Especially, after transfer of in vitro developed embryos in vitrified group, four of six recipient animals were pregnant and three of them were ongoing-pregnant by manual palpation at 250 days after transfer. This result demonstrates that MVC method is very appropriate freezing method for the Hanwoo in vitro matured oocytes and that ovum bank can be maintained efficiently by MVC cryopreservation method.

• Korean Journal of Animal Reproduction
• /
• v.23 no.2
• /
• pp.127-132
• /
• 1999

This study was carried out to investigate the general characteristics such as volume, sperm concentration, sperm motility, sperm abnormality on whole semen, removed seminal plasma(RSP) semen and fractional semen of small dogs, and the effect of temperature and preservatio time and cryopreservation on motility of whole and removed seminal plasma semen. Multiple ejaculates were collected from small dogs by the digital manipulation of penis. 1. Average sperm concentration, sperm motility and abnormal sperm rates of whole semen and RSP semen were 5.07$\pm$2.32$\times$10$^{6}$ cells/$m\ell$, 95.42$\pm$2.65%, 4.42$\pm$0.157% and 4.69$\pm$3.27~4.25$\pm$3.65$\times$10$^{6}$ cells/$m\ell$, 91.17$\pm$3.85~88.52$\pm$3.85%, 6.57$\pm$0.43~5.54$\pm$0.52%, respectively. 2. Average semen volume per ejaculate, sperm concentration, sperm motility and abnormal sperm rate of 1st fractional semen were 0.92$\pm$0.7$m\ell$, 4.57$\pm$0.78$\times$10$^{6}$ cells/$m\ell$, 10.72$\pm$3.21% and 5.50$\pm$0.70%. Average semen volume per ejaculate, sperm concentration, sperm motility and abnormal sperm rate of 2nd fractional semen were 2. 14$\pm$0.19$m\ell$, 2.01$\pm$0.12$\times$10$^{6}$ cells/$m\ell$, 95.44$\pm$4.21% and 4.31$\pm$0.53%. Average semen volume per ejaculate, sperm concentration, sperm motility and abnormal sperm rate of 3rd fractional semen were 2.66$\pm$0.23$m\ell$, 2.35$\pm$0.21$\times$10$^{6}$ cells/$m\ell$, 90.71$\pm$2.63%, 6.33$\pm$0.91%, respectively. 3. Motility of whole semen and RSP semen were higher at 2$0^{\circ}C$ than at 4$^{\circ}C$ or 37$^{\circ}C$. When preservation temperature was 2$0^{\circ}C$, sperm motility were 98.32% at 1 hr, 92.15% at 5 hrs, 90.23% at 10 hrs 82.08% at 15 hrs 70.07% at 20 hrs 60.02% at 20 hrs 37.19% at 40 hrs respectively. 4. Average sperm motility of frozen 2nd fraction semen and RSP semen were 33.3$\pm$8.7, 54.7$\pm$9.5%, respectively. Sperm motility was significantly higher in frozen 2nd fraction semen and RSP semen compared with control group.