• KSBB Journal
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• 제15권5호
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• pp.438-443
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• 2000

맥주의 숙성기간을 단축시키기 위해 네 종류의 고정화용 담체를 활용한 고정화 효모 반응기를 시험하였다. 전발효가 끝난 Green Beer (GB)를 효모를 제거하고 열처리할 경우 처 리온도가 높을수록 전구체인 $\alpha$-acetolactate의 diacety 1로의 전 환율이 낮고 전환속도도 빨라 $70^{\circ}C$이상의 온도에서는 4분이면 충분하였다 GB 중의 산소농도는 전구체에서 DA로의 변환율에 매우 큰 영향을 끼쳤는데 그 농도가 낮을수록 전환율이 낮았으며 0.1 ppm 이하의 농도에서는 거의 대부분의 전 구체가 DA이외의 물칠로 전환되었다. 열처리한 전발효 맥주 를 HAN, G-2, FLO, GDC 담체 column에 $\alpha$, 체류시간 80-150분으로 통과시킬 경우 diacety I 농도를 상품 맥주의 품질로 적합한 0.1 ppm이하로 떨어뜨릴 수 있어서 시험한 담체 모두 맥주 숙성용 효모고정화용 답체로의 실용화 가능 성이 있었다. 이 때 세라믹 담체 column의 경우 GDC 담체 column에 비해 미발효된 잔당의 발효에 의한 새로운 DA 전 구체의 생성이 많았다. 위와 같은 방법으로 생산한 맥주를 공장에서 기존의 방법으로 생산한 맥주와 맛을 비교한 결과 미세한 차이가 있으나 불쾌한 느낌을 주는 이취등은 발견되지 않아 고정화 효모 반응기에 의한 고속 숙성공정의 현장적 용 가능성이 높음을 확인할 수 있었다.

• Clinical and Experimental Reproductive Medicine
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• 제19권1호
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• pp.15-29
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• 1992

In the present study, it was aimed to find the role of calcium on the maturation of mouse follicular oocytes as well as for the role of calcium inhibitors, $Ni^{2+}$ and $La^{3+}$. Mouse follicular oocytes were cultivated in different media at $37^{\circ}C$, in 100% humidified $CO_2$ incubator for 3 and 17 hrs. The results were as follows; 1. There was no differences in GVBD between the control and experimental groups during the 3 hr culture. 2. Mouse oocytes were matured to higher rate in MHBS rather than HTF for 17 hr culture. 3. Maturation rate was significantly lower in $Ca^{2+}$-free and $Ca^{2+}$ 0.4 mM which were tested, compared to other calcium concentration used in the present study. 4. Calcium inhibitor, $Ni^{2+}$, it showed highest degeneration rate at all calcium concentrations and additionally in $Ni^{2+}$ $100{\mu}M$ treated group next. Maturation rate was significantly decrease as the $Ca^{2+}$ inhibitor concentration increased. 5. In all Lanthanum treated groups of calcium-free, degeneration were significantly high treated groups at 0.4 mM $Ca^{2+}$ concentrations degeneration rates of all group were significantly lower than that of the control but maturation rates were not significantly different in any group. In lanthanum $100{\mu}M$ treated group at 0.4 mM and 0.8 mM calcium concentration, its maturation rate was significantly higher than that of the control. Maturation rates of all groups of lanthanum treated at 1.71 mM calcium concentration were not significantly different among groups. 6. In the calcium treated group (0.4mM-1.7 mM), the presence of phosphate does not seem to be needed for oocyte maturation. However, the presence of phosphate at $Ca^{2+}$ 0.8 mM only seems to stimulated maturation.

• Asian-Australasian Journal of Animal Sciences
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• 제14권5호
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• pp.693-696
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• 2001

The effect of replacement of in vitro maturation medium completely with the buffalo follicular fluid (buFF) on in vitro oocyte maturation of buffalo oocytes was studied. 5 to 8 buffalo cumulus oocyte complexes were cultured in a single drop with each of the eight media studied i.e., M199+steer serum (10% v/v), M199+steer serum (10% v/v)+PMSG, M199+buFF (10% v/v), M199+buFF (10% v/v)+PMSG, M199+buFF (50% v/v), M199+buFF (50% v/v)+ PMSG, buFF (100%) and buFF+PMSG at $39^{\circ}C$ and 5% $CO_2$ in air for 24 h. Supplementation of M199 with Steer serum alone resulted in IVM rate of 35% only. When the above medium was supplemented with PMSG, the maturation rate rallied to 82%. Significant increase in the maturation rates were observed when M199 was supplemented with increasing levels of buFF. A further increase in the maturation rate was also obtained when PMSG was incorporated into the medium of M199 supplemented with buFF. The rate of maturation was to the tune of 91% when oocytes were matured in buFF alone which was increased non significantly on the addition of PMSG. Highest maturation rate (97%) obtained with M199+buFF (50%v/v)+PMSG did not differ significantly from that obtained by either M199+buFF (10%v/v)+PMSG or buFF+PMSG. It is suggested that buFF alone without any supplementation can form the effective in vitro maturation medium for buffalo oocytes.

• Clinical and Experimental Reproductive Medicine
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• 제21권2호
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• pp.191-200
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• 1994

The influence of hypoxanthine and ovarian steroids on the meiotic maturation process of mouse oocytes was investigated for the qualified application of culture medium in in vitro fertilization(IVF). Mouse oocytes were cultured in hypoxanthine and various ovarian steroids(progesterone, estradiol-17${\beta}$ and testosterone) and their effects on the oocyte maturation had been observed. When mouse oocytes were cultured in the various concentration(1-4mM) of hypoxanthine, meiotic maturation of cumulus cell-enclosed oocytes was inhibited by presence itself, which was a dose-dependent effect in meiotic arrest of mouse oocytes. The presence of progesterone, estradiol-17${\beta}$ and testosterone have made the mouse oocyte mature properly. Meanwhile maturation of cumulus cell-enclosed oocyte was severely inhibited by 3 hoursculture in the media of progesterone supplemented with hypoxanthine. However the continuous presence lasting 24 hours of progesterone even supplemented with hypoxanthine had got rid of the inhibition of oocytes maturation. Not only estradiol-17${\beta}$ supplemented with hypoxanthine but also testosterone supplemented with hypoxanthine exert the severe inhibition of the maturation of cumulus cell-enclosed oocytes for 3-hours culture. However the continuous presence lasting 24 hours of estradiol-17${\beta}$ and testosterone even supplemented with· hypoxanthine had relieved the inhibition of oocytes maturation. These results make us suggest that hypoxanthine inhibits the mouse oocyte maturation, particularly markedly in conjunction with ovarian steroids for short period, which indicated some sort of the synergistic inhibitory retationship between the ovarian steroids and hypoxanthine.

• 한국수정란이식학회지
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• 제10권2호
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• pp.115-120
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• 1995

This study was carried out to investigate On in vitro fertilization, survival rate and developmental rate of rapidly frozen bovine immature oocytes. Immature oocytes cultured for 1, 12, 24, 48 hours in 20% FCS + TCM-199 medium and thereafter rapidly freezing-thawed oocytes inseminated with capacitated sperm. The immature oocytes following dehydration by 1.5M DMSO + 2.0M glycerol + 0.25M sucrose + TCM 199 media + 20% FGS were directly plunged into liquid nitrogen and thawes in 3$0^{\circ}C$ water. Rapid freezing embryos co-cultured in 20% FCS + TCM-199 media containing hormones(21U/mL PMSG, 21U /mL hGG and 1 $\mu$g /mL 17$\beta$-estradiol) and cumulus cells(1 x 105-6 cells). Survival rate was defined as development rate on in vitro culture or FDA-test. The results are summarized as follows ; 1. The in vitro maturation and fertilization rate of immature bovine oocytes on in vitro maturation period(1, 12, 24, 48 hrs) before rapid freezing4hawed were 57.1%, 45.7%, 37.1%, 25.7% and 40.0%, 31.4%, 20.0%, 11.4%, respectively. 2. The survival rate of immature bovine oocytes on in vitro maturation period(1, 12, 24, 48 hrs) before rapid freezing-thawed were 33.3%, 26.7%, 20.0%, and 10.0%, respectively. The survival rate of rapid freezing4hawed immature oocytes was significantly lower than that of non-freezing oocytes. 3. The survival rate of rapid freezing4hawed excellent and good bovine embryos co-cultured in 20% FCS + TCM-199 media containing hormones(PMSG, hCG, 17$\beta$-estradiol) and cumulus cells 4 to 5 hrs and 20 to 24 hrs were 35.0%, 15.0% and 25.0%, 15.0% and 40.0%, 20.0% and 30.0%, 15.0%, respectively. The survival rate of embryos co-cultured in TCM-199 media containing hormones and cumulus cells was significantly higher than that of non co-culture.

• 한국가축번식학회지
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• 제12권2호
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• pp.120-124
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• 1988

Rat oocyte-cumulus complexes were cultured in verious media in order to induce maturation division in vitro. When the complexes were cultured in mKRB containing estrous rat serum (ERS) and FCS of 5% instead of BSA higher proportions(83.3 and 86.7%) of oocytes matured to metaphase II in 20h compared to control(75%) and t도 maturation rates in mKRB plus FCS were generally higher than those in mKRB plus ERS. Fertilization and early cleavage rates in vitro of the intact oocytes matured in mKRB containing BSA and FCS were generally higher than those of cumulus-removed oocytes and these rates were higher in mKRB containing 5% FCS than those in mKRB containing BSA. These results indicate that maturation rate in vitro was greatly increased by the addition of FCS instead of BSA to mKRB solution and the presence of cumulus cells around oocytes prior to sperm insemintion may be responsible for the increase of in vitro fertilization and early cleavage rates.

• 한국가축번식학회지
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• 제15권3호
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• pp.221-224
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• 1991

This study was undertaken to evaluate the effect of rabbit peritioneal fluid(rPF) on in vitro maturtion of porcine follicular oocytes. From does 20h after hCG injection, rPF was aspirated aseptically at laparatomy, and then centrifuged, filtrated, and preincubated immediately for 12h. Porcine follicular oocytes isolated from ovaries of slaughtered animals were incubated in TCM-HEPES＋10% FCS, TCM-HEPES＋rPF(v/v, 50/50), or rPE only and examined the nuclear maturation after aceto-orcein or hochest staining. After identifying the optimal incubation time, this experiment was repeated for 5 times. Under the TCM-HEPES containing hormones and serum codition, the time range of porcine follicular oocyte maturation was 38 to 44 hours and the optimal time of maturation of follicular oocyte in vitro was 42 hour cultivation, respectively. The maturatin rates(89.4% and 92.7%) of porcine follicular oocytes cultured in the media with 50% rPF or only rPF were signifciantly higher thanthat (84.6%) of oocytes cultured with TCM-HEPES, respectively. These results suggest that the unknown components(s) of rPF promoted in vitro maturation of porcine follicular oocytes.

• 한국발생생물학회지:발생과생식
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• 제25권2호
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• pp.75-82
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• 2021

We studied steroid metabolites produced from red-spotted grouper ovarian follicles during maturation. Oocytes with 350-500 ㎛ diameter were in vitro incubated in the presence of [³H] 17α-hydroxyprogesterone as a precursor. Steroid metabolites were extracted from incubated media and oocytes. The extracts were separated and identified using thin layer chromatography, high performance liquid chromatography and gas chromatography-mass spectrometry. The identified metabolites were androstenedione (A₄), testosterone (T) and estrone (E₁). The metabolites of A₄ was dominant in all size of oocytes and it was the highest in 480 ㎛ diameter oocytes. The metabolites of two progestins, 17α,20β-dihydroxy-4-pregnen-3-one and 17α,20α-dihydroxy-4-pregnen-3-one were detected in the oocytes less than 480 ㎛ diameter although they were not identified definitely. In the oocytes of 480 ㎛ diameter, metabolite of progestin was the highest, and germinal vesicle (GV) was still in the middle of cytoplasm. In the oocytes of 500 ㎛ diameter, GV was began to migrate and the major metabolites were A₄ and E₁. The metabolite of E₁ was detected in all size of oocytes and it was higher than that of E₂. These results suggest that oocytes of 480 ㎛ diameter are the transitional stage involving steroidogenic shift to final oocyte maturation and potential function of E₁ during maturation process.

• 한국가축번식학회지
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• 제21권2호
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• pp.123-130
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• 1997

Normal maturation of the mammalian oocytes is prerequisite for the fertilization and the early embryonic development. We have been tested the effects of purine and its de novo synthetic inhibitor, azaserine(Aza) on the maturation of germinal vesicle(GV) and germinal vesicle breakdown(GVBD) mouse oocytes. Denude-immature oocytes were cultivated in the media containing adenosine, guanosine, and/or azaserine, and checked the matruation stage by monitoring the prominent morphological changes. In GV stage oocytes, GV was arrested temporarily by the adenosine(1.0%) and protractedly by the guanosine(65.9%, P<0.001). The regression was increased significantly at the adenosine(90%, P<0.001) but decreased at the guanosine(1.6%, P<0.05). Inhibiting the de novo synthesis of purine, nuclear maturation rate was increase(90.4% : 96.7%), but GV arrest was significantly increased by cotreatment with guanosine(P<0.001). Polar body extraction significantly was increased at the Aza(P<0.05), but not in others. In GVBD oocytes, adenosine itself did not affect GVBD arrest. Guanosine, on the other hand, elevated GVBD arrest rate(P<0.001), but co-treated with Aza, decreased GVBD arrest(P<0.001). Aza increased GVBD arrest rate(20.2%, P<0.05) compared with control. From those results, we know that guanosine shows more prominent effect on the inhibition of nuclear maturation at the GV stage, and of the 1st polar body extrusion at the GVBD stage. Adenosine showed the cytoplasmic toxicity at GV stage oocyte. Our data speculate that cytoplasmic cAMP level is auto-regulated by endogenous adenylate cyclase while GVBD is inhibited by guanosine, since purine toxicity is not observed in the GVBD stage. And it is showed that purine metabolism is concerned with nuclear maturation, that the amounts of purine metabolism is not even during the oocyte maturation.

• Asian-Australasian Journal of Animal Sciences
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• 제8권2호
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• pp.123-127
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• 1995

In vitro embryo production (IVP) is affected by various factors during in vitro maturation, fertilization, and development. In this experiment, the effect of ovary type, quality of follicular oocyte, medium used for fertilization, presence of hormone in medium, sperm concentration on in vitro maturation and fertilization were examined for effective IVP. In vitro maturation was carried out using TCM-199 supplemented with 15% FCS and hormones in 5% $CO_2$ incubator for 24h. In vitro fertilization was performed with frozen-thawed sperm in modified mTALP medium containing 0.3% BSA, $10{\mu}g/ml$ heparin, and 5mM/ml caffeine for 24h. The fertilized embryos were co-cultured on monolayer of cumulus cells in TCM-199. When oocytes were collected from functionally active and inactive ovaries, maturation rate was 76.9 and 7.7%, respectively. When oocytes were classified morphologically to good and poor grades, maturation rate was 75 and 58.8%, respectively. FSH + LH + $E_2$ (86.4%) showed higher maturation rate than control (53.0%) and FSH (73%). The fertilization rate was 28.2, 100 and 91.7% in $1.6{\times}10^5$, $5.0{\times}10^5$ and $10.0{\times}10^5$ sperm concentration per ml. When oocytes were fertilized in mTALP and BO media, fertilization and cleavage rates of oocytes in mTALP were higher (84.3 and 56.9%) than those (67.4 and 23.3%) in BO medium. In this experiment, in vitro maturation, fertilization and development of oocytes were affected by type of ovary, grade of oocyte, hormones, sperm concentration and fertilization medium.