호밀 사일리지 제조 시 팽이버섯 수확 후 배지 첨가수준이 in vitro 반추위 발효특성 및 건물소화율에 미치는 영향

강한별,조웅기,조수정,이신자,이성실,문여황 경상대학교 농업생명과학연구원 2017 농업생명과학연구 Vol.51 No.5

본 연구는 팽이버섯 수확 후 배지를 첨가하여 호밀 사일리지 제조 시 팽이버섯 수확 후 배지를 에너 지원으로서 사용하기 위하여 in vitro 반추위 발효실험이 수행되었다. 공시 사일리지는 출수기의 호밀 에 팽이버섯 수확 후 배지 첨가비율(0%, 20%, 40%, 60%)에 따라 제조하여 6주일간 발효시켰다. In vitro 배양액 제조를 위한 반추위액은 농후사료와 볏짚을 40:60의 비율로 급여한 반추위 cannula가 시 술된 Holstein 수소 2두로 부터 채취하였다. In vitro 실험은 발효시간대를 3, 6, 9, 12, 24 및 48시간 으로 설정하고, 각 처리구별로 3반복으로 발효특성과 건물소화율을 측정하였다. In vitro 배양액의 pH 는 발효시간이 길어짐에 따라 낮아지는 경향이었으며, 48시간 경과 시에는 버섯수확 후 배지 60% 첨가 구가 타 처리구에 비해 유의적(p<0.05)으로 낮았다. 미생물 성장율은 배양시간이 경과함에 따라 증가 하는 경향이었으며, 발효 48시간 경과 시에는 버섯수확 후 배지 20% 첨가구가 타 처리구에 비해 유의 적으로(p<0.05) 높았다. Gas발생량은 48시간 발효 시에 대조구가 타 처리구에 비해 유의적(p<0.05)으 로 높았다. 건물소화율은 버섯수확 후 배지의 첨가비율이 높을수록 높았는데, 발효 24시간 및 48시간에 는 R-60구가 처리구 중에서 가장 높았으며(p<0.05), 대조구에서는 전 발효기간 동안 건물소화율이 현 저히 낮은(p<0.05) 상태에 있었다. In vitro 반추위내 발효실험의 결과와 버섯수확 후 배지의 활용성을 고려할 때, 호밀사일리지 제조 시 팽이버섯 수확 후 배지 첨가비율을 원물기준으로 60%수준이 가장 긍 정적인 것으로 나타났다. 향후 대사시험이나 사양시험을 통하여 가축사료로써의 최적 대체 비율을 규명 해야 할 것으로 사료되는 바다. The in vitro experiment was conducted to ensure the supplemental level of spent Flammulina velutipes mushroom substrates(SMS) as an energy source in manufacturing of rye silage. Rye harvested at heading stage was ensiled with spent mushroom substrates of 0%(Control), 20%(R-20), 40%(R-40) and 60%(R-60) as fresh matter basis for 6week. The rumen fluid for preparation of in vitro solution was collected from two cannulated Holstein bulls fed a 40:60 concentrate:timothy diet. The experiment was conducted by 3, 6, 9, 12, 24, and 48 hrs of ncubation time with 3 replications. The silages were evaluated fermentation characteristics and dry matter digestibility(DMD) in vitro. The pH of in vitro solution was inclined to decrease with elapsing the incubation time, and that of the R-60 was significantly(p<0.05) lower than the other treatment at 48 hr of incubation. The microbial growth in vitro was inclined to increase with elapsing the incubation time, and that of the R-20 was significantly(p<0.05) greater than the Control at 48 hr of incubation. Gas production was greater(p<0.05) in the Control than the other treatments at 48 hr of incubation. In vitro dry matter digestibility(IVDMD) was higher with increasing the supplemental level of SMS, and was significantly(p<0.05) lower in the Control compared with other treatments throughout whole incubation time. The IVDMD for R-60 was the highest(p<0.05) among treatments at 24 hr and 48 hr of incubation. Considering of above results and the availability of SMS, SMS could be supplemented by 60% in fresh matter basis for rye silage fermentation.

In vitro ruminal fermentation of fenugreek (Trigonella foenum-graecum L.) produced less methane than that of alfalfa (Medicago sativa)

Niu Huaxin,Xu Zhongjun,Yang Hee Eun,McAllister Tim A,Acharya Surya,Wang Yuxi 아세아·태평양축산학회 2021 Animal Bioscience Vol.34 No.4

Objective: The objective of this study was to compare fenugreek (FG) with alfalfa (Alf) in ruminal fermentation and methane (CH4) production in vitro. Methods: Whole-plant FG harvested at 11- and 15-wk and Alf harvested at early and mid-bloom maturities, alone or as 50:50 mixture of FG and Alf at the respective maturity, were assessed in a series of 48-h in vitro batch culture incubations. Total fermentation gas and methane gas production, dry matter (DM) disappearance, volatile fatty acids, microbial protein and 16S RNA gene copy numbers of total bacteria and methanogens were determined. Results: Compared to early bloom Alf, FG harvested at 11-wk exhibited higher (p<0.05) in vitro DM and neutral detergent fibre disappearance, but this difference was not observed between the mid-bloom Alf and 15-wk FG. Regardless plant maturity, in vitro ruminal fermentation of FG produced less (p<0.001) CH4 either on DM incubated or on DM disappeared basis than that of Alf during 48-h incubation. In vitro ruminal fermentation of FG yielded similar amount of total volatile fatty acids with higher (p<0.05) propionate percentage as compared to fermentation of Alf irrespective of plant maturity. Microbial protein synthesis was greater (p<0.001) with 11-wk FG than early bloom Alf as substrate and 16S RNA gene copies of total bacteria was higher (p<0.01) with 15-wk FG than mid-bloom Alf as substrate. Compared to mid-bloom Alf, 15-wk FG had lower (p<0.05 to 0.001) amount of 16S RNA methanogen gene copies in the whole culture during 48-h incubation. Conclusion: In comparison to Alf, FG emerges as a high quality forage that can not only improve rumen fermentation in vitro, but can also remarkably mitigate CH4 emissions likely due to being rich in saponins. Objective: The objective of this study was to compare fenugreek (FG) with alfalfa (Alf) in ruminal fermentation and methane (CH₄) production <i>in vitro</i>.Methods: Whole-plant FG harvested at 11- and 15-wk and Alf harvested at early and mid-bloom maturities, alone or as 50:50 mixture of FG and Alf at the respective maturity, were assessed in a series of 48-h <i>in vitro</i> batch culture incubations. Total fermentation gas and methane gas production, dry matter (DM) disappearance, volatile fatty acids, microbial protein and 16S RNA gene copy numbers of total bacteria and methanogens were determined.Results: Compared to early bloom Alf, FG harvested at 11-wk exhibited higher (p<0.05) <i>in vitro</i> DM and neutral detergent fibre disappearance, but this difference was not observed between the mid-bloom Alf and 15-wk FG. Regardless plant maturity, <i>in vitro</i> ruminal fermentation of FG produced less (p<0.001) CH₄ either on DM incubated or on DM disappeared basis than that of Alf during 48-h incubation. <i>In vitro</i> ruminal fermentation of FG yielded similar amount of total volatile fatty acids with higher (p<0.05) propionate percentage as compared to fermentation of Alf irrespective of plant maturity. Microbial protein synthesis was greater (p<0.001) with 11-wk FG than early bloom Alf as substrate and 16S RNA gene copies of total bacteria was higher (p<0.01) with 15-wk FG than mid-bloom Alf as substrate. Compared to mid-bloom Alf, 15-wk FG had lower (p<0.05 to 0.001) amount of 16S RNA methanogen gene copies in the whole culture during 48-h incubation.Conclusion: In comparison to Alf, FG emerges as a high quality forage that can not only improve rumen fermentation <i>in vitro</i>, but can also remarkably mitigate CH₄ emissions likely due to being rich in saponins.

Development and pregnancy rates of Camelus dromedarius-cloned embryos derived from in vivo- and in vitro-matured oocytes

Son Young-Bum,Jeong Yeon Ik,Jeong Yeon Woo,Olsson Per Olof,Hossein Mohammad Shamim,Cai Lian,Kim Sun,Choi Eun Ji,Sakaguchi Kenichiro,Tinson Alex,Singh Kuhad Kuldip,Rajesh Singh,Noura Al Shamsi,Hwang Wo 아세아·태평양축산학회 2022 Animal Bioscience Vol.35 No.2

Objective: The present study evaluated the efficiency of embryo development and pregnancy of somatic cell nuclear transfer (SCNT) embryos using different source-matured oocytes in Camelus dromedarius. Methods: Camelus dromedarius embryos were produced by SCNT using in vivo- and in vitro- matured oocytes. In vitro embryo developmental capacity of reconstructed embryos was evaluated. To confirm the efficiency of pregnancy and live birth rates, a total of 72 blastocysts using in vitro- matured oocytes transferred into 45 surrogates and 95 blastocysts using in vivo- matured oocytes were transferred into 62 surrogates by transvaginal method. Results: The collected oocytes derived from ovum pick up showed higher maturation potential into metaphase II oocytes than oocytes from the slaughterhouse. The competence of cleavage, and blastocyst were also significantly higher in in vivo- matured oocytes than in vitro- matured oocytes. After embryo transfer, 11 pregnant and 10 live births were confirmed in in vivo- matured oocytes group, and 2 pregnant and 1 live birth were confirmed in in vitro- matured oocytes group. Furthermore, blastocysts produced by in vivo-matured oocytes resulted in significantly higher early pregnancy and live birth rates than in vitromatured oocytes. Conclusion: In this study, SCNT embryos using in vivo- and in vitro-matured camel oocytes were successfully developed, and pregnancy was established in recipient camels. We also confirmed that in vivo-matured oocytes improved the development of embryos and the pregnancy capacity using the blastocyst embryo transfer method. Objective: The present study evaluated the efficiency of embryo development and pregnancy of somatic cell nuclear transfer (SCNT) embryos using different source-matured oocytes in Camelus dromedarius.Methods: Camelus dromedarius embryos were produced by SCNT using in vivo- and in vitro- matured oocytes. In vitro embryo developmental capacity of reconstructed embryos was evaluated. To confirm the efficiency of pregnancy and live birth rates, a total of 72 blastocysts using in vitro- matured oocytes transferred into 45 surrogates and 95 blastocysts using in vivo- matured oocytes were transferred into 62 surrogates by transvaginal method.Results: The collected oocytes derived from ovum pick up showed higher maturation potential into metaphase II oocytes than oocytes from the slaughterhouse. The competence of cleavage, and blastocyst were also significantly higher in in vivo- matured oocytes than in vitro- matured oocytes. After embryo transfer, 11 pregnant and 10 live births were confirmed in in vivo- matured oocytes group, and 2 pregnant and 1 live birth were confirmed in in vitro- matured oocytes group. Furthermore, blastocysts produced by in vivo-matured oocytes resulted in significantly higher early pregnancy and live birth rates than in vitromatured oocytes.Conclusion: In this study, SCNT embryos using in vivo- and in vitro-matured camel oocytes were successfully developed, and pregnancy was established in recipient camels. We also confirmed that in vivo-matured oocytes improved the development of embryos and the pregnancy capacity using the blastocyst embryo transfer method.

Acclimatization of in vitro Gymnaster koraiensis Using Paclobutrazol and Transplanting Medium

Seon Mi Kim,Chang Hee Lee,Ki Sun Kim 한국원예학회 2005 Horticulture, Environment, and Biotechnology Vol.46 No.4

An efficient acclimatization of micropropagated Gymnaster koraiensis was developed through paclobutrazol pretreatment in vitro. Microshoots were induced on the MS medium containing a combination of 0.2 ㎎ㆍL?¹ thidiazuron and 0.1 ㎎ㆍL?¹ α-naphthaleneacetic acid, followed by culture on MS media containing combinations of 0.0?2.0 ㎎ㆍL?¹ paclobutrazol (PBZ) and 0.0?0.1 ㎎ㆍL?¹ indole-3-butyric acid (IBA) for 4 weeks. PBZ pretreatments in vitro at 0.5?1.0 ㎎ㆍL?¹ seemed to be more effective on the number of primary roots and fresh weight of root regardless of IBA concentration. Thus, PBZ pretreatment in vitro at 0.0, 0.5, 1.0, or 2.0 ㎎ㆍL?¹, respectively, was evaluated for in vitro root development and for acclimatization of plantlets in a greenhouse. Concerning in vitro rooting after 6 weeks of culture, root diameter and chlorophyll contents were increased in a PBZ concentration-dependent manner, whereas leaf area, leaf length, leaf shape index, and shoot and root length were markedly decreased. As PBZ concentration increased, stomatal index increased in adaxial leaf surface but it kept constant in abaxial surface. The plantlets obtained from each PBZ pretreatment in vitro were acclimatized in three commercial media under greenhouse condition. Soil mixture (peatmoss:perlite = 7:3, v/v) was the most suitable transplanting medium for acclimatization of this species regardless of in vitro PBZ concentration. The plantlets pretreated with PBZ at 0.5 ㎎ㆍL?¹ during in vitro culture showed the heavier fresh weight and dry weight than any other pretreated with PBZ concentrations. As a result, leaf morphology such as leaf shape index, stomatal index, and number of leaves seemed not to be significantly changed in PBZ-pretreated plantlets compared with normal plantlets after 16 weeks of acclimatization period. In conclusion, in vitro PBZ pretreatment at 0.5 ㎎ㆍL?¹ enhanced in vitro rooting and in vivo acclimatization of micropropagated G. koraiensis.

The antioxidant icariin protects porcine oocytes from age-related damage in vitro

윤재욱,Lee Seung-Eun,Park Yun-Gwi,Kim Won-Jae,Park Hyo-Jin,Park Chan-Oh,김소희,Oh Seung-Hwan,Lee Do-Geon,Pyeon Da-Bin,김은영,박세필 아세아·태평양축산학회 2021 Animal Bioscience Vol.34 No.4

Objective: If fertilization does not occur within a specific period, the quality of unfertilized oocytes in the oviduct (in vivo aging) or in culture (in vitro aging) will deteriorate over time. Icariin (ICA), found in all species of Epimedium herbs, has strong antioxidant activity, and is thought to exert anti-aging effects in vitro. We asked whether ICA protects oocytes against age-related changes in vitro. Methods: We analyzed the reactive oxygen species (ROS) levels and expression of antioxidant, maternal, and estrogen receptor genes, and along with spindle morphology, and the developmental competence and quality of embryos in the presence and absence of ICA. Results: Treatment with 5 μM ICA (ICA-5) led to a significant reduction in ROS activity, but increased mRNA expression of glutathione and antioxidant genes (superoxide dismutase 1 [SOD1], SOD2, peroxiredoxin 5, and nuclear factor erythroid 2‐like 2), during aging in vitro. In addition, ICA-5 prevented defects in spindle formation and chromosomal alignment, and increased mRNA expression of cytoplasmic maturation factor genes (bone morphogenetic protein 15, cyclin B1, MOS proto‐oncogene, serine/threonine kinase, and growth differentiation factor‐9). It also prevented apoptosis, increased mRNA expression of antiapoptotic genes (BCL2-like 1 and baculoviral IAP repeat-containing 5), and reduced mRNA expression of pro-apoptotic genes (BCL2 antagonist/killer 1 and activation of caspase-3). Although the maturation and cleavage rates were similar in all groups, the total cell number per blastocyst and the percentage of apoptotic cells at the blastocyst stage were higher and lower, respectively, in the control and ICA-5 groups than in the aging group. Conclusion: ICA protects oocytes against damage during aging in vitro; therefore, it can be used to improve assisted reproductive technologies. Objective: If fertilization does not occur within a specific period, the quality of unfertilized oocytes in the oviduct (<i>in vivo</i> aging) or in culture (<i>in vitro</i> aging) will deteriorate over time. Icariin (ICA), found in all species of <i>Epimedium</i> herbs, has strong antioxidant activity, and is thought to exert anti-aging effects <i>in vitro</i>. We asked whether ICA protects oocytes against age-related changes <i>in vitro</i>.Methods: We analyzed the reactive oxygen species (ROS) levels and expression of antioxidant, maternal, and estrogen receptor genes, and along with spindle morphology, and the developmental competence and quality of embryos in the presence and absence of ICA.Results: Treatment with 5 μM ICA (ICA-5) led to a significant reduction in ROS activity, but increased mRNA expression of glutathione and antioxidant genes (superoxide dismutase 1 [<i>SOD1</i>], <i>SOD2</i>, peroxiredoxin 5, and nuclear factor erythroid 2‐like 2), during aging <i>in vitro</i>. In addition, ICA-5 prevented defects in spindle formation and chromosomal alignment, and increased mRNA expression of cytoplasmic maturation factor genes (bone morphogenetic protein 15, cyclin B1, MOS proto‐oncogene, serine/threonine kinase, and growth differentiation factor‐9). It also prevented apoptosis, increased mRNA expression of antiapoptotic genes (BCL2-like 1 and baculoviral IAP repeat-containing 5), and reduced mRNA expression of pro-apoptotic genes (BCL2 antagonist/killer 1 and activation of caspase-3). Although the maturation and cleavage rates were similar in all groups, the total cell number per blastocyst and the percentage of apoptotic cells at the blastocyst stage were higher and lower, respectively, in the control and ICA-5 groups than in the aging group.Conclusion: ICA protects oocytes against damage during aging <i>in vitro</i>; therefore, it can be used to improve assisted reproductive technologies.

사료 접종 방법에 의한 in vitro 반추위 발효 상성 변화

유대겸,문준범,김한빈,양성재,박중국,이세영,서자겸 강원대학교 동물자원공동연구소 2019 동물자원연구 Vol.30 No.3

The objective of this study was to investigate the effect of different feed inoculation method on rumen fermentation in an in vitro. Three experimental treatments were used: control (CON, direct dispersion of feed (2 g) in rumen fluid), combinations of direct dispersion (1 g) and nylon bag (DNB, pore size: 50 μm, 1 g), and nylon bag (NB, 2 g). An in vitro fermentation experiment was carried out using strained rumen fluid for 48 h incubation time and timothy was used as a substrate. At the end of the incubation, in vitro dry matter digestibility (IVDMD), in vitro neutral detergent fiber digestibility (IVNDFD), pH, volatile fatty acids (VFA), ammonia nitrogen (NH3-N), and microbial community were evaluated and gas production was estimated at 3, 6, 12, 24, 48 h incubation periods. Gas production was higher in CON than DNB and NB at 6 and 12 h incubation time (p<0.01). There were no differences in final gas production, pH, NH3-N concentration, total VFA production, and VFA profiles among treatments. The IVDMD was lowest in CON (p<0.01) but the IVNDFD was not differed by feed distribution methods. There were no significant differences in general bacteria and fungi. Protozoa count was highest in NB treatment among treatments (p<0.01). The abundance of cellulolytic bacteria, Ruminococcus flavefaciens and Fibrobacter succinogenes, was highest in the CON among treatments (p<0.01). 생물학적 평가 방법 중 in situ와 in vitro 평가 방법은 비용 및 시간을 절약할 수 있으며 동물 복지 측면에서 제약이 적어 주목되고 있는 방법이다. In vitro 실험에서 사용되는 사료 접종법으로 배양병에 직접 사료를 담는dispersion 방법은 실제 반추위 발효 상황에 근접한 조건에서 실험할 수 있는 장점이 있지만, 시간과 노동력이 많이 소모된다. 반대로 사료 주머니를 이용한 방법은 실험 수행의 편이성이 있지만 첨가제 실험 수행의 제약이 있으며, 미생물 활성과 섬유소 소화율이 저하되는 단점이 있다. 본 실험은 dispersion 방법과 사료 주머니 방법을 조합하여 보다 편리한 방법으로 dispersion에 가까운 높은 재현성의 발효 성상과 소화율 평가 가능할 것이라고 생각하여 in vitro 발효 성상과 미생물 조성을 비교하였다. Dispersion 방법과 nylon bag을 조합한 실험에서 건물 소화율은 세척 과정에서 미생물 및 사료 입자의 유실로 과대평가될 수 있지만 이론적 최대 가스 발생량 Vmax,가스 발생량에 대한 분해 속도 상수 Kg에서 유의적 차이가 나타나지 않으며, IVNDFD, pH, NH3-N, total VFA발생량 그리고 VFA 조성에서도 유의적 차이가 나지 않았다. Real-time PCR을 이용한 미생물 조성 분석에서는 general bacteria의 절대량에서 유의적 차이가 나지 않았다. 상기의 결과를 종합해 볼 때, in vitro 실험에서 사료 주머니의 사용은 충분히 적용가능하며 일반적인 방법인 dispersion과 비교해 볼 때, 영양소 이용에 있어 반추미생물의 사료 접촉을 방해하지 않을 것으로 생각된다. 하지만 in vitro bach culture 방법은 배양조건에서 제약이 있어, 향후 연속배양 방법과 in situ의 실험을 통해 본 실험의 실험결과에 대한 추가적인 규명이 필요할 것으로 사료된다.

9L 쥐 뇌종양에 있어서 체내 BrdU 표식지표와 체외 BrdU 표식지표의 비교

어환,長島正,松谷雅生,高倉公朋 대한신경외과학회 1992 Journal of Korean neurosurgical society Vol.21 No.12

The study of in vivo bromodeoxyuridine labeling index(BrdU LI) has become an established method alongside conventional histopathologic study for more complete and reliable evaluation of the biological behavior of tumors. However, in vivo BrdU LI study was limited because of requiring patient selection and patient consent In order to compare in vitro BrdU LI with in vivo one, the authors used 15 rats whose brains were stereotactically implanted with 9L glioma cell suspension, and incorporated BrdU into tumors in vivo or in vitro. In vivo BrdU LI was 18.3± 2.1 % ; in vitro LI of tumors incorporated BrdU within 30 minutes after tumor removal was 11.8± 2.6% ; in vitro LI of tumors incorporated 2 hours after removal was 9.7± 0.8% ; in vitro LI of tumors incorporated 4 hours after removal was 6.5± 3.0%. In vitro LIs were significantly lower than in vivo LI(p<0.05), and in vitro LI decreased gradually as the time of BrdU labeling passed But there were not statistically significant among in vitro LIs. In conclusion, it is essential to incorporate BrdU into tumor cells as soon as possible after tumor removal for in vitro BrdU LI. and in vitro LI is considered to take the place of in vivo LI for the evaluation of proliferating activity of tumor cells.

시험관내에서 인공배양한 제 3기 자충 및 성충을 이용한 구충효능 선발시험

지차호,박승준,Jee, Cha-ho,Park, Seung-jun 대한수의학회 1998 大韓獸醫學會誌 Vol.38 No.3

The in vitro screening tests against the in vitro cultivated $L_3$ of Ascaris suum (in vitro $L_3$), which were cultivated from the embryonated egg to third-stage larva on 7 days in culture(DIC) and the in vivo rat's lung-derived $L_3$ of Ascaris suum (in vivo $L_3$), which were recovered from the lungs of rat on 7 days after infection, carried out in order to compare the anthelmintic efficacy of in vitro $L_3$ and that of in vivo $L_3$ in RPMI medium 1640 with 5% bovine calf serum. And also a screening test of efficacy against adult worms of Trichuris suis performed. The efficacies of screening tests were as follows : 1. The screening efficacies of abamectin and ivermectin against the in vitro $L_3$ were all 100% at the 10ppm concentration in RPMI medium 1640 on 5 DIC. 2. The screening efficacies of abamectin and ivermectin against the in vivo $L_3$ were all 100% at the 20ppm on 5 DIC or at 40ppm on 3 DIC. 3. The screening efficacies of abamectin and ivermectin against the adult worms of Trichuris suis were all 100% at 20ppm on 4 DIC. And therefore, the in vitro cultivated $L_3$ of Ascaris suum were used in the screening test as well as the in vivo rat's lung-derived $L_3$ of Ascaris suum. And also the adult worms such as Trichuris suis and filaroids which is small size and difficult to cultivate to vitro, were used in the screening test in vitro.

Development of a Physiologically Relevant Population Pharmacokinetic <i>in Vitro</i>–<i>in Vivo</i> Correlation Approach for Designing Extended-Release Oral Dosage Formulation

Kim, Tae Hwan,Shin, Soyoung,Bulitta, Jü,rgen B.,Youn, Yu Seok,Yoo, Sun Dong,Shin, Beom Soo American Chemical Society 2017 Molecular Pharmaceutics Vol.14 No.1

<P>Establishing a level A in vitro in vivo correlation (IVIVC) for a drug with complex absorption kinetics is challenging. The objective of the present study was to develop an IVIVC approach based on population pharmacokinetic (POP-PK) modeling that incorporated physiologically relevant absorption kinetics. To prepare three extended release (ER) tablets of loxoprofen, three types of hydroxypropyl methylcellulose (HPMC 100, 4000, and 15000 cps) were used, as drug release modifiers, while lactose and magnesium stearate were used as the diluent and lubricant, respectively. An in vitro dissolution test in various pH conditions showed that loxoprofen dissolution was faster at higher pH. The in vivo pharmacokinetics of loxoprofen was assessed following oral administration of the different loxoprofen formulations to Beagle dogs (n = 22 in total). Secondary peaks or shoulders were observed in many of the individual plasma concentration vs time profiles after ER tablet administration, which may result from secondary absorption in the intestine due to a dissolution rate increase under intestinal pH compared to that observed at stomach pH. In addition, in vivo oral bioavailability was found to decrease with prolonged drug dissolution, indicating site-specific absorption. Based on the in vitro dissolution and in vivo absorption data, a POP-PK IVIVC model was developed using S-ADAPT software. pH dependent biphasic dissolution kinetics, described using modified Michaelis Menten kinetics with varying V-max and site-specific absorption, modeled using a changeable absorbed fraction parameter, were applied to the POP-PK IVIVC model. To experimentally determine the biphasic dissolution profiles, of the ER tablets, another in vitro dissolution test was conducted by switching dissolution medium pH based on an in vivo estimate of gastric emptying time. The model estimated, using linear regression, that in vivo initial maximum dissolution rate (Vmax(0)(in vivo)) was highly correlated (r(2) > 0.998) with in vitro (V-max(0)(in vitro)), indicating that in vivo dissolution profiles obtained from POP-PK modeling could be converted to in vitro dissolution profiles and vice versa. Monte Carlo simulations were performed for model validation, and prediction errors for C-max and AUC were all within the acceptable range (90 to 110%) according to the FDA guidelines. The developed model was successfully applied for the prediction of in vivo pharmacokinetics of a loxoprofen double-layered tablet using the in vitro dissolution profile. In conclusion, a level A IVIVC approach was developed and validated using population modeling that accounted for pH-dependent dissolution and site-specific absorption. Excellent correlations were observed between in vitro and in vivo dissolution profiles. This new approach holds great promise for the establishment of IVIVCs for drug and formulation development where absorption kinetics strongly depend on complex physiologically absorption processes.</P>

한약재 추출물이 반추위 in vitro 건물소화율, 휘발성 지방산 생성 및 미생물 성장률에 미치는 영향

문여황 경상대학교 농업생명과학연구원 2009 농업생명과학연구 Vol.43 No.6

This study was conducted to investigate the effect of herbal (Obtusifolia, Cinnamon, Chinese pepper, Licorice) extracts on the rumen fermentation in vitro. Comparing to the control, in vitro dry matter digestibility was significantly (P<0.05) decreased at zero hour in the Cinnamon and the Chinese pepper, and at three hour after supplementation in the Licorice. The ratio of volatile fatty acids were significant (P<0.05) differences at 3 hour after fermentation only, acetic acid was higher (P<0.05) in the control compare to the herbal extract treatments, but the ratios of butyrate, iso-butyrate, iso-valerate and valerate were lowest in the control. The growth rate of rumen microbes in vitro was significantly (P<0.05) higer in the herbal extract treatments excluding the Obtusifolia than the control during three hour fermentation, but was not significant difference among treatments in the other fermentation times. From above results, even though the extracts of Cinnamon, Chinese pepper and Licorice inclined to inhibit the activity of rumen microbes during early fermentation period, but did not affect on the growth rate of rumen microbes in vitro. 본 연구는 한약재 (결명자, 계피, 산초, 감초) 추출물의 반추위내 발효와 미생물 활성에 대한 효과를 구명하기 위하여 수행되었다. In vitro 건물소화율은 계피와 산초 추출물 첨가구에서는 0시간대, 감초 추출물 첨가구에서는 3시간대에 대조구에 비해 현저히 (P<0.05) 낮았다. 한약재 추출물 첨가에 따른 발효 시간대별 휘발성 지방산 조성의 변화는 3시간대에서만 처리간 유의성이 인정되었는데, acetate 비율은 대조구가 천연 추출물 첨가구보다 유의적으로 높았으나, butyrate, isobutyrate, isovalerate 및 valerate은 대조구에서 가장 낮았다 (P<0.05). 미생물 성장율은 발효 3시간대에서 결명자 첨가구를 제외한 한약재 첨가구에서 대조구에 비해 유의적 (P<0.05)으로 높았으나 다른 발효 시간대에서는 차이가 없었다. 이상의 결과로부터 한약재로 사용되고 있는 계피, 산초 및 감초 추출물을 in vitro 반추위 배양액에 첨가하였을 때, 발효초기에 반추위 미생물의 활성을 억제하는 경향은 있었으나 미생물 성장에 대한 억제 효과는 없는 것으로 나타났다.