This study evaluated the effect of lactic acid bacteria (LAB, a mixture of Enterococcus faecium and Lactobacillus plantarum) supplementation, the storage temperature, and storage period on the fermentation characteristics and in vitro ruminal digestibility of a total mixed ration (TMR). The TMR was prepared into two groups, namely, CON (control TMR without the LAB) and ML (supplementing a mixture of E. faecium and L. plantarum in the ratio of 1% and 2% (v/w), respectively). Both groups were divided and stored at 4°C or 25°C for 3, 7, and 14 d fermentation periods. Supplementing LAB to the TMR did not affect the chemical composition of TMR except for the lactate and acetate concentration. Storage temperatures affected (p<0.05) the chemical composition of the TMR, including pH, lactate, and acetate contents. The chemical composition of TMR was also affected (p<0.05) by the storage period. During in vitro rumen fermentation study, the ML treatment showed lower (p<0.05) dry matter digestibility at 24 h incubation with a higher pH compared to the CON. There was no difference in the in vitro dry matter digestibility (IVDMD) of TMR between the CON and ML treatment however, at 24 h, ML treatment showed lower (p<0.05) IVDMD with a higher pH compared to the CON. The effects of storage temperature and period on IVDMD were not apparent at 24 h incubation. In an in vivo study using Holstein steers, supplementing LAB to the basal TMR for 60 d did not differ in the final body weight and average daily gain. Likewise, the fecal microbiota did not differ between CON and ML. However, the TMR used for the present study did include a commercial yeast in CON, whereas ML did not; therefore, results were, to some extent, compromised in examining the effect of LAB. In conclusion, storage temperature and period significantly affected the TMR quality, increasing acetate and lactate concentration. However, the actual effects of LAB supplementation were equivocal.

This study was conducted to evaluate the effect of lactic acid bacteria (LAB) inoculation to domestically-cultivated Italian ryegrass (IRG) on silage fermentation and in vitro ruminal fermentation. There were six treatments based on the LAB inoculants: 1) no addition of LAB (negative control: NC), additions of 2) commercially-available LAB (positive control: PC), 3) Lactobacillus plantarum (LPL), 4) L. paracasei (LPA), 5) L. acidophilus (LA), and 6) L. pentosus (LPT). All treatments were inoculated at a concentration of 106 CFU/g and ensiled for 3, 7, 21, and 42 days in triplicate and analyzed for nutritive values when ensiling was terminated. Day 42 silage from all treatments were also examined for in vitro ruminal fermentation. After 42 days, LAB-inoculated silages had higher (P<0.05) lactic acid concentration compared to the NC. In terms of nutritive values, the silages treated with LPA, LA, and LPT showed higher (P<0.05) crude protein and lower (P<0.05) neutral detergent fiber and acid detergent fiber content compared to the rest of the treatment. In vitro ruminal dry matter degradability was not affected by LAB addition. However, LAB-treated IRG had shown higher (P<0.05) ammonia-N compared with that of the NC. LPA had shown the highest (P<0.05) volatile fatty acid concentration among the LAB examined. In conclusion, the addition of a single strain of LAB appeared to produce a quality IRG silage compared with the NC and the PC. Among the strains examined, LPA seemed to be superior to the others.

본 연구는 국화과 추출물 첨가가 in vitro 반추위 발효성상 및 메탄 발생에 미치는 영향을 구명하고자 수행하였다. 반추위액은 cannula 장착된 한우 암소 1두에서 채취하였으며, 공시축의 사양관리는 timothy와 농후사료를 6:4 비율로 하였다. 배양액은 Mcdougall buffer 10mL, 위액 5mL 섞어 50mL serum bottle에 혐기상태로 분주하였다. 티모시 0.3g를 각각 넣고 국화과 추출물인 국화, 민들레, 씀바귀, 제비쑥, 해바라기를 기질의 5%를 첨가한 뒤 발효시간대별(3, 6, 9, 12, 24, 48 및 72시간)로 7처리 3반복 수행하였다. 배양액 pH값은 6.29~7.44으로 반추위 적정 pH범위에 속하였다. 건물 소화율은 발효시간대가 늘어날수록 처리구에서 대조구에 비해 유의적(p<0.05)으로 증가하였다. 총가스 발생량은 발효 48시간대에서 민들레, 씀바귀, 제비쑥, 해바라기에서 대조구에 비해 유의적(p<0.05)으로 높게 측정 되었다. 이산화탄소 발생량은 발효 48시간대에서 민들레, 씀바귀, 제비쑥, 해바라기에서 대조구에 비해 유의적(p<0.05)으로 높게 측정 되었으며, 메탄 발생량도 발효 48시간대에서 민들레, 씀바귀, 제비쑥, 해바라기에서 대조구에 비해 유의적(p<0.05)으로 높게 측정 되었다. 미생물 성장량은 발효 48시간대에서 제비쑥과 해바라기 처리구에서 대조구에 비해 유의적(p<0.05)으로 높게 측정되었다. 발효 48시간대 에서 Acetic acid 생성량이 민들레, 씀바귀, 제비쑥, 해바라기 처리구에서 대조구에 비해 유의적(p<0.05)으로 높았다. 발효 12시간대에서 Propionic acid 생성량은 씀바귀, 제비쑥, 해바라기 처리구에서 대조구에 비해 유의적(p<0.05)으로 높았다. 발효 24시간대에서 A/P ratio는 민들레, 제비쑥, 해바라기 처리구에서 대조구에 비해 유의적(p<0.05)으로 감소하였다. 본 연구의 결과에 의해 국화과 추출물 첨가가 반추위 발효에 부정적인 영향을 미치지 않는 것으로 생각되나 메탄 발생량을 감소시키는데 영향이 없는 것으로 생각된다.

This study was conducted to examine the effect of corn (Zea mays L.) - soybean (Glycine max L.) silage prepared by intercropping method on the nutritive value of the silage, in vitro rumen fermentation characteristics, dry matter degradability, as well as milk yield and milk composition of dairy cows. In a couple of experiments intercropped corn-soybean silage (CSBS) was compared with corn silage (CS) and/or Italian ryegrass hay (IRG). Numerically, CSBS had higher crude protein, ether extract, and lactic acid contents compared to CS. In vitro rumen fermentation analysis demonstrated that up to a 24-h incubation period, both CS and CSBS showed higher total gas production, ammonia N concentration, and dry matter degradability compared to IRG (p<0.05). The investigation on animals was conducted in a commercial dairy farm located in Gyeongju, South Korea, employing 42 Holstein cows that were divided into 2 group treatments: CS and CSBS in a completely randomized design. Although no significant difference was observed in milk yield, animals fed on CSBS showed significantly higher milk protein (p<0.05) and milk fat content (p<0.01), compared to animals fed on CS. Taken together, our findings indicate that corn-soybean silage that is cultivated, harvested, and prepared through intercropping can improve the protein content of the silage, and can also enhance in vitro rumen fermentation, dry matter degradability, and performance of dairy cattle.

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.

This study was aimed to evaluate the dose-response the effects of nano-encapsulated conjugated linoleic acids(CLAs) on in vitro ruminal fermentation profiles. A fistulated Holstein cow was used as a donor of rumen fluid. Nano-encapsulated CLAs(LF, 5% of nano-encapsulated CLA-FFA; HF, 10% of nano-encapsulated CLA-FFA; LT, 5% of nano-encapsulated CLA-TG; HT, 10% of nano-encapsulated CLA-TG) were added to the in vitro ruminal fermentation experiment. In the in vitro ruminal incubation test, the total gas production on incubation with nano-encapsulated CLAs was increased significantly according to the incubation time, compared with the control(p<0.05). The tVFA concentrations on addition of LF and HT were significantly higher than that of the control(p<0.05). Thus, nano-encapsulated CLAs might improve the ruminal fermentation characteristics without adverse effects on the incubation process. In addition, the population of Butyrivibrio fibrisolvens which is closely related to ruminal biohydrogenation was increased by adding HT, while decreased by adding LF at 12 h incubation. These results showed that nano-encapsulated CLA-FFA could be applied to enhance CLA levels in ruminants by maintaining the stability of CLA without causing adverse effects on ruminal fermentation profiles considering the optimal dosage.

The study was conducted to evaluate the effects of microbial culture supplements on ruminal fermentation and fermentative quality of Italian ryegrass silage (IRGS) both in vitro and in situ. Three species of microbes (Lactobacillus casei (LC), Bacillus subtilis (BS), and Saccharomyces cerevisiae (SC)) were used in this study. They were applied to IRGS at 30 days after silage manufacture. Various items were measured using in vitro and in situ incubation technique after each microbial supplement was inoculated into IRGS at 0.5×104 CFU/g. In the first experiment, in vitro ruminal fermentation characteristics of IRGS were evaluated at 0, 12, 24, 48, and 72 hours after microbes were inoculated into IRGS. In the second experiment, in situ fermentation characteristics were investigated at 0, 1, 3, and 5 days after the inoculation of each microbial supplement. In vitro ruminal NH3-N content was significantly (p<0.05) increased in LC-, BS-, and SC-IRGS at 12 hrs post incubation compared to that in control IRGS. In vitro ruminal total VFA concentration and dry matter digestibility (DMD) of IRGS were not significantly difference among LC-, BS-, and SC-IRGS, although they were numerically increased in LC-IRGS than those of the other IRGS. In addition, this study evaluated the fermentation characteristics and in situ DMD of IRGS with the lapse of incubation time up to 5 days. Throughout the incubation times from 1 day to 5 days, the pH value was significantly (p<0.05) lower in BS-, LC-, and SC-IRGS than that in control IRGS. Lactate was significantly (p<0.05) higher, and significantly (p<0.05) butyrate was lower in LC-IRGS than that in other treatments at 0 day. It was higher (p<0.05) in control IRGS than that of BS-, LC-, and SC-IRGS at 1-5 days. In situ DMD tended to increase in BS-, LC-, and SC-IRGS compared to that in control IRGS. Especially, DMD was higher in SC-IRGS than that in other treatments at 0 day. It tended to be higher in LC-IRGS at all incubation time. Taken together, these results suggest that it might be useful to select a microorganism by considering the feeding time of IRGS to ruminants because organic acids and DMD of IRGS were affected by the incubation time of each microorganism with IRG silage, especially for L. casei decreased the content of acetate and butyrate in IRGS.

This study was conducted to evaluate effects of supplementation of Bifidobacterium ruminantium on in vitro ruminal fermentation and methane production. Ruminal fermentation characteristics of Timothy hay (C1), whole barley (C2), Timothy hay + B. ruminantium (T1) and whole barley + B. ruminantium (T2) were evaluated at 0, 3, 6, 9, 12, 24 and 48 h incubation at 39℃. The amount of B. ruminantium culture added into T1 and T2 was 0.3 ml. The pH values ranged from 5.99 to 6.83 in all the treatments. Concentration of NH3-N of C2 and T2 was higher than C1 and T1 at 48 h (p<0.05). The total gas production of C2 and T2 was higher than C1 and T1 at 9 h (p<0.05). The total methane production of treatments with B. ruminantium was not significantly different at 24 and 48 h (p<0.05). Concentration of lactic acid was significantly different between both substrates (p<0.05). Therefore, B. ruminantium supplementation was determined to be insignificant in the in vitro ruminal fermentation and methane production, while a further study was required to investigate relation to lactic acid production with different forage sources.

본 시험은 질산염함유 식물 추출물을 이용하여 in vitro 반추위 발효성상 및 반추위 메탄 발생에 미 치는 영향을 알아보기 위해 수행되었다. 공시재료는 감자, 당근, 배추, 상추 및 시금치 추출물을 사용하 였으며, 발효 기질은 2mm로 분쇄된 timothy 0.3g을 사용하였다. In vitro 실험은 조사료(timothy) 및 배합사료를 6:4의 비율로 급여한 반추위 cannnula가 시술된 한우 암소에서 채취하였다. 반추위액과 McDougall’s buffer를 1:2의 비율로 혼합한 발효액을 0.3g timothy와 식물 추출물이 담긴 50mL serum bottle에 혐기상태로 20mL 분주한 뒤, 39℃에서 9, 12, 24, 48시간 동안 발효하였다. pH는 발 효 시간이 증가함에 따라 점점 감소하며, 6.31~6.96으로 정상범위였다. 건물 소화율은 9시간 배추 추 출물구에서 유의적으로 낮았고, 암모니아 농도는 감자, 배추, 상추 추출물구에서 대조구에 비해 첨가구 에서 유의적으로(p<0.05) 낮았으며, 반추위 미생물 성장량은 발효 24시간대의 당근 추출물구에서 높았 다(p<0.05). Acetate와 propionate 농도는 대조구에 비해 첨가구에서 유의적으로 낮았으며(p<0.05) butyrate 농도는 첨가구간에 발효시간별로 각각 다른 양상을 보였다. 총 가스 발생량은 발효 12시간 및 24시간대에서 배추, 상추, 시금치 추출물구에서 유의적으로(p<0.05) 낮았으며, 메탄 발생량은 감자, 배 추, 시금치 추출물구가 대조구에 비해 유의적으로(p<0.05) 낮았고, 이산화탄소 발생량 또한 첨가구에 비해 대조구에서 유의적으로(p<0.05) 낮았다. 결과적으로 본 시험에 사용된 5종의 질산염 함유 식물 추 출물은 반추위 발효에 악영향을 미치지 않으면서 메탄저감 효과를 나타내었다. 특히 배추 추출물은 반 추위내 메탄발생을 저감시킬 수 있는 식물추출물로서 가능성이 높은 것으로 사료된다.

The objective of this study was to determine the effect of bacterial inoculation (Lactobacillus plantarum or combo inoculant mixed with Lactobacillus plantarum and Lactobacillus buchneri) and addition of fibrolytic enzyme on chemical compositions and fermentation characteristics of whole crop barley (WCB) and triticale (TRT) silage, their ruminal in vitro fermentation, and digestibility. In TRT silage, enzyme addition significantly (p<0.01) decreased NDF content compared to no enzyme addition treatment. Organic acids such as lactate and acetate contents in WCB and TRT silages were significantly (p<0.01) higher compared to those in the control. Particularly, lactate content was the highest in L. plantarum treatment. Fibrolytic enzyme treatment on both silages had relatively higher lactic acid bacteria content, while mold content was lower in both treatments compared to that in the control. In vitro dry matter digestibility was generally improved in WCB silages. It was higher (p<0.01) in TRT with mixed treatment of L. plantarum, L. buchneri, and enzyme compared to others. In vitro ruminal acetate production was relatively higher in treatments with both enzyme and inoculant additions compared to that in the control. Therefore, the quality of silage and rumen fermentation could be improved by inoculants (L. plantarum and L. buchneri) regardless whether whole crop barley (WCB) or triticale (TRT) silage was used. Although it was found that fibrolytic enzyme addition to both silages had various quality and rumen fermentation values, further study is needed

The in vitro experiment was conducted to ensure the supplemental level of spent Flammulina velutipes mushroomsubstrates (SMS) as an energy source in manufacturing of whole crop sorghum silage. Sorghum harvested at heading stage wasensiled with spent mushroom substrates of 20% (S-20), 40% (S-40) and 60% (S-60) as fresh matter basis for 6 week. Theexperiment was conducted by 3, 6, 9, 12, 24, 48 hrs of incubation time with 3 replications. The silages were evaluatedfermentation characteristics and dry matter digestibility (DMD) in vitro. The pH of in vitro solution was inclined to decrease withelapsing the incubation time, and that of the S-20 was significantly (P<0.05) lower than the other treatment at 48 hr ofincubation. Gas production was greater (P<0.05) in the S-20 than the other treatments at 6 and 12 hrs of incubation. Themicrobial growth in vitro was inclined to decrease following 24 hr of incubation, and thereafter sustained the similar levels. Invitro dry matter digestibility (IVDMD) was lowered by increasing the supplemental level of spent mushroom substrate, and was alow level in the S-60 throughout whole incubation time. Althoughthe IVDMD for S-40 was steadily increased from 9 hr ofincubation and reached to similar level with the S-20 at 48 hourof incubation, however SMS for whole crop sorghum silagefermentation might as well add about 20 to 30% in fresh matterbasis when considering DMD.

The objective of this study was to know the in vitro effects of supplemental anthelmintic plant extracts on the inhibition of protozoa for reducing methane production in the rumen. A fistulated Holstein cow was used as a donor of rumen fluid. The plant extracts (Lonicera japonica, Zanthoxylum piperitum, Pyrethrum, Torreya nucifera, Ruta graveolens) known to have anthelmintic effect were added to the in vitro fermentation bottles containing the rumen fluid and medium. The rumen protozoal population was depressed by the addition of Pyrethrum, Torreya nucifera and Ruta graveolens. The methane production was also significantly (p<0.05)reduced by addition of Pyrethrum (2.20 ml/g DM), Torreya nucifera (2.36 ml/g DM) and Ruta graveolens (2.20 ml/g DM). The microbial growth in the treatments of Ruta graveolens or anthoxylum piperitum was the greatest after 12 h and 24 h incubations, respectively. The results of this study indicated that anthelmintic plant extracts appeared to reduce methane production by inhibition of ruminal protozoa related with the methanogens living endosymbiotic in protozoal cells.

본 시험은 효소제 및 효모추출물 급여가 사양성적, in vitro 반추위 발효 및 혈액성상에 미치는 영향을 구명하기 위해 23개월령의 한우 거세우 48두를 공시하여 사료용 첨가제로 농후사료의 0.1% 수준으로 급여하여 출하시까지 대조구(무첨가), 처리구 1(효소제), 처리구 2(효모추출물), 처리구 3(효소제+효모추출물)으로 구분하여 급여하였다. 사양성적에서는 급여 4개월과 8개월의 일당증체량과 사료요구율이 처리구 1에서 가장 향상되었고 (p<0.05), 대조구에서 가장 낮았다(p<0.05). 도축성적에서는 육량지수를 포함하여 처리구 1>처리구 3>처리구 2>대조구 순으로 나타났다. in vitro 배양액에 따른 처리구별 pH는 대조구>처리구 2>처리구 3>처리구 1 순으로 유의적(p<0.05)인 차이를 나타냈고, VFA와 사료소화율은 처리구 1과 3에서 높았고 (p<0.05), 대조구에서는 낮았다(p<0.05). 혈액성상에서는 면역 첨가제 급여군인 처리구 2와 3에서 급여기간에 따른 백혈구 수치가 유의적(p<0.05)으로 감소하였는데, 이는 면역글로블린 G의 유의적(p<0.05)인증가에 따라서 상대적으로 나타난 것으로 판단된다. 결과적으로, 한우 비육후기 거세우의 사료첨가제 급여 시험에서 처리구 1은 급여 4개월부터 대조구 대비 사양성적 및 도체성적을 향상시켰고, 처리구 2는 급여 2개월부터 면역에 관련된 면역글로블린 G의 혈액성상을 개선시키는 것으로 분석되었다.

This study is conducted to screen compounds affecting ruminal fermentation under in vitro incubation. Saponin, chitosan, metformin, zinc acetate, zinc chloride, zinc oxide, zinc sulfate, ascorbic acid, oil-coated ascorbic acid, nicotinic acid, eastern herb cocktail, charcoal extract and garlic sources (lyophilized and extract) were added to rumen culture fluid at 1.25% of substrate (100% timothy) volume at 24 h incubation. pH, total gas, ammonia, VFAs were measured. Ascorbic acid increased total gas production indicating fermentation level. All zinc compounds significantly decreased (p<0.05) although zinc sulfate increased propionate of volatile fatty acid (p<0.05). In following experiment, ascorbic acid, oil-coated ascorbic acid, garlic lyophilized, herb cocktail and zinc sulfate were added to rumen culture fluid at 2.5% of substrate with 3, 6, 12 and 24 h incubation. Zinc sulfate decreased both ruminal fermentation and VFAs production but ascorbic acid enhanced total gas production. Ascorbic acid increased fermentation regardless of supplement concentration although excessive zinc sulfate decreased fermentation. These results suggest that optimal level of trace compounds might affect ruminal fermentation in ruminant.

Substantial efforts have been made to manipulate ruminal environment in a hope to enhance ruminal fermentation efficiency for better ruminant productivity. Some of examples are methane inhibitors, antibiotics, microbial enzymes, fatty acids and/or lipid feeding, buffering agents, ionophores and probiotics. Of these efforts, the non-ionic surfactant (NIS) has been known for its stimulation to release enzymes from a range of anaerobic microbes. A series of studies were conducted 1) to evaluate NIS diluted with water and ethanol on in vitro ruminal fermentation and 2) to determine the influence of diluted NIS on digestibility of feedstuffs. In first experiment (Exp. 1), NIS was diluted with water or ethanol to measure its effects on in vitro microbial growth, ruminal enzyme activities and gas production by mixed ruminal microbial culture. The NIS was diluted with water or ethanol separately in a 1:5 ratio (w/v). Water and ethanol-diluted NIS with wheat flour were added with rice straw-based mixed ruminal microbial cultures at the rate of 2 ㎎ NIS/16 ㎖ McDougall buffer plus 4 ml ruminal fluid solution. The mixed ruminal microbial culture was run without any NIS addition as control. Addition of NIS either diluted with water or ethanol has significantly reduced the gas production in mixed ruminal microbial culture at 12 and 24 h of incubation. At 48 h post incubation, gas production was the highest with the addition of ethanol diluted NIS followed by water-diluted NIS and control. Carboxy methyl cellulase activity in rice straw-based mixed ruminal bacterial culture was significantly higher with the addition of ethanol-diluted NIS compared with the addition of water-diluted NIS and control at 24 and 72 h post incubation. In second in vitro experiment (Exp. 2), effects of addition of ethanol diluted NIS on dry matter (DM) digestibility of alfalfa hay, gas production, pH and cellular growth in mixed ruminal microbial culture were examined. Alfalfa hay based mixed ruminal microbial culture without any NIS addition was run as a control. The pH of mixed ruminal microbial culture was significantly lower than control at all post incubation sampling hours. In vitro DM digestibility of alfalfa hay was significantly higher with the addition of NIS compared with control. Gas production was significantly less with NIS addition compared with control at all post incubation sampling hours. Microbial growth in mixed ruminal microbial culture was significantly increased with the addition of NIS compared to control.

본 연구는 반추위 보호 choline의 급여가 in vitro 반추위 발효특성과 착유우의 유생산성에 미치는 영향을 검토하기 위해 수행되었다. 시험구 배치는 기초사료 첨가구(T1), 기초사료 +23 g/일 보호처리를 하지 않은 choline과 부형제 첨가구(T2) 및 기초사료+ 25.56 g/일 보호 choline 첨가구(T3)의 3처리로 하였다. 보호 choline의 첨가가 in vitro 배양액의 pH에 미치는 영향은 3시간을 제외한 전 배양시간에서