• Title/Summary/Keyword: In Vitro Fermentation

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Interactions between Entodinium caudatum and an amino acid-fermenting bacterial consortium: fermentation characteristics and protozoal population in vitro

  • Tansol Park;Zhongtang Yu
    • Journal of Animal Science and Technology
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    • v.65 no.2
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    • pp.387-400
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    • 2023
  • Ruminal protozoa, especially entodiniomorphs, engulf other members of the rumen microbiome in large numbers; and they release oligopeptides and amino acids, which can be fermented to ammonia and volatile fatty acids (VFAs) by amino acid-fermenting bacteria (AAFB). Studies using defaunated (protozoa-free) sheep have demonstrated that ruminal protozoa considerably increase intraruminal nitrogen recycling but decrease nitrogen utilization efficiency in ruminants. However, direct interactions between ruminal protozoa and AAFB have not been demonstrated because of their inability to establish axenic cultures of any ruminal protozoan. Thus, this study was performed to evaluate the interaction between Entodinium caudatum, which is the most predominant rumen ciliate species, and an AAFB consortium in terms of feed degradation and ammonia production along with the microbial population shift of select bacterial species (Prevotella ruminicola, Clostridium aminophilum, and Peptostreptococcus anaerobius). From an Ent. caudatum culture that had been maintained by daily feeding and transfers every 3 or 4 days, the bacteria and methanogens loosely associated with Ent. caudatum cells were removed by filtration and washing. An AAFB consortium was established by repeated transfers and enrichment with casamino acids as the sole substrate. The cultures of Ent. caudatum alone (Ec) and AAFB alone (AAFB) and the co-culture of Ent. caudatum and AAFB (Ec + AAFB) were set up in three replicates and incubated at 39℃ for 72 h. The digestibility of dry matter (DM) and fiber (NDF), VFA profiles, ammonia concentrations, pH, and microscopic counts of Ent. caudatum were compared among the three cultures. The co-culture of AAFB and Ent. caudatum enhanced DM degradation, VFA production, and Ent. caudatum cell counts; conversely, it decreased acetate: propionate ratio although the total bacterial abundance was similar between Ec and the Ec + AAFB co-culture after 24 h incubation. The ammonia production and relative abundance of C. aminophilum and P. anaerobius did not differ between AAFB alone and the Ec + AAFB co-culture. Our results indicate that Ent. caudatum and AAFB could have a mutualistic interaction that benefited each other, but their interactions were complex and might not increase ammoniagenesis. Further research should examine how such interactions affect the population dynamics of AAFB.

In-vitro Anti-thrombosis Activity of R4-Nuruk Made from Rhizopus oryzae KSD-815 (Rhizopus oryzae KSD-815를 이용하여 제조된 R4 누룩의 항혈전 활성)

  • Kim, Mi-Sun;Lee, Ye-Seul;Kim, Jong Sik;Shin, Woo-Chang;Sohn, Ho-Yong
    • Microbiology and Biotechnology Letters
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    • v.43 no.2
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    • pp.169-174
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    • 2015
  • Nuruk has been used as fermentation starter in the alcohol industry for some time in Korea. Various bioactivities, such as antiproliferative and anti-obesity, of R4 nuruk made from Rhizopus oryzae KSD-815 have been previously reported. In this study, the hot water and ethanol extract of R4 nuruk and their subsequent organic solvent fractions were prepared, and their antithrombosis activities were evaluated. The ethylacetate fraction showed strong anti-coagulation activity, and the ethylacetate and butanol fraction from hot water extract demonstrated platelet aggregation inhibitory activity, without hemolysis against human RBC. Our results suggest that R4 nuruk has the potential to act as a new anti-thrombosis agent.

High-Level Production of High-Purity Human and Murine Recombinant Prion Proteins Functionally Compatible to In Vitro Seeding Assay

  • Hwang, Hae-Gwang;Kim, Dae-Hwan;Lee, Jeongmin;Mo, Youngwon;Lee, Se-Hoon;Lee, Yongjin;Hyeon, Jae Wook;Lee, Sol Moe;Cheon, Yong-Pil;Choi, Eun-Kyoung;Kim, Su Yeon;Lee, Yeong Seon;Son, Young-Jin;Ryou, Chongsuk
    • Journal of Microbiology and Biotechnology
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    • v.28 no.10
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    • pp.1749-1759
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    • 2018
  • Recombinant (rec) prion protein (PrP) is an extremely useful resource for studying protein misfolding and subsequent protein aggregation events. Here, we report mass production of high-purity rec-polypeptide encoding the C-terminal globular domain of PrP; (90-230) for human and (89-231) for murine PrP. These proteins were expressed as His-tagged fusion proteins in E. coli cultured by a high cell-density aerobic fermentation method. RecPrPs recovered from inclusion bodies were slowly refolded under reducing conditions. Purification was performed by a sequence of metal-affinity, cation-exchange, and reverse-phase chromatography. The current procedure yielded several dozens of milligrams of recPrP per liter with >95% purity. The purified recPrPs predominantly adopted an ${\alpha}$-helix-rich conformation and were functionally sufficient as substrates to measure the seeding activity of human and animal prions. Establishment of a procedure for high-level production of high-purity recPrP supports the advancement of in vitro investigations of PrP including diagnosis for prion diseases.

Effect of the Level of Carbohydrates on Bio-hydrogenation and CLA Production by Rumen Bacteria When Incubated with Soybean Oil or Flaxseed Oil In vitro (Soybean Oil 및 Flaxseed Oil 첨가 배양시 탄수화물 첨가수준에 의한 반추미생물의 Bio-hydrogenation과 CLA 생성에 미치는 효과)

  • 최성호;임근우;김광림;송만강
    • Journal of Animal Science and Technology
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    • v.48 no.4
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    • pp.521-532
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    • 2006
  • An in vitro study was conducted to examine the effect of addition level of carbohydrates on fermentation characteristics, and bio-hydrogenation of unsaturated fatty acids by mixed rumen bacteria when incubated with soybean oil or flaxseed oil. Four levels(0%, 0.3%, 0.6% and 0.9%, w/v) of the mixed carbohydrates(glucose, cellobiose, soluble starch, 1:1:1, in weight basis) and oil sources(soybean or flaxseed oil, 60mg in 150ml culture solution) were added to the mixed solution of strained rumen fluid with artificial saliva(1:4, v/v), and incubated anaerobically for 12 hours at 39℃. pH and ammonia-N concentration were lower by increasing the substrate levels at all incubation periods(P<0.05~P<0.001). The propionate proportion increased(P<0.001), but acetic acid and butyric acid decreased(P<0.001) with the substrate level at 6 and 12 h incubations. Oil sources did not influence the proportions of individual VFA. At the end of incubation, the proportions of C18:0(P<0.01), C18:1(P<0.001) and trans-11C-18:1(P<0.01) were reduced but those of C18:2(P<0.001) and C18:3(P<0.01) were enhanced by the addition of flaxseed oil compared to addition of soybean oil. The proportions of C18:0 and total CLA were reduced(P<0.01) but those of trans-11-C18: (P<0.05) and C18:2(P<0.01) were increased with the substrate level when incubated with soybean oil or flaxseed oil. There were interactions(P<0.05) in the proportions of C18:1, C18:2 and C18:3(P<0.01) between oil source and substrate level. The proportions of cis-9, trans-11-CLA and trans-10, cis-12-CLA tended to reduce with substrate level, although there was no significant difference between treatments.

Chemical Composition, Degradation Characteristics and Effect of Tannin on Digestibility of Some Browse Species from Kenya Harvested during the Wet Season

  • Osuga, I.M.;Abdulrazak, S.A.;Ichinohe, T.;Fujihara, T.
    • Asian-Australasian Journal of Animal Sciences
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    • v.18 no.1
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    • pp.54-60
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    • 2005
  • A study was conducted with the objective of evaluating the nutritive value of some browse species from Kenya. The species evaluated included: Bauhinia alba, Bauhinia variegata, Bridelia micrantha, Calliandra calothyrsus, Carisa edulis, Cratylia argentea, Gliricidia sepium, Lantana camara, Maerua angolensis, Sesbania micrantha and S. sesban. The browses were evaluated by their chemical composition including phenolics, in vitro gas production and tannin activity (tannin bioassay). All the species had high crude protein content (149-268 g/kg DM) and low NDF content (239-549 g/kg DM). The feeds had varying contents of total extractable tannins (TET) ranging from low (3-22 mg/g DM), moderate (42-58 mg/g DM) and high (77-152 mg/g DM). Calliandra calothyrsus had the highest tannin content. Significant (p<0.05) variation in gas production was recorded among the species. Sesbania micrantha had the highest (p<0.05) potential gas production while Gliricidia sepium had the highest (p<0.05) rate of gas production. Use of polyethylene glycol (PEG 6000), to assess the adverse affect of tannins, indicated that tannins in browse species with high tannin content had inhibitory effects on rumen microbial fermentation as indicated by the gas production. Estimated organic matter digestibility and metabolizable energy also increased with PEG addition. The results of this study indicate that such Kenyan browse species have the potential to be used as feed supplements for ruminant animals.

Physiological Effects of Formulations Added with Black Garlic Extract on Skin Care: Oxidative Stress, Tyrosinase and Elastase Activities (흑마늘 추출물을 첨가한 화장품의 기능성 평가: Tyrosinase와 Elastase 저해 활성 및 산화 스트레스에 미치는 영향 중심으로)

  • Jung, Eun-Young;Hong, Yang-Hee;Kim, Seon-Hee;Suh, Hyung-Joo
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.39 no.5
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    • pp.662-668
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    • 2010
  • We evaluated the physiological effects of formulations added with black garlic extract on skin care. Black garlic was made by spontaneous fermentation for 40 days at $60{\sim}70^{\circ}C$ and 85~95% RH without any additives. When black garlic extract was added to formulation, the tyrosinase activity was decreased significantly (p<0.05, p<0.01, p<0.001). Base formulation inhibited slightly elastase activity (<5%), while black garlic formulation had about 8-fold higher elastase inhibitory activity (p<0.01). It was observed that formulations, with or without black garlic extract, decreased thiobarbituric acid reactive substances (TBARS) and hydrogen peroxide which were increased by UVB irradiation, although there were no significant differences of these contents between black garlic formulation and base formulation. In conclusion, black garlic formulation had high inhibitory activities for tyrosinase and elastase, suggesting that black garlic may have beneficial properties as a material for cosmeceuticals.

Phylogenetics, Safety and In Vitro Functional Properties of Bacillus Species Isolated from Iru, a Nigerian Fermented Condiment

  • Adewumi, Gbenga Adedeji;Grover, Sunita;Isanbor, Chukwuemeka;Oguntoyinbo, Folarin Anthony
    • Microbiology and Biotechnology Letters
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    • v.47 no.4
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    • pp.498-508
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    • 2019
  • Bacillus species were isolated from iru, a traditional fermented condiment in Nigeria. Polyphasic approach was used to evaluate the phylogenetic relationship and strain sub-type of the isolated species. Additionally, the phylogenetic profiles of the species isolated from iru were compared with those of bacilli isolated from different continents. The phylogenetic diversity analysis was performed using the combination of 16S rRNA gene sequencing, ITS-PCR, ITS-PCR-RFLP, and M13 RAPD-PCR. The analysis revealed that Bacillus subtilis U170B and B. subtilis U146A isolated from iru were the closest relatives of strains belonging to the phylogeny of B. subtilis sensu stricto and were related to other bacilli isolated from different continents that had functional benefits. The two isolated species exhibited resistance to acidic pH (pH 2.0). The survival rates of B. subtilis U170B, B. subtilis U146A, and B. clausii UBBC-07 (commercial probiotic strain) cultured at pH 2.0 for 3 h were 33.45, 12.44, and 9.53%, respectively. The strains were highly tolerant to bile salts [0.3% (w/v)]. B. subtilis U170B exhibited the highest cell viability (43.45%) when cultured for 3 h in the presence of bile salts, followed by B. subtilis U146A (25%) and B. clausii UBBC-07 (18.94%). B. subtilis U170B and B. subtilis U146A did not exhibit haemolytic activity and were susceptible to different antibiotics. Additionally, these two strains exhibited weak antagonistic activity against B. cereus. The diverse wild strains of B. subtilis can be used as a safe multifunctional starter culture for the industrial production of condiments with health benefits.

Effects of Dietary Potential Acid Production Value on Productivity in Dairy Cows

  • Kim, E.T.;Lee, S.S.;Kim, H.J.;Song, J.Y.;Kim, C.H.;Ha, Jong-K.
    • Asian-Australasian Journal of Animal Sciences
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    • v.25 no.5
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    • pp.653-658
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    • 2012
  • This study was conducted to estimate the potential acid production value (PAPV) of major diets and to determine the relationship between dietary PAPV and dairy production traits. Estimation of PAPV of major cattle feeds was based on an in vitro technique, which determined the degree of Ca dissociation from $CaCO_3$. Data on feeds and production traits were collected on 744 multiparous lactating Holstein dairy cows from five different farms. Grains had high PAPV with variable protein sources and by-products. High PAPV feedstuffs had a higher total gas production and lower pH compared to those with low PAPV. Dietary PAPV had a positive correlation with intake of dry matter, NDF, ADF, milk yield and milk solid production but a negative correlation with milk protein and milk fat concentration. Current results indicate that dietary PAPV can be utilized in predicting dairy production traits.

Isolation of antibiotic resistant Lactobacillus spp from fermented milk and starters, and detection of transmissible R plasmids (유산균 발효유 및 starter에서 항생제내성 유산균 분리 및 전달성 R plasmids 분포조사)

  • Baek Kui-Jeong;Seo Heyng-Seok;Roh Young-Sun;Yang Hae-Dong;Hur Boo-Hong;Seo Lee-Won;Joung Dong-Suk;Song Hee-Jong
    • Korean Journal of Veterinary Service
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    • v.29 no.3
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    • pp.277-285
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    • 2006
  • Plasmids are covalently closed circular molecules of DNA that are stably inherited and replicate somewhat independently of the bacterial chromosome. Genes carried on plasmids can mediate a wide variety of important functions, including antibiotics (R plasmids) and heavy metals resistance, toxins production, cell penetration, iron chelation, complement resistance, and metabolic characteristics such as sucrose and lactose fermentation. Fifty strains of lactobacilli were isolated from 26 staters and 29 fermented milk products. They were classified 27 strains as Lactobacillus paracasei subsp paracasei, 11 stains as Lactococcus lactis subsp cremoris, 6 strains as L delbrueckii subsp lactis, 4 strains as L acidophius, and 2 strains as L delbrueckii subsp bulgaricus. All of these strains were examined for drug resistance and transferability of R plasmids. All of the isolates were sensitive to Am, C, CF, E, NB, P, T, and Te. But resistant to SXT 94% (47 strains), K 66% (33 strains), S 56% (28 strains), ENR 50% (25 strains), NOR 38% (19 strains) CIP 38% (19 strains), GM 16% (8 strains), and N 14% (7 strains), in order. And 32 different resistant patterns were found. The most frequently encountered patterns were CIP-ENR-K-NOR-S-SXT (5 strains). In vitro R plasmids transfer experiment, 57 antibiotic resistant strains which were not transfer to the recipient 2 Escherichia coli strains by conjugation, These results indicate that Lactobacillus in internal trade market' stater recognize R factor but transmissible R plasmid is not existed.

In Vitro Profiling of Bacterial Influence and Herbal Applications of Lonicerae Flos on the Permeability of Intestinal Epithelial Cells (장내 상피세포 점막 투과성에 대한 유산균 및 금은화의 효과)

  • Lee, Sin-Ji;Lee, Myeong-Jong;Jung, Ji-Eun;Kim, Ho-Jun;Bose, Shambhunath
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.41 no.7
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    • pp.881-887
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    • 2012
  • This study was designed to examine influence of bacteria and herbal applications of Lonicerae Flos on the permeability of intestinal epithelial cells. The HCT-116 human intestinal cell was used as an in vivo model of "leaky gut". Dextran sodium sulphate (DSS) was used to induce an increase in the permeability of epithelial cell tight junctions. Probiotics including Lactobacillus acidophilus, Lactobacillus plantarum, Salomonella spp. and Staphylococcus aureus were used to evaluate bacterial influence on the intestinal epithelial cells. Herbal extract Lonicerae Flos was also assessed to find out what kind of effects it has when used alone or fermented with Lactobacillus plantarum. The permeability of intestinal cell lines was assessed using transepithelial electrical resistance (TEER) and horseradish peroxide (HRP) assessments. This data provides evidence for the probiotic benefits of intestinal permeability. The data also provides promising evidence of herbal effects on reducing intestinal permeability, especially when fermented with probiotics.