• Animal Bioscience
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• 제34권3_spc호
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• pp.463-470
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• 2021

Objective: This experiment was conducted to find out the immunological effects of wheat phytase when long-chain inorganic polyphosphate (polyP) treated with wheat phytase was added to a macrophage cell line, Raw 264.7, when compared to intact long-chain polyP. Methods: Nitric oxide (NO) production of Raw 264.7 cells exposed to P700, a long-chain polyP with an average of 1,150 phosphate residues, treated with or without wheat phytase, was measured by Griess method. Phagocytosis assay of P700 treated with or without phytase in Raw 264.7 cells was investigated using neutral red uptake. The secretion of tumor necrosis factor α (TNF-α) by Raw 264.7 cells with wheat phytase-treated P700 compared to intact P700 was observed by using Mouse TNF-α enzyme-linked immunosorbent assay kit. Results: P700 treated with wheat phytase effectively increased NO production of Raw 264.7 cells by 172% when compared with intact P700 at 12 h exposure. At 5 mM of P700 concentration, wheat phytase promoted NO production of macrophages most strongly. P700, treated with wheat phytase, stimulated phagocytosis in macrophages at 12 h exposure by about 1.7-fold compared to intact P700. In addition, P700 treated with wheat phytase effectively increased in vitro phagocytic activity of Raw 264.7 cells at a concentration above 5 mM when compared to intact P700. P700 dephosphorylated by wheat phytase increased the release of TNF-α from Raw 264.7 cells by 143% over that from intact P700 after 6 h exposure. At the concentration of 50 μM P700, wheat phytase increased the secretion of cytokine, TNF-α, by 124% over that from intact P700. Conclusion: In animal husbandry, wheat phytase can mitigate the long-chain polyP causing damage by improving the immune capabilities of macrophages in the host. Thus, wheat phytase has potential as an immunological modulator and future feed additive for regulating immune responses caused by inflammation induced by long-chain polyP from bacterial infection.

• Journal of Animal Science and Technology
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• 제63권1호
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• pp.114-124
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• 2021

The objective of this study was to characterize the enzymatic hydrolysis of lipopolysaccharide (LPS) by wheat phytase and to investigate the effects of wheat phytase-treated LPS on in vitro toxicity, cell viability and release of a pro-inflammatory cytokine, interleukin (IL)-8 by target cells compared with the intact LPS. The phosphatase activity of wheat phytase towards LPS was investigated in the presence or absence of inhibitors such as L-phenylalanine and L-homoarginine. In vitro toxicity of LPS hydrolyzed with wheat phytase in comparison to intact LPS was assessed. Cell viability in human aortic endothelial (HAE) cells exposed to LPS treated with wheat phytase in comparison to intact LPS was measured. The release of IL-8 in human intestinal epithelial cell line, HT-29 cells applied to LPS treated with wheat phytase in comparison to intact LPS was assayed. Wheat phytase hydrolyzed LPS, resulting in a significant release of inorganic phosphate for 1 h (p < 0.05). Furthermore, the degradation of LPS by wheat phytase was nearly unaffected by the addition of L-phenylalanine, the inhibitor of tissue-specific alkaline phosphatase or L-homoarginine, the inhibitor of tissue-non-specific alkaline phosphatase. Wheat phytase effectively reduced the in vitro toxicity of LPS, resulting in a retention of 63% and 54% of its initial toxicity after 1-3 h of the enzyme reaction, respectively (p < 0.05). Intact LPS decreased the cell viability of HAE cells. However, LPS dephosphorylated by wheat phytase counteracted the inhibitory effect on cell viability. LPS treated with wheat phytase decreased IL-8 secretion from intestinal epithelial cell line, HT-29 cell to 14% (p < 0.05) when compared with intact LPS. In conclusion, wheat phytase is a potential therapeutic candidate and prophylactic agent for control of infections induced by pathogenic Gram-negative bacteria and associated LPS-mediated inflammatory diseases in animal husbandry.

• Animal Bioscience
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• 제36권10호
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• pp.1604-1611
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• 2023

Objective: The aim of this study was to investigate the protective effect of wheat phytase as a structural decomposer of inflammatory nucleotides, extracellular adenosine triphosphate (ATP), and uridine diphosphate (UDP) on HT-29 cells. Methods: Phosphatase activities of wheat phytase against ATP and UDP was investigated in the presence or absence of inhibitors such as L-phenylalanine and L-homoarginine using a Pi Color Lock gold phosphate detection kit. Viability of HT-29 cells exposed to intact- or dephosphorylated-nucleotides was analyzed with an EZ-CYTOX kit. Secretion levels of pro-inflammatory cytokines (IL-6 and IL-8) in HT-29 cells exposed to substrate treated with or without wheat phytase were measured with enzyme-linked immunosorbent assay kits. Activation of caspase-3 in HT-29 cells treated with intact ATP or dephosphorylated-ATP was investigated using a colorimetric assay kit. Results: Wheat phytase dephosphorylated both nucleotides, ATP and UDP, in a dose-dependent manner. Regardless of the presence or absence of enzyme inhibitors (L-phenylalanine and L-homoarginine), wheat phytase dephosphorylated UDP. Only L-phenylalanine inhibited the dephosphorylation of ATP by wheat phytase. However, the level of inhibition was less than 10%. Wheat phytase significantly enhanced the viability of HT-29 cells against ATP- and UDP-induced cytotoxicity. Interleukin (IL)-8 released from HT-29 cells with nucleotides dephosphorylated by wheat phytase was higher than that released from HT-29 cells with intact nucleotides. Moreover, the release of IL-6 was strongly induced from HT-29 cells with UDP dephosphorylated by wheat phytase. HT-29 cells with ATP degraded by wheat phytase showed significantly (13%) lower activity of caspase-3 than HT-29 cells with intact ATP. Conclusion: Wheat phytase can be a candidate for veterinary medicine to prevent cell death in animals. In this context, wheat phytase beyond its nutritional aspects might be a novel and promising tool for promoting growth and function of intestinal epithelial cells under luminal ATP and UDP surge in the gut.

• Journal of Animal Science and Technology
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• 제44권4호
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• pp.407-418
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• 2002

Phytase 공급원으로 밀과 밀기울의 첨가가 육계 생산성 및 P이용률에 미치는 영향을 측정하기 위해 5주간의 사양 실험을 실시하였다. 갓 부화한 1,000수의 병아리(Ross$^{(R)}$)를 20pen에 50수씩(암수 각 25수) 공시하여 5처리 4반복으로 완전임의 배치하였다. T1은 대조구로 정상수준의 nonphytate-P(NPP)를 함유하였고, T2는 T1 －0.1% NPP의 低 NPP구, T3는 T2 + 사료 kg당 600IU microbial phytase(NOVO$^{(R)}$)구, T4는 T2 + 밀과 밀기울로 사료 kg당 600IU plant phytase 공급구, T5는 T2 + 밀과 수침 후 건조 처리한 밀기울로 사료 kg당 600IU plant phytase 공급구 였다. NPP 수준을 0.1% 감소시킨(T2) 처리에서 생산성과 사료효율이 감소되었고 식물성 phytase 처리시(T4와 T5)에서는 생산성이 회복되었다. 식물성 phytase 처리구들(T4와 T5)이 microbial phytase 처리구(T3)보다 더 나은 결과를 보였다. 정상 밀기울구(T4)와 수침 후 건조 처리한 밀기울구(T5)와는 차이를 보이지 않았다. 폐사율은 低  NPP(T2)에서 가장 높았다. 육성 사료에서 조지방과 조회분 이용률에 일반 밀기울 처리구(T4)가 가장 높았다. Ca과 P의 이용률 또한 T4가 가장 높고 T3와 T5가 다음으로 높았다. Phytase 처리구 (T3, T4와 T5) 들은 Mg, Fe와 Zn의 이용률을 크게 증가시켰다. Ca, P, Mg, Fe와 Zn의 배설량은 microbial phytase가 가장 적었다. 혈청내 Ca과 Mg의 함량은 低 NPP에서 가장 높았고, 경골내 조회분 함량은 T2와 T3가 T1, T4와 T5보다 낮았다. 그러나 경골내 Ca 함량은 T1과 T2가 다른 처리구들보다 높았다. 경골내 P과 Mg의 함량은 T1구가 가장 높았다. 결론적으로 밀기울을 통한 식물성 phytase의 공급은 P 이용률 향상에 효과적이었다. 수침 후 건조처리하는 것은 이용률 향상에 도움이 되지 않았다.

• Asian-Australasian Journal of Animal Sciences
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• 제16권2호
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• pp.239-247
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• 2003

Two experiments were conducted with broilers to investigate the feasibility of microbial phytase replacing partial inorganic phosphorus supplementation and the synergistic effects of xylanase (320 FTU/kg) supplementation alone or in combination with phytase (750 U/kg) replacing 0.08% dietary inorganic phosphorus, on the growth performance and utilization of nutrients in broilers fed wheat-based diets. In Experiment 1, 540 broilers were fed five diets for 6 weeks. Diets C0 and C1 were corn-based diets and 0.08% inorganic P supplementation was replaced with 750 U phytase/kg feed in Diet C1. Diets W0, W1 and W2 were wheat-based diets supplemented with microbial phytase 0, 750, 750 U/kg feed and 0, 0.08% and 0.16% dietary inorganic P were replaced, respectively. In Experiment 2, 432 broilers were divided into four treatments to determine the synergistic effects of supplemental xylanase and phytase replacing 0.08% inorganic P. Four experimental diets were arranged according to a $2{\times}2$ factorial design. The results indicated that addition of phytase increased the digestibility of phytic P by 31.0 to 55%, dramatically decreased the excretion of phytic P and total P by 31.6 to 55.0% and 13.8 to 32.9%, respectively (p<0.01). It is feasible to completely replace 0.08% inorganic phosphorus supplementation with microbial phytase 750 U/kg in corn- or wheat-based diets for broilers. Addition of xylanase alone or in combination with phytase replacing 0.08% dietary inorganic P, increased body weight gain and feed utilization efficiency of broilers fed wheat-based diets (p<0.10) and decreased overall mortality (p<0.10). In the groups of birds supplementing xylanase 320 FTU/kg feed, a marked elevation of the dietary AME was observed (p<0.05). Addition of phytase replacing 0.08% dietary inorganic phosphorus, concurrently with xylanase supplementation had additive effects on the apparent digestibility of dietary phytic P and overall feed conversion ratio (p<0.05).

• Animal Bioscience
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• 제35권6호
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• pp.892-901
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• 2022

Objective: This study was performed to investigate the potential effect of wheat phytase on long-chain inorganic polyphosphate (polyP)-mediated interleukin 8 (IL-8) signaling in an intestinal epithelial cell line, HT-29 cells. Methods: Cell viability and the release of the pro-inflammatory cytokine IL-8 in HT-29 cells exposed to polyP1150 (average of 1,150 phosphate residues) treated with or without wheat phytase were measured by the EZ-CYTOX kit and the IL-8 ELISA kit, respectively. Also, the activation of cellular inflammatory factors NF-κB and MAPK (p38 and ERK 1/2) in HT-29 cells was investigated using ELISA kits. Results: PolyP1150 negatively affected the viability of HT-29 cells in a dose-dependent manner. However, 100 mM polyP1150 dephosphorylated by wheat phytase increased cell viability by 1.4-fold over that of the intact substrate. Moreover, the 24 h exposure of cells to enzyme-treated 50 mM polyP1150 reduced the secretion of IL-8 and the activation of NF-κB by 9% and 19%, respectively, compared to the intact substrate. PolyP1150 (25 and 50 mM) dephosphorylated by the enzyme induced the activation of p38 MAPK via phosphorylation to 2.3 and 1.4-fold, respectively, compared to intact substrate, even though it had little effect on the expression of ERK 1/2 via phosphorylation. Conclusion: Wheat phytase could attenuate polyP1150-induced IL-8 release in HT-29 cells through NF-κB, independent of MAP kinases p38 and ERK. Thus, wheat phytase may alleviate inflammatory responses including hypercytokinemia caused by bacterial polyP infection in animals. Therefore, wheat phytase has the potential as an anti-inflammatory therapeutic supplement in animal husbandry.

• Asian-Australasian Journal of Animal Sciences
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• 제15권7호
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• pp.1036-1039
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• 2002

The effect of germination on phytase activity in wheat NEAU123, triticale5305 and rye2 was studied in the present study. Germination significantly increased phytase activity by 2.04 times for wheat NEAU123 (3 d), 1.82 times for triticale 5305 (1 d) and 2.45 times for rye2 (1 d), respectively. It was safe for phytase in fresh malts kilned for 2 h at $40^{\circ}C$. Phytase in cereal seeds had strong heat stability. There was no loss of phytase activity in cereal seeds heated at $70^{\circ}C$ for 1 h, a little loss (${\leq}$5.46%) at $80^{\circ}C$ or $90^{\circ}C$. Even heated at $100^{\circ}C$, the phytase activity in wheat NEAU123, triticale5305 and rye2 remained 89.47%, 86.44% and 104.64%, respectively.

• 한국식품과학회지
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• 제54권4호
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• pp.422-430
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• 2022

본 연구는 통밀가루에 다량 함유되어 있는 미네랄의 체내 이용률을 높여 기능성 소재로의 응용가능성을 조사하고자 phytase와 cellulase를 처리한 통밀가루 미네랄의 체내이용률에 대하여 연구하였다. 이에 통밀가루의 최적 효소 처리 조건을 확립하기 위하여 효소농도별에 따른 이화학적 특성과 phytate 및 식이섬유소의 함량을 분석하였으며, 미네랄 강화 소재로의 이용 가능성을 확인하기 위해 머핀을 제조 후 품질특성 및 미네랄의 체내 이용률을 확인하였다. 통밀가루 건중량의 2, 10 및 20%로 2시간 동안 반응시킨 phytase와 cellulase 단일 효소처리한 시료와 20%의 cellulase/phytase를 5:5 비율로 복합 처리한 시료를 제조하였다. 효소 처리 통밀가루의 총 식이섬유소의 함량은 효소 농도에 따라 감소하였고 특히, 복합 처리군에서 가장 크게 감소하였다. 더불어 phytate 함량은 모든 효소에서 농도가 증가할수록 감소하였다. 효소처리 농도가 증가할수록 철분, 칼슘, 아연의 체내 이용률이 증가하였는데 특히, 철분의 체내 이용률은 cellulase를 처리하였을 때 약 3.3배로 가장 크게 증가하였고, 칼슘은 cellulase와 phytase를 복합으로 처리하였을 때 약 5.1배로 가장 증가하였고, 아연은 phytase를 처리하였을 때 약 2.8배로 가장 크게 증가하였다. 미네랄의 이용률이 가장 높았던 20% phytase, cellulase, cellulase/phytase 처리한 통밀가루를 이용하여 대체 비율(0, 25, 50, 75 및 100%)에 따른 머핀의 품질특성 및 미네랄 체내 이용률을 확인한 결과, 효소 처리 통밀가루로 제조한 머핀의 부피는 무처리군으로 제조한 머핀보다 대체율이 높을수록 증가하였으며, 조직감의 경우 hardness는 감소하였고, springiness와 cohesiveness는 증가하는 경향을 보였다. 그리고 효소처리 통밀가루로 제조한 머핀에서 대체 비율이 증가할수록 철분, 칼슘, 아연의 체내 이용률은 무처리군 머핀보다 증가하였는데, Cellulase 처리군의 철분 체내 이용률은 2.3배, cellulase/phytase 복합 처리군에서 칼슘은 3.7배, phytase 처리군에서 아연은 2.3배로 가장 크게 증가하였다. 이상의 결과로부터 통밀가루에 phytase와 cellulase를 처리한 후 미네랄의 이용률이 증가함을 확인하였고, 효소처리 통밀가루로 제조한 머핀의 품질의 특성이 향상되어 미네랄 이용률 증진 소재로 응용이 가능함을 확인하였다.

• Animal Bioscience
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• 제34권8호
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• pp.1365-1374
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• 2021

Objective: This study was conducted to investigate the synergistic effect of exogenous multienzyme and phytase on growth performance, nutrients digestibility, blood metabolites, intestinal microflora, and morphology in broilers fed corn-wheat-soybean meal diets. Methods: A 2×2 factorial design was used in this study. Four dietary treatments consisted of i) basal diets (corn-wheat-soybean meal based diets without multi-enzyme and phytase), ii) basal diets with phytase (0.05%), iii) basal diets with exogenous multi-enzyme (0.05%), and iv) basal diets with exogenous multi-enzyme including phytase (0.05%). A total of 480 broiler chickens (Ross 308 - one day old) were weighed and allotted to thirty-two cages (15 birds per cage), and chicks were randomly allocated to four dietary treatments. Results: The body weight gain and feed conversion rate were improved by supplementation of exogenous multi-enzyme containing phytase during the finisher period (p<0.05). The birds fed diets with exogenous multi-enzyme containing phytase had a significantly greater digestibility of dry matter, gross energy, crude protein, calcium, and phosphorus compared with birds fed non-supplemented diets (p<0.05). The chickens fed diets with exogenous multi-enzyme containing phytase showed a higher concentration of Ca and P in the serum (p<0.05). The population of Lactobacillus spp., Escherichia coli, and Clostridium were not affected in the ileum and cecum of chickens fed enzyme-supplemented diets. The dietary supplemental exogenous multi-enzyme containing phytase showed a significant improvement in villus height, crypt depth, and villus height and crypt depth ratio, compared to basal diets or dietary supplemental phytase (p<0.05). Conclusion: The supplementation of the exogenous multi-enzyme containing phytase synergistically improved the growth performance, nutrients digestibility, and villus height of the small intestine of broiler chickens fed a corn-wheat-soybean meal based diets.

• Asian-Australasian Journal of Animal Sciences
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• 제16권3호
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• pp.394-402
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• 2003

Individual and combined supplementation of phosphorus-adequate, wheat-based broiler diets with exogenous phytase and xylanase was evaluated in three experiments. The effects of the enzyme combination in lysine-eficient diets containing wheat and sorghum were more pronounced than those of the individual feed enzymes. The inclusion of phytase plus xylanase improved (p<0.05) weight gains (7.3%) and feed efficiency (7.0%) of broilers (7-28 days post-hatch) and apparent metabolisable energy (AME) by 0.76 MJ/kg DM. Phytase plus xylanase increased (p<0.05) the overall, apparent ileal digestibility of amino acids by 4.5% (0.781 to 0.816); this was greater than the responses to either phytase (3.6%; 0.781 to 0.809) or xylanase (0.7%; 0.781 to 0.784). Absolute increases in amino acid digestibility with the combination exceeded the sum of the individual increases generated by phytase and xylanase for alanine, aspartic acid, glutamic acid, glycine, histidine, isoleucine, phenylalanine, threonine, tyrosine and valine. These synergistic responses may have resulted from phytase and xylanase having complementary modes of action for enhancing amino acid digestibilities and/or facilitating substrate access. The two remaining experiments were almost identical except wheat used in Experiment 2 had a higher phytate concentration and a lower estimated AME content than wheat used in Experiment 3. Individually, phytase and xylanase were generally more effective in Experiment 2, which probably reflects the higher dietary substrate levels present. Phytase plus xylanase increased (p<0.05) gains (15.4%) and feed efficiency (7.0%) of broiler chicks from 4-24 days post-hatch in Experiment 2; whereas, in Experiment 3, the combination increased (p<0.05) growth to a lesser extent (5.6%) and had no effect on feed efficiency. This difference in performance responses appeared to be 'rotein driven'as the combination increased (p<0.05) nitrogen retention in Experiment 2 but not in Experiment 3; whereas phytase plus xylanase significantly increased AME in both experiments. In Experiments 2 and 3 the combined inclusion levels of phytase and xylanase were lower that the individual additions, which demonstrates the benefits of simultaneously including phytase and xylanase in wheat-based poultry diets.