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1.
Org Biomol Chem ; 22(8): 1714-1720, 2024 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-38318943

RESUMO

Ratiometric fluorescent assays have a built-in correction factor which enhances assay accuracy and reliability. We have developed fluorescent ratiometric supramolecular tandem assays for phosphatase and phytase enzymes using a mixture of three molecular components. One of the molecules is a tetra-cationic fluorescence quencher called CalixPyr which can bind and quench the polyanionic pyrene fluorophore, CMP, that emits at 430 nm. Polyphosphates can disrupt the CMP/CalixPyr complex and alter the fluorescence intensity (responsive signal). CalixPyr has no effect on the fluorescence emission of cationic pentamethine cyanine fluorophore, cCy5, which emits at 665 nm and acts as a non-responsive reference signal. The continuous ratiometric fluorescent assay for alkaline phosphatase monitored hydrolytic consumption of adenosine triphosphate (ATP). The continuous ratiometric fluorescent assay for phytase activity monitored hydrolytic consumption of phytate. With further development this latter assay may be useful for high throughput assessment of phytase activity in individual batches of fortified animal feed. It is likely that the three-molecule mixture (CMP, CalixPyr, cCy5) can become a general assay platform for other enzymes that catalyse addition/removal of phosphate groups from appropriate molecular substrates.


Assuntos
6-Fitase , Monoéster Fosfórico Hidrolases , Animais , 6-Fitase/metabolismo , Reprodutibilidade dos Testes , Fosfatase Alcalina/metabolismo , Hidrólise , Corantes Fluorescentes/química
2.
Poult Sci ; 103(4): 103511, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38340661

RESUMO

An experiment was conducted to evaluate the effects of phytase in calcium (Ca) and available phosphorous (avP)-reduced diet on growth performance, body composition, bone health, and intestinal integrity of broilers challenged with Eimeria maxima and Eimeria acervulina. A total of 672 14-day-old male broilers were allocated to a 2 × 4 factorial arrangement with 6 replicates per treatment and 14 birds per replicate. Two factors were Eimeria challenge and 4 dietary treatments: 1) a positive control (PC; 0.84% Ca and 0.42% avP); 2) a negative control (NC; 0.74% Ca and 0.27% avP); 3) NC + 500 FTU/Kg of phytase (NC + 500PHY); and 4) NC + 1,500 FTU/Kg of phytase (NC + 1500PHY). On d 14, birds in the Eimeria-challenged groups received a solution containing 15,000 sporulated oocysts of E. maxima and 75,000 sporulated oocysts of E. acervulina via oral gavage. At 5 d postinoculation (DPI), the challenged birds showed a higher (P < 0.01) FITC-d level than the unchallenged birds. While the permeability of the NC group did not differ from the PC group, the phytase supplementation groups (NC + 500PHY and NC + 1500PHY) showed lower (P < 0.05) serum FITC-d levels compared to the NC group. Interaction effects (P < 0.05) of Eimeria challenge and dietary treatments on feed intake (FI), mucin-2 (MUC2) gene expression, bone ash concentration, and mineral apposition rate (MAR) were observed. On 0 to 6 and 0 to 9 DPI, Eimeria challenge decreased (P < 0.01) body weight (BW), body weight gain (BWG), FI, bone mineral density (BMD), bone mineral content (BMC), bone area, fat free bone weight (FFBW), bone ash weight, bone ash percentage and bone ash concentration; and it showed a higher FCR (P < 0.01) compared to the unchallenged group. The reduction Ca and avP in the diet (NC) did not exert adverse effects on all parameters in birds, and supplementing phytase at levels of 500 or 1,500 FTU/Kg improved body composition, bone mineralization, and intestinal permeability, with the higher dose of 1,500 FTU/Kg showing more pronounced enhancements. There was an observed increase in FI (P < 0.01) when phytase was supplemented at 1,500 FTU/Kg during 0 to 6 DPI. In conclusion, results from the current study suggest that dietary nutrients, such as Ca and avP, can be moderately reduced with the supplementation of phytase, particularly in birds infected with Eimeria spp., which has the potential to save feed cost without compromising growth performance, bone health, and intestinal integrity of broilers.


Assuntos
6-Fitase , Eimeria , Minerais , Masculino , Animais , Cálcio/metabolismo , Fósforo , Galinhas , Densidade Óssea , Fluoresceína-5-Isotiocianato , Dieta/veterinária , Cálcio da Dieta/metabolismo , Suplementos Nutricionais/análise , Aumento de Peso , Composição Corporal , Ração Animal/análise
3.
Poult Sci ; 103(4): 103520, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38364607

RESUMO

Apparent ileal digestibility (AID) of P, apparent total tract retention (ATTR) of P, and phytic acid disappearance in canola meal were evaluated in the presence of increasing levels of exogenous phytase. In Experiment 1, a precision-fed rooster assay was used to determine phytic acid (myo-inositol 1,2,3,4,5,6-hexakis; InsP6) and inositol phosphate (InsP6-3; InsP-P) disappearance in conventional and cecectomized Leghorn roosters. Roosters were crop intubated with 25 g of canola meal mixed with 0, 500, 1,000, or 2,000 FTU/kg of exogenous phytase. In Experiment 2, InsP6 and InsP-P disappearance and AID and ATTR of P were determined using ad libitum-fed broiler chickens. Treatments consisted of semi-purified diets containing 45% canola meal as the sole source of P. Phytase was added to increase phytase activity by 0, 500, 1,000, or 2,000 FTU/kg. Experiments contained 6 replicates per treatment. Canola meal contained a high phytase activity (1,630 FTU/kg as-fed) due to contamination with a commercially available phytase at the feed mill from which the canola meal was sourced. In Experiment 1 with precision-fed roosters, there was no effect (P > 0.05) of phytase or bird type on InsP6 and InsP-P disappearance; however, phytase linearly reduced (P < 0.05) InsP3 concentrations in excreta. In Experiment 2 with ad libitum-fed chickens, phytase linearly increased (P < 0.05) ileal InsP6 and InsP-P disappearance, and phytase had a quadratic effect (P < 0.05) on excreta InsP6 and InsP-P disappearance. Increasing dietary phytase activity resulted in a linear increase (P < 0.05) in AID of P and phytase had a quadratic effect (P < 0.05) on ATTR of P. In conclusion, titration of high levels of phytase (1,600 to 3,600 FTU/kg as-fed) reduced InsP3 concentrations in precision-fed roosters but did not affect overall phytic acid hydrolysis, which was 78% or greater for all treatments; however, increasing the total phytase activity from 700 to 2,700 FTU in ad libitum-fed broiler chickens increased phytic acid disappearance and P digestibility.


Assuntos
6-Fitase , Brassica napus , Animais , Masculino , Galinhas , Ácido Fítico , Digestão , Ração Animal/análise , Suplementos Nutricionais , Dieta/veterinária , Fenômenos Fisiológicos da Nutrição Animal
4.
Vet Med Sci ; 10(2): e1379, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38358071

RESUMO

BACKGROUND: As a major part of the cost of broiler farms is related to their feed, breeders are looking for diets at a reasonable price. One of these affordable ingredients is sesame meal (SM), which can be used to replace soybean meal (SBM) in diets. OBJECTIVES: This study aims to investigate the effect of replacing SBM with SM in diets with different levels of calcium (Ca) and phytase (Phy) during the grower (11-24 days), finisher (25-42 days) and whole phases (11-42 days) on the growth performance, carcass characteristics and blood serum parameters of broiler chickens. METHODS: A total of 600 1-day-old Ross 308 broiler male chicks were randomly dispensed to 12 dietary treatments with five replicates (10 birds per replicate) based on a completely randomized design in a factorial arrangement of 3 × 2 × 2 with 3 levels of SM (0%, 10% and 20%), 2 levels of Ca (standard (0.87% and 0.79% for grower and finisher phases, respectively) and 0.2% higher than the standard amount) and 2 levels of Phy (0 and 700 FTU/kg diet). RESULTS: During the study, the significant effects of SM, Ca and Phy on the daily average body weight gain (DAWG) and daily feed intake (DFI) were observed, whereas for the feed conversion ratio, only the effect of SM was significant (p < 0.05). During the finisher and whole phases, the SM and Ca levels influenced the DAWG and DFI, whereas the interaction between Phy and Ca was also significant for DAWG (p < 0.05). The main effects of SM and Ca on the relative weights of breast, heart and liver were significant (p < 0.05). Application of 20% of SM significantly reduced serum Ca concentration (p < 0.05). CONCLUSION: Generally, the inclusion of more than 10% of SM in the broilers diet is not recommended. In addition, the use of Phy and Ca levels 0.2% higher than standard in broilers diet could improve the birds' performance.


Assuntos
6-Fitase , Sesamum , Animais , Masculino , Cálcio , Galinhas , Dieta/veterinária , Farinha , Aumento de Peso
5.
Environ Sci Technol ; 58(8): 3858-3868, 2024 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-38356137

RESUMO

Phytate, the principal P storage in plant seeds, is also an important organic P in soils, but it is unavailable for plant uptake. However, the As-hyperaccumulator Pteris vittata can effectively utilize soluble Na-phytate, while its ability to utilize insoluble Ca/Fe-phytate is unclear. Here, we investigated phytate uptake and the underlying mechanisms based on the phytase activity, nutrient uptake, and expression of genes involved in As metabolisms. P. vittata plants were cultivated hydroponically in 0.2-strength Hoagland nutrient solution containing 50 µM As and 0.2 mM Na/Ca/Fe-phytate, with 0.2 mM soluble-P as the control. As the sole P source, all three phytates supported P. vittata growth, with its biomass being 3.2-4.1 g plant-1 and Ca/Fe-phytate being 19-29% more effective than Na-phytate. Phytate supplied soluble P to P. vittata probably via phytase hydrolysis, which was supported by 0.4-0.7 nmol P min-1 g-1 root fresh weight day-1 phytase activity in its root exudates, with 29-545 µM phytate-P being released into the growth media. Besides, compared to Na-phytate, Ca/Fe-phytate enhanced the As contents by 102-140% to 657-781 mg kg-1 in P. vittata roots and by 43-86% to 1109-1447 mg kg-1 in the fronds, which was accompanied by 21-108% increase in Ca and Fe uptake. The increased plant As is probably attributed to 1.3-2.6 fold upregulation of P transporters PvPht1;3/4 for root As uptake, and 1.8-4.3 fold upregulation of arsenite antiporters PvACR3/3;1/3;3 for As translocation to and As sequestration into the fronds. This is the first report to show that, besides soluble Na-phytate, P. vittata can also effectively utilize insoluble Ca/Fe-phytate as the sole P source, which sheds light onto improving its application in phytoremediation of As-contaminated sites.


Assuntos
6-Fitase , Arsênio , Pteris , Poluentes do Solo , 6-Fitase/metabolismo , Pteris/metabolismo , Ácido Fítico/metabolismo , Raízes de Plantas/química , Raízes de Plantas/metabolismo , Biodegradação Ambiental
6.
J Microbiol Methods ; 217-218: 106890, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38272400

RESUMO

Research into phytase production is useful for improving the efficiency of animal production, reducing environmental impact, and contributing to the development of sustainable and efficient animal production systems. This study aims to investigate the potential of yeast strains for phytase biosynthesis in nutrient media. Phytase is a phosphomonoesterase (E.C 3.1.3.8) catalyzing in a ladder-like manner the dephosphorylation of phytic acid and its salts, with various resulting myo-inositol phosphates and phosphoric acid. Yeasts of the genera Saccharomyces, Zygosaccharomyces, Candida, and Pichia were evaluated in a two-step screening procedure for phytase production. One hundred and eighteen strains were screened in the first stage, which was conducted on four types of solid culture media containing calcium phytate as the selected background. On PSM medium, many strains were found to form halos as early as the 24th hour of development. Several strains with significant potential for enzyme production were evaluated in the second step of the screening. It was conducted in a liquid culture medium. In conclusion, the strain C. melibiosica 2491 was selected for further studies when cultured in a YPglu culture medium. Further research will focus on finding suitable conditions that increase the biosynthesis of the enzyme, which is of significant technological and practical interest for animal nutrition.


Assuntos
6-Fitase , Saccharomyces cerevisiae , Animais , Pichia , Candida , Fosfatos de Inositol , Ácido Fítico
7.
Sci Rep ; 14(1): 612, 2024 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-38182617

RESUMO

The occurrence of organically bound phosphorus (P) as phytate in plant-based feeding material is a challenge for livestock farming due to limited utilization during the digestion by the animal. Its excretion into the environment through the manure pathway, poses a challenge, due to increased eutrophication and restrictions for P. Hence, while the routine supplementation of phytase enzymes in monogastric diets is common practice, metabolically triggering endogenous plant enzymes by wet-treatment prior to feeding can also lead to a better utilization of phytate bound P and increased digestibility by the animal. Nonetheless, traditional quantification of residual phytate content in plant material is both labor- and chemical-intense. The aim of this study is, therefore, to predict the remaining phytate content during wet-treatment through a straightforward and flexible methodological approach based on real-time analysis. For this, rye bran is used as a model substrate. A partial least squares regression algorithm relates the infrared spectra to the concentrations and predict the amount of P species that are transferred from the bran matrix to the liquid phase. By applying a mass balance for P and considering the effect of water compression, the amount of residual phytate content in rye bran at different time points of wet-treatment is determined. Results are compared to wet chemical methods, resulting in a RMSEP of 0.28 gphytate∙100 gbran-1. In addition, the study demonstrates the feasibility of this approach and provides insights into phytate degradation in plant residuals. The method holds the potential for further applications for the screening and investigation of feed material conditioning and also offers the possibility to employ various real-time analytical techniques for assessing phytate remnants in biological samples during wet-treatment.


Assuntos
6-Fitase , Ácido Fítico , Animais , Alimentos , Agricultura , Algoritmos
8.
Microb Cell Fact ; 23(1): 9, 2024 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-38172920

RESUMO

BACKGROUND: Existing plasmid systems offer a fundamental foundation for gene expression in Cupriavidus necator; however, their applicability is constrained by the limitations of conjugation. Low segregational stabilities and plasmid copy numbers, particularly in the absence of selection pressure, pose challenges. Phytases, recognized for their widespread application as supplements in animal feed to enhance phosphate availability, present an intriguing prospect for heterologous production in C. necator. The establishment of stable, high-copy number plasmid that can be electroporated would support the utilization of C. necator for the production of single-cell protein from CO2. RESULTS: In this study, we introduce a novel class of expression plasmids specifically designed for electroporation. These plasmids contain partitioning systems to boost segregation stability, eliminating the need for selection pressure. As a proof of concept, we successfully produced Escherichia coli derived AppA phytase in C. necator H16 PHB- 4 using these improved plasmids. Expression was directed by seven distinct promoters, encompassing the constitutive j5 promoter, hydrogenase promoters, and those governing the Calvin-Benson-Bassham cycle. The phytase activities observed in recombinant C. necator H16 strains ranged from 2 to 50 U/mg of total protein, contingent upon the choice of promoter and the mode of cell cultivation - heterotrophic or autotrophic. Further, an upscaling experiment conducted in a 1 l fed-batch gas fermentation system resulted in the attainment of the theoretical biomass. Phytase activity reached levels of up to 22 U/ml. CONCLUSION: The new expression system presented in this study offers a highly efficient platform for protein production and a wide array of synthetic biology applications. It incorporates robust promoters that exhibit either constitutive activity or can be selectively activated when cells transition from heterotrophic to autotrophic growth. This versatility makes it a powerful tool for tailored gene expression. Moreover, the potential to generate active phytases within C. necator H16 holds promising implications for the valorization of CO2 in the feed industry.


Assuntos
6-Fitase , Cupriavidus necator , Cupriavidus necator/metabolismo , 6-Fitase/genética , 6-Fitase/metabolismo , Dióxido de Carbono/metabolismo , Plasmídeos/genética , Regiões Promotoras Genéticas , Escherichia coli/genética , Escherichia coli/metabolismo
9.
Vet Med Sci ; 10(1): e1344, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38227704

RESUMO

BACKGROUND: Enzyme combinations, particularly phytase (PHY) with various carbohydrases and proteases, are utilized in commercial broiler production to enhance nutrient and energy bioavailability. OBJECTIVE: A feeding study was undertaken to determine whether the efficiency of an Escherichia coli-derived PHY and a feed enzyme complex (FEC) derived from Bacillus spp. containing carbohydrase and protease as main activities in broiler chickens is dependent on diet quality. A total of 900 male one-day-old broiler chickens (Ross 308) were assigned to a 2 × 3 factorial arrangement of the treatments with 2 different nutrient density diets, standard nutrient diet (SN diet) and a low-nutrient diet (LN diet; -100 kcal/kg for AMEn and -5% for crude protein [CP] and limiting amino acids), and 3 enzyme treatments (control [no enzymes], PHY and PHY + FEC). Each treatment group was composed of 6 replicates of 25 birds each. RESULTS: The LN diet caused a decrease in performance index, tibia length and diameter, tibia calcium content and jejunal villus surface area (VSA). The interaction effects between diet and enzyme supplementation were observed (p < 0.05) on overall average daily gain (ADG), performance index, tibia ash content and jejunal villus height (VH) and VSA, with the favourable benefits of PHY + FEC treatment being more pronounced in the LN diets. Regardless of dietary nutrient density, supplementation with PHY alone or combined with FEC enhanced (p < 0.05) final body weight, overall ADG and jejunal villus height (VH)/crypt depth, with the highest values observed in the PHY + FEC group. The PHY + FEC treatment also improved (p < 0.05) overall feed conversion ratio, apparent ileal digestibility of dry matter, organic matter, CP, and energy, and tibia phosphorus content compared to the control treatment. CONCLUSIONS: The results indicate that the simultaneous addition of PHY and FEC to the LN diets improved the growth rate, bone mineralization and gut morphology.


Assuntos
6-Fitase , Suplementos Nutricionais , Glicosídeo Hidrolases , Animais , Masculino , Galinhas , 6-Fitase/metabolismo , 6-Fitase/farmacologia , Peptídeo Hidrolases/farmacologia , Calcificação Fisiológica , Escherichia coli , Digestão , Dieta/veterinária , Nutrientes , Ração Animal/análise
10.
Plant Cell Environ ; 47(2): 600-610, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37885374

RESUMO

Ectomycorrhizal fungi (ECMFs) that are involved in phosphorus mobilisation and turnover have limited ability to mineralise phytate alone. The endofungal bacteria in the ectomycorrhizal fruiting body may contribute to achieving this ecological function of ECMFs. We investigated the synergistic effect and mechanisms of endofungal bacteria and ECMF Suillus grevillea on phytate mineralisation. The results showed that soluble phosphorus content in the combined system of endofungal bacterium Cedecea lapagei and S. grevillea was 1.8 times higher than the sum of C. lapagei and S. grevillea alone treatment under the phytate mineralisation experiment. The S. grevillea could first chemotactically assist C. lapagei in adhering to the surface of S. grevillea. Then, the mineralisation of phytate was synergistically promoted by increasing the biomass of C. lapagei and the phosphatase and phytase activities of S. grevillea. The expression of genes related to chemotaxis, colonisation, and proliferation of C. lapagei and genes related to phosphatase and phytase activity of S. grevillea was also significantly upregulated. Furthermore, in the pot experiment, we verified that there might exist a ternary symbiotic system in the natural forest in which endofungal bacteria and ECMFs could synergistically promote phytate uptake in the plant Pinus massoniana via the ectomycorrhizal system.


Assuntos
6-Fitase , Micorrizas , Pinus , Micorrizas/metabolismo , Pinus/metabolismo , Fósforo/metabolismo , 6-Fitase/metabolismo , Ácido Fítico/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Bactérias/metabolismo
11.
Poult Sci ; 103(2): 103296, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38042042

RESUMO

A total of 360 Ross 708 male broiler chicks were used in an 8 treatment and 9 replicate cage study to explore the influence of day-old chick weight on the efficacy of exogenous phytase. Treatments were arranged as a 2 × 4 factorial with the factors being diet (a positive (PC) and negative control (NC) varying in nutrient density fed without or with 2 concentrations of exogenous phytase) and chick weight (light; <38.5 g or heavy; >42 g). Chicks were sourced from the same breeder flock, with light and heavy chicks being selected from the naturally occurring heterogeneity in the population. The diets were corn-soybean meal based and the PC was formulated to meet the nutrient requirements of male broiler chicks. The NC was formulated to contain 120 kcal/kg, 0.5, 0.18, and 0.18% less apparent metabolizable energy (AME), crude protein (CP), calcium (Ca), and phosphorus (P), respectively, than the PC. Amino acid (AA) density in the NC was also reduced in line with the reduction in CP and the manufacturers' recommendations for the effect of phytase on amino acid digestibility. Phytase at either 1,000 FYT/kg or 3,000 FYT/kg was added only to the NC to create a total of 4 diets. Water and mash feed were available ad libitum and were offered to 8 replicate cages, each containing 5 chicks. The experiment was conducted over a period of 3 wk and diets were offered in 2 phases (starter from d 1 to 10 and grower from d 11 to 21). Growth performance was monitored at the end of each diet phase and on d 21 ileal digesta were collected for estimation of apparent digestibility of energy (DE), nitrogen (N), Ca, P, dry matter (DM), and AA. There were no statistically significant interactions between diet and day-old chick weight for any of the measured parameters. Light chicks had significantly lower weight gain (approx. 5%) at both d 10 and d 21 compared with heavy chicks. This effect was principally associated with reduced feed intake and there was no significant effect of chick weight on feed conversion ratio (FCR). Chick weight had no effect on ileal nutrient digestibility. The reduction in nutrient density from the PC to the NC generated a significant reduction in weight gain (around 12%) and a significant increase in FCR (1.68 vs. 1.83). This effect was associated with a significant reduction in ileal DE (approximately 150 kcal/kg) and in the digestibility of several AA. Exogenous phytase significantly increased weight gain, reduced FCR and generated a significant increase in the ileal digestibility of energy, N, P, and several AA. Although chick weight and diet did not interact statistically, heavy chicks benefited more than light chicks from high doses of exogenous phytase across almost all measured end points which was confirmed by regression analysis. In conclusion, light chicks have inferior performance outcomes than heavy chicks principally because of reduced feed intake, but putatively not in digestive capacity per se. Exogenous phytase is effective in improving performance and nutrient digestibility in nutrient deficient diets. The effect of chick weight per se, and also breeder flock age, on the utility of supra-nutritional inclusion concentrations of exogenous phytase warrants further study.


Assuntos
6-Fitase , Galinhas , Animais , Masculino , Galinhas/fisiologia , Ração Animal/análise , Dieta/veterinária , Cálcio da Dieta/metabolismo , Aumento de Peso , Aminoácidos/metabolismo , Digestão , Fenômenos Fisiológicos da Nutrição Animal , Suplementos Nutricionais/análise
12.
Bioprocess Biosyst Eng ; 47(1): 39-55, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37962643

RESUMO

Phytase enzyme found in plants, animals, and microorganisms is mainly involved in catalyzing the systematic removal of a phosphate group from phytic acid. Enzyme immobilization is one of the cost-effective methods for the wide usage of enzymes in the industrial sector. This paper reports the covalent immobilization of phytase on glutaraldehyde-activated aluminum oxide beads. The immobilization yield, efficiency, and activation energy were found to be 47.8%, 71.5%, and 15.78 J/mol, respectively. The bound enzyme displayed a shift in pH optima from 5.5 to 4.5, which is more beneficial to increase digestibility in comparison with the free enzyme. Immobilized phytase retained 42.60% of its activity after 1.0 h incubation at 80 °C, whereas free enzyme retained only 4.20% of its activity. Thermodynami increase in half-lives, D-values, enthalpy and free energy change after covalent immobilization could be credited to the enhanced stability. Immobilized phytase could be reused for five consecutive cycles retaining 51% of its initial activity with sodium phytate. The immobilized phytase was also found effective to hydrolyze the soybean meal, thus increasing the digestibility of poultry feed. The hydrolyzing reaction of soybean meal was carried out for six consecutive cycles and immobilized phytase retained nearly 50% of activity till the fifth cycle. The amount of phosphorus released after treatment with immobilized phytase was far higher than that from free phytase. Immobilization on this support is significant, as this support can sustain high mechanical resistance at high pH and temperature. This considerable stability and reusability of the bound enzyme may be advantageous for its industrial application.


Assuntos
6-Fitase , Aspergillus oryzae , 6-Fitase/química , Aspergillus oryzae/metabolismo , Células Imobilizadas/metabolismo , Farinha , Fosfatos , Ácido Fítico/metabolismo
13.
Poult Sci ; 103(1): 103248, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38006776

RESUMO

The objective of this study was to estimate phosphorus (P) equivalency of exogenous phytase relative to P from monosodium phosphate (MSP) in broiler chickens. To attain this objective, the impact of dietary MSP or exogenous phytase on growth performance, ileal digestibility of P, and bone characteristics in broiler chickens was assessed. Six experimental diets consisting of a P-deficient basal diet based on corn and soybean meal, basal diet plus 0.9 or 1.8 g/kg of inorganic P from MSP, and basal diet plus 500, 1,000, or 2,000 FYT/kg of exogenous phytase. Calcium to P ratio in all diets was maintained at 1.5:1. A total of 576 male broiler chickens (Cobb 500; initial BW = 190 ± 17 g) on d 8 post hatching were allotted to the 6 dietary treatments in a randomized complete block design using BW as a blocking factor. Each dietary treatment contained 8 replicate cages with 12 birds per cage. On d 11 post hatching, 7 birds from each cage were euthanized by CO2 asphyxiation and dissected for the collection of ileal digesta. On d 18, ileal digesta were also collected from the remaining 5 birds in each cage. The left femur and tibia were collected from the bird with the median BW on d 11 and 18 from each cage, and analyzed for bone breaking strength and bone ash. Weight gain and gain-to-feed ratio linearly or quadratically increased (P < 0.05) in every period as the inclusion rate of MSP or phytase increased. Ileal digestibility of P linearly increased (P < 0.05) on d 11 with increasing MSP, but there was no significant effect on d 18. Increasing phytase concentration linearly increased (P < 0.05) ileal digestibility of P on d 11 and 18. Increasing dietary MSP or phytase linearly increased ileal digestible P concentration in diets (P < 0.05). Bone breaking strength and bone ash linearly or quadratically increased (P < 0.05) with increasing inclusion rate of MSP or phytase on d 11 and 18. The equivalency of 1,000 FYT/kg of exogenous phytase based on dietary ileal digestible P concentration was 1.5 and 1.2 g/kg of inorganic P in diets on d 11 and 18, respectively. The current results showed that the supplementation of MSP or exogenous phytase can increase growth performance, ileal P digestibility, bone breaking strength, and bone ash in young broiler chickens.


Assuntos
6-Fitase , Fósforo na Dieta , Animais , Masculino , Fósforo , Fosfatos , Galinhas , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Dieta/veterinária , Digestão , Suplementos Nutricionais
14.
Appl Biochem Biotechnol ; 196(2): 790-803, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37204550

RESUMO

The aims of the study were to degrade the anti-nutritional factors (ANFs) such as phytic acid, glycinin, and ß-conglycinin and improve the values of soybean meal (SBM). Firstly, in this study, a strain PY-4B which exhibited the best enzymatic activities of protease (403.3 ± 17.8 U/mL) and phytase (62.9 ± 2.9 U/mL) was isolated and screened among the isolates. Based on the analysis of physiological and biochemical characteristics and 16S rDNA sequence, the strain PY-4B was identified and named as Pseudomonas PY-4B. Next, Pseudomonas PY-4B was applied to fermentation of SBM. The results showed that the contents of glycinin and ß-conglycinin were decreased by 57-63%, and the phytic acid was remarkably degraded by 62.5% due to the fermentation of SBM by Pseudomonas PY-4B. The degradation of glycinin and ß-conglycinin resulted in increase of contents of water-soluble proteins and amino acids in fermented SBM. Moreover, Pseudomonas PY-4B exhibited no hemolytic activity and slight inhibitory effect on the growth of pathogen Staphylococcus aureus and the wide range of pH tolerance (3 to 9). In summary, our study indicates that isolated strain Pseudomonas PY-4B is a safe and applicable strain and has the ability to effectively degrade the ANFs (phytic acid, glycinin, and ß-conglycinin) in SBM by fermentation.


Assuntos
6-Fitase , 6-Fitase/metabolismo , Peptídeo Hidrolases/metabolismo , Fermentação , Ácido Fítico/metabolismo , Farinha , Endopeptidases/metabolismo , Ração Animal/análise
15.
Poult Sci ; 103(2): 103330, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38113707

RESUMO

Three broiler experiments were conducted to estimate the Ca equivalency of a novel phytase using direct and indirect methods. All 3 experiments employed 4 concentrations of limestone to create 4 reference diets, deficient in nonphytate P, with increasing dietary Ca. Phytase was supplemented to the lowest Ca reference diet at 350, 700, 1,400, or 2,800 FYT/kg in experiment (Exp.) 1 and Exp. 2 and at 500, 1,000, 2,000, or 4,000 FYT/kg in Exp. 3. Broilers were fed from d 8 to 10 and 20 to 24, 19 to 21, or 7 to 10 and 7 to 21 posthatching in Exp. 1, 2, or 3, respectively. Diet did not affect growth performance or tibia ash in Exp. 1. Reducing the dietary Ca linearly (P < 0.05) increased body weight gain (BWG) and feed intake (FI) in Exp. 2 or Exp. 3. Feed conversion ratio (FCR) was decreased (linear or quadratic, P < 0.05) as dietary Ca was reduced in Exp. 2 or Exp. 3 (d 7-21). Tibia ash percent linearly (P < 0.05) decreased as dietary Ca decreased in Exp. 3 but only from d 7 to 21 and phytase increased (linear or quadratic, P < 0.05) FI and BWG, and decreased FCR. In Exp. 1 (d 8-10) and Exp. 2, apparent ileal digestibility (AID), total tract retention, and apparent digested and retained Ca or P increased (linear or quadratic, P < 0.05) as dietary Ca decreased. Phytase increased (linear or quadratic, P < 0.05) AID and apparent digested and retained Ca or P in Exp. 1 or Exp. 2. Due to the nature of the effect of dietary Ca on performance or tibia ash, it was not possible to use the indirect method to estimate the Ca equivalence of phytase in the current experiments. The total and digestible Ca equivalence of phytase could be estimated using the direct method. These experiments highlight challenges to consider when designing experiments to estimate the Ca equivalency for phytase in the future.


Assuntos
6-Fitase , Cálcio , Animais , Carbonato de Cálcio , Galinhas , Tíbia , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Dieta/veterinária , Cálcio da Dieta
16.
Poult Sci ; 103(2): 103360, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38160615

RESUMO

The current study aimed to quantify the digestible calcium (Ca) equivalency of a new phytase (HiPhorius) in broiler chickens. A total of 1,152 male Cobb 500 broiler chickens were used in an experiment in which 8 diets consisting of graded levels of Ca supplied with limestone or graded levels of the phytase were fed. The 8 dietary treatments were based on a corn-soybean meal diet containing 5.1 g/kg of Ca and 5.1 g/kg of phosphorus (P) as negative control (NC); the NC + 1.3, 2.6, or 3.9 g/kg of Ca from limestone; and the NC + 500, 1,000, 2,000, or 4,000 FYT/kg of phytase. Birds were fed the experimental diets for 3 d (from d 7 to 10) or 14 d (from d 7 to 21) to determine apparent ileal digestibility (AID) and apparent total tract retention (ATTR) of dry matter (DM), CP, Ca, and P. In the 10-day-old birds, increasing the levels of Ca decreased the AID of P and Ca (linear, P < 0.05). Increasing the levels of phytase quadratically improved (P < 0.05) the AID of Ca and P. The AID of DM and CP in the younger birds quadratically increased (P < 0.05) as the levels of Ca increased. There were linear and quadratic effects (P < 0.05) of increasing phytase level for the AID of DM and a linear effect (P < 0.05) for the AID of CP. In the 21-day-old birds, increasing Ca levels resulted in a linear decrease (P < 0.05) in the AID of CP and P, and a quadratic decrease (P < 0.05) in the ATTR of CP. Increasing phytase levels linearly and quadratically (P < 0.05) improved the AID and ATTR of CP, P, and the ATTR of Ca. The analyzed phytase activity in the diets supplemented with phytase ranged from 1,520 to 7,661 FYT/kg. The digestible Ca equivalence for dietary phytase at 1,520 to 7,661 FYT/kg ranged from 1.55 to 2.02 g/kg in the 10-day-old birds fed for 3 d and from 0.52 to 0.64 g/kg in 21-day-old birds fed for 14 d. The results showed the reduction in Ca level that could be accommodated by phytase supplementation, which is markedly different between the younger and older birds. Feeding duration influenced the impact of phytase supplementation on Ca and P digestibility, with better efficacy of phytase observed in the 10-day-old birds fed for 3 d. Also, the results showed the extra-phosphoric effects of phytase on the utilization of other essential nutrients such as protein and Ca.


Assuntos
6-Fitase , Carbonato de Cálcio , Animais , Masculino , Cálcio , Galinhas/metabolismo , 6-Fitase/metabolismo , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Digestão , Cálcio da Dieta , Dieta/veterinária , Fósforo , Nutrientes
17.
Poult Sci ; 103(2): 103326, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38157789

RESUMO

Two experiments were conducted to determine the impact of Ca, phytase, sampling time, and age on the digestibility (AID) of Ca and P and the expression of their transporters. Cobb 500 male chicks (N = 600) were used in each experiment and allocated to cages with 10 (Exp 1, 8-11 d) or 5 (Exp 2, 21-24 d) birds/cage and 10 (Exp 1) or 20 (Exp 2) reps/treatment. Treatments were a 2 × 3 factorial arrangement, with low (LOW) or standard (STD) Ca level and 3 phytase (PHY) levels (0, 300, or 3,000 FYT/kg). Ileal digesta were collected at 8, 12, 24, 48, and 72 h, and jejunum tissues at 12, 48, and 72 h after the start of feeding experimental diets. In Exp 1, there was no effect of Ca or phytase on the AID of Ca at 8, 12, or 24 h. Phytase increased the AID of P (P < 0.05) at all time points, and the magnitude was influenced by Ca. At 12 h, the mRNA level of P (NaPi-IIb) and Ca (CaSR) transporters was greatest in the LOW diets without phytase (Ca × PHY, P ≤ 0.06). In Exp 2, the STD diet decreased the AID of Ca and P (P < 0.05) at 8, 24, 48, or 72 h. Phytase increased the AID of Ca (P < 0.05) at 8, 12, and 24 h, and decreased the AID of Ca (quadratic, P < 0.05) in the STD diet (48 h). The AID of P (P < 0.05) increased with phytase at all sampling times. At 48 h, 3,000 FYT/kg decreased (P < 0.05) mRNA expression of NaPi-IIb and Ca transporter ATP2B1 in the STD diet (Ca × PHY, P < 0.05). In conclusion, to avoid adaptation of broilers to Ca and P deficiencies, the optimal time on experimental diets is ≤ 48 h for young broilers and ≤ 24 h in older birds due to up- or down-regulation of Ca and P transporters in response to dietary Ca, P, and phytase.


Assuntos
6-Fitase , Galinhas , Animais , Masculino , Galinhas/fisiologia , 6-Fitase/metabolismo , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Dieta/veterinária , Minerais , Nutrientes , Suplementos Nutricionais , RNA Mensageiro/genética , Digestão
18.
An Acad Bras Cienc ; 95(4): e20191162, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38088696

RESUMO

Male broiler chickens (384), Cobb 500, were housed in metabolic cages to assess the efficacy of phytase in diets with low and high phytate-phosphorus on the performance, bone physical characteristics, tissue and serum mineral deposits. Birds were distributed in four treatments with a 2 x 2 factorial arrangement in a completely randomized block design. Experimental diets based on maize-soybean meal were T1 - diet low phytate-phosphorus; T2 - diet low phytate-phosphorus and phytase (500 FTU/kg); T3 - diet high phytate-phosphorus; T4 - diet high phytate-phosphorus and phytase (500 FTU/kg). Feed intake, body weight, weight gain and feed conversion ratio were assessed. Two left tibias per experimental unit were analyzed for physical characteristics and mineral concentration; a section of skinless breast muscle and blood were collected to measure the concentration of calcium, phosphorus and sodium. Results showed interaction between bone stiffness and serum calcium. The inclusion of phytase in diets with low and high phytate-phosphorus did not alter performance, bone resistance and flexibility, mineral deposits in the tibia and breast muscle, but increased bone stiffness after 22 days of age. It also provided a higher serum calcium rate in broilers fed diets with low phytate-phosphorus up to 32 days of age.


Assuntos
6-Fitase , Fósforo , Animais , Masculino , 6-Fitase/metabolismo , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Cálcio/metabolismo , Galinhas/fisiologia , Dieta/veterinária , Suplementos Nutricionais , Minerais , Fósforo/metabolismo , Ácido Fítico
19.
Int J Mol Sci ; 24(24)2023 Dec 06.
Artigo em Inglês | MEDLINE | ID: mdl-38139020

RESUMO

Organic phosphorus (OP) is an essential component of the soil P cycle, which contributes to barley nutrition after its mineralization into inorganic phosphorus (Pi). However, the dynamics of OP utilization in the barley rhizosphere remain unclear. In this study, phytin was screened out from six OP carriers, which could reflect the difference in OP utilization between a P-inefficient genotype Baudin and a P-efficient genotype CN4027. The phosphorus utilization efficiency (PUE), root morphological traits, and expression of genes associated with P utilization were assessed under P deficiency or phytin treatments. P deficiency resulted in a greater root surface area and thicker roots. In barley fed with phytin as a P carrier, the APase activities of CN4027 were 2-3-fold lower than those of Baudin, while the phytase activities of CN4027 were 2-3-fold higher than those of Baudin. The PUE in CN4027 was mainly enhanced by activating phytase to improve the root absorption and utilization of Pi resulting from OP mineralization, while the PUE in Baudin was mainly enhanced by activating APase to improve the shoot reuse capacity. A phosphate transporter gene HvPHT1;8 regulated P transport from the roots to the shoots, while a purple acid phosphatase (PAP) family gene HvPAPhy_b contributed to the reuse of P in barley.


Assuntos
6-Fitase , Hordeum , Fósforo/metabolismo , Hordeum/genética , Hordeum/metabolismo , 6-Fitase/metabolismo , Ácido Fítico/metabolismo , Genótipo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo
20.
Biomolecules ; 13(12)2023 12 10.
Artigo em Inglês | MEDLINE | ID: mdl-38136641

RESUMO

A study evaluated nine kinetic data and four kinetic parameters related to growth, production of various phytase activities (PEact), and released phosphate ion concentration ([Pi]) from five lactic acid bacteria (LAB) strains cultivated in three types of media: phytate (IP6), milling stage rice bran (MsRB), and whitening stage rice bran (WsRB). Score ranking techniques were used, combining these kinetic data and parameters to select the most suitable LAB strain for each medium across three cultivation time periods (24, 48, and 72 h). In the IP6 medium, Lacticaseibacillus casei TISTR 1500 exhibited statistically significant highest (p ≤ 0.05) normalized summation scores using a 2:1 weighting between kinetic and parameter data sets. This strain also had the statistically highest levels (p ≤ 0.05) of produced phosphate ion concentration ([Pi]) (0.55 g/L) at 72 h and produced extracellular specific phytase activity (ExSp-PEact) (0.278 U/mgprotein) at 48 h. For the MsRB and WsRB media, Lactiplantibacillus plantarum TISTR 877 performed exceptionally well after 72 h of cultivation. It produced ([Pi], ExSp-PEact) pairs of (0.53 g/L, 0.0790 U/mgprotein) in MsRB and (0.85 g/L, 0.0593 U/mgprotein) in WsRB, respectively. Overall, these findings indicate the most promising LAB strains for each medium and cultivation time based on their ability to produce phosphate ions and extracellular specific phytase activity. The selection process utilized a combination of kinetic data and parameter analysis.


Assuntos
6-Fitase , Lactobacillales , Oryza , Fosfatos , Biopolímeros , Ácido Láctico , Íons
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