Application Of Phytase Research Articles

Phosphorus utilization in starter pigs fed high-moisture corn-based liquid diets steeped with phytase

The application of phytase in conventional dry swine diets has been shown to improve P availability and utilization. The effectiveness of phytase may be further improved by steeping feedstuffs with phytase before feeding. A study was conducted to determine the value of steeping high-moisture corn (HMC) with phytase in P-deficient liquid diets for starter pigs. A total of 384 pigs were weaned at 19 to 23 d of age and 6.7 ± 0.1 kg of BW. Pigs were randomly assigned to pens, with 8 barrows and 8 gilts per pen and 5 pens per dietary treatment (only 4 pens for the control treatment). The 5 dietary treatments (all HMC-based 3-phase feeding programs) were 1) negative control with no added phytase, 2 and 3) negative control with phytase added to the HMC to achieve 62.5 or 125 phytase units (FTU)/kg of HMC (DM basis) of phytase added to the HMC and allowed to steep for 24 h before feeding, and 4 and 5) negative control with the same amount of phytase added to the base mix without steeping before feeding. Total P content (88% DM basis) averaged 0.49% in phase I and 0.37% in phase II and III diets. Individual pig BW and per pen ADFI were measured on a weekly basis. Apparent total tract digestibility of DM, OM, CP, P, and Ca were measured using titanium dioxide as an indigestible marker in phase III diets. At the end of the study (7 wk postweaning), 4 pigs from each pen were killed for assessment of body composition, breaking strength and mineral content of metacarpals, total and soluble P content in duodenal digesta, and urinary P content. There was no effect of added phytase on ADG, ADFI, or G:F. The soluble P:total P ratio in duodenal digesta was increased with the addition of phytase (P < 0.05). Steeping HMC with phytase resulted in greater digestibility of DM and CP (P < 0.01). A trend toward increased digestibility of Ca with added phytase was observed (P = 0.07), but there was no effect of dietary treatment on P digestibility. Urinary P content was considerably greater in pigs fed diets with exogenous phytase (P < 0.05). Additional phytase resulted in increased P and Ca content in the empty body (P < 0.05). Metacarpal content of P (P < 0.05) and Ca (P = 0.07) and breaking strength (P < 0.05) were improved with added phytase. Despite a lack of effect on P digestibility, added phytase improved retention of Ca and P in starter pigs fed P-deficient HMC-based liquid diets. There was little benefit from steeping HMC with phytase before feeding.

Biochemical properties, molecular characterizations, functions, and application perspectives of phytases

As a kind of enzyme widely existing in eukaryotic species, especially in grains and oil seeds, phytases play an important role in the degradation of some phosphates containing organic molecules. So far, phytases derived from various species have been successfully used as animal feed additives. It has also been experimentally verified that phytases have a potential use in generating crop germplasm with high phosphorus use efficiency, based on their biochemical role in releasing Pi from the phytate and its derivatives. In this paper, the biochemical properties, molecular characterizations, functions and the potential application perspective of phytases are reviewed and commented on, aiming at the further exploration of the biochemical and molecular characterizations, and promotion of the application of phytases, a kind of important enzyme possessing potential use in animal feeding and creation of high P use crop cultivars, in the future.

Impact of soaking and phytase treatments on phytic acid, calcium, iron and zinc in faba bean fractions

SummaryWith the aim to maximise phytic acid removal and minimise losses of dry matter and minerals (Ca, Fe, Zn) in faba bean, fractions of faba beans (whole faba beans, dehulled faba beans, hulls) and flours, were soaked in demineralised water at 10 °C (SDW), NaAc buffer of pH 3.5 at 10 °C (SAB), and 500 U L−1 phytase of pH 5.5 at 50 °C (SPS). In whole seeds and flour, SAB removed 20–51% phytic acid from seeds, and SPS 100% from flour depending upon different preheating treatments; dry matter, Ca, Fe, and Zn losses were 1.8–18%, 20–61%, 33–61% and 8–51%. In dehulled seeds flour, phytic acid could be removed 100% by SPS treatments. In dehulled seeds and flour, losses of dry matter, Ca, Fe were 2.4–18.6%, 12–63%, 11–43%, and Zn apparent gain was 12–73% respectively. In hulls flour, SPS removed 80–82% phytic acid, and SAB 48–50% depending upon different preheating treatments; dry matter, Ca, Zn losses were 6.4–28.7%, 16–46%, 60–72%, and Fe apparent gain was 2–73%, respectively. Phytase application is an effective method to rapidly remove phytic acid while preserving relatively more dry matter and minerals than other soaking approaches in faba bean fractions.

Immobilization of Avocado Phytase on Epoxy-Activated Sepabead EC-EP and its Application in Soymilk Phytate Hydrolysis

There is a great demand for using phytases to reduce phytate content in animal feed stuffs and food for human consumption. Industrial application of phytase requires efficient methods to immobilize the enzyme, yielding a biocatalyst with high activity and stability compared to the free enzyme. In the present study, phytase was immobilized on Sepabead EC-EP and then used in the biodegradation of soymilk phytate. The immobilized enzyme exhibited an activity of 0.1 U per g of carrier and activity yield of 70.83%. Optimum temperature and pH for the immobilized enzyme were 55 degrees C and 5.5, respectively. Both the enzymes were stable between pH 3.0-8.0 and below 70 degrees C. Kinetic parameters(K(m) and V(max)) and the usability of the immobilized enzyme were determined. The immobilized enzyme hydrolyzed 65% of soymilk phytate in 8 h at 60 degrees C, as compared with 56% hydrolysis observed for the native enzyme over the same period of time.

Strain improvement and up scaling of phytase production by Aspergillus niger NCIM 563 under submerged fermentation conditions

Combination of physical and chemical mutagenesis was used to isolate hyper secretory strains of Aspergillus niger NCIM 563 for phytase production. Phytase activity of mutant N-1 and N-79 was about 17 and 47% higher than the parent strain. In shake flask the productivity of phytase in parent, mutant N-1 and N-79 was 6,181, 7,619 and 9,523 IU/L per day, respectively. Up scaling of the fermentation from shake flask to 3 and 14 L New Brunswick fermenter was studied. After optimizing various fermentation parameters like aeration, agitation and carbon source in fermentation medium the fermentation time to achieve highest phytase activity was reduced considerably from 14 days in shake flask to 8 days in 14 L fermenter. Highest phytase activity of 80 IU/ml was obtained in 1% rice bran-3.5% glucose containing medium with aeration 0.2 vvm and agitation 550 rpm at room temperature on 8th day of fermentation. Addition of either bavistin (0.1%), penicillin (0.1%), formalin (0.2%) and sodium chloride (10%) in fermented broth were effective in retaining 100% phytase activity for 8 days at room temperature while these reagents along with methanol (50%) and ethanol (50%) confer 100% stability of phytase activity at 4 degrees C till 20 days. Among various carriers used for application of phytase in feed, wheat bran and rice bran were superior to silica and calcium carbonate. Thermo stabilization studies indicate 100% protection of phytase activity in presence of 12% skim milk at 70 degrees C, which will be useful for its spray drying.

In Vitro Solubility of Calcium, Iron, and Zinc in Rice Bran Treated with Phytase, Cellulase, and Protease

Absorption of minerals is inhibited by phytic acid, fiber, and protein because of the chelates formed. Response surface method (RSM) was used in this study to evaluate the effect of application of commercial phytase, protease, and cellulase in rice bran on the in vitro solubility of calcium (IVCa), iron (IVFe), and zinc (IVZn). It is shown that IVCa and IVZn were significantly improved by the application of phytase and cellulase, and the models of two second-order polynomials are recommended for prediction, with coefficients at R(2) = 0.86 and R(2) = 0.88, respectively. Interaction between protease and cellulase also significantly affected IVCa and IVZn. Cellulase had more efficiency than phytase on IVCa. Differing in its effect on Ca and Zn, phytase had a significant linear correlation with IVFe, and none of the other processing parameters exerted a significant effect. The largest increment for IVFe, IVCa, and IVZn occurred in the treatment with applications of phytase at 2.5 U g(-1) and protease and cellulase at 0.2% and 1%, respectively, which were 3.3, 3.6, and 4.3 times, respectively, that of the untreated material.

Evaluation of Heat-Stable Phytases in Pelleted Diets Fed to Broilers from Day Zero to Thirty-Five During the Summer Months

Supplementing phytase to broiler diets has been shown to improve phytate P digestibility in commercial broilers. Broiler rations are typically pelleted to promote improved broiler performance, but high pelleting temperatures can inactivate phytase enzymes. Before the introduction of heat-stable phytase products, phytase enzymes were generally added postpelleting to limit the effect of pelleting temperature on phytase activity. Therefore, the objective of this study was to evaluate postpelleting phytase activity of 3 concentrations of 2 heat-stable phytase enzymes and their efficacy when fed to 0- to 35-d-old broilers. After pelleting the phytase-supplemented diets at 93.3°C, the retained phytase activities of the 2 enzymes were similar, suggesting equivalent heat stability. Supplementing phytase to deficient nonphytate P diets improved FCR by 3% (P ≤ 0.05) compared with the FCR of birds fed an unsupplemented deficient diet. No differences in tibia ash (TA) were detected between the 2 phytase sources; however, TA of birds fed supplemental phytase at the 2 greatest concentrations was improved (P ≤ 0.05) compared with the TA of birds fed nonphytate P-deficient diet. These results suggest that the prepelleting inclusion of heat-stable phytase enzymes may be a viable alternative to postpellet application of phytase for improving P utilization in broilers.

Effects of Phytate and Phytase on the Performance and Immune Function of Broilers Fed Nutritionally Marginal Diets

The effects of phytate and phytase on broiler performance and cellular and humoral immunity was assessed by using 504 Cobb 500 female broilers fed nutritionally marginal diets. At 1 d of age, the chicks were randomly allocated to 6 treatments in a 2 × 3 factorial arrangement including 2 levels of phytate P (0.22 or 0.44%) and 3 dose rates of an Escherichia coli-derived phytase [0, 500, or 1,000 phytase units (FTU)/kg of feed]. Both low- and high-phytate diets were corn and soy based with the same nutritional specifications (AME 2,900 kcal/kg, CP 21.00%, Ca 0.78%, and nonphytate P0.28%), differing only in the concentration of phytate P. The immune status of birds was measured at 14, 21, and 28 d of age. The results revealed that phytase improved feed intake, BW, and feed conversion ratio (P < 0.05), whereas high dietary phytate depressed bird performance (P < 0.05). There was no interaction between phytate and phytase on bird performance. Feeding phytase increased the percentages of erythrocyte rosette-forming cells and erythrocyte-antibody complement cells by 3.03 and 1.83% at d 21, and by 2.76 and 2.20% at d 28, respectively (P < 0.05). The percentages of CD4+ CD8+ T lymphocyte subsets were also increased by phytase (P < 0.05), without affecting the ratio of CD4+ and CD8+. The levels of intestinal secretory IgA were improved with phytase at d 14, 21, and 28 (P < 0.05). Antibodies against Newcastle disease virus vaccine were enhanced at d 21 and 28 in the high-phytate diets with phytase addition (P < 0.05). Increasing the phytase dose to 1,000 FTU/kg did not improve immune function further than 500 FTU/kg. The results suggest that application of phytase in nutritionally marginal diets could enhance lymphocyte numbers and the seral and mucosal antibodies of 1- to 28-d-old broilers, suggesting that both phytate and phytase may have a role in gastrointestinal health and immune competence.

Phytate: impact on environment and human nutrition. A challenge for molecular breeding

Phytic acid (PA) is the primary storage compound of phosphorus in seeds accounting for up to 80% of the total seed phosphorus and contributing as much as 1.5% to the seed dry weight. The negatively charged phosphate in PA strongly binds to metallic cations of Ca, Fe, K, Mg, Mn and Zn making them insoluble and thus unavailable as nutritional factors. Phytate mainly accumulates in protein storage vacuoles as globoids, predominantly located in the aleurone layer (wheat, barley and rice) or in the embryo (maize). During germination, phytate is hydrolysed by endogenous phytase(s) and other phosphatases to release phosphate, inositol and micronutrients to support the emerging seedling. PA and its derivatives are also implicated in RNA export, DNA repair, signalling, endocytosis and cell vesicular trafficking. Our recent studies on purification of phytate globoids, their mineral composition and dephytinization by wheat phytase will be discussed. Biochemical data for purified and characterized phytases isolated from more than 23 plant species are presented, the dephosphorylation pathways of phytic acid by different classes of phytases are compared, and the application of phytase in food and feed is discussed.

Efficacy of Phytases on Egg Production and Nutrient Digestibility in Layers Fed Reduced Phosphorus Diets

The effects of phytases on the performance of layers and the ileal nutrient digestibility of corn-, soybean-, and by-product meal-based diets were assessed with 320 Hy-Line brown layers from 23 to 28 wk of age. Layers were grouped randomly into 5 treatments, with 8 replicates per treatment and 8 layers per replicate. The 5 diets consisted of a positive control diet with adequate Ca (3.30%), total P (0.50%), and nonphytate P (NPP; 0.28%), and a negative control diet with Ca reduced by 0.12%, total P reduced by 0.14%, NPP reduced by 0.13%, and 3 phytases (phytase A derived from Aspergillus niger, and phytases B and C derived from Escherichia coli) supplemented at 300 phytase units/kg of feed, respectively. Egg production and feed intake were recorded daily, and eggshell quality and ileal nutrient digestibility were measured at the end of a 6-wk feeding period. The results revealed that the reduction of Ca and P from the positive control diet significantly depressed feed intake, egg mass, eggshell hardness, and the digestibility of N, Ca, P, and amino acids (P < 0.05). Phytase supplementation in the negative control diet improved the digestibility of P and Ca by 11.08 and 9.81% (P < 0.05), respectively, whereas it improved the digestibility of amino acids by 2 to 8% (P < 0.05). However, the digestibility of most amino acids was not restored to the levels of the positive control diet by the application of phytases. Supplementing phytases in the negative control diet improved the rate of lay, egg mass, and egshell quality to the levels of birds fed the positive control diet. These results suggest that supplementing phytases can improve the digestibility not only of Ca and P, but also of amino acids in layers fed a corn-, soybean-, and by-product-based diet.

Present Scenario and Future Prospects of Phytase in Aquafeed - Review -

Aquaculture pollution is a major concern among the entrepreneurs, farmers and researchers. Excess discharge of phosphorus and nitrogen into the water bodies is the principal pollutant responsible for this. Plant-based feed ingredients due to its high phytic acid content enhances both nitrogen and phosphorus discharge thereby increasing the pollution level. Dietary phytase treatment is probably the best answer to address this problem. This review explains the nature and properties of phytate, its interactions with other nutrients and the application of phytase in aquafeed to reduce the pollution. This review also covers the different biotechnological aspects for lowering the phytic acid level in the common aquafeed ingredients, as an alternate approach to controlling the pollution level. Some of future research needs have also been highlighted to attract the attention of more researchers to this area.

Biotechnological production and applications of phytases

Phytases decompose phytate, which is the primary storage form of phosphate in plants. More than 10 years ago, the first commercial phytase product became available on the market. It offered to help farmers reduce phosphorus excretion of monogastric animals by replacing inorganic phosphates by microbial phytase in the animal diet. Phytase application can reduce phosphorus excretion by up to 50%, a feat that would contribute significantly toward environmental protection. Furthermore, phytase supplementation leads to improved availability of minerals and trace elements. In addition to its major application in animal nutrition, phytase is also used for processing of human food. Research in this field focuses on better mineral absorption and technical improvement of food processing. All commercial phytase preparations contain microbial enzymes produced by fermentation. A wide variety of phytases were discovered and characterized in the last 10 years. Initial steps to produce phytase in transgenic plants were also undertaken. A crucial role for its commercial success relates to the formulation of the enzyme solution delivered from fermentation. For liquid enzyme products, a long shelf life is achieved by the addition of stabilizing agents. More comfortable for many customers is the use of dry enzyme preparations. Different formulation technologies are used to produce enzyme powders that retain enzyme activity, are stable in application, resistant against high temperatures, dust-free, and easy to handle.

Evaluation of Nonuniform Application of Phytase (Simulated) on Growth Performance and Bone Quality in Broiler Chicks

The postpellet application of phytase can be variable and can lead to high coefficients of variation (CV). Two trials were conducted to evaluate the effects of high CV of phytase application (simulated by feeding 2 to 4 times the recommended level of phytase fed every second, third, or fourth day), when added to a standard broiler diet deficient in nonphytate phosphorus (NPP) and fed to 0- to 21-d-old male broilers. A standard curve of graded levels of NPP was included in both trials to determine the NPP requirement. Weight gain, feed efficiency, tibia breaking strength, and percentage tibia ash were assessed in both trials. Trial 1 was designed to evaluate the effect of a high variation (103% CV; 23,000 phytase units (PTU)/kg fed every other day) of added phytase fed on alternate days compared with low variation of added phytase fed daily (0% CV; 11,500 PTU/kg fed daily) and to determine the optimal phytase inclusion level and the NPP requirement. Trial 2 was conducted to determine the maximum variation of phytase addition fed alternate days and reevaluate the NPP requirement. When tibia ash was used as the response criterion, the NPP requirement was determined to be 0.42% in both experiments, and the optimal phytase level was determined to be 14,630 PTU/kg feed. The results of these studies also suggest that feeding a high variation diet of added phytase (103% CV) was not detrimental to the parameters evaluated. Only when the CV exceeded 103% was bone quality negatively affected.

Phytase enzymology, applications, and biotechnology.

Phytases are phosphohydrolases that initiate the step-wise removal of phosphate from phytate. These enzymes have been widely used in animal feeding to improve phosphorus nutrition and to reduce phosphorus pollution of animal waste. The potential of phytases in improving human nutrition of essential trace minerals in plant-derived foods is being explored. This review covers the basic biochemistry and application of phytases, and emphasizes the emerging biotechnology used for developing new effective phytases with improved properties.

Application of Phytase, Microbial or Plant Origin, to Reduce Phosphorus Excretion in Poultry Production

In order to prevent pollution from animal waste, the excretion of nutrients should be reduced through proper nutritional management. Among the many nutrients of concern, such as N, P, Cu, Zn and K, P is one of the most concerned nutrients to be managed. Seven feeding trials, three with layers and four with broilers, were conducted to determine if microbial phytase supplementation can reduce non-phytate phosphorus (NPP) level in diets and results in concomitant reductions of P excretion. The results showed that microbial phytase can be successfully used to achieve these purposes. Activity of natural phytase in certain plant feedstuffs is high enough to be considered in feed formulation. Three experiments have been conducted to study the characteristics of plant phytase and its application to feeding of broilers. Selected brands of wheat bran could be successfully used as a source of phytase in broiler feeding. (Asian-Aust. J. Anim. Sci. 2003. Vol. 16, No. 1 : 124-135)

Solid-state fermentation for production of phytase by Rhizopus oligosporus.

Solid-state fermentation of coconut oil cake has been carried out with Rhizopus oligosporus for the production of phytase. Phytase is used commercially in the animal feed industry to improve animal performance because there is a substantial and growing interest among swine and poultry producers in the application of phytase to improve the nutritional quality in animal feeds. Demonstrated benefits include improved feed yield ratios and reduction in the environmental costs associated with the disposal of animal wastes. We report the production of extracellular phytase by R. oligosporus under solid-state fermentation using coconut oil cake as substrate. Maximal enzyme production (14.29 U/g of dry substrate) occurred at pH 5.3, 30 degrees C, and 54.5% moisture content after 96 h of incubation. The addition of extra nutrients to the substrate resulted in inhibition of product formation. The results indicate the scope for production of phytase using coconut oil cake as solid substrate without additional nutrients.

Simultaneous application of phytase and xylanase to broiler feeds based on wheat: in vitro measurements of phosphorus and pentose release from wheats and wheat‐based feeds

An in vitro procedure that simulated digestion in growing broilers was tested to predict phosphorus availability and arabinoxylan hydrolysis in samples of nine wheat varieties and in a wheat-based diet. Amounts of dialysable phosphorus freed from wheat samples correlated with activities of endogenous phytase (R = 0.913; p < 0.0001), whereas amounts of pentoses released were correlated with viscosities of the digested samples (R = 0.899; p < 0.0001). Differences in phosphorus release resulting from graded levels of microbial phytase added to feeds that were either autoclaved or not autoclaved revealed a decreasing role of endogenous phytase in dephosphorylation as levels of microbial phytase supplementation grew. Amounts of pentoses released from feeds containing two different xylanase preparations reflected literature data on different in vivo efficacies of those preparations. Simultaneous addition of phytase and xylanase affected phosphorus release in a manner that depended upon the form of xylanase preparation used (liquid or powder). There was a positive influence of acid protease on both phytate and arabinoxylan hydrolysis in feeds supplemented with phytase. Effects observed by the in vitro procedures corresponded to in vivo phenomena described in the literature. © 1999 Society of Chemical Industry

Simultaneous application of phytase and xylanase to broiler feeds based on wheat: feeding experiment with growing broilers

In a 2 week study with broilers, graded levels of phytase added to feeds containing 6.70 g kg−1 of Ca, 4.80 g kg−1 of total phosphorus along with the recommended xylanase dosage (400 FXU kg−1) improved body weight gains quadratically (R2 = 0.55; p < 0.0001). Xylanase supplementation to feeds containing 0, 600 or 1000 units kg−1 of phytase significantly improved gains (p = 0.0072) and decreased viscosities of intestinal contents (p < 0.0001). High phytase concentrations in feeds (1000 FTU kg−1) increased intestinal viscosities, whereas, in the presence of xylanase, graded levels of phytase linearly reduced viscosities below the levels attained with xylanase fed as a sole supplemental enzyme (p = 0.033). Contents of ash in the toes were increased by phytase supplementation (p = 0.0011), the effect being more pronounced in birds consuming feeds that also contained xylanase. Improvements in performance and in bone mineralisation resulting from phytase and xylanase supplementation were lower than those observed in the control diets containing 4.50 g kg−1 of available phosphorus. The in vitro dialysable phosphorus predicted 71% of the variability observed in body gains, whereas amounts of pentoses freed from feed samples in vitro were highly correlated (R = 0.919) with feed efficiencies when the data were subjected to Hanes transformation. © 1999 Society of Chemical Industry

Simultaneous application of phytase and xylanase to broiler feeds based on wheat:in vitro measurements of phosphorus and pentose release from wheats and wheat-based feeds

An in vitro procedure that simulated digestion in growing broilers was tested to predict phosphorus availability and arabinoxylan hydrolysis in samples of nine wheat varieties and in a wheat-based diet. Amounts of dialysable phosphorus freed from wheat samples correlated with activities of endogenous phytase (R = 0.913; p < 0.0001), whereas amounts of pentoses released were correlated with viscosities of the digested samples (R = 0.899; p < 0.0001). Differences in phosphorus release resulting from graded levels of microbial phytase added to feeds that were either autoclaved or not autoclaved revealed a decreasing role of endogenous phytase in dephosphorylation as levels of microbial phytase supplementation grew. Amounts of pentoses released from feeds containing two different xylanase preparations reflected literature data on different in vivo efficacies of those preparations. Simultaneous addition of phytase and xylanase affected phosphorus release in a manner that depended upon the form of xylanase preparation used (liquid or powder). There was a positive influence of acid protease on both phytate and arabinoxylan hydrolysis in feeds supplemented with phytase. Effects observed by the in vitro procedures corresponded to in vivo phenomena described in the literature. © 1999 Society of Chemical Industry