A study was conducted to determine the efficacy of phytase, an enzymic cocktail, and a waste Aspe... more A study was conducted to determine the efficacy of phytase, an enzymic cocktail, and a waste Aspergillus niger mycelium to hydrolyze phytate present in corn-soybean meal diets. One hundred turkey poults were assigned to dietary treatments for 2 wk (Days 7 to 21). Dietary treatments included: 1) NRC (1994) diet (NRC), with recommended concentration of 0.6 % availa-ble P (aP) and 1.2 % Ca; 2) Phytase diet (PHYT), 1,000 units phytase/kg diet, 0.16 % aP, and 0.84 % Ca; 3) cocktail diet (COC), 1,000 units of phytase/kg diet plus acid phosphatase (100 units /g of diet), acid protease (42 units/g of diet), pectinase (2.94%), 0.16 % aP, and 0.84% Ca; 4) Fungal mycelium diet (MYC), 5 % mycelium, 0.16 % aP, and 0.84 % Ca; and 5) a positive control diet
A study was conducted to evaluate the effects of dietary supplementation of phytase B (product of... more A study was conducted to evaluate the effects of dietary supplementation of phytase B (product of the Aspergillus niger phyB gene expressed in Trichoderma reesei) on feed intake, laying performance, eggshell quality, and on phosphorus and calcium balance in laying hens. Seventy-two, 40 weeks old Hy Line Brown hens were fed for 14 weeks the following four phosphorus-deficient (0.12% nonphytate phosphorus, NPP), maize-soybean meal-based diets: (1) calcium-deficient (2.8% Ca) control diet; (2) diet 1 + phytase B at the activity of 2.5 acid phosphatase units (AcPU/kg); (3) control diet (3.8% Ca); (4) diet 3 + phytase B at the activity of 2.5 AcPU/ kg. Each dietary treatment was fed to 18 cages of hens, 1 hen/cage kept in individual cages. Hens fed the NPP- and Ca-deficient diets consumed more feed (P < 0.01) and excreted less calcium (P < 0.01) than those receiving P-deficient diets with the standard calcium level. There were no effects of calcium level on feed utilization, egg ma...
The effect of different phytases on the performance response, nutrient utilization and tibia char... more The effect of different phytases on the performance response, nutrient utilization and tibia characteristics of chickens was investigated. The five pelleted diets were the following: positive control (PC) with added monocalcium phosphate; negative control (NC) formulated with equivalency values of phytase for Ca and digestible P; and three further diets where different phytases were individually added to the NC diet at 500 FTU/kg. The phytases were derived either from Aspergillus (phytase I), or E. coli (phytases II and III). Compared to PC, the performance parameters, as well as apparent metabolizable energy (AMEn), mineral retention, bone breaking force and tibia mineral content were suppressed by the reduction of dietary Ca and digestible P. All phytases enhanced the overall body weight gains and feed conversion ratio in comparison with NC, but none outperformed PC. Only phytase II improved AMEn compared to NC and PC group. However only phytase I outperformed NC group in terms of mineral retention and P retention was higher than phytase II and III. No significant differences were observed in fat digestibility and N retention. Bone strength among phytases did not differ and all improved this parameter compared to the NC diet. However, even though all phytases enhanced tibia minerals content, the improvement was less pronounced with phytase III. Moreover, the differences in all analysed tibia minerals between phytase III and II were significant suggesting that even among 6-phytases derived from and expressed in the same organism, different efficacy or mode of action can occur.
A study was conducted to determine whether P in a low phytic acid mutant barley (Hordeum vulgare ... more A study was conducted to determine whether P in a low phytic acid mutant barley (Hordeum vulgare L.; MB) containing the lpa 1-1 allele is more available than P in a near-isogenic, wild-type barley (NB). The MB contained 0.21% non-phytate P (nP) (estimated available P; aP) and 0.35% total P (tP), whereas NB contained 0.11% aP and 0.35% tP. A completely randomized design was used with 150 1-d-old male poults randomly assigned to five treatments (six pens of five poults per treatment) for 21 d. The five treatments were 1) a NB diet containing 0.30% aP, 0.41% tP, and 1.0% Ca; 2) a MB diet containing 0.36% aP, 0.41% tP, and 1.0% Ca; 3) a NB diet similar to Diet 1 but with KH 2 PO 4 added to increase the aP to 0.36% (0.47% tP) to match the aP in Diet 2; 4) a MB diet containing 0.60% aP, 0.86% tP, and 1.2% Ca; and 5) a NB diet containing 0.60% aP, 0.92% tP, and 1.20% Ca. Performance and bone ash were significantly lower (P < 0.05) in poults fed Diet 1 compared with those in poults fed Diet 2. Performance and bone ash were similar (P > 0.05) in poults fed Diets 2 and 3 and in poults fed Diets 4 and 5. Poults fed Diet 1 retained 13.9% more P than did poults fed Diet 2 (P < 0.05). Poults fed Diets 2 and 4 retained 11.9 and 4.9% more P than poults fed Diets 3 and 5, respectively (P < 0.05). Poults fed MB diets excreted 41% less P than did poults fed NB diets when barley was the sole source of phytic acid in the diet. Results of the current study indicate that P in MB is more available than P in NB, and decreasing the phytate content did not compromise the nutritional value of MB.
At low levels of dietary calcium and phosphorus (2.8 and 0.36%, respectively), the degree of phyt... more At low levels of dietary calcium and phosphorus (2.8 and 0.36%, respectively), the degree of phytate conversion raised by adding phytase B to feeds that had already been supplemented with 3-or 6-phytase A, modulated production parameters, and egg shell quality of Hy Line Brown laying hens fed maize-soyabean meal from week 40 to 54. Feed consumption, laying rate, mean egg weight and shell breaking strength depended on the type of phytase A added to feeds only at a low phytate conversion degree. Egg production and feed per egg conversion ratio did not depend on the dietary supplements studied.
The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface met... more The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface method were found to correlate with their capabilities to synthesize intracellular enzymes which degrade phytates (phytase and acid phosphatase). Another high correlation was observed between phytase and acid phosphatase activities bound to the cell walls of mycelia.
The effect of initial solid and moisture contents, temperature and time of incubation on the prod... more The effect of initial solid and moisture contents, temperature and time of incubation on the production of polygalacturonase, phytase, acid phosphatase, xylanase and β-glucanase by Aspergillus niger 377-4 during solid state fermentation was studied. Parameters of enzyme synthesis were optimized using statistical experimental designs. It was shown that the capacity of strain to synthesize the aforementioned enzymes could be modified within a wide range by culture parameters selection. The optimal polygalacturonase production efficiency was achieved with the initial medium mass of 19.9 g and humidity of 59.9%, after 77.7 h of incubation at 28.9 o C. The best combination of culture parameters for phytase synthesis was: initial medium mass 19.9 g, moistures 50%, temperature 33 o C and incubation time 83.9 h. The highest activity of acid phosphatase was obtained after 81.3 h of incubation at 27 o C, with initial substrate mass of 17.8 g and moistness content of 60%. The initial solid and...
Prepar ations of 6-phytase A (EC 3.1.3.26) and phytase B (acid phosphatase, EC 3.1.3.2) were appl... more Prepar ations of 6-phytase A (EC 3.1.3.26) and phytase B (acid phosphatase, EC 3.1.3.2) were applied alone and combined in the preparation of dough to estimate their catalytic potential for myo-inositol liberation from rye fl our in the breadmaking technology. The experimental bread samples were ground a er baking and subjected to determination of myo-inositol bioavailability by an in vitro method that simulated digestion in a human alimentary tract, followed by measurements of myo-inositol transport through enterocyte--like diff erentiated Caco-2 cells to determine its bioaccessibility. Myo-inositol content was measured by a high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) technique. The concentration of myo-inositol in the dialysates of control bread was 25.3 µg/mL, whereas in the dialysates of bread sample baked with 6-phytase A, the concentration increased to 35.4 µg/mL, and in the bread baked with phytase B to 64.98 µg/mL. Simultaneous application of both enzymes resulted in myo-inositol release of 64.04 µg/mL. The highest bioaccessibility of myo-inositol, assessed by the measurement of the passage through the Caco-2 monolayer was determined in the bread baked with the addition of 6-phytase A. Enzymatically modifi ed rye bread, particularly by the addition of 6-phytase A, may be therefore a rich source of a highly bioaccessible myo--inositol.
Phytase is well studied and explored, however, little is known about its effects on the microbial... more Phytase is well studied and explored, however, little is known about its effects on the microbial ecology of the gastrointestinal tract. In total, 400 one-day-old female Ross 308 chicks were randomly distributed to four experimental groups. The dietary treatments were arranged as a 2 × 2 complete factorial design, with the factors being adequate (PC) or insufficient calcium (Ca) and digestible phosphor (dP)(NC) and with or without 5000 phytase units (FTU)/kg of Escherichia coli 6-phytase. The gastrointestinal tract pH values, ileal microbial communities and short-chain fatty acid concentrations in the digesta were determined. The reduction in Ca and dP concentration significantly affected pH in the crop and caeca, and addition of phytase to the NC resulted in a pH increase in the ileum. The reduction in Ca and dP concentration significantly lowered, while phytase supplementation increased ileal total bacterial counts. Additionally, the deficient diet reduced butyrate- but increased ...
Investigating intestinal physiology in vitro remains challenging due to the lack of an effective ... more Investigating intestinal physiology in vitro remains challenging due to the lack of an effective primary enterocyte culture system. Recently developed protocols for growing organoids containing crypts and villus from adult mouse intestinal epithelium in Matrigel present an attractive alternative to the classical techniques. However, these approaches require the use of sophisticated and expensive serum-free medium supplemented with epithelial growth factor (EGF), Wnt agonist (R-spondin 1), and bone morphogenetic protein inhibitor (Noggin) in high concentrations. Here we demonstrate that is possible to use an isolated chicken embryonic intestinal epithelium to create such an organoid culture. Structures formed in Matrigel matrix in the first two days following isolation survive and enlarge during ensuing weeks. They have the appearance of empty spheres and comprise cells expressing cytokeratin (an epithelial cell marker), villin (a marker of enterocytes), and Sox-9 (a transcription fa...
The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface met... more The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface method were found to correlate with their capabilities to synthesize intracellular enzymes which degrade phytates (phytase and acid phosphatase). Another high correlation was observed between phytase and acid phosphatase activities bound to the cell walls of mycelia.
A study was conducted to determine the bioavailability of P in a low phytic acid mutant barley (H... more A study was conducted to determine the bioavailability of P in a low phytic acid mutant barley (Hordeum vulgare L.; MB) containing the lpa 1-1 allele and a near-isogenic wild-type hybrid barley (NB). By analysis, the MB contained 0.21% non-phytate P (nP) and 0.35% total P (tP), whereas NB contained 0.11% nP and 0.35% tP. Non-phytate P was considered to be available to poultry. Reagent grade KH 2 PO 4 was used as a standard source to determine relative availability. The experimental design consisted of 10 dietary treatments fed to six replicate pens of six chicks each. Dietary treatments included a low available P (aP) basal diet (0.20% aP; 1.0% Ca; Diet 1) supplemented with 0.04, 0.08, and 0.12% P from reagent-grade KH 2 PO 4 (Diets 2, 3, and 4, respectively), MB (Diets 5, 6, and 7, respectively), and NB (Diets 8, 9, and 10, respectively). Diets were fed ad libitum for 14 d. Feed intake was not affected (P > 0.10) by source or level of P. Body weight gain was not influenced (P > 0.05) by P level, but was affected (P < 0.05) by P source. Feed conversion was influenced (P < 0.05) by both P source and level. Based on non-linear regression analysis (assuming KH 2 PO 4 = 100%) and using percentage toe ash as the response variable, bioavailability of P was determined to be 49.3 ± 17% for MB and 28.0 ± 15% for NB. Based on the results of toe ash, P in MB was 1.8 times more available to chicks than P in NB. These results compare favorably with analyzed nP values, which indicated that MB contains 1.9 times as much nP as NB (0.21 vs. 0.11%).
Abstract Commercial microbial phytase (Natuphos®5000, 1000 units/kg) was fed to turkey hens grown... more Abstract Commercial microbial phytase (Natuphos®5000, 1000 units/kg) was fed to turkey hens grown to market age as a partial substitute for inorganic phosphate in corn-soybean meal diets. Available P (aP) levels followed NRC recommendations or were reduced by 0.15%. Dietary Ca levels remained at levels recommended by NRC. This process revealed that phytase improved feed intake and body weight gains of turkey hens grown to market age. Partial substitution of phytase for inorganic P did not influence body composition, toe ash, or bone breaking strength. Partial substitution of phytase for aP did not result in increased P retention from commercial corn-soybean meal diets. Results suggest that too high a P concentration, too wide a Ca:P ratio, or both prevented Natuphos® phytase from increasing P digestibility under commercial type conditions employed in this study.
An ion chromatography technique (HPAEC-PAD) was employed for determination of inositol in plant c... more An ion chromatography technique (HPAEC-PAD) was employed for determination of inositol in plant components of feeds as an alternative to a microbiological assay. The study included analysis of total, free and dialysable inositol released from samples by an in vitro procedure simulating digestion in the gastrointestinal tract of birds. Inositol was separated on the high-performance anionexchange CarboPack MA1 column and monitored by the pulsed amperometric detection (PAD). As compared to the HPAEC-PAD technique, microbiological assay produced 2-3 fold lower values of total inositol, probably due to myo-inositol stereoselectivity of Saccharomyces cerevisiae ATCC 9080 analytical strain. The overall correlation coefficient was 0.989, 0.941 and 0.736 for total, free and dialysable inositol, respectively. Microbiological method of analysis rather than the HPAEC-PAD technique may be recommended for determination of dialysable myo-inositol released from feed samples during in vitro digestions that simulate gastrointestinal tract of birds. The HPAEC-PAD technique was more sensitive, convenient and suitable for total inositol determination.
Green tea catechins are classified as (-)-epiforms (2R, 3R) or (-)-forms (2S, 3R) with respect to... more Green tea catechins are classified as (-)-epiforms (2R, 3R) or (-)-forms (2S, 3R) with respect to stereoisomerism. The (-)-forms (2S, 3R) in catechin preparations obtained from green tea amounts to approximately 10% of total catechins, whereas the other 90% are (-)-epiforms (2R, 3R). ). This study investigated the effect of catechin preparations containing 10, 20 and 30% (-)-forms (2S, 3R) on the oxidative and hydrolytic stability of butter. For comparison, butter with δ-tocopherol and BHT and butter without stabilizer were analysed. Butter stability was examined under conditions of refrigeration (8 o C) and freezing (-22 o C) and at temperature of 50 o C and 100 o C. Catechin preparations were more efficient butter stabilizers than BHT and δ-tocopherol. Thermal modification of catechins that led to the generation of 20% of (-) forms (2S, 3R) improved their antioxidative efficacy, but longer treatment leading to the formation of 30% of (-) forms (2S, 3R) decreased their antioxidative activity. The hydrolytic stability of butter, however, increased as the amount of (-) forms (2S, 3R) increased.
World Journal of Microbiology & Biotechnology, 1993
Acid phosphatase present in preparations ofAspergillus niger phytase accelerated dephosphorylatio... more Acid phosphatase present in preparations ofAspergillus niger phytase accelerated dephosphorylation of sodium phytate. Its influence on the reaction rate and distribution ofmyo-inositol phosphates was most apparent at low pH value (2.5) and when acid-hydrolysed substrate was de-esterified enzymatically. With partly purified phytase, the predominant inositol form was tetraphosphate but a preparation having acid phosphatase activity caused an even distribution of lower inositol phosphates after a few hours.
An in vitro method was developed to predict inorganic P release from maize-soyabean poultry feeds... more An in vitro method was developed to predict inorganic P release from maize-soyabean poultry feeds containing supplemental phytase (EC31·3·8), and to quantify the effect of acid phosphatase (EC 3·1·3·2), fungal protease (EC 3·4·23·6) and Aspergillus niger cellulase (EC 3·2·1·4) on phytate dephosphorylation. Pepsin (EC3·4·23·1) and pancreatin digestion periods were preceded by a 30 min pre-incubation at pH 5·25 to simulate digestion in the crop of poultry. Pancreatin digestion was carried out in dialysis tubing, with a ratio of about 1:25 (v/v) between the digesta and dialysing medium, to simulate gradient absorption from the duodenum. The feed:water ratio was kept within physiological limits and a constant proportion of feed weight to digestive enzymes was maintained. There was a linear response to increasing dosages of phytase up to 1000 phytase units (FTU)/kg feed, and to increasing phosphate concentration in feeds. In vivo validation was performed with growing turkeys (1–3 weeks) ...
The rate of phytate P removal from feed (level of dephosphorylation, DL) and the extent to which ... more The rate of phytate P removal from feed (level of dephosphorylation, DL) and the extent to which the molecule of phytic acid is deprived of phosphate moieties (conversion degree, CD) were studied in vitro and in a feeding trial with broilers fed corn-soybean diets. In the in vitro model, phytase A asymptotically increased DL and CD. Phytase B influenced DL only at low dosages of phytase A [0 or 250 phytase activity units (FTU)/kg], but it enhanced CD irrespective of phytase A activity. In the feeding trial, 3-phytase A and 6-phytase A (at 750 FTU/kg) exerted similar effects on broiler performance and similarly influenced bone mineralization, P retention, and Ca retention. Phytase B [6,400 acid phosphatase activity units (ACPU)/kg] enhanced feed intake, BW gain (BWG), toe ash, and P retention but not the retention of
A study was conducted to determine the efficacy of phytase, an enzymic cocktail, and a waste Aspe... more A study was conducted to determine the efficacy of phytase, an enzymic cocktail, and a waste Aspergillus niger mycelium to hydrolyze phytate present in corn-soybean meal diets. One hundred turkey poults were assigned to dietary treatments for 2 wk (Days 7 to 21). Dietary treatments included: 1) NRC (1994) diet (NRC), with recommended concentration of 0.6 % availa-ble P (aP) and 1.2 % Ca; 2) Phytase diet (PHYT), 1,000 units phytase/kg diet, 0.16 % aP, and 0.84 % Ca; 3) cocktail diet (COC), 1,000 units of phytase/kg diet plus acid phosphatase (100 units /g of diet), acid protease (42 units/g of diet), pectinase (2.94%), 0.16 % aP, and 0.84% Ca; 4) Fungal mycelium diet (MYC), 5 % mycelium, 0.16 % aP, and 0.84 % Ca; and 5) a positive control diet
A study was conducted to evaluate the effects of dietary supplementation of phytase B (product of... more A study was conducted to evaluate the effects of dietary supplementation of phytase B (product of the Aspergillus niger phyB gene expressed in Trichoderma reesei) on feed intake, laying performance, eggshell quality, and on phosphorus and calcium balance in laying hens. Seventy-two, 40 weeks old Hy Line Brown hens were fed for 14 weeks the following four phosphorus-deficient (0.12% nonphytate phosphorus, NPP), maize-soybean meal-based diets: (1) calcium-deficient (2.8% Ca) control diet; (2) diet 1 + phytase B at the activity of 2.5 acid phosphatase units (AcPU/kg); (3) control diet (3.8% Ca); (4) diet 3 + phytase B at the activity of 2.5 AcPU/ kg. Each dietary treatment was fed to 18 cages of hens, 1 hen/cage kept in individual cages. Hens fed the NPP- and Ca-deficient diets consumed more feed (P < 0.01) and excreted less calcium (P < 0.01) than those receiving P-deficient diets with the standard calcium level. There were no effects of calcium level on feed utilization, egg ma...
The effect of different phytases on the performance response, nutrient utilization and tibia char... more The effect of different phytases on the performance response, nutrient utilization and tibia characteristics of chickens was investigated. The five pelleted diets were the following: positive control (PC) with added monocalcium phosphate; negative control (NC) formulated with equivalency values of phytase for Ca and digestible P; and three further diets where different phytases were individually added to the NC diet at 500 FTU/kg. The phytases were derived either from Aspergillus (phytase I), or E. coli (phytases II and III). Compared to PC, the performance parameters, as well as apparent metabolizable energy (AMEn), mineral retention, bone breaking force and tibia mineral content were suppressed by the reduction of dietary Ca and digestible P. All phytases enhanced the overall body weight gains and feed conversion ratio in comparison with NC, but none outperformed PC. Only phytase II improved AMEn compared to NC and PC group. However only phytase I outperformed NC group in terms of mineral retention and P retention was higher than phytase II and III. No significant differences were observed in fat digestibility and N retention. Bone strength among phytases did not differ and all improved this parameter compared to the NC diet. However, even though all phytases enhanced tibia minerals content, the improvement was less pronounced with phytase III. Moreover, the differences in all analysed tibia minerals between phytase III and II were significant suggesting that even among 6-phytases derived from and expressed in the same organism, different efficacy or mode of action can occur.
A study was conducted to determine whether P in a low phytic acid mutant barley (Hordeum vulgare ... more A study was conducted to determine whether P in a low phytic acid mutant barley (Hordeum vulgare L.; MB) containing the lpa 1-1 allele is more available than P in a near-isogenic, wild-type barley (NB). The MB contained 0.21% non-phytate P (nP) (estimated available P; aP) and 0.35% total P (tP), whereas NB contained 0.11% aP and 0.35% tP. A completely randomized design was used with 150 1-d-old male poults randomly assigned to five treatments (six pens of five poults per treatment) for 21 d. The five treatments were 1) a NB diet containing 0.30% aP, 0.41% tP, and 1.0% Ca; 2) a MB diet containing 0.36% aP, 0.41% tP, and 1.0% Ca; 3) a NB diet similar to Diet 1 but with KH 2 PO 4 added to increase the aP to 0.36% (0.47% tP) to match the aP in Diet 2; 4) a MB diet containing 0.60% aP, 0.86% tP, and 1.2% Ca; and 5) a NB diet containing 0.60% aP, 0.92% tP, and 1.20% Ca. Performance and bone ash were significantly lower (P < 0.05) in poults fed Diet 1 compared with those in poults fed Diet 2. Performance and bone ash were similar (P > 0.05) in poults fed Diets 2 and 3 and in poults fed Diets 4 and 5. Poults fed Diet 1 retained 13.9% more P than did poults fed Diet 2 (P < 0.05). Poults fed Diets 2 and 4 retained 11.9 and 4.9% more P than poults fed Diets 3 and 5, respectively (P < 0.05). Poults fed MB diets excreted 41% less P than did poults fed NB diets when barley was the sole source of phytic acid in the diet. Results of the current study indicate that P in MB is more available than P in NB, and decreasing the phytate content did not compromise the nutritional value of MB.
At low levels of dietary calcium and phosphorus (2.8 and 0.36%, respectively), the degree of phyt... more At low levels of dietary calcium and phosphorus (2.8 and 0.36%, respectively), the degree of phytate conversion raised by adding phytase B to feeds that had already been supplemented with 3-or 6-phytase A, modulated production parameters, and egg shell quality of Hy Line Brown laying hens fed maize-soyabean meal from week 40 to 54. Feed consumption, laying rate, mean egg weight and shell breaking strength depended on the type of phytase A added to feeds only at a low phytate conversion degree. Egg production and feed per egg conversion ratio did not depend on the dietary supplements studied.
The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface met... more The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface method were found to correlate with their capabilities to synthesize intracellular enzymes which degrade phytates (phytase and acid phosphatase). Another high correlation was observed between phytase and acid phosphatase activities bound to the cell walls of mycelia.
The effect of initial solid and moisture contents, temperature and time of incubation on the prod... more The effect of initial solid and moisture contents, temperature and time of incubation on the production of polygalacturonase, phytase, acid phosphatase, xylanase and β-glucanase by Aspergillus niger 377-4 during solid state fermentation was studied. Parameters of enzyme synthesis were optimized using statistical experimental designs. It was shown that the capacity of strain to synthesize the aforementioned enzymes could be modified within a wide range by culture parameters selection. The optimal polygalacturonase production efficiency was achieved with the initial medium mass of 19.9 g and humidity of 59.9%, after 77.7 h of incubation at 28.9 o C. The best combination of culture parameters for phytase synthesis was: initial medium mass 19.9 g, moistures 50%, temperature 33 o C and incubation time 83.9 h. The highest activity of acid phosphatase was obtained after 81.3 h of incubation at 27 o C, with initial substrate mass of 17.8 g and moistness content of 60%. The initial solid and...
Prepar ations of 6-phytase A (EC 3.1.3.26) and phytase B (acid phosphatase, EC 3.1.3.2) were appl... more Prepar ations of 6-phytase A (EC 3.1.3.26) and phytase B (acid phosphatase, EC 3.1.3.2) were applied alone and combined in the preparation of dough to estimate their catalytic potential for myo-inositol liberation from rye fl our in the breadmaking technology. The experimental bread samples were ground a er baking and subjected to determination of myo-inositol bioavailability by an in vitro method that simulated digestion in a human alimentary tract, followed by measurements of myo-inositol transport through enterocyte--like diff erentiated Caco-2 cells to determine its bioaccessibility. Myo-inositol content was measured by a high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) technique. The concentration of myo-inositol in the dialysates of control bread was 25.3 µg/mL, whereas in the dialysates of bread sample baked with 6-phytase A, the concentration increased to 35.4 µg/mL, and in the bread baked with phytase B to 64.98 µg/mL. Simultaneous application of both enzymes resulted in myo-inositol release of 64.04 µg/mL. The highest bioaccessibility of myo-inositol, assessed by the measurement of the passage through the Caco-2 monolayer was determined in the bread baked with the addition of 6-phytase A. Enzymatically modifi ed rye bread, particularly by the addition of 6-phytase A, may be therefore a rich source of a highly bioaccessible myo--inositol.
Phytase is well studied and explored, however, little is known about its effects on the microbial... more Phytase is well studied and explored, however, little is known about its effects on the microbial ecology of the gastrointestinal tract. In total, 400 one-day-old female Ross 308 chicks were randomly distributed to four experimental groups. The dietary treatments were arranged as a 2 × 2 complete factorial design, with the factors being adequate (PC) or insufficient calcium (Ca) and digestible phosphor (dP)(NC) and with or without 5000 phytase units (FTU)/kg of Escherichia coli 6-phytase. The gastrointestinal tract pH values, ileal microbial communities and short-chain fatty acid concentrations in the digesta were determined. The reduction in Ca and dP concentration significantly affected pH in the crop and caeca, and addition of phytase to the NC resulted in a pH increase in the ileum. The reduction in Ca and dP concentration significantly lowered, while phytase supplementation increased ileal total bacterial counts. Additionally, the deficient diet reduced butyrate- but increased ...
Investigating intestinal physiology in vitro remains challenging due to the lack of an effective ... more Investigating intestinal physiology in vitro remains challenging due to the lack of an effective primary enterocyte culture system. Recently developed protocols for growing organoids containing crypts and villus from adult mouse intestinal epithelium in Matrigel present an attractive alternative to the classical techniques. However, these approaches require the use of sophisticated and expensive serum-free medium supplemented with epithelial growth factor (EGF), Wnt agonist (R-spondin 1), and bone morphogenetic protein inhibitor (Noggin) in high concentrations. Here we demonstrate that is possible to use an isolated chicken embryonic intestinal epithelium to create such an organoid culture. Structures formed in Matrigel matrix in the first two days following isolation survive and enlarge during ensuing weeks. They have the appearance of empty spheres and comprise cells expressing cytokeratin (an epithelial cell marker), villin (a marker of enterocytes), and Sox-9 (a transcription fa...
The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface met... more The ability of eight strains of Aspergillus niger to produce citric acid by the solid surface method were found to correlate with their capabilities to synthesize intracellular enzymes which degrade phytates (phytase and acid phosphatase). Another high correlation was observed between phytase and acid phosphatase activities bound to the cell walls of mycelia.
A study was conducted to determine the bioavailability of P in a low phytic acid mutant barley (H... more A study was conducted to determine the bioavailability of P in a low phytic acid mutant barley (Hordeum vulgare L.; MB) containing the lpa 1-1 allele and a near-isogenic wild-type hybrid barley (NB). By analysis, the MB contained 0.21% non-phytate P (nP) and 0.35% total P (tP), whereas NB contained 0.11% nP and 0.35% tP. Non-phytate P was considered to be available to poultry. Reagent grade KH 2 PO 4 was used as a standard source to determine relative availability. The experimental design consisted of 10 dietary treatments fed to six replicate pens of six chicks each. Dietary treatments included a low available P (aP) basal diet (0.20% aP; 1.0% Ca; Diet 1) supplemented with 0.04, 0.08, and 0.12% P from reagent-grade KH 2 PO 4 (Diets 2, 3, and 4, respectively), MB (Diets 5, 6, and 7, respectively), and NB (Diets 8, 9, and 10, respectively). Diets were fed ad libitum for 14 d. Feed intake was not affected (P > 0.10) by source or level of P. Body weight gain was not influenced (P > 0.05) by P level, but was affected (P < 0.05) by P source. Feed conversion was influenced (P < 0.05) by both P source and level. Based on non-linear regression analysis (assuming KH 2 PO 4 = 100%) and using percentage toe ash as the response variable, bioavailability of P was determined to be 49.3 ± 17% for MB and 28.0 ± 15% for NB. Based on the results of toe ash, P in MB was 1.8 times more available to chicks than P in NB. These results compare favorably with analyzed nP values, which indicated that MB contains 1.9 times as much nP as NB (0.21 vs. 0.11%).
Abstract Commercial microbial phytase (Natuphos®5000, 1000 units/kg) was fed to turkey hens grown... more Abstract Commercial microbial phytase (Natuphos®5000, 1000 units/kg) was fed to turkey hens grown to market age as a partial substitute for inorganic phosphate in corn-soybean meal diets. Available P (aP) levels followed NRC recommendations or were reduced by 0.15%. Dietary Ca levels remained at levels recommended by NRC. This process revealed that phytase improved feed intake and body weight gains of turkey hens grown to market age. Partial substitution of phytase for inorganic P did not influence body composition, toe ash, or bone breaking strength. Partial substitution of phytase for aP did not result in increased P retention from commercial corn-soybean meal diets. Results suggest that too high a P concentration, too wide a Ca:P ratio, or both prevented Natuphos® phytase from increasing P digestibility under commercial type conditions employed in this study.
An ion chromatography technique (HPAEC-PAD) was employed for determination of inositol in plant c... more An ion chromatography technique (HPAEC-PAD) was employed for determination of inositol in plant components of feeds as an alternative to a microbiological assay. The study included analysis of total, free and dialysable inositol released from samples by an in vitro procedure simulating digestion in the gastrointestinal tract of birds. Inositol was separated on the high-performance anionexchange CarboPack MA1 column and monitored by the pulsed amperometric detection (PAD). As compared to the HPAEC-PAD technique, microbiological assay produced 2-3 fold lower values of total inositol, probably due to myo-inositol stereoselectivity of Saccharomyces cerevisiae ATCC 9080 analytical strain. The overall correlation coefficient was 0.989, 0.941 and 0.736 for total, free and dialysable inositol, respectively. Microbiological method of analysis rather than the HPAEC-PAD technique may be recommended for determination of dialysable myo-inositol released from feed samples during in vitro digestions that simulate gastrointestinal tract of birds. The HPAEC-PAD technique was more sensitive, convenient and suitable for total inositol determination.
Green tea catechins are classified as (-)-epiforms (2R, 3R) or (-)-forms (2S, 3R) with respect to... more Green tea catechins are classified as (-)-epiforms (2R, 3R) or (-)-forms (2S, 3R) with respect to stereoisomerism. The (-)-forms (2S, 3R) in catechin preparations obtained from green tea amounts to approximately 10% of total catechins, whereas the other 90% are (-)-epiforms (2R, 3R). ). This study investigated the effect of catechin preparations containing 10, 20 and 30% (-)-forms (2S, 3R) on the oxidative and hydrolytic stability of butter. For comparison, butter with δ-tocopherol and BHT and butter without stabilizer were analysed. Butter stability was examined under conditions of refrigeration (8 o C) and freezing (-22 o C) and at temperature of 50 o C and 100 o C. Catechin preparations were more efficient butter stabilizers than BHT and δ-tocopherol. Thermal modification of catechins that led to the generation of 20% of (-) forms (2S, 3R) improved their antioxidative efficacy, but longer treatment leading to the formation of 30% of (-) forms (2S, 3R) decreased their antioxidative activity. The hydrolytic stability of butter, however, increased as the amount of (-) forms (2S, 3R) increased.
World Journal of Microbiology & Biotechnology, 1993
Acid phosphatase present in preparations ofAspergillus niger phytase accelerated dephosphorylatio... more Acid phosphatase present in preparations ofAspergillus niger phytase accelerated dephosphorylation of sodium phytate. Its influence on the reaction rate and distribution ofmyo-inositol phosphates was most apparent at low pH value (2.5) and when acid-hydrolysed substrate was de-esterified enzymatically. With partly purified phytase, the predominant inositol form was tetraphosphate but a preparation having acid phosphatase activity caused an even distribution of lower inositol phosphates after a few hours.
An in vitro method was developed to predict inorganic P release from maize-soyabean poultry feeds... more An in vitro method was developed to predict inorganic P release from maize-soyabean poultry feeds containing supplemental phytase (EC31·3·8), and to quantify the effect of acid phosphatase (EC 3·1·3·2), fungal protease (EC 3·4·23·6) and Aspergillus niger cellulase (EC 3·2·1·4) on phytate dephosphorylation. Pepsin (EC3·4·23·1) and pancreatin digestion periods were preceded by a 30 min pre-incubation at pH 5·25 to simulate digestion in the crop of poultry. Pancreatin digestion was carried out in dialysis tubing, with a ratio of about 1:25 (v/v) between the digesta and dialysing medium, to simulate gradient absorption from the duodenum. The feed:water ratio was kept within physiological limits and a constant proportion of feed weight to digestive enzymes was maintained. There was a linear response to increasing dosages of phytase up to 1000 phytase units (FTU)/kg feed, and to increasing phosphate concentration in feeds. In vivo validation was performed with growing turkeys (1–3 weeks) ...
The rate of phytate P removal from feed (level of dephosphorylation, DL) and the extent to which ... more The rate of phytate P removal from feed (level of dephosphorylation, DL) and the extent to which the molecule of phytic acid is deprived of phosphate moieties (conversion degree, CD) were studied in vitro and in a feeding trial with broilers fed corn-soybean diets. In the in vitro model, phytase A asymptotically increased DL and CD. Phytase B influenced DL only at low dosages of phytase A [0 or 250 phytase activity units (FTU)/kg], but it enhanced CD irrespective of phytase A activity. In the feeding trial, 3-phytase A and 6-phytase A (at 750 FTU/kg) exerted similar effects on broiler performance and similarly influenced bone mineralization, P retention, and Ca retention. Phytase B [6,400 acid phosphatase activity units (ACPU)/kg] enhanced feed intake, BW gain (BWG), toe ash, and P retention but not the retention of
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