O objetivo deste trabalho foi avaliar a disponibilidade de Fe, Cu e Zn em presença de mesocarpo de babaçu, pois o mesocarpo contem compostos antinutricionais (fitato e lignina) que podem diminuir a disponibilidade de nutrientes minerais. As concentrações de Fe, Cu e Zn encontradas no mesocarpo de babaçu foram 17, 7,0 e 2,6 µg g-1, respectivamente. Para um melhor entendimento das interações dos elementos com o mesocarpo, foi realizada a extração de substâncias complexantes com soluções de NaOH (pH 7 e 12). Análises por espectrometria de absorção molecular indicaram que o principal complexante presente no extrato era o fitato. Soluções de Fe, Cu e Zn (10 a 300 mg L-1) foram adicionadas aos extratos, para que, após agitação e centrifugação, fossem determinadas as recuperações desses elementos. Os resultados mostraram que os elementos formam complexos com o fitato e a solubilidade desses complexos é dependente da razão elemento/fitato, quanto maior essa razão menor é a solubilidade. Também foi avaliada a interação dos elementos com o sólido remanescente da extração. Fe, Cu ou Zn foram adicionados ao mesocarpo lavado. Não foi possível afirmar que houve interação do mesocarpo lavado com Fe e Cu devido à precipitação decorrente da hidrólise. Porém, o Zn não sofreu hidrólise e teve baixas recuperações, indicando que houve interação com o sólido. Além disso, pode ser observado que ao adicionar maiores concentrações de Zn obtiveram-se melhores recuperações, provavelmente devido à saturação dos sítios de ligação. Para os estudos de disponibilidade, foi utilizado o procedimento de digestão simulada in vitro da US Pharmacopeia. Fe e Zn não foram extraídos durante a digestão do mesocarpo, portanto esses elementos não estariam disponíveis para absorção pelo organismo. Entretanto, cerca de 120 µg L-1 de Cu foi extraído do mesocarpo. Foram realizadas digestões do mesocarpo com adição de Fe, Cu e Zn e apenas 48% do Fe, 65% do Cu e 75% do Zn foram recuperados. Digestões contendo essa mesma concentração dos elementos foram realizadas com adições de Ca e Mg. A presença do Ca diminuiu a recuperação de Fe e Zn. Para simular uma situação próxima ao real, foram realizadas digestões gastrointestinais de leite, mistura de leite e mesocarpo e mistura de leite e lignina. Apesar do ferro presente no leite apresentar baixa disponibilidade, quando o mesmo foi misturado ao mesocarpo observou-se um aumento da concentração de Fe solúvel. Compostos presentes no leite ou no mesocarpo podem aumentar a solubilidade dos elementos. Não foi possível observar se havia alguma alteração da disponibilidade do Cu presente no leite na presença do mesocarpo, pois a concentração de Cu no leite é muito baixa. A fração de Zn solúvel na mistura de leite e mesocarpo permaneceu a mesma, porém a fração dialisável foi praticamente nula. A adição de lignina ao leite aumentou a extração dos elementos. Entretanto, esses elementos continuaram não sendo dialisados / The aim of this study was to evaluate the availability of Fe, Cu and Zn in the presence of mesocarp of babassu because it contains antinutritional compounds (phytate and lignin) that may decrease the availability of mineral nutrients. The concentrations of Fe, Cu and Zn found in the mesocarp of babassu were 17, 7.0 and 2.6 mg g-1, respectively. For a better understanding of elements interactions with mesocarp, it was performed the extraction of complexing substances with NaOH solutions (pH 7 and 12). Analysis by molecular absorption spectrometry indicated that the main complexing present in the extract was the phytate. Solutions of Fe, Cu and Zn (10 to 300 mg L-1) were added to the extracts for determining the recoveries of these elements, after agitation and centrifugation. The results showed that the elements form complexes with phytate and the solubility of these complexes is dependent on the ratio element / phytate, the greater this ratio the lower the solubility. The elements interaction with the remaining solid from extraction was also evaluated. Fe, Cu or Zn were added to the washed mesocarp. It was not possible to affirm that there was interaction of washed mesocarp with Fe and Cu due to precipitation derived from hydrolysis. However, Zn did not suffer hydrolysis and had low recoveries, indicating that there was an interaction with the solid. Furthermore, it can be seen that adding higher concentrations of Zn, better recoveries were obtained, probably due to saturation of the binding sites. For the studies of availability, it was used an in vitro simulated digestion procedure from U.S. Pharmacopeia. Fe and Zn were not extracted during digestion of the mesocarp, therefore these elements would not be available for absorption by the organism. However, about 120 mg L-1 of Cu was extracted from the mesocarp. Mesocarp digestions were performed with addition of Fe, Cu and Zn and only 48% of Fe, 65% of Cu and 75% of Zn were recovered. Digestions containing the same concentration of the elements were performed with additions of Ca and Mg. The presence of Ca decreased the recovery of Fe and Zn. For simulating a real situation, gastrointestinal digestion of milk, milk and mesocarp mixture and milk and lignin mixture were performed. Although the iron present in milk had low availability, when it was mixed with mesocarp it was observed an increase in the concentration of soluble Fe. Compounds present in milk or in mesocarp can increase the solubility of the elements. It was not possible to observe if there was any change in the availability of Cu present in milk in the presence of mesocarp, because the Cu concentration in the milk is very low. The fraction of soluble Zn in milk and mesocarp mixture remained the same, but the dialysable fraction was practically null. Lignin addition increases the milk elements extraction. However, these elements remain not dialyzed

O objetivo deste trabalho foi avaliar a disponibilidade de Fe, Cu e Zn em presença de mesocarpo de babaçu, pois o mesocarpo contem compostos antinutricionais (fitato e lignina) que podem diminuir a disponibilidade de nutrientes minerais. As concentrações de Fe, Cu e Zn encontradas no mesocarpo de babaçu foram 17, 7,0 e 2,6 µg g-1, respectivamente. Para um melhor entendimento das interações dos elementos com o mesocarpo, foi realizada a extração de substâncias complexantes com soluções de NaOH (pH 7 e 12). Análises por espectrometria de absorção molecular indicaram que o principal complexante presente no extrato era o fitato. Soluções de Fe, Cu e Zn (10 a 300 mg L-1) foram adicionadas aos extratos, para que, após agitação e centrifugação, fossem determinadas as recuperações desses elementos. Os resultados mostraram que os elementos formam complexos com o fitato e a solubilidade desses complexos é dependente da razão elemento/fitato, quanto maior essa razão menor é a solubilidade. Também foi avaliada a interação dos elementos com o sólido remanescente da extração. Fe, Cu ou Zn foram adicionados ao mesocarpo lavado. Não foi possível afirmar que houve interação do mesocarpo lavado com Fe e Cu devido à precipitação decorrente da hidrólise. Porém, o Zn não sofreu hidrólise e teve baixas recuperações, indicando que houve interação com o sólido. Além disso, pode ser observado que ao adicionar maiores concentrações de Zn obtiveram-se melhores recuperações, provavelmente devido à saturação dos sítios de ligação. Para os estudos de disponibilidade, foi utilizado o procedimento de digestão simulada in vitro da US Pharmacopeia. Fe e Zn não foram extraídos durante a digestão do mesocarpo, portanto esses elementos não estariam disponíveis para absorção pelo organismo. Entretanto, cerca de 120 µg L-1 de Cu foi extraído do mesocarpo. Foram realizadas digestões do mesocarpo com adição de Fe, Cu e Zn e apenas 48% do Fe, 65% do Cu e 75% do Zn foram recuperados. Digestões contendo essa mesma concentração dos elementos foram realizadas com adições de Ca e Mg. A presença do Ca diminuiu a recuperação de Fe e Zn. Para simular uma situação próxima ao real, foram realizadas digestões gastrointestinais de leite, mistura de leite e mesocarpo e mistura de leite e lignina. Apesar do ferro presente no leite apresentar baixa disponibilidade, quando o mesmo foi misturado ao mesocarpo observou-se um aumento da concentração de Fe solúvel. Compostos presentes no leite ou no mesocarpo podem aumentar a solubilidade dos elementos. Não foi possível observar se havia alguma alteração da disponibilidade do Cu presente no leite na presença do mesocarpo, pois a concentração de Cu no leite é muito baixa. A fração de Zn solúvel na mistura de leite e mesocarpo permaneceu a mesma, porém a fração dialisável foi praticamente nula. A adição de lignina ao leite aumentou a extração dos elementos. Entretanto, esses elementos continuaram não sendo dialisados / The aim of this study was to evaluate the availability of Fe, Cu and Zn in the presence of mesocarp of babassu because it contains antinutritional compounds (phytate and lignin) that may decrease the availability of mineral nutrients. The concentrations of Fe, Cu and Zn found in the mesocarp of babassu were 17, 7.0 and 2.6 mg g-1, respectively. For a better understanding of elements interactions with mesocarp, it was performed the extraction of complexing substances with NaOH solutions (pH 7 and 12). Analysis by molecular absorption spectrometry indicated that the main complexing present in the extract was the phytate. Solutions of Fe, Cu and Zn (10 to 300 mg L-1) were added to the extracts for determining the recoveries of these elements, after agitation and centrifugation. The results showed that the elements form complexes with phytate and the solubility of these complexes is dependent on the ratio element / phytate, the greater this ratio the lower the solubility. The elements interaction with the remaining solid from extraction was also evaluated. Fe, Cu or Zn were added to the washed mesocarp. It was not possible to affirm that there was interaction of washed mesocarp with Fe and Cu due to precipitation derived from hydrolysis. However, Zn did not suffer hydrolysis and had low recoveries, indicating that there was an interaction with the solid. Furthermore, it can be seen that adding higher concentrations of Zn, better recoveries were obtained, probably due to saturation of the binding sites. For the studies of availability, it was used an in vitro simulated digestion procedure from U.S. Pharmacopeia. Fe and Zn were not extracted during digestion of the mesocarp, therefore these elements would not be available for absorption by the organism. However, about 120 mg L-1 of Cu was extracted from the mesocarp. Mesocarp digestions were performed with addition of Fe, Cu and Zn and only 48% of Fe, 65% of Cu and 75% of Zn were recovered. Digestions containing the same concentration of the elements were performed with additions of Ca and Mg. The presence of Ca decreased the recovery of Fe and Zn. For simulating a real situation, gastrointestinal digestion of milk, milk and mesocarp mixture and milk and lignin mixture were performed. Although the iron present in milk had low availability, when it was mixed with mesocarp it was observed an increase in the concentration of soluble Fe. Compounds present in milk or in mesocarp can increase the solubility of the elements. It was not possible to observe if there was any change in the availability of Cu present in milk in the presence of mesocarp, because the Cu concentration in the milk is very low. The fraction of soluble Zn in milk and mesocarp mixture remained the same, but the dialysable fraction was practically null. Lignin addition increases the milk elements extraction. However, these elements remain not dialyzed

The objective of this thesis was to investigate the effect of age and feeding length on phytase efficacy in broiler chickens during the starter phase. Two studies were carried out to evaluate this objective.<br>Study 1 was a randomized complete block design with 4 × 5 factorial arrangements of treatments. There were four diets; a positive control (PC), negative control (NC) and two phytase supplemented diets with inclusion levels of 1,000 and 2,000 phytase units/kg. There were five age and duration of feeding groups; Three 2-d feeding lengths terminated at d 8, 14, and 22 (d 6 to 8, d 12 to 14, and d 20 to 22), a 5-d feeding length terminated at d 14 (d 9 to 14) and a 16-d feeding length terminated on d 22 (d 6 to 22). Growth performance and sample collections were collected at the end of each phase i.e. d 8, 14 and 22. There was a difference (P < 0.01) in weight gain, feed intake, and feed efficiency between birds fed the PC diets and birds fed the NC diets across all groups as birds on the NC diets had lower performance (P < 0.05) than birds on the PC diet. However, birds fed the phytase supplemented diets had higher (P < 0.05) growth performance compared with birds fed the NC diet across all groups. Similarly, phosphorus (P) and calcium (Ca) digestibility and retention of birds fed the NC were lower (P < 0.05) as compared with birds fed the PC diet while birds fed the phytase supplemented diets had higher mineral digestibility and retention (P < 0.05) compared with birds on the NC diet. Age effect was evaluated by comparing the performance of birds fed the experimental diets for 2 d until d 8, 14, and 22. Birds fed until d 14 had the highest impact of the NC diet on mineral utilization, and the largest improvement of phytase on mineral utilization as compared with birds fed until d 8 and 22. Similarly, when feeding length effect was considered, birds fed for a shorter period had greater response to phytase (P < 0.05) on nutrient utilization than birds fed for a longer period at d 14 and 22. Tibia ash was higher (P < 0.05) in birds fed phytase supplemented diets for a longer period (i.e. 16 d) compared with birds fed or 2 or 5 d. The results from this study observed that age and duration of feeding influenced phytase efficacy especially in younger birds fed for a short period. However, it could not be determined if feeding birds for a short period at different ages in the starter phase would have a similar effect.<br>In study 2, the effects of age and feeding low P diets to birds for a short period of time on phytase efficacy and super dosing were evaluated at two critical points in the starter phase. This study had 3 × 2 × 2 factorial arrangements of treatments comprising 3 diets; a PC, NC, and a NC with phytase supplemented at 2,000 phytase units/kg; and 2 ages (i.e d 14 and 22) and 2 feeding lengths (i.e 2-d and 5-d). Thus, birds were fed the experimental diets from d 12 to 14, 9 to 14, 20 to 22, and 17 to 22 respectively. Results observed were similar to the first study. Birds fed the NC diet had lower (P < 0.01) performance as compared with birds fed the PC diets across all age and feeding length groups. Similarly, birds fed diets with the super dose level of phytase had greater growth performance (P < 0.01) compared with birds fed the NC diets. When age effect was considered, birds fed for 2 or 5 d until d 14 had the greatest improvements of phytase on nutrient utilization and bone mineralization compared with birds fed for both periods until d 22. When effect of feeding was considered, birds fed for 2-d at both ages had greater responses to phytase in performance and nutrient utilization compared with birds fed for 5-d at both ages. Plasma myo-inositol was higher (P < 0.01) in birds fed the super dose level of phytase compared with birds fed the NC diet.<br>In summary, we could conclude that the efficacy of phytase both at 1,000 and 2,000 FTU/kg was higher in birds fed for 2 d until d 14 as compared with the other groups. This could potentially help in designing studies to evaluate new phytase products or for comparing the efficacy of phytase from various sources. Feeding broiler chickens during the suggested time phase would potentially reveal the maximum efficacy of the phytase product.<br>

Accurate quantification of phosphorus (P) fractions in manures and agricultural runoff is vital to understanding P dynamics in the environment. Phytate is an organic form of P, with 6 phosphate groups, which is found in manures, but is difficult to quantify due to its affinity for complex formation with aluminum (Al) and iron (Fe). The objective of the first study was to measure if high concentrations of aluminum (Al) and iron (Fe) hinder accurate quantification of phytate in dairy manure and broiler litter when measured by high performance ion chromatography (HPIC). In this study dairy manure and broiler litter samples were spiked with Al, Fe, and phytate. Samples were alkaline extracted, acidified,cleaned up and filtered, and then phytate was analyzed with HPIC. High concentrations of Fe did not hinder phytate recovery in manure or litter samples. While phytate recovery was close to 100% at typical manure and litter Al concentrations, high concentrations of Al inhibited phytate recovery in litter samples and in some manure samples. Overall, alkaline extraction of dairy manure and broiler litter and analysis with HPIC proved to be a relatively accurate, fast and cheap within normal Al and Fe ranges, compared to the commonly used NMR method. The developed method was then used to measure the concentration of phytate P (Pp) rather than the entire phytate molecule (6 moles Pp per mole phytate) in runoff waters and in an adsorption study. Phytate P has not been previously measured in runoff, due to the low concentrations of Pp found in runoff and the complexity Pp extraction from runoff. Runoff treatments were dairy manure with and without added Pp (DM, DM+Pp), broiler litter with and without added Pp (BL, BL+Pp), and control, with and without two levels of Pp (control, control Pp low, control Pp high). Runoff was collected under simulated rainfall and analyzed for total and dissolved reactive P (DRP). Runoff was also separated into sediment (>0.45"m) and liquid (<0.45"m and lyophilized) fractions for Pp analysis by high performance ion chromatography (HPIC). The new method for Pp analysis recovered 70% of Pp spiked into runoff samples. Phytate P was found only on the sediment fraction of runoff and was not detected in the liquid fraction, even when it was lyophilized and extracted. This agreed with the adsorption study, which showed strong adsorption of Pp. Phytate P loss from control Pp high was significantly greater than Pp loss from control. �When control Pp high was removed from analysis, Pp loss from BL, BL+Pp, DM, and DM+Pp was significantly greater than from control. Phytate P in the manures appeared to behave differently than pure Pp salts, likely to do the formation of protein-phytate and cation-phytate complexes in the manures. Phytate P had no effect on DRP in runoff for any treatment, indicating no significant release of inorganic P through competitive binding. The majority of P lost in runoff was in fractions other than DRP and Pp in the sediment fraction. Efforts to control the erosive loss of soil during rainfall events appear to be the best way to reduce total P loss, irrespective of the presence of Pp. / Master of Science

Orientador: Gabriela Alves Macedo / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-14T10:38:34Z (GMT). No. of bitstreams: 1 Schons_PatriciaFernanda.pdf: 1271406 bytes, checksum: 668e62b416afd72ff47587e7425cecd0 (MD5) Previous issue date: 2009 / Resumo: Este trabalho visou avaliar o efeito do fungo Paecilomyces variotti e das enzimas fitase e tanase sobre fatores antinutricionais, taninos e fitatos presentes no sorgo tipo vassoura. A enzima tanase produzida pelo fungo Paecilomyces variotti apresentou atividade enzimática de 626U/g, além desta enzima, estavam presentes no extrato enzimático, com menor atividade protease, carboximetilcelulase, polimetilgalacturonase e fitase. A fitase empregada foi comercial (NATUPHOS ¿ BASF) e apresentou atividade de 660U/g. O sorgo utilizado neste estudo possui 13,6% de proteína, 4,2% de lipídeos, 1,7% de cinzas e 80,5% de carboidratos, elevada concentração de compostos fenólicos, especialmente taninos condensados e 226mg/100g de sorgo de fósforo inorgânico. O sorgo foi preparado e tratado com Paecilomyces variotti, tanase e fitase em diferentes combinações. As respostas diminuição de tanino e aumento de fósforo foram avaliados, os melhores resultados foram obtidos quando a farinha de sorgo foi tratada com tanase e fitase durante 5 dias. Por meio de planejamento experimental foi feito estudo do tratamento de sorgo com as enzimas fitase e tanase, o qual indicou a relação entre sorgo:água, como sendo a variável mais significativa para o processo. O processamento (maceração e fervura) e adição das enzimas foram efetivos na redução de taninos e aumento de fósforo, quando comparamos estes resultados com o sorgo cru, porém, as diferentes concentrações das enzimas não tiveram efeito sobre os parâmetros avaliados. Baseado nos resultados expostos, o sorgo para a elaboração das dietas foi tratado com a concentração inferior de enzimas estudada (100U/Kg). O estudo in vivo mostrou que a dieta elaborada com sorgo tratado enzimaticamente foi efetivo na melhoria da digestibilidade aparente de fósforo, na diminuição da excreção de fósforo, resultaram em melhores índice bioquímico para glicose e colesterol, apresentaram uma menor atividade de enzimas hepáticas aspartato aminotrasnferase (AST) e alanina aminotransferase (ALT), comparando com sorgo cru. No entanto, não teve efeito significativo na melhoria do ganho de peso, da digestibilidade aparente de proteína, na concentração de fósforo, ferro e cálcio plasmático, bem como na concentração de mineral contido no fêmur / Abstract: This work aimed assess the fungus Paecilomyces variotti and enzymes phytase, tannase effects on the antinutritional factors, tannins and phytates present in sorghum. The tannase produced by the fungus Paecilomyces variotti has enzymatic activity of 626U/g, in addition this enzyme were present in the enzymatic extract protease, in less activity protease, carboxymetilcellulase, polymetilgalacturonase and phytase. The phytase used is commercially (NATUPHOS ¿ BASF) and has activity of 660U/g. The sorghum utilized in this study has 13.6% protein, 4.2% of lipids, 1.7% ash and 80.5% carbohydrate, high concentration of phenolic, mainly condensed tannins and inorganic phosphorus 226mg/100g sorghum. The sorghum was prepared and treated by using Paecilomyces variotti, tannase and phytase in different combinations. The decrease in the tannins and increase in the phosphorus were evaluated, the best results were obtained when sorghum flour was treated with tannase and phytase for 5 days. Through experimental design were made a study about sorghum treated by the enzymes tannase and phytase, which indicated the relationship between sorghum:water, as the most significant variable in the process. The processing (soaking and boiling) and addition of enzymes were effective in reducing tannin and increase phosphorus, when comparing these results with the raw sorghum, however, different concentrations of the enzymes had no effect on the parameters evaluated. Based on the results above, the sorghum for the preparation of the diets were treated with lower concentration of enzymes studied (100U/Kg). The in vivo assay showed that the diet prepared using sorghum treated enzimaticaly was effective in improving digestibility of phosphorus, reducing the phosphorus excretion, resulting in better biochemical index for glucose and cholesterol, showed a lower activity of liver enzymes aspartate aminotransferase (AST) and alanin aminotransferase (ALT), compared with raw sorghum. However, it had no significant effects in improve weight, the apparent digestibility of protein, and the concentration of phosphorus, iron and calcium in the plasma and mineral content in the femur / Universidade Estadual de Campi / Ciência de Alimentos / Mestre em Ciência de Alimentos

The role of dietary phytate (inositol hexaphosphate) in the regulation of intestinal mucosa phytase was investigated in chicks. Seven-day-old chicks were grouped by weight into six blocks of three cages with six birds per cage. Three purified diets [a chemically defined casein diet, a chemically defined casein diet plus sodium phytate (20 g/kg diet) and a chemically defined casein diet plus sodium phytate (20 g/kg diet) and microbial phytase (1000 units/kg diet)] were randomly assigned to cages within each block. Chicks were fed experimental diets from 8 to 22 days of age then killed, and duodenal mucosa and left tibia removed. Phytase activity in duodenal mucosa, growth performance and bone ash content were determined. Addition of phytate to the chemically defined casein diet reduced (p < 0.05) the Vmax of the duodenal brush border phytase, but the Km of the enzyme was not affected. Addition of phytate also reduced (p < 0.05) weight gain, feed intake, feed efficiency and percentage ash. Addition of microbial phytase fully restored the feed efficiency (p < 0.05), but Vmax and body weight gain were only partially restored (p < 0.05). In conclusion, it would seem that dietary phytates non-competitively inhibit intestinal mucosa phytase.

The objective of this dissertation was to investigate areas that needed further research with regards to phosphorus, phytate, and phytase in monogastric nutrition. To fulfill this objective, a total of 6 studies were carried out.<br>Study 1 was designed to evaluate the additivity of apparent ileal digestibility (AID) and standardized ileal digestibility (SID) of phosphorus (P) in mixed dies containing corn and soybean meal (SBM) and fed to broiler chickens. There were 7 dietary treatments in a randomized complete block design with body weight (BW) serving as the blocking factor. Treatments consisted of 4 semi-purified diets prepared to contain either corn or SBM as the sole source of P with or without phytase supplementation at 1,000 FYT/kg; 2 mixed diets containing corn and SBM with or without phytase addition; and a P-free diet (PFD) formulated to determine the basal endogenous loss (BEL) of P. A total of 512 day-old broiler chicks were fed a commercial starter diet for 21 days while experimental diets were fed for 3 days (day 22-24 post hatching). The BEL of P in broiler chickens as determined by the PFD was 166 mg/kg dry matter (DM). The SID of P in corn and SBM was 52.2 and 65.4 %, respectively. Phytase addition improved (P < 0.05) both the AID and SID of P in the ingredient and mixed diets. There was no difference between the predicted and determined AID and SID of P in the mixed diets and were thus additive. It was concluded that the AID and SID of P in corn and SBM were additive with or without phytase addition. However, we could not be certain if the assumption of additivity will hold true in mixed diets containing ingredients with a higher phytate content and if age of birds affected the BEL of P.<br>In the 2nd study, the additivity of AID and SID of P in mixed diets containing corn and canola meal (CCM) was investigated in broiler chickens at 2 ages. A total of 588 broiler chickens was used in this study. Dietary treatments were arranged as a 2 × 3 × 2 factorial with 2 ages (day 13 and 21), 3 diets (corn, canola meal, and CCM), and 2 levels of phytase (0 and 1,000 FYT/kg) in a randomized complete block design. A PFD was fed to birds at both ages to determine the effects of age on the BEL of P. A commercial starter diet was fed from day 1 to 10 or 18 post hatching and then experimental diets were fed for 3 days until day 13 or 21, respectively. The AID and SID of P was higher (P < 0.05) in birds at day 13 as compared with birds at day 21 regardless of phytase addition. Similarly, the BEL of P in younger birds was higher (P < 0.01) than in older birds at d 21. Regardless of age or phytase supplementation, the predicted and determined AID and SID of P in the CCM diets were similar and thus additive. In conclusion, age influenced the BEL of P and the utilization of minerals in broiler chickens while the assumption of additivity held true when mixed diets containing CCM was fed to birds at both ages with or without phytase addition.<br>In order to evaluate the efficacy of a new consensus phytase variant, the 3rd and 4th studies investigated the responses of broiler chickens to varying concentrations of phytate phosphorus (PP) and the new consensus bacterial 6-phytase variant (PhyG) in the starter and grower phases, respectively. Responses evaluated included growth performance, tibia ash, AID and total tract retention (TTR) of nutrients. A total of 1,152 and 768 day-old male broiler chickens were used in the starter and grower phases, respectively. Dietary treatments were arranged as a 3 × 5 + 1 factorial with a nutrient-adequate positive control (PC) (2.8 g PP/kg) and 15 nutrient-reduced negative control (NC) diets with 3 levels of phytate (NC1, NC2, and NC3 with 2.3, 2.8, and 3.3 g PP/kg respectively), and 5 levels of PhyG (0, 500, 1,000, 2,000, and 4,000 FYT/kg). Rice bran served as the main source of PP in the experimental diets. All treatments had 6 replicates with 12 birds/cage in the starter phase (day 1 to 11 post hatching) and 8 birds/cage in the grower phase (day 12 to 23 post hatching). Birds fed the PC had greater responses (P < 0.05) as compared with birds fed the NC2 diets without phytase at the starter and grower phases. There was no interaction between PP and PhyG on responses of birds in the starter phase but there was an interaction effect (P < 0.05) on the AID of some amino acids (AA) in birds at the grower phase. Increasing levels of PP reduced (P < 0.05) the growth performance, Ca and P utilization of birds but had no effect on the tibia ash in the starter phase and grower phases. Phytase supplementation improved (P < 0.05) the growth performance, AID and TTR of nutrients, and tibia ash of birds in the starter and grower phases. In conclusion, increasing PP levels reduced some responses of birds in the starter and grower phase while PhyG inclusion improved all responses of broiler chickens at both phases.<br>The time effects of phytase on the P utilization of growing and finishing pigs was investigated in the 5th and 6th studies, respectively using growth performance, apparent total tract digestibility (ATTD) of nutrients, P excretion, and plasma mineral concentrations as evaluation parameters. In both studies (Exp 1 and 2), treatments were arranged as a 3 × 4 factorial in a randomized complete block design with 3 corn-SBM based diets including a P-adequate PC, a low-P NC, and NC + 1,000 FYT/kg; and 4 sampling time points at days 7, 14, 21, and 28 in Exp 1 and days 14, 26, 42, and 55 in Exp 2. Ninety-six growing pigs were used in both trials with an average BW of 19.8 ± 1.16 kg in Exp 1 and 49.8 ± 3.21 kg in Exp 2. Each treatment had 8 replicates evenly divided between barrows and gilts, and 4 pigs per pen. There was an interaction (P < 0.01) between time and diet on some growth performance parameters in pigs in Exp 1 but none in Exp 2. Pigs fed the PC had greater (P < 0.05) growth performance and ATTD of nutrients as compared to pigs fed the NC in both trials. Phytase supplementation improved (P < 0.05) the ATTD of P and Ca in pigs as compared with pigs fed the NC. There was an interaction (P < 0.05) between time and diet on the total and water-soluble P lost from pigs in Exp 1. Supplementing the NC with phytase reduced the water-soluble P by 45, 32, and 35 % over the growing, finishing and the entire grow-finish period, respectively. In Exp 2, plasma concentrations of P were increased (P < 0.05) over time. In conclusion, phytase improved the responses of growing pigs however, some of these responses were influenced over time.<br>In summary, the AID and SID of P in corn, SBM, and canola meal are additive in complete diets with or without the inclusion of phytase. The BEL of P could be affected by the age of birds. A new consensus phytase variant seems to be efficient in improving the productivity of broiler chickens regardless of the presence of varying concentrations of PP and the growth phase of the birds. Phytase supplementation improves the P utilization and reduces the P loss of growing and finishing pigs however, this may be influenced by time within each growth phase.<br><br>

Successful implementation of dietary P management strategies demand improved understanding of P digestion dynamics in ruminants and this is not possible without a reliable and accurate phytate (Pp) quantification method. The objective of the first study was to develop a robust, accurate, and sensitive method to extract and quantify phytate in feeds, ruminant digesta and feces. Clean-up procedures were developed for acid and alkaline extracts of feed, ruminant digesta and feces and clarified extracts were analyzed for Pp using high performance ion chromatography (HPIC). The quantified Pp in acid and alkaline extracts was comparable for feed but alkaline extraction yielded greater estimates of Pp content for digesta and feces than did acid extraction. Extract clean-up procedures successfully removed sample matrix interferences making alkaline extraction compatible with HPIC. The developed method was applied to investigate the disappearance of Pp from the large intestine of dairy heifers. Eight ruminally- and ileally-cannulated crossbred dairy heifers were used and each heifer was infused ileally with 0, 5, 15, or 25 g/d Pp and total fecal collection was conducted. On average 15% of total Pp entering the large intestine was degraded but the amount of infused Pp did not influence the degradability of Pp. Net absorption of P from the large intestine was observed. A feeding trial was conducted to investigate the effect of dietary Pp supply on ruminal and post-ruminal Pp digestion. Six ruminally-and ileally-cannulated crossbred lactating cows were used and dietary treatments were low (0.10% Pp), medium (0.18% Pp), and high (0.29% Pp) Pp, and a high inorganic P (Pi; 0.11% Pp; same total P content as high Pp). Ruminal Pp digestibility increased linearly with dietary Pp. As in the infusion study, net disappearance of Pp from the large intestine was only 16% of total Pp entering the large intestine and not influenced by dietary Pp. Fecal P excretion increased linearly with increasing dietary Pp but was not affected by form of dietary P. In lactating cows Pp digestibility was not affected by dietary Pp and fecal P excretion was regulated by total dietary P rather than by form of dietary P. / Ph. D.

Phytic acid, also known as myo-inositol hexakisphosphate or Ins(1,2,3,4,5,6)P6, is the major storage form of phosphorus in plant seeds. Phytic acid is poorly digested by non-ruminant animals such as swine and poultry, and it chelates mineral cations including calcium, iron, zinc, and potassium, classifying it as an anti-nutrient. The excretion of unutilized phytic acid in manure translates to an excess amount of phosphorus runoff that can lead to eutrophication of lakes and ponds. Understanding the phytic acid biosynthetic pathway will allow for the development of low phytic acid (lpa) soybeans by the down-regulation of specific genes. The goal of this research was to elucidate the pathway(s) for phytic acid biosynthesis in soybean (Glycine max). We have isolated several myo-inositol phosphate kinase genes in soybean as possible candidates for steps in the biosynthetic pathway. We have characterized the genes for four myo-inositol(1,3,4)P3 5/6-kinases (GmItpk1-4), one myo-inositol(1,4,5)P3 6/3/5-kinase (GmIpk2), and one myo-inositol(1,3,4,5,6)P5 2-kinase (GmIpk1). We have examined expression in developing seeds and other tissues by Northern blot analysis and quantitative RT-PCR. We have expressed all six genes as tagged fusion proteins in E. coli, and verified enzyme activity on the proposed substrates. For each enzyme, we have conducted biochemical characterization to determine enzyme kinetics and substrate specificities. We have verified in vivo activity of GmIpk2 and GmIpk1 by complementing yeast mutants in the respective genes. Our studies indicate the likelihood that three of the genes may be involved in phytic acid biosynthesis: GmItpk3, GmIpk2 and GmIpk1. For future work, to more fully understand the contribution of each kinase gene to phytic acid biosynthesis, an RNA interference approach will be employed. The gene sequences identified in this study will be used to construct silencing vectors for use in future transformation of soybean embryogenic cultures to determine the effects of down-regulation on myo-inositol phosphate profiles. / Ph. D.

Sucrose, raffinose, and stachyose make up the majority of the carbohydrates in soybean seeds. While sucrose is a desirable component of soybean seeds, raffinose and stachyose are considered to be anutritive factors and eliminating or reducing them appears to be a beneficial endeavour. The major objective of this study was to determine the genetic mechanism controlling accumulation of soluble saccharides in soybean seeds. An experimental soybean line, V99-5089, with high sucrose (14.6%) content coupled with low amounts of raffinose (0.5%) and stachyose (0.4%) was the center of this study. Three populations were studied and segregation patterns were observed in F2:3 populations. All three sugars were extracted by an aqueous procedure and quantified by high pressure liquid chromatography (HPLC) using a NH2 column and refractive index (RI) detector. Segregation of seeds from F2:3 plants indicated a single, partially recessive gene reduced stachyose content of soybean seeds from about 4% to less than 1%. Estimates of genetic variability indicate the presence of sufficient additive variation in addition to the putative major gene to warrant selection. Raffinose and stachyose were positively correlated to each other and each was negatively correlated with sucrose while there was not a significant correlation between total sugar content and the amount of any of the individual sugars. Agronomic traits evaluated do not appear to be adversely effected by the reduction of stachyose content. Additionally, a negative relationship was observed between inorganic phosphorus and stachyose content of soybean seeds but a relationship was not observed between stachyose and phytate phosphorus or between inorganic phosphorus and phytate phosphorus. / Master of Science