Soy flour was inoculated with Lactobacillus delbrueckii ssp. lattis R0187, and incubated for 8 h, to evaluate the protein hydrolysis and identify peptides generated by this fermentation, and the impact on ACE and trypsin inhibitory activities. Aqueous protein extracts prepared from different fermentation time periods showed a decrease in soluble protein content (from 2.83 to 0.02 mg/mL), while soluble inorganic nitrogen and free amino acid contents increased (from 0.029 to 0.062% w/w, and from 0.75 to 0.90% w/w, respectively). The protein extracts were analyzed by SDS-PAGE; proteolysis was observed after 5 h incubation of inoculated soy flour, suggesting that glycinin, beta-conglycinin, and trypsin inhibitors, were hydrolyzed. Peptides were isolated by tricine-SDS-PAGE, and analyzed by MS/MS; fragments of soy anti-nutritional factors (Kunitz and Bowman-Birk trypsir, inhibitors), as well as of other soybean proteins, were identified, confirming that these proteins were hydrolyzed. The protein extracts at time 0 h and 8 h were analyzed by RP-HPLC; one fraction was analyzed by MS/MS, which identified peptides from Lactobacillus species. Determination of trypsin inhibitory activity showed less inhibition of the enzyme with inoculated soy flour compared to the control (un-inoculated soy flour), confirming the deactivation of trypsin inhibitors by fermentation. Determination of ACE inhibitory activity showed a higher inhibition with the control (86% +/- 3.0) compared to inoculated soy flour (66% +/- 7.6).

Soybean production and consumption has increased in recent decades. However, trypsin inhibitor activity and “beany” flavour are two drawbacks limiting the utilization of soybean. In the present study, micronization, ethanol washing, and enzymatic hydrolysis (alone or in combination) were used to treat soybean. Micronization at 100 °C and 135 °C decreased the activity of both trypsin inhibitors (53% and 80% respectively), and lipoxygenase (51% and 99%, respectively). Ethanol increased the trypsin inhibitor activity while alcalase hydrolysis decreased its activity. Different treatment combinations affected trypsin inhibitor activity, with micronization having a major influence. “Beany” flavour related volatiles (hexanal, (E)- 2-hexenal, 1-hexanol, heptanal, (E)-2-octenal, (E)-2-nonenal, (E,E)-2,4-nonadienal, 2,4-decadienal, (E,E)-2,4-decadienal, 1-octen-3-ol, 2-pentylfuran and 3-octen-2-one) were significantly decreased with micronization. Ethanol effects varied with different volatiles. Soybean micronized at 135°C and washed with 65% ethanol was recommended for soybean processing due to its low trypsin inhibitor activity and low “beany” related volatile content.

Digestive proteases were isolated from the pancreas of the stomachless cunner fish (Tautogolabrus adspersus) and characterized in terms of their physicochemical properties, their ability to hydrolyze native pectin methylesterase (PME) from orange and polyphenol oxidases (PPO) from mushroom and the ability of the cunner enzyme(s) to maintain the stability of orange juice cloud. / The cunner trypsin fraction exhibited exceptional capacity to hydrolyze native proteins versus the bovine trypsin. Incubation of native PME with cunner or bovine trypsin resulted in a loss of 75% or 35% in PME activity respectively. Similarly, a 75% or 55% loss in PPO activity was observed after treatment with cunner and bovine trypsin respectively. Bovine trypsin, however, hydrolyzed the heat-denatured PME and PPO better than the cunner trypsin. Also, there was no reactivation of both PME and PPO activity after treatment with either the cunner or bovine enzyme during storage at 4$ sp circ$C for 3 weeks. However, PPO retained up to 20% or 50% of the initial activity after treatment with cunner or bovine trypsin, respectively. / A 3 x 3 factorial design involving the factors of temperature, enzyme concentration and incubation time carried out gave an r$ sp2$ of 0.92 and 0.95 for cunner and bovine trypsin treated PME respectively. On the other hand, an r$ sp2$ of 0.91 and 0.94 was obtained for the combined effects using cunner and bovine trypsin for PPO inactivation. Validation of the model of PME inactivation measured as the % cloud remaining revealed that the cunner trypsin fraction upheld the cloud stability of cloud juice better than bovine trypsin, with cunner trypsin retaining more than 90% of the cloud whereas the juice treated with bovine trypsin only resulted in a 70% retention of the juice cloud. (Abstract shortened by UMI.)

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2002. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Missouri-Columbia, 1996. / Typescript. Vita. Includes bibliographical references (leaves 135-140). Also available on the Internet.

Thesis (Ph. D.)--University of Missouri-Columbia, 1996. / Typescript. Vita. Includes bibliographical references (leaves 135-140). Also available on the Internet.

Thesis (Ph.D) - University of Queensland, 2006. / Includes bibliography.

Orientador: Maria Ligia Rodrigues Macedo / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-23T15:21:13Z (GMT). No. of bitstreams: 1 Guimaraes_LaysCordeiro_M.pdf: 5932501 bytes, checksum: 2972555d649882421e88bba9ccd5dfea (MD5) Previous issue date: 2013 / Resumo: O resumo poderá ser visualizado no texto completo da tese digital / Abstract: The abstract is available with the full electronic document / Mestrado / Bioquimica / Mestra em Biologia Funcional e Molecular

<p>For this dissertation, four experiments were conducted to evaluate the effect of dietary addition of exogenous protease and amylase enzymes on growth performance and nutrient utilization in broiler chickens. An additional fifth experiment was designed to determine the role of central and peripheral appetite regulators in birds fed diets deficient in dietary phosphorus (P). This arose from consistent reports in literature of a direct effect of dietary P concentration on feeding response in broiler chickens. </p><p>Experiment 1 examined the growth performance and protein utilization responses of broiler chickens to purified trypsin inhibitors (TI) and exogenous protease additions. Experimental diets were arranged as a 2 × 2 factorial with factors being dietary TI (1,033 or 10,033 TIU/g) and exogenous protease (0 or 15,000 PROT/kg). Protease supplementation improved BW gain (<i>P < </i>0.01) and gain to feed ratio (<i>P < </i>0.05) of birds. The relative weight of pancreas increased (<i>P < </i>0.05) with added TI on d 14 and 21 but was reduced (<i>P < </i>0.001) with protease supplementation. Apparent ileal digestibility (AID) of all amino acids (AA), except methionine, decreased (<i>P < </i>0.001) with added TI, but increased (<i>P < </i>0.05) with protease supplementation. Duodenal trypsin and chymotrypsin activities were reduced (<i>P < </i>0.05) with added TI but increased <i>(P < </i>0.01) with protease supplementation. It was concluded that dietary addition of purified TI negatively affects nutrient utilization by broiler chickens and that the efficacy of the exogenous protease might be independent of dietary TI concentration. A follow-up experiment was conducted (Experiment 2) to evaluate the impact of TI and exogenous protease supplementation on endogenous AA loss in broiler chickens. Four diets were arranged as a 2 × 2 factorial with factors being dietary TI (0 or 8,000 TIU/g) and exogenous protease (0 or 15,000 PROT/kg). There was no effect of TI, exogenous protease, or their interaction on growth performance of birds. Endogenous nitrogen (N) loss and all AA (except Cys) increased (P < 0.05) due to added dietary TI. Exogenous protease had no effect on endogenous loss of N and all AA. The AID of Ca, Fe, Mg, Mn, and Cu was reduced (P < 0.05) by added dietary TI. Protease supplementation improved the AID of Cu (P < 0.01) and K (P < 0.05). Secretion of crude mucin and sialic acid (g/kg DM intake) increased (P < 0.05) with increased dietary TI and was not recovered by protease supplementation. It was concluded from this study that TI increases the endogenous loss of AA, reduces the digestibility of minerals in broiler chickens, and that exogenous protease had no effect on endogenous AA flow, irrespective of added dietary TI. </p><p>In Experiment 3, the responses of broiler chickens fed corn-soybean meal-based diets to dietary α-amylase supplementation during 4 growth phases were evaluated. Birds were assigned to 8 treatment diet in a 2 × 4 factorial arrangement of 2 dietary levels of α-amylase supplementation (0 or 80 kilo-Novo alpha amylase units (KNU) per kg diet) and 4 post hatching growth phases (d 0 to 11, d 11 to 21, d 21 to 42, or d 42 to 56). Body weight gain and feed efficiency of birds improved (<i>P</i> < 0.01) with α-amylase supplementation. There were main effects of α-amylase, growth phase and interaction (<i>P</i> < 0.01) on AID of starch. The total tract retention (TTR) of starch increased (<i>P</i> < 0.05) with amylase supplementation but was not different across growth phases. Amylase supplementation improved (<i>P</i> < 0.05) gross energy utilization in birds, and specifically, during d 11 to 21 post hatching, the viscosity of jejunal digesta and pancreatic amylase activity increased (<i>P</i> < 0.01) with amylase supplementation. The conclusion from the study was that the growth phase of birds may affect the response to exogenous amylase. Following the result of this study, Experiment 4 was conducted to evaluate the effect of amylase supplementation on starch and energy digestibility at various intestinal sites in broiler chickens. Experimental diets comprised 3 concentrations of α-amylase supplementation (0, 80, or 160 KNU/kg diet) and sampling was done on 4 intestinal sites: anterior jejunum (AJ), posterior jejunum (PJ), anterior ileum (AI) and posterior ileum (PI). There were linear and quadratic (<i>P</i> < 0.01) responses of increasing α- amylase supplementation on starch and energy digestibility at the PJ and AI, with only linear effects on TTR of starch (<i>P</i> < 0.05). A linear increase in starch disappearance and digestible energy (kcal/kg) was observed (<i>P</i> < 0.01) with digesta flow from AJ to PJ with increasing amylase supplementation, which may be related to the observed decrease in the viscosity of the jejunal digesta (<i>P</i> < 0.05). Results from this experiment demonstrate the efficacy of exogenous amylase to improve starch, and energy digestibility in broiler chickens, with the highest impact observed in the posterior jejunum.</p><p>A final study (Experiment 5) was conducted to evaluate the impact of dietary phosphorus (P) concentration on hypothalamic molecular regulation of appetite by broiler chickens. Birds were randomly assigned to 3 experimental diets which contained 1.2 (P-deficient), 2.8 (P-marginal) or 4.4 (P-adequate) g/kg non-phytate P (nPP). A decrease in feed intake and BW gain was observed (P < 0.001) in birds fed the P-deficient diet. There was upregulation (P < 0.05) in the mRNA expression of Sodium-phosphate cotransporter (NaPi-IIb), anorexia-related hypothalamic cholecystokinin receptor (CCKAR) and melanocortin receptors (MC3R and MC4R) in birds fed P-deficient diets, whereas cholecystokinin (CCK) mRNA was downregulated (P < 0.01). It may be concluded that a deficiency in dietary P decreases feed intake in broiler chickens by altering the expression of anorexigenic genes in the gut and hypothalamus.</p>

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