Browsing by Subject "Functionality"

Now showing 1 - 3 of 3
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    Allergenicity and functionality of soy protein subjected to limited enzymatic hydrolysis and Maillard-induced glycation
    (2016-08) Hinnenkamp, Chelsey
    Increasing consumer demand for dietary protein coupled with the need for functional proteins is driving the growth of the soy protein market. However, the expanded use of soy protein is limited by its distinction as one of the “Big 8” food allergens. Thus, there exists a need for a functional, hypoallergenic soy protein ingredient. Previous protein modification approaches have demonstrated varying levels of success in producing a functional, hypoallergenic protein ingredient, with limitations including feasibility, consumer acceptability, loss of functionality, and residual immunoreactivity. A potential solution is to combine two controlled modification approaches, limited enzymatic hydrolysis with partial Maillard-induced glycation. We hypothesize that optimization of limited enzymatic hydrolysis and Maillard-induced glycation conditions will reduce the allergenicity of soy protein, while maintaining its functionality. Thus, the objectives of this study were twofold: (1) to optimize limited enzymatic hydrolysis and partial Maillard-induced glycation conditions to produce a hypoallergenic soy protein, and (2) to determine the effect of limited and targeted hydrolysis and further glycation on protein functionality. Limited (< 8%), targeted hydrolysis was achieved using alcalase, pepsin, and an alcalase/pepsin combination to produce, respectively, ASPH, PSPH, and AP_SPH samples with degrees of hydrolysis (% DH) ranging from 3.8 – 6.8%. Two levels of glycation were achieved, with 68 – 75% remaining free amines, for the mild condition and 55 – 68% for the advanced condition. Changes in immunoreactivity was monitored following qualitative, western blot, and quantitative, ELISA immunoassays, utilizing sera from seven individual with high levels (>3.5 kU/L) of soy protein specific IgE. A significantly higher overall L*a*b* color difference and visible browning confirmed that the advanced condition, compared to the mild condition, had propagated to the final stages of the Maillard reaction. To avoid browning, loss in nutritional quality, and formation of advanced glycation endproducts (AGE) associated with the final stages of the Maillard reaction, the mild glycation conditions were selected for functionality testing. The solubility, emulsification capacity (EC), emulsification activity index (EAI), emulsion stability (ES), gel strength, and water holding capacity (WHC) were tested for soy protein isolate (SPI) and hydrolysate (SPH) samples, as well as the glycated samples and glycation controls. Pepsin targeted the hydrolysis of glycinin, and alcalase targeted the hydrolysis of β-conglycinin. Limited enzymatic hydrolysis significantly reduced immunoreactivity, with reductions in immunoreactivity ranging from 20 – 75%, depending on the enzyme used and the variability among the sera tested. Maillard-induced glycation predominately modified the subunits of β-conglycinin, while having a variable effect on immunoreactivity. In some cases, the negative effect of glycation was due to partial denaturation (unfolding), which exposed epitopes originally hidden in interior of the protein. The solubility, at neutral pH and a 2.5% protein concentration, of SPH samples (66 – 68% protein solubility) decreased in comparison, to that of SPI (96% protein solubility). The effect of limited enzymatic hydrolysis and partial Maillard-induced glycation on emulsification and gelling properties was highly variable with factors such as enzyme choice, % DH, peptide profile, and extent and location of glycation influencing the observed results. For example, while hydrolysis reduced the ES and increased EAI, glycation improved the ES and had an opposite effect on EAI. Similarly, both modifications reduced gel strength of soy protein, while the WHC of PSPH and AP_SPH was improved upon glycation. Overall, this study demonstrated that limited enzymatic hydrolysis and partial Maillard-induced glycation, under specific conditions require further optimization to successfully reduce immunoreactivity (> 90% reduction to claim a hypoallergenic product) and improve functionality. Results highlighted the importance to consider and target individual variability in the immunoreactivity with the various modified proteins, when optimizing conditions to produce a hypoallergenic product. Additional research to optimize modification conditions and to understand the structural impact of these modifications on the protein structure is needed to achieve the goal of developing a hypoallergenic, functional soy protein ingredient.
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    Chemical Characterization, Functionality, and Baking Quality of Intermediate Wheatgrass (Thinopyrum intermedium)
    (2017-05) Rahardjo, Citra
    Environmental problems have exacerbated the challenges of food production. Soil erosion, run-off from irrigation, and greenhouse gas emissions have significantly impacted the ecosystem through their cumulative effects over decades. As the population increases, these aforementioned problems will grow in scale and scope. Thus, it is important to address these concerns by investigating sustainable solutions and conducting research on new approaches to food supply. Specifically, the development of perennial crops for food use would have environmental benefits such as reduced soil and water erosion. Further, the usage of perennial crops for food applications would provide alternatives to the current and ever diminishing food supply, provide incentives to farmers for planting these crops, and address consumers’ growing interest in sustainable food systems. One of several perennial crops available for potential food use is intermediate wheatgrass (IWG) (Thinopyrum intermedium). Little is known about the chemical and functional characteristics of IWG. Therefore, the overall objective of this work was to characterize the chemical and functional properties of IWG grains from multiple breeding lines for food applications, namely bread baking. Sixteen IWG experimental lines along with one bulk IWG sample and two wheat controls (Arapahoe and commercial hard red wheat) were analyzed for proximate composition following standard methodologies. Dietary fiber, total starch content, and percent damaged starch were determined using Megazyme kits. Amylose/amylopectin ratio and their molecular weight distribution were determined using size exclusion chromatography. Gluten forming proteins profile and molecular distribution were determined using gel electrophoresis and size exclusion chromatography. Dough rheology was assessed using a farinograph and a texture analyzer equipped with a Kieffer rig, while starch pasting properties were monitored using a rapid visco analyzer. Bread baking tests were performed following the AACCI 10-10.03 method. Compared to wheat controls, IWG samples had higher protein, dietary fiber, and ash contents, yet were lower in starch content and deficient in high molecular weight glutenins (HMWG), important protein components responsible for dough strength and elasticity. Specifically, wheat controls had more high molecular weight polymeric proteins (HMWPP), while IWG samples had more albumins and globulins. The ratios of amylose to amylopectin among the IWG samples and the wheat controls were similar. However, percent damaged starch was higher in the wheat controls than in IWG samples. On the other hand, the soluble to insoluble dietary fiber ratio was higher in wheat controls than in IWG samples. Dough rheology data showed that IWG dough was weaker than that of the controls. Farinograph and Kieffer data demonstrated that doughs made from wheat controls were more stable, more resistant to extension, and more extensible than doughs made from IWG samples. In terms of starch pasting properties, wheat controls had higher peak, hold, and final viscosities than the IWG samples, indicating the superiority of wheat controls over IWG samples as viscosity builders. The starch pasting properties data illustrated the samples’ behavior upon heating and cooling treatments, which are important characteristics to consider when evaluating IWG for commercial applications. Even though IWG had similar specific volumes to one of the wheat controls (hard red wheat), both wheat controls had a higher rising capability due to the wheat’s gluten network forming ability. Deficiency in HMWG and high fiber content of IWG samples, contributed to the poor gluten network and consequently inferior baking quality. Overall, results of this work suggest that IWG has a superior nutritional profile as compared to wheat, but poses challenges for baked products that require dough rising properties. Further studies on IWG, such as investigating the effect of conditioners on enhancing protein functionality, determining the effect of fiber on dough development, effect of blending with wheat, and exploring other food applications would enhance its potential utilization as a food crop. This research and future efforts will support breeders in their current screening and future breeding efforts for the development of IWG lines suitable for food applications.
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    The Effect of Limited Enzymatic Hydrolysis and Maillard-Induced Glycation on Sodium Caseinate Allergenicity, Bioactivity, and Functionality
    (2016-06) Liang, Lu
    With superior functional properties, especially emulsification properties, high nutritional value, and excellent physiological benefits, sodium caseinate has been widely used in the formulation of many food and beverage products. However, the major drawback of the expanded utilization of sodium caseinate is its allergenicity. Therefore, in order to prevent a decrease in the market value of casein ingredients, different protein modification techniques have been explored to reduce the allergenicity of casein. Extensive enzymatic hydrolysis (> 25% DH) can reduce the allergenicity of a protein. However, this technique is detrimental to both flavor and functionality. Limited enzymatic hydrolysis, on the other hand, can maintain the functionality, but may only reduce the immunoreactivity to a certain extent. Another protein modification technique that may reduce the allergenicity of proteins is Maillard-induced glycation. Glycation, however, should be properly controlled to prevent the formation of mutagenic compounds and brown pigments with adverse effects on functionality, digestibility, and bioactivity. It was hypothesized that combining the two mechanisms, proteolysis and Maillard glycation, under controlled and mild conditions, will have a dual effect on reducing allergenicity by altering potential epitopes, while maintaining nutritional quality, bioactivity and functionality. Thus, the objectives of this study were twofold: (1) Determine the effect of several proteolysis conditions and Maillard-induced glycation conditions on the allergenicity of sodium caseinate. (2) Determine the effect of the dual modification approach on the bioactivity and emulsification properties of sodium caseinate. Sodium caseinate isolate (SCI) was hydrolyzed using three enzymes separately, alcalase, neutrase and trypsin, following the pH-stat method, while maintaining limited degree of hydrolysis (DH 3.6-17 %). Different enzyme hydrolysis conditions, including enzyme to substrate ratio, incubation temperature, pH, and time were investigated. The sodium caseinate hydrolysate (SCH) with the highest emulsifying properties (emulsification capacity, stability and activity) and ACE-inhibitory activity was tested for remaining allergenicity. The same SCH was further glycated with maltodextrins at 60°C, water activity (aw) of 0.49, 0.63, or 0.79, and a 2:1 ratio of protein to maltodextrin over a period of 24-120 h. Sodium caseinate isolate was also glycated to determine the effect of glcation alone on modifying emulsification properties and allergenicity. Loss in free amine groups along with the formation of Amadori compounds and browning were monitored. Flouresence compound formation was monitored to determine progression of the reaction into intermediate stages before formation of brown pigments. Urea-PAGE gel was utilized to confirm the glycation formation. Select glycated samples were tested for allergenicity, emulsifying properties, and ACE-inhibitory bioactivity. ELISA was performed to determine the immunoreactivity of the samples using sera from five allergenic donors following ELISA. Trypsin was better than Alcalase and Neutrase in producing hydrolysates that retained emulsification properties. This might be due to trypsin’s narrow and unique specificity, where large molecular weight peptides with better amphiphilic properties were generated compared to those generated by Alcalase and Neutrase. Limited hydrolysis (5% DH) resulted in pronounced ACE inhibitory activity and reduced immunoreactivity (6%-45%). Differences in the reduced immunoreactivity were attributed to the sensitivity of sera used and the variation in milk-protein-specific IgE among the different sera that could react with the epitopes present in casein proteins samples and newly generated epitopes upon hydrolysis. Among all the glycation conditions studied, optimum conditions for the desired objectives were determined to be 96 h of incubation at 0.63 aw, in a 2:1 ratio of casein protein to maltodextrin, due to a moderate Amadori compounds formation, as noted by adsorbace at 304 nm, limited % amino group blockage (9.7%), minimal progression to intermediate stages, as noted by % fluorescence intensity, and limited progression to advances stages of the Maillard reaction, as noted by minimal browning measured at 420 nm. No significant (P < 0.05) effect on ACE inhibitory activity, emulsification properties and immunoreactivity was observed post glycation at at 0.63 aw. This is likely due to insufficient Amadori compounds formation and the interference from free maltodextrin and unadsorbed intact sodium caseinate, which can destabilize the emulsion through depletion flocculation. Results from this work, for the first time, provided valuable insights into the effect of proteolysis conditions and Maillard-induced glycation conditions on the allergenicity, emulsification properties, and ACE-inhibitory activity of sodium caseinate. Combining two different protein modification techniques under controlled and mild conditions could maintain the emulsification properties and ACE-inhibitory activity, but had no dual effect on reduction of immunoreactivity. Therefore, further optimization of glycation conditions needs to be conducted for the development of hypoallergenic casein ingredient with acceptable functionality and bioactivity.