Physicochemical Properties of Rice Endosperm Proteins Extracted by Chemical and Enzymatic Methods

Rice proteins are nutritional, hypoallergenic, and healthy for human consumption. Efficient extraction with approved food‐grade enzymes and chemicals are essential for commercial production and application of rice protein as a functional ingredient. Rice endosperm proteins were isolated by alkali, salt, and enzymatic methods and evaluated for extractability and physicochemical properties. Alkali (RP_A) and salt (RP_S) methods extracted 86.9 and 87.3% of proteins with 65.9 and 58.9% yield, respectively. The enzymatic methods with Termamyl (RP_ET) and amylase S (RP_EA) extracted 85.8 and 81.0% proteins with 85.2 and 86.2% yield, respectively. Enthalpy values of RP_A (1.79 J/g), RP_S (1.22 J/g), RP_ET (nondetectable), and RP_EA (0.17 J/g), determined by differential scanning calorimetry, demonstrated that the varying level of denaturation of proteins depends on the method of extraction. Surface hydrophobicity data supported this observation. Alkali‐ and salt‐extracted proteins had higher solubility and emulsifying properties than those of enzyme‐extracted proteins. Comparatively, more favorable protein composition, lower surface hydrophobicity, higher solubility, and a lower degree of thermal denaturation of alkali‐ and salt‐extracted proteins contributed to higher emulsifying and foaming properties than those of enzyme‐extracted proteins; therefore, alkali‐ and salt‐extracted proteins can have enhanced functional use and a potential starting material for preparing tailored rice protein isolates.     

Hydrophobicity,Solubility, and Emulsifying Properties of Enzyme-Modified Rice Endosperm Protein

Rice endosperm protein was modified to enhance solubility and emulsifying properties by controlled enzymatic hydrolysis. The optimum degree of hydrolysis (DH) was determined for acid, neutral, and alkaline type proteases. Solubility and emulsifying properties of the hydrolysates were compared and correlated with DH and surface hydrophobicity. DH was positively associated with solubility of resulting protein hydrolysate regardless of the hydrolyzing enzyme, but enzyme specificity and DH interactively determined the emulsifying properties of the protein hydrolysate. The optimum DH was 6–10% for good emulsifying properties of rice protein, depending on enzyme specificity. High hydrophobic and sulfhydryl disulfide (SH-SS) interactions contributed to protein insolubility even at high DH. The exposure of buried hydrophobic regions of protein that accompanied high-temperature enzyme inactivation promoted aggregation and cross-linking of partially hydrolyzed proteins, thus decreasing the solubility and emulsifying properties of the resulting hydrolysate. Due to the highly insoluble nature of rice protein, surface hydrophobicity was not a reliable indicator for predicting protein solubility and emulsifying properties. Solubility and molecular flexibility are the essential factors in achieving good emulsifying properties of rice endosperm protein isolates.     

Recovery and characterization of α-zein from corn fermentation coproducts

Zeins were isolated from corn ethanol coproduct distiller's dried grains (DDG) and fractionated into α- and β γ-rich fractions. The effects of the ethanol production process, such as fermentation type, protease addition, and DDG drying temperature on zein recovery, were evaluated. Yield, purity, and molecular properties of recovered zein fractions were determined and compared with zein isolated from corn gluten meal (CGM). Around 29-34% of the total zein was recovered from DDG, whereas 83% of total zein was recovered from CGM. Process variations of cooked and raw starch hydrolysis and fermentation did not affect the recovery, purity, and molecular profile of the isolated zeins; however, zein isolated from DDG of raw starch fermentation showed superior solubility and film forming characteristics to those from conventional 2-stage cooked fermentation DDG. Protease addition during fermentation also did not affect the zein yield or molecular profile. The high drying temperature of DDG decreased the purity of isolated zein. SDS-PAGE indicated that all the isolated α-zein fractions contained α-zein of high purity (92%) and trace amounts of β and γ-zeins cross-contamination. Circular dichroism (CD) spectra confirmed notable changes in the secondary structure of α-zeins of DDG produced from cooked and raw starch fermentation; however, all the α-zeins isolated from DDG and CGM showed a remarkably high order of α-helix structure. Compared to the α-zein of CGM, the α-zein of DDG showed lower recovery and purity but retained its solubility, structure, and film forming characteristics, indicating the potential of producing functional zein from a low-value coproduct for uses as industrial biobased product.     

Preparation of Rice Endosperm Protein Isolate by Alkali Extraction

Rice endosperm extraction conditions were optimized by response surface methodology. The optimum alkali extraction conditions were pH 11.0 at 40°C for 3 hr with 8:1 solvent‐to‐solid ratio. The maximum protein yield was 43.1% at these conditions. As the extraction pH was increased from 9.0 to 12.0, protein extractability and content increased but the solubility and emulsifying properties of the extracted protein decreased. The extracted protein was recovered by either isoelectric precipitation (IEP) or ultrafiltration (UF). Ultrafiltering the supernatant with a 5‐kDa hollow fiber membrane concentrated the protein from 1.8 to 16% (dry basis) and the resulting solution was spray‐dried to produce a protein concentrate (RP_UF) with 71% protein. Although RP_IEP contained higher protein (86%) than RP_UF (71%), RP_UF showed higher solubility and emulsifying properties. The solubility of RP_UF was higher (37%) than RP_IEP (15%). RP_UF also demonstrated higher emulsion activity (0.414) and stability (22.4 min) compared with the emulsion activity (0.282) and stability (15.5 min) of RP_IEP. Higher solubility and the soluble nonprotein components of RP_UF contributed to higher emulsifying properties than RP_IEP. The UF provided milder extraction conditions with improved emulsifying properties than conventional IEP.     

Glycosylation and Deamidation of Rice Endosperm Protein for Improved Solubility and Emulsifying Properties

Rice endosperm protein was prepared by alkali-extraction method and subsequently modified by controlled glycosylation (RP_Glu, RP_XG), deamidation (RP_DA), and enzymatic hydrolysis by alcalase (RP_Alc) methods. The RP_Glu and RP_XG were prepared by Maillard type glycosylation with D-glucose and xanthan gum, respectively. The glycosylation improved the emulsion activity (0.721) and stability (26.8 min) of the protein but did not show a substantial improvement in solubility (39.7%). The rice protein modified by controlled alkali-deamidation (RP_DA) showed highest solubility (68%), emulsion activity (0.776), and emulsion stability (24 min) among the three protein modification methods evaluated in this study. The alcalase treatment to 1.8% DH (RP_Alc) slightly improved solubility (33%), emulsion activity (0.468), and emulsion stability (17.5 min) compared with unmodified rice protein (RP), which had 18% solubility, 0.266 emulsion activity, and 14.7 min emulsion stability. The glycosylation and deamidation methods were more effective than the controlled enzymatic hydrolysis by alcalase in improving solubility and emulsifying properties of rice endosperm protein. Glycosylated and deamidated rice endosperm proteins can find application in enhancing emulsifying properties in suitable products.     

Two fraction extraction of α-zein from DDGS and its characterization

Zein was recovered from corn distiller's dried grains with solubles (DDGS) by a modified method using 70% (w/w) aqueous 2-propanol (70-IPA) or 70% (v/v) aqueous ethanol (70-EtOH) solvents, and a commercial method using 88% (w/w) aqueous 2-propanol (88-IPA). Yield, purity, and film properties of the isolated zein were determined. The modified procedure extracted two fractions of zeins: a mostly α-zein fraction, and a mostly γ-zein fraction. The modified method increased α-zein yield from 4% to 14%. Enzyme cellulase pretreatment did not improve zein yield, but grinding did. The α-zein fraction showed electrophoretic bands at 40, 22, 19, and 10 kDa, corresponding to α-zein dimer, α₁-zein, α₂-zein, and δ-zein, respectively. The α-zein of DDGS retained its film forming capability. The α-zein film of unmodified DDGS was cloudy and rough, unlike the clear and smooth films of α-zeins isolated from corn gluten meal and enzyme-treated DDGS.