Changes Occurring in Protein Body Structure and α-Zein During Cornflake Processing

Zeins, which comprise the majority of proteins in corn, are located in spherical organelles called protein bodies. Changes in protein body shape and release of encapsulated α-zeins as a result of cornflake processing (conventional pressed or extrusion flaking) were investigated. Size-exclusion chromatography, SDS-PAGE, and protein solubility tests showed that, upon cooking, zein proteins form large, disulfide-bound polymers, many of which were insoluble in nonreducing solvents. Transmission electron microscopy with immunogold staining revealed that cooking had no effect on protein body structure in corn, but after processing to cornflakes, protein body structure was altered. In conventional pressed cornflakes, the protein bodies were flattened, partially fused together, and α-zeins were to some degree released, whereas in the extruded flakes, protein bodies were completely disrupted and α-zeins dispersed. These results suggest that zeins in cornflakes, particularly extruded ones, are not confined to rigid protein bodies but can interact with each other and other components in the system. The disruption of protein bodies, zein release, and the chemical changes that proteins undergo during processing are speculated to be determinants of texture in ready-to-eat corn-based breakfast cereals.     

Immunochemical and Electrophoretic Analysis of the Modification of Wheat Proteins in Extruded Flour Products

Antibodies specific for wheat proteins were used to identify protein fractions modified during extrusion of Hard Red Spring wheat flour (14% protein) under four different combinations of extrusion conditions (18 and 24% feed moisture and 145 and 175°C die temperature). Antibody binding was assessed on immunoblots of proteins extracted from flour and extrudates separated by SDS‐PAGE. Antibodies to high molecular weight glutenin subunits (HMW‐GS) and to B‐group low molecular weight glutenin subunits (LMW‐GS) recognized intact subunits from both flour and extrudates. Antibodies to C‐group LMW‐GS had diminished binding to extruded proteins. Glutenin‐specific antibodies also recognized protein in the extrudates migrating as a smear at molecular weights higher than intact subunits, indicating cross‐linked proteins. Antibodies recognized albumins or globulins in flour but not in extrudates, evidence that these fractions undergo significant modification during extrusion. Acid‐PAGE and antibody reaction of gliadins extracted in 1M urea and in 70% ethanol revealed total loss of cysteine‐containing α, β, γ‐gliadins but no obvious effects on sulfur‐poor ω‐gliadins, suggesting gliadin modification involves replacing intramolecular disulfides with intermolecular disulfide cross‐links. Identifying protein fractions modified during different extrusion conditions may provide new options for tailoring extrusion to achieve specific textural characteristics.     

Extrusion-induced changes to the chemical profile and viscosity generating properties of citrus fiber

Each of 8 variants in extrusion conditions was applied to a commercially available citrus fiber. Extrusion under conditions where the specific mechanical energy (SME) exceeded 400 kJ·kg(-1) was able to solubilize up to 30% of the fibers. Where the SME was ～200 kJ·kg(-1) the degree of fiber solubilization was between 8 and 12%. All extruded fibers showed a loss of water-retaining capacity compared to the reference fiber, and this was attributed to the disruption of the integrated cell wall structure during the extrusion process. Nevertheless, within the 8 extruded variants there was a wide range of viscosity generating capacity which depended on the level of SME to which the fibers were subjected. The SME also had a pronounced effect on the nature of the solubilized fibers in terms of both their monosaccharide composition and their molecular weight profile. Both pectic and hemicellulosic polysaccharides were solubilized. It is concluded that extrusion has promise as a physical process for manipulating both the technological functionality and the health promoting properties of dietary fibers.     

Isolation and Characterization of Zein from Corn Distillers' Grains and Related Fractions

Corn distillers' grains with solubles (CDGS), the major coproduct of fermentation of corn to produce ethanol, were extracted with 0.1M NaOH, 0.1% dithiothreitol (DTT), and 0.5% SDS yielding 35% of the total nitrogen and ≈25% of the protein nitrogen. Gel electrophoresis revealed that the extractable proteins contained zein plus other proteins similar to the extractable proteins from corn flour. Although difficult to extract, the proteins isolated from the fermentation coproducts appeared undegraded and apparently survived gelatinization, fermentation, distillation, and drying during the production of ethanol. Extraction of CDGS with 60% ethanol at 60°C yielded 1.5–3.9% of crude zein. When the ethanol contained DTT, yields of crude zein were increased to 3.2–6.6%. Protein contents of the crude zeins were only 37–57%, indicating that lipids and pigments were coextracted with the ethanol. Gel electrophoresis showed that the protein fractions extracted by ethanol contained primarily α-zein whereas the proteins extracted by ethanol + DTT contained α- + β-zein. Further confirmation of the presence of zein in the crude prolamin preparations was obtained by amino acid analyses. The amino acid compositions of the crude zeins paralleled those of commercial zein and α-zein.     

Extrusion Chemistry of Wheat Flour Proteins: II. Sulfhydryl-Disulfide Content and Protein Structural Changes

Effects of twin-screw extrusion conditions on wheat flour proteins were studied, using a two-level fractional factorial experimental design (11 and 14% protein content, 160 and 185°C, 16 and 20% moisture, 300 and 500 rpm screw speed, mass flow rate of 225 and 400 g/min). Total protein detectable by solid-phase bicinchoninic acid assay decreased slightly after extrusion, with greatest protein loss at 16% moisture and 160°C. Sulfhydryl content of both flours increased after extrusion at 185°C and 16% moisture with moderate specific mechanical energy (SME ≈ 400–600 kJ/kg) or 160°C and 16% moisture with high SME (SME > 1,000 kJ/kg). Disulfide bonds increased under comparable conditions but with moderate shear (SME = 510–540 kJ/kg). At 20% moisture and either temperature, sulfhydryl and total thiol contents decreased without corresponding increases in disulfides. Reversed-phase HPLC indicated gliadins were the fractions most affected by extrusion; high molecular weight glutenin subunits also were affected. Changes in gliadins were extensive at 185°C and 16% moisture and were minimal at 160°C and 20% moisture. SDS-PAGE confirmed the disappearance of protein bands and appearance of new material at low and high molecular weights, presumably resulting from polypeptide fragmentation followed by random radical recombination. Both protein fragmentation and cross-linking appeared to involve free radicals.     

Changes in Disulfide and Sulfhydryl Contents and Electrophoretic Patterns of Extruded Wheat Flour Proteins

Twin‐screw extrusion of wheat flour and the effects on the flour proteins were studied using flour samples containing 9, 20, and 30% protein. Vital gluten containing 70% protein was used to achieve the flour protein levels. The three flour samples were extruded with a twin‐screw extruder at a combination of processing parameters (exit die temperatures of 120, 140, and 160°C, and screw speeds of 240, 320, and 400 rpm). Increasing extruder exit die temperatures resulted in increased sulfhydryl content of the 9 and 20% protein content flour samples, but appeared to have little or no effect on the 30% protein content flour sample. Similarly, disulfide content decreased, albeit disproportionately, following the same trend. Both sulfhydryl and disulfide contents of extruded samples were lower than those of the nonextruded samples and could imply denaturation of protein, aggregation through intermolecular disulfide bonds, or oxidation during extrusion processing. Total cysteine content of extruded samples decreased by ≈16% relative to nonextruded samples, but otherwise remained almost unchanged among all extruded samples. The loss of total cysteine in extruded samples could represent the generation of hydrogen sulfide, volatile organic compounds, or flavor compounds during extrusion. SDS‐PAGE analysis of total proteins showed a shift from the higher to lower molecular weight regions for certain protein bands. Both depolymerization and protein aggregation occurred at higher shear forces during extrusion.     

Effect of γ-zein on the rheological behavior of concentrated zein solutions

Zein, the prolamin of corn, is attractive to the food and pharmaceutical industries because of its ability to form edible films. It has also been investigated for its application in encapsulation, as a drug delivery base, and in tissue scaffolding. Zein is actually a mixture of proteins, which can be separated by SDS-PAGE into α-, β-, γ-, and δ-zein. The two major fractions are α-zein, which accounts for 70-85% of the total zein, and γ-zein (10-20%). γ-Zein has a high cysteine content relative to α-zein and is believed to affect zein rheological properties. The aim of this study was to investigate the effect of γ-zein on the often observed phenomena of zein gelation. Gelation affects the structural stability of zein solutions, which affects process design for zein extraction operations and development of applications. The rheological parameters, storage modulus (G') and loss modulus (G″), were measured for zein solutions (27% w/w solids in 70% ethanol). β-Mercaptoethanol (BME) was added to the solvent to investigate the effect of sulfhydryl groups on zein rheology. Modulus data showed that zein samples containing γ-zein had measurable gelation times under experimental conditions, contrary to samples with no γ-zein, where gelation was not detected. Addition of BME decreased the gelation time of samples containing γ-zein. This was attributed to protein unfolding. SEM images of zein microstructure revealed the formation of microspheres for samples with relatively high content of α-zein, whereas γ-zein promoted the formation of networks. Results of this work may be useful to improve understanding of the rheological behavior of zein.     

SME‐Arrhenius Model for WSI of Rice Flour in a Twin‐Screw Extruder

We have modeled a rice extrusion process focusing specifically on the starch gelatinization and water solubility index (WSI) as a function of extrusion system and process parameters. Using a twin‐screw extruder, we examined in detail the effect of screw speed (350–580 rpm), barrel temperature, different screw configurations, and moisture content of rice flour on both extrusion system parameters (product temperature, specific mechanical energy [SME], and residence time distribution [RTD]) and extrudate characteristics (expansion, density, WSI, and water absorption index [WAI]). Changes in WSI were monitored to reveal a relationship between the reaction kinetics during extrusion and WSI. Reaction kinetics models were developed to predict WSI during extrusion. WSI followed a pseudo first‐order reaction kinetics model. It became apparent that the rate constant is a function of both temperature and SME. We have developed an adaptation of the kinetic model based on the Arrhenius equation that shows better correlations with SME and distinguishes data from different screw configurations. This adaptation of the model improved predictability of WSI, thereby linking the extrusion conditions with the extruded product properties.     

Phenolic Compounds and Antioxidant Activity of Extruded Nixtamalized Corn Flour and Tortillas Enriched with Sorghum Bran

Sorghum bran (SB) is a good source of phenolic compounds with high antioxidant capacity that increases the antioxidant activity (AOX) of tortillas prepared with extruded nixtamalized corn flour. The objective of this research was to study the effects of bran addition (0, 5, or 10%) before (ENBESB) or after (ENAFSB) extrusion, in the features and composition of baked tortillas in terms of total phenolic compounds (TPC), AOX, color (L , a , b, hue, chroma, and E value), and tortilla firmness. It was possible to retain more than 81.8 and 89.9% of TPC and AOX, respectively, in ENBESB‐10% flour. Tortillas prepared with ENAFSB‐10% flour retained more than 92 and 76% of TPC and AOX, respectively, compared with ENBESB. However, tortillas elaborated with ENAFSB flour showed a higher firmness and lower flexibility than counterparts produced from ENBESB. The use of extrusion to produce nixtamalized corn flours and the strategy of adding the SB to the corn meal before extrusion were essential to retain TPC and AOX and, additionally, to enhance texture of tortillas.     

Extrusion conditions affect chemical composition and in vitro digestion of select food ingredients

An experiment was conducted to determine the effects of extrusion conditions on chemical composition and in vitro hydrolytic and fermentative digestion of barley grits, cornmeal, oat bran, soybean flour, soybean hulls, and wheat bran. Extrusion conditions altered crude protein, fiber, and starch concentrations of ingredients. Organic matter disappearance (OMD) increased for extruded versus unprocessed samples of barley grits, cornmeal, and soybean flour that had been hydrolytically digested. After 8 h of fermentative digestion, OMD decreased as extrusion conditions intensified for barley grits and cornmeal but increased for oat bran, soybean hulls, and wheat bran. Total short-chain fatty acid production decreased as extrusion conditions intensified for barley grits, soybean hulls, and soybean flour. These data suggest that the effects of extrusion conditions on ingredient composition and digestion are influenced by the unique chemical characteristics of individual substrates.     

Extrusion of Cross‐Linked Hydroxypropylated Corn Starches II. Morphological and Molecular Characterization

A series of cross‐linked hydroxypropylated corn starches were extruded with a Leistritz micro‐18 co‐rotating extruder. Extrusion process variables including moisture (30, 35, and 40%), barrel temperature (60, 80, and 100°C), and screw design (low, medium, and high shear) were investigated. Scanning electron microscopy (SEM) of extruded starches showed a gel phase with distorted granules and granule fragments after extrusion at 60°C. After extrusion at 100°C only a gel phase was observed with no granular structures remaining. High performance size exclusion chromatography (HPSEC) equipped with multiangle laser light‐scattering (MALLS) and refractive index (RI) detectors showed extruded starches degraded to different extents, depending on extrusion conditions. The average molecular weight of the amylopectin of unextruded native corn starch was 7.7 × 10⁸. Extrusion at 30% moisture, 100°C, and high shear reduced the molecular weight of amylopectin to 1.0 × 10⁸. Hydroxypropylated normal corn starch extruded at identical conditions showed greater decreases in amylopectin molecular weight. With the addition of cross‐linking, the amylopectin fractions of the extruded starches were less degraded than those of their native and hydroxypropylated corn starch counterparts. Similarly, increasing moisture content during extrusion lowered amylopectin degradation in the extruded starches. Increasing temperature during extrusion of cross‐linked hydroxypropylated starches at high moisture content (e.g., 40%) lowered amylopectin molecular weights of the extruded starches, whereas increasing extrusion temperature at low moisture content (30%) resulted in less degraded molecules. This difference was attributed to the higher glass transition temperatures of the cross‐linked starches.     

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.     

Effect of extrusion temperature on the solubility and molecular weight of lentil bean flour proteins containing low cysteine residues

Lentil flour was extruded at die temperatures of 135, 160, and 175 degrees C. The soluble protein content in the extrudates decreased by 40.1% in the extracting buffer (1% sodium dodecyl sulfate in 50 mM sodium phosphate buffer, pH 6.9) as the extrusion die temperature was increased to 175 degrees C. The most insoluble proteins in the extrudates extruded at die temperatures of up to 175 degrees C could be resolubilized by using sonication. The total disulfide content and sulfhydryl content in the extrudates decreased. The SDS-PAGEs showed that the molecular weight distribution of proteins in the lentil flour changed little before and after extrusion as well as during reduction. The results from this study show that the extrusion temperature had less effect on the solubility and molecular weight of the lentil proteins, which contain a lower level of cysteine residues than wheat proteins.     

Characterization and Optimization of Extrusion Cooking for the Manufacture of Third‐Generation Snacks with Winter Squash (Cucurbita moschata D.) Flour

The aim of this work was to study the effects of barrel temperature (BT, 93.5–140.5°C), feed moisture (FM, 21.3–34.7%), and winter squash flour content (SFC, 0.43–15.6%) on physicochemical properties of microwave‐expanded third‐generation snack foods obtained by extrusion. Physicochemical properties used for optimization were expansion index (EI), penetration force (PF), specific mechanical energy (SME), and total color difference (ΔE). Response surface methodology was used for the analysis of data. The highest values of EI and lowest values of PF were found at high BT and low FM. The lowest values of SME were obtained at high levels of FM throughout the range of BT and SFC, whereas the highest values of ΔE were obtained at high SFC and low FM. Increasing levels of SFC increased ΔE values, whereas EI and SME values decreased. The best processing conditions (EI > 6.0, PF < 9.5 N, SME < 172 kJ/kg, and ΔE < 18) were found in the range of BT, 122–141°C; FM, 24.7–29.5%; and SFC, 0–10.9%. Under optimal process conditions, the retention of total carotenoids was higher than 60%. It is possible to manufacture third‐generation snack foods with good physicochemical properties, which could bring a health benefit because of the presence of carotenoids and dietary fiber in winter squash flour.     

Waxy Soft White Wheat: Extrusion Characteristics and Thermal and Rheological Properties

Waxy wheat flour was analyzed for its thermal and rheological properties and was extruded to evaluate its potential for extruded products. Normal soft white wheat flour was analyzed with the same methods and same extrusion conditions to directly compare differences between the two types of flour. Through DSC analysis, waxy wheat flour was found to have a higher gelatinization peak temperature of 66.4°C than normal wheat at 64.0°C, although the transition required 2.00 J/g less energy. Rapid visco‐analysis indicated that the waxy wheat flour pasted much more quickly and at lower temperatures than the normal wheat flour. Preliminary extrusion experiments were conducted to determine the optimal screw profile for waxy wheat with respect to maximum radial expansion. The optimum screw profile was used for extrusion trials with varying flour moisture (15–25% wb) and extruder screw speed (200–400 rpm) while monitoring process conditions including back pressure and specific mechanical energy. Physical properties of the extrudates were then studied. The radial expansion ratios of the waxy wheat extrudates exceeded those of the normal wheat extrudates by nearly twice as much, and it was observed that the waxy wheat flour took less energy in the form of fewer shear screw elements to expand. The waxy wheat extrudates also exhibited significantly higher water solubility and less water absorption than the normal wheat extrudates owing to solubilizing of the extrudates. The results of our study indicate that waxy wheat flour may be a viable ingredient for creating direct expanded products with less energy.     

Characteristics and Protein Subunit Composition of Flour Mill Streams from Different Commercial Wheat Classes and Their Relationship to White Salted Noodle Quality

Proximate characteristics and protein compositions of selected commercial flour streams of three Australian and two U.S. wheats were investigated to evaluate their effects on the quality of white salted noodles. Wheat proteins of flour mill streams were fractionated into salt‐soluble proteins, sodium dodecyl sulfate (SDS)‐soluble proteins, and SDS‐insoluble proteins with a sequential extraction procedure. SDS‐soluble proteins treated by sonication were subsequently separated by nonreducing SDS polyacrylamide gel electrophoresis (SDS‐PAGE). There was a substantial amount of variation in distributions of protein content and protein composition between break and reduction mill streams. SDS‐insoluble proteins related strongly to differences in protein quantity and quality of flour mill streams. The soluble protein extracted by SDS buffer included smaller glutenin aggregates (SDS‐soluble glutenin) and monomeric proteins, mainly gliadin (α‐, β‐, γ‐, and ω‐types) and albumin and globulin. SDS‐soluble proteins of different flour mill streams had similar protein subunit composition but different proportions of the protein subunit groups. Noodle brightness (L) decreased and redness (a) increased with increased SDS‐insoluble protein and decreased monomeric gliadin. Noodle cooking loss and cooking weight gain decreased with increased glutenin aggregate (SDS‐soluble glutenin and SDS‐insoluble glutenin) and decreased monomeric gliadin. Noodle hardness, springiness, cohesiveness, gumminess, chewiness, tensile strength, breaking length, and area under the tensile strength versus breaking length curve increased with increased glutenin aggregate. Monomeric gliadin contributed differently to texture qualities of cooked noodles from glutenin aggregate. Monomeric albumin and globulin were not related to noodle color attributes (except redness), noodle cooking quality, and texture qualities of cooked noodles. The results suggested that variation in protein composition of flour mill streams was strongly associated with noodle qualities.     

Factors Affecting Yield and Composition of Zein Extracted from Commercial Corn Gluten Meal

Twelve corn gluten meal samples obtained from six wet-milling plants were processed into zein. Zein was extracted using 88% aqueous isopropyl alcohol at pH 12.5, followed by chilling. Protein recovery ranged from 21.3 to 32.0%, and protein purity ranged from 82.1 to 87.6%. Protein recovery increased as the protein purity increased (r = 0.76) (P < 0.01). One of the major factors influencing extraction yield was protein composition; especially α-zein content, which ranged from 53.4 to 64% of the total protein in the corn gluten meal samples. The intensity of red color of the corn gluten meal was negatively correlated with protein recovery and zein purity (r = -0.66 and -0.72, respectively) (P < 0.02).     

Effects of Calcium Hydroxide and Processing Conditions on Corn Meal Extrudates

The effects of added calcium hydroxide (0.0, 0.15, 0.25, and 0.35%) and processing conditions, feed moisture content (mc) (16, 18, and 20%) and barrel temperature (130 and 150°C) on characteristics of corn meal extrudates were studied. Extruder screw speed was maintained at 130 rpm. Corn meal was extruded with a single-screw extruder (Brabender model GNF 1014/2) with a screw compression ratio of 3:1. The highest values (P < 0.05) for radial expansion and the lowest values for density and breaking force of extrudates were found for the treatment with 0.00% calcium hydroxide extruded at 16% feed mc and 130°C barrel temperature. This treatment was statistically different from the other treatments. Best values for radial expansion of samples extruded with added calcium hydroxide were for the samples with 0.15% calcium hydroxide at 18% feed mc and 130°C barrel temperature, followed by the sample with 0.35% calcium hydroxide at 16% feed mc and 130°C barrel temperature. Water absorption index and water solubility index were affected by calcium hydroxide and extrusion conditions evaluated. Extrudates had large numbers of flattened and sheared granules. Increases in calcium hydroxide increased extrudate yellowness. The combined action of calcium hydroxide and extrusion conditions completely modified the organized structure of the starch and suggest the formation of a starch-calcium complex (crystalline region). The texture of the extruded products was crispy after puffing.     

Using an In‐Line Slit‐Die Viscometer to Study the Effects of Extrusion Parameters on Corn Melt Rheology

An in‐line slit‐die viscometer (SDV) was used to measure the viscosity of a melt extrudate independently of the extruder operating conditions. The melt produced by extrusion of the corn grits followed a power law rheological model. The viscosity of the melt and extrusion parameters such as specific mechanical energy (SME), torque, and die pressure decreased with increasing moisture content. The degree of starch gelatinization increased when barrel temperature increased from 90 to 130°C. At temperatures higher than 130°C, most of the starch had gelatinized. The increase in barrel temperature, however, resulted in small changes in the apparent viscosity of the melt, until a maximum of ≈130°C. At a constant feed rate, SME increased and torque decreased when screw speed increased due to the shear thinning behavior of the melt. At a constant screw speed, the torque increased and SME decreased with increasing feed rate. This was due to a decrease in apparent viscosity of the melt at higher feed rates. SME is not an independent extrusion variable and should be used with caution either when predicting the effect of thermomechanical treatment of the product or as the key and only variable for controlling the food extrusion process.     

Effect of Specific Mechanical Energy on Properties of Extruded Protein‐Starch Mixtures

The effect of the specific mechanical energy (SME) during extrusion of a protein‐starch mixture was studied by analyzing the glass transition temperature (T_g) and starch gelatinization. We found that the SME values of 344 to 2108 kJ/kg did not significantly change the T_g of the product. To explain the insensitivity of T_g to SME in spite of reported fragmentation taking place during extrusion, we studied the effect of the molecular weight (MW) on T_g in a model system consisting of dextrans of varying molecular weights. We found that the effect of the molecular weight on the T_g reached a plateau at 6.7 × 10⁴. Because the reported size of the fragments created during the extrusion process is larger than this, we were able to explain the apparent insensitivity of T_g to SME in the protein‐carbohydrate matrix studied. However, we found that starch gelatinization varied with SME, the degree of gelatinization being higher for systems exposed to higher SME.