Rheological Studies on the Reaction of Zein with Polyethylenemaleic Anhydride

There continues to be interest in developing solvent‐resistant articles from biobased renewable materials to successfully compete with petrochemical products. It was previously shown that reaction of zein with polyethylenemaleic anhydride (PEMA) provides articles that are solvent resistant. The gelation kinetics for the reaction of PEMA with zein was investigated rheologically to better understand this chemistry. The reaction of the nucleophilic groups on zein with the anhydrides on PEMA is the main cause for the gelation reaction. The gelation time was defined as being the point when the elastic modulus (G′) and viscous modulus (G″) cross. In this work, the rate of reaction, in terms of time to gelation, was studied in N,N‐dimethylformamide solution for which the amount of PEMA, the reaction temperature, and the overall reaction concentration were varied. Exponential relationships were found between the gelation time and % PEMA, temperature, and % solids, as well as between elastic modulus with either % PEMA or % solids. The concentration of PEMA had the largest impact on gelation time, for which going from 2.5% PEMA to 6% PEMA reduced the gelation time from 63,114 to 1,576 s. The temperature dependence of this gelation reaction was well described by an Arrhenius plot with an apparent activation energy of 50.5 kJ/mol.     

Extraction and Film Properties of Kafirin from Coarse Sorghum and Sorghum DDGS by Percolation

The high cost of kafirin and zein restricts their use for bioplastic and food applications. Effective, simple, and rapid kafirin/zein isolation processes are required. Here a percolation‐type aqueous ethanol solvent extraction process from coarse meals (grits) and coarse sorghum distillers dried grains and solubles (DDGS) for kafirin and zein isolation employing a low ratio of extractant to meal (2.5:1) was investigated, which is potentially applicable in the grain bioethanol industry. Postextraction filtration times were more than twice as fast using coarse meals compared with fine flours. Washing the meals prior to extraction to remove starch improved protein preparation purity to 73–85% compared with 68–72% for unwashed meals. Hence, no subsequent filtration or centrifugation step is required to clean up the kafirin/zein solution prior to solvent evaporation. With a single extraction step, kafirin/zein yields were 48% (protein basis) for DDGS and 53–70% for washed sorghum/maize meals. Cast films were used as a model bioplastic system to evaluate extracted kafirin/zein functional properties. DDGS kafirin films had rough surfaces but had the lowest water uptake and in vitro digestibility, owing to heat‐induced disulfide crosslinking during DDGS processing. Extraction by percolation using coarse meal/DDGS has potential to improve kafirin/zein viability.     

Production of Textile Fibers from Zein and a Soy Protein-Zein Blend

Zein fibers were successfully prepared by a wet-spinning technique from a zein suspension formulated with 15% zein, 60% water, 22% 0.4N sodium hydroxide, and 3% urea by weight. Tenacities were measured with an Instron machine, and flexibility was determined by noting the smallest diameter rod around which a fiber could be looped without breaking. After spinning, the tenacity of the zein fibers was improved by modification with several agents: acetic anhydride for acetylation, glutaraldehyde, and dialcohols for cross-linking and physical stretching. The tenacity and flexibility of the fibers were measured at 11, 65, and 100% rh. Untreated fibers had tenacities of 3.41, 2.65, and 0.17 g/tex at 11, 65, and 100% rh, respectively. A combination of chemical treatments (20% glutaraldehyde and 95% acetic anhydride) and 115% stretching increased tenacities to 6.89. 6.56, and 1.17 g/tex at 11, 65, and 100% rh, respectively. Control zein fibers had flexibilities of 5 and 2.5 mm at 11 and 65% rh, respectively, whereas the treated fibers had flexibilities of 1.5 mm at both humidities. Extrusion of zein fibers was not successful. Zein-soy protein mixture could be extruded, but properties of wet-spun fibers of zein-soy blends were not much improved over those of soy protein alone. The tenacity of blended soy protein-zein fibers was greater than that of soy protein fibers at 11% rh.     

Reducing the brittleness of zein films through chemical modification

Zein protein is a major coproduct of biofuel from corn. To reduce the brittleness of zein films, a new type of zein-based biomaterial, was synthesized by chemical modification of zein with lauryl chloride through an acylation reaction. The final products were confirmed by (1)H NMR, FT-IR analysis, and SDS-PAGE. Thermal analysis detected no microphase separation in the synthesized polymer matrix. As the content of lauryl moiety increased, the glass transition temperatures of modified zein decreased by as large as 25.8 °C due to the plasticization effect of the lauryl moiety. In addition, mechanical and surface properties of cast films from acylated zein were also investigated. The elongation at break of modified zein sheet was increased by about 7-fold at the high modification level with some loss of mechanical strength. The surfaces of modified zein films were as uniform as unmodified zein film but more hydrophobic, further suggesting that no microphase separation happened during the film formation process. This work indicated the potential of these new biomaterials in the development of biodegradable food packaging materials and delivery systems.     

Development of New Method for Extraction of α‐Zein from Corn Gluten Meal Using Different Solvents

A modified procedure for the extraction of α‐zein from corn gluten meal was developed and compared against a commercial extraction method. The modification involved raising the concentration of alcohol in solvent and removing the precipitate by centrifugation. Five organic solvent mixtures were compared using the modified extraction procedure developed along with the reductant sodium bisulfite and NaOH. The modified procedure precipitated most of the non‐α‐zein protein solids by increasing the concentration of alcohol. The supernatant had α‐zein‐rich fraction, resulting in higher yield of α‐zein than the commercial method when cold precipitated. The commercial extraction procedure had a zein yield of 23% and protein purity of 28% using 88% 2‐propanol solvent. The three best solvents, 70% 2‐propanol, 55% 2‐propanol, and 70% ethanol, yielded ≈35% of zein at protein purity of 44% using the modified extraction procedure. Zeins extracted using the novel method were lighter in color than the commercial method. Densitometry scans of SDS‐PAGE of α‐zein‐rich solids showed relatively large quantities of α‐zein with apparent molecular weights of 19,000 and 22,000 Da. The α‐zein‐rich solids also had small amounts of δ‐zein (10,000 Da) because it shares similar solubility properties to α‐zein. A solvent mixture with 70% 2‐propanol, 22.5% glycerol, and 7.5% water extracted significantly less zein (≈33%) compared to all other solvents and had α‐zein bands that differed in appearance and contained little to no δ‐zein.     

Optimizing Extraction of Zein and Glutelin‐Rich Fraction During Sequential Extraction Processing of Corn

This study was conducted to improve yields and qualities of corn protein co‐products produced by the sequential extraction process (SEP), a process using ethanol to fractionate corn in producing fuel ethanol. A two‐stage extraction protocol was evaluated to recover zein and subsequently recover a glutelin‐rich fraction (GRF). After the simultaneous oil‐extraction and ethanol‐drying step of SEP, zein was extracted from the anhydrous‐ethanol‐defatted, flaked corn by using 70% (v/v) ethanol at 60°C for 1.5 hr in a shaking water bath. Zein was recovered by ultrafiltering and then drying in a vacuum‐oven. Zein yield was 65% of the available zein in the flaked corn. SDS‐PAGE band patterns of the recovered zein closely resembled that of commercial zein. After zein extraction, the GRF was extracted using 45% ethanol and 55% 0.1M NaOH at 55°C for 2 hr. The extract was concentrated by ultrafiltration and then freeze‐dried. GRF yield was ≈65% of the available protein. Freeze‐dried GRF contained 90% crude protein (db), which classified the protein as a protein isolate. As with the protein concentrate from the original SEP, the GRF isolate was highly soluble in water at pH ≥ 7, had good emulsifying and foaming properties, formed stable emulsions, and was heat‐stable.     

New (1)h NMR procedure for the characterization of native and modified food-grade starches

A novel, fast, and straightforward procedure is presented for the characterization of starch (the largest energy component in food) and modified starches (such as octenyl succinic anhydride (OSA)-modified starches used as a dispersing agent in the food industry). The method uses (1)H NMR to measure the degree of branching and also, for modified starches, the degree of chemical substitution. The substrate is dissolved in dimethyl-d(6) sulfoxide; addition of a very low amount of deuterated trifluoroacetic acid (d(1)-TFA) to the medium gives rise to a shift to high frequency of the exchangeable protons of the starch hydroxyl groups, leading to a clear and well-defined (1)H NMR spectrum, which provides an improved way to determine the degrees of both branching and chemical substitution. Measurements of the size and molecular weight distributions by multiple-detector size exclusion chromatography show that degradation by TFA does not affect the accuracy of the method.     

Solvent Extraction of Zein from Dry‐Milled Corn

Batch extraction of zein from dry‐milled whole corn with ethanol was optimum with 70% ethanol in water, an extraction time of 30–40 min, and temperature of 50°C. High yields (60% of the zein in corn) and high zein contents in the extracted solids (50%) were obtained at a solvent‐to‐solids ratio of 8 mL of 70% ethanol/g of corn. However, zein concentration in the extract was higher at lower ratios. Multiple extraction of the same corn with fresh ethanol resulted in a yield of 85% after four extractions, whereas multiple extractions of fresh corn with the same ethanol resulted in high (15 g/L) zein concentration in the extract. Optimum conditions for batch extraction of zein were 45°C, with 68% ethanol at a solvent‐to‐solids ratio of 7.8 mL/g for an extraction time of 55 min. Column extractions were also best at 50°C and 70% ethanol; a solvent ratio of 1 mL/g resulted in high zein concentrations in the extract (17 g/L) but yields were low (20%).     

Glycemic response to a food starch esterified by 1-octenyl succinic anhydride in humans

To evaluate the glycemic response to a food starch esterified by 1-octenyl succinic anhydride (OSA), 30 healthy nondiabetic adult subjects were studied in a double-blind crossover design. After an overnight fast, subjects consumed a product containing either 25 g of glucose or 25 g of OSA-substituted starch. Finger-prick capillary blood was obtained at baseline and 15, 30, 45, 60, 90, and 120 min postprandial for glucose measurement. After OSA treatment, the rise in blood glucose was reduced (P < 0.05) at 15 and 30 min and tended (P < 0.08) to be lower at 45 min. Mean peak rise in glucose was reduced 19% (P < 0.01) by OSA (3.30 +/- 0.19 versus 2.66 +/- 0.16 mmol/L) compared to glucose, but time to peak did not differ between treatments. Net incremental area under the curve was also lower (P < 0.05) on OSA compared to glucose. Minimal effects on gastrointestinal symptoms (intensity and frequency of nausea, cramping, distention, and flatulence) were noted for both products, with no clinically significant difference between products. In conclusion, starch substitution with OSA attenuated the postprandial glycemic excursion compared to an equivalent glucose challenge and was well tolerated by fasting healthy adult subjects.     

Starch Acetate‐Maleate Mixed Ester Synthesis and Characterization

A new starch acetate‐maleate mixed ester with different degrees of substitution (DS) for each ester group was prepared. High‐amylose (70%) corn starch was first reacted at 123°C with ≈4.5 equivalents of acetic anhydride for different times, and then the intermediate products were further reacted at 80°C with 0.27–1.1 equivalents of maleic anhydride for a fixed time. ¹H NMR and alkaline saponification were used to determine DS values of acetyl and maleate groups. Increasing the reaction time from 60 to 120 min for starch and acetic anhydride increased DS of the acetyl group, while DS of the maleate group increased with increasing amounts of maleic anhydride. There was a good agreement between the two methods for DS determination. The reaction efficiency of acetylation increased from ≈30 to 50% with reaction time, and maleation decreased from ≈30 to 20% with increasing amounts of maleic anhydride. The mixed esters were characterized by ¹H NMR, ¹³C NMR, FTIR, DSC, and TGA. Characteristic peaks in ¹H NMR, ¹³C NMR, and FTIR spectra indicated the presence of acetyl and maleate groups in starch molecules. The exothermic peak in the DSC curve and the increase in thermal stability from the TGA curve were attributed to thermal cross‐linking of the double bond of the maleate group.     

Effects of a Dual‐Frequency Frequency‐Sweeping Ultrasound Treatment on the Properties and Structure of the Zein Protein

We investigated the effects of a dual‐frequency frequency‐sweeping ultrasound (DFFSU) treatment on the functional properties and structure of zein. The solubility of ultrasound‐treated zein proteins increased slightly but significantly as the treatment time increased. The results showed that the DFFSU treatment had an obvious influence on the mean particle size and the size distribution. A significant (P < 0.05) increase in the size of the particles with respect to time was observed after a sonication time of more than 20 min in zein solutions. Differential scanning calorimetry results showed that sonication alters the thermal behavior of zein. Circular dichroism spectra showed a small increase in the percentage of ordered structure elements within the protein molecule. After 60 min of ultrasonication, the percentage of α‐helix structures increased by 0.9%, whereas the percentage of β‐sheets and β‐turns decreased by 0.5%. Microstructural analyses by scanning electron microscopy showed that several microholes appeared in the zein following ultrasonic pretreatment. Under the conditions investigated in this study, DFFSU treatment was found to affect the studied functional properties of the zein protein. This technology could be used to obtain improved functional properties in some protein samples.     

Preparation and Characterization of Starch Particles for Use in Pickering Emulsions

Particle‐stabilized emulsions, called Pickering emulsions, can be produced by using starch particles. In this work we studied how the properties of the starch particles affect the droplet size and creaming of such emulsions. In the study, various sizes of starch particles were generated by two different methods and used to stabilize Pickering emulsions. Sedimentation according to Stokes’ law was used to separate small and large starch granules. Acid hydrolysis was another method used to obtain smaller particles. All samples were modified with octenyl succinic anhydride (OSA) to increase their hydrophobicity with a level of OSA substitution between 1.8 and 3.1%. The size of starch particles was the main factor influencing emulsion droplet sizes. Furthermore, the droplet size decreased as the starch concentration increased. Using small starch particles with sizes <10 μm produced stable emulsions with smaller droplet size compared with larger sizes of starch particles, >10 μm. When subjected to acid hydrolysis, smaller starch particles were generally obtained, which could subsequently create smaller emulsion droplets. The emulsion index increased for the acid‐hydrolyzed starch owing to the size reduction of starch particles. The shape of the starch seemed to have a minor impact on the droplet size and the creaming of Pickering emulsions.     

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.     

Composition and Reactivity of A‐ and B‐type Starch Granules of Normal,Partial Waxy,and Waxy Wheat

Wheat has great potential to make inroads into starch markets with the advent of partial waxy and waxy starches of diverse composition and properties. The majority of isolated starch utilized in food applications is chemically modified to improve starch properties according to the intended use. Therefore, it is critical to understand factors that affect wheat starch reactivity. This work investigated the relative reactivities of normal, partial waxy, and waxy wheat starches and their respective A‐ and B‐type starch granule fractions. Native starch isolated from four closely related soft wheat lines (normal, partial waxy, and full waxy) was modified through 1) substitution (propylene oxide analog) and 2) cross‐linking (phosphorus oxychloride) reactions to generate both types of modified starch products for each wheat line. Characterization of the unmodified starch fractions confirmed compositional differences among the cultivars and their respective granule types. In cross‐linking reactions, B‐type granules were slightly more reacted than A‐type granules for all cultivars, while the waxy starch generally exhibited higher reactivity compared with normal and partial waxy starches. For the substituted starches, no differences in reactivity were observed among the cultivars or between the two granule types.     

Octenyl succinic anhydride modified early indica rice starches differing in amylose content

Seven early indica rice starches with different amylose contents were modified by octenyl succinic anhydride (OSA) in aqueous suspension systems to evaluate the effect of amylose contents on starch esterification. The crystalline structure and pasting properties of starches were investigated using X-ray diffraction and a Rapid Visco Analyzer (RVA). The results indicated that the amylose content had a positive impact on the OSA modification. As the amylose content increased from 0 to 39.6%, the degree of substitution increased from 0.024 to 0.030 and the reaction efficiency increased from 62.8 to 77.5%. X-ray diffraction scans confirmed that the amylose was mainly present in the amorphous domain of the granule and was highly substituted after the OSA treatment. The RVA profiles demonstrated that the OSA starches had higher viscosities than their native counterparts. Moreover, negative correlations were observed between the amylose content and the major RVA parameters (e.g., peak viscosity, hot paste viscosity, cool paste viscosity, and breakdown viscosity).     

Effects of Extrusion and Starch Removal Pretreatment on Zein Proteins Extracted from Corn Gluten Meal

In this study, the structure and selected properties of zeins extracted from corn gluten meal (CGM) pretreated by extrusion and removal of starch were investigated. The structure and properties of the zeins from pretreated CGM changed significantly. Pretreatments can decrease the extraction yields of zeins and change the granule shape and size of zein aggregates. The studies indicated that extrusion and removal of starch can significantly decrease the thermal enthalpy (ΔH₁ and ΔH₂) of zein from 1.94 ± 0.20 to 0.19 ± 0.10 and from 107.20 ± 0.80 to 78.62 ± 2.30 and J/g, respectively. The SDS‐PAGE results confirmed that the molecular weight of zeins from CGM was 24,000 and 27,000, and the molecular weight of zeins did not change with the pretreatment. On the other hand, the circular dichroism spectroscopy results showed that the processing of extrusion and removal of starch can change the secondary structure content of β‐sheets and β‐turns; these results indicated that extrusion and removal of starch can significantly break the secondary structure of zeins. Furthermore, extrusion and removal of starch can change the sulfhydryl content of zeins. The obtained results provided some fundamental information that is useful for further modification of CGM to improve its functional properties and industrial applications.     

Physical properties of octenyl succinic anhydride modified rice,wheat, and potato starches

The physical properties of octenyl succinic anhydride (OSA) starches prepared from rice, wheat, and potato starches were studied. Rice and wheat OSA starches had significantly higher peak viscosity (PV), hot paste viscosity (HPV), and cool paste viscosity (CPV), but potato OSA starch had only significantly higher CPV, relative to the native starch. The gel hardness was higher with lower degree of substitution (DS) but lower with higher DS OSA compared to native starch. The swelling volumes (SV) of rice and wheat OSA starches were significantly higher compared to native starch, but the SV of potato OSA starch was slightly lower at high DS. The gelatinization temperature (GT) of rice OSA starches was sharply lower at low DS; for wheat OSA starch it was slightly lower even at high DS, but potato OSA starches had higher GT than the native starch. The enthalpy of all the OSA starches decreased gradually with increased DS. This study showed that the magnitude of changes in physical properties of OSA-modified starches depends not only on their DS but also on the botanical origin of the native starches.     

Isolation of Zein Using 100% Ethanol

Traditionally, zein is isolated and recovered from corn gluten meal (GCM) using aqueous alcohol as the solvent. Recovery of zein from this solvent is inconvenient and costly. Zein is insoluble in 100% ethanol at room temperature, but it is soluble at 120°C in ethanol. Absolute ethanol effectively extracted zein from CGM, distillers dried grains (DDG), and ground corn. Zein was extracted from CGM with absolute ethanol in a high‐pressure reactor at 130°C. After extracting at 130°C for 45 min, the solution was pumped out of the extractor and allowed to cool. Upon cooling, the zein precipitated from solution. The precipitate was removed from the solution and air‐dried, resulting in 14% recovery of the starting material. The recovered precipitate had an average protein content of >90% on a dry basis, accounting for ≈20% of the CGM protein and recovered ≈35% of its zein. No differences were seen in the amount of zein extracted from CGM samples that were hand‐collected off the dewatering screen and gently dried, versus commercial CGM samples. The commercial CGM did produce a greater amount of solubles. The extraction procedure also worked at temperatures as low as 90°C. The lower temperature did produce lower yields of extracted zein. The zein extracted at the lower temperatures was less brown, but zein extracted at either temperature was almost fully soluble in traditional zein solvents.     

Improved Isolation of Zein from Corn Gluten Meal Using Acetic Acid and Isolate Characterization as Solvent

An improved means of isolating zein is needed to develop new uses for corn zein. We have measured the yield of zein and evaluated the ability of acetic acid to remove zein from corn gluten meal, distillers dried grains, and ground corn using acetic acid as solvent. Acetic acid removed zein more quickly, at lower temperatures, and in higher yields when compared with alcoholic solvents. After 60 min at 25°C, ≈50% of the zein in corn gluten meal was removed. A step change in yield from 43 to 50% occurs as the extraction temperature is increased from 40 to 55°C after mixing for 30 min at 25% solids. The protein composition of the zein removed from corn gluten meal using acetic acid is very similar to that of commercial zein by SDS‐PAGE. The zein obtained from corn gluten meal using acetic acid had higher amounts of fatty acids and esters according to IR analysis, leading to slightly lower protein content. Films made from zein extracted from corn gluten meal using acetic acid had lower tensile strength (≈60% lower) than films produced from commercial zein. Fibers with very small diameter (0.4–1.6 μm) can be produced by electrospinning using the AcOH solution obtained after corn gluten meal extraction.     

Chemistry and physical properties of melt-processed and solution-cross-linked corn zein

Corn zein was cross-linked with glutaraldehyde (GDA) using glacial acetic acid (HAc) as catalyst. The objectives are to evaluate the swelling characteristics of GDA cross-linked zein gels in water, ethanol, and their combinations. Similar formulations, upon solvent evaporation, form films. The mechanical properties of the films are compared to compression molded tensile bars from GDA melt-processed zein as a second objective. Chemistry of the cross-linking reaction was based on the aldehyde binding characteristics defined by use of fluorescence spectroscopy; sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to demonstrate the cross-linking reaction; FTIR to observe absorption differences of the cross-linked product; differential scanning calorimetry, dynamic mechanical analysis and thermogravimetric analysis to assess thermal properties; and the use of Instron Universal Testing Machine to evaluate mechanical properties. A reaction mechanism for acid catalyzed GDA cross-linking of zein is proposed. Thermal and mechanical properties of tensile bars cut from either film or formed by compression molding were similar, where both showed increased tensile strengths, ductility and stiffness when compared with unmodified controls. Samples that were reacted with 8% GDA by weight based on weight of zein from either process retained their integrity when tensile bars from each were subjected to boiling water for 10 min or soaking in either water or HAc for 24 h. The melt-processed, cross-linked zein is a more environmentally friendly method that would eliminate the need for HAc recovery.