Wet‐Milling Characteristics of 10 Lines from Germplasm Enhancement of Maize Project Compared with Five Corn Belt Lines

The use of corn (Zea mays L.) hybrids with high grain yield and starch extractability has steadily increased in the processing industry. In light of widespread corn seed industry participation in the Germplasm Enhancement of Maize Project (GEM), which seeks to enhance exotic germplasm, future hybrids may contain more exotic sources in genetic backgrounds. It is necessary to establish and monitor physical, compositional, and milling characteristics of the new exotic breeding materials to determine the processing value. The present study was conducted to determine the wet‐milling characteristics of a set of GEM lines compared with typical Corn Belt lines. Ten GEM lines introgressed with exotic materials from Argentina, Chile, Cuba, Florida, and Uruguay and previously identified as having different starch yields, three commercial inbred lines, and two public inbred lines (B73 and Mo17) were analyzed using both near‐infrared transmittance (NIT) and a 100‐g wet‐milling procedure. There were statistical differences (P < 0.05) in the yield of wet‐milled fractions (starch, fiber, gluten, and germ). The GEM lines AR16035:S19‐227‐1‐B and CUBA117:S1520‐562‐1‐B had similar or better starch yield and starch recovery than B73 and the other adapted inbred lines, indicating that they may be useful in improving the proportion of extractable starch present in kernels of hybrids. Residual protein levels in the starch and gluten fractions were 0.26–0.32% and 38–45%, respectively. The starch yield of GEM lines from wet milling correlated positively with starch content from NIT and was negatively correlated with protein content of the corn kernels. Oil content in the germ varied from 50 to 60%. Our results indicate that incorporating GEM lines in a breeding program can maintain or even improve wet‐milling characteristics of Corn Belt materials if lines with appropriate traits are used.     

A 10‐g Laboratory Wet‐Milling Procedure for Maize and Comparison With Larger Scale Laboratory Procedures

A very small scale laboratory procedure (≈10 g) is needed to test wet‐milling characteristics of corn when amounts of corn available for testing are quite limited. The objective of this study was to downscale 100‐g laboratory wet‐milling methods already widely used to measure wet‐milling properties of 10 g of corn. A Standard 100‐g procedure, a Modified 100‐g procedure, and an Experimental 10‐g procedure were compared using three corn hybrids with known differences in wet‐milling properties. All three procedures ranked most fraction yields (all except for germ) of the three hybrids the same. Germ separation was conducted differently for each procedure and probably accounts for these differences. Flotation and screening methods were likely affected by germ density and germ size, and hand‐picking the germ was efficient in recovering a pure germ fraction. The two 100‐g procedures were performed very similarly except for fiber recovery. The Modified 100‐g procedure was more efficient in recovering fiber because of intensive washing. Hybrid effects on the starch/gluten separation were more pronounced when the Experimental 10‐g procedure was used, which may allow for more discrimination among hybrids. Although most fraction yields are too small to run replicates for analytical tests, the Experimental 10‐g procedure will be useful in measuring milling efficiency of early generations of corn hybrids where limited samples are available, such as when valuable recombinant proteins are expressed for therapeutics and industrial enzymes.     

Farinograph Responses for Wheat Flour Dough Fortified with Wheat Gluten Produced by Cold‐Ethanol or Water Displacement of Starch

The objective of this research was to identify and define mixing characteristics of gluten‐fortified flours attributable to differences in the method for producing the gluten. In these studies, a wheat gluten concentrate (W‐gluten) was produced using a conventional process model. This model applied physical water displacement of starch (dispersion and screening steps), freeze‐drying, and milling. W‐gluten was the reference or “vital” gluten in this report. An experimental W‐concentrate was produced using a new process model. The new model applied coldethanol (CE) displacement of starch (dispersion and screening steps), freeze‐drying, and milling. Freeze‐drying was used to eliminate thermal denaturation and thereby focus on functional changes due only to the separation method. The dry gluten concentrates were blended with a weak, low‐protein (9.2%), soft wheat flour and developed with water in a microfarinograph. We found that both water and cold‐ethanol processed gluten successfully increased the stability (St) and improved mixing tolerance index (MTI) to create in the blended flour the appearance of a breadbaking flour. Notably, in the tested range of 9–15% protein, the St for CE‐gluten was always higher then the St for W‐gluten. Furthermore, the marginal increase in St (slope of the linear St vs. protein concentration) for the CE‐gluten was ≈57% greater than that for the W‐gluten. The slope of the MTI vs. protein data was lower for the CE‐gluten by 24%. Flour fortified with CE‐gluten exhibited higher water absorption (up to 1.8% units at 13.5% P) than flour fortified with W‐gluten.     

Effect of Stress Cracks on Corn Wet‐Milling Yields

U.S. No. 2 yellow dent corn was randomly probe‐sampled from rail cars being shipped to a wet‐milling plant from a Corn Belt local elevator. The probe samples were blended together and kernels were sorted into four levels of stress cracks (0, 1, 2, or multiple). Each level of stress cracking was then laboratory wet‐milled in triplicate. The only statistically observed differences were in total fiber and in protein content of the gluten meal fraction. The starch yield difference between zero stress cracked corn and multiple stress cracked corn was smaller (0.8%) than would be expected if stress cracking were an indicator of damage to the wet‐milling characteristics of the corn.     

Laboratory Wet Milling of Corn: Milling Fraction Correlations and Variations Among Crop Years

Several coproducts result from fractionating corn in the wet‐milling process. Because small changes in product composition and milling characteristics can have a major impact on coproduct yields and values, testing is done to anticipate final product yields. Using small sample size and controlled conditions, a laboratory wet‐milling method proved to be a useful tool for wet milling and genetics industries. A wet‐milling process (100‐g batches) was used for data collection. Data collected during 11 years (1994–2004) were observed for samples used as benchmarks to verify process precision and accuracy and determine correlations among wet‐milling yields. More than 400 milling tests were performed on benchmark samples. Data from benchmark samples also were pooled. Coefficients of variation were low (<6%) for mean yields; year‐to‐year standard deviations of benchmark sample yield means were homogenous and implied precision of the procedure. Some differences were detected in mean yields among years (P ≤ 0.05) for benchmark data due to combined effects of hybrid and environment. A negative correlation (r = –0.58) was observed between starch and gluten yield for pooled benchmark data. Four years (2002–2005) of milling data from commercially available hybrids were analyzed using the milling procedure. For pooled commercial data, the correlation between starch and fiber yield was (r = –0.80); correlation between starch and gluten was (r = –0.76).     

Relationship Between Wheat (Triticum aestivum L.) Grain Hardness and Wet‐Milling Quality

Grain hardness variation has large effects on many different end‐use properties of wheat (Triticum aestivum). The Hardness (Ha) locus consisting of the Puroindoline a and b genes (Pina and Pinb) controls the majority of grain hardness variation. Starch production is a growing end‐use of wheat. The objective of this study was to estimate the differences in starch yield due to natural and transgenically conditioned grain hardness differences. To accomplish this goal, a small‐scale wet‐milling protocol was used to characterize the wet‐milling properties of two independent groups of isogenic materials varying in grain hardness and in Pin expression level. The first group of lines consisted of hard/soft near‐isogenic lines created in cultivars Falcon or Gamenya in which lines carried either the Pina‐D1a (functional) or the Pina‐D1b (null) alleles of Pina. The second group of lines consisted of Pina, Pinb, or Pina and Pinb overexpressing lines created in Hi‐Line, a hard red spring wheat. Soft near‐isogenic lines had higher starch extractability than the hard Pina null counterparts. This difference in starch extractability was more pronounced between Hi‐Line and its transgenic isolines, with highest levels of extractable starch observed in the transgenic isoline with intermediate grain texture. The results demonstrate that the Ha locus and puroindoline expression are both linked to wet‐milling starch yield and that selection for increased Ha function increases starch yield through the enhanced separation of starch granules and the protein matrix during wet milling.     

Physical,Compositional, and Wet‐Milling Characteristics of Mexican Blue Maize (Zea mays L.) Landrace

Mexico has the largest diversity of genetic resources for maize in the world, with about 59 different landraces. However, little is known about their wet‐milling characteristics. The aim of this study was to determine whether 15 Mexican blue maize (Zea mays L.) genotypes of Elotero de Sinaloa landrace collected in the northwestern region of Mexico have suitable wet‐milling properties. Great variability of physical, compositional, and wet‐milling characteristics among these blue maize genotypes was observed. The FAUAS‐457 and FAUAS‐488 maize genotypes had similar starch yield and starch recovery as reported for the wet‐milling industry, which indicated that they may be useful as a source of extractable starch. Residual protein levels in the starch fractions were in the range of 0.39–0.68%, and total solids recovery exhibited a mean value of 98.8%, indicating acceptable efficacy of the wet‐milling process. This process afforded starches from blue maize genotypes with low protein contents. Wet‐milling fractions correlated with the physical and chemical properties of the kernels. Our results indicate that Mexican blue maize genotypes contain characteristics that make them appropriate and utilizable at the industrial level, and they can also be valuable for improving wet‐milling characteristics of maize through breeding programs.     

White Pan Bread and Sugar‐Snap Cookies Containing Wheat Starch Phosphate,A Cross‐Linked Resistant Starch

Pup‐loaf bread was made with 10, 30, and 50% substitution of flour with wheat starch phosphate, a cross‐linked resistant starch (XL‐RS4), while maintaining flour protein level at 11.0% (14% mb) by adding vital wheat gluten. Bread with 30% replacement of flour with laboratory‐prepared XL‐RS4 gave a specific volume of 5.9 cm³/g compared with 6.3 g/cm³ for negative control bread (no added wheat starch), and its crumb was 53% more firm than the control bread after 1 day at 25°C, but 13% more firm after 7 days. Total dietary fiber (TDF) in one‐day‐old bread made with commercial XL‐RS4 at 30% flour substitution increased 3–4% (db) in the control to 19.2% (db) in the test bread, while the sum of slowly digestible starch (SDS) plus resistant starch (RS), determined by a modified Englyst method, increased from 24.3 to 41.8% (db). The reference amount (50 g, as‐is) of that test bread would provide 5.5 g of dietary fiber with 10% fewer calories than control bread. Sugar‐snap cookies were made at 30 and 50% flour replacement with laboratory‐prepared XL‐RS4, potato starch, high‐amylose (70%) corn starch, and commercial heat‐moisture‐treated high‐amylose (70%) corn starch. The shape of cookies was affected by the added starches except for XL‐RS4. The reference amount (30 g, as‐is) of cookies made with commercial XL‐RS4 at 30% flour replacement contained 4.3 g (db) TDF and 3.4 g (db) RS, whereas the negative control contained 0.4 g TDF and 0.6 g RS. The retention of TDF in the baked foods containing added XL‐RS4 was calculated to be >80% for bread and 100% for cookies, while the retention of RS was 35–54% for bread and 106–113% for cookies.     

Laboratory Procedure to Wet‐Mill 100 g of Grain Sorghum into Six Fractions

A small‐scale (100 g of grain) procedure was developed to wet‐mill grain sorghum into six fractions by modifying the procedure of Eckhoff et al (1996). The wet‐milling process was repeated five times on commercial grain sorghum, and the mean yield (69.4%) of starch (≤0.3% protein) varied by 0.3%, whereas the yields of fiber, gluten, and germ plus bran fractions varied by 5–6%. The starch fraction accounted for ≈95% of that in the grain, while the total solids recovered was 99.0%. Four other samples of grain sorghum gave 92–95% recoveries of starches and 98.2–99.8% recoveries of total solids. All grain sorghum starches had lightness (L*) values and pasting curves nearly equal to those of a commercial maize starch.     

Influence of Starch and Gluten Characteristics on Rheological Properties of Wheat Flour Gel at Small and Large Deformation

Starch and gluten were isolated from 10 wheat cultivars or lines with varied amylose content. The rheological properties of 30% wheat flour gel, starch gel, and the gel of isolated gluten mixed with common starch were determined in dynamic mechanical testing under shear deformation, creep‐recovery, and compression tests under uniaxial compression. Variation of wheat samples measured as storage shear modulus (G′), loss shear modulus (G″), and loss tangent (tan δ = G″/G′) was similar between flour and starch gels and correlated significantly between flour and starch gel. The proportion of acetic acid soluble glutenin exhibited a significant relationship with tan δ of gluten‐starch mixture gel. The small difference in amylose content strongly affected the rheological parameters of flour gels in creep‐recovery measurement. Wheat flour gel with lower amylose content showed higher creep and recovery compliance that corresponded to the trend in starch gel. Compressive force of flour gel at 50 and 95% strain correlated significantly with that of starch gel. Gel mixed with the isolated gluten from waxy wheat lines appeared to have a weaker gel structure in dynamic viscoelasticity, creep‐recovery, and compression tests. Starch properties of were primarily responsible for rheological changes in wheat flour gel.     

Observations on the Quality Characteristics of Waxy (Amylose‐Free) Winter Wheats

Previous investigations have suggested waxy (amylose‐free) wheats (Triticum aestivum L.) possess weak gluten properties and may not be suitable for commercial gluten extraction. This limitation could prevent the use of waxy wheat as a source of unique starch, because gluten is a by‐product of the wheat starch purification process. Fifty waxy wheat lines were used to determine the extent to which gluten protein and other grain quality related traits might vary and, consequently, allow the development of waxy wheat with acceptable gluten properties. Among the waxy lines, significant variation was observed for all measured quality traits with the exception of flour protein concentration. No waxy entries statistically equaled the highest ranking nonwaxy entry for grain volume weight, falling number, flour yield, or mixograph mix time. No waxy lines numerically exceeded or equaled the mean of the nonwaxy controls for falling number, flour yield, or mixograph mix time. For grain and flour protein related variables, however, many waxy lines were identified well within the range of acceptability, relative to the nonwaxy controls used in this study. Approximately 50% of the waxy lines did not differ from the highest ranking nonwaxy cultivar for grain and flour protein concentrations. Forty‐three (86%) of the tested waxy lines were not sig‐nificantly different from the nonwaxy line with the highest mixograph mixing tolerance, 22/50 (44%) of the waxy wheat lines did not differ from the highest ranking nonwaxy line in gluten index scores, and 17/50 (34%) did not differ from the highest ranking nonwaxy line in extracted wet gluten. All waxy experimental lines produced gluten via Glutomatic washing. The quality of the gluten, as measured both by mixograph and gluten index, varied widely among the waxy lines tested. These observations suggest that weak gluten is not a natural consequence of the waxy trait, and waxy cultivars with acceptable gluten properties can be developed.     

Use of Proteases to Reduce Steep Time and SO2 Requirements in a Corn Wet‐Milling Process

To eliminate the diffusion barriers associated with enzyme addition during conventional steeping, we have developed a two‐stage milling procedure to evaluate the effects of enzyme addition on corn wet milling. The current study compares the effects of the addition of commercially available enzyme preparations during conventional steeping to their comparable addition in the two‐stage procedure. Results are presented in terms of yields of fiber, starch, germ, and gluten. The results demonstrate that the application of enzymes to the normal steeping step of wet milling is not an effective means of decreasing the steeping time or sulfur dioxide usage. Only when specific enzymes are added to the hydrated ground corn, using the modified two‐stage procedure, are enzymes effective in decreasing the steeping time and sulfur dioxide requirements. The overall steeping time with the two‐stage modified procedure ranges from 6 to 8 hr, representing a 67–83% reduction over the conventional process. The modified process greatly decreases, and possibly eliminates, the need for sulfur dioxide addition, while producing starch yields and quality equivalent to that from the conventional process.     

Application of Protease and High‐Intensity Ultrasound in Corn Starch Isolation from Degermed Corn Flour

The conventional corn wet‐milling process requires a long steeping time and has environmental and health concerns from the use of SO₂. A recently proposed two‐stage enzymatic milling procedure with the first stage of water soaking and coarse grinding of corn and the second stage of incubating with enzymes has been shown to reduce the soaking time and possibly eliminate the need for SO₂ addition. This current work explored the applications of protease and high‐intensity ultrasound in the second stage of the two‐stage enzymatic milling for corn starch isolation to further shorten the process time without use SO₂. of The starch yield from sonication alone was 55.2–67.8% (starch db) as compared with 53.4% of the water‐only control with stirring for 1 hr and 71.1% of the conventional control with SO₂ and lactic acid steeping for 48 hr. Protease digestion alone for 2 hr was not effective (45.8–63.9% yield) in isolating corn starch, but the starch recovery was increased to 61.2–76.1% when protease was combined with sonication. The preferred combination was neutral protease digestion for 2 hr followed by sonication at 75% amplitude for 30 min. The results demonstrated that combinations of high‐intensity ultrasound and neutral protease could replace SO₂ and shorten the steeping time in the enzymatic wet‐milling process for corn starch isolation.     

Mixograph Responses of Gluten and Gluten‐Fortified Flour for Gluten Produced by Cold‐Ethanol or Water Displacement of Starch from Wheat Flour

The total protein of gluten obtained by the cold‐ethanol displacement of starch from developed wheat flour dough matches that made by water displacement, but functional properties revealed by mixing are altered. This report characterizes mixing properties in a 10‐g mixograph for cold‐ethanol‐processed wheat gluten concentrates (CE‐gluten) and those for the water‐process concentrates (W‐gluten). Gluten concentrates were produced at a laboratory scale using batter‐like technology: development with water as a batter, dispersion with the displacement fluid, and screening. The displacing fluid was water for W‐gluten and cold ethanol (≥70% vol, ‐12°C) for CE‐gluten. Both gluten types were freeze‐dried at ‐10°C and then milled. Mixograms were obtained for 1) straight gluten concentrates hydrated to absorptions of 123–234%, or 2) gluten blended with a low protein (9.2% protein) soft wheat flour to obtain up to 16.2% total protein. The mixograms for gluten or gluten‐fortified flour were qualitatively and quantitatively distinguishable. We found differences in the mixogram parameters that would lead to the conclusion of greater stability and strength for CE‐gluten than for W‐Gluten. Differences between the mixograms for these gluten types could be markedly exaggerated by increasing the amount of water to the 167–234% range. Mixograms for evaluation of gluten have not been previously reported in this hydration range. Mixograms for fortification suggest that less CE‐gluten than W‐gluten would be required for the same effect.     

Textural Comparisons of Gluten‐Free and Wheat‐Based Doughs,Batters, and Breads

Studies were conducted with two newly developed gluten‐free bread recipes. One was based on corn starch (relative amount 54), brown rice (25), soya (12.5), and buckwheat flour (8.5), while the other contained brown rice flour (50), skim milk powder (37.5), whole egg (30), potato (25), and corn starch (12.5), and soya flour (12.5). The hydrocolloids used were xanthan gum (1.25) and xanthan (0.9) plus konjac gum (1.5), respectively. Wheat bread and gluten‐free bread made from commercial flour mix were included for comparison. Baking tests showed that wheat and the bread made from the commercial flour mix yielded significantly higher loaf volumes (P < 0.01). All the gluten‐free breads were brittle after two days of storage, detectable by the occurrence of fracture, and the decrease in springiness (P < 0.01), cohesiveness (P < 0.01), and resilience (P < 0.01) derived from texture profile analysis. However, these changes were generally less pronounced for the dairy‐based gluten‐free bread, indicating a better keeping quality. Confocal laser‐scanning microscopy showed that the dairy‐based gluten‐free bread crumb contained network‐like structures resembling the gluten network in wheat bread crumb. It was concluded that the formation of a continuous protein phase is critical for an improved keeping quality of gluten‐free bread.     

Effect of Lactic Acid on Protein Solubilization and Starch Yield in Corn Wet‐Mill Steeping: A Study of Hybrid Effects

To better understand the role of lactic acid (LA) in corn wet‐milling, steeping studies were performed on different yellow dent corn hybrids using four different solutions containing LA, sulfur dioxide (SO₂), a combination of LA and SO₂, or no added chemicals. Although there was variation in protein solubilization among the hybrids, protein release was consistently higher when LA was included in the steepwater than when it was excluded (both with and without SO₂). Several groups have reported that starch recoveries are improved when steepwater contains LA. To explore the relationship between protein solubilization and starch yield as effected by LA, several yellow dent hybrids were steeped in 0.20% SO₂ and 0.50% LA‐0.20% SO₂ solutions and milled to recover starch by a 100‐g laboratory corn wet‐milling procedure. In all instances, both starch yields and protein solubilization were enhanced in solutions containing LA. These results support the hypothesis that direct dissolution of the endosperm protein matrix by LA contributes to the improved starch recoveries.     

Starch and By‐Products from a Laboratory‐Scale Barley Starch Isolation Procedure

Starch was isolated from three different barleys with normal, highamylose, or high‐amylopectin (waxy) starch. The laboratory‐scale starch isolation procedure included crushing of grains, steeping, wet milling, and sequential filtration and washing with water and alkali, respectively. Yield and content of starch, protein, and dietary fiber, including β‐glucan, were analyzed in isolated starch and in the by‐products obtained. Starch yield was 25–34%, and this fraction contained 96% starch, 0.2–0.3% protein, and 0.1% ash. Most of the remaining starch was found in the coarse material removed by filtration after wet milling, especially for the high‐amylose barley, and in the starch tailings. Microscopy studies showed that isolated starch contained mostly A‐granules and the starch tailings contained mostly B‐granules. Protein concentration was highest in the alkali‐soluble fraction (54%), whereas dietary fiber concentration was highest in the material removed by filtration after alkali treatment for the normal and waxy barleys (55%). The β‐glucan content was especially high for the waxy barley in this fraction (26%). The study thus showed that it was possible to enrich chemical constituents in the by‐products but that there were large differences between barleys. This result indicates a need for modifications in the isolation procedures for different barleys to obtain high yields of starch and different by‐products. Valuable by‐products enriched in β‐glucan or protein, for example, may render starch production more profitable.     

Pasting Properties and Surface Characteristics of Starch Obtained from an Enzymatic Corn Wet‐Milling Process

Recently, we reported the development of an enzymatic corn wet‐milling process that reduces or eliminates sulfur dioxide requirements during steeping, considerably reduces steep time, and produces starch yields comparable to that of conventional corn wet‐milling. The best results so far, using the enzymatic corn wet‐milling procedure, were achieved when a particular protease enzyme (bromelain) was used. In this study, pasting properties and surface characteristics of starch obtained from six different enzyme treatments (three glycosidases [β‐glucanase, cellulase, and xylanase] and three proteases [pepsin, acid protease, and bromelain]) using the enzymatic corn wet‐milling procedure were evaluated and compared with those from starch obtained using the conventional corn wet‐milling procedure. Significant effects from enzymatic milling were observed on all the three starch pasting properties (peak, shear thinning, and setback). The setback viscosities of starch from all enzyme treatments were significantly lower compared with those of the control sample, indicating that starch polymers from enzymatic corn wet‐milling do not reassociate to the same extent as with the control. Comparison between bromelain treatment and the control sample showed that starch samples obtained from bromelain treatment are very similar to control starch in water‐binding capacity, molecular breakdown, and time to swell when cooked in water. Significant effects from enzymatic milling were observed on the surface characteristics of starch granules. The glycosidase treatments, especially the β‐glucanase samples, showed holes in the starch granules. No visual differences were observed in starch granules between bromelain and control samples.     

Phosphorus Concentrations and Flow in Maize Wet‐Milling Streams

Marketing of coproducts such as corn gluten meal (CGM) and corn gluten feed (CGF) is important to the maize wet‐milling industry. High phosphorus concentrations could lead to limited markets for CGF due to its potential to increase phosphorus in animal wastes. The objective was to measure the concentration and flow of phosphorus in the wet‐milling process and identify streams that could be altered. Samples were taken from 21 process streams of three facilities and the phosphorus content of each was determined. Flow of phosphorus was simulated using a computer model for a 2,700 tonne/day (105,000 bu/day) wet‐milling plant. Phosphorus concentrations of streams varied from <10 mg/kg to >14,000 mg/kg. Phosphorus content of many streams differed significantly among facilities. Flow of phosphorus (kg/day) varied dramatically among streams. However light steepwater, light gluten, and process water streams (5,960, 3,080, and 970 kg/day, respectively) accounted for much of the phosphorus flow. Modification of these streams could reduce phosphorus content of coproducts. The high phosphorus content of either CGF or CGM could be reduced markedly if phosphorus was reduced in the appropriate streams.