Successive Reduction Dry Milling of Normal and Waxy Corn: Grain, Grit, and Flour Properties Sheetal Thakur, Amritpal Kaur, Narpinder Singh, and Amardeep Singh Virdi

C: Food Chemistry

Dry milling of different corn types resulted in varied proportions of germ, pericarp, grit and flour. Grit and flour produced during different reduction stages varied in particle size and chemical constituents, hence applications in food industry. In this study, recovery of different fractions and variation in physicochemical and pasting properties of grit and flour fractions obtained during 3 successive reduction dry millings of 2 normal (African tall, HQPM1) and 1 waxy corn (IC 550353) were evaluated. Waxy corn grains had the highest L∗ , a∗ , b∗ , ash, fat, and protein content and the lowest weight. Waxy and African tall gave the highest recovery of germ and pericarp, respectively. Waxy corn showed lower grit and flour recovery as compared to normal corn. Flour fractions showed higher L∗ and lower a∗ and b∗ values than grit fractions. Particle size of grit and flour fractions ranged from 840 to 982 μm and 330 to 409 μm, respectively. Fractions with larger particle size showed lower L∗ value. The b∗ value showed positive correlation with yellow pigment content. Grit and flour from the 1st reduction stage showed higher ash and fat content. Protein content was correlated positively with ash content and negatively with L∗ value. Grit and flour fractions with higher protein content had lower pasting viscosities. Pasting viscosities were higher for flours than their corresponding grits. Protein profiling of grit and flour fractions from different stages showed quantitative and qualitative differences in medium (22, 28, and 35 kDa) and low molecular weight (16, 17, and 19 kDa) polypeptides and were related to grit and flour yield.

Abstract:

Keywords: dry milling, particle size, pasting, protein, SDS-PAGE, waxy corn, yellow pigment content (YPC)

The variation in particle size, physicochemical, pasting and protein profiling of grit and flour fractions indicated differences in their suitability for various products. The high grit yield from normal corn varieties indicated their better suitability for the dry milling industry, as these industries prefer varieties that gave higher grit yield that is used for production of extruded products. Lower paste viscosities and retrogradation (indicated by setback viscosity) of grit and flour from waxy corn indicated their suitability for products where better mouth feel and refrigerated stability is required. The results of this work are useful for the corn milling industry and those involved in utilization of corn milled products.

Practical Application:

Introduction

In different studies, the effect of corn varieties (Nago and others 1997; Sandhu and others 2007) and milling conditions (MartinezFlores and others 1998) on particle size, physicochemical, and pasting properties of milled fractions have been evaluated. Liu (2009) reported the effect of particle size on color characteristics (L∗ , a∗ , and b∗ ) and protein content of ground corn fractions. The physicochemical properties of whole corn flour and dehulleddegermed corn flour were evaluated by Housson and Ayernor (2002) to predict the functional properties of products derivable from milled flours. Paulsan and Hill (1985) reported that the vitreousness was commonly associated with hardness and dry milling behavior. Rao and Rao (2006) studied the dry milling characteristics of corn grains, which were dried in a domestic microwave oven. Grit produced from different corn types vary in composition, particle size, and end-use suitability (Singh and others 2009). Lin and others (2001) reported that amylopectin in grit portion of dry milled corn had a significantly higher proportion of long chains and a postulated lower extent of chain branching than its flour counterpart. Mestres and others (1991) found that the chemical compositions (ash and protein content) and physical properties MS 20141521 Submitted 9/10/2014, Accepted 3/27/2015. Authors are with (sphericity or dent kernel percentage) could be used to predict dry Dept. of Food Science and Technology, Guru Nanak Dev Univ., Amritsar, Punjab, milling characteristics (semolina quality and quantity) of different India. Direct inquiries to author Singh (E-mail: [email protected]). yellow dent corn hybrids. Paulsen and Hill (1985) observed that yields of large flaking grits were significantly increased in corn with Corn kernel is composed of 3 main parts: germ, endosperm, and pericarp (Watson, 2003). The pericarp is the outermost covering of the grain that constitutes about 5% to 6% and the germ constitutes approximately 1.1% of the grain weight. The endosperm is the main storage tissue of the grain and constitutes up to 85% of the grain weight (Watson 2003). Dry milling of corn is done to reduce endosperm into grit of different particle sizes which is used in the production of various products like breakfast cereals and snacks (Singh and others 2014). Various methods of corn dry milling are used to obtain grit and flour fractions. Nago and others (1997) used the method of grinding whole corn grains to obtain whole corn flour. On the other hand, Mestres and others (2003) suggested milling of dehulled grains into flour. Another method involved initial dehulling and degerming of the grains to obtain grits that are finally milled into flour (Mestres and others 2003). The main objective of dry milling is to get the maximum recovery of grit with minimum contamination of hull, germ, pericarp and tip cap (Singh and others 2011). The corn varieties that produce higher amounts of fine flour are not preferred for dry milling.

C1144

Journal of Food Science r Vol. 80, Nr. 6, 2015

R  C 2015 Institute of Food Technologists

doi: 10.1111/1750-3841.12895 Further reproduction without permission is prohibited

Corn: grain, grit and flour properties . . . mal and waxy corn varieties and the variation in physicochemical, pasting and protein profile amongst grit and flours fractions obtained from 3 successive reduction dry millings of normal and waxy corn varieties.

Materials and Methods Materials African tall and HQPM1 normal corn varieties were procured from Palampur, Himachal Pradesh and waxy corn (IC 550353) was procured from J&K from 2011 to 2012 harvests.

Figure 1–Flow chart of 3 successive reduction dry milling of corn.

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low breakage susceptibilities and high-test weight. The various physicochemical parameters like ash and protein have significant correlation with grit and flour obtained from various reduction stages (Shevkani and others 2014). The literature indicated that no detailed study has compared dry milling behavior of normal and waxy corn and composition and protein profile of their fractions. Previous studies on physicochemical and extrusion behavior of corn fractions from dry milling revealed a significant variation in extrusion behavior of fractions obtained during successive reduction milling of corn (Singh and others 2009; Shevkani and others 2014). This study was carried out to evaluate the physicochemical and protein profiling of nor-

Corn: grain, grit and flour properties . . . Table 1a–Physicochemical properties of grains of different corn varieties. Variety African tall HQPM1 Waxy corn

HGW (g)

L∗

a∗

b∗

Ash content (%)

Protein content (%)

Fat content (%)

31.08b ± 0.54 20.34a ± 0.53 19.30a ± 0.28

57.97a ± 0.58 69.08b ± 0.25 69.70b ± 0.39

4.68b ± 0.31 2.48a ± 0.17 7.30c ± 0.26

18.06a ± 0.60 22.28b ± 0.42 29.16c ± 0.20

1.35a ± 0.05 1.25a ± 0.05 1.62b ± 0.02

8.62a ± 0.48 8.05a ± 0.53 11.03b ± 0.26

4.37a ± 0.02 5.55ab ± 0.05 5.91ab ± 0.99

C: Food Chemistry

Values with similar superscripts in a column do not differ significantly (p < 0.05).

Table 1b–Composition of germ and pericarp collected after successive reduction dry milling of different corn varieties. Corn type

Fraction

Reduction Stage

African tall

Germ Pericarp Germ Pericarp Germ Pericarp

R1 R3 R1 R3 R1 R3

HQPM1 Waxy

Ash content (%) 3.60d 1.98a 3.67d 2.45bc 2.25b 2.82c

± ± ± ± ± ±

0.30 0.04 0.36 0.12 0.23 0.04

Protein content (%) 9.50a 10.72b 9.24a 10.18b 11.38c 11.86c

± ± ± ± ± ±

0.57 0.74 0.57 0.15 0.70 0.39

Fat content (%) 7.22b 6.41ab 5.65a 8.62c 5.96a 9.46d

± ± ± ± ± ±

0.19 0.58 0.55 0.42 0.76 0.34

Values with similar superscripts in a column do not differ significantly (p < 0.05).

Hundred grain weight (HGW) Corn grains were randomly selected from the 3 varieties and 100 corn grains were counted and weighed and results were expressed in grams. All of the measurements were in triplicate. Color characteristics Color parameters (L∗ , a∗ , and b∗ ) of grains and their milled grit and flour fractions were determined using an Ultra Scan VIS Hunter Lab (Hunter Assoc. Laboratory Inc., Reston, Va., U.S.A.). A glass cell containing sample was placed above the light source, covered with a white plate and L∗ , a∗ , and b∗ color values were recorded. The instrument (45°/0° geometry, 10° observer) was calibrated against a standard red colored reference tile (Ls = 25.54, as = 28.89, bs = 12.03). The L∗ value indicates the lightness, 0 to 100 representing dark to light. The a∗ value gives the degree of the red-green color, with a higher positive a∗ value indicating more red. The b∗ value indicates the degree of the yellow–blue color, with a higher positive b∗ value indicating more yellow.

through the 32 mesh was designated as (R1F1) flour (500 μm), and flour (R2F2) (500 μm) and flour (R3F3) (500 μm) and that which passed with 1 mL of 0.2% iodine solution, and final volume was raised to C1146 Journal of Food Science r Vol. 80, Nr. 6, 2015

100 mL with distilled water. The mixture was kept at room temperature for 15 min, and maximum absorbance spectra from 450 to 800 nm were scanned with a spectrophotometer (Lambda Bio 35, Perkin Elmer, Norwalk, Conn., U.S.A.) to determine λmax . Blue values of iodine–starch complexes at 680 nm were measured according to the method of Takeda and others (1983).

step from 50 to 95 °C at 6 °C/min (after an equilibration time of 1 min at 50 °C), a holding phase at 95 °C for 5 min, a cooling step from 95 to 50 °C at 6 °C/min and a holding phase at 50 °C for 2 min. Peak viscosity (PV), breakdown viscosity (BDV) (peak viscosity − trough viscosity), final viscosity (FV), setback viscosity (SB) (final viscosity – trough viscosity) and pasting temperature (PT) were recorded. Each sample was analyzed in triplicate.

Pasting properties Pasting properties of finely ground fractions were studied by using a Rapid Visco Analyser (Newport Scientific Pvt. Ltd., War- Gel electrophoresis riewood, NSW 2102, Australia). Viscosity profiles of flours were Total proteins from milled whole grain and milled grit and recorded using flour suspensions (3.5 g of flour plus 24.5 g of dis- flour obtained during different reduction stages were extracted by tilled water). The temperature–time conditions included a heating the method described by Kawakatsu and others (2008). Briefly,

Figure 2–Bar diagrams showing (A) total recovery of germ, pericarp, grit and flour during dry milling (B) recovery of grit fractions during different reduction stages and (C) recovery of flour fractions during different reduction stages (HQ, HQPM1; AT, African Tall).

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Corn: grain, grit and flour properties . . .

0.31 0.53 0.02 1.35 0.19 0.32 0.39 0.15 0.01 0.16 1.98 0.22 0.84 0.15 2.36 2.32 0.57 0.73 ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± 4.99b 2.09a 2.70ab 3.85ab 3.25ab 5.05b 5.61b 2.18a 3.98ab 5.59b 5.22b 5.45b 7.32c 3.80ab 6.71bc 8.22c 5.19b 7.47c 0.07 0.03 0.08 0.33 0.05 0.44 0.25 0.25 0.27 0.18 0.12 0.38 0.12 0.07 0.08 0.46 0.33 0.35 ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

939d 982e 886c 330a 338a 348a 930d 971e 857c 355a 332a 349a 932d 840c 844c 409b 408b 406b

4.25 3.75 3.10 6.10 1.85 2.40 6.25 3.40 4.10 1.80 6.20 6.50 6.90 6.40 3.90 7.50 7.70 2.45

1.77cd 0.71a 1.14b 1.60c 1.25b 1.32b 1.26b 0.58a 0.81ab 1.29b 0.94b 1.28b 2.25d 1.40bc 1.25b 2.61e 1.48bc 2.08d

0.02 0.03 0.53 0.04 0.05 0.05 0.06 0.04 0.09 0.03 0.01 0.05 0.74 0.28 0.17 0.06 0.02 0.08

Ash content (%) Particle size (μm) Yellow pigment content (ppm)

0.92a 0.53a 0.54a 1.97b 0.86a 0.86a 16.7e 20.7f 22.6g 17.1e 20.0f 19.7f 3.27c 1.71b 1.81b 4.38d 2.63c 3.82d

0.18 0.06 0.10 0.11 0.06 0.06 0.09 0.10 0.13 0.15 0.24 0.13 0.01 0.07 0.05 0.12 0.02 0.01

Statistical analysis The data were subjected to two-way analysis of variance (ANOVA) and least significant difference (LSD) test with Minitab statistical software (version 14.12.0, Minitab, State College, Pa., U.S.A.). Pearson correlation coefficients (r) for determining the relationships between various properties were also calculated.

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± 13.21b 12.33b 13.74b 8.96a 9.18a 10.05a 28.4f 34.59g 34.44g 25.14e 28.85f 29.53f 21.08d 21.03d 21.63d 18.55c 18.17c 18.42c Flour

Grit Waxy corn

Flour

Grit HQPM1

Flour

C1148 Journal of Food Science r Vol. 80, Nr. 6, 2015

Values with similar superscripts in a column do not differ significantly (p < 0.05).

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± 0.64b 0.45b 0.65b -0.24a -0.16a −0.08a 4.73f 6.71g 6.93g 2.20c 2.90d 3.10d 3.50e 3.1d 3.65e 2.2c 1.82c 1.95c ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± 84.94fg 85.69g 84.71fg 89.17h 90.15h 89.4h 79.05d 78.93d 77.48c 85.56g 85.83g 83.74f 72.38a 75.50b 72.34a 76.57c 80.82e 79.54d R1 R2 R3 R1 R2 R3 R1 R2 R3 R1 R2 R3 R1 R2 R3 R1 R2 R3 Grit African tall

Fraction

Reduction stage

L∗

0.17 0.85 0.91 0.78 0.28 0.16 0.25 0.41 0.71 0.33 0.40 1.12 0.23 0.63 0.97 0.14 0.34 0.13

a∗

0.07 0.10 0.21 0.05 0.04 0.03 0.15 0.23 0.27 0.03 0.05 0.02 0.06 0.08 0.16 0.03 0.03 0.10

b∗

0.44 0.33 0.63 0.10 0.10 0.07 0.44 1.39 0.50 0.17 0.29 0.42 0.16 0.16 0.21 0.13 0.04 0.16

Results and Discussion

Corn type

Table 2–Hunter color parameters and composition of grit and flour from 3 successive reduction dry milling of different corn varieties.

Protein content (%)

C: Food Chemistry

100 mg of flour from different reduction stages was poured into preautoclaved Eppendorf tubes (ET) and 1.0 mL of extraction buffer (66 mM Tris buffer [pH 6.8], 8 M urea, 4% sodium dodecyl sulfate [SDS], 2% β-mercaptoethanol, 20% glycerol, 15-μL protease inhibitor cocktail [Sigma Aldrich USA]) was added. Eppendorf tubes containing flour and extraction buffer were vortexed gently and kept at 25 °C for overnight on an orbital shaker. SDSPAGE analysis was carried out according to the modified method of Laemmli (1970). A total of 15 μL of protein solution was mixed with an equal volume of sample buffer (66 mM Tris buffer [pH 6.8], 4% SDS, 4% β-mercaptoethanol, 20% glycerol) and loaded onto wells after a brief spin. Then 12% resolving and 5% stacking gels were prepared for SDS-PAGE analysis. The electrophoresis was performed at 120 V constant current for stacking and 30-mA constant current for resolving gels. SDS-PAGE gels were soaked in Coommasie brilliant blue-R250 dye (50% methanol; 10% glacial acetic acid; 0.2% w/v CBBR-250) to stain the resolved proteins. Destaining was carried out by soaking stained gels in destaining solution (20% methanol and 12% glacial acetic acid) followed documentation by using a HP Scanjet G4010 scanner at 600 dots per inch resolution.

8.43b 8.12b 8.79b 7.68ab 7.49a 8.05ab 8.47b 8.27b 8.59b 8.41b 8.19b 8.48b 11.29d 11.14d 11.70d 11.05d 10.31c 11.64d

Fat content (%)

Corn: grain, grit and flour properties . . .

Grain characteristics The grains of 2 dents and 1 waxy corn varieties were evaluated for 100 grain weight (HGW), hunter color parameters, ash content, fat content, and protein content (Table 1a). The highest HGW was observed for African tall (31.08 g) and the lowest for waxy corn (19.30 g) (Table 1a). Mestres and others (1995) reported HGW in a range from 19.3 to 31.5 g for different corn varieties. Various Hunter color parameters amongst corn varieties varied significantly. L∗ value of grains from different corn varieties ranged from 57.97 to 69.70 (Table 1a). African tall grains were white colored with the lowest L∗ (indicator of lightness) value (57.97). HQPM1 and waxy corn grains, respectively, were yellow and yellowish golden with insignificant difference in L∗ value (69.08 and 69.70, respectively). The highest L∗ value for waxy corn indicated its lighter color grains than that from other corn varieties. The lowest a∗ value was observed for HQPM1 (2.48) and b∗ value for African tall (18.06) (Table 1a). de la Parra and others (2007) observed that lutein concentration was the highest in yellow corn and the lowest in white corn. The higher “b∗ ” value has been reported to be an indication of the protein content (Singh and others 2009) and waxy corn with higher b∗ value also had higher protein content. Waxy corn grains had the highest ash (1.62%), fat (5.91%), and protein (11.03%) content (Table 1a). Collins and others (2003) also reported the higher ash, fat, and protein content of waxy corn than normal corn. Milling fractions The dry milling of 3 corn varieties gave significantly different yield of germ and pericarp. Grit and flour obtained during 3 successive reduction stages also varied significantly. Waxy corn gave

significantly higher germ (27.61%) and lower pericarp (9.52%) yield than that of African tall (germ = 5.16%, pericarp = 13.5%) and HQPM1 (germ = 5.58%, pericarp = 12.20%) (Figure 2a). Grit and flour produced from different corn types have been reported to vary in yield and composition (Singh and others 2009). Grit yields of 23.72%, 44.82%, and 31.44%, respectively, during 1st, 2nd, and 3rd reduction stage was obtained for African tall. HQPM1 and waxy corn gave grit yields of 20.30% and 14.85%, respectively during 1st, 41.96% and 71.37%, respectively, during 2nd, and 37.72% and 10.63%, respectively during 3rd reduction stage. African tall gave flour yield of 44.88%, 36.93%, and 18.18%, respectively, and HQPM1 gave the flour yield of 45.10%, 42.39%, and 12.50%, respectively, during 1st, 2nd, and 3rd reduction stage. Waxy corn gave the highest flour yield of 63.49% during 2nd reduction stage and the lowest of 7.93%, during 3rd reduction stage. Total grit and flour yield during 3 reduction stages was the lowest for waxy corn and the highest for African tall (Figure 2a). Total recovery of grit and flour was 64.58% and 14.66%, 61.53% and 18.87%, respectively for African tall and HQPM1. Whereas waxy corn gave total grit and flour recovery of 52.57% and 9.01%, respectively. Lower recovery of grit from waxy corn may be attributed to lower endosperm to pericap plus germ ratio. The recovery of grit from 3 varieties during the 2nd reduction stage was higher than that obtained during 1st and 3rd reduction stage (Figure 2b). The flour recovery from African tall and HQPM1 decreased with each successive reduction stage and waxy corn had the highest flour recovery in 2nd reduction stage and the lowest in 3rd reduction stage (Figure 2c). These differences in recovery of fractions may be due to difference in grain hardness, which may had been the major characteristics that affected the yield of coarse fraction. Another main reason for variation in grit and flour yield may be the difference in proteins present in endosperm portion of corn grain. Waxy corn that showed higher accumulation of total proteins gave the lowest total grit and flour yield whereas, African tall with less accumulation of proteins showed higher total grit and flour yield (Table 2, Figure 2a). These results were further

Physicochemical properties L∗ , a∗ , and b∗ value of various milling fractions obtained from different corn varieties is shown in Table 2. The statistical analysis revealed significant differences in L∗ , a∗ , and b∗ values amongst different corn varieties (Table 4a, 5a). L∗ , a∗ , and b∗ values of grit and flour obtained from different varieties also differed significantly. The L∗ values of flour (89.4 to 90.15) from African tall were significantly higher than that of grit (84.71 to 85.69) (Table 2). Similarly, L∗ values of flour from HQPM1 (83.74 to 85.83) and waxy corn (76.57 to 80.82) were significantly higher than their grit fraction (77.48 to 79.05 and 72.34 to 75.50, respectively). The a∗ (−0.24 to −0.08) and b∗ (8.96 to 10.05) Figure 3–SDS-PAGE analysis of total proteins of grit obtained from successive reduction dry milling of different corn varities. Total proteins from corn grit were extracted using 50 mM Tris HCl, pH 6.8, 8 M urea, 2% β-ME, 20% glycerol, and 15-μL/mL protease inhibitor cocktail. Whole grain was used as control for evaluating the differential spatial distribution of total storage proteins.

3rd Reduction

2nd Reduction

1st Reduction

Whole Grain

Waxy 3rd Reduction

2nd Reduction

1st Reduction

Whole Grain

HQPM1 3rd Reduction

2nd Reduction

1st Reduction

Whole Grain

kDa

Marker

African Tall

validated by SDS-PAGE analysis of grit and flour obtained during 3 successive reduction stages. Interestingly, the grit fractions from African tall and HQPM1 showed accumulation of high molecular weight polypeptides, whereas, grit fractions from waxy corn showed higher accumulation of low molecular weight polypeptides of 32, 30, 26, 25, and 22 kDa along with high molecular weight polypeptides (Figure 3). Higher accumulation of total protein in waxy corn may be attributed to higher accumulation of low molecular weight polypeptides since these appeared to be higher in grit fractions obtained from 3 reduction stages (Table 2, Figure 3). However, the protein profiling of flour obtained during all reduction stages did not show significant difference in accumulation of polypeptides ranging from 15 to 61 kDa (Figure 4). The protein profiling revealed the dependence of grit and flour yield on the presence or absence of some protein subunits (Figure 3 and 4). Pan and others (1996) observed that harder endosperm corn gave higher prime grit yield than soft corn with lower density. Differences in the ability of starch granules to get separated from the protein matrix may be a reason for the difference in endosperm products recovery and the degree of adhesion between starch and protein matrix that appeared to be related to corn hardness, as soft corn produces a larger amount of fine fractions and lower of grit fractions (Singh and others 2009).

205.0

97.4 66.0

43.0

29.0

20.0

87 82 75 74 66 61 55 50 46 42 38 35 32 30 26 25 22 19 15

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Corn: grain, grit and flour properties . . .

2nd Reduction

1st Reduction

3rd Reduction

2nd Reduction

43.0

29.0

20.0

C1150 Journal of Food Science r Vol. 80, Nr. 6, 2015 Waxy

97.4 66.0

61 51 45 43

39 36 35 33 31 29 28 26 25

24 22

20

17

15

14.3

Figure 4–SDS-PAGE analysis of total proteins of flour obtained from the successive reduction dry milling of different corn varities. Total proteins from corn flour were extracted by using 50-mM Tris HCl, pH 6.8, 8 M urea, 2% β-ME, 20% glycerol, and 15-μL/mL protease inhibitor cocktail.

Flour

Grit

Flour

Grit

R1 R2 R3 R1 R2 R3 R1 R2 R3 R1 R2 R3 R1 R2 R3 R1 R2 R3

Reduction stages ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

BV 0.095h 0.130l 0.120k 0.088g 0.107j 0.131l 0.089g 0.082f 0.101i 0.106j 0.097h 0.101i 0.039b 0.064e 0.051d 0.008a 0.050d 0.043c

0.01 0.02 0.01 0.03 0.05 0.06 0.03 0.04 0.01 0.01 0.08 0.04 0.03 0.06 0.02 0.02 0.03 0.03

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± 0.01 1.24 0.05 0.05 0.03 0.05 0.60 0.01 0.01 0.02 0.09 0.01 0.01 0.06 0.03 0.09 0.16 0.01

λmax (nm) 587g 582e 581de 582e 575b 597h 574a 573a 577c 576c 579d 583f 575b 577c 576bc 575b 575b 576bc

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

6.50 52.00 11.50 54.50 14.00 3.50 2.50 19.00 2.50 8.00 30.00 8.00 1.00 0.01 0.50 1.00 2.00 0.01

PV (cP) 1133c 1397e 1319d 1661h 1698h 1573g 1269d 1764i 1309d 1488f 1773i 1289d 64.00a 63.00a 59.00a 124.0b 105.0b 84.00a

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

2.50 1.00 0.50 5.50 1.50 2.00 2.00 2.00 8.00 4.00 1.00 1.00 0.58 1.00 0.50 0.1 0.01 0.01

BDV (cP) 135f 17c 1.0a 137f 96e 45d 8.0b 40d 0.5a 1.0a 0.5a 1.0a 2.0ab 7.0b 6.0b 0.5a 1.0a 1.0a

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

16.50 39.50 36.50 46.50 47.00 13.50 10.50 86.50 11.00 6.50 8.00 4.50 1.00 1.00 1.00 3.00 4.00 0.50

FV (cP) 2430b 3286c 3130c 3599e 3756f 3737f 3413d 4479h 3816f 4152g 4793i 3599e 95.00a 116.0a 108.0a 164.0a 180.0a 133.0a

1432b 1906c 1808c 2073d 2154d 2209e 2152d 2735h 2500g 2650h 2957i 2307f 29.00a 46.00a 43.00a 38.00a 75.00a 50.00a

Values with similar superscripts in a column do not differ significantly (p < 0.05). BV, blue value; λmax , lambda max; PV, peak viscosity; BDV, breakdown viscosity (peak viscosity − trough viscosity); FV, final viscosity; SB, setback (final viscosity – trough viscosity); PT, pasting temperature.

Waxy corn

HQPM1

African tall

Flour

Fraction

Grit

Corn type

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

7.50 13.50 27.50 76.50 31.50 19.00 3.00 65.50 21.50 5.50 10.50 19.00 0.01 0.01 1.00 0.50 2.00 0.50

SB (cP)

78.0e 79.6f 81.1g 76.3cd 78.0e 79.6f 82.0gh 81.1g 84.3i 81.1g 82.8h 84.3i 73.9a 75.5c 78.6e 74.6a 75.5c 77.0d

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

0.02 0.02 0.08 0.05 0.13 0.02 0.10 0.05 0.10 0.45 0.10 0.15 0.53 0.93 0.87 0.10 0.09 1.20

PT (°C)

values of flour from African tall were significantly lower than its counterpart grit fraction (a∗ = 0.45 to 0.65 and b∗ = 12.33 to 13.74). HQPM1 and waxy corn showed the similar variation in a∗ and b∗ values where flour had significantly lower a∗ and b∗ values than grit (Table 2). Higher L∗ values of African tall indicated that grit and flour of this variety were lighter in color in comparison to those fractions from other varieties. L∗ , a∗ , and b∗ values of 81.45, 2.86, and 23.49, respectively, for corn grits has been reported earlier (Jamin and Flores 1998). Highest YPC of HQPM1 might be attributed to the presence of higher xanthophylls as reported earlier (Singh and others 2011). Above mentioned statement is also confirmed by a strong positive correlation between YPC and b∗ value (r = 0.905, P ࣘ 0.005). Particles size analysis revealed the average particle size between 842 and 982 μm for grit and 330 and 409 μm for flour from different corn varieties. The particle size of flour fractions was observed to be significantly lower than its counterpart grit fractions (Table 2). Particle size showed a negative correlation with L∗ (r = −0.485, P ࣘ 0.05) (Table 6). The significantly higher L∗ value and lower a∗ and b∗ values of flour than grit was due to difference in particle size. Liu (2009) reported that as the particle size increased, the L∗ value decreased, a∗ and b∗ values increased which showed that fractions of smaller particle size were relatively lighter, less red and less yellow than fractions with larger particle size.

Table 3–Blue value, λmax , and pasting properties of grit and flour obtained from 3 successive reduction dry milling of different corn varieties.

3rd Reduction

HQPM1

2nd Reduction

1st Reduction

African Tall

3rd Reduction

kDa 1st Reduction

C: Food Chemistry Marker

Corn: grain, grit and flour properties . . .

Proximate composition Ash content (%), protein content (%), and fat content (%) of germ, pericarp, grit, and flour obtained from different corn varieties are shown in Table 1b and 2. The germ and pericarp obtained from 3 varieties showed a significant variation in ash, fat, and protein content (Table 1b). Ash content of germ from HQPM1 and African tall was significantly higher than that from waxy corn. Ash content of pericarp from African tall was significantly lower than other 2 varieties. Protein content of germ and pericarp from African tall and HQPM1 did not show any significant variation, whereas protein content of germ and pericarp from waxy corn was significantly higher than other 2 varieties. Fat content of germ was significantly lower than pericarp in HQPM1 and waxy corn (Table 1b). The statistical analysis showed significant difference in ash content of both grit and flour fractions from different reduction stages amongst different corn varieties (Table 4a, 5a). Ash content of grit and flour from African tall ranged from 0.71% to 1.77%. Ash

3rd Reduction

2nd Reduction

1st Reduction

Whole Grain

Waxy 3rd Reduction

2nd Reduction

Whole Grain

HQPM1 3rd Reduction

2nd Reduction

1st Reduction

Marker

Whole Grain

kDa

African Tall

1st Reduction

A

97.4 66.0

43.0

42

29.0 25 22 20.0 14.3

B

Figure 5–(A) Corn ethanol (70%) soluble proteins extracted from whole grain and grit obtained from 3 successive reduction dry milling stages. (B) Densitometric scaning of ethanol (70%) soluble 25 and 22 kDa poplypeptides of corn grit. The scanning of respective polypeptide bands was accomoplished by using AlphaEase FC Software v 6.0.0.

content of grit and flour from HQPM1 and waxy corn was 0.58% to 1.29% and 1.25% to 2.61%, respectively (Table 2). Ash content of corn flour from different varieties was reported in the range from 0.19% to 1.66% by Sandhu and others (2007). The higher ash content of waxy corn may be due to higher germ recovery than normal corn (Table 2 and Figure 2a). The statistical analysis showed significant difference in protein content of fractions from different reduction milling stages (Table 4a, 5a). Ash content was observed to be positively correlated with protein content (r = 0.688, P ࣘ 0.005) (Table 6). Protein content of the grit and flour fractions obtained from waxy corn (10.31 to 11.70%) was significantly higher than that from African tall (7.49 to 7.79%) and HQPM1 (8.05 to 8.59%) (Table 2). Liu (2009) reported that the corn fractions of larger particle size had higher protein content than those of smaller particle size. The statistical analysis showed significant difference in fat content of the grit and flour fractions obtained from 3 reduction stages of all the varieties (Table 4a). Protein content was negatively correlated to L∗ (r = −0.726, P ࣘ 0.005) (Table 6) and similar relationship has been observed earlier (Shevkani and others 2014). Fat content of the grit and flour fractions from African tall, HQPM1 and waxy corn ranged from 2.09% to 5.05%, 2.18% to 5.61%, and 3.80% to 8.22%, respectively (Table 2). Fat content was observed to be the highest for the grit and flour fractions from waxy corn. Alexander (1987) reported protein and fat content of 5.2% and 2% for corn flour.

Blue value and λmax The statistical analysis revealed a significant difference in blue value amongst different corn varieties at different reduction stages of grit and flour (Table 4b and 5b). Blue value of grit and flour fraction ranged from 0.088 to 0.131, 0.082 to 0.106, and 0.008 to 0.064, respectively for African tall, HQPM1, and waxy corn (Table 3, 4b, 5b). The λmax values of grit and flour ranged from 575 to 597, 572 to 583, and 575 to 577 nm, respectively, for African tall, HQPM1, and waxy corn (Table 3). The λmax value of grit and flour from waxy corn was the lowest. The λmax value of grit and flour from African tall was significantly higher which also showed the higher BDV. The iodine binding capacity allows estimation of amylose content or the ratio of long chains to short ones in amylopectin (Fiedorowicz and Rebilas, 2002). High iodine binding capacity values indicates larger amounts of long chains in the amylopectin (Lu and others 2013). Pasting properties PV, BDV, FV, SB, and PT of different milling fractions obtained from 3 reduction stages of corn varieties are shown in Table 3. The statistical analysis showed significant difference in pasting properties of the grit and flour fractions obtained from different corn varieties during 3 successive reductions milling (Table 4b, 5b). PV represents the point of maximum swelling of starch granules. The flour fractions from African tall and waxy corn from various reduction stages showed significantly higher PV than their counterpart grit fractions. PV of the flour fractions from HQPM1 and African tall ranged from 1573 to 1698 cP and 1289 to 1773 cP, respectively and its counterpart grit fractions ranged from 1133 to 1397 cP and 1269 to 1764 cP, respectively. Similarly, waxy corn flours showed PV between 84 and 124 cP and their counterpart grit fractions showed between 59 and 64 cP (Table 3). Grit as well as flour from waxy corn showed the lowest PV among all the varieties (Table 3). High crystallinity in waxy corn may have been responsible for its lower PV and PT than normal corn varieties as observed earlier (Singh and others 2006). Vol. 80, Nr. 6, 2015 r Journal of Food Science C1151

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Corn: grain, grit and flour properties . . .

Corn: grain, grit and flour properties . . . A strong negative correlation of BDV with protein content has been observed earlier (Singh and others 2014). It was reported that the presence of proteins increased the resistance of starches to shear thinning. BDV of waxy corn in each reduction stage was the lowest among all 3 varieties which may be due to the absence or least amount of amylose content as indicated by the lowest blue value and λmax . The results revealed that in the presence of higher protein, flour pastes showed lower BDV and higher paste stability (Shevkani and others 2014). FV (indicates the ability of flour to form a viscous paste) of flour (3599 to 3756 cP) was higher than that of grit (2430 to 3286 cP) from African tall. Similar trend for SB (measure of retrogradation tendency of flour upon cooling of the cooked pastes) of flour (2073 to 2209 cP) and grit (1432 to 1906 cP) of African tall was observed (Table 3). HQPM1 flour from 1st and 2nd reduction stage showed higher FV (4152 and 4793cP, respectively) than grit from similar stages (3413 and 4479 cP, respectively). HQPM1 also showed higher SB for flour (2650 and 2957 cP, respectively)

Figure 6–(A) Corn ethanol (70%) and β-ME (2%) soluble proteins extracted from whole grain and grit obtained from successive reduction dry milling of different corn varieties. (B) Densitometric scanning of ethanol (70%) and β-ME (2%) soluble 25 and 22 kDa polypeptides of corn grit. The scanning was carried out as described in Figure 5.

3rd Reduction

2nd Reduction

1st Reduction

Whole Grain

Waxy 3rd Reduction

2nd Reduction

1st Reduction

Whole Grain

HQPM1 3rd Reduction

2nd Reduction

1st Reduction

kDa

Whole Grain

African Tall

A Marker

C: Food Chemistry

PV showed negative correlation with ash content and protein content (Table 6). Sandhu and others (2007) also observed a negative correlation of protein and ash content with peak viscosity of corn flour. Singh and others (2014) also reported that the significant negative correlation of protein content with PV indicating the protective role of proteins on the integrity of the starch granules, making them resistance to mechanical shearing. BDV in corn starches has been attributed to the extent of disintegration of swollen starch granules in the absence of amylose and lipids which assist in maintaining granule integrity (Singh and others 2006). BDV of grit and flour fractions from African tall ranged from 1 to 137 cP. Grit and flour from HQPM1 and waxy corn had BDV of 0.5 to 40 cP and 0.5 to 07 cP, respectively (Table 3). The highest BDV of flour from African tall indicated its paste instability (Sandhu and others 2007). Grit and flour from waxy corn and HQPM1 had the lowest BDV which may be due to low amylose content which revealed from the lowest blue value and λmax . BDV had negative correlation with protein content (Table 6).

97.4 66.0

43.0 42 29.0 25 20.0 14.3

B

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Corn: grain, grit and flour properties . . .

Variety Stage Interaction Variety Fraction Interaction

DF

L∗

a∗

b∗

YPC

PS

AC

PC

FC

2 2 4 2 1 2

56.97∗∗ 1.53 0.67 353.48∗∗ 260.54∗∗ 3.13∗

55.61∗∗ 0.87 1.13 337.24∗∗ 192.46∗∗ 35.22∗∗

348.97∗∗ 4.44∗ 3.63∗ 638.30∗∗ 74.35∗∗ 1.39

3316∗∗ 5.59∗ 29.18∗∗ 1087∗∗ 0.60 4.06∗

0.01 0.07 0.03 0.25 3684∗∗ 20.72∗∗

26.91∗∗ 18.70∗∗ 0.57 18.31∗∗ 6.55∗ 1.53

352.56∗∗ 11.67∗∗ 1.10 318.64∗∗ 12.37∗∗ 2.68∗

20.78∗∗ 13.54∗∗ 1.03 15.43∗∗ 7.81∗ 0.4

DF, degree of freedom; YPC, yellow pigment content; PS, particle size; AC, ash content; PC, protein content; FC, fat content. ∗ P < 0.05; ∗∗ P < 0.005.

Table 4b–F values from ANOVA analysis of the data (variety versus stage and variety versus fraction) reported in Table 3.

Variety Stage Interaction Variety Fraction Interaction

DF

B.V

ƛ max

PV

BDV

FV

SB

PT

2 2 4 2 1 2

302.44∗∗ 25.24∗∗ 11.56∗∗ 125.61∗∗ 1.84 4.51∗

24.50∗∗ 7.62∗∗ 3.97∗ 17.29∗∗ 2.26 1.73

719.65∗∗ 13.22∗∗ 6.65∗∗ 580.62∗∗ 16.15∗∗ 7.02∗∗

83.62∗∗ 21.98∗∗ 24.16∗∗ 25.34∗∗ 0.61 3.82∗

887.85∗∗ 11.69∗∗ 5.75∗∗ 724.50∗∗ 16.05∗∗ 5.25∗

966.62∗∗ 9.07∗∗ 3.88∗ 958.22∗∗ 17.72∗∗ 6.14∗∗

336.28∗∗ 57.05∗∗ 1.06 113.66∗∗ 4.18∗ 1.51

DF, degree of freedom; BV, blue value; λmax , lambda max; PV, peak viscosity; BDV, breakdown viscosity; FV, final viscosity; SB, setback; PT, pasting temperature. ∗ P < 0.05; ∗∗ P < 0.005.

Table 5a–F values from ANOVA analysis of the data (variety versus stage and fraction) reported in Table 2.

African tall HQPM1 Waxy corn

Stage Fraction Interaction Stage Fraction Interaction Stage Fraction Interaction

DF

L∗

a∗

b∗

YPC

PS

AC

PC

FC

2 1 2 2 1 2 2 1 2

3.95∗ 234.36∗∗ 0.2 15.95∗∗ 512.54∗∗ 0.43 80.65∗∗ 544.15∗∗ 13.58∗∗

2.89 234.68∗∗ 2.5 172.19∗∗ 2089.12∗∗ 33.68∗∗ 36.34∗∗ 1179.12∗∗ 11.30∗∗

17.43∗∗ 516.79∗∗ 3.82∗ 116.53∗∗ 217.42∗∗ 5.36∗ 11.65∗∗ 1600.39∗∗ 7.59∗

92.44∗∗ 120.83∗∗ 21.21∗∗ 1300.07∗∗ 226.35∗∗ 177.16∗∗ 1027.47∗∗ 2005.61∗∗ 123.50∗∗

193.53∗∗ 109176∗∗ 296∗∗ 156.16∗∗ 58782∗∗ 254.39∗∗ 113.21∗∗ 25852∗∗ 103.33∗∗

15.90∗∗ 0.13 5.90∗ 139.32∗∗ 128.75∗∗ 27.47∗∗ 8.12∗ 9.93∗ 2.45

11.13∗∗ 43.29∗∗ 0.11 13.17∗∗ 3.24 1.37 17.09∗∗ 8.07∗ 3.14

12.20∗∗ 7.21∗ 11.99∗∗ 5.88∗ 17.91∗∗ 6.89∗ 8.60∗∗ 2.23 0.08

DF, degree of freedom; YPC, yellow pigment content; PS, particle size; AC, ash content; PC, protein content; FC, fat content. ∗ P < 0.05; ∗∗ P < 0.005.

Table 5b–F values from ANOVA analysis of the data (variety versus stage and fraction) reported in Table 3.

African tall HQPM1 Waxy corn

Stage Fraction Interaction Stage Fraction Interaction Stage Fraction Interaction

DF

BV

ƛ max

PV

BDV

FV

SB

PT

2 1 2 2 1 2 2 1 2

907.25∗∗ 90.77∗∗

2743.84∗∗ 185.59∗∗

2592.76∗∗ 44778.50∗∗

3847.05∗∗ 1328.95∗∗

26456.90∗∗ 150393.63∗∗

20846.95∗∗ 119551.52∗∗

149.12∗∗ 100.36∗∗ 0.15 44.99∗∗ 0.26 11.84∗∗ 80.87∗∗ 26.11∗∗ 10.68∗∗

174.64∗∗ 145.21∗∗ 378.06∗∗ 134.91∗∗ 1501.95∗∗ 1237.69∗∗ 182.99∗∗

3960.60∗∗ 157.21∗∗ 374.45∗∗ 14.25∗∗ 569.30∗∗ 461.73∗∗ 511.21∗∗

2351.66∗∗ 30234.82∗∗ 1462.50∗∗ 2086.47∗∗ 617.69∗∗ 6621.62∗∗ 369.41∗∗

370.38∗∗ 759.48∗∗ 1164.52∗∗ 745.92∗∗ 6.21∗ 76.54∗∗ 4.88∗

12056.41∗∗ 275113.58∗∗ 58403.42∗∗ 59610.97∗∗ 991.34∗∗ 10264.66∗∗ 737.69∗∗

8235.67∗∗ 19984.05 6775.56∗∗ 9066.92∗∗ 1431.50∗∗ 1350.00∗∗ 298.50∗∗

DF column as given in Table 5a is required to be inserted in Table 5b. DF, degree of freedom; BV, blue value; λmax , lambda max; PV, peak viscosity; BDV, breakdown viscosity; FV, final viscosity; SB, setback; PT, pasting temperature. ∗ P < 0.05; ∗∗ P < 0.005.

than that of grit (2152 and 2735 cP, respectively) from 1st and 2nd reduction stage (Table 3). Grit and flour fractions from waxy corn showed much less SB as compared to fractions from African tall and HQPM1 which may be attributed to the absence of amylose, because during swelling, amylose interacts with amylopectin (Tester and Morrison 1990). The lower FV and SB of grit than flour indicated the lower tendency of former to retrograde. PT of grit and flour fraction from African tall, HQPM1, and waxy corn ranged from 76.30 to 81.10 °C, 81.10 to 84.35 °C and 73.95 to 78.60 °C, respectively (Table 3). The highest PT of grit and flour fractions was observed for HQPM1 and the lowest from waxy corn. PT provides an indication of the minimum temperature required to cook flour. Seetharaman and others (2001) reported that

the pasting temperatures for Argentinian corn landraces ranged from 74.9 to 84.7 °C. Yu and others (2012) reported that proteins may significantly influence the physicochemical and pasting properties.

SDS-PAGE analysis of corn proteins Ethanol (70%) soluble proteins both under reducing and nonreducing conditions were sequentially extracted from grit obtained from 3 succesive reduction stages. Extratcion of ethanol soluble proteins under nonreducing conditions from the whole grain and grit fractions of African tall and waxy corn revealed the presence of 42, 25, and 22 kDa polypeptide subunits. However, HQPM1 did not show the presence of these ethanol soluble proteins

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Table 4a–F values from ANOVA analysis of the data (variety versus stage and variety versus fraction) reported in Table 2.

Corn: grain, grit and flour properties . . . Table 6–Pearson correlation coefficients between the various properties of grit and flour from 3 successive reduction dry milling of different corn varieties. L∗ a∗ b∗

C: Food Chemistry

PS YPC AC PC λmax PV BDV FV SB PT BV

−0.660∗∗ −0.476∗ −0.485∗ −0.726∗∗ 0.585∗ 0.761∗∗ 0.542∗ 0.688∗∗ 0.650∗∗ 0.749∗∗

a∗ 0.934∗∗ 0.415∗ 0.733∗∗

b∗

0.905∗∗

−0.533∗

−0.451∗

−0.476∗

−0.538∗ 0.572∗

YPC

−0.530∗

AC

0.688∗∗ −0.674∗∗

0.541∗ 0.596∗∗ 0.764∗∗

PC

−0.721∗∗ −0.732∗∗ −0.773∗∗ −0.686∗∗

−0.968∗∗ −0.467∗ −0.941∗∗ −0.923∗∗ −0.586∗ −0.828∗∗

BV

PV

FV

SB

PT

0.985∗∗ 0.970∗∗ 0.663∗∗ 0.829∗∗

0.997∗∗ 0.761∗∗ 0.817∗∗

0.794∗∗ 0.802∗∗

0.654∗∗

0.562∗

PS, particle size; YPC, yellow pigment content; AC, ash content; PC, protein content; BV, blue value; λmax , lambda max; PV, peak viscosity; BDV, breakdown viscosity; FV, final viscosity; SB, setback; PT, pasting temperature. ∗ P < 0.05; ∗∗ P < 0.005.

(Figure 5a). Grit from all reduction stages of African tall and waxy corn showed higher proportion of 22 and 25 kDa polypeptides as compared to their whole grain (Figure 5a and 5b). Whereas, under reducing conditions, ethanol soluble proteins obtained from grit of African tall and waxy corn from 3 successive reduction stages showed the presence of 25, 22 and 18 kDa polypeptides (Figure 6a). The absence of 25 kDa and the presence of 22, 18, 14, and 13 kDa polypeptides in HQPM1 grit was remarkable (Figure 6a). Densitometric scanning of each band was also carried out to quantitatively analyze the accumulation of these proteins. The scanning revealed that the polypeptides of 25 and 22 kDa under reducing conditions was differential and reduction stage dependent. African tall showed increased accumulaton of 25 and 22 kDa polypeptides. Whereas, storage of 25 and 22 kDa polypeptides in grit of 2nd and 3rd reduction stage of waxy variety was higher than that in grit of 1st reduction stage and lower than that of whole grain for 22 kDa polypeptides (Figure 6a and 6b). Zein proteins were reported to affect the physicochemical properties of corn flour (Singh and others 2011). α-prolamin, (α-zein) possesses lesser cysteine residues as compred to the γ -kafirin thus affect the pasting and rheological properties of corn flour (Oom and others 2008) and a positive corelation was observed among the pop corn expansion volume and α-Zein proteins and glutelins (Borras and others 2006; Mejia and others 2007; Schober and others 2008). Waxy corn grit and flour showed higher proteins compared to their counterpart fractions from African tall and HQPM1 (Table 2). To further validate these finding, proteins extracted from whole grain as well as from grits and flours obtained during the 3 succesive reduction stages was further subjected to SDS-PAGE analysis under very strong denaturing conditions by using extraction buffer (50 mM Tris HCl [pH 6.8], 8 M urea, 2% β-ME, 20% glycerol and 15-μL/mL protease inhibitor cocktail). Analysis of total proteins from grit and flour obtained after diffferent reduction stages revealed prominent variation in the medium and low molecular weight proteins (Figure 3 and 4). Polypeptides of 74, 66, 55, 46, 38, 32, and 30 kDa were only present in grit of 3 reduction stages of different corn varieties (Figure 3) and accumulation of these polypeptides was cultivars dependent. Accumulation of 66 and 50 kDa polypeptides was higher in grit from HQPM1 as compared to that of African tall and waxy corn (Figure 3). Whereas, higher accumulation of 25 and 22 kDa polypepdides was observed in grit of waxy corn (Figure 3). Total proteins in flour obtained during 3 successive reduction stages revealed variation in storage

C1154 Journal of Food Science r Vol. 80, Nr. 6, 2015

of 39, 36, 31, 28, 25, 24, 22, 17, and 15 kDa polypeptides (Figure 4). However, accumulation of these polypeptides was independent of reduction stages and no quantitative differences were observed (data not shown) (Figure 4). The presence of low molecular weight polypeptides of 19 and 15 kDa in flour and high molecular weight polypeptides of 87, 82, 75, 74, and 66 kDa in grit of 3 successive reduction stages showed significantly different composition of polypeptides in grit and flour of all varities (Figure 3 and 4). These results further confirmed the differential distributuion of corn seed storage proteins in different locations of seed during grain filling and may be responsible for varied pasting properties of corn grit and flour (Table 3) since the amylose content in all the reduction stages was almost identical as indicated by the blue value and λmax (Table 3) and the physicochemical properties of grit and flour were significantly varied (Table 3, 4a,b, and 5a,b). Aqueous ethanol soluble proteins known as zein in corn and their presence significanly affected the physicochemical properties of flour, therefore, changes in yield as well as physicochemical properties of grit and flour produced at different reduction stages may be attributed to the differential accumulation of these proteins at different locations of grain. Therefore, these findings suggests that the reduction, fractionation and identification of grit higher in protein can be used for making of gluten-free products. Earlier studies carried out by Mejia and others (2007), Schober and others (2008), and Erickson and others (2012) have demonstarted the specific role of zein proteins in processing of gluten-free products.

Conclusions The physicochemical, pasting and protein characteristics of grit and flour fractions obtained during 3 successive reductions milling of degermed grains of 2 normal (African tall, HQPM1) and 1 waxy corn differed significantly. Grain characteristics (HGW, hunter color parameters, ash, protein, and fat content) varied significantly amongst the 3 varieties. Dry milling of grains of 3 varieties gave varied grit and flour recovery at different reduction stages. Waxy corn with the lowest HGW and the highest a∗ and b∗ values gave the lowest grit and flour recovery. Particle size was larger for grit fractions than flour fractions. Grit and flour fractions obtained from HQPM1 had the highest YPC while from African tall had the lowest. Grit and flour fractions from different reduction stages showed variable amount of polypeptides. The presence of low molecular weight polypeptides of 19 and 15 kDa in flour and high molecular weight polypeptides of 87, 82, 75, 74 and 66 kDa

in grit of 3 successive reduction stages significantly differentiate the composition of polypeptides in grit and flour fractions from 3 varities. The results revealed that mealy protion of the grain that mainly consituted flour had higher amount of low molecular weight polypeptides while vitreous portion that constituted grit had higher amount of high molecular weight polypeptides.

Acknowledgments ST acknowledges UGC-BSR for providing financial assistance in the form of Fellowship. NS acknowledges Science and Engineering Research Board for providing funds in the form of a research project (SERB/SR/SO/PS/13/2011). Authors acknowledges Mr Jaghmohan Singh, Kulwant Nutrition, Batala for allowing the use of dry milling facilities.

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Corn: grain, grit and flour properties . . .