Project Title: Characterization of GEM line starches for resistant starch
development and biofuel production
Prepared by Hongxin Jiang, Hanyu Yangcheng and Jay-lin Jane, Department of Food Science and
Human Nutrition, Iowa State University, Ames, IA 50011
Project Overview
This report serves to document research conducted under a cooperative agreement
between ARS and Iowa State University. Specific objectives of this research project are to
(1) characterize the molecular structure and starch granule formation and develop
analytical methods to identify germplasm for high digestibility and resistant starch, (2)
characterize and develop utilizations of GEM line starch to improve nutritional value to
humans and animals, and (3) characterize GEM line normal and waxy starch for biofuel
production and to improve the yield of ethanol.
Maize endosperms of G/G, G/H, H/G, and H/H with 3, 2, 1, and 0 doses of
high-amylose modifier (HAM) gene(s) were produced by self- and inter-crossing of
GEMS-0067 (G) (a homozygous double mutant of amylose-extender (ae) and HAM genes)
and H99ae (H) (an ae single-mutant). The content of amylopectin decreased from 31.1%
to 11.8% with the increase in the HAM gene dosage from 0 to 3. The G/H starch had
similar amylose and large molecular-weight intermediate components (IC) contents (54.2%
and 7.5%, respectively) to the H/G starch (55.8 % and 7.5%, respectively) but had a larger
content of small molecular-weight IC (19.9%) than the H/G starch (14.6%). Analytical
results showed that an increase in the HAM-gene dosage of the endosperm of ae-mutant
maize increased the content of amylose and the content as well as the branch
chain-length of the small molecular-weight IC, but had little effect on the branch
chain-length of the amylopectin and large molecular-weight IC and the structure of
amylose.
Four normal GEM-lines and nine waxy GEM-lines were selected for an ethanol
production study using a cold-fermentation process, and the starch of six waxy lines
(08GEM05036, 05037, 05039, 05040, 05041 and 05042) was isolated for chemical and
structural analyses. HPLC analysis showed a range of ethanol yield from 16.49%
(08GEM05042) to 18.62% (08GEM05036) for the 13 GEM lines. The total starch content
of the 6 selected lines, ranging from 59.31% (08GEM05042) to 68.71% (08GEM05036), is
proportional to their ethanol yield, with a strong linear correlation (R2=0.94, p<0.01). The
digestive rate of dry-grind waxy corn was also highly correlated to ethanol yield (R2=0.93,
p<0.01), however, there was no correlation between the digestive rate of isolated starch
and ethanol yield. The enzymatic hydrolysis of isolated starch showed different kinetics
from that of the dry-grind corn, likely resulting from the effects of cell wall structure and
protein matrix of the dry-grind corn. Molecular weight and gyration radii of starch
molecules, branch chain length of amylopectin, thermal and pasting properties were also
studied, and results are shown in this report.
During this report period, five manuscripts and four abstracts were published. The
abstract “Starch-granule development in high-amylose maize” authored by H. Jiang et al.
1
was selected as the best student paper competition Finalist Award at AACCI annual
meeting.
Publications and Presentations
1) Jiang, H., Campbell, M., Blanco, M., and Jane, J. 2010. Characterization of maize
amylose-extender (ae) mutant starches: Part II. Structures and properties of starch
residues remaining after enzymatic hydrolysis at boiling-water temperature.
Carbohydrate Polymers. 80(1), 1-12.
2) Jiang, H., Horner, H., Pepper, T., Blanco, M., Campbell, M., and Jane, J. 2010.
Formation of elongated starch granules in high-amylose maize. Carbohydrate
Polymers. 80(2), 534-539.
3) Jiang, H., Jane, J., Acevedo, D., Green, A., Shinn, G., Schrenker, D., Srichuwong,
S., Campbell, M., and Wu, Y. 2010. Variations in starch physicochemical properties
from a generation-means analysis study using amylomaize V and VII parents.
Journal of Agricultural and Food Chemistry. 58(9), 5633-5639.
4) Jiang, H., Campbell, M., and Jane J. 2010. Characterization of maize
amylose-extender (ae) mutant starches: Part III. Structures and properties of the
Naegeli dextrins. Carbohydrate Polymers. 81(4), 885-891.
5) Jiang, H., Blanco, M., Campbell, M., and Jane J. 2010. Resistant-starch formation in
high-amylose maize during the kernel development. Journal of Agricultural and
Food Chemistry. (In press).
6) Jiang, H. and Jane, J. Oct 24-27, 2010. Starch-granule development in
high-amylose maize. Annual Meeting of American Association of Cereal Chemists
in Savannah, GA, USA.
7) Jiang, H., Horner, H. T., Pepper, T., Campbell, M. and Jane, J. July 29-31, 2010.
Formation of resistant starch and elongated starch granules in high-amylose maize
starch. Plant Polysaccharides and Applied Glycoscience Workshop, International
Carbohydrate Symposium organized by the Japanese Society of Applied
Glycoscience. Tokyo, Japan.
8) Jiang, H., Campbell, M., and Jane, J. July 17-20, 2010. Development of elongated
starch granules in high-amylose maize. IFT Annual Meeting & Food Expo in
Chicago, IL, USA.
9) Jiang, H. and Jane, J. Jun 7-9, 2010. Resistant starch in high-amylose maize starch.
7th Corn Utilization and Technology Conference in Atlanta, GA, USA
Objectives
Objective1: Characterize the molecular structure and starch granule formation and develop
analytical methods to identify germplasm for high digestibility and resistant starch;
Objective 2: Characterize and develop utilizations of GEM line starch to improve nutritional
value to humans and animals;
Objective 3: Characterize GEM line normal and waxy starch for biofuel production and to
improve the yield of ethanol.
2
Progress made in 2010
Objestive 1 & 2
Endosperm starch of corn lines, G/G, G/H, H/G, and H/H, that containing different
doses of high-amylose modifier (HAM) gene(s) was isolated and analyzed using a
Sepharose CL-2B gel-permeation column (Fig.1), following the method reported by Jane
and Chen(1992).
2
G/G
Absorbance (unit)
Absorbance (unit)
2
1.5
1
0.5
0
G/H
1.5
1
0.5
0
10
15
20
25
30
35
40
45
10
15
20
Fraction number
H/G
30
35
40
45
35
40
45
H/H
2
Absorbance (unit)
Absorbance (unit)
2
25
Fraction number
1.5
1
0.5
0
1.5
1
0.5
0
10
15
20
25
30
35
40
45
10
Fraction number
15
20
25
30
Fraction number
Fig. 1. Sepharose CL-2B gel-permeation profiles of the whole starches isolated from kernels of self- and
inter-crossed lines between GEMS-0067 (G) and H99ae (H).
, blue value;
, total carbohydrate.
The starch samples were then fractionated into amylopectin, amylose, and
intermediate component (IC) of large and small molecular-weights using 1-butanol
precipitation and gel permeation chromatography. Gel-permeation profiles of mixtures of
amylopectin and IC are shown in Fig.2. The content of amylopectin decreased from 31.1%
to 11.8% with the increase in the HAM gene dosage, from 0 to 3. The G/G starch had more
amylose (63.7%) and less small molecular-weight IC (16.8%) than the G/H starch (54.2%
and 19.9%, respectively) but had a similar content of the large molecular-weight IC to the
G/H starch (~7.5%).(Table1)
3
Absorbance (unit)
Absorbance (unit)
G/G
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0
10
15
20
25
30
35
40
G/H
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0
10
45
15
20
H/G
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0
10
15
20
25
30
25
30
35
40
45
Fraction number
Absorbance (unit)
Absorbance (unit)
Fraction number
35
40
45
H/H
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0
10
15
20
Fraction number
25
30
35
40
45
Fraction number
Fig. 2. Sepharose CL-2B gel-permeation profiles of amylopectin/IC mixtures of starches isolated from
kernels of self- and inter-crossed lines between GEMS-0067 (G) and H99ae (H).
, blue value;
,
total carbohydrate.
Table 1 Amylopectin, amylose, and intermediate component (IC) contents of starches
isolated from kernels of self- and inter-crossed lines between GEMS-0067 (G) and H99ae (H)
Sample
G/G
G/H
H/G
H/H
a
Amylopectina
(%)
Amylose
(%)
11.8  1.5
18.3  1.6
22.1  0.8
31.1  1.6
63.7  1.8
54.2  0.6
55.8  0.6
42.1  1.9
IC
Largeb
(%)
7.8  0.6
7.5  1.2
7.5  0.0
14.9  1.4
Smallb (%)
16.8  1.2
19.9  1.9
14.6  0.6
11.8  0.5
Amylopectin, amylose, and IC contents were determined using normal butanol precipitation method and
Sepharose Cl-2B gel-permeation chromatography followed by total carbohydrate determination.
b
Large molecular-weight IC molecules were in fractions 19 to 30 (see Fig. 2); Small molecular-weight IC
molecules were in fractions 31 to 45 (see Fig. 2).
Amylopectin, and large and small molecular-weight IC molecules were collected from
different GPC fractions. A fluorophore-assisted capillary-electrophoresis was used for the
analysis of branch-chain length distribution (Fig.3-5, Table2). Amylose molecular weight
was analyzed using Shodex SB-804 and SB-803 analytical columns (Showa Denko K.K.,
Tokyo, Japan), and the molecular-weight distributions are shown in Fig.6 and Table 3.
4
G/G
G/H
6
Normalized molar (%)
Normalized molar (%)
6
5
4
3
2
1
0
5
4
3
2
1
0
6
14
22
30
38
46
54
62
70
6
14
22
Degree of polymerization
H/G
38
46
54
62
70
62
70
H/H
6
Normalized molar (%)
Normalized molar (%)
6
30
Degree of polymerization
5
4
3
2
1
0
5
4
3
2
1
0
6
14
22
30
38
46
54
62
70
6
14
22
Degree of polymerization
30
38
46
54
Degree of polymerization
Fig. 3. Branch chain-length distributions of amylopectin from starches of self- and inter-crossed lines
between GEMS-0067 (G) and H99ae (H). The amylopectin was collected from GPC fractions 11-18 (see Fig.
G/G
5
4.5
4
3.5
3
2.5
2
1.5
1
0.5
0
G/H
5
Normalized molar (%)
Normalized molar (%)
2).
4
3
2
1
0
6
14
22
30
38
46
54
62
6
70
14
22
Normalized molar (%)
Normalized molar (%)
H/G
5
4.5
4
3.5
3
2.5
2
1.5
1
0.5
0
6
14
22
30
38
46
54
30
38
46
54
62
70
62
70
Degree of polymerization
Degree of polymerization
62
H/H
5
4.5
4
3.5
3
2.5
2
1.5
1
0.5
0
70
6
Degree of polymerization
14
22
30
38
46
54
Degree of polymerization
Fig. 4. Branch chain-length distributions of the large molecular-weight IC from starches of self- and
inter-crossed lines between GEMS-0067 (G) and H99ae (H). The large molecular-weight IC was collected
from GPC fractions19-30 (see Fig. 2).
5
G/G
3.5
3
2.5
2
1.5
1
0.5
0
6
14
22
30
38
G/H
4
Normalized molar (%)
Normalized molar (%)
4
46
54
62
3.5
3
2.5
2
1.5
1
0.5
0
70
6
14
22
Degree of polymerization
H/G
38
46
54
62
70
62
70
H/H
4
3.5
Normalized molar (%)
Normalized molar (%)
4
30
Degree of polymerization
3
2.5
2
1.5
1
0.5
3.5
3
2.5
2
1.5
1
0.5
0
0
6
14
22
30
38
46
54
62
70
6
Degree of polymerization
14
22
30
38
46
54
Degree of polymerization
Fig. 5. Branch chain-length distributions of the small molecular-weight IC from starches of self- and
inter-crossed lines between GEMS-0067 (G) and H99ae (H). The small molecular-weight IC was collected
from GPC fractions 31-45 (see Fig. 2).
Table 2 Molar-based branch-chain-length distributions of amylopectins separated from
starches of self- and inter-crossed lines between GEMS-0067 (G) and H99ae (H)a
Average CLb
DP≤12
DP13-24
DP25-36
DP≥37
Sample
(DP)
(%)
(%)
(%)
(%)
G/G
26.2
16.2  0.0
46.3  0.5
13.5  0.5
23.9  1.1
G/H
26.3
13.6  0.5
48.3  0.5
14.7  0.7
23.4  0.3
H/G
26.9
13.7  0.0
46.9  0.3
14.7  0.5
24.7  0.9
H/H
27.1
13.2  0.1
47.0  0.7
14.9  0.2
25.0  1.0
a The
b
amylopectin was collected from GPC fractions 11-18 (see Fig. 2).
CL = chain length; DP = degree of polymerization.
Although the G/G starch consisted of 63.7% amylose, which was greater than the G/H
(54.2%), H/G (55.8%), and H/H (42.1%) starches, amylose of all four lines showed similar
molecular-size distributions, with average molecular-sizes of DP 731-817. After a
debranching reaction, the molecular-size distribution of the debranched amylose reduced
and the average molecular size ranged from DP 417 to 487. The average branch
chain-lengths of the amylopectin of all four lines were similar (DP 26.2-27.1), which were
slightly shorter than the large molecular-weight IC counterpart (DP 27.3-27.9). The
average branch chain-lengths of the small molecular-weight IC were DP 35.4, 32.6, 29.1,
and 30.8 for the G/G, G/H, H/G, and H/H, respectively.
6
7
5
0.15
4
0.1
3
2
0.05
1
0
5
35
45
0.15
4
0.1
3
2
0.05
1
0
55
0
15
25
Elution time (min)
0.25
7
6
0.2
5
0.15
4
0.1
3
2
0.05
1
0
Normalized RI signal (%)
DP 1076
Log Mw
Normalized RI signal (%)
55
D
0.25
35
45
7
DP 1143
6
0.2
5
0.15
4
0.1
3
2
0.05
1
0
25
45
Elution time (min)
C
15
35
Log Mw
25
6
0.2
0
15
7
Log Mw
0.2
DP 1018
B
0.25
6
Log Mw
Normalized RS signal (%)
DP 1160
Normalized RI signal (%)
A
0.25
0
55
0
15
25
Elution time (min)
35
45
55
Elution time (min)
Fig. 6. Molecular-weight distributions of amylose of starches isolated from the self- and inter-crossed lines
between GEMS-0067 (G) and H99ae (H). A, G/G; B, G/H; C, H/G; and D, H/H. Solid line: before
isoamylase-debranching; dashed line: after isoamylase-debranching; linear line: a standard curve with
molecular sizes of DP 2, 3, 4, 7, 75, 141, 292, 617, 1148, and 2346.
Table 3 Average molecular-weights of amyloses separated from starches
of self- and inter-crossed lines of GEMS-0067 (G) and H99ae (H)
Before debranching
After debranching
Sample
a
(DP )
(DP)
G/G
817 ± 0
452 ± 1
G/H
731 ± 8
445 ± 10
H/G
795 ± 1
487 ± 7
H/H
756 ± 2
417 ± 1
a
DP = degree of polymerization
Objective 3
Thirteen GEM lines were planted in the field of North Central Regional Plant
Introduction Station (Ames, IA) in the 2009 crop year. Corn kernels of the lines were ground
using a cyclone mill and passed through a 0.5mm screen. The dry-grind corn was used for
raw-starch ethanol fermentation using raw-starch hydrolyzing enzymes (Novozyme,
Denmark) for 4 days. The supernatant was collected for analysis of the ethanol yield. The
ethanol yields of the 13 GEM lines are shown in Table 4. Among all the samples,
08GEM05036 gave the highest ethanol yield, 18.62%, whereas 08GEM05041 gave the
7
least yield of 16.49%.
Table 4 Ethanol yield of GEM line corn usingraw-starch fermentation
a
Sample
Ethanol yield(%)
08GEM04701(N)a
17.75±0.18
08GEM04702(N)
18.17±0.50
08GEM04703(N)
17.19±0.00
08GEM04704(N)
17.90±0.03
08GEM05036(W)
18.62±0.06
08GEM05037(W)
18.34±0.03
08GEM05038(W)
18.31±0.05
08GEM05039(W)
17.85±0.33
08GEM05040(W)
17.65±0.01
08GEM05041(W)
16.49±0.37
08GEM05042(W)
16.74±0.25
08GEM05043(W)
18.16±0.29
08GEM05044(W)
17.96±0.26
N, normal corn; W, waxy corn
Six waxy GEM lines, including the two lines that had the largest ethanol yields
(08GEM05036 and 5037), the two of the least ethanol yields (08GEMS05041 and 5042)
and the other two in between, were selected from the 13 lines for further physicochemical
analyses. Corn kernels were degerminated and decorticated, and endosperm starches
were isolated following the method reported by Li e.tal (2008). Total starch contents of the
selected waxy corn lines are shown in Table 5.
Table 5 Total starch contents of GEM waxy corn lines
Sample
Starch (%)
08GEMS05036
68.71±2.40
08GEMS05037
68.38±0.21
08GEMS05039
63.98±0.35
08GEMS05040
62.51±0.45
08GEMS05041
59.31±0.79
08GEMS05042
59.87±0.80
Starch hydrolysis rates of dry-grind corn and isolated starch samples were investigated
using the same enzymes that were used for the fermentation (raw-starch hydrolyzing
enzymes, Novozyme, Denmark). The percentage of hydrolysis at 1, 3, 6, 10, 24, 48 and 72
hours were determined. The starch hydrolysis showed a typical 1st order reaction, and thus
8
the reaction constant “K” was obtained by plotting Ln (starch %) against reaction time
(Table 6). Starch-hydrolysis curves are shown in Figure 7 and 8, with B73 as a control.
Hydrolysis rate of dry-grind corn
100
90
80
Hydrolysis(%)
70
60
50
40
08GEMS5036
08GEMS5037
08GEMS5039
08GEMS5040
08GEMS5041
08GEMS5042
B73
30
20
10
0
Time(hr) 0
5
10
15
20
25
30
35
40
45
50
55
60
65
70
Fig.7 Starch hydrolysis of dry-grind waxy corn using raw-starch hydrolyzing enzymes
Hydrolysis rate of starches
100.00
90.00
80.00
Hydrolysis(%)
70.00
60.00
50.00
08GEM05036
08GEM05037
08GEM05039
08GEM05040
08GEM05041
08GEM05042
B73
40.00
30.00
20.00
10.00
0.00
Time(hr) 0
5
10
15
20
25
30
35
40
45
50
55
60
65
Fig.8 Starch hydrolysis of isolated waxy corn starch using raw-starch hydrolyzing enzymes
9
70
Table 6 Starch-hydrolysis constants of dry-grind corn and isolated starches of GEM
waxy lines
Constant(K)a
Sample
Dry-grind
Starches
08GEM05036
0.0572
0.0598
08GEM05037
0.0550
0.0620
08GEM05039
0.0489
0.0548
08GEM05040
0.0473
0.0627
08GEM05041
0.0425
0.0582
08GEM05042
0.0426
0.0532
B73
0.0109
0.0180
aK
was obtained by measuring the slope of the line plotting ln (starch%) against reaction time.
The amylopectin branch-chain length distribution was analyzed, and results are
shown in Table 7 and Figure 9. Molecular weight and gyration radii of the amylopectin
are shown in Table 8. Pasting properties of the waxy starch samples were analyzed
using an RVA, and the results are shown in Table 9. Thermal properties of the waxy
starch samples were analyzed using a differential scanning calorimeter (DSC), and the
gelatinized starches were stored at 4°C for 7days and rescanned to determine starch
retrogradation. Results of starch thermal properties are shown in Table 10.
JMP Version 8 software (SAS Software Institute, Cary, NC, USA) was used for the
statistical analyses of the experimental data that were collected from at least duplicate
measurements. Pairwise correlation was calculated using the Pearson-product
moment approach. Correlations between different starch physiochemical properties
are shown in Table 11, and only significant linear relationship are reported.
Table 7 Branch-chain length distribution of amylopectin isolated from the GEM waxy
line starch
Sample
DP<12
DP13-24
DP25-36
DP>37
AVE CL
08GEM05036
23.81±1.39
49.43±1.76
12.93±1.04
13.83±2.11
21.41±0.75
08GEM05037
22.50±0.27
47.71±0.08
13.46±1.28
16.34±0.93
22.37±0.38
08GEM05039
22.93±0.01
48.96±0.37
12.08±0.63
16.03±1.01
22.48±0.39
08GEM05040
23.08±0.34
46.94±1.49
12.50±0.15
17.47±1.99
23.02±0.62
08GEM05041
22.30±0.24
45.33±1.22
11.84±0.56
20.53±2.02
24.27±1.07
08GEM05042
22.43±0.45
47.08±0.05
11.57±0.73
18.92±1.13
23.40±0.61
10
08GEM05036
7
08GEM05037
7
7
6
6
5
5
5
Peak Area
Peak Area
Peak Area
6
4
4
3
3
4
3
2
2
2
1
1
1
0
0
6
9
12
15
18
21
24
27
30
33
36
39
42
45
48
51
54
57
60
63
66
69
7
08GEM05040
0
6 10 14 18 22 26 30 34 38 42 46 50 54 58 62 66 70 74 78
DP
6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60 63 66 69 72
DP
DP
08GEM05041
7
7
6
5
5
Peak Area
Peak Area
6
5
Peak Area
6
4
4
3
3
3
2
2
1
1
1
0
0
6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60 63 66 69 72 75
DP
08GEM05042
4
2
0
08GEM05039
6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60 63 66 69 72 75
DP
Figure 9 Branch-chain length distribution of GEM waxy starch amylopectin.
11
6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60 63 66 69 72
DP
Table 8 Amylopectin Molecular weight and gyration radii of GEM waxy starch
Sample
08GEMS05036
Mw(×109)a
1.28±0.03d
Rz(nm)b
385.87±12.85
ρ(g/mol/nm3)c
22.28±1.69
08GEMS05037
1.47±0.04
421.43±4.91
19.60±0.41
08GEMS05039
1.41±0.02
413.00±1.84
20.04±0.00
08GEMS05040
1.78±0.08
434.40±7.07
21.71±0.13
08GEMS05041
1.36±0.03
400.00±0.99
21.20±0.70
08GEMS05042
1.67±0.05
429.33±7.35
21.05±0.82
a
Weight-average molecular weight
b
z-average radius of gyration
c
Density(ρ) = Mw/ Rz3
d
± Standard deviation
Table 9 Pasting properties of GEM waxy starch
Pasting
Peak
Sample
Temperature
(RVU)
08GEM05036
68.5°C
213.9±4.4
Hold
(RVU)
66.8±7.6
Final
(RVU)
92.7±0.5
Set-back
(RVU)
25.9±8.1
08GEM05037
69.8°C
253.3±6.5
72.5±8.2
103.9±6.9
31.4±1.3
08GEM05039
68.8°C
254.0±8.7
78.5±1.3
105.3±4.7
26.8±3.4
08GEM05040
71.0°C
235.4±1.6
76.0±1.4
106.1±1.7
30.2±3.1
08GEM05041
71.7°C
222.3±
74.3±
99.0±
24.8±
08GEM05042
72.3°C
216.1±11.8
81.7±1.5
107.3±3.7
25.6±5.2
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Table 10 Thermal properties of GEM waxy starch
Native starch
Sample
To(℃)
Tp(℃)
Tc(℃)
⊿H(J/g)
Retrograded starch
To(℃)
Tp(℃)
b
Retrogradation(%)
Tc(℃)
⊿H(J/g)
08GEMS05036 62.3±0.2 69.6±0.1 76.1±0.8 14.3±0.9
43.3±2.1 58.0±0.2 67.5±0.8 1.4±0.1
9.44±0.11
08GEMS05037 63.6±0.2 70.1±0.4 75.2±0.3 14.6±0.0
41.4±0.9 56.4±2.0 64.8±1.2 2.5±0.7
17.08±4.81
08GEMS05039 64.1±0.7 69.7±0.8 74.9±1.3 14.4±0.5
41.2±0.1 54.5±0.1 62.7±0.0 2.8±0.1
19.50±1.70
08GEMS05040 64.3±0.4 70.2±0.1 76.8±0.3 15.3±0.2
40.3±1.7 54.8±0.2 62.9±0.0 2.8±0.2
17.96±1.12
08GEMS05041 65.8±0.2 71.9±0.1 77.8±0.9 15.4±0.8
42.2±0.3 55.1±0.1 64.3±0.7 5.1±0.3
33.50±0.49
08GEMS05042 65.0±0.5 72.9±0.4 79.1±0.3 16.4±0.4
43.0±0.8 55.1±0.0 64.3±0.4 4.9±0.3
29.57±1.41
41.6±0.3 53.4±0.9 62.9±0.8 6.3±0.5
51.22
B73
63.38
67.74
73.32
12.3
a
Samples (2.0-3.0 mg, dsb) and deionized water (6.0-9.0 µl) were used for the analysis; T o, T p, T c and ΔH are onset, peak, conclusion temperature, and
enthalpy change, respectively.
b
Values were calculated from two replicates; ±Standard deviation.
13
Table 11 Correlation coefficients(R-square) between different starch physiochemical properties and ethanol production of the six GEM
waxy lines
Ethanol Y(%)
Starch(%)
Retro.(%)
Log(Mw)
Log(Rz)
Flour K
Starch K
DP<12
DP>37
AVECL
To(℃)
Ethanol Y(%) Starch(%) Retro.(%)
1
0.94**(+) 0.94**(-)
1
0.81*(+)
1
Log(Mw)
Log(Rz)
1
0.86**(+)
1
To(℃)
Tp(℃) Pasting T Peak Vis Final Vis
AVECL
0.90**(-) 0.91**(-) 0.78*(-) 0.77*(-)
0.83*(-) 0.86**(-)
0.70*(-)
0.9**(+) 0.93**(+) 0.76*(+) 0.70*(+)
0.67*(+)
0.92**(+)
0.79*(-) 0.82*(-) 0.86**(-)
0.69*(-)
Flour K Starch K DP<12 DP>37
0.93**(+)
0.91**(-)
0.80*(-)
0.81*(-)
0.71*(-) 0.92**(+)
1
1
1
Tp(℃)
0.70*(-)
0.70*(-)
1
0.99***(+)0.95***(+) 0.68*(+) 0.82*(+)
1
0.97***(+)
0.75*(+)
1
0.69*(+)
1
Pasting T
Peak Vis
Final Vis
0.81*(+)
1
1
1
a. Ethanol Y(%): ethanol yield of dry-grind corn fermentation; Starch (%): total starch content of dry-grind corn; Retro. (%): percentage of retrogradation;
Log (Mw): Log value of starch molecular weight; Log (Rz): Log value of starch gyration radii; Flour K: flour digestive constant; Starch K: starch digestive
constant; DP<12: proportion of branch-chain length<12DP; DP>37: proportion of branch-chain length>37DP; AVECL: average branch-chain length
(DP); To(°C): onset gelatinization temperature; T p(°C): peak gelatinization temperature; Pasting T: pasting temperature; Peak Vis: peak viscosity(RVU);
Final Vis: final viscosity(RVU)
b. (+) means the correlation is positive; (-) means the correlation is negative; * means p<0.05; ** means p<0.01; *** means p<0.001
14