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Starch damage content determination: Amperometric method vs enzymatic

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Conference Paper · August 2008

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STARCH DAMAGE CONTENT DETERMINATION: AMPEROMETRIC METHOD
VS ENZYMATIC METHOD

J.T. McAllister1, C.K. Black1, O. Le Brun2, J. Algeldeh2, A. Dubat2 and J.F. Panozzo1
1
Department of Primary Industries, Horsham, Victoria, Australia
2
CHOPIN Technologies, Villeneuve-La-Garenne, France

INTRODUCTION
Starch damage content is an important parameter in evaluating flour quality. When a grain sample is
milled to produce flour, a proportion of the starch granules are mechanically damaged. Starch
granules may be cracked, fractured or crushed by the action of the mill. The starch damage
content of the flour has a considerable effect on the bread making process. On the positive
side, damaged starch has a greater capacity to absorb water than native starch. It is also
preferentially attacked by α- and β-amylases, making more sugars available during the
breadmaking process. On the negative side, too much starch damage may result in stickiness
of the dough, undesirable colour of the baked product and other problems. (Dubat 2007,
Simmonds 1989). For good quality flour, the starch damage should be optimised for dough
handling and end product usage.

There are two main techniques for quantifying starch damage content: enzymatic methods
(for example, Megazyme, Farrand and Audidier) and amperometric methods (SDmatic®).
The purpose of this study is to demonstrate that a good relationship can be established
between the enzymatic and amperometric methods.

MATERIALS AND METHODS

Thirty flour samples, covering a wide range of starch damage content values (2.7-8.5%,
Megazyme), were analysed by the Megazyme enzymatic method (AACC 76-31) and then by
the SDmatic® amperometric method (AACC 76-33). Samples were analysed in duplicate.
The amperometric method (Figure 1) is based on the work carried out by Medcalf and Gilles
(1965) which established that the iodine absorption capacity of a flour is proportional to the
starch damage content.

eInto the reaction bowl, transfer:

Carefully weigh out 1±0.1g of 3±1g potassium iodide
flour in the sample spoon f 3±1g boric acid
120mL distilled water
1 drop of 0.1M sodium thiosulfate
ePlace the reaction bowl and
spoon into place in the SDmatic®

When the solution reaches 35ºC (3-5min) the flour is

automatically transferred into the reaction bowl

3min after the flour is transferred to the reaction bowl, the SDmatic® displays the
starch damage content of the sample (5 different units including two with options)
Total analysis time: Approximately 10 minutes
®
Figure 1. SDmatic method- operating procedure.

260
During the SDmatic® analysis, iodine is electrochemically produced using the probe. The
liberated iodine results in an electrical current being produced which is measured using a pair
of electrodes. When the flour is introduced into the reaction vessel, the damaged starch will
absorb the iodine and the measured current will decrease. The greater the drop in current, the
higher the starch damage content.

The Megazyme method (figure 2) involves the hydrolysis of the damaged starch granules
using fungal alpha amylase. The resultant maltosaccharides and dextrins are completely
degraded to glucose by mean of amyloglucosidase. The liberated glucose is then reacted
further with a glucose oxidase/peroxidise reagent mixture and quantified spectroscopically.
The higher the glucose reading, the greater the starch damage.

Weigh 100±10mg of flour in a 12mL tube

d
Equilibrate at 40ºC for 2-5min
d
Add 1mL of fungal alpha amylase solution (50U/mL) pre-equilibrated at 40ºC. Vortex
d
Incubate at 40ºC for precisely 10min
d
Stop the enzymatic reaction by adding 8mL of diluted sulphuric acid (0.2% v/v)
d
Centrifuge at 3000rpm for 5min
d
Add 0.1mL aliquot to test tube. Add 0.1mL amyloglucosidase solution
d
Incubate at 40ºC for 10min
d
Add 4mL of GOPOD reagent
d
Incubate at 40ºC for 20min
d
Measure absorbance at 510nm (spectrophotometer)
d
Convert the absorbance to starch damage %

Total Analysis time: approximately 50min

Figure 2. Megazyme method- operating procedure.

RESULTS AND DISCUSSION

From the results obtained (Figure 3) there is a very significant correlation between the two
methods (R2=0.94). The SDmatic® can be set up to display a predicted Megazyme value for
each sample. A comparison between the SDmatic® predicted starch damage and the actual
Megazyme starch damage is shown in Figure 4. The mean average difference (expressed as
an absolute value) between the Megazyme reference value and the value predicted between
the SDmatic® is 0.3% (enzymatic units) while the median is 0.2%. By way of comparison:
during this study, the differences observed between two replicates reached as high as 0.4%
using the reference Megazyme method.

261
In comparing the two techniques, the SDmatic® method is much simpler than the Megazyme
method and requires far less training and operator skill. The SDmatic® method utilises no
enzymes, therefore there are no issues with contamination or loss of activity. For small
samples numbers, where the data is required rapidly, the SDmatic® is ideal. The SDmatic®
method can be further simplified by making bulk solutions for analysis rather than measuring
out reagents for each test (Figure 1). The SDmatic® is supplied with a working calibration.
However, if desired, a new calibration can be created using a suitable reference method on a
new set of samples. Thereafter, the SDmatic® may be used in preference to the reference
method, depending on laboratory requirements.
9.0

8.0
2
y = 0.0853x - 15x + 662.02
(AACC 76-31 reference values)

7.0 2
R = 0.9434
Damaged starch %

6.0

5.0

4.0

3.0

2.0

1.0

0.0
89 90 91 92 93 94 95 96 97
Damaged starch - AI% (SDmatic AACC 76-33)

Figure 3. Damaged Starch Content - Relationship between enzymatic and SDmatic®

methods.

9.0

8.0
AACC 76-31 damaged starch %

y = 0.9981x - 0.0862
7.0 2
R = 0.9434
(reference values)

6.0

5.0

4.0

3.0

2.0
2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0
AACC 76-31 damaged starch % (SDmatic prediction)

Figure 4. Damaged Starch Content - Relationship between reference values and values
predicted by the SDmatic®.

262
 CONCLUSION
This study demonstrates that there is a strong correlation between the SDmatic® and
Megazyme methods for determining starch damage content. It is therefore possible to benefit
from the simplicity, speed and precision of the SDmatic®.

REFERENCES
Dubat, A. (2007) Cereal Foods World 52:319-323.
Megazyme Starch Damage Assay procedure, AACC Method 76-31, ICC Method No. 164,
02/2008.
Medcalf, D. and Gilles, K. (1965) Cereal Chem. 42:546-557.
Rogers D.E., Gelroth J.A., Langemeier J.M. and Ranhotra G.S. (1994) Cereal Chem. 71:578-
581.
Simmonds, D.H. (1989) Fundamental aspects of wheat quality: starch and non-starchy
polysaccharides, Wheat and Wheat Quality in Australia, Australia, CSIRO. 215-242.

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