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Preliminary study of ratio of amylose and amylopectin as indicators of glycemic index and in vitro enzymatic hydrolysis of rice and wheat starches

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Abstract

Background: Wheat and rice form the staple food of large majority of the population throughout the world. Starch is a carbohydrate which is a homopolymer consisting of a large number of D-glucose units joined by α- glycosidic bonds. Starch can be separated into two fractions-amylose and amylopectin. Amylose, Amylopectin ratio is an indicator of Glycemic Index. Starches with lower amylose content will have higher glycemic indexes. Invitro digestion of starch provides efficient means of analysing carbohydrate digestibility and hence glycemic properties of foods. Invitro digestion indicates how a given food item is likely to behave in vivo, in terms of rate and extent of sugar release from starch, by stimulating physiological processes occurring in the mouth, stomach and small intestine. The rate of starch hydrolysis which gives rise to sustained release of reducing sugars would be preferred by the diabetics.Methods: In our study, starches of wheat and rice were first isolated. Amylose and Amylopectin ratio was determined to predict the glycemic index of both. Wheat and rice having the same amylose and amylopectin ratio were selected for our study. These isolated starches were subjected to invitro enzymatic hydrolysis by salivary and pancreatic amylases. Reducing sugars released after hydrolysis and incubation of 0, 5, 10, 15, 20 and 30 minutes were estimated by Folin-Wu method. Statistical analysis was carried out in the form of unpaired student’s t-test to find significant difference between means of reducing sugars release by wheat and rice during enzymatic hydrolysis.Results: Having compared the starches of wheat and rice having the same amylose-amylopectin ratio, it was seen that the rice had low digestibility than the wheat starch. Rice starch releases less amount reducing sugars gradually while the wheat starch releases more reducing sugars rapidly in a short period of time. This can be due to amylopectin A which might be present in larger quantities in wheat starch than in rice starch which assists in rapid digestion of wheat starch.Conclusions: Our findings revealed that rice starch released less reducing sugars gradually over a period of time while wheat starch released more reducing sugars rapidly. So, rice is the best option for diabetics for consumption because of its comparatively low release of reducing sugars as compared to wheat.
International Journal of Research in Medical Sciences | September 2018 | Vol 6 | Issue 9 Page 3095
International Journal of Research in Medical Sciences
Murugadass G et al. Int J Res Med Sci. 2018 Sep;6(9):3095-3098
www.msjonline.org
pISSN 2320-6071 | eISSN 2320-6012
Original Research Article
Preliminary study of ratio of amylose and amylopectin as indicators of
glycemic index and in vitro enzymatic hydrolysis of rice
and wheat starches
Gowri Murugadass, Kavita Dipnaik*
INTRODUCTION
Starch is a carbohydrate which is a homopolymer
consisting of a large number of D-glucose units joined by
α- glycosidic bonds. It is the most common carbohydrate
in human diets and is contained in large amounts in such
staple foods as potatoes, wheat, maize, rice, and cassava.
The white, granular, organic chemical is a soft, white,
tasteless powder that is insoluble in cold water, alcohol,
or other solvents. The basic chemical formula of the
Department of Biochemistry, Lokmanya Tilak Municipal Medical College, Sion, Mumbai-400022, Maharashtra, India
Received: 05 July 2018
Accepted: 31 July 2018
*Correspondence:
Dr. Kavita Dipnaik,
E-mail: kavita.dipnaik@yahoo.com
Copyright: © the author(s), publisher and licensee Medip Academy. This is an open-access article distributed under
the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial
use, distribution, and reproduction in any medium, provided the original work is properly cited.
ABSTRACT
Background: Wheat and rice form the staple food of large majority of the population throughout the world. Starch is
a carbohydrate which is a homopolymer consisting of a large number of D-glucose units joined by α- glycosidic
bonds. Starch can be separated into two fractions-amylose and amylopectin. Amylose, Amylopectin ratio is an
indicator of Glycemic Index. Starches with lower amylose content will have higher glycemic indexes. Invitro
digestion of starch provides efficient means of analysing carbohydrate digestibility and hence glycemic properties of
foods. Invitro digestion indicates how a given food item is likely to behave in vivo, in terms of rate and extent of
sugar release from starch, by stimulating physiological processes occurring in the mouth, stomach and small intestine.
The rate of starch hydrolysis which gives rise to sustained release of reducing sugars would be preferred by the
diabetics.
Methods: In our study, starches of wheat and rice were first isolated. Amylose and Amylopectin ratio was determined
to predict the glycemic index of both. Wheat and rice having the same amylose and amylopectin ratio were selected
for our study. These isolated starches were subjected to invitro enzymatic hydrolysis by salivary and pancreatic
amylases. Reducing sugars released after hydrolysis and incubation of 0, 5, 10, 15, 20 and 30 minutes were estimated
by Folin-Wu method. Statistical analysis was carried out in the form of unpaired student’s t-test to find significant
difference between means of reducing sugars release by wheat and rice during enzymatic hydrolysis.
Results: Having compared the starches of wheat and rice having the same amylose-amylopectin ratio, it was seen that
the rice had low digestibility than the wheat starch. Rice starch releases less amount reducing sugars gradually while
the wheat starch releases more reducing sugars rapidly in a short period of time. This can be due to amylopectin A
which might be present in larger quantities in wheat starch than in rice starch which assists in rapid digestion of wheat
starch.
Conclusions: Our findings revealed that rice starch released less reducing sugars gradually over a period of time
while wheat starch released more reducing sugars rapidly. So, rice is the best option for diabetics for consumption
because of its comparatively low release of reducing sugars as compared to wheat.
Keywords: Amylase, Amylopectin, Amylose, Glycemic index, Starch
DOI: http://dx.doi.org/10.18203/2320-6012.ijrms20183651
Murugadass G et al. Int J Res Med Sci. 2018 Sep;6(9):3095-3098
International Journal of Research in Medical Sciences | September 2018 | Vol 6 | Issue 9 Page 3096
starch molecule is (C6H10O5)n. Starch can be separated
into two fractionsamylose and amylopectin.1
Amylose is a helical polymer made of α-D-glucose units,
bound to each other through α(1→4) glycosidic bonds.
The carbon atoms on glucose are numbered, starting at
the aldehyde (C=O) carbon, so, in amylose, the 1-carbon
on one glucose molecule is linked to the 4-carbon on the
next glucose molecule byα(1→4) bonds.1
Amylopectin is a highly branched molecule, with (1→4)-
linked α-D-glucosyl units in chains joined by (1→6)
linkages. It consists of three types of branch chains. A-
chains are those linked to other chains (B- or C-) by their
reducing ends through α -D-(1→6) linkages, but they are
not branched themselves. B-chains are those linked to
another B-chain or a C-chain, but B-chains are branched
by A-chains or other B-chains at O-6 of a glucosyl unit.
Each amylopectin molecule has only one C-chain, which
carries the sole reducing end of the molecule.1
The ability of a meal preparation to raise the blood
glucose is called as glycemic index. Not all complex
carbohydrates are digested at the same rate within the
intestine. Hence, their hyperglycemic effect varies from
immediate to a rather slower one. The glycemic index is
high for bread, potatoes and table sugar and low for
legumes whole grain cereals. Simple carbohydrates such
as glucose, sucrose have a high glycemic index compared
to complex carbohydrates such as starch. This may be
because the digestion and absorption of complex
carbohydrates is slow and they slowly contribute to an
increase in blood sugar. When carbohydrates occur in
combined form with fats, fibre and protein, the glycemic
index is low.2,3
Amylases catalyse the hydrolysis of starch into maltose.
In the digestive systems of humans and many other
mammals, an alpha-amylase called ptyalin is produced by
the salivary glands, whereas pancreatic amylase is
secreted by the pancreas into the small intestine.
METHODS
In this study, wheat and rice having the same amylose
and amylopectin content were selected. To check the
amylose and amylopectin content, first the starch was
extracted using procedures described by Kurusawa et al,
and Badenhuizen where the rice and wheat flour were
suspended in 0.2% sodium hydroxide solution at 5oC to
remove protein. It was changed repeatedly until it gave
negative biuret test. The residue was washed with
distilled water and suspended in 0.1N sodium chloride
solution under a toluene/xylene layer. It was agitated for
3 hours and allowed to stand overnight to remove all
protein without degrading the starch. The upper layer of
protein containing toluene complex was removed by
decantation and discarded. The starch residue was
washed with distilled water. Finally, to get the final
product, the water and fat are removed using alcohol.
Ether can be used to get the dried product as it is very
volatile and evaporates faster than alcohol. The fine white
powdery product is subjected to standard confirmatory
test i.e. iodine test to establish that it is starch.4,5
Then the estimation of amylose and amylopectin content
of starch was carried out by Mc-Cready and Hassid
method i.e. 100mg of isolated starch sample was
dispersed in 10ml of distilled water. 0.5ml of 1N NaOH
solution was added and the solution was warmed in a
boiling water bath for about 5 minutes. It was then
diluted to 100ml in a volumetric flask. 1ml of this diluted
solution was transferred into 50 ml of volumetric flask
and neutralised with 1ml of 1N HCl. 1ml of iodine
reagent was then added and the solution was diluted to
50ml with distilled water. The blue coloured developed
was read in a colorimeter at 660nm.6
After that, the starches were subjected to invitro
enzymatic hydrolysis using salivary amylase and
pancreatic amylase under optimum in vitro conditions
(optimum temperature: 37oC and optimum pH: 6.9).
Reducing sugars released after hydrolysis and incubation
for 0, 5, 10, 15, 20 and 30 minutes were estimated by
Folin-Wu method by standard graph obtained from
standardisation process.7 Standardisation of Folin-Wu
method for reducing sugars estimation was carried out
using appropriate concentration range of glucose for
standard graph. Statistical analysis was carried out in the
form of unpaired student’s t-test to find significant
difference between means of reducing sugars release by
wheat and rice during enzymatic hydrolysis.8
RESULTS
Table 1 depicts the concentration of amylopectin and
amylose in both wheat and rice are represented in g%. In
both wheat and rice, amylopectin and amylose content
are exactly the same. The amylopectin content in rice is
80%-85% and that in wheat is 75% according to Guan-
Xing Chen et al.9 The value obtained in this wheat and
rice sample is slightly higher than the reference range.
The normal range of amylose content in rice is from 3% -
20% according to Abas A et al.10 And according to Guan-
Chen X et al, wheat usually consists of 25% amylose.9
According to Behall KM et al, compared to amylose,
amylopectin led to a greater increase in blood sugar and
insulin levels.11
Table 1: Concentration of amylopectin and amylose in
wheat and rice starch.
Cereal types
Amylose In g%
Wheat
12.7
Rice
12.7
Table 2 shows the amylose and amylopectin ratio in
wheat and rice. As, the amylopectin and amylose contents
are same in wheat and rice, their ratios are also same in
Murugadass G et al. Int J Res Med Sci. 2018 Sep;6(9):3095-3098
International Journal of Research in Medical Sciences | September 2018 | Vol 6 | Issue 9 Page 3097
both of them. Hence, both the starches have similar
glycemic index.
Table 2: Ratio of amylose and amylopectin in wheat
and rice starch.
Cereal types
Ratio of amylose and amylopectin
Wheat
0.137±0.007
Rice
0.137±0.001
Table 3: Reducing sugars released in mg% by wheat
and rice starch.
Time in
minutes
Reducing sugars released in mg% by
WHEAT
RICE
0
12.5±0
4.0±0
5
19.5±0.1
5.83±0.1
10
24.25±0
7.66±0.1
15
29.25±0
9.83±0.07
20
32.5±0.01
12.0±0
30
37.5±0.07
15.0±0
Table 4: Reducing sugars released in mg% by wheat
and rice starch in the following time intervals.
Time
interval
Reducing sugars released in mg% by
WHEAT
RICE
0-5
6.75±0
1.83±0.007
5-10
4.5±0.001
1.83±0.007
10-15
5.0±0.1
2.0±0
15-20
3.25±0.017
2.16±0.014
20-30
4.25±0
3.0±0
Table 3 highlights the reducing sugars released in mg%
on enzymatic hydrolysis by salivary and pancreatic
amylases by wheat and rice starch in 0, 5, 10, 15, 20, 30
minutes. Wheat starch releases reducing sugars rapidly
than the rice starch, which releases the reducing sugars
gradually over a period of time.
Table 5: Statistical analysis of reducing sugars
released by wheat and rice starch.
Reducing sugars
released by wheat
starch vs rice starch
during time intervals
Calculated
‘t’ value
Table ‘t’
value
Signifi-
cance*
0 - 5
935.6
3.18
S
5 - 10
593.3
3.18
S
10 - 15
0
3.18
NS
15 - 20
104.8
3.18
S
20 - 30
0
3.18
NS
*Where, S = Significant; NS = Non-significant
Table 4 highlights the reducing sugars released in mg%
on enzymatic hydrolysis by wheat and rice starch during
the time intervals 0-5, 5-10, 10-15, 15-20 and 20-30
minutes. During the time interval wheat released high
amounts of reducing sugars while rice released low
amounts. As time progressed the reducing sugars released
by wheat decreased while that of rice increased. So,
wheat releases more sugar into the blood more rapidly
while rice releases sugars gradually.
Table 5 shows the shows the statistical analysis (t-test) of
reducing sugars released by wheat and rice starch.
DISCUSSION
The wheat and rice starches selected had the same
amylose-amylopectin ratio, hence their glycemic index
were also same. The extracted starches were subjected to
invitro enzymatic hydrolysis by salivary amylase and
pancreatic amylase and the reducing sugars released in
mg% during the time intervals 0-5, 5-10, 10-15, 15-20
and 20-30 minutes were estimated. It was seen that the
reducing sugars released by wheat starch was highest for
the time interval 0-5 minutes but subsequently falls as
time progresses. But the opposite is seen in rice starch, it
releases reducing sugars lowest during the time interval
0-5 minutes but subsequently increases as time
progresses. This establishes the fact that wheat is digested
faster than the rice, this can be due to a specific type of
amylopectin i.e. amylopectin A which might be present in
large amounts in wheat than in rice which helps in rapid
digestion of wheat starch which in turn increases the
blood sugar level rapidly as told by William Davis in his
book “Wheat belly”.12
Hence, wheat starch has higher rate of digestibility of
starch as compared to rice.
Limitation: Different varieties of wheat and rice samples
having different amylase-amylopectin ratio were not
studied for their digestibility behaviour. Also, the
digestibility behaviours of different cultivars of wheat
and rice have not been studied
CONCLUSION
Amylose and amylopectin ratio can predict the glycemic
index of wheat and rice. Having compared the starches of
wheat and rice having the same amylose-amylopectin
ratio, it was seen that the rice had low digestibility than
the wheat starch. Rice starch releases less amount
reducing sugars gradually while the wheat starch releases
more reducing sugars rapidly in a short period of time.
This can be due to amylopectin A which might be present
in larger quantities in wheat starch than in rice starch
which assists in rapid digestion of wheat starch. Rice is
the best option for diabetics for consumption because of
its comparatively low digestibility than wheat, as
sustained slow release of reducing sugars is desirable in
diabetics.
Funding: No funding sources
Conflict of interest: None declared
Ethical approval: Not required
Murugadass G et al. Int J Res Med Sci. 2018 Sep;6(9):3095-3098
International Journal of Research in Medical Sciences | September 2018 | Vol 6 | Issue 9 Page 3098
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Cite this article as: Murugadass G, Dipnaik K.
Preliminary study of ratio of amylose and
amylopectin as indicators of glycemic index and in
vitro enzymatic hydrolysis of rice and wheat
starches. Int J Res Med Sci 2018;6:3095-8.
... It was well reported that white rice has a glycemic index of 70 or higher [16] where it contains amylase and amylopectin starches where the rice amylase is lighter and fine when the amylase is higher but it is much creamier and sticker when the amylopectin is high [17] [18]. Other studies have found a significant negative relationship between rice amylose content and [21] and a positive relationship between food carbohydrates of rice and wheat and glycemic index [22] [23]. ...
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