Download Analysis of Total Sugar and Free Amino Acids in Wheat Grains

Lasbela. Uni. J. Sci. Tech., vol.3, pp. 10-13, 2014
ISSN 2306-8256
RESEARCH ARTICLE
Analysis of Total Sugar and Free Amino Acids in Wheat Grains
Samples from Five Different Regions of Balochistan
Farida Behlil1*,Samiullah2, Muzaffar Khan1,Shafia Muzaffar1, Musarat Raiz1 and Shazia A Jan1
1. Department of Chemistry, Sardar Bahadur Khan Women University, Brewery Road Quetta,
Pakistan.
2. Department of Chemistry, University of Balochistan, Sariab Road Quetta, Pakistan.
Abstract:-Wheat is an important cereal enriched with starch, proteins and fibers. In this analysis five
different samples of wheat grains grown in different ecological regions (Jaffarabad, Kohlu, Loralai, Quetta and Sibi) of
Balochistan were studied for total sugar and free amino acids. These parameters result in differences between all the
samples. Maximum concentration of both total sugar and amino acids were shown by wheat grain sample of Loralai
while minimum concentration of total sugars were shown by the sample from Sibi and minimum concentration of
amino acids were shown by sample from Jaffarabad. The differences in genotype and environmental conditions were
considered to be responsible for the variation of total sugar and free amino acids concentration in wheat grain samples
taken from different regions of Balochistan.
Key Words: Wheat, total sugar, amino acids, Balochistan.
(Tobeh and Jamaati-e-Somarin, 2011).
Wheat grains consist of the sugars,
amino acids, fats, vitamins, minerals and crude
fibers. Sugars are the most important and chief
component of wheat (Hussain and Qamar,
2007). The anthrone method for determination
of sugars has been known for many years.
Glucose is normally used to establish a
calibration curve for the anthrone method and
total sugars are expressed in terms of glucose
(Cerning-Beroard, 1975; Kumar, 2011). Another
component of wheat is amino acids which are
the monomers of proteins. Amino acids in food
components are present in free form or bound
through peptide linkage or non-peptide bonded
polymers. Analysis of the total amino acid in
food components involve protein hydrolysis by a
variety of methods that take into account
variations in permanence of individual amino
acids (Peace and Gilani, 2005). Ninhydrin is
frequently used for the identification of free
amino and carboxylic acid groups of proteins
and peptides (Friedman, 2004).
MATERIALS AND METHODS
INTRODUCTION
Wheat is a cereal grain that belongs
to the genus Triticum. In 2010 it was considered
as the third most abundant cereal plant after
maize and rice. Pakistan occupies a very
important position in wheat production. It is an
important primary food, consumed 65% by
human, 21% by animals, 8% as seeds and 6%
used in industries. It is cultivated in all provinces
of Pakistan (Noorka et al., 2009; Khan et al.,
2010).
Balochistan province is situated in the
southwest of Pakistan. The climate of
Balochistan varies from extreme cold winter to
hot summers in highlands, plain regions have
mild winter and summers are hot and dry (Khan
and Aslam, 2004; Weiss et al., 2012). Amongst
the environmental factors, temperature has a
strong effect on the growth of wheat. Wheat
growth consists of germination, requires cold
temperature (3 ᵒC) while maturation process
needs normal temperature (25-33 ᵒC). It takes
about several weeks for the formation of grain
The samples of wheat grains used in this
analysis were collected from five different
*Corresponding author: [email protected]
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regions of Balochistan such as Jaffarabad,
Kohlu, Loralai, Quetta and Sibi. These samples
were washed and then dried in oven. For
analysis these grain samples were grinded to
form flour by using electrical grinder.
To obtain standard graph, 0.5 ml of each
standard leucine solution and 0.5 ml of each of
the samples solution were taken in separate test
tubes and their volume were made 1ml by using
distill water. Then 1ml of 2% ninhydrine and
1ml of 10% pyridine solution were added to all
test tubes. All the test tubes were heated in
boiling water bath for 30 minutes and result was
obtained in violet color. The solutions of all the
test tubes were made the volume up to 50 ml by
distilled water. The absorbance was recorded by
UV-VIS spectrophotometer at 570 nm.
For the preparation of sample solution 2
grams of flour of each sample was taken and
dissolved in 75% solution of ethanol in separate
test tubes and were filtered after 24 hours. Then
these solutions were used for the analysis of
total sugar and free amino acids by using
following methods.
RESULTS AND DISCUSSION
ANALYSIS OF THE TOTAL SUGAR
Wheat is a cereal plant cultivated largely
than any other cereal in the world. It is planted
in majority of sub-tropical regions and almost all
the temperate countries of the world. In Pakistan
it is cultivated in all the provinces. It is a
primary dietary source in the world, containing
all the basic nutrients like proteins, free amino
acids,
fats,
carbohydrates
and
micronutrients.Wheat also play very important
role in the economy of every country. The wheat
nutritive value, grain yield, grain texture and
size greatly depends upon the genotype and
enviromental conditions. It depends upon soil
composition, fertilizers, stress factors like
humidity and drought.These factors are different
even in the same country as well as in the same
province. Pakistan is a country having areas
ranging from very cold to very hot enviroment
(Zeb et al., 2006).
The analysis of total sugar is estimated by
anthrone method (Cerning-Beroard, 1975).
A stock solution of glucose (0.01%) was
prepared. Glucose standards of 0.1, 0.2, 0.3, 0.4,
0.5, 0.6, 0.7, 0.8, 0.9, and 1 ml and 84%
Sulphuric acid, 0.2% Anthrone were also
prepared.
METHOD
The glucose standards and 0.1 ml of
each of the sample solution were taken in
separate test tubes and their volume was made to
1ml by adding distilled water. 4 ml of 0.2%
anthrone solution was then added to each test
tube and stirred. The test tubes were heated in
boiling water bath for (20) minutes and cooled.
Blue green colour was observed in all the test
tubes. The absorbencies of all the solutions
(glucose standards and samples) were observed
by UV-visible spectrophotometer at 620nm.
The aim of the study was to analyze the
chemical composition of wheat cultivated in
five different regions of Balochistan i.e
Jaffaraabad, Kohlu, Loralai, Quetta and Sibi.
The difference was observed in the chemical
composition of wheat samples from five
different region of Balochistan. The maximum
value (0.068 mg/2g) of free amino acid was
shown by the sample from Loralai while
minimum value (0.068 mg/2g) was shown by
the sample from Jaffarabad. The samples from
other regions also showed a marked difference
in their free amino acid concentration as shown
in table I.
ANALYSIS OF TOTAL FREE AMINO
ACIDS
The analysis of total amino acids in wheat
samples is taken by Ninhydrine method (Pandey,
1984).
Standard solutions of Leucine with
different concentration 1, 2, 4, 6, 8, and 10 mg
were made. 10% Pyridine, and 2% Ninhydrine
solutions were also prepared.
METHOD
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Similarly the maximum concentration of
total sugar (0.09 mg/2g) was shown by the
sample from Loralai and the minimum
concentration of total sugar (0.048 mg/2g) was
shown by the sample from Sibi. The samples
from other regions also showed a marked
difference in their total sugar contents as shown
in table II.
optimum temperature around 25-33°C which
allow the grain to stores greater amount of sugar
and other nutrients while the Sibi which is the
hottest region provide high temperature during
grain formation as a result decrease the time of
grain maturation which affect the accumulation
of sugar and other nutrients. Similar results were
also conducted by Buriro et al., 2011 by using
five different varieties of wheat grains.
Temperature is also responsible for high
amino acid concentration in wheat sample from
Loralai and low concentration in wheat sample
form Jaffarabad. The same result was also found
by Balla et al, 2009 that the high temperature
treatments at grain formation stage destroy the
quality of wheat grains. But along the
temperature change this difference may be due
change in the salinity of the different areas of
Balochistan which is the cause of formation of
less sugar and other nutrients which was also
determined by Rahman et al., 2009. The water
availability and nutrients in the water also affect
the
chemical
composition
and
other
characteristics of wheat as the water stress also
showed significant decline in wheat quality due
decrease in nutrient availability in water as
determined by Noorka et al., 2009. The type and
amount of fertilizers is also responsible for this
change in the concentration of amino acids and
total sugar because fertilizers provide the basic
elements required for the chemical parameters as
the same result was obtained by the Sameen et
al., 2002.
Table I: Total free amino acids in wheat
samples
Regions
Jaffarabad
Absorbance
(nm)
0.14
Total free amino acids
(mg/2g of sample)
0.014
Sibi
0.21
0.044
Quetta
0.17
0.026
Kohlu
0.23
0.052
Loralai
0.27
0.068
Table II: Total sugars in wheat samples.
Samples
Absorbance
(nm)
Total sugar
(mg/2g of sample)
Jaffarabad
1.8
0.068
Sibi
1.50
0.048
Quetta
2.2
0.09
Kohlu
2.0
0.084
Loralai
2.3
0.099
Among the environmental factors,
temperature is the most important contributing
factor controlling wheat germination and wheat
quality. Wheat consists of two phases; one is
wheat germination which requires the
temperature above 3 °C and another is wheat
grain formation which require temperature 25-33
°C. The 25-33 °C temperature is required for
grain maturation for several weeks. In the grain
maturation the sucrose is converted into starch
and the sugar of leaves, stems and other parts
transfer to the grain and store in it (Tobeh and
Jamaati-e-Somarin, 2011). The results of this
analysis showed maximum sugar concentration
in Loralai region because during grain formation
in the month of March and April it provide
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(Received 24th July 2014, Accepted 3rdOctoberr 2014)
Article can be viewed online at www.luawms.edu.pk
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