Effect of flour processing on soy beverage

1. Effect of flour processing on the quality characteristics of a soy-based
beverage
SARA ARIF1
, ASIF AHMAD1
, TARIQ MASUD1
, NAUMAN KHALID2
, IMRAN HAYAT3
,
FARZANA SIDDIQUE1
, & MUHAMMAD ALI1
1
Department of Food Technology, PMAS-Arid Agricultural University, Rawalpindi, Pakistan, 2
Department of Global
Agricultural, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo, Japan, and
3
Faculty of Agricultural, Azad Jammu & Kashmir University, Rawalakot, Pakistan
Abstract
Four treatments (roasting, germination, autoclaving and an application of 0.5% EDTA þ 0.5% sodium hydroxide) were used to
reduce the beany flavour of soya beans to produce a soy-based beverage. While germination significantly increased the protein
level as compared to the other treatments, the maximum reduction of the beany flavour was achieved by the 0.5%
EDTA þ 0.5% sodium hydroxide application. The soya beans that underwent this treatment were used during the second phase
for optimized beverage formulation. During the second phase, a beverage was prepared according to different formulations and
analysed for chemical composition and total viable count during a two-month storage period. During storage, the beverage
samples exhibited variations in several parameters. The acidity, reducing sugars and total sugars increased, while the ascorbic
acid, total soluble solids and pH decreased. Overall, chemical and microbial analyses showed the stability of the product during
the storage period.
Keywords: soya bean, beany flavour, nutritious beverage, proximate analysis, microbial analysis
Introduction
Malnutrition due to protein deficiency is a serious
problem among the masses in many developing
countries, where the diet is generally based on cereals
and, thus, fails to supply sufficient amounts of certain
essential amino acids. The problem of protein
malnutrition is most prevalent in South Asia, where
the proportion of under-nourished children is
much higher than that of even Sub-Saharan Africa
(Mehrotra 2006). This type of malnutrition may lead
to many disorders, such as weight and muscle loss,
retarded growth, kwashiorkor and marasmus (Laghari
et al. 2010). Thus, the utilization of protein-rich
ingredients in the development of nutritious products
is indispensable to combat the problems of protein
malnutrition.
The soya bean may provide a solution to protein
malnutrition. In addition to being a rich source of
protein, the soya bean contains essential amino acids
as well as appreciable amounts of lipids, carbo-
hydrates, fibre and vital minerals (Gandhi 2009). The
protein content of the dry soya bean is approximately
40%, with a good balance of essential amino acids, as
well as protein quality comparable to that of animal
proteins, such as poultry and beef (Jooyandeh 2011).
These nutritional constituents in the soya bean
together with other components, such as isoflavones,
phytosterols, steroidal saponins and other non-
isoflavone phenolic acids, play a significant role in
the reduction of several chronic diseases, such as
cardiovascular disease, cancer and osteoporosis
(Jooyandeh 2011; Mora-Escobedo et al. 2009).
Conventionally, the soya bean is processed into
products such as miso, natto and tofu. It is also used to
produce soya flour, soy sauce, soy protein meat
ISSN 0963-7486 print/ISSN 1465-3478 online q 2012 Informa UK, Ltd.
DOI: 10.3109/09637486.2012.687365
Correspondence: Nauman Khalid, Department of Global Agricultural, Graduate School of Agricultural and Life Sciences, University of Tokyo,
Japan. Tel: þ 81-80-3385-0786. E-mail: nauman_khalid120@yahoo.com
International Journal of Food Sciences and Nutrition,
December 2012; 63(8): 940–946
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2. mimics, tempeh, soya bean oil, soya milk, soya
yogurt and soya cheese (Vishwanathan et al. 2011;
Champagne et al. 2009). The soya bean also has
applications in various infant formulas, gluten-free
breads as well as protein-enriched cookies (Moore et al.
2006; Marco and Rosell 2008). Another important
aspect of the efficient utilization of the soya bean is the
production of soy-based beverages, as beverages are
highly economical and widely used food products
among all age groups (Vishwanathan et al. 2011).
However, the critical problem associated with the use
of the soya bean is its undesirable beany flavour (Endo
et al. 2004). Native chemical substances such as
isoflavones, lipoxygenase enzymes and phenolic sub-
stances are responsible for this beany flavour (Carra˜o-
Panizzi et al. 1999; Anthon and Barrett 2001). To
utilize the soya bean for product development, it is
very important to remove these off-flavouring com-
pounds during processing. This study was, therefore,
undertaken to eliminate the undesirable beany flavour
in the development of a soy-based beverage and to
evaluate the performance of that beverage.
Materials and methods
Raw materials and chemicals
The soya beans (CV: Ajmeri) were procured from the
National Agriculture Research Council in Islamabad,
Pakistan. The seeds of the soya beans were cleaned to
remove dust and other extraneous material and stored
in polyethylene bags until they were put to use. Sugar,
hydrocolloids and other ingredients (of commercial
grade quality) for the preparation of the soy-based
beverage were purchased from the local market. All the
chemicals used in the analysis were of analytical grade
purity.
First phase: reduction of the beany flavour
The aim of the first phase trials was to reduce the
beany flavour of the soya beans to an acceptable level.
The seeds were rinsed prior to undergoing the
following four different treatments: roasting (T1) for
15 min, germination (T2) for 48 h, autoclaving (T3) at
1218C for 15 min and chemical treatment (T4) of
ethylenediaminetetraacetic acid (EDTA) 0.5% þ
sodium hydroxide (NaOH 0.5%) for 4 h at a
temperature of 60 ^ 28C. The seeds were then
blanched (except for the autoclaved samples) at
1008C for 10 min, followed by wet grinding. The
resultant paste was then heated at boiling temperature
for 20 min. The soy paste was then dried and finally
ground into flour.
Proximate analysis of the soya bean flour
The crude protein, crude fat, crude fibre carbohydrate
content and ash content were determined using the
standard methods of AOAC (1990).
Sensory evaluation of the soya bean flour
A sensory evaluation of the soya bean flour during the
first phase was carried out to determine the level of
reduction of the beany flavour in the treated samples.
This evaluation was carried out by a panel of trained
judges using a nine-point Hedonic scale, where nine
was the highest score and one was the lowest score for
flavour. Uniform conditions were maintained during
the evaluation process, and the samples were evaluated
one at a time (Larmond 1977). Before commencing
the evaluation process, a 15-min briefing session was
conducted with all members of the panel. The data
were analysed for variance using the MSTATC
statistical package (when significant differences, i.e.
p , 0.05, were found), and Duncan’s multiple range
test (DMRt) was used to determine the differences
among the means (Steel et al. 1997).
Second phase: Beverge preparation
The best treatment (T4) from the first phase in terms
of the maximum reduction of the beany flavour was
selected and utilized to produce a beverage according
to the formulation shown in Table I. Syrup (water and
sugar) was prepared and pasteurized, and then
stablizers, emulsifiers and other ingredients were
Table I. Treatments and formulation of the beverage.
Treatment
codes
Concentration
of soya bean (%)
Stabilizers
(CMC þ Guargum)
(1:1) (%)
Sugar
(g/200 mL)
Ascorbic acid
(g/200 mL)
Citric acid
(g/200 mL)
Flavour
(orange)
(mL/200 mL)
Emulsifier
(Lecithin) (%)
T1 5 0.6 24 0.03 0.24 0.24 1.0
T2 5 0.8 24 0.03 0.24 0.24 1.0
T3 5 1.0 24 0.03 0.24 0.24 1.0
T4 7 0.6 24 0.03 0.24 0.24 1.0
T5 7 0.8 24 0.03 0.24 0.24 1.0
T6 7 1.0 24 0.03 0.24 0.24 1.0
T7 9 0.6 24 0.03 0.24 0.24 1.0
T8 9 0.8 24 0.03 0.24 0.24 1.0
T9 9 1.0 24 0.03 0.24 0.24 1.0
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3. added according to the given formulation. These
materials along with the soya bean flour were blended
at high speed. Finally, flavour was added and the whole
mixture was blended again. Due to the sensitivity of
the flavour, the blended beverage content was filled at
room temperature in pre-sterlized glass bottles. The
beverage was stored in these pre-sterlized bottles for
two months at room temperature.
Proximate analysis of the beverage
The acidity, sugars, ascorbic acid, total soluble solids
and pH of the beverage were determined by following
the standard methods of AOAC (1990) at 15-day
intervals for a period of 2 months.
Microbiological analysis
Total viable count. The total viable count of the
beverage was determined by the serial dilution method
explained by Fagbemi and Ijah (2006) at 15-day
intervals over the 2-month period.
Statistical analysis
The data for various experimental parameters were
analysed by using Minitab software version 13.0
(Minitab, Inc., USA). The general linear model
technique was used to analyse the variance of these
experimental parameters. In case of significant
differences among treatments, DMRt as described by
Steel et al. (1997) was used.
Results and discussion
Proximate analysis of the soya bean flour (first phase)
The results regarding the proximate composition of
the soya bean flour are presented in Table II. The
effects of different treatments on the protein, crude fat,
crude fibre, ash and carbohydrate contents were found
to be significant (p , 0.05). The different treatments
applied to the soya bean flour resulted in the increase
in ash contents, with the maximum ash contents
observed in the roasted soya bean flour (T1); this
might have been due to the concentration of minerals
through the removal of moisture during the roasting
process. The increased ash content may be explained
by the removal of moisture and the digestion of some
organic material due to the exposure to dry heat
(Rajaram and Janardhanan 1992). The autoclaving
treatment resulted in a slight increase in fat content
due to the breakdown of the lipid–protein complex in
the soya beans. Higher crude fat may favour the
lipoxygenase activity, but this possibility of lipoxygen-
ase activity was managed through high-temperature
blanching for a slightly longer period of time. The flour
obtained from the germinated soya bean seeds (T2)
showed the highest amount of crude protein content.
This increase in protein may be attributed to the
synthesis of enzyme proteins or compositional changes
following the degradation of other constituents.
Germination has been reported to improve the
nutrient profile of soya beans, especially the amino
acid content (Mostafa et al. 1987). Several treatments
resulted in the decrease in the crude fibre content of
the flour; the lowest amount of crude fibre was
detected in the flour of the autoclaved soya beans (T3),
and this might have been due to the heating of the
seeds under pressure. The maximum carbohydrate
amount was observed in the flour of the autoclaved
soya beans (T3), which was due to the decreases in the
ash and crude fibre contents of the flour.
Sensory evaluation of the soya bean flour (first phase)
The results regarding the beany flavour of the soya
bean flour differed among the treatments. The results
of DMRt shown in Table III revealed that the T4 soya
flour received the highest scores (p , 0.05) and the T2
flour received the lowest scores. These low scores for
the germinated soya bean flour might have been due to
the soaking of the seeds in water prior to germination
Table II. Effect of different treatments on chemical composition (%) of the soya bean flour.
Treatments code Ash content (%) Crude fat (%) Crude protein (%) Crude fibre (%) Carbohydrate content (%)
T0 6.40b
20.83b
37.48b
13.37a
14.19b
T1 6.97a
23.50a
37.14b
12.76a
19.63bc
T2 6.43b
21.50b
40.25a
12.90a
18.92c
T3 5.70c
23.54a
36.75b
10.70b
23.31a
T4 6.60b
21.33b
38.27b
12.83a
20.97b
Note: Values with different superscript letters in a column are statistically significant (p , 0.05).
T0 (Control); T1 (Roasting); T2 (Germination); T3 (Autoclaving); T4 (EDTA þ NaOH).
Table III. Sensory evaluation of soya bean flour for beany flavour.
Treatment Scores
T0 (Control) 3.00b
T1 (Roasted flour) 3.67b
T2 (Germinated flour) 2.67b
T3 (Autoclaved flour) 3.67b
T4 (EDTA þ NaOH treated flour) 7.00a
Note: Values with different superscript letters in a column are
statistically significant (p , 0.05).
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4. that triggered the oxidation reactions of the isoflavones
to develop some off-flavour compounds. The high
scores obtained by the T4 flour may have been due to
the application of EDTA, which is a chelating agent
and also has antioxidant properties; this may protect
against the production of the off-flavour by controlling
the oxidation of phenolic compounds. These results
are well supported by the study of Jacobsen et al.
(2008), who explained the use of EDTA as an
antioxidant. On the basis of sensory evaluation, the T4
flour was found to be the best among all four
treatments, and therefore, it was selected for the
preparation of the soy-based beverage.
Physicochemical analysis of the beverage (second phase)
Acidity. The results regarding the acidity of the
beverage are presented in Table IV. The results of
the present study indicated that the acidity of the
beverages was significantly ( p , 0.05) affected by the
interaction between the storage and treatments. Slight
variations in acidity levels were observed in different
Table IV. Physicochemical analysis of the beverage.
Treatment
codes Storage
Acidity
(%)
Reducing
sugars (%)
Non-reducing
sugars (%)
Total
sugars (%)
Ascorbic acid
(mg/100 mL)
TSS
(brix) PH
T1 S1 0.06n
0.42p,q
12.13a– e
12.36 150.6b,c
12.08 4.20
S2 0.09n
0.68o
11.92a– e
12.46 143.8f
11.36 4.13
S3 0.47f,g
1.20i
11.04b– g
12.53 138.4k,l
10.75 3.62
S4 1.57a,b
1.73j
10.63e– g
12.54 136.8m,n
10.33 3.24
S5 1.77a
2.46e,f
7.46h
12.70 128.7t,u
10.07 3.20
T2 S1 0.07n
0.22r
12.20a– e
12.43 148.1d
12.16 4.04
S2 0.24j– m
0.74n,o
11.70a– g
12.40 142.3g
11.56 3.83
S3 0.75e
1.16lc
11.33a– g
12.50 137.7lm
10.93 3.53
S4 1.40c
1.78j
10.75c– g
12.53 135.30p
10.43 3.23
S5 1.47b,c
2.43f
10.27g
12.85 129.3t
9.73 2.73
T3 S1 0.06n
0.23r
12.27a– d
12.50 152.1a
12.43 4.23
S2 0.07n
0.74n,o
11.84a– f
12.58 144.5e,f
11.77 3.83
S3 0.34g –j
1.44k
11.21a– g
12.65 136.4n,o
11.35 3.43
S4 0.96d
1.97h
10.72d– g
12.69 134.8p
10.78 3.03
S5 0.98d
2.56d,e
10.29f,g
13.73 128o,u
10.41 2.91
T4 S1 0.07n
0.32q,r
12.53a,b
12.86 150.4c
13.31 4.20
S2 0.16l– n
0.85n
12.11a– e
12.96 140.8h,i
12.91 3.82
S3 0.25i– m
1.17lk
11.84a– f
13.01 138.6k,l
12.33 3.61
S4 0.35g –j
1.83i,j
11.45a– g
13.28 135.0p
11.83 3.00
S5 0.82e
2.72b,c
11.01b– g
13.80 127.7u
11.33 2.91
T5 S1 0.07n
0.43p,q
12.56a,b
12.89 151.0b,c
13.20 4.65
S2 0.12m,n
0.68o
12.10a– e
13.09 143.4f
12.63 4.20
S3 0.25i– m
1.27lc
11.87a– e
13.40 139.0j,k
12.08 3.91
S4 0.32h– j
2.133g
11.33a– g
13.53 136.4n,o
11.53 3.60
S5 0.38g –i
2.70b,c
11.00b– g
13.84 130.7s
11.00 3.51
T6 S1 0.08n
0.33r
12.54a,b
12.87 150.9b,c
13.45 4.47
S2 0.18l– n
0.78n,o
12.37a,b
13.15 143.8f
12.95 4.33
S3 0.28i– l
1.47k
11.79a– g
13.27 139.0j,k
12.33 4.20
S4 0.43f –h
2.20g
11.55a– g
13.75 136.4n,o
11.73 3.81
S5 0.52f
2.76b
11.07b– g
13.54 130.9s
10.95 3.43
T7 S1 0.08n
0.21r
12.51a,b
12.93 152.8a
14.40 5.00
S2 0.09n
0.54p
12.33a– c
13.01 145.6e
13.83 4.13
S3 0.27i– l
1.48k
11.95a– g
13.15 139.0j,k
13.23 3.82
S4 0.32h– k
1.90h,i
11.67a– g
13.41 136.1n,o
12.58 3.32
S5 0.46f,g
2.90a
11.07b– g
13.81 132.5r
12.06 3.16
T8 S1 0.09n
0.43p,q
12.50a,b
12.94 151.6a,b
14.18 5.23
S2 0.19k –n
0.67o
12.42a,b
13.09 147.4d
13.47 4.50
S3 0.23j– m
1.47k
11.95a– g
13.42 143.6f
12.61 3.93
S4 0.38g –i
1.85h –j
11.70a– g
13.55 140.0i,j
12.08 3.71
S5 0.47f,g
2.63c,d
11.18a– g
13.91 132.5r
11.52 3.13
T9 S1 0.08n
0.32q,r
12.54a,b
12.98 152.4a
15.14 5.13
S2 0.09n
0.98m
12.67a
13.35 148.2d
14.56 4.72
S3 0.18l– n
1.53k
12.25a– d
13.52 145.3e
13.96 4.19
S4 0.24j– m
2.20g
11.49a– f
13.62 141.2g,h
13.11 3.80
S5 0.35g –j
2.80a,b
11.21a– f
12.70 133.7q
12.40 3.55
Note: Values with different superscript letters in a column are statistically significant ( p , 0.05).
T1 (soya bean 5% þ mixed stablizer 0.6%), T2 (Soya bean 5% þ mixed stablizer 0.8%), T3 (Soya bean 5% þ mixed stablizer 1%), T4 (soya bean
7% þ mixed stablizer 0.6%), T5 (soya bean 7% þ mixed stablizer 0.8%), T6 (Soya bean 7% þ mixed stablizer 1%), T7 (soya bean 9% þ mixed
stablizer 0.6%), T8 (soya bean 9% þ mixed stablizer 0.8%), T9 (soya bean 9% þ mixed stablizer 1%); S1 (day 0), S2 (day 15), S3 (day 30),
S4 (day 45), S5 (day 60).
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5. treatments, which may have been due to different
concentrations of soya bean flour. The acidity
increased linearly as a function of storage during the
two-month storage period. The increase in acidity may
have been the result of sucrose degradation during
storage as a result of the chemical and enzymatic
activity that produced acid, thereby making the
beverage more acidic with the passage of time.
During storage, the loss of the buffering capacity of
solids, such as proteins and sugars, may also have
resulted in the increased acidity of the beverage. Such
a fluctuation in acidity may also be attributed to
the degradation of soy proteins by the actions of
micro-organisms (Adeyemi et al. 1991).
Reducing sugars. The results regarding the reducing
sugars of the beverage (Table IV) indicated that the
beverage was significantly ( p , 0.05) affected by the
interaction between the treatment and storage
intervals. Variations in the reducing sugar values were
observed in different treatments, which might be
attributed to variable amounts of different reducing
sugars in the soya bean flour, such as mannose, xylose
and ribose. There was a significant difference in the
reducing sugars at all storage intervals. The results
showed that the production of reducing sugar increased
as the length of the storage period increased. This
increase in the concentration of reducing sugars was the
result of the hydrolysis of oligosaccharides. The
inversion of sucrose and the hydrolysis effect of
temperature were well explained by Stein et al. (1986).
Non-reducing sugars. The results indicated that the
non-reducing sugars of the beverage were significantly
(p , 0.05) affected by the interaction between storage
and treatments (Table IV). Variations in the non-
reducing sugar values of the beverage were observed in
different treatments, which might be ascribed to the
variation in soya bean concentrations as well as the
presence of significant amounts of sucrose in the soya
bean seeds. The non-reducing sugars decreased as a
function of storage. These results may have been due
to the degradation of these non-reducing sugars by
micro-organisms during storage (Fuleki et al. 1994).
Total sugars. The interaction between the storage
interval and treatments showed a non-significant
(p . 0.05) effect on the total sugars of the beverage
(Table IV). The results showed that the total sugar
contents of different beverages differed in various
treatments with different amounts of soya bean flour.
This was due to the fact that non-reducing sugars were
actually converted into reducing sugars, thus
maintaining the level of total sugar to a non-
significant difference.
Ascorbic acid. The results pertaining to the ascorbic
acid contents of the beverage showed a significant
( p , 0.05) effect of the interaction between the
storage interval and treatments (Table IV). Variations
in ascorbic acid levels were observed in beverages with
different amounts of soya bean flour, which might have
been due to the variable amounts of ascorbic acid in
different soya bean concentrations. The ascorbic acid
content of the beverage decreased as a function of
storage throughout the two-month storage period.
This reduction in ascorbic acid was attributed to
conversion into other substances during storage.
0.00E+00
5.00E+03
1.00E+04
1.50E+04
2.00E+04
2.50E+04
3.00E+04
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
S1
S2
S3
S4
S5
T1 T2 T3 T4 T5 T6 T7 T8 T9
Effect of treatments and storage intervals
Totalviablecount
Figure 1. Total viable count of the soy-based beverages. T1 (soya bean 5% þ mixed stablizer 0.6%), T2 (Soya bean 5% þ mixed stablizer
0.8%), T3 (Soya bean 5% þ mixed stablizer 1%), T4 (soya bean 7% þ mixed stablizer 0.6%), T5 (soya bean 7% þ mixed stablizer 0.8%), T6
(Soya bean 7% þ mixed stablizer 1%), T7 (soya bean 9% þ mixed stablizer 0.6%), T8 (soya bean 9% þ mixed stablizer 0.8%), T9 (soya bean 9%
þ mixed stablizer 1%); S1 (day 0), S2 (day 15), S3 (day 30), S4 (day 45), S5 (day 60).
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6. The results of the present study are closely
aligned with previous research (Costa et al. 2003)
that showed a decrease in ascorbic acid as a function of
storage.
Total soluble solids. The results revealed a non-
significant effect of the interaction between storage
interval and treatments for the total soluble solids
(TSS) of the beverage (Table IV). The results
indicated that the TSS differed in various treatments
with different amounts of soya bean flour. The
increasing trend of TSS may have been due to
the increased soya bean concentrations. The results of
the present study showed that TSS decreased as the
storage time increased. This decrease in TSS may have
been due to the effect of the storage temperature and
chemical changes during storage as well as the
destruction of solid contents by micro-organisms.
This trend is in line with Mugula et al. (2001), who
explained that TSS decreases as a function of storage
in both pasteurized and unpasteurized beverages made
up of sorghum.
pH. The interaction between treatments and storage
intervals showed a non-significant ( p . 0.05) effect on
the pH level of the beverage (Table IV). The results
regarding the pH of the soy beverage indicated
variations in different treatments, which may have
been due to different concentrations of soya bean
flour. The pH of the beverage decreased as the storage
duration increased. This decrease in pH during
storage might have been the result of increased acid
production by micro-organisms. The acidification of
soy beverages occurs due to the protein degradation by
micro-organisms resulting in a drop in pH over the
course of the storage period.
Microbiological analysis
Total viable count. The results regarding the total
viable count of the beverage are depicted in Figure 1,
which shows variations in the total viable count of
different treatments. The total viable count slightly
increased with increasing concentrations of soya bean
flour, and the maximum total viable count was
observed in T9. The increased total viable count in
T9 may have been the result of the increased
concentration of soya bean flour; this confirms the
finding of Viegas et al. (1985) that increasing the
amount of soya flour leads to an increase in the total
viable count. The total viable count of the beverage
varied significantly at all storage intervals. This
variation may be attributed to changes in acidity and
pH during the storage period. The increase in acidity
limited the microbial growth.
Conclusions
As a rich source of protein as well as other nutrients,
the soya bean has great potential to be transformed
into various value-added products to combat the
problems of malnutrition. It is possible to utilize a
combination of chemical treatments, such as EDTA
and NaOH, to reduce the beany flavour of soya beans
to a significant level, and such treated soya beans could
be used to manufacture nutritious beverages. Vari-
ations in the soya bean concentrations in the beverage
samples introduced small compositional changes in
the final products. Such nutritious soy-based bev-
erages have the potential to remain stable for a fairly
long period of time.
Declaration of interest: The authors report no
conflict of interest. The authors alone are responsible
for the content and writing of the paper.
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