The application of Stevia as sugar substitute in carbonated drinks using Response Surface Methodology
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J. Trop. Agric. and Fd. Sc. 40(1)(2012): 23– 34 K. Saniah and M. Sharifah Samsiah
The application of Stevia as sugar substitute in carbonated drinks
using Response Surface Methodology
(Penggunaan Stevia sebagai pengganti gula di dalam minuman berkarbonat
menggunakan Kaedah Gerak Balas Permukaan)
K. Saniah* and M. Sharifah Samsiah**
Keywords: Stevia, carbonated drinks, response surface methodology, sensory evaluation,
nutritional value
Abstract
A central composite design and response surface methodology were used to
establish the optimum level of Stevia sweetener as a replacement of sucrose in
carbonated drinks. In this study, Stevia-sucrose combinations in the range of
0.2–0.5% (Stevia) and 0–54% (sucrose) were the independent variables and
their effects on sensory acceptability and physico-chemical profile of product
were investigated. Based on the response surface and superimposed plots, the
desired sensory quality of orange flavoured carbonated drink was obtained by
incorporating 0.43% Stevia and 33.13% sucrose in syrup. Results also revealed
that the amount of calorie and carbohydrate was reduced by 42.9%, indicating
that Stevia has a good potential to be exploited as an alternative sweetener for
carbonated drinks.
Introduction moved beyond choosing food and drinks
Carbonated beverages are generally simply to maintain everyday health (Geuns
sweetened, flavoured, acidified, artificially 2003).
coloured and carbonated, and sometimes There are only a few low calorie
chemically preserved. The major ingredient sweeteners as well as herb-based sweetening
of carbonated soft drink is sugar, where agents that have been introduced into the
sucrose is the most common sweetener market to meet consumers needs. Aspartame
used in the product. Lately, the increasing is not stable at higher temperatures and
consumption of sugar is known as a loses its sweetening ability. Likewise,
common factor associated with the rising saccharin has limited accessibility because
number of diabetics and increase in of certain restrictions by the Food and
obesity. The health concerns together with Drug Administration (FDA) as well as the
the continuous increase of sugar prices Malaysian Food Act and Regulations (Anon.
drives people to look at other alternative 2000). Since December 2008, the US FDA
sweeteners or low calorie-based sugar for approved the Stevia-derived sweetener Reb
their daily intake. There is an increasing A as generally recognized as safe (GRAS)
demand in society for more sugar substitutes for its use in foods and beverages (Puri et.
in processed products. Consumers have also al. 2011). Stevia rebaudiana is a perennial
*MARDI Station, Johor Bahru, No. 13, Jalan Bakti, Larkin Industrial Area, 80350 Johor Bahru, Johor, Malaysia
**Food Technology Research Centre, MARDI Headquarters, Serdang, P.O. Box 12301,
50774 Kuala Lumpur, Malaysia
Authors’ full names: Saniah Kormin and Sharifah Shamsiah Mohamed
E-mail: saniah@mardi.gov.my
©Malaysian Agricultural Research and Development Institute 2012
23Stevia in carbonated drinks
herb with claimed medicinal and culinary purchased from PureCircle Sdn Bhd, Negeri
characteristics. Stevioside and rebaudioside Sembilan. The effect of two independent
A, the sweet components in Stevia, are variables (percentages of sucrose and Stevia)
approximately 200–300 times sweeter than in syrup formulation were studied. Response
sucrose (Soejarto et al. 1982; Hanson and de surface methodology (RSM) using 22 central
Oliveira 1993). composite design (Khuri and Cornell 1987)
Stevia plant extracts as well as was used to evaluate the effect of different
stevioside have been used for a long time formulations on the sensory and physico-
as a sweetener in Japan, America, Asia and chemical profiles of carbonated drink. Each
some other countries. Stevia contributes of the variables to be optimized was coded
about 40% of the sweetener market in at three levels: -1 (minimum), 0 (medium)
Japan (Lester 1999). It is officially used and +1 (maximum). The minimum and
as a low calorie sweetener and dietary maximum levels for sucrose and Stevia in
supplement (Mizutani and Tanaka 2002). syrup are shown in Table 1. The complete
Stevia sweetener is heat stable up to 200 ºC, design consisted of 13 experimental points
acid-stable and not fermentable, making it which included five replications of the
suitable for use in a wide range of products centre point (Table 2).
including baked/cooked foods (Puri et al.
2011) as well as acidified beverages. Stevia Sensory evaluation
can partially replace sucrose in certain types Trained panellists were used for the sensory
of food products including non-carbonated evaluation session. The training session
drinks (Saniah et al. 2009), traditional cakes was started with a screening test which
(Zainun et. al. 2009) and confectionery ttaught the candidates the test process while
jellies (Sharifah Samsiah and Latifah 2009), weeding out unsuitable nondiscriminators.
without significantly affecting the sensory The screening tests aimed to determine
qualities. differences among candidates in the ability
Furthermore, Stevia is non-calorific, to discriminate the different levels of
able to maintain good dental health and intensity for each attribute. Panellists were
suitable for diabetic patients (Geuns carefully introduced to the attributes, the
2003). However, the problem of the bitter terminology used to describe them as well
taste in Stevia is of concern in consumer as the scale method used to indicate or
acceptability (Saniah et al. 2009). Thus, the measure the degree of liking for a product.
aim of the present study is to investigate Acceptance test was used to determine the
the possibility of fully or partially replacing ‘affective status’ of a product or how well
sucrose content in carbonated drinks with it is liked by the panellists. Panellists were
Stevia for achieving maximum consumer asked to rank the samples in ascending order
acceptance and quality. ascording to the level of sensory attributes
including sweetness, sourness, bitter-after
Materials and methods taste and carbon dioxide taste. Panellists
Sample preparation and experimental who were able to rank the samples correctly
design were accepted for the sensory evaluation
Carbonated drink was developed by
incorporation of flavoured syrup or cordial, Table 1. Factors and levels for response surface
cooled water and carbon dioxide gas using a study
carbonated drink machine. Orange flavoured Factor Low level High level
syrup was first prepared according to (–1) (+1)
Hamzah (1987) with modifications on the
X2 (A) – Sucrose (%) 0% 54%
amount of sucrose and Stevia substitution as
X1 (B) – Stevia (%) 0.2% 0.5%
sweetening agent. Stevia (Sweta brand) was
24K. Saniah and M. Sharifah Samsiah
Table 2. Experimental design for the formulation of flavoured syrup
Std. run no. Coded independent variables Decoded independent variables
X1 X2 Stevia (%) Sucrose (%)
1 –1 –1 0.20 0.00
2 1 –1 0.20 54.00
3 –1 1 0.50 0.00
4 1 1 0.50 54.00
5 –1 0 0.35 0.00
6 1 0 0.35 54.00
7 0 –1 0.20 27.00
8 0 1 0.50 27.00
9 0 0 0.35 27.00
10 0 0 0.35 27.00
11 0 0 0.35 27.00
12 0 0 0.35 27.00
13 0 0 0.35 27.00
the session. The preparation of samples The sensory quality of carbonated
and training procedures were carried out drinks with Stevia was also compared with
according to Meilgaard et al. (1999) with commercial samples. The degree of liking
slight modifications. (DOL) for viscosity, orange taste, sourness,
A total of 13 samples of carbonated sweetness, bitter after-taste, CO2 gas
drinks were subjected to sensory evaluation content as well as overall acceptability were
by 13 trained panellists. A sample of 40 ml evaluated.
was served and coded with three digits
chosen at random. Sensory attributes Proximate analysis
evaluated were the degree of liking (DOL) Crude fibre, sugar and fat contents of the
for taste, texture, bitter after-taste, sourness, samples were determined according to
sweetness and overall acceptability. All AOAC (1985). The Kjeldahl method of
panellists evaluated the samples using a determining total nitrogen was based on
7-point category hedonic scale (1 = dislike Tecator Kjeltec System 1026 and Pearson
extremely; 4 = neither like nor dislike; 7 = (1976). The crude protein was calculated as
like extremely) as described by Meilgaard et N X 6.25. Total carbohydrate was estimated
al. (1999). according to Nergiz and Otles (1993).
Models with a greater coefficient of Energy was calculated using the factors 4.0,
determination (R2) and adjusted coefficient 4.0 and 9.0 kcal/g for protein, carbohydrate
of determination (R2adj) and, consequently, and fat respectively (Abdurahman et al.
greater F values; non-significance (p >0.05) 1998). Ascorbic acid content (mg/g sample)
in lack of fit and good results in the was determined using direct titration method
residual analysis were considered as good according to Suntornsuk et al. (2002).
performances to provide predictive models
for the responses. The Design-Expert 6.0.8 Results and discussion
(DX6) Statistical Program (Stat-Ease Inc. Model fitting from RSM
2000) was used to develop the experimental Using RSM, the relationship among the
plan for RSM. This software was also used variables, i.e. sucrose and Stevia were
for regression analysis of the data obtained, expressed mathematically in the form of
to estimate the coefficients of the regression polynomial model, which gave response
equation and to perform the analysis of as a function of relevant variables. The
variance (ANOVA). independent and dependent variables were
25Stevia in carbonated drinks fitted to the second order model equation evaluation result shows that the models for and examined for the goodness of fit. sweetness, sourness, bitter after-taste and The analysis of variance was performed overall acceptability had coefficients of to test the adequacy of the model. The determination (R2) above 0.8 and 0.9. This parameters found to be significant at the indicated that nearly 80–90% of the total probability level (p) equal or less than 5% variation can be explained or accounted for were considered in order to obtain the fitted by the models. All models also presented models to predict the responses. In this case, a non-significant lack of fitness (p >0.05), the responses for bitter after-taste, sourness, which is desirable for the model as sweetness, carbon dioxide gas content and significant lack-of-fit indicates that there overall acceptability were significant at might be contributions in the regressor- 0.01% level. The evaluation of the effects response relationship that are not accounted on DOL for orange taste and texture for by the model. (viscosity) indicated that a linear term was not significant, where the probability Effect of sucrose and Stevia on level (p) was above 5% (data not shown). carbonated drinks Therefore, these attributes were not adequate Viscosity It can be observed that the and have not been used for the optimization viscosity of carbonated drink depended on study. The coefficient of determination or the amount of sucrose added, as its quadratic R2 suggested that for a good fit model, R2 effect was positive at p
K. Saniah and M. Sharifah Samsiah of sucrose added. Potter (1986) previously Total soluble solid content It was noticed that the sugar in beverages not observed (Figure 1 and Table 4) that the only contributes to sweetness but also adds total soluble solid content of carbonated body and mouthfeel. For this reason, if drink depended on the amount of sucrose Stevia is used as a sweetener, the addition added, as its 2F1 (2 factor interaction) of thickening agent such as carboxymethyl effect was positive at p
Stevia in carbonated drinks
Design-Expert Software Design-Expert Software
Bitter after taste Sweetness
• Design points above predicted values • Design points above predicted values
• Design points below predicted values • Design points below predicted values
5.3 5.5 5.2
6
3.46 3.82
X1 = A : Sucrose 6 X1 = A : Sucrose 5.5
X2 = B : Stevia X2 = B : Stevia
4.5 5
Bitter after taste
4 4.5
Sweetness
3.5 4
3 3.5
0.50 54.00 0.50 54.00
45.00 45.00
0.42 36.00 0.42 36.00
0.35 27.00 0.35 27.00
0.27 18.00 0.27 18.00
9.00 9.00
B : Stevia 0.20 0.00 A : Sucrose B : Stevia 0.20 0.00 A : Sucrose
Figure 2. Response surface plot for bitter Figure 3. Response surface plot for sweetness
after-taste
Design-Expert Software Design-Expert Software
OA Sourness
• Design points above predicted values • Design points above predicted values
• Design points below predicted values • Design points below predicted values
5.6 6 5.6
3.54 4.0 6
X1 = A : Sucrose 5.5 X1 = A : Sucrose
5.5
X2 = B : Stevia 5 X2 = B : Stevia
5
4.5
4.5
4
Sourness
4
OA
3.5
3 3.5
0.50 54.00 0.50 54.00
45.00 45.00
0.42 36.00 0.42 36.00
27.00 0.35 27.00
0.35 18.00 18.00
B : Stevia 0.27 B : Stevia 0.27 9.00 A : Sucrose
9.00 A : Sucrose 0.20 0.00
0.20 0.00
Figure 4. Response surface plot for Figure 5. Response surface plot for sourness
overall acceptability
coefficients of determination (R2) of 0.9085. bitter after-taste in mango drink (Saniah et
The Rebaudioside A component is known al. 2009).
to have bitter taste affecting overall panel Figure 5 shows surface plot generated
sensory score. Preliminary study (data not from predictive equation for degree of
shown) indicated a slight bitter after-taste liking of sourness. It can be observed that
in carbonated drinks when the amount of the sourness was insignificantly depended
Stevia added was more than 0.5%. At this on the concentration of sucrose and Stevia
level, the product was still acceptable but (p >0.05).
with the right combination of sucrose. The The concentration of sucrose and Stevia
different levels of sucrose significantly had positive effect on degree of liking of
influenced (pK. Saniah and M. Sharifah Samsiah
A minimum acceptable sensory score sucrose enhanced the sensory perception
of 5.0 for all attributes was considered by contributing to both taste and texture of
essential to establish the range of Stevia product.
-sucrose levels for carbonated drink
preparation. Generally, the lowest sensory Optimization of basic formulation and
score for overall acceptability (below 4.0) verification of the model
was obtained from 0% level of sucrose Graphical optimization function was used
(Table 3). Data showed that carbonated to superimpose the contour plots generated
drinks containing medium to maximum level from five response variables. Superimposing
of sucrose and Stevia had highest scores of of sensory contour (Figure 7) resulted in
this attribute. It was found that addition of an acceptable beverage of minimum five
sensory scores, where the suitable range of
Design-Expert Software
CO2 content
Stevia and sucrose was shown as feasible
• Design points above predicted values
• Design points below predicted values
region.
5.2
3.13
6 The multiple response optimizations
X1 = A : Sucrose
X2 = B : Stevia
5.5 were also performed using the numerical
5 optimization function. The desired goals
4.5 for the factors and responses are set. In
CO2 content
4 this optimization study, the percentage of
3.5 sucrose and Stevia was set as ‘in range’. It is
desirable to have a maximum sensory score
0.50 54.00
0.42
45.00
36.00
for all responses, particularly the degree of
0.35
0.27
27.00
18.00 liking (DOL) for bitter after-taste, sourness,
B : Stevia 9.00 A : Sucrose
0.20 0.00 sweetness, carbon dioxide content and
Figure 6. Response surface plot for carbon
overall acceptability. Therefore, the goals
dioxide content for the responses under investigation were
Design-Expert Software Overlay Plot
Overlay Plot 0.50
Bitter after taste
Sweetness
Sourness 5
Sourness
CO2 content
OA
• Design Points
X1 = A : Sucrose 0.42 CO2 content 5
X2 = B : Stevia Bitter after taste: 5.19588
Sweetness: 5.5626
Sourness: 5.27403
CO2 content: 5.31055
B : Stevia
Co
CO22content
content
OA: 55 5.59871
0.35 Sourness
X1 = 5A : Sucrose 33.13
X2 = OA
B : 5Stevia 0.43
Bitter
Bitterafter
aftertaste
taste5
OA 5
0.27
Sweetness 5
Sourness 5
0.20 Bitter after taste 5
0.00 9.00 18.00 27.00 36.00 45.00 54.00
A : Sucrose
Figure 7. Superimposed plots of the five response variables of orange
flavoured carbonated drink
29Stevia in carbonated drinks
Table 6. Possible optimal solutions for orange flavoured carbonated drink
Sucrose Stevia Bitter after- Sourness Sweetness CO2 Overall Total soluble Viscosity
taste content acceptability solid
33.13 0.43 6.44 6.15 6.20 6.43 6.87 0.88 8.94
set as ‘maximize’. The possible optimal and acceptable. Only a few samples show an
solutions for the percentage and combination error between 10% and 20%.
of sucrose and Stevia for orange flavoured
syrup are shown in Table 6. Effect of Stevia on consumer preference
From the optimization tool, it is and nutritional value
clear that the best DOL for sensory test is The sensorial acceptability of carbonated
obtainable when 33.13% of sucrose was drink with Stevia as compared with similar
used in combination with 0.43% Stevia. sugar rich commercial carbonated drink was
With this formulation, the amount of evaluated. Generally, panellists gave higher
sucrose can be reduced up to 39%. The score to commercial sample of orange-
incorporation of 33.13% sucrose and 0.43% flavoured carbonated drink particularly
Stevia suggested to obtain the optimum for overall acceptability, carbon dioxide
score of sensory attributes evaluated with content, sweetness, orange taste and
the desirability of 0.859. The optimum viscosity with a sensory score of 5.82, 5.91,
scores for bitter after-taste, sourness, 5.55, 5.73 and 5.82 respectively (Table 8).
sweetness and carbon dioxide content However, carbonated drink with Stevia
were 6.44, 6.15, 6.20 and 6.43 or close gave comparable resuls with no significant
to ‘like slightly’ respectively. The overall difference, except for DOL of carbon
acceptability score was 6.87 or close to ‘like dioxide content and viscosity (Table 8).
moderately’ (Table 6). This might be due to the different carbon
In order to verify the adequacy of dioxide gas flow rate applied by commercial
the model developed, several confirmation companies (usually up to 12 bar) as
runs were performed involving different compared to 4 bar used in this study, since
percentages of sucrose and Stevia within the pressure is needed to force more CO2
the range of the levels defined previously. into solution. The solubility of CO2 in water
Using the point prediction capability of the is greater at a lower temperature of liquid.
software, the DOL for orange taste, texture, A nutritious balanced diet is the
bitter after-taste, sourness, sweetness, carbon key to good health and the changes in
dioxide content and overall acceptability nutritional quality is also associated with
of the selected experiments were predicted greater changes in consumer acceptance.
together with the 95% prediction interval. Table 9 shows the effect of adding Stevia
The predicted and the actual sensory score in carbonated drink on the proximate
were compared and the residual and the composition as compared to control and
percentage error were calculated. These are commercial samples. The higher amount of
presented in Table 7. The calculated results carbohydrate was detected in the control and
indicated that the models developed were commercial samples which were mostly due
highly accurate, particularly for DOL of to addition of sucrose in the formulation.
bitter after-taste, carbon dioxide content and The combination of sucrose-Stevia in
overall acceptability, where the percentage the drink fomulation greatly reduced the
errors for other responses were less then carbohydrate content from 14% in the
10%. The percentage errors for other control sample to 8%. Stevia is considered
sensory attributes were considered as small to be a non-caloric sweetener. In line with
30K. Saniah and M. Sharifah Samsiah
Table 7. Predicted and actual value of sensory scores for confirmation run
Responses Factors Predicted value Actual value Residual % Error
Viscosity S1 4.5 5.2 0.7 13.50%
S2 4.41 4.45 0.04 0.90%
S3 5.19 5.36 0.17 3.17%
Orange taste S1 5.06 5.4 0.34 6.30%
S2 3.54 3 –0.54 –18.00%
S3 5.06 4.82 –0.24 –4.98%
Sourness S1 5.27 5.4 0.13 2.40%
S2 3.88 3.36 –0.52 –15.47%
S3 4.74 5 0.26 5.20%
Sweetness S1 5.56 5.19 –0.37 –7.13%
S2 3.72 3.09 –0.63 –20.42%
S3 4.95 4.27 –0.68 –15.92%
Bitter after-taste S1 5.2 5.3 0.1 1.90%
S2 3.42 3.64 0.22 6.00%
S3 4.88 4.82 -0.06 –1.24%
CO2 content
S1 5.31 5 –0.31 6.20%
S2 3.69 3.82 0.13 3.40%
S3 4.48 4.73 0.25 5.29%
Overall acceptability
S1 5.27 5.4 –0.13 2.41%
S2 3.41 3.09 –0.32 –10.36%
S3 5.27 4.9 –0.37 –7.55%
S1 = 33.16% sucrose, 0.43% Stevia; S2 = 0.00% sucrose, 0.20% Stevia
S3 = 54.00% sucrose, 0.50% Stevia
Table 8. The degree of liking (DOL) of carbonated drink with Stevia vs commercial carbonated drink
Sample Viscosity Orange Sourness Sweetness Bitter CO2 Overall
taste after-taste gas content acceptability
Carbonated drink 5.2 5.4 5.4 5.19 5.3 5 5.4
with Stevia
Commercial 5.82 5.73 5.45 5.55 ND 5.91 5.82
sample
ND = Not detected
Table 9. Nutritional value of carbonated drink with Stevia vs control sample (n = 3)
Sample Protein as Fat Total Energy Dietary fibre Sugar content Vitamin C
N x 6.25 (% w/v) Carbohydrate (kcal/100 ml) (% w/v) (% w/v) (mg/100 g)
(% w/v) (% w/v)
Carbonated drinkStevia in carbonated drinks
that, sucrose-Stevia based carbonated Conclusion
drink provides only 32 kcal/100 ml of Central composite design and response
calorie content, significantly lower than the surface methodology can be used for the
amount detemined in the control sample (56 purpose of optimizing the formulation
kcal/100 ml). As can be observed, generally of Stevia-based carbonated drinks. This
the amount of nutrient components in study has shown that the average panellists
carbonated drink was very low and/or below generally responded with a high level of
human requirements. acceptance for carbonated drinks containing
Results from the nutritional analysis 33.13% sucrose and 0.43% Stevia in syrup
indicated that the replacement of sugar formulation. Results indicated that Stevia
with Stevia provides a healthier choice of can partially replace the amount of sucrose
carbonated drink to the global population, without drastically affecting the physical
particularly those concerned about health properties and consumer acceptability.
and nutrition. Related to these findings, Reducing the concentration of sucrose as
substituting sugar with low calorie well as increasing the amount of Stevia
sweeteners may be effective in weight decreased the viscosity and total soluble
management or preventing metabolic solid of the carbonated drink. Furthermore,
disorders such as obesity. Stephen et al. the addition of 0.43% Stevia significantly
(2010), studied the effects of preloads reduced the level of carbohydrates and
containing Stevia, aspartame and sucrose on calories by 42.9% in the carbonated drink.
food intake, satiety, and postprandial glucose
and insulin levels. He observed that Stevia References
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220–222
33Stevia in carbonated drinks Abstrak Kaedah gerak balas permukaan dan reka bentuk komposit berpusat telah digunakan dalam penentuan tahap optimum pemanis Stevia sebagai pengganti gula di dalam minuman berkarbonat. Dalam kajian ini kesan penggunaan kombinasi Stevia – sukrosa dengan julat 0.2 – 0.5% (Stevia) dan 0–54% (sukrosa) terhadap tahap penerimaan nilai rasa dan profil fizikal dan kimia bagi produk telah dikaji. Berdasarkan keputusan gerak balas permukaan dan pertindanan rajah, kualiti yang dikehendaki bagi minuman berkarbonat berperisa oren dapat diperoleh dengan menggabungkan 0.43% Stevia dan 33.13% sukrosa di dalam sirap. Keputusan menunjukkan nilai kalori serta karbohidrat menurun sebanyak 42.9%. Ini menggambarkan Stevia mempunyai potensi yang baik untuk diekploitasi sebagai pemanis alternatif bagi minuman berkarbonat. Accepted for publication on 3 April 2012 34
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