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Optimization of Hydrocolloid Extraction From Wild Sage Seed (Salvia macrosiphon) Using Response Surface

Aram Bostana; Seyed M. A. Razavia; Reza Farhoosha a Department of Food Science and Technology, Ferdowsi University of Mashhad (FUM), Khorasan, Iran Accepted uncorrected manuscript posted online: 29 June 2010 Online publication date: 29 June 2010

To cite this Article Bostan, Aram , Razavi, Seyed M. A. and Farhoosh, Reza(2010) 'Optimization of Hydrocolloid Extraction

From Wild Sage Seed (Salvia macrosiphon) Using Response Surface', International Journal of Food Properties, 13: 6, 1380 1392, doi: 10.1080/10942910903079242, First posted on: 29 June 2010 (iFirst) To link to this Article: DOI: 10.1080/10942910903079242 URL: http://dx.doi.org/10.1080/10942910903079242

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International Journal of Food Properties, 13:13801392, 2010 Copyright Taylor & Francis Group, LLC ISSN: 1094-2912 print / 1532-2386 online DOI: 10.1080/10942910903079242

OPTIMIZATION OF HYDROCOLLOID EXTRACTION FROM WILD SAGE SEED (SALVIA MACROSIPHON) USING RESPONSE SURFACE Aram Bostan, Seyed M.A. Razavi, and Reza Farhoosh
Department of Food Science and Technology, Ferdowsi University of Mashhad (FUM), Khorasan, Iran
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The effect of temperature (2580 C), water to seed ratio (25:185:1) and pH (39) on the yield, apparent viscosity and emulsion stability index of wild sage seed hydrocolloid was investigated. The generated quadratic model showed that the optimum conditions for maximizing the responses were when temperature was 25 C, water to seed ratio was 51:1 and pH was 5.5. All hydrocolloid solutions (1% w/v) showed shear thinning behavior in different extraction conditions, consistency coefcient and ow behavior index varied from 4.455 to 9.435 (Pa.sn ), and 0.317 to 0.374, respectively. Besides, the chemical compositions of the seed and extracted gum were determined at optimal conditions. Keywords: Wild sage seed, Hydrocolloid, Extraction, Optimization, Response Surface Methodology (RSM).

INTRODUCTION The food industry has seen a large increase in use of hydrocolloids in recent years. According to the safety, availability and low process costs, plant seeds have a good potential as new sources of hydrocolloids. Most seeds contain starches as the principal reserve food stored for use by the embryonic plant, but many seeds contain other polysaccharide polymers with gum-like functional properties which have served as a useful source of commercial hydrocolloids.[1] The genus Salvia (Labiatae) contains more than 700 species, which about 200 out of them exist in Iran and is probably found in neighboring countries. Plants belonging to this genus are pharmacologically active and have been used in folk medicine all around the world. Wild sage seed (Salvia macrosiphon) is a small, rounded seed, which readily swells in water to give mucilage,[2] but very few formal studies have looked at this little seed, only the composition of essential oil of this species has been reported by Matloubi-Moghaddam et al.[3] and recently computer image analysis and physic-mechanical properties of the seed investigated by Razavi et al.[4] Food hydrocolloids are used for thickening, gelling, lm forming and stabilizing purposes. Many food products such as sauces, syrups, ice cream, instant foods, beverages
Received 10 January 2009; accepted 31 May 2009. Address correspondence to Seyed M.A. Razavi, Department of Food Science and Technology, Ferdowsi University of Mashhad (FUM), Khorasan, P.O. Box 91775 1163, Iran. E-mail: s.razavi@um.ac.ir

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and confectionaries, marshmallows, and candies contain hydrocolloids in their formulations. The common property of hydrocolloids is that they impart viscosity or thickening to the aqueous solutions. The rheological behavior of hydrocolloids governs the quality of the end product, as well as the design and evaluation of process equipments.[5] Previous studies showed that the degree of thickening can be inuenced by the extraction conditions.[611] Hydrocolloids are also added to control the stability of different emulsions such as salad dressings where the effect of extraction conditions on emulsion stability is important to be considered.[12] Preliminary tests showed that extraction temperature, pH and water to seed ratio have signicant inuence on the yield, apparent viscosity and emulsion stability index of wild sage seed hydrocolloids. Thus, it is essential to optimize the extraction process in order to obtain the highest yield and quality of this hydrocolloid. The general practice of determining these optima is by varying one parameter while keeping the other at an unspecied constant level. The major disadvantage of this single variable optimization is that it does not include interactive effects among the variables; thus, it does not depict the net effects of various parameters on the reaction rate. In order to overcome this problem, when many factors and interactions affect desired variables, response surface methodology (RSM) is an effective tool for optimizing the process.[7] RSM is an effective statistical method that uses a minimum of resources and quantitative data from an appropriate experimental design to determine and simultaneously solve a multivariate equation.[13] Response surface experiments attempt to identify the response that can be thought of as a surface over the explanatory variables_ experimental space. It usually uses an experimental design such as central-composite experimental design (CCED) to t an empirical, full second-order polynomial model. A central-composite experimental design, coupled with a full polynomial model, is a very powerful combination that usually provides an adequate representation of most continuous response surfaces over a relatively broad factor domain.[14] Many researchers have used RSM to optimize hydrocolloid extraction.[611] Negligible information is available so far on the extraction and functional properties of wild sage seed hydrocolloid and this paper deals with optimum extraction conditions of wild sage seed hydrocolloid as a novel gum source and some chemical and rheological characterization. MATERIAL AND METHODS Sample Preparation The wild sage seeds used in this study were obtained from a local market in Mashhad, Iran. The seeds were cleaned manually to remove all foreign matter such as dust, dirt, stones and chaff. All chemicals used were reagent grade unless otherwise specied. Gum Extraction Wild sage seed gum was extracted from whole seeds using distilled water (water to seed ratio of 25:185:1) at pH 39. The pH was monitored continuously and adjusted by 0.1 mol/L NaOH and HCl, respectively, while the temperature of the aqueous system ranged from 2580 C and was controlled within 2.0 C using an adjustable temperature controlled water bath. Water was preheated to a designated temperature before the seeds were added. Extraction was carried out in three stages; in the rst stage, the seeds (40 g)

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were mixed with 1000 ml water (25:1 W:S) at a specic pH and temperature and enough time (20 min) was given that complete water absorption was occurred. A soaking time of 20 min was selected based on the yield of preliminary trials. Separation of the gum from the swelled seeds was done by passing the seeds through a laboratory extractor (Model 412, Pars Khazar Com., Iran). Crude gum was collected and residual seeds immersed in remaining of water in two stages, according to water to seed ratio proposed for each run, and again was passed through the extractor. The collected crude gum from the different stages was mixed, ltered and dried overnight in a forced convection oven (Model 4567, Kimya Pars Com., Iran) at 70 C.[15] The dried gum was then grounded, ltered and used for analysis.

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Experimental Design The optimization method based on RSM involved three major steps: design of experiment using statistical approach, coefcient estimation based on mathematical model and response prediction and nally model adequacy check. The models were tested with analysis of variance (ANOVA) with 95% degree of condence. The RSM outputs such as contour and 3D graphic surface plots provide the optimum and most inuential variables for hydrocolloid extraction. In this paper, a central-composite experimental design, with three variables, was used to study the response pattern and to determine the optimum combination of variables. The effect of the independent variables x1 (temperature, T), x2 (water to seed ratio, W:S) and x3 (pH), at ve variation levels on the responses is shown in Table 1.
Table 1 The central composite experimental design and results for yield, apparent viscosity and ESI of crude hydrocolloid extract of wild sage seed. Factors Run 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Temperature ( C) 52.50 (0) 69.00 (+1) 52.50 (0) 36.00 (1) 52.50 (0) 52.50 (0) 69.00 (+1) 36.00 (1) 52.50 (0) 52.50 (0) 52.50 (0) 36.00 (1) 52.50 (0) 36.00 (1) 52.50 (0) 69.00 (+1) 52.50 (0) 69.00 (+1) 24.75 (1.68) 80.25 (+1.68) W:S 54.95 (0) 37.10 (1) 54.95 (0) 37.10 (1) 54.95 (0) 54.95 (0) 72.80 (+1) 72.80 (+1) 54.95 (0) 54.95 (0) 24.93 (1.68) 37.10 (1) 54.95 (0) 72.85 (+1) 54.95 (0) 72.80 (+1) 84.97 (+1.68) 37.10 (1) 54.95 (0) 54.95 (0) pH 6.00 (0) 4.22 (1) 8.99 (+1.68) 7.78 (+1) 6.00 (0) 6.00 (0) 7.78 (+1) 4.22 (1) 6.00 (0) 6.00 (0) 6.00 (0) 4.22 (1) 6.00 (0) 7.78 (+1) 3.01 (1.68) 4.22 (1) 6.00 (0) 7.78 (+1) 6.00 (0) 6.00 (0) Yield (%) 10.45 9.35 9.74 8.9 10.12 10.45 12.2 10.3 10.4 10.35 7.04 8.5 9.95 10 10.3 12.9 10.93 9.9 9.8 13.69 Responses Viscosity1 (mPa.s) 290 207 264 355 293 290 227 273 300 298 311 277 299 198 230 234 198 331 301 274 ESI (min) 310 213 252 188 316 360 298 371 300 300 239 393 340 308 226 172 174 255 450 220

1 Apparent viscosity was determined at 25 C and shear rate 122 s1

for 1% (w/v) hydrocolloid solution.

HYDROCOLLOID EXTRACTION FROM WILD SAGE SEED

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Six replicates at the centre of the design were used to estimate a pure error sum of squares. The responses functions (y) measured were yield, apparent viscosity and emulsion stability index (ESI). Different models were tted to the responses and their adequacy checked, nally the best model was selected and related coefcients were reported. These values were related to the coded variables (xi , i = 1, 2, and 3) by a second degree polynomial using the equation below:
2 2 2 y = 0 + 1 x1 + 2 x2 + 3 x3 + 11 x1 + 22 x1 + 33 x3 + 12 x1 x2 + 13 x1 x3 + 23 x2 x3 . (1)

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The coefcients of the polynomial were represented by 0 (constant term), 1 , 2 and 3 (linear effects), 11 , 22 and 33 (quadratic effects), and 12 , 13 and 23 (interaction effects). The analysis of variance (ANOVA) tables were generated and the effect and regression coefcients of individual linear, quadratic and interaction terms were determined. The signicances of all terms in the polynomial were judged statistically by computing the F-value at a probability (p) level of 0.05. The regression coefcients were then used to make statistical calculation to generate contour maps from the regression models. The data were analyzed using the Design-Expert Software (Version 6.0.2 , 2000, Stat-Ease, Inc., UK). Flow Behavior The ow behavior of crude hydrocolloid extract of sage seed was determined using a rotational viscometer (Bohlin Model Visco 88, Bohlin Instruments, UK) equipped with a heating circulator (Julabo, Model F12-MC, Julabo Labortechnik, Germany). Bob and cup measuring spindle (C30) was used during measurements according to the viscosity of dispersion. Prepared samples (1% w/v) were loaded into the cup and allowed to equilibrate for 10 min at desired temperature (25 C) and were then subjected to a programmed logarithmic shear rate ramp increasing from 14 to 300 s1 during 3 min. The shear stressshear rate data of wild sage seed gums was tested for power law model as follow (16): = k n , (2)

where, is the shear stress (Pa); is the shear rate (s1 ); k is the consistency coefcient (Pa.sn ); and n is the ow behavior index (dimensionless).

Evaluation of Extracted Gum Extraction yield. The yield of the extracted gum for various extraction conditions was determined by weighting the dried extracted gums and calculating the percentage based on the weight of the seeds.[17] Apparent viscosity. For Non-Newtonian uids, viscosity depends on the shear rate, therefore the apparent viscosity (a ) of crude hydrocolloid extract was calculated by viscometer software at a given shear rate (122s-1) and 25 C for 1% (w/v) solutions.

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Emulsion stability index (ESI). Oil in water emulsions were prepared by mixing 20% canola oil with an aqueous phase containing 0.5% wild sage seed hydrocolloid. The hydrocolloid powder was dissolved in distilled water by stirring at room temperature and kept in a refrigerator overnight to ensure complete hydration. Emulsion was prepared by homogenizing oil and aqueous phase using a high speed blender (Sanyo, Japan). The ESI for the emulsions were determined by the turbidimetric methods.[18] Freshly prepared emulsions (1 ml) were pipetted out at 0 and 10 min after homogenization and diluted with 99 ml SDS (1 g.kg1 ). Absorbance of the nal dispersion was measured at 500 nm (Spectrophotometer, UV-160A, Shimadzu, Japan). The ESI (min) was determined as follows: ESI =
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A0 t. A

(3)

where A0 is the absorbance of the diluted emulsion immediately after homogenization; A is the change in absorbance between 0 and 10 min (A0 _A10 ); and t is the time interval, 10 min in this case. Analytical Methods To analyze the chemical composition of the wild sage seeds and that of the extracted gum at optimum condition, moisture content of the seed and the extracted gum at optimum condition was determined by the vacuum oven method (temperature 70 C and pressure 250 mbar) until a constant mass was obtained,[17] total nitrogen and ash contents were determined in duplicate according to AOAC methods (19): 2.0612.062 (modied Kjeldahl method) and 923.03 (direct method of ash determination in ours), respectively. The Kjeldahl factor used was 6.25. Fat was extracted by a semi-continuous procedure using a Soxhlet device, and diethyl ether and hexane were used as the extraction solvent. Crude ber (i.e., cellulose, lignin, and part of the total hemi-cellulose) was also determined according to AOAC method No. 920.86. Total carbohydrates were determined as the difference between 100 and the sum of the other components. All measurements were performed at least in triplicate and results expressed as mean SD. RESULTS Flow Behavior The dispersions of the hydrocolloid from wild sage seeds showed non-Newtonian pseudoplastic behavior over the entire extraction conditions. Based on the values obtained for R2 , the power law model was well tted to experimental ow curves (Table 2). Both parameters of the power law model, k and n are shown in Table 2 along with the apparent viscosity values at 46.16 s1 . Values of the ow behavior index (n) were below unity conrming a shear-thinning behavior of the wild sage seed extracts at all extraction conditions tested. The ow behavior ranged from 0.317 to 0.374 with the mean value as 0.347, which showed the high shear thinning tendency of the hydrocolloid solutions. The extraction conditions had no signicant effect on the ow behavior index. The values of consistency index, k, ranged from 4.455 to 9.435 Pa sn , while mean value of k was 6.355 Pa sn . The extraction conditions signicantly affected the consistency index, increasing pH and descreasing temperature and water to seed ratio had a negative impact on parameter k.

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As the polysaccharide polymers enhance the thickening properties of the solution, probably this is due to impurities (non polysaccharide compounds) which were probably extracted at higher temperatures and water to seed ratios. Marcotte et al. (16) also reported that an increase in temperature will decrease the consistency coefcient of the hydrocolloids. The apparent viscosity values of sage seed hydrocolloid ranged from 373 to 694 mPa.s at different extraction conditions, while Cui et al. (10) reported the range of 16.74 to 148.50 for axseed gum at the same concentration (1% w/w) and shear rate (46.16 s1 ). Statistical Analysis and the Model Fitting The experimental data for yield, apparent viscosity and emulsion stability index of the extracted hydrocolloid under different treatment conditions are presented in Table 1. The analysis of variance of different models showed that adding terms up to quadratic will signicantly improved the model (Table 3), therefore a quadratic model is the most appropriate model for the three responses. Model validating parameters and the coefcients of equation terms, which is an empirical relationship between response and the test variables in the regression models after implementing the stepwise ANOVA for response variables, are presented in Table 4. The statistical analysis indicated that the proposed regression model for yield, apparent viscosity and ESI was adequate, possessing no signicant lack of t and with satisfactory values of the R2 (coefcient of determination) for all the responses. The R2 values were 0.982, 0.989 and 0.892 for yield, apparent viscosity and ESI, respectively (Table 4). The closer the value of R2 to the unity, the better the empirical model ts the actual data.[6] Furthermore, the predicted-R2 is in reasonable agreement with adjusted-R2 for all three responses. Inuence of Variables on Yield The effect of different extraction conditions on hydrocolloid yield has been reported by the coefcient of the second order polynomials (Table 4) and to aid visualization, the response surfaces for yield are shown in Fig. 1. As shown in Table 4, all variables except linear and quadratic terms of pH and interaction between temperature and pH had significant effect on yield (P < 0.05). The variables with the largest effect on yield were the linear terms of water: seed ratio and temperature, respectively. Higher temperature and water to seed ratio resulted in higher yield due to enhanced mass transfer rate (Table 1 and Fig. 1). Similar results were obtained by Wu et al.,[6] Koocheki et al.,[8] Razavi et al.,[9] Sepulveda et al.,[15] Cui et al.,[10] and Singthong et al.[11] Li et al.[20] reported that the effect of extraction temperature was substantial, but the water to solid ratio had a slight effect on extraction yield. Also, the effect of pH on yield was not signicant, Koocheki et al.[8] and Cui et al.[10] also reported the minor effect of pH on extraction yield, but Razavi et al.,[9] Furuta et al.,[21] and Somboonpanyakul et al.[22] reported that pH has a signicant effect on the yield. In constant pH (pH = 6.0), increasing water to seed ratio increased the yield. This effect was more pronounced at higher temperatures (Fig. 1a). Increased in water to seed ratio in all pH increased the yield except the alkaline pH (Fig. 1c). Figure 1b shows the changes of hydrocolloid yield with temperature and pH at a constant water to seed ratio (W:S = 51:1). It was clear that increasing temperature increased the hydrocolloid yield, but pH had no signicant effect on yield, however, at high W:S, increasing the pH decreased the yield (Fig. 1c). In this study, the lowest yield of gum (7.04%) was obtained at lowest water to seed ratio (24.93:1) at temperature of 52.5 C and pH of 6.00,

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Table 4 ANOVA results showing the variables as a linear, quadratic and interaction terms on each response variables and coefcients for the prediction models. Yield SS 39.029 294.95 12.74 17.12 0.072 3.86 3.03 0.12 0.0027 0.7449 0.0261 0.1906 279.87 104.00 36509.00 0.9895 0.9800 0.9351 2.27 2370.45 10.50 14.25 35.50 882.00 1624.5 10082.00 < 0.0001 < 0.0001 0.2134 75.89 2809.32 3977.73 2.29 13.96 16.61 0.1900 < 0.0001 < 0.0001 0.0007 < 0.0001 < 0.0001 0.1506 < 0.0001 < 0.0001 0.3337 1634.55 13416.01 2298.71 10.94 31.34 12.97 < 0.0001 < 0.0001 < 0.0001 51.90 0.68 4.12 8.81 36.63 25.14 12.00 54.50 28.25 320.41 36788.48 6.36 231.88 1117.51 19331.93 9104.76 0.0002 0.9428 0.6661 0.3521 0.0023 0.0193 0.3451 0.0011 0.0418 0.1244 1152.00 23762.00 6384.50 11731.24 8821.24 2910.00 1.92E005 0.8925 0.7958 0.3421 12.05 71808.83 < 0.0001 36125.13 < 0.0001 P value Coefcient SS P value Coefcient Apparent viscosity ESI SS 97420.51 P value 0.0009

Source

DF

Coefcient

10.29

1 1 1

0.97 1.12 0.073

1 1 1 1.09 0.008 0.48 0.70 0.40 0.21

0.52 0.46 0.092

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Model 0 Linear 1 2 3 Quadratic 11 22 33 Interaction 12 13 23 Residual Lack of t Pure error Total R2 Adjusted R2 Predicted R2 CV Press

1 1 1 10 5 5 19

0.37 0.031 0.24

0.9825 0.9668 0.8984 2.57 4.06

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(a)
15 13 Yield (%w/w) 9 7 Yield (%w/w) 55.00 B: W/S Ratio 40.00
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(b)
14 12.75 11.5 10.25 9

11

85.00 70.00 80.00 66.25 52.50 38.75 A: Temperature

9.00 80.00 7.50 66.25 6.00 52.50 C: pH 4.50 38.75 A: Temperature 3.00 25.00

25.00 25.00

(c)
11.5 10.125 Yield (%w/w) 8.75 7.375 6

9.00 7.50 C: pH 6.00 4.50

85.00 70.00 55.00 40.00 B: W/S Ratio 3.00 25.00

Figure 1 Response surface for the effect of temperature, water to seed ratio and pH on yield of wild sage seed hydrocolloid; (a) T and W:S at pH = 6.0; (b) T and pH at W:S = 51:1; (c) W:S and pH at T = 55 C.

while the highest value (12.2%) was obtained at the highest temperature (80 C), water to seed ratio of 55:1 and pH of 6.00 (Table 1). Inuence of Variables on Apparent Viscosity The result of analysis of variance and response surface plots showed that the apparent viscosity of hydrocolloid solution was signicantly (P < 0.05) affected by the linear, quadratic and the cross terms between all variables (Table 3), except the quadratic effect of temperature that was no signicant. Based on the sum of squares, water to seed ratio and interaction of water to seed ratio and pH had the largest effect on apparent viscosity (Table 3). Figure 2a reveals that at low water to seed ratio, increasing the temperature decreased the apparent viscosity, whereas at higher W:S, temperature had greater effect. At low pH (37), increasing temperature decreased the apparent viscosity afterwards the apparent viscosity became constant (Fig. 2b). Increasing water to seed ratio at acidic conditions increased the apparent viscosity, while at alkaline conditions it caused reduction in apparent viscosity (Fig. 2c). Koocheki et al. (8), Razavi et al. (9) and Ibanez and Ferrero (23) have reported increasing of pH led to decrease in apparent viscosity. Apparent viscosity varied between 198 to 355 mPa.s in different extraction conditions, the lowest apparent viscosity observed when W:S, T and pH were 72.8, 36 and

HYDROCOLLOID EXTRACTION FROM WILD SAGE SEED

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(a)
Apparent Viscosity(mPa.s)

(b)
350 307.5 265 222.5 180

Apparent Viscosity(mPa.s)

310 272.5 235 197.5 160

80.00

66.25 A: Temperature

25.00 40.00 52.50 55.00 38.75 70.00 B: W/S Ratio 25.00 85.00

80.00 66.25 52.50 4.50 6.00 7.50 C: pH 3.00

A: Temperature

38.75 25.00 9.00

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(c)
Apparent Viscosity(mPa.s)

390 311.25 232.5 153.75 75

85.00 70.00 55.00 4.50 6.00 7.50 C: pH 3.00

B: W/S Ratio

40.00 25.00 9.00

Figure 2 Response surface for the effect of temperature, water to seed ratio and pH on apparent viscosity (at 25 C, 1% (w/v) solution and shear rate 122 s1 ) of wild sage seed hydrocolloid. (a) T and W:S at pH = 6.0; (b) T and pH at W:S = 51:1; and (c) W:S and pH at T = 55 C.

7.78, respectively, and the highest obtained when W:S, T and pH were 37.1, 36 and 7.78, respectively (Table 1). It can be concluded that water to seed ratio substantially affected the apparent viscosity. Inuence of Variables on Emulsion Stability Index The results showed that only temperature had signicant linear effect on ESI and linear effect of water to seed ratio and pH were not signicant (Table 4). When the temperature increased (Fig. 3a), the ESI decreased maybe it was due to decreasing in hydrocolloid solution viscosity as temperature increased. In quadratic terms, only pH and W:S had signicant effect and in cross action interaction between pH and temperature, and pH and water to seed ratio had signicant effect on ESI (p < 0.05). Increasing temperature at lower pH (37) decreased ESI, while at higher pH, it had no signicant effect (Fig. 3b). Elliptical contour observed in Fig. 3c, demonstrated the perfect interaction between pH and water to seed ratio. The maximum predicted value indicated by the surface was conned in the smallest ellipse in the contour diagram (6). In this study, ESI varied between 172 and 450 min under different extraction conditions. The lowest ESI observed when T, W:S and

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(a)
435 356 277 198 119

(b)
521 400.75 280.5 160.25 40

ESI

85.00 70.00 55.00 B: W/S Ratio 40.00 25.00

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80.00 66.25 52.50 38.75 A: Temperature 25.00

ESI

9.00 7.50 6.00 C: pH 4.50 3.00 52.50

80.00 66.25

38.75 A: Temperature 25.00

(c)
521 400.75 280.5 160.25 40

ESI

9.00 7.50 6.00 C: pH 4.50 3.00 25.00 40.00 55.00

85.00 70.00

B: W/S Ratio

Figure 3 Response surface for the effect of temperature, water to seed ratio and pH on ESI (20% O/W emulsion) of wild sage seed hydrocolloid. (a) T and W:S at pH = 6.0; (b) T and pH at W:S = 51:1; and (c) W:S and pH at T = 55 C

pH were 69 C, 72.8:1 and 4.22, respectively, while the highest ESI obtained when W:S and pH were 54.95 and 6.00 at the lowest T (24.75 C). Optimization and Verication Optimization of the extraction procedure was based upon the following: higher extraction yield, apparent viscosity and ESI. The suitability of the models for predicting optimum response values was tested under the conditions: extraction temperature 25 C, water to seed ratio 50.95:1 and pH 5.53. This set of conditions was determined to be optimum by the RSM optimization approach and was also used to validate experimentally and predict the values of the responses using the models (Table 5). The experimental and predicted values were found to be not statistically different at 5% level of signicance, indicating that the model was adequate for the extraction process. Most of researchers found higher temperature for optimum extraction,[9,10,15] whereas our study showed that the crude hydrocolloid of sage seed could be extracted in ambient temperature. The yield of crude hydrocolloid of wild sage seed was 9.97%, which was more than values reported at optimum condition for Flaxseed gum[10] and

HYDROCOLLOID EXTRACTION FROM WILD SAGE SEED Table 5 Predicted and experimental values of responses at optimum extraction condition of sage seed hydrocolloid. Experimental value Response variables Yield (%) Apparent viscosity (mPa.s) ESI (min) Predicted value 9.97 316 450 Mean 10.1 312 403 SD 0.05 14.11 27.2

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Table 6 Chemical compositions of the seeds of wild sage and the hydrocolloid powder. Composition (w/w%)
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Seed 5.13 0.002 3.38 0.005 3.848 0.002 4.32 0.005 0.78 0.000 82.54

Hydrocolloid powder 6.72 0.001 0.85 0.0004 8.172 0.008 2.84 0.009 1.67 0.00 79.75

Moisture Lipid Ash Protein Crude ber Carbohydrate

less than Qodumeh seed[8] and Basil seed.[9] The crude hydrocolloid extracted under the optimum conditions was further analyzed for chemical compositions as shown in Table 6. In this research, the crude hydrocolloid powder of sage seed contained 79.75% carbohydrates, 2.84% proteins, 0.85% lipid, 6.72% moisture, 1.67% crude ber, and 8.172% ash (Table 6). CONCLUSION The extraction conditions had signicant effects on the yield, apparent viscosity and ESI of wild sage seed crude hydrocolloid. Increasing the temperature and water to seed ratio increased the yield of extracted hydrocolloid, while increasing the temperature decreased the ESI and also at high water to seed ratio and alkaline conditions increased the apparent viscosity. The inuence of pH on yield and ESI was signicant (P < 0.05). Increasing pH at low water to seed ratio increased the apparent viscosity, whereas it decreased at high water to seed ratio. In addition, increasing pH at low temperature decreased the apparent viscosity, while it increased at high temperature. Optimum conditions (temperature 25 C, water to seed ratio at 51:1 and pH at 5.5) for the extraction procedure of crude hydrocolloid from wild seeds were identied. All hydrocolloid solutions (1% w/v) showed shear thinning behavior under different extraction conditions. The hydrocolloid dried powder contain 6.72% moisture, 0.85% lipid, 8.172% ash, 2.84% protein, 1.67% crude ber, and 79.75% carbohydrate.

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