f o o d a n d b i o p r o d u c t s p r o c e s s i n g 8 6 ( 2 0 0 8 ) 227–231

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Response surface optimization of wheat germ oil yieldby supercritical carbon dioxide extraction

Ping Shaoa,∗, Peilong Suna, Yanjie Yingb

a College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310032, Chinab Faculty of Electronic and Information Engineering, Zhejiang Wanli University, Ningbo 315100, China

a r t i c l e i n f o

Article history:

Received 25 December 2006

Accepted 18 April 2007

Keywords:

Supercritical carbon dioxide

Wheat germ oil

Response surface methodology

Extraction

a b s t r a c t

The supercritical fluid extraction (SFE) of wheat germ oil was studied. Response surface

methodology (RSM) was used to optimize the parameters of the supercritical carbon dioxide

extraction. Independent variables were operating temperature (40, 50 and 60 ◦C), pressure

(20, 27.5 and 35 MPa) and flow rate (15, 20 and 25 L/h). The response and variables were fitted

well to each other by multiple regressions. All the independent parameters and quadratic of

temperature and pressure affected the oil yield significantly. The maximum wheat germ oil

yield to be about 10.15% by SFE were obtained when SFE was carried out at 35 MPa of pressure,

50 ◦C of temperature, 22.5–25 L/h of solvent flow rate and 1 h of extraction time. The humidity

of wheat germ influenced negatively the extraction process. A comparison between the

relative qualities by SFE and by organic solvent extraction using hexane was made. The

quality of wheat germ oil extracted by SFE was similar to that of oil extracted by hexane.

The experimental results indicated that SFE technique reduced solvent consumption and

extraction time with no adverse effect on the extraction yield and fatty acid composition of

the oil.

© 2008 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

1

Sns(nevt2pus

sgbp

aggregation (Malecka, 2002). Although SC-CO2 extraction of

0d

. Introduction

upercritical carbon dioxide (SC-CO2) extraction is an alter-ative to supplement or to substitute the conventionaleparation systems (distillation or liquid solvent extraction)Lang and Wai, 2001). SC-CO2 offers the advantages of usingon-toxic, non-explosive and cost effective solvent. It enablesxtraction at low temperatures and complete removal of sol-ent from the final product based on the use of fluids atemperatures and pressures above the critical values (Hauthal,001). These conditions make the supercritical fluid presenteculiar physicochemical properties between the gas and liq-id states, which offer them with exceptional characteristicsolvent.

Several researchers have studied SC-CO2 extraction ofeed oil from a wide range of seed species such as cornerm (List et al., 1984a,b), cottonseed (List et al., 1984a,b),

orage seed (Molero Gomez and Martinez de la Ossa, 2002),alm kernel meal (Nik Norulaini et al., 2004) and wheat germ

∗ Corresponding author.E-mail address: pingshao325@tom.com (P. Shao).

960-3085/$ – see front matter © 2008 The Institution of Chemical Engioi:10.1016/j.fbp.2007.04.001

(Reverchon and Marrone, 2001). Wheat germ is a by-productof the wheat milling industry. Germ constitutes about 2–3%of the wheat grain and can be separated in a fairly pureform from the grain during the milling process (Krings andBerger, 2001). Wheat germ contains about 11% oil (Dunfordand Zhang, 2003). Wheat germ oil is used in products suchas foods, biological insect control agents, pharmaceuticalsand cosmetic formulations. Wheat germ processing presentschallenges due to its high content of polyunsaturated fattyacids and bioactive compounds. These compounds are proneto oxidation and degradation under the conditions usedfor conventional edible oil extraction and refining meth-ods (Krings et al., 2000). Resent studies have demonstratedthat wheat germ oil has several important physiologicaleffects which include the ability to lower plasma choles-terol, to reduce cholesterol absorption and to inhibit platelet

lipids has been extensively studied in the laboratory, muchless studies have been reported on total oil yield of wheat

neers. Published by Elsevier B.V. All rights reserved.

r o c e s s i n g 8 6 ( 2 0 0 8 ) 227–231

Table 1 – Independent variables and their levels forcentral composite design

Independent variables Variable levels

−1 0 +1

Temperature, X1 (◦C) 40 50 60Pressure, X2 (MPa) 20 27.5 35

228 f o o d a n d b i o p r o d u c t s p

germ in SC-CO2 by response surface methodology to ourknowledge.

Knowledge of the total oil yield in SC-CO2 is useful for thedesign and development of a process. The classical methodof a single dimensional search involves changing one vari-able while fixing the others at a certain level is laborious andtime consuming especially. Response surface methodologyhas increasing been used for optimizing purpose due to itsefficiency and less data requirement (Osbome and Armacost,1996). Thus, the main objective of the study was to determineand explain the effects of pressure, temperature, flow rate andtime on the yield of oil as well as develop a model equationthat will predict and determine the optimum conditions fortotal oil yield.

2. Experimental

2.1. Materials and reagents

The raw material used for the processes was wheat germ sup-plied by Daping Oil and Fats Company (Anhui, China). Thewheat germ was crushed to particle size of 0.75 mm and thenexposed to microwave heating for 3 min. The temperature ofthe heated bran reached 100 ◦C and was removed from theoven, cooled to 25 ◦C, and stored at 4 ◦C for the stabilization.The carbon dioxide used in SFE was 99.5% (w/w) pure. As aconventional extraction solvent n-hexane (99%) was used.

2.2. Supercritical CO2 extraction

Supercritical CO2 extraction was carried out using HA121-50 extraction system (Nantong, Jiangsu, China). Thermostaticbaths were switched on to reach the operating temperaturerequired for extraction. Gas CO2 was introduced into a com-pressor. The extraction vessel was 1000 ml volume capable ofoperating up to 50 MPa and 75 ◦C with the circulation of heatedwater. The independent variables were temperature (40, 50and 60 ◦C), pressure (20, 27.5 and 35 MPa) and flow rate (15, 20and 25 L/h). After 200 g sample was placed in extraction vessel,the extraction temperature, pressure and flow rate were con-trolled automatically and maintained for 60 min. When thedesired pressure, temperature and flow rate were reached, theextraction was started. The oil dissolved in the supercriticalCO2 was separated from the carbon dioxide and collected inthe separator. The oil yield was determined gravimetrically asdisplayed in

total oil yield (%) =(

extracts (g)feed materials (g)

)× 100 (1)

Conventional extraction was carried out using hexane in aSoxhlet apparatus for 20 h (with a fraction wheat germ size of0.75 mm and humidity less than 0.35%) to guarantee the max-imum extraction efficiency. These values are considered veryimportant to establish an indisputable basis for comparisonto the high-pressure process.

2.3. Experimental design

A central composite design was employed to study the

response, namely wheat germ oil yield. The independent vari-ables were X1, X2 and X3 representing temperature, pressureand flow rate, respectively. The settings for the independent

Flow rate, X3 (L/h) 15 20 25

variables were as follows (low and high values): temperatureof 40 and 60 ◦C; pressure of 20 and 35 MPa; flow rate of 15and 25 L/h. Each variable to be optimized was coded at threelevels: −1, 0 and +1. Three replicates at the centre (0, 0, 0)of the design were performed to allow the estimation of thepure error. The central composite design is shown in Table 1.All experiments were carried out in a randomized order tominimize the effect of unexpected variability in the observedresponse due to extraneous factors.

As for the optimization for wheat germ oil yield, theresponses were analyzed using Matlab 6.5 software. Aquadratic polynomial regression model was assumed for pre-dicting responses. The model proposed for each response of Ywas

Y = A0 + A1X1 + A2X2 + A3X3 + A12X1X2 + A13X1X3

+A23X2X3 + A11X21 + A22X2

2A33X23 (2)

where A0 is a constant; A1, A2, and A3 are linear coefficients;A12, A13, and A23 are cross-product coefficients; and A11, A22

and A33 are quadratic coefficients.The goodness of fit of the model was evaluated by the

coefficient of determination R2 and the analysis of variance(ANOVA). Quadratic polynomial equations were obtained byholding one of the independent variables at a constant valueand changing the level of the other variables.

2.4. Gas chromatography–mass spectrometry analysis

Wheat germ oil after methyl esterification was dissolvedin n-hexane and fatty acid composition was determined bythe following procedure (AOAC, 1995). A Shimada QP2010gas chromatograph with a mass spectrometer (GC–MS),detector model (electron impact, 70 eV) and a MXT-5(30 m × 0.25 �m × 0.25 mm) capillary column was used. Thedetector and injection temperature were carried out at 250 ◦C.Helium was the carrier gas. The oven temperature was pro-grammed from 180 ◦C for 3 min, and then from 180 to 280 ◦C at20 ◦C min−1. The final temperature was maintained for 10 min.Spectra of the compounds were obtained and compared withthose in the US. National Institute of Standards and Technol-ogy (NIST) library. The weight compositions of the oil werecomputed from the GC peak areas without using any correc-tion factors.

3. Results and discussion

3.1. Model fitting

In order to develop response surface equation to predict the

percentage of oil yield in the range of studies conducted, theexperimentally determined oil yield percentages were fitted toEq. (1) and the parameters of the equation were evaluated. The

f o o d a n d b i o p r o d u c t s p r o c e s s i n g 8 6 ( 2 0 0 8 ) 227–231 229

Table 2 – Central composite design and experiment data

Run Independent variables Responses

Temperature, X1 (◦C) Pressure, X2 (MPa) Flow rate, X3 (L/h) Experimental (%) Predicted (%)

1 0 −1 −1 7.13 7.222 0 −1 1 8.20 8.103 0 1 −1 9.94 10.044 0 1 1 10.20 10.115 −1 0 −1 7.18 7.156 −1 0 1 7.38 7.547 1 0 −1 7.87 7.718 1 0 1 8.24 8.279 −1 −1 0 6.83 6.78

10 −1 1 0 9.05 8.9811 1 −1 0 7.14 7.2112 1 1 0 9.79 9.8513 0 0 0 8.74 8.6414 0 0 0 8.66 8.6415 0 0 0 8.52 8.64

yauispi

wowpqs

Y

Tipttat

ields of wheat germ oil obtained for each of the experimentsre listed in Table 2 along with the predicted total oil yield val-es. In the experiment, we also found that the extraction yield

s higher within 30 min of extraction for all the experimentstudied. This may be due to the fact that the extractible com-onents were easily accessible to the solvent. The extraction

n this period was fast.Table 2 shows the responses. The response and variables

ere fitted to each other by multiple regressions. A good fit wasbtained. The regression coefficients of the response functionith statistical analysis are given in Table 3. All the inde-endent parameters (temperature, pressure and feed flow),uadratic of temperature and pressure affected the oil yieldignificantly:

=

8.64000 + 0.32500X1 + 1.21000X2 + 0.23750X3

+0.10750X1X2 + 0.04250X1X3 − 0.20250X2X3

− 0.81875X21 + 0.38125X2

2 − 0.15375X23

100(3)

he value for R2 and small P (probability) (P < 0.05 when signif-cant) were 0.9783 and 0.0001, respectively (Table 4). Also, theredicted results, according to models for oil yield, were closeo the observed experimental responses. These indicated thathe generated models adequately explained the data variationnd significantly represented the actual relationships between

he reaction parameters.

Table 3 – Estimated coefficients of the second-orderregression model for oil yield

Coefficient Values Sum of squares P value

A1 0.32500 0.84500 0.002267A2 1.21000 11.71280 0.00100A3 0.23750 0.45125 0.008597A12 0.10750 0.046225 0.237888A13 0.04250 0.007225 0.618921A23 −0.20250 0.164025 0.052893A11 −0.81875 2.475144 0.000188A22 0.38125 0.536683 0.006023A33 −0.15375 0.087283 0.124915

3.2. Effects of parameters

Many parameters can influence the separation performanceof wheat germ oil extraction. Eq. (2) shows that wheat germoil yield has a complex relationship with independent vari-ables that encompass both first and second-order polynomialsand may have more than one maximum point. The best wayof expressing the effect of any parameter on the yield withinthe experimental space under investigated was to generateresponse surface plots of the equation. The three-dimensionalresponse surfaces were plotted in Figs. 1 and 2 as a functionof the interactions of any two of the variables by holding theother one at middle value. Both plots in Figs. 1 and 2 show therelationships with respect to the effects of each variable.

Contour plot and response surface curve showing predictedresponse surface of oil yield as a function of pressure and flowrate was shown in Fig. 1. It showed that at temperature 50 ◦Cthe oil yield of wheat germ increased with increase in pres-sure. The oil yield increased from about 7.13% to 10.00% asthe pressure was increased from 20 to 35 MPa. These resultsare consistent with presented in other study about the wheatgerm oil extraction (Wang and Xue, 2003; Chen et al., 2002). Theoptimum pressure for the maximum yield of oil was around35 MPa. At lower pressure, the solubility of oil affected by vaporpressure of the oil, apparently at this stage CO2 relatively actas an ideal gas that does not have any special characteristic ofa solvent. However, at high pressures the solubility of the oilincreased due to the increase in density of CO2. As the density

increases, the distance between molecules decreases and theinteraction between oil and CO2 increases, leading to greateroil solubility in CO2 (Molero Gomez and Martinez de la Ossa,

Table 4 – Analysis of variance showed the processingvariables on the response considered

Source Degree offreedom

Sum ofsquares

Meansquare

P value

Model 9 16.49067 1.832297 0.0001Linear 3 13.00905 4.33635 0.0001Quadratic 3 3.264148 1.088049 0.0005Cross product 3 0.217475 0.072492 0.1469Lack of fit 3 0.1039 0.034633 0.2746

R2 0.9783

230 f o o d a n d b i o p r o d u c t s p r o c e s s i n g 8 6 ( 2 0 0 8 ) 227–231

Fig. 1 – Contour plot and response surface curve showingpredicted response surface of oil yield as a function ofpressure and feed flow (temperature 50 ◦C, time 60 min).

Fig. 2 – Contour plot and response surface curve showingpredicted response surface of oil yield as a function oftemperature and feed flow (pressure 22.5 MPa, time 60 min).

ide. At the wheat germ humidity 3.17% and 7.24% for 60 min,the extraction oil yield was 9.7% and 9.15%, respectively. Oilyield reached its maximum value after 60 min in the case of

Fig. 3 – Influences of wheat germ humidity on the

2002). So, the pressure factor is dominant in determining themass transfer rate and the diffusion of oil in wheat germ.

Fig. 2 shows the effects of temperature and flow rate onyield at pressure 27.5 MPa (coded value of pressure 0). Theincrease yield with increase in flow rate could also be seen inthe figure and remain constant at high values gradually. Whenflow rate varied from 15 to 25 L/h, the oil yield reached from8.2% up to about 8.7% with pressure increase possibly due tothe decrease of resistance force in mass transfer. It also can beseen that the maximum oil yield together with the minimumconsumption of solvent was achieved at about 22.5 L/h (codedvalue 0.5). A higher flow rate will give a somewhat higher yieldbut with a much higher solvent consumption; a lower flow ratewill reduce solvent consumption but produce notably loweryields. Therefore, the flow rate of 22.5 L/h gave a maximumyield with minimum solvent consumption.

From Fig. 2, it could be seen that the best extraction yieldwas reached at an operating temperature of 50 ◦C. At this tem-perature some 10% of the highest oil yield is achieved at 35 MPaand 22.5 L/h. Consequently, operating temperature 40 ◦C, pres-sure 35 MPa and flow rate 22.5 L/h are considered to be theefficient operating conditions.

No significant differences were detected in the extractionyields obtained when the extraction processes were carried

out with partially hydrated wheat germ and almost fully dehy-drated samples. As can be seen in Fig. 3, the humidity of

wheat germ influenced negatively the extraction processesdue to the variation of the solvent capacity of carbon diox-

extraction oil yield by SFC-CO2 (temperature, 50 ◦C; flowrate 22.5 L/h; pressure 35 MPa).

f o o d a n d b i o p r o d u c t s p r o c e s s

Table 5 – Fatty acid compositiona (%) of wheat germ oilsextracted with supercritical carbon extraction andsolvent extraction

Fatty acid composition (%) Extraction method

SC-CO2 Hexane (Soxhlet, 12 h)

Palmitic acid 17.20 17.28Stearic acid 1.27 1.20Oleic acid 18.47 18.39Linoleic acid 55.63 55.75Linolenic acid 7.43 7.38

Conditions of SC-CO2: Pressure 35 MPa, temperature 50 ◦C and flowrate 22.5 L/h.

Sma

hcabe

4

RohtiSpaea

A

TAa

r

a GC area percentage.

C-CO2 with different humid material. After reaching maxi-um values, the curve for the oil yield of wheat germ attainsplateau in the cases.

Table 5 shows the free acid of wheat germ obtained usingexane as solvent, as well as the extracted by supercriticalarbon dioxide. The composition of linoleic acid and oleiccid reached up to 55.63% and 18.47%, respectively. As cane observed, the content in fatty acids did not depend on thextraction method.

. Conclusions

esponse surface methodology was successfully applied forptimization of wheat germ oil yield parameters by SFE. Theigh regression coefficients of second-order polynomial ofhe response showed that model fitted data well. The max-mum wheat germ oil yield was obtained to be 10.15% byFE are obtained when SFE was carried out at 35 MPa ofressure, 40 ◦C of temperature, solvent flow rate 22.5–25 L/hnd extraction time 1 h. The quality of wheat germ oilxtracted by SFE was similar to that of oil extracted by hex-ne.

cknowledgements

his work was supported by Natural Science Foundation ofnhui Province (No. 03041302). And the authors gratefullycknowledge the contribution of the following to this work:

i n g 8 6 ( 2 0 0 8 ) 227–231 231

Prof. D.R. Ma for her assistances in operating SFE system andDr. M.H. He for his valuable comments on this manuscript.

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