Available online at www.sciencedirect.com Journal of Integrative Agriculture 2012,11(1):151-158 ScienceDirect January 2012 RESEARCH ARTICLE Response Surface Optimization of Nigella glandulifera Freyn Seed Oil Yield by Supercritical Carbon Dioxide Extraction ZHANG Jun-ping.2,HOU Xi-lin1.2,YU Tian',LI Ying1.2 and DONG Hai-yan.2 i State Key Laboratory of Crop Genetics Germplasm Enhancement,Nanjing Agriculture University,Nanjing 210095,P.R.China Key Laboratory of Southern Vegetable Crop Genetic Improvement,Ministry of Agriculture,Nanjing 210095,P.R.China Abstract Supercritical carbon dioxide (SC-CO2)extraction was employed to extract oil from Nigella glandulifera Freyn seed in this study.Response surface methodology(RSM)was applied to evaluate the effects of the process parameters(pressure, temperature,and CO,flow rate)on oil yield of N.glandulifera seed.A Box-Behnken design was used to optimize the extraction parameters.The analysis of variance indicated that the linear coefficients of pressure and CO,flow rate,the quadratic term coefficients of pressure and temperature and the interactions between pressure and temperature,as well as temperature and CO,flow rate,had significant effects on the oil yield(P<0.05).The optimal conditions to obtain the maximum oil yield from N.glandulifera seed were pressure 30.84 MPa,temperature 40.57C,and CO,flow rate 22.00 L h-1. Under these optimal conditions,the yield of oil was predicted to be 38.19%.The validation experiment results agreed with the predicted values.The fatty acid composition of N.glandulifera seed oil extracted using SC-CO2 was compared with that of oil obtained by Soxhlet method.The results showed that the fatty acid compositions of oil extracted by the two methods were similar.Identification of oil compounds with gas chromatography-mass spectrometry(GC-MS)showed that the contents of unsaturated fatty acids linoleic acid(48.30%),oleic acid(22.28%)and saturated fatty acids palmitic acid (16.65%).stearic acid (4.17%)were the most abundant fatty acids in seed oil from N.glandulifera Key words:supercritical carbon dioxide extraction,Nigella glandulifera Freyn seed oil,response surface methodology gas chromatography-mass spectrometry,fatty acids asthma (Xiao et al.2002).They contain about 35- INTRODUCTION 42%oil.which is rich in linoic acid and oleic acid. Linoic acid and oleic acid are unsaturated fatty acids, Nigella glandulifera Freyn,a plant of Nigella genus in and are considered to be beneficial to the health of Ranunculaceae family,is widely distributed in Xinjiang, mankind,so its oil has high value for exploitation and Yunnan,and Tibet of China.The seeds of N.glandulifera utilization and can be used in foods,pharmaceuticals, are well-known as a Uighur's traditional medicine and and cosmetic formulations and so on. food,which are included in Pharmacopoeia of the Oil extraction using supercritical carbon dioxide(SC- People's Republic of China from 1997 to now.These CO,)has gained increasing attention over the traditional seeds are believed to have diuretic,analgesic, techniques,like steam distillation and solvent extraction spasmolytic,galactagogue,and bronchodilator func- SC-CO,has the advantages of using nontoxic,nonex- tions to cure edema,urinary calculus,and bronchial plosive and volatile solvent,which protects extracts from Received 28 October,2010 Accepted 4 July,2011 ZHANG Jun-ping,E-mail:xj2005zhangip@126.com;Correspondence HOU Xi-lin,Tel:+86-25-84395917,E-mail:hxl@njau.edu.cn 2012,CAAS.All rights reserved.Published by ElsevierLtd
Journal of Integrative Agriculture 2012, 11(1): 151-158 January 2012 © 2012, CAAS. All rights reserved. Published by Elsevier Ltd. RESEARCH ARTICLE Response Surface Optimization of Nigella glandulifera Freyn Seed Oil Yield by Supercritical Carbon Dioxide Extraction ZHANG Jun-ping1, 2, HOU Xi-lin1, 2, YU Tian1, LI Ying1, 2 and DONG Hai-yan1, 2 1 State Key Laboratory of Crop Genetics & Germplasm Enhancement, Nanjing Agriculture University, Nanjing 210095, P.R.China 2 Key Laboratory of Southern Vegetable Crop Genetic Improvement, Ministry of Agriculture, Nanjing 210095, P.R.China Abstract Supercritical carbon dioxide (SC-CO2) extraction was employed to extract oil from Nigella glandulifera Freyn seed in this study. Response surface methodology (RSM) was applied to evaluate the effects of the process parameters (pressure, temperature, and CO2 flow rate) on oil yield of N. glandulifera seed. A Box-Behnken design was used to optimize the extraction parameters. The analysis of variance indicated that the linear coefficients of pressure and CO2 flow rate, the quadratic term coefficients of pressure and temperature and the interactions between pressure and temperature, as well as temperature and CO2 flow rate, had significant effects on the oil yield (P<0.05). The optimal conditions to obtain the maximum oil yield from N. glandulifera seed were pressure 30.84 MPa, temperature 40.57°C, and CO2 flow rate 22.00 L h-1. Under these optimal conditions, the yield of oil was predicted to be 38.19%. The validation experiment results agreed with the predicted values. The fatty acid composition of N. glandulifera seed oil extracted using SC-CO2 was compared with that of oil obtained by Soxhlet method. The results showed that the fatty acid compositions of oil extracted by the two methods were similar. Identification of oil compounds with gas chromatography-mass spectrometry (GC-MS) showed that the contents of unsaturated fatty acids linoleic acid (48.30%), oleic acid (22.28%) and saturated fatty acids palmitic acid (16.65%), stearic acid (4.17%) were the most abundant fatty acids in seed oil from N. glandulifera. Key words: supercritical carbon dioxide extraction, Nigella glandulifera Freyn seed oil, response surface methodology, gas chromatography-mass spectrometry, fatty acids INTRODUCTION Nigella glandulifera Freyn, a plant of Nigella genus in Ranunculaceae family, is widely distributed in Xinjiang, Yunnan, and Tibet of China. The seeds of N. glandulifera are well-known as a Uighur’s traditional medicine and food, which are included in Pharmacopoeia of the People’s Republic of China from 1997 to now. These seeds are believed to have diuretic, analgesic, spasmolytic, galactagogue, and bronchodilator functions to cure edema, urinary calculus, and bronchial asthma (Xiao et al. 2002). They contain about 35- 42% oil, which is rich in linoic acid and oleic acid. Linoic acid and oleic acid are unsaturated fatty acids, and are considered to be beneficial to the health of mankind, so its oil has high value for exploitation and utilization and can be used in foods, pharmaceuticals, and cosmetic formulations and so on. Oil extraction using supercritical carbon dioxide (SCCO2 ) has gained increasing attention over the traditional techniques, like steam distillation and solvent extraction. SC-CO2 has the advantages of using nontoxic, nonexplosive and volatile solvent, which protects extracts from Received 28 October, 2010 Accepted 4 July, 2011 ZHANG Jun-ping, E-mail: xj2005zhangjp@126.com; Correspondence HOU Xi-lin, Tel: +86-25-84395917, E-mail: hxl@njau.edu.cn
152 ZHANG Jun-ping et al. thermal degradation and solvent contamination(Brunner The regression analysis of the response function with 1994).There are reports on SC-CO,extraction as an statistical analysis are given in Table 2.Statistical test- excellent alternative to the use of chemical solvents in ing of the model was performed in the form of ANOVA the extraction of oils from different plants such as corn Here,the value for Fand P(probability)(P<0.05 when (List et al.1984),soybean (Friedrich et al.1982)and significant)were 80.5708 and 0.0001,respectively,and cotton seeds(Kuk and Hron 1994;Bhattacharjee et al. the lack-of-fit of 0.7117 was not significant (P>0.05). 2007),cocoa beans(Saldafia et al.2002),green tea indicating that the generated model adequately explained (Chang et al.2000),and ginseng (Wang et al.2001). the data variation and significantly represented the ac- However,no studies have been reported on the oil ex- tual relationship between the reaction parameters.The traction from N.glandulifera seed by SC-CO.extrac- determination coefficient R2=0.9904 indicating that tion to our knowledge.Response surface methodol- 99.04%of the variability in the response could be ex- ogy(RSM)which combines mathematics with statis- plained by the model.From the P-values of each model tics is often used to design experiments,build models, term,we concluded that the linear coefficients of pres- and evaluate the effects of factors (Yue et al.2008). sure and CO.flow rate and the quadratic terms of pres- The main advantage of RSM is the small number of sure and temperature,had highly significant effects on experimental trials needed to evaluate multiple param- the oil yield at the 1%level(P<0.01).The interactions eters and their interactions(Chow et al.1998),and it between pressure and temperature,as well as tempera- has been used successfully in food processing opera- ture and CO,flow rate,had significant effects on the tions(Hierro and Santa-Maria 1992;Reverchon 1997; oil yield at the 5%level (P<0.05). Lee et al.2000;Huang et al.2008;Liu et al.2009). SC-CO,was used to extract seed oil from N.glanelulifera Response surface analysis in this work.The effects of independent factors (pressure,temperature,and CO,flow rate)on the oil From Eq.(1)we can see that the oil yield of N.glandulifera yield of N.glandulifera seed were investigated.RSM seed has a complex relationship with independent was employed to build a model between the oil yield variables.The best way of expressing the effects of and these independent factors as well as to develop a independent variables on the oil yield within the experi- model equation that will predict and determine the opti- mental space under investigation is to generate response mum conditions for the oil yield surface plots of the equation.The three-dimensional response surfaces curves and corresponding contour RESULTS AND DISCUSSION plots were obtained using the Design Expert and are shown in Figs.1,2,and 3 to illustrate the relationship between independent variables and the oil yield. Model fitting Fig.I shows response surface curve and its con- tour plot for the effects of pressure and temperature on Oil yields obtained from all the experiments are listed in the oil yield and their interaction at a fixed flow rate of Table 1.The experimental data were used to calculate 20 L h.The extraction pressure and temperature the coefficients of the second-order polynomial equation. showed a quadratic effect on the response.At low The application of RSM offered,based on parameter pressure,the oil yield was increased with the increase estimates,an empirical relationship between the response of pressure.This is most likely due to the improve- variable,and the test variables under consideration.By ment of oil solubility resulted from the increased CO, applying multiple regression analysis on the experimen- density with the rise of pressure(Lee et al.2000).When tal data,the response variable and the test variables were the pressure was increased to levels greater than ap- related by the following second-order polynomial proximately 30 MPa,the negative quadratic effect be- equation: gan to have an impact.Such effect of pressure is not Y=36.81+0.78x-0.014x,+1.27x,+0.41xx2-0.099xx3 unexpected,when the pressure becomes too high,a +0.32x-2.16x2-1.64x2+0.031x2 (1) reduction in the solvent diffusivity and mass transfer 2012,CAAS.All rights reserved.Published by Elsevier Ltd
152 ZHANG Jun-ping et al. © 2012, CAAS. All rights reserved. Published by Elsevier Ltd. thermal degradation and solvent contamination (Brunner 1994). There are reports on SC-CO2 extraction as an excellent alternative to the use of chemical solvents in the extraction of oils from different plants such as corn (List et al. 1984), soybean (Friedrich et al. 1982) and cotton seeds (Kuk and Hron 1994; Bhattacharjee et al. 2007), cocoa beans (Saldaña et al. 2002), green tea (Chang et al. 2000), and ginseng (Wang et al. 2001). However, no studies have been reported on the oil extraction from N. glandulifera seed by SC-CO2 extraction to our knowledge. Response surface methodology (RSM) which combines mathematics with statistics is often used to design experiments, build models, and evaluate the effects of factors (Yue et al. 2008). The main advantage of RSM is the small number of experimental trials needed to evaluate multiple parameters and their interactions (Chow et al. 1998), and it has been used successfully in food processing operations (Hierro and Santa-Maria 1992; Reverchon 1997; Lee et al. 2000; Huang et al. 2008; Liu et al. 2009). SC-CO2 was used to extract seed oil from N. glanelulifera in this work. The effects of independent factors (pressure, temperature, and CO2 flow rate) on the oil yield of N. glandulifera seed were investigated. RSM was employed to build a model between the oil yield and these independent factors as well as to develop a model equation that will predict and determine the optimum conditions for the oil yield. RESULTS AND DISCUSSION Model fitting Oil yields obtained from all the experiments are listed in Table 1. The experimental data were used to calculate the coefficients of the second-order polynomial equation. The application of RSM offered, based on parameter estimates, an empirical relationship between the response variable, and the test variables under consideration. By applying multiple regression analysis on the experimental data, the response variable and the test variables were related by the following second-order polynomial equation: Y=36.81+0.78x1 -0.014x2 +1.27x3 +0.41x1 x2 -0.099x1 x3 . +0.32x2 x3 -2.16x1 2 -1.64x2 2 +0.031x3 2 (1) The regression analysis of the response function with statistical analysis are given in Table 2. Statistical testing of the model was performed in the form of ANOVA. Here, the value for F and P (probability) (P<0.05 when significant) were 80.5708 and 0.0001, respectively, and the lack-of-fit of 0.7117 was not significant (P>0.05), indicating that the generated model adequately explained the data variation and significantly represented the actual relationship between the reaction parameters. The determination coefficient R2 =0.9904 indicating that 99.04% of the variability in the response could be explained by the model. From the P-values of each model term, we concluded that the linear coefficients of pressure and CO2 flow rate and the quadratic terms of pressure and temperature, had highly significant effects on the oil yield at the 1% level (P<0.01). The interactions between pressure and temperature, as well as temperature and CO2 flow rate, had significant effects on the oil yield at the 5% level (P<0.05). Response surface analysis From Eq. (1) we can see that the oil yield of N. glandulifera seed has a complex relationship with independent variables. The best way of expressing the effects of independent variables on the oil yield within the experimental space under investigation is to generate response surface plots of the equation. The three-dimensional response surfaces curves and corresponding contour plots were obtained using the Design Expert and are shown in Figs. 1, 2, and 3 to illustrate the relationship between independent variables and the oil yield. Fig. 1 shows response surface curve and its contour plot for the effects of pressure and temperature on the oil yield and their interaction at a fixed flow rate of 20 L h-1. The extraction pressure and temperature showed a quadratic effect on the response. At low pressure, the oil yield was increased with the increase of pressure. This is most likely due to the improvement of oil solubility resulted from the increased CO2 density with the rise of pressure (Lee et al. 2000). When the pressure was increased to levels greater than approximately 30 MPa, the negative quadratic effect began to have an impact. Such effect of pressure is not unexpected, when the pressure becomes too high, a reduction in the solvent diffusivity and mass transfer
Response Surface Optimization of Nigella glandulifera Freyn Seed Oil Yield by Supercritical Carbon Dioxide Extraction 153 Table 1 Box-Behnken design and response for the oil yield of N.glandulifera seed Trial no. Factors Responses Pressure (x,MPa) Temperature(x,C) CO:flow rate (x,Lh) Predicted(%) Observed (% -1(25) 0(40の 1(22) 35.270 35.280 2 -1(25) 0(40) -1(18) 32.358 32.540 -1(25) 1(45) 0(20) 31.906 31.818 -1(25) -1(35) 0(20) 32.765 32.662 0(30) 0(40) 0(20) 36.377 36.811 6 0(30) -1(35) -1(18) 34.345 34.266 0(30) 0(40) 0(20) 36.799 36.811 8 0(30 0(40) 0(20) 37.091 36.811 9 0(30) 0(40) 0(20) 37.099 36.811 0(30) 1451 -1(18) 33.690 33.597 11 0(30) 0(40) 0(20) 36.687 36.811 0(30) -1(35) 1(22) 36.075 36.168 13 0(30) 1(45) 1(22) 36.704 36.783 14 1(35) 0(40) -1(18) 34.299 34.290 5 1(35) -1(35) 0(20) 33.309 33.398 1(35) 1(45) 0(20) 34.085 34.188 > 1(35) 0(40) 1(22) 36.817 36.636 Table 2 Analysis of variance for the fitted quadratic polynomial model Source Sum of squares Mean square F-value P-value Model 51.5472 9 5.7275 80.5708 <0.0001 4.8221 4.8220 67.8341 <0.0001 0.0015 1 0.0015 0.0209 0.8891 12.9388 12.9388 182.0156 <0.0001 x 0.6684 1 0.6683 9.4014 0.0182 r 0.03881 1 0.0388 0.5460 0.4840 x 0.4122 0.4122 5.7981 0.0469 2 19.5706 19.5706 275.3080 <0.0001 x22 11.3029 11.3029 159.0027 <0.0001 x 0.0041 1 0.0041 0.0581 0.8164 Residual 0.4976 7 0.0711 Lack of fit 0.1324 0.0441 0.4833 0.7117 Pure error 0.3652 0.0913 Cor total 52.0448 g R-squared:0.9904 Adj R-squared:0.9781 coefficient will occur,which will offset the increase in CO,flow rate is shown in Fig.2 at the fixed extraction extraction rate caused by higher CO,density(Clifford temperature of 40C.How pressure affect the oil yield 1999;Xu et al.2008;Liu et al.2009).At low has been described in Fig.1.The CO,flow rate showed temperature,the oil yield was increased with the in- a positive and significant effect on the oil yield of crease of temperature.This is most likely due to the N.glandulifera seed.At a definite extraction pressure, increased mass transfer speed.However,at higher tem- the yield of oil was increased slightly with the increase perature levels(approximately 40C),the oil yield was of the CO,flow rate,and nearly reached a peak at the decreased with the further increase of temperature.This highest flow rate tested.It might be due to the de- is most likely due to the reduced density of carbon crease of resistance force in mass transfer with the dioxide,with a consequent reduction of solute solubility. increase of flow rate (Yin et al.2005).So,the opti- For a volatile solute,there is competition between its mum flow rate for the maximum yield ofoil was around solubility in SC-CO,and its volatility (Pourmortazavi 22 L h". and Hajimirsadeghi 2007).In this case,the optimum Fig.3 shows the effect of the extraction tempera- extraction pressure and temperature for the maximum ture and the CO,flow rate on the oil yield at a fixed yield of oil were around 30 MPa and 40C. extraction pressure of 30 MPa.How temperature and The interaction between the extraction pressure and CO,flow rate affect the oil yield has been described in 2012,CAAS.All rights reserved.Published by Elsevier Ltd
Response Surface Optimization of Nigella glandulifera Freyn Seed Oil Yield by Supercritical Carbon Dioxide Extraction 153 © 2012, CAAS. All rights reserved. Published by Elsevier Ltd. coefficient will occur, which will offset the increase in extraction rate caused by higher CO2 density (Clifford 1999; Xu et al. 2008; Liu et al. 2009). At low temperature, the oil yield was increased with the increase of temperature. This is most likely due to the increased mass transfer speed. However, at higher temperature levels (approximately 40°C), the oil yield was decreased with the further increase of temperature. This is most likely due to the reduced density of carbon dioxide, with a consequent reduction of solute solubility. For a volatile solute, there is competition between its solubility in SC-CO2 and its volatility (Pourmortazavi and Hajimirsadeghi 2007). In this case, the optimum extraction pressure and temperature for the maximum yield of oil were around 30 MPa and 40°C. The interaction between the extraction pressure and CO2 flow rate is shown in Fig. 2 at the fixed extraction temperature of 40°C. How pressure affect the oil yield has been described in Fig. 1. The CO2 flow rate showed a positive and significant effect on the oil yield of N. glandulifera seed. At a definite extraction pressure, the yield of oil was increased slightly with the increase of the CO2 flow rate, and nearly reached a peak at the highest flow rate tested. It might be due to the decrease of resistance force in mass transfer with the increase of flow rate (Yin et al. 2005). So, the optimum flow rate for the maximum yield of oil was around 22 L h-1. Fig. 3 shows the effect of the extraction temperature and the CO2 flow rate on the oil yield at a fixed extraction pressure of 30 MPa. How temperature and CO2 flow rate affect the oil yield has been described in Table 1 Box-Behnken design and response for the oil yield of N. glandulifera seed Trial no. Factors Responses Pressure (x1, MPa) Temperature (x2, °C) CO2 flow rate (x3, L h-1) Predicted (%) Observed (%) 1 -1 (25) 0 (40) 1 (22) 35.270 35.280 2 -1 (25) 0 (40) -1 (18) 32.358 32.540 3 -1 (25) 1 (45) 0 (20) 31.906 31.818 4 -1 (25) -1 (35) 0 (20) 32.765 32.662 5 0 (30) 0 (40) 0 (20) 36.377 36.811 6 0 (30) -1 (35) -1 (18) 34.345 34.266 7 0 (30) 0 (40) 0 (20) 36.799 36.811 8 0 (30) 0 (40) 0 (20) 37.091 36.811 9 0 (30) 0 (40) 0 (20) 37.099 36.811 10 0 (30) 1 (45) -1 (18) 33.690 33.597 11 0 (30) 0 (40) 0 (20) 36.687 36.811 12 0 (30) -1 (35) 1 (22) 36.075 36.168 13 0 (30) 1 (45) 1 (22) 36.704 36.783 14 1 (35) 0 (40) -1 (18) 34.299 34.290 15 1 (35) -1 (35) 0 (20) 33.309 33.398 16 1 (35) 1 (45) 0 (20) 34.085 34.188 17 1 (35) 0 (40) 1 (22) 36.817 36.636 Table 2 Analysis of variance for the fitted quadratic polynomial model Source Sum of squares df Mean square F-value P-value Model 51.5472 9 5.7275 80.5708 <0.0001 x1 4.8221 1 4.8220 67.8341 <0.0001 x2 0.0015 1 0.0015 0.0209 0.8891 x3 12.9388 1 12.9388 182.0156 <0.0001 x1x2 0.6684 1 0.6683 9.4014 0.0182 x1x3 0.03881 1 0.0388 0.5460 0.4840 x2x3 0.4122 1 0.4122 5.7981 0.0469 x1 2 19.5706 1 19.5706 275.3080 <0.0001 x2 2 11.3029 1 11.3029 159.0027 <0.0001 x3 2 0.0041 1 0.0041 0.0581 0.8164 Residual 0.4976 7 0.0711 Lack of fit 0.1324 3 0.0441 0.4833 0.7117 Pure error 0.3652 4 0.0913 Cor total 52.0448 16 R-squared: 0.9904 Adj R-squared: 0.9781
154 ZHANG Jun-ping et al. Y(oil yield,%) 36.9 1.00 35.625 0.50 34.35 8 33.075 0.00 R图 31.8 -0.50 1.00 1.00 69 59少2水】 0.50 0.50 0.00 0.00 -1.00 0.50 -0.50 X(pressure,MPa) -1.00 -0.50 0.00 0.50 1.00 X(temperature,C) -1.00-1.00 Xi(pressure,MPa】 Fig.1 Response surface curve and its contour plot for the effects of pressure and temperature at a constant CO,flow rate of 20 L h!on the oil yield. Y(oil yield.%) 38.2 1.00 36.775 35.35 372257 33.92 0.00 35352☐ 32.5 34439 362885 100 -0.50 1.00 35352 0.50 0.50 334766 0.00 0.00 -0.50 -0.50 -1.00 (flow rate,Lh) -1.00-1.00 (pressure,MPa) -1.00 -0.50 0.00 0.50 1.00 Xi(pressure,MPa) Fig.2 Response surface curve and its contour plot for the effects of pressure and CO,flow rate at a constant extraction temperature of 40C on the oil yield. Figs.1 and 2,respectively.At a definite extraction CO,flow rate. temperature,the CO,flow rate displayed a linear effect By solving the inverse matrix with Software Design on the response in Fig.3,there was a sharp increase of Expert,the optimum levels of the tested factors were oil yield from about 33.7 to 36.0%as the flow rate extraction pressure at 30.84 MPa,extraction tempera- was increased from 18 to 22 L h. ture at 40.57C,and flow rate at 22 L h.Under these From these three-dimensional response surface conditions,the maximum predicted yield was 38.19%, curves and corresponding contour plots,it is evident and the observed yield was about 37.87-38.25%,the that the CO,flow rate is the most significant factor experimental values agreed with the predicted values. affecting the oil yield in SC-CO,extraction,followed by the extraction pressure and temperature.The inter- Verification of the predictive model actions of extraction pressure and temperature,as well as extraction temperature and CO,flow rate are signifi- In order to validate the model Eq.(1),a total of 15 cant for the oil extraction from N.glandulifera seed. verification experiments were carried out under differ- compared to the interaction of extraction pressure and ent combinations of pressure,temperature,and time 2012.CAAS.All rights reserved.Published by Elsevier Ltd
154 ZHANG Jun-ping et al. © 2012, CAAS. All rights reserved. Published by Elsevier Ltd. Fig. 2 Response surface curve and its contour plot for the effects of pressure and CO2 flow rate at a constant extraction temperature of 40°C on the oil yield. Fig. 1 Response surface curve and its contour plot for the effects of pressure and temperature at a constant CO2 flow rate of 20 L h-1 on the oil yield. Figs. 1 and 2, respectively. At a definite extraction temperature, the CO2 flow rate displayed a linear effect on the response in Fig. 3, there was a sharp increase of oil yield from about 33.7 to 36.0 % as the flow rate was increased from 18 to 22 L h-1. From these three-dimensional response surface curves and corresponding contour plots, it is evident that the CO2 flow rate is the most significant factor affecting the oil yield in SC-CO2 extraction, followed by the extraction pressure and temperature. The interactions of extraction pressure and temperature, as well as extraction temperature and CO2 flow rate are significant for the oil extraction from N. glandulifera seed, compared to the interaction of extraction pressure and CO2 flow rate. By solving the inverse matrix with Software Design Expert, the optimum levels of the tested factors were extraction pressure at 30.84 MPa, extraction temperature at 40.57°C, and flow rate at 22 L h-1. Under these conditions, the maximum predicted yield was 38.19%, and the observed yield was about 37.87-38.25%, the experimental values agreed with the predicted values. Verification of the predictive model In order to validate the model Eq. (1), a total of 15 verification experiments were carried out under different combinations of pressure, temperature, and time
Response Surface Optimization of Nigella glandulifera Freyn Seed Oil Yield by Supercritical Carbon Dioxide Extraction 155 y (oil yield,% 1.00 382 37.025 0.50 ●373729 35.85 i 34.675 0.00 36677 33.5 -0.50 3307内 07④ 1.00 1.00 ●5866 0.50 0.50 0.00 0.00 34☒ -0.50 -1.00 (flow rate.Lh) 0.50 -1.00-1.00 -1.00 -0.50 0.00 0.50 1.00 X(temperature,C) (temperature,C) Fig.3 Response surface curve and its contour plot for the effects of temperature and CO,flow rate at a constant extraction pressure of 30 MPa on the oil yield. and the result is shown in Fig.4.The plot demon- 37.9 strates that the experimental points are evenly distrib- 37.7 uted around the diagonal of horizontal and vertical axis, which indicates that the experimental values are in good 37.5 1-0.9852r+0.573 agreement with the predicted values,and also suggests d that the predicted second order polynomial model is 37.3 accurate and reliable.Thus,a statistically significant 37.1 multiple regression relationship between the indepen- 37 373 37.5 37.7 37.9 dent variables (pressure,temperature,and CO,flow Observed value (% rate)and the response variable(oil yield)was established. The second order polynomial model could therefore be Fig.4 Observed values vs.predicted values for model verification. effectively used to represent the relationship among the parameters selected. consistent with the effect of N.glandulifera seed in activating blood circulation.The essential fatty acids Fatty acid composition of N.glandulifera seed oil like linoleic acid are not easily synthesized in the human body and must be supplied externally through the The fatty acid composition of N.glandulifera seed oil diet,and N.glandulifera seed oil can be a good nutri- extracted by SC-CO,and by Soxhlet method was de- tional supplement as a source of linoleic acid. termined by gas chromatography-mass spectrometry (GC-MS)and is shown in Table 3.The result shows that the fatty acid composition of oil extracted by the CONCLUSION two methods is similar,and the content of linoleic acid and oleic acid reached up to 48.30 and 22.28%, RSM was successfully applied for optimization of SC- respectively.As can be observed,the composition and CO,extraction parameters for N.glandulifera seed oil content in fatty acids do not depend on the extraction yield.The response surface plots indicated that the method.The oil contained 12 fatty acids,among them, three factors(pressure,temperature and CO,flow rate) linoleic acid was the most abundant unsaturated fatty significantly influenced the oil yield,independently and acids.This work also shows that N.glandulifera Freyn interactively.The optimum process parameters were seed oil is a rich source of linoleic acid.Linoleic acid obtained as:pressure 30.84 MPa,temperature 40.57C, has the ability to inhibit platelet aggregation,which is and CO,flow rate 22 L h.Under these conditions,the 2012,CAAS.All rights reserved.Published by Elsevier Ltd
Response Surface Optimization of Nigella glandulifera Freyn Seed Oil Yield by Supercritical Carbon Dioxide Extraction 155 © 2012, CAAS. All rights reserved. Published by Elsevier Ltd. consistent with the effect of N. glandulifera seed in activating blood circulation. The essential fatty acids like linoleic acid are not easily synthesized in the human body and must be supplied externally through the diet, and N. glandulifera seed oil can be a good nutritional supplement as a source of linoleic acid. CONCLUSION RSM was successfully applied for optimization of SCCO2 extraction parameters for N. glandulifera seed oil yield. The response surface plots indicated that the three factors (pressure, temperature and CO2 flow rate) significantly influenced the oil yield, independently and interactively. The optimum process parameters were obtained as: pressure 30.84 MPa, temperature 40.57°C, and CO2 flow rate 22 L h-1. Under these conditions, the Fig. 4 Observed values vs. predicted values for model verification. and the result is shown in Fig. 4. The plot demonstrates that the experimental points are evenly distributed around the diagonal of horizontal and vertical axis, which indicates that the experimental values are in good agreement with the predicted values, and also suggests that the predicted second order polynomial model is accurate and reliable. Thus, a statistically significant multiple regression relationship between the independent variables (pressure, temperature, and CO2 flow rate) and the response variable (oil yield) was established. The second order polynomial model could therefore be effectively used to represent the relationship among the parameters selected. Fatty acid composition of N. glandulifera seed oil The fatty acid composition of N. glandulifera seed oil extracted by SC-CO2 and by Soxhlet method was determined by gas chromatography-mass spectrometry (GC-MS) and is shown in Table 3. The result shows that the fatty acid composition of oil extracted by the two methods is similar, and the content of linoleic acid and oleic acid reached up to 48.30 and 22.28%, respectively. As can be observed, the composition and content in fatty acids do not depend on the extraction method. The oil contained 12 fatty acids, among them, linoleic acid was the most abundant unsaturated fatty acids. This work also shows that N. glandulifera Freyn seed oil is a rich source of linoleic acid. Linoleic acid has the ability to inhibit platelet aggregation, which is Fig. 3 Response surface curve and its contour plot for the effects of temperature and CO2 flow rate at a constant extraction pressure of 30 MPa on the oil yield