Enhanced Extraction of Oil from Flaxseed (Linum usitatissimum L.) Using Microwave Pre-treatment
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1 Journal of Oleo Science Copyright 2015 by Japan Oil Chemists Society doi : /jos.ess15099 Enhanced Extraction of Oil from Flaxseed (Linum usitatissimum L.) Using Microwave Pre-treatment Guangyue Ren 1*, Wei Zhang 1, Shangde Sun 2*, Xu Duan 1 and Zhenshan Zhang 2 1 College of Food and Bioengineering, Henan University of Science and Technology, Luoyang, , China 2 School Food Science & Engineering, Henan University of Technology, Zhengzhou , China Abstract: The effect of microwave (MW) pre-treatment on the extraction of flaxseed oil was investigated by hot extraction (HE). Nine MW pre-treatments were established, combining three MW radiation intensities (12, 18 and 24 W/g) and three MW radiation times of pre-treatment (90, 150 and 210 s). Extraction yield increased significantly with MW pre-treatments of flaxseed, and a max oil extraction yield (78.11%) can be obtained using MW pre-treatment at 18 W/g for 210 s. Scanning electronic microscopy showed that the microstructure of treated samples (18 W/g and 210 s) was modified compared with that of untreated samples. The fatty acid compositions (palmitic acid 5.85±0.01%, stearic acid 3.00±0.01%, oleic acid 17.64±0.07%, linoleic acid 16.16±0.06%, and linolenic acid 57.37±1.32%) of the oils extracted by the MW pre-treatments HE were similar with that of the conventional HE method. Results showed that fatty acid compositions of flaxseed oil were not affected by MW pre-treatments. Key words: microwave pre-treatment, hot extraction, oil, flaxseed 1 INTRODUCTION Flax Linum usitatissimum L. had been cultivated as early as 6000 BC when it was first found in Eastern Turkey. At present, flax has mainly been cultivated in Canada, Argentina, America, China and India 1. The stem of flax can be used to produce the fibre of linen, and flaxseeds linseeds can be used for oil and feed preparation. Therefore, flax can be classified into fibered flax fibre-rich and oily flax oil-rich. Flaxseed usually contains about 40 oil, 30 dietary fibre, 20 protein, 4 ash, and 6 moisture, and riched in some excellent physiologically active components lignans, minerals, and vitamin, etc. Therefore, flaxseed is one of the most popular oilseeds for human consumption and has been received increasing interest 2. Flaxseed oil riched in alpha linolenic acid ALA 55, an omega-3 fatty acid, which is 5.5 times higher than that of other vegetable oils, such as, walnuts and canola oil 2. In human intestine, ALA can be metabolized to form eicosapentaenoic acid EPA and docosahexaenoic acid, which can reduce the risk of lifestyle diseases 3. Previous studies also found that flaxseed oil has positive effects by decreasing the risk of many diseases such as hyperlipidemia 4, mammary cancer 5, atherosclerosis 6, and cardiovascular disease 7. The flaxseed oil can also be used for industrial purposes, such as, paints, coatings, varnishes, inks, cos- metics and linoleum 8, 9. Conventional extraction methods of flaxseed oil are pressing and solvent extraction. Pressing is often associated with lower yield, while solvent extraction also presents some disadvantages, such as, long extraction time, high operation costs, environment pollution, and security problems. Therefore, some improved extraction techniques, for example, supercritical fluid extraction, aqueous enzymatic extraction, and ultrasound-assisted extraction techniques have been attracted much attention. Because microwave MW can reduce processing times, save energy, and deliver energy directly to materials through molecular interaction with the electromagnetic field 10, 11. Therefore, MW-assisted and MW pre-treatment have been used for extracting oil from many agri-food materials 12, such as, Chilean hazelnuts 13, citrus peels 14, and sesame 15. However, there is no literature to report extracting oil from flaxseed by MW-assisted and MW pre-treatment methods. The aim of this study was to investigate the effects of MW pre-treatment prior to the oil extraction by hot extrac- Abbreviations: ALA, alpha linolenic acid; FAME, fatty acid methyl esters; HE, hot extraction; MW, microwave; SEM, scanning electron microscope; t MW, microwave radiation time; t EX, solvent extraction time * Correspondence to: Guangyue Ren; Shangde Sun, College of Food and Bioengineering, Henan University of Science and Technology, Luoyang, , China; School Food Science & Engineering, Henan University of Technology, Zhengzhou , China guangyueyao@163.com; sunshangde@hotmail.com Accepted June 19, 2015 (received for review April 28, 2015) Journal of Oleo Science ISSN print / ISSN online
2 G. Ren, W. Zhang and S. Sun et al. tion HE method on the microstructure, yield of oils, and oil quality of flaxseed. 2 EXPERIMENTAL 2.1 Materials The flaxseed was acquired from Zhangjiakou region located in the north of China. The seeds were cleaned by airing and sieving, and then all foreign material and broken seeds were removed manually. The initial water content of the flaxseed was determined by oven drying at for 24 h. The initial water content of the seeds was 6.21 wet basis. An analytical grade n-hexane used in the extraction was purchased from Beijing Beihua Chemical Company Beijing, China. 2.2 Microwave pre-treatment For each MW pre-treatment, 30 g of flaxseeds were arranged in a petri dishes Φ90 mm, covered with another petri dishes Φ95 mm, and then placed on the centre of the turntable plate of the microwave variable power oven WG900DSL23DK6, Galanz, China. Samples were MWtreated at a frequency of 2450 MHz. Nine MW pre-treatments were established, combining three MW radiation intensities 12, 18 and 24 W/g and three MW radiation times t MW of pre-treatment 90, 150 and 210 s. Samples were collected and cooled to room temperature prior to the breaking treatment, and the water content was determined by the method above. Flaxseed samples were dried in an oven at 60 for 6 h, and used as control processing untreated samples. 2.3 Breaking treatment Treated and untreated samples were crushed into powder in a mixer B-400, Büchi Labortechnik AG, Switzerland with a size range of mm. The resulted powder was kept in a vacuum dryer until the oil extraction. 2.4 Oil extraction Flaxseed oil was obtained by Büchi s HE solvent method. Extraction technique of the HE contains three main steps: extraction, flushing, and drying. In the first step, solvent extraction time t EX was 30 min with the 11 and 4 heating grade respectively in lower and upper heater; in the second step, heater does not work; in the third step, drying time was 10 min with 8 heating grade in lower heater. For each extraction processing, flaxseed powder was mixed well by manual quartering method, and then weighted 5 g and packed by filter paper. Four samples were prepared and put into Büchi s sample tube, respectively. HE was operated with n-hexane as extraction solvent. Later, all the extracted oil was collected for analysis. Fine particles in the extracted oil were separated by filtration, and these filtered crude oils were subsequently centrifuged in a centrifuge LG10-2.4A, Beijing medical centrifuge factory, China at 5000 rpm for 15 min to remove the precipitation. 2.5 Oil extraction yield determination The yield of flaxseed oil was calculated as follows: yield W e 100 W t Where: W e is the mass of flaxseed oil extracted from the sample g and W t is the mass of total oil in the sample g. W t was determined under standard conditions by Büchi s Soxhlet extraction method B-811, Büchi LabortechnikAG, Switzerland with hexane for 8 hours 16. The mass of total oil in the 100 g flaxseed water content, 6.21, based on dry weight was g average standard deviation. 2.6 Scanning electron microscopy SEM analysis The micrographic appearances of the MW-treated and untreated powder samples were observed by SEM S- 3400N, Hitachi Instruments Ltd., Japan. The samples were fixed on the silicon wafer and coated with gold to avoid charging under the electron. The thickness of Aucoatings was about 100 nm. An accelerating potential of 30 kv was used during micrography. 2.7 GC analysis Fatty acid compositions of the oils extracted by the MW pre-treatments HE and the conventional HE method were determined by GC after derivatization to fatty acid methyl esters FAME. The preparation of FAME was performed according to the ISO 5509 standard method 17. FAME separation and identification were carried out on GC CP-3800, Varian Inc, Walnut Creek, CA equipped with a flame ionization detector and capillary column HP-Innowax 30 m 0.32 mm 0.25 μm. The amount of each sample injected was 1.0 μl. Nitrogen, at a constant flow 1.0 ml/min, was used as the carrier gas and a spilt/spiltless injector was used with a split ratio of 20:1. The injector temperature was 250 and the detector temperature was 270. The column temperature was programmed form 100 to 180 at 20 /min, and then to 230 at 10 /min and held at 230 for 5 min. FAME was identified by comparison with the standard FAME Sigma, USA. FAME was quantified as percentages of the total methyl ester peak areas. 2.8 Statistical analysis All the experiments including water content, MW pretreatment, HE, and GC were carried out in quadruple, and the average values were reported. And the repeatability was satisfactory of all experiments for the MW pre-treatment, HE, and GC. The analysis of variance was carried out based on the experimental data by using the SAS statistical package and 1044
3 Enhanced extraction of oil using microwave pre-treatment p values were used to determine the extent of effect of MW pre-treatment on the yield and quality of extracted oil. A probability of p 0.05 was considered significant. 3 RESULTS AND DISCUSSION 3.1 Effect of MW pre-treatment on flaxseed powder microstructure In order to illuminate the influence of MW pre-treatment on the microstructure of the seeds and to explain the extraction mechanism, SEM was applied in this study. Figure 1 shows the flaxseed powder contain oil-bearing microstructures. Figure 1A shows the approximate intact structure and the oil in the form of globules on the surface of the untreated seed. The oil on the surface can be easily extracted, but the near intact structure and adhering membranes make oil in interior distilled difficultly. Figure 1B shows the effect of MW pre-treatment on the microstructure of flaxseed under the processing condition 18 W/g and 210 s. Results showed that the modification of the seed structure, exfoliation takes place and in porosity, large number of thin broken seed fragment and smaller lipid globules appeared on the surface of the exfoliation. Therefore, the oil in these thin broken fragments can be easily extracted. MW heating vaporizes the water of the seeds, and increases the pressure in its interior, which result in the disintegration of the material. These results demonstrated that the application of MW pre-treatment can rupture the flaxseed structure and improve the efficiency of the extraction of oil from seeds. 3.2 Effect of MW pre-treatment on extraction yield of flaxseed oil The effect of MW pre-treatments on the extraction yield of oil was investigated to extract oil by HE method with using MW pre-treatment and untreated flaxseed samples. The oil extraction yields were shown in Table 1. It can be seen form Table 1, oil extraction yield from flaxseeds increased significantly using MW processing and extracted oil yields were obtained under the processing condition 18 W/g, 210 s and 24 W/g, 210 s using MW pre-treatment, which were higher than that of untreated sample. However, when the t MW increased to 210 s, extraction yields of 18 W/g MW radiation intensity were similar with that of 24 W/g. These results showed that the effect of MW pre-treatment is similar at 18 W/g. Therefore, 18 W/g and 210 s were selected as the MW pretreatment conditions to extract oil for test analysis. 3.3 Effect of MW pre-treatment on the fatty acid composition of flaxseed oil The fatty acid compositions of flaxseed oils extracted by MW-treated 18 W/g and 210 s and untreated samples were given in Table 2 and Fig. 1. Five main fatty acids of flaxseed oil Table 2 and Fig. 2 were similar to that found in previous reports 16. And there were no remarkable differences p 0.05 between MWtreated sample and untreated sample. These indicate that the effect of MW pre-treatment on the fatty acid composition of flaxseed oil was negligible. Similar effects of MW treatment on fatty acid composition were also found in other seeds, such as, Chilean hazelnuts 13, soya bean 18, peanut seeds 19, sunflower seed 20 and pumpkin seeds 21. Linolenic acid was the most abundant fatty acid in flaxseed oil. The contents of linolenic acid of MW pre-treatment and untreated seeds were and 58.46, re- A B Fig. 1 SEM images of flaxseed powder contain oil-bearing: A untreated flaxseed powder, B MW-treated (18 W/g and 210 s) flaxseed powder. 1045
4 G. Ren, W. Zhang and S. Sun et al. Table 1 Effect of MW pre-treatments on extraction yield of flaxseed oil (standard deviation in parentheses). MW radiation intensity / W/g MW radiation time / s Extraction yield / % Moisture content / % (d. b.) ( 3.32) 5.47 ( 0.26) ( 2.92) 5.06 ( 0.31) ( 3.51) 4.27 ( 0.23) ( 2.39) 4.38 ( 0.15) ( 2.87) 4.02 ( 0.17) ( 2.56) 3.84 ( 0.13) ( 0.87) 3.68 ( 0.12) ( 1.32) 3.34 ( 0.21) ( 1.56) 3.15 ( 0.13) control ( 0.55) 5.43 ( 0.21) Table 2 Fatty acid compositions of the flaxseed oil (standard deviation in parentheses). Fatty acid compositions/ % MW pre-treatment/ % Control/ % Palmitic C16: ( 0.01) 5.34 ( 0.00) Stearic C18: ( 0.01) 3.05 ( 0.01) Oleic C18:1 n ( 0.07) ( 0.06) Linoleic C18:2 n ( 0.06) ( 0.04) Linolenic C18:3 n ( 1.32) ( 1.27) Saturated fatty acids (S) Unsaturated fatty acids (U) Polyunsaturated fatty acids (P) P/S U/S n-3/n n-3, omega-3 fatty acid; n-6, omega-6 fatty acid; n-9, omega-9 fatty acid. spectively Table 2. Furthermore, the unsaturated fatty acids U, polyunsaturated fatty acids P, P/S, U/S and n-3/n-6 decreased slightly in the oil extracted by the MW pre-treatment, which may be due to the inactivation of the oxidative enzyme. Fig. 2 A gas chromatogram of the extraction flaxseed oil after MW pre-treatment (18 W/g and 210 s). n-3, omega-3 fatty acid; n-6, omega-6 fatty acid; n-9, omega-9 fatty acid. 4 CONCLUSION In this work, the effect of MW pre-treatments on flaxseed was investigated. Results showed that the flaxseed oil extraction yield increased significantly with MW pre-treatments compared with untreated flaxseed. A max oil extraction yield can be obtained using MW pre-treatment at 18 W/g for 210 s. SEM results indicated that the microstructure of flaxseed was modified after the MW- 1046
5 Enhanced extraction of oil using microwave pre-treatment treatment. The effect of MW pre-treatment on the fatty acid compositions of flaxseed oils extracted were negligible. Therefore, the MW pre-treatment may be considered as an effective and feasible method for the production of flaxseed oil. ACKNOWLEDGMENTS Supported by National Natural Science Foundation of China , U , and Science and Technology Project of Henan Province of China References 1 Wang, B.; Li, D.; Wang, L. J.; Huang, Z. G.; Zhang, L.; Chen, X. D.; Mao, Z. H. Effect of moisture content on the physical properties of flaxseed. Int. J. Food Eng. 3, Herchi, W.; Arráez-Román, D.; Trabelsi, H.; Bouali, I.; Boukhchina, S.; Kalle, H.; Segura-Carretero, A.; Fernández-Gutierrez, A. Phenolic Compounds in Flaxseed: a Review of Their Properties and Analytical Methods. An Overview of the Last Decade. J. Oleo Sci. 63, Visentainer, J. V.; Souza, N. E.; Makoto M.; Hayashi C.; Franco M. R. B. Influence of diets enriched with flaxseed oil on the α-linolenic, eicosapentaenoic and docosahexaenoic fatty acid in Nile tilapia Oreochromis niloticus. Food Chem. 90, Vijaimohan, K.; Mallika Jainu, K. E.; Sabitha, S.; Subramaniyam, C.; Anandhan, C. S.; Devi, S. Beneficial effects of alpha linolenic acid rich flaxseed oil on growth performance and hepatic cholesterol metabolism in high fat diet fed rats. Life Sci. 79, Chen, J.; Wang, L.; Thompson, L. U. Flaxseed and its components reduce metastasis after surgical excision of solid human breast tumor in nude mice. Cancer Lett. 234, Prasad, K. Dietary flax seed in prevention of hypercholesterolemic atherosclerosis. Atherosclerosis 132, Lanzmann-Petithory, D.; Pueyo, S.; Renaud, S. Primary prevention of cardiovascular diseases by alphalinolenic acid. Am. J. Clin. Nutr. 76, Oomah, B. D.; Mazza, G. Fractionation of flaxseed with a batch dehuller. Ind. Crop. Prod. 9, Zhang, Z. S.; Li, D.; Wang, L. J.; Ozkan, N.; Chen, X. D.; Mao, Z. H.; Yang, H. Z. Optimization of ethanol water extraction of lignans from flaxseed. Sep. Purif. Technol. 57, Thostenson, E. T.; Chou, T. W. Microwave processing: Fundamentals and applications. Compo. Part A-Appl. S. 30, Lokman, I. M.; Rashid, U.; Zainal, Z.; Yunus, R.; Taufiq- Yap, Y. H. Microwave-assisted biodiesel production by esterification of palm fatty acid distillate. J. Oleo Sci. 63, Sumi, T.; Horikoshi, S. Microwave synthesis, extraction, improvement and degradation in oil chemistry. J. Oleo Sci. 62, Uquiche, E.; Jeréz, M.; Ortíz, J. Effect of pretreatment with microwaves on mechanical extraction yield and quality of vegetable oil from Chilean hazelnuts Gevuina avellana Mol. Innov. Food Sci. Emerg. 9, Bousbia, N.; Vian, M. A.; Ferhat, M. A.; Meklati, B. Y.; Chemat, F. A new process for extraction of essential oil from Citrus peels: Microwave hydrodiffusion and gravity. J. Food Eng. 90, Papadakis, E. N.; Vryzas, Z.; Papadopoulou-Mourkidou, E. Rapid method for the determination of 16 organochlorine pesticides in sesame seeds by microwave-assisted extraction and analysis of extracts by gas chromatography mass spectrometry. J. Chromatogr. A 1127, Zhang, Z. S.; Wang, L. J.; Li, D.; Jiao, S. S.; Chen, X. D.; Mao, Z. H. Ultrasound-assisted extraction of oil from flaxseed. Sep. Purif. Technol. 62, ISO. Animal and vegetable fats and oils-preparation of methyl esters of fatty acids Method ISO International Organization for Standardization, Geneva, pp Takagi, S.; Ienaga, H.; Tsuchiya, C.; Yoshida, H. Microwave roasting effects on the composition of tocopherols and acyl lipids within each structural part and section of a soya bean. J. Sci. Food Agric. 79, Yoshida, H.; Hirakawa, Y.; Tomiyama, Y.; Nagamizua, T.; Mizushina, Y. Fatty acid distributions of triacylglycerols and phospholipids in peanut seeds Arachis hypogaea L. following microwave treatment. J. Food Compos. Anal. 18, Anjum, F.; Anwar, F.; Jamil, A.; Iqbal, M. Microwave roasting effects on the physico-chemical composition and oxidative stability of sunflower seed oil. J. Am. Oil Chem. Soc. 83, Yoshida, H.; Tomiyama, Y.; Hirakawa, Y.; Mizushina, Y. Microwave roasting effects on the oxidative stability of oils and molecular species of triacylglycerols in the kernels of pumpkin Cucurbita spp. seeds. J. Food Compos. Anal. 19,
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