Chemical Engineering and Processing 50 (2011) 1207–1213 Contents lists available at ScienceDirect Chemical Engineering and Processing: Process Intensification journal homepage: www.elsevier.com/locate/cep Extraction of oil from Moringa oleifera kernels using supercritical carbon dioxide with ethanol for pretreatment: Optimization of the extraction process Hoang N Nguyen a,∗ , Pag-asa D Gaspillo a , Julius B Maridable a , Roberto M Malaluan b , Hirofumi Hinode c , Chris Salim c , Ha K.P Huynh d a Department of Chemical Engineering, College of Engineering, De La Salle University, 2401 Taft Avenue, 1004 Manila, Philippines Department of Chemical Engineering Technology, School of Engineering Technology, Iligan Institute of Technology, Mindanao State University, Andres Bonifacio Avenue, Tibanga, 9200 Iligan City, Philippines c Department of International Development Engineering, Graduate School of Science and Engineering, Tokyo Institute of Technology, 2-12-1-I4-1, Ookayama, Meguro-ku, Tokyo 152-8550, Japan d Department of Inorganic Chemistry, Faculty of Chemical Engineering, Ho Chi Minh City University of Technology, 268 Ly Thuong Kiet Street, District 10, Ho Chi Minh City, Viet Nam b a r t i c l e i n f o Article history: Received 26 March 2011 Received in revised form July 2011 Accepted August 2011 Available online 27 August 2011 Keywords: Moringa oleifera seed oil Supercritical carbon dioxide Optimization Ethanol for pretreatment Response surface methodology a b s t r a c t The study involved the extraction of oil from Moringa oleifera (MO) kernels using supercritical fluid extraction (SFE) technique The arrays of operating parameters are extraction condition, loading configuration, and SC-CO2 with ethanol addition for substrate pretreatment It identifies a combination of operating parameters that provide a higher yield The experiments were conducted in the pressure range of 15–30 MPa, 35–60 ◦ C temperature interval, average particle size of 0.16–1.12 mm and CO2 flow rate of 0.5 m3 /h Adding 10% EtOH for substrate pretreatment, increased 10% in the yield No significant difference in yield and fatty acid content between the oil from SC-CO2 -EtOH and Soxhlet extraction with n-hexane was detected Loading the seeds in multiple-staged trays increased to as much as 26.89% in the yield against randomly packed configuration Response surface methodology, predicted an optimal oil yield of 37.84% at a pressure of 28.97 MPa, 44.30 ◦ C temperature, and particle size of 0.54 mm A crossover pressure point of Ben oil was found within the range 22.5–30 MPa © 2011 Elsevier B.V All rights reserved Introduction Moringa oleifera which has its origin in India is locally wellknown in the Philippines as Malunggay and is also popular in other countries in Asia, Africa, South America, the Caribbean and Oceania [1,2] The food derived from the plant is high in nutritional value [3,4] In more recent years, other uses of the plant cited medicinal applications and environmental applications [5–9] M oleifera seed oil, also known as Ben oil, has been reported to contain very high oleic oil (70% oleic acid) and smells a pleasant peanut-like fragrance [10,11] Oleic acid, a mono-unsaturated fatty acid, has strong oxidative stability when compared to polyunsaturated fatty acids essential for longer storage and at high temperature frying process Ben oil is more stable than canola oil, soybean oil, and palm olein when used in frying [12] Blending Ben oil with sunflower oil and soybean oil enhances the oxidative stability of the mixture [13] Furthermore, ingesting mono-unsaturated fatty acid reduces the risk of developing coronary heart disease ∗ Corresponding author Tel.: +63 5240563; fax: +63 5240563 E-mail address: ngochoang.ibft@gmail.com (H.N Nguyen) 0255-2701/$ – see front matter © 2011 Elsevier B.V All rights reserved doi:10.1016/j.cep.2011.08.006 as opposed to saturated and trans fatty acids [12,13] Extraction of oil from M oleifera seeds by cold pressing, aqueous enzymatic methods, or the traditional extraction process with the use of conventional solvent such as n-hexane, petroleum ether was already studied [10,14] However, non-solvent methods such as cold press method or enzyme extraction were reported to produce low oil yield While extraction using n-hexane or light petroleum ether propels high yield, the possible thermal degradation of the light components in the oil and the incomplete elimination of the toxic solvents are some of the drawbacks of the traditional extraction process Using supercritical carbon dioxide (SC-CO2 ) for extraction overcomes the disadvantages of the traditional solvent extraction SC-CO2 is a likely substitute to organic solvents because it is nontoxic, has flexible properties, and easy to separate from extracted oil by depressurization In addition, its density, solubility, diffusivity, and viscosity can be varied widely with the change in pressure and temperature [15–18] Successes in using SC-SO2 to extract oil from the seeds of sunflower, palm, hazel, jojoba, rape and peach were reported [19–24] However, no literature has been published yet regarding the extraction of Ben oil using SC-CO2 and supercritical carbon dioxide with ethanol addition for substrate pretreatment (SC-CO2 -EtOH) In this paper, the M oleifera seed oil extraction was 1208 H.N Nguyen et al / Chemical Engineering and Processing 50 (2011) 1207–1213 investigated using SC-CO2 and SC-CO2 -EtOH The study also examined the influence of modifying the loading of raw materials into the extraction vessel vis-à-vis the yield of the product The yield was compared in a multi-layered (similar to multiple stages packed bed) packing arrangement of the ground kernels to that of randomly charged (similar to random packed bed) configuration The extraction yield and the fatty acid component of Ben oil extracted using SC-CO2 , SC-CO2 -EtOH and n-hexane were compared Optimization of the process parameters was also included in the study Materials and methods 2.1 Material and sample preparation Air dried M oleifera seeds were sourced from Ilocos Norte and Ilocos Sur situated in the Northern part of the Philippine archipelago The seeds were decoated and the good quality kernels that were not moldy or breached were selected MO kernels were ground a few hours before extraction to minimize the effect of oxidation of the oil The ground kernels contained 8.7% moisture in weight The ground kernels were sieved to average particle sizes of 0.16 mm, 0.32 mm, 0.64 mm and 1.12 mm Commercial CO2 supplied by SUGECO (a local company) Analytical grade ethanol (99.9% v/v, EtOH), standard chemical for fatty acid analysis such as fatty acid methyl ester of myristic acid (C14:0 ), palmitic acid (C16:0 ), palmitoleic acid (C16:1 ), stearic acid (C18:0 ), oleic acid (C18:1 ), linoleic acid (C18:2 ), linolenic acid (C18:3 ), arachidic acid (C20:0 ), cis-11-eicosenoic acid (C20:1 ), behenic acid (C22:0 ), tetracosanoic acid (C24:0 ) and other chemicals such as n-hexane (GC analytical grade), methyl acetate (99.5%), acetic acid (99.7%), dehydrated diethyl ether (99.5%), sodium methoxide (0.5 M) in dry methanol, solid sodium were all supplied by Sigma–Aldrich, Inc and Wako Pure Chemical Industries, Ltd., Japan 2.2 Supercritical extraction methods The extraction using SC-CO2 was carried out in a pilot plant (fabricated in Akico, Japan) at the Department of Chemical Engineering Technology, Mindanao State University – Iligan Institute of Technology, Iligan City, Philippines Seventy grams of the ground kernels was loaded into a 500 mL-extractor by two methods The first method was randomly packed where the ground kernels were dumped into the extractor forming a layer The other method was loading the ground kernels arranged in multi-layers as illustrated in Fig 1a The number of layers and the thickness of each layer were varied as presented in the first two columns in Table The distance between layers was 10 mm The schematic illustration of the pilot plant is shown in Fig 1b The CO2 was first liquified before passing to a high pressure pump (with maximum capacity of 35 MPa) The CO2 liquid was then heated until it reached supercritical state In this work, the pressure and temperature of the SC-CO2 were varied from 15 to 30 MPa and from 35 to 60 ◦ C, respectively The supercritical CO2 flowed through the extractor and the SC-CO2 – oil solution from the extractor passed through the expansion valve where the oil was separated from CO2 The oil was collected in a vessel while the CO2 , passed through a rotameter at a fixed rate of 0.5 m3 /h (equivalent to 0.45 kg/h) before being released to the atmosphere In the SCCO2 -EtOH process, the EtOH was added directly to the extractor together with the ground kernels The substrate were soaked with EtOH in the actual processing environment for about 45 before extraction commenced which was conducted until no significant change in oil yield was observed to nearly h of continuous run [25] In this study, the percentage of EtOH is expressed as the initial weight of EtOH versus that of 500 mL CO2 at the processing condition 2.3 Solvent extraction Soxhlet extraction using n-hexane was conducted following the methods of analysis by the Association of Official Analytical Chemists (AOAC) [26] About g of MO ground kernels was packed in filter paper before loading in the Soxhlet extractor Extraction time was h The oil was obtained by evaporating the solvent at 100 ◦ C until a constant weight was attained The oil yield is a ratio of the weight of the oil with that of the MO kernel 2.4 GC analysis The analysis of fatty acid components of MO oil was conducted at Department of International Development Engineering, Graduate School of Science and Engineering, Tokyo Institute of Technology, Tokyo, Japan The oil was converted to fatty acid methyl esters using the method of Christie [27] Up to mg of oil was dissolved by mixing 0.5 mL sodium-dried diethyl ether, 20 L methyl acetate and 40 L sodium methoxide After min, the reaction was stopped by adding L acetic acid Nitrogen was used to evaporate the solvent and mL hexane was then added The solid precipitate was separated from the solution by using centrifuge Hitachi CRG Series R20A2 at 4000 r/min for The solution was injected into the capillary column SP-2560 (100 m × 0.25 mm ID, 0.25 m film) of Fig (a) The packing arrangement in the extractor unit (b) Flow scheme of the CO2 supercritical fluid extraction pilot plant (F: flow meter; H1: freezer; H2: heater; H3: heat exchanger; PI: pressure indicator; ST: stirrer; TI: temperature indicator; V1, V2, V3: gas valve) H.N Nguyen et al / Chemical Engineering and Processing 50 (2011) 1207–1213 Table Coded levels and real values of independent variables Factor Symbols coded Temperature (◦ C) Pressure (MPa) Particle size (mm) X1 X2 X3 Levels −1 35.00 15.00 0.16 47.50 22.50 0.64 60.00 30.00 1.12 the Gas Chromatograph GC-17A Shimadzu with helium as carrier gas Operating temperature program for column was set at 140 ◦ C for then increased to 240 ◦ C at ◦ C/min and withheld for 15 2.5 Experiment design for optimization of operating condition Response surface methodology was employed to optimize the extraction process using SC-CO2 with EtOH addition for substrate pretreatment The main parameters considered are pressure, temperature, particle size, EtOH concentration, solvent flow rate and extraction time EtOH concentration was fixed at 10% w/w as a consequence of the discussion in Section 3.2 The CO2 flow rate was fixed at 0.5 m3 /h because of the limitation of the apparatus which worked unstable at the flow rate higher than 0.5 m3 /h Consequently, extraction time was fixed at h in which no significant change in oil yield was observed In this study, the optimization of the extraction process was based from three parameters: pressure, temperature and particle size In Table 1, X1 , X2 and X3 represent three independent variables: temperature, pressure and average particle size of MO ground kernels, respectively Temperature was varied from 35 ◦ C to 60 ◦ C Pressure range was from 15 MPa to 30 MPa while the average particle size was from 0.16 mm to 1.12 mm The estimated range of the three factors was divided into three levels Coded level is the midpoint of each factor range Coded levels −1 and correspond to the lowest value and the highest value of the experimental parameter, respectively A central composite design method was used to design the experiment Table shows all the twenty experiments which included eight factorial points, four axial points and one center point Factorial points have all possible combinations of the +1 and −1 levels of the three factors The axial points have all of the factors set to except one factor, which has the value +/− Alpha Alpha, the axial distance, has a coded value for Face Centered type Center point has all Table Central composite design for experiment and results of oil yield Runs Coded variables X1 10 11 12 13 14 15 16 17 18 19 20 −1 −1 −1 0 −1 0 1 0 −1 1209 levels set to coded level The center point was repeated six times to estimate experimental error while other points were duplicated All experiments were conducted at CO2 flow rate of 0.5 m3 /h with 10% by weight EtOH and used multi-layer packed bed with layers Results and discussion 3.1 Effect of the number of layers and the thickness of material layers on oil yield Table shows the oil yield of SC-CO2 -EtOH using random packing and multiple layers packing arrangement The results for random packing method as presented are equivalent to a layer with 100 mm thickness The remainder belongs to multiple layers packed bed All runs were triplicated at a pressure of 30 MPa, temperature of 47.5 ◦ C, with average particle size of 0.32 mm A 10% by weight EtOH was added for substrate pretreatment Significant increase in oil yield was observed with a maximum of 26.89%, when multiple-layered packing method was employed This result agrees with those of Rubio-Rodriguez et al [28] on SFE of omega-3 rich oil in hake by-products However, their study did not include the effect of neither thickness of sample layer nor number of layers on extraction yield Table shows that decreasing the thickness of the sample layer but increasing the number of layers of the packed bed enhanced the extraction yield The multi-layer packed bed has prevented the formation of preferential channels thus allowing the SC-CO2 to be distributed uniformly in the extractor It allowed the solvent to penetrate covering wide surface areas of the ground kernels thus promoting better extraction of the oil [28] The gaps between layers are akin to the liquid re-distributors in multiple-staged packed bed absorption column purposely employed to constantly wet the surface of the packings It was noted, however, that when the amount of the ground kernels was increased as exhibited by its increasing thickness in a layer, the extraction of oil slowed down as exhibited by the corresponding reduction in oil yield [29] The reverse is true when the thickness of the kernels in the tray layer was reduced It can be concluded that the thinner the thickness of the kernels in a layer at multiple stages, the less resistance of mass transfer, thus increasing the yield However, the operating efficiency of the extractor system for multiple stages as a whole was low obviously because of the gaps in between layers In a configuration of ten layers, the operating efficiency exhibited is only 66.7% far from the acceptable value of 75% 3.2 Effect of ethanol ratio on oil yield Oil yield X2 −1 1 −1 0 0 0 −1 −1 −1 0 X3 −1 −1 1 0 0 0 −1 −1 −1 1 22.41 36.13 34.55 37.82 21.64 33.13 32.78 34.62 32.92 27.71 32.65 16.93 32.58 8.23 6.71 35.20 31.90 32.59 35.67 33.09 Co-solvent EtOH in SC-CO2 enhanced extraction yield was reported by several authors Casas et al [30] and Lee et al [31], have stated that ethanol have swollen the cellular structure thus facilitating the SC-CO2 penetration Others such as Guclu-Ustundag Table Oil yield of SFE using random packing and multiple stages packed bed (extraction conditions were at pressure of 30 MPa, temperature of 47.5 ◦ C, average particle size of 0.32 mm, CO2 flow rate of 0.5 m3 /h and 10% by weight EtOH) Number of layers 10 Thickness of a layer (mm) Oil yield (% wt.) 10 20 25 50 100 40.01 38.01 37.25 34.49 29.24 ± ± ± ± ± 0.71 0.57 0.72 0.78 0.97 Recoverya (%) Working efficiency of extractor (%) 100.0 95.0 93.1 86.2 73.1 66.7 71.4 83.3 90.9 100.0 a The recovery is the ratio of oil yield which is extracted in each experiment in comparison with the highest oil yield (40.01%) 1210 H.N Nguyen et al / Chemical Engineering and Processing 50 (2011) 1207–1213 Table Oil yield with varying initial EtOH ratio (extraction conditions were at pressure of 22.5 MPa, temperature of 47.5 ◦ C, average particle size of 1.12 mm, CO2 flow rate of 0.5 m3 /h) With co-solvent 0% EtOH 10% EtOH 15% EtOH Oil yield 28.71 ± 0.67 31.90 ± 0.51 31.74 ± 0.82 Table Analysis of variance of three polynomial models Type of model F Pa Lack of fit-Pa Linear model Quadratic model Cubic model 19.02 1298.87 2.29