Study on enzyme-assisted aqueous extraction of oil from soybean

Transcription

1 8 Journl Scientific & Industril Reserch J SCI IND RES VOL 69 NOVEMBER 2010 Vol. 69, November 2010, pp Study on enzyme-ssisted queous extrction oil from soyben Jun-Qing Qin*, De-Hui Qin, Xing-Mo Xie nd Wen-Wu Zhou College Phrmceuticl Science, Zhejing University Technology, Hngzhou , Chin Received 20 My 2010; revised 03 August 2010; ccepted 06 August 2010 This study presents enzyme-ssisted queous extrction (EAAE) method for oil extrction from soyben. Prmeters for EAAE oil with 1398 neutrl protese were optimized through single-fctor nd orthogonl test. Soyben oil (yield, 80.2% w/ w) ws chieved under following optimized conditions: enzyme (conc., 840 IU / g soyben; ph, 7.0); enzymtic hydrolysis (temp., 45 C; time, 1.5 h); rtio petroleum ether to soyben oil, 4:1(v/w); extrction (temp., 30 C; time, 15 min; ph, 4.5); nd centrifugtion force, 1400 g. Keywords: Aqueous Extrction Enzymtic hydrolysis Orthogonl test, Soyben Oil Introduction Aqueous extrction method hs been successfully pplied to soybens, sunflower seeds, penuts nd coconut oils 1,2. Oil cn be seprted from queous phse fter breking emulsion 3-5. Until now, high cost enzymtic processing nd difficulties in queous seprtion fter enzymtic hydrolysis is involved 6. Industrilized enzyme with low price is the bsis enzyme-ssisted extrction process This study presents enzyme-ssisted queous extrction (EAAE) method for oil extrction from soyben by using chep enzyme to reduce cost processing, dopting simple method for two-phse oil-wter seprtion without using high-speed centrifugtion, nd mintining degrees hydrolyztion trnsformtion. Experimentl Section Mterils Soyben (moisture, 12.4; oil, 18.6; nd crude protein 40.5 % w/w) ws obtined from locl mrket. Neutrl protese (EC , B. subtilis) (csein s substrte) ws provided by Wuxi Enzymes Preprtion Fctory (Chin). Other chemicls were nlyticl grde. Preprtion Soyben Suspension Rw soyben (150 g) ws soked in distilled wter Author for correspondence Tel: ; Fx: E-mil: qjq@zjut.edu.cn (0 ml, ph 9.0.) t room temperture (RT) for 1 h, nd then grinded fter dding two times volume wter. Soyben suspension ws filtered with sieve (100 mesh). Filter residue (4.5% w/w totl oil) ws wshed with two times volume wter. Filtrte ws obtined for further nlysis. Enzyme-Assisted Aqueous Extrction (EAAE) Oil from Soyben Suspension Soyben suspension ws hydrolyzed with 1398 neutrl protese t suitble ph through controlling temperture nd enzymtic hydrolysis (EH), which ws stopped through djusting solution ph to inctivte enzyme. Oil ws extrcted from queous phse by centrifuging two-phse with orgnic solvent. In order to reduce opertion error, sme group soyben suspension ws divided into numbers different smples; ech hving 450 ml suspension. For fixing oil extrction process conditions, medium ws extrcted t 25 C nd ph 4.5 for 20 min under continuous stirring followed by centrifugl seprtion t 0 g for 15 min. Then, totl system splitted into four lyers (oil-solvent mixing phse, protein-orgnic creming lyer, queous phse nd solid lyer) from top to bottom. Soyben oil ws obtined fter evportion nd solvent ws recovered from oil-solvent mixing phse. Amount oil nd wter ws determined by reported method 11. All nlyses were conducted three times

2 QIAN et l : ENZYME-ASSISTED EXTRACTION OF OIL 861 Totl yield extrction oil, % Totl yield extrction oil / % A Stirring, r/min Mss extrction oil / g Totl Totl yield yield extrction extrction oil / % oil, % B 10 8 Mss 6 4 extrction oil / 2g The mount enzyme enzyme, / ( IU/g IU/g soyben) soyben Totl Totl yield extrction oil oil, / % % C ph Totl yield extrction oil, % D Time enzyme hydrolysis, h Totl yield extrction oil, % 62 E o Temperture, o C - -, Totl yield extrction oil; - -, Mss extrction oil Fig.1 Effect on extrction oil by enzymtic hydrolysis : A) Stirring (soyben, 50 g; enzyme conc., 480 IU/g soyben; ph, 7.0; Temp., 45 C; time, 1 h); B) Amount enzyme (soyben, 50 g; ph, 7.0; Temp., 45 C; time, 1 h); C) ph (soyben, 50 g; enzyme conc., 480 IU/g soyben; Temp., 45 C; time, 1 h); D) Time (soyben, 50 g; enzyme conc., 480 IU/g soyben; ph, 7.0; Temp., 45 C); nd E) Temperture (soyben, 50 g; enzyme conc., 480 IU/g soyben; ph, 7.0; time, 1 h) under sme experimentl conditions. 9.5 gi /100 g. All qulity indictors were excellent s compred to solvent extrction method. Results nd Discussion Qulity Extrcted Oil from Enzyme-Assisted Aqueous Extrction (EAAE) Method Chrcteristics extrcted oil were found s follows: impurities, 0.13% w/w; cid vlue, 2.9 mg KOH/ kg; peroxide vlue, 15 meq /kg; nd iodine vlue, Enzymtic Hydrolysis for Extrction Oil from Soyben Suspension Under EH (Fig. 1A), continuous stirring increses mss trnsfer, resulting in good extrction rte oil [mss extrcted oil, 4.85 g (yield, 52.3% w/w)]. But stirring rte should be controlled t 300 rpm. Without using

3 862 J SCI IND RES VOL 69 NOVEMBER 2010 Totl yield extrction oil, % Totl yield extrction oil oil, / % % Totl yield extrction oil, % A C E Temperture, o C Time extrction/min / B D ph milk Amount Petroleum ether ech 100g rw mteril / ml Size Size centrifugtion, centrifugtion / g g Time centrifugtion, g - -, Totl yield extrction oil; - -, Mss extrction oil Totl yield extrction oil, % Totl yield extrction oil, % Totl Totl yield yield extrction extrction oil / % oil, % F Mss extrction oil / g Mss extrction oil / g Mss extrction oil / g Fig. 2 Effect on extrction oil : A) Temperture (soyben, 50 g; pet ether vol, 100 ml; ph, 4.5; time, 15 min); B) ph (soyben, 50 g; pet ether vol, 100 ml; temp., 35 C; time, 20 min); C) Extrction time (soyben, 50 g; pet ether vol, 100 ml; ph, 4.5; temp., 35 C); D) Amount solvent (soyben, 50 g; ph, 4.5; temp., 35 C; time, 20 min); E) Rte centrifugtion (soyben, 50 g; pet ether vol, 100 ml; ph, 4.5; extrction time, 15 min; temp., 35 C; centrifugtion time, 15 min); nd F) Time centrifugtion (soyben, 50 g; pet ether vol, 100 ml; ph, 4.5; extrction time, 15 min; temp., 35 C; centrifugtion size, 0 g) enzyme (Fig. 1B), extrction rte oil ws very low, only 1.59% w/w oil soyben suspension ws relesed. Extrction rte oil significntly incresed, with incresing mount enzyme until 0 IU /g soyben. EH t ph (Fig. 1C) gve high extrction oil. Extrction yield oil ws relted to combined stte oil molecule nd protein molecule in suspension. Extrction yield oil incresed with incresing time EH nd reched mximum t 2 h, nd then decresed between h (Fig. 1D). Optimized temperture EH ws 45 C (Fig. 1E). Thus optimized conditions EH (Fig. 1) were: enzyme, 0

4 QIAN et l : ENZYME-ASSISTED EXTRACTION OF OIL 863 Tble 1A Orthogonl test enzymtic hydrolysis conditions (soyben, 50 g; pet ether, 100 ml; temp., 25 C; ph, 4.5; nd extrction time, 20 min) Run No. Amount enzyme Temperture ph Time Oil yield IU/g soyben C h % A B C D R Degree influence: R B>C>A>D R , R 2 nd R 3 indicte level fctors corresponding to totl oil yield; R indictes poor Tble 1B Anlysis vrince components orthogonl test results in Tble 1A Sources Sum Degree Squres sum F Significnce vrince squres freedom verge devition A b S A = B S B = * C S C = Error (D) S E = Totl 122. F 0.05 (2, 2) =19.0; F 0.10 (2, 2) =9.0; b A, B nd C indicte fctors IU/g soyben; temperture, 45 C; ph, 7.5; stirring, under 300 rpm; nd rection time, 2 h. Under optimized conditions, oil yield from soyben suspension ws 75.8 % w/w, which is.0% w/w totl oil soyben. Optimiztion Enzyme-Assisted Extrction Oil from Soyben Suspension After selection EH conditions, oil extrction conditions (extrction temperture, medium ph, extrction time, mount orgnic solvent, centrifugtion force nd centrifugtion time on extrction oil) were optimized (Fig. 2). Optimum temperture for extrction oil ws found to be 35 C (Fig. 2A). Serious creming could hppen between solution nd orgnic solvent t higher temperture, which ws not beneficil to phse seprtion. And so, orgnic solvent ws esily voltilized t higher temperture. Optiml suspension ph (fter EH for extrction oil) ws 4.5, due to be close to isoelectric point soyben protein (Fig. 2B). Optiml time extrction oil ws found to be 20 min (Fig. 2C). Extrction yield oil decresed with longer extrction time. Extrction yield oil incresed with incresing mount orgnic solvent until petroleum ether (300 ml/ 100 g suspension) for extrction oil. After tht, it nerly unchnged even fter incresing mount orgnic solvent (Fig. 2D). Entire system looks like ltex fter being extrction with petroleum ether nd must be

5 8 J SCI IND RES VOL 69 NOVEMBER 2010 Tble 2A Orthogonl test extrction oil conditions Run No. Amount pet Temperture Time Oil yield ether / ml C h ph % A B C D b R Degree influence: R A>C>B>D R Amount pet ether is volume pet ether dded to emulsion (900 ml) from 100 g soyben for extrction oil b, R 2 nd R 3 indicte level fctors corresponding to totl oil yield; R indictes poor Tble 2B Anlysis vrince components orthogonl test results in Tble 2A Sources Sum Degree Squres sum F Significnce vrince squres freedom verge devition A b S A = * B S B = C S C = Error (D) S E = Totl F 0.05 (2, 2) =19.0; F 0.10 (2, 2) =9.0; b A, B nd C indicte fctors seprted by centrifugtion. For present system, low centrifugtion force could mke phse seprtion well. Similr results were obtined using different centrifugtion force (350 g g). Centrifugtion force 1400 g ws chosen for two-phse seprtion (Fig. 2E). Optiml extrction yield ws obtined fter centrifugtion for 10 min (Fig. 2F). Thus optimized extrction conditions for EAAE oil were 300 ml petroleum ether /100 g soyben t 35 C nd ph 4.5 for 20 min, nd 1400 g centrifugtion force for 10 min. Averge totl yield extrcted oil ws 71.5 % (w/w) under optimized conditions both EH nd extrction oil. However it ws lower thn expected, becuse EH nd extrction occurred in complicted system. Thus, it ws necessry to optimize conditions EH nd oil extrction by using orthogonl test (OT). Orthogonl Test (OT) OT Enzymtic Hydrolysis Conditions Influencing fctors (mount enzyme, temperture nd time EH, nd ph medium) were selected for OT (Tble 1A) nd test results on EH were obtined with Anlysis vrince (ANOVA) (Tble 1B). Temperture ws found highly significnt fctor (Tble 1). Effect time on EH ws wekest ll fctors (Tble 1). Optimized conditions EH were found s follow: enzyme, 840 IU/g soyben; temperture, 45 C; ph, 7.0; nd time, 1.5 h. OT Extrction Oil Conditions

6 QIAN et l : ENZYME-ASSISTED EXTRACTION OF OIL 865 Similr to OT EH conditions, influencing fctors (mount enzyme, temperture nd time EH, nd ph medium) were selected for OT (Tble 2A) for oil extrction conditions nd test results were obtined with ANOVA (Tble 2B). Amount petroleum ether ws highly significnt fctor (Tble 2). Effect medium ph on EH ws wekest ll fctors (Tble 2A). Optimized conditions extrction oil were s follows: extrction oil with petroleum ether, 400 ml / 100 g soyben; temperture, 30 C; ph, 5.0; nd time, 15 min. Averge totl extrction yield (80.2%, w/w) totl oil soyben ws chieved under optimized conditions EH nd extrction oil. Conclusions Low cost enzyme-ssisted queous extrction oil from soyben hs been found techniclly vible process. Chep 1398 neutrl protese ws used for enzymtic hydrolysis soyben suspension. Enzymtic hydrolysis significntly incresed extrction rte oil from soyben suspension. Process prmeters were optimized using single-fctor test nd orthogonl test. Totl soyben oil (v 80.2% w/w) ws chieved under optimum conditions. This environment friendly process hs potentil to be utilized in oil industry. Acknowledgement Authors thnk Wuxi Enzymes Preprtion Fctory (Chin) for supply 1398 neutrl protese. References 1 Chbrnd R M & Gltz C E, Destbiliztion the emulsion formed during the enzyme-ssisted queous extrction oil from soyben flour Enzyme Microb Tech, 45 (2009) Jing L, Aqueous enzymtic extrction penut oil nd protein hydrolystes, FBP, 8 (2009) Kwku T D & Yoshiyuki O, Enzyme-ssisted queous extrction sheft: A rurl pproch, AOCS, (1995) Che Mn Y B & Subrdiyono A B, Aqueous enzymtic extrction coconut oil, AOCS, 73 (1996) Sengupt R & Bhttchryy D K, Enzymtic extrction mustrd seed rice brn, AOCS, 6 (1996) Wng Z X, Shi y & Lin L etc, Study on the Enzymtic Process Simultneously Prepring Soy Oil nd Soy Protein Hydrolyste from Full Ft Soy Ben, J WuXi Univ Light Ind, 13 (1994) Li B S, Peng Z Y & Song Z X, Enzymtic pretretment for oil extrction from oil plnts, J Chinese Cerels Oils Assoc, 12 (1997) Dominguez H, Nunez M J & Lem J M, Enzymtic pretretment to enhnce oil extrction from fruits nd oil seed : review, J Food Chem, 49 (1994) Mour J M L N & Johnson L A, Two-Stge Countercurrent enzyme-ssisted queous extrction processing oil nd protein from soybens, J Am Oil Chem Soc, 86 (2009) Virginie N, Enzyme-ssisted queous extrction oleosomes from soybens (Glycine mx), J. Agric. Food Chem, 56 (2008) Wuxi University Light Industry & Tinjin University Light Industry, Food Anlysis (Light Industry Press, Bei Jing) 1983, -225.