Articles

FT-Raman Spectroscopy Quantification of Biodiesel in a ProgressiveSoybean Oil Transesterification Reaction and Its Correlation with

1H NMR Spectroscopy Methods

Grace Ferreira Ghesti,* Julio Lemos de Macedo, Ineˆs Sabioni Resck, Jose´ Alves Dias, andSılvia Claudia Loureiro Dias*

Laboratorio de Catalise, Instituto de Quı´mica, UniVersidade de Brası´lia, caixa postal 4478,Brasılia-DF, 70904-970, Brazil

ReceiVed December 29, 2006. ReVised Manuscript ReceiVed May 29, 2007

Biodiesel fuel (fatty acid esters) has become more and more attractive due to its environmental benefits.Transesterification is the most common and important method for making biodiesel from vegetable oils oranimal fats. Several studies have focused on the development and improvement of analytical methods formonitoring biodiesel production and determining the fuel quality. Analytical procedures reported in the literatureinclude chromatographic methods (e.g., gas chromatography, high-performance liquid chromatography, gelpermeation chromatography, etc.) and spectroscopic methods [e.g.,1H and13C NMR, near infrared, Fouriertransform infrared spectroscopy, and recently, Fourier transform (FT)-Raman]. The study presented in thispaper expands our previous research, in which FT-Raman spectroscopy combined with partial least squares(PLS) multivariate analysis was successfully applied to the quantification of soybean oil/ethyl ester mixtures.The FT-Raman/PLS methods developed by our group were used to monitor and quantify a transesterificationreaction process involving soybean oil and ethanol to produce fatty acid ethyl esters (biodiesel) over 22 hcatalyzed by a heterogeneous Lewis acid catalyst. The results were successfully correlated with two1H NMRspectroscopic methods reported in the literature and a new1H NMR method proposed in this work that can beeasily extended to other vegetable oils. The correlation coefficients (R2) obtained from the linear fit betweenFT-Raman measurements and the above1H NMR methods were 0.9974, 0.9847, and 0.9972, respectively.

1. Introduction

Biodiesel is defined by the American Society for Testing andMaterials (ASTM) as a fuel comprised of monoalkyl esters oflong-chain fatty acids derived from vegetable oils or animalfats meeting the requirements of ASTM D 6751.1,2 Biodieselhas distinct advantages when compared to petroleum-deriveddiesel fuel (petrodiesel). It is derived from renewable resources;is biodegradable; is nontoxic; has low emission profiles, a higher

flash point, and excellent lubricity; and can be used either pureor blended with petrodiesel fuel.1,3,4

The use of vegetable oils as fuel has been known since theParis Exposition in 1900.4 However, due to its higher molecularmass and kinematic viscosity, its direct use in diesel enginesresulted in several operational problems (e.g., poor atomization,carbon deposits due to incomplete combustion, oil ring sticking,lubricating problems, etc.).1,4,5 To solve these problems, fourpossible solutions were investigated in literature: transesteri-

* Corresponding authors. Phone: 55-(61)-3307-2162. Fax: 55-(61)-3368-6901. E-mail: scdias@unb.br (S.C.L.D.) and grace@unb.br (G.F.G.).

(1) Ma, F.; Hanna, M. A.Bioresour. Technol.1999, 70, 1-15.(2) ASTM D 6751-03a.Annu. Book ASTM Stand.2005, 05.04, 609-

614.

(3) Knothe, G.J. Am. Oil Chem. Soc.1999, 76, 795-800.(4) The Biodiesel Handbook; Knothe, G., Gerpen, J. V., Krahl, J., Eds.;

American Oil Chemists’ Society Press: Champaign, IL, 2005.(5) Meher, L. C.; Sagar, D. V.; Naik, S. N.Renew. Sustain. Energy ReV.

2006, 10, 248-268.

VOLUME 21, NUMBER 5 SEPTEMBER/OCTOBER 2007

© Copyright 2007 American Chemical Society

10.1021/ef060657r CCC: $37.00 © 2007 American Chemical SocietyPublished on Web 07/17/2007

fication (also called alcoholysis), pyrolysis, dilution with con-ventional petroleum-derived diesel fuel, and microemulsifi-cation.1,4-12

The transesterification reaction is the only method that leadsto products defined as biodiesel (i.e., alkyl esters) and is by farthe most usual method to make biodiesel.1,4 In transesterification,the triacylglyceride (TAG) molecules found in animal fats orvegetable oils (e.g., soybean,12-14 peanut,12 rapeseed,1 etc.)reacted with an alcohol (e.g., methanol,12 ethanol,12-14 etc.) inthe presence of a catalyst (e.g., NaOH,12-14 H2SO4,12 lipase,11

etc.) to form esters and glycerol (Figure 1).1,3-5,10,12In additionto the advantages previously mentioned, biodiesel fuel producedby transesterification also has similar properties to petroleum-derived diesel fuel (e.g., cetane number, viscosity, molecularmass, density, etc.),15-18 and no diesel engine modifications arerequired.19

In Brazil, ethanol is a less expensive alcohol than methanoldue to its high production volume from biomass sugarcane. Inthis sense, the biodiesel produced from ethanol (fatty acid ethylesters or FAEE) can be seen as a truly renewable fuel. In 2003,the Brazil Federal Government started its own biodiesel pro-grams (PROBIODIESEL and PNPB), which intend to stimulatethe scientific and technologic development of biodiesel and toimplement the sustainable production and use of biodiesel,respectively, through the gradual increase of renewable resourcesin the Brazilian energetic matrix.20 The development andoptimization methods for biodiesel production in Brazil and inother countries have motivated a great number of publicationsand patents.21 The analytical procedures reported in the literaturefor the determination of fuel quality and the monitoring of

biodiesel production22 include chromatographic methods [e.g.,gas chromatography (GC),23 high-performance liquid chroma-tography,24 gel permeation chromatography (GPC),22,25etc.] andspectroscopic methods [e.g., nuclear magnetic resonance(NMR),14,26,27near-infrared (NIR),3,27Fourier transform infraredspectroscopy (FTIR),13,25 and recently, FT-Raman spectros-copy28].

Spectroscopic techniques are fast, easily adapted in routineprocess analysis, and allow nondestructive measurements of thesamples22,29versus time-consuming chromatographic methods.NMR spectroscopy has become one of the most powerfultechniques to investigate and identify the structure of chemicalcompounds and dynamics of molecular systems in almost allbranches of chemistry.30,31 Although NMR spectroscopy doesnot present the detection limit accuracy for the quantificationof minor components as chromatographic methods do, NMR isa suitable method to monitor a chemical reaction, since smallamounts are required to obtain a quantitative spectrum withsignificant information related to the substances of interest inthe reaction media.32

Alternatively, the employment of vibrational spectroscopictechniques (NIR, FTIR, and Raman) in quality-control monitor-ing has been growing quickly due to several qualities (e.g., fastmeasurements, easy handling, accuracy, reliability, possibilityof on-line monitoring with fiber-optic probes, etc.).22,33 Forexample, NIR spectroscopy is being used to analyze free fattyacid contents in oils,34,35 and Raman spectroscopy has beenwidely used in the pharmaceutical36 and polymer industries.37

Uni- and multivariate regression analyses have been widely usedto develop calibration models based on vibrational spectroscopicdata.38-40 Actually, all reports in the literature until nowdescribing vibrational spectroscopy methods to monitor andquantify biodiesel fuel production were based on regressionanalysis.3,13,27

Recently, our group reported the advantages of FT-Ramanspectroscopy to quantify the concentration of ethyl esters in

(6) Schwab, A. W.; Bagby, M. O.; Freedman, B.Fuel 1987, 66, 1372-1378.

(7) Dasari, M. A.; Goff, M. J.; Suppes, G.J. Am. Oil Chem. Soc.2003,80, 189-192.

(8) Schwab, A. W.; Dykstra, G. J.; Selke, E.; Sorenson, S. C.; Pryde, E.H. J. Am. Oil Chem. Soc.1988, 65, 1781-1786.

(9) Bagby, M. O.; Freedman, B.; Schwab, A. W.Seed Oils for DieselFuels: Sources and Properties; ASAE Paper No. 87-1583; AmericanSociety of Agricultural Engineers: St. Joseph, MI, Dec 1987.

(10) Schuchardt, U.; Sercheli, R.; Vargas, R. M.J. Braz. Chem. Soc.1998, 9, 199-210.

(11) Jackson, M. A.; King, J. W.J. Am. Oil Chem. Soc.1996, 73, 353-356.

(12) Freedman, B.; Pryde, E. H.; Mounts, T. L.J. Am. Oil Chem. Soc.1984, 61, 1638-1643.

(13) Zagonel, G. F.; Peralta-Zamora, P.; Ramos, L. P.Talanta 2004,63, 1021-1025.

(14) Neto, P. R. C.; Caro, M. S. B.; Mazzuco, L. M.; Nascimento, M.G. J. Am. Oil Chem. Soc.2004, 81, 1111-1114.

(15) Demirbas¸, A. Energy ConVers. Manage.2002, 43, 2349-2356.(16) Fukuda, H.; Kondo, A.; Noda, H.J. Biosci. Bioeng.2001, 92, 405-

416.(17) Pryde, E. H. J.Am. Oil Chem. Soc.1984, 61, 1609-1610.(18) Barnwal, B. K.; Sharma, M. P.Renew. Sustain. Energy ReV. 2005,

9, 363-378.(19) Saka, S.; Kusdiana, D.Fuel 2001, 80, 225-231.(20) Programa Nacional de Produc¸ ao e Uso de Biodiesel Home Page.

http://www.biodiesel.gov.br (accessed May 2007).(21) Pinto, A. C.; Guarieiro, L. L. N.; Rezende, M. J. C.; Ribeiro, N.

M.; Torres, E. A.; Lopes, W. A.; Pereira, P. A. P.; Andrade, J. B.J. Braz.Chem. Soc.2005, 16, 1313-1330.

(22) Knothe, G.Trans. ASAE2001, 44, 193-200.(23) Plank, C.; Lorbeer, E.J. Chromatogr., A1995, 697, 461-468.(24) Holcapek, M.; Jandera, P.; Fischer, J.; Prokesˇ, B. J. Chromatogr.,

A. 1999, 858, 13-31.(25) Dube, M. A.; Zheng, S.; McLean, D. D.; Kates, M.J. Am. Oil Chem.

Soc.2004, 81, 599-603.(26) Geldard, G.; Bre´s, O.; Vargas, R. M.; Vielfaure, F.; Schuchardt, U.

F. J. Am. Oil Chem. Soc.1995, 72, 1239-1241.(27) Knothe, G.J. Am. Oil Chem. Soc.2000, 77, 489-493.(28) Ghesti, G. F.; Macedo, J. L.; Braga, V. S.; Souza, A. T. C. P.;

Parente, V. C. I.; Figuereˆdo, E. S.; Resck, I. S.; Dias, J. A.; Dias, S. C. L.J. Am. Oil Chem. Soc.2006, 83, 597-601.

(29) Drago, R. S.Physical Methods for Chemists, 2nd ed.; SaundersCollege Publishing: New York, 1992; pp 162-192.

(30) Silverstein, R. M.; Bassler, G. C.; Morril, T. C.SpectrometricIdentification of Organic Compounds, 7th ed.; John Wiley & Sons: NewYork, 2005; pp 165-202.

(31) Engelhardt, G.; Michel, D.High-Resolution Solid-State NMR ofSilicates and Zeolites; John Wiley & Sons: New York, 1987; p 1.

(32) Morgenstern, M.; Cline, J.; Meyer, S.; Cataldo, S.Energy Fuels2006, 20, 1350-1353.

(33) Cooper, J. B.; Wise, K. L.; Jensen, B. J.Anal. Chem.1997, 69,1973-1978.

(34) Sato, T.Biosci. Biotechnol. Biochem.2002, 66, 2543-2548.(35) Zhang, H.-Z.; Lee, T.-C.J. Agric. Food Chem.1997, 45, 3515-

3521.(36) Vankeirsbilck, T.; Vercauteren, A.; Baeyens, W.; Van der Weken,

G. TrAC, Trends Anal. Chem.2002, 21, 869-877.(37) Chalmers, J. M.; Everall, N. J.TrAC, Trends Anal. Chem.1996,

15, 18-25.(38) Cooper, J. B.; Wise, K. L.; Groves, J.; Welch, W. T.Anal. Chem.

1995, 67, 4096-4100.(39) Yu, Z.; Ma, C. Y.; Yuen, S. N.; Phillips, D. L.Food Chem.2004,

87, 477-481.(40) Ampiah-Bonney, R. J.; Walmsley, A. D.Analyst1999, 124, 1817-

1821.

Figure 1. Scheme of a generic transesterification reaction betweenone triacylglyceride (TAG) molecule and three ethanol (EtOH)molecules (stoichiometric relation) to form three molecules of ethylesters (FAEE or biodiesel) and one glycerol (GLY) molecule.

2476 Energy & Fuels, Vol. 21, No. 5, 2007 Ghesti et al.

known standard mixtures containing soybean oil.28 The presentpaper extends that work, and FT-Raman/partial-least-squares(PLS) calibration models were used to monitor and quantify aprogressive transesterification reaction with soybean oil andethanol. The results obtained were correlated with three1H NMRspectroscopic methods, two of them already reported by otherauthors14,41 and a new approach reported here.

2. Experimental Section

2.1. Materials. Commercial refined soybean oil (Soya), NaOH(Vetec, 99%), NaCl (Vetec, 99.5%), and concentrated HCl (Vetec,38%) were used as received. Ethanol (Vetec, 99.8%) was driedover 3A molecular sieves (Aldrich) for at least 24 h before theexperiments, and MgSO4‚7H2O (Vetec, 98.0%) was dried at 300°C for 4 h.

2.2. Preparation of the Heterogeneous Catalyst.Cerium tris-dodecylsulfate was used as a heterogeneous Lewis acid catalystfor the transesterification reaction. The synthesis, characterization,and modification have been described elsewhere.42,43For all reactionprocedures, the catalyst was activated in a muffle furnace at 100°C for 4 h.

2.3. Preparation of Biodiesel and Soybean Oil/BiodieselStandard Mixtures. Ethyl esters were prepared by transesterifi-cation according to conditions suggested in the literature12 for base-catalyzed reactions. The reaction was performed in a 50 mL glassround-bottom flask containing 20.00 g of soybean oil, 6.34 g ofanhydrous ethanol (1:6 oil-to-alcohol mole ratio), and 0.20 g ofNaOH (1% w/w of oil). The system was kept stirring at 80°Cunder reflux for 90 min. Then, the product was cooled to roomtemperature, washed several times with consecutive aqueoussolutions of HCl (0.5 wt %) and NaCl (5 wt %), and dried overanhydrous magnesium sulfate and residual alcohol was removedin a rotary evaporator at 70°C. Molecular weights of biodieseland soybean oil were calculated using the composition of fatty acidsobtained from the literature.44 The quality of the product wasverified by NMR (section 2.6) and GC (GC-17A Shimadzuchromatograph with a flame ionization detector and polydimeth-ylsiloxane column, CBPI PONA-M50-042). Since no significantcontaminants were observed, the biodiesel was treated as 100%ethyl ester. Six soybean oil/ethyl ester samples (0:100, 20:80, 40:60, 60:40, 80:20, and 100:0%) were prepared by weighing thefeedstock (soybean oil) and the ethyl ester standards produced(biodiesel).

2.4. Transesterification Reaction.The heterogeneous transes-terifications were carried out in 50 mL glass round-bottom flaskskept under stirring at 100°C under reflux conditions. A total ofsix reactions (2, 6, 10, 14, 18, and 22 h) were made under identicalconditions in order to reproduce a continuous 22 h reaction. Foreach run, 10.00 g of soybean oil, 15.85 g of anhydrous ethanol(1:30 oil-to-alcohol mole ratio), and 1.00 g of the catalyst (10%w/w of oil) were used. After the reaction, the system was cooledto room temperature, centrifuged to remove the catalyst, washedthree times with a 5 wt %NaCl solution, and dried over anhydrousmagnesium sulfate; residual alcohol was removed in a rotaryevaporator at 70°C; and then the system was kept in a refrigerator

for FT-Raman and NMR analyses. The reaction conditions usedabove were determined to be ideal for this catalyst according toGhesti.45

2.5. FT-Raman/PLS Analysis.FT-Raman spectra were recordedon a Bruker FRA 106/S module attached to a Bruker Equinox 55spectrometer using a 1 cmquartz cuvette with a mirrored surfacetoward the scattering direction (128 scans and 4 cm-1 resolution).The laser excitation (Nd:YAG) and laser power were 1064 nm and250 mW, respectively, and the signal was detected by a liquid N2

cooled Ge detector. All spectra were recorded at room temperature.The complete procedure of the method developed and theoreticalexplanations were described in a previous publication.28 Formultivariate analysis, PLS-1 (OPUS-NT Quant software, fromBruker) methods were used, also as described previously.28

2.6. 1H NMR Measurements. NMR experiments were per-formed at 7.05 T using a Varian Mercury Plus NMR spectrometerequipped with 5 mm Varian probes (ATB and SW) using CDCl3

as solvent.1H (300 MHz) spectra were recorded with a pulseduration of 45°, a recycle delay of 1.36 s and 16 scans. The spectrawere referenced to tetramethylsilane (δ ) 0.0 ppm).13C (75.46MHz) spectra were recorded with a pulse duration of 45°, a recycledelay of 0.28 s and 300 scans. The spectra were referenced to CDCl3

(δ ) 77.0 ppm). The two-dimensional experiment heteronuclearmultiple quantum correlation (HMQC) was obtained with the fieldgradient mode, and the attached proton test (APT) pulse sequencewas used to distinguish13C NMR multiplicities.

3. Results and Discussion

3.1.1H NMR Methods. The first reports involving biodieselsynthesis by transesterification and1H NMR analysis wereprimarily focused on yield determination of progressive metha-nolysis26,27or ethanolysis14,41 reactions (i.e., transesterificationof a vegetable oil or animal fat with methanol or ethanol,respectively). In an article by Knothe,27 the rate of oil conversionto fatty acid methyl ester (FAME) was also studied by removingaliquots through the reaction and analyzing by1H NMR. Thislatter work was expanded by recent papers,32,46 where NMRspectroscopy was used to elucidate aspects related to the kineticsand the mechanism of biodiesel production. The transesterifi-cation process to produce biodiesel fuel from TAGs follows astepwise mechanism of consecutive reversible reactions: (1)conversion of TAG into diacylglyceride (DAG) and FAME, (2)conversion of DAG into monoacylglyceride (MAG) and FAME,and (3) conversion of MAG into glycerol (GLY) and FAME.47

Morgenstern and co-workers32 calculated, from initial rates ofFAME formation, an activation energy of 27.2 kJ mol-1 forthe rate-determining step (DAG to MAG) in the multistepmechanism proposed by Freedman and co-workers.47 It has beenshown48 that13C NMR can provide valuable information aboutthe acyl positional distribution of TAG (triacylglyceride) invegetable oils. Jin and co-workers,46 by using1H and13C NMRto identify the positional isomers of DAGs and MAGs formedduring the transesterification reaction, reported that the metha-nolysis pathway occurs preferentially throughsn-1,3-DAGs tosn-1-MAG intermediates. These results showed that NMRspectroscopy can be successfully applied in all steps of biodieselproduction.

(41) Silva, C. L. M. Obtenc¸ ao de EÄ steres Etı´licos a Partir da Transes-terificacao do OÄ leo de Andiroba com Etanol. M.S. Thesis, University ofCampinas, Campinas, SP, Brazil, 2005.

(42) Ghesti, G. F.; Macedo, J. L.; Dias, J. A.; Dias, S. C. L. Green LewisAcid Catalysts for Biodiesel Production by Transesterification. BrazilianPatent Appl. 325, 2007.

(43) Ghesti, G. F.; Macedo, J. L.; Parente, V. C. I.; Dias, J. A.; Dias, S.C. L. To be submitted for publication, 2007.

(44) Gunstone, F. D.Fatty Acid and Lipid Chemistry; Aspen Publishers,Inc.: Gaithersburg, MD, 1999; p 76.

(45) Ghesti, G. F. Estudo de Catalisadores para Obtenc¸ ao de Biodieselpor Transesterificac¸ ao e Determinac¸ ao do Rendimento por EspectroscopiaRaman. M.S. Thesis, University of Brası´lia, Brasılia, DF, Brazil, 2006.

(46) Jin, F.; Kawasaki, K.; Kishida, H.; Tohji, K.; Moriya, T.; Enomoto,H. Fuel 2007, 86, 1201-1207.

(47) Freedman, B.; Butterfield, R. O.; Pryde, E. H.J. Am. Oil Chem.Soc.1986, 63, 1375-1380.

(48) Mannina, L.; Luchinat, C.; Emanuele, M. C.; Segre, A.Chem. Phys.Lipids 1999, 103, 47-55.

FT-Raman Spectroscopy Quantification of Biodiesel Energy & Fuels, Vol. 21, No. 5, 20072477

As described in the literature,49 the transesterification reactioncatalyzed by Lewis acid sites occurs through the coordinationof acyl groups of the triacylglyceride molecule to lowestunoccupied molecular orbitals of catalytic active centers. Thiscoordination increases acyl group polarization and forms acarbocation that undergoes alcohol nucleophilic attack. Thetetrahedral intermediary so formed eliminates the diacylglyceridemolecule and produces an ester.

Quantification of biodiesel by NMR can be made by simpleequations,26,27,41by building an analytical curve14 or by usingan internal standard (e.g., acetone).32 Ethyl ester quantificationby 1H NMR spectroscopy is more complex than methyl esterquantification due to a superimposition of the glyceryl meth-ylenic hydrogens in soybean oil and the-OCH2 from ethylester in biodiesel (see Figures 1-3). Figure 3 shows the1HNMR spectra of pure soybean oil (doublet of doublets), pureethyl esters (quartet), and a 40:60 (m/m) soybean oil/ethyl estermixture from 4.00 to 4.40 ppm, where it illustrates thedifficulties created from signals overlapping caused by partialconversion. To overcome this problem, two methods havealready been proposed by Neto and co-workers,14 who preparedan analytical curve, and Silva,41 who used an equation. In thenext paragraphs, we will describe the use of both methods, anda third alternative is suggested in this work.

To clarify the methods to be described, the following notationwas used (see Figure 2 and 3 for further clarification): (i)ITAG

) integration of glyceryl methylenic hydrogens at 4.25-4.35ppm; (ii) ITAG+EE ) integration of glyceryl methylenic hydro-gens and-OCH2 of ethoxy hydrogens superimposed at 4.10-4.20 ppm; and (iii)IRCH2 ) integration ofR-acyl methylenichydrogens in soybean oil and ethyl esters at 2.20-2.40 ppm.

The analytical curve was built by plotting known concen-trations of soybean oil/ethyl ester mixtures (see experi-mental section 2.3) versusITAG/ITAG+EE ratios. Values of 0 and1 were attributed to pure ethyl esters and pure soybean oil inITAG/ITAG+EE, respectively. The correlation coefficient (R2) soobtained was 0.9867.

The equation proposed by Silva41 and presented below (eq1) was applied to the same standard mixtures of soybean oiland ethyl ester prepared (see experimental section 2.3) todetermine soybean oil conversion into biodiesel (CEE):

A plot of predicted values versus real values for eq 1 showedanR2 ) 0.9977. However, in our1H NMR spectral analyses,45

it was observed that intermediary compounds formed duringtransesterification46 persisted in biodiesel fuel after purificationsteps, which regularly causes the signals from-OH groups inmono- and diacylglyceride species to overlap withRCH2 signalsat 2.20-2.40 ppm (Figure 4).50 Their presence was alsoobserved at 3.50-3.80 ppm.50 To use the integrated values ineq 1, all spectra were treated by deconvolution and subtractionsteps to ensure reliable results. To avoid these time-consumingprocedures, a new equation is proposed in this work (eq 2),whereR-acyl methylenic hydrogens (RCH2) are not taken intoaccount:

As one can see, it is obvious that the above equation can besimplified to%CEE ) 100[(ITAG+EE - ITAG)/(ITAG+EE + 2ITAG)].However, to facilitate the following explanations, it was left inthe original form. The numbers 4 and 6 in eq 2 are related tofour glyceryl methylenic hydrogens present in TAG molecules

(49) Parshall, G. W.; Ittel, S. D.Homogeneous Catalysis, 2nd ed.;Wiley: New York, 1992; pp 270-275.

Figure 2. Full 1H NMR spectra from (a) pure soybean oil and (b)pure ethyl ester.

Figure 3. 1H NMR spectra at 4.00-4.40 ppm region from (a) puresoybean oil, (b) 40:60 (m/m) soybean oil/ethyl esters, and (c) pure ethylester.

Figure 4. 1H NMR spectra at 2.20-2.40 ppm region from (a) pureethyl ester and (b) product from transesterification reaction after 2 h(* indicates NMR signals from intermediate species).

%CEE ) 100((ITAG+EE - ITAG)

IRCH2) (1)

%CEE ) 100( 4(ITAG+EE - ITAG)

4(ITAG+EE - ITAG) + 6(2ITAG)) (2)

2478 Energy & Fuels, Vol. 21, No. 5, 2007 Ghesti et al.

and to six hydrogens formed in three ethyl ester products (seeFigure 1), respectively. The number of glyceridic hydrogensassigned at 4.35-4.10 ppm was obtained from the litera-ture,32,46,48,50and also from our own 2D heteronuclear correlationin the HMQC experiment (not shown), where four glyceridichydrogens at 4.35-4.10 ppm were correlated with the CH2O-carbon signal at 62.0 ppm and one glyceridic hydrogen at 5.20-5.33 ppm was correlated with the CHO- carbon signal at 68.8ppm. The APT experiment (not shown) and computationalsimulation analysis were used to discriminate and correctlyassign carbon ressonances. The differenceITAG+EE - ITAG (i.e.,integration from the superimposition between glyceryl meth-ylenic hydrogens of TAG molecules and the-OCH2 of ethoxyhydrogens from ethyl ester molecules at 4.10-4.20 ppm minusthe integration of glyceryl methylenic hydrogens at 4.25-4.35ppm) in eq 2 is associated with ethyl ester formation. To simplifyeq 2, the areas from glyceryl methylenic groups for soybeanoil at 4.25-4.35 and 4.10-4.20 ppm were assumed to be equal(see Figure 3), that is, 2ITAG. Although this is not exactly true,because the ratio between the two areas is 0.98 instead of 1.00,the values obtained from eq 2 on standard soybean oil/ethylester mixtures, considering hydrogen areas to be equivalent ornot, were very similar (see Table 1). Indeed, a plot of biodieselconcentration predicted by eq 2 versus the true concentration,using both calculations in Table 1, resulted in the samecorrelation coefficient (R2 ) 0.9983).

3.2. FT-Raman/PLS Methods.Knothe’s work,3 the firstreport linking vegetable oil transesterification and vibrationalspectroscopic analysis, described biodiesel quantification andfuel quality assessment by NIR spectroscopy using a fiber-opticprobe. The major spectral differences between soybean oil andtheir respective methyl esters were distinguished at 4425-4430and 6005 cm-1, where it was observed that methyl estersdisplayed bands while soybean oil triacylglyceride moleculesexhibited only shoulders. These regions were analyzed forbiodiesel and contaminant quantifications (glycerol and metha-nol) in the spectra obtained from soybean oil/methyl ester andcontaminant/methyl ester mixtures by PLS calibration modelswith excellent results for both spectral regions. In a second NIRpaper,27 Knothe successfully applied the calibration models builtin the first paper to a transesterification reaction in progressand correlated, with good agreement, with1H NMR spectros-copy. Although FTIR spectra of vegetable oil and biodiesel arealmost identical, Zagonel and co-workers13 developed ananalytical procedure, based on CdO stretching vibrationdisplacement from 1746 cm-1 (TAGs) to 1735 cm-1 (ethylesters), to monitor soybean oil ethanolysis by FTIR multivariate

analysis models. The results obtained in a progressive transes-terification reaction by the calibration model, built from FTIRspectra of standard mixtures of triolein and ethyl esters analyzedby principal component analysis and PLS regression, werecorrelated with GPC analyses showing only small differences(R2 ) 0.9837).

Raman spectra of soybean oil and their corresponding ethylesters showed several differences: (i) a peak at 2932 cm-1

(νCH2), whereas the soybean oil spectrum displays only ashoulder; (ii) νCdO displacement from 1748 (soybean oil) to1739 cm-1 (ethyl esters); and (iii) bands at 861 (assigned asνR-CdO and νC-C) and 372 cm-1 (assigned asδCO-O-C).28

Raman spectra present well-defined bands and less congestionthan FTIR and NIR spectra, and as a consequence, there is asmaller possibility of band overlapping, making quantitativemeasurements simpler.51,52

As mentioned earlier, in our previous work,28 FT-Raman/PLS methods showed good prediction capabilities when appliedto known samples treated as unknown. A data set of 63 spectrafrom 21 standard soybean oil/ethyl ester mixtures (triplicatemeasurements) was used, and from the numerous correlationsand regression methods tested for analysis, FT-Raman/PLSmethods at the CH stretching region (3100-2740 cm-1, mainlythe peak at 2932 cm-1) could be distinguished from the othersdue to their predicted capabilities: (i)R2 ) 0.9994 for the modelusing 21 samples averaged from 63 measurements as a data set(model 4, notation used in ref 28) and (ii)R2 ) 0.9993 for themodel using all 63 measurements as a data set (model 5). Adetailed description of both methods was given elsewhere,28 andno further discussion will be mentioned here. The two modelswere applied to the heterogeneous transesterification reaction,and similar results were obtained. A plot of model 4 predictedvalues versus model 5 predicted values results in anR2 )0.9976. However, when compared to the predicted values foundby 1H NMR methods, model 4 was slightly better then model5. Although multivariate analysis generally requires a largenumber of samples to build reliable calibration models (at least20 samples for each component), the articles of Knothe3,27usingNIR/PLS, and Zagonel et al.13 using FTIR/PLS, were success-fully applied to progressive transesterification reactions withsmaller data sets in their models. In our case, the effects ofincreased data sets on FT-Raman/PLS models led to increasedpredicted capabilities and linearity (see Table 2).

3.3. Correlation Between FT-Raman/PLS and1H NMRMethods. In preceding articles dealing with spectroscopicmeasurements of biodiesel content in transesterification reac-

(50) Jie, M. S. F. L. K.; Lam, C. C.Chem. Phys. Lipids1995, 77, 155-171.

(51) Baran´ska, H.; Łbudzin´ska, A.; Terpin´ski, J.Laser Raman Spectros-copy: Analytical Applications; Ellis Horwood Limited: Chichester, U.K.,1987; pp 151-168.

(52) Skoog, D. A.; Holler, F. J.; Nieman, T. A.Principles of InstrumentalAnalysis; Saunders College Publishing: Philadelphia, PA, 1997; pp 429-443.

Table 1. Biodiesel True Concentration Values and Those Predictedby Eq 2 with and without Assumption of Six Standard Soybean

Oil/Ethyl Esters Mixtures

true CEE

values (wt %)

predictedCEE

values (wt %)without assumptiona

predictedCEE

values (wt %)with assumptionb

0.00 0.00 0.6320.45 20.64 21.1940.39 42.46 42.9059.85 60.62 60.9479.62 76.80 77.00

100.00 99.40 99.41

a CEE values calculated from eq 2 without assuming that glycerylmethylenic hydrogens areas at 4.25-4.35 and 4.10-4.20 ppm are identical.b CEE values calculated from eq 2 assuming that glyceryl methylenichydrogens areas at 4.25-4.35 and 4.10-4.20 ppm are identical.

Table 2. PredictedCEE Values by Raman/PLS Models withIncreased Data Set for Standard Soybean Oil/Ethyl Ester Mixtures

of Known Composition Treated as Unknown

predictedCEE values (wt %)

trueCEE

values (wt %)a

6 samplesdata set size

(R2 ) 0.9990)

11 samplesdata set size

(R2 ) 0.9991)

21 samplesdata set size

(R2 ) 0.9997)

0.00 3.73 3.71 0.52a

20.01 25.91 25.72 21.88a

49.82 51.23 51.34 50.28a

79.69 80.52 80.42 80.64a

100.00 99.64 99.18 99.31

a Values obtained from ref 28.

FT-Raman Spectroscopy Quantification of Biodiesel Energy & Fuels, Vol. 21, No. 5, 20072479

tions, the results obtained were correlated with other techniquesto validate their values. Knothe27 correlated NIR spectroscopyand two1H NMR equations with good agreement. Neto andco-workers14 correlated their1H NMR analytical curves withviscosity measurements, obtaining a linear correlation coefficient

of 0.9981. Zagonel and co-workers13 used GPC to validate theirFTIR/PLS methods, obtaining anR2 ) 0.9837 between bothtechniques. Figure 5 displays FT-Raman/PLS (model 4) and1H NMR methods applied to a progressive heterogeneoustransesterification reaction. The results obtained from all fourmethods (FT-Raman/PLS and three1H NMR methods) showedgood correlation (Figure 5) with each other, indicating theirmutual validation. Indeed, the linear fit between FT-Ramanmeasurements and all NMR methods tested (i.e., analyticalcurve, eq 1, and eq 2) showedR2 values of 0.9974, 0.9847, and0.9972, respectively (Figure 6). The difference observed betweenbiodiesel yield values from eq 1 and the other methods after 2h of reaction is associated with integration difficulties producedby signal superimposition from glyceridic intermediates (MAGsand DAGs) andR-acyl CH2.

Although both methods presented in this work do not attendthe detection limits required by ASTM to determine low levelsof contaminants in biodiesel fuel as chromatographic methodsdo, spectroscopic techniques allows a faster analysis of thereaction by easily identifying the formation of peaks or bandsrelated to ethyl ester products. These methods, when coupledto multivariate analysis, can be applied to fast reversiblereactions, such as transesterification,3,13,14,26,27,32,46to evaluatea new catalyst potentiality to produce biodiesel. In conclusion,FT-Raman/PLS models are an attractive alternative to rapidlymonitor biodiesel synthesis due to their inherent qualities andtheir reduced costs when compared with other techniques,including NMR. However,1H NMR methods, especially wheninvolving simple equations, are more easily adapted to othervegetable oils’ methanolysis or ethanolysis reactions. Bothprocedures described here were successfully correlated with eachother and validate by reported methods, evidencing and increas-ing their potential for biodiesel synthesis and quality monitoring.

Acknowledgment. Acknowledgement is given to CNPq, CAPES/PQI, FINATEC, Finep-CTPetro, Finep-CTInfra 970/01, UnB-IQ,UnB-IG, and FAPDF/SCDT/CNPq.

EF060657R

Figure 5. Biodiesel yield from a heterogeneous transesterificationreaction determined by four methods:1H NMR analytical curve, eq 1,eq 2, and FT-Raman/PLS model.

Figure 6. Plot from FT-Raman/PLS model predicted reaction valuesvs 1H NMR analytical curve, eq 1, and eq 2 predicted reaction values(R2 ) 0.9974, 0.9847, and 0.9972, respectively).

2480 Energy & Fuels, Vol. 21, No. 5, 2007 Ghesti et al.