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Journal of Chromatography A, 1196–1197 (2008) 147–152
Contents lists available at ScienceDirect
Journal of Chromatography A journal homepage: www.elsevier.com/locate/chroma
Green procedure with a green solvent for fats and oils’ determination Microwave-i Microw ave-integrated ntegrated Soxhlet using limonene followed by microw microwave ave Clevenger Cleveng er distillation Matthieu Virot a , Valerie ´ Tomao a , Christian Ginies a , Franco Visinoni b , Farid Chemat a,∗ a b
UMR 408 S´ecurit´ ecurit´e et Qualit´e des Produits d’Origine V´eg´ eg´etale, etale, INRA, Universit´e d’Avignon et des Pays du Vaucluse, 84000 Avignon, France Milestone s.r.l., via fatebenefratelli, 1/5, Sorisole, BG, Italy
a r t i c l e
i n f o
Article history: Available online 22 April 2008 Keywords: Microwave Limonene Soxhlet Clevenger Extraction
Currently there are only two kinds of solvents that can be used in chemistry: solvents obtained from petroleum industry and solvents of agricultural origin so-called “bio-solvents”. Hexane has been been used used fordecades fordecades for fats fats andoils’ determ determin inati ation on in food food prodproducts  .. It offers satisfactory extractions of fats and oils due to its low boiling point added to its low polarity. However, many works dealt dealt with with the the toxic toxic and and hazar hazardou douss effect effectss of this this solve solvent nt [2–5] and several several investig investigation ationss wer were e also achieve achieved d usingalternativ usingalternative e solvents solvents withthe aimof effectiv effective e andgreenerextractionproced andgreenerextractionprocedures,thatare ures,thatare safe for usersand more more environ environment mentallyfriendl allyfriendly y [1,6–9] [1,6–9].. Nevertheless less and and despit despite e its ranki ranking ng on top top of the the list list of hazar hazardou douss solve solvent nts, s, n-hexane is still the solvent of choice for fats and oils’ extractions. Limonene, which is the solvent used in this study is a major by-product of the citrus fruits industry [10,11] [10,11].. This monoterpenic monoterpenic molecule molecule (Fi Fig. g. 1) is the the major major compon componentof entof essent essential ial oils oils extra extract cted ed from citrus peels and plays an important role in the ﬁeld of ﬂavour and fragrances for many years thanks to its physical and chemical properties [11–13] [11–13].. Table 1 lists and compares the relevant properties ties of d-limonene -limonene and n-hexane. -hexane. The growin growing g interest interest in limonene limonene has emerged since its cleaner and degreaser qualities were recog-
nised and taken into consideration  .. This compound has thus been found to be a valuable alternative to halogenated hydrocarbons and/or conventional degreaser products traditionally used in industries and household cleaning. Liu and Mamidipally recently demonstrated that the industrial extraction of oil from rice bran was possible by using d-limonene instead of the regular n-hexane [15,16].. [15,16] Limon Limonene ene is a main main compo componen nentt of citrus citrus essent essential ial oils oils whic which h are commonly extracted from their matrix by using distillation. The Cleveng Clevenger er apparatu apparatuss has been used for decades decades in hydrodi hydrodistill stillation ation in order to extract and measure essential oils contained in plants [17–21].. This process is generally achieved [17–21] achieved in several hours [20,21] butis interest interestingin ingin thefact thatthe compoun compounds ds areextractedat areextractedat low tempera temperature ture (azeotr (azeotropic opic distillat distillation) ion) as compared compared to the high boiling point of essential oils and are thus not destroyed. Recently, an improved Clevenger apparatus using microwave energy has been suggested  .. This technique has been applied to extract essential oils and so, limonene (more than 90% of the composition of essential oils) from orange peels. This microwave extraction can be considered as an effective approach since it offers, among others, ers, short short extra extracti ction on times times (only (only 30min again against st 3 h in conv convent ention ional al method), low cost, low production of by-products (compared with convent conventiona ionall distillat distillation)and ion)and also a more more environ environment mentally ally friendly friendly extraction extraction procedure. We recently developed a new Soxhlet assisted by microwave energy  called microwave-integrated Soxhlet (MIS). This new device has shown to ensure rapid, efﬁcient and green extraction
M. Virot et al. / J. Chromatogr. A 1196–1197 (2008) 147–152
earth or stones were then removed from olive seeds before being randomized in plastic ﬂasks and stored at −18 ◦ C until use. 2.3. Moisture and volatile matter content
Fig. 1. Structure of limonene.
procedures. First, the experiments offered promising outcomes in terms of results compared with conventional Soxhlet extraction which offered quite similar results in terms of gravimetric determination and fatty acids composition. In addition extractions were performedin only 32min as comparedwith the 8 h required forthe Ofﬁcial extraction procedure. Furthermore, MIS can be considered as a green technology since energy used is reduced and since there is a possible solvent recycling of up to 90%. Thisinvestigation is a useful andgreen procedurecombining the two improved apparatus as described before. The aim of the study was to evaluate the possible extraction of fats and oils from olive seedsusing d-limoneneas solvent andcombining thisachievement by using microwave energy for both the extraction step and the cleaning step needed during the procedure. Extraction step of oils from olive seeds was thus investigated using the MIS and elimination step of the solvent from the medium was carried out by using the microwave Clevenger. Extracted oils were then compared withoils obtained using both conventional Soxhlet and MIS extraction procedures performed with n-hexane in term of crude extract (quantitative results) and fatty acid composition (qualitative comparison). 2. Experimental
2.1. Materials and reagents Solvents used during extraction experiments ( d-limonene or n-hexane) were of analytical grade and were supplied by VWR International (Darmstadt, Germany). Methanol, n-heptane, sodium chloride, sodium hydroxide and BF3 –methanol reagent (20% solution in methanol) used for the preparation of fatty acid derivatives were all of analytical grade and were also purchased from VWR International. Various glasswares, Soxhlet apparatus and extraction thimbles used in extractions and fatty acid methyl ester preparations were supplied by Legallais (Montferrier-sur-Lez, France). To release the solvent in conventional procedure, a rotary evaporator (Heidolph, laborata 2000) was utilized. To release solvent and/or water traces, an electrical oven “Aria guidata NS 9000” purchased from I.S.Co (Milan, Italy) was used. To evaluate percentage of crude oil in samples, an analytical balance APX-200 (Denver Instruments) was also employed. 2.2. Sample collection In these investigations, oil was extracted from olives Aglandau (Vaucluse, France) samples collected from local oil mill during harvest period for oil production. Odd materials such as leaves, Table 1 Relevant properties of n-hexane and d-limonene
Boiling point ( ◦ C) Density (g/mL) Heat of vaporization (kJ/kg) Dielectric constant ( ε) Odor Environmental impact Toxic
68.7 0.6603 334 2.0 Petroleum High Yes
175.5 0.8411 353 2.3 Citrus Low No
Samples studied were dried in an electrical oven before extraction as described: amounts of about 10 g of olive samples were placed on a desiccated and tarred capsule in an electrical oven at 80 ◦ C for 2 h. The capsule was then removed from the oven and cooled to room temperature in a desiccator before weighing. The above-describedprocedure wasrepeatedat every 2 h until the ratio m/m (%) was less than 10%. An average moisture of about 55% was found in our samples and was in agreement with that intended. 2.4. Extraction procedures Extractions performed, whatever the solvent and whatever the procedure used, were repeated at least three times and the mean yield values were reported. Yield of oil extracted was expressed as a percentage of the weight of oil obtained after extraction relative to the weight of dry sample used for extraction, as described hereinafter: % oilcontent =
2.4.1. Conventional Soxhlet extraction Extraction of fatsand oilsfrom saidolivesampleswas carried out according to an Ofﬁcial method . Soxhlet extractions were thus performed using 30g of olive sample weighed to the nearest 10 mg after grinding in an electrical mill. The amounts of ground samples were loaded in a 33 × 100 cellulose cartridge and transferred in the extraction chamber of a 200 mL capacity Soxhlet apparatus. Cotton wool was placed on the top of the cartridge in order to avoidtransfer of sample particles in thedistillation ﬂask. TheSoxhlet apparatus was ﬁtted with a condenser and placed on a 500 mL capacity distillation ﬂask containing 300 mL of solvent. Samples were thenextractedunder reﬂuxduring4 h. Thereafter, the cellulose thimble was removed from Soxhlet, cooled to room temperature in a desiccator and its content was then milled before being loaded again in the cellulose cartridge. Extraction was then performed during further 2 h and the above-described procedure was repeateduntila total extractiontime of8 h (4h + 2 h + 2 h). After the extraction, the major solvent contained in the distillation ﬂask was eliminated with a vacuum rotary evaporator. The content of the distillation ﬂask was then transferred in a smaller tarred ﬂask and concentrated to dryness with a vacuum rotary evaporator during 1 h at 80 ◦ C. The ﬂask was cooled to room temperature in a desiccator and weighed to the nearest mg. The above procedure was repeated for 30 min until a difference between two consecutive weights was smaller than 10% (m/m). Mean values were then reported before analysis step. 2.4.2. Microwave-integrated Soxhlet extraction MIS extractions have been performed in a Milestone DryDist microwave oven. The multi-mode microwave reactor has a maximum delivered power of 1000 W variable in 10W increments by usinga twinmagnetron(2 × 800 W,2.45 GHz). Duringexperiments, time, temperature, pressure and power were controlled with the “easy WAVE” software package. Temperature was monitored by a shielded thermocouple (ATC-300) inserted directly intothe sample container and by an infrared sensor outside the reactor. A traditional glass round-bottom ﬂask suited for microwave reactions was used as base vessel. It contains a polytetraﬂuoroethylene/graphite (Weﬂon, Milestone, Italy) stir bar capable of
M. Virot et al. / J. Chromatogr. A 1196–1197 (2008) 147–152
absorbing microwaves at the bottom of the vessel. The use of such a stir bar allows diffusion of heat created by the microwaves to the surroundings and is particularly useful when using transparent to microwave radiations’ solvents, i.e. which are not able to absorb microwaves, such as n-hexane or d-limonene (see low dielectric constants in Table 1). Extraction procedures performed with MIS were carried out as explained in previous article . They were performed in three steps, namely extraction time, leaching time and cleaning step (which canbe assimilated as thesolventelimination step or extract drying). Optimal settings evaluated in our previous investigation were also used to extract oil from olive seeds in this investigation (13 min for the extraction time, 17 min for the leaching time and 720W for the microwave irradiation power). 2.4.3. Microwave Clevenger distillation Microwave Clevenger distillations were carried out following MIS extraction steps. To perform, the extraction tube used in MIS procedures was removed from the device and distilled water was added to the mixture composed of extracted oil and limonene. The distillation ﬂask was then ﬁtted with Clevenger glassware (instead of the Soxhlet glassware) and the mixture was thus heated by means of microwaves. During azeotropic distillation of the binary water–limonene mixture, limonenewas eliminated fromthe distillation ﬂask. Both used limonene and extracted oil were recovered separately by phase separation. Limonene was recovered from the water layer by phase separation in the “separating funnel” of the Clevenger glassware and the extracted oil was recovered from the water layer by phase separation in the distillation ﬂask. 2.5. Preparation of fatty acid methyl ester derivatives AOCS Ofﬁcial method Ce 2-66 was used to prepare fatty acid methyl ester (FAMEs) derivatives . A basic catalysis was employed during derivatization procedures by using a deﬁned amount of 0.5M methanolic sodium hydroxide solution added to a speciﬁc amount of extracted oil. The mixture was then heated during 10 min and an amount of 5 mL of BF 3 –methanol reagent was then added to the solution. The ﬂask was allowed to heat again during 2 min and 5 mL of n-heptane was added to the solution. After 1 min, the ﬂask was removed from heat and 15mL of saturated sodium chloride solution were added. The ﬂask was then stoppered and shook vigorously during 15s. A small amount of the ﬂoating n-heptanelayer was then removed and transferred in a test tube containing a small quantity of anhydrous sodium sulphate. Samples were then ﬁltered through a 0.2 m cellulose regenerated ﬁlter (Alltech Associates, Deerﬁeld, IL, USA) before being injected directly in a gas chromatograph. Analyses, after derivation procedures, were performed in triplicate for each oil samples and mean values were reported. 2.6. Chromatographic analysis FAMEs were separated, quantiﬁed and identiﬁed by gas chromatography coupled with mass spectrometry (GC/MS). Analyses were performed by using a Shimadzu QP2010 (Kyoto, Japan) gas chromatography. The instrument wasequippedwith a CP-Wax capillarycolumn 30m × 0.32mm × 0.5 m (Varian) andthe velocityof the carrier gas (He) was at 47 cm/s. Injections of 2 L of the various samples were carried out with a split mode (ratio 1:15) and the injector temperature was set at 250 ◦ C. Oven temperature was initially 60 ◦ C for 1 min and then progressed at a rate of 20 ◦ C/min from 60 to 180 ◦ C and then increased from 180 to 230 ◦ C at a rate of 4 ◦ C/min. The temperature was then held at 230 ◦ C for 15 min. The mass spectra were recorded at 3 scan/s from 50 to 400 a.m.u.
Fig.2. Proposedextraction procedureusing limonene:microwave-integratedSoxhlet extraction (a) followed by microwave Clevenger distillation (b).
and theionisation mode was e.i. at 70eV. Identiﬁcation of common fatty acids was performed using the NIST’98 [US National Institute of Standards and Technology (NIST), Gaithersburg, MD, USA] mass spectral database. 3. Results and discussion
Extraction performedwith limonenehas alreadybeen described by authors [15,16]. Limonene appears as a good extraction solvent since its properties look like hexane in terms of polarity and thus afﬁnity for fats and oils. The main problem of using such a solvent is the higher energy consumption related to solvent recovery and thus, to solvent elimination, during the evaporating step due to its higher boiling point (175 ◦ C) when compared with n-hexane (69 ◦ C). More energy is de facto needed after extraction procedures to eliminate solvent when using d-limonene instead of n-hexane. This investigation is to propose a valuable extraction procedure of oil fromolive seeds followed by an originalelimination of limonene in the distillation ﬂask thus avoiding the use of large energy. As can be seenin Fig.2, extraction steps were performed using MIS device. Then, to eliminate d-limonene from the distillation ﬂask, we used the property that terpenes are traditionally extracted from their matrix by using a technique called steam or hydrodistillation thus inducing the use of an azeotropic distillation to below the boiling point of limonene under the boiling point of the water (97.4 ◦ C). 3.1. Quantitative determination Extracted oils obtained using different procedures were compared in terms of qualitative determination. Yields of crude extract were thus checked and compared for both methods using gravimetric determination. These results are listed in Table 2. As can be seen in this table, extracted mass of crude oil was higher using the new method than conventional Soxhlet and MIS procedures Table 2 Comparison of yields forolive oil extractionfunctions of procedureand solventused
Yield (%) a b c
Proposed procedure c
Conventional Soxhlet extraction using n-hexane. MIS extraction using n-hexane. MIS extraction with d-limonene followed by microwave Clevenger distillation.
M. Virot et al. / J. Chromatogr. A 1196–1197 (2008) 147–152
Fig. 3. Schematic representation of the different fatty acids found in the analyzed olive oil samples.
using n-hexane. This effect has already been noted by Liu and Mamidipally, and might be due to the slightly more polar nature of d-limonene compared with n-hexane [15,16]. As a consequence more compounds can be extracted from the matrix. A higher dissolving ability of limonene for triglycerides might also be pointed out by the higher temperature used to boil this solvent which can produce a lower viscosity of the analytes in the matrix and, in consequence, a better diffusion rate of the solute from the solid phase to the solvent. 3.2. Qualitative determination Extracted oils were converted into FAME derivatives and fatty acids composing the extracted oils were then separated and identiﬁed by using GC/MS. The composition of the analyzed olive oil contents, in term of chemical composition and relative amount of fatty acid functions of solvent and procedure used is listed in Table 3. The main fatty acids extracted using the new pro-
posed procedure are oleic (C18:1), palmitic (C16:0) and linoleic (C18:2) acids. These three fatty acids represent more than 90% of the total fatty acid composition of the extracted oil. Other fatty acids such as palmitoleic (C16:1), stearic (C18:0), linolenic (C18:3) or arachidic (C20:0) acids were also noted with a less predominant peak area. Myristic (C14:0), pentadecanoic (C15:0), margaric (C17:0), margaroleic (C17:1), nonadecyclic (C19:0), gadoleic(C20:1) and behenic (C22:0) acids were found in trace levels. The composition proﬁle of olive oil extracted with the new procedure was in good agreement with what was intended and with literature [26–30]. Results obtained with the presented method were almost similar with those obtained by both conventional Soxhlet and MIS extraction procedures. The sum percentages of saturated, mono- and poly-unsaturated fatty acids were also in line with those of several tables dealing with thefattyacid composition of olive oil found in literature . As can be seen in Table 3, the oleic/linoleic ratio was always higher than the minimal value of 7 [30,31]. As a conclusion, it canbe said that theproportionof the dif-
Table 3 Comparison of fatty acids compositions for olive oil extraction functions of procedure and solvent used
SFAs: Saturated fatty acids; MUFAs: mono-unsaturated fatty acids; PUFAs: poly-unsaturated fatty acids. a Conventional Soxhlet extraction using n-hexane. b MIS extraction using n-hexane. c MIS extraction using -limonene followed by microwave Clevenger distillation. d
M. Virot et al. / J. Chromatogr. A 1196–1197 (2008) 147–152
ferentfatty acidsas well as the proportion of SFAs,PUFAs,or MUFAs has not been affected by the unusual conditions used in our experiment, in other words, microwave energy and the use of limonene as solvent do not involve extraneous effects and/or artefacts on the composition of the extracted oils. The proportion of the different fatty acids has been simplistically represented in Fig. 3 (% axis has been cut at 30% in order to see the relative proportion of all of the fatty acids implicated). 3.3. Solvent recycling Limonene recovered from water layer in the funnel of the Clevenger apparatus was analyzed by GC/MS in order to check if extraction procedures increased the production of oxidation compounds in the solvent. The chromatogram proﬁle of both unused solvent and solvent of extraction were almost the same. Oxidation compounds such as carveol, carvone or limonene epoxides were found in trace levels in both solvents. As a consequence, the solvent of extraction recovered after experiment can be re-used for another extraction according to Fig. 4. 3.4. Green approach of the proposed procedure The green aspect of the proposed investigation can be pointed out readily in three points: 1st aspect : Our previous investigation dealing with MIS device  has shown that extractions were carried out in a shortened time compared with conventional extraction procedure which is a key parameter in terms of time and energy saving. The solvent recycling possibilities using MIS instead of Soxhlet apparatus have also been pointed out. Microwave energy is, in addition of that, the only heating source used and needed to perform extraction. As a consequence, the extraction step of the proposed procedure is clearly advantageous in term of time, solvent and energy saving. 2nd aspect : The microwave Clevenger apparatus is presented as a green process since it allows reduction of time and energy required for limonene distillation step. Currently, energy that can be used to eliminate limonene from the distillation ﬂask is reduced by using azeotropic distillation technique (temperature of evaporation diminished from about 175 ◦ C to less than 100 ◦ C). The use
of microwave energy to perform enhances the conventional distillation technique and is a clean and fast method for limonene release. 3rd aspect : The proposed approach using a green solvent to perform extraction is useful and can be considered as a nice alternative to conventional petroleum solvent where toxicity for both operator and environment is reduced. Furthermore, the use of a by-product of the industry as solvent, its possible recycling and life-cycle extension is original and of increasing interest for many chemistry experiments. This useful and safe procedure may lead to numerousinvestigations and/or alternatives to conventional chemistry procedures that are often hazardous. 3.5. Safety considerations Soxhlet and Clevenger apparatus are simple to use and their functioning can be readily understood. However, operators have to be careful since the use of such solvents with a high boiling point may be dangerous. Indeed, it should be taken into account that the combined use of microwaves together with such a solvent requires a stringent controlled handling. In addition, solvent boiling and release have to be controlled and always stirred to avoid hazardous effects of the distillation and/or extraction procedures. Finally, microwave energy is simple to use but can pose hazards when employed carelessly. Several knowledge and much abilities have to be taken into account before performing these kinds of experiments. Thecombined useof solvent with a high boiling point added to microwave energy needs a reﬂective, planned and approved approach. 4. Conclusion
The aim of the present study was to investigate an alternative procedure for the determination of fats and oils using microwave energy. Two original devices were combined to attempt this aim with good results in term of gravimetric determination (yields of crude oil) and fatty acid composition. As a consequence, it can be said that the proposed investigation is a valuable and effective method for fats and oils’ determination in olive seeds. In addition, the green aspect of the total procedure has to be taken into account since research concerning new alternatives and new solvents in chemistry is at the moment, for earth and environment protection, a key challenge that we cannot disregard. The use of a bio-solvent, its possible recycling by recovery from water phase and its combined use with MIS and microwave Clevenger procedures, which allow waste reduction, short operating time and energy saving, offers a nice approach that can be considered as a departure to one of the responses of Global Problems. This total procedure is a ﬁrst approach and we hope that this work will be the basis of further investigations in the ﬁeld of green extraction. References
Fig. 4. Extraction procedure used for experiments (MW: microwave).
 L.A. Johnson, E.W. Lusas, J. Am. Oil Chem. Soc. 60 (1983) 229.  P. Bavazzano, V. Li donni, A. Baldasseroni, Med. Lav. 84 (1993) 115.  L. Simonsen, H. Johnsen, S.P. Lund, E. Matikainen, U. Midtgard, A. Wennberg, Scand. J. Work Environ. Health 20 (1994) 1.  A. Baldasseroni, P. Bavazzano, E. Buiatti, E. Lanciotti, C. Lorini, S. Toti,A. Biggeri, Int. Arch. Occup. Environ. Health 76 (2003) 260.  P. Manini, R. Andreoli, W.M.A. Niessen, J. Chromatogr. A 1058 (2004) 21.  A.P. Gandh, K.C. Joshi, J. Krishna, V.S. Parihar, D.C. Srivastav, P. Raghunadh, J. Kawalkar, S.K. Jain, R.N. Tripathi, J. Food Sci. Technol. 42 (2005) 352.  A.P. Gandhi, K.C. Joshi, K. Jha, V.S. Parihar, D.C. Srivastav, P. Raghunadh, J. Kawalkar, S.K. Jain, R.N. Tripathi, Int. J. Food Sci. Technol 38 (2003) 369.  M.J. Noh, E.S. Choi, S.H. Kim, K.-P. Yoo, Y.H. Choi, Y.W. Chin, J. Kim, Kor. J. Chem. Eng. 14 (1997) 109.  S. Espinosa, M.S. Diaz, E.A. Brignole, Lat. Am. Appl. Res. 35 (2005) 321.
M. Virot et al. / J. Chromatogr. A 1196–1197 (2008) 147–152
 S.M. Njoroge, H. Koaze, P.N. Karanja, M. Sawamura, Flavour Fragrance J. 20 (2005) 80.  B. Mira, M. Blasco, A. Berna, S. Subirats, J. Supercrit. Fluids 14 (1999) 95.  S. Selli, T. Cabaroglu, A. Canbas, J. Food Compos. Anal. 17 (2004) 789.  E. Guenther, The Essential Oils, 3, R. E. Kreiger Publishing, New York, 1974.  T. Toplisek, R. Gustafson, Precision Cleaning 3 (1995) 4.  S.X. Liu, P.K. Mamidipally, Cereal Chem. 82 (2005) 209.  P.K. Mamidipally, S.X. Liu, Eur. J. Lipid Sci. Technol. 106 (2004) 122.  M.R.J.R. Albuquerque, T.L.G. Lemos, O.D.L. Pessoa, E.P. Nunes, R.F. Nascimento, E.R. Silveira, Flavour Fragrance J. 22 (2007) 249.  C. Capetanos, V. Saraglou, P.D. Marin, A. Simic, H.D. Skaltsa, J. Serb. Chem. Soc. 72 (2007) 961.  O.O. Okoh, A.A. Sadimenko, A.J. Afolayan, J. Appl. Sci. 7 (2007) 3806.  M. Negahban,S. Moharramipour,F. Seﬁdkon,J. Stored Prod.Res.43 (2007) 123.  N. Tabanca,F. Demirci, B. Demirci, D.E.Wedge,K.H. Can Baser,J. Pharm. Biomed. Anal. 45 (2007) 714.  A.M. Ferhat, B.Y. Meklati, J. Smadja, F.Chemat, J. Chromatogr.A 1112 (2006) 121.
 M. Virot, V. Tomao, G. Colnagui, F. Visinoni, F. Chemat, J. Chromatogr. A 1174 (2007) 138.  ISO 659-1988 (E),International Organization forStandardization (ISO), Geneva, 1988.  Ofﬁcial Method Ce 2-66, American Oil Chemists’ Society, Champaign, IL, 1989.  E. Beˇster, B. Butinar, M. Buˇcar-Miklavˇciˇc, T. Golob, Food Chem. 108 (2008) 446.  P. Pinelli,C. Galardi, N. Mulinacci,F.F. Vincieri,A. Cimato, A. Romani,Food Chem. 80 (2003 ) 331.  J.A. Perez-Serradilla, ´ R. Japon-Juj ´ an,M.D. ´ Luque de Castro, Anal. Chim. Acta 602 (2007) 82. ´ M.J. Motilva, M.J. Tovar, M.P. Romero, Food Chem. 85 (2004)  J.R. Morello, 357.  H. Manai, F. Mahjoub-Haddada, I. Oueslati, D. Daoud, M. Zarrouk, Sci. Hortic. 115 (2008) 252.  A. Ranalli, G. Modesti, M. Patumi, G. Fontanazza, Food Chem. 69 (2000) 37.