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Refined corn oil aromatization by Citrus aurantium peel essential oil
Industrial Crops and Products 32 (2010) 202–207
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Review
Refined corn oil aromatization by Citrus aurantium peel essential oil Iness Jabri Karoui ∗ , Wissem Aidi Wannes, Brahim Marzouk Aromatic and Medicinal Plants Unit, Center of Biotechnology of the Technopol Borj-Cedria, BP. 901, 2050 Hammam-Lif, Tunisia
a r t i c l e
i n f o
Article history: Received 18 October 2009 Received in revised form 23 April 2010 Accepted 29 April 2010
Keywords: Oil aromatization Monterpenes Bitter orange peel Limonene
a b s t r a c t Corn oil was submitted to dynamic headspace to eliminate volatile compounds remained after refining process. The optimization of extraction parameters leads to an important deodorization after 4 h of extraction with residual aroma content of about 0.901 g/ml of deodorized corn oil. Different peel quantities and different incubation times were used during this experiment while oil volume, incubator temperature and shaking speed were hold constant. Essential oil components retained in corn oil were mainly represented by monoterpene hydrocarbons and limonene was the major one (ranging from 92.57% to 96.11%). Samples containing 15 g of Citrus peel and incubated for 1 h, showed the highest total volatiles with 2.4 mg/ml and limonene represented 2.3 mg/ml. Fatty acid analysis showed that aromatization did not affect fatty acid composition. © 2010 Elsevier B.V. All rights reserved.
Contents 1. 2.
3.
4.
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Materials and methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1. Plant material . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2. Lipid matrix . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.3. Refined corn oil deodorization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.4. Refined corn oil aromatization process . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.5. Peel essential oil characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.6. GC–FID analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.7. GC–MS analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.8. Compounds identification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.9. Lipid matrix characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.9.1. Fatty acid methyl esters (FAMEs) preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.9.2. Fatty acid analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.10. Statistical analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Results and discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1. Corn oil deodorization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2. Refined corn oil aromatization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3. Corn oil fatty acid composition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4. Peel essential oil composition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5. Evaluation of lipid matrix and aroma interaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
∗ Corresponding author. Tel.: +216 71 430 855; fax: +216 79 412 638. E-mail address: [email protected] (I.J. Karoui). 0926-6690/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.indcrop.2010.04.020
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1. Introduction
2.2. Lipid matrix
Flavour is considered as one of the most important sensory properties determining the food acceptance by the consumer (Guichard, 2006). Many recent investigations projected developing advanced technologies to improve sensory qualities of food products and to satisfy consumer. In general, food aroma consists of many volatile compounds and depends on the composition and the structure of the matrix, that influence their retention and release (Seuvre et al., 2006). In addition, not only aroma compounds nature and concentration are important in food flavouring but also their binding behaviour is of great practical importance. Therefore, aroma interactions with non-volatile macromolecules such as sugars, proteins and lipids have been thoroughly reviewed (Bakker, 1995; Druaux et al., 1998; Van Ruth et al., 2008). De Roos (2003) studied also, the effect of food texture and microstructure on flavour retention and release. Many studies have been done on the protein–flavour interactions. Several ones demonstrated reversible, non-specific, hydrophobic binding of most volatile compounds tested (ketones, alcohols, aldehydes, terpenes) with proteins such as bovine serum albumin and -lactoglobulin (Guichard, 2006). As regards, the interactions of aroma compounds with sugars, added mono- and disaccharides usually increase the vapour pressure (Bakker, 1995). Polysaccharides, on the other hand, usually decrease aroma compounds volatility (Le Thanh et al., 1992). Among all the food ingredients, lipids have probably the strongest organoleptic impact (Piraprez et al., 1998). For instance, Ebeler et al. (1988), studying menthone and isoamyl acetate in soybean oil, found lipids to significantly increase perceived flavour intensity. Seed oil composition has been studied extensively (Conte et al., 2004). Recently, more interest is given to its minor compounds (volatiles, chlorophylls, phenolics, etc.) (Shahidi, 2002). In order to improve quality of crude oils, refining process is necessary to remove undesirable compounds (phospholipids, free fatty acids, pigments and volatile compounds) responsible for off-flavours; although some unwanted non-volatile compounds might remain at the end of the process (Ruiz-Méndez et al., 1997). To improve the taste of oil and to satisfy consumer preferences, one of the key discoveries was the incorporation of aromatic plants such as lavender, thyme and menthe (Maldao-Martins et al., 2004; Bensmira et al., 2007). In the same case, the main objective of this work was the aromatization of refined corn oil by bitter orange peel. The latter was selected as aromatic plant material for our work because of its role in the promotion of the appetite, besides its medicinal properties such as regulation effect on some central nervous disorders, anxiety treatment and sedative effect (De Moreas Pultrini et al., 2006). Moreover, Tunisia is well known for its important production of Citrus fruits and such survey leads also to the valorization of citrus peel as already studied (Rehman, 2006).
Refined corn (Zea mays L.) oil was selected for this study owing to its common use in Tunisia. The process alkali refining of this oil includes four steps: degumming, neutralization, bleaching and deodorizing. Samples belonging to the same lot, composed of: crude, degummed, bleached and alkali-refined corn oil, were purchased from a local refinery located at Oued Ellil (North West of Tunisia) then stored at cold (4 ◦ C) in the dark following the results reported by Marzouk and Riahi (2000), Riahi and Marzouk (2000) and Naz et al. (2005).
2. Materials and methods 2.1. Plant material Bitter orange (Citrus aurantium L.) is a plant that belongs to Rutaceae family. Its fruit pulp is acidic and the albedo is bitterer (Chapot and Praloran, 1955). Fresh fruits used in our research, were originate from an orchard located at the Cap-Bon region (North East of Tunisia); they have been harvested from four selected trees, peeled then the peel was used as an oil aromatizing agent. Bitter orange skin was composed of two constituents: flavedo and albedo; its essential oil stored in flavedo glands, was easily extracted by mechanic ways and has a very characteristic pleasant odor (Chapot and Praloran, 1955).
2.3. Refined corn oil deodorization Alkali-refined samples were selected owing to their very low volatiles level. They were extracted by dynamic headspace method (Druaux et al., 1998) with splashed gas N2 (Strip-trap) according to published data (Dhifi et al., 2002). In fact, nitrogen gas was bubbled through 40 ml of oil at a flow rate corresponding to a constant pressure of 0.4 bar. The 6-methyl-5-hepten-2-one was added in the sample as an internal standard in order to quantify the aroma residue. Volatile compounds were stripped with nitrogen for 4 h (time fixed after a kinetic survey during 1 h 15 min, 2 h, 3 h and 4 h), trapped on 50 mg of activated charcoal (0.5–0.85 mm, 20–35 mesh ASTM, Merck, Schuchardt, Germany), eluted 120 with 1 ml of diethyl-ether and then analyzed. Oil was maintained at a temperature about 36 ± 1 ◦ C for preserving flavour according to earlier reports (Dhifi et al., 2002). 2.4. Refined corn oil aromatization process The aromatization was used to improve the organoleptic profile of corn oil. For this, deodorized refined corn oil was put in a 500 ml vial and used as lipid matrix. Bitter orange peels were cut into small pieces then homogenized in the oil in an incubator shaker (Excella E-24/24R Benchtop, New Brunswick Scientific CO., INC.) with dynamic plate and hermetical lid. Incubation period (1 h, 2 h and 3 h) and peel quantity (5 g, 10 g and 15 g) were optimized while oil volume (40 ml), incubator temperature (20 ◦ C) and shaking speed (100 rpm) remained constant. After, flavoured oils were filtered (peels were carried out of oil) and an internal standard (6-methyl-5-hepten-2-one) was added. Volatile compounds were extracted using the same headspace parameters of deodorization method since these optimized conditions permitted to obtain an aroma maximum extraction. The aim of the extraction step was to evaluate peel essential oil retention by the lipid matrix. 2.5. Peel essential oil characterization Peel composition was studied in order to confirm later their retention in corn oil. Peel volatiles were extracted by hydrodistillation method. For this, fresh peels (50 g) were immersed in double their volume of distilled water and essential oils were extracted during 90 min; this time was fixed after a kinetic survey during 30 min, 60 min, 90 min and 120 min. Essential oils were recovered from the emulsion with diethyl-ether (v/v) and added with an internal standard, the 6-methyl-5-hepten-2-one, in order to quantify them. After a concentration at 35 ◦ C, using a Vigreux column, essential oils obtained were subsequently analyzed by gas chromatography. All experiments were made in triplicates. 2.6. GC–FID analysis Peel essential oil as well as deodorized and flavoured corn oil volatile compounds, were analyzed by GC using a Hewlett–Packard
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6890 apparatus (Agilent Technologies, Palo Alto, CA, USA), equipped with a flame ionization detector (FID) and an electronic pressure control (EPC) injector. A polyethylene glycol capillary column (HP Innowax, 30 m × 0.25 mm i.d., 0.25 m film thickness) and an apolar HP-5 column (30 m × 0.25 mm, 0.25 m film thickness) were used. The flow of the carrier gas (N2 ) was 1.6 ml/min and the split ratio in the injector was 60:1. The analysis was performed using the following temperature programme: oven temperature isotherm at 35 ◦ C for 10 min, from 35 to 205 ◦ C at the rate of 3 ◦ C min−1 , and isotherm at 205 ◦ C during 10 min. Injector and detector temperatures were held, respectively, at 250 and 300 ◦ C. Surfaces of peaks and percentages of the different compounds were determined using the same HP chemstation cited above. 2.7. GC–MS analysis A HP-5890 series II coupled to a HP-5972 mass spectrometer with electron impact ionization (70 eV) and a HP-5 MS capillary column (30 m × 0.25 mm, 0.25 m film thickness) were used. Column temperature was programmed to rise from 50 to 240 ◦ C at a rate of 5 ◦ C min−1 ; transfer line temperature was 250 ◦ C. The carrier gas was helium with a flow rate of 1.2 ml/min and a split ratio of 60:1. Scan time and mass range were 1 s and 40–300 m/z, respectively.
Fig. 1. Quantitative evaluation of peel essential oil retained in deodorized refined corn oil after different incubation times (1 h, 2 h or 3 h = 1H, 2H or 3H, respectively); total volatile compound contents with different subscripts (a–e) were significantly different at p < 0.05 (Duncan test).
2.10. Statistical analyses Data were subjected to statistical analysis using statistical program package STATISTICA (Statsoft, 1998). Percentage of each volatile compound and fatty acids was the mean of three replicates ± standard deviation and the differences between individual means were deemed to be significant at p < 0.05.
2.8. Compounds identification The identity of the oil components was assigned by comparison of their retention indices relative to (C8–C22) n-alkanes with those of literature or with those of authentic compounds available in our laboratory. Further identification was made by matching their recorded mass spectra with those stored in the Wiley/NBS mass spectral library 167 of the GC–MS data system and other published mass spectra (Adams, 2001). 2.9. Lipid matrix characterization 2.9.1. Fatty acid methyl esters (FAMEs) preparation Crude, degummed, bleached, alkali-refined and flavoured corn oil fatty acids were transformed into their FAMEs after a transmethylation according to the method described by Cecchi et al. (1985), using sodium methylate. An aliquot (50 g of oil) was prepared in a screw tube; then, 2 ml of hexane and 0.5 ml of sodium methylate (3% in methanol) were added. After stirring during 1 min and standing for 2 min, the mixture was neutralized by addition of 0.2 ml of H2 SO4 (1N). After washing the mixture with 1.5 ml of distilled water, the superior phase containing FAMEs was poured off, concentrated under N2 stream and analyzed by gas chromatography (GC). Experiments were made in triplicates. 2.9.2. Fatty acid analysis FAMEs were analyzed by GC, using the same apparatus previously described; the flow of the carrier gas (N2 , U) was 1.6 ml/min and the split ratio was 60:1. The detector and injector temperatures were set at 275 and 250 ◦ C, respectively. The initial oven temperature was held at 150 ◦ C for 1 min−1 , increased at a rate of 15 ◦ C min−1 to 200 ◦ C and then held there for 3 min and finally ramped at 2 ◦ C min−1 for 184–242 ◦ C. Surfaces of peaks and percentages of the different compounds were determined using HP chemstation (Rev.A.0401) software. The latter also permits to control the analytic parameters (flow, temperature, beginning and end of the analyses, etc.). Fatty acids were identified by comparing their retention times with those of authentic standards analyzed in the same conditions.
3. Results and discussion 3.1. Corn oil deodorization The aim of this step was to remove residual volatiles responsible for off-flavours remained at the end of refining process in order to obtain a completely deodorized lipid matrix and consequently to be able to evaluate only peel essential oil quantity retained. Headspace parameters optimization permitted to reach a considerable aroma residue when extraction period was about 1 h 15 min, 2 h and 3 h. However, samples submitted to 4 h extraction showed the best deodorization result with residual aroma of about 3.4% (0.901 g/ml of oil). 3.2. Refined corn oil aromatization Refined oil deodorized during 4 h was used as lipid matrix. Different shaking periods (1 h, 2 h and 3 h) and variable peel quantities (5 g, 10 g and 15 g) were experimented. Temperature of incubator was held at 20 ◦ C because of the essential oil volatility that increases at high temperatures (Weel et al., 2004; De Roos, 2006). Due to a fixed oil volume used (40 ml), peel quantity used did not pass 15 g; In fact, 20 g of peel were experimented and the homogenization was partial. Shaker speed was 100 rpm. GC profiles showed that the percentages of peel essential oil compounds retained were comparable for all the samples studied. Especially hydrocarbon compounds were readily retained on lipid matrix with a large contribution of monoterpene hydrocarbons such as limonene, ␣-pinene, -pinene, sabinene and ␣-terpinene with a total percentage ranging from 96% to 99%. The only oxygenated compound retained was 1,8-cineole with a proportion inferior to 1% (Table 1). In addition, all samples showed high proportions of limonene varying from 92.57% to 96.11%. However, all the other compounds forming the minor fraction of peel essential oils were retained. Peel quantity and incubation time effects on peel total volatiles retention in corn oil were studied. Table 1 and Fig. 1 pointed out the highest contents of some volatiles retained in oils after 1 h of incu-
0.00e 0.00e 0.00f 0.00e 0.01f 0.00e 0.00a 0.01e ± ± ± ± ± ± ± ± 0.006 0.002 0.001 0.01 0.39 0.002 0.01 0.42 ␣-Pinene -Pinene Sabinene ␣-Terpinéne Limonéne 1–8 Cinéole Others Total volatiles
0.01 0.003 0.001 0.02 1.09 0.01 0.03 1.17
± ± ± ± ± ± ± ±
1.10 0.01 ± 0.00abc 1.21 0.00bc 0.30 0.004 ± 0.00de 0.38 0.00de 0.12 0.00bcde 0.13 0.001 ± 0.00def 1.68 0.02 ± 0.00cd 1.49 0.00c de de 93.39 1.09 ± 0.21 95.69 0.32 0.60 0.01 ± 0.00c 0.72 0.00c a a 0.01 2.80 0.004 ± 0.001 0.39 d d 100 1.14 ± 0.17 100 0.27
%
205
bation and mainly limonene. However, samples incubated for 2 h or 3 h showed decrease retention. This could be explained by volatile fraction partial lost when shaking period was long. In addition, the contents of total volatile compounds retained in corn oil were higher when peel quantity was more important. Furthermore, the highest contents of aroma (2.4 mg/ml) and limonene (2.3 mg/ml) were detected in the samples incubated with 15 g of peel for 1 h. The results obtained indicated that peel quantity and incubation time affected aroma retention in the lipid matrix. It is worth noting that this work, to the best of our knowledge, is the first one dealing with aromatization optimization conditions of corn oil by C. aurantium. 3.3. Corn oil fatty acid composition As can be seen in Table 2, fatty acid composition pointed out the abundance of three fatty acids (palmitic C16:0, oleic C18:1 and linoleic C18:2) which accounted 96.92% fraction in crude oil, 97.45% in degummed oil, 96.42% in bleached oil, 96.26% in refined oil and 97.08% flavoured oils. The unsaturated fatty acids (C18:1 and C18:2) were more abundant than the saturated ones with the highest rate of the linoleic acid varying from 56% to 57%, followed by the oleic one ranging from 26.12% to 27.46%. The palmitic acid was present at a percentage varying from 13% to 14.5%. These results showed that there were not significant differences among different samples, permitting to deduce that refining process did not affect fatty acid composition but affected essentially minor compounds of oil such as volatiles, pigments and phenols, as indicated in previous work (Lanzon et al., 1994). In addition, aromatization process did not also change fatty acid composition but improved oil quality. 3.4. Peel essential oil composition
Values followed by the same small letter did not share significant differences at 5% (Duncan test).
% mg/ml %
1.49 0.02 ± 0.00a 0.9 0.01 ± 0.00ab 1.06 0.01 ± 0.00abc 1.26 0.01 ± 0.00cd 0.44 0.01 ± 0.00bc 0.68 0.01 ± 0.00de 1.23 0.55 0.01 ± 0.00bc 0.37 0.004 ± 0.00de 0.29 0.01 ± 0.00a 0.89 0.01 ± 0.00cde 0.20 0.01 ± 0.00cd 0.32 0.01 ± 0.00ab 1.35 0.20 0.002 ± 0.00a 0.11 0.001 ± 0.00cde 0.10 0.002 ± 0.00abc 0.17 0.001 ± 0.00ef 0.05 0.002 ± 0.00abcd 0.10 0.002 ± 0.00ab 0.33 1.16 0.03 ± 0.00b 1.84 0.02 ± 0.00c 1.51 0.01 ± 0.00de 1.07 0.04 ± 0.01a 1.81 0.03 ± 0.00b 1.68 0.01 ± 0.00e 1.62 b cd e a bc f 93.48 1.82 ± 0.15 95.75 1.39 ± 0.18 95.79 1.01 ± 0.08 92.83 2.31 ± 0.50 95.91 1.65 ± 0.02 95.20 0.55 ± 0.04 92.58 0.39 0.01 ± 0.00b 0.71 0.01 ± 0.00b 0.92 0.004 ± 0.00d 0.38 0.02 ± 0.00a 0.86 0.01 ± 0.00b 0.69 0.00 ± 0.00de 0.53 a a a a a a 2.74 0.006 ± 0.00 0.34 0.005 ± 0.00 0.33 0.04 ± 0.00 3.41 0.02 ± 0.01 0.73 0.02 ± 0.00 1.32 0.01 ± 0.00 2.36 b cd d a bc e 100 1.90 ± 0.12 100 1.45 ± 0.15 100 1.09 ± 0.06 100 2.40 ± 0.41 100 1.73 ± 0.01 100 0.59 ± 0.03 100
3 h (incubation time) 2 h (incubation time)
mg/ml %
1 h (incubation time)
mg/ml % mg/ml % mg/ml % mg/ml % mg/ml mg/ml % mg/ml
10 g (peel quantity)
1 h (incubation time) 3 h (incubation time) 2 h (incubation time) 1 h (incubation time)
5 g (peel quantity) Compounds
Table 1 Contents (mg/ml of oil) and percentages (%) of peel volatile retained in deodorized corn oil.
2 h (incubation time)
3 h (incubation time)
15 g (peel quantity)
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Peel essential oil composition was determined and permitted to conclude on a large contribution of terpenes (2.71 mg/g of peel). The major monoterpene hydrocarbons were represented by limonene, ␣-pinene, -pinene, sabinene and ␣-terpinene with a total percentage of about 92.8%. These results are in agreement with those found by Dugo et al. (1993). The limonene importance in citrus peel has been confirmed by several studies; its percentage varied from 90% to 95% (Dugo et al., 1993; Saidani and Marzouk, 2003) which is in conformity with our result (90.25%). Limonene is one of the most common terpenes in nature and is the majority constituent of an essential oil series. Its pleasant citric fragrance is commonly used as a flavouring in foods and drinks, for which it is classified in the U.S. Code of Federal Regulation as safe. Tests in animals have proven the effectiveness of limonene against some types of cancer including gastric, mammary, pulmonary adenoma and liver. Limonene also has been shown to be effective in relieving gastroesophageal reflux disorder and occasional heartburn (Thiago et al., 2009). 3.5. Evaluation of lipid matrix and aroma interaction To evaluate volatile compounds retention by lipid matrix during the aromatization experiments, peel essential oil composition was used as a reference, after their extraction by hydrodistillation using different peel quantities (5 g, 10 g and 15 g). A great interest was given to only compounds retained. Based on the results presented in Figs. 2 and 3, the quantity of total volatile compound retained was considerable in the major part of samples when headspace method and oil matrix were used. This could be explained by the high hydrophobic matrix used and the relatively lipophilic character of the most aroma compounds that leads to a higher affinity of these latter to corn oil. These results were in conformity with those found by Piraprez et al. (1998). Fur-
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Table 2 Fatty acid composition (%) of corn oil during refining and aromatization steps. Fatty acid % C16:0 C16:1 C18:0 C18:1 C18:2 C18:3 C20:0 C20:1
Crude oil 13.4 0.07 1.52 27.46 56.06 0.87 0.28 0.14
± ± ± ± ± ± ± ±
Degummed oil c
0.6 0.01h 0.1d 1.20b 1.9a 0.06e 0.01f 0.05g
14.46 0.08 1.10 26.12 56.87 0.9 0.23 0.16
± ± ± ± ± ± ± ±
c
0.9 0.03g 0.2d 0.93b 2.4a 0.02d 0.03e 0.01f
Bleached oil 13.19 0.13 1.35 26.20 57.03 0.98 0.29 0.21
± ± ± ± ± ± ± ±
c
0.5 0.01g 0.3d 1.5b 1.8a 0.20d 0.0e 0.03f
Refined oil 13.81 0.09 1.49 26.36 56.09 1.23 0.21 0.18
± ± ± ± ± ± ± ±
Flavoured oil c
1.3 0.05h 0.1d 1.2b 2.2a 0.05e 0.04f 0.02g
14.55 0.06 1.22 26.33 56.20 1.00 0.27 0.1
± ± ± ± ± ± ± ±
1.53c 0.00 0.02d 1.02b 1.6a 0.13e 0.06f 0.01
Values followed by the same small letter did not share significant differences at 5% (Duncan test).
92.57% to 96.11%. Fatty acid composition analysis showed nearly the same proportions after all the refining steps and after aromatization. The total quantity of peel volatile compounds retained was considerable when headspace method and oil matrix were used with comparison to hydrodistillation method. These results incite us to pursue this research by determining the antioxidant and antiradical potentialities of flavoured corn oil and the maximum aroma conservation period in this oil.
References Fig. 2. Peel volatile compounds quantity extracted by hydrodistillation; total volatile compound contents with different subscripts (a–c) were significantly different at p < 0.05 (Duncan test).
Fig. 3. Peel volatile compounds quantity in corn oil extracted by headspace after different incubation times (1 h, 2 h or 3 h = 1H, 2H or 3H, respectively); total volatile compound contents with different subscripts (a–g) were significantly different at p < 0.05 (Duncan test).
thermore, Seuvre et al. (2006), studying aroma retention in water and oil, pointed out aroma retention by lipids and indicated that due to the hydrophobic character of major volatile compounds, polar ones were the most soluble in water; this was in conformity with our results. Headspace system is a method using a cold extraction at a temperature of about 36 ± 1 ◦ C that permits an important retention of non-polar compounds which constituted the largest part of retained volatiles. 4. Conclusion Aroma analysis indicated that peel compounds especially monoterpene hydrocarbons such as limonene, ␣-pinene, -pinene, sabinene and ␣-terpinene were retained in deodorized refined corn oil with a total percentage ranging from 96% to 99%. Limonene was the main compound detected with a percentage varying from
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Review
Refined corn oil aromatization by Citrus aurantium peel essential oil Iness Jabri Karoui ∗ , Wissem Aidi Wannes, Brahim Marzouk Aromatic and Medicinal Plants Unit, Center of Biotechnology of the Technopol Borj-Cedria, BP. 901, 2050 Hammam-Lif, Tunisia
a r t i c l e
i n f o
Article history: Received 18 October 2009 Received in revised form 23 April 2010 Accepted 29 April 2010
Keywords: Oil aromatization Monterpenes Bitter orange peel Limonene
a b s t r a c t Corn oil was submitted to dynamic headspace to eliminate volatile compounds remained after refining process. The optimization of extraction parameters leads to an important deodorization after 4 h of extraction with residual aroma content of about 0.901 g/ml of deodorized corn oil. Different peel quantities and different incubation times were used during this experiment while oil volume, incubator temperature and shaking speed were hold constant. Essential oil components retained in corn oil were mainly represented by monoterpene hydrocarbons and limonene was the major one (ranging from 92.57% to 96.11%). Samples containing 15 g of Citrus peel and incubated for 1 h, showed the highest total volatiles with 2.4 mg/ml and limonene represented 2.3 mg/ml. Fatty acid analysis showed that aromatization did not affect fatty acid composition. © 2010 Elsevier B.V. All rights reserved.
Contents 1. 2.
3.
4.
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Materials and methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1. Plant material . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2. Lipid matrix . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.3. Refined corn oil deodorization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.4. Refined corn oil aromatization process . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.5. Peel essential oil characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.6. GC–FID analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.7. GC–MS analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.8. Compounds identification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.9. Lipid matrix characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.9.1. Fatty acid methyl esters (FAMEs) preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.9.2. Fatty acid analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.10. Statistical analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Results and discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1. Corn oil deodorization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2. Refined corn oil aromatization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3. Corn oil fatty acid composition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4. Peel essential oil composition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5. Evaluation of lipid matrix and aroma interaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
∗ Corresponding author. Tel.: +216 71 430 855; fax: +216 79 412 638. E-mail address: [email protected] (I.J. Karoui). 0926-6690/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.indcrop.2010.04.020
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1. Introduction
2.2. Lipid matrix
Flavour is considered as one of the most important sensory properties determining the food acceptance by the consumer (Guichard, 2006). Many recent investigations projected developing advanced technologies to improve sensory qualities of food products and to satisfy consumer. In general, food aroma consists of many volatile compounds and depends on the composition and the structure of the matrix, that influence their retention and release (Seuvre et al., 2006). In addition, not only aroma compounds nature and concentration are important in food flavouring but also their binding behaviour is of great practical importance. Therefore, aroma interactions with non-volatile macromolecules such as sugars, proteins and lipids have been thoroughly reviewed (Bakker, 1995; Druaux et al., 1998; Van Ruth et al., 2008). De Roos (2003) studied also, the effect of food texture and microstructure on flavour retention and release. Many studies have been done on the protein–flavour interactions. Several ones demonstrated reversible, non-specific, hydrophobic binding of most volatile compounds tested (ketones, alcohols, aldehydes, terpenes) with proteins such as bovine serum albumin and -lactoglobulin (Guichard, 2006). As regards, the interactions of aroma compounds with sugars, added mono- and disaccharides usually increase the vapour pressure (Bakker, 1995). Polysaccharides, on the other hand, usually decrease aroma compounds volatility (Le Thanh et al., 1992). Among all the food ingredients, lipids have probably the strongest organoleptic impact (Piraprez et al., 1998). For instance, Ebeler et al. (1988), studying menthone and isoamyl acetate in soybean oil, found lipids to significantly increase perceived flavour intensity. Seed oil composition has been studied extensively (Conte et al., 2004). Recently, more interest is given to its minor compounds (volatiles, chlorophylls, phenolics, etc.) (Shahidi, 2002). In order to improve quality of crude oils, refining process is necessary to remove undesirable compounds (phospholipids, free fatty acids, pigments and volatile compounds) responsible for off-flavours; although some unwanted non-volatile compounds might remain at the end of the process (Ruiz-Méndez et al., 1997). To improve the taste of oil and to satisfy consumer preferences, one of the key discoveries was the incorporation of aromatic plants such as lavender, thyme and menthe (Maldao-Martins et al., 2004; Bensmira et al., 2007). In the same case, the main objective of this work was the aromatization of refined corn oil by bitter orange peel. The latter was selected as aromatic plant material for our work because of its role in the promotion of the appetite, besides its medicinal properties such as regulation effect on some central nervous disorders, anxiety treatment and sedative effect (De Moreas Pultrini et al., 2006). Moreover, Tunisia is well known for its important production of Citrus fruits and such survey leads also to the valorization of citrus peel as already studied (Rehman, 2006).
Refined corn (Zea mays L.) oil was selected for this study owing to its common use in Tunisia. The process alkali refining of this oil includes four steps: degumming, neutralization, bleaching and deodorizing. Samples belonging to the same lot, composed of: crude, degummed, bleached and alkali-refined corn oil, were purchased from a local refinery located at Oued Ellil (North West of Tunisia) then stored at cold (4 ◦ C) in the dark following the results reported by Marzouk and Riahi (2000), Riahi and Marzouk (2000) and Naz et al. (2005).
2. Materials and methods 2.1. Plant material Bitter orange (Citrus aurantium L.) is a plant that belongs to Rutaceae family. Its fruit pulp is acidic and the albedo is bitterer (Chapot and Praloran, 1955). Fresh fruits used in our research, were originate from an orchard located at the Cap-Bon region (North East of Tunisia); they have been harvested from four selected trees, peeled then the peel was used as an oil aromatizing agent. Bitter orange skin was composed of two constituents: flavedo and albedo; its essential oil stored in flavedo glands, was easily extracted by mechanic ways and has a very characteristic pleasant odor (Chapot and Praloran, 1955).
2.3. Refined corn oil deodorization Alkali-refined samples were selected owing to their very low volatiles level. They were extracted by dynamic headspace method (Druaux et al., 1998) with splashed gas N2 (Strip-trap) according to published data (Dhifi et al., 2002). In fact, nitrogen gas was bubbled through 40 ml of oil at a flow rate corresponding to a constant pressure of 0.4 bar. The 6-methyl-5-hepten-2-one was added in the sample as an internal standard in order to quantify the aroma residue. Volatile compounds were stripped with nitrogen for 4 h (time fixed after a kinetic survey during 1 h 15 min, 2 h, 3 h and 4 h), trapped on 50 mg of activated charcoal (0.5–0.85 mm, 20–35 mesh ASTM, Merck, Schuchardt, Germany), eluted 120 with 1 ml of diethyl-ether and then analyzed. Oil was maintained at a temperature about 36 ± 1 ◦ C for preserving flavour according to earlier reports (Dhifi et al., 2002). 2.4. Refined corn oil aromatization process The aromatization was used to improve the organoleptic profile of corn oil. For this, deodorized refined corn oil was put in a 500 ml vial and used as lipid matrix. Bitter orange peels were cut into small pieces then homogenized in the oil in an incubator shaker (Excella E-24/24R Benchtop, New Brunswick Scientific CO., INC.) with dynamic plate and hermetical lid. Incubation period (1 h, 2 h and 3 h) and peel quantity (5 g, 10 g and 15 g) were optimized while oil volume (40 ml), incubator temperature (20 ◦ C) and shaking speed (100 rpm) remained constant. After, flavoured oils were filtered (peels were carried out of oil) and an internal standard (6-methyl-5-hepten-2-one) was added. Volatile compounds were extracted using the same headspace parameters of deodorization method since these optimized conditions permitted to obtain an aroma maximum extraction. The aim of the extraction step was to evaluate peel essential oil retention by the lipid matrix. 2.5. Peel essential oil characterization Peel composition was studied in order to confirm later their retention in corn oil. Peel volatiles were extracted by hydrodistillation method. For this, fresh peels (50 g) were immersed in double their volume of distilled water and essential oils were extracted during 90 min; this time was fixed after a kinetic survey during 30 min, 60 min, 90 min and 120 min. Essential oils were recovered from the emulsion with diethyl-ether (v/v) and added with an internal standard, the 6-methyl-5-hepten-2-one, in order to quantify them. After a concentration at 35 ◦ C, using a Vigreux column, essential oils obtained were subsequently analyzed by gas chromatography. All experiments were made in triplicates. 2.6. GC–FID analysis Peel essential oil as well as deodorized and flavoured corn oil volatile compounds, were analyzed by GC using a Hewlett–Packard
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6890 apparatus (Agilent Technologies, Palo Alto, CA, USA), equipped with a flame ionization detector (FID) and an electronic pressure control (EPC) injector. A polyethylene glycol capillary column (HP Innowax, 30 m × 0.25 mm i.d., 0.25 m film thickness) and an apolar HP-5 column (30 m × 0.25 mm, 0.25 m film thickness) were used. The flow of the carrier gas (N2 ) was 1.6 ml/min and the split ratio in the injector was 60:1. The analysis was performed using the following temperature programme: oven temperature isotherm at 35 ◦ C for 10 min, from 35 to 205 ◦ C at the rate of 3 ◦ C min−1 , and isotherm at 205 ◦ C during 10 min. Injector and detector temperatures were held, respectively, at 250 and 300 ◦ C. Surfaces of peaks and percentages of the different compounds were determined using the same HP chemstation cited above. 2.7. GC–MS analysis A HP-5890 series II coupled to a HP-5972 mass spectrometer with electron impact ionization (70 eV) and a HP-5 MS capillary column (30 m × 0.25 mm, 0.25 m film thickness) were used. Column temperature was programmed to rise from 50 to 240 ◦ C at a rate of 5 ◦ C min−1 ; transfer line temperature was 250 ◦ C. The carrier gas was helium with a flow rate of 1.2 ml/min and a split ratio of 60:1. Scan time and mass range were 1 s and 40–300 m/z, respectively.
Fig. 1. Quantitative evaluation of peel essential oil retained in deodorized refined corn oil after different incubation times (1 h, 2 h or 3 h = 1H, 2H or 3H, respectively); total volatile compound contents with different subscripts (a–e) were significantly different at p < 0.05 (Duncan test).
2.10. Statistical analyses Data were subjected to statistical analysis using statistical program package STATISTICA (Statsoft, 1998). Percentage of each volatile compound and fatty acids was the mean of three replicates ± standard deviation and the differences between individual means were deemed to be significant at p < 0.05.
2.8. Compounds identification The identity of the oil components was assigned by comparison of their retention indices relative to (C8–C22) n-alkanes with those of literature or with those of authentic compounds available in our laboratory. Further identification was made by matching their recorded mass spectra with those stored in the Wiley/NBS mass spectral library 167 of the GC–MS data system and other published mass spectra (Adams, 2001). 2.9. Lipid matrix characterization 2.9.1. Fatty acid methyl esters (FAMEs) preparation Crude, degummed, bleached, alkali-refined and flavoured corn oil fatty acids were transformed into their FAMEs after a transmethylation according to the method described by Cecchi et al. (1985), using sodium methylate. An aliquot (50 g of oil) was prepared in a screw tube; then, 2 ml of hexane and 0.5 ml of sodium methylate (3% in methanol) were added. After stirring during 1 min and standing for 2 min, the mixture was neutralized by addition of 0.2 ml of H2 SO4 (1N). After washing the mixture with 1.5 ml of distilled water, the superior phase containing FAMEs was poured off, concentrated under N2 stream and analyzed by gas chromatography (GC). Experiments were made in triplicates. 2.9.2. Fatty acid analysis FAMEs were analyzed by GC, using the same apparatus previously described; the flow of the carrier gas (N2 , U) was 1.6 ml/min and the split ratio was 60:1. The detector and injector temperatures were set at 275 and 250 ◦ C, respectively. The initial oven temperature was held at 150 ◦ C for 1 min−1 , increased at a rate of 15 ◦ C min−1 to 200 ◦ C and then held there for 3 min and finally ramped at 2 ◦ C min−1 for 184–242 ◦ C. Surfaces of peaks and percentages of the different compounds were determined using HP chemstation (Rev.A.0401) software. The latter also permits to control the analytic parameters (flow, temperature, beginning and end of the analyses, etc.). Fatty acids were identified by comparing their retention times with those of authentic standards analyzed in the same conditions.
3. Results and discussion 3.1. Corn oil deodorization The aim of this step was to remove residual volatiles responsible for off-flavours remained at the end of refining process in order to obtain a completely deodorized lipid matrix and consequently to be able to evaluate only peel essential oil quantity retained. Headspace parameters optimization permitted to reach a considerable aroma residue when extraction period was about 1 h 15 min, 2 h and 3 h. However, samples submitted to 4 h extraction showed the best deodorization result with residual aroma of about 3.4% (0.901 g/ml of oil). 3.2. Refined corn oil aromatization Refined oil deodorized during 4 h was used as lipid matrix. Different shaking periods (1 h, 2 h and 3 h) and variable peel quantities (5 g, 10 g and 15 g) were experimented. Temperature of incubator was held at 20 ◦ C because of the essential oil volatility that increases at high temperatures (Weel et al., 2004; De Roos, 2006). Due to a fixed oil volume used (40 ml), peel quantity used did not pass 15 g; In fact, 20 g of peel were experimented and the homogenization was partial. Shaker speed was 100 rpm. GC profiles showed that the percentages of peel essential oil compounds retained were comparable for all the samples studied. Especially hydrocarbon compounds were readily retained on lipid matrix with a large contribution of monoterpene hydrocarbons such as limonene, ␣-pinene, -pinene, sabinene and ␣-terpinene with a total percentage ranging from 96% to 99%. The only oxygenated compound retained was 1,8-cineole with a proportion inferior to 1% (Table 1). In addition, all samples showed high proportions of limonene varying from 92.57% to 96.11%. However, all the other compounds forming the minor fraction of peel essential oils were retained. Peel quantity and incubation time effects on peel total volatiles retention in corn oil were studied. Table 1 and Fig. 1 pointed out the highest contents of some volatiles retained in oils after 1 h of incu-
0.00e 0.00e 0.00f 0.00e 0.01f 0.00e 0.00a 0.01e ± ± ± ± ± ± ± ± 0.006 0.002 0.001 0.01 0.39 0.002 0.01 0.42 ␣-Pinene -Pinene Sabinene ␣-Terpinéne Limonéne 1–8 Cinéole Others Total volatiles
0.01 0.003 0.001 0.02 1.09 0.01 0.03 1.17
± ± ± ± ± ± ± ±
1.10 0.01 ± 0.00abc 1.21 0.00bc 0.30 0.004 ± 0.00de 0.38 0.00de 0.12 0.00bcde 0.13 0.001 ± 0.00def 1.68 0.02 ± 0.00cd 1.49 0.00c de de 93.39 1.09 ± 0.21 95.69 0.32 0.60 0.01 ± 0.00c 0.72 0.00c a a 0.01 2.80 0.004 ± 0.001 0.39 d d 100 1.14 ± 0.17 100 0.27
%
205
bation and mainly limonene. However, samples incubated for 2 h or 3 h showed decrease retention. This could be explained by volatile fraction partial lost when shaking period was long. In addition, the contents of total volatile compounds retained in corn oil were higher when peel quantity was more important. Furthermore, the highest contents of aroma (2.4 mg/ml) and limonene (2.3 mg/ml) were detected in the samples incubated with 15 g of peel for 1 h. The results obtained indicated that peel quantity and incubation time affected aroma retention in the lipid matrix. It is worth noting that this work, to the best of our knowledge, is the first one dealing with aromatization optimization conditions of corn oil by C. aurantium. 3.3. Corn oil fatty acid composition As can be seen in Table 2, fatty acid composition pointed out the abundance of three fatty acids (palmitic C16:0, oleic C18:1 and linoleic C18:2) which accounted 96.92% fraction in crude oil, 97.45% in degummed oil, 96.42% in bleached oil, 96.26% in refined oil and 97.08% flavoured oils. The unsaturated fatty acids (C18:1 and C18:2) were more abundant than the saturated ones with the highest rate of the linoleic acid varying from 56% to 57%, followed by the oleic one ranging from 26.12% to 27.46%. The palmitic acid was present at a percentage varying from 13% to 14.5%. These results showed that there were not significant differences among different samples, permitting to deduce that refining process did not affect fatty acid composition but affected essentially minor compounds of oil such as volatiles, pigments and phenols, as indicated in previous work (Lanzon et al., 1994). In addition, aromatization process did not also change fatty acid composition but improved oil quality. 3.4. Peel essential oil composition
Values followed by the same small letter did not share significant differences at 5% (Duncan test).
% mg/ml %
1.49 0.02 ± 0.00a 0.9 0.01 ± 0.00ab 1.06 0.01 ± 0.00abc 1.26 0.01 ± 0.00cd 0.44 0.01 ± 0.00bc 0.68 0.01 ± 0.00de 1.23 0.55 0.01 ± 0.00bc 0.37 0.004 ± 0.00de 0.29 0.01 ± 0.00a 0.89 0.01 ± 0.00cde 0.20 0.01 ± 0.00cd 0.32 0.01 ± 0.00ab 1.35 0.20 0.002 ± 0.00a 0.11 0.001 ± 0.00cde 0.10 0.002 ± 0.00abc 0.17 0.001 ± 0.00ef 0.05 0.002 ± 0.00abcd 0.10 0.002 ± 0.00ab 0.33 1.16 0.03 ± 0.00b 1.84 0.02 ± 0.00c 1.51 0.01 ± 0.00de 1.07 0.04 ± 0.01a 1.81 0.03 ± 0.00b 1.68 0.01 ± 0.00e 1.62 b cd e a bc f 93.48 1.82 ± 0.15 95.75 1.39 ± 0.18 95.79 1.01 ± 0.08 92.83 2.31 ± 0.50 95.91 1.65 ± 0.02 95.20 0.55 ± 0.04 92.58 0.39 0.01 ± 0.00b 0.71 0.01 ± 0.00b 0.92 0.004 ± 0.00d 0.38 0.02 ± 0.00a 0.86 0.01 ± 0.00b 0.69 0.00 ± 0.00de 0.53 a a a a a a 2.74 0.006 ± 0.00 0.34 0.005 ± 0.00 0.33 0.04 ± 0.00 3.41 0.02 ± 0.01 0.73 0.02 ± 0.00 1.32 0.01 ± 0.00 2.36 b cd d a bc e 100 1.90 ± 0.12 100 1.45 ± 0.15 100 1.09 ± 0.06 100 2.40 ± 0.41 100 1.73 ± 0.01 100 0.59 ± 0.03 100
3 h (incubation time) 2 h (incubation time)
mg/ml %
1 h (incubation time)
mg/ml % mg/ml % mg/ml % mg/ml % mg/ml mg/ml % mg/ml
10 g (peel quantity)
1 h (incubation time) 3 h (incubation time) 2 h (incubation time) 1 h (incubation time)
5 g (peel quantity) Compounds
Table 1 Contents (mg/ml of oil) and percentages (%) of peel volatile retained in deodorized corn oil.
2 h (incubation time)
3 h (incubation time)
15 g (peel quantity)
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Peel essential oil composition was determined and permitted to conclude on a large contribution of terpenes (2.71 mg/g of peel). The major monoterpene hydrocarbons were represented by limonene, ␣-pinene, -pinene, sabinene and ␣-terpinene with a total percentage of about 92.8%. These results are in agreement with those found by Dugo et al. (1993). The limonene importance in citrus peel has been confirmed by several studies; its percentage varied from 90% to 95% (Dugo et al., 1993; Saidani and Marzouk, 2003) which is in conformity with our result (90.25%). Limonene is one of the most common terpenes in nature and is the majority constituent of an essential oil series. Its pleasant citric fragrance is commonly used as a flavouring in foods and drinks, for which it is classified in the U.S. Code of Federal Regulation as safe. Tests in animals have proven the effectiveness of limonene against some types of cancer including gastric, mammary, pulmonary adenoma and liver. Limonene also has been shown to be effective in relieving gastroesophageal reflux disorder and occasional heartburn (Thiago et al., 2009). 3.5. Evaluation of lipid matrix and aroma interaction To evaluate volatile compounds retention by lipid matrix during the aromatization experiments, peel essential oil composition was used as a reference, after their extraction by hydrodistillation using different peel quantities (5 g, 10 g and 15 g). A great interest was given to only compounds retained. Based on the results presented in Figs. 2 and 3, the quantity of total volatile compound retained was considerable in the major part of samples when headspace method and oil matrix were used. This could be explained by the high hydrophobic matrix used and the relatively lipophilic character of the most aroma compounds that leads to a higher affinity of these latter to corn oil. These results were in conformity with those found by Piraprez et al. (1998). Fur-
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Table 2 Fatty acid composition (%) of corn oil during refining and aromatization steps. Fatty acid % C16:0 C16:1 C18:0 C18:1 C18:2 C18:3 C20:0 C20:1
Crude oil 13.4 0.07 1.52 27.46 56.06 0.87 0.28 0.14
± ± ± ± ± ± ± ±
Degummed oil c
0.6 0.01h 0.1d 1.20b 1.9a 0.06e 0.01f 0.05g
14.46 0.08 1.10 26.12 56.87 0.9 0.23 0.16
± ± ± ± ± ± ± ±
c
0.9 0.03g 0.2d 0.93b 2.4a 0.02d 0.03e 0.01f
Bleached oil 13.19 0.13 1.35 26.20 57.03 0.98 0.29 0.21
± ± ± ± ± ± ± ±
c
0.5 0.01g 0.3d 1.5b 1.8a 0.20d 0.0e 0.03f
Refined oil 13.81 0.09 1.49 26.36 56.09 1.23 0.21 0.18
± ± ± ± ± ± ± ±
Flavoured oil c
1.3 0.05h 0.1d 1.2b 2.2a 0.05e 0.04f 0.02g
14.55 0.06 1.22 26.33 56.20 1.00 0.27 0.1
± ± ± ± ± ± ± ±
1.53c 0.00 0.02d 1.02b 1.6a 0.13e 0.06f 0.01
Values followed by the same small letter did not share significant differences at 5% (Duncan test).
92.57% to 96.11%. Fatty acid composition analysis showed nearly the same proportions after all the refining steps and after aromatization. The total quantity of peel volatile compounds retained was considerable when headspace method and oil matrix were used with comparison to hydrodistillation method. These results incite us to pursue this research by determining the antioxidant and antiradical potentialities of flavoured corn oil and the maximum aroma conservation period in this oil.
References Fig. 2. Peel volatile compounds quantity extracted by hydrodistillation; total volatile compound contents with different subscripts (a–c) were significantly different at p < 0.05 (Duncan test).
Fig. 3. Peel volatile compounds quantity in corn oil extracted by headspace after different incubation times (1 h, 2 h or 3 h = 1H, 2H or 3H, respectively); total volatile compound contents with different subscripts (a–g) were significantly different at p < 0.05 (Duncan test).
thermore, Seuvre et al. (2006), studying aroma retention in water and oil, pointed out aroma retention by lipids and indicated that due to the hydrophobic character of major volatile compounds, polar ones were the most soluble in water; this was in conformity with our results. Headspace system is a method using a cold extraction at a temperature of about 36 ± 1 ◦ C that permits an important retention of non-polar compounds which constituted the largest part of retained volatiles. 4. Conclusion Aroma analysis indicated that peel compounds especially monoterpene hydrocarbons such as limonene, ␣-pinene, -pinene, sabinene and ␣-terpinene were retained in deodorized refined corn oil with a total percentage ranging from 96% to 99%. Limonene was the main compound detected with a percentage varying from
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