Determination of Volatile Compounds in Apple Pomace by Stir Bar ...

C: Food Chemistry

Determination of Volatile Compounds in Apple Pomace by Stir Bar Sorptive Extraction and Gas Chromatography-Mass Spectrometry (SBSE-GC-MS) Roberto Rodr´ıguez Madrera and Belén Suárez Valles

Abstract: A method based on stir bar sorptive extraction combined with gas chromatography-mass spectrometry detection (SBSE-GC-MS) has been optimized with the aim of applying it to the analysis of apple pomace. The method allowed the identification of 124 volatile compounds after 3 h of extraction with a precision (RSDs) ranging between 2% (linalool) and 11% (ethyl hexanoate). Its use in analyzing varietal apple pomace revealed the interest of this substrate as regards its content in aromas. From a semi-quantitative point of view, the higher content in aldehydes and esters of the Blanquina variety is worth highlighting, as are the greater concentration of acids in the Clara variety and the higher content of terpenes and norisoprenoides in the Coloradona variety. In contrast, the Ernestina and Perico varieties presented the lowest levels of aromas. Keywords:

apple, aroma, GC-MS, pomace, SBSE

Practical Application: The analysis of varietal apple pomaces showed the importance of this type of waste in the food

industry, both for its content in aromas such as for its use as substrate in biotechnological processes.

Introduction

(Regulation [EC] nr 110/2008 of the European Parliament of the Council). On the other hand, the flavors used in the agrifood industry represent around 25% of the food additives market (Longo and Sanromán 2006). In this respect, apple pomace could be an interesting product for the sector on account of its content in aromas. In consequence, a method for producing aromas via biotechnological procedures has been proposed (Almonsino and others 1996). The volatile fraction of fruits is formed by compounds belonging to different chemical families such as alcohols, acids, esters, carbonylic compounds, or terpenes, among others. Various analytical methods are used to study aromas in solid matrices of this kind, generally consisting of a stage of extraction or concentration of the aromas followed by a gas chromatographic analysis. Thus, classic methods such as extraction-distillation, the principal disadvantages of which are the use of organic solvents and tedious sample preparations with possible formation of several artifacts, have sometimes been used (Ruberto and others 2008). In other cases, methods based on headspace analysis (dynamic or static) and purge and trap, whose principal disadvantage is the limitation of use to the most volatile compounds, have been proposed (Rowan and others 2009). One of the most successful techniques for analyzing aromas in foods has been solid phase microextraction (SPME). SPME employs a fused silica fiber coated with one or more polymers capable of retaining the analytes of interest by absorption and/or adsorpMS 20110659 Submitted 5/26/2011, Accepted 8/11/2011. Authors are with tion. Subsequently, the retained aromas are thermally desorbed ´ Area de Tecnolog´ıa de los Alimentos, Servicio Regional de Investigación y Desarrollo Agroalimentario (SERIDA), 33300-Villaviciosa, Asturias, Spain. Direct inquiries to for analysis by means of chromatographic techniques. SPME has several advantages, such as little or no sample preparation, the abauthor Rodr´ıguez Madrera (E-mail: [email protected]). sence of organic solvents and good reproducibility. Nevertheless,

Cider-making is one of the main agrifood industries in Asturias (northern Spain) with an annual average production of over 50 million liters, its principal residue being pomace (the mass resulting from the pressing of the apple to obtain juice). Although the amount of the extracted juice varies in relation to several factors such as apple variety, state of ripeness, type of pressing or the application of different enzymatic treatments, a must yield of around 65% to 70% (v/m) is usually estimated. This results in a bulky residue with a great potential for application. A number of different uses for this residue have been proposed: in the synthesis of pectolytic enzymes (Berovic and Ostroversnik 1997) as a substrate for the production of fructofuranosidase (Hang and Woodams 1995), as an important source of antioxidant polyphenols (Diñeiro Garc´ıa and others 2009) or in the extraction of pectin (May 1990), this last use being the only industrial application to date (Gullón and others 2007). Moreover, one of the usual agrifood-processing uses of fruit pomaces is the production of spirits according to traditional methods, which gives rise to quality products of recognized prestige. Fruit marc spirits have a high alcoholic strength and are characterized by notable levels of aroma compounds recovered during the distillation of the fermented pomace

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Journal of Food Science r Vol. 76, Nr. 9, 2011

R C 2011 Institute of Food Technologists doi: 10.1111/j.1750-3841.2011.02406.x

Further reproduction without permission is prohibited

the limited amount of stationary phase on the fused silica fiber means it is not sensitive enough on occasions (Kataoka and others 2000). Baltussen and others (1999) introduced stir bar sorption extraction (SBSE), a variation on SPME consisting of the use of magnetized bars covered with an absorbent polymer. SBSE presents the same advantages as SPME, but in addition its sensibility increases around 100-fold because it uses a greater amount of stationary phase. SBSE has been successfully applied in recent years in the analysis of aromas in different matrices of vegetable origin (Bicchi and others 2003; Salinas and others 2004; Malowicki and others 2008). The aim of the present study was to develop an analytical method based on SBSE followed by gas chromatographic analysis and mass spectrometry (GC-MS) and to apply it to the study of the aromas in apple pomace.

Material and Methods

splitless mode raising the temperature from –40 to 320 ◦ C at a rate of 10 ◦ C/s. The gas chromatograph was an Agilent 7890A equipped with an MSD 5975C (Agilent Technologies, Palo Alto, Calif., U.S.A.). The column employed was a FFAP (50 m × 0.2 mm i.d., 0.25 μm, Agilent Technologies). The oven temperature program was as follows: 40 ◦ C (5 min) rising to 220 ◦ C (25 min) at a rate of 3 ◦ C/min. Helium was used as carrier gas at a flow rate of 1 mL/min. Identification was carried out by comparing spectra with those registered in a Wiley 138K mass spectral library and linear retention indices and confirmed by pure standards, when available. Analyses were carried out in SIM mode and the results were expressed as microgram of internal standard (2-octanol) per kilogram of wet pomace.

Statistical analysis The influence of the factors studied in the optimization of the extraction method (stirring time and ionic strength) on the recovery of aromas was evaluated by means of a t-test for independent samples. The program used was SPSS version 15.0.

Standards All standards were of analytical quality, with at least 98% purity. The standard working solutions were prepared by dilution of compounds in water. Ultra pure water was obtained from a Milli-Q Results and Discussion system from Millipore (Mildford, Conn., U.S.A.). Optimization of the extraction method The efficiency of the extraction by SBSE is affected by different Samples variables: extraction time, stirring speed, and temperature, addition A sample of multi-varietal apple pomace from an industrial cellar of inert salts or other modifiers such as methanol, pH, quantity, and 5 mono-varietal apple pomaces from cider apples belonging and dilution of the sample, among others (Prieto and others 2010). to different cider apple categories: Blanquina (acid), Clara (bitter), In addition, to obtain effective, reproducible extraction with stir Coloradona (sweet-bitter), Ernestina (sweet), and Perico (semiacid), harvested in the year 2007, were analyzed. To facilitate conservation, all the pomaces (composed of skin, Dry pomace pulp and seeds) were dried until constant weight at 60 ◦ C for 48 h. Pomace moisture content ranged between 70% and 76%. Rehydration

Aroma extraction Figure 1 shows the process used to obtain the extract described subsequently. At the time of carrying out the analysis, dried pomace was rehydrated and mixed with double its amount of water to obtain so-called prehomogenized pomace. An aliquot of this sample (4.5 g) was then transferred to a 50 mL vial containing 50 mg of ascorbic acid and 2.8 ng of 2-octanol (internal standard, IS) and 30 mL of water. The mixture was homogenized in a Polytron PT 10–35 (Kinematica AG, Littau, Switzerland) during 2 min. Then the polydimethylsiloxane coated stir bar (0.5 mm film thickness, 20 mm length, Twister, Gerstel GmbH & Co., Mülheim an der Ruhr, Germany) was added and the extraction was performed at room temperature, stirring at 700 rpm for 3 h. After the extraction, the stir bars were rinsed with distilled water, dried with a cellulose tissue and then placed in desorption tubes. All analyses were carried out in triplicate.

Pomace-Water (1:3) 30 min

Wet pomace

Pomace-Water (1:2) Crushed 3 min

Pre-homogenized pomace

Homogenization Ascorbic acid

Pomace-Water (3:20)

I.S.

2 min

Chromatographic analysis Stir bar The desorption tubes were placed in a thermal desorption unit (TDU, Gerstel GmbH & Co.) and the stir bars were desorbed Extraction (3h) from 25 ◦ C (1 min) to 295 ◦ C (10 min) at a rate of 60 ◦ C/min in solvent vent mode (helium flow: 50 mL/min). The desorbed aromas were cryofocussed in a CIS-4 programmable temperature Chromatographic analysis vaporizing (PTV) inlet (Gerstel GmbH & Co.) at –40 ◦ C using liquid nitrogen, the inlet liner being packed with quartz wool. Analytes were transferred into the chromatographic column in Figure 1–Extraction procedure. Vol. 76, Nr. 9, 2011 r Journal of Food Science C1327

C: Food Chemistry

Aromas in apple pomace by SBSE-GC-MS . . .


C: Food Chemistry

bars, the sample mixture must be well homogenized and in liquid form, which favors the distribution of the analytes between the matrix and the stationary phase. The fact that the sample is made up of skin, pulp, and seeds, which have dissimilar compositions, means that the pomace must be properly homogenized to guarantee adequate sampling. For this reason, subsequent to prior testing, it was decided to add double the amount of water to the wet pomace followed by a first crushing, thereby obtaining a homogeneous, compact puree (prehomogenized pomace), thus favoring the subsequent handling of the sample (Figure 1). Water was then added to aliquots of prehomogenized pomace and the particle size was reduced in a Polytron PT 10–35. This favored the suitable transfer of the analytes to the aqueous phase for their subsequent extraction. Furthermore, the stirring of the bar was uniform, thus increasing extraction reproducibility. This prior treatment was found to be indispensable to obtain homogeneous samples and prefixed the level of dilution and the amount of sample (pomace : water, 3 : 20). On the other hand, desorption of the analytes extracted with the stir bar should be complete to ensure good reproducibility of the method and to avoid the existence of a memory effect in successive extractions. Taking into account the compounds present in the sample and the limitations of use of stir bars, a maximum desorption temperature of 295 ◦ C during 10 min was chosen. When the stir bars were desorbed a second time under these conditions, the chromatogram only showed peaks due to the polydimethylsiloxane (PDMS) film. However, as no volatiles from the pomace were detected, this allowed us to discard any memory effect and avoid preconditioning of the stir bar between successive extractions. Using stir bars causes their physical degradation, with losses of PDMS, and the deposition of non-volatile substances. This diminishes the extraction capacity and leads to a lack of reproducibility between bars depending on the number of extractions that have been carried out. To avoid this drawback, it was decided to use an internal standard. 2-Octanol was thus selected taking into account its absence in the samples and the PDMS-water distribution coefficient (log KO/W = 2.72), which favors an effective extraction (David and others 2003). The extraction of this compound under the final conditions was good, with a reproducibility of 5%, while the relative areas to the internal standard compensated the loss of reproducibility when the bars began to deteriorate, thus extending their useful life. Bearing in mind previous studies based on SBSE in similar matrices, it was decided to perform the extractions at room temperature (22 ± 2 ◦ C) at a stirring speed of 700 rpm (Bicchi and others 2003; Salinas and others 2004). The influence of the other parameters studied (stirring time and ionic strength) was determined by comparing the relative areas to the internal standard for the chromatographic peaks. To avoid interferences in the estimation of the peak areas, SIM mode was used, employing characteristic m/z relationships for each compound. To evaluate the influence of stirring time, relative areas from 2 extraction times (1 and 3 h) were compared for 16 analytes: ethyl hexanoate, methyl octanoate, ethyl decanoate, 2-phenylethyl acetate, amyl alcohols, 1-octanol, 2-phenylethanol, linalool, farnesol (isomer 2), α-farnesene, β-ionone, acetophenone, 6-methyl-5-heptenone, γ -decalactone, octanoic acid, and decanoic acid, covering the entire volatile range and the distinct chemical families (Figure 2a and 2b). The results showed different behaviors for the analytes under study. A significant increase in extraction (P < 0.05) was detected C1328 Journal of Food Science r Vol. 76, Nr. 9, 2011

after 3 h for ethyl decanoate, 1-octanol, farnesol, α-farnesene, linalool, and β-ionone, while a significant decrease (P < 0.05) in extraction efficacy was only detected for ethyl hexanoate and 2-phenylethyl acetate. No significant differences (P > 0.05) were found for the rest of the aromas studied as regards extraction time. It was also noted that a longer extraction time (3 h) favored equilibrium in the distribution of analytes between the 2 phases and improved extraction reproducibility. Under these conditions, the reproducibility of the method ranged between 2% (linalool) and 11% (ethyl hexanoate), values similar to those obtained with SBSE in other matrices (Salinas and others 2004; Gomes Zuin and others 2005). Subsequently, the influence of ionic strength on extraction efficacy was evaluated by adding 3 g NaCl and stirring for 3 h. In this case, a significant increase was only detected in the extraction (P < 0.05) of ethyl hexanoate, methyl octanoate, linalool, and 2-phenylethyl acetate, which could be due to a reduction in their solubility in the aqueous phase (Giordano and others 2009). In contrast, a significant decrease in extraction (P < 0.05) was detected for α-farnesene, farnesol, β-ionone, 1-octanol, amyl alcohols, γ -decalactone, and ethyl decanoate, which could have different causes: the occupation of the surface of the PDMS by salt ions (Portugal and others 2008) or an “oil effect,” via which the presence of a salt in solution promotes the movement of non-polar compounds to the surface of the aqueous phase, thus minimizing their interaction with the PDMS (Gomes Zuin and others 2005). As can be seen, the use of NaCl did not improve the extraction of the analytes or the detection of new ones, so it was decided not to use NaCl. The analytical method allowed us to detect and identify 124 volatile compounds in apple pomace samples (Table 1), belonging to different chemical families, and found in the aroma of apples by other methods (Young and others 2004; López and others 2007; Xiaobo and Jiewen 2008; Reis and others 2009; Rowan and others 2009). Taking into account our interest in applying the analytical method to study the volatile compounds in apple pomace before and after alcoholic fermentation, known amounts of 9 aroma components (1-octanol, linalool, methyl octanoate, ethyl decanoate, acetophenone, decanoic acid, γ -decalactone, 2-phenylethyl acetate, and 6-methyl-5-hepten-2-one), representing the different chemical families, were added at 2 levels of addition (Table 2). The standard additions were extracted under the optimized conditions. The results for both matrices showed no significant differences (P > 0.05) between the slopes of the regression lines, with a suitable linearity (R2 > 0.99). Thus, systematic errors such as the matrix effect may be discarded.

Apple pomace analysis The optimized method was applied to the analysis of 5 apple pomaces, all derived from mono-varietal mixtures of cider apples. Figure 3 shows the chromatogram obtained for the Perico variety. Semi-quantitative analysis was used for determination of the volatile compounds in apple pomace. Concentrations were calcuar ea lated as follows: P eISa k area × IS concentration. Total of 33 relevant compounds from an odorant and technological standpoint from different chemical families were semi-quantified (Table 3). As can be seen, 10 carbonylic compounds were semi-quantified. Worth highlighting among these because of their organoleptic relevance are the aliphatic aldehydes (2-heptenal, 2-octenal, 2-nonenal, and 2.4-decadienal isomers), responsible for herbaceous odors as observed in grape pomace distillates by Williams

and Strauss (1978). The presence of unbranched chain aldehydes and ketones is associated with the action of different enzymes on apple polyunsaturated fatty acids, mainly linoleic and linolenic acids (Fauconnier and Marlier 1997). However, the most abundant carbonylic compound in all cases was benzaldehyde, with a strong smell of bitter almonds, derived from the hydrolysis of the amygdalin present in the seeds that form part of the apple pomace. Free fatty acids are considered as the main precursors of esters, alcohols and volatile aldehydes in apple (Fellman and others 2000). Octanoic, nonanoic, decanoic, and dodecanoic acids were detected in all the analyzed samples, decanoic acid being the acid extracted in greater amounts in all cases. On the other hand, the

highest concentrations of these acids were detected in the Clara variety. Another important group of aroma compounds are esters, whose presence in apples is due to the esterification of the corresponding alcohols and carboxylic acids through the action of an alcohol acyl-CoA transferase (Fellman and others 2000). Esters are usually associated with fruity odors and noticeable differences between pomaces were observed in this study. The pomaces of the Blanquina variety were found to present a major abundance of esters in general, except for ethyl butyrate, more abundant in the Coloradona variety (Table 3). Furthermore, all the samples showed the presence of γ -decalactone, a compound responsible

Figure 2–(A) Influence of time and ionic strength on the extraction of aromas in apple pomace (mean and standard deviation). (B) Influence of time and ionic strength on the extraction of aromas in apple pomace (mean and standard deviation).

Vol. 76, Nr. 9, 2011 r Journal of Food Science C1329

C: Food Chemistry



Table 1–Volatile compounds detected in apple pomace by SBSE-GC-MS.

C: Food Chemistry

Alcohols Amyl alcohols 1-Hexanol 1-Octen-3-ol 1-Octanol Z-2-octen-1-ol 1-Decanol Phenylmethanol 2-Phenylethanol Aldehydes and ketones Hexanal Heptanal 2-Hexenal Octanal 1-Hydroxy-2-propanone 1-Octen-3-one 2-Heptenal 6-Methyl-5-hepten-2-one 2,6-Dimethyl-5-heptenal 3-Methyl-2-cyclopenten-1-one 2-Nonanone Nonanal 2-Octenal 2-Decanone (E,E)-2,4-heptadienal Decanal 3,5-Octadien-2-one Benzaldehyde 2-Nonenal (E,E)-2,4-octadienal 6-Methyl-3,5-heptadien-2-one 2-Undecanone (Z)-2-decenal Acetophenone (E,E)-2,4-nonadienal 2-Undecenal (E,Z)-2,4-decadienal (E,E)-2,4-decadienal 3-Phenylpropenal Benzophenone Acids Acetic acid Propanoic acid Crotonic acid Hexanoic acid Octanoic acid Nonanoic acid Decanoic acid Benzoic acid Dodecanoic acid Terpenes and norisoprenoids Linalool β-cyclocitral Safranal β-farnesene β-citral isomer 1 β-citral isomer 2 Camphene α-farnesene β-damascenone Nerylacetone Nerolidol isomer 1 6,10-Dimethyl-5,9-undecadien-2-ol β-ionone Nerolidol isomer 2 Pseudoionone isomer 1 Pseudoionone isomer 2 2-Methyl-6-methylene-1,7-octadien-3-one Farnesol isomer 1 Farnesol isomer 2

Retention time

Linear retention index

Identification

Descriptor

22.53 29.37 33.65 38.21 40.80 46.19 50.60 51.66

1145 1276 1372 1482 1516 1683 1806 1834

1.2 1.2 1.2 1.2 1.2 1.2 1.2 1.2

malt burnt herbaceous mushroom oily fatty green fat floral rose floral

16.99 22.19 23.68 27.18 27.34 27.65 28.79 29.21 30.12 30.80 31.80 31.9 33.55 36.00 36.22 36.64 37.50 37.54 38.06 40.16 40.26 40.65 42.43 42.62 44.53 46.49 46.85 48.46 56.49 70.45

1043 1139 1165 1231 1234 1240 1264 1272 1291 1306 1329 1333 1368 1428 1433 1444 1465 1466 1478 1527 1530 1538 1578 1582 1633 1692 1703 1747 1968 2403

1.2 1.2 1.2 1 1.2 1.2 1 1.2 1 1 1.2 1.2 1 1 1 1 1 1.2 1 1 1 1.2 1.2 1.2 1 1 1 1 1.2 1

grass fat citrus green. apple green fat cheese mushroom almond fruity green fat melon green fruity green green fat green walnut orange nuts fat orange peel tallow fruity mushroom almonds cucumber fat cucumber cinnamon coconut orange green tallow floral almonds green fat sweet pungent fried fried balsamic

34.10 37.28 43.65 49.00 56.18 59.54 62.77 67.96 69.44

1382 1459 1607 1762 1959 2063 2165 2331 2374

1.2 1.2 1 1.2 1.2 1.2 1.2 1.2 1.2

vinegar pungent rancid burnt sweat sweat cheese green fat rancid fat urine fat

37.80 41.64 42.70 43.54 43.75 45.58 45.83 46.61 48.80 49.99 51.46 52.98 53.13 55.83 56.43 59.28 61.47 64.25 65.52

1472 1560 1526 1604 1610 1665 1672 1696 1757 1790 1829 1869 1873 1949 1967 2054 2124 2213 2255

1.2 1 1 1 1 1 1 1 1.2 1 1 1 1 1 1 1 1 1.2 1.2

floral lavender mint herbaceous sweet citric sweet wood lemon lemon camphor wood sweet apple roses honey floral wood floral fruity sweet violet raspberry wood floral floral floral (Continued)

C1330 Journal of Food Science r Vol. 76, Nr. 9, 2011

Aromas in apple pomace by SBSE-GC-MS . . . abundant in the apple pomace extracts, its content in the pomace of the Coloradona variety being noteworthy. It should also be noted that this compound not only has a relevant aroma contribution (floral odor), but that it presents significant antimicrobial activity against human and plant pathogens (Hornby and others 2001; Jabra-Rizk and others 2006). Moreover, other terpenes such

Table 1–Continued

Esters Isoamyl acetate Isobutyl 2-methylbutanoate Ethyl hexanoate Hexyl acetate Isoamyl 2- methylbutanoate 2-Methylbutyl 2- methylbutanoate Z-3-hexen-1-yl acetate Hexyl propanoate Methyl octanoate Butyl hexanoate Hexyl butanoate Hexyl 2- methylbutanoate Ethyl octanoate Isoamyl hexanoate Z-3-hexen-1-yl 2- methylbutanoate Ethyl nonanoate Methyl decanoate Hexyl hexanoate Methyl benzoate Ethyl decanoate Diethyl succinate Ethyl benzoate Ethyl 9-decenoate Phenylmethyl acetate Ethyl phenylacetate Methyl salicylate Methyl dodecanoate 2-Phenylethyl acetate Ethyl 2,4-decadienoate Ethyl dodecanoate Isoamyl decanoate Butyl benzoate 2-Phenylethyl propanoate Isoamyl ethyl succinate 2-Phenylethyl isobutanoate 2-Phenylethyl butanoate Isoamyl phenylacetate E-11,13-dimethyl-1,2-tetradecen-1-yl acetate Ethyl tetradecanoate Hexyl benzoate Ethyl cinnamate 2-Phenylethyl hexanoate Ethyl hexadecanoate Ethyl 9-hexadecenoate Ethyl octadecanoate Ethyl oleate Methyl linoleate Ethyl linoleate Lactones γ -butyrolactone 3-Methyl- γ -butyrolactone γ –decalactone γ –dodecalactone Others Pentylbenzene Furfural 5-Methylfurfural Furfuryl alcohol Naphtalene 2-Methylbenzotiazol

Retention time

Linear retention index

Identification

Descriptor

18.76 21.72 24.42 26.26 26.75 26.93 28.22 29.71 31.71 32.85 33.02 33.26 33.87 34.97 35.37 38.16 40.48 41.22 41.45 42.24 43.03 43.19 44.06 45.32 47.39 47.70 48.38 48.60 49.67 49.70 50.30 50.45 51.30 51.30 53.71 54.06 54.89 56.97 57.01 57.59 59.17 60.42 63.44 64.05 70.19 70.65 70.88 72.41

1077 1130 1178 1212 1222 1226 1252 1283 1327 1353 1357 1363 1377 1403 1413 1481 1535 1551 1556 1574 1592 1595 1619 1657 1718 1726 1745 1751 1781 1781 1798 1802 1824 1825 1888 1897 1921 1982 1983 2000 2051 2091 2187 2206 2396 2409 2416 2461

1.2 1 1.2 1.2 1 1 1.2 1 1.2 1 1.2 1 1.2 1 1 1 1 1.2 2 1.2 1.2 1.2 1 1.2 1 1 1 1.2 1 1.2 1 1 1 1 1 1 1 1 1.2 1 1.2 1 1.2 1 1 1 1 1.2

banana fruity apple peel fruit herbaceous apple apple herbaceous fruity orange fruity fruity strawberry fruity fat fruity herbaceous sweet mulberry wine orange peel peach lettuce raisin grape wine fruity floral camomile floral fruity sweet mint coconut fat roses honey pear leaves brandy coconut balsamic fruity rose fruity rose floral rose honey ether woody balsamic cinnamon honey banana pineapple fat mild waxy fat

41.86 45.20 59.73 66.65

1565 1654 2069 2292

1.2 1 1.2 1.2

caramel sweet peach fruity sweet

33.15 34.76 39.42 42.80 46.30 72.21

1360 1398 1511 1586 1687 2455

1 1.2 1.2 1.2 1 1

caramel burnt sugar caramel burnt tar -

1 Identification by linear retention indices and/or Mass Spectrum (quality > 85). 2 Confirmed with pure standards.


C: Food Chemistry

for the peach aroma. Lactones impart characteristics fruity odors, on account of which these compounds are of major interest to the industry as flavorings (Albrecht and others 1992). Some of the constituent of vegetables with major aromatic potential are terpenes and norisoprenoids. From the 8 analyzed compounds belonging to this group, an isomer of farnesol was the most

Aromas in apple pomace by SBSE-GC-MS . . . different microorganisms (Fisher and others 1999) through the action of lipoxygenases on linoleic and linolenic acids (Manzel and Schreier 2007).

C: Food Chemistry

as linalool, ß-cyclocitral, and citral, and norisoprenoids such as ßionone and ß-damascenone, all of which are responsible for fruity and floral aromas, were detected in all the samples (Guillot and others 2006). In agreement with other studies on apple aroma (Rowan and others 2009; López and others 2007), alcohols constituted the group of least relevance. Although the principal pathway of the formation of alcohols in food products is the fermentation of carbohydrates and amino acids, in the case of some alcohols such as 1-hexanol, which is responsible for herbaceous aromas, their presence in apple products is varietal (Young and others 1996). 1-hexanol, 1-octanol and 1-octen-3-ol were the most abundant alcohols in all cases. In this respect, it is important to emphasize that 1-octen-3-ol (intense mushroom-like odor) is considered an off-flavor in apple juice and its presence is usually associated with

Conclusions The proposed analytical method has been successfully used to determine volatile compounds in apple pomace, allowing the detection of 124 compounds, 33 of which belonging to different chemical families were semi-quantified. The reproducibility of the method ranged between 2% (linalool) and 11% (ethyl hexanoate). Its use in the analysis of varietal apple pomaces revealed the interest of this substrate as regards its content in aromas. From a qualitative and semi-quantitative point of view, carbonylic compounds and esters were the most important in all the cases.

Table 2–Regression lines in fermented and unfermented pomaces from standard additions.

1-Octanol Linaool Methyl octanoate Ethyl decanoate 2-Phenylethyl acetate 6-Methyl-5-hepten-2-one Acetophenone γ -decalactone Decanoic acid

Unfermented pomace

Fermented pomace

Addition m/z

range (mg/L)

slope

intercept

r2

slope

intercept

r2

84 93 74 88 104 55 105 85 60

0 to 0.282 0 to 0.267 0 to 098 0 to 0557 0 to 0352 0 to 0397 0 to 0165 0 to 0150 0 to 3731

0.328 0.575 9.436 2.148 3.815 0.414 0.833 6.972 0.150

0.006 0.009 0.004 0.001 0.072 0.031 0.009 0.055 0.263

0.999 0.999 1.000 1.000 0.997 0.997 0.997 0.992 0.994

0.328 0.572 9.510 2.097 3.790 0.414 0.895 6.480 0.152

0.015 0.003 0.022 0.549 0.042 0.027 0.005 0.037 0.980

0.997 1.000 1.000 0.996 0.999 0.998 1.000 0.998 0.990

Abundance

I.S.

6

22

11+13

16

9000000

8000000

5

7000000

6000000

7

5000000

25

8 28

4000000

29 3000000

10

2000000

4

1000000

15.00

20.00

25.00

26 1

30.00

20 32 9

31

17

24

21

30

3

2

Time-->

15

23

14 33

27 19

18 12

35.00

40.00

45.00

50.00

55.00

60.00

65.00

Figure 3–Chromatogram obtained with the selected extraction conditions for the apple pomace from the Perico variety displaying the analyzed aromas. I.S. Internal standard (2-octanol). Peak references are in Table 3.



Alcohols 1-Hexanol 1-Octen3–ol 1-Octanol Aldehydes and ketones 2-Hexenal 2-Heptenal 2-Octenal 2-Nonenal E,Z-2,4-decadienal E,E-2,4-decadienal 6-Methyl-5-hetpen-2-one 3,5-Octadien-2-one 6-Methyl-3,5-heptadienone Benzaldehyde Acids Octanoic acid Nonanoic acid Decanoic acid Dodecanoic acid Terpenes y norisoprenoids Linaool β-cyclocitral citral β-damascenone nerylacetone β-ionone Pseudo-ionone Farnesol (isomer 2) Esters and lactones Methyl octanoate Methyl decanoate Ethyl octanoate Ethyl decanoate Ethyl benzoate Hexyl butyrate 2-Phenylethyl acetate γ -decalactone

Id

m/z

Blanquina (acid)

Clara (bitter)

Coloradona (sweet-bitter)

Ernestina (sweet)

Perico (semi-acid)

1 2 3

56 57 84

38.84(0.78) 462.00(22.00) 57.83(4.01)

97.87(0.81) 147.11(13.33) 60.66(4.31)

65.46(5.34) 93.32(4.20) 24.74(1.16)

102.22(6.96) 66.65(2.54) 20.90(0.98)

34.22(1.33) 185.57(17.58) 27.00(3.88)

4 5 6 7 8 9 10 11 12 13

83 55 83 70 81 81 55 95 109 106

3.35(0.16) 543.89(4.04) 548.51(25.80) 156.92(0.52) 518.18(7.13) 325.13(8.29) 66.46(0.32) 26.92(0.12) 52.27(0.80) 3074.58(60.16)

9.06(0.24) 165.00(8.84) 331.53(3.53) 162.8(5.41) 297.06(20.20) 286.08(15.22) 128.65(0.69) 43.57(0.82) 147.81(1.43) 2274.46(32.53)

7.93(0.33) 124.79(5.37) 224.87(0.19) 116.81(2.30) 213.10(6.69) 207.98(9.07) 180.26(2.61) 28.17(1.00) 100.39(4.02) 1001.03(114.12)

3.33(0.23) 118.33(2.63) 192.55(2.41) 75.26(0.27) 193.73(0.77) 177.88(1.88) 138.68(7.93) 17.24(1.18) 69.38(1.62) 1338.75(53.45)

6.29(0.41) 355.82(34.54) 358.48(17.91) 103.15(2.35) 328.44(25.75) 201.54(5.45) 59.35(4.52) 19.85(0.08) 24.09(0.23) 1560.65(133.50)

14 15 16 17

60 60 60 60

243.18(9.55) 189.57(3.20) 711.25(7.86) 309.05(3.83)

593.78(2.72) 525.29(0.49) 2406.82(31.61) 589.17(44.65)

463.21(42.34) 224.15(11.05) 1933.10(180.03) 476.59(36.53)

374.61(12.28) 125.14(1.50) 1946.22(15.17) 460.82(6.76)

207.18(17.07) 147.65(10.56) 744.44(64.09) 238.04(20.56)

18 19 20 21 22 23 24 25

93 137 69 69 69 177 69 69

32.41(0.66) 12.41(0.20) 222.63(7.42) 71.02(1.28) 646.30(7.22) 168.64(7.18) 106.03(3.37) 1795.70(129.93)

31.71(0.89) 11.37(0.34) 172.02(15.74) 27.83(1.8) 471.85(13.37) 184.14(0.36) 248.86(0.06) 2163.03(65.19)

21.62(0.35) 16.10(0.24) 206.48(3.34) 20.77(1.40) 583.59(32.22) 231.84(6.8) 363.16(6.98) 3663.09(367.63)

9.11(1.10) 15.28(0.31) 136.21(1.42) 13.95(1.26) 389.28(5.89) 192.76(0.85) 255.01(0.08) 1283.62(32.80)

23.21(0.48) 13.61(0.18) 156.45(16.28) 38.42(3.81) 490.04(39.45) 138.21(1.10) 70.05(4.44) 784.04(66.17)

26 27 28 29 30 31 32 33

74 74 88 88 105 71 104 85

15.94(0.79) 52.57(0.75) 284.96(10.65) 1678.08(203.36) 375.83(3.48) 9.68(0.16) 319.29(9.83) 22.23(0.12)

4.99(0.38) 11.54(1.53) 275.10(0.71) 535.18(32.85) 91.49(5.99) 34.58(0.86) 165.35(0.39) 33.17(2.22)

nd 4.23(0.27) 178.53(15.32) 300.81(16.54) 188.37(3.62) 291.84(25.87) 49.53(2.24) 28.46(1.73)

nd nd 129.68(9.03) 200.51(11.42) 102.54(3.26) 21.93(1.07) 283.14(8.68) 46.84(0.43)

8.78(0.74) 19.77(2.47) 169.44(21.64) 257.66(37.28) 69.21(3.90) 12.32(1.22) 182.42(6.35) 22.99(0.13)

Id = Identification in Figure 3, m/z = ion selected in SIM mode, nd = not detected.

Acknowledgment Financial support for this study was managed by the Natl. Inst. of Research and Agro-Food Technology (INIA) and co-financed with ERDF and ESF funds (project RTA2009–00113-00–00).

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C: Food Chemistry

Table 3–Semi-quantitative concentrations of volatile compounds in varietal apple pomaces. Mean and standard deviation expressed as microgram of 2-octanol/kg of wet pomace.


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