Postharvest Biology and Technology 45 (2007) 1119

Postharvest quality of integrated and organically produced apple fruit

E. R th a, , A. Berna a , K. Beullens a , S. Yarramraju b , J. Lammertyn a , A. Schenk a , B. Nicola a o
a

Flanders Centre of Postharvest Technology/BIOSYST-MeBioS, Katholieke Universiteit Leuven, Willem de Croylaan 42, Leuven B-3001, Belgium b Laboratory for Pharmaceutical Analysis, Katholieke Universiteit Leuven, O & N2, Herestraat 49, PB923, Leuven B-3000, Belgium Received 28 June 2006; accepted 17 January 2007

Abstract Apple cv. Jonagold fruit were picked in three different regions of Belgium. In each region one organic and one integrated orchard with identical climatic and soil characteristics was sampled. Fruit were stored in air and under CA conditions (1% O2 , 2.5% CO2 ) at 1 C for 6 months. The acoustic stiffness, rmness, soluble solids contents, acid and sugar contents, and the aroma prole were measured. Quality parameters were analysed right after harvest and storage. At both times an additional shelf-life experiment was carried out to simulate the conditions in the commercial chain. The quality attributes of apples coming from different regions and different production systems did not differ signicantly, neither at harvest nor after storage. There was a considerable softening during storage in air and shelf-life, but not under CA conditions. Immediately after harvest, high malic acid, quinic acid and sucrose contents were observed, while glucose and citric acid contents were higher after storage. The aroma prole changed during shelf-life, except for apples stored in air, which even immediately after storage already had an aroma prole comparable to that after shelf-life. The volatile responsible for the typical apple aroma (2-methylbutyl acetate) had the highest relative abundance after CA storage and subsequent shelf-life, followed by apples immediately after CA storage. The effect of storage conditions on the quality of the apples was in general much larger than that of the possible effects of the production system. Aroma proles of air-stored and CA-apple converge during shelf-life conditions. 2007 Elsevier B.V. All rights reserved.
Keywords: Apple; Aroma; Integrated; Organic; Quality; Taste

1. Introduction Due to the increasing importance of food safety, environmentally friendly production systems (organic and integrated crop management) are becoming more important in food research. The integrated crop management system is an ecologically safer method aiming at minimising the undesirable side effects and use of agrochemicals (replacing them with environmentally friendly alternatives) to enhance the environment and human health. Organic farming seeks to eliminate the use of agricultural chemicals and to apply natural materials both in plant protection and in nutrition supply (Ferree and Warrington, 2003). Organic products have much lower amounts of pesticide residues compared to integrated or conventional ones (Baker et al., 2002). Besides the presumed health properties, these systems claim to yield products which have a better internal quality (avour,

Corresponding author. Tel.: +32 16 322384; fax: +32 16 322955. E-mail addresses: erika.roth@biw.kuleuven.be, g.erikaroth@gmail.com (E. R th). o 0925-5214/$ see front matter 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.postharvbio.2007.01.006

texture, vitamins). Apples grown organically may have a lower growth rate (Lampkin, 1990) and may have smaller fruit due to smaller cells and less intercellular spaces. Fruit size very much depends on nutrition supply, which also remarkably inuences postharvest quality (Harker et al., 1997; Sams, 1999). Nitrogen, phosphorus, potassium, and calcium in particular have been reported to have pronounced effects on texture (Sams, 1999). Storage potential depends mainly on season and site (related to variable cropping and climatic factors), but these interact with orchard management systems and nutrition supply (Sharples, 1980). Factors prior to harvest, such as environment, cultural practices, agrochemicals and nutrition, have an impact on avour of fruit and vegetables through the effects of the abovementioned parameters on plant development (Mattheis and Fellman, 1999). Previous research has shown differences between the two production systems in their postharvest behaviour (R th et al., o 2001a, 2001b, 2004), but these differences were mainly due to different ripening stages at harvest. The critical impact of harvest maturity on fruit quality has been stressed also by Harker (2004) and Mattheis and Fellman (1999). The objective of our research was to examine whether there is a statistical difference in the

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texture, sugar and acid contents, aroma volatiles and postharvest behaviour of apples coming from integrated versus organic orchards. Special attention was paid to harvesting all apples at a similar maturity stage. 2. Materials and methods

sured rst. Ten apples were randomly selected for the analysis of taste compounds and aroma volatiles. Apples after harvest were kept in a cool place for half a day before the start of the cold storage. The required conditions in the CA-chambers (1 C, 1 kPa O2 , 2.5 kPa CO2 ) were reached after 2448 h after the start of cooling. 2.2. Methods

2.1. Material Six apple (Malus domestica Borkh. cv. Jonagold) orchards were chosen (three organic and three integrated) in three different production regions of Belgium to provide raw material for the experiment. Orchards were situated within 20 km of Leuven in the region of Bierbeek (Region I), Molenbeek-Wersbeek (Region II) and Assent (Region III). The organic and integrated orchards within a production region were located close to each other so that the effect of microclimate and soil type could be reduced. Organic orchards followed the rules (EG 2092/91) laid down by the Blik (Division of Integra for Organic Production). Integra bvba is the certication and inspection body for agriculture and food processing in Belgium. The integrated orchards were treated according to the standards for integrated production in Belgium (Belgisch Staatsblad MB 1996-03-01/31). The year of planting, rootstock and spacing is shown in Table 1. Apples were picked at the same maturity stage, which was determined by the change and value of the Streif index calculated in the last 2 weeks before harvest. According to the Streif index and the background colour, apples of similar size were picked at about their commercially optimal maturity for storage (Streifoptimal = 0.07 according to Streif, 1996) in September 2004. The Streif index is an indirect measure of the maturity and can be calculated from three quality attributes, as below: Streif = F (kg) SSC (%)S 2.2.1. Stiffness measurement Acoustic stiffness (Schotte et al., 1999) was measured using the Aweta acoustic rmness sensor (Aweta DFT, Version 0.0.0.7., Nootdorp, The Netherlands). The stiffness s was related to the characteristic frequency according to Chen and De Baerdemaeker (1993): s = f 2 m2/3 with s the acoustic rmness index (s2 kg2/3 ), f the rst resonance frequency of the vibration spectrum, and m the fruit mass (kg). Stiffness was measured at harvest and immediately after storage and after the corresponding shelf-life experiments. All apples were labelled and measured at harvest while only 20 apples per batch were measured each consecutive time. Five parallel measurements were done on each apple at their equator and the mean stiffness per apple was calculated from these data. In all measurements the apples were treated as individuals, thus all parameters measured are known for each apple. 2.2.2. Firmness measurement The Magness-Taylor rmness was measured using a TAXT2 Texture Analyser (Stable Micro Systems Ltd., Godalming, Surrey, UK) with a cylindrical probe (11 mm diameter). The maximum force needed to penetrate the esh over a distance of 8 mm at a speed of 8 mm/s was measured at two opposite sites on the equator of the fruit. The apple was peeled at the point of penetration before the measurement. The rmness of 20 apples was measured at harvest and immediately after storage and after the subsequent shelf-life experiments. 2.2.3. Soluble solids contents The juice that was released by the rmness test was used to measure soluble solids (SSC) with a digital refractometer (Atago Co. Ltd., Tokyo, Japan) and expressed as %. 2.2.4. Sugar and acid contents Immediately after measuring the MT-rmness and soluble solids contents, apples were sliced, frozen in liquid N2 and

with F the rmness (kg), SSC the soluble solids content (%) and S the starch index (CTIFL starch conversion chart). From every orchard 40 apples were used for measurements at harvest, 40 apples were stored for 6 months under controlled atmosphere conditions (1 C, 1 kPa O2 , 2.5 kPa CO2 ) and another set of 40 apples under normal air (1 C, 21 kPa O2 , 0.03 kPa CO2 ). Half of each set of 40 apples were analysed immediately after harvest and just after storage, while the other half was kept for 14 days under shelf-life conditions (20 C) to simulate the conditions in the commercial chain. In total, 720 apples were used in this experiment. At each measurement time, the basic parameters (stiffness, rmness, SSC, weight) of the 20 apples were meaTable 1 Year of planting, rootstock and spacing of trees in the studied orchards Orchard Production system Rootstock Planting year Spacing (m m) Region I Organic M9 1997 1.25 3.4 Region I Integrated M9 1996 1.25 3.25

Region II Organic M9 1998 1.25 3.4

Region II Integrated M9 2002 1 3.5

Region III Organic M9 1995 1.25 3.5

Region III Integrated M9 1996 1.25 3.45

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stored at 80 C for subsequent analysis of individual apple fruit. Frozen powder of apple tissue (0.1 g) was extracted three times using ethanol (80%, v/v, 500 l/extraction), then dried and stored frozen (20 C). Water-diluted samples were made from this frozen extract and analysed with two Agilent 1100 Series HPLC instruments (Agilent Technologies Inc., Santa Clara, CA, USA). The HPLC for analysis of sugars (sorbitol, sucrose, glucose and fructose) was equipped with an Aminex Resin Based column (HPX-87C, 300 mm 7.8 mm; Bio-Rad, USA) using water (water for HPLC, VWR) as mobile phase. The HPLC for analysis of organic acids (citric, malic and quinic acid) was equipped with an Alltech Prevail Organic Acid column (250 mm 4.6 mm; Alltech Associates Inc., Deereld, IL, USA) using potassium dihydrogen phosphate (pH was adjusted to 2.5 using metaphosphoric acid) as mobile phase. A refractive index detector and diode array detector was used for the analysis of sugars and acids, respectively (Beullens et al., 2006; Rudnitskaya et al., 2006). 2.2.5. Aroma volatiles The aroma prole of apples was investigated using solid phase microextraction gas chromatography coupled to mass spectrometry (SPME-GC-MS). The GC (6890N, Agilent Technologies Inc., USA) used in this study was coupled to a single quadrupole mass spectrometer (Mass selective detector, Agilent). Vials were manipulated by a multipurpose sampler (MPS2, Gerstel, M lheim an der Ruhr, Germany) equipped with a heated u tray of 25 housing positions. The autosampler syringe was equipped with an SPME bre coated with a 65 m thick layer of PDMS/DVB (Supelco Co., Bellefonte, USA). The bre was preconditioned at 250 C for 1 h. The chromatographic column used was a diphenylpolysiloxane (5%) dimethylpolysiloxane (95%) coated capillary column (Optima-5-MS, 30 m 0.25 mm i.d. with 0.25 m lm thickness, Macherey-Nagel, D ren, Geru many). The parameters used in the SPME-GC-MS measurement were based on previous studies of the Flanders Centre of Postharvest Technology of the Katholieke Universiteit Leuven (Berna et al., 2004, 2005; Saevels et al., 2004). The ion source and quads temperatures of the MS were maintained at 250 and 150 C, respectively. The ionisation energy used was 70 eV. The total ion current was recorded after a solvent delay of 2.0 min at a rate of 4.15 scans per second with a scan range of 10350 m/z. The obtained spectra were identied using the reference mass spectra from the NIST library (National institute of standards and technology). The carrier gas used was helium at the ow of 1.2 mL/min (63.1 kPa). The purge ow used was 50 mL/min for 0.25 min. The injection port temperature was maintained at 250 C. The initial GC oven temperature was programmed at 40 C and increased to 165 C at a rate of 5 C/min and subsequently increased to 250 C at a rate of 50 C/min and maintained there for 15 min. The total GC run time was 42 min. Apple homogenate was used for headspace sampling as this reduced the time to reach static equilibrium of volatile analytes between the matrix and headspace compared to intact apples. Also, homogenate can be frozen quickly and stored until GCMS analysis. A further advantage is that the homogenate is closer to the masticated fruit pulp experience when eating. Sat-

urated NaCl solution (36%, w/v, in water) was added to the homogenate to avoid possible enzyme action after the apple is ruptured. NaCl was selected over CaCl2 due to the intensity of the volatiles salting out effect and also because it was observed that the slurry resulting when using CaCl2 was more viscous than that with NaCl, which might affect the time to reach static equilibrium. Therefore, 25 mL of saturated NaCl solution was added to 50 g of unpeeled apple (cut at random positions) and homogenized for 1 min using a high-speed blender (Waring Products Inc., USA). Subsequently 25 mL of water was added to the thick homogenate and blended for 1 min to decrease the viscosity and help to release the volatiles. This homogenate was immediately transferred to 15 mL Falcon tubes, frozen in liquid nitrogen and stored under 80 C until analysis. Sample preparation was performed as fast as possible once the apple was cut. This is not only due to the possible enzyme reaction but also the NaCl-induced volatile salting out effect may cause loss of volatiles of interest. On the day of the analysis, frozen samples were thawed in a water bath at 35 C for 15 min just before the analysis. Before routine analyses, the incubation temperature of the homogenate, the incubation time and the SPME extraction time were optimised using apples from the same batch. The optimal conditions were 35 C for 120 min followed by 10 min extraction time; extending the incubation time up to 14 h at 40 C did not result in appreciable changes in the volatile composition. A purge time of 0.25 min (purge ow of 50 mL/min) was chosen to get well resolved GC peaks. 2.2.6. Statistical analysis Differences between the parameters of objects were analysed by using Tukeys Studentised range test on the means by SAS System 8.0 (SAS Institute, Heidelberg, Germany). Principal component analysis (PCA), using the Unscrambler software (Version 9, CAMO AS Trondheim, Norway) was used to discriminate between production methods, growing regions and storage regimes. 3. Results and discussion 3.1. Overall quality The complete dataset was analysed by means of principal component analysis (PCA) (Fig. 1A and B) based on the correlation matrix. The aim was to see which quality attributes were mainly responsible for the differences between the objects. The rst two principal components (PC1 and PC2) explain 54% of the variability in the data set. No separation due to region or production system was observed. When the score plot of the PCA (Fig. 1A) was further investigated, a separation along the rst PC was observed between apples at harvest and after storage except for apples immediately after CA storage as their quality was very close to freshly harvested ones. Apples stored under different atmospheres form separate groups. Apples immediately after storage or at harvest also differed from their shelf-life counterparts. The loading plot of the analysis (Fig. 1B) shows that aroma volatiles, rmness and acid contents are aligned with

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liminary statistical analysis did not show a signicant regional effect on these parameters, the orchards of three regions were pooled for this analysis. No signicant difference due to the production system was observed. Firmness decreased signicantly during storage in air, but not during CA storage. As the acoustic stiffness is more inuenced by water loss (Hertog et al., 2003), a signicant decrease was visible also after CA storage, but it was less pronounced than in air. These results are in line with those of Aaby et al. (2002). During shelf-life the stiffness decreased signicantly in time, while the loss in rmness was not signicant. The soluble solids contents changed only slightly, both during cold storage and during shelf-life conditions. Weight loss was the highest after the shelf-life exposure of apple stored in air. The total acidity decreased signicantly, while the total aroma volatiles increased signicantly during storage in air. They did not change signicantly under CA conditions. The total aroma volatiles production signicantly increased also during shelf-life except for apples stored in air, which already immediately after removal from storage had a high aroma production. The lowest aroma production was observed at harvest, while the most volatiles were produced after 6 months storage in air. As a result of suppressed aroma synthesis during storage, the aroma intensity of apple fruit after CA storage was much lower than that after storage in air. 3.2. Sugar and acid contents The chemical composition of apple juices can vary, depending on cultivar, production region, maturity and various other horticultural practices (Leonard and Mayer, 1983). Due to possible micro-climate and soil differences between the different growing regions, a difference in taste parameters was expected. However, no regional effect on the taste parameters (sugar- and acid-contents) was found in our data. The most important taste components in apple are sugars such as glucose, fructose and sucrose and organic acids such as malic and citric acid (Ackermann et al., 1992; F leki et al., 1994; u Del Campo et al., 2005). Malic acid is the characteristic acid of apple, and hence is mainly responsible for the sour sensation of apple. The concentration of quinic acid has been also analysed as it is known to impact astringency to apple fruit and plays an important role in the sensation of the sweet taste. The average concentration of acids and sugars in the apple fruit coming from different orchards is indicated in Table 3. As the previous analysis did not show any effects of region on these parameters, orchards of different regions were pooled. No discrimination between production systems could be observed; neither in the case of fresh apples, nor in apples after shelf-life except for the citric acid contents of air-stored apples (integrated apples had higher citric acid contents). This result is in contrast to results of Reganold et al. (2001) who found organic apples to be less tart and sweeter than their integrated pairs both at harvest and also after storage. As no information is given in this work about the harvest maturity of apples coming from different production systems, the difference experienced by Reganold et al. (2001) could be the result of different maturity at harvest.

Fig. 1. Score plot (A) and loading plot (B) coming from principal component analysis of overall apple quality (taste, aroma and texture). On the score plot (A) different symbols indicate the scores for the rst two principal components (PC1, PC2) of fresh and stored apple fruit before and after shelf-life. The loading plot (B) represents the linear combination used for PC1 and PC2 with the different symbols referring to the loadings of texture, taste and aroma parameters. The rst two PCs explain respectively 39 and 15% of the observed variation.

the rst principal component axis. Freshly harvested fruit before and after shelf-life, as well as CA-stored fruit before shelf-life were characterised by low aroma volatile contents, but were rm and high in acids. Air-stored fruit before and after shelflife as well as CA-stored fruit after shelf-life, were soft, low in acids, but high in aroma volatiles. The second principal axis is mainly characterised by sugar and aroma composition, and separates stored apples before and after shelf-life. All aroma volatiles were higher in stored apples except 2-hexenal, which is a characteristic compound of freshly harvested apples (Mattheis et al., 1991; Delippi et al., 2005) and apples immediately after removal from CA, imparting a fresh, grassy aroma for the apple. It can be considered as a freshness indicator. Tukeys Studentised range test on the means (Table 2) was used to analyse the differences with respect to overall quality parameters (rmness, stiffness, soluble solids contents, weight loss, total acidity, total aroma volatiles) between objects. As pre-

E. R th et al. / Postharvest Biology and Technology 45 (2007) 1119 o Table 2 Mean values of main quality attributes at harvest and during storage of organic and integrated apple fruit Sampling time At harvest, fresh At harvest, shelf-life Air storage, fresh Air storage, shelf-life CA storage, fresh CA storage, shelf-life Production system Organic Integrated Organic Integrated Organic Integrated Organic Integrated Organic Integrated Organic Integrated Firmness (N/cm2 ) 73.2 a 73.2 a 70.9 ab 72.9 a 48.6 c 50.3 c 45.9 c 45.4 c 75.2 a 74.5 a 66.4 b 72.1 a Stiffness (104 s2 kg2/3 ) 30.6 a 29.8 a 19.8 d 21.5 d 11.6 f 12.1 f 5.4 g 6.1 g 27.7 bc 26.4 c 14.7 e 13.6 ef SSC (%) 13.10 ab 12.77 b 13.61 a 13.59 a 13.60 a 13.52 ab 13.38 ab 13.06 ab 13.70 a 13.46 ab 13.20 ab 13.68 a Weight loss (%) 2.93 c 2.11 cd 5.22 b 4.61 b 8.26 a 8.05 a 1.40 d 1.23 d 4.63 b 4.49 b Total aciditya (mmol/mL) 0.056 ab 0.057 a 0.054 ab 0.058 a 0.042 de 0.040 def 0.031 f 0.033 ef 0.049 abcd 0.051 abc 0.042 cde 0.047 bcd

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Total aroma (sum of peak area of 20 volatiles) 1.10 108 c 7.19 107 c 8.73 108 a 5.40 108 b 7.96 108 a 8.44 108 a 8.35 108 a 8.03 108 a 1.16 108 c 1.01 108 c 5.79 108 b 5.51 108 b

Fruit coming from different orchards were pooled as no regional effect was observed in the preliminary analysis. Tukeys Studentised range test was used to perform a pair wise test of the means per quality attribute. Means (within the same column) with the same letter are not signicantly different at the 95% condence interval. SSC: soluble solids content. a Total acidity was calculated as the sum of acids present (in mmol) in 1 mL apple juice. Each acid was considered with 1 of dissociation (due to the low pH of the apple).

Fructose was the dominating sugar representing 5570% of the total sugars, followed by sucrose, glucose and nally sorbitol. This is in agreement with results of Suni et al. (2000). The main acid was malic acid contributing about 9598% of the total acid contents. Both quinic acid and citric acid only amounted to a trace to up to 5% of the total acid contents. The amount of fructose did not change considerably in time, while the amount of sucrose decreased during storage in both air and under CA, in line with results of Gorin (1973) and Suni et al. (2000). At the same time, glucose and sorbitol contents showed an increase as a function of storage time. These results agree with results of Suni et al. (2000), who reported that storage had only minor a effect
Table 3 Mean concentrations of taste compounds of organic and integrated apple fruit Sampling time At harvest, fresh At harvest, shelf-life Air storage, fresh Air storage, shelf-life CA storage, fresh CA storage, shelf-life Threshold values (mg/mL) Production system Organic Integrated Organic Integrated Organic Integrated Organic Integrated Organic Integrated Organic Integrated Quinic acid (mg/mL) 0.214 a 0.180 ab 0.186 ab 0.183 ab 0.167 bc 0.147 bcd 0.107 d 0.134 cd 0.102 d 0.105 d 0.130 cd 0.121 d 0.375a Malic acid (mg/mL) 7.351 ab 7.515 a 7.120 ab 7.589 a 5.412 cde 5.197 def 4.109 f 4.315 ef 6.449 abc 6.767 ab 5.526 cde 6.185 bcd 0.2145b

on the average content of sugars and acids of apple compared to the effect of cultivar. In contrast to our results, Lee and Wrolstad (1988) showed a decreasing sorbitol content during storage of apples. Sorbitol is the principal photosynthate translocated from leaves to fruit and is metabolised to other sugars after the uptake by the fruit (Loescher, 1987). It is characteristic of apple that fructose is produced in abundance while relatively little glucose is formed due to the conversion of sorbitol to fructose by sorbitol dehydrogenase (Ber ter and Studer Feust, 1997). u This is conrmed by our results. The increase in the level of sorbitol during storage may be attributed to the anaerobic conversion of fructose into sorbitol (Ackermann et al., 1992).

Citric acid (mg/mL) 0.039 ef 0.046 ef 0.033 f 0.046 ef 0.126 b 0.150 a 0.073 cd 0.084 c 0.065 d 0.070 cd 0.046 ef 0.057 de 0.4419b

Sorbitol (mg/mL) 0.636 d 0.626 d 0.748 d 0.794 d 1.410 abc 1.406 abc 1.649 a 1.591 ab 1.238 c 1.187 c 1.205 c 1.351 bc 13.68b

Glucose (mg/mL) 12.552 d 13.132 d 14.681 cd 17.674 abc 17.134 bc 19.818 ab 10.860 d 14.365 cd 19.877 ab 21.249 a 19.767 ab 21.576 a 11.03b

Sucrose (mg/mL) 28.184 a 28.868 a 29.657 a 27.392 a 20.569 b 16.952 bcd 19.275 bc 17.436 bcd 15.220 bcd 14.247 cd 13.279 d 12.791 d 6.84b

Fructose (mg/mL) 61.236 ab 62.341 ab 68.868 a 67.822 ab 59.758 ab 61.020 ab 61.552 ab 62.658 ab 59.914 ab 57.471 b 65.571 ab 63.177 ab 5.70b

Fruit coming from different orchards were pooled as no regional effect was observed in preliminary analysis. Tukeys Studentised range test was used to perform a pair wise test of the means per quality attribute. Means (within the same column) with the same letter are not signicantly different at the 95% condence interval. a Estimated as the mean of the threshold for malic and citric acid. b Amerine et al. (1965).

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ones (Fig. 2A) than in the previous analysis. The rst two PCs explain 98% of the variability in the data; PC1 and PC2 represent acidity and sweetness, respectively. The loading plot (Fig. 2B) shows that malic acid is mostly responsible for the variability in the data (determining the rst PC), followed by fructose content (which determines the second PC). Sucrose, a storage carbohydrate, having a positive loading like malic acid, is present in higher amounts in fruit at harvest than in stored fruit. This shows that apples at harvest are more tart and have a different carbohydrate pattern than stored fruit. Acids and sugars present in smaller amounts do not pay an important role in separating the different treatments based on their taste compounds. 3.3. Aroma volatiles The most abundant aroma volatiles of Jonagold apples consisted of three alcohols (mainly 1-hexanol) responsible for only 48% of total volatile production, two aldehydes (2-hexenal, hexanal; making up 4065% of the total volatile production), 13 esters (mainly acetates like hexyl acetate, 2-methylbutyl acetate, butyl acetate; making up 3050% of total produced volatiles), one sesquiterpene (alpha farnesene) and one aromatic hydrocarbon (1-methoxy-4-(2-propenyl) benzene). The list of compounds is given in Table 4. To avoid the errors due to measurement conditions, relative peak areas were used for the statistical analysis. The relative peak area was calculated by dividing each peak area by the total peak area of the 20 listed peaks (Table 4.) In order to evaluate differences in aroma as perceived by humans, a PCA was carried out on the relative peak areas
Fig. 2. Score plot (A) and loading plot (B) of PCA coming from principal component analysis of weighted taste parameters, using perception thresholds as weights, as indicated in the loading plot. On the score plot (A) different symbols indicate the scores for the rst two principal components (PC1, PC2) of fresh and stored apple fruit before and after shelf-life. The loading plot (B) represents the linear combinations of the parameters indicated for calculating PC1 and PC2. The rst two PCs explain respectively 91 and 7% of the observed variation. Table 4 The 20 most characteristic aroma compounds of Jonagold apple with their retention times (RT) RT (min) 2.18 2.66 2.96 3.47 3.88 4.10 4.18 4.98 5.43 5.67 6.44 6.59 9.10 9.68 10.66 15.85 16.00 17.41 22.24 26.13
a b c d

Compound 1-Butanol n-Propyl acetate 2-Methyl-1-butanol 2-Methylpropyl acetate Hexanal Propyl propanoate Butyl acetate 2-Hexenal 1-Hexanol 2-Methylbutyl acetate Butyl propanoate Pentyl acetate Butyl butanoate Hexyl acetate 2-Methylbutyl butanoate Hexyl butanoate + butyl hexanoate 1-Methoxy-4-(2-propenyl) benzene 2-Methylhexyl butanoate Hexyl hexanoate Alpha farnesene

Perception thresholds (ng/g) 500a 2000b 300a 5a 57c 66a 17a 500a 11b 25a 43b 100a 2a 17a 250a + 700d 6a

Malic and quinic acid contents decreased during storage under both conditions. As malic acid is the major substrate for respiration it decreased less under CA conditions than under air, due to suppressed respiration because of the low oxygen atmosphere. The data set was also analysed by means of PCA (Fig. 2). The amount of sugars and malic acid determines the balance of sweetness and tartness in the fruit. The more malic acid and the less sugars present, the stronger the avour. However, as the taste perception depends not only on the measured concentration of the individual compounds resulting in sweet and sour taste, but also among others, on the human perception threshold values, the data were analysed by weighting the taste compounds with the reciprocal value of their perception thresholds (Table 3; Amerine et al., 1965; Hong et al., 2005). Because a taste threshold value for quinic acid was lacking, the average threshold for malic and citric acid was applied to quinic acid. Fig. 2 shows the score and loading plot of a PCA on the weighted taste compounds. Apple fruit at harvest were less clearly separated from stored

Aaby et al. (2002). Dixon and Hewett (2000). van Gemert and Nettenbreier (1977). Takeoka et al. (1990).

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Fig. 3. Score plot (A) and loading plot (B) of the PCA based on the relative areas of the 16 abundant peaks (weighted by their perception thresholds) in case of organic and integrated apples, coming from different regions at harvest. On the score plot (A) different symbols indicate the scores for the rst two principal components (PC1, PC2) of organic and integrated apple fruit. The loading plot (B) represents the linear combinations of the parameters indicated for calculating PC1 and PC2. The rst two PCs explain respectively 88 and 8% of the observed variation.

Fig. 4. Score plot (A) and loading plot (B) of PCA based on the weighted relative peak area of the most important 16 volatiles of Jonagold apple, using perception thresholds as weights. On the score plot (A) different symbols indicate the scores for the rst two principal components (PC1, PC2) of fresh and stored apple fruit before and after shelf-life. The loading plot (B) represents the linear combinations of the parameters indicated for calculating PC1 and PC2. The rst two PCs explain respectively 91 and 6% of the observed variation.

weighted by their perception thresholds (Table 4; van Gemert and Nettenbreier, 1977; Takeoka et al., 1990; Dixon and Hewett, 2000; Aaby et al., 2002). As threshold values were only available for 16 out of our 20 compounds, only these 16 compounds were included in the analysis. When analysing the score plot obtained by the PCA based on the relative areas of 16 abundant peaks (weighted by their perception thresholds) at harvest, the different orchards could be distinguished, but no clear effect of production system or region was observed (Fig. 3). Hence, the statistical analysis focused on interpreting the effects of storage regime and shelf-life on both production systems and regions and would show up any possible discrimination. In Fig. 4, the score plot of the full PCA analysis is shown. A clear separation of the storage times and regimes was obtained, where the rst two PCs explained 97% of the variability in the data set. From the score plot (Fig. 4A) it is clear that the perceived aroma prole of apples at harvest was similar to that of apples stored in CA immediately after removal from storage,

although a shift along PC3 can be observed. When analysing the loading plot of PC3PC1 (plot not shown) it appears that the relative contribution of 2-hexenal was higher for freshly harvested apples than for apple fruit after CA storage. Aldehydes are the major sources of ester synthesis and they are mostly abundant in unripe fruit (Mattheis et al., 1991; Delippi et al., 2005; Komthong et al., 2006). As CA storage suppresses the synthesis of aroma volatiles (Dixon and Hewett, 2000; Tough and Hewett, 2001; Fellman et al., 2003; Saquet et al., 2003; Saevels et al., 2004), it is plausible that the aroma prole of CA-stored apples is closer to freshly harvested apples than to that of apples stored in air. Fruit after storage in CA had a more typical apple-like aroma as they were richer in 2-methylbutyl acetate and also could be perceived as having a fresher, greenish avour than air-stored apples due to their higher aldehyde contents and the more dominant presence of 2-methylhexyl butanoate (responsible for an apple-, grapefruit-like aroma). After shelf-life, the potentially perceived aroma of apples from different storage regimes was more similar than immediately after removal from storage due to the on-going aroma synthesis. Similar observations were made

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E. R th et al. / Postharvest Biology and Technology 45 (2007) 1119 o

by Saevels et al. (2004). After shelf-life exposure, CA-stored fruit lost their fresh avour sensation as a result of reduced aldehyde content, but retained their apple-like aroma. Air-stored apples were characterised by an apple-pear-oral-like aroma due to their predominant hexyl acetate. The effect of shelf-life on the relative aroma prole of fresh and stored apples is different. This might be due to the different metabolic processes during shelflife of freshly harvested apple fruit as compared to the shelf-life of either CA- or air-stored apple fruit. In the case of freshly harvested fruit the change mainly happens along PC1, while for the CA-stored fruit a shift along PC2 is observed. From the loading plot (Fig. 4B) it is clear that the majority of the variance is due to ve aroma volatiles. Hexyl acetate is more abundant in the perceived aroma after storage in air, and following subsequent shelf-life, than in CA. These results are supported by the data of Saevels et al. (2004). Butyl and hexyl acetate and methyl branched esters characterise apples after shelf-life, and are partly responsible for the sweet, fruity avour sensation of Jonagold apples. They become more abundant as fruit ripen. As the production of volatile compounds changes dramatically as ripening progresses, the observed tendencies can be signs of different ripening stages of apples after storage and the subsequent shelf-life experiment. Previous analysis (not shown) on relative peak areas without perception thresholds gave similar results as compounds important in separation of the objects have low threshold values (even a low concentration of them can be perceived easily), with the exception of butyl acetate which was replaced by 2-methylhexyl butanoate (that has a lower perception threshold). When analysing the relative abundance of aroma volatiles, the volatile that increased the most during storage in air was hexyl acetate. This compound may, hence, be considered as an indicator for maturity of Jonagold apple. The predominant esters are the acetates (hexyl acetate, butyl acetate, 2-methylbutyl acetate) as Jonagold belongs to the ester type of apples based on its volatile production (Mattheis et al., 1995; Dixon and Hewett, 2000; Komthong et al., 2006). Propyl-acetate and propyl-propanoate were compounds absent at harvest and also immediately after removal from CA storage. In line with the results of Fellman et al. (2003), after shelf-life or air storage these compounds were synthesized and can thus be considered as ripeness indicators. Bourn and Prescott (2002), Magkos et al. (2003) and Trewavas (2004), based on a literature survey, stated that there was no strong evidence that organic and conventional food differ in their concentration of nutrients, which is in agreement with our results. They are also in agreement with those of Hansen (1981), Baker (1992) cited by Trewavas (2004) stating that the public cannot easily distinguish organic products from conventional ones. Acknowledgements Authors are grateful to the Research Council of K.U. Leuven and the IWT (VIS 10427) for their nancial support. Thanks also to Christine Frank, Ele Dekempeneer and Bert Vandebosch for their valuable assistance.

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