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Changes in some odour-active compounds in paclobutrazol-treated ‘Starkspur Golden’ apples at harvest and after cold storage
Postharvest Bidogy and Technology ELSEVIER
Postharvest Biology and Technology 11 (1997) 39946
Changes in some odour-active compounds in paclobutrazol-treated ‘Starkspur Golden’ apples at harvest and after cold storage Anna
Rizzolo a,*, Costanza
Visai b, Maristella
Vanoli b
r’Istiruto Sperimentale per la Valorizzazione Tecnologica dei Prodotti Agricoli (I. V. T.P.A.), cia Venetian, 26-20133 Milano. Itall h Istituto di Coltivarioni Arhoree, Universith degli Studi, via Celoria, 2-20123 Milano. Ital?
Accepted 21 January 1997
Abstract Apple trees (M&s domestica Borkh. cv ‘Starkspur Golden’) were treated 25 days after full bloom (DAFB) with paclobutrazol (PBZ) as a soil-drench application. After 2 years, fruit were picked 158 (commercial harvest), 172 and 181 DAFB from treated and untreated trees, with fruit taken at commercial harvest being stored in air at 2°C and 90% R.H. During ripening and after 5 and 7 months of storage, treated and untreated apples were analysed for odour-active volatiles, by using either static or dynamic headspace samplings and capillary gas chromatography. The results were expressed in terms of Odour Units (Uo) in order to relate the relative importance of each compound to overall aroma. The average Uo values at harvest and after storage showed that only half of the monitored volatiles were present in concentrations great enough to contribute to overall aroma. PBZ treatment enhanced the ripening processes, which are associated with volatile evolution at harvest and after 5 months of air storage, and may have a specific effect on the metabolism of pentyl acetate and ethyl butanoate. 0 1997 Elsevier Science B.V. Keywords:
Apples;
Paclobutrazol;
Volatiles;
Ripening;
Air storage
1. Introduction Apple flavour is a complex combination of taste and odour sensations. Although taste and texture are important for apple quality, the presence of * Corresponding 2365377. 0925-5214/97!$17.00
author.
Tel.: + 39 2 70630256;
6 1997 Elsevier
PII SO925-5214(97)01417-S
fax: + 39 2
Science B.V. All rights
reserved
trace amounts of volatiles, responsible for the characteristic odour, gives fruit much of their perceived quality (Dimick and Hoskin, 1983). Various studies have shown that the production of volatile compounds in climacteric fruits, such as apples, is related to the climacteric rise in respiration (Salunkhe and Do, 1976). However, little information exists about their evolution, and
40
A. Rizzolo et al. /Postharvest
Biology and Technology I1 (1997) 39-46
especially on whether the few volatiles which have a significant impact on apple sensory quality behave in the same way as any other odourless compounds. Numerous factors such as variety, climate, date of harvest and storage (Dimick and Hoskin, 1983; Rizzolo et al., 1995) affect the levels of apple volatile compounds. Usually, apples destined for storage are harvested prior to the onset of the climacteric in order to delay the formation of ethylene and ripening. Paclobutrazol [/3-[(4chlorophenyl) methyl - CI- ( 1, 1- dimethylethyl) - 1H1,2,4-triazol-1 -ethanol] (PBZ) has been widely shown to be active in controlling the vegetative growth of many fruit crops; generally it reduces ethylene production, delays fruit maturation and ripening, enhances fruit set and limits water loss (Steffens and Wang, 1986). In fruit after cold storage in air, PBZ also causes a lower ethylene production, and a delayed CO, climacteric (Elfving et al., 1987; Luo et al., 1987). However, the type of effect strictly depends on the cultivar and the time of PBZ application. As an example, Wang and Steffens (1987) found no differences in respiration and ethylene evolution between untreated and PBZ-treated ‘Spartan’ apples. Few researchers have considered the influence of growth regulators on aroma compounds during apple fruit ripening and storage. Bangerth (1986) observed in aminoethoxyvinylglycine-treated ‘Golden Delicious’ apples no climacteric-like increase in CO, production and a greatly reduced emission of volatiles during cold storage. Yahia et al. (1990) found that daminozide adversely affected volatile composition in ‘Cortland’ and ‘McIntosh’ apples, delaying the onset of climacteric ethylene production and the formation of five odour-active volatiles ((E)-2-hexenal, methyl, butyl, hexyl hexanoate and hexyl 2-methylbutanoate) during fruit maturation and after storage in low-ethylene controlled atmosphere. Etcher (personal communication) found in ‘Starkspur Golden’ apples that soil-drench PBZ treatment was effective in controlling vegetative growth up to 3 years after the treatment, reducing internode length and leaf size, and producing fruit with reduced size and length/diameter ratio com-
pared with the control apples. For the fruit shape and size the effect of the treatment was very noticeable in the first year, but this diminished gradually over the next 2 years. Rizzolo et al. (1993), studying the quality of these ‘Starkspur Golden’ apples during fruit growth (from 50 to 180 days after full bloom), showed that even after 2 years, soil-drench PBZ treatment had an influence on the starch content and on total volatile evolution, specially during maturation and ripening, whilst it did not affect titratable acidity and soluble solids. It is not known whether PBZ has adverse effects also on the qualitative composition of apples during ripening and after cold storage. The purpose of this research was to study how soil-drench PBZ application on ‘Starkspur Golden’ apple trees affected the composition of volatiles during maturation and after air storage, paying particular attention to some odour-active compounds.
2. Materials and methods PBZ treatment and storage Apple trees, 17 year old (Malus domestica Borkh. cv ‘Starkspur Golden’) grafted on MM1 11 and grown in the Experimental Orchard of the University of Milan (Montanaso Lombardo, PO Valley), were used. One group of five trees was treated 25 days after full bloom (DAFB, full bloom occurring on April 16) with PBZ as a soil-drench around the trees (3 g/tree). A second group of five untreated trees was used as control. After 2 years, five apples/tree were picked 158 (commercial harvest), 172 and 181 DAFB (FB occurred on April 16) from treated and untreated (control) trees, when soluble solids were 1 l- 11.5” Brix and titratable acidity was 6.3 meq/lOO ml of juice for early picked apples, 3.5 meq/lOO ml of juice for 172 DAFB fruit and 2 meq/lOO ml of juice for the apples from the last harvest. Fruits were pooled into two replications and analysed for volatile composition by static headspace/capillary gas chromatography on a Carbowax 20 M column (25 m x 0.5 mm i.d., 1 pm film thickness) according to the conditions reported by Rizzolo et
A. Rizzolo
et al. /Postharvest
Biology and Technology
al. (1992). Each sample was obtained by pooling a slice per fruit and mincing the combined tissue. At commercial harvest (158 DAFB), 80 fruits from control .and treated trees were collected and stored in air at 2°C and 90% R.H. After 5 and 7 months of storage, 20 apples per treatment, divided into two replications, were analysed for volatile compound evolution during post-storage ripening at 18°C by dynamic headspaceicapillary gas chromatography on a 12 m x 0.33 mm i.d. Carbowax 20 M column (0.5 pm film thickness) following the conditions described in previous papers (Rizzolo et al., 1992; Vanoli et al., 1995). Volutile ident$cation and quarhjication Identification of volatile compounds was performed according to their retention times, comparing them with those of commercial standards and/or with the data obtained by gas chromatography/mass spectrometric analysis using a Hewlett&Packard 5870B Quadrupole Mass Spectrometer operated in the electron ionization mode at 70 eV and identified via National Bureau of Standards, Washington, D.C. library match. Volatiles were quantified using flame ionization detection by relating the peak area of each compound to that of an external standard (hexyl acetate) and were expressed as pggikg of fruit flesh for static headspace, and as pg/kg per h for dynamic headspace. To relate the relative importance of each compound to overall aroma, concentrations were converted into Uo. This representation of volatile pattern is an approximation, but has a practical use in selecting the most important contributors to aroma (Guadagni et al., 1966). Uo were calculated from concentrations using odour thresholds reported in the literature (Herrmann, 1991), according to: Uo = C/OT, where C = concentration in the sample and OT = odour detection threshold concentration. Compounds with Uo equal to or greater than 1.0 significantly contribute to aroma as they are above their odour threshold concentration.
I1 (1997) 39.-46
41
3. Results Volatile composition The odour-active compounds monitored in ‘Starkspur Golden’ apples and their odour threshold concentrations are listed in Table 1. Some compounds, such as butanol, hexanol, butyl propanoate, hexyl butanoate, butyl hexanoate and acetaldehyde show high odour threshold concentration (above 100 ppb) so they could contribute to aroma only if they are present in very high amounts. Other compounds, such as ethyl butanoate and ethyl 2-methylbutanoate could contribute to aroma even if they are present in very low quantities, as their odour threshold concentrations are below 1 ppb. In considering the average odour units (Av Uo) at harvest and after storage (Table 2), only half of the compounds monitored actually contributed to the overall aroma. The main odour contributors were ethyl butanoate, hexyl acetate, ethyl 2methylbutanoate, butyl acetate, hexanal and Table 1 Odour-active volatiles monitored in ‘Starkspur and their Odour Thresholds (OT) (Herrmann, Volatiles
OT(ppb)
Esters Butyl acetate Pentyl acetate Hexyl acetate (Z)-3-hexenyl acetate Propyl propanoate Butyl propanoate Ethyl butanoate Propyl butanoate Hexyl butanoate 3-methylbutyl butanoate Ethyl 2-methylbutanoate Hexyl 2-methylbutanoate Butyl hexanoate
18 250 21 0.1 22 250
Aldehydes Acetaldehyde Hexanal (E)-2-hexenal
120 4.5 17
Alcohol.~ Butanol 3-penten-2-01 Hexanol
500 4.2 150 _
66 5 2 2 57 200
I
Golden’ 1991)
apples
42
A. Rizzolo
et al. /Postharvest
Biology and Technology
Table 2 Average odour unit values (Av Uo) of volatile compounds in ‘Starkspur Golden’ apples at harvest, and after 5 and 7 months of air storage Compounds
Av Uo Harvest
Ethyl butanoate Hexyl acetate Ethyl 2-methylbutanoate Butyl acetate 3-penten-2-01 Hexanal Pentyl acetate Propyl butanoate (E)-2-hexenal Hexyl 2-methylbutanoate Acetaldehyde (Z)-3-hexenyl acetate Propyl propanoate Butanol Hexanol Hexyl butanoate+ butyl hexanoate Butyl propanaote 3-methylbutyl acetate
Storage 5 months
7 months
IS 31 6 3 1.1 1 1 0.8 0.3 0.27 0.2 0.15 0.1 0.09 0.03 0.03
10 48 9 2 0.2 2 3 0 1.5 0.1 0.05 0.06 0 0.06 0.14 0.05
9 24 38 1 0 2 2 0 0.8 0.002 0.05 0.003 0 0.04 0.08 0.02
0.01 0
0.04 0.05
11 (1997) 39-46
increasing amounts of hexanal, even if they were always below its odour threshold (Uo < 1); with ripening, control fruits produced decreasing quantities of hexanal, with Uo > 1 at 158 and 181 DAFB. A characteristic compound of ‘Golden’ apple aroma, 3-penten-2-01, was always detectable from organoleptic analysis in control fruits, while in PBZ-treated apples it contributed to the aroma only at 18 1 DAFB. Differences were also found among the esters. In PBZ-treated fruits, pentyl acetate was detected only at 172 DAFB at very low amounts and not significant for odour. Ethyl butanoate levels increased although the amounts at 158 and 172 DAFB were much lower than in control fruits. Control apples, on the other hand, produced increasing amounts of pentyl acetate, always 158 DAFB loguo ii,rl,
-;-~TYI~
0 0 _,,5 I-
___-. hex.4
pentyl acetate; other compounds (3-penten-2-01, (E)-2-hexenal and propyl butanoate) were present in amounts equal or close to the odour threshold concentration. All the other volatiles monitored had Uo values too low to contribute to the overall aroma and for this reason they will be not considered in the discussion. The major odour contributors changed from harvest to storage (Table 2). At harvest more than half of the total Uo were made up of ethyl butanoate, after 5 months of storage hexyl acetate, and after 7 months ethyl 2-methylbutanoate. In addition, with storage, propyl butanoate and 3-penten-2-01 no longer contributed to aroma, while (E)-2-hexenal increased and became detectable. Influence of PBZ treatment at harvest The PBZ treatment influenced the trend of some odour-active volatile compounds during ripening (Fig. 1). PBZ-treated apples produced
3p201
butac
penac
hexac
etbut
probut
e2mb
172 DAFB logIJO 24 T_-----
~--
1,5 +--
_::: -:‘oT~_E
TL!!
-I,5 hexal
_,,5 ~ _... hexal
3p201
..__
butac
penac
hexac
etbut
probut
e2mb
penac
hexac
etbut
probut
e2mb
.___~~~~~~~ 3p201
butac
Fig. 1. Odour-active compounds (log Uo) of control (W) and PBZ-treated (0) ‘Starkspur Golden’ apples at 158, 172 and 181 DAFB. No line drawn on x axis means that the compound is absent. Compound abbreviations: hexal = hexanal, 3p201= 3-penten-2-01, butac = butyl acetate, penac = pentyl acetate, hexac = hexyl acetate, etbut = ethyl butanoate, probut = propyl butanoate, e2mb = ethyl 2-methylbutanoate.
A. Rirzolo et al. /Postharoest Biology and Technology I I (1997) 39-46
present in amounts above its odour threshold, and very high levels of ethyl butanoate at 158 and 172 DAFB (about 200-220 Uo). At 181 DAFB, this ester was produced in an amount similar to that in PBZ-treated fruits (about 8 Uo). Butyl acetate and hexyl acetate increased with harvest both in control and in PBZ-treated apples; the latter fruits, however, developed higher Uo of these esters at 158 and 181 DAFB. Propyl butanoate followed an increasing trend with ripening both in PBZ-treated and control apples, exceeding the odour threshold concentration only at the last harvest date (181 DAFB), with control apples producing the higher amount. Ethyl 2-methylbutanoate had an increasing trend in control and PBZ-treated fruits; this ester was always higher in PBZ-treated apples. The main volatile compound which contributed to odour was ethyl butanoate in control fruits at 158 and 172 DAFB and hexyl acetate in control fruit at 18 1 DAFB and in PBZtreated apples at 158, 172 and 181 DAFB. Influence (f PBZ treatment after air storage Propyl butanoate was not detected in control and PBZ-treated apples after 5 and 7 months of air storage. PBZ-treated apples stored for 5 months (Fig. 2) with post-storage ripening at 18°C developed a relatively constant amount of hexanal and ethyl butanoate, comparable with those of control fruit. (E)-2-hexenal and butyl acetate showed a decreasing trend in PBZ-treated apples, while control fruit produced increasing amounts of these compounds, reaching the higher Uo values on the 8th day of post-storage ripening, when the fruits were fully matured. Pentyl and hexyl acetate were almost constant during post-storage ripening in PBZ-treated apples, while in control fruits they steeply increased on the 8th day. At the end of the post-storage ripening, only control apples produced 3-penten-2-01, and only PBZ-treated fruit developed ethyl 2-methylbutanoate. Comparing the Uo values on the 8th day of post-storage ripening, i.e. when fruits were fully
log
“0
PBZ-treated
43
apples
2.50 --/
Control apples
Fig. 2. Odour-active compounds (log Uo) of control and PBZtreated ‘Starkspur Golden’ apples after five months of air storage and 1 (open area), 2 (horizontal rules). 3 (diagonal rules). 7 (vertical rules) and 8 (filled area) days of post-storage ripening. No line drawn on x axis means that the compound is absent. Compound abbreviations: hexal = hexanal. t2hexal = (E)-2-hexenal. 3~201 = 3-penten-2-ol. butac = butyl acetate. penac = pentyl acetate, hexac = hexyl acetate. etbut = ethyl butanoate. e2mb = ethyl 2-methylbutanoate.
ripe (Fig. 2), the main contributing compound to odour in control fruit was hexyl acetate, while ethyl 2-methylbutanoate was the main compound in PBZ-treated fruits. Fully ripe control fruit were characterized by higher amounts of (E)-2-hexenal, 3-penten-2-01, butyl acetate, pentyl acetate and hexyl acetate. After 7 months of storage (Fig. 3) control and PBZ-treated apples did not emit 3-penten-201. PBZ-treated apples produced butyl acetate, pentyl acetate, hexyl acetate, (E)-2-hexenal. ethyl 2-methylbutanoate and ethyl butanoate with a behaviour similar to control fruits. In addition, (E)-2-hexenal was above the odour threshold only in PBZ-treated apples. Comparing the Uo values at the end of the post-storage ripening (Fig. 3), in both control and PBZ-treated apples the main contributing compound was ethyl 2-methylbutanoate.
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Biology and Technology 11 (1997) 39-46
4. Discussion
The odour-active compounds monitored in ‘Starkspur Golden’ apples have all been shown to be odour-active in ready-to-eat ‘Golden Delicious’ apple aroma (Rizzolo et al., 1989) and in a combined apple extract of various cultivars (Cunningham et al., 1986). However, by converting concentrations into Uo, not all these compounds can be shown to contribute to the formation of the aroma. Except for 3-penten-2-01, all the alcohols monitored were present in concentrations far below their odour thresholds. The same was found for the propanoates and butanoates of higher molecular weight. Thus, all these compounds could not significantly contribute to the aroma. The contributing volatiles collected from ‘Starkspur Golden’ apples treated with PBZ differed from those in control fruits considerably at harvest and after 5 months of storage in air. This is consistent with the findings of other authors (Elfving et al., 1987; Steffens et al., 1991), showPBZ-treated
apples
Control apples
0.5 0
Fig. 3. Odour-active compounds (log Uo) of control and PBZtreated ‘Starkspur Golden’ apples after 7 months of air storage and 1 (M), 2 (0) and 3 (W) days of post-storage ripening. No line drawn on x axis means that the compound is absent. Compound abbreviations: hexal = hexanal, t2hexal= (E)-2hexenal, butac = butyl acetate, penac = pentyl acetate, hexac = hexyl acetate, etbut = ethyl butanoate, e2mb = ethyl 2-methylbutanoate.
ing the influence of soil drench treatment with PBZ on ethylene production, fruit maturation and ripening, and gibberellin composition over a 3 year experimental period. In agreement with our previous findings on ‘Starkspur Golden’ apples (Vanoli et al., 1995) 172 DAFB control fruits had a volatile composition typical of ripe apples, i.e. a higher content of acetate esters, responsible for the ether-like, sweet, fruity characteristic aroma of ‘Golden’ apples, together with a high amount of ethyl butanoate, responsible for a powerful ether-like/fruity odour. 181 DAFB control apples showed a composition typical of overripe fruits, having the highest amounts of ethyl 2-methylbutanoate and detectable amounts of propyl butanoate, the former having a powerful diffusive green-fruity odour (Panasiuk et al., 1980; Willaert et al., 1983). Control apples picked at commercial harvest produced the highest quantities of hexanal and the lowest amounts of acetate esters; this pattern indicates that the commercial harvest time corresponds to unripe aroma quality (Panasiuk et al., 1980; Paillard, 1981; Willaert et al., 1983). PBZ treatment severely influenced the volatile composition at each harvest date, changing the odour profile. Treated apples never developed pentyl acetate, which gives a sweet-fruity odour. Hexanal was present in low amounts below its odour threshold; this aldehyde is responsible for a fatty-green, grassy odour. 3-penten-2-01, typical of ripe ‘Golden’ apples, exceeded the odour threshold only at the last harvest date. Contrary to control fruits, PBZ-treated apples always had as the main contributing odour compounds hexyl acetate (sweet, fruity) and ethyl 2-methylbutanoate (green-fruity). These patterns are close to those in control 181 DAFB fruits; thus, PBZtreated apples have an aromatic pattern more related to ripeness/overripeness at each harvest date. The fact that PBZ-treated apples produced less hexanal, no pentyl acetate, less ethyl butanoate, and more butyl acetate, hexyl acetate and ethyl 2-methylbutanoate could indicate that PBZ has a selective effect on the metabolism of these compounds.
A. Rir-_olo et 01. / Posthunwt
Biology
Differences between control and PBZ-treated apples were also found after storage in air. After 5 months at the end of post-storage ripening, control apples showed an aromatic pattern characteristic of ripe ‘Starkspur Golden’ apples. They developed all the acetates with hexyl acetate as the main contributing odour compound, all the C6 aldehydes characteristic of ripe stored apples and 3-penten-2-01 (Visai et al., 1993). PBZ-treated apples produced these compounds in lower amounts, except for hexanal, which was slightly higher, and 3-penten-2-01, which was not produced. Furthermore, only PBZ-treated apples emitted a large amount of ethyl 2-methylbutanoate, which was the main contributing compound, making it seem as if they had been stored for a longer period. In fact, this ester is a character impact compound for Delicious apples (Flath et al.. 1967), and according to Yahia et al. (1990) is a postharvest-induced volatile, present in very high concentrations in apples ripened off the tree and after long periods of cold storage. By prolonging the storage time up to 7 months there was a considerable change in the aroma composition of control apples, as all acetates decreased, 3-penten-2-01 was not detected and ethyl 2-methylbutanoate developed to high levels (92 Uo). the latter being the main contributing compound. The volatile pattern of PBZ-treated fruits did not change by prolonging the storage time and was similar to the one reported for control fruit after 7 months of storage. Our findings could indicate that PBZ treatment enhanced the ripening processes which are associated with volatile evolution both at harvest and after cold storage. In addition. PBZ may have a specific effect on the metabolism of a few odouractive compounds. i.e. pentyl acetate and ethyl butanoate. As PBZ-treated apples had an aromatic pattern related to ripeness already at the first harvest, and further storage would only increase the overripe aspect of aroma, treated apples should be consumed on the fresh market or be stored only for a short period. Further studies are needed to understand the mode of action of PBZ treatment, and to find out if PBZ has a specific and selective effect on the metabolism of the odour-active volatile compounds.
and Technolox),
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45
References Bangerth, F., 1986. Effect of ethylene biosynthesis and/or action on the production of volatile flavour substances of stored apple fruit. Acta Hortic., 179: 82% 824. Cunningham, D.G., Acree, T.E.. Barnard, J., Butts, R. and Braell, P.. 1986. Analysis of apple volatiles. Food Chem., 19: 137-147. Dimick. P.S. and Hoskin, J.C.. 1983. Review of apple flavour: state of the art. Crit. Rev. Food Sci. Nutr., 18: 387 ~409. Elfving. D.C.. Chu, C.L.. Lougheed, E.C. and Cline. R.A., 1987. Effects of daminozide and paclobutrdzol treatments on fruit ripening and storage behaviour of ‘McIntosh’ apple. J. Am. Sot. Hortic. Sci., 112: 910 -915. Flath. R.A. Black, D.R.. Guadagni, D.G., McFadden, W.H. and Schultz. T.H., 1967. Identification and organoleptic evaluation of compounds in Delicious apple essence. J. Agric. Food Chem., IS: 29- 3.5. Guadagni, D.G., Buttery, R.G. and Harris. J.. 1966. Odoul intensities of hop oil components. J. Sci. Food Agric.. 17: 142. 144. Herrmann. K.. 1991. Die Aromastoffe des Obstes. Teil I: Allgemeine Angaben zu den Aromastoffen. ihren Schwellenwerten und ihrer Zusammensetzung. Erwerbsobstbau. 33: 4 7. Luo. Y.. Wainwright. H. and Moore. K.G.. 1987. Effect of orchard application of paclobutrazol on the post-harvest ripening of apples. J. Hoi-tic. Sci.. 62: 295 301. Paillard. N.M.N.. 1981. Factors influencing flavour formation in fruits. 1n: P. Schreier (Editor). Flavour 81. Walter de Gruyter. Berlin, pp. 479-507. Panasiuk. 0.. Talley. F.B. and Sapers. G.M.. 1980. Correlation between aroma and volatile composition of Mclntosh apples. J. Food Sci., 45: 989~ 991. Rizzolo, A.. Teleky-Vamossy. Gy. and Polesello. A.. 1989. Capillary GCsensory analysis of volatile compounds developed from ripening apple fruit. J. High Rcsol. Chromatogr., 12: 824 827. Rizzolo. A.. Polesello, A. and Polesello. S., 1992. Use of headspace capillary GC to study the development of xolatilc compounds in fresh fruit. J. High Resol. Chromatogr.. 15: 472-477. Rizzolo. A.. Visai, C. and Vanoli, M., 1993. Influence of paclobutrdzol on the quality of apples during growth. Acia Hortic.. 329: I40 142. Rizzolo. A.. Visai. C. and Vanoli. M., 1995. Volatile compounds during fruit ripening: state of the art. In: COST94 Workshop on Ripening and Senescence, Federal Research Centre for Nutrition. Karlsruhe, June 22 23. 1992. in press. Salunkhe. D.K. and Do, J.Y.. 1976. Biogenesis of aroma constituents of fruits and vegetables. Crit. Rev. Food Sci. Nutr., 6: I61 190. Steffens. G.L. and Wang, S.Y., 1986. Biochemical and physiologxal alterations in apple trees caused by a gibberelhn biosynthesis inhibitor. paclobutrazol. Acta Hortic., 179: 433 442
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Steffens, G.L., Stafford, A.E. and Lin, J.T., 1991. Influence of an inhibitor of gibberellin biosynthesis, paclobutrazol, on apple seed gibberellin content. Physiol. Plant, 83: 366-372. Vanoli, M., Visai, C. and Rizzolo A., 1995. The influence of harvest date on the volatile composition of Starkspur Golden apples. Postharvest Biol. Technol., 6: 2255234. Visai, C., Rizzolo, A. and Vanoli, M., 1993. Sviluppo delle sostanze volatili nelle mele durante la conservazione frigorifera. Riv. Frutticoltura, 55: 57760. Wang, C.Y. and Steffens, G.L., 1987. Postharvest responses of
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Spartan apples to preharvest paclobutrazol treatment. Hortic. Sci., 2: 276-278. Willaert, G.A., Dirinck, P.J., De Pooter, H.L. and Schamp, N.N., 1983. Objective measurement of aroma quality of Golden Delicious apples as a function of controlled atmosphere storage time. J. Agric. Food Chem., 31:809-813. Yahia, E.M., Acree, T.E. and Liu, F.W., 1990. The evolution of some odour-active volatiles during the maturation and ripening of apples on the tree. Lebensm. Wiss. Technol., 23: 488-493.
Postharvest Biology and Technology 11 (1997) 39946
Changes in some odour-active compounds in paclobutrazol-treated ‘Starkspur Golden’ apples at harvest and after cold storage Anna
Rizzolo a,*, Costanza
Visai b, Maristella
Vanoli b
r’Istiruto Sperimentale per la Valorizzazione Tecnologica dei Prodotti Agricoli (I. V. T.P.A.), cia Venetian, 26-20133 Milano. Itall h Istituto di Coltivarioni Arhoree, Universith degli Studi, via Celoria, 2-20123 Milano. Ital?
Accepted 21 January 1997
Abstract Apple trees (M&s domestica Borkh. cv ‘Starkspur Golden’) were treated 25 days after full bloom (DAFB) with paclobutrazol (PBZ) as a soil-drench application. After 2 years, fruit were picked 158 (commercial harvest), 172 and 181 DAFB from treated and untreated trees, with fruit taken at commercial harvest being stored in air at 2°C and 90% R.H. During ripening and after 5 and 7 months of storage, treated and untreated apples were analysed for odour-active volatiles, by using either static or dynamic headspace samplings and capillary gas chromatography. The results were expressed in terms of Odour Units (Uo) in order to relate the relative importance of each compound to overall aroma. The average Uo values at harvest and after storage showed that only half of the monitored volatiles were present in concentrations great enough to contribute to overall aroma. PBZ treatment enhanced the ripening processes, which are associated with volatile evolution at harvest and after 5 months of air storage, and may have a specific effect on the metabolism of pentyl acetate and ethyl butanoate. 0 1997 Elsevier Science B.V. Keywords:
Apples;
Paclobutrazol;
Volatiles;
Ripening;
Air storage
1. Introduction Apple flavour is a complex combination of taste and odour sensations. Although taste and texture are important for apple quality, the presence of * Corresponding 2365377. 0925-5214/97!$17.00
author.
Tel.: + 39 2 70630256;
6 1997 Elsevier
PII SO925-5214(97)01417-S
fax: + 39 2
Science B.V. All rights
reserved
trace amounts of volatiles, responsible for the characteristic odour, gives fruit much of their perceived quality (Dimick and Hoskin, 1983). Various studies have shown that the production of volatile compounds in climacteric fruits, such as apples, is related to the climacteric rise in respiration (Salunkhe and Do, 1976). However, little information exists about their evolution, and
40
A. Rizzolo et al. /Postharvest
Biology and Technology I1 (1997) 39-46
especially on whether the few volatiles which have a significant impact on apple sensory quality behave in the same way as any other odourless compounds. Numerous factors such as variety, climate, date of harvest and storage (Dimick and Hoskin, 1983; Rizzolo et al., 1995) affect the levels of apple volatile compounds. Usually, apples destined for storage are harvested prior to the onset of the climacteric in order to delay the formation of ethylene and ripening. Paclobutrazol [/3-[(4chlorophenyl) methyl - CI- ( 1, 1- dimethylethyl) - 1H1,2,4-triazol-1 -ethanol] (PBZ) has been widely shown to be active in controlling the vegetative growth of many fruit crops; generally it reduces ethylene production, delays fruit maturation and ripening, enhances fruit set and limits water loss (Steffens and Wang, 1986). In fruit after cold storage in air, PBZ also causes a lower ethylene production, and a delayed CO, climacteric (Elfving et al., 1987; Luo et al., 1987). However, the type of effect strictly depends on the cultivar and the time of PBZ application. As an example, Wang and Steffens (1987) found no differences in respiration and ethylene evolution between untreated and PBZ-treated ‘Spartan’ apples. Few researchers have considered the influence of growth regulators on aroma compounds during apple fruit ripening and storage. Bangerth (1986) observed in aminoethoxyvinylglycine-treated ‘Golden Delicious’ apples no climacteric-like increase in CO, production and a greatly reduced emission of volatiles during cold storage. Yahia et al. (1990) found that daminozide adversely affected volatile composition in ‘Cortland’ and ‘McIntosh’ apples, delaying the onset of climacteric ethylene production and the formation of five odour-active volatiles ((E)-2-hexenal, methyl, butyl, hexyl hexanoate and hexyl 2-methylbutanoate) during fruit maturation and after storage in low-ethylene controlled atmosphere. Etcher (personal communication) found in ‘Starkspur Golden’ apples that soil-drench PBZ treatment was effective in controlling vegetative growth up to 3 years after the treatment, reducing internode length and leaf size, and producing fruit with reduced size and length/diameter ratio com-
pared with the control apples. For the fruit shape and size the effect of the treatment was very noticeable in the first year, but this diminished gradually over the next 2 years. Rizzolo et al. (1993), studying the quality of these ‘Starkspur Golden’ apples during fruit growth (from 50 to 180 days after full bloom), showed that even after 2 years, soil-drench PBZ treatment had an influence on the starch content and on total volatile evolution, specially during maturation and ripening, whilst it did not affect titratable acidity and soluble solids. It is not known whether PBZ has adverse effects also on the qualitative composition of apples during ripening and after cold storage. The purpose of this research was to study how soil-drench PBZ application on ‘Starkspur Golden’ apple trees affected the composition of volatiles during maturation and after air storage, paying particular attention to some odour-active compounds.
2. Materials and methods PBZ treatment and storage Apple trees, 17 year old (Malus domestica Borkh. cv ‘Starkspur Golden’) grafted on MM1 11 and grown in the Experimental Orchard of the University of Milan (Montanaso Lombardo, PO Valley), were used. One group of five trees was treated 25 days after full bloom (DAFB, full bloom occurring on April 16) with PBZ as a soil-drench around the trees (3 g/tree). A second group of five untreated trees was used as control. After 2 years, five apples/tree were picked 158 (commercial harvest), 172 and 181 DAFB (FB occurred on April 16) from treated and untreated (control) trees, when soluble solids were 1 l- 11.5” Brix and titratable acidity was 6.3 meq/lOO ml of juice for early picked apples, 3.5 meq/lOO ml of juice for 172 DAFB fruit and 2 meq/lOO ml of juice for the apples from the last harvest. Fruits were pooled into two replications and analysed for volatile composition by static headspace/capillary gas chromatography on a Carbowax 20 M column (25 m x 0.5 mm i.d., 1 pm film thickness) according to the conditions reported by Rizzolo et
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et al. /Postharvest
Biology and Technology
al. (1992). Each sample was obtained by pooling a slice per fruit and mincing the combined tissue. At commercial harvest (158 DAFB), 80 fruits from control .and treated trees were collected and stored in air at 2°C and 90% R.H. After 5 and 7 months of storage, 20 apples per treatment, divided into two replications, were analysed for volatile compound evolution during post-storage ripening at 18°C by dynamic headspaceicapillary gas chromatography on a 12 m x 0.33 mm i.d. Carbowax 20 M column (0.5 pm film thickness) following the conditions described in previous papers (Rizzolo et al., 1992; Vanoli et al., 1995). Volutile ident$cation and quarhjication Identification of volatile compounds was performed according to their retention times, comparing them with those of commercial standards and/or with the data obtained by gas chromatography/mass spectrometric analysis using a Hewlett&Packard 5870B Quadrupole Mass Spectrometer operated in the electron ionization mode at 70 eV and identified via National Bureau of Standards, Washington, D.C. library match. Volatiles were quantified using flame ionization detection by relating the peak area of each compound to that of an external standard (hexyl acetate) and were expressed as pggikg of fruit flesh for static headspace, and as pg/kg per h for dynamic headspace. To relate the relative importance of each compound to overall aroma, concentrations were converted into Uo. This representation of volatile pattern is an approximation, but has a practical use in selecting the most important contributors to aroma (Guadagni et al., 1966). Uo were calculated from concentrations using odour thresholds reported in the literature (Herrmann, 1991), according to: Uo = C/OT, where C = concentration in the sample and OT = odour detection threshold concentration. Compounds with Uo equal to or greater than 1.0 significantly contribute to aroma as they are above their odour threshold concentration.
I1 (1997) 39.-46
41
3. Results Volatile composition The odour-active compounds monitored in ‘Starkspur Golden’ apples and their odour threshold concentrations are listed in Table 1. Some compounds, such as butanol, hexanol, butyl propanoate, hexyl butanoate, butyl hexanoate and acetaldehyde show high odour threshold concentration (above 100 ppb) so they could contribute to aroma only if they are present in very high amounts. Other compounds, such as ethyl butanoate and ethyl 2-methylbutanoate could contribute to aroma even if they are present in very low quantities, as their odour threshold concentrations are below 1 ppb. In considering the average odour units (Av Uo) at harvest and after storage (Table 2), only half of the compounds monitored actually contributed to the overall aroma. The main odour contributors were ethyl butanoate, hexyl acetate, ethyl 2methylbutanoate, butyl acetate, hexanal and Table 1 Odour-active volatiles monitored in ‘Starkspur and their Odour Thresholds (OT) (Herrmann, Volatiles
OT(ppb)
Esters Butyl acetate Pentyl acetate Hexyl acetate (Z)-3-hexenyl acetate Propyl propanoate Butyl propanoate Ethyl butanoate Propyl butanoate Hexyl butanoate 3-methylbutyl butanoate Ethyl 2-methylbutanoate Hexyl 2-methylbutanoate Butyl hexanoate
18 250 21 0.1 22 250
Aldehydes Acetaldehyde Hexanal (E)-2-hexenal
120 4.5 17
Alcohol.~ Butanol 3-penten-2-01 Hexanol
500 4.2 150 _
66 5 2 2 57 200
I
Golden’ 1991)
apples
42
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Table 2 Average odour unit values (Av Uo) of volatile compounds in ‘Starkspur Golden’ apples at harvest, and after 5 and 7 months of air storage Compounds
Av Uo Harvest
Ethyl butanoate Hexyl acetate Ethyl 2-methylbutanoate Butyl acetate 3-penten-2-01 Hexanal Pentyl acetate Propyl butanoate (E)-2-hexenal Hexyl 2-methylbutanoate Acetaldehyde (Z)-3-hexenyl acetate Propyl propanoate Butanol Hexanol Hexyl butanoate+ butyl hexanoate Butyl propanaote 3-methylbutyl acetate
Storage 5 months
7 months
IS 31 6 3 1.1 1 1 0.8 0.3 0.27 0.2 0.15 0.1 0.09 0.03 0.03
10 48 9 2 0.2 2 3 0 1.5 0.1 0.05 0.06 0 0.06 0.14 0.05
9 24 38 1 0 2 2 0 0.8 0.002 0.05 0.003 0 0.04 0.08 0.02
0.01 0
0.04 0.05
11 (1997) 39-46
increasing amounts of hexanal, even if they were always below its odour threshold (Uo < 1); with ripening, control fruits produced decreasing quantities of hexanal, with Uo > 1 at 158 and 181 DAFB. A characteristic compound of ‘Golden’ apple aroma, 3-penten-2-01, was always detectable from organoleptic analysis in control fruits, while in PBZ-treated apples it contributed to the aroma only at 18 1 DAFB. Differences were also found among the esters. In PBZ-treated fruits, pentyl acetate was detected only at 172 DAFB at very low amounts and not significant for odour. Ethyl butanoate levels increased although the amounts at 158 and 172 DAFB were much lower than in control fruits. Control apples, on the other hand, produced increasing amounts of pentyl acetate, always 158 DAFB loguo ii,rl,
-;-~TYI~
0 0 _,,5 I-
___-. hex.4
pentyl acetate; other compounds (3-penten-2-01, (E)-2-hexenal and propyl butanoate) were present in amounts equal or close to the odour threshold concentration. All the other volatiles monitored had Uo values too low to contribute to the overall aroma and for this reason they will be not considered in the discussion. The major odour contributors changed from harvest to storage (Table 2). At harvest more than half of the total Uo were made up of ethyl butanoate, after 5 months of storage hexyl acetate, and after 7 months ethyl 2-methylbutanoate. In addition, with storage, propyl butanoate and 3-penten-2-01 no longer contributed to aroma, while (E)-2-hexenal increased and became detectable. Influence of PBZ treatment at harvest The PBZ treatment influenced the trend of some odour-active volatile compounds during ripening (Fig. 1). PBZ-treated apples produced
3p201
butac
penac
hexac
etbut
probut
e2mb
172 DAFB logIJO 24 T_-----
~--
1,5 +--
_::: -:‘oT~_E
TL!!
-I,5 hexal
_,,5 ~ _... hexal
3p201
..__
butac
penac
hexac
etbut
probut
e2mb
penac
hexac
etbut
probut
e2mb
.___~~~~~~~ 3p201
butac
Fig. 1. Odour-active compounds (log Uo) of control (W) and PBZ-treated (0) ‘Starkspur Golden’ apples at 158, 172 and 181 DAFB. No line drawn on x axis means that the compound is absent. Compound abbreviations: hexal = hexanal, 3p201= 3-penten-2-01, butac = butyl acetate, penac = pentyl acetate, hexac = hexyl acetate, etbut = ethyl butanoate, probut = propyl butanoate, e2mb = ethyl 2-methylbutanoate.
A. Rirzolo et al. /Postharoest Biology and Technology I I (1997) 39-46
present in amounts above its odour threshold, and very high levels of ethyl butanoate at 158 and 172 DAFB (about 200-220 Uo). At 181 DAFB, this ester was produced in an amount similar to that in PBZ-treated fruits (about 8 Uo). Butyl acetate and hexyl acetate increased with harvest both in control and in PBZ-treated apples; the latter fruits, however, developed higher Uo of these esters at 158 and 181 DAFB. Propyl butanoate followed an increasing trend with ripening both in PBZ-treated and control apples, exceeding the odour threshold concentration only at the last harvest date (181 DAFB), with control apples producing the higher amount. Ethyl 2-methylbutanoate had an increasing trend in control and PBZ-treated fruits; this ester was always higher in PBZ-treated apples. The main volatile compound which contributed to odour was ethyl butanoate in control fruits at 158 and 172 DAFB and hexyl acetate in control fruit at 18 1 DAFB and in PBZtreated apples at 158, 172 and 181 DAFB. Influence (f PBZ treatment after air storage Propyl butanoate was not detected in control and PBZ-treated apples after 5 and 7 months of air storage. PBZ-treated apples stored for 5 months (Fig. 2) with post-storage ripening at 18°C developed a relatively constant amount of hexanal and ethyl butanoate, comparable with those of control fruit. (E)-2-hexenal and butyl acetate showed a decreasing trend in PBZ-treated apples, while control fruit produced increasing amounts of these compounds, reaching the higher Uo values on the 8th day of post-storage ripening, when the fruits were fully matured. Pentyl and hexyl acetate were almost constant during post-storage ripening in PBZ-treated apples, while in control fruits they steeply increased on the 8th day. At the end of the post-storage ripening, only control apples produced 3-penten-2-01, and only PBZ-treated fruit developed ethyl 2-methylbutanoate. Comparing the Uo values on the 8th day of post-storage ripening, i.e. when fruits were fully
log
“0
PBZ-treated
43
apples
2.50 --/
Control apples
Fig. 2. Odour-active compounds (log Uo) of control and PBZtreated ‘Starkspur Golden’ apples after five months of air storage and 1 (open area), 2 (horizontal rules). 3 (diagonal rules). 7 (vertical rules) and 8 (filled area) days of post-storage ripening. No line drawn on x axis means that the compound is absent. Compound abbreviations: hexal = hexanal. t2hexal = (E)-2-hexenal. 3~201 = 3-penten-2-ol. butac = butyl acetate. penac = pentyl acetate, hexac = hexyl acetate. etbut = ethyl butanoate. e2mb = ethyl 2-methylbutanoate.
ripe (Fig. 2), the main contributing compound to odour in control fruit was hexyl acetate, while ethyl 2-methylbutanoate was the main compound in PBZ-treated fruits. Fully ripe control fruit were characterized by higher amounts of (E)-2-hexenal, 3-penten-2-01, butyl acetate, pentyl acetate and hexyl acetate. After 7 months of storage (Fig. 3) control and PBZ-treated apples did not emit 3-penten-201. PBZ-treated apples produced butyl acetate, pentyl acetate, hexyl acetate, (E)-2-hexenal. ethyl 2-methylbutanoate and ethyl butanoate with a behaviour similar to control fruits. In addition, (E)-2-hexenal was above the odour threshold only in PBZ-treated apples. Comparing the Uo values at the end of the post-storage ripening (Fig. 3), in both control and PBZ-treated apples the main contributing compound was ethyl 2-methylbutanoate.
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Biology and Technology 11 (1997) 39-46
4. Discussion
The odour-active compounds monitored in ‘Starkspur Golden’ apples have all been shown to be odour-active in ready-to-eat ‘Golden Delicious’ apple aroma (Rizzolo et al., 1989) and in a combined apple extract of various cultivars (Cunningham et al., 1986). However, by converting concentrations into Uo, not all these compounds can be shown to contribute to the formation of the aroma. Except for 3-penten-2-01, all the alcohols monitored were present in concentrations far below their odour thresholds. The same was found for the propanoates and butanoates of higher molecular weight. Thus, all these compounds could not significantly contribute to the aroma. The contributing volatiles collected from ‘Starkspur Golden’ apples treated with PBZ differed from those in control fruits considerably at harvest and after 5 months of storage in air. This is consistent with the findings of other authors (Elfving et al., 1987; Steffens et al., 1991), showPBZ-treated
apples
Control apples
0.5 0
Fig. 3. Odour-active compounds (log Uo) of control and PBZtreated ‘Starkspur Golden’ apples after 7 months of air storage and 1 (M), 2 (0) and 3 (W) days of post-storage ripening. No line drawn on x axis means that the compound is absent. Compound abbreviations: hexal = hexanal, t2hexal= (E)-2hexenal, butac = butyl acetate, penac = pentyl acetate, hexac = hexyl acetate, etbut = ethyl butanoate, e2mb = ethyl 2-methylbutanoate.
ing the influence of soil drench treatment with PBZ on ethylene production, fruit maturation and ripening, and gibberellin composition over a 3 year experimental period. In agreement with our previous findings on ‘Starkspur Golden’ apples (Vanoli et al., 1995) 172 DAFB control fruits had a volatile composition typical of ripe apples, i.e. a higher content of acetate esters, responsible for the ether-like, sweet, fruity characteristic aroma of ‘Golden’ apples, together with a high amount of ethyl butanoate, responsible for a powerful ether-like/fruity odour. 181 DAFB control apples showed a composition typical of overripe fruits, having the highest amounts of ethyl 2-methylbutanoate and detectable amounts of propyl butanoate, the former having a powerful diffusive green-fruity odour (Panasiuk et al., 1980; Willaert et al., 1983). Control apples picked at commercial harvest produced the highest quantities of hexanal and the lowest amounts of acetate esters; this pattern indicates that the commercial harvest time corresponds to unripe aroma quality (Panasiuk et al., 1980; Paillard, 1981; Willaert et al., 1983). PBZ treatment severely influenced the volatile composition at each harvest date, changing the odour profile. Treated apples never developed pentyl acetate, which gives a sweet-fruity odour. Hexanal was present in low amounts below its odour threshold; this aldehyde is responsible for a fatty-green, grassy odour. 3-penten-2-01, typical of ripe ‘Golden’ apples, exceeded the odour threshold only at the last harvest date. Contrary to control fruits, PBZ-treated apples always had as the main contributing odour compounds hexyl acetate (sweet, fruity) and ethyl 2-methylbutanoate (green-fruity). These patterns are close to those in control 181 DAFB fruits; thus, PBZtreated apples have an aromatic pattern more related to ripeness/overripeness at each harvest date. The fact that PBZ-treated apples produced less hexanal, no pentyl acetate, less ethyl butanoate, and more butyl acetate, hexyl acetate and ethyl 2-methylbutanoate could indicate that PBZ has a selective effect on the metabolism of these compounds.
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Biology
Differences between control and PBZ-treated apples were also found after storage in air. After 5 months at the end of post-storage ripening, control apples showed an aromatic pattern characteristic of ripe ‘Starkspur Golden’ apples. They developed all the acetates with hexyl acetate as the main contributing odour compound, all the C6 aldehydes characteristic of ripe stored apples and 3-penten-2-01 (Visai et al., 1993). PBZ-treated apples produced these compounds in lower amounts, except for hexanal, which was slightly higher, and 3-penten-2-01, which was not produced. Furthermore, only PBZ-treated apples emitted a large amount of ethyl 2-methylbutanoate, which was the main contributing compound, making it seem as if they had been stored for a longer period. In fact, this ester is a character impact compound for Delicious apples (Flath et al.. 1967), and according to Yahia et al. (1990) is a postharvest-induced volatile, present in very high concentrations in apples ripened off the tree and after long periods of cold storage. By prolonging the storage time up to 7 months there was a considerable change in the aroma composition of control apples, as all acetates decreased, 3-penten-2-01 was not detected and ethyl 2-methylbutanoate developed to high levels (92 Uo). the latter being the main contributing compound. The volatile pattern of PBZ-treated fruits did not change by prolonging the storage time and was similar to the one reported for control fruit after 7 months of storage. Our findings could indicate that PBZ treatment enhanced the ripening processes which are associated with volatile evolution both at harvest and after cold storage. In addition. PBZ may have a specific effect on the metabolism of a few odouractive compounds. i.e. pentyl acetate and ethyl butanoate. As PBZ-treated apples had an aromatic pattern related to ripeness already at the first harvest, and further storage would only increase the overripe aspect of aroma, treated apples should be consumed on the fresh market or be stored only for a short period. Further studies are needed to understand the mode of action of PBZ treatment, and to find out if PBZ has a specific and selective effect on the metabolism of the odour-active volatile compounds.
and Technolox),
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45
References Bangerth, F., 1986. Effect of ethylene biosynthesis and/or action on the production of volatile flavour substances of stored apple fruit. Acta Hortic., 179: 82% 824. Cunningham, D.G., Acree, T.E.. Barnard, J., Butts, R. and Braell, P.. 1986. Analysis of apple volatiles. Food Chem., 19: 137-147. Dimick. P.S. and Hoskin, J.C.. 1983. Review of apple flavour: state of the art. Crit. Rev. Food Sci. Nutr., 18: 387 ~409. Elfving. D.C.. Chu, C.L.. Lougheed, E.C. and Cline. R.A., 1987. Effects of daminozide and paclobutrdzol treatments on fruit ripening and storage behaviour of ‘McIntosh’ apple. J. Am. Sot. Hortic. Sci., 112: 910 -915. Flath. R.A. Black, D.R.. Guadagni, D.G., McFadden, W.H. and Schultz. T.H., 1967. Identification and organoleptic evaluation of compounds in Delicious apple essence. J. Agric. Food Chem., IS: 29- 3.5. Guadagni, D.G., Buttery, R.G. and Harris. J.. 1966. Odoul intensities of hop oil components. J. Sci. Food Agric.. 17: 142. 144. Herrmann. K.. 1991. Die Aromastoffe des Obstes. Teil I: Allgemeine Angaben zu den Aromastoffen. ihren Schwellenwerten und ihrer Zusammensetzung. Erwerbsobstbau. 33: 4 7. Luo. Y.. Wainwright. H. and Moore. K.G.. 1987. Effect of orchard application of paclobutrazol on the post-harvest ripening of apples. J. Hoi-tic. Sci.. 62: 295 301. Paillard. N.M.N.. 1981. Factors influencing flavour formation in fruits. 1n: P. Schreier (Editor). Flavour 81. Walter de Gruyter. Berlin, pp. 479-507. Panasiuk. 0.. Talley. F.B. and Sapers. G.M.. 1980. Correlation between aroma and volatile composition of Mclntosh apples. J. Food Sci., 45: 989~ 991. Rizzolo, A.. Teleky-Vamossy. Gy. and Polesello. A.. 1989. Capillary GCsensory analysis of volatile compounds developed from ripening apple fruit. J. High Rcsol. Chromatogr., 12: 824 827. Rizzolo. A.. Polesello, A. and Polesello. S., 1992. Use of headspace capillary GC to study the development of xolatilc compounds in fresh fruit. J. High Resol. Chromatogr.. 15: 472-477. Rizzolo. A.. Visai, C. and Vanoli, M., 1993. Influence of paclobutrdzol on the quality of apples during growth. Acia Hortic.. 329: I40 142. Rizzolo. A.. Visai. C. and Vanoli. M., 1995. Volatile compounds during fruit ripening: state of the art. In: COST94 Workshop on Ripening and Senescence, Federal Research Centre for Nutrition. Karlsruhe, June 22 23. 1992. in press. Salunkhe. D.K. and Do, J.Y.. 1976. Biogenesis of aroma constituents of fruits and vegetables. Crit. Rev. Food Sci. Nutr., 6: I61 190. Steffens. G.L. and Wang, S.Y., 1986. Biochemical and physiologxal alterations in apple trees caused by a gibberelhn biosynthesis inhibitor. paclobutrazol. Acta Hortic., 179: 433 442
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Steffens, G.L., Stafford, A.E. and Lin, J.T., 1991. Influence of an inhibitor of gibberellin biosynthesis, paclobutrazol, on apple seed gibberellin content. Physiol. Plant, 83: 366-372. Vanoli, M., Visai, C. and Rizzolo A., 1995. The influence of harvest date on the volatile composition of Starkspur Golden apples. Postharvest Biol. Technol., 6: 2255234. Visai, C., Rizzolo, A. and Vanoli, M., 1993. Sviluppo delle sostanze volatili nelle mele durante la conservazione frigorifera. Riv. Frutticoltura, 55: 57760. Wang, C.Y. and Steffens, G.L., 1987. Postharvest responses of
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Spartan apples to preharvest paclobutrazol treatment. Hortic. Sci., 2: 276-278. Willaert, G.A., Dirinck, P.J., De Pooter, H.L. and Schamp, N.N., 1983. Objective measurement of aroma quality of Golden Delicious apples as a function of controlled atmosphere storage time. J. Agric. Food Chem., 31:809-813. Yahia, E.M., Acree, T.E. and Liu, F.W., 1990. The evolution of some odour-active volatiles during the maturation and ripening of apples on the tree. Lebensm. Wiss. Technol., 23: 488-493.