Postharvest Biology and Technology 35 (2005) 167176

Effect of commercial conditioning and cold quarantine storage

treatments on fruit quality of Rouge La Toma grapefruit
(Citrus paradisi Macf.)
Andrea Biolattoa, , Daniel E. Vazquezb , Ana M. Sanchoa ,
Fernando J. Carduzaa , Norma A. Pensela
a Centro de Investigacion de Agroindustria, Instituto Tecnologa de Alimentos, Instituto Nacional de Tecnologa Agropecuaria (INTA),
CC 77 (B1708WAB) Moron, Buenos Aires, Argentina
b Estacion Experimental Agropecuaria-Concordia, Instituto Nacional de Tecnologa Agropecuaria (INTA), Entre Ros, Argentina

Received 28 March 2004; accepted 23 August 2004

Abstract

We evaluated the effect of cold quarantine treatments on the development of CI and temperature conditioning on the induction
of low temperature tolerance in Rouge La Toma grapefruit (Citrus paradisi Macf.), and their effects on acetaldehyde, ethanol
and d-limonene contents and sensory characteristics. Various treatments, non-conditioned + quarantine at 2 C and 85% RH for
18 days + storage at 13 C and 85% RH for 4 days or 17 days + marketing period at 20 C and 85% RH for 7 days; conditioned
at 15 C and 85% RH for 7 days + quarantine at 2 C and 85% RH for 18 days + storage at 13 C and 85% RH for 4 days or
17 days + marketing period at 20 C and 85% RH for 7 days; storage at 13 C and 85% HR for 22 days or 35 days + marketing
period at 20 C and 85% RH for 7 days (control treatments), were assayed. By the end of the simulated marketing period,
the conditions did not promote chilling injury development in Rouge La Toma grapefruit. After the simulated marketing
period, acetaldehyde, ethanol and d-limonene contents were not affected in fruit stored under treatments that included cold
quarantine. In some cases, treatments that included temperature conditioning significantly increased acetaldehyde and ethanol
levels, however, the amounts detected were comparable with fresh grapefruit juice. In general, the storage times involved in the
treatments assayed, did not promote increases in the acetaldehyde and ethanol levels. Conversely, fruit stored at non-chilling
temperatures (control treatments) had higher levels of d-limonene compared to those which underwent treatments that included
cold quarantine. In addition, a significant increase in d-limonene levels between the initial time of the treatments and the end of the
marketing period was observed, for all treatments, with the highest and most variables levels observed in fruit stored under control
treatments. Sensory characteristics, such as sweet, acid and bitter taste and typical flavor intensity, in general, were not affected
by the postharvest handling practices applied. Therefore, it can be concluded that the cold quarantine treatment and temperature

Corresponding author. Tel.: +54 11 4621 0446; fax: +54 11 4621 2012.
E-mail address: abiolatto@cnia.inta.gov.ar (A. Biolatto).

0925-5214/$ see front matter 2004 Elsevier B.V. All rights reserved.
doi:10.1016/j.postharvbio.2004.08.002
168 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176
conditioning may have important commercial applications for Rouge La Toma grapefruit (Citrus paradisi Macf.) without
adversely affecting its quality.
2004 Elsevier B.V. All rights reserved.
Keywords: Citrus paradisi; Postharvest treatments; Acetaldehyde; Ethanol; d-Limonene; Sensory characteristic
1. Introduction
Rouge La Toma, a natural mutation of grapefruit
(Citrus paradisi Macf.) cultivars selected in Salta, a
northwestern province of Argentina, is a pigmented va-
riety appreciated by the consumer because of its flavor
and general appearance. Quarantine treatment for pest
disinfestations of fruit commodities is essential for in-
ternational trade. Cold quarantine treatment, which in-
volves the exposure of fruit to near-freezing tempera-
tures for a specified period, is a procedure accepted for
Mediterranean fruit fly Ceratitis capitata (Wied) dis-
infestation of citrus fruit by the regulatory agencies of
most importing countries and is currently applied on
a commercial scale. However, many citrus cultivars,
particularly grapefruit, are sensitive to chilling injury
(CI) (Schiffmann-Nadel et al., 1971; Grierson, 1974;
Chalutz et al., 1981) and may develop peel injury when
exposed to cold treatment, which greatly reduces the
fruit marketability. The primary cause of chilling injury
is thought to be plant cell membrane damage. This dam-
age initiates a cascade of secondary reactions, which
may include: ethylene production, increased respira-
tion, reduced photosynthesis, interference in energy
production, accumulation of toxic compounds such as
ethanol and acetaldehyde, and disruption at the cellular
and subcellular structure (Lyons, 1973; Wang, 1982).
Moreover, most of the CI is limited to the flavedo, the
portion of the rind that contains large amounts of es-
sential oils such as d-limonene (Obenland et al., 1996),
which is an abundant volatile component in citrus fruit
and important for grapefruit flavor (Sun and Petracek,
1999). Several postharvest heat treatments have been
reported to induce fruit tolerance to cold temperature
and to reduce the development of CI symptoms during
cold storage and cold quarantine treatments (Hatton,
1990; Wang, 1993, 1994; Lurie, 1998a, 1998b; Porat
et al., 2000, 2003). In citrus, these heat treatments
include prestorage conditioning for either 3 days at
21 C or 7 days at 16 C (Grierson, 1974; Hatton and
Cubbedge, 1982a, 1982b, 1983; McDonald, 1986),
hot water dips (HWD) at 53 C for 23 min (Wild
and Hood, 1989; Rodov et al., 1995; Schirra and
Dhallewin, 1997; Shirra et al., 1997), and curing for 3
days at 36 C under water-saturated conditions (Ben-
Yehoshua et al., 1989; Rodov et al., 1995; Mulas
et al., 1996). The objective of the present study was
to evaluate the effect of cold quarantine treatment on
the development of CI and temperature conditioning on
the induction of low temperature tolerance in Rouge
La Toma grapefruit (Citrus paradisi Macf.), and their
effect on acetaldehyde, ethanol and d-limonene levels
and sensory characteristics.
2. Materials and methods
2.1. Plant material and postharvest treatments
The variety of grapefruit (Citrus paradisi Macf.)
used was Rouge La Toma, a natural mutation selected
in the Argentinian northern province of Salta, harvested
in June 1999. Grapefruit subjected to temperature con-
ditioning were harvested 7 days earlier than fruit dis-
tributed among other treatments. Fruit were degreened
for 80 h with 3.5 mg/kg ethylene and 1.5 mg/kg CO2
at a temperature of 26 C and 90% relative humidity.
Then, all fruit were washed, disinfected with 0.5/100 g
sodium orthophenylphenate (SOPP), rinsed, dried and
coated with a polyethylene-based wax (18/100 g solid
matter) containing 5000 mg/L thiabendazole (TBZ).
After that, medium size fruit, free from defects, were
selected, and placed into boxes (40 individually num-
bered fruits) before being transported from the packing-
house to our laboratory, located about 1200 km south-
east (a 36 h trip), in refrigerated transport at 7 1 C.
Six commercial treatment groups of six boxes each
were assayed: (T1) non-conditioned + quarantine at
2 C and 85% RH for 18 days + storage at 13 C and
85% RH for 17 days; (T2) conditioned at 15 C and
 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176
85% RH for 7 days + quarantine at 2 C and 85% RH
for 18 days + storage at 13 C and 85% RH for 17
days; (T3) storage at 13 C and 85% HR for 35 days;
(T4) non-conditioned + quarantine at 2 C and 85%
RH for 18 days + storage at 13 C and 85% RH for
4 days; (T5) conditioned at 15 C and 85% RH for 7
days + quarantine at 2 C and 85% RH for 18 days +
storage at 13 C and 85% RH for 4 days; (T6) storage
at 13 C and 85% HR for 22 days. For all treatments,
at the end of storage, fruit were maintained at 20 C
and 85% RH for 7 days to simulate a 1 week marketing
period (SMP).
Each treatment group was then divided into two sub-
groups of three boxes each (replications). Fruit boxes
from the first subgroup were used for physiological
disorders, while the fruit of the second subgroup were
sampled for acetaldehyde, ethanol and d-limonene lev-
els and sensory characteristics.
2.2. d-Limonene extraction and analysis from
grapefruit peel
Each d-limonene level determination was per-
formed using the grapefruit peel (flavedo) from six
fruit randomly taken from one box. At each sampling
time, three replicates were analyzed for each treat-
ment. Isolation and quantification of the d-limonene
was performed as described by Biolatto et al. (2001).
The methodology consists of chopping, into 0.5 cm
pieces, the same quantity of flavedo from each fruit.
Then the six fruit flavedo pieces were mixed, and
4 g of fresh weight (FW) of this pool was homoge-
nized 3 with n-pentane (1 g of FW/4 mL), adding
500 L of 400 g/mL solution of internal standard,
lauric acid methyl ester. The homogenates were de-
canted, and the organic layer was dried with anhy-
drous Na2 SO4 and then concentrated to 0.5 mL un-
der nitrogen at room temperature before being ana-
lyzed. The extracts were analyzed with a Shimadzu
series 14B gas-liquid chromatograph (GLC), equipped
with a flame ionization detector (FID) and a glass cap-
illary column (J&W Scientific, Folsom, CA) coated
with Carbowax 20 M (30 m 0.53 mm i.d., 1.0 m
film thickness). The carrier gas flow rate was 5 mL/min
N2 . The injection volume was 2 L, and the split
ratio was 10/1. The injector and detector tempera-
tures were 270 and 280 C, respectively. The follow-
ing column temperature-programming sequence was
169
used: an initial of 75 C was maintained for 5 min
before being increased to 200 C at 40 C/min, then
raised at 10 C/min to 240 C and held for 15 min.
Each peak area on the gas chromatogram was calcu-
lated automatically with a Class-VP Software. For cap-
illary gas-liquid chromatographymass spectrometry
(GLCMS) a Hewlett-Packard 5989A mass spectrom-
eter was used with a column similar to that used above.
Peak identification was confirmed by comparing the
retention times and mass spectra with those of an au-
thentic sample. Quantitative determinations were based
on the known amount of added standard.
2.3. Acetaldehyde and ethanol extraction and
analysis from fresh grapefruit juice
Each acetaldehyde and ethanol determination was
performed on fruit juice pools. Each pool was obtained
by squeezing six randomly chosen fruit from one box.
An aliquot from each pool was transferred to 27 mL
vials equipped with crimp-top caps with TFE/silicone
septa seals. At each sampling time, three replicates
were analyzed for each treatment.
Acetaldehyde and ethanol levels were determined
by headspace analysis on a Shimadzu series 14B
gas-liquid chromatograph (GLC), equipped with a
flame ionization detector (FID) and a model HSS-2B
headspace sampler. A Supelcowax glass capillary col-
umn (30 m 0.53 mm i.d. 0.50 m film thickness)
was used with a 25 kPa nitrogen head pressure. Juice
samples were equilibrated in the headspace sampler
for 45 min at 80 C prior to injection. The injection
volume was 0.4 mL, and the split ratio was 5/1. Oven
temperature programming was 40 C for 5 min and then
raised to 30 C/min at 180 C. The FID detector ampli-
fier range setting was for high sensitivity, and the tem-
perature was 250 C. Concentrations were calculated
with the use of regression equations, determined by in-
jecting five different concentrations of each component
added to a juice base to obtain a peak height calibration
curve. The juice base was prepared by reconstitution
to 11.8 Brix of concentrated juice (pumpout) from an
evaporator that contained no added flavor fractions.
2.4. Sensory evaluations
An eight member sensory panel was recruited from
the staff at Instituto Tecnologa de Alimentos of INTA.
170 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176
Sensory training was carried out prior to assessment
sessions. During these sessions, members of the panel
were trained using scales and fruit similar to those in-
volved in the assessment sessions. All of the panelists
had worked on profiling tests for more than 3 years.
This panel was used to determine the intensity of the
different tastes that form part of the typical fruit fla-
vor. Judges evaluated the samples using a descriptive
and comparative test and determined sweet, acid and
bitter taste and typical flavor intensity using a 10 cm
non-structured scale where the lower extreme means
extremely weak and the upper extreme means ex-
tremely strong. The tests were performed in a condi-
tioned assay room for sensory analysis under IRAM
20003 (ISO 8589:1988) and the judges were trained
under IRAM 20002 (ISO 6658:1985).
The responses of the panelists were recorded by
measuring the distance in cm (110) from the left side
of the scale for each attribute. The results are expressed
as the mean of these numeric values assigned by each
judge.
2.5. Reagents
Acetaldehyde, ethanol, d-limonene and lauric acid
methyl ester were purchased from SigmaAldrich of
Argentina SA.
2.6. Statistical analysis
The experimental procedure consisted of a com-
pletely randomized design, with three replicates per
treatment, with each box being a replicate. Data for
acetaldehyde, ethanol, and d-limonene level were an-
alyzed using a fixed effects model (Eq. (1)) by means
of the general linear model (PROC GLM) procedure of
SAS v.8 (SAS, 2000).
T-Tukey was applied as the test a posteriori with a
level of significance of 95% on analytical and sensory
characteristic to determine the effects of the different
treatments and processes.
3. Results
By the end of the simulated marketing period, the
conditions applied in the present research did not in-
duce the development of CI (data not shown).
The results of the present study indicate that after the
simulated marketing period, no significant differences
in acetaldehyde and ethanol concentrations were ob-
served between treatments that included cold quaran-
tine (T1 and T4) and those developed at the non-chilling
temperature (control, T3 and T6) (Tables 1 and 2).
However, after the simulated marketing period, fruit
subjected to temperature conditioning (T2) had signif-
icantly higher levels of acetaldehyde as compared to
fruit without temperature conditioning (T1) and con-
trol fruit (T3) (Table 1). The same behavior was ob-
served for ethanol levels when T5 was compared with
T4 and T6 (Table 2). Conversely, Tables 1 and 2 show
the T-Tukey test for the acetaldehyde and ethanol lev-
els found at the initial time and at the end of the mar-
keting period for each treatment. It can be observed
that the time involved in these treatments did not pro-
mote an accumulation in the acetaldehyde and ethanol
levels, except for T5. Although a statistical difference
in ethanol levels at the initial time and at the end of
the marketing period was found in T5, this difference
could be related to the variability in the initial amount
of ethanol in the fruit.
Results from flavedo of Rouge La Toma grape-
fruit, shown in Tables 1 and 2, revealed no significant
differences in the d-limonene level between treatments
either with or without temperature conditioning (T2
versus T1 and T5 versus T4) at the end of the mar-
keting period. It is important to note that fruit sub-
jected to control treatments, T3 and T6, had higher
levels of d-limonene than those of the other treatments
after the simulated marketing period. Moreover, d-
limonene levels increased significantly between the ini-
tial time and the end of the simulated marketing period
(Tables 1 and 2), for all treatments, with the highest and
most variables levels in those fruit stored under control
treatments (T3 and T6) (Fig. 1a and b).
After the simulated marketing period, in general,
no significant changes in either the sweet, the acid or
the bitter taste intensity and typical grapefruit flavor
were detected in any treatments assayed, except for T1
versus T3 (Table 3).
4. Discussion
Rouge La Toma is an important pigmented grape-
fruit variety, and is widely used in local and export
 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176 171

Table 1
Influence of treatments that included 42 days of postharvest handling on chemical parameters in Rouge La Toma grapefruita,b
Treatments Storage (days) Acetaldehyde (mg/100 mL) Ethanol (mg/100 mL) d-Limonene (mg/100 g)
T1 1 0.55 0.10(a) 18.38 4.14(a) 388.64 80.85 (bcd)
42 0.32 0.02 b (bc) 17.66 3.60 a (a) 473.29 15.21 ab (ab)
T2 1 0.46 0.05 (ab) 13.42 3.93 (a) 287.06 34.24 (d)
42 0.48 0.08 a (ab) 20.63 7.16 a (a) 423.52 52.10 b (abc)
T3 1 0.52 0.03 (a) 13.64 2.92 (a) 323.68 35.40 (cd)
42 0.25 0.02 b (c) 13.38 1.96 a (a) 525.28 21.52 a (a)

Source of variance d.f. Mean square

Analysis of variance
Treatment 2 1.15 3360.59 10555.60*
Time 1 11.25*** 1941.68 89340.46***
Treatment time 2 3.48** 2964.98 5151.81
Error 12 0.36 1819.65 2061.75
a Time 1: initial time 1 day after refrigerated transport and before treatment; Time 42: at the end of simulated marketing period. T1: non-
conditioned + quarantine at 2 C and 85% RH for 18 days + storage at 13 C and 85% RH for 17 days + marketing period at 20 C and 85% RH
for 7 days; T2: conditioned at 15 C and 85% RH for 7 days + quarantine at 2 C and 85% RH for at 20 C and 85% RH for 18 days + storage at
13 C and 85% RH for 17 days + marketing period at 20 C and 85% RH for 7 days; T3: storage at 13 C and 85% HR for 35 days + marketing
period at 20 C and 85% RH for 7 days.
b Means with letters without parenthesis indicate significant difference (P < 0.05) relate to comparisons of the effect of treatments. Means

with letters in parenthesis indicate significant difference (P < 0.05) relate to comparisons of the influence of the time in storage, within each
treatment.
P < 0.05.
P < 0.01.
P < 0.001.

commerce. Nevertheless, to our knowledge, these are
the first data providing evidence on its postharvest han-
dling and cold tolerance characteristics.
Postharvest peel pitting diminishes the external
quality and consequently the value of fruit for market
(Petracek et al., 1995, 1998). In general, peel disor-
ders in citrus fruit are induced by a wide array of biotic
and abiotic factors in the field or during postharvest
handling and storage. Factors that have been reported
to reduce the chilling injury incidence include: ethy-
lene degreening prior to cold storage (Grierson, 1974),
waxing (Davis and Harding, 1959) and application of
the fungicide thiabendazole (TBZ) (Schiffman-Nadel
et al., 1972, 1975; Petracek et al., 1998; Schirra et al.,
2000), delayed cooling (Hatton and Cubbedge, 1982a,
1982b, 1983; Chalutz et al., 1985; Porat et al., 2000) and
high humidity during storage (Pantastico et al., 1968;
Porat et al., 2004). In addition, the incidence of CI is
determined mainly by the storage temperature and by
the length of time during which the fruit is exposed to
the CI-inducing temperature (Schiffmann-Nadel et al.,
1971; Hatton and Cubbedge, 1982a, 1982b).
In the present research, we found that Rouge
La Toma grapefruit did not develop CI under the
posthavest handling practices applied, at the end of sim-
ulated marketing period. Our result differs with those
of Chalutz et al. (1985) who in comparable conditions
of cold quarantine (2.2 C for 16 days) showed that
Marsh grapefruit developed CI, mostly in the form
of slight peel pitting on 2.6% of the fruit. This may
be explained by the waxing with polyethylene-based
TBZ containing wax, ethylene degreening and condi-
tioning at 15 C for 7 days prior to cold quarantine.
Another reason for the lack of CI in Rouge La Toma
could be explained by a greater resistance of the culti-
var to CI development, under the cold quarantine con-
ditions assayed (2 C for 18 days). Furthermore it is
known that the susceptibility of grapefruit to CI when
exposed to temperatures below 10 C varies throughout
the harvesting season, i.e. grapefruit harvested during
midseason are generally more resistant to CI than fruit
harvested either earlier or later in the season (Harvey
and Rygg, 1936; Grierson, 1974; Purvis et al., 1979).
It is important to note that grapefruit employed in this
172 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176

Table 2
Influence of treatments that included 29 days of postharvest handling on chemical parameters in Rouge La Toma grapefruita,b
Treatments Storage (days) Acetaldehyde (mg/100 mL) Ethanol (mg/100 mL) d-Limonene (mg/100 g)
T4 1 0.56 0.04 (a) 19.10 2.82 (ab) 292.00 15.03 (c)
29 0.33 0.03 a (bc) 15.54 1.02 b (ab) 504.19 54.41 b (b)
T5 1 0.46 0.09 (ab) 12.79 5.05 (b) 353.42 29.13 (c)
29 0.37 0.13 a (bc) 23.79 1.88 a (a) 560.15 36.95 ab (ab)
T6 1 0.47 0.06 (ab) 19.15 5.82 (ab) 318.34 46.58 (c)
29 0.25 0.03 a (c) 11.72 2.38 b (b) 616.90 20.75 a (a)

Source of variance d.f. Mean square

Analysis of variance
Treatment 2 1.87* 1263.48 8395.53*
Time 1 7.53** 0.025 257391.17***
Treatment time 2 4.51** 14150.36** 3981.31
Error 12 0.47 1291.73 1333.34
a Time 1: initial time 1 day after refrigerated transport and before treatment; time 29: at the end of simulated marketing period; T4: non-
conditioned + quarantine at 2 C and 85% RH for 18 days + storage at 13 C and 85% RH for 4 days + marketing period at 20 C and 85% RH
for 7 days; T5: conditioned at 15CircC and 85% RH for 7 days + quarantine at 2CircC and 85% RH for 18 days + storage at 13 C and 85% RH
for 4 days + marketing period at 20 C and 85% RH for 7 days; T6: storage at 13 C and 85% HR for 22 days + marketing period at 20 C and
85% RH for 7 days.
b Means with letters without parenthesis indicate significant difference (P < 0.05) relate to comparisons of the effect of treatments. Means

with letters in parenthesis indicate significant difference (P < 0.05) relate to comparisons of the influence of the time in storage, within each
treatment.
P < 0.05.
P < 0.01.
P < 0.001.

study were harvested during midseason (June) when opment of postharvest pitting was caused by the re-
the mean low temperature in the field drops below duction of fruit internal O2 and increase of CO2 levels.
10 C. Low O2 may stimulate anaerobic respiration (Biale and
In addition, Petracek et al. (1995, 1998), have shown Young, 1962). Compounds produced by anaerobiosis
that postharvest pitting of different species and culti- such as alcohols and aldehydes may then stimulate pit-
vars is stimulated by high temperature storage of waxed ting (Petracek et al., 1998). We did not observe an in-
fruit. Results published by Petracek et al. (1998), re- crease in acetaldehyde and ethanol levels, or the devel-
lated peel pitting in grapefruit with coating formula- opment of pitting of individual fruit. In accordance with
tion and gas permeability, suggesting that the devel- Petracek et al. (1998), these results can be explained
in part by the application of polyethylene-based wax,
which is more permeable compared to shellac-based
Table 3
Sensory characteristic of treatments that included 42 days of posthar-
wax (Petracek et al., 1998).
vest handling in Rouge La Toma grapefruita,b In contrast, Davis (1971), Lyons (1973), Davis
Treatment Sweet Acid Bitter Typical grapefruit
et al. (1974), Eaks (1980) and Pantastico et al. (1968),
taste taste taste flavor have reported acetaldehyde and ethanol accumulation
in fruit during chilling. However, in the present research
T1 3.16 a 2.12 b 5.17 a 4.99 a
T2 3.07 a 2.75 ab 4.39 a 4.32 a the treatments that included quarantine did not show
T3 2.15 a 3.29 a 4.92 a 4.06 a acetaldehyde and ethanol accumulation. Although dif-
a Samples presented at room temperature using a scale of 1: ex-
ferent grapefruit cultivar and conditions were studied,
tremely weak; 9: extremely strong.
these results are in agreement with those obtained by
b Means with letters indicate significant difference (P < 0.05) re- Murata and Ku (1966), Eaks (1980), Schirra (1992),
late to comparisons of the effect of treatments. Arras and Usai (1993), Schirra et al. (1998), Kyung
 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176
Fig. 1. (a) d-Limonene levels of Rouge La Toma grapefruit from
different treatments after 1 day refrigerated transport; 18 days of
quarantine at 2 C; 18 days of quarantine at 2 C + 17 days of stor-
age at 13 C; 18 days of quarantine at 2 C + 17 days of storage
at 13 C + 7 days of marketing period at 20 C. Vertical bars in-
dicate S.E. (n = 3). (
) Storage at 13 C and 85% HR for 35
days + marketing period at 20 C and 85% RH for 7 days; ()
conditioned at 15 C and 85% RH for 7 days + quarantine at 2 C
and 85% RH for 18 days + storage at 13 C and 85% RH for 17
days + marketing period at 20 C and 85% RH for 7 days; () non-
conditioned + quarantine at 2 C and 85% RH for 18 days + storage at
13 C and 85% RH for 17 days + marketing period at 20 C and 85%
RH for 7 days. (b) d-Limonene levels of Rouge La Toma grapefruit
from different treatments after 1 day refrigerated transport; 18 days
of quarantine at 2 C; 18 days of quarantine at 2 C + 4 days of storage
at 13 C; 18 days of quarantine at 2 C + 4 days of storage at 13 C + 7
days of marketing period at 20 C. Vertical bars indicate S.E. (n = 3).
(
) Storage at 13 C and 85% HR for 22 days + marketing period at
20 C and 85% RH for 7 days; () conditioned at 15 C and 85% RH
for 7 days + quarantine at 2 C and 85% RH for 18 days + storage at
13 C and 85% RH for 4 days + marketing period at 20 C and 85%
RH for 7 days; () non-conditioned + quarantine at 2 C and 85% RH
for 18 days + storage at 13 C and 85% RH for 4 days + marketing
period at 20 C and 85% RH for 7 days.
173
and Yong (1998) and Schirra and Cohen (1999), who
have shown, in different species and cultivars of fruit,
that ethanol and acetaldehyde can accumulate as a re-
sult of storage conditions inducing chilling injury.
Among heat treatments that enhance cold toler-
ance and can be commercially used to reduce CI dam-
age (Hatton, 1990; Wang, 1993, 1994; Lurie, 1998a,
1998b), in the present study we selected temperature
conditioning. This treatment was chosen due to the
discoveries of Grierson (1974), Hatton and Cubbedge
(1982a, 1982b, 1983) and McDonald (1986), who
found that temperature conditioning of grapefruit prior
to low temperature treatment gave some success. How-
ever, in this study the effectiveness of temperature con-
ditioning was not observed because Rouge La Toma
grapefruit did not show development of CI under the
conditions assayed.
The occurrence of off-flavours in stored citrus fruit
has been related to high accumulation of volatiles such
as acetaldehyde and ethanol in the juice (Cohen et al.,
1990; Ke and Kader, 1990). Storage temperature and
storage period at which fruit are held affect both fruit
respiratory demand and coating permeability to gases.
These factors contribute to alter the atmosphere and
thus may affect volatile compounds (Baldwin et al.,
1995), and that, in turn, impact on flavor. Therefore,
the highest acetaldehyde and ethanol levels in T2 and
T5, at the end of the marketing period, might be a result
of temperature conditioning (15 C for 7 days). Porat
et al. (2000) showed that extended heat treatments, like
temperature conditioning (21 C for 3 days and 16 C
for 7 days) affected internal quality parameters in Star
Ruby grapefruit.
It is important to stress that, in general, the sensory
characteristic were not affected by the postharvest treat-
ments applied, excepting for the significant differences
observed between T1 and T3 in acid taste. This result
could be due to a difference in the acid content of the
juice. Nevertheless, it should be noted that the amounts
of acetaldehyde and ethanol detected for all treatments
assayed are comparable with those found by Nisperos-
Carriedo and Shaw (1990) in fresh orange juice, and
by Cadwallader and Xu (1994) in fresh grapefruit
juice.
Low temperatures, used for pest disinfestations,
are often very injurious to citrus and can cause
severe injury to the rind, disrupting the oil glands and
promoting the d-limonene emanation. According to
174 A. Biolatto et al. / Postharvest Biology and Technology 35 (2005) 167176
Obenland et al. (1996, 1997), postharvest heat treat-
ments, used to induce cold tolerance and to reduce the
development of CI symptoms, decrease the release of
d-limonene. They observed that lemons after 3 weeks
of cold storage at 1 C at which moderate to severe CI
was apparent, showed some increase in d-limonene
levels. However, in our study the d-limonene level
in flavedo tissue from Rouge La Toma grapefruit
did not show significant differences between fruit
subjected to treatments that included cold quarantine
treatment with or without temperature conditioning
(T2 and T5, T1 and T4, respectively), at the end of the
marketing period. Accordingly, no visible symptoms
of CI were observed in both cases.
It is known that under refrigerated conditions it
is possible to retard biochemical and physiological
changes associated with senescence (Couey, 1982;
Wang, 1994). In the current research, fruit stored at
optimum temperature storage (control treatments, T3
and T6) revealed higher d-limonene levels compared
to those under refrigerated conditions (T1, T2, and T4,
T5), at the end of the marketing period. Moreover,
a significant increase in d-limonene levels through-
out storage time (Fig. 1a and b) was observed in fruit
stored under control treatments. Although ethylene is
considered a growth regulator capable of accelerat-
ing the maturation-senescence processes of citrus fruit
(Ben-Yehoshua et al., 1995; Leievre et al., 1997;
Salveit, 1999), the results observed in control treat-
ments compared to the other treatments suggest that
the biosynthesis of d-limonene might be regulated by
storage conditions. Thus, d-limonene may be a can-
didate as a senescence indicator or predictor, because
higher temperature storage and/or longer storage time
promotes senescence. The same conclusion was in-
ferred by Sun and Petracek (1999) using Marsh white
grapefruit, when their results showed the opposite be-
havior in the d-limonene level (decrease with higher
temperature and longer time). Therefore, further spe-
cific studies are required to elucidate this point.
5. Conclusions
Cold treatments are an effective quarantine method
to kill eggs and larvae in grapefruit and are commer-
cially applied by various citrus-importing countries.
While the applications of cold quarantine treatments
may be problematic for disinfecting fruit cultivars sus-
ceptible to CI, the application of heat treatment before
cold disinfestation may overcome the risk of potential
fruit damage from CI after quarantine treatment. Based
on results, it can be concluded that cold quarantine
treatment and temperature conditioning have an im-
portant commercial application for Rouge La Toma
grapefruit (Citrus paradisi Macf.) without adversely
affecting its quality.
Acknowledgements
The study described herein is a part of a major
project financed by the Phytosanitary Association of
Argentina Northwest (AFINOA), that includes produc-
ers from the States of Tucuman, Salta, Jujuy and Cata-
marca of Argentina. AFINOA financed the develop-
ment of a Sanitary Barriers System in order to secure
to the admission of Argentina Northwest region into
the Citric Bank, in addition to achieving an integral
system for the prevention, control and eradication all
plagues and diseases that could affect the free com-
mercialization of vegetable products of this region. We
thank Taryn Kinahan and Betina Biolatto for their help-
ful advice and discussions. We thank Gabriela Grigioni
from Instituto Tecnologa de Alimentos-INTA, Caste-
lar, for her helpful advice.
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