Sample origin and characteristics
Analyses were performed on two lots of pears (Pyrus
communis L. var. S. Bartolomeu), corresponding to two
drying seasons, 2008 and 2009. Each lot was composed by
fresh pear fruits and traditionally dried fruits (direct
exposure to solar radiation) (Traditional). The samples
from 2009 were also processed using three different
methodologies: in a large glass greenhouse with air convection
(GH1), in a small greenhouse with natural convection
(GH2) and using a hot air tunnel in the absence of
light (HAT). All fruits were harvested in an orchard located
at the village of Ervedal da Beira (Oliveira do Hospital,
Portugal) at the commercial maturity stage (Fig. 1a). On
average, the fresh fruits had a mass of 64 g, with a diameter
of 4.3 cm and height of 6.5 cm. The moisture content was
78%.
For fresh pear analysis, some fruits were frozen just
after harvesting. The remaining fruits were processed
according to the different methodologies under study.
According to the traditional sun-drying process, the
fresh fruits were peeled and allowed to dry in an open
space with sunshine incidence, as described by Ferreira
et al. [5] (Fig. 1b). After being peeled, fresh pears were
also allowed to dry in two greenhouses that differed in their
size, structure and location. GH1 was provided with air
convection, with a flux of 900 m3 h-1, it was located at
ground level and, in comparison to GH2, had the largest
cargo capacity. Its area was 6.3 m2 (3.19 m long by 1.93 m
wide), and the height in the sides was 1.24 m, whereas in
the middle was 1.97 m. The structure was aluminium with
horticulture glass and there were two roof windows for air
extraction. The drying time for GH1 was approximately
7 days (Fig. 1c). The GH2 greenhouse consisted of insulated
glass panels and had the form of a box (1.20 m height
by 1.20 m long and 1.00 m wide). Inside, it was composed
by a U-shaped structure, parallel to the floor, where pears
were let to dry. GH2 was located on a roof of a building
and contained a step structure coated with reflective film,
where pears were allowed to dry. This interior structure
allowed a greater incidence of the light and consequently
promoted the drying process by natural air convection, with
the air entering in the lower level (first step) and leaving in
the higher level, by means of a girandole. The drying time
for GH2 was approximately 5 days (Fig. 1d), 2 days less
that the drying time of GH1.
For hot air tunnel drying (HAT), after peeled, pears were
let to dry at a constant temperature of 40 C and an air flow
of 1.2 m s-1 for about 7 days (Fig. 1e). The drying tunnel
was 2 m long, and the air convection was promoted by a
ventilator placed at the entrance of the tunnel. In this
structure, pears were not exposed to light, although the air
was heated by a solar collector, and therefore the sun was
also the main source of energy.
Independently of the type of processing used, the processed
fruits reached a mass of 10–14 g with a moisture
content of 20% and had a maximum width of 2.5–3.4 cm,
height of 3.9–4.7 cm and thickness of 1.2–1.5 cm. The
sensory descriptive profile of the traditional product, when
evaluated by a sensory panel test, is the following: brown–
red uniform colour (Fig. 1b), sweet and slight acid, with a
not too hard–not too soft texture and presenting elasticity.
Concerning GH1, it was considered very similar to the
traditional product, with the same rate of global appreciation,
but with lower colour uniformity (Fig. 1c), sweetness
and elasticity. GH2 was similar to GH1, although obtaining
a lower global appreciation, possibly due to an even lower
acidity (Fig. 1d). HAT presented the sensory profile close
to the traditional product concerning colour uniformity,
sweetness and toughness but it was not appreciated due to
the colour tonality (yellow-orange) (Fig. 1e) and slight less
acidity. This product, although having the accurate organoleptic
characteristics except the colour, was rejected by
the panellists due to the inaccurate colour when compared
with the traditional product.
Before analysis, the pulp of the fresh and dried pears
was collected, ground and freeze-dried.
Amino acids analysis
The methodology for acid hydrolysis of protein material
was adapted from Zumwalt et al. [22]. To a test tube with a
screw cap with PTFE coating was rigorously weighed
10 mg of freeze-dried material. To each sample, 2 mL
hydrochloric acid (HCl) 6 M was added; the content of the
tubes was frozen in liquid nitrogen, evacuated during 1 min
with a vacuum pump and refilled with nitrogen. After
thawing, the suspension was sonicated for 5 min in an
ultrasound bath at room temperature. The procedure of
freezing–vacuum–sonication was repeated twice. The
hydrolysis took place during 24 h at 110 C using a heating
block. After cooling to room temperature, 500 lL of the
Fig. 1 The S. Bartolomeu pear.
a Fresh pears and pears dried by
different technologies:
b Traditional, c GH1, d GH2
and e HAT
Eur Food Res Technol (2011) 233:637–646 639
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internal standard solution (norleucine 5.0 mM in HCl
0.1 M) was added and the tubes content was evaporated to
dryness under vacuum in a centrifugal evaporator. The
resulting material was dissolved in 1 mL HCl 0.1 M and
filtered with 0.45 lm filters. For the analysis of free amino
acids, 10 mg of each sample was suspended in 2 mL of a
solution of HCl 0.1 M and spiked with 500 lL of the
internal standard solution. The suspension was left stirring
for several hours and then was filtered with 0.45 lm filters.
The solutions containing the released amino acids were
dried under vacuum using a centrifugal evaporator.
The derivatization of amino acids for GC analysis was
performed according to the methodology described by
MacKenzie et al. [23]. The resultant solid residue was
dissolved in 200 lL of a solution of 3 M HCl in isobutanol.
This solution was prepared by adding 270 lL of acetyl
chloride per mL of dry isobutanol; the isobutanol was dried
with calcium hydride, distilled and stored with molecular
sieves 4 A ° . The mixture was heated to 120 C for 10 min
and, after shaking in a vortex, was heated for further
30 min. After cooling to ambient temperature, the excess of
reagent was evaporated under vacuum using a centrifugal
evaporator. Then, 200 lL of a solution of 0.2 mg mL-1
BHT prepared in ethyl acetate was added and the solvent
was removed under vacuum in a centrifugal evaporator.
Afterwards, 100 lL of heptafluorobutyric anhydride was
added and the mixture was heated during 10 min at 150 C.
After cooling to room temperature, the excess of solvent
was removed under vacuum and the material obtained was
dissolved in 50 lL of ethyl acetate and analysed immediately
or frozen at -20 C until analysis.
Separation of amino acids was achieved by gas chromatography,
carried out in a PerkinElmer Clarus 400
instrument (PerkinElmer, Massachusetts, USA) equipped
with a flame ionisation detector (FID). The injector was
kept at 250 C and the detector at 260 C. Hydrogen was
used as carrier gas. A DB-1 (30 m, 0.25 mm i.d. and
0.15 lm thickness) fused-silica capillary column (J & W
Scientific) was used with the following temperature programme:
1 min hold at 70 C, increase to 170 C at 2.0 C/
min and then to 250 C (5 min hold) at 16 C min-1. The
compounds were identified by their retention times and
chromatographic comparison with authentic standards.
Quantification was based on the internal standard method
using L-norleucine, and the calibration curves were built for
18 amino acids. For asparagine (Asn) and aspartic acid
(Asp), as well as for glutamine (Gln) and glutamic acid
(Glu), the methodology does not allow the distinction
between the amide and carboxylic acid functions. As such,
those amino acids were quantified together as Asx and Glx,
respectively. Also, the methodology used did not allow the
detection of His. The limit of quantification of analysed
amino acids was determined to be ten times the value of the
residual signal peaks.