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.