Sample preparation and storage
Single strength orange juice (11.2 Brix, pH 3.7, titratable acidity
0.8%) was prepared by reconstituting frozen Brazilian orange
concentrate (65 Brix) with water (1:5, w/w). The juice was immediately
subjected to pasteurisation (92 C, 30 s) followed by filling
at 85 C into monolayer PET bottles (500 mL), cap twisting and
inversion of the bottles to disinfect the bottle tops. The bottles
were then cooled to ambient temperature by submerging them
into a circulating and chlorinated water tank. After processing,
the juices were stored at 20 and 28 C for 32 weeks, at 35 C for
12 weeks and at 42 C for 8 weeks, in dark incubators (IPP500;
Memmert, Schwabach, Germany).
At each sampling time (Fig. 1), three bottles were taken from
storage, mixed and divided uniformly into smaller tubes
(+30 mL). These tubes were immediately frozen in liquid nitrogen
and stored at 80 C until use. At the time of analysis, tubes were
thawed in a circulating water bath at 25 C.
2.2. C