Fermentation dynamics was assessed gravimetrically measuring
the weight loss of fermentation medium due to CO2 release as
described earlier [12]. The samples for physico-chemical analysis
(ca. 20 mL aliquots) were taken every 24 h of fermentation. The
non dissolved solids concentration was determined as follows: ca.
10 g of fermentation media (with an accuracy of 0.001 g) were filtered
through pre-dried (105 C, 4 h) and pre-weighted filter paper,
the residual dissolved solids were washed with ca. 250 mL of distilled
water. The filters with remaining non-dissolved solids were
dried in an oven at 60 C for 12 h next at 105 C for 4 h. Afterwards
the filters were cooled to room temperature in a desiccator and
weighted with an accuracy of 0.001 g. The difference in the weight
of the filter with and without of non-dissolved particles gave the
concentration of them in the fermentation medium which was
expressed as grams of non-dissolved solids per kilogram. For HPLC,
reducing sugar and dissolved solids analysis the samples of fermentation
media were centrifuged at 6000 rpm (5243g) at 4 C using
MPW-351R centrifuge (MPW Med. Instruments, Poland) and clear
supernatants were taken for analyses. The reducing sugars concentration
(as glucose) was determined using the DNS method [26].
The dissolved solids content (apparent extract) was determined
by the density measurement of the clarified fermentation medium
as described previously [29].