2.4. Fermentation conditions
The fermentations were carried out in glass jars with a working
volume of 2 L (including the inoculum). The substrate comprised of
hydrolyzed henequen leaf juice diluted to 6, 8 and 10 degrees Brix
(Bx) with distilled water. The Brix scale is used in the sugar and
winemaking industry to indicate the percent of sugar by weight
in a solution, in our case the soluble solids present in the henequen
juice were not only sugars but also other compounds not yet identified
(organic acids, saponins among others). It was necessary to
dilute the juice as the concentration of soluble solids varies between
6 and 12 Bx approximately according to the time of year.
(In preliminary experiments 12 Bx was found to have the maximum
of soluble solids obtained in henequen leaf juice.) In order
to establish a constant initial concentration of soluble solids, the
three dilutions were complemented with industrial molasses until
12 Bx was reached in all cases. Ammonium sulphate was added as
nitrogen source at a concentration of 1.5 g L1. The pH was not adjusted
but was measured as henequen juice always has a pH value
of 4.7 ± 0.2. After inoculation, the fermentations were incubated at
35 ± 2 C in darkness, without stirring, and samples were collected
every 12 h to monitor the evolution of the fermentation process.
The soluble solids concentration of the samples was measured
every 12 h and the reducing sugars were analyzed on initiation
and completion of fermentation. Previous experiments confirmed
that the concentration of soluble solids stabilizes at its lowest
point between 46 and 48 h, and foaming also ceases at this point.
Our fermentations were carried out over a period of 48 h.