c-PGA production were obtained by calculation of regression equation.
The optimum levels of the variables in their actual values
were as follows: glycerol (x1) = 17.6 g l1
, glutamic acid (x4) =
59.6 g l1
, yeast extract (x7) = 2.7 g l1
, and K2HPO4 (x11) =
2.3 g l1
. The predicted response as c-PGA production under the
optimized condition was 48.5 g l1
.
3.4. Validation of the statistically optimized values
For verification of the adequacy of the quadratic model for predicting
c-PGA production, the statistically optimized levels were
validated by conducting fermentor-scale experiment under the
optimized conditions. As shown in Fig. 4, c-PGA production was
peaked at 24 h of fermentation. The maximum c-PGA concentration
(48.7 g l1
) obtained from the validation experiment gives
only 0.4% differences from the statistically predicted response
(48.5 g l1
). This result indicates that the regression model developed
in this study should be in good agreement between the actual
and predicted responses.
On the other hand, the maximum dry cell weight (0.88 g l1
),
viscosity (645 cP), and molecular weight (62.0 kDa) were obtained
at 24, 30, and 36 h of fermentation, respectively. However, molecular
weight distribution of c-PGA decreased sharply after the value
reached to its maximum. Troy (1973) and Birrer et al. (1994) previously
observed a decrease in the medium viscosity when B.
licheniformis was cultivated, and Goto and Kunioka (1992) reported
degradation of c-PGA from cell-free culture broth, thereby suggesting
the existence of c-PGA depolymerase, which could be responsible
for the breakdown of c-PGA.
4. Conclusions
The statistical approaches proved to be efficient for the optimization
of c-PGA production by B. subtilis RKY3. A c-PGA production
in fermentor-scale experiment was quite similar to the
predicted response, and the maximum c-PGA concentration
was only 0.4% different from the statistically predicted response.
During validation study, a decrease in molecular weight of the cPGA
was observed after the latter half of fermentation. The characteristics
of c-PGA depolymerase from B. subtilis RKY3, which
could be responsible for decreases in molecular weight of cPGA
would be further examined to improve the molecular
weight of c-PGA.
Refer