3. Results and discussion
In the present study, the effect of NaCl addition on the growth
and PHA production by C. necator was evaluated.
3.1. Effect of NaCl at different concentrations on the growth of
C. necator
PHA yield not only depends on the amount of PHA stored within
the bacterial cell but it is also attributed to the number of cells accumulating
PHA. Therefore, it was essential to achieve high cell numbers
before the bacteria started accumulating PHA. The initial
increase in OD (Tables 1a–1e) and CDW was mainly due to cell
growth (Fig. 1). It was evident from CDW measurements, that
bacteria were able to have a reasonable growth of above 5 g/l CDW
by 24 h in all the cases except for salt concentrations of 12 g/l and
15 g/l NaCl, which resulted in much lower CDW of 1.9 g/l and 2 g/l
at 24 h, respectively (Fig. 1). Also, theCDWfor the fermentation with
9 g/l NaCl was 6% higher at 24 h than for the control (no addition of
NaCl) and 12% and 10% higher than 6.5 g/l and 3.5 g/l NaCl fermentations,
respectively. For fermentations with NaCl concentrations
of 12 g/l and 15 g/l the CDW was 69–70% lower than compared to
the control at 24 h, indicating an inhibitory effect at higher salt concentrations
demonstrated by the lower cell growth of C. necator cells.
However, in terms of CDW, the control demonstrated higherCDWat
24 than 3.5 and 6.5 g/l NaCl fermentations.
Fig. 2 shows the stirring speed profile for all the experiments
and indicated the bacterial behaviour and oxygen demands at
increased NaCl concentrations. DO was controlled at 40% throughout
each experiment by maintaining the stirring speed to avoid any
oxygen limitation for bacterial growth. Fig. 2 shows the stirring
speed in the fermenter and it was observed that at the beginning
of the control fermentation, the stirring speed was 200 rpm as
compared to 300 rpm for all other fermentations and 600 rpm for
the 15 g/l NaCl fermentation. It is a known phenomenon that
increasing the salinity of any media decreases its oxygen solubility.
Therefore, at fermentations with increased salt concentrations, the
stirring speed also showed to have to increase to maintain the DO
concentration as close to 40% as possible to cope with low oxygen
solubility at higher salt concentrations. The DO readings (data not
shown) back up this phenomenon that had not been investigated
previously. DO concentration decreased to approximately 28% for
the 15 g/l NaCl fermentation (data not shown), whilst the stirring
speed was at its maximum at 600 rpm for the first 10 h period
(Fig. 2), indicating that high salt concentrations seemed unfavourable
for growth of C. necator due to oxygen unavailability. The 9 g/l
NaCl fermentation showed a peak in stirring speed between 20 and
30 h with its maximum at 24–26 h, which was also the time of
maximum PHA accumulation for that fermentation (Fig. 2;
Table 1d).
It was difficult to compare the growth rate in the salt media
with measurements of OD and CDW only, as these measurements
not only depend on cell growth but also on PHA accumulation.