The time course study indicated that PHB is a growthassociated
product in A. fertilissima, and its accumulation
increased significantly when the culture reached the
exponential phase until the stationary phase. The maximum
value, 6.4% (dcw), was achieved on the 14th day of
cultivation. After 20 days, the decrease in the PHB content
might be due to the mobilization of PHB for cellular metabolism to regain the full capacity of photoautotrophic
growth as a source of carbon storage material (Stal 1992).
A pH of 8.5 and a temperature range of 28–32°C were
found optimum for PHB accumulation.
A. fertilissima is quite flexible in utilizing carbon
sources. Intracellular accumulation of PHB, however,
appeared to be carbon source-specific (Table 2), and
maximum rise, i.e. an ∼6-fold increase in PHB content
(up to 34% dcw), was observed in cultures supplemented
with 3 g L−1 citrate. As observed by Chen et al. (2010), the
addition of citrate decreased the activity of phosphofructokinase
by chelating Mg2+, which blocks the glycolytic
pathway, especially the reaction from fructose-6-phosphate
to fructose-1,6-bisphosphate, thus resulting into the elevated
pool of fructose-6-phosphate. An ultimate rise in glucose-6-
phosphate pool was observed by the conversion of fructose-6-
phosphate to glucose-6-phosphate by isomerase, which led to
the increased activity of 6-phosphoglucose dehydrogenase,
the first enzyme of the pentose phosphate pathway (PPP), thus
facilitating the production of a more reduced cofactor,
NADPH via PPP. NADPH is reported to be a prerequisite
for the activity of the enzyme acetoacetyl-CoA reductase for
conversion of acetoacetyl-CoA to β-hydroxybutyryl-CoA.
Therefore, the increased PHB accumulation following citrate
supplementation could be due to the enhanced availability of
NADPH.
Citrate supplementation was found to inhibit biomass
concentration significantly (Table 2). Contrary to this,
fructose supplementation was stimulatory for biomass
production. By increasing the concentration up to
10 g L−1, growth and PHB production increased up to
2.39 and 0.38 g L−1, respectively, against 0.50 and
0.03 g L−1 in the control culture. A further increase in
fructose concentration inhibited the growth of A. fertilissima.
This might be due to the increased osmotic pressure
at high fructose concentration and the imbalance between
glycolysis and metabolic oxidation in cyanobacterial cells
(Tabandeh and Vasheghani-Farahani 2003). The stimulatory
effect of acetate on PHB accumulation, i.e., 27.1% (dcw)
against 6.4% control, could be due to a direct utilization of
acetate to increase the intracellular acetyl-CoA pool at the
expense of free CoA by means of the usual pathway
operating in cyanobacteria (Gibson 1981; Panda and
Mallick 2007). Glucose utilization in cyanobacteria, however,
occurs via PPP. Thus, the positive effect of glucose on
PHB production could be attributable to the increased
supply of the reduced cofactor, NADPH (Lee et al. 1995).
Similar explanation could also be valid for the increased
PHB contents in fructose-, sucrose-, and maltosesupplemented
cultures. Interestingly, co-feeding A. fertilissima
cultures with 3 g L−1 citrate and 3 g L−1 acetate
stimulated PHB accumulation up to 65.9% (dcw). This
could be explained by the combined effects of the precursor
available in plenty, i.e., acetate, and the cofactor NADPH
The time course study indicated that PHB is a growthassociatedproduct in A. fertilissima, and its accumulationincreased significantly when the culture reached theexponential phase until the stationary phase. The maximumvalue, 6.4% (dcw), was achieved on the 14th day ofcultivation. After 20 days, the decrease in the PHB contentmight be due to the mobilization of PHB for cellular metabolism to regain the full capacity of photoautotrophicgrowth as a source of carbon storage material (Stal 1992).A pH of 8.5 and a temperature range of 28–32°C werefound optimum for PHB accumulation.A. fertilissima is quite flexible in utilizing carbonsources. Intracellular accumulation of PHB, however,appeared to be carbon source-specific (Table 2), andmaximum rise, i.e. an ∼6-fold increase in PHB content(up to 34% dcw), was observed in cultures supplementedwith 3 g L−1 citrate. As observed by Chen et al. (2010), theaddition of citrate decreased the activity of phosphofructokinaseby chelating Mg2+, which blocks the glycolyticpathway, especially the reaction from fructose-6-phosphateto fructose-1,6-bisphosphate, thus resulting into the elevatedpool of fructose-6-phosphate. An ultimate rise in glucose-6-phosphate pool was observed by the conversion of fructose-6-phosphate to glucose-6-phosphate by isomerase, which led tothe increased activity of 6-phosphoglucose dehydrogenase,the first enzyme of the pentose phosphate pathway (PPP), thusfacilitating the production of a more reduced cofactor,NADPH via PPP. NADPH is reported to be a prerequisitefor the activity of the enzyme acetoacetyl-CoA reductase forconversion of acetoacetyl-CoA to β-hydroxybutyryl-CoA.Therefore, the increased PHB accumulation following citratesupplementation could be due to the enhanced availability ofNADPH.Citrate supplementation was found to inhibit biomassconcentration significantly (Table 2). Contrary to this,fructose supplementation was stimulatory for biomassproduction. By increasing the concentration up to10 g L−1, growth and PHB production increased up to2.39 and 0.38 g L−1, respectively, against 0.50 and0.03 g L−1 in the control culture. A further increase infructose concentration inhibited the growth of A. fertilissima.This might be due to the increased osmotic pressureat high fructose concentration and the imbalance betweenglycolysis and metabolic oxidation in cyanobacterial cells(Tabandeh and Vasheghani-Farahani 2003). The stimulatoryeffect of acetate on PHB accumulation, i.e., 27.1% (dcw)against 6.4% control, could be due to a direct utilization ofacetate to increase the intracellular acetyl-CoA pool at theexpense of free CoA by means of the usual pathwayoperating in cyanobacteria (Gibson 1981; Panda andMallick 2007). Glucose utilization in cyanobacteria, however,occurs via PPP. Thus, the positive effect of glucose onPHB production could be attributable to the increasedsupply of the reduced cofactor, NADPH (Lee et al. 1995).Similar explanation could also be valid for the increasedPHB contents in fructose-, sucrose-, and maltosesupplementedcultures. Interestingly, co-feeding A. fertilissimacultures with 3 g L−1 citrate and 3 g L−1 acetatestimulated PHB accumulation up to 65.9% (dcw). Thiscould be explained by the combined effects of the precursoravailable in plenty, i.e., acetate, and the cofactor NADPH
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