Part B | 12.5
354 Part B Tools and Methods in Marine Biotechnology
The nonheterocystous marine cyanobacterium
Phormidium valderianum (which is known to produce
hydrogen under argon atmosphere) was immobilized in
combination with two other nonmarine cells (viz., the
extreme halophile Halobacterium halobium and salt
tolerant Escherichia coli) for stable, long-term biohydrogen photo-evolution under an alternating light/dark
illumination cycle (6 h on/18 h off) in a PBBR by
Patel and Madamwar [12.44], as well as by Bagai and
Madamwar [12.43].
The former workers used Ca-alginate or agar-agar
as the immobilization matrix for cell entrapment in
a 50 mL working volume PBBR (basically a packed
column) continuously fed with N-free growth medium –
comparatively better results were obtained with alginate. The problem of cell and enzyme leakage from
alginate beads (leading to loss of H 2 production)
was overcome by using glutaraldehyde as a crosslinking agent. H 2 photoevolution rate increased with
medium feed rate up to a maximum, and then declined
markedly. Stable, long-term H 2 production for 11 d was
demonstrated.
Using the same combination of microorganisms,
in a similar PBBR, but with a different immobilization matrix, viz., PVA cross-linked with glutaraldehyde,
Bagai and Madamwar [12.43] demonstrated continued
H 2 production for over 60 d. In both these studies for
photohydrogen production, compared with a free-cell
system, the immobilized system was much more productive, stable, and long lasting.
Sabu et al. [12.46] and Kumar and Chandrasekaran
[12.45] all reported the production of the therapeutically and industrially important enzyme L-glutaminase
by Ca-alginate immobilized marine microorganisms in
glass-column PBBRs, but whereas the former employed
the fungus Beauveria bassiana the latter used the bacterium Pseudomonas for that purpose. Although the
PBBRs used in these studies differed in dimensions
(i. e., column height and diameter) with the one used
by Kumar and Chandrasekaran [12.45] being larger
(height 45 cm and internal diameter 3:6 cm), the basic principle of reactor construction was the same –
immobilized viable cell beads were aseptically packed
into the column with a perforated Teflon disk placed at
the top of the packing to prevent bed expansion during
operation.
Sabu et al. [12.46] pumped the medium from the
bottom of the packed column and collected the effluent from the top. This was, however, reversed by Kumar
and Chandrasekaran [12.45]. The inlet and outlet flow
rates were kept equal to maintain a constant liquid level
at just above the bed level. Now B. bassiana being
aerobic with strong aeration dependence, air was introduced through a sparger at the bottom of the packed
column after passing through a bacterial filter. Remarkably, Sabu et al. [12.46], noted that the maximum
enzyme production rate was observed in the absence
of forced aeration, and any increase in aeration rate
caused a decrease in enzyme productivity. In order to
explain this finding, the authors posited that with external aeration, air bubbles fill the void spaces of the
packing beads, thus reducing the contact between the
cell-laden beads and the liquid medium. They also
found that enzyme productivity increased with increasing bed height/decreasing flow rate, leading them to
conclude that increase in residence time either with increase in bed height or with decrease in flow rate, in turn
increases the contact time between the immobilized
beads and the medium, thereby causing an increase in
productivity.
Kumar and Chandrasekaran [12.45] reported that
enzyme production declined with an increase in dilution
rate, irrespective of the substrate concentration used.
Again, at all dilution rates examined, enzyme yield increased with increase in substrate concentration. It was
noted that their continuous-flow PBBR system could
be operated for 120 h without any decline in enzyme
productivity.
Kumar et al. [12.47] developed a nitrifying PBBR
immobilized with an indigenous NBC (nitrifying bacterial consortium) (comprising predominantly marine
species, e.g., Nitrosospira) for rapid nitrification in
brackish water and marine hatchery systems. They
examined nitrification performances of the PBBR integrated into a recirculating aquaculture system (RAS)
viz., a Penaeus monodon (a marine crustacean –
shrimp) recirculating maturation system. The RAS
(Fig. 12.22) comprises a shrimp maturation tank (MT),
an overhead tank (OHT), Six PBBRs (designated as R1
through R6) and a collection tank (CT). Fluid from the
MT was pumped into the OHT from where it flowed
through the reactors (R1–R6) connected serially by
gravitation and finally collected in the CT, from where
the treated water was recirculated back to the MT.
Pumping was controlled by an automated water level
controller installed in the OHT. A regulator valve was
connected to the OHT to maintain the flow through the
reactors.
All six PBBRs (R1–R6) (working volume 20 L
each) were constructed identically, consisting of a fiberglass shell with a perforated Perspex plate carrying nine
equidistantly fixed polyvinylchloride (PVC) pipes (air-
354 Part B Tools and Methods in Marine Biotechnology
The nonheterocystous marine cyanobacterium
Phormidium valderianum (which is known to produce
hydrogen under argon atmosphere) was immobilized in
combination with two other nonmarine cells (viz., the
extreme halophile Halobacterium halobium and salt
tolerant Escherichia coli) for stable, long-term biohydrogen photo-evolution under an alternating light/dark
illumination cycle (6 h on/18 h off) in a PBBR by
Patel and Madamwar [12.44], as well as by Bagai and
Madamwar [12.43].
The former workers used Ca-alginate or agar-agar
as the immobilization matrix for cell entrapment in
a 50 mL working volume PBBR (basically a packed
column) continuously fed with N-free growth medium –
comparatively better results were obtained with alginate. The problem of cell and enzyme leakage from
alginate beads (leading to loss of H 2 production)
was overcome by using glutaraldehyde as a crosslinking agent. H 2 photoevolution rate increased with
medium feed rate up to a maximum, and then declined
markedly. Stable, long-term H 2 production for 11 d was
demonstrated.
Using the same combination of microorganisms,
in a similar PBBR, but with a different immobilization matrix, viz., PVA cross-linked with glutaraldehyde,
Bagai and Madamwar [12.43] demonstrated continued
H 2 production for over 60 d. In both these studies for
photohydrogen production, compared with a free-cell
system, the immobilized system was much more productive, stable, and long lasting.
Sabu et al. [12.46] and Kumar and Chandrasekaran
[12.45] all reported the production of the therapeutically and industrially important enzyme L-glutaminase
by Ca-alginate immobilized marine microorganisms in
glass-column PBBRs, but whereas the former employed
the fungus Beauveria bassiana the latter used the bacterium Pseudomonas for that purpose. Although the
PBBRs used in these studies differed in dimensions
(i. e., column height and diameter) with the one used
by Kumar and Chandrasekaran [12.45] being larger
(height 45 cm and internal diameter 3:6 cm), the basic principle of reactor construction was the same –
immobilized viable cell beads were aseptically packed
into the column with a perforated Teflon disk placed at
the top of the packing to prevent bed expansion during
operation.
Sabu et al. [12.46] pumped the medium from the
bottom of the packed column and collected the effluent from the top. This was, however, reversed by Kumar
and Chandrasekaran [12.45]. The inlet and outlet flow
rates were kept equal to maintain a constant liquid level
at just above the bed level. Now B. bassiana being
aerobic with strong aeration dependence, air was introduced through a sparger at the bottom of the packed
column after passing through a bacterial filter. Remarkably, Sabu et al. [12.46], noted that the maximum
enzyme production rate was observed in the absence
of forced aeration, and any increase in aeration rate
caused a decrease in enzyme productivity. In order to
explain this finding, the authors posited that with external aeration, air bubbles fill the void spaces of the
packing beads, thus reducing the contact between the
cell-laden beads and the liquid medium. They also
found that enzyme productivity increased with increasing bed height/decreasing flow rate, leading them to
conclude that increase in residence time either with increase in bed height or with decrease in flow rate, in turn
increases the contact time between the immobilized
beads and the medium, thereby causing an increase in
productivity.
Kumar and Chandrasekaran [12.45] reported that
enzyme production declined with an increase in dilution
rate, irrespective of the substrate concentration used.
Again, at all dilution rates examined, enzyme yield increased with increase in substrate concentration. It was
noted that their continuous-flow PBBR system could
be operated for 120 h without any decline in enzyme
productivity.
Kumar et al. [12.47] developed a nitrifying PBBR
immobilized with an indigenous NBC (nitrifying bacterial consortium) (comprising predominantly marine
species, e.g., Nitrosospira) for rapid nitrification in
brackish water and marine hatchery systems. They
examined nitrification performances of the PBBR integrated into a recirculating aquaculture system (RAS)
viz., a Penaeus monodon (a marine crustacean –
shrimp) recirculating maturation system. The RAS
(Fig. 12.22) comprises a shrimp maturation tank (MT),
an overhead tank (OHT), Six PBBRs (designated as R1
through R6) and a collection tank (CT). Fluid from the
MT was pumped into the OHT from where it flowed
through the reactors (R1–R6) connected serially by
gravitation and finally collected in the CT, from where
the treated water was recirculated back to the MT.
Pumping was controlled by an automated water level
controller installed in the OHT. A regulator valve was
connected to the OHT to maintain the flow through the
reactors.
All six PBBRs (R1–R6) (working volume 20 L
each) were constructed identically, consisting of a fiberglass shell with a perforated Perspex plate carrying nine
equidistantly fixed polyvinylchloride (PVC) pipes (air-