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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
Alginate, which consists of mannuronic and guluronic dimers, is widely used in
scientific research due to its exceptional biocompatibility and biodegradability.
Numerous investigations have been conducted to investigate the impact of alginate
makeup, pureness, and concentration on encapsulated cells’ viability and peptide
synthesis. Typically, it is well accepted that the purification of alginates is of utmost
importance to guarantee their biocompatibility [87]. However, there is ongoing
discussion on the appropriateness of the monomer composition of the capsules. In
recent times, there has been a notable advancement in the production of innovative
materials, encompassing oligochitosans, cellulose sulfate, pectin, and various
synthetic polymers. The application of a suitable semipermeable membrane to
coat the produced capsule matrix is an area of research that is now gaining attention.
The outer membrane will facilitate the bidirectional passage of nutrients, oxygen,
and waste, while impeding the ingress of immune cells, antibodies, and other
constituents of the immune response [88]. The membrane chemistry of Alginate-PLL
complex has received much research attention, making it the most widely investigated. Nevertheless, alternate polycations such as polyethylene glycol, poly-vinyl
alcohol, and poly-L-ornithine have also been examined in this situation. The
selection of appropriate encapsulated cells plays a crucial role in determining the
efficacy and viability of biological applications. Numerous cellular entities have
been extensively investigated as prospective contenders for sustained release of
medicinal substances. Once the appropriate polymers and therapeutic cells have
been chosen, it becomes necessary to build technologically optimized microcapsules
in order to achieve a functioning drug delivery system [89]. The selection of the
microencapsulation technique and microencapsulation system will be contingent
upon the chosen biomaterials. Consequently, distinct manufacturing techniques will
be employed depending on whether natural or synthetical polymers are selected. In
the first step, cells that produce therapeutic products are mixed with a solution of
sodium alginate, typically at concentrations of 1.6% (w/v) or 3% (w/v). Using
extrusion, the cell and gel suspension is introduced into a jellifying solution, such as
CaCl
or BaCl2. The interaction between sodium alginate, a part of the brown algae
2
cell wall and the jellifying solution induces the polymerization of the matrix due to
the ionic exchange, resulting in the formation of microbeads composed of calcium or
barium alginate. After complete gelation, microbeads containing cells are chemically
cross-linked with poly-L-lysine (PLL). Following a washing procedure, the beads
undergo an additional coating with a layer of alginate. The latter is important in
order to tackle the issue of immunological rejection, as the polycations employed in
the creation of the semipermeable membrane have an affinity for inflammatory cells,
hence triggering cells death of the captured cells. Once manufactured, microcapsules
can be utilized in the therapeutic intervention of many medical conditions.
4.12.2 Microcapsule evaluation
A key focus within the domain of cell microencapsulation pertains to the endorsement and optimization of both the biomaterials and technology employed in
constructing immobilization systems. Additionally, there is a need to develop and
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
refine assays and techniques that facilitate the analysis and replication of these
devices [90]. The proliferation of sophisticated procedures and assays in recent times
has significantly expanded the possibilities for systematizing this therapeutic
strategy. Extensive study is being conducted to investigate the purity and biocompatibility of various materials. Multiple procedures are being developed to facilitate
the beneficiation of alginates. The methodologies used in this research include the
utilization of chemical reagents and dialysis, the initiation of mortality in Jurka cells,
and the evaluation of the growth of lymphocytes by assessment. The approach
described above is a rapid and highly responsive methodology utilized to identify
potential contaminants that can induce fibrosis in alginate samples [91]. Prior
research has established heightened cytokine levels, such as tumor necrosis factor
and human monocyte upon activation by alginates of diverse compositions. After
the biomaterials have been characterized, their shape and in vivo biocompatibility as
microcapsules must be evaluated. Several techniques are constantly refined and
developed to help researchers get around these problems. Among them are photoelectron spectroscopy with x-rays (PES), atomic force microscopes (AFM), confocal
laser scanning microscopy (CLSM), and enhanced nuclear magnetic resonance
(NMR). These methods allow for a thorough study of the microcapsules and the
parts that make them up. The CLSM method looks at how alginate and PLL are
arranged in space within whole microcapsules [92]. Furthermore, NMR and AFM
methods are used together to provide solid proof of the capsules’ surface topography
and the cross-linking properties between alginate and the gelling ion. Key parameters for producing biocompatible capsules are calculated using FT-IR and x-ray
photoelectron spectroscopy, and the reactivity of the microcapsules is evaluated in
the immediate post-transplant period. The comprehensive examination of histological and immunopathological aspects and in vivo antigenicity assessment signifi-
cantly enhances the advantages of testing for biocompatibility for anchoring devices.
Different studies have been designed to test their mechanical stability to make
microcapsules stronger. A texture analyzer gadget can measure how resistant
microcapsules are to compression [93]. An alternative approach to enhance
comprehension of the mechanical characteristics of polymer systems involves
subjecting the particles to destabilizing forces, such as exposure to swollen solution
and application of shear stresses. An osmotic pressure test has been used to develop
a new way to measure the strength of microcapsules. This test entails subjecting the
microcapsules to a range of hypotonic solutions with varying concentrations and
subsequently measuring the proportion of capsules that have ruptured as a
percentage [94]. Another area of interest pertains to assessing the permeability
characteristics of the semipermeable microcapsules, namely the entry and egress
behavior. The viability of a membrane is contingent upon the extent to which
control can be exercised over the selective exclusion of molecules based on size and
the rate at which diffusion occurs. This control is crucial for regulating the survival
and metabolic efficiency of the graft. The transport of species over a membrane,
referred to as membrane permeability, is influenced by two key parameters: the
equilibrium partition coefficient, which is a thermodynamic parameter, and the
diffusion coefficient, which is a kinetic parameter. Several assays of immobilization
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
devices have been optimized to assess the permeability characteristics, specifically
the permeability to hemoglobin and immunoglobulin G (IgG) [95]. The tests used in
this study involve the technique of mass refusal chromatography, which involves
using dextran references with different molecular weights and calculating the
coefficient of mass transfer ratio. The latest technique was successfully used in the
first evaluation of microcapsules consisting of three different hurdle substances,
including poly-L-ornithin, polymethylene hydrochloride, and PLL. The PLO small
capsules had the highest level of success among each of the options. In future
research, it is recommended that the optimization of encapsulated cells should
include an evaluation of their mechanical stability, diffusion, and permeability
qualities in vivo. The resolution of these challenges will have a substantial influence
on the uniformity of immune isolation gadgets during the FDA phase [96].
4.12.3 Potential health benefits
The therapeutic applications of cell microencapsulation technology encompass a
diverse array of areas, which can be categorized into five distinct groups. The
categories mentioned above include the management of classical Mendelian
disorders, therapeutic approaches for cancer, diseases affecting the central nervous
system (CNS), the development of artificial tissues, and several other uses. Given the
current severe scarcity of donor organs, researchers have explored the utilization of
donor cells and tissues (both human and non-human) as potential therapeutic agents
[97]. Xenografts have emerged as a significant source of apprehension because of the
potential transmission of infectious organisms, including porcine endogenous
retrovirus (PERV), from the donor to the recipient. Moreover, using genetically
engineered cells has created novel opportunities for addressing conventional
Mendelian illnesses, including cancer. Genes can serve as templates, while cells
can function as reactors to produce and release the desired product. Additionally,
capsules can be employed as immunoselection vehicles to deliver drugs within a
living organism. Biosafety is a crucial factor to consider while employing genetically
modified cells. Utilizing genetically modified cells necessitates a delicate equilibrium
between ensuring the safety and stability of gene expression [98]. The scientific
foundation for various clinical trials has been established by utilizing small and large
animal models. These trials encompass a range of applications, such as the
encapsulation of allogeneic islets to address diabetes, the encapsulation of cytochrome P450 enzyme-expressing cells to combat pancreatic cancer, and the
immobilization of retinal pigmented epithelial (hRPE) cells on gelatin microcarriers
for the treatment of advanced Parkinson’s disease in patients [99].
4.12.4 Modern developments in cell encapsulation
Researchers are looking into the possibility of using settled cells that release the
granulocyte macrophage-colony-stimulating factor (GM-CSF) as an immunomodulatory or adjuvant in the context of immunization for both humans and animals
as a result of the rapid developments in cell biology, gene treatment, and chemical
engineering and pharmaceutical technology. Live vaccines can potentially be used in
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
cancer immunotherapy and the prevention of infectious illnesses [100]. Another
potential use case is immobilizing genetically modified cells that produce therapeutic
antibodies. Various cell types have been employed in this methodology, encompassing hybridomas, skin fibroblasts, keratinocytes, myogenic cells, and hepatocytes.
Notably, in certain instances, antibodies generated through both in vitro and in vivo
methods maintain the specificity and affinity of the original parenteral antibody
without any observed idiotypic reaction in animals that create ectopic antibodies
[101]. In a similar vein, the utilization of encapsulated cells that produce retroviral
vectors has been examined to achieve sustained in vivo gene transfer, given the
requirement for frequent vector administration to ensure effective delivery of the
therapeutic gene. Significant endeavors are being undertaken by scientists in the
advancement of a bioartificial pancreas by the process of immobilizing pancreatic
islets within polymer microcapsules. Several research groups have focused their
efforts on developing a neovascularized environment suitable for the implantation of
encapsulated islets [102]. While not universally endorsed by all specialists, this
methodology can enhance the interaction between the bloodstream and immobilized
cells, potentially improving their long-term performance and functionality. This
strategy can be readily implemented by employing the encapsulation technique of
cells that secrete angiogenic factors or by utilizing gelatin microspheres impregnated
with the administration of basic FGF or VEGF, which stands for vascular
endothelial growth factor [103].
4.13 Immobilization of microalgae
The utilization of algae for commercial purposes has a substantial historical
background, with numerous algae species employed as food, feed, and important
chemical resources. Various kinds of macroalgae, such as Iridaea, Porphyra,
Gigartina, Caulerpa, and Ulva, are commonly collected for human consumption
or the extraction of hydrocolloids. The cultivation of Porphyria, commonly referred
to as ‘Nori’ in Japan, is a significant sector within the aquaculture industry of Japan
[104]. Furthermore, the industries manufacturing carrageenan, agar, and alginic acid
from macroalgae have been firmly established. Certain microalgae, including
specific Nostoc and Aphanizomenon species, are procured from natural habitats
for human consumption. However, most microalgae utilized in commercial applications are cultivated. Several microalgae species are known for their potential to
produce valuable compounds [105].
For instance, Dunaliella is recognized for synthesizing β-carotene, while Chlorella
and Spirulina are commonly utilized for protein production. Spoehr and Milner (1949)
proposed that the cultivation of microalgae on a large scale may address the issue of
protein deficiencies worldwide. The rationale behind their positive outlook stemmed
from algae having a crude protein content surpassing 50% and demonstrating a
biomass production of approximately 25 tonnes per hectare per year. In addition,
wastewater high in nitrogen and phosphorus is also considered a useful substrate for
algae production. Cultivating algae in wastewater presents a dual benefitoftreating
the wastewater while generating algal biomass. Biomass has the potential to be used
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
for a range of applications, such as the production of nutritional supplements and food
additives for aquaculture, as well as for animal and human feed [106].
Additionally, it can be harnessed for energy production, such as biogas and fuels,
and for applications in agriculture, such as fertilizers and soil conditioners. One of the
primary obstacles in the utilization of microalgal biomass is in the process of
harvesting or separating the algal biomass from the discharged water after treatment.
Considerable attention has been dedicated to developing appropriate technology for
the extraction of microalgae, encompassing several methods such as basic sand
filtering and more energy-demanding centrifugation techniques. The practice of
autoflocculation, which involves the self-aggregation of cyanobacteria by the cessation
of aeration and subsequent decantation, has also been seen [107]. Experiments have
also been conducted on the utilization of ultrasonic waves or ultrasound for the
harvesting of microalgae. In this approach, the separating process relies on the gentle
aggregation generated by acoustic forces, which is subsequently followed by an
intensified sedimentation phase. In the present context, the immobilization of algal
cells has been suggested as a potential solution to address the challenge of harvesting
and to preserve the valuable algal biomass for subsequent processing [108].
4.13.1 Techniques for immobilization
Among the several techniques employed for immobilizing cells and enzymes,
entrapment stands out as the predominant approach for algal immobilization.
This specific application utilizes a range of natural polymers, including collagen,
which is agar, agarose, or cellulose, and alginate, which and carrageenan, alongside
synthetic polymers such as acrylamide, polyurethane, and polyvinyl [109]. In the
algal immobilization process, it is important to acknowledge that the often-used
natural gels contain alginate and carrageenan.
The gel is commonly converted into functioning biological catalysts pellets by the
introduction of microalgae cells in solution to an aquatic solution that contains the
gelling component. As mentioned above, the substance is subsequently transformed
into small spherical particles by applying controlled droplet formation techniques,
passing via a nozzle or orifice, and introduced into a solution containing interacting
salts [110]. The droplets are later rendered stable through polymerization or other
forms of cross-linking, resulting in biocatalyst beads containing the imprisoned
organisms. As an illustration, alginate droplets have the potential to be rendered
stable through the introduction of bivalent ions, such as calcium. Conversely,
carrageenan beads are commonly subjected to cross-linking processes utilizing K+
ions. Monodisperse calcium alginate beads are generated within the size range of
0.2–1.0 mm through the laminar jet break-up, ensuring sterility and reproducibility.
Implementing an in situ cleaning procedure for the nozzles is undertaken to ensure
the completion of multiple batch process cycles and achieve a throughput level of up
to 5.3 l h
−1
[111]. The beads were subjected to analysis, revealing that the relative
disparity in the average diameter across several nozzles was below 0.3%. Recently, a
study reported the comobilization of microalgae with the bacteria Az spirillum
Brasiliense. This comobilization was observed to enhance the growth, pigment, and
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
lipid contents, In addition to the cells and population density of the entangled algae.
The results of this study indicate that the novel immobilized biocatalyst exhibited
greater efficacy in the removal of nitrogen and phosphorus compared to the use of
immobilized algae alone. Since 1990, half of the papers about immobilized microalgae have focused on their application in the removal of nitrogen and phosphorus
from wastewater [112]. The remaining reports have explored the utilization of
immobilized algae for the accumulation and removal of metals. Recently, there has
been a notable development in the field of conductometric biosensors, namely in
utilizing immobilized microalgae. Fiber optic biosensors are employed to detect
environmentally hazardous substances, including heavy metals, herbicides, and
other toxic compounds.
4.13.2 Use of cryopreserved algae
Research indicates that immobilization significantly affects the productivity of algal
cells. Specifically, studies have shown a remarkable enhancement in hydrogen
production from the filamentous alga Anabaena, with a threefold increase observed
following immobilization. This demonstrates the profound impact that immobilization
can have on algal cell function and output. Brouwers has demonstrated that
immobilization notably enhances the production of ammonia and hydrocarbons in
Mastigocladus laminosus [113]. The immobilization of ‘Chlamydomonas reinhardtian’
cells in Ba-alginate offers a reliable and efficientapproachforthephotoproductionof
ammonia. The study demonstrated that the generation of glycolate in cells entrapped
in alginate was twice as high as in cells that were not entrapped and were free-living
[114]. Leon and Galvan investigated glycerol production in C. reinhardtian cells fixed
in Ca-alginate. The immobilized cells exhibited a production rate of 8 g l
their free-living counterparts achieved a invention rate of 4 g l
−1
. The comparative
−1
,whereas
competence of arrested cells and thylakoid vesicles of the microalga C. reinhardtia was
assessed concerning their free-existing foils, specifically in hydrogen peroxide generation [28]. Hydrogen peroxide (H
) is a highly effective and environmentally
2O2
friendly fuel in various applications such as rocket propulsion, motors, and heating
systems. The molecule in question is generated through a catalytic cycle within the
photosystems, wherein a redox mediator, namely Me viologen, undergoes reduction
by the electrons derived from water [105]. The Ca-alginate-entrapped cells maintained
a photoproduction rate of 33 μmol H
/mg Chl./h for several hours under ideal
2O2
conditions, exhibiting an energy conversion efficiency of 0.25%.
On the other hand, the immobilized cyanobacteria, specifically Anabaena
variabilis and Anacystis nidulans, demonstrated reduced catalase activity and
attained productivities of 151 and 61 μmol hydrogen peroxide. A novel photobioreactor has been developed to enhance Maren nine production, as indicated by
the findings of this study. In a recent study, the microalgae species Butyraceous
braunii and B. protuberans were immobilized as entire cells within alginate beads
during airlift batch culture. This immobilization technique led to a significant
increase in rest phase development, during which both resting and hydrocarbon
output are substantially enhanced [104].
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
4.13.3 Removal of nitrogen and phosphorous
Numerous publications have been dedicated to examining the application of
immobilized microalgae to remove nitrogen and phosphorus from wastewater,
specifically focusing on its potential as a tertiary treatment method. Typically,
immobilized cells exhibit more efficiency in removing nitrogen and phosphorus than
their free-living counterparts [115]. Furthermore, the removal of phosphate is shown
to occur at a slower rate than that of nitrogen. Moreover, a progressive decrease in
efficiency was also noted from the initial cycle to the succeeding ones. In 1986,
Jeanfils and Thomas conducted a study involving alginate-immobilized Scenedesmus
obliquus. Their findings indicated that the efficiency of nitrite uptake was not
impacted by immobilization, except for a prolonged lag phase found in the
immobilized cells compared to the free cells. The available research supports the
notion that culture age has a negligible effect on nitrogen removal [116]. In contrast
to those mentioned earlier, previous batch culture investigations on phosphate
uptake by alginate-entrapped Chlorella Emersonian have demonstrated that the cells
in the exponential growth phase exhibit a phosphate removal rate from the medium
that is five times higher than that of cells in the late stationary phase. Significant
variations in reactor performance can be observed when immobilized cells of
varying ages are enclosed in Ca-alginate and placed within a small-scale packedbed reactor. The rate of nitrogen intake in immobilized S. obliquus was significantly
enhanced under conditions of nitrogen scarcity. Rai and Mallick (1992) observed a
greater rate of uptake for nitrogen and phosphorus in immobilized Chlorella and
Anabaena compared to their free-living counterparts [117]. In the following inves-
tigations, it was observed that immobilized algae with a cell density of 0.1 g dry
weight per liter exhibited the highest efficacy in nutrient and metal removal within a
pH range spanning from 6.0 to 8.0. Furthermore, using chitosan as a substrate for
supporting algae has shown promise in enhancing the process of wastewater
detoxification. The research done by Vilchez and Vega shows that using alginateentrapped C. reinhardtian cells offers a dependable and efficient method for
removing nitrogenous contaminants in wastewater [118]. To determine the most
favorable operating parameters for the immobilized cells, many criteria were
considered, such as the concentration of the matrix, the loading of cells, the
temperature, and the pH. In the case of C. reinhardtian cells, using a 3% alginate
concentration effectively addresses challenges related to substrate transport. This
concentration level facilitates the attainment of the appropriate physical properties
of the beads. This study aims to evaluate the efficacy of foam-immobilized devices in
the removal of phosphate [119]. This study compared the performance of Phormium
laminose in batch bioreactors to that of continuous flow bioreactors. Beds in
funneled columns and beds housed in Erlenmeyer flasks were the first fluidized-bed
designs to be invented. The bioreactors were treated to a continuous illumination
employing cool white fluorescent lamps with 100 μmol photon m
2s−1
irradiation.
The bottom half of each bioreactor was immersed in a thermostatically controlled
water bath to keep the temperature within at a constant 45 °C. Preboiled, cleaned,
and dried foam cubes (now 5 mm in size) were mixed with algae suspension. The
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
adsorption immobilization procedure was placed over two months. The foam cubes
were put in the bioreactors once fully colonized [120]. Despite the excellent removal
of nitrogen and phosphorus from the system by cyanobacteria immobilized on
polymer foams, the utilization of fluidized-bed reactors led to the development of a
heterogeneous system, rendering it unsuitable for laboratory standardization in most
cases. However, numerous researchers have investigated the removal of nitrogen
and phosphorus in packed-bed reactors. Limitations in the operation of these
reactors include inadequate light penetration, lack of cell mixing, and the presence of
gaseous fluxes. Kaya and Picard have successfully devised an innovative immobilized algal system for wastewater biotreatment. The present study involved the
cultivation of Scenedesmus bicellularis, a green microalga obtained from a secondary
decantation tank. The microalga was cultured in a synthetic medium for 12 days
[121]. Following centrifugation, the harvested cells were immobilized on alginate
screens. The screens were placed within a photochamber fully saturated with a
relative humidity of 100%. The photochamber had a photoperiod of 16 h and was
illuminated with an intensity of 150 μEm
2s−1
. Following 48 h of nutrient
deprivation, the immobilized cells were employed in the elimination of ammonium
and orthophosphate from a synthetic secondary wastewater effluent within a
flexiglass reactor. A second study additionally revealed that intermittent carbon
dioxide (CO
) enrichment expedites the process of tertiary wastewater treatment.
2
Nevertheless, a significant issue associated with using gel materials pertains to the
challenge of preserving the structural stability of alginate gels for an extended
period, typically beyond a few weeks [121]. While agarose gel-based reactors have
demonstrated greater stability compared to other types of reactors, it is important to
note that these matrices, composed of polysaccharides, are very susceptible to
microbial degradation when exposed to natural environments. In order to address
these challenges, Robinson put out a novel design for a hollow-fiber reactor. A
diverse range of sizes of hollow-fiber cartridges can be found in the commercial
market. Robinson developed a hollow-fiber reactor consisting of 50 cylindrical tubes
made of polysulfone. These tubes were bundled together and securely enclosed
within a transparent cartridge measuring 20 cm long. Each fiber possessed an
internal diameter (ID) of 1.1 mm, and the fiber wall exhibited many perforations in
the form of pores. One potential method involves the use of algae cells contained
within the fiber lumen of a cartridge. Nutrients can be introduced either through the
shell space or in the reverse direction, depending on the specific requirements of the
experimental setup. The initial findings indicate a rapid fall in the phosphate uptake
rate within a short timeframe [122]. This decline is not attributed to decreased
activity but to settling biomass within the reactor. Nevertheless, the settling rates of
cells were seen to be considerably reduced when they were immersed in a solution
containing 1% Na-alginate. The investigation focused on eliminating nitrate and
phosphate from wastewater at 43 °C. This was achieved by providing a constant
supply of a diluted growth medium for seven days, followed by introducing
secondary-treated sewage for an additional 12 days [123]. Under identical conditions, the nitrogenous and phosphate ions eliminated from secondary-treated
sewage were recorded as 0.25 and 0.12 mmol d
−1l−1
. The utilization of thermophilic
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
cyanobacteria in a wastewater purification system offers distinct advantages due to
their capacity to withstand elevated temperatures, hence minimizing the risk of
contamination. The coimmobilization of Azospirillum brasilense within small
alginate beads led to significant enhancements in the growth, pigment and lipid
content, as well as the cell and population size of both Chlorella species. The
simultaneous mobilisation of both microorganisms was found to be more efficient in
the removal of nitrogen and phosphorus as compared to the use of just immobilized
microalgae [124]. In a 6-day timeframe, this comobilization method removed up to
100% of ammonium, 15% of nitrate, and 36% of phosphorus, with some variation
between wastewater types. Using microalgae alone, on the other hand, led to lower
removal rates: 75% for ammonium, 6% for nitrate, and 19% for phosphorous.
4.13.4 Disposal of metals
Microalgae have garnered significant attention in the biotechnology field due to
their potential applications in removing heavy metals and radionuclides from
emissions and effluents. Furthermore, it is plausible to extract valuable elements
such as gold, silver, and uranium alongside the detoxifying process by appropriately
treating the algal biomass enriched with these metals. Numerous studies have been
conducted on the sequestration of metals by microalgae-immobilized systems [125].
One particularly intriguing aspect is the development of ‘AlgaSORB’ at New
Mexico State University. During this procedure, the algae are densely packed
within a columnar structure composed of solid silica gel. The microorganisms are
rendered non-viable by encapsulating the algae within a solid matrix. However, the
cell walls of these organisms continue to serve as a rich source of binding sites that
can effectively sequester heavy metal ions from the surrounding solution. The
current work explored extracting mercury from an aqueous solution utilizing a
packed-bed reactor (PBR) that included Chlorella emersonii immobilized inside
alginate and agarose gels [126]. The process of microalgae cultivation using wastewater for biomass production was reported in figure 4.14.
Figure 4.14. Integrated process of microalgae cultivation using wastewater for biomass production and its
applications in biofuels, animal feed, and biofertilizers.
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Introduction to Pharmaceutical Biotechnology, Volume 2 (Second Edition)
The reactors were made with chromatography columns filled with 200 gel
particles between 4 and 6 mm across. The point of this study was to find out
what happens to the mercury removal process when the cell packing density, the
amount of mercury coming in, and the flow rate change. The investigation revealed
that using agarose as an immobilizing matrix, as opposed to alginate, led to a
reduction in the amounts of mercury volatilization. A laboratory-scale algal column
reactor was created, utilizing Chlorella vulgaris, a species of green microalgae, with a
combined volume of 75 ml of alginate-algal beads. The reactor was employed for the
remediation of copper (Cu) and nickel (Ni) at a concentration of 30 mg l
−1
, utilizing
a hydraulic retention time (HRT) of 30 min. After the loading process of a 4 l metal
solution was concluded, it was noted that a significant proportion of copper (Cu)
and nickel (Ni), namely over 97% and 91%, respectively, were effectively extracted
from the wastewater. It was discovered that up-flow is preferable to down-flow
for keeping the flow rate stable. Gardea-Torresdey et al [127] immobilized
Synechococcus sp. PCC7942 biomass in a silica-polymer matrix in their experiment.
The ability of the immobilized biomass to bind metals was then studied under
continuous flow conditions. The experimental findings demonstrated that the highest
degree of adsorption was observed for lead (Pb), followed by cadmium (Cd), with
copper (Cu) and nickel (Ni) exhibiting comparable levels of adsorption [128]. A
concentration of 0.2 M hydrochloric acid was determined to be efficacious in the
retrieval of the metals that had been adsorbed. A series of experiments were
undertaken to ascertain the feasibility of many metal binding and stripping cycles
by the immobilized biomass. The elements Cd, Cu, and Ni were shown to undergo
sorption and desorption processes three times, whereas Pb exhibited as many as six
sorption and desorption events. The study investigated the metallic elimination
effectiveness of the Scenedesmus acutus and Chlorella vulgaris, immobilized in foam
polyurethane and κ-carrageenan gel matrices. The effectiveness was evaluated in
both fluidized-bed and packed-bed reactors. The tolerance of immobilized cells to
heavy metals such as Cd, Cr, and Zn is greater than that of free cells, suggesting a
significant potential for their application in wastewater treatment procedures. Singh
and Prasad constructed a column named ‘AlgaSORB’ by combining silica, a
polymer matrix (poly-N-xylene-N,N′-di cyclohexyl ethylenediamine dibromide),
and a green microalga (Spirogyra) [129]. This column exhibited distinctive ion
selectivity for copper (Cu) over other metal ions in the waste samples [130].
Standardizing factors such as flow rate, pH, and equilibrium time also impact the
outcomes. The system’s kinetics are advantageous, as it does not exhibit any issues
related to clumping, clogging, or leaching of the stationary support. The present
method possesses appealing characteristics that make it suitable for single-column
ion chromatography due to its exceptional durability, straightforwardness, and costeffectiveness. An experimental study was conducted to evaluate the effectiveness of a
novel sorption system utilizing Chlorella sorokiniana immobilized on the biomatrix
of Luffa cylindrica, commonly known as a vegetable sponge. The objective was to
investigate the system’s capability to remove Cd and Ni from a contaminated
aqueous medium under continuous liquid flow conditions in a column setup, and the
immobilized Chlorella sorokiniana exhibited notably superior efficiency compared to
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