either carrier-bond or carrier-free enzymes. Detailed methodology has been
published in several reviews (Brady and Jordaan 2009; Hanefeld et al. 2009) and a
brief summary of each technique is summarized in this chapter.
Immobilization on a support depends on the properties of both the enzyme and
the carrier material. A typical and straightforward method is physisorption (i.e.,
physical adsorption). It is a reversible process commonly based on hydrophobic or
ionic interactions, van der Waals forces, and hydrogen bonding. This method is very
simple and requires only mixing the enzyme and the carrier material under appropriate pH conditions and ionic strength. Because of weak interactions between the
enzyme and the carrier, several factors like temperature fluctuations, changes in the
pH, and ionic concentrations may cause desorption of the enzyme (Ahmad and
Sardar 2015). Spontaneous desorption is the main disadvantage. It can be controlled,
however, and utilized for regeneration and reloading the support with fresh enzyme.
The cost of the enzyme is often a primary factor in the overall cost of the
immobilized catalysts (Mohamad et al. 2015).
Chemisorption (i.e., covalent binding) is a widely used strategy for enzyme
immobilization. It is mostly irreversible once covalent bonds have been formed
between the enzyme and the carrier. Side chain functional groups of the enzyme
like amino, thiol, carboxylic, imidazole, and phenolic are involved in the binding;
however, it is important to verify that the groups recruited for immobilization are
not essential for the catalytic activity of the enzyme. The surface of the support is
commonly activated by electrophilic groups, e.g., epoxide, which react with
nucleophiles on the protein. Covalent immobilization prevents enzyme release
from the carrier surface into the reaction system, which results in higher stability.
Conformational changes and orientation of the enzyme after the attachment to the
support may cause an increase or decrease in the specific activity. Compared to the
physical adsorption, covalent binding is irreversible and if the enzyme activity is
decayed, it cannot be easily desorbed and the support cannot be simply regenerated
(Mohamad et al. 2015).
Entrapment involves inclusion of an enzyme in a polymer network (gel lattice)
such as an organic polymer or a silica sol–gel, or a membrane device such as a
hollow fiber or a microcapsule. Entrapment requires the synthesis of the polymeric
network in the presence of the enzyme (Homaei et al. 2013). Encapsulation of an
enzyme is the formation of a membrane-like physical barrier around an enzyme
preparation. Encapsulated enzymes are enclosed within the internal capsule phase by
a semipermeable membrane coating or a porous polymeric network structure
(Koyani and Vazquez-Duhalt 2016). Entrapment and encapsulation restrict the
movement of the enzyme and only the substrate and products can pass through.
By adequate selection of the polymer material and modification of the immobilization process, it is possible to create an ideal microenvironment for the enzyme with
optimal pH and polarity. The entrapped and encapsulated enzyme is either free
molecule or additionally covalently attached to the support. Entrapment and encapsulation of the enzyme in polymer network can improve the mechanical stability,
prevent enzyme leaching, and protect the enzyme against external environment. The
main disadvantage of these methods is mass transfer limitations. Large substrates
16 Enzyme-Based Nanomaterials in Bioremediation
349
published in several reviews (Brady and Jordaan 2009; Hanefeld et al. 2009) and a
brief summary of each technique is summarized in this chapter.
Immobilization on a support depends on the properties of both the enzyme and
the carrier material. A typical and straightforward method is physisorption (i.e.,
physical adsorption). It is a reversible process commonly based on hydrophobic or
ionic interactions, van der Waals forces, and hydrogen bonding. This method is very
simple and requires only mixing the enzyme and the carrier material under appropriate pH conditions and ionic strength. Because of weak interactions between the
enzyme and the carrier, several factors like temperature fluctuations, changes in the
pH, and ionic concentrations may cause desorption of the enzyme (Ahmad and
Sardar 2015). Spontaneous desorption is the main disadvantage. It can be controlled,
however, and utilized for regeneration and reloading the support with fresh enzyme.
The cost of the enzyme is often a primary factor in the overall cost of the
immobilized catalysts (Mohamad et al. 2015).
Chemisorption (i.e., covalent binding) is a widely used strategy for enzyme
immobilization. It is mostly irreversible once covalent bonds have been formed
between the enzyme and the carrier. Side chain functional groups of the enzyme
like amino, thiol, carboxylic, imidazole, and phenolic are involved in the binding;
however, it is important to verify that the groups recruited for immobilization are
not essential for the catalytic activity of the enzyme. The surface of the support is
commonly activated by electrophilic groups, e.g., epoxide, which react with
nucleophiles on the protein. Covalent immobilization prevents enzyme release
from the carrier surface into the reaction system, which results in higher stability.
Conformational changes and orientation of the enzyme after the attachment to the
support may cause an increase or decrease in the specific activity. Compared to the
physical adsorption, covalent binding is irreversible and if the enzyme activity is
decayed, it cannot be easily desorbed and the support cannot be simply regenerated
(Mohamad et al. 2015).
Entrapment involves inclusion of an enzyme in a polymer network (gel lattice)
such as an organic polymer or a silica sol–gel, or a membrane device such as a
hollow fiber or a microcapsule. Entrapment requires the synthesis of the polymeric
network in the presence of the enzyme (Homaei et al. 2013). Encapsulation of an
enzyme is the formation of a membrane-like physical barrier around an enzyme
preparation. Encapsulated enzymes are enclosed within the internal capsule phase by
a semipermeable membrane coating or a porous polymeric network structure
(Koyani and Vazquez-Duhalt 2016). Entrapment and encapsulation restrict the
movement of the enzyme and only the substrate and products can pass through.
By adequate selection of the polymer material and modification of the immobilization process, it is possible to create an ideal microenvironment for the enzyme with
optimal pH and polarity. The entrapped and encapsulated enzyme is either free
molecule or additionally covalently attached to the support. Entrapment and encapsulation of the enzyme in polymer network can improve the mechanical stability,
prevent enzyme leaching, and protect the enzyme against external environment. The
main disadvantage of these methods is mass transfer limitations. Large substrates
16 Enzyme-Based Nanomaterials in Bioremediation
349