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T. Polley and U. Ghosh
an immobilized enzyme to harsh environmental conditions such as large ranges of
pH and temperature (Cipolatti et al. 2014). In the last decades, several methods have
been used for immobilization of enzymes. These techniques are commonly classified into four wide categories; that is, covalent binding, adsorption, entrapment, and
cross-linking (Dwevedi et al. 2016). In general, the chemical and physical properties
of the supports used in combination with the biochemical properties of the enzyme
play a key role in selecting the most appropriate immobilization technique (Mateo
et al. 2007).
Immobilized enzymes offer more advantages, when compared to free enzymes,
like enhanced stability against various denaturing conditions, higher catalytic
activity, easier product and enzyme recovery, continuous operation of enzymatic
processes, reusability, and reduced susceptibility to microbial contamination (Asftraf
et al. 2010). However, limitations in applications of immobilized enzymes include
high cost and low yield (Matto et al. 2007). The effective support materials for an
immobilized enzyme should be low cost and provide an adequate large surface area,
together with the least diffusion limitation in the transport of substrate and product
(Krajewska et al. 2004). Natural polymers used as carrier materials in immobilization
technology, such as alginate, carrageenan, agarose, chitin, and chitosan, along with
their application in the treatment of various pollutants, have the advantages of being
nontoxic, biocompatible, and biodegradable (Zille et al. 2003). Used tea is cheap,
easily available, and nontoxic biological waste material. The utilization of used tea
as solid matrix will not only reduce the disposal as well as pollution problem but will
also help the immobilized enzyme to be cost-effective. The present study deals with
i. Immobilization of alkaline protease using polyphenol extracted used tea as solid
matrix.
ii. Characterization of immobilized enzyme to temperature & pH optima, storage
stability, and reusability.
iii. Comparative study of thermal stability and pH stability of free and immobilized
enzyme.
2 Materials and Methods
2.1 Microorganism and Inoculum Preparation
Alternaria alternata TUSGF1 strain was originally isolated from poultry farm soil
and identified as producer of alkaline protease according to Polley et al. (2018). For
inoculum preparation, the strain was grown on PDA agar slant at 30 °C for 7 days
incubation.
T. Polley and U. Ghosh
an immobilized enzyme to harsh environmental conditions such as large ranges of
pH and temperature (Cipolatti et al. 2014). In the last decades, several methods have
been used for immobilization of enzymes. These techniques are commonly classified into four wide categories; that is, covalent binding, adsorption, entrapment, and
cross-linking (Dwevedi et al. 2016). In general, the chemical and physical properties
of the supports used in combination with the biochemical properties of the enzyme
play a key role in selecting the most appropriate immobilization technique (Mateo
et al. 2007).
Immobilized enzymes offer more advantages, when compared to free enzymes,
like enhanced stability against various denaturing conditions, higher catalytic
activity, easier product and enzyme recovery, continuous operation of enzymatic
processes, reusability, and reduced susceptibility to microbial contamination (Asftraf
et al. 2010). However, limitations in applications of immobilized enzymes include
high cost and low yield (Matto et al. 2007). The effective support materials for an
immobilized enzyme should be low cost and provide an adequate large surface area,
together with the least diffusion limitation in the transport of substrate and product
(Krajewska et al. 2004). Natural polymers used as carrier materials in immobilization
technology, such as alginate, carrageenan, agarose, chitin, and chitosan, along with
their application in the treatment of various pollutants, have the advantages of being
nontoxic, biocompatible, and biodegradable (Zille et al. 2003). Used tea is cheap,
easily available, and nontoxic biological waste material. The utilization of used tea
as solid matrix will not only reduce the disposal as well as pollution problem but will
also help the immobilized enzyme to be cost-effective. The present study deals with
i. Immobilization of alkaline protease using polyphenol extracted used tea as solid
matrix.
ii. Characterization of immobilized enzyme to temperature & pH optima, storage
stability, and reusability.
iii. Comparative study of thermal stability and pH stability of free and immobilized
enzyme.
2 Materials and Methods
2.1 Microorganism and Inoculum Preparation
Alternaria alternata TUSGF1 strain was originally isolated from poultry farm soil
and identified as producer of alkaline protease according to Polley et al. (2018). For
inoculum preparation, the strain was grown on PDA agar slant at 30 °C for 7 days
incubation.