biologia - Studia

M.. DRĂGAN-BULARDA,. R. CARPA, M. MIŞCA, V.-CĂLEAN
glucose from amylose, respectively, many maltodextrins and limit dextrin (with
ramifications through α-1.6 bonds), some maltose and glucose from amylopectin
(Baig et al., 1984; Banu et al., 1987; Dan, 2000; Jurcoane, 2000; Malhontra et al.,
2000; Iorga and Câmpeanu, 2004; Drăgan-Bularda and Samuel, 2008).
The α-amylase produced by bacteria is a metalloenzyme with high molecular
mass of about 50,000 Da, characterized by the presence in its structure of some
inorganic ions, related to the polypeptide chains (Zarnea et al.,1988). These are Ca 2+
(1-3) ions for stabilizing the structure and 1-2 Zn 2+ ions that determine the dimer
formation of the enzyme (www.resb.org). Due to the fact that it is a protein, the
biosynthesis of the alpha-amylase takes place at ribosome level and is secreted by the
bacteria in the culture medium (Drăgan-Bularda and Samuel, 2008).
In the last decades, more and more microorganisms were used as potential
sources for various biotechnologies (Zarnea and Dumitru, 1994). Bacterial α-amylase,
which are usually produced by bacteria belonging to Bacillus genus (Nigam and Singh,
1995) are widely used in the starch-processing industry for the hydrolysis of starch
(Lonsane and Ramesh, 1990). There were tested conditions for optimizing the
biosynthesis of some enzymatic complexes (protease-amylase complex) by using
Bacillus subtilis strains (Bahrim and Dan, 1994; Dan et al., 1994), respectively, there
was studied the influence of the input of aminoacids on the biosynthesis of some
enzymes, among these also the alpha-amylase on Bacillus subtilis strains, or the effect
of some water-soluble vitamins on the biosynthesis in case of fermentation with
Bacillus subtilis strains. In most cases the enzymatic process is inhibited by high
substrate and product concentration and also instability of the enzyme repetitive or
prolonged use. Immobilization is an important technique for continuous and repeated
use of enzymes in industrial application and also rapid separation of the enzyme
from the reaction medium. The general methods employed for immobilization are
entrapment, microencapsulation, copolymerization, cross-linking, physical adsorption,
chemical attachment and covalent binding (Gangadharan et al., 2009).
Tests were conducted for immobilizing some microbial hydrolases with
protease and amylase activity on collagen support (Tcacenco et al., 1994). α-amylase
produced by Bacillus amyloliquefaciens ATCC 23842 was immobilized in calcium
alginates. Also, alginate beads have been successfully used for entrapment of α-
amylase of Bacillus subtilis and effectively used for starch hydrolysis (Rajagopalan and
Krishnan, 2008). Immobilization is a very effective alternative in overcoming problem
of instability and repetitive use of enzymes. Entrapment method of immobilization is
advantageous over other methods that do not involve chemical modification of the
enzyme (Gangadharan et al., 2009).
It should be emphasized that the application of α-amylase in different
industrial processes demands specific characteristics including thermostability,
particular pH activity profiles, pH stability and Ca-independency, consequently,
demand for novel amylases is increasing worldwide, as the variety of enzyme
applications is increasing in various industrial sectors (Pandely et al., 2000). Industrially
120