Enzyme Technology
Preparation of enzymes from clarified solution
In many cases, especially when extracellular
enzymes are being prepared for sale, the clarified solution is simply
concentrated, preservative materials added, and sold as a solution or as a dried
preparation. The concentration process chosen will be the cheapest which is
compatible with the retention of enzyme activity. For some enzymes rotary
evaporation can be considered, followed if necessary by spray drying. The most
popular method, though, is ultrafiltration, whereby water and
low molecular weight materials are removed by passage through a membrane under
pressure, enzyme being retained. Ultrafiltration differs from conventional
filtration and microfiltration with respect to the size of particles being
retained (< 50 nm diameter). It uses asymmetric microporous membranes with a
relatively dense but thin skin, containing pores, supported by a coarse strong
substructure. Membranes possessing molecular weight cut-offs from 1000 to
100,000 and usable at pressure up to 2 MPa are available.
There are a number
types of apparatus available. Stirred cells represent the simplest configuration
of ultrafiltration cell. The membrane rests on a rigid support at the base of a
cylindrical vessel which is equipped with a magnetic stirrer to combat
concentration polarisation. It is not suitable for large scale use but is useful
for preliminary studies and for the concentration of laboratory column eluates.
Various large-scale units are available in which membranes are formed into wide
diameter tubes (1 - 2 cm diameter) and the tubes grouped into cartridges. These
are not as compact as capillary systems (area/volume about 25 m−1) and
are very expensive but are less liable to blockage by stray large particles in
the feedstream. Cheaper thin-channel systems are available (area/volume about
500 m−1) which use flat membrane sandwiches in filter press
arrangements of various designs chosen to produce laminar flow across the
membrane and minimise concentration polarisation. Capillary membranes represent
a relatively cheap and increasingly popular type of ultrafiltration system which
uses micro-tubular membranes 0.2 - 1.1 mm diameter and provides large membrane
areas within a small unit volume (area/volume about 1000 m−1).
Membranes are usually mounted into modules for convenient manipulation. This
configuration of membranes can be scaled up with ease. Commercial models are
available that give ultrafiltration rates of up to 600 L hr−1.
The
steady improvement in the performance, durability and reliability of membranes
has been a boon to enzyme technologists, encouraging wide use of the various
ultrafiltration configurations. Problems with membrane blockage and fouling can
usually be overcome by treatment of membranes with detergents, proteases or,
with care, acids or alkalis. The initial cost of membranes remains considerable
but modern membranes are durable and cost-effective. Ultrafiltration, done
efficiently, results in little loss of enzyme activity. However, some
configurations of apparatus, particularly in which solutions are recycled, can
produce sufficient shear to damage some enzymes.
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This page was established in 2004 and last updated by Martin
Chaplin on
6 August, 2014
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