Isolated enzymes
Clear advantages
less wasteful --> don't produce excess biomass
more efficient --> work at higher concentrations that if whole microorganisms were used
more specific --> no unwanted enzymes present
maximise efficiency --> can be given ideal conditions for maximum product formation
less downstream processing --> pure product is produced
Most isolated enzymes are extracellular as they're generally easier and cheaper to use than intracellular
they are secreted so easier to isolate
each microorganisms produces few extracellular enzymes so its easy to identify and isolate the desired one
they tend to be more robust that intracellular ones (adapted to cope in greater variations of pH and temperature)
But intracellular enzymes are sometimes isolated and used because there's a wider range of them and in some cases they're the only ideal one
benefits of using very specific intracellular enzymes sometimes outweigh the smaller costs of using an extracellular one
glucose oxidase for food preservation
asparaginase for cancer treatment
penicillin acylase for converting natural penicillin to semi-synthetic drugs
Immobilised enzymes
isolated enzymes are more efficient that using whole organisms but often very wasteful
enzymes aren't cheap and at the end of process are easily lost
increasingly, industries are using immobilised enzymes instead (attached to an inert support system which the substrate is passed over and the products are formed)
because they are held stationary during the process they can recovered from the reaction mixture and reused --> they don't contaminate the end product so downstream processing is needed less
Advantages
can be reused so are cheaper
easily separated from reactants so less downstream processing - cheaper
more reliable because there's a high degree of control over the process
they have a greater temperature tolerance as they're less easily denatured
easily manipulated so can keep bioreactors running continuously
Disadvantages
reduced efficiency as the immoblising process may reduce their activity rate
higher initial costs of materials as immobilised enzymes are more expensive than free enzymes
higher initial costs of the bioreactor as they system is different
more technical issues as the systems are more complex
How are they immobilised?
Adsorption to inorganic carriers (cellulose, silica)
Adv = cheap, simple, used for many processes, enzymes very accessible
Disadv = enzymes can be lost from matrix easily
Covalent or ionic bonding to inorganic carrier (covalent to carrier with amino group or ionic to polysaccharides)
Adv = cost varies, enzymes are strongly bound, very accessible to substrate, pH and substrate concentration have little effect on activity
Disadv = cost varies, active site may be modified in the process
Entrapment in a matrix (polysaccharides, gelatin)
Adv = widely applicable to different processes
Disadv = may be expensive, can be difficult, diffusion to and from active site can be slow, effect on activity can vary
Membrane entrapment in microcapsules (polymer-based semi-permeable membranes)
Adv = relatively simple, relatively small effect on activity, widely applicable
Disadv = relatively expensive, diffusion to and from active site can be slow
Uses of immobilised enzymes
Penicillin acylase --> make semi-synthetic penicillins from natural penicillin to use against penicillin resistant strains or for people who're allergic to penicillin
Glucose isomerase --> used to convert glucose to fructose as its sweeter so less needs to be used in foods to make them taste the same
Lactase --> used to make lactose-free dairy products for lactose intolerant people (converts lactose to glucose and galactose)
Aminoacylase --> used to produce pure samples of L-amino acids
Glucoamylase --> used to complete the breakdown of starch to glucose syrup