6.4.8 immobilised enzymes

• product must be isolated from enzymes before use = expensive • setting up process is expensive • immobilised enzymes often less active than free enzymes = process is slower

• don’t mix with product, so extraction costs are lower • enzymes easily reused • continuous process is made easier, as no cells requiring nutrients, reproducing & releasing waste products • enzymes surrounded by immobilising matrix, protecting them from extreme conditions = wider pH range & higher temperature used without denaturing

• adsorption • covalent bonding • entrapment • membrane separation

• enzyme molecules bound to supporting surface with active site exposed = accessible to substrate • combination of hydrophobic interactions & ionic links • surfaces include: clay, porous carbon, glass beads & resin negatives: - active site may be distorted by additional interactions = affects enzyme activity - bonding forces not always strong, so enzyme can become detached/leak into reaction mixture

• enzyme molecules bonded to supporting surface by strong covalent bonds (eg. clay) • enzymes bonded using cross-linking agent, possibly linking them in chain negatives: - production of covalent boding can be expensive & distort active site = reduces activity positives: - enzymes less likely to leak/become detached

• enzyme molecules trapped in matrix which doesn’t allow free movement • schools often use calcium alginate beads to immobilise enzymes • industrial processes may use cellulose mesh positives: - enzyme molecules unaffected & remain fully active negatives: - substrate molecules must diffuse into entrapment matrix & product molecules must be able to diffuse out = method only suitable for processes where substrate/products are relatively small

• enzyme molecules separated from reaction mixture by partially permeable membrane negatives: - product/substrate molecules must be small enough to pass through membrane by diffusion - access to enzymes may limit reaction rate