04 Immobilisation of Enzymes


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04 Immobilisation of Enzymes

  1. 1. Immobilisation of enzymes• Immobilisation of an enzyme in alginate beads (e.g. amylase): – Amylase enzyme is mixed with a solution of sodium alginate – This mixture is dripped (usually through syringe) into a solution of calcium chloride – The sodium ions are displaced by the calcium ions – formation of hard, insoluble beads of calcium alginate, in which are trapped the molecules of amylase – The alginate beads are left to harden further – they are rinsed – Beads are normally placed in a suitable container to create a column of beads – Suspension of starch can then be trickled down the column and collected in a beaker ALBIO9700/2006JK
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  3. 3. • Advantages of immobilisation: – Enzyme can be recovered after use using a very coarse filter rather than a molecular filter – Enzyme does not contaminate product – Immobilisation may enhance stability (thermostability or pH-stability) of the enzyme molecule as it is supported – Substrate can be easily passed through the enzyme several times ALBIO9700/2006JK
  4. 4. Advantages of immobilisation Disadvantages of immobilisation1. Easier to separate enzyme and 1. Immobilisation may alter shape ofproducts enzyme2. Allows catalysis in unfavourable 2. May alter catalytic abilitymedia3. Increases stability and can be 3. Enzyme may become detachedmanipulated easily4. Allows continuous 4. Expensiveproduction/enzyme used for longer5. Enzyme can be recovered andreused6. Enzyme does not contaminateproduct/no purification required ALBIO9700/2006JK
  5. 5. The use of dipsticks and biosensors in the quantitative measurement of glucose• One way to determine the quantity of insulin needed by diabetics is to test a sample of urine with a glucose dipstick• The dipstick has the enzyme glucose oxidase - oxidises glucose into hydrogen peroxide and gluconic acid• Hydrogen peroxide will oxidise indicator chemical on dipstick – colour change• Colour change related to concentration of glucose in urine sample – colour chart• http://www.irvingcrowley.com/cls/urin.htm ALBIO9700/2006JK
  6. 6. • Biosensor: a device which uses a biological material, such as an enzyme, a cell or an antibody to detect or measure a chemical compound• Reaction between the biological material and the chemical being measured brings about a change which is converted to an electrical signal by an appropriate transducer• Electrical signal is then amplified to give a read-out on a digital display ALBIO9700/2006JK
  7. 7. • The glucose biosensor works as follows: – It contains a layer of immobilised glucose oxidase enzyme – Enzyme binds with any glucose in the blood, which is oxidised with dissolved oxygen from the solution to form hydrogen peroxide and gluconic acid – An electrode (usually platinum) measures the drop in oxygen concentration as it is used to make hydrogen peroxide – electrode generates an electrical signal – The size of the electrical signal is proportional to the concentration of glucose in the blood – A digital readout gives the user a figure for the glucose concentration ALBIO9700/2006JK
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