RP 1 - Rate of Enzyme Controlled Reaction

• The rate of reaction of an enzyme-controlled reaction is influenced by different factors: the temperature, pH, concentration of the substrate, and the concentration of the enzyme.
• The effect of each of these can be determined by changing a single variable and measuring its effect on the rate of reaction.
• It is important to keep all other variables constant so that they do not influence the results.
• In this method, the named variable is temperature.
• Make two control samples: take two flat bottomed tubes, add 5cm3 of milk suspension to each tube, add 5cm3 of distilled water to one tube, and add 5cm3 of hydrochloric acid to the other tube.
• Take three test tubes and measure 5cm3 milk into each, place in water bath at 10°C for 5 minutes to equilibrate, add 5cm3 trypsin to each test tube simultaneously and start the timer immediately, record how long it takes for the milk samples to completely hydrolyse and become colourless, repeat steps 2-3 at temperatures of 20°C, 30°C, 40°C and 50°C, find the mean time for the milk to be hydrolysed at each temperature and use this to work out the rate of reaction.
• In emergency, risk level is broken glass, cuts from sharp object, take care when handling glass objects, elevate cuts, apply pressure, do not remove glass from wound, seek medical assistance.
• Hydrochloric acid may cause harm/irritation to eyes or in cuts, wear eye protection, avoid contact with skin, tie up long hair, wash off skin immediately, flood eye/cuts with cold water.
• Hot liquids can cause scalding, handle with care, use tongs to remove boiling tubes from water bath, wear eye protection, keep away from the edge of the desk, run burn under cold water, seek medical assistance.
• Enzymes can cause allergies, avoid contact with skin/eyes, wear eye protection, seek assistance.
• Graph: plot a graph of rate of reaction against temperature.
• Milk contains a protein called casein which, when broken down, causes the milk to turn colourless.
• Trypsin is a protease enzyme which hydrolyses the casein protein.
• As the temperature increases from 10°C, kinetic energy increases so more enzyme-substrate complexes form, meaning that the rate of reaction increases up to the optimum temperature.
• At temperatures beyond the optimum, bonds in the enzyme tertiary structure break, which changes the shape of the active site, meaning that the substrate and enzyme are no longer complementary.