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To determine if the rate of reaction of free enzymes is higher that the rate of reaction in immobilised enzymes in different pH concentrations of solution.

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AS Level Human Biology Investigation into the effects of pH on free and immobilised amylase By: Laura Carolan-Munslow Aim: To determine if the rate of reaction of free enzymes is higher that the rate of reaction in immobilised enzymes in different pH concentrations of solution. Introduction: Enzymes are biological catalysts that help biological reactions, which normally occur very slowly at biological temperatures, to occur faster. Enzymes do this by lowering the activation energy (the energy required to get the reaction started, and break some bonds initially). By lowering the activation energy, the reactions can occur much faster. Most enzymes are globular proteins, (though some are RNA enzymes). Protein enzymes have a 3D formation, which has to remain intact for the enzyme to function correctly. If this 3D shape is destroyed, or even altered, the enzyme cannot function. When this happens the enzyme is said to be denatured. An enzyme can be denatured in various ways; the main factors that affect the activity of the enzyme are pH, temperature, and substrate concentration. If the pH is higher or lower than the optimum level, then the enzyme will denature. If the temperature is too high, the bonds of the 3D shape will break, and thus the enzyme will denature. Every enzyme has an optimum pH, (the pH at which the enzyme will function best) at this pH the highest reaction rate will be recorded. This will depend on the 3D bonds of the enzyme. Above the optimum pH of the enzyme, the conditions will cause ionization of the enzyme, which results in the decrease of the number of reactions, due to the breaking of the bonds holding the 3D shape together. ...read more.


Bead size - When making the beads it is hard to control the size. During this process, effort must be made to ensure that they are roughly the same size, do not contain any air bubbles (this can be checked, if the beads float in the calcium chloride, they contain air bubbles), and don't have "tails" ("tails" are created when the beads have been released into the calcium chloride to near to the surface). By controlling these factors, the rate of the reaction isn't affected. Temperature - The temperature must be kept constant throughout the experiment, as increasing/decreasing temperature, affects the enzymes rate of reaction. If the temperature is increased during the experiment, at a certain temperature (around 40�C) the enzymes will denature. Calcium Chloride - The concentration of this solution must remain constant to ensure that the beads of agar and amylase set properly. This solution should also be refreshed each time the experiment is carried out. Volume of enzyme (free and immobilised) -must remain constant, or the rate of reaction times will vary. Buffer Solution - should not be varied within each buffer test, as the rate of reaction times will vary greatly with each buffer. Substrate concentration - the rate of reaction for the enzyme will increase as the substrate concentration increases, so therefore, the concentration of the starch molecules in solution must remain constant. Sodium Alginate - the amount drawn up into the syringe must remain 3 cm3 throughout the experiment, or bead formation, size, and rate of reaction will be different. ...read more.


In beaker A the immobilized enzymes gave a slower reaction time than the free enzymes in beaker B. Evaluation The reliability of the results depends on a number of factors. Firstly, the accuracy of the experimental equipment must be taken into account. The scales used were very old, and therefore, could have a lesser degree of accuracy than what is needed for the experiment. Therefore, the solutions of starch and amylase, may have had different concentrations as they were made up, which would increase the rate of reaction. sodium alginate, and the free enzyme amylase had to be mixed in the 5cm syringes, it was impossible to get the solution thoroughly mixed, and therefore, some beads contained more free enzyme than others. This would also vary the time taken for the starch to be broken down into maltose. Some of the immobilised amylase beads had tails, this was unavoidable, although they were released further from the surface of the calcium chloride, they still developed tails, due to the pressure of the syringe being inadequate. The room temperature varied throughout the experiment, as it was done over several days, and some days were hotter than others. The effect of temperature on enzymes, can increase or decrease the rate of reaction, depending on how hot the temperature is. As the temperature never went above 40 degrees, (15 degrees, or lower) the rate of reaction didn't decrease due to this factor. However, the reaction rate may have varied slightly with increase or decrease in temperature, either slowing it down, or making it faster. Laura Munslow Page 1 Human Biology ...read more.

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