Enzymes are also sensitive to pH most enzymes perform optimally between pH of 6-8, pH breaks ionic and hydrogen bonds denaturing the enzyme. Concentration of enzyme to substrate and substrate to enzyme either speeds up or slows down a reaction. If there is more enzyme than substrate then the substrate will limit the reaction, if there is more substrate than enzyme the enzyme will limit the reaction (Todd & Gomez, 2001). Alteration of the active site causes the reaction to either slow down or speed up. Alteration of temperature, pH, and concentration will not cause a change in enzyme activity.
Changes in temperature, pH and concentration will have an effect on enzyme activity by feeding up or slowing down the reactions. Materials and Method Part 1 of the experiment was the effect of temperature on enzyme activity. This was carried out using 7 test tubes, a phosphate buffer (pH 7), potato juice, and catechist. Also an ice water bath and at 100 C, and a hot water bath at 500 C were used to show how temperature changes the speed of enzyme reaction. A spectrophotometer was used to show the changes by testing the light absorbency of the solutions.
Three test tubes were filled with 3 ml of pH 7 phosphate buffer; one was placed in the ice bath, one room temperature (ICC), ND the other in the hot water bath. Two test tubes were filled with 3 ml of potato juice one was placed in the ICC ice bath the other was placed in the ICC hot water bath. Then two more tubes were filled with 3 ml of catechist, one was placed in the ice water bath the other in the hot water bath. After 10 minutes 10 drops of catechist and 10 drops of potato juice were added to the buffers of corresponding temperatures. The tubes stood for 5 minutes mixing periodically during this time.
Then color change was observed. Part 2 of the experiment was the effect of pH on enzyme activity. There were 7 test tubes used in this part. Also phosphate buffers 4,5,6,7,8,9,and 10 were individually added to the corresponding tube. Each tube was filled with 3 ml of the buffer and then 10 drops of catechist and 10 drops of potato juice were added. Tubes stood for 5 minutes and were inverted at 1 minute intervals. The color change was observed in each tube. Part 3 was the effect of enzyme concentration on enzyme activity. This part required 4 test tubes labeled A, B, C, and D.
Tube A was filled with 3 ml + 20 drops of pH 7 buffer, tube B received 3 ml + 15 drops of pH 7 buffer and drops of potato juice, tube C had 3 ml + 10 drops of buffer and 10 drops of potato juice, and tube D had 3 ml of buffer and 20 drops of potato juice. Then 10 drops of catechist were added to each tube and the stood for 4 minutes mixing the contents at 1 minute intervals. Color change was then observed. Part 4 was the effect of substrate concentration on enzyme activity. Eight test tubes were labels with numbers corresponding to the table, Tube 1 was filled with 5 ml + 47 drops of pH 7 buffer and 1 drop of catechist.
Tube 2 received 5 ml + 46 drops of buffer and 2 drops of catechist. Tube 4 had 5 ml + 44 drops of buffer and 4 drops of catechist. Tube 8 had 5 ml +40 drops of buffer and 8 drops of catechist. Tube 16 had 5 ml + 32 drops of buffer and 16 drops of catechist. Tube 24 had 5 ml + 24 drops of buffer and 24 drops of catechist. Tube 32 had 5 ml + 16 drops of buffer and 32 drops of catechist. Tube 48 had 5 ml of buffer and 48 drops of catechist. Then 30 drops of potato juice was added to all 8 tubes. The tubes stood for 5 minutes and were inverted to mix at 1 minute intervals. The color change was then observed.
Results Part 1 resulted in the room temperature tube having the highest amount of reduce produced which means it had the fastest reaction. We know this because the color was the darkest in this tube. The ICC tube had the slowest reaction because it was the lightest color and the ICC tube had a reaction that was in between the other two speeds. Part 2 resulted in the pH 7 tube having the most product formed and there for the fastest reaction. The following list is in order from the fastest reaction (most product formed) to the slowest reaction (least product formed): pH 8, pH 9, pH 6, pH 5 all the way to no color change in pH 4 and pH 10.