Temerature and Ph Factors That Affect the Enzymatic Activity of Salivary Amylase

Onion DNA Synthesis

Gaduena, Kristine Joy B., Garong, Maria Ana Therese DR., Guballa, Paulyn Aura Mae V.*, Iglesia, Christina Bien Paula DJ.

3PSY4, Department of Psychology, College of Science, UST, Manila

paulynguballa@gmail.com

Abstract

DNA, also known as deoxyribonucleic acid is one of the essential biochemical molecules of an organism that serves as the repository of genes. The objectives of the experiment are to isolate DNA from the onion sample, to subject it to acid hydrolysis and to perform qualitative color reactions. DNA was isolated from onion by disrupting its cell membrane and other cellular components. From the experiment,the isolate was found to have a thread- like structure.. Spectroscopy was applied to test the purity of the DNA sample. In the experiment, the DNA isolate had an absorbance of .607/ .282 which indicates that the isolate was pure. The sample was subjected to acid hydrolysis and qualitative color reaction test that includes test for deoxyribose, test for phosphate, test for pyrimidine and test for purines. In the experiment, the hydrolysate produced dark brown and colorless layer which is a positive result for test for deoxyribose, clear colorless solution with yellow precipitate for test for phosphate which is also a positive result, and yellow residue for test for purine which indicates the presence of adenine however it produced a white cloudy solution for test for pyrimidines which is a negative result.

Keywords: Salivary amylase, enzymatic activity, pH, temperature, optimum activity

Introduction

Methodology

        For the temperature, 2 mL of the enzyme solution was put in a large test tube. This was labelled as 4°C. In a separate test tube, 2 ml of the buffered starch solution was added. Both test tubes were then incubated for ten minutes in an ice bath (4°C). The solution was mixed immediately after, then three drops of the mixture were quickly taken and two drops of the iodine solution were simultaneously added onto a spot plate. This was the zero minute. After a minute of interval, three drops were again taken of the mixture and two drops of the iodine solution were simultaneously added onto the second well. This was the one minute. It was repeated until a light yellow-colored solution was observed. The time was noted. For the other temperatures, the steps were repeated following the desired incubation temperature. The reciprocal of time (, min-1) was plotted versus the temperature (T). The optimum temperature of the amylase was determined.[pic 1]

        For the pH, 1 mL of acetate buffer (pH 4) was mixed with 1 mL of 2% unbuffered starch in a large test tube. In a separate large test tube, 2 mL of the enzyme solution was added. Both test tubes were then incubated for ten minutes in a 37°C water bath. The solution was immediately mixed and three drops of the mixture were quickly taken and simultaneously, two drops of the iodine solution were added onto a spot plate (first well). This was the zero minute. The step was repeated until a light yellow-colored solution was observed. The time was noted. For the other pH, the steps were repeated using the appropriate buffer. Then, the reciprocal of time (, min-1) was plotted versus the buffer pH. The optimum pH of the amylase was determined.[pic 2]