Starch Hydrolysis by Amylase

1379 Words Jan 31st, 2014 6 Pages
Experiment 2: Starch Hydrolysis by Amylase

Theoretical Background
Polymers of carbohydrates are called polysaccharides, and make up some of the most important naturally occurring compounds [1]. They have thousands of monosaccharide units linked to each other by oxygen bridges. They include starch, glycogen, and cellulose, all three of which yield only glucose when completely hydrolyzed [2].

A

B

Figure 1. Starch (amylose) (A) and cellulose (B)

Starch occurs naturally in plants, which use it to storage glucose units for energy. It is often found in seeds and tubers (e.g., potatoes). It consists of two kinds of polymers of glucose. The simpler kind is called amylose, and it makes up about
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Figure 2: Schematic diagram of the components of a spectrophotometer.
The arrows indicate the pathway of light [7]

Transmittance is the ratio of the transmitted light energy (I) to the incident light energy (I0); percent transmittance is 100X ratio (I/I0). Transmittance, however, is not proportional to solute concentration, so it is usually converted into absorbance which is proportional to solute concentration. Digital spectrophotometers have readouts for both percent transmittance and absorbance, but the absorbance is measured always [7].

% T = (I / I0) x 100
ABS. = log 10 (100 / % T)

APPARATUS
Equipment
1. Test tubes
2. Pipettes
3. Pasteur pipettes
4. Beaker, 250 mL
5. Test tube rack
6. Plastic cuvettes

Buffers and Chemicals
1. Human salivary enzyme
2. Starch solution, 20 g/L
3. HCl stopping solution, 0.1N HCl
4. Iodine reagent stoch solution (in aqueous solution)
Iodine: 5 g/L ; KI: 50 g/L; Dilute to 1:100
5. Potassium phosphate buffers
KH2PO4 (monobasic phosphate) (MW=136.1 g/mol)
K2HPO4.3H2O (dibasic phosphate) (MW=228.23 g/mol)

PROCEDURE
Preparation of 20 g/L starch solution
1. Mix 20 g of soluble potato starch in approx. 50 mL of cold water.
2. While stirring, add the slurry to approx. 900 mL of gently boiling water in a large beaker.
3. Mix well and cool the gelatinized starch solution to room temperature.
4. Add more water to bring
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