Alp Enzyme Lab Report

1270 Words 5 Pages
Introduction
This experiment looks at the effects of pH, enzyme concentration, and temperature of the catalytic rate of different enzymes including alkaline phosphatase (ALP). Some of the enzymes used are involved in human digestion while others are involved in clinical tests for certain diseases (Wilson, D’Alessio, Schmitt Lavin, Keith, & BIOL 1500 Team, 2015). The ALP enzyme is used to diagnose diseases such as bone cancer, jaundice and cirrhosis of the liver. This enzyme also catalyzes the hydrolysis of a phosphate group connected to lipids in vivo or to proteins (Wilson et al., 2015).
There are three major reasons that enzymes are important. The first reason is that enzymes speed up chemical reactions, and every chemical reaction
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Enzymes have optimal temperature, and a majority of enzymes have an optimal temperature between twenty and forty degrees Celsius. Once the temperature is too high, enzymes will denature, but before the enzymes denature, the rate of reaction doubles for every ten degrees Celsius increase in temperature for most enzymes (Wilson et al., 2015). Hₒ: The temperature of the solution will not affect the rate of reaction. Hₐ: If the temperature is low, the rate of reaction will not be fast.
Materials and Methods
The first step of part one of the experiment is to prepare solution E by adding 6.5 mL of solution B, which is para nitro phenolphosphate (pNPP), with 6.5 mL of distilled water (Wilson et al., 2015). Next, four cuvettes need to be labeled and prepared according to Table 1. Solution D, which is a high concentration of the ALP enzyme, is not to be added yet. A strip of broad-range pH paper is to be placed on a clean and dry watch glass, and place a drop of each solution on the pH paper immediately after mixing the solution. The pH of each is then recorded in Table 2 (Wilson et al., 2015).
Table 1: Mixing Instructions for pH
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This will set the absorbance at zero for the solution being used. Solution D is not to be added until the readings in the spectrophotometer are going to be done. To mix in solution D, cover the cuvette with parafilm and invert to mix. After the solution is mixed, place the cuvette into the spectrophotometer. Record in table 2 the absorbance readings every minute for five minutes (Wilson et al., 2015). The second and third parts will be prepared together. Solution A is an alkaline buffer, and solution B is a substrate that contains .003 M disodium p-nitrophenyl phosphate (dNPP). 15 mL of both solution A and solution B need to be mixed to form solution F (Wilson et al., 2015). The eight cuvettes need to be labeled at the top with 1a, 1b, 2a, 2b, 3a, 3b, 4a, and 4b. Now, the blank cuvette 1a needs to be prepared with the solutions specified in Table 3. The prepared 1a cuvette is used to 0 the spectrophotometer a 405 nm, and make sure not to touch the blank button again. Next, 3 mL of solution F needs to be added to each of the cuvettes. To prepare for the temperature experiment place cuvette 1b in a 4°C refrigerator, leave 2b at room temperature, place 3b in a 32°C water bath, and place 4b in a 60°C water bath. Before beginning the next step, make sure the data is ready to be recorded, and make sure to have a small square parafilm ready (Wilson et al.,

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