Oxidase Enzyme Lab

And finally into test tube 3, I pipetted 1.0 ml turnip extract and 4.0 ml of water. The contents of test tube 1 was poured into a spectrometer tube and labeled it “B” for blank. “B” tube was now inserted it into the spectrometer. An adjustment to the control knob was made to zero the absorbance reading on the spectrometer since one cannot continue the experiment if the spectrometer is not zeroed. A combination of two people and a stop watch was now needed to not only record the time of the reaction, but to mix the reagents in a precise and accurate manner. As my partner recorded the time, I quickly poured tube 3 into tube 2. I then poured tube 2 into the experiment spectrometer tube labeled “E” and inserted it into the spectrometer. A partner then recorded the absorbance reading for every 20 seconds for a total of 120 seconds. After the experiment, a brown color in the tube should be observed to indicate the reaction was carried out. Using sterile techniques, any excess liquid left was disposed

In this lab or experiment, the aim was to determine the following factors of enzymes: (1) the effects of enzymes concentration the catalytic rate or the rate of the reaction, (2) the effects of pH on a particular enzyme, an enzyme known and referred throughout this experiment as ALP (alkaline phosphate enzyme) and lastly (3) the effects of various temperatures on the reaction or catalytic rate. Throughout the experiment 8 separate cuvettes and tubes are mixed with various solutions (labeled as tables 1,3 & 4 in the apparatus/materials sections of the lab) and tested for the effects of the factors mentioned above (concentration, pH and temperature). The tubes labeled 1-4 are tested for pH with pH paper and by spectrophotometer, cuvettes 1a-4a was tested for concentration and cuvettes labeled 1b-4b was tested for temperature in four different atmospheric conditions (4ºC, 23ºC, 32ºC and 60ºC) to see how the enzyme solution was affected by the various conditions. After carrying out the procedures the results showed that the experiment followed the theory for the most part, which is that all the factors work best at its optimum level. So, the optimum pH that the enzymes reacted at was a pH of 7 (neutral), the optimum temperature that the reactions occurs with the enzymes is a temperature of 4ºC or

The Effects of Varied Temperatures, pH Values, Enzyme Concentrations, and Substrate Concentrations on the Enzymatic Activity of Catecholase

Students will be observing normal catalase reaction, the effect of temperature on enzyme activity, and the effect of pH on enzyme activity in this experiment. The enzymes will all around perform better when exposed in room temperature than when it is exposed to hot and cold temperatures. This is based on the fact that the higher the temperature, the better the enzymes will perform, but as the temperature reaches a certain high degree, the enzymes will start to denature, or lose their function.

The role of an enzyme is to catalyse reactions within a cell. The enzyme present in a potato (Solanum Tuberosum) is catechol oxidase. In this experiment, the enzyme activity was tested under different temperature and pH conditions. The objective of this experiment was to determine the ideal conditions under which catechol oxidase catalyses reactions. In order to do this, catechol was catalyzed by catechol oxidase into benzoquinone at diverse temperatures and pH values. The enzyme was exposed to its new environment for 5 minutes before the absorbance of the catechol oxidase was measured at 420 nm using a spectrophotometer. The use of a spectrophotometer was crucial for the collection of data in this experiment. When exposed to hot and cold temperatures, some enzymes were found to denature causing the activity to decrease. Similarly, when the pH was too high or low, then the catechol oxidase enzyme experienced a significant decrease in activity. It can be concluded after completing this experiment that the optimal pH for catechol oxidase is 7 and that the prime temperature is 20º C. Due to the fact that the catechol oxidase was only tested under several different temperatures and pH values, it is always possible to get a more precise result by decreasing the increments between the test values. However, our experiment was able to produce accurate results as to the

Temperature can affect the reaction of catechol oxidase by speeding up or slowing down the reaction. I was able to see what happened to the absorbance after changing the temperature of the catechol oxidase solution. I did this by heating and cooling the solutions to measure the absorbances in hot, cold, warm, and room temperature. Then the data was compared to see how the temperature effected the solution. The catechol oxidase solutions reacted best in room temperature (twenty-three degrees Celsius) and the worst in the cold (zero degrees Celsius). I concluded that temperature really does affect the way catechol oxidase reacts.

Within the experiment, pure catechol was mixed with different concentrations of catechol oxidase and the rate at which each solution produced benzoquinone was measured. The amount of benzoquinone made throughout the trials was measured by using a colorimeter to measure the level of “brownness” of the liquid. The colorimeter worked by shining a light through the liquid and then measuring that light on the other side to see how much of it was absorbed. In this experiment, absorbance of blue light was measured because blue light is absorbed by the color brown. The amount of blue light absorbance was measured every 15 seconds for five minutes. Because enzymes speed up reactions, more enzymes would cause the reaction to be even faster.1

In order to see the effects of pH and temperature on the enzymatic reaction of catechol oxidase when separated from potato tissue. We used a spectrophotometer to measure how much blue light energy is absorbed by benzoquinone. Benzoquinone is a product of catechol when it has been oxidized by different temperatures and pHs. We hypothesized that the benzoquinone absorbance rate would be faster when the pH added to the cuvettes were greater than the pH of the potato tissue. The pH of the potato tissue was pH 6. Our results show that pH 7 had the faster absorbance rate, slightly slower at pH 4, and slowest at pH

Catechol, in the presence of oxygen is oxidized by catechol oxidase to form benzoquinone (Harel et al., 1964). Bananas and potatoes contain catechol oxidase that acts on catechol which is initially colorless and converts it to brown (Harel et al., 1964). In this experiment, the effect of pH on the activity of catechol oxidase was conducted using buffers ranging from pH2 to pH10. Two trials were conducted due to the first trial results being altered by an external factor. The results were acquired by taking readings every 2 minutes for 20 minutes from a spectrophotometer and then recorded on to the table. The data collected in the table were then made into graphs to illustrate the influence of pH on the catechol oxidase catalyzed reaction. After analysis, the data revealed that pH did have a significant influence on the enzyme as recorded by absorbance per minute. However, the data was collected was not accurate due to external factors, thus the results are debatable and should be experimented again for validation.

The purpose of this experiment is to learn the effects of a certain enzyme (Peroxidase) concentration, to figure out the temperature and pH effects on Peroxidase activity and the effect of an inhibitor. The procedure includes using pH5, H202, Enzyme Extract, and Guaiacol and calibrating a spectrophotometer to determine the effect of enzyme concentration. As the experiment continues, the same reagents are used with the spectrophotometer to determine the temperature and pH effects on Peroxidase activity. Lastly, to determine the effect of an inhibitor on Peroxidase, an inhibitor is added to the extract. It was found that an increase in enzyme concentration also caused an increase in the reaction rate. The reaction rate of peroxidase increases at 40oC. Peroxidase performed the best under pH5 and declined as it became more basic. The inhibitor (Hydroxy-lamine) caused a decline in the reaction rate. The significance of this experiment is to find the optimal living conditions for Peroxidase. This enzyme is vital because it gets rid of hydrogen peroxide, which is toxic to living environments.

An isomer of catechol hydroxyquinone was used to determine the specificity of an enzyme called catecholase. Second experiment was performed to determine the effect of temperature on an enzyme. Enzyme catecholase was exposed to different temperatures. The third experiment was

The wavelength was set to 540 nanometers. It was set to 540 nanometers in order to produce a green light that would be absorbed by the benzoquinone for the reasons previously stated. There were five different tube samples that each held a total of 5mL of liquid. 1 mL of enzyme and 2 mL of EDTA were added to Tube 1. 1 mL of enzyme and 2 mL of PTU were added to Tube 2. 1 mL of enzyme and 2 mL of Citric Acid were added to Tube 3. 1 mL of enzyme and 2 mL of dH2O were added to Tube 4. Tube 4 was the control tube considering it contained no chelating agents.

Enzymes are high molecular weight molecules and are proteins in nature. Enzymes work as catalysts in biochemical reactions in living organisms. Enzyme Catecholase is found on in plants, animals as well as fungi and is responsible for the darkening of different fruits. In most cases enzymatic activities are influenced by a number of factors, among them is temperature, PH, enzyme concentration as well as substrate concentration (Silverthorn, 2004). In this experiment enzyme catecholase was used to investigate the effects of PH and enzyme concentration on it rate of reaction. A pH buffer was used to control the PH, potato juice was used as the substrate and water was used as a solvent.

After measuring equal amounts of distilled water and either adding or subtracting catechol which we referred to as the substrate some reactions was seen immediately. After which we were able to get data that supported my original hypothesis that in the addition of substrate and an enzyme the reaction would be present in varying degrees dependent on whether a temperature change was provided or not. In the second part of the experiment we were testing the inhibition action of Catechol Oxidase at different levels in several tubes of varying samples of potato extract, phenylthiourea (PTU) and distilled water. The experiment showed that (PTU) bonded with the extract and the water causing a reaction whereas there was no reaction in tube # 1 where there was an equal amount of everything in the tube. And test tube # 3 was the control tube where the (PTU) was eliminated as to observe if there was any reaction at all. Of course with the whole experiment we had to be very careful as to add the catechol last to ensure no premature reaction. It was hypothesized that (PTU) is a non -competitive inhibitor and doubling the substrate will have no reversal effect.

This experiment is designed to analyze how the enzyme catalase activity is affected by the pH levels. The experiment has also been designed to outline all of the directions and the ways by which the observation can be made clearly and accurately. Yeast, will be used as the enzyme and hydrogen peroxide will be used as a substrate. This experiment will be used to determine the effects of the concentration of the hydrogen peroxide versus the rate of reaction of the enzyme catalase.