Succinate Dehydrogenase Lab Report

Understanding the activity of enzymes in different muscle types can aid greatly in obtaining more information about other processes such as metabolism of the tissues (Anderson et al., 2012). There are many different methods in order to achieve this information based in two different major categories but the most convenient method is one called continuous assay. This process includes the use of a spectrophotometer to continuously monitor the assay. This method allows for an easy way to calculate the initial rate of reaction as one can establish time points easily. In the lab performed the continuous method was used in order to determine the measurement of the activity enzyme succinate dehydrogenase (SDH). SDH is most commonly found in mitochondria. It is present in many different muscle tissues including the heart, red, and white tissue. In the following lab the enzyme was tested and measured in these three muscle types in the Oncorhynchus mykiss in order to determine which type contained the highest and the lowest activity. This enzyme is involved in multiple processes such as involved in both the Krebs cycle and the electron transport chain (ETC) (Smith, 2014). Its role in

Succinate dehydrogenase is an enzyme found in the mitochondrial inner membrane. The enzyme catalyzes the reaction of oxidizing its substrate, succinate, into fumarate via the removal of hydrogen ions from succinate. This oxidation is vital in the Krebs cycle.

The preparation for the experiment started by gathering the solutions of enzyme Peroxidase, substrate hydrogen peroxide, the indicator guaiacol and distilled water. Two small spectrometer tubes and three large test tubes with numbered labels. In addition, one test tube rack, one pipet pump and a box of kimwipes were also gathered. Before the experiment, the spectrometer must be set up to use by flipping the power switch to on. Following, the machine was warmed up for 10 minutes and the filter lever was moved to the left. In addition, I set the wavelength to 500 nm with the wavelength control knob. Before the experiment, I had to create the blank solution by pipetting 0.1 ml of guaiacol, 1.0 ml of turnip extract and 8.9 ml water into tube #1. Following the creation of the blank, a control 2% solution was created.

We used TLC analysis to identify each product obtained from the dihydroxylation reactions by spotting a TLC plate with the product of our reaction, a solution of cis-cyclohexane, trans-cyclohexane, and a 50:50 mixture of the two. We then placed the plate in a beaker with ethyl acetate saturating the atmosphere to allow the TLC plate to develop. Finally, we compared Rf values of the components of the mobile phase, after the phase was completed. 100% ethyl acetate was used instead of 100% Hexane or a mixture of Ethyl Acetate, because ethyl acetate has high polarity and can separate the components of a mixture to elution, unlike hexane, which is non-polar, and therefore unable to separate the components of the mixture. A 50:50 mixture of both would not work, because the polar and non-polar compounds would neutralize the mixture, and thereby not separate the components of the mixture.

The optimum pH level would be pH 7. This is because this is where the highest amount of enzyme activity is taking place.

2. We measured 1 mL of turnip peroxidase (the enzyme) and 3 mL of neutral buffer (pH corresponding to the test tube number i.e. pH 5 in test tube 5) with a syringe and disposed it into tubes 3, 5, 6, 7, 8, and 10

In this lab we experimented with pH, spectrophotometry, and enzymes like catecholase. We used spectrophotometry to detect how enzyme activity would change under different pH, temperature, enzyme concentration, and substrate concentration. Enzymes are important for biochemical reactions because they speed up the process and allow the organism to continue living. The enzyme used in the experiment is catecholase and it catalyzes a reaction between catechol and water. We analyzed the samples in the spectrophotometry and it is measured by absorbance and transmittance and this will allow us to see which sample will have the largest concentration of molecules. The first experiment involved enzyme concentration and enzyme activity; in this experiment

In this laboratory exercise, studies of enzyme catalase, which accelerates the breakdown of hydrogen peroxide into water and oxygen. The purpose was to isolate catalase from starch and measure the rate of activity under different conditions. The laboratory was also conducted in association with a second laboratory that measured the effects of an inhibitor on the enzymes.

Enzymes are catalysts that function to speed up reactions; for example, the enzyme sucrose speeds up the hydrolysis of sucrose, which breaks down into glucose and fructose. They speed up reactions but are not consumed by the reaction that is taking place. The most important of the enzyme is the shape as it determines which type of reaction the enzyme speeds up. Enzymes work by passing/lowering and energy barrier and in doing so; they need to bind to substrates via the active. Once they do, the reaction speeds up so much more quickly than it would without the enzyme. Coenzymes and cofactors aid the enzyme when it comes to binding with the substrate. They change the shape of the active site so the substrate can bind properly and perform its function.

In the experiment we used Turnip, Hydrogen Peroxide, Distilled Water, and Guaiacol as my substances. On the first activity, Effect of Enzyme concentration of Reaction Rate for low enzyme concentration, we tested three concentrations of the turnip extract, and hydrogen peroxide. For the Turnip Extract I used 0.5 ml, 1.0 ml, and 2.0 ml. For hydrogen peroxide we used 0.1 ml, 0.2 ml, and 0.4 ml. We used a control to see the standard, and used a control for each enzyme concentration used. The control contains turnip extract and the color reagent, Guaiacol. We prepared my substrate tubes separately from the enzyme tubes. My substrate tube

reaction rate increases. If the temperature of an enzyme gets to high the reaction rate will slow

Organisms cannot depend solely on spontaneous reactions for the production of materials because they occur slowly and are not responsive to the organism's needs (Martineau, Dean, et al, Laboratory Manual, 43). In order to speed up the reaction process, cells use enzymes as biological catalysts. Enzymes are able to speed up the reaction through lowering activation energy. Additionally, enzymes facilitate reactions without being consumed (manual,43). Each enzyme acts on a specific molecule or set of molecules referred to as the enzyme's substrate and the results of this reaction are called products (manual 43). As a result, enzymes promote a reaction so that substrates are converted into products on a faster pace (manual 43). Most enzymes are proteins whose structure is determined by its sequence of its amino acids. Enzymes are designed to function the best under physiological conditions of PH and temperature. Any change of these variables that change the conformation of the enzyme will destroy or enhance enzyme activity(manual, 43).

Cell fractionation is a very important procedure in cell biology and can be very useful for studying different organelles. By fractionating, we mean separating or dividing the cell into different component parts.

The purpose of this lab experiment was to determine the relationship between temperature and the rate of enzymatic activity in yeast cells. In the lab, the temperature was the independent variable. The temperatures consisted of 6°C, 24°C, 34°C, 46°C. The dependent variable in the experiment was the rate of enzymatic activity in yeast cells. The temperatures were tested by using a LabQuest and pressure probe that tested pressure inside the plastic test tube. LabQuest graphed the data and created a line of best fit that was used to determine the slope of the graph. The slope of the graph represents the rate of enzymatic activity. The slope was found for each temperature in 2 different trials. Then, the rate of enzyme activity (kPa/sec) for each temperature in the 2 trials were averaged. These averages were used to develop a graph that shows the relationship between temperature and the rate of enzyme activity. According to the results of the experiment, as the temperature increases, the rate of enzymatic activity decreases. Each enzyme has its own optimal temperature in which it can function efficiently.

We could go even depeer in the explanation of metabolism reaction but this is just an