paraphrase all graphs
Hasanlari
biolabdiscuusion.docxIntroduction
The subject of the experiment circled around enzymes and to explain the methods taken and my hypothesis, essential background material like the fact that an enzymes are organic catalysts. Enzymes play a major role in the progression of a biological reaction. With the presence of enzymes, a speed in a reaction is noticed and this is due to the fact that the components of an enzyme are made up of large globular proteins. The way an enzyme works is that it effects Affects the Ea, activation energy, by lowering it and resulting into an increase in the reaction itself, and can be observed by a standard curve. To follow the steps taken in this experiment we take in count the fact that in an enzyme reaction the elements that are taken in consideration are the enzyme itself, which is a complex structure made through the folding of polypeptide chains, the second component are the substrates involved, and the third are the products formed, which in this experiment are P-Nitrophenol Phosphate. To test the different variations our experiments would result in, we use tools like spectrophotometer for the results of optical density. The observation of the optical density increasing if the addition of P-Nitrophenol to increase the solution’s concentration, is a question addressed in this experiment. The hypothesis formed for this experiment is that the reaction with the presence of an enzyme will occur in a faster rate than a reaction that has no enzyme present. This hypothesis was tested through out a series of tests and graphs that are plotted after the data was collected using a timely mannered experiment with the technique of diluting buffers, serums and solutions.
Materials and Methods (protocol)
In this experiment, a detailed protocol was followed referring to the lab manual that included the materials for part B that was a standard curve and we used the buffer that was 10.0 in pH and 0.1 M (Sodium carbonate-bicarbonate buffer) and 0.005mM (P-Nitrophenol standard) was used and to conduct this part the spectrophotometer was used along with 6 test tubes.
In parts C, D and E the materials used were the 0.1M, pH 10.0 buffer that was Sodium carbonate-bicarbonate buffer, the solution was P-Nitrophenyl phosphate substrate solution, the serum with alkaline phosphate, and used to conduct these parts was a spectrophotometer. In part C water was part of the materials with 4 test tubes. Part D needed 12 test tubes and part E was preformed with 7 test tubes.
To preform this experiment a prepared written protocol was followed with the steps of the methods that were unique to each part. For part B the end product we wanted was to obtain the data for creating a standard curve, this was done by preparing 5 dilutions (0.0025mM, 0.005mM, 0.01mM, 0.025mM and 0.05mM) that would result in the standard curve from the optical densities from solutions consisting of the buffer (Sodium carbonate-bicarbonate buffer) and the P-Nitrophenol standard.
In the following part of the lab, part C, we use water for sample 1 and serum in sample 2. This part is where we test the enzyme effect on the reaction rate. 0.4mL of water is used in sample 1B and sample 2B contains 0.4mL of serum to see the enzyme alkaline phosphate vs. no enzyme effect.
For part D we increased the enzyme by 0.1mL, 0.2mL, 0.4mL, 0.6mL, 0.8mL and 1.0mL and changed the volumes of the enzymes. The result observed in this part is the effect volume change had on an enzyme.
The last part in this experiment concluded in part E changed concentrations of substrates by 1.0mL, 2.0mL, 2.6mL, 2.8mL, 3.2mL and 3.6mL with the enzyme being constant at a volume of 0.4mL.
Discussion
a) paragraph 1 – The objective of this experiment is to test and prove the effects of enzyme on a reaction and if the pace will change in result of the presence of enzymes in that reaction. This is conducted by a series of tests and we observe is the increasing of substrate levels have a direct effect on the reaction’s rate or of the enzyme concentrations’ variation will effect the speed of the reaction. The effect of alkaline phosphate enzyme is acknowledged by the results and data collected from this lab. After arranging the data into tables and further expanding this comparison into graphs we visually proved the hypothesis that were in the introduction true. Increasing P-Nitrophenol increased the absorbance (O.D.) which proved part B and the standard volume dilutions increasing statement was shown in the graph. Part C’s other graph showed clear increase with the 2nd sample’s presence of enzyme and the 1st sample had no enzyme hence progressing in a slower rate. Our original graph does proof this too, a slight increase is shown for sample 2, yet for errors discussed in paragraph 5 our data is not for referring to in this analysis. As we can see in part D (other) graph the optical density shows increase as the enzyme volume increases as well. So in part E when adding the amount of substrate to the same amount of enzyme would result in a faster reaction.
b) paragraph 2 – The methods used in this experiment provided the data we needed to further determine the validity of our hypothesis. One tool that was an essential factor is the spectrophotometer. All the optical density values in this lab report was obtained through the readings of the spectrophotometer. These optical densities were how we determined if the enzyme was effecting Affecting the rate of the reaction. Through those optical density values the graphs could be plotted in a time, concentration fashion for viable data. As to how useful this method was, there was some limitations. An example is that the fact that the spectrophotometer is the same one that obtained all the results for the continuous parts of this experiment, so any fluctuating of this tool caused the parts to have somewhat not 100% accurate results.
c) paragraph 3 – As stated in paragraph a, our hypothesis was met and in this paragraph a closer look at why that is right will be further analyzed. In the results of part C you can see the increase of the reaction with the presence of an enzyme and the slow pace in comparison to the rate of reaction with no enzyme. For the reaction with the enzyme the reaction barely progressed as to the reaction with enzyme the values were increasing forming a linear line in the graph. The values for sample 1 would increase 0.01 and return to its initial value decreasing that 0.01 units through the reading of optical density (O.D.). This resulted in the plotting of a straight line almost parallel with the x axis. In sample 2 an increase of 0.018 occurred immediately and just increased after that. This is due to the presence of an enzyme, speeding up the reaction rate. The comparison of the two samples could be made by the first time measurement that was clear that the sample with enzyme started with a higher value that the sample without enzyme.
Adding enzyme concentrations added the values of O.D. was what was conducted in part D. A reading of 0.050 of sample 2 where 0.2 of serum was added and a reading of 0.068 of sample 3 where 0.4 of serum was added, and both of these readings at the initial. The jump is shown with this data and the enzyme seems to be doing it’s speeding up from the start. This correlates with the hypothesis where the increase of an enzyme volume (mL) will result in the increase of the rate of the reaction within the ten-minute period.
Relating the substrate with the enzyme is what was preformed in part E. This complex resulted in the increasing optical density graph. This is expressed when looking at the data of the 6 test tubes, where test tube 1 containing 1.0mL of substrate had a 0.013 at initial and a test tube like 4 that had 2.8mL of substrate formed a reading of 0.033. The graph agrees with the original hypothesis.
e) paragraph 5 – After each graph that was not viable to use for this experimental lab report I put in a paragraph expressing the errors that occurred in this laboratory. The most noticeable error that affected our lab report data was the readings of the spectrophotometer. It’s two knobs was mixed up with and that resulted in a glitch in our results, although all the other steps from the prepared protocol was followed precisely. The column for mixing in our tables made that clear, yet the wiping of the cuvette with the constant back and forth and the quick addition of enzyme so it will not run longer that the calculated stop watch may not have been conducted every time. The blanking of the spectrophotometer was established and the steps that followed that was surely accurate though. The other error with the spectrophotometer is that it would from reading to reading fluctuate and that would have caused an increase or decrease in our data results therefore effecting the slope of our graphs.
f) paragraph 6 – The relevance of my results in this experiment is at a major factor here. Enzymes as shown in my discussion is a crucial substance to the biochemical reactions that occur in our human bodies. Enzymes work they way they do because of the fact that they lower the Ea, activation energy, in a reaction for the least amount of energy to be exerted. So this regulates our body’s energy so we don’t have to use up high energy in our bodies without the presence of enzymes. Another factor enzyme help regulating the human body with is the time it would take without an enzyme. A helpful contribute enzymes preform in our bodies is the activation of functions and even the inhibition of functions through the allosteric function an enzyme has. Regulating the human body is also done with the feedback inhibition an enzyme prevents.
