Titration and Buffers Lab
Write a brief objective for this experiment in your own words.
The amino acids A and B used in this lab are from the following list: glycine, aspartic acid, glutamic acid and lysine. Look up the structure of each of these in the Biochemistry text.
Identify the R group (the variable part) of each and draw the structural formulae (acidic, basic and zwitterionic) of each as the titration proceeds. Note that some of the amino acids will have three forms and others will have four.
Prepare a graph of your results, plotting mL of NaOH versus pH. Plot each of the two titrations (not the dry run) using standard graph paper or Excel. Remember to label the graph axes and provide enough tick marks along each axis to be useful. It is easy to forget these steps when using software; write the information in by hand, if necessary.
From a titration curve it is easy to discern the inflection point (pI). This is the point when 1 equivalent of base has been added. In this experiment you use 10 mL of 0.1 M amino acid or 1 millimole of amino acid (fully protonated). As stated in the introduction, the completely protonated amino acid can donate two protons during the titration.
It will take one equivalent (1 millimole) of base to titrate the first proton (on the acid group) and another equivalent to titrate the second proton (on the amine group). Find the pI and label it on each graph.
Note- An equivalent refers to the moles of OH- or H+ added with each addition of NaOH. It is determined by multiplying the volume of NaOH added by the concentration of the NaOH.
Since the pKacid is the midpoint on the first leg, you can find the pKacid at 0.5 equivalents of base. Likewise, the pKamine can be found at 1.5 equivalents of base. Find and label the pKacid and pKamine for each graph (if possible).
Calculate the pI by finding the average between the average pKacid value and the average pKamine value. This should be a better method of determining the pI of the amino acid.
Identify your amino acid from the following list: glycine, aspartic acid, glutamic acid and lysine. Look up the “true” pI of the amino acid from a reliable source (list your source) and calculate a percent error between your pI value and the “true” pI value.
Write down the final pH of your buffer measured by pH-meter.
Does the pH determined by Henderson-Hasslebach equation method agree with the desired pH of 7.5? If not, can you explain why?
The final pH of the buffer measured by the pH meter was 6.5. The pH of 6.5 does not agree with the desired pH of 7.5. It appears that our group’s calculations were off by a little and this could have been due to weighing out the incorrect amount of buffer.