Titration of Amino Acids Lab Con
Transcript of Titration of Amino Acids Lab Con
Titration of Amino Acids
Introduction
Materials0.1 N NaOH0.1 N HCl0.1 M glycine solution0.1 M lysine solution0.1 M aspartic acid solutionNeutralized formaldehyde
Procedures1. Take two pipettes and fill the first with 0.1N
HCl and the second with 0.1N NaOH2. Into each of the two beakers, introduce 10.0
mL of the amino acid solution and add 10mL of distilled water and measure the resulting pH of the solution
3. Titrate the first solution with 0.1N HCl adding 2.0 mL at a time and determining the pH after each addition, until a total of 10.0 mL is reached (for glycine and aspartic acid) or 20.0 mL (for lysine). In addition, measure the pH at 5mL and 15mL volumes
Procedures4. Titrate the second solution in the same manner
as the 1st using instead, 0.1N NaOH, until 10.0 mL is reached (for glycine and lysine) or 20.0 mL is reached (for aspartic acid)
5. Plot the pH (ordinate) vs. The equivalent acid/base (abscissa). One mL of acid/base = 0.1 mEq of acid/base. (Show how this value was obtained).
6. Repeat the above procedure, but add 5.0mL of neutralized formaldehyde solution into each of the amino acid solutions before determining the pH of the solution. Titrate the solutions.
Procedures
7. Plot pH vs. the equivalent acid/base on the same graph as above. Construct your titration curves on graphing paper. Solve for the pI and pK values of your amino acid.
8. Record your results as follows
Glycine: Acid VS Base Data
HCl Gly w/o HCHO
HCl Gly w/ HCHO
NaOH Gly w/o HCHO
NaOH Gly w/ HCHO
0 5.15 5.02 6.91 4.57
2 2.96 2.97 9.28 6.06
4 2.62 2.56 9.69 6.57
5 2.52 2.43 9.75 6.92
6 2.35 2.33 9.98 7.21
8 2.13 2.12 10.26 7.84
10 1.99 1.99 10.58 8.88
Glycine: Acid VS Base Graph
0 2 4 5 6 8 100
2
4
6
8
10
12
HCl Gly w/o HCHO HCl Gly w/ HCHO NaOH Gly w/o HCHO NaOH Gly w/ HCHO
Aspartic Acid: Acid VS Base DataHCl Asp w/o
HCHOHCl Asp w/
HCHONaOH Asp w/o HCHO
NaOH Asp w/ HCHO
0 4.47 4.49 4.5 4.52 8.62 7.12 4.01 4.014 9.66 8.2 3.64 3.645 9.96 8.53 3.47 3.646 10.18 8.72 3.3 3.478 10.52 9.16 2.95 3.11
10 10.9 9.63 2.59 2.7512 11.43 10.34 2.32 2.4514 12.16 11.32 2.12 2.2215 12.37 11.58 2.04 2.1116 12.51 11.78 1.96 2.0318 12.68 11.99 1.84 1.9120 12.8 12.14 1.76 1.81
Aspartic Acid: Acid VS Base Graph
0 2 4 5 6 8 10 12 14 15 16 18 200
2
4
6
8
10
12
14
HCl Asp w/o HCHO HCl Asp w/ HCHO NaOH Asp w/o HCHO NaOH Asp w/ HCHO
Lycine: Acid VS Base DataHCl Lys w/o
HCHOHCl Lys w/
HCHONaOH Lys w/o HCHO
NaOH Lys w/ HCHO
0 8.79 6.25 9.04 6.362 7 5.5 9.54 6.764 2.55 3 10 7.165 2.3 2.58 10.3 7.386 2.14 2.32 10.58 7.668 1.94 2.04 11.03 8.36
10 1.81 1.87 11.42 9.3512 1.67 1.7414 1.59 1.6515 1.55 1.6116 1.53 1.5718 1.47 1.5220 1.42 1.47
Aspartic Acid: Acid VS Base Graph
0 2 4 5 6 8 10 12 14 15 16 18 200
2
4
6
8
10
12
HCl Lys w/o HCHO HCl Lys w/ HCHO NaOH Lys w/o HCHO NaOH Lys w/ HCHO
Questions
Question1. What can account for
Sorensen’s discovery that the endpoint of titration between an amino acid and a standard alkali (without formaldehyde) is not reached?
AnswerFormaldehyde in excess readily
combines with free unprotonated amino groups of amino acids to give dimethylol derivatives, and the proton titrated can be treated directly. Formaldehyde makes the solution more acidic when titrated with a standard base
Question2. Compare the values obtained
when the amino acid was titrated with HCl both in the absence and presence of formaldehyde. How do you account for this?
AnswerIn the presence of formaldehyde,
titration curve was slightly lower. This is due to the fact that formaldehyde ties down the amino groups, making the carboxyl hydrogen ion more available
Question3. At which pH will an amino acid
exert its maximum buffering capacity? Why? Where in your graph is the buffering region for your amino acid?
AnswerAn amino acid exerts its
maximum buffering capacity at the plateau near or on its pKa values because it is in this area that there is equal concentration of proton donors and acceptors.
Question4. From the titration curve of an
amino acid, can you determine the nature of its R group i.e., basic, acidic or neutral?
AnswerYes, the nature of the R group
can be determined from the titration of an amino acid through the determination of the pKa values for each dissociable group of an amino acid by extrapolating the midpoint of each buffering region or plateau within the curve.
AnswerSince the ionic form of the amino
acid present in an aqueous solution is dependent upon solutions pH, if:pH > pKa: basic (negatively charged)pH < pKa: acidic (positively charged)pH = pKa: zwitterion (no charge)
isoelectric point