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DETERMINATION OF THE VALUES

OF AN AMINO ACID
Experiment 2
Ebru AKHARMAN, Gebze Technical University, Turkey
AIM:
1. To determine the titration curve for an amino acid.

2. To use this curve to estimate the values of the ionizable groups of the amino acid.

3. To understand the acid base behaviour of an amino acid.

In this experiment, unknown amino acid is identified via an acid base titration.

INTRODUCTION: MATERIAL AND METHODS:


Playing an important role in the structure meter
and function of proteins, amino acids are
perhaps one of the most important Unknown amino acid
biomolecules as proteins themselves NaOH
participate in nearly every physiological
event with in a cell. In order to understand Beaker
the acid base properties of proteins and Distilled water
their resulting behaviour as polyionic
macromolecules with each containing at 10 ml of one of the unknown amino acid
least one amino and carboxylic acid group, solutions are placed into a 250 ml beaker.
amino acids are classified as amphoteric This solutions is titrated with 0,1 M NaOH.
substances and react with both acids and of unknown amino acid solution is
bases. Amino acids are weak polyprotic
measured before adding NaOH. 1,0 ml NaOH
acids. All of the amino acids have an acidic
group (COOH) and a basic group (NH2 ) is added in the solution. After each addition,
attached to the carbon, and also they beaker is mixed and measured the .
contain ionizable groups that act as weak
acids or bases, giving off or taking an The titration is continued until the
protons when the is altered. The at reaches 11. After each measurement, data is
which the net charge of an amino acid is recorded.
zero is called the isoelectric point, or the .
The isoelectric point is always half way RESULTS:
between the values. The Formula for
is: The results obtained in the experiment is
+ + tabulated. The result is placed on the graph
= and this application helps to find unknown

amino acid.
: nth value
Graph 1: Values of pH graph

If we want to found unknown amino acid.

When the concentration of the


unprotonated form equals that af the
unprotonated form, the ratio of their
concentrations equals 1, and log 1 = 0.
Hence, can be defined as the at
which the concentrations of the protonated
and unprotonated forms of a particular
ionizable species are equal.

The also equals the at which the


ionizable group is at its best buffering
capacity, that is the at which the
solution resists changes in most
Table 1: Valus of pH table effectively.

If you want to show this data graphically:


Graph 2: Calculate of pI graph

[ ]
= + log
[]

= []. Thus,
[ ]
=1
[]
log 1 = 0
=
1 + 2
=
2
2,61 + 10,34
=
2
= 6, 48
DISCUSSION: RESOURCES:
When testing, it is important to know the http://vlab.amrita.edu/?sub=3&brch=63&si
amount of test material. In this experiment, m=1336&cnt=1
the pKa of an unknown amino acid was
calculated and the identity of this amino https://www.chem.fsu.edu/chemlab/bch40
acid was estimated. Before describing this 53l/Protein%20Characterization/AA%20Ti
process, I would like to talk about the tration/index.html
properties of amino acidsAll of the amino
acids have an acidic group (COOH) and a
basic group (2 ) attached to the carbon,
and also they contain ionizable groups that
act as weak acids or bases, giving off or
taking an protons when the pH is altered.
The pH at which the net charge of an amino
acid is zero is called the isoelectric point, or
the pI. The isoelectric point is always half
way between the values.The amino
acid is estimated by utilizing the pI value. In
this experiment NaOH is added until the
unknown amino acid is pH 11. After each
addition of NaOH, the pH is measured and
the data recorded. Graphics are generated
from the obtained data. The pi is then
calculated using the Formula:

+ +
=

The amino acid is estimated from the pI


values table. The obtained value of 6.48 is
close to the proline.