CHM 215

BIOCHEMISTRY LABORATORY

Experiment 2

AMINO ACID TITRATION

SUBMITTED BY ŞEVVAL SARGIN

Course instructor: Dr. ESRA AYDEMİR

Submitted to: ŞEVVAL SARGIN

EXPERIMENT DATE: 30.11.2020

SUBMISSION DATE: 02.12.2020

Biruni University
Istanbul
2020
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TABLE OF CONTENTS

1. OBJECTIVE........................................................................................................................3
2. THEORY.............................................................................................................................3
3. APPARATUS......................................................................................................................5
4. PROCEDURE.....................................................................................................................6
5. RESULTS............................................................................................................................7
6. DISCUSSION......................................................................................................................8

REFERENCES......................................................................................................................10
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1. OBJECTIVE

Glycine is an apolar amino acid that is structurally the simplest of the 20 amino acids found in
proteins. Its side chain consists of only one hydrogen atom. The aim of this experiment is to
measure the pKa value of the ionized group, to find the buffer regions of the amino acid and
to learn the information between the net charge of the amino acid and the pH of the solution.

2. THEORY

2.1. Amino Acid Titration

Amino acids, building blocks of peptides, proteins and enzymes in living organisms
They are of particular importance among various chemical substance groups.
Having both amine group (-NH2) and carboxylic acid group (-COOH) in its molecule
organic compounds are called amino acids. An amino acid generally R (NH2) CH-COOH can
be shown as [1]. Gradually adding protons to the environment or is the removal. At very low
pH the dominant glycine ion is complete as it is protonated. When the pH is increased slowly,
carboxyl first group loses protons. The proton donor at the midpoint of the first stage and
zwiterion is present in equal amounts. The pH of the midpoint of the titration is protonated is
equal to the pKa value of the group. pK1 (pKa value of the carboxyl group of glycine) =2.34
The second important point in titration is the first proton
the loss is completed and the second one begins it is at this point that glycine is largely
dipos. pH = pI. Stage 3 is the amino group proton of glycine loses. PK2, which is the
midpoint of this phase (pKa value of the amino group of glycine) = 9.6 [2]

2.2. Titration Of Glycine

Glycine consists of an amine and a single carbon attached to the carboxyl group. The small
size helps it to function as a flexible linkage signal in proteins and enables the formation of
coils, recognition sites in cell membranes and enzymes, a modifier of molecular activity and
an osmoprotective formation by conjugation of hormone precursors and glycine elongation.
[3] Purpose: To identify unknown amino acids by determining the pKa values from the acidic
and basic properties of amino acids with a titration curve.
The titration curve is determined by observing the pH value by adding acid or alkali to a
certain volume of sample. Glycine has two buffering ranges. These are shown as rectangular
areas with a pH range of +/- 1 centered at pH 2.34 and pH 9.6.Glycine can be used as a buffer
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solution in these pH ranges. Glycine pI: 5.97. So, the titration curve is determined by adding
acid or alkali to a sample of a certain volume and observing the pH value.

3. APPARATUS

3.1. Equipment
 Micropipette
 Micropipette Tips
 Beaker
 pH meter
 Spatula
 Measuring Tube
 Burette
 Sensitive Scales
 Distilled Water
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3.2. Chemicals
 NaOH
 HCL

3.3. Samples
 Glycine

4. PROCEDURE

First of all we weigh 0.1 M 50 ml (0.375 grams) of Glycine and pour it into a beaker. Then
we add 50 ml of distilled water to the beaker containing Glycine. Then we mix the solution
well. We label Glycine as 0.1 M 50 milliliters. pH meter we wash it each time and use it to
calculate the pH of the Glycine solution, but we are very careful. And we add small amounts
of HCl into the beaker using a micropipette to monitor the pH drop. We repeat this process
until we reach a pH of 1.55. Starting with a low pH Glycine level, we add 0.5 M 50 ml of
NaOH into the burette and drop 1 ml of NaOH into the beaker which is completely white
acidic Glycine. We make sure to take note of the pH levels for and according to every other
milliliter dripping. Use 26 ml to chart.

5. RESULTS

By adding HCL and NaOH, we changed the pH value on the experiment.

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0 pH(1,55)
1 mL 1,81 14 mL 8,4
2 mL 1,93 15 mL 8,96
3 mL 2 16mL 9,09
4 mL 2,05 17 mL 9,32
5 mL 2,2 18 mL 9,47
6 mL 2,3 19 mL 9,62
7 mL 2,4 20 mL 9,72
8 mL 2,52 21mL 9,94
9 mL 2,72 22mL 10,2
10mL 2,86 23mL 10,54
11 mL 3,15 24mL 11,2
12 mL 3,52 25mL 11,6
13 mL 4

6. DISCUSSION

Every time we use the pH meter, we must wash it and clean it well. Any chemical residue left
on it may interfere with the measurements made at the end of the experiment. This may cause
us to move away from the data we normally need to access. Also, attention should be paid to
the buffer zones of glycine. Pl value must be calculated correctly. Because this allows to
create an accurate chart.

REFERENCES

[1] DOĞAN, A. (2001). Amino Asitlerin Mikroskopik Denge Sabitleriyle İlgili

Çalışmalar. Gazi Üniversitesi Gazi Eğitim Fakültesi Dergisi, 21(2).

[2] file:///C:/Users/Hp/Downloads/BiyokimyaI-4.pdf ( YTU / Yrd.Doç.Dr. Emel Ordu)

[3] Hall, JC (1998). Glisin. Parenteral ve Enteral Beslenme Dergisi , 22 (6), 393-398.