DEMOTICA, Kin P.

Date performed: January 31, 2018

Chemistry 160.1 Section 6L Date submitted: February 7, 2018

EXERCISE 2. pH and Buffer Systems

Post-Laboratory Report

I. DATA

Table 2.1. Effect of concentration of buffer on buffering capacity.

Actual pH

Buffer used Concentration Before addition After addition of

ΔpH
(M) of NaOH NaOH

0.005 4.7 10.8 6.1

Acetate buffer 0.05 4.5 4.7 0.2

0.1 4.5 4.6 0.1

0.005 7.5 10.7 6.1

Phosphate
0.05 6.9 7.0 0.1
buffer
0.1 6.8 6.9 0.1

Table 2.2. Effect of adding NaOH to buffer with different pH

Actual pH

Test tube no. [A-]/[HA] Calculated Before addi- After addition ΔpH
pH tion of NaOH of NaOH

1 0.089/1 6.2 5.8 6.2 0.4

Phosphate 2 0.49/1 7.2 6.8 6.9 0.1

buffer
3 0.9071/1 8.2 7.7 8.0 0.3

4 0.08/1 3.7 3.6 3.9 0.3

Acetate 5 0.48/1 4.7 4.6 4.8 0.2

buffer 6 0.90/1 5.7 5.6 7.6 1.1

Table 2.3. Effect of adding HCl to buffer with different pH
Actual pH

Test tube no. [A-]/[HA] Calculated Before addi- After addition ΔpH
pH tion of HCl of HCl

1 0.089/1 6.2 5.9 2.3 -3.6

Phosphate 2 0.49/1 7.2 6.7 2.4 -4.3

buffer
3 0.9071/1 8.2 7.7 4.2 -3.5

4 0.08/1 3.7 3.6 2.1 -1.5

Acetate 5 0.48/1 4.7 4.6 4.2 -0.4

buffer 6 0.90/1 5.7 5.6 5.0 -0.6

Table 2.4. Titration of unknown amino acid.

mL KOH pH mL KOH pH
0.0 1.5 8.5 2.1
0.5 1.4 9.0 2.2
1.0 1.5 9.5 2.4
1.5 1.5 10.0 2.6
2.0 1.5 10.5 3.1
2.5 1.5 11.0 8.1
3.0 1.5 11.5 9.8
3.5 1.6 12.0 10.5
4.0 1.6 12.5 11.2
4.5 1.6 13.0 11.5
5.0 1.6 13.5 11.7
5.5 1.6 14.0 11.8
6.0 1.7 14.5 11.9
6.5 1.9 15.0 12.0
7.0 1.9
7.5 2.0
8.0 2.0

II. CALCULATIONS

To prepare the phosphate buffer with different molarities for the experiment, the following
calculations were done.
 !.!!# % ('# ())
For 0.005 M, V1 = = 1.25 mL of 0.1M phosphate buffer
!.+ %

!.# % ('# ())

For 0.5 M, V1 = = 12.5 mL of 0.1M phosphate buffer
!.+ %

!.+! % ('#())
For 0.10 M, V1 = = 25 mL of 0.1M phosphate buffer
!.+ %
III. Discussion

A buffer system is a solution composed of a weak acid or weak base and its conjugate base or
conjugate acid, respectively, and has the ability to resist drastic changes in its pH level when
limited amounts of base or acid is added to it (Brown, 2005). According to Brown, applications of
buffer systems in nature involve regulation of pH in living systems, while in a laboratory, they
are used to create environments with stable pH.

These changes in the pH is minimized due to the equal proportion of the acid and its conjugate
base or vice versa. For example, when H+ is added to the buffer, the weak base component of
the buffer reacts with the ion to form the acid component of the buffer. Whereas when OH- is
added, the weak acid react with it to form the base component. Unless large quantities of H+ and
OH- are added, the concentration of the base and acid do not change, thus, changes in pH is
negligible (Bettelheim, 2007).

For this laboratory exercise, two buffers with varying pH levels were studied: acetate buffer,
composed of CH3COOH and CH3COO-, with a pH of 4.7 and phosphate buffer, composed of
HPO42- and H2PO4-, with a pH of 7.2.

Upon addition of small amount of acid (H3O+):

H3O+ + HPO42- H2O + H2PO4- H3O+ + CH3COO- H2O + CH3COOH

Upon the addition of small amount of base (OH-):

OH- + H2PO4- H2O + H2PO4- OH- + CH3COO- H2O + CH3COOH

The first experiment done on these buffer systems was to test their buffering capacity with
respect to their concentrations in molarity. As shown in Table 2.1, buffers with a highest
concentration (0.10 M) resisted pH change most effectively, with respect to the ones with lower
concentrations. This is expected since it is known that as buffer concentrations increase, its
buffering capacity, or the ability to resist pH change, also increases (Brown, 2005).

Along with the effect of buffer concentrations, the effect of the addition of strong base and
strong acid to the buffer was also tested. When a strong base such as NaOH is added, the OH-
ions is expected to react with the acid component of the buffer, thus favoring a forward reaction.
In our data as seen in Table 2.2, the buffer that produced the least change was test tube 2. In
theory, a buffer with equal concentrations of acid and base components is more efficient
maintaining its pH. In other words, the more equal the concentrations of the acid and base in a
buffer, the more its buffering capacity is (Brown, 2005). However, during our experiment, this
was not the case, which may have been due to some errors.

As long as the concentration of the weak acid is higher than the conjugate base, the pH in the
buffer will be maintained since the OH- ions from NaOH interact with the H+ ions to form water.
Thus, any change in pH is minimized due to the neutralization process. However, when the
conjugate base concentration is higher than the weak acid, buffer capacity is lowered since
there is less H+ to neutralize the additional OH- ions from NaOH (Bettelheim, 2007).

On the other hand, the addition of a strong acid to the buffer donates protons to the conjugate
base. If the concentration of the conjugate base in the buffer is higher than the weak acid, then
the H+ ions from HCl will be neutralized effectively, thus, minimizing changes in pH. This results
to a higher buffer capacity. However, when there are higher weak acid concentrations, the
additional H+ ions from HCl will interact with the small amount of conjugate base and water
molecules to form more H3O+, thus, dramatically decreasing the pH. This is supported by the
data in Table 2.3.

In addition to testing the strength of buffering capacity, the experiment also involved the titration
of an amino acid with 0.1 M KOH. KOH was used for practical reasons such as its cheaper cost
compared to NaOH. Before the titration, the pH of the amino acid solution was first adjusted to
1.5. This is to make sure that the amino acid will go through its first ionic form in pKa1 where pH
is low.

14.0

12.0 pKa2

10.0

8.0
pH

6.0 IpH

4.0

2.0
pKa1
0.0
0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0 16.0
mL KOH

Figure 2.1. Titration curve of an unknown amino acid sample.

Based on the graph, pKa1 is 2.2 and pKa2 is 11.2. Theoretically, the buffer action of this amino
acid is observable within the ranges of 1.2 – 3.2 pH and 10.2 and 12.2 pH. The IpH was
computed using the average of the pKa1 and pKa2 giving the value of 6.7. The amino acid during
this point exists as a zwitterion, with both positive and negative charges. Thus, at this point, any
addition of strong acid or base can react with it by either receiving a proton or donating a proton
(Bettelheim, 2007). The identity of the unknown acid could be serine based on the Pka1 alone.
The pKa2 is difficult to determine since the graph is not similar to a normal titration curve. This
error can be attributed to the wrong handling and usage of the pH meter.

Living organisms rely on buffers to maintain their homeostasis, providing optimum environments
and pH levels for various biochemical processes inside the cell (Berg et al., 2006). A small
change in the pH level of the body of an organism can tip the balance of the organism and can
even be fatal. In practical applications, on the other hand, scientists use buffers to create stable
aqueous solutions with optimum pH levels. They use buffers to study biomolecules and mimic
the natural environment they are in so they can be observed in their natural behavior.
REFERENCE:

Berg, J., Tymoczko, J., Stryer, L. and Gumport, R. (2006). Biochemistry. 5th ed. Basingstoke:

W.H.Freeman & Co Ltd.

Bettelheim, F. (2007). Introduction to General, Organic and Biochemistry. Brooks/Cole by

Thomson Learning

Brown, C. (2005). Introduction to Biochemistry. McGraw-Hill nternational.

Ionization of Amino Acids. (n.d.). Retrieved February 12, 2015, from Chemistry UIUC:
http://butane.chem.uiuc.edu/cyerkes/Chem104ACSpring2009/Genchemref/Ionization_AA.htm