pH Measurement and Buffer Preparation

Jyle Fraium L. Dino​, Franc Ison P. Dy, Demi Louise L. Go

Vheverli Whendel P. Go, Marc Lennard G. Ip
Group 4 2A Medical Technology Biochemistry Laboratory

ABSTRACT
A 250 mL Acetate buffer of pH 4.7 with a molar concentration of 0.5 M was prepared using 7.20 mL of Glacial
CH3COOH and 2.5 g of NaOH. In order to obtain the desired pH 4.7 of the acetate buffer, either 6M HCl or 6M NaOH
were added while being monitored by a pH meter. Other samples such as distilled water and shampoo were also
subjected in Electrometric determination using a pH meter and gave pH readings of 7.362 and 3.841 respectively. The
samples, distilled water and acetate buffer, were subjected to color determination by referring to the table of the pH
ranges of various acid-base indicators. The distilled water gave the colors of yellow in thymol blue, violet in
bromophenol blue, blue in bromocresol green, violet in bromocresol purple, orange in phenol red, red in methyl red,
yellow in methyl orange, and colorless in phenolphthalein and indicating a pH range of 7.0 - 7.5. The distilled water
gave the colors of yellow in thymol blue, violet in bromophenol blue, green in bromocresol green, yellow in
bromocresol purple,yellow in phenol red, orange in methyl red, yellow in methyl orange, and colorless in
phenolphthalein and indicating a pH range of 5.0.

INTRODUCTION the buffer capacity of the prepared buffer

The hydrogen ion concentration of a solution is solutions.
referred to as the pH, which is potential of
Hydrogen. It is the measure of alkalinity or EXPERIMENTAL
acidity of a solution. The pH scale is a logarithmic A. Test Compound/s or (Sample/s) used
scale with values from 1 to 14, wherein the Sodium Hydroxide (NaOH) pellets, Glacial Acetic
increasing or decreasing of two values are by Acid (CH3COOH), Distilled H2O
factors of 10 [1]. Seven is the neutral point with B. Procedure
values below it being indicators of acidity and 1. Preparation of Acetate Buffer
values above the neutral point being indicators of In order to prepare 250 mL 0.5 M Acetate
basicity. The lower the pH value, the more acidic buffer, certain computations were made to
a solution is; the higher the pH value, the more acquire the necessary values for the preparation
alkaline it is. of the Acetate buffer. 2.5g of Sodium Hydroxide
Buffer solutions are aqueous solutions that (NaOH) pellets were obtained and weighed
resist changes in pH level when acids or alkali of through an analytical balance. It was placed
small amounts are added. They contain inside a beaker and after which, 7.20 mL of
components that remove any hydrogen or Glacial Acetic Acid (CH3COOH) was introduced.
hydroxide ions from the added amounts, and This was mixed until the NaOH pellets were
usually buffer solutions are composed of a weak dissolved fully and then transferred into a 250
acid and its conjugate base or vice versa; a weak mL volumetric flask. Subsequently, distilled water
base and its conjugate acid [2]. was added up to the 250 mL mark of the flask.
The Henderson-Hasselbach equation is used to Furthermore, it was stirred and mixed
relate the pH of a buffer solution of an acid to its completely.
acid dissociation constant (​pka​) as well as molar
concentrations. The equation is said to be valid Computations:
when it has the acid and conjugate base’s Number of Moles required:
equilibrium concentrations [3]. It is used to moles solute
M = L solution
compute for the pH of aqueous solutions.
The objectives of the experiment are to be able Moles​Buffer = (0.250L) x (0.5M) = 0.125
to prepare different buffer solutions, determine moles
the pH of buffers and samples colorimetrically
using liquid indicators and electrometrically using
the pH meter and lastly, to know how to calculate
Actual moles produced:
Henderson Hasselbach’s equation
 [conjugate base]
pH = pKA + log [acid]
Using a graduated cylinder, 120 mL of
conc. HCl was measured in order to prepare 250
4.7 = 4.7 - log [CH3COO−]
[CHCOOH] mL of 6.0 M HCl. This was then transferred and
[CH3COO−]
[CH3COOH]
= antilog (4.7 - 4.7) labelled appropriately.
[CH3COO−]
= 1
[CH3COOH] 1 Computations:
Total moles produced = 2 moles MxL=MxL
(12.2M) x (L) = (6.0M) (0.250L)
L conc HCl = 0.12 L or 120 mL
Mole adjustment:
Moles [CH3COO​-​] 3. Electrometric Determination of pH
1 mole
2 moles
= x
0.125 moles ​ he pH of the assigned prepared buffer was
T
​ .0625 moles
x =​ 0 measured using a pH meter. Before obtaining the
pH of the acetate buffer, the pH meter was first
Moles [CH3COOH] calibrated at pH 7 by immersing the electrode in
1 mole
= x distilled water. Afterwards, the electrode was
2 moles 0.125 moles
lifted and dried with a paper towel. The electrode
​ .0625 moles
x =​ 0 was then submerged in the acetate buffer. The
expected reading of the solution must be around
Glacial CH3COOH preparation: 4.7 for the acetate buffer. HCl or NaOH were
M = 17.4 M added in order to obtain the desired pH of the
Moles required = 0.125 moles acetate buffer.
L=x
M = moles solute
L solution
L solution = 0.125 moles
17.4 M
= 0.0718 L or 7.20
mL

NaOH preparation:
MW of NaOH = 40g/mol
Moles required = 0.0625 moles
Mass = x
mass (g)
Moles = M W
X = (0.0625 moles) x (40g/mol) = 2.5g
NaOH

Figure 2​. pH meter with the desired pH of the

acetate buffer

4. Colorimetric Determination of pH
​ he colors of various acid-base indicators at
T
certain pH were determined by referring to the
table of the pH ranges.

Figure 1. NaOH being mixed with Glacial Acetic RESULTS AND DISCUSSION
Acid
2. Preparation of Reagents
 Table 1. Results of the electrometric
1. Electrometric Determination of pH determination of pH

​ he pH meter is used to measure the

T
electrochemical properties of liquids, pastes, and Samples pH [ H+ ]
semi-solids. The instrument points out the
Distilled 7.362 4.35 × 10−8
hydrogen ion concentration in a test solution by Water
responding to the potential developed by an
electrical cell. The common pH meter consists of Assigned 3.841 1.44 × 10−4
electrodes, the glass and the reference. Sample
Fluctuations on readings with the slightest (shampoo)
addition of HCl and NaOH can be observed. It
Buffer 4.711 1.94 × 10−5
also displays sensitivity to a small amount of [ H +
prepared
] and [ OH − ] ions because of the electrode. An
example is when an addition of HCl will decrease
the pH, while an addition of NaOH will increase
2. Colorimetric Determination of pH
the pH of the buffer. As a result, the pH is more
accurate in reading pH levels compared to a pH
The colorimetric determination of pH showed
paper.
the varying color changes an acid-base indicator
undergoes when added to a solution of certain
The value of [ H + ]was computed by obtaining
pH. Therefore, this property of an acid-base
the antilog go the negative pH, 10−pH . For the
indicator can be utilized to identify different
acetate buffer, the pH was manipulated using
substances by narrowing their pH range. In
6.0M HCl solution and 6.0M NaOH dilution until
addition, acid-base indicators also show
the desired pH (pH 4.70) was obtained.
molecular characteristics of a substance such as
the color changes in molecules. This can be
A titration curve is a graphical representation
caused by changes in electron confinement; more
of a titration in which the volume of titrant is
confinement makes the light absorbed darker,
potted on X-axis and the pH of the solution is
while less confinement makes the light absorbed
plotted on the Y-axis. The base is added in small
lighter.
increments and the pH of the acid in each step is
determined with a pH meter. In the experiment,
the acetic acid, which is a weak acid, is titrated
with a strong base, NaOH. The NaOH is added as
small increments of equal volume and at each
step, the pH of the solution is measured. The
Table 2. Results of the colorimetric
graph reveals the pKa of the acid or the pH at
determination of pH
which the system consists of the proton donor
concentration and proton acceptor. This
Acid-base pH pH pH pH pH pH pH Distill Buffer
relationship between from the
indicator 2.0 3.0 5.0 7.0 7.5 8.0 12.0 ed Prepa
Henderson-Hasselbalch equation. Thus, a
H 2O red
titration curve also reveals the buffer action of
weak acid and its conjugate base. Thymol blue O Y Y Y Y B B Y Y
Bromophenol Y Y G G B B B G G
blue
Bromocresol Y Y G B B B B B G
green
Bromocresol Y Y Y V V V V V Y
purple
Phenol red Y Y Y O O R R O Y
Methyl red R R O Y Y Y Y Y O
Figure 3.​ Titration Curve of Acetic Acid Methyl R R O Y Y Y Y Y O
orange
phenolphthal C C C C C C P C C 3. Determination of Buffer Capacity
ein
pH 7.3 4.7
O=orange, Y=yellow, B=blue, G=green,
V=violet, R=red, C=colorless, P=pink
The experiment was focused on the Acetate
buffer with a ph of 4.7 or 5.
For thymol blue, the buffer indicated a yellow
coloration. Thymol blue has pH range of 1.2-2.8
at acid range. In this range orange is the color
below and yellow is color above. Thymol blue has
pH range of 8.0-9.6 at basic range. In this range
yellow below and blue is color above.
For bromophenol blue, the buffer indicated a
green coloration. Bromophenol has a pH range of
3.0-4.6 and yellow indicates it is below pH range
and green indicates above the pH range.
At bromocresol green, the buffer also indicates
a green coloration. Bromocresol green has pH
range of 3.8-5.4 wherein yellow indicates it is
below pH range and green indicates above the pH
range.
For bromocresol purple, the buffer indicates a
yellow coloration. Bromocresol purple has pH
range of 5.2-6.8. Yellow color indicates below pH
range and violet indicates above pH range. The
color of the buffer might have been affected by
other environmental factors hence turning pink to
violet.
For phenol red, the pH range is 6.8 to 8.2. The
acetic buffer showed a yellow color. Yellow for
the lower pH while red for the higher pH. Since
the acetic buffer has a pH of 4.70, it had a
yellow color. In distilled water which has a pH of
7.3, it produced an orange color.
The pH range of methyl red is 4.4 to 6.2. This
indicator produced an orange color in the acetic
buffer. A red color with methyl red indicator
signifies that it is low in pH while a yellow color
signifies that it is high in pH. In distilled water
which has a pH of 7.3, it produced a yellow color.
Methyl orange produced an orange color when
mixed on acetic buffer. Its pH range is 3.1 to 4.4.
The pH is said to be too low when it produces
red. The pH is said to be too high when it
produces a yellow color. In distilled water which
has a pH of 7.3, it produced a yellow color.
Phenolphthalein showed no color change until
the pH range is 8.3 to 10. This indicator produced
a colorless solution on both the acetic buffer and
distilled water.
Buffer capacity is used to measure the ability of
a solution to withstand changes in pH through
absorbing or desorbing H​+ or OH​- ions. Effects on
pH change varies upon adding an acid or a base
to a buffer system, depending on the initial pH
and the capacity of the buffer to resist change in
pH. Buffer capacity is the moles of an acid or
base needed to change the pH of a solution by
one unit, divided by the pH change and the
volume of buffer in liters. A buffer only causes
drastic change in the pH of a solution once a
complete reaction has occurred. The pH becomes
less resistant to change and causes a more
significant change in pH as the buffer is depleted.
In addition, buffer capacity is the ability of a
solution to resist change, which is dependent on
the ratio of the acid to the base and the
concentration of the buffer.
The following formula is used in calculating for
the buffer capacity:
REFERENCES
[1] Which of your friends needs to learn this
term? (n.d.). Retrieved from
http://www.businessdictionary.com/definition/pH
-scale.html
[2] Clark, J. (2002). Buffer Solutions. Retrieved
from
https://www.chemguide.co.uk/physical/acidbasee
qia/buffers.html
[3] Henderson-Hasselbalch equation and the pH
of buffer solution. (n.d.). Retrieved from
http://www.chembuddy.com/?left=pH-calculation
&right=pH-buffers-henderson-hasselbalch
[4] ​Easy Biology Class. (n.d.). Retrieved from
https://www.easybiologyclass.com/titration-curv
e-of-a-weak-acid-and-its-pka-biochemistry-notes
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