pH Measurement and Buffer Preparation

J.M. Matila, R.M. Mayor, P.M. Mojica, F.M. Pasion, C. Navarro

Group 5, 2F-PH, Faculty of Pharmacy, UST, Manila

ABSTRACT
A buffer is an aqueous solution containing a weak acid and its conjugate base or a weak base and its conjugate
acid. The objectives of the experiment were to prepare the assigned phosphate buffer solutions with desired pH
and concentrations, to determine samples colorimetrically using liquid indicators and electromagnetically using the
pH meter. The amount of the buffer components was computed using the Henderson-Hasselbach Equation. A
volume of 500 mL of 6M HCl and 6M NaOH were prepared to adjust the buffer solution with a concentration of
0.50 M to its desired pH of 7.0. The sample which is Sprite, with pH 3.4, was subjected to colorimetric
determination giving the result colors of light yellow for thymol blue, violet for bromophenol blue, blue for
bromocresol green, gray for bromocresol purple, yellow for phenol red, light orange for methyl red, orange for
methyl orange, and colorless for phenolphthalein.

INTRODUCTION
Studying and understanding the essence of buffer functioning in optimal conditions. When our body is
capacity and pH measurement is very intrinsic. Few in equilibrium, our multiple self-regulating control
people did know that buffer capacity and pH helps mechanisms are able to operate in ideal conditions
maintain the equilibrium of some enzymatic to efficiently deliver nutrients to our cells and to
activities inside our body. On top of that, what is a dissolve and eliminate waste products through the
buffer? A buffer is a solution that consists of either a bodys natural elimination channels. Over
weak acid-conjugate base combination or weak acidification of the body, on the other hand, is a
base-conjugate acid combination that resists dangerous condition that weakens all body systems,
sudden change in the pH whenever small amounts of and can give rise to an internal environment
acid or base is added. A pH is the measure of the conducive to disease and even death. One of the
hydrogen ion concentration in a solution. Solutions bodys pH control mechanisms is called the acid-base
with a high concentration of hydrogen ions have a buffer system. In this experiment, we will further
low pH and solutions with a low concentration of H+ understand the importance and the essence buffers
ions have a high pH. This may seem like a confusion and its other benefits in human body.
way to express these relationships. It has a
logarithmic nature whereas pH is expressed as the METHODOLOGY
negative logarithm of the hydrogen ion Preparation of Buffers
concentration of a particular solution. On the other
hand, a solutions buffer capacity is the ability of a
buffer solution to resist change. It is the strength of a
buffer to resist any changes brought by an acid or
base that is being introduced to the buffer solution
What makes our knowledge about buffer solution
important is because, buffer solution greatly affects
how our body is going to function. We have to
understand that the human body is designed to
maintain a very subtle pH balance in its fluid, tissue
and its system. As most biochemical reactions
essential to life take place in an aqueous
environment, however, it is our blood plasma and
interstitial fluids surrounding the cells that are most
sensitive to acid-alkaline imbalance. Whereas these
fluids are needed and must be maintained in a
narrow range of pH. The physiological pH is 7.35 and
7.45. In this pH our bodys immune system is
Figure 1.1 Weighing of reagents
The assigned buffer for the group was the phosphate
buffer. To prepare this, 6.9 grams of sodium
orthophosphate and 5 grams of sodium phosphate
anhydrous dibasic were weighed using a triple beam
balance.
Figure 1.2 Dissolving of components
After being weighed, each of the components was
put separately into erlenmeyer flasks both filled with
50mL of distilled water. These were both dissolved
by shaking until no solid components were to be
found in the solution.
Figure 1.2 Mixing of buffer components
After dissolving, the 50mL solution of distilled water
and sodium orthophosphate was mixed with the
50mL solution of distilled water and sodium
phosphate anhydrous basic.
Electrometric Determination of pH
For the electrometric determination of pH, the pH
meter was calibrated first to pH 7 by submerging it
to distilled water which is known to have the said
pH. After this, the pH of 20-mL of the following
samples was measured: prepared buffer solution
and assigned sample (Sprite).
Figure 2.1 Measuring pH of assigned solution
To do this, the pH meter was submerged to the
beaker with 20-mL of Sprite, which was the assigned
solution to the group, and the final reading was
recorded.

Figure 2.2 Measuring and adjusting pH of buffer
With the buffer solution, it first read a pH of 4.4 and
to adjust this to the desired pH of 7.00 portions of
a basic solution 6.0M NaOH which was prepared
beforehand was added to the buffer solution and
each time, the pH was checked until pH 7 was
reached. A total of 4 mL of NaOH was added to
attain the said pH.
Colorimetric Determination of pH
Figure 3.1 Labelling of acid-base indicators
For the colorimetric determination of pH, test tubes
were firstly prepared and labelled with the acid-base
indicators: thymol blue, bromophenol blue,
bromocresol green, bromocresol blue, bromocresol
purple, phenolphthalein, methyl orange, and methyl
red.
Figure 3.2 Mixing of Acid-Base Indicators and Buffer
Solution
5-mL of the buffer prepared was added using a
pipette to 2 drops of each acid-base indicator
already filled into the test tubes. The mixture was
mixed vigorously using a vortex mixer and the final
resulting colors of each of the solution were
recorded. This was also done using the assigned
sample to the group and the results were recorded,
as well.
RESULTS AND DISCUSSION
1. Preparation of the Buffer
Each group was assigned a buffer to
prepare, and the group was tasked to phosphate
buffer with pH of 7.0. Buffers are aqueous systems
that tend to resist changes in their pH when small
amounts of acid (H+) or base (OH-) are added [1].
= + log[ ]
7 = 7.21 + log[ ]
7 7.21 = log[ ]
7 7.21
 0.617 3. Colorimetric determination of pH
=
1 Table 2: Colorimetric Test for the Buffers
Acid-Base pH pH pH pH pH pH pH Buffer
0.617 + 1 = 1.617() Sprite
Indicators 2.0 3.0 5.0 7.0 8.0 10.0 12.0 prepared
Thymol blue SP Y LY LY CL LB LB Y LY
0.50 Bromophenol blue Y G BV V Pu LV V Y V
0.25 = 0.125
1 Bromocresol
Y Y B B B SB Pu Y B
green
[A] Bromocresol
Y Y Y Gr PU PU Y Y GR
0.617 1 1000 purple
0.125 = 95.39 Phenol red Y LY Y Y P P O Y Y
1.617 0.50 1
Methyl red P P P LO LY Y B R LO
[HA] Methyl orange R R O O O O P CL O
Phenolphthalein CL CL CL CL CL DP FP P CL
pH
1 1 1000 Y-Yellow, G-Green, R-Red, P-Pink, B-Blue, V-Violet, O-
0.125 = 154.61
1.617 0.50 1 Orange, CL- Colorless, SP-Salmon Pink, FP- Fuchsia
Pink, SB- Sky Blue
2. Electrometric Determination of pH
Acid-Base Indicators (also known as pH
Table 1.a: pH of Assigned Sample indicators) are substances which change color with
Using the pH meter, this measures the pH. They are usually weak acids or bases. Indicators
Sample pH [H+] are used in titration solutions to signal the
3.98x10-4 completion of the acid-base reaction. [2]
1. Sprite 3.4
M Thymol blue has the range of 1.2 - 2.8 and in this
2. Sting 5.01x10-4 range, Red is the color below the pH range but in the
3.3
(energy drink) M
experiment at pH of 2.0 it is Salmon Pink and Yellow
2.51x10-4 when it is above the pH range.
3. Green tea 3.6
M
7.94x10-4 Bromophenol blue has the pH range of 3.0 4.6
4. Apple juice 3.1
M and if the pH is below the range, the color is yellow
electrochemical properties of liquids, pastes and and blue violet if its above the range. Bromocresol
semi-solids. The sample Sprite has a pH of 3.4, the green has the pH range of 3.8 5.4 and the color
group computed for the concentration of the [H+] below the pH range is Yellow and if above the pH
ion using the formula: pH= - log[H+] ; [H+]= antilog(- range is Blue this is showed from pH of 3.0 it is
pH). yellow and in pH of 5.0 the color changes to blue.
Bromocresol purple has the pH range of 5.2 6.8.
Table 1.b: pH of Buffer solution
The color Yellow indicates that the pH is below the
The pH meter showed an accurate reading for the
Samples pH [H+]
buffer solution prepared by the group. The
Distilled Water 5 1x10-5 M
phosphate buffer prepared had pH of 6.7 so the
Buffer Prepared 7 1x10-7 M
group decided to add a base, 6M NaOH solution.
range and Purple or violet indicates a higher pH
Gradually adding the base into solution, the pH
value.
meter reading showed fluctuations even with the
slightest addition of NaOH. Adding NaOH into the Phenol red has the pH range of 6.8 8.4. In pH
buffer will increase the pH, but if HCl is added the pH of 7.0 the color is still yellow this is maybe because it
will decrease, the phosphate buffer prepared is in the range and have higher pH than the lower
needed an increase thats why the group chose to value of the range which is 6.8. The color changes to
add the NaOH or the base. pink in pH of 8.0, if the pH is above the range, it will
be red but since the pH of 8.0 is still within the range
it appeared pinkish in color.
Methyl red has the pH range of 4.8 6.0, if the
pH is below the pH range it is color Pink, while if its
above the range it is color Yellow. Methyl orange
have the pH range of 3.2 4.4, if the pH is below the
range the color is Red and if its above the range, the
color is Orange.
Phenolphthalein is a colorless, weak acid which
dissociates in water forming magenta anions. The pH
range is 8.2 10.0, thats why from the pH of 8.0 to
pH of 2.0 it is colorless.
The phosphate buffer with pH of 7.0 prepared
by the group is approximately within those acid-base
indicators which has pH range of 6.8 8.4.
Reference:
[1] Lehninger A.L., Nelson D.L.,and Cox, M.M. (1993). Principles of Biochemistry 2nd ed. New York: Worth Publishers
[2] Retrieved from: (chemistry.bd.psu.edu, 2017),
CONCLUSION
In conclusion, buffers are aqueous systems that tend
to resist changes in their pH when a small amount of
acid or base is added. The number of moles of strong
acid or strong base needed to change the pH of 1
liter of buffer by one unit is called the buffer
capacity. The larger the buffer capacity, the more
resistant the buffer is to changes in pH. The most
effective pH range for any buffer is at or near the pH
where the acid and salt concentrations are equal
(that is pKa). For the maximum buffer capacity, pKa=
pH 1. The pH meter showed accurate pH readings
of the prepared buffer solution. It showed
fluctuations in readings with the slightest addition of
HCl and NaOH. In the experiment, the group
achieved to do the objectives that were given.