pH Measurement and Buffer Preparation
Laguitan, R., Lim, J., Luna, K., Magday, X., Manaois, K.
ABSTRACT
The objective of the experiment is to prepare a buffer solution, electrometrically
determine the pH level using the pH meter and subject the prepared buffer solution to
colorimetric determination using acid-base indicators. A 50 ml phosphate buffer solution
having a pH of 7.5 with a molar concentration of 0.50 M was prepared using 0.117
grams of primary sodium phosphate monohydrate NaH2PO4 and 0.4429 grams of
secondary sodium phosphate heptahydrate Na2HPO4. The pH level of the prepared
buffer solution was subjected through electrometric determination. Using a pH meter,
parts of either 6.0 M HCl or 6.0 M NaOH solution were added drop by drop adjusting
the buffer solution to its desired pH of 7.5. Colorimetric determination of pH was later
then done. Assigned samples were micellar water, distilled water, and the prepared
buffer solution. 200 μl of each corresponding sample was pipetted on eight spots of the
microplate. Different buffer solutions having a narrow range of pH values were
gathered as well. Drops of each acid-base indicators namely thymol blue, bromophenol
blue, bromocresol green, bromocresol purple, phenol red, methyl red, methyl orange,
and phenolphthalein were added in the microplate. Variation of colors were noticed.
The buffer prepared having a pH of 7.5 exhibited a white color with phenolphthalein,
yellow for methyl red, yellow for thymol blue, orange for methyl orange, blue for
bromocresol green, yellow for phenol red, blue violet for bromophenol blue, and blue
for bromocresol purple. Eventually, through colorimetric determination, the sample,
micellar water was found out to have a 6.0 pH level.
INTRODUCTION used to neutralize the excess hydrogen
(H+) and hydroxyl ions (OH-), thus
pH is a method of expressing the resisting the changes in pH of the
alkalinity (low hydrogen concentration) solution and helps to maintain a near pH
or the acidity (high hydrogen by adding it to the solution.
concentration) of a solution and was first
introduced by a Danish biochemist Søren The Henderson-Hasselbach
Peter Lauritz Sørensen in 1909. pH is a equation points the relationship of the
short term for "power of hydrogen" pH of a solution to the pK of an acid. It
wherein the "p" means potenz, a German shows the ratio of the concentration of
word for power and the H is the symbol the acid and its conjugate base that also
in the periodic table for hydrogen. determines if the aqueous solution used
is an effective buffer or not. The
On the other hand, a buffer equation provides an obsolete way to
system is a solution that is a mixture of a think about buffer solutions and pH:
weak acid and its conjugate base, or a pH = pKa + log ab
weak base and its conjugate acid. It is
In line with this, the students III. Colorimetric Determination of
prepared an H3PO4 buffer solution and pH
calibrated the pH of the buffer according
to the desired pH which is 7.5. The
assigned sample which was micellar
water was diluted with 20 mL distilled
water for accuracy.
Methodology
I. Preparation of Reagents
The reagents prepared were 500
mL of 6.0 M HCl and 6.0 M NaOH. For
6.0 M HCl, the computed volume of Image 2: Addition of different acid-base indicators to
concentrated HCl was diluted first. For each spot.
6.0 M NaOH, the computed amount were
dissolved in 300 mL of distilled water. Different microliters of primary
Both reagents were poured into separate phosphate groups were pipetted on eight
amber bottles and were labeled spots. Each spot were treated with the
accordingly. following chronologically:
Phenolphthalein, Methyl red, Thymol
II. Preparation of Buffers blue, Methyl orange, Bromocresol green,
NaH2PO4 and NaHPO4 were Phenol red, Bromophenol blue,
measured and placed in a beaker to be Bromocresol purple. Each spot is treated
dissolved in water. After completely with different acid-base indicators
dissolving, the pH was adjusted to the producing different colors depending on
desired pH by using the buffers prepared the reaction of the acid-base indicator.
beforehand. It was then transferred to a The colors were then noted.
50 mL volumetric flask and filled to
volume with water. Once mixed well, it Results
was transferred to a beaker and labeled
accordingly. The results gathered from the
experiment are tabulated below.
Table 1. Electrometric Determination of pH
Table 1 presents the pH level of
distilled water, assigned sample, and
Image 1: Adjusting the pH to the desired pH using the
pH meter.
buffer preparation through the pH meter
which are 7.0, 6.0, and 7.5 respectively.
Besides that, the hydrogen ion Discussion
concentration was also determined. 1 x
10-7 for the distilled water, 1 x 10-6 for the In the experiment, the group
assigned sample, and 1 x 10-8 also for made a phosphate buffer of 7.5 pH using
the buffer preparation. NaH2PO4 and Na2HPO4. Before measuring
the pH using a pH meter, it was
Table 2. Colorimetric Determination of pH calibrated in order to ensure its accuracy.
The group used the electrometric
method to measure the pH of the buffer
because it gives a specific number, which
is more accurate as compared to the
colorimetric method which gives a range
of values. On the first reading, the pH
meter read 7.3 pH which was not the
same as the calculated 7.5 theoretical
pH. This might be due to environmental
factors, thus the group adjusted the pH
of the buffer using 6.0M of NaOH and
diluted it with distilled water. Distilled
water did not change the pH of the
buffer.
The group then subjected the
Table 2 shows the color change selected sample of Micellar Water to a
that occurred to an acid-base indicator colorimetric method of pH reading using
when added to solutions with different several pH indicators. It was then found
pH levels. out that the Micellar Water was around
6.0 pH.
References
Bathan, G., Crisostomo, A., Daya, M.,
De Guia, R., Farrow, F., Gabona,
M. et al. (2017). Laboratory
Manual in General Biochemistry.
C&E Publishing, Inc.
Blacks, B. (2014). PH Colour Chart to
determine the pH level. England
Journal of Education, 68, 1-3.
Norby, J. (2000). “’The origin and the
meaning of the little p in pH’’.
Journal of Trends in the
Biochemical Sciences, 25 (1),
36-37.
Image 3. Microplate with distilled water, assigned
sample, and buffer prepared.
