CPB30103- Biochemical Engineering

Experiment 1: Preparation of Buffer Solution

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SUMMARY
Buffer solution is an aqueous solution consisting of a mixture of a weak acid and its conjugate
base, or vice versa. Its pH changes very little when a small or moderate amount of strong acid or
base is added to it and thus it is used to prevent changes in the pH of a solution. Buffer solution
are used as a means of keeping pH at a nearly constant value in a wide variety of chemical
application such as to maintain the correct pH for enzymes in organisms to work. In this
experiment, we were preparing buffer solution using 0.1 M of citric acid and 0.1 M sodium
citrate. The amount for each solution needed to prepare buffer solution were calculated and its
value were 91.32ml for citric acid and 158.69ml for sodium citrate. Then the pH of buffer
solution was compared with the theoretical value that has been calculated using HendersonHasselbalch equation. The second experiment was conducted to test the effectiveness of the
buffer solution which using hydrochloric acid and sodium hydroxide that been poured into
respectively 100ml beaker containing distilled water. The experiment was repeated by replacing
distilled water with the buffer solution and the pH values were taken and recorded. During the
experiment, there were errors that might affect our results such as the amount of citric acid and
sodium citrate where the theoretical amounts were 91.32ml and 158.69ml respectively but the
amount been poured into the beaker were 91ml and 159ml respectively. Then, the pH values
taken also might been affected by the unwell stirred during the mixing process and the pH
electrode was not well cleaned. We also make some recommendation to improve this experiment
in the future such as before using any apparatus, the apparatus need to be clean up to ensure the
apparatus is contamination-free. At the end of the experiment, we find that pH value is affected
from the number of hydrogen ions or hydroxyl ions in a solution and the effectiveness of buffer
solution can be evaluate based on the more A and HA molecules available, the less of an effect
addition of a strong acid or base will have on the pH of a system. The other factor of
effectiveness is the absolute concentrations of the acid and conjugate base or vice versa.

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Experiment 1: Preparation of Buffer Solution

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OBJECTIVES
1.
2.
3.
4.

To study the acid and base chemistry.

To prepare a buffer solution using citric acid and sodium citrate.
To calculate the preparation of buffer solution by using Henderson-Hasselbalch equation.
To test the effects of added acid or base act on the pH of the solutions of distilled water

and buffer solution.

5. To determine the effectiveness of buffer solution by mixing with strong acid and strong
base whether the solution resist to the pH changes.
6. To determine the theory and error of pH measurement.
7. To define buffer capacity and the factors affecting the buffer capacity.

DATA AND RESULTS

pH READING
INITIAL

AFTER ADDING
HCI

AFTER ADDING
NaOH

DISTILLED WATER

7.46

1.53

8.34

BUFFER
SOLUTION

5.62

4.78

4.92

DISCUSSION

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Based on the experiment of preparation of buffer solution that had been conducted, in the
part (a) preparation of the buffer solution, the buffer solution was prepared by mixing 0.1 M of
citric acid solution, the buffer agents form (HA) with 0.1 M of sodium citrate solution, its
conjugate base (A-). The initial pH value of buffer solution that was calculated using the
HendersonHasselbalch equation and obtained from the experiment by pH electrode are 5.00 and
5.62, respectively. The difference between the expected and obtained pH value was 0.62. The
difference occurred because of small inaccuracy amount of citric acid and sodium citrate needed
that poured into the measuring cylinder where for example the actual amount of citric acid and
sodium citrate solution are 91.32 mL and 158.69 mL, respectively while the amount of citric acid
and sodium citrate that was taken by us during conducting the experiment are 91 mL and 159
mL, respectively.
In part (b) effectiveness of the buffer solution, the recorded value of 7.46 was the initial
pH value for the distilled water that was obtained which it indicated that the distilled water had
been well maintained. Based on the theory, the pH value of distilled water is between 5.6 and
7.0. The pH value might change and easily influenced by the present of CO2 in atmosphere and if
it was left exposed to the atmosphere where dissolved CO2 reacted slowly with water to give the
bicarbonate and hydronium ions.
From the experiment, firstly, 50 mL of the distilled water was added into the strong acid,
1.0 mL of 1.0 M HCl in the 100 mL beaker. The pH value obtained is 1.53 which meant the
distilled water has turn into acid solution. Next, at the same time, 25 mL of the buffer solution
was also added into strong base, 1.0 mL of 1.0 M NaOH in the other 100 mL beaker, the pH
value obtained is 8.34. From the result, it could be obviously seen as the deionized water was
turned into acid and base solutions as the strong acid and strong base was added respectively.
The procedure above was repeated by replacing distilled water with the prepared buffer
solution. From the experiment, firstly, 25 mL of the buffer solution was added into the strong
acid, 1.0 mL of 1.0 M HCl in the 100 mL beaker where the pH value changed from 5.62 to 4.78.
It showed the difference occurred was 0.84 where it could be considered as quite small of the
difference. Next, at the same time, 25 mL of the buffer solution was also added into strong base,
1.0 mL of 1.0 M NaOH in the other 100 mL beaker, the pH value changed from 5.62 to 4.92. The
difference also could be considered as quite small which was 0.70. The small different that was

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occurred might be came from several errors that happened during conducting the pH electrode or
handling the solutions.
The error of pH measurements occurred gave a slight different as the calculated one. The
distilled water and prepared buffer solution may have a little contamination because of not stirred
well during mixing the solutions with acid or base that causing the pH value to become 7.46
instead of 7.00 for distilled water and for buffer solution, 5.62 instead of 5.00. Next, the error
occurred while handling the distilled water and buffer solution with the pH electrode. The data
was taken before it stabled or the pH electrode itself was not well cleaned. Then, the parallax
error might happen such as the position of eyes was uneven with level of solutions or liquids in
the beaker during the chemical was taken where finally, the amount or value of the solutions
might not be accurate as desired.
Our buffer solutions that was composed of a weak acid and its conjugate base was still
tend to resist changes in the pH value as acid or base was added, 4.78 and 4.92, respectively
because addition of H+ had given a little effect because it was absorbed. The initial pH of buffer
was 5.62 where the difference of the pH value was only between 0.70 and 0.85 which could be
considered small in changes. The characteristics and process of the buffer solution could be seen
even though the pH value of buffer solution obtained was different from the calculation.
However, the distilled water solutions was not tend to resist changes in the pH value as acid or
base was added. The distilled water turns into acid and base solutions when were being added
with HCl and NaOH, respectively.

CONCLUSION

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The objective of this experiment is to study more about the buffer solution and the pH
measurement. Through this experiment, buffer solution was prepared and been reacted with the
acid and base in order to study about the effectiveness. The result from that was compared to
earlier experiment where the acid and base were reacted with the distilled water. From the result
obtained, it can be conclude that the pH value be affected from the number of hydrogen ions (H+)
or Hydroxyl ions (OH-) that are present in the solutions. Furthermore, in our study of the
effectiveness of the buffer solution, the reason behind it is from the ratio of weak acid and its
conjugate base [A-]/[HA] that consist in the solution.The more A- and HA molecules available,
the less of an effect addition of a strong acid or base will have on the pH of a system. The other
factor that affect the effectiveness is the absolute concentrations of the acid and conjugate base or
vice versa. This experiment can be said as successful.
As the recommendation, in order to obtain more accurate and desirable result and data of
this experiment in the future, it is necessary to clean up the apparatus first before using it to
ensure the apparatus is contamination-free. Next, to avoid parallax error that caused an error to
the experiment, during observing and reading the values of liquids or solutions, it is necessary
for the position of eyes must be in the same level and parallel as the meniscus of the measuring
cylinder. Then, the pH electrode must be rinsed using distilled water each time handling the pH
electrode and changing the solutions to ensure there is no solutions residue (acid or base
solutions) on the pH electrode so that desirable and accurate pH value can be obtained.

QUESTIONS

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Experiment 1: Preparation of Buffer Solution

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1. Explain the theory of pH measurement.

pH measurement basically depends on the pH sensitive electrode which usually made
from glass, a reference electrode, temperature element to provide a temperature signal to
the pH analyzer. pH electrode has the ability to produce a voltage proportional to pH of
the solution when it is connected with the solution. Besides that, the reference electrode is
design to maintain the voltage at any temperature and to complete the measuring circuit
within the solution.. It provides a known reference potential for the pH electrode. The
difference in the potentials of the pH and reference electrodes provides a millivolt signal
proportional to pH. Most pH sensors are designed to produce a 0 mV signal at 7.0 pH,
with a (theoretically ideal) slope (sensitivity) of -59.16 mV / pH at 25C.
2. Define buffer capacity and the factors affecting the buffer capacity.
Buffer capacity is a level of the efficiency of a buffer in resisting changes in pH.
Conventionally, the buffer capacity ( ) is expressed as the amount of strong acid or base, in
gram-equivalents, that must be added to 1 liter of the solution to change its pH by one unit.
The factors that affecting buffer capacity are :
Ratio of the salt to the acid or base. Buffer capacity is define optimal when the pH = pKa
Total buffer concentration which mean the total concentration of acid or base to reduce the
concentration of buffer solution.
3. Discuss the importance of pH buffer in industrial system.
For every industrial system, there are specific importance of the pH buffer for each sectors,
for example
i.

Pharmaceutical industries
Most of the medicines are prepared in aqueous solution of different chemicals so these
aqueous solutions require a constant pH in order to assure the stability and clinical
effectiveness of a medicines and this is done through buffers. Buffers are also added in
pharmaceuticals to improve patient comfort and to make longer transportation of medicines
possible. Apart from this buffers are also used to:

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Experiment 1: Preparation of Buffer Solution

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Maintain some drug or medicine in ionized form as ionized forms are more soluble in
aqueous solutions.
Maintain some drug or medicine in un-ionized form as un-ionized forms are more soluble
in lipids.
Maintain the stability of drugs in different aqueous solutions as many drugs are
vulnerable to hydrolysis of aqueous solutions.
Maintain the pH of most of the drugs or medicine near to neutral otherwise that specific
drug or medicine may cause irritation in body tissues.

REFERENCES
1. Reach Devices. 2013. Biological Buffer.
[http://www.reachdevices.com/Protein/BiologicalBuffers.html]. Accessed October 15
2016.
2. Clark, Jim. 2016. Buffer Solution.
[http://www.chemguide.co.uk/physical/acidbaseeqia/buffers.html]. Accessed October 15
2016.
3. About Education. 2016. Henderson Hasselbalch Equation and Example.
[http://chemistry.about.com/od/acidsbase1/a/hendersonhasselbalch.htm]. Accessed
October 15 2016.
4. ChemCollective. Predicting the pH of a Buffer.
[http://chemcollective.org/activities/tutorials/buffers/buffers4]. Accessed October 15
2016.
5.

Chem Collective: Buffer Capacity. Carnegic Mellon University . Retrieved October 8,

2016 ,from http://chemcollective.org/activities/tutorials/buffers/buffers5

6. 2.) The Pharmaceutics and Compounding Laboratory: Buffer and Buffer Capacity.
Retrieved October 8, 2016, from http://pharmlabs.unc.edu/labs/ophthalmics/buffers.htm

APPENDICES

CPB30103- Biochemical Engineering

Experiment 1: Preparation of Buffer Solution

CALCULATION
Equation to calculate pH value
+
H

K=
+
H

+
H

log 10
+
H

log 10

pH= p K a +log
Calculate volume

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Experiment 1: Preparation of Buffer Solution

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pH=5
p K a=4.76

5=4.76+ log

0.24=log

[ sodium citrate ]
[ citric acid ]

[ sodium citrate ]
[ citric acid ]

1.7378 [ citric acid ] =[ sodium citrate ]

For each volume of citric acid, 1.7378 volumes of sodium citrate must be added. Thus, the
total volume is 2.7378

sodium citrate=

1.7378
500 ml=317.37 ml
2.7378

( 317.37
2 )

158.69 ml
citric acid=

( 182.63
2 )

91.31 ml

1
500 ml=182.63 ml
2.7378

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Experiment 1: Preparation of Buffer Solution

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