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BIO3101 BIOCHEMISTRY: PRATICAL 1 INTRODUCTON TO THE LAB

INSTRUMENTS I

1. INTRODUCTION

Lab instrumentations are very important in various type of field, for example

biology, chemistry, physic etc. Different lab apparatus may carry a similar

function. Thus, as students, we should know the proper procedures of using

these lab instruments while conducting the experiments. This is to minimize the

accident that often occurs in the laboratory and to increase the accuracy to get

the result. Thus, the students will be exposed to the concepts and theories of

applications of several instruments that frequently used in the laboratory in this

practical. Therefore, students can differentiate the usage of different type of

apparatus. This enable the students to understand which lab instruments should

be used in each experiment.

2. OBJECTIVES

i. To recognize the names and the usages of the lab instruments that

frequently used in the laboratory.

ii. To identify the correct procedures to handle the lab instruments while

conducting the experiments.

iii. To understand the importance of safety while conducting the

experiments in the laboratory.

3. MATERIALS AND APPARATUS

3.1 Materials to measure the density of a solution:

-70ml of sample solution

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3.2 Apparatus to measure the density of a solution:

-Electronic balance, 100ml beakers, pipette, pipette bulb, graduated cylinder,

Erlenmeyer flask

3.3 Materials to measure the density of solid:

-Water, real object (stone)

3.4 Apparatus to measure the density of solid:

-Electronic balance, graduated cylinder, 100ml beaker

4. METHOD

4.1 Method to measure the density of a solution:

1. A 70ml of sample solution was measured and poured into a beaker

labelled as beaker A.

2. A second beaker was labelled as beaker B.

3. The weight of an empty beaker B was measured using an electronic

balance and recorded.

4. 20ml of the sample solution was measured and transferred into the

beaker B by using a pipette. Then, the weight of beaker B with the

sample solution was measured and recorded.

5. A third beaker was labelled as beaker C.

6. The weight of an empty beaker C was measured and recorded.

7. Another 20ml of the sample solution was transferred from beaker A to

beaker C by using a graduated cylinder. Then, the weight of beaker C

with the sample solution was measured and recorded.

8. A fourth beaker was labelled as beaker D.

9. The weight of an empty beaker D was measured and recorded.

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10. Another 20ml of sample solution from beaker A was transferred to

beaker D by using the Erlenmeyer flask. Then, the weight of beaker D

with the sample solution was measured and recorded.

11. All the weight measured was tabulated in Table 1.

12. The weight of the solution was determined by the formula :

Weight of the sample solution (g)

= Weight of the beaker with sample solution (g) Weight of the empty

beaker (g)

13. The density of the sample solution was calculated using the formula

below:

Density of sample solution (g/ml)

= Weight of sample solution (g) / Volume of sample solution (ml)

4.2 Method to measure the density of object:

1. The weight of a stone is determined by using an electronic balance

and recorded.

2. A graduated cylinder was half-filled with water and the volume of

the water was recorded in Table 2.

3. The stone was put gently inside the graduated cylinder which half-

filled with water to prevent the water spill out and to avoid the

damaging of graduated cylinder.

4. The final volume of the water was observed and recorded in Table

2.

5. The volume of the stone was determined by using the formula:

Volume of object (ml)

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= Volume of water with object (ml) Volume of water without

object (ml)

6. The density of the stone was calculated by this formula below:

Density of object (g/ml)

= Weight of the object (g) / Volume of object (ml)

5. RESULT

5.1 Table 1: Measuring the density of a solution

Pipette

Graduated

Cylinder

Erlenmeyer Flask

Weight of empty

beaker (g)

51.08 50.22 51.08

Weight of beaker

with sample

solution (g)

70.70 69.77 65.13

Weight of solution

(g)

19.62 19.55 14.05

Density of solution

(g/ml)

0.98 0.98 0.70

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5.1.1 Figure 1: Measuring the density of a solution

5.2 Table 2: Measuring the density of object

Weight of object (g) 5.06

Volume of water with object (ml) 27.00

Volume of water without object (ml) 25.00

Volume of object (ml) 2.00

Density of object (g/ml) 2.53

5.2.1 Figure 2: Measuring the density of object

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6. DISCUSSION

1. The weight of solution that gained from the pipette is the highest

compared to the graduated cylinder and Erlenmeyer flask although the volume

of solution is fixed at 20ml. Besides, the density of solution that gained from

pipette also is the highest among these three lab instruments. The density of the

solution can be defined as mass per unit volume (g/ml). The accuracy of the

pipette is the highest, followed by graduated cylinder and the Erlenmeyer flask.

The volume of pipette is ranged from less than 1 ml to about 100 ml with

tolerances of less than 0.2%. While the graduated cylinder is ranged in the size

from 10ml to 1000 ml with tolerances about 1%. Erlenmeyer flask are used for

the purpose of mixing, transporting, and reacting, but not for accurate

measurements. The volumes stamped on the sides of Erlenmeyer flask are

approximate and accurate to within about 5%. Thus, the volume measured by

the pipette is more accurate to 20ml, while the volume of solution measured by

graduated cylinder and Erlenmeyer flask may higher and not accurate to 20ml.

The lower the volume of solution, the higher the density of solution.

2. Beaker, Erlenmeyer flask and graduated cylinder are few examples of

lab instruments that are suitable to transfer 50ml of a reagent from its storage to

the bench. A synthesis experiment that requires 35ml of an acid solution can be

measured by using a graduated cylinder. 10ml of reagent can be measured as

accurately as possible by using a pipette for the purpose of titration experiment.

3. Water displacement method is used to measure the volume of solid

particle which have irregular shape, for example, stone, sand, etc. Volume can

be defined as the amount of 3-dimensional space an object occupies.

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4. The observation must be done at an eye level and read at the bottom of

a meniscus of the liquid level in order to obtain the volume of solution

accurately.

7. CONCLUSION

The names and the usages of the lab instruments that frequently used in the

laboratory were recognized. Besides, the correct procedures to use the lab

instruments were followed while conducting the practical. Moreover, safety is

important in the laboratory, all the rules and regulations were identified and

followed strictly.

8. REFERENCE

1. Dartmouth College, 1997-2000, Retrieved from

https://www.dartmouth.edu/~chemlab/techniques/flasks.html

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BIO3101 BIOCHEMISTRY: PRATICAL 2 INTRODUCTON TO THE LAB

INSTRUMENTS II

1. INTRODUCTION

There are basic lab instruments in the laboratory. However, some biochemistry

experiments require the use of special tools and special instruments. This is

because the biochemical experiment often involve the study of cells and

organelles, which are the basic unit form of the life. There are several types of

macro biological molecules and micro biological molecules in the cells of

organisms. To further study these biological components, the cell structures

must be disrupted. Thus, special instruments are needed to break down the cells

and separate the components based on their molecular weight, size of molecules

and density of the molecules. Therefore, early knowledge of these specializes

tools will be exposed to the students in this practical.

2. OBJECTIVES

i. To measure the pH of samples using pH meter and litmus paper, hence

compare both results.

ii. To determine the maximum wavelength of bromophenol blue using the

spectrophotometer.

iv. To investigate the relationship between the concentration of

bromophenol blue and the absorbance of bromophenol blue.

3. MATERIALS AND APPARATUS

3.1 Materials to measure the pH of samples:

Distilled water, tap water, vinegar, orange juice, soda water, milk, soap,

shampoo, litmus paper

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3.2 Apparatus to measure the pH of samples:

pH meter, beakers

3.3 Materials to determine the wavelength of bromophenol blue:

Bromophenol blue

3.4 Apparatus to determine the wavelength of bromophenol blue:

Cuvettes, spectrophotometer, beaker

3.5 Materials to investigate the relationship between absorbance and

concentration:

Bromophenol blue

3.6 Apparatue to investigate the relationship between absorbance and

concentration:

Cuvettes, spectrophotometer, beakers

4. METHOD

4.1 Method to measure pH of samples:

1. The sample solutions were prepared in each beaker with label.

2. The measuring probe of the pH meter was rinsed with the distilled water

before experiment was carried out.

3. The measuring probe of pH meter was immersed slightly into beaker

that containing the distilled water to obtain good contact between distilled water

and electrode.

4. The measuring probe of pH meter was then immersed in the different

sample solutions and stirred gently to obtain a stable pH.

5. The experiment was then repeated by using litmus paper.

6. The reading of each pH value shown by the pH meter and litmus paper

were recorded in Table 1 respectively.

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4.2 Method to determine the maximum wavelength of bromophenol blue:

1. The absorbance of 0.02mg/ml bromophenol blue was measured for

every 25nm within the wavelength from the range of 525nm to 650nm. The

absorbance was recorded in Table 2.

2. Distilled water was used as a reference for every time a record is taken.

3. A graph of absorbance against wavelength was plotted in graph 1. The

maximum wavelength of bromophenol blue was determined from the plotted

graph.

4.3 Method to investigate the relationship between absorbance and

concentration

1. The spectrophotometer was set with the maximum wavelength of

bromophenol blue that determined from previous practical.

2. The absorbance for each of the different bromophenol blue

concentrations was measured and recorded in Table 3.

3. A graph of absorbance against concentration of bromophenol blue

concentration was plotted in graph 2.

4. The absorbance of solution X with unknown concentration was

measured. The concentration of solution X was determined from the graph 2.

5. RESULT

5.1 Table 1: Measuring pH of samples

Samples pH meter (pH) Litmus paper (pH)

Distilled water 6.42 5

Tap water 6.23 6

Vinegar 1.88 2

Orange juice 2.30 3

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Soda water 8.72 10

Milk 6.57 7

Soap 7.44 7

Shampoo 5.97 6

5.1.1 Figure 1: Measuring the pH of sample solution using litmus paper

5.2 Table 2: Absorbance of bromophenol blue within the range of 525nm-

650nm

Wavelength (nm)

1st reading of

absorbance

(O.D.)

2nd reading of

absorbance

(O.D.)

3rd reading of

absorbance

(O.D)

Average of

absorbance

(O.D)

525 0.587 0.603 0.661 0.617

550 1.085 1.116 1.062 1.088

575 1.730 1.725 1.692 1.716

600 1.850 1.891 1.885 1.875

625 0.343 0.348 0.328 0.340

650 0.005 0.027 0.033 0.022

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5.2.1 Figure 2: Absorbance of bromophenol blue within the wavelength

range of 525nm-650nm

5.2.2 Graph 1: Absorbance against wavelength

0

0.5

1

1.5

2

2.5

500 520 540 560 580 600 620 640 660

Aso

rban

ce /

O.D

Wavelength / nm

Graph of Absorbance Against Wavelength

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5.3 Table 3: Absorbance of various concentration of bromophenol blue

Concentration

(mg/ml)

1st reading of

absorbance

(O.D.)

2nd reading of

absorbance

(O.D.)

3rd reading of

absorbance

(O.D)

Average of

absorbance

(O.D)

0.0025 0.297 0.296 0.296 0.296

0.005 0.562 0.562 0.562 0.562

0.01 1.094 1.092 1.093 1.093

0.015 1.431 1.431 1.430 1.431

0.02 1.950 1.952 1.954 1.952

Larutan X 1.231 1.230 1.231 1.231

5.3.1 Figure 3: Absorbance of various concentration of bromophenol blue

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5.3.2 Graph 2: Absorbance against various concentration of bromophenol

blue

6. DISCUSSION

1. From the result above, we can indicate that vinegar and orange juice are

acidic among the tested samples solution in Table 1. The pH of vinegar and

orange juice are 1.88 and 2.30 respectively which are measured by pH meter.

In addition, both samples of solution vinegar and orange juice show acidity

with the pH 2 and 3 respectively which tested using litmus paper. Besides,

soda water and soap are basic samples solution. The pH of soda water and

soap shown by the pH meter are 8.72 and 7.44 respectively. Using the litmus

paper, the pH of soda water and soap is 10 and 7 respectively. pH is a

numeric scale used to specify the acidity or alkalinity of an aqueous solution.

In chemistry, pH also defined as negative logarithm to base 10 of the

concentration of the hydrogen ion. An aqueous solution with the pH value

0.296

0.562

1.093

1.431

1.952

0

0.5

1

1.5

2

2.5

0.0025 0.005 0.01 0.015 0.02

Ab

sorb

ance

(O

.D.)

Concentration of bromophenol blue(mg/ml)

Graph of Absorbance Against Concentration

Absorbance

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that less than 7 is acidic while an aqueous solution with the pH value that

more than 7 is basic and the pH 7 indicates the aqueous solution is neutral.

2. Besides, there are differences between the pH meter and litmus paper

recording. A pH meter has a membrane that allows hydrogen ions (H+) to

pass through and create a voltage. The pH meter associates each voltage

with a particular pH value. The higher the concentration of acid, the more

hydrogen ions will pass through the membrane, thereby the changing

voltage is created. This voltage change will be interpreted as a higher pH

value. Litmus paper are impregnated with pH indicator molecules that

change colour upon contact with solution of a particular pH. The litmus

paper is then compared to a standard chart where the colours are compared

and then associated with a pH value. The accuracy of pH meter is higher

than litmus paper. Litmus paper is good for quick qualitative work, however

it fails at highly accurate quantitative work. The particular colours shown

by litmus paper indicate certain values and each measurement is only

accurate within a unit or two.

3. Spectrophotometry is a method to measure how much a chemical substance

absorbs light by measuring the intensity of light as a beam of light passes

through sample solution. Transmittance is the ratio of the intensity of the

transmitted light (I) to the intensity of the incident light (Io). While the

absorbance is defined as the negative logarithm of the transmittance which

is a measure of the amount of light absorbed at a particular wavelength as

the light passes through a sample of substance.

4. From the graph of absorbance of bromophenol blue against the wavelength

plotted, the graph is a bell-shaped. From the graph 1, we can analyse that

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absorbance reading is increasing from the wavelength of 525nm to 600nm.

However, the absorbance reading is then decreasing from the wavelength of

600nm to 650nm. The peak absorbance of bromophenol blue that obtained

from the graph 1 is 600nm which is 1.875O.D. The result obtained shows

the maximum absorbance of bromophenol blue occurs in the wavelength of

600nm.

5. A maximum wavelength that obtained in Graph 1, 60nm is set as in the

spectrophotometer to determine the absorbance readings of different

concentration of bromophenol blue, which are 0.0025mg/ml, 0.005 mg/ml,

0.01 mg/ml, 0.015 mg/ml, 0.02 mg/ml and solution X. A straight line should

be obtained in the Graph 2. This indicates the absorbance of bromophenol

blue is directly proportional to the concentration of bromophenol blue. The

higher the concentration of bromophenol blue, the higher the absorbance of

bromophenol blue. Therefore, the graph has obey the Beer-Lambert Law

where A=ebc

A= Absorbance / L mol-1cm-1

e= Length of cuvette path which contains the sample / cm

c= Concentration of compound in sample ( mol/l )

6. From the Table 3, the absorbance of solution X is 1.231 O.D. In order to

determine the concentration of solution X from the Graph 2, we need to

draw a horizontal line start from 1.231 O.D. at the y-axis entitled

Absorbance to meet a point at the straight line, then draw a vertical line

to the x-axis which entitled Concentration of bromophenol blue.

Therefore, the concentration of solution X is estimated as 0.0124 mg/ml.

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7. However, there are random errors might occur during the experiment as all

points should be in the straight line. The random error that might occurs is

the surface of the cuvette is not clear. For example, the fingerprints of

student printed at the clear and transparent surface of the cuvette and thus

affect the amount of light passing through. Hence, the reading absorbance

is affected. In order to reduce the random error, the surface of cuvette should

wiped by the clean tissue paper before place it into the spectrophotometer.

Besides, the fingers should put on the other side of the cuvette instead of the

clear side of the cuvette.

7. CONCLUSION

The uses of the special lab instruments should be known by the students. This

is because these specialized tools are important to study biochemical experiment.

Moreover, some precaution steps must be taken in order to obtain an accurate

result.

8. REFERENCE

1. Philip J. Carlson, 2004, PH Meter Versus PH Paper, Retrieved from

http://www.ehow.com/about_5840578_ph-meter-versus-ph-paper.html

2. Stephen Gallik, Ph. D, 2011, Transmittance and Absorbance, Retrieved

from http://cellbiologyolm.stevegallik.org/node/7

3. Gore, MG. 2000. Spectrophotometry and Spectrofluorimetry: A

Practical Approach. Oxford University Press, New York. 368 p.