EXPERIMENT 6

COLORIMETRIC METHOD OF ANALYSIS

Group 2:Falgui, Lim, Pumbaya, Tiunayan, Mr. JanMark Soriano

Villaraza

2PHAR1 PHARM121LAB

OBJECTIVE: To determine the concentration of a solution by colorimetric method using the

Spectronic 20 Spectrophotometer

MATERIALS:
ferric chloride powder
Unknown solution of ferric chloride
Spectronic 20 Spectrophotometer

PROCEDURES:
A. Operation of the Spectronic 20 Spectrophotometer

1. Switch on the instrument lamp by means of the lamp switch located at the front of the
instrument.
2. Place a clean cuvette containing the solvent in the sample compartment of the
instrument. Adjust the wavelength to the required value. (595-650)
3. Turn the scale by means of the knob located in front of the instrument until the scale
reading zero
4. The instrument is now ready for the standard and sample readings.

B. Preparation of Standard ferric chloride Solution

1. Dissolve exact 100mg of ferric chloride powder, USP grade or its equivalent, accurately
weigh in a 100ml volumetric flask and make up to volume with distilled water. This is the
standard solution
2. Prepare several dilutions from the stock solution as follows

standard stock solution - 100 mg - 100 ml

C. Preparation of the Unknown solution

1. Prepare the dilutions of the unknown solutions close to the concentrations of the
above standard solution.

Note: The instructor prepares the samples sufficient for the number groups in the class.

D. To read the solutions in the absorbance values

1. Remove the cuvette containing the solvent replacing with the standard solutions one
after the other. Note down the readings.
 2. Repeat the procedure with the sample or unknown solutions per group.
3. Record all results on the table.

SPECTROPHOTOMETER ANALYSIS
ferric chloride wavelength= 650 nm

water G2 G3 G4 G5 G6 G7 G8
0mg/100 0.5mg/ 1mg/ 1.5mg/ 2mg/ 2.5mg/ 3mg/ 3.5mg/
Concentrations ml 100ml 100ml 100ml 100ml 100ml 100ml 100ml
of standard per
100mL
100.7 50.9 52.2 80.7 44.9 43.5 56.9 42.9
Transmittance
of standard
Questions
1. Differentiate Colorimetry and Spectrophotometry
● According to Hzdg (n.d.), colorimetry is a process of utilizing a colorimeter to
observe the measurement of color by human perception through Beer-Lambert’s
Law stating that a solute’s measured concentration should be proportional to the
said solute’s absorbance. According to Drawell (2023), spectrophotometry is
defined as the utilization of a spectrophotometer to measure the transmitted
light’s absorption and it’s wavelength.
2. Give the important principles of Spectrophotometric analysis.
● Absorption and Transmission: Measuring how a material interacts with light,
either absorbing, reflecting, scattering, or transmitting it.
● Beer-Lambert Law: Establishing a linear relationship between absorbance and
concentration of an absorbing species in a solution.
● Instrumentation: Comprising light sources, monochromators, sample holders, and
detectors for accurate measurement.
● Nature’s Inert Property: Based on the fact that every compound, that is present in
nature, has a property to absorb, transmit, or reflect light (electromagnetic
radiation) at a certain wavelength where spectrophotometry now deals with the
measurement of the interaction of light with these materials.
● Components: essentially consists of four important components: a light/radiation
source, a collimator, a monochromator, and a detector. The monochromator
comprehends a fixed entrance slit, a dispersing element such as a prism or a
diffraction grating, and a moving exit slit.

3. Draw a basic spectrophotometer. Label the parts and give their functions.
1. Pilot Lamp:
● Description: A small light source typically used to illuminate the scales or dials on
the spectrometer, making them easier to read.
● Function: Helps the user see and operate the controls, especially in low-light
conditions.
2. Needle:
● Description: A pointer or indicator, often part of an analog gauge, that moves over
a scale to show measurements.
● Function: Indicates the value of the measured variable, such as absorbance or
transmittance, by pointing to the corresponding value on the scale.
3. Mirror:
● Description: A reflective surface used to direct the light path within the
spectrometer.
● Function: Reflects and directs the light beam through the optical components of
the spectrometer, ensuring the light reaches the sample and the detector.
4. Gauge:
● Description: An instrument or device with a scale and a pointer used to measure
and display a specific parameter.
● Function: Displays measurements such as transmittance or absorbance, helping
the user to read and interpret the results.
5. Sample Compartment:
● Description: The area or holder where the sample to be analyzed is placed.
● Function: Holds the sample in the path of the light beam, allowing the
spectrometer to measure the sample's absorbance or transmittance.
6. Wavelength Control:
● Description: A dial, knob, or electronic control used to select the wavelength of
light the spectrometer uses.
● Function: Allows the user to choose the specific wavelength of light for the
analysis, enabling measurements at different points of the spectrum.
7. Power Switch/Zero Control:
 ● Description: A combined control used to turn the spectrometer on and off and to
set the baseline measurement to zero.
● Function: Powers the device and calibrates it by setting the initial reading to zero,
ensuring accurate measurements.
8. Transmittance/Absorbance Control:
● Description: A control used to switch between measuring transmittance (the
amount of light passing through the sample) and absorbance (the amount of light
absorbed by the sample).
● Function: Allows the user to toggle between and measure either transmittance or
absorbance, providing flexibility in the type of analysis performed.
Product: Cupric Sulfate Batch No.: N/a

Form: Liquid Lot No.: N/a

Quality Control Sheet

Date Performed: Date Submitted:

A. Procedure (You can have it in bullet form or in number form, you can also add
flowchart or flow diagram)

Procedure:
Procedure for Determining the Concentration of Ferric Chloride Solution Using the Spectronic
20 Spectrophotometer

Objective: To determine the concentration of a solution by the colorimetric method using the
Spectronic 20 Spectrophotometer.

Materials:
- Ferric chloride powder
- Unknown solution of ferric chloride
- Spectronic 20 Spectrophotometer
- Cuvettes
- 100 ml volumetric flasks
- Distilled water
- Analytical balance
- Pipettes

Procedure:

A. Operation of the Spectronic 20 Spectrophotometer

1. Switch on the instrument lamp using the lamp switch located at the front of the instrument.
2. Place a clean cuvette containing the solvent (distilled water) in the sample compartment of
the instrument.
3. Adjust the wavelength to 650 nm using the wavelength knob (specific to ferric chloride,
confirm optimal wavelength with instructor).
4. Turn the scale knob located at the front of the instrument until the scale reading is zero with
the solvent cuvette in place. The instrument is now ready for standard and sample readings.

B. Preparation of Standard Ferric Chloride Solution

1. Accurately weigh 100 mg of ferric chloride powder using an analytical balance.
2. Dissolve the weighed ferric chloride powder in a 100 ml volumetric flask and make up to
volume with distilled water to prepare the standard stock solution (1 mg/ml).
3. Prepare several dilutions from the standard stock solution as follows to obtain a range of
concentrations:
 - 0 mg/100 ml: Use 100 ml distilled water.
- 0.5 mg/100 ml: Pipette 0.5 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.
- 1 mg/100 ml: Pipette 1 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.
- 1.5 mg/100 ml: Pipette 1.5 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.
- 2 mg/100 ml: Pipette 2 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.
- 2.5 mg/100 ml: Pipette 2.5 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.
- 3 mg/100 ml: Pipette 3 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.
- 3.5 mg/100 ml: Pipette 3.5 ml of standard stock solution into a 100 ml volumetric flask and
dilute to volume with distilled water.

C. Preparation of the Unknown Solution

1. Prepare the dilutions of the unknown solution close to the concentrations of the above
standard solutions.

D. Measurement of Absorbance
1. Remove the cuvette containing the solvent and replace it with the cuvette containing
the first standard solution. Note the absorbance reading.
2. Repeat step 1 for each of the standard solutions and record the absorbance readings.
3. Repeat the procedure with the sample or unknown solutions for each group, noting
down the absorbance readings.
4. Record all results in a table similar to the one below.
E. Absorbance Results
A. Water (0mg/ml) - 0
B. Ferric Chloride (0.5mg/100ml) - 0.29
C. Ferric Chloride (1mg/100ml) - 0.28
D. Ferric Chloride (1.5mg/100ml) - 0.09
E. Ferric Chloride (2mg/100ml) - 0.35
F. Ferric Chloride (2.5mg/100ml) - 0.36
G. Ferric Chloride (3mg/100ml) - 0.24
H. Ferric Chloride (3.5mg/100ml) - 0.37

B. Graphs or Computations (Please include here your Graphical or Chart or

Computation if APPLICABLE)
C. Disposition of the Batch (Please indicate here PASSED or FAILED, then provide
JUSTIFICATION)

According to the Beer-Lambert Law, the graph of their Transmittance

versus concentration will show a linear relationship where transmittance
decreases logarithmically as concentration increases. The graph is expected to
show an exponential decay, starting high at low concentrations and dropping off
rapidly as concentration increases. In line with this, the actual results of the
experiment did not show the expected result which marks a FAILED disposition
as the graph showed a non-linear relationship, with fluctuating results.