This procedure is to be applied at the time of the Operation and Calibration of a visible spectrophotometer.
It is the policy of Xyz Limited that the written procedure shall be followed for the operation and calibration of a visible spectrophotometer and its use monitored to ensure that the instrument gives consistently accurate and reproducible results in compliance with regulatory requirements.
RESPONSIBILITY & ACCOUNTABILITY
Persons
along with their responsibilities & accountabilities are given below:
|
S. No |
Designation |
Responsibility |
|
01 |
Executive -
Corporate Quality Assurance |
To prepare SOP and follow the same |
|
02 |
Manager – Quality Assurance |
To ensure implementation of SOP. |
|
03 |
Section - in charge |
To ensure implementation of SOP. |
|
04 |
Manager
- Technical |
To ensure implementation of SOP. |
|
05 |
VP - Technical |
To ensure implementation of SOP. |
|
06 |
Assistant - General Manager |
To ensure implementation of SOP. |
5.0 PROCEDURE
5.1 PRELIMINARY CHECK
5.1.1 Check and ensure that the instrument is clean and suitable for starting
the operation. If it is not, clean and ensures suitability.
5.1.2 The instrument should be located at the ambient temperature.
5.1.3 Check the calibration status of an instrument before use.
5.2 CLEANING OF CUVETTE
5.2.1 Never use a brush to clean the inside of the cuvette.
5.2.2 Clean the Cuvette using Purified water or any organic solvent (e.g. Acetone or
Methanol) & dry the cuvette
5.2.3 Add about 1 ml of the solution to be measured. Tilt and turn the cuvette so that
the solution has contact with all the surfaces. Discard the solution and repeat
this rinse once more.
5.2.4 Fill the cuvette about ¾ full of the solution you wish to test.
5.2.5 Wipe the outside of the cuvette with a lint-free cloth, or tissue paper to remove any
moisture or fingerprints from the outside surface.
5.3 PRECAUTIONS
5.3.1 Do not touch the fine surfaces, always use the rough surface of the cells to hold.
5.3.2 Ensure cleanliness of the cells before and after use.
5.3.3 Use Purified water to rinse the cells.
5.3.4 Do not use directly Purified water to rinse cells after the use of solutions
prepared with immiscible solvents, use ethanol or methanol first
and then water.
5.3.5 Use a lint-free cloth duster for wiping the cells.
5.3.6 Use keeping cells in one direction in the cell holder throughout the test to
minimize the error in the blank and sample solutions.
5.3.7 Do not shake solutions of samples immediately before measurement.
5.3.8 The temperature of all solutions/reagents used in the test should not differ by
more than 0.5 ºC.
5.3.9 The solvent in the reference cell should be of the same batch as that used to
prepare the solutions
Operations of Spectrophotometer
Make sure the panel is closed and cuvette holders are empty
before starting.
Before
starting check power supply is On.
Turn the spectrophotometer "ON" by flipping the switch on the right side of
the instrument and allow 15-20 minutes for the instrument to warm up.
Set the appropriate wavelength with a wavelength setting knob from the front
panel of the instrument.
Set the zero in the air with a zero-setting knob. Fill the cuvettes with
blank / Solvent solution, wipe with a lint-free cloth duster and place it in the
cell holders & set 100% Transmittance with 100 setting knob. And also
adjust the sensitivity with a sensitivity knob from the front panel of the
instrument.
Then remove the cuvette, and discard the blank solution. Rinse the cuvette with the sample
solution. Then fill the cuvette with sample solution & replace it in the cell
holder. The monitor will display the Absorbance/Transmittance of the sample
solution at the wavelength displayed.
Switch “OFF” the instrument when not in use.
Maintain Logbook of Visible Spectrophotometer on XYZ/CQA/SOP-089/FR-01 Logbook for Visible Spectrophotometer (Appendix I)
6.0 CALIBRATION PROCEDURE
6.0.1 CALIBRATION FREQUENCY: Half Yearly and After Every Maintenance Job
6.1 WAVELENGTH ACCURACY
6.2 LINEARITY OF ABSORBANCE
6.3 SPECIFICATION OF CELLS
6.1 WAVELENGTH ACCURACY
6.1.1 Preparation
of 0.15M CoCl2.6H2O:
Prepare a stock solution of 1.5M CoCl2.6H2O by dissolving 17.84 g of CoCl2.6H2O in 50 ml of water. Pipette out 10.0ml of stock solution to the 100ml volumetric flask
and make up the volume with water up to the mark (0.15M CoCl2.6H2O).
6.1.2 Set the wavelength to 600nm and put the blank to set the 100% transmittance.
6.1.3 Clean, then rinse and fill the cuvette with 0.15M CoCl2.6H2O standard solution
and note the transmittance.
Repeat
the same for 650nm, 700nm, 750nm, 800nm, and 850nm.
|
Sr. No. |
Wavelength in nm |
Transmittance in % |
|
1.0 |
600 |
119.8 - 114.6 |
|
2.0 |
650 |
155.6 - 162.0 |
|
3.0 |
700 |
170.9 - 177.9 |
|
4.0 |
750 |
160.1 - 166.7 |
|
5.0 |
800 |
161.7 - 168.3 |
|
6.0 |
850 |
130.1 - 135.5 |
Acceptance
criteria:
Maximum Transmittance should be observed at 700 ±50nm.
6.1.5 Discard the solution after the calibration.
6.1.6 Record the calibration data as per XYZ/CQA/SOP-089/FR-02 Appendix II (Page 2 of
4).
6.2 LINEARITY OF ABSORBANC
6.2.1 Preparation of the stock solution
Prepare a stock solution of 1.5M CoCl2.6H2O by dissolving 17.84 g of CoCl2.6H2O in 50 ml of water. Working standard solution:
Make 0.15M CoCl2.6H2O solution
Pipette
out 5.0ml of stock solution to the 50ml volumetric flask and make up the volume
with water up to the mark
Make 0.12M CoCl2.6H2O solution
Pipette
out 4.0ml of stock solution to the 50ml volumetric flask and make up the volume
with water up to the mark.
Make 0.09M CoCl2.6H2O solution
Pipette
out 3.0ml of stock solution to the 50ml volumetric flask and make up the volume
with water up to the mark.
Make 0.06M CoCl2.6H2O solution
Pipette
out 2.0ml of stock solution to the 50 ml volumetric flask and make up the volume
with water up to the mark.
Make 0.03M CoCl2.6H2O solution
Pipette
out 1.0ml of stock solution to the 50ml volumetric flask and make up the volume
with water up to the mark
|
Sr.
No. |
concentration |
Transmittance
at 550 nm |
|
1 |
0.1500 |
44.1
to 45.9 |
|
2 |
0.1200 |
51.9
to 54.0 |
|
3 |
0.0900 |
60.8
to 63.2 |
|
4 |
0.0600 |
70.6
to 73.4 |
|
5 |
0.0300 |
83.3
to 86.7 |
Acceptance
criteria
Transmittance
should be ±2%.
The
linearity coefficient ‘r’ obtained from the linearity graph of CoCl2.6H2O for
different levels should not be less than 0.99.
6.2.3 Record the calibration data as per XYZ/CQA/SOP-089/FR-02 (Appendix II)
6.3 SPECIFICATION
OF CELL
In Photometry mode select transmittance mode. Set the wavelength at 400 nm. Then
set ‘zero’ to get 100 % transmittance with air blank in the cell holders. Then
place the cuvettes in the sample compartment filled with Milli Q water using an air blank and read the transmittance. Similarly repeat the operation for 450nm,
500nm, and 550nm. The transmittance values should meet the requirement as given
in the following table.
Acceptance
criteria
|
WAVELENGTH |
LIMIT OF % TRANSMITTANCE |
|
400nm |
Not Less Than 73% |
|
450nm |
Not Less Than 76% |
|
500nm |
Not Less Than 79% |
|
550nm |
Not Less Than 82% |
6.3.2 Record the calibration data as per XYZ/CQA/SOP-089/FR-02 (Appendix II) page 4
of 4.
6.3.3 Summarize all the data on XYZ/CQA/SPO-089/FR-03 Visible Spectrophotometer
Calibration Summary Sheet Appendix III
7.0 GENERATION OF INSTRUMENT CALIBRATION NUMBER
Generate the Instrument Calibration number on the calibration datasheet as INSCALXXYYZZZ
Where INS denotes Instrument, CAL denotes Calibration
XX
denotes the year, YY denotes the Month ZZZ denotes the sequence number.
8.0 ABBREVIATIONS
INS
CAL: Instrument
Calibration
XYZ: Xyz Limited
SOP: Standard Operating Procedure
CQA: Corporate Quality Assurance
T: Transmittance
Appendix & Logbook: - Download
