SOP For Operation and Calibration Of Visible Spectrophotometer

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 ÂșC.

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 CALInstrument Calibration 

XYZ: Xyz Limited

SOP: Standard Operating Procedure

CQA: Corporate Quality Assurance 

T: Transmittance

Appendix & Logbook: - Download

BANTI SINGH

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