Operation and Calibration of HPLC Pump for column washing


1.0       OBJECTIVE

  1.       To lay down a procedure for the operation and calibration of High Pump for column washing Make:  Shimadzu.

2.0 RESPONSIBILITY

2.1       Chemist / Executive – Quality Control – to follow the procedure.

2.2        Head – Quality Control for implementation and compliance.

3.0 PROCEDURE

  • 3.1 OPERATION
  • Ensure that the working area is clean and check the calibration status of the instrument.  
  • Switch on the power to the instrument, wait for initialization.
  • Keep the solvent line tube in appropriate solvents.
  •  Open the drain valve 2 turns anti clock wise direction.
  •  Click on ‘purge’ then pump starts flow.
  • Close the drain valve after completion of purge.
  • Connect  the column to the column inlet tube
  • Set the appropriate flow  rate by pressing function ‘Key’
  • Enter the appropriate flow rate and then press ‘Enter’.
  • Displays the new set volume of flow rate.
  • Press the pump key to start the pumping of set flow rate.
  • After completion of washing of the column again click on pump to stop the flow.
  • Enter the column washing details as per Annexure – 1 of  SOP QC073.
  • Switch off the instrument after usage.
  • 3.2  CALIBRATION
  • 3.2.1 FLOW ACCURACY TEST
  • Fix the C18, 75 x 4.6 mm column to the system.
  • Keep degassed methanol (HPLC grade) in solvent line and prime thoroughly, and stabilize for about 30 min. with a flow rate of 1 ml/min.
  • Take a 10 ml class ‘A’ calibrated dry volumetric flask.
  • When the flow and pressure are stable, insert the outlet tubing into the volumetric flask and immediately start a calibrated stopwatch.
  • Stop the stopwatch button when the meniscus reaches the 10 ml mark on the flask.
  • Record the elapsed time in seconds.
  • Calculate the flow rate using the following equation:   

10 ml * 60

                                        Calculated flow rate =       ————————-

                                                                            Measured Time in seconds

  • Record the calculated flow rate in Annexure – 1
  •  Similarly test the flow rate accuracy with flow rates of 0.5 and  2.0 ml/min and record the calculated flow rates in Annexure –1.
  •  Acceptance criteria: ± 0.02 ml/min of set flow rate.
  •  Calibration Schedule: Once three month and after any major maintenance job
4.0        ABBREVIATIONS
  • HPLC         –            High performance liquid chromatography.
  • HOD           –            Head of Department

5. 0      REFERENCES

Nil

6. 0      ANNEXURES            

  • ANNEXURE –  1      –  Calibration Record for Wash Pump.

ANNEXURE –  1 – Calibration Record for Wash Pump

Instrument No. Make 
Calibration date Next due date 
Model Page no1of  1

STOP WATCH ID:______________    CALIBRATION DUE DATE OF STOP WATCH:____________

VOLUMETRIC FLASK NO:___________________

1.           FLOW ACCURACY CALIBRATION RECORD

  Set flow rateObserved elapsed Time(sec) to collect 10 ml  Flow Rate       
 Result    (10*60 /Time in (sec)
Difference of Flow   Rate in ml
       0.5   ml/min       
        1.0  ml/min         
    2.0 ml/min             

Acceptance Criteria : + 0.02 ml/min of set flow rate

Results :                                Passes/Fails

Performed By  :________                                                               Verified By   : _______

Date               :________                                                           Date          : ________

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out /  Change )

Google photo

You are commenting using your Google account. Log Out /  Change )

Twitter picture

You are commenting using your Twitter account. Log Out /  Change )

Facebook photo

You are commenting using your Facebook account. Log Out /  Change )

Connecting to %s