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SOP for Operation and Calibration of Gas chromatography (THERMO-TRACE)

Objective
The objective of this Standard Operating Procedure is to lay down the procedure for the operation and calibration of Gas chromatography (THERMO-TRACE)

Scope
This SOP covers the operation and calibration procedure of Gas chromatography (THERMO-TRACE).

Responsibility
QC Officer: Responsible for the operation and maintenance of the instrument as per the procedure.
Head of Department: Responsible for maintenance, timely as per schedule.
QA Manager: Review the records and governing the document



Procedure

Operation
  • Ensure that the instrument is clean and free from dust. Wipe all the traces of solvent/water/moisture by dry cloth.
  • Properly install a suitable capillary column in the GC oven injector & selected detector.
  • Open the Gas pressure valve from the distribution panel. (N2 cylinder for N2 set 60 psi for N2).
  • Check the backpressure of column.
  • Switch on the mains power, stabilizer, GC power & computer power.
  • Double click on the Chrom-card icon from the desktop, chrom-card window display on the screen, double click on Konark icon, select the method from edit G.C parameters icon, then. Apply the chromatographic condition and enter ok open run signal. After completion of the run, open the instrument in offline, then go to the reprocessing menu and select file and click on ok button, integrate peak, after integration enter the component
  • name then click on ok, go to edit method than go to in report parameters and click on ok to view the print layout print chromatogram by clicking printer icon.
  • Condition the column at 280°C for 10 min. Enter the programming condition of the sample through micro syringe, when G.C shows ready signal.
  • Cool the G.C after completion of the analysis & switch it off. Shut down the computer, mains power supply. Close the gas supply from the distribution panel & from cylinder.

Calibration Procedure

Calibration of Flow rate
  • Open the column oven compartment.
  • Remove the column if attached to the injector port –1 (to be calibrated).
  • Connect the soap film flow meter (side arm) and injector port outlet with the help of Teflon or a rubber tube.
  • Fill the pipette bulb partially with a soap solution and attach to the bottom of the flow meter.
  • Open the knob of carrier gas (N2) and set up proper pressure (i.e. 500 kpa or 5 Kg/cm2) in the carrier gas pressure controller.
  • Check for leaks at each and every point of attachment using soap solution.
  • Open the knob of carrier gas flow controller & allow carrier gas to flow through the corresponding digital flow control.
  • Adjust the carrier gas flow rate with flow control (30 ml/min).


  • Gentle squeeze the bulb to force a soap film up into the gas stream. Start the stopwatch as soon as the film reaches to zero ml mark. Stop the watch when the film reaches to 30 ml marking. Note down the time require to reach the film from 0 to 30 ml mark. Calculate the flow rate using the following formula.


  • Similarly calibrate the flow rate after the time interval of 20 min, 40 min, & 60 min and find out the difference between the readings.
  • Acceptance criteria: observed flow rate of the equipment should be within ±2ml/min of set flow rate.

Calibration of FPD

1. Detector precision and consistency of relative retention time:
  • Preparation of pesticide standard (0.2ppm): Take 100µl of standard solution (100ppm) of Methyl parathion in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 2 ml of this solution in 10 ml volumetric flask and dilute to mark with acetone.
  • Inject 2.0µl of pesticides standard 5 times and observe area and RT.
ACCEPTANCE CRITERIA:
The deviation of RT ± 0.2 min
The deviation of area ± 10%

2. Detector Linearity:
Prepare the five different concentration solution as follow to check the detector linearity.
  • Preparation of pesticide standard 1 (0.025 ppm): Take 100µl of standard solution (100 ppm) of Methyl parathion in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1ml of this solution in 10ml volumetric flask and dilute to mark with acetone. Take 5ml of this solution in 10ml volumetric flask and dilute to mark with acetone. Take further 5ml in 10ml volumetric flask and dilute to mark with acetone.
  • Preparation of pesticide standard 2 (0.05 ppm): Take 100µl of standard solution (100 ppm) of Methyl parathion in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1ml of this solution in 10ml volumetric flask and dilute to mark with acetone. Take further 5ml in 10ml volumetric flask and dilute to mark with acetone.
  • Preparation of pesticide standard 3 (0.1 ppm): Take 100µl of standard solution (100 ppm) of Methyl parathion in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1ml of this solution in 10ml volumetric flask and dilute to mark with acetone.


  • Preparation of pesticide standard 4 (0.2 ppm): Take 100µl of standard solution (100 ppm) of Methyl parathion in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1ml of this solution in 5ml volumetric flask and dilute to mark with acetone.
  • Preparation of pesticide standard 5 (0.4 ppm): Take 100µl of standard solution (100 ppm) of Methyl parathion in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 2ml of this solution in 5ml volumetric flask and dilute to mark with acetone


  • Inject 3 replicates injection of the above solution and observes area.
  • Plot the graph of linearity and calculate the correlation coefficient.
Acceptance criteria:
Correlation coefficient minimum 0.99

CHROMATOGRAPHIC CONDITION:
Name of the instrument: GC [Thermo Trace ultra]
Column: CPSIL-8CB [30m X 0.25 mm ID X 0.25µm]
                                                          @ 15°C/min
Oven Temp.: 130°C (3min)   270°C (10 min)
Injector Temp.: 250°C
Detector Temp.: 300°C
Carrier Gas: N2 [2 ml] H2 (115ml) AIR (120ml
Injection Volume: 2.0µl
Split Ratio: Split less

Calibration of NPD

1. Detector Precision and consistency of relative retention time: 
  • Preparation of pesticide standard (0.2 ppm): Take 100 mL of standard solution (100 ppm) of Chlorpyrifos in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 2 ml of this solution in 10 ml volumetric flask and dilute to mark with acetone.
  • Inject 2.0µL of above pesticide standard 5 times and observe area and RT.

ACCEPTANCE CRITERIA:
The deviation for RT ± 0.2 min and deviation for area ± 10%.

2. Detector Linearity:
Prepared the five different concentrate solution as follow to check the Detector Linearity
  • Preparation of pesticide standard 1 (0.025ppm): Take 100 mL of standard solution (100 ppm) of Chlorpyrifos in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1 ml of this solution in 10 ml volumetric flask and dilute to mark with acetone. Take 5 ml of this solution in 10 ml of volumetric flask and dilute to mark with hexane. Take further 5 ml in 10 ml volumetric flask and dilute to mark with acetone.
  • Preparation of pesticide standard 2 (0.050ppm): Take 100 mL of standard solution (100 ppm) of Chlorpyrifos in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1 ml of this solution in 10 ml volumetric flask and dilute to mark with acetone. Take further 5 ml in 10 ml of volumetric flask and dilute to mark with acetone.
  • Preparation of pesticide standard 3 (0.1ppm): Take 100 mL of standard solution (100 ppm) of Chlorpyrifos in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 1 ml of this solution in 10 ml volumetric flask and dilute to mark with acetone.


  • Preparation of pesticide standard 4 (0.2ppm): Take 100 mL of standard solution (100 ppm) of Chlorpyrifos in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 2 ml of this solution in 10 ml volumetric flask and dilute to mark with acetone.
  • Preparation of pesticide standard 5 (0.4ppm): Take 100 mL of standard solution (100 ppm) of Chlorpyrifos in 10 ml volumetric flask containing 5ml acetone and dilute to mark with same solvent. Take 2 ml of this solution in 5 ml volumetric flask and dilute to mark with acetone.

  • Inject 3 replicates injection of the above solution and observe Area
  • Plot the graph of linearity and calculate the correlation-coefficient

CHROMATOGRAPHIC CONDITION
Name of Instrument: GC [THERMO]
Column: CPSIL-8CB(30.0 M x 0.25 X 0.25mm)
Oven Temp.: 130°C (3min hold)@5°C/min --> 270°C (10 min hold)
Detector: 300 °C(NPD)
Injector: 250 °C
Carrier Gas: N2 [2.0ml/min ]
Injection volume: 2 mL
Split Ratio: split less

Acceptance criteria:
Correlation-coefficient Minimum 0.99

Calibration of Oven
Calibration of oven to be done by calibration department.

Frequency of calibration: Quarterly

Routine Maintenance
  • Clean the all, out side part of instrument.
  • Clean the injector and injector liner. Change the glass wool of injector liner. Change the septa of injector port if required.
  • Check all connection of gases for leakage.

Annexure
Nil

Abbreviation
GC: Gas chromatography
QC: Quality Control
QA: Quality Assurance

Revision History
Nil

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