SOP for Operating Procedure for Dionex Summit HPLC Quaternary Gradient System

OBJECTIVE

To lay down the procedure for operating the HPLC Quaternary Gradient system for better and error-free use.

SCOPE

This procedure is applicable for the HPLC Quaternary Gradient system installed at the Quality Control Laboratory at Pharmainfo Limited.

RESPONSIBILITY

Execution: Executive and above - QC department

Checking: Asst. Manager and above- QC department

ACCOUNTABILITY

Head of the QC Department

PROCEDURE

• Ensure that the instrument is visually clean and calibrated.
• Make necessary entries in respective instrument log cards as per reference SOP.
• Prepare mobile phase as per Standard test procedure or analytical report of respective sample to be analyzed.
• Prepare the appropriate rinsing solution for an autosampler. Degas the solution by ultrasonic treatment or by applying a vacuum.
• Place the tubes A, B, C, and D leading to the pump in suitable solvents or mobile phase.

• Turn on the P680 binary gradient pump main unit’s power.
• Turn on the mains of ASI 100 auto sampler main unit’s power.
• Turn on the TCC 100 column oven main unit’s power.
• Turn on the UVD 170 U detector main unit’s power.
• After conforming that all the units are switched on, turn the PC power ON.
• Place the tube leading to the pump, and open the drain valve, which is located in the right-hand side of the pump, select the pump that you are using, then press Purge ON. Once the air bubbles are removed /new solvent comes through the pump then press purge off to stop purging.
• Ensure that the chromeleon server is ON.
• Double-click on the chromeleon icon placed on the desktop.
• The main menu includes the username and password. Give the appropriate username and password.
• If your username and password are correct chromeleon open the chromeleon browser screen.
• Open the QC0TEST2_local (Data source name)/HP 407 or HP 408 /Panel folder for HP407 instrument use.
• Open the panel for HP407/408 instrument and check that the pump, sampler, column oven & UV detectors are connected to the chromeleon software. Use the pump settings for flow changing or purging. Use the sampler settings for wash, prime syringe or wash vial selection. Use the column oven settings for the temperature setting of the column oven. Use the detector settings for the lamp ON/OFF command.
• Make a program file as per the procedure given below:

1. Click on File-new-program File and then press ok.
2. Select the instrument name and then press next.
3. Give the temperature of the desired value for the column oven and then press next.
4. Give the desired temperature for the sampler chillier and then press next.
5. In pump options, select the Multi-step gradient or isocratic that you want and then press next.
6. In the sampler option given the desired setting for the sampler and then press next.
7. In the Acquisition option, select the pump pressure acquisition, column oven acquisition & UV acquisition that you want to desire. Give the run time in this menu & press next –next.
8. In UV option, select the desired channel and wavelength according to your requirement and then press next.
9. Select the review and press finish to review the programme file. Select the save and press finish to save the programme. Press Ctrl +S to save the programme. Select the desired path for saving the programme.

• Make a method file as per the procedure given below

1. Click on File-new-Method File and then press ok.
2. Give the minimum area for detection & then select File–Save as a command.
3. Then save the method file at an appropriate location.

ALSO READ: Qualification of High Performance Liquid Chromatography

• Make a sequence file as per the procedure given below:

1. Click on File-new-Sequence (Using Wizard) and then press Next.
2. Select the instrument name and then press next. Give the temperature of the desired value for the column oven and then press next.
3. Select the no. of vials, number of injections per vial, Start position,n and then press apply command next. After giving apply command, you can also see the rack preview and then press next.
4. Generate standards from a template or import them via clip board or another sequence menu select the no. of vials, number per vial, start position and then press the apply command. After giving the apply command, you can also see the rack preview. Then press next.
5. Select the appropriate Program file, Quantitation Method, Report file & preferred channel for that sequence. Then press next.
6. Given the sequence name, sequence title, Data source name, Sequence Directory (Sequence path) and then press finish and done.
7. After pressing done command, your blank sequence will be opened. Insert or append samples according to your requirements. Then give the sample names, type, position, injection volume, Programme file, Method file, comment, Sample ID, Weight, Dilation Factor according to your preparation and requirement.
8. Then press Ctrl + S for saving the sequence.

• Run a sequence

1. Select the sequence, then press Ctrl+B (Batch-Edit).Then add the sequence in batch list. Then press ready check to check the system. If any errors come in ready check command, then resolve that error. If the warning is coming, then it doesn’t matter in running the sequence.
2. If ready check is ok, then press start to start the sequence. When you press start button then sample no.1 which is in the sequence will automatically running and it’s colour will become green in running mode. One by one all the samples in the sequence are finished automatically and all the sample status is finished after completion of sequence.

• QNT Method editing:

1. For open a chromatogram, double-click on the samples which have finished status in sequence.
2. When you double-click on the samples that have finished status, the default report format that you have selected in that sequence will be opened up. Then select the QNT-Editor mode for modifying a method file.
3. In QNT –Editor mode, select the detection sheet for giving integrating parameters such as minimum area, Valley to Valley on, Inhibit integration ON/OFF. Fronting sensitivity factor, Tailing Sensitivity Factor etc.
4. Select the peak table sheet for peak naming and giving Retention Window.
5. If your integration is proper in all the samples, then press File -> Save command to save the QNT method file.

• Reporting:

1. If your integration is proper in all the samples of sequence, then select the Printer Lay out mode for viewing & printing of the report.
2. In Printer Lay Out mode, if you want to see the Integration of a current sample, select the integration sheet, if you want to see the peak analysis report select the Peak analysis sheet. If you want to see the RSD of standards injected in a sequence, select the Summary Sheet for RSD variation of standards.
3. For printing of report, select the appropriate sheet you want to print & then press the file -> Printing (Ctrl +P) command for printing.

ANNEXURES

Nil

ABBREVIATIONS

HPLC: High-Performance Liquid Chromatograph

UV: Ultraviolet

RSD: Relative Standard Deviation

API: Active Pharmaceutical Ingredient

STP: Standard Test Procedure

SOP: Standard Operating Procedure

No. : Number

Dept. : Department

QC: Quality Control

Tr.: Trainee

Asst. : Assistant

REVISION HISTORY

Nil

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