SOP for HPLC Analysis & Documentation

OBJECTIVE

To lay down a procedure for analysis to be carried out by HPLC and its documentation.

SCOPE

This procedure is applicable for analysis to be carried out by High-Performance Liquid Chromatography in Quality Control Laboratory.

RESPONSIBILITY

Execution: Chemist – QC department.

Checking: Chemist and above – QC department

PROCEDURE

• Collect the sample along with the protocol/datasheet for analysis from a section in charge.
• Read the STP and protocol/datasheet carefully for tests to be performed by HPLC, reagents used, HPLC parameters, System suitability parameters, standard preparation, and test preparation etc.
• Select the HPLC system and column (as specified in STP/protocol/datasheet) for analysis.
• Use only HPLC-grade solvents in the preparations.

Preparation of Mobile Phase:

• Use the appropriate grade of reagents as stated in the STP.
• Weigh the reagents accurately, dissolve and dilute them with solvent as per the individual STP.
• Adjust the pH with the appropriate reagent as per the STP.
• Mix thoroughly the buffer and solvents with the ratio mentioned in the individual STP and adjust the pH if it is mentioned in the STP.
• Follow the same procedure for the gradient buffer also and do the preparation separately.
• Prepare the quantity of buffer according to the sample load.
• Filter the mobile phase with appropriate membrane filter paper.
• Do the filtration without delay.
• Degas the mobile phase for about 5 minutes by using a sonicator.
• Use the mobile phase within 5 days from the date of preparation.
• Label the mobile phase with the following information, product name, and Test prepared on and use before as per the annexure-QCD30/A01-00.
• If any visual impurities are found within 5 days discard the mobile phase and prepare freshly.

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The sequence of Injections:

1. System suitability (if applicable)
2. Standard Solution (In replicates, if mentioned in STP)
3. Sample Solution (In replicates, if mentioned in STP)

• Before starting the analysis trial run carried out is required to check the Baseline noise, RT, system suitability parameters, etc
• On completion, flush the system with lab water for 15 minutes followed by the organic solvent used in the mobile phase: water (50: 50)
• After completing the analysis of one sample, make the entries in the Column usage log book and in the instrument log book.

ABBREVIATION

SOP - Standard Operating Procedure

STP - Standard test procedure

QC - Quality Control

HPLC - High-Performance Liquid Chromatography

ANNEXURE

Nil

REVISION HISTORY

Nil

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