1) SOP for Operating High-Performance Liquid Chromatography (HPLC)

2) Purpose

The purpose of this Standard Operating Procedure (SOP) is to provide detailed instructions for the calibration, operation, and maintenance of High-Performance Liquid Chromatography (HPLC) equipment, as well as for sample preparation and data analysis.

3) Scope

This SOP applies to all personnel involved in the use of HPLC within the pharmaceutical manufacturing and quality control areas.

4) Responsibilities

• Operators are responsible for calibrating, operating, and maintaining HPLC equipment according to this SOP.
• Supervisors are responsible for overseeing the use of HPLC and ensuring compliance.
• Quality Assurance (QA) personnel are responsible for verifying that procedures are followed correctly and addressing any deviations.

5) Procedure

1. Preparation
   1. Ensure the HPLC area is clean and free of contaminants.
   2. Verify the availability of reagents, solvents, and calibration standards.
   3. Check the cleanliness and readiness of the HPLC equipment.
2. Calibration
   1. Turn on the HPLC system and allow it to equilibrate as per the manufacturer’s instructions.
   2. Prepare calibration standards according to the method requirements.
   3. Run calibration standards to establish a calibration curve.
   4. Document the calibration process in the calibration logbook.
3. Sample Preparation
   1. Prepare samples according to the method requirements, ensuring correct concentrations and volumes.
   2. Filter and degas samples if necessary.
   3. Label samples clearly and document preparation details in the batch record.
4. Operation
   1. Load samples into the HPLC autosampler or manually inject samples as per the method.
   2. Start the HPLC run and monitor the chromatogram for any abnormalities.
   3. Ensure that system suitability criteria are met before analyzing samples.
   4. Document all observations and any adjustments made during the run in the relevant logbook.
5. Data Analysis
   1. Analyze the chromatographic data using appropriate software.
   2. Quantify the analyte concentrations using the calibration curve.
   3. Document the results in the batch record and ensure all calculations are accurate.
6. Troubleshooting
   1. If the HPLC system malfunctions, stop the operation immediately and troubleshoot the issue.
   2. Refer to the manufacturer’s manual for troubleshooting guidelines.
   3. Document the issue and the corrective actions taken in the maintenance logbook.
   4. Resume operation once the issue is resolved and the system is functioning properly.
7. Cleaning and Maintenance
   1. Flush the HPLC system with appropriate solvents to prevent cross-contamination.
   2. Clean all parts thoroughly as per the manufacturer’s instructions.
   3. Document the cleaning process in the cleaning logbook.
   4. Perform regular maintenance checks and document them in the maintenance logbook.

6) Abbreviations, if any

• SOP: Standard Operating Procedure
• QA: Quality Assurance
• HPLC: High-Performance Liquid Chromatography

7) Documents, if any

• Calibration Logbook
• Cleaning Logbook
• Maintenance Logbook
• Batch Record

8) Reference, if any

• Current Good Manufacturing Practice (cGMP) guidelines
• Manufacturer’s operation and maintenance manual

9) SOP Version

Version 1.0