validation activities. It ensures the column performs consistently with defined chromatographic parameters and meets quality standards for reproducibility and accuracy.

2. Scope

This SOP is applicable to all new and used HPLC columns procured or reused in the Analytical Method Development (AMD) department. It includes qualification for reversed-phase, normal-phase, ion-exchange, and size-exclusion columns.

3. Responsibilities

• Analyst: Responsible for conducting the qualification test as per approved protocol and maintaining logs.
• Section Head: Reviews column qualification data and approves suitability for method use.
• QA Representative: Ensures traceability and compliance with applicable guidelines.

4. Accountability

The Head of Analytical Method Development is accountable for ensuring that all HPLC columns are qualified, documented, and maintained as per cGMP and ICH Q2(R1) guidelines before deployment.

5. Procedure

5.1 Column Identification and Documentation

1. Record column details in the HPLC Column Inventory Log (Annexure-1): manufacturer, type, dimensions, particle size, lot number, and serial number.
2. Assign a unique Column ID code for internal tracking and traceability.

5.2 Qualification Criteria

Each column must demonstrate acceptable performance for the following parameters using a test mixture or designated standard solution:

1. Retention time (RT) reproducibility (±2%)
2. Theoretical plate number (N) within specification limits
3. Tailing factor (Tf) ≤ 2.0
4. Resolution (Rs) ≥ 2.0 between critical peaks

5.3 Qualification Procedure

1. Condition the new column per the manufacturer’s recommendation using mobile phase or buffer.
2. Inject a qualification standard mixture containing analytes with known retention properties.
3. Perform six replicate injections and record chromatographic parameters.
4. Document and evaluate data against acceptance criteria.

5.4 Qualification of Used Columns

1. Used columns must be evaluated before reuse after extended storage or use in sensitive methods.
2. Flush with appropriate solvent to remove residuals.
3. Perform the same qualification test used for new columns.
4. Document requalification results in Column Qualification Log (Annexure-2).

5.5 Rejection and Disposal

• If column fails to meet acceptance criteria in two consecutive tests, mark it “Unqualified”.
• Tag and move rejected column to the designated area for return, disposal, or reclamation as per SOP.

6. Abbreviations

• HPLC: High-Performance Liquid Chromatography
• RT: Retention Time
• N: Number of Theoretical Plates
• Tf: Tailing Factor
• Rs: Resolution

7. Documents

1. Annexure-1: HPLC Column Inventory Log
2. Annexure-2: HPLC Column Qualification Log
3. Column Qualification Summary Sheet

8. References

• ICH Q2(R1): Validation of Analytical Procedures
• USP General Chapter <621> – Chromatography
• 21 CFR Part 211

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name	Ritika Sinha	Pradeep Shetty	Dr. Anamika Deshmukh
Designation	AMD Analyst	QA Reviewer	Head – Analytical Method Development
Department	Analytical Method Development	Quality Assurance	Analytical Method Development

11. Annexures

Annexure-1: HPLC Column Inventory Log

Column ID	Type	Dimension	Lot No.	Make	Date Received
COL-HPLC-001	C18	250×4.6mm, 5µ	L202301	Waters	05/04/2025

Annexure-2: HPLC Column Qualification Log

Column ID	RT (min)	Theoretical Plates	Tailing Factor	Resolution	Status
COL-HPLC-001	5.84	8495	1.08	2.21	Qualified

Revision History

Revision Date	Revision No.	Details	Reason	Approved By
01/06/2025	2.0	Enhanced documentation and disposal instructions	Periodic review	Dr. Anamika Deshmukh
12/07/2022	1.0	Initial Issue	New SOP	QA Head