using advanced instrumental techniques such as LC-MS/MS, PDA, FTIR, and NMR.

2. Scope

This SOP applies to all drug substances and drug products tested within the Analytical Method Development (AMD) department, where degradation peaks are observed but not identified during routine HPLC analysis.

3. Responsibilities

• Analytical Scientist: Plans the identification strategy, selects instruments, and interprets spectral data.
• Instrument Analyst: Operates LC-MS, FTIR, and other instruments under GMP environment.
• QA Officer: Verifies documentation, compliance with ICH Q3A/B guidelines, and ensures traceability.
• Head – AMD: Approves reports and identification conclusions for submission to regulatory authorities.

4. Accountability

The Head of AMD is accountable for ensuring that all unknown degradation products are systematically identified and qualified using validated analytical techniques.

5. Procedure

5.1 Initial Observation and Qualification

1. During forced degradation or stability studies, if an unidentified peak is detected (typically >0.05% for APIs or >0.1% for products), initiate identification protocol.
2. Assign a unique peak code and record chromatographic data including retention time, area %, and peak shape.
3. Record the observation in Annexure-1: Degradation Observation Log.

5.2 Isolation of Unknown Peak

1. Enrich the sample using:
   • Extended exposure (higher degradation)
   • Preparative HPLC (if required)
2. Confirm presence and separation using PDA detector and calculate peak purity angle vs threshold.

5.3 Identification via LC-MS/MS

1. Inject sample into LC-MS with conditions optimized for:
   • Electrospray ionization (ESI) or APCI mode
   • Positive/negative polarity scan
2. Obtain:
   • Molecular ion (M⁺ or [M+H]⁺)
   • Fragmentation pattern
   • Mass-to-charge ratio (m/z)
3. Compare spectra with mass spectral libraries or theoretical fragments of API.

5.4 Confirmatory Techniques

1. If required, use FTIR to detect functional groups and NMR (1H/13C) for structural assignment.
2. Match observed shifts to theoretical structures.
3. Document spectra and results in Annexure-2: Structural Characterization Report.

5.5 Qualification and Reporting

1. Assess toxicological significance of the impurity per ICH Q3A/Q3B.
2. Determine whether qualification is needed based on identification threshold.
3. Include all results and spectra in identification report.

6. Abbreviations

• LC-MS: Liquid Chromatography – Mass Spectrometry
• FTIR: Fourier Transform Infrared Spectroscopy
• NMR: Nuclear Magnetic Resonance
• PDA: Photodiode Array
• SOP: Standard Operating Procedure

7. Documents

1. Degradation Observation Log – Annexure-1
2. Structural Characterization Report – Annexure-2

8. References

• ICH Q3A(R2) – Impurities in New Drug Substances
• ICH Q3B(R2) – Impurities in New Drug Products
• ICH Q1A(R2) – Stability Testing Guidelines
• ICH Q2(R2) – Validation of Analytical Procedures

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: Degradation Observation Log

Sample ID	RT (min)	Peak Area (%)	Detection Date	Observed By
DEG-PROD-024	7.58	0.14%	15/05/2025	Sunita Reddy

Annexure-2: Structural Characterization Report

Peak Code	m/z	Major Fragments	Proposed Structure	Identification Method
UK-058	321.1	145.0, 203.0	Hydroxy degradation of API	LC-MS/MS

Revision History:

Revision Date	Revision No.	Details	Reason	Approved By
04/05/2025	2.0	Added requirement for confirmatory techniques and annexure formats	Compliance with ICH Q3A/Q3B