ingredients (APIs) and finished drug products.

2. Scope

This SOP applies to the Analytical Method Development (AMD) department and is relevant to impurity profiling studies for new drug substances, generics, and stability testing in line with ICH Q3A and Q3B guidelines.

3. Responsibilities

• Analytical Chemist: Develops the method, performs impurity spiking and forced degradation, and evaluates specificity and sensitivity.
• Reviewer: Verifies method performance including resolution, linearity, and detection limits.
• QA Officer: Reviews documentation and ensures compliance with regulatory expectations.
• Head – AMD: Approves the method for validation and regulatory submission.

4. Accountability

The Head of AMD is accountable for ensuring that all related substances methods are scientifically sound, stability-indicating, and suitable for detecting impurities at regulatory thresholds (e.g., 0.05%, 0.1%, 0.2%).

5. Procedure

5.1 Preliminary Planning and Impurity Identification

1. Review:
   • Synthetic process impurities
   • Degradants from forced degradation
   • Known pharmacopoeial impurity standards
2. Set reporting, identification, and qualification thresholds based on ICH guidelines.
3. Document impurity profiles in Annexure-1: Impurity Planning Sheet.

5.2 Chromatographic Method Development

1. Start with reversed-phase C18 column (150 mm × 4.6 mm, 5 µm).
2. Select mobile phase:
   • A: Aqueous buffer (e.g., phosphate pH 3–4)
   • B: Acetonitrile or methanol
   • Use gradient elution if impurities vary in polarity
3. Wavelength: Choose based on impurity UV absorbance or API λmax.
4. Flow rate: 1.0 mL/min; Injection: 20 µL.
5. Record in Annexure-2: Chromatographic Conditions Log.

5.3 Preparation of Solutions

1. API Standard: Prepare 100 µg/mL solution.
2. Individual Impurity Standards: Prepare stock and dilute to 0.05%, 0.1%, 0.2% w.r.t API.
3. Spiked Sample: Mix API and impurities to assess resolution and quantification.
4. Forced Degradation Samples: Prepare samples under acid, base, oxidation, heat, and light.
5. Document preparation in Annexure-3: Solution Preparation Log.

5.4 Evaluation of System Suitability and Specificity

1. Check:
   • Resolution between API and nearest impurity ≥ 2.0
   • Tailing factor ≤ 2.0
   • Plate count ≥ 2000
2. Overlay chromatograms of blank, API, spiked, and degradation samples.
3. Confirm no co-elution of impurities or degradants.
4. Record data in Annexure-4: System Suitability Report.

5.5 Determination of Detection and Quantification Limits

1. Inject decreasing concentrations of impurities (e.g., 0.1%, 0.05%, 0.02%, 0.01%).
2. Determine LOD (S/N ≥ 3) and LOQ (S/N ≥ 10).
3. Ensure acceptable RSD at LOQ level (≤10%).
4. Document results in Annexure-5: LOD and LOQ Summary.

5.6 Linearity and Accuracy

1. Perform linearity from LOQ to 150% level for each impurity.
2. Acceptance criteria: R² ≥ 0.999 for each impurity.
3. Accuracy: Spike impurities at 50%, 100%, and 150% levels and recover (acceptable: 80–120%).
4. Document in Annexure-6: Linearity and Recovery Log.

5.7 Robustness and Method Finalization

1. Evaluate robustness with small variations:
   • ±0.1 mL/min flow rate
   • ±2°C column temperature
   • ±5% in mobile phase ratio
2. Confirm method remains suitable under altered conditions.
3. Finalize method with all performance parameters documented in Annexure-7: Final Method Summary.

6. Abbreviations

• HPLC: High-Performance Liquid Chromatography
• LOD: Limit of Detection
• LOQ: Limit of Quantification
• ICH: International Council for Harmonisation
• SOP: Standard Operating Procedure

7. Documents

1. Impurity Planning Sheet – Annexure-1
2. Chromatographic Conditions Log – Annexure-2
3. Solution Preparation Log – Annexure-3
4. System Suitability Report – Annexure-4
5. LOD and LOQ Summary – Annexure-5
6. Linearity and Recovery Log – Annexure-6
7. Final Method Summary – Annexure-7

8. References

• ICH Q3A (R2) – Impurities in New Drug Substances
• ICH Q3B (R2) – Impurities in New Drug Products
• ICH Q2 (R1) – Validation of Analytical Procedures
• USP <621> – Chromatography

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: Impurity Planning Sheet

Impurity Name	Type	Threshold (%)	Standard Available
Impurity A	Process	0.1%	Yes
Impurity B	Degradant	0.2%	Yes

Annexure-2: Chromatographic Conditions Log

Column	Mobile Phase A	Mobile Phase B	Gradient Program	Flow Rate
C18, 150×4.6 mm	Phosphate buffer pH 3.5	Acetonitrile	10–90% B in 15 min	1.0 mL/min

Annexure-3: Solution Preparation Log

Solution ID	Component	Concentration	Prepared By
SPK-001	API + Imp A + Imp B	100 + 0.1 + 0.2 µg/mL	Rajesh Kumar

Annexure-4: System Suitability Report

Parameter	Observed	Specification	Status
Resolution (API vs Imp A)	2.4	≥ 2.0	Pass

Annexure-5: LOD and LOQ Summary

Impurity	LOD (%)	LOQ (%)	RSD @ LOQ
Impurity A	0.015	0.05	6.2%

Annexure-6: Linearity and Recovery Log

Impurity	R²	Recovery (%)	Status
Impurity B	0.9991	98.7%	Pass

Annexure-7: Final Method Summary

Column	Mobile Phase	Gradient	Detection	Run Time
C18	Buffer:ACN	10–90% B in 15 min	UV @ 245 nm	20 min

Revision History:

Revision Date	Revision No.	Details	Reason	Approved By
04/05/2025	2.0	Included gradient method and expanded impurity recovery annexures	Annual Review