Characterization of products formed by forced degradation of Amlodipine Besylate using LC/MS/MS

Significance of the Topic

Amlodipine besylate is a widely used dihydropyridine calcium channel blocker in cardiovascular therapy. Understanding its degradation behavior under stress conditions is essential for ensuring drug safety, defining shelf life, and guiding formulation and manufacturing processes. Forced degradation studies reveal potential impurities and degradation pathways that impact efficacy and toxicity.

Study Objectives and Overview

This work aimed to generate and characterize the degradation products of amlodipine besylate induced by acidic, basic, oxidative, and thermal stress. Using a high-sensitivity triple quadrupole LC/MS/MS system (Shimadzu LCMS-8080), the study identified major degradation species, quantified degradation extent, and elucidated structural features of the products.

Methodology and Instrumentation

Stress Conditions:

• Acidic hydrolysis: 5 M HCl at 80 °C for 6 h (resulting in ~60 % degradation)
• Basic hydrolysis: 1 M NaOH at 80 °C for 2 h (~25 % degradation)
• Oxidative: 30 % H₂O₂ at 80 °C for 6 h (~20 % degradation)
• Thermal: 80 °C for 48 h (minimal, ~0.25 % degradation)

Sample Preparation:
Amlodipine stock solutions (1000 ppm) were mixed with stress reagents, heated, neutralized or diluted to 200 ppm, and analyzed. Thermal samples were directly prepared from oven-aged solid drug.

Used Instrumentation:

• Shimadzu LCMS-8080 triple quadrupole mass spectrometer with Hot Source Induced Desolvation (HSID)
• Shim-pack XR ODS column (75 × 3 mm, 2.2 µm)
• UV detection at 238 nm
• Mobile phase: 10 mM ammonium acetate (pH 7.0) and acetonitrile gradient
• ESI interface: nebulizing gas 2 L/min, heating gas 15 L/min, curtain gas 2.4 L/min

Main Results and Discussion

LC/MS/MS chromatograms and precursor/product ion scans revealed distinct degradation profiles:

• Acidic and oxidative conditions produced a common impurity at m/z 407, matching the European Pharmacopeia Impurity D.
• Basic hydrolysis generated four primary products (m/z 395, 349, 381, 351) with unique fragmentation patterns.
• Product ions at m/z 180 and 167 were consistently observed across all stress types, indicating conserved core fragments.

Oxidative degradation also shared some product ions with acidic degradation (e.g., m/z 230), suggesting similar oxidative pathways.

Benefits and Practical Applications

The rapid LC/MS/MS approach enables simultaneous detection and structural inference of multiple low-level degradation compounds without lengthy isolation. This supports stability-indicating method development, impurity profiling, and regulatory compliance for drug substances and formulations.

Future Trends and Opportunities

Advances in high-resolution MS and data-driven spectral interpretation will further improve sensitivity and confidence in degradation product identification. Integration of in-silico prediction tools with LC/MS workflows could accelerate impurity risk assessment and formulation optimization.

Conclusion

A robust LC/MS/MS method was established for forced degradation analysis of amlodipine besylate. Distinct degradation products were mapped under acidic, basic, oxidative, and thermal stress, with major impurities structurally characterized. The workflow offers fast, sensitive, and informative data for stability studies and quality control.

Reference

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3. Patil K.R. et al. Stability-Indicating LC Method for Amlodipine and Olmesartan. J Chromatogr Sci, 2010.
4. ICH Q1A(R2) Stability Testing of New Drug Substances and Products, 2003.