Extending Column Lifetime in Pharmaceutical Methods with High pH-Stable InfinityLab Poroshell HPH Chemistries

Significance of the Topic

High-performance liquid chromatography (HPLC) is indispensable in pharmaceutical method development and quality control. Traditional silica-based columns degrade rapidly under high-pH conditions, limiting analysis of basic compounds and reducing column lifetime. The Agilent InfinityLab Poroshell 120 HPH-C18 column introduces an organically modified superficially porous material that remains stable across a broad pH range, offering enhanced robustness and prolonged service life.

Objectives and Study Overview

The study aims to evaluate the chemical and mechanical stability of the sub-3 μm InfinityLab Poroshell 120 HPH-C18 column under typical and stress conditions. Comparative experiments were conducted using low-pH and high-pH mobile phases, and against other superficially porous, hybrid, and conventional columns. Key performance metrics included retention time stability, peak shape, column efficiency, and lifetime under accelerated degradation protocols.

Methodology and Used Instrumentation

• Instrumentation: Agilent 1260 Infinity LC system comprising G1312B Binary Pump, G1367C Automatic Sampler, G1316C Thermostatted Compartment, and G4220A DAD (254 nm).
• Columns tested: InfinityLab Poroshell 120 HPH-C18 (2.1×50 mm and 4.6×50 mm, 2.7 μm), Poroshell 120 EC-C18, and competitive superficially porous and hybrid columns.
• Analytes: mixtures of acidic, neutral, and basic compounds (e.g., quinine, nortriptyline, phenol, acetophenone).
• Mobile phases: 0.2% formic acid; 0.1% ammonium hydroxide (pH 10.5); 10 mM ammonium bicarbonate (pH 10); 30 mM phosphate buffer (pH 8) in 40% methanol.
• Tests: Gradient and isocratic injections, van Deemter measurements at 25 °C, stress protocols at elevated temperatures (50–65 °C).

Main Results and Discussion

• Low-pH performance: Over 2,000 injections with 0.2% formic acid showed consistent retention and peak shape, confirming robustness under routine acidic conditions.
• High-pH performance: In 0.1% ammonium hydroxide (pH 10.5), the HPH-C18 column endured >1,500 injections with stable retention times; minor peak-shape changes occurred for early eluting analytes.
• Ammonium bicarbonate buffer (pH 10): Over 2,000 injections, HPH-C18 maintained retention; a non-Agilent column exhibited nortriptyline coelution by 500 injections, indicating superior selectivity and stability of HPH-C18.
• Stress degradation (10 mM bicarbonate, 50 °C): Two-step isocratic test revealed HPH-C18 lost 10% efficiency after ~4,000 mL buffer; EC-C18 lost 25% at 500 mL; non-Agilent hybrid and superficially porous columns degraded faster.
• Phosphate buffer at pH 8 (65 °C): HPH-C18 retained >90% efficiency after 3,000 mL; EC-C18 lost 40% at 1,000 mL, demonstrating enhanced high-pH chemical stability.
• Van Deemter analysis: Both HPH-C18 and EC-C18 exhibited reduced plate heights of ~1.6, confirming comparable packing quality and efficiency.

Benefits and Practical Applications

• Extended column lifetime under high-pH conditions reduces operating costs and downtime.
• Improved robustness for analysis of basic pharmaceuticals enhances method reliability.
• Compatibility with volatile buffers and LC/MS workflows facilitates seamless integration into MS-based assays.
• Low back pressure and high efficiency enable faster separations with sub-3 μm performance.

Future Trends and Possibilities

Continued innovation in bonding chemistries and superficially porous particle design may further extend pH stability beyond pH 10, enabling direct analysis of highly basic or unstable compounds. Integration with automated high-throughput platforms and advanced buffer systems avoiding phosphate or carbonate could streamline pharmaceutical screening and impurity profiling. Development of even thinner porous shells and novel organic modifiers may enhance mass-transfer kinetics and column longevity.

Conclusion

The Agilent InfinityLab Poroshell 120 HPH-C18 column successfully combines the benefits of superficially porous particles with high-pH chemical stability. It significantly outperforms conventional EC-C18, hybrid, and competing superficially porous columns in lifetime and retention stability under both routine and stress conditions. This technology offers a robust solution for modern pharmaceutical HPLC and LC/MS applications, improving method reliability and throughput.

References

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