Reproducing the TCM Method for the Characteristic Chromatogram of Guilin Xiguashuang

Importance of the Topic

Characteristic chromatograms are essential for quality control of traditional Chinese medicines (TCMs) as specified in the Chinese Pharmacopeia (ChP). Ensuring consistent retention times and peak patterns across different liquid chromatography (LC) platforms is critical for regulatory compliance and reliable analytical results.

Aims and Study Overview

The study aimed to reproduce the ChP method for obtaining the characteristic chromatogram of Guilin Xiguashuang using an Agilent 1290 Infinity II LC system equipped with Intelligent System Emulation Technology (ISET) and an InfinityLab Poroshell HPH-C18 column. Emulation of Waters ACQUITY UPLC H-Class and I-Class systems was evaluated to confirm cross-platform method transferability.

Methodology and Instrumentation

The method followed the ChP monograph for Guilin Xiguashuang, using:

• Mobile phase A: 0.2% phosphoric acid + 0.2% triethylamine
• Mobile phase B: acetonitrile
• Gradient program from 20% to 95% B over 43 minutes, 45-minute run time, 6-minute post-run
• Flow rate: 0.2 mL/min; column temperature: 30 °C; injection volume: 1 µL; detection: UV at 254 nm

Five InfinityLab Poroshell 120 columns were screened, including EC-C18, SB-C18, CS-C18, Aq-C18, and HPH-C18 chemistries. The Agilent 1290 Infinity II modules comprised a binary pump, multisampler, column compartment, and diode array detector, controlled by OpenLab CDS 2.8 with ISET 4.

Main Results and Discussion

Column screening demonstrated that the HPH-C18 column provided the same elution order as the original Waters UPLC BEH C18 phase and delivered all relative retention times (RRTs) within ±10% of ChP requirements. Emulation of Waters ACQUITY UPLC H-Class yielded complete compliance, while I-Class emulation produced one peak (peak 12) slightly outside tolerance due to differences in dwell volume.

Adjusting dwell volume by configuring mixer and sample loop settings within ISET improved separation of critical peaks 7 and 8. Using a 100 µL sample loop in H-Class emulation enhanced resolution (Rs7,8 increased from 1.4 to 1.9), demonstrating the importance of instrument parameter matching for accurate method transfer.

Benefits and Practical Applications

• Enables ChP-compliant characteristic chromatogram analysis of Guilin Xiguashuang on Agilent LC platforms without modifying the original method.
• Ensures reproducibility of RRTs and peak resolution across different vendor systems.
• Reduces need for method redevelopment when switching instruments, saving time and resources in pharmaceutical quality control laboratories.

Future Trends and Opportunities

Advancements in system emulation technologies will facilitate broader method transfer among various LC platforms. Further optimization of dwell volume and mixer configurations could enhance resolution for other complex TCMs. Integration of automated parameter checks and AI-driven method adaptation may streamline cross-platform compliance and accelerate analytical workflows.

Conclusion

The Agilent 1290 Infinity II LC with ISET and InfinityLab Poroshell HPH-C18 column successfully reproduced the ChP characteristic chromatogram for Guilin Xiguashuang, meeting all RRT specifications when emulating Waters ACQUITY UPLC H-Class. Proper configuration of dwell volume and sample loop settings is crucial for accurate method transfer, enabling reliable quality control across LC systems.

Reference

China National Medical Products Administration and National Health Commission. Chinese Pharmacopeia. 2025 Edition, Vol 1: Guilin Xiguashuang.