Analysis of Saponins in Notoginseng using Agilent Poroshell 120, 4 μm Columns

Significance of the Topic

Notoginseng saponins such as notoginsenoside R1, ginsenosides Rg1, Re and Rb1 are key bioactive constituents in traditional Chinese medicine for treating blood disorders and require reliable quantification by HPLC for quality control.

Objectives and Study Overview

The study aimed to adapt the China Pharmacopoeia regulated HPLC method for notoginseng saponin analysis to an Agilent Poroshell 120 superficially porous 4 µm column, compare it against conventional 3.5 and 5 µm totally porous columns and a 2.7 µm Poroshell column, and to shorten analysis time via a shorter column format and higher flow rates.

Methodology and Instrumentation

• Agilent 1290 Infinity LC System with Binary Pump, Autosampler, Thermostatted Column Compartment and Diode Array Detector.
• Columns tested: Poroshell 120 EC-C18 4.6×150 mm 4 µm; Poroshell 120 EC-C18 4.6×100 mm 4 µm; Poroshell 120 EC-C18 4.6×100 mm 2.7 µm; ZORBAX Eclipse Plus C18 4.6×100 mm 3.5 µm; ZORBAX Eclipse Plus C18 4.6×150 mm 5 µm.
• Mobile phase: water (A) and acetonitrile (B) with gradient programs optimized for each column length.
• Conditions: 30 °C, flow rates of 1.0 or 1.5 mL/min, injection volumes 10 µL or 6.7 µL, detection at 203 nm.

Main Results and Discussion

• Transition to the 4 µm Poroshell column improved resolution of Rg1/Re from Rs=1.51 to Rs=2.31 and halved peak width compared with the 5 µm column.
• The 4 µm format achieved efficiency similar to the 3.5 µm column but with lower backpressure, suitable for low-pressure instruments.
• Shortening to a 100 mm column reduced run time by over 30% (60 to 40 min) with proportional injection volume adjustment, and 30 min runs at 1.5 mL/min remained acceptable (Rs≈1.62).

Benefits and Practical Applications

• Seamless replacement of existing CHP method without parameter adjustments.
• Faster throughput and reduced solvent consumption supports cost-effective quality control.
• Compatibility with instruments limited to 200 bar backpressure expands accessibility.

Future Trends and Potential Applications

• Increasing adoption of superficially porous particles for high-throughput and green chromatography.
• Integration with mass spectrometry for enhanced selectivity and sensitivity.
• Development of miniaturized and automated analytical workflows for routine QA/QC.

Conclusion

The study demonstrates that an Agilent Poroshell 120 EC-C18 4 µm column fully meets China Pharmacopoeia requirements for notoginseng saponin analysis, offering improved resolution, lower solvent use and faster turnaround on both high- and low-pressure LC systems.

References

• Notoginseng, China Pharmacopoeia, 2015 edition.
• Zhu S.; Zou K.; Fushimi H.; Cai S.; Komatsu K. Comparative study on triterpene saponins of ginseng drugs. Planta Medica 2004, 70, 666–677.