Analysis of Polar Compounds in Plant Material

Significance of the Topic

Polar phytochemicals such as stachydrine are key bioactive constituents in Traditional Chinese Medicine herb Leonurus japonicus. Their high polarity and presence in a complex plant matrix make analysis by reversed-phase chromatography challenging. A robust, fast method for quantitation of stachydrine supports quality control, regulatory compliance, and research on efficacy of herbal supplements.

Study Objectives and Overview

This work aimed to develop both isocratic and gradient hydrophilic interaction chromatography (HILIC) methods using an Agilent InfinityLab Poroshell 120 HILIC-Z column for direct separation and quantitation of stachydrine in L. japonicus extract. Detection was achieved with evaporative light scattering (ELSD) to compensate for weak UV absorbance.

Applied Instrumentation

• Agilent 1260 Infinity II binary pump
• Agilent 1260 Infinity II vial sampler
• Agilent 1260 Infinity II multicolumn thermostat
• Agilent 1290 Infinity II ELSD
• Agilent OpenLAB software
• Ultralow dispersion kit

Methodology

Reagents included HPLC-grade acetonitrile, 10 mM ammonium acetate buffer, and purified water. Standards of stachydrine hydrochloride were prepared at 0.5 mg/mL in 70 % ethanol. Plant extract was obtained by refluxing 1 g of dried L. japonicus powder in 70 % ethanol for 2 hours, adjusting the weight loss with fresh solvent, and filtering through a 0.2 µm PTFE membrane. Chromatographic conditions: Poroshell 120 HILIC-Z, 2.1 × 100 mm, 2.7 µm; mobile phase A: 10 mM ammonium acetate in water; phase B: acetonitrile; flow rate: 0.30 mL/min; column temperature: 30 °C; injection volume: 2 µL. ELSD settings: nebulizer and evaporator temperature 40 °C, gas flow 1.6 SLM, data rate 40 Hz.

Main Results and Discussion

Under isocratic conditions (90 % acetonitrile, 10 % buffer) a complete separation of stachydrine from co-eluting polar impurities was achieved in 8 minutes. Gradient elution (95 % to 60 % B over 10 minutes) provided similar resolution with minimal run time extension. Chromatograms showed a clear, symmetric stachydrine peak free from matrix interferences.

Benefits and Practical Applications

The HILIC-Z platform enables direct injection of crude plant extracts without derivatization or extensive cleanup, reducing analysis time and solvent usage. Low backpressure of superficially porous particles supports high throughput. The method is suited for routine quality control in herbal product manufacturing and research laboratories.

Future Trends and Potential Uses

Expansion of HILIC techniques to profile additional polar alkaloids and glycosides in botanical matrices is anticipated. Integration with mass spectrometry detection will enhance selectivity and sensitivity for multi-component analysis. Development of automated sample preparation could further streamline workflows.

Conclusion

The developed HILIC method on the Agilent InfinityLab Poroshell 120 HILIC-Z column provides rapid, reliable quantitation of stachydrine in Leonurus japonicus extracts. Both isocratic and gradient protocols deliver excellent peak shape and resolution, supporting robust analytical workflows in natural product research and quality assurance.

References

No formal literature references were provided in the source document.