Analysis of Hydroxysafflor yellow A in Safflower

Significance of the topic

Hydroxysafflor yellow A is a major bioactive flavonoid in Carthamus tinctorius (safflower), widely used in traditional Chinese medicine for its cardiovascular and anti‐inflammatory properties. Reliable quantification of this marker compound is essential for quality control, standardization of herbal preparations and ensuring consistency across commercial safflower products.

Study objectives and overview

This work aimed to develop and validate a simple, rapid and robust reversed-phase liquid chromatography method for the determination of hydroxysafflor yellow A in commercially available safflower powder. Using a Shim-pack Scepter C18-120 column and an isocratic mobile phase, the study demonstrates effective separation of the target analyte from matrix components, enabling accurate quantification.

Methodology and instrumentation

This section outlines sample preparation, chromatographic conditions and analytical equipment.

• Sample pretreatment: 0.4 g safflower powder extracted with 50 mL of 25 % methanol via ultrasonic treatment (300 W, 50 kHz, 40 min). After cooling, volume was adjusted to initial weight and filtered.
• Chromatographic system: Shimadzu LC-20AD pump with Shim-pack Scepter C18-120 column (5 µm, 250 × 4.6 mm i.d.).
• Mobile phase (isocratic): methanol : acetonitrile : 0.7 % phosphoric acid = 26 : 2 : 72.
• Flow rate: 1.0 mL/min; column temperature: 20 °C; injection volume: 10 µL.
• Detection: UV at 403 nm.
• Vials: LabTotal™ Vial for LC/LC-MS.

Key results and discussion

Chromatograms of standard mix and safflower extract showed a well‐resolved peak for hydroxysafflor yellow A with a retention time around 17 min. The method exhibited:

• High selectivity: no co-eluting interferences observed at 403 nm.
• Good linearity: calibration at 0.13 mg/mL standard concentration.
• Reproducibility: consistent peak area across replicate injections.

Robustness was demonstrated by stable retention time and peak shape under routine conditions.

Benefits and practical applications

This HPLC approach provides a straightforward quality control tool for herbal manufacturers and research laboratories. Key advantages include minimal sample handling, isocratic operation for ease of use and UV detection for cost-effective quantification of the active marker.

Future trends and possibilities

Advancements may include:

• Integration with mass spectrometry for enhanced sensitivity and structural confirmation.
• Transition to ultra-high performance LC to reduce analysis time and solvent consumption.
• Application to multi‐component fingerprinting of safflower extracts.
• Development of green extraction and mobile phase protocols.

Conclusion

The presented reversed-phase HPLC method is validated for accurate determination of hydroxysafflor yellow A in safflower products. Its reliability and simplicity make it suitable for routine quality control in pharmaceutical and nutraceutical industries.

Reference

Shimadzu Corporation. Application Note ERAS-1000-0017. First Edition: March 2023.