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Developing and Validating a Method for Separating Flavonoid Isomers in Common Buckwheat Sprouts Using HPLC-PDA.

Davin Jang1, Young Sung Jung2, Mi-Seon Kim3

  • 1Graduate School of Biotechnology, Kyung Hee University, Yongin 17104, Korea. davin1031@khu.ac.kr.

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Summary

This study optimized a high-performance liquid chromatography (HPLC) method for separating and quantifying flavonoid isomers in buckwheat sprout extract. The validated method accurately and precisely analyzes key flavonoids, enabling their monitoring in sprouts.

Keywords:
chromatographic separationcommon buckwheat sproutflavonoid isomerquercetin-3-O-robinobiosidevalidation

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Area of Science:

  • Analytical Chemistry
  • Plant Biochemistry
  • Nutraceutical Science

Background:

  • Buckwheat sprouts are a rich source of bioactive flavonoids, including orientin, vitexin, and rutin, along with their isomers.
  • Accurate quantification of these flavonoid isomers is crucial for understanding their health benefits and for quality control.
  • Existing analytical methods may not offer sufficient resolution for separating closely related flavonoid isomers.

Purpose of the Study:

  • To develop and validate a high-performance liquid chromatography (HPLC) method for the optimal separation and simultaneous quantification of flavonoid isomers in common buckwheat sprout extract (CSE).
  • To establish robust analytical parameters including linearity, precision, accuracy, limit of detection, and limit of quantification for key flavonoids.

Main Methods:

  • Optimization of HPLC conditions, including column temperature (40 °C), mobile phase composition (0.1% acidic water and acetonitrile), and flow rate (1 mL/min).
  • Validation of the method using standards of orientin, isoorientin, vitexin, isovitexin, and rutin, assessing linearity, accuracy (recovery 96.67-103.60%), and precision (RSD < 5.40%).
  • Application of the validated HPLC method to analyze and quantify flavonoid isomers present in common buckwheat sprout extract.

Main Results:

  • Optimal HPLC conditions were determined, achieving good resolution and symmetry for separating flavonoid isomers.
  • The method demonstrated excellent linearity (R² = 0.9999) over the tested concentration range (6.25-100.00 μg/mL) for all target flavonoids.
  • High accuracy and precision were confirmed, with recovery rates between 96.67-103.60% and relative standard deviations below 5.40%.

Conclusions:

  • The developed and validated HPLC method provides a reliable and efficient means for the simultaneous separation and quantification of flavonoid isomers in buckwheat sprouts.
  • This analytical technique is suitable for routine monitoring of flavonoid content in buckwheat sprouts, supporting research and quality assurance in the food and nutraceutical industries.