This document introduces a large-scale separation method using Agilent Prep-C18 columns, which can be used to separate large amounts of anthocyanins and re-analyze the collected fractions to demonstrate purity.
This document describes the purification and identification of anthocyanins using Agilent Prep-C18 columns. It highlights the excellent scalability of these columns, allowing for method development on analytical scale columns while achieving high resolution and purity on preparative scales. The text explains that anthocyanins are highly retained on C18 columns and require high concentrations of organic solvents for extraction.
The application demonstrates the scalability of the Agilent Prep-C18 column using blueberry extract. By using the ChemStation fraction collection software with specific threshold and slope settings, fractions were obtained that were significantly purer than the separation would seem to allow. The document includes chromatograms showing the separation of Delphinidin-3-galactoside and Delphinidin-3-glucoside.
| Column Dimensions (Prep) | 21.2 x 250 mm, 10 µm |
|---|---|
| Flow Rate (Prep) | 21.2 mL/min |
| Flow Rate (Analytical) | 1.0 mL/min |
| Injection Volume (Prep) | 2000 µL |
| Injection Volume (Analytical) | 100 µL |
Fraction 1 is greater than 99% pure Delphinidin-3-galactoside.
Fraction 2 is greater than 97% pure Delphinidin-3-glucoside.
The ChemStation fraction collection software is used to obtain pure fractions.