| Items | Conditions | ||
|---|---|---|---|
| HPLC system | 1260 (Infinity Quaternary LC System, Agilent Technologies, Waldbronn, Germany) | ||
| Column | ODS-H80 (ODS-H80, 4.6×250 mm, YMC Co., Ltd., Kyoto, Japan) | ||
| Mobile phase | Time (min) | 1% formic acid in H2O | 1% formic acid in ACN:MeOH (75:25) |
| 0 | 75 | 25 | |
| 1 | 60 | 60 | |
| 14 | 60 | 40 | |
| 22 | 40 | 60 | |
| 23 | 11 | 89 | |
| 45 | 11 | 89 | |
| Flow rate | 0.5 mL/min | ||
| Column temp. | 30°C | ||
| Injection volume | 5 μL | ||
| Detector | UV (280 nm) | ||