HPLC-UV Quantification of Phthalides

The quantification of five phthalides of VOC in formulation samples was determined by a validated reversed-phase HPLC-UV method (Shimdzu, Japan). The HPLC system consisted of a LC-20AT pump, an SPD-M20A PAD-visible detector, a SIL-20A injector with a 20 μL loop and an LC solution workstation. The mobile phase consisted of a mixture of ultrapure water (A) and acetonitrile (B) (0~10 min, 38~42% B; 10~36 min, 42~45% B; 36~55 min, 45~48% B; 55~60 min, 48~38% B) was delivered with a flow rate of 1 mL/min, and separation was performed using an Akzo Nobel Kromasil 100-5-C18 column (4.6 mm × 250 mm, 5 µm) with a sample injection volume of 20 µL. The analysis was performed at 30 °C with UV detection at 280 nm for senkyunolide A(SA) and Z-ligustilide (ZL), and 230 nm for neocnnolide (NOL), n-butylphthalide (NBP) and butenylphthalide (BP) [22 (link)].