10atvp auto sampler

The liquid chromatographic system consisted of two LC-20AT pumps, a Shimadzu 10ATvp auto sampler, a CTO-10Avp column oven, a SPD-20A detector and a CBM-10Avp Plus chromatographic station (Shimadzu, Kyoto, Japan). Analyses were performed on a Shim-pack VP-DOS C18 analytical column (250 mm×4.6 mm i.d., particle size 5 μm, Shimadzu, Kyoto, Japan), which was protected by a ODS guard column (Security Guard, Phenomenex^®, USA) at 30 °C. The mobile phase was composed of methanol/water (45:55, v/v) at a flow rate of 0.50 mL/min. The detection wavelength was set at 218 nm. Under these conditions, triptolide eluted at approximately 15 min and the internal standard at 25 min (Fig. 3).

A HPLC system consisted of a LC-10AD pump, a DGU-14 AM degasser, a Shimadzu 10ATvp auto-sampler and a CTO-10Avp column oven (Shimadzu Corporation, Kyoto, Japan) was employed to achieve simultaneous detection of all analytes. Separation was performed on a Shimadzu 2010 liquid chromatograph–mass spectrometer (Shimadzu Corporation) with a LUNA C18 column (150 mm × 2 mm, 5 μm, Phenomenex®, Los Angeles, CA, USA). The mobile phase was consisted of 0.1 mmol/L ammonium chloride solution (A) and acetonitrile (B) and the flow rate was set at 0.2 mL/min. A gradient elution program was used as follows: 0.04 → 1.5 min, B% 25 → 25; 1.5 → 12.0 min, B% 25 → 45; 12.0 → 19.0 min, B% 45 → 90; 19.0 → 22.0 min, B% 90 → 90; 22.0 → 23.0 min, B% 90 → 25; 23.0 → 29.0 min, B% 25 → 25. Quantification was performed using SIM mode with [M+Cl]⁻ peak which was modified from our previous method²¹ (link): m/z 1143.3 for Rb1; m/z 1113.3 for Rb2; m/z 1113.3 for Rc; m/z 981.4 for Rd; m/z 835.4 for Rg1; m/z 981.4 for Re; m/z 819.3 for Rg3; m/z 955.3 for Ro; m/z 815.4 for digoxin. The concentration range in plasma was 5–2000 ng/mL for Rb1 and Rb2, 5–1000 ng/mL for Rc, Rd, Rg1, Re and Ro, and 5–500 ng/mL for Rg3. The calibration curve of all ginsenosides detected in tumor showed good linearity over the concentration range of 5–500 ng/mL.