Eclipse plus c18 rapid resolution hd

Urinary 11-dehydrothromboxane B2 was measured by LC–MS/MS method. The extraction of 11-dehydroythromboxane B2 from urine samples was performed following solid-phase extraction (13 (link)). The separation of 11-dehydroythromboxane B2 and IS from endogenous substances was achieved using ZORBAX Eclipse Plus C18 Rapid Resolution HD (2.1 × 50 mm, 1.8 μm) and mobile phase consisting of a mixture of 2 mM ammonium formate with 0.1% formic acid (A) and methanol with 0.1% formic acid (B) in a gradient program mode. The details of the method, method validation data (Supplementary Table S2), and LC–MS/MS chromatograms for 11-dehydroythromboxane B2 (Supplementary Figure S7) are provided in the supplementary file.

Identification of aglycones after enzymatic hydrolysis by snailase was carried out by a 1290 Infinity II LC System equipped with a ZORBAX Eclipse Plus C18 Rapid Resolution HD (1.8 μm, 2.1 × 150 mm) connected to a G7117C diode array detector and subsequently followed by a 6545 LC/Q-TOF (Agilent, USA) with a multimode ion source. The flow rate was set to 0.3 ml/min, and the column oven was operated at 35°C.
The same mobile phase as for HPLC analysis was used with the following gradients, namely, 1 min at 5 vol% (B), 4 min from 5 to 15 vol% (B), 6 min to 53 vol% (B), 4 min to 100 vol% (B), 6.5 min at 100 vol% (B), 0.5 min to 5 vol%, and post run 5 min at 5 vol% (B). The wavelength during analysis was 290 nm. All solvents were of LC-MS grade.
Mass spectrums were recorded in ESI-negative mode between m/z 100 and 1,000 Da (scan rate: 2 spectra/s) after a 5 min solvent cut. The mass spectrometer was operated as follows: gas temp.: 350°C, gas flow: 10 l/min, nebulizer: 40 psig, vaporizer: 220°C, VCharge: 1.75 kV, VCap: 1 kV, fragmentor: 180 V, and skimmer: 75 V.
Flavonoid aglycones were identified according to the retention time, substance-specific UV absorbance, and the accurate mass of the molecular ion. All identified flavonoid aglycones and relevant data per plant extract by mass spectrometric analysis are presented in Supplementary Table S23.