Zorbzax 300 sb c18 column

LC–Q–TOF–MS analysis was carried out using an Agilent Technologies (Santa Clara, CA, USA) 1290 Infinity Series liquid chromatograph coupled with an Agilent Technologies 6500 iFunnel Q–TOF LC/MS device equipped with an electrospray ionization Agilent Technologies Jet Stream ion source. Chromatographic separation was achieved on an Agilent ZORBZAX 300 SB-C18 column (4.6 mm × 250 mm) (Agilent, Santa Clara, CA, USA). Injection volume was 20 µL. The mobile phase consisted of 0.1% formic acid in Milli-Q water (solvent A) and 0.1% formic acid in 100% acetonitrile (solvent B) at a flow rate of 0.5 mL/min. The mobile-phase gradient (10–60% B) was applied. Source nitrogen gas temperature was 325 °C, sheath gas flow was 12 L/min, and nebulizer pressure was 40 psig. Voltages were set at 4000 (capillary) and 175 V (fragmentor). Positive ions were acquired in the range of 100–3200 m/z for MS scans. Internal mass correction was enabled by using two reference masses at 121.0509 and 922.0098 m/z. Data acquisition and instrument control were performed using Agilent MassHunter Workstation software (B.06.01 SPI).

DAKS1 purity was analyzed by HPLC on an Agilent ZORBZAX 300 SB-C18 column (4.6 mm × 250 mm) under linear gradient elution conditions by using acetonitrile as the organic modifier, and trifluoroacetic acid (TFA) as the volatile buffer. Eluent A consisted of 0.1% TFA in distilled water (v/v), and eluent B consisted of 0.1% TFA in 100% acetonitrile (v/v). Gradient elution was carried out according to the following process: 0–20 min, B 5–50%; 20–22 min, B 50–90%; and 22–25 min, B 90%. Flow rate was 1 mL/min. UV absorbance was monitored at 214 nm.