Meteoric core c18 bio

The extracted and purified mixtures of lipopeptides were dissolved in acetonitrile and then filtered using a 0.22 μm membrane filter. For analyses, the UHPLC Dionex UltiMate 3000 system with DAD detector at a wavelength of 214 nm was used. For separations, the reverse-phase column YMC Meteoric Core C18 Bio (150 × 4.6 mm) with a particle size of 2.7 μm was utilized. The injection volumes were 20 μL. All the standard lipopeptides (iturin A, fengycin, surfactin) were purchased from Sigma Aldrich, Germany. The concentration of standards was 0.5 mg/mL. The eluent A consisted of 0.025% TFA in acetonitrile and eluent B was 0.025% water solution of TFA. The separation strategy was optimalized using gradient elution and at the 0–5 min mark 100% of eluent B was used with the flow rate of 0.5 mL/min; the linear gradient followed from 5 to 10 min, using 0–10% of eluent A with the flow rate of 0.8 mL/min, and finally, from 10 to 75 min, 10–90% of eluent A with the flow rate of 1.5 mL/min was performed.

For analysis, the UHPLC Dionex UltiMate 3000 system with DAD detector at a wavelength of 346 nm (Thermo Scientific, Rockford, IL, USA) was used. For separations, the reverse-phase column YMC Meteoric Core C18 Bio (150 × 4.6 mm) with a particle size of 2.7 μm was utilized. The injection volumes were 5 μL. The specificity of the method was studied using standard samples of berberine (purchased from Sigma Aldrich, Germany, B3251). In the range of concentrations between 1.0 and 0.025 mg/mL. For the mobile phase, water (eluent A) and acetonitrile (eluent B) (A/0626/17, Fisher Scientific, Loughborough, Leics, United Kingdom) was used, using gradient elution. In the time of 0–2 min, 80% of eluent A was used with the flow rate range of 1 to 0.1 mL/min, then 50% of eluent A was used using the flow from 0.1 to 1 mL/min. The repeatability of sample measurements was carried out using three replicates of the same sample and was expressed as percentage of relative standard deviation (%RSD). The quantity of berberine was calculated by comparing their peak areas with those of standard solutions.