Yl 9100 series hplc instrument

Chromatographic analysis was performed using an YL-9100 series HPLC instrument equipped with a sample injector and a photodiode array ultraviolet/visible (PDA-UV/Vis) detector (Young Lin, Korea). The analysis was conducted using Phenomenex Gemini NX-C18 column (4.6 mm × 250 mm; 5 μm, Phenomenex Inc., CA, USA) as the stationary phase, with at injection volume of 20 μL at room temperature. The mobile phase was composed of water (containing 0.1% formic acid) (A) and acetonitrile (B), with the following gradient system: 0–5 min linear change from 10% B to 10% B, 5–10 min linear change from 10% B to 20% B, 10–60 min linear change from 20% B to 40% B, 60–80 min linear change from 40% B to 70% B, 80–85 min linear change from 70% B to 100% B, and 85–90 min held at 100% B. The flow rate for HPLC analysis was set to 0.7 mL/min while the detection wavelength was set to 210 and 254 nm. Each sample was analyzed three times. Standard samples for HPLC were dissolved in 50% methanol, and NJ20 was prepared at a concentration of 10 mg/mL. For the quantitative analysis of compounds 1–9, standard compounds were prepared at concentrations between 0.0125 and 0.5 mg/mL, and calibration curves were constructed by plotting peak areas against the known concentrations of the standard compounds. The slope, intercept, and correlation coefficient (R) were calculated by linear regression analysis.

To prepare a sample using the extract, 10 mg of water extract of ARP was dissolved in 1 ml distilled water and then filtered through a 0.45 μm syringe filter. Reference compounds such as (+)-catechin (99% purity, Cat. C1251) and 4-hydroxybenzoic acid (99% purity, Cat. H20059) were purchased from Sigma-Aldrich Chemical Co. The concentrations of the compounds were prepared as 0.125, 0.25, and 0.5 mg/ml in methanol for quantitative and qualitative analysis. High-performance liquid chromatography (HPLC) separation was performed using a YL-9100 series HPLC instrument equipped with a sample injector and UV/V detector (YoungLin. Republic of Korea). A YMC ODS-pack column (10 mm × 150 mm I. D., 5 μm, Japan) was used for separation. The ARP water extract sample (10 mg/ml) was injected at 20 μl. Subsequently, 10 μl of each compound was injected, and all sample injections were performed three times. For the HPLC analysis, 0.1% formic acid added to water (A)/acetonitrile(B) was used as the mobile phase with a gradient elution method: 0–10 min, an isocratic 5% B; 10–40 min, gradient from 5% B to 15% B; 40–55 min, gradient from 15% B to 100% B; 55–60 min held at 100% B. Flow rate was 0.7 ml/min, and detection wavelength was 254 and 280 nm.