Agilent 1200 rr hplc instrument

Analyses were carried out on an Agilent 1200 RR HPLC instrument with a DAD detector (Agilent, Waldbronn, Germany) using a reversed-phase LiChrospher RP-18 (Agilent) analytical column (250 × 4 mm i.d.; 5 µm particle size). The mobile phase consisted of solvent A (1% v/v solution of orthophosphoric acid in water) and solvent B (acetonitrile). Separation was achieved according to the following scheme: 90–80% A 0–5 min, 80% A 5–10 min, 80–70% A 10–20 min, 70–30% A 20–30 min, 30–0% A 30–35 min. Detection wavelengths were set at 280 and 330 nm, and the flow rate was 1 mL/min. The injection volume was 5 μL, while the column temperature was maintained at 25 °C. The compounds 4′-O-methylhypolaetin-7-O-[6‴-O-acetyl-β-d-allopyranosyl (1→2)-β-d-glucopyranoside (HYP), isoscutellarein 7-O-[6‴-O-acetyl-ß-d-allopyranosyl (1→2)]-ß-d-glucopyranoside (ISC 1) and 4′-O-methylisoscutellarein-7-O-[6‴-O-acetyl-β-d-allopyranosyl-(1→2)]-β-d-glucopyranoside (ISC 2) were previously isolated in our laboratory [2 ]. The concentrations of compounds in the investigated extracts were determined using calibration curves (r > 0.9997). The results are presented as milligrams per gram of dry weight (mg/g dw).

Analyses were performed on Agilent 1200 RR HPLC instrument (Agilent, Waldbronn, Germany), on a reverse-phase Zorbax SB-C18 (Agilent, Waldbronn, Germany) analytical column (150 mm × 4.6 mm i.d.; 5 μm particle size) according to the previously described procedure [17 (link)]. Identification of the marker compound (gentiopicroside) was achieved by comparing their UV spectra and retention time with those from authentic standards, while the concentration was determined from the peak areas by using the equation for linear regression obtained from calibration curves (correlation coefficient was 0.998).