D2000

The content of vitamin C in Chinese chive was estimated by using the previous methods [14 (link),47 (link)]. The tissues of Chinese chive were derivatized with 1,2-o-phenylenediamine after extracting with trichloroacetic acid solution. The vitamin C content was determined by HPLC (D-2000, Hitachi, Ltd., Tokyo, Japan; excitation at 350 nm, emission at 430 nm, respectively). Quantification of vitamin C content was carried out by external calibration with L-ascorbic acid and expressed as mg 100 g⁻¹ FW.

Ten micrograms of anti-CR-1 Ab was analyzed by anion exchange column Inertsil AX column (ø4.6 mm × 150 mm, GL Sciences, Tokyo, Japan). The elution was performed using a linear gradient of NaCl concentration from 0 to 1 M in 10 mM Tris-HCl, pH 8.0, for 30 min at a flow rate of 0.5 mL/min. The absorbance was monitored at 280 nm. The chromatography was performed with the HPLC system D2000 (Hitachi, Tokyo, Japan).

The analysis of AOFE was performed using an HPLC system (Hitachi D2000, Tokyo, Japan), which consisted of an L-2130 HTA pump, an L-2200 autosampler, and an L-2455 photodiode array detector. All solvents used were HPLC grade, and all reagents were analytical grade. The extracted compounds of AOFE and the kojic acid standard (Sigma-Aldrich, St. Louis, MO, USA) were separated using an Inertsil ODS-2 C18 column (250 mm × 4.6 mm, 5 μm) with a gradient mobile phase of 0.1% formic acid in ddH₂O (solvent A) and 0.1% formic acid in acetonitrile (solvent B). The analysis involved a gradient elution at a flow rate of 1 mL/min: 0–10 min, 10% B; 10–24 min, 10% B to 20% B; 24–30 min, 20% B to 22% B; 30–35 min, 22% B to 25% B; 35–35.1 min, 25% B to 10% B; 35.1–45 min, 10% B. Injection volume was 10 μL. The column temperature was maintained at 30 °C, and the absorbance of the eluate was monitored continuously at 280 nm. Chromatographic peaks were identified by spiking the samples with standard compounds, and their UV spectra and retention times were matched with standard compounds.