G7129a vial sampler

For quantitative analysis, an Agilent 1260 system equipped with a G7112C Quat Pump, a G7129A Vial sampler, and a G7117C diode array detector (DAD) was used (Agilent, Santa Clara, CA, USA). The chromatographic parameters were the same as described in Section 3.4. The chromatograms were recorded at 254 nm.

The analysis was performed on an Agilent 1260 Infinity II LC system (Agilent Technologies UK Ltd., Stockport, UK) equipped with 1260 Quat Pump G7111B, a G7129A Vialsampler and a G7121A fluorometric detector (excitation wavelength 280 nm, emission wavelength 350 nm). The fluorescence detector gain was set at 10 with an attenuation setting of 100. The ODS-3 analytical column (250 mm × 4.6 mm internal diameter, 5 μm packing; Inertsil^™, GL Sciences, JP; InertClone^™, Phenomenex, Torrance, USA) protected by a precolumn (SecurityGuard^™ Guard Cartridge Kit, Phenomenex, Torrance, USA), was used to separate and quantify enfuvirtide at 25 °C. The mobile phase was composed of solvent A (deionised water with 0.1% v/v phosphoric acid, pH 2.5) and solvent B (ACN). A gradient with a flow rate of 1 mL/min was applied in the following way: 35–45% B (0–3 min), 45–75% B (3–5.5 min), 75% B (5.5–7 min), 75–35% B (7–9 min), as shown in Table 1. The injection volume was 30 µL, followed by six needle washes.Table 1

Chromatographic condition (gradient)

Time (min)	% Solvent A	% Solvent B	Flow (mL/min)
0.0	65.0	35.0	1.0
3.5	55.0	45.0	1.0
5.5	75.0	75.0	1.0
7.0	35.0	35.0	1.0

To identify allantoin in yam extract, HPLC was conducted using an Agilent 1260 infinity II quaternary system equipped with a G7129A vial sampler and a WR G7115 Adiode array detector (Agilent, Waldbronn, Germany) and a ZORBAXNH2 column (4.6 × 150 mm, 5-micron). Chromatographic separation was performed using a gradient solvent system consisting of acetonitrile (HPLC grade, Merck, Darmstadt, Germany) (B) and water (HPLC grade, Merck) (A). The gradient program was as follows: 0 min, 25% B; 5 min, 17% B; 10 min, 17% B. The injection volume was 10 μL and the column eluent was monitored at UV 200 nm while chromatography was performed at 30 °C with a flowrate of 1.0 mL/min. The HPLC pattern of allantoin in yam water extract has been reported in the literature [9 (link),26 (link)].