Agilent 1200 liquid chromatography system

The extraction and determination methods of MC-LR were determined by HPLC, according to Zhao et al. [30 ]. Sample pretreatment ((solid-phase extraction (SPE)): (1) Filtration: Absorb 12 mL algal liquid and use the water filter of 0.45 μm membrane to collect the filtrate. (2) Activation: SPE column was activated with 6 mL of methanol and 6 mL of high purity water. (3) Enrichment: 10.0 mL of filtrate was added for enrichment after activation of the SPE column. (4) Elution: After enrichment, impurities were removed with 6 mL of high purity water and 6 mL of 10% methanol, and then MC-LR was washed twice with 80% methanol (3 mL each time) to elute MC-LR. The eluent was concentrated to less than 1 mL with a rotary evaporator, and the volume was fixed to 1 mL with high purity water. The samples were obtained through the above operations.
MC-LR was determined with the Agilent 1200 liquid chromatography system (Agilent Technologies Inc., Palo Alto, CA, USA) Chromatographic conditions: Chromatographic column: C18 reversed-phase column with a column length of 250 mm, inner diameter of 4.6 mm and filler particle size of 5 μm; chromatographic column temperature: 40 °C; mobile phase: high purity water (containing 0.05% trifluoroacetic acid): methanol = 65:35; flow rate: 1.0 mL/min; detector: UV detector, wavelength 238 nm.

BSYZ-E was qualitatively analyzed by HPLC. Agilent 1200 liquid chromatography system (Santa Clara, USA), equipped with ELSD, a quaternary solvent delivery system, a column temperature controller, and an autosampler, was used for chromatographic analysis. BSYZ-E, BSYZ-F and the mixed standard solution were separated on Phenomenex Luna-C18 column (250 mm × 4.6 mm, 5 μm) at 40 °C. The mobile phase was composed of 0.1% formic acid in water (A) and 0.1% formic acid in methanol (B). Programmed gradient elution was performed as follow: 0–30 min, 10–30% B; 30–40 min, 30–50% B; 40–70 min, 50–100% B. The flow rate was 1 mL/min. Monitoring was performed at 254 nm with PDA detector. Chromatographic data were recorded and processed with Allchrom Plus Client/Server software.

Following partial reduction, SE-HPLC was used with the in-line UV signal detector and was set at 280 nm to confirm the integrity of the full-size antibody and absence/presence of protein aggregates, using an Agilent 1200 Liquid chromatography system (Agilent Technologies, Santa Clara, CA, USA). The column used was TSKgel G3000SWXL 7.8 mm ID × 30 cm (TOSOH Bioscience, Tokyo, Japan). The mobile phase used was 0.15 M potassium phosphate buffer pH 6.5, flow rate was 0.5 mL/min, 10 µL was used as an injection volume, and measurements were performed in triplicate and repeated twice with two different vials.

The identification and quantification of phenolic compounds in cladode extracts was performed in triplicate (an analysis for each of the three extractions/replicas) using an Agilent 1200 Liquid Chromatography system (Agilent Technologies, Palo Alto, CA, USA) and the chromatographic conditions and column were the same already reported by Sabella et al. [13 (link)]. The identification of phenolic compounds was confirmed by a TOF LC/MS system (Agilent 6320, Agilent Technologies, Palo Alto, CA, USA), equipped with a dual ESI interface operating in negative ion mode [13 (link)].
The identified phenolic compounds were quantified by the external standard method using a six-point calibration curve of p-hydroxybenzoic acid (0.5–100 mg/L), rutin (0.5–50 mg/L), isorhamnetin (0.5–50 mg/L), and narcissin (0.5–50 mg/L).