Lc 10atvp pump

NRG was assayed by HPLC using a system consisting of a quaternary LC-10AT VP pump equipped with UV/VIS detector, SPD-10AVP column oven (Shimadzu), a Rheodyne injector, and chromatography data system software (CLASS-VP Ver 6.14 SP1). Chromatographic separation was carried out on column (LiChrospher®100 RP-18 (5 μm), Merck, Darmstadt, Germany) with a mobile phase consisting of a 7:3 ratio of methanol and water with 0.1 % glacial acetic acid at ambient temperature (25 ± 0.5°C) having a flow rate of 1 ml/min. Samples (20 μl) were filtered through microfilters having 0.45 μm pore size and Rheodyne injector was used for injecting the samples which were then examined at 289 nm [22 (link)].

Each of the five lipids tested (i.e. Ovucire®, Precirol®, Compritol 888 ATO®, Geleol®, and Suppocire) was added to duplicate glass vials mixed with excess amount of PV, and the vials were vigorously vortexed before being incubated at 40 °C for the duration of the solubility research. To keep the medicine suspended, vials were regularly vortexed and carefully sealed. Samples were taken at 24, 48, and 72 h to attain equilibrium. The vials were whirled in a temperature-controlled centrifuge for 30 min at 5000 rpm, before sampling (Eppendorf centrifuge 5804R). The supernatant was put into a tared 5-mL volumetric flask, and diluted with 66% vol/vol chloroform and methanol mixture (Persson et al., 2013 (link)).
On a Shimadzu LC-10ATVP pump, SPD-M10 AVP with PDA detector, HPLC analysis of PV in collected samples was carried out. At room temperature, separation was accomplished using the phenomenex C18 (250 × 4.60); 5 particle size column and a mobile phase made up of 0.5% acetic acid and acetonitrile (35:65) at a flow rate of 1 mL/min. Prior to use, the mobile phase was filtered through a 0.45-m nylon filter (Millipore, USA) and sonicated to remove any gas. At 266 nm, the detection was carried out. A 20-loop sample injector valve was used to introduce the standard and sample. CLASS-VP software was used to measure peak areas during the data collecting process (Kumar et al., 2019 ).

Qualitative and quantitative detection of GLSs in samples after IEC, GPC, and TLC was performed using an isocratic HPLC system. This system consisted of an LC 10ATvp pump and UV–VIS detector from Shimadzu Corporation (Kyoto, Japan) and a SpectraSYSTEM AS3000 autosampler from Thermo Separation Products (Piscataway, NJ, USA), as well as a HILIC column containing sulfoalkylbetaine zwitterionic functional groups (150 × 4.6 mm, 5 μm, EC 150/4.6 Nucleodur HILIC 5 μm, Macherey-Nagel, Düren, Germany). HPLC measurements were performed according to the procedure of Wade et al. [103 (link)]. After injection of 20 μL of sample, GLSs were isocratically eluted with 15 mM ammonium formate in 70% (v/v) acetonitrile at pH 5 and at a flow rate of 0.5 mL/min and detected at 229 nm in a UV detector. Chromatograms were evaluated with Clarity Lite™ 2.1 software (DataApex, Prague, Czech Republic). The purity of the final preparations was controlled by an additional run on HILIC and was calculated as the ratio of the peak area of individual GLSs to the total area of the peaks detected in the chromatogram at 229 nm. The amount of individual GLSs was calculated from the peak areas at 229 nm relative to the peak of the internal standard (potassium sorbate) using the relative response factor (evaluated in our laboratory).

The monosaccharide composition analysis was determined by HPLC performed on a Shim-pak VP-ODS column (150 × 4.6 mm i.d) with a guard column of a Shimadzu HPLC system (LC-10 ATvp pump and UV-Vis detector) and monitored by UV absorbance at 245 nm. The CVG sample (2mg), initially was methanolyzed with 2M HCl at 80°C for 16 h, and then hydrolyzed with 2 M TFA (1 mL) at 120°C for 1 h. Elution was carried out at a flow rate of 1.0 mL/min at room temperature. The hydrolysate was derivatized to be 1-phenyl-3methyl-5-pyrazolone (PMP) derivatives and subsequently analyzed by HPLC [50 (link)]. D-Gal, D-Ara, D-Fuc, D-Rha, D-Man, D-Xyl, D-Glc, D-GlcA and D-GalA were used as sugar standards.

The concentration of ZL006 in samples was measured via HPLC conducted by using a LC-10ATVP pump and SPD-10AVP UV detector (Shimadzu, Kyoto, Japan). The mobile phase was consisted of methanol and ammonium acetate buffer solution (0.25 mol/L, pH 6.0) (35:65, v/v) and the flow rate was set at 1.0 mL/min and the detection wavelength was 284 nm. The HPLC was calibrated with standard solutions of 1–100 μg/mL of ZL006 dissolved in methanol (Y = 48030X-17645, correlation coefficient of R² = 0.9997). The limit of quantification was 0.5 ng/mL, and the coefficients of variation were all within 3.5%.