Lc 30ad pump

The Reverse phase HPLC (RP-HPLC) system (Waters, United States of America) equipped with a rheodyne 7725i injector having 20 µL loop, 3 Lines degasser (Volume 400 μL), LC-30AD pump, C18 column (5 µm particle size, 250 × 4.6″) UV/Vis detector was used for the analysis. The method developed was isocratic with the mobile phase of Methanol: Water (1% acetic acid) - 60:40 v/v/v. A Milli-Q academic water purification system (Bedford, MA, United States of America) equipped with 0.22 μm Millipak express filter. The pH of the solvent B was adjusted at 2.4 by using 1% (v/v) glacial acetic acid for better ionization. The mobile phase was filtered through 0.45 mm pore size (Millipore) membrane filter followed by sonication to degas the solvent. Whatman syringe filters (NYL 0.45 μm) were used for the filtration of the sample. The temperature of the column was kept at 25°C and the injection volume was 20 µL. The total run time was set at 10 min. The flow rate was set at 1.0 ml/min and the λ_max was set at 230 nm for maximum absorption of the compound. Quantitative estimation was performed with Empower 2 software programs using the external calibration method. Typical chromatograms of andrographolides standard and an Andrographis paniculata extract sample are shown in Figure 1.