I had already tried DMSO 100%, H20 and methanol mixtures(50/50 v/v) to prepare the stock of drug, but still no multiple relationships between 10ug and 100ug these two concentrations on HPLC chromatography . Maybe is the solubility of this drug? But it appears to be completely dissolved.
spectrometer: shimadzu LC-MS/MS 8030 system
Column: purospher star RP-18 (2.1x100mm,2um, merck)
mobile phase: methanol /0.2% acetic acid solution (40/60 v/v)
Hi Ruby Huang - The issue with linearity between 10 µg and 100 µg concentrations may stem from solubility, detector saturation, or column overload. Ensure the drug is fully dissolved by sonication and filtration, and confirm the injection volume isn’t too high to avoid detector saturation. Test intermediate concentrations to verify the linear range of your system and optimize the mobile phase ratio (e.g., increase methanol to 50:50). Clean the column with a gradient wash and check MS parameters for consistency. Finally, prepare fresh stock solutions, mix thoroughly, and run a serial dilution to establish a reliable calibration curve. Thanks, Ahmed!
To improve the relationship between 10 µg and 100 µg concentrations in HPLC, verify the drug's solubility limits, optimize method parameters, and check for potential interference or matrix effects. Experimenting with different solvent systems and reviewing relevant literature may also help enhance your results.
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