Objective To investigate the
linear relationship between the
quantity of unknown minor impurity and its HPLC
peak area in rifaputin.Method A Hypersil-C18(4.6mm×150mm,5μm);the mobile phase was a mixture of acetonitrile and 0.1 mol/L monobasic potassium phosphate(50∶50),adjusted by 2mol/L sodium hydroxide to a pH of 6.5±0.1;detected at a wavelength 254nm.A series of rifaputin solutions containing 0.09～2.2mg/ml were prepared and 10μl of the solutions was injected into the chromatograph respectively.The peak area of
impurities were recorded.Let x0 equal to the quantity of impurities in 0.09mg/ml solution,then other solutions contain 2x0～24.44x0 of impurities.Calculate the relationship coefficients between the quantity of impurities which were substituted by the
coefficient of x0 and the peak area.Results The linear range was 0.36 to 8.8μg/ml;the relationship coefficent was 0.9997.Conclusion The relationship between the quantity of impurities and the peak area was linear;besides,mathematics derivative revealed that relative coefficient was related to x0 coefficient,but not to x0 itself.Therefore,the
Method of calculating relationship coefficient was scientific,reliable,simple and direct.This method provided a new way for other drugs to investigate linear relationship between unknown impurity and peak area of HPLC.
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