Optimization of HPLC Conditions for Analysis of Hirudin
In order to study the optimal chromatographic conditions for the HPLC analysis of hirudin,we compared the elution conditions,detection wavelengths and separation effects of the HPLC methods for recombinant hirudin,fitted the linear equations of hirudin concentration and peak area,and optimized the hirudin chromatographic analysis conditions.The results showed that the optimal conditions for the analysis on the ODSC18 column were as follows:gradient elution,mobile phases A and B elution program:0～60 min,B:0％～100％,injection volume of 20 μL,column temperature of 35℃,flow rate of 1 mL/min,and wavelength of 205 nm.And the optimal conditions for the analysis on the TSKgelG2000SW column were as follows:mobile phase's phosphate buffer(0.02 mol/L,pH 7.0),flow rate of 0.5 mL/min,and wavelength of 205 nm,injection volume of 20 μL.The study showed that the concentration of hirudin had a good linear correlation with the peak area,and the analytical effect of the TSKgelG2000SW column was better than that of the ODSC18 column,which could be used for the analysis of the active peptides in animals.
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维普
