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Gradient elution is usually recommended in the complexity of the biological matrix for compounds separation, but it needs long time for column cleaning (using high organic solvents) and rebalance (returning column to initial condition). The run time is usually more than 7 min, which will occupy large instrument resource. Therefore, how to shorten the run time and increase the output rate of instruments without losing resolution during sample analysis in nonclinical PK study is important.
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