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The HPLC was set at a flow rate of 0.25 ml min 1 with 99% buffer A from 0 to 3 min, a linear gradient to 99% buffer A/1% buffer B (100% methanol) from 3 to 20 min, 80% buffer A/20% buffer B from 20 to 21 min, a linear gradient to 30% buffer A/70% buffer B from 21 to 28 min at 0.35 ml min 1 , 99% buffer A/1% buffer B from 28 to 31 min and a linear gradient to 99% buffer A from 31 to 37 min at 0.25 ml min 1