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The gradient was developed by mixing increasing proportions of
absolute methanol to 10% methanol plus 0.5% acetic acid in water
The gradient of absolute
methanol was programmed to 20% over the first
15 min, from 20% to 45% for between 15 min
and 22 min, from 45% to 54% for between
22 min and 33 min, from 54% to 100% for between
33 min and 34 min, and 100% for between
34 min and 50 min, at 1 ml min¨C1 flow at 40C.
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