Table 1.

HPLC gradient solvent profile^a

Time (min)	% Solvent A	% Solvent B	% Solvent C	Flow (ml/min)	Curve
0	100	0	0	0.4	1
5	60	40	0	0.4	6
7	10	90	0	0.4	6
9	0	0	100	0.4	1
11	0	0	100	0.4	1
12	100	0	0	0.4	1
20	100	0	0	0.4	1

^a Waters Alliance 2795 HPLC. Solvent A, 95% acetonitrile–5% water with 0.1% formic acid and 0.005% trifluoroacetic acid. Solvent B, 20% methanol–80% water with 0.1% formic acid. Solvent C, water, with 0.1% formic acid. Curve 6 is a linear gradient; curve 1 is a step change to the indicated percent solvent.