| Step 1: |
70-min gradient from 0% to 80% solvent B |
|
10 min held at 80% solvent B |
| Steps 2-13: |
Each of these steps follows the sequence: |
|
5 min of 100% solvent A |
|
2 min of x% solvent Ca |
|
3 min of 100% solvent A |
|
10-min gradient from 0% to 10% solvent B |
|
90-min gradient from 10% to 45% solvent B |
| Step 14: |
5-min 100% solvent A wash |
|
20-min 100% solvent C wash |
|
5-min 100% solvent A wash |
|
10-min gradient from 0% to 10% solvent B |
|
90-min gradient from 10% to 45% solvent B |
| Step 15: |
5-min 100% solvent A wash |
|
20-min 100% solvent D wash |
|
5-min 100% solvent A wash |
|
10-min gradient from 0% to 10% solvent B |
|
90-min gradient from 10% to 45% solvent B |
| aThe 2-minute solvent C percentages (x) in Steps 2-13 are as follows: 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 90%, 100%,
and 100%, respectively.
|