Twenty ll were injected into a C18 column (150 3.9 mm i.d., 5 lm size, Waters) and the mobile phase consisting of solvent A (acetonitrile), solvent B (methanol) and solvent C (bi-distilled water) was eluted at 1.0 ml/min for 50 min as follows: isocratic flow 60% A, 35% B and 5% C for 5 min, gradient flow 60% A and 40% B to 8 min, maintained at 60% A and 40% B for 10 min, and then gradient flow 22% A and 78% B to 20 min, maintained isocratically to 35 min, reverted to initial conditions to 45 min, and then isocratic conditions that were kept to equilibrate the column to 50 min.