4. Using a 20 cm long C18 bonded HPLC column filled with 5m size particles and a mobile phase consisting of 50% methanol in water, Vitamins B2 and B3 elute at 4.2 and 4.4 minutes respectively with corresponding base peak widths of 12.6 and 13.2 seconds respectively. The mobile phase solvent peak elutes at 2 minutes. Calculate:

a. the retention factors (k) for both vitamins

b. the selectivity ()

c. the number of theoretical plates (N) and the Height Equivalent to a Theoretical Plate (H)

d. the resolution Rs

e. Identify the parameter (N, k, ) that is responsible for this low resolution and the practical step you would take to improve the separation?