Answer Happy

a # Morphine and morphine- 3-B-D-glucuronide were separated using two different 50 x 4.6 mm columns with 3 um particle. Column A was C18-silica run at 1.4 mL/min and column B was pure silica run at 2.0 mL/min. Separation using column A was achieved using a mixture of 10 mM ammonium acetate (pH 3.0) and acetonitrile (98:2, v/v). Morphine and morphine-3-3-D-glucuronide eluted at 1.5 and 2.8 min, respectively. Calculate the retention volume and explain the order of elution. [6 marks] b. Find the retention factor k for each analyte on column A given that t = 0.65 min. [4 marks] Analytes were separated with column B using a 5 min gradient starting at 90 % acetonitrile in water and ending at 50 % acetonitrile in water. Both solvents contained 10 mM ammonium formate. Morphine and morphine-3-3-D-glucuronide eluted at 1.3 and 2.7 min. Explain the order of elution. Why does the gradient go from high to low acetonitrile? [6 marks] Calculate k for each analyte on column B given that to is 0.50 min. [4 marks]