Question

Determination of Benzoic Acid and Caffeine in Soft Drinks via Liquid Chromatography.

Q1: This experiment is reverse phase chromatography. What does that tell you about the polarity of the mobile and stationary phases? (2 pts)

Q2: Which compound do you expect to elude first from the column and why? (2 pts)

Q3: What is the definition of an isocratic separation? (2 pts)

Q4: What is an acceptable value for the resolution of a column? (2 pts)

Q5: Do you expect this method or be more sensitive or less sensitive than UV-visible spectroscopy? (2 pts)

Answer 1

Q1. Reverse phase chromatography is a type of chromatography which uses hydrophobic i.e. non polar stationary phase and hydrophilic or polar mobile phase. This type of method of separation of solution depends on the reversible adsorption and hence is called reverse phase.

Q2. Since polar components of mixture does not reacts with non polar components, they tends to elute first. In our case caffeine is more polar than benzoic acid. So caffeine must elute first from the column.

Q3. Isocratic separation is a method of separating a mixture by keeping the constant composition of mobile phase.

Q4. Resolution value of the column gives us the quantitative measure of how well elution peaks can be observed in a separation. In general, resolution value of the column must be greater than or equal to 1 to identify the peaks properly.

Q5. UV-visible spectroscopy is more sensitive but it has many issues like it can only determine the components, will absorb light for impurities also etc. On the other hand, reverse phase chromatography is much better in terms of determination and separation of components.