AX = F AS

[X]
[S]

Chem 251 Exam 4 Fall 2005

NAME:

Questions on the exam are worth 10 points each unless otherwise noted. Show your setup for all
calculations.

Peak Identification
1.
2.
3.
4.
5.

Cytidine
Uridine
Xanthosine
Guanosine
Adenosine

1. Given the
HPLC
chromatogram
above for a
mixture of
nucleosides,
calculate the resolution between the Guanosine and Adenosine peaks using your best estimate of the
required parameters from the chromatogram.

2. What is a theoretical plate? What are some factors that determine N?

3. What is gradient elution and why would you need to use it? Give two examples of types of
gradients used in chromatography.

Peak Identification
1.
2.
3.
4.
5.

Barbital (0.25 mg/ml)

Phenobarbital (0.1 mg/ml)
Talbutal (0.3 mg/ml)
Amobarbital (0.25 mg/ml)
Mephobarbital (0.1 mg/ml)

4. Given the HPLC chromatogram above for a mixture of barbiturates, calculate the number of
theoretical plates based on the Amobarbital peak using your best estimate of the required parameters
from the chromatogram.

5. Referring to the chromatogram at the top of the page, assuming that barbital is more polar than
phenobarbital which is more polar than talbutal, etc, was this experiment run under normal or reverse
phase conditions? (2 point question)

6. Short Answer: (2 points for each answer)

a. A resolution of _________ is the minimum for baseline peak separation.
b. Which HETP is better, 0.5 mm or 1.0 mm?
c. The main purpose of chromatography is _______________________________________________
d. The most common HPLC detector is:_________________________
e. In liquid chromatography, the mobile / stationary phase is always liquid. (circle one).
7. (8 pt quest) Give the best match for the terms in the first list with the characteristics in the second:
____ normal phase chromatography

A. One common stationary phase is DEAE

____ reverse phase chromatography

B. Mobile phase is more polar than stationary

phase

____ size exclusion chromatography

C. Water in sample may cause interference because

of its attraction to the stationary phase.

____ ion-exchange chromatography

D. Smallest solute particles elute slowly.

8. A known mixture of compounds X and Y gave the following chromatographic results:

Compound
X
Y

Concentration (M)
0.0842
0.0599

Peak Area (arbitrary units)

423
347

Next, a solution containing an unknown concentration of compound X was analyzed: To 10.0 mL of

the unknown, 10.0 mL of a solution containing 0.146 M "Y" was added, and the mixture was diluted to
25.0 mL in a volumetric flask. This mixture yielded a chromatogram with two peaks with areas of 553
("X") and 339 ("Y"). What is the concentration of "X" in the unknown?

9. Give the best match for the terms in the first list with the characteristics in the second: (8 pt quest)
____ TCD

A. Detects the current change as the eluent

passes through a flame.

____ ECD

B. Detects the current change as the eluent

passes through a stream of high energy
electrons.

____ FID
C. Detects the current change due to the
difference in heat conductance of mobile phase
and analyte.
All of the above are: _______________________________________.

10 Give definitions of and describe the differences between WCOT, SCOT, and PLOT columns.
(6 pt quest)

11. Sketch a simple diagram of a GC. (6 pt quest)

12. A weak acid, HA, has a pKa of 6.5. If you are utilizing ion-exchange chromatography with an
anion exchange resin, is pH 8.0 a good pH to facilitate this acid binding to the stationary phase? Why
or why not? Hint: Which form predominates at pH 8.0?