Page 261 - Vogel's TEXTBOOK OF QUANTITATIVE CHEMICAL ANALYSIS

THIN-LAVER CHROMATOGRAPHV - THE RECOVERV OF SEPARATEO SUBSTANCES BV ELUTION TECHNIOUES 8.9

8.8 DETERMINATION OF ASPIRIN, PHENACETIN AND CAFFEINE IN A MIXTURE
High performance liquid chromatography is used for the separation and
quantitative analysis of a wide variety of mixtures, especially those in which the
components are insufficiently volatile and/or thermally stable to be separated
by gas chromatography. This is illustrated by the following method which may
be used for the quantitative determination of aspirin and caffeine in- the common
analgesic tablets, using phenacetin as internal standard; where APC tablets are
available the phenacetin can also be determined by this procedure.
Sample mixture. A suitable sample mixture is obtained by weighing out
accuratelyabout 0.601 g of aspirin, 0.076 g of phenacetin and 0.092 g of caffeine.
Dissolve the mixture in 10 mL absolute ethanol, add 10 mL of 0.5 M ammonium
formate solution and dilute to 100 mL with de-ionised water.

Sohent (mobile phase). Ammonium formate (0.05M) in 10 per cent (v/v)
ethanol-water at pH 4.8. Use a flow rate of 2 mL min-' with inlet pressure of
about 117 bar (1 bar = 105 Pa).
Column. 15.0 cm x 4.6 mm, packed with a 5 pm silica SCX (strong cation
exchanger) bonded phase.
Detector. UV absorbance at 244 nm (or 275 nm).
Procedure. Inject 1 pL of the sample solution and obtain a chromatogram.
Under the above conditions the compounds are separated in about 3 minutes, the
elution sequence being (1) aspirin; (2) phenacetin; (3) caffeine. Measure peak
areas with an integrator and normalise the peak area for each compound (i.e.
express each peak area as a percentage of the total peak area). Compare these
results with the known composition of the mixture; discrepancies arise because
of different detector response to the same amount of each substance.
Determine the response factors (r) for the detector relative to phenacetin
( = 1) as internal standard by carrying oui three runs, using 1 pL injection, and
obtaining the average value of r.

Peak area of compound/Mass of compound
Relative response factor, r =
Peak area of standard/Mass of standard
Correct the peak areas initially obtained by dividing by the appropriate response
factor and normalise the corrected values. Compare this result with the known
composition of the mixture.

8.9 THIN-LAYER CHROMATOGRAPHY - THE RECOVERY OF SEPARATED SUBSTANCES
BY ELUTION TECHNIQUES
The purpose of the experiment is to illustrate the elution technique for the
recovery of pure substances after their separation by thin-layer chromatography.
The experiment can be readily extended to include the quantitative determination
of the recovered substances.
Apparatus. Prepared silica gel plates.
Chromatographic tank (see Fig. 8.6).
Drummond (or similar) micropipette.