Analysis of Caffeine in Dietary Products by Multiple Injection Capillary Electrophoresis - Caffeine: Chemistry, Analysis, Function and Effects - page 158

d n 0 t 2 n g | 4

Figure 9.2

Capillary electrophoresis separations of caffeine and acetaminophen for
different conditions. Representative electropherograms of caffeine and
acetaminophen in the absence (A) and presence (B) of 15 mg mL -1
sulfated b-cyclodextrin (S-b-CD). The separations were performed in a
fused silica capillary of dimensions: 50 mm 6 65 6 75 cm; Applied
voltage 25 kV; Hydrodynamic injection (0.1 psi) for 10 seconds; BGE
50 mM phosphoric acid (pH 7.0) containing 10% ACN (v/v %), 15 mg mL -1
S-b-CD and 20 mM NaCl; capillary temperature 20 o C; UV detection at
270 nm. Peaks: 0, water; 1, caffeine; 2, acetaminophen.

in the presence of sulfated b-cyclodextrin (see Figure 9.2), which are excellent
conditions for the conversion of the method to multiple-injection CE mode I
(Amini et al 2008). Acetaminophen (Figure 9.1(B)) was selected as an injection
marker based on the fact that acetaminophen as a very weak acid (pK a # 9)
co-migrated with caffeine at the selected pH, Figure 9.2 (A). In addition,
acetaminophen like caffeine exhibited a satisfactory UV absorption at 270 nm.
Detection at this wavelength contributed to the specificity of the method since
spectral interferences of background and other substances present in the
extract samples were suppressed. In order to further prevent the matrix effects,
as well as to improve the solubility of caffeine, the extract was diluted. The
extent of dilution was dependent on the caffeine concentration in the sample.
These

measures also facilitated the application of the multiple-injection

technique, where interferences are not desirable.
On the basis of previous experience sulfated b-CD (S-b-CD), as an effective
complexing agent, was added to the BGE for the separation of the IM and
caffeine (Amini et al 2006), Figure 9.1 (C). b-cyclodextrin is a cyclic oligo-
saccharide containing seven D -(+)-glucose units linked by a-1,4-glycosidic
bonds, Figure 9.1 (C). It possesses a central hydrophobic cavity and a
hydrophilic outside. The hydrophobic cavity of the CD is able to form
inclusion complexes with hydrophobic compounds of appropriate size.
As Figure 9.2 (B) shows, base-line separation was achieved in less than
fifteen minutes in the presence of 15 mg mL -1 S-b-CD. This concentration was
considered to be adequate for the separation and conversion of the method to
multiple-injection CE mode I (Amini et al 2008). This multiple-injection CE
mode is considered to be suitable for compounds that exhibit small migration