Sample prep-free determination of Caffeine in shampoo by using TLC-MS

Abstract

The analysis of shampoo for caffeine determination is an excellent application to demonstrate the easy handling and good performance of TLC-MS (thin layer chromatography – mass spectrometry) using the elution-based approach. High matrix tolerance of TLC makes chromatographic separation fast and economical, as only a minimum of sample preparation is needed and direct sample application on the HPTLC Si60 F254 MS-grade plate is possible. Caffeine was extracted with the TLCMS Interface from Camag and afterwards analysed with an ESI mass detector.

Results

After chromatographic separation caffeine could be identified with an hRf value of 54. For quantification UV peak area was used. A six point calibration and a triple determination of caffeine in the shampoo sample results in an average concentration of 94 mg corresponding to the usual amount of 0.1% active ingredient in the formulation. Figure 1a shows the scan at 254 nm. Beside the bands of caffeine, matrix compounds appear at the starting point and close to the solvent front (Fig. 1b).

For confirmation the separated caffeine was eluted with the TLC-MS Interface into the mass spectrometer. The mass spectrum shows the protonated molecule at m/z 195.2 (Fig. 2).

 

UV spectrum of caffeine, 1b: bands of caffeine standard

Figure 1a: UV spectrum of caffeine, 1b: bands of caffeine standard (1-3, 7-9) and separated shampoo (4-6), chromatographic data shown on next page.
 

Mass spectrum and structure of caffeine recorded at an hRf value of 54 at track 4

Figure 2: Mass spectrum and structure of caffeine recorded at an hRf value of 54 at track 4.

 

Conclusion

These results demonstrate that caffeine can easily be analysed qualitative as well as quantitative by TLC-MS. Due to excellent chromatographic separation no interference of the analyte caffeine and the matrix occurs, although just a minimum of sample preparation was performed.

Application data

Chromatography
Plate HPTLC Silica gel 60 F254 MS-grade, 20x10 cm
Sample preparation 15 min. stirring in 2-Propanol, filtration (0.45 μm)
Sample application ATS 4 sample applicator (Camag), 6 mm bandwise
Application volume 0.5 – 5 μL
Mobile phase 2-Propanol / n-Heptane / Water 7/3/1 (v/v/v)
Migration distance 5 cm
Migration time 50 min
Extraction
Extraction equipment “TLC-MS Interface” from Camag
Extraction solvent Acetonitrile / Water 95/5 (v/v) + 0.1% Formic acid
Extraction flow 0.1 mL/min
Detection
Documentation equipment documentation unit Reprostar / Digistore (Camag)
Wavelength UV 254 nm
Staining Anisaldehyde sulphuric acid reagent
MS equipment single-quadrupole mass spectrometer
MS detection ESI (+) mode MS (m/z 100 – 500)

Chromatographic data

Track Compound Conc. [mg/mL] Application volume [μL] hRf Detected mass m/z
1-3 Caffeine standard 0.10 0.1, 0.3, 0.5 54 195.2
4-6 Shampoo - 5.0 54 195.2
7-9 Caffeine standard 0.10 0.8, 1.0, 2.0 54 195.2

Ordering information

Description Product No.
HPTLC Silica gel 60 F254 MS-grade, 20x10 cm 1.00934
Acetonitrile hypergrade for LC-MS LiChrosolv® 1.00029
2-Propanol gradient grade for liquid chromatography LiChrosolv® 1.01040
N-Heptane gradient grade for liquid chromatography LiChrosolv® 1.04390
Formic acid for analysis EMSURE® 1.00264
Water for chromatography (LC-MS Grade) LiChrosolv® 1.15333
Millex®-FH filter, 0.45 μm hydrophobic PTFE, 25 mm, non-sterile SLFH025NS

 

Materials