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Phenolics
Sample:
1 g milled sample is sonicated with 10 mL methanol for 5 min.
The extract is filtered through a 0.2 m syringe filter.
Standards (optional):
2 mg echinacoside, 2 mg cynarine, 1 mg chlorogenic acid,
1 mg caffeic acid, 0.5 mg caftaric acid, and 0.5 mg chicoric
acid are dissolved in 10 mL methanol each.
Derivatization reagent:
Natural Products reagent (NP reagent): 1 g diphenylborinic
acid aminoethylester is dissolved in 200 mL ethyl acetate.
Chromatographic conditions:
Stationary phase: HPTLC plates silica gel 60 F254 (Merck), 10x10 cm or
20x10 cm.
Mobile Phase: Ethyl acetate, ethylmethyl ketone, formic acid, water
(15:9:3:3)
Sample application: 5 L of test solution and 2 L of each standard are applied as
8 mm bands, at least 2 mm apart, 8 mm from lower edge of
plate.
Development: 10x10 cm or 20x10 cm Twin Trough Chamber, saturated for
20 min (filter paper), 5 mL (respectively 10 mL) developing
solvent per trough, developing distance 60 mm from lower
edge of plate. The plate is then dried with a hair dryer (cold
air) for 5 min.
Detection: NP reagent: the plate is heated at 100C for 3 min, then
dipped while still hot in NP reagent and dried in a stream of
cold air. Examination under UV 366 nm.
11/03 Please contact CAMAG for more application notes and products for analysis of herbals! F-24A
APPLICATION NOTES
Results:
Compare to the chromatograms below:
Roots
1 2 3 4 5 6 7 8 9 1: Echinacea purpurea root
2: Echinacea pallida root
3: Echinacea angustifolia root
4: Echinacea atrorubens root
5: Caftaric acid, cynarine, chicoric
acid (with increasing Rf)
6: Echinacoside, chlorogenic acid,
caffeic acid (with increasing Rf)
7: Liatris punctata root (adulterant)
8, 9: Parthenium integrifolium root
Echinacea purpurea root contains caftaric and chicoric acids. (adulterant)
Echinacea pallida root contains echinacoside.
Echinacea angustifolia root contains echinacoside and
a low amount of cynarine.
Echinacea atrorubens root shows none of the markers.
The profiles of the adulterants dont match any of the standards.
11/03 Please contact CAMAG for more application notes and products for analysis of herbals! F-24A