APPLICATION NOTES

HPTLC Identification of Goldenseal (Hydrastis canadensis)

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Sample:
In a test tube 0.25 g powdered drug are extracted with 4 mL
methanol - water (8:2) for 30 min in an ultrasonic bath at
room temperature. The mixture is filtered and the residue
washed twice with 2 mL methanol each. In a 20 mL
volumetric flask filtrate and washings are combined and filled
up to the mark with methanol. This is the test solution.
Note: Samples are sensitive to light and heat. Store in the
refrigerator in colored vials.
Standards (optional):
In a 50 mL volumetric flask, 25 mg (1R,9S)--hydrastine HCl,
1 mg hydrastinine HCl, 1.2 mg palmatine chloride and 1.2 mg
berberine chloride are dissolved in methanol and filled up to
the mark.
Derivatization reagent (optional):
Ninhydrin reagent: 0.6 g ninhydrin are dissolved in 190 mL
isopropanol, then 5 mL acetic acid are added.
Chromatographic conditions:
Stationary phase:

HPTLC plates silica gel 60 F254 (Merck), 10x10 cm or

20x10cm.

Mobile Phase:

Ethyl acetate, methanol, formic acid, water (50:10:6:3)

Sample application:

5 L test solution and standard are applied as 8 mm bands,

min. 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:

a) UV 254 nm
b) UV 366 nm
c) Ninhydrin reagent: the plate is immersed in reagent for
1 s, dried in stream of cold air, then heated at 110C for
2 min. Examination in white light.

06/03

Please contact CAMAG for more application notes and products for analysis of herbals!

F-15

APPLICATION NOTES

Results:
Compare to the chromatograms below:

1 a UV 254 nm

1 b UV 366 nm

HPTLC of golden seal

1

Track 1: Hydrastinine
Track 2: Hydrastine
Track 3: Golden seal (extract)
Track 4: Palmatine chloride
Track 5: Berberine chloride
Track 6: Mixture of standards
Note: Palmatine must not be seen in the sample!

1 c Ninhydrin reagent / white light:

06/03

Please contact CAMAG for more application notes and products for analysis of herbals!

F-15

APPLICATION NOTES

Comparison of Goldenseal and common adulterants:

10

11

2a

10

11

2b

10

11

2c

06/03

Please contact CAMAG for more application notes and products for analysis of herbals!

F-15

APPLICATION NOTES

10

11

2 b UV 366 nm overexposed
Caption Figures 2a-c
Track 1:

Standard mixture: hydrastinine, hydrastine, palmatine, berberine (with increasing Rf) 5 L

Track 2:

Rumex crispus

Track 3:

Mahonia aquafolium

Track 4:

Coptis chinensis

Track 5:

Xanthorrhiza sp. leaves

Track 6:

Hydrastis canadensis (root) authenticated

Track 7:

Xanthorrhiza sp. steam

Track 8:

Xanthorrhiza root

Track 9:

Berberis nervosa

Track 10:

Berberis vulgaris

Track 11:

Standard mixture: hydrastinine, hydrastine, palmatine, berberine (with increasing Rf) 3 L

06/03

Please contact CAMAG for more application notes and products for analysis of herbals!

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