Source: Royal Botanic Gardens, Kew.^[1]

European vervain herb (aerial parts) HPTLC ID - UV 254 nm

European vervain herb (aerial parts) (Verbena officinalis)

Lane Assignments Lanes, from left to right (Track, Volume, Sample):

1. 3 µL European vervain herb 1
2. 5 µL European vervain herb 1
3. 7 µL European vervain herb 1
4. 3 µL European vervain herb 1 (Ph.Eur. extraction)
5. 5 µL European vervain herb 1 (Ph.Eur. extraction)
6. 7 µL European vervain herb 1 (Ph.Eur. extraction)
7. 5 µL Rutin
8. 5 µL Arbutin
9. 3 µL European vervain herb 2
10. 5 µL Lemon verbena leaf 1
11. 7 µL Lemon verbena leaf 1
12. 5 µL Lemon verbena leaf 1
13. 5 µL Lemon verbena leaf 2
14. 5 µL Lemon verbena leaf 3
15. 5 µL European vervain herb 3 

Reference Sample(s) Reference: Dissolve 1 mg of rutin in 5 mL of methanol; Dissolve 1 mg of arbutin in 5 mL of methanol. 

Stationary Phase Stationary phase, i.e. Silica gel 60, F254 

Mobile Phase Formic acid, acetic acid, water, ethyl acetate 11:11:27:100 (v/v/v/v) 

Sample Preparation Method Sample: Mix 500 mg of powdered sample with 5 mL of methanol and sonicate for 10 minutes, then centrifuge or filter the solutions and use the supernatants / filtrates as test solutions.

Derivatization reagent: Anisaldehyde reagent, Preparation: 170 mL of ice-cooled methanol are mixed with 20 mL of acetic acid, 10 mL of sulfuric acid and 1 mL of anisaldehyde, Use: Dip (time 0, speed 5), heat at 100°C for 5 min 

Detection Method Saturated chamber; developing distance 70 mm from lower edge; relative humidity 33% 

Other Notes Images presented in this entry are examples and are not intended to be used as basis for setting specifications for quality control purposes.

System suitability test: Rutin: light yellow zone at Rf ~ 0.34 (white RT). Arbutin: light brown zone at Rf ~ 0.46 (white RT).

Identification : Compare result with reference images. The fingerprint of the test solution is similar to that of the corresponding botanical reference sample. Additional weak zones may be present. Under UV 254 nm the chromatogram of the test solution shows a quenching zone at Rf ~ 0.40 and at Rf ~ 0.53. Under white RT there is a greyish violet zone at Rf ~ 0.40 right above the position of reference rutin, and another greyish violet zone below rutin at Rf ~ 0.22. Just above the position of reference arbutin there is a grey zone at Rf ~ 0.54 (blue arrows). Around Rf ~ 0.07 there are two yellow zones.

Test for adulteration: The chromatogram of Lemon verbena leaf does not show a zone at Rf ~ 0.40 (red arrow).

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  European vervain herb (aerial parts) HPTLC ID - UV 254 nm

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  European vervain herb (aerial parts) HPTLC ID - Anisaldehyde reagent, white RT

Source: HPTLC Association ^[6]