Foeniculum vulgare HPTLC ID - Vanillin/H2SO4 Reagent UV 365 nm

Fennel (seed) (Foeniculum vulgare)

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

1. 1 μL Eugenol ~0.1% in CH3OH
2. 3 μL Foeniculum vulgare-1 (seed)
3. 3 μL Foeniculum vulgare-2 (seed)
4. 3 μL Foeniculum vulgare-3 (seed)
5. 3 μL Foeniculum vulgare-3 (seed)
6. 3 μL Foeniculum vulgare-4 (seed)
7. 3 μL Foeniculum vulgare-5 (seed)
8. 3 μL Foeniculum vulgare-6 (seed)

Reference materials used here have been authenticated by macroscopic, microscopic &/or TLC studies according to the reference source cited below held at Alkemists Laboratories, Costa Mesa, CA. 

Stationary Phase Silica gel 60, F254, 10 x 10 cm HPTLC plates 

Mobile Phase toluene: ethyl acetate [9.5/0.5] 

Sample Preparation Method 0.5g+5ml dichloromethane, sonicate/centrifuge/decant, evaporate to dryness with N2, qs 1.0 ml Toluene 

Detection Method Vanillin/H2SO4 Reagent -> 110° C 5 min -> UV 365 nm 

Reference see British Pharmacopoeia, 2003

Source: Elan M. Sudberg, Alkemist Laboratories ^[2]

Anise fruit (seed) HPTLC ID - NP and PEG reagent, UV 366 nm (Image electronically enhanced)

Anise fruit (seed) (Pimpinella anisum)

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

1. 1 µL Anise fruit 1
2. 3 µL Anise fruit 1
3. 6 µL Anise fruit 1
4. 3 µL Anise fruit 2
5. 1 µL Caraway fruit 1
6. 3 µL Caraway fruit 1
7. 6 µL Caraway fruit 1
8. 3 µL Caraway fruit 2
9. 3 / 1µL Rutin, Caffeic acid (with incr. Rf)
10. 3 µL Bitter Fennel fruit 1
11. 3 µL Bitter Fennel fruit 2
12. 3 µL Sweet Fennel fruit 1
13. 3 µL Sweet Fennel fruit 2
14. 3 µL Wild Fennel fruit
15. 3 µL Fennel tea 

Reference Sample(s) Reference: Dissolve 3 mg of rutin in 1 mL of methanol. Dissolve 1 mg of caffeic acid in 1 mL of methanol. 

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

Mobile Phase Ethyl acetate, formic acid, water 15:1:1 (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: NP and PEG reagent; Preparation NP reagent: 1 g of natural products reagent in 200 mL ethyl acetate; Preparation PEG reagent: 10 g of polyethyleneglycole 400 in 200 mL dichloromethane; Use: Heat for 3 min to 100°C, dip (time 0, speed 5) in NP reagent while still hot, dry, dip in PEG reagent. 

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: orange fluorescent zone at Rf ~ 0.07; Caffeic acid: light bluish fluorescent zone at Rf ~ 0.77.

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. In the upper part of the chromatogram there are three prominent zones: a light blue zone at Rf ~ 0.88, a yellow zone at Rf ~ 0.76 and a light blue zone at Rf ~ 0.61. In the lower part of the chromatogram there is a sequence of three zones (yellow, light blue, yellowish) between Rf ~ 0.15 and 0.26. Right above the application position there is a yellow zone.

Test for adulteration: In the middle of the chromatogram there are neither yellow zones at Rf ~ 0.32 and Rf ~ 0.38 (red arrows, Caraway fruit) nor a faint dark blue zone at Rf ~ 0.32 (blue arrow, Bitter Fennel fruit) nor a light blue zone at Rf ~ 0.53 (yellow arrow, Sweet Fennel fruit).

Source: HPTLC Association ^[3]

Bitter Fennel (fruit) HPTLC ID - NP and PEG reagent, UV 366 nm (image electronically enhanced)

Bitter Fennel (fruit) (Foeniculum vulgare ssp. vulgare var. vulgare)

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

1. 1 µL Bitter Fennel fruit 1
2. 3 µL Bitter Fennel fruit 1
3. 6 µL Bitter Fennel fruit 1
4. 3 µL Bitter Fennel fruit 2
5. 1 µL Sweet Fennel fruit 1
6. 3 µL Sweet Fennel fruit 1
7. 6 µL Sweet Fennel fruit 1
8. 3 µL Sweet Fennel fruit 2
9. 3 / 1µL Rutin, caffeic acid (with incr. Rf)
10. 3 µL Wild Fennel fruit
11. 4 µL Fennel tea
12. 3 µL Anise fruit 1
13. 3 µL Anise fruit 2
14. 3 µL Caraway fruit 1
15. 3 µL Caraway fruit 2 

Reference Sample(s) Reference: Dissolve 3 mg of rutin in 1 mL of methanol. Dissolve 1 mg of caffeic acid in 1 mL of methanol. 

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

Mobile Phase Ethyl acetate, formic acid, water 15:1:1 (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: NP and PEG reagent; Preparation NP reagent: 1 g of natural products reagent in 200 mL ethyl acetate; Preparation PEG reagent: 10 g of polyethylene glycol 400 in 200 mL dichloromethane; Use: Heat the plate at 100°C for 5 min, dip (time 0, speed 5) in NP reagent while still hot, dry in air, dip in PEG reagent. 

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: orange zone at Rf ~ 0.07; Caffeic acid: light bluish zone at Rf ~ 0.77.

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. The chromatogram of the test solution shows a dark blue zone at Rf ~ 0.78 slightly above the position of reference caffeic acid and a dark blue zone at Rf ~ 0.31 (white arrows).

Test for other species: No yellow zone is seen at Rf ~ 0.17 (red arrows, Anise fruit, Sweet Fennel fruit). A light blue zone is neither seen at Rf ~ 0.53 (yellow arrow, Sweet Fennel fruit) nor at Rf ~ 0.62 (yellow arrow, Anise fruit, Caraway fruit). No brownish zones are seen at Rf ~ 0.32 and 0.38 (green arrows, Caraway fruit).

Source: HPTLC Association ^[4]