Zingiber officinale – essential oil

1. Scope
This method identifies essential oils of dried roots and rhizomes of Zingiber officinale Roscoe
by HPTLC fingerprint.

2. Source of method
Ilona Trettin, HPTLC Association

3. Procedure
Test solution Dissolve 10.0 µL of essential oil in 1.0 mL of toluene.
Reference solutions 0.25 µL/mL of isoeugenol and 0.5 mg/mL of isoeugenyl acetate in
toluene.
Optional: 0.2 µL/mL of geraniol, 1.0 µL/mL of citral, and
0.5 mg/mL of each of 6-gingerol and 6 shogaol in toluene.
Stationary phase HPTLC Si 60 F254 (Merck)
Application volume 15 tracks, band length 8 mm, track distance 11.4 mm, distance
from left edge 20 mm, distance from lower edge 8 mm, application
volume 2.0 µL of reference solutions and test solution.
Developing solvent Ethyl acetate, toluene 5:95 (v/v)
Developing distance 70 mm from lower edge
Saturation time 20 min, with a saturation pad
Relative humidity 33%, saturated MgCl2
Temperature 22 ± 5°C
Derivatization reagent Anisaldehyde reagent
Preparation: Slowly and carefully mix 170 mL of ice-cooled
methanol with 20 mL of acetic acid and 10 mL of sulfuric acid.
Allow the mixture to cool to room temperature, then add 1 mL of
anisaldehyde (p-methoxy benzaldehyde).
Use: Derivatize (Derivatizer: 3 mL, blue nozzle, spraying level 3),
heat plate at 100°C for 3 min.
Detection A) Underivatized, UV 254 nm
B) Underivatized, UV 366 nm
C) Underivatized, white light RT
D) Derivatized, UV 366 nm
E) Derivatized, white light RT

4. Results
Note: These chromatographic fingerprints are representative of the samples used in this
analysis. Fingerprints obtained may vary from sample to sample. Analysts must validate the
most appropriate fingerprint for their identity standard.
System suitability test (SST) (under UV 254 nm after derivatization)

International Association for the Advancement of High Performance Thin Layer Chromatography
A non-profit organization dedicated to the promotion of HPTLC in plant analysis and other analytical fields • www.hptlc-association.org
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  isoeugenol: greyish zone at RF ~ 0.36
 isoeugenyl acetate: greyish zone at RF ~0.40
Specific markers (under white light after derivatization)
 6-gingerol: brownish-purple zone at RF ~0.02
 geraniol: purple zone at RF ~0.13
 6-shogaol: brownish-purple zone at RF ~0.19
 citral: brownish-purple zone at RF ~0.33
Different origins

Figure 1: HPTLC fingerprints UV 254 nm (A), UV 366 nm (B) and white light (C) prior to derivatization, and UV 366
nm after derivatization (D), and white light after derivatization (E).

Track Sample Origin

1 Isoeugenol, isoeugenyl acetate (with increasing RF) --
2 6-gingerol, 6-shogaol (with increasing RF)
3 Geraniol, citral (with increasing RF)
4-9 Zingiber officinale – essential oil of roots and rhizomes Unknown

Version Revision history Released by

1 Created by: TD/16 Mar 2020 M Sharaf / 7 Apr 2020

International Association for the Advancement of High Performance Thin Layer Chromatography
A non-profit organization dedicated to the promotion of HPTLC in plant analysis and other analytical fields • www.hptlc-association.org
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