Authenticity of Essential Oils: A HPTLC Fingerprint Method

Ilona Trettin, Application Specialist (CAMAG), Melanie Broszat, Scientific Business Development Manager (CAMAG), Monika Baumle, Product Manager TLC, Matthias Nold, Product Manager Reference Materials

Due to the increasing popularity of natural essential oils as products for personal care, pharmaceuticals, or for aromatherapy, the global market for essential oils is growing rapidly.1 Unfortunately, this increases the likelihood of adulteration by the addition of cheaper oils or oil constituents to maximize profits. Since there are so many possible ways adulteration can occur, it is a challenge to analyze these products. An efficient method for the detection of such adulteration is the use of chromatographic fingerprinting methods, in particular, gas chromatography or as shown in this article, High-Performance Thin-Layer Chromatography (HPTLC).

To help analytical labs test for essential oil adulteration, we offer reference materials from HWI pharma services for twenty-five commonly used essential oils. Table 1 shows the ten most recent additions that were subjects of this study. These products are secondary reference materials, traceable to HWI primary reference materials quantified by qNMR (quantitative NMR) or mass balance. Similar to the vanilla extract reference materials shown in the article "Vanilla – Natural or Out of the Reaction Flask", for each product, a quantitative value is provided for at least one of the major components, while several other key components are qualitatively verified by GC-FID. You will find the complete range on our webpage for Essential Oils.

As mentioned, HPTLC is a fast and efficient fingerprinting method and is therefore the perfect alternative or complement for GC to test essential oils for authenticity. Fingerprints for a range of essential oil secondary reference standards from HWI are shown below, obtained with the standard HPTLC method for identification of essential oils (submitted to Ph. Eur. for evaluation) by comparison of RF values and colors of reference substances and matching zones in the oils.

While not all potential components of essential oils are detectable with this method (e.g., camphene, menthofurane, a-pinene, β-pinene or g-terpinene), the variety of detection methods (UV and chemical derivatization) increases the chance to detect adulteration.

Chromatographic conditions

Table 1. Essential Oil Reference Materials recently added.
Table 2.Reference Standards
* HWI primary reference material

Individual Reports for Each Oil

Bergamot Oil (cat. no. 05941501 batch: HWI01935)

Documentation under white light and UV 366 nm after derivatization with anisaldehyde reagent.

Bergamot Oil

Figure 1. From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073, batch: BCBS7535V); 2: linalool, 1 µL/mL (RF 0.25) (cat. no. 00350190, batch: HWI01135‑1); 3: 1.8 cineole, 10 µL/mL (RF 0.35) (cat. no. 00020590, batch: HWI00654‑2); 4: linalyl acetate, 2 µL/mL (RF 0.52) (cat. no. 49599, batch: BCBS8744V); 5: bergamot oil, 20 µL/mL (cat. no. 05941501, batch: HWI01935).

Cinnamon Oil (cat. no. 06031501 batch: HWI01937)

Documentation: under UV 254 nm prior to derivatization*, under white light and UV 366 nm after derivatization.

*Deviation from the method with additional detection mode

Cinnamon Oil

Figure 2.From left to right: HPTLC chromatograms under UV 254 nm prior to derivatization, under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 1 mg/mL (RF 0.15) (cat. no. 83073, batch: BCBS7535V); 2: linalool, 1 µL/mL (RF 0.25) (cat. no. 00350190, batch: HWI01135-1); 3: 1.8-cineole, 10 µL/mL (RF 0.35) (cat. no. 00020590, batch: HWI00654-2); 4: eugenol, 0.5 µL/mL (RF 0.37) (cat. no. 01050595, batch: HWI00301-1), 5: isoeugenyl acetate, 1 mg/mL (RF 0.40) (cat. no. 07055, batch: BCBV6080); 6: cinnamon oil, 30 µL/mL (cat. no. 06031501, batch: HWI01937).

Dwarf Pine Oil (cat. no. 06001501 batch: HWI01934)

Documentation: under UV 366 nm prior to derivatization*, under white light and UV 366 nm after derivatization

*Deviation from the method with additional detection mode

Dwarf Pine Oil

Figure 3.From left to right: HPTLC chromatograms under UV 366 nm prior to derivatization, under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073, batch: BCBS7535V); 2: bornyl acetate, 1.5 µL/mL (RF 0.50) (cat. no. 00400585, batch: HWI01713); 3: dwarf pine oil, 100 µL/mL (cat. no. 06001501, batch: HWI01934).

Frankincense Oil (cat. no. 05971501 batch: HWI011941)

Documentation: under white light and UV 366 nm after derivatization.

Frankincense Oil

Figure 4.From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073, batch: BCBS7535V); 2: (-)‑menthol, 1.5 mg/mL (RF 0.22) (cat. no. 00580590, batch: HWI00289-1); 3: linalool, 1 µL/mL (RF 0.25) (cat. no. 00350190, batch: HWI01135-1); 4: terpinen‑4‑ol, 1 µL/mL (RF 0.27) (cat. no. 03900590, batch: HWI00630-3); 5: frankincense oil, 200 µL/mL (cat. no. 05971501, batch: HWI01941).

Lemon Oil (cat. no. 05981501 batch: HWI01936)

Documentation: under UV 254 nm prior to derivatization*, under white light and UV 366 nm after derivatization.

*Deviation from the method with additional detection mode

Lemon Oil

Figure 5.From left to right: HPTLC chromatograms under UV 254 nm prior to derivatization, under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073, batch: BCBS7535V); 2: (R)‑(+)‑limonene, 120 µL/mL (main zone at RF 0.44; zones below due to impurities) (cat. no. 00590590, batch: HWI01323-2); 3: lemon oil, 80 µL/mL (cat. no. 05981501, batch: HWI01936).

Mint Oil (cat. no. 06011501 batch: HWI01952)

Documenttion: UV 254 nm prior derivatization*, under white light and UV 366 nm after derivatization.

*Deviation from the method with additional detection mode

Mint Oil

Figure 6.From left to right: HPTLC chromatograms under UV 254 nm prior to derivatization*, under white light and UV 366 nm after derivatization. Track 1: (-)‑menthol, 3 mg/mL (RF 0.22) (cat. no. 00580590, batch: HWI00289-1); 2: 1.8‑cineole, 10 µL/mL (RF 0.35) (cat. no. 00020590, batch: HWI00654-2); 3: (-)‑carvone, 0.35 mg/mL (RF 0.38) (cat. no. 00290595, batch: HWI00236-1); 4: (-)‑menthone, 20 µL/mL (main zone at RF 0.49; zones below due to impurities) (cat. no. 04660585, batch: HWI01363-5); 5: mint oil, 10 µL/mL (cat. no. 05981501, batch: HWI01936).

Patchouli Oil (cat. no. 05591501 batch: HWI01634-1)

Documentation: with the TLC Visualizer 2 under white light and UV 366 nm after derivatization.

Patchouli Oil

Figure 7.From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073, batch: BCBS7535V); 2: linalool, 1 µL/mL (RF 0.25) (cat. no. 00350190, batch: HWI01135-1); 3: 1.8‑cineole, 10 µL/mL (RF 0.35) (cat. no. 00020590, batch: HWI00654-2); 4: patchoulol, 4 mg/mL (RF 0.35) (cat. no. 05690595, batch: HWI01638-1); 5: patchouli oil, 20 µL/mL (cat. no. 05591501, batch: HWI01634-1).

Peppermint Oil (cat. no. 05541501 batch: HWI01631-1)

Documentation: under UV 254 nm prior derivatization*, under white light and UV 366 nm after derivatization.

*Deviation from the method with additional detection mode

Peppermint Oil

Figure 8.From left to right: HPTLC chromatograms under UV 254 nm prior to derivatization, under white light and UV 366 nm after derivatization. Track 1: (-)‑menthol, 3.0 mg/mL (RF 0.22) (cat. no. 00580590, batch: HWI00289-1); 2: 1.8‑cineole, 10 µL/mL (RF 0.35) (cat. no. 00020590, batch: HWI00654-2); 3: (-)‑carvone, 0.35 mg/mL (RF 0.38) (cat. no. 00290595, batch: HWI00236-1); 4: (-)‑menthone, 20 µL/mL (main zone at RF 0.49; zones below due to degradations below) (cat. no. 04660585, batch: HWI01363-5); 5: peppermint oil, 20 µL/mL (cat. no. 05541501, batch: HWI01631-1).

Pine Needle Oil (cat. no. 05991501 batch: HWI01932)

Documentation: under white light and UV 366 nm after derivatization.

Pine needle oil

Figure 9.From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073 batch: BCBS7535V); 2: (+)‑borneol, 0.5 mg/mL (RF 0.18) (cat. no. 68878, batch: 14627); 3: (-)‑carvone, 0.35 mg/mL (RF 0.38) (cat. no. 00290595, batch: HWI00236-1); 4: bornyl acetate, 1.5 µL/mL (RF 0.50) (cat. no. 00400585, batch: HWI01713); 5: pine oil, 100 µL/mL (cat. no. 05991501/a>, batch: HWI01932).

Sage Oil (cat. no. 06021501 batch: HWI01940)

Documentation: under white light and UV 366 nm after derivatization.

Sage oil

Figure 9.From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL (RF 0.15) (cat. no. 83073 batch: BCBS7535V); 2: (+)‑borneol, 0.5 mg/mL (RF 0.18) (cat. no. 68878, batch: 14627); 3: linalool, 1 µL/mL (RF 0.25) (cat. no. 00350190, batch: HWI01135-1); 4: 1.8-cineole, 10 µL/mL (RF 0.35) (cat. no. 00020590, batch: HWI00654-2); 5: bornyl acetate, 0.5 µL/mL (RF 0.50) (cat. no. 00400585, batch: HWI01713); 6: sage oil, 20 µL/mL (cat. no. 05581501, batch: HWI01633).

Spruce Needle Oil (cat. no. 05961501 batch: HWI01933)

Documentation: under white light and UV 366 nm after derivatization.

Spruce needle oil

Figure 10. From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 1 mg/mL (RF 0.15) (cat. no. 83073 batch: BCBS7535V); 2: (+)‑borneol, 0.5 mg/mL (RF 0.18) (cat. no. 68878, batch: 14627); 3: linalool, 1 µL/mL (RF 0.25) (cat. no. 00350190, batch: HWI01135-1); 4: bornyl acetate, 3 µL/mL (RF 0.50) (cat. no. 00400585, batch: HWI01713); 5: spruce needle oil, 100 µL/mL (cat. no. 05961501, batch: HWI01933).

Tea Tree Oil (cat. no. 05511501 batch: HWI01628-1)

Documentation: under white light and UV 366 nm after derivatization.

Tea tree oil

Figure 11.From left to right: HPTLC chromatograms under white light and UV 366 nm after derivatization. Track 1: (+)‑α‑terpineol, 0.5 mg/mL ( RF ; 0.15 ) (cat. no. 83073, batch: BCBS7535V); 2: terpinen-4-ol, 2 µL/mL ( RF ; 0.27 ) (cat. no. 03900590, batch: HWI00630-3); 3: 1.8‑cineole, 10 µL/mL ( RF ; 0.35 ) (cat. no. 00020590, batch: HWI00654-2); 4: tea tree oil, 15 µL/mL (cat. no. 05511501, batch: HWI01628-1).

Ylang Ylang Oil (cat. no. 05951501 batch: HWI01993)

Documentation: under UV 254 nm prior to derivatization*, under white light and UV 366 nm after derivatization.

*Deviation from the method with additional detection mode

Ylang ylang oil

Figure 12. From left to right: HPTLC chromatograms under UV 254 nm prior to derivatization, under white light and UV 366 nm after derivatization. Track 1: linalool, 1 µL/mL ( RF 0.25 ) (cat. no. 00350190, batch: HWI01135-1); 2: geranyl acetate, 0.5 µL/mL (RF 0.50) (cat. no. 01290190, batch: HWI01134); 3: ylang ylang oil, 15 µL/mL (cat. no. 05951501, batch: HWI01993).

Materials
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