Source: MOBOT, Tropicos.org^[1]

“… Cannabis sativa can be identified by microscopic structures on the surface of the plant, namely, by trichomes. Two types of trichomes occur and can be observed with a binocular microscope with a magnification factor of 40: (a) Non-glandular trichomes are numerous, unicellular, rigid and curved hairs, with a slender pointed apex: - Cystolithic trichomes found on the upper surface of the cannabis leaves have a characteristic bear claw shape and may have calcium carbonate crystals (cystoliths) visible at their bases. Frequently, the trichome is broken and the cystolith freed; - Non-cystolithic trichomes occur mainly on the lower side of the leaves, bracts and bracteoles and lack the enlarged base; - The simultaneous presence of these bear claw-shaped trichomes on the upper surface and the fine, slender non-cystolithic trichomes on the lower surface of the leaves is a characteristic of cannabis. (b) Glandular trichomes. They occur as: - Sessile glands, i.e. trichomes without stalk, which are generally found on the lower epidermis; - Small bulbous glandular trichomes with one-celled stalks; - Long multicellular stalks on the bracteoles surrounding the female flowers (multicellular stalked glandular trichomes). These are mainly associated with the flower structures (pistillate plants being particularly rich in these structures) but they can also be found on the underside of the leaves and occasionally on the stems of young plants. Some plants possess trichomes that may be confused with those present on Cannabis sativa and care should be taken in definitive identification. However, the combination of cystolithic hairs on the leaf upper surface and longer trichomes and sessile glands on the lower surface, which is unique to Cannabis sativa, enables positive identification of even fragmented material.” Source: Recommended methods for the identification and analysis of cannabis and cannabis products, United Nations Office on Drugs and Crime, Vienna, 2009. (ST/NAR/40) [5]

Transverse section in the midrib of the leaflet Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[6]	Upper epidermis of the leaflet Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[7]	Lower epidermis of the leaflet Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[8]

University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]]

Non-glandular trichrome Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[9]	Cystolithic trichrome Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[10]	Epidermal cells with cystolithic trichrome Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[11]

University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]]

Cannabis spp. Internal Standard Solution: 1 mg/mL 4-androstene-3,17-dione in a mixture of chloroform and methanol (1:9). Test Sample Preparation: 100 mg ground plant material in 3 mL of Internal Standard Solution, macerate at room temperature for 1 h,sonicate for 5 min, filter, and use the filtrate. Column: 15-m x 0.25-mm, 0.25-um film, J&W Scientific DB-1 Oven Program: 170°C for 1 min, increase to 250°C at 10°C/min, hold at 250° for 3 min. Injection Port: 240°C Detector: FID, 260°C Carrier Gas: Helium Injection volume: 1 uL Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA [12]

Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[13]	Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[14]	Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[15]	Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA[16]

University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]] [[Category:National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA]]

Cannabis spp. (rhizome) HPTLC ID - White RT

Cannabis spp. (rhizome) HPTLC ID

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

1. 2 uL Δ9-Tetrahydrocannabinol (Δ9-THC or THC)
2. 2 uL Δ9-Tetrahydrocannabinolicacid (THCA)
3. 5 uL THC-type Cannabis
4. 5 uL Intermediate-type Cannabis
5. 5 uL Fiber-type Cannabis
6. 2 uL Cannabidiolic acid (CBDA)
7. 2 uL Cannabidiol (CBD)
8. 2 uL Cannabichromene (CBC)
9. 2 uL Cannabigerol (CBG)
10. 2 uL Cannabinol (CBN)
11. 2 uL Tetrahydrocannabivarin (THCV)
12. 5 uL Intermediate-type Cannabis decarboxylated 

Reference Sample(s) Reference Standard Solutions: 1.0 mg/mL of each Δ9-Tetrahydrocannabinol, Δ9-tetrahydrocannabinolicacid, and cannabidiol in methanol.

Reference Sample Preparations:100 mg of powdered plant material in 10 mL of dichloromethane, sonicate for 1 h, filter, and evaporate the extract to dryness under nitrogen. Dissolve the residue in 10 mL of methanol. 

Stationary Phase Stationary Phase:TLC, Silica gel 60RP-18 F254, 150 um (Sorbent Technologies) 

Mobile Phase Mobile Phase:0.1% glacial acetic acid in a mixture of methanol and water (75:25) 

Sample Preparation Method Test Sample Preparation: Prepare test sample as described under Reference Sample Preparations and apply 5uL. Derivatization reagent: Fast blue reagent– 5 mg/mL fast blue B salt in water. 

Detection Method Development:Saturated chamber, developing distance 70 mm from lower edge of the plate; relative humidity 33%, temperature 25°.

Detection: Heat plate at 70°C for 2 min, dip (time 5, speed 5) in Derivatization reagent, dry, and examine under white light. 

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.

Identification: Compare Test Sample Preparation chromatogram with chromatograms of Reference Standard Solutions and Reference Sample Preparations. The Test Sample Preparation chromatogram is similar to that of the THC-type Reference Sample Preparation chromatogram. Additional weak zones may be present. The most intense zones in the chromatogram of the Test Sample Preparation are a purple zone in the lower-third section and a purple zone in the upper-third section of the chromatogram corresponding to THC and THCA, respectively,in the chromatograms of the Reference Standard Solutions and the THC-type Reference Sample Preparation.

The Test Sample Preparation chromatogram exhibits no zones or a minor zone at about one-third the chromatogram and at an Rf corresponding to cannabidiol in the chromatograms of the Reference Standard Solutions (cf.Intermediate-type Cannabis and Fiber-type Cannabis). The Test Sample Preparation chromatogram exhibits minor zones in the upper-third of the chromatogram, above the zone corresponding to THCA in the chromatograms of the Reference Standard Solutions (cf. Intermediate-type Cannabis and Fiber-type Cannabis).

Source: National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS 38677, USA ^[17]