สารบัญ

Thai name  สารสกัดกัญชา (SAN SAKAT KANCHA)

Category  Anti-emetic; anti-epileptic; analgesic.

Cannabis Extract is the soft extract prepared from the extraction of the powdered, dried and mature pistillate inflorescences of Cannabis sativa L. by means of supercritical fluid (carbon dioxide) extraction or ethanol extraction.
It contains not less than 80.0 per cent and not more than 120.0 per cent of the labelled amounts of tetrahydrocannabinol (C₂₁H₃₀O₂) and cannabidiol (C₂₁H₃₀O₂); the labelled amounts of tetrahydrocannabinol and cannabidiol are not less than 30.0 per cent and 1.0 per cent, respectively, calculated on the dried basis.

Description  Yellowish brown to dark brown paste or viscous liquid; odour, characteristic.

Other relevant information  Cannabinol may be found in some samples as it is a degraded product of tetrahydrocannabinol due to improper storage.

Packaging and storage  Cannabis Extract shall be kept in tightly closed containers, preferably in amber glass containers, protected from light, and stored at a temperature not exceeding 25º.

Labelling  The label on the container states the amounts of tetrahydrocannabinol and cannabidiol.

Identification

     A. To 50 mg of the sample in a screw-capped, amber test-tube, add 10 mL of a mixture of 9 volumes of methanol and 1 volume of chloroform, sonicate for 30 minutes and allow to cool to room temperature.
Filter through a 0.45-μm polyvinylidene fluoride filter (Vertical^® or equivalent is suitable.) until a clear solution is obtained. Fold a filter paper into a quarter and open it partly to form a funnel, place a few drops of the solution into the centre of the
paper and allow the filter paper to dry. Add a very small amount of a 1 per cent w/w mixture of Fast Blue B salt and anhydrous sodium sulfate and a few drops of a 10 per cent w/v solution of sodium bicarbonate: a purple red colour develops at the centre of the filter paper.
     (Note This colour is a combination of the colours of the different cannabinoids which are the major components of cannabis: tetrahydrocannabinol is red, cannabidiol is orange and cannabinol is purple.)

     B. Carry out the test as described in the “Thin-Layer Chromatography” (Appendix 3.1).
     Standard solution A  A solution containing 500 μg per mL of Tetrahydrocannabinol RS in methanol.
     Standard solution B  A solution containing 1 mg per mL of Cannabidiol RS in methanol.
     Standard solution C  A solution containing 1 mg per mL of Cannabinol RS in methanol.
     Test solution  To 30 mg of the sample in a screw-capped, amber test-tube, add 2 mL of methanol. Filter through a 0.45-μm polyvinylidene fluoride filter (Vertical^® or equivalent is suitable.). 
     Adsorbent  Octadecylsilyl silica gel F254 (a precoated plate by Sorbent  Technologies, Inc. or equivalent is suitable).
     Mobile phase  Methanol, water and glacial acetic acid (75:25:0.1).
     Application  Apply 1 μL of each of Standard solutions and Test solution, as 10-mm bands.
     Development and drying  Allow the solvent front to ascend 7 cm above the line of application. Dry the developed plate in air.
     Detection  Examine the plate under ultraviolet light (254 nm). Spray the plate with Fast Blue B salt TS and observe under visible light.
     Results  When examined under ultraviolet light (254 nm), the test solution shows two quenching bands (tetrahydrocannabinol and cannabidiol) or three quenching bands (tetrahydrocannabinol, cannabidiol and cannabinol) at the lower third of the chromatogram corresponding in R_f to the bands shown by the standard solutions.  When examined under visible light, the test solution shows a red band due to tetrahydrocannabinol, an orange band due to cannabidiol and/or a purple band due to cannabinol, corresponding in colour and R_f to the bands obtained from the standard solutions A, B and C, respectively.

     C. The chromatogram of the Assay preparation shows several peaks, two of which correspond to the tetrahydrocannabinol peak and the cannabidiol peak of the Standard preparations, as obtained in the Assay.

Loss on drying  Not more than 10.0 per cent w/w after drying at 105º for 2 hours (Appendix 4.15).

Heavy metals  Complies with the requirements described under “Limit Test for Heavy Metals in Herbal Drugs and Herbal Drug Preparations” (Appendix 5.2); see also under “Arsenic and Heavy Metals” in the General Notices.

Pesticide residues  Complies with the requirements in the “Pesticide Residues” (Appendix 7.22H).

Assay Carry out the determination as described in the “Liquid Chromatography” (Appendix 3.5).
     Diluent  Prepare a mixture of the equal volumes of acetonitrile and water.
     Standard stock preparation A  Dissolve an accurately weighed quantity of Tetrahydrocannabinol RS in Diluent to obtain a stock solution having a known concentration of about 1 mg per mL.
     Standard stock preparation B  Dissolve an accurately weighed quantity of Cannabidiol RS in Diluent to obtain a stock solution having a known concentration of about 500 μg per mL.
     Internal standard preparation  Dissolve an accurately weighed quantity of Diazepam RS in Diluent to obtain a stock solution having a known concentration of about 500 μg per mL. Dilute this solution with Diluent to obtain a solution having
a final concentration of about 50 μg per mL.
     Standard preparations  Dilute Standard stock preparation A and Standard stock preparation B, quantitatively, and stepwise with Diluent to obtain five solutions having known concentrations of about 2, 25, 50, 75, and 100 μg each of tetrahydrocannabinol and cannabidiol per mL. Upon dilution, add sufficient amount of Internal standard preparation so that each final solution contains diazepam at a concentration of about 50 μg per mL.
     System suitability preparation  Dilute Standard stock preparation A and Standard stock preparation B, quantitatively, and stepwise with Diluent to obtain a solution having known concentrations of about 50 μg each of tetrahydrocannabinol and cannabidiol per mL. Upon dilution, add sufficient amount of Internal standard preparation so that each final solution contains diazepam at a concentration of about 50 μg per mL.
     Assay preparation  Transfer about 50 mg of Cannabis Extract, accurately weighed, to a screw-capped, amber test-tube, add 10 mL of a mixture of 9 volumes of methanol and 1 volume of chloroform, sonicate for 30 minutes and allow to cool to room temperature. Filter through a 0.45-μm polyvinylidene fluoride filter (Vertical^® or equivalent is suitable.) until a clear solution is obtained.  Evaporate 75.0 μL of the filtrate to dryness under a fume hood. Dissolve the residue in 2.0 mL of Internal standard preparation, sonicate for 2 minutes and filter through a polyvinylidene fluoride membrane having a 0.45-μm porosity.
     (Note In case the concentrations of tetrahydrocannabinol and cannabidiol in the Assay preparation are beyond the concentration ranges of standard curve, allow to adjust the volume of the filtrate taken for evaporation.)
     Mobile phase A  Dissolve 3.153 g of ammonium formate in a 10 per cent v/v solution of acetonitrile and dilute with the same solvent to 1000.0 mL.  Adjust with formic acid to a pH of 3.75±0.05.
     Mobile phase B  Prepare a 90 per cent v/v solution of acetonitrile.
The step gradient of mobile phases is as follows:

 Chromatographic system
     Detector  Ultraviolet light (228 nm)
     Column  A stainless steel column (15 cm × 4.6 mm), packed with octadecylsilane chemically bonded to porous silica or ceramic microparticles (3.5 μm) equipped with a similarly packed guard column (5 mm × 3.9 mm)
     Column temperature  30°±1°
     Flow rate  1.0 mL per minute.
     (Note  Adjustments are not recommended for gradient elutions, as stated in “Chromatographic Separation Techniques” (Appendix 3.9), unless it is necessary to obtain the relative retention times of about 0.2, 0.6 and 1 for diazepam, cannabidiol and tetrahydrocannabinol, respectively.)
     System suitability
         Sample  System suitability preparation
     Suitability requirements
         Symmetry factor  Not more than 2.0 for the tetrahydrocannabinol peak and the cannabidiol peak.
         Relative standard deviations  Chromatograph five injections of System suitability preparation, and record peak areas. The analytical system is suitable for conducting this Assay if the relative standard deviation for the area ratios of tetrahydrocannabinol to diazepam and cannabidiol to diazepam are not more than 2.0 per cent for the tetrahydrocannabinol peak and the cannabidiol peak, respectively.
     Procedure  Separately inject about 30 μL each of Standard preparations into the chromatograph, record the chromatograms and measure the areas of the diazepam, cannabidiol and tetrahydrocannabinol peaks, respectively. Calculate the area ratios of cannabidiol to diazepam and the area ratios of tetrahydrocannabinol to diazepam. Plot the standard curves of the calculated area ratios of cannabidiol and tetrahydrocannabinol against their respective concentrations. Inject about 30 μL of Assay preparation into the chromatograph, record the chromatogram and measure the peak areas and calculate the area ratios, as with the Standard preparation.
     Calculation  By reference to the standard curve, calculate the contents of tetrahydrocannabinol (C₂₁H₃₀O₂) and cannabidiol (C₂₁H₃₀O₂) in a portion of the Extract taken.
     Other requirements  Complies with the requirements described under “Extracts” (Appendix 1.16).