The CBD fraction of column chromatography can be distilled (bp 187-190° C/2 mm; pale yellow resin) to purify it. Isomerization can be accomplished with any of several solvents and acids. Alcohol and sulfuric acid isomerizes only 50-60% of CBD to THC; p-TolueneSulfonic Acid (p-TSA) in petroleum ether or other light, non-polar solvent will convert 90% of CBD to THC upon refluxing 1 hour at 130° F.
(16, 17) Reflux 3 gr CBD in 100 ml dry benzene for 2 hours with 200 mg p-TSA monohydrate until the alkaline Beam test (5% KOH in ethanol) is negative (no color). Separate the upper layer, wash it with 5% sodium bicarbonate, wash again with water, and strip the solvent. The remaining viscous oil should give a negative reaction to the Beam test. The crude THC can be purified by distillation (bp 169-172° C/0.03 mm), or by chromatography in 25 ml pentane on 300 gr alumina. Elute with pentane 95:5 ether to yield fraction of CBD and THC. Combine the THC fractions and distill (bp 175-178° C/1 mm). Reflux 2 gr CBD in 35 ml cyclohexane, and slowly add a few drops of sulfuric acid.
Continue to reflux until the Beam test is negative. Separate the sulfuric acid from the reaction mixture. Wash the solution twice with aqueous sodium bicarbonate, the twice again with water. Purify by chromatography, or distill (bp 165° C/0.01 mm). Another method is to reflux a mixture of 6 gr dry pyridine hydrochloride and 3 gr CBD at 125° C until the Beam test is negative. Wash the reaction mixture with water to remove the pyridine, then extract the mixture with ether. Wash the ether with water, evaporate the ether, and distill the residue i.v. Similarly, reflux 3 gr CBD in 150 ml ethanol with 50 ml 85% phosphoric acid until the Beam test is negative. Alternatively, reflux 3 gr CBD in 100 ml absolute ethanol containing 0.05% HCl for 19 hours. Extract the ether, wash the ether with water, dry, evaporate, and chromatograph on 400 gr alumina to yield: (a) 0.5 gr 1-EthoxyHexaHydro-CBN (EHH-CBN: mp 86-87° C); elute with pentane 98:2 ether. Repeated chromatography will separate the less polar forms. (c) 0.5 gr EHH-CBN, eluted with pentane 93:7 ether. It can be isomerized to THC by refluxing in benzene for 2 hours. Cool the reaction mixture, wash it with water; separate, dry, and strip the solvent layer i.v. CBD also can be isomerized by irradiation of a cyclohexane solution in a quartz vessel with a mercury lamp (235-265 nm) for 20 minutes. (18-20) THC gives an acetate (ATHC) which is as potent as THC. Wohlner, et al., prepared ATHC by refluxing the crude distillate of cannabis oil with approximately 3 volumes of acetic anhydride. Cahn prepared ATHC thus: add 150 ml acetyl chloride (dropwise with stirring and cooling) to 185 gr crude resin in 500 ml dry pyridine. Crystals may separate during the addition, or on standing a few hours at room temperature. Pour the mixture into dilute hydrochloric acid/ice. Wash this solution with dilute acid, then with aqueous sodium carbonate, and again with water. Strip the solvent and distill the residue (240-270 C°/20 mm). The mixture of acetylated cannabinoids is separated by dissolving 2 gr in 100 ml benzene and chromatography over silica (150-200 mesh).
Combine the washings and the original effluent solutions, then strip the benzene i.v. The material remaining on the column contains CBD and other cannabinoid acetates which can be recovered with ethanol and worked up. (21) Colorimetric tests are the simplest method of identifying cannabinoids. Hundreds more sophisticated analytical methods have been developed, as a review of Chemical Abstracts will reveal. It gives a purple color with 5% ethanolic KOH, based on the oxidation of CBD, CBG, etc., and their acids to hydroxyquinones. Only two plants (Rosemary and Salvia) out of 129 common species tested give a weakly positive reaction. Among some 50 pure vegetable substances such as mono- and sesqui-terpenes, aromatics, etc., only juglone, embelin, and alkyl dioxyquinone develop a color reaction close to that of Cannabis. The reaction is not always dependable; it can be absent if the ethanol is hot. (22, 23) A modification of the Beam test uses absolute ethanol saturated with gaseous hydrogen chloride.
When added to an extract of suspect material, it gives a cherry red color which disappears if water is added.