A one-pot cannabinoid analysis

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  • Published: Jan 4, 2010
  • Author: Steve Down
  • Channels: HPLC
thumbnail image: A one-pot cannabinoid analysis

Cannabis is the most popular drug of abuse in Europe with an estimated 4 million adults using it daily. It has been the subject of official confusion in the UK where the government does not seem to be able to make up its mind on the classification grade. It was downgraded from class B to class C in 2004 but was reclassified to class B in January 2009, where it resides with amphetamines.

One reason given for the U-turn is the emergence of skunk, a stronger variety of cannabis that contains higher levels of the active ingredient tetrahydrocannabinol (THC). The constant evolution of street cannabis has put pressure on analytical labs to measure the content of the active cannabinoids in seized specimens.

Forensic organisations are not the only people interested in the chemical make up of Cannabis sativa plants. Clinical labs are also concerned about the cannabinoid composition, due to its medicinal uses.

The common procedure for measuring the cannabinoids in cannabis is GC but it has some disadvantages. The inherent heating process brings about decomposition of the acidic cannabinoids via decarboxylation. The fact that this degradation is only partial further complicates matters. The resultant neutral products require derivatisation before they can be analysed by GC.

A logical alternative would be HPLC because this does not generally require the input of heat. Several published HPLC procedures exist but they appear to be restricted by inefficient separation of the target cannabinoids or have not been validated. In an attempt to correct this, researchers from Belgium have developed and validated an HPLC-DAD method for the determination of the major cannabinoids in cannabis.

Benjamin De Backer and co-researchers from the universities of Liege and Ghent extracted cannabinoids from seized samples of drug-type cannabis. Their method was geared towards Delta9-tetrahydrocannabinol (THC), THC acid, cannabidiol (CBD), cannabidiol acid, cannabigerol (CBG), cannabigerol acid, cannabinol and Delta8-tetrahydrocannabinol.

These compounds are either related in the cannabinoid synthetic pathway or are the principal decarboxylation or isomerisation products. They were selected because they are important for the classification of cannabis phenotypes or for monitoring the psychotropic potency.

The team used design of experiment protocols and predictive multilinear models to find the best chromatographic conditions for optimal separation of the target cannabinoids. With a C18 column and an aqueous methanol gradient, these were determined to be a starting level of 68% methanol in aqueous 50 mM ammonium formate (pH 5.19), rising linearly to 90.5% over 25 minutes at a flow rate of 0.3 ml/min.

Under these conditions, the predicted and real chromatograms were very similar for model compounds, producing good separation between the peaks. The only potential overlap occurred with CBD and CBG at relatively high concentrations of each of more than 10%. The excellent separation was confirmed for cannabinoids extracted from herbal cannabis and cannabis resin.

The procedure was optimised using accuracy profiles based on â-expectation tolerance intervals, as well as calculating values for trueness (expressed as relative bias %), precision and measurement uncertainty. All of the values were deemed to be acceptable.

The new optimised method brings about the separation of 8 key cannabinoids within 25 minutes and allows for their identification and measurement in cannabis samples. It was demonstrated for cannabis drug and fibre.

De Backer predicted that the method would be useful for the identification of plant phenotypes, quality control of medicinal specimens and the estimation of the psychoactive potency.

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.



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