Introduction

Cannabidiol (CBD) is a non-psychotropic cannabinoid from the Cannabis sativa L. (Cannabaceae) plant. Researchers have shown that CBD’s medicinal properties may be of use for treating a variety of conditions including pain, inflammation, epilepsy, and cancer.1,2 Recently, the US Food and Drug Administration (FDA) approved the first cannabis-derived drug (Epidiolex®) to treat children with rare forms of epilepsy.

Changes in the legal status of cannabis for medical use, along with the increasing number of therapeutic applications for CBD, has resulted in rapidly growing demand for highly pure CBD. As such, it is crucial to develop standardized CBD products free of unwanted components to ensure patient safety. CBD manufacturers will require efficient, cost-effective methods that can produce high yields of impurity-free products. In this article, we describe a rapid and reproducible method for large-scale purification of pure CBD using Centrifugal Partition Chromatography (CPC).

Challenges of producing highly pure CBD

Chromatographic purification of natural compounds can be challenging due to the complex nature of the starting materials. Crude cannabis oil contains about 400 potentially active compounds. Phytocannabinoids are compounds found in the sticky resin of trichomes, that densely cover the surfaces of female inflorescences and, to a lesser degree, the foliage of male and female hemp plants. For the safe use of CBD in possible treatments, it must be purified from unwanted compounds in a hemp extract, such as tetrahydrocannabinol (THC), the main psychoactive chemical in the plant.

While traditional chromatography techniques, such as high-performance liquid chromatography (HPLC) - preparative scale and flash chromatography are effective for other applications, they are not always adapted for large-scale CBD purification. The major reason is that the purification of CBD by these methods is a two-step process requiring silica resin, an expensive consumable that must be replaced frequently due to irreversible absorption of a variety of compounds during the separation process. In addition, these methods use large quantities of solvents to elute natural compounds. These factors make the use of tools like HPLC or flash chromatography time consuming and cost prohibitive for large-scale production.

What is CPC?

CPC is a liquid purification technique that does not require traditional solid supports like silica. Instead, it relies on two immiscible liquid phases. One serves as the mobile phase or the eluent, and the other as the stationary phase, which replaces the silica-based column in flash chromatography and HPLC. The stationary phase is retained in the column by a centrifugal field, generated by rotation of the CPC column. Once injected, the compounds to purify are eluted by the mobile phase flow in order of their partition coefficients, defined by their relative affinity for each of the CPC liquid phases (Figure 1). The reusable CPC column consists of several stacked disks, each engraved with numerous twin cells, linked by ducts. This configuration provides better retention of the stationary phase, allows for higher elution flow rates, and improves separation times.

Advantages of CPC for CBD purification

CPC offers numerous benefits for CBD purification. It can be used to purify complex mixtures, such as crude extracts of cannabis, in just one step. By relying on a liquid stationary phase, CPC columns do not need to be replaced like traditional silica-based columns and cartridges used in preparative HPLC and flash chromatography methods. The CPC column can be loaded with different solvents to create the column needed, greatly reducing costs. Purification parameters can be adjusted according to the targeted compound or desired purity level to achieve THC-free extracts, highly purified phytocannabinoids, or pharmaceutical-grade products.

CPC is also easy to scale up for processing milligrams to kilograms of product efficiently, whereas preparative HPLC and flash chromatography may require substantial stationary phase changes to optimize the purification methodology as it moves to a larger scale.

Another key advantage of CPC, besides the possibility to work with higher elution flow rates, therefore faster run times, is the lack of irreversible absorption of compounds. In addition, CPC uses significantly less solvent resulting in reduced consumable costs.

High recoveries of pure CBD using CPC

In addition to cost and time savings, manufacturers need a method that yields highly pure product with high recoveries. To demonstrate the benefits of CPC for CBD purification, 5 grams of crude hemp extract were injected into a Gilson CPC 250 PRO and PLC 2250 Purification System (Figure 2). Using this one-step method, they obtained 205 milligrams of over 99% pure CBD, as determined by HPLC analysis.

Summary

As applications and medical uses for CBD expand, the demand for pure CBD will continue to grow. Manufacturers need efficient, cost-effective methods to meet these demands. CPC offers high yields, purity, and is adaptable from lab-scale to industrial-scale use. By eliminating the need for silica-based columns and greatly reducing solvent use, CPC is a more economical approach to CBD purification than traditional chromatography techniques.

References

1. Russo, E.B. (2017) Cannabidiol claims and misconceptions. Trends Pharm. Sci. http://dx.doi.org/10.1016/j.tips.2016.12.004.

2. Zhornitsky, S. and Potvin, S. (2012) Cannabidiol in humans: the quest for therapeutic targets. Pharmaceuticals. http://dx.doi.org/10.3390/ph5050529.