Supplementary Material for: Pilot-scale Preparation of Broad-spectrum CBD: Extraction Optimization and Purification using Centrifugal Partition Chromatography

Introduction: Cannabinoids, a class of compounds found in Cannabis sativa L. have wide range of pharmacological properties. While Δ9-tetrahydrocannabinol (Δ9-THC) is strictly regulated due to its psychoactive effects, cannabidiol (CBD), a non-psychoactive compound, is permitted in certain countries. This study aimed to optimize the preparation of ethanolic cannabis extracts using response surface methodology (RSM) and to develop an effective system for removing Δ9-THC through centrifugal partition chromatography (CPC) to produce broad-spectrum CBD (hemp extract containing CBD and other compounds with minimal or no Δ9-THC).
Methods: Three variables and six responses were assessed to optimize extraction conditions. Predictions were made using Design-Expert® software, and experimental conditions were identified using the Box‒Behnken design. Extracts were analyzed via HPLC and a chromameter. Optimal conditions were used for pilot-scale extraction, and the CPC process was optimized by determining the partition coefficient of target cannabinoids in various solvent systems and the maximum sample load.
Results: The optimal extraction conditions were -31°C, 33 min, and a sample-to-solvent ratio of 1:8% w/v, with a desirability value of 0.576. Temperature was the most influential factor. Although the total yield decreased, this condition provided the highest concentration of light-colored cannabinoids and was successfully scaled up for three other cannabis samples. The optimal CPC solvent system, consisting of hexane/0.1% FA in ACN/20 mM ammonium formate at a ratio of 10/6.5/3.5% v/v, demonstrated a yield recovery of 89.3 ± 0.21% w/w with a maximum load of 5 grams’ sample per run. The resulting broad-spectrum CBD extract had a high CBD content (73.3 ± 0.37% w/w) and minimal Δ9-THC content (0.2 ± 0.00% w/w)
Conclusion: The BBD-RSM optimization of ethanolic cannabis extraction provided the highest cannabinoid concentration with a short extraction time and desirable appearance. The CPC process successfully separated Δ9-THC, yielding a high-purity, broad-spectrum CBD extract.