Workflow for Cannabinoids Analysis in Cannabis Using A Chromolith® Monolithic Silica HPLC Column Providing Low Backpressure and Extended Column Lifetime

by | Jun 23, 2023

Discover an HPLC-DAD workflow to determine 14 cannabinoids in hemp bud samples using a monolithic silica-based Chromolith® HPLC column.

This article from Issue 14 of the Analytix Reporter describes an HPLC-DAD workflow to determine 14 cannabinoids in hemp bud samples using a monolithic silica-based Chromolith® HPLC column, offering good selectivity and efficiency along with demonstrating solid robustness.


The legal use of recreational and medical cannabis is expanding globally along with hemp-based products (based on cannabidiol), for health and wellness. Hemp is defined legally in various geographies as cannabis varieties with limits on total tetrahydrocannabinol (THC) content. To ensure consumer safety, cannabis and hemp products need to be tested to determine accurate potency of the active cannabinoid constituents. Cannabis products in the market range from plants to distillates, and edibles to cosmetics. This broad variety of matrices underscores the need for robust columns and high throughput analytical methods.

This work provides a complete HPLC-DAD (high performance liquid chromatography-diode array detection) workflow for cannabinoids analysis using robust Chromolith® HighResolution (HR) HPLC columns which are based on monolithic silica. Chromolith® HPLC columns enable fast and cost-efficient separations due to low column backpressure and the very high robustness of the column. The low backpressure allows fast separation at high flow rates with the same mobile phase consumption per sample compared to slower low flow-rate methods. The workflow offers the following:

  • Detailed hemp bud sample preparation for HPLC-UV analysis
  • Fast and cost-efficient separation with Chromolith® monolithic silica HPLC columns with low back pressure to determine potency of hemp bud sample
  • Demonstration of robustness of Chromolith® column
  • Separation of 14 cannabinoids within 10 minutes
  • Calibration curve preparation using certified reference materials (CRMs)

Experimental Conditions
In this work, hemp bud samples were analyzed to determine their potency. Sample preparation involved ethanol extraction of cannabinoids from plant material. The extract was then analyzed applying an HPLC-UV method and using a Chromolith® HR RP-18e monolithic silica HPLC column. Quantitation was performed utilizing a 6-point calibration curve obtained from HPLC-UV analysis of standard solutions prepared from CRMs. Peaks were identified using the retention times from a chromatogram of a 14 cannabinoids mix. Cannabinoid peaks were also verified by comparing UV absorption spectra of both samples and standards. Furthermore, robustness of the monolithic silica based Chromolith® was demonstrated via retention time stability and separation efficiency after 1400 injections.

Results and Discussion
Hemp bud sample was homogenized at low temperature to prevent analyte degradation using cryo-cup grinder followed by double extraction with ethanol. Resulting solution was diluted, filtered, and subjected to HPLC-DAD analysis. Calibration curves were obtained by analyzing solutions prepared from CRMs. Cannabinoids in hemp bud extract were identified based on retention time match with standards and cross verified with UV absorption spectra.

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