Advancing Cannabinoid Profiling with UHPLC-MS Chromatography

A team comprising members of the Brazil’s Federal University of Espírito Santo, the National Institute of Forensic Science and Technology, and the Brazilian Federal Police (both in Porto Alegre, Brazil) optimized the extraction process of six cannabinoids in cannabis oil and marijuana samples. The researchers also performed an analytical validation of a quantitative and qualitative method for seven cannabinoids using ultrahigh-performance liquid chromatography coupled with low-resolution mass spectrometry (UHPLC-LTQ-MS). The optimization showed that ethyl acetate is the most suitable solvent for oil extraction. A paper based on their work was published in ACS Omega (1).

Historically, the cultivation Cannabis sativa L. has been done for different purposes, such as medicinal use and as a drug, among others (2,3). The plant’s wide applicability is attributed to its various constituents, with over 550 compounds identified, especially a class of terpenophenolic compounds known as cannabinoids (2,4). Cannabinoids produce effects through their interaction with specific receptors located within different parts of the central nervous system. Simply put, cannabinoids regulate how cells communicate—how they send, receive, or process messages (5). While there have been previous studies using high-performance liquid chromatography (HPLC) and UHPLC to identify and quantify cannabinoids, mainly coupled to spectrophotometric detection, it is rare that a complete validation of the methodology is performed, resulting in a failure to improve chromatographic separation for various isomers, and a lost opportunity to enhance the extraction process by evaluating different variables (6-9), thus inspiring this Brazilian study.

For the marijuana samples, the researchers performed extraction using the same solvent and sonication and vortex agitation for 10 and 7.5 min, respectively. In the presented method, no matrix effect was observed, where limit of quantification (LOQ) ranged between 1 and 5 ng mL^-1 and limit of detection (LOD) between 0.3 and 1.5 ng mL^-1. Recovery ranged from 84.6 to 107.6% in oil and from 80.6 to 105.9% in marijuana. Precision was evaluated in three ways: within a single day (relative standard deviation [RSD]: 3.14-10.87% in oil, 3.25-10.14% in marijuana), across different days with a second analyst (RSD: 1.98-10.71% in oil, 4.65-12.81% in marijuana), and laboratories with a third analyst (RSD: 5.59-13.94% in oil, 4.65-13.56% in marijuana). The proposed method offers high sensitivity, selectivity, and precision, being adequate and satisfactory for the quantification of cannabidiol (CBD), cannabinol (CBN), cannabichromene (CBC), cannabidiolic acid (CBDA), Δ⁹-tetrahydrocannabinol (Δ⁹-THC), and Δ⁹-tetrahydrocannabinol acid (Δ⁹-THCA) (1).

“The extraction,” reported the authors of the paper, “demonstrated a satisfactory yield for all of the cannabinoids evaluated, standing out for its speed and simplicity across the different matrices analyzed.” They additionally noted that LC coupled with MS “proved to be an exceptional technique for quantitative cannabinoid analysis, offering high sensitivity and selectivity.” The researchers believe that their results contribute to the advancement of analytical methodologies for cannabinoid quantification as well as establish a solid foundation for future research in the field (1).

Read More on Similar Topics:

Study Shows Py-GC-MS Outperforms Traditional GC-MS for THC Edible Forensics

References

1. de Almeida, J. V. M.; Conceição, N. S.; Pereira, A. R. et al. Development and Validation of a Cannabinoid Quantification Method in Oil and Marijuana by UHPLC-MS. ACS Omega 2025, 10 (47), 57385-57393. DOI: 10.1021/acsomega.5c07689
2. dos Santos, N. A.; Romão, W. Cannabinomics Studies – Cannabis – A State of the Art About the Millenary Plant: Part I. Forensic Chem. 2023, 32, 100470. DOI: 10.1016/j.forc.2023.100470
3. Nascimento, I. R.; Costa, H. B.; Souza, L. M. et al. Chemical Identification of Cannabinoids in Street Marijuana Samples Using Electrospray Ionization FT-ICR Mass Spectrometry. Anal. Methods 2015, 7, 1415, DOI: 10.1039/C4AY02355B
4. Rock, E. M.; Parker, L. A. Constituents of Cannabis sativa. In Cannabinoids and Neuropsychiatric Disorders; Murillo-Rodriguez, E.; Pandi-Perumal, S. R.; Monti, J. M. (eds), Springer, 2021, DOI: 10.1007/978-3-030-57369-0_1
5. Cannabinoids. Alcohol and Drug Foundation website. https://adf.org.au/drug-facts/cannabinoids/ (accessed 2025-12-10)
6. Carvalho, V. M.; de Almeida, F. G.; de Aguiar, A. F. L. et al. Facing the Forensic Challenge of Cannabis Regulation: A Methodology for the Differentiation between Hemp and Marijuana Samples. Braz. J. Anal. Chem. 2022, 9, 162, DOI: 10.30744/brjac.2179-3425.ar-42-2021
7. Hsu, Y. H.; Fang, M. C.; Huang, S. C. et al. Determination of cannabinoids in hemp oil based cosmetic products by LC-tandem MS. J. Food Drug Anal. 2021, 29, 502, DOI: 10.38212/2224-6614.3370
8. Kim, S. Y.; Kim, J. Y.; Kwon, W. et al. Method Development for Simultaneous Determination of Amphetamine Type Stimulants and Cannabinoids in Urine Using GC–MS. Microchem. J. 2013, 110, 326. DOI: 10.1016/j.microc.2013.04.004
9. Hädener, M.; König, S.; Weinmann, S. Quantitative Determination of CBD and THC and their Acid Precursors in Confiscated Cannabis Samples by HPLC-DAD. Forensic Sci. Int. 2019, 299, 142. DOI: 10.1016/j.forsciint.2019.03.046