Meirelles João Victor, Estevam Débora Cristina Diniz, Dos Santos Vanessa Farelo, Pereira Henrique Marcelo Gualberto, Saint Pierre Tatiana D, Veiga-Junior Valdir F, Padilha Monica Costa
Departamento de Química, Pontifícia Universidade Católica do Rio de Janeiro, Rio de Janeiro 22451-045, RJ, Brazil.
Laboratório Brasileiro de Controle de Dopagem (LBCD), Instituto de Química, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-909, RJ, Brazil.
Biomolecules. 2025 Feb 8;15(2):246. doi: 10.3390/biom15020246.
The market is experiencing steady global growth. herbal extracts (CHE) are interesting and sought-after products for many clinical conditions. The medical potential of these formulations is mainly attributed to neutral cannabinoids, such as cannabidiol (CBD), tetrahydrocannabinol (THC), and cannabinol (CBN), and their non-standardized content poses a significant fragility in these pharmaceutical inputs. High-resolution mass spectrometry portrays a powerful alternative to their accurate monitoring; however, further analytical steps need to be critically optimized to keep up with instrumental performance. In this study, Full Factorial and Box-Behnken designs were employed to achieve a multivariate optimization of CBD, THC, and CBN extraction from human and veterinary commercial CHE using a minimum methanol/hexane 9:1 volume and short operational times. A predictive model was also constructed using the Response Surface Methodology and its accuracy was validated. Agitation and sonication times were identified as the most significant operational extraction parameters, followed by the extraction mixture volume. The final setup of the optimized method represented a significantly faster and cheaper protocol than those in the literature. The selected neutral cannabinoids quantification was conducted using ultra high-performance liquid chromatography coupled to high-resolution tandem mass spectrometry (UHPLC-HRMS/MS) with a precision of <15%, accuracy of 69-98%, sensitivity of 23-39 ng kg, and linearity regarding pharmaceutical requirements. State-of-the-art levels of analytical sensitivity and specificity were achieved in the target quantification due to high-resolution mass spectrometry. The developed method demonstrated reliability and feasibility for routine analytical applications. As a proof-of-concept, it enabled the efficient processing of 16 samples of commercial CHE within a three-hour timeframe, showcasing its practicality and reproducibility, and highlighting its potential for broader adoption in similar scenarios for both human and veterinary medicines.
市场正经历着稳定的全球增长。草药提取物（CHE）对于许多临床病症来说都是有趣且受欢迎的产品。这些制剂的医学潜力主要归因于中性大麻素，如大麻二酚（CBD）、四氢大麻酚（THC）和大麻酚（CBN），而它们未标准化的含量在这些药物原料中构成了显著的脆弱性。高分辨率质谱法是对其进行精确监测的有力替代方法；然而，需要对进一步的分析步骤进行严格优化以跟上仪器性能。在本研究中，采用全因子设计和Box-Behnken设计，使用最低甲醇/己烷9:1体积比并缩短操作时间，对从人和兽医商业CHE中提取CBD、THC和CBN进行多变量优化。还使用响应面方法构建了预测模型并验证了其准确性。搅拌和超声处理时间被确定为最显著的操作提取参数，其次是提取混合物体积。优化方法的最终设置代表了一种比文献中的方法显著更快且更便宜的方案。使用超高效液相色谱与高分辨率串联质谱联用（UHPLC-HRMS/MS）对选定的中性大麻素进行定量，精密度<15%，准确度为69 - 98%，灵敏度为23 - 39 ng/kg，符合药物要求的线性度。由于高分辨率质谱法，在目标定量中实现了分析灵敏度和特异性的先进水平。所开发的方法在常规分析应用中显示出可靠性和可行性。作为概念验证，它能够在三小时内高效处理16个商业CHE样品，展示了其实用性和可重复性，并突出了其在人和兽用药品类似场景中更广泛应用的潜力。
