Use of an “Intrinsic” Internal Standard to Quantitate CBD in Hemp Seed Samples

Use of an “Intrinsic” Internal Standard to Quantitate CBD in Hemp Seed Samples

Hemp is a non-psychoactive form of the Cannabis sativa L. plant that is used for a wide range of applications and whose chemical composition is therefore of ongoing interest. Hemp seeds, in particular, are used as both a food and pharmaceutical source, with a nutritional profile thought to offer several health benefits. This has resulted in a growing demand for rapid and reliable analytical methods for characterizing the compounds found in hemp seed products.

A Range of Approaches
A number of methods have been used to analyze the cannabinoids found in C.sativa and its derivatives. Cannabidiol (CBD), the primary cannabinoid in C.sativa, is typically analyzed using conventional methods with chromatographic separation like HPLC or GC coupled to an array such as UV or FID, or mass spectrometer. Quantitative approaches that do not require a chemical standard, such as ERETIC, have also been used. But these approaches have limitations, including the range of applications to which they are suited, the need for time-consuming pre-analysis preparation, and the potential thermal conversion of cannabinoids. Overall, there is a need for more innovative approaches to metabolite profiling, particularly in the absence of reference standards.

One such approach is quantitative NMR (qNMR). Like other NMR methods, qNMR provides the benefits of allowing direct analysis of samples without chemical or physical pre-treatment that may modify its molecular composition. It is fast, accurate, and highly specific method that can deliver all of the information needed in a single spectrum. But so far, it has not been used widely to analyze hemp seed products.

但是，QNMR确实需要使用内部或外部标准。这可能会带来一个缺点，因为库存解决方案的准备时间可能很耗时。用作内部标准的有机化合物可能会引入样品污染。

为了克服这些问题，意大利卡拉布里亚大学的一个团队开发了一种QNMR方法。该方法在同位素不纯净的氘化氯仿中使用未标记的氯仿作为“内在”内标。

A New Standard
The team’s approach centers on using isotopically impure deuterated chloroform, with 99.5% purity and therefore containing 0.5% of unlabeled residual chloroform. The resonance from this nondeuterated chloroform can then be used as an internal standard for calibrating spectra. Its weight in dissolved samples is known, therefore it can simultaneously be used to quantitate the analyte in the sample.
The researchers used the approach to quantitate the CBD content of hemp seed flour, cakes, and oil. Hemp seed cakes are composed of fiber and oily residues produced as a by-product of oil extraction from the seeds and are sometimes used in animal feed.

该团队使用了带有Z轴梯度线圈和温度控制单元的5 mM BBO探针的Bruker Avance 300 NMR光谱仪。他们首先创建了一种库存解决方案，其中将纯CBD样品溶解以生成校准曲线。使用此解决方案，团队评估了几个参数，以确认该方法的可靠性，例如线性范围，准确性，检测限和量化限制，然后再进行测试样品。他们确定其样品的CBD浓度在蛋糕提取物中为1.65 mg/ml，在大麻种子粉提取物中的CBD浓度为1.28 mg/ml，在大麻籽油中的CBD浓度为2.16 mg/ml。

研究人员强调了该技术提供详细的光谱信息的能力，而无需在矩阵采样或色谱方法中进行化学或物理操作。取而代之的是，矩阵直接在氯仿溶液中提取。该方法不需要将有机化合物用作内部标准，这可以引入样品污染，并且也很快且经济。他们说，它可能可以用于测试大麻种子产品中的其他大麻素。

Bruker Avance Technology
在这个study, the researchers used a Bruker Avance NMR spectrometer. The latest generation of Avance technology from Bruker is the Avance Neo, which incorporates a range of innovative technology to deliver enhanced digital control, improved dynamic range and greater flexibility and scalability than ever before.
This includes its multi-receive capabilities, in which each NMR channel acts as both transmitter and receiver. This allows each channel to effectively operate as an independent spectrometer and multiple experiments to be conducted simultaneously. The Avance Neo also comes with Bruker’s industry-standard TopSpin software and is compatible with user control via the cloud.

全球大麻行业通过开放新的应用领域（如药物患者治疗）来显示出巨大的增长率。CBD是该地区最有希望的组成部分之一。通过提供最具创新的技术，Bruker使大麻行业能够确保拥有真实，一致的原材料和加工商品的安全和声音供应链，包括在医疗应用中有效的功效。必威手机客户端
Bruker does not support, encourage, or intend that its products or services be used in connection with any illegal use, cultivation or trade of cannabis or cannabis products. Bruker products are intended to be used only in compliance with all applicable laws in a manner that promotes public safety and in connection with any lawful and approved scientific or medical research activities.

参考
Siciliano C, Bartella L, Mazzotti F et al. 1H NMR quantification of cannabidiol (CBD) in industrial products derived from Cannabis sativa L. (hemp) seeds. IOP Conf. Series: Materials Science and Engineering 2019; 572:012010. doi: 10.1088/1757-899X/572/1/012010.