PRELIMINARY ANALYSIS OF FLAVONOID CONTENT IN LEGAL CANNABIS PRODUCTS
Author: Luke Khoury, Senior Scientist
In July 2020, Sonoma Lab Works completed a preliminary analysis of legal cannabis products to measure flavonoid profiles, an aspect of the cannabis metabolome which has long been unaddressed in the industry. All of the samples included in this small study have already undergone California compliance testing and are accessible to consumers throughout the state. The LC-MS/MS based assay has been developed to provide sensitive quantitation of these phytochemicals in order to begin documenting their prevalence within the variety of cannabis products currently in the market. It is important to provide consumer education of common products, especially in an industry slowly emerging from total prohibition. Furthermore, quality control laboratories not only serve as critical checkpoints in the supply chain but often yield opportunities to discover and share new information from the data that is generated. Quantitative flavonoid analysis will enable new insight into the metabolomic expression of cannabis and can be translated into new product development and breeding strategies.
What are flavonoids, though? And why should consumers be aware of them? Flavonoids are a subset of polyphenols, which comprise a family of diverse chemical classes of natural products that typically exhibit bioactivity. Flavonoid biosynthesis is ubiquitous among plant species and their biological functions are diverse. Ranging from regulating plant development, pigmentation, and UV protection, to an array of roles in defense and signaling between plants and microorganisms.1 Blueberries, onions, tomatoes, lettuce, celery, tea, and red wine all contain flavonoids. Perhaps surprisingly, some of those are also found in cannabis.2-6 Flavonoids have undergone investigation for decades regarding their roles in the health benefits associated with plant-rich diets. These include neuroprotective, antioxidant, and anticancer properties as demonstrated in several animal models.7-9 More recently, the biosynthesis of prenylated flavones unique to cannabis, cannflavin A and B, is being investigated. They are known for their potent anti-inflammatory properties and by understanding the metabolomic expression of these compounds, researchers aim to precisely engineer their production for therapeutic applications.10 The marked health benefits of plants are often overlooked but while probing the deep complexity of cannabis chemistry, it is refreshing to be reminded that these types of compounds can be found throughout common dietary lifestyles. For a more detailed review of flavonoids and other natural products in cannabis, see Flavonoids, Alkaloids, Terpenoids.11
Upon entering a typical California cannabis dispensary, one immediately notices the overwhelming number of different products available. Innovation in the cannabis space is seemingly endless and the proliferation of ideas translated into new products signals the rapid growth of the industry. However, not all products are created equally. Therefore, it is important to be aware of what various flowers, prerolls, concentrates, tinctures, capsules, topicals, and edibles contain. These types of products were investigated in this study. Although they do not span all product types, they represent several of those regularly consumed. The quantitative assay measured apigenin, baicalin, beta-sitosterol, cannflavin A, chrysin, orientin, rutin, vitexin, and wogonin in 17 samples. The analyte panel consists of flavonoid aglycones, glycoside complements, and a phytosterol. The flower and preroll samples showed similar flavonoid profiles and concentrations. Both contained beta-sitosterol (approximately 2000 µg/g) and cannflavin A (approx. 200 µg/g). A variety of other flavonoids were also detected including apigenin, orientin, rutin, and vitexin. The flower and preroll samples resulted in total mass percent concentrations of 0.23% and 0.25%, respectively. On the other hand, concentrates showed highly variable profiles that appeared dependent on specific manufacturing processes and formulations. Vape cartridges, hydrocarbon extracts (HCE), rosin, hashish, and whole plant extracts (WPE) all together contained combinations of beta-sitosterol, cannflavin A, rutin, and vitexin with total concentrations ranging from 0.26% to undetectable amounts. The hash-rosin sample contained the greatest concentration of cannflavin A (approx. 2000 µg/g) among all samples in the study. Notably, a THC-dominant WPE (a non-inhalable concentrate orally consumed for medical applications) contained the highest concentration of flavonoids of all concentrate samples. Two tinctures, olive oil- and MCT oil- based, were tested and resulted in strikingly different profiles. It is well established that olive oil contains various plant sterols.12 This was reflected in the olive oil-based tincture total concentration of 0.38%, which was primarily composed of beta-sitosterol (approx. 3750 µg/g). Cannflavin A was also detected, albeit at a significantly lower concentration (approx. 30 µg/g). Comparatively, the MCT oil-based tincture did not contain detectable amounts of the screened compounds. This result may reflect the formulation of this specific product, not all MCT-oil based tinctures. Similarly, capsules have various formulations due to the different types of cannabis extracts used during manufacturing. The WPE-based capsule tested here, revealed a total concentration of 0.07% and contained beta-sitosterol (approx. 680 µg/g) and cannflavin A (approx. 20 µg/g). A topical sample resulted in a total concentration of 0.15% and contained beta-sitosterol (approx. 1410 µg/g) and chrysin (approx. 40 µg/g). Different topicals likely have a range of flavonoid profiles. They are often formulated using a combination of cannabis and non-cannabis ingredients. This product, in particular, was formulated using a WPE but also contained other botanical ingredients and waxes from various natural sources. Lastly, a chocolate edible sample was analyzed and resulted in 0.13% total concentration. Cocoa naturally contains flavonoids13,14, however, only beta-sitosterol was observed in this sample (approx. 1300 µg/g). Chocolate edible formulations may contain cocoa from various sources with different chemical profiles. Additionally, cocoa contains several flavonoids and other polyphenols not screened for in this method.
Embraced by the age of biotechnology, cannabis research takes on the challenge of deconvoluting the intricate expression of the plant. This preliminary review merely scratches the surface of a long unaddressed aspect of the cannabis metabolome in the legal market. It serves primarily as an incentive to collaborate and discover new applications. For example, correlating consumer responses to more broadly quantified cannabis formulations translates into new product development. Strain development and cultivation strategies are also improved through newly accessible data. Future studies are necessary that measure expanded analyte panels while encompassing all product types, formulations, cultivation, and manufacturing methodologies. In science, finding answers to important questions generally opens the door to new possibilities but also stimulates even more questions. The cannabis industry certainly has many more questions to answer.
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