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Nonterpenoid Chemical Diversity of Cannabis Phenotypes Predicts Differentiated Aroma Characteristics

Abstract


The recent increase in legality of Cannabis Sativa L. has led to interest in developing new varieties with unique aromatic or effect-driven traits. Selectively breeding plants for the genetic stability and consistency of their secondary metabolite profiles is one application of phenotyping. While this horticultural process is used extensively in the cannabis industry, few studies exist examining the chemical data that may differentiate phenotypes aromatically.


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To gain insight into the diversity of secondary metabolite profiles between progeny, we analyzed five ice water hash rosin extracts created from five different phenotypes of the same crossing using comprehensive 2-dimensional gas chromatography coupled to time-of-flight mass spectrometry, flame ionization detection, and sulfur chemiluminescence detection.


These results were then correlated to results from a human sensory panel, which revealed specific low-concentration compounds that strongly influence sensory perception. We found aroma differences between certain phenotypes that are driven by key minor, nonterpenoid compounds, including the newly reported 3-mercaptohexyl hexanoate. We further report the identification of octanoic and decanoic acids, which are implicated in the production of cheese-like aromas in cannabis.


These results establish that even genetically similar phenotypes can possess diverse and distinct aromas arising not from the dominant terpenes, but rather from key minor volatile compounds. Moreover, our study underscores the value of detailed chemical analyses in enhancing cannabis selective breeding practices, offering insights into the chemical basis of aroma and sensory differences.


Introduction


The cultivation of Cannabis sativa L. has rapidly evolved over the past few decades, resulting in varieties with highly diverse morphological, chemical, and sensory characteristics.

(1−7)


Apart from the medicinal use of cannabis and the need for pharmaceutical-grade cultivars (i.e., genetic stability and reproducibility), there is a desire to influence cannabis offspring to produce unique secondary metabolite profiles.

(8,9)


Cannabis’ vast genetic diversity enables a spectrum of aromas ranging from sweet, savory, or prototypical.

(4,8,10−13)


In particular, many cultivators aim to produce varieties that express unique aromatic and flavor characteristics that can significantly impact consumer preferences. (4)


Volatile organic compounds (VOCs) make up part of cannabis’s secondary metabolite profile and are responsible for the aroma it produces.

(7,14)


These compounds are found in different plants, fruits, and vegetables. (15−20) In cannabis, the aromatic compounds discussed often include monoterpenes, sesquiterpenes, and their respective terpenoids. However, recent studies show that low-concentration compounds such as volatile sulfur compounds and recently discovered flavorants (nonterpenoid-derived VOCs) are the primary source of the unique and diverse aromas produced by cannabis. (10,21)


While these compounds are known in the context of aroma identification, they have rarely been studied in the framework of selective breeding. For instance, minimal studies exist comparing the secondary metabolite profiles of cannabis progeny. (22) This chemical information could potentially shed light on the chemodiversity of siblings and how they inherit the aroma characteristics of their parents, as well as yield information about phenotypic distribution. (23)


Selective breeding can be utilized in a variety of different ways depending on the application (e.g., hash oil, aroma distillation, and fiber production). (8,24,25) One of the key techniques used when selectively breeding cannabis is colloquially known as “phenohunting”─the process of propagating two varietals with desirable traits, as shown in Figure 1.


Breeders will often produce hundreds of different progeny from a single cross, followed by conducting analysis on their morphological, growth, and secondary metabolite characteristics, specific to their final application. (3,23) After selection of the phenotypes that best exhibit the desired traits, these chosen phenotypes are further propagated through cloning or self-pollination to ensure genetic stability and consistency in their lineage.

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