Twelve vape oil liquid samples were tested for constituents. Terpenes, terpenoids, flavor and fragrance agents, cannabinoids, and many other additives were detected (Tables 2 and 3). We observed that the flavor names and cannabinoids listed on their respective package did not necessarily correspond with the terpene types and cannabinoids found in samples. For example, product type Blue Dream is high in myrcene, known for its relaxing and sedative effects and Sour Diesel is high in both myrcene and limonene, a combination known for its energizing and stress-relieving effects (Erickson, 2019), but we did not find the stated terpene types in those samples. Therefore, the package descriptions may merely serve as a marketing tool to attract consumers who seek for those added benefits.

Vape oil liquid samples consisted of small amounts of terpenes from below 1–7% based on the peak areas of the total compounds found in one sample (Figure 1). The most common terpenes and natural extracts found were Caryophyllene (12 samples), Alpha-Bisabolol (11 samples), Linalool (10 samples), Alpha-Humulene (9 samples), Caryophyllene oxide (8 samples), D-Limonene (8 samples), Phytol (8 samples), Fenchol (6 samples), Nerolidol (6 samples), Selina-3,7(11)-diene (6 samples), Squalene (6 samples), Vitamin E (6 samples), Beta-Myrcene (5 samples), and Gamma-Selinene (5 samples). Among these commonly found terpenes, Caryophyllene, D-Limonene, Alpha-Humulene were found at higher percentage compared to other terpenes. These major terpenes we found are consistent with the ones described in USP from cannabis plants (Sarma et al., 2020). In general, typical Cannabis plants can have as many as 140 different terpenes (containing carbon and hydrogen) and terpenoids (containing carbon, hydrogen and oxygen) including monoterpenoids (C10), sesquiterpenoids (C15), diterpenoids (C20), and triterpenoids (C30) (Brenneisen and ElSohly, 2007). In this study we found around 60 various terpenes, terpenoids, flavor and fragrance agents in the twelve tested samples. Although most of these 60 terpenes maybe natural substances carried over through extraction process from cannabis plants, it’s possible that some of the terpenes, especially some flavor and fragrance agents such as Valencene, Menthone, Benzyl Alcohol, D-Carvone, and Triacetin were purposely added into the extracted vape oil to enhance the flavor. Adding flavors into e-liquids for nicotine vaping is a common practice despite the possible health implications (Erythropel et al., 2019) and it is an increasing trend that various terpenes, flavor and fragrance agents are being added into cannabis vape products (Erickson, 2019).

The most common cannabinoids found in this study were Delta9-Tetrahydrocannabinol (Delta9-THC) (12 samples), Cannabinol (CBN) (12 samples), Cannabicitran (CBT) (12 samples), Cannabigerol (CBG) (11 samples), Tetrahydrocannabivarin (THCV) (11 samples), Cannabichromene (CBC) (10 samples), (6aR,9R)-delta10-THC (7samples), and Cannabifuran (5 samples). Delta9-THC was the most dominant cannabinoid in most of the samples with average around 50–60% peak areas of the total compounds found in one sample and with a maximum of 73% peak area (Figure 1). CBN as an oxidation and degradation byproduct of the Delta9-THC was consistently found in all twelve samples ranging from 1 to 20% peak areas of the total compounds. This reveals that the quality of tested vape products has great variations depending on the age of the product, original packing and storage conditions. The amount of CBC, CBG, THCV, CBT and Delta10-THCs also varied from sample to sample.

In two samples (F18CTS035 and F18CTS046) obtained in September and December of 2018, respectively (one year before the EVALI outbreak), the major content was Delta9-THC with greater than 70% peak areas of the total compounds found. CBN as the second major constituent had similar levels of around 5–6% peak areas in both samples and this indicates that vape products may have longer than 12 months of shelf life, especially stored at lower temperatures. Major terpenes and other additives were at much lower levels than cannabinoids with around 7% peak area in one sample and 4% peak area in the second sample (Figure 1).

In the ten samples collected in 2019 during the EVALI outbreak period, Delta9-THC varied significantly ranging from 4.5 to 70% based on the peak areas of the total compounds found in one sample. Five of the samples contained approximately 40–50% of Delta9-THC peak areas in those samples. In the samples with lower levels of cannabinoids, three samples contained more than 30% Vitamin E Acetate (VEA) peak areas of the total compounds with small amount of Vitamin E; two samples contained more than 25% of Medium Chain Glycerides (MCTs) peak areas; three samples contained Polyethylene Glycols (PEGs) with two of them greater than 10% peak areas of the total compounds found; one sample had more than 20% CBN peak area; and one sample had more than 3.5% caryophyllene peak area (Figure 1). In these variety of samples, minor cannabinoids such as Delta8-THC, CBT, (6aR, 9R)-delta10-THC/(6aR,9S)-delta10-THC, 9(R)-delta6a,10a-THC/9(S)-delta6a,10a-THC, Hexahydrocannabinol, Cannabidivarol (CBDV), Delta8-Tetrahydrocannabivarin, and Exo-THC were detected. Usually these minor cannabinoids are seen in the degradation of Delta9-THC or as byproducts of the extraction process in very small amounts (Hudalla, 2020).

Delta10-THCs ((6aR, 9R)-delta10-THC, (6aR,9S)-delta10-THC) have no pharmacological effects and 9(R)-delta6a,10a-THC and 9(S)-delta6a,10a-THC have very low or very limited psychoactivity. These byproducts are not well studied for their efficacy and toxicity and therefore, the long-term health effects in cannabis products are unknown to consumers (Hudalla, 2020; Williams, 2020). Delta8-THC exists in small amounts in cannabis plants (<1%). The identified Delta8-THC isomers may have resulted from raw cannabis plant material from the extraction or post extraction processing using ethanol, hydrocarbons, or CO₂ to extract/purifying cannabis oil, and remove waxes and chlorophyll (Wilhelm; Hudalla, 2020). Delta8-THC has become increasingly popular and drawn attention in recent days when CBD became nationally legalized. CBD can be easily converted to Delta 8-THC with addition of catalysts (e.g., p-toluenesulfonic acid) in a solvent mixture (Barrie Webster and LeonardSarna, 2004). However, the conversion process is unpredictable in producing other byproducts such as other minor cannabinoids that are not found in natural cannabis plants. Delta 8-THC is not currently covered under the current California cannabis regulations and it has almost two thirds of the psychoactivity compared to Delta9-THC (Inverse.com, 2020). In one sample (F1912013-004), in addition to VEA, more than 30% Delta8-THC and 10% 9(S)-delta6a,10a-THC peak areas were detected, indicating likely adulteration with the synthetic form of Delta8-THC. In these ten vape cartridges, the CBN ranged from 1–20% and most of the samples had 3–5% peak areas. The sample with 20% CBN peak area was half full in an unused cartridge and had sign of drying. This was likely due to a poor sample seal or storage problem.

Vitamin E and VEA were found in six and three of the samples, respectively. Vitamin E, also known as tocopherols, is well known for its antioxidant properties. The most active form of alpha-tocopherol is commonly found in plant material, especially in plants with high oil content and hemp seed (Callaway, 2004). It is essential for plant development and help to provide the major antioxidant function for free radical damages (Muñoz and Munné-Bosch, 2019). Therefore, it is possible to have small amount of Vitamin E in cannabis plant that is coextracted and carried into the vape oil product. However, according to Brenneisen et al., Vitamin K was the only Vitamin found in cannabis plants (Brenneisen and ElSohly, 2007). Large amount of VEA were found in three cartridge samples. VEA is also called α-tocopheryl acetate and is a synthetic form of Vitamin E [EFSA, 2016]. It has a similar appearance to cannabis vape oil. It is commonly added to THC vaping liquids to dissolve/dilute or thicken them as a cutting agent to cut down the cost. Recent studies by the Centers for Disease Control and Prevention (CDC), the US Food and Drug Administration (USFDA), state investigators and research institutes have concluded that VEA is strongly associated with EVALI (Chand et al., 2019; Blount et al., 2020; Matsumoto et al., 2020; Muthumalage et al., 2020). After the EVALI outbreak, VEA is currently banned from cannabis products in many states (Boudi et al., 2019; Gibbons, 2021). In the samples containing large amounts of VEA, Vitamin E was also found and at higher amount than the other three samples with Vitamin E alone. They are probably the byproduct of VEA as VEA is sensitive to hydrolysis and breaks down to free Vitamin E and acetic acid [EFSA, 2016].

Medium-chain triglycerides (MCTs) are getting more attention in recent years for their health benefits as quick energy sources which are less likely to be stored as fat. They are also used as the supplement among athletes and bodybuilders as well as to aid weight loss (Mary Jane Brown, 2020). Triglyceride is simply the technical term for fat and has two main purposes either the body will burn it for energy or store as body fat. MCTs contain two or three fatty acids that have a chain length of 6–12 carbon atoms and they include caproic acid or hexanoic acid (C6), caprylic acid or octanoic acid (C8), capric acid or decanoic acid (C10), and lauric acid or dodecanoic acid (C12). Food sources rich for commercial extraction of MCTs include palm kernel oil and coconut oil (Mary Jane Brown, 2020). Like VEA, MCTs are added to THC e-liquid as dilute or thickening agent based on their appearance and claimed health benefits, especially in counterfeit products (Chand et al., 2019; Muthumalage et al., 2020). MCTs are generally regarded safe by the FDA as food additives under certain limitations (USFDA, 1938a). However, little is known about how they affect the respiratory tract and its local immune-inflammatory functions when used in vape products. MCTs were also found in bronchoalveolar lavage fluid in EVALI patients from CDC’s study (Blount et al., 2019; Blount et al., 2020). In the current study, we found two samples containing around 25 and 39% MCT peak areas with the major compounds as 2-(Decanoyloxy)propane-1,3-diyl dioctanoate, 2-(Octanoyloxy)propane-1,3-diyl bis(decanoate), and Glycerol Tricaprylate.

Polyethylene Glycols (PEGs) were also found in three vape cartridge samples and they contained 2–15% total peak areas including Tetraethylene Glycol, Pentaethylene Glycol, Hexaethylene Glycol, Heptaethylene glycol, Nonaethylene Glycol, Octaethylene Glycol, Decaethylene Glycol, and Undecaethylene Glycol. Propylene glycol (PG) and vegetable glycerin (VG) are two of the primary solvents heavily used in nicotine e-liquid as the thinning agents. PEGs have also been found in vape products (Traboulsi et al., 2020). PEG 400 is a low molecular weight grade of PEG that is widely used in cosmetics and pharmaceutical formulations as a solvent/lubricant due to its low oral and dermal toxicity. Even though they are safe as food additives (USFDA, 1938b), studies have shown that during vaping, PG and PEG 400 produced high levels of toxic compounds-acetaldehyde and formaldehyde when heated to 230°C. In addition, PEG 400 produced significantly higher levels of acetaldehyde and formaldehyde than PG, MCT, and VG (Troutt and DiDonato, 2017). Samples containing PEGs were also found in EVALI patients (USFDA, 2020c).

In these vape oil samples, we also found a set of fatty acids, mainly unsaturated fatty acids including linoleic acid, linolenic acid, oleic acid, and their methyl ester or ethyl ester derivatives (Table 3). These acids are commonly found in cannabis, and are especially rich in cannabis seeds (Callaway, 2004; Brenneisen and ElSohly, 2007). They have some terpene functions and offer flavors such as green, fruity, waxy, citrus, aldehydic soapy, creamy, and coconut (The Good Scents Company I). Other compounds we found were Benzyl Alcohol (1 sample) and Butylated Hydroxytoluene (1 sample) used as preservatives; and Triacetin (1 samples) reported to function as a cosmetic biocide, plasticizer, and solvent in cosmetic formulations. Finally, a small portion of the compounds in each sample could not be identified using Wiley11/NIST 2017 Mass Spectral libraries and Cayman library.