Cannabinoids, notably THC and CBD have been the focus of studies in the pharmaceutical industry (15). Additional cannabinoids have received less attention regarding their safety and pharmacokinetics, yet are becoming a new focus for the Cannabis industry (18). Cannabinoids are synthesized in the Cannabis plant from the parent molecule CBGA (19), which is converted through synthase activities to both CBDA and THCA (20). Therefore, in the Cannabis plant, THC and CBD are primarily present in their carboxylic acid forms THCA and CBDA, respectively. These acidic cannabinoids, including CBGA (the parent molecule) are thermally unstable and are decarboxylated when exposed to UV light or heat (21). Supercritical carbon dioxide processing enables low heat extraction of the native molecules CBDA and THCA (16). This low heat extraction technology has allowed higher availability of the acidic forms in products for their potential use as therapeutics in this growing market. Sales of Cannabis related products including cannabis-derived herbal extracts for pets has increased by 1,000% between September 2016 and 2017 (19). The interest in cannabinoids for companion animals has expanded to include consideration of their potential benefits including modifying behavioral problems (15), alleviating seizure frequency for idiopathic epilepsy (10), and relieving osteoarthritic pain thus improving life quality for dogs (11).

As the demand for cannabinoid containing products increases, a better understanding of their safety, dosage, and pharmacokinetic properties is needed. There is a dearth of studies regarding dosage for efficacy and safety in pets, particularly cats. Deabold et al. (16) evaluated the pharmacokinetic and safety of CBD/CBDA-rich hemp on eight healthy cats and dogs over a 12-week period, with a twice daily fixed dosage of 2 mg/kg provided either in a fish oil base or soft chew, respectively. Throughout the course of the study, all cats had normal blood count and blood chemistry profiles, except for one cat who showed a persistently elevated serum ALT concentration. The main adverse effects were excessive licking and head shaking, with occurrence rates of 35.4 and 25.2%, respectively.

In a more recent study, Kulpa et al. assessed the safety and tolerability of cannabinoids in cats with 11 escalating dosages each separated with a minimum of a 3-day period (15). Both CBD and THC were tested in 2 treatment groups with dosage escalating from 2.8 to 30.5 mg/kg and 3.8 to 41.5 mg/kg, respectively. In addition, a mixture of CBD and THC was tested in one treatment group at escalating dosages of 1.2–13 mg/kg and 0.8–8.4 mg/kg, respectively. Adverse effects were noted within 24 h following the first dosage of 2.8 mg/kg CBD and 3.8 mg/kg THC, and for each dosage thereafter. Ataxia, membrana nictitans prolapse, and blepharospasm were associated with THC alone, while emesis, hypersalivation, and lethargy were associated with CBD alone. It is noteworthy to mention however, that statistical inference in that study was carried out only following the 11th dosage of CBD. In our study, CBD and THC were tested in combination with CBDA, THCA and CBGA, with a CBD and CBDA mean dosage of ~1.37 and 1.13 mg/kg, respectively; and 25-fold less amounts of CBG, CBGA, THC and THCA. All cats had normal blood counts and serum chemistry profiles within the reference range, with no observed physical or behavioral adverse effects, despite some decrease in the blood counts over the week. This may be related to the timing of blood drawing. Blood samples were taken at 7 am in the pre study phase, and ~1 pm in the afternoon for the 1 week follow up phase. Additionally, the serum chemistry alterations observed for blood proteins and cholesterol may have also been related to timing of blood draw, which is a limitation of this study as a placebo group was not included. Interestingly, serum ALT activity did increase in the population by a mean of ~18 U/L, similarly to the observations of Deabold et al. (16), hence unlike dogs where rises in ALP are observed, cats appear to have slight rises in the ALT hepatic enzyme. Markers of kidney compromise were unchanged, yet the more sensitive renal marker s-dimethylarginine was not examined in this acute dosing study. Of note in our population was a statistically significant decrease in RBC (and hemoglobin), neutrophil, eosinophil, and monocyte counts which were still within normal ranges for all cats. It is difficult to determine whether this was due to cannabinoids or the base paste ingredients; however, a prior study of long term dosing of a fish oil based CBD/CBDA-rich hemp extract similar in nature showed no differences in these parameters, suggesting a possible paste effect (16). A recent publication in guinea pigs undergoing lipopolysaccharide stimulation showed that using 50 mg/kg body weight orally resulted in a mild suppression of neutrophil counts in brochoalveolar lavage samples in a pulmonary model (22). Therefore, based on physical examination data, our dosage of 1.37 mg/kg of CBD and 1.13 mg/kg of CBDA compared to that of Deabold et al. (1 mg/kg of CBD and 1 mg/kg of CBDA) and Kulpa et al. (2.8 mg/kg of CBD) appears to be a range of safe tolerable dosage for cats with superior absorption (15, 16). Despite the presence of adverse behavioral effects in Deabold et al. which may potentially be related to the oil base, and in Kulpa et al. using CBD isolate alone in a medium chain triglyceride (MCT) base at ~12 mg/kg, this product shows promise regarding short term dosing safety (15, 16).

Our CBD dosage was slightly higher than that of Deabold et al., and the Cmax and AUC were five times higher (282.0 ± 149.4 vs. 43 ± 9 ng/mL, and 908.5 ± 528.1 vs. 164 ± 29, respectively) (Table 1), indicating greater CBD absorption in our study (16). In contrast, the other parameters including Tmax, t $\frac{1}{2}$, MRT were similar to those of Deabold et al. (16) (2 ± 0 vs. 2 ± 0.6 h, 2.1 ± 1.1 vs. 1.5 ± 0.1 h and 3.8 ± 1.0 vs. 3.5 ± 1.4 h, respectively). However, Kulpa et al. (15) reported 24-h pharmacokinetic data only following the ninth dose consisting of 25 mg/kg CBD or 34 mg/kg THC. Of these data, only values for Tmax were reported, ranging from 2 to 4 h with an average of 3.3 h. This high Tmax value is not surprising given the high CBD concentration tested in that study.

The higher apparent CBD absorption in our study may be due to either the matrix used to dose the cannabinoids or the presence of other cannabinoids in addition to CBD. Deabold et al. suggested that CBD absorption was higher in dogs compare to cats, in part due to the different matrix used (16). In their study, the 2 mg/kg dose of an equal mix of CBD and CBDA in fish oil resulted in a 24-h Cmax of ~43 ng/mL with a 2-h half-life for cats. Providing the same hemp extraction incorporated into a soft chew resulted in a Cmax over 250 ng/mL in dogs with a similar half-life, suggesting that the matrix can have a profound effect on absorption. The much higher CBD absorption in our study may be related to the matrix used which consisted of a food-based paste, composed in part of soy oil, dextrose, and dried chicken liver. The serum CBD concentration at the 2-h time in our study was similar to that of Kulpa et al. (280 vs. 256 ng/mL, respectively), despite that CBD dosage in Kulpa et al. was 18 times greater than our study (15). A pure medium chain triglyceride or fish oil-based matrix appeared to have caused inferior absorption in both studies (see Table 3 for comparative purposes). In most studies, animals are fasted before administration of CBD (Table 4), and thus the potential effects of feeding status prior to cannabinoid administration should not be ignored. In a recent meta-analysis, a greater absorption of CBD was reported for fed compared to fasted human individuals (24) (Table 4).

Hemp cannabinoid extract absorption kinetics may be dependent on the cannabinoid mixture being presented to the subject. Greater plasma cannabinoid and metabolite levels after administration of the CBD/THC mixture were found compare to CBD, or THC alone, suggesting a potential pharmacokinetic interaction between CBD and THC known as the “entourage effect” (15). In human seizure patients, it is becoming evident that when using whole plant hemp extract doses can be significantly reduced when compared to CBD isolate alone, further supporting this concept that minor cannabinoids or terpenes from the product may be inducing a synergistic effect with CBD (25). In our study, CBD was administered along with CBDA and in the presence of small concentrations of THC, THCA and CBGA as discussed earlier, and CBD absorption was far greater than that reported by Kulpa et al. (15). In addition, Pellesi et al. found greater absorption of THCA and CBDA compared to THC and CBD in humans (24). The higher absorption of those acidic forms illustrates their promising medicinal uses, as the unheated acidic forms will eliminate the psychoactive effects of THC (26).

So far, no study has been reported on the safety and tolerability for the acidic forms of cannabinoids in cats. Herein, CBDA, along with THCA and CBGA were administrated with dosage of 1.13, 0.05, 0.03 mg/kg, respectively. The addition of small quantity of minor cannabinoids did not lead to a higher Cmax for CBD (282 ng/mL), but may have resulted in a much greater CBDA mean concentration of 1,011 ng/mL. In fact, Anderson et al. suggested that cannabis extracts mixture can provide a natural vehicle to enhance plasma CBDA concentrations due to cannabinoid-cannabinoid interactions, thus potentially improving its absorption (27). The acidic forms of cannabinoids may become an important focus in the field and deserves further consideration to fully understand their functions and effectiveness, as tissue levels may not correspond to short lived serum levels.

Although there is safety data for chronic dosing extensive cannabinoid examination during twice daily dosing was not performed (16). Pharmacokinetic software allows for a serum steady state concentration to be predicted based on the assumption that steady state is achieved after 5 half-lives of administration. This should be similar to the serum concentrations that were measured 6 h after the 13th dose of administration on a twice daily regimen. We found that the serum concentrations were ~50% of what would be predicted to be the steady state for all cannabinoids except for THC and THCA. This suggests differential hepatic cannabinoid metabolism which is in its infancy regarding companion animals. Though speculative, it appears that there may be some induction of the cytochrome p450 system leading to higher than expected biliary or urinary excretion after a week of twice daily dosing. This is further supported with the 24-h pharmacokinetics results showing a maximal serum concentration of 7-COOH-CBD at ~40 ng/mL which increased to over 150 ng/mL by 1 week, suggesting an upregulation of hepatic metabolism. More importantly, these levels even after 1 week of dosing appear to be far less than what is observed as the major metabolite in rodents and humans (28, 29). These data and one other study on hepatic microsomes from a cat liver suggest alternative metabolism via the cytochrome p450 enzymatic pathways which has been proposed to be 4–6 hydroxylation or carboxylation (17). The physiological effects of these metabolites are currently unknown and there are no commercially available standards to evaluate metabolic disposal through the cytochrome p450 system to standardize evaluation of these metabolic byproducts.

In conclusion, this is the first comprehensive evaluation of hemp-derived cannabinoids from a specific product that is rich in both CBD and CBDA. Like observed in other species, there appears to be superior absorption of the acidic forms of cannabinoids when compared to their decarboxylated counterparts. Our observation of superior absorption of the acids is also evident for CBD in this specific formulation when compared to the two other studies utilizing CBD in either fish oil or MCT base (15, 16). More importantly with this superior absorption, we did not recognize any adverse events associated with a 1-week administration of the product. Nonetheless, findings from one CBD-rich hemp product cannot be extrapolated to other products due to the strain differences in cannabinoid production. Although we observed no clinically relevant alterations in complete blood count and serum chemistry parameters, it is still prudent to evaluate hepatic enzymes when deciding to treat due to a very mild rise in serum ALT.