POTENTIAL VETERINARY CLINICAL APPLICATIONS

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1 TECHNICAL REPORT for USE by VETERINARIANS ONLY Robert J. Silver DVM, MS, CVA Chief Medical Officer RxVitamins for Pets HempRx Active Ingredient: Whole hemp plant essential oil: 15 mg/ml/0.5 mg/drop: 375 mg/bottle, 25 mls/bottle Inactive Ingredients: Organic hemp seed oil CLINICAL SNAPSHOT Anecdotal reports from veterinarians regarding the use of hemp and its derivatives in dogs and cats support its value for: o Anxiety Behavioral modification Social dysfunction Training o Mild-Moderate pain Arthritis Adjunctive use with other pain management tools o Neurologic support Tremors and tonic-clonic contractions Breakthrough seizures with idiopathic epilepsy Head trauma Neuropathy o Pet Hospice End of Life comfort Quality of Life management o Neoplasia Palliation of pain used concurrently with other modalities Support Quality of Life Appetite POTENTIAL VETERINARY CLINICAL APPLICATIONS (8) 1. Pain, Inflammation and Immunomodulation 2. Epilepsy a. Effective for both acute and chronic pain by centrally and peripherally modulating nocioception b. CBD affects T-cells resulting in a mild generalized immunosuppressive effect c. CBD has been found to have potential benefit for arthritis and psoriasis in humans a. CBD attentuates seizures in experimental models of epilepsy in animals RxVitamins: HempRx Technical Report January

2 3. Anxiolytic b. THCV inhibits CB1 receptor activity resulting some anticonvulsant activity a. CBD exerts benzodiazepam-independent activity, postulated to be via post-synaptic 5-HT1A receptors 4. Neuroprotection a. CBD acts as an antioxidant and as such has been suggested for Alzheimer s, Parkinson s and Huntington s diseases. 5. Anti-emesis a. CBD in animal models has been found to be effective for the control of vomiting that is unresponsive to 5-HT-3 agonists such as metoclopramide or ondansetron 6. Diabetes Mellitus a. CBD inhibits development of diabetes in experimental models of diabetes in mice. Reduction of pancreatic inflammation and antioxidant effects are credited with this benefit 7. Bone formation 8. Cancer a. Cannabinoids stimulate the stem cells responsible for fracture healing and bone formation, as well as reducing bone loss by controlling bone reabsorption a. Many of the cannabinoids have anti-apoptotic effects and reduce neoplastic proliferation in selected tumor cell lines b. Anecdotal reports from both human and veterinary patients indicate the potential for complete remission and possibly even cure of a number of different neoplastic diseases 9. Anti-microbial a. Both CBC and CBG have potent anti-bacterial effects including against MERSA (MIC of mcg/ml) CANNABIS FAQ: HEMP VS MARIJUANA Hemp essential oil contains legal, minor, non-psychotropic amounts of Δ-9- tetrahydrocannabinol (THC) <0.3% DM which makes it legal by Federal law to be used throughout the United States and Canada. The Federal Farm Act of 2013 allows each state to establish legal domestic hemp farms as commercial operations, state by state. (Currently 27 states have laws allowing the legal cultivation of hemp.) Hemp essential oil contains multiple plant molecules, many of which are unique to the hemp plant, but many of the terpenes are shared by other plants such as hops, pine, lemon, lavender and many more. Phytocannabinoids have been found in plants other than Cannabis sativa L, although Cannabis has the widest distribution and highest phytocannabinoid content of any plant. RxVitamins: HempRx Technical Report January

3 Hemp and marijuana are derived from different cultivars of the Cannabis sativa L. plant. The marijuana cultivar contains substantial amounts of THC, and is psychotropic. Marijuana can be grown legally only in the 23 states that have passed laws to allow its medicinal or recreational use. The hemp cultivar contains insubstantial amounts of THC, for which reason it is non-psychotropic, and can be grown legally in the United States. Hemp is known for its fiber and oil content which can be used to make food, clothing, cloth, paper and structural materials for building and more. The hemp plant contains over 200 different plant molecules that include flavonoids, terpenes and phytocannabinoids. The primary phytocannabinoid found in hemp differs than the phytocannabinoid found in marijuana. The dominant hemp phytocannabinoid is cannabidiol, (CBD) which is in the same family of molecules as THC. But, CBD is not psychotropic like THC, and has been found, like THC, to possess potent medicinal benefits. It has also been found that CBD can help to antidote the toxicity of THC. Marijuana cultivars have historically been low in CBD and high in THC. Because of the recognized benefits of CBD, many marijuana growers are now breeding their plants to contain much higher CBD content. Research in the 1990s identified membrane receptors for THC and determined many of the mechanisms of action of CBD, most of which are not receptor-dependent. It was found that the body produces its own endogenous cannabinoids, termed endocannabinoids, which are ligands for membrane-based cannabinoid receptors. The endocannabinoids also have non-receptor dependent mechanisms of action. The endocannabinoids, their receptors and the enzymes involved in recycling the receptorligand complexes have been termed the Endocannabinoid System (ECS) which is a newly-discovered system within the body that has very specific functions as regards the nervous system repair and maintenance, pain management, immune function and more. It has subsequently been found that the endocannabinoid system is very primitive, and has been found in animals as primitive as the sea squirt. Phytopharmacologists suggest that the biological effects of the Cannabis sativa L plant, whether it be expressed as marijuana with high THC or hemp with high CBD, are actually the result of all of the individual chemical constituents of the plant working together in concert to create a unified result. The term given to this unified effect of multiple plant constituents is the: Entourage Effect. (7) This group effect is present not just in the cannabis plant, but is the same mechanism of action whereby all herbal medicines work. When an individual constituent of a plant is isolated and made into a drug, not uncommonly, adverse side effects will arise which are different than when the whole plant medicine is used. ADMINISTRATION Appropriate dosages to give for specific conditions to a variety of veterinary species as determined through pharmacological research are still pending. Funding agencies are nervous about providing funding for cannabis research due to the 70 year federal prohibition of Cannabis sativa L., whether it be marijuana or hemp. Empirical Determination: RxVitamins: HempRx Technical Report January

4 This author established empirical doses from multiple sources as there are no doseresponse studies to refer to in the veterinary literature. 1. One method to estimate effective dosages utilized cannabinoid analysis of the two leading brands of legal hemp-based commercial products available to pet owners in capsules over the internet. We conducted third party laboratory analysis of the content of both products to determine the amount of cannabidiol being recommended for a given weight. This author then reverse-engineered a numerical mg/kg CBD dosage from this. Anecdotal reports of pet-owners using these products with observed clinical benefits for a number of different conditions were used to cross-check the reverse-engineered dosages to arrive at a firm empirical dose for these products. Interestingly, the dosage which has been used successfully by these two products was quite low when compared to published dosages for laboratory subjects. 2. Other sources of dosing information include the clinical reports from a Board certified veterinary oncologist and other veterinarians who were given trial bottles of HempRx, donated by RxVitamins, so as to determine the most effective dosing range for most patients. 3. This author also used reports from veterinarians regarding the effective doses that they used with other commercially-available consumer products such as biscuits and other pet-edibles. By matching up the weight of the animal that had demonstrated response to a certain dosage of biscuit or liquid as expressed in milligrams of cannabidiol (CBD), a dosage estimate was derived. Multiple determinations were used to create an average effective dosing range for hemp extracts. 4. Over time, once concentration-standardized hemp products, such as HempRx are used more clinically by veterinarians, a more precise guide to veterinary administration will be developed. As funding becomes available, veterinary pharmacological research is absolutely essential for the continued safe and effective use of hemp-based extracts. There is one pharmacokinetic study in dogs, published in 1988 that contains information regarding the kinetics of cannabidiol in the dog. (11). 5. The range of effective amounts to give of hemp extract, based on its cannabidiol content, as a result of the first three methodologies described above is: 0.1 mg/kg BID 0.5 mg/kg BID NOTE: It is recommended to start at the low end of the dosage scale, which is comparable to the dosages recommended for commercially-available hemp capsules sold over the internet by the two large Canna companies. Give the dosage 1-2 weeks for full effect. If the desired effect is not seen after two weeks, then it is recommended to increase the dosage to: 0.25 mg/kg BID. RxVitamins: HempRx Technical Report January

5 If, after two weeks that dose is still ineffective for the condition you are using it for, then go to the high end of the dosage range, 0.5 mg/kg BID. Doses as high as 1-2 mg/kg BID may be needed for more severe conditions. The only side-effect reported in a few patients is some mild sedation. Sedation is not consistently achieved in all patients for a given dose, but seems to be more dependent upon the individual s constitution. NOTE: At this point in time (early 2016), the cost of good-quality organicallygrown hemp extract is quite high. Economically, it makes more sense to start dosing at a lower, less consumptive level, and then increment it up if the lower dosing scale isn t working. With the commercial capsules, many dogs respond well to the label dosages, but the companies encourage feedback from their petowning customers because they know that individual response to hemp-dosing can be quite variable, and sometimes you need a higher dose to effect change. LEGAL CONSIDERATIONS Cannabis sativa L. has been prohibited from being grown or sold in the United States for over 70 years. Shortly following the end of the prohibition of alcohol in the US in the 1930s, cannabis became prohibited, illegal, and stigmatized. The psychotropic nature of the higher phytocannabinoid content of THC in marijuana is the reason for this prohibition. Up until that time, cannabis could be found in many Pharmacopoeia in the United States and was included in a number of medical remedies prior to the 1930 s era that introduced prohibition, including veterinary remedies, primarily in horses and livestock. Hemp is not Marijuana Cannabis sativa L. has two main cultivars, the high THC, low fiber marijuana, and the low THC, high fiber hemp. The plants look very similar, and the seeds of both plants look identical. The similarity in external appearances between these two related plants led to the prohibition of the growing and commercialization of hemp in the United States, in spite of how economically valuable hemp is as a crop. There are some conspiracy thinkers who suggest that hemp was made illegal so as to allow a few oligarchs with large lumber holdings to profit. Hemp is a renewable resource that can be made into building materials, clothing and paper (as well as being a source of protein and healthy omega three oils), thus competing with the lumber industry, which is not sustainable, and is poorly renewable as a resource. It takes many years for a forest to grow. You can have several hemp harvests annually. Commercial hemp production was a major reason for the colonization of the New World by the maritime European nations during that time period. Hemp was used for sail cloth, and cordage for the tall-masted ships, as well as for clothing and paper. The uniforms of the Revolutionary war soldiers were made from hemp. Betsy Ross used hemp to sew the first American flag. Our founding fathers, George Washington, Thomas Jefferson, Thomas Paine, Benjamin Franklin, all produced or used hemp in their work. RxVitamins: HempRx Technical Report January

6 With the passage of the Farm Act of 2013 in 2014, hemp began to be grown again in the United States. Prior to this, all of the hemp used in the US has been imported from Canada, Europe and China. Since the Farm Act of 2013 passage there are, as of Winter 2015, 27 states that have passed laws allowing the commercial cultivation of hemp under the regulation and jurisdiction of each state s Department of Agriculture. The number of acres of hemp produced by American farmers grew by 1000% in 2015 over Experts predict that hemp production has the potential to become a significant American industry, benefiting the agricultural segment as well as food, paper, cloth, building materials, pharmaceutical and nutraceutical industries. Paper made from hemp is acid free, unlike paper made from wood, and unlike wood-based paper will last for centuries. Hemp seed oil is a healthy source of omega 3 fatty acids, and hemp seed cake contains high quality highly digestible protein. Currently it s estimated that total hemp sales in the United States are around $452 Million annually. Hemp grows organically and is generally resistant to common pests and diseases. It requires little fertilizer or pesticide for production. Hemp doesn t deplete the soil, and in fact can be used for phyto-remediation due to the fact that its deep roots pull toxic substances such as heavy metals, pesticides, solvents, crude oil and even radiation from the soil. As an example, it was used to decontaminate the Chernobyl nuclear disaster site. Hemp captures carbon, rebuilds the soil, and is drought-tolerant requiring 1/3 the water of corn. The hemp cultivar from which the HempRx essential oil is derived is grown organically under the sanction and supervision of the Colorado Department of Agriculture to ensure that the plants contain less than the legal limit of 0.3% DM THC. This specific cultivar has been bred over 10 years to produce a high content of the terpene linalool, which is also found in large amounts in the lavender plant (Lavandula angustifolia). The hemp cultivar that is the source of the essential oil in HempRx is propagated from one crop to the next by means of cloning, versus allowing them to go to seed. This decreases the genetic variation from crop to crop which helps to ensure a consistent supply of hemp extract with little variation from crop to crop in the phytochemical constituents of its essential oil extract. The mature plants are tested by the Colorado Department of Agriculture and any plants containing higher than 0.3% DM THC are destroyed. The plant material is then dried and run through a Super Critical CO 2 extraction process. Liquid carbon dioxide, under pressure, efficiently extracts all of the fat-soluble molecules in the hemp plant, to form a waxy solid. At normal pressures and temperatures the liquid CO 2 evaporates without any residue remaining, leaving only the desired plant molecules: Flavonoids, terpenes and phytocannabinoids. RxVitamins: HempRx Technical Report January

7 THE ENDOCANNABINOID SYSTEM (ECS) Following the determination of the structure of the first cannabinoid Δ-9- tetrahydrocannabinol (THC) in 1964, researchers started looking for the membrane receptors that could mediate the activity of the cannabinoids. In 1988, the first cannabinoid receptor was described in the rat brain using a radioactive-labeled THC derivative. This receptor, termed Cannabinoid Receptor 1 (CB1), was determined to be a G-protein coupled receptor with the highest density in the rat cerebral cortex, hippocampus, hypothalamus, cerebellum, basal ganglia, brain stem, spinal cord and amygdala. This receptor is present in all vertebrate species, indicating that that the endocannabinoid system has been in existence for over 500 million years. Primitive cannabinoid receptors have been described in animals as evolutionarily primitive as the Sea Squirt (Ascidiacea) which is a marine invertebrate filter feeder. The endocannabinoid system (ECS) consists of 1) the cannabinoid ligand, which binds to the cannabinoid receptor, 2) the receptor itself, and 3) the enzymes that synthesize and degrade the ligands. The CB1 Receptor The CB1 receptor is found in its highest concentrations on neurons that release gamma amino butyric acid (GABA), the main inhibitory neurotransmitter. It is located near the synapse. The discovery of this endocannabinoid receptor was a water-shed moment in neurophysiology in that it led to the discovery of the body s own endogenous cannabinoid molecules (endocannabinoids) and the previously undiscovered endocannabinoid system. Mechoulam, who discovered THC, also discovered the first endocannabinoid, which he called Anandamide after the Sanskrit word for bliss. Anandamide binds to the CB1 receptor and creates similar effects as the phytocannabinoids that are naturally-occurring in the cannabis plant. A second endocannabinoid was subsequently discovered, 2- arachidonoyl glycerol (2-AG). There are several other compounds currently under investigation as potential endocannabinoids. The endocannabinoid receptors evolved along with the endocannabinoids to constitute a naturally-occurring cellular communication system, which is the endocannabinoid system. It is sheer coincidence that the phytocannabinoids found in the cannabis plant resemble the endocannabinoids enough to activate the cannabinoid receptors. The cannabinoid receptor CB1 is the most abundant G protein-coupled receptor expressed in the brain, with particularly dense expression in (rank order): the substantia nigra, globus pallidus, hippocampus, cerebral cortex, putamen, caudate, cerebellum and amygdala. This distribution has been determined for the human brain. Detailed studies in the dog using PCR technology are needed, but none have yet been conducted, as of this writing. (4, 5) RxVitamins: HempRx Technical Report January

8 The endocannabinoid system s major homeostatic functions were summarized by DiMarzo as: Relax, Eat, Sleep, Forget and Protect. The Endocannabinoid system has an effect on embryological development, neural plasticity, neuroprotection, immunity, inflammation, apoptosis, carcinogenesis, pain, emotional memory, hunger, feeding and metabolism. (10) The endogenous agonists for cannabinoid receptors are long-chain polyunsaturated fatty acids (eicosanoids) that are derivatives of arachidonic acid, and have varying degrees of selectivity for either one or both of the cannabinoid receptors. Endocannabinoids are unlike other neurotransmitters in that they are lipid versus aqueous in nature. They also are not stored, but are manufactured ad hoc from the cellular membrane. Endocannabinoids are released as calcium levels increase inside the neuron or when G- coupled protein receptors are activated. Endocannabinoids function as neuroprotectants by virtue of their antioxidant activity and by inhibiting calcium influx and excessive glutamate production. There are both cannabinoid receptor-dependent and cannabinoid receptor-independent actions of endocannabinoids. Activities that are cannabinoid receptor-dependent include cognition, memory, appetite control, emesis, motor behavior, sensory, anxiety, and autonomic and neuroendocrine processes. Endocannabinoids induce hypotension and bradycardia, inhibit cell growth, affect energy metabolism and modulate immune responses, as well as being involved in fat accumulation, glucose and lipid metabolism. Endocannabinoids can also exert proinflammatory actions such as enhancing the cellular migration of eosinophils, neutrophils and natural killer T cells (3) Endocannabinoids use a previously undiscovered form of neuronal communication: Retrograde signaling, which is the opposite to the normal direction of neurotransmitter release from presynaptic neuron to reception on the postsynaptic neuron. Endocannabinoids released from the postsynaptic neuron actually bind at CB1 receptors on the presynaptic GABA neurons to modulate neuronal activity. This novel discovery of retrograde signaling was termed: Depolarization-Induced Suppression of Inhibition or DSI. DSI helps to explain a number of previously unexplained aspects of brain activity. When you temporarily dampen inhibition, a form of learning called long-term potentiation occurs, which is a process by which information is stored through the strengthening of synapses. It was also found that CB1 receptors can, in some cases, block presynaptic cells from releasing excitatory neurotransmitters. This is true in the cerebellum where endocannabinoids located on excitatory synapses help to regulate neurons involved with motor and proprioceptive control of movement. This helps to explain, in part, the static ataxia uniquely observed in dogs only. The canine species have the highest density of CB1 receptors in the cerebellum of any other species studied to date. RxVitamins: HempRx Technical Report January

9 Cannabis Research in Dogs Research performed in the 1970 s by the Department of Defense, explored whether marijuana could be weaponized. Dogs were administered radioactive-labeled THC intravenously at escalating dosages. As a result, researchers found that dogs, as compared to pigeons, monkeys, guinea pigs, rats and mice, had the highest concentration of THC (now known to be bound to CB1 receptors) in the cerebellum. The canine molecular layer was found to be more dense than the molecular layer in any of the other species studied. The hippocampal formation was also very dense in specific locations. (4) Previous work had found that the minimum dose of THC administered IV to create static ataxia was 0.5 mg/kg IV.(5) Tolerance to the behavioral effects of THC in the dog developed after daily injections were given. McMillan found that a dose of 2 mg/kg IV produced marked static ataxia, evidenced by swaying movements, hypersensitivity to moving objects and a prance-like foot placement. However, some dogs in this study group developed tolerance rapidly after the first administration of 2 mg/kg of THC. Subsequent injections continued to increase the degree of tolerance to THC in this study group. The magnitude of tolerance developed in these canine studies was in excess of 100 fold. (9) CB1 receptors are found primarily in the central nervous system, but also have been found in the GI tract (perhaps explaining why we see appetite stimulation with cannabis administration), cardiovascular system and reproductive system. In the dog, localization of the CB1 receptors was found in the hippocampus, structures of the skin including mast cells, hair follicles and salivary glands. (6) CB2 Receptors A second, G-protein coupled receptor for cannabinoids is the CB2 receptor. These receptors have been found to be strongly expressed in cells of the immune system, including the microglia, the peripheral nervous system and the organs. CB2 immunoreactivity was found in the B cell zones of lymphoid follicles in the dog, as well as in structures of the skin including mast cells, and hair follicles. (6) CB2 receptors are up-regulated during the early phases of inflammation in cells of the CNS and peripheral tissues, suggesting a role for cannabinoids in the management of inflammatory conditions of those tissues. Non-CB Receptor-Dependent Activity In addition to the receptor-dependent mechanism of action of the cannabinoids, terpenes and terpenoids, their activity can also be mediated through non-receptor dependent interactions. The endocannabinoids exert multiple pharmacological effects through a number of different mechanisms not restricted to modulation of the endocannabinoid system through receptor-ligand binding. A partial list of these non-receptor dependent actions include: (1) Transient receptor potential (TRP) channel activation Peroxisome proliferator-activated receptor λ (PPAR λ) GPR55 Abnormal-CBD receptor 5-hydroxytryptamine receptor subtype 1A (5-HT1A) Glycine α1 and α1β receptors Adenosine membrane transporter phospholipase A1 RxVitamins: HempRx Technical Report January

10 Lipoxygenase (LOX) and cyclooxygenase-2 (COX-2) enzymes Calcium modulation Inhibition of anandamide inactivation by CBD, CBG and CBC Terpenes and terpenoids exert strong biological effects by themselves, and have been found to interact synergistically with phytocannabinoids in the treatment of pain, inflammation, depression, anxiety, addiction, epilepsy, cancer, fungal and bacterial infections (including MERSA) (7) Phytocannabinoids and Liver Metabolism There is no data for veterinary species regarding the effect of the phytocannabinoids on liver enzyme metabolism. Typically the impact of a botanical compound on liver metabolism is what determines herb-drug interactions. There are a few published studies of the interaction between human liver enzyme systems and the phytocannabinoids, mostly derived from in vitro data. The in vitro data support that cytochrome P-450 (CYP-450) enzymes are important contributors to the first-pass metabolism of the following phytocannabinoids: THC, CBD, and CBN. CYP-450 enzymes also contribute to the second pass metabolism of these three exogenous cannabinoids. The authors of this study go on to state in their conclusion: Studies of THC, CBD and CBN inhibition and induction of major human CYP- 450 isoforms generally reflect a low risk of clinically-significant drug interactions with most use, but specific human data are lacking. (12) BIOMEDICAL ACTIVITY of PHYTOCANNABINOIDS & TERPENES There are many plant constituents in cannabis of medicinal interest. Of most interest are the phytocannabinoids, which consist of more than 100 terpenophilic compounds, found mainly in cannabis, but recently have been described in several other plants in the family Linaceae (Flax), and Asteraceae (Echinaceae and Helichrysum). Other phytoconstituents such as terpenes, terpenoids, and flavonoids also contribute to the medicinal profile of cannabis. Cannabinoids exist in the plant mainly as carboxylic acids, which are called cannabinoid acids and are all non-psychotropic. The acidic form is converted to neutral molecular analogs by light, heat and combustion. (2) The phytocannabinoid that has gotten the most attention in this plant is Δ-9 THC, which has both psychotropic and medicinal qualities, and, subsequently, has resulted in its value, notoriety and illegality. However, the other phytocannabinoids, which are divided into multiple classes based on chemical structure, are not psychotropic, but contain the majority of the medicinal properties of this plant. TABLE ONE: Major and Minor Cannabinoids (2) Δ-9-tetrahydrocannabinol (Δ-9 THC) MAJOR CANNABINOID Analgesic (reduces pain), anti-inflammatory, antioxidant, brochodilatory, improves symptoms of Alzheimer s disease, benefit duodenal ulcers, muscle relaxant, anti-itch, cholestatic jaundice. Δ-9 Tetrahydrocannabinolic acid THCA is the acidic or carboxylated form of THC. It RxVitamins: HempRx Technical Report January

11 (Δ-9 THCA) Δ-9 Tetrahydrocannabivarin (Δ-9 THCV) Δ-8 Tetrahydrocannabinol (Δ-8 THC) Δ-8 Tetrahydrocannabinolic acid (Δ-8 THCA) Cannabidiol (CBD) MAJOR CANNABINOID Cannabidiolic acid (CBDA) Cannabichromene (CBC) Cannabigerol (CBG) Cannabidivarin (CBDV) Cannabinol (CBN) is the predominant cannabinoid in psychoactive strains. It is non-psychotropic until activated or decarboxylated, smoked or cooked at temperatures greater than 245 F. Also has medicinal benefits, similar but also separate and different than THC. Anti-inflammatory, anti-convulsant, analgesic properties, anti-oxidant, neuroprotective in model of Parkinson s in one study, improved glucose tolerance and insulin sensitivity in vivo Stable in air, much less psychotropic than Δ-9 THC; At low doses, Δ-8 THC (0.001 mg/kg PO was found to induce appetite stimulation without psychotropic effects. The carboxylated (acidic) form of Δ-8 THC Anti-anxiety, anticonvulsant, Parkinson s disease, Huntington s disease, psychosis, MS, Alzheimer s, cytotoxic for breast cancer, effective against MRSA, reduces oily skin, treatment of addiction Acidic form of CBD, carboxylated form of CBD. Has medicinal properties but not well studied at this point in time. Anti-inflammatory, analgesic, antifungal, antidepressant, Anandamide reuptake inhibitor Anti-fungal, GABA uptake inhibitor (calming), antidepressant, analgesic, anti-inflammatory, reduces scales in psoriasis, effective against MRSA Anti-convulsant Sedative, effective versus MERSA, helps with burns, reduces scales in psoriasis, helps with breast cancer. May be a degradation product of THC or CBD. Other, equally important phytoconstituents of cannabis are the terpenes and terpenoids. These organic compounds are produced by a variety of plants. It is thought they serve a protective function for these plants. They are a significant component in plant essential oils. These molecules are responsible for the aroma of cannabis, and because they, like cannabinoids, are lipophilic, they also cross the blood-brain barrier and contribute to the medicinal benefits of cannabis. RxVitamins: HempRx Technical Report January

12 The US FDA considers terpenes and terpenoids to be Generally Recognized as Safe (GRAS), as they are flavor and fragrance components common to human and pet diets. Cannabinoids, terpenes and terpenoids are all produced in the same glandular structure on the cannabis plant, the trichome, from the same chemical precursor, geranyl pyrophosphate. Hops (Humulus lupus) is a member of the same Cannabaceae Family as cannabis, and they share many of the same terpenes and terpenoids such as β-myrcene, β-pinene, humulone, and β-caryophyllene. Cannabinoids are virtually odorless, emitting only a slight pitch-pine scent. The biological effects of cannabis are due to interactions among the many various phytoconstituents of cannabinoids, terpenes and terpenoids. This phytochemical interaction has been termed the Entourage Effect, and is believed to explain the multiple biological activities of the cannabis plant, and the differences that are seen in bioactivity of the different strains of the cannabis plant. The Entourage Effect states that the potency of the whole plant extract is the sum total of the interaction of all of the plant constituents involved, and is different than the effect of any individual plant component alone. Strains are subsets of the cannabis sativa L. genome, which contain differing distributions of fiber, phytocannabinoids, terpenes and terpenoids. Strains are much like breeds of dogs. All are Canis familiaris, but there are definite differences between a Chihuahua and a Saint Bernard, in spite of the similarity of 99% of their shared genome. HEMP Rx ESSENTIAL OIL ANALYSIS TEST ID: W41 JL DATE: 1/8/2016 SAMPLE: Order (375 mg CBD in each Bottle) TYPE: CBD-RICH HEMP OIL Tincture POTENCY RESULTS (Per 25 ml bottle) Wt. (%) (mg/g) CBD CBG CBN <0.01 <0.1 THC CBC THC-A <0.01 <0.1 CBD-A MAX THC MAX CBD TOTAL ACTIVE THC:CBD 1.25 VISUAL INSPECTION Clear, light green tincture Tested By: Jenna Lubieniecki Reviewed By: Raj Gupta, Ph.D., Lab Director RxVitamins: HempRx Technical Report January

13 All cannabinoids in their acid forms, those ending in '-A', can be converted to their non-acid forms through a process called decarboxylation when the sample is heated. The molecules lose mass through this process, and thus to find the total theoretical active cannabinoids you must multiply the acid forms by 87.7%. For example, THC-A can be converted to active THC using the formula: THC-A x = THC. And so the Max THC for the sample is: Max THC = (THC-A x 0.877) + THC. This method has been validated according to the principles of the International Conference on Harmonisation. TERPENE RESULTS Wt. (%) Wt. (%) d-limonene Myrcene Eucalyptol α-bisabolol α-humulene Phytol α-pinene Linalool β-caryophyllene Nerdilol RESIDUAL SOLVENTS RESULTS ppm ppm Propane Not Detected Pentane Not Detected Isobutane Not Detected Isopropanol Not Detected Butane Not Detected Hexane Not Detected Ethanol Not Detected* Acetone Not Detected * Limit = 5000 ppm PESTICIDES RESULTS ppm ppm Acequinocyl Not Detected Spinosad Not Detected Pyrethrium Not Detected Spirotetramat Not Detected Spiromesifen Not Detected Bifenazate Not Detected Abamectin Not Detected Fenoxycarb Not Detected N/A Imidacloprid N/A Not Detected Paclobutrazol Not Detected None Detected (Criteria < 10 ppm) HEAVY METAL RESULTS REFERENCES 1. Izzo AA, Borelli F, Capasso, R, Di Marzo V, Mechoulam R. Non-psychotropic plant cannabinoids: new therapeutic opportunities from an ancient herb. Trends in Pharmacological Sciences;2009;30(10): American Herbal Pharmacopoeia: Cannabis Inflorescence: Standards of Identity, Analysis, and Quality of Control (2013); 3. Svizenska I, Dubovy P, Sulcova A. Cannabinoid receptors 1 and 2 (CB1 and CB2), their distribution, ligands and functional involvement in nervous system structures: A short review. Pham Biochem Beha 2008; 90: Herkenham M, Lynn AB, Little MD, Johnson MR, Melvin LS, de Costa BR, Rice KC. Cannabinoid receptor localization in the brain. Proc Natl Acad Sci. 1990;87: RxVitamins: HempRx Technical Report January

14 5. Martin BR, Dewey WL, Harris LS, Beckner JS. 3 H Δ 9 Tetrahydrocannabinol tissue and subcellular distribution in the central nervous system and tissue distribution in peripheral organs of tolerant dogs. J Pharm Exper Ther.1976;196(1): Campora L, Miragliotta V, Ricci E, Cristino L, DiMarzo V, Albanese F, della Valle MF, Abramo F. Cannabinoid receptor type 1 and 2 expression in the skin of healthy dogs and dogs with atopic dermatitis. Am J Vet Res 2012;73: Russo EB. Taming THC: potential cannabis synergy and phytocannabinoidterpenoid entourage effects. Brit J Pharmacol 2011;163: Boothe, DM. The High Points of Medical Marijuana. Western States Veterinary Conference Proceedings, February McMillan DE, Dewey WL and Harris LS. Characteristics of Tetrahydrocannabinol tolerance. Annals NY Acad Sci. (1971)191: McParland JM, Guy GW and Di Marzo V. Care and Feeding of the Endocannabinoid System: A Systematic Review of Potential Clinical Interventions that Upregulate the Endocannabinoid System.(2014)PLoS ONE (9)3:e doi: /journal.pone Samara E, Bialer M, Mechoulam, R. Pharmacokinetics of Cannabidiol in Dogs. (1988) Drug Metabolism and Disposition. 1988;16(3) Stout S, Cimino NM. Exogenous cannabinoids as substrates, inhibitors, and inducers of human drug metabolizing enzymes: a systematic review. Drug Metabolism Reviews, 2014;46(1): RxVitamins: HempRx Technical Report January