A closer look at some of the current in vitro evidence suggesting that cannabinoids possess potent activity against a range of gram-positive bacteria.

The Role of Cannabinoids in the Global Fight Against Antibiotic Resistance

A significant contribution to our understanding of how cannabidiol (CBD) exerts its clinical efficacy has recently been published online by researchers at the University of Colorado (1). The evidence for how CBD affects more than 20 membrane targets had potentially implicated that the influence of CBD on membrane fluidity was responsible for its pharmacological effects, but the actual mechanism had not been demonstrated until now. The key finding from the multi-omic systems level analysis in human cell lines is that CBD partitions into cellular membranes, which disrupts cholesterol homeostasis along with membrane spanning ion channels and receptors, triggering a myriad of downstream consequences. CBD affected cytosolic calcium, cholesterol transport and storage, cellular energy sensors, and oxidative stress responses in a dose-dependent manner (1). In addition, CBD-exposed cells were sensitive to extracellular cholesterol, which induced cell death, or apoptosis, in a dose dependent manner.

Cholesterol interacts with the fatty acid tails of phospholipids to moderate the biophysical properties of the plasma membrane, reducing fluidity and making the membrane less permeable to small water-soluble molecules that would otherwise freely cross. While cholesterol’s bad press has led to the widespread prescription of statins that block synthesis of cholesterol, it is essential for both membrane permeability and also in metabolism as the starting point for the synthesis of bile salts and steroid hormones including cortisol, estrogen, and testosterone. Cholesterol makes up around 25–30% of the lipid content of the cell membrane and it is the dominant lipid in the brain. Cells either synthesize cholesterol from acetyl coenzyme A (acetyl-CoA) or take it up from the food we eat. While cholesterol deficiency results in several diseases, we are just beginning to investigate and appreciate the regulatory role that cholesterol plays in its relationship to cellular homeostasis, inflammation, and pain much like how we think of CBD today.

Of the proposed list of CBD targets, 22 are membrane-localized channels and receptors. Mutations in one voltage-dependent sodium channel, NaV1.1, are responsible for epilepsy syndromes including Dravet syndrome for which Epidiolex is prescribed (2). Many other proposed targets are calcium channels or receptors, in fact postsynaptic calcium mobilization is the probable mechanism for the anticonvulsant activity of CBD (3). The recent study brings strong evidence to a previously proposed theory that this interaction of CBD with membrane-localized channels is indirect through biophysical alteration of the lipid bilayer with cholesterol as the effector molecule (4,5).

Growing evidence suggests that pain sensation caused by inflammation is in part regulated by the pro- or anti-nociceptive actions of lipid mediators which include endogenous cannabinoids as well as agonists of peroxisome proliferator-activated receptor-α (PPAR-α), platelet activating factor (PAF), and various products of oxidative polyunsaturated fatty acids (PUFAs) metabolism (6). An increasing number of lipid molecules have been shown to suppress the inflammatory process, restore homeostasis in damaged tissues, and attenuate pain sensitivity by regulating neural pathways that transmit nociceptive signals from the periphery of the body to the central nervous system (7). The endocannabinoids, anandamide and 2-AG, are synthesized locally on-demand to mitigate the effects of exogenous and endogenous pro-analgesic agents by attenuating nociceptor excitability and contrasting local pro-inflammatory signals from another group of bioactive lipids, including membrane cholesterol (7).

Let’s take a closer look at how loss of cholesterol homeostasis mitigates a wide range of physiological change. Normally, cholesterol levels are tightly regulated between the plasma membrane (PM) sensors and the endoplasmic reticulum (ER). When cholesterol is depleted, low density lipoprotein (LDL) molecules transporting cholesterol bind to receptors and enter the cell through endocytosis to reach lysosomes where the cholesterol is released from the LDL and delivered to the PM and ER membranes. If there is excess cholesterol, it is removed from the PM and delivered to the ER where it is esterified for storage in lipid rafts and not available for normal cellular processes (8). When this normal feedback inhibition of cholesterol levels is disrupted there is subsequent activation or deactivation of raft-associated proteins, such as death receptor proteins, protein kinases, and calcium channels. Furthermore, increased production of cholesteryl esters, has been reported across human tumors compared with normal tissue.

Changes in cholesterol metabolism play a role in carcinogenesis. It is well established that cancer cells have a much higher concentration of cholesterol and the levels of cholesterol determine their resistance to chemotherapeutic drugs (9). This increased capacity to esterify and accumulate cholesterol in tumor cells has been associated with a higher growth rate, suggesting a link between cholesteryl ester production and cell proliferation (10,11). There does not appear to be a correlation with the levels of cholesterol consumed, but rather de novo synthesis from dysregulation of the cholesterol pathway. Several anti-cancer drugs are able to suppress growth and induce apoptosis of tumor cells through alteration of cholesterol containing lipid raft contents via disrupting lipid raft integrity and thereby sensitizing cancer cells to therapeutics and also influencing cancer drug resistance (12). Similar findings have been reported for cancer cells treated with CBD, essentially robbing cancerous cells of their energy source, cholesterol.

In earlier reporting, primary microglial cells treated with CBD resulted in a concentration dependent induction of apoptosis that was not counteracted by CB1 or CB2 receptor antagonists but was attenuated by a lipid raft disruptor. This finding suggests that CBD induces a pro-apoptotic effect in primary microglia through lipid raft coalescence and elevated expression of GM1 ganglioside, involved in neuron repair, and caveolin-1, a structural component of lipid rafts (13). In the case of unregulated cholesterol accumulation in cancer cells, CBD is eliciting the same apoptotic outcome as anti-cancer drugs without the toxic side effects. Likewise, statins have proven therapeutic in some cancers—including gastric, hepatic, prostatic, and esophageal—through their inhibition of cholesterol synthesis (14).

CBD treatment was associated with high levels of Ca

 ions can damage cells through disruption of neural circuitry through excitotoxicity which can occur in neurodegenerative diseases or brain trauma such as traumatic brain injury (TBI) or stroke even causing cell death. Cellular calcium homeostasis is necessary for stress responses in metabolic organs including the heart, liver, and adipose tissue. Multiple lines of evidence demonstrate that inappropriate consumption of nutrients and chronic metabolic diseases leads to dysregulation of cytosolic and organelle Ca

 fluxes and disrupted equilibrium in metabolic tissues and immune cells, which in turn alters organelle homeostasis, signaling pathways and autophagy (15). Cellular Ca

 homeostasis fails to maintain its adaptive integrity in the face of structural and signaling defects introduced by chronic metabolic challenges whether through over consumption of nutrients or through treatment with CBD.

The effects rendered by CBD treatment cannot escape the interrelatedness of all processes in the human body. In a recent study by Olivas-Aguierre and colleagues (2019) the treatment of acute lymphoblastic leukemia of T-cells (T-ALL) with CBD resulted in mitochondrial dysfunction and cell death by altering their calcium homeostasis, which was directly demonstrated as the inhibitor of mitochondrial Ca

 uptake protected the T-ALL cells from CBD-induced cell death (16). Importantly, and of even further interest open to speculation, is that healthy, resting T-cells were resistant to the effects of CBD and avoided cell death! These data provide provocative evidence for CBD as a candidate for chemotherapy trials.

We now understand that not only does CBD intercalate into the plasma membrane, disrupting ion channels, but it also activates internalization of PM cholesterol which in turn destabilizes the ability of mitochondria to manage its own calcium homeostasis affecting multiple cellular processes including Ca

 signaling the downstream cell death. CBD also causes the translocation of other membrane proteins leading to decreased cellular energy (ATP) production. At play here is the translocation of a key mitochondrial membrane protein, hexokinase I (HK1). Loss of functioning HK1 results in the decoupling of glycolysis from mitochondrial respiration through diminished transformation of glucose to glucose 6-P (17). Through multi-omic profiling in combination with fluorescence resonance energy transfer (FRET)-based biosensor screening, the Colorado group’s results confirm an earlier report by Alharris and colleagues (2019) that CBD-induces mitochondrial dysfunction (18).

Mitochondrial dysfunction leading to apoptosis or cell death, reported in the Colorado study, has also previously been shown in pancreatic b-cells with associated insulin resistance prior to cell death a hallmark in type 2 diabetes (19). Findings from another relevant study this time involving neurons and platelet-activating factor (PAF), which is also influenced by calcium levels and PM cholesterol is worth citing here (20). PAF binds to its receptor (PAFR) located in the PM and nuclear membrane which mobilizes Ca

 and activates numerous signaling pathways including inflammation, which can result in cell death. If cells are treated with statins or CBD to reduce the level of cholesterol, then PAFR is displaced, PAF doesn’t bind to its receptor and neurons become resistant to apoptosis. This is a significant finding in light of another study where deletion of PAFR in a mouse knockout experiment abolished the inflammatory response and neuronal apoptosis caused by TBI (21). As a side note, Etizolam, a benzodiazepine analog, is a PAF-receptor antagonist used to treat anxiety.

The widespread self-medication with CBD, now accessible in a dizzying array of consumables readily available to the public, has preceded any clear understanding of the mechanisms behind CBD’s apparent benefit across a wide range of ailments. The far-reaching cellular disruptions observed in the Colorado study were corroborated by transcriptomics, metabolomics, and proteomics showing the downstream impact from disruption in cholesterol biosynthetic and regulatory pathways including decoupling of glycolysis from mitochondrial respiration, upregulation in oxidative stress response, increase in Ca

 cytosolic levels, and the increased production and storage of cholesterol sensitizing cells to apoptosis making cholesterol the effector molecule of CBD across many intracellular pathways rather than any specific target.

The Colorado study has shown that not all CBD effects on cells are beneficial. While there had been earlier reports from human clinical trials of the now Food and Drug Administration (FDA)-approved CBD product, Epidiolex, indicating that CBD treatment could lead to risk of liver injury as indicated by increased levels of aminotransferase activity and inhibition of P-450 (CYP) enzymes (22), the molecular mechanisms of that potential toxicity were not fully understood (23).

It should be mentioned that the Colorado study was carried out on two immortalized human cell lines: SK-N-BE(2) neuroblastoma cells and HaCaT human keratinocyte cells with CBD treatment in the micromolar range which would be less than the prescription dosing of Epidiolex, but probably more than self-administered CBD dosing. Their analyses based on transcriptomics showed that the enriched transcription factors shared oxidative stress as a stimulus, resulting in upregulation of oxidative stress response including fatty acid and cholesterol biosynthetic gene transcripts. It was known that CBD upregulates mRNA transcripts in microglial cells involved in fatty acid metabolism and cholesterol biosynthesis (24). This alteration in fatty acid metabolism was also shown in a mouse chronic alcohol feeding study where CBD reduced the infiltration of liver cells with fat, a distinct benefit (25).

If anything, this study has brought attention to the need for basic research on cannabinoids and the biological systems they effect to clarify why endocannabinoids are synthesized locally on an as needed basis in nanomolar amounts and then are quickly degraded. Fundamentally, the endocannabinoid system presents a unique opportunity to look at how a lack of essential nutrients, specifically lipid-based foods, or an overabundance of low-quality nutrient intake impacts synaptic plasticity and resulting outcomes in physical and mental health.

https://www.researchgate.net/publication/341935024_Multi-Omic_Analysis_Reveals_Cannabidiol_Disruption_of_Cholesterol_Homeostasis_in_Human_Cell_Lines

 e02882. DOI: 10.7554/eLife.02882 (2014).

, is Chief of Science at CLIP Labs in San Diego, California. Direct correspondence to: 

Articles

Op-ed on a new study related to how cannabidiol (CBD) exerts its clinical efficacy.

New Study Links Cannabidiol’s Multitudinous Effects to Disruption of Cholesterol Homeostasis

In September the United States House of Representatives will be voting on an extensive cannabis legalization bill, which has been labeled the Marijuana Opportunity, Reinvestment, and Expungement (MORE) Act. Majority Whip James Clyburn (D-SC) has said in an email blast to members that the chamber is anticipated to take up the MORE Act during the “September work period” (1).

This historic bill would federally deschedule cannabis, expunge the records of individuals with prior marijuana convictions, and also impose a federal 5% tax on sales. This added revenue would then be reinvested in communities most impacted by the drug war. Aside from these legislation terms, the MORE Act would create a new pathway to resentence individuals who were incarcerated for marijuana offenses, help protect immigrants from being denied US citizenship over cannabis, and restrain federal agencies from denying public benefits or security clearances because of its use.

Representative Earl Blumenauer (D-OR), co-founder of the Congressional Cannabis Caucus said (1), “As people across the country protest racial injustices, there’s even greater urgency for Congress to seize this historic opportunity and finally align our cannabis laws with what the majority of Americans support, while ensuring restorative justice.”

Earlier this month, the American Civil Liberties Union, NAACP, Human Rights Watch, Drug Policy Alliance (DPA), ordered for a floor vote on the MORE Act to take place by the end of September. “Advancing this bill meets the principles of this political moment where millions of Americans are demanding changes to the status quo,” said Queen Adesuyi, policy manager for DPA (1). “House Democrats have the incredible opportunity to move a bill that enjoys bipartisan support while responding to the calls for justice reform and racial justice reverberating across the country.”

“We are facing challenging times in our fight for racial equity and law enforcement reform, and passing the MORE Act is crucial to addressing our country’s history of injustice,” Representative Barbara Lee (D-CA), co-chair of the Congressional Cannabis Caucus said (1). “This is the next step in our goal of ending the harmful prohibition of cannabis in 2021.”

If the bill is passed by the House it still needs to pass the hurdle of a Republican-controlled Senate. Majority Leader Mitch McConnel (R-KY) has become an advocate for hemp (2) but has held a relentless opposition to broader cannabis reform.

There is the possibility that the MORE Act passing in the House might ignite the Senate to bend on their stance a little bit and take up cannabis reform legislation such as the Strengthening the Tenth Amendment Through Entrusting States (STATES) Act. This bill would grant states the opportunity to set their own cannabis policies without the fear of federal intervention. The MORE Act is gaining traction in support through Representative Matt Gaetz (R-FL) as the sole GOP cosponsor on the bill who approves removing marijuana from the Controlled Substances Act (CSA) and ending federal prohibition (3).

“Passage of The MORE Act is essential in order to truly right the wrongs of federal marijuana criminalization, and to once and for all allow the majority of states that have legalized cannabis for either medical or adult-use to embrace these policies free from the threat of undue federal prosecution or interference,” said Paul Armentano the Deputy Director of NORML (2).

While no official date has been set for this exciting floor vote, leading democrats have been hinting that the strongest step toward federal legalization of cannabis never before taken by Congress—the MORE Act—vote will take place around September 21st (4).

 Since this article was originally posted, an announcement was made that the vote will be delayed until after the election in November 2020.

https://www.marijuanamoment.net/house-to-vote-on-historic-marijuana-legalization-bill-next-month-leadership-announces/

https://www.marijuanamoment.net/mitch-mcconnell-says-legal-hemp-industry-off-to-a-rocky-start/

https://www.marijuanamoment.net/gop-congressman-will-vote-yes-on-marijuana-legalization-bill-despite-reparations-concerns/

https://www.leafly.com/news/politics/marijuana-legalization-more-act-news

The U.S. House of Representatives will be voting on a historic cannabis legalization bill in September, titled the Marijuana Opportunity, Reinvestment, and Expungement (MORE) Act.

US House of Representatives to Vote on Momentous Cannabis Legalization Bill in September

The mechanics of grow light technology is continuously advancing to better suit the needs of growers. Through these constant improvements, cannabis cultivators are able to better provide the proper necessities for the best cannabis growth. Here, Noah Miller, CEO of Black Dog LED, delves into the world of grow light technology and how it will continue to evolve.

How did you and your company get involved in the cannabis industry?

 Black Dog LED was purpose-built to be involved in the cannabis industry from the very beginning—we did not pivot into the industry. The founder wanted a quality light that was less dangerous to use in a home environment than high-pressure sodium vapor (HPS) lights—the most common type of lighting available at the time being used to flower cannabis. He tried some of the early light-emitting diode (LED) grow lights on the market, but was sorely disappointed, and felt he could do better. Fast forward 10 years and we are on our fifth generation of lights and have developed the most unique spectrum focused on growing high-quality cannabis.

What is the correct spectrum for flowering cannabis plants?

 This question is almost impossible to answer since the permutations available to us are too many to test in a lifetime. Creating the perfect spectrum is something that may take another 50 years, however, with the research available to us we have been able to engineer a spectrum that accomplishes the goals of most indoor cannabis cultivators. These primary goals are, of course, yield and quality, but also include controlling plant morphology (growth pattern) so that the plant is easier to control and flower indoors as well as other benefits we can achieve with a properly engineered spectrum.

What is the difference between your LED grow lights and others that are on the market?

 The main difference between our lights and the others is the countless hours of scientific research spent in identifying what makes cannabis plants grow and flower. Our team comes from a scientific background and we understand that different wavelengths of light result in different effects within a plant. As such, we sought to develop a grow light that focused on flower yield and quality, which differs from other spectrums that focus on growth, which would be a better fit for leafy greens.

Can you provide further detail on the lighting spectrum your grow light technology produces?

 In the past, grow lighting has been provided by technologies designed to illuminate areas for humans, which also happened to grow plants. Fluorescent/induction lights work for very short plants, but are inefficient and lack the intensity necessary to grow taller plants. High-intensity discharge (HID) lighting provides this intensity and better efficiency but limits plant growth due to inadequate spectral coverage and intense heat. All of these technologies were designed for illuminating areas for people, not plants. LED technology allows the creation of lighting with both high efficiency and a completely tuned light spectrum.

Our product is the result of many years of research—and even more importantly years of researching cannabis grows—to fine-tune an LED light spectrum for growing plants that maximize yield, quality, and efficiency.

, which is how efficiently a plant can carry out photosynthesis at different wavelengths of light. By carefully controlling ratios of different colors of light we can not only efficiently fuel plant growth but also affect their morphology, creating denser, more-compact plants that waste less of their energy creating stems, leaving more energy for flowers and fruits.

Photosynthesis isn't the only light-driven process in plants. Secondary metabolites (compounds) are created in a process called 

, many based on colors of light not used directly for photosynthesis. Our spectrum includes ultraviolet (UV) and infrared (IR) light to stimulate plants to create these quality-enhancing compounds. While UV and IR are both outside of the parabolic aluminized reflector (PAR) lighting region, they are present in natural sunlight and are critical to achieving plants' full potential.

How did you research LED grow lights in the cannabis industry? What other research is your company working on related to cannabis cultivation?

 We researched our lights against competitors in highly-controlled testing environments, growing the same strain of plants, in rooms of the same dimensions, and focused on ensuring that all variables were controlled as much as possible so that the only notable difference would be the effectiveness of the light and its spectrum. We’ve repeated these tests hundreds of times, focusing on different variables like leaf surface temperature, to hone in on our final product which, we believe, is the most advanced spectrum ever created and is backed by research. We’ve also recently opened an R&D center focused on Farm Bill—compliant cannabis hemp flower, focusing on the unique light spectrum needed to grow hemp—a close cousin of cannabis but with lower tetrahydrocannabinol (THC) yield.

What should people know to be able to determine the ideal LED grow light setup or what will work best for their plants?

 Each grower’s setup is unique, and a large part of our work involves visiting our customers’ grow sites in advance to view the facility, map out their grow room, and set up lights in a pattern that maximizes lighting exposure to boost yield per square foot.

To do this, we’ve developed a propriety light mapping software. This is completely different from the rest of the market who use traditional computer-aided design (CAD) software which is designed for buildings and general construction but not tailored to the unique needs of cannabis cultivators.

And, of course, each grower must be cognizant of the laws and regulations within their state, county, and town as they can vary dramatically across the US and globally.

How do you address the different limits or electricity caps that some states (such as Massachusetts) have instituted for cannabis cultivation sites?

 Black Dog LED is uniquely positioned to help with these challenges, and we support further environmental stewardship in the cannabis industry around cultivation. By working closely with our customers, we have found that we can still help them achieve their cultivation goals while working within the confines of different state regulations. This entails a discovery phase upfront that explores the needs and goals of an organization as these can be different. Yes, all operations want to grow cannabis, but their unique goals can help dictate the lighting deployment and affect energy use.

By taking the time to learn about each commercial project allows us to ensure the success of the customer. Most customers are surprised by our desire to dig into the details, as they are used to organizations that just want to sell them lights, but we feel this partnership with the customer is critical in helping the customer get the most out of their lighting infrastructure purchase.

Are there any differences in working with cultivators in the US compared to Canada?

 Canada tends to have larger, more well-funded operations than in the United States due to the legal climate up there. The rush to build out in Canada early on occurred primarily before LED had really caught on in the commercial cannabis industry. Further, there has been a major correction in the market with some operations even being mothballed recently. We feel that Canada will have a large opportunity to retrofit newer LED technology in the next 1-3 years. The operators that make this transition to the new tech will have a competitive advantage over the companies producing cannabis at a higher cost due to older, less efficient technology.

What are the next steps Black Dog LED plans on taking in growing their LED line? Will the company expand to other areas with cannabis production?

 We’re always adjusting our lights based on our latest research findings and hope to announce our latest product soon. That said, our immediate focus is getting through COVID-19 which has been one of the most impactful events of our lives that we’ll look back on in the same view as The Great Depression.

We’re also continuing to develop research based on hemp flower and have recently announced our hemp grow room, which is currently taking a majority of our focus.

How has the COVID-19 pandemic affected your company?

 Like many others, COVID-19 forced us to rethink how we approach business in a way that protects our employees while keeping the business running. In the immediate sense, that meant shifting to remote work several weeks before the state began its mandates—and we were fortunate to already have a strong work from home culture and processes in place. As for business, we’ve actually seen an increase in our home growing kits due to everyone having more time to devote to new hobbies, including home growing and indoor gardening.

How have you assisted in aiding the community during the COVID-19 pandemic?

 Being a smaller company with somewhat limited resources. We have focused on our community of growers and especially our Black Dog LED family of employees. For the grower community, we have doubled down on providing education online with no paywall or any purchase requirements. We want to do anything we can do to help people cultivate safely at home during this time. For our employees, we have given them the ability to participate in making decisions about how we address these unprecedented times as well as allowing them to make their own decisions based on their level of comfort and personal risk. While the time has been difficult, our team has truly risen to the occasion and we couldn’t ask for a better group of people to be working as we navigate these challenges.

What methods of protection are you taking to keep workers and customers safe?

 Like many others, we have transitioned to primarily remote operations with only essential activities such as shipping and receiving taking place in-person, and only when necessary. Remote work has long been a part of our company’s DNA since, living in Colorado, snowstorms and blizzards will often shut down roadways and force us to work from home anyway. We’re also limiting nearly all in-person meetings—opting instead for Zoom calls which are nearly as effective—and do not plan on attending trade shows or large gatherings until it is safe to do so.

With grow light technology constantly advancing, Noah Miller, CEO of Black Dog LED, discusses the research behind grow light technology and its future in cannabis.

The Science Behind LED Lights

In a recent press release, Bruker (Billerica, Massachusetts) and Purity-IQ (Mississauga, Canada) announced their joint partnership with a goal to create a new testing system aimed at helping the medical cannabis industry in Canada (1). The Purity-IQ Cannabis Fingerprint

 incorporates Bruker’s expertise in nuclear magnetic resonance (NMR) based screening technology with Purity-IQ’s genomics capabilities to provide traceability and identity testing throughout the cannabis supply chain.

Currently, there are no standardized regulations in effect between states or countries to regulate quality control in cannabis. Without these regulations in place, it makes it difficult for consumers to make informed decisions about the products they are buying. According to the two companies, uniting these technologies will help give the supply chain an opportunity to progress the cannabis industry forward to a consistent and compliant platform.

With regulatory standards continuously shifting, cannabis producers and manufacturers need to be able to ensure product quality and consistency of strain and dosage. Also affected are the retailers such as pharmacies and dispensaries who need to protect their brand and offer consumers a product that can be trusted.

According to the press release, identifying the genomic profile and chemical compounds in a cannabis cultivar is the key to being able to ensure dosing assurance and consistent quality. Genomics are used to establish cannabis identity and heredity. NMR identifies the consistency and purity of the product at a molecular level, which offers traceability, authenticity, and identity throughout the supply chain from plant to finished product. This creates a unique chemical “fingerprint,” using the Purity-IQ Global Cannabis Registry to devise new Cannabis Authenticity and Purity Standards (CAPS).

In addition, the announcement stressed the advantages of using NMR compared to other technologies available on the market. NMR is an untargeted analysis method that reportedly provides an extensive view on the metabolome. With the use of NMR, it allows for an easier determination of the relative concentration of substances in a mixture accurately and rapidly.

“A genetic fingerprint alone is not enough to produce a full picture. To generate a complete fingerprint, cannabis strains must also be profiled using NMR technology,” said Dr. Henry Stronks, President of Bruker Canada, in the press release (1). “Pattern recognition allows us to identify peaks that correspond to the pharmacological properties of the cannabis, its geographical location, strain, hybrid and quality. We believe this is the first step for the cannabis industry in providing safe, uniform and effective products founded in combined genomics and metabolomics testing.”

Terry Dennis, Chief Marketing Officer at Purity-IQ also commented on the collaboration with Bruker (1). “CAPS recognition will identify and differentiate products, build intellectual property, ensure authenticity, drive confidence in product quality claims, allow access to new markets, and protect brand reputation. There is a whole host of other sectors that would benefit from the combination of genetic and metabolomic fingerprinting, particularly in the food and beverage industry in the fight against food fraud,” said Dennis. “This will support the whole value chain and, one day, consumers will have full transparency, trust and confidence in all products they buy.”

Through this newly formed partnership, both Bruker and Purity-IQ plan to apply their technology to better aid the cannabis industry and promote consistency along with compliancy. Bruker’s 400 MHz NMR spectrometer and its Fourier 80 benchtop NMR system are installed at the University of Guelph in Canada, where Bruker’s and Purity-IQ’s scientists plan to collaborate in research and development with the Natural Health Products (NHP) Research Alliance. The NHP is a collaborative body based at the University of Guelph, creating novel DNA-based ingredient identity tools founded on a global standard biological reference materials DNA library for natural health products.

https://www.bruker.com/news/bruker-and-purity-iq-sign-partnership-agreement-to-transform-the-cannabis-regulatory-landscape.html

Bruker and Purity-IQ announced their joint partnership with a goal to create a new testing system aimed at helping the medical cannabis industry in Canada.

Bruker and Purity-IQ Partner to Transform Cannabis Regulatory Landscape

® a multimedia platform that focuses on educating the legal cannabis industry about the science and technology of analytical testing, quality control, extraction, and cultivation, will host its first 

Hemp Science & Technology Virtual Symposium. Brian C. Smith, Ph.D.,

 founder, CEO, and chief technical officer of Big Sur Scientific, will serve as the two-day event’s program chair.

The two-day virtual symposium will take place Oct. 28-29, 2020 and is free to attend. The symposium will feature talks from hemp scientists, researchers, and industry experts on a wide array of topics, including hemp analysis, cultivation, and extraction.

“During this health crisis, it’s more important than ever to offer resources and virtual events to our community that share knowledge and help the hemp industry continue to push forward,” said Mike Hennessy Jr., president and CEO of MJH Life Sciences™, parent company of 

®. “We are excited to offer a new dedicated event for hemp professionals to learn more about the science and technology that can improve their work or lead to better products for consumers.”

® to offer the Hemp Science & Technology Symposium to our industry,” Smith said. “I have three state-of-the art symposia planned: New Developments in Hemp Testing, Optimizing Hemp Extractions and Overcoming Hemp Growing Challenges. Each session will feature talks by industry experts, and a roundtable discussion where attendees can ask the experts questions. Don’t miss this opportunity to learn how to improve your products and operations and advance your career.”

Details on speakers and presentation titles will be announced in the coming weeks. To register, click 

® is a multimedia platform that focuses on educating the legal cannabis industry about the science and technology of analytical testing, quality control, extraction, and cultivation. The magazine is accompanied by an online component and provides relevant information and tutorials for all members of the cannabis industry. 

™, the largest privately held, independent, full-service medical media company in North America dedicated to delivering trusted health care news across multiple channels.


Alexandra Ventura, 609-716-7777, ext. 121


Cannabis Science and Technology® will host its first Hemp Science & Technology Virtual Symposium. Brian C. Smith, PhD, founder, CEO, and chief technical officer of Big Sur Scientific, will serve as the two-day event’s program chair.

Cannabis Science and Technology® to Host Hemp Science & Technology Virtual Symposium

Hemp is a naturally variable, inhomogeneous material. Analyzing one or a few cuttings from a grow will not give data representative of a field. Here I propose a scientifically based representative sampling plan for hemp grows detailing how to collect composite samples, how many to collect, and how to analyze them. I want this industry to commit itself to the goal of analyzing one plant per 1000. Technologies available to handle the resultant increased sample load are reviewed.

With passage of the 2018 Farm Bill and promulgation of the United States Department of Agriculture’s (USDA) interim final rule in October 2019, it is now legal to grow hemp provided the total tetrahydrocannabinol (THC) level in dried plant material does not exceed 0.3% dry weight (1–3). To enforce this law, extensive testing of hemp crops is being performed around the country by state departments of agriculture. The USDA has issued a sampling guidelines document (3). The USDA sampling document defines a “lot” of hemp plants as a (3) “…contiguous area in a field, greenhouse, or indoor growing structure containing the same variety or strain of cannabis throughout.”

My concerns with this sampling plan are noted below, along with my proposal for an improved hemp representative sampling plan. First however, we must address the topic of sampling error, something a good representative sampling plan tries to minimize.

The entirety of anything that we wish to sample, be it all the people in the US or all the plants in a hemp grow, is called a 

 (4). In a perfect world, we would measure the quantity of interest in the entire population. For example, asking the opinion of everyone in the US for a poll, or analyzing every hemp plant in a field. However, this is impractical given the enormity of the task. Instead, we choose a subset of the population from which to collect data, for example, we could choose to ask 1000 Americans their opinion for a poll or choose to analyze one plant per 1000 in a hemp grow. Sampling error rears its ugly head because there is always a chance the population subset we choose to analyze will not give us the same result as if we had analyzed the entire population.

Sampling error is different from the random error discussed in a previous column (5). Random error is caused by things we have no control over such as instrumental and environmental fluctuations. Random error is minimized by more and better testing (5). When testing a hemp field then for cannabinoid content, the amount of error in the final measurement is a combination of random error and sampling error (assuming systematic error is zero [5]).

I am not a statistician but a chemist, so I refer you to these references if you desire a detailed mathematical treatment of margin of error and population subset size (6,7). However, everything I have read on the subject points to the same conclusion: the way to minimize sampling error is to maximize the size of the population subset that is analyzed. Intuitively this hopefully makes sense. There are more than 300 million people in the United States. When running an opinion poll, the more people you talk to the better the chances you will gather data that is representative of the nation. When it comes to hemp fields, when taking one or a few cuttings there is a high chance that the plants sampled are not representative of an entire grow. The way around this is to analyze as many hemp plants in a field as is practicable.

The limiting factor in performing representative sampling for hemp field testing is time and expense. Therefore, we must find a happy medium between minimizing sampling error while not generating an inordinate number of samples for testing. I believe the best compromise here is achieved if we test one hemp plant per 1000 in a field. I want our industry to commit itself to this testing goal.

The USDA hemp sampling document (3) presents equations for calculating the minimum number of cuttings to collect in a lot. Example calculations are given and are summarized in tables showing the minimum number of cuttings to collect based on the acreage of hemp plants. In the examples given, one cutting per acre or less is collected per lot. The document calls for compositing all cuttings into a single sample. Typically, one aliquot of this composite sample is tested (8–10).

The following sections summarize my concerns about this approach.

The calculations for the minimum number of cuttings to collect are based on this assumption (3): “…the proportion of hemp plants having THC content greater than the acceptable hemp THC level…are based on past experience in the same or similar growing areas.”

The example calculations assume the proportion of hot hemp plants is 0.01 (3).

One problem with this assumption is that in some areas of the country hemp is being planted for the first time this year, so the needed historical data does not exist. Additionally, since it only became legal nationwide to plant hemp as of January 2019 (1), there is little historical data for the rest of the country upon which to base an assumption of the proportion of hot plants. Lastly, even if we had enough historical data, given the vicissitudes of the weather, every growing season is going to be different in every area. Absent any peer-reviewed data to the contrary, I do not believe it is safe to assume that any given growing season will be predictive of any future growing season. Rather than basing a sampling scheme assuming we know the proportion of hot plants, would it not make more sense to measure that proportion of hot plants instead?

My concern here is that, on average, the USDA sampling document calls for collecting one cutting per acre or less (3). The problem is that hemp farmers plant at different densities. For example, Farmer x may grow 2000 plants/acre, while Farmer y may grow 4000 plants/acre. Remember that the population being sampled is the hemp plants, not the area of the field they occupy. Basing the number of samples to collect based on acreage is like choosing the number of people to include in an opinion poll based on a state’s area. Using this logic, you would talk to more people in Alaska rather than New Jersey for a poll, even though New Jersey has 12 times as many people as Alaska (11).

Our goal should be to achieve the same level of sampling error for all lots analyzed. To do this, the number of samples to collect needs to be determined by the number of plants in a plot, not their acreage. In our examples then, and following the goal of sampling one plant per 1000, Farmer x’s plot should be sampled at a rate of 2 plants/acre, while Farmer y’s plot should be sampled at a rate of 4 plants/acre. Only by basing the sample size on the number of plants in a plot can we assure the best representative sample is obtained.

Where on the plant should we collect cuttings? The USDA sampling document states that cuttings are to be taken from the top 1/3 of the plant (3). Work by myself and others has clearly shown that cuttings from the same hemp plant vary in cannabinoid content (12) as seen in 

The top of a hemp plant normally has the highest cannabinoid content, and hence only collecting plant tops means a sample will be collected that will be biased towards containing more THC than the field in general. This makes it more likely that its total THC content will be above 0.3% and that the farmer will lose his crop.

Ultimately, I believe that the plant parts sampled need to reflect how the crop will be harvested. For smokable hemp, the buds are the article of commerce, which means that harvesting the top 1/3 of the plant may be appropriate for this type of harvest. However, when growing hemp for extraction, the entire plant may be ground up and utilized. In this case cuttings from the top, middle, and bottom would be appropriate.

Composite Samples Are Not Necessarily Uniform

The USDA’s sampling document calls for compositing all the cuttings from a lot into one sample (3).

There are two problems with this. First, the regulations assume that every composite sample is homogeneous. Where is the peer-reviewed science on this? A simple visual inspection of ground hemp samples will show they are not uniform. In my own experience, I have found that composite samples are not uniform. I have taken composite hemp samples and had their cannabinoid content measured in triplicate at an ISO licensed, state certified laboratory (13). I found cannabinoid concentration variation across the samples well above the margin of error as reported by the laboratory, which means these composite samples were not homogeneous, and that sampling error exists for composite samples.

The second problem with composite samples is only testing them once. This assumes the sample is uniform. Wouldn’t it be better to test multiple aliquots and reduce the sampling error rather than only testing one aliquot and assuming the sampling error is zero? When it comes to science, I strongly feel that actual measurements are always better than assumptions. At minimum three aliquots of composite samples should be tested and an average and a standard deviation calculated to get a handle on the sampling error.

Another concern I have is that different states are sampling in different ways (8-10). Some states only collect a specific number of cuttings regardless of grow size. Some states are using sampling based on acreage and not number of plants. Some states give specific details on how to walk a field and collect samples, but there is no agreement on what pattern to walk. Lastly, some states are testing using liquid chromatography (8,9), while others are using gas chromatography (10). Until there is nationwide standardization on how to collect and analyze hemp samples, different states will obtain different results on the same samples.

A Proposed Hemp Representative Sampling Plan

Patricia Atkins and I have written in this journal on the importance of representative sampling in cannabis testing (14,15). I offer here my representative sampling proposal for hemp. This proposed plan does not necessarily conform with regulations promulgated by the USDA or any state’s department of agriculture. My proposal is based on my understanding of statistics, my 30+ years experience in chemical analysis, and my PhD in physical chemistry.

The gist is this. Rather than assuming we know the proportion of hot plants, why not use statistical sampling techniques to generate enough data to get an excellent estimate of the average total THC value for a hemp lot? Rather than only collecting one composite sample per lot, assuming that sample is uniform, and then only testing it once, why not collect multiple composite samples from a lot, test each of them multiple times, and maximize the amount of data we have on a lot minimizing the sampling error?

The number of samples to collect from any given lot needs to be based on the number of plants in that lot as the plants are the population we are testing and care about. I want our industry to commit itself to sampling one hemp plant per 1000 planted.

My own instinct on this is that sampling locations should be chosen randomly. This is to make sure that location does not bias the sample. For our 10,000 plant example plot this means 10 locations should be chosen at random. However, there is work indicating that collecting samples according to a pattern is a good idea as well (15). I am persuadable on what might be the best approach here.

What Parts of the Plant Should Be Sampled?

Research has shown that the cannabinoid content from different parts of the same plant varies appreciably (Figure 1). As stated above, the portion of the plant tested should correspond to how the plant will be harvested. If only plant tops are to be harvested, then only plant tops should be tested. If the entire plant is to be harvested then cuttings from the top, middle, and bottom of plants should be collected and tested.

Since cannabinoid content varies from plant to plant, taking a cutting from only one plant at a given location might not be representative of that spot. To overcome this problem, cuttings from three plants at a given location should be collected to obtain a more representative sample. If harvesting just plant tops, the upper 1/3 of nine plants from a given location should be sampled. If harvesting the entire plant, then a cutting from the top, middle, and bottom on each of three plants for nine cuttings total should be collected. The cuttings from a given location should be combined to comprise a composite sample representative of that spot. For a 10,000 plant hemp lot then, 10 composite samples should be collected from 10 locations.

The law clearly states that dried hemp is the material to be tested. In my own work I dry hemp cuttings at 140 °F in a standard food dehydrator overnight. This gets the material down to about 5% moisture. I grind about 5 g of dried cuttings in a coffee grinder for 1 min to homogenize the sample. There are other ways of drying and grinding hemp. I welcome input on this. The most important thing is that we, as an industry, standardize how we prepare hemp samples for analysis (16).

How Many Aliquots of Each Sample Should Be Tested?

The problem with compositing all the cuttings from a lot and testing only one aliquot were discussed above. At minimum, any composite sample should be tested three times and the results averaged to obtain a more representative picture of the composite. Using our example then, each of the 10 composite samples should be tested three times for a total of 30 data points.

For a given hemp lot, the average total THC value for a lot should be calculated. For our example hemp plot, this would mean calculating the average of 30 values. The standard deviation (17) of these values should be calculated as well. This number is a measure of the variance in the data, and it is important because it takes into account sampling error.

The “acceptable hemp THC level” as defined by the USDA is calculated by adding the margin of error for total THC for a given method to 0.3 wt. % total THC (2). For example, if the margin of error is 0.04 wt. % for an analyzer (18), the acceptable hemp THC level for this analyzer would be 0.34%. If the average total THC value for a given lot is within a 95% confidence interval (17) of the acceptable hemp THC level for an analyzer, it should be considered legal. For hemp with total THC values above this level it should be considered illegal.

For compliance purposes the law states that a hemp grow should be sampled within 15 days of harvest (1,2). However, for the purposes of managing a hemp grow to maintain compliance, I recommend farmers sample and test their crops more often. One approach is to begin testing about halfway through the growing season, and to test a lot once per week. I highly recommend tracking your results over time to spot trends. As harvest approaches, sampling a couple of times a week is a good idea.

For our 10,000 plant plot, I recommended collecting 10 composite samples and performing 30 analyses total. Sending these samples out to a third party laboratory is not practical because it can take weeks to get results and may cost thousands of dollars to analyze one weeks’ worth of samples. This means that hemp farmers need to perform their own testing to manage their grows to maintain compliance.

In a previous column, I compared different technologies for cannabis potency testing (19) using the criteria of speed, accuracy, cost, and the ability to perform representative sampling. By necessity, any technique that is going to be used for representative sampling must be fast, easy, and inexpensive to make handling the representative sampling analysis load feasible.

I showed (19) that chromatography can accurately analyze a given sample but it is slow and expensive to use. As a result, it is not amenable to representative sampling. Another problem with chromatography is its difficulty of use, which is why some states require degreed scientists to run these instruments (19). I believe these problems with chromatography are why we are not performing proper representative sampling in this industry.

Spectroscopy is fast and inexpensive, but because it is a secondary technique it is not necessarily as accurate as chromatography (20). There exist spectroscopic analyzers capable of an accuracy or ±0.04 wt.% accuracy for the determination of total THC in dried, ground hemp (18). Since this is several times lower than the legal total THC limit of 0.3%, these analyzers are of sufficient accuracy to measure total THC in hemp. This technology is fast enough, inexpensive enough, and accurate enough to handle the sample load required to allow true representative sampling of hemp lots for farmers and regulators.

For state departments of agriculture, I recommend a two-tier approach to efficiently handle the sample load necessitated by proper representative sampling. Infrared spectroscopy could be used as an initial screen of samples. If a lot tests as being legal that is the end of it. If a lot tests hot, samples should be subsequently tested by chromatography to confirm the analysis. The advantage of this approach means that time and money are not wasted analyzing legal samples by chromatography.

There is natural variability amongst hemp plants, hence their grows need to be sampled in a representative manner. There are issues with how hemp fields are currently sampled. Here I propose that one plant per every 1000 should be analyzed. This is a reasonable compromise between minimizing sampling error and not generating an overwhelming number of samples to test. How to collect, prepare, analyze the samples, and report the data was discussed. Technologies available to handle the increased sample load resulting from representative sampling were covered.

115th United States Congress, Senate Bill S.2667,” Hemp Farming Act of 2018.

https://www.federalregister.gov/documents/2019/10/31/2019-23749/establishment-of-a-domestic-hemp-production-program

https://www.ams.usda.gov/sites/default/files/media/SamplingGuidelinesforHemp.pdf

https://en.wikipedia.org/wiki/Sampling_error

https://en.wikipedia.org/wiki/Sample_size_determination

https://agriculture.delaware.gov/plant-industries/hemp-program

Massachusetts State Hemp Plane, Massachusetts Department of Agricultural Resources, Submitted to USDA March 9, 2020.

Kentucky Department of Agriculture Standard Operating Procedure KDA-HEMP-20190204-1, issued Feb. 4, 2019.

https://en.wikipedia.org/wiki/List_of_states_and_territories_of_the_United_States_by_population

M. Giese, M. Lewis, L. Giese, and K. Smith, 

https://en.wikipedia.org/wiki/Confidence_interval

California Bureau of Cannabis Control Regulations, Section 5719.

, is Founder, CEO, and Chief Technical Officer of Big Sur Scientific. He is the inventor of the BSS series of patented mid-infrared based cannabis analyzers. Dr. Smith has done pioneering research and published numerous peer-reviewed papers on the application of mid-infrared spectroscopy to cannabis analysis, and sits on the editorial board of 

. He has worked as a lab director for a cannabis extractor, as an analytical chemist for Waters Associates and PerkinElmer, and as an analytical instrument salesperson. He has more than 30 years of experience in chemical analysis and has written three books on the subject. Dr. Smith earned his PhD on physical chemistry from Dartmouth College.

Here a scientifically based representative sampling plan for hemp grows is presented, detailing how to collect composite samples, how many to collect, and how to analyze them.

A Proposed Representative Sampling Plan for Hemp Grows

This article reviews the chemical makeup of cannabis extracts and how they affect the human body. From terpenes to cannabinoids, this article takes a deep dive into the chemistry of cannabis extracts. It also demonstrates the opportunity for discovery and what we have yet to uncover.

Cannabis is widely known for tetrahydrocannabinol (THC) and cannabidiol (CBD), two prominent compounds found in relatively high concentrations in the flowers of 

 (1). As the legal cannabis industry develops, product offerings have expanded exponentially. Smoking cannabis flower is no longer the only option available to consumers, though it does maintain its position as the majority of product sales (

Cannabis extraction has emerged as the gateway to product development. Various solvents are used to produce a number of different cannabis extracts, which can be sold as inhalable products or used to infuse ingestible and topical products. The chemical profile of the extract is dependent on the solvent used during the extraction, the parameters used during the extraction, as well as the input material.

 (2) is optimal for efficient cannabinoid extraction. The solubility of ethanol allows for all grades of cannabis plant matter to be used in the extraction, with nearly 100% of cannabinoids able to be extracted within 20 min. However, by optimizing for cannabinoids this extraction method sacrifices terpenoids, flavonoids, and other light compounds within cannabis that are lost in the process.

Warm or Room Temperature Ethanol Extracts

Warm or room temperature ethanol extracts contain:

⦁ Cannabinoids (THC, CBD, cannabigerol [CBG], and so on)

⦁ Plant pigments such as chlorophyll and carotenoids

Because of the plant pigments and waxes, further refinement is necessary to remove these undesirable by-products. In turn, there is more loss of the delicate flavonoids and terpenoids leaving a highly concentrated extract full of phytocannabinoids. Some higher temperature ethanol extraction simultaneously decarboxylates the tetrahydrocannabinolic acid (THCA) turning it into THC. While we know the aforementioned compounds are found in the extract, we are still unsure of about 20–40% of the chemical makeup of cannabis extract depending on the cannabinoid potency. It is thought that warm or room temperature ethanol extracts are full of medically beneficial compounds and are best used in tinctures, capsules, and sublingual products.

The cold temperatures of cryogenic ethanol extraction allow for efficient extraction of cannabinoids without the by-products such as pigments or waxes. The result is a high potency cannabinoid fraction that is not decarboxylated. Advanced cold extraction techniques are able to produce inhalable products called 

, which is a hard-candy consistency product used as an inhalable. It is made up of 80–90% THCA crystals.

Ethanol extracts are growing in popularity because of their efficiency and potency, but lack in versatility. These extracts are often used to make high potency distillates as well as a variety of distillate infused ingestible products (

Hydrocarbon extraction started as a low-tech but innovative extraction method in early medical and illicit markets. Its products, known as “concentrates” or “dabs,” have been a popular choice for many experienced cannabis users because of the amplified flavor and higher concentration of cannabinoids.

Hydrocarbon solvents are used under cold temperatures and under relatively low pressures. This is fast and captures the desirable components of the cannabis plant.

⦁Terpenoids, especially mono-terpenes and mono-terpenoids

Because of the higher potency of cannabinoids and terpenoids, hydrocarbon extracts often have potencies over 75% cannabinoids (

While hydrocarbons quickly extract terpenes, aromatic hydrocarbons found in the plant, and cannabinoids extensive post-processing is required to remove the solvent from the extract. This requires more time and explosion-proof extraction and solvent removal equipment. The gravity of scalability and safety of this extraction method is significant.

Carbon dioxide is used in a number of other industries including hops oil extraction. Because of the nature of the cannabis plant, and the familiar similarities to hops, it was clear that 

 would be a good fit for cannabis (4). CO

 is easily manipulated into different phases using pressure and temperature. Naturally, this makes CO

 one of the most versatile solvents in cannabis extraction.

Despite requiring a longer extraction time, 

 from the plant because of its various phase properties (5). Subcritical and Supercritical CO

 are the two most popular phases for cannabis extraction. Supercritical fluids maintain the properties of both liquid and gas. This gives CO

 the ability to freely flow through the plant material, like a gas, while penetrating the trichomes and surface of the plant material, like a liquid. Subcritical CO

 is operated at a lower temperature becoming a less efficient solvent overall, but focusing primarily on the volatile aromatic compounds such as terpenoids and terpenes. CO

 is more efficient with extracting the full terpene profile of the plant including monoterpenes, sesquiterpenes, and a wide array of terpenoids, as demonstrated in a terpene fraction (

 pressures can be ramped up to the supercritical phase to increase the efficiency of cannabinoid extraction. With that, the extraction time is dramatically reduced but more undesirable components such as waxes and pigments are also extracted. This reduces the crude potency of the extract, but a wax removal process known as “winterization” can be utilized to remove the lipids. This increases the potency of the extract. The terpene fraction and cannabinoid fractions are blended together to create high quality extract for vapes (

In addition to the extraction solvent and parameters, the starting plant material also dictates the end result of the extract’s chemical profile. If starting with material that is low in terpenes but high in cannabinoids, the end result will be a low terpene high cannabinoid extract. If decarboxylated plant material is extracted with hydrocarbons, a THCA crystalline product will not be possible to produce as the THCA has already been converted to THC. There are many variables to the process that must be considered prior to extraction.

While testing methods have advanced significantly since the dawn of the modern era of cannabis, there remains a wide frontier of opportunity to discover and learn. Even in 2020, 10–30% of the total mass of a cannabis extract remains unidentified on a typical certificate of analysis (COA). This demonstrates that we are unable to identify a relatively large portion of the extract’s chemical makeup, or that it is not economically viable to do so. Generally, it is known that there are compounds such as esters, flavonoids, various pigments, alkaloids, and so on within cannabis extract. Despite the potential to influence experiences with cannabis, these compounds have yet to capture public interest or significant scientific inquiry with respect to product profiles and consumption.

Over time, discoveries have been made which demonstrate that each compound affects the human experience with cannabis. THC can enable euphoria, CBD reduces inflammation, limonene is energizing, 

 (7), linalool is relaxing—yet none of these compounds work the same together as in isolation. Each compound influences the behavior of the entire network to provide synergistic effects. Through studying cannabis interactions with the human body, scientists have unveiled entire networks of systems within the human body, namely the endocannabinoid system (ECS). Our understanding of cannabis chemistry remains limited, and this new frontier provides ample opportunity for research and insights that can enable people to get more from this plant.

https://en.wikipedia.org/wiki/Cannabis_sativa

https://www.edenlabs.com/coldfinger/ethanol-extraction-process/

https://www.edenlabs.com/co2-extraction/co2-extraction-process/

A review of the chemical makeup of cannabis extracts and how they affect the human body. From terpenes to cannabinoids, this article takes a deep dive into the chemistry of cannabis extracts.

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