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2020 EMERALD CONFERENCE POSTER PRESENTATIONS

Presenter:
Arun Apte

Company:
Cloud LIMS

Abstract:
With no federal oversight for the cannabis market, the challenge of regulating this industry is entirely on the states. However, the cannabis industry is expanding rapidly, and the governing bodies are not able to keep pace. Hence, some states have well-established regulations in place, and some are still in the process in the process of setting up. In such a scenario, the most viable option is to adopt the best practices and regulations from testing and calibration laboratories in other industries such as ISO/IEC 17025:2017. The ISO 17025 standard enables laboratories to demonstrate that they operate competently and generate accurate results. Therefore, several states have mandated the ISO17025 standard. Although ISO 17025 is a well-known standard for testing and calibration industry, however, it is new to the cannabis industry. Cannabis laboratories find it challenging to get accredited to ISO 17025 as they are hiring the staff who are not only new to the industry but also lack experience operating in a regulated laboratory environment, which in turn, delays accreditation, and subsequent production timelines. The talk will highlight how a Laboratory Information Management System (LIMS) can ease testing laboratories workload and help streamline the process by creating an efficient method to control documents, manage testing and training records, and manage equipment logs, calibration schedules —a primary requirement of ISO 17025. The automation, in turn, facilitates managing external audits and assessments.

Presenter:
Benjamin Katchman

Company:
PathogenDx

Abstract:
As the cannabis industry expands and becomes more heavily regulated, the need for screening tools to detect microbial contamination increases. To date, screening has focused on the raw product and there has been little emphasis on the actual facilities in which that product is processed, which generates the potential to be a source for “follow-on” contamination prior to sale. The following case study was performed to demonstrate the utility of high-resolution environmental screening in a cannabis production and processing facility. Samples were collected for assessment of microbial contamination across 11 locations throughout the facility. To compare the effectiveness of screening methods, each sample was assessed by traditional microbiological plating and by the PathogenDx “EnviroX” microarray, which interrogates for the presence of a set of 50 bacteria and fungi. The study was based on 56 swab samples. Each tested positive for bacterial and fungal contamination when analyzed by both methods. Throughout, the most prevalent species detected were Pseudomonas spp., Golovinomyces spp., and Cladosporium spp. While there were variations in the microbial species present within each of the rooms tested, EnviroX was found to detect contamination with higher sensitivity, and with much higher level of speciation, than traditional plating. It was seen that EnviroX microarray testing revealed temporal and spatial relationships not seen with plating. In summary, the data suggest that standardized monitoring deployed before, during and between grows may be key to preventing the spread of contamination, thus providing a higher yield of clean, safe product for the consumer.

Presenter:
Benjamin Katchman

Company:
PathogenDx

Abstract:
Cannabis, food, water, air and agricultural safety have grown to require testing of complex panels of pathogen contamination (bacterial and fungal). The size of the supply chain for each and the widening range of pathogens which must be tested on each are, we argue, no longer sustainable via the current “one-off” approaches to microbial testing. In particular, we argue that the historical gold standard of plate-based culture is time and labor intensive and lacks the sensitivity and specificity required to maintain public safety when large-scale, multiple-pathogen analysis is required. We show here that new technology (multiplexed PCR coupled to hybridization on a DNA probe microarray) can now detect and quantify a variety of bacterial and fungal pathogens inexpensively and simply: <8 hours without enrichment. For example, PathogenDx’s DetectX array contains probes for E. coli (pathogenic and non-pathogenic), S. entericaA. nigerA. fumigatusA. terreus, and A. flavus; with a LOD of 1 CFU/gram at 90% sensitivity and 100% specificity compared to plating. The PathogenDx QuantX DNA microarray contains probes for Total Aerobic, Bile Tolerant Gram Negative, Total Enterobacteriaceae/ Coliform and Total Yeast and Mold and reports values as CFU/gram. We demonstrate here linearity for all probes (R2 ≥ 0.9), probe specificity and concordance in comparison to plating with a dynamic range of 102 to 106 CFU/gram. In summary, we feel that DNA microarray technology of this kind could be a breakthrough:  allowing simple, fast, multiplexed microbial detection at sensitivity and specificity unmatched by other DNA platforms such as qPCR.

Presenter:
Chris Denicola

Company:
Prism Scientific

Abstract:
Potency testing is routine and boring but cannabis and hemp companies rely on it for business decisions and financial security. As with most things in the cannabis and hemp space, the analytical services companies repurposed off the shelf technology for quick monetary gains with little thought to how the products would actually be used. For potency testing, simple C18 HPLC columns were pushed out into the marketplace with hasty application notes demonstrating efficacy. However, in practice, its not that simple. The C18 phase needs help to resolve the various cannabinoids and that help comes in the form of mobile phase additives like formic acid and ammonium formate. These additives are critical to the separation, identification, and quantitation companies need but sublet changes in the amounts of these additives, either from an error by the technician or the evaporation of the solvents, can lead to erroneous results. What we will show in this poster is an analytical method for potency testing that does not require mobile phase additives. This will save time and money for analytical testing labs, reduce errors, and allow for extraction and manufacturing companies to add in- house testing with reduced risk or need of a skilled chromatographer. The secondary goal is to show that this method transfer to prep scale without modification. If a company is using prep LC or flash LC, the incorporation of this stationary phase will simplify the purifications or THC remediation.

Presenter:
Chris Denicola

Company:
Prism Scientific

Abstract:
During this presentation, Denicola will outline future-looking strategies CBD companies should utilize to reduce confusion for costumers and keep THC levels down in CBD products (without compromising the Entourage Effect) in preparation for FDA crackdown. A common concern for new consumers is the risk of failing a drug test following CBD consumption. Based on solubility and dissolution testing, we know that one serving of a CBD tincture testing at 0.3% THC will have three milligrams of THC. This would come as a shock for most consumers, especially if the product label reads “zero THC.” Denicola argues that future FDA regulations are heading towards the legal percentage of THC levels consumed per day instead of per dose, which has the potential to change the CBD landscape entirely. Consumers are also looking for clarity and consistency in terms of expected effects and suggested dose. This has been Denicola’s focus while formulating Santeer’s line of 95% soluble and 99% bioavailable tablets that have different effects based on the terpene formulation. Lastly, Denicola will touch on the butterfly effect of utilizing solubility-enhancing chemicals in CBD products and discuss how to leverage technology to achieve optimal solubility and identify the right volume and profile of terpenes for specific purposes.

Presenter:
Klause Schoene

Company:
Sartorius

Abstract:
The two-fold objective of this study was to 1) demonstrate that aqueous mobile phase preparations from purchased lab grade bottled water yielded comparable results to in-house prepared water (Sartorius Arium purified water Type 1 and Type 3) and that 2) using filters (Sartorius Minisart® RC, SRP, NY & NY Plus, all 0.2 µm) yielded comparable results to unfiltered specimens under the same mobile phase conditions. A blank matrix specimen (homogenized nettle leaf (Urtica dioica)) was spiked for calibrators and controls and analyzed without cannabinoid spikes (blanks, n=5) using designated mobile phases for each water type to assess any specimen affects from the water. In addition, a true cannabis specimen was filtered using each of the four different filter types (Sartorius Minisart® RC, SRP, NY & NY Plus, all 0.2 µm) and analyzed (n=5) using the corresponding calibration curves and controls.

Presenter:
Sareeta Nerkar

Company:
Pickering Labs

Abstract:
The legalization of hemp under the 2018 Farm Bill brought new opportunities as well as new challenges to both growers of industrial hemp and manufacturers of hemp-containing products. The Farm Bill classifies hemp as the plant Cannabis sativa L. and all its derivatives with a delta-9 tetrahydrocannabinol (THC) of not more than 0.3 % concentration. To comply with Federal laws, all producers need to test their products to determine THC content as well as the concentration of other cannabinoids, particularly CBD, that are associated with the pharmacological activity of Cannabis sativa L. plant. A new HPLC method with post-column derivatization was developed to analyze cannabinoids in hemp and hemp-containing edible products. This post-column method is based on reaction with Fast Blue Salt reagent under basic conditions. Detection at 475 nm is performed using a UV/Vis detector. The method utilizes a simple extraction procedure with no additional clean-up and is suitable for analysis of the major neutral cannabinoids as well as cannabinoid acids with high sensitivity and selection of detection.

Presenter:
Sareeta Nerkar

Company:
Pickering Labs

Abstract:
The Agricultural Improvement Act (2018 Farm Bill) makes hemp production and distribution legal under Federal law. Hemp has a wide range of possible applications, including the production of fibers, paper, certain foods, supplements, cosmetics and even the recently FDA-approved drug Epidiolex. Legally available hemp and hemp products need to be tested for the presence of pesticides, heavy metals, residual solvents, and other harmful substances. Similarly, classified crops are routinely tested for Mycotoxins, including Aflatoxins and Ochratoxin A. Pickering Laboratories developed an easy and sensitive method to analyze Aflatoxins B1, B2, G1, G2 and Ochratoxin A in hemp and hemp-containing edible products. Mycotoxins are isolated using immunoaffinity clean-up columns and analyzed with fluorescence detection. To increase the sensitivity of Aflatoxins B1 and G1, an in-line photochemical reactor (UVE ) is installed before the detector. This method allows laboratories to easily determine these Mycotoxins at low ppb levels by HPLC and post-column derivatization.

Presenter:
Frank Sikora

Company:
University of Kentucky

Abstract:
A proficiency testing program for hemp was initiated in our Division of Regulatory Services in 2018. Hemp samples are ground to pass a 1 mm screen and homogenized. Two 10 g samples are sent to each laboratory in each shipment. Two shipments occurred in the fall of 2018 and 2019. Laboratories are asked to report their results and methods used for THC, THCA, total THC (THC + THCA x 0.877), CBD, CBDA, total CBD (CBD + CBDA x 0.877), CBN, and moisture in triplicate. Individualized reports are sent to each lab evaluating the lab’s precision and accuracy of their analyses. Summary reports, certificates of analysis, and a guide for interpreting reports are also available on the web at www.rs.uky.edu by clicking on Hemp PT in the menu bar. Data in the program is analyzed with robust statistics following the procedure on page 53 of ISO 13528:2015(E) to determine Z scores for each laboratory which is a measure of how close their result is to the robust mean. Precision of a lab’s triplicate results is evaluated with HorRat(r) values. The program currently has 67 laboratories in the program with 34 that are public and 33 that are private. The program is a valuable tool for laboratories to determine their measurement uncertainty and evaluate whether material is legal based on the 2018 Farm Bill definition of hemp being less than 0.3% THC on a dry weight basis. Data will be presented with a focus on total THC.

Presenter:
Sven Reister

Company:
Qiagen

Abstract:
The detection of pathogen contamination in cannabis samples is an essential part of product quality surveillance. Still, standardization and workflow integration are pertinent challenges in cannabis industry. Here we present a real time PCR-based workflow, that gives investigators a much faster and more robust tool for detection of pathogenic contamination compared to classic culture based typing. In this presentation we introduce different approaches for automated sample processing as well as data interpretation. In addition, we will provide insights into development of an pan aspergillus screening assay, as well as our custom assay development capabilities. In short a generic screening assay for 4 Aspergillus strains namely A.Fumigatus, A.terreus, A.Niger and A. Flavus was designed using the QIAGEN UCP (Ultra-Clean Production) Probe real-time PCR chemistry. These assays were multiplexed with FAM probes and combined with an internal control system labelled in HEX. All 4 assays show high linearity throughout a broad input range as well as a Limit of Detection <10 copies per reaction. Furthermore, we have validated these performance parameters in the presence of PCR inhibitors like Hematin. In order to ease real-time PCR data analysis an automated analysis template for QIAGEN’s operating and analysis software of the real-time cycler Rotor-Gene Q, Q-Rex was developed to enable automated and user unbiased analysis that is also available for detection of Salmonella and Shiga toxin producing E.Coli. The proposed workflow and methodologies will help to improve workflow robustness for microbial contamination testing and allow an easy transfer of methodologies to novice labs starting with molecular biologybased detection workflows.

Presenter:
Charles Johnson

Company:
Napro Research

Abstract:
As the popularity and variety of concentrated cannabis and hemp products on the market increases, so do the potential challenges associated with providing reliable pesticide analysis to ensure that the end user is consuming a safe product. The list of pesticides in many recent regulations for cannabis products can be extensive and vary greatly from location to location in terms of acceptable limits. Many action levels are in the part per billion range which creates great challenges for laboratories due to ion suppression and interference that occurs as a result of the matrix. The vast diversity of potential matrices requires a thorough understanding of how their components may interfere with analysis and how to overcome the analytical challenges they present based on matrix type. There are an abundance of application notes and publications describing validated methods for cannabis plant material but few are focused on more refined cannabis products. Refined cannabis and hemp oils, such as distillate, and the plethora of products formulated from these oils (eg. vape oils) may contain increased pesticide residue concentrations due to the extraction/manufacturing process. This work assesses the many analytical options that are available for sample preparation as well as different approaches for accomplishing an accurate quantitation of pesticides included in the Canadian and California regulations.

Presenter:
Geoffrey Rule

Company:
Millipore Sigma

Abstract:
Testing of cannabis for pesticide residues is mandated by regulations in many states and countries. The reporting limits and thresholds vary widely and in some cases are in the low ppb range. Cannabis, and derivative products, are challenging matrices for pesticide analysis due to high cannabinoid and terpene content, and a broad range of other possible constituents. An understanding of matrix suppression effects and selection of internal standards can aid in accurate quantitation of pesticides in cannabis and related products. Matrix effects resulting from the presence of high levels of cannabinoid compounds are studied using the tee-infusion methodology and results are used to suggest where particular isotopically labelled (SIL) pesticide internal standards may be most advantageous for quantitation. In addition, it is shown how issues related to calibration of individual pesticides and the manner in which validation studies are conducted may have inordinate influence on quantitation. We focus primarily on the requirements for pesticide testing and validation, calibration, regression, quantitation, and recovery. Using five different SIL pesticides, distributed across the chromatographic run, we illustrate important aspects of internal standard choice as they relate to matrix suppression effects, curve linearity, and dynamic range. Studies suggest an additional and often more challenging aspect of quantitation, that of sample clean-up and chromatography. We outline experiments for evaluation of both recovery and suppression effects of pesticides and illustrate how use of SIL internal standards can result in accurate quantitation despite poor recovery.

Presenter:
Douglas Duncan

Company:
Chem History

Abstract:
Consistent cannabinoid data is a critical indicator of laboratory performance and viability. ChemHistory has shown as much as a 40% relative difference between laboratories in the Oregon recreational market. This presentation shows how laboratory management uses low-cost analytics provided by Confident Cannabis to improve quality management systems. ChemHistory will present a “bird’s eye view” of data collected from recreational marijuana, medicinal marijuana, and industrial hemp Oregon markets. Using this data, we will show historical trends that are used to find anomalous data. Finally, ChemHistory will show 2019 Industrial hemp results evaluated using the new USDA THC action level. While few pre-harvest tests would fail USDA rules, a significant amount of post-harvest plant material and hemp products would fail.

Presenter:
Loy Jones

Company:
Agilent

Abstract:
Streamlining Cannabis Analysis by ICP-MS to Ensure Accuracy and Productivity with Enhanced Software Techniques ICP-MS is a proven instrument for reliable analysis of routine cannabis and cannabis related products for toxic metals and any additional metals of interest. The ability to obtain accurate results using unique software tools, as well as monitoring the associated analysis remotely allows ease of multi-tasking with sample preparation and other analytical techniques. Using advanced acquisition techniques and remote monitoring of the analysis assures precise and accurate data and that the cannabis samples are running smoothly to allow for a productive workflow

Presenter:
Elizabeth Almasi

Company:
Agilent

Abstract:
The importance of safety and quality testing of cannabis and hemp is paramount in order to satisfy both consumers’ need and regulatory requirements. The implementation of the testing protocols requires considerable time and substantial technical expertise. To reduce the time and effort needed to create optimized, fully functional analyses, we will show how eMethods allow to achieve analytical productivity faster, greatly reducing or eliminating the developmental effort. The eMethods deliver expert analytical protocols for potency, pesticide residue, mycotoxins, heavy metals, residual solvents and terpenes testing. Beside the complete instrumental and data handling methods, information on sample preparation and best practices are also provided along with a comprehensive list of all the necessary components needed for the application. The pesticide residue analysis workflow will be detailed as an example of an Agilent eMethod.

Presenter:
Roberto Federico-Perez

Company:
Milestone Inc.

Abstract:
Cannabis plant species are susceptible to the accumulation of heavy metals as part of their growth and metabolism. To guarantee consumer safety and meet rapidly expanding state requirements, trace metal analysis plays a significant role as a quality control parameter in the processing of cannabis and their derivatives. Sample preparation steps are critical to generate reliable results prior to the use of traditional spectroscopic methods for metals testing (ICP-MS/ICP-OES). Microwave digestion is a sample prep technique that provides high temperatures in short times to perform a full thermal decomposition of cannabis matrices. This process is assisted by the use of acids to completely solubilize metals in solution. As the industry grows, there is an ever present need to have sample prep solutions that offer the highest throughput to keep up with the demands of the lab. In this work, we present recovery studies on cannabis flower and derivative matrices that show a successful performance in terms of full analyte recovery (90-105%) and great reproducibility (%RSD < 3) for two microwave digestion approaches: traditional closed-vessel rotor-based systems and Single Reaction Chamber (SRC) technology. We will discuss the differences in operation and capabilities of these two techniques, and how to achieve the highest throughput with each system type.

Presenter:
Adam Floyd

Company:
Think 20 Labs

Abstract:
Modern interest in terpenes and terpene analysis has grown significantly as the recreational and medicinal use of cannabis has expanded across the globe. Terpenes are found in significant concentrations in cannabis. There is research that suggests terpenes provide a synergistic “entourage” effect, compounding the suggested medicinal benefits from THC and CBD usage. Terpenes are built from repeating five carbon units called isoprene. Their distinction as monoterpenes, sesquiterpenes, diterpenes, etc. is dependent on the number of isoprene units utilized in their biosynthesis. Because of this, the mass spectrum produced by terpenes share many common ions. The use of mass spectrometry allows for the separation and quantification of co-eluting peaks. This in turn allows for the rapid analysis of samples as chromatographic separation becomes less critical. Traditional use of FID (flame ionization detectors) demands complete chromatographic separation and extended run times. In order to fully utilize the benefits of mass spectrometry in this complex analysis careful consideration must be taken in selection quantitation and qualifier ions. The objective of this study was to produce a fast, accurate, and precise method for analyzing terpenes in cannabis products using gas chromatography/mass spectrometry. There are over 200 terpenes found in cannabis, developing a method to analyze such an extensive list of analytes is not reasonable for a production laboratory. Speed and precision are critical in production laboratories, thus developing methodologies that meet these requirements are critical. This method focuses on the most common terpenes found in cannabis.

Presenter:
Charles Harb

Company:
RingIR

Abstract:
In the last decade we have seen an enormous increase in the knowledge around cannabis. The traditional view of THC as only active ingredient has shifted toward a consideration of all components and how they interact. The differences in the effects that various cultivars produce is attributed to this and often referred to as the entourage effect. Most of the phytochemical variation in cannabis is found in a class of secondary metabolites called terpenes. The presented research will demonstrate a novel technique that can quickly and reliable quantify many terpenes at once and will help create a better understanding of the differences between cultivars.

Presenter:
Barbara Fisher

Company:
Green Thumb Industries

Abstract:
One of the most important aspects of medical cannabis is testing. Cultivators, processors, and manufacturers of cannabis infused products rely on independent testers to accurately convey potency information to the consumer. Today, there are no agencies holding independent testing labs accountable and/or authenticating their data. Therefore, it’s essential to have internal testing to validate. Here we elucidate the importance of labeling HPLC peaks in order to prevent false identification of cannabinoids, Cannabicyclol (CBL), degradation product, of cannabichromene (CBC), and delta10-THC. Delta10-THC is an isomer of delta9-THC and can easily be confused with other cannabinoids without monitoring retention time shifts and calibrating on a regular basis. Emerging synthetic chemistry encourages method modification and the discovery of more rare cannabinoids. With no functional guidelines to follow and labs holding their methods proprietary to be competitive in a federally illegal climate, transparency needs to be encouraged and embraced.

Presenter:
Rodney Nash

Company:
Omni International, Inc.

Abstract:
Chronic pain is a growing epidemic and public health issue that continue to affect the aging population in the US. Managing pain is particularly difficult with patients who suffer from cancer-associated pain, neuropathic pain, as well central pain states. These conditions are typically treated inadequately with opioids, antidepressants, and anticonvulsive drugs. There is growing evidence that cannabinoids natural products are beneficial for a range of clinical conditions, including pain and inflammations. The use of cannabinoids in highly regulated prescription drugs is fast approaching and the development of such drugs requires well planned medical trials as well as very stringent guidelines for contaminants such as heavy metals, pesticides, mycotoxins, etc. However, federal guidelines have not kept up with this emerging industry. Currently, each state is left to manage their own set of regulations, resulting in an industry that has great uncertainty and chaos as it attempts to satisfy the growing demand for its products. Here we demonstrates in detail the importance of sample homogenization and how this critical process can influence potency quantitation of cannabinoids. Here we demonstrate a fast and simple analytical approach to homogenization and sample extraction to quantify 11 cannabinoids using HPLC. Using the Omni Bead Ruptor-96 we demonstrate the impact that homogenization has on the determination of cannabinoid potency. Potency is greatly impacted by the homogeneity of the sample, and the size (in microns) of each particle. Here we show the Omni Bead Ruptor-96 (BR-96) can consistently produce a homogenate that is ideal for measuring cannabinoid

Presenter:
Brittany Leffler

Company:
CEM Corporation

Abstract:
With the legalization of hemp in the 2018 Farm Bill there is a need to confirm that the product is indeed hemp, and meets the criteria of low THC. Having a harmonized method to accurately determine the cannabinoid content in hemp products is important to this industry. The extraction of cannabinoids from hemp is challenging due to the easy conversion of THCA to THC. Existing techniques do not offer a method that is rapid, simple, and efficient. In this study, a new extraction system, the EDGE, combines the processes of pressurized fluid extraction and dispersive solid phase extraction is explored. This new method offers efficient extraction of cannabinoids from hemp in less than 10 minutes in one simple process. Different analytes beyond cannabinoids, such as pesticides and mycotoxins can also be extracted with this new method. The EDGE method offers a rapid, simple, and efficient solution for hemp testing.

Presenter:
Andrew Defries

Company:
Rosales Consulting Group

Abstract:
Secondary metabolites that we collectively know as Cannabinoids are organic small molecules biosnthesized from isoprenoid building blocks in Cannabis. Our casual use of the term cannabinoid(s) and exclusive association of these molecules with cannabis has blind sided our exploration and exploitation of the diversity of molecules available from medicinal plants. Natural products derived from non-cannabis plants that impinge on the Endo Cannabinoid System (ECS) have been coined as phyto-cannabinoids. Through the use of chemical informatics (cheminformatics) we have taken inventory of the physicochemical properties from a myriad of cannabinoids and phyto-cannabinoids cited in published literature. These physicochemical properties were used to create several models to provide insight into the extraction, separation and isolation of pure compounds from botanical starting material. Similarities and differences in the biosynthetic routes of diverse cannabinoids and phyto-cannabinoids give us leads for the cultivation, breeding, and genetics of cannabis and non-cannabis plants to be used in formulations directed at the ECS. We will highlight several areas of opportunity in these areas.

Presenter:
James Brennan

Company:
LabWare

Abstract:
The cannabis industry faces many challenges, including highly detailed testing, varied regulatory scrutiny, and financial pressure as the market grows increasingly competitive. Cannabis testing laboratories demand sophisticated, enterprise-level informatics solutions that support compliance. This presentation reviews obstacles to efficient data management facing cannabis laboratories and examines options available to those evaluating laboratory informatics systems. These include in-house developed systems and commercial laboratory software, such as LIMS, ELN, QMS, and mobile apps.

Presenter:
Oliver Buettel

Company:
Analytik Jena

Abstract:
“Vaping” of cannabis concentrates via electronic vaporizers is the method preferred by more than 30% of cannabis users. Recently, the use of vape cartridges has been associated with severe lung disease, probably caused by contaminations introduced during illicit manufacture of black market products. This study evaluates the release of toxic metals from vape cartridges obtained from various sources, and their impact on the quality of cannabis products consumed.

Presenter:
John Mills

Company:
BioMerieux

Abstract:
Rapid Salmonella spp. and STEC detection in cannabis products presents unique challenges. The industry is continually seeking faster time to result, further challenging the limits of available methodologies. There is a significant need for rapid and robust detection methods to decrease the risk of inaccurate results and optimization to combat naturally occurring PCR-inhibitory properties. For this study, the SLM (Salmonella spp.) and EH1 (STX/EAE) GENE-UP assays were evaluated with an artificial contamination study of multiple cannabis matrices. This study also included some enrichment optimization to mitigate PCR inhibition. Eight cannabis or cannabis related products were inoculated with two levels of both a Salmonella spp. and Escherichia coli O111 to determine the performance of the GENE-UP assay for the detection of these target organisms in cannabis products. For the low level, <2 CFU/test portion of each organism was spiked into each product. Approximately 50 CFU/test portion was inoculated into all products for the high level. To investigate exclusivity, a competitive (non-target) inoculation was also investigated. The GENE-UP® Salmonella (SLM) and STEC (EH1) assays demonstrated equivalency to the confirmation plating methods for the detection of both Salmonella spp and STECS for all products evaluated. These data validate that the GENE-UP system is an acceptable method for screening cannabis flower and THC infused products for the presence of Salmonella spp and STECS.

Presenter:
Niranjan Aryal

Company:
Certus Analytics

Abstract:
Distribution of cannabinoids across cannabis samples Mike Tunis, Niranjan Aryal* Certus Analytics, 26359 Jefferson Ave Suite G, Murrieta, CA 92562 *Corresponding author: [email protected] With the legalization of cannabis in several countries and different states of the United States, enthusiasm for the knowledge of chemistry on the cannabis flowers and different cannabis-derived products has increased. Potency, which is attributed by the cannabinoid contents, has attracted both recreational and medicinal cannabis community. While about 100 of more than 400 secondary metabolites produced by the Cannabis plant are categorized as the cannabinoids, the understandings of their chemical structures, and biosynthetic pathways are still evolving. We measured and analyzed 10 major cannabinoids in flowers and cannabis-derived products from various sources; farmers, distributors, extractors and cannabis-products manufacturer. Correlation study for the cannabinoids from more than 100 Marijuana flower samples showed that THCA is positively correlated to CBGA, and the THCA derivatives such as delta 8 and 9 THC are positively correlated to THCA. Interestingly, THCV was positively correlated to delta 8 THC (0.959 Pearson correlation). This may be a result of feedback regulation in the cannabinoid biosynthetic pathway. However, there requires analysis of much larger dataset and relative gene-expression studies before concluding. We also built a principal component analysis for cannabinoid data obtained from analyzing about 850 samples including concentrates and other edible products. CBG, THCV and CBG formed a component while CBDA, THCA, CBGA, CBC and delta 8 and 9 THC formed another component. Interestingly CBN formed a third component staying equidistant from both first and second component. This may be because of the tendency of a cannabinoid to co-elute with the other during the extraction process, given that there was no addition of synthesized cannabinoids. Establishment of clear chemical structure for major and minor cannabinoids and analysis of mega dataset across testing labs may lead us towards more conclusive answers.

Presenter:
Neya Jourabchian

Company:
CSA LA, LLC

Abstract:
As the cannabis industry grows, so does the amount of data that is being collected by the third party laboratories responsible for analyzing pesticides and other contaminants in a myriad of matrices being introduced daily. With sophisticated instrumentation ranging from high performance liquid chromatography to tandem mass spectrometry, the data being acquired has proved invaluable in analyzing trends in the industry. CannaSafe, which analyzes a quarter to a third of the products being sold in California, will show trending data on most frequently encountered pesticides from the R&D and Compliance markets, spanning the different counties and regions of the state. Additionally, we will elucidate on potential dangers, degradation products, and the importance of instrument method development to support accurate analyses.

Presenter:
Nikhil Kumar

Company:
Canalysis Laboratories

Abstract:
Microbial pathogen testing of cannabis is a rapidly evolving aspect of regulatory testing. An increasing number of jurisdictions requires compliance testing to include detection of four species of Aspergillus (A. flavus, A. fumigatus, A. niger, and A. terreus) that can cause disease in individuals with underlying health issues. Pulmonary aspergillosis, an invasive lung disease, has been reported in immunocompromised medical cannabis patients, with multiple cases resulting in death despite antifungal interventions. In some patients, a few inhaled Aspergillus spores can lead to irreversible organ damage. Therefore, it is crucial that the detection system for Aspergillus must be robust, specific, and highly sensitive. Here, we examine modifications to standard qPCR workflows that third-party testing labs can implement for accurate and rapid detection of Aspergillus in cannabis. A method is described that incorporates Bio-Rad’s Free DNA Removal Solution into the iQ-Check Aspergillus Real-Time PCR Kit, resulting in the degradation of free DNA from dead or lysed Aspergillus. Further, a comparison is made across samples spiked with low levels of Aspergillus and enriched for either 24 or 48 hours, demonstrating the benefits of a 48-hour enrichment time. Together, these implementations can be added onto existing qPCR workflows to improve enrichment and detection of Aspergillus.

Presenter:
Colton Myers

Company:
Restek

Abstract:
Terpenes are a class of organic compounds that are comprised of isoprene units based on 5 carbons. These compounds are produced by a variety of plants and generally have a strong odor. Different terpenes, and ratios of terpenes, can be found in different cannabis chemovars. Not only are terpenes identified in cannabis for their aromas and flavors, but they may also play a major role in the entourage effect. The entourage effect is believed to be created by the synergistic interaction between cannabinoids, terpenes, and other naturally occurring compounds, which gives each cannabis chemovar its own unique experience. This research is focused on evaluating injection techniques for analyzing terpenes via gas chromatography – mass spectrometry (GC-MS). These injection techniques include Headspace-Syringe (HS-Syringe), HS-Solid Phase Microextraction (HS-SPME), Direct Immersion-SPME (DI-SPME), and Liquid Injection (LI).

Presenter:
Melissa Chandler

Company:
UCT

Abstract:
With the mounting interest in hemp and cannabis products for medicinal and recreational use around the world, the need for suitable analytical methods to identify and determine the concentration of cannabinoids is essential for ensuring consumer safety. Traditional analyses for measuring the potency of cannabinoids in cannabis and hemp samples have focused mainly on 5 primary analytes: THC, THC-A, CBD, CBD-A, and CBN. As the industry continues to expand and evolve, more attention is being directed towards additional, although less prevalent, cannabinoids that have been shown to exhibit physiological effects. Thus, simple yet selective analytical methodologies are needed to adequately separate and identify a wide range of cannabinoids present in the hemp and cannabis products which exist on the market, including plant products, edibles, oils, topicals and extracts. This poster will outline a simple and robust method for the detection and quantitation of 16 cannabinoids in hemp and cannabis samples using an isocratic HPLC method coupled with UV detection. Baseline separation of all 16 cannabinoids, including that of the critical pair Δ9-THC and Δ8-THC, was successfully achieved using a Selectra® C18 column. Examples of hemp flower and oil samples analyzed using the analytical method will also be presented.

Presenter:
Geoffrey Faden

Company:
MAC-MOD Analytical

Abstract:
Cannabis samples are complex and contain several target compound classes that are of interest from analytical and regulatory perspectives. Cannabinoids and terpenes are major components of Cannabis flowers and are therefore of primary importance for pharmacological benefits. Regulatory requirements for testing are varied and may require determination of several key cannabinoids, for example1: ∆9-Tetrahydrocannabinol (∆9-THC), Cannabidiol (CBD), ∆9-Tetrahydrocannabinolic acid A (THCA-A), Cannabidiolic acid (CBDA), Cannabigerol (CBG), Cannabinol (CBN). Terpenes are another target class of interest due to potential synergistic effects with cannabinoids and potential use for fingerprinting cultivars. This poster summarises work performed to develop LC-MS compatible methods for the analysis of an extended list of 10 cannabinoids of interest and for fingerprinting terpene content. This work is to identify quick method development strategies to enhance method development activities and can support new cannabinoids and terpenes that may be of interest and added to the current list of major compounds.

Presenter:
Douglas Stevens

Company:
Waters Corporation

Abstract:
Terpenes are among the more abundant compounds produced by hemp. These compounds play a variety of roles including deterring herbivores or attracting pollinating species. They are largely responsible for the aroma profile of the bulk plant material and have been shown to contribute to physiological responses associated with the use of hemp-based consumer products. As such terpenes play an important role in the quality of products derived from hemp. They are also relatively volatile and can be effected by the type of processing used to extract them as well as in shipment and during storage. Therefore, it is important to quantify the terpenes present in hemp and related products in order to assess the quality and value of the plant material. In this work a simple microextraction of plant material, followed by liquid injection into an EI GC/MS/MS system was used to assess the potential advantages provided by this instrument configuration and workflow. The use of GC/MS/MS was proposed for terpenes analysis due to its common use for residual pesticide analysis. The goal is to perform the trace analysis of residual pesticides to ensure the safety of hemp as well as to analyze for terpenes to monitor the quality of the product all on a single instrument. Among the method elements investigated were the chromatographic separation and multiplexed mass spectral acquisition modes. Method development examples will be included as well as information on prospective improvements for routine use in future methods. Quantitative results will also be presented.

Presenter:
Stephanie Moon

Company:
Evio Labs

Abstract:
The increasing demands of the hemp and cannabis production industry on testing laboratories has created a demand for a robust, reliable, and accurate quantitation method for a larger number of cannabinoid analytes compared to the previous needs of the industry. In Oregon, more stringent ODA regulations on THC-A and D9-THC concentrations for pre-harvest hemp compliance testing has influenced the hemp market toward production of flower strains that produce the cannabinoid CBG-A over strains that produce CBD-A. CBG-A dominant strains have shown lower THC concentrations compared to CBD-A dominant strains. This HPLC method was developed on the Shimadzu 2030 Prominence-I using a PDA detector. The mobile phase is acidified methanol and water at a flow rate of 0.8ml/min. Methanol is a cheaper, greener solvent than acetonitrile or other strong organic solvent mobile phase options. Another advantage to methanol mobile phase is a clean baseline.The analytical range of the method is 0.5ppm-600ppm for THC-A, D9THC, CBD-A, CBD, CBG-A, and CBG and 0.5ppm-100ppm for CBC, CBN, CBD-VA, and CBD-V. This method uses high pressure achieved using a 2.1 mm column to elute 10 analytes in just over 10 minutes. The entire method is 13 minutes which includes a 1 minute 100% methanol clean out period and a 1 minute starting mobile phase equilibrium period. This method allows for reliable quantification and identification of 10 analytes in a variety of matrices including cannabis, hemp, concentrates, extracts, topicals, and edible products. This method has proven its efficacy and reliability during the harvest season of 2019.

Presenter:
Michael Miller

Company:
Cayman Chemical

Abstract:
High-quality reference materials with established metrological traceability are an essential element for measurement accuracy during routine analytical checks as well as for cannabis product quality testing and profiling. A Certified Reference Material (CRM), produced by an ISO 17034 accredited Reference Material Producer, provides the metrological traceability, homogeneity, and stability to validate its overall accuracy. Single analyte CRMs are typically the go-to reference when preparing a calibration and control standards for testing by various chromatographic methods. However, complications can arise from preparing stock standards from single component CRMs, which may result in traceability and measurement accuracy being compromised. A multi-component CRM mixture was designed containing the ten most prevalent phytocannabinoids found in Cannabis plants: cannabidivarin (CBDV), cannabidiolic acid (CBDA), cannabigerolic acid (CBGA), cannabigerol (CBG), cannabidiol (CBD), cannabinol (CBN), tetrahydrocannabinolic acid A (THCA-A), Δ9-tetrahydrocannabinol (Δ9-THC), Δ8-tetrahydrocannabinol (Δ8-THC), and (±)-cannabichromene (CBC). A pre-made, multi-component CRM bypasses the additional steps required to prepare in-house multi-component stock solutions using single component CRMs. A comparison of the concentration values from the multi-component CRM solution against an in-house, multi-component solution prepared using single component CRMs demonstrates that a premade, multi-component CRM solution provides more accurate concentrations compared to stock standards created using single component CRMs.

Presenter:
Jacob Jalali

Company:
PerkinElmer

Abstract:
Cannabis oil is a concentrated extract from cannabis sativa plant. Pesticide analysis in cannabis oil, has been a challenge in past few months due to high percentage of cannabinoids in matrices and heavy ion suppression. Recent changes in regulatory guidelines for the quality control of medicinal cannabis and cannabis for recreational use has highlighted some interesting analytical challenges in the analysis of pesticides in cannabis oil. This study will detail an analytical UHPLC-MS-MS method for low ppb level quantitation of CA regulated pesticides in cannabis oil. It is anticipated that this study will help support method development efforts towards improved robustness and quantitative accuracy in this new and emerging industry. In this research, cannabis oil with spiked deuterated internal standards, is extracted with acetonitrile and after filtration, supernatant is transferred to HPLC vial for LC-MS-MS analysis. A mix of 33 deuterated internal standards were used across the run for correction of any matrix effect and achieving a good recovery for pesticides. PerkinElmer Quasar SPP Pesticides HPLC column (100×4.6mm, 2.7um) is used for separation of pesticides. Cannabis oil samples were prepared and analyzed at Napro Research (Sacramento, CA). The UHPLC for this investigation was a PerkinElmer LX-50. The MS/MS instrument was a QSight 420. Investigating mobile phase solvents, flow rates, injection volume, peak shapes, injection modes, ionization source parameters and cannabis strain-base product ion scans, were part of this research. Good accuracy, recovery and long-term stability were found on spiked oil samples, by using this analytical method.

Presenter:
Christopher Pauli

Company:
Frontrange Biosciences

Abstract:
Genotyping methods for Cannabis have advanced greatly in the last few years, with marker assisted breeding and genotyping by sequencing being most commonly used. This poster will focus on applying a method of variant identification analysis, referred to as high resolution melting (HRM), which is currently being used to guide the breeding of our hemp population through identifying the genotypic diversity in various crosses and parental lines. We will explain how the process of HRM analysis works and some background on the development work that went into creating these breeding tools. With the genotypes of various genes involved in cannabinoid and terpenoid biosynthesis identified via HRM, we are applying this knowledge to rapidly breed new varieties and unique combinations of traits through genetic selection. The markers presented will focus on the total accumulation of cannabinoids and the ratios of those cannabinoids, as well as some discussion towards our developing markers to build on our nascent understanding of terpene biosynthesis. In addition to the data presented on the individual markers, we will show how understanding the combination of genotypes from multiple markers can be applied as a group to work as a fingerprint of Cannabis, that can also be applied in other various breeding applications.

Presenter:
Saba Hariri

Company:
PerkinElmer

Abstract:
Although, medicinal use of cannabis was legalized since 2001, acquisition, cultivation, possession and consumption of cannabis only became legal in 2018 under the federal Cannabis Act. Owing to the excessive use of chemicals during cannabis plant cultivation, Health Canada issued regulatory acceptable limits for pesticides, residual organic solvents, among others. We developed a fully validated analytical sample preparation method with a PerkinElmer liquid chromatography (LX50) coupled with the tandem mass spectrometer for a complete analysis of all 96 pesticides outlined in the Health Canada regulations for cannabis. A developed method on the PerkinElmer QSight 420™ series was used to accurately quantify all 96 pesticides, including such compounds like cyfluthrin, kinoprene, and quintozene which are conventionally analyzed by gas chromatography were all detected on this single platform, PerkinElmer LC-MS/MS system. The method entailed the use of selected deuterated pesticide standards to correct for MS signal enhancements or suppressions, annulling any false positives. LOQs recorded for all pesticides were less than 20%, including those typically analyzed by gas chromatography, exceeded Health Canada action limits for pesticides in cannabis flower. Pesticides percent recoveries from the cannabis matrix, at 2 different concentration levels, were within 70-120%. The use of a single analytical LC-MS/MS platform invalidates the need for the use of a GC-MS/MS and therefore provides an effective cost–saving opportunity for research scientists and laboratories engaged with pesticides analysis in cannabis, in general.

Presenter:
Teresa Moreno

Company:
Callaghan Innovation

Abstract:
Cannabinoid decarboxylation is a non-enzymatic process by which the naturally occurring acidic forms of cannabinoids such as THCA, CBGA or CBDA are converted into the neutral compounds (THC, CBG or CBD) which are the preferred form for many cannabis derived products. Decarboxylation is a relatively slow process that occurs naturally in the plant, but can be accelerated by exposure to heat, light and oxygen. Industrially, this is easily performed by heating the plant material in an oven at temperatures above about 120°C. Kinetically, this is a complex reaction system with interactions between the different cannabinoids that requires the right balance between reaction temperature and time to achieve the desired conversion while minimising degradation of neutral cannabinoids. Moreover, the loss of moisture occurring during the process, coupled with the formation of neutral cannabinoid forms that are more readily soluble in non-polar solvents such as CO2, will impact the subsequent extraction and affect the yield and composition of the resin obtained. In this work, the effect of several parameters (i.e. temperature, time, presence of oxygen and amount of plant material) on the decarboxylation of THCA, CBDA and CBGA in hemp has been investigated. Reaction kinetics have been studied using different models of increasing complexity: from a simple approach considering only the three decarboxylation reactions separately, to a more complex model which accounts for the further conversion and loss of cannabinoids observed at high temperatures. The difference between these models has been analysed, and a comparison with previously reported results has been performed. CO2 extraction of decarboxylated and non-decarboxylated plant material was carried out and the effect on yield and composition of the extract is discussed.

Presenter:
Brent Trela

Company:
Ball Corp.

Abstract:
Beverages containing cannabinoids such as tetrahydrocannabinol (THC) and/or cannabidiol (CBD) are rapidly growing segments in marijuana and ready to drink (RTD) markets. It has been suggested that certain container materials may decrease cannabinoid potency in the beverages which they contain. Cannabinoid concentration is either regulated to be within a certain range and/or important to truth in labeling statements and consumer confidence. This work measured the concentration over time of several aqueous preparations of emulsified CBD and THC during storage in glass and containers with various surface coating materials, including aluminum beverage cans. Among cannabinoid preparations, beverage matrix and storage conditions, there were significant differences. There were generally little to no significant differences in stability among the coating materials and glass, although there were a few exceptions. If claims are made of potency losses attributed to certain packaging materials, it should be in reference to controls stored under similar conditions of time, temperature, O2 and UV exposure, etc.

Presenter:
Wade Bontempo

Company:
SepSolve Analytical

Abstract:
Laboratories seeking to maximise productivity in the analysis of cannabis products face numerous challenges due to their complex matrices containing hundreds of volatile organic compounds (VOCs) spanning many orders of magnitude in concentration. Most laboratories adopt one-dimensional gas chromatography coupled with mass spectrometry (GC-MS) in an attempt to tackle this challenge. However, this does not provide adequate separation, resulting in numerous coelutions which interfere with the compounds of interest and cause difficulties in quantitation. Furthermore, separate methods and/or systems are usually required for the analysis of specific target compounds – e.g. residual solvents or terpenes. Here, we describe a combined approach for the analysis of residual solvents and terpenes using a flexible analytical system. The system utilizes gas chromatography with parallel detection by flame ionization detection and time-of-flight mass spectrometry, as well as an optional secondary separation (or GC×GC) for complex samples in need of additional resolving power. We will demonstrate how this approach can not only maximise productivity, but also improve data quality.

Presenter:
Gregory Matthews

Company:
High Grade Hemp Seed, Inc.

Abstract:
As cannabis becomes mainstream, new and more sophisticated applications of biotechnology will come be incorporated into breeding programs. Traditionally, cannabis breeders have simply flowered their males and females at approximately the same time and let nature take care of the rest. Now that more serious agronomy is involved, there are multi-year breeding projects being undertaken and long term pollen and seed storage are becoming necessary. How can we ensure the integrity of old or improperly stored pollen or seeds? A chemical solution that is in use in other agricultural crops, 2,3,5- triphenyltetrazolium chloride (TTC), is also applicable to the Cannabis plant. Demonstrated here is an easy and quick method for using TTC to determine viability. An obvious color change of clear to red signifies that the material is performing metabolic processes and is therefore alive. This technology has shown clear results and will have an impact on cannabis breeding programs by ensuring the viability of stored pollen and seed.

Presenter:
Thu Huynh

Company:
Hygiena

Abstract:
The growing cannabis industry has resulted in a proliferation of state regulations to ensure the quality and safety of cannabis products. Cannabis suppliers must label the level of total tetrahydrocannabinol (THC) and cannabidiol (CBD) in their products. Additionally, hemp suppliers in the United States must show their products contain <0.3% THC. Several methods for potency measurement have been developed, including high-performance liquid chromatography (HPLC). However, there are drawbacks. For instance, HPLC is time intensive, requires sample clean-up, advanced training, and can be costly. We developed an enzyme-linked immunoassay (ELISA) capable of detecting the psychoactive compound, D9-tetrahydrocannabinol (D9-THC) from cannabis and hemp products. The D9-THC ELISA measures %THC in a variety of products by altering the dilution to accurately measure the broad range of THC levels among disparate sample types. Validation studies were performed on market products and spiked materials, and the %THC recovered was 85-113%. A D9-THC proficiency sample was measured to contain 390.218 μg/mL, which was within the acceptable range of 291-540 μg/mL. A hemp oil proficiency sample was also tested and shown to contain 0.6989 mg/mL, which was within the acceptable range of 0.612-1.14 mg/mL. This assay may be useful to the cannabis and hemp value chain from growers to manufacturers of edibles who need rapid, cost-effective screening methods for measuring D9-THC concentration.

Presenter:
Jessica Westland

Company:
Agilent

Abstract:
Food products containing cannabis extract (edibles) have emerged as a popular option of cannabis use in the legalized market. While gummies are one of the most sought after cannabis edibles, not all gummies are the same. Varying factors in ingredients, recipes, and even production materials make a difference in the final product. Oregon state regulates that all cannabis flower sold must be screened for about 60 pesticides commonly used in cannabis cultivation (along with potency levels). Edibles, on the other hand, undergo various lab tests first as a flower and then as a butter. As regulations expand to include the end product (the gummy itself), the need for an efficient, reliable, and cost-effective sample preparation method is needed. The commonly utilized QuEChERS sample preparation method was compared to pass through SPE protocols with different chemistries to investigate the extraction of pesticides from a cannabis infused gummy in the state of Oregon.

Presenter:
Jessie McGinn

Company:
AltaSci Labs

Abstract:
The need for precise quantitation of pesticide residues in medicinal cannabis is vital for ensuring patients are not subjected to inhaling toxic compounds, the side effects of which are largely unstudied and unknown. However, the combination of a complex matrix and low action limits (≤ 10 µg/kg) present a significant challenge for analysts looking to quantitate pesticides in cannabis flower. In particular, great care must be taken during the sample preparation process as certain pesticides (e.g. spinosad) can be lost at this step and lead to low recoveries. This can be detrimental to patient safety as it could lead to an underestimation of the level of pesticides present in the sample or even a false negative result with potentially harmful consequences. Despite these challenges an innovate dispersive solid-phase extraction (dSPE) product produced by United Chemical Technologies has helped to remove challenging matrix interference while preserving the pesticides in the sample when used as part of a QuEChERS sample preparation procedure. In combination with LC-MS/MS analysis our method is capable of detecting pesticides in cannabis flower with excellent recoveries (70-120%) and low repeatability/reproducibility (≤ 20%) while being able to detect pesticides well below the action limits (LOD = 1µg/kg). The use of the novel dSPE cleanup product and corresponding extraction procedure has helped to ensure that our lab results for the regulated pesticides in medicinal cannabis flower is accurate, conforms to Connecticut testing standards, and ultimately helps to ensure patient safety.

Presenter:
Chris Leija

Company:
Millipore Sigma

Abstract:
Cannabinoid potency and profiling is critical to the quality control of cannabis and hemp products. Defined testing and certification are required in many states at the federal level for hemp. Dozens of HPLC-UV methods have been proposed for the analysis cannabinoids in hemp and cannabis. However, many of these methods offer limited characterization of major cannabinoids, result in high cost per analysis, or require high-end HPLC systems. To address these shortcomings, we have developed a complete cannabinoid workflow that prioritizes low cost analysis, broad compatibility, and characterization of all major cannabinoids. Both gradient and isocratic methods were developed on Supelco Ascentis Express Fused Core columns using cannabinoid solutions that have been certified as ISO 17034 Certified Reference Materials (CRMs). 17 cannabinoids were separated using isocratic conditions on the Ascentis Express C8 (150x3mm, 2.7um) in under 10 minutes. A shorter run time of 7 minutes was achieved by incorporating a simple gradient on the Ascentis Express C18 (150x3mm, 2.7um). To further reduce costs, a methanol-based method was developed that resolves 14 cannabinoids on the Chromolith HR RP-18 (50 x 2mm) column. The unique mesopore and macropore structure of monolithic HPLC columns make it well suited for handling complex matrices and significantly extends column lifetimes. This cannabinoid workflow offers a streamlined solution for sample preparation, preparation of standards, and chromatographic separation.

Presenter:
Melissa Wilcox

Company:
Regis Technologies

Abstract:
In recent years, there has been a growing interest in the development of therapies and other consumer products derived from cannabis and hemp, including cannabidiol (CBD). With the passage of the US 2018 Farm Bill, hemp was removed from the Controlled Substances Act, which means that cannabis plants and derivatives that contain no more than 0.3 percent tetrahydrocannabinol (THC) on a dry weight basis are no longer controlled substances under federal law. The small amount of THC found naturally in hemp plants can sometimes exceed the legal limit of 0.3% when extracted and concentrated along with CBD. Therefore, it is important to have a means of removing THC from CBD extracts in order to comply with Federal regulations. Herein, we demonstrate the ability of supercritical fluid chromatography (SFC) to separate CBD and THC with two different column stationary phases, 2-ethylpyridine and 4-ethylpyridine. Simple changes in mobile phase composition can switch the elution order of these compounds, providing flexibility for both analysis and purification. 4-ethylpyridine is also shown to resolve CBD and THC from their inactive, acid forms.

Presenter:
Ashlee Gerardi

Company:
Restek

Abstract:
The LC-UV Analysis of 19 Cannabinoids of Interest in Cannabis Products More than 100 cannabinoids have been isolated from cannabis in addition to the five most commonly tested: THC, THCA, CBD, CBDA, and CBN. While methods have been published that show the separation of these major cannabinoids, many do not take into account the possibility of interference from other cannabinoids that may be present. This is most problematic in concentrates where minor cannabinoids can be enriched to detectable levels that were not observed in the flower. In this study, the LC-UV separation of 19 cannabinoids of interest was performed while monitoring for the potential impact from minor cannabinoids. The method was originally developed on a Raptor ARC-18 150 x 4.6 mm, 2.7 µm column, but was also translated to two other column dimensions: 150 x 2.1 mm, 2.7 µm and 100 x 3.0 mm, 1.8 µm. The ability to use multiple column dimensions enables a fit-for-purpose method that can highlight one or more of the following benefits: reduced solvent consumption, increased sample throughput, MS compatibility, increased resolution, or increased sensitivity. The method was applied to cannabis products such as concentrates, oils, edibles, and personal care products.

Presenter:
Jeremy Melanson

Company:
National Resource Council Canada

Abstract:
A growing list of countries have thriving medicinal cannabis programs and countries such as Canada have legalized cannabis for recreational purposes. Most regulated markets have stringent testing requirements to ensure safety of consumers. Arguably the most challenging requirement is the pesticide test, and the list of pesticides to be tested for varies by State and Country. For instance, the Canadian regulations require mandatory testing for 96 pesticides with method limits of quantitation (LOQ) down to 10 ppb. This generally requires a combination of liquid chromatography – tandem mass spectrometry (LC-MS/MS) and gas chromatography – tandem mass spectrometry (GC-MS/MS) to achieve sufficient sensitivity over the wide range of pesticides. While measuring most pesticides at the low-ppb level with this instrumentation is relatively straightforward in pure solution, the high complexity of the cannabis extract combined with the roughly 7 order of magnitude excess of cannabinoids makes this test very challenging. In the case of LC-MS/MS, a co-eluting major cannabinoid will suppress virtually all signal for poorly ionizing pesticides. This presentation will describe the method development and validation of GC-MS/MS and LC-MS/MS methods for pesticides in dried cannabis. Using matrix-matched calibration to compensate for matrix effects, the methods achieve the LOQs for the 96 pesticides listed in the Canadian regulations. The need to adjust chromatography to shift poorly ionizing pesticides peaks from those of major cannabinoids will be highlighted. While these methods are designed to meet Canadian regulatory requirements, the methods are applicable to several State’s testing requirements that generally have higher pesticide limits.

Presenter:
James Laucius

Company:
High Purity Natural Products

Abstract:
The surface area of unfertilized female flowers of Cannabis sativa is covered with trichomes, which are the site of biosynthesis of cannabinoids and terpenes. Terpenes are volatile hydrocarbons that confer the characteristic aroma to C. sativa. Anecdotal evidence indicates that some terpenes may have therapeutic effects when consumed with cannabinoids; the so-called “entourage effect” suggests that terpenes work synergistically with cannabinoids to produce a psychoactive effect different from one caused by ingesting tetrahydrocannabinol alone. Consumers purchase dried Cannabis flower products not only by their cannabinoid content, but also by their aroma, which can vary greatly among different strains. As such, it has become an industry standard to analyze the cannabinoid and terpene profiles of legal cannabis products. There is little peer-reviewed literature on C. sativa terpene analysis and as the legal industry grows, there is an increasing need for more efficient and higher throughput protocols to meet the demand. Therefore, the objective of this work was to develop a method of analysis using gas chromatography mass spectrometry that could maintain or exceed the level of sensitivity that existing protocols offered while significantly decreasing the runtime to increase throughput. Female clones of Cannabis sativa ‘Wife’ were grown to maturity, and apical flowers were harvested and prepared for extraction. The developed method produced excellent separation of different terpenes allowing for quantification while significantly reducing runtime compared to peer-reviewed protocols.

Last Year's Conference Poster Presentations

Presenter(s): Laura Breit

Root Engineers

How can you optimize your grow while balancing needs for infrastructure optimization and the cost of technology? As pressure increases in the industry for efficiencies and cost savings, it is important for both growers and investors to have a strategy that focuses on both upfront costs and long-term operation effects. This poster focuses on optimizing some of your largest and most critical investments: heating, ventilation and air conditioning (HVAC) technologies. By providing a knowledgeable look at the technologies accessible all sizes of operations, and describing the relative economics for various systems, we will analyze profitability impacts as they relate to your business. Brought to life by real-world case studies, plus a discussion of codes and regulations that have an impact on decision making, attendees will leave this discussion with practical knowledge to apply to the technology selection process.

Presenter(s): Laura Breit

Root Engineers

As cannabis flower consumption falls to less than 50 percent of the market share in North America, it is clear that a number of consumers prefer the ease and convenience of extracts and edibles. Cannabis businesses preparing for the future of the industry – driven by changing consumer tastes – are finding that the right design to support their extraction operation is increasingly critical to business performance. A wrong step in the early development stages can be extremely costly, especially if operations are facing inspection failure or lack of ability to scale with growth.

This poster will explore hydrocarbon vs. non-hydrocarbon extraction methods, and the different regulatory considerations that go along with producing extracts. Attendees will develop a strong understanding of the different federal, state, and local governing bodies and codes, as well as dive into the space classification and permitting processes.

Presenter(s): Matthew Sica

ANSI National Accreditation Board (ANAB)

In the absence of industry standard methods for cannabis testing, robust and rigorous method validation is critical to ensure test methods are fit for intended use. Method accuracy, precision, range and detection limits must be evaluated prior to the introduction of any test method into the laboratory.

Method validation should be a planned activity in all cannabis testing laboratories. The scope of the method being validated, and acceptance criteria, must be defined before method validation is initiated. Validation activities must consider the entire scope of the method, including sample matrices and dilutions. Use of controls in method validation demonstrates and maintains the integrity of the validation.

The test method must meet the specified acceptance criteria to be considered fit for its intended use. Changes to methods must be evaluated and, where necessary, method validation should be repeated to ensure the altered method continues to fit the intended use.

Validation provides confidence that the methods used for cannabis testing generate accurate and reliable results. Industry standard methods could then be developed from these validated methods.

Presenter(s): Kimberley Russell

Bruker

Cannabis growers invest substantial time and resources into optimized strains of medicinal and recreational products. Cannabis is known as an efficient bio-accumulator plant. Nutrients (e.g., P, K), secondary nutrients (e.g., Ca, Mg, S) and micro nutrients (e.g., Cu, Fe, Mn, Mo, Zn) in natural or manufactured agents allow crops to develop into large, appetizing and potent products. Conversely, heavy metals (e.g., As, Cd, Hg, Pb) present in outdoor or indoor growth media can damage crops and develop hazardous products. Recent interest in optimized sustainable agriculture has led many organizations to monitor plant uptake of nutrients and heavy metals using battery operated portable X-ray fluorescence (pXRF). It is an elemental analysis method of choice because it is nondestructive and can be taken into the field, greenhouse or lab. PXRF is typically used for rapid screening of elemental nutrients and heavy metals in plant tissue, solids, powders or liquids for immediate actionable results. Real-time nutrient diagnostics and heavy metal screening with portable elemental analyzers help cultivators optimize plant characteristics and maintain product safety at any stage of the grow. This capability enables growers to make better decisions about their products in real time without having to send large quantities of samples out to a lab. The technology of pXRF will be described; the tradeoffs between screening samples as-is and analyzing samples which have been prepared will be discussed; and data correlated with ICP analysis will be presented.

Presenter(s): Edward Palumbo

Charlotte’s Web Inc.

In an industry associated with poor method transparency and inflated testing results, there is a demonstrated need for validated and verified potency assays to quantitatively determine cannabinoid concentration levels in a variety of hemp-derived sample types. There are several reverse phase (RP) HPLC-DAD methods reported in the literature; however, these methods are lacking in several ways. First, many methods do not adequately utilize robust method validation procedures, as outlined within AOAC and ICH guidelines. Additionally, most published HPLC methods are characterized by exorbitant run times, as well as extensive sample preparation and sub-dilution steps. To further compound this efficiency issue, most methods require multiple preparation methods and calibration curves due to the inherent variability in cannabinoid concentration, as well as matrix composition, across multiple sample types.

Through an extensive method development, validation and verification process, we have developed a RP HPLC-DAD method to test several different sample types across a wide concentration range. The assay provides for the quantitative determination of seven phytocannabinoids common to hemp: cannabidiolic acid (CBDA), cannabigerol (CBG), cannabidiol (CBD), cannabinol (CBN), delta-9 tetrahydrocannabinol (Δ9-THC), cannabichromene (CBC), and delta-9 tetrahydrocannabinolic acid (Δ9-THCA). In effect, it eliminates the need for additional HPLC runs or sample sub-dilutions through diode array detection (DAD) at multiple UV wavelengths. Furthermore, the method reduces HPLC run times and sample preparation procedures while significantly lowering the limit of detection (LOD) and limit of quantitation (LOQ) for minor cannabinoids. As a component of the verification process, orthogonal chromatographic methods were utilized, super critical chromatography (SFC), to augment the validity of cannabinoid testing results.

Presenter(s): Kyle Boyar

Medicinal Genomics

As the cannabis industry is on track to become a $50B+ industry, it would benefit from having access to a freely available high quality, reference genome to help standardize cultivar types, properly identify and classify them, provide a foundation for marker assisted breeding projects, and reduce the risk of IP that throttles the entire industry. As seen in many other economically important crops, advanced genetic tools can accelerate the production yields, focusing on narrowly defined traits. The basis for these genetic tools is access to a usable, reference genome. Medicinal Genomics has recently blockchain published the most contiguous and complete reference genome for Cannabis sativa L. More recently, the team has made further improvements in the assembly, boosting the N50 contig length to over 7.6Mb with less than 400 contigs representing the genome and a BUSCO score of >98%. To demonstrate the usability of this assembly, the team has completed both RNA-Seq and whole genome methylation studies to identify unique transcripts along with corresponding regulatory elements. A few examples include the identification of the regulatory regions for the Edestin and Cs2S genes along with two other sites important for seed production. We observe the CBCA, THCA and CBDA synthase gene clusters have been phased onto respective contigs demonstrating tandem repeat expansions. Several gene families that contain up to 30 paralogs, including the important THCA Synthase and CBCA Synthase genes have been identified. Interestingly, many of these sequences contain full length genes with most variation occurring at the 3’ and 5’ termini. By using RNA-Seq, we are actively annotating and identify which sequences are expressed and biologically active. To ensure the highest quality assembly is public and open access, we recruited 3 independent reviewers to review, comment, challenge, and improve the assembly published on the DASH blockchain, enshrining those reviews as part of the open publication process.

Presenter(s): Stephanie Balderston

Stillwater Foods

Cannabis infused products, particularly hemp derived, are being rapidly developed and brought to market in a test and learn approach. Certain food systems, such as ready to drink (RTD) beverages, are highly dynamic systems and rapidly gaining popularity in the cannabinoid space. The formulation and execution of water-soluble cannabinoids into RTD beverages is in its early stages for understanding the effects of thermal processing, pH, storage conditions and shelf life on cannabinoid stability.  Due to the lack of access to reliable laboratory testing, there is an added level of complexity to the production of consistent, stable, and safe products. We performed a 12-week shelf life study of RTD beverages to evaluate CBD concentration through utilization of liquid chromatography and chemical extraction methods. At the time of production, the beverages were bottled into amber and clear glass bottles, a common practice for a wide array of products that are susceptible to oxidation, including cannabinoids. Both bottle types were stored in a dark refrigerated environment and a commercial retail refrigerated environment, with constant fluorescent light exposure. Upon conclusion of the study there was no significant change in CBD concentration over time in relation to final packaging and storage conditions. As the cannabinoid industry continues to grow, it is important to provide safe and consistent products to consumers. Maintaining a high level of integrity in a competitive market will only fuel more innovation and establish key standards that will legitimize the use of cannabinoids as a functional food.  

 

Presenter(s): Brittany Lefler

CEM Coporation

With the use of cannabis for medicinal purposes becoming legal in numerous states, there is a need to confirm the safety of these products. Confirming that banned pesticides are not present in these products is one of the biggest concerns in this industry. The extraction of pesticides from cannabis can be challenging due to the high levels of interfering chemicals in the plant. Existing techniques do not offer a method that is rapid, simple, and efficient. In this study, a new extraction system, the EDGE, combines the processes of pressurized fluid extraction and dispersive solid phase extraction is explored. This new method offers efficient extraction of pesticides from cannabis in less than 10 minutes in one simple process. Different analytes beyond pesticides, such as cannabinoids and terpenes can also be extracted with this new method. The EDGE method offers a rapid, simple, and efficient solution for cannabis testing.

Presenter(s): Nathaly Reyes

Restek Coproration

While methods have been published that show the separation of major cannabinoids, many do not take into account the possibility of interference from minor cannabinoids. This is most problematic in concentrates where minor cannabinoids can be enriched to detectable levels. Additionally, some terpenes have been shown to absorb UV light at the same wavelength as cannabinoids, which can result in an additional source of interference. In this study, the LC-UV separation of 16 cannabinoids of interest was performed while monitoring for the potential impact from minor cannabinoids and terpenes on reported potency values. The method is applied to commercially available cannabis, cannabis concentrates, cannabis-infused edibles, and CBD oils.

 

Presenter(s): Lex Dreiter

WHITE LABEL AR

Our work modeling laboratory architecture and scientific data in three dimensional space and presentation in one-to-one real world scale has been widely reported. Using marker-less Augmented Reality we’re able to place complex laboratory systems in various configurations in exact scale. With marker based Augmented Reality we are able to overlay digital real-time cloud-based data on real world objects such as labels. We will show the applications of Augmented Reality in Cannabis Research, Testing, Cultivation, Extraction and Products – especially labeling and marketing

Presenter(s): Cheryl Denman

ANSI-ASQ National Accreditation Board (ANAB)

80% of cannabis testing laboratories are required to transition to the 2017 edition of the ISO/IEC 17025 standard in calendar year 2019. At this writing, just 20% of laboratories providing cannabis testing in North America have transitioned to the new 17025 standard. All of the remaining laboratories are still accredited to the old 2005 version. In the spirit of sharing information and transferring knowledge at Emerald Conference, this poster will present the changes and review the impact of ISO/IEC 17025:2017 transition specifically for cannabis testing labs and their customers. Included in this poster is the new ISO/IEC 17025:2017 structure and content. Key changes to ISO/IEC 17025 including those requirements that are new, substantially revised and deleted are thoughtfully presented. Finally, a crosswalk comparison will be posted and also made available as handouts. These valuable tools are timely and will aid labs in the navigation of the new standard during reassessment or surveillance activities in 2019.

Presenter(s): Simon Erridge1 , Marie Miller2, Tamara Gall1, Antonio Constanzo3, Barbara Pacchetti3, Mikael H Sodergren1,3

1Department of Surgery and Cancer, Imperial College, London, UK 2 Imperial Clinical Trials Unit, Imperial College, London, UK 3 Emmac Life Sciences, London, UK

Background Surgeons are seeking to reduce epidural and opioid use in the postoperative period due to their significant side-effect profiles. There is evidence that cannabis-derived medicinal products (CDMPs) have anti-emetic properties and in combination with opioids have synergistic analgesic effects in part signalling through the delta and kappa opioid receptors. The aim of this patient and public involvement (PPI) programme was to determine perception of perioperative CDMPs in our local population and to inform design and clinical end-points of a double-blinded randomised controlled clinical trial(RCT). Methods The programme used qualitative data from focus group(n=14) and semi-structured interviews(n=4). Qualitative data analysis was based on the framework methodology. Verbatim transcriptions were coded categorically into analytical frameworks for thematic analysis. Emergent themes and associated degree of consensus/dissent were determined. Results The participants were composed of a representative population cohort of patients and relatives (M:F=1:1, age range 33-85). Most common coding categories included in thematic analysis framework were side-effect profile, trial schedule of events and safety. Consensus across the PPI programme was that potential benefits of CDMPs were attractive compared to the known risk profile of perioperative opioid use. Decrease in opioid-dependence was agreed to be an appropriate clinical end-point for a RCT and there was high concurrence of a therapeutic schedule of 5 days. Participants identified constipation and nausea as the most significant opioid side-effects. Negative CDMP perceptions included addiction, dysphoria, and adverse effects in psychiatric sub-populations. Sub-lingual or oral administration was the most acceptable route of administration, with some expressing that inhalation delegitimises therapeutic properties. All participants found randomisation with placebo acceptable in coadministration with current analgesic gold standard.

Presenter(s): Peter JW Stone, Jennifer Hitchcock

Agilent Technologies Inc.

To enable low level pesticide and mycotoxin analytical measurements of cannabis-derived plant material by LC/TQ techniques, appropriate sample preparation routines are needed to efficiently remove analytes from suppressive background interferents found in original plant matrix. Sample preparation is required to yield sufficiently high recoveries of all analytes during safety testing under State regulations and must be relatively simple and inexpensive to undertake. Multiresidue pesticides and mycotoxins are examples of such analytes, therefore to meet California stipulations, these analyte recoveries must fall between 70-130%. This poster outlines such a sample preparation routine specifically developed for removing these analytes from plant matrix and which adheres to the Californian requirements. Herein we will detail each stage of the sample preparation process, inclusive of analytical instrumentation and acquisition methodology which incorporates a unique in-situ automated final dilution stage. Typical method detection limits (MDLs) obtained using this workflow for each of the LC/TQ amenable pesticide and mycotoxin analytes will be tabulated against the State action list to demonstrate analytical performance to less than 50% of the sensitivity requirements. The acquisition method chromatographic cycle time is 11 minutes, from sample to sample.

Presenter(s): Sareeta Nerkar, Maria Ofitserova

Pickering Laboratories

As medical and recreational Cannabis use gains broader acceptance, regulations are being put in place to mandate the testing of consumer products containing Cannabis. Legally available Cannabis plant and cannabis-containing edible products are tested for presence of pesticides, heavy metals, residual solvents and other harmful substances. State regulations have established maximum allowed levels for total Aflatoxins and Ochratoxin A in cannabis products, and laboratories are looking for methods to analyze these compounds.

We present easy and sensitive method to analyze Aflatoxins B1, B2, G1, G2 and Ochratoxin A in cannabis plant and edible products. Mycotoxins are isolated using immunoaffinity clean-up columns and analyzed with fluorescence detection. To increase sensitivity of Aflatoxins B1 and G1, an in-line photochemical reactor is installed before the detector. This method utilizes standard HPLC equipment and allows laboratories to easily determine Mycotoxins at levels below the limits established by state regulations.

 

Presenter(s): Laura Breit

Root Engineers

In this poster, we will explore how vapor pressure differentials (VPD) affect transpiration rates and, ultimately equipment sizing. An understanding of VPD at different temperatures and relative humidities, all of which influences maximum plant growth. Too little and peak growth rates are not achieved and plants cannot adequately cool themselves; too much and peak growth rates are also reduced, the plant stomata will close limiting transpiration, which can result in issues like tip burn and rot. In fact, our experience has shown that a set point difference of just 10 percent can almost double the necessary size of the required HVAC system for maintaining your optimal growth environment, which impacts first costs and energy costs.

 

Presenter(s): Melissa Wilcox & Edward Franklin

Regis Technologies

The global cannabis industry is growing rapidly, with many countries and US states adding regulatory frameworks for medical and recreational cannabis programs. Quality control is an essential component in protecting health and safety of the consumer in this emerging market and there is increasing demand upon cannabis testing laboratories for analytical determination of multiple cannabinoids. Current regulations concerning potency vary by jurisdiction, but usually require testing for the active forms of THC and CBD. In addition to those, many require testing for the acid forms, THCA and CBDA, along with other cannabinoids like CBG, CBGA, THCV, CBC and CBN. As regulations evolve, and as research interests in minor cannabinoids expand, it is important to have robust analytical methods in place that are capable of meeting those needs. Herein, the baseline resolution of 17 cannabinoids by high performance liquid chromatography (HPLC) with UV detection is described. Chromatographic method development was performed with particular attention to speed of analysis and a means for changing selectivity to improve resolution of critical pairs, as needed.

 

Presenter(s): Melissa Wilcox1, Edward Franklin1, Guilia Mazzoccanti2

 1Regis Technologies, Inc., 8210 Austin Avenue, Morton Grove, IL 60053, USA2Dipartimento di Chimica e Tecnologie del Farmaco – Sapienza Università di Roma, p.le Aldo Moro 5, 00185 Roma, Italy

Membrane proteins are frequently the targets for large families of binding ligands, including small drug molecules. The intrinsic chirality of protein drug receptors, which consist of L-amino acids, often governs specific pharmacological response to ligands with particular stereochemical geometries. Characterization of the potency, efficacy, and safety of individual enantiomeric ligands is therefore important in the early stages of drug development.Naturally occurring cannabinoids often contain chiral molecules that target cannabinoid receptors expressed in mammalian central and peripheral nervous systems. While certain isomers are pyschotropically active, others are not, but may have important therapeutic properties. Stereoselective analysis of endogenous cannabinoids and well as phytocannabinoids may prove essential for cannabis-related research, phenotype determination, and quality control of medicinal cannabis used in therapeutic treatment.Herein, chromatographic resolution of the natural racemic coupound, cannabichromene, in plant extract, along with the synthetic racemic (+/-)-Δ9-trans-tetrahydrocannabinol is demonstrated using supercritical fluid chromatography (SFC) coupled with mass spectrometry (MS).

Presenter(s): Maria Ofitserova, Sareeta Nerkar

Pickering Laboratories

Broader acceptance of medical cannabis use increases the need for analytical methods capable of determining the active compounds of cannabis. A new HPLC method with post-column derivatization was developed to analyze cannabinoids in cannabis plants as well as in cannabis-containing edible products. This post-column method is based on reaction with Fast Blue Salt reagent, a well-known color-forming reaction that is used in drug tests to detect cannabinoids via test-tube and thin-layer chromatography. Detection at 475 nm is performed using a UV/Vis detector. Our method implements a simple extraction with acidified water/acetonitrile followed by QUECHERS sample clean-up. The same procedure is applicable to both plant materials and edible products containing cannabis. The method is suitable for analysis of the major neutral cannabinoids such as THC, CBD and CBG as well cannabinoid acid (THCA) with high sensitivity and selectivity of detection.

Presenter(s): Jacob Jalali

PerkinElmer

Demands for accurate quantitation of pesticides in cannabis have gained significant prominence in recent times among cannabis industry stakeholders and government regulatory institutions among others. This in part may be due to the complexity and difficulty in treatment of the cannabis matrix for pesticide analysis. In addition, the more stringent pesticides limits in cannabis products in certain states within the United States have also broadened the analytical challenge. This study provides an analytical procedure for effective, reliable and accurate quantitation of Emerald proficiency tests for pesticide screening. The analytical method target is to identify & quantify all the present pesticides in Emerald efficiency tests using the PerkinElmer LX50, an ultra-high performance liquid chromatograph for effective separation with a PerkinElmer Quasar SPP Pesticides column (100×4.6mm, 2.7um) and the QSight 220 tandem mass spectrometer for accurate quantitation. All the reported result using this analytical method were in the acceptable range and Emerald badged were achieved by PerkinElmer.

Presenter(s): DJ Tognarelli

JASCO

As more states continue to legalize medicinal and recreation marijuana use, companies will consider entering or expanding into cannabis market. One of the most important considerations is how they will test their product. The results can determine the efficiency of their production, determine the value of their product and potentially determine the direction of the business. Cannabinoid potency is one of the most common tests required and in-house HPLC can offer immediate results in a more cost-effective solution. We have developed a sub 5-minute HPLC analysis for rapid potency results.Currently terpene analysis is performed by gas chromatography requiring a second instrument increasing the lab operation cost. Here we show how this can be overcome by also performing the terpene analysis on the HPLC showing the versatility of one instrument doing the job of two. With the addition of a mass spectrometer, the analysis of the cannabinoids and terpenes can be combined into one quick and easy analysis.

Presenter(s): Douglas Townsend

PerkinElmer Inc.

In the past few years, hemp extract has experienced a meteoric rise in popularity as a proposed alternative treatment to a range of medical conditions and lifestyle diseases. Coinciding with this surge is a rapidly growing number of administration methods, formulations and available products for the consumer. While hemp extracts have demonstrated evidence for medicinal value there remains many uncertainties surrounding the legality, quality and safety of these products.

Central to these issues is the accurate and reproducible manufacturing of hemp extracts and their formulated derivatives. Currently, there is very little in-process monitoring during the manufacturing of hemp extract products. As such, the industry has adopted a reactionary approach to manufacturing which can lead to inferior quality, irreproducible batches, lengthy processes and an overall inefficient use of laboratory resources.

Infrared spectroscopic determination of cannabinoid content in during each stage of the manufacturing process offers many advantages over traditional methods for potency determination. Users of this technique benefit from the ability to obtain accurate potency values in less than 30 seconds, without the need for sample preparation or hazardous solvents. The speed and simplicity of infrared spectroscopy is ideal for manufacturing staff who must make rapid and chemically informed decisions to ensure consistency in their products.

Chemometric models for cannabinoid content in hemp products will be presented and discussed, highlighting the utility of infrared spectroscopy as an in-process tool for potency determination.

Presenter(s): John Robinson

Avicanna, Inc.

The federal legal status of cannabis has led to a severe deficit in the quality and quantity of data supporting the safety, consistency, stability and pharmacokinetic profile of commercialized cannabis products. This limitation on research has led to regulations that do not mandate the basic studies normally required to bring a consumer product to market. Given this lack of oversight, self-imposed industry standards are needed to ensure that therapeutic, cannabinoid-based products are of the same quality and potency at the time of purchase as the day they were packaged. In many products, the cannabinoids will begin to degrade in as little as a week and show even greater signs of degradation at 25 days, resulting in as much as 15% lower potency. Similar to stability, not all formulations are equal in terms of bioavailability and absorption. A lead candidate for the treatment of neurological disorders, AVCN319301, is a cannabinoid oral formulation currently under development. Animal studies have shown improved bioavailability relative to administration of other formulations that are currently available from licensed producers in Canada. In comparison to the 93.10ng/mL peak plasma concentration of commercially available formulations comprised of MCT & Resin at 4 hours, and MCT & CBD isolate of 91.98 ng/mL at 6 hours, preliminary testing of AVCN319301 demonstrated a superior peak plasma concentration of 1046.99 ng/mL at 1 hour. In addition to focusing on bioavailability and absorption, ongoing testing and formulation enhancement to maximize product stability is currently underway.

 

Presenter(s): John Mills

bioMerieux, Inc.

The bioMérieux GENE-UP® Real Time PCR Pathogen Detection System has been evaluated for use with the GeneProof Aspergillus PCR Kit for the detection of Aspergillus fumigatus, Aspergillus terreus, Aspergillus flavus and Aspergillus niger.  This study encompassed pure culture testing of each target species, proficiency testing samples, and spiking studies with cannabis flower. The test method evaluation included the optimization of the lysis step which incorporated the GENE-UP Lysis Kit with patented Magic Cap. Use of this ready-to-use kit significantly simplifies sample preparation prior to PCR and increases the ease of use for laboratories which are required to screen aerosol products for Aspergilli. Following overnight incubation of the test sample, 20ul of enrichment is introduced into the lysis tube and then vortexed @2200 RPM for 15 minutes. The lysate is then added to a PCR tube with GeneProof PCR mix and analyzed on the GENE-UP system. A review of the PCR curves for each target organism, in all of the samples tested, exhibited good overall amplification and performance for each of the test samples analyzed during this investigation. These data demonstrate that the GeneProof Aspergillus PCR Kit used with the patented GENE-UP Lysis Kit and GENE-UP Real Time PCR System is a suitable method for the screening of cannabis and cannabis related products for the presence of Aspergillus spp.

Presenter(s): Colton Myers

Restek Corporation

With more states legalizing medical and recreational use of cannabis, there is a need for analytical methodologies to ensure consumers receive safe products. Currently, there is a growing trend in cannabis concentrate products. These concentrates are created by extracting the chemical compounds, such as Delta-9-Tetrahydrocannabinol (THC) and Cannabidiol (CBD), from cannabis flower. Manufacturers use a variety of extraction solvents in their processes. This study focused on analyzing residual solvents in cannabis concentrates with a like-USP 467 sample preparation technique. Subsequently, residual solvents were extracted with the more “classical” approach of HS-Syringe and then compared to extractions with a large volume HS-SPME approach (i.e., SPME Arrow). Comparison results and a simple solution for analyzing these complex matrices will be presented.

Presenter(s): Randy Reed

The Lehua Group USA

Mature cannabis markets are trending towards “terpier” products where taste, effect and smell are key features. Products like Terp Sauce, HTFSE, CDT (Cannabis Derived Terpenes), Live Resin, etc. all have one thing in common, they’re all rich in extracted cannabis terpenes. Many of these products aim to extract and preserve the native chemovar which is where the majority of the value lies in the product.

Chemovar is an extension of the term “chemotype” which was first coined by Dr. Rolf Santesson and his son Johan in 1968, defined as, “…chemically characterized parts of a population of morphologically indistinguishable individuals.” (1)

The term has been adopted and adapted to chemovar in the cannabis industry by such thought leaders like Dr. Raphael Mechoulam, Dr. Ethan Russo, Dr. Dedi Meiri, et al.

Chemovar can be described as a more accurate term for “Strains or Cultivars” i.e OG Kush, Northern Lights, Kona Gold, etc. It also accounts for Indica and Sativa nomenclature originally used to describe cannabis cultivars physical characteristics. (2) Thus, chemovar can be used to describe the chemical fingerprint of API’s found in cannabis.

“With this approach, it may be possible to move away from the current system of cannabis cultivars, with often vague and unsubstantiated characteristics dominated by Sativa/Indica labeling, toward a new classification based on chemical varieties (or “chemovars”) with a complex, but well-defined and reproducible chemical profile. (3)

“Chemovar drift” is not yet formally defined in cannabis nomenclature but a concept that most cannabis industry stakeholders and non-stakeholders alike intuitively understand. Delivery method and API (Active Pharmaceutical Ingredient) concentration aside, “Chemovar Drift” is speculated to be a reason why many cannabis products, namely extracts don’t taste, smell or feel like the input material.

That is, through the lens of a chemovar and all things being equal, does the output match the input?

By characterizing the native chemovar via GC-MS as baseline input material chromatogram and applying the same method to the final product, the concept of “Chemovar Drift” can be better understood.

What we’ve discovered over the past several years of operating a commercial cannabis CO2 extraction lab is that minimizing “Chemovar Drift” requires careful attention to detail and a multiphased method.

The cannabis matrix is complex w/ hundreds of API’s of varied affinities and volatilities.

API extraction can be optimized by “tuning” the solvent system for specific phases i.e liquid, gaseous, solid and supercritical to fractionate constituents w/ high efficiency. The term Multi-Phase Extraction (MPE) refers to the use of sub-critical phases and super-critical phases of CO2. Additionally, the use of ethanol in combination with CO2 constitutes the basis of MPE nomenclature.

The addition of ethanol to a CO2 solvent system increases the solvents systems polar affinity and thus the systems solubility spectrum. A modified MPE CO2 solvent system can span a wider range of solubilities than it’s individual parts and can dramatically increase mass transfer and extraction efficiency of “full spectrum extracts”.

CO2 extractions ability to fractionate enables the technology to extract and preserve the native chemovar by removing the terpene fractions and cannabinoid fractions separately. The volatilities and polarities of these compounds allow for efficient separation from the matrix. Fractionation is at the core of the MPE method because it enables the manufacturer to extract in a manner that minimizes de-naturing and degradation of native chemovars. The ability to fractionate in primary extraction allows for optimized downstream processing and retention of native chemovars not found in other commercially viable extraction methods.

Once the fractions are extracted they can be refined further or reconstructed into a number of products that retain the native chemovar with minimal chemovar drift.

It’s speculated that extracted products that retain the native chemovar are valued higher in mature cannabis marketplaces. We’ve seen this play out first hand in blind tasting panel competitions where top prizes are awarded to extracts that execute this method well. Our laboratory has taken top prize in Washington states most competitive open extract categories as “Best THC Oil”, “Best CBD Live Resin”, “Most Potent CBD Live Resin” and “Best THC Cartridge”. Other CO2 labs utilizing similar methods are also winning these highly competitive categories.

With the many methods available to process cannabis into concentrates, MPE via CO2 is superior in its ability to extract and preserve the native chemovar. Additionally, there are many other benefits associated w/ CO2 extraction such as safety, clean label, scalability, and regulatory ease. This method can be applied across equipment platforms w/ minimal modifications to the machines. That said some are better than others for this purpose.

All of our primary extraction is done on an Eden Labs 20L Hi-flow. We’ve found this system to be well built for commercial, 24/7 operation and excellent for fractionation. Chemotyping analytics by a third party, Confidence Analytics in WA state, using a Shimadzu HPLC Nextera-i for cannabinoid characterization and a Shimadzu GC-MS QP2020 for terpenes.

(1) Keefover-Ring K, Thompson JD, and Linhart YB. 2009. Beyond six scents: defining a seventh Thymus vulgaris chemotype new to southern France by ethanol extraction. Flavour and Fragrance Journal, 24(3): 117-122. doi:10.1002/ffj.1921
(2) Cultivar to Chemovar II—A Metabolomics Approach to Cannabis Classification
(3) Pollio A (2016) The name of Cannabis: a short guide for nonbotanists, Cannabis and Cannabinoid Research 1:1, 234–238, DOI: 10.1089/can.2016.0027.

Presenter(s): Geoff Faden

MAC-MOD Analytical

The determination of active components of cannabis is of increasing interest for quality control purposes, strain identification and for the determination of potency. Cannabis samples are complex and contain several target compound classes that are of interest from analytical and regulatory perspectives. Cannabinoids are a major compound class and are therefore of primary importance.

The work presented in this poster aims to develop methods for the routine separation of major cannabinoid components. A standard mix containing a total of 10 cannabinoids was used in this study. Regulatory requirements for testing are varied and may require determination of several key components included in this study, such as ∆9-Tetrahydrocannabinol (∆9-THC), cannabidiol (CBD), ∆9-Tetrahydrocannabinolic acid A (THCA), Cannabidiolic acid (CBDA), Cannabigerol (CBG) and Cannabinol (CBN). Samples were screened using a six-column screening protocol, to identify a suitable stationary phase. Stationary phases screened included a C18 phase and five Novel ACE Excel phases (C18-AR, C18-PFP, C18-Amide, CN-ES and SuperC18), specially developed to provide alternative modes of analyte retention and orthogonal selectivity. The mobile phase conditions were selected to be LC-MS compatible. The most promising result from the screening experiment was then optimised to provide complete separation of all ten analytes. Finally, the method was translated to a small format UHPLC column format to provide a method capable of determining these major cannabinoid components in <4 minutes.

 

Presenter(s): Savino Sguera

Digamma Consulting

The amount of data collected worldwide has grown exponentially over the last decade, and nowhere is this more apparent than with regards to cannabis. What began as only scant publications on studies performed using seized products has become caches of information such as chemical and genetic profiles, user experiences, growth and process yields, and contamination assays. In our study we employed machine learning and neural network algorithms on available data to derive meaningful insights that serve to elucidate the unknown and provide a framework of how to proceed in the coming decade.

 

Presenter(s): Avinash Dalmia

PerkinElmer

Over 30 states in the U.S. have legalized the use of recreational or medical cannabis because of therapeutic benefits for ailments such as cancer, multiple sclerosis, and ALS. Recently, government in Canada passed a law to allow its residents to use cannabis for medical and recreation purposes. Apart from Oregon state regulatory limits for about 59 pesticides in cannabis, the state of California has issued more stringent regulatory limits for about 65 pesticides residues in cannabis flower, edibles and oil. Normally pesticide analysis in cannabis and other food matrices is done by both GCMS and LCMS since some nonpolar and chlorinated pesticides are difficult to ionize with electrospray and APCI ion sources used in LCMS systems. LCMSMS method with electrospray and APCI source was used for low level analysis of all pesticides (including the very hydrophobic and chlorinated pesticides analyzed by GCMS) and mycotoxins in cannabis. The overall sensitivity for pesticides and mycotoxins was between 1-500 ppb in cannabis, well below regulatory limits set by the states of Oregon and California for pesticides and mycotoxins analysis in cannabis. The ability to screen and quantitate all pesticides, including the very hydrophobic and chlorinated GCMS amenable, in cannabis with LCMSMS only with dual ESI/APCI source makes this a novel way of screening and quantitation of pesticides in cannabis and different matrices with a single instrument. Long term stability data for pesticide analysis in cannabis was collected using a triple quadrupole mass spectrometer fitted with dual electrospray ionization source and atmospheric chemical ionization source ( APCI) and combined with a heated and self cleaning laminar flow interface. Long term stability data for pesticide analysis in cannabis showed that response RSD over 1 week for majority of pesticides was between 2 to 10 %. These results demonstrated that the heated self-cleaning interface with laminar flow interface in an LCMSMS system would reduce maintenance needs of the LCMSMS based method for pesticide analysis in dirty matrices such as cannabis. A simple solvent extraction method was used for extraction of pesticides and mycotoxins from dried cannabis flower. The recovery of all of pesticides and mycotoxins was between 70-120 % with RSD less than 20 %.

Presenter(s): DJ Tognarelli1, John Burchell1, Chris Densicola2v

1JASCO Incorporated, 28600 Mary’s Court, Easton, MD 21601; 2 Prism Sciences, Morrison, CO, USA

Preparative SFC is often the preferred technique for preparative purifications for its short analysis times, low solvent consumption and easy fraction recovery. The use of a semi-prep SFC-MS in this application provides analytical analysis of cannabinoids and terpenes with identification using a mass spectrometer and the direct scale-up to purification of the cannabinoids using mass directed fraction collection. The combined use of this system offers cannabinoid potency, impurity identification, terpene analysis, and scale-up purification for production of standards.

Presenter(s): David Kohler

ES Industries

Supercritical fluid chromatography (SFC) is a powerful chromatographic technique for the separation and isolation of complex mixtures from natural products. It has been useful in the areas of preparative chromatography and rapid analysis chromatography. Many chemicals can potentially be used as a supercritical mobile phase for SFC, however virtual all current practitioners of SFC use carbon dioxide (CO2 ) which offers several advantages, particularly when compared to preparative liquid chromatography. In addition, it is miscible with a wide range of organic solvents and it is nonflammable. CO2-based SFC is particular well suited to the area of preparative chromatography where it can be easily removed after fractionation, enabling the rapid recovery of isolated, pure compounds. In addition, any residual amounts of CO2 in isolated products are considered to be non-toxic.

At ES Industries we have developed a new phase and SFC chromatography column optimized for SFC separations and isolation of Tetrahydrocannabinolic acid (THCA) and Cannabidiolic acid (CBDA) from Cannabis. This new column/phase – Chromega NP-III has similar separation characteristics to 2-Ethyl pyridine a stationary phase and column traditionally used for separation and isolation of THCA and CBDA, however Chromega NP-III is able to rapidly separate both THCA and CBDA using minimal amount of ethanol as modifier solvent for CO2 mobile phase used in SFC chromatography. Traditional 2-ethyl pyridine columns require high levels of ethanol to obtain similar separations to the new Chromega NP-III column. For this presentation examples showing the superior separation performance of the new Chromega NP-III column will shown.

Presenter(s): Adam Floyd

PerkinElmer

The high demand for the analysis of cannabis products has catapulted the need for fast, accurate, and precise testing. Both quality (terpenes) and safety (residual solvents) are of paramount importance to the community. Traditionally, this analysis has been done with liquid injection in relatively long chromatographic run times. These new methods allow for the quick and clean analysis of cannabis products using headspace (HS), gas chromatography (GC) and mass spectrometry (MS).

Like all botanicals and plants found in nature, cannabis contains terpenes that are the aromatic oils in cannabis varieties with distinctive flavors and aromas. There have been up to 140 different types of terpenes reported in cannabis, but multiple studies suggest there are approximately 17 that are most common and can be used for examining chemovar (chemotype: those strains that have chemical properties that differ from each other’s). Among them are monoterpenes, diterpenes, and sesquiterpenes, which are characterized by the number of repeating units of a 5-carbon molecule called isoprene, the structural hallmark of all terpenoid compounds.

The use of HS-Gas Chromatography for the testing of residual solvents has been validated by several agencies including environmental, forensics and pharmaceutical. In addition to enhancing more accurate results, the technique of headspace allows for rapid, “clean” analysis ensuring productivity and ease of operation.

This presentation will focus on a method that allows for the analysis of both of these groups of compounds using HS/GC/MS to enhance sample throughput and laboratory productivity.

 

Presenter(s): Robert Kerr

Sorbent Technologies, Inc.

Coming Soon.

Presenter(s): Markus Roggen

Complex Biotech Discovery Ventures

CBDV is a young research venture that seeks to add fundamental chemical insight to the field of cannabis production. We present our latest findings in understanding, controlling and optimizing cannabis production in analytics, processing, extraction and formulation. The collaborative effort of the multidisciplinary research team at CBDV led to a diverse set of insight in every stage of production. For example, in-process analytics via IR, and an overview of factors controlling ethanol extraction.Using this insight, a multitude of process optimizations have led to a rethinking in cannabis production and products.

Presenter(s): Thu Huynh, Ph.D., Beatrice Kachel, Ph.D., and Wondu Wolde-Mariam

Helica Biosystems Inc

The rapidly expanding cannabis industry has resulted in a proliferation of state regulations to ensure the quality and safety of cannabis products. For example, suppliers must provide potency information to notify customers regarding the level of total tetrahydrocannabinol (THC) and cannabidiol (CBD) in their products. Cannabis must also be tested for specific mycotoxins, such as total aflatoxins (AFs), to confirm the absence of these potent human carcinogens produced by toxigenic fungi present in the soil. Several methods for potency and mycotoxin testing have been developed, including thin layer chromatography, gas chromatography, and high pressure liquid chromatography (HPLC), which are reliable and accurate for quantification of specific compounds. However, there are drawbacks. For example, HPLC is time intensive, requires sample clean-up, and can be costly. In this study, we developed enzyme-linked immunoassays (ELISAs) capable of detecting total AFs (B1, B2, G1, and G2) and the psychoactive compound, D9-tetrahydrocannabinol (D9-THC) from cannabis products. Our total AF low matrix ELISA has a standard curve ranging from 0.02-0.4ng/mL, while the D9-THC ELISA has a range of 500-8000ng/mL. The D9-THC ELISA measures %THC in a variety of cannabis products by altering the dilution to meet different %THC ranges, thus offering flexibility to meet variance among cannabis samples. Validation studies were performed on market products and the %THC recovered was 88-104%, while recoveries were 83-95% for total AFs. These assays may be useful to the cannabis value chain from growers to manufacturers of edibles who need rapid, cost-effective screening methods for measuring total AFs and THC.

Presenter(s): Frank Gallagher

Leaflyte Life Sciences, Cal Grow LLC

Coming soon.

Presenter(s): Zeyead Gharib

Harvest Direct Enterprises LLC

This session is to educate the cannabis community about the concept/theory behind Lossless Activation – and to explore why it’s better than traditional methods, and what it means for the future of cannabis and medicine. Lossless Activation is one of the hottest emerging trends in the cannabis industry and for good reason. Through this technology we can begin to capture the specific medical benefits of each distinct cannabis strain into non-smoked forms. LACY allows us to decarboxylate cannabinoids while maintaining the full plant profile we started with. The result is precisely dosed cannabis products such as capsules, inhalers, or topicals – that can for the first time ever capture the characteristics and effects of the strain we started with. This means we can target specific cannabis profiles towards specific ailments, creating a safe, effective, and stigma-free medicine that is plant-based and holistic. Essentially, making cannabis a real medicine.

Presenter(s): Catharine Layton

Waters Corporation

Cannabinoid profiling and potency determination is required for characterization of consumer products containing cannabis derived ingredients. With the massive influx of new product formulations to the market, the number of samples submitted to testing laboratories has increased significantly. Cannabinoid content is traditionally assessed via HPLC (High-Performance Liquid Chromatography) methodology, which can take up to 30 minutes per sample analysis. Traditional HPLC methods can be easily converted to methods which employ sub 2-micron column particle sizes, such as those utilized in UPLC (Ultra-Performance Liquid Chromatography). This poster will demonstrate the transfer of an isocratic HPLC cannabinoid potency method to a UPLC method using geometric scaling principles. The final method provides a dramatic increase in the number of samples that can be tested per day, solvent-savings, and preserves selectivity, linearity and resolution for the chromatographic separation of 16 cannabinoids.

Presenter(s): Patricia Atkins

SPEX CertiPrep

The QuEChERS method has been shown to be practical for pesticide analysis on a number of different sample types and is increasingly being employed on more difficult matrices. Unfortunately some matrices either by their nature or their economic value like cannabis can be difficult to analyze with just the QuEChERS method alone. These cannabis samples can show lower recoveries of pesticides and other target analytes than are often observed with more traditional agricultural products. An improved combined extraction and clean-up method is proposed in which both the extraction and dispersive solid phase extraction (DSE) steps are combined and heated using a pressurized fluid extraction and Adding heat and pressure to the process increases the efficiency leading to better sample clean-up and DSE improved analyte recoveries. In this study, a new combined extraction system was optimized to increase sample processing throughput, efficiency and recovery in a one-step process. Different analytes including pesticides, cannabinoids and terpenes were examined to determine improvement of recovery and method efficiency of the combined extraction apparatus. The new method showed marked improvement in sample clean-up, throughput and sample extraction recovery for cannabis testing.

Presenter(s): Thomas D. Kiselaka, Robert Scott McKinley Ph.D.ab, Guido F. Verbeck Ph.D.a

a Department of Chemistry, The University of North Texas, 1155 Union Circle #305070 Denton, TX, United States bLaCore Labs, 1801 Industrial Pkwy, Van Alstyne, TX, United States

Lipophilic cannabinoids and terpinoids have been shown to have a therapeutic effect in the treatment of numerous medical conditions. However, both classes of compounds have poor water solubility leading to a low bioavailability, limiting the overall efficiency of the medical treatment. Previous studies have found that the use of liposomes have improved the bioavailability of the lipophilic compound, however the bile salts and the low pH of the stomach in the gastrointestinal tract continues to limit the efficiency of drug delivery. The synthesis of nano-liposomes, 20-30nm in diameter, increases the permeability of the organic soluble compounds. Furthermore, with the use of a polar amphiphilic surfactants, such as Kolliphor RH40, transfersomes can be made, again increasing the permeability of the compound due to the elastic characteristic of the transfersome. This study has shown that the formation of nano-transfersomes with diameters below 25 nm containing cannabinoids and terpinoids can be achieved using the thin-film hydration method coupled with sonification. A parallel artificial membrane permeability assay (PAMPA) mimicking the gastrointestinal tract was used to determine the permeability of the compounds. The permeability of the nano-transfersomes containing both cannabinoids and terpinoids across the artificial gastrointestinal tract membrane was enhanced by an order of magnitude. This report allows researchers to further study the optimal diameter of nano-transfersomes for permeability across the GI membrane by changing the diameter of the nano-transfersomes. In addition, other PAMPA models can be used to determine the permeability of nano-transfersomes across other membranes, such as the skin for topical treatments.

Presenter(s): Patricia Atkins

SPEX CertiPrep

The cannabis industry has taken the scientific world by storm, flooding the market with new products. Recently, concerns have arisen about safety of this, unregulated market, resulting in many new labs testing for cannabinoid potency, pesticides, bacteria/mold and other potential contaminants. Recreational cannabis and hemp are part of the C.sativa species, with different cultivars resulting in unique cannabinoid profiles. However, due to United States prohibitions on cannabis growth and sales, it has been difficult for the emerging cannabis industries to develop methodologies and standards to validate their testing using certain varieties of cannabis. Hemp in many cases has become a substitute matrix for the cannabis testing methods.

Pesticides are a large concern for the cannabis industry since there are thousands of potential pesticides available around the world and only a handful of regulations or guidance as to their use with cannabis. While many methodologies exist for the extraction of botanicals for pesticides, there are no validated methods or certified reference materials for use by the cannabis industry. This study examines the process of creation of cannabis CRM and the challenges of the production and use of a hemp based cannabis CRM.

Presenter(s): Taylor Trah, Blake Grauerholz

OutCo

The cannabis industry has taken the scientific world by storm, flooding the market with new products. Recently, concerns have arisen about safety of this, unregulated market, resulting in many new labs testing for cannabinoid potency, pesticides, bacteria/mold and other potential contaminants. Recreational cannabis and hemp are part of the C.sativa species, with different cultivars resulting in unique cannabinoid profiles. However, due to United States prohibitions on cannabis growth and sales, it has been difficult for the emerging cannabis industries to develop methodologies and standards to validate their testing using certain varieties of cannabis. Hemp in many cases has become a substitute matrix for the cannabis testing methods.

Pesticides are a large concern for the cannabis industry since there are thousands of potential pesticides available around the world and only a handful of regulations or guidance as to their use with cannabis. While many methodologies exist for the extraction of botanicals for pesticides, there are no validated methods or certified reference materials for use by the cannabis industry. This study examines the process of creation of cannabis CRM and the challenges of the production and use of a hemp based cannabis CRM.

 

Presenter(s): Chris Denicola

Prism Sciences

In this poster, we intend to show that novel materials that we created can be used to selectively retain neutral and acidic cannabinoids with the purpose of streamlined extractions and the simplification of the sample prep for pesticide analyses. One of the issues cannabis and hemp testing labs are having with pesticide testing is the contamination of the mass spec (loss of overall signal) by the high concentration of cannabinoids, mainly THC. If we can removed only the cannabinoids, then labs will be able to have a cleaner sample, less interferences, and less instrument downtime due to maintenance and cleaning. It should also mean that with less cannabinoids in the sample, a reduced dilution can be used resulting in higher data quality.

With this work, we will directly show that the use of these materials on crude hemp plant extracts (also acting as a proxy for hemp plant samples) and decarboxylated hemp oil results in higher data quality and no residual cannabinoids in the samples. With no cannabinoids in the processed samples, we can make 2 assumptions. The first is that there will be less downtime for the mass spec and therefore more runs per day/month resulting in higher revenues. The second is that when scaled up and applied to hemp and cannabis extraction, the resultant oil will be clear of chlorophyll, waxes, and most importantly, pesticides.

Last year at Emerald Conference, I was speaking to an attendee about this work and a different attendee said it was impossible (maybe a bit harsher)…so, to keep this independent, the work is being done by Dr. Kevin Schug’s lab at the University of Texas at Arlington.

 

Presenter(s): Loy Jones

Agilent Technologies, Inc.

Cannabis-based products are available in a wide variety of formulations ranging from dry plant material, plant concentrates including waxes and distillates, and in fused products such as foods and candies. Given the variety of sample matrices, existing sample preparation procedures developed for inductively coupled plasma-based techniques can be applied to cannabis products. For example, trace element analysis of plant and nutritional supplement materials is a well-established application. Following acidic digestion to break down the primary components of the plant-based samples, inductively coupled plasma–mass spectrometry (ICP-MS)orICP-optical emission spectrometry (OES) is often used for quantitative analysis because of the multielement capability, speed, and robustness of each technique. ICP-OES is suited to the analysis of mineral and micronutrients such as K,Ca,Mg,Cu,Fe,Mn,Zn,Cu,Mo, and Ni—vital elements required for plant growth.
When the required analytes also include trace elements such as As, Se, Cd, Pb, and Hg, which may require lower detection limits, ICP-MS offers greater sensitivity, delivering detection limits and accurate analysis down to nanogram-per-liter (part-per-trillion) levels.

 

Presenter(s): Justin Steimling

Restek Corporation

Mycotoxins, secondary metabolites produced by fungi, are among the most toxic contaminants in cannabis and other agricultural products that can cause disease and death in humans and other animals. In the analysis of mycotoxins, using immunoaffinity columns (IACs) to reduce matrix effects and eliminate potential sources of interference for LC-MS/MS analysis is common. When not used, significant matrix interferences have been shown to elute near target mycotoxins resulting in an adverse effect on measured ion ratios. Herein, alternative approaches to IACs including dilute-and-shoot, dispersive sold phase extraction (dSPE) and pass-through SPE are evaluated and applied to the analysis of aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, and ochratoxin A in commercially available CBD oils by LC-MS/MS.

Presenter(s): Joseph Evans, Dave McCurdy

Trufolia Labs

Headspace/GC is the analysis of choice for residual solvents in cannabis extracts. Headspace techniques require sample dissolution, or a solid extract is placed inside a headspace vial; i.e. Full Evaporation Technique (FET). Henry’s Law specifies that the solvents will exist in equilibrium within the gas and solution phase, or gas and melted-sample phase, and the solvent mass ratio between the two phases is dependent on the unique partition coefficient of the liquid matrix. FET assumes 100% solvent partitioning to the gas phase and therefore matrix matching for external standards is not required. However, when using FET on solid extracts > 20mg (aliquots > 20 mg are required because of inhomogeneous residual solvent distribution) there is an incomplete transfer of the solvents within the matrix into the gas phase. Solvent concentrations are then subject to partition coefficient variations resulting from different sample matrices and therefore not directly quantifiable using an unmatched external standard. Experimentation showed matrix differences that cause significant partition coefficient variation. A round-robin study with a cannabis shatter also showed large concentration differences for butane reported by 5 different laboratories using FET. Concentration differences, in part were because of differences due to sample size variations between labs: Mass differences changed the partitioning of butane between sample phases. Conclusions suggest that FET is not a good method of choice for analysis of cannabis extracts and that additional method development is needed for analysis using a dissolution technique that ensures matrix matching calibration techniques

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