Separation of cannabinoids for clinical use

PrepChrom C-700: Isolation of cannabidiol (CBD) and tetrahydrocannabinol (THC) from other cannabinoids extracted from cannabis

PrepChrom WordPressThe separation of cannabinoids is an important process which is necessary for clinical studies. The aim of these studies is to investigate the effect of the isolated cannabinoids on patients suffering from chronic illnesses and side effects caused by chemotherapy. The relaxation of laws concerning medical use of cannabis all around the world, especially the United States, has increased the interest in determining the effect of cannabinoids on the human body. Clinical studies require large amount of highly pure separated cannabnnoids. This can easily be achieved with the PrepChrom C-700.

Introduction

Cannabis is an extremely versatile plant and has been used for many purposes throughout history. Some specific sorts are used for fibers in the textile and paper industry because of their long stems. It is a useful source of foodstuffs, such as hemp oil and seed. Due to the cannabinoids, cannabis is also a popular recreational drug. These substances can cause mental and physical effects when consumed. The impact of the cannabinoids on the human body is of great interest for the pharmaceutical sector. The research emphasis lies in treating the side effects of chemotherapy, inflammatory diseases like multiple sclerosis or degenerative illnesses such as Parkinson’s disease.[3] AI FAME GmbH, a Swiss company, is a pioneering company to extract the plant-based active substances and make them water-soluble for improved, further processing.

Here, we aim to separate three cannabinoids, i.e. cannabidiol, tetrahydrocannabinol and tetrahydrocannabinolic acid in high purity from the extract.

Experimental

Equipment: PrepChrom C-700

Sample: Water-soluble cannabis extract. Cultivated, extracted and provided by AI FAME GmbH

Preparation: The chromatographic separation is performed with 1 mL of the water-soluble cannabis extract diluted in 1 mL methanol : water (1:1)

Separation: Method parameters
Column:  Sepacore® C-18 80 g
Particle size:  40-63 μm
Flow rate:  40 mL/min
Sample loop:  2 mL
Detection:  253, 270 nm and SCAN 200-600 nm

Gradient: methanol (A) : water(B)
50 % – 90 % methanol (A) in 20 min
90 % – 90 % methanol (A) in 10 min
90 % – 95 % methanol (A) in 5 min
95 % – 95 % methanol (A) in 10 min

Results

Cannabidiol (CBD) bearing a 1,3-diol functional group on its benzene ring, is the first substance to be eluted when applying reversed phase conditions (Figure 1), hence, is the most polar cannabinoid in the sample. The second compound, eluting after 27 minutes, is tetrahydrocannabinol (THC). The last substance to be eluted is tetrahydrocannabinolic acid (THCA), a form of cannabinoid which easily decarboxilates in to THC when exposed to light or heat. Assignment of the signals and identification was done by comparison with reference HPLC data provided by AI FAME GmbH.

Cannabinoide SN finalFigure 1: Methanol : water separation of the extracted cannabinoids using the PrepChrom C-700. Detection at 270 nm, 254 nm, and 200-600 nm SCAN.

 

 

Conclusion

The three cannabinoids extracted by AI FAME GmbH could easily be separated and separately collected with the PrepChrom C-700.

The water-solubility of the extract enables the use of aqueous solvents that have inherent economic and ecologic advantages over common organic solvents.

The growing interest in the clinical use of cannabinoids is unbreakable. Furthermore, cannabis is decriminalized and it is allowed to exploit the potential of its valuable substances. BUCHI offers straight forward solutions to separate and purify small to large size batches of the precious extracts.

Acknowledgment

AI FAME GmbH, Switzerland, is thanked for providing the extract and for the fruitful analytical discussion.

References

[1] Sutton, IR. Daeniken, P. (2006). Cannabinoids in the management of intractable chemotherapy-induced nausea and vomiting and cancer-related pain. The Journal of Supportive Oncology.

[2] State Medical Marijuana Laws. National Conference of State Legislatures. (16.3.2015). http://www.ncsl.org/research/health/state-medical-marijuana-laws.aspx

[3] Kogan, NM. Mechoulam, R. (2007) Cannabinoids in health and disease. Dialogues Clin Neurosci. 2007 Dec; 9(4): 413–430.

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EPA compliant sample preparation for environmental analysis

Current regulatory developments in the US market have led to increased use of the BUCHI Syncore in contract labs

Environmental_landscape lr

Industrialization, globalization, economic growth and an expanding green conscience demand a qualitative and quantitative leap in environmental protection and analysis. This development is widely supported, globally accepted, and increasingly impacts local as well as global regulation and law. Numerous countries have implemented strict regulations or have tightened their demand for an environmentally sustainable economy during recent years.

One of the latest and most prominent examples for tightened legislation is the United States Environmental Protection Agency (US EPA). The US EPA has not only lowered the threshold values for numerous harmful emissions for both industry and research, but has also started to tightly regulate laboratories responsible for testing such emissions. This has created quite a significant environmental testing industry, mostly consisting of customer contract laboratories specialized in testing various environmental samples in accordance to US EPA regulations. Recent news from several US states indicate that the agency has started to enforce those regulations and closely monitor affected institutions. This has already led to several substantial and publicly released monetary fines in 2014. The main point of interest of the US EPA is the organic solvent balance of affected companies. Solvents purchased and used by companies must be accounted for upon disposal, discrepancies are attributed to loss of solvent to the atmosphere. Among other factors, organic solvent emissions are a critical influence on air pollution and the greenhouse effect. Companies which fail to verify their solvent turnover are subject to further EPA investigations.

This situation becomes important once contract labs have reached a certain size. With dozens or hundreds of mandatory solvent exchanges and evaporations every day, the solvent emission factor becomes a serious aspect for every company in this segment.

For more than 75 years the family owned Swiss company, BÜCHI Labortechnik AG, has been known as a leading provider for solutions in rotary and parallel evaporation. Sustainability is one of BUCHI’s core values, a creed that has resulted in many environmentally friendly innovations and products which aim to conserve energy and water resulting in the smallest possible impact on the environment. One such product is the Syncore® Analyst, a highly productive parallel evaporator combined with the latest SVR (solvent vapor recovery system) technology, which allows simultaneous drying or concentration of up to 12 samples to a pre-defined residual volume at low temperatures while applying gentle vacuum conditions.

The Syncore® is tailored to the demands of modern high-throughput environmental laboratories. High analyte recoveries, even of sensitive compounds such as SVOCs, are obtained by means of a chilled zone at the bottom of each sample vessel. When used in combination with the programmable BUCHI vacuum controller and the individual sample sealing work quality is increased by automating processes to produce reproducible results and eliminate the chance of samples going to dryness or being cross-contaminated. High analyte recoveries are further obtained using a chilled Flushback module. The Flushback module partially condenses the solvent vapor at the top part of the sample vessel generating a continuous rinsing along the glass wall. Thus, adsorption of analytes at the glass wall is avoided.

The solvent vapor is condensed at the primary condenser and collected, a post-pump secondary condenser makes sure no solvent vapors escape the evaporation system. As a result, solvent recoveries greater than 95% are achieved. This not only eliminates harmful solvent emissions inside the laboratory, it is also beneficial to the environment and saves money as the recovered solvent can be reused.

Automated parallel sample concentration with high analyte and solvent recovery is achieved with the Syncore® Analyst. The Syncore® technology reduces manual labor and is fully compliant with the latest amendments to the regulations by the US EPA Solvent Emission Program, yielding solvent recoveries greater than 95%.

Current regulatory developments in the US market have therefore led to increased use of the BUCHI Syncore in these contract labs. This increased popularity of the Syncore not only proves that the combination of fast parallel evaporation, low cost per sample, and high solvent recovery is unrivaled, but also that the instrument is fully compliant with the latest emission regulations.

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Voice of the customer:
“If you’re looking to take your laboratory to a better level of quality, efficiency, and environmental friendliness, the Syncore Analyst is the way to go”
>> Download Case Study

Lionel Thomas, Laboratory Manager,
Accutest Laboratories, NJ


Solutions for alcoholic beverages

From raw material to the drinks you taste

Different alcoholic drinksIn this modern, digital world consumer product experiences are rapidly communicated to a global audience. More than ever before, the value of a brand is reflected in the quality and consistency of every single product introduced into the marketplace despite the possible variation in the raw material quality or the production process. The production of alcoholic beverages with consistent taste and quality requires experience and quality control at all stages of production, from raw material inspection to final product analysis.

BUCHI understands how to assist the manufacturers of alcoholic beverages in meeting your goals and offers solutions for the entire production cycle.

Production cycle step Sample matrix Analyte Analysis technology
Raw material e.g. barley, hops, grapes Moisture, protein, nitrogen, alpha and beta bitter acid, storage index NIRSolutions
Malting, fermentation,
wine pressing,
distillation
Malt, grains Total nitrogen, soluble nitrogen, moisture, acid, alcohol, starch NIRSolutions
Final product Wine, beer, spirits Alcohol, ethyl carbamate, volatile acids, sulfur dioxide, protein Steam distillation, Kjeldahl,
solid phase extraction (SPE)

For more than 50 years, BUCHI has been developing ingenious solutions for direct steam distillation and Kjeldahl applications. Our solutions include innovative products for the entire process workflow, customized application support, practical apps, and professional maintenance. Tailor-made hardware, software support, and precalibrations underpin the NIRSolutions concept.

Explore BUCHI’s solutions covering

  • the broadest range of applications for steam distillation (alcohol, SO2, volatile acids, protein)
  • more than 50 years of experience in protein determination according to Kjeldahl
  • combined parallel solid phase extraction (SPE) and parallel evaporation for the determination of ethyl carbamate
  • the multi-component quality check of raw materials and final products applying the NIR technology
  • the real-time in-line process monitoring enabled by NIR, VIS and camera sensors

By collaborating closely with our customers, we do everything in our power to make our products, systems, solutions, applications and services as sustainable as possible for people and the environment.

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Voice of the customer:
“The solution for Kjeldahl as well as sulfur dioxide determination in one single instrument is excellent. Moreover the support for application development is very much appreciated.”

Dr. S.S. Marwaha, Chief Executive Officer,
Punjab Biotechnology Incubator, India.


Slip melting point determination for the quality control of lipstick or palm stearin

m565Measuring the slip melting point (SMP) is a popular and important method used for the quality control of waxy compounds in the cosmetic or food industries.

Such applications can be easily performed using the standard BUCHI M-565 Melting Point instrument. The obtained results show a high reproducibility and the process can be automatically recorded for later analysis if required.

Two application notes clearly demonstrate that SMP measurements can be reliably performed for the quality control of lipstick or palm stearin following the USP recommendations.

Application Note #67: Determination of Pesticides in Soil

SpeedExtractor E-916

SpeedExtractor E-916

This Application Note #67 describes a fast and reliable way to extract organochlorine pesticides (OCP) from soil with the SpeedExtractor E-916. After reducing the volume with the Syncore Analyst the extracts were cleaned by using Florisil. The cleaned extracts were concentrated again and quantified by GC-ECD.

High pressure extraction is used for a wide range of applications in environmental analysis. Recognition by the US EPA method SW-846 3545 and SW-846 6860 reflects the excellent results in comparison to classic Soxhlet extraction.

The following analytes are particularly suitable: PAH, BNA, PCB, dioxins and furanes, TPH, pesticides and many more.

Introduction:
A pesticide is any substance or mixture of substances intended for, preventing, destroying, repelling, or mitigating any pest. There are three main types of pesticides: organochlorine, organophosphate and carbamates. OCP can cause severe health problems, are persistent and tend to bio-accumulate. Therefore the use of some OCP is banned by the Stockholm convention.

Experimental:
Instrumentation: SpeedExtractor E-916 with 20 mL cells, Syncore Analyst, Thermo Trace GC Ultra
Samples: CRM 847-050 and CRM 804-050 by R.T. Corporation.

Depending on the expected values of the OCP in the samples 1 to 5g of sample were weighed and mixed with sand. The mixture was transferred to the cell and two surrogates were added prior to the extraction. The cells were extracted and 150 mL Syncore® vessels with appendix were used as collection vials.

After the extraction, 1 mL of Internal Standard (IS) was added to the extracts. After concentration on Syncore® Analyst a clean-up with Florisil was performed. The volume of the cleaned solution was reduced again and quantification of 16 different pesticides was performed by GC-ECD.

A fourfold extraction of the samples was done. Two blanks were extracted in parallel on two positions not used for samples. In addition post-extractions of the samples were to show the performance of the method.

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Application Note #64: Determination of Pesticides in Spices By Extraction

Determination of Pesticides in Spices

Determination of Pesticides in Spices

Application Note #64 describes a fast and reliable way to extract organochlorine and other pesticides in chilli and paprika spice samples with the SpeedExtractor E-916. After the extraction the extract was cleaned up via GPC and analyzed by GC-MS/MS. Contact us to request this Application Note.

Introduction:
A pesticide is any substance or mixture of substances intended for preventing, destroying, repelling or mitigating any pest. Pesticides can be classified by their chemical structure (organochlorines, organophosphates and carbamates) or by their target (herbicides, insecticides, fungicides, rodenticides, pediculocides and biocides).

The adverse health effects depend on the type of pesticide. Organophosphates and carbamates for example affect the nervous system. Others may irritate the skin or eyes. Some pesticides may be carcinogens. Others may affect the hormone or endocrine system in the body.

Experimental:
Instrumentation:
SpeedExtractor E-916 with 20 ml cells, GPC Gilson 233XL, Varian 3800 with MS-1200 QQQ

Samples:
Paprika and Chilli Powder with incurred pesticide content.

3 g sample was weighed in a beaker and 6 ml of water was added. The mixture was placed in the refrigerator over night. The next day 3 g of diatomaceous earth was added and mixed in well.

The sample was transferred into a 20 ml extraction cell. A fourfold extraction was carried out using the parameters shown within the Application Note. A blank and a spiked sample were extracted as well (QC test). After the extraction a GPC clean-up was performed and the pesticide content was analysed by GC-MS/MS. For comparison the samples were also extracted with a Dionex ASE® 200.

Nitrogen & Protein Determination in Milk By Digestion

Protein Determination in Milk

Protein Determination in Milk

Nitrogen and Protein Determination in Milk by Digestion with Hydrogen Peroxide and Sulfuric Acid

Application Note #54 introduces a simple and very fast procedure for protein determination in milk and provides detailed steps for completing the process. BUCHI’s Kjeldahl application database lists more than 100 Kjeldahl and Non-Kjeldahl applications using dedicated BUCHI equipment: Download #54 and all our other Kjeldahl Application Notes online.

The sample is digested with hydrogen peroxide and sulfuric acid using the SpeedDigester K-436 or K-439, followed by distillation and titration with the Kjeldahl Sampler System K-370/K-371. The determined protein contents correspond to the results obtained with the Kjeldahl method.

Introduction:
The digestion with hydrogen peroxide 30% instead of Kjeldahl tablets for nitrogen and protein determination is a very fast (30 min instead of 85 min (K-439) or 100 min (K-436)) and environmentally friendly (free of any heavy metal) alternative to the classical Kjeldahl methods.

Instrumentation:
SpeedDigester K-436, K-439, with H2O2 suction module, Kjeldahl Sampler System K-370/K-371

Samples:
Whole milk UHT, partially skimmed milk UHT, and chocolate milk beverage: labeled protein contents 3 g/100ml (whole milk) and 3.5 g/100ml (partially skimmed and chocolate milk). Protein determinations according to the classical Kjeldahl method, measure 3.15% for the whole milk, 3.19% for the partially skimmed milk, and 3.30% for the chocolate milk beverage.

Determination:
Approx. 5 g of the homogenized sample were weighed directly into a sample tube. A portion of 20 ml of sulfuric acid was added, and the digestion was started using the parameters specified (See complete note for data). Initially, 15 ml of hydrogen peroxide 30% was added to the capillary funnel 2 minutes after the start of the digestion. Then, another 15 ml of hydrogen peroxide 30% was added to the capillary funnel 10 minutes after the start of the digestion. The method was verified by using 0.18 g tryptophan as the reference substance.