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Cannabis sativa L. is an extremely variable species. Even within the same cultivar plants can significantly differ in the content and profile of cannabinoids. Therefore, the best method for production of uniform plants and standardized raw material is vegetative propagation using clones. The aim of this study was to determine the content of cannabidiolic acid (CBDA), cannabidiol (CBD), Δ9-tetrahydrocannabinolic acid (Δ9-THCA), Δ9-tetrahydrocannabinol (Δ9-THC), cannabichromene (CBC), cannabigerol (CBG), and cannabinol (CBN) in clonally propagated plants of industrial hemp. One hundred and thirty-nine plants representing 17 different hemp genotypes were regenerated in vitro, hardened, and grown in a vegetation hall until harvest. Single plants of each accession were analyzed using high-performance liquid chromatography with UV/diode-array detection (HPLC-DAD/UV). The results revealed significant variability in the total cannabinoid content (0.55–5.18% in dry weight) among tested genotypes and within the Epsilon 68 cultivar. The highest content of total CBD (4.410%) was recorded for EPS/40 genotype, while the level of total Δ9-THC was below the allowed threshold (0.3%). Therefore, we can conclude that some clonally propagated plants provided reproducible hemp material as a potential source of cannabidiol. The results of this study will be useful for breeding and early selection of hemp genotypes.

Influence of agroclimatic conditions on active substances content in hemp cultivated in the South-East Baltic region Abstract: Due to legal regulations prohibiting cultivation of cannabis (Cannabis indica Lam.) in many countries, industrial hemp (Cannabis sativa L.) remains the main source of active substances with potential application in the pharmaceutical industry. To assess the possibility of using the varieties of industrial hemp for this goal, and their adaptation to the habitat conditions and the agricultural technology appropriate for them, we investigated three monoecious varieties of hemp (‘Futura 75,’ ‘KC Dora’ and ‘Tygra’) at different sowing densities (60 germinating seeds·1 m2 or 180 germinating seeds·1 m2) and nitrogen fertilization levels (0, 30, 60, 90 kg·ha−1). In none of the tested hemp varieties registered as fibrous did the concentration of Δ9-THC exceed 0.2%, satisfying the requirements of European legislation for industrial hemp varieties. The tested varieties did not differ significantly in the cannabidiol (CBD) and Δ9- tetrahydrocannabinol (Δ9 -THC) content in the dry matter of inflorescences. Agronomic factors such as sowing density or nitrogen fertilization did not modify the content of CBD and Δ9-THC. This result is very helpful for farmers, because it allows them to select hemp varieties of dual-purpose production (stems and inflorescences or stems and seeds) adapted to the South-East Baltic environment.

The aim of the experiments was to determine the effect of sowing density and nitrogen fertilisation on the cannabidiol (CBD) and Δ9-tetrahydrocannabinol (Δ9-THC) yields in selected cultivars of fibre hemp. A three-factor field experiment was conducted in 2014-2016. Factor A: cultivar (Futura 75, KC Dora and Tygra), factor B: sowing density (60 and 180 seeds/m2) and factor C: nitrogen fertilisation (0, 30, 60 and 90 kg/ha). The CBD yield ranged from 10.52 g/m2 in cv. Tygra to 10.99 g/m2 in cv. KC Dora. Among the examined cultivars, the highest yield of Δ9-THC in cv. KC Dora was observed. Sowing density did not modify the CBD yield, but increasing the density from 60 to 180 seeds/m2 caused an increase in the yield of Δ9-THC in cvs. KC Dora and Tygra. Nitrogen fertilisation significantly influenced the yields of CBD and Δ9-THC. In both cases, yield increased until the rate of 60 kg N/ha.

Synthetic seed technology and cold storage methods provide genetic uniformity, paving the way for pest- and disease-free plants that are easy to handle. The aim of this study was to develop protocols for the cold storage of nonencapsulated and alginate-capsulated explants of Cannabis sativa L. Axillary shoot tips derived from in vitro-grown plants that were used as explants and stored for up to 9 months at 4 °C in the dark. Somatic seeds were produced in 3% sodium alginate and Murashige and Skoog (MS) medium salt and stored for up to 3 months. After 6 months of cold storage, the highest regrowth of 45% was recorded for the nonencapsulated explants. The recovery of somatic seeds was 90% under the same storage condition after 3 months. Well-developed, regenerated plants from encapsulated explants successfully acclimatized.

Commonly used clinical strategies against coronavirus disease 19 (COVID-19), including the potential role of monoclonal antibodies for site-specific targeted drug delivery, are discussed here. Solid lipid nanoparticles (SLN) tailored with tocilizumab (TCZ) and loading cannabidiol (CBD) are proposed for the treatment of COVID-19 by oral route. TCZ, as a humanized IgG1 monoclonal antibody and an interleukin-6 (IL-6) receptor agonist, can attenuate cytokine storm in patients infected with SARS-CoV-2. CBD (an anti-inflammatory cannabinoid and TCZ agonist) alleviates anxiety, schizophrenia, and depression. CBD, obtained from Cannabis sativa L., is known to modulate gene expression and inflammation and also shows anti-cancer and anti-inflammatory properties. It has also been recognized to modulate angiotensin-converting enzyme II (ACE2) expression in SARS-CoV-2 target tissues. It has already been proven that immunosuppressive drugs targeting the IL-6 receptor may ameliorate lethal inflammatory responses in COVID-19 patients. TCZ, as an immunosuppressive drug, is mainly used to treat rheumatoid arthritis, although several attempts have been made to use it in the active hyperinflammatory phase of COVID-19, with promising outcomes. TCZ is currently administered intravenously. It this review, we discuss the potential advances on the use of SLN for oral administration of TCZ-tailored CBD-loaded SLN, as an innovative platform for managing SARS-CoV-2 and related infections.

The bioavailability levels of cannabidiol (CBD) and tetrahydrocannabinol (THC) determine their pharmacological effects. Therefore, for medical purposes, it is essential to obtain extracts containing the lowest possible content of the psychogenic component THC. In our extract, the CBD/THC ratio was 16:1, which is a high level compared to available medical preparations, where it is, on average, 1:1. This study assessed the bioavailability and stability of CBD and THC derived from Cannabis sativa L. with reduced THC content. The extract was orally administered (30 mg/kg) in two solvents, Rapae oleum and Cremophor, to forty-eight Wistar rats. The whole-blood and brain concentrations of CBD and THC were measured using liquid chromatography coupled with mass spectrometry detection. Much higher concentrations of CBD than THC were observed for both solvents in the whole-blood and brain after oral administration of the Cannabis sativa extract with a decreased THC content. The total bioavailability of both CBD and THC was higher for Rapae oleum compared to Cremophor. Some of the CBD was converted into THC in the body, which should be considered when using Cannabis sativa for medical purposes. The THC-reduced hemp extract in this study is a promising candidate for medical applications.

Solid Lipid Nanoparticles (SLN) and Nanostructured Lipid Carriers (NLC) are receiving increasing interest as an approach to encapsulate natural extracts to increase the physicochemical stability of bioactives. Cannabis extract-derived cannabidiol (CBD) has potent therapeutic properties, including anti-inflammatory, antioxidant, and neuroprotective properties. In this work, physicochemical characterization was carried out after producing Compritol-based nanoparticles (cSLN or cNLC) loaded with CBD. Then, the determination of the encapsulation efficiency (EE), loading capacity (LC), particle size (Z-Ave), polydispersity index (PDI), and zeta potential were performed. Additionally, the viscoelastic profiles and differential scanning calorimetry (DSC) patterns were recorded. As a result, CBD-loaded SLN showed a mean particle size of 217.2 ± 6.5 nm, PDI of 0.273 ± 0.023, and EE of about 74%, while CBD-loaded NLC showed Z-Ave of 158.3 ± 6.6 nm, PDI of 0.325 ± 0.016, and EE of about 70%. The rheological analysis showed that the loss modulus for both lipid nanoparticle formulations was higher than the storage modulus over the applied frequency range of 10 Hz, demonstrating that they are more elastic than viscous. The crystallinity profiles of both CBD-cSLN (90.41%) and CBD-cNLC (40.18%) were determined. It may justify the obtained encapsulation parameters while corroborating the liquid-like character demonstrated in the rheological analysis. Scanning electron microscopy (SEM) study confirmed the morphology and shape of the developed nanoparticles. The work has proven that the solid nature and morphology of cSLN/cNLC strengthen these particles’ potential to modify the CBD delivery profile for several biomedical applications.

The high interest in non-psychoactive cannabidiol increases the need for efficient and straightforward cannabidiol (CBD) extraction methods. The research aimed to compare simple methods of cannabinoid extraction that do not require advanced laboratory equipment. This work assesses the content of total CBD and Δ9-tetrahydrocannabinol (Δ9-THC) in popular solvents such as water and ethanol extracts. Hemp raw material was analyzed with Gas Chromatography with a Flame Ionization Detector (GC-FID), while extracts were tested by High-Performance Liquid Chromatography (HPLC). The female inflorescences of three varieties of industrial hemp were tested: Futura 75, KC Dora, and Tygra (different sowing and N fertilization densities). Tygra (T/10/30) showed the highest content of CBD (0.064%) in water extracts. However, in 80% tincture from Futura 75 (F/30/30), a higher CBD content of 1.393% was observed. The use of 96% ethanol for extraction and ultrasound enabled the highest CBD content to be obtained: 2.682% in Futura 75 (F/30/30). Cold water extraction showed no effect on Δ9-THC content, while hot water extraction increased content from 0.001% in KC Dora to 0.002% in Futura 75 (F/30/30) and Tygra, but the changes were statistically insignificant. Application of 80% ethanol revealed the significantly highest content of Δ9-THC in KC Dora, from 0.026% (K/30/90) to 0.057% (K/30/30), as well as in Tygra (T/30/30) (0.036%) and Futura 75 (F/30/30) (0.048%). The use of ethanol extraction in combination with ultrasound could be an efficient method of obtaining cannabinoids.

Industrial hemp (Cannabis sativa L., family Cannabaceae) is a multi-purpose crop, used in the production of food, nutraceuticals, cosmetics and medicines. Therefore, development of new varieties with specific chemical profiles is necessary. In vitro culture methods could be complementary to conventional breeding and a useful tool for large-scale propagation. Strong apical dominance is considered as one of the factors contributing to the recalcitrance of industrial hemp in shoot proliferation. In this study, we tested the polar transport inhibitors N-1-naphtylphtalamic acid (NPA) and 2,3,5-triiodobenzoic acid (TIBA) to enhance shoot regeneration as the result of suppression of apical dominance and to develop in vitro protocols for Diana, Finola and Fedora 17 cultivars. Shoot tips derived from epicotyls were cultivated on Murashige and Skoog medium (MS) supplemented with meta-topolin (mT) and NPA, and also thidiazuron (TDZ) with a combination of TIBA and NPA. The results showed that the combination of TDZ with NPA (1–5 mg L−1) and TDZ with TIBA (0.5–2.5 mg L−1) increased the response of explants and the multiplication rate, but the effect was genotype-dependent and malformations were observed. To optimize the developed protocol, a two-step procedure with shortened time of exposure to inhibitors and reduced concentrations of them was applied. Shoots were rooted on media containing indole-3-butyric acid (IBA) and then successfully acclimatized. The obtained results will be useful in micropropagation of recalcitrant industrial hemp varieties.