Stoilkovska Gjorgievska, Veronika

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Stoilkovska Gjorgievska, Veronika
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Stoilkovska Gjorgievska, Veronika

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    Item type:Publication,
    Differentiation of Cannabis seeds employing digital morphological screening and infrared spectroscopy coupled with multivariate modeling
    (Elsevier, 2024-05-01)
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    Trajkovska, Ana
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    Cultivation of Cannabis for medicinal purposes primarily relies on seed propagation with expected variations in yield, cannabinoid content, growth rate and seed material non-uniformity. This study aims to employ digital methods for morphological analysis and infrared spectroscopy, combining them with multivariate analysis to characterize and differentiate Cannabis seeds. Morphological traits of 100 seeds from both commercial Cannabis specimens and wild-growing local varieties were analyzed using the high-throughput phenotyping software in addition to their collected infrared spectra in the mid-IR region. Subsequently, a chemometrics approach by means of Principal Component Analysis (PCA), Hierarchical Cluster Analysis (HCA), and Partial Least Square-Discriminatory Analysis (PLS-DA) was applied. The statistical indicators of the PLS-DA model (R2X=0.99, R2Y=0.63, Q2=0.64) demonstrate strong predictive capabilities for the differentiation of Cannabis seed specimens based on morphological attributes. The score scatter plot clearly shows a distinct grouping pattern, primarily driven by seed size. Wild-type seeds predominantly cluster into group 1, characterized by smaller diameters, while commercial seeds cluster into group 2. By analysing spectral data, in contrast to the expected differentiation based on secondary metabolites (cannabinoids) in the seeds, differentiation was based on the macronutrient profile with characteristic bands at 3275 cm−1, 2921 cm−1, 2852 cm−1, 1743 cm−1, 1630 cm−1, 1532 cm−1, 1459 cm−1, 1239 cm−1, 1157 cm−1, 1094 cm−1, 1018 cm−1, identified as the most distinctive spectral features. The PCA model (R2X=0.88 and Q2=0.85) was composed of 5 principal components explaining 88% of the spectral variations. The loading plot of the PLS-DA model reveals the distinctive spectral features for both groups (lipid and carbohydrate bands - group 2 samples, protein and water content - group 1 samples). The developed models have the potential for application for rapid screening of quality parameters of Cannabis seeds and their differentiation.
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    Item type:Publication,
    Cannabinoid production in various Cannabis sativa L. in vitro cultures
    (Springer Nature, 2025-01-02)
    Ana Trajkovska;
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    Oliver Tusevski;
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    Iskra Davkova;
    The main goal of this study was to develop an efficient micropropagation protocol for cannabis (Cannabis sativa L.) and to summarize the influence of plant growth regulators on cannabinoid production in various in vitro cultures. Three in vitro culture lines were obtained during cannabis micropropagation and referred to calli, shoots, and plantlets. Shoot multiplication was examined on apical segments isolated from in vitro seedlings and cultivated on solid MS/B5 medium supplemented with various concentrations (0.05-1.0 mg·L− 1) of thidiazuron (TDZ). Callogenesis was induced from leaf explants of seedlings in the presence of 1.0 mg·L− 1 TDZ and 0.5 mg·L− 1 1-naphthaleneacetic acid (NAA), while further biomass production was evaluated upon TDZ (0.5-2.0 mg·L− 1). The shoot regenerative potential was assessed on various concentrations (0.1-1.0 mg·L− 1) of indole-3-butyric acid (IBA). The HPLC-DAD analysis of cannabinoids revealed that cannabidiolic acid (CBDA), cannabidiol (CBD), cannabinol (CBN), ∆9-tetrahydrocannabinol (∆9-THC), and ∆9-tetrahydrocannabinolic acid (∆9-THCA) were the main identified cannabinoids in calli, shoots and regenerated plantlets. The superior production of Δ9-THC was observed in multiple shoots cultured on 0.05 mg·L− 1 TDZ (49.23 mg·g− 1), callus cultures treated with 1.0 mg·L− 1 TDZ (3.92 µg·g− 1), as well in plantlets regenerated on hormone-free medium (80.1 mg·g− 1). The Δ9-THC content in the leaves of 6-month-old regenerated plantlets was significantly higher (1.6-fold) than that of the mother plant. Cannabinoid contents in the inflorescences of the mother plant were higher or even comparable to those observed for acclimatized plantlets. In conclusion, cannabis in vitro cultures could be proposed as promising systems for cannabinoid production.