Plant cells contain a wide range of interesting secondary metabolites, which are used as natural pigments and flavoring agents in foods and cosmetics as well as phyto‐pharmaceutical products. However, conventional industrial extraction from whole plants or parts of them is limited due to environmental and geographical issues. The production of secondary metabolites from in vitro cultures can be considered as alternative to classical technologies and allows a year‐round cultivation in the bioreactor under optimal conditions with constant high‐level quality and quantity. Compared to plant cell suspensions, differentiated plant in vitro systems offer the advantage that they are genetically stable. Moreover, the separation of the biomass from culture medium after fermentation is much easier. Nevertheless, several investigations in the literature described that differentiated plant in vitro systems are instable concerning the yield of the target metabolites, especially in submerged cultivations. Other major problems are associated with the challenges of cultivation conditions and bioreactor design as well as upscaling of the process. This article reviews bioreactor designs for cultivation of differentiated plant in vitro systems, secondary metabolite production in different bioreactor systems as well as aspects of process control, management, and modeling and gives perspectives for future cultivation methods.
The process of galanthamine and related alkaloids production by Leucojum aestivum shoot culture in a temporary immersion system was studied. It was established that temporary immersion approach is prospective for development of a biosynthetic process for obtaining valuable Amaryllidaceae alkaloids. Both immersion frequency and temperature had significant effect on biomass accumulation and the yields of galanthamine and related alkaloids. The maximal yield of galanthamine was achieved at the cultivation of L. aestivum shoot culture in temporary immersion RITA ® system at immersion frequency 15 min flooding and 8 h stand-by periods, at 26°C. Data on the relationships in the biological system "Nutrient medium-L. aestivum shoot culture-galanthamine" are presented as well.
Shoot culture of summer snowflake (Leucojum aestivum L.) was successfully cultivated in an advanced modified glass‐column bioreactor with internal sections for production of Amaryllidaceae alkaloids. The highest amounts of dry biomass (20.8 g/L) and galanthamine (1.7 mg/L) were achieved when shoots were cultured at 22°C and 18 L/(L·h) flow rate of inlet air. At these conditions, the L. aestivum shoot culture possessed mixotrophic‐type nutrition, synthesizing the highest amounts of chlorophyll (0.24 mg/g DW (dry weight) chlorophyll A and 0.13 mg/g DW chlorophyll B). The alkaloids extract of shoot biomass showed high acetylcholinesterase inhibitory activity (IC50 = 4.6 mg). The gas chromatography–mass spectrometry (GC/MS) profiling of biosynthesized alkaloids revealed that galanthamine and related compounds were presented in higher extracellular proportions while lycorine and hemanthamine‐type compounds had higher intracellular proportions. The developed modified bubble‐column bioreactor with internal sections provided conditions ensuring the growth and galanthamine production by L. aestivum shoot culture.
Hairy root cultures of Salvia tomentosa were initiated by transformation with Agrobacterium rhizogenes. To prevent necrosis in the explants and to protect young hairy roots, Amberlite XAD-4 resin, in combination with a temporary immersion cultivation system, was applied. HPLC analyzes showed that the resin adsorbed more than 93% of the released phenolic acids and 100% of the released flavonoids. The decreased content of the released phenolics significantly reduced their destructive effects on the plant tissues, prevented, and speeded up the appearance of hairy roots.
The Amaryllidaceae alkaloid galanthamine (Gal) is a long‐acting, selective, reversible, and competitive acetylcholinesterase inhibitor used for the treatment of early‐ to mid‐stage Alzheimer's disease, poliomyelitis, and other neurological diseases. Currently, Gal is produced by extraction from plants such as daffodils (Narcissus cultivar Carlthon), snowflake (Leucojum aestivum), “red‐tubed lily” (Lycoris radiata), and Ungernia victoria, and alternatively by chemical synthesis. Due to the increased demand by the generic pharmaceutical companies and the limited availability of plant sources, the biosynthesis of Gal by plant in vitro systems has been considered as an alternative approach for its sustainable production. The present article reviews the state of the art of in vitro Gal biosynthesis including growth regulators, medium components, culture conditions, elicitation, and bioreactor systems. It may be used as a starting point for further studies in this area leading to a progress in biotechnological production of this valuable alkaloid.
Nettle (Urtica dioica L.), tansy (Tanacetum vulgare L.), bladder campion (Silene vulgaris (Moench) Garcke, waterpepper (Polygonum hydropiper L.), common centaury (Centaurium erythraea Pers.) and rose hip fruit (Rosa canina L. cv. Plovdiv 1) were used for preparation of different water extracts (infusion, decoction and microwave extract) and ethanol (tincture) extracts. Carotenoids (lutein, lycopene and β-carotene), tocopherols (α-, γ- and δ-), organic acids (ascorbic, malic, fumaric and citric), five macro- and three microelements, sugars and uronic acids content in the obtained extracts were analyzed. Among the investigated plants, stinging nettle, bladder campion and rose hip fruit were evaluated as most potential with respect to bioactive compounds and microelements. The results showed that the selected six medicinal plants and their extracts can be presented as sources of dietary fibers and micronutrients, which may encourage further application as food supplements and beverages as well as to motivate plant use as a dietary alternative in different foods. The present study is a first detailed analysis with respect to sugar content of decoction, infusion and tincture of S. vulgaris.
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