Lycium schweinfurthii, a wild shrub of the Solanaceae family, has received increasing attention in the last decade for its therapeutic potential in traditional medicine due to its diverse array of secondary metabolites, including phenolic substances and terpenoids. The aim of this study was to investigate the accumulation of phenolics, flavonoids, and the terpenoid lupeol in L. schweinfurthii cell suspension shake flask cultures and a single-use 2-dimensional rocking motion bioreactor. Three different media formulations were compared for in vitro cell cultures. Various parameters, such as biomass accumulation, settled cell volume, cell viability (assessed via a 2,3,5-triphenyl tetrazolium chloride assay), and sucrose consumption were determined as indicators of cell activity and growth. Total phenolic and flavonoid contents were estimated spectrophotometrically, lupeol was quantified via High-Performance Thin Layer Chromatography (HPTLC). Although a higher fresh biomass concentration of 464 g L− 1 was obtained in MS medium supplemented with a combination of each, 1 mg L− 1 of 2,4-Dichlorophenoxyacetic acid (2,4-D) and 1-Naphthaleneacetic acid (NAA), the rocking-motion bioreactor cultivation was performed with 2 mg L− 1 NAA due to its superior reproducibility in viability, productivity, and content of bioactive compounds (e.g., phenolics, flavonoids, lupeol). A final fresh biomass concentration of 185 g L− 1 was achieved in a 16 L cultivation scale with a notable increase in the concentration of phenolics (1.4-fold) and flavonoids (1.7-fold). Most importantly, the concentration of lupeol, a pentacyclic triterpenoid known for its anti-inflammatory, antibacterial, and anti-atherogenic properties, exhibited a remarkable 5.5-fold increase in the bioreactor cultivation (585 µg g− 1) compared to shake flask cultivations (106 µg g− 1). The current study demonstrated the profound impact of media composition and non-limited fed-batch conditions in a rocking-motion bioreactor on the accumulation of bioactive compounds. The findings are also relevant to other plant cell cultures.
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Oksana Sytar,
Diaa Mamdouh Ahmed Mohammed,
Iryna Smetanska
Lycium schweinfurthii is a traditional medicinal plant grown in the Mediterranean region. As it is used in folk medicine to treat stomach ulcers, it took more attention as a source of valuable secondary metabolites. The in vitro cultures of L. schweinfurthii could be a great tool to produce secondary metabolites at low costs. The presented study aimed to introduce and optimize a protocol for inducing callus and cell suspension cultures as well as estimating phenolic, flavonoid compounds, and antioxidant activity in the cultures of the studied species. Three plant growth regulators (PGRs) were supplemented to MS medium solely or in combination to induce callus from leaf explants. The combination between 2,4-dichlorophenoxy acetic acid (2,4-D) and 1-naphthyl acetic acid (NAA) induced callus in all explants regardless of the concentration. The highest fresh weight of callus (3.92 g) was obtained on MS medium fortified with 1 mg L−1 of both 2,4-D and NAA (DN1) after 7 weeks of culture. DN1 was the best medium for callus multiplication regarding the increase in fresh weight and size of callus. Otherwise, the highest phenolics, flavonoids, and antioxidant activity against DPPH free radicals were of callus on MS fortified with 2 mg L−1 NAA (N2). The cell suspension cultures were cultivated on a liquid N2 medium with different sucrose concentrations of 5–30 g L−1 to observe the possible effects on cells’ multiplication and secondary metabolite production. The highest fresh and viable biomass of 12.01 g was obtained on N2 containing 30 g L−1 sucrose. On the other hand, the cell cultures on N2 medium of 5 and 30 g L−1 sucrose produced phenolics and flavonoids, and revealed
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Diaa Mamdouh Ahmed Mohammed,
Ahmed M. M. Gabr,
Iryna Smetanska
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