Selective Impact of MTMS-Based Xerogel Morphology on Boosted Proliferation and Enhanced Naphthoquinone Production in Cultures of Rindera graeca Transgenic Roots

Abstract

In situ extraction is a method for separating plant secondary metabolites from in vitro systems of plant biomass cultures. The study aimed to investigate the MTMS-based xerogels morphology effect on the growth kinetics and deoxyshikonin productivity in xerogel-supported in vitro culture systems of Rindera graeca hairy root. Cultures were supplemented with three types of xerogel, i.e., mesoporous gel, microporous gel, and agglomerated precipitate, in the disintegrated or monolithic form. Structure, oil sorption capacity, and SEM analyses for xerogel-based additives were performed. Application of monolithic macroporous xerogel resulted in the highest biomass proliferation, i.e., 5.11-fold fresh biomass increase after four weeks of the screening culture. The highest deoxyshikonin production (i.e., 105.03 µg) was noted when hairy roots were maintained with particles of disintegrated mesoporous xerogel. The detailed kinetics investigations (6-week culture) revealed the highest growth of hairy root biomass and secondary metabolite production, equaling 9.46-fold fresh weight biomass and 204.08 µg deoxyshikonin, respectively. MTMS-based xerogels have been recognized as selective biocompatible scaffolds for boosting the proliferation of transgenic roots or for productivity enhancement of naphthoquinones without detrimental effects on biomass growth, and their successful applicability in in situ removal of secondary plant metabolites has been experimentally confirmed.

Effect of Silica Xerogel Functionalization on Intensification of Rindera graeca Transgenic Roots Proliferation and Boosting Naphthoquinone Production

“…As we demonstrated previously [14,23], the application of MTMS-based xerogel materials (with methyl groups on the surface) can significantly enhance plant secondary metabolism with preserved growth ability. Another key aspect of the interaction between the plant biomass and applied biomaterial is the chemical structure of the latter, including the presence of specific chemical residues or substituents [29][30][31][32].…”

“…All samples were treated in a Sonorex ultrasonic scrubber (Bandelin, Berlin, Germany) to enhance the extraction efficiency until the solvent color faded. Obtained extracts were dried under reduced pressure (0.1 bar) using Rotavapor ® R-100 (Buchi, Flawil, Switzerland) and stored in a freezer at −20 • C. A detailed procedure used for secondary metabolite extraction from R. graeca transgenic roots is available in previous reports, e.g., in [14].…”

“…The TEOS-based materials used both directly for the cultivation of transgenic roots and for further functionalization were characterized by a uniform pore morphology of nanometric size (Figure 1), with a well-crosslinked and chemically uncomplicated structure of silicon oxide [37]. The small size of the pores results in the creation of high capillary pressure, which positively affects the kinetics of sorption, including the extraction of secondary metabolites [14].…”

“…Furthermore, the efficient production of plant-derived therapeutics from natural sources is often restricted by seasonal variability and geographical, ecological, or political restrictions [12]. The application of modern biotechnological techniques, i.e., in vitro cultures of plant organs, like roots or shoots, allows the maintenance of plant biomass independently from the aspects mentioned above, but with plentiful options of parameters optimization and the scaling-up of bioprocess, it allows the exceeding of productivity observed in wild plants [13][14][15]. One of the practical bioengineering techniques widely recognized as a high-potential solution for the intensification of secondary plant metabolite production is in situ extraction [16][17][18][19].…”

“…The impact of individual organic groups in the structure of experimental xerogel materials on plant biomass growth and secondary metabolite production is still not fully known. Due to that, in this study, we individually and independently introduced several chemical groups to the surface of silica-based xerogel as biomaterial that is non-toxic against plant biomass used in our previous studies [14,23]. The TEOS-based xerogel was used as a basic material for the implementation of functional groups on the surface of silica xerogel.…”