Remarkable enhancement of flavonoid production in a co-cultivation system of Isatis tinctoria L. hairy root cultures and immobilized Aspergillus niger.

The dried roots of Isatis tinctoria L. are highly traded in the pharmaceutical industry due to their notable anti-influenza efficacy. For the first time, I. tinctoria hairy root cultures (ITHRCs) were co-cultured with two immobilized live GRAS (Generally Recognized as Safe) fungi, i.e. Aspergillus niger and Aspergillus niger, for the elevated production of pharmacologically active flavonoids. Immobilized A. niger (IAN) was exhibited as the superior elicitor in the plant-fungus co-cultivation system. The highest flavonoid production (3018.31 ± 48.66 µg/g DW) were achieved in IAN-treated ITHRCs under the optimal conditions of IAN spore concentration ca.104 spores/mL, temperature 30 °C, initial pH value of media 7.0 and time 72 h, which remarkably increased 6.83-fold relative to non-treated control (441.91 ± 7.35 µg/g DW). Also, this study revealed that IAN elicitation could trigger the sequentially transient accumulation of signal molecules and intensify the oxidative stress in ITHRCs, which both contributed to the up-regulated expression of associated genes involved in flavonoid biosynthetic pathway. Moreover, IAN could be reused at least five cycles with satisfactory performance. Overall, the coupled culture of IAN and ITHRCs is a promising and effective approach for the enhanced production of flavonoids, which allows for the improved applicability of these valuable compounds in pharmaceutical fields.

Ultraviolet radiation for flavonoid augmentation in Isatis tinctoria L. hairy root cultures mediated by oxidative stress and biosynthetic gene expression.

Search of cost-effective strategies that can enhance the accumulation of phytochemicals of pharmaceutical interest in plant in vitro cultures is an essential task. For the first time, Isatis tinctoria L. hairy root cultures were exposed to ultraviolet radiation (ultraviolet-A, ultraviolet-B, and ultraviolet-C) in an attempt to promote the production of pharmacologically active flavonoids. Results showed that the maximum flavonoid accumulation (7259.12 ±â€¯198.19 µg/g DW) in I. tinctoria hairy root cultures treated by 108 kJ/m2 dose of UV-B radiation increased 16.51-fold as compared with that in control (439.68 ±â€¯8.27 µg/g DW). Additionally, antioxidant activity enhancement and cell wall reinforcement were found in the treated I. tinctoria hairy root cultures, indicating the positive-feedback responses to oxidative stress mediated by ultraviolet-B radiation. Moreover, the expression of chalcone synthase gene was tremendously up-regulated (up to 405.84-fold) in I. tinctoria hairy root cultures following ultraviolet-B radiation, which suggested chalcone synthase gene might play a crucial role in flavonoid augmentation. Overall, the present work provides a feasible approach for the enhanced production of biologically active flavonoids in I. tinctoria hairy root cultures via the simple supplementation of ultraviolet-B radiation, which is useful for the biotechnological production of these high-added value compounds to fulfil the ever-increasing demand in pharmaceutical fields.

Enhanced Production of Two Bioactive Isoflavone Aglycones in Astragalus membranaceus Hairy Root Cultures by Combining Deglycosylation and Elicitation of Immobilized Edible Aspergillus niger.

A cocultivation system of Astragalus membranaceus hairy root cultures (AMHRCs) and immobilized food-grade fungi was established for the enhanced production of calycosin (CA) and formononetin (FO). The highest accumulations of CA (730.88 ± 63.72 µg/g DW) and FO (1119.42 ± 95.85 µg/g DW) were achieved in 34 day-old AMHRCs cocultured with immobilized A. niger (IAN) for 54 h, which were 7.72- and 18.78-fold higher than CA and FO in nontreated control, respectively. IAN deglycosylation could promote the formation of CA and FO by conversion of their glycoside precursors. IAN elicitation could intensify the generation of endogenous signal molecules involved in plant defense response, which contributed to the significantly up-regulated expression of genes in CA and FO biosynthetic pathway. Overall, the coupled culture of IAN and AMHRCs offered a promising and effective in vitro approach to enhance the production of two health-promoting isoflavone aglycones for possible nutraceutical and pharmaceutical uses.

Degradation of lignin in birch sawdust treated by a novel Myrothecium verrucaria coupled with ultrasound assistance.

Combined treatment of a novel fungal endophyte Myrothecium verrucaria coupled with ultrasound assistance was conducted to enhance lignin degradation in birch sawdust. The optimum treatment conditions were confirmed as the materials to liquid ratio 1:20, temperature 30°C, time 4days and pH 7, respectively. The results showed that the combined treatment led to the lignin degradation reaching 67.95±2.14%, while the lignin degradation were 45.50±2.12% and 13.75±0.66% with separate fungal treatment and ultrasound treatment, respectively. Moreover, SEM and FTIR analysis indicated that combined treatment significantly altered surface morphology and chemical structure of birch sawdust. The combined treatment greatly increased lignin removal during short time in mild environment. Therefore, these results demonstrated that the combined treatment of fungal endophyte coupled with ultrasound assistance has the high potential for the removal lignin in lignocellulose.

Production of Laccase by a New Myrothecium verrucaria MD-R-16 Isolated from Pigeon Pea [Cajanus cajan (L.) Millsp.] and its Application on Dye Decolorization.

The present study was conducted to screen a laccase-producing fungal endophyte, optimize fermentation conditions, and evaluate the decolorization ability of the laccase. A new fungal endophyte capable of laccase-producing was firstly isolated from pigeon pea and identified as Myrothecium verrucaria based on a ITS-rRNA sequences analysis. Meanwhile, various fermentation parameters on the laccase production were optimized via response surface methodology (RSM). The optimal fermentation conditions were a fermentation time of five days, temperature 30 °C and pH 6.22. Laccase activity reached 16.52 ± 0.18 U/mL under the above conditions. Furthermore, the laccase showed effective decolorization capability toward synthetic dyes (Congo red, Methyl orange, Methyl red, and Crystal violet) in the presence of the redox mediator ABTS, with more than 70% of dyes decolorizing after 24 h of incubation. Additionally, the activity of laccase was relatively stable with pH (4.5-6.5) and a temperature range of 35-55 °C. Therefore, the high laccase production of the strain and the new fungal laccase could provide a promising alterative approach for industrial and environmental applications.