Improving Flavonoid Accumulation of Bioreactor-Cultured Adventitious Roots in Oplopanax elatus Using Yeast Extract.

Oplopanax elatus is an endangered medicinal plant, and adventitious root (AR) culture is an effective way to obtain its raw materials. Yeast extract (YE) is a lower-price elicitor and can efficiently promote metabolite synthesis. In this study, the bioreactor-cultured O. elatus ARs were treated with YE in a suspension culture system to investigate the elicitation effect of YE on flavonoid accumulation, serving for further industrial production. Among YE concentrations (25-250 mg/L), 100 mg/L YE was the most suitable for increasing the flavonoid accumulation. The ARs with various ages (35-, 40-, and 45-day-old) responded differently to YE stimulation, where the highest flavonoid accumulation was found when 35-day-old ARs were treated with 100 mg/L YE. After YE treatment, the flavonoid content increased, peaked at 4 days, and then decreased. By comparison, the flavonoid content and antioxidant activities in the YE group were obviously higher than those in the control. Subsequently, the flavonoids of ARs were extracted by flash extraction, where the optimized extraction process was: 63% ethanol, 69 s of extraction time, and a 57 mL/g liquid-material ratio. The findings provide a reference for the further industrial production of flavonoid-enriched O. elatus ARs, and the cultured ARs have potential application for the future production of products.

Optimizing elicitation strategy of salicylic acid for flavonoid and phenolic production of fed-batch cultured Oplopanax elatus adventitious roots.

Oplopanax elatus is a valuable medicinal plant, but its plant resource is lacking. Adventitious root (AR) culture of O. elatus is an effective way for the production of plant materials. Salicylic acid (SA) exerts enhancement effect on metabolite synthesis in some plant cell/organ culture systems. To clarify the elicitation effect of SA on fed-batch cultured O. elatus ARs, this study investigated the effects of SA concentration, and elicitation time and duration. Results showed that flavonoid and phenolic contents, and antioxidant enzyme activity obviously increased when the fed-batch cultured ARs were treated with 100 µM SA for 4 days starting on day 35. Under this elicitation condition, total flavonoid and phenolic contents reached 387 rutin mg/g DW and 128 gallic acid mg/g DW, respectively, which were significantly (p < 0.05) higher than those in the SA-untreated control. In addition, DPPH scavenging and ABTS+ scavenging rates, and Fe2+ chelating rate also greatly increased after SA treatment, and their EC50 values were 0.0117, 0.61, and 3.34 mg/L, respectively, indicating the high antioxidant activity. The findings of the present study revealed that SA could be used as an elicitor to improve the flavonoid and phenolic production in fed-batch O. elatus AR culture.

Hepatoprotective Effect of Oplopanax elatus Nakai Adventitious Roots Extract by Regulating CYP450 and PPAR Signaling Pathway.

O. elatus Nakai is a traditional medicine that has been confirmed to exert effective antioxidant and anti-inflammatory functions, and is used for the treatment of different disorders. However, its potential beneficial effects on drug induced hepatotoxicity and relevant molecular mechanisms remain unclear. This study investigated the protective effect and further elucidated the mechanisms of action of O. elatus on liver protection. O. elatus chlorogenic acids-enriched fraction (OEB), which included chlorogenic acid and isochlorogenic acid A, were identified by HPLC-MS/MS. OEB was administrated orally daily for seven consecutive days, followed by a single intraperitoneal injection of an overdose of APAP after the final OEB administration. The effects of OEB on immune cells in mice liver were analyzed using flow cytometry. APAP metabolite content in serum was detected using HPLC-MS/MS in order to investigate whether OEB affects CYP450 activities. The intestinal content samples were processed for 16 s microbiota sequencing. Results demonstrated that OEB decreased alanine aminotransferase, aspartate aminotransferase contents, affected the metabolism of APAP, and decreased the concentrates of APAP, APAP-CYS and APAP-NAC by inhibiting CYP2E1 and CYP3A11 activity. Furthermore, OEB pretreatment regulated lipid metabolism by affecting the peroxisome proliferator-activated receptors (PPAR) signaling pathway in mice and also increased the abundance of Akkermansia and Parabacteroides. This study indicated that OEB is a potential drug candidate for treating hepatotoxicity because of its ability to affect drug metabolism and regulate lipid metabolism.

Analysis of Differentially Expressed Genes in Tissues of Camellia sinensis during Dedifferentiation and Root Redifferentiation.

Tissue culture is very important for identifying the gene function of Camellia sinensis (L.) and exploiting novel germplasm through transgenic technology. Regeneration system of tea plant has been explored but not been well established since the molecular mechanism of tea plant regeneration is not clear yet. In this study, transcriptomic analysis was performed in the initial explants of tea plant and their dedifferentiated and redifferentiated tissues. A total of 93,607 unigenes were obtained through de novo assembly, and 7,193 differentially expressed genes (DEGs) were screened out from the 42,417 annotated unigenes. Much more DEGs were observed during phase transition rather than at growth stages of callus. Our KOG and KEGG analysis, and qPCR results confirmed that phase transition of tea plant was closely related to the mechanism that regulate expression of genes encoding the auxin- and cytokinin-responsive proteins, transcription factor MYB15 and ethylene-responsive transcription factor ERF RAP2-12. These findings provide a reliable foundation for elucidating the mechanism of the phase transition and may help to optimize the regeneration system by regulating the gene expression pattern.

Different Catabolism Pathways Triggered by Various Methylxanthines in Caffeine-Tolerant Bacterium Pseudomonas putida CT25 Isolated from Tea Garden Soil.

The degradation efficiency and catabolism pathways of the different methylxanthines (MXs) in isolated caffeine-tolerant strain Pseudomonas putida CT25 were comprehensively studied. The results showed that the degradation efficiency of various MXs varied with the number and position of the methyl groups on the molecule (i.e., xanthine > 7-methylxanthine ≈ theobromine > caffeine > theophylline > 1-methylxanthine). Multiple MX catabolism pathways coexisted in strain CT25, and a different pathway would be triggered by various MXs. Demethylation dominated in the degradation of N-7-methylated MXs (such as 7-methylxanthine, theobromine, and caffeine), where C-8 oxidation was the major pathway in the catabolism of 1-methylxanthine, whereas demethylation and C-8 oxidation are likely both involved in the degradation of theophylline. Enzymes responsible for MX degradation were located inside the cell. Both cell culture and cell-free enzyme assays revealed that N-1 demethylation might be a rate-limiting step for the catabolism of the MXs. Surprisingly, accumulation of uric acid was observed in a cell-free reaction system, which might be attributed to the lack of activity of uricase, a cytochrome c-coupled membrane integral enzyme.