HPLC quantitative analysis of protocatechuic acid contents in 11 Phellinus mushroom species collected in Thailand

Abstract Phellinus mushrooms have been traditionally used for various medicinal purposes. Protocatechuic acid, which was previously reported to be a component in some Phellinus mushrooms, has some pharmacological effects. This study aimed to validate a HPLC method for the quantitative analysis of the protocatechuic acid contents in the extracts from different Phellinus mushroom species collected in Thailand. HPLC was carried out using a C18 column and the gradient mobile phases of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. Method validation was performed to assure the linearity, accuracy, precision, limit of detection and limit of quantitation of the analytical method. The linearity range of protocatechuic acid was 1 - 10 µg/ml. The average recovery was 104.16%. The method was shown to be precise with the RSD of repeatability and intermediate precision at less than 3%. The protocatechuic contents in 11 Phellinus mushrooms were in the range of less than 0.0099 - 0.4121 %w/w of the extract. The developed HPLC method was reliable and suitable for the quantitative analysis of protocatechuic acid content in Phellinus mushrooms.

Screening for proteins related to the biosynthesis of hispidin and its derivatives in Phellinus igniarius using iTRAQ proteomic analysis.

BACKGROUND: Hispidin (HIP) and its derivatives, a class of natural fungal metabolites, possess complex chemical structures with extensive pharmacological activities. Phellinus igniarius, the most common source of HIP, can be used as both medicine and food. However, the biosynthetic pathway of HIP in P. igniarius remains unclear and we have a limited understanding of the regulatory mechanisms related to HIP. In this work, we sought to illustrate a biosynthesis system for hispidin and its derivatives at the protein level. RESULTS: We found that tricetolatone (TL) is a key biosynthetic precursor in the biosynthetic pathway of hispidin and that its addition led to increased production of hispidin and various hispidin derivatives. Based on the changes in the concentrations of precursors and intermediates, key timepoints in the biosynthetic process were identified. We used isobaric tags for relative and absolute quantification (iTRAQ) to study dynamic changes of related proteins in vitro. The 270 differentially expressed proteins were determined by GO enrichment analysis to be primarily related to energy metabolism, oxidative phosphorylation, and environmental stress responses after TL supplementation. The differentially expressed proteins were related to ATP synthase, NAD binding protein, oxidoreductase, and other elements associated with electron transfer and dehydrogenation reactions during the biosynthesis of hispidin and its derivatives. Multiple reaction monitoring (MRM) technology was used to selectively verify the iTRAQ results, leading us to screen 11 proteins that were predicted to be related to the biosynthesis pathways. CONCLUTION: These findings help to clarify the molecular mechanism of biosynthesis of hispidin and its derivatives and may serve as a foundation for future strategies to identify new hispidin derivatives.

The phytochemistry and pharmacology of medicinal fungi of the genus Phellinus: a review.

Phellinus Quél is one of the largest genera of Hymenochaetaceae, which is comprised of about 220 species. Most Phellinus macro-fungi are perennial lignicolous mushrooms, which are widely distributed on Earth. Some Phellinus fungi are historically recorded as traditional medicines used to treat various diseases in eastern Asian countries, especially China, Japan and Korean. Previous phytochemical studies have revealed that Phellinus fungi produce diverse secondary metabolites, which mainly contain polysaccharides, flavones, coumarins, terpenes, steroids, and styrylpyranones. Pharmacological documents have demonstrated that Phellinus mushrooms and their compounds have a variety of bioactivities, such as anti-tumor, immunomodulation, anti-oxidative and anti-inflammation, anti-diabetes, neuro-protection, and anti-viral effects. This review surveys the literature reporting the isolation, characterization, and bioactivities of secondary metabolites from the fungi of the genus Phellinus, focusing on studies published in the literature up to April 2020. Herein, a total of more than 300 compounds from 13 Phellinus species and their isolation, characterization, chemistry, pharmacological activities, and relevant molecular mechanisms are comprehensively summarized.

Influences of carbon and nitrogen source addition, water content, and initial pH of grain medium on hispidin production of Phellinus linteus by solid-state fermentation.

This study aimed to evaluate the effects of certain grain medium conditions, such as carbon and nitrogen source addition, water content, and initial pH, on the hispidin production of Phellinus linteus using solid-state fermentation. The results showed that the highest hispidin yield, specific productivity of hispidin, and total content of hispidin were associated with sucrose and malt extract addition, 55% water content, and an initial pH of 5.5, which resulted in 0.290, 0.233, 0.301, and 0.296 mg/g dry weight of mycelium; 0.261, 0.191, 0.257, and 0.227 µg/mg week-1 specific productivity; and 46.01, 40.67, 58.85, and 55.06 mg/kg dry weight brown rice medium, respectively. Additionally, two combinations of culture conditions with pearl barley and black rice medium were tested for hispidin production. The highest hispidin yield, specific productivity of hispidin, and total hispidin content for pearl barley medium fermented using P. linteus were 3.88-fold, 4.86-fold, and 3.60-fold higher than those for the control (brown rice medium), respectively. Overall, this study shows that P. linteus hispidin production can be enhanced using solid-state fermentation, with optimal medium conditions.