Quality Evaluation of Tremella fuciformis Polysaccharides from Different Sources / 中国实验方剂学杂志

Objective:To establish a method for the determination of polysaccharide and monosaccharide composition of Tremella fuciformis, and to analyze the difference of polysaccharide content in T. fuciformis from different sources and cultivation methods, so as to provide reference for the quality determination.Method:High performance size exclusion chromatography coupled with multi-angle laser light scattering and refractive index detection (HPSEC-MALLS-RID) was employed to determine the content and relative molecular weight distribution of T. fuciformis polysaccharides. The monosaccharide types and proportions of T. fuciformis polysaccharides were analyzed by 1-phenyl-3-methyl-5-pyrazolone (PMP) precolumn derivative high performance liquid chromatography (HPLC).Result:The weight-average relative molecular weight (Mw) and the content of polysaccharides in T. fuciformis cultivated by cut-log from different sources were distributed in 2.618×106-3.503×106 Da and 307.12-609.06 g·kg-1, respectively. These two parameters of polysaccharides in T. fuciformis with substitute cultivation from different sources were 2.723×106-3.886×106 Da and 366.38-647.37 g·kg-1, respectively. The T. fuciformis polysaccharides mainly consisted of mannose, glucuronic acid, glucose, galactose, xylose and fucose, their ratios in samples with cut-log and substitute cultivation were 4.4∶0.7∶1.0∶0.2∶1.4∶1.6 and 4.4∶0.8∶1.0∶0.1∶1.5∶1.5, respectively. The contents of the above six monosaccharides in 39 batches of T. fuciformis from different sources were mannose of 36.71-191.31 g·kg-1, glucose of 10.46-76.10 g·kg-1, galactose of 1.00-6.72 g·kg-1, xylose of 16.73-70.54 g·kg-1, glucuronic acid of 9.74-32.12 g·kg-1, fucose of 17.16-68.20 g·kg-1.Conclusion:The content of polysaccharides in T. fuciformis from different sources has a certain difference, the developed method can be used as a routine method for the quality evaluation of polysaccharides in T. fuciformis.

An Enzymolysis-Assisted Agrobacterium tumefaciens-Mediated Transformation Method for the Yeast-Like Cells of Tremella fuciformis

Agrobacterium tumefaciens-mediated transformation (ATMT), as a simple and versatile method, achieves successful transformation in the yeast-like cells (YLCs) of Tremella fuciformis with lower efficiency. Establishment of a more efficient transformation system of YLCs is important for functional genomics research and biotechnological application. In this study, an enzymolysis-assisted ATMT method was developed. The degradation degree of YLCs depends on the concentration and digestion time of Lywallzyme. Lower concentration (≤0.1%) of Lywallzyme was capable of formation of limited wounds on the surface of YLCs and has less influence on their growth. In addition, there is no significant difference of YLCs growth among groups treated with 0.1% Lywallzyme for different time. The binary vector pGEH under the control of T. fuciformis glyceraldehyde-3-phosphate dehydrogenase gene (gpd) promoter was utilized to transform the enzymolytic wounded YLCs with different concentrations and digestion time. The results of PCR, Southern blot, quantitative real-time PCR (qRT-PCR) and fluorescence microscopy revealed that the T-DNA was integrated into the YLCs genome, suggesting an efficient enzymolysis-assisted ATMT method of YLCs was established. The highest transformation frequency reached 1200 transformants per 106 YLCs by 0.05% (w/v) Lywallzyme digestion for 15 min, and the transformants were genetically stable. Compared with the mechanical wounding methods, enzymolytic wounding is thought to be a tender, safer and more effective method.

An Improved Total RNA Extraction Method for White Jelly Mushroom Tremella fuciformis Rich in Polysaccharides

An improved method for extracting high quality and quantity RNA from a jelly mushroom and a dimorphic fungus—Tremella fuciformis which is especially rich in polysaccharides, is described. RNA was extracted from T. fuciformis mycelium M1332 and its parental monokaryotic yeast-like cells Y13 and Y32. The A260/280 and A260/230 ratios were both approximately 2, and the RNA integrity number was larger than 8.9. The yields of RNA were between 108 and 213 µg/g fresh wt. Downstream molecular applications including reverse transcriptional PCR and quantitative real-time PCR were also performed. This protocol is reliable and may be widely applicable for total RNA extraction from other jelly mushrooms or filamentous fungi rich in polysaccharides.

Protection of phosphate esterification polysaccharides from Tremella fuciformis on hematopoietic function in radiation-injured mice / 中草药

Objective: To study the effect of phosphate esterification polysaccharides from Tremella fuciformis (PEPTF) on hematopoietic function in radiation-injured mice. Methods: Rats were randomly divided into six groups: control, model, polysaccharides from T. fuciformis (10 mg/kg) positive control, low-, mid-, and high-PEPTF (10, 30, and 60 mg/kg) groups. Before irradiated with 137Cs γ-ray, rats were ip administered once daily for 3 d. On the day 7 after irradiation, the rats were sacrificed. Colony-forming unit of spleen, number of nucleated cells in bone marrow, number of white blood cells, spleen index, and thymus index were used as observation indexes to investigate the effect of PEPTF on hematopoietic function of mice. Results: Compared with the model group, the number of the nucleated cells in bone marrow, the number of white blood cells, spleen index, and thymus index of mice increased markedly. Conclusion: PEPTF have the protective effect on the hematopoietic function of radiation-injured mice.

Análise preliminar do tratamento de Diabetes mellitus com Polissacarídeo de Tremella fuciformis / Preliminary analysis of the treatment of Diabetes mellitus with Tremella fuciformis polysaccharide

Introdução: O Diabetes mellitus é uma desordem metabólica que ocorre no mundo inteiro, caracterizada por hiperglicemia associada a problemas com secreção ou ação da insulina ou, ainda por alterações no metabolismo intermediário de carboidratos, proteína e lipídeos. O cogumelo Tremella fuciformis (Basideomycota) foi isolado no Brasil e levado para a China onde tem sido produzido e utilizado como alimento por apresentar diversas atividades biológicas como anti-inflamatório, anti-mutagênico, reduz hipertensão, reduz diabetes entre outras propriedades relacionadas com imunidade. A Embrapa de Brasília tornou isolar o cogumelo e enviou a universidade para os testes preliminares. Objetivo: O presente trabalho teve como objetivo induzir a diabetes em ratos e realizar tratamento com polissacarídeo (PS) extraído do cogumelo gelatinoso Tremella fuciformis, desenvolvido em frascos no laboratório. Método: Ratos, no total de 28 animais foram divididos em 4 grupos sendo: 1-Normal) grupo recebendo dieta normal e água; 2-Sacarose) grupo recebendo ração normal e uma solução de sacarose 20% por 65 dias; 3-Sacarose + PS) grupo recebendo ração normal + solução de sacarose (65dias) e por gavagem PS na concentração de 1mmol (nos últimos 20 dias sendo uma vez/semana) ; 4-idem ao grupo 3, entretanto PS com concentração de 2mmol. No período, os ratos (uma vez/semana) foram pesados e submetidos a teste de glicose (fita Accu Check) e, depois sacrificados com dose letal de analgésico e coleta do sangue para testes de glicose, colesterol, triglicérides, creatinina e HDL. Resultados: Os resultados parciais demonstraram que os animais submetidos à dieta com sacarose aumentaram o peso e glicemia. Já os animais submetidos a tratamento com o PS permaneceram com o peso praticamente constante e, redução da glicemia. A creatinina tem sido um parâmetro para diagnosticar problemas renais, mas os animais em todos os tratamentos, não apresentaram diferença estatística. Para colesterol os animais submetidos ao tratamento com o PS apresentaram valores abaixo dos encontrados no grupo com sacarose, entretanto, com triglicérides e HDL apresentaram diminuição, mas houve, aumento em relação aos animais do grupo normal, indicando controle nos lipídeos. Por conclusão os resultados preliminares demonstraram que o PS extraído do cogumelo Tremella fuciformis apresenta grande potencial como produto natural contra diabetes.

Quality control of Tremella fuciformis polysaccharide by HPGPC / 中草药

Objective: To develop a high performance gel permeation chromatography (HPGPC) method for quality control of Tremella fuciformis polysaccharide. Methods: The separations were carried out with two series-wound columns including Shodex Sugar KS-804 (300 mm x 8 mm, 7 μm) and KS-805 (300 mm x 8 mm, 17 μm), water as mobile phase at flow rate 0.8 mL/min, column temperature at 40°C, injection volume 40 μL, and differential index detector. Results: T. fuciformis polysaccharide peak was located within 14.40-15.30 min and corresponding molecular weight was 1.26 × 106-1.39 × 106. The linearity was good during 0.01-2.0 mg/mL (r = 0.999). The polysaccharide concentration was 1.1-1.5 mg/mL and 0.8-1.1 mg/mL with precipitation of T. fuciformis polysaccharide and blue dextran as reference, respectively. LOD and LOQ were 0.8 and 2 μg, respectively. Conclusion: The method is simple, quick, and accurate, and has good repeatability and high sensitivity, which is fitful for quality control.

The Neuroprotective and Neurotrophic Effects of Tremella fuciformis in PC12h Cells

We examined the neuroprotective and neurotrophic effects of Tremella fuciformis. The neurotrophic effects of the hot water extract of T. fuciformis was evaluated by microscopically monitoring its potency to induce neurite outgrowth in PC12h cells. The hot water extract of T. fuciformis promoted neurite outgrowth in PC12h cells in this study, superior to other natural substances which was reported previously. When cells were treated with the hot water extract of T. fuciformis prior to beta-amyloid peptide treatment (active domain of A peptide 25~35 treated), toxicity was significantly diminished (p<0.01). These results suggest that T. fuciformis might potentially be used as a precautionary agent in neurodegenerative disease, such as Alzheimer's disease, etc.