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Analysis of Monosaccharide Composition of Polysaccharides in ganoderma atrum by Capillary Gas Chromatography

Analysis of Monosaccharide Composition of Polysaccharides in...

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中国食品科学技术学会第五届年会暨第四届东西方食品业高层论坛

作者： Yi Chen Ming-YongXie~* Shao-PingNie Chang Li Yuan-Xing Wang (State Key Laboratory of Food Science and Technology,Nanchang University,Nanchang 330047 People’s Republic of China)

A simple and sensitive gas chromatography(GC)-based method was developed for the analysis of monosaccharide composition of purified polysaccharides in ganoderma atrum (***).The polysaccharide was firstly hydrolyzed to give the constituent monosaccharides, which were subsequently derived into acetylated *** separation of the derivatized monosaccharides was efficiently carried out by GC,using a capillary column packed with a (5%phenyl)methylpolysiloxane stationary *** concentrations of monosaccharides were determined based on the measured peak area *** stable derivatives of the most common monosaccharides of these glycoconjugates(Ara,Rha,Xyl,Fuc,Gal,Man,GIc,and Gala)could be separated and quantitatively and reproducibly determined with a high degree of sensitivity level in the presence of acetyl inositol as the inner *** method fulfilled the established criteria for *** limits of detection(LOD)and quantification(LOQ)were down to 0.013 mg/mL and 0.043 mg/mL *** intermediary precision values(expressed as RSD) were less than 10%.The mean recovery of the method was 100+3%,with the RSD values of less than 5%.In order to prove the validity of the above method,high performance liquid chromatography(HPLC)was also used to compare with *** determined concentrations of monosaccharides obtained from the two chromatographic methods were found close to each other within acceptable error *** study demonstrated that the developed method could normally be recommended for oligosaccharide analysis,and could also be used in the reproducible quantification of monosaccharides in biological and biochemical laboratories,where HPLC chromatography or MS method may not be readily available.

关键词： ganoderma atrum polysaccharide monosaccharide composition acetylated aldononitriles gas chromatography

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Expression,purification and functional characteristic of a novel fungal immunomodulatory protein from ganoderma atrum

Expression,purification and functional characteristic of a n...

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中国菌物学会2015年学术年会

作者： Hui Xu Ying-Yu Kong Wei-Ran Cong Xin-Ping Zhang Xuan-Wei Zhou School of Agriculture and Biology Shanghai Jiao Tong University Key Laboratory of Urban Agriculture（South） Ministry of Agriculture Plant Biotechnology Research CenterSchool of Agriculture and BiologyShanghai Jiaotong University Engineering Research Center of Cell ＆ Therapeutic Antibody Ministry of EducationChina

ganoderma atrum fungal immunomodulatory protein（FIP-gat） is a new member of the FIP family. The FIP-gat protein was composed 111 amino acids, and the sequences of homologous rate was 86% with FIP-glu（LZ-8）. Based on the cloning FIP-gat gene from G. atrum, we used a p ET30 a expression vector to express FIP-gat gene as a His-tagged fusion protein in Escherichia coli BL21. Then, the recombinant protein was analyzed and identified by the SDS-PAGE and LC/Q-TOF-MS. Further, the rF IP-gat was induced with IPTG and purified using a Ni-NTA agarose resin column. The purified r FIP-gat migrated with the predicted molecular mass on SDS-PAGE and was detected using specific antibodies. Finally, its bioactivity was investigated by tetrazolium cell proliferation assay and agglutination test in human breast cancer cells MDA-MB-231. The results showed that the rF IP-gat protein could be expressed in E. coli BL21, accounted for 59.5% of the total soluble protein, and the yield of recombinant protein was 29.75 mg/L protein. Meanwhile, the r FIP-gat could reduce cell viability dose-dependently;the median inhibitory concentration was 23.921 μg/m L;at 5 μg/mL, the r FIP-gat could agglutinate MDA-MB-231 cells. These results suggest that E. coli BL21 cells can produce active r FIP-gat and that the protein has potential anti-cancer bioactivity in MDA-MB-231 cells.

关键词： ganoderma atrum fungal immunomodulatory protein prokaryotic expression MTT assay agglutinating activity

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