The National Edible Mushroom Center of Henan University has made progress in the structural identification and immunomodulatory effect of Xiuzhen mushroom polysaccharide

Responsible Editor: Food Science

Funded by the major public welfare projects of Henan Province: Key Technology and Industrial Demonstration of Deep Processing of Edible Mushrooms and the Creation of Health Foods (201300110200) and Research on Precision Nutrition and Health Foods (CXJD2021006), in cooperation with the team of researcher Zhang Yan of Hebei Food Inspection Research Institute, On the structure identification of the polysaccharide from Pleurotus chinensis, the result was published in the journal Carbohydrate Polymers under the title of "A glucomannogalactan from Pleurotus geesteranus : Structural characterization, chain conformation and immunological effect" (instant IF: 10.

Pleurotus geesteranus is one of the most rapidly developing varieties in China's edible mushroom industry.

Results and Discussion

Chemical composition and homogeneity analysis

Table 1 shows the yield, neutral sugar content, uronic acid content, and monosaccharide composition of PGP and its purified fractions

The molecular weight of PGP-1c was calculated from a dextran standard curve (Log M w=−1.

In the UV spectrum, PGP-1c has no absorption peaks at 260 and 280 nm, which further proves that PGP-1c does not contain free proteins and nucleic acids

The results of methylation analysis of PGP-1c are shown in Table 2

In ¹ H NMR (Fig.

In order to obtain the chemical shift information of all H and C of each major fragment of PGP-1c, so as to determine the linking sequence of each residue fragment, 2D NMR was further measured, through the spectral correlation signal and the coupling between ¹ H and ¹³ C nuclei , assigning chemical shifts to individual residues

After the residue fragments with corresponding chemical shifts were determined, the connectivity between different residues in PGP-1c and the structural information of the molecular backbone were further analyzed and determined by HMBC

Due to the magnetic field generated by the resonance transition, the signal of fragment a is very weak, and it is difficult to generate junction sites with other fragments in HMBC.

Molecular chain conformation of PGP-1c

In Figure 4, the 3D structure of PGP-1c is in the shape of mountains (or islands) of different sizes (10-100 nm in height, 24.

In Figure 5A, the XRD spectrum of PGP-1c showed 5 distinct sharp and narrow peaks at 14.

In Fig.

To explore the effect of PGP-1c on the viability of macrophages RAW 264.
7, MTT method was used to evaluate the effect of PGP-1c in the dose range of 12.
5-800 μg/mL
.
The results (Fig.
6A) showed that PGP-1c had no toxic effect on RAW 264.
7 cells in the range of 12.
5-400 μg/mL, and the concentrations of 100, 200 and 400 μg/mL were selected for the next experiment
.

Immunomodulatory activity of PGP-1c

In Figure 6B, compared with the blank group, PGP-1c could significantly stimulate RAW 264.
7 cells to secrete NO in the concentration range of 100-400 μg/mL, and showed an obvious dose-effect relationship
.
After 100 μg/mL of PGP-1c acted on the cells for 24 h, the NO secretion reached 12.
03 ± 0.
13 μM, an increase of 3.
08 μM compared with the blank group, and was 54.
45% of the LPS positive control group.
The increase in NO content indicated that PGP-1c may It has good immune-promoting effect and the activity of inhibiting the growth of tumor cells
.
In Figure 6C, compared with the blank control group, different concentrations (200-400 μg/mL) of PGP-1c could significantly promote the secretion of IL-6 from RAW 264.
7 cells ( P < 0.
01) in a dose-dependent manner
.
In Fig.
6D, PGP-1c (100-400 μg/mL) significantly promoted the release of TNF -α in RAW 264.
7 cells , with the PGP-1c activity at 100 and 200 μg/mL being extremely significant ( P < 0.
001)
.
The results of PGP-1c promoting the release of cytokines from RAW 264.
7 cells indicated that PGP-1c has a good immune-enhancing ability
.

Conclusion

A purified polysaccharide PGP - 1c with a molecular weight of 20.
9 kDa was obtained from Xiuzhen Mushroom .
(1,2,6- α -Gal and 1,2,6- α - 3 - O Me-Gal) galactose units are linked by α- (1→6) backbone and β -D- Man p- (1→,→3) -α -D-Glc p- (1→ and a small amount of Fuc p as side chains consist of 48% branched glucomannan .
In addition, chain conformational characterization revealed that PGP- 1c has a long branched and stacked multi-chain network structure, as well as a triple-helix structure with anti-tumor immune activity characteristics .
By examining the effect of PGP-1c on the NO and cytokine secretion capacity of macrophages, it was found that it has 3- O - CH The ₃ -group, triple-helix and multi-branched polysaccharides have good immunomodulatory activity .

About the corresponding author

Kang Wenyi , Ph.
D.
, second-level professor, Director of National Edible Fungus Processing Technology Research and Development Center of Henan University, Director of Henan Functional Food Engineering Technology Research Center, Director of Henan International Joint Laboratory for Functional Research of Medicinal and Edible Resources, Henan Edible Mushroom Chairman of the Technology Innovation Strategic Alliance of the Deep Processing Industry, Member of the National Professional Standardization Technical Committee, Vice Chairman of the Edible Fungus Postharvest and Processing Industry Branch of the Chinese Society of Mycology, Standing Committee Member of the Nutrition and Health Food Branch of the Chinese Nutrition Society, and post-marketing of the World Federation of Chinese Medicines Executive director of the re-evaluation professional committee, expert member of the State Food and Drug Administration, leading analysis of special food sampling monitoring, deputy director of the post-doctoral innovation team of traditional Chinese medicine in Henan Province, and industrial technology reform expert in Henan Province
.
At present, the main research directions are the material basis of medicinal and edible resources, the efficacy evaluation of key biologically active molecules and their mechanism of action, as well as the detection methods and quality management of raw materials and their derived products
.
Currently presided over 1 sub-project of the "13th Five-Year" national key R&D plan, 2 health food special projects of the Food and Drug Administration, 1 major public welfare project of Henan Province and more than 10 provincial and ministerial projects; published more than 400 research papers, including SCI More than 100 high-level papers
.
Based on this work foundation, he won the second prize of Henan Province Science and Technology Progress Award and the special prize of China Chamber of Commerce in 2020
.

About the corresponding author

Zhang Yan , male, researcher, postdoctoral fellow in medicine, vice president of Hebei Food Inspection Research Institute, expert enjoying special allowances from the State Council, first-level candidate of Hebei Province's "Three-Three-Three" Talent Project, Hebei Province Top Young Talent, International Organization for Standardization Biology Expert of the Bioanalytical Methods Working Group of the Technical Committee, member of the 2nd National Food Safety Standard Review Committee, Deputy Editor-in-Chief of Food Science and Human Wellness (incorporated by SCI), Deputy Editor-in-Chief of Journal of Food Quality (incorporated by SCI), National Market Member of the expert group of the Beijing Winter Olympics Food Supply Safety Coordination Group of the State Administration of Supervision, and member of the Beijing 2022 Winter Olympics Food Safety Guarantee Expert Committee
.
In recent years, the comrade has undertaken more than 30 national-level scientific research projects and provincial and ministerial-level scientific research projects such as the "13th Five-Year Plan" and "Twelfth Five-Year Plan", published more than 200 high-level academic papers, participated in the compilation and publication of 24 monographs, authorized the national There are 9 invention patents and 8 computer software copyright registrations; 54 national standards, 11 industry standards, and 65 local standards have been formulated and revised, and 25 scientific research achievements have won provincial and ministerial-level scientific and technological progress awards
.

About the first author

Liang Zhenhua , a 2019 graduate student of Henan University, mainly focuses on the structure and biological activity of polysaccharides
.
He has won the National Scholarship and the title of "Three Good Students in Henan Province", and has published 6 SCI papers in journals such as Carbohydrate Polymers, Food Science and Human Wellness as the first author and co-first author
.