Purification, Structural Characterization and Immunomodulatory Effects of Polysaccharides from Amomumvillosum Lour. on RAW 264.7 Macrophages
Abstract
:1. Introduction
2. Results and Discussion
2.1. Preparation and Physicochemical Features of Polysaccharides
2.2. Structural Characterization of Polysaccharides
2.2.1. Monosaccharide Composition Identification
2.2.2. Determination of Homogeneity and Molecular Weight
2.2.3. Fourier Transform Infrared (FT-IR) Spectroscopy Analysis
2.2.4. Determination of Triple Helical Structure
2.2.5. Surface Microscopic Analysis
2.2.6. Methylation Analysis
2.2.7. Nuclear Magnetic Resonance (NMR) Analysis
2.3. Immunomodulatory Activities
2.3.1. Effects of Polysaccharides on Cell Viability and NO Production of Macrophages
2.3.2. Effect of AVPG-2 on Cytokines Secretion of Macrophages
2.3.3. Effects of AVPG-2 on Polarization of Macrophages
2.3.4. Effects of AVPG-2 on Phagocytic Capacity of Macrophages
3. Materials and Methods
3.1. Materials and Reagents
3.2. Extraction and Purification of Polysaccharides
3.3. Preliminary Chemical Analysis
3.4. Determination of Homogeneity and Molecular Weight
3.5. Monosaccharide Composition Identification
3.6. UV Spectroscopy and FT-IR Spectroscopy Analysis
3.7. Determination of Triple-Helical Structure
3.8. Surface Microscopic Analysis
3.9. Methylation Analysis
3.10. NMR Spectroscopy Analysis
3.11. Immunomodulatory Activity
3.11.1. Cell Culture
3.11.2. Assay of Cell Viability
3.11.3. Assay of NO
3.11.4. Determination of Cytokines
3.11.5. Real Time (RT)-PCR
3.11.6. Assay of Macrophages Phagocytosis
3.12. Statistical Analysis
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
Sample Availability
References
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Methylated Sugars | Linkage | AVPG-1 (Molar Ratios) | AVPG-2 (Molar Ratios) |
---|---|---|---|
2,3,6-Me3-Glcp | →4)-Glcp-(1→ | 0.65 | 0.36 |
2,6-Me2-Glcp | →3,4)-Glcp-(1→ | 0.16 | ND |
2,3,6-Me3-Galp | →4)-Galp-(1→ | 0.09 | 0.04 |
2,3,4-Me3-Xylp | Xylp-(1→ | 0.04 | ND |
2,3,5-Me3-Araf | Araf-(1→ | 0.06 | 0.09 |
1,2,3,4-Me4-Glcp | →6)-Glcp | ND | 0.09 |
2,4,6-Me3-Galp | →3)-Galp-(1→ | ND | 0.03 |
2,4-Me2-Galp | →3,6)-Galp-(1→ | ND | 0.17 |
2,3-Me2-Araf | →5)-Araf-(1→ | ND | 0.06 |
2,3-Me2-Xylp | →4)-Xylp-(1→ | ND | 0.10 |
2,3,4-Me3-Rhalp | Rhap-(1→ | ND | 0.02 |
Peak | Glycosyl Residue | H-1/C-1 | H-2/C-2 | H-3/C-3 | H-4/C-4 | H-5/C-5 | H-6/C-6 |
---|---|---|---|---|---|---|---|
Chemical shifts of residues of AVPG-1 | |||||||
A | -4)-α-d-Glcp-(1- | 5.38/99.70 | 3.62/71.01 | 3.95/72.92 | 3.64/73.25 | 3.76/72.26 | 3.41, 4.03 /68.98 |
B | -3,4)-β-d-Glcp-(1- | 4.45/101.61 | 3.35/72.80 | 3.80/80.18 | 3.74/73.55 | 3.56/73.40 | 3.70, 3.94 /60.44 |
C | β-d-Xylp-(1- | 4.71/103.10 | 3.76/75.83 | 4.03/71.18 | 3.41/71.67 | 3.70, 4.12/67.49 | |
D | -4)-α-d-Galp-(1- | 5.22/108.35 | 3.56/78.60 | 3.26/72.21 | 3.69/79.54 | 4.11/76.60 | 3.80, 4.00 /67.54 |
E | α-l-Araf-(1- | 5.20/109.20 | 4.19/80.85 | 3.92/75.86 | 4.11/83.77 | 3.36, 4.20/62.46 | |
F | α-d-GlcpA-(1- | 5.26/97.60 | 4.14/80.37 | 3.69/79.68 | 3.82/70.89 | 4.04/61.64 | -/179.22 |
Chemical shifts of residues of AVPG-2 | |||||||
A | -6)-β-d-Glcp | 4.62/95.84 | 3.22/71.19 | 3.45/75.81 | 3.87/69.30 | 4.09/68.42 | 3.70, 3.94/61.23 |
B | -4)-α-d-Glcp-(1- | 5.40/99.70 | 3.62/71.55 | 3.94/73.19 | 3.61/76.66 | 3.75/76.47 | 3.41, 3.98/68.70 |
C | -3)-α-d-Galp-(1- | 5.39/101.63 | 3.62/71.43 | 3.70/79.94 | 3.40/69.51 | 3.93/73.11 | 4.12, 4.19/68.14 |
D | -4)-α-d-Galp-(1- | 5.22/108.27 | 3.50/71.36 | 3.26/72.73 | 3.67/72.58 | 3.40/72.25 | 3.71, 3.82/60.94 |
E | -3,6)-β-d-Galp-(1- | 4.41/103.09 | 3.50/71.63 | 3.70/80.03 | 3.53/73.81 | 3.36/69.57 | 3.71, 4.09/68.50 |
F | α-l-Araf-(1- | 5.23/109.18 | 4.19/81.19 | 3.80/76.31 | 4.11/76.43 | 3.45, 3.80/60.36 | |
G | α-l-Araf-(1- | 5.20/92.01 | 4.20/81.25 | 3.91/76.63 | 4.11/76.52 | 3.45, 3.80/60.36 | |
H | -5)-α-l-Araf-(1- | 5.07/98.92 | 4.11/83.85 | 3.92/76.49 | 4.19/81.18 | 3.68, 3.91/76.67 | |
I | -4)-β-d-Xylp-(1- | 4.49/102.72 | 3.34/73.17 | 3.55/73.97 | 3.94/76.49 | 3.39, 4.10/62.89 | |
J | α-l-Rhap-(1- | 4.71/100.70 | 3.90/73.37 | 4.11/80.40 | 3.41/72.03 | 4.01/69.88 | 1.23/16.42 |
K | α-d-GlcpA-(1- | 5.26/97.61 | 4.14/80.60 | 3.92/76.47 | 4.19/80.94 | 3.98/68.66 | 177.05 |
L | -4)-α-d-GalpA | 5.06/107.43 | 3.74/79.84 | 3.63/71.49 | 4.10/83.89 | 3.92/76.46 | 175.38 |
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Zhou, Y.; Qian, C.; Yang, D.; Tang, C.; Xu, X.; Liu, E.-H.; Zhong, J.; Zhu, L.; Zhao, Z. Purification, Structural Characterization and Immunomodulatory Effects of Polysaccharides from Amomumvillosum Lour. on RAW 264.7 Macrophages. Molecules 2021, 26, 2672. https://doi.org/10.3390/molecules26092672
Zhou Y, Qian C, Yang D, Tang C, Xu X, Liu E-H, Zhong J, Zhu L, Zhao Z. Purification, Structural Characterization and Immunomodulatory Effects of Polysaccharides from Amomumvillosum Lour. on RAW 264.7 Macrophages. Molecules. 2021; 26(9):2672. https://doi.org/10.3390/molecules26092672
Chicago/Turabian StyleZhou, Yang, Chunguo Qian, Depo Yang, Cailin Tang, Xinjun Xu, E-Hu Liu, Jingtang Zhong, Longping Zhu, and Zhimin Zhao. 2021. "Purification, Structural Characterization and Immunomodulatory Effects of Polysaccharides from Amomumvillosum Lour. on RAW 264.7 Macrophages" Molecules 26, no. 9: 2672. https://doi.org/10.3390/molecules26092672