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. 2017 Nov 17;22(11):1992.
doi: 10.3390/molecules22111992.

A New UPLC-MS/MS Method for the Characterization and Discrimination of Polysaccharides from Genus Ephedra Based on Enzymatic Digestions

Yong-Gang Xia  1 , Tian-Long Wang  2 , Li-Ming Sun  3 , Jun Liang  4 , Bing-You Yang  5 , Hai-Xue Kuang  6
Affiliations

A New UPLC-MS/MS Method for the Characterization and Discrimination of Polysaccharides from Genus Ephedra Based on Enzymatic Digestions

Yong-Gang Xia et al. Molecules. .

Abstract

Ephedra sinica polysaccharides have been reported to possess important activities, so quality evaluation of polysaccharides from the genus Ephedra is urgent. In this study, enzymatic digestions were performed to establish multiple saccharide fingerprints by ultra-performance liquid chromatography with electrospray ionization triple quadrupole linear ion trap mass spectrometry (UPLC-ESI-TQ-MS/MS) based on a multiple-reaction monitoring in negative mode. Under optimum UPLC-ESI⁻-TQ-MS/MS conditions, excellent separation and quantification of 21 constituents were achieved within 20 min on a solid core column with a 1.6 μm particle using pre-column derivatization with a PMP reagent. This method, coupled with enzymatic digestions and principal component analysis, has been successfully applied to characterize and discriminate Ephedra polysaccharides attributed to different species and plant parts. The results suggest that the proposed analytical strategy could achieve a quality evaluation of plant polysaccharides from traditional Chinese medicines.

Keywords: UPLC-MS/MS; enzymatic digestion; genus Ephedra; polysaccharides.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
(A) Condensation reaction with PMP; (B) characteristic fragmentation pathways of bis-PMP derivatives of monosaccharides in negative ESI-MS/MS mode. Peaks were as follows: 1, Man; 2, GlcN; 3, ManN; 5, Rib; 6, Rha; 8, GalN; 11, GlcUA; 12, GalUA; 14, GlcNAc; 16, D2-Glc; 17, Glc; 18, GalNAc; 19, Gal; 20, Ara; 21, Fuc.
Figure 2
Figure 2
MRM ion chromatograms and 22 reference standards. Peaks 1–3, 5, 6, 8, 11, 12, 15 and 16–21 are the same as in Figure 1. Other peaks are as follows: 4, maltoheptaose; 7, maltohexaose; 9, maltopentaose; 10, maltotetraose; 13, maltotriose; 15, lactose; 22, 5-HMF.
Figure 3
Figure 3
Typical MRM ion chromatograms of E. sinica root polysaccharides treated by multiple enzymatic digestions. (A), α-amylase; (B), β-(1→3)-d-glucanase, (C), pectinase; (D), cellulose; (E), endo-1,4-β-xylanase. (a,b) were due to polysaccharide samples with and without enzymatic digestions, respectively.
Figure 3
Figure 3
Typical MRM ion chromatograms of E. sinica root polysaccharides treated by multiple enzymatic digestions. (A), α-amylase; (B), β-(1→3)-d-glucanase, (C), pectinase; (D), cellulose; (E), endo-1,4-β-xylanase. (a,b) were due to polysaccharide samples with and without enzymatic digestions, respectively.
Figure 4
Figure 4
PCA score plots of 20 polysaccharide samples from multiple enzymatic digestions. (A) α-amylase; (B) β-(1→3)-d-glucanase, (C) pectinase; (D) cellulose; (E) endo-1,4-β-xylanase. (a) E. sinica (stem); (b) E. sinica (root); (c) E. intermedia (stem); (d) E. equisetina (stem).
See this image and copyright information in PMC

References

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