Method for rapidly detecting monosaccharide composition in polysaccharide

Abstract

The invention discloses a method for rapidly detecting monosaccharide composition in polysaccharide, and relates to the technical field of food analysis. The method comprises the following steps: firstly hydrolyzing polysaccharide by adopting TFA, then derivatizing PMP, and finally analyzing the monosaccharide composition of the derivatized polysaccharide hydrolysate by adopting UHPLC QQQ MS (Ultra High Performance Liquid Chromatography QQQ Mass Spectrometry). According to the method provided by the invention, the detection analysis of 18 monosaccharides can be quickly completed within 7 minutes, and the detection limit is also lower than that of a mainstream monosaccharide detection method HPAEC-PAD.

Description

Technical Field

[0001] This invention relates to the field of food analysis technology, and in particular to a method for rapidly detecting the monosaccharide composition in polysaccharides. Background Technology

[0002] Carbohydrates are a class of substances widely found in nature and are an important class of biomolecules besides proteins and nucleic acids. Due to the diversity and complexity of carbohydrate structures, current research on carbohydrates lags far behind that on nucleic acids and proteins.

[0003] Monosaccharides are the smallest building blocks of carbohydrates. Current methods for monosaccharide detection include capillary electrophoresis, thin-layer chromatography (TLC), gas chromatography (GC), liquid chromatography (LC), and liquid chromatography-mass spectrometry (LC-MS). Capillary electrophoresis has poor separation efficiency for monosaccharides; TLC is simple to operate but has low resolution and poor sensitivity, making it suitable only for semi-quantitative or qualitative analysis; GC and LC require derivatization before determination, resulting in long detection times (up to 30 minutes for a single neutral sugar sample); high-performance anion exchange chromatography combined with pulsed amperometric detector (HPAEC-PAD) can detect 12 neutral sugars and uronic acids within 30 minutes without derivatization, making it the primary method for monosaccharide composition determination. However, with an increasing number of monosaccharide types, HPAEC-PAD cannot achieve complete separation. As carbohydrate research deepens and the demand for diverse monosaccharide types grows, higher requirements are placed on the efficient determination of monosaccharide composition. Therefore, developing a method for the simultaneous and rapid detection of mixtures of multiple monosaccharides is of great significance. Summary of the Invention

[0004] In view of this, the main objective of the present invention is to provide a method for rapidly detecting the monosaccharide composition in polysaccharides, so as to solve the problems existing in the prior art.

[0005] To achieve the above objectives, the technical solution of the present invention is as follows:

[0006] A method for rapidly detecting the monosaccharide composition in polysaccharides includes the following steps:

[0007] S1: Hydrolyze the polysaccharide into monosaccharides to obtain a polysaccharide hydrolysate;

[0008] S2: After derivatizing the polysaccharide hydrolysate or the monosaccharide standard mixed solution with PMP, dry it, redissolve it in water, wash it with chloroform, take 200 μL of the aqueous layer and add 200 μL of methanol, and pass it through a 0.22 μm nylon membrane to obtain the test solution;

[0009] S3: The content of various monosaccharide components in the test solution was determined by UHPLC QQQ MS;

[0010] The chromatographic conditions are as follows:

[0011] The chromatographic column was an Agilent Poroshell HPH C18 UHPLC column, model 1.9μm, 2.1×50mm;

[0012] The chromatographic elution conditions were as follows: Solvent A was 5% acetonitrile / water containing ammonium acetate, pH 8.0; Solvent B was 95% acetonitrile / water.

[0013] The elution program was as follows: 0-5 min, 11% B; 5-5.3 min, 11%-99% B; 5.3-6 min, 99% B; 6-6.3 min, 99%-11% B; 6.3-7 min, 11% B.

[0014] Mass spectrometry conditions are:

[0015] The ion source is an electrospray ion source, which operates in positive ion mode;

[0016] Sheath gas temperature 300℃, sheath gas flow rate 12L / min, dryer gas temperature 290℃, atomizer pressure 30psi, dryer flow rate 11L / min, capillary voltage 1800V, nozzle voltage 1500V.

[0017] Further, in step S1, the hydrolysis specifically involves: mixing 1 mg of polysaccharide with 200 μL of LTFA solution, sealing and hydrolyzing at 100°C for 2 hours, filtering, vacuum drying, and redissolving in water to obtain a polysaccharide hydrolysate;

[0018] The TFA concentration is 4M.

[0019] Further, in step S2, the PMP derivation includes:

[0020] Mix 5 μL of polysaccharide hydrolysate or monosaccharide standard solution with 100 μL of 1-phenyl-3-methyl-5-pyrazolone-methanol solution and ammonia, shake well, react at 70 °C in the dark for 45 min, centrifuge, vacuum dry, redissolve in 250 μL of ultrapure water, wash three times with 250 μL of chloroform, take 200 μL of the aqueous layer and add 200 μL of methanol, and pass through a 0.22 μm nylon membrane to obtain the test solution.

[0021] Furthermore, in step S2, the concentration of the polysaccharide hydrolysate is 1 mg / mL;

[0022] The monosaccharide standards include ribose, arabinose, xylose, rhamnose, fucose, galactosamine, glucosamine, fructose, mannose, glucose, galactose, mannuronic acid, glucuronic acid, galacturonic acid, acetylglucose, acetylglucose, muramic acid, and acetylglucan.

[0023] The concentration of the standard solution is 5×10. 6 1×10 6 5×10 5 1×10 5 5×10 4 1×10 4 5×10 3 1×10 3 5×10 2 1×10 2 5×10, 10 ng / mL;

[0024] The concentration of the 1-phenyl-3-methyl-5-pyrazolone-methanol solution is 0.2M;

[0025] The ammonia concentration is greater than 25%.

[0026] Furthermore, in step S3, the concentration of ammonium acetate in solvent A is 25 mM.

[0027] Furthermore, in step S3, the chromatographic conditions of UHPLC also include a column temperature of 35°C, a flow rate of 0.5 mL / min, and an injection volume of 2 μL.

[0028] Further, in step S1, the polysaccharides include, but are not limited to, arabinoxylan, xyloglucan, Dextran, starch, glycogen, galactomannan, RG I pectin and RG pectin;

[0029] The present invention also aims to provide the application of all the above methods in the detection of polysaccharides, wherein in step S1, the hydrolysis method is not unique and can be determined according to the polysaccharide structure type.

[0030] In existing patented technologies, such as Technology 1 (a method for high-throughput screening of lactic acid bacteria producing extracellular polysaccharides and identifying the monosaccharide composition of their extracellular polysaccharides): HPLC-ESI-MS / MS can complete the qualitative and quantitative analysis of 7 monosaccharides within 20 minutes; Technology 2 (a high-performance liquid chromatography method for monosaccharide analysis): HPLC can complete the qualitative and quantitative analysis of 11 monosaccharides within 51 minutes. In comparison, this invention can complete the identification of a mixture of 18 monosaccharides within 7 minutes, and the detection time and the number of monosaccharides are both superior to those reported in existing patents.

[0031] The linear range, detection limit, and quantitation limit of UHPLC QQQ MS were determined. This method was compared with ion chromatography (HPAEC-PAD) to detect the monosaccharide composition of polysaccharides with different structures. The results showed that the present invention can complete the detection of 18 monosaccharides in 7 minutes. The main monosaccharide composition obtained by the invention was basically consistent with the detection results of ion chromatography, which takes 30 minutes to complete the detection. This indicates that UHPLC QQQ MS is a more efficient monosaccharide detection technology.

[0032] The beneficial effects of this invention include at least the following:

[0033] (1) The present invention can completely separate 18 kinds of monosaccharide mixture within 7 minutes, and the detection time and the types of monosaccharides are superior to the existing technology;

[0034] (2) The detection limit of the present invention is lower than that of the mainstream monosaccharide detection method, ion chromatography (HPAEC-PAD), and can detect monosaccharides at lower concentrations. Attached Figure Description

[0035] Figure 1 This is a flowchart for monosaccharide detection in this invention.

[0036] Figure 2 The response strength of 18 monosaccharide derivatives under different fragmentor voltages (voltage range 200V, interval 50V).

[0037] Figure 3 The response strength of 18 monosaccharide derivatives under different fragmentor voltages (voltage range 90V, interval 10V).

[0038] Figure 4 The response strength of 18 monosaccharide derivatives at different fragmentor voltages (voltage range 9V, interval 1V).

[0039] Figure 5 MS / MS plots of 18 monosaccharide derivatives at different Collision Energy voltages.

[0040] Figure 6 MRM diagrams for 18 monosaccharides.

[0041] Figure 7 MRM diagrams of monosaccharides in different polysaccharides were determined using liquid chromatography-mass spectrometry.

[0042] Figure 8 Elution curves for detecting monosaccharides in different polysaccharides using ion chromatography. Detailed Implementation

[0043] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0044] Furthermore, the technical solutions of the various embodiments of the present invention can be combined with each other, but only if they are based on the ability of those skilled in the art to implement them. When the combination of technical solutions is contradictory or cannot be implemented, it should be considered that such combination of technical solutions does not exist and is not within the scope of protection claimed by the present invention.

[0045] The following specific embodiments illustrate the solution proposed in this invention:

[0046] Example 1: Determination of Monosaccharides by UHPLC QQQ MS

[0047] 1. Preparation of monosaccharide standard solution

[0048] Preparation of monosaccharide standard solutions: Weigh 10 mg each of ribose (Rib), arabinose (Ara), xylose (Xyl), rhamnose (Rha), fucose (Fuc), galactosamine (GalN), glucosamine (GlcN), fructose (Fru), mannose (Man), glucose (Glc), galactose (Gal), mannuronic acid (ManA), glucuronic acid (GlcA), galacturonic acid (GalA), N-acetylglucose (GlcNAc), N-acetylglucose (GalNAc), muramic acid (Mur), and N-acetylglucan (NAM) into centrifuge tubes, add 1 mL of ultrapure water, and prepare a 1×10⁻⁶ solution. 7 A monosaccharide mixed standard solution was prepared by serially diluting the mixed standard solution with ultrapure water to obtain a 5 × 10⁶ ng / mL solution. 6 1×10 6 5×10 5 1×10 5 5×10 4 1×10 4 5×10 3 1×10 3 5×10 2 1×10 2 Mixed standard solutions of different concentrations: 5×10 and 10 ng / mL.

[0049] 2. 1-Phenyl-3-methyl-5-pyrazolone (PMP) Derivatization of Monosaccharides

[0050] Take 5 μL of monosaccharide mixed standard solution, add 100 μL of 0.2 M PMP-methanol solution and 100 μL of ammonia (>25%), shake well, react at 70 °C in the dark for 45 min, centrifuge (1000 rpm, 5 min), vacuum dry, redissolve in 250 μL of ultrapure water, extract three times with 250 μL of chloroform, take 200 μL of the aqueous layer and add 200 μL of methanol, and detect through a 0.22 μm nylon membrane.

[0051] 3. Detection by UHPLC QQQ MS method

[0052] Chromatographic conditions: Agilent 1290 Infinity ultra-high performance liquid chromatography system and Agilent 6475QQQ mass spectrometer; Agilent Poroshell HPH C18 UHPLC column (1.9 μm, 2.1 × 50 mm); column temperature 35℃. Elution conditions were as follows: Solvent A was 5% acetonitrile / water (containing 25 mM ammonium acetate), pH 8.0; Solvent B was 95% acetonitrile / water; flow rate 0.5 mL / min; injection volume 2 μL; elution program as follows: 0-5 min, 11% B; 5-5.3 min, 11%-99% B; 5.3-6 min, 99% B; 6-6.3 min, 99%-11% B; 6.3-7 min, 11% B.

[0053] Mass spectrometry conditions: Electrospray ionization (ESI) was used as the ion source, and measurements were performed in positive ion mode using multiple reaction monitoring (MRM) acquisition mode. The mass spectrometry parameters were as follows: sheath gas temperature 300℃, sheath gas flow rate 12 L / min, drying gas temperature 290℃, nebulizer pressure 30 psi, dryer flow rate 11 L / min, capillary voltage 1800 V, and nozzle voltage 1500 V. The mass scan range for monosaccharides was set to m / z 50–1700. The fragmentator voltage and collision energy voltage for each monosaccharide were optimized, and the optimized voltages were used for testing.

[0054] 4. Examination of linear relationships

[0055] Standard solutions of monosaccharides at different concentrations were determined according to the chromatographic and mass spectrometric conditions described in step 3 of the method. Linear regression was performed using the peak area and mass concentration logarithms of the MRM chromatogram as the Y-axis and X-axis, respectively, to plot a standard curve for each monosaccharide, thus obtaining the linear equation. The correlation coefficient R was ensured to be within the linear range. 2 >0.99 and meets the requirement that the minimum concentration is not less than the limit of quantitation.

[0056] 5. Investigation of detection limit and quantitation limit

[0057] Limit of detection (LOD): The lowest concentration of the analyte that the analytical method can detect under specified experimental conditions; Limit of quantification (LOQ): The lowest concentration of the analyte that the analytical method can quantitatively determine. A mixed standard solution of 18 monosaccharides was stepwise diluted, and 2 μL was injected into a UHPLC QQQ MS. The LOD was calculated based on a signal-to-noise ratio (S / N) of 3, and the LQ was calculated based on a S / N of 10.

[0058] 6. UHPLC QQQ MS method applied to the detection of polysaccharides.

[0059] Hydrolysis of polysaccharides: Weigh 1.00 mg of each of the eight polysaccharides (arabinoxylan, xyloglucan, Dextran, starch, glycogen, galactomannan, RG I pectin and RG pectin), add 200 μL of 4MTFA, and hydrolyze in a sealed container at 100 °C for 2 h. Dry under vacuum and reconstitute with water to obtain a 1.00 mg / mL polysaccharide hydrolysate.

[0060] Derivatization and UHPLC QQQ MS determination of polysaccharide hydrolysate: Take 5 μL of polysaccharide hydrolysate, derivatize with PMP, extract the aqueous layer and pass it through a membrane, and detect it by UHPLC QQQ MS method, referring to steps 2 and 3.

[0061] (1) Selection of Fragmentor voltage and Collision Energy voltage for 18 monosaccharide derivatives

[0062] Fragmentor voltage determines the mass spectrometry response intensity of a substance. To obtain the highest mass spectrometry response intensity for different monosaccharide derivatives, the fragmentor voltage for 18 monosaccharide derivatives was optimized. Measurements were performed at 50V intervals within a voltage range of 100V-300V. The mass spectrometry intensities at different fragmentor voltages for different monosaccharide derivatives are shown below. Figure 2 Rib, Ara, Xyl, Rha, Fuc, GalN, GlcN, Fru, Man, Glc, Gal, ManA, GlcA, GalA, GlcNAc, and GalNAc exhibited the highest signal strength at 150V, while Mur and NAM showed the highest signal strength at 200V. The voltages on either side of the highest signal strength were selected as the further Fragmentor voltage optimization range. Therefore, the further Fragmentor voltage optimization range for Rib, Ara, Xyl, Rha, Fuc, GalN, GlcN, Fru, Man, Glc, Gal, ManA, GlcA, GalA, GlcNAc, and GalNAc is 100V-190V, and for Mur and NAM, it is 150V-240V. Measurements were performed at 10V intervals, and the results are shown in [Figure 1]. Figure 3Rib, Ara, Rha, Glc, Gal, GlcNAc, and NAM have the highest signal strength at 160V; Xyl, Fuc, GlcN, GalN, Man, ManA, GlcA, GalA, and GalNAc have the highest signal strength at 170V; Fru has the highest signal strength at 150V; and Mur has the highest signal strength at 190V. To further refine the fragmentation voltage of each monosaccharide derivative, the further optimized fragmentation voltage range for Rib, Ara, Rha, Glc, Gal, GlcNAc, and NAM was 155V-164V; for Xyl, Fuc, GlcN, GalN, Man, ManA, GlcA, GalA, and GalNAc, it was 165V-174V; for Fru, it was 145V-154V; and for Mur, it was 185V-194V. Measurements were performed at 1V intervals, and the signal intensity at different voltages is shown in [Figure showing signal strength at different voltages]. Figure 4 The voltage with the highest response intensity was taken as the fragmentor voltage for each monosaccharide, and the selected fragmentor voltages for each monosaccharide are shown in Table 1.

[0063] The collision energy (V) determines the degree of fragmentation of a substance in mass spectrometry. A suitable fragmentation voltage is needed to obtain abundant fragment information while ensuring the presence of the parent ion. Three fragmentation voltages (20V, 25V, and 30V) were used to test 18 monosaccharide derivatives; the mass spectra are shown below. Figure 5 The Collision Energy voltages for each monosaccharide selection under the condition of ensuring the presence of the parent ion are shown in Table 1.

[0064] Table 1. MRM ion pairs for each monosaccharide

[0065]

[0066]

[0067] (2) Isolation of 18 monosaccharide derivatives

[0068] Figure 6As can be seen, the total time for monosaccharide elution and column equilibration was 7 min, but the elution of 18 monosaccharides was completed within 4 min. First, based on the mass-to-charge ratio (m / z), pentoses, hexoses, deoxyhexoses, glycosamines, uronic acids, N-acetyl sugars, muramic acids, and N-acetylmuramic acids can be distinguished. Second, considering the elution time, the peak elution times for pentoses were Rib (1.361 min), Xyl (2.814 min), and Ara (2.989 min); the peak elution times for hexoses were Fru (0.356 min), Man (1.137 min), Glc (2.424 min), and Gal (2.773 min); the peak elution times for deoxyhexoses were Rha (1.494 min) and Fuc (3.354 min); and the peak elution times for glycosamines were... The elution times of GlcN (1.394 min) and GalN (2.565 min) were as follows: uronic acid (0.863 min), GlcA (1.411 min), and GalA (1.660 min); acetylglucosamine (2.457 min) and GalNAc (2.706 min) were as follows: GlcNAc (2.457 min) and GalNAc (2.706 min); muramic acid (0.763 min) and acetylglucosamine (1.486 min) were as follows: muramic acid (0.763 min) and acetylglucosamine (1.486 min).

[0069] (3) Linear range, detection limit and quantitation limit of monosaccharides

[0070] Table 2 summarizes the linear range, limit of detection, and limit of quantitation for 18 monosaccharide standards. Among them, the linear ranges for Xyl, Ara, Rha, Fuc, GlcN, GalN, Man, Gal, GlcA, GalA, GlcNAc, and GalNAc are all within 50–1 × 10⁻⁶. 6 The linear range for ManA and Mur is 100–1 × 10 ng / mL. 6 The linear range for Fru and Glc was 500–1 × 10⁻⁶ ng / mL. 6 The linear range for Rib and NAM was 1000–1 × 10⁻⁶ ng / mL. 6 ng / mL. Linear equations R for each monosaccharide. 2 The limit of detection (LOD) is >0.99, the limit of quantitation (LOQ) is 0.7-212.77 ng / mL, and the limit of quantitation (LOQ) is 2.32-709.22 ng / mL, indicating that this method has good linearity within this linear range.

[0071] Table 218 linear ranges, limits of detection, and limits of quantitation for liquid chromatography-mass spectrometry (LC-MS) analysis of monosaccharides.

[0072]

[0073]

[0074] (4) Polysaccharides

[0075] Figure 7 The MRM diagrams of monosaccharide composition in polysaccharides with different structures were determined using UHPLC-QQQ MS. The results of each monosaccharide composition are shown in Table 3. The recoveries of Ara and Xyl in arabinoxylan monosaccharide composition reached 98.21%, Xyl and Glc in xyloglucan monosaccharide composition reached 90.1%, Glc in dextran monosaccharide composition reached 96.96%, Glc in glycogen monosaccharide composition reached 97.34%, Glc in starch monosaccharide composition reached 96.76%, and Man and Gal in galactomannan monosaccharide composition reached 98.59%. The main monosaccharide components of RG I pectin were Ara, Rha, Gal, and GalA, with a total recovery rate of 95.04%. The main monosaccharide components of RG type pectin were Xyl, Ara, Rha, Fuc, Gal, and GalA, with a total recovery rate of 98.3%. The results of UHPLC QQQ MS determination of the main monosaccharide composition of each polysaccharide verified the structural type of the corresponding polysaccharide, and the relative standard deviation of the content of all monosaccharides was between 0.00% and 0.59%.

[0076] Table 3. Monosaccharide composition (%) of polysaccharides with different structures determined by UHPLC QQQ MS method

[0077]

[0078]

[0079] Comparative Example 1: HPAEC-PAD Method for Determination

[0080] The polysaccharide hydrolysate was obtained in the same manner as in Example 1. The hydrolysate was diluted to 20 μg / mL and filtered through a Dionex membrane. TM Detected using ICS-6000. Chromatographic conditions: CarboPac TM PA20 analytical column (3 mm × 150 mm, Dionex, CA). Mobile phase: A is 0.25 mol / L sodium hydroxide, B is ultrapure water, C is 1 mol / L sodium acetate, D is ultrapure water; elution conditions are shown in Table 4. Column temperature: 30℃, component temperature: 25℃, flow rate: 0.5 mL / min, injection volume: 10 μL.

[0081] Table 4 ICS 6000 Elution Procedure

[0082] Time (min) Flow rate (mL / min) A(％) B(％) C(％) 0.0 0.5 0.8 99.2 0.0 21.0 0.5 0.8 99.2 0.0 21.1 0.5 5.8 94.2 5.0 30.0 0.5 20.8 79.2 20.0 30.1 0.5 80.0 20.0 0.0 50.0 0.5 80.0 20.0 0.0 50.1 0.5 0.8 99.2 0.0 60.0 0.5 0.8 99.2 0.0

[0083] Figure 8The elution curves for determining the monosaccharide composition of polysaccharides with different structures were obtained using the HPAEC-PAD method, and the results are shown in Table 5. The HPAEC-PAD method is a commonly used method for monosaccharide composition determination. The difference between the monosaccharide recoveries obtained by this method and those obtained by the UHPLC QQQ MS method was within 5%, indicating that the UHPLC QQQ MS method is relatively reliable and practical. Furthermore, the HPAEC-PAD method can detect concentrations as low as 250 ng / mL, while the UHPLC QQQ MS method can detect concentrations as low as 50 ng / mL. Overall, the UHPLC QQQ MS method has a wide detection range, accurate results, short measurement time, and good separation even when multiple monosaccharides are present simultaneously, making it suitable for determining the monosaccharide composition of food polysaccharides.

[0084] Table 5. Monosaccharide composition (%) in polysaccharides with different structures determined by HPAEC-PAD method.

[0085]

[0086]

[0087] It should be noted that, in this document, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Unless otherwise specified, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes that element.

[0088] The sequence numbers of the above embodiments of the present invention are for descriptive purposes only and do not represent the superiority or inferiority of the embodiments.

[0089] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. A method for rapidly detecting the monosaccharide composition in polysaccharides, characterized in that, Includes the following steps: S1: Hydrolyze the polysaccharide into monosaccharides to obtain a polysaccharide hydrolysate; S2: After derivatizing the polysaccharide hydrolysate or the monosaccharide standard mixed solution with PMP, the solution is dried, reconstituted with water, washed with chloroform, and the aqueous layer is collected and an equal volume of methanol is added. The solution is then passed through a 0.22 μm nylon membrane to obtain the test solution. S3: The content of various monosaccharide components in the test solution was determined by UHPLC QQQ MS; The chromatographic conditions are as follows: The chromatographic column was an Agilent Poroshell HPH C18 UHPLC column, model 1.9μm, 2.1×50mm; The chromatographic elution conditions were as follows: Solvent A was 5% acetonitrile / water containing ammonium acetate, pH 8.0; Solvent B was 95% acetonitrile / water. The elution program was as follows: 0-5 min, 11% B; 5-5.3 min, 11%-99% B; 5.3-6 min, 99% B; 6-6.3 min, 99%-11% B. 6.3-7 min, 11% B; Mass spectrometry conditions are: The ion source is an electrospray ion source, which operates in positive ion mode; Sheath gas temperature 300℃, sheath gas flow rate 12L / min, dryer gas temperature 290℃, atomizer pressure 30psi, dryer flow rate 11L / min, capillary voltage 1800V, nozzle voltage 1500V.

2. The method for rapid detection of monosaccharide composition in polysaccharides according to claim 1, characterized in that, In step S1, the hydrolysis specifically involves mixing 1 mg of polysaccharide with 200 μL of TFA solution, sealing and hydrolyzing at 100°C for 2 hours, filtering, vacuum drying, and reconstitution with water to obtain a polysaccharide hydrolysate. The TFA concentration is 4M.

3. The method for rapid detection of monosaccharide composition in polysaccharides according to claim 1, characterized in that, In step S2, the PMP derivation specifically includes: Mix 5 μL of polysaccharide hydrolysate or monosaccharide standard solution with 100 μL of 1-phenyl-3-methyl-5-pyrazolone-methanol solution and 100 μL of ammonia water, shake well, react at 70 °C in the dark for 45 min, centrifuge, vacuum dry, redissolve in 250 μL of ultrapure water, wash three times with 250 μL of chloroform, take 200 μL of the aqueous layer and add 200 μL of methanol, filter through a 0.22 μm nylon membrane to obtain the test solution. The concentration of the polysaccharide hydrolysate is 1 mg / mL; The monosaccharide standards include ribose, arabinose, xylose, rhamnose, fucose, galactosamine, glucosamine, fructose, mannose, glucose, galactose, mannuronic acid, glucuronic acid, galacturonic acid, N-acetylglucose, N-acetylglucose, muramic acid, and N-acetylglucan. The standard solution concentration is 5 × 10⁻⁶. 6 1×10 6 5×10 5 1×10 5 5×10 4 1×10 4 5×10 3 1×10 3 5×10 2 1×10 2 5×10, 10 ng / mL; The concentration of the 1-phenyl-3-methyl-5-pyrazolone-methanol solution is 0.2M; The ammonia concentration is greater than 25%.

4. The method for rapid detection of monosaccharide composition in polysaccharides according to claim 1, characterized in that, In step S3, the concentration of ammonium acetate in solvent A is 25 mM.

5. The method for rapid detection of monosaccharide composition in polysaccharides according to claim 1, characterized in that, In step S3, the chromatographic conditions for UHPLC also include a column temperature of 35°C, a flow rate of 0.5 mL / min, and an injection volume of 2 μL.

6. The method for rapid detection of monosaccharide composition in polysaccharides according to claim 1, characterized in that, The polysaccharides include arabinoxylan, xyloglucan, Dextran, starch, glycogen, galactomannan, RG I type pectin, and RG type pectin.

7. The application of any one of the methods of claims 1-6 in the detection of polysaccharides.

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