An HG-type pectin, its preparation method and application

Abstract

The present invention discloses an HG-type pectin, its preparation method and application. The present invention extracts a crude extract from plant raw materials and conducts multi-stage separation and purification, and screens out an HG-type pectin with a single component having the activity of improving dyslipidemia. In addition, the present invention constructs an animal model to verify that the HG-type pectin has the effect of improving dyslipidemia. The HG-type pectin of the present invention has broad application prospects in the fields of foods, drugs, etc. for preventing, improving or treating dyslipidemia.

Description

Technical Field

[0001] The present invention belongs to the technical field of natural product extraction, and particularly relates to an HG-type pectin, a preparation method thereof, and an application thereof. Background Art

[0002] As an active ingredient of certain functional edible and medicinal plants, pectin has various pharmacological activities such as liver protection, anti-inflammatory, immunomodulatory, antioxidant, and anti-tumor. The relationship between its biological activity and structural characteristics is manifested in aspects such as molecular weight, monosaccharide composition, glycosidic bond and its linkage mode, functional groups, and spatial configuration. The structural characteristics of pectin not only affect its physical and chemical properties such as solubility, stability, and bioavailability, but also affect its biological activity. At the same time, the composition and fine structure of pectin vary greatly due to differences in extraction conditions.

[0003] However, there are still some problems in current pectin extraction and application technologies. For example, after pectin is extracted, it still needs to be purified to remove impurities such as pigments and inorganic salts. Appropriate purification agents and methods need to be selected during the purification process, otherwise it will lead to the loss of pectin and the change of its structure. Therefore, there is still a need to study new pectin extraction and purification processes to obtain pectin components with specific structures and high biological activities, thereby promoting the sustainable development of the pectin industry. Summary of the Invention

[0004] To solve at least some of the above-mentioned technical problems in the prior art, the present invention screens out an HG-type pectin with a single component having the activity of improving dyslipidemia by extracting a crude extract from plant raw materials and performing multi-stage separation and purification. Specifically, the present invention includes the following contents.

[0005] In the first aspect of the present invention, there is provided an isolated HG-type pectin, the main chain of which has a glycosidic bond as shown in →4)-α-D-GalAp-(1→, and the terminal α-D-GalAp-(1→ is connected to the main chain through →3,4)-α-D-GalAp-(1→, and the HG-type pectin has the structure shown in Formula I:

[0006] Formula I;

[0007] Wherein, n is a natural number selected from 1 to 30.

[0008] In certain embodiments, the HG-type pectin according to the present invention has a molecular weight of 12931 Da.

[0009] In the second aspect of the present invention, there is provided a pharmaceutical composition, which includes the HG-type pectin according to the present invention.

[0010] In certain embodiments, the pharmaceutical composition according to the present invention, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

[0011] In certain embodiments, the pharmaceutical composition according to the present invention, wherein the pharmaceutically acceptable carrier comprises at least one of a diluent, a filler, an absorbent, a wetting agent, a binder, a disintegrant, a lubricant, a sweetening agent, a preservative, and an antioxidant.

[0012] A third aspect of the present invention provides a method for preparing HG-type pectin according to the present invention, which comprises the following steps:

[0013] (1) Extracting a plant raw material with an aqueous solvent at 50 - 90 °C for 100 - 200 min to obtain an extract, adding a precipitant for precipitation to obtain a crude extract;

[0014] (2) Subjecting the crude extract to multi-stage separation and purification to obtain the HG-type pectin.

[0015] In certain embodiments, the preparation method according to the present invention, wherein the plant raw material comprises at least one of seabuckthorn, citrus, lemon, grapefruit, apple, pear, cherry, carrot, and ginseng.

[0016] In certain embodiments, the preparation method according to the present invention, wherein further comprising a step of crushing the plant raw material before the extraction step.

[0017] In certain embodiments, the preparation method according to the present invention, wherein the precipitant comprises at least one of ethanol, methanol, propanol, isopropanol, acetone, and methyl ethyl ketone.

[0018] In certain embodiments, the preparation method according to the present invention, wherein the multi-stage separation and purification comprises protein removal, resin adsorption, anion exchange chromatography, and non-ionic gel chromatography separation.

[0019] In certain embodiments, the preparation method according to the present invention, wherein the multi-stage separation and purification comprises:

[0020] Preliminarily removing proteins and pigments in the crude extract to obtain a first purified product;

[0021] Separating the first purified product on an anion chromatography column, collecting the eluted fractions, and taking the fraction capable of lowering blood lipid levels as the second purified product;

[0022] Using water as the mobile phase, passing the second purified product through a non-ionic gel filtration column to obtain a third purified product, and taking the fraction capable of lowering blood lipid levels as the fourth purified product;

[0023] Using water as the mobile phase, passing the fourth purified product through a non-ionic gel filtration column again to obtain the HG-type pectin.

[0024] In the fourth aspect of the present invention, there is provided the use of HG-type pectin in the preparation of a drug for preventing, treating or improving dyslipidemia, wherein the main chain has a glycosidic bond as shown in →4)-α-D-GalAp-(1→, and the terminal α-D-GalAp-(1→ is connected to the main chain through →3,4)-α-D-GalAp-(1→, and the HG-type pectin has the structure shown in Formula I:

[0025] Formula I;

[0026] Wherein, n is selected from natural numbers from 1 to 30.

[0027] In the present invention, a crude extract is extracted from a plant raw material and subjected to multi-stage separation and purification, and an HG-type pectin with a single component having the activity of improving dyslipidemia is screened. In addition, by constructing an animal model, the present invention verifies that the HG-type pectin has the effect of improving dyslipidemia. The HG-type pectin of the present invention has broad application prospects in the fields of foods, drugs, etc. for preventing, improving or treating dyslipidemia. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1-2 Shows the separation and screening of HG-type pectin (named "SPC2A").

[0029] Figure 3-4 Shows the results of the structural and compositional analysis of SPC2A.

[0030] Figure 5-7 Shows the nuclear magnetic resonance analysis results of SPC2A.

[0031] Figure 8-10 Shows the morphological, three-dimensional structure and thermogravimetric analysis results of SPC2A.

[0032] Figure 11-13 Shows the effects of SPC2A on the body weight gain and dyslipidemia of zebrafish.

[0033] Figure 14-17 Shows the effects of SPC2A on the lipid accumulation in the liver of zebrafish. DETAILED DESCRIPTION OF THE INVENTION

[0034] The various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be construed as a limitation of the present invention, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present invention.

[0035] It should be understood that the terms used in the present invention are only for describing specific embodiments and are not intended to limit the present invention. Additionally, for the numerical ranges in the present invention, it should be understood that the upper and lower limits of the range and each intermediate value therebetween are specifically disclosed. Each intermediate value within any stated value or stated range and each smaller range between any other stated value or intermediate value within the stated range are also included in the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0036] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although the present invention only describes preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In case of conflict with any incorporated document, the content of this specification shall prevail.

[0037] HG-type pectin

[0038] In one aspect of the present invention, there is provided an HG-type pectin, the main chain of which has a glycosidic bond as shown by →4)-α-D-GalAp-(1→, and the terminal α-D-GalAp-(1→ is connected to the main chain through →3,4)-α-D-GalAp-(1→, and the HG-type pectin has the structure shown in Formula I:

[0039] Formula I;

[0040] Wherein,

[0041] n is selected from natural numbers from 1 to 30.

[0042] In a preferred embodiment, n is selected from natural numbers from 1 to 30, preferably from 1 to 25, more preferably from 1 to 20, still more preferably from 1 to 15, and even more preferably from 1 to 13, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13. In a specific embodiment, n is 13.

[0043] The present invention has carried out structural analysis and characterization on the obtained HG-type pectin. In a specific embodiment, high performance gel permeation chromatography (HP-GPC) is used for molecular weight determination, and the HG-type pectin described in the present invention has a weight average molecular weight of 12931 Da.

[0044] In a specific embodiment, by performing infrared spectroscopy analysis on the HG-type pectin, detection is carried out in the IR mode, and the scanning wavenumber range is 4000 cm -1 -400 cm -1, the number of scans is 32 times. The HG-type pectin of the present invention has an O-H stretching vibration peak of saccharide molecules at 3412 cm -1 ; has a stretching vibration peak of amide carbonyl C=O produced by uronic acid at 1740 cm -1 ; has a stretching vibration peak of free carboxylic acid carbonyl at 1608 cm -1 ; has a stretching vibration peak of C-O at 1417 cm -1 ; has a pyran ring absorption peak at 1048 cm -1 and 1018 cm -1 ; and has an α-glycosidic bond absorption peak between 890 - 970 cm -1 .

[0045] In a specific embodiment, the specific chemical structure of the HG-type pectin is determined by nuclear magnetic resonance analysis, and 1 H NMR, 13 C NMR, COSY, NOESY, HSQC, HMBC and TOCSY are measured with a 600 MHz nuclear magnetic resonance spectrometer. The results show that the HG-type pectin of the present invention has a hydrogen spectrum signal between 3.0 - 5.5 ppm, and has a sugar ring proton signal at δ 3.2 - 4.0 ppm, and an anomeric proton peak distributed in the region of 4.3 - 5.8 ppm, including signal peaks of δ 5.71, 5.23, 5.02, 4.85, 4.52.

[0046] In a specific embodiment, the HG-type pectin of the present invention has an anomeric carbon region between δ 93 - 180, wherein the HG-type pectin of the present invention has anomeric carbon signal peaks including δ 108.18, 101.34, 100.38, 97.48, 93.62, and signal peaks distributed in the region of 60 - 85 ppm, including signal peaks of δ 72.1, 71.31, 70.70, 71.69, 72.7, 82.55, 80.28, 71.83, 69.4, 70.05, 79.15, 72.65, 72.82, 73.28, 79.19, 72.75, 69.6, 75.3, 79.12, 72.7.

[0047] In a specific embodiment, the HG-type pectin of the present invention has an anomeric carbon signal peak at δ100.38 and an anomeric hydrogen signal peak at δ5.02 in the HSQC spectrum. The signals of H1-2 are 5.02 / 3.67; the signals of H2-3 are 3.67 / 3.93; the signals of H3-4 are 3.93 / 4.32; the signals of H4-5 are 4.32 / 4.99. Among them, H1, H2, H3, H4, and H5 are 5.02, 3.67, 3.93, 4.32, and 4.99 respectively, and the corresponding C1-5 are 100.38, 69.40, 70.05, 79.15, and 72.65.

[0048] In a specific embodiment, the HG-type pectin of the present invention has an anomeric hydrogen integration with a ratio of →4)-α-D-GalAp-(1→ and →3,4)-α-D-GalAp-(1→ of 3:1 in the H spectrum.

[0049] In a specific embodiment, according to the one-dimensional and two-dimensional NMR spectra, the anomeric hydrogen of the glycosidic bond →4)-α-D-GalAp-(1→ of the HG-type pectin of the present invention has a correlation signal peak with its own C4, indicating the existence of the linkage mode of →4)-α-D-GalAp-(1→4)-α-D-GalAp-(1→. And the anomeric hydrogen of →4)-α-D-GalAp-(1→ has a correlation peak with the C4 of the glycosidic bond →3,4)-α-D-GalAp-(1→ respectively, indicating the existence of the glycosidic bond →4)-α-D-GalAp-(1→3,4)-α-D-GalAp-(1→.

[0050] Pharmaceutical composition

[0051] In one aspect of the present invention, there is provided a pharmaceutical composition comprising the HG-type pectin described in the present invention and a pharmaceutically acceptable carrier.

[0052] In the present invention, the pharmaceutically acceptable carrier includes at least one of diluents, fillers, absorbents, wetting agents, binders, disintegrants, lubricants, sweeteners, preservatives, and antioxidants. Among them, examples of diluents include, but are not limited to, normal saline, aqueous buffer solutions, solvents, dispersion media, etc.; fillers include, but are not limited to, starch, lactose, mannitol, microcrystalline cellulose, etc.; absorbents include, but are not limited to, calcium sulfate, calcium hydrogen phosphate, calcium carbonate, etc.; wetting agents include, but are not limited to, water, ethanol, etc.; binders include, but are not limited to, hypromellose, povidone, microcrystalline cellulose, etc.; disintegrants include, but are not limited to, croscarmellose sodium, crospovidone, surfactants, low-substituted hydroxypropyl cellulose, etc.; lubricants include, but are not limited to, magnesium stearate, talc, polyethylene glycol, sodium lauryl sulfate, colloidal silicon dioxide, talc, etc.; sweeteners include, but are not limited to, sucralose, acesulfame, saccharin, sucrose, xylitol, mannitol, sorbitol, glucose, fructose, aspartame, etc.; preservatives include, but are not limited to, parabens, chlorobutanol, phenol, sorbic acid, etc.; antioxidants include, but are not limited to, ascorbic acid, methionine, etc.

[0053] In the present invention, the pharmaceutically acceptable carrier participates in transporting or delivering a drug from one organ or a certain part of the body to another organ or another part of the body. Each carrier is "acceptable", which means that it is compatible with other components of the formulation (such as HG-type pectin) and does not harm the patient.

[0054] Preparation method

[0055] In one aspect of the present invention, there is provided a method for preparing the HG-type pectin according to the present invention, which includes steps (1) and (2), and the following is a detailed description.

[0056] Step (1) of the present invention is a step of extracting a plant raw material with an aqueous solvent at 50 - 90 °C for 100 - 200 min to obtain an extract, and adding a precipitant for precipitation to obtain a crude extract.

[0057] In a preferred embodiment, the plant raw material is mixed with 5 - 20 times (preferably 6 - 19 times, more preferably 7 - 18 times, even more preferably 8 - 17 times, such as 8, 9, 10, 11, 12, 13, 14, 15, 16, 17 times) of an aqueous solvent, and subjected to a crushing treatment (for example, cell wall disruption is carried out at a speed of 5000 - 15000 r / min for 20 - 60 s), and extraction is carried out at 50 - 90 °C, preferably 52 - 88 °C, more preferably 54 - 86 °C, further preferably 56 - 84 °C, even more preferably 58 - 82 °C, even more preferably 60 - 80 °C, such as 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80 °C for 100 - 200 min, preferably 110 - 190 min, more preferably 120 - 180 min, such as 120, 130, 140, 150, 160, 170, 180 min to obtain an extract, which is filtered (for example, filtered through a 100 - 300 mesh sieve), the filtrate is centrifuged, and the supernatant is retained and concentrated to 5 - 10 times (such as 5, 6, 7, 8, 9, 10 times) the volume of the original fresh medicinal material, a precipitant is added for precipitation, and it is left to stand in the dark at room temperature for more than 8 h, then centrifuged again to discard the supernatant, the precipitate is added with water until completely dissolved, the alcohol smell is completely volatilized and concentrated to 3 - 8 times (such as 3, 4, 5, 6, 7, 8 times) the volume of the original fresh medicinal material to obtain a crude extract. It can be understood that in order to better extract the active ingredients, the extraction process can be repeated several times.

[0058] In the present invention, the aqueous solvent includes but is not limited to distilled water, deionized water, reverse osmosis water, ultrapure water, etc.; the plant raw material includes but is not limited to seabuckthorn, citrus, lemon, grapefruit, apple, pear, cherry, carrot, ginseng, etc.; the precipitant includes but is not limited to ethanol, methanol, propanol, isopropanol, acetone, methyl ethyl ketone, etc.

[0059] Step (2) of the present invention is the step of performing multi - stage separation and purification on the crude extract to obtain the HG - type pectin. In a preferred embodiment, the multi - stage separation and purification includes protein removal, resin adsorption, anion - exchange chromatography, and non - ionic gel chromatography separation.

[0060] In a preferred embodiment, the Sevag method is used for the protein removal process. In a specific embodiment, the crude extract obtained in step (1) and chloroform - n - butanol are mixed and shaken for 20 - 40 minutes to obtain an aqueous phase. It can be understood that in order to better remove proteins, the above - mentioned mixing and shaking processes can be repeated.

[0061] In a preferred embodiment, resin adsorption includes adding the above aqueous phase to a macroporous adsorption resin, adjusting the pH to acidic (e.g., pH 3, 4, 5, 6), decolorizing at 50 - 70 °C (preferably 52 - 68 °C, more preferably 54 - 66 °C, e.g., 54, 56, 58, 59, 60, 62, 64, 66 °C) for 70 - 150 min (preferably 80 - 140 min, more preferably 90 - 130 min, e.g., 90, 100, 110, 120, 130 min), filtering to remove the macroporous resin, centrifuging to remove impurities, and freeze-drying to obtain a first purified product.

[0062] In a preferred embodiment, the first purified product is dissolved in an aqueous solvent, and after removing impurities using a 0.45 μM aqueous filter membrane, it is loaded onto an anion chromatography column (examples include but are not limited to DEAE-cellulose DE-52 column, TSKgel BioAssist Q column, TSKgel DEAE-5PW column, TSKgel DEAE-2SW column, TSKgel DEAE-3SW column, TSKgel SuperQ-5PW column, TSKgel QAE-2SW column, Hamilton RCX-30 column, Hypersil SAX column, etc.). It is eluted with an aqueous solvent, 0.1 M NaCl, 0.3 M NaCl, 0.5 M NaCl, and 0.7 M NaCl respectively. The eluted product is detected for the peak position and the content of the purified product in each tube by the phenol-sulfuric acid method, an elution curve is plotted, the main fractions of the eluate are collected, the supernatant is dialyzed (3500 Da), and different purified products are obtained by freeze-drying.

[0063] To obtain a purified product with pharmacological activity, the fractions capable of downregulating blood lipid levels are screened, and the fractions capable of downregulating blood lipid levels are used as the second purified product for subsequent purification steps. The fractions capable of downregulating blood lipid levels particularly refer to purified products that can significantly reduce the amount of triglyceride and / or total cholesterol.

[0064] In a preferred embodiment, using water as the mobile phase, a non-ionic gel filtration column (examples include but are not limited to Sephacryl S-300 HR column, polystyrene gel chromatography column, porous silica gel column, Bio-Beads series gel chromatography column, Toyopearl series gel chromatography column, etc.) is used to further separate and purify the aqueous solution of the second purified product. The product is detected for the peak position and the content of the purified product in each tube by the m-hydroxybenzene method, the purified fractions are collected by combining, and a third purified product is obtained. The fractions capable of downregulating blood lipid levels are used as the fourth purified product.

[0065] It is understandable that the fourth purified product can be directly used as the HG-type pectin of the present invention, or as a preferred technical solution, using water as the mobile phase, passing the fourth purified product through a non-ionic gel filtration column again to obtain HG-type pectin with higher content and purity.

[0066] Application

[0067] One aspect of the present invention provides the use of the HG-type pectin of the present invention in the preparation of a drug for preventing, treating or improving dyslipidemia.

[0068] In the present invention, the terms "prevent, treat or improve" refer to therapeutic treatment and preventive or prophylactic measures, the purpose of which is to prevent or slow down (reduce) undesired physiological changes or disorders, such as the increase in blood lipids. Beneficial or desired clinical outcomes include, but are not limited to, the following whether detectable or undetectable outcomes, including the remission of symptoms, the reduction of the degree of disease, the stabilization of the disease state (i.e., no deterioration), the delay or slowdown of disease progression, the improvement or alleviation of the disease state, and the alleviation (whether partial or complete). Those in need of treatment include those who already suffer from hyperlipidemia or lipid-related diseases or those who need to prevent or improve hyperlipidemia or lipid-related diseases.

[0069] In the present invention, dyslipidemia includes at least one of abnormal total cholesterol, abnormal total triglyceride, abnormal low-density lipoprotein cholesterol, abnormal high-density lipoprotein cholesterol, abnormal apolipoprotein A1, and fatty liver.

[0070] Example 1

[0071] The following exemplarily shows the extraction and screening process of SPC2A.

[0072] 1. Extraction of SPC2A

[0073] Add fresh seabuckthorn berries to 12 times the amount of distilled water, break the wall in a blender at a speed of 9000 r / min for 40 s, and heat and extract at 70 °C for 150 minutes, repeating the heat extraction twice. Filter the extract with a 200-mesh silk cloth, centrifuge the filtrate, retain the supernatant, concentrate it to eight times the volume of the original fresh medicinal material, add ethanol (95%, v / v) four times the volume of the supernatant for precipitation, let it stand in the dark at room temperature for more than 8 h, centrifuge again to discard the supernatant, dissolve the precipitate in water until completely dissolved, thoroughly volatilize the alcohol smell and concentrate it to five times the volume of the original fresh medicinal material to obtain a crude extract. The extraction rate under this process is 1.2%, and the purity is 54%.

[0074] 2. Separation and purification of SPC2A

[0075] Remove the protein mixture in the crude extract according to the Sevag method. Take the crude extract solution, add 1 / 4 volume of chloroform-n-butanol (volume ratio 4:1), shake well for 30 minutes, and then separate the aqueous phase from the chloroform phase. Add the aqueous phase to another 1 / 4 volume of chloroform-n-butanol solution and repeat this process twice. Leave the aqueous layer in the separating funnel. Use macroporous adsorption resin to remove the pigments in the crude extract solution. Add 1 / 5 mass of macroporous adsorption resin, adjust the pH to 5, decolorize at 60 °C for 110 min, filter to remove the macroporous resin, then centrifuge the crude extract solution to remove impurities, and freeze-dry to obtain the crude seabuckthorn extract, with a purity of 67% at this time.

[0076] Dissolve the crude seabuckthorn extract in distilled water and separate it on a DEAE-cellulose DE-52 chromatographic column. Weigh 10 g of the crude seabuckthorn extract, dissolve it in 50 mL of distilled water, filter through a 0.45 μM aqueous filter membrane to remove impurities, and then load the sample. Elute with distilled water, 0.1 M NaCl, 0.3 M NaCl, 0.5 M NaCl, and 0.7 M NaCl respectively. Detect the peak position and sugar content of each of the 700 eluted products by the phenol-sulfuric acid method and draw the elution curve. Collect the main fractions of the eluate, dialyze the supernatant (3500 Da), and freeze-dry to obtain the purified components SPA, SPB, SPC, and SPD ( Figure 1 of A). Using hyperlipidemic zebrafish larvae induced by a high-fat diet as a model and triglyceride and total cholesterol as indicators, it was screened that the SPC component has the best effect on regulating blood lipids ( Figure 2 ).

[0077] Further purify SPC on a Sephacryl S-300HR column using triple-distilled water as the mobile phase (flow rate 1.0 mL / min). Weigh 40 mg of SPC, dissolve it in 2 mL of triple-distilled water, filter through a 0.22 μM aqueous filter membrane to remove impurities, and then load the sample. Detect the peak position and sugar content of each of the 40 eluted products by the m-hydroxydiphenyl method and draw the elution curve of SPC on the ion-exchange column. Combine and collect according to the elution curve, and after multiple purifications, collect the same components and freeze-dry to obtain the purified fractions (SPC1, SPC2) ( Figure 1 of B). SPC2 is separated and purified again on a Sephacryl S-300 HR column using distilled water solution as the mobile phase to obtain the final product SPC2A ( Figure 1 of C). Using the hyperlipidemic zebrafish larvae model in the same way, it was screened that SPC1 and SPC2 have comparable effects on regulating total cholesterol (p<0.05), but the SPC2 component has a more significant effect on regulating total triglyceride (p<0.01) ( Figure 2 of C, D).

[0078] Example 2

[0079] The structure and composition analysis of SPC2A are shown below.

[0080] 1. Protein content detection

[0081] Accurately weigh 50 mg of bovine serum albumin and dissolve it in an appropriate amount of distilled water, make up the volume to 100 mL and shake well. Add 0 μL, 0.4 μL, 0.8 μL, 1.2 μL, 1.6 μL, 2.0 μL, 2.4 μL of the protein mother liquor into 6 clean test tubes respectively, and make up the total volume to 15 μL with deionized water. Then add 5 mL of Coomassie Brilliant Blue solution, shake well, place it at room temperature for 5 min, and measure the absorbance at a wavelength of 595 nm. With the protein concentration as the abscissa and the absorbance as the ordinate, draw a standard curve. The regression equation of the straight line is y = 14.178x + 0.0117, and the fitting degree R² = 0.9944. According to the standard curve, the protein content in the purified sample is calculated to be 0.001 mg / mL. The protein removal of the crude seabuckthorn extract can effectively remove most of the proteins in SPC2.

[0082] 2. Ultraviolet spectrum analysis

[0083] Accurately weigh 1.00 mg of SPC2A and dissolve it in distilled water to form a solution with a concentration of 1 mg / mL. Using distilled water as a control, detect the absorption light curve of the sample in the range of 190 - 400 nm, and observe the absorbance values of SPC2A at 280 nm and 260 nm. Conduct a full-wavelength scan of SPC2A, showing that there are no characteristic absorption peaks at 230 nm and 260 nm ( Figure 3 A), indicating that SPC2A contains almost no protein, which is consistent with the results of the Coomassie Brilliant Blue assay.

[0084] 3. Molecular weight analysis

[0085] High performance gel permeation chromatography was used for molecular weight determination. Chromatographic conditions (chromatographic column: BRT105 - 103 - 101 (8×300 mm); detector: differential refractive index detector RID - 20A; mobile phase: 0.05 M NaCl solution; flow rate: 0.7 mL / min; column temperature: 40°C, injection volume: 25 μL). Inject the samples in ascending order of relative molecular weight, record the retention time, draw the standard curves of lgMp - RT (Mp peak molecular weight), lgMw - RT (Mw weight average molecular weight), lgMn - RT (Mn number average molecular weight), obtain the molecular weight calculation formula, and detect the samples under the same above conditions.

[0086] The chromatogram of SPC2A shows a single symmetric peak ( Figure 3B), indicating that its composition is single and the purity is good. According to the molecular weight determination method, a linear regression equation was established using dextrans with a series of molecular weights as standards, and the calibration curves of lgMp-RT (peak molecular weight), lgMw-RT (weight-average molecular weight), and lgMn-RT (number-average molecular weight) were obtained. The equation of the lgMp-RT calibration curve is: y = -0.1345x + 9.4624, R² = 0.9967; the equation of the lgMw-RT calibration curve is: y = -0.1361x + 9.514, R² = 0.997; the equation of the lgMn-RT calibration curve is: y = -0.1354x + 9.4855, R² = 0.9965. According to the standard curve, the calculation formula was obtained and the molecular weight of each sample was calculated. Substituting the retention time of 39.694 min in the SPC2A chromatogram into the regression equation, the weight-average molecular weight of SPC2A was calculated to be approximately 12931 Da. The dispersity (Mw / Mn) is used to describe the width of the molecular weight distribution. According to HP-GPC analysis, the dispersity of SPC2A is 1.001, close to 1, indicating that its molecular weight distribution is narrow and the molecular weights are relatively uniform.

[0087] 4. Infrared Spectroscopy Analysis

[0088] The purified sample was mixed with potassium bromide at a mass ratio of 1:100 and ground into a powder. After pressing the sample into a tablet using a tablet press, it was detected in IR mode, and the scanning wavenumber range was 4000 cm -1 -400 cm -1 , and the number of scans was 32 times.

[0089] Analysis of the infrared spectrum of SPC2A found that the broad peak at approximately 3412 cm -1 is the characteristic O-H stretching vibration peak between sugar molecules; the stretching vibration of the amide carbonyl C=O at 1740 cm -1 is the absorption peak produced by uronic acid; the absorption peak at 1608 cm -1 is the stretching vibration of the free carboxylic acid carbonyl; the absorption peak at 1417 cm -1 is the stretching vibration of C-O ( Figure 4 A). In addition, the prominent absorption peaks at 1048 cm -1 and 1018 cm -1 are the pyran rings in the SPC2A structure, and the weaker absorption peaks between 890 - 970 cm -1 are α-glycosidic bonds.

[0090] 5. Monosaccharide Composition Analysis

[0091] After hydrolyzing SPC2A with trifluoroacetic acid, the monosaccharide composition in the hydrolysis product was determined using an ion chromatograph. Chromatographic conditions (the chromatographic column was Thermo Scientific™ Dionex™ CarboPac™ PA100; mobile phase A was 20 mM NaOH; mobile phase B was 100 mM NaOH + 500 mM NaAc; the flow rate was 1 mL / min; the column temperature was 30°C). The main monosaccharides in the acid hydrolysate of the crude extract sample analyzed by ion chromatography were GalA (galacturonic acid), with only small amounts of Rha (rhamnose) and Ara (arabinose) present ( Figure 4 of B).

[0092] Example 3

[0093] The structural analysis of SPC2A is shown below.

[0094] 1. Methylation analysis

[0095] Weigh 20 mg of SPC2A and make up the volume to 4 mL with triple-distilled water. Add 800 μL of 0.2 M morpholineethanesulfonic acid (MES) and 1.6 mL of 500 mg / mL carbodiimide (EDC) solution, and react at 25°C for 3 h. Add 4 mL of 4 M imidazole-HCl, weigh 168 mg of sodium borotritide and pour it directly into the reaction solution, and react at room temperature overnight. The next day, add 2 mL of glacial acetic acid under ice bath conditions to terminate the reaction. After desalting the reaction solution by dialysis, freeze-dry it.

[0096] Take 2 mg of the freeze-dried SPC2A in 0.5 mL of DMSO (dried with molecular sieve), dissolve it ultrasonically in a stoppered glass tube. Under nitrogen protection, add 20 mg of dry NaOH, ultrasonically dissolve it completely, and then continue to add 0.3 mL of methyl iodide under nitrogen protection. Stir and react at room temperature in the dark for 2.5 h, add 0.3 mL of ultrapure water and stir for 0.5 h to terminate the reaction. Continue to add 1 mL of dichloromethane to wash the reaction tube and transfer it all to a clean glass test tube. Add 3 mL of ultrapure water to the glass tube for extraction, take the lower organic phase, extract with ultrapure water repeatedly 3 times. After adding an excess of NaSO4 to adsorb the excess water in the lower layer, transfer it to a clean glass tube and dry it with nitrogen. Repeat the above reaction 3 times until the hydroxyl groups are completely substituted.

[0097] Add 1 mL of 2 M TFA to the methylated sample above, react at 110 °C for 2 h. After hydrolysis is completed, place it in an evaporating dish, heat it in a water bath at 70 °C until it is evaporated to dryness, add a small amount of methanol and evaporate it to dryness. Repeat this process 5 times until the pH of the aqueous solution is neutral. Then add 0.3 mL of ultrapure water and 1 drop of 1% ammonia water to adjust the pH to alkaline. Add 5 mg of NaBD4 to the hydrolysis product, mix well, stir at room temperature, and carry out a reduction reaction for 12 h. After the reaction is completed, adjust the pH to neutral with acetic acid. After evaporating the reaction solution to dryness, add methanol and wash and evaporate it to dryness repeatedly. Repeat this process 3 times. Then add 1 mL of acetic anhydride:pyridine (1:1 = v / v) and react at 90 °C for 2 h, add 1 mL of ultrapure water to terminate the reaction. Then extract twice with 1 mL of dichloromethane. After sufficient shaking, collect the dichloromethane phase and evaporate it to dryness. Finally, add 0.6 mL of dichloromethane to redissolve it, filter it with a 0.22 μm organic filter membrane, and perform GC-MS analysis. Detection is carried out using a gas chromatography quadrupole time-of-flight high-resolution mass spectrometer 8890-7250. Capillary column: HP-5MS (0.25 μm, 30 m × 0.25 mm).

[0098] 2. Nuclear magnetic resonance analysis

[0099] Weigh 50 mg of the SPC2A sample, dissolve it in 0.5 mL of heavy water and freeze-dry it. Subsequently, dissolve the freeze-dried powder in 0.5 mL of heavy water again and continue to freeze-dry it. Repeat the above process to fully exchange the labile hydrogen. Then dissolve the sample in 0.5 mL of heavy water and place it at room temperature of 25 °C for measurement on a nuclear magnetic resonance spectrometer at 600 MHz 1 1H NMR, 13 13C NMR, COSY, NOESY, HSQC, HMBC and TOCSY.

[0100] The hydrogen spectrum signals are mainly concentrated between 3.0 - 5.5 ppm. The signals of δ3.2 - 4.0 ppm are the proton signals of the sugar ring. The main anomeric proton peaks δ5.71, 5.23, 5.02, 4.85, 4.52 are concentrated in the region of 4.3 - 5.8 ppm ( Figure 5 A of).

[0101] The carbon spectrum analysis is carried out at 1313C NMR (201 MHz, D2O): The signals of 13C NMR are mainly concentrated between 60 - 120 ppm. By observing the 13C NMR spectrum, it can be seen that the main anomeric carbon signal peaks are δ108.18, 101.34, 100.38, 97.48, 93.62. The anomeric carbon region is mainly between δ93 - 180. And δ72.1, 71.31, 70.70, 71.69, 72.7, 82.55, 80.28, 71.83, 69.4, 70.05, 79.15, 72.65, 72.82, 73.28, 79.19, 72.75, 69.6, 75.3, 79.12, 72.7. The main signal peaks are distributed in the 60 - 85 ppm region ( Figure 5 B). According to the results of monosaccharide composition, the purified product is mainly composed of galacturonic acid, indicating that it is mainly galacturonan, that is, HG-type pectin.

[0102] Through the HSQC spectrum ( Figure 6 B), the anomeric carbon signal can be observed as δ100.38, and the corresponding anomeric hydrogen signal in the HSQC spectrum is δ5.02. Through HH-COSY ( Figure 6 A), the signal of H1-2 is 5.02 / 3.67; the signal of H2-3 is 3.67 / 3.93; the signal of H3-4 is 3.93 / 4.32; the signal of H4-5 is 4.32 / 4.99; indicating that H1, H2, H3, H4, H5 are δ5.02, 3.67, 3.93, 4.32, 4.99 respectively, and the corresponding C1-5 are 100.38, 69.40, 70.05, 79.15, 72.65. Therefore, this signal belongs to the glycosidic bond →4)-α-GalAp-(1→. According to similar rules and combined with HMBC ( Figure 7 A) and TOCSY ( Figure 7 B), all glycosidic bond signals are assigned, as shown in Table 1.

[0103] It is found from the integration of anomeric hydrogen in the 1H spectrum that the ratio of →4)-α-D-GalAp-(1→ and →3,4)-α-D-GalAp-(1→ is about 3:1. Therefore, in the HMBC spectrum, according to the one-dimensional and two-dimensional NMR spectra, the glycosidic bond signals of SPC2A are assigned; the anomeric hydrogen of the glycosidic bond →4)-α-D-GalAp-(1→ has a correlation signal peak with its own C4; indicating the existence of the linkage mode of →4)-α-D-GalAp-(1→4)-α-D-GalAp-(1→. The anomeric hydrogen of →4)-α-D-GalAp-(1→ has correlation peaks with the C4 of the glycosidic bond →3,4)-α-D-GalAp-(1→ respectively, indicating the existence of the glycosidic bond →4)-α-D-GalAp-(1→3,4)-α-D-GalAp-(1→.

[0104] In summary, the main glycosidic bond structure of HG pectin is as follows: the main chain connection is the glycosidic bond of →4)-α-D-GalAp-(1→, and the terminal α-D-GalAp-(1→) is connected to the main chain through the O-3 bond of →3,4)-α-D-GalAp-(1→, and the structural formula is as follows: Figure 7 C, where n is 13.

[0105] Table 1

[0106]

[0107] Example 4

[0108] The morphology, three-dimensional structure, and thermogravimetric analysis of SPC2A are shown below.

[0109] 1. Scanning Electron Microscope Analysis

[0110] About 5 mg of dried SPC2A was taken and adhered to a conductive carbon film containing a double-sided adhesive, placed in the sample chamber of an ion sputtering instrument, and sprayed with gold for about 40 s. After the sample was taken out, it was placed in the observation chamber of an ultra-high resolution scanning electron microscope with an accelerating voltage of 5 kV.

[0111] Under the microscope of 500x and 1000x, the samples are mainly in the form of flakes with relatively smooth surfaces. When further magnified to 2000x, it can be seen that some flake samples have a small amount of attachments on their surfaces. After magnification to 5000x, the smooth surface is more obvious ( Figure 8 ).

[0112] 2. Atomic Force Microscopy Analysis

[0113] The ultrastructure of the sample was measured using a Bruker ICON atomic force microscope. SPC2A was dissolved in ethanol water (20 μg / mL), placed in a 60°C water bath, and shaken for 120 min. The treated solution (10 μL) was placed on a mica sheet and dried at 25°C for 12 h. AFM scanning was performed at room temperature in the tapping mode with a scanning range of 5*5 μm.

[0114] Planar and three-dimensional images of SPC2A Figure 9 As shown. Among them, SPC2A is mainly linear and a small amount of irregular round particles, randomly distributed and of different sizes. The height of SPC2A is -4.8-3.8 nm, which is inconsistent with the average height of single-chain polysaccharides, indicating that SPC2A is easier to aggregate and entangle in the solvent, and its higher aggregates may be due to the existence of van der Waals forces and other forces between its chains. It can be seen that the uneven surface structure of SPC2A has sample aggregation and stacking.

[0115] 3. Congo Red Analysis

[0116] The Congo red experiment was used to determine whether SPC2A has a triple-helix chain structure. Weigh 5 mg of SPC2A and add 10 mL of distilled water and an equal volume of 50 μmol / L Congo red reagent. After mixing, prepare NaOH solutions with different concentrations. Specifically, gradually add 2 mol / L NaOH solution to increase the final concentration of NaOH in the solution from 0.0 mol / L to 0.5 mol / L, and scan with a UV-visible recording spectrometer to measure the maximum absorption wavelength under each NaOH concentration condition. Plot a graph with the NaOH concentration as the abscissa and the maximum absorption wavelength as the ordinate. Among them, deionized water is used instead of the HG-type pectin solution as a blank control.

[0117] The results are as Figure 10 shown in Figure A. Compared with the blank Congo red solution, the maximum absorption wavelength of the sample in Congo red basically did not change. That is, in the range of NaOH concentration from 0.1 - 0.5 mol / L, in the same NaOH solution, after adding the HG-type pectin, the maximum absorption wavelength of Congo red did not change significantly, that is, there was no obvious red shift in absorption, indicating that the HG-type pectin has a disordered structure.

[0118] 4. TG / DSC Thermogravimetric Analysis

[0119] The thermogravimetric analyzer was used to analyze the thermal stability of the sample. Accurately weigh 3 mg of SPC2A and place it in a crucible. After pressing, use a thermogravimeter to increase the temperature from 25°C to 800°C at a heating rate of 10°C / min under nitrogen protection.

[0120] From the thermogravimetric curve Figure 10 shown in Figure B, it can be seen that the overall thermogravimetric curve is divided into three stages: the first weight loss stage is 25 - 206.4°C, and SPC2A mainly loses free water with a weight loss rate of 36.3%; the second stage is around 206.5 - 301.9°C, and there is an obvious weight loss step in this stage with a weight loss rate reaching 45.4%; the third stage is around 302.0 - 787.0°C, and there is no obvious weight loss step in this stage with a weight loss rate reaching 69.4%; the total weight loss rate of the sample is 69.4%. This indicates that in this temperature range, the HG-type pectin decomposed and the chemical bonds were damaged.

[0121] It can be seen from the differential scanning calorimetry curve that SPC2A shows a downward trend from room temperature to 259.7 °C, indicating that SPC2A exhibits exothermic behavior at this temperature, and shows an upward trend from 259.8 - 275.5 °C, indicating that SPC2A exhibits endothermic phenomenon at this temperature. Then SPC2A shows a linear downward trend from 275.6 - 602.4 °C, indicating that SPC2A exhibits exothermic behavior at this temperature, and shows an upward trend from 602.5 - 679.9 °C, indicating that SPC2A exhibits endothermic phenomenon at this temperature. Finally, the sample shows a downward trend from 680.0 - 707.0 °C, indicating that SPC2A exhibits exothermic behavior at this temperature, and shows a linear upward trend from 707.1 - 787.0 °C, indicating that SPC2A exhibits endothermic phenomenon at this temperature.

[0122] Example 5

[0123] The following shows the efficacy evaluation of SPC2A.

[0124] 1. Experimental method

[0125] Three-month-old male zebrafish of the AB strain were purchased from the Chinese Zebrafish Resource Center (Wuhan, China) and raised in a zebrafish culture system with the breeding conditions controlled at a temperature of 28 °C, 0.2% edible sea salt, pH 6.9 - 7.2, and conductivity of 480 - 510 mS·cm -1 , in water with a hardness of 53.7 - 71.6 mg / L CaCO3, with a 14 h:10 h light-dark cycle per day. 180 adult male zebrafish were divided into 6 groups, namely the blank control group, the high-fat diet group (HFD), the low-dose SPC2A group (SPC2A-10) (aqueous solution of HG pectin), the medium-dose SPC2A group (SPC2A-50), the high-dose SPC2A group (SPC2A-100), and the positive group (bezafibrate), with 30 fish in each group. Throughout the experimental period, the zebrafish in the blank control group were continuously fed a normal diet (ND) composed of brine shrimp larvae bait, and the zebrafish in the HFD group were continuously fed a high-fat diet prepared by adding 7% (w / w) egg yolk powder and 7% (w / w) cholesterol (Shanghai Feixi Biotechnology Co., Ltd.) to the normal diet. The other four groups were fed a high-fat diet in the first 10 days; on the 11th day, the zebrafish in the bezafibrate group began to be fed the positive drug group prepared by adding bezafibrate to the high-fat diet (70 mg / kg per fish), and the SPC2A groups began to be fed the SPC2A treatment groups prepared by adding SPC2A to the high-fat diet (10, 50, 100 mg / kg per fish). Feed twice a day for 50 days. The administration method is as Figure 11 in A.

[0126] At the end of the 60-day experimental period, the zebrafish were fasted overnight, anesthetized in an ice-water mixture, weighed, and the fish tails were quickly cut off with a blade between the anal fin and the caudal fin. Blood from the dorsal aorta was aspirated into a centrifuge tube using a micropipette. The centrifuge tube was pre-rinsed with sodium heparin and left to stand at room temperature for 15 min. Plasma was collected by centrifugation at 1500 g for 5 min. The zebrafish were dissected under a stereomicroscope, and the liver was quickly separated and placed into a microcentrifuge tube, frozen in liquid nitrogen, and stored at -80 °C for kit detection. Another part of the liver was fixed in 4% paraformaldehyde for pathological observation.

[0127] 2. Effects on zebrafish body weight gain and dyslipidemia

[0128] Examination of zebrafish body weight and dyslipidemia showed that compared with zebrafish in the normal diet group, zebrafish in the high-fat diet group had enlarged abdomens and wider body shapes. Drug treatment could alleviate this trend of obesity, especially in the high-dose SPC2A group and bezafibrate ( Figure 11 B). At the same time, compared with the blank control group, the body weight of zebrafish in the HFD group increased significantly, while SPC2A intervention significantly reduced the body weight gain caused by the high-fat diet ( Figure 12 A). It is worth noting that there were no significant differences in feeding time among the groups ( Figure 12 B). This indicates that SPC2A can alleviate dyslipidemia caused by a high-fat diet without affecting appetite and energy intake. By observing the plasma lipid profile, it was found that compared with the blank control group, the levels of total cholesterol (TC), total triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) in the plasma of the HFD group were significantly increased, while the level of high-density lipoprotein cholesterol (HDL-C) was significantly decreased ( Figure 13 ). Compared with the HFD group, SPC2A intervention could significantly reduce the levels of TC and LDL-C and increase the level of HDL-C, and there was no obvious dose-dependent relationship (p < 0.05). However, SPC2A intervention could significantly reduce the TG level in a dose-dependent manner (p < 0.05).

[0129] 3. Effects on lipid accumulation in the zebrafish liver

[0130] Long-term feeding with a high-fat diet can induce lipid metabolism disorders in zebrafish, and the liver is prone to hepatic steatosis, increasing the susceptibility to liver injury. In this example, the liver weight of zebrafish in the high-fat diet group increased compared with that of zebrafish in the normal diet group, and SPC2A treatment further reduced the liver weight of zebrafish in the high-fat diet group ( Figure 14 A). Compared with the N blank control group, the liver weight index (liver weight / body weight) of zebrafish induced by a high-fat diet increased significantly, and bezafibrate and SPC2A treatment inhibited the increase in the liver weight index ( Figure 14B). There were significant statistical differences in the liver TC and TG levels between the blank control group and the HFD group of zebrafish. SPC2A and bezafibrate could reduce the levels of liver TC and TG. In zebrafish with dyslipidemia induced by a high-fat diet, the effect of SPC2A on reducing TG was more significant than that on reducing TC ( Figure 14 C and D). Compared with the normal group, the activities of plasma AST and ALT in the HFD group of zebrafish increased, while the activities of ALT and AST in the plasma of zebrafish after SPC2A intervention were significantly lower than those in the HFD group ( Figure 14 E and F). These results indicate that the use of SPC2A can regulate dyslipidemia in zebrafish and protect liver injury to a certain extent.

[0131] To further confirm the effect of SPC2A on liver lipid accumulation in zebrafish with dyslipidemia, the appearance and microstructure of the zebrafish liver were examined microscopically. As Figure 15 shown, from the perspective of the liver morphology under the microscope, the livers of zebrafish in the HFD group were swollen, with a lighter color, showing fatty degeneration. However, the color and morphology of the livers of zebrafish in the SPC2A intervention group showed a tendency to recover, and the liver state was relatively healthy. HE staining showed that the hepatocytes of zebrafish in the HFD group were significantly damaged, producing a large number of lipid droplets. SPC2A could significantly reduce the liver fatty degeneration and ballooning lesions caused by a high-fat diet and reduce the lipid droplet accumulation in the liver ( Figure 16 ). Nile red staining further showed the lipid accumulation in the livers of zebrafish in the HFD group. In the liver sections of the SPC2A group, this characteristic was significantly reduced (p < 0.0001) ( Figure 17 ).

[0132] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions recorded in the foregoing embodiments, or perform equivalent replacements on some of the technical features. And these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. An HG-type pectin, characterized in that, Its main chain has a glycosidic bond as shown by →4)-α-D-GalAp-(1→, and the terminal α-D-GalAp-(1→ is connected to the main chain through →3,4)-α-D-GalAp-(1→. The HG-type pectin has the structure shown in Formula I: Formula I; Wherein, n is 13, and the weight-average molecular weight of the HG-type pectin is 12,931 Da.

2. A pharmaceutical composition, characterized in that, It includes the HG-type pectin described in Claim 1.

3. The pharmaceutical composition according to claim 2, wherein The pharmaceutical composition further includes a pharmaceutically acceptable carrier, and the pharmaceutically acceptable carrier includes at least one of a diluent, a filler, an absorbent, a wetting agent, a binder, a disintegrant, a lubricant, a sweetening agent, a preservative, and an antioxidant.

4. The preparation method of HG-type pectin according to claim 1, characterized in that, It includes the following steps: (1) Using an aqueous solvent to extract plant raw materials at 50 - 90 °C for 100 - 200 min to obtain an extract, adding a precipitant for precipitation to obtain a crude extract; (2) Subjecting the crude extract to multi-stage separation and purification to obtain the HG-type pectin.

5. The preparation method according to claim 4, characterized in that, The plant raw materials include at least one of seabuckthorn, citrus, lemon, grapefruit, apple, pear, cherry, carrot, and ginseng.

6. The preparation method according to claim 4, characterized in that, The multi-stage separation and purification include protein removal, resin adsorption, anion exchange chromatography, and non-ionic gel chromatography separation.

7. The preparation method according to claim 4, characterized in that, The multi-stage separation and purification include: Preliminarily removing proteins and pigments in the crude extract to obtain a first purified product; Separating the first purified product on an anion chromatography column, collecting the eluted fractions, and using the fraction capable of lowering blood lipid levels as the second purified product; Using water as the mobile phase, passing the second purified product through a non-ionic gel filtration column to obtain a third purified product, and using the fraction capable of lowering blood lipid levels as the fourth purified product.

8. The preparation method according to claim 4, characterized in that, The precipitant includes at least one of ethanol, methanol, propanol, isopropanol, acetone, and methyl ethyl ketone.

9. Use of HG pectin in the preparation of a medicament for preventing, treating or improving dyslipidemia, characterized in that, Its main chain has a glycosidic bond as shown by →4)-α-D-GalAp-(1→, and the terminal α-D-GalAp-(1→ is connected to the main chain through →3,4)-α-D-GalAp-(1→. The HG-type pectin has the structure shown in Formula I: Formula I; Wherein, n is 13, and the weight-average molecular weight of the HG-type pectin is 12,931 Da.

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