CN107456459B - Roughhaired holly root polysaccharide and application thereof in preparation of anticomplement medicines - Google Patents

Roughhaired holly root polysaccharide and application thereof in preparation of anticomplement medicines Download PDF

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CN107456459B
CN107456459B CN201610393196.4A CN201610393196A CN107456459B CN 107456459 B CN107456459 B CN 107456459B CN 201610393196 A CN201610393196 A CN 201610393196A CN 107456459 B CN107456459 B CN 107456459B
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陈道峰
卢燕
温泉
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Abstract

The invention belongs to the field of traditional Chinese medicines, and relates to three homogeneous polysaccharides in a roughhaired holly root aqueous extract and application thereof in preparation of anticomplement medicines. The invention separates three radix Ilicis Asprellae homogeneous polysaccharides GDS1, GDS2 and GDS3 from radix Ilicis Asprellae which is a traditional Chinese medicine for clearing away heat and toxic material, and the results of in vitro tests prove that the prepared radix Ilicis Asprellae polysaccharides GDS1, GDS2 and GDS3 have obvious inhibition effects on cell hemolysis caused by activation of classical pathway and alternative pathway of complement, and have obvious inhibition effects on both classical pathway and alternative pathway of complement activation, wherein CH50The values are respectively 0.134 +/-0.024 mg/mL, 0.187 +/-0.031 mg/mL and 0.098 +/-0.011 mg/mL; AP (Access Point)50The values were 0.297. + -. 0.037mg/mL, 0.312. + -. 0.033mg/mL and 0.195. + -. 0.030mg/mL, respectively. The roughhaired holly root polysaccharide provided by the invention can be used as an active ingredient for further preparing natural anticomplement medicines.

Description

Roughhaired holly root polysaccharide and application thereof in preparation of anticomplement medicines
Technical Field
The invention belongs to the field of traditional Chinese medicines, relates to polysaccharide, and particularly relates to natural homogeneous polysaccharide in a traditional Chinese medicine holly and application thereof in preparation of anticomplement medicines.
Background
Respiratory system virus infection (such as SARS, H1N1 and other virus infections) is easy to cause acute lung injury, which in turn leads to Acute Respiratory Distress Syndrome (ARDS), which not only has strong infectivity, but also has high lethality rate and great harmfulness. Studies have shown that the pathogenesis of the disease is characterized by excessive complement system activation, production of a number of bioactive complement products, vasodilation, release of inflammatory mediators, and induction of systemic inflammatory responses, which results in tissue damage. Therefore, it is considered that inhibition of excessive activation of the complement system is a preferred clinical treatment for respiratory viral infection diseases.
In clinical practice, the traditional Chinese medicine for clearing heat and removing toxicity is often used for treating respiratory system virus infection diseases, and has good effects of inhibiting inflammation and removing heat. The earlier stage research of the subject group shows that the heat-clearing and detoxifying traditional Chinese medicines such as selfheal, houttuynia cordata and bupleurum polysaccharide all show good anticomplementary activity. Therefore, the search for complement inhibition from heat-clearing and detoxifying traditional Chinese medicines is attracting attention of those skilled in the art due to their efficiency and pertinence.
The radix Ilicis Asprellae is dry root of Ilex asprella (hook. et Arn.) of Ilex asprella of Aquifoliaceae (Aquifoliaceae), is a commonly used heat-clearing and toxicity-removing medicine in Lingnan of China, and has the effects of relieving sore throat, relieving swelling, removing blood stasis and relieving pain. The roughhaired holly root is widely applied clinically, and a plurality of Chinese patent medicines containing the roughhaired holly root, such as qingkailing, roughhaired holly root throat clearing granules and the like, can be used for treating tonsillitis, pneumonia and pulmonary infection caused by cold and fever, and have obvious effect. Modern pharmacological research shows that the roughhaired holly root has obvious functions of antipyresis, analgesia, anti-inflammation, immunity regulation and the like, but no research report about the anticomplementary activity of the roughhaired holly root and polysaccharide thereof is available so far.
Disclosure of Invention
The invention aims to provide a component with anticomplementary activity in traditional Chinese medicinal materials, and particularly relates to roughhaired holly root polysaccharide and application thereof in preparing anticomplementary medicines.
The invention carries out active guide separation on the aqueous extract of the traditional Chinese medicine roughhaired holly root to prepare roughhaired holly root polysaccharide, wherein the roughhaired holly root polysaccharide comprises three homogeneous polysaccharides which are respectively named as GDS1, GDS2 and GDS 3. In vitro hemolysis experiment shows that the polysaccharide of roughhaired holly root has excellent complement inhibiting activity and may be used as active medicine component in developing and preparing anticomplement medicine.
In the invention, the traditional Chinese medicine roughhaired holly root is selected from dry root of ilex aegilops taux (hook. et arn.) of Aquifoliaceae (Aquifoliaceae).
The roughhaired holly polysaccharide GDS1, GDS2 and GDS3 have the following structural characteristics:
(1) GDS1 is a heteropolysaccharide composed mainly of 7 monosaccharides, containing a small amount of glucuronic acid, and having a molar ratio of sugar residues of xylose (Xyl), rhamnose (Rha), arabinose (Ara), mannose (Man), glucose (Glc), galactose (Gal), galacturonic acid (GalA) of 1.0:2.7:8.8:3.1:28:24.8: 3.4; the molecular weight is 92.77 KDa; specific rotation degree
Figure BDA0001009626750000021
CD spectrum: 202nm (-4.594); protein content: 0.64 percent; content of uronic acid: 8.8 percent; sugar content 95.8%; the main connection mode is as follows: the main linkage modes of galactose are 1,3, 4-linkage, 1, 4-linkage and end linkage, and a small amount of 1,4, 6-linkage and 1, 3-linkage galactose; the primary mode of arabinose attachment is 1, 3-linked and end-linked arabinose; the mannose is linked in a 1, 4-linkage manner; the connection mode of glucose is 1,2,3, 4-connection and end connection, and the connection mode of rhamnose is 1, 3-connection and end connection; in addition, it also contains a small amount of 2, 3-linked xylose. Wherein the end-linked galactose, 1,3, 4-linked galactose and 1, 3-linked arabinose are the main components;
(2) GDS2 is a heteropolysaccharide composed mainly of 7 monosaccharides, containing a small amount of glucuronic acid, with a molar ratio of sugar residues xylose (Xyl), rhamnose (Rha), arabinose (Ara), mannose (Man), glucose (Glc), galactose (Gal), galacturonic acid (GalA) of 1.0:1.5:2.3:1.6:1.8:4.6: 1.7; the molecular weight is 16.22 KDa; specific rotation degree
Figure BDA0001009626750000022
CD spectrum 201nm (-2.242), 223nm (1.882); protein content: 0.64 percent; content of uronic acid: 15.5 percent; sugar content 93.6%; the main connection mode is as follows: the connection mode of galactose is 1,3, 4-connection, 1, 3-connection and end connection; the connection mode of arabinose is 1, 3-connection and end connection; mannose is linked in a 1, 4-linkage and end-linkage manner, and a small amount of 1, 3-linked mannose; the connection mode of glucose is 1,2,3, 4-connection, 1, 6-connection and end connection, and the connection mode of rhamnose is 1, 3-connection and end connection; in addition, it also contains 2, 3-linked xylose;
(3) GDS3 is a heteropolysaccharide composed mainly of 4 monosaccharides, containing small amounts of xylose and arabinose, and having a molar ratio of sugar residues of mannose (Man), glucose (Glc), galactose (Gal), galacturonic acid (GalA) of 1.6:1.0:1.2: 0.9; the molecular weight is 21.84 KDa; specific rotation degree
Figure BDA0001009626750000031
CD spectrum: 194nm (1.882), 230nm (0.993), 209nm (-2.158); the protein content is 0.64%; content of uronic acid: 19.8 percent; sugar content 92.3%; the main connection mode is as follows: 1, 4-linked mannose, galactose also predominates on 1, 4-and terminal linkages, glucose on 1,3,4, 6-and 1, 4-linkages, and in addition, contains small amounts of 1,3, 4-linked arabinose and terminal-linked xylose. Wherein 1, 4-linked mannose, 1, 4-linked galactose and 1,3,4, 6-linked glucose are the main components.
The roughhaired holly root polysaccharide (GDS1, GDS2 and GDS3) is prepared by the following method:
extracting radix Ilicis Asprellae with 95% ethanol under reflux, volatilizing ethanol from the residue, extracting the residue with hot water for 3 times, filtering, concentrating the water extract to appropriate volume, adding 4 times of anhydrous ethanol, filtering to obtain precipitate, redissolving with water, adding trichloroacetic acid to remove free protein, centrifuging, adjusting the supernatant to neutral with 1.0mol/L NaOH, concentrating to appropriate volume, dialyzing, concentrating the dialysate to appropriate volume, and freeze drying to obtain fluffy crude polysaccharide; (ii) a
Dissolving the crude polysaccharide in distilled water under magnetic stirring, centrifuging, and separating the supernatant in DEAE-52 chromatographic column. Eluting with 0.1, 0.2, 0.4, 0.8 and 1.6mol/L NaCl solution, collecting fractions, combining the same fractions according to sulfuric acid-phenol method, concentrating, and freeze drying to obtain 5 secondary components: d1, D2, D3, D4 and D5. Further separating D1 and D4 with significant anti-complement activity;
d1 and D4 were dissolved in distilled water, and the solution was separated by Sephacryl S-200 column (1.5 m.times.1.5 cm). Eluting with distilled water, and collecting each fraction. The same fractions were pooled, concentrated and lyophilized to obtain the polysaccharides GDS1, GDS2 and GDS3, based on the results of the sulfuric acid-phenol method for color development and anticomplementary activity assay.
The in vitro test proves that the prepared roughhaired holly root polysaccharides GDS1, GDS2 and GDS3 have obvious effect on cell hemolysis caused by activation of classical and alternative complement pathwaysHas significant inhibitory effect and has significant anticomplementary effect, wherein, CH50The values are respectively 0.134 +/-0.024 mg/mL, 0.187 +/-0.031 mg/mL and 0.098 +/-0.011 mg/mL; AP (Access Point)50The values were 0.297. + -. 0.037mg/mL, 0.312. + -. 0.033mg/mL and 0.195. + -. 0.030mg/mL, respectively.
The invention provides a new component with anticomplementary activity, in particular to roughhaired holly root polysaccharides GDS1, GDS2 and GDS3, which can be further used for preparing anticomplementary drugs.
Drawings
FIG. 1 is an HPGPC chromatogram of GDS1(A), GDS2(B), and GDS3(C), wherein,
a TSK-GEL GMPWXL GEL column (300X 7.6 mm); eluent: 0.1% NaCl; flow rate: 1.0mL/min, DAD210nm assay.
FIG. 2 is an HPCE chromatogram of GDS1(A), GDS2(B), and GDS2(C), wherein,
high efficiency capillary columns (60 cm. times.75 μm); buffer solution: 0.01mol/L boric acid-KOH buffer, pH 10.
FIG. 3 is a graph showing differences of GDS1(A), GDS2(B), and GDS3(C), wherein,
a TSK-GEL GMPWXL GEL column of 300X 7.6 mm; eluent: 0.1% NaCl; flow rate: 0.5mL/min, RID.
Detailed Description
EXAMPLE 1 preparation of the Roughhaired holly root polysaccharides GDS1, GDS2 and GDS3
Crushing 19Kg of roughhaired holly root medicinal material, performing reflux extraction for 3 times by using 95% ethanol, volatilizing ethanol in filter residue, leaching the filter residue for 3 times by using hot water, filtering, combining extracting solutions, concentrating to a proper volume, adding anhydrous ethanol with 4 times of volume, standing for about 3 hours, filtering to obtain a precipitate, redissolving the precipitate by using water, adding trichloroacetic acid to ensure that the concentration is 15% to remove free protein, centrifuging, adjusting the supernatant to be neutral by using 1.0mol/L NaOH, concentrating to a proper volume, dialyzing for 3 days by using flowing water by using a dialysis membrane with molecular weight cut-off of 3000, concentrating the dialyzate to a proper volume, and performing freeze drying to obtain about 15.8g of fluffy crude polysaccharide. Weighing 12.0g of crude polysaccharide, dissolving in distilled water, centrifuging, and separating the supernatant in DEAE-52 column. Eluting with 0.1, 0.2, 0.4, 0.8 and 1.6mol/L NaCl solution to obtain eluateThe volume was larger than 2 column volumes (about 0.5L), the flow rate was about 2mL/min, fractions were collected, and after color development with a sulfuric acid-phenol method, the absorbance value was measured at 490 nm. According to the results of sugar color reaction and ultraviolet detection, the same fractions are combined, concentrated to a certain volume, and then concentrated to a proper volume after dialysis, and then freeze-dried to obtain 5 secondary components: d1, D2, D3, D4 and D5. Activity tracking shows that D1 and D4 have better anticomplementary activity (CH of D1)50The value is 0.201 +/-0.041 mg/mL; CH of D450The value is 0.193. + -. 0.027 mg/mL);
d1(950mg) was dissolved in distilled water and separated by Sephacryl S-200 column (100X 1.6 cm). Eluting with distilled water as eluent, controlling flow rate at constant flow pump to be about 0.6mL/min, and collecting each fraction. After the color development of a sulfuric acid-phenol method separation tube, detecting an absorbance value at 490nm, detecting the hemolysis inhibition rate of the gel by a corresponding separation tube, merging the same fractions according to the detection result, concentrating to a proper volume, and freeze-drying to obtain homogeneous polysaccharides GDS1(130mg) and GDS2(110 mg);
d4(500mg) was completely dissolved in distilled water, and the solution was separated by Sephacryl S-200 column (100X 1.6 cm). Eluting with distilled water as eluent, controlling flow rate to about 0.6mL/min with constant flow pump, collecting absorbance value detected at 490nm after color development of sulfuric acid-phenol method separation tube by automatic fraction receiver, detecting hemolysis inhibition rate with corresponding separation tube, combining same fractions according to detection result, concentrating to appropriate volume, and freeze drying to obtain homogeneous polysaccharide GDS3(150 mg);
the GDS1, GDS2 and GDS3 were all homogeneous polysaccharides (as shown in FIGS. 1-3) detected by High Performance Gel Permeation Chromatography (HPGPC), High Performance Capillary Electrophoresis (HPCE) and differential detection method.
Example 2 structural characterization of the Roughhaired Holly root polysaccharides (GDS1, GDS2 and GDS3)
(1) Determination of molecular weight
The molecular weights of GDS1, GDS2 and GDS3, which are calculated by Wyatt eighteen-angle laser light scattering, are 92.77KDa, 16.22KDa and 21.84KDa respectively by adopting a TSK-GEL GMPWXL GEL column (300 multiplied by 7.6mm) and a mobile phase of 0.01mol/L NaCl, a flow rate of 0.8mL/min and a column temperature of 25 ℃.
(2) Results of elemental analysis
GDS1:C,40.30%;H,6.68%;N,0.56%。
GDS2:C,37.80%;H,5.88%;N,0.57%。
GDS3:C,29.91%;H,5.45%;N,0.67%。
(3) Specific rotation and CD spectrum
GDS1:
Figure BDA0001009626750000051
CD:202nm(-4.594);GDS2:
Figure BDA0001009626750000052
Figure BDA0001009626750000053
CD:201nm(-2.242),223nm(1.882);GDS3:
Figure BDA0001009626750000054
CD:194nm(1.882),230nm(0.993),209nm(-2.158)
(4) Total sugar, uronic acid and protein content determination
The total sugar content of GDS1 determined by sulfuric acid-phenol method is 95.8%; the total sugar content of GDS2 was 93.6%; GDS3 total sugar content 92.3%;
the uronic acid content of GDS1 determined by m-hydroxy biphenyl method is 8.8%; the uronic acid content of GDS2 was 15.5%; uronic acid content of GDS3 19.8%;
the protein content of GDS1 determined by Coomassie brilliant blue method is 0.64%; the protein content of GDS2 was 0.86%; the protein content of GDS3 was 1.54%;
(5) sugar composition analysis
Hydrolyzing GDS1, GDS2 and GDS3 with 2.0mol/L TFA at 100 deg.C for 6 hr, hydrolyzing to obtain product, adding appropriate amount of NaBH4Reducing, acetylating acetic anhydride to prepare an alditol acetate derivative, and carrying out gas phase composition analysis;
GDS1 is a heteropolysaccharide composed mainly of 7 monosaccharides, with xylose (Xyl), rhamnose (Rha), arabinose (Ara), mannose (Man), glucose (Glc), galactose (Gal), galacturonic acid (GalA) 1.0:2.7:8.8:3.1:2.8:24.8:3.4 on a molar ratio of sugar residues, and also a small amount of glucuronic acid;
GDS2 is a heteropolysaccharide composed mainly of 7 monosaccharides, with xylose (Xyl), rhamnose (Rha), arabinose (Ara), mannose (Man), glucose (Glc), galactose (Gal), galacturonic acid (GalA) 1.0:1.5:2.3:1.6:1.8:4.6:1.7 on a molar ratio of sugar residues, and also a small amount of glucuronic acid;
GDS3 is a heteropolysaccharide composed mainly of 4 monosaccharides with a molar ratio of mannose (Man) to glucose (Glc) to galactose (Gal) to galacturonic acid (GalA) of 1.6:1.0:1.2:0.9, and further containing small amounts of xylose and arabinose;
(6) methylation analysis
Methylating GDS1, GDS2 and GDS3, depolymerizing the methylated product with formic acid for 4h, hydrolyzing with 2mol/LTFA at 100 deg.C for 6h, and hydrolyzing with NaBH4Reducing and acetylating acetic anhydride to obtain partially methylated alditol acetate derivative, and performing GC-MS analysis;
the GDS1 is mainly connected by standard map judgment in the following ways: the main linkage modes of galactose are 1,3, 4-linkage, 1, 4-linkage and end linkage, and a small amount of 1,4, 6-linkage and 1, 3-linkage galactose; the primary mode of arabinose attachment is 1, 3-linked and end-linked arabinose; the mannose is linked in a 1, 4-linkage manner; the connection mode of glucose is 1,2,3, 4-connection and end connection, and the connection mode of rhamnose is 1, 3-connection and end connection; in addition, it also contains a small amount of 2, 3-linked xylose. Wherein the end-linked galactose, the 1,3, 4-linked galactose and the 1, 3-linked arabinose are the main components;
the GDS2 has the following main connection modes: the connection mode of galactose is 1,3, 4-connection, 1, 3-connection and end connection; the connection mode of arabinose is 1, 3-connection and end connection; mannose is linked in a 1, 4-linkage and end-linkage manner, and a small amount of 1, 3-linked mannose; the connection mode of glucose is 1,2,3, 4-connection, 1, 6-connection and end connection, and the connection mode of rhamnose is 1, 3-connection and end connection; in addition, it also contains 2, 3-linked xylose;
the GDS3 has the following main connection modes: 1, 4-linked mannose, galactose also predominates on 1, 4-and terminal linkages, glucose on 1,3,4, 6-and 1, 4-linkages, and in addition, contains small amounts of 1,3, 4-linked arabinose and terminal-linked xylose. Wherein 1, 4-linked mannose, 1, 4-linked galactose and 1,3,4, 6-linked glucose are the main components.
Example 3 classical pathway complement inhibition assay
0.1mL of complement (guinea pig serum) was taken, and Barbital Buffer (BBS) was added to prepare a 1:10 solution, which was diluted in duplicate with BBS to a 1:20, 1:40, 1:80, 1:160, 1:320, 1:640, and 1:1280 solution. Dissolving 1000 parts of hemolysin 1, complement with various concentrations and Sheep Red Blood Cell (SRBC) 2% 0.1mL respectively in 0.3mLBBS, mixing, placing in 37 deg.C water bath for 30min, placing in low temperature high speed centrifuge, and centrifuging at 5000rpm and 4 deg.C for 10 min. 0.2mL of the supernatant from each tube was placed in a 96-well plate, and the absorbance at 405nm was measured. The experiment was performed with a full hemolysis group (0.1mL of 2% SRBC in 0.5mL of triple distilled water). And (4) calculating the hemolysis rate by taking the absorbance of the three-distilled water-soluble blood vessel as a total hemolysis standard. Complement dilution was plotted on the X-axis and percent hemolysis was plotted on the Y-axis. The lowest complement concentration that achieves a similar high hemolysis rate is chosen as the critical complement concentration required to ensure that the system is hemolyzed properly. And (2) uniformly mixing the complement with critical concentration with each homogeneous polysaccharide sample, adding a proper amount of BBS, hemolysin and 2% SRBC, carrying out water bath at 37 ℃ for 30min, putting the mixture into a low-temperature high-speed centrifuge, centrifuging the mixture at 5000rpm and 4 ℃ for 10min, then respectively taking 0.2mL of supernatant of each tube, putting the supernatant into a 96-well plate, and measuring the absorbance at 405 nm. The test is carried out by setting a test sample control group, a complement group and a whole hemolysis group. And subtracting the absorbance value of the corresponding test sample control group from the absorbance value of the test sample, and calculating the hemolysis rate. The concentration of the test sample on the X-axis and the inhibition rate of hemolysis on the Y-axis were plotted to calculate the Concentration (CH) of the test sample required for 50% inhibition of hemolysis50) (as shown in table 1).
Example 4 in vitro anti-alternative complement pathway assay
0.2mL of complement (human serum) was taken, and added with AP diluent (barbital buffer, pH 7.4, containing 5mM Mg)2+8mM EGTA) was prepared as a 1:1 solution, diluted in duplicate to 1:2, 1:4, 1:8, 1:16, 1:32, 1:64 and 1:128 solutions. Taking 0.15mL of complement and 0.15mL of AP diluent at each concentration0.15mL of release solution and 0.20mL of 0.5% rabbit red blood cell (RE), mixing, placing in a low-temperature high-speed centrifuge after 30min of 37 ℃ water bath, and centrifuging for 10min at 5000rpm and 4 ℃. 0.20mL of the supernatant from each tube was placed in a 96-well plate, and the absorbance was measured at 405 nm. The experiment was performed with a full hemolysis group (0.20mL of 0.5% RE in 0.3mL of triple distilled water). And (4) calculating the hemolysis rate by taking the absorbance of the three-distilled water-soluble blood vessel as a total hemolysis standard. Complement dilution was plotted on the X-axis and percent hemolysis was plotted on the Y-axis. The lowest complement concentration that achieves a similar high hemolysis rate is chosen as the critical complement concentration required to ensure that the system is hemolyzed properly. The determined critical concentration of complement and each homogeneous polysaccharide sample is mixed, in 37 ℃ water bath 10min, adding 0.20 mL0.5% RE. Placing each tube in a 37 ℃ water bath for 30min, placing the tube in a low-temperature high-speed centrifuge, centrifuging the tube at 5000rpm and 4 ℃ for 10min, respectively taking 0.20mL of supernatant of each tube, placing the tube in a 96-well plate, and measuring the absorbance at 405 nm. The test is carried out by setting a test sample control group, a complement group and a whole hemolysis group. And subtracting the absorbance value of the corresponding test sample control group from the absorbance value of the test sample, and calculating the hemolysis rate. The concentration of the test sample on the X-axis and the inhibition rate of hemolysis on the Y-axis were plotted to calculate the concentration of the test sample required for 50% inhibition of hemolysis (AP)50) (as shown in table 1).
TABLE 1 complement Activity of three homogeneous polysaccharides of Roughhaired Holly root
Figure BDA0001009626750000081
CH50And AP50The values are expressed as: mean ± SD (n ═ 3).

Claims (5)

1. Use of radix Ilicis Asprellae polysaccharide GDS1, GDS2 or GDS3 in preparing complement inhibiting medicine; the polysaccharide of the roughhaired holly root is a polysaccharide of the roughhaired holly root, wherein,
the roughhaired holly root polysaccharide GDS1 has the structural characteristics that heteropolysaccharide mainly consists of 7 monosaccharides, the molar ratio of sugar residues is Xyl, Ara, Man, Glc, Gal, GalA, 1.0, 2.7, 8.8, 3.1, 2.8, 24.8 and 3.4, the molecular weight is 92.77KDa, and the specific rotation degree is [ α ]]D 25=-25.0(c 0.5,H2O), CD profile: 202nm (-4.594); protein content: 0.64 percent; furfuralAcid content: 8.8 percent; sugar content 95.8%;
the roughhaired holly root polysaccharide GDS2 has the structural characteristics that heteropolysaccharide mainly consists of 7 monosaccharides, the molar ratio of sugar residues is Xyl, Ara, Man, Glc, Gal, GalA, 1.0, 1.5, 2.3, 1.6, 1.8, 4.6, 1.7, the molecular weight is 16.22KDa, and the specific rotation degree is [ α ]]D 25=-37.0(c 0.5,H2O), CD profile: 201nm (-2.242), 223nm (1.882); protein content: 0.86 percent; content of uronic acid: 15.5 percent; sugar content 93.6%;
the roughhaired holly root polysaccharide GDS3 is structurally characterized in that heteropolysaccharide mainly comprises 4 monosaccharides, the molar ratio of sugar residues is Man to Glc to Gal to GalA to 1.6 to 1.0 to 1.2 to 0.9, the molecular weight is 21.84kDa, and the specific rotation degree is α]D 25=58(c 0.5,H2O), CD profile: 194nm (1.882), 230nm (0.993), 209nm (-2.158); protein content: 1.54 percent; content of uronic acid: 19.8 percent; the sugar content was 92.3%.
2. The use as claimed in claim 1, wherein said polysaccharide of roughhaired holly root, GDS1, GDS2 or GDS3, has a significant inhibitory effect on the hemolysis of cells induced by the activation of the classical and alternative complement pathways.
3. Use according to claim 2, characterized in that said roughhaired holly root polysaccharide GDS1 has CH50The value is 0.134. + -. 0.024mg/mL, AP50The value was 0.297. + -. 0.037 mg/mL.
4. Use according to claim 2, characterized in that said roughhaired holly root polysaccharide GDS2 has CH50The value was 0.187. + -. 0.031mg/mL, AP50The value was 0.312. + -. 0.033 mg/mL.
5. Use according to claim 2, characterized in that said roughhaired holly root polysaccharide GDS3 has CH50The value is 0.098 +/-0.011 mg/mL, AP50The value was 0.195. + -. 0.030 mg/mL.
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