CN114395003B - Glucoside derivative of epigallocatechin gallate or salt thereof, preparation method and application - Google Patents
Glucoside derivative of epigallocatechin gallate or salt thereof, preparation method and application Download PDFInfo
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- CN114395003B CN114395003B CN202210112686.8A CN202210112686A CN114395003B CN 114395003 B CN114395003 B CN 114395003B CN 202210112686 A CN202210112686 A CN 202210112686A CN 114395003 B CN114395003 B CN 114395003B
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
- C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
- C07H19/056—Triazole or tetrazole radicals
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P39/00—General protective or antinoxious agents
- A61P39/06—Free radical scavengers or antioxidants
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
Abstract
The invention provides a glucoside derivative of epigallocatechin gallate or a salt thereof, a preparation method and application thereof, and relates to the technical field of pharmacology. The invention solves the technical problems of low stability of epigallocatechin gallate (EGCG), low blood-brain barrier penetration rate, brain targeting and poor bioactivity of the EGCG, and achieves the technical effects of improving the stability, the blood-brain barrier penetration rate, the brain targeting and the bioactivity of the EGCG.
Description
Technical Field
The invention relates to the technical field of pharmacology, in particular to a glucoside derivative of epigallocatechin gallate or a salt thereof, a preparation method and application.
Background
Green tea is tender leaf of camellia plant tea, and its main effective components include green tea polyphenol, green tea polysaccharide and caffeine, in which the most important component is green tea polyphenol (green tea polyphenols, GTP), also called tea tannin and catechin, and is the total name of catechin, flavonoid, phenolic acid and flower colour compound in the tea, and its main effective component is 13% -30% of dry weight of tea and 2% -5% of fresh leaf.
The green tea polyphenol takes catechin as a main component, accounts for 60% -80% of the total phenol content, and the catechin mainly comprises the following substances:
epigallocatechin gallate (EGCG), which is the component with highest catechin content and accounts for 60% of catechin content, has the following structural formula:
epigallocatechin (EGC) has the structural formula shown below:
epicatechin gallate (ECG) having the structural formula:
gallocatechin (GC) having the following structural formula:
and Epicatechin (EC), the structural formula is shown below:
at present, a great deal of researches show that EGCG has the biological activities of resisting tumor, inflammation, oxidation, aging, obesity, ultraviolet, blood pressure and blood sugar, blood fat and heart and cerebral vascular system diseases, and regulating endocrine and immune systems. In particular, the neuroprotective effects of EGCG are becoming increasingly important. 6-hydroxydopamine is dopaminergic neurotoxin, and in vitro experimental research shows that EGCG can resist 6-hydroxydopamine induced death of rat PC12 cells (rat adrenal pheochromocytoma cells as nerve cell model) and human glioma cells SH-SY5Y cells.
However, limitations of EGCG molecular structure and physicochemical properties thereof limit its efficacy and application, e.g., EGCG has good water solubility and poor lipid solubility, resulting in low blood brain barrier penetration when applied to neuroprotection; meanwhile, EGCG is extremely unstable, so that the EGCG is easily oxidized into quinone compounds harmful to human bodies in a solution, and the stability of the EGCG is seriously affected by environmental factors such as temperature, oxidants, neutral or alkaline environments. Therefore, in order to improve the stability, blood brain barrier penetration, brain targeting and bioactivity of EGCG, reasonable chemical modification of the molecular structure of EGCG is required.
In view of this, the present invention has been made.
Disclosure of Invention
One of the purposes of the invention is to provide a glucoside derivative of epigallocatechin gallate or a salt thereof, which has higher stability, high blood brain barrier penetration rate, better brain targeting and bioactivity.
The second object of the present invention is to provide a method for producing a glucoside derivative of epigallocatechin gallate or a salt thereof.
The invention further aims to provide an application of the glucoside derivative of epigallocatechin gallate or the salt thereof in preparing brain cell antioxidant drugs.
In order to achieve the above object of the present invention, the following technical solutions are specifically adopted:
in a first aspect, the present invention provides a glucoside derivative of epigallocatechin gallate or a salt thereof, comprising a structure represented by general formula (1):
wherein a is selected from integers from 0 to 2;
b and c are independently selected from integers from 0 to 3.
Further, the glucoside derivative of epigallocatechin gallate or a salt thereof includes a structure represented by the general formula (2):
wherein a is selected from integers from 0 to 1;
b and c are independently selected from integers from 0 to 3.
Further, the glucoside derivative of epigallocatechin gallate or a salt thereof includes a structure represented by the general formula (3):
wherein a, b and c are independently selected from integers from 0 to 2.
Further, the glucoside derivative of epigallocatechin gallate or a salt thereof includes a structure represented by the general formula (4):
wherein a, b and c are independently selected from integers from 0 to 2.
Further, the glucoside derivative of epigallocatechin gallate or the salt thereof is any one of the following structures:
and
in a second aspect, the present invention provides a method for preparing a glucoside derivative of epigallocatechin gallate or a salt thereof, comprising the steps of:
performing click reaction on 6-azido-6-deoxyglucose and terminal acetylized epigallocatechin gallate to obtain a glucoside derivative of the epigallocatechin gallate or a salt thereof;
wherein the terminal acetylized epigallocatechin gallate comprises a structure represented by the general formula (5):
wherein e is selected from integers from 0 to 2;
m and n are independently selected from integers from 0 to 3.
Further, the click reaction temperature is 10-45 ℃, and the reaction time is 1-24 hours;
further preferably, the preparation method of the 6-azido-6-deoxyglucose comprises the following steps:
the hydroxyl on the 6-position of glucose is substituted by azido to obtain the 6-azido-6-deoxyglucose;
further preferably, the preparation method of the 6-azido-6-deoxyglucose comprises the following steps:
the hydroxyl on the 6-position of glucose is substituted with p-toluenesulfonyl chloride, and then the p-toluenesulfonate group is substituted by azido, so that the 6-azido-6-deoxyglucose is obtained.
Further, the preparation method of the terminal acetylized epigallocatechin gallate comprises the following steps:
and substituting hydroxyl groups on the epigallocatechin gallate with propynyl groups to obtain the terminal-alkynylated epigallocatechin gallate.
Further, the terminal-alkynylated epigallocatechin gallate includes at least one of 5-propynylepigallocatechin gallate, 4',4 "-dipropynylepigallocatechin gallate and 5,4',4" -tripropynylepigallocatechin gallate.
In a third aspect, the invention provides an application of a glucoside derivative of epigallocatechin gallate or a salt thereof in preparing a brain cell antioxidant medicament.
Compared with the prior art, the invention has at least the following beneficial effects:
according to the glucoside derivative or the salt of the epigallocatechin gallate, which is provided by the invention, the 6-position covalent bonding of catechin and glucosyl is modified, and the 6-position of glucose extends outside the six-membered ring molecular structure, so that the steric hindrance is minimum, and the recognition and bonding of glucosyl and GLUT1 are very facilitated; meanwhile, compared with catechin, the modified catechin provided by the invention has the advantages of lower cytotoxicity, wider neuroprotection threshold, higher blood brain barrier penetration rate and better oxidation protection activity on brain cells, and can improve the stress capability of nerve cells on oxidation injury and reduce the possibility of the nerve cells on oxidation injury.
The preparation method of the glucoside derivative or the salt of the epigallocatechin gallate provided by the invention has the advantages of simple process and convenient operation.
The application of the glucoside derivative of the gallocatechin gallate or the salt thereof in preparing the brain cell antioxidant medicament can improve the stability, the blood brain barrier penetration rate, the brain targeting and the bioactivity of the medicament.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are needed in the description of the embodiments or the prior art will be briefly described, and it is obvious that the drawings in the description below are some embodiments of the present invention, and other drawings can be obtained according to the drawings without inventive effort for a person skilled in the art.
FIG. 1 is a reaction scheme for preparing end-alkynylated epigallocatechin gallate provided in example 4 of the present invention;
FIG. 2 is a reaction scheme for preparing 6-azido-6-deoxyglucose provided in example 4 of the present invention;
FIG. 3 is a reaction scheme for preparing glu-EGCG according to example 4 of the invention;
FIG. 4 is a reaction scheme for preparing 2glu-EGCG according to example 4 of the invention;
FIG. 5 is a reaction scheme for preparing 3glu-EGCG according to example 4 of the invention;
FIG. 6 is a graph showing the effect of the glucoside derivative of EGCG on the viability of PC12 cells and bEnd.3 cells, obtained in the test example of the invention;
FIG. 7 shows the resistance of the glucoside derivative of EGCG obtained in the test example of the invention to H by PC12 cells 2 O 2 Oxidized cell viability effect graph;
FIG. 8 shows H-reduction of PC12 cells by the glucoside derivative of EGCG obtained in the test example of the invention 2 O 2 Flow data graph of intracellular Reactive Oxygen Species (ROS) capacity generated by oxidative damage.
Detailed Description
The technical solutions of the present invention will be clearly and completely described in connection with the embodiments, and it is apparent that the described embodiments are some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The invention is further illustrated by the following examples. The materials in the examples were prepared according to the existing methods or were directly commercially available unless otherwise specified.
According to a first aspect of the present invention, there is provided a glucoside derivative of epigallocatechin gallate or a salt thereof, comprising a structure represented by general formula (1):
wherein a is selected from integers from 0 to 2, and a is 0, 1 or 2;
b and c are independently selected from integers from 0 to 3, b is 0, 1,2 or 3, c is 0, 1,2 or 3.
According to the glucoside derivative or the salt of the epigallocatechin gallate, which is provided by the invention, the 6-position covalent bonding of catechin and glucosyl is modified, and the 6-position of glucose extends outside the six-membered ring molecular structure, so that the steric hindrance is minimum, and the recognition and bonding of glucosyl and GLUT1 are very facilitated; meanwhile, compared with catechin, the modified catechin provided by the invention has the advantages of lower cytotoxicity, wider neuroprotection threshold, higher blood brain barrier penetration rate and better oxidation protection activity on brain cells, and can improve the stress capability of nerve cells on oxidation injury and reduce the possibility of the nerve cells on oxidation injury.
In a preferred embodiment, the glucoside derivative of epigallocatechin gallate or the salt thereof of the present invention includes a structure represented by the general formula (2):
wherein a is selected from integers from 0 to 1, and a is 0 or 1;
b and c are independently selected from integers from 0 to 3, b is 0, 1,2 or 3, c is 0, 1,2 or 3.
In a preferred embodiment, the glucoside derivative of epigallocatechin gallate or the salt thereof of the present invention includes a structure represented by general formula (3):
wherein a, b and c are independently selected from integers from 0 to 2, a being 0, 1 or 2; b is 0, 1 or 2; c is 0, 1 or 2.
In a preferred embodiment, the glucoside derivative of epigallocatechin gallate or the salt thereof of the present invention includes a structure represented by the general formula (4):
wherein a, b and c are independently selected from integers from 0 to 2, a being 0, 1 or 2; b is 0, 1 or 2; c is 0, 1 or 2.
The glucoside derivative of epigallocatechin gallate or the salt thereof is any one of the following structures:
/>
and
the catechin of the invention is modified by covalent bonding with the 6-position of the glucosyl group, so that the penetration rate of the catechin to the blood brain barrier and the antioxidant protective activity to brain cells can be improved, the modified catechin has lower cytotoxicity and wider nerve protection threshold value than the protocatechuic, the stress capability of the nerve cells for resisting oxidative damage can be improved, and the possibility of the nerve cells for being damaged by oxidation can be reduced.
According to a second aspect of the present invention, there is provided a method for producing a glucoside derivative of epigallocatechin gallate or a salt thereof, comprising the steps of:
and (3) performing click reaction on the 6-azido-6-deoxyglucose and the terminal acetylized epigallocatechin gallate to obtain a glucoside derivative of the epigallocatechin gallate or a salt thereof.
Wherein the terminal-alkynylated epigallocatechin gallate comprises a structure represented by the general formula (5):
wherein e is selected from integers from 0 to 2, and e is 0, 1 or 2;
m and n are independently selected from integers from 0 to 3, m is 0, 1,2 or 3, n is 0, 1,2 or 3.
In the present invention, the click reaction temperature is 10-45deg.C, which is typically but not limited to, for example, 10deg.C, 15deg.C, 20deg.C, 25deg.C, 30deg.C, 35deg.C, 40deg.C, 45deg.C; the reaction time is 1-24 hours, typical but non-limiting reaction times being, for example, 1h, 2h, 4h, 5h, 10h, 15h, 24h.
The click reaction temperature and time defined by the invention can obtain better reaction effect and higher product yield and purity.
In the invention, the 6-azido-6-deoxyglucose can be a commercial product or can be prepared by the following preparation method: the preparation method of the 6-azido-6-deoxyglucose comprises the following steps:
the hydroxyl on the 6-position of glucose is substituted by azido to obtain 6-azido-6-deoxyglucose, and the structural formula is as follows:
in a preferred embodiment, the method for preparing 6-azido-6-deoxyglucose comprises the steps of:
the hydroxyl on the 6-position of glucose is substituted with p-toluenesulfonyl chloride, and then the p-toluenesulfonate group is substituted by azido, so that the 6-azido-6-deoxyglucose is obtained.
The preparation method provided by the invention can better obtain the 6-azido-6-deoxyglucose.
In the invention, the end-alkynylated epigallocatechin gallate can be a commercial product or can be prepared by the following preparation method: and substituting hydroxyl on epigallocatechin gallate with propynyl to obtain the terminal acetylized epigallocatechin gallate.
In a preferred embodiment, the terminal-alkynylated epigallocatechin gallate of the present invention includes at least one of 5-propynylepigallocatechin gallate, 4',4 "-dipropynylepigallocatechin gallate, and 5,4',4" -tripropynylepigallocatechin gallate.
The structural formula of the 5-propyne epigallocatechin gallate is as follows:
the structural formula of the 4', 4' -dipropargininyl epigallocatechin gallate is as follows:
the structural formula of the 5,4' -tripropyleepigallocatechin gallate is as follows:
the terminal acetylized epigallocatechin gallate of the invention can also be selected from any one of the following structural formulas:
and +.>
According to the third aspect of the invention, the application of the glucoside derivative of epigallocatechin gallate or the salt thereof in preparing brain cell antioxidant medicines is provided, the technical problems of low stability, low blood brain barrier penetration rate, brain targeting and poor bioactivity of the epigallocatechin gallate (EGCG) are solved, and the technical effects of improving the stability, blood brain barrier penetration rate, brain targeting and bioactivity of the EGCG are achieved.
The invention is further illustrated by the following examples. The materials in the examples were prepared according to the existing methods or were directly commercially available unless otherwise specified.
Example 1
A glucoside derivative of epigallocatechin gallate (EGCG), 5- (1, 2,3, triazole-6-deoxyglucose) epigallocatechin gallate, abbreviated as glu-EGCG, has the following molecular structural formula, and is not reported in the literature:
example 2
A glucoside derivative of epigallocatechin gallate (EGCG), 4' - (1, 2,3, triazole-6-deoxyglucose) epigallocatechin gallate, abbreviated as 2glu-EGCG, has the molecular structural formula as follows, and is not reported in the literature:
example 3
A glucoside derivative of epigallocatechin gallate (EGCG), 5,4' - (1, 2,3, triazole-6-deoxyglucose) epigallocatechin gallate, abbreviated as 3glu-EGCG, has the molecular structural formula as follows, and is not reported in the literature:
example 4
This example is a process for preparing a glucoside derivative of EGCG of example 1, example 2 and example 3, the reaction routes of which are shown in fig. 1, fig. 2, fig. 3, fig. 4 and fig. 5, comprising the steps of:
(1) Taking EGCG as a starting material, and selectively attaching propynyl to the 5-hydroxyl of EGCG by nucleophilic substitution to obtain 5-propynylepigallocatechin gallate (compound 1 in figure 1); or, taking EGCG as a starting material, and selectively connecting propynyl to the 4 'and 4' hydroxyl of EGCG through nucleophilic substitution to obtain 4', 4' -dipropylenic epigallocatechin gallate (compound 2 in figure 1); or, using EGCG as starting material to replace hydroxyl at 5 and 4', 4' positions of EGCG to obtain 5,4' -tripropyleepigallocatechin gallate (compound 3 in figure 1);
the specific reaction comprises the following steps:
synthesis of 5-propyne epigallocatechin gallate (Compound 1 in FIG. 1):
EGCG (1.4 g,3 mmol) was dissolved in 15mL dry DMF, under nitrogen protection, 60% NaH (180 mg,4.5 mmol) was added under ice bath, the reaction was stirred at temperature for 1h, 3-bromopropyne (0.24 mL,3 mmol) was slowly added under ice bath, the temperature was raised to 90℃and then the reaction was carried out overnight; TLC monitoring reaction, after the reaction, the reaction solution is decompressed and concentrated to obtain a crude product, and silica gel column chromatography is used for purification (dichloromethane/methanol) to obtain 459.12mg of light pink powder with the yield of 30.82%;
synthesis of 4', 4' -Epigallocatechin gallate (Compound 2 in FIG. 1):
EGCG (2.8 g,6 mmol) was dissolved in 25mL dry DMF, nitrogen protected, stirred at 0deg.C, then 60% NaH (480 mg,12 mmol) was added, the reaction was carried out for 1h with stirring at a maintained temperature, then 3-bromopropyne (0.96 mL,12 mmol) was slowly added at 0deg.C, the temperature was raised to 90deg.C and then the reaction was carried out overnight; TLC monitoring reaction, after the reaction, the reaction solution is decompressed and concentrated to obtain a crude product, and silica gel column chromatography is used for purification (dichloromethane/methanol) to obtain 506.30mg of light pink powder with the yield of 15.78%;
synthesis of 5,4',4 "-tripropyleepigallocatechin gallate (compound 3 in fig. 1):
EGCG (5.6 g,12 mmol) was dissolved in 45mL dry DMF, under nitrogen protection, 60% NaH (1.44 g,36 mmol) was added under ice bath, the reaction was carried out for 1h with stirring at the temperature, 3-bromopropyne (2.88 mL,36 mmol) was slowly added under ice bath, and after raising the temperature to 90℃the reaction was carried out overnight; TLC monitoring reaction, after the reaction, the reaction solution is decompressed and concentrated to obtain a crude product, and silica gel column chromatography is used for purification (dichloromethane/methanol) to obtain 708.8mg of light pink powder, and the yield is 10.32%;
(2) Glucose is taken as a raw material, p-toluenesulfonyl chloride is used for replacing hydroxyl at the 6-position of glucose to obtain glucose with p-toluenesulfonate (a compound 4 in fig. 2), and azide is used for replacing the p-toluenesulfonate to obtain 6-azide-6-deoxyglucose (a compound 5 in fig. 2);
the specific reaction comprises the following steps:
synthesis of 6-O-p-toluenesulfonyl-glucose (Compound 4 in FIG. 2):
glucose (2.00 g,11.10 mmol) was dissolved in 40mL of dry pyridine, nitrogen protected, and stirred at 0deg.C to give a reaction solution; then p-toluenesulfonyl chloride (2.54 g,13.32 mmol) is dissolved in 10mL of dry pyridine, the mixture is added dropwise into the reaction liquid at the temperature of 0 ℃, and after the dropwise addition, the reaction liquid is moved to room temperature for reaction for 18h, and the color of the reaction liquid is changed from colorless to pale yellow; TLC monitoring reaction, after the reaction is completed, the reaction solution is decompressed and concentrated to obtain a crude product, and silica gel column chromatography is used for purification (dichloromethane/methanol) to obtain 1.62g of white oily solid with the yield of 43.65%;
synthesis of 6-azido-6 deoxyglucose (Compound 5 in FIG. 2):
compound 4 (1.50 g,4.50 mol) was dissolved in 20mL of DMF to obtain a reaction solution, naN was added 3 (1.46 g,22.5 mmol) was added to the reaction mixture, and the mixture was heated to 50℃for two days; TLC monitoring reaction, after reaction is completed, the reaction solution is filtered by diatomite to remove NaN which is not completely reacted 3 Concentrating under reduced pressure to obtain crude product, purifying by silica gel column chromatography (dichloromethane/methanol) to obtain 376.33mg of white powdery solid with 40.76% yield;
(3) The compound 5 and the compound 1 are subjected to click reaction to obtain a molecular glucose modified EGCG compound glu-EGCG (EGCG glucoside derivative of example 1), and the specific reaction is shown in figure 3 and comprises the following operation steps:
compound 5 (82.07 mg,0.4 mmol) and Compound 1 (198.57 mg,0.4 mmol) were dissolved in 4mL DMF, sodium ascorbate (47.55 mg,0.24 mmol) was added slowly to the reaction mixture with stirring, and cuprous iodide (25.71 mg,0.135 mmol) was added rapidly to the reaction mixture for 3h at room temperature; TLC monitoring reaction, after reaction completion, vacuum concentration gave crude product, silica gel column chromatography purification (ethyl acetate/methanol) to give slightly pink white powdery solid, which was purified by preparative high performance liquid chromatograph (Agilent Technologies 1260 info II) to give 58.72mg of slightly yellowish powdery solid product, yield 20.92%, characterized as follows:
1 H NMR(CD 3 OD,400MHz)δ7.97(d,1H,J＝20.4Hz,CH-N),6.92(s,2H,C 2 ″-H,C 6 ″-H),6.50(s,2H,C 2 ′-H,C 6 ′-H),5.96(s,2H,C 6 -H,C 8 -H),5.54(brs,1H,C 2 -H),5.19(s,2H,OCH 2 ),5.06(d,1H,J＝3.6Hz,C 1 ″′-H),4.97(s,1H,C 3 -H),4.45-4.44(m,2H),4.10-4.06(m,1H),3.58-3.69(m,1H),3.17-3.04(m,2H),3.02-2.96(m,1H,C 4 -H a ),2.87-2.83(m,1H,C 4 -H b )；
13 C-NMR(CD 3 OD,125MHz)δ165.7(C＝O),156.5(C-5),156.4(C-7),155.8(C-9),150.4(C-3″),150.4(C-5″),145.3(C-3′),145.3(C-5′),137.8(C-4″),132.4(C-4′),129.4(C-1′),128.4(C-N),125.7(C-1″),125.6(C＝N),109.0(C-2″),109.0(C-6″),105.4(C-2′),105.4(C-6′),97.9(C-10),96.8(C-6),95.2(C-8),94.5(C-1″′),77.1(C-2),74.6,74.4,73.3,71.2,69.0(C-3),64.3(OCH 2 ),51.1(C-6″′),25.4(C-4)；
the mass spectrum is as follows: HR-MS (ESI) m/z calculated for C 31 H 31 N 3 O 16 [M-H] - 700.17,found 700.15；
(4) The compound 5 and the compound 2 are subjected to click reaction to obtain two molecules of glucose modified EGCG compound 2glu-EGCG (EGCG glucoside derivative of example 2), and the specific reaction is operated as follows in the following steps of FIG. 4:
compound 5 (184.65 mg,0.9 mmol) and Compound 2 (240.51 mg,0.45 mmol) were dissolved in 9mL DMF, sodium ascorbate (53.49 mg,0.27 mmol) was added slowly to the reaction with stirring, and cuprous iodide (22.85 mg,0.12 mmol) was added rapidly to the reaction solution and reacted at room temperature for 4h; TLC monitoring reaction, after reaction completion, the reaction solution was concentrated under reduced pressure to obtain a crude product, and silica gel column chromatography was performed (ethyl acetate/methanol) to obtain a slightly pink white powdery solid, which was further performed with a preparative high performance liquid chromatograph (Agilent Technologies 1260Infinity II) to obtain 127.72mg of a slightly yellowish powdery solid product, yield 30.08%, characterized as follows:
1 H NMR(CD 3 OD，400MHz)δ7.94(dd,2H,J＝27.8,32.8Hz,CH-N),6.91(s,2H,C 2 ″-H,C 6 ″-H),6.53(s,2H,C 2 ′-H,C 6 ′-H),5.97(d,2H,J＝1.2Hz,C 6 -H,C 8 -H),5.56(brs,1H,C 2 -H),5.49(s,1H),5.19(s,2H,OCH 2 ),5.11(s,2H,OCH 2 ),5.07(dd,2H,J＝3.6Hz,C 1 ″′-H),5.01(s,1H,C 3 -H),4.56-4.42(m,4H),4.07-4.06(m,1H),3.58-3.69(m,2H),3.17-3.02(m,4H),2.99-2.84(m,2H,C 4 -H)；
13 C-NMR(CD 3 OD,125MHz)δ165.6(C＝O),156.5(C-5),156.5(C-7),155.6(C-9),150.4(C-3″),150.4(C-5″),150.2(C-3′),150.2(C-5′),143.7(C-4″),137.8(C-4′),134.9(C-1′),132.8(C-1″),125.6(C-N),125.5(C-N),125.5(C＝N),125.4(C＝N),109.0(C-2″),109.0(C-6″),105.7(C-2′),105.7(C-6′),97.9(C-10),96.8(C-6),95.2(C-8),94.5(C-1″′),92.6(C-1″′′),76.8(C-2),76.3,74.6,74.3,73.3,72.1,71.5,71.2,69.8,68.8(C-3),64.4(OCH 2 ),64.3(OCH 2 ),53.4(C-6″′),51.14(C-6″′′),25.3(C-4)；
the mass spectrum is as follows: HR-MS (ESI) m/z calculated for C 40 H 44 N 6 O 21 [M-H] - 943.26,found 943.30.
(5) The compound 5 and the compound 3 are subjected to click reaction to obtain a trimolecular glucose modified EGCG compound 3glu-EGCG (EGCG glucoside derivative of example 3), and the specific reaction has the following operation steps in FIG. 5:
compound 5 (277 mg,1.35 mmol) and Compound 3 (257.57 mg,0.45 mmol) were dissolved in 15mL DMF, sodium ascorbate (53.49 mg,0.27 mmol) was added slowly to the reaction with stirring, and cuprous iodide (38 mg,0.2 mmol) was added rapidly to the reaction solution and reacted at room temperature for 6h; TLC monitoring reaction, after reaction completion, the reaction solution was concentrated under reduced pressure to obtain a crude product, and silica gel column chromatography was performed (ethyl acetate/methanol) to obtain an off-white powdery solid, which was further purified by a preparative high performance liquid chromatograph (Agilent Technologies 1260 informativity II) to obtain 160.25mg of an off-yellow powdery solid product, yield 30%, mass spectrum of the product was as follows:
HR-MS(ESI):m/z calculated for C 49 H 57 N 9 O 26 [M-H] - 1187.12,found 1187.32.
comparative example 1
EGCG has the structure shown below:
comparative example 2
The glycoside derivative of EGCG glucose disclosed in prior art CN108467418A is modified by the 1-site linkage of EGCG to glucose.
The connection mode of the 1-position of glucose in CN108467418A can influence the affinity of glucose and GLUT1, but the 6-position of glucose extends outside a six-membered ring of a molecular structure in examples 1,2 and 3 of the invention is modified by covalent bonding with the 6-position of a glucose group, so that the steric hindrance is minimum, and the recognition and the bonding of the glucose group and the GLUT1 are very favorable.
Test examples
The glu-EGCG of example 1 and the 2glu-EGCG of example 2 were tested for cytotoxicity on PC12 cells and bEnd.3 cells and their resistance to PC2 cells H using PC12 cells (rat adrenal pheochromocytoma cells) as a neural cell model and bEnd.3 cells (mouse brain microvascular endothelial cells) as a blood brain barrier model 2 O 2 Oxidative damage ability EGCG of comparative example 1 was used as a control experiment.
(1) Cytotoxicity examination:
the test was performed by the conventional procedure using PC12 cells and bEnd.3 cells. The results showed that EGCG showed no significant decrease in cell viability for PC12 cells and bEnd.3 cells in the concentration range of 2.5. Mu.M-20. Mu.M (indicating no significant toxicity), but the glucoside derivatives of EGCG of example 1 and example 2 showed no significant decrease in cell viability in the broader concentration range of 2.5. Mu.M-320. Mu.M, and the results showed that the glucoside derivatives of EGCG of example 1 and example 2 were both less cytotoxic than EGCG, see FIG. 6.
(2) Test for ability to resist oxidative damage to nerve cells
The test was performed using PC12 cells according to the conventional procedure. The test group had physical mixtures of EGCG, EGCG+glucose (1:1 and 1:2 molar ratio) of comparative example 1, and glu-EGCG of example 1 and 2glu-EGCG of example 2, the results are shown in FIG. 7;
when the concentration increased to 20. Mu.M, each group began to promote anti-H in the cells 2 O 2 The effect is improved, and the antioxidant stress capability is enhanced along with the increase of the concentration; physical mixing of glucose with EGCG has no effect on EGCG antioxidant capacity; however, the antioxidant capacity of two EGCG analogues (glu-EGCG and 2 glu-EGCG) is obviously stronger than that of EGCG, and the antioxidant stress capacity of 2glu-EGCG is stronger than that of glu-EGCG, and the two EGCG analogues have difference significance at the concentration of 60 mu M; the results in FIG. 7 show that there is a significant difference in the concentration of glu-EGCG at 10. Mu.M, and that 20-60. Mu.M is the effective concentration range for the significant difference; the difference of 60 mu M is most obvious, and the 2glu-EGCG has the strongest antioxidant stress capability; when the sample concentration is higher than 60. Mu.M, there is no significant difference.
(3) ROS index evaluation of neuronal cell resistance to oxidative damage
The test was performed using PC12 cells according to the conventional procedure. Test group had blank, H 2 O 2 Physical mixtures of EGCG and EGCG+glucose (1:1 and 1:2 molar ratio), and glu-EGCG from example 1 and 2glu-EGCG from example 2, see FIG. 8;
the results showed that EGCG, as well as glu-EGCG of example 1 and 2glu-EGCG of example 2, increased in antioxidant stress capacity and decreased in H with increasing concentration 2 O 2 Intracellular Reactive Oxygen Species (ROS) produced by oxidative damage become more potent; meanwhile, glu-EGCG and 2glu-EGCG have obviously better H reduction than EGCG 2 O 2 Intracellular ROS capacity produced by oxidative damage, and 2glu-EGCG reduces H 2 O 2 Oxidative damage produces intracellular ROS with greater capacity than glu-EGCG.
In summary, the glucoside derivatives of EGCG of examples 1 and 2 of the present invention can improve stability, blood brain barrier penetration, brain targeting, and bioactivity of EGCG.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some or all of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit of the invention.
Claims (12)
1. A glucoside derivative of epigallocatechin gallate or a salt thereof, characterized by having a structure represented by the general formula (1):
wherein a is selected from integers from 0 to 2;
b and c are independently selected from integers from 0 to 3.
2. The glucoside derivative of epigallocatechin gallate or a salt thereof according to claim 1, wherein the glucoside derivative of epigallocatechin gallate or a salt thereof has a structure represented by general formula (2):
wherein a is selected from integers from 0 to 1;
b and c are independently selected from integers from 0 to 3.
3. The glucoside derivative of epigallocatechin gallate or a salt thereof according to claim 1, wherein the glucoside derivative of epigallocatechin gallate or a salt thereof has a structure represented by general formula (3):
wherein a, b and c are independently selected from integers from 0 to 2.
4. The glucoside derivative of epigallocatechin gallate or a salt thereof according to claim 1, wherein the glucoside derivative of epigallocatechin gallate or a salt thereof has a structure represented by general formula (4):
wherein a, b and c are independently selected from integers from 0 to 2.
5. The glucoside derivative of epigallocatechin gallate or a salt thereof according to any one of claims 1 to 4, wherein the glucoside derivative of epigallocatechin gallate or a salt thereof is any one of the following structures:
and
6. a method for producing the glucoside derivative of epigallocatechin gallate or the salt thereof according to any one of claims 1 to 5, comprising the steps of:
performing click reaction on 6-azido-6-deoxyglucose and terminal acetylized epigallocatechin gallate to obtain a glucoside derivative of the epigallocatechin gallate or a salt thereof;
wherein the terminal acetylized epigallocatechin gallate has a structure represented by the general formula (5):
wherein e is selected from integers from 0 to 2;
m and n are independently selected from integers from 0 to 3.
7. The method of claim 6, wherein the click reaction is performed at a temperature of 10-45 ℃ for a time of 1-24 hours.
8. The preparation method of the 6-azido-6-deoxyglucose according to claim 6, which comprises the following steps:
the hydroxyl group on the 6-position of glucose is substituted by azido, so that the 6-azido-6-deoxyglucose is obtained.
9. The preparation method of the 6-azido-6-deoxyglucose according to claim 6, which comprises the following steps:
the hydroxyl on the 6-position of glucose is substituted with p-toluenesulfonyl chloride, and then the p-toluenesulfonate group is substituted by azido, so that the 6-azido-6-deoxyglucose is obtained.
10. The method of claim 6, wherein the method of preparing the terminal-alkynylated epigallocatechin gallate comprises the steps of:
and substituting hydroxyl groups on the epigallocatechin gallate with propynyl groups to obtain the terminal-alkynylated epigallocatechin gallate.
11. The method according to claim 10, wherein the terminal-alkynylated epigallocatechin gallate is at least one of 5-propynylepigallocatechin gallate, 4',4 "-dipropynylepigallocatechin gallate and 5,4',4" -tripropynylepigallocatechin gallate.
12. Use of a glucoside derivative of epigallocatechin gallate or a salt thereof according to any one of claims 1-5 in preparing a brain cell antioxidant medicament.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108467418A (en) * | 2018-01-31 | 2018-08-31 | 云南农业大学 | Epigallo-catechin gallate (EGCG) glycosides derivatives and its application |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108467418A (en) * | 2018-01-31 | 2018-08-31 | 云南农业大学 | Epigallo-catechin gallate (EGCG) glycosides derivatives and its application |
Non-Patent Citations (4)
| Title |
|---|
| A short route for the synthesis of "sweet" macrocycles via a click-dimerization-ring-closing metathesis approach;Doerner, Simon，等;《Chemical Communications 》;第22卷;2852-2854及Supporting Information * |
| PREPARATION AND CHARACTERIZATION OF THE 6-ALDEHYDO DERIVATIVES OF AMYLOSE AND WHOLE STARCH;D. M. CLODE，等;《CARBOHYDRATE RESEARCH》;第第17卷卷;365-373 * |
| Sonochemistry: A Powerful Way of Enhancing the Efficiency of Carbohydrate Synthesis;Shenglou Deng，等;《J. Org. Chem.》;第71卷;5179 -5185及Supporting Information * |
| 表没食子儿茶素没食子酸酯（EGCG）衍生物的合成及生物活性研究;张盼;《吉林大学硕士学位论文》;全文 * |
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