Cinnamic acid derivative, preparation method therefor and use thereof

Abstract

The present invention belongs to the technical field of medicine, and disclosed are a cinnamic acid derivative, a preparation method therefor, and a use thereof. The structural formula of the cinnamic acid derivative is as shown in Formula 1A or Formula 1B, the number of substituents R being 0 to 5; Rs at different substitution sites are independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, ester, carboxyl, hydroxyl and allyloxy. The present invention, by means of improving the chemical structure of a cinnamic acid derivative and a preparation method therefor, designs a novel cinnamic acid derivative, which can be used to prepare a drug, for example, to have a therapeutic effect by means of inhibiting the MAPK signaling pathway, to exert a therapeutic effect by means of inhibiting ferroptosis, or to exert a detoxifying effect by means of activating UDP-glucuronosyltransferase, and in particular can be used to treat liver damage.

Description

A cinnamic acid derivative and its preparation method and application

Technical field

[0001] The present invention belongs to the field of medical technology, and more specifically relates to a cinnamic acid derivative and a preparation method and application thereof. [Background Technology]

[0002] Cinnamon bark is a traditional Chinese medicinal herb, consistently listed as a top-grade herb in ancient compendiums of materia medica. According to the 2015 edition of the Chinese Pharmacopoeia, the National Compilation of Traditional Chinese Medicine Standards, and the Ministry of Health Drug Standards, there are 565 Chinese patent medicines using cinnamon as a medicinal ingredient. Cinnamic acid, one of the main active monomeric components of cinnamon, has been shown to possess a variety of physiological activities, including antioxidant, anticancer, anti-inflammatory, antibacterial, antiallergic, and antithrombotic properties. Its effects on acute liver injury have also been reported in limited numbers. Cinnamic acid is also a very important intermediate in fine organic synthesis, and its derivatives have a wide range of applications in food, pharmaceuticals, and daily chemicals. They can be used to synthesize important drugs such as those for treating coronary heart disease, bone relaxants, local anesthetics, antiseptics, and hemostatics. Furthermore, many cinnamic acid derivatives, particularly those containing phenolic hydroxyl groups, are recognized antioxidants with strong free radical scavenging properties. Muhammad et al. reported that the cinnamic acid derivative 2,4-dihydroxy-5-methoxycinnamic acid exhibited DPPH free radical scavenging activity comparable to that of the antioxidants α-tocopherol and vitamin C in the concentration range of 2–50 μg / mL. In anti-glycation experiments, the compound exhibited moderate activity similar to that of the standard drug rutin. Based on their antioxidant potential, cinnamic acid derivatives such as ethylhexyl methoxycinnamate (cinnamyl ester), isoamyl p-methoxycinnamate (aminooxy ester), octyl cinnamyl ester, and cinnamyl cinnamyl ester are widely used in cosmetics. Furthermore, some natural and synthetic derivatives of cinnamic acid exhibit skin-whitening and anti-aging properties, particularly 4-hydroxycinnamic acid, which is currently listed as a new drug candidate for the treatment of hyperpigmentation. Cinnamic acid and its derivatives have also been widely reported in the prevention and treatment of diabetes and its complications, with mechanisms including stimulation of insulin secretion, improvement of pancreatic β-cell function, and inhibition of hepatic gluconeogenesis. According to a recent review, several reported cinnamic acid derivatives have demonstrated in vitro efficacy superior to positive controls or clinical therapeutic drugs, including anti-tumor, anti-tuberculosis (cinnamic acid itself has also been used to treat pulmonary tuberculosis), anti-Staphylococcus aureus, anti-malarial, and dual inhibition of human acetylcholinesterase (hAChE) and butyrylcholinesterase (hBuChE) (potential therapeutic activity for Alzheimer's disease). Furthermore, toxicology studies have confirmed the safety of these derivatives, indicating potential for further development.

[0003] On the other hand, acetaminophen (APAP) is an effective antipyretic and analgesic, but APAP overdose is the main cause of drug poisoning and acute liver failure. APAP overdose consumes a large amount of glutathione (GSH), leading to excessive accumulation of the intermediate metabolite N-acetyl-p-benzoquinoneimine (NAPQI). NAPQI covalently binds to sulfhydryl groups in cellular proteins, especially mitochondrial proteins, causing hepatocellular damage and liver necrosis. Oxaliplatin (OXA) is a third-generation platinum anticancer drug. Liver damage is a common and serious adverse reaction of OXA, with the main clinical manifestations being splenomegaly, thrombocytopenia, abnormal liver function, and portal hypertension. Its pathogenesis is closely related to OXA-induced activation of the liver DNA damage repair pathway and oxidative stress, causing damage to the liver sinusoidal endothelial cells, and further leading to platelet aggregation and vascular occlusion in the liver sinusoids.

[0004] [Summary of the invention]

[0005] In response to the above-mentioned defects or improvement needs of the prior art, the object of the present invention is to provide a cinnamic acid derivative and its preparation method and application. By improving the chemical structure of the cinnamic acid derivative and its preparation method, the new cinnamic acid derivative designed can be used to prepare drugs, for example, by inhibiting the MAPK signaling pathway to exert a therapeutic effect, by inhibiting ferroptosis to exert a therapeutic effect, by activating UDP-glucuronyl transferase to exert a detoxification effect, and is particularly useful for treating liver damage.

[0006] To achieve the above object, according to one aspect of the present invention, a cinnamic acid derivative is provided, characterized in that its structural formula is as shown in Formula 1A or Formula 1B:

[0007] And exclude formula 18:

[0008] For Formula 1A or Formula 1B:

[0009] The number of substituents R is 0 to 5; when the number of substituents R is 1 to 5, the substitution sites correspond to any one to all five of positions 2, 3, 4, 5, and 6 of the benzene ring; R at different substitution sites is independently selected from the group consisting of fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, ester, carboxyl, hydroxyl, and allyloxy.

[0010] The dashed and solid lines represent carbon-carbon single bonds or carbon-carbon double bonds.

[0011] As a further preferred embodiment of the present invention, the structural formula of the cinnamic acid derivative is any one of Formula 5 to Formula 17, and Formula 19 to Formula 30:

[0012] According to another aspect of the present invention, the present invention provides a method for synthesizing the above-mentioned cinnamic acid derivative, characterized in that it comprises the following steps:

[0013] (1) Cinnamic acid as shown in Formula 3A is used as a reaction substrate, and oxalyl chloride undergoes a substitution reaction under the action of a catalytic amount of DMF to obtain cinnamoyl chloride as shown in Formula 4A;

[0014] In Formula 3A, the number of substituents R1 is 0 to 5; when the number of substituents R1 is 1 to 5, the substitution sites correspond to any one to all five of positions 2, 3, 4, 5, and 6 of the benzene ring; R1 at different substitution sites is independently selected from the group consisting of fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, and ester;

[0015] Dashed and solid lines represent carbon-carbon single bonds or carbon-carbon double bonds;

[0016] (2) subjecting the cinnamoyl chloride obtained in step (1) as shown in Formula 4A to a substitution reaction with compound KD to generate a corresponding cinnamic acid derivative, the structural formula of which satisfies Formula 1A; or, subjecting the cinnamoyl chloride obtained in step (1) as shown in Formula 4A to a substitution reaction with compound MDG to generate a corresponding cinnamic acid derivative, the structural formula of which satisfies Formula 1B;

[0017] According to another aspect of the present invention, the present invention provides a method for synthesizing the above-mentioned cinnamic acid derivative, characterized in that it comprises the following steps:

[0018] S1: Cinnamic acid represented by the formula Cinnamic acids undergoes a substitution reaction with allyl bromide under alkaline conditions to produce a cinnamate represented by formula 2; then, the cinnamate represented by formula 2 is hydrolyzed under alkaline conditions and then acidified to produce cinnamic acid represented by formula 3B;

[0019] In the formula Cinnamic acids, the number of hydroxyl substituents is 1 to 5, and the substitution sites correspond to any one to all five of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; the number of substituents R2 is 0 to 4, and the sum of the number of hydroxyl substituents and substituents R2 does not exceed 5; when the number of substituents R2 is not 0, the substitution sites of the substituents R2 correspond to any one or more of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring except the hydroxyl substitution site; and R2 at different substitution sites is independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, and ester group;

[0020] Dashed and solid lines represent carbon-carbon single bonds or carbon-carbon double bonds;

[0021] S2: using the cinnamic acid (as shown in Formula 3B) obtained in step S1 as a reaction substrate and reacting it with oxalyl chloride in the presence of a catalytic amount of DMF to produce cinnamoyl chloride (as shown in Formula 4B);

[0022] S3: subjecting the cinnamoyl chloride obtained in step S2 as shown in Formula 4B to a substitution reaction with compound KD to generate a corresponding cinnamic acid derivative, the structural formula of which satisfies Formula 1A; alternatively, subjecting the cinnamoyl chloride obtained in step S2 as shown in Formula 4B to a substitution reaction with compound MDG to generate a corresponding cinnamic acid derivative, the structural formula of which satisfies Formula 1B;

[0023] According to another aspect of the present invention, the present invention provides the use of the above-mentioned cinnamic acid derivatives in the preparation of drugs for treating liver damage.

[0024] As a further preferred embodiment of the present invention, the drug for treating liver injury is a drug for treating acute liver injury.

[0025] According to the last aspect of the present invention, the present invention also provides various applications of a cinnamic acid derivative including Formula 18, wherein the structural formula of the cinnamic acid derivative is shown in Formula 1A or Formula 1B:

[0026] For Formula 1A or Formula 1B:

[0027] The number of substituents R is 0 to 5; when the number of substituents R is 1 to 5, the substitution sites correspond to any one to all five of positions 2, 3, 4, 5, and 6 of the benzene ring; R at different substitution sites is independently selected from the group consisting of fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, ester, carboxyl, hydroxyl, and allyloxy.

[0028] Dashed and solid lines represent carbon-carbon single bonds or carbon-carbon double bonds;

[0029] in:

[0030] The first application is the use of the cinnamic acid derivative in the preparation of drugs that exert therapeutic effects by inhibiting the MAPK signaling pathway;

[0031] The second application is the use of the cinnamic acid derivative in the preparation of a drug that exerts a therapeutic effect by inhibiting ferroptosis;

[0032] The third application is the use of the cinnamic acid derivative in the preparation of a drug that activates UDP-glucuronyl transferase to exert a therapeutic effect.

[0033] Through the above technical solutions conceived by the present invention, compared with existing technologies, the present invention integrates drug design concepts such as structural assembly and bioisostericity to design and synthesize multiple cinnamic acid derivatives having the chemical structures represented by Formulas 1A and 1B. These newly synthesized cinnamic acid derivatives are structurally stable and non-cytotoxic within an effective concentration range (as illustrated in the Examples below, experiments with cinnamic acid derivatives at concentrations of 10 μM to 1000 μM have demonstrated non-toxicity and efficacy at a concentration of 1 mM, with efficacy at a concentration of 10 μM).

[0034] Specifically, the present invention can achieve the following beneficial effects:

[0035] (1) The present invention provides novel cinnamic acid derivatives with specific chemical structures, which satisfy the general formulas 1A and 1B. The following examples of the present invention utilize high-resolution mass spectrometry, proton nuclear magnetic resonance spectroscopy, carbon nuclear magnetic resonance spectroscopy, and other techniques to comprehensively analyze the synthesized cinnamic acid derivatives and determine their chemical structures. The chemical structures of some cinnamic acid derivatives (e.g., compounds 5–17, 19–30) are shown below:

[0036] (2) The present invention provides two synthetic routes for the cinnamic acid derivatives having a specific chemical structure, depending on whether the raw materials are commercially available. They are as follows.

[0037] Among them, R1 and R2 can both be 0.

[0038] The starting material of route (a) is a compound of formula 3A, which can be directly purchased through commercial channels, wherein R1 is any one or more of fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, and ester groups (≥2, the same or different groups), and the substitution site is any one or more of positions 2, 3, 4, 5, and 6; the starting material of route (b) is a compound of formula Cinnamic Acid compounds (wherein, the number of hydroxyl groups is ≥1, and the corresponding substitution sites are any one or more of positions 2, 3, 4, 5, and 6; R2 at different substitution sites is independently selected from fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, and ester groups; the substitution sites are the remaining sites among positions 2, 3, 4, 5, and 6; and when R2 is ≥2, they can be a combination of the same or different groups) are first prepared to obtain the intermediate compound of formula 3B (the compound of formula 3B is not commercially available). Compounds of formula 3A and formula 3B can be used as reaction substrates, respectively, and undergo substitution reactions with oxalyl chloride under the action of a catalytic amount of DMF to obtain compounds of formula 4A and formula 4B. They continue to undergo substitution reactions with KD to generate the corresponding cinnamic acid derivatives satisfying formula 1A; if they undergo substitution reactions with MDG, the corresponding cinnamic acid derivatives satisfying formula 1B are generated.

[0039] Taking route (b) as an example (steps 3 to 4 below are common to route (a) and are described together), step 1: cinnamic acid containing a hydroxyl substituent (≥1 hydroxyl group) on the benzene ring undergoes a substitution reaction with allyl bromide under alkaline conditions (i.e., reaction i in the synthetic route) to produce cinnamate 2; step 2: cinnamate 2 obtained from step 1 above undergoes a hydrolysis reaction under alkaline conditions (i.e., reaction ii in the synthetic route), and then undergoes acidification (i.e., reaction iii in the synthetic route) to obtain cinnamate 2 containing an allyloxy group on the benzene ring. Step 3: Cinnamic acid 3B with or without substituents, or cinnamic acid 3B with an allyloxy substituent (≥1 allyloxy group) on the benzene ring, purchased directly from commercial channels, is used as a reaction substrate and reacted with oxalyl chloride in the presence of a catalytic amount of DMF to undergo a substitution reaction (i.e., reaction iv in the synthetic route); Step 4: KD or MDG is used as a reaction substrate and reacted with cinnamoyl chloride 4 to undergo a substitution reaction (i.e., reaction v in the synthetic route) to produce the corresponding cinnamic acid derivatives 5–30.

[0040] In the present invention, cinnamoyl chloride is used as one of the reaction substrates, and pyridine is used as a base to undergo substitution reactions with KD or MDG, respectively, to successfully obtain the target cinnamic acid derivatives. The above synthetic routes ensure the preparation of the target cinnamic acid derivatives having the structures shown in Formulas 1A and 1B. As illustrated in the comparative examples below, the inventors also attempted ester condensation reactions promoted by DCC or EDCI during the development process, but neither method successfully provided the target cinnamic acid derivatives.

[0041] (3) The present invention used APAP-induced acute liver injury (ALI) cell models and mouse models for preliminary screening and found that these cinnamic acid derivatives had anti-ALI effects. An OXA-induced ALI model was also constructed to further verify the anti-ALI effects of these cinnamic acid derivatives. As shown in the examples below, compounds 16 and 28 showed stronger anti-liver injury effects than the positive drug N-acetyl-L-cysteine ​​(NAC).

[0042] NAC is currently the only clinically approved antidote for APAP-induced ALI. However, it is only effective for patients within eight hours of APAP poisoning and is ineffective for patients with advanced APAP poisoning or severe liver damage. Excessive use can also prolong liver recovery and regeneration. The cinnamic acid derivative developed by this invention exhibits a stronger effect than the positive control, NAC.

[0043] In addition, among the common enzymes involved in drug metabolism, UGT enzyme (uridine diphosphate glucuronyl transferase) accounts for the second largest proportion and is one of the most important enzymes in the human phase II metabolism. UGT enzyme is a membrane protein bound to the endoplasmic reticulum, which plays a catalytic role in the process of transferring glucuronic acid from UDP-glucuronic acid (UDPGA) to other molecules (usually hydrophobic molecules). UGT enzyme uses glucuronic acid as a glycosyl donor and widely catalyzes the binding reaction of endogenous (such as bilirubin, fatty acids, steroid hormones, compounds in food, drugs, environmental pollutants, etc.) and exogenous chemical substances. Usually, the compound after the binding reaction is more water-soluble and easy to be excreted from the body, thereby playing a detoxification role. It also participates in the metabolic clearance process of many drugs. Therefore, the new cinnamic acid derivatives discovered by the present invention that can be used for diseases related to MAPK signaling pathway activation, ferroptosis reaction, and liver detoxification-related mechanisms are of great significance to the development of related new drugs.

[0044] As illustrated in the examples below, the present invention uses APAP-induced ALI cell and mouse models to demonstrate that multiple cinnamic acid derivatives have anti-ALI effects, which are stronger than the positive control NAC. OXA-induced cell and mouse ALI models were also constructed to further verify the anti-ALI effects of the above-mentioned compounds (i.e., cinnamic acid derivatives satisfying general formulas 1A and 1B, represented by compound formulas 16 and 28). Further mechanistic studies have shown that the effects of the cinnamic acid derivatives of the present invention are related to inhibiting MAPK signaling pathway activation and ferroptosis reactions in the body, and enhancing UDP-glucuronyl transferase activity. The present invention has discovered new cinnamic acid derivatives related to the preparation of drugs for treating diseases related to MAPK signaling pathway activation, ferroptosis reactions, and UDP-glucuronyl transferase activation, which is of great significance for the development of related new drugs.

[0045] Over the past 50 years, researchers have been diligently developing potentially effective treatments for APAP-induced ALI. Promising compounds reported so far include 4-methylpyrazole (4MP, a CYP450 and JNK inhibitor), calmangafodipir (CMFP, a SOD mimetic), metformin, and methylene blue. However, these active molecules have yet to become marketed drugs. Providing more candidate anti-ALI active molecules would undoubtedly enrich preclinical research and significantly advance the development of related drugs. Many other molecules and natural products have also been reported to be effective in ALI models, but most studies have significant flaws in their experimental design. For example, mice were pretreated with compounds before APAP modeling and then their efficacy was assessed, sometimes even for a month. This is completely inconsistent with clinical practice and has hindered the further development of these compounds. Consequently, more than 50 years after NAC was approved for the clinical treatment of ALI, no other drugs remain on the market.

[0046] In the present invention, as illustrated in the Examples below, we first established an experimental model of APAP poisoning for 1 hour (at which point liver damage has already occurred), and then administered the test compound, similar to the situation in patients seeking clinical treatment. The hepatoprotective effects of the compound were then evaluated on this basis. Simultaneously, we established a high-dose APAP-induced acute liver failure (ALF) model in mice, which can simulate ALF patients with excessive APAP intake or patients with advanced APAP poisoning at standard doses. These patients are essentially incurable clinically, and even NAC treatment is ineffective. Using compounds of Formula 16 and Formula 28 as examples, survival experiments based on the ALF mouse model showed that treatment with CK16 and CG28 significantly prolonged the survival of mice, while NAC did not. These results demonstrate that both CK16 and CG28 have stronger hepatoprotective effects than NAC. From a mechanism of action perspective, the novel cinnamic acid derivatives CK16 and CG28 alleviate APAP hepatotoxicity by activating UDP-glycosyltransferases and enhancing APAP glucuronidation, while simultaneously inhibiting MAPK signaling pathway activation and ferroptosis. Compared to NAC, they are more effective and have completely different mechanisms, offering certain advantages and novelty. CK16 and CG28 are also the only two UGT activators discovered so far, with no compounds with similar mechanisms reported.

Brief Description of the Drawings

[0047] Figure 1 shows the cytotoxic activity screening of cinnamic acid derivatives. Each grayscale column in the figure corresponds to the experimental results using different concentrations of KD, MDG, and compounds 5 to 30. The first blank column corresponds to the negative control group in which DMSO (<0.1%) was applied. For the remaining gray columns, "-" indicates that the corresponding compound (e.g., KD, compound 5, compound 9, NAC, etc.; the same below) was not applied; "+" indicates that the corresponding compound was applied. The specific value indicates that the corresponding compound was applied and the compound concentration met the value.

[0048] Figure 2 shows the screening of cinnamic acid derivatives for their anti-APAP-induced liver injury activity in a mouse model. The figure shows the injection of multiple test compounds one hour after APAP injection. The experiment was terminated 23 hours later, and samples were collected to measure serum alanine aminotransferase and aspartate aminotransferase levels in each experimental group. The "Control" group in the figure represents normal mice, the "APAP" group represents the model group, and the "NAC" group represents the positive control group.

[0049] Figure 3 shows the anti-APAP-induced liver injury activity of cinnamic acid derivatives CK16 and CG28. Figure 3A shows the treatment of mice over the experimental time period; Figure 3B shows the serum alanine aminotransferase and aspartate aminotransferase levels in mice treated with NAC (50, 100, 200, and 300 mg / kg); Figure 3C shows the serum alanine aminotransferase and aspartate aminotransferase levels in mice treated with CK16, CG28, or NAC; Figure 3D shows representative liver photographs and H&E-stained liver sections; Figure 3E shows the quantification of liver necrosis area in different groups; Figure 3F shows representative images of liver TUNEL, HMGB1, and p-MLKL staining; Figure 3G shows mice treated with CK16, CG28, or NAC 1 hour after intraperitoneal injection of APAP (600 mg / kg), and the 24-hour survival rate of each group was recorded. Taking A in Figure 3 as an example, multiple test compounds were injected 1 hour after APAP injection, and the experiment was terminated 23 hours later to collect samples for testing. The data in B to F in Figure 3 also follow this experimental design.

[0050] Figure 4 shows the anti-oxaliplatin-induced liver injury activity of cinnamic acid derivatives CK16 and CG28. Figure 4 (A) shows serum alanine aminotransferase and aspartate aminotransferase levels in mice treated with different groups in an in vivo oxaliplatin experiment; Figure 4 (B) shows the activity of CK16, CG28, or NAC in an in vitro oxaliplatin-induced acute liver injury cell model.

[0051] Figure 5 shows that the cinnamic acid derivatives CK16 and CG28 inhibit MAPK signaling pathway activation in vivo. Western blot analysis in Figure 5 examined the expression of p-P38, P38, p-ERK1 / 2, ERK1 / 2, p-JNK, and JNK in the livers of control mice, APAP-treated mice, and mice treated with CK16 (or CG28) 24 hours after APAP treatment. β-actin was used as a protein control.

[0052] Figure 6 shows that cinnamic acid derivatives CK16 and CG28 inhibit ferroptosis in vivo. Figure 6 (A) shows immunohistochemical staining of 4-HNE expression in liver tissue (scale bar: 80 μm); Figure 6 (B) shows immunohistochemical staining of DAB-enhanced Prussian blue expression in liver tissue (scale bar: 200 μm); Figure 6 (C) shows Western blot analysis of GPX4, FSP1, SLC7A11, TfR1, FTL, FTH1, FPN, and FABP4 expression in the liver of control mice, APAP group mice, and CK16 or CG28 group mice after 24 hours of APAP treatment, with β-actin as a protein expression control; Figure 6 (D) shows RT-PCR detection of FGF21, DHODH, FTH1, GPX4, SLC7A11, DHFR, TfR1, FPN, FTL, and GCH1 mRNA expression in mouse liver.

[0053] Figure 7 shows the enhancement of UDP-glucuronosyltransferase activity by the cinnamic acid derivatives CK16 and CG28. Figure 7 (A) shows the kinetics of glucuronidation of the fluorescent substrate in mouse donor liver microsomes (MLMs, 0.05 mg / mL) at 37°C, as well as the inhibition of UGT activity in MLMs by CK16 or CG28 and the nonselective UGT ligand diclofenac. Under blank reaction conditions, the cofactor UDPGA was replaced with vehicle (assay buffer). Dose-response curves for UGT activation in MLMs by CK16 or CG28 were generated, and the percentage activity at each concentration was calculated by comparison with the activity in reactions containing vehicle alone. Figure 7 (B) shows the levels of APAP-Cys, APAP-Gluc, and APAP-Sulf in mouse serum. [Specific implementation method]

[0054] In order to make the objectives, technical solutions and advantages of the present invention more clearly understood, the present invention is further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely for the purpose of explaining the present invention and are not intended to limit the present invention. In addition, the technical features involved in the various embodiments of the present invention described below may be combined with each other as long as they do not conflict with each other.

[0055] Compound KD used in Example 1 below was extracted and isolated from the natural plant Anoectochilus roxburghii using a conventional method (Journal of Ethnopharmacology 2007, 114, 141–145). Compound MDG was purchased from a reagent store. Its Chinese name is methyl-α-D-pyranoglucopyranoside, and its CAS number is 97-30-3. The raw materials, compound 3A and cinnamic acids, were also purchased from a reagent store.

[0056] Example 1: Synthesis and structural identification of cinnamic acid derivatives

[0057] (1) Synthesis of cinnamic acid derivatives. The synthetic route is as follows:

[0058] The synthetic routes and key reaction raw materials used in formulas 5-30 of the present invention are shown in the following table:

[0059] Taking cinnamic acid derivative 28 as an example, the substrate raw materials and synthetic intermediates required in its synthesis process are as follows:

[0060] The reactions in the synthetic route are carried out as follows:

[0061] (i-iii) Synthesis of cinnamic acid 3B (with ≥1 allyloxy substituent on the benzene ring)

[0062] A dry reaction flask was charged with a magnetite, cinnamic acid (30 mmol, 1.0 equiv.), and potassium carbonate (x mmol. Assuming a compound has one hydroxyl group and one carboxyl group, x = 180 mmol = 6.0 equiv.; assuming a compound has two hydroxyl groups and one carboxyl group, x = 270 mmol = 9.0 equiv.; taking the preparation of cinnamic acid derivative 28 as an example, cinnamic acid derivative 28 has one allyloxy group in its structure, and the raw material cinnamic acid for synthesizing 28 contains one hydroxyl group and one carboxyl group, then 180 mmol, i.e., 6.0 equiv., of potassium carbonate should be added). 120 mL of acetonitrile was added as the reaction solvent, and allyl bromide (x mmol, equivalent to the amount of potassium carbonate; dissolved in an appropriate amount of acetonitrile solvent) was slowly added by injection in portions under stirring. The reaction flask was then transferred to an 80°C oil bath and heated under reflux for overnight reaction. The next day, the reaction flask was moved to room temperature for cooling, and the reaction was monitored by TLC spot plate. After the reaction was complete, the reaction solution was filtered, the filter residue was washed with ethyl acetate, and the filtrate (a mixed solution of acetonitrile and ethyl acetate) was collected and concentrated under the conditions of a rotary evaporator. The concentrate was dissolved with ethyl acetate, and extracted three times with ethyl acetate and water as two phases. The ethyl acetate layer was collected, dried over anhydrous sodium sulfate, filtered, and concentrated on a rotary evaporator to obtain cinnamate 2, which can be directly used in the next step without further purification treatment.

[0063] The cinnamate 2 (25 mmol, 1.0 equiv.) obtained in the previous step was added to a dry reaction flask, a magnetic bar was added, and a mixed solvent of methanol and water (40 mL:40 mL) was added and stirred at room temperature. Sodium hydroxide (100 mmol, 4.0 equiv.) was slowly added to the reaction system in batches. After the addition was complete, the reaction flask was moved to a 60°C oil bath and heated under reflux for overnight reaction. The next day, the reaction flask was moved to room temperature for cooling, and the reaction was monitored by TLC spot plate. After the reaction was complete, preliminary concentration was performed under the conditions of a rotary evaporator to remove the methanol solvent. Subsequently, the concentrate was dissolved with dichloromethane and extracted three times with dichloromethane and water as two phases. The aqueous layer was collected and placed in a separatory funnel. 1M HCl aqueous solution was added to adjust the pH to 1-2. An appropriate amount of ethyl acetate was added and extracted three times. The ethyl acetate layer was collected and the aqueous layer was spotted by TLC to observe whether the organic matter in the aqueous layer was completely extracted by ethyl acetate. Anhydrous sodium sulfate was added to the collected ethyl acetate layer for drying, filtration, and concentration on a rotary evaporator to obtain cinnamic acid 3B, which can be directly used in the next step without further purification treatment.

[0064] (iv) Synthesis of cinnamoyl chloride 4A or 4B (with ≥1 allyloxy substituent on the benzene ring)

[0065] Commercially available cinnamic acid 3A (15 mmol, 1.0 equiv.) or cinnamic acid 3B (15 mmol, 1.0 equiv.) prepared via the above steps was added to a dry reaction flask. A magnetic bar was added, followed by 40 mL of anhydrous dichloromethane and a catalytic amount of N,N-dimethylformamide (0.75 mmol, 0.05 equiv.). Oxalyl chloride (45 mmol, 3.0 equiv.) was slowly added dropwise by injection. The reaction solution was stirred at room temperature for 2-6 hours and monitored by TLC plate until the reaction was complete. Note that gas is generated in the reaction system, so the gas balance should be carefully maintained during the addition process. After completion of the reaction, the solution was concentrated on a rotary evaporator to remove the solvent, yielding cinnamoyl chloride 4A or 4B, which was used directly in the next step without further purification.

[0066] (v) Synthesis of cinnamic acid derivatives 5–30

[0067] KD (10 mmol, 1.0 equiv.) or MDG (10 mmol, 1.0 equiv.) was added to a dry reaction flask, a magnet was added, dry anhydrous pyridine (100 mmol, 10.0 equiv.), and an appropriate amount of anhydrous dichloromethane was added as solvent. The reaction flask was stirred at -10°C. Cinnamoyl chloride 4A (11 mmol, 1.1 equiv.) or 4B (11 mmol, 1.1 equiv.) prepared in the above step was dissolved in anhydrous dichloromethane and slowly added dropwise to the reaction flask by injection. The reaction flask was then moved to room temperature and stirred overnight. The next day, TLC was performed to monitor the reaction. After completion, a small amount of methanol was added to quench the reaction system and concentrated on a rotary evaporator. The concentrate was dissolved in dichloromethane and transferred to a separatory funnel, and 1 M The solution was adjusted to pH 5-6 with aqueous HCl solution, and extracted with dichloromethane and water for about six times until the dichloromethane layer no longer contained the target compound. The dichloromethane layer was collected, dried over anhydrous sodium sulfate, filtered, and concentrated on a rotary evaporator to obtain a crude product. An appropriate amount of dichloromethane was added to dissolve the crude product, and 100-200 mesh silica gel was added for mixing. The sample was separated by column chromatography (200-300 mesh silica gel, petroleum ether: ethyl acetate = 3:1-1:4) to obtain cinnamic acid derivatives 5-30, respectively.

[0068] (II) Structural identification of cinnamic acid derivatives 5–30

[0069] The structure of cinnamic acid derivatives 5–30 was determined by comprehensive analysis of high-resolution mass spectrometry, H NMR spectra, and C NMR spectra. The results are as follows:

[0070] Methyl ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3yl)oxy)tetrahydro-2H-pyran-2-yl)cinnamate (5)

[0071] White solid, 1.69 g, 42.9% yield; mp (melting point): 185–187°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1H NMR(400MHz,MeOD)δ7.73(d,J=16.1Hz,1H),7.65–7.59(m,2H),7.41(dt,J=4.5,1.7Hz,3H),6.58(d,J=16 .0Hz,1H),4.65(ddt,J=6.6,4.2,2.0Hz,1H),4.56(dd,J=11.9,2.2Hz,1H),4.50(dt,J=10.3,1.2Hz,1H), 4.47–4.43(m,1H),4.41(d,J=7.8Hz,1H),4.37(dd,J=11.9,6.0Hz,1H),3.57(ddd,J=9.5,5.9,2.1Hz,1H) ,3.42–3.34(m,2H),3.25–3.19(m,1H),2.85(dd,J=18.0,6.5Hz,1H),2.64(ddd,J=18.0,2.1,0.9Hz,1H). 13 C NMR(150MHz,MeOD)δ178.4,168.4,146.6,135.7,131.6,130.0,129.3,118.6,104.0,77.7,76.7,76.1,75.5,74.8,71.5,64.6,36.0.HRMS(ESI):calcd.For C 19 H 22 O9Na[M+Na] + :417.1156, found 417.1163.

[0072] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl 3-phenylpropionate (6)

[0073] White solid, 1.06 g, 26.7% yield; mp: 173-175°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(400MHz,MeOD)δ7.29–7.25(m,2H),7.23–7.21(m,2H),7.19–7.16(m,1H),4.56(ddt,J=6.6,4.7,2.0Hz,1H),4.46 (dd,J=11.9,2.2Hz,1H),4.45–4.42(m,1H),4.38(dd,J=10.3,4.7Hz,1H),4.34(d,J=7.8Hz,1H),4.17(dd,J=11.9,6. 0Hz,1H),3.44(ddd,J=9.7,6.0,2.2Hz,1H),3.34(t,J=9.1Hz,1H),3.28(dd,J=9.7,8.9Hz,1H),3.17(dd,J=9.2,7.8H z,1H),2.94(t,J=7.6Hz,2H),2.83(dd,J=18.0,6.6Hz,1H),2.68(t,J=7.6Hz,2H),2.62(ddd,J=18.0,2.1,0.9Hz,1H). 13 C NMR (150 MHz, MeOD) δ177.0,173.0,140.6,128.1,128.0,125.9,102.6,76.3,75.3,74.7,74.0,73.3,70.0,63.1,35.4,34.7,30.5.HRMS(ESI):calcd.for C 19 H 24 O9Na[M+Na] + :419.1313, found 419.1312.

[0074] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-fluorophenyl)acrylate (7)

[0075] White solid, 1.66 g, 40.2% yield; mp: 114-116°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(600MHz,MeOD)δ7.71(d,J=16.0Hz,1H),7.69–7.64(m,2H),7.17–7.12(m,2H),6.53(d,J=16.0 Hz,1H),4.65(ddt,J=6.6,4.8,2.0Hz,1H),4.55(dd,J=11.9,2.2Hz,1H),4.51–4.48(m,1H),4.44(d d,J=10.4,4.7Hz,1H),4.41(d,J=7.8Hz,1H),4.36(dd,J=11.9,6.0Hz,1H),3.58–3.54(m,1H),3.40 –3.34(m,2H),3.23–3.19(m,1H),2.85(dd,J=18.0,6.6Hz,1H),2.63(ddd,J=18.1,2.1,0.9Hz,1H). 13 C NMR (150MHz, MeOD) δ178.4, 168.3, 165.4 (d, J = 249Hz), 145.3, 132.2 (d, J = 3Hz), 131.5 (d, J = 9Hz), 118.5 (d, J=3Hz),116.9(d,J=22.5Hz),104.0,77.7,76.7,76.1,75.5,74.8,71.5,64.6,36.0.HRMS(ESI):calcd.for C 19 H 21 FO9Na[M+Na] + :435.1062, found 435.1059.

[0076] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-chlorophenyl)acrylate (8)

[0077] White solid, 1.23 g, 28.7% yield; mp: 146-148°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(400MHz,MeOD)δ7.69(d,J=16.0Hz,1H),7.64–7.57(m,2H),7.44–7.38(m,2H),6.57(dd,J=16.0,1. 0Hz,1H),4.65(ddt,J=6.6,4.3,2.0Hz,1H),4.55(dd,J=11.9,2.2Hz,1H),4.49(dd,J=10.3,1.9Hz,1H) ,4.46–4.43(m,1H),4.41(d,J=7.8Hz,1H),4.36(dd,J=11.9,6.0Hz,1H),3.56(ddd,J=8.6,5.9,2.3Hz, 1H),3.42–3.33(m,2H),3.25–3.17(m,1H),2.85(dd,J=18.0,6.5Hz,1H),2.63(dd,J=17.9,2.1Hz,1H). 13 C NMR(100MHz,MeOD)δ178.5,168.2,145.2,137.5,134.6,130.9,130.3,119.6,104.2,77.8,76.8,76.2,75.6,74.9,71.6,64.8,36.2.HRMS(ESI):calcd.for C 19 H 21 ClO9Na[M+Na] + :451.0766, found 451.0769, 453.0742.

[0078] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-bromophenyl)acrylate (9)

[0079] White solid, 1.19 g, 25.3% yield; mp: 125-127°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(600MHz,MeOD)δ7.68(d,J=16.0Hz,1H),7.59–7.50(m,4H),6.64–6.57(m,1H),4 .64(ddd,J=6.7,4.7,2.4Hz,1H), 4.55(dd,J=11.9,2.3Hz,1H), 4.49(dd,J=10.5,1. 7Hz,1H),4.44(dt,J=10.4,4.9Hz,1H),4.41(d,J=7.8Hz,1H),4.36(dd,J=11.9,6.0 Hz,1H),3.55(ddd,J=8.6,6.0,2.3Hz,1H),3.41–3.33(m,2H),3.21(t,J=8.3Hz,1H), 2.85(dd,J=18.1,6.5Hz,1H), 2.63(dd,J=18.1,1.9Hz,1H). 13 C NMR(150MHz,MeOD)δ178.4,168.1,145.1,134.8,133.2,130.9,125.6,119.6,104.0,77.7,76.7,76.1,75.5,74.8,71.5,64.7,36.0.HRMS(ESI):calcd.for C 19 H 21 BrO9Na[M+Na] + :495.0261, found 495.0264, 497.0243.

[0080] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-(trifluoromethyl)phenyl)acrylate (10)

[0081] White solid, 1.27 g, 27.4% yield; mp: 202-204°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR (600MHz, MeOD) δ7.81(d,J=8.1Hz,2H),7.77(d,J=16.0Hz,1H),7.70(d,J=8.1Hz,2H),6.71(d,J=16.0H z,1H),4.65(ddt,J=6.8,4.3,2.0Hz,1H),4.57(dd,J=12.0,2.2Hz,1H),4.49(dd,J=10.4,1.7Hz,1H),4.44 (dd,J=10.3,4.6Hz,1H),4.42(d,J=7.8Hz,1H),4.38(dd,J=11.9,5.9Hz,1H),3.57(ddd,J=8.6,5.8,2.2Hz ,1H),3.41–3.36(m,2H),3.22(t,J=8.2Hz,1H),2.85(dd,J=18.0,6.5Hz,1H),2.64(dd,J=18.1,2.0Hz,1H). 13 C NMR (150MHz, MeOD) δ178.4,167.8,144.5,139.5,132.8(q,J=31.5Hz),129.8,126.9(q,J=4.5Hz),125 .4(d,J=270Hz),121.6,104.0,77.9,76.7,76.1,75.5,74.8,71.5,64.8,36.0.HRMS(ESI):calcd.for C 20 H 21 F3O9Na[M+Na] + :485.1030, found 485.1042.

[0082] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(naphthalen-1-yl)acrylate (11)

[0083] Pale yellow solid, 1.19 g, 26.9% yield; mp: 140-142°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1H NMR(600MHz,MeOD)δ8.57(d,J=15.8Hz,1H),8.19(dd,J=8.6,1.1Hz,1H),7.94(d,J=8.2Hz,1H),7.93–7.88(m,1H),7.85(dt,J=7.3,0.8Hz,1 H),7.60(ddd,J=8.5,6.9,1.4Hz,1H),7.54(ddd,J=8.0,6.8,1.1Hz,1H),7.53–7.48(m,1H),6.63(d,J=15.7Hz,1H),4.66(ddt,J=6.7,4.8,2 .0Hz,1H),4.62(dd,J=11.9,2.3Hz,1H),4.52(ddd,J=10.3,1.8,0.9Hz,1H),4.45(d,J=4.8Hz,1H),4.46–4.39(m,2H),3.60(ddd,J=9.6,6.0 ,2.3Hz,1H),3.43–3.38(m,2H),3.27–3.21(m,1H),2.84(dd,J=18.1,6.6Hz,1H),2.64(ddd,J=18.1,2.1,0.9Hz,1H).HRMS(ESI):calcd.for C 23 H 24 O9Na[M+Na] + :467.1313, found 467.1308.

[0084] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(p-tolyl)acrylate (12)

[0085] White solid, 2.05 g, 50.2% yield; mp: 172-174°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(400MHz,MeOD)δ7.63–7.56(m,1H),7.39(dd,J=8.4,2.1Hz,2H),7.18–7.09(m,2H), 6.40(d,J=16.0Hz,1H),4.56(ddt,J=6.6,4.3,2.0Hz,1H),4.47(dd,J=11.9,2.2Hz,1H) ,4.41(dd,J=10.3,1.8Hz,1H),4.36(d,J=4.8Hz,1H),4.33(d,J=7.8Hz,1H),4.27(dd,J =11.9,6.0Hz,1H),3.49(ddd,J=8.6,6.1,2.2Hz,1H),3.37–3.25(m,2H),3.20–3.10(m, 1H), 2.76 (dd, J=18.0, 6.5Hz, 1H), 2.55 (dd, J=18.0, 2.0Hz, 1H), 2.26 (s, 3H). 13 C NMR(100MHz,MeOD)δ178.4,168.6,146.6,142.3,132.9,130.7,129.3,117.5,10 4.0,77.7,76.6,76.1,75.5,74.7,71.5,64.6,36.0,21.5.HRMS(ESI):calcd.for C 20 H 24 O9Na[M+Na] + :431.1313, found 431.1310.

[0086] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-methoxyphenyl)acrylate (13)

[0087] White solid, 0.90 g, 21.2% yield; mp: 138-140°C; column chromatography eluent: petroleum ether / ethyl acetate (1:6 v / v). 1H NMR(600MHz,MeOD)δ7.68(d,J=15.9Hz,1H),7.58–7.54(m,2H),6.98–6.93(m,2H),6.42(d,J=16.0Hz,1H),4.64( ddt,J=6.7,4.8,2.0Hz,1H),4.53(dd,J=11.9,2.2Hz,1H),4.49(ddd,J=10.3,1.9,0.9Hz,1H),4.44(dd,J=10.4, 4.7Hz,1H),4.40(d,J=7.8Hz,1H),4.35(dd,J=11.9,6.1Hz,1H),3.83(s,3H),3.55(ddt,J=8.1,3.6,2.2Hz,1H), 3.40–3.33(m,2H),3.21(td,J=7.7,1.5Hz,1H),2.85(dd,J=18.1,6.6Hz,1H),2.63(ddd,J=18.0,2.0,0.8Hz,1H). 13 C NMR(150MHz,MeOD)δ178.4,168.8,163.2,150.0,146.4,131.0,128.2,115.8,115.4 ,104.0,77.7,76.6,76.1,75.5,74.7,71.5,64.5,55.9,36.0.HRMS(ESI):calcd.for C 20 H 24 O 10 Na[M+Na] + :447.1262, found 447.1258.

[0088] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-(allyloxy)phenyl)acrylate (14)

[0089] Pale yellow solid, 1.91 g, 42.5% yield; mp: 129-131°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR (400MHz, MeOD) δ7.67(d,J=15.9Hz,1H),7.54(dd,J=8.7,1.5Hz,2H),6.96(dd,J=8.5,1.4Hz,2H),6.46–6.36(m,1H),6.05(ddt,J=17 .1,10.4,5.2Hz,1H),5.40(dq,J=17.3,1.7Hz,1H),5.26(dq,J=10.7,1.5Hz,1H),4.64(ddt,J=6.6,4.3,2.0Hz,1H),4.58(dq,J=5.1,1.5H z,2H),4.54(dd,J=11.9,2.2Hz,1H),4.49(dd,J=10.3,1.9Hz,1H),4.47–4.40(m,1H),4.41(d,J=7.8Hz,1H),4.35(dd,J=11.9,6.1Hz,1H) ,3.56(ddd,J=8.7,6.2,2.2Hz,1H),3.44–3.31(m,2H),3.22(t,J=8.3Hz,1H),2.85(dd,J=18.0,6.5Hz,1H),2.63(dd,J=17.8,2.0Hz,1H). 13 C NMR(100MHz,MeOD)δ178.4,168.8,162.1,146.4,134.5,131.0,128.4,117.8,116.2,11 5.9,104.0,77.9,76.6,76.1,75.5,74.7,71.5,69.8,64.5,36.0.HRMS(ESI):calcd.for C 22 H 26 O 10 Na[M+Na] + :473.1418, found 473.1413.

[0090] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(3-(allyloxy)-4-methoxyphenyl)acrylate (15)

[0091] Pale yellow solid, 2.32 g, 48.3% yield; mp: 115-117°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1H NMR (400MHz, MeOD) δ7.64(dd,J=15.9,5.6Hz,1H),7.23–7.14(m,2H),7.02–6.94(m,1H),6.41(dd,J=16.0,1.5Hz,1H),6.08(ddt,J=17.2,10.6,5.4H z,1H),5.42(dq,J=17.3,1.7Hz,1H),5.26(dq,J=10.6,1.5Hz,1H),4.64(d dt,J=6.7,4.3,2.1Hz,1H),4.60(dt,J=5.4,1.6Hz,2H),4.54(dd,J=11.9, 2.2Hz,1H),4.49(dd,J=10.2,1.9Hz,1H),4.45(d,J=4.7Hz,1H),4.40(d,J=7.8Hz,1H),4.35(dd,J=11.9,6.0Hz,1H),3.86(s,3H),3 .55(ddd,J=8.6,6.0,2.3Hz,1H),3.44–3.34(m,2H),3.26–3.17(m,1H),2.85(dd,J=18.0,6.5Hz,1H),2.63(dd,J=17.9,2.1Hz,1H). 13 C NMR(100MHz,MeOD)δ178.3,168.7,153.3,149.6,146.6,134.7,128.6,124.3,117.9,116.2,113.7 ,112.9,104.0,77.7,76.6,76.1,75.5,74.8,71.5,70.9,64.5,56.5,36.0.HRMS(ESI):calcd.for C 23 H 28 O 11 Na[M+Na] + :503.1524, found 503.1521.

[0092] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-(allyloxy)-3,5-dimethoxyphenyl)acrylate (16)

[0093] Pale yellow solid, 2.23 g, 43.7% yield; mp: 113-115°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1H NMR (600MHz, MeOD) δ7.66(d,J=15.9Hz,1H),6.92(s,2H),6.52(d,J=15.9Hz,1H),6.04(ddt,J=16.5,10.5,5.9Hz,1H),5.29( dd,J=17.2,1.7Hz,1H),5.15(dq,J=10.4,1.3Hz,1H),4.64(ddt,J=6.7,4.4,2.1Hz,1H),4.54(dd,J=11.9,2.2Hz,1H),4.49(d t,J=6.3,1.5Hz,3H),4.44(dd,J=10.4,4.7Hz,1H),4.40(d,J=7.8Hz,1H),4.36(dd,J=12.0,6.0Hz,1H),3.86(s,6H),3.55(d dd,J=8.6,5.9,2.3Hz,1H),3.42–3.33(m,2H),3.24–3.18(m,1H),2.85(dd,J=18.1,6.6Hz,1H),2.63(dd,J=18.0,2.0Hz,1H). 13 C NMR(150MHz,MeOD)δ178.3,168.5,155.0,146.7,139.9,135.6,131.5,118.1,117.9,106.7 ,104.1,77.7,76.7,76.1,75.5,75.1,74.8,71.5,64.5,56.7,36.0.HRMS(ESI):calcd.for C 24 H 30 O 12 Na[M+Na] + :533.1629, found 533.1630.

[0094] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(3,4-bisallyloxyphenyl)acrylate (17)

[0095] Yellow solid, 2.08 g, 41.0% yield; mp: 155-157°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1H NMR (600MHz, MeOD) δ7.64(d,J=15.9Hz,1H),7.22(d,J=2.1Hz,1H),7.16(dd,J=8.4,2.1Hz,1H),6.97(d,J=8.4Hz,1H),6.41(d,J=15.9Hz ,1H),6.08(dddt,J=17.2,10.4,6.7,5.2Hz,2H),5.43(m,2H),5.27(m,2H),4.64(ddt,J=6.7,4.8,2.0Hz,1H),4.62(dt,J=5.2,1.6Hz,4H ), 4.54(dd,J=11.9,2.3Hz,1H), 4.49(ddd,J=10.4,1.9,0.9Hz,1H), 4.44(dd,J=10.4,4.8Hz,1H), 4.40(d,J=7.8Hz,1H), 4.35(dd,J=11. 9,6.1Hz,1H),3.58–3.52(m,1H),3.42–3.33(m,2H),3.24–3.18(m,1H),2.85(dd,J=18.1,6.6Hz,1H),2.63(ddd,J=18.0,2.2,0.9Hz,1H). 13 C NMR(150MHz,MeOD)δ178.3,168.7,152.3,150.0,146.6,134.8,134.5,128.9,124.1,117.8,117.7,116.3 ,114.8,114.2,104.0,77.7,76.7,76.1,75.5,74.8,71.5,71.0,70.7,64.5,36.0.HRMS(ESI):calcd.for C 25 H 30 O 11 Na[M+Na] + :529.1680, found 529.1684. Methyl ((2R,3S,4S,5R)-3,4,5-trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)cinnamate (18)

[0096] White solid, 1.24 g, 38.3% yield; mp: 107-109°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1H NMR(400MHz,MeOD)δ7.71(d,J=16.0Hz,1H),7.62–7.58(m,2H),7.43–7.38(m,3 H),6.55(dd,J=16.0,0.9Hz,1H),4.68(d,J=3.7Hz,1H),4.51(dd,J=11.9,2.2Hz ,1H),4.34(dd,J=11.9,5.9Hz,1H),3.79(ddd,J=10.1,5.9,2.2Hz,1H),3.65(t, J=9.2Hz,1H),3.45(d,J=3.8Hz,1H),3.42(s,3H),3.36(dd,J=10.1,8.9Hz,1H). 13 C NMR(100MHz,MeOD)δ168.5,146.5,135.7,131.6,130.0,129.3,118.6,101.3,75.0,73.5,71.9,71.1,64.9,55.6.HRMS(ESI):calcd.for C 16 H 20 O7Na[M+Na] + :347.1101, found 347.1104.

[0097] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-p-fluorophenyl)acrylate (19)

[0098] White solid, 1.01 g, 29.6% yield; mp: 106-108°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1 H NMR (400MHz, MeOD) δ7.73–7.60(m,3H),7.14(t,J=8.7Hz,2H),6.50(d,J=16.0Hz,1H),4.68(d,J=3.7Hz,1H),4.50(dd,J=11.9,2.2Hz,1H),4. 34(dd,J=11.9,5.9Hz,1H),3.79(ddd,J=10.1,5.9,2.2Hz,1H),3.65(t,J=9.3Hz,1H),3.45(d,J=3.8Hz,1H),3.42(s,3H),3.39–3.35(m,1H). 13C NMR (100MHz, MeOD) δ168.3, 165.4 (d, J = 248Hz), 145.1, 132.2 (d, J = 3Hz), 131.5 (d, J = 9Hz), 118.5 (d,J=2Hz),116.9(d,J=22Hz),101.3,75.0,73.5,71.9,71.1,65.0,55.6.HRMS(ESI):calcd.for C 16 H 19 FO7Na[M+Na] + :365.1007, found 365.1016.

[0099] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-chlorophenyl)acrylate (20)

[0100] White solid, 1.03 g, 28.8% yield; mp: 107–109°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1 H NMR (600MHz, MeOD) δ7.68(d,J=16.0Hz,1H),7.63–7.58(m,2H),7.43–7.39(m,2H),6.57(d,J=16.1Hz,1H),4.68(d,J=3.7Hz,1H),4.50(dd,J=11.8,2. 2Hz,1H),4.34(dd,J=11.8,5.9Hz,1H),3.80–3.76(m,1H),3.64(dd,J=9.6 ,8.9Hz,1H),3.45–3.42(m,1H),3.42(s,3H),3.35(dd,J=10.1,8.9Hz,1H). 13 C NMR(150MHz,MeOD)δ168.2,144.9,137.3,134.4,130.7,130.2,119.5,101.3,75.0,73.5,71.9,71.1,65.0,55.6.HRMS(ESI):calcd.for C 16 H 19 ClO7Na[M+Na] + :381.0712, found 381.0709, 383.0677.

[0101] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-bromophenyl)acrylate (21)

[0102] White solid, 0.79 g, 19.7% yield; mp: 113–115°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1 H NMR (600MHz, MeOD) δ7.66(d,J=16.0Hz,1H),7.58–7.52(m,4H),6.58(d,J=16.0Hz,1H),4.68(d,J=3.8Hz,1H),4.50(dd,J=11.8,2.2Hz,1H),4.34(dd ,J=11.8,5.9Hz,1H),3.78(ddd,J=10.1,5.9,2.1Hz,1H),3.64(dd,J=9.6, 8.9Hz,1H),3.45–3.42(m,1H),3.41(s,3H),3.35(dd,J=10.1,8.8Hz,1H). 13 C NMR(150MHz,MeOD)δ168.2,145.0,134.8,133.2,130.9,125.5,119.6,101.3,75.0,73.5,71.9,71.1,65.0,55.6.HRMS(ESI):calcd.for C 16 H 19 BrO7Na[M+Na] + :425.0206, found 425.0201,427.0181.

[0103] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-(trifluoromethyl)phenyl)acrylate (22)

[0104] White solid, 1.03 g, 26.0% yield; mp: 99–101°C; column chromatography eluent: petroleum ether / ethyl acetate (1:1 v / v). 1H NMR (600MHz, MeOD) δ7.80(d,J=8.1Hz,2H),7.75(d,J=16.1Hz,1H),7.70(d,J=8.1Hz,2H),6.69(d,J=16.0Hz,1H),4.68(d,J=3.8Hz,1H),4.52(dd,J=11 .9,2.2Hz,1H),4.36(dd,J=11.9,5.9Hz,1H),3.80(ddd,J=10.1,5.8,2.2Hz ,1H),3.68–3.62(m,1H),3.46–3.41(m,1H),3.42(s,3H),3.41–3.33(m,1H). 13 C NMR(150MHz,MeOD)δ167.8,144.4,139.5,132.7(q,J=31.5Hz),129.7,126.9(q,J=4.5Hz),1 25.4(d,J=270Hz),121.6,101.3,75.0,73.5,71.9,71.1,65.1,55.6.HRMS(ESI):calcd.for C 17 H 19 F3O7Na[M+Na] + :415.0975, found 415.0982.

[0105] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(naphthalen-1-yl)acrylate (23)

[0106] Pale yellow solid, 0.81 g, 21.6% yield; mp: 110–112°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(600MHz,MeOD)δ8.56(d,J=15.8Hz,1H),8.16(dd,J=8.6,1.1Hz,1H),7.93(d,J=8.2Hz,1H),7.93–7.88(m,1 H),7.84(dt,J=7.2,0.9Hz,1H),7.58(ddd,J=8.4,6.8,1.4Hz,1H),7.53(ddd,J=8.0,6.8,1.2Hz,1H),7.53–7.47 (m,1H),6.61(d,J=15.7Hz,1H),4.71(d,J=3.7Hz,1H),4.57(dd,J=11.8,2.2Hz,1H),4.40(dd,J=11.8,6.0Hz,1H ),3.85–3.82(m,1H),3.67(dd,J=9.7,8.9Hz,1H),3.48–3.44(m,1H),3.44(s,3H),3.40(dd,J=10.1,8.8Hz,1H). 13 C NMR (150MHz, MeOD) δ167.0,141.7,133.8,131.3,131.1,130.5,128.5,126.7,125.9,125. 2,124.8,122.6,119.7,99.94,73.7,72.1,70.6,69.8,63.7,54.3.HRMS(ESI):calcd.for C 20 H 22 O7Na[M+Na] + :397.1258, found 397.1261.

[0107] ((2R,3S,4S,5R)-3,4,5-trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(p-tolyl)acrylate (24)

[0108] White solid, 1.53 g, 45.2% yield; mp: 155–157°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1H NMR (400MHz, MeOD) δ7.67(dd,J=16.7,2.2Hz,1H),7.47(dq,J=8.1,1.7Hz,2H),7.25–7.17(m,2H),6.47(dt,J=15.9,1.3Hz,1H),4.68(d,J=3.7 Hz,1H),4.50(dd,J=11.9,2.2Hz,1H),4.33(dd,J=11.9,5.9Hz,1H),3.79(ddd,J=10.1,5.9,2.1Hz ,1H),3.65(td,J=9.3,1.5Hz,1H),3.48–3.41(m,1H),3.41(s,3H),3.39–3.33(m,1H),2.34(s,3H). 13 C NMR(100MHz,MeOD)δ168.6,146.5,142.2,132.9,130.7,129.3,117.5,101.2,75.0,73.4,71.9,71.1,64.9,55.6,21.5.HRMS(ESI):calcd.for C 17 H 22 O7Na[M+Na] + :361.1258, found 361.1254.

[0109] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-methoxyphenyl)acrylate (25)

[0110] White solid, 1.19 g, 33.5% yield; mp: 118–120°C; column chromatography eluent: petroleum ether / ethyl acetate (1:4 v / v). 1 H NMR (400MHz, MeOD) δ7.66 (dd, J=16.0, 2.1Hz, 1H), 7.57–7.51 (m, 2H), 6.95 (dd, J= 9.4,2.3Hz,2H),6.43–6.36(m,1H),4.68(d,J=3.7Hz,1H),4.49(dd,J=11.9,2.1Hz ,1H),4.32(dd,J=11.8,5.9Hz,1H),3.82(s,3H),3.78(ddd,J=10.2,5.9,2.2Hz,1H ),3.64(td,J=9.3,1.7Hz,1H),3.45–3.42(m,1H),3.41(s,3H),3.38–3.33(m,1H). 13C NMR(150MHz,MeOD)δ168.9,163.2,146.3,131.0,128.2,115.9,115.4,101.3,75.0,73.5,71.9,71.1,64.8,55.9,55.6.HRMS(ESI):calcd.for C 17 H 22 O8Na[M+Na] + :377.1207, found 377.1207.

[0111] ((2R,3S,4S,5R)-3,4,5-trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-(allyloxy)phenyl)acrylate (26)

[0112] Pale yellow solid, 0.92 g, 24.1% yield; mp: 112–114°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1 H NMR (400MHz, MeOD) δ7.65(d,J=16.0Hz,1H),7.53(dd,J=8.7,1.7Hz,2H),6.95(dd,J=8.7,1.5Hz,2H),6.40(dd,J=1 5.9,1.0Hz,1H),6.05(dddd,J=17.6,10.5,5.5,4.7Hz,1H),5.40(dq,J=17.3,1.7Hz,1H),5.26(dq,J=10.5,1.5Hz, 1H),4.68(d,J=3.8Hz,1H),4.61–4.54(m,2H),4.49(dd,J=11.9,2.2Hz,1H),4.32(dd,J=11.9,5.9Hz,1H),3.78(dd d,J=10.1,5.9,2.2Hz,1H),3.64(t,J=9.2Hz,1H),3.44(d,J=3.7Hz,1H),3.41(s,3H),3.35(dd,J=10.1,8.9Hz,1H). 13 C NMR(100MHz,MeOD)δ168.9,162.1,146.3,134.5,131.0,128.4,117.8,116.2,116.0,101.3,75.1,73.5,71.9,71.1,69.8,64.8,55.6.HRMS(ESI):calcd.for C 19 H 24 O8Na[M+Na] +:403.1363, found 403.1357.

[0113] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(3-(allyloxy)-4-methoxyphenyl)acrylate (27)

[0114] Pale yellow solid, 0.85 g, 20.8% yield; mp: 103–105°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1 H NMR(600MHz,MeOD)δ7.63(dd,J=15.9,1.7Hz,1H),7.20(m,1H),7.17(dt,J=8.3,2.1Hz,1H),7.01–6.95(m,1H),6.40(dd,J=15.9,1.5Hz,1H),6.13– 6.03(m,1H),5.45–5.38(m,1H),5.26(m,1H),4.68(d,J=3.7Hz,1H),4.63 –4.58(m,2H),4.49(dd,J=11.8,2.2Hz,1H),4.33(dd,J=11.8,5.9Hz,1H), 3.87(s,3H),3.78(ddd,J=10.1,5.9,2.2Hz,1H),3.64(t,J=9.3Hz,1H),3.46–3.41(m,1H),3.42(s,3H),3.36(dd,J=10.0,8.9Hz,1H). 13 C NMR(150MHz,MeOD)δ167.5,152.0,148.2,145.2,133.4,127.2,123.0,116.5,114.8,11 2.3,111.5,99.9,73.7,72.1,70.5,69.8,69.6,63.4,55.0,54.2.HRMS(ESI):calcd.for C 20 H 26 O9Na[M+Na] + :433.1469, found 433.1471.

[0115] ((2R,3S,4S,5R)-3,4,5-trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-(allyloxy)-3,5-dimethoxyphenyl)acrylate (28)

[0116] Pale yellow solid, 0.99 g, 22.5% yield; mp: 102–104°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1 H NMR(600MHz,MeOD)δ7.64(d,J=15.9Hz,1H),6.92(s,2H),6.51(d,J=15.9Hz,1H),6.05(ddt,J= 17.2,10.4,6.0Hz,1H),5.29(dq,J=17.2,1.6Hz,1H),5.16(ddt,J=10.4,1.8,1.2Hz,1H),4.68 (d,J=3.7Hz,1H),4.52–4.46(m,3H),4.34(dd,J=11.9,5.7Hz,1H),3.86(s,6H),3.80–3.76(m, 1H),3.64(dd,J=9.7,8.9Hz,1H),3.45–3.43(m,1H),3.42(s,3H),3.36(dd,J=10.1,8.9Hz,1H). 13 C NMR(150MHz,MeOD)δ167.2,153.6,145.2,138.5,134.2,130.2,116.7,116.5,10 5.3,99.9,73.7,73.6,72.1,70.5,69.7,63.5,55.3,54.3.HRMS(ESI):calcd.for C 21 H 28 O 10 Na[M+Na] + :463.1575, found 463.1580.

[0117] ((2R,3S,4S,5R)-3,4,5-Trihydroxy-6-methoxytetrahydro-2H-pyran-2-yl)methyl (E)-3-(3,4-bisallyloxyphenyl)acrylate (29)

[0118] Pale yellow solid, 0.95 g, 21.7% yield; mp: 106–108°C; column chromatography eluent: petroleum ether / ethyl acetate (1:2 v / v). 1H NMR(600MHz,MeOD)δ7.62(d,J=15.9Hz,1H),7.21(d,J=2.0Hz,1H),7.14(dd,J=8.4,2.0Hz,1H),6.96 (d,J=8.4Hz,1H),6.40(d,J=15.9Hz,1H),6.08(m,2H),5.43(m,2H),5.26(m,2H),4.68(d,J=3.8Hz,1H ),4.61(m,4H),4.49(dd,J=11.9,2.2Hz,1H),4.33(dd,J=11.9,5.9Hz,1H).3.78(ddd,J=10.1,5.8,2 .2Hz,1H),3.64(dd,J=9.6,8.9Hz,1H),3.45–3.42(m,1H),3.42(s,3H),3.36(dd,J=10.1,8.9Hz,1H). 13 C NMR(150MHz,MeOD)δ167.5,150.9,148.6,145.1,133.4,133.1,127.5,122.8,116.5,116.3,11 4.9,113.4,112.8,99.9,73.7,72.1,70.5,69.8,69.6,69.3,63.4,54.3.HRMS(ESI):calcd.for C 22 H 28 O9Na[M+Na] + :459.1626, found 459.1624.

[0119] ((2R,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(((R)-5-oxotetrahydrofuran-3-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl(E)-3-(3-fluorophenyl))acrylate (30)

[0120] White solid, 1.58 g, 38.3% yield; mp: 180–182°C; column chromatography eluent: petroleum ether / ethyl acetate (1:3 v / v). 1H NMR(400MHz,MeOD)δ7.70(d,J=16.0Hz,1H),7.46–7.36(m,3H),7.15(ddq,J=9.0,5.6,2.7Hz,1H),6.61(d,J=16.0Hz,1H),4.65(ddt,J=6.5, 4.2,2.0Hz,1H),4.56(dd,J=11.9,2.2Hz,1H),4.49(dd,J=10.2,1.8Hz,1H),4.45(d,J=4.6Hz,1H),4.41(d,J=7.8Hz,1H),4.37(dd,J=11.9,6. 0Hz,1H),3.56(ddd,J=9.5,5.9,2.1Hz,1H),3.41–3.33(m,2H),3.25–3.18(m,1H),2.86(dd,J=18.0,6.5Hz,1H),2.63(dd,J=18.4,2.1Hz,1H). 13 C NMR (100MHz, MeOD) δ178.4, 168.0, 164.5 (d, J = 244Hz), 145.1 (d, J = 3Hz), 138.1 (d, J = 8Hz), 131.8 (d, J = 8Hz), 125.5 (d, J = 3Hz ),120.3,118.2(d,J=21Hz),115.3(d,J=22Hz),104.0,77.7,76.7,76.1,75.5,74.7,71.5,64.7,36.0.HRMS(ESI):calcd.for C 19 H 21 FO9Na[M+Na] + :435.1062, found 435.1068.

[0121] Comparative Example

[0122] In addition to the preparation method of Example 1 above, in order to prepare cinnamic acid derivatives 5-30, the inventors also tried two common ester condensation methods, namely, DCC and EDCI-promoted ester condensation reactions, but both failed to successfully produce the target cinnamic acid derivatives. Taking compound 5 as an example:

[0123] Method 1: EDCI-promoted ester condensation reaction: EDCI (1-ethyl-3-(3-dimethylpropylamino)carbodiimide, 0.34 mmol, 1.7 equiv.) and 4-DMAP (4-dimethylaminopyridine, 0.02 mmol, 0.1 equiv.) were added to a dry reaction flask. Anhydrous dichloromethane (4 mL) was added as solvent. The reaction flask was moved to 0°C and triethylamine (0.4 mmol, 2.0 equiv.) was slowly added. Subsequently, cinnamic acid (0.3 mmol, 1.5 equiv.) was slowly added in portions. Stirring was continued at 0°C for 15 min. MDG (0.2 mmol, 1.0 equiv.) was added and the reaction flask was moved to room temperature and stirred overnight. The next day, TLC plate was spotted, but no target product 5 was found.

[0124] Method 2: DCC-promoted ester condensation reaction: Cinnamic acid (0.24 mmol, 1.2 equiv.) was added to a dry reaction flask, a magnetic particle was added, and anhydrous dichloromethane solvent (4 mL) was added as a solvent. The mixture was stirred at 0°C, and MDG (0.2 mmol, 1.0 equiv.) was added. Subsequently, DMAP (0.02 mmol, 0.1 equiv.) and DCC (dicyclohexylcarbodiimide, 0.3 mmol, 1.5 equiv.) were added to the reaction flask in sequence. The reaction flask was moved to room temperature and continued to stir overnight. The next day, TLC plate was spotted, but no target product 5 was found.

[0125] Example 2: Determination of cytotoxic and anti-hepatic damage activities of cinnamic acid derivatives

[0126] (1) Using cell viability assay technology (CCK-8 kit), the cytotoxic effects of cinnamic acid derivatives were first evaluated. Primary immortalized mouse normal liver cell line AML12 was used as target cells. AML12 cells were treated with cinnamic acid derivatives (10, 100, 1000 μM) for 48 hours, and then the proportion of viable cells was detected by adding CCK-8 reagent. The results showed that except for compounds 21, 22, and 23, whose cell survival rate was less than 50% at a concentration of 1000 μM, the other compounds had no cytotoxic effects at all concentrations, indicating that this class of compounds is generally safe (Figure 1).

[0127] (2) C57BL / 6J mice were intraperitoneally injected with 300 mg / kg APAP to establish an acute liver injury model in mice. One hour after APAP injection, different cinnamic acid derivatives (200 mg / kg) and NAC (300 mg / kg) were intraperitoneally injected into the mice. The experiment was terminated 24 hours after APAP treatment, and relevant mouse samples were collected for relevant tests. 24 hours after APAP injection, the serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels of mice were significantly increased. Among them, the increased ALT activity indicates the degree of liver inflammatory activity, and AST indicates that liver cells have undergone necrosis, which is a recognized marker of severe liver injury. Judging from the AST results, compounds 5, 12, 13, 14, 15, 16, 18, 21, 22, 27, 28, 29, and 30 reduced the AST levels elevated by APAP treatment to varying degrees. Compounds 12, 16, 22, 28, and 29 also reduced the ALT levels elevated by APAP treatment to varying degrees. Compounds 16 and 28, in particular, exhibited a potent protective effect against APAP-induced acute liver injury, as evidenced by the simultaneous reduction of ALT and AST levels, even approaching that of the positive control, NAC. This demonstrates the therapeutic potential of compounds 16 and 28 against APAP-induced acute liver injury (Figure 2). In the figure, the "Control" group represents normal mice, the "APAP" group represents the model group, and the "NAC" group represents the positive control group.

[0128] The inventors of the present invention previously reported on a cinnamic acid derivative (compound MCGP) as shown in Formula 18. It is primarily used to treat acetaminophen (APAP)-induced drug-induced liver injury and carbon tetrachloride-induced chemical liver injury. Its mechanism of action may be related to inhibiting oxidative stress, inhibiting hepatocyte apoptosis, and promoting liver regeneration (reference: Front. Pharmacol. 2022, 13, 873938). The administration method described in that paper differs significantly from that of the present invention: MCGP was used for pre-administration 10 days before establishing an ALI mouse model, whereas the present invention administers the cinnamic acid derivative (Formulas 1A–1B) after ALI modeling has already occurred in the mice. This experimental protocol is more consistent with clinical practice, as most ALI patients in clinical practice are unlikely to anticipate the onset of ALI and therefore do not take medication in advance. More importantly, in the experiments of the present invention, the anti-hepatic injury effect of MCGP was weak, while several compounds described in the present invention (such as compounds 5, 12, 13, 14, 15, 16, 21, 22, 27, 28, 29, 30) showed stronger anti-hepatic injury effects than MCGP.

[0129] Example 3: Determination of the effects of representative compound 16 (denoted as CK16) and compound 28 (denoted as CG28) on liver injury by inhibiting MAPK signaling pathway activation, inhibiting ferroptosis, and activating UDP-glucuronyl transferase

[0130] (1) To confirm the effects of CK16 and CG28 on APAP-induced acute liver injury, mice were treated with different concentrations of CK16 and CG28 (100, 200, and 300 mg / kg) 1 hour after intraperitoneal injection of 300 mg / kg APAP (see Figure 3A). The solvent was 2% DMSO and 98% corn oil (volume ratio: 2:98) as a control. The administration concentration of NAC was explored and it was found that only at 300 mg / kg, NAC significantly alleviated the increase in serum ALT and AST induced by APAP. Therefore, we used 300 mg / kg NAC as a positive reference in the following experiments (see Figure 3B). Intraperitoneal injection of CK16 and CG28 1 hour after APAP injection significantly reduced the increase in ALT and AST levels in mice caused by APAP (see Figure 3C). H&E stained histology showed that CK16 and CG28 significantly improved APAP-induced liver hemorrhage, fatty degeneration, hepatocyte swelling and necrosis (see Figure 3D and E). Furthermore, immunofluorescence staining of liver specimens revealed that the number of TUNEL-positive cells in mice treated with CK16 and CG28 was significantly lower than that in the APAP-treated group. CK16 and CG28 treatment significantly inhibited the translocation of HMGB1 to the cytoplasm in APAP-treated mice. Immunochemical staining of mouse liver tissues revealed that p-MLKL expression was significantly reduced after CK16 and CG28 administration compared with the APAP model group (Figure 3, F). We also examined the effects of CK16 and CG28 on APAP-induced mortality by intraperitoneal injection of a lethal dose of APAP (600 mg / kg). Mouse survival was monitored every 4 hours for 24 hours after administration. During this observation period, CK16 and CG28 administration significantly improved mouse survival, and the survival rate of mice in the CK16 and CG28-treated groups was significantly higher than that in the NAC group, demonstrating that CK16 and CG28 mice exhibited a stronger mitigation effect against APAP-induced liver injury than NAC (Figure 3, G). Taken together, these results indicate that CK16 and CG28 can effectively ameliorate APAP-induced acute liver injury in mice.

[0131] (2) Furthermore, the anti-liver injury effects of CK16 and CG28 were tested using an oxaliplatin (OXA)-induced liver injury model. OXA was dissolved in 5% glucose solution (mass percentage) and injected intraperitoneally at 8 mg / kg into mice to induce an acute liver injury model. CK16 and CG28 were dissolved in 2% DMSO and 98% corn oil (volume ratio: 2:98) and then injected intraperitoneally at a dose of 100 mg / kg 1 hour after OXA treatment. NAC was dissolved in normal saline and injected intraperitoneally at a dose of 300 mg / kg 1 hour after OXA treatment. The experiment lasted for three days, with administration once a day. After three days of OXA action, the liver and blood were collected for further experiments. The results showed that intraperitoneal injection of CK16 and CG28 could significantly reduce the increase in ALT and AST levels in mice caused by OXA (see Figure 4A), indicating that CK16 and CG28 have a certain protective effect on OXA-induced acute liver injury in mice. A liver injury model was established by treating the AML12 cell line with OXA (35 μM). AML12 cells were then co-treated with CK16 and CG28 (6.25, 12.5, 25, 50, and 100 μM) for 24 hours. The NAC (10 mM) treatment group served as a positive control. CCK-8 reagent was then added to measure the proportion of viable cells. The results showed that CK16 and CG28 exhibited a modest ameliorative effect on the OXA-induced in vitro cell liver injury model (see Figure 4, B). These results demonstrate that the cinnamic acid derivatives CK16 and CG28 possess anti-oxaliplatin-induced liver injury activity.

[0132] (3) An APAP-induced liver injury model was used to evaluate the inhibitory effects of cinnamic acid derivatives CK16 and CG28 on MAPK signaling pathway activation and ferroptosis in vivo.

[0133] 24 hours after receiving APAP, the MAPK family proteins (P38, ERK1 / 2, and JNK) in the liver tissue of the mice in the APAP model group were significantly phosphorylated, indicating the activation of this pathway, while the phosphorylation of MAPK family proteins in the CK16 or CG28 treatment groups was significantly inhibited (see Figure 5).

[0134] Immunohistochemical analysis showed that 4-HNE was significantly upregulated in the APAP model group, while CK16 or CG28 could significantly prevent this upregulation (see Figure 6A), indicating that CK16 and CG28 can reduce lipid accumulation and the occurrence of ferroptosis in the liver of APAP-treated mice. In addition, CK16 or CG28 can significantly reduce the accumulation of iron ions (see Figure 6B). The expression of ferroptosis-related genes was detected by qRT-PCR. CK16 and CG28 significantly changed the expression of ferroptosis-related genes. Specifically, the expression of FTH1, DHODH, FGF21, SLC7A11, FTL, DHFR, FPN and GCH1 was increased. These genes are negative regulators of iron oxidation (see Figure 6C). Immunoblotting results showed that compared with the APAP model group, CK16 and CG28 increased the expression levels of GPX4 and FSP1, while the expression levels of these two proteins were basically similar to those of the control group mice. Consistent with the qRT-PCR results, CK16 and CG28 also significantly increased the expression levels of other proteins negatively correlated with ferroptosis in the liver, such as SLC7A11, FTL, FTH1, FPN, and CD71 (see Figure 6D). These results indicate that CK16 and CG28 can alleviate APAP-induced hepatocyte ferroptosis and improve liver damage.

[0135] (4) The activation effect of cinnamic acid derivatives CK16 and CG28 on UGT enzyme activity was detected using a UGT enzyme activity assay kit. The present invention first used HPLC-MS / MS to analyze the APAP metabolites of each experimental group, including APAP-Cys, APAP-Sulf and APAP-Gluc. The detection was completed by a CRO company (Wuhan Hongren Biopharmaceutical Co., Ltd.) that specializes in providing drug development technology services. The detection of UDP-glycosyltransferase is currently difficult. The present invention used a UGT activity assay kit (Abcam, UK, ab273331) to determine the effect of the compound on UGT activity. The results showed that CK16 and CG28 can enhance the activity of UGT enzymes in vitro (see A in Figure 7). We collected mouse blood 2 hours after APAP injection (1 hour after compound injection) to detect changes in APAP metabolites. The results showed that the APAP-cys level in the CK16 or CG28 treatment group was significantly lower than that in the APAP model group, and the APAP-cys level in the NAC treatment group was higher. Meanwhile, treatment with CK16 or CG28 significantly increased APAP-gluc levels in mice but had little effect on APAP-sulf. NAC treatment had no effect on APAP-gluc levels in mice but significantly reduced APAP-sulf levels (see Figure 7, B). The APAP-induced liver injury model confirmed that the cinnamic acid derivatives CK16 and CG28 can activate UGT enzyme activity in vivo.

[0136] Results and Analysis:

[0137] The new cinnamic acid derivatives synthesized by the present invention have stable structures and are non-cytotoxic within the effective concentration range. In a cell and two mouse acute liver injury models, multiple cinnamic acid derivatives showed a strong effect in alleviating acute liver injury. Among them, the representative compounds CK16 and CG28 had stronger anti-liver injury effects than the positive drug N-acetyl-L-cysteine ​​(NAC). The representative compounds CK16 and CG28 can exert their effects by inhibiting the activation of the MAPK signaling pathway and ferroptosis response in the body and activating UDP-glucuronyl transferase.

[0138] Although Example 3 verifies the anti-liver injury effects of representative compounds CK16 and CG28 by inhibiting MAPK signaling pathway activation, inhibiting ferroptosis reaction, and activating UDP-glucuronyl transferase, it is not limited to the treatment of liver injury. The present invention can also be applied to other diseases that have similar mechanisms of action to exert therapeutic effects (it can be used alone or in combination with other drugs). This is because MAPK signaling pathway activation and ferroptosis are common stages of many diseases in organisms, and play a role in the occurrence, development, and drug treatment of many diseases. UDP-glucuronyl transferase is one of the most important enzymes in human phase II metabolism, responsible for approximately 40-70% of endogenous and heterologous reactions, making the compound after the binding reaction more water-soluble and easy to be excreted from the body. It participates in the metabolic clearance process of many drugs, thereby playing a detoxifying role, and the associated physiological and pathological processes are also relatively wide.

[0139] While Example 3 only addresses representative compounds CK16 and CG28, other cinnamic acid derivatives satisfying Formulas 1A and 1B can also mitigate APAP-induced liver injury to varying degrees. This is because they are structurally similar to CK16 and CG28, sharing the same glycoside-cinnamoyl backbone. The hydroxyl group within the glycoside structure and the carbonyl group within the cinnamoyl structure are key to the anti-ALI effects of these compounds. Furthermore, reducing the time interval between APAP induction and administration can enhance their therapeutic effects on liver injury.

[0140] It will be easily understood by those skilled in the art that the above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A cinnamic acid derivative, characterized in that, Its structural formula is shown in Formula 1A or Formula 1B as follows: And exclusion formula 18: for formula 1A or formula 1B: the substituent R is from 0 to 5; when the substituent R is from 1 to 5, the substitution sites correspond to any 1 to all 5 of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; the Rs at different substitution sites are independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, trifluoromethoxy, 2,3-phenyl, methyl, methoxy, ester group, carboxyl group, hydroxyl group, allyloxy group; the solid and dashed lines represent a carbon-carbon single bond or a carbon-carbon double bond.

2. The cinnamic acid derivative according to claim 1, characterized in that, The structural formula of the cinnamic acid derivative is specifically any one of Formula 5 to Formula 17 and Formula 19 to Formula 30:

3. The method for synthesizing the cinnamic acid derivative according to claim 1 or 2, characterized in that, comprises the following steps: (1) Using cinnamic acid as shown in Formula 3A as a reaction substrate, a substitution reaction occurs with oxalyl chloride under the action of a catalytic amount of DMF to obtain cinnamoyl chloride as shown in Formula 4A; In Formula 3A, the substituent R 1 is from 0 to 5; when the substituent R 1 is from 1 to 5, the substitution sites correspondingly are any 1 to all 5 of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; R at different substitution sites 1 is independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, ester group; the solid and dashed lines represent a carbon-carbon single bond or a carbon-carbon double bond; (2) React the cinnamoyl chloride shown in Formula 4A obtained in step (1) with compound KD to produce a corresponding cinnamic acid derivative, and the structural formula of this cinnamic acid derivative satisfies Formula 1A; or react the cinnamoyl chloride shown in Formula 4A obtained in step (1) with compound MDG to produce a corresponding cinnamic acid derivative, and the structural formula of this cinnamic acid derivative satisfies Formula 1B; 4. The method for synthesizing the cinnamic acid derivative according to claim 1 or 2, characterized in that, comprises the following steps: S1: Under alkaline conditions, cinnamic acid as shown in the formula Cinnamic acids undergoes a substitution reaction with allyl bromide to form a cinnamate as shown in the formula 2; then, the cinnamate as shown in the formula 2 is hydrolyzed under alkaline conditions and then acidified to obtain cinnamic acid as shown in the formula 3B; In the formula Cinnamic acids, the number of hydroxyl substituents is from 1 to 5, and the substitution sites correspond to any 1 to all 5 of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; the substituent R 2 is from 0 to 4, and the sum of the number of hydroxyl substituents and the substituent R 2 does not exceed 5; when the substituent R 2 is non-zero, the substitution site of the substituent R 2 corresponds to any 1 or several of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring except the hydroxyl substitution sites; and, R 2 at different substitution sites are independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, oxytrifluoromethyl, 2,3-phenyl, methyl, methoxy, ester group; the solid and dashed lines represent a carbon-carbon single bond or a carbon-carbon double bond; S2: Use the cinnamic acid shown in Formula 3B obtained in Step S1 as a reaction substrate, and carry out a substitution reaction with oxalyl chloride under the action of a catalytic amount of DMF to obtain cinnamoyl chloride shown in Formula 4B; S3: React the cinnamoyl chloride shown in Formula 4B obtained in Step S2 with compound KD to form a corresponding cinnamic acid derivative, and the structural formula of this cinnamic acid derivative satisfies Formula 1A; or react the cinnamoyl chloride shown in Formula 4B obtained in Step S2 with compound MDG to form a corresponding cinnamic acid derivative, and the structural formula of this cinnamic acid derivative satisfies Formula 1B; 5. Use of a cinnamic acid derivative in the preparation of a drug that exerts a therapeutic effect by inhibiting the MAPK signaling pathway, characterized in that, The structural formula of the cinnamic acid derivative is shown in Formula 1A or Formula 1B as follows: for formula 1A or formula 1B: the substituent R is from 0 to 5; when the substituent R is from 1 to 5, the substitution sites correspond to any 1 to all 5 of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; the Rs at different substitution sites are independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, trifluoromethoxy, 2,3-phenyl, methyl, methoxy, ester group, carboxyl group, hydroxyl group, allyloxy group; the solid and dashed lines represent a carbon-carbon single bond or a carbon-carbon double bond.

6. Use of a cinnamic acid derivative in the preparation of a drug that exerts a therapeutic effect by inhibiting ferroptosis, characterized in that, The structural formula of the cinnamic acid derivative is shown in Formula 1A or Formula 1B as follows: for formula 1A or formula 1B: the substituent R is from 0 to 5; when the substituent R is from 1 to 5, the substitution sites correspond to any 1 to all 5 of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; the Rs at different substitution sites are independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, trifluoromethoxy, 2,3-phenyl, methyl, methoxy, ester group, carboxyl group, hydroxyl group, allyloxy group; the solid and dashed lines represent a carbon-carbon single bond or a carbon-carbon double bond.

7. Use of a cinnamic acid derivative in the preparation of a drug that exerts a therapeutic effect by activating UDP-glucuronosyltransferase, characterized in that, The structural formula of the cinnamic acid derivative is shown in Formula 1A or Formula 1B as follows: for formula 1A or formula 1B: the substituent R is from 0 to 5; when the substituent R is from 1 to 5, the substitution sites correspond to any 1 to all 5 of the 2nd, 3rd, 4th, 5th, and 6th positions of the benzene ring; the Rs at different substitution sites are independently selected from: fluorine, chlorine, bromine, iodine, trifluoromethyl, trifluoromethoxy, 2,3-phenyl, methyl, methoxy, ester group, carboxyl group, hydroxyl group, allyloxy group; the solid and dashed lines represent a carbon-carbon single bond or a carbon-carbon double bond.

8. Use of the cinnamic acid derivative according to claim 1 or 2 in the preparation of a drug for treating liver injury.

9. The use according to claim 8, characterized in that, the drug for treating liver injury is a drug for treating acute liver injury.

Images