4-alkoxyl substituted 2,6-dihydroxybenzoic acid esters of d-limonene or fenchol and their pharmaceutical use

Abstract

4-alkoxy-substituted 2,6-dihydroxybenzoic acid ester compound of d-camphor or fenchol, the structure conforms to general formula (I) wherein: R 1 is or R 2 is -H, -NO2; R 3 alkyl of 1-4 carbon atoms or the compound has good anti-inflammatory effect, and can be used for preparing drugs for treating inflammation-related diseases or stroke injury, cerebral edema, senile dementia.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of pharmacy, and provides a class of 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right camphor or fenchol ester compounds and pharmaceutically acceptable salts thereof and their medical uses. BACKGROUND

[0002] Stroke seriously endangers human health. Right camphor ((+)-borneol) has a clear anti-cerebral ischemia effect (Journal of Biomedical Research, 2017, 31:306-314), and its mechanism of action may be related to its inhibition of inflammatory response. In the process of inflammatory response, macrophages play an important role in starting, maintaining and resolving inflammatory response. Lipopolysaccharide (LPS) is one of the main components of the cell wall of gram-negative bacteria, and has strong immune stimulating ability. RAW264.7 macrophages, as a kind of murine immune cells, will be activated when stimulated by external factors (such as LPS), and secrete a large number of inflammatory factors (such as NO, etc.), resulting in inflammatory response. The amount of inflammatory factors can indirectly reflect the severity of inflammation, and is a quantitative indicator of the severity of inflammation. Right camphor can inhibit the increase of NO in the LPS-induced RAW264.7 macrophage inflammation model. However, right camphor has low oral bioavailability, and when it is made into an injection, it is difficult to dissolve in water, and a large amount of organic solvent needs to be added, which increases the difficulty of drug preparation and the risk of clinical medication.

[0003] Chinese invention patent 2021113312181 discloses 2,6-dihydroxybenzoic acid right camphor or fenchol ester compounds and their medical uses, the structure of which conforms to the general formula wherein: R = -H, -OH, -NR 1 R 2 , or -CONR 3 R 4 ; R 1 , R 2 = -H, acyl of 1-4 carbon atoms or alkyl of 1-6 carbon atoms, or -COR 5 ; R 3 , R 4 , R 5 = -H or alkyl of 1-4 carbon atoms. Such drugs show good efficacy in reducing stroke damage.

[0004] Among them: Chinese invention patent 2021113312181 embodiment compound 2 (2, 4, 6-trihydroxybenzoic acid d-camphor alcohol ester, control compound 1) has good efficacy in reducing stroke damage, the applicant alkylates the 4-position hydroxyl group, and unexpectedly finds that the ester compound of 4-alkoxy substituted 2, 6-dihydroxybenzoic acid d-camphor alcohol or fenchol has good inhibitory effect on inflammation, and the drug efficacy is significantly higher than that of control compound 1 (see examples 2, 3). On this basis, the introduction of nitro group at its 3 position can further increase the drug efficacy (see examples 2, 3).

[0005] Cerebral edema is a non-specific pathological swelling of the brain, which can develop gradually in a focal or diffuse manner after any type of neurological injury. Cerebral edema can be secondary to blood-brain barrier disruption, local inflammation, vascular changes or cellular metabolic changes. Cerebral edema is considered one of the most common causes of elevated intracranial pressure (ICP), which has been identified as a predictor of poor outcome in patients with traumatic brain injury (TBI), stroke and other intracranial lesions. Cerebral edema secondary to cerebral hemorrhage, especially vasogenic cerebral edema, is considered a central link in post-hemorrhagic injury and one of the main causes of exacerbation of the patient's condition and death. Brain tissue will also develop cerebral edema under ischemic and hypoxic conditions. Reperfusion causes secondary injury, and the water content in the brain increases rapidly, forcing the brain volume to increase, which increases the risk of ischemic stroke. Cell toxicity edema occurs in the early stage of ischemia, and then endothelial cells swell, the tight junction structure between endothelial cells is damaged, the blood-brain barrier permeability changes, and vasogenic edema is formed, and the production of toxic products and inflammatory mediators further aggravates the edema.

[0006] Fenchol can help protect the brain from Alzheimer's disease pathology. Fenchol significantly reduces excess Aβ accumulation and neuronal death by stimulating FFAR2 signaling, a gut microbiome sensing mechanism. Zombie cells in the brains of people with Alzheimer's disease stop replicating and slowly die, accumulate in diseased and aging organs, create a destructive inflammatory environment, and send stress or death signals to neighboring healthy cells, which eventually also become harmful zombie cells or die (Frontiers in Aging Neuroscience, 2021). However, fenchol has low oral bioavailability, and when made into an injection, it is difficult to dissolve in water, requiring the addition of a large amount of organic solvent, which increases the difficulty of drug preparation and the risk of clinical use. SUMMARY

[0007] To solve the above technical problems in the prior art, the present application provides a kind of 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound and its pharmaceutically acceptable salt and its pharmaceutical use.The drugs have good anti-inflammatory effect, and can be used for preparing drugs for treating inflammation-related diseases and / or stroke injury, cerebral edema and senile dementia.

[0008] The technical scheme of the present application is as follows:

[0009] The first object of the present application is to provide a 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound, which has a structure according to general formula (I) wherein: R 1 is R 2 is -H, -NO2;

[0010] R 3 is an alkyl group with 1-4 carbon atoms or Further, wherein: R 3 is an alkyl group with 1 carbon atom.

[0011] Further, the structure is any of the following:

[0012]

[0013]

[0014] The second object of the present application is to provide a pharmaceutically acceptable salt of the aforementioned 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound.

[0015] The third object of the present application is to provide a pharmaceutical composition comprising the aforementioned 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound or the pharmaceutically acceptable salt of the aforementioned 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound.

[0016] The fourth object of the present application is to provide the use of the aforementioned 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound or the pharmaceutically acceptable salt of the aforementioned 4-alkoxy-substituted 2,6-dihydroxybenzoic acid right clonal or fenchol ester compound or the aforementioned pharmaceutical composition in the preparation of a drug for treating inflammation-related diseases and / or stroke injury, cerebral edema and senile dementia.

[0017] The fifth object of the present application is to provide a drug for treating inflammation-related diseases and / or stroke injury, cerebral edema, and senile dementia, the effective component of which comprises the aforementioned ester compound of 4-alkoxy-substituted 2,6-dihydroxybenzoic acid with fenchyl alcohol or a pharmaceutically acceptable salt of the aforementioned ester compound of 4-alkoxy-substituted 2,6-dihydroxybenzoic acid with fenchyl alcohol.

[0018] The present application has the following advantages over the prior art:

[0019] The compound of the present application has good anti-inflammatory and stroke injury-reducing effects, and can be used for preparing a drug for treating inflammation-related diseases or stroke injury. It is to be noted that in a cell model, the drug efficacy of the ester compound of fenchyl alcohol of the present application is significantly better than that of the control compound 1, and the drug efficacy of the ester compound of fenchyl alcohol of the present application is significantly better than that of fenchyl alcohol. In an acute inflammation model induced by carrageenan, the ester compound of fenchyl alcohol of the present application has a significantly better inhibitory effect on rat foot swelling caused by carrageenan than the control compound 1. It is suggested that etherification of the hydroxyl group at the 4-position plays an important role in improving the anti-inflammatory effect of the compound. In a stroke MCAO model, the compound of the present application shows good therapeutic effect. It is suggested that the compound has a good application prospect in preparing a drug for treating inflammation-related diseases or stroke injury.

[0020]

[0021] Control compound 1. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 Inhibitory effect of the target compound of fenchyl alcohol on the increase of IL-1β and TNF-α in a RAW264.7 macrophage inflammation model induced by lipopolysaccharide;

[0023] Figure 2 Inhibitory effect of the target compound of fenchyl alcohol on the increase of IL-1β and TNF-α in a RAW264.7 macrophage inflammation model induced by lipopolysaccharide;

[0024] Figure 3 Inhibitory effect of the target compound 1 on rat foot swelling caused by carrageenan;

[0025] Figure 4 Effect of the test substance on the symptoms of neurological defects;

[0026] Figure 5 Effect of the test substance on the cerebral infarction area (%);

[0027] Figure 6 Effect of the test substance on the water content (%) of brain tissue. DETAILED DESCRIPTION

[0028] The present application will be further explained by the following examples, but the examples do not limit the present application in any form.

[0029] Synthesis of the target compound of Example 1

[0030] 1) Synthesis of dextro-menthyl 2,6-dihydroxy-4-methoxybenzoate (Target compound 1)

[0031] Synthesis route:

[0032]

[0033] Synthesis method:

[0034] Dextro-menthyl 2,4,6-trihydroxybenzoate 1.53 g (5 mmol) was dissolved in 5 mL of dichloromethane, and 1,8-diazabicycloundec-7-ene (DBU, 10 mmol) 1.52 g was added dropwise. After stirring for 5 min, dimethyl sulfate (DMS, 5 mmol) 0.63 g was slowly added dropwise, and the reaction was allowed to proceed at room temperature for 2 h. After the completion of the reaction, 50 mL of ethyl acetate was added, and the organic layer was collected and dried. The product was purified by silica gel chromatography (petroleum ether: ethyl acetate = 100:1) to obtain a white powder at a yield of 67%. 1 H NMR (400 MHz, DMSO-d6) δ (ppm): δ 5.92 (s, 2H), 4.97 (d, J = 9.6 Hz, 1H), 3.67 (s, 3H), 2.37-2.26 (m, 1H), 2.13 (td, J = 10.0, 9.6, 5.4 Hz, 1H), 1.73-1.62 (m, 2H), 1.24 (dd, J = 31.3, 11.2 Hz, 2H), 1.04 (dd, J = 13.7, 3.6 Hz, 1H), 0.87 (s, 3H), 0.83 (d, J = 6.8 Hz, 6H). 13 C NMR (101 MHz, DMSO-d6) δ 170.78, 164.79, 162.32, 97.28, 93.85, 81.13, 55.78, 49.12, 47.86, 44.80, 36.97, 28.09, 27.48, 20.07, 19.22, 13.96.

[0035] 2) Synthesis of dextro-menthyl 2,6-dihydroxy-4-ethoxybenzoate (Target compound 2): The synthesis was performed according to the synthesis method of target compound 1, using dextro-menthyl 2,4,6-trihydroxybenzoate and diethyl sulfate as starting materials, and a white solid was obtained.

[0036] 1H NMR (400 MHz, DMSO-d6) δ 5.89 (s, 2H), 4.96 (ddd, J = 10.0, 3.5, 2.0 Hz, 1H), 3.94 (q, J = 7.0 Hz, 2H), 2.32 (ddt, J = 13.5, 9.5, 3.7 Hz, 1H), 2.13 (td, J = 10.1, 9.2, 5.5 Hz, 1H), 1.67 (dt, J = 14.1, 4.2 Hz, 2H), 1.27 - 1.25 (m, 3H), 1.23 - 1.20 (m, 2H), 1.03 (dd, J = 13.7, 3.5 Hz, 1H), 0.89 - 0.80 (m, 9H). 13 C NMR (101 MHz, DMSO-d6) δ 170.82, 164.10, 162.36, 97.06, 94.21, 81.12, 63.78, 49.11, 47.86, 44.80, 36.97, 28.09, 27.48, 20.07, 19.22, 14.97, 13.95.

[0037] 3) Synthesis of dextro-menthyl 2,6-dihydroxy-4-isopropoxybenzoate (Target compound 3):

[0038] To dextro-menthyl 2,4,6-trihydroxybenzoate 1.53 g (5 mmol) was dissolved in 5 ml of acetonitrile, potassium carbonate (6 mmol) and potassium iodide (1 mmol) were added, after stirring for 5 min, 2-chloropropane (7.5 mmol) was added dropwise, and it was reacted at 60°C for 5 h. After the reaction was completed, 50 ml of ethyl acetate was added, washed with saturated brine, and the organic layer was collected and dried, and purified by silica gel chromatography (petroleum ether: ethyl acetate = 100: 1) to obtain a white powder at a yield of 57%. 1 H NMR (400 MHz, DMSO-d6) δ 5.88 (s, 2H), 4.96 (ddd, J = 9.9, 3.5, 2.1 Hz, 1H), 4.51 (p, J = 6.1 Hz, 1H), 2.32 (ddt, J = 13.8, 9.7, 3.9 Hz, 1H), 2.14 (ddd, J = 12.8, 8.3, 3.2 Hz, 1H), 1.73 - 1.63 (m, 2H), 1.23 - 1.18 (m, 8H), 1.03 (dd, J = 13.7, 3.6 Hz, 1H), 0.87 (s, 3H), 0.83 (d, J = 6.8 Hz, 6H). 13C NMR (101 MHz, DMSO-d6) δ 170.80, 163.15, 162.43, 96.81, 95.00, 81.10, 70.03, 49.11, 47.85, 44.79, 36.98, 28.09, 27.48, 22.40, 22.24, 20.06, 19.21, 18.12, 13.94.

[0039] 4) Synthesis of dextroveratryl 2,6-dihydroxy-4-isobutyloxybenzoate (Target Compound 4):

[0040] Reference to the synthesis method of Target Compound 3, synthesized with dextroveratryl 2,4,6-trihydroxybenzoate and 1-chloro-2-methylpropane as raw materials, white solid. 1 H NMR (400 MHz, DMSO-d6) δ 5.90 (s, 2H), 4.96 (ddd, J = 9.9, 3.6, 1.8 Hz, 1H), 3.66 (d, J = 6.5 Hz, 2H), 2.32 (ddt, J = 13.7, 9.5, 4.0 Hz, 1H), 2.14 (ddd, J = 12.7, 9.3, 3.3 Hz, 1H), 1.94 (dq, J = 13.2, 6.6 Hz, 1H), 1.72 - 1.63 (m, 2H), 1.29 - 1.17 (m, 2H), 1.08 - 0.99 (m, 1H), 0.91 (s, 3H), 0.90 (s, 3H), 0.87 (s, 3H), 0.84 (s, 3H), 0.82 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 170.80, 163.15, 162.43, 96.81, 95.00, 81.10, 70.03, 49.11, 47.85, 44.79, 36.98, 28.09, 27.48, 22.40, 22.24, 20.06, 19.21, 18.12, 13.94.

[0041] 5) Synthesis of dextroveratryl 2,6-dihydroxy-4-benzyloxybenzoate (Target Compound 5): Reference to the synthesis method of Target Compound 3, synthesized with dextroveratryl 2,4,6-trihydroxybenzoate and chlorobenzyl as raw materials, white solid. 1H NMR (400 MHz, Chloroform-d) δ 7.41 - 7.25 (m, 5H), 6.12 (s, 2H), 5.25 (dt, J = 9.9, 3.2 Hz, 1H), 5.05 (s, 2H), 2.52 (ddt, J = 14.1, 10.1, 3.9 Hz, 1H), 1.79 (tdt, J = 25.5, 9.4, 4.1 Hz, 3H), 1.53 (dddd, J = 14.1, 12.3, 4.8, 2.2 Hz, 1H), 1.31 (ddd, J = 13.4, 7.0, 3.2 Hz, 1H), 1.19 (dd, J = 14.2, 3.6 Hz, 1H), 0.98 - 0.91 (m, 9H). 13 C NMR (101 MHz, Chloroform-d) δ 170.04, 165.48, 136.11, 128.78, 128.33, 127.59, 95.24, 83.61, 70.14, 49.05, 48.15, 44.66, 37.05, 28.19, 28.09, 19.83, 18.97, 13.75.

[0042] 6) Synthesis of dextromethorphan 2,6-dihydroxy-3-nitro-4-methoxybenzoate (Target compound 6):

[0043] Synthetic route:

[0044]

[0045] Synthetic method:

[0046] Dextromethorphan 2,6-dihydroxy-4-methoxybenzoate (target compound 1) (1.00 mmol) was dissolved in 2 ml of acetic anhydride, and a mixture of concentrated nitric acid and acetic anhydride (volume ratio = 1:5) was slowly added dropwise under ice bath conditions until the solution color turned orange yellow. After 5 min, water was added to quench, and the aqueous phase was extracted with 50 ml of ethyl acetate, washed with 100 ml of saturated brine three times, and the organic layer was collected and dried. Purification by silica gel chromatography (petroleum ether: ethyl acetate = 100:1) gave a light yellow powder with a yield of 72%. 1H NMR (400 MHz, Chloroform-d) δ 6.10 (s, 1H), 5.26 (dt, J = 9.7, 3.0 Hz, 1H), 3.91 (s, 3H), 2.52 (ddt, J = 14.0, 9.9, 4.0 Hz, 1H), 1.81 (ddd, J = 16.4, 7.8, 4.0 Hz, 2H), 1.49 (dd, J = 15.1, 3.3 Hz, 1H), 1.36 - 1.24 (m, 1H), 1.17 (dd, J = 14.1, 3.6 Hz, 1H), 0.97 - 0.91 (m, 9H). 13 C NMR (101 MHz, DMSO-d6) δ 169.62, 162.49, 155.94, 154.50, 124.67, 98.18, 92.25, 81.97, 57.14, 49.17, 44.75, 36.72, 28.02, 27.42, 20.04, 19.19, 13.94.

[0047] 7) Synthesis of dextromethorphan 2,6-dihydroxy-3-nitro-4- isopropoxybenzoate (Target compound 7): Refer to the synthesis method of target compound 6, starting from dextromethorphan 2,6-dihydroxy-4-isopropoxybenzoate (target compound 3) to synthesize, light yellow powder. 1 H NMR (400 MHz, DMSO-d6) δ 6.19 (s, 1H), 4.99 (dt, J = 9.7, 2.9 Hz, 1H), 4.61 (hept, J = 6.0 Hz, 1H), 2.32 (ddt, J = 13.8, 9.8, 3.9 Hz, 1H), 2.09 (ddd, J = 12.9, 9.2, 4.1 Hz, 1H), 1.73 - 1.63 (m, 2H), 1.28 - 1.17 (m, 8H), 1.08 (dd, J = 13.7, 3.4 Hz, 1H), 0.89 - 0.81 (m, 9H). 13 C NMR (101 MHz, DMSO-d6) δ 169.77, 162.33, 154.64, 154.32, 125.62, 97.67, 93.69, 81.99, 72.83, 49.16, 47.91, 44.76, 36.73, 28.02, 27.43, 21.94, 20.04, 19.19, 13.92.

[0048] 8) Synthesis of dextromethorphan 2,6-dihydroxy-3-nitro-4- isobutoxybenzoate (Target compound 8): Refer to the synthesis method of target compound 6, starting from dextromethorphan 2,6-dihydroxy-4-isobutoxybenzoate (target compound 4) to synthesize, light yellow powder. 1H NMR (400 MHz, DMSO-d6) δ 6.20 (s, 1H), 4.97 (ddd, J = 9.9, 3.6, 1.8 Hz, 1H), 3.66 (d, J = 6.5 Hz, 2H), 2.33 (ddt, J = 13.7, 9.5, 4.0 Hz, 1H), 2.15 (ddd, J = 12.7, 9.3, 3.3 Hz, 1H), 1.95 (dq, J = 13.2, 6.6 Hz, 1H), 1.73 - 1.63 (m, 2H), 1.30 - 1.17 (m, 2H), 1.09 - 0.99 (m, 1H), 0.91 (s, 3H), 0.90 (s, 3H), 0.87 (s, 3H), 0.84 (s, 3H), 0.82 (s, 3H).

[0049] 9) Synthesis of 2,6-dihydroxy-3-nitro-4-benzyloxybenzoic acid veryl ester (Target compound 9): Refer to the synthesis method of target compound 6, and synthesize from 2,6-dihydroxy-4-benzyloxybenzoic acid veryl ester (target compound 5) as a raw material, a light yellow powder. 1 H NMR (400 MHz, DMSO-d6) δ 6.19 (s, 1H), 5.26 (dt, J = 9.9, 3.2 Hz, 1H), 5.06 (s, 2H), 2.53 (ddt, J = 14.1, 10.1, 3.9 Hz, 1H), 1.79 (tdt, J = 25.5, 9.4, 4.1 Hz, 3H), 1.53 (dddd, J = 14.1, 12.3, 4.8, 2.2 Hz, 1H), 1.31 (ddd, J = 13.4, 7.0, 3.2 Hz, 1H), 1.19 (dd, J = 14.2, 3.6 Hz, 1H), 0.98 - 0.91 (m, 9H).

[0050] 10) Synthesis of 2,6-dihydroxy-4-methoxybenzoic acid fenchyl ester (target compound 10):

[0051] Synthesis route:

[0052]

[0053] Synthesis method:

[0054] To 2,4,6-trihydroxybenzoic acid fenchyl ester 1.53 g (5 mmol) dissolved in 5 mL of dichloromethane, 1,8-diazabicycloundec-7-ene (DBU, 10 mmol) 1.52 g was added dropwise, and after stirring for 5 min, dimethyl sulfate (DMS, 5 mmol) 0.63 g was slowly added dropwise, and the reaction was allowed to proceed at room temperature for 2 h. After the reaction was completed, 50 mL of ethyl acetate was added, and the organic layer was collected and dried. The solvent was removed under reduced pressure, and the product was purified by silica gel chromatography (petroleum ether: ethyl acetate = 100:1) to obtain a white solid. 1 HNMR (400 MHz, DMSO-d6) δ 5.92 (s, 2H), 4.49 (d, J = 1.8 Hz, 1H), 3.67 (s, 3H), 1.98 - 1.87 (m, 1H), 1.69 (d, J = 3.8 Hz, 1H), 1.61 (h, J = 4.1 Hz, 2H), 1.48 - 1.35 (m, 1H), 1.13 (d, J = 39.6 Hz, 5H), 1.03 (s, 3H), 0.74 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 171.15, 164.93, 162.61, 96.68, 93.85, 87.40, 55.79, 48.56, 48.34, 41.18, 29.87, 27.04, 25.93, 20.68, 19.75.

[0055] 11) Synthesis of 2,6-dihydroxy-4-ethoxybenzoic acid fenchyl ester (Target compound 11): Referring to the synthesis method of target compound 10, 2,4,6-trihydroxybenzoic acid fenchyl ester and diethyl sulfate were used as raw materials to synthesize a white solid. 1 HNMR (400 MHz, DMSO-d6) δ 5.90 (s, 2H), 4.50 (d, J = 1.9 Hz, 1H), 4.01 - 3.86 (m, 2H), 1.99 - 1.88 (m, 1H), 1.69 (d, J = 4.1 Hz, 1H), 1.66 - 1.57 (m, 2H), 1.48 - 1.35 (m, 1H), 1.31 - 1.21 (m, 3H), 1.17 (dd, J = 10.1, 1.6 Hz, 1H), 1.08 (s, 3H), 1.07 - 1.04 (m, 1H), 1.03 (s, 3H), 0.74 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 171.24, 164.27, 162.73, 96.34, 94.18, 87.37, 63.83 (d, J = 11.6 Hz), 48.50 (d, J = 7.5 Hz), 48.33, 41.20 (d, J = 10.5 Hz), 29.87 (d, J = 10.1 Hz), 27.00 (d, J = 6.1 Hz), 25.95 (d, J = 5.7 Hz), 20.66, 19.73, 14.96.

[0056] 12) Synthesis of 2,6-dihydroxy-4-isopropoxybenzoic acid fenchyl ester (Target compound 12):

[0057] To 2,4,6-trihydroxybenzoic acid fenchyl ester 1.53 g (5 mmol) was dissolved in 5 ml of acetonitrile, potassium carbonate (6 mmol) and potassium iodide (1 mmol) were added, after stirring for 5 min, 2-chloropropane (7.5 mmol) was added dropwise, and it was reacted at 60°C for 5 h. After the reaction was completed, 50 ml of ethyl acetate was added, washed with saturated brine, and the organic layer was collected and dried, and purified by silica gel chromatography (petroleum ether: ethyl acetate = 100:1) to obtain a white powder. 1 H NMR (400 MHz, DMSO-d6) δ 5.91 (s, 2H), 4.49 (d, J = 1.9 Hz, 1H), 3.65 (d, J = 6.5 Hz, 2H), 1.93 (ddd, J = 12.2, 8.1, 3.6 Hz, 2H), 1.68 (d, J = 3.9 Hz, 1H), 1.59 (d, J = 10.2 Hz, 2H), 1.41 (ddd, J = 12.6, 6.2, 2.3 Hz, 1H), 1.19 (d, J = 5.1 Hz, 2H), 1.08 (s, 3H), 1.02 (s, 3H), 0.90 (d, J = 6.8 Hz, 6H), 0.73 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 171.23, 164.52, 162.73, 96.36, 94.27, 87.39, 74.28 (d, J = 16.1 Hz), 48.47, 48.34, 41.17, 29.84, 28.07, 27.25 - 26.85 (m), 25.93, 20.66, 19.58 (d, J = 31.2 Hz).

[0058] 13) Synthesis of 2,6-dihydroxy-4-isobutoxybenzoic acid fenchyl ester (Target compound 13): It was synthesized by referring to the synthesis method of target compound 12, using 2,4,6-trihydroxybenzoic acid fenchyl ester and 1-chloro-2-methylpropane as raw materials, and a white solid was obtained. 1H NMR (400 MHz, DMSO-d6) δ 5.92 (s, 2H), 4.95 (ddd, J = 9.9, 3.6, 1.8 Hz, 1H), 3.66 (d, J = 6.5 Hz, 2H), 2.32 (ddt, J = 13.7, 9.5, 4.0 Hz, 1H), 2.15 (ddd, J = 12.7, 9.3, 3.3 Hz, 1H), 1.94 (dq, J = 13.2, 6.6 Hz, 1H), 1.72 - 1.63 (m, 2H), 1.29 - 1.17 (m, 2H), 1.08 - 0.99 (m, 1H), 0.91 (s, 3H), 0.90 (s, 3H), 0.87 (s, 3H), 0.84 (s, 3H), 0.82 (s, 3H).

[0059] 14) Synthesis of 2,6-dihydroxy-4-benzyloxybenzoic acid fenchyl ester (Target compound 14): Refer to the synthesis procedure of target compound 12, starting from 2,4,6-trihydroxybenzoic acid fenchyl ester and chlorobenzyl to synthesize, white solid. 1 H NMR (400 MHz, DMSO-d6) δ 7.41 - 7.25 (m, 5H), 6.00 (s, 2H), 5.04 (s, 2H), 4.49 (d, J = 1.7 Hz, 1H), 1.92 (tdd, J = 8.5, 6.6, 6.2, 3.8 Hz, 1H), 1.69 (d, J = 3.8 Hz, 1H), 1.64 - 1.57 (m, 2H), 1.47 - 1.34 (m, 1H), 1.20 - 1.13 (m, 1H), 1.05 (d, J = 22.1 Hz, 7H), 0.74 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 171.10, 163.92, 162.57, 137.06, 129.01, 128.50, 128.19, 96.87, 94.64, 87.40, 48.56, 48.34, 29.86, 27.03, 25.93, 20.69, 19.74.

[0060] 15) Synthesis of 2,6-dihydroxy-3-nitro-4-methoxybenzoic acid fenchyl ester (Target compound 15):

[0061] Synthetic route

[0062]

[0063] Synthetic procedure:

[0064] To 2,6-dihydroxy-4-methoxybenzoic acid fenchyl ester (target compound 10) (1.00 mmol) was dissolved in 2 ml acetic anhydride, a mixture of concentrated nitric acid and acetic anhydride (volume ratio = 1:5) was slowly added dropwise at ice bath condition until the color of the solution was orange yellow, 5 min later, water was added to quench, the water phase was extracted with 50 ml ethyl acetate, and then washed with 100 ml saturated brine for three times, the organic layer was collected and dried, and then purified by silica gel chromatography (petroleum ether: ethyl acetate = 100:1) to obtain a light yellow powder. 1 H NMR (400 MHz, DMSO-d6) δ 6.16 (s, 1H), 4.52 (d, J = 1.8 Hz, 1H), 3.81 (s, 3H), 1.89 (dt, J = 10.6, 6.2 Hz, 1H), 1.70 (d, J = 3.8 Hz, 1H), 1.62 (d, J = 10.4 Hz, 2H), 1.43 (td, J = 12.0, 6.3 Hz, 1H), 1.28 - 1.18 (m, 2H), 1.09 (s, 3H), 1.03 (s, 3H), 0.75 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 170.15, 162.77, 156.13, 155.09, 124.48, 97.28, 92.23, 88.17, 57.14, 48.62, 48.30, 41.16, 29.85, 26.97, 25.91, 20.67, 19.70.

[0065] 16) Synthesis of 2,6-dihydroxy-3-nitro-4-ethoxybenzoic acid fenchyl ester (target compound 16): Referring to the synthesis method of target compound 15, 2,6-dihydroxy-4-ethoxybenzoic acid fenchyl ester (target compound 11) was used as the raw material to synthesize a light yellow powder. 1 H NMR (400 MHz, DMSO-d6) δ 6.16 (s, 1H), 4.52 (d, J = 1.8 Hz, 1H), 3.81 (s, 3H), 1.89 (dt, J = 10.6, 6.2 Hz, 1H), 1.70 (d, J = 3.8 Hz, 1H), 1.62 (d, J = 10.4 Hz, 2H), 1.43 (td, J = 12.0, 6.3 Hz, 1H), 1.28 - 1.18 (m, 2H), 1.09 (s, 3H), 1.03 (s, 3H), 0.75 (s, 3H).

[0066] 17) Synthesis of 2,6-dihydroxy-3-nitro-4-isopropoxybenzoic acid fenchyl ester (Target compound 17): It was synthesized with 2,6-dihydroxy-4-isopropoxybenzoic acid fenchyl ester (Target compound 12) as a raw material according to the synthesis method of Target compound 15, and was a light yellow powder. 1 H NMR (400 MHz, DMSO-d6) δ 6.15 (s, 1H), 4.49 (d, J = 1.9 Hz, 1H), 3.65 (d, J = 6.5 Hz, 2H), 1.92 (ddd, J = 12.2, 8.1, 3.6 Hz, 2H), 1.68 (d, J = 3.9 Hz, 1H), 1.59 (d, J = 10.2 Hz, 2H), 1.42 (ddd, J = 12.6, 6.2, 2.3 Hz, 1H), 1.19 (d, J = 5.1 Hz, 2H), 1.08 (s, 3H), 1.02 (s, 3H), 0.90 (d, J = 6.8 Hz, 6H), 0.73 (s, 3H).

[0067] 18) Synthesis of 2,6-dihydroxy-3-nitro-4-benzyloxybenzoic acid fenchyl ester (Target compound 18): It was synthesized with 2,6-dihydroxy-4-isobutyloxybenzoic acid fenchyl ester (Target compound 14) as a raw material according to the synthesis method of Target compound 15, and was a light yellow powder. 1 H NMR (400 MHz, DMSO-d6) δ 7.42 - 7.25 (m, 5H), 6.17 (s, 1H), 5.04 (s, 2H), 4.49 (d, J = 1.7 Hz, 1H), 1.92 (tdd, J = 8.5, 6.6, 6.2, 3.8 Hz, 1H), 1.69 (d, J = 3.8 Hz, 1H), 1.64 - 1.57 (m, 2H), 1.47 - 1.34 (m, 1H), 1.20 - 1.13 (m, 1H), 1.05 (d, J = 22.1 Hz, 7H), 0.74 (s, 3H).

[0068] Inhibitory effect of the compound of Example 2 on IL-1β, TNF-α increase in a lipopolysaccharide-induced RAW264.7 macrophage inflammation model

[0069] The experiment was set up blank group, model group, drug group (youkang alcohol, fenchol, control compound 1, target compound 1-target compound 18). The logarithmic growth period of RAW264.7 mouse peritoneal macrophage was diluted according to the ratio of 1:9 and inoculated in 6-hole plate, each group was added with 1 mL of culture solution containing cells, and adherent growth was carried out for 2 h. After the cells adhered, the culture medium of the drug group was sucked out (blank group and model group were not treated), and the prepared drug solution (4 μg / mL) 1 mL was added, and the culture was continued for 30 min. After 30 min, the blank group was added with 1 mL of complete culture medium; the model group and the drug group were added with 2.4 μg / mL lipopolysaccharide (LPS) solution 1 mL, so that the final concentration of LPS was 1.2 μg / mL. After 24 h of cell treatment (2 μg / mL), the 6-hole plate was taken out from the incubator, the cell culture supernatant was collected, centrifuged at 3000 rpm at 4°C for 10 min, and the content of IL-1β and TNF-α was determined according to the kit instructions.

[0070] The results show that: at the concentration of 2 μg / mL, youkang alcohol, fenchol, control compound 1 have no significant inhibitory effect on the increase of IL-1β and TNF-α of RAW264.7 macrophage inflammation model induced by lipopolysaccharide. The inhibitory effect of youkang alcohol series target compounds on the increase of IL-1β and TNF-α of RAW264.7 macrophage inflammation model induced by lipopolysaccharide is significantly stronger than that of control compound 1 or youkang alcohol. The inhibitory effect of fenchol series target compounds on the increase of IL-1β and TNF-α of RAW264.7 macrophage inflammation model induced by lipopolysaccharide is significantly stronger than that of fenchol Figure 1 、 Figure 2 ).

[0071] Example 3

[0072] SD rats were randomly divided into groups, 6 rats in each group. The sham operation group and the model group were injected with the same amount of solvent intravenously, and each drug group (control compound 1, target compound 1, target compound 6, target compound 15) was injected with each compound 20 μmol / kg intravenously. 0.5 h after administration, the sham operation group was subcutaneously injected with normal saline 100 μL at the toes of the hind limbs, and the other groups were injected with 1% carrageenan 100 μL. The toe volume of the rats was measured at 0 h, 0.5 h, 1 h, 2 h, and 4 h after the injection of carrageenan, and the rat foot swelling rate was calculated.

[0073] The results show that: control compound 1, target compound 1, 6, and 15 have a significant inhibitory effect on rat foot swelling caused by carrageenan. The inhibitory rate of target compound 6 on rat foot swelling caused by carrageenan is significantly stronger than that of control compound 1 Figure 3 ).

[0074] Example 4 Protective effect of target compound on rat focal cerebral ischemia-reperfusion model

[0075] The middle cerebral artery occlusion (MCAO) cerebral ischemia-reperfusion model was prepared by middle cerebral artery thread embolization method. The animals were anesthetized with gas (isoflurane), and first placed in the induction box of MSS-3 small animal anesthetizing machine for anesthesia, then fixed on the mouse plate connected with the breathing mask in supine position, the skin was disinfected, the right common carotid artery, external carotid artery, internal carotid artery were separated, the vagus nerve was gently stripped, the external carotid artery was ligated and cut. The proximal end of the common carotid artery was clamped, a 2438-A5 thread embolus (the top end is hemispherical, the front end 5-6 mm is coated with silicone) was inserted from the distal end of the ligation line of the external carotid artery, passed through the common carotid artery bifurcation into the internal carotid artery, and then slowly inserted to about 20 mm from the bifurcation, blocking the blood supply of the middle cerebral artery, the skin of the neck was sutured, disinfected, and put back into the cage. After 90 min of ischemia, the rats were induced and fixed on the mouse plate again, the skin of the neck was cut, the thread embolus was gently pulled out to restore blood supply for reperfusion, the skin of the neck was sutured, disinfected, and put back into the cage for feeding.

[0076] 4.1 Preparation of cerebral ischemia model

[0077] The middle cerebral artery occlusion (MCAO) cerebral ischemia-reperfusion model was prepared by middle cerebral artery thread embolization method. The animals were anesthetized with gas (isoflurane), and first placed in the induction box of MSS-3 small animal anesthetizing machine for anesthesia, then fixed on the mouse plate connected with the breathing mask in supine position, the skin was disinfected, the neck was incised in the middle, the right common carotid artery, external carotid artery, internal carotid artery were separated, the vagus nerve was gently stripped, the external carotid artery was ligated and cut. The proximal end of the common carotid artery was clamped, a 2438-A5 thread embolus (the top end is hemispherical, the front end 5-6 mm is coated with silicone) was inserted from the distal end of the ligation line of the external carotid artery, passed through the common carotid artery bifurcation into the internal carotid artery, and then slowly inserted to about 20 mm from the bifurcation, blocking the blood supply of the middle cerebral artery, the skin of the neck was sutured, disinfected, and put back into the cage. After 90 min of ischemia, the rats were induced and fixed on the mouse plate again, the skin of the neck was cut, the thread embolus was gently pulled out to restore blood supply for reperfusion, the skin of the neck was sutured, disinfected, and put back into the cage for feeding.

[0078] 4.2 Evaluation of neurological deficit symptoms

[0079] The neurological deficit symptoms were evaluated by the modified Bederson 5-point method.

[0080] 0: When the tail was lifted, both forelimbs of the animal were stretched towards the floor, and there was no other behavioral defect

[0081] 1: When the tail was lifted, the surgical (left) side of the animal showed wrist and elbow flexion, shoulder internal rotation, elbow abduction, and close to the chest wall

[0082] 2: When the animal was placed on a smooth flat plate, the resistance decreased when the surgical side shoulder was pushed to the opposite side

[0083] 3: When the animal walked freely, it circled or turned to the opposite side of the surgery

[0084] 4: Limb flaccid paralysis, no spontaneous activity of the limbs

[0085] 4.3 Measurement of cerebral infarction area

[0086] The method reported in the literature was used. The animals were anesthetized with 10% chloral hydrate, decapitated, and the brain was removed, with the olfactory bulb, cerebellum, and lower brain stem removed. The surface of the brain was washed with normal saline, and the residual water was absorbed. The brain was placed at -80°C for 7 min, and then immediately cut into coronal sections at 2 mm intervals. The brain slices were incubated in TTC (20 g / L) prepared with normal saline at 37°C for 90 min. The normal brain tissue was dyed dark red, and the ischemic brain tissue was pale. After washing with normal saline, the brain slices were arranged in order from front to back, and the residual water was absorbed. The photographs were taken. The right ischemic area (white area) and the right area were circled, and the percentage of cerebral infarction area was calculated using the following formula.

[0087]

[0088] 4.4 Statistical analysis

[0089] The quantitative data were expressed as mean ± standard error. The cerebral infarction area and neurological deficit score were analyzed by one-way ANOVA, and the significance of the difference between the two groups was determined by Scheffe's test. The mortality rate and body weight were analyzed by ANOVA. The Stata statistical software was used for analysis, and the difference P < 0.05 was defined as significant.

[0090] 4.5 Effect of the test substance on neurological deficit symptoms

[0091] The effect of the test substance on neurological deficit symptoms is shown in Table 4.5. Figure 4 Each test substance group improved the neurological deficit symptoms. The target compound 1 group and the target compound 6 group were significantly better than the control compound group.

[0092] 4.6 Effect of the test substance on the cerebral infarction area (%)

[0093] The effect of the test substance on the cerebral infarction area (%) is shown in Table 4.6. Figure 5 Each test substance group improved the cerebral infarction area. The target compound 1 group and the target compound 6 group were significantly better than the control compound group.

[0094] Example 5 Anti-cerebral edema effect of the target compound in a rat autologous arterial blood infusion method cerebral hemorrhage model

[0095] 10% chloral hydrate was used to anesthetize the rats intraperitoneally (dose of 4 μl / g), the rats were in prone position, the skull was cut in the middle of the head top, a 1.5 mm diameter hole was drilled at 0.2 mm in front of the anterior fontanel, 3 mm right of the midline, and the depth reached the surface of the dura mater. The microsyringe was vertically inserted into the hole for about 6 mm until the target point. The operation group slowly injected 50 μL of fresh non-anticoagulant arterial blood into the brain (equivalent to the position of the caudate nucleus) within 20 min, the sham operation group slowly injected 50 μL of normal saline into the brain within 20 min, and the needle was slowly pulled out after 10 min. A total of 5 groups were set, namely the sham operation group, the model group, the control compound 1 group (5 mg / kg), the target compound 1 group (5 mg / kg), and the target compound 6 group (5 mg / kg). The animals in each group were injected with drugs once at 2 h, 1 d, and 2 d after the operation. The animals were sacrificed by decapitation at 3 d after the operation, the right cerebral hemisphere of each rat was weighed (wet weight), and was placed in a 80°C constant temperature oven, and was taken out after 24 h and weighed again (dry weight). The water content of the brain tissue = {(wet weight-dry weight) / wet weight}x100%. The effects of the tested substances on the water content (%) of the brain tissue of the rats with cerebral infarction are shown in Table 1. Figure 6 The target compound 1 group and the target compound 6 group were significantly better than the control compound 1 group.

[0096] The above only describes the preferred embodiments of the present application, and it should be noted that for ordinary skilled persons in the art, several improvements and refinements can be made without departing from the principles of the present application, and these improvements and refinements should also be considered as the protection scope of the present application.

Claims

1. Ester compounds of 4-alkyloxy-substituted 2,6-dihydroxybenzoic acid with d-menthol or fenchol, characterized in that The ester compound structure conforms to general formula (I) wherein: R 1 is R 2 R is -H, -NO2; R 3 alkyl of 1 to 4 carbon atoms or 2. The ester compound of 4-alkoxy-substituted 2, 6-dihydroxybenzoic acid with d-menthol or fenchol according to claim 1, characterized in that, R 3 alkyl is an alkyl group of 1 carbon atom.

3. The ester compound of 4-alkoxy-substituted 2,6-dihydroxybenzoic acid with d-menthol or fenchol according to claim 1, characterized in that, The structure is any of the following:

4. A pharmaceutically acceptable salt of the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in claims 1-3.

5. A pharmaceutical composition, characterized by, The pharmaceutical composition comprises the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in any one of claims 1-3 or the pharmaceutically acceptable salt of the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in claim 4.

6. Use of the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in claims 1-3 or the pharmaceutically acceptable salt of the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in claim 4 or the pharmaceutical composition as claimed in claim 5 in the preparation of a medicament for treating inflammation-related diseases and / or brain stroke injury, brain edema; the inflammation-related diseases are lipopolysaccharide-induced macrophage inflammation and / or carrageenan-induced footpaw swelling.

7. A medicament for treating an inflammation-related disease and / or brain stroke injury, brain edema, characterized by, The active ingredient comprises the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in claims 1-3 or the pharmaceutically acceptable salt of the ester compound of 4-alkoxyl-substituted 2,6-dihydroxybenzoic acid with d-camphor or fenchol as claimed in claim 4; The inflammation-related diseases are lipopolysaccharide-induced macrophage inflammation and / or carrageenan-induced footpaw swelling.

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