Succinimide derivatives, their preparation methods and applications

Abstract

This application discloses succinimide derivatives in the field of organic chemistry, with the general structural formula shown in Formula V. During preparation, compound I reacts with compound II to yield compound III; compound III reacts with compound IV in the presence of a condensing agent to yield succinimide derivative V. The succinimide derivatives of this application can be used in the preparation of sedative, tranquilizing, nootropic, anticonvulsant, antiepileptic, antineuralgia, anti-anxiety, antidepressant, antidementia, and / or brain injury protective drugs.

Description

[0001] This application claims priority to Chinese Patent Application No. 2024104773680, filed on April 19, 2024, entitled "Succinimidide Derivatives and Preparation Methods Thereof and Applications Thereof", the entire contents of which are incorporated herein by reference. Technical Field

[0002] This invention belongs to the field of organic chemistry, and specifically relates to succinimide derivatives, their preparation methods, and applications. Background Technology

[0003] Amber, the resin of ancient pines (Pinus spp.) in the Pinaceae family, is a representative traditional Chinese medicine for its calming and sedative effects. The *Chinese Materia Medica* summarizes amber's effects as "calming the nerves and relieving convulsions"; "dispersing blood stasis and stopping bleeding"; "promoting urination and relieving strangury"; and "removing corneal opacity and improving eyesight." The formula "Xiao'er Hupo Zhenjing Wan" (Children's Amber Convulsion Pill) contains amber, cinnabar, musk, gastrodia elata, borneol, and arisaema cum bile as its main ingredients. This formula is used to treat acute and chronic infantile convulsions, feverish convulsions, and coma. Succinic acid is a compound with a succinic acid structure, widely found in natural products. Modern pharmacological studies have shown that succinic acid derivatives possess a wide range of pharmacological activities, including anti-epileptic, anti-inflammatory, anti-tumor, and antibacterial effects.

[0004] Gastrodin (GAS) is a small-molecule active compound extracted from the rhizome of Gastrodia elata Blume. Its chemical name is 4-hydroxybenzyl alcohol-4-O-β-D-glucopyranoside, and it is the main pharmacologically active component of the traditional Chinese medicine Gastrodia elata. Studies have shown that gastrodin possesses pharmacological effects such as neuroprotection, anti-inflammation, analgesia, sedation and hypnosis, anticonvulsant effects, and improvement of behavioral disorders.

[0005] Borneol (Borneolum Syntheticum) has a long history of use in my country. The "Newly Revised Materia Medica" records its effects as "treating evil qi in the heart and abdomen, wind-dampness accumulation, deafness, improving eyesight, and removing redness and pterygium in the eyes." Modern pharmacology shows that borneol has the effects of opening the orifices and refreshing the mind, clearing heat and detoxifying, and relieving pain. Tambe et al. found that borneol has the effects of protecting the heart and brain and anticonvulsant, and can significantly inhibit the occurrence of epilepsy in pentylenetetrazol-ignited mice and inhibit the expression of neurochemical stress and neuroinflammatory markers in the brain [Tambe R, Naunyn Schmiedebergs Arch Pharmaco, 2016, 389(5):467].

[0006] Based on the excellent pharmacological activities of active ingredients such as succinic acid, gastrodin, and borneol in sedation, tranquilization, anticonvulsant, and neuroprotection, the synthesis of a series of succinimide derivatives, including succinimide-amino acid-gastrodin ester, succinimide-amino acid-vanillyl ester, and succinimide-amino acid-borneol ester, is of positive significance for screening new chemical drugs with therapeutic effects in anti-epileptic, anticonvulsant, memory improvement, anti-anxiety, anti-depression, anti-dementia, and brain injury improvement. Summary of the Invention

[0007] The present invention aims to provide a succinimide derivative with anti-epileptic, anti-convulsant, anti-neuralgia, memory-improving, anti-anxiety, anti-depressant, anti-dementia and / or brain injury-improving effects.

[0008] The succinimide derivatives in this scheme have the general structural formula shown in Formula V:

[0009]

[0010] Wherein: R1 is H, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, branched, straight-chain C1-8 alkyl, cycloalkyl C m H 2m-1 (m=3-6), methylcycloalkyl C m+1 H 2m+1 (m=3-6), substituted allyl, substituted phenyl, substituted benzyl, heterocyclic, methoxymethyl, hydroxymethyl, 2-hydroxyethyl, methylthioethyl, heterocyclic; R2 is methyl, ethyl, isopropyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, branched, straight-chain C1-8 alkyl, cycloalkyl C m H 2m-1 (m=3-6), methylcycloalkyl C m+1 H 2m+1 CH2 (m=3-6), substituted allyl, substituted cinnamyl, rac-2-camphenyl, (-)-2-camphenyl, (+)-2-camphenyl, rac-menthyl, (-)-menthyl, (+)-menthyl, substituted phenyl, substituted benzyl, heterocyclic, 2-methylpiperidine, dimethylamino, diethylamino, diisopropylamino, 2,2-dimethyl-3-(propylamino)propyl-1-yl, N,2-dimethyl-1-phenylpropyl-2-amino; R3 is H, methyl, ethyl, phenyl; R4 is H, methyl, ethyl, phenyl, spirocyclopentyl; or R3R4 constitute cyclohexyl, phenyl, (1R,4S)-bicyclo[2,2,1]hexane-2-enyl; X is O, NH or piperazine; n is 0~3.

[0011] Further, R1 is hydrogen, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, 2-methyl-n-butyl, neopentyl, n-hexyl, 2-methyl-n-pentyl, 3-methylpentyl, 4-methylpentyl, 2-hexyl, 4-methylpentan-2-yl, 3-methylpentan-2-yl, 3-pentyl, n-heptyl, n-octyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopropanemethyl, cyclobutanemethyl, cyclopentanemethyl, cyclohexanemethyl, allyl, 3-methyl-2-buten-1-yl, substituted phenyl, substituted benzyl, methoxymethyl, hydroxymethyl, 2-hydroxyethyl, methylthioethyl, or aromatic heterocyclic rings.

[0012] Furthermore, the structural formula of R1 is as follows:

[0013]

[0014] Furthermore, the structural formula of R2 is as follows:

[0015]

[0016]

[0017] Furthermore, the R3 or R4 structure is shown below;

[0018]

[0019] Alternatively, R3 and R4 together form cyclohexyl, phenyl, or (1R,4S)-bicyclo[2,2,1]hexane-2-enyl, with the following structural formula:

[0020]

[0021] Regarding the above-mentioned succinimide derivatives, this application also provides a method for preparing succinimide derivatives, comprising the following steps:

[0022] The reaction of compound I with compound II yields compound III; the reaction of compound III with compound IV in the presence of a condensing agent yields succinimide derivative V.

[0023]

[0024] Furthermore, the specific preparation process of the general formula III compound is as follows: the general formula I compound and the general formula II compound are added sequentially to the reaction vessel at a molar ratio of 1.0 to 2.0:1.0, and reacted at 70°C to 100°C until the general formula I compound and / or the general formula II compound disappear; the solvent is removed by vacuum concentration, water is added to the residue to precipitate it, the residue is filtered, and dried at 30-60°C to obtain the general formula III compound.

[0025] Further, in the preparation of succinimide derivative V: Compound of general formula III and compound of general formula IV, condensing agent and catalyst are added sequentially to the reaction vessel. The molar ratio of compound of general formula III to compound of general formula IV is 1.0:1.0 to 1.5. The reaction is carried out at 5℃-40℃ until compound of general formula III and / or compound of general formula IV is consumed. After washing with pure water, the mixture is extracted with ethyl acetate, concentrated under vacuum to remove ethyl acetate, and the organic phase is dried, concentrated under vacuum, and purified by column chromatography.

[0026] Furthermore, the condensing agent is one of DCC, DIC, EDC, BOP, PyBOP, HATU, HBTU, TBTU, ethyl chloroformate, phenyl chloroformate, isopropyl chloroformate, 1-n-propylphosphonic anhydride, etc., preferably EDC.

[0027] Furthermore, the catalyst is one of Et3N, DIEA, DMAP, DPPY, 2,6-dimethylpyridine, DABCO, DBU, etc., with DMAP being preferred.

[0028] Furthermore, the stereoisomers of the succinimide derivatives or mixtures of different stereoisomers thereof.

[0029] Furthermore, the succinimide derivatives are used in the preparation of sedative, tranquilizing, nootropic, anticonvulsant, antiepileptic, antineuralgia, anti-anxiety, antidepressant, antidementia, and / or brain injury protective drugs.

[0030] Furthermore, the use of stereoisomers of the succinimide derivatives or mixtures of different stereoisomers in the preparation of sedative, tranquilizing, nootropic, anticonvulsant, antiepileptic, antineuralgia, anti-anxiety, antidepressant, antidementia, and / or brain injury protective drugs. Attached Figure Description

[0031] Figure 1 A diagram showing the effects of the compound on normal zebrafish.

[0032] Figure 2 The effect of the compound on PTZ-induced seizures in zebrafish is shown in the figure.

[0033] Figure 3 Comparative images of neuronal apoptosis in the CA1 region of mouse hippocampus. Detailed Implementation

[0034] The following detailed description illustrates the specific implementation method:

[0035] The preparation method of succinimide derivative V, and the reaction formula are shown below:

[0036]

[0037] Specific method: Compound of general formula I and compound of general formula II are added sequentially to a reaction vessel at a molar ratio of 1.0-2.0:1.0. The reaction is carried out at 70-100℃ for 3-10 hours, and the reaction is monitored by TLC until the starting material disappears. The solvent is removed by vacuum concentration, and water is added to the residue to precipitate it. The residue is filtered and dried at 40-60℃ to obtain compound of general formula III.

[0038] Compound of general formula III and compound of general formula IV, condensing agent, and catalyst were added sequentially to a reaction vessel in a molar ratio of 1.0:(1.0–1.5):(1.0–1.5):(0.1–0.5). The reaction was carried out at 5–40 °C for 12–24 hours until the starting material disappeared. The mixture was then washed with pure water, extracted with ethyl acetate, concentrated under vacuum to remove the solvent, dried with a drying agent, concentrated, and purified by column chromatography to obtain compound of general formula V. The catalyst used was DMAP, and the condensing agent used was EDC.

[0039] The invention will be better understood through the following description, which is merely illustrative and advantageous embodiments of the invention are not limited thereto.

[0040] Example 1

[0041] Succinimide-O-methylserine Borneol Ester (RJ-1)

[0042]

[0043] Preparation of succinimide-O-methylserine: Potassium succinimide (5.0 g, 50.5 mmol), O-methylserine (6.6 g, 55.5 mmol), and 60 mL of dioxane were added to a 100 mL reaction flask. The mixture was heated to 100 °C and reacted for 6 hours. All solvents were then evaporated under reduced pressure. 20 mL of water was added to the residue, and the mixture was stirred for 20 min. The mixture was then filtered, and the off-white solid collected was succinimide-O-methylserine.

[0044] Preparation of succinimide-O-methylserine borneol ester: Succinimide-O-methylserine (2.0 g, 9.95 mmol) was added to 80 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. Borneol (1.53 g, 9.95 mmol), EDC (2.85 g, 14.92 mmol), and DMAP (243 mg, 1.99 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (50 mL * 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The dichloromethane phase was dried over anhydrous sodium sulfate and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent. The product was 2.42 g of colorless oil; 72% yield.

[0045] Example 2

[0046] Succinimide-glycine 4-hydroxybenzyl ester (RJ-2)

[0047]

[0048] The preparation method of succinimide-glycine is the same as in Example 1, except that the raw material is changed to glycine, and the other operation steps are the same.

[0049] Succinimide-glycine (1.56 g, 9.95 mmol) was added to 80 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. 4-hydroxybenzyl alcohol (1.23 g, 9.95 mmol), EDC (2.85 g, 14.92 mmol), and DMAP (243 mg, 1.99 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (50 mL x 2). The organic phase was collected and washed once each with 20 mL of aqueous phase and brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% ethyl acetate / petroleum ether-15% methanol / dichloromethane as the eluent.

[0050] The product was 0.91 g of an off-white solid; yield 42%. 1 H NMR (400MHz, CDCl3) δ7.28 (d, J = 7.2 Hz, 2H), 6.84 (d, J = 7.2 Hz, 2H), 5.11 (s, 2H), 4.28 (s, 2H), 2.79 (s, 4H); 13C NMR (101MHz, CDCl3) δ176.3,166.6,158.7,129.4,129.1,115.4,115.0,67.5,39.6,28.2; HRMS (ESI): Exactmass calcd for C 13 H 13 NO5, [M+H] + ,264.0866.Found 264.0860.

[0051] Example 3

[0052] Succinimide-Isoleucine Borneol Ester (RJ-3)

[0053]

[0054] The preparation method of succinimide-isoleucine is the same as in Example 1, except that the raw material is replaced with isoleucine, and the other operation steps are the same.

[0055] In a 100 mL reaction flask, succinimide-isoleucine (1.24 g, 4.98 mmol) and 50 mL of dichloromethane were added and dissolved with stirring. Borneol (0.768 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 1.9 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent.

[0056] The product was 1.18 g of a colorless oil (68% yield). Major isomer: minor isomer = 2.0:1.0.

[0057] 1 H NMR (400MHz, CDCl3) δ4.90-4.83(m,1H),4.53(d,J=8.0Hz,1H),2.76(s,4H),2.47-2.32(m,2H),1.73-1.68(m,4H),1.28- 1.17(m,3H),1.11(d,J=6.4Hz,1H),0.97(d,J=7.2Hz,3H),0.88(s,3H),0.86(s,3H),0.82(d,J=6.4Hz,3H),0.76(s,3H); 13C NMR (101MHz, CDCl3) δ176.5,168.7,81.5,56.8,48.7,47.7,44.7,36.4,34.0,27.9,27.5,27.2,19.6,18.8,15.6,13.4,11.3; HRMS (ESI): Exact mass calcd for C 20 H 31 NO4, [M+Na] + ,372.2145.Found 372.2153.

[0058] Example 4

[0059] Succinimide-valine borneol ester (RJ-4)

[0060]

[0061] The preparation method of succinimide-valine is the same as in Example 1, except that the raw material is changed to valine, and the other operation steps are the same.

[0062] Succinimide-valine (991 mg, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. Borneol (0.768 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent.

[0063] The product was 1.24 g of a colorless oil; yield 74.2%. Major isomer:minor isomer = 3.0:1.0.

[0064] 1 H NMR (400MHz, CDCl3) δ4.91-4.88(m,1H),4.41(d,J=8.8Hz,1H),2.76(s,4H),2.72-2.64(m,1H),2.38-2.31(m,1H),1.77-1.66(m ,4H),1.28-1.19(m,3H),1.15(d,J=6.8Hz,3H),1.05-0.98(m,1H),0.89(s,3H),0.86(s,3H),0.84(d,J=7.2Hz,3H),0.79(s,3H);13 C NMR (101MHz, CDCl3) δ176.4,168.5,81.4,58.4,48.7,47.7,44.7,36.5,28.0,27.9,27.6,27.2,21.2,19.6,19.4,18.8; HRMS (ESI): Exact mass calcd for C 19 H 29 NO4, [M+Na] + ,358.1989.Found358.1998.

[0065] Example 5

[0066] Succinimide-alanine borneol ester (RJ-5)

[0067]

[0068] The preparation method of succinimide-alanine is the same as in Example 1, except that the raw material is changed to alanine, and the other operation steps are the same.

[0069] Succinimide-alanine (851 mg, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. Borneol (0.768 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent.

[0070] The product was 1.18 g of a colorless oil; yield 77.3%. Major isomer:minor isomer = 2.0:1.0.

[0071] 1 H NMR (400MHz, CDCl3) δ4.99-4.82(m,2H),2.75(s,4H),2.40-2.32(m,1H),1.76-1.68( m,4),1.31-1.17(m,2H),1.03-1.00(m,2H),0.89(s,3H),0.86(s,3H),0.8s(s,1.5H); 13C NMR (101MHz, CDCl3) δ176.1,169.2,81.6,48.8,48.3,47.8,44.8,36.5,28.1,27.9,27.1,19.6,18.8,14.2,13.3; HRMS (ESI): Exact mass calcd for C 17 H 25 NO4, [M+Na] + ,330.1676.Found 330.1685.

[0072] Example 6

[0073] Succinimide-valine 4-hydroxybenzyl ester (RJ-6)

[0074]

[0075] The preparation method of succinimide-valine is the same as in Example 1, except that the raw material is changed to valine, and the other operation steps are the same.

[0076] Succinimide-valine (991 mg, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. 4-hydroxybenzyl alcohol (0.617 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once each with 20 mL of aqueous phase and brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 50% ethyl acetate / petroleum ether as the eluent.

[0077] The product was 0.53 g of a colorless oil; yield 34.6%. 1 H NMR (400MHz, CDCl3) δ7.17 (d, J = 8.4Hz, 2H), 6.78 (d, J = 8.4Hz, 2H), 5.07 (dd, J1 = 12.0Hz, J2 = 20.0Hz, 2H), 4. 72(dd,J1=5.6Hz,J2=10.4Hz,1H),2.72(s,4H),2.15-2.05(m,2H),1.28-1.21(m,2H),0.90(t,J=7.2Hz,3H); 13CNMR(101MHz, CDCl3)δ176.8,169.0,156.1,130.3,127.1,115.4,67.5,52.7,29.7,28.0,19.5,13.3; HRMS(ESI): Exact mass calcd for C 16 H 19 NO5, [M+H] + ,306.1336.Found306.1335.

[0078] Example 7

[0079] Succinimide-Isoleucyl-4-fluorobenzylamine (RJ-7)

[0080]

[0081] The preparation method of succinimide-isoleucine is the same as in Example 1, except that the raw material is replaced with isoleucine, and the other operation steps are the same.

[0082] Succinimide-isoleucine (1.06 g, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. EDC (1.43 g, 7.46 mmol), HOBt (673 mg, 4.98 mmol), and DMAP (122 mg, 1.0 mmol) were added. After 10 min, 4-fluorobenzylamine (0.622 g, 4.98 mmol) was added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL * 2). The organic phase was collected and washed successively with saturated sodium bicarbonate solution, aqueous phase, and saturated brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 1.8 g of crude product. The crude product was subjected to silica gel column chromatography, yielding 1.15 g of a colorless oily substance; 72.5% yield.

[0083] 1 H NMR (400MHz, CDCl3) δ7.15(dd,J1=5.2Hz,J2=8.8Hz,2H),6.93(t,J=8.8Hz,2H),4.38(dd,J1=6.0Hz,J2=14.5Hz,2H),4.32((t,J=11.6Hz, 1H),4.21(dd,J1=5.6Hz,J2=14.4Hz,1H),2.67(s,4H),2.53-2.50(m,1H),1.20-1.16(m,1H),0.90(d,J=6.8Hz,3H),0.76(d,J=7.6Hz,3H); 13C NMR (101MHz, CDCl3) δ177.5,168.4,163.3,1609,133.9,133.8,129.4,129.3,115.6,115.4,62.5,42.8,32.0,25.3,15.6,10.0; 19 F NMR (376MHz, CDCl3) δ-115.0; HRMS (ESI): Exact mass calcd for C 17 H 21 FN₂O₃,[M+H] + ,321.1609.Found321.1601.

[0084] Example 8

[0085] Succinimide-Valylbenzylamine (RJ-8)

[0086]

[0087] The preparation method of succinimide-valine is the same as in Example 1, except that the raw material is changed to valine, and the other operation steps are the same.

[0088] Succinimide-valine (0.991 g, 4.98 mmol) was added to 50 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. EDC (1.43 g, 7.46 mmol), HOBt (673 mg, 4.98 mmol), and DMAP (122 mg, 1.0 mmol) were added. After 10 min, benzylamine (0.622 g, 4.98 mmol) was added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL * 2). The organic phase was collected and washed successively with saturated sodium bicarbonate solution, aqueous phase, and saturated brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 1.5 g of crude product. The crude product was subjected to silica gel column chromatography with 50% ethyl acetate / petroleum ether as the eluent.

[0089] The product was 1.05 g of colorless oil; yield 73.4%. 1 H NMR (400MHz, CDCl3) δ7.55-7.26 (m, 5H), 4.52 (dd, J1=6.4Hz, J2=14.8Hz, 1H), 4.39-43 3(m,2H),2.83-2.74(m,1H),2.77(s,4H),1.07(d,J=6.4Hz,3H),0.82(d,J=6.4Hz,3H); 13C NMR (101MHz, CDCl3) δ177.4,168.2,137.9,128.7,128.0,127.5,63.9,43.6,27.9,26.6,19.7,19.3; HRMS (ESI): Exact masscalcd for C 16 H 20 FN₂O₃,[M+H] + ,289.1547.Found 289.1541.

[0090] Example 9

[0091] Succinimide-valine borneol ester (RJ-9)

[0092]

[0093] The preparation method of succinimide-valine is the same as in Example 1, except that the raw material is changed to valine, and the other operation steps are the same.

[0094] Succinimide-valine (991 mg, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. Borneol (0.768 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added. The reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent. The product was 1.20 g of colorless oil; yield 72.6%.

[0095] 1 H NMR(400MHz, CDCl3)δ4.90-4.86(m,1H),4.72(dd,J1=6.0Hz,J2=9.6Hz,1H),2.76(s,4H),2.37-2.29(m,1H),2.14- 2.09(m,2H),1.74-1.65(m,3H),1.30-1.25(m,4H),0.94(t,J1=7.2Hz,3H),0.88(s,3H),0.86(s,3H),0.82(s,3H); 13C NMR (101MHz, CDCl3) δ176.4,169.0,81.6,52.7,48.7,47.7,44.8,36.2,29.7,28.0,27.8,27.2,19.6,18.8,13.4; HRMS (ESI): Exact mass calcd for C 19 H 29 NO4, [M+Na] + ,358.1989.Found 358.1998.

[0096] Example 10

[0097] Succinimide-phenylalanine borneol ester (RJ-10)

[0098]

[0099] The preparation method of succinimide-phenylalanine is the same as in Example 1, except that the raw material is changed to phenylalanine, and the other operation steps are the same.

[0100] Succinimide-phenylalanine (1.23 g, 4.98 mmol) was added to 50 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. Borneol (0.768 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent. The product was 1.49 g of colorless oil; yield 78.1%.

[0101] 1 H NMR (400MHz, CDCl3) δ7.27-7.13(m,5H),5.03(dd,J1=5.6Hz,J2=11.2Hz,1H),4.97-4.94(m,1H),3.50-3.40(m,2H),2.61-2.49(m,4H) ,2.41-2.33(m,1H),1.74-1.68(m,3H),1.29-1.21(m,2H),1.04(dd,J1=3.2Hz,J2=13.6Hz,1H),0.90(s,3H),0.87(s,3H),0.82(s,3H); 13C NMR (101MHz, CDCl3) δ176.2,168.4,136.6,128.8,128.5,126.9,81.9,53.7,48.9,47.8,44.8,27.1,19.6,18.8,13.4; HRMS (ESI): Exact mass calcd forC 23 H 29 NO4, [M+Na] + ,406.1989.Found 406.1999.

[0102] Example 11

[0103] Succinimide-Leucine Borneol Ester (RJ-11)

[0104]

[0105] The preparation method of succinimide-leucine is the same as in Example 1, except that the raw material is changed to leucine, and the other operation steps are the same.

[0106] Succinimide-leucine (1.06 g, 4.98 mmol) was added to 50 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. Borneol (0.768 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent. The product was 1.33 g of colorless oil; 76.5% yield.

[0107] 1 H NMR (400MHz, CDCl3) δ4.90-4.85(m,1H),4.82-4.77(m,1H),2.74(s,4H),2.37-2.30(m,1H),2.18-2.10(m,1H),1.95-1.88(m,1H),1.76-1.63(m, 3H),1.46-1.39(m,1H),1.32-1.15(m,2H),1.00(dd,J1=3.2Hz,J2=13.6H z,1H),0.93(s,3H),0.92(s,3H),0.88(s,3H),0.85(s,3H),0.79(s,2H); 13C NMR (101MHz, CDCl3) δ176.5,169.3,81.6,51.4,48.8,47.8,44.7,36.6,36.4,28.1,27.9,27.1,25.1,23.0,21.2,19.6,18.8,13.3; HRMS (ESI): Exact mass calcd for C 20 H 31 NO4, [M+Na] + ,372.2145.Found 372.2155.

[0108] Example 12

[0109] Succinimide-leucyl-4-fluorobenzylamine (RJ-12)

[0110]

[0111] The preparation method of succinimide-leucine is the same as in Example 1, except that the raw material is changed to leucine, and the other operation steps are the same.

[0112] Succinimide-leucine (1.06 g, 4.98 mmol) was added to 50 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. EDC (1.43 g, 7.46 mmol), HOBt (673 mg, 4.98 mmol), and DMAP (122 mg, 1.0 mmol) were added. After 10 min, 4-fluorobenzylamine (0.622 g, 4.98 mmol) was added, and the reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL * 2). The organic phase was collected and washed successively with saturated sodium bicarbonate solution, aqueous phase, and saturated brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 1.8 g of crude product. The crude product was subjected to silica gel column chromatography with 50% ethyl acetate / petroleum ether as the eluent.

[0113] The product was 1.25 g of white solid; yield 83.1%; 1 H NMR (400MHz, CDCl3) δ7.24-7.20(m,2H),7.01(t,J=8.4Hz,1H),6.48(brs,1H),4.80(dd,J1=5.2Hz,J2=10.8Hz,1H),4.44(m,2H),2. 73(s,4H),2.34-2.26(m,2H),1.88-1.82(m,1H),1.74-1.67(m,2H),1.42-1.35(m,1H),0.92(d,J=8.0Hz,3H),0.88(d,J=8.0Hz,3H);13 C NMR (101MHz, CDCl3) δ177.1,168.6,163.4,160.9,133.6,133.6,129.4,129.3,115.6,115.4,53.7,43.0,41.9,36.7,28.0,27.0,25.3,23.0,21.3; 19 FNMR (376MHz, CDCl3) δ-114.9; HRMS (ESI): Exact mass calcd forC 17 H 21 FN₂O₃,[M+H] + ,321.1609.Found 321.1604.

[0114] Example 13

[0115] Succinimide-valine 4-hydroxybenzyl ester (RJ-13)

[0116]

[0117] The preparation method of succinimide-valine is the same as in Example 1, except that the raw material is changed to valine, and the other operation steps are the same.

[0118] Succinimide-valine (991 mg, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. 4-hydroxybenzyl alcohol (0.617 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4–6 h until the starting material disappeared. The pH was adjusted to 4–5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL * 2). The organic phase was collected and washed once each with 20 mL of aqueous phase and brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 2.0 g of crude product. The crude product was subjected to silica gel column chromatography with 50% ethyl acetate / petroleum ether as the eluent.

[0119] The product was 0.45 g of a colorless oil; yield 30.3%. 1 H NMR (400MHz, CDCl3) δ7.16 (d, J = 8.0Hz, 2H); 6.78 (d, J = 8.0Hz, 2H), 5.10-5.02 (m, 2H), 4.41 (d ,J=8.8Hz,1H),2.70(s,4H),2.72-2.62(m,1H),1.10(d,J=6.4Hz,3H),0.82(d,J=6.4Hz,3H); 13C NMR (101MHz, CDCl3) δ176.7,168.3,156.0,130.3,115.5,67.1,58.2,27.9,27.7,21.0,19.3; HRMS (ESI): Exactmass calcd for C 16 H 19 NO5, [M+H] + ,306.1336.Found 306.1330.

[0120] Example 14

[0121] Succinimide-Isoleucine-4-hydroxybenzyl ester (RJ-14)

[0122]

[0123] The preparation method of succinimide-isoleucine is the same as in Example 1, except that the raw material is replaced with isoleucine, and the other operation steps are the same.

[0124] Succinimide-isoleucine (1.06 g, 4.98 mmol) was added to 50 mL of dichloromethane in a 100 mL reaction flask and dissolved with stirring. 4-hydroxybenzyl alcohol (0.617 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4–6 h until the starting material disappeared. The pH was adjusted to 4–5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL * 2). The organic phase was collected and washed once each with 20 mL of aqueous phase and brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 1.5 g of crude product. The crude product was subjected to silica gel column chromatography with 50% ethyl acetate / petroleum ether as the eluent.

[0125] The product was 0.57 g of a colorless oil; yield 36.4%. 1 H NMR (400MHz, CDCl3) δ7.16(d,J=8.4Hz,2H),6.78(d,J=8.4Hz,2H),5.90(brs,1H),5.09-5.01(m,2H),4.55-4.49(m,1H),2.68(d, J=3.2Hz,4H),2.45-2.41(m,1H),1.40-1.33(m,1H),1.05(J=6.4Hz,2H),0.92(J=7.2Hz,1H),0.82(dd,J1=6.2Hz,J2=16.0Hz,3H); 13C NMR (101MHz, CDCl3) δ176.8,168.4,156.1,130.3,127.2,115.4,67.2,57.7,33.1,28.0,25.7,16.9,10.9; HRMS (ESI): Exact masscalcd for C 17 H 21 NO5, [M+Na] + ,342.1317.Found 342.1325.

[0126] Example 15

[0127] Succinimide-Isoleucine Vanillyl Ester (RJ-15)

[0128]

[0129] The preparation method of succinimide-isoleucine is the same as in Example 1, except that the raw material is replaced with isoleucine, and the other operation steps are the same.

[0130] Succinimide-isoleucine (1.06 g, 4.98 mmol) was added to a 100 mL reaction flask and dissolved in 50 mL of dichloromethane with stirring. Vanillin (0.767 g, 4.98 mmol), EDC (1.43 g, 7.46 mmol), and DMAP (122 mg, 1.0 mmol) were added, and the reaction was carried out at room temperature for 4–6 h until the starting material disappeared. The pH was adjusted to 4–5 with 2.0 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL * 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain the crude product. The crude product was subjected to silica gel column chromatography with 70% ethyl acetate / petroleum ether as the eluent. The product was 0.46 g of colorless oil, with a yield of 26.4%.

[0131] The ratio of product to isomer is approximately 2:1; 1 H NMR (400MHz, CDCl3) δ6.90-6.80(m,3H),5.75(s,1H),5.11-4.99(m,2H),4.55-4.48(m,1H),3.87(s,3H),2.70(s,4H) ,2.45-2.41(m,1H),1.40-1.33(m,1H),1.05(J=6.4Hz,2H),0.92(J=7.2Hz,1H),0.82(dd,J1=6.2Hz,J2=16.0Hz,3H); 13CNMR(101MHz, CDCl3)δ176.6,168.4,146.5,145.8,127.2,121.9,114.3,111.3,67.4,57.6,55.9,33.8,28.0,25.7,16.8,10.9; HRMS(ESI): Exact mass calcd for C 18 H 23 NO6, [M+H] + ,350.1598.Found 350.1591.

[0132] Example 16

[0133] Succinimide-glycine ethyl ester (RJ-16)

[0134]

[0135] In a 100 mL reaction flask, potassium succinimide (5.0 g, 50.5 mmol), ethyl glycine (5.72 g, 55.5 mmol), and 60 mL of dioxane were added. The mixture was heated to 100 °C and reacted for 6 hours. All solvent was then evaporated under reduced pressure. 20 mL of water was added to the residue, and the mixture was stirred for 20 min. The mixture was filtered, and the off-white solid collected was succinimide-ethyl glycine. The crude product was subjected to silica gel column chromatography with 60% ethyl acetate / petroleum ether as the eluent. The product was 9.4 g of a pale yellow solid, with a yield of 90.3%.

[0136] 1 H NMR (400MHz, CDCl3) δ4.24 (s, 2H), 4.20 (q, J = 7.2Hz, 2H), 2.79 (s, 4H), 1.27 (t, J = 7.2Hz, 3H); HRMS (ESI): Exact mass calcd for C8H 11 NO4, [M+H] + ,186.0671.Found186.0665.

[0137] Example 17

[0138] Succinimide-phenylglycine borneol ester (RJ-17)

[0139]

[0140] The preparation method of succinimide-phenylglycine is the same as in Example 1, except that the raw material is changed to phenylglycine, and the other operation steps are the same.

[0141] Succinimide-phenylglycine (1.0 g, 4.23 mmol) was added to a 100 mL reaction flask and dissolved with stirring in 50 mL of dichloromethane. Borneol (0.651 g, 4.23 mmol), EDC (1.23 g, 6.44 mmol), and DMAP (103 mg, 0.84 mmol) were added. The reaction was carried out at room temperature for 4-6 h until the starting material disappeared. The pH was adjusted to 4-5 with 2 mol / L hydrochloric acid, and the mixture was extracted with dichloromethane (30 mL x 2). The organic phase was collected and washed once with 20 mL of aqueous phase and once with brine. The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain 1.8 g of crude product. The crude product was subjected to silica gel column chromatography with 10%-40% dichloromethane / ethyl acetate as the eluent. The product was 1.25 g of colorless oil; yield 80.2%.

[0142] 1 H NMR (400MHz, CDCl3) δ7.51-7.49(m,2H),7.35-7.32(m,3H),5.84(d,J=2.4Hz,1H),4.98-4.91(m,1H),2.74(s,4H),2.42-2.3 1(m,1H),1.69-1.62(m,3H),1.49-1.46(m,1H),1.24-1.03(m,3H),0.88(s,3H),0.84(s,3H),0.82(s,1.5H),0.77(s,1.5H); 13 C NMR (101MHz, CDCl3) δ175.8,167.6,134.1,130.0,128.6,128.4,82.3,56.6,48. 9,47.0,44.8,36.6,36.2,28.2,27.8,27.0,19.6,18.8,13.4; HRMS(ESI):Exact mass calcd forC 22 H 27 NO4, [M+Na] + ,392.1832.Found 392.1841.

[0143] Bioactivity studies of succinimide derivatives

[0144] Succinimide derivatives are used to treat convulsions and epilepsy. This project uses the maximum electroconvulsive assay (MES) to further study and evaluate the antiepileptic activity of the drug, applies neurotoxicity experiments to evaluate the safety of the compound, uses the classic chemical model of pentylenetetrazole to study and evaluate the anticonvulsant pharmacological mechanism of the drug, employs the elevated maze test, open field test, and fear box to examine anxiety-like behavior and suggestible conditioned memory behavior in mice, observes the growth and morphological changes of hippocampal neurons in epileptic mice after drug administration using HE staining technology, and conducts hepatotoxicity tests to examine the effects of the compound on liver function in mice.

[0145] (1) Maximum Electroconvulsive Test (MES) and Neurotoxicity Test

[0146] MES Method: Mice were initially screened before the experiment, and only qualified mice were used in the next stage. The screening method was as follows: One day before the experiment, the mice were subjected to electrical stimulation at 50V and 50Hz. Two electrodes were clamped between the mice's ears, and the stimulation was applied for 0.2s. Mice that exhibited hind limb rigidity were designated as reserve mice for later experiments. The method for determining the anticonvulsant effect was as follows: After the test compound was dissolved, it was administered by gavage. 0.5 hours after administration, the mice were stimulated with ear electrodes at 50V and 50Hz for 0.2s. If the mice did not exhibit hind limb rigidity, it indicated that the test compound had anticonvulsant activity at that dose.

[0147] Neurotoxicity test method: Before the experiment, mice were trained on an XZC-4B rotating bar with a radius of 0.4 cm and a rotation speed of 24 rap / min. Training was conducted continuously for 2–3 days, 1–3 times per day, to ensure that all mice used in this model were adapted to the experimental conditions before drug administration. Mice that did not fall off the fatigue tester within 3 minutes or fell off no more than 3 times were selected as neurotoxicity test subjects. Then, qualified mice were randomly divided into groups, and a double-blind drug administration method was used. Mice in the drug administration group were administered 100 mg / kg of the test compound via intraperitoneal injection. At 0.5, 1, and 2 hours after administration, the mice were stably placed on the fatigue tester and rotated at 24 rap / min. If the test mouse did not fall off within 3 minutes or fell off no more than 3 times, it indicated that the compound had no neurotoxicity at that dose; if the number of falls exceeded 3 times, it indicated that the test compound had neurotoxicity at that dose. The number of mice exhibiting neurotoxic reactions was recorded to preliminarily evaluate the safety of the compound.

[0148] In the MES model, the antiepileptic activity of the compound was initially evaluated 0.5 h after intraperitoneal administration; in the neurotoxicity model, the neurotoxicity of the compound was evaluated 0.5 h after intraperitoneal administration (Table 1). As shown in Table 1, the compound did not show neurotoxicity at the experimental doses.

[0149] Table 1. Results of MES and neurotoxicity experiments on succinimide derivatives (n=3)

[0150]

[0151]

[0152] a Number of mice that did not experience forced seizures / Number of experimental mice;

[0153] b MES analysis was performed 0.5 h after intraperitoneal injection of each compound;

[0154] c Number of mice that did not develop neurotoxicity / Number of experimental mice;

[0155] In the MES model, the antiepileptic activity of the compounds was preliminarily evaluated 0.5 h after intraperitoneal administration. As shown in Table 1, the succinimide derivatives could protect mice from tetany under stimulation and had certain antiepileptic activity. In the neurotoxicity model, the neurotoxicity of the compounds was evaluated 0.5 h after intraperitoneal administration. As shown in Table 1, the compounds did not show neurotoxicity at the experimental doses.

[0156] (2) Experimental acute epilepsy model induced by pentylenetetrazol

[0157] Method 1: Mice were randomly divided into a treatment group (50 mg / kg), a positive control group, and a model group, with 3 mice in each group. The treatment group was injected intraperitoneally with the test compound, the control group was injected intraperitoneally with the positive drug sodium valproate, and the model group was administered physiological saline by gavage. 30 minutes later, the mice were injected subcutaneously with pentylenetetrazol 85 mg / kg. The test animals were placed in a cage alone and observed for 30 minutes. The latency time, seizure severity, number of clonic seizures, number of tonic seizures, and number of deaths of mice in each group were recorded (Table 2).

[0158] Table 2. Effects of succinimide derivatives on PTZ (85 mg / kg)-induced acute epileptic seizures in mice (n=3)

[0159]

[0160]

[0161] Data are expressed as mean ± standard deviation;

[0162] a Number of mice with generalized tonic-clonic seizures / Number of mice tested;

[0163] b Number of mice exhibiting clonic contractions / Number of mice tested;

[0164] cNumber of dead mice / Number of test mice;

[0165] The PTZ dose for the model group and the fourteen compound groups was 85 mg / kg.

[0166] Method 2: Mice were randomly divided into a treatment group (25 mg / kg, 75 mg / kg), a positive control group (sodium valproate, VPA), and a model group, with 3 mice in each group. The treatment group was injected intraperitoneally with the test compound, the control group was injected intraperitoneally with the positive drug VPA, and the model group was given physiological saline by gavage. 30 min later, the mice were subcutaneously injected with 85 mg / kg of pentylenetetrazol. The test animals were placed in a cage alone and observed for 30 min. The latency time, seizure severity, number of clonic seizures, number of tonic seizures, and number of deaths of mice in each group were recorded (Table 3).

[0167] Table 3. Effects of succinimide derivatives on PTZ (85 mg / kg)-induced acute epileptic seizures in mice (n=3)

[0168]

[0169]

[0170] Data are expressed as mean ± standard deviation;

[0171] a Number of mice with generalized tonic-clonic seizures / Number of mice tested;

[0172] b Number of mice exhibiting clonic contractions / Number of mice tested;

[0173] c Number of dead mice / Number of test mice.

[0174] According to Table 2, in the antagonistic experiment against pentylenetetrazol-induced acute seizures, compared with the model group, most succinimidide derivatives at the same dose prolonged the duration of clonic seizures, reduced the severity of clonic seizures, inhibited tonic seizures, and reduced mouse mortality. According to Table 3, most succinimidide derivatives at different doses also prolonged the duration of clonic seizures, reduced the severity of clonic seizures, inhibited tonic seizures, and reduced mouse mortality. This indicates that succinimidide derivatives may have the effect of inhibiting or reducing pentylenetetrazol-induced seizures in mice.

[0175] (3) Pentylenetetrazole-induced chronic epilepsy model in mice

[0176] Methods: Experimental mice were divided into a blank control group, a positive control group (VPA), and an epilepsy group. The blank control group was given the same volume of physiological saline. Mice in the epilepsy group were injected intraperitoneally with PTZ (35 mg / kg) every other day for 10 consecutive times. The seizures of the mice were observed and recorded for 30 minutes. When the mice exhibited 3 consecutive seizures after 3 consecutive subcutaneous injections of PTZ, the mice were considered successfully ignited and could be used in later experiments (Table 4). The successfully ignited mice were randomly divided into a drug administration group (25 mg / kg, 50 mg / kg, 75 mg / kg), a positive control group, and a model group, with 6 mice in each group. The drug administration group was injected intraperitoneally with the test compound, the positive control group was injected intraperitoneally with the positive drug sodium valproate, and the model group was administered physiological saline by gavage. 30 minutes later, the mice were injected subcutaneously with PTZ (35 mg / kg) and observed in a separate cage for 30 minutes. The latency time of clonic seizures, the number of clonic seizures, the number of tonic seizures, and the number of deaths were recorded for each group (Table 5).

[0177] Table 4. Effects of PTZ (35 mg / kg) on ​​chronic epileptic seizures in mice (n=6)

[0178]

[0179] Data are expressed as mean ± standard deviation.

[0180] Table 5. Effects of succinimide derivatives on PTZ (35 mg / kg)-induced chronic epileptic seizures in mice (n = 6)

[0181]

[0182] Data are expressed as mean ± standard deviation;

[0183] a Number of mice with generalized tonic-clonic seizures / Number of mice tested;

[0184] b Number of mice exhibiting clonic contractions / Number of mice tested;

[0185] c Number of dead mice / Number of test mice.

[0186] During the pentylenetetrazol modeling process, the duration of clonic seizures in mice gradually shortened until the model was successfully established. In the anti-pentylenetetrazole chemically induced seizure model experiment, compared with the model group, the duration of clonic seizures in mice in the positive drug group was significantly prolonged, and the severity of clonic seizures was reduced. Compared with the model group, the low, medium, and high doses of succinimide-valine borneol ester (RJ-9) all prolonged the duration of clonic seizures in mice. Compared with the model group, the high dose of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) prolonged the duration of clonic seizures in mice.

[0187] (4) Pentylenetetrazole-induced epilepsy model in zebrafish

[0188] Zebrafish juveniles were used in the experimental group, and the model group was administered PTZ. Mice in the positive control group were administered VPA (500 μM) + PTZ. The drug groups were administered succinimide-valine borneol ester (RJ-9) and succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14), respectively. After administration, the swimming behavior of zebrafish in each group was detected using a ZebraBox, and the total swimming distance and swimming speed were recorded. The effects of the compounds on epileptic behavior in zebrafish were evaluated.

[0189] In this experiment, four groups of juvenile zebrafish treated with the drug the previous day and one group of untreated zebrafish were transferred to 15 mmol PTZ and immediately placed in 48-well plates, one fish per well. The 48-well plates were then placed in a zebrafish behavior tracking system. After approximately 10 minutes of rest in darkness, zebrafish behavior was recorded for 30 minutes, with data collected every 5 minutes to track and record the behavioral trajectories of each group. Measurements using a zebrafish behavior analyzer showed that the compounds succinimide-valine borneol ester (RJ-9) and succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) did not produce toxic effects on zebrafish at the experimental doses. Figure 1 ).

[0190] Studies showed that the total movement distance of juvenile zebrafish in the PTZ model group increased, and their movement trajectory diagrams showed a state of hyperexcitation. Compounds RJ-9 and RJ-14 at 50 μM significantly reduced the movement distance of zebrafish exhibiting epileptic symptoms, demonstrating significant anti-epileptic activity. Figure 2 ).

[0191] (5) Elevated Cross Maze (EPMT) Test

[0192] On day 7 post-drug administration, behavioral assessments of epileptic mice were conducted using EPMT in a room with controlled sound and light. The elevated cruciate maze consisted of two opposing, equal-length open arms and a closed arm connected by a central area. All four arms were 50cm x 10cm long, and the central area was 10cm x 10cm, constructed from blue high-grade medical-grade organic board. The bottom of the maze was 50cm above the ground. At the start of the experiment, mice were placed in the maze with their heads facing away from the maze users, positioned in the central area facing the open and closed arms. The mouse's head and forelimbs entering an arm were considered "entering," and exiting an arm was considered "exiting." After each mouse's experiment, all four arms were carefully wiped with 75% ethanol, and mouse excrement was cleaned to avoid affecting subsequent experiments. Video analysis on a computer showed the number of times the mouse entered the open arm and the duration of its stay within 10 minutes, and the corresponding percentages were calculated to reflect anxiety-like behaviors.

[0193]

[0194] Table 6. Ratio of time and number of times mice entered the elevated cruciform maze on day 7 after drug administration.

[0195]

[0196]

[0197] Data are expressed as mean ± standard deviation.

[0198] In the elevated cross maze (EPMT) test, as shown in Table 6, on day 7 of drug administration, compared with the model group, the time taken for mice to enter the open arms of the elevated cross maze was increased in the positive control group, the low, medium, and high dose groups of succinimide-valine borneol ester (RJ-9), and the low and medium dose groups of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14). Compared with the model group, the number of times mice entered the open arms of the elevated cross maze was increased in the positive control group, the low, medium, and high dose groups of succinimide-valine borneol ester (RJ-9), and the low and high dose groups of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14). These results indicate that succinimide derivatives have the effect of improving anxiety-like behavior in PTZ-induced epilepsy model mice.

[0199] (6) Open Field (OFT) Test

[0200] The open field test (OFT) was performed on day 7 after drug administration. The open field test chamber was divided into nine equal-sized grids. Mice were placed in the central area, and the operator immediately moved away from the open field. The entire experiment was conducted in a well-ventilated, quiet, and dimly lit indoor space. A video analysis system was used to record the mice's movement distance and dwell time in the central area for 5 minutes to reflect anxiety-like behaviors. After each mouse's experiment, the experimental equipment was cleaned with 75% ethanol, and mouse excrement was removed.

[0201] Table 7. Distance and dwell time of mice in the central area of ​​the open field on day 7 after drug administration.

[0202]

[0203] Data are expressed as mean ± standard deviation;

[0204] * This indicates a significant difference from the model group (P < 0.05);

[0205] ## This indicates a significant difference compared to the control group (P < 0.01).

[0206] Mice with lower anxiety levels tended to move longer distances and stay longer in the central open area of ​​the open field. Table 7 shows that on day 7 of administration, compared to the control group, the model group mice moved less distance in the central open field area (P<0.01). Compared to the model group, the positive control drug and low, medium, and high doses of succinimide derivatives all increased the distance moved in the central open field area of ​​PTZ-induced epilepsy mice. Specifically, 50 mg / kg succinimide-valine borneol ester (RJ-9) and 25 mg / kg succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) significantly increased the distance moved in the central open field area of ​​PTZ-induced epilepsy mice (P<0.05), with statistically significant differences.

[0207] Table 7 shows that on day 7 of drug administration, compared with the control group, the model group mice had a significantly shorter dwell time in the central region of the open field (P<0.05), which was statistically significant. Compared with the model group, the positive control drug and low, medium, and high doses of succinimide derivatives all increased the dwell time in the central region of the open field in PTZ-induced epilepsy mice. Among them, the high dose of succinimide-valine borneol ester (RJ-9) significantly increased the dwell time in the central region of the open field in PTZ-induced epilepsy mice (P<0.05), which was statistically significant. The results indicate that succinimide derivatives can improve anxiety-like behavior, improve motor ability, and increase exploratory behavior in PTZ-induced epilepsy model mice.

[0208] (7) Fear Box (FCT) Experiment

[0209] The experiment was divided into two phases:

[0210] ① Training period: Mice were placed on the partition of the electric shock generator in the fear chamber for 3 minutes to adapt. The electric shock was applied at a magnitude of 0.35 mA, with an interval of 62 seconds and a duration of 2 seconds, repeated 3 times. ② Testing period: Mice were placed in the fear chamber in the same environment as the training period for 3 minutes to adapt without stimulation. The rigidity time and frequency of the mice in the fear chamber for 6 minutes were analyzed using a video analysis system. After each mouse experiment, mouse excrement was removed, the area was wiped with 75% ethanol, and allowed to dry completely before subsequent experiments.

[0211] Table 8. Duration and frequency of rigidity in the fear box during the training period for mice in each group on day 7 after drug administration.

[0212]

[0213]

[0214] Data are expressed as mean ± standard deviation;

[0215] *This indicates a significant difference from the model group (P < 0.05); ** This indicates a significant difference from the model group (P < 0.01); *** This indicates a significant difference from the model group (P < 0.001);

[0216] ## This indicates a significant difference compared to the control group (P < 0.01). ### This indicates a significant difference compared to the control group (P < 0.001).

[0217] Table 8 shows that during the Fear Box (FCT) training period, compared with the model group, the PTZ-induced epilepsy mice in the positive control group, the low, medium, and high dose groups of succinimide-valine borneol ester (RJ-9), and the low and medium dose groups of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) all showed an increase in the time of rigidity in the fear box. The high dose group of succinimide-valine borneol ester (RJ-9) showed a significant increase in the time of rigidity in the fear box (P<0.0001), indicating a statistically significant difference. The positive control group and the low, medium, and high dose groups of succinimide derivatives all showed an increase in the number of rigidity episodes in the fear box.

[0218] During the Fear Box (FCT) experiment, compared with the control group, the model group mice showed a significant decrease in both the duration and number of periods of immobility within the fear box (P < 0.01). Compared with the model group, the positive control group and the low, medium, and high dose groups of succinimide derivatives all showed an increase in both the duration and number of periods of immobility in the fear box. Among these, the positive control group and the medium dose group of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) showed a significant increase in the duration of immobility in the fear box (P < 0.0001). The medium dose group of succinimide-valine borneol ester (RJ-9) and the high dose group of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) also showed a significant increase in the duration of immobility in the fear box (P < 0.01), and the differences were statistically significant. The positive control group, the low and medium dose groups of succinimide-valine borneol ester (RJ-9), and the medium and high dose groups of succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) significantly increased the number of rigid immobilities in the fear box (P < 0.0001); the high dose group of succinimide-valine borneol ester (RJ-9) significantly increased the number of rigid immobilities in the fear box (P < 0.01), and the differences were statistically significant.

[0219] Numerous brain nuclei are involved in fear memory, among which scene-based fear memory is related to the activity of hippocampal neurons in mice. In PTZ-induced epilepsy mice, seizures lead to hippocampal damage and neuronal apoptosis. Measuring the establishment and extinction of scene-based memories in PTZ-induced epilepsy mice using the fear box experiment is beneficial for exploring the protective effect of succinimide derivatives on hippocampal neurons in PTZ-induced epilepsy mice.

[0220] Table 8 shows that compared with the control group, the model group had a reduced duration and frequency of immobility in PTZ-induced epilepsy mice, indicating a decline in scene-related fear memory. This phenomenon may be due to neuronal damage in PTZ-induced epilepsy mice. Compared with the low, medium, and high dose groups of succinimide derivatives, continuous treatment with succinimide derivatives in PTZ-induced epilepsy mice increased the duration and frequency of immobility, suggesting that succinimide derivatives can improve scene-related fear memory impairment in PTZ-induced epilepsy model mice, indicating that it has a certain protective effect on hippocampal neurons.

[0221] (8) Results of HE staining of mouse hippocampus

[0222] HE staining, 200×, succinimide-valine borneol ester (RJ-9), succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14), Figure 3 (Left: 25mg / kg, Center: 50mg / kg, Right: 75mg / kg; Left, Center, and Right represent the three positions respectively).

[0223] Depend on Figure 3 It was found that, compared with the control group, the model group mice experienced more severe neuronal apoptosis in the CA1 region of the hippocampus. Compared with the model group, the positive control drug and low, medium, and high doses of succinimide derivatives all reduced neuronal apoptosis in the CA1 region of the hippocampus in mice, with the reduction being more significant in the positive control drug group. This indicates that succinimide derivatives can improve neuronal apoptosis in the CA1 region of the hippocampus in PTZ-induced epilepsy model mice.

[0224] (9) Compound hepatotoxicity detection

[0225] ALT and AST are mainly found within hepatocytes. When hepatocytes are damaged or destroyed, these enzymes leak from the cells into the bloodstream, leading to increased blood concentrations. Serum ALT and AST are early markers of acute liver injury, possessing high specificity and sensitivity, and are most commonly used clinically to reflect the degree of liver function impairment; their levels are directly proportional to the degree of liver damage. Therefore, this experiment detected liver ALT and AST in mice after drug administration. The detection method is as follows: After the experiment, blood was collected by enucleation, stored at 4°C for 2 hours, and then centrifuged at 3000 r / min (centrifugation radius 60 mm) for 5 minutes at 4°C to obtain serum. The levels of ALT and AST in the serum were identified using a multi-mode microplate reader, following the instructions for the alanine aminotransferase (ALT) and aspartate aminotransferase (AST) kits.

[0226] As shown in Table 9, the average ALT and AST levels in the model group were higher than those in the control group. The results indicate that the compounds succinimide-valine borneol ester (RJ-9) and succinimide-isoleucine 4-hydroxybenzyl ester (RJ-14) have no significant liver toxicity.

[0227] Table 9 Effects of succinimide derivatives on liver function in mice

[0228]

[0229] In summary, the study found that succinimide derivatives have a relatively high safety profile and significant anticonvulsant, antiepileptic, antineural pain, antidepressant, neuroprotective, brain injury protective, memory-improving, and anti-anxiety activities, making them worthy of further development and use.

[0230] The above descriptions are merely embodiments of the present invention, and common knowledge regarding specific structures and characteristics is not elaborated upon here. It should be noted that those skilled in the art can make various modifications and improvements without departing from the structure of the present invention, and these should also be considered within the scope of protection of the present invention. These modifications and improvements will not affect the effectiveness of the present invention or the practicality of the patent. The scope of protection claimed in this application should be determined by the content of its claims, and the specific embodiments described in the specification can be used to interpret the content of the claims.

Claims

1. Succinimidide derivatives, characterized in that: Its general structural formula is shown in equation V: ； Where: R3 is H; R4 is H; X is O; n is 0 or 1; When R2 is (+)-2-camphenyl, R1 is methoxymethyl, hydrogen, isopropyl, methyl, n-propyl, benzyl, isobutyl or phenyl; When R2 is 4-hydroxyphenyl or 3-methoxy-4-hydroxyphenyl, R1 is methoxymethyl, hydrogen, isopropyl, methyl, n-propyl or isobutyl.

2. The succinimide derivative according to claim 1, characterized in that: The succinimide derivatives are RJ-1, RJ-2, RJ-3, RJ-4, RJ-5, RJ-6, RJ-9, RJ-10, RJ-11, RJ-13, RJ-14, RJ-15 or RJ-17; The structural formula of succinimide derivative RJ-1 is shown below: ； The structural formula of the succinimide derivative RJ-2 is shown below: ； The structural formula of the succinimide derivative RJ-3 is shown below: ； The structural formula of the succinimide derivative RJ-4 is shown below: ； The structural formula of the succinimide derivative RJ-5 is shown below: ； The structural formula of the succinimide derivative RJ-6 is shown below: ； The structural formula of the succinimide derivative RJ-9 is shown below: ； The structural formula of the succinimide derivative RJ-10 is shown below: ； The structural formula of the succinimide derivative RJ-11 is shown below: ； The structural formula of the succinimide derivative RJ-13 is shown below: ； The structural formula of the succinimide derivative RJ-14 is shown below: ； The structural formula of the succinimide derivative RJ-15 is shown below: ； The structural formula of the succinimide derivative RJ-17 is shown below: 。 3. The method for preparing the succinimide derivative according to claim 1, characterized in that: The succinimide derivative V is obtained by reacting a compound of general formula III with a compound of general formula IV under a condensing agent, as shown in the following reaction formula: ； The compound of general formula III is obtained by reacting the compound of general formula I with the compound of general formula II, as shown in the following reaction formula: ； The specific preparation process of the compound of general formula III is as follows: the compound of general formula I and the compound of general formula II are added to the reaction vessel in a molar ratio of 1.0 to 2.0:1.0, and reacted at 70-100°C until the compound of general formula I and / or the compound of general formula II disappear; the solvent is removed by vacuum concentration, water is added to the residue to precipitate, the residue is filtered, and dried at 30-60°C to obtain the compound of general formula III.

4. The method for preparing succinimide derivatives according to claim 3, characterized in that: In the preparation of succinimide derivative V: Compound of general formula III and compound of general formula IV, condensing agent and catalyst are added sequentially to the reaction vessel and reacted. The molar ratio of compound of general formula III to compound of general formula IV is 1.0:1.0~1.

5. The reaction is carried out at 5℃-40℃ until compound of general formula III and / or compound of general formula IV is consumed. After washing with pure water, the mixture is extracted with ethyl acetate, concentrated under vacuum to remove ethyl acetate, and the organic phase is dried, concentrated and purified by column chromatography.

5. The method for preparing succinimide derivatives according to claim 4, characterized in that: The condensing agent is DCC, DIC, EDC, BOP, PyBOP, HATU, HBTU, TBTU, ethyl chloroformate, phenyl chloroformate, isopropyl chloroformate, or 1-n-propylphosphonic anhydride.

6. The method for preparing succinimide derivatives according to claim 5, characterized in that: The catalyst is: Et3N, DIEA, DMAP, DPPY, 2,6-dimethylpyridine, DABCO, or DBU.

7. The racemic mixture of the succinimide derivative according to claim 1.

8. The use of the succinimide derivative of claim 1 and / or the racemic mixture of the succinimide derivative of claim 7 in the preparation of anticonvulsant, antiepileptic, anti-anxiety and / or brain injury protective drugs.

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