Corydalis bungeana tetrahydropyrrolizine [2, 3-c] quinoline alkaloid compound as well as preparation and application thereof

Abstract

The invention relates to a tetrahydro pyrrololizine [2, 3-c] quinoline alkaloid compound with a novel structure, a derivative of the tetrahydro pyrrololizine [2, 3-c] quinoline alkaloid compound, a preparation method of the tetrahydro pyrrololizine [2, 3-c] quinoline alkaloid compound, and an application of the tetrahydro pyrrololizine [2, 3-c] quinoline alkaloid compound and the derivative of the tetrahydro pyrrololizine [2, 3-c] quinoline alkaloid compound. The new compound, namely, the ungeanoline G, provided by the invention, is obtained by extracting and separating from a Corydalis bungeana Turcz which is a Corydalis bungeana belonging to papaveraceae, and the new compound, namely, the ungeanoline G, provided by the invention, is obtained by extracting and separating from the Corydalis bungeana Turcz which is a plant belonging to papaveraceae. The invention further provides a method for artificially synthesizing the bungeanoline G, and a series of derivatives of the bungeanoline G are prepared. A biological activity experiment shows that the compound has acetylcholin esterase and butyrylcholin esterase inhibition activity and can be applied to preparation of drugs for treating or improving various neurodegenerative diseases such as Alzheimer's disease, vascular dementia and myasthenia gravis.

Description

Technical Field

[0001] The present invention relates to a novel tetrahydropyrrolizino[2,3-c]quinoline alkaloid compound and a preparation method thereof, as well as use of the tetrahydropyrrolizino[2,3-c]quinoline alkaloid compound as an acetylcholinesterase and / or butyrylcholinesterase inhibitor for treating or improving neurodegenerative diseases such as Alzheimer's disease, vascular dementia, myasthenia gravis and the like. Background Art

[0002] Alzheimer's disease (AD) is a chronic neurodegenerative disease that seriously affects the health of the elderly and has become a global public health problem. According to the World Health Organization (WHO), more than 55 million people worldwide suffer from dementia, and AD is the most common form of dementia, accounting for 60-70% of all cases. Natural alkaloids are an important source for the development of anti-AD drugs. Among the four anti-AD drugs approved by the US FDA, the skeleton structures of two of them, rivastigmine and galantamine, are derived from natural alkaloids. In recent years, the discovery of new anti-AD drugs from cholinesterase inhibitors has become a hot spot in the development of AD drugs. The latest research shows that dual inhibitors of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) have better efficacy and fewer side effects in the treatment of AD than single AChE inhibitors. One of the important reasons is that BChE plays a vital role in regulating acetylcholine levels in the brains of patients with advanced AD. At present, the research and development of anti-AD drugs is overly dependent on the structures of known bioactive compounds. Although some target compounds have been found in the process of anti-AD lead compound development, they are often just simple structural modifications of existing cholinesterase inhibitors. Therefore, the discovery of novel dual cholinesterase inhibitors with unique carrier skeletons from natural alkaloids has important research significance and value for the treatment of AD.

[0003] Corydalis bungeana Turcz. is the dried whole herb with roots of Corydalis bungeana Turcz., a plant of the genus Corydalis in the family Papaveraceae. It is also known as Corydalis bungeana, Corydalis bungeana, and Corydalis bungeana. It tastes bitter and is cold in nature. It has the effects of clearing away heat and detoxifying, dispersing knots and relieving swelling. It is mainly used for epidemic colds and sore throats. Modern studies have shown that Corydalis bungeana has the effects of inhibiting the body's immune function, anti-inflammatory, sedative and hypnotic. Alkaloids are the main biologically active ingredients in Corydalis bungeana. It is reported that Corydalis bungeana alkaloids have multiple pharmacological effects such as analgesia and anti-inflammatory, sedative and hypnotic, antiviral, antibacterial, immune suppression, and cytotoxicity.

[0004] The tetrahydropyrrolizino[2,3-c]quinoline alkaloid compound (bungeanoline G) of the present invention is a new compound extracted and separated from corydalis for the first time, which has not been reported in other natural sources, and there is no report on the compound and its pharmacological activity in the existing literature. Because bungeanoline G has a unique 6 / 6 / 5 / 5 tetracyclic fused tetrahydropyrrolizino[2,3-c]quinoline skeleton, the present invention chemically synthesizes bungeanoline G and prepares its derivatives. Pharmacological activity tests show that the compound and its derivatives have significant acetylcholinesterase and butyrylcholinesterase inhibitory activities, and can be used in the preparation of drugs for treating or improving diseases such as Alzheimer's disease, vascular dementia, and myasthenia gravis. Summary of the invention

[0005] One of the purposes of the present invention is to provide a novel tetrahydropyrrolizino[2,3-c]quinoline alkaloid compound and a preparation method thereof. These compounds have significant acetylcholinesterase and butyrylcholinesterase inhibitory activity and have application prospects in the development of therapeutic drugs for neurodegenerative diseases that are currently receiving much attention, such as Alzheimer's disease, vascular dementia, myasthenia gravis, etc.

[0006] To achieve the above-mentioned object, the present invention provides a series of tetrahydropyrrolizino[2,3-c]quinoline alkaloid derivatives, and pharmaceutically acceptable salts, solvates, hydrates or crystalline forms thereof, the general formula (I) of which is as follows:

[0007]

[0008] in,

[0009] R1-R4 are independently H, halogen, C 1-3 Alkyl, hydroxyl, C 1-3 One or two or more alkoxy groups;

[0010] n is 1, 2, or 3.

[0011] In some embodiments, R1-R4 of compound I are each independently one or two or more of H, F, Cl, Br, I, methyl, ethyl, propyl, isopropyl, hydroxy, methoxy, ethoxy, propoxy, and isopropoxy; further, R1-R4 are each independently one or two or more of H, F, Br, methyl, ethyl, propyl, hydroxy, methoxy, ethoxy, and propoxy; preferably, R1-R4 are each independently one or two or more of H, Br, methyl, ethyl, methoxy, and ethoxy.

[0012] In some embodiments, n of Compound I is independently 1, 2, or 3; preferably, n is independently 1 or 2.

[0013] In some embodiments, compound I is a racemate, or one or both of 7aR and 7aS, as shown in the following formulas: I-achiral, I-7aS and I-7aR:

[0014]

[0015] In some embodiments, the preferred compounds include one or two or more of the following:

[0016]

[0017]

[0018] In some embodiments, the salt of Compound I and a pharmaceutically acceptable acid includes one or more of HCl, HBr, trifluoroacetic acid, sulfuric acid, formic acid, acetic acid, succinic acid, fumaric acid, p-toluenesulfonic acid, methanesulfonic acid, trifluoromethanesulfonic acid, and citric acid, and its general formula is shown in IX:

[0019]

[0020] Another object of the present invention is to provide a composition comprising any one or more compounds of the present invention and pharmaceutically acceptable adjuvants, carriers, diluents and / or other active compounds, which can be used to prepare a pharmaceutical preparation containing at least one compound of the present invention.

[0021] The present invention also relates to the use of any one of the compounds I or a pharmaceutical composition thereof as an active ingredient in the preparation of a drug for treating or improving diseases such as Alzheimer's disease, vascular dementia, myasthenia gravis, etc. by inhibiting acetylcholinesterase and / or butyrylcholinesterase.

[0022] Compounds according to the present invention or salts generated with pharmaceutically acceptable acids can be made into suitable galenic dosage forms, such as oral, injection, spray administration compositions, etc., according to acceptable pharmaceutical procedures. Pharmaceutical compositions according to the present invention include effective amounts of compounds of the present invention, and suitable pharmaceutically acceptable carriers or diluents, which are well known in the art. Carriers can be any inert raw materials, organic or inorganic, suitable for enteral, percutaneous or parenteral administration, such as water, gelatin, gum arabic, lactose, microcrystalline fiber starch, calcium hydrogen phosphate, magnesium stearate, talcum, colloidal silicon dioxide, etc. The composition can also contain other pharmaceutically active agents, and conventional additives, such as stabilizers, wetting agents, emulsifiers, flavoring agents, buffers, etc.

[0023] The composition according to the present invention can be prepared into solid or liquid dosage forms for oral administration, such as tablets, capsules, powders, syrups; prepared into sterile solutions, suspensions or emulsions for parenteral administration; prepared into dry powder preparations, sterile solutions, suspensions or emulsions for spray administration.

[0024] Another object of the present invention is to provide the use of the compounds of the present invention in the preparation of treatments for various types of neurodegenerative diseases. The dosage of a specific compound will vary depending on its potency, mode of administration, age and weight of the patient, and severity of the condition being treated.

[0025] The present invention discloses a novel tetrahydropyrrolizino[2,3-c]quinoline alkaloid compound and derivative thereof, and a preparation method and use thereof. The novel compound bungeanoline G provided by the present invention is extracted and separated from Corydalis bungeana Turcz. of the Papaveraceae family. The present invention also proposes a method for artificially synthesizing bungeanoline G, and prepares a series of derivatives of bungeanoline G. Biological activity experiments show that the compound has acetylcholinesterase and butyrylcholinesterase inhibitory activity, and can be used in the preparation of drugs for treating or improving various neurodegenerative diseases such as Alzheimer's disease, vascular dementia, and myasthenia gravis. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 , natural compound I-1 1 H- 1 H COSY and key HMBC;

[0027] Figure 2 , characterization of BChE inhibitory activity of representative compound I;

[0028] Figure 3 , Characterization of AChE inhibitory activity of representative compound I;

[0029] Figure 4 , schematic diagram of the structure of compound I of the preferred embodiment of the present invention. DETAILED DESCRIPTION

[0031] The following will describe the implementation methods of the present invention in detail with reference to the accompanying drawings and embodiments, so that the implementation process of how the present invention applies technical means to solve technical problems and achieve technical effects can be fully understood and implemented accordingly.

[0032] It should be understood that when used in this specification and the appended claims, the terms "include" and "comprises" indicate the presence of described features, integers, steps, operations, elements and / or components, but do not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components and / or combinations thereof.

[0033] It should also be understood that the terms used in the description of the embodiments of the present invention are only for the purpose of describing the specific embodiments and are not intended to limit the embodiments of the present invention.

[0034] The present invention also reports the preparation method of the above-mentioned compound I. With reference to similar literature reports or the specific examples shown below, experienced researchers can easily select the appropriate reaction conditions for the chemical reaction. The raw materials required for such reactions can be purchased or prepared by themselves according to conventional methods.

[0035] According to the conditions disclosed in the present invention, the following method is provided:

[0036] Natural product extraction methods

[0037] The invention provides a method for preparing a natural compound bungeanoline G (I-1), and the steps are as follows: grinding the dried whole herb with roots of corydalis quinata to obtain a medicinal material powder, heating and extracting with 50% to 90% ethanol at 60 to 70°C for 6 to 24 hours, filtering the extract with an 80-mesh filter, and concentrating the extract under reduced pressure to obtain an extract; the extract is dissolved in 0.01 to 0.2 mol / L sulfuric acid water, adjusting the pH to 2 to 3, adding petroleum ether at a volume ratio of 1:1 to 1:2 to extract, and taking a water layer. The obtained water layer is adjusted to a pH of 9 to 10 with 0.01 to 2 mol / L sodium hydroxide, and dichloromethane is added at a volume ratio of 1:1 to 1:2 to extract, and a dichloromethane layer is obtained. The dichloromethane layer is separated by multi-step high-efficiency preparative liquid chromatography to obtain the compound bungeanoline G (I-1).

[0038] The present invention also provides a method for artificially synthesizing the above-mentioned compound I or a pharmaceutically acceptable salt thereof, and the synthesis method is as follows:

[0039] Synthesis method 1

[0040]

[0041] wherein R1-R4 and n are the same as defined in the above claims 1-4.

[0042] Reaction a

[0043] Substrate 1 (1 mmol, 1.0 molar equivalent) was dissolved in THF (10 ml), followed by the addition of substrate 2 (1.5 molar equivalent relative to substrate 1), Pd(PPh3)2Cl2 (0.03 molar equivalent relative to substrate 1), CuI (0.03 molar equivalent relative to substrate 1), PPh3 (0.03 molar equivalent relative to substrate 1) and triethylamine (2 molar equivalent relative to substrate 1), and the mixture was stirred and dissolved at room temperature. After the reaction system was replaced with a nitrogen atmosphere, it was heated to 60°C and stirred for 12 hours. After the reaction was complete as monitored by LC-MS, the reaction solution was cooled to room temperature, filtered, the filtrate was concentrated by rotary evaporation, and the concentrate was purified by silica gel column chromatography to obtain product 3 (as substrate 3).

[0044] Reaction b

[0045] Substrate 3 (1 mmol, 1.0 molar equivalent) was dissolved in acetone (15 ml), and water (5 ml), mercuric sulfate (1.5 molar equivalent relative to substrate 3) and (mass concentration 98%) concentrated sulfuric acid (2 molar equivalent relative to substrate 3) were added in sequence under stirring, and stirred at 30° C. for 72 hours. After the reaction was completed as monitored by LC-MS, saturated sodium carbonate solution was added to the reaction solution to neutralize sulfuric acid to pH 8, and extracted 3 times with ethyl acetate (10 ml of ethyl acetate each time), and the organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the corresponding product 4 (as substrate 4).

[0046] Reaction c

[0047] Substrate 4 (1 mmol, 1.0 molar equivalent) was dissolved in methanol (10 ml), acetic acid (4 molar equivalent relative to substrate 4) and benzylamine (4 molar equivalent relative to substrate 4) were added, and the mixture was stirred at 25°C for 8 hours, and sodium cyanoborohydride (4 molar equivalent relative to substrate 4) was added, and the mixture was stirred at 25°C for 16 hours. Water (10 ml) was added to the reaction solution and stirred for 0.5 hours, and ethyl acetate (10 ml ethyl acetate was used each time) was washed 3 times, and the organic phase was discarded. Concentrated hydrochloric acid (mass concentration 38%) was added to the aqueous phase to adjust the pH to 2, and palladium carbon (specification 10wt%, 0.2 mass equivalent relative to substrate 4) was added, and the mixture was stirred at 25°C in hydrogen at normal pressure for 16 hours. Saturated sodium carbonate solution was added to the reaction solution until the pH was 8, and ethyl acetate (10 ml ethyl acetate was used each time) was extracted 3 times, and the organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product I.

[0048] Reaction

[0049] Substrate 4 (1 mmol, 1.0 molar equivalent) was dissolved in methanol (10 ml), acetic acid (4 molar equivalents relative to substrate 4) and p-methoxybenzylamine (4 molar equivalents relative to substrate 4) were added, and the mixture was stirred at 25°C for 2 hours. Sodium cyanoborohydride (4 molar equivalents relative to substrate 4) was added, and the mixture was stirred at 25°C for 16 hours. Methanol was removed from the reaction solution under reduced pressure, saturated sodium carbonate solution was added to pH 8, and the mixture was extracted 3 times with ethyl acetate (10 ml each time), and the organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the corresponding product 5 (as substrate 5).

[0050] Reaction

[0051] The substrate 5 (1 mmol, 1.0 molar equivalent) was dissolved in acetonitrile (5 ml) and water (20 ml), and 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ, 1.5 molar equivalent relative to substrate 5) was added, and the mixture was heated to 70°C and stirred for 12 hours. The acetonitrile was removed from the reaction solution under reduced pressure, and 2N hydrochloric acid (5 ml) and an equal volume of ethyl acetate were added. The mixture was separated, and the organic phase was extracted three times with 2N hydrochloric acid (5 ml each time). The organic phase was discarded, the acid-water phases were combined, and the pH was adjusted to 8 with a saturated sodium carbonate solution, and the mixture was extracted three times with ethyl acetate (5 ml each time). The organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product I.

[0052] When all R1-R4 in compound I are not Br, and at least one is H, the derivative of compound I can be further synthesized by using synthesis method 2 as shown below:

[0053] Synthesis method 2

[0054]

[0055] Br in compound Ia may be substituted at any or any two positions of R2-R4, R refers to other R1-R4 substituents other than those substituted by Br under reaction condition f, R' in Ib refers to the group after replacing the Br group under reaction condition g; "alkyl boronic acid or boronic ester" refers to C 1-3 Alkylboronic acid or boric acid ester; Compounds Ia and Ib are within the scope of the definition of Compound I.

[0056] Reaction f

[0057] Compound I (substrate I) (1 mmol, 1.0 molar equivalent) was dissolved in acetic acid (10 ml), N-bromosuccinimide (NBS, 1.2 molar equivalent relative to substrate I) was added under ice bath, stirred at room temperature for 1 hour, and saturated sodium sulfite solution was added to the reaction solution to quench the reaction. The reaction solution was concentrated under reduced pressure, saturated sodium carbonate solution was added to alkaline (pH 8), extracted 3 times with dichloromethane (10 ml each time), the organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product Ia (as compound Ia).

[0058] Reaction g

[0059] Compound Ia (1 mmol, 1.0 molar equivalent) was dissolved in dioxane (10 ml), and water (2 ml), C 1-3 Alkylboronic acid or boric acid ester substrate (3 molar equivalents relative to substrate Ia), Pd(PPh3)4 (0.05 molar equivalents relative to substrate Ia) and potassium carbonate (3 molar equivalents relative to substrate Ia) were stirred at 90°C for 16 hours under nitrogen protection. The reaction solution was cooled to room temperature and filtered, the filter cake was washed with ethyl acetate, and the filtrate was washed 3 times with saturated brine (10 ml each time), dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product Ib.

[0060] When compound I is a chiral monomer, the racemate can be separated by chiral chromatography:

[0061] Split method

[0062] The racemic compound I is dissolved in methanol and separated and purified by supercritical fluid chromatography through BH (chiral chromatography column). The mobile phase A is CO2, and B is (volume concentration 0.1%) triethylamine-methanol. The gradient elution condition is 0-12min: 80% A. The chiral monomers IS and IR are obtained by collecting according to the chromatographic peaks.

[0063] When Compound I forms a salt with a pharmaceutically acceptable acid (HX) (these acids refer to one or more of HCl, HBr, trifluoroacetic acid, sulfuric acid, formic acid, acetic acid, succinic acid, fumaric acid, p-toluenesulfonic acid, methanesulfonic acid, trifluoromethanesulfonic acid, and citric acid), Compound I is mixed with an appropriate amount of HX:

[0064]

[0065] Reaction h

[0066] Compound I (1 mmol, 1.0 molar equivalent) was dissolved in dioxane or tetrahydrofuran or dichloromethane or ethyl acetate (5 ml), HX (1.2 molar equivalent relative to substrate I) was added, stirred at room temperature for 1 hour, concentrated under reduced pressure, the precipitated solid was filtered and dried to obtain the target product IX.

[0067] The following examples are intended to illustrate the present invention rather than to further limit the present invention. The present invention can be implemented in any manner described in the present invention.

[0068] Preparation Example of Natural Compound I-1 of the Present Invention:

[0069] Compound preparation and structure identification:

[0070] In the following preparation examples, the preparation system includes Waters Alliance, including e2695 separation unit, 2998 PDA detector, data processing is performed by empower 3; Waters AutoP automatic purification system, including 2545 separation unit, 2767 sample manager, 2489 dual wavelength detector; Novase HPLC industrial grade high pressure liquid phase preparative chromatography and HIPERSEP software. Reagents including chromatographic grade methanol and acetonitrile were purchased from Fisher Scientific (Loughborough, UK), chromatographic grade formic acid, acetic acid, ammonia, triethylamine, ammonium formate, sodium monohydrogen phosphate, sodium dihydrogen phosphate were purchased from J&K Technology (Hebei, China), laboratory water was from Milli-Q ultrapure water purification system (Billerica, MA, USA), and preparation grade methanol was purchased from Shanghai Xingke High Purity Solvent Co., Ltd. (Shanghai, China). Preparative columns and semi-preparative columns: C18HCE (15 μm, 100 mm × 250 mm), C18HCE (10 μm, 100 mm × 325 mm), C18CE (7 μm, 50 mm × 250 mm), FC8HL (3.5 μm, 30 mm × 150 mm), C18HCE (5 μm, 10 mm × 150 mm) and BH (5 μm, 4.6 mm × 150 mm) (Dalian Sipu Precision Industry Co., Ltd.).

[0071] The nuclear magnetic resonance spectra used for structural identification were determined by Bruker AVIII-600 nuclear magnetic resonance spectrometer (Bruker, Germany), and the compounds were dissolved in deuterated methanol (MeOD) or deuterated chloroform (CDCl3). The mass spectrometry was performed by Agilent 1290 Infinity LC / 6540 Q-TOF MS liquid chromatography-mass spectrometry system for sample separation and analysis.

[0072] The preparation and compound identification steps are as follows:

[0073] (1) Extraction of medicinal materials: Take the dried whole herb with roots of Corydalis eryngii, and crush it to obtain a total of 100 kg of medicinal material powder. Use 1000 liters of 70% ethanol by volume and heat it at 70°C for 6 hours. After filtering the extract through an 80-mesh filter, remove the solvent by rotary evaporation to obtain an extract. The extract is dissolved in 0.01 mol / L sulfuric acid water, and a 2% sulfuric acid aqueous solution is added to adjust the pH to 2. Petroleum ether is added in a 1:1 volume ratio to extract the water layer. The obtained water layer is adjusted to a pH of 10 with 2 mol / L sodium hydroxide, and dichloromethane is added in a 1:1 volume ratio to extract the water layer. The obtained dichloromethane layer is the crude alkali.

[0074] (2) The crude alkali obtained in step (1) was separated and purified in the first dimension using C18HCE (15um, 100×250mm, reverse phase chromatography column) at a flow rate of 320mL / min. The mobile phase A was formic acid-methanol (volume concentration 0.1%), and the mobile phase B was formic acid-water (volume concentration 0.1%). The gradient elution conditions were: 0-8min: 25%A (volume ratio, the same below), 8-23min: 40%A, 23-38min: 80%A, 38-55min: 100%A. The distillate was taken at a flow time of 4.2-26.0min and concentrated as a total alkaloid sample of Corydalis bispinosa.

[0075] (3) The total alkaloids of bitter herbs obtained in step (2) were separated and purified in the second dimension using C18HCE (10um, 100×325mm, reverse phase chromatography column) at a flow rate of 320mL / min. The mobile phase A was formic acid-methanol (volume concentration 0.1%), and the mobile phase B was formic acid-water (volume concentration 0.1%). The gradient elution conditions were: 0-5min: 20%A, 5-50min: 20-45%A, 50-60min: 95%A. Eleven sub-fractions F1 ( 5.7~10.7min), F2 (10.7~17.1min), F3 (17.1~20.9min), F4 (20.9~25.7min), F5 (25.7~29.2min), F6 (29.2~33 .0min), F7 (33.0~36.3min), F8 (36.3~40.4min), F9 (40.4~44.4min), F10 (44.4~55.6min) and F11 (55.6~60min).

[0076] (4) The sub-fraction F4 obtained in step (3) was separated and purified in the third dimension using C8CE (7 μm, 50×250 mm, reverse phase chromatography column), the mobile phase A was (volume concentration 0.1%) 25% ammonia-methanol, B was (volume concentration 0.1%) 25% ammonia-water, the gradient elution conditions were 0-5 min: 5% A, 5-50 min, 5-95% A, 50-60 min: 95% A, and 15 sub-fractions were collected according to the corresponding distillate outflow time to obtain F4-1 (4.0-7.8 min), F4-2 (7.8-10.1 min), F4-3 (10.1-13.4 min), F4-4 (15.3-17.9 min), F4-5 (16.7-18.9 min), F4-6 (17.8-20.9 min), F4-7 (18.9-21.3 min), F4-8 (19.1-23.4 min), F4-9 (20.2-24.7 min), F4-10 (21.3-25.9 min), F4-11 (22.4-24.8 min), F4-12 (23.5-25.7 min), F4-13 (24.6-26.9 min), F4-14 (25.8-27.9 min), F4-15 (26.7-28.9 min), F4-16 (25.8-29.9 min), F4-17 (26.8-30.9 min), F4-18 (25.8-31.9 min), F4-19 (26.9-32.9 min), F4-21 (26.9-33.9 min), F4-22 (26.9-34.9 min), F4- (13.4~18.1min), F4-5 (18.1~21.3min), F4-6 (21.3~23.7min), F4-7 (23.7~28.5min), F4-8 (28.5~30.7min), F4-9 (30.7~33.6min), F4-10 (33.6~35.0min), F4-11 (35.0~37.6min), F4-12 (37.6~39.5min), F4-13 (39.5~43.9min), F4-14 (43.9~46.1min), F4-15 (46.1~57.0min).

[0077] (5) The subfraction F4-13 obtained in step (4) was separated and purified in the fourth dimension using FC8HL (3.5 μm, 30×100 mm, reverse phase chromatography column), the mobile phase A was (20 mM) ammonium formate-95% methanol / water (volume concentration), B was (20 mM) ammonium formate-water, the gradient elution conditions were 0-2.38 min: 50% A, 2.38-13.88 min: 50%-100% A, 13.88-32 min: 100% A, and 10 subfractions were collected according to the corresponding distillate outflow time to obtain F4-13-1 (1.5-2. 5min), F4-13-2 (9.1~9.6min), F4-13-3 (9.6~11.0min), F4-13-4 (11.0~11.6min), F4-13-5 (11.6~12.4min), F4-13-6 (12 .4~13.6min), F4-13-7 (13.6~14.5min), F4-13-8 (14.5~16.2min), F4-13-9 (16.2~16.9min), F4-13-10 (16.9~19.2min).

[0078] (6) The subfraction F4-13-10 obtained in step (5) was separated and purified in the fifth dimension by C18HCE (5 μm, 10×150 mm, reverse phase chromatography column). The mobile phase A was formic acid-methanol (volume concentration 0.1%), and the mobile phase B was formic acid-water (volume concentration 0.1%). The gradient elution conditions were 0-5 min: 25% A, 5-15 min: 25-45% A, 15-20 min: 45% A, and the compound I-1 (t R =6.83min), named bungeanoline G.

[0079] (7) Compound I-1 obtained in step (6) was separated and purified by supercritical fluid chromatography on a BH column (5 μm, 10×250 mm, chiral column), with mobile phase A being CO2 and mobile phase B being triethylamine-methanol (volume concentration 0.1%), and gradient elution conditions being 0 to 12 min: 80% A, and compound I-1a (t R =5.0min) and I-1b(t R =6.9 min), and were named (-)-(R)-bungeanoline G and (+)-(S)-bungeanoline G, respectively.

[0080] The above compounds have the following physical and chemical properties and spectral characteristics:

[0081] Bungeanoline G: yellow powder; 1 H(600MHz) and 13 C (150 MHz) NMR data are shown in Table 1; HRESIMS m / z 211.1234 [M+H] + (calcd for C 14 H 15 N2,211.1235); 1a:-31(c 0.42,MeOH); 1b:37(c0.78,MeOH); ECD(MeOH) 1a:λ max (Δε)255(-2.74),349(0.04); 1b:λ max (Δε)255(4.52),347(-0.05).

[0082] Artificial synthesis method of compound I of the present invention

[0083] All compounds 1H-NMR spectra were collected on a Brucker AVANCE III 400MHz instrument, with tetraethylsilane (TMS) as the internal standard. Liquid chromatography-mass spectrometry (LC-MS) analysis was performed on a Waters Alliance e2695-ZQ2000 system, reporting m / z values. Unless otherwise noted, all solvents were used directly, all ratios of mixed solvents refer to volume ratios (v / v), and all temperatures are in degrees Celsius (°C).

[0084] The following non-limiting examples and pharmacological experiments will further illustrate the present invention. Unless otherwise specified, the compounds are racemates or mixtures of diastereomers.

[0085] Example 1

[0086]

[0087] 1.1 Synthesis of 4-chloro-3-(5-chloropent-1-yn-1-yl)quinoline

[0088] According to the steps and conditions described in reaction a of the above-mentioned synthesis method of the compound of the general formula 1, 4-chloro-3-bromo-quinoline and 5-chloropent-1-yne are used as substrates to obtain the target product with a yield of 85%. 1 H-NMR (400MHz, CDCl3) δ8.82(s,1H),8.22(dd,J=8.4,0.9Hz,1H),8.09(dd,J=8.4,0.5Hz,1H),7.74(ddd,J=8.4,6.9,1.4 Hz,1H),7.65(ddd,J=8.2,7.0,1.2Hz,1H),3.80(t,J=6.3Hz,2H),2.77(t,J=6.8Hz,2H),2.15(p,J=6.6Hz,2H).LCMS[M+H] + m / z 264.06.

[0089] 1.2 Synthesis of 5-chloro-1-(4-chloroquinolin-3-yl)pentan-2-one

[0090] According to the steps and conditions described in reaction b of the above-mentioned synthesis method of the compound of the general formula 1, 4-chloro-3-(5-chloropent-1-yn-1-yl)quinoline is used as a substrate to obtain the target product with a yield of 57%. 1H-NMR (400MHz, CDCl3) δ8.73(s,1H),8.26(d,J=8.1Hz,1H),8.15(d,J=8.1Hz,1H),7.78(t,J=7.0Hz,1H ),7.67(t,J=7.2Hz,1H),4.11(s,2H),3.60(s,2H),2.82(t,J=6.2Hz,2H),2.21–2.04(m,2H).LCMS[M+H] + m / z 282.00.

[0091] 1.3 Synthesis of 7a,8,9,10-tetrahydro-7H-pyrrolizino[2,3-c]quinoline (Bungeanoline G, I-1)

[0092] According to the steps and conditions described in reaction c of the above-mentioned synthesis method of the compound of general formula 1, 5-chloro-1-(4-chloroquinoline-3-yl)pentan-2-one is used as a substrate to obtain the target product I-1 with a yield of 27%. 1 H-NMR (400MHz, CDCl3) δ8.48 (s, 1H), 8.00 (t, J = 11.4Hz, 1H), 7.93 (dd, J = 8.4, 0.9H z,1H),7.59(ddd,J=8.4,6.8,1.4Hz,1H),7.38(ddd,J=8.2,6.8,1.2Hz,1H),4.29–4 .18(m,1H),3.91(ddd,J=10.1,5.7,3.3Hz,1H),3.55–3.43(m,1H),3.30(dd,J=15. 9,10.3Hz,1H),3.15(dd,J=15.9,4.3Hz,1H),2.12–1.93(m,3H),1.47–1.37(m,1H). 13 C-NMR (101MHz, CDCl3) δ156.60,148.06,145.44,128.44,127.69,123.28,122.06,119.16,117.24,65.69,51.05,30.67,25.85.HRMS(ESI):calcdfor C 14 H 14 N2[M+H] + m / z 211.1235, found 211.1186.

[0093] Example 2

[0094]

[0095] 2.1 Synthesis of 4-chloro-3-(5-chloropent-1-yn-1-yl)-6-methoxyquinoline

[0096] According to the steps and conditions described in reaction a of the above-mentioned synthesis method of the compound of the general formula 1, 3-bromo-4-chloro-6-methoxyquinoline and 5-chloropent-1-yne are used as substrates to obtain the target product with a yield of 69%. 1 H-NMR (400MHz, CDCl3) δ8.68(s,1H),7.97(d,J=9.1Hz,1H),7.43(d,J=2.7Hz,1H),7.38(d,J=2.8Hz,1H), 7.36(d,J=2.8Hz,1H),3.98(s,3H),3.80(t,J=6.3Hz,2H),2.77(t,J=6.8Hz,2H),2.20–2.10(m,2H).[M+H] + m / z 264.06.

[0097] 2.2 Synthesis of 5-chloro-1-(4-chloro-6-methoxyquinolin-3-yl)pentan-2-one

[0098] According to the steps and conditions described in reaction b of the above-mentioned synthesis method of the compound of the general formula 1, 4-chloro-3-(5-chloropent-1-yn-1-yl)-6-methoxyquinoline is used as a substrate to obtain the target product with a yield of 35%. 1 H-NMR (400MHz, CDCl3) δ8.58(s,1H),8.04(d,J=9.1Hz,1H),7.45(d,J=2.7Hz,1H),7.41(dd,J=9.1,2.8Hz, 1H),4.08(s,2H),3.98(s,3H),3.60(t,J=6.2Hz,2H),2.80(t,J=6.9Hz,2H),2.16–2.06(m,2H).LCMS[M+H] + m / z 312.09.

[0099] 2.3 Synthesis of 4-chloro-6-methoxy-3-((1-(4-methoxybenzyl)pyrrolidin-2-yl)methyl)quinoline

[0100] According to the steps and conditions described in reaction d of the above-mentioned synthesis method of the compound of the general formula 1, 5-chloro-1-(4-chloroquinoline-3-yl)pentan-2-one is used as a substrate to obtain the target product with a yield of 32%. 1H-NMR (400MHz, CDCl3) δ8.63 (s, 1H), 7.98 (d, J = 9.1Hz, 1H), 7.46 (d, J = 2.7Hz ,1H),7.36(dd,J=9.2,2.8Hz,1H),7.29(d,J=8.4Hz,2H),6.88–6.83(m,2H), 4.09(d,J=12.8Hz,1H),3.98(s,3H),3.82–3.77(m,3H),3.38(dd,J=14.5,6. 0Hz,2H),3.07–2.83(m,3H),2.33–2.18(m,1H),1.77–1.65(m,4H).LCMS[M+H] + m / z 397.25.

[0101] 2.4 Synthesis of 2-methoxy-7a,8,9,10-tetrahydro-7H-pyrrolizino[2,3-c]quinoline (I-2)

[0102] According to the steps and conditions described in reaction e of the above-mentioned synthesis method of the compound of general formula 1, 5-chloro-1-(4-chloroquinoline-3-yl)pentan-2-one is used as a substrate to obtain the target product I-2 with a yield of 27%. 1 H-NMR (400MHz, CDCl3) δ8.31(s,1H),7.87(d,J=9.2Hz,1H),7.20(dd,J=8.8,2.4Hz,1H),7.11(d,J=2.5Hz,1H),4.25–4.10(m,1H),3.87–3. 75(m,4H),3.41(dd,J=17.4,8.9Hz,1H),3.22(dd,J=15.9,10.3Hz,1H),3.05(dd,J=15.9,4.3Hz,1H),2.07–1.87(m,3H),1.44–1.34(m,1H). 13 C-NMR (101MHz, CDCl3) δ155.94,155.22,143.48,142.90,129.51,120.17,119.61,117.78,100.22,65.57,54.43,50.82,30.87,25.82.HRMS(ESI):calcd for C 15 H 16 N2O[M+H] + m / z 241.1341, found 241.1286.

[0103] Example 3

[0104]

[0105] 3.1 Synthesis of 4-chloro-3-(6-chlorohex-1-yn-1-yl)quinoline

[0106] According to the steps and conditions described in reaction a of the above-mentioned synthesis method of the compound of the general formula 1, 3-bromo-4-chloro-quinoline and 6-chlorohex-1-yne are used as substrates to obtain the target product with a yield of 78%. 1 H-NMR (400MHz, CDCl3) δ8.83(s,1H),8.24(dd,J=8.4,0.9Hz,1H),8.11(d,J=8.4Hz,1H),7.75(ddd,J=8.4,7.0,1.4Hz,1H),7.66(ddd, J=8.2,7.0,1.2Hz,1H),3.65(t,J=6.5Hz,2H),2.62(t,J=6.9Hz,2H),2.05(tt,J=13.1,6.7Hz,2H),1.87(dt,J=9.6,7.0Hz,2H).[M+H] + m / z 278.06.

[0107] 3. Synthesis of 2,6-chloro-1-(4-chloro-6-methoxyquinolin-3-yl)hexan-2-one

[0108] According to the steps and conditions described in reaction b of the above-mentioned synthesis method of the compound of the general formula 1, 4-chloro-3-(5-chloropent-1-yn-1-yl)-6-methoxyquinoline is used as a substrate to obtain the target product with a yield of 75%. 1 H-NMR (400MHz, CDCl3) δ8.70(s,1H),8.24(dd,J=8.4,0.7Hz,1H),8.13(d,J=8.4Hz,1H),7.76(ddd,J=8.4,7.0, 1.3Hz,1H),7.72–7.61(m,1H),4.07(s,2H),3.62–3.48(m,2H),2.73–2.56(m,2H),1.87–1.73(m,4H).LCMS[M+H] + m / z 296.06.

[0109] 3.3 Synthesis of 4-chloro-3-((1-(4-methoxybenzyl)piperidin-2-yl)methyl)quinoline

[0110] According to the steps and conditions described in reaction d of the above-mentioned synthesis method of the compound of the general formula 1, 6-chloro-1-(4-chloro-6-methoxyquinolin-3-yl)hexan-2-one was used as a substrate to obtain the target product with a yield of 53%. 1H-NMR (400MHz, CDCl3) δ8.70(s,1H),8.22(dd,J=8.4,0.9Hz,1H),8.08(dd,J=8.4,0.6Hz,1H),7.71(ddd,J=8.4, 6.9,1.4Hz,1H),7.62(ddd,J=8.2,6.9,1.2Hz,1H),7.27–7.25(m,2H),6.88–6.76(m,2H),4.03(d,J=13.4Hz,1H), 3.79(s,3H),3.57(d,J=13.4Hz,1H),3.47(dd,J=13.0,4.0Hz,1H),3.09–2.99(m,1H),2.95(d,J=2.0Hz,1H),2.8 5(dt,J=11.3,4.4Hz,1H),2.39–2.28(m,1H),1.76–1.66(m,1H),1.61–1.50(m,3H),1.44–1.34(m,2H).LCMS[M+H] + m / z 381.30.

[0111] 3.4 Synthesis of 7,7a,8,9,10,11-hexahydroindolizino[2,3-c]quinoline (I-3)

[0112] According to the steps and conditions described in reaction e of the above-mentioned synthesis method of the compound of general formula 1, using 4-chloro-3-((1-(4-methoxybenzyl)piperidin-2-yl)methyl)quinoline as a substrate, the target product I-3 can be obtained with a yield of 52%. 1 H-NMR (400MHz, CDCl3) δ8.27(s,1H),8.11(dd,J=19.3,9.3Hz,2H),7.61(t,J=6.9Hz,1H),7.34(dd,J=16.8,9.6Hz,1H),4.69(d,J=12.9Hz,1H),3.87(d t,J=11.8,5.8Hz,1H),3.41(td,J=12.6,2.8Hz,1H),3.27(dd,J=15.4,9.4H z,1H),2.75(dd,J=15.3,9.2Hz,1H),2.08–1.88(m,3H),1.77–1.56(m,3H). 13 C-NMR (101MHz, CDCl3) δ154.89,145.61,140.23,130.43,126.27,124.41,122.8 4,118.04,114.82,65.53,47.75,32.35,32.23,25.65,23.46.HRMS(ESI):calcd for C15 H 16 N2[M+H] + m / z 225.1392, found 225.1342.

[0113] Example 4

[0114]

[0115] 4.1 Synthesis of 4-chloro-3-(6-chlorohex-1-yn-1-yl)-6-methoxyquinoline

[0116] According to the steps and conditions described in reaction a of the above-mentioned synthesis method of the compound of the general formula 1, 3-bromo-4-chloro-6-methoxyquinoline and 6-chlorohex-1-yne are used as substrates to obtain the target product with a yield of 75%. 1 H-NMR (400MHz, CDCl3) δ8.68(s,1H),7.98(d,J=9.1Hz,1H),7.43(d,J=2.7Hz,1H),7.37(dd,J=9.1,2.8Hz,1H),3.9 8(s,3H),3.65(t,J=6.5Hz,2H),2.62(t,J=6.9Hz,2H),2.05(dt,J=14.6,6.7Hz,2H),1.91–1.80(m,2H).LCMS[M+H] + m / z 308.10.

[0117] 4. Synthesis of 2-6-chloro-1-(4-chloro-6-methoxyquinolin-3-yl)hexan-2-one

[0118] According to the steps and conditions described in reaction b of the above-mentioned synthesis method of the compound of the general formula 1, 4-chloro-3-(6-chlorohex-1-yn-1-yl)-6-methoxyquinoline is used as a substrate to obtain the target product with a yield of 59%. 1 H-NMR (400MHz, CDCl3) δ8.56 (s, 1H), 8.03 (d, J = 9.1Hz, 1H), 7.45 (d, J = 2.7Hz, 1H), 7.41 (dd, J = 9.1, 2.8 Hz,1H),4.05(s,2H),3.98(s,3H),3.58–3.49(m,2H),2.66–2.56(m,2H),1.84–1.75(m,4H).LCMS[M+H] + m / z 326.11.

[0119] 4.3 Synthesis of 4-chloro-6-methoxy-3-((1-(4-methoxybenzyl)piperidin-2-yl)methyl)quinoline

[0120] According to the steps and conditions described in reaction d of the above-mentioned synthesis method of the compound of the general formula 1, 6-chloro-1-(4-chloro-6-methoxyquinolin-3-yl)hexan-2-one was used as a substrate to obtain the target product with a yield of 32%. 1 H-NMR (400MHz, CDCl3) δ8.54(s,1H),7.96(d,J=9.1Hz,1H),7.44(d,J=2.7Hz,1H),7.35(dd,J=9.1,2.8 Hz,1H),7.27(d,J=4.2Hz,2H),6.85–6.80(m,2H),4.02(d,J=13.4Hz,1H),3.98(d,J=4.9Hz,3H),3.79( s,3H),3.56(d,J=13.4Hz,1H),3.44(dd,J=12.7,3.7Hz,1H),3.06–2.89(m,2H),2.87–2.77(m,1H),2.3 9–2.27(m,1H),1.70(dq,J=9.7,6.1Hz,1H),1.55(dd,J=10.9,5.3Hz,3H),1.43–1.32(m,2H).LCMS[M+H] + m / z 411.24.

[0121] 4.4 Synthesis of 2-methoxy-7,7a,8,9,10,11-hexahydroindolizino[2,3-c]quinoline (I-4)

[0122] According to the steps and conditions described in reaction e of the above-mentioned synthesis method of the compound of the general formula 1, using 4-chloro-6-methoxy-3-((1-(4-methoxybenzyl)piperidin-2-yl)methyl)quinoline as a substrate, the target product I-4 can be obtained with a yield of 56%. 1 H-NMR (400MHz, CDCl3) δ8.30(s,1H),7.94(s,1H),7.40(s,1H),7.36(d,J=7 .6Hz,1H),4.72(d,J=13.1Hz,1H),4.17–4.03(m,1H),3.92(s,3H),3.66–3.5 5(m,1H),3.35(dd,J=15.7,9.9Hz,1H),2.79(dd,J=15.7,8.5Hz,1H),2.09(d ,J=4.6Hz,1H),1.99(d,J=12.4Hz,2H),1.70(ddd,J=28.7,16.6,9.3Hz,3H). 13C-NMR (101MHz, CDCl3) δ156.74,155.98,136.61,132.93,123.88,123.67,117.56,1 14.10,103.03,65.46,55.84,47.53,32.85,31.50,25.72,23.09.HRMS(ESI):calcd for C 16 H 18 N2O[M+H] + m / z 255.1497, found 225.1440.

[0123] Example 5

[0124]

[0125] According to the steps and conditions described in reaction f of the above-mentioned general formula compound synthesis method 2, using I-1 and an appropriate amount of NBS as substrates, the target products I-5, I-6, I-7, and I-8 can be obtained with yields of 68%, 23%, 12%, and 10%, respectively.

[0126] 5.1 2-Bromo-7a,8,9,10-tetrahydro-7H-pyrrolizino[2,3-c]quinoline (I-5):

[0127] 1 H-NMR (400MHz, CDCl3) δ8.46(s,1H),8.05(d,J=2.0Hz,1H),7.88(d,J=9.1Hz,1H),7.64(dd,J=9.1,2.1Hz,1H),4.26(dt,J=10.4,5.3Hz,1H),3.9 5–3.85(m,1H),3.47(dd,J=16.9,9.4Hz,1H),3.30(dd,J=16.1,10.4Hz, 1H),3.16–3.10(m,1H),2.13–1.99(m,3H),1.43(dt,J=10.9,3.5Hz,1H). 13 C-NMR (101MHz, CDCl3) δ156.88,147.27,146.23,132.29,130.88,125.41,121.21,119.18,117.84,66.90,51.79,31.70,31.45,26.89.HRMS(ESI):calcd for C 14 H 13 BrN2[M+H] + m / z 289.0340, found 289.0275.

[0128] 5.2 3-Bromo-7a,8,9,10-tetrahydro-7H-pyrrolizino[2,3-c]quinoline (I-6):

[0129] 1 H-NMR (400MHz, CDCl3) δ8.42(s,1H),8.03(d,J=2.1Hz,1H),7.94(d,J=9.1Hz,1H),7.66(dd,J=9.1,2.1Hz,1H),4.36–4.23(m,1H),3.96–3. 85(m,1H),3.58–3.45(m,1H),3.29(dd,J=16.2,10.5Hz,1H),3.12(dd,J=16.2,4.9Hz,1H),2.18–2.09(m,3H),1.44(dt,J=11.0,7.7Hz,1H). 13 C-NMR (101MHz, CDCl3) δ157.60,145.07,143.58,133.40,128.68,125.54,121.46,118.25,118.22,67.41,51.01,31.51,30.91,26.95.HRMS(ESI):calcd forC 14 H 13 BrN2[M+H] + m / z 289.0340,found 289.0260.

[0130] 5.3 4-Bromo-7a,8,9,10-tetrahydro-7H-pyrrolizino[2,3-c]quinoline (I-7):

[0131] 1 H-NMR (400MHz, CDCl3) δ8.50(s,1H),7.82(ddd,J=9.6,7.9,1.2Hz,2H),7.11(dd,J=8.3,7.5Hz,1H),4.21–4.10(m,1H),3.83–3.74(m ,1H),3.34(td,J=9.5,7.9Hz,1H),3.21(dd,J=16.1,10.4Hz,1H),3.06(dd,J=16.1,4.4Hz,1H),2.00–1.88(m,3H),1.37–1.26(m,1H). 13C-NMR (101MHz, CDCl3) δ157.98,147.21,146.31,132.43,124.92,124.28,123.16,121.46,119.31,66.98,52.17,31.72,31.36,26.89.HRMS(ESI):calcd forC 14 H 13 BrN2[M+H] + m / z 289.0340, found 289.0257.

[0132] 5.4 2,4-Dibromo-7a,8,9,10-tetrahydro-7H-pyrrolizino[2,3-c]quinoline (I-8):

[0133] 1 H-NMR (400MHz, CDCl3) δ8.46(s,1H),7.92(d,J=2.1Hz,1H),7.90(d,J=2.1Hz,1H),4.16(dq,J=15.6,5.0Hz,1H),3.83–3.70(m,1H),3.2 9(tt,J=12.5,6.2Hz,1H),3.20(dd,J=16.3,10.4Hz,1H),3.05(dd,J=16.3,4.5Hz,1H),2.05–1.90(m,3H),1.31(qd,J=11.1,7.3Hz,1H). 13 C-NMR (101MHz, CDCl3) δ157.04,147.33,145.09,134.95,126.04,125.34,122.39,119.75,116.52,67.07,52.00,31.70,31.29,26.89.HRMS(ESI):calcd forC 14 H 12 Br2N2[M+H] + m / z 368.9425, found 368.9347.

[0134] Example 6

[0135]

[0136] According to the steps and conditions described in reaction g of the above-mentioned synthesis method of the compound of general formula 2, the target product I-9 can be obtained with a yield of 39% using I-5 and methylboronic acid as substrates. 1H-NMR (400MHz, CDCl3) δ8.43(s,1H),7.89(d,J=8.7Hz,1H),7.68(s,1H),7.41(d,J=8.6Hz,1H),4.30–4.15(m,1H),3.94–3.84(m,1H),3.48(dd,J=1 7.7,8.9Hz,1H),3.28(dd,J=15.9,10.3Hz,1H),3.13(dd,J=15.8,4.2Hz, 1H), 2.50 (s, 3H), 2.05 (ddt, J=20.1, 11.4, 4.0Hz, 3H), 1.49–1.38 (m, 1H). 13 C-NMR (101MHz, Acetone) δ157.04,147.87,146.09,133.97,130.81,129.37,121.84,120.22,118.42,66.55,52.36,31.85,26.83,21.83.HRMS(ESI):calcd forC 15 H 16 N2[M+H] + m / z 225.1392 found 225.1339.

[0137] Example 7

[0138] According to the steps and conditions described in reaction h of the above-mentioned general formula compound synthesis method 3, I-9 and formic acid are used as substrates to obtain the target product I-9-HCOOH with a yield of 95%. 1 H-NMR (400MHz, CDCl3) δ8.36(s,1H),8.26(s,1H),7.93–7.80(m,2H),7.52(dd,J=8.7,1.9Hz,1H),4.39–4.26(m,1H),4.00(ddd,J=10.5,8.4,2. 5Hz,1H),3.55(d,J=7.5Hz,1H),3.37–3.24(m,1H),3.18(dd,J=16.1,4.9Hz,1H),2.50(s,3H),2.18–2.09(m,3H),1.50(dt,J=11.2,3.6Hz,1H). 13C-NMR(101MHz,Acetone)δ162.89,157.72,145.58,142.99,134.28,131.66,126.89, 122.32,120.73,117.36,67.06,51.12,31.30,30.73,26.63,20.72.HRMS(ESI):calcd for C 15 H 16 N2[M+H] + m / z 225.1392 found 225.1339.

[0139] Example 8

[0140]

[0141] According to the steps and conditions described in reaction g of the above-mentioned synthesis method of the compound of general formula 2, the target product I-9 can be obtained with a yield of 43% using I-5 and ethylboric acid as substrates. 1 H-NMR (400MHz, CDCl3) δ8.37 (s, 1H), 7.88 (d, J = 8.7Hz, 1H), 7.63 (s, 1H), 7.40 (dd,J=8.7,1.8Hz,1H),4.25–4.12(m,1H),3.91–3.80(m,1H),3.50–3.41(m,1H ),3.23(dd,J=15.9,10.3Hz,1H),3.07(dd,J=15.9,4.3Hz,1H),2.73(q,J=7.6H z,2H),2.03–1.91(m,3H),1.38(dt,J=10.5,6.7Hz,1H),1.25(t,J=7.6Hz,3H). 13 C-NMR (101MHz, Acetone) δ157.55,147.11,144.98,140.52,130.26,128.65,120. 59,120.23,118.06,66.77,52.00,31.70,29.06,26.87,15.49.HRMS(ESI):calcd for C 16 H 18 N2[M+H] + m / z 239.1548 found 239.1494.

[0142] Example 9

[0143]

[0144] According to the steps and conditions described in reaction h of the above-mentioned general formula compound synthesis method 3, I-10 and formic acid are used as substrates to obtain the target product I-10-HCOOH with a yield of 93%. 1 H-NMR (400MHz, CDCl3) δ8.36 (s, 1H), 8.32 (s, 1H), 7.95 (d, J = 8.8Hz, 1H), 7.87 (s, 1 H),7.60(dd,J=8.7,1.9Hz,1H),4.41–4.37(m,1H),4.04(ddd,J=10.6,8.3,2.3Hz, 1H),3.65(td,J=9.5,7.3Hz,1H),3.40–3.27(m,1H),3.19(dd,J=16.1,5.0Hz,1H), 2.83(q,J=7.6Hz,2H),2.19–2.08(m,3H),1.60–1.47(m,1H),1.31(t,J=7.6Hz,3H). 13 C-NMR(101MHz,Acetone)δ163.58,158.31,144.69,141.73,140.88,131.24,125.92,121 .28,120.89,116.95,67.36,50.72,31.19,30.45,28.53,26.69,15.01.HRMS(ESI):calcd for C 16 H 18 N2[M+H] + m / z 239.1548 found 239.1494.

[0145] Natural product I-1 1 H-NMR and 13 The C-NMR data are shown in Table 1. The key two-dimensional NMR information of the compounds is as follows: Figure 1 shown.

[0146] Table 1. Natural product I-1 1 H-NMR and 13 C-NMR data (δ H and δ C , MeOD)

[0147]

[0148]

[0149] Activity test example:

[0150] Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) were purchased from Sigma-Aldrich, and physostigmine, galantamine, acetylcholine iodide (ATCI) and butyrylcholine iodide (BTCI) were purchased from Aladdin Reagent Co., Ltd. 5,5-Dimercaptobis-2-nitrobenzoic acid (DTNB) was purchased from Shanghai Merrill Biochemical Technology Co., Ltd. Ethylenediaminetetraacetic acid disodium (EDTA) and sodium dodecyl sulfate (SDS) were purchased from Solebao Technology Co., Ltd. Bovine serum albumin (BSA) was purchased from J&K Technology Co., Ltd. Na2HPO4 and NaH2PO4 were purchased from Sinopharm Chemical Reagent Co., Ltd. The buffer was prepared as follows: phosphate buffer A (0.1 M, pH = 7.4, containing 1% BSA), phosphate buffer B (0.1 M, pH = 7.4, containing 1 mM EDTA), and phosphate buffer C (0.1 M, pH = 7.4).

[0151] The AChE and BChE inhibitory activity of compound I was detected by the modified Ellman method. The detection principle is as follows: AChE or BChE hydrolyzes the substrate iodinated acetylcholine (ATCI) or iodinated thiobutyrylcholine (BTCI) to generate a product, thiocholine, which reacts with the colorimetric reagent 5,5-dimercaptobis-2-nitrobenzoic acid (DTNB) to generate a yellow compound. The amount of the product thiocholine is indirectly determined by colorimetry, so that the amount of the hydrolyzed product thiocholine reflects the activity of AChE or BChE. Physostigmine (PSM) and galantamine (GLM) were used as positive controls. The specific detection operation is as follows: Step 1, add 12.5 μL of 0.50U / mL AChE solution or 0.10U / mL BChE solution (dissolved in phosphate buffer A) to a 96-well plate, and add 12.5 μL of phosphate buffer A to the experimental control group. In the second step, 15 μL of phosphate buffer B (blank group), physostigmine or galantamine (positive control), and compound I (experimental group) were added, and all samples were dissolved in phosphate buffer B; 15 μL of phosphate buffer B was added to the experimental control group. The highest final concentration of physostigmine and galantamine was 10 μM, 4-fold dilution, 7 concentration points (10 μM, 2.5 μM, 0.625 μM, 0.1563 μM, 0.0391 μM, 0.001 μM, 0.0025 μM); the highest final concentration of compound I was 100 μM, 4-fold gradient dilution, 7 concentration points (100 μM, 25 μM, 6.25 μM, 1.563 μM, 0.391 μM, 0.098 μM, 0.024 μM). The third step is to shake evenly, add 25 μl of 2 mM ATCI or BTCI solution (dissolved in phosphate buffer C), mix well and incubate in a 37°C constant temperature incubator for 20 minutes. The fourth step is to add 25 μl of 6 mM DTNB solution (dissolved in phosphate buffer C, containing 2% SDS by mass), and finally add 22.5 μl of phosphate buffer B. After shaking evenly, measure the absorbance value (A) at a wavelength of 412 nm and calculate the enzyme inhibition rate. Enzyme activity inhibition rate (%) = [(A 空白组 -A 阳性控制 )-(A 实验组 -A 实验控制组 )] / (A 空白组 -A 阳性控制 )×100.

[0152] Table 2. AChE and BChE inhibitory activity data of compound I

[0153]

[0154] Note: a Indicates that the compound was not tested due to solubility issues.

[0155] The AChE and BChE inhibitory activity data of compound I are shown in Table 2. The dose-dependent inhibition curves of compound I on AChE and BChE are shown in Table 2. Figure 2 and Figure 3 As shown, the test results show that 6 compounds have dual inhibitory activity against AChE and BChE, among which compounds I-1 and I-7 have the most significant activity (the inhibitory activity against AChE and BChE is less than 1 μM); and another 5 compounds have significant AChE inhibitory activity. Therefore, the compounds of the present invention can be used to develop lead compounds for treating or improving neurodegenerative diseases such as Alzheimer's disease, vascular dementia, and myasthenia gravis.

Claims

1. A tetrahydropyrrolizino[2,3-c]quinoline alkaloid compound, comprising one or two or more of the compound represented by the general structural formula I and its pharmaceutically acceptable salts, solvates, hydrates or crystalline compounds: in, R1-R4 are each independently H, halogen (one or more of bromine, chlorine, fluorine, iodine), C 1-3 Alkyl, hydroxyl, C 1-3 One or two or more alkoxy groups; n is 1, 2 or 3.

2. The compound according to claim 1, characterized in that R1-R4 of compound I are each independently one or two or more of H, F, Cl, Br, I, methyl, ethyl, propyl, isopropyl, hydroxyl, methoxy, ethoxy, propoxy, and isopropoxy; More preferably, R1-R4 are independently one or two or more of H, F, Br, methyl, ethyl, propyl, hydroxyl, methoxy, ethoxy, and propoxy; More preferably, R1-R4 are independently one or two or more of H, Br, methyl, ethyl, methoxy, and ethoxy; In compound I, n is independently 1, 2 or 3; preferably, n is independently 1 or 2.

3. The compound according to claim 1 or 2, characterized in that Compound I is a racemate (I-achiral), or one, two, or three of 7aR or 7aS, as shown in the following formulas I-achiral, I-7aS, and I-7aR; 4. The compound according to claim 1, characterized in that The preferred compounds include one or more of the following:

5. The compound according to any one of claims 1 to 4, characterized in that The compound can form a salt with a pharmaceutically acceptable acid, which refers to one or more of HCl, HBr, trifluoroacetic acid, sulfuric acid, formic acid, acetic acid, succinic acid, fumaric acid, p-toluenesulfonic acid, methanesulfonic acid, trifluoromethanesulfonic acid, and citric acid; The compound includes its corresponding enantiomeric mixture, one or more of its enantiomeric monomers, and one or more of their pharmaceutically acceptable salts, solvates, hydrates or various crystal forms.

6. Use of a compound according to any one of claims 1 to 5 as an active ingredient in the preparation of a drug for treating or improving a neurodegenerative disease, or as an active ingredient in the preparation of a cholinesterase inhibitor; The neurodegenerative disease includes one or more neurodegenerative diseases such as Alzheimer's disease, vascular dementia or myasthenia gravis.

7. A pharmaceutical composition comprising the compound of any one of claims 1 to 5 or one or more of its pharmaceutically acceptable salts, solvates, hydrates or crystalline compounds as active ingredients, and any pharmaceutically acceptable excipients and / or other active compounds.

8. Use of the pharmaceutical composition according to claim 7, characterized in that: The disease to which the pharmaceutical composition is applicable is one or more of two or more neurodegenerative diseases such as Alzheimer's disease, vascular dementia or myasthenia gravis.

9. A method for preparing the compound I-1 according to claim 4 from the Chinese medicinal material Kudiding, characterized in that: The following steps are involved: (1) Extraction of medicinal materials: Take the whole dried herb with roots of Corydalis eryngii, crush it to obtain medicinal material powder; use 50% to 90% ethanol by volume to heat and extract at 60 to 70°C for 6 to 24 hours, filter the extract with a 70-80 mesh filter, and then rotary evaporate the filtrate to remove the solvent to obtain an extract; dissolve the extract in 0.01 to 0.2 mol / L sulfuric acid water, add 2% sulfuric acid aqueous solution to adjust the pH to 2 to 3, add petroleum ether at a volume ratio of 1:1 to 1:2 (petroleum ether: sulfuric acid aqueous solution) to extract, and take the water layer; adjust the pH of the obtained water layer to 9 to 10 with 0.01 to 2 mol / L sodium hydroxide, add dichloromethane at a volume ratio of 1:1 to 1:2 (dichloromethane: water layer with pH 9 to 10) to extract, and the obtained dichloromethane layer is the crude alkali; (2) The crude alkali obtained in step (1) is separated and purified in the first dimension using C18HCE (reverse phase chromatography column), wherein the mobile phase A is formic acid-methanol (volume concentration 0.1%-2%), and the mobile phase B is formic acid-water (volume concentration 0.1%-2%), and the gradient elution conditions are: 0-8 min: 25% A (volume ratio, the same below), 8-23 min: 40% A, 23-38 min: 80% A, 38-55 min: 100% A, and the sample is taken from 4.2-26 min, and concentrated as a total alkaloid sample of Corydalis bispinosa L.; (3) The total alkaloids of bitter herbs obtained in step (2) were separated and purified in the second dimension using C18HCE (reverse phase chromatography column), the mobile phase A was formic acid-methanol (volume concentration 0.1%-2%), B was formic acid-water (volume concentration 0.1%-2%), the gradient elution conditions were: 0-5 min: 20% A (volume ratio, the same below), 5-50 min: 20-45% A, 50-60 min: 95% A, and 11 sub-fractions F1 (5.7-10 .7min), F2 (10.7~17.1min), F3 (17.1~20.9min), F4 (20.9~25.7min), F5 (25.7~29.2min), F6 (29.2~33.0m in), F7 (33.0~36.3min), F8 (36.3~40.4min), F9 (40.4~44.4min), F10 (44.4~55.6min) and F11 (55.6~60min); (4) The sub-fraction F4 obtained in step (3) was separated and purified in the third dimension using C8CE (reverse phase chromatography column), wherein the mobile phase A was (volume concentration 0.1% to 2%) ammonia-methanol with a mass concentration of 20 to 25%, and the mobile phase B was (volume concentration 0.1% to 2%) ammonia-water with a mass concentration of 20 to 25%, and the gradient elution conditions were 0 to 5 min: 5% A (volume ratio, the same below), 5 to 50 min: 5 to 95% A, and 50 to 60 min: 95% A. The 15 sub-fractions F4-1 (4.0 to 7.8 min), F4-2 (7.8 to 10.1 min), and F4-3 (10.1 to 13.4 min) were collected in sequence according to the corresponding distillate outflow time. , F4-4 (13.4~18.1min), F4-5 (18.1~21.3min), F4-6 (21.3~23.7min), F4-7 (23.7~28.5min), F4-8 (28.5~30.7min), F4-9 (30.7~33.6min), F4 -10(33.6~35.0min), F4-11(35.0~37.6min), F4-12(37.6~39.5min), F4-13(39.5~43.9min), F4-14(43.9~46.1min), F4-15(46.1~57.0min); (5) The subfraction F4-13 obtained in step (4) was separated and purified in the fourth dimension using FC8HL (reverse phase chromatography column), the mobile phase A was (5-50 mM) ammonium formate-95% methanol / water (volume concentration), B was (5-50 mM) ammonium formate-water, and the gradient elution conditions were 0-2.38 min: 50% A (volume ratio, the same below), 2.38-13.88 min: 50%-100% A, 13.88-32 min: 100% A, and 10 subfractions were collected in sequence according to the chromatographic peaks, namely F4-13-1 (1.5-2.5 min ), F4-13-2 (9.1~9.6min), F4-13-3 (9.6~11.0min), F4-13-4 (11.0~11.6min), F4-13-5 (11.6~12.4min), F4-13-6 (12.4 ~13.6min), F4-13-7 (13.6~14.5min), F4-13-8 (14.5~16.2min), F4-13-9 (16.2~16.9min), F4-13-10 (16.9~19.2min); (6) The subfraction F4-13-10 obtained in step (5) was separated and purified in the fifth dimension by C18HCE (reverse phase chromatography column), the mobile phase A was formic acid-methanol (volume concentration 0.1% to 2%), B was formic acid-water (volume concentration 0.1% to 2%), the gradient elution conditions were 0 to 5 min: 25% A (volume ratio, the same below), 5 to 15 min: 25 to 35% A, and the natural product I-1 (t R =6.58~7.07min), named bungeanoline G; (7) The natural product I-1 obtained in step (6) was separated and purified by supercritical fluid chromatography using a BH (chiral chromatography column), wherein the mobile phase A was CO2, and B was triethylamine-methanol (volume concentration 0.1% to 2%), and the gradient elution condition was 0 to 12 min: 80% A (volume ratio, the same below), and the natural product 1a (t R =4.0~6.5min) and 1b(t R =6.5~8.0min), named as (-)-(R)-bungeanoline G and (+)-(S)-bungeanoline G, respectively.

10. A method for synthesizing the compound I or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 5, characterized in that: The synthesis method is one or more of the following: Synthesis method 1 In the reaction formula, R1-R4 and n are respectively the same as defined in any one of claims 1-4 above; Reaction a The substrate 1 (1 mmol, 1.0 molar equivalent) is dissolved in THF (10-100 ml), and then substrate 2 (relative to substrate 1, 1-3 molar equivalent), Pd(PPh3)2Cl2 (relative to substrate 1, 0.01-0.1 molar equivalent), CuI (relative to substrate 1, 0.01-0.1 molar equivalent), PPh3 (relative to substrate 1, 0.01-0.1 molar equivalent) and triethylamine (relative to substrate 1, 1-5 molar equivalent) are added, and the mixture is stirred and dissolved at room temperature; after the reaction system is replaced with a nitrogen atmosphere, it is heated to 30-60° C. and stirred for 5-24 hours; after the reaction is completed as monitored by LC-MS or a thin layer chromatography plate (TLC), the reaction solution is cooled to room temperature, filtered, and the filtrate is concentrated by rotary evaporation, and the concentrate is purified by silica gel column chromatography to obtain product 3 (as substrate 3); Reaction b The substrate 3 (1 mmol, 1.0 molar equivalent) is dissolved in acetone (10-50 ml), and water (1-10 ml), mercuric sulfate (relative to substrate 3, 1-3 molar equivalent) and concentrated sulfuric acid (mass concentration 98%) (relative to substrate 3, 1-3 molar equivalent) are added in sequence under stirring, and stirred at 25-40° C. for 24-72 hours; after the reaction is completed as monitored by LC-MS or TLC, a saturated sodium carbonate solution is added to the reaction solution to neutralize the sulfuric acid to a pH of 8-9, and ethyl acetate (10-50 ml of ethyl acetate is used each time) is extracted 2-5 times, and the organic phase is dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the corresponding product 4 (as substrate 4); Reaction c Substrate 4 (1 mmol, 1.0 molar equivalent) was dissolved in methanol (2-20 ml), acetic acid (1-5 molar equivalent relative to substrate 4) and benzylamine (1-5 molar equivalent relative to substrate 4) were added, and the mixture was stirred at 25°C for 1-12 hours. Sodium cyanoborohydride (1-5 molar equivalent relative to substrate 4) was added, and the mixture was stirred at 25°C for 2-24 hours. Water (2-20 ml) was added to the reaction mixture, and the mixture was stirred for 0.5-1 hour. Ethyl acetate (2-20 ml ethyl acetate was used each time) was added. ) for 2-5 times, the organic phase was discarded, concentrated hydrochloric acid (mass concentration 38%) was added dropwise to the aqueous phase to adjust the pH to 2-3, palladium carbon (specification 10wt%, relative to substrate 4, 0.1-2 mass equivalent) was added, and stirred at 25°C in hydrogen for 3-24 hours; saturated sodium carbonate solution was added to the reaction solution until the pH was 8-9, and ethyl acetate (2-20 ml of ethyl acetate was used each time) was extracted 2-5 times, the organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product I; Reaction Substrate 4 (1 mmol, 1.0 molar equivalent) was dissolved in methanol (2-20 ml), acetic acid (relative to substrate 4, 1-5 molar equivalent) and p-methoxybenzylamine (relative to substrate 4, 1-5 molar equivalent) were added, and the mixture was stirred at 25°C for 1-5 hours. Sodium cyanoborohydride (relative to substrate 4, 1-5 molar equivalent) was added, and the mixture was stirred at 25°C for 2-24 hours. Methanol was removed from the reaction solution under reduced pressure, saturated sodium carbonate solution was added to pH 8-9, and ethyl acetate (2-20 ml each time) was used for extraction 2-5 times. The organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the corresponding product 5 (as substrate 5). Reaction e The substrate 5 (1 mmol, 1.0 molar equivalent) was dissolved in acetonitrile (2-20 ml) and water (8-80 ml), and 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ, 1-3 molar equivalent relative to the substrate 5) was added, and the mixture was heated to 70°C and stirred for 1-12 hours. The acetonitrile was removed from the reaction solution under reduced pressure, and 2N hydrochloric acid (2-5 ml) and an equal volume of ethyl acetate were added. The mixture was separated, and the organic phase was extracted 2-5 times with 2N hydrochloric acid (1-5 ml each time). The organic phase was discarded, and the acid-water phases were combined and the pH was adjusted to 8-9 with a saturated sodium carbonate solution. The mixture was extracted 2-5 times with ethyl acetate (2-20 ml each time). The organic phase was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product I. Alternatively, when all R1-R4 in compound I are not Br and at least one is H, compound I can be further modified by synthetic method 2 as shown below: Synthesis method 2 Br in compound Ia can be substituted at any or any two positions of R2-R4, R refers to other R1-R4 substituents other than those substituted by Br under reaction condition f, R' in Ib refers to the group after replacing the Br group under reaction condition g; "alkyl boronic acid or boric ester" refers to C 1-3 Alkylboronic acid or boric acid ester; Compounds Ia and Ib are within the scope of Compound I; Reaction f The substrate I (1 mmol, 1.0 molar equivalent) is dissolved in acetic acid (1-10 ml), N-bromosuccinimide (NBS, 1-5 molar equivalent relative to the substrate I) is added under ice bath, and stirred at room temperature for 0.5-1 hour. A saturated sodium sulfite solution is added to the reaction solution to quench the reaction; the reaction solution is concentrated under reduced pressure, a saturated sodium carbonate solution is added to a pH range of 8-9, and extracted with dichloromethane (2-20 ml each time) 2-5 times, and the organic phase is dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product Ia (as compound Ia); Reaction g Compound Ia (1 mmol, 1.0 molar equivalent) was dissolved in dioxane (1-10 ml), and water (1-5 ml), C 1-3 Alkylboronic acid or borate substrate (relative to substrate Ia, 1-5 molar equivalents), Pd(PPh3)4 (relative to substrate Ia, 0.01-0.1 molar equivalents) and potassium carbonate (relative to substrate Ia, 1-5 molar equivalents) are prepared under nitrogen protection and stirred at 50-100°C for 8-24 hours; the reaction solution is cooled to room temperature and filtered, the filter cake is washed with ethyl acetate, the filtrate is washed 2-5 times with saturated brine (2-20 ml each time), dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by silica column chromatography to obtain the target product Ib.

11. The synthesis method according to claim 10, characterized in that: When compound I is a chiral monomer, the racemate can be separated by chiral chromatography: Split method The racemic compound I is dissolved in methanol, and separated and purified by supercritical fluid chromatography through BH (chiral chromatography column), the mobile phase A is CO2, B is (volume concentration 0.1% to 2%) triethylamine-methanol, the gradient elution condition is 0 to 12min: 80% A, and the chiral monomers IS and IR are obtained by collecting according to the chromatographic peaks; And / or, when compound I forms a salt with a pharmaceutically acceptable acid (HX) (these acids refer to one or more of HCl, HBr, trifluoroacetic acid, sulfuric acid, formic acid, acetic acid, succinic acid, fumaric acid, p-toluenesulfonic acid, methanesulfonic acid, trifluoromethanesulfonic acid, and citric acid), the synthesis method is as follows: Synthesis method 3 Reaction h Compound I (1 mmol, 1.0 molar equivalent) is dissolved in dioxane or tetrahydrofuran or dichloromethane or ethyl acetate (1-10 ml), HX (relative to substrate I, 1-3 molar equivalent) is added, stirred at room temperature for 0.5-2 hours, concentrated under reduced pressure, and the precipitated solid is the target product IX.

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