A β-carboline alkaloid derivative, its extraction method and application; a monoamine oxidase inhibitor

Abstract

This invention provides β -Carboline alkaloid derivatives, their extraction methods and applications, and a monoamine oxidase inhibitor. This invention provides... β The carbaline alkaloid derivatives, with the structure shown in Formula 1, possess a novel structure and exhibit excellent monoamine oxidase inhibitory activity. This can slow the depletion of monoamine neurotransmitters in the brain, mitigate oxidative damage, and contribute to the treatment of neurodegenerative diseases. Furthermore, they can be used to prepare monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases, showing broad application prospects. (Formula 1)

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine technology, specifically relating to a... β - Carboline alkaloid derivatives, their extraction methods and applications, and a monoamine oxidase inhibitor. Background Technology

[0002] Monoamine oxidase (MAO) is a class of flavoproteins found on the outer mitochondrial membrane, and it exists in two subtypes: monoamine oxidase A (MAO-A) and monoamine oxidase B (MAO-B). MAO-A preferentially reacts with substrates such as serotonin and adrenaline, while MAO-B preferentially reacts with substrates such as benzylamine. β - Phenylethylamine. The control of motor, mood regulation, perception, and cognitive functions, as well as the occurrence of related diseases, are closely related to monoaminergic signaling mechanisms. In vivo, MAO-A is mainly associated with depression, while MAO-B is associated with Parkinson's disease. Excessive activity of monoamine oxidase in the brain leads to excessive metabolism of monoamine neurotransmitters and the generation of excessive hydrogen peroxide, inducing nervous system disorders and oxidative damage. Oxidative damage is one of the causes of many neurodegenerative diseases, such as depression, Parkinson's disease, and Alzheimer's disease. In particular, excessively high levels of monoamine oxidase have been detected in brain slices from deceased patients with depression and Parkinson's disease. Therefore, inhibiting the activity of monoamine oxidase can slow down the consumption of monoamine neurotransmitters in the brain, reduce oxidative damage, and achieve the goal of treating neurodegenerative diseases such as depression and Parkinson's disease.

[0003] Tangut white thorn ( Nitraria tangutorum Bobr. is a wild plant used for both food and medicine; its fruit is an important plant resource, rich in polysaccharides, β The fruit of *Nitraria tangutorum* contains carboline alkaloids, flavonoids, and phenolic acids, possessing high economic, ecological, and medicinal value. In traditional Chinese medicine, it is commonly used to treat spleen and stomach weakness, indigestion, neurasthenia, hyperglycemia, dizziness, colds, and insufficient lactation. Currently, research on extracts from *Nitraria tangutorum* fruit is limited, and there are no reports of extracting and isolating substances with monoamine oxidase inhibitory activity from it. Summary of the Invention

[0004] The purpose of this invention is to provide a β -Carboline alkaloid derivatives, their extraction methods and applications, and a monoamine oxidase inhibitor. This invention provides... β -Carboline alkaloid derivatives have good monoamine oxidase inhibitory activity and can be used to prepare monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases.

[0005] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0006] This invention provides β -Carboline alkaloid derivatives, having the structure shown in Formula 1:

[0007] Formula 1.

[0008] The present invention provides the technical solution described above. β - The extraction method for carboline alkaloid derivatives includes the following steps:

[0009] (1) The juice of Tangut white thorn was subjected to alcohol precipitation to obtain an extract;

[0010] (2) Dissolve the extract in water, and use pure water and 40% ethanol aqueous solution as eluents in sequence. Perform macroporous resin column chromatography on the extract to obtain the elution fraction of 40% ethanol aqueous solution.

[0011] (3) Using MCI resin as the stationary phase and methanol-water as the eluent, the eluted fraction of the 40% (v / v) ethanol aqueous solution was separated by column chromatography under gradient elution conditions to obtain four components, denoted as Fr. 40-1 ~Fr 40-4 The volume fraction of methanol in the eluent of step (3) is 0%~80%.

[0012] (4) Using MCI resin as the stationary phase and methanol-water as the eluent, Fr was removed under gradient elution conditions. 40-1 The components were separated by column chromatography, yielding 8 components, denoted as Fr. 40-1-1 ~Fr 40-1-8 The volume fraction of methanol in the eluent of step (4) is 0%~50%.

[0013] (5) Using LH-20 gel as the stationary phase and methanol-water as the eluent, Fr was removed under gradient elution conditions. 40-1-7 The components were separated by column chromatography, yielding 10 components, denoted as Fr. 40-1-7-1 ~Fr 40-1-7-10 The volume fractions of methanol in the eluent of step (5) are 0%, 20%, 40%, and 60%, respectively.

[0014] (6) Using C18 bonded silica gel as the stationary phase and acetonitrile-water as the eluent, Fr was removed under isogradient elution conditions. 40-1-7-9 The components were separated by column chromatography to obtain the structure shown in Formula 1. β -Carboline alkaloid derivatives; the eluent contains 15% acetonitrile by volume.

[0015] Preferably, in step (1), the reagent used for alcohol precipitation is an aqueous ethanol solution; the volume fraction of the aqueous ethanol solution is 50%~75%.

[0016] Preferably, the alcohol precipitation is performed 3 to 4 times.

[0017] Preferably, the alcohol precipitation time for each precipitation is 10-12 hours, and the volume ratio of the ethanol aqueous solution to the Tangut thorn juice is 2-4:1.

[0018] Preferably, in step (3), the volume fraction of methanol in the eluent used for column chromatography separation is 0%, 20%, 40% and 80% respectively.

[0019] Preferably, in step (4), the volume fraction of methanol in the eluent used for column chromatography separation is 0%, 10%, 20%, 30%, 40% and 50% respectively.

[0020] Preferably, in step (2), the chromatographic column used for macroporous resin column chromatography is an AB-8 macroporous resin column.

[0021] The present invention provides the technical solution described above. β - Application of carboline alkaloid derivatives in the preparation of monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases.

[0022] This invention provides a monoamine oxidase inhibitor, comprising the components described above. β -Carboline alkaloid derivatives.

[0023] This invention provides a structure having the structure shown in Formula 1 β - Carboline alkaloid derivatives, with novel structures and excellent monoamine oxidase inhibitory activity, can slow down the consumption of monoamine neurotransmitters in the brain, mitigate oxidative damage, and achieve therapeutic effects on neurodegenerative diseases. Furthermore, they can be used to prepare monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases, showing broad application prospects. Examples show that... β -Carboline alkaloid derivatives have good MAO-A inhibitory activity, IC50 50 The concentration was 151.4 µmol / L; it also exhibited good MAO-B inhibitory activity, IC50... 50 The value is 122.3 µmol / L.

[0024] The present invention also provides β This invention relates to a method for extracting carboline alkaloid derivatives. Using *Nitraria tangutorum* fruit as raw material, the method employs ethanol extraction combined with a series of column chromatography separations to obtain the derivatives. β -Carboline alkaloid derivatives. The extraction method described in this invention is simple and rapid, and the resulting... β-Carboline alkaloid derivatives have high purity, reaching 95% or higher. Attached Figure Description

[0025] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0026] Figure 1 for β - Tangutoid LIII, a carboline alkaloid derivative 1 H-NMR spectrum;

[0027] Figure 2 for β - Tangutoid LIII, a carboline alkaloid derivative 13 C-NMR spectrum;

[0028] Figure 3 for β -HSQC spectrum of Tangutoid LIII, a carboline alkaloid derivative;

[0029] Figure 4 for β -HMBC spectrum of Tangutoid LIII, a carboline alkaloid derivative;

[0030] Figure 5 for β - Tangutoid LIII, a carboline alkaloid derivative 1 H- 1 H COSY spectrum;

[0031] Figure 6 for β - NOESY spectrum of Tangutoid LIII, a carboline alkaloid derivative;

[0032] Figure 7 for β - TOCSY spectrum of Tangutoid LIII, a carboline alkaloid derivative;

[0033] Figure 8 for β (-)-HRESIMS spectrum of Tangutoid LIII, a carboline alkaloid derivative;

[0034] Figure 9 for β-IR spectrum of Tangutoid LIII, a carboline alkaloid derivative;

[0035] Figure 10 for β - UV spectrum of Tangutoid LIII, a carboline alkaloid derivative. Detailed Implementation

[0036] This invention provides β -Carboline alkaloid derivatives, having the structure shown in Formula 1:

[0037] Formula 1.

[0038] The present invention β -The molecular formula of carboline alkaloid derivatives is C 32 H 38 N2O 13 With a novel structure and good monoamine oxidase inhibitory activity, it can slow down the consumption of monoamine neurotransmitters in the brain, reduce oxidative damage, and achieve the treatment of neurodegenerative diseases. Furthermore, it can be used to prepare monoamine oxidase inhibitors or drugs for the treatment of neurodegenerative diseases, showing broad application prospects.

[0039] The present invention also provides the technical solutions described above. β - The extraction method for carboline alkaloid derivatives includes the following steps:

[0040] (1) The juice of Tangut white thorn was subjected to alcohol precipitation to obtain an extract;

[0041] (2) Dissolve the extract in water, and use pure water and 40% ethanol aqueous solution as eluents in sequence. Perform macroporous resin column chromatography on the extract to obtain the eluted fraction of 40% ethanol aqueous solution.

[0042] (3) Using MCI resin as the stationary phase and methanol-water as the eluent, the elution fraction of the 40% ethanol aqueous solution was separated by column chromatography under gradient elution conditions to obtain four components, denoted as Fr. 40-1 ~Fr 40-4 The volume fraction of methanol in the eluent of step (3) is 0%~80%.

[0043] (4) Using MCI resin as the stationary phase and methanol-water as the eluent, Fr was removed under gradient elution conditions. 40-1 The components were separated by column chromatography, yielding 8 components, denoted as Fr. 40-1-1 ~Fr 40-1-8 The volume fraction of methanol in the eluent of step (4) is 0%~50%.

[0044] (5) Using LH-20 gel as the stationary phase and methanol-water as the eluent, Fr was removed under gradient elution conditions. 40-1-7 The components were separated by column chromatography, yielding 10 components, denoted as Fr. 40-1-7-1 ~Fr 40-1-7-10 The volume fractions of methanol in the eluent of step (5) are 0%, 20%, 40%, and 60%, respectively.

[0045] (6) Using C18 bonded silica gel as the stationary phase and acetonitrile-water as the eluent, Fr was removed under isogradient elution conditions. 40-1-7-9 The components were separated by column chromatography to obtain the structure shown in Formula 1. β -Carboline alkaloid derivatives; the eluent contains 15% acetonitrile by volume.

[0046] In this invention, unless otherwise specified, all raw materials used are commercially available products well known to those skilled in the art or prepared using methods well known to those skilled in the art.

[0047] This invention involves ethanol precipitation of Tangut thorn juice to obtain an extract. This invention also involves sequentially crushing and concentrating Tangut thorn fruits to obtain Tangut thorn juice. In a specific embodiment of this invention, the crushing method is juicing. This invention involves crushing Tangut thorn fruits to obtain crushed material; the crushed material is then concentrated to obtain Tangut thorn juice.

[0048] After obtaining the juice of *Nitraria tangutorum*, the present invention performs alcohol precipitation on the juice to obtain an extract. The reagent used for alcohol precipitation in the present invention can be an aqueous ethanol solution. In the present invention, the volume fraction of the aqueous ethanol solution can be 50%~75%; in specific embodiments of the present invention, the volume fraction of the aqueous ethanol solution can be 50%, 55%, 60%, 65%, 70%, or 75%. The number of alcohol precipitations in the present invention can be 3~4 times; in specific embodiments of the present invention, the number of alcohol precipitations can be 3 or 4 times. The present invention does not have special requirements for the temperature of alcohol precipitation; conventional operating temperatures in the art are sufficient. In embodiments of the present invention, alcohol precipitation is carried out at room temperature (20~35℃). In the present invention, the time for each alcohol precipitation can be independently 10~12 hours, and the volume ratio of aqueous ethanol solution to *Nitraria tangutorum* juice can be independently 2~4:1; in specific embodiments of the present invention, the time for each alcohol precipitation is independently 10 hours, 11 hours, or 12 hours, and the volume ratio of aqueous ethanol solution to *Nitraria tangutorum* juice is independently 2:1, 3:1, or 4:1. After each alcohol precipitation, the present invention can perform solid-liquid separation, collect the liquid material, and use the liquid material as the material to be processed for the next extraction; after the final alcohol precipitation, the obtained liquid material is concentrated to obtain the extract. In the present invention, the solid-liquid separation method can be filtration; the concentration can be vacuum distillation. The alcohol precipitation extraction method of the present invention can remove proteins, pectin, and polysaccharides from Tangut white thorn juice; at the same time, ethanol can be recovered by distillation to obtain an extract containing... β - Crude extract of carboline alkaloid derivatives.

[0049] After obtaining the extract, the present invention dissolves the extract in water, and sequentially uses pure water and a 40% (v / v) ethanol aqueous solution as eluents to perform macroporous resin column chromatography, obtaining a 40% (v / v) ethanol aqueous solution eluent. In the present invention, the resin column used for macroporous resin column chromatography can be an AB-8 macroporous resin column. In a specific embodiment of the present invention, the extract is loaded onto an AB-8 macroporous resin column, and sequentially uses pure water, a 40% (v / v) ethanol aqueous solution, and a 95% (v / v) ethanol aqueous solution as eluents for column chromatography separation, respectively obtaining a pure water eluent, a 40% (v / v) ethanol aqueous solution eluent, and a 95% (v / v) ethanol aqueous solution eluent. In the present invention, the column chromatography separation is performed under gradient elution conditions. Subsequent further separation processes do not involve the pure water eluent and the 95% (v / v) ethanol aqueous solution eluent, and will not be described in detail here.

[0050] After obtaining the 40% (v / v) ethanol-water aqueous solution eluent, this invention uses MCI resin as the stationary phase and methanol-water as the eluent, and performs column chromatography separation of the 40% (v / v) ethanol-water aqueous solution eluent under gradient elution conditions to obtain four components, denoted as Fr. 40-1 ~Fr 40-4 In a specific embodiment of the present invention, the elution fraction of the 40% (v / v) ethanol aqueous solution is dissolved in water and then loaded onto an MCI gel chromatography column for column chromatography separation. In this invention, the volume fraction of methanol in the eluent used for column chromatography separation is 0%~80%; in a specific embodiment of the present invention, the volume fraction of methanol in the eluent used for column chromatography separation is successively 0%, 20%, 40%, and 80%, that is, the eluent used for column chromatography separation is successively pure water, 20% methanol-water, 40% methanol-water, and 80% methanol-water. After elution with 80% methanol-water as the eluent, the present invention can use pure methanol as the eluent to wash the chromatographic column used for column chromatography separation. In a specific embodiment of the present invention, the elution fraction of the 40% (v / v) ethanol aqueous solution is loaded onto an MCI gel chromatography column, and gradient elution is performed successively using pure water, 20% methanol-water, 40% methanol-water, 80% methanol-water, and pure methanol as eluents.

[0051] Get Fr 40-1 After componentization, this invention uses MCI resin as the stationary phase and methanol-water as the eluent to remove Fr under gradient elution conditions. 40-1 The components were separated by column chromatography, yielding 8 components, denoted as Fr. 40-1-1 ~Fr 40-1-8 In this invention, the volume fraction of methanol in the eluent used for column chromatography separation is 0%~50%; in a specific embodiment of this invention, the volume fraction of methanol in the eluent used for column chromatography separation is successively 0%, 10%, 20%, 30%, 40%, and 50%, that is, the eluent used for column chromatography separation is successively pure water, 10% methanol-water, 20% methanol-water, 30% methanol-water, 40% methanol-water, and 50% methanol-water. After elution with 50% methanol-water as the eluent, this invention can use pure methanol as the eluent to wash the chromatographic column used for column chromatography separation. In a specific embodiment of this invention, Fr is used... 40-1 The components were loaded onto an MCI gel chromatography column and eluted sequentially using pure water, 10% methanol-water, 20% methanol-water, 30% methanol-water, 40% methanol-water, 50% methanol-water and pure methanol as eluents.

[0052] Get Fr 40-1-7 After fractionation, this invention uses LH-20 gel as the stationary phase and methanol-water as the eluent to remove Fr under gradient elution conditions. 40-1-7The components were separated by column chromatography, yielding 10 components, denoted as Fr. 40-1-7-1 ~Fr 40-1-7-10 In this invention, the volume fraction of methanol in the eluent is 0%, 20%, 40%, and 60%, respectively. In a specific embodiment of this invention, the eluent used for column chromatography is, in sequence, pure water, 20% methanol-water, 40% methanol-water, and 60% methanol-water. After elution with 60% methanol-water, this invention can use pure methanol as the eluent to wash the chromatographic column used for column chromatography. In a specific embodiment of this invention, Fr... 40-1-7 The components were loaded onto an LH-20 gel chromatography column and eluted sequentially using pure water, 20% methanol-water, 40% methanol-water, 60% methanol-water and pure methanol as eluents.

[0053] Get Fr 40-1-7-9 After componentization, this invention uses C18 bonded silica gel as the stationary phase and acetonitrile-water as the eluent to remove Fr under isogradient elution conditions. 40-1-7-9 The components were separated by column chromatography to obtain the structure shown in Formula 1. β -Carboline alkaloid derivatives. In this invention, the volume fraction of acetonitrile in the eluent is 15%. In a specific embodiment of this invention, Fr... 40-1-7-9 The components were loaded onto a preparative liquid chromatography (PCC) Megress C18 column and eluted using 15% acetonitrile-water as the eluent in an isogradient elution process.

[0054] This invention uses the fruit of *Nitraria tangutorum* as raw material, employs ethanol extraction, and combines this with a series of column chromatography separation methods to obtain the desired product. β -Carboline alkaloid derivatives. The extraction method described in this invention is simple and rapid, and the resulting... β -Carboline alkaloid derivatives have high purity, at 95%.

[0055] The present invention also provides the technical solutions described above. β - Application of carboline alkaloid derivatives in the preparation of monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases.

[0056] The present invention β -Carboline alkaloid derivatives have good MAO-A inhibitory activity, IC50 50 The concentration was 151.4 µmol / L; it also exhibited good MAO-B inhibitory activity, IC50... 50 With a concentration of 122.3 µmol / L, it can slow down the consumption of monoamine neurotransmitters in the brain and reduce oxidative damage, thus enabling the treatment of neurodegenerative diseases such as depression and Parkinson's disease. Furthermore, it can be used to prepare monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases.

[0057] The present invention also provides a monoamine oxidase inhibitor, comprising the components described in the above technical solution. β -Carboline alkaloid derivatives. In this invention, the monoamine oxidase inhibitor may include an active ingredient and pharmaceutical excipients; the active ingredient is as described in the above technical solution. β -Carboline alkaloid derivatives.

[0058] The technical solutions of this invention will be clearly and completely described below with reference to the embodiments thereof. Obviously, the described embodiments are only a part of the embodiments of this invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0059] The room temperature conditions described in the examples are 20~35℃.

[0060] Example 1

[0061] (1) Take 110 kg of Tangut white thorn fruit, juice it, and obtain crushed material; concentrate the crushed material under reduced pressure to obtain 25 L of Tangut white thorn juice. Add 50 L of ethanol aqueous solution (volume fraction of 75%) to the Tangut white thorn juice, precipitate with alcohol at room temperature for 12 h, filter after the precipitation, and collect the supernatant; repeat the above alcohol precipitation operation 3 times, and distill the supernatant obtained from the last alcohol precipitation under reduced pressure to recover ethanol and obtain the extract.

[0062] (2) Dissolve the extract obtained in step (1) in water, and load the resulting extract onto an AB-8 macroporous resin column. Use pure water, 40% ethanol-water and 95% ethanol-water as eluents for gradient elution. After the elution, the pure water elution fraction (denoted as Fr0) and the 40% ethanol-water elution fraction (denoted as Fr) are obtained respectively. 40 ) and the 95% ethanol-water elution fraction (denoted as Fr) 95 ).

[0063] (3) Take the Fr obtained in step (2) 40 The components were dissolved in water, and the resulting solution was then loaded onto an MCI gel chromatography column. A gradient elution was performed sequentially with pure water, 20% methanol-water, 40% methanol-water, 80% methanol-water, and pure methanol. Four fractions were obtained after the elution, denoted as Fr. 40-1 ~Fr 40-4 .

[0064] (4) Take the Fr obtained in step (3) 40-1The components were re-loaded onto the MCI gel chromatography column from step (3), and eluted sequentially with pure water, 10% methanol-water, 20% methanol-water, 30% methanol-water, 40% methanol-water, 50% methanol-water, and pure methanol. After the elution, eight components were obtained, denoted as Fr. 40-1-1 ~Fr 40-1-8 .

[0065] (5) Take the Fr obtained in step (4) 40-1-7 The fractions were loaded onto an LH-20 gel chromatography column and eluted sequentially with pure water, 20% methanol-water, 40% methanol-water, 60% methanol-water, and pure methanol. Ten fractions were obtained after the elution process, denoted as Fr. 40-1-7-1 ~Fr 40-1-7-10 .

[0066] (6) Take the Fr obtained in step (5) 40-1-7-9 The sample was loaded onto a Megress C18 preparative liquid chromatography column and isocratic eluted with a 15% (v / v) acetonitrile aqueous solution to obtain one of the components from *Nitraria tangutorum*. β -Carboline alkaloid derivatives (125.8 mg), the aforementioned β -The carboline alkaloid derivative was named Tangutoid LIII.

[0067] Test Example 1

[0068] Using nuclear magnetic resonance hydrogen spectroscopy (NMR) 1 H-NMR, carbon nuclear magnetic resonance (NMR) 13 C-NMR, heteronuclear single quantum correlation spectroscopy (HSQC), heteronuclear multi-bond correlation spectroscopy (HMBC), hydrogen-hydrogen correlation spectroscopy (C-NMR), heteronuclear single quantum correlation spectroscopy (HSQC), heteronuclear multi-bond correlation spectroscopy (HMBC), hydrogen-hydrogen correlation spectroscopy (HMBC) 1 H- 1 The structure of the compound Tangutoid LIII prepared in Example 1 was determined using H COSY, NOESY, TOCSY, HRESIMS, infrared (IR), and ultraviolet (UV) spectroscopy. The results are as follows: Figures 1-10 As shown, the compound Tangutoid LIII 1 H-NMR and 13 C-NMR data in DMSO- d The NMR data obtained from the measurements are shown in Table 1.

[0069] Table 1. Compound Tangutoid LIII 1 H-NMR (600 MHz) and 13 C-NMR (150 MHz) data

[0070]

[0071] The specific spectral data of compound Tangutoid LIII are as follows: white powder, [ α ] D 20.1 -35.70 (c0.118, MeOH); UV (MeOH) λ max nm (log ε ): 315.00 (0.2059), 291.50 (0.1895), 223.50(0.3970), 201.50 (0.2937); IR (KBr) ν max : 3385, 2933, 1692, 1630, 1604, 1514,1395, 1262, 1170 cm -1 ; NMR spectroscopic date see table 1; HRESIMS m / z 673.2256[MH]- (calculated for C 32 H 37 N2O 14 673.2250).

[0072] Furthermore, the compound Tangutoid LIII has a high purity of 95%; the structural formula of the compound Tangutoid LIII is shown below:

[0073] .

[0074] Test Example 2

[0075] The monoamine oxidase inhibitory activity of the compound Tangutoid LIII obtained in Example 1 was tested, and the specific procedures are as follows:

[0076] (1) Monoamine oxidase extraction

[0077] Place 10g of commercially available pork liver in a juicer cup, add 400g of 0.3mol / L sucrose aqueous solution pre-cooled to 4℃, and start juicing to obtain pork liver slurry. Then, centrifuge the pork liver slurry at 1000g (2100r / min) and 4℃ for 10min to remove surface foam and collect the supernatant. Next, centrifuge the supernatant at 10000g (6800r / min) and 4℃ for 30min to collect the precipitate. Add 25mL of phosphate-buffered saline (PBS, 0.1mol / L, pH=7.4) pre-cooled to 25℃ to the precipitate, aliquot, and store at -80℃ to obtain monoamine oxidase enzyme solution.

[0078] (2) Evaluation of monoamine oxidase inhibitor activity

[0079] The compound Tangutoid LIII obtained in Example 1 was added to dimethyl sulfoxide (DMSO) to obtain compound solutions with gradient concentrations, which were then set aside. 2 μL of the compound solutions were mixed with 100 μL of PBS aqueous solution to obtain a compound-PBS composite solution.

[0080] Take two identical petri dishes and label them Reaction System A and Reaction System B, respectively. Add 8 μL of monoamine oxidase enzyme solution and 102 μL of compound-PBS composite solution to each of Reaction System A and Reaction System B, and incubate at 37°C for 15 min. Add 40 μL of Amplex Red solution (0.25 mg / mL), 10 μL of horseradish peroxidase solution (0.3 U / mL), and 40 μL of 0.1 mmol / L 5-hydroxytryptamine hydrochloride solution (as MAO-A substrate) to Reaction System A. Add 40 μL of Amplex Red solution to Reaction System B. Red solution (0.25 mg / mL), 10 μL horseradish peroxidase solution (0.3 U / mL), and 40 μL benzylamine hydrochloride solution (0.1 mmol / L as MAO-B substrate) were used. Reaction system A and reaction system B were reacted at 37 °C for 24 min. After the reaction, the difference in optical density before and after the reaction was measured at 570 nm. The inhibition rate I of compound Tangutoid LIII on MAO-A or MAO-B was calculated.

[0081] The inhibition rate I is calculated using the following formula: I=1 D1 / D2 and ΔD1 represent the difference in optical density before and after the reaction in the experimental group, while ΔD2 represents the difference in optical density before and after the reaction in the blank group without the addition of monoamine oxidase solution.

[0082] The half-maximal inhibitory concentrations (IC50) of compound Tangutoid LIII against MAO-A and MAO-B were calculated using Graphpad 8.0.2. 50 IC 50 The test results are shown in Table 2.

[0083] Table 2. Inhibitory activities of compound Tangutoid LIII for MAO-A and MAO-B.

[0084]

[0085] Note: IC in Table 2 50 The test results are the average value ± SD (n=3).

[0086] As shown in Table 2, compound Tangutoid LIII exhibits excellent inhibitory activity against both MAO-A and MAO-B. Specifically, the IC50 of Tangutoid LIII's inhibitory activity against MAO-A is [not specified in the original text]. 50 The value was 151.4 µmol / L, and the IC50 value for the inhibitory activity against MAO-B was 151.4 µmol / L. 50 The value is 122.3 µmol / L.

[0087] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A kind β -Carboline alkaloid derivatives, characterized in that... It has the structure shown in Equation 1: Formula 1.

2. The claim 1 β -A method for extracting carboline alkaloid derivatives, characterized in that, Includes the following steps: (1) The juice of Tangut white thorn was subjected to alcohol precipitation to obtain an extract; (2) Dissolve the extract in water, and use pure water and 40% ethanol aqueous solution as eluents in sequence. Perform macroporous resin column chromatography on the extract to obtain the elution fraction of 40% ethanol aqueous solution. (3) Using MCI resin as the stationary phase and methanol-water as the eluent, the eluted fraction of the 40% (v / v) ethanol aqueous solution was separated by column chromatography under gradient elution conditions to obtain four components, denoted as Fr. 40-1 ~Fr 40-4 The volume fraction of methanol in the eluent of step (3) is 0%~80%. (4) Using MCI resin as the stationary phase and methanol-water as the eluent, Fr was removed under gradient elution conditions. 40-1 The components were separated by column chromatography, yielding 8 components, denoted as Fr. 40-1-1 ~Fr 40-1-8 The volume fraction of methanol in the eluent of step (4) is 0%~50%. (5) Using LH-20 gel as the stationary phase and methanol-water as the eluent, Fr was removed under gradient elution conditions. 40-1-7 The components were separated by column chromatography, yielding 10 components, denoted as Fr. 40-1-7-1 ~Fr 40-1-7-10 The volume fractions of methanol in the eluent of step (5) are 0%, 20%, 40%, and 60%, respectively. (6) Using C18 bonded silica gel as the stationary phase and acetonitrile-water as the eluent, Fr was removed under isogradient elution conditions. 40-1-7-9 The components were separated by column chromatography to obtain the structure shown in Formula 1. β -Carboline alkaloid derivatives; the eluent contains 15% acetonitrile by volume.

3. The extraction method according to claim 2, characterized in that, In step (1), the reagent used for alcohol precipitation is an aqueous ethanol solution; the volume fraction of the aqueous ethanol solution is 50%~75%.

4. The extraction method according to claim 2 or 3, characterized in that, The alcohol precipitation is performed 3 to 4 times.

5. The extraction method according to claim 2 or 3, characterized in that, The alcohol precipitation time was 10-12 h for each precipitation, and the volume ratio of ethanol aqueous solution to Tangut white thorn juice was 2-4:

1.

6. The extraction method according to claim 2, characterized in that, In step (3), the volume fraction of methanol in the eluent used for column chromatography separation is 0%, 20%, 40% and 80% respectively.

7. The extraction method according to claim 2, characterized in that, In step (4), the volume fraction of methanol in the eluent used for column chromatography separation is 0%, 10%, 20%, 30%, 40% and 50% respectively.

8. The extraction method according to claim 2, characterized in that, In step (2), the chromatographic column used for macroporous resin column chromatography is an AB-8 macroporous resin column.

9. The claim 1 β - Application of carboline alkaloid derivatives in the preparation of monoamine oxidase inhibitors or drugs for treating neurodegenerative diseases.

10. A monoamine oxidase inhibitor comprising the β-carboline alkaloid derivative of claim 1.

Images