A matrine alkaloid, its preparation method and use

Abstract

The present invention discloses a matrine alkaloid, its preparation method and use. The matrine alkaloid is selected from any one of new compounds 1 to 16, and is obtained by extraction and separation from the rhizome of Sophora tonkinensis or obtained by artificial synthesis. The present invention also provides a preparation method of the matrine alkaloid, and verifies that the matrine alkaloid has a significant effect in preventing and treating TSWV and poisoning the western flower thrips which is the transmission vector of TSWV.

Description

Technical Field

[0001] The present invention relates to the technical field of pesticides for crops, in particular to matrine alkaloids and their preparation methods and uses. Background Art

[0002] Plant virus diseases are the second largest category of plant diseases after plant fungal diseases. They have strong specificity, great harm, and are difficult to control. The economic losses caused worldwide each year are as high as $60 billion, of which only food crops amount to $20 billion. Tomato spotted wilt virus (TSWV) is one of the top 10 recognized plant viruses globally and is also the only virus in the genus Tomato spotted wilt virus that can infect plants. TSWV can infect 1,090 plant species in 84 families, especially severely infecting crops such as soybean, peanut, celery, pumpkin, pepper, potato, and tomato, and causing the most serious harm in temperate and subtropical regions. In China, TSWV was first detected in peppers in 2003, and then the harm range has been expanding year by year, and it has been reported in various horticultural plants in Sichuan, Yunnan, Guangzhou, Shandong, Yunnan, Hebei, Ningxia Hui Autonomous Region, Shaanxi, Qinghai and other places. At present, the harm of TSWV to crop production is becoming increasingly serious, and greenhouse, greenhouse, and field-cultivated tomatoes are all damaged. In severe cases, the yield loss can exceed 80% or even result in a complete harvest failure. TSWV can be transmitted by a variety of thrips and mechanically, and Frankliniella occidentalis is the main transmission vector. Frankliniella occidentalis has the characteristics of a wide host range, miscellaneous feeding habits, and strong reproductive ability, and once it carries the virus, it will carry the virus for life. TSWV can replicate and proliferate in Frankliniella occidentalis, greatly enhancing the virus transmission efficiency. Currently, with the frequent cross-infection of Frankliniella occidentalis and TSWV, and the variability of the TSWV virus, the current control measures can only inhibit or reduce the proliferation of the virus in plants to a certain extent, and cannot completely eliminate TSWV. In view of the current serious harm of plant virus disease TSWV, the still lack of agents against plant virus TSWV, the increasing drug resistance of the virus, and the environmental pollution caused by pesticide residues, the development of efficient, environmentally friendly, and safe anti-TSWV virus agents has become the focus and hotspot of the research and development of new pesticides against plant virus TSWV. Summary of the Invention

[0003] The purpose of the present invention is to provide matrine alkaloids and their preparation methods and uses. The present invention extracts a new matrine alkaloid from the rhizome of Sophora tonkinensis, and through efficacy tests, it has been proved to have the characteristics of effectively preventing and controlling TSWV and killing insects.

[0004] The technical solution of the present invention: matrine alkaloids, wherein the matrine alkaloids are selected from any one of Compounds 1 to 16 with the following chemical structures:

[0005]

[0006] The matrine alkaloids are obtained by extraction and isolation from the roots and rhizomes of Sophora tonkinensis or by artificial synthesis.

[0007] The method for preparing the matrine alkaloids as described above comprises the following steps:

[0008] (1) Take the dried roots and rhizomes of Sophora tonkinensis, reflux and extract with methanol, combine the extraction solutions and concentrate to obtain an extract, and subject the extract to acid-base treatment to obtain total alkaloids;

[0009] (2) Subject the total alkaloids to silica gel column chromatography with 300 - 400 mesh, and use dichloromethane:methanol with a volume ratio of 100:0 - 0:100 as the mobile phase for gradient elution to obtain fractions A, B, C, D, and E;

[0010] (3) Subject fraction D to MCI column chromatography, and use methanol:water with a volume ratio of 0:100 - 100:0 as the mobile phase for gradient elution to obtain D-1, D-2, D-3, and D-4;

[0011] (4) Subject D-1 to silica gel column chromatography with 300 - 400 mesh, and use petroleum ether:acetone:diethylamine with a volume ratio of 100:1:0.125 - 1:1:0.005 as the mobile phase for gradient elution to obtain compound 14 and compound 12;

[0012] (5) Subject D-2 to methanol gel purification and then to semi-preparative HPLC purification, and use methanol:water with a volume ratio of 21:79 as the mobile phase for elution to obtain compound 6, compound 7, and compound 1;

[0013] (6) Subject D-3 to silica gel column chromatography with 300 - 400 mesh, and use dichloromethane:methanol with a volume ratio of 50:1 as the mobile phase for gradient elution, and then subject it to methanol gel purification to obtain compound 8, compound 9, compound 4, and compound 5;

[0014] (7) Subject D-4 to silica gel column chromatography with 300 - 400 mesh, and use petroleum ether:acetone:diethylamine with a volume ratio of 50:1:0.125 - 1:1:0.005 as the mobile phase for gradient elution to obtain compound 10 and compound 15;

[0015] (8) Subject fraction E to silica gel column chromatography with 300 - 400 mesh, and use dichloromethane:methanol with a volume ratio of 20:1 - 0:100 as the mobile phase for gradient elution to obtain E-1, E-2, E-3, E-4, and E-5;

[0016] (9) E-1 was subjected to silica gel column chromatography (300 - 400 mesh), gradient elution was performed using petroleum ether:acetone:diethylamine with a volume ratio of 25:1:0.005 - 1:1:0.005 as the mobile phase, then purified by methanol gel, further purified by semi-preparative HPLC, and finally eluted using acetonitrile:water with a volume ratio of 17:83 as the mobile phase to obtain Compound 11;

[0017] (10) E-3 was purified by methanol gel, then subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was performed using petroleum ether:acetone:diethylamine with a volume ratio of 10:1:0.005 - 1:1:0.005 as the mobile phase to obtain Compound 16;

[0018] (11) E-4 was subjected to silica gel column chromatography (300 - 400 mesh), gradient elution was performed using dichloromethane:methanol with a volume ratio of 25:1 as the mobile phase, then purified by semi-preparative HPLC, and eluted using methanol:water with a volume ratio of 34:66 as the mobile phase to obtain Compound 2 and Compound 3;

[0019] (12) E-5 was purified by methanol gel, then purified by semi-preparative HPLC, and eluted using methanol:water with a volume ratio of 36:64 as the mobile phase to obtain Compound 13.

[0020] In the aforementioned preparation method, in step (1), Sophora tonkinensis is a plant of the genus Sophora in the legume family.

[0021] In the aforementioned preparation method, in step (1), the reflux extraction was performed 3 times, each time for 3 hours.

[0022] In the aforementioned preparation method, in step (1), the acid-base treatment specifically is as follows: first, the pH value was adjusted to 1 - 3 with 0.1% hydrochloric acid, then non-alkaloid components were removed by petroleum ether extraction. The acid aqueous layer was then adjusted to pH 9 - 11 with 25% - 28% concentrated ammonia water, and then the basic aqueous layer was successively extracted with dichloromethane and n-butanol.

[0023] In the aforementioned preparation method, in step (2), fraction A was eluted with dichloromethane:methanol with a volume ratio of 50:1, fraction B was eluted with dichloromethane:methanol with a volume ratio of 20:1, fraction C was eluted with dichloromethane:methanol with a volume ratio of 10:1, fraction D was eluted with dichloromethane:methanol with a volume ratio of 5:1, and E was eluted with dichloromethane:methanol with a volume ratio of 2:1.

[0024] In the aforementioned preparation method, in step (3), D-1 was eluted with methanol:water with a volume ratio of 20:80, D-2 was eluted with methanol:water with a volume ratio of 40:60, D-3 was eluted with methanol:water with a volume ratio of 60:40, and D-4 was eluted with methanol:water with a volume ratio of 80:20.

[0025] In the aforementioned preparation method, in step (4), compound 14 is obtained by eluting with petroleum ether:acetone:diethylamine at a volume ratio of 20:1:0.005, and compound 12 is obtained by eluting with petroleum ether:acetone:diethylamine at a volume ratio of 10:1:0.005.

[0026] In the aforementioned preparation method, in step (7), compound 10 is obtained by eluting with petroleum ether:acetone:diethylamine at a volume ratio of 10:1:0.005, and compound 15 is obtained by eluting with petroleum ether:acetone:diethylamine at a volume ratio of 2:1:0.005.

[0027] In the above-mentioned preparation method, in step (8), E-1 is obtained by eluting with dichloromethane:methanol at a volume ratio of 15:1, E-2 is obtained by eluting with dichloromethane:methanol at a volume ratio of 10:1, E-3 is obtained by eluting with dichloromethane:methanol at a volume ratio of 5:1, E-4 is obtained by eluting with dichloromethane:methanol at a volume ratio of 2:1, and E-5 is obtained by eluting with dichloromethane:methanol at a volume ratio of 1:1.

[0028] In the aforementioned preparation method, in step (10), the compound 16 is obtained by eluting with petroleum ether:acetone:diethylamine in a volume ratio of 5:1:0.005.

[0029] The matrine alkaloids prepared by the above method are white powders. The H NMR spectra of compounds 1 to 16 ( 1 H-NMR) and carbon spectra ( 13 C-NMR) data are shown in Tables 1 to 6, where Table 1 is the hydrogen spectrum of compound 1 ( 1 H-NMR) and carbon spectra ( 13 C-NMR) data, Table 2 is the hydrogen spectrum of compound 2-compound 6 ( 1 H-NMR) data, Table 3 is the hydrogen spectrum of compound 7-compound 11 ( 1 H-NMR) data; Table 4 is the hydrogen spectrum of compound 12-compound 16 ( 1 H-NMR) data; Table 5 is the carbon spectra of compound 2-compound 9 ( 13 C-NMR) data; Table 6 is the carbon spectra of compound 10-compound 16 ( 13 C-NMR) data.

[0030] Table 1. Compound 1 1 H and 13 C NMR data (δ in ppm, Jin Hz)

[0031]

[0032]

[0033] a 600 MHz in CDCl 3

[0034] Table 2. For compounds 2 - 6 1 1H NMR data (δ in ppm, J in Hz)

[0035]

[0036]

[0037] a 150 MHz in CDCl 3 ; b 150 MHz in CD 3 OD.

[0038] Table 3. For compounds 7 - 11 1 1H NMR data (δ in ppm, J in Hz)

[0039]

[0040]

[0041] a 150 MHz in CDCl 3 ; b 150 MHz in CD 3 OD.

[0042] Table 4. For compounds 12 - 16 1 1H NMR data (δ in ppm, J in Hz)

[0043]

[0044]

[0045] a 150 MHz in CDCl 3 ; b 150 MHz in CD 3 OD.

[0046] Table 5. For compounds 2 - 9 13 13C NMR data (δ in ppm)

[0047]

[0048]

[0049] a 150 MHz in CDCl 3 ; b 150 MHz in CD 3 OD.

[0050] Table 6. 13 C NMR data (δ in ppm) of Compounds 9 - 16

[0051] Position <![CDATA[10 a > <![CDATA[11 a > <![CDATA[12 a > <![CDATA[13 a > <![CDATA[14 a > <![CDATA[15 a > <![CDATA[16 a > 2 46.2 44.5 57.1 50.2 50.2 55.8 56.2 3 23.1 22.3 20.8 19.7 19.8 20.8 22.4 4 27.3 35.0 27.6 24.1 24.1 26.5 37.5 5 113.9 208.4 35.6 115.0 114.6 34.1 71.6 6 171.7 169.4 63.7 152.2 152.0 63.4 70.7 7 205.3 45.6 42.6 113.8 113.4 60.1 71.5 8 36.3 26.9 26.8 23.9 22.0 208.9 34.0 9 29.0 19.5 20.6 19.6 19.7 40.0 21.5 10 46.3 47.4 57.2 49.6 49.6 56.5 56.4 11 64.3 64.2 59.7 148.6 148.9 52.5 56.7 12 25.3 30.0 27.0 30.2 29.7 26.6 19.0 13 17.5 17.1 20.8 23.0 23.6 18.4 21.6 14 33.1 31.0 33.9 34.7 32.8 32.7 32.6 15 169.2 172.2 173.9 175.2 173.7 169.2 173.3 17 122.2 60.9 40.8 134.8 135.1 41.4 47.0 <![CDATA[OCH 3 > - - 51.5 - 51.6 - -

[0052] a 150 MHz in CDCl 3 ; b 150 MHz in CD 3 OD.

[0053] After the above structural identification, Compounds 1 - 16 are as follows:

[0054] Compound 1: White powder; [α]25D – 102.5 (c 0.12, MeOH); UV (MeOH) λ max (logε): 201 (4.36) nm; IR (KBr) ν max 2935, 1710, 1640, 1470, 1334, 1196, 1101 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Table 1 and Table 1; HRESIMS m / z 571.3134 [M + H] + , (calcd for C 30 H 43 N 4 O 7 , 571.3126).

[0055] Compound 2: Colorless transparent crystals; mp 182 - 83 °C; [α]25D + 65.9 (c 0.04, MeOH); UV (MeOH) λ max (logε): 200 (4.17) nm; IR (KBr) ν max 2922, 1624, 1454, 1415, 1385, 1129 cm -1 ; 1 H and 13 C NMR data (CDCl 3, 600 and 150 MHz) are shown in Tables 2 and 5; HRESIMS m / z 315.1674 [M+Na] + (calcd for C 16 H 24 N 2 O 3 Na, 315.1679).

[0056] Compound 3: White powder; [α]25D +67.4 (c 0.13, MeOH); UV (MeOH) λ max (logε): 201 (3.99) nm; IR (KBr) ν max 3409, 2926, 1622, 1455, 1385, 1178, 1127 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 2 and 5; HRESIMS m / z 279.1695 [M+H] + (calcd for C 15 H 23 N 2 O 3 , 279.1703).

[0057] Compound 4: White powder; [α]25D +61.8 (c 0.09, MeOH); UV (MeOH) λ max (logε): 200 (4.05) nm; IR (KBr) ν max 3394, 2943, 1627, 1541, 1447, 1372, 1125 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 2 and 5; HRESIMS m / z 342.1794 [M+Na] + (calcd for C 17 H 25 N 3 O 3 Na, 342.1788).

[0058] Compound 5: White powder; [α]25D +87.7 (c 0.12, MeOH); UV (MeOH) λ max (logε): 200 (3.97) nm; IR (KBr) ν max2959, 1664, 1414, 1351, 1041, 1026 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 2 and 5; HRESIMS m / z 313.1526 [M+H] + (calcd for C 16 H 22 N 2 O 3 Na, 313.1523).

[0059] Compound 6: Colorless oil; [α]25D +59.6 (c 0.04, MeOH); UV (MeOH) λ max (logε): 201 (4.14) nm; IR (KBr) ν max 2940, 1683, 1635, 1412, 1385, 1332, 1206, 1180, 1132 cm -1 ; 1 H and 13 C NMR data (CD 3 OD, 600 and 150 MHz) are shown in Tables 2 and 5; HRESIMS m / z 299.1358 [M+Na] + (calcd for C 15 H 20 N 2 O 3 Na, 299.1366).

[0060] Compound 7: White powder; [α]25D +51.5 (c 0.03, MeOH); UV (MeOH) λ max (logε): 200 (4.10), 238 (3.58) nm; IR (KBr) ν max 2947, 1682, 1647, 1384, 1205, 1180, 1133, 1085, 801, 722 cm -1 ; 1 H and 13 C NMR data (CD 3 OD, 600 and 150 MHz) are shown in Tables 3 and 5; HRESIMS m / z 299.1357 [M+H] + (calcd for C 15 H 20 N 2 O 3 Na, 299.1366).

[0061] Compound 8: white powder; [α]25D +73.8 (c 0.12, MeOH); UV (MeOH) λ max (logε): 201 (3.95), 231 (3.55) nm; IR (KBr) v max 2937, 1748, 1672, 1634, 1449, 1310, 1222, 1013 cm -1 ; 1 Hand 13 13C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 3 and 5; HRESIMS m / z 359.1581 [M+H] + (calcd for C 17 H 24 N 2 O 5 Na, 359.1577).

[0062] Compound 9: white powder; [α]25D –29.0 (c 0.10, MeOH); UV (MeOH) λ max (logε): 200 (3.78) nm; IR (KBr) v max 2938, 1736, 1669, 1670, 1645, 1452, 1213, 1015 cm -1 ; 1 H and 13 13C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 3 and 5; HRESIMS m / z 359.1581 [M+Na] + (calcd for C 17 H 24 N 2 O 5 Na, 359.1577).

[0063] Compound 10: colorless transparent crystal; mp 225-226 °C; [α]25D +45.5 (c 0.10, MeOH); UV (MeOH) λ max (logε): 208 (3.65), 250 (3.95) nm; IR (KBr) v max 2945, 1710, 1639, 1480, 1404, 1253 cm -1 ; 1 H and 13 13C NMR data (CDCl 3, 600 and 150 MHz) see Tables 3 and 6; HRESIMS m / z 299.1370 [M+H] + (calcd for C 15 H 20 N 2 O 3 Na, 299.1366).

[0064] Compound 11: White powder; [α]25D +36.9 (c 0.07, MeOH); UV (MeOH) λ max (logε): 201 (4.25), 240 (2.52) nm; IR (KBr) ν max 2932, 1671, 1644, 1459, 1396, 1367, 1273, 1180 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) see Tables 3 and 6; HRESIMS m / z 301.1515 [M+Na] + (calcd for C 15 H 22 N 2 O 3 Na, 301.1523).

[0065] Compound 12: Colorless needle crystals; mp 195 - 196 °C; [α]25D +65.5 (c 0.03, MeOH); UV (MeOH) λ max (logε): 200 (3.91) nm; IR (KBr) ν max 2936, 1737, 1634, 1444, 1385, 1202, 1166, 1096 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) see Tables 4 and 6; HRESIMS m / z 332.1952 [M+Na] + (calcd for C 16 H 27 N 3 O 3 Na, 332.1945).

[0066] Compound 13: White powder; UV (MeOH) λ max (logε): 220 (4.12), 251 (3.07), 296 (4.13) nm; IR (KBr) νmax 3375, 2919, 1670, 1567, 1475, 1382 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 4 and 6; HRESIMS m / z 260.1760 [M+H] + (calcd for C 15 H 22 N 3 O, 260.1757).

[0067] Compound 14: White powder; UV (MeOH) λ max (logε): 221 (3.87), 251 (3.13), 297 (3.87) nm; IR (KBr) ν max 2927, 2358, 1733, 1635, 1594, 1557, 1557, 1201 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 4 and 6; HRESIMS m / z 275.1755 [M+Na] + (calcd for C 16 H 23 N 2 O 2 , 275.1754).

[0068] Compound 15: White powder; [α]25D +55.3 (c 0.04, MeOH); UV (MeOH) λ max (logε): 200 (4.25), 259 (3.51) nm; IR (KBr) ν max 2932, 1671, 1664, 1396, 1273, 1180, 1140, 1084 cm -1 ; 1 Hand 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Tables 4 and 6; HRESIMS m / z 285.1580 [M+Na] + (calcd for C 15 H 22 N 2 O 2 Na, 285.1574).

[0069] Compound 16: white powder; [α]25D +40.8 (c 0.06, MeOH); UV (MeOH) λ max (logε): 201 (4.12) nm; IR (KBr) ν max 3426, 2927, 1626, 1415, 1384, 1131, 1040 cm -1 ; 1 H and 13 C NMR data (CDCl 3 , 600 and 150 MHz) are shown in Table 4 and Table 6; HRESIMS m / z 303.1675 [M+Na] + (calcd for C 15 H 24 N 2 O 3 Na, 303.1679).

[0070] The present invention also provides a pharmaceutical composition, comprising the matrine alkaloid or its salt as described above and an agriculturally acceptable carrier thereof.

[0071] The pharmaceutical composition is a pharmaceutical preparation. The pharmaceutical preparation includes: powder, wettable powder, emulsifiable concentrate, granule, suspension, soluble liquid, milk powder, aqueous solution, smoke agent, seed coating agent, microcapsule agent, etc.

[0072] The present invention also provides the application of the matrine alkaloid as described above or the pharmaceutical composition as described above in the preparation of a pesticide or insecticide against plant virus TSWV.

[0073] Compared with the prior art:

[0074] The present invention uses the roots and rhizomes of Sophora tonkinensis as raw materials, adopts a new extraction and separation method, the preparation method is simple and easy to operate, new compounds are extracted therefrom, the structures are novel, and they are identified as matrine alkaloids. Through experimental verification, the matrine alkaloids have strong inhibitory effects on tomato spotted wilt virus in terms of treatment and prevention, and have strong insecticidal effects on the virus vector Frankliniella occidentalis. Specific embodiments

[0075] The following examples are used to further illustrate the present invention, but do not serve as a basis for limiting the present invention.

[0076] Example 1:

[0077] The matrine alkaloid is any one of Compounds 1 to 16 with the following chemical structures:

[0078]

[0079] The matrine alkaloids are obtained by extraction and separation from the roots and rhizomes of Sophora tonkinensis or by artificial synthesis.

[0080] A method for preparing matrine alkaloids comprises the following steps:

[0081] (1) Take 45 kg of dry roots and rhizomes of Sophora tonkinensis, reflux extract with methanol three times, each time for 3 hours, combine the extraction solutions and concentrate to obtain an extract. The extract is treated with acid and alkali, that is, first adjust the pH value to 2 with 0.1% hydrochloric acid, then extract with petroleum ether to remove non-alkaloid components. The acid aqueous layer is then adjusted to pH 10 with 26% concentrated ammonia water, and then the alkaline aqueous layer is successively extracted with dichloromethane and n-butanol to obtain 1.7 kg of total alkaloids;

[0082] (2) Subject the total alkaloids to silica gel column chromatography with 300 - 400 mesh, and perform gradient elution with dichloromethane:methanol with a volume ratio of 100:0 - 0:100 as the mobile phase. Elute with a volume ratio of 50:1 to obtain fraction A, with a volume ratio of 20:1 to obtain fraction B, with a volume ratio of 10:1 to obtain fraction C, with a volume ratio of 5:1 to obtain fraction D, and with a volume ratio of 2:1 to obtain fraction E;

[0083] (3) Separate fraction D by MCI column chromatography, and perform gradient elution with methanol:water with a volume ratio of 0:100 - 100:0 as the mobile phase. Elute with a volume ratio of 20:80 to obtain D-1, with a volume ratio of 40:60 to obtain D-2, with a volume ratio of 60:40 to obtain D-3, and with a volume ratio of 80:20 to obtain D-4;

[0084] (4) Subject D-1 to silica gel column chromatography with 300 - 400 mesh, and perform gradient elution with petroleum ether:acetone:diethylamine with a volume ratio of 100:1:0.125 - 1:1:0.005 as the mobile phase. Elute with a volume ratio of 20:1:0.125 to obtain 23.2 mg of compound 14, and with a volume ratio of 10:1:0.005 to obtain 52.7 mg of compound 12;

[0085] (5) Purify D-2 by methanol gel and then by semi-preparative HPLC, and elute with methanol:water with a volume ratio of 21:79 as the mobile phase to obtain 27.7 mg of compound 6, 21.6 mg of compound 7, and 9.3 mg of compound 1;

[0086] (6) Subject D-3 to silica gel column chromatography with 300 - 400 mesh, and perform gradient elution with dichloromethane:methanol with a volume ratio of 50:1 as the mobile phase, and then purify by methanol gel to obtain 9.5 mg of compound 8, 13.2 mg of compound 9, 24.3 mg of compound 4, and 8.5 mg of compound 5;

[0087] (7) D-4 was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was carried out with petroleum ether:acetone:diethylamine in a volume ratio of 50:1:0.125 - 1:1:0.005 as the mobile phase. Compound 10 (23.5 mg) was obtained by elution with a volume ratio of 10:1:0.125, and compound 15 (8.4 mg) was obtained by elution with a volume ratio of 2:1:0.005;

[0088] (8) Fraction E was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was carried out with dichloromethane:methanol in a volume ratio of 20:1 - 0:100 as the mobile phase. E-1 was obtained by elution with a volume ratio of 15:1, E-2 was obtained by elution with a volume ratio of 10:1, E-3 was obtained by elution with a volume ratio of 5:1, E-4 was obtained by elution with a volume ratio of 2:1, and E-5 was obtained by elution with a volume ratio of 1:1;

[0089] (9) E-1 was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was carried out with petroleum ether:acetone:diethylamine in a volume ratio of 25:1:0.005 - 1:1:0.005 as the mobile phase. Then it was separated by methanol gel and further purified by semi-preparative HPLC. Finally, elution was carried out with acetonitrile:water in a volume ratio of 17:83 as the mobile phase to obtain 34.1 mg of compound 11;

[0090] (10) E-3 was purified by methanol gel and then subjected to silica gel column chromatography (300 - 400 mesh). Gradient elution was carried out with petroleum ether:acetone:diethylamine in a volume ratio of 10:1:0.005 - 1:1:0.005 as the mobile phase. Compound 16 (46.9 mg) was obtained by elution with a volume ratio of 5:1:0.005;

[0091] (11) E-4 was subjected to silica gel column chromatography (300 - 400 mesh) and eluted with dichloromethane:methanol in a volume ratio of 25:1 as the mobile phase. Then it was purified by semi-preparative HPLC and eluted with methanol:water in a volume ratio of 34:66 as the mobile phase to obtain 36.2 mg of compound 2 and 43.9 mg of compound 11.

[0092] (12) E-5 was purified by methanol gel and then by semi-preparative HPLC. Elution was carried out with methanol:water in a volume ratio of 36:64 as the mobile phase to obtain 26.1 mg of compound 13.

[0093] A pharmaceutical composition comprising the above-mentioned matrine alkaloids or their salts and a pesticidally acceptable carrier.

[0094] The present invention also provides the use of the above-mentioned matrine alkaloids or pharmaceutical composition in the preparation of a pesticide or insecticide against plant virus TSWV.

[0095] Experimental Example 1:

[0096] The matrine-type alkaloids obtained in Example 1 were studied and verified for their anti-TSWV activity. The specific method is as follows:

[0097] a. The host was the common tobacco cultivar K326 (Nicotiana tabacum cv. K326), and the seeds were provided and cultivated in a glass greenhouse by this experiment. The tomato spotted wilt virus (TSWV) source was provided by this laboratory. Ningnanmycin was purchased from Heilongjiang Deqiang Biology Co., Ltd.

[0098] b. Inoculating the virus and applying the drug:

[0099] Applying the drug after inoculating the virus: Select tobacco seedlings at the 5-6 true leaf stage with consistent growth, and inoculate TSWV by rubbing on the 4th and 5th true leaves. After 10 min, rinse with water. After 24 h, apply 1 mL of compounds 1-16 with a concentration of 100 μg / mL respectively, smear evenly, and rinse with water after 10 min; there were 2 positive controls. One positive control was to first rub and inoculate TSWV, and then apply DMSO (diluted to 100 μg / mL with sterile water) after 24 h for calculating the inhibition rate; the other positive control was to first rub and inoculate TSWV, apply ningnanmycin after 24 h, and then apply DMSO as a blank control after 24 h. The plants grew in a pest-free greenhouse. The experiment was repeated 3 times, and the number of necrotic spots was counted 3 days after smearing to calculate the inhibition rate.

[0100] Inhibition rate (%) = (number of necrotic spots in the positive control - number of necrotic spots in the treatment) / number of necrotic spots in the positive control × 100% Applying the drug before inoculating the virus: Select tobacco seedlings at the 5-6 true leaf stage with consistent growth, and apply 1 mL of compounds 1-16 with a concentration of 100 μg / mL respectively on the 4th and 5th true leaves, smear evenly, and rinse with water after 10 min. After 6 h, inoculate TSWV by rubbing and rinse with water after 10 min. There were 2 positive controls. One positive control was to first apply DMSO (100 μg / mL), and then inoculate TSWV after 6 h for calculating the inhibition rate; the other positive control was to first apply ningnanmycin, and then inoculate TSWV after 6 h, and only apply DMSO as a blank control. The tobacco seedlings grew in a pest-free greenhouse, and the experiment was repeated 3 times. The number of necrotic spots was counted 3 days after inoculating the virus, and the inhibition rate was calculated. The detection results are shown in Table 7.

[0101] Table 7 Results of the anti-TSWV activity of compounds 1-16

[0102]

[0103]

[0104] a Average value of three times; b Ningnanmycin was used as a positive control.

[0105] The in vivo anti-TSWV test showed that compound 12 (100 μg / mL) exhibited a strong preventive effect in the treatment of first applying the drug and then inoculating the virus, with an inhibition rate of 78.0%, which was 65.0% higher than the positive control Ningnanmycin.

[0106] Experimental Example 2:

[0107] The insecticidal activity of the above-mentioned matrine alkaloids was studied and verified as follows:

[0108] Test target: Western flower thrips (Frankliniella occidentalis) is a sensitive strain that has been raised indoors for many years. Before the experiment, pseudopupae of western flower thrips were collected, and after they emerged, adults that emerged on the same day were selected for this test.

[0109] Culture conditions: The culture conditions for the test targets and post-test targets are temperature 27±1°C, relative humidity 70±5%, and photoperiod 16 / 8h (L / D).

[0110] Instruments and equipment: electronic balance (sensitivity of one ten-thousandth), spray tower, 100ml beaker, glass test tube, measuring cylinder, culture dish, Para film, sponge, filter paper, pipette, tweezers, brush, microscope, etc.

[0111] Test reagents: The content of new compounds is 100%.

[0112] Test concentration: 200mg / L.

[0113] Preparation of medicine: original medicine: weigh the required amount using a 1 / 10,000 electronic balance; solvent: N,N-dimethylformamide (DMF), 0.2%; emulsifier: Tween 80, 0.2%; add clean water to dilute to the required concentration.

[0114] Test method:

[0115] The test compound was dissolved in N,N-dimethylformamide and diluted to the required concentration with distilled water containing TW-80 (2mL / L), and 4 replicates were set. A filter paper with a diameter of 180mm was placed at the bottom of a 200mm diameter petri dish. Fresh scallion segments were cut into 120mm lengths, placed in different concentrations of drug solution and soaked for 30s, then taken out and dried under natural conditions, and placed in the prepared petri dishes, 5 roots per dish, and 50 healthy and uniform-sized adult thrips were selected and placed in the petri dishes, which were sealed with plastic wrap and pierced with 20 holes to keep the petri dishes ventilated. The petri dishes were placed in the above-mentioned artificial climate box for breeding. The positive control was ethyl spinetoram, and the negative control was a solution (CK) of equal amounts of N,N-dimethylformamide and TW-80 concentrations.

[0116] Methodology:

[0117] After 48 h, the survival of Frankliniella occidentalis adults treated with insecticides at various concentrations was observed and recorded. When observing, the insect body was gently touched with a fine brush, and if it did not move twice, it was recorded as the death of Frankliniella occidentalis. The recording results are shown in Table 8.

[0118] Table 8 Insecticidal effects of some compounds on Frankliniella occidentalis

[0119]

[0120] The results in Table 8 above showed that Compounds 4, 10 and 12 showed good activities against Frankliniella occidentalis at the tested concentrations.

[0121] Experimental Example 3:

[0122] The insecticidal activities of Compounds 4, 10 and 12 against Frankliniella occidentalis were further evaluated. The specific method was as follows:

[0123] Six concentrations of Compounds 4, 10 and 12, namely 200 mg / L, 100 mg / L, 50 mg / L, 25 mg / L, 12.5 mg / L and 6.25 mg / L, were set respectively, and the anti-aphid activity was studied by the method of Experimental Example 2.

[0124] Data statistical analysis: All experimental data were analyzed using DPS V 7.0 statistical software. The mortality rate was calculated based on the experimental data, and the virulence regression equation, correlation coefficient (r), LC 50 、LC 90 and its 95% confidence limits were obtained, as shown in Table 9.

[0125] Table 9 Activity test results of Compounds 4, 10 and 12 against Frankliniella occidentalis 50 Activity test results of Compounds 4, 10 and 12 against Frankliniella occidentalis

[0126]

[0127] It can be seen from Table 9 that Compounds 4, 10 and 12 have high insecticidal activities.

Claims

1. Matrine alkaloids, characterized in that: the matrine alkaloids are selected from any one of Compounds 1 to 14 with the following chemical structures:

2. The preparation method of the matrine alkaloids according to Claim 1, characterized in that: it comprises the following steps: (1) Take Sophora tonkinensis root and rhizome, reflux and extract with methanol, combine the extraction solutions and concentrate to obtain an extract, and subject the extract to acid-base treatment to obtain total alkaloids; the acid-base treatment is specifically as follows: first adjust the pH value to 1-3 with 0.1% hydrochloric acid, then extract with petroleum ether to remove non-alkaloid components, adjust the pH of the acid aqueous layer to 9-11 with 25% - 28% concentrated ammonia water, and then extract the alkaline aqueous layer with dichloromethane and n-butanol in sequence; (2) Subject the total alkaloids to silica gel column chromatography with 300 - 400 mesh, and perform gradient elution with dichloromethane:methanol with a volume ratio of 100:0 - 0:100 as the mobile phase to obtain Fraction A, Fraction B, Fraction C, Fraction D and Fraction E; Fraction A is eluted with dichloromethane:methanol with a volume ratio of 50:1, Fraction B is eluted with dichloromethane:methanol with a volume ratio of 20:1, Fraction C is eluted with dichloromethane:methanol with a volume ratio of 10:1, Fraction D is eluted with dichloromethane:methanol with a volume ratio of 5:1, and Fraction E is eluted with dichloromethane:methanol with a volume ratio of 2:1; (3) Subject Fraction D to MCI column chromatography separation, and perform gradient elution with methanol:water with a volume ratio of 0:100 - 100:0 as the mobile phase to obtain D-1, D-2, D-3 and D-4; D-1 is eluted with methanol:water with a volume ratio of 20:80, D-2 is eluted with methanol:water with a volume ratio of 40:60, D-3 is eluted with methanol:water with a volume ratio of 60:40, and D-4 is eluted with methanol:water with a volume ratio of 80:20; (4) Subject D-1 to silica gel column chromatography with 300 - 400 mesh, and perform gradient elution with petroleum ether:acetone:diethylamine with a volume ratio of 100:1:0.125 - 1:1:0.005 as the mobile phase to obtain Compound 14 and Compound 12; Compound 14 is eluted with petroleum ether:acetone:diethylamine with a volume ratio of 20:1:0.005, and Compound 12 is eluted with petroleum ether:acetone:diethylamine with a volume ratio of 10:1:0.005; (5) Subject D-2 to methanol gel purification and then to semi-preparative HPLC purification, and perform elution with methanol:water with a volume ratio of 21:79 as the mobile phase to obtain Compound 6, Compound 7 and Compound 1; (6) Subject D-3 to silica gel column chromatography with 300 - 400 mesh, and perform elution with dichloromethane:methanol with a volume ratio of 50:1 as the mobile phase, and then subject it to methanol gel purification to obtain Compound 8, Compound 9, Compound 4 and Compound 5; (7) D-4 was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was performed with petroleum ether:acetone:diethylamine at a volume ratio of 50:1:0.125 - 1:1:0.005 as the mobile phase to obtain compound 10; the compound 10 was eluted with petroleum ether:acetone:diethylamine at a volume ratio of 10:1:0.005; (8) Fraction E was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was performed with dichloromethane:methanol at a volume ratio of 20:1 - 0:100 as the mobile phase to obtain E-1, E-2, E-3, E-4, and E-5; E-1 was eluted with dichloromethane:methanol at a volume ratio of 15:1, E-2 was eluted with dichloromethane:methanol at a volume ratio of 10:1, E-3 was eluted with dichloromethane:methanol at a volume ratio of 5:1, E-4 was eluted with dichloromethane:methanol at a volume ratio of 2:1, and E-5 was eluted with dichloromethane:methanol at a volume ratio of 1:1; (9) E-1 was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was performed with petroleum ether:acetone:diethylamine at a volume ratio of 25:1:0.005 - 1:1:0.005 as the mobile phase, followed by separation on a methanol gel and purification by semi-preparative HPLC. Finally, elution was performed with acetonitrile:water at a volume ratio of 17:83 as the mobile phase to obtain compound 11; (10) E-4 was subjected to silica gel column chromatography (300 - 400 mesh), and gradient elution was performed with dichloromethane:methanol at a volume ratio of 25:1 as the mobile phase, followed by purification by semi-preparative HPLC and elution with methanol:water at a volume ratio of 34:66 as the mobile phase to obtain compound 2 and compound 3; (11) E-5 was purified by a methanol gel and then by semi-preparative HPLC, and elution was performed with methanol:water at a volume ratio of 36:64 as the mobile phase to obtain compound 13.

3. A pharmaceutical composition comprising the matrine alkaloid or its salt according to claim 1 and a pesticidally acceptable carrier.

4. Use of the matrine alkaloid according to claim 1 or the pharmaceutical composition according to claim 3 in the preparation of a pesticide against plant virus TSWV.

5. Use of compounds 2, 3, 4, 6, 7, 10, 11, 12, 13, 14 in the matrine alkaloid according to claim 1 in the preparation of an insecticide or use of the pharmaceutical composition according to claim 3 containing compounds 2, 3, 4, 6, 7, 10, 11, 12, 13, 14 in the preparation of an insecticide.

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