Application of beta-costus acid in preparation of medicine for resisting plant pathogenic fungi

Abstract

The invention discloses application of beta-costus acid in preparation of a medicine for resisting plant pathogenic fungi, and belongs to the technical field of new application of natural compounds. The beta-costus acid disclosed by the invention can be used for inhibiting phytophthora nicotianae, fusarium oxysporum, alternaria alternata, colletotrichum gloeosporioides, colletotrichum caffeophyllum and botrytis cinerea, and shows a very good inhibition effect on the mycelial growth of the phytophthora nicotianae, the fusarium oxysporum and the colletotrichum gloeosporioides, and the bacteriostasis rates respectively reach 94.64%, 72.55% and 70.46%. The beta-costus acid has the effect of resisting the plant pathogenic fungi and can be used as a new choice of the medicine for resisting the plant pathogenic fungi.

Description

Technical Field

[0001] The invention belongs to the technical field of new applications of natural compounds, and specifically relates to the application of beta-costus acid in the preparation of anti-plant pathogenic fungi drugs. Background Art

[0002] Laggera crispata (Vahl) Hepper & J.RI Wood is a perennial herbaceous plant of the genus Laggera of the Asteraceae family. According to the "Southern Yunnan Materia Medica", Laggera crispata can treat wind-heat toxicity, painful acne, scars, warts, epilepsy, and blood-wind ringworm. The whole plant of Laggera crispata is used as medicine, with a spicy taste, slightly warm nature, and cold nature. It has the effects of detoxification, swelling, blood circulation, cancer removal and stagnation. It is mainly used to treat upper respiratory tract infections, tonsillitis, pharyngitis, mumps, stomatitis, tracheitis, carbuncles, boils, burns, scalds, snake bites and trauma. According to modern pharmacological studies, Laggera crispata has expectorant and anti-tumor effects; modern clinical use is used to treat glanditis, upper respiratory tract infections, tonsillitis, stomatitis, bronchitis and painful swellings.

[0003] Its active ingredient, β-costus acid, has been disclosed to have in vitro acaricidal activity and can be used to treat Varroa mite disease in bee colonies. However, there are no reports that β-costus acid or Hexaphyllum pterophyllum extract has anti-plant pathogenic fungal activity. Summary of the invention

[0004] The purpose of the present invention is to provide the application of beta-costus acid in the preparation of anti-plant pathogenic fungi drugs.

[0005] To achieve the above object, the present invention provides the following technical solutions:

[0006] One of the technical solutions of the present invention: application of β-costus acid in the preparation of anti-plant pathogenic fungi drugs.

[0007] The second technical solution of the present invention: application of β-costus acid in the preparation of drugs against tobacco blight.

[0008] The third technical solution of the present invention: the use of β-costus acid in the preparation of drugs against tobacco oxysporum Fusarium.

[0009] Technical solution four of the present invention: application of β-costus acid in the preparation of drugs against citrus gloeosporioides anthracnose.

[0010] The structural formula of β-costus acid is:

[0011]

[0012] The beneficial technical effects of the present invention are as follows:

[0013] The invention discloses that beta-costus acid can inhibit tobacco phytophthora, tobacco oxysporum Fusarium, tobacco red spot pathogen, citrus gloeosporium Colletotrichum, coffee fruit anthracnose and tomato gray mold, and shows good inhibitory effect on the hyphae growth of tobacco phytophthora, tobacco oxysporum Fusarium and citrus gloeosporium Colletotrichum, with the inhibition rates reaching 94.64%, 72.55% and 70.46% respectively. It shows that beta-costus acid has the function of resisting plant pathogenic fungi and can be used as a new choice of anti-plant pathogenic fungi drug. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 This is a graph showing the inhibitory effects of different concentrations of β-costus acid on tobacco Phytophthora, tobacco oxysporum and citrus gloeosporium. DETAILED DESCRIPTION

[0015] Now, various exemplary embodiments of the present invention are described in detail, and this detailed description should not be considered as a limitation of the present invention, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present invention. It should be understood that the terms described in the present invention are only for describing specific embodiments and are not used to limit the present invention.

[0016] In addition, for the numerical range in the present invention, it is understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0017] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the invention pertains. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention.

[0018] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0019] Example 1

[0020] Extraction of β-costus acid:

[0021] The wild-collected Hexaphyllum pterocarpa was dried and crushed (5 kg), then soaked in methanol reagent, extracted three times at room temperature, and finally concentrated under reduced pressure to obtain 8 L of crude extract. The crude extract was extracted with equal amounts of petroleum ether and ethyl acetate (each used 3 times) to obtain a petroleum ether layer (68 g) and an ethyl acetate layer (55 g). The ethyl acetate layer was then subjected to silica gel column chromatography using a silica gel column (10×100 cm), eluted with petroleum ether-acetone (100:1-1:1), and each sample obtained by the gradient was subjected to thin layer chromatography and then heated with sulfuric acid for color development (some samples had a darker tailing phenomenon during color development, making it difficult to distinguish between different samples, which could be improved by adding 0.1% formic acid to the developing agent). The same components were combined to obtain 8 parts (numbered as Fr.1-Fr.8 in the order of elution).

[0022] The Fr.4 (45.6 g) fraction was first purified by MCI column chromatography with a mobile phase of 10%-90% methanol-water gradient elution. When the sample flushed out under the condition of 10% methanol-water became significantly less or no sample was flushed out, the mobile phase was replaced with 20% methanol-water. The same was true for the rest of the process. Each time the mobile phase concentration was changed, 10% methanol was added to finally obtain 8 parts (4A-4H). Fraction 4D (10.5 g) was obtained when the ratio of methanol to water was 50%. The main thin layer chromatography spot plate was heated with persulfate and the color turned dark blue (the developing agent was petroleum ether: ethyl acetate = 5:1 and the display agent was 10% concentrated sulfuric acid). Fraction 4D was further eluted by normal silica gel column chromatography (the mobile phase was petroleum ether-ethyl acetate = 40:1-4:1) gradient elution to obtain 5 fractions (4D1-4D5). Fraction Fr.4D2 (3.9 g) was obtained when the ratio of petroleum ether: ethyl acetate was 10:1. This part continued to be eluted by Sephadex LH-20 gel column chromatography (the eluent was a mixed solution of methanol-dichloromethane = 6:4) to remove certain impurities and pigments. The purified sample was then purified by HPLC (liquid phase conditions: the chromatographic column was YMC-pack ODS-A (5 μm, 10×250 mm), mobile phase was 58% acetonitrile water, flow rate was 3 mL / min, tR=59 min), and finally the target product was obtained.

[0023] Structure confirmation:

[0024] Example 1 The target product obtained by 13 C-NMR and DEPT detection showed that the compound had 15 carbon signals, including 1 methyl, 8 methylene, 2 methine and 4 quaternary carbons. Combined with ESI-MS, its molecular formula was determined to be C 15 H 22 O2, molecular weight 234 (m / z: 233 [MH] - ).

[0025] The NMR data are as follows: 1 H-NMR(500MHz, CD3Cl3)δ:2.3(1H,m,H-1),2.01(1H,m,H-1),1.58(1H,m,H-2),1.52(1H,m,H-2),1.5 9(1H,m,H-3),1.34(1H,m,H-3),1.89(1H,d,J=11.3Hz,H-5),1.66(1H,m,H-6),1.22(1H,m,H-6),2.5 3(1H,m,H-7),1.61(1H,m,H-8),1.46(1H,m,H-8),1.59(1H,m,H-9),1.28(1H,m,H-9),6.31(1H,s,H- 13),5.68(1H,s,H-13),0.55(3H,s,H-14),4.40(1H,d,J=1.5Hz,H-15),4.70(1H,d,J=1.5Hz,H-15). 13 C NMR (150MHz, CDC l3 ,)δ:36.8(C-1),23.4(C-2),41.8(C-3),150.7(C-4),49.8(C-5),29.9(C-6),39.3(C-7),27.3(C- 8),41.0(C-9),36.8(C-10),145.3(C-11),172.5(C-12),124.8(C-13),16.4(C-14),105.5(C-15).

[0026] After analysis, the product extracted in Example 1 was β-costus acid.

[0027] The antibacterial activity of the β-costus acid extracted in Example 1 was evaluated, and the results are shown in Table 1. As shown in Table 1, at a mass concentration of 100 μg / mL, β-costus acid has an inhibitory effect on the mycelial growth of six tested plant pathogens, including tobacco Phytophthora, tobacco oxysporum Fusarium, tobacco brown spot pathogen, citrus gloeosporium Colletotrichum, coffee berry anthracnose and tomato gray mold. It shows a good inhibitory effect on the mycelial growth of tobacco Phytophthora, tobacco oxysporum Fusarium and citrus gloeosporium Colletotrichum, with the inhibition rates reaching 94.64%, 72.55% and 70.46%, respectively.

[0028] Table 1 Inhibition rate of β-costus acid on six plant pathogenic fungi (%)

[0029]

[0030] Determination of the toxicity equation of β-costus acid against plant pathogenic fungi

[0031] The toxicity equation of tobacco phytophthora, tobacco oxysporum and citrus gloeosporium, with an inhibition rate of β-costus acid of more than 50%, was determined. The inhibitory effect of each concentration of β-costus acid is shown in Figure 1 , the calculation results are shown in Table 2.

[0032] Table 2 Inhibitory activity of β-costus acid on the mycelial growth of three plant pathogens

[0033] Plant pathogenic fungi Virulence regression equation <![CDATA[EC 50 (μg / mL)]]> <![CDATA[Correlation coefficient (R 2 )]]> Phytophthora nicotianae y=1.713x+3.1137 12.56 0.979 Fusarium oxysporum y=1.3822x+2.6427 51.29 0.9277 Colletotrichum citri y=1.3297x+2.7718 47.86 0.9581

[0034] As can be seen from Table 2, the EC values ​​of β-costus acid against Phytophthora nicotianae, Fusarium oxysporum and Colletotrichum citri gloeosporioides are 50 The values ​​were 12.56, 51.29 and 47.86 μg / mL respectively.

[0035] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. Application of β-costus acid in the preparation of drugs against plant pathogenic fungi.

2. Application of β-costus acid in the preparation of drugs against tobacco blight.

3. Application of β-costus acid in the preparation of drugs against tobacco oxysporum fungus.

4. Application of β-costus acid in the preparation of drugs against citrus gloeosporioides anthracnose.

Images