KR20070065930A - Anti-pinewood nematode compound containing cinnamaldehyde extracted from cinnamomum cassia and its structurally-similar compounds - Google Patents

Abstract

An anti-pinewood nematode composition comprising cinnamaldehyde is provided to show strong nematicidal effect against pine tree nematode by treating the composition mainly comprising the cinnamaldehyde and its' structurally similar compounds of methyl-trans-cinnamaldehyde and 2-methoxy-cinnamaldehyde, thereby significantly decreasing the individuals of the pine tree nematode. The anti-pinewood nematode composition comprises cinnamaldehyde extracted from Cinnamomum cassia, which is prepared by pulverizing Cinnamomum cassia, adding 5 times weight of 70% hydrogenated alcohol regarding the pulverized Cinnamomum cassia thereto and extracting it at a temperature of 4-40 deg.C for 12 hours. In the composition, the concentration of the cinnamaldehyde is 0.1-50 vol.%.

Description

Anti-pinewood nematode compound containing cinnamaldehyde extracted from Cinnamomum cassia and its structurally-similar compounds.

FIG. 1 shows the chemical structure (C ₆ H ₅ CH = CHCHO (C ₉ H ₈ O), molecular weight: 132.16) of the purified aldehyde isolated from cinnamon.

2 shows the mass spectrometry spectra of cinnamic aldehydes isolated and purified from cinnamon.

3 shows the chemical structure of -Methyl- trans- cinnamaldehyde (formula: C ₆ H ₅ CH = C (CH ₃ ) CHO, molecular weight: 146.19).

Figure 4 shows the chemical structure of 2-Methoxycinnamaldehyde (structure: CH ₃ OC ₆ H ₄ CH = CHCHO, molecular weight: 162.19).

There are many types of pathogens and pests in the soil. Among them, phytoparasites are known to directly and indirectly affect the growth and development of plants. Among them, pine re-nematodes, which are infected with pine, a representative species of Korean forests, cause pine death in a short time, are mediated by the pine salted sky, have a very rapid spreading rate, and have no apparent preventive or therapeutic drugs. It is known as one of the most important harmful nematodes along with root-knot nematodes, which are parasitic and inhibit the growth of crops. Pine re-nematodes are taxonomy belonging to leaf nematodes and pine re-nematodes, and 49 species (Proc. Helminthol. Soc. Wahs. 55 (1), pp 1-11, 1988) have been reported worldwide. South Korea currently has Bursaphelenchus xylophilus (Bursaphelenchus xylophilus) and similar jaeseonchung (Bursaphelenchus mucronatus ) only two species have been reported (intuitive pine nematode, National Forest Research Institute, 2005). Among them, pine re-nematodes are infected in the body of the sea salt, and then infested with 1-2 year old pine shoots.They grow inside the pine to block the catheter of host plants, block the water rise, and secrete cellulase. It is known to kill host plants in a short time of about 90 days. The pine re-nematode disease was first discovered in Japan in 1905, but it was not until 1972 that the causative pest proved to be a pine re-nematode, and now it is known that most pine forests in Japan died. In China, it was gradually spread after the outbreak was confirmed in Nanjing in 1982. In Taiwan, it was known to have been invaded in 1985 and suffered almost complete annihilation. In Korea, the damage caused by pine nematode disease, which is estimated to be caused by infected wood, has been increasing rapidly since it was confirmed at Busan Geumjeongsan in 1988. As of September 2005, the area has reached 22,525 ha, 49 municipalities in the country. It has also been reported that about 1 million pines have been harvested due to their damages to date (Korea Forest Service, National Forest Research Institute, and Southern Forest Service, 2005).

As a preventive method of pine nematode disease, there are methods such as aviation and ground drug spraying, and water injection, etc. As a control method, a method of controlling the burning of dead wood, burning, pulverizing and fumigating is used. However, the drug dispersing method using mep emulsion has the advantage of being applicable to a large area of forest as a pesticide for the saltwater saltwater, which is a medium, but has a high possibility of causing a wide range of environmental pollution. On the other hand, there is no drug currently developed in Korea, which is a method of directly preventing pine re-nematode, and it is known that a drug such as Abamectin is being tested (pine re- nematode control workshop, Southern Forest Service, 2005). In the case of Levamis, Mesulfenfos and Morantel tartrate, three types are available on the market, but due to their high price, they are limited to the ornamental water. In addition, the currently used control method is a combination of physical and chemical methods, as mentioned above, but the disadvantages that concern about the side effects due to wildfire, thermal damage, high control failure potential and the toxicity of Metam sodium used for fumigation (Intuitive pine worm nematode, National Forest Research Institute, 2005).

On the other hand, the market size of the world's pesticides in recent years has been gradually decreasing except Eastern Europe, with USD 25.2 billion in 1992, USD 30.7 billion in 1998, USD 30 billion in 1999, USD 22.9 billion in 2000, and USD 279.4 billion in 2001. , Woodmackenzie, 2002). Similarly, the domestic pesticide market continued to decline to 1.3 trillion won in 2001, 930 billion won in 2002, 9270 billion won in 2003, and 920 billion won in 2004. In 2005, the use of chemical synthetic pesticides in the control of pine nematodes, along with the possibility of adverse effects on the environment and human beings, combined with the spread of negative perceptions on the use of chemical pesticides and the government's control of chemical pesticide use in the future, It is expected to remain constant.

On the other hand, in Korea, products with natural nematode suppressing effects are partially sold in the agricultural sector for the purpose of reducing root-knot density, but most of them are imported from surfactants, nematode premature fungi and India, China, etc. Oil (main ingredient: Azadirachtin) or Pyrethrins (main ingredient: Pyrethrins) is used as a raw material, and it is difficult to determine the exact amount of use due to the uncertainty of distribution and sales volume, and the effect of reducing the root gland nematode density by soil irrigation Compared to the chemical pesticides that have been used is known to fall significantly. Moreover, it is understood that the present application or control drug product has not been developed so far to control pine nematode disease by using the active ingredient derived from natural products.

Currently, there are two types of pine reef nematode control agents that directly control pineal nematode or pine re nematode, but all of them are chemical synthetic pesticides. Among them, three kinds of Levamis, Mesulfenfos, and Morantel tartrate which are commercially available in Japan are excellent. On the other hand, due to the high cost along with the possibility of adverse effects on the human body and other ecosystems, it is being restricted to use in pine re-nematode disease site. In addition, most of the pine re-nematode control drugs currently in use are imported drugs. As mentioned above, drugs such as Abamectin are being tested in Korea, but it may take some time for commercialization. Meanwhile, in the first half of 2005, pine re-nematode control drugs mainly composed of organophosphorus fosthiazate were marketed in Korea, but it was originally found to control root-knot nematodes parasitic in agricultural horticultural crops, and it has been confirmed that it has side effects on silkworms. The effect has not yet been proven in the field of re-nematode disease.

In the present invention, to find out the active ingredient by searching for a new natural product having a strong nematicidal activity among the natural products widely used as edible and herbal medicine, and to enable it to be applied as a natural pesticide having an effective insecticide against pine re- nematode. To this end, Cinnamic aldehyde, a known ingredient, was isolated and purified from cinnamon, which was found to be excellent in nematicidal effect among 150 kinds of herbal medicines that were primarily searched, and confirmed that it was an active ingredient exhibiting activity against pine re-nematodes.

The technical problem to be achieved by the present invention is the rise of -Methyl- trans- cinnamaldehyde and 2-Methoxycinna-maldehyde, which are structurally similar and commercially available compounds with strong nematicidal activity of cinnamic aldehyde isolated and purified from cinnamon. The effect is to provide a control agent with nematicidal activity against pine re-nematode, which is currently rapidly spreading nationwide.

The present invention is a pine re-nematode control agent containing -Methyl- trans- cinnamaldehyde (Aldrich) and 2-Methoxycinnamaldehyde (Aldrich), which are structurally similar and commercially available compounds with purified aldehyde isolated from cinnamon. More specifically, the main component is a cinnamic aldehyde obtained from an extract made by drying the bark of cinnamon belonging to the camphor family, finely pulverized, and adding an organic solvent. The present invention relates to a control agent which exhibits a strong insecticidal effect against pine re-nematode by adding -Methyl- trans- cinnamaldehyde (98%, Aldrich) and 2-Methoxycinnamaldehyde (96%, Aldrich).

Cinamic aldehyde has been confirmed by the presence of Morris, etc. from cinnamon (J. Assoc. Off. Anal. Chem., (56) 1037, 1973), irrelevant to the present invention, the cinnamic aldehyde of the present invention Is a vegetable oil obtained from cinnamon with the chemical formula C ₆ H ₅ CH = CHCHO (C ₉ H ₈ O), and a patent related to pine re-nematode control agent using the same has not been reported yet. In the present invention, in addition to the cinnamic aldehyde separated and purified from cinnamon, the same substance obtained by chemical synthesis can be used.

In the present invention, in order to search for a natural-derived compound having an excellent pine re-nematode insecticidal effect, various kinds of plant extracts were treated to determine the extent to which the pine re-nematode density was reduced to search for nematicidal activity. As a result, the extract of cinnamon was found to be superior to the nematicidal activity against pine re-nematode compared to other natural extracts, it was confirmed that the cinnamic aldehyde is an active ingredient.

Example 1

The manufacturing method of the cinnamic aldehyde from the cinnamon performed by this invention is as follows.

1 kg of Cinnamomum cassia was pulverized, and 5% of 70% hydrous alcohol was added, followed by extraction at 4-40 ° C. for 12 hours, followed by filtration using Whatman 42 filter paper, and the filtrate was dried at 65 ° C. using a rotary vacuum evaporator. To give 115 g of dry weight.

It was suspended again in purified water and the suspension was partitioned with hexane. This fraction was separated and purified by column chromatography and silica gel chromatography using silica gel to obtain the target compound, cinnamic aldehyde. However, the method for obtaining the target compound is not limited to the above purification method.

IR, mass spectrometry and boiling point measurement results of the cinnamic aldehyde, the target compound obtained by the above purification method, are as follows.

1. IR Spectrum measurement result

Carbonyl streching  frequency C = O: 1679 ^cm-1

Aromatic C = C streching  frequency: 1495 ^cm-1

Aromatic C = C-H and olefin C-H sp2  Carbon

                H peak streching  frequency: 3028 ^cm-1 , 3061 ^cm-1

Aromatic out of plane bending: 751 ^cm-1

Aldehyde  - CHO C-H straching  frequency: 2815 ^cm-1

2. Mass Spectrum Measurement Results (see FIG. 2)

M / S (m / z): 132 (M ⁺ , molecular ion), 131 (100, M-H), 115 (2),

           103 (39, M- CHO ), 77 (28, phenyl group),

           63 (5), 53 (1), 51 (23), 43 (6)

As a result of mass spectrometry, the molecular weight peak was 132, which is a typical peak that shows that the aldehyde-containing compound easily loses hydrogen atoms, and the peak of 103 (39, M-CHO) and 77 (28, phenyl group). It was confirmed that this material is a cinnamic aldehyde.

3. Boiling Point Measurement Results

Measured value: 100 ℃ / 10mmHg, 248 ℃ / 760mmHg

It was also found that the boiling point was within the range of known aldehydes.

Experimental Example 1: Insecticidal effect of cinnamic aldehyde and structurally similar compounds -Methyl- trans- cinnamaldehyde and 2-Methoxycinnamaldehyde against pine re-nematode

To verify the effectiveness of pine re-nematode insecticides by treatment with cinnamic aldehyde isolated from cinnamon and structurally similar -Methyl- trans- cinnamaldehyde (98%, Aldrich) and 2-Methoxycinnamaldehyde (96%, Aldrich) Drug efficacy experiments were conducted. The Bursaphelenchus xylophilus used for testing was used in the experiment was collected from the pine fragment Bursaphelenchus xylophilus suspicious neck Gyeongsangbuk Gumi Seonsan seupryeri one hill, isolated from cinnamon, the purified aldehyde, and dynamic scenarios -Methyl- trans -cinnamaldehyde, 2-Methoxycinnamaldehyde Was diluted with ethanol and treated three times on each concentration in 24-Well plate and the insecticidal rate was calculated by analyzing the average nematode density after 24 hours under 25 ℃ and shading conditions. The pine nematode was removed from the suspicion of infection, and the exposed sapwoods were collected and separated by Baermann's funneling technique. The numbers were determined by Leica S6E anatomical microscope.

Insecticidal Effect of Soil Root-knot Nematodes on Cinnamic Aldehyde and -Methyl- trans- cinnamaldehyde and 2-Methoxycinnamaldehyde Isolated from Cinnamon Treatment Insecticide Rate (%) No treatment 6.28 CA, 0.005% (50 ppm) 4.55 CA, 0.01% (100 ppm) 26.67 CA, 0.02% (200 ppm) 87.53 CA, 0.03% (300 ppm) 100.00 CA, 0.05% (500 ppm) 100.00 CA, 0.1% (1,000 ppm) 100.00 CA: -MC = 1: 1, 0.005% (50 ppm) 9.52 CA: -MC = 1: 1, 0.01% (100 ppm) 22.22 CA: -MC = 1: 1, 0.02% (200 ppm) 77.25 CA: -MC = 1: 1, 0.03% (300 ppm) 100.00 CA: -MC = 1: 1, 0.05% (500 ppm) 100.00 CA: -MC = 1: 1, 0.1% (1,000 ppm) 100.00 CA: 2-MC = 1: 1, 0.005% (50 ppm) 6.25 CA: 2-MC = 1: 1, 0.01% (100 ppm) 20.00 CA: 2-MC = 1: 1, 0.02% (200 ppm) 85.71 CA: 2-MC = 1: 1, 0.03% (300 ppm) 100.00 CA: 2-MC = 1: 1, 0.05% (500 ppm) 100.00 CA: 2-MC = 1: 1, 0.1% (1,000 ppm) 100.00 CA: -MC: 2-MC = 1: 1: 1, 0.005% (50 ppm) 11.11 CA: -MC: 2-MC = 1: 1: 1, 0.01% (100 ppm) 37.36 CA: -MC: 2-MC = 1: 1: 1, 0.02% (200 ppm) 97.65 CA: -MC: 2-MC = 1: 1: 1, 0.03% (300 ppm) 100.00 CA: -MC: 2-MC = 1: 1: 1, 0.05% (500 ppm) 100.00 CA: -MC: 2-MC = 1: 1: 1, 0.1% (1,000 ppm) 100.00

* CA, cinnamic aldehyde; -MC, -Methyl- trans -cinnamaldehyde;

  2-MC, 2-Methoxycinnamaldehyde

* Insect killing rate (%) = 1-((nematode after treatment / nematode before treatment) × 100)

As a result of the experiment, the pine insecticidal insecticidal effect according to the concentration of -Methyl- trans- cinnamaldehyde and 2-Methoxycinnamaldehyde, which were isolated and purified from cinnamon, was shown in Table 1. Treatment with 0.02% alone showed a significant insecticidal effect of 77% -97%, and treatment with 0.03% showed a strong insecticidal effect of 100% in both treatments alone and mixed. In particular, the strongest nematicidal effect was found in the experimental group treated with the same amount of 1: 1, 1: Methyl aldehyde, -Methyl- trans- cinnamaldehyde and 2-Methoxycinnamaldehyde at a concentration of 0.02%. It was shown to be most effective for the control of pine re-nematodes.

 Example 2

Table 2 shows the composition of pineal worm nematode control agent mainly containing cinnamic aldehyde, purified from cinnamon, -Methyl- trans- cinnamaldehyde and 2-Methoxycinnamaldehyde.

ingredient Content (Unit: Volume%) Example 1 -Methyl- trans- cinnamaldehyde (98%, Aldrich) 2-Methoxycinnamaldehyde (96%, Aldrich) ethanol 30 30 30 10 Sum 100

Experimental Example 2: Pine re-nematode control effect of pine re-nematode control agent mainly composed of cinnamic aldehyde, -Methyl- trans -cinnamaldehyde and 2-Methoxycinnamaldehyde isolated and purified from cinnamon

The purpose of this study was to examine the control effect of pine re-nematode control agent on the site of pine worm nematode control agent consisting mainly of cinnamic aldehyde, -Methyl- trans- cinnamaldehyde and 2-Methoxycinnamaldehyde, purified from cinnamon. Field experiments were conducted on pine re-nematode infections. Re-neutral nematode infection and population determination of the pine was used in the same manner as in Experiment 1. The control test using the pine re-nematode control agent of Example 2 was carried out using intra-water injection of the stock solution by perforation. First, several holes were drilled about 10 cm deep at equal intervals in a 1.5 m above ground region of a pine tree having a diameter of 20 ± 2 cm and a diameter of 20 ± 2 cm with a perforator, and a control agent of Example 2 was injected with a certain amount and then sealed with silicon. Yield per treatment was 10 and 3 repetitions, and the change of the number of re-nematogens in pine was taken 30 grams of saplings in the treatment and untreated areas in the same manner as in Experiment 1 before and 4 weeks after treatment. Nematodes were isolated and speculatively determined using. The experimental results are shown in Table 3.

Control effect of pine re-nematode of Example 2 by treatment on pine re-nematode infected tree Treatment Average Nematode Density (30g) Average Nematode Density (30g) Control is (%) No treatment 8,215 10,766 0% Example 2 50 ml 5,542 4,167 24.81% Example 2 100 ml 12,036 1,621 86.53% Example 2 200 ml 7,763 889 88.54%

* Controlled value (%) = 1-((Nematode number before treatment / nematode number after treatment) × 100)

As a result, it was confirmed that 4 weeks after treatment with pine re-nematode control agent of Example 2, the number of re-nematodes in pine was significantly reduced compared to no treatment, and more than 100 ml per week in 20 ± 2 cm thorax diameter pine used in this experiment. It has been shown that re-nematode infected with pine tree can be effectively controlled by the control agent treatment. This result is judged to be a result of the pine re-nematode control effect is somewhat lower in the field test, which is a relatively bad condition than the indoor experiment conducted in Experimental Example 1 (control effect of more than 97% at 200 ppm treatment). However, the pine re-nematode control agent of Example 2 was far more than the control standard (Rural Development Administration notification 2005-3, April 25, 2005) which is more than 50% of the standard of biochemical pesticides against pine re-nematodes in the processing more than 100ml of the stock solution. It was shown that the high control value of more than 86% indicates that the pineal re-nematode control agent of Example 2, which is mainly composed of cinnamic aldehyde and -Methyl- trans- cinnamaldehyde, 2-Methoxycinnamaldehyde, isolated from cinnamon, can be used as a biochemical pesticide. It was.

Cinnamic aldehyde isolated and purified from cinnamon of the present invention showed a strong nematode effect against pine re-nematode in laboratory experiments, along with cinnamic aldehyde, -Methyl- trans -cinnamaldehyde and 2- The nematode control agent containing methoxycincinaldehyde as a main ingredient in pine reinfected nematode significantly reduced the population of pine re-nematodes and showed a clear control effect against pine re-nematodes, a direct cause of pine re-nematode disease.

Claims (4)

Cinnamomum cassia ( Cinnamomum cassia ) is a control agent for the purpose of treating and preventing pine re- nematodes containing filtrate after extraction for 12 hours at 4-40 ℃ by adding 70% hydrous alcohol of 5 times the weight. The method of claim 1, wherein the concentration of cinnamic aldehyde is contained in an amount of 0.1 to 50% by volume of the control agent for the purpose of treating and preventing pine re-nematode. The method of claim 1, wherein the concentration of -Methyl- trans -cinnamaldehyde is contained in an amount of 0.1 to 50% by volume of the control agent for the purpose of treating and preventing pine re- nematodes. The method of claim 1, wherein the concentration of 2-Methoxycinnamaldehyde is contained in an amount of 0.1 to 50% by volume of the control agent for the purpose of treating and preventing pine re- nematodes.

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