KR930000525B1 - Herbicide composition and process for it - Google Patents

Abstract

A plant growth inhibiting agent is prepd. by (a) fermenting hygromycin producing streptomyces olivovirdis KCTC 8426P in the medium contg. 10 g soluble starch, 20 G glucose, 25 g soybean meal, 1 g beef extract, 14 g yeast extract, 2 g NaCl and 0.5 g K2HPO4 (pH 7.3), (b) seperating hygromycin from the fermented broth and (c) mixing the obtd. hygromycin with additives to obtain the final product. The obtd. product contg. hygromycin is useful as herbicide or plant growth inhibiting agent.

Description

Herbicides and methods of preparing plant growth inhibitors and compositions thereof

The present invention relates to a herbicide composition produced by one round of soil actinomycetes, in particular, a new pollution-free herbicide and plant growth inhibitor composition having hygromycin (Hygro mycin) as an active ingredient, which has been previously used as a livestock repellent and feed additive. It is about.

Factors that influence agricultural productivity include a number of factors such as breeding improvement of crops, establishment of cultivation techniques and fertilizers, development of pesticides, and application to agricultural infrastructure. Among them, the development and use of pesticides, especially herbicides, which are used in terms of crop protection, are becoming more important as modernization of agriculture, large-scale farming and energy-saving cultivation methods are introduced. The average yield loss of each crop by actual weeds usually yields about 20-30% yield loss. If the loss of pests is included, a reduction of 30-45% is expected.

For this reason, much research has been focused on the search for herbicide active materials for herbicide development. To date, 571 kinds of herbicides have been developed and some of them have been put to practical use (Hodogaya Co. Ltd.). , Short review of herbicides, Zenkoku Nosan Kyoiku Kyokai Publishing Co. Ltd., 1986). However, most of the compounds reported above are organic chemical synthetic substances, which poses serious social problems such as pollution of the natural environment due to residual toxicity and safety problems to the human body.

The present invention is to propose a solution that can solve not only the pollution prevention of the natural environment as well as the above problems, but also the stability problems of the human body.

The inventors of the present invention have been searching for new substances that can be used as herbicides or plant growth regulators among pesticides that are essential for improving agricultural productivity, and hygromycin, which is used as an insect repellent or a feed additive for livestock, strongly increases the chloroplast formation of various plants. The present invention was found to be the first to discover a new and groundbreaking fact that it has the effect of inhibiting the growth of plants, depending on the concentration of the treatment, as well as the use of it to produce stable and pollution-free herbicides or plant growth regulators. It was completed.

Accordingly, an object of the present invention is to provide a new use of hygromycin, a known substance that has been produced from microorganisms and has been used as an insecticide or feed additive in livestock.

Hereinafter, the present invention will be described in detail.

The present invention relates to a herbicide composition comprising hygromycin as an active ingredient. The present invention also includes a plant growth inhibitor composition comprising the hygromycin as an active ingredient.

Hygromycin according to the present invention uses a product obtained by culturing soil actinomycetes, such as Streptomvces olivoviridis KCTC 842P, in a conventional actinomycetes batch.

Referring to the present invention in more detail as follows.

According to the present invention, isolated and confirmed Streptomyces oriboviridis KCTC 8426P, a kind of soil anthrax, obtained hygromycin as a culture product therefrom, and 10% of hygromycin as an active ingredient, and other supplements, interfaces Herbicide compositions or plant growth inhibitory compositions containing 90% of other ingredients such as active agents and weighting agents are prepared. In the present invention, in order to achieve the above object, after separating about 3,000 strains of actinomycetes from rice fields, fields, fields, and forest soils of Korea, the plant is being searched for potent herbicidally active substances or plant growth inhibitors from the culture medium. Streptomyces orivoviridis KCTC 8426P (Streptomy ces olivoviridis KCTC 8426P), a strain that produces substances that strongly inhibit chloroplast formation, was isolated and identified the morphological, cultural and physiological characteristics of the strains. Was separated and subjected to extraction purification. The characteristics of the strain used in the present invention are as follows.

1) Morphological characteristics

The Streptomyces oriboviridis KCTC 8426P (hereinafter referred to as "carrier strain") was incubated at 28 ° C for 14 days according to ISP (International Streptomyces Project) regulations. The spores showed good growth, and the shape of spores observed by electron microscopy was elliptical, and the surface showed a spikey shape and the size of spores was 1.0-1.2 × 0.5-0.6㎛. The chain of spores had more than 50 spores connected in a straight line.

2) Cultural characteristics according to various media conditions

Table 1 shows the culture characteristics of the strain after incubation at 28 ° C. for 14 days under various media conditions.

TABLE 1

Cultural characteristics according to various media preparations

The growth of this strain was generally good in various media, but the growth of strain and growth of mycelia were the most favorable in oatmeal agar media, and the color of the mycelia showed white with grayish white to red light. However, the production of water-soluble pigments was not recognized.

3) Physiological characteristics

In order to investigate the physiological characteristics of this strain, after incubation at 28 ° C. for 14 days using Schilling Gerbrid agar medium, the hydrolysis capacity, gelatin liquefaction capacity, peptonization of milk, and melanin pigment production were investigated. 2 is shown.

TABLE 2

Various physiological characteristics

Moreover, the result of having investigated the magnetization ability of the carbon source after culture | cultivating for 28 days and 28 degreeC using the sealing gerbrid agar medium is as Table 3, and. In other words, cellulose and rhamnos were not used at all, and salicin was not used at all, but L-arabinose, I-inositol, and mannitol were used.

TABLE 3

Sugar use characteristics

4) About fungicides

The cell wall of the strain Streptomyces oriboviridis 8426P strain was examined by Becker's method (B. Backer et. Al., Applied Micro-biology Vol. 12, pp. 421-423, 1964). Minofymeric acid (LL-diaminopimellic acid) and glycine (glycin) were detected and the cell wall was found to be type I (type I). In addition, a characteristic pattern was not recognized as a result of examining the sugar component of procytosis according to Lechevalier's method (Lechevalier, L.P., Journal of Laboratory and Clinical Medicine, Vol. 71, No. 934, 1968).

As a result of examining the physiological and biochemical characteristics of the strain of chloroplast-forming inhibitors, the strain was identified as the same strain as Streptomyces olivoviridis ATCC 15882. In addition, this strain was identified by ISP (International Streptomyces Project) standard, Burgy's Manual of Determination Bacteriology, 8th edition, The Actinomycetes volume 2, and recent literature on actinomycetes. .

In the present invention, the culture of the production strain was carried out in accordance with the general method of culturing actinomycetes, but in the case of liquid culture, it is preferable to shake shake culture or aeration stirring. As a medium component, media commonly used for actinomycetes were used, for example, glucose, fructose, mannitol, inositol, sucrose, starch, and soybean oil as carbon sources, and soybean meal, peptone, gravy, yeast and yeast extract as nitrogen sources. Inorganic salts such as corn steep liquor, corn steep sound, ammonium sulfate, and ammonium acetate are used as salts, phosphates, and calcium carbonates. Moreover, a trace amount metal salt may be added as needed. The acidity (pH) of the medium is preferably near neutral, and the temperature of the medium is 24-35 ° C., particularly around 28 ° C., and the production of the present chloroplast-forming inhibitor in the culture medium changes with time. When cultured with a triangular frying flask containing medium, the production of hygromycin used in the present invention reached the highest level when cultured for about 90-140 hours.

The hygromycin produced in this way was mainly present in the culture medium and easily eluted in butanol, and the RF value was weak when the methanol / chromoform = 1: 1 solution was used as a developing solvent in thin layer chromatography plate analysis using silica 60F plate. 0.56 and did not react with ninhydrin solution. In addition, the active material purified by thin layer chromatography was collected using a single peak section showing activity using high performance liquid chromatography to obtain a pure purified product. The UV spectrum of the purified active material thus obtained showed 270 nm at neutral pH and shoulders at 300 nm and 214 nm, respectively, and no change in spectrum at acidic pH, but at 285 nm, 340 nm and alkaline pH. Each shift occurred at 250 nm.

In addition, the hygromycin used in the present invention showed mainly antimicrobial activity against gram negative bacteria as shown in Table 4.

TABLE 4

Antibacterial spectrum of this substance

As a result, the antimicrobial spectrum and UV spectrum NMR spectrum results of the material produced by this strain were examined, and the results showed that the chloroplast formation inhibitor was the same substance as the known hygromycin (United States). Patent Office, Patent No. 3,100, 176, 1963). However, pore-formed hygromycin substances are used only as insect repellents or feed additives for poultry and livestock (Shiba Shoji, Bioactive Natural Substances, Section 480, OTC Publication, 1979), plant growth inhibitory or chloroplast formation. Documents reported as inhibitors were first discovered by the present invention in the absence of any conditions.

Previous reports on hygromycin indicate that Maria del Carmen and Juan Modolell, Eur. J. Biochem., 107, 409-414, 1980 and Minoru Yoshida (Minoru Yoshida et. Al., Agric. Biol. Chem., 50, 143-149, 1986), et al. Describe ribosomal peptidyl transferase and K88 antigen synthesis for hygromycin A. Only as a new inhibitor for K88 antigen synthesis has been reported, and J. Le boul and J. Davies, The J. Antibiotics, 35, 527-528, 1982) hygromycin Enzymatic modification by enzyme of B was described only, and production strain was also reported as Streptomyces hygroscopicus in the above literature. Meanwhile, Kiyoshi Isono et.al., the J, Antibiotics, Ser.A., 10: 21-30, 1951) revealed that streptomyces was the same substance for hygromycin and homomycin. The strain Noboritoensis (Streptomyces noboritoensis) also only reported that the foreign material. In the present invention, however, the inhibitor of chloroplast formation was found in the culture medium of Streptomyces orivoris KCTC 8426P (Streptomyces olivoviridis KCTC 8426P), which is completely different from the conventional production strain and use for producing hygromycin described above. It was soon discovered that chloroplast formation inhibitors were hygromycin. Hygromycin obtained as a culture product of the microorganism Streptomyces oriboviridis in the present invention has a new effect, unlike the existing effect of inhibiting plant growth due to chloroplast formation inhibition, poor growth and inhibition, in particular in the Examples to be described later As shown in the present invention, growth inhibition occurs when the hygromycin is 50 µg per 1 ml of the total composition, and at 50 µg or more per 1 ml, a significant growth inhibitory effect is evident. Represents herbicidal activity.

Therefore, the herbicide or plant growth inhibitory composition of the present invention can be prepared to be used very effectively for the purpose of herbicide or plant growth inhibition by the use concentration of hygromycin.

As such, unlike the conventional herbicides, the herbicide or plant growth inhibitory composition containing hygromycin according to the present invention is manufactured from a culture product of microorganisms which are harmless to the human body at all, so that the side effects on use or the herbicide of the crop are used. It can solve the problems caused by pesticide pollution.

Hereinafter, the present invention will be described in detail based on the examples, but the present invention is not necessarily limited to the examples.

Example 1

To produce this inhibitor of chloroplast formation, 10 g of water-soluble starch, 20 g of glucose, 25 g of soybean wheat, 1 g of broth, 4 g of yeast extract and inorganic salts (NaCl 2 g, K ₂ HPO ₄ 0.5 g) were added to 1 liter of water and adjusted to pH 7.3. After sterilization, inoculated with Streptomyces orivoviridis KCTC 8426P (Streptomyces olivovirdis) KCTC 8426P) strain, and incubated for 5 days at 28 ℃. After the incubation was completed, the culture solution was collected, and the sample was concentrated to 1/20 using a vacuum concentrator. The concentrated culture solution and the same amount of methanol were added to denature the protein. The precipitated protein was filtered out, and extracted three times with butanol. The extract was dried and subjected to silica gel thin layer chromatography, and developed to examine the activity of each of the separated bands, and the active bands (RF = 0.56) were scraped off from the silica gel plate to extract the active substance with methanol. The methanol solution having the activity was concentrated in a small amount, and about 5 times of petroleum ether was added to precipitate the active material. The precipitated active material was collected and finally separated and purified using high performance liquid chromatography. The column used a C ₁₈ reversed phase column and 15% acetnitryl demand as a solvent, and the active substance was separated hygromycin gently at about 17 minutes.

Example 2

In order to confirm the inhibitory effect of chloroplast formation and plant growth inhibitory effect, the culture solution is recovered after culturing as described in Example 1. Place a 9-cm-sized Sha-Lewi filter paper, add 0.5 ml of culture medium, 4.5 ml of distilled water, 1 ml of culture medium, and 4 ml of distilled water, respectively, and soak the filter paper with radish, Chinese cabbage, alfalfa seeds, and monocotyledonous rice, barley, and seed seeds. Of 10 grains on top of each other and grown for 5 days in the plant incubator (myeong, 30 ℃, 3000 tox, 14 hours-25 ℃, 10 hours cancer) and observed the degree of germination, growth and chloroplast formation. Get the same parentheses.

TABLE 5

In other words, the amount of hair growth and growth of the plant to be examined was not significantly different from the control group in which 0.5 ml of sa-red sugar was added compared to the control group, but yellowing was inhibited by inhibiting chloroplast formation. In addition of 1ml culture medium, all of the plants examined showed good germination, but growth was inhibited due to growth inhibition, and chloroplast formation was completely inhibited.

Example 3

In order to check the effect of plant growth inhibition and chloroplast formation by germinating soil before germination, test with 3cm diameter, fill the soil, seed the radish seeds, and cover them with the same soil. A 1 ml solution per test tube is dispersed using a pipette of a solution of the present substance prepared at a predetermined concentration as shown in Table 6. After dispersal, the test tube is grown in a greenhouse controlled at 30 ° C for 2 weeks, and the growth and inhibition of chloroplast formation are examined. When tested according to the above examples, the results are obtained as shown in Table 6 below.

TABLE 6

According to the present invention, in the case of herbicide composition, the treatment concentration is 50-500 µg / ml of hygromycin, as in the measurement results of the above examples, and the known additives such as surfactants, adjuvants and extenders, etc. The same effect can be obtained by mixing and preparing in the form of these granular, powdery, liquid or the like.

At this time, the herbicide composition showed an effect on both dicotyledonous weeds, monocotyledonous weeds, or was shown to be superior to herbicide activity than in boxy weeds.

Claims (1)

Receiving starch 10g, glucose 20g, Soy binmil 25g, juicy 1g, 14g of yeast extract and inorganic dyes _{(Ncal 2g, K 2 HPO 4} , 0.5g) was added to a Streptomyces duck 1ℓ water is adjusted sterile PH ₃ bobiri disk ( Streptomyces olivovirdis) Plant growth inhibitor composition containing hygromycin obtained from the culture product of KCTC 8426P as an active ingredient.