CN104585167B - The preparation of extraction fromginkgoseed coat nano-capsule - Google Patents

Abstract

The invention discloses the preparation of a nanocapsule of ginkgo exocarp extract. The steps are: (1) dissolving 0.3 g of ammonium persulfate (initiator) in 60 g of water; (2) weighing 1.5 g of ginkgo exocarp extract (3) Dissolve 18g OP-50 and 3g sodium lauryl sulfate in 150g 2% polyvinyl alcohol aqueous solution, add mixed monomer, and disperse into an emulsion; (4) Heating the emulsion to maintain a temperature of 70 ℃, add the first part of initiator to obtain seed emulsion; dropwise add 4.5g of styrene, 4.5g of butyl acrylate, 3g of acrylic acid mixed monomer and the second part of initiator, add auxiliary agents and discharge; the microcapsules are discharged under high temperature and It can not produce precipitation at low temperature and has the characteristics of good physical and chemical stability. The preparation uses water as a dispersion medium, does not contain or contains little organic solvents, conforms to the development direction of current pesticide formulations, and meets the requirements of current agricultural sustainable development.

Description

Preparation of nanocapsules of Ginkgo biloba extract

technical field

The invention relates to the preparation of a pesticide, in particular to the preparation of a ginkgo exotesta extract nanocapsule.

Background technique

As the social and ecological problems caused by pesticides have attracted increasing attention, the use and function of chemical pesticides have been widely questioned. In the 1980s and 1990s, many experts at home and abroad believed that the use of chemical pesticides would come to an end, and would be gradually replaced by biological control, physical control and comprehensive control. But the facts show that in the foreseeable historical period, pesticide and chemical control methods will still not be completely replaced. Since 2000, China's Ministry of Agriculture and local governments at all levels have successively issued relevant laws and regulations, prohibiting the use of many highly toxic and high residue chemical pesticides (such as carbofuran, methyl parathion, etc.). According to the "Twelfth Five-Year Plan" of the pesticide industry, in 2015, high-efficiency, safe, economical and environmentally friendly pesticide varieties should account for more than 50% of the total output. Therefore, the development of biopesticides that are highly effective against harmful organisms, safe for non-target organisms, easy to decompose, and the decomposition products are not harmful to the environment is currently a field of extensive research and application ^[5] , and botanical pesticides have become one of the more ideal alternative products. one.

Botanical pesticides refer to pesticides that use certain parts of plants or extract their effective active ingredients to make pesticides that have antibacterial or insecticidal effects. The formation of plant-derived active ingredients and their widespread existence in nature are the result of adapting to the process of biological evolution. In the control of crop diseases and insect pests, they are environmentally friendly, pollution-free, generally low in toxicity, easy to degrade, and have no residue. It has the advantages of making diseases and insects produce corresponding resistance to pesticides, and is the preferred pesticide variety for the production of pollution-free agricultural products ^[6] .

There are many ways to research and develop botanical pesticides, mainly as follows:

(1) Extract directly from plants with rich plant resources, low cost, large biological harvest, and high content of active ingredients. If the complex active ingredients have strong activity and are difficult to achieve artificial synthesis, the plant itself or its extracts can be directly processed into pesticide products.

(2) Fully artificial biomimetic synthesis method For plant species that are relatively scarce in plant resources and conflict with traditional Chinese medicine materials, have little content in plants but high biological activity and relatively simple compound structures, artificial synthesis or modification can be used to add other Structure, synthesis of compounds with higher or higher activity, including directed synthesis and template synthesis.

(3) Compounding According to the theory of compatibility of traditional Chinese medicines, scientific and reasonable compatibility of traditional Chinese medicines can produce obvious synergistic effects or produce new active ingredients. Most of the plants in botanical insecticides are also the objects of traditional Chinese medicine. Therefore, drawing on the theory of traditional Chinese medicine, compounding botanical insecticides can also improve their control effect on harmful organisms.

In recent years, at home and abroad, attention has been paid to the extraction and utilization of active substances in the outer testa of Ginkgo biloba. Current experimental research shows that the active substances extracted from the outer testa of Ginkgo biloba have anti-allergic, anti-inflammatory, antibacterial, anti-viral, anti-aging, and anti-plant diseases and insect pests effects.

The development and utilization of ginkgo testa as a biopesticide is the goal of producing green food and ensuring human health. The acid components in the outer testa of Ginkgo biloba are the main components of biopesticides. They have good application effects in the control of plant diseases and insect pests. They have a wide range of control, do not pollute the environment, have no pesticide residues, and are safe for humans and animals. Long-term use of this pesticide will not cause pests. drug resistance, its production equipment is simple, and its production cost is low. Production practice shows that the effect of Ginkgo biloba exotesta extract on controlling bean aphids is 70%, that of beetles is 85%, that of Spodoptera litura is 90%, and that of cabbage caterpillar, rice borer and cotton aphid is 100%. The effects of controlling grape powdery mildew, flower black spot, apple anthracnose, pear scab and peach brown rot are all above 80%.

Microencapsulated pesticides are microcapsule preparations formed by coating solid and liquid pesticides in capsule wall materials using microcapsule technology. Pesticide microcapsules can be used to reduce the toxicity of pesticides to humans and animals, reduce evaporation, extend the duration of efficacy, reduce the content of pesticides in the environment, avoid excessive phytotoxicity, and increase the application range of pesticides.

At present, there are few successful cases of combining Ginkgo biloba exotesta extract with microencapsulated pesticides.

Contents of the invention

In view of the above problems, the present invention proposes a preparation of nanocapsules of Ginkgo biloba exotesta extract, the microcapsules can not produce precipitation at high temperature and low temperature, and have good physical and chemical stability. The preparation uses water as a dispersion medium, does not contain or contains little organic solvents, conforms to the development direction of current pesticide formulations, and meets the requirements of current agricultural sustainable development.

In order to achieve the above-mentioned technical purpose, the present invention adopts the preparation of a kind of ginkgo exotesta extract nanocapsules, comprising the following steps:

(1) Dissolve 0.3g ammonium persulfate (initiator) in 60g water (divided into two parts according to 4:1);

(2) claim that 1.5g ginkgo exocarp extract is dissolved in 21g styrene, 24g butyl acrylate, 0.6g acrylic acid (both liquid) mixed monomer;

(3) Dissolve 18g of OP-50 and 3g of sodium lauryl sulfate in 150g of 2% polyvinyl alcohol aqueous solution, put it into a container, stir, and add mixed monomers dissolved with ginkgo exocarp extract at the same time, and disperse into an emulsion;

(4) Heating the emulsion to keep the temperature at 70°C, adding the first part of initiator, reacting for half an hour, and keeping it warm for one hour to obtain the seed emulsion; add dropwise 4.5g of styrene, 4.5g of butyl acrylate, 3g of acrylic acid mixed monomer and the second Add two parts of initiator dropwise for half an hour, keep warm for one hour, cool down, add additives and discharge.

In the present invention, the auxiliary agent is ethylene glycol.

In the present invention, the auxiliary agent is a mixing auxiliary agent of ethylene glycol and tributyl phosphate.

In the step (3) of the present invention, gelatin: sodium alginate = 1:2 can be used instead of polyvinyl alcohol.

In step (3) of the present invention, gelatin: polyvinyl alcohol = 1:2 can be used instead of polyvinyl alcohol.

According to the mechanism of the preparation of pesticide microcapsules, the present invention has screened out a better ratio of protective colloid - gelatin: sodium alginate = 1:2, and a suitable ratio of auxiliary agents to prepare the effective Ginkgo exocarp extraction The nanocapsule preparation of the substance, the microcapsule can not produce precipitation at high temperature and low temperature, and has the characteristics of good physical and chemical stability. The preparation uses water as a dispersion medium, does not contain or contains little organic solvents, conforms to the development direction of current pesticide formulations, and meets the requirements of current agricultural sustainable development.

Description of drawings

What shown in Fig. 1 is Fig. 1 microcapsule particle size distribution figure among the present invention;

Shown in Fig. 2 is the HPLC spectrogram of standard substance C13:0 among the present invention;

What shown in Fig. 3 is the standard curve figure that reversed-phase high-performance liquid chromatography measures standard substance C13:0 among the present invention;

Shown in Fig. 4 is the HPLC spectrogram of ginkgo exocarp n-hexane extract in the present invention;

Shown in Fig. 5 is the HPLC spectrogram of ginkgolic acid microcapsules of the present invention;

Shown in Fig. 6 is the comparison figure of the corrected death rate of the 3rd instar larva of Plutella xylostella in different concentrations of Ginkgo biloba exotesta extract microcapsules in the present invention;

detailed description

Example 1

The present invention will be described in further detail below in conjunction with the accompanying drawings and specific embodiments.

The first choice of the present invention needs to carry out the preparation of water-suspended nanocapsules:

Composition by weight ratio: Pesticide: 0.2-5%

Mixed monomer: 5-30%

Initiator: 0.05-1%

Deionized water: balance

Additives: 12-25% (preferred)

Followed by the screening of emulsifiers

⑴CMC (Critical Micelle Concentration) - the lowest emulsifier concentration that can form micelles

①Relationship with the size of the hydrophobic group: it decreases with the increase of the number of carbon atoms in the hydrophobic group

② If there are unsaturated bonds on the hydrocarbon group, the CMC will increase

③The CMC increases when the hydrocarbon chain contains polar groups

④ When the hydrogen atoms on the hydrocarbon chain are replaced by fluorine atoms, the CMC decreases

⑤The closer the hydrophilic group is to the middle of the hydrocarbon chain, the greater the CMC

⑥ The CMC of the ionic emulsifier is larger than that of the non-ionic emulsifier

⑵ cloud point

The cloud point is the phenomenon that when the aqueous solution of nonionic emulsifier is heated to a certain temperature, the solution changes from transparent to cloudy. It is a characteristic parameter of nonionic emulsifiers. Ionic _emulsifiers have no cloud point; Emulsion polymerization t _c -t ≥ 10 ° C. In addition, the assembly of hydrophilic groups leads to a rise in cloud point.

(3) HLB value

Large HLB value, hydrophilic; H-LB value small, lipophilic

⑷ Triple point (Craft characteristic) - the characteristic parameter of ionic emulsifier

At a specific temperature, the three phases of true solution, solid, and micelles exist simultaneously, and this temperature is the triple point. Generally speaking, when selecting an ionic emulsifier, it should be ensured that the triple point t _k is lower than the reaction temperature and the minimum storage temperature t, that is, tt _k ≥ 10°C.

A kind of preparation of ginkgo exocarp extract nanocapsule, comprises the steps:

(1) Dissolve 0.3g ammonium persulfate (initiator) in 60g water (divided into two parts according to 4:1);

(2) claim that 1.5g ginkgo exocarp extract is dissolved in 21g styrene, 24g butyl acrylate, 0.6g acrylic acid (both liquid) mixed monomer;

(3) Dissolve 18g of OP-50 and 3g of sodium lauryl sulfate in 150g of 2% polyvinyl alcohol aqueous solution, put it into a container, stir, and add mixed monomers dissolved with ginkgo exocarp extract at the same time, and disperse into an emulsion;

(4) Heating the emulsion to keep the temperature at 70°C, adding the first part of initiator, reacting for half an hour, and keeping it warm for one hour to obtain the seed emulsion; add dropwise 4.5g of styrene, 4.5g of butyl acrylate, 3g of acrylic acid mixed monomer and the second Add two parts of initiator dropwise for half an hour, keep warm for one hour, cool down, add additives and discharge.

In the present invention, the auxiliary agent is ethylene glycol.

In the present invention, the auxiliary agent is a mixing auxiliary agent of ethylene glycol and tributyl phosphate.

In the step (3) of the present invention, gelatin: sodium alginate = 1:2 can be used instead of polyvinyl alcohol.

In step (3) of the present invention, gelatin: polyvinyl alcohol = 1:2 can be used instead of polyvinyl alcohol.

Choice of protectant

In the vinyl monomer emulsion polymerization system, animal glue, gelatin, polyvinyl alcohol, polyvinylpyrrolidone, cellulose derivatives, acrylate and other substances can be used as protective colloids, especially the application of polyvinyl alcohol is the most widely.

The application of polyvinyl alcohol as a protective colloid in the emulsion polymerization of vinyl monomers began in the mid-1930s, especially in the preparation of wood and paper adhesives. The use of protective colloids can increase the initial viscosity of the emulsion in a wet state. Viscosity, mechanical properties of the coating film (such as: shear strength and yield strength) are also greatly improved ^[60] .

Using gelatin: sodium alginate = 1:2 to replace polyvinyl alcohol has the best effect, followed by gelatin: polyvinyl alcohol = 1:2. The effect of using all polyvinyl alcohol is better than that using all gelatin and sodium alginate The effect is good.

Auxiliary selection

The development of pesticide adjuvants in my country started from the research and development of emulsifiers. There are four types of pesticide emulsifiers: nonionic, anionic, cationic, and zwitterionic surfactants, and the most commonly used ones are nonionic, anionic, or a mixture of nonionic and anionic. The main types are alkylphenol polyoxyethylene ether (octylphenol polyoxyethylene ether-OP, nonylphenol polyoxyethylene ether-NP), benzylphenol polyoxyethylene ether (Nongru BP, BC), styrene-based Phenol polyoxyethylene ether (Nongru 600, Nongru BS, Nongru 1601 and 1602, Ningru No. 32), alkylphenol polyoxyethylene ether formaldehyde condensate (Nongru No. 700, Ningru No. 36), castor oil Polyoxyethylene ether (BY), fatty alcohol polyoxyethylene ether (Nongru No. 200), fatty alcohol polyoxyethylene ester, polyol fatty acid ester and its ethylene oxide adduct (Span, Tween series), etc. Ionic surfactants and alkylbenzene sulfonate (Nongru No. 500, DBS-Ca), fatty alcohol polyoxyethylene ether sulfate (AES). It is mainly used for emulsification and dispersion of pesticide emulsifiable concentrate, microemulsion, water emulsion, milk powder, suspoemulsion and other emulsion systems.

The effect of adding 0.5g ethylene glycol to the finished microcapsules prepared by every 20ml is the best, followed by 0.8g ethylene glycol and 0.2g tributyl phosphate mixed auxiliary, 1.5g ethylene glycol, the least effective It is 1.0g of ethylene glycol, and the particle size is very uneven.

Physical properties and characterization of microcapsules (see Figure 1)

The particle size of pesticide microcapsules varies greatly, but generally speaking, D ₉₀ is required to be less than 50 μm, because if the particle size is too large, it is easy to block the pesticide application equipment, and the stability of the microcapsule suspension is also greatly affected, and it is easy to produce knots. lumps and sediment. The study found that the main factors affecting the particle size distribution of microcapsules are emulsification conditions, chemical structure of reaction raw materials, polymerization temperature, viscosity, type of surfactant used, stirring speed and so on. The control of these conditions is particularly important in the preparation process of microcapsules. At present, most of the researches on pesticide microcapsules focus on these aspects. The characterization of particle size distribution can be obtained through microscope observation and statistics, and can also be measured by Malvern particle size analyzer;

Using the ratio of gelatin: sodium alginate = 1:2 as the protective colloid, the particle size classification of microcapsules is 7-40 μm, and the effect is better.

Determination of thermal storage stability

The experiment is carried out according to the determination method of pesticide heat storage stability, that is, the microcapsule sample to be tested is injected into the bottle at first, the bottle is refrigerated with an ice-salt bath, and sealed with a parafilm. Place the sealed bottle in a constant temperature oven at a temperature of 54±2°C, heat it for 14 days, and then take it out. After taking it out, cool it to room temperature. At the same time, wipe off the moisture and other impurities on the outer surface of the bottle and weigh it. If the quality of the microcapsule suspension sample to be tested does not change, check whether flocculent precipitation will occur.

Low Temperature Stability Determination

Carry out experiment by pesticide low-temperature stability assay method, get the microcapsule sample to be tested and be placed in the 50ml beaker, be placed in refrigerator and be cooled to 0 ℃, place and keep 1h, stir once every 15min, each time lasts 15s, observe Changes in appearance. Then put the beaker back into the refrigerator again, and keep it at a constant temperature of 0°C for 7 days. After 7 days, take out the beaker containing the sample and let it stand still to return to room temperature. See if the reagent remains uniform during the temperature recovery process.

Determination of active ingredients in microcapsules

1 Experimental materials and methods

1.1 Preparation of ginkgolic acid standard solution

Precisely weigh 10.0 mg of ginkgolic acid C13:0 single product, fully dissolve it in methanol to a constant volume of 10 mL volumetric flask, prepare a 1.0 mg/ml control solution, shake well, and set aside.

1.2 Preparation of sample solution

Weigh 0.1157g of ginkgolic acid extract, dissolve in methanol, dilute to 25mL, shake well, filter with 0.45um organic microporous membrane, and take the filtrate as the sample solution of ginkgolic acid extract.

Weigh 0.4765g of ginkgo extract microcapsules, dilute with methanol, dilute to a 25mL volumetric flask, shake well, filter with a 0.45um organic microporous membrane, and take the filtrate as the sample for ginkgo phenolic acid extract microcapsules liquid.

Analytical method of ginkgolic acid

Using high performance liquid chromatography:

Chromatographic column: Waters Xbridge C18 (150mm×4.6mm, 5μm);

Mobile phase: methanol: 3% glacial acetic acid solution (90:10);

Detection wavelength: 310nm;

Flow rate: 1mL/min;

Column temperature: 30°C

Ginkgolic acid standard spectrum

The ginkgolic acid standard was analyzed by high performance liquid chromatography, and the results are shown in Figure 2, which shows that the total ginkgolic acid can be completely separated under chromatographic conditions, and the retention time of ginkgolic acid C13:0 is 15.138min. Determination of standard curve

The sample solution of the prepared ginkgolic acid C13:0 standard was filtered through a 0.45um organic filter, and the injection volume was 3ul, 5ul, 7ul, 9ul, 11ul, 13ul, and the peak area was measured. The standard curve in Fig. 3 is made with the peak area Y (volt·second) and the injection volume X (ug), the regression equation is Y=0.9481X-0.876, and the correlation coefficient R ² =0.9993.

Content of ginkgolic acid in ginkgolic acid extract

Using the above-mentioned chromatographic conditions to measure the content of ginkgolic acid in the n-hexane extract of ginkgo exocarp, the HPLC analysis spectrum is shown in Figure 4. According to the calculation method of the total ginkgolic acid in the literature ^[62] , the ginkgolic acid in the extract is obtained. The content is 35.32%.

Content of ginkgolic acid in microcapsules

Using the above-mentioned chromatographic conditions to measure the content of ginkgolic acid in the n-hexane extract microcapsules of Ginkgo biloba ^exotesta , the HPLC analysis spectrum is shown in Figure 5. The content of ginkgolic acid is 2.22%.

Toxicity determination of ginkgo extract microcapsules

The comprehensive toxicity of pesticides to Plutella xylostella was determined by dipping insects and leaves. According to the pre-test results, prepare the test agent into aqueous solution with 5-7 concentration gradients, dip the third-instar larvae of Plutella xylostella one by one in the order from low concentration to high concentration, and the immersion time is 5 seconds. The excess medicinal liquid on the body was put into a petri dish (Φ9cm) covered with filter paper for moisturizing, 10 heads each. After soaking the worms, dip the prepared cabbage leaf dish (Φ1.5cm) in the liquid medicine for 2-3 seconds, take it out, absorb the excess liquid medicine with absorbent paper, dry it, and put it into the corresponding Plutella xylostella handles in Petri dishes. Each concentration treatment was repeated 3 times, and water treatment was used as the control. Each treatment petri dish was placed in the incubator, and the death of the diamondback moth larvae was checked after 72 hours, and the death rate and corrected death rate were calculated. For the data obtained from the biological assay, the toxicity regression equation and the lethal concentration (LC50) of the tested agent were calculated by extreme value analysis.

Determination of Insecticidal Activity of Ginkgo Acid Extract

Take 2 g of ginkgo extract, prepare solutions with different concentration gradients, and use the method 1.1 to determine its insecticidal activity.

data analysis

Analyze and process data with Excel and SPSS data processing software

bioassay results

The indoor toxicity of 6% Ginkgo biloba extract microcapsules to the 3rd instar larvae of Plutella xylostella was measured by the method of soaking worms and leaves. The results are shown in Figure 6. The microcapsules have good insecticidal activity against the 3rd instar larvae of Plutella xylostella xylostella. Showed obvious insecticidal activity at 48h after chemical treatment, and the corrected mortality rates of each concentration (100mg/L, 200mg/L, 500mg/L, 1000mg/L, 5000mg/L) were respectively 13.3%, 23.3% at 48h. %, 30%, 50%, and 66.7%, there were significant differences among them. By 72h, the mortality rate continued to rise, reaching 26.7%, 33.3%, 49.4%, 66.7%, and 89.3%. The differences among treatments were obvious.

As can be seen from the table, the poisonous activity of the ginkgolic acid extract to diamondback moth after being processed into microcapsules is significantly higher than that of the ginkgolic acid extract. This shows that the processing of ginkgolic acid extract into microcapsules is more conducive to the play of ginkgolic acid insecticidal activity.

Claims (3)

1. the preparation of extraction fromginkgoseed coat nano-capsule, it is characterised in that comprise the steps:

(1) 0.3g Ammonium persulfate. is dissolved in 60g water, by first part of initiator: second part of initiator is that 4:1 is divided into two parts；

(2) 1.5g extraction fromginkgoseed coat is claimed to be dissolved in 21g styrene, 24g butyl acrylate, 0.6g acrylic acid mix monomer；

(3) 18gOP-50,3g sodium lauryl sulphate is dissolved in 150g2% polyvinyl alcohol water solution, puts into container, stirring, it is simultaneously introduced the mix monomer dissolved with extraction fromginkgoseed coat, is dispersed into emulsion；

(4) emulsion heating keeps temperature 70 C, adds first part of initiator, reacts half an hour, is incubated one hour, prepares seed emulsion；Dropping 4.5g styrene, 4.5g butyl acrylate, 3g acrylic acid mix monomer and second part of initiator, drip half an hour, be incubated one hour, cooling, addition auxiliary agent discharging, and auxiliary agent is ethylene glycol or ethylene glycol and tributyl phosphate mixed aid.

2. the preparation of extraction fromginkgoseed coat nano-capsule as claimed in claim 1, it is characterised in that in described step (3), gelatin can be used: sodium alginate=1:2 substitutes polyvinyl alcohol.

3. the preparation of extraction fromginkgoseed coat nano-capsule as claimed in claim 1, it is characterised in that in described step (3), gelatin can be used: polyvinyl alcohol=1:2 substitutes polyvinyl alcohol.