A non-toxic insect control method using ozone and carbon dioxide synergistically

Abstract

This invention provides a non-toxic insecticidal method using ozone and carbon dioxide in combination, comprising the following steps: Step 1: Place the items to be insecticidal into a sealed insecticidal chamber; Step 2: After sealing the chamber, evacuate the air to 0.2–0.9 bar and maintain this low pressure for a period of time; Step 3: Introduce ozone at a concentration of 40–160 ppm and control the pressure inside the chamber at 1.0–1.7 bar for 3–20 minutes; Step 4: Evacuate the air inside the chamber again to 0.2–0.9 bar and maintain this low pressure for a period of time; Step 5: Introduce high-concentration carbon dioxide into the chamber and control the pressure inside at 1.0–1.7 bar for 10–30 minutes; Step 6: After insecticidal treatment, exhaust the air completely, restore normal pressure, and remove the insecticidal items. This invention has the advantages of being non-toxic, highly efficient, and rapid in insecticidal treatment. It can reduce the oxidative damage of ozone to insecticidal items and equipment, has low requirements for the strength and performance of the equipment used, and has wide applicability.

Description

[Technical Field]

[0001] This invention relates to a non-toxic insecticidal method using ozone and carbon dioxide in synergy. [Background Technology]

[0002] Currently, fumigation is a commonly used method for pest control, but most of the gases used are toxic, including methylene bromide, ethylene oxide, hydrogen cyanide, and phosphine, some of which are carcinogenic and face elimination. In recent years, the use of environmentally friendly and non-toxic carbon dioxide (CO2) for pest control has become increasingly popular. However, using CO2 alone requires a long time. Goerke used 10% CO2 to kill insects in greenhouses for 12 hours a day for a week, which proved effective (K. Goerke et al., Response of aphids and greenhouse plants to insecticidal concentration of carbon dioxide, J. Plant Disea and Prot, 2005, 112, 5, 50). Wang used 60% CO2 to treat post-harvest asparagus in an open environment, and pest control took 48 hours or longer (LXWang et al., Effect of high CO2 treatment and MA packing on sensory quality and physiological-biochemical characteristics of green asparagus during postharvest storage. Horticulturae, 2020, 6, 84). Navarro first evacuated the pest control space and then treated the mites with CO2 at room temperature and pressure. A low concentration of CO2 (30%) could kill 89% of the mites' eggs, but it took 16 hours (SNavarro et al., Integrated storage pest control method using vacuum or CO2 in transportable system, Integrated protection of stored production, IOBC Builerin, 2002, 25(3), 207). Sadeghigt, when subjected to ambient temperature and pressurized CO2 at 0.5 atmospheres, can kill 90% of insects within 24 hours (GR Sadeghite et al., High-pressure carbon dioxide use to control dried apricot pest, Foods, 2021, 10, 1190).Shimma pressurized CO2 to 5–15 atmospheres, achieving insect control in 1.8 days (SH Shima et al., Comparative effect of CO2 in modified temperature and pressure condition on adults and larvae of the ed flourbeetle. Coleopterist Bull, 2020, 74(1), 127). Serrani directly pressurized CO2 to 1.8 atmospheres, achieving effectiveness in as little as 90 minutes, but only after 120 minutes did it show insecticidal effects (Serrani et al., Industrial application of carbon dioxide and high pressure as a method to pest control on foodstuff raw materials, Adv AgriHort Ento, 2020, 3, 38). It is evident that CO2 insecticidal operations, whether at normal pressure, pressurized, or depressurized, require sterilization times exceeding 2 hours. In addition, ozone, another environmentally friendly and non-toxic gas, has also been used for fumigation and pest control in recent years. Sharma's review shows that due to the high oxidizing power and concentration of ozone, general equipment is easily oxidized and products are easily deteriorated, which limits the effectiveness of ozone in pest control.

[0003] In recent years, many people have used ozone mixed with carbon dioxide for sterilization. Goerke et al., Golestan et al., Mun. Ent. Zool, 2016, 11, 169, recently reported that CO2 mixed with ozone could kill insects after 7 days. Sekhon et al. used carbon dioxide mixed with ozone for ham mite control, requiring 96 hours to kill the insects. Sadeghi's experiment with mixed carbon dioxide and ozone gas also required 24 hours to kill the insects. This method of using ozone and carbon dioxide mixed gas simultaneously is relatively effective, but it is still limited to economical spaces with a diameter of less than two meters due to the limitations of vacuum or pressure-resistant equipment. Furthermore, the application of ozone mixed with carbon dioxide can reduce the partial pressure of ozone and carbon dioxide (reducing concentration) or cause mutual interference, and the insecticidal efficiency of ozone or carbon dioxide is directly proportional to the concentration. Therefore, the mixed gas method still needs improvement.

[0004] Ozone's insecticidal mechanism primarily involves destroying unsaturated fatty acids, proteins, and polysaccharides on the cell membrane surface and altering permeability, leading to death. Carbon dioxide's insecticidal mechanism, as discovered by Tsao and Caozp, affects the respiratory system and ATP and energy metabolism, causing insect death, reducing NADPH enzyme levels, and killing insects at any stage of their life cycle, including eggs. These two insecticidal gases operate on completely different mechanisms. [Summary of the Invention]

[0005] The technical problem to be solved by the present invention is to provide a non-toxic insecticidal method using ozone and carbon dioxide in combination. This method has the advantages of being non-toxic, highly efficient and rapid in killing insects. It can reduce the oxidative damage of ozone to insecticidal items and equipment, has low requirements for the strength and performance of the application equipment, and has wide applicability.

[0006] This invention is implemented as follows:

[0007] A non-toxic insecticidal method using ozone and carbon dioxide in synergy, comprising the following steps:

[0008] Step 1: Place the items to be exterminated into the sealed insect extermination chamber;

[0009] Step 2: After sealing the insect-killing chamber, evacuate the air to 0.2-0.9 bar and maintain this low pressure for a period of time;

[0010] Step 3: Introduce ozone at a concentration of 40–160 ppm and control the pressure inside the insecticidal chamber at 1.0–1.7 bar for 3–20 minutes to kill insects;

[0011] Step 4: Evacuate the gas in the insecticidal chamber back to 0.2-0.9 bar and maintain the low pressure for a period of time;

[0012] Step 5: Introduce high-concentration carbon dioxide into the insecticidal chamber and control the pressure inside the chamber to 1.0–1.7 bar for 10–30 minutes to kill insects.

[0013] Step 6: After the insect extermination is complete, vent the gas completely and restore normal pressure, then remove the insect extermination items.

[0014] Furthermore, in step 2, after sealing the insect-killing chamber, the air is evacuated to 0.2-0.9 bar, vibrated by a vibrator, and maintained at low pressure for 1.5-2.5 minutes.

[0015] Furthermore, in step 3, the ozone enters from the bottom of the insect-killing chamber.

[0016] Furthermore, in step 4, the low pressure is maintained for 1.5-2.5 minutes.

[0017] Furthermore, in step 5, the concentration of carbon dioxide is 99.5%, and the carbon dioxide enters from the bottom of the insecticidal chamber.

[0018] Furthermore, the pumping pressure in steps 2 and 4 is 0.3 to 0.7 bar.

[0019] Furthermore, the ozone is humidified to 5-95% RH before being introduced into the insecticidal chamber.

[0020] Furthermore, the carbon dioxide is humidified to 5-95% RH before being introduced into the insecticidal chamber.

[0021] Furthermore, the method of the present invention can be applied to kill organisms with respirators or respiratory organs.

[0022] The present invention has the following advantages:

[0023] This invention is designed to operate a system that can simultaneously use ozone and carbon dioxide, integrating two insecticidal gases with different mechanisms into one. It uses a two-stage non-toxic gas insecticidal scheme. First, ozone treatment is used to damage the respiratory tract of insects and increase their hydrophilic permeability. Then, high-concentration carbon dioxide (pressurized) is used. The two work together to kill insects more efficiently and quickly. 100% of insects can be killed in just 20 minutes. The insecticidal efficiency is far higher than that of existing technologies. The short ozone introduction time can reduce oxidative damage to insecticidal items and equipment.

[0024] Furthermore, since the entire system only requires moderate decompression, it can avoid damage to fruits caused by low or high pressure when the pests are being treated. At the same time, it has lower requirements for equipment strength and performance, and can increase the size of the pest control chamber, even allowing an entire shipping container to enter the pest control chamber, greatly expanding its applicability.

Detailed Implementation Methods

[0025] This invention relates to a non-toxic insecticidal method using ozone and carbon dioxide in synergy, the method comprising the following steps:

[0026] Step 1: Place the items to be exterminated into the sealed insect extermination chamber;

[0027] Step 2: After sealing the insect-killing chamber, evacuate the air to 0.2-0.9 bar and maintain this low pressure for a period of time;

[0028] Step 3: Introduce ozone at a concentration of 40–160 ppm and control the pressure inside the insecticidal chamber at 1.0–1.7 bar for 3–20 minutes to kill insects;

[0029] Step 4: Evacuate the gas in the insecticidal chamber back to 0.2-0.9 bar and maintain the low pressure for a period of time;

[0030] Step 5: Introduce high-concentration carbon dioxide into the insecticidal chamber and control the pressure inside the chamber to 1.0–1.7 bar for 10–30 minutes to kill insects.

[0031] Step 6: After the insect extermination is complete, vent the gas completely and restore normal pressure, then remove the insect extermination items.

[0032] Preferably, in step 2, after sealing the insect-killing chamber, the air is evacuated to 0.2-0.9 bar, vibrated by a vibrator, and maintained at low pressure for 1.5-2.5 minutes.

[0033] Preferably, the ozone in step 3 enters from the bottom of the insecticidal chamber.

[0034] Preferably, the low pressure is maintained for 1.5-2.5 minutes in step 4.

[0035] Preferably, the concentration of carbon dioxide in step 5 is 99.5%, and the carbon dioxide enters from the bottom of the insecticidal chamber.

[0036] Preferably, the pumping pressure in steps 2 and 4 is 0.3 to 0.7 bar.

[0037] Preferably, the ozone is humidified to 5-95% RH before being introduced into the insecticidal chamber.

[0038] Preferably, the carbon dioxide is humidified to 5-95% RH before being introduced into the insecticidal chamber.

[0039] The method of this invention is applied to killing organisms with respirators or respiratory organs.

[0040] The technical solution of the present invention will be clearly and completely described below with reference to specific embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer shall apply. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased commercially.

[0041] The method of this invention can be applied to organisms and insect eggs with respirators or respiratory organs. Specific experiments were conducted using scale insects and insect eggs. Since the surfaces of the respiratory organs of scale insects or other organisms that exchange gases are moist, ozone operation does not require additional humidification. When the items to be killed require a humidified environment, ozone and carbon dioxide can be used to humidify the environment to a suitable level before placing them into the insect-killing chamber for insect control.

[0042] Example 1

[0043] A non-toxic insecticidal method using ozone and carbon dioxide in synergy, comprising the following steps:

[0044] Step 1: Transfer the item containing scale insects to be exterminated into a glass bottle and seal the bottle opening with meltblown nonwoven fabric. The filter membrane of the meltblown nonwoven fabric has a pore size of about 10um, which simulates the hiding effect of insects or other organisms hiding in the concave parts of the fruit, the stem, or the bottom of the packaging (this method was tested with stacked custard apples or wax apples, and the results were no different). Then place the glass bottle into a sealed insect extermination chamber.

[0045] Step 2: After sealing the insecticidal chamber, evacuate the air to 0.2-0.9 bar and maintain this low pressure for 2 minutes, and confirm that the insecticidal chamber is sealed.

[0046] Step 3: Introduce ozone at a concentration of 40-160 ppm from the bottom of the insecticidal chamber and control the pressure inside the chamber to 1.0-1.7 bar for 3-20 minutes to kill insects.

[0047] Step 4: Expel the ozone from the insecticidal chamber and evacuate the gas in the insecticidal chamber back to 0.2-0.9 bar, maintaining the low pressure for 2 minutes;

[0048] Step 5: Introduce high-concentration carbon dioxide from the bottom of the insecticidal chamber and control the pressure inside the chamber to 1.0–1.7 bar for 10–30 minutes to kill insects.

[0049] Step 6: After the insect extermination is complete, vent the gas completely and restore normal pressure, then remove the insect extermination items.

[0050] Step 7: Spray the insects with 0.5% methylene blue solution (pale blue) or 0.05% neutral red solution (pale red). Wait 60 minutes; if the color does not fade, the insects are dead. All values ​​are expressed as mean ± SD. Anova or other appropriate statistical analysis was used; P < 0.05 was considered statistically significant.

[0051] Example 2: Relationship between ozone concentration and insecticidal efficacy

[0052] Using the same experimental method as in Example 1, the concentration of introduced ozone was adjusted (ozone concentration 40-160 ppm); in step 2, the insecticidal chamber was evacuated to 0.3 bar; in step 3, the pressure after ozone introduction was controlled at 1.3 bar, and insecticidal treatment lasted for 20 minutes; in step 4, the insecticidal chamber was evacuated to 0.3 bar; in step 5, the pressure after carbon dioxide introduction was controlled at 1.3 bar, and insecticidal treatment lasted for 20 minutes. The specific results of the relationship between ozone concentration and insecticidal efficacy are shown in Table 1 below:

[0053] Table 1

[0054] Insect mortality rate / ozone 40ppm 70ppm 100ppm 130ppm 160ppm Mortality rate of mealybugs (%) 81±11 93±7 100±0 100±0 100±0 Mortality rate of cottony cushion scale (%) 76±10 98±3 100±0 100±0 100±0

[0055] As shown in Table 1, ozone concentrations of 70 ppm or higher are effective in killing scale insects. (When the ozone concentration is 70 ppm, 90 minutes after ozone treatment in step 3, 100% of the insects are still dead.)

[0056] Example 3: Relationship between ozone pressure and insecticidal efficacy

[0057] The experimental method was the same as in Example 2, but the vacuum pressure in step 2 was adjusted (0.2–0.9 bar); the concentration of ozone introduced was 70 ppm, and the other parameters were the same as in Example 2. The specific results regarding the relationship between ozone pressure and insecticidal efficacy are shown in Table 2 below:

[0058] Table 2

[0059] Vacuuming (bar) 0.2 0.3 0.4 0.5 0.6 0.9 Mortality rate of mealybugs (%) 100±0 91±5 82±10 71±13 52±14 30±17 Mortality rate of cottony cushion scale insects (%) 100±0 96±3 85±12 68±17 48±16 37±18

[0060] As shown in Table 2, when the vacuum pressure in step 2 of the insecticidal chamber is reduced to 0.5 bar, 70 ppm ozone can effectively kill insects. However, to completely eliminate the insect population, the vacuum pressure needs to be reduced to 0.2 bar or lower (when the vacuum pressure is 0.3 or 0.4 bar, the insects are still 100% dead 90 minutes after ozone treatment). It is evident that the lower the vacuum pressure in step 2, the higher the ozone pressure after reaching 1.3 bar, and the higher the ozone insecticidal efficiency. Besides the low-pressure effect, the insecticidal efficiency is also directly proportional to the total amount of ozone used. Experiments at 0.4 and 0.5 bar further demonstrate that the respiratory organs of insects treated with ozone are severely damaged, making them prone to death. Considering factors such as equipment and efficiency, the example uses 0.3 bar as the experimental standard.

[0061] Example 4: Ozone treatment time and insecticidal efficacy

[0062] Using the same experimental method as in Example 2, the ozone insecticidal time in step 2 was adjusted (3-30 minutes); the ozone concentration was 70 ppm, and the ozone level was increased to 1.0 bar; all other parameters remained the same as in Example 2. The specific results regarding the relationship between ozone treatment time and insecticidal efficacy are shown in Table 3 below:

[0063] Table 3

[0064] Ozone treatment time (minutes) 3 5 10 15 20 30 Mortality rate of mealybugs (%) 61±16 71±10 91±7 100±0 100±0 100±0 Mortality rate of cottony cushion scale insects (%) 64±14 68±9 87±9 100±1 100±0 100±0

[0065] As can be seen from Table 3 above, even if the vacuum pressure is 0.3 bar in step 2, and ozone is subsequently introduced to 1.0 bar, using 70 ppm ozone for only 5 minutes can effectively damage the insect's respirator or even kill it. Moreover, the 5 minutes of dried ozone has little effect on the oxidation of items or equipment.

[0066] Example 5: Insect-killing efficacy of ozone synergistically with carbon dioxide

[0067] The same experimental method as in Example 1 was used, but the carbon dioxide insecticidal time was adjusted (10-50 minutes); in step 2, the insecticidal chamber was evacuated to 0.3 bar; in step 3, the pressure after ozone introduction was controlled at 1.0-1.4 bar, and the insecticidal time was 5 minutes; in step 4, the insecticidal chamber was evacuated to 0.3 bar; in step 5, the pressure after introducing 99.5% carbon dioxide was controlled at 1.4 bar, and the insecticidal time was 10-50 minutes.

[0068] The specific results of the synergistic insecticidal efficiency of ozone and carbon dioxide at an ozone pressure of 1.0 bar are shown in Table 4 below:

[0069] Table 4

[0070] CO2 insecticidal time (minutes) 10 15 20 30 40 Mortality rate of mealybugs (%) 81±6 91±8 100±1 100±0 100±0 Mortality rate of cottony cushion scale insects (%) 72±11 88±11 97±4 100±0 100±0

[0071] The specific results of the synergistic insecticidal efficiency of ozone and carbon dioxide at an ozone pressure of 1.4 bar are shown in Table 5 below:

[0072] Table 5

[0073] CO2 insecticidal time (minutes) 10 15 20 30 40 Mortality rate of mealybugs (%) 89±8 100±1 100±0 100±0 100±0 Mortality rate of cottony cushion scale insects (%) 92±11 100±0 100±0 100±0 100±0

[0074] This experiment shows that even if the ozone exposure time is reduced to 5 minutes, the insecticidal efficiency of ozone at 1.0 bar combined with carbon dioxide can completely kill insects in about 30 minutes. If the ozone application is increased to 1.4 bar, the insects can be completely killed in 15 minutes (no difference in mortality rate between groups, P>0.05).

[0075] Under the same experimental conditions as in Example 5, with an ozone pressure of 1.4 bar, silkworm eggs were used, and carbon dioxide was introduced for 20 minutes to kill the insects. The experimental group of silkworm eggs had zero hatching. It can be seen that under this experiment, ozone can also cause damage to silkworm eggs, allowing carbon dioxide to enter the eggs or stagnate in the gas chamber, thus destroying the hatching process.

[0076] In summary, the method of the present invention can be effectively used to kill other organisms with respirators or respiratory organs in a non-toxic manner, such as rats, cockroaches, ants, spiders, beetles, horseflies, geckos, silkworms, insect eggs, etc.

[0077] The applicant discovered that scale insects have a waxy substance on their surface, which makes them difficult for water-based reagents to adhere to. However, ozone treatment increases the water solubility and permeability of scale insects. Therefore, dead scale insects treated with ozone are easily stained by water-based dyes such as methylene blue and neutral red, which is used to identify the survival of scale insects. Simultaneously, the applicant found that ozone treatment increases the water solubility and permeability of scale insects, significantly reducing their water resistance. Scale insects with low water resistance are more sensitive to carbon dioxide, and the insecticidal efficiency of carbon dioxide is related to its initial concentration.

[0078] Therefore, this invention proposes a two-stage rapid insect control scheme. First, ozone treatment damages the insect's respiratory tract and increases its hydrophilic permeability. Then, high-concentration carbon dioxide is used to quickly kill the insect. Furthermore, by using a moderate decompression method (i.e., first evacuating to 0.3-0.7 bar), followed by pressurization to a moderate high pressure (ozone or carbon dioxide introduced at a pressure of 1.1-1.7 bar), the medium-pressure equipment is easy to manufacture and can achieve a pressure chamber with a diameter of over 4 meters (length unlimited). Decompression solves the problem of insecticidal gas not easily entering gaps, while pressurization solves the problem of insufficient gas partial pressure. For example, an insecticidal chamber that can withstand moderate decompression of 0.3 bar can naturally withstand moderate high pressure up to 1.7 bar, with a pressure difference of 1.4 bar between decompression and moderate pressurization.

[0079] This method, which involves a brief ozone treatment (minimum 5 minutes) under reduced pressure (0.3–0.7 bar) followed by the introduction of carbon dioxide (1.1–1.7 bar), reduces the oxidative damage of ozone to items and equipment. It also utilizes ozone's lipid-degrading properties to increase sensitivity to carbon dioxide. Furthermore, the pressure difference of one atmosphere (e.g., 0.3–1.3 bar) between low and high pressure is equivalent to forcibly applying one atmosphere (1.0 bar) or a higher concentration of insecticidal gas. This allows even reduced-pressure equipment to achieve near-normal pressure or even higher efficiency than vacuum equipment (e.g., 0.3–1.7 bar = 1.4 bar), thus reducing the requirements for the strength and performance of the insecticidal chamber equipment.

[0080] Therefore, the design of this invention can simultaneously operate ozone and carbon dioxide systems, integrating two insecticidal gases with different mechanisms into one. It employs a two-stage, non-toxic gas insecticidal scheme: first, ozone treatment damages the insect's respiratory tract and increases its hydrophilic permeability; then, high-concentration carbon dioxide (pressurized) is used. The two work together to more efficiently and quickly kill the insects, achieving 100% insect elimination in just 20 minutes, far exceeding the efficiency of existing technologies. Furthermore, because the entire system only requires moderate depressurization, its structure is simple, with lower requirements for equipment strength and performance, allowing for a larger insecticidal chamber—even allowing an entire shipping container to enter. Container transport is currently the most common method of trade, making container-based insect control the most effective and economical method for concentrating products.

[0081] While specific embodiments of the present invention have been described above, those skilled in the art should understand that the specific embodiments described are merely illustrative and not intended to limit the scope of the present invention. Equivalent modifications and variations made by those skilled in the art in accordance with the spirit of the present invention should be covered within the scope of protection of the claims of the present invention.

Claims

1. A non-toxic insecticidal method using ozone and carbon dioxide in synergy, characterized in that: The method steps are as follows: Step 1: Place the items to be exterminated into the sealed insect extermination chamber; Step 2: After sealing the insect-killing chamber, evacuate the air to 0.2 ~ 0.9 bar and maintain this low pressure for a period of time; Step 3: Introduce ozone at a concentration of 40-160 ppm and control the pressure inside the insecticidal chamber at 1.0-1.7 bar for 3-20 minutes to kill insects; Step 4: Evacuate the gas in the insecticidal chamber back to 0.2 ~ 0.9 bar and maintain the low pressure for a period of time; Step 5: Introduce high-concentration carbon dioxide into the insecticidal chamber and control the pressure inside the chamber to 1.0 ~ 1.7 bar for 10 ~ 30 minutes to kill insects; the concentration of carbon dioxide in step 5 is 99.5%, and the carbon dioxide enters from the bottom of the insecticidal chamber; Step 6: After the pest control is completed, vent the gas completely and restore normal pressure, then remove the pest control items. Insects are organisms that have respiratory organs or respiration apparatus.

2. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: In step 2, after sealing the insect-killing chamber, the air is evacuated to 0.2 to 0.9 bar, and the chamber is vibrated by a vibrator and maintained at low pressure for 1.5 to 2.5 minutes.

3. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: In step 3, the ozone enters from the bottom of the insect-killing chamber.

4. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: In step 4, maintain the low pressure for 1.5-2.5 minutes.

5. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: The suction pressure in steps 2 and 4 is 0.3 to 0.7 bar.

6. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: The ozone is humidified to 5-95%RH and then introduced into the insecticidal chamber.

7. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: The carbon dioxide humidifies the air to 5-95% RH before being introduced into the insecticidal chamber.

8. The non-toxic insecticidal method using ozone and carbon dioxide synergistically according to claim 1, characterized in that: This method is applied to killing organisms with respiratory organs.