Sterilization Characteristics of Ozone-mist Spray for Chemical FreeAgriculture

K. Ebihara1,2, F. Mitsugi2, T. Ikegami2, Y. Yamashita3, Y. Hashimoto3, T. Yamashita3,S. Kanazawa4, H. D. Stryczewska5, J. Pawlat5, S. Teii6, and T.-L. Sung6

1Environment and Energy Laboratory, Japan2Department of Computer Science and Electrical Engineering, Kumamoto University, Japan

3Sanwa hi-tech Co. Ltd, Japan4Department of Electrical and Electronic Engineering, Oita University, Japan

5Lublin University of Technology, Poland6Lunghwa University of Science and Technology, Taiwan

Abstract—We have developed an ozone-mist spray system for chemical free agricultural sterilization. This environmentally friendlysystem is the movable and backpack type in which the high density ozone is generated by surface dielectric barrier discharge inoxygen. The ozone gas is injected into the water-mist which is produced by the water spray nozzle. The ozone-mist is sprayed overharmful insects, viruses and bacteria living on/in the plants. The ozone solubility of the ozone-mist water is over 5ppm in the caseof standard treatment conditions (oxygen flow rate 1 L/min, mist water flow rate 0.3 L/min). Many kinds of harmful insects weresterilized to evaluate the sterilization characteristics and operation management by changing conditions such as ozone concentration,oxygen flow rate, mist water flow rate, distance between nozzle and biological objects using various structures of the spray nozzlesystem. Typical experiment shows that the survival rate in the case of aphids (small insects of a few mm size) becomes practicalvalue of about 10% for very short spray time of 10 sec.

Keywords—Ozone-mist, sterilization, spray, chemical-free, harmful insect

I. INTRODUCTION

Ozone (O3) is created naturally in the upper atmospherethrough ultraviolet radiation and is also produced in nature bylightning. Ozone as nature’s most powerful oxidizer convertsorganic materials into their base compounds. Natural ozonehas played an important role in purification and sterilization ofatmosphere, ground and water since time immemorial. Recentmass production in large-scale agricultural farming supportedby monoculture, agricultural chemicals, high mechanizationetc. resulted in loss of biological diversity, fungus resistance,degradation and erosion of soil, pesticide residues and pollu-tion (air, soil and water). In order to solve these global envi-ronmental issues occurring in human life, ozone has attractedmuch interest in alternative technology for sterilization andpurification in agricultural management [1]–[6]. Ozone is oneof alternatives to agricultural chemicals and helps promote amore sustainable agricultural system. The electric discharge inair can provide various state of ozone available for sterilizationin agriculture.

Agricultural sterilization using ozone have been studied forsoil treatment and pest control. Choi showed that the dielectricbarrier discharges at atmospheric pressure sterilized E. coliwith 99.9% effectively in air and that ozone molecules werethe dominant germicidal species [7]. Qui showed that at dosageequivalent to 250 kg of ozone/ha, the parasitic nematodes and

Corresponding author: Kenji Ebiharae-mail address: kankyo@mild.ocn.ne.jp

Presented at the 2014 International Symposium on Electrohydrodynamics(ISEHD 2014), in June 2014

free-living nematodes in soil were reduced by 68% and 52%respectively [8]. On the other hand, there are few studies ondetailed study describing the ozone sterilization of harmfulinsects such as aphids and worms living on plants.

We have studied fundamentals of ozone sterilization and itsapplication to sterilization in agriculture [9]–[20]. The DNA orRNA genome contained in bacteria and viruses is oxidized anddestroyed by ozone under appropriate treatment conditions.Our previous experiments showed that ozone treatment of 5%wt. ozone breaks the structure of the λ-E. coli DNA [9]–[19].

We have developed an ozone-mist sterilization which iscarried on the back of farmers [20]. This backpack type systemis designed for agricultural farms located in remote rural areaswhere ethnic groups have engaged in chemical-free agricultureunder poor infrastructure of electrical power supply.

Technological requirements for this chemical-free steriliza-tion system are followings;

1) kill living small insects, worms, bacteria and viruses athigh sterilization rate (∼90%) in short operation time(10∼20 sec);

2) oxygen gas and water as original materials producean ozone-mist creating pesticide radicals which killmicroorganism;

3) secure farmer health from ozone exposure;4) leave no harmful residues on the produce.The ozone-mist of this system is generated by injecting

high dense ozone gas into sterilization space surrounded withwater-mist. Chemical reaction between ozone and water in theozone-mist provides ozone derivative radicals such as hydroxylradical (OH · ) and the superoxide radical anion (O ·–

2 ) whichhave high redox potential.

Ebihara et al. 11

Fig. 1. Outline of ozone-mist sterilization system.

Fig. 2. Ozone-mist sterilization system (1: Ozone generator, 2: High-frequency source(Inverter), 3: Li-ion battery, 4: O2 vessel, 5: Water pump,6: Water tank, 7: Ozone-mist spray).

We report here sterilization characteristics of this ozone-mist sterilization system. Harmful insects such as aphidsand worms are tested under various sterilization conditionsincluding ozone concentration, mixture rates between ozoneand mist-water, treatment time, and separation distance fromtargets living on the plants.

II. EXPERIMENTAL

Our system of the ozone-mist sterilization is composed ofan ozone generator, a water-mist supply system, an AC-DCinverter connected to Li-ion battery, a lightweight oxygenvessel (FRP) with a pressure regulator, a water tank and amovable spray rod with ozone-mist nozzle [20]. Fig. 1 showsthe outline of our sterilization system and Fig. 2 is a pictureof the latest prototype system (size: height 620 mm, width360 mm, depth 180 mm, weight 15 kg).

Fig. 3. Ozone-mist structure.

High dense ozone which is suitable to sterilize insects,worms, bacteria and viruses is generated by surface dielectricbarrier discharge (SDBD) [21]–[23].

The HV electrode of the SDBD is embedded inside the di-electric plate and molded. The dielectric barrier discharge wasgenerated on the surface electrode placed on the dielectricsAl2O3 (146×146 mm). The AC electric power for the highfrequency generator (15 kHz, Vp−p = 8 kV) was supplied fromthe AC-DC inverter. The SDBD system was cooled by air.

Although the concentration of the ozone generator changeswith oxygen gas flow rate, a concentration of 70 g/m3 atan oxygen flow rate of 1 liter/min was used for most ofsterilization experiments. In the practical use at agriculturalfield, the ozone concentration is expected to be adjusted bychanging oxygen flow rate at a fixed high frequency voltage(Vp−p= 8 kV).

Fig. 3 shows the outline of the ozone-mist structure. Water-mist was produced by supplying the water to four mist nozzlesplaced concentrically. High dense ozone was injected into thewater-mist through an ozone gas nozzle.

When ozone reacts with water-mist, highly unstable andrapid decomposition occurs. The elementary reactions follow:

OH · +O3 −−→ HO2 · +O ·−2 (1)

HO2 · −−→ H+ +O ·−2 (2)

O ·−2 +O3 −−→ O2 +O ·−

3 (3)O ·−

3 +H+ −−→ HO3 (4)HO3 −−→ O2 +OH · (5)

O3 +OH · −−→ HO2 · +O2 (6)

Initially, the superoxide radical anion (O ·–2 ) and the hy-

droperoxyl radical (HO2 · ) are formed and then lead to thegeneration of the highly reactive hydroxyl radical (OH · ). Theozonide radical ion (O ·–

3 ) is formed as an intermediate reac-tion product [24]. The ozone and generated reactive radicalshave strong oxidation mechanism due to high redox potentialcomparable to fluorine F (2.87 V); OH · (2.86 V) and O3(2.07 V). A half-life time of ozone in aqueous solution wasmeasured to be in the range from 2 min to 165 min dependingon the conditions such as temperature, pH value and gasdynamics [24].

The generated ozone-mist was sprayed on the target samplessuch as insects and plants as illustrated in Fig. 3.

The separation distance (d) between ozone nozzle exit andthe target was kept at about 20 mm. The ozone solubilityof the ozone-mist water is over 5ppm in the case of oxygen

12 International Journal of Plasma Environmental Science & Technology, Vol.10, No.1, MARCH 2016

TABLE IINSECTS TREATED

InsectAphid Red aphid (Uroleucon nigrotuberculatum)

Cotton aphid (Gossypii glover)Crysanthermum aphid (Mocrosiphonella Sanbolnl)Rose aphid (Sitobion ibarae)

Tobacco worm Lasioderma serricorne(Fabricius)

Green rice leaf hopperCaterpillar

Fig. 4. Aphids after treatment (Inset is microscopic photo of red aphid).

flow rate 1 L/min and mist water flow rate 0.3 L/min. Ozone-mist sterilization was performed for some kinds of aphids andworms listed in Table I.

III. RESULTS AND DISCUSSION

Fig. 4 shows a typical photo of the red aphids after ozone-mist treatment. In this experiment, the target samples such asaphids were captured on the mesh sheet set at the bottom of atest PET cup (top diameter 90 mm, bottom diameter 60 mm).

We took photos of the aphids on the mesh every 30 min aftertreatment. The dead aphids remained at the same places on thesheet and alive aphids escaped from the sheet. We counted thenumber of dead aphids (including apparent suspended death)lying on the sheet. The survival rate defined as (alive number/total number) is a dominant factor to evaluate the sterilizationsystem.

Fig. 5 shows the variation of the survival rates as a functionof the time (t) after sterilization when the samples were treatedfor different treatment times (T = 5 sec, 10 sec, 20 sec).A survival rate of 10% at t = 180 min is attained for thepractical operation time of T = 10 sec where the samplenumber (N ) under the test was N = 20. Longer treatmenttime of T = 20 sec gives complete death (N = 25) due toaccumulative increase of the gaseous ozone and its derivativeradicals taken into aphid body. In the case of short treatmenttime of T = 5 sec, 7 aphids in N = 26 were alive (survival rate= 26.9%). Although immediately after the treatment most ofaphids showed apparent death, some aphids gradually survived

Fig. 5. Survival rate as a function of time (t) after treatment at varioustreatment times (T ).

and finally died in a few days. When only the gaseous ozoneunder same conditions (70 g/m3 O3, 1 L/min) without water-mist was used, most of aphids died during very short treatmenttime of 5-10 sec. However, long-term exposure to high densegaseous ozone by this method causes serious damage onhuman body.

We performed statistical analysis on the results obtainedthrough the ozone-mist sterilization operation. Fig. 6 showsthe normal distribution (F ) of the survival rates when redaphids were treated during 10 sec. Eleven sterilization data(denoted by black square in Fig. 6) were taken at sametreatment conditions of an ozone concentration of 70 g/m3

at oxygen gas flow of 1 L/min and a mist-water flow rate of0.3 L/min. This profile shows that a most probable value of thesurvival rate is 13% with mean 13.3 and standard deviationof 12.6%. The deviation is mostly caused by body growingstages (larva, imago) and spatial non-uniformity of ozone-mist sprayed on the targets staying on the bottom mesh sheet(diameter 60 mm).

Plant viruses including tobacco mosaic viruses and cucum-ber mosaic viruses can be transmitted by aphids. Gaseousozone and its derivative radicals are taken into the aphidbody through spiracles, travel via the trachea through the bodyand reach the cells. As a result, the aphid may be killed bymolecular ozone and its derivative radicals acting on proteinsand organelles of the cell.

Concentration of ozone derivative radicals in ozone-mistmatter is one of essential factors to control sterilization effectof pest in agriculture. Kanazawa developed the terephthalatedosimetry for in-situ OH radical evaluation in liquids [25],[26]. Fluorescence of the 2-hydroxylterephthalic acid (HTA)was measured by using terephthalic acid (TA) as an OH radicalscavenger. This method was introduced to estimate OH radicalconcentration of the ozone-mist matter which was gener-ated by our sterilization system. Our preliminary experimentsshowed OH radicals concentration of the ozone-mist water

Ebihara et al. 13

Fig. 6. Normal distribution profile of survival rate for aphids treated during10 sec. Data (black square) were obtained at 70 g/m3 ozone with an oxygenflow rate of 1 L/min and mist-water flow rate of 0.3 L/min.

was estimated to be 7ppb (4.5×10−7 M) under an ozone gasconcentration (at SDBD tube) of 70 g/m3 (35,000ppm). Thisvalue of OH radical concentration is lower than the publisheddata of 0.05-1.4ppm for the TA solution (350 L/min) intowhich ozone gas (concentration: 1000-1000ppm) was drawnthrough a venturi tube [27]. We promote further detailed studyon OH and other radical measurement of the ozone-mist matterunder various sterilization conditions.

Since aphids have hydrophobic surface skin, droplets ofwater containing ozone gas are shed. Considering adsorptionmechanism of ozone and the related radicals into aphid body,the ozone-mist sterilization developed here is advantageouscomparing to the conventional ozonated water spray method.

IV. CONCLUSION

We developed a portable ozone-mist spray system to controlharmful insects (pests) in agriculture. The survival rate ofaphids using this ozone-mist sterilization system was stud-ied. On-site generation of ozone derivative radicals such ashydroxyl radical (OH · ), superoxide radical anion (O ·–

2 ), andhydroperoxyl radical (HO2 · ) in the ozone-mist process playseffective role on increasing the sterilization rate. In the case ofred aphids, the survival rate was 10% at short treatment timeof 10 sec. The ozone-mist spray sterilization is a promisingmethod to secure farmer health from ozone exposure and torealize chemical-free agriculture at low sterilization cost.

ACKNOWLEDGMENT

We would like to express sincere thanks to Lunghwa Univer-sity of Science and Technology (Taiwan), Taiwan AgriculturalResearch Institute (Taichung), National Pingtung Universityof Science and Technology (Taiwan), Livestock ResearchInstitute of Council of Agriculture (Ilan, Taiwan) for their kindsupport to promote our R&D project.

We also thank President Wang of Yunnan Science-Technology Trade Co. Ltd and Mr.Dai of Hongta TobaccoGroup Co. Ltd. for their encouragement extended to our

research and thoughtful support of tobacco farm experiment atYunnan (China), and Dr. Feng Haiying of Tsinghua Universityand Mrs. Yao Shuen of Shanghai Representative Office ofKumamoto Prefecture for their international coordination andcordial support to progress our research and development inChina.

This work has been performed under the promotion projectfor global technology cooperation of the Ministry of Economy,Trade and Industry of Japan.

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