초록

○ 연구결과
- 화학적방제체계 : 참다래 저장병 예방 우수약제 4종 선발, 우수 약제 방제 적기 및 횟수 구명, 예방약제 최적 살포프로그램 개발
- 천연물농약개발 : 천연항균성 우수 Eucalyptus 수종 선발, 최적 추출법 구명, 항균활성원인물질 화학적 구조동정, 현장검증을 통한 효과 입증
- 물리적 방제기술 : TiO₂ 광촉매 및 오존 항균활성구명, 최적 처리조건 도출, TiO₂ 광촉매와 오존복합처리 시제품개발 (특허출원), 현장검증을 통한 효과 입증

Abstract ▼

Kiwifruit (Chinese gooseberry, Actinidia deliciosa (A. Chev,) C. F. Liang et A. R. Ferguson) has been grown in southern regions of Korea Since the early 1980s. Kiwifruit is currently cultivated on approximately 1,200 ha in Korea and great amount of the fruits has been imported from foreign countries

Kiwifruit (Chinese gooseberry, Actinidia deliciosa (A. Chev,) C. F. Liang et A. R. Ferguson) has been grown in southern regions of Korea Since the early 1980s. Kiwifruit is currently cultivated on approximately 1,200 ha in Korea and great amount of the fruits has been imported from foreign countries due to increasing demand. Kiwifruit can be maintained more than 4 months when stored at 0±1℃. However, fruit rot diseases caused a severe loss of kiwifruits during storage, transportation, marketing and consumption after harvest. More than 7 fungi were reported to be associated with postharvest fruit rots of kiwifruit. Among the fungi, Botryosphaeria sp. and Phomopsis sp. were reported to be the major causal organisms of postharvest fruit rots of kiwifruit. fruit rots of kiwifruit caused by Phomopsis sp. was named as stem-end rot by Beraha. Sommer and Beraha reported its telemorph state as Diaporthe actinidiae. In Korea, Park et al. reported Phomopsis sp. as one of the major fruit rot pathogens of kiwifruit. Ripened rot of kiwifruit caused by Botryosphaeria dothidea were also reported in Korea. Fungal pathogens isolated from fruit rots were identified as Botryosphaeria dothidea, Diaporthe actinidiae, Botrytis sp., Collectotrichum sp., Penicillium sp., Fusarium sp., and Pestalotiopsis sp.. Among them, Botryosphaeria dothidea and Diaporthe actinidiae were mainly detected at the rates of 83.3％ and 11.9％, respectively. Whereas Diaporthe actinidiae was reported to be dominant fungal pathogen rather than Botryospmeria dothidea in New Zealand, Botryosphaeria dothidea was found to be dominant pathogen in Korea. The fungal pathogen infects kiwifruits during the whole growing season, and heavily during rainy season (July and August) of Korea. Intensive application of fungicides just before or after rainy season can effectively control the fungal infection. Current control methods against these fungi include prehavest sprays with fungicides. Fungicides have been effective in controlling postharvest fungal infection. However, fungicides are under review in many countries due to their possible health risks. As the number of approved fungicides is reduced, there is increased need for technologies that can safely and effectively inhibit fungal growth, reduce postharvest losses and extend market quality. Postharvest dips in fungicides are also effective, but there are persistent residues. Prolonged and intensive use of fungicides may also create a selection pressure for fungicide-resistant pathogens. Curing during prestorage period is commercially applied to control the pathogens, but success is sometimes variable, possibly due to inadequate control of factors, such as relative humidity during the curing process. Experimental control methods such as hot water dips, application of natural coatings and biological control have been attempted, but all theses methods have their shortcomings.
Ozone is a strong oxidant and has been used to disinfest drink water since 1893. The interactions between gaseous ozone and plant pathogens have been extensively studied. Ozone treatment has been attempted to control postharvest fungal pathogens of several fruits.
The photocatalytic activity of TiO₂ for the degradation of organic water pollution is a well-established phenomenon. The reactive oxygen species generated by the TiO₂ photocatalytic reactions under UV irradiation cause various damages to living organisms. Since Matsunaga et al. reported the microbiocidal effect of TiO₂ photocatalytic reactions, research work on TiO₂ photocatalytic killing has been intensively conducted on a wide spectrum of organisms including viruses, bacteria, fungi, algae, and cancer cells. In TiO₂-photocatalytic oxidation processes oxygen is widely used as an additional oxidizing agent. The disadvantage is the slow electron transfer from TiO₂ to O₂. Ozone should be a potential oxidant for photocatalytic oxidation processes.
Until now, few studies on TiO₂ photocatalytic ozonation (the combination of TiO₂ and O₃) have been attempted to degrade organic compounds in water.
There is interest in the use of natural products as safer and more acceptable alternatives to current fungicides. Antimicrobial substances originated from plant have been extensively studied for their use as an agrochemical with highly selective activity against some plant pathogens or as leading molecules for the synthesis of new chemical fungicides. Eucalyptus trees have been known to produce several natural substances having antagonistic activities against several pathogenic microorganisms.
In this research proposal, development of eco-friendly control techniques for integrated pest management of postharvest diseases on kiwifrutis was attempted to reduced the fungal spoilage and improve quality control of kiwifruits during long-term storage in cold room. For this purpose, three main strategies were applied as followings ; 1) establishment of optimum fungicide application programs, 2) development of natural antifungal compounds extracted from Eucalyptus trees, and 3) development of TiO₂ photocatalytic ozonation techniques.
1. Establishment of optimum fungicide application programs, To date, Benomyl WP and Thiophanate WP are the only registered fungicides in Korea for the control of the fruit diseases of kiwifruit. Screening of effective fungicides which can substitute for Benomyl WP and Thiophanate WP and determination of their optimal application program were attempted in this research.
Among the several candidate fungicides tested, Viclozolin WP, Flusilazole WP, Tebuconazole WP and Iprodione WP effectively inhibited the mycelial growths of the causal organisms on laboratory conditions. Flusilazole WP, Tebuconazole WP and Iprodione WP were more effective than Thiophanate WP and as effective as Benomyl WP when sprayed four times at 10-day-intervals from June 16. The control efficacies of the fungicides varied depending upon the spray schedules.
Two-year field performance demonstrated that four times application of the fungicieds at the middle of June was most effective in controlling postharvest decay on kiwifruits. Alternative use of these fungicides is required to avoid emergence of fungicide-resistant pathogens.
2. Development of natural antifungal compounds extracted from Eucalyptus trees
This research investigated the antimicrobial properties of compounds isolated from cold-tolerant eucalyptus trees Eucalyptus darlympleana against the fungal pathogens causing kiwifruit soft rot decay.
Five-year-old trees of Eucalyptus darlympleana were used for the investigation. The tree was proved to be highly cold tolerant and successfully grown under the low temperature of winter seasons in Korea.
Crude methanol extract of Eucalyptus darlympleana inhibited aerial mycelium growth of several plant pathogenic fungi including kiwifruit soft rot pathogens. For chemical identification of antifungal substances, EtOAc soluble fraction and Bu-OH soluble fraction of the leaf extract was investigated. Two phenolic compound (3,4-dihydroxybenzoic acid and gallic acid) and three flavonoid compounds (quercetin, Quercetin -3-O-rhamnoside (quercitrin), Quercetin-3-O-glucoside (isoquercitrin)) were isolated and identified from the two fractions of Eucalyptus darlympleana leaf extract.
Among the five compounds, gallic acid significantly inhibited aerial mycelium growth of Botrytis cinerea over 1,000 ㎎/ℓ concentrations. No inhibitory effect of gallic acid on mycelium growth was found in Botryosphaeria dothidea and Diaporthe actinidiae. Gallic acid has been extracted from several plants and found to have strong antioxidant activity. Many researches on gallic acid currently concentrate antioxidant properties against low-density lipoprotein oxidation in animal cells. The compound was also found to have strong antimicrobial activity against pathogenic bacteria such as Staphylococcus aureus. Little information is available on antifungal activity of the compound so far. This study demonstrated that gallic acid has antifungal activity, but minimum inhibitory concentration (MIC) against fungal growth is much higher than MIC against bacterial growth. Botrytis rot caused by the fungus, Botrytis cinerea is one of the most important postharvest diseases of kiwifruit. Gallic acid also inhibited spore germination in a similar manner with aerial mycelium growth. Spore germination was severely inhibited at 5,000 ㎎/ℓ in all the tested pathogens, and completely in Botrytis cinerea. Since Botrytis cinerea is major fungal pathogen causing serious economic losses in kiwifruit industry and was found to be sensitive to the compound, gallic acid as a natural product can be safer and more acceptable alternative to current fungicides controlling stem-end rot of kiwifruit. In addition to direct spoilage of diseased tissue, fruit infected by the fungus produces small amounts of ethylene in cool storage. Ethylene released by infected fruit could in principle cause further losses by softening adjacent sound fruit. It is speculated that gallic acid as a strong antioxidant agent can scavenge the oxidants and subsequently prevent ethylene production in infected fruit. This can delay fruit softening and reduced soft rot decay.
Systemic investigation on the speculation is further needed. This is the first report on the isolation of gallic acid from Eucalyptus darylmpleana, which has antifungal activity against mycelium growth and spore germination of Botrytis cinerea.
3. Development of TiO₂ photocatalytic ozonation techniques In this research, TiO₂ photocatalytic ozonation was attempted to disinfect the fungal pathogens causing postharvest spoilage of kiwifruit and to decompose residues of the fungicides sprayed at preharvest or postharvest for the fungal spoilage control.
Device for TiO₂ photocatalytic ozonation was invented for the treatment of kiwifruits to prevent fungal spoilage and degrade residual fungicide.
Degradation of Methylene Blue by oxidative reactions of ozonation and/or TiO₂ photocatalysis was examined to find out their efficacies of organic matter decomposition. The degradation rate of TiO₂ photocatalytic ozonation (TiO₂/O3) is about 3 times higher than the degradation by ozonation (O3) under same condition without TiO₂ and 12 times higher compared to elimination by TiO₂ treatment. This result showed that both processes TiO₂/O₂ and H₂O/O₃ occurred during the combined photocatalytic ozonation that more rapidly eliminated the dye than individual process. The degradation rate of TiO₂/O₃ process is about 2.3 times higher than the sum of the degradation rate of TiO₂/O₂ and H₂O/O₃. The synergistic degradation implies that there were additional reactions leading to the faster decomposition of the dye. The higher oxidation rate might be due to a photocatalytic-induced decay of ozone, initiated by the combination of TiO₂ and UV illumination.
Ozonation was more effective than TiO₂ photocatalysis in germicidal activity of the fungal spore. Germination of the spores was more apparently inhibited by TiO₂ photocatalytic ozonation than ozonation (O₃) or TiO₂ phoptocatalytic oxidation alone. TiO₂ photocatalytic ozonation synergistically inhibited conidial germination as found in Methylene Blue degradation. About 95% of conidia were successfully disinfected by the exposure to TiO₂ photocatalytic ozonation process within 5 min.
Fungicide flusilazole was effectively decomposed by TiO₂ photocatalytic ozonation. The concentration curve was approximately linear. The decomposition rate determined by regression was calculated as 1.71 μM/min (P=0.001). Approximately 70% of the flusilazole (initial concentration of 100 ㎎/ℓ) was decomposed in the photocatalytic ozonation reactor within 120 min.
TiO₂ photocatalytic ozonation successfully controlled fungal spoilage of kiwifruits and showed higher efficacy of disease control than fungicide (flusilazole) treatment. Examination of whole storage period showed that TiO₂ photocatalytic ozonation most successfully prevented fungal spoilage when kiwifruits were treated with the device just after harvest. This result clearly demonstrates that TiO₂ photocatalytic ozonation can successfully retard subsequent decay during cold storage and distribution in markets.
The efficient control of fungal spoilage and removal of residual fungicides indicate that TiO₂ photocatalytic ozonation is an very attractive method for postharvest disease control of kiwifruits with the reported benefits of alternative to chemical control methods.
Now, postharvest decay of kiwifruit is definitely controlled by IMP programs including the new informations and technologies induced from this research project.

목차 Contents

• 표지 ... 1
• 제출문 ... 2
• 요약문 ... 3
• Summary ... 12
• Contents ... 18
• 목차 ... 19
• 제1장 연구개발과제의 개요 ... 21
• 제1절 연구개발의 목적 ... 21
• 제2절 연구개발의 필요성 ... 22
• 제3절 연구개발의 범위 ... 26
• 제2장 국내외 기술개발 현황 ... 27
• 제3장 연구개발수행 내용 및 결과 ... 30
• 제1절 농약사용량 절감을 위한 참다래 저장병 화학적 방제체계 확립 ... 30
• 1. 참다래 저장병 화학적 방제체계 확립을 위한 저장병 예방약제 선발 및 살포 적기 구명예방약제 선발 및 살포 적기 구명 ... 30
• 1) 참다래 저장병 예방약제 실내선발 ... 30
• 2) 참다래 저장병 예방약제 약효 포장검정 ... 31
• 3) 참다래 저장병 예방약제 살포적기 구명 ... 34
• 2. 참다래 저장병 예방약제 적정 살포횟수 구명 ... 40
• 3. 참다래 저장병 예방약제 최적 살포 프로그램 정립 및 포장 검증 ... 45
• 4. 농약사용량 절감을 위한 참다래 저장병 화학적 방제체계 확립 결과 요약 ... 49
• 제2절 천연항균성 물질을 이용한 환경친화형 참다래 저장병 방제기술 개발 ... 50
• 1. 참다래 저장병 천연물농약 개발을 위한 Eucalyptus 수목의 엽육 추출액 대량회수 최적조건 구명 ... 50
• 1) Eucalyptus 수목의 엽육 추출액의 참다래 저장병균에 대한 항균활성 검정 ... 50
• 2) Eucalyptus 수목의 엽육 추출 방법에 따른 항균 효과 검증 ... 53
• 3) Eucalyptus 부위별, 시기별 추출액의 참다래 과실무름병원균에 대한 항균 활성 검정 ... 55
• 2. Eucalyptus 엽육 추출액 대량회수 최적조건 구명 ... 57
• 1) 항균물질 추출 및 분리에 적합한 최적 용매 및 전개조건 구명 ... 57
• 2) Eucalyptus darlympleana 엽육 추출액의 chloroform과 butanol fraction 혼합물에 대한 참다래 저장병원균의 균사생장억제 효과 조사 ... 60
• 3) 참다래 저장병원균에 대한 Eucalyptus darlympleana 추출액 butanol fraction의 항균 활성 검증 ... 62
• 3. Eucalyptus darlympleana 엽육 추출액에서 항균원인물질 화학적 구조동정 및 항균활성 검정 ... 64
• 1) Eucalyptus darlympleana 엽육 추출액에서 항균원인물질 화학적 구조동정 ... 64
• 2) Eucalyptus darlympleana 엽육 추출액에서 분리한 항균원인물질의 참다래 과실무름병균 억제효과 검증 ... 83
• 3) 참다래 저장병원균에 대한 Eucalyptus darlympleana 추출액 butanol fraction의 항균 활성 검증 ... 87
• 4. 천연항균성 물질을 이용한 환경친화형 참다래 저장병 방제기술 개발연구 결과 요약 ... 91
• 제3절 무공해적 참다래 저장병 물리적 방제기술 개발 ... 93
• 1. TiO2 광촉매를 이용한 참다래 저장병 방제 효과 검증 및 최적 처리조건 확립 ... 93
• 1) TiO2 광촉매 처리수를 이용한 살균시스템 제작 ... 93
• 2) TiO2 광촉매 최적 처리조건 확립 ... 97
• 3) 참다래 저장병균에 대한 TiO2 광촉매의 살균효과 ... 109
• 2. TiO2 광촉매와 오존 복합 처리를 위한 시제품 제작 및 참다래 저장병 방제 효과 ... 114
• 1) TiO2 광촉매와 오존 복합 처리 시스템 설계 및 시작품 제작 ... 114
• 2) 참다래 과실무름병균에 대한 TiO2 광촉매의 살균효과 ... 117
• 3) TiO2 광촉매 처리에 의한 살균제 flusilazole 분해효과 조사 ... 121
• 3. 참다래 과실무름병균에 대한 TiO2 광촉매와 오존복합처리장치 시제품을 이용한 현장적용 실험 ... 125
• 4. 무공해적 참다래 저장병 물리적 방제기술 개발 연구 결과 요약 ... 128
• 제4장 목표 달성도와 관련분야에의 기여도 ... 130
• 1. 연구개발목표 달성도 ... 130
• 2. 관련 분야에의 기여도 ... 131
• 제5장 연구개발결과의 활용계획 ... 135
• 제6장 연구개발과정에서 수집한 해외과학기술정보 ... 137
• 제7장 참고문헌 ... 139
• 끝페이지 ... 145