가시박(Sicyos angulatus L.)은 기존의 자연식생을 교란시켜 자연과 농업생태계를 위협하는 교란종으로 2009년 환경부에 의해 생태계교란 야생식물로 지정되어 방제를 위한 연구가 진행되어오고 있다. 본 연구는 가시박 종자 내에서 발아억제를 유도하는 화학물질을 찾아내고 그 천연물질을 이용하여 작물의 생육조절 활성을 알아보았으며 또한 이를 이용한 친환경 제초제 개발 연구를 위해 본 실험을 진행하였다. 연구에 사용된 가시박은 경상북도 구미시 해평면(낙동강)과 영주시 평온면(내성천) 일원에서 총 4회 시료를 채취한 뒤 동결건조하여 보관하였으며, 이 가시박 종자를 물과 함께 분쇄하여 10%, 20%의 농도로 상추에 처리하였다. 그 결과 초장, 엽수, 엽폭, 엽장 등이 무처리구에 비해 가시박 종자 추출물(이하 BSE) 처리구에서 큰 폭으로 감소됨을 확인하였다. 또한 BSE 20%에서 페놀계 물질이 다량 함유 되어 있음을 확인하였고 이로 인해 상추의 생체량 또한 큰 폭으로 감소되었으리라 사료된다. 또한 BSE를 TLC와 column chromatography를 통해 여러 차례 분리, 정제하여 GC-MS를 이용하여 물질 동정을 진행한 결과 2-linoleoyl glycerol이 억제활성을 가진 물질임을 확인하였다. 상추 생육억제 기작을 확인하기 위하여 2-dimensional electrophoresis (2-DE)와 LC-MS/MS를 이용하여 단백질 동정을 진행하였고, 총 57 spots이 확인되었으며, 그 중 39 spots은 무처리구에 비해 20% BSE 처리구에 비해 발현량이 감소하였으며, 18 spots은 발현량이 증가함을 확인하였다. 그 중 chlorophyll a/b binding protein and oxygen-evolving enhancer protein 1등이 무처리구에 비해 BSE 20% 처리구에서 발현량이 크게 감소함을 확인하였고, 이는 광합성 색소 합성이 감소함을 의미하고 이러한 이유로 상추의 생육이 억제되었을 것이라 판단된다. 실제로 BSE처리구의 gibberellins의 합성량이 감소하였고, 스트레스 관련 호르몬인 abscisic acid, salicylic acid, jasmonic acid의 합성량이 증가함을 확인하였다. 필수 원소인 Ca, Fe, Mg, K, S, Mo의 흡수량은 무처리구가 많았고, Cu, Zn, Na의 양은 BSE처리구에서 많음을 확인하였다. 본 연구의 결과는 BSE는 잡초의 생육을 억제하고 광합성 기작을 억제하며, 호르몬과 영양적인 변화를 조절함으로써 친환경 제초제로서의 연구 가능성을 확인하였다.
Bioherbicides from plant extracts are an effective and environmentally friendly method to prevent weed growth. The present investigation was aimed at determining the inhibitory effect of bur cucumber seed extracts (BSE) on lettuce plant growth. Bur cucumber seeds were ground with water, and two different concentrations of seed extracts (10% and 20%) were prepared and applied to lettuce plants. Decreased plant height, number of leaves, leaf length, leaf width, an Prod. Type: FTPd leaf area were found in lettuce exposed to BSE as compared with controls. A significant reduction in lettuce biomass was observed in 20% BSE-treated plants due to the presence of higher amounts of phenolic content in the extracts. Moreover, a significant inhibitory chemical, 2-linoleoyl glycerol, was identified in BSE extracts. The mechanism of plant growth inhibition was assayed in lettuce proteins by 2-dimensional electrophoresis (2-DE) and the LC-MS/MS method. In total, 57 protein spots were detected in plants treated with 20% BSE and control plants. Among these, 39 proteins were down-regulated and 18 proteins were upregulated in plants exposed to 20% BSE as compared with controls. The presence of low levels of chlorophyll a/b binding protein and oxygen-evolving enhancer protein 1 in BSE-exposed plants reduced photosynthetic pigment synthesis and might be a reason for stunted plant growth. Indeed, the plantgrowth stimulating hormone gibberellin was inhibited, and synthesis of stress hormones such as abscisic acid, jasmonic acid, and salicylic acid were triggered in lettuce by the effects of BSE. Uptake of essential nutrients, Ca, Fe, Mg, K, S, and Mo, was deficient and accumulation of the toxic ions Cu, Zn, and Na was higher in BSE-treated plants. The results of this study suggest that extracts of bur cucumber seeds can be an effective eco-friendly bioherbicide for weed control that work by inhibiting mechanisms of photosynthesis and regulating phytohormones and nutritional elements.
Abstract iii1. Introduction2. Materials and Methods2.1. Bur cucumber extract preparation and seed germination2.2. Bur cucumber extract and lettuce plant growth2.3. Isolation and identification of proteins2.3.1. Protein sample preparation2.3.2. Two-Dimensional gel electrophoresis2.3.3. Image and data analysis2.3.4. In-gel protein digestion2.3.5. Identification of proteins by LC-MS/MS2.4. Identification of phytohormones2.4.1. GA extraction and quantification2.4.2. Determination of absicsic acid (ABA)2.4.3. Determination of salicylic acid (SA)2.4.4. Determination of jasmonic acid (JA)2.5. Nutrient analysis2.6. Total phenolic compound content in BSE2.7. Identification of plant growth inhibiting chemical from BSE2.8. Statistical analysis3. Results and Discussion3.1. Phytotoxic effect of bur cucumber on plant growth3.2. Inhibition of photosynthetic mechanisms in plants3.3. Hormonal control in plant growth3.4. BS- induced regulation of nutrient uptake by lettuce4. Conclusion5. ReferenceAbstract (in Korean)