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Monitoring of Photodynamic Herbicides

#B-706


Tetrapyrrole-Dependent Photodynamic Herbicides and Production of Devices for Monitoring of Herbicide Action

Tech Area / Field

  • AGR-PPR/Plant Protection/Agriculture
  • BIO-CHM/Biochemistry/Biotechnology

Status
3 Approved without Funding

Registration date
23.03.2001

Leading Institute
National Academy of Sciences of the Republic of Belarus / Institute of Biophysics and Cell Engineering, Belarus, Minsk

Supporting institutes

  • Belarussian State University / Institute of Physical Chemical Problems, Belarus, Minsk

Collaborators

  • Institut fur Pflanzengenetic und Kulturpflanzenforschung, Germany, Gatersleben\nPhilipps Universität / Fachbereich Biologie/Botanik, Germany, Marburg\nLudwig-Maximilians-Universität / Botanisches Institut, Germany, Munich\nUniversity of Illinois / Department of Natural Resources and Environmental Sciences, USA, IL, Urbana

Project summary

The main purpose of the project is the elaboration of the conception of novel effective tetrapyrrole-dependent photodynamic herbicide (TDPH) design, the development of a new technology for estimation of herbicide activities and production of the computerized pilot devices for monitoring of TDPH action.

High priority of the project is due to substantial contribution of herbicides to modern plant production providing cost-effective increase in agricultural productivity. Without chemical control of weeds major crops yield penalties up to 40% and more. A new class of herbicides, peroxidizing or more exactly, TDPH, has become an attractive subject of interest of modern agrochemistry and academic science as to how they provide an excellent method of weed control and extremely low use rates ensure environmental safety [Peroxidizing Herbicides. Bouger and Wakabayashi (eds.), (1999)]. TDPH have a common mode of action i.e. inhibition of chlorophyll (Chl) biosynthesis through binding to their target enzymes and accumulation of cyclic tetrapyrroles that are strong photosensitizers. They cause light-induced formation of reactive oxygen species (ROS) which oxidize the photosynthetic pigments (photobleaching phenomenon), proteins and nucleic acids as well as cellular membranes by lipid peroxidation. The only commercially available TDPH are the inhibitors of protoporphyrinogen oxidase (Protox), for example such as oxyfluorfen, oxadiazone and the most commonly used acifluorfen. Inhibition of Protox results in accumulation of photodynamic protoporphyrin IX (Proto, up to 20 nmol/g fresh weight) and then ROS in cytosol. Although there is a high level of Protox inhibitor patenting (431 for 1986–1995), the market of TDPH is limited by the similar selectivities of most of these compounds and high toxicity of the inpidual TDPH. Design of commercial TDPH having different target enzymes in Chl route can results in synthesis of novel more environmentally safe and effective herbicides controlling another weed spectrum than Protox inhibitors thereby considerably extending their capabilities. Monitoring and screening of TDPH is based on comparative evaluation of biochemical characteristics of plants treated with inpidual TDPH, such as accumulation of ROS, peroxidation of cellular membrane lipids, responses of the most important cellular proteins on photooxidation triggered by herbicides. However biochemical examinations are very labor and time consuming, required expensive chemicals and skilled specialists. Development of a new simple technology for estimation of TDPH activities based on correlation between biochemical and physical characteristics of plants treated with TDPH, production of the devices for realization of the technology is very important and attractive task.

Many still experimental TDPH attacking other sites in Chl pathway have been developed. Two of them, glutafen, showing excellent activity against weeds in cereal crops, and cesium chloride, having only theoretical significance, are the most interesting. Partial inhibition of uroporphyrinogen decarboxylase (Urode) by cesium chloride in greening plants leads to an unique opportunity to over-produce photodynamic protochlorophyllide (Pchlide, up to 100 nmol/g fresh weight) resulting in strong plant photooxidation. Phenanthroline, a component of glutafen, activates 5-aminolevulinic acid (ALA) production, inhibits Mg-cyclase and pheophytinizes Chl a that result in accumulation of photodynamic Mg-Protoporphyrin IX, its monomethyl ester – Mg-Proto(E), (up to 30 nmol/g fresh weight) and pheophytin a (Pheo, up to 50 nmol/g fresh weight). Unlike Protox TDPH under action of cesium chloride and glutafen excess of tetrapyrroles accumulates and localizes exclusively in chloroplasts.

It is very important to carry out comparative study of experimental TDPH and commercial Protox inhibitors as prototype in relation to their herbicidal activity to detect the considerable promise herbicide targets in Chl pathway and work out a conception for design a novel effective TDPH. The main task of the project is an analysis of the important biochemical characteristics of weed plants under photooxidation triggered by acifluorfen, cesium chloride and glutafen. This work will be first executed.

Theoretically TDPH may be expected to be able to function only as non-selective compounds, because they act via Chl biosynthetic pathway common to all green plants. However, all of these compounds were found to have selective herbicidal effect. Selectivity mainly due to differences in antioxidative defense of inpidual plants to remove ROS formed during herbicide action. To cope with ROS higher plants have developed complex protection mechanisms acting through carotenoids, low molecular weight antioxidants such as ascorbate (AsA), glutathione (GSH), a-tocopherol (TP) and the main plant protected enzymes, ascorbate peroxidase (APX) and catalase (CAT), scavenged H2O2, superoxide dismutases (SOD), scavenged superoxide anions, and glutathione reductase (GR) restored the level of GSH [Oxidative Stress and the Molecular Biology of Antioxidant Defenses. J.Scandalios (eds.), 1997]. Under action of acifluorfen, cesium chloride and glutafen ROS are formed in different cell compartments contained specific antioxidative systems. It could result in differences in manifestation of plant resistance to inpidual TDPH. The important task of the project is an analysis of activity of antioxidative systems in crop plants and weeds under photooxidation triggered by above-mentioned TDPH.

The physiological response of plants sensitive to TDPH is manifested in a rapid wilting and browning of shoots, bleach out of the pigments and retardation of growth due to a strong interference of herbicides with biochemical processes in plant cells. The important task of the project is the development of the principal new technology for estimation of TDPH activities by registration of electrical capacitance of plants after herbicide application and establishing of the correlation between changes of electrical characteristics and the above-mentioned biochemical ones. The new technology will be based on the elaboration worked up by one of the participants of the project [Certificate of Authorship (1982) SU No1027526] and showed strong correlation between electrical capacitance of plants, evaluated with high precision, and such traditional plant characteristics as surface, volume and mass that are strongly changed after TDPH application. The new technology will first allow to exert an express control of crop plants and weeds after TDPH application by remote measurement of their electrical characteristics in all planed experiments. By application of the technology the estimation of TDPH activities, their monitoring during all time of plant vegetation will be carried out. The demonstration of the technology as well as production of the pilot devices for monitoring of TDPH action in laboratory tests, computerization of the devices, patent application with foreign collaborators will be realized. It is planned also to make the preliminary work for future introduction of the technology and the devices into practice.

The impact of the proposed project on the progress in the field:

– The conception of design of novel effective TDPH will allow considerably increase the list of commercial herbicides with a new spectrum of selectivity and more environmentally safe than Protox inhibitors.

– The technology and devices for TDPH monitoring will correspond to the level of the world’s standards. They will open the new market niches of devices monitored TDPH action on base of changing of electrical parameters of plants. Their implementation will save both time and labour for estimation of TDPH action, simplify their screening as well as monitoring doing the last available for all involved in plant production.

Participants of the project.

11 main researches involved with the project present two groups. The group of device production and TDPH monitoring (Research Institute for Physical Chemical Problems of the Belarusian State University – 5 scientists, 4 Ph. Dr.) will be engaged in the development of the technology of TDPH monitoring, production and computerization of the pilot devices for monitoring of TDPH action. It is consists of weapons scientists and engineers related to missile technologies. The group of biochemical analysis (Institute of Photobiology of the National Academy of Science of Belarus – 6 scientists, 2 doctors of sciences and 4 Ph. Dr.) will be engaged in the study of biological responses of plants to TDPH application and the development of the technology of TDPH monitoring. They are concerned with the study of the mechanisms of regulation of biosynthesis of Chl precursors, their localization, and native state in vivo, participation in photodamage of plants, activity of plant antioxidative system, design of the new TDPH. Both groups have a high qualification in their spheres documented by their publications (more than 450) and 47 CA.

Expected results.

Solving of 7 main tasks will allow to receive the following results:

1. The most active TDPH and the peculiarities of their phytotoxicity will be detected.

2. The most photosensitive enzymes of Chl biosynthesis pathway and peculiarities of their responses to photooxidation due to action of inpidual TDPH will be established.

3. The most photosensitive proteins of photosynthetic apparatus under action of inpidual TDPH will be detected.

4. The mechanisms of resistance of the crop plant species to acifluorfen, cesium chloride, and glutafen and the nature of the herbicide selectivity will be established.

5. The program and the methods of laboratory tests of the technology and pilot devices, monitored TDPH action, will be worked out.

6. The new technology for estimation of TDPH activities will be developed. Demonstration of the technology in laboratory tests will be carried out.

7. Computerized pilot devices for monitoring of TDPH action will be produced. Patent application for the devices with the foreign collaborators and demonstration of the devices will be realized.

Scientific significance of the project is in the development of the conception of a new effective TDPH design. Commercial and industrial significance of the project involves appearance in the future on the market novel commercial environmentally safe herbicides with a new spectrum of selectivity, production of the computerized pilot devices for monitoring of TDPH action in field, switch from pilot to batch-production and then to commercial production of the devices and their sale.

Application of the results.

The conception of new effective TDPH design, the technology and devices for monitoring of TDPH action, developed as results of the realization of the project, will find a broad application in research laboratories of plant protection and agriculture. A batch-production of devices is planned to be launched in Small State Enterprise “Design Bureau “Pribor”” having necessary basic equipment and specialists. With help of the foreign collaborators it is planned to find foreign partners to launch commercial production of the devices and introduce them to the market.

Meeting ISTC goals and objectives.

The project fully conforms to goals and objectives of ISTC and:

– provides the weapons scientists and engineers related to missile technologies opportunities to redirect their talents to peaceful activities;


– promotes integration of these scientists into the international scientific community;
– supports basic and applied researches, technology developments for peaceful purposes in fields of biotechnology and environmental monitoring;
– reinforces the transition to market-based economies and gives opportunity to weapons scientists and engineers to go to self-financing;
– contributes to the solution of national and international environmental problems.

Scope of activities.

Duration of the project is 30 months. Participants: Institute of Photobiology of the National Academy of Sciences of Belarus and Research Institute for Physical Chemical Problems of the Belarusian State University. Researches into the following 7 tasks are planned:

– Study of peroxidation of cellular lipids, generation of total ROS and hydrogen peroxide in weeds treated with acifluorfen, cesium chloride and glutafen.


– Detection of activities and content of the Chl biosynthesis enzymes in weeds treated with acifluorfen, cesium chloride and glutafen.
– Study of photodegradation of the main proteins of photosynthetic apparatus in weeds treated with acifluorfen, cesium chloride and glutafen.
– Tracing of the activity of the antioxidative systems in weed and crop plants treated with acifluorfen, cesium chloride and glutafen.
– Development of the program and the methods of laboratory tests of the technology and pilot devices monitored TDPH action.
– Development of the technology for estimation of TDPH activities based on measurement of electrical parameters of plants.
– Production of the computerized pilot devices for monitoring of TDPH action on weed and crop plants.

Role of foreign collaborators.

The long-term cooperation with lider scientists in related researches from Germany and Switzerland is planned. They have provided the consultations in the design of the project and in preparing of the work plan. The following activities are planned:

– information exchange in the course of the project implementation;


– providing of comments to the annual and final reports submitted by project participants to the ISTC;
– shared use of certain equipment, chemicals; performance of joint investigations;
– common testing and evaluation of technology and devices developed in the course of the project;
– participation in technical monitoring of project activities performed by ISTC staff;
– assistance and material support for project participants to join international meetings;
– conduction of joint seminars and workshops.

Technical approach and methodology.

As model test-systems two monocotyledon crop plants, barley (Hordeum vulgare L.) and maize (Zea Mays), sensitive to inpidual TDPH, and two dicotyledon weed species, chickweed (Stellaria media L.) and rape (Raphanus raphanistrum L.), resistant to TDPH, will be used. Seeds of weed plants will be collected in fields. Seedlings will be grown in vegetation vessels, then treated with TDPH, placed into darkness for accumulation of tetrapyrroles and then illuminated for development of photooxidation. Preliminary study of plant photooxidation under glutafen and cesium chloride action was done. Strong correlation between electrical capacitance of plants and plant surface, volume and mass that are strong changed after TDPH application, will be a base for the development of the technology for estimation of TDPH activities.


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