Utilizing Derivatives of Microbial Metabolites and ... - ACS Publications

Chapter 22

Utilizing Derivatives of Microbial Metabolites and Plant Defenses To Control Diseases 1

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Downloaded by LOUISIANA STATE UNIV on November 15, 2014 | http://pubs.acs.org Publication Date: March 12, 1993 | doi: 10.1021/bk-1993-0524.ch022

H. V. Morton and R. Nyfeler 1

Ciba-Geigy Corporation, P.O. Box 18300, Greensboro, NC 27419 Ciba-Geigy, Ltd., Basle, Switzerland 2

The phenylpyrroles are a new class of agricultural fungicides related to the natural antibiotic pyrrolnitrin, which is produced by Pseudomonas spp. Pyrrolnitrin fulfills the main requirements of natural products, namely it has interesting biological activity and possesses a simple enough structure to allow the synthesis of analogues. A second area offering a new approach to disease control is that of systemic acquired resistance (SAR) utilizing chemicals with no direct bactericidal or fungicidal effect. Agents which induce SAR include the rice blasticide probenazole and the immunization compound 2,6-dichloroisonicotinic acid and its ester.

Opportunities for Natural Products A question frequently heard today is: "What is the role of chemicals in the future of crop protection?" We view our business objectives as inseparable from our environmental objectives and thus integrated pest management (IPM) is one of our goals. These are fundamental elements that help drive trie discovery, development, production, and sale of our products. The following discussion illustrates the chal lenges and opportunities for (a) using natural products or their derivatives, and how these may be optimized, and (b) the use or chemicals to stimulate the host plant resistance mechanisms. Such strategies offer new approaches to disease con trol and a possible means to address the growing public concern for the environ ment. It should also be recognized that IPM win oe the primary strategy of pest control and that chemical crop protection remains an integral feature oiTPM. To maintain the value of chemicals in crop protection, the following four objectives must be fulfilled. 1. Continue and even accelerate the search for novel compounds which are effective at very low doses; are selective for the target organisms; show a moderate persistence in the environment; and are nontoxic to humans, nontarget organisms, and the environment 2. Strengthen the search for methods to delay or eliminate the induction and appearance of resistance to crop protection chemicals. 3. Improve application technology, concentrating on optimization of for mulations, placement of the products, and timing o f applications. 4. Extend and improve the Integrated Pest Management approach. This paper will address only the first point, namely the search for new chemicals. Such a search is influenced by:

0097-6156/93/0524-0316$06.00/0 © 1993 American Chemical Society In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.

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MORTON & NYFELER 1. 2.


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Microbial Metabolites and Plant Defenses

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The availability of relevant biological test systems. A battery of tests to investigate the fate of the compounds in the environment, and to study the metabolism in plants and animals and the toxicological properties. An ample supply of novel compounds.

The quality and relevance of these tests and the diversity of the chemical structures are the "key elements to finding safe, effective crop protection com pounds. While a review of crop protection chemistry shows that synthetic chemicals have clearly dominated the natural products, the main reasons to search for natural prod ucts as crop protection chemicals are: 1. Natural products provide an alternative source of new structures, 2. Since natural products are formed and degraded in nature, there is a reasonable chance they will fit IPM, and 3. An understanding of the role of secondary metabolites in natural pro cesses can lead to new crop protection principles. There are two approaches to searching for new secondary metabolites from plants and microorganisms: (a) random screening, and (b) utilizing a biorational approach, e.g., allelochemicals and induced or preformed metabolites which con tribute to the resistance of plants against diseases and insects. Typical examples of these would be phytoalexins or insect antifeedants. Many microorganisms that are evaluated as biocontrol agents owe their activity, at least in part, to secondary metabolites. Isolation anastructure determi nation of these active principles can result in new interesting lead structures. Table I shows an overview of the natural products which have gained some importance in crop protection. Secondary metabolites from microorganisms show a broader spectrum of activity than those derived from plants. A review of the 8th edition of "The Pesticide Manual" shows that there is a total of 14 natural products out of approximately 570 chemicals listed. Table T. The Direct I Jse of Natural Products in Crop Protection Active Against Plants

Microorganisms

Insects

Derived From Plants Nicotine Rotenone Pyrethrins Derived From Microorganisms Gibberellic Acid Bialaphos

Blasticidin S Cycloheximide Kasugamycin Pimancin Polyoxins B and D Vafidamycin Streptomycin

from

Delta-endotoxin Bacillus subtihs Avermectin B1

Japan has been a pioneer in finding uses for natural products in crop protec tion. The most widely used of these are shown in Table H

In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.

318

PEST CONTROL WITH ENHANCED ENVIRONMENTAL SAFETY


Table II.

Agricultural Antibiotics Used in Crop Protection in Japan

Name Blasticidins

Introduction 1962

Kasugamycin Polyoxins Validamycin Streptomycin

1965 1968 1972 1957

Produced By Streptomyces griseochrbmoeenes S. kasugaensis S. cacaoi S. hvgroscopicus S. griseus

Tons Sold In 1986 11 91 76 172 175

Reference: Noyaku Yoran (1) With the exception of streptomycin, all of these compounds have been dis covered and developed in Japan. It is worth noting that all of these antibiotics are produced by fermentation from Streptomyces species. One of the main reasons these products are not used outside of Japan is that the purity of the products, respective to the by-products, is not sufficient to satisfy the stringent western registration guidelines. EPA requires that any by-product that is present at more than 0.1% of the total amount has to be identified. The following is an example of a project CIBA-GEIGY undertook using a natural product as a lead structure. Pyrrolnitrin, a secondary metabolite from Pseudomonas pvrrocinia. was isolated by Arima. et al., in 1965 (2). This compound showed interesting antifungal activity and was developed as a topical antimycotic for human use. In the late 1970's pyrrolnitrin was found to be nighly active against a range of phytopathogenic fungi in our fungicide screen. The inter esting biological activity, combined with an apparently simple structure, made pyr rolnitrin an ideal lead structure for optimization (Figure 1).

I Figure 1. Pyrrolnitrin Despite the simple looking structure, the compound proved to be quite a challenge for the chemists. The best total synthesis of pyrrolnitrin still required seven steps and afforded the product in only 20% total yield from the reaction sequence (Figure 2). Fortunately, we have discovered a novel procedure to prepare new analogues of pyrrolnitrin. From readily available starting materials, the desired products were prepared in a single step and with good yields. Out of a large optimization program in which the substitutes X ana E (Figure 2) were broadly varied, there resulted a number of highly active fungicides belonging to the phenylpyrrole chemical class.


22. MORTON & NYFELER


319

(a) The Production of Pyrrolnitrin: (3) 7 Steps a' N02

Total Yield: 20%

N I

H


(b) Analogues with New Synthetic Method: (4)

E

+

Base Ether/DMSO

E = CN COOH Yield: 50-80% NO, Figure 2. Synthetic Approaches to Pyrrolnitrin Production H

C

3 ~^

^2S02NC ^—OK*

3

H

The phenylpyrrole fungicides are protectant fungicides with a broad spec trum of activity which allows them to be used as both seed treatments and foliar fungicides. Fenpiclonil (see Table IV) has recently been introduced as a cereal seed treatment in Europe. In the U.S., we are developing a more active analogue, CGA-173506 4-(2,2-difluoro-l, 3-benzodioxol-4-yl) pyrrole-3-carbonitrile (Figure 3, Table III). Chemical class: Chemical name:

Phenylpyrrole 4-(2,2-difluoro-1,3-benzodioxol-4-yl) pyrrole-3-carbonitrile

Structural formula:

Figure 3. CGA-173506: A new phenylpyrrole fungicide with broad-spectrum disease control


320

PEST CONTROL WITH ENHANCED ENVIRONMENTAL SAFETY


Table III.

In Vitro Spectrum of Activity of CGA-173506 E C 50 (ppm a.i.) Fungal Pathogen

Oomvcetes: >100 Phytophthora infestans >100 Pythium ultimum 10.3 Aphanomvces laevis Ascomvcetes: 0.18 Ophiobolus graminis 0.07 Monilini^ructicola 6.05 Venturia inaequalis Basidiomvcetes: 0.04 Rhizoctonia solani 0.10 Pellicularia~sasakii 0.22 Sclerotiumroltsii Deuteromvcetes: Helminthosporium teres 0.05 0.15 Alternarnia solani 0.02 Botrytis cinerea 0.18 Fusarium culmorum 0.20 Cercospora arachidTcola With its spectrum and degree of activity, protectant mode of action, and lim ited toxicity, CGA-173506 is a major breakthrough. The key to this optimization program was the marked increase in light stability of the cyanopyrroles (See Table Table TV. Modification of Pyrrolnitrin for improved Light Stability Pyrrolnitrin

Test Light stability T 1/2 (suntest- lamp)

27 min

48 hrs.

Apple: 6 ppm Bean: 200 ppm

6 ppm 6 ppm

4%

91%

Botrvtis activity Greenhouse E C 80 Field activity (100 g/hl)

E. Stamm, CIBA-GEIGY, unpublished


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MORTON & NYFELER


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New Crop Protection Principles Now we will turn to a somewhat more speculative approach: regulation of host parasite interactions, in particular the phenomenon of systemic acquired resistance (SAR) first described by Ross (5) on tobacco. In a variety of plant species, the development of necrotic lesions in response to pathogen infection leads to induction of generalized disease resistance in infected tissues. SAR is character ized by the development of a disease-resistant state in plants that have reacted hypersensitively to previous infection by tobacco mosaic virus. Salicylic acid has beenjim^icated as the endogenous signal that activates the resistant state Malamy At CIBA-GEIGY we have been trying to synthesize compounds which will induce local and systemic resistance in plants. 2,6-dichloroisonicotinic acid (CGA-41396) andits ester (CGA-41397) are examples of chemistries which induce host plant resistance (Figure 4). Ward, et at (7) have shown that these products also induce SAR gene expression by following nine classes of RNAs (by gel blot hybridization) that are coordinately induced concomitantly with the onset of SAR.

CGA-41396

CGA-41397

Figure 4. Chemical structures of isonicotinic acid derivatives The onset of SAR is characterized by a decrease in the size of lesions of foliar pathogens. This induction response takes about six days to develop in the case of T M V in tobacco as described by Ross (4). Isonicotinic acid was shown by Ward, et a l (7) to impact lesion size within four days and reached a maximum of steady-state RNA at six days after infection. Practical experience in field trials has shown these isonicotinic acids must be applied preventively to provide acceptable disease control. It is of interest to note that monocotyledonous plants express this resistance for longer than dicotyledon ous plants, which require applications every seven to ten days to maintain the expression of induced resistance. These products are effective against bacteria, fungi, and viruses, as well as Pyricularia oryzae (rice blast) via seed-treatment. The isonicotinic acid derivatives perform best against the powdery and downy mildews, while they are weaker against thymatrophic parasites (those that kill tis sues in advance of their own cells and then invade the Killed tissues) such as Rhizoctonia solani and Septoria spp. Unfortunately the amount of phytotoxicity associated with foliar sprays of CGA-41396 and CGA-41397 precludes their commercial development. CIBAGEIGY is currently working with a second generation of products which are safer to plants. There are several opportunities offered by products which result in SAR. One of these possibilities is for plant breeders to use such products to accentuate host plant resistance to plant pathogens. We view this chemistry as resulting in exciting new forms of disease control in the future. Literature Cited 1.

Noyaku Yoran, 1986 and 1988 published by Japan Plant Protection Association.

2. Arima, K; Imanaka, H; Kousaka, M ; Fukuda, A; Tamura, G. J. Antibiotics 1965, 18, 211-219.


322 3. D.

PEST CONTROL WITH ENHANCED ENVIRONMENTAL SAFETY Gosteli, J. Helv. Chim. Acta 1972, 55, 451-460.

4. Van Leusen, A . M ; Siderius, H; Hoogenboom, B. E ; van Leusen, Tettrahed. Letters 1972, 5337-5340. 5.

Ross, A. F. Virology 1961, 14, 340-358.

6.

Malamy, J; Carr, J. P; Klessig, D. F; and Raskin, I. Science 1990, 10021004.

7.

Ward, E . R; Uknes, S. J; Williams, S. C; Dincher, S. S; Winderhold, D. L ; Alexander. D. C; Ahl-Goy, P; Metraux, J. P; and Ryals, J. A . Plant Cell 1991, 3, 1005-1094.


RECEIVED September 1, 1992


Utilizing Derivatives of Microbial Metabolites and ... - ACS Publications

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