Insect hormones and their derivatives as insecticides

SESSION X INSECTICIDES BASED ON INSECT HORMONES Bull. Org. mond. Sandte 11971, 44, 381-389 Bull. Wld Hlth Org. Insect Hormones and Their Derivatives as Insecticides WILLIAM S. BOWERS' The hormonal control of moulting, reproduction, and diapause in insects has little or no relationship to any similar phenomena in other animals, and the hormones involved in these processes are unlike any known hormones of vertebrates. The availability ofpure chemicals with high biological activity has permitted an asto- nishing increase in research on insect hormones. At present, understanding of insect endocrinology is far too incomplete to justify much speculation about the possibility of using insect hormones as insecticides. However, the preliminary studies discussed in this paper give reason for hope, and the results justify further effort. Man's contest with insects for food and fibre has been on a nearly equal footing. In the past, the omnivorous characteristics and nomadic habits of man, together with a low population density, enabled him to cope with insect-mediated famine and pestilence. Today the nutritional needs of an expanding population, existing for the most part in urban concentrations, must be satisfied by the intensive cultivation or rearing of relatively few domesticated fruits, grains, and animals. This specialization has generated an abundance of high quality food but has also permitted maximum competition by insect pests. Cultural, biological, and chemical methods of insect control have become a major concern of the agricultural scientist. To date, the use of toxic chemicals has been the basis of our most successful efforts to control insects. However, the potential environmental hazards of many of these chemicals, together with the rapid development of resistance to them by insects, necessitates a more enlightened approach to chemical methods of control. If more satisfactory chemical insecticides are to be developed, they must, in addition to being effective, meet certain new criteria. They must not pose immediate or long-term hazards to human populations, domestic animals, or wildlife. Ideally their insecticidal effects should be specific-that is, they should act only against a given target pest. 1 Senior Insect Physiologist, Insect Physiology Laboratory, Entomology Research Division, Agricultural Research Ser- vice, US Department of Agriculture, Beltsville, Md., USA. HORMONAL REGULATION OF INSECT DEVELOPMENT Fig. 1 illustrates hormonal regulation of insect metamorphosis as exemplified by the life-cycle of the yellow mealworm, Tenebrio molitor L. Each immature moult (larva to larva) takes place in the presence of the moulting hormones (ecdysones) and the juvenile hormones. The ecdysones are necessary for all moulting to occur, but the juve- Fig. 1 Hormonal regulation of insect development * Ad 02CDOQo ADUJLT LARVAE X~PUPA * JH = juvenile hormone; MH = moulting hormone 2651 381- W. S. BOWERS Fig. 2 Structure of juvenile hormones * K 'N 'N H20H "N0 C'N H20CH3 'N 'N 'N H2N(C2H5 N H20H /\ A f ~~H20CH13 Mr 0 / OOCH3 VI - H20C H2CH20C H2CH20C4Hg K H2CH2CH3 KIU -0C H(OC H2C H2)2OC H2 C H3 0 P OC H2C!CH OCH2CH2CH3 K --CH20 / 0-- C°X-X\\0 0v / OOCH3 / OOCH3 IO / \ OOCH3 X X - ~~X'CI,Brx I~ ~ ~ ~ V r OOCKH3 o / n XXI 0 K ~~~~~xxn 0 0 / 0 CH302C WHO 10235 * References: compounds l-lll, Schmialek (1961); compounds IV & V, Bowers (1963); compound VI, Bowers (1965); compound VII,'Bowers (1966); compound Vil, Cerny (1967); compounds IX & X, Slama et al. (1968); compound XI, Romanuk (1967); compounds XII, Roller (1967); compound XIII, Meyer et al. (1968); compounds XIV-XVIII, Bowers (1968); compounds XIX- XXIII, Bowers (1969). 382 INSECT HORMONES AND THEIR DERIVATIVES AS INSECTICIDES nile hormones are present only when the genetic "6programming" of the insect requires growth without maturation or differentiation. Thus, juvenile hor- mones (JH) prevent metamorphosis during immature life. When the insect reaches the point of metamor- phosis to the adult reproductive stage, it moults to a pupal form and finally to the adult. Both of these moults must take place in the absence of JH. The original idea of a hormone-based insecticide was founded on the discovery of Williams (1956) that treatment of the pupal stage of an insect with a hormone extract would disrupt morphogenesis during the pupa-adult moult and produce an inter- mediate form of insect incapable of survival. Although additional lethal effects of JH on insects have been discovered, the original idea of disrupting adult development by treatment with JH remains valid and is still the basis of most biological assays for these hormones. CHEMISTRY OF JUVENILE HORMONES The first chemical discovered to have JH activity was famesol (I, Fig. 2), which was extracted from the faeces of Tenebrio by Schmialek (1961). Subse- quently, the methyl ether (II) and the diethyl amine (III) of farnesol were found to have greater biolo- gical activity than the parent alcohol (Schmialek, 1963). In our laboratory we began structural modification work and found that while hexahydro- farnesol (IV) and its methyl ether were active, several simple saturated alcohols and their methyl ethers were active, especially dodecyl methyl ether (V) (Bowers & Thompson, 1963). Continuing these studies, we determined sufficient of the chemical and biological properties of the JH in cecropia extract to permit the synthesis of trans,trans-10,11- epoxy methyl farnesenate (VI), which we discovered to be extremely active in all of our biological assay systems (Bowers et al., 1965) and which we feel predicted the structure of the natural cecropia juvenile hormones. Following the discovery of JH activity in conifers (Slama & Williams, 1965) we isolated from the balsam fir and identified a monocyclic sesquiterpenoid compound (VII), which we called juvabione (Bowers et al., 1966), and which shows a relatively high specific JH activity only against members of the Pyrrhocoridae, an insect family that contains important pests such as the cotton stainers. Subsequently, other investigators (Cerny et al., 1967; Slama et al., 1968) isolated Fig. 3 Juvenile-hormone-active para-substituted aromatic terpenoid ethers R .o R' R=CH3,CH2C I3 R -C H31C H2 C H3 C)_-1 iorCOOCH3 another naturally occurring analogue of juvabione (VIII) and synthesized several aromatic analogues (IX, X) with similar activity for these insects. Com- pound XI was isolated (Romanuk et al., 1967) from a hydrochlorination reaction mixture (Law et al., 1966) and was found to be selectively active in the nanogram range against pyrrhocorids. Roller et al. (1967) characterized one of the natural juve- nile hormones from the cecropia moth (XII) and Meyer et al. (1968) characterized a second homo- logous juvenile hormone from the same moth. During an investigation of the possibility of increasing the activity of JH compounds with conventional insecticide synergists, we found that several synergists-such as piperonyl butoxide (XIV), sesamex (XV), and Niagara 16388 (XVI)-possessed appreciable JH activity. Since the chemistry of the synergists could in no way be related to the previously known active terpenoids, we combined chemical features of the synergists with the terpenoids to see if the JH activity could be increased. The aromatic ethers of epoxy farnesol were prepared (XVII, XVIII) and were discovered to be about as active as the synergists (Bowers, 1968). However, when the terpenoid portion was shortened by one isoprene unit (sesamyl geranyl ether epoxide, XIX) the activity was in- creased tremendously. Working on the hypothesis that the ethyl branches of the cecropia hormones might result in increased activity, we prepared compounds XX, XXI, and XXII. These " hybrid" combinations proved to be the most active of all the compounds studied and in our biological assays they were uniformly active at subnanogram levels (Bowers, 1969). Several non-methylenedioxy aromatic derivatives were prepared and were found to possess relatively high JH activity. Fig. 3 shows the structures of several para-substituted aromatic geranyl ethers with significantly high juvenile hormone activity. 2 383 W. S. BOWERS LETHAL EFFECTS OF INSECT HORMONES It is clear that the presence or absence of JH at certain critical periods of insect development pro- foundly affects the direction of this development. During the moulting process the presence of JH prevents cellular differentiation and, hence, matu- ration. In the absence of JH morphogenesis and maturation proceed towards the adult stage. The application of a JH chemical to a stage of develop- ment that is due to undergo maturation results in the formation of an intermediate that is incapable of further development and dies. Therefore, the classical notion of a JH insecticide was based simply upon supplying the hormone to the insect at a time when it must normally be absent. One of the earliest proposals for the use of JH as an insecticide was made by Williams (1956), who sug- gested that it be applied to insects in the pupal stage, a procedure that results in the formation of a pupa-adult intermediate. Many of the biological assays for JH are based on this phenomenon- for example, the Tenebrio genitalia assay developed in our laboratory is based on the retention of pupal genitalia in an otherwise normal adult beetle following the topical application of the candidate hormonal compound to the abdomen of the pupa. For ultimate sensitivity, the hormone treatment must be applied during the early phase of pupal development, since JH chemicals appear to exert their effects by preventing the expression of genetic information dealing with metamorphosis. Even large doses of JH cannot reverse differentiation. Therefore, in anticipation of the use of a JH chemical for control purposes the time of application must be given careful consideration. If JH chemicals are to be used for control purposes, it might be best to restrict them, at least initially, to situations in which they can be applied during the most sensitive periods in the insect's development or in which continuotls Table 1 Effect of juvenile hormone (JH) on the confused flour beetle Concentration of JH required to kill 100 % of the insects Compound Topical Concentration application a in diet b (ng/pupa) (ppm) o -HOOCH3 500 1000 0 C OOCH3 250 100 -0 <N S30 100 10 /:t/ 10 1 <0 ,- 0 / 1 0.1-1.0 a Pupa-adult intermediates or second pupae produced. b No normal adults produced. 384 INSECT HORMONES AND THEIR DERIVATIVES AS INSECTICIDES contact throughout the life cycle is possible. Where continuous contact is possible by the use of resi- dual films, baits, or fumigation the maximum lethal effect can be obtained. From the theoretical point of view, some obvious applications would be the treatment of stored grain, of mosquito and housefly breeding sites, and of soil against soil insects. Even field applications might be effective where elimi- nation of the first generation of a multiple-brooded pest would avoid the production of subsequent generations that would have more seriotus economic consequences. To illustrate the activity of a range of JH lhormonlal chemicals, Table I lists the lethal doses and con- centrations of these compounds for a serious stored-grain pest, the confused flouLr beetle (Lin- published data, this laboratory). Table 2 gives similar data for two imliportanlt lepidopterous pests of stored grain the Indian meal moth, Plodiai interpulictella, and the tobacco moth, Ephistia elitella) (unpublished data, this laboratory). It should be recognized that the dietary concen- trations of compounds shown in Tables I and 2 are the effective concentrations required to kill 100l% of the insects without delaying pupation. Higher concentrations, although ultimately lethal, prolong larval life, resulting in increased feeding and consequently greater damage. At the same time it should be noted that an approximately 10-fold increase in the effective concentration is required to produce an extension of the larval feeding period, so that rather gross formulation errors would be necessary to do serious damage. From the results of these tests it is apparent that the methylenedioxy aromatic terpenoid ethers are more active than the cecropia JH, and this has consistently been our experience with other species of economic importance, such as the Mexican bean beetle, the Colorado potato beetle, the cigarette beetle, and the black carpet beetle. The effects of these highly active "hybrids" compounds are shown in Table 3. The housefly, the milkweed bug, and the yellow mealworm were treated topically during their most sensitive stages- namely, the pupal or last nymphal instar. The hornworms and earworms were fed treated diets throughout larval life, while the yellow fever mosquito larvae (4edes aegypti) were exposed as third and fourth instar larvae by incorporation of the compound into the aqueous larval mediunm (unpublished data, this laboratory). ble 2 Effect of juvenile hormone (JH) compounds on the Indian meal moth and the tobacco moth Concentration of JH in the diet (ppm) required to kill 1 00 % of the insects Compound without delaying pupationa Indian meal moth Tobacco moth OOCH3 0 OQtOC CH3 4de~07 0 10-100 10 10-100 <1.0 10-100 91_ .0 a No normal adults produced. 385 W. S. BOWERS Table 3 Effect of sesamyl geranyl ether epoxide homologues on insects Insect and stage Dose orconcentration Effect housefly pupae milkweed bug nymphs Tenebrio pupae hornworm larvae corn earworm larvae mosquito larvae 0.5 ipg 1 0.0 ng 0.5 ng 0.1 ppm 0.5 ppm 0.1 ppm The lethal morphogenetic activity of these syn- thetic hormones is clearly effective against insects of different orders at dosage levels equal to or below those of conventional poisons. However, their effective use will require a more perceptive approach to methods of application than is necessary with present control methods. pupa-adult intermediates nymph-adult intermediates pupa-adult intermediates no pupation no adult emergence no adult emergence The fumigant action of JH chemicals (Bowers, 1969) is now known to be effective against numerous species, including the yellow mealworm, the con- fused flour beetle, the Mexican bean beetle, the tobacco hornworm, the Indian meal moth, the cotton stainer, and fruit flies. The concentrations shown in Table 4 for lethal effects on mealworms Table 4 Lethal effects of the fumigant action of juvenile hormone (JH) compounds on pupae of the yellow mealworm * Compound Amount of compound (9g)c oplaced in a 2 x 9-cm Petri dish o OOCH3 3t0 IOOCH 70° 1.0 10.0 1.0 0.1 * The effects were the production of pupa-adult intermediates or second pupae, which died. 386 INSECT HORMONES AND THEIR DERIVATIVES AS INSECTICIDES 387 Table 5 Percentage hatch of Mexican bean beetle eggs following topical treatment Percentage hatch after treatment with the following concentrations Compound (ppm)a 1 000 100 10 1 0 0 0 / 0 ~~ ~~ ~~~~~~~02 76 K <o/ 0- 0 3 33 o 0 <0 / - 1 4 41 K a Exposure by dipping in acetone-water (3-1) test solution for 5 seconds (Walker & Bowers, 1970). Table 6 Percentage hatch of Mexican bean beetle eggs following fumigation* Percentage hatch after exposure to the vapour from the following Compound amounts of compound (Ag)placed in a 2 x 9-cm Petri dish 100 10 1 0.1 0 0~~ Karathion 0 0 0 2 72 73 0-"' K 0 ~~ ~~ ~~~ ~~~~~0036 76 0 0 Kil / 05 24 78 K parathion - 0 1 79 * Walker & Bowers (1970). W. S. BOWERS by fumigant action are, with slight differences, valid for all the insects mentioned above. The period of embryogenesis is another develop- mental stage that is susceptible to the application of a JH chemical (Riddiford & Williams, 1967; Slama & Williams, 1966; Walker & Bowers, 1970). Although very little is known about the normal endocrine processes that occur within the insect egg, the application of juvenile hormones by contact or fumigation is lethal to the embryo. Tables 5 and 6 show the effect of several methylenedioxy " hybrid " compounds on the eggs of the Mexican bean beetle by dipping and by fumigation (Walker & Bowers, 1970). In Table 6 the effectiveness of these chemicals is compared with that of parathion, which is known to be an effective ovicidal fumigant for this species. The lethal action of JH compounds on insect eggs significantly extends the usefulness of such chemi- cals when an insect population of mixed ages is to be treated. Direct contact of the chemical with the eggs of several insect species is unnecessary, since the treatment of gravid females is sufficient to prevent the hatch of eggs produced subsequently (Riddiford & Williams, 1967; Slama & Williams, 1966). Numerous other aspects of insect biology that are subject to control by JH offer possibilities for pest-control applications. Many adult insects that enter diapause in order to survive periods of envi- ronmental stress such as winter or severe heat or drought can be brought out of diapause by treat- ment with JH compounds (Bowers & Blickenstaff, 1966; Connin et al., 1967). Without the physiological protection of diapause, few insects might be expected to survive. Juvenile hormones also regulate lipid metabolism, the production of sex pheromones, and development of the ovaries. Further elucidation of juvenile hormone chemistry, the isolation and identification of other natural juvenile hormones from additional insect species to serve as molecular models, and an understanding of the biosynthesis and metabolism of these hormones will undoubtedly increase their usefulness. Any serious discussion of the use of hormones for insect control must not omit consideration of the insect moulting hormones, whose biological activities are equally interesting and potentially use- ful. Three naturally occurring insect hormones have been identified (Horn, 1970); their structures are shown in Fig. 4. The potentially useful features of these polyhydroxy steroids include the fact that they Fig. 4 Natural insect ecdysones OH a-ecdysone 20-hydroxyecdysone 20,26-dihydroxyecdysone R' = Rp" = H R'= OH, R"= H R'= R" = OH Fig. 5 Synthetic ecdysone analogue, 22,25-bisdeoxyecdysone Table 7 Concentrations of 22,25-bisdeoxyecdysone in the diet or medium required to give approximately 75 % inhibition in the assay system * Assay system Concentration Larval development confused flour beetle German cockroach housefly yellow fever mosquito Reproduction housefly ovarian development housefly reproduction confused flour beetle reproduction 500-750 ppm 150-300 ppm 15-25 ppm 20-30 x 10-3 ppm 0.10-0.15 % 0.05-0.10 % 0.50-0.75 % * Robbins et al. (1970) 388 INSECT HORMONES AND THEIR DERIVAT[VES AS INSECTICIDES 389 interfere with immature moulting, ovarian develop- ment, embryogenesis, and diapause. Although the chemistry of the natural ecdysones is somewhat complex, it has been possible to prepare simpler synthetic analogues, which are much more potent inhibitors. One of these, 22,25-bisdeoxyecdysone (Fig. 5), inhibits immature development, reproduc- tion, and diapause in a wide range of insect pests. Table 7 indicates the effective range of concentrations of this ecdysone analogue when incorporated into the diet or medium (Robbins et al., 1970). Unlike JH, which exerts its lethal morphogenetic activity during the penultimate and ultimate moults, the ecdysones are able to interfere with the earlier moults of several insects. Reproduction is inhibited in two ways, dependent upon dosage. High dosages interfere with ovarian development, the effects being irreversible, while lower concentrations bring about reduced oviposition accompanied by low egg via- bility. CONCLUSION To enjoy biological success all living organisms must feed, mate, and reproduce. The fundamental biochemical processes underlying these require- ments have changed little in about 2 thousand million years. Consequently, the development of insecticides having specificity of action against insects must take advantage of alternative biological pro- cesses that have resulted from divergent evolution. In insects the most clearly recognized biological " innovations " are the hormonal control mecha- nisms. Insect hormones appear to be unique regu- lators of such vital processes as moulting, repro- duction, and diapause and as metabolic regulators they have no counterpart in higher animals. The high biological activity and specificity of these hor- mones, together with their low toxicity for higher animals, hold promise for their ultimate utilization for insect control. REFERENCES Bowers, W. S. (1968) Science, 161, 895 Bowers, W. S. (1969) Science, 164, 323 Bowers, W. S. & Blickenstaff, C. C. (1966) Science, 154, 1673 Bowers, W. S., Fales, H. M., Thompson, M. J. & Uebel, E. C. (1966) Science, 154, 1020 Bowers, W. S. & Thompson, M. J. (1963) Science, 142, 1469 Bowers, W. S., Thompson, M. J. & Uebel, E. C. (1965) Life Sci., 4, 2323 Cerny, V., Dolejs, L., Labler, L., Sorm, F. & Slama, K. (1967) Colln Czech. cheni. Commnun. Engl. Edn, 32, 3926 Connin, R. V., Jantz, 0. K. & Bowers, W. S. (1967) J. econ. Ent., 60, 1752 Horn, D. H. S. (1970) In: Jacobson, M. & Crosby, D. G., ed., Naturally occurring insecticides, New York, Dekker Law, J. H., Yuan, C. & Williams, C. M. (1966) Proc. nat. Acad. Sci. (Wash.), 55, 576 Meyer, A. S., Schneiderman, H. A., Hanzman, E. & Ko, J. H. 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