Allelopathy and the Secret Life of Ailanthus altissima - Arnoldia

Allelopathy and the Secret Life of Ailanthus altissima - Arnoldia

Allelopathy and the Secret Life of Ailanthus altissima " Rod M. Heisey - Although the reputation of the tree-of-heaven as an ornamental has declin...

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Allelopathy and the Secret Life of Ailanthus altissima

"

Rod M. Heisey

-

Although the reputation of the tree-of-heaven as an ornamental has declined over the past century, investigations now underway may discover a new role for the species

as

the

source

of

a

natural herbicide.

Ailanthus altissima (Simaroubaceae) has been extremely successful in invading and dominating certain habitats since its introduction to the United States in 1784. In parts of the northeastern United States, especially in southern Connecticut, southern Pennsylvania, and the lower Hudson Valley of New York, A. altissimaforms nearly pure stands that are resistant to invasion by other tree species.’ A number of characteristics contribute to the invasiveness and success of Ailanthus altissima, often called tree-of-heaven. First, the versatility of its reproduction methods provides a decided advantage. The trees regularly produce large crops of winged seeds that are widely dispersed by the wind. A. altissima can also spread rapidly by sprouting from stumps or from its wide-ranging lateral roots, particularly in openings or at the edges of forested areas. Another of its advantageous characteristics is the extremely rapid growth rate that enables it to outcompete many other species, especially when reproducing from root or stump sprouts. Average heights reported for one-year-old trees in south and central Pennsylvania were 1.3 feet for seedlings, 2.7 feet for root sprouts, and 6.0 feet for stump sprouts; two-year-old trees averaged 3.9 feet, 5.6 feet, and 9.2 feet, respectively.2 These rates make A. altissima one of the fastest-growing trees in the temperate zone.

Allelopathy: The Secret Weapon? Another contributor to the invasiveness and success of Ailanthus altissima may be a secondary metabolite that provides competitive superiority through a process known as "allel-

opathy." Many plants produce chemical compounds that have no apparent role in life processes or plant structure; hence, these compounds are called secondary metabolites. As techniques for identifying naturally produced chemicals have improved in recent decades, it has become apparent that plants manufacture a great diversity of secondary metabolites, including terpenoids, alkaloids, glycosides, flavonoids, coumarins, quinones, saponins, and phenolic compounds. Humans have found a variety of uses for some of these compounds, including menthol, a terpenoid produced by mint; nicotine, an alkaloid produced by tobacco; caffeme, an alkaloid produced by the coffee plant and other species; and salicin, a phenolic compound having analgesic properties, from the willow tree. Why do plants produce secondary metabolites ? An early hypothesis suggesting that they were simply waste products of normal metabolism has been largely discounted, since it does not explain the wide variety of secondary metabolites. Much evidence now indicates that some of these allelochemicals, as they are termed, play a defensive role for the producer organism, protecting plants from herbivores by making the plant tissues toxic, perhaps, or by reducing their palatability.3 Other compounds have antimicrobial effects and may protect plants from invasion by pathogens. Another role that secondary metabolites may play is that of allelopathy (Greek, allelo-, of one another; patheia, suffering), the inhibition of one plant’s growth by another through the production and release of toxic chemicals into the environment. Many secondary metabolites have

29

A

typical stand of Ailanthus altissima showing a

sparse

been shown to inhibit seed germination or plant growth in laboratory tests, and entire books have been written attributing a broad range of effects to allelopathy;4 however, some researchers question how widespread or important it really is in natural habitats.~ Two examples where the argument for allelopathy seems most convincing are the purple sage (Salvia leucophylla) in the coastal sage scrub community of California and the black walnut (Juglans nigra) in the eastern Umted States.~ But in fact, allelopathy has not yet been proven to exist in any plant to all researchers’ satisfaction.

Testing for Allelopathic Effects My research has focused on two areas: ( 1)determining whether Ailanthus altissima actually is allelopathic under natural conditions, and (2) evaluating the potential of its secondary metabolite as a natural herbicide. Members of the Simaroubaceae, mcluding Ailanthus, produce a class of bitter-tasting secondary metabolites called quassinoids, which exhibit a wide range of biological activity

understory with scattered Ailanthus root sprouts

including negative effects on insects, fungi, protozoa, viruses, and cancer cells.’ In China A. altissima has long been used as medicine and as insect repellent.~ The first publications on allelopathy by A. altissima were by Mergen (1959) and Voigt and Mergen ( 1962), who reported that water extracts of foliage and stems were injurious to tree seedlings of other species. The major phytotoxic compound produced by A. altissima was recently identified as a quassinoid compound called ailanthone.9 A major tool for research on allelopathy is the bioassay, a test that allows us to isolate phytotoxic compounds and quantify their effects under controlled laboratory conditions. A good bioassay should possess high sensitivity, give reproducible results, and take a relatively short time to perform. I usually use seeds of garden cress (Lepidium sativum) for bioassays, because they germinate rapidly and are very sensitive to phytotoxins. A basic bioassay involves placing garden cress seeds on filter paper in petri dishes, treating them with plant extracts, and then incubating them under standard conditions. At

30 A

Chemical structure of ailanthone (molecular weight 376), the phytotoxic compound produced by Ailanthus altissima. Allanthone is extremely bitter and belongs to the class of compounds called quassmoids. the end of the incubation period, the growth of the radicle (the initial root formation) of the

treated

seedlings is compared to that of control seedlings that received only deionized or distilled

deliver biologically effective amounts of the toxin from Ailanthus altissima trees to nearby soil. Stemflow collars were placed around A. altissima trunks, and rain was collected as it flowed down the trees. At the same time, precipitation was collected m open areas nearby to serve as a control. The water samples were then tested using the cress seed bioassay. Surprisingly, stemflow stimulated more cress radicle growth than either control precipitation or deionized water.’1 In retrospect this result was not unreasonable. The outer bark of A. altissima (low in ailanthone) probably prevents the toxin from being leached from the highailanthone inner bark in large enough quantities to inhibit plant growth; the bark may also contribute inorganic nutrients or growth hormones to the stemflow, thereby offsetting the effect of any ailanthone that does reach the soil. In any case, the results certainly did not support stemflow as a mechanism responsible for allelopathy under natural conditions.

water.

In order to learn whether ailan-

thone could indeed serve as an allelopathic agent for Ailanthus altissima, I first needed to find out where in the tree the phytotoxic compound is found. This was important because the location helps determine the quantity of toxin released into the soil as well as the release mechanism. I

began by assaying

water

of different A. altissima tissues, using the method described above. Phytotoxic effects were highest for the inner bark of the trunk and the bark of roots and branches, intermediate for leaves, and lowest for the thin outer bark of the trunk and for the wood of the trunk and roots.l° These results suggested two possible release mechanisms for the phytotoxin: (1) extraction of toxin from bark and foliage by rain, followed by stemflow down branches and trunks; or (2) exudation from roots. Tests were designed to learn which of these mechanisms, if either, could extracts

water extracts of Ailanthus altissima root bark on of barnyard grass (BYGR, Echinochloa crusgalli), corn (Zea mays), and garden cress (Lepidum satmum/. The seeds were moistened with (left to mghtJ deiomzed water for control or a water extract of A. altissima root bark correspondmg to 1 gram of bark m 5000 and 500 milhhters, respectmely. Both concentrations of extract caused considerable mhlbition of radicle growth of all three species compared to the control.

Bioassay of seeds

31

Effect of addmg Ailanthus altissima leaflets and root bark to soil on growth of garden c,ress The pots rec.emed (left to mght) no bark (=control); 2 grams of leaflets and 2 grams of root bark from which ailanthone had been extracted with methanol, and 0.5, 1, and 2 grams of non-extracted root bark The non-extracted bark caused obmous inhibition of cress growth, but bark from which the allanthone was removed with methanol stimulated growth compared to the control contammg no bark. The leaflets reduced cress growth shghtly, butt much less than the bark.

To test whether significant amounts of ailanthone could be released by Ailanthus altissima roots, some roots were added to soil m petri dishes. Control dishes contained identical soil, but no roots. The dishes containing roots were stored m a refrigerator (to retard degradation of the toxin by soil microorganisms) for 6 or 13 days to provide time for the toxin from the roots to exude into the soil. The dishes were then removed from refrigeration, seeds of garden cress were placed on the soil near the roots, and radicle growth was measured 3 days later. The results showed that significant amounts of ailanthone had been released from the roots. Exposure of the soil for 6 days to fine roots (less than three millimeters in diameter/ reduced cress radicle growth to 50 percent of that in the control soil, and exposure to larger roots (five to ten millimeters in diameter) reduced cress radicle growth to 74 percent. In soil stored for 13 days, cress radicle growth was reduced by exposure to fine roots to 33 percent of growth in the control soil and to 74 percent by exposure to

larger roots. In another test Ailanthus altissima root bark and leaves were added separately to soil in pots and the effect measured on garden cress seeds. Root bark strongly inhibited growth of the seeds, whereas leaves had a much weaker effect. Dry root bark added in quantities of 0.2, 0.4, and

0.8 percent of dry soil weight reduced cress seedling emergence to 39, 21, and 5 percent respec-

that m pots containing no root bark; shoot biomass was reduced to 55, 25, and 5 percent of biomass in the control pots. Dry leaflets added to soil m similar quantities only reduced cress emergence to 94, 88, and 93 percent of that in control pots and shoot biomass to 81, 81, and 64 percent. These results support the hypothesis that exudation of ailanthone from A. altissima roots is a mechanism whereby allelopathy could occur, whereas exudation from leaves probably is not. The experiments described so far had a serious weakness: the Ailanthus altissima tissues used had been removed from the trees and injured by cutting or drying. It could therefore be objected that the experiments did not mimic natural situations. Another investigation was performed to assess more realistically the potential for allelopathy caused by root exudation of ailanthone. Soil within two centimeters of A. altissima roots was collected m a twentyyear-old stand of trees and assayed usmg garden cress seeds. Control soil was collected from a nearby forested area containing few A. altissima trees. Cress radicle growth in soil from near A. altissima roots was 85 percent of radicle growth in control soil. The bioassay was repeated because the difference was so small; in the

tively of cress

32

Effect of treatments of ailanthone pre-emergence, photographed 6 days after spray (top row of flats), and post-emergence, 5 days after spray (bottom row of flats). Apphcation rates are (left to mghtJ 0 (control), 0.5, 1, 2, 4, and 8 kilograms of ailanthone per hectare Plant species are (front to back of flats) redroot pigweed (Amaranthus retroflexus), garden cress, velvetleaf, foxtail ~Setaria glaucal, barnyard grass, corn, and (m post-emergence flats only) seedlmgs of Ailanthus altissima. All plants were killed m the postemergence treatments, even at 0 5 kg/ha, except velvetleaf, which was m~ured at the higher rates of apphcation, and A. altissima seedlmgs, which remarkably showed no m~ury even at 8 kg/ha

second test,

cress

A. altissima roots

radicle was

growth in soil near 77 percent of control

radicle growth. The investigations of root exudation provide evidence that ailanthone may be released into the rhizosphere of Allanthus altissima in amounts sufficient to influence the growth of other plants. Before concludmg that A. altisslma is allelopathic, however, another factor had to considered. Many organic compounds are rapidly degraded by soil microorganisms; juglone, for example-the allelopathic compound from black walnut-can be degraded rapidly by soil bacteria to concentrations below which phytotoxicity would occur.’2 If the same is true of ailanthone, its biological effectiveness could be greatly reduced. The persistence of ailanthone in soil was therefore examined. In one investigation, a solution of ailanthone was mixed with soil in petri dishes. In some dishes, soil that had been sterilized by autoclaving was used, whereas

nonsterile soil was used in other dishes. The dishes were then incubated at 25 degrees Centigrade for time periods ranging from 0 to 21 days, and the soil was subsequently tested for phytotoxicity with a cress seed bioassay. Strong toxicity persisted for 21 days in the dishes containing sterile soil; by contrast, it persisted for only 2 or 3 days and rapidly disappeared thereafter in the dishes containing nonsterile soil. A similar pattern was observed when powdered Ailanthus altissima root bark was mixed with soil and incubated. These results clearly demonstrate that the toxic effects of ailanthone in soil are short-lived, probably because of microbial degradation, and raise questions about allelopathic potential of A. altissima under natural conditions. Ailanthus altissima as a Herbicide Regardless of its ecological role, ailanthone is a very powerful herbicidal compound: in the standard garden cress bioassay, radicle growth

33

typically reduced to 50 percent by a solution containing only 0.7 milligrams of ailanthone per liter (0.7 parts per million~.’3 Ailanthone is therefore being evaluated for commercial is

use, since

a

natural herbicide could have several

advantages over synthetic ones: (1) rapid degradation of the herbicide in soil or water, resulting in less environmental pollution; (2) reduced dependence on

down into the seed zone. To test postemergence herbicidal effects, the crude extract or purified ailanthone was sprayed directly onto emerged seedlings of weeds and crop species. Strong herbicidal effects resulted from both pre- and post-emergence applications, but the post-emergence effects were especially striking: even the lowest application rate of ailanthone

fossil fuels since the herbicide could be made biosynthetically rather than from petrochemicals ; and, perhaps (3), lower toxicity of the herbicide to nontarget organisms. Ailanthone can be described as a broad-spectrum herbicide that is toxic to many plants, both weeds and crop species. It has its greatest effect on annual plants shortly after they have emerged, but it also has a

significant

pre-emergence

effect. Ailanthone is toxic to both monocots and dicots, but dicots tend to be the more sensitive. It has a very low degree of selectivity; however, Ailanthus altissima seedlings and certain species in the Malvaceae such as cotton (Gossypium hirsutum) and velvetleaf (Abutilon theophrasti) are

resistant.

Initial investigations of the herbicidal effects of Allanthus altissima were made in the greenhouse using a crude extract of root bark. Later, after the herbicidal compound had been identified, purified ailanthone was used. Both the crude extract and the purified ailanthone were sprayed onto the surface of soil sown with weeds and crop species to test for pre-emergence herbicidal effects. The soil was then watered so that the herbicidal material would be carried

Field trial of post-emergence spray with extract of A. altissima bark mne days after apphcation. The plot at top (control) received no extract, and the plot belowrecemed the eqmvalent of 1 1 kilogram of ailanthone per hectare. The predommant weed is Galinsoga ciliata. The crop plants are (front row to back row) corn, cauhflower (Brassica oleracea var. rtalica/, tomato (Lycopersicon esculentuml, and green bean (Phaseolus vulgans/. A sigmficant reduction m weed populatlon of the treated plot is apparent, but m~ury to the crops is also evident.

34

A mature A~lanthus altissima m August Showmg the charac,tenstrc compound leaves, palm-like appearance, and abundant clusters of mpening samaras.

(equivalent to 0.5 kilogram per hectare) caused complete mortality of most plant species tested. The most recent tests of ailanthone were conducted in outdoor field plots. Because large amounts of the herbicidal material were required and isolation of pure ailanthone is expensive and time-consuming, a crude extract of Ailanthus altissima trunk bark was used.

gmmg

a

somewhat

Weeds and crops were planted in the field and sprayed after emergence with an extract containing a known amount of ailanthone. Symptoms of damage were evident on many weeds and crop species within a few days of spraying. As demonstrated previously in the laboratory, ailanthone does not persist long in the soil, so new weeds germinated and some injured weeds

35

recovered within

a few weeks of spraying. A of single application ailanthone would therefore be insufficient to control weeds over an entire growing season. Future research will investigate ways to extend the herbicidal effects over a longer time and to minimize toxicity to crops.

Conclusion Despite the positive results of many laboratory investigations, we do not yet have enough information to state unequivocally that Ailanthus altissima is allelopathic: too little is known of the complex interactions and potentially mitigating circumstances that occur in the natural environment. However, from an evolutionary standpoint it makes little sense that A. altissima would expend the energy to produce a compound unless it somehow conferred a selective advantage. It is certain that ailanthone has powerful herbicidal effects and may have evolved to inhibit competing plants, but it may also have other functions. Anecdotal evidence suggests that it is toxic to some fungi and may therefore function to protect A. altissima against fungal pathogens. It might also act as a feedmg deterrent to herbivores because of its extremely bitter taste, a possibility suggested by the fact that few animals feed on A. altissima plants. Clearly, there is much we have yet to learn about ailanthone and the secret role it plays m the life of A. altissima.

References

Bell,

physiological role(s) of secondary (natural) products. The Biochemistry of Plants, ed. P. K Stumpf and E. E. Conn. NY: Academic

E. A. 1981 The

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3 4

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Pelgler

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1977, Bell 1981. 1984, Putnam and Tang 1986.

of extracts from tree-of-heaven (Ailanthus altissima) Amencan Journal of Botany 77: 662-670

1990b. Evidence for allelopathy by tree-of-heaven (Ailanthus altissima) Journal of Chemical Ecology 16: 2039-2055. . 1996. Identification of an allelopathic compound from Ailanthus altissima (Simaroubaceae) and characterization of its herbicidal activity. .

Journal of Botany 83: 192-200. Hoffmann,J J., Jolad, L. K Hutter, and S. P. McLaughhn. 1992. Glaucarubolone glucoside, a potent fungicidal agent for the control of grape downy mildew Journal of Agricultural and Food Chemistry 40’ 1056-1057. ~ Hu, S. Y. 1979. Allanthus. Arnoldia 39: 25-50. Ilhck, J. S , and E F. Brouse. 1926 The Ailanthus Tree m Pennsylvama. Bulletm 38. Harmsburg, PA. Pennsylvania Department of Forests and Amemcan

S. D.

Waters.

Klocke, J. A,

M. Amsawa, S. S. Handa, A. D Kmghorn, G. A. Cordell, and N. R. Farnsworth. 1985. Growth mhibitory, msecticidal, and antifeedant effects of some antileukemic and cytotoxic quassmoids on two species of agricultural pests. Expementia 41: 379-382.

B. 1925. Antagomsm of the walnuts (Juglans mgra L. and J. cmerea L.) m certain plant

Massey, A.

Swam

associations.

Rice

F. 1959. A

5 Harper

Mergen,

1977

6

Muller and Muller 1964, Muller and del Moral 1966, Massey 1925, Fisher 1978.

7

Klocke et al. 1985, Polonsky et al. 1989; Hoffmann et al. 1992; Trager and Polonsky 1981, Pierre et al. 1980, Ogura et al 1977.

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8

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Ogura, M., G.

12

Schmidt 1988.

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11

Phytopathology 15: 773-784. principle m the leaves of

toxic

Ailanthus Botamcal Gazette 121: 32-36. H, and R. del Moral. 1966. Soil toxicity induced by terpenes from Salma leucophylla Bulletm of the Torrey Botanical Club 93: 130-

Muller,

9

10

pme growth under conditions. Soil Science

Society of Amenca Journal 42: 801-803. Harper, J. L. 1977. Population Biology of Plants NY: Academic Press, 369-381. Heisey, R. M. 1990a. Allelopathic and herbicidal effects

Endnotes 2

Juglone mhibits

certam moisture

1988.

A. Cordell, A. D. Kinghorn, and N R. Farnsworth. 1977. Potential anticancer agents. VI. Constituents of Ailanthus excelsa (Simaroubaceae). Lloydia 40: 579-584.

R. 1993. A defense of ailanthus. American

Horticultumst

72/2/:

38-43.

36

M. Robert-Gero, C. Tempete, and J. Polonsky 1980. Structural requirements of quassinolds for the inhibition of cell transformation Biochemical and Biophysical Research Commumcations 93: 675-686.

Plasmodmm falciparum m vitro. Amencan Journal of Tropical Medicme and Hygiene 30.

Pierre, A.,

Polonsky, J., S. C. Bhatnagar, D. C Griffiths, J. A. Pickett, and C. M. Woodcock. 1989. Activity of quassinolds as antifeedants against aphids Journal of Chemical Ecology 15: 993-998 Putnam, A.R., and C. S. Tang, eds. 1986. The Science of Allelopathy. NY: Wiley Rice, E. L. 1984. Allelopathy NY: Academic Press. Schmidt, S. K. 1988. Degradation of ~uglone by soil bacteria. Journal of Chemical Ecology 14: 1561-1571

Secondary compounds as protective agents. Annual Remew of Plant Physiology 28: 479-501. Trager, W., and J. Polonsky. 1981. Antimalarial activity of quassinolds against chloroquine-resistant Swain,

T. 1977.

531-537.

Voigt,

G.

K., and F. Mergen. 1962 Seasonal variation in toxicity of ailanthus leaves to pme seedlings

Yang,

R.

Z., and

C. S.

control

m

Botanical Gazette 123: 262-265.

Tang. 1988 Plants used for pest Chma: hterature review. Economic

Botany 42: 376-406.

Rod Heisey, professor of biology at Penn State Umversity, grew up on a farm in Lancaster County, Pennsylvania. While a graduate student at the University of California, Davis, and a postdoctoral researcher at the Pesticide Research Center of Michigan State University, he became mterested in allelopathy and the use of natural products for controlling agricultural pests. His research focuses on plant ecology and microbial ecology, with emphasis on allelopathy and natural-

product pesticides.

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