ON THE TOXICOLOGY OF THE TUTU PLANT

WILLIAM W. FORD

From the Bacteriological Laboratory, Johns Hopkins University

. Received for publication, June 23, 1910

One of the most interesting of the poisons to be obtained in

pure condition within recent years is tutin, tutu, or the toot poison,
the active principle of several species of Coriarie, a group of plants
found, especially in New Zealand and causing the most serious
pecuniary loss to the agriculturalist there, because of the many
deaths which occur in stock from eating the various species. The
Coriaria are shrubs or small trees which grow to a height of three
or four feet in certain varieties, or to a height of twenty to twenty-
five feet in others. Both their succulent branches and their
delicious berries are eaten with avidity by the domestic animals.
The native Maoris have been familiar with the poisonous charac-
ter of these plants since early times and from their language the
term toot or tutu is derived. The early settlers in New Zealand
quickly recognized the danger their stock was subjected to, and
Lauder Lindsay (1) states that in 1863 the colonists uniformlylost
a quarter of their animals from this cause alone, the percentage
sometimes even reaching 75, and that the difficulty of keeping
stock alive proved one of the greatest barriers to the settlement
and development of the land. Lindsay further states that the
animals brought originally by Captain Cook to New Zealand,
sheep and goats, died in some mysterious way, and he identifies
their symptoms as coming from “toot poisoning.” The incoming
Europeans quickly differentiated two varieties of poisonous
Coriarial, one the shrub proper Coriaria thymifolia, called by the
Maoris tutu-papa or tutu-heu-heu (ground toot); the other the
small tree or bush Coriaria ruscifolia, tutu, pohou, tupakihi,
74 WILLIAM W. FORD

(tree toot), and they further soon found that the tutu-berries
were by no means an infrequent cause of death among human
beings.
The various species of Coriarial are identical in their poisonous
action and no animal is naturally immune. Cattle and sheep
especially suffer most severely, but larger animals are susceptible.
An elephant belonging to a traveling menagerie, for instance, died
of toot poison in seven hours, and the skeleton of this animal is
now in the Colonial Museum in Wellington. Birds also are sensi-
live to the poison, and domestic fowls show symptoms of toot-
poisoning from eating the berries. The number of cases on record
of poisoning in man is not large, possibly twenty or twenty-five,
but this is not surprising since the deadly character of the plant
has been so well known for many years, and it presents no di.ffi-
culties of recognition.
Besides Coriaria ruscifolia and Coriaria thymifolia, already
mentioned, Coriaria angustifolia also is occasionally found in
New Zealand and all three species are called tutu by the natives
and by the settlers. Plants belonging to the same order are found
in other countries having the same latitude, the best known of
these being Coriaria m.yrtifolia, which is abundant in Europe. It
is known in Germany as Gerberstrauch, or dyer’s bush, and in
France as redoul. It is largely used in the tanning of leather and
as a black dye, and has considerable commercial value. The leaves
are occasionally used to adulterate Alexandrian senna, and death
has resulted therefrom. The active principle of thisspecies is
Coriamyrtin, a crystalline glucoside belonging to the pikrotoxin
group of poisons and first isolated by the French chemist Riban (2).
Coriaria ruscifolia is found in China, where it yields a black stain
made use of by shoemakers, and the fruit of another species found
in the Himalaya mountains is eaten. On this continent the spe-
cies thymifolia is found in South America where it is known as the
“ink plant,” the juice of the fleshy petals being used as an ink
under the name” chanchi’ ‘(3). The same species grows in Mexico,
but no record of its poisonous action on animals has been found.
I suspect, however, that some of the obscure cases of cattle poison-
ing described are due to this plant both in Mexico and in the
 ON THE TOXICOLOGY OF THE TUTU PLANT 75

extreme southern part of the United States. The plants can be

kept alive in this latitude only under artificial conditions in green-
houses, and while several representatives of the genus have from
time to time been brought to the New York Botanical Garden
and to Boston, none of these are alive at the present time. In
London, however, in the Royal Botanical Gardens there are
several specimens of Coriaria including both the Coriaria myrti-
folia of Europe and the New Zealand varieties. In this latter
country, although it is by all odds the most poisonous of all the
plants found there, yet it serves also a useful purpose to the natives.
The Maoris use the fruit as the source of a non-intoxicating
beverage, musical instruments are made from the hollow stems,
the juice is used for tattooing, and, because of its sweet taste,
is mixed with drinking water and with jellies made from sea weed.
In animals poisoned by these plants the symptoms are largely
referable to the nerve centers and consist of stimulated and then
impaired respirations, tetanic convulsions and coma. The symp-
toms usually make their appearance within a short time after
the plant is eaten, and lead to the death of the animal in a few
hours. In man, recovery from severe poisoning occurs, but im-
pairment of memory may result. Very many remedies have been
tried without marked success. Bleeding is looked upon by stock-
owners as the best method of affording relief and in human beings
this has been combined with the use of stimulants, emetics,
compulsory exercise, and chloroform to control the convulsions.
The only drug which can be said to act as an antidote is lime or
some of the other alkalis. In vitro tutu is destroyed by alkalis
and the administration of lime has been carried out by Dr. James,
of Wellington, in a case of poisoning from eating the berries (4).
Lindsay states that carbonate of ammonia is a valuable remedy
in animals suffering from toot poison. The earlier attempts to
isolate the active principle of the toot plant by Skey (5), Hughes( 6),
and Cristie (7) were not successful, but in 1900 T. H. Easterfield,
Professor of Chemistry in Victoria College, Wellington, and B. C.
Aston, Chemist to the Department of Agriculture, obtained a
crystalline glucoside from all three of the New Zealand species
of Coriarise (8). The fresh young shoots of the plants were finely
76 WILLIAM W. FORD

divided, the juice expressed, filtered, evaporated to a thick syrup,

nearly neutralized by sodium carbonate and then shaken up with
ether. A crystalline deposit occurred on evaporation of the
ether and after further purification by recrystallization from alco-
hol, this material was found to be highly toxic for animals re-
producing in them the symptoms found in animals accidentally
poisoned by the plants. This substance has been named tutu by
its discoverers. It is, according to these authors a colorless, odor-
less compound occurring in oblique-ended prisms. It is volatile,
has an intense and lasting bitter taste and gives the chemical
reactions of a glucoside. That is, it reduces Fehling’s solution
only after inversion by mineral acids, but then very strongly.
It has the percentage composition of C17H20O4. It is highly
toxic, 0.129 gram killing a pig weighing 17 kilos in 5 hours, 0.01
gram killing a kitten weighing 1 kilo in 40 minutes and 1 milligram
(.001 gram) causing a severe illness with convulsions in a cat
weighing 2 kilos.
According to J. A. Gilruth, (9), Chief Government Veterinarian
of New Zealand, the symptoms in the poisoned animals consist
of tetanic convulsions, accelerated breathing, nausea, vomiting,
spasm of the glottis and respiratory muscles, the animals succumb-
ing in the height of the convulsions. The post-mortem examina-
tion reveals nothing beyond a few halmorrhages in the submucous
coat of the stomach and the stoppage of the heart in diastole.
Prof. Marshall states that the pharmacological action of tutin
is “closely allied to that of coriamyrtin, it producing salivation,
a fall in the frequency of the pulse, increased respiratory activity
followed by convulsions for the most part clonic and limited
to the forepart of the body.” Marshall thinks that the poison
produces its effect by acting on the medulla oblongata and basal
ganglia of the brain (10).
Recently through the kindness of B. C. Aston, Chief Chemist
of the Department of Agriculture of New Zealand, I received a
small quantity of crystalline tutin. The extreme toxicity of the
substance for rabbits and guinea pigs and its localization in the
brain and spinal cords of these animals, seem worthy of re-
porting.
 .

ON THE TOXICOLOGY OF THE TUTU PLANT 77

TOXICITY

A small quantity of pure crystalline tutin, 0.15 grams, was dis-

solved in 30 cubic centimeters of distilled water to which a little
ethyl alcohol had been added. The pure crystals dissolve with
considerable difficulty in water, but the addition of the alcohol
renders solution possible, although some time, 18 to 24 hours, is
necessary before solution is complete. One cubic centimeter of
this solution thus represents .005 grams or 5 milligrams of the
pure poison. On the administration of this poison to both guinea
pigs and rabbits, characteristic effects are almost at once apparent.
With 1aige doses the animals are thrown into violent convulsions
within fifteen to thirty minutes. The respirations are labored,
the head is retracted, and the limbs, especially the fore limbs,
show peculiar clonic spasms. The animals run rapidly about their
cages as though making the most violents attempts to escape, and
even when the effect of the poison becomes more pronounced and
the animals sink into a profound coma, the rhythmic movements
of the fore legs are maintained almost to the moment of death.
The full effect of the poison comes on usually in about two
hours, at which time the convulsions are most violent, and the
animals succumb to the intoxication in from four to five hours.
Death may occur much earlier from large doses, and rarely the
animals sink into a profound coma, from which they never recover,
without showing the preliminary stage of convulsions. With
sublethal doses the animals show a peculiar somnolence, with
occasional rapid movements of the fore legs, the symptoms pass-
ing off rapidly, and at the end of 24 hours the animals appear
quite well, with no loss of weight, with natural movements and
good appetite. There are no late effects and no evidence of a
chronic intoxication such as develops when poisons are adminis-
tered which produce degenerative changes of the parenchymatous
organs. Guinea pigs and rabbits are equally susceptible to the
poison, the convulsions, however, being somewhat more general-
ized in the larger animals, the guinea pigs always showing the
peculiar running movement due to the clonic spasms of the fore
limbs.
78 WILLIAM W. FORD

The toxicity of the poison for both rabbits and guinea pigs is
very high, so high indeed as to make tutin rank as one of the most
poisonous of organic substances, certainly one of the most toxic
of the glucosides. The minimum fatal dose of guinea pigs of 250
gram, weight is about half a milligram, as can be seen from the
following table:
TABLE I

Toxicity of Tutin for Guinea Pigs

DOSAGE WEIOBT EFFECT

1 cc. stock solution

545 grams Death 2 hours
5 milligrams
0.5 cc.
o grams Death 2 hours
24 milligrams
0.2 cc. 290 grams Death 2 hours
1 milligram
0.1 cc.
275 grams Death 2 hours
4 milligram
0.05 cc.
315 grams
5 Somnolence and convulsive movements of
4 milligram fore-limbs. Complete recovery

With rabbits the minimum fatal dose is somewhat higher but

bears a definite ratio to the body weight. For an animal of 1200
grams it may be considered about 2 milligrams.

TABLE II

Toxicity of Tutin for Rabbits

DOSAGE WEIGHT EFFECT

1 cc. stock solution

1370 grams Death 2 hours
5 milligrams
0.5 cc.
1395 grams Death 2 hours
24 milligrams
0.2 cc.
1095 grams Somnolence, no convulsions. Recovery.
1 milligram

RESISTANCE TO HEAT

Tutin is far more resistant to heat than the majority of organic

poisons, and may be compared to some of the snake venoms in
 ON THE TOXICOLOGY OF THE TUTU PLANT 79

this respect. It withstands boiling for half an hour, the boiled

material showing apparently no loss in toxicity. Thus one cubic
centimeter of our stock solution was boiled half an hour and, when
administered to a guinea pig weighing 370 grams, killed it in
violent convulsions in half an hour.

PERMANENCE OF ACTION

Tutin solutions preserve their toxicity for long periods of time

without apparent deterioration. Our first tests were made in
February, 1909. The same solution was again tested in Novem-
ber, nine months later, and found to exhibit the same or even a
slightly increased toxicity, the solution being apparently slightly
more concentrated from a little evaporation of the solvent. The
slightly increased toxicity can be seen from the following
table:

TABLE III

Toxicity of Tatin Subutions After Standing

DOSE WEIGHT OF GUINEA PIG EFFECT

lee. 5 -
oOO grains Death 2 hours
5 milligrams
cc
450 grams Death 2 hours
i milligrams
re cc.
260 grams Death 2 hours
4 milligram
cc.
300 grams Death 2 hours
milligram 5
4cc. S -
30 grams No effect
milligram 5
pQcC. 5
400 grams No effect
ie milligram [

This same solution was tested in June, 1910, nearly a year and
a half after its original preparation, and found to Jiave its charac-
teristic toxicity.
80 WILLIAM W. FORD

LOCATION OF TUTIN IN THE TISSUES

In animals dead from tutin intoxication, it is possible to locate

the poison accurately in the nerve structures, to which it is closely
bound. The poison may be identified by its chemical reactions.
It fails to reduce Fehling’s solution untreated, but after hydroly-
sis with mineral acids (preferably strong hydrochloric acid) gives
a characteristic reduction with the deposition of the yellow oxide
of copper. By this reaction it can be detected in the tissues and it
is almost entirely limited to the brain and spinal cord. If these
structures be emulsified with salt solution and boiled with hydro-
chloric acid, after neutralization, they give the reduction of Feh-
ling’s solution characteristic of the glucoside. Indeed these tissues
seem to be loaded with the poison. Controls made with the brains
and spinal cords of normal animals give but slight reduction of
Fehling’s solution, or none, even after prolonged boiling with
hydrochloric acid. At times a little yellowish-green deposit is
found in the bottom of the tubes, but when this is separated by
filtration, decomposed by acetic acid and tested by ammonia for
copper (cupra-ammonium oxide) the test is absolutely negative.
Moreover, if the tissues of the brain and spinal cord are emulsified
in salt solution and then filtered, the clear filtrate from normal
animals gives no trace of reduction of Fehling’s solution. Similar
clear filtrates from both the brain and spinal cord of animals
dead of tutin give the characteristic deposit of cuprous oxide when
tested with the reagent. The blood of inoculated animals may
show at times a faint trace of the poison, but the various organs,
liver, kidney, ‘spleen and adrenals, are quite free from it or at
least show only those traces which can be accounted for by the
amount of blood which they may contain.
The poison is clearly located then in the brain and spinal cord,
and is closely bound to these tissues. If the brain and spinal cords
of animals dead of tutin be removed aseptically and emulsified,
this emulsion of tutin-loaded nerve tissue may be given to animals
without the slightest effect. Not only is the poison localized in
these structures, but it is bound to them in such a way as to be
completely neutralized. Thus the brain of a 500 gram guinea
 ON THE TOXICOLOGY OF THE TUTU PLANT 81

pig killed by the administration of five milligrams of tutin in half

an hour, was emulsified in salt solution and the thick emulsion
administered subcutaneously to a guinea pig weighing 400 grams.
No effect whatever seemed to follow the inoculation. The same
experiment was carried out in a number of animals, but in no case
could any poisonous action be detected. Ocular inspection’ of
the amount of reduction of Fehling’s solution produced by the
nerve structures of the animals dead of tutin intoxication, as
compared with the apparent amount of reduction produced by
definite quantities of tutin would lead one to believe that the
quantity of the glucoside present in the brain emulsioiis used in
these experiments is much more than a fatal dose of the active
poison and yet the animals showed no symptoms whatever,
even the somnolence and the peculiar clonic spasms of the fore
limbs produced by sublethal doses being absent. The poison
seems to be so closely bound to the nerve structures or to be so
altered in its chemical constitution either by the loss of certain
constituents or by the assumption of some of the constituents
of the tissues that it is no longer toxic. It may provisionally be
spoken of as detoxified tutin.
Attempts to imitate in vitro this union of poison and nerve sub-
stance were unsuccessful. Aseptically removed brains and spinal
cords fail to detoxify tutin and when emulsified with it and intro-
duced subcutaneously into animals they die with all the signs of
tutin intoxication. Various methods were employed. The tissues
were emulsified with tutin solutions and injected at once, were pre-
served on ice for 18 hours and injected, were kept at 37#{176}
C. for the
same length of time and injected, and in all instances the effect
of the emulsion of nerve tissues and tutin was quite like that of
the pure poison. Thus the brain of a normal guinea pig emulsified
with one cubic centimeter of our stock tutin solution, and then
kept on ice over night, was fatal to a 400 gram guinea pig in two
hours, while another mixture of brain and one-half cubic centi-
meter of the solution, kept in the thermostat 18 hours killed a 370

The quantity of tutin at my disposal was not so large as to admit of a series

of quantitative estimations in respect to the point at issue.
82 WILLIAM W. FORD

gram pig in the same length of time. Not even a minimum fatal
dose of the poison will be neutralized outside the body, since an
emulsified brain mixed with two drops of tutin solution, represent-
ing the lowest limits of dosage of the poison for guinea pigs, and
left in the thermostat over night, killed a 305 gram guinea pig
in two hours.
The brains of animals removed from the body contain but
relatively small amounts of blood serum and it was thought possi-
ble that some constituent of the serum might be necessary to
effect a union of tutin and the nerve tissues, but no evidence
could be adduced to show that blood serum has such an action
‘ ‘ in vitro.” Thus one and one-half cubic centimeters of fresh
normal guinea pig serum mixed with two drops of tutin solution
killed a 291 gram pig in two hours, and the same quantity of serum
added to a brain emulsion and two drops of stock tutin solution,
the mixture being kept on ice for 48 hours, killed a 420 gram guinea
pig in two hours with the typical symptoms of tutin intoxication.
We are apparently unable to imitate outside the animal organ-
ISIII that peculiar process which in the body permits this powerful
nerve poison to be localized and bound to the nerve structures and
in such a combination to be completely detoxified.

IMMUNITY EXPERIMENTS

Gilruth (bc. cit.) has pointed out that animals recovering from
non-fatal doses of the toot poison exhibit no subsequent resist-
ance when inoculated with the fatal doses for normal animals.
He reports the administration of one milligram of tutin to a cat
weighing two kilograms and the production of convulsions and an
illness lasting 24 hours. The animal recovered, but subsequently
succumbed when inoculated with 3 milligrams of the poison.
The attempt was made to produce an immunity to this poison
in both rabbits and guinea pigs, but with both species of animals
the introduction of small non-fatal doses seemed to have no effect
whatever in heightening the resistance of the animals to the action
of fatal doses. After the effect of the poison wears off the animals
are susceptible to apparently the same degree as are normal ani-
 ON THE TOXICOLOGY OF THE TUTU PLANT 83

mals. There is no increased resistance, the animals dying of

fatal doses for normal animals, and there is no evidence that the
poison accumulates in the system. Even after the introduction
of a number of small doses, over a fairly long period of time, it
still requires a minimum fatal dose to bring on a fatal intoxica-
tion. The following tables indicate these points:

TABLE IV

Treatment of Rabbit

DATE DOSAGE WEIGHT RESULT

March 30 1 milligram j 1095 grams Somnolence and recovery

May 4 14 milligrams 1295 grams Somnolence and recovery

May 31 2 milligrams 1450 grams [Typical symptoms of tutin intoxica-

tion and death in 2 hours

TABLE V

Treatment of Guinea Pmq

DATE DOSAGE WEIGHT RESULT

May 10 1 milligram 315 grams #{149}No

appreciable effect
(Death in 2 hours, with typical
May 31 2 milligrams symptoms

Other animals treated with small doses of tutin at varying

intervals likewise showed no evidence of increased resistance, all
succumbing as soon as a normal fatal dose was administered.
Even the administration of the detoxified tutin in the form of the
emulsion of the brain of an animal killed with this poison, failed
to induce any immunity. Thus on November 18th a 500 gram
guinea pig was inoculated with one cubic centimeter of tutin
solution and died in violent convulsions. The brain of this animal
was removed, emulsified and injected into a guinea pig weighing
420 grams. No effect followed the injection and on December 10th
one drop of the tutin solution representing milligram and on
December 23rd two drops, representing milligram were admin-
istered, also without effect. On January 10th, 18 days after the
last inoculation, the animal succumbed to 3 drops of the solution,
representing about 1 milligram, a fatal dose for animals of its
weight.
84 WILLIAM W. FORD

The failure to immunize animals to this glucoside is not without

significance, and is in line with the work of Bashford, (11) and
Ehrlich as to the character of the substances to which immunity
can be produced. (12) It is Ehrlich’s doctrine that the organic
poisons of a firm fixed structure such as the crystalline glucosides
form so intimate a union with the cells of the body that these
cells fail to throw off any immune substances and the attempts
of Bashford to immunize against a number of glucosides resulted
in complete failure. It has been shown by the investigations
upon the Amanita-hcrrnolysin, (13) however, that there are certain
glucosides against which an artificial immunity can be produced,
and the question arises, therefore, as to why it is that such a
marked difference is shown by different representatives of this
group. It may be pointed out that the Arnanita-hrmoiysin has
many features in common with the true toxins. It is thermo-
labile, deteriorates rapidly on standing, is destroyed by acids
and shows a definite latent period in its action upon animals.
Tutin, however, is thermostabile, does not deteriorate on stand-
ing, acts upon animals without a latent period and forms an
intimate union with the issues of the body upon which its
poisonous action is most pronounced. Tutin may be said to
follow the general rule enunciated by Ehrlich as to the impossi-
bility of immunizing against glucosides, while the Amanita-
hcemolysin is an exception to this rule. Possibly the ease with
which a poison may be broken up into its various components
may be a more important determining factor in immunity
production than the fact that it gives proteid rather than gluco-
cidal reactions.
CONCLUSIONS

In conclusion, then, we have in tutu a crystalline glucoside

whose toxicity will place it among the most poisonous of organic
poisons, at least of poisonous glucosides. The poison does not
deteriorate on standing and resists boiling. In the body it fonts
an intimate union with the nerve structures and in so combining
is completely detoxified. This union and detoxification cannot
be imitated outside the animal body. The administration of
 ON TIlE TOXICOLOGY OF THE TUTU PLANT 85

doses causing symptoms of intoxication but not acutely fatal is

not followed by secondary degenerative changes and the animals so
treated recover completely. Finally such recovered animals
show no increased susceptibility to the poison and exhibit no
immunity against the action of normally fatal doses.

BIBLIOGRAPHY

(1) Lindsay, Lauder: On the toot poison of New Zealand. Report of the British
Association’s Thirty-second Meeting. Cambridge, 1862. Also, Contri-
butions to New Zealand Botany. London, 1868.
(2) Riban, M. J. : Surle principe toxique du Coriaria Myrtifobia (Redoul). Compt.
Rend. de l’Acad. des Sciences. Paris, 1863, p. 798. (Cited from Easter-
field and Aston.) Also, Recherches experimentales sur Ic principe
toxique du redoul (Coriaria myrtifolia). Paris, 1863. Also, Sur le coria-
myrtin et ses d#{233}riv#{233}s.
Compt. Rend. de I’Acad. des Sciences. Paris, 1866.
(3) Jameson: On the ink plant of New Granada. Proc. Linn. Soc., vol. vii, p. 120.
(4) Easterfield and Aston : (bc. cit.).
(5) Skey: On the extraction of the poisonous principle of the tutu plant (Con-
aria ru.scifolia). Transactions of the New Zealand Institute, 1869, pp.
153, 399, 400.
(6) Hughes: On certain properties of the tutu plant (Coriaria ru.scifolia). Trans..
actions of the New Zealand Institute, 1870, 237.
(7) Cristie: On the physiological action of Coriaria ruscifolia, the tutu poison
of New Zealand. New Zealand Medical Journal, July and October, 1890.
(8) Easterfield and Aston: Tutu, parts i and ii. New Zealand Department of
Agriculture, Wellington, New Zealand, 1900 and 1901. Also studies on the
chemistry of the New Zealand Flora. Transactions of the New Zealand
Institute, 1900.
(9) Easterfield and Aston: (boc. cit.).
(10) Marshall C. R.: The pharmacological action of tutu, the toot plant of
New Zealand. Transactions of the Royal Society of Edinburgh, vol.
xlvii, part ii, No. 13.
(11) Bashford: Arch. Internat. de Pharm. et de Ther. 1901, 8, p. 101; 9, p. 451.
(12) Ehrlich: Chemical constitution and pharmacological action: Collected
studies on immunity. New York, 1906, p. 433.
(13) Ford: The toxines and antitoxines of poisonous mushrooms. The Journal
of Infectious Deseases, vol. iii, No. 2, April, 1906, pp. 191-224. Also anti-
bodies to glucosides, with special reference to Rhus Toxicodendron.
The Journal of Infectious Diseases, vol. iv, No. 4, October, 1907.
See also
Abel and Ford: On the poisons of Amanita Phalloides. The Journal of
Biological Chemistry, vol. ii, No. 4, January, 1907.
Fitchett and Malcolm: On the physiological action of tutin. Quarterly
Journal of Experimental Physiology, vol. ii, p. 335, October, 1909.
Fitchett: Transactions of the New Zealand Institute, vol. xli, p. 286.