1

The Discovery of Ivermectin and Other Avermectins

W. C. CAMPBELL, R. W. BURG, M. H. FISHER, and R. A. DYBAS

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Merck Institute for Therapeutic Research, Rahway, NJ 07065

The avermectins were initially detected in a

Downloaded via 64.114.52.119 on March 3, 2024 at 19:17:16 (UTC).

program in which thousands of microbial fermenta-

tion products were tested in mice for activity
against the nematode, Nematospiroides dubius.
Among the few preparations showing activity in this
assay, was the product of a microorganism isolated
from soil by workers at The Kitasato Institute.
The microorganism was classified as a new species
of actinomycete, Streptomyces avermitilis. Its
anthelmintic activity was shown to reside in 8
closely related macrocyclic lactones, named aver-
mectins, which were also found to possess activity
against free-living and parasitic arthropods. One
of the natural components, avermectin B , is now 1

being evaluated as a pesticide for the control of

mites of citrus and cotton crops and control of the
Red Imported Fire Ant. A chemical derivative,
22,23-dihydroavermectin B , or ivermectin, has been
1

developed as an antiparasitic agent. It is being

marketed for use in cattle, horses and sheep and is
expected to become available for swine and dogs.

The t i t l e of t h i s s e s s i o n , Guided and Serendipitous Discovery

(within the symposium, Approaches to R a t i o n a l Synthesis o f
P e s t i c i d e s ) focuses a t t e n t i o n on the nature of the d i s c o v e r y ,
r a t h e r than the outcome. The d i s c o v e r y of the avermectin
f a m i l y of compounds was by no means s e r e n d i p i t o u s ; those who
were seeking found what they sought. I t i s the purpose of
t h i s paper to r e c o r d the manner of the seeking and the manner

0097-6156/ 84/ 0255-0005S06.00/ 0

© 1984 American Chemical Society

Magee et al.; Pesticide Synthesis Through Rational Approaches

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
6 PESTICIDE SYNTHESIS THROUGH RATIONAL APPROACHES

of assessing and enhancing what was found. The discovery of

i v e r m e c t i n i s part of the l a r g e r s t o r y of the discovery of the
avermectin family of compounds: the avermectins are produced
by a microorganism, and ivermectin i s a chemical m o d i f i c a t i o n
of one of those substances.
The i n i t i a l o b j e c t i v e of the search was an anthelmintic
with p r o p e r t i e s r a d i c a l l y d i f f e r e n t from those of known
anthelmintics. What was found was an a n t h e l m i n t i c whose
p r o p e r t i e s were indeed markedly d i f f e r e n t — not only i n terms
of chemical s t r u c t u r e and e f f i c a c y against helminths, but a l s o
i n the extension of the p o t e n t i a l u t i l i t y of the c l a s s to the
c o n t r o l of arthropod p a r a s i t e s of animals. Further, the
compounds turned out to have s t r i k i n g a c t i v i t y against a
v a r i e t y of f r e e - l i v i n g and p l a n t - p a r a s i t i c nematodes and
arthropods — and so i t has come about that a new l i v e s t o c k
a n t h e l m i n t i c has become an agenda item i n t h i s symposium on
agricultural pesticides.

Primary Screening and Microbiology

Several i n v i t r o assays f o r d e t e c t i n g fermentation products

w i t h a n t h e l m i n t i c a c t i v i t y had been run without success,
p r i m a r i l y because of the l a r g e number of t o x i c compounds which
had to be e l i m i n a t e d . F i n a l l y , the d e c i s i o n was made to use
an i n v i v o assay i n mice with the hope that the mice would
screen out the n o n - s p e c i f i c t o x i c compounds.
The nematode, Nematospiroides dubius, was s e l e c t e d f o r
the assay. Infected mice were fed f o r s i x days with m i l l e d
Purina Lab Chow which had been mixed with the fermentation
product to be t e s t e d . The mice were then fed a normal d i e t
and, at 14 days p o s t i n f e c t i o n , f e c a l p e l l e t s were examined f o r
the presence of eggs. I f eggs were absent on three successive
days, the mice were s a c r i f i c e d and t h e i r small i n t e s t i n e s
examined f o r the presence of worms.
This assay was r e l a t i v e l y s u c c e s s f u l i n achieving the
goal of e l i m i n a t i n g n o n s p e c i f i c t o x i c a c t i v i t i e s . Among the
many thousands of c u l t u r e s tested i n t h i s assay, about 1% were
a c t i v e i n the f i r s t t e s t . A l l a c t i v e c u l t u r e s were regrown,
and about 20% of the c u l t u r e s i n i t i a l l y s e l e c t e d were a c t i v e .
Among the c u l t u r e s confirmed a c t i v e , most were s t i l l extremely
t o x i c and were not pursued f u r t h e r . However, during the
t e s t i n g of a l a r g e c o l l e c t i o n of c u l t u r e s which had been
r e c e i v e d from The K i t a s a t o I n s t i t u t e of Tokyo, Japan, the
choice of the i n v i v o assay was f i n a l l y j u s t i f i e d . Among one

Magee et al.; Pesticide Synthesis Through Rational Approaches

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
1. C A M P B E L L ET AL. Discovery of Ivermectin 1

group of 50 K i t a s a t o c u l t u r e s submitted f o r assay was a

c u l t u r e bearing the number 0S-3153. The r e s u l t s of the e a r l y
assays of t h i s c u l t u r e are summarized i n Table I. I t was
scored as f u l l y a c t i v e (no eggs, no worms) i n the f i r s t t e s t .
Marked r e d u c t i o n i n mouse body weight i n d i c a t e d that the
fermentation product was e i t h e r unpalatable or t o x i c .
C u l t u r e 0S-3153 was regrown on the o r i g i n a l medium
(designated KH) and on a second, u n r e l a t e d medium (JH) (Table
I, Experiment No. 2). The c u l t u r e grown on the second medium
was very t o x i c , r e s u l t i n g i n host death on the day that the
f i r s t f e c a l p e l l e t was examined f o r eggs. That grown on the
o r i g i n a l medium was a l s o t o x i c , causing severe suppression of
weight gains; but, again, there were no eggs or worms.
In the t h i r d t e s t , the a c t i v i t y was t i t r a t e d u s i n g s e r i a l
2 - f o l d d i l u t i o n s . A l l four l e v e l s were f u l l y a c t i v e , and t h i s
time there was l i t t l e s i g n of t o x i c i t y . A few c u l t u r e s had
been confirmed a c t i v e p r e v i o u s l y , but none had e x h i b i t e d a
s e p a r a t i o n of t o x i c i t y from a c t i v i t y , l e t alone a c t i v i t y over
at l e a s t an 8 - f o l d range. A f t e r an i n a u s p i c i o u s s t a r t , the
a c t i v i t y of c u l t u r e OS-3153 had been f i r m l y e s t a b l i s h e d .
Success had been achieved i n the quest f o r a fermentation
product with a n t h e l m i n t i c a c t i v i t y . In f a c t , as events would
soon prove, the newly found product had an even broader
a c t i v i t y than had been a n t i c i p a t e d .
The c u l t u r e , now bearing the product number C-076 and the
Merck c u l t u r e c o l l e c t i o n number MA-4680, was submitted f o r
taxonomic s t u d i e s . I t s c h a r a c t e r i s t i c s , i n c l u d i n g a brownish-
gray spore mass c o l o r , smooth spore s u r f a c e , s p i r a l
sporophores born as side branches on the a e r i a l mycelia and
the p r o d u c t i o n of melanoid pigments, were u n l i k e those of any
p r e v i o u s l y d e s c r i b e d species of Streptomyces. The c u l t u r e was
named Streptomyces a v e r m i t i l i s , the Streptomyces "capable of
separating from worms".
Based on a c o r r e l a t i o n of a n t h e l m i n t i c a c t i v i t y and HPLC
a n a l y s i s of the t o t a l avermectin complex, i t was estimated
that the t h i r d fermentation contained a minimum of 9 yg/ml.
Improvement of the medium increased the y i e l d by the o r i g i n a l
c u l t u r e (MA-4680) to 120 yg/ml. A high-producing i s o l a t e
(MA-4848) obtained from t h i s c u l t u r e produced n e a r l y 500 yg/ml
of t o t a l avermectins. Thus, t h i s c u l t u r e y i e l d e d r e a d i l y to
medium improvement and i s o l a t e s e l e c t i o n to produce r e l a t i v e l y
l a r g e amounts of the avermectins. Accounts of the e a r l y
fermentation s t u d i e s and taxonomy have been p u b l i s h e d (1-2).

Magee et al.; Pesticide Synthesis Through Rational Approaches

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
8 PESTICIDE SYNTHESIS T H R O U G H RATIONAL APPROACHES

Table I . Summary of the F i r s t Assays of

C u l t u r e OS-3153.

Feed Mouse
Expt. Eaten Weight
No. Medium Dose ' (g) (g) Activity

1 KH 50 13 14 A
2 KH 50 25 15 A
JH 50 13 - Dead
3 KH 50 25 22 A
25 25 25 A
12 25 29 A
6 25 28 A
JH 50 25 22 SA
25 25 29 SA
12 25 — ΝΑ

» Ml of broth per 25 g of feed.

(2) A, a c t i v e - no eggs or worms; SA, s l i g h t l y a c t i v e - no

eggs, worms present; NA, not a c t i v e .

Magee et al.; Pesticide Synthesis Through Rational Approaches

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
1. C A M P B E L L ET A L . Discovery of Ivermectin 9

Chemistry

I s o l a t i o n . The avermectin complex, c o n s i s t i n g of f o u r major

a n c o u r w e r
components designated A^ , A , ^ l a * ^2a
a 2 a * ^ l°
w a s e x t r a c t e (
homologs designated A ^ , A ^, ^ i b * ^2b* 2 ^ with
acetone from the mycelia of Streptomyces a v e r m i t i l i s (3).
Solvent p a r t i t i o n and a d s o r p t i o n on granular carbon produced
an o i l y residue c o n t a i n i n g 5% 16% A , 20% Β and 15% Β ·
2 χ 2

Separation of the A components from the Β components was

achieved by p a r t i t i o n chromatography with hexane-methylene
chloride-methanol (10:10:1) over Sephadex LH-20. A^ was
separated from A u s i n g Sephadex LH-20 and a s o l v e n t system
2

c o n t a i n i n g hexane-toluene-methanol (6:1:1). B^ was separated

from Β2 e i t h e r by c r y s t a l l i z a t i o n from ethylene g l y c o l or by
Sephadex LH-20 chromatography using hexane-toluenemethanol
(3:1:1). S i m i l a r chromatography was a l s o used to separate the
lower homologs and p u r i t y was e s t a b l i s h e d by reverse-phase
HPLC a n a l y s i s .

S t r u c t u r e Determination. The s t r u c t u r e s of a new group of

p e s t i c i d a l sixteen-membered lactones named milbemycin B^, B , 2

and Ββ were d e s c r i b e d i n 1975 (4) on the b a s i s of X-ray

a n a l y s i s . A l a t e r p u b l i c a t i o n (5) gave d e t a i l s of i s o l a t i o n
and s t r u c t u r e s of t h i r t e e n milbemycins, with s p e c t r a l data.
The avermectins were discovered i n 1975 and p a r t - s t r u c t u r e s ,
1ο
J
deduced from proton and C NMR s p e c t r a and t h e i r mass
s p e c t r a l fragmentation p a t t e r n s , suggested a c l o s e
r e l a t i o n s h i p with the milbemycins ( 6 ) . Methanolysis of aver­
mectin A gave an aglycone and a 6:1 mixture of
2 -and
-methyl-Loleandroside. The recovery of t h i s g l y c o s i d e i n more
than 100 mol. % y i e l d demonstrated the presence of two
i d e n t i c a l sugars i n the molecule. Further s p e c t r a l exami­
n a t i o n i n d i c a t e d the attachment of an a-L-oleandrosyl-ot-L-
oleandrosyloxy d i s a c c h a r i d e to the 13-ct-position of the
macrolide r i n g . A chemical proof f o r the p o i n t of attachment
and i d e n t i t y of the d i s a c c h a r i d e was provided by o z o n o l y s i s
C
and i s o l a t i o n of the d i s a c c h a r i d e attached to the fragment l l
C
through 1 4 . The s t r u c t u r e s of the e i g h t components are shown
i n F i g . 1 and a l l c o n t a i n the same d i s a c c h a r i d e s u b s t i t u e n t at
the 13-α-position. They vary at C-5 with hydroxy or methoxy
groups, at C-23 with an a x i a l M i y d r o x y group on a 22,23-olefin
and a t C-25 with i s o p r o p y l or s e c - b u t y l groups i n c o n t r a s t to
the methyl and e t h y l s u b s t i t u e n t s at the 2 5 - p o s i t i o n of the
milbemycins. X-ray a n a l y s i s (7) of B 2 a aglycone and B^ both a

Magee et al.; Pesticide Synthesis Through Rational Approaches

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
 QCH,

α series R
•v
Δ, R = CH 2 5 = CH(CH )C H
3 2 5 A 2 R = CH
5 3
5 3
m
Β, R = Η
5
b series R 2 5 = CH(CH ) 3 2 B 2 R = H
5
H
a
m
H 2 (1 atm) ( 0 P )
3 3 RhCI
0 C H , 25°, 18 h 8 5 %
3
Η
Χ
m

C
4' °v H 3
H
a:
Ο
C
ο
X

25
δ

Magee et al.; Pesticide Synthesis Through Rational Approaches

>
r
>

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.

no
22, 23-Dihydroavermectin B }

ivermectin η
χ
m
in
F i g u r e 1. S t r u c t u r e s of the Avermectins.
1. CAMPBELL E T A L . Discovery of Ivermectin 11

confirmed the s t r u c t u r e and, through the L-oleandrose,

e s t a b l i s h e d the absolute stereochemistry.

Parasitological Evaluation

Following the demonstration of e f f i c a c y i n the

Nematospiroides-mouse assay, and the a s s o c i a t e d
m i c r o b i o l o g i c a l and chemical research d e s c r i b e d above, much
work was needed to determine the r e l a t i v e a n t h e l m i n t i c
e f f i c a c y of both the n a t u r a l avermectin components and the
d e r i v a t i v e s . For t h i s purpose two types of bioassay were
employed. In one, compounds were t e s t e d i n s m a l l l a b o r a t o r y
animals i n f e c t e d with nematodes other than N. dubius. For
example, t e s t m a t e r i a l s were given to j i r d s (Meriones
unguiculatus) i n f e c t e d with T r i c h o s t r o n g y l u s c o l u b r i f o r m i s ,
and the animals were subsequently k i l l e d f o r determination of
worm burden ( 8 ) . The use of that h o s t - p a r a s i t e combination
f o r a n t h e l m i n t i c t e s t i n g had been reported by P a n i t z and Shum
(9) and has proved u s e f u l i n the e v a l u a t i o n of a v a r i e t y of
a n t h e l m i n t i c s , i n c l u d i n g the avermectins. In the other,
compounds were tested a g a i n s t a v a r i e t y of nematodes i n sheep.
These were s m a l l s c a l e t e s t s , done i n c o n j u n c t i o n with the
small-animal t e s t i n g and p r o v i d i n g important i n f o r m a t i o n on
the e f f i c a c y of the t e s t substances i n a ruminant host.
Compounds of s p e c i a l i n t e r e s t were s i m i l a r l y t e s t e d against
helminths i n sheep, and o c c a s i o n a l l y i n other h o s t s , u s i n g
l a r g e r numbers of t e s t animals (10-12).
As the remarkable potency and unique s t r u c t u r e of the
avermectins became apparent, t e s t i n g was extended to organisms
other than helminths. The f i r s t t e s t against an i n s e c t was
done using the Confused F l o u r B e e t l e , T r i b o l i u m confusum, and
the i n c o r p o r a t i o n of the t e s t substance i n t o the f l o u r i n
which the b e e t l e s l i v e d . In t h i s f a s h i o n the i n s e c t i c i d a l
a c t i v i t y of the avermectins was demonstrated (13) and was
followed independently and almost immediately by the
demonstration of e f f i c a c y against a p a r a s i t i c i n s e c t (14).
E f f i c a c y against p a r a s i t i c i n s e c t s was f u r t h e r e s t a b l i s h e d by
t e s t s u s i n g the rodent bot, Cuterebra sp., i n mice (D. A.
O s t l i n d , unpublished) and was shown to extend to some
p a r a s i t i c a c a r i n e s (15-16). Tests against the trematodes
Schistosoma mansoni and F a s c i o l a h e p a t i c a and the cestode
Hymenolepis diminuta i n l a b o r a t o r y animals f a i l e d to show
e f f i c a c y (D. A. O s t l i n d , unpublished d a t a ) . T h i s i s i n accord
w i t h r e p o r t s that the avermectins d i s r u p t GABA-mediated nerve

Magee et al.; Pesticide Synthesis Through Rational Approaches

ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
 PESTICIDE SYNTHESIS T H R O U G H RATIONAL APPROACHES

transmission i n nematodes and arthropods and that f l u k e s and

tapeworms do not employ GABA as a neurotransmitter (17).

A g r i c u l t u r a l Chemical E v a l u a t i o n

The r e s u l t s (above) against the Confused F l o u r B e e t l e

( T r i b o l i u m confusum), rodent bot (Cuterebra spp.), and the
e c t o p a r a s i t i c l a r v a of the sheep b l o w f l y ( L u c i l i a cuprina)
were s u f f i c i e n t l y encouraging to suggest that the avermectins
may possess general b i o l o g i c a l a c t i v i t y against arthropod
pests and, i n p a r t i c u l a r , those of importance i n crop
p r o t e c t i o n (18). To i n v e s t i g a t e t h i s p o t e n t i a l and to expand
our in-house c a p a b i l i t i e s , a research program was e s t a b l i s h e d
w i t h the Boyce Thompson I n s t i t u t e f o r P l a n t Research to t e s t
the avermectin d e r i v a t i v e s i n t h e i r m i t i c i d e and i n s e c t i c i d e
screens. In a l l , n e a r l y s e v e n t y - f i v e r e l a t e d s t r u c t u r e s ,
n a t u r a l products and semisynthetic d e r i v a t i v e s , were evaluated
i n the greenhouse f o r t o x i c i t y t o a spectrum of arthropod
p e s t s . Of these, avermectin B^, the major component of the
fermentation process, was determined to be the most promising
candidate as an a g r i c u l t u r a l p e s t i c i d e .
R e s u l t s from these l a b o r a t o r y s t u d i e s demonstrated that
avermectin B^ had high t o x i c i t y f o r the twospotted s p i d e r mite
(Tetranychus u r t i c a e ) on bean p l a n t s . When a p p l i e d i n
s o l u t i o n d i r e c t l y onto adult and nymphal s p i d e r mite
populations on f o l i a g e , avermectin B^ was shown to be 50-200
times as potent as commercially a v a i l a b l e a c a r i c i d e s , with an
LC 9 0 of 0.02-0.03 ppm. A d d i t i o n a l t e s t s on f o l i a g e with
i n s e c t s i n the order Lepidoptera, Coleoptera, Homoptera,
Orthoptera, D i p t e r a , Isoptera and Hymenoptera confirmed the
broad spectrum a c t i v i t y and potency of the avermectin f a m i l y
of compounds and avermectin B^ i n p a r t i c u l a r . Table II pro-
v i d e s LC(JQ values f o r avermectin B^ f o r the c o n t r o l o f l a r v a l
forms of s e v e r a l of these i n s e c t s i n f o l i a r residue assays
(18).

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ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
1. C A M P B E L L ET AL. Discovery of Ivermectin 13

T a b l e II. E f f i c a c y of F o l i a r Residues of Avermectin

B^ Against Adult mites and L a r v a l
a

Insects.

L C m
Insect 9 0 (PP )

Twospotted s p i d e r mite 0.02-0.03

Tomato hornworm 0.02
Colorado potato b e e t l e 0.03
Mexican bean b e e t l e 0.2
Cabbage looper 0.75-1.2
Southern armyworm 6.0

On the b a s i s of the e f f i c a c y demonstrated i n the green-

house and l a b o r a t o r y s t u d i e s avermectin was s e l e c t e d f o r
development and assigned the Merck development code number
MK-936. Avermectin B^ has been evaluated worldwide f o r
e f f i c a c y against mites and i n s e c t s a f f e c t i n g a number
of a g r i c u l t u r a l crops i n c l u d i n g c i t r u s , c o t t o n , apples, pears,
vegetables, potatoes, t r e e nuts, and grapes. Under f i e l d use
c o n d i t i o n s i t has been observed that e x c e l l e n t c o n t r o l of a
number of economically important pests i n c l u d i n g the c i t r u s
rust and red mite, twospotted s p i d e r mite, broad mite,
Colorado potato b e e t l e , diamond back moth, pear p s y l l a , and
Liriomyza leafminers can be achieved at extremely low
a p p l i c a t i o n r a t e s of MK-936 i n the range of 0.005-0.03 l b per
acre (5.5 - 33 g per h e c t a r e ) . For f o l i a g e a p p l i c a t i o n s a
0.15 EC (1.8% w/v) e m u l s i f i a b l e concentrate f o r m u l a t i o n has
been developed. F i e l d s t u d i e s have shown that the f o r m u l a t i o n
i s non-phytotoxic to a l l target crops on which i t has been
evaluated i n c l u d i n g many v a r i e t i e s of s e n s i t i v e ornamental
plants.
During the course of the development program, samples of
avermectin B^ were provided to a number of outside agencies
f o r e v a l u a t i o n i n s p e c i a l i z e d assays. As a consequence, i t
was discovered i n t e s t i n g conducted by the USDA l a b o r a t o r y f o r
Insects A f f e c t i n g Man and Animals, G a i n e s v i l l e , F l o r i d a , that
the red imported f i r e ant (Solenopsis i n v i c t a ) i s among the
most s u s c e p t i b l e species of i n s e c t s to the t o x i c a c t i o n of
avermectin B ^ . When a p p l i e d i n a corn g r i t b a i t , avermectin

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14 PESTICIDE SYNTHESIS THROUGH RATIONAL APPROACHES

B^ at r a t e s as low as i n 25 to 50 mg per acre has been

e f f e c t i v e i n c o n t r o l l i n g f i r e ant i n f e s t a t i o n s i n large s c a l e
t r i a l s i n the southern United S t a t e s . A submission f o r
r e g i s t r a t i o n of MK-936 f o r t h i s a p p l i c a t i o n has been made.
The avermectin n a t u r a l products are p e s t i c i d e s possessing
novel chemistry and mode of a c t i o n . C r o s s - r e s i s t a n c e has not
been observed i n l a b o r a t o r y or f i e l d s t u d i e s with mites
andinsects t o l e r a n t to commercially a v a i l a b l e organophosphate,
carbamate, c h l o r i n a t e d hydrocarbon and p y r e t h r o i d p e s t i c i d e s .

S y n t h e t i c Program

S t r u c t u r e - A c t i v i t y R e l a t i o n s h i p s . Compounds of the Β s e r i e s
were g e n e r a l l y more potent than those of the A s e r i e s . Thus
an u n s u b s t i t u t e d hydroxy group at the 5 - p o s i t i o n i s a c t i v i t y
enhancing (19). D i f f e r e n c e s i n potency between the 1- and 2-
s e r i e s v a r i e d among p a r a s i t e s , but i n most instances the 1-
s e r i e s was more potent. Reduction of the 22,23-olefin had
l i t t l e e f f e c t on a c t i v i t y but f u r t h e r r e d u c t i o n caused a
s u b s t a n t i a l decrease i n a c t i v i t y . The monosaccharides were
two- to f o u r f o l d l e s s a c t i v e than the parent compounds w h i l e
the aglycones were more than t h i r t y f o l d l e s s a c t i v e , Table I I I .
A c e t y l a t i o n at the 4 " - p o s i t i o n caused no change i n a c t i v i t y
whereas a c e t y l a t i o n at the 5- or 23- p o s i t i o n caused a
c o n s i d e r a b l e decrease i n a c t i v i t y . D i a c e t a t e s and t r i a c e t a t e s
showed s i m i l a r l y reduced a c t i v i t y , Table IV (20).

Ivermectin. E a r l y b i o l o g i c a l s t u d i e s demonstrated that while

avermectin B^ was more a c t i v e than avermectin B by o r a l 2

a d m i n i s t r a t i o n , the converse was true when the compounds were

given p a r e n t e r a l l y . Furthermore, avermectin B^ was much l e s s
e f f e c t i v e against Cooperia species when given p a r e n t e r a l l y
than by o r a l treatment. Avermectin B had g e n e r a l l y lower
2

a c t i v i t y against Haemonchus s p e c i e s . Examination of the B^

and B s t r u c t u r e s revealed that the d i f f e r e n c e s centered on
2

the 22,23-position. Avermectin B^ i s a 22,23-olefin whereas

i n avermectin B t h i s bond i s hydrated with the hydroxyl group
2

at the 2 3 - p o s i t i o n . The conformation of the r i n g bearing

these f u n c t i o n a l i t i e s i s d i f f e r e n t and i t was reasoned that
b i o a c t i v i t y might be l i n k e d to conformation. It therefore
became an important o b j e c t i v e to s y n t h e s i z e 22,23-
dihydroavermectin B^ which r e q u i r e d f o r i t s s y n t h e s i s the
s e l e c t i v e r e d u c t i o n of one of f i v e o l e f i n s . However, only the

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1. CAMPBELL ET AL. Discovery of Ivermectin 15

Table I I I . A c t i v i t y of Avermectin D e r i v a t i v e s against Adult

G a s t r o i n t e s t i n a l Helminths of Experimentally
Infected Sheep on O r a l A d m i n i s t r a t i o n .

a
Efficacy
Structure Dose, mg/kg H.c. ° O.c. T.a. T.c.° C. spp. O.c.

Al 0.1 2 2 0 0 2 0
A 2
0.1 3 3 3 3 0 3
Bl 0.05 3 3 3 3 3 3
B 2
0.1 0 3 3 3 3 3
H2A1 0.3 3 2 0 1 0 3
H Bi
2 0.1 3 3 3 3 3 3
BiMS 0.15 2 2 3 3 3 0
B MS
2 0.2 1 1 3 3 3 3
H2B MS 1 0.3 3 3 3 3 2 3
H BiAG
2 3.0 1 2 3 3 1 3
H4B1 0.2 0 0 1 0 0 3

a
0 = < 50%, 1 = 5 0 - 7 4 % , 2 = 7 5 -•90%, 3 = > 90% e f f i c a c y .
Abbreviations used: H.c. Haemonchus con t o r t u s ; O . c ,
O s t e r t a g i a c i r c u m c i n c t a ; T.a., T r i c h o s t r o n g y l u s a x e i ;
T . c , T r i c h o s t r o n g y l u s c o l u b r i f o r m i s ; C. spp., Cooperia
spp.; O . c , Oesophagostomum columbianum.

*MS = monosaccharide, AG = aglycon, H = 22,23-dihydro 2

d e r i v a t i v e , H 4 = 3,4,22,23-tetrahydro d e r i v a t i v e .

Benzimidazole r e s i s t a n t .

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16 PESTICIDE SYNTHESIS THROUGH RATIONAL APPROACHES

Table IV. D e r i v a t i v e s of Avermectin A~ and B^ and Anthelmintica

A c t i v i t y against T r i c h o s t r o n g y l u s c o l u b r i f o r m i s i n G e r b i l s

H 0R 5

anthelmintic
R5 R23 acta
H CH 3 H 0.05
H H H 0.0125
CH3CO CH 3
H 0.0625
H CH 3 CH CO
3 0.25
CH3CO CH 3
CH CO
3 0.5
CH3CO CH CO3 CH CO
3 0.5

aMinimal dose(mg/kg) needed to remove > 83% of the worm burden.

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1. CAMPBELL ET AL. Discovery of Ivermectin 17

22, 2 3 - o l e f i n i s c i s - s u b s t i t u t e d , suggesting the use of

Wilkinson's homogenous c a t a l y s t (Ph^P^RhCl known to be very
s e n s i t i v e to the s t e r i c environment of an o l e f i n . Hydrogéna-
t i o n of avermectin B^ f o r 20 hours using Wilkinson's c a t a l y s t
i n benzene or toluene at 25°C under one atmosphere of
hydrogen gave 22,23-dihydroavermectin B^ i n 85% y i e l d (Figure
1). T h i s compound was s e l e c t e d f o r development as a broad-
spectrum a n t i p a r a s i t i c agent f o r animals on the b a s i s of i t s
o v e r a l l e f f i c a c y by o r a l and p a r e n t e r a l routes and f o r i t s
improved s a f e t y p r o f i l e (19-21). 22,23-Dihydroavermectin B^,
c o n t a i n i n g at l e a s t 80% of 22,23-dihydroavermectin B and not
l a

more than 20% of 22,23-dihydroavermectin B ^ has been assigned

the non-proprietary name i v e r m e c t i n . The compound was
subjected to a l a r g e i n t e r n a t i o n a l program of development,
which l i e s beyond the scope of t h i s paper, and which included
e f f i c a c y t r i a l s and s a f e t y assessment i n sheep, c a t t l e ,
horses, swine and dogs. T h i s development program r e s u l t e d i n
the i n t r o d u c t i o n of ivermectin as a commercial a n t i p a r a s i t i c
agent i n 1981. For c a t t l e , sheep and horses, the dosage
recommended f o r general a n t i p a r a s i t i c use i s 0.2 mg/kg; f o r
swine the dosage i s 0.3 mg/kg. The compound i s used both
o r a l l y and p a r e n t e r a l l y — the formulation and route of
a d m i n i s t r a t i o n depending on the host species being t r e a t e d .

Discussion

The d i s c o v e r y of the avermectins, by v i r t u e of the wide

spectrum of the compounds, and t h e i r extreme potency and novel
mode of a c t i o n , met the i n i t i a l o b j e c t i v e of f i n d i n g an
a n t h e l m i n t i c with r a d i c a l l y d i f f e r e n t c h a r a c t e r i s t i c s . The
avermectins are not a c t i v e against a l l groups of helminths —
they have not been reported a c t i v e against f l u k e s or tapeworms
— but they are a c t i v e against a l l nematode groups that have
been t e s t e d , and indeed there i s no c l e a r evidence that any
species of any genus of nematode i s r e f r a c t o r y to the a c t i o n
of i v e r m e c t i n . In at l e a s t one instance (adult D i r o f i l a r i a
immitis) a p a r t i c u l a r l i f e c y c l e stage i s r e f r a c t o r y while
other stages of the same species are s u s c e p t i b l e . The
occurrance of antinematodal and antiarthropod a c t i v i t y i n a
s i n g l e chemical c l a s s , i s not e n t i r e l y unprecedented. The
organophosphates are a c t i v e against p a r a s i t e s of both groups,
but t h e i r spectrum of a c t i v i t y against nematodes i s r e l a t i v e l y
narrow. The s a l i c y l a n i l i d e compounds are a c t i v e against
c e r t a i n nematodes and arthropods but are used p r i m a r i l y
against f l u k e s .

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ACS Symposium Series; American Chemical Society: Washington, DC, 1984.
 PESTICIDE SYNTHESIS THROUGH RATIONAL APPROACHES

The discovery of avermectins r e s u l t e d from the d e l i b e r a t e

choice of fermentation products as the prime source of
substances to be submitted f o r anthelmintic screening. Many
f a c t o r s were c r i t i c a l t o the success of the venture, i n c l u d ­
i n g the submission of novel actinomycte i s o l a t e s by workers
at The K i t a s a t o I n s t i t u t e i n Japan, the s e l e c t i o n of the i n
v i v o screen, the r a p i d i s o l a t i o n and i d e n t i f i c a t i o n of the
a c t i v e p r i n c i p l e , the assessment of a n t i p a r a s i t i c p r o p e r t i e s ,
and the enhancement of b i o l o g i c a l p r o p e r t i e s by s y n t h e t i c
chemical m o d i f i c a t i o n o f the s t r u c t u r e . The names of those
r e s p o n s i b l e f o r these c o n t r i b u t i o n s may be found i n the e a r l y
papers published on the s u b j e c t , and are l i s t e d elsewhere
according to s c i e n t i f i c d i s c i p l i n e (2). The discovery of the
avermectins thus rested on e m p i r i c a l t e s t i n g — as d i d the
d i s c o v e r y of a l l other s u c c e s s f u l a n t h e l m i n t i c s and ectopara-
siticides. Such d i s c o v e r i e s are nevertheless a t t r i b u t a b l e t o
d e l i b e r a t e and f a r - f r o m - a r b i t r a r y choices made during the
i n i t i a l conception and subsequent operation of the screening
program.
The i d e n t i f i c a t i o n of a biochemical mode of a c t i o n that
appears to d i f f e r profoundly from that of previous
a n t i p a r a s i t i c agents has provided a t o o l f o r new approaches to
understanding and e x p l o i t i n g the b a s i c biochemical pathways of
animal and p l a n t p a r a s i t e s . The b i o l o g i c a l p r o p e r t i e s of the
avermectins have opened new p o s s i b i l i t i e s f o r the study of
low-dose drug d e l i v e r y systems, and many aspects of
nematodology, entomology and acarology, as w e l l as
c o n t r i b u t i n g d i r e c t l y t o the c o n t r o l of many l i v e s t o c k
p a r a s i t e s and a g r i c u l t u r a l pests.

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 PESTICIDE SYNTHESIS THROUGH RATIONAL APPROACHES

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RECEIVED April 10, 1984

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ACS Symposium Series; American Chemical Society: Washington, DC, 1984.