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REVIEW
From methylene blue to chloroquine: a brief review
of the development of an antimalarial therapy
Kristine Krafts &Ernst Hempelmann &
Agnieszka Skórska-Stania
Received: 16 February 2012 /Accepted: 27 February 2012 / Published online: 13 March 2012
#Springer-Verlag 2012
Abstract Malarial treatment is widely and readily avail-
able today. However, there was a time in the not-so-
distant past when malaria was a deadly disease with no
known cause or cure. In this article, we trace the origins
of an antimalarial therapy from the discovery of the
nature of the malarial parasite through the development
of chloroquine. We dedicate this article to Johann
“Hans”Andersag, the scientist who developed chloro-
quine, on the 110th anniversary of his birth, 16 Febru-
ary 1902.
Elucidation of the cause of malaria
In 1887, the Polish pathologist Czesław Chęciński ap-
plied a combination of methylene blue and eosin to
blood smears and discovered the daisy-like and sickle-
shaped parasites we now know as Plasmodium malariae
and Plasmodium falciparum.Untilthistime,therewere
many hypotheses regarding the causative agent of this
disease, from poisonous air (a theory mentioned as early
as 1476 by Leonardo Bruni (Bruni 1476)) to bacteria
(Klebs and Tommasi-Crudeli 1879). Three years after
Chęciński’s stain was published, the German physician
Ernst Malachowski discovered a method of polychrom-
ing methylene blue that, when used in combination with
eosin, revealed not only a range of colors within leuko-
cytes, but also the elusive purple-red nucleus of the
malarial parasite (Krafts et al. 2011). The discovery of
the malarial nucleus provided definitive proof that the
malarial organism was a eukaryote. A search for an
effective antiparasitic malarial treatment could now be-
gin in earnest.
Paul Ehrlich (1854–1915)
Paul Ehrlich (Fig. 1), a highly renowned German physician
and scientist, developed a new and effective stain for blood
in 1880 which he termed the neutralen Farbkörper or “neu-
tral stain.”This stain incorporated methylene blue and acid
fuchsin and allowed differentiation between the different
types of white blood cells. Among his remarkable accom-
plishments was his discovery that certain dyes could be used
as drugs which would kill a specific organism while leaving
other tissues unharmed.
In 1891, Ehrlich discovered that methylene blue fell
into this category of “magic bullet”drugs, in this case
targeting the malarial organism (Fig. 2). Until this point
in time, the primary treatment for malaria was quinine
(Fig. 3a), a natural substance derived from the cinchona
tree of South America and thus limited in supply. Ehr-
lich’s introduction of methylene blue (Fig. 3b), a syn-
thetic compound, allowed large-scale production of
antimalarial therapy, unlimited by the supply or location
of natural resources.
K. Krafts (*)
Department of Pathology,
University of Minnesota School of Medicine,
Duluth, MN, USA
e-mail: kristinekrafts@gmail.com
E. Hempelmann
Department of Pharmacology, Witwatersrand University,
Johannesburg, South Africa
A. Skórska-Stania
Department of Crystal Physics and Crystal Chemistry,
Faculty of Chemistry, Jagiellonian University,
Kraków, Poland
Parasitol Res (2012) 111:1–6
DOI 10.1007/s00436-012-2886-x
Author's personal copy
Fig. 1 Paul Ehrlich in 1910
Fig. 2 Figure from Guttmann and Ehrlich’s1891 paper “Ueber die Wirkung des Methylenblau bei Malaria”(On the Action of Methylene Blue on
Malaria) showing the action of methylene blue on the organism in an infected patient (Guttmann and Ehrlich 1891)
Fig. 3 Structures of relevant antimalarial drugs. aQuinine. bMethy-
lene blue. cChloroquine
2 Parasitol Res (2012) 111:1–6
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That same year, Gutmann and Ehrlich successfully
treated two patients—a domestic servant and a sailor—
with methylene blue (Guttmann and Ehrlich 1891). Ehr-
lich’s student, Röhl, continued antimalarial trials with
methylene blue at Bayer in Eberfield. However, methy-
lene blue was found to be insufficiently effective to
supplant quinine as an antimalarial drug. Röhl began
testing the company’s antimalarial compounds in birds
(Figs. 4and 5) and devised a new compound in which
one methyl group was replaced by an aminoalkyl group.
Scientists at Bayer began synthesizing a large number
of compounds, the most successful of which was termed
quinacrine (subsequently marketed as mecaprine and
atabrine), which they synthesized in 1931.
Hans Andersag (1902–1955)
Johann “Hans”Andersag (Fig. 6) was born February 16,
1902, in Lana (Meran) to Johann Andersag and Cre-
sienz Andersag (née Tribus) (Fig. 7). Johann, the short
form of the name Johannes, is derived from the Hebrew
name Yehochanan יוחנן meaning “Yah weh (JH W H ) is
gracious.”Popular in Northern Europe, especially in
Germany, the name Johannes has several variants in-
cluding, Hans (diminutized to Hänschen or Hänsel),
Hannes, Jens, and Jan. Later in his life, Andersag
changed his given name from Johann to the shortened
form Hans.
Andersag studied chemistry in Munich from 1921 to
1927 and worked at Bayer in Elberfeld until 1955 (Fig. 8).
He was married to Else Andersag née Nouvortne.
In July, 1934, Andersag modified atabrine by replac-
ing its acridine ring with a quinoline ring. The resulting
compound, which would later be termed chloroquine
(Fig. 3c), was found to have high antimalarial activity,
and unlike methylene blue or atabrine, did not discolor
skin and eyes.
Andersag began with two compounds: oxaloacetic acid
diethylester and m-chloroaniline (Fig. 9). His process con-
sisted of the following steps (Fig. 10):
1. Condensation of m-chloroaniline with oxaloacetic acid
diethylester
2. Saponification and thermal decarboxylation
3. Reaction with phosphorus oxychloride (POCl
3
)
Fig. 4 Bayer laboratory (Pharmakologisches Labor) (courtesy, Bayer
AG Archiv)
Fig. 5 Testing antimalarial compounds using birds (Pharmakologisches
Labor) (Courtesy, Bayer AG Archiv)
Fig. 6 Hans Andersag (courtesy, Bayer AG Archiv)
Parasitol Res (2012) 111:1–6 3
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4. Substitution by a diamino group
Andersag made a salt of the base using 2,4,-dihydroxy-
benzoic acid. This salt received the name resochin, being the
RESOrcinate of a 4-aminoCHINolin.
Resochin was tested in 1935 by Bayer against blood-
induced vivax malaria in four paretics at a psychiatric clinic
in Düsseldorf but was found to be “too toxic for practical
use in humans”(Coatney 1963). Subsequently, Bayer
shelved the drug for more than 10 years, a decision that
would come to be known as “the resochin error.”
Meanwhile, Andersag continued to modify the drug to
minimize the toxic effect, producing the compound 3-
methylresochin (Dünschede 1971). This compound was
named “sontochin”and was tested at the Institute for Trop-
ical Diseases in Hamburg. By the end of 1939, over 1,100
patients with malaria had been treated with sontochin.
Both drugs, resochin and sontochin, were patented in
November 1939 (Reichspatentamt, Patentschrift Nr
683692) and later issued to The Winthrop Chemical Com-
pany through their IG Farben cartel arrangement with Bayer
(US Patent 2 233 970). Clinical trials with sontochin were
also conducted jointly by French and German scientists in
Tunisia in 1941–1943, with very impressive results. In May,
1943, drug supplies and accompanying data were handed
over by the French authorities to the Allied Forces. During
the following years, resochin was rediscovered; it was given
the name “chloroquine”by EK Marshall in November 1945
(Coatney 1963).
Chloroquine was a first-line antimalarial therapy
for many years. As is the case for most antimicrobial
drugs, resistant strains of the target organism eventually
Fig. 7 Birth certificate of
Johann “Hans”Andersag,
pages 1 and 2. (Courtesy, parish
register of Lana a.d.E.).
Translation: Parish of Lana: List
of the 1902 born and baptized
children. Name and first name:
ANDERSAG, Johann Josef.
Born: 16 February 1902, 10 ½
at night. Baptized 17 February
1902, 1 in the afternoon. Name
of the father: Johann Andersag,
born in Lana 24. Aug 1861, son
of Johann and Anna. Position of
the father: Tenant. Name of the
mother: Cresienz nee Tribus,
born in Tisens 27. Aug. 1866,
daughter of Michael and Josefa
Fig. 8 Andersag (left) at Pharmazeutisch-wissenschaftliches Labor,
Elberfeld, 6 December 1953 (courtesy, Bayer AG Archiv)
4 Parasitol Res (2012) 111:1–6
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developed. However, since the mechanism of the drug
(inhibition of hemozoin biocrystallization) involves a
host-derived drug target (which cannot be modified by
the malarial parasite), it took over 20 years for resistant
forms of malaria to develop (Hempelmann 2007). For
those 20 years, Andersag’s drug saved countless lives
Fig. 9 a,bPages from Andersag’s laboratory notebook detailing his method for the synthesis of chloroquine. (courtesy, Bayer AG Archiv)
Fig. 10 Synthesis of
chloroquine by Andersag
Parasitol Res (2012) 111:1–6 5
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and it continues to be an effective antimalarial treatment
nearly 80 years later.
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