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Investigation of some medicinal plants traditionally used for treatment of malaria in Kenya as potential sources of antimalarial drugs

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Malaria is a major public health problem in many tropical and subtropical countries and the burden of this disease is getting worse, mainly due to the increasing resistance of Plasmodium falciparum against the widely available antimalarial drugs. There is an urgent need for discovery of new antimalarial agents. Herbal medicines for the treatment of various diseases including malaria are an important part of the cultural diversity and traditions of which Kenya's biodiversity has been an integral part. Two major antimalarial drugs widely used today came originally from indigenous medical systems, that is quinine and artemisinin, from Peruvian and Chinese ancestral treatments, respectively. Thus ethnopharmacology is a very important resource in which new therapies may be discovered. The present review is an analysis of ethnopharmacological publications on antimalarial therapies from some Kenyan medicinal plants.
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Investigation of some medicinal plants traditionally used for treatment of malaria
in Kenya as potential sources of antimalarial drugs
C.N. Muthaura
a,
, J.M. Keriko
b
, S. Derese
c
, A. Yenesew
c
, G.M. Rukunga
a
a
Centre for Traditional Medicine and Drug Research, Kenya Medical Research Institute, P.O. Box 54840, Nairobi, Kenya
b
Department of Chemistry, Jomo Kenyatta University of Agriculture and Technology, P.O. Box 62000, Nairobi, Kenya
c
Department of Chemistry, University of Nairobi, P.O. Box 30197, Nairobi, Kenya
article info
Article history:
Received 19 February 2010
Received in revised form 1 November 2010
Accepted 9 November 2010
Available online 21 November 2010
Keywords:
Ethnopharmacology
Kenyan antimalarial plants
Antiplasmodial screening
abstract
Malaria is a major public health problem in many tropical and subtropical countries and the burden of
this disease is getting worse, mainly due to the increasing resistance of Plasmodium falciparum against
the widely available antimalarial drugs. There is an urgent need for discovery of new antimalarial agents.
Herbal medicines for the treatment of various diseases including malaria are an important part of the cul-
tural diversity and traditions of which Kenya
0
s biodiversity has been an integral part. Two major antima-
larial drugs widely used today came originally from indigenous medical systems, that is quinine and
artemisinin, from Peruvian and Chinese ancestral treatments, respectively. Thus ethnopharmacology is
a very important resource in which new therapies may be discovered. The present review is an analysis
of ethnopharmacological publications on antimalarial therapies from some Kenyan medicinal plants.
Ó2010 Elsevier Inc. All rights reserved.
1. Malaria in Kenya
Malaria constitutes one of the biggest health problems in trop-
ical Africa and is slowly spreading to hitherto non-malaria areas
(Trape, 2002). The emergence of resistant parasites, changes in cli-
matic conditions over a large part of Africa, changes in land use and
population migration (Foster, 1991; Ridley, 1997) are extending
the areas of malaria transmission, which requires innovative strat-
egies for malaria and the mosquito vector control. Malaria is also
endemic in South-east Asia, in Central and South America and Oce-
ania. After the African countries, India and Brazil are presently the
regions of highest endemicity in the World (WHO, 1997). It is esti-
mated that the malaria incidence range between 350 and 500 mil-
lion cases with 90% of these being in tropical Africa (WHO, 2005).
In Kenya, more than 90% of malaria is caused by Plasmodium fal-
ciparum (Khaemba et al., 1994) transmitted by Anopheles gambiae
which is the most widespread in Africa and difficult to control.
Each year, there are over 8.2 million malaria infections in Kenya
(Jean-Marie, 2002) mostly due to inadequate medical care, failure
to use insecticide treated nets and increased resistance of the par-
asites to drugs. The disease accounts for 30% of all the outpatient
cases and 19% of all admissions, 5.1% of whom die, and 72 children
below the age of 5 years die daily (DMS, 2006; WHO, 1996;
Mouchet, 1999).
The disease is endemic in the lowlands, particularly the coastal
strip and Lake Victoria basin where transmission is sufficiently in-
tense. Both incidence and prevalence of infection reach more than
90% of the population within 10–12 weeks after the beginning of
the rainy season (Hoffman et al., 1996).
Malaria transmission patterns in Kenya are influenced by sev-
eral factors such as rainfall, relative humidity, vector species, and
intensity of biting, altitude and presence of susceptible new human
hosts. Patterns of endemicity are described in terms of stable,
unstable, epidemic and malaria-free zones (MoH, 1992); stable
malaria occurs in zones which have continuously high transmis-
sion rate throughout the year and includes the coastal strip and
western parts of Kenya. The population exhibits a high degree of
immunity and epidemics are highly unlikely. Unstable malaria oc-
curs where there is seasonal endemicity with one or two annual
transmission peaks and includes parts of Eastern Province and
some parts of the Rift Valley. Epidemic malaria occurs in highland
areas bordering endemic zones. Since 1988 there has been sporadic
highland epidemics and considerable child mortality reported in
Uasin Gishu, Nandi, Kericho, Kisii and Nyamira districts. Malaria-
free zones include all land that lies at attitudes 1600 m above sea
level, but due to the critical epidemiological situation of this dis-
ease very few areas are safe (MoH, 1992). A major impact of the
disease was documented in the highlands of East Africa, where
the spread of chloroquine (CQ) resistance was probably the only
factor likely to explain the changing epidemiology of malaria in
areas of low and unstable transmission, despite initial claims that
it could be attributed to global warming (Shanks et al., 2000).
0014-4894/$ - see front matter Ó2010 Elsevier Inc. All rights reserved.
doi:10.1016/j.exppara.2010.11.004
Corresponding author. Address: Kenya Medical Research Institute, P.O. Box
70174, 00400 Nairobi, Kenya. Fax: +254 20 2720030.
E-mail address: cmuthaura@kemri.org (C.N. Muthaura).
Experimental Parasitology 127 (2011) 609–626
Contents lists available at ScienceDirect
Experimental Parasitology
journal homepage: www.elsevier.com/locate/yexpr
2. The flora and fauna of Kenyan biodiversity
The vegetation and animal life of Kenya reflect the variety of its
topography and climate, which also leads to a wide variation in
malaria and subsequent disease epidemiology (Ochola, 2003).
From the coastal (Kwale and Kilifi disticts) mangrove swamps
and rain forest to the vast plains of the hinterland covered with
grass giving way to typical tropical savanna and mountain forest.
The highland areas to the west of the country are densely forested
with bamboo, dense undergrowth and timber. The equatorial al-
pine climate around Mount Kenya (3000–5200 m above sea level),
the second highest mountain in Africa, borders Meru district and
influences the natural conditions in the district leading to a wide
variety of microclimates and agroecological zones with a wide
range of flora and fauna. Wildlife of great variety is to be found
in Kenya, both in the sparsely populated areas and in the National
Parks and Reserves that have been created for its protection. For-
ests occupy about 2–3% of Kenya’s land area and yet, they are res-
ervoirs of biological diversity (genes, species and ecosystems).
Many of the plant species have medicinal value, with an estimate
of over 400 plant species used for management of common dis-
eases in East Africa (Kokwaro, 1993; Gachathi, 1989). The forests
like Kakamega, Aberdares, Mount Kenya and Arabuko Sokoke have
exemplary display of flora and fauna. The biological diversity they
carry is important because it contributes directly to the well being
of Kenyans, especially those in the rural areas, and indirectly to the
mainly agricultural economy. Harversting of medicinal plants and
other forest products is strictly controlled by the Kenya Wildlife
Service that oversees the conservation of the forests and its wild-
life. However, human expansionist demands can be expected to
wreak environmental deterioration and biotic destruction well into
the next century. Kenya’s strategy for conservation of forests in-
volves intensification of timber and other non-wood products out-
side forest areas (Njuguna et al., 2000).
The Taita Hills, in the coastal region of Kenya (south-east Kenya,
03
o
20
0
S, 38
o
150
0
E) is the northernmost extreme of the Eastern Arc
Mountains, a biodiversity hotspot chain of mountains that run
from south-eastern Kenya to southern Tanzania (Lovett and Was-
ser, 1993) and boosts an extremely high diversity of flora and fauna
(Mittermeier et al., 1998). Indigenous cloud forest in the Taita Hills
currently covers an area of 430 ha, reflecting 98% forest reduction
over the last 200 years, mainly due to clearance for agricultural
purposes (Myers et al., 2000). Forest clearance is less widespread
at present, but despite the small size of the 12 remaining indige-
nous forest fragments, they are of global conservation importance,
holding numerous rare and endemic plants and animals. Conserva-
tion effort is being put in place through a multi-disciplinary ap-
proach that seeks better legal and law enforcement co-ordination
between the Government, Local Authorities, scientists and the local
community (Githiru and Lens, 2004).
3. The role of herbal medicine for treatment of malaria
The use of traditional and herbal remedies seems to be the
alternative choice of treatment in countries where malaria is ende-
mic (Sofowora, 1982; Rasoanaivo et al., 1992; Gessler et al., 1995).
In the Third World, 80% of people are thought to rely on herbal
remedies (Zirihi et al., 2005; WHO, 2002). Local medicinal plants
continue to be used in the treatment of malaria and the evaluation
of antimalarial activity of medicinal plants against P. falciparum has
been extensively studied (O’Neill et al., 1985; Carvalho et al., 1991;
Gakunju et al., 1995; Gessler et al., 1994; Basco et al., 1994;
Andrade-Neto et al., 2003; Tran et al., 2003; Simonsen et al.,
2001). Reviews for those studies have been undertaken in many
countries: in South Africa (Pillay et al., 2008), in West Africa (Soh
and Benoit-Vical, 2007), in Brazil (Krettli et al., 2001) and else-
where (Kaur et al., 2009).
In Asia, Latin America and Africa, the extensive use of natural
plants as primary health remedies, due to their pharmacological
properties, is quite common (Conco, 1991; Phillipson et al.,
1987). The Brazilian flora, just like the Kenyan which is transversed
by the equator and rich in biodiversity offers a wide variety of bio-
active substances (Brandão et al., 1992). Many people in these
areas rely on traditional medicine for the treatment of many infec-
tious diseases, including malaria (de Mesquita et al., 2007). South-
East Asia is home to the most drug resistant parasites in the world,
both P. falciparum and Plasmodium vivax (WHO, 2001) and tradi-
tional medicine is also widely practiced. Vietnamese traditional
medicine is similar in many respects to the traditional Chinese
school having incorporated Chinese and Indian teaching (Nghiem,
2002).
In Africa herbal medicines are an important part of the culture
and traditions of its people and its biodiversity has played major
specific roles in the cultural evolution of human societies (Mugabe
and Clark, 1998). Apart from their cultural significance, traditional
medicines have been accessible and affordable and most people in
Kenya especially in rural areas use traditional medicine and
medicinal plants to treat many diseases including malaria (Njoroge
and Bussmann, 2006). It is estimated that there is one traditional
healer for every 200–400 people in Uganda, one of East African
countries. This contrasts sharply with the availability of trained
medical personnel for which the ratio is 1:20,000 or less (WHO,
2002a).
The role of ethnopharmacology is to give direction on the plant
species for selection as well as data for plant preparation, posology,
effects and side effects which could provide specific targets for iso-
lation of active compounds and pharmacological investigation in
the quest for development of new pharmaceuticals (Cox and Balick,
1994). Recent work on African plants used in the treatment of ma-
laria is very encouraging. It is striking how many different plants
are reported by herbalists to cure malaria. When we compare anti-
malarial species being used in Kenya with those recently reported
to be used in Ivory Coast, Ghana, Madagascar, and Sudan, only
Azadirachta indica (Neem tree) is shared (Benoit-Vical et al.,
1998; Addae-Kyereme et al., 2001; Rasoanaivo et al., 1999; El Tahir
et al., 1999). However, the chemical composition of the various
plant constituents is affected by the climatic conditions and the
locality under which the plant species are growing (Gessler et al.,
1995). The challenge will be to translate herbal medicine practice
with these plants into an evidence-based monotherapy or
combined therapy as suggested by Rasoanaivo et al. (1999). There
is need therefore, to corroborate with traditional healers and
clinicians for observational retrospective treatment-outcome and
prospective clinical study of a traditional medicine. The administra-
tion of a traditional treatment (e.g. a plant preparation) as a decoc-
tion/concoction, and the systematic follow up of the outcome in a
clinical study with the effect of a rapid and complete cure, without
failure and or serious side effects, would lead to further research of
the product with a view to isolating active constituents that would
form the basis of a monotherapy or combination therapy.
The use of medicinal herbs in traditional human societies has
primarily been for treatment of active or acute conditions whereas
in most instances, the reported use of medicinal herbs by animals
appears to be prophylactic (Hart, 2005). During epidemics some
medicinal preparations may be used as preventive and other prep-
arations used as tonics to enhance better health.
The traditional culture of the Maasai pastoralists of Kenya has
outlived the present day development. They are known to ingest
wild plant materials as foods, as regular ingredients of milk and
meat based soups and as herbal medicines or as flavours and tonics
for prevention or allevation of a range of common ailments (Johns
610 C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626
et al., 1999). Perhaps the most widespread prophylactic use of
medicinal herbs in humans is the use of spices in food preparation.
Evidence for the antimicrobial effects of spices in reducing the
growth of ingested foodborne pathogens and in the preservation
of meat, has been well documented (Billing and Sherman 1998).
There are no available causal prophylaxis drugs to prevent ma-
laria in endemic areas. Primaquine, the only drug specifically
developed to inhibit the liver infection, has been curtailed by the
associated toxicity, poor compliance, and increased risk of hemoly-
sis when administered to persons with glucose-6-phosphate
dehydrogenase deficiency (Carraz et al., 2006). The existence of
medicinal plants used for prophylaxis also seems to be rare.
Strychnopsis thouarsii an endemic plant in Madagascar has been
used for malaria prophylaxis (Boiteau, 1986). Carraz et al. (2006)
isolated a compound tazopsine from the plant that had activity
against the erythrocytic and the initial pre-erythrocytic phase of
malaria infection. However, on derivativation of tazopsine to
achieve N-cyclopentyl-tazopsine (NCP-tazopsine), the latter was
shown to be specifically active against the in vitro liver stage but
inactive against the blood forms of the malaria parasite, placing
this lead molecule into a novel category of antimalarial compounds
with true causal prophylactic activity. Ampelozyziphus amazonicus
is another plant used in some regions of the Amazon to prevent
malaria infection (Brandão et al., 1992). Using newly established
methods to screen for drugs that inhibit sporozoites and/or liver
stage parasites, Andrade-Neto et al. (2008) demonstrated evident
activity of its ethanolic root bark extract against pre-erythrocytic
forms of Plasmodium berghei sporozoite cultures in hepatoma cells
in vitro and in vivo.
Demonstration of efficacy for prophylactic use has been prob-
lematic because the behaviour is based on reducing the severity
or likelihood of adverse events in the future. In antimalarial drug
discovery programs, routine screening for anti-pre-erythrocytic
stage activity is seldom carried out, partly because these stages
are clinically silent, but also because access to pre-erythrocytic
parasites and purified infected hepatocytes is costly and restricted
to a few laboratories. However, demonstration of efficacy for the
treatment of acute infections has been straightforward because
plant extracts or isolated compounds can be tested in vitro or
in vivo using laboratory experimental models, which have been
easier to develop (Hart, 2005).
Traditional medicines are a potential rich source of new drugs
against malaria and other infectious diseases and given the
remarkable antimalarial properties of Cinchona bark that have been
known for more than 300 years, resulting in the discovery of qui-
nine (Camacho et al., 2000) and the more recent development of
artemisinin derivatives has re-affirmed the potential of plant spe-
cies to provide effective drugs for the treatment of malaria. Arte-
misinin, a sesquiterpene lactone was isolated from the herb
Artemisia annua in China in 1971 and was highly unusual as it con-
tained an endoperoxide moiety in contrast to known antimalarial
drugs (Wright, 2005). Its discovery heralded a new era in antima-
larial drug development as the compound and its synthetic deriv-
atives such as artemether and artesunate were rapidly effective
against parasites resistant to other antimalarials. The compounds
had gametocyticidal activity, thus were able to reduce transmis-
sion to the mosquito vector (Wright and Warhurst, 2002).
Despite the cost and adverse effects, a standard treatment for
severe malaria in Africa is the intravenous administration of qui-
nine (WHO, 2001) and resistance against the drug in Africa has
not been reported (Le Bras et al., 2006). Resistance of P. falciparum
to current antimalarial drugs (Trape, 2002), coupled with unavail-
ability and unaffordability of these agents (Bathurst and Hentschel,
2006), in addition to lack of new therapeutic agents (Benoit-Vical,
2005; Mutabingwa, 2005) has led the Government of Kenya to
provide free and subsidized Artemisinin based combination ther-
apy (ACT) in public health facilities and private pharmacies,
respectively, as first line of treatment for uncomplicated malaria.
However, hopes that artemisinins will have a major impact on
malaria have been tempered by a recent study in which a number
of clinical isolates of P. falciparum from malaria patients showed
resistance to artemether (Jambou et al., 2005). The World Health
Organization have called for an immediate halt to the marketing
and sale of malaria medicines that contain only artemisinin or
one of its derivatives (WHO, 2006) and recommended the use of
ACTs to minimize the risk of resistance development. Even so,
there is evidence that resistance to lumefantrine–artemether
may be developing in Zanzibar after a short period of use (Sisowath
et al., 2005). This underlines the urgent need for continued search
of new antimalarial drugs from medicinal plants.
It has long been recognized that natural product structures have
the characteristics of high chemical diversity, biochemical specific-
ity, and other molecular properties that make them favourable
structures for drug discovery, and which serve to differentiate
them from libraries of synthetic and combinatorial compounds
(Clardy and Walsh, 2004).
The new drug discovery approaches need to take into account
some specific concerns, in particular, the requirement for new
therapies to be inexpensive and simple to use, as well as the need
to limit the cost of drug research in as much as the new therapy
would be a drug of mass treatment, affordable to the poor who
are most vulnerable to the disease. (Bathurst and Hentschel, 2006).
A number of ethnobotanical surveys have described several
plant species traditionally used for the management of malaria in
Kenya (Muthaura et al., 2007c,d). However, only about 20% of the
plants with claimed bioactivities have been subjected to bioassay
screening (Houghton, 2001). Among the currently ongoing efforts
is the discovery of new antimalarial leads from natural products
(Rukunga et al., 2007; Murata et al., 2008; Yenesew et al., 2003;
2004; Oketch-Rabah et al., 2000). Kenya is rich in green tropical
vegetation cover and its biodiversity nature and long history of tra-
ditional plant uses are claimed to possess medicinal value. The
search for new bioactive plant products can follow three different
ways of approach for the selection of medicinal plants: random,
chemotaxonomical and ethnopharmacological where the chances
for research success are greater (Trotter et al., 1982; Elisabetsky
and Wannmacher, 1993). This review is an analysis of ethnophar-
macological publications describing research into antimalarial
therapies.
4. Methods and selection of articles
The articles selected concerned studies on in vitro or in vivo
antimalarial activity of medicinal plants from Kenya. The publica-
tions cited were sourced from databases such as PubMed and have
used classical methodologies such as the continuous culture of P.
falciparum strains (Trager and Jensen, 1976); the in vitro antimalar-
ial activity tests using radio-isotopic methods (Desjardins et al.,
1979); the in vivo antiplasmodial assays with the reference for
blood schizonticidal activity of plant extracts using the classical
4-day suppressive test by Peters and Robinson (1992) on a rodent
malaria model. The cytotoxicity concerned in vitro known cell-lines
using standard protocols for example, 3-(4,5-Dimethylthiazol-2-
yl)-2,5-diphenyltetrazolium bromide (MTT) assay (Mosmann,
1983). MTT reduction is one of the most frequently used methods
for measuring cell proliferation and cytotoxicity assays. The ratio of
cytotoxicity to biological activity is defined as selectivity index (SI)
and it is generally considered that biological efficacy is not due to
the in vitro cytotoxicity when SI P10 (Vonthron-Senecheau et al.,
2003).
C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626 611
5. Antiplasmodial screening
The development of a culture system for P. falciparum (Trager
and Jensen, 1976) was quickly exploited to test antimalarial agents
in vitro (Desjardins et al., 1979). The publication of O’Neill et al.
(1985) was among the first report that demonstrated the use of
an in vitro assay using P. falciparum for the bioassay-guided frac-
tionation of plant extracts in the search for new antimalarial drugs.
He determined the ratio of in vitro antimalarial activity and in vitro
toxicity before any in vivo work. From this there followed many
research papers (Weenen et al., 1990; Gakunju et al., 1995;
Omulokoli et al., 1997; Muregi et al., 2003).
These tools have enabled many scientific teams to evaluate the
efficacy of Kenyan medicinal plants. The evaluations sought to
establish the rationale for the use of medicinal plants traditionally
used for treatment of malaria and the potential of their constitu-
ents as possible new antimalarial agents or leads to new antimalar-
ial compounds.
Ethnobotanical studies on some Kenyan plants with relevance
to plants used for preparation of remedies against malaria have
been undertaken (Muthaura et al., 2007c,d; Orwa et al., 2007;
Njoroge and Bussmann, 2006) and several ethnobotanical screen-
ing programs have been conducted on Kenyan medicinal plants
(Rukunga et al., 2009; Gathirwa et al., 2008; Muthaura et al.,
2007a,b; Irungu et al., 2007; Muregi et al., 2003). In addition some
plants with known antiplasmodial activity were evaluated in indi-
vidual studies leading to isolation of antiplasmodial compounds
(Rukunga et al., 2008; Yenesew et al., 2004; Kuria et al., 2002;
Oketch-Rabah et al., 1997). Table 1 shows plant species that have
been bioassayed for in vitro antiplasmodial properties in Kenya
using tritiated [
3
H]-hypoxanthine incorporation method (Desjar-
dins et al., 1979) and whose IC
50
were less than 5
l
g/ml, a value
at which plant extracts were considered to be highly active with
potential for isolation of active constituents (Clarkson et al.,
2004). In vitro cytotoxicity was measured on known cell lines
(viz. Vero E6 cells or African green monkey kidney cells and KB
cells, human oral epidermoid cancer cell line) using standard pro-
tocols such as the MTT assay (Mosmann, 1983). Table 2 shows
plant species that have been bioassayed for in vivo antimalarial
properties using a murine model infected with P. berghei (Peters
and Robinson 1992) and whose suppression of parasitaemia was
greater than 30%, a value at which plant extracts were considered
to be partially active (Muriithi et al., 2002; Carvalho et al., 1991).
Several compounds with antiplasmodial properties were isolated
through bioassay-guided fractionation of active plant extracts as
well as through phytochemical analysis and some of them are
illustrated in Table 3.
5.1. Herbal leads from Meru district
Muthaura et al. (2007b), Kirira et al. (2006) and Gathirwa et al.
(2007, 2008) in different studies evaluated ethnobotanically de-
scribed antimalarial plants, collected in Meru district for their anti-
plasmodial properties. In the first study Muthaura et al. (2007b)
evaluated 10 plants against CQ sensitive (D6) and resistant (W2)
P. falciparum clones. Four of these (40%) were found to have IC
50
<5
l
g/ml for either CQ sensitive (D6) or resistant (W2) clones
and 8 (80%) had IC
50
<10
l
g/ml. This is within the range compara-
ble to antiplasmodial activity reported for ethanolic extracts of A.
annua (IC
50
, 3.9
l
g/ml; KI resistant strain) and A. indica (IC
50
,
4.1–7.2
l
g/ml; FcBI resistant strain) in the in vitro microdilution
assay (O’Neill et al., 1985; Benoit-Vical et al., 1996; Udeinya,
1993) from which potent antiplasmodial compounds (artemisinin
and nimbolide, respectively) have been isolated. The four active ex-
tracts were the water stem bark extract of Boscia angustifolia A.
Rich. (Capparaceae) (IC
50
, 1.42
l
g/ml), methanol whole plant ex-
tract of Schkuhria pinnata (Lam) O. Ktze (Compositae) (IC
50
,
1.30
l
g/ml), methanol whole plant extract of Fuerstia africana
T.C.E. Fries (Lamiaceae) (IC
50
, 0.98
l
g/ml) and the water whole
plant extract of Ludwigia erecta (L.) Hara (Onagraceae). (IC
50
,
0.93
l
g/ml).
B. angustifolia has been reportedly used for malaria and epilepsy
in Mali and the methanol leaf extract had IC
50
of 37.6
l
g/ml against
CQ sensitive (3D7) P. falciparum strain (Bah et al., 2007). The root
bark of the plant is reported to have antibacterial activity against
a range of organisms and the presence of alkaloids and saponins
were demonstrated by Hassan et al. (2006).Koch et al. (2005) re-
ported antiplasmodial activity of chloroform stem bark extract
(IC
50
>10
l
g/ml) for the same species collected in Kajiado district
of Kenya. Polar constituents in the water extract seem to be more
active than organic extracts since Muthaura et al. (2007b) reported
a much lower activity for the methanol extract as compared to the
water extract for the same species collected in Meru district.
S. pinnata whole plant is reported to be used for malaria
(Njoroge and Bussmann, 2006); leaf decoction has been used for
malaria in Zimbabwe (Watt and Breyer-Brandwijk, 1962) and in
Peru (Ramirez et al., 1988). Munoz et al. (2000) reported antimalar-
ial activity and Pacciaroni et al. (1995) demonstrated the presence
of sesquiterpenes lactones.
Koch et al. (2005) have reported the use of F. africana for malaria
by the Maasai and confirmed its antiplasmodial activity (IC
50
,
3.76
l
g/ml; D6 sensitive strain) in addition to isolation of a previ-
ously known compound ferruginol, which exhibited antiplasmo-
dial activity (IC
50
, 1.95
l
g/ml; D6 strain) but was cytotoxic to KB
cells.
The decoction of L. erecta whole plant has been used for malaria
in East Africa (Kokwaro, 1993). There is no literature information
about this plant but the findings are interesting in that the results
show high in vitro antiplasmodial activity for water extract, which
was about five times more active than the methanol extract, but it
was the latter that showed a higher suppression of parasitaemia in
P. berghei infected mice. However, the authors observed that the
water extracts were generally less active than the methanol ex-
tracts in antiplasmodial in vitro tests, albeit water is mostly the for-
mulation that has been used in traditional medicine. On the other
hand, the water extracts exhibited relatively high suppression of
parasitaemia in P. berghei infected mice giving credence to consis-
tent reports by traditional healers that these plants are effective in
treating malaria in humans. The authors noted that the ratio of
sensitivity for the two clones was much less than that of CQ for
the four plant extracts suggesting no cross-resistance with the lat-
ter and consequently phytochemical studies of these extracts is
underway in the authors
0
laboratories in order to identify the ac-
tive principles. The SI for the extracts were high suggestive of safer
therapy.
In the second study Kirira et al. (2006) evaluated 10 plants
against CQ sensitive (NF54) and resistant (ENT30) P. falciparum
strains. Two of these (20%) were found to have IC
50
<5
l
g/ml for
CQ sensitive (NF54) strain and these were the methanol stem bark
extract of Fagaropsis angolensis (Engl.) Dale (Rutaceae) and the
methanol stem bark extract of Zanthoxylum usambarense (Engl.)
Kokwaro (Rutaceae).They were not toxic in the brine shrimp nau-
plii assay, indicative of selective toxicity. Nitidine, the most com-
mon anti-malarial benzophenanthridine alkaloid in Zanthoxylum
species (Gakunju et al., 1995), has been previously isolated from
Z. usambarense (Kato et al., 1996) and F. angolensis (Khalid and
Waterman, 1985). This may explain the observed antiplasmodial
activity.
Gathirwa et al. (2007, 2008) in two independent studies evalu-
ated 10 ethnobotanically described antimalarial plants for in vitro
antiplasmodial and in vivo antimalarial assays, singly as well as
612 C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626
Table 1
Plant extracts exhibiting high antiplasmodial activity (IC
50
<5
l
g/ml) and their cytotoxic activity.
Family Plant species Plant parts
tested
Extraction solvent; antiplasmodial
activity IC
50
(
l
g/ml) and strains
of Plasmodium falciparum used
Cytotoxicity IC
50
(
l
g/ml)
and cell-lines used
cell-lines used
Selectivity
index:
(cytotoxicity/
activity)
References
Amaranthaceae Sericocomopsis
hildebrandtii Schinz
Root bark CHCl
3
; 3.78 (D6) >20 (KB) >5.29 Koch et al. (2005)
Asteraceae Artemisia afra Jacq. Leaves MeOH; 3.98 (W2) Water; 4.65 (W2) 594.85 (Vero E6) 149.46 Gathirwa et al.
(2007
Gutenbergia cordifolia Benth. Leaves CHCl
3
; 4.40 (D6) 0.2 (KB) 0.045 Koch et al. (2005)
Apocynaceae Catharanthus roseus (L.) Don Leaves MeOH; 4.65 (D6) 167.52 (Vero E6) 36.03 Gathirwa et al.
(2007)
Burseraceae Commiphora schimperi (O. Berg)
Engl.
Inner bark CHCl
3
; 4.63 (D6) >20 (KB) >4.32 Koch et al. (2005)
Canellaceae Warburgia stuhlmannii Engl. Stem bark MeOH; 1.81 (D6), 2.33 (W2) 233 (Vero E6) 128.7 (D6) Muthaura et al.
(2007a)
W. ugandensis Sprague Stem bark CH
2
Cl
2
; 2.2 (NF54), 1.4 (KI) 0.34 (L6) 0.24 (K1) Irungu et al. (2007)
Capparaceae Boscia angustifolia A. Rich. Stem bark Water; 1.42 (D6), 4.77 (W2) 6720.0 (Vero E6) 4732.4 (D6) Muthaura et al.
(2007b)
Capparidaceae B. salicifolia Oliv. Stem bark MeOH; 1.04 (D6) Water; 3.65 (D6) 304.92 (Vero E6) 293.19 Gathirwa et al.
(2007)
Celastraceae Maytenus arbutifolia
(A. Rich.) Wilczek
Root Water; 4 (M24), 4 (K67) nd nd Gakunju et al.
(1995)
M. undata (Thunb.) Blakelock Leaves Water; 0.95 (D6), 1.90 (W2) 3645.7 (Vero E6) 3837.6 (D6) Muthaura et al.
(2007a)
M. putterlickioides (Loes) Excell and
Mendoca
Root bark MeOH; 4.41 (D6) 112.4 (Vero E6) 25.5 Muthaura et al.
(2007a)
Compositae Schkuhria pinnata
(Lam) O. Ktze
Whole plant MeOH; 1.30 (D6) 1 61.5 (Vero E6) 124.2 Muthaura et al.
(2007b)
Vernonia lasiopus O. Hoffm. Leaves CHCl
3
; 1.2 (K39), 3.4 (V1/S), 1.7
(NF54),
3.6 (ENT30) EtOAc; 1.0 (K39), 1.6
(V1/S),
1.6 (NF54), 1.4 (ENT30)
nd nd Muregi et al.
(2003)
V. lasiopus O. Hoffm Root bark CH
2
Cl
2
; 4.9 (NF54), 4.7 (KI) >90 (L6) >10.7 Irungu et al. (2007)
Cyperaceae Cyperus articulatus L. Rhizome MeOH; 4.84 (NF54) nd nd Rukunga et al.
(2009)
Euphorbiaceae C. robusta Pax, E, FZ. Leaves MeOH; 3.41 (D6) 460.29 (Vero E6) 134.98 Gathirwa et al.
(2007)
Phyllanthus reticulatus Poir. Leaves Water; 1.7 (K67) nd nd Omulokoli et al.
(1997)
Suregada zanzibariensis Baill. Leaves Water; 1.5 (K67), 1.5 (ENT36), nd nd Omulokoli et al.
(1997)
Flueggea virosa (Willd.) Voigt Leaves MeOH; 2.28 (D6), 3.64 (W2) 682.6 (Vero E6) 299.3 (D6) Muthaura et al.
(2007a)
Fabaceae Acacia mellifera (Vahl) Benth. Inner bark CHCl
3
; 4.48 (D6) >20 (KB) >4.46 Koch et al. (2005)
Lamiaceae Fuerstia africana T.C.E. Fries Whole plant MeOH; 0.98 (D6), 2.40 (W2) 954.7 (Vero E6) 974.2 (D6) Muthaura et al.
(2007b)
F. africana T.C.E. Fries Leaf CHCl
3
; 3.76 (D6) >20 (KB) >5.32 Koch et al. (2005)
Leguminosae Erythrina abyssinica DC. Roots Acetone; 0.49 (D6), 0.64 (W2) nd nd Yenesew et al.
(2003a)
Meliaceae Ekebergia capensis Sparrm. Inner bark CHCl
3
; 3.97 (D6) >20 (KB) >5.04 Koch et al. (2005)
E. capensis Sparrm. Stem bark CHCl
3
; 3.9 (K39), EtOAc; 4.7 (K39),
4.9 (ENT30), MeOH; 4.6 (K39),
Water; 3.9 (K39)
nd nd Muregi et al.
(2004)
Turraea robusta Gurke Root bark MeOH; 2.09 (D6) 24.38 (Vero E6) 11.83 Gathirwa et al.
(2008)
T. robusta Guerke Root bark MeOH; 2.4 (NF54), 3.5 (KI) nd nd Irungu et al. (2007)
Mimosaceae Albizia gummifera (JF Gmel.) C.A. Sm.Stem bark MeOH; 4.8 (ENT22), 4.0 (ENT30), 1.8
(KIL9)
nd nd Ofulla et al. (1995)
Onagraceae Ludwigia erecta (L.) Hara Whole plant MeOH; 4.10 (D6), Water; 0.93 (D6),
1.61 (W2)
3283.6 (Vero E6) 3530.7 (D6) Muthaura et al.
(2007b)
Rutaceae Fagaropsis angolensis (Engl.) Dale Stem bark MeOH; 4.68 (NF54) nd nd Kirira et al. (2006)
Zanthoxylum chalybeum Engl. Root bark MeOH; 3.14 (ENT30), Water; 2.32
(ENT30)
nd nd Rukunga et al.
(2009)
Z. usambarense (Engl.) Stem bark MeOH; 3.20 (NF54) nd nd Kirira et al. (2006)
Toddalia asiatica (L.) Lam. Root bark MeOH; 0.78 (K39), 0.7 (K67), 1.46
(M24),
nd nd Gakunju et al.
(1995)
Rhamnaceae Rhamnus prinoides L
´H
´erit. Root bark CHCl
3;
3.53 (D6) >20 (KB) >5.67 Koch et al. (2005)
Ranunculaceae Clematis brachiata Thunb. Root bark CHCl
3
; 4.15 (D6) >20 (KB) >4.82 Koch et al. (2005)
Simaroubaceae Harrisonia abyssinica Oliv Stem bark CH
2
Cl
2
; 5.6 (NF54), 4.4 (KI) 32.8 (L6) 7.5 Irungu et al. (2007)
Verbenaceae Clerodendrum myricoides
(Hochst.) Vatke
Root bark MeOH; 4.0 (V1/S) nd nd Muregi et al.
(2004)
Zygophyllaceae Balanites aegyptiaca (L.) Del. Root bark CHCl
3
; 3.49 (D6) >20 (KB) >5.73 Koch et al. (2005)
nd = not done.
CQ sensitive strains: D6, NF54, K39, K67, M24.
CQ resistant strains: W2, KI, V1/S, ENT30, ENT36, KIL9.
C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626 613
Table 2
In vivo antimalarial activity of plants exhibiting suppression of parasitaemia >30% in P. berghei infected mice.
Family Plant species Plant part Parasite strain Dose mg/kg/day
(route)
Extract % Chemo
suppression
References
Amaranthaceae Cyathula schimperana Igifashi Root bark P. berghei – ANKA 100 (ip) MeOH 45.08 Gathirwa et al. (2007)
Anacardiaceae Rhus natalensis Krauss Leaves 100 (ip)
100 (ip)
MeOH
Water
56.24
83.15
Sclerocarya birrea (A. Rich.) Hochst. Stem bark 100 (ip)
100 (ip)
100 (oral)
100 (oral)
MeOH
Water
MeOH
Water
63.49
66.51
42.04
40.21
Gathirwa et al. (2008)
Lannea schweinfurthii (Engl.) Engl. Stem bark 100 (ip)
100 (ip)
100 (oral)
100 (oral)
MeOH
MeOH
Water
91.37
83.08
51.66
48.43
Apocynaceae Catharanthus roseus (L.) Don Leaves 100 (ip)
100 (ip)
MeOH
Water
34.6
42.36
Gathirwa et al. (2007)
Asteraceae Artemisia afra Jacq. Leaves 100 (ip)
100 (ip)
MeOH
Water
77.45
70.25
Caesalpinaceae Caesalpinia volkensii Harms Seed P. berghei – NK65 500 (oral) MeOH 33.3 Muregi et al. (2007b)
Canellaceae Warburgia stuhlmannii Engl. Stem bark P. berghei – ANKA 100 (ip)
100 (ip)
MeOH
Water
52.95
84.95
Muthaura et al. (2007a)
Capparidaceae Boscia salicifolia Oliv. Stem bark 100 (ip)
100 (ip)
MeOH
Water
86.50
43.75
Gathirwa et al. (2007)
Celastraceae Maytenus acuminata (L.f.) Loes. Root bark P. berghei – NK65 500 (oral) Water 33 Muregi et al. (2007a)
M. acuminata (L.f.) Loes Root bark
Leaves
500 (oral) MeOH 41.5
36.6
Muregi et al. (2007b)
M. heterophylla (Eckl. and Zeyh.) Root bark 500 (oral) Water 49 Muregi et al. (2007a)
M. undata (Thunb.) Blakelock Leaves P. berghei – ANKA 100 (ip)
100 (ip)
MeOH
Water
76.29
47.74
Muthaura et al. (2007a)
M. putterlickioides (Loes.) Excell and Mendoca Root bark 100 (ip
100 (ip)
MeOH
Water
78.66
45.31
Compositae Sphaeranthus suaveolens (Forsk.) DC. Whole plant 100 (ip)
100 (ip)
MeOH
Water
46.74
45.38
Muthaura et al. (2007b)
Schkuhria pinnata (Lam) O. Ktze Whole plant 100 (ip)
100 (ip)
MeOH
Water
49.9
64.22
Vernonia lasiopus O. Hoffm. Root bark P. berghei – NK65 500 (oral) Water 54 Muregi et al. (2007a)
V. lasiopus O. Hoffm. Root bark 500 (oral) MeOH 59.3 Muregi et al. (2007b)
Euphorbiaceae Boscia angustifolia A. Rich. Stem bark P. berghei – ANKA 100 (ip) MeOH 60.12 Muthaura et al. (2007b)
Clutia abyssinica Jaub and Spach Leaves 100 (ip)
100 (ip)
MeOH
Water
40.45
71.69
C. robusta Pax, E, FZ. Leaves 100 (ip) Water 42.35 Gathirwa et al. (2007)
Flueggea virosa (Willd.) Voigt Leaves 100 (ip)
100 (ip)
MeOH
Water
70.91
35.15
Muthaura et al. (2007a)
Guttiferae Harunganamadagascariensis Poir Leaves 100 (ip)
100 (ip)
MeOH
Water
53.13
88.04
Lauraceae Ocotea usambarensis Engl. Stem bark 100 (ip)
100 (ip)
MeOH
Water
42.20
31.70
Muthaura et al. (2007b)
Lamiaceae Fuerstia africana T.C.E. Fries Whole plant 100 (ip)
100 (ip)
MeOH
Water
61.85
43.16
Meliaceae Ekebergia capensis Sparrm. Stem bark P. berghei – NK65 500 (oral) MeOH 33.3 Muregi et al. (2007b)
Turraea robusta Guerke Root bark P. berghei – ANKA 100 (ip)
100 (ip)
100 (oral)
100 (oral)
MeOH
Water
MeOH
Water
78.2
63.81
52.24
48.83
Gathirwa et al. (2008)
Mimosaceae Albizia gummifera (JF Gmel.) C.A. Sm. Stem bark P. berghei – NK65 500 (oral) MeOH 31.7 Muregi et al. (2007b)
Moraceae Ficus sur Forssk. Root bark
Stem bark
Leaves
500 (oral)
500 (oral)
500 (oral)
MeOH
MeOH
MeOH
48.8
34.1
43.9
Olacaceae Ximenia americana L. Root bark P. berghei – ANKA 100 (ip)
100 (ip)
MeOH
Water
36.49
21.55
Gathirwa et al. (2007)
Onagraceae Ludwigia erecta (L.) Hara Whole plant 100 (ip)
100 (ip)
MeOH
Water
65.28
49.64
Muthaura et al. (2007b)
Rhamnaceae Rhamnus prinoides L
´Hérit Root bark
Leaves
P. berghei – NK65 500 (oral) Water 51 33 Muregi et al. (2007a)
R. prinoides L
´H
´erit Root bark
Leaves
500 (oral) MeOH 34.1
43.9
Muregi et al. (2007b)
R. staddo A. Rich. Root bark 500 (oral) MeOH 48.1
R. staddo A.Rich. Root bark
Leaves
500 (oral) Water 42 33 Muregi et al. (2007a)
Rubiaceae Vangueria acutiloba Robyns Leaves P. berghei – ANKA 100 (ip) Water 39.02 Muthaura et al. (2007b)
Rutaceae Toddalia asiatica (L.) Lam. Root bark
Leaves
P. berghei – NK65 500 (oral) MeOH 59.3
37.0
Muregi et al. (2007b)
614 C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626
in combination with other extracts. In the first study the methanol
and water leaves extract of Artemisia afra Jacq. (Asteraceae), the
methanol and water stem bark extract of Boscia salicifolia Oliv.
(Capparidaceae), the methanol leaves extract of Catharanthus ro-
seus (L.) G. Don (Apocynaceae) and the methanol leaves extract
of Clutia robusta Pax, E, FZ. (Euphorbiaceae) gave IC
50
s<5
l
g/ml
for either CQ sensitive (D6) or resistant (W2) P. falciparum clones.
All the extracts showed a SI > 10. The pounded leaves of B. salicifo-
lia form a remedy for fever in cattle and the stem bark has been
used for backache (Kokwaro, 1993; Beentje, 1994). In Zimbabwe,
Kazembe and Nkomo (2010) have confirmed its traditional use as
a mosquitocide while Walter and Séquin (1990) reported the pres-
ence of flavonol glycosides from the leaves. C. roseus root bark is
reportedly used to treat gastrointestinal problems among the Luo
of Kenya (Johns et al., 1995; Kokwaro, 1993) and malaria or fever
in Brazil (Brandão et al., 1985). The leaves of this species are the
only source of the indolomonoterpenic alkaloids, vincristin (leuro-
cristine) and vinblastin (vincaleucoblastine), which are anticancer
agents (Ferreres et al., 2010). Weenen et al. (1990) reported insig-
nificant antiplasmodial activity of C. robusta from Tanzania, and the
Maasai use it for wound healing (Beentje, 1994).
Of particular interest was the A. afra which grows in upland
bushes and forest edges in Kenya. Its closely related species, A. an-
nua, is known to contain potent antimalarial molecule artemisinin,
which proved to be active against CQ sensitive and resistant P.fal-
ciparum (Phillipson and Wright, 1991). A. afra is reported to show
activity against P. falciparum (Kraft et al., 2003; Van Zyl and Viljoen,
2003; Clarkson et al., 2004). It was interesting to note that the
methanol and water extracts showed twofold activity for the resis-
tant strain W2 (3.98 and 4.65
l
g/ml, respectively) compared to the
sensitive strain D6 (9.04 and 11.23
l
g/ml, respectively) and that it
was the less polar extract that was more active. The constituents of
an extract may have different mechanisms of action. An alkaloidal
fraction from Cissampelos andromorpha DC. (Menispermaceae), a
Brazilian medicinal plant, was found to be active against CQ-resis-
tant (K1) but inactive against CQ sensitive (Palo Alto) P. falciparum
strains (Fischer et al., 2004).
In vivo parasitaemia suppression of P. berghei in mice for the
methanol and water extracts was significant (77% and 70%, respec-
tively). Hot water extracts of A. annua had no activity on rodent
malaria parasite P. berghei in contrast with ether extracts, which
led to isolation of artemisinin (Wright, 2005), casting doubt to
the similarity of the active constituents in the two plant species.
Indeed, there are no reports that A. afra contains artemisinin or
any of its derivatives and this was confirmed by a recent study
by Van der Kooy et al. (2008) who found out that artemisinin is
an important chemical marker to differentiate between the two
species.
Kraft et al. (2003) demonstrated the activity of a lipophilic (pet-
rol ether/ethylacetate 1:1) extract of A. afra against a CQ-sensitive
(PoW) and resistant (Dd2) P. falciparum strains (IC
50
, 8.9 and
15.3
l
g/ml, respectively). Several flavanoids and sesquiterpene
lactones were isolated from this plant through bioassay-guided
fractionation; with acacetin (1), genkwanin (2) and 7-methoxyac-
acetin (3) exhibiting the best in vitro antiplasmodial activity
(IC
50
, 4.3–12.6
l
g/ml). The exact mechanism of antimalarial action
of flavonoids is unclear but some flavonoids have been shown to
inhibit the influx of
L
-glutamine and myoinositol into infected
erythrocytes (Elford, 1986). Exiguaflavanone A and exiguaflava-
none B from A. indica exhibited similar in vitro antiplasmodial
activities (IC
50
, 4.6 and 7.0 l
l
g/ml, respectively) (Chanphen et al.,
1998). In contrast to artemisinin from A. annua none of the com-
pounds isolated from A. afra showed extraordinary activity, and
the authors concluded that the activity of A. afra was due to the
complex mixture of substances, which may act additively or syner-
gistically (Kraft et al., 2003).
5.2. Herbal leads from Kwale district
Five ethnobotanically described antimalarial plants, collected in
Kwale district of the Kenyan Coast were evaluated against CQ sen-
sitive (D6) and resistant (W2) P. falciparum clones (Muthaura et al.,
2007a). Four of these (80%) were found to have IC
50
<5
l
g/ml for
either CQ sensitive (D6) or resistant (W2) clones and these were
the methanol leaves extract of Flueggea virosa (Willd.) Voigt
(Euphorbiaceae), methanol stem bark extract of Warburgia stuhl-
mannii Engl. (Canellaceae), water leaves extract of Maytenus undata
(Thunb.) Blakelock (Celastraceae) and methanol root bark extract
of Maytenus putterlickioides (Loes.) Excell and Mendoca (Celastra-
ceae). The methanol extracts of the Maytenus species were cyto-
toxic to Vero E6 cells in comparison to the other extracts. These
species are widely used in folk medicine as antitumour, antiasth-
matic and for stomache problems. Dihydroagarofuran sesquiter-
pene alkaloids mayteine, putterine A and putterine B have been
isolated from the root bark extract of M. putterlickioides (Schane-
berg et al., 2001). Quinone methide triterpenoids are a diverse
group of secondary metabolites from a related species Maytenus
ilicifolia (Buffa Filho et al., 2002), which have revealed potential
anti-tumour and anti-microbial activities (Bavovada et al., 1990).
The various biological activities have been attributed to those di-
verse secondary metabolites among them maytansinoids (Reider
and Roland, 1984). W. stuhlmannii is found on the Kenyan Coast
and a dichloromethane stem bark extract from its close variety
found inland, Warburgia ugandensis, had a similar antiplasmodial
activity (IC
50s
1.4 and 2.2
l
g/ml for CQ sensitive (NF54) and resis-
tant (KI) P. falciparum strains, respectively (Irungu et al., 2007). A
cytotoxic sesquiterpene, characterized as muzigadial, was isolated
from W. ugandensis. It showed in vitro trypanocidal activity and
was highly toxic in the brine shrimp assay (Olila et al., 2001).
M. undata and F. virosa are reported in South Africa having good
in vitro antiplasmodial activity (Clarkson et al., 2004) and bergenin,
an isocoumarin isolated from methanol leaves extract of F. virosa
has been reported to be antiprotozoal (Nyasse et al., 2004). The
in vivo antimalarial results in mice for the four plant extracts were
much lower than would be expected from their high activity
in vitro. The results may not necessarily correlate as reported by
Gessler et al. (1995). There is, however, good correlation in all of
the clinically used antimalarial drugs and Peters’ 4-day test is
therefore regarded as a good screen for in vivo activity. It could
be that crude extracts with their complex mixture of several
Table 2 (continued)
Family Plant species Plant part Parasite strain Dose mg/kg/day
(route)
Extract % Chemo
suppression
References
Verbenaceae Clerodendrum eriophyllum Guerke Root bark P. berghei – ANKA 100 (ip)
100 (ip)
MeOH
Water
90.13
61.54
Muthaura et al. (2007b)
C. myricoides (Hochst.) Vatke Root bark P. berghei – NK65 500 (oral) MeOH 31.7 Muregi et al. (2007b)
ip = intraperitoneal.
CQ sensitive strain: P. berghei – ANKA.
CQ resistant strain: P. berghei – NK65.
C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626 615
Table 3
Antiplasmodial activity of constituents from some Kenyan medicinal plants.
Constituents Plant species
(family)
Antimalarial
activity
IC
50
in
l
g/ml (
a
)or
l
M(
b
) and (strains
of Plasmodium
falciparum used)
Cytototoxicity
CC
50
(
l
g/ml) and
(celllines used)
Chemical structure References
Flavonoids:
acacetin (1)
Artemisia afra
Jacq.(Asteraceae)
5.5
a
(PoW),
12.6 (Dd2)
nd
O
O
OCH3
H
HO
OH
Kraft et al.
(2003)
Genkwanin (2) 5.5
a
(PoW),
8.1 (Dd2)
O
O
OH
H
H3CO
OH
7-Methoxyacacetin (3) 4.3
a
(PoW),
7.0 (Dd2)
O
O
OCH
3
H
H3CO
OH
Flavonones:
5-deoxyabyssinin II (19)
Erythrina abyssinica
DC.(Leguminosae)
13.6
b
(D6),
13.3 (W2)
nd
O
O
HO
OH
OCH 3
Prenyl
H
H
H
Yenesew
et al.
(2004)
Abyssinin III (20) 5.8
b
(D6),
5.2 (W2)
O
O
HO
OH
Prenyl
OH
H
OH
Prenyl
Abyssinone IV (21) 5.4
b
(D6),
5.9 (W2)
O
O
HO
OH
Prenyl
Prenyl
H
H
H
Abyssinone V (22) 4.9
b
(D6),
6.1 (W2)
O
O
HO
OH
Prenyl
Prenyl
H
OH
H
Sigmoidin A (23) 5.8
b
(D6),
5.9 (W2)
O
O
HO
OH
Prenyl
OH
Prenyl
OH
H
616 C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626
Table 3 (continued)
Constituents Plant species
(family)
Antimalarial
activity
IC
50
in
l
g/ml (
a
)or
l
M(
b
) and (strains
of Plasmodium
falciparum used)
Cytototoxicity
CC
50
(
l
g/ml) and
(celllines used)
Chemical structure References
Terpenoids: diterpene
ajugarin-1 (17)
Ajuga remota
Benth (Labiatae)
23.0
a
(FCA20/GHA) >88 (A431)
O
OAc
O
AcO
O
Kuria et al.
(2002)
Triterpenoid limonoid
-deacetoxy-7-oxogedunin (4)
Ekebergia capensis
Sparm.(Meliaceae)
6
a
(FCR-3) nd
O
O
O
O
O
O
Murata
et al.
(2008)
2-Hydroxymethyl-2,3,22,23-
tetrahydroxy-2,6,10,15,19,-
23-hexamethyl-6,10,14,-
18-tetracosatetraene (5)
18
a
(FCR-3),
7 (K1)
OH
HO
OH
OH
OH
Tritepene ergosterol-
5,8-endoperoxide (18)
Ajuga remota
Benth (Labiatae)
8.2
a
(FCA20/
GHA)
>23 (A431)
HO
O
O
H
Kuria et al.
(2002)
Sesquiterpenes:
mustakone (6)
Cyperus articulatus
L. (Cyperaceae)
0.14
a
(NF54),
0.25 (ENT30)
nd Rukunga
et al.
(2008)
Corymbolone (7) 1.07
a
(NF54),
1.92 (ENT30)
Spermine alkaloids:
budmunchiamine K1 (12)
Albizia gummifera
J.F. Gmel C.A. Sm
(Mimosaceae)
0.09
a
(NF54),
0.73 (ENT30)
nd
N
N
N
N
CH3
(CH2)
5(CH2)8
H
O
HCH3
CH3CH 3
Rukunga
et al.
(2007)
6-Hydroxybudmunchiamine
K2 (13)
0.15
a
(NF54),
0.91 (ENT30)
N
N
N
N
CH3
(CH
2)5(CH2
)8
CH3
O
H
CH3
CH3
OH
(continued on next page)
C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626 617
Table 3 (continued)
Constituents Plant species
(family)
Antimalarial
activity
IC
50
in
l
g/ml (
a
)or
l
M(
b
) and (strains
of Plasmodium
falciparum used)
Cytototoxicity
CC
50
(
l
g/ml) and
(celllines used)
Chemical structure References
5-Normethylbudmunchiamine
K3 (14)
0.10
a
(NF54),
0.93 (ENT30)
N
N
N
N
H
(CH
2)5(CH2)8
CH3
H
O
HCH3
CH3
6-Hydroxy-5-normethylbudmun-
chiamineK4 (15)
0.17
a
(NF54), 0.88
(ENT30)
N
N
N
N
H
(CH2)5(CH2)8
CH3
O
H
CH3
CH3
OH
9-Normethylbudmunchiamine
K5 (16)
0.12
a
(NF54),
0.89 (ENT30)
N
N
N
N
CH3
(CH
2
)5(CH2)8
CH3
H
O
HCH3
H
Alkaloid: nitidine (10) Toddalia asiatica
(L.) Lam. (Rutaceae)
0.042
a
– 0.052
(S – R)
nd
N
O
O
MeO
MeO
X
Gakunju
et al.
(1995)
Coumarins: 2
0
epicycloisobrachy-
coumarinone epoxide (8)
Vernonia
brachycalyx O.
Hoffm.(Asteraceae)
160
b
(3D7),
54 (Dd2)
313 (Human
lymphocytes)
O
O
O
H
CH
3
O
O
Oketch-
Rabah
et al.
(1997)
Cycloisobrachycoumarinone
epoxide (9)
111
b
(3D7),
54 (Dd2)
1810 (Human
lymphocytes)
O
O
O
CH3
H
O
O
5,7-dimethoxy-8-(3
1
-hydroxy-
3
1
-methyl-1
1
-butene)-
coumarin (11)
Toddalia asiatica
(L.) Lam. (Rutaceae)
16.2
a
(K39),
8.8 (VI/S)
nd
O
OH
O
OMe
MeO
Oketch-
Rabah
et al.
(2000)
618 C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626
compounds could explain their high in vitro activity due to poten-
tial synergism but the fragile chemical structure could be upset
due to biotransformation of the constituents or poor bioavailability
of the active constituents in vivo, and the results may also be
encumbered by other host factors.
5.3. Herbal leads from Kajiado district (Maasai community)
The Maasai maintain a rich traditional culture and herbal rem-
edies are an integral part of their culture. Some elder Maasai are to
be seen hawking herbal concoctions (miti ni dawa) for enhancing
better health in some of the towns outside Maasailand. Koch
et al. (2005) evaluated 21 ethnobotanically described antimalarial
plants, collected from the Maasai community of Kajiado district in
southwestern Kenya for antiplasmodial and cytotoxic properties.
Nine of the plants (42%) prepared in chloroform had IC
50
<5
l
g/
ml against CQ sensitive (D6) P. falciparum clone. These were: root
bark extract of Balanites aegyptiaca (L.) Del. (Zygophyllaceae), root
bark extract of Sericocomopsis hildebrandtii Schinz (Amarantha-
ceae), inner bark extract of Commiphora schimperi (O. Berg) Engl.
(Burseraceae), inner bark extract of Acacia mellifera (Vahl) Benth.
(Fabaceae), leaf extract of Fuerstia africana T.C.E. Fries (Lamiaceae),
leaf extract of Gutenbergia cordifolia Benth. (Asteraceae), root bark
extract of Clematis brachiata Thunb. (Ranunculaceae), inner bark
extract of Ekebergia capensis Sparrm. (Meliaceae) and root bark ex-
tract of Rhamnus prinoides L
´H
´erit. (Rhamnaceae). The cytotoxicities
of the 9 plant extracts except that of G. cordifolia on KB cells
showed IC
50
>20
l
g/ml and thus considered to have good SI for
the parasites (Likhitwitayawuid et al., 1993).
Muregi et al. (2004) found similar in vitro antiplasmodial
activity for E. capensis collected fom Kisii district in Kenya. A
limonoid compound (4), (Oxogedunin) (Bevan et al., 1963) and a
triterpenoid (5), (2-hydroxymethyl-2,3,22,23-tetrahydroxy-2,6,-
10,15,19,23-hexamethyl-6,10,14,18-tetracosatetraene) (Nishiyama
et al., 1996, 1999) both isolated from the plant showed potent IC
50
activities as low as 6 and 7
l
M, respectively (Murata et al., 2008).
Limonoids are produced by species of Meliaceae. One well
known representative from this family is A. indica, the Neem tree,
widely used as an antiplasmodial plant in Asia. Rochanakij et al.
(1985) identified nimbolide as the active antimalarial principle of
the Neem tree (EC
50
, 0.95 ng/ml, P. falciparum K1). Gedunin (IC
50
,
720 ng/ml, P. falciparum D6) and its dihydro derivative were also
found to be active in vitro (IC
50
, 2630 ng/ml) (MacKinnon et al.,
1997). Their antimalarial activities may be related to the presence
Table 3 (continued)
Constituents Plant species
(family)
Antimalarial
activity
IC
50
in
l
g/ml (
a
)or
l
M(
b
) and (strains
of Plasmodium
falciparum used)
Cytototoxicity
CC
50
(
l
g/ml) and
(celllines used)
Chemical structure References
Phenylanthraquinones:
joziknipholone A (24) and
Bulbine frutescens
(Jacq.)
Rowley
(Asphodelaceae)
0.14
a
(K1) 16.3 (L6), 10
(L5178y)
HO
O
Me
OH
HO
OP
O
OH
HO
HO
OH
OO
OMe
Me
O
HO
P
Me
HO
OH
O
Me
OMe
H
S
P
Bringmann
et al.
(2008)
B(25) 0.23
a
(K1) 17.4 (L6),
8.7 (L5178y)
HO
O
MeOH
HO
OP
O
OH
HO
HO
O
HOO
OMe
HO
Me
HO
OH
O
Me
OMe
H
S
P
Me
O
M
nd = not done.
CQ sensitive strains: PoW, K39, NF54, 3D7, FCA/20GHA, D6, FCR-3, HB3, S.
CQ resistant strains: Dd2, ENT30, V1/S, KI, W2, R.
C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626 619
of reactive groups on the ring like the carbonyl group and unsatu-
ration in C-1/C-2 positions. Benoit-Vical et al. (2003) found similar
in vitro antiplasmodial activity between iridal, a triterpenoidic
compound extracted from Iris germanica L. and an extract of A. in-
dica on P. falciparum, but there was no correlation between CQ and
iridal sensitivity against P. falciparum, indicating that the modes of
action of iridal and CQ should be different. Compound (5) showed
strong in vivo parasitemia suppression of 52.9%, at a dose of
500 mg/kg (which was unusually high concentration for a pure
compound) against a CQ-tolerant (NK65) P. berghei strain, consis-
tent with in vitro activity (IC
50
)of18
l
M and 7
l
M against CQ-sen-
sitive (FCR-3) and resistant (K1) P. falciparum strains, respectively
an indication of no cross resistance with CQ (Murata et al., 2008).
The total methanol extract of E. capensis showed in vivo parasita-
emia suppression of 33% at the same dose and parasite strain
(Muregi et al., 2007b).
B. aegyptiaca was previously reported as not possessing anti-
plasmodial activity (Weenen et al., 1990) however, Koch et al.
(2005) reported good activity. The discrepancy may be attributed
to plant parts tested, extraction procedures, georeference and sea-
sonal variation; factors which can account for the differences in ob-
served antiplasmodial activities between the various studies.
Antileishmanial and larvicidal properties have been reported
(Wiesman and Chapagain, 2006). Saponins have been isolated from
various parts of B. aegyptiaca,C. schimperi, Carissa edulis, C. brachi-
ata and Clerodendrum myricoides which may play part in antiplas-
modial activity of these plants (Chhabra et al., 1984; Reed, 1986;
Johns et al., 1999; Mohamed et al., 2002; Speroni et al., 2005).
5.4. Herbal leads from Kilifi and Tharaka districts
Rukunga et al. (2009) evaluated 12 ethnobotanically described
antimalarial plants, used traditionally for anti-malarial therapy in
Kilifi and Tharaka districts against CQ sensitive (NF54) and resis-
tant (ENT30) P. falciparum strains using
3
[H]-hypoxanthine
in vitro assay. Two of these (17%) were found to have IC
50
<5
l
g/
ml for either CQ sensitive (NF54) or resistant (ENT30) strains and
these were the methanol and water root bark extract of Zanthoxy-
lum chalybeum Engl. (Rutaceae) and the methanol rhizome extract
of Cyperus articulatus L. (Cyperaceae). Both plants were most often
cited by all the traditional healers interviewed as the most effec-
tive antimalarial remedies among the Kilifi and Tharaka people.
Both the water and methanol extracts of Z. chalybeum exhibited
higher antiplasmodial activity for the resistant strain than for the
sensitive strain suggesting a different mode of action for the con-
stituents as compared to CQ. Traditionally the stem bark, root bark
and leaves of the plant is reported to be used for malaria (Kokwaro,
1993; Gessler et al., 1994, 1995; Beentje, 1994) and species of Z.
chalybeum from different regions of Tanzania are reported to exhi-
bit high in vitro antiplasmodial activity (Gessler et al., 1994). Quin-
oline alkaloids from the root bark have been reported (Kato et al.,
1996).
The leaves of C. articulatus are reportedly used in Guinea for
cerebral malaria (Akendengue, 1992). The roots have antiplasmo-
dial activity and traditionally have been used for treatment of ma-
laria in Nigeria (Etkin, 1997). In Kenya, the rhizomes were used to
prepare traditional medicine for fever, convulsions and malaria
(Kokwaro, 1993). The extracts from the rhizomes have been shown
to selectively inhibit NMDA receptor-mediated neurotransmission
and also have anticonvulsant properties (Bum et al. 2001; 1996).
Two Sesquiterpenes, mustakone (6) and corymbolone (7) isolated
from the chloroform extract of the rhizomes of C. articulatus exhib-
ited significant antiplasmodial properties. Mustakone was approx-
imately ten times more active than corymbolone against the CQ
sensitive (NF54) P. falciparum strain (Rukunga et al., 2008). The
presence of the
a
,b-unsaturated ketone function may probably
contribute to the observed high activity, a phenomenon that has
been observed in some other sesquiterpenes isolated from the clo-
sely related species C. rotundus (Weenen et al., 1990). Some eudes-
mane sesquiterpenes, from the ethyl acetate leaves extract of
Melampodium camphoratum have been reported to show activity
in the hemin degradation assay (Chaturvedula et al., 2004). The
authors concluded that the results of this study suggested that
mustakone can be used as a ‘‘marker compound’’ in the standard-
ization of herbal medicines for malaria prepared from C. articulatus.
One of the plants from Kilifi, screened by Kirira et al. (2006)
among others from Meru district above, was A. indica or Neem
(Meliaceae), a plant growing in the coastal region of Kenya and
widely used around the world for malaria treatment, but showed
weak antiplamodial activity (IC
50
> 250
l
g/ml) for water and
methanol leaves extracts against CQ sensitive (NF54) and resistant
(ENT30) P. falciparum strains. Similar findings were reported by
Ofulla et al. (1995) for water leaves extract of A. indica
(IC
50
> 250
l
g/ml) collected elsewhere in Kenya. Studies in other
countries showed good in vitro antiplasmodial activity against P.
falciparum on extracts from Sudan (El Tahir et al., 1999), the Ivory
Coast (Benoit-Vical et al., 1996), India (Dhar, 1998) and Thailand
(Rochanakij et al., 1985). Tests showed that the cytotoxicity of
crude Neem extracts was lower than synthesized molecules (Bad-
am et al., 1987). In vivo studies in mice with P. berghei have been
uniformly disappointing (Ekanem, 1971; Obih and Makinde,
1985). Many reasons could explain these poor in vivo activities. It
may be that mice do not metabolise Neem extracts in the same
way as humans. Murine Plasmodium have different properties
and sensitivities compared with human P. falciparum and the 4-
day suppressive test may not be sufficient to evaluate plant ex-
tracts (Willcox and Bodeker, 2004).
5.5. Herbal leads from Kisii district
Muregi et al. (2003, 2004) in two separate studies evaluated 22
ethnobotanically described antimalarial plants, collected in Kisii
district against CQ sensitive (K39 and NF54) and resistant (ENT30
and V1/S) P. falciparum strains. In the first and second study, one
(4.5%) and two (9%) plant species, respectively showed IC
50
s
<5
l
g/ml for either CQ sensitive or resistant P. falciparum strains.
These were Vernonia lasiopus O. Hoffm. (Compositae), Ekebergia
capensis Sparrm. (Meliaceae) and C. myricoides (Hochst.) Vatke
(Verbenaceae). V. lasiopus was particularly active, with chloroform
and ethyl acetate extracts exhibiting IC
50
<4
l
g/ml for all the
strains tested. Irungu et al. (2007) reported a similar finding
(IC
50
<5
l
g/ml) for the dichloromethane root bark extract of V. lasi-
opus against CQ resistant (KI) P. falciparum strain. The leaf infusion
is traditionally used for malaria in Uganda (Katuura et al., 2007;
Hamill et al., 2000). V. brachycalyx, a closely related species has
been used for malaria by the Maasai, the Kipsigis, and other tribes
in East Africa for treatment of parasitic diseases (Beentje, 1994).
Two isomeric 5-methylcoumarins, 2
0
-epicycloisobrachycoumari-
none epoxide (8) and cycloisobrachycoumarinone epoxide (9),
with some antiplasmodial and antileishmanial activities have been
isolated from V. brachycalyx root extracts through bioactivity-
guided fractionation (Oketch-Rabah et al., 1997) as well as 16,17-
dihydrobrachycalyxolide which was also antiplasmodial and
antileishmanial but inhibited human lymphocytes at the same
concentration, indicating that the antiprotozoal activity was due
to general toxicity (Oketch-Rabah et al., 1998). Oxygenated sesqui-
terpene lactones are the most abundant secondary metabolites of
the genus Vernonia, and artemisinin, which is effective against
multi-drug resistant strains of P. falciparum and isolated from A.
annua belong to the same class of compounds (Oketch-Rabah,
1996; Trigg, 1989). Muregi et al. (2007b) investigated the in vivo
activity of some of the plants above and demonstrated a
620 C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626
parasitaemia suppression of 59.3% for V. lasiopus methanol root
bark extract against mice infected with P. berghei strain NK65.
5.6. The Pokot and the antimalarial leads
The Pokot traditionally are herders and inhabit a highland
plateau west of Rift Valley in Kenya. They are endowed with a rich
cultural heritage and herbal remedies are widely practiced due to
inaccessibility of modern medicine. A Pokot herbalist aged 90 years
described the use of 14 plants for the treatment of malaria and fever.
Gakunju et al. (1995) went onto evaluate their antiplasmodial activ-
ity and to isolate an antimalarial compound nitidine (10) from one of
the active plant extracts, Toddalia asiatica (L.) Lam. (Rutaceae). Of the
14 ethnopharmacologically described plants two (14%) had IC
50
<5
l
g/ml and those were T. asiatica and Maytenus arbutifolia (A Rich.)
Wilczek (Celastraceae). The active fraction of T. asiatica, with a mean
IC
50
of 51.6 ng/ml against CQ resistant (ItD12, FCR3, FCB) strains,
was more active than quinine against Kenyan parasites (IC
50
, ca.
120 ng/ml) (Pasvol et al., 1991). In addition to being antiplasmodial,
nitidine is a well-known cytotoxic agent which has received consid-
erable attention as a potential anticancer drug following the discov-
ery of potent antileukemic activity in mice (Messmer et al., 1972).
Subsequent chemical development of this compound has further
improved its potency against CQ resistant P. falciparum isolates
in vitro (Waigh, 2002). Oketch-Rabah et al. (2000) isolated a couma-
rin derivative, 5,7-dimethoxy-8-(3
0
-hydroxy-3
0
-methyl-1
0
-butene)-
coumarin (11) from the same plant which exhibited antiplasmodial
activity. The in vivo antimalarial activity of the methanolic root bark
extract showed 59.3% parasitaemia suppression of P. berghei in mice
at a dose of 500 mg/kg/day (Muregi et al., 2007b). T. asiatica is wide-
spread in Kenya and field interviews with local herbalists most often
cite this plant as important in traditional therapy against malaria
(Muthaura et al., 2007c,d; Orwa et al., 2007; Katuura et al., 2007).
5.7. Herbal leads from other places in Kenya
Omulokoli et al. (1997) evaluated four ethnobotanically de-
scribed antimalarial plants, collected in Coast and Western Prov-
inces of Kenya against CQ sensitive (K67) and resistant (ENT36)
P. falciparum strains. Two of the plants Phyllanthus reticulatus Poir.
and Suregada zanzibariensis Baill. both in Euphorbiaceae family
showed IC
50
<5
l
g/ml for the CQ sensitive (K67) and resistant
(ENT36) P. falciparum strains. Phytochemical screening revealed
classes of compounds such as alkaloids, steroids, and triterpenoids
in the two plant extracts (Omulokoli et al., 1997) and also flavonoid
glycosides (Lam et al., 2007).
Irungu et al. (2007) evaluated 14 ethnobotanically described
plants, traditionally used for antimalarial therapy in Eastern,
Central and Coast Provinces of Kenya for their ability to inhibit
the in vitro proliferation of CQ sensitive (NF54) and resistant (K1)
P. falciparum strains using
3
[H]-hypoxanthine in vitro assay. Four
of these (28.5%) were found to have IC
50
<5
l
g/ml for either CQ
sensitive (NF54) or resistant (KI) strains and these were the diclo-
romethane stem bark extract of Harrisonia abyssinica Oliv (Sima-
roubaceae), methanol root bark extract of Turrea robusta Guerke
(Melianthaceae), dichloromthane stem bark extract of W. uganden-
sis Sprague (Canellaceae) and dichloromethane stem bark extract
of V. lasiopus O. Hoffm (Compositae). The family Simaroubaceae
is known to contain quassinoids which are heavily oxygenated lac-
tones, possessing a wide spectrum of biological activities, and
among the antiplasmodial compounds isolated is simalikalactone
D from Simaba guianesis with IC
50
<1.7 ng/ml (Cabral et al.,
1993). The activity of these compounds is due to the methylene–
oxygen bridge.
Ofulla et al. (1995) evaluated 4 ethnobotanically described anti-
malarial plants from Kenya against CQ resistant (ENT22, ENT30,
KIL9) P. falciparum isolates. One of them, Albizia gummifera metha-
nol stem bark extract was found to have IC
50
<5
l
g/ml for all the
isolates. Bioassay-guided fractionation of the methanol extract
led to isolation of five bioactive known spermine alkaloids namely:
Budmunchiamine K(1) (12), 6-Hydroxybudmunchiamine K(2) (13),
5-Normethylbudmunchiamine K(3) (14), 6-Hydroxy-5-normethy-
budmunchiamine K(4) (15) and 9-Normethybudmunchiamine
K(5) (16) (Rukunga and Waterman, 1996). These alkaloids exhib-
ited activities against CQ sensitive (NF54) and resistant (ENT30)
with IC
50s
ranging from 0.09 to 0.91
l
g/ml. With the exception of
(15), the alkaloids were further evaluated for in vivo activity
against P. berghei infected mice and showed suppression of para-
sitaemia ranging from 43% to 72% (Rukunga et al., 2007). Similar
bioactive spermine alkaloids, budmunchiamines L4 and L5 (IC
50
,
14.0 and 15.0
l
M, respectively) have been isolated from the meth-
anol stem bark and leaves extracts of Albizia adinocephala. The ex-
tracts were found to inhibit the malarial enzyme plasmepsin II
(Ovenden et al., 2002). Alkaloids have been successfully used for
the treatment of parasitic infections. The outstanding example is
quinine from Cinchona succirubra (Rubiaceae) used for the treat-
ment of malaria for more than three centuries.
Kuria et al. (2001) picked out Ajuga remota as the most com-
monly used herbal medicine to treat malaria in Kenya during field
trips to herbalists’ practices in an area about 200 miles around Nai-
robi but similarly to Muregi et al. (2004) findings, the antiplasmo-
dial activity was moderate (IC
50
>10
l
g/ml). In a follow up study
Kuria et al. (2002) isolated ajugarin-1 (17) and ergosterol-5,8-
endoperoxide (18) from A. remota and evaluated their in vitro anti-
plasmodial activity. Ajugarin-1 was moderately active, with IC
50
of
23.0
l
M, against the CQ sensitive (FCA20/GHA) P. falciparum strain.
Ergosterol-5,8-endoperoxide was about 3 times as potent. Both
ajugarin-1 and ergosterol-5,8-endoperoxide did not exhibit cyto-
toxicity against A431 (skin carcinoma) cell line. Ajugarin-1 is inter-
esting in view of its unusual chemical structure (epoxide ring)
when compared to conventional antimalarial compounds. Such a
compound can be considered as a lead compound to synthesize
new pharmaceutically important derivatives with possibly higher
antimalarial activities. Ergosterol-5,8-endoperoxide has been
found to inhibit the growth of protozoan parasite of Trypanoso-
matidae family such as Trypanosome cruzi and various Leishmania
species by interfering with the integrity of the cell membrane (Liñ-
ares et al., 2006). The broad action of ergosterol endoperoxide
against several protozoal parasites and mycobacteria opens up
new possibilities for application in the treatment of several dis-
eases. Possibly the mechanism of action of ergosterol endoperoxide
could involve the formation of reactive oxygen species, that causes
disruption of the parasite membrane (Correa et al., 2006).
Among the five Erythrina species present in Kenya, E. abyssinica
is the most widely used in traditional medicine where it has been
used for treatment of malaria and microbial infections (Kokwaro,
1993). The antiplasmodial activities of the stem and root bark ex-
tracts of E. abyssinica have been reported (Yenesew et al., 2003a,
2004). The acetone extract of the roots were found to have IC
50
<5
l
g/ml for either CQ sensitive (D6) or resistant (W2) P. falcipa-
rum strain. Compounds isolated which showed antiplasmodial
activity included chalcones, isoflavonoids and flavonones, with
the latter showing higher antipasmodial activities. Some of the ac-
tive flavonones were 5-deoxyabyssinin II (19), Abyssinin III (20),
Abyssinone IV (21), Abyssinone V (22) and Sigmoidin A (23)
(Yenesew et al., 2004).
And from the roots of South African plant species Bulbine frutes-
cens (Jacq.) Rowley (Asphodelaceae) widely cultivated in other
parts of the world for aesthetic purposes, phenylanthraquinone
knipholone compounds with remarkable antiplasmodial activities
have been reported (Abegaz et al., 2002). Further work by Bring-
mann et al. (2008) led to isolation of two unprecedented dimeric
C.N. Muthaura et al. / Experimental Parasitology 127 (2011) 609–626 621
phenylanthraquinones, namely joziknipholones A (24) and B (25),
that showed high antiplasmodial activity against CQ resistant
(K1) P. falciparum strain. The antiplasmodial activity appears to
be associated intrinsically with the complete molecular array of a
phenylanthraquinone including the stereogenic axis (Abegaz
et al., 2002). The two compounds exhibited low antitumoural
and cell cytotoxicity activities against murine leukemic lymphoma
L5178y cells and rat skeletal myoblast (L6) cells, respectively.
5.8. Herbal leads and synergism
In a study for in vitro antiplasmodial activity of some plants
used in Kisii, against malaria and their CQ potentiation effects,
Muregi et al. (2003) observed that the ethyl acetate leaves extract
of V. lasiopus with CQ against the multi-drug resistant (V1/S) P. fal-
ciparum isolate showed synergistic effects. Cepharanthine, a bisb-
enzylisoquinoline alkaloid isolated from Stephania erecta was
shown to synergize with CQ by significantly reducing the IC
50
for
the CQ resistant (W2) clone but not low enough to the level of
the sensitive (D6) clone (Tamez et al., 2005). In the in vitro anti-
plasmodial combination tests between different extracts for some
plants from Meru, Gathirwa et al. (2008) showed that interaction
was mainly additive, while in the in vivo antimalarial tests some
synergistic interactions were observed resulting in unusually high
parasitaemia suppression for the extracts (p> 0.05) when com-
pared to CQ. This may explain why traditional healers use a con-
coction of different plants in preparation of herbal remedies.
Synergistic interactions between the components of individual
or mixtures of plant extracts are considered to be a vital part of their
therapeutic efficacy; however until recently there has been very
little evidence to demonstrate what herbal practitioners have al-
ways believed. The ‘‘isobole method’’ of Berenbaum (1989) seems
to be one of the most practicable experimentally and also the most
demonstrative method among all those so far proposed for the proof
of synergy effects. For example over the last two decades it has been
suggested that the efficacy of A. annua plant extract derives from a
synergistic effect and that it is a combination of constituents in the
plant which produce the total antiplasmodial activity. Several poly-
methoxyflavones such as casticin, artemetin, chrysosplenetin,
chrysosplenol-D and circilineol (Bhakuni et al., 2001) may contrib-
ute to the in vitro activity of artemisinin against P. falciparum (Bhak-
uni et al., 2001; Elford et al., 1987; Phillipson, 1999; Liu et al., 1992).
However, data concerning the mechanism related to the synergistic
effects have been scarce mainly due to the complexity of very many
compounds in a plant extract. Bilia et al. (2002) demonstrated that
the synergism of the flavonoids is related to the assisted activation
of artemisinin before its interaction with hemin.
In Mali, Nauclea latifolia was associated with Mitragyna inermis
or Guiera senegalensis and Feretia apodanthera with M. inermis (Azas
et al., 2004). In modern drug therapy synergistic interaction is to be
preferred because the magnitude of the drug efficacy is many times
enhanced with a minimal risk of selecting mutant, resistant para-
site populations (White and Olliaro, 1996). Resistance develop-
ment by infective agents to traditional therapies may be difficult
due to the activities of a complex mixture of many constituents
either in a single plant or in a mixture of plants. Drug combination
has been the way forward to counter parasite resistance in antima-
larial chemotherapy (White and Olliaro, 1996) and to minimize the
risk of resistant development (Anne et al., 2001; Olliaro and Taylor,
2003).
6. Conclusions
The ethnopharmacology approach used in search for new
antimalarial compounds appears to be predictive, and whereas
all the crude extracts are used daily for malaria treatment and
show high in vitro activity, only a few extracts showed good
in vivo efficacy, although in vivo assays reported are few in compar-
ison to the in vitro assays. The cytotoxicity and toxicity tests in ani-
mal models are even fewer albeit folk medicine has been in use for
centuries without apparent serious side effects. However, informa-
tion in this area may be lacking due to non-reporting and the pos-
sible potential genotoxic effects that follow prolonged use of the
more popular herbal remedies are a cause for alarm.
In vitro screening programmes, using the ethnobotanical ap-
proach, are important in validating the traditional use of herbal rem-
edies and for providing leads in the search fornew active principles.
Using bioassay-guided fractionation, several compounds having
biological activity have been isolated and identified. The number
of active plant extracts and compounds identified from only a small
sampling of Kenya’s medicinal flora, using a series of in vitro tests,
further emphasises the potential areas for future work.
Ideally, extracts and compounds effective at the blood stage of
the malaria parasite should have strong in vitro antiplasmodial
and in vivo antimalarial activities with good selectivity for the ma-
laria parasites in cytotoxic assays. No clinical trials have been re-
ported in the country for even the more popular medicinal plants
like A. indica, whose products are to be found dotting many super-
market shelves in both upmarket and downmarket trading areas.
The significance of the reported research lies not only in serving
to promote the value of the Kenyan flora and the quest for new
antimalarial molecules but also the need for traditional healthcare.
There is a growing awareness on the usefulness of traditional
foods and natural products and it may not be long, before a draft
policy which is already under discussion is implemented through
an act of parliament, for integration of traditional medicine with
allopathic medicine in public health facilities. This would lead to
well-controlled clinical trials that would be invaluable for locating
new antimalarial and other drugs.
Acknowledgments
This work received financial support from UNICEF/UNDP/World
Bank/WHO special programme for Research and Training in Trop-
ical Diseases (TDR). We thank the Director, KEMRI for allowing
publication of this study.
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... 17,20 More than 53000 plant species are currently reported to be used in herbal medicine worldwide, of which more than 10% are indigenous to Africa. 21 The African medicinal plants, most of which belong to the Leguminosae and Asteraceae families are used traditionally for the treatment of infectious diseases [22][23][24][25][26][27][28][29][30][31][32] and other non-communicable diseases (NCDs) such as cancer and diabetes. 21,23,27,33,34 Some of these plants have been reported to elaborate anti-infective secondary metabolites including alkaloids, flavonoids, isoflavonoids and anthraquinones, 30,[35][36][37][38][39][40] justifying their use in traditional medicine. ...
... 2,101 Symptomatic treatment is a common practice for most viral infections. 2,11 Some of the marketed antiviral drugs include antihepatitic drugs such as ribavirin (26), entecavir (27), sofosbuvir (28) and dasabuvir (29), and antiretrovirals such as abacavir (30), lamivudine (31) and zidovudine (32) used for the treatment and prevention of HIV ( Figure 6). 101 Figure 6: Some commercially available antiviral drugs ...
... for the treatment of infectious diseases[22][23][24][25][28][29][30][31][32] and other noncommunicable diseases.21,23,33,34 such as diabetes and cancer. ...
Thesis
Full-text available
Two approaches for the synthesis of prenylated isoflavones were explored: the 2,3-oxidative rearrangement/cross metathesis approach, using hypervalent iodine reagents as oxidants and the Suzuki-Miyaura cross-coupling/cross metathesis approach. Three natural prenylated isoflavones: 5-deoxy-3′-prenylbiochanin A (59), erysubin F (61) and 7-methoxyebenosin (64), and non-natural analogues: 7,4′-dimethoxy-8,3′-diprenylisoflavone (126j) and 4′-hydroxy-7-methoxy-8,3′-diprenylisoflavone (128) were synthesized for the first time via the 2,3-oxidative rearrangement/cross metathesis approach, using mono- or diallylated flavanones as key intermediates. The reaction of flavanones with hypervalent iodine reagents afforded isoflavones via a 2,3-oxidative rearrangement and the corresponding flavone isomers via a 2,3-dehydrogenation. This afforded the synthesis of 7,4′-dimethoxy-8-prenylflavone (127g), 7,4′-dimethoxy-8,3′-diprenylflavone (127j), 7,4′-dihydroxy-8,3′-diprenylflavone (129) and 4′-hydroxy-7-methoxy-8,3′-diprenylflavone (130), the non-natural regioisomers of 7-methoxyebenosin, 126j, erysubin F and 128 respectively. Three natural prenylated isoflavones: 3′-prenylbiochanin A (58), neobavaisoflavone (66) and 7-methoxyneobavaisoflavone (137) were synthesized for the first time using the Suzuki-Miyaura cross-coupling/cross metathesis approach. The structures of 3′-prenylbiochanin A (58) and 5-deoxy-3′-prenylbiochanin A (59) were confirmed by single crystal X-ray diffraction analysis. The 2,3-oxidative rearrangement approach appears to be limited to the substitution pattern on both rings A and B of the flavanone while the Suzuki-Miyaura cross-coupling approach appears to be the most suitable for the synthesis of simple isoflavones or prenylated isoflavones whose prenyl substituents or allyl groups, the substituents that are essential precursors for the prenyl side chains, can be regioselectively introduced after the construction of the isoflavone core. The chalcone-flavanone hybrids 146, 147 and 148, hybrids of the naturally occurring bioactive flavanones liquiritigenin-7-methyl ether, liquiritigenin and liquiritigenin-4′-methyl ether respectively were also synthesized for the first time, using Matsuda-Heck arylation and allylic/benzylic oxidation as key steps. The intermolecular interactions of 5-deoxy-3′-prenylbiochanin A (59) and its two closely related precursors 106a and 106b was investigated by single crystal and Hirshfeld surface analyses to comprehend their different physicochemical properties. The results indicate that the presence of strong intermolecular O-H···O hydrogen bonds and an increase in the number of π-stacking interactions increases the melting point and lowers the solubility of isoflavone derivatives. However, the strong intermolecular O-H···O hydrogen bonds have a greater effect than the π-stacking interactions. 5-Deoxy-3′-prenylbiochanin A (59), erysubin F (61) and 7,4′-dihydroxy-8,3′-diprenylflavone (129), were tested against three bacterial strains and one fungal pathogen. All the three compounds were inactive against Salmonella enterica subsp. enterica (NCTC 13349), Escherichia coli (ATCC 25922), and Candida albicans (ATCC 90028), with MIC values greater than 80.0 μM. The diprenylated isoflavone erysubin F (61) and its flavone isomer 129 showed in vitro activity against methicillin-resistant Staphylococcus aureus (MRSA, ATCC 43300) at MIC values of 15.4 and 20.5 μM, respectively. 5-Deoxy-3′-prenylbiochanin A (59) was inactive against this MRSA strain. Erysubin F (61) and its flavone isomer 129 could serve as lead compounds for the development of new alternative drugs for the treatment of MRSA infections.
... Artemisia afra commonly called African wormwood [5]- [14] is a member of the Compositae (Asteraceae) family [5] [6] [9] [10] [13]- [18]. It is one of the most widely used in African medicinal plants with essential oil components [5] [7] [19]. ...