M.T.H. Khan and A. Ather (eds.)
Lead Molecules from Natural Products
r2006 Elsevier B.V. All rights reserved.
The medicinal potential of black seed
(Nigella sativa) and its components
HALA GALI-MUHTASIB,NAHED EL-NAJJAR,REGINE SCHNEIDER-STOCK
The seeds of Nigella sativa L., commonly known as black seed, have been used in traditional medicine by
many Asian, Middle Eastern and Far Eastern Countries to treat headache, coughs, abdominal pain,
diarrhea, asthma, rheumatism and other diseases. The seeds of this plant are the most extensively studied,
both phytochemically and pharmacologically. The aqueous and oil extracts of the seeds have been shown
to possess antioxidant, antiinﬂammatory, anticancer, analgesic and antimicrobial activities. Thymoqui-
none, the most abundant constituent of black seed essential oil, has been shown to be the active principle
responsible for many of the seed’s beneﬁcial effects. This review paper describes the seed, its chemical
components and popular uses in traditional medicine. The paper also discusses the medicinal potential and
therapeutic values of some of the individual components present in the extracts of the seeds.
Keywords: medicinal plants, nigella sativa, black seed, thymoquinone, N. sativa oil
Abbreviations: BP, benzo(a)pyrene; TQ
, dithymoquinone; DOX, doxorubicin; GC, gas chromatography;
HPLC, High-performance liquid chromatography; IL, interleukin; NSO, Nigella sativa oil; PGE2, prosta-
glandin E2; TLC, thin layer chromatography; THQ, thymohydroquinone; TOH, thymol; TQ, thymoquinone;
TNF, tumor necrosis factor.
Plants are natural factories for the production of chemical compounds, many of
which are used to promote health and ﬁght diseases and some of them are marketed
as food or herbal medicines (Dubick, 1986). Herbal medicines have long been viewed
as a source of curative remedy based on religious and cultural traditions (Huxtable,
1992;Ghazanfer, 1994). The use of indigenous plant medicines in developing coun-
tries became a World Health Organization policy since 1970. Of the 520 new drugs
approved in the period 1983–1994 by either the US Food and Drug Administration
or comparable entities in other countries, 30 drugs came directly from natural
product sources, 173 were either semi-synthetics or synthetics originally modeled on
a natural parent product (De Smet, 1997).
Nigella sativa is an annual herb of the Ranunculaceae family, which grows in coun-
tries bordering the Mediterranean Sea, Pakistan and India. This widely distributed plant
is native to Arab countries and other parts of the Mediterranean region (Jansen, 1981).
For thousands of years, this plant has been used in many Asian, Middle Eastern and
Far Eastern Countries as a spice and food preservative as well as a protective and health
remedy in traditional folk medicine for the treatment of numerous disorders (Chopra et
al., 1956;Nadkarni, 1976). The seed of this plant is commonly known as black seed and
is referred to by the prophet Mohammed as having healing powers. The seeds are
commonly eaten alone or in combination with honey and in many food preparations.
The oil prepared by compressing the seeds of N. sativa is used for cooking. Black seed is
also identiﬁed as the curative black cumin in the Holy Bible, and is described as the
Melanthion of Hippocrates and Discroides and as the Gith of Pliny (Chopra et al., 1956;
Nadkarni, 1976). Other names for the seed include black caraway seed, Habbatu Sawda
and Habatul Baraka ‘‘the Blessed Seed’’.
N. sativa plant is one of the most extensively studied, both phytochemically and
pharmacologically. The extracts of N. sativa seeds have been used by patients
to suppress coughs (Mahfouz et al., 1960), disintegrate renal calculi (Hashem and
El-Kiey, 1982), retard the carcinogenic process (Hassan and El-Dakhakhny, 1992;
Worthen et al., 1998), treat abdominal pain, diarrhea, ﬂatulence and polio (Enomoto
et al., 2001), exert choleretic and uricosuric activities (El-Dakhakhny, 1965), anti-
inﬂammatory (Chakravarty, 1993;Houghton et al., 1995) and antioxidant effects
(Mansour et al., 2002). Besides, the essential oil was shown to have antihelminthic
(Agarwal et al., 1979), antinematodal (Akhtar and Riffat, 1991), antischistosomal
(Mahmoud et al., 2002), antimicrobial (Hanafy and Hatem, 1991;Aboul-Ela et al.,
1996) and antiviral (Salem and Hossain, 2000) effects. In addition, the crude oil
prepared from the seeds produce a variety of pharmacological actions such as anti-
histaminic (El-Dakhakhny, 1965;Mahfouz et al., 1965;Chakravarty, 1993), diuretic
and antihypertensive (El-Tahir et al., 1993b;Zaoui et al., 2000), hypoglycemic
(Al-Hader et al., 1993), antioxytocic (Aqel and Shaheen, 1996), antinociceptive
(Abdel-Fattah et al., 2000), respiratory stimulation (El-Tahir et al., 1993a), hemato-
logical (Enomoto et al., 2001) hepatoprotective (Daba and Abdel-Rahman, 1998)
and immunopotentiating (Swamy and Tan, 2000) effects. The latter pharmacological
properties appear to be involved in the beneﬁcial effects of N. sativa oil on headache,
ﬂatulence, blood homeostasis abnormalities, rheumatism and related inﬂammatory
diseases (Boulos, 1983). Moreover, the seeds are believed to have carminative,
stimulatory and diaphoretic properties and are used in the treatment of bronchial
asthma and eczema (Boulos, 1983). This chapter will review the medicinal potential
of N. sativa seed extracts and emphasize the reasons for its long history of use in
folklore medicine in Mediterranean countries.
II. Chemical constituents and active principles in N. sativa seeds
Millions of people in the Mediterranean region and on the Indian subcontinent use
the oil from the seed of N. sativa daily as a natural protective and curative remedy.
Lead molecules from natural products: discovery and new trends134
The seeds are very rich and diverse in chemical composition. They contain amino
acids, proteins, carbohydrates, ﬁxed and volatile oils (Khan, 1999). Many of the
pharmacological activities mentioned above have been attributed to quinone con-
stituents in the seed. As early as 1956, Chopra et al. found that thymoquinone (TQ)
(Figure 1) is the main active constituent of the volatile oil of the black seed. Mahfouz
and El-Dakhakhny (1960) were the ﬁrst to report on the isolation of ‘nigellone’ from
the oil of N. sativa seed, using Girard’s reagent. Nigellone was later found to possess
antihistaminic properties in relatively low concentrations (Mahfouz et al., 1965).
El-Dakhakhny (1963) was able to isolate the constitutive components of N. sativa
seeds from its essential oil, among which TQ was later shown to be the main con-
stituent of the volatile oil (Houghton et al., 1995). In addition, El-Dakhakhny de-
termined that the ‘nigellone’ isolated earlier was a dimer of TQ, which was later
named dithymoquinone (TQ
)(Figure 1). The latter compound was shown to be
formed via photodimerization of TQ as a consequence of exposure to sunlight during
separation and extraction of the quinones from the seed. El-Fatatry (1975) reported
the isolation of thymohydroquinone (THQ) from N. sativa seed volatile oil. In an-
other study (Aboutabl et al., 1986), the chemical composition of the black seed of N.
sativa was found to contain a ﬁxed oil (30%) and a volatile oil (average 0.5%,
maximum 1.5%). The volatile oil was found to contain 54% TQ and many mono-
terpenes such as p-cymene and a-pinene, TQ
In recent years, the seeds of N. sativa have been subjected to a range of phyto-
chemical investigations. They have been shown to contain more than 30% (w/w) of a
ﬁxed oil with 85% of total unsaturated fatty acid (Houghton et al., 1995). The seeds
also contain alkaloids of unknown pharmacological actions, such as nigellidine,
Fig. 1. Chemical structures of thymol (a), thymoquinone (b) and dithymoquinone (c) (Me:
The medicinal potential of black seed (Nigella sativa) 135
nigellimine and nigellicine (ur-Rahman et al., 1985), saponins and crude ﬁber as well
as minerals such as calcium, iron, sodium and potassium. Other constituents of the
volatile oil include thymol (TOH) (Figure 1)(Aboutabl et al., 1986). Recently, the
presence of TQ, TQ
and TOH in N. sativa seed was conﬁrmed using thin layer
chromatography (TLC) and normal phase high-performance liquid chromatography
(HPLC) methods (Abou-Basha et al., 1995;Aboul-Enein and Abou Basha, 1995).
The content of TQ in N. sativa seed oil samples, obtained from different origins, was
measured by gas chromatography (GC) analysis and found to be in the range of
0.13–0.17% w/v of the oil (Houghton et al., 1995). The seeds are also rich in proteins;
when whole N. sativa seeds were fractionated using SDS-PAGE, they were found to
contain a number of protein bands ranging from 10 to 94 kDa molecular mass (Haq
et al., 1999). An HPLC method for quantifying the putative pharmacologically ac-
tive constituents (TQ, TQ
, THQ and TOH) in the oil of N. sativa seed was recently
described by Ghosheh et al. (1999). In this procedure, the four compounds men-
tioned were separated and quantiﬁed in commercial N. sativa seed oil with good
resolution, reproducibility and sensitivity. Both heat and light are known to affect
the levels of the constituents in the oil. Since various storage and manufacturing
conditions are expected to make a difference in the amounts of the quinone con-
stituents of the oil, the analytical HPLC method described by Ghosheh et al. (1999)
can be used to quantify the levels of the above constituents in the oil and seed
extracts of N. sativa under different manufacturing conditions. The protocol is also
useful as a quality control method for the determination of pharmacologically active
quinones in N. sativa seed oil. Using TLC, the oil of black seed was found to contain
TQ and the terpenoid components carvacrol, t-anethole and 4-terpineol (Burits and
Bucar, 2000). GC-MS analysis of the essential oil obtained from six different samples
of N. sativa seeds and from a commercial ﬁxed oil showed that the qualitative
composition of the volatile compounds was almost identical. Differences were
mainly restricted to the quantitative composition (Burits and Bucar, 2000).
In conclusion, N. sativa seeds contain ﬁxed oils and volatile oils, which are rich
sources of quinones, unsaturated fatty acids, amino acids and proteins and contain
traces of alkaloids and terpenoids. Most of the studies on the biological effects of
N. sativa have dealt with its crude extracts in different solvents; however, some
studies used its active principles. Among the components isolated from the volatile
oil of N. sativa, TQ has been shown to be the principal active ingredient (Mahfouz
and El-Dakhakhny, 1960) and thus is the most studied of all. In what follows, the
physiological, antioxidant, antimicrobial, analgesic, antiinﬂammatory and chemo-
preventive effects of black seed with a special emphasis on TQ will be discussed.
III. Physiological effects of N. sativa and its component TQ
The oil extract of black seed has been shown to exert effects on various systems in-
cluding the respiratory, cardiovascular, gastric and uterine and smooth muscle. The
effects of intravenous administration of volatile oil and of TQ were investigated on the
respiratory system of the guinea pig (El-Tahir et al., 1993a). The latter compounds were
found to increase the intratracheal pressure in the dose range of 4–32 ml/kg and
1.6–6.4 mg/kg, respectively. Although N. sativa oil (NSO) significantly increased the
Lead molecules from natural products: discovery and new trends136
respiratory rate of guinea pigs, TQ was without any effect. The effects of NSO were
significantly antagonized by treatment of the animals with antihistamines such as at-
ropine and reserpine, suggesting that the oil-induced respiratory effects were mediated
via the release of histamine and indirect activation of muscarinic and cholinergic mech-
anisms (El-Tahir et al., 1993a). This also suggested that the removal of TQ from black
seed oil might provide a potential centrally acting respiratory stimulant (El-Tahir et al.,
1993a). This same group demonstrated that the intravenous administration of NSO
(4–32 ml/kg) or TQ (0.2–1.6 mg/kg) to rats decreased the arterial blood pressure and the
heart rate in a dose-dependent manner (El Tahir et al., 1993b), suggesting that the oil
may possess antihypertensive effects. The cardiovascular depressant effects of the oil
were significantly antagonized by atropine and cyproheptadine, suggesting that these
effects were mediated mainly centrally via indirect and direct mechanisms that involved
both 5-hydroxy tryptaminergic and muscarinic mechanisms (El-Tahir et al., 1993b).
NSO has also been shown to increase bile secretion in dogs and uric acid in rats as well
as protect guinea pigs against histamine-induced bronchospasm (El-Dakhakhany,
1982). The fatty and petroleum extracts shortened bleeding time and inhibited ﬁbrin-
olytic activity in rabbits (Ghoneim et al., 1982). In a recent study, the crude extract of N.
sativa seeds was found to exhibit spasmolytic and bronchodilator activities mediated
possibly through calcium channel blockade and this activity was concentrated in the
organic fraction of the extract (Gilani et al., 2001).
Traditionally N. sativa plant has been in use in many Middle Eastern countries as
a natural remedy for diabetes. Significant reduction in blood glucose and cholesterol
levels in humans following the use of the plant was reported by Bamosa et al. (1997).
The oil of this plant has a great potential in the treatment of diabetic animals because
of its combined hypoglycemic (Al-Hader et al., 1993;Zaoui et al., 2002a) and
immunopotentiating properties (Haq et al., 1999). A plant extract mixture compris-
ing N. sativa, myrrh, gum Olibanum, gum asafetida and aloe was found to lower
blood glucose in streptozotocin diabetic rats (Al-Awadi et al., 1991). In an attempt
to elucidate the mechanism of this antidiabetic action, the rate of gluconeogenesis in
isolated hepatocytes as well as the activity of pyruvate carboxylase and phosphoenol
pyruvate carboxykinase in rat liver homogenates was examined. It was found that
the plant extracts significantly decreased hepatic gluconeogenesis, suggesting that it
may prove to be a useful therapeutic agent in the treatment of non-insulin-dependent
diabetes mellitus. Similar insulinotropic effects of NSO were recently observed in
streptozotocin plus nicotinamide-induced diabetes mellitus in hamsters (a model of
type 2 diabetes) orally fed with the oil (Fararh et al., 2002). In this study, positive
immunoreactivity for the presence of insulin was observed in the pancreases from
oil-treated vs. non-treated hamsters using immunohistochemical staining, suggesting
that the hypoglycemic effect of NSO resulted, partly, from a stimulatory effect on
beta cell function with consequent increase in serum insulin level. The ability of NSO
to lower blood glucose concentrations was later conﬁrmed in streptozotocin diabetic
rats following 2, 4 or 6 weeks of treatment (El-Dakhakhny et al., 2002b). In addition,
the effects of NSO, nigellone and TQ were studied on insulin secretion of isolated rat
pancreatic islets. The blood glucose-lowering effect of NSO was not paralleled by a
stimulation of insulin release. The data indicated that the hypoglycemic effect of
NSO might be mediated by extrapancreatic actions, to be elucidated, rather than by
stimulated insulin release (El-Dakhakhny et al., 2002b).
The medicinal potential of black seed (Nigella sativa) 137
In many Arab countries N. sativa and its derived products are consumed abusively
for traditional treatment of blood homeostasis abnormalities and as a treatment for
dyslipidemia (Zaoui et al., 2002a). Several studies support the use of NSO extract for
the treatment of thrombosis and dyslipidemia (Labhal et al., 1997;Enomoto et al.,
2001;Zaoui et al., 2002a). The puriﬁed components (2-(2-methoxypropyl)-5-methyl-
1,4-benzenediol, thymol and carvacrol) obtained from the methanol-soluble portion of
NSO showed inhibitory effects on arachidonic acid-induced platelet aggregation and
blood coagulation. Interestingly, some aromatic compounds present in the extract
were found to be more potent than aspirin, which is well known as a remedy for
thrombosis (Enomoto et al., 2001). In addition, an aqueous suspension of N. sativa
seeds was found to decrease the serum total lipids and body weight in Psammomys
obesus sand rat (Labhal et al., 1997). Analogous results, accompanied by decreases in
serum lipid levels have also been observed in rats chronically treated with N. sativa
ﬁxed oil (Zaoui et al., 2002b). Animals were treated daily with an oral dose of 1 ml/kg
body weight of the N. sativa seed ﬁxed oil for 12 weeks. The serum cholesterol,
triglycerides and the count of leukocytes and platelets decreased significantly by
15.5%, 22%, 35% and 32%, respectively, compared to the control values. Hematocrit
and hemoglobin levels increased significantly by 6.4% and 17.4%, respectively (Zaoui
et al., 2002a), suggesting that the oil inﬂuences blood homeostasis.
N. sativa is also used in Arab folk medicine as a diuretic and hypotensive plant. In
an attempt to experimentally support the above traditional uses of the plant, a study
was conducted on the diuretic and hypotensive effects of the dichloromethane ex-
tract of N. sativa seeds in the spontaneously hypertensive rat (Zaoui et al., 2000). An
oral dose of either N. sativa extract (0.6 mL/kg/day) or furosemide (5 mg/kg/day)
significantly increased diuresis by 16% and 30%, respectively, after 15 days of
treatment. The urinary excretions of Cl
and urea were also increased
after 15 days of treatment. In the same rat study, a comparison between N. sativa
and nifedipine found mean arterial pressure to be decreased by 22% and 18% in the
N. sativa- and nifedipine-treated rats, respectively, suggesting that N. sativa extract
may play a role in decreasing blood pressure.
Evidence indicates that NSO has a protective role against gastric ulcers (El-
Dakhakhny et al., 2000b). Oral administration of NSO for 2 weeks in rats produced a
significant increase in gastric mucin content and glutathione level and a significant
decrease in gastric mucosal histamine content without significant changes in free acidity
and peptic activity of the gastric juice (El-Dakhakhny et al., 2000b). Ethanol admin-
istration, however, produced 100% ulcer induction accompanied by a reduction in free
acidity, mucin content and glutathione level without any significant changes in peptic
activity. When animals were pretreated with NSO before ulcer induction by ethanol, a
protection ratio of 53.56% was noted as compared to the ethanol group (El-Dakhakhny
et al., 2000b).TheprotectiveactionofNSOwasbelievedtobethroughtheincreaseof
the cytoprotective mucin content and/or decrease of histamine.
A ﬁnal physiological effect of NSO includes its potential as an antioxytocic agent.
Aqel and Shaheen (1996) tested the effects of NSO on the uterine smooth muscle of
rats and guinea pigs in vitro using isolated uterine horns. The volatile oil was found
to inhibit the spontaneous movements of rat and guinea pig uterine smooth muscle
and also the contractions induced by oxytocin stimulation (Aqel and Shaheen, 1996).
These effects were concentration dependent and reversible by tissue washing.
Lead molecules from natural products: discovery and new trends138
IV. Antimicrobial and antiparasitic effects of N. sativa oil
Extracts of N. sativa have shown promising effects against bacteria, fungi, viruses,
parasites and worms. In 1975, the puriﬁed compound THQ from NSO was found to
have high antimicrobial effect against Gram positive microorganisms (El-Fatatry,
1975). In later studies, seed extracts of N. sativa were found to inhibit the growth of
Escherichia coli,Bacillus subtilis and Streptococcus feacalis (Saxena and Vyas, 1986).
The antimicrobial activity of N. sativa was further established against several species
of pathogenic bacteria and yeast (Topozada et al., 1965;Hanafy and Hatem, 1991).
In the latter study, ﬁlter paper discs impregnated with the diethyl ether extract of
N. sativa seeds caused concentration-dependent inhibition of Gram-positive
Staphylococcus aureus and Gram-negative Pseudomonas aeruginosa and E. coli and
a pathogenic yeast Candida albicans. The extract showed antibacterial synergism
with streptomycin and gentamicin and showed additive antibacterial action with
spectinomycin, erythromycin, tobramycin, doxycycline, chloramphenicol, nalidixic
acid, ampicillin, lincomycin and sulfamethoxyzole–trimethoprim combination. In-
terestingly, the extract successfully eradicated a non-fatal subcutaneous staphylo-
coccal infection in mice when injected at the site of infection (Hanafy and Hatem,
1991). Recently, crude extracts of N. sativa showed promising antimicrobial effects
against bacterial isolates with multiple resistances against antibiotics (Morsi, 2000).
The most effective extracts were the crude alkaloid and water extracts.
The antiparasitic actions of NSO have been well documented by several researchers
(Agarwal et al., 1979;Akhtar and Riffat, 1991;Abdel-Salam et al., 1993;Mahmoud
et al., 2002). The antihelminthic activities of NSO were studied by Agarwal et al. (1979)
who reported that the essential oil from the seeds of N. sativa showed pronounced
activity even in 1:100 dilutions against tapeworms and earthworms. Anticestodal effects
of N. sativa seeds were studied in children infected naturally with the respective worms.
A single oral administration of 40 mg/kg of N. sativa ethanolic extract reduced the
percentage of the fecal eggs without producing any adverse side effects in the doses
tested (Akhtar and Riffat, 1991). When given orally to Schistosoma mansoni-infected
mice, a 2-week treatment with NSO reduced the number of S. mansoni worms in the
liver and decreased the total number of ova deposited in both the liver and the intestine
(Mahmoud et al., 2002). Furthermore, it increased the number of dead ova in the
intestinal wall and reduced the granuloma diameters markedly (Mahmoud et al., 2002).
When NSO was administered in combination with praziquantel, the drug of choice for
the treatment of schistosomiasis, the most prominent effect was a further lowering of the
dead ova number over that produced by praziquantel alone. These changes were cor-
related mainly with the ability of NSO to improve liver function and the immunological
system of infected mice and partly to its antioxidant effects (Mahmoud et al., 2002). The
protection is also due to the ability of NSO and TQ to reduce the cytogenetic damage
induced by schistosomiasis infection (Aboul-Ela, 2002). Karyotyping of bone marrow
and spleen cells of infected mice showed that the main chromosomal abnormalities were
gaps, fragments and deletions especially in chromosomes 2, 6 and some in chromosomes
13 and 14. Treatment with NSO or TQ for 7 days was found to reduce the percentage of
chromosomal aberrations and the incidence of deletions and tetraploidy compared to
the control level. Thus, NSO may be improving the therapeutic efﬁcacy of S. mansoni
infection by decreasing the induced chromosomal abnormalities.
The medicinal potential of black seed (Nigella sativa) 139
The antiviral effect of NSO was only recently investigated using murine cytome-
galovirus as a model (Salem and Hossain, 2000). The cytomegalovirus is a herpes-
virus that causes disseminated and fatal disease in immunodeﬁcient animals
(Reynolds et al., 1993) similar to that caused by human cytomegalovirus in
immunodeﬁcient humans (Smith and Brennessel, 1994). Intraperitoneal administra-
tion of NSO to mice strikingly inhibited the virus titers in spleen and liver on day 3 of
infection. The difference in the viral load in spleen and liver of the control and NSO-
treated mice was very high, 45 10
vs. 7 10
and 23 10
vs. 3 10
for liver and
spleen, respectively. This antiviral effect coincided with an increase in serum level of
interferon-gamma and increased numbers of CD4
helper T cells and suppressor
function and numbers of macrophages. On day 10 of infection, the virus titer was
undetectable in spleen and liver of NSO-treated mice, while it was detectable in
control mice (Salem and Hossain, 2000). The antiviral effects of NSO were more
potent than the action of Chinese traditional herbal medicine hochuekki – against
murine cytomegalovirus (Hossain et al., 1999).
V. Anticancer effects of N. sativa and its components
The active principles in NSO have been found to exert antineoplastic effects both
in vitro and in vivo using various models of carcinogenesis. In what follows, the
anticancer effects of NSO and its components will be discussed.
V.A. In vitro effects
Black seed preparations (TQ and TQ
) have been demonstrated to have significant
antineoplastic activity against various tumor cells in vitro (Salomi et al., 1991, 1992;
Swamy and Tan, 2000). The active principles of N. sativa showed 50% cytotoxicity
against Ehrlich ascites carcinoma, Dalton’s lymphoma ascites and Sarcoma-180 cells
at a concentration of 1.5, 3 and 1.5 mg, respectively, with little activity against
lymphocytes (Salomi et al., 1991). In vitro cytotoxicity was also demonstrated against
human pancreatic adenocarcinoma, uterine sarcoma and leukemic cell lines (Salomi
et al., 1992). The growth inhibitory activity was found to be related to the extract’s
ability to inhibit DNA synthesis as measured by the incorporation of tritiated thy-
midine into cells. These ﬁndings were later conﬁrmed by Worthen et al. (1998) who
assayed the in vitro cytotoxicity of a crude gum, a ﬁxed oil and two puriﬁed com-
ponents of N. sativa seed, TQ and TQ
, on several parental and multidrug resistant
human tumor cell lines. Although as much as 1% w/v of the gum or oil was devoid of
cytotoxicity, both TQ and TQ
were cytotoxic for all of the tested cell lines (IC
to 393 mM). Interestingly, the multidrug resistant cell variants that are over 10-fold
more resistant to the standard antineoplastic agents doxorubicin and etoposide were
sensitive to TQ and TQ
(Worthen et al., 1998). The ethyl acetate fraction of
N. sativa seeds (identiﬁed as CC-5) was later found to exhibit significant growth
inhibition on a variety of cancer cell lines without inhibiting the growth of normal
human endothelial cells (Swamy and Tan, 2000). The ED
values of the extract
showed increased sensitivity towards Hep G2, LL/2 and Molt4 cell lines compared
with SW620 and J82 cell lines. Badary and Gamal El-Din (2001) also showed that
Lead molecules from natural products: discovery and new trends140
TQ inhibited the survival of ﬁbrosarcoma cells with IC
of 15 mM by inhibiting the
H thymidine into cells. The cellular mechanism of antineoplastic
activity of TQ was only recently investigated (Shoieb et al., 2003). In this study, the
cellular mechanisms of TQ-induced cytotoxicity in parental and cisplatin-resistant
osteosarcoma human breast adenocarcinoma, human ovarian adenocarcinoma and
Madin–Darby canine cell lines have been examined. The cisplatin-resistant cells were
the most sensitive to TQ treatment, while the canine cell lines were the least sensitive.
A dose of 25 mM of TQ induced apoptosis of osteosarcoma cells 6 h after treatment.
This dose also decreased the number of cells in S-phase and increased cells in
-phase, indicating cell cycle arrest at G
. These results suggest that TQ induces cell
cycle arrest and apoptosis in cancer cells. Interestingly, non-cancerous cells are rel-
atively resistant to the apoptotic effects of TQ (Shoieb et al., 2003). In our labo-
ratories, we have recently investigated the effects of TQ on the proliferation and
cytotoxicity of a panel of primary, benign and malignant mouse and human epi-
dermal keratinocytes and colon cells. Although lower doses of TQ were found to
exert no effects on the morphology or proliferation of normal cells, they inhibited
cellular proliferation of benign and malignant cells, conﬁrming the selectivity of this
compound to cancer cells (Gali-Muhtasib et al., 2004). The growth-inhibitory effects
of TQ against colon cancer cells were found to be mainly due to the ability of this
compound to induce G1 cell cycle arrest and apoptosis. The apoptotic effects of TQ
are modulated by Bcl-2 protein and are linked to and dependent on p53 (Gali-
Muhtasib et al., 2004). Our data support the strong potential for using the agent TQ
in the prevention or therapy against colon cancer. We are presently testing the
potency of TQ in the 1,2-dimethyl hydrazine mouse model of colon carcinogenesis
by administering it in drinking water, intraperitoneally or by gavage.
V.B. In vivo effects
Several studies have shown that NSO and TQ retard the carcinogenic process in
animals. The active principles of N. sativa seeds containing fatty acids were found to
completely inhibit the Ehrlich ascites carcinoma in mice (Salomi et al., 1991, 1992).
A dose of 100 mg/kg body weight (b.w.) of N. sativa extract delayed the onset
of papilloma formation and reduced the mean number of papillomas per mouse
(Salomi et al., 1991). Intraperitoneal administration of N. sativa (10 mg/kg b.w.) 30
days after subcutaneous administration of 20-methylcholanthrene-induced soft tis-
sue sarcoma restricted tumor incidence to 33.3% compared to 100% in methyl-
cholanthrene-treated controls (Salomi et al., 1991). In vivo Ehrlich ascites carcinoma
tumor development was completely inhibited by the active principle at the dose of
2 mg per mouse per day for 10 days (Salomi et al., 1992). Furthermore, NSO was
reported to possess a protective effect on chemical-induced carcinogenesis in hamster
cheek pouch (Hassan and El-Dakhakhny, 1992). In another study, the administra-
tion of a dose of 1 mg of TQ twice weekly for 4 weeks demonstrated powerful
chemopreventive effects against benzo(a)pyrene (BP)-induced forestomach tumors
(Badary et al., 1999). TQ inhibited both BP-induced forestomach tumor incidence
and multiplicity by 70% and 67%, respectively. More recently, this same group
(Badary and Gamal El-Din, 2001) demonstrated that the administration of 0.01% of
TQ in drinking water 1 week before and after 20-methylcholanthrene treatment
The medicinal potential of black seed (Nigella sativa) 141
significantly inhibited ﬁbrosarcoma tumor incidence and tumor burden by 43% and
34%, respectively. Moreover, TQ delayed the onset of methylcholanthrene-induced
ﬁbrosarcoma tumors that appeared at 12 weeks and produced less methyl-
cholanthrene-induced mortality. The possible modes of anticarcinogenic actions of
TQ in the above two studies were suggested to be through its antioxidant and
antiinﬂammatory activities, coupled with enhancement of detoxiﬁcation processes.
In a recent study, the effect of CC-5 (ethyl acetate fraction of NSO) was evaluated
for its in vivo antitumor activity against intraperitoneally implanted murine P388
leukemia and subcutaneously implanted Lewis lung carcinoma cells in BDF1 mice
(Kumara and Huat, 2001). At doses of 200 and 400 mg/kg b.w., the fraction pro-
longed the life span of these mice by 153% compared to DMSO-treated control
mice. The antitumor activity of a 21-day treatment of CC-5 against subcutaneously
implanted LL/2 was tested and found to produce a 60–70% tumor inhibition rate. A
triterpene saponin was isolated from the CC-5 fraction and identiﬁed to be a-he-
derin. This compound was found to exert more potent anticancer effects compared
to the commonly used anticancer drug, cyclophosphamide. When a-hederin was
given i.p. at doses of 5 and 10 mg/kg b.w. to mice with formed tumors, it produced
significant dose-dependent tumor inhibition rate values of 50% and 71%, respec-
tively, on day 15, compared to 42% on day 15 in the cyclophosphamide (CP)-treated
group. The underlying mechanism(s) of antitumor activity of a-hederin is not deﬁned
yet (Kumara and Huat, 2001). The protective effect of Nigella grains on carcino-
genesis induced by methylnitrosourea in Sprague Dawley rats was recently inves-
tigated (Mabrouk et al., 2002). When given orally (0.2 g ground Nigella grains) alone
or with honey, a 6-month treatment reduced MNU-induced inﬂammatory reaction
in lung and skin and MNU-induced colon adenocarcinomas by 80% (Mabrouk
et al., 2002). There was an associated elevation of malondialdehyde and nitric oxide
in sera obtained from methylnitrosourea-treated animals, which was lowered by
ingestion of N. sativa grains. Interestingly, combined oral treatment of honey and N.
sativa grains protected 100% against methylnitrosourea-induced oxidative stress,
carcinogenesis and abolished the nitric oxide and malondialdehyde elevations shown
in sera of animals that did not receive these nutrients (Mabrouk et al., 2002).
TQ has also been shown to improve the therapeutic index of several anticancer
agents and to protect non-tumor tissues from chemotherapy-induced damage. TQ
protected against ifosfamide-induced Fanconi syndrome in rats and enhanced its
antitumor activity in Ehrlich ascites carcinoma-bearing mice (Badary, 1999). The
disease Fanconi syndrome is characterized by wasting off glucose, electrolytes and
organic acids along with elevated serum creatinine and urea as well as decreased
creatinine clearance rate (Brade et al., 1986). The changes in renal function observed
in the rat model of Fanconi syndrome correlate well with the nephrotoxic effects of
ifosfamide observed in man. Oral supplementation of TQ (5 mg/kg/day) with drink-
ing water rendered rats significantly less susceptible to ifosfamide-induced renal
abnormalities. It also corrected for the damage induced by ifosfamide on phospho-
rus, glucose, serum creatinine and urea levels and significantly normalized creatinine
clearance rate. This effective dose of TQ was found to be very safe (Badary et al.,
1998). TQ protected the kidney against ifosfamide-induced damage through an
antioxidant mechanism, since it significantly prevented ifosfamide-induced renal
glutathione depletion and lipid peroxide accumulation. In mice bearing Ehrlich
Lead molecules from natural products: discovery and new trends142
ascites carcinoma xenograft, TQ (10 mg/kg/day) administered in drinking water sig-
nificantly enhanced the antitumor effect of ifosfamide. Furthermore, mice treated
with ifosfamide in combination with TQ showed less body weight loss and mortality
rate compared to ifosfamide single therapy. This ﬁnding is in full agreement with
previous ﬁndings that TQ potentiates cisplatin antitumor activity and pro-
tects against cisplatin-induced nephrotoxicity in mice and rats (Badary et al., 1997;
El-Daly, 1998), carbon tetrachloride-induced hepatotoxicity and lipid peroxidation
(Al-Gharably et al., 1997;Nagi et al., 1999) in mice and doxorubicin (DOX)-induced
cardiotoxicity (Al-Shabanah et al., 1998) in mice. In this context, N. sativa seed
extract was shown to protect against cisplatin-induced myelosuppression in mice
(Nair et al., 1991). Moreover, recent investigations by Nagi and Mansour (2000)
showed that oral administration of TQ (10 mg/kg/day) with drinking water starting 5
days before a single i.p. injection of DOX (15 mg/kg) and continuing during the
experimental period ameliorated the DOX-induced cardiotoxicity in rats. TQ also
protected against the nephropathy and oxidative stress induced by DOX in rats
(Badary et al., 2000). Although DOX is a potent cancer chemotherapeutic agent
against several malignancies, its clinical efﬁcacy is limited because of severe cytotoxic
side effects, the most serious being cardiotoxicity (Cortes et al., 1975). Experimen-
tally DOX induces hyperlipidemic nephropathy in rats associated with hypo-
albuminemia, hypoproteinemia, elevated serum urea, hyperlipidemia and a high
urinary excretion of protein and albumin. The nephropathy observed in this model
resembles histologically and clinically the focal and segmental glomerulosclerosis
that occurs in humans (Zima et al., 1997). There is increasing evidence that free
radical generation by DOX is involved in the primary pathogenic mechanism of
DOX-induced nephropathy in rats (Bertani et al., 1986). Treatment of rats with TQ
(10 mg/kg per day) supplemented with the drinking water for 5 days before DOX,
and daily thereafter significantly lowered serum urea and serum and kidney levels of
triglycerides and total cholesterol. It also suppressed DOX-induced proteinuria and
albuminuria (Badary et al., 2000). In both studies, TQ’s protective effects against
DOX damage to the heart and kidney was found to be mainly due to its superoxide-
scavenging and antilipid peroxidation effects.
In conclusion, the ability of TQ to enhance the therapeutic index of anticancer
drugs and provide protection from cytotoxicity induced by these agents strengthens
the potential use of this readily available drug as a cytoprotective agent. This pro-
tection documented by several investigators enforces its preclinical evaluation in
combination with anticancer agents.
VI. Antiinﬂammatory and immunomodulatory effects of N. sativa
N. sativa and its derived products have been traditionally used as a treatment for
rheumatism, liver diseases and related inﬂammatory disorders. The effect of black
seed on the immune system has been investigated by several researchers (Houghton
et al., 1995;El-Dakhakhny et al., 2000a;Haq et al., 1995;Al-Ghamdi, 2001). All
studies have shown that the oil and its most abundant component, TQ, inhibit many
inﬂammatory mediators, and, thus, may be useful in ameliorating inﬂammatory and
autoimmune conditions. Chakravarty (1993) reported that the N. sativa-derived
The medicinal potential of black seed (Nigella sativa) 143
nigellone, the carbonyl polymer of TQ, was very effective at low concentrations in
inhibiting histamine release from rat peritoneal mast cells in vitro. He suggested that
the mechanism of action is mainly due to the ability of TQ to decrease intracellular
calcium by inhibiting protein kinase C and partly due to its ability to inhibit oxi-
dative energy metabolism.
Several studies point to the effect of N. sativa on the human immune system
(El-Kadi and Kandil, 1986;El-Kadi et al., 1987). The seeds were found to produce
an increase in the ratio of helper to suppressor T cells and to enhance natural killer
cell activity in normal volunteers (El-Kadi and Kandil, 1986). In vitro studies showed
that the crude ﬁxed oil and pure TQ were potent inhibitors of eicosanoid generation,
namely thromboxane B
and leucotriene B
, by inhibiting both cyclooxygenase and
lipoxygenase, respectively (Houghton et al., 1995). Thromboxane B
has been im-
plicated in the mechanism of hepatocyte plasma membrane bleb formation, which is
an early event in hepatocyte injury when exposed to oxidative stress (Horton and
Wood, 1990). In another study, N. sativa enhanced the production of IL-3 by human
lymphocytes and had a stimulatory effect on macrophages (Haq et al., 1995). Be-
sides, the immunomodulatory effect of N. sativa puriﬁed proteins was found in
mixed lymphocyte cultures and caused increased secretion in the levels of the
cytokines IL-1band IL-8 (Haq et al., 1999). Moreover, the ﬁxed oil increased the
release of PGE
, inhibited the release of leukotrienes and histamine from normal
and sensitized guinea pig lungs. Other pieces of evidence include the inhibition
of TNF-aproduction in murine septic peritonitis by TQ (El-Dakhakhny et al.,
2000a) and the unique immunomodulatory properties of the ethyl acetate (CC-5)
fraction of N. sativa at non-cytotoxic doses (Swamy and Tan, 2000). The ability of
TQ to modulate cytokines and enhance the immune system has been implicated as
the main reason for its protective effect against schistosome egg infection in the liver
(Mahmoud et al., 2002).
In an attempt to determine the immunomodulatory role of TQ, the effect of this
compound on the production of nitric oxide (NO) by rat peritoneal macrophages
was investigated (El-Mahmoudy et al., 2002). It was found that it reduced produc-
tion of NO in supernatants of lipopolysaccharide-stimulated macrophages without
affecting the cell viability. The protein and mRNA levels of inducible nitric oxides-
ynthase in peritoneal macrophages were also decreased by TQ. Immunoﬂuorescence
staining of inducible nitric oxide synthase in macrophages showed decreased
immunoreactivity for inducible nitric oxide synthase after TQ treatment.
The antiinﬂammatory effect of N. sativa has been found to be comparable to that
of 100 mg/kg aspirin (Al-Ghamdi, 2001). In conclusion, the pharmacological activ-
ities of N. sativa documented by several researchers support its use in folk medicine
to reduce inﬂammation.
VII. Antioxidant and hepatoprotective effects of N. sativa
Health food stores sell N. sativa seeds as a natural remedy for a variety of complaints
including liver diseases (Boulos, 1983). The hepatoprotective effects of TQ have been
well documented and have been found to be related to its strong antioxidant po-
tentials. In fact, the antioxidant and free radical scavenging properties of many
Lead molecules from natural products: discovery and new trends144
plants have been found to play an important role in their hepatoprotective activity
(Kiso et al., 1984;Valenzuela et al., 1986;Navaro et al., 1993;Thabrew et al., 1995).
Oxidant stress can increase the susceptibility to irreversible injury by oxidative in-
toxication and by free radicals that can result in lipid peroxidation, protein oxida-
tion, protein inactivation, disturbance in calcium homeostasis and consequent loss of
cell viability (Masaki et al., 1989;Shertzer et al., 1994). Most of the hepatoprotective
drugs belong to the group of free radical scavengers and their mechanism of action
involves membrane stabilization, neutralization of free radicals and immunomod-
ulation. The ﬂavanolignan mixture, silymarin and its most active constituent, silybin,
obtained from the plant Silybum marianum have been studied intensively for their
antihepatotoxic effects (Vogel, 1977;Miguez et al., 1994). They are now used clin-
ically in the treatment of many liver diseases (Fernandez et al., 1995).
The oil of N. sativa and TQ are known to possess strong antioxidant activities
(Aboul-Enein et al., 1999;Nagi and Mansour, 2000;Meraletal.,2001;El-Dakhakhny
et al., 2002a;Mahmoud et al., 2002); TQ has been shown to inhibit non-enzymatic
peroxidation in ox brain phospholipid liposomes (Houghton et al., 1995) with a potency
that is 10 times higher than NSO. Using TLC screening methods, Burits and Bucar
(2000) showed that TQ and NSO components, namely, carvacrol, t-anethole and
4-terpineol possess strong radical-scavenging properties. Moreover, TQ showed ex-
tremely high superoxide anion radical-scavenging abilities (as effective as superoxide
dismutase against superoxide) in pure chemical systems (Nagi and Mansour, 2000). This
high scavenging power of TQ was responsible for its protective effects against DOX-
induced cardiotoxicity in rats (Nagi and Mansour, 2000). In a recent study, TQ was
observed to be metabolized by liver DT diaphorase to dihydrothymoquinone, a phe-
nolic metabolite that acts as a radical scavenger and inhibits lipid peroxidation in vitro
(Mansour et al., 2002). TQ and dihydrothymoquinone acted not only as superoxide
anion scavengers, but also as general free radical scavengers with IC
values in the
nanomolar and micromolar ranges, respectively (Mansour et al., 2002). Treatment of
mice with TQ orally for 5 successive days produced significant reductions in hepatic
superoxide dismutase, catalase, and glutathione peroxidase activities (Mansour et al.,
2002). Moreover, TQ significantly reduced hepatic and cardiac lipid peroxidation as
compared with the respective control group. The most comprehensive evidence on the
antioxidant effects of NSO and its components came from the studies conducted by
Kruk et al. (2000) and El-Dakhakhny et al. (2002a). They showed that TOH, TQ and
exhibit antioxidant properties and acted as scavengers of various reactive oxygen
species. TOH, for example, acted as
quencher, while TQ and TQ
oxide dismutase-like activity removing O
2.Thesamegroup(El-Dakhakhny et al.,
2002a) also showed that NSO as well as nigellone and TQ exert inhibitory actions on the
production of leukotriene-type mediators of inﬂammation in vitro. Whereas TQ exerts a
strong inhibitory activity (IC
: 0.3 mg/ml), nigellone is far less active (IC
: 12 mg/ml),
possibly due to the loss of antioxidative activity through polymerization. The high
antioxidative action of NSO and its components suggests their importance for the
treatment of various diseases occurring with participation of reactive oxygen species.
In an attempt to evaluate the hepatoprotective effects of TQ, Daba and Abdel-
Rahman (1998) studied its ability to protect against oxidative stress caused by tert-butyl
hydroperoxide in isolated rat hepatocytes and compared it to the effects of the known
hepatoprotective agent silybin. The toxicity of tert-butyl hydroperoxide was manifested
The medicinal potential of black seed (Nigella sativa) 145
by the loss of cell viability and the progressive depletion of intracellular glutathione and
leakage of cytosolic enzymes, alanine transaminase and aspartic transaminase in isolated
rat hepatocytes treated with this compound. Preincubation of cells with 1 mM of either
TQ or silybin resulted in protection against tert-butyl hydroperoxide-induced toxicity as
evidenced by decreased leakage of alanine transaminase and aspartic transaminase and
increased cell viability. Silybin was slightly more potent in preventing loss of cell viability
and enzyme leakage, but both compounds prevented tert-butyl hydroperoxide-induced
depletion of glutathione to the same extent (Daba and Abdel-Rahman, 1998). Hepato-
protective effects of TQ were also documented against carbon tetrachloride-induced
toxicity (Mansour, 2000). In this study, oral administration of TQ in drinking water,
starting 5 days before carbon tetrachloride injection and continuing during the experi-
mental period, ameliorated the hepatotoxicity induced by carbon tetrachloride, as evi-
denced by a significant reduction in the elevated levels of serum enzymes as well as a
significant decrease in the hepatic malonaldehyde content and a significant increase in
the total sulfhydryl content. While oral administration of TQ in a single dose (100 mg/
kg) resulted in significant hepatoprotection against carbon tetrachloride-induced toxicity
in male Swiss albino mice, dihydrothymoquinone (IC
: 0.34), the reduction metabolite
of TQ, was found to be more potent than TQ (IC
: 0.87) in protecting against carbon
tetrachloride-induced hepatotoxicity (Nagi et al., 1999). Similar hepatoprotective effects
in the same system (carbon tetrachloride-induced hepatotoxicity) were obtained fol-
lowing a 4-weeks’ oral intake of NSO in male albino rats (El-Dakhakhny et al., 2000c).
Recently, it was shown that N. sativa seeds given orally every day for 2 months de-
creased the lipid peroxidation, increased the antioxidant defense system and prevented
the lipid peroxidation-induced liver damage in experimentally induced diabetic rabbits
(Meral et al., 2001), suggesting that the seed may be used in diabetic patients to prevent
VIII. Analgesic and antinociceptive effects of N. sativa
The analgesic and antinociceptive effects of N. sativa were only recently reported
(Abdel-Fattah et al., 2000;Al-Ghamdi, 2001) and the mechanisms by which they
occur are not fully understood. Evidence, however, points to the potential of using
the aqueous extract of N. sativa as an analgesic agent. N. sativa crude aqueous
suspension was found to produce significant increase in the hot plate reaction time in
mice (indicating analgesic effects); however, it had no effect on yeast-induced py-
rexia. The absence of antipyretic effect suggests that the constituents of these seeds
may not inhibit the synthesis of prostaglandins (Al-Ghamdi, 2001). This was in
agreement with the ﬁndings of Abdel-Fattah et al. (2000) who showed that the oral
administration of NSO (50–400 mg/kg) dose-dependently suppressed the nociceptive
responses caused by thermal, mechanical and chemical nociceptive stimuli in mice. In
this study, the systemic administration and the i.c.v. injection of NSO attenuated the
response in not only the early phase, but also the late phase of the chemical test. In
another study, upon using several opioid receptor antagonists, it was demonstrated
that NSO and TQ produce antinociceptive effects through indirect activation of the
supraspinal opioid systems (Abdel-Fattah et al., 2000). It remains unclear if TQ
antinociception in the chemical test is due to its direct interaction with opioid
Lead molecules from natural products: discovery and new trends146
receptors, since no information is available regarding the in vitro opioid receptor
binding of TQ. However, the difference in receptor antagonist sensitivity of TQ
antinociception between the early and late phases raises the possibility that the
antinociceptive action of TQ in the chemical test is mediated by mechanisms other
than direct stimulation of opioid receptors located in the central nervous system.
Other mechanisms that could explain the antinociceptive effects of NSO include its
inhibitory effect on the inﬂammatory mediators. Further experiments are needed to
clarify the mechanisms underlying the antinociceptive action of NSO and TQ.
IX. Are N. sativa seeds or its components safe to consume?
The toxicity properties of TQ and THQ were investigated in male rats whereby the
drugs were dissolved in propylene glycol, injected i.p. into 30 male rats and LD
determined (El-Dakhakhny, 1965). Using this protocol, TQ (LD
: 10 mg/kg b.w.)
was found to be more toxic than THQ (LD
: 25 mg/kg b.w.).
In more recent studies, the oral administration of aqueous extracts of the seeds of
N. sativa for 14 days has been shown to cause no toxicity symptoms in male Sprague-
Dawley rats (Tennekoon et al., 1991). The safety of consuming N. sativa seeds was
also recently reported by Al-Homidan et al. (2002) whereby the seeds did not affect
the growth of 7-day-old Hibro broiler chicks when fed to them at 20 and 100 g/kg of
the diet for 7 weeks.
Although several studies have reported the safety of consuming N. sativa seeds, a
recent comprehensive investigation has shown that the plant is relatively unsafe if
consumed for prolonged periods of time (Zaoui et al., 2002b). LD
by single doses, orally and intraperitoneally administered in mice were 28.8 and 2.06
ml/kg b.w., respectively. Treatment of animals with a daily oral dose of 1 ml/kg b.w.
of NSO for 12 weeks resulted in significant slowdown of the body weight in
N. sativa-treated animals compared to untreated control animals. Changes in key
hepatic enzymes levels and histopathological modiﬁcations (heart, liver, kidneys and
pancreas) were not observed in rats treated with N. sativa after 12 weeks. However,
the serum cholesterol, triglyceride and glucose levels and the count of leukocytes and
platelets decreased significantly, compared to control values, while hematocrit and
hemoglobin levels increased significantly. The decrease in body weight in N. sativa-
treated rats was thought to be related to the decrease in serum lipids and glucose
levels as a consequence of a possible reduction in food intake by NSO administration
(Zaoui et al., 2002b). Interestingly, no evidence of toxicity was noted in 10 times this
dose in mice, suggesting only a seeming margin of safety for the used therapeutic
doses of N. sativa.
In this regard, it is worth mentioning that TQ is both an irritant and a potent
elicitor of allergic contact dermatitis (Steinmann et al., 1997;Zedlitz et al., 2002).
The use of ethnobotanical drugs among Asians as complementary medicine is prev-
alent and is also gaining increasing popularity in the West. More than 25% of
The medicinal potential of black seed (Nigella sativa) 147
currently used drugs are derived directly from plants; while the other 25% are
chemically altered natural products. Evidence indicates that N. sativa seeds have a
potential medicinal value and are relatively safe to consume. Future research should
focus on the mechanisms by which N. sativa seeds exert their medicinal effects. With
the increased understanding of its mechanism of bioactivity, the incorporation of
this medicinal herb as complementary medicine into mainstream medical science can
be achieved in the future.
We thank the Deutsche Forschungsgemeinschaft for supporting the in vitro and in
vivo testing of the role of thymoquinone in colon cancer prevention and therapy
(Germany: DFG 477/6-1 and 477/7-1).
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