Black Seed
Novin Ghaffari
2023-05-05
1 Overview
Black seed, Nigella sativa, is a annual flowering herb
native to Western Asia and Eastern Europe. It has a long history of
medicinal use dating back to the ancient world. Modern scholarship and
clinical evidence has found some evidence to support medicinal effects
and elucidate mechanisms of action.
Black seeds
2 Etymology, Taxonomy, History
One study recovered black seed seeds from a pilgrim flask found
in north-central Turkey dating to the Old Hittite period (1650BC). A
GC-MS analysis of the seeds found the content of the essential oil
similar to modern samples (1).
Black seed and its oil has been recovered from the tomb of
Tutankhamun dating to 1325BC. This, along with references to its
medicinal usage, indicates the importance of black seed in ancient
Egyptian society (2).
In Hebrew the word for black seed is (קֶצַח) [qeṣaḥ], possibly
deriving from a Ugaritic word. Black seed is mentioned in the bible in
Isaiah 28:25. Black seed occurs under several names in Arabic, including
(حبة البركة) [ḥabbat al-baraka] and (الحبة السوداء) [al-ḥabbat
al-sawdā’]. In the hadith corpus, Muhammad is reputed to have
recommended black seed as a catch-all cure for every ailment, “except
death.”
In Hindi black seed is referred to as ‘kalonji’ (कलौंजी). It has
a number of uses in Ayurveda, the traditional Indian system of medicine.
In India, black seed is also popular as a culinary condiment for its
intense, unique flavor. In ancient Greek the seeds were referred to as
‘melanthion’ (μελᾰ́νθῐον) and were recommended as medicines by both
Hippocrates and Discorides for a host of ailments. Later the Roman-Greek
physician Galen wrote about the benefits of black seed. In Persian black
seed is referred to as (سیاه دانه) [siyāhdāne]. It was recommended by
the Iranian polymaths Ibn Sina (Avicenna) in his highly influential
‘Canon of Medicine’ and Biruni in his compendium of medicines (3).
The largest producers of black seed are India, Iran, Pakistan,
Turkey, Syria, and Egypt. The largest importers are Brazil, Canada,
Colombia, European Union, Ecuador, Japan, Malaysia, Mexico, South
Africa, and the USA (3).
3 Taste, Appearance, Flavor, and Aroma
Seeds are angular, trigonus, and somewhat elongated; they are
black on the outside and white inside. The aroma and flavor are complex,
being described as a cross between onion, thyme, black pepper, and
sesame seed flavors (4).
4 Active Constituents
One study reported the composition of black seeds as:
- protein: (\(26.7\%\), (5))
- fat: (\(28.5\%\), (5))
- carbohydrates: (\(24.9\%\), (5))
- fiber: (\(8.4\%\), (5))
- total ash: (\(4.8\%\), (5))
The Nigella sativa seeds contain a fatty oil that is abundant in unsaturated fatty acids:
- linoleic acid: (\(50\%-60\%\), (5))
- oleic acid: (\(20\%\), (5))
- dihomolinoleic acid: (\(10\%\), (5))
- eicodadienoic acid: (\(3\%\), (5))
Saturated fatty acids, primarily palmitic and stearic acid, constitute about 30% of the fatty oil fraction (5).
The following trace metals and minerals have been detected in
Nigella sativa:
- Calcium (Ca) (6)
- Copper (Cu) (5),
- Iron (Fe) (5), (6)
- Phosphorus (P) (5), (6)
- Potassium (K) (6)
- Zinc (Zn) (5), (6)
The following elements are constituents of the fatty and essential
oils (5), (7):
df = data.frame(
Group = c(
rep("fixed Oil",6),
rep("terpenes",11),
rep("alkaloids",15),
rep("coumarins",3),
rep("saponins",3),
rep("flavonoids",4),
rep("phenolics",4)
),
Subgroup = c(
rep("unsaturated fatty acids",4),
rep("saturated fatty acids",2),
rep("aliphatic",11),
rep("isoquinoline alkaloids",4),
rep("pyrazole alkaloids",2),
rep("dolabellane diterpenes",9),
"hydroxy coumarin",
"methoxy coumarin",
"oxy coumarin",
"steroidal",
rep("triterpenes",2),
"flavonoidal pigments",
rep("flavonoidal glycosides",3),
rep("acid phenolics",4)
),
Actives = c(
"dihomolinoleic acid",
"eicodadienoic acid",
"linolenic acid",
"oleic acid",
"palmitic acid",
"stearic acid",
"thymoquinone",
"dithymoquinone (nigellone)",
"thymohydroquinone",
"p-cymene",
"α-pinene",
"carvacrol",
"carvone",
"limonene",
"4-terpineol",
"citronellol",
"anethol",
"nigellimine",
"nigellimine N-Oxide",
"nigellicimine",
"nigellicimine N-Oxide",
"nigellicine",
"nigellidine",
"nigellamine A1",
"nigellamine A2",
"nigellamine A3",
"nigellamine A4",
"nigellamine A5",
"nigellamine B1",
"nigellamine B2",
"nigellamine C",
"nigeglamine",
"6-methoxy-coumarin",
"7-hydroxy-coumarin",
"7-oxy-coumarin",
"α-hederin",
"steryl glucosides",
"acetyl-steryl-glucoside",
"quercetin",
"kaempferol 3-glucosyl galactosyl glucoside",
"quercetin 3-galactosyl glucoside",
"trigillin quercetin-3-glucoside",
"vanillic acid",
"hydroxybenzoic acid",
"syringic acid",
"p-cumaric acids"
)
)
One survey of several studies characterizing the essential oil
of Nigella sativa found that thymoquinone ranged from
0.6%-52.6% and p-cymene ranged from 14.1%-60.5% as a percentage
of the total composition (8).
thymoquinone = c(4.47, .79,11.27,.6,42.4,3,52.6)
pcymene = c(56.67,49.48,36.2,14.8,14.1,60.5,25.8)
writeLines("Correlation of Thymoquinone and p-cymene compositions")
corr = as.numeric(format(cor(thymoquinone,pcymene), digits = 3))
corr## Correlation of Thymoquinone and p-cymene compositions
## [1] -0.553
Another study examined the variation in terpenoid content over
the life cycle of Nigella sativa. Almost none of the
characteristic monoterpenes of the seeds are present in the flowers.
Flower extracts were found to mainly contain 2E-hexanal, 2E-hexenol, and
t-caryophyllene. Terpene accumulation begins after pollination at around
30 days. \(\gamma\)-terpinene is the
first terpene to begin increasing in concentration, almost immediately
after pollination and seed formation. Thymoquinone, thymohydroquinone,
and carvacrol begin increasing at day 50. At day 55 \(\gamma\)-terpinene levels begin dropping
off. Thymoquinone appears to peak at day 65, other constituents continue
to increase through the end of the plant-seed life cycle (75 days).
These trajectories of terpene concentrations are depicted below in
Figure 2 from the paper (9):
Reprinted from “Distribution of Primary and Specialized Metabolites in Nigella sativa Seeds, a Spice with Vast Traditional and Historical Uses”, by Botnick, I. et al., 2012, Molecules, p. 10164, Figure 2
5 Active Effects
5.1 Anti-Asthmatic & Bronchodilator
5.1.1 [Salem et al. 2017]
One study examined the bronchodilator and anti-asthmatic effects
of Nigella sativa seeds in 76 18-65 year olds with asthma.
Participants had to meet inclusion criteria. Of the total, 24 were
assigned to control and given placebo (500mg capsule twice daily), 26 to
crushed black seed 1g/day (500 mg capsule twice daily), 26 to crushed
black seed 2g/day (2 500mg capsules twice daily). The day patients were
assigned to their groups and given their treatments during their first
visit (visit 0, week 0) and subsequently checked on twice (visit 1, week
6 and visit 2, week 12). Clinical assessment, spirometry, measurement of
fractional exhaled nitric oxide (FeNO) and serum ‘total immunoglobulin
E’ (IgE) were measured on all visits. Cytokines were measured on visits
0 and 2.
Asthma symptoms were measured via the Asthma control test (ACT).
Other measures used included forced expiratory volume at 1 second
(FEV1), forced vital capacity (FVC), forced expiratory volume
(FEV25-75%), peak expiratory flow (PEF), fractional exhaled nitric oxide
(FeNO), and immunoglobulin E (IgE) levels. For cytokine analysis, 8 ml
of blood was taken, and levels of the following cytokines (IL -
interleukin and IFN - interferon) were checked: IL-4, IL-10, IL-17,
IFN-\(\gamma\), and eotaxin.
The administration of Nigella sativa had a pronounced
effect (\(p<.01\)) in improving ACT
scores at weeks 6 and 12 as compared to controls. Additionally, treated
groups had a much lower rate of severe exacerbation than the control
group. The administration of Nigella sativa also improved
several measures of respiration. In particular the 2g/day dose was found
to improve FEV1 and FEV25-75%, while both Nigella sativa groups
showed gains in PEF over controls (\(p\) values \(<.05\)). From the cytokines measured,
only IFN-\(\gamma\) was significantly
different from baseline in the treatment groups, showing a statisticall
significant (\(p=.03\)) increase. Low
dose Nigella sativa was associated with statistically
significant reductions in FeNO by week 12, whereas high dose was
associated with statistically significant reductions in IgE by week
12.
For information see the study (10).
5.1.2 [He & Xu 2020]
A couple of Chinese researchers conducted a meta-analysis on
trials of Nigella sativa as an asthma treatment. The study
specifically sought randomized controlled trials comparing Nigella
sativa to placebo. All studies used common outcome measures of the
score on the asthma control test (ACT), forced expiratory volume at 1
second (FEV1), peak expiratory flow (PEF). In addition, all studies
considered looked at two common cytokines (IL-4 and IFN-\(\gamma\)).
The researchers searched PubMed, EMbase, Web of science, EBSCO,
and Cochrane library databases. In the end 4 studies were included and a
random effects model was used for the meta-analysis. The following 4
studies were included:
- Boskabady et al 2007 (\(n=29\))
- Barlianto et al 2017 (\(n=28\))
- Koshak et al 2017 (\(n=80\))
- Salem et al 2017 (\(n=50\), didn’t include 2g/day group)
While all these studies met the inclusion criteria and were placebo-controlled randomized trials, there were notable differences (e.g., different age ranges and different format like crushed seeds or extracts).
Overall the authors found the following effects: *
ACT: significant improvements in ACT score (\(p=.01\)) * FEV1:
significant improvements in FEV1 (\(p=.04\)) * PEF:
differences were trending toward significance (\(p=.17\)) * IFN-\(\gamma\): differences were
trending toward significance (\(p=.16\)) * IL-4: effects
on IL-4 did not show any significance (\(p=.5\))
For more see the analysis (11).
5.2 Anticancer
For Nigella sativa in general and its primary active
thymoquinone in particular anticancer effects have been documented and
studied in the peer-reviewed literature.
5.2.1 [Salomi et al. 1991]
An early study examined the anti-carcinogenic capacity of
Nigella sativa alongside saffron, both reputed for their
anticancer properties. Inbred Swiss albino mice were used for the study
with 10 mice in each treatment group in each experiment, 3 experiments
total (i.e. 30 total per treatment group over all experiments). A
methanolic extract of Nigella sativa and a ethanolic-aqueous
extract of Crocus sativus were created and used. Two separate
experiments were conducted. Black seed and saffron were tested topically
in one and internally in the other. In the topical experiment
dimethylbenz[a]anthracene (DMBA) was used to induce skin carcinogenesis
and papilloma formation. In the internal experiment
20-methylcholanthrene (MCA) was administered subcutaneously to induce
soft tissue sarcoma. The application of DMBA was preceded by the
administration of 100 mg/kg of black seed or saffron extract in
treatment groups. MCA was administered by subcutaneous injections, and
30 days after chemical carcinogenesis treatment groups received 100
mg/kg of either black seed intraperitoneally or saffron by oral gavage.
Controls received saline in the internal experiment and DMSO in the
topical experiment.
For the skin carcinogenesis experiment, papilloma measurements
were taken in weeks 6, 9, and 12 weeks. The following percent of animals
had papillomas:
- week 6:
- control: \(56.6\%\)
- black seed: \(33.3\%\)
- saffron: \(26.6\%\)
- week 9:
- control: \(76.6\%\)
- black seed: \(33.3\%\)
- saffron: \(26.6\%\)
- week 12:
- control: \(90\%\)
- black seed: \(76.6\%\)
- saffron: \(50\%\)
The following are the mean number of papillomas per animal with standard deviations * week 6: + control: \(0.80 ± 0.5\) + black seed: \(0.26 ± 0.13\) + saffron: \(0.53 ± 0.35\) * week 9: + control: \(2.0 ± 0.6\) + black seed: \(0.53 ± 0.26\) + saffron: \(0.57 ± 0.27\) * week 12: + control: \(2.4 ± 0.5\) + black seed: \(1.3 ± 0.6\) (\(p<.02\)) + saffron: \(1.0 ± 0.3\)(\(p<.01\))
For the internal experiments, assessments of tumors were taken
at weeks 8 and 12. By week 12 100% of controls had tumors. However many
treated animals did not, so another measurement for those animals was
taken at week 20. Mean tumor diameter was measured only on weeks 8 and
12. The percentages were:
- week 8:
- control: \(56.6\%\)
- black seed: \(0\%\)
- saffron: \(0\%\)
- week 12:
- control: \(76.6\%\)
- black seed: \(33.3\%\)
- saffron: \(10\%\)
- week 20:
- black seed: \(56.6\%\)
- saffron: \(43\%\)
Mean tumor diameters and standard deviations were: * week 8: + control: \(0.83 ± 0.41\) + black seed: \(-\) + saffron: \(-\) * week 12: + control: \(3.15 ± 0.52\) + black seed: \(1.47 ± 0.48\) + saffron: \(0.93 ± 0.26\)
For details on this study see (12).
5.2.2 [Salomi et al. 1992]
A study directly tested the anticancer properties of Nigella
sativa on several cancer cell lines, both cultured cells and cells
transplanted into mice. Interestingly, the study found Nigella
sativa had some inhibitory effect on lymphocyte-derived cell lines
but was more potent against other cell lines.
For the cytotoxicity studies in mice, Swiss albino mice were intraperitoneally transplanted with Dalton’s lymphoma ascites cells (DLA), Ehrlich ascites carcinoma cells (EAC), or Sarcoma-180 cells (S-180). Mice were treated with varying levels of the black seed extract (.5-50 \(\mu\)g/ml). The following concentrations were 50% cytotoxic to each cell line (i.e. \(50\%\) dead cells observed in assays):
- DLA: 3 \(\mu\)g/ml
- EAC: 1.5 \(\mu\)g/ml
- S-180: 1.5 \(\mu\)g/ml
By contrast doses of .28 mg/ml were needed for \(50\%\) cytotoxicity to lymphocytes. In addition, the metabolism of nucleosides by the DLA lines were monitored. A drastic decrease in thymidine uptake was noted, indicating a possible mechanism of action. \(50\%\) inhibition in uptake was achieved with doses of .1 \(\mu\)g/ml and \(75\%\) inhibition with .2 \(\mu\)g/ml. The following effects were seen on average life span between controls, 50 mg/kg, and 100 mg/kg treatment groups:
- EAC:
- control: \(19.0 f 1.71\)
- 50mg/kg: \(31.18 f 3.78\) (\(p<.001\))
- 100mg/kg: - (no tumors) (\(p<.001\))
- DLA:
- control: \(24.38 f 4.96\)
- 50mg/kg: \(34.3 f 4.5\) (\(p<.001\))
- 100mg/kg: \(45.08 f 5.2\) (\(p<.001\))
For the cultured cell study, KB (human oral carcinoma) cells and K-562 (human myelogenous leukemia). At .5 \(\mu\)g/ml \(60\%\) inhibition of the KB cell line was achieved. By contrast, at doses of 10 \(\mu\)g/ml, the extract only retards K-562 growth in the first 24h and then growth of the cell line resumes.
For more details see the study (13).
5.2.3 [Worthen et al 1998]
One study evaluated Nigella sativa (gum and oil) as
well as thymoquinone and dithymoquinone for anticancer activity via
cytotoxicity against multidrug resistant (MDR) human tumor cell lines.
Both thymoquinone and dithymoquinone were cytotoxic against all cell
lines tested. Both parental cell lines and MDR descendents, which were
10 times more resistant to standard chemotherapy agents doxorubicin
(DOX) and etoposide (ETP), were equally sensitive to thymoquinone and
dithymoquinone. The addition of quinine potentiated the impacts of DOX
and ETP on the MDR cell lines and reversed resistance. It had no effect
on thymoquinone and dithymoquinone. The inclusion of DMSO, a free
radical scavenger, reduced the cytotoxicity of DOX by 39%. It had no
effect on thymoquinone and dithymoquinone. This study suggests that
thymoquinone and dithymoquinone are cytotoxic in a manner that is
independent of free radical generation. For more info see (14).
5.3 Anti-COVID-19
5.3.1 [Rahman 2020]
This literature review study raises the prospect of black seed
oil being a powerful auxiliary treatment alongside zinc.
In humans, most zinc is stored intracellularly, with .1% of zinc
found in the plasma. Excess free zinc ions can be toxic cells, so the
body has a system of zinc transporter proteins that maintain
intracellular homeostasis and balance intracellular and external zinc
levels. Free zinc ions play an important role in activating certain
immune functions and have inhibitory effects on viral replication.
During the pandemic there was much controversy over two proposed
treatments: chloroquine and hydroxychloroquine. These compounds serve as
ionophores, enhancing the cellular entry of ZN2+ ions. The
author identifies several terpene compounds in Nigella sativa
oil that can mimic chloroquine’s ionophore capacity. The most promising
constituent the author identifies is nigellimine.
The study notes that CD4+ and CD8+ lymphocyte levels are a potent
predictor of COVID-19 outcomes. Black seed oil has been noted to boost
levels of these lymphocytes.
For more see (15).
5.3.2 [Koshak et al. 2021]
An open-label study in Saudi Arabia found promising results for
the use of Nigella sativa oil in the treatment of mild
COVID-19. Adults 18 and older admitted to King Abdulaziz University
Hospital in Jeddah with mild COVID-19 were recruited for the study
between May 1 and September 31, 2020. Infections were confirmed via
polymerase tests within a week of onset of symptoms. A total of 87
patients were in the control group and 86 in the treatment arm who
received 500 mg of Nigella sativa oil (MARNYS® Cuminmar) twice
daily.
The main outcome of interest was recovery with a 14 day window, with recovery defined as 3 consecutive days symptom-free. The number of days to recovery was secondary outcome of interest. The following results were found:
- recovery rate:
- \(35\%\) (control group)
- \(63\%\) (treatment group)
- \(p=.001\)
- recovery time:
- \(12.4 ± 2.9\) (days, control group)
- \(10.7 ± 3.3\) (days, treatment group)
- \(p=.001\)
For more see (16).
5.3.3 [Xu et al. 2021]
A Chinese research team combined computational and in-vitro
experimental methods for a study on thymoquinone ability to prevent
SARS-CoV-2 (as well as SARS-CoV and NL63) entry into cells via
ACE-2.
Computational modeling studies showed that thymoquinone had a
strong binding affinity for the ACE-2 receptor, effectively blocking the
S1 subunit from binding to the receptor.
To prevent the spread of a real pathogen from the lab and to
facilitate the study, pseudoviruses were created by loading SARS-CoV-2,
SARS-CoV, and NL63 spike proteins into virus particles containing
luciferase reporter genes. This way pseudovirus replication in cell
cultures could be measured via fluorescence imaging. Thymoquinone had an
IC50 of 5 \(\mu\)M for SARS-CoV-2,
7.598 \(\mu\)M for SARS-CoV, and 6.019
for NL63 pseudovirus.
For more see (17).
5.3.4 [Baig & Srinivasan 2022]
A molecular docking study funded by the Indian Ministry of
Science and Technology and performed at the B.S. Abdur Rahman Institute
of Science & Technology, Chennai examined the binding potential of
various Nigella sativa oil constituents to various proteins of
the COVID-19 virus. Chloroquine and its docking potentials were used as
base cases.
The following COVID-19 proteins were tested for docking. The
black cumin constituent with the highest docking potential is listed in
parentheses:
- main protease (Alpha—hederin −8.8 (Kcal/Mol))
- papain-like protease (Nigellamine A2 -10.5 (Kcal/Mol))
- its helicase (Alpha—hederin −10.5 (Kcal/Mol))
- RNA-dependent RNApolymerase (Rutin −9.6 (Kcal/Mol))
- RNA-binding protein (Alpha—hederin −8 (Kcal/Mol))
- Endoribonuclease (Alpha—hederin −10.1 (Kcal/Mol))
- receptor-binding domain (Alpha—hederin −7.8 (Kcal/Mol))
- RNA-binding domain of nucleocapsid phosphoprotein (Alpha—hederin −9 (Kcal/Mol))
A handful of constituents for each protein showed high docking
potential (low binding energy). The authors note an attractive aspect of
this is black seed oil is well-tolerated whereas excessive zinc
consumption can be harmful; hence by acting as an ionophore, black seed
oil in conjunction with zinc could increase its efficacy without
increasing its dose. For more see (18).
5.3.5 [Bencheqroun et al. 2022]
A double-blind, placebo-controlled study using a Nigella
sativa oil formulation ThymoQuinone Formula (TF), which is an
enteric-coated capsule standardized to 1.7% thymoquinone, examined its
capacity to treat COVID-19. The study had a multi-center design, with
trials taking place at the following institutions:
- RESPIRE Research, LLC, Palm Springs, CA, USA
- United Memorial Medical Center, Houston, TX, USA
- L&A Morales Clinic, Miami, FL, USA
Subjects were recruited based on several criteria, including
being 18 years old or older and having mild to moderate symptoms
(indicated by a FLU-PRO Plus assessment score of 3 or greater on at
least 2 symptom scores). Sample sizes were small with only a total of 42
subjects completing all the study protocols with valid FLU-PRO scores.
Primary outcomes safety/tolerability and time-to-sustained clinical
response (SCR). SCR was defined as exhibiting all symptom scores less
than or equal to 2 for 3 days. Several other outcomes of interest were
also examined.
The following were observed:
- adverse events no significant different in adverse events indicating acceptable safety and tolerance
- SCR a difference of 2-2.5 days in SCR that was not statistically significant but was suggestive of being so under a larger sample size
- viral load viral load was not statistically significant at any time point measured, but there was a suggestive difference at day 7 and especially at day 14, indicating a larger sample size may be needed
- T cells there was a statistically significant increase in cytotoxic CD8+ (\(p=.042\)) and CD4+ (\(p=.042\))
- symptom burden there was a statistically significant reduction in total symptom burden (\(p<.001\)) as well as of several individual symptoms
Additional exploratory in-vitro research found that TQF and
thymoquinone had inhibitory effects against several strains of
SARS-CoV-2 (‘omicron’,‘delta’,‘uk’,‘brazil’,‘614G’)
For more see (19).
5.3.6 [Sherwani et al. 2022]
One study examined several health-related impacts and anti-COVID of Nigella sativa oil through in-vitro and in-silico methods. For the study, black cumin seeds were crushed and a methanol extraction was prepared. The following aims were examined:
- free radical scavenging: two standard free radical
scavenging assays were performed to determine antioxidant capacity of
the extract
- DPPH
- \(H_2O_2\)
- red blood cell membrane stability: RBC membrane
stability is adversely affected by COVID-19 and lead to lingering RBC
membrane dysfunction; COVID patients’ RBCs have shown altered lipid
metabolism and increased vulnerability to oxidative damage. Mitigating
abilities of the extract were examined via the following
- heat-induced hemolysis inhibition
- hyposaline-induced hemolysis inhibition
- advanced glycation end products: diabetic patients
are at increased risks with COVID-19; hyperglycemia results in increased
glycation of biomolecules forming AGEs, and AGE formation is associated
with increased inflammatory response and adverse outcomes
- browning intensity
- protein aggregation index
- AGEs-specific fluorescence tests
- molecular docking studies: in-silico molecular docking studies were performed to assess the anti-viral capacity of several phytoconstituents present in the extract
The following results were observed:
- \(H_2O_2\): the \(H_2O_2\) assay found that at concentrations of 400-600 \(\mu\)g/ml the extract had comparable scavenging potential to 100-200 \(\mu\)g/ml of ascorbic acid (\(p<.001\) vs control); at 600 \(\mu\)g/ml the extract had \(70.28\%\) scavenging capacity
- DPPH: the DPPH assay found that at concentrations of 400-600 \(\mu\)g/ml the extract had comparable scavenging potential to 200 \(\mu\)g/ml of ascorbic acid (\(p<.001\) vs control); at 600 \(\mu\)g/ml the extract had \(67.33\%\) scavenging capacity.
- heat-induced hemolysis protection: the study found at concentrations of 400-600 \(\mu\)g/ml the extract had comparable RBC membrane protecting effects on heat-induced hemolysis to 100-200 \(\mu\)g/ml of aspirin (\(p<.001\) vs control); the mean inhibition of heat-induced hemolysis at 600 \(\mu\)g/ml was \(57.86\%\)
- hyposaline-induced hemolysis protection: the study found at concentrations of 400-600 \(\mu\)g/ml the extract had comparable RBC membrane protecting effects on hyposaline-induced hemolysis to 100-200 \(\mu\)g/ml of diclofenac sodium (\(p<.001\) vs control); the mean inhibition of heat-induced hemolysis at 600 \(\mu\)g/ml was \(38.72\%\)
- browning intensity mitigation: the methanol extract significantly reduced browning intensity of glycated BSA at values of 200 \(\mu\)g/ml and over; this was more significant for 400-600 \(\mu\)g/ml (\(p\leq.01\)), and at 600 \(\mu\)g/ml it reduced browning intesity by \(43.91\%\)
- protein aggregation index: at concentrations of 100 \(\mu\)g/ml and over, the extract significantly inhibited protein aggregation; the effect was more significant at 300-600 \(\mu\)g/ml (\(p\leq.01\)), and maximum inhibition was \(51.38\%\)
- AGE fluorescence intensity: at concentrations of 100 \(\mu\)g/ml and over the methanol extract exhibited significant reduction of the AGEs fluorescence intensity; at 400-600 \(\mu\)g/ml the effect was more significant (\(p\leq.01\)), and maximum inhibition was \(52.18\%\)
- molecular docking: the following constituents of
the extract were tested and found to have varying levels of binding
affinity to spike protein components, indicating anti-COVID-19 capacity:
- thymoquinone
- dithymoquinone
- nigellicine
- pentadecanoic acid
- octadecadienoic acid
- (Z)6-pentadecen-1-ol
- 9,12-octadecadien-1-ol
- dioctyl phthalate
For more details on the study see (20).
5.4 Antifungal
5.4.1 [Bita et al. 2012]
Researchers in Romania collected 20 different pathological
strains of Candida albicans from patients at the Craiova
Emergency Hospital. The strains were cultured for in-vitro
experimentation of the anti-fungal capacity of Nigella sativa
seeds. Aqueous, chloroform, and methanolic extracts were taken from
Nigella sativa seeds. Methanolic extracts were found to have
the strongest inhibitory effects against Candida albicans
followed by chloroform. Aqueous extractions showed little to no
antifungal activity (21).
5.4.2 [Rogozhin et al. 2011]
Antimicrobial peptides (AMP) are key element of the immune
systems of plants and animals. Defensins are an important family of
AMPs. Russian and Uzbekistani researchers collected Uzbekistani black
seed specimens. The team used complex chromatographic techniques to
isolate two defensins from Nigella sativa seeds: Ns-D1 and
Ns-D2.
The antifungal activity of both defensins was tested against several fungal species to determine \(\mathrm{IC}50\) concentrations. The following results were obtained:
- Aspergillus niger:
- Ns-D1: 3.5 \(\mu\)g/ml
- Ns-D2: 3.5 \(\mu\)g/ml
- Bipolaris sorokiniana:
- Ns-D1: 3.0 \(\mu\)g/ml
- Ns-D2: 1.8 \(\mu\)g/ml
- Botrytis cinerea:
- Ns-D1: 27.4 \(\mu\)g/ml
- Ns-D2: 13.7 \(\mu\)g/ml
- Fusarium culmorum:
- Ns-D1: 6.9 \(\mu\)g/ml
- Ns-D2: 6.9 \(\mu\)g/ml
- Fusarium graminearum:
- Ns-D1: 6.9 \(\mu\)g/ml
- Ns-D2: 6.9 \(\mu\)g/ml
- Fusarium oxysporum
- Ns-D1: 9.5 \(\mu\)g/ml
- Ns-D2: 5.3 \(\mu\)g/ml
The team also tested the antibacterial activity of these Nigella
sativa defensins. For more see (22).
5.5 Immunomodulatory, Anti-Inflammatory, and Antioxidant
5.5.1 [Chakravarty 1993]
An early study examined the impact of nigellone on histamine
release in-vitro. Nigellone is a carbonyl polymer of thymoquinone; it
retains many of the same pharmacological properties but is lower in
cytotoxicity than thymoquinone. The trial on peritoneal rat mast cells
demonstrated reductions in histamine release with nigellone addition.
The primary mechanisms of action appeared to decreased intracellular
calcium via inhibition of uptake and protein kinase C inhibition.
Additionally there was some evidence of mild inhibition of oxygen energy
metabolism. For more details see (23).
5.5.2 [Umar et al. 2012]
A dysregulation and imbalance between pro-inflammatory and
anti-inflammatory cytokines is a major hallmark of rheumatoid arthritis.
This rat study sought to determine the impact of thymoquinone
administration on cytokine balance in a collagen-induced model of
arthritis. A total of 18 Wistar rats were divided into 3 groups.
Controls were not immunized and administered saline. The arthritis group
was administered a collagen immunization to induce arthritis. The
thymoquinone group was administered thymoquinone after immunization, at
a dose of 5mg/kg for 21 days after.
After the experiment rats were sacrificed and cartilage was
extracted and tested. Levels of several important antioxidants (e.g.,
GSH, SOD, CAT) and several cytokines (interleukins (IL), interferon
(IFN), tumor necrosis factor (TNF)). The following effects were
observed:
- Arthritic Severity: while thymoquinone did not prevent the onset of arthritic symptoms, it did reduce the intensity of onset and mititgated some effects
- TBARS: a measure of lipid peroxidation in cartilege, increased in the arthritis group but thymoquinone greatly inhibited this effect (\(p<.05\))
- Glutathione: glutathione was significantly reduced in the arthritis group, but this was reversed to a significant degree with thymoquinone administration (\(p<.05\))
- SOD: SOD levels were drastically reduced in the arthritic group, but this was markedly improved with thymoquinone administration (\(p<.01\))
- Catalase: catalase activity was greatly reduced in the arthritic group, but this was partly reversed to a significant degree with thymoquinone (\(p<.01\))
- Nitrite: nitrite levels were more than doubled in the arthritic group, this was reduced to a highly significant degree with thymoquinone (\(p<.001\))
- IL-1\(\beta\): pro-inflammatory, levels increased significantly in the arthritic group (\(p<.05\)), which was significantly counteracted with thymoquinone administration (\(p<.05\))
- IL-6: pro-inflammatory, levels increased significantly in the arthritic group (\(p<.05\)), which was significantly counteracted with thymoquinone administration (\(p<.01\))
- IL-10: anti-inflammatory, levels decreased significantly in the arthritic group (\(p<.05\)), which was significantly counteracted with thymoquinone administration (\(p<.05\))
- IFN-\(\gamma\): pro-inflammatory, levels increased significantly in the arthritic group (\(p<.05\)), which was significantly counteracted with thymoquinone administration (\(p<.01\))
- TNF-\(\alpha\): pro-inflammatory, levels increased significantly in the arthritic group (\(p<.05\)), which was significantly counteracted with thymoquinone administration (\(p<.05\))
- PGE2: pro-inflammatory, levels increased significantly in the arthritic group (\(p<.05\)), which was significantly counteracted with thymoquinone administration (\(p<.05\))
For more details see (24).
5.5.3 [Umar et al. 2015]
Tumor necrosis factor-\(\alpha\) (TNF-\(\alpha\)) is a major pro-inflammatory
cytokine produced by monocytes and macrophages that is closely linked to
the etiology of rheumatoid arthritis. It induces the production of other
pro-inflammatory cytokines, such as interleukin (IL) 6 and IL-8, which
in turn induce chronic inflammation and tissue destruction. This process
activates fibroblasts in the joints that produce chemokines, matrix
metalloproteinases, and adhesion molecules, furthering inflammation and
cartilege destruction.
This study examined the effects of thymoquinone, a key active in
Nigella sativa, on TNF-\(\alpha\) induced pro-inflammatory cytokine
response. Fibroblast cell cultures were obtained from rheumatoid
arthritis patients who had undergone joint replacement surgery.
Fibroblasts were serum starved for 2 hours, treated with thymoquinone (1
or 5 μM) for 2 hours, and then treated with TNF-\(\alpha\).
The following outcomes were observed:
- IL-6: 10-fold increase when treated with TNF-\(\alpha\), this effect was reversed in a dose dependent manner by thymoquinone administration
- IL-8: 44-fold increase when treated with TNF-\(\alpha\), this effect was reversed in a dose dependent manner by thymoquinone administration
- ICAM-1: 32-fold increase when treated with TNF-\(\alpha\), this effect was reversed in a dose dependent manner by thymoquinone administration
- VCAM-1: 10-fold increase when treated with TNF-\(\alpha\), this effect was reversed in a dose dependent manner by thymoquinone administration
- Cad11: 2-fold increase when treated with TNF-\(\alpha\), this effect was reversed in a dose dependent manner by thymoquinone administration
In addition, thymoquinone was found to inhibit some, but not all, of the downstream (from TNF-\(\alpha\)) signaling pathways, including p38, JNK, and ASK1.
For more details see the study (25).
5.5.4 [Fallahi et al. 2016]
The main cause of death in respiratory illness is immune system
dysregulation, resulting in overexpression of pro-inflammatory cytokines
leading to inflammation and respiratory failure from poor oxygenation.
In particular, the cytokine interleukin (IL) 13 has been linked to
inflammation of the lungs and airways. MicroRNA (miRNA) 126 and its
overexpression have been directly implicated in IL-13 overactivity.
A rat study induced a cytokine storm in rats via ovalbumin
sensitization. The sensitization produced inflammatory outcomes that
were reversed with the administration of thymoquinone and \(\alpha\)-hederin. The study divided 40 rats
into 4 groups:
- Control: control animals were injected with saline solution
- Sensitized: sensitized animals were injected with ovalbumin
- Thymoquinone: thymoquinone animals were administered 3 mg/kg thymoquinone alongside ovalbumin
- \(\alpha\)-hederin: \(\alpha\)-hederin animals were administered .02 mg/kg \(\alpha\)-hederin alongside ovalbumin
Statistically significant increases in miRNA-126 and IL-13 were
observed in sensitized rats. These increases were reversed to
statistically significant degree by the administration of thymoquinone
and of \(\alpha\)-hederin.
For more details see (26).
5.5.5 [Bordoni et al 2019]
There are a host of volatile bioactive compounds in Nigella
sativa that vary by cultivar, growing conditions/season, extraction
method, and storage. Hence this study compared the anti-inflammatory and
antioxidant effects of fresh extracted oil (FEO) to stored extracted oil
(SEO). FEO was found to have significantly (33%) higher thymoquinone
content.
Three assays were used to determine antioxidant capacity of the
FEO and SEO:
- total antioxidant assay (TAA)
- 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) assay
- chemiluminescence assays
- luminol-amplified: CAT equivalent antioxidant capacity
- lucigenin-amplified: SOD equivalent antioxidant capacity
The overall antioxidant capacity of SEO was greater than that of FEO; the study found the following results: * (TAA): 11.273 ± 0.935 (SEO) and 6.103 ± 0.446 (FEO) (\(p=1.255\times{}10^{-7}\)) * (DPPH) assay: 9.895 ± 0.817 (SEO) and 4.727 ± 0.324 (FEO) (\(p=2.891\times{}10^{-14}\)) * chemiluminescence assays + luminol-amplified: 140.68 ± 4.66 (SEO) and 132.78 ± 11.33 (FEO) (\(p>.05\)) + lucigenin-amplified: 0.0138 ± .0000154 (SEO) and 0.0137 ± .0004175 (FEO) (\(p>.05\))
The study also examined the ability of Nigella sativa to
modulate inflammatory cytokine production. Cytokines release was tested
in-vitro using Simpson–Golabi–Behmel syndrome human pre-adipocytes to
construct a low-grade inflammation model. An array of important
pro-inflammatory cytokines, particularly interleukins (IL), were
surveyed. FEO had a significant impact on lowering IL-6 while SEO had a
significant impact on lowering IL-1\(\beta\). Interestingly, while FEO lowered
IL-6, concentrated thymoquinone did not have the same effect, indicating
that high thymoquinone in combination with the other matrix of elements
in Nigella sativa oil were responsible for these effects.
For more see (27).
5.6 Weight Loss, hyperglycemia, dyslipidemia, and metabolic syndrome
5.6.1 [Farzaneh et al. 2014]
A randomized, double-blind, placebo-controlled trial examined
the effects of Nigella sativa supplementation in combination
with aerobic exercise on lipid profile and on VO2 max in
sedentary, overweight-to-obese women. A total of 20 women were recruited
for the study in Kerman, Iran, with a mean age of \(34.31 ± 7.9\) years and a BMI \(\geq{}25\) kg/m2. Half (\(n=10\)) were assigned to placebo and half
to the treatment group. Only women who had led a sedentary lifestyle for
2 or more years and a total cholesterol level \(\geq{}200\) mg/dl were included. There were
many exclusion criteria (e.g. history of smoking, cardiovascular
disease, diabetes, etc.).
Both groups were instructed in aerobic exercise that involved 10 mins slow running, 10 mins stretching, 30 mins aerobic training, and 10 mins cool down. This training regiment was completed for 3 days week for 8 weeks, and participants were instructed to avoid any other form of strenuous exercise. Subjects were instructed not to deviate from their standard diet. In addition to exercise protocols, subjects were instructed to consume either 2g of Nigella sativa powder or placebo each day for the 8 week duration. Subjects consumed 1g before breakfast and 1g in the afternoon prior to eating.
Blood samples were taken before and after the 8 week training
and supplementation period. The following \(p\) values were obtained for pre and post
trial period
- Triglycerides:
- placebo pre-post difference: (\(p<.001\))
- black seed pre-post difference: (\(p<.001\))
- placebo-black seed difference: (\(p=.42\))
- Total Cholesterol:
- placebo pre-post difference: (\(p<.001\))
- black seed pre-post difference: (\(p=.002\))
- placebo-black seed difference: (\(p=.43\))
- LDL:
- placebo pre-post difference: (\(p=.001\))
- black seed pre-post difference: (\(p<.001\))
- placebo-black seed difference: (\(p=.01\))
- HDL:
- placebo pre-post difference: (\(p=.305\))
- black seed pre-post difference: (\(p=.004\))
- placebo-black seed difference: (\(p=.01\))
- VO_2_ Max:
- placebo pre-post difference: (\(p=.002\))
- black seed pre-post difference: (\(p=.008\))
- placebo-black seed difference: (\(p=.35\))
- BMI:
- placebo pre-post difference: (\(p=.099\))
- black seed pre-post difference: (\(p=.001\))
- placebo-black seed difference: (\(p=.24\))
In particular, this suggests Nigella sativa
particularly helps in lowering cholesterol. For more details see (28).
5.6.2 [Amin et al. 2015]
A study conducted a randomized controlled trial examining the
efficacy of black seed and turmeric, alone and in combination, for
controlling several cardiovascular health metrics in patients with
metabolic syndrome. The study took place in Pakistan and recruited 250
males with a waist circumference \(>90\) cm and at least 3 other markers of
metabolic syndrome. Patients were assigned either to control (500 mg of
ispaghula husk, 1.5 g/day), black seed (500 mg, 1.5 g/day), turmeric
(800mg, 2.4 g/day), or black seed + turmeric (300 mg + 500 mg, 900
mg/day + 1.5 g/day) groups. Blood samples were taken at baseline, 4
weeks, and 8 weeks; several markers of metabolic health were tested:
- fasting blood glucose
- c-reactive protein
- total cholesterol
- LDL
- HDL
- triglycerides
In addition the following measurements were also taken:
- blood pressure
- BMI
- body fat %
- waist circumference
- hip circumferences
At weeks 4 and 8, as compared to baseline, each treatment had the statistically significant improvements on the following:
- placebo:
- week 4: BMI
- week 8: BMI, bodyfat %
- black seed:
- week 4: BMI, bodyfat %, waist circumference
- week 8: BMI, bodyfat %, waist circumference, total cholesterol, triglycerides
- turmeric:
- week 4: BMI, bodyfat %, waist circumference, hip circumference
- week 8: BMI, bodyfat %, waist circumference, hip circumference
- black seed + turmeric:
- week 4: BMI, bodyfat %, waist circumference, hip circumference, total cholesterol, LDL, triglycerides, fasting blood glucose, diastolic BP, systolic BP
- week 8: BMI, bodyfat %, waist circumference, hip circumference, total cholesterol, LDL, HDL, triglycerides, fasting blood glucose, diastolic BP, systolic BP
The study showed that while black seed and turmeric individually
can have positive impacts on metabolic syndrome, their combination (even
at 60% of the dosage of each), has synergistic impacts on a broader
range of parameters affecting metabolic syndrome. For more details see
(29).
5.6.3 [Mahdavi et al. 2015]
A double-blind, randomized, placebo-controlled study examined
effects of Nigella sativa oil (NSO) alongside a low-calorie
diet on weight loss and markers of cardiovascular health. Obese women
with BMI between 30-35 kg/m2. Women were randomly assigned to
a low calorie diet and 3g/day (1g with each meal) of placebo or NSO for
8 weeks. A total of 43 women participated in the treatment group and 41
in the placebo group. NSO administration was associated with the
following significant changes in weight and cardiometabolic markers:
- weight loss:
- placebo: \(-3.6\%\)
- NSO: \(-6.0\%\)
- significance: (\(p<.01\))
- waist circumference:
- placebo: \(-3.4\%\)
- NSO: \(-6.9\%\)
- significance: (\(p<.01\))
- triglycerides:
- placebo: \(-1.4\%\)
- NSO: \(-14.0\%\)
- significance: (\(p<.01\))
- v-LDL:
- placebo: \(-7.0\%\)
- NSO: \(-14.0\%\)
- significance: (\(p<.01\))
For more, see (30).
5.6.4 [Namazi et al. 2015]
A double-blind, placebo-controlled study looked at the effect of
Nigella sativa oil (NSO) consumed concurrently with a
low-calorie diet intervention on obese women. The study recorded overall
weight loss alongside several markers of oxidative stress after 8 weeks
of low-calorie dieting with either 3g/day of placebo or NSO. A total of
25 women in the treatment group and 24 in the placebo group completed
the study. The study found the following effects:
- weight loss:
- placebo: \(-1.40 ± 1.90\)kg
- NSO: \(-4.80 ± 1.50\)kg
- significance: (\(p<.01\))
- red blood cell SOD:
- placebo: \(-3.30 ± 109.80\)U/gHb
- NSO: \(88.98 ± 87.46\)U/gHb
- significance: (\(p<.01\))
- glutathione peroxidase: no significant difference
- lipid peroxidation: no significant difference
For more see the study (31).
5.6.5 [Ahmad et al. 2017]
In this study 30 albino rats were divided into three groups of
10: A, B, C. Group A were controls. Group B were fed a cholesterol
supplement inducing hyperlipidemia. Group C were fed the cholesterol
supplement in addition to 500mg/kg of black seed powder. Serum levels of
cholesterol, triglycerides, LDL, and HDL were all measured. The
following were mean levels for each Group
- Group A:
- Cholesterol: \(72.30±4.45\)
- Triglycerides: \(108.41±1.86\)
- LDL: \(8.51±0.34\)
- HDL: \(6.36±0.36\)
- Group B:
- Cholesterol: \(189.10±5.16\)
- Triglycerides: \(287.03±4.17\)
- LDL: \(71.11±0.81\)
- HDL: \(3.36±0.24\)
- Group A:
- Cholesterol: \(107.48±11.56\)
- Triglycerides: \(138.98±15.23\)
- LDL: \(35.72±12.30\)
- HDL: \(4.88±0.87\)
Improvements in lipid profile in group C over group B were
statistically significant (\(p<.05\)). For the original study see
(32).
5.6.6 [Safi et al. 2021]
A randomized, placebo-controlled, double-blind crossover trial
examined the benefits Nigella sativa on weight loss and several
anthropometric parameters. Participants were overweight women 25-55
years old with BMI 27-35 kg/m2 who passed the exclusion
criteria. The study consisted of two 8 week treatment periods with a 4
week washout in between. Treatment dosages were 2 g/day of black seed or
placebo (paraffin oil) in the form of 1g capsules taken twice daily.
Women were placed on individualized iso-caloric diets. The outcomes of
interest were several anthropometric and body composition indices linked
to obesity along with appetite.
The following effects were observed:
- body weight: \(p<.001\) and Cohen’s \(d=.6\)
- BMI: \(p<.001\) and Cohen’s \(d=.5\)
- waist circumference: \(p=.02\) and Cohen’s \(d=.4\)
- hip circumference: \(p=.049\) and Cohen’s \(d=.7\)
- body fat percentage: \(p<.001\) and Cohen’s \(d=.2\)
- body fat mass: \(p=.001\) and Cohen’s \(d=.2\)
- visceral fat area: \(p=.011\) and Cohen’s \(d=.2\)
- appetite: improvements all 4 measures of appetite: hunger, satiety, fullness, prospective consumption
For more details see the study (33).