Phytochemical composition,therapeutical and pharmacological potential of Nigella sativa:a review

Nivedita Manoharan,Dheepthi Jayamurali,Rajeshwari Parasuraman,Sathya Narayanan Govindarajulu＊

1Department of Physiology,Dr.ALM Post Graduate Institute of Basic Medical Sciences,University of Madras,Taramani,Chennai,Tamil Nadu 600113,India.

Abstract Plants naturally produce chemical compounds,which are used to endorse health and fight against diseases and have been used traditionally.Nigella sativa(N.sativa)has a broad spectrum of usage in traditional folk medicines and it’s a well-known medicinal herb in ancient Ayurveda,Siddha,Chinese,Arab and Unani Tibb.N.sativa stimulates the body with its natural vitalizing process and cures the disease.Phytochemically,N.sativa seed contains a wide range of fixed oils,proteins,alkaloids,saponin and essential oils.Most of the medicinal properties of N.sativa are due to the presence of the quinone constituent.Among the constituents of N.sativa,thymoquinone is one of the predominant bioactive compounds.The aim of this review is to explore the phytochemical,therapeutical and pharmacological potentials of N.sativa.N.sativa reported to have many therapeutical and pharmacological actions which includes antioxidant,antimicrobial,antidiabetic,anticancer,anti-inflammatory,immunomodulator,cardioprotective,antihyperlipidemic,pulmonary protective,hepatoprotective,nephroprotective,gastroprotective,diuretics,anti-osteoporotic,dermatologic,neuroprotective effects and also has stimulative action on the reproductive system.N.sativa seeds and its derivatives have to be isolated and further investigations should be done using animal models and clinical trials to understand its novel molecular mechanism of action.So,N.sativa and the plant derived constituents can be used in the production of new drug and to treat several diseases.

Keywords:Nigella sativa,Phytochemistry,Therapy,Pharmacology

Background

Plants are known for their medicinal properties for centuries and they have been used globally to cure and avert diseases,infections,and infestation[1].There are numerous plants which have active bioactive compounds with therapeutic characteristic[2].Plant species with a long history of pharmacological and phytochemical properties leads to the safe use of the herb and development of drug[3].In recent years,there has been increasing interest in the usage of natural products because they are cost effective,they have various therapeutic properties and also because of the connotation of side effects to man-made drugs[4].In the course of time,almost 25%of drugs are reliable plants based;another 25% are chemically altered plant based products[5,6].Among several medicinal plants,Nigella sativa(N.sativa)has wide-ranging therapeutic effect and they are also reported for their significant uses[7].N.sativanot only known for their medicinal properties,but they are also widely used as spices and flavoring agents in a variety of foods[8].N.sativahas broad antiquity in different civilization as a miracle herb for promoting the body for its natural healing process[9].The herb was considered to cure any disease except death in prophetic medicine[5]and it is first mentioned inBook of Isaiahin 8thcentury B.C.E.[10].This miraculous herb was considered as“the herb from heaven”by the ancient herb specialists Avicenna(10thcentury)[8].Archeological evidence reported that in the tomb of Tutankhamen the herb was sited[10].N.sativais traditionally used in Ayurveda,Siddha,Chinese,Arab and Unani Tibb[2,11].

Nigellais a genus of about 20 species of annual plants in the family Ranunculaceae(Table 1).The common titles applied to members of this genus are“devil-in-a-bush”or“love in a mist”[12].N.sativagrows in loamy soils mainly in South Europe,North Africa,and South-West Asia and also cultivated in Middle East Mediterranean region[11,13].It is an annual flowering plant which grows up to 20-90 cm tall,finely divided leaves,the leaf segment narrowly linear or threadlike.The flowers are delicate,and usually colored white,yellow,pink,pale blue or pale purple with 5-10 petals.The ripe fruit has large and inflated capsule which is composed of 3-7 united follicles,each containing numerous seeds.The seeds are small dicotyledons,trigonus,angular,regular tubercle,2-3.5 mm×1-2 mm black externally and white inside,slightly aromatic with pungent bitter taste(Figure 1)[7,12]

Table 1 Taxonomy of N.sativa

Biological activities and therapeutical uses ofN.sativawere widely studied in the ancient period.N.sativaseeds and oil are traditionally used to treat several diseases.N.sativawas well-known for its medicinal properties as they are used in rheumatism,jaundice,dyspepsia,ulcer,helminthic infections,piles,diuresis,bronchitis,asthma,epilepsies,paralysis,inflammatory diseases,fatty liver,diabetes,cardiovascular diseases,conjunctivitis,ammenorhea,anorexia,pain such as chronic headaches and back pain.They are also applied directly in blister,nasal abscesses,orchitis,eczema,and swollen joints[2,7,8,14,15].Most of the pharmacological and therapeutical properties ofN.sativaare due to the presence of the quinone constituent,in which thymoquinone(TQ)is a one of the major bioactive compounds of the essential oil[2].The aim of this review is to explore the phytochemical,therapeutical,and pharmacological potentials ofN.sativa.

Phytochemical composition

In different varieties of seeds,various bioactive compounds were identified and stated in many studies.Among them,the most important bioactive compounds are TQ (30%-48%), thymohydroquinone,dithymoquinone,p-cymene(7%-15%),carvacrol(6%-12%),4-terpineol(2%-7%),t-anethol(1%-4%),sesquiterpene longifolene(1%-8%),α-pinene and thymol etc.N.sativaalso contains some other compounds in trace amounts.The seeds have two types of alkaloids; that is isoquinoline alkaloids-nigellicimine and nigellicimine-N-oxide,and pyrazol alkaloids or indazole ring bearing alkaloids which include nigellidine and nigellicine.The seeds also contain α-hederin(water soluble pentacyclic triterpene)and saponin(potential anticancer agent)[18,19].In trace amounts other compounds like carvone,limonene,citronellol were also found.The seeds ofN.sativaalso contain protein(26.7%),fat(28.5%),carbohydrates(24.9%),crude fiber(8.4%)and total ash(4.8%).The seed also contains good amount of various vitamins and minerals like Cu,P,Zn,Fe and carotene which is converted by the liver to vitamin A[18,20].

In some research these seeds are also reported to contain rich unsaturated fatty acids,mainly linoleic acid(50%-60%),oleic acid(20%),eicodadienoic acid(3%)and dihomolinoleic acid(10%).Saturated fatty acids(palmitic,stearic acid)are about 30% or less.α-sitosterol is a major sterol,which is 54% of the total sterols,followed by stigmasterol(6.6%-20.9% of total sterols)[21-23].In few other studies it is reported that the other components includes nigellone,avenasterol-5-ene,avenasterol-7-ene,campesterol,cholesterol,citrostadienol,cycloeucalenol,gramisterol,lophenol,obtusifoliol,stigmastanol,stigmasterol-7-ene,β-amyrin, butyro-spermol, cycloartenol,24-methylenecycloartanol,taraxerol,tirucallol,3-O-(β-D-xylopyranosyl(1→3)-α-L-rhamnopyranosyl(1→2)-α-L-arabino-pyranosyl)-28-O-(α-L-rhamnopyranosyl-(1→4)-β-glucopyranosyl(1→6)-β-D-gluco-pyranosyl)hederagenin,volatile oil(0.5%-1.6%),fatty oil(35.6%-41.6%),oleic acid,esters of unsaturated fatty acids with C15 and higher terpenoids,esters of dehydrostearic and linoleic acid,aliphatic alcohol,β-unsaturated hydroxy ketone,hederagenin glycoside,melanthin,melanthigenin,tannin,resin,protein,reducing sugar,glycosidal saponin,3-O-(β-Dxylopyranosyl-(1→2)-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranosyl)-11-methoxy-16,23-dihydroxy-28-methy-lolean-12-enoate,stigma-5,22-dien-3-β-Dgluco-pyranoside, cycloart-23-methyl-7, 20,22-triene-3β,25-diol,nigellidine-4-O-sulfite,N.mines A3,A4,A5,C,N.mines A1,A2,B1,and B2[20,24,25-27].The phytochemical composition of some of the components ofN.sativa(Table 2)(Figure 2)

Table 2 Chemical composition of N.sativa seeds

Figure 2 Chemical structure of components

Pharmacological activity of N.sativa

In last few decades,extensive research was conducted onN.sativaseed extracts and its bioactive compounds to study its therapeutical and pharmacological actions on multiple diseases.The following are the medleys of some research.

Antioxidant activity

Essential oil ofN.sativawas tested for its antioxidant activity by thin-layer chromatography screening methods,in which TQ and the components carvacrol,t-anethole,and 4-terpineol confirmed upright radical scavenging property.They were also effective·OH radical scavenging agents in the assay for non-enzymatic lipid peroxidation in liposomes and the deoxyribose degradation assay.This indicates thatN.sativaseeds can be used in the treatment of various diseases[42].TQ,the natural main constituent ofN.sativaoil(NSO),and a synthetic structurally related tert-butyl hydroquinone(TBHQ)were studied in vitro.Both TQ and TBHQ powerfully inhibited iron-dependent microsomal lipid peroxidation in a concentration-dependent manner with median inhibitory concentration(IC50)values of 16.8 and 14.9 mM,respectively.TQ was more active than TBHQ as a superoxide anion scavenger with IC50 of 3.35 mM compared to 18.1 mM for TBHQ[43].BothN.sativaand essential oil was effective in enhancing the antioxidant indices against potassium bromate induced oxidative stress by significantly increasing the glutathione-S-transferase,glutathione reductase,and glutathione peroxidase(GSH-Px)[17].

Likewise,the methanolic extract and volatile oil fractionated fromN.sativaseed oil in atherogenic suspension fed rats have been reported to have replenished the plasma total antioxidant power by an average of 88% against free radical[44].The modulatory effect of TQ on erythrocyte lipid peroxidation and antioxidant status during 1,2-dimethylhydrazine (DMH)-induced colon carcinogenesis after initiation in male Wistar rats was studied in which pretreatment of TQ refurbished the increased level of malondialdehyde(MDA)and conjugated diene levels,and augmentation of enzyme activities like catalase(CAT),GSH-Px,and superoxide dismutase(SOD)activities due to exposure to DMH.TQ was a useful compound preventing DMH-induced erythrocyte damages[45].

Antimicrobial activity

N.sativaextract(NSE)is reported to have effective antibacterial activity against gram-positive(Staphylococcusaureus) and gram-negative(Pseudomonas aeruginosaandEscherichia coli)species.The extract showed a synergistic effect with streptomycin and gentamycin and also showed addictive action with spectinomycin,erythromycin,tobramycin,doxycycline,chloramphenicol,nalidixic acid, ampicillin, lincomycin, and sulphamethoxyzole-trimethoprim combination.These outcomes suggested the antimicrobial efficacy of the extract[46].The methanolic extract ofN.sativaseed shows a greater inhibition zone for gram-positive bacteria(Streptococcus pyogene)and gram-negative bacteria(Pseudomonas aeruginosa,Klebseilla pneumoniaeandProteus vulgaris)[47].TQ revealed a significant antibacterial activity majorly against gram-positive cocci with MICs values ranging from 8 to 32 μg/mL showed the minimum biofilm inhibition concentration at 22 and 60 μg/mL forStaphylococcus aureusandStaphylococcus epidermidis,respectively[48].

The effect of essential oil and extract ofN.sativamainly TQ proved its potent antifungicidal activity on different dermatophyte strains likeTrichophyton entagrophytes,Microsporum canisandMicrosporum gypseum[49].The aqueous extract ofN.sativaexhibits an inhibitory effect against candidiasis and the study also validates that a 5-fold decrease in candida in kidneys,8-fold in the liver,and 11-fold in the spleen was observed in post-treated mice[50].Two novel defensins Ns-D1 and Ns-D2 were isolated and sequenced fromN.sativaseeds.These defensins show strong antifungal activity towards several phytopathogenic fungi[51].

Using murine cytomegalovirus model,antiviral effect of NSO was studied in which NSO exhibited complete inhibition of virus titer in the liver and spleen on day 3 of infection and coincided with the serum level of rising interferon-γ(IFN-γ),macrophages,and CD4+T cells.On the 10thday of infection,the virus titer was undetectable in the liver and spleen of NSO treated mice.The result exhibited a striking antiviral activity against murine cytomegalovirus mediated by increase in macrophages and IFN-γ production[52].

NSO was given forSchistosoma mansoniinfected mice for two weeks,which showed diminished number of worms in the liver and also reduced in ova deposition in the liver and intestine.The infected mice also produced a rise in serum activity of L-alanine aminotransferase(ALT),gamma-glutamyl transferase,with a low increase in alkaline phosphatase level,while reduced serum albumin level,administration of NSO was partially successful.When NSO along with praziquantel was administered,it showed a reduction in the dead ova number in comparison with praziquantel alone seen.The results suggested that NSO plays a role in the modification caused bySchistosoma mansoni[53].Anti-plasmodial activity of various plant extract was performed among thatN.sativashowed a 100%inhibition of the parasite growth(Plasmodium falciparum)at a concentration≤50 μg/mL[54].Efficacy of NSO againstHymenolepis nana,a causative agent of zoonosis diseases mice was used in this study to check the inhibitory effect of NSO dose-dependent against the causative agent.The efficacy of the 5 mL/kg NSO dose againstHymenolepis nanaattained 100% 14 days after treatment;the efficacy of the 2.5 mL/kg NSO does attain an efficacy of 100% 21 days after treatment.It indicates that NSO unveils efficacy againstHymenolepis nana[55].

Anti-diabetic activity

The effect ofN.sativaand TQ on histopathological changes of the sciatic nerve was evaluated.The treatment withN.sativaand TQ showed a sharp decrease in elevated serum glucose and an increase in lowered serum insulin concentration in streptozotocin(STZ)induced diabetic rats.The treatment withN.sativaand TQ increased the area of insulin immunoreactive beta cells significantly.Only fewer morphological changes were seen in diabetic animals treated withN.sativaand TQ.The ultra-structural features of axon also showed a remarkable improvement suggesting the utility ofN.sativaand TQ as a potential treatment on peripheral neuropathy in STZ induced diabetic rats[56].

The effect of hydrochloric extract of NSE at high dosage did not exhibit any effect.But this extract at low doses has a hypoglycemic effect and ameliorative effect on the regeneration of pancreatic islets and maybe also is used as a therapeutic agent in the management of diabetes mellitus[57].N.sativaand its constituent TQ possess maximum anti-diabetic potential towards hepatic gluconeogenesis,glucose absorption in the intestine,blood glucose level,simulates glucose induced secretion of insulin from beta cells in the pancreas,improves glucose tolerance as efficiently as metformin so far it has not shown significant adverse effects and has very low toxicity[58].

Anti-cancer activity

This study investigates the anticancer activity of NSO and petroleum ether extract ofN.sativaagainst a human lung cancer cell line.NSO(0.10,0.25,0.50,and 1.00 mg/mL)and NSE(0.25,0.50,and 1.00 mg/mL)were added to the cell line and kept for 24h.NSE and NSO significantly reduce the viability of cells and alter the cellular morphology of A-549 cells in a concentration dependent manner[59].In another study,NSE obtained by various seed thermal processing was examined in-vitro for their anti-proliferative activity in mouse colon carcinoma cells and TQ content.At various thermal processing 25°C,50°C,100°C,or 150°C,the seeds were heated at 50°C,100°C,or 150°C produced oil with a strong ability to inhibit tumor cell growth and a higher TQ content and inhibits the NF-κB signaling pathway[60].The research suggesting that TQ induced apoptosis through a p53-independent pathway with the expression of p21 and cell cycle arrest at S phase in human colon cancer cells[61].TQ treated to DLD-1(human colon cancer cell line)increased the phosphorylation of mitogen-activated protein kinases extracellular signal-regulated kinase and c-Jun N-terminal kinase,but not of p38.It provides the evidence linking the pro-oxidant effects of TQ with its apoptotic effects in colon cancer and shows a protective role of mitogen-activated protein kinases[62].

Anti-inflammatory property

NSO,nigellone,and TQ were evaluated for their effect on the formation of 5-lipoxygenase(5-LO)product from polymorphonuclear leukocytes.IC50 values of inhibition of 5-LO for NSO and TQ were similar at 25 μg/mL and 0.2 μg/mL.The IC50 value of 5-hydroxy-eicosa-tetra-enoic acid production from NSO,nigellone,and TQ are 24 μg/mL,11.9 μg/mL,and 0.36 μg/mL respectively.It shows part of the effect of oil,nigellone,and TQ in ameliorating inflammatory disease[63].The NSO and TQ both inhibit the eicosanoid generation in leukocytes through the inhibition of cyclo-oxygenase and 5-LO pathways of arachidonate metabolism.This shows that NSO and TQ can be used in the treatment of rheumatism and related inflammatory diseases[64].In a comparative study of ethanolic extract ofN.sativaand diclofenac sodium(an anti-inflammatory drug),the extract significantly reduces the paw inflammatory response in albino rats.The ethanolic extract ofN.sativahas a potent anti-inflammatory effect in albino rats however,this effect is moderately less but prolonged than that produced by diclofenac sodium[65].Experimental autoimmune encephalomyelitis is a well-established animal model for multiple sclerosis.NSO treated animals ameliorated the clinical signs and suppressed inflammation and also enhanced remyelination in the hippocampus.N.sativacould be used as a protective agent or an aide in treatment for encephalomyelitis even when the treatment started after the appearance of the first clinical signs[66].

Immunomodulatory activity

A report showed that alloxan induced diabetic rats showed a significant increase in monocytes and granulocytes and a significant decrease in the proliferation capacity of lymphocytes and tumour necrosis factor α(TNF-α),interleukin(IL)-4,and IL-8 levels in the diabetic group.When these rats were treated withN.sativainduced significant amelioration in monocytes and granulocytes,with a significant increase in lymphocyte TNF-α,IL-4,and IL-8 levels.It showed the potential effect ofN.sativaas an immunomodulator[67].At different concentrations,ethanolic extract ofN.sativa,TQ and dexamethasone has effect on cell viability,proliferation,IL-4,and IFN-γ secretion in nonstimulated,phytohemagglutinin and (concavaline A)-stimulated splenocytes.Dexamethasone treated animals indicated suppression in cell viability,IFN-γ,and IL-4 secretion in non-stimulated and stimulated splenocytes while extract and TQ reduced the viability and inhibited the proliferation of stimulated and non-stimulated splenocytes concentration-dependently.The higher concentration of extract and TQ reduced the secretion of IL-4/IFN-γ ratio in both stimulated and non-stimulated cells,while the higher concentration of TQ produced same effect on stimulated cells.N.sativaand TQ showed a cytotoxic inhibitory effect on rat splenocytes,T-helper 1 and T-helper 2 cytokines concentration-dependently.It shows the cytotoxic and immunomodulatory effect ofN.sativaand TQ[68].Likewise,dexamethasone induced immune suppressed male rabbits showed a decreased in phagocytic activity while NSE reported to increase the phagocytic activity by stimulating the immune cells and increase the immune potentiality[69].The radioprotective effect ofNigellacrude oil against the hemopoietic adverse effect of gamma irradiation was evaluated.NSO administered before irradiation significantly normalized the increased MDA concentration and plasma GSH-Px,CAT,and erythrocyte SOD activities and produced significant regeneration in spleen and thymus lymphoid follicles,thus having a promising natural radioprotective activity against the immunosuppressive and oxidative effect of ionizing radiation[70].

Cardioprotective activity

The effect ofN.sativasupplementation for two months to normal rats on cardiac hemodynamics in vivo,the ionotropic and chronotropic properties of the isolated hearts in vitro,and the cardiac responsiveness to progressive adrenergic stimulation by isoproterenol were studied in which the results indicated the intrinsic cardiac contractile properties without evidence of increased cardiac workload or energy consumption in vivo makesN.sativaan inotriopic agent[71].This shows the cardioprotective effect of NSO on lead induced cardiotoxicity.The results showed produced significant normalization of the physiological parameters and restored the histological structure and decreased the cyclooxygenase-2 expression of the heart.NSO intake has cardioprotective potential through its ability to decrease pro-inflammatory cytokines,oxidative stress,and cardiac tissue damage[72].Myocardial injury was induced in rats by cisplatin(CP)and showed a significant increase in congestion,edema,and pyknotic nuclei in myocardial fibres and decrease the expression of antiapoptotic protein B cell lymphoma-2.CP along with TQ was administered and it showed attenuating cardiomyocyte necrosis and apoptosis by increasing B cell lymphoma-2 expression[73].

Anti-hyperlipidemic effects

N.sativawas given to hyperlipidemia rats and there was significantly reduced cholesterol and triglycerides with adequate safety profile.This shows the effectiveness ofN.sativato treat hyperlipidemia[74].A similar study was conducted in which hyperlipidemic rats were treated with the methanolic extractN.sativaand volatile oil.Both extract and volatile oil have TQ common which effectively ameliorate the hypolipidemic activity via hepatic 3-hydroxy-3-methylglutaryl-coenzyme A reductase pathway.Methanolic extract and volatile oil both can be used as a hypolipidemic agent[75].Another study also confirmsN.sativaantihyperlipidemic effect by a significant decrease in total cholesterol,low-density lipoprotein,and triglyceride[76].

Pulmonary protective activity

The study examines the beneficial effect of NSO on pulmonary aspirations in experimental lung injury in rats.The volatile oil treated rats showed significant inhibition of inflammatory pulmonary responses,reduced peribronchial inflammatory cell infiltration,alveolar septal infiltration,alveolar edema,alveolar exudate,alveolar macrophages,interstitial fibrosis,granuloma,and necrosis formation.It also showed a significant reduction in the activity of inducible nitric oxide synthase and a rise in surfactant protein D in lung tissue of different pulmonary aspiration models afterN.sativatherapy.N.sativacan subside lung injury and have potential clinical use[77].The relaxant effect of n-hexane,dichloromethane,methanol,and the aqueous fraction ofN.sativa(0.8,1.2,1.6 and 2.0 g%)was compared with theophylline(0.2,0.4,0.6 and 0.8 mM).The relaxant effect of different concentrations of methanol and dichloromethane fraction and n-hexane was significantly greater.It showed the potent effect ofN.sativawas mainly due to lipid fractions[78].The effect of nigellone and TQ on the trachea and their influence on respiratory clearance was studied.In the study,the effect of Ba++carbachol and leukotriene induced tracheal contraction and the transport of fluorescence dye rhodamin B concerning ciliary action was also studied using microdialysis.Nigellone and the higher concentration of TQ had a concentration dependent effect on the trachea and the contraction induced by leukotrience-d4were inhibited by nigellone and TQ.The mucociliary effect of nigellone was higher than TQ.It is suggested that nigellone is more effective than TQ and can be used in the treatment of respiratory diseases[79].

Gastro-protective activity

The NSO showed a promising effect on necrotizing enterocolitis.The histopathology,apoptosis evaluation,and severity of bowel damage were significantly lower and tissue GSH-Px and superoxide levels were preserved where MDA and myeloperoxidase(MPO)were lowered in necrotizing enterocolitis.This shows the potent antioxidant, anti-inflammatory;antiapoptotic properties of NSO and thus seems to be a potentially promising agent for protecting intestinal tissues from severe damage[80].The gastric effect ofN.sativapowder was noticed in alcohol induced gastric lesions in mice.There was a significant increase in gastric secretion whereas a significant decrease in gastric volume and reported a vigorous improvement in gastric disorders treated byN.sativa[81].The anti-ulcer effect ofN.sativaaqueous suspension against necrotizing agent which induces gastric ulcer was studied.N.sativasignificantly ameliorates the ulcer severity and basal gastric ulcer secretionN.sativatreatment appears to reinforce the mucosal barrier,which is the first line of defense against endogenous and exogenous ulcerogenic agents.The anti-ulcer effect ofN.sativais possibly prostaglandin-mediated or via its antioxidant and anti-secretory activities[82].A report on murine colitis induced by administration of dextran sodium sulfate and treated with TQ showed amelioration of colitis-related damage and significant reduction in colonic MPO activity and MDA levels and elevation in glutathione levels.It can be concluded that the antioxidant and anti-inflammatory effects of TQ can ameliorate inflammatory bowel disease[83].

Hepato-protective effect

In hepatic ischemia-reperfusion injury,N.sativawas infused intraperitoneally and there was a significant reduction in aminotransferase,ALT, lactate dehydrogenase levels,total oxidative status(TOS),oxidative stress index(OSI),and MPO and a significant increase in total antioxidant capacity(TAC)and histological tissue damage was milder inN.sativatreated rats[84].In a similar study,bile duct ligation induces cholestatic liver injury in rats on treatment withN.sativadecreased the liver enzymes level,TOS,OSI,and MPO and increased TAC levels significantly.N.sativatreated rats with biliary obstruction resulted in inhibition of necro-inflammation through attenuation of improved neutrophil infiltration and oxidative stress in liver tissue[85].In a study about CP induced hepatotoxicity,CP treatment caused an increase in ALT,aminotransferase,and lipid peroxidation,and decreased antioxidants and activities of various carbohydrate metabolism and membrane enzymes.NSO administration to CP rats showed reduced liver damage.NSO protected the CP induced hepatotoxicity by improving energy metabolism and strengthening antioxidant defense mechanisms[86].In this study,cadmium induces hepatotoxicity which significantly increased antioxidant enzymatic activities and protein carbonyl it also reduced glutathione content,and pretreatment with TQ showed an attenuating effect on protein oxidation and rejuvenation of depleted antioxidants of the cellular fraction.It shows the modulatory role of TQ in the liver[87].

Nephro-protective effect

A study was conducted to in vestigate about the gentamicin induced nephrotoxicity and the protective effect of NSO against gentamicin.Plasma creatinine and urea levels are significantly decreased in both higher and lower dosages ofN.sativaalso there were a significant reduction in MDA and nitric oxide generation and increased SOD and GSH-Px activities and renal damage was also reduced histopathologically.This impliesN.sativaacts as a potent scavenger of free radicals to prevent gentamycin induced nephrotoxicity[88].In similar study on paracetamol induced nephrotoxicity and protective role of ethanolic extract ofN.sativa.There was a significant decrease in creatinine,urea,MDA and significant increase in SOD and glutathione levels in the kidney.It suggested that the anti-inflammatory and antioxidant effect of NSE,as a potential therapeutic candidate for nephrotoxicity[89].Hyperlipidemic nephropathy induced by doxorubicin in rats showed a significant decrease in serum urea,triglycerides,total cholesterol and there was an elevation of non-protein sulfhydryl content and CAT activity upon TQ treatment in the kidney.The data suggest that due to the high antioxidant potential of TQ there was a suppression of doxorubicin induced nephropathy and TQ might be used as a protective agent of proteinuria and hyperlipidemia associated nephrotic syndrome[90].The report on TQ in cadmium induced nephrotoxicity,in rats there was a marked reduction in cadmium toxicity and the architecture of the kidney was preserved and in immunohistochemical analysis.TQ also produced a marked decrease in the expression of NF-кB in renal tissues and there was a significant decrease in apoptotic cells and lipid peroxidation.The finding suggested the nephro-protective potential of TQ might be due to anti-oxidant and anti-apoptotic properties[91].

Diuretic like effect

The diuretic effect of aqueous extract ofN.sativain rats in dose dependent manner(i.e.,10,30 and 50 mg/kg)with the reference group received furosemide(10 mg/kg),showed a significant diuretic,kaliuretic and natriuretic effects with no change in urinary pH.The diuretic index value showed good diuretic activity in NSE.The higher dosage of NSE significantly elevated the urinary volume and modified the concentration of urinary electrolytes;there were no signs of acute toxicity.N.sativahas a strong potential to be used as a diuretic agent[92].

Anti-osteosporotic effect

Osteoporotic effect ofN.sativawas studied in STZ induced diabetic rats.STZ treated rats showed a significant reduction in body weight and serum alkaline phosphate.Radiological changes were observed in femur like decreased bone density,bone softening,and cortical thinning.Histological changes like degeneration of osteoblast and osteocytes,multiple osteoporotic cavities,reduced collagen fibres,and irregular bone surface were also observed.Morphometrically,it showed a significant reduction in osteoblast number and immunohistochemically,there was reduced osteopontin protein in the bone matrix.InN.sativatreated rats,there was the improvement of biochemical, radiographic, histological,morphometrical,and immunohistochemical.N.sativaameliorates diabetic changes in bone[93].A similar study was conducted on STZ induced diabetic rats which can cause diabetic osteopenia.The analysis showed combined treatment withN.sativaand human parathyroid hormone showed enhanced bone mass and biomechanical behavior[94].The protective role ofN.sativaon osteoporosis produced by ovariectomy in rats.In ovariectomy rats,there was a reduction in plasma calcium level and elevation in alkaline phosphate,amino terminal collagen type 1 telopeptide,MDA,nitrates,TNF-α,and IL-6.These changes were reversed inN.sativatreated rats and the histological changes of tibias shown discontinuous eroded bone trabeculae with widened spaces in ovariectomy rats and a decrease in both cortical and trabecular bone thickness were also reversed inN.sativatreated.There was no mononuclear cellular infiltration and congestion of blood vessels was seen inN.sativatreated rats.This showsN.sativaas a potent anti-osteoporotic agent,which can be due to its high content of unsaturated fatty acid as well as antioxidant and anti-inflammatory properties[95].

Neuro-protective effect

A study was conducted to investigate the effect of hydro-alcoholic extract ofN.sativaand TQ on lipopolysaccharide induced depression-like behavior in rats.In whichN.sativawas given in two different doses(200 and 400 mg/kg of NSE)and TQ(40 mg).Two hours before conducting the forced swim test,lipopolysaccharide(100 μg/kg)was administered and NSE and TQ was administered the day before starting the experiment.In open field apparatus,N.sativaand TQ treated showed a lowered immobility time,crossing the number of peripheral and higher central crossing numbers when compared to lipopolysaccharide alone.It shows thatN.sativaof hydro-alcoholic extract and TQ have a beneficial effect on anti-depression[96].Repeated administration NSO for four weeks exhibited significant motor activity which shows the anxiolytic behavior.The concentration of serotonin(5-hydroxytryptamine)was increased and there was a decrease in the levels of 5-hydroxyindoleacetic acid in the brain and tryptophan levels were significantly increased in the brain and plasma.This study shows thatN.sativacan be a promising drug for antidepressant and anxiolytic[97,98].To study the role of gamma-aminobutyric acid-releasing(GABAergic)and nitriergic modulation in the anxiolytic effect of TQ,stress was given by immobilizing the rats for 6 hours and behavioral analysis was tested through the elevated plus maze and light/dark test.TQ was administered in two different doses(10 and 20 mg/Kg).TQ in higher doses decreased plasma nitrite and reversal of decreased brain gamma-aminobutyric acid(GABA)content[99].Using the Morris water maze test scopolamine induced spatial memory impairment and the effect ofN.sativaof hydro-alcoholic extract was evaluated.Two different doses of NSE were given(200 and 400 mg/kg)and scopolamine(2 mg/kg)was induced 30 min before Morris water maze test.InN.sativa(400 mg/kg)there was significantly shorter travel path level and latency when compared with scopolamine.In brain cortical tissues,acetylcholinesterase activity showed a significantly increase in scopolamine group and significantly decrease in NSE treated when compared to scopolamine group.After inducing scopolamine,MDA concentration was higher and decreased cortical thiol content was observed.In pretreated with NSE showed significant elevation in thiol content and reduction in MDA.N.sativaof hydro-alcoholic extract showed decreased acetylcholinesterase activity and oxidative stress in the brain.It shows the folk belief about the beneficial effect ofN.sativa[100].During neonatal and juvenile growth on learning and memory,the effect ofN.sativawas investigated.In which neonates were tested in morris water maze,where reduced time latency and distance traveled to reach the platform and there was a decrease in MDA concentration in hippocampal tissues and elevated total thiol content in the brain.This shows the positive effect of learning and memory in neonatal and juvenile.This may be due to the antioxidant effect ofN.sativa[101].NSO enhancement of spatial working memory performance was assessed using a radial arm maze.NSO treated rats exhibited no hindrance in the spatial memory performance.The results exhibited a declined mean number of errors was observed in NSO treated rats.It shows the ability of NSO to enhance learning and memory[10].The possible beneficial effect ofN.sativain comparison to methylprednisolone on experimental spinal cord injury was studied.Spinal cord injury was performed by placing an aneurysm clip extradural at T11-12.In spinal cord injury,there was a significant elevation of MDA and protein carbonyl levels and reduced SOD,GSH-Px,and CAT enzyme activities when compared to control.The study suggested the NSO neuroprotective and restorative effect on secondary pathochemical actions after spinal cord injury[102].

Ethanolic extract ofN.sativawas shown to have antiparkinson activity in chlorpromazine induced rats.The NSE was given in two different doses(200 and 400 mg/kg)and it reduced the cataleptic score.Levodopa,carbidopa,andN.sativaelevated the depleted level of reduced glutathione and total protein and decreased the elevated levels of thiobarbituric acid reactive substances and nitrite preferably at higher doses of NSE.Due to its anti-cataleptic and neurochemical response,N.sativais shown to have antiparkinson activity[103].The study to investigate the role of TQ on unilateral intrastriatal 6-hydroxydopamine-lesioned rats induces Hemi-Parkinson was conducted.After 1 week of the lesion,apomorphine caused contralateral rotations,a reduction in the number of neurons on the left side of substantia nigra pars compacta was observed,MDA and nitrite levels in the midbrain were increased and SOD were reduced in 6-hydroxydopamine lesions.But in pretreated TQ there was significant progress in turning behavior,prevented the loss of substantia nigra pars compacta neurons,and lowered MDA levels.The effect of TQ may be partially due to the attenuation of lipid peroxidation and it can provide benefit along with other therapies,in neurodegenerative disorder including Parkinson’s disease[104].Another study was conducted to investigate the protective role of TQ against Alzheimer’s amyloid-β peptide(Aβ).Cultured hippocampal and cortical neurons were treated with Aβ1-12 and TQ.TQ treated efficiently attenuated Aβ1-12 induced toxicity by enhancing cell viability.TQ inhibited mitochondrial membrane potential depolarization and reactive oxygen species generation caused by Aβ1-12.TQ also restored synaptic vesicle recycling inhibition,partially reversed firing activity.The study suggested that TQ may be a promising drug for Alzheimer’s disease[105].

The current study suggested the epileptiform ofN.sativain penicillin induced epilepsy and the mediation of neurotransmitters in epilepsy. In electroencephalographic data pretreatedN.sativasuppressed penicillin induced seizures by abolishing 85% high voltage spike discharges and there was the elevation of serotonin and reduced dopamine in the cerebral cortex,cerebellum,caudate nucleus,and midbrain.Decreased norepinephrine was observed only in the cerebral cortex.N.sativaadministration selectively alters the monoamine level in the different brain regions and similar to the anticonvulsive pattern shown by diazepam.The study suggests thatN.sativahas promising anticonvulsant action[106].Another study was conductedN.sativaaqueous extract,fixed oil,volatile oil,and its components TQ and p-cymene using pentylenetetrazole(PTZ)and maximal electroshock(MES)induced convulsions.All the constituents ofN.sativashowed a protective effect against PTZ-induced convulsion except fixed oil.Volatile oil and its constituent effectively suppressed MES-induced convulsion.All constituents showed a varying degree of minimal neurological deficit in the chimney test and TQ may be responsible for this act.GABAergic transmission is indicated as a pathway in the antiepileptic effect through the GABAAreceptor.Combination of valproate with TQ showed potentiation of valproate and reduction of median effective dose in both PTZ and MES induced convulsions.TQ can be used as an adjunct with valproate in seizure therapy[107].In another study action of intra-cerebroventricular injection of TQ in PTZ induced convulsion.TQ was administered in two different doses 200 and 400 μM in PTZ induced convulsion and resulted in prolongation of onset and reduced duration of tonic-clonic seizures and the lethality of doses was 45% and 50% respectively.The study also includes about GABAAreceptor and opioid receptors and the action of TQ through flumazenil and naloxone.TQ showed anticonvulsant activity,maybe an opioid receptor-mediated increase in GABAergic tone[108].

In a study with the albino mice,the analgesic effect of ethanolic extract ofN.sativawas assessed using acetic acid induced writhing test.When compared to diclofenac sodium,N.sativapossessed significant analgesic activity[109].In another study to assess the antinociceptive effect of NSO and TQ on dose dependent effect in response to the hot-plate test,tail-pinch test,acetic acid,and in the early phase of the formalin test,TQ showed a promising action.TQ attenuates the nociceptive response in the early and late phases of the formalin test.Naloxone significantly blocked NSO and TQ induced antinociception in the early phase of the formalin test.Furthermore,naloxone(μ1opioid receptor antagonist),naloxonazine or к-opioid receptor antagonist,nor-binaltorphimine significantly reversed TQ induced antinociception in the early phase but not in the late phase of the formalin test while naltrindole(δ-opioid receptor antagonist)did not affect either phase.This shows that NSO and TQ might have act via the supraspinal opioid system,particularly μ1andк-opioid receptor subtypes[110].

Effect on male reproduction system

A study was conducted to evaluate the role of TQ on testicular toxicity induced by methotrexate.In methotrexate,there was an elevation in TAC,and MPO activity and TQ treated animals showed a reduced TAC and prevent the increase in MPO activity.In histological studies,methotrexate showed an interstitial space dilatation,edema,severe disruption of the seminiferous epithelium,and reduced diameter of seminiferous dilatation,and TQ treated reversed the changes significantly.The study suggests that TQ may decrease the destructive effects of methotrexate on testicular tissues[111].Another study investigated the protective effect ofN.sativaagainst cyclophosphamide-induced toxicity on reproduction.The cyclophosphamide-induced alone showed a reduced weight of testis,epididymides,and sperm count.InN.sativatreated there was increased sperm count may be due to the antioxidant property of the plant which acts on the germline.N.sativarestored the histological architecture of testis and protective property against oxidative damage induced by cyclophosphamide.N.sativaalso promotes acrosome reaction during fertilization[112].The study examines the protective role of TQ on unilateral testicular ischemia reperfusion(I/R)injury in mice.In testicular torsion,there was increased TOS,OSI,and MDA levels.In TQ treated torsion group there was decrease in TOS,OSI,and MDA and did not affect total oxidative capacity and MPO activity,significantly reduced histological damage associated with I/R injury[113].The study was conducted to know the effect of TQ on testosterone and leutinizing hormone levels in STZ induced diabetic rats.There was a decrease in testosterone and leutinizing hormone in diabetic rats and TQ treated has resulted in increased testosterone and leutinizing hormone level was not significant with other groups[114].

A study was conducted to investigate the effect of alcoholic extract ofN.sativaon the fertility of male rats.At two different doses(0.5 and 1.5 g/kg)N.sativawas given and the results revealed that significant decrease of excitation time and the significant increase in body weight,reproductive parameters(seminiferous tubules thickness and diameters,the account of spermatogonia,primary and secondary spermatocytes,spermatids,free spermatozoa,the account of Sertoli and Leydig cells,diameter of Leydig cells and the height of epithelial cells entirely covered epididymal caudal),protein concentration,testosterone and follicle stimulating hormone and decrease in leutinizing hormone and cholesterol.This study suggested that daily oral administration improves male fertility[115].In a similar study,the alcoholic extract ofN.sativawas evaluated in the reproductive system of the male rat.The extract was given at two doses(200 and 400 mg/kg body weight)and the results showed a significant difference in testes and epididymides weight,sperm count,epididymal sperm reserve,daily sperm production,blood testosterone concentration,leutinizing hormone,and fertility index in both the lower dose group and the higher group as compared to the control group.The study suggested that a higher dose could increase fertility potential,leutinizing hormone,and testosterone concentration in male rats[116].Another study was conducted to examine the role of aqueous extract ofN.sativa(300 mg/kg)for 60 days.The treated group showed a significant increase in reproductive organ weight,sperm motility,and count in cauda epididymides,testicular ducts,and secretary activities of seminal vesicle and ventricular prostate and also increased spermatogenesis activity observed in seminiferous tubule.The results confirm that long term ingestion ofN.sativaincreases the effect on fertility and reproductive system in male rats[117].

Effect on female reproductive system

A study was conducted to evaluate the effect ofN.sativaon experimentally induced I/R injury in rat ovaries.After 3 hours of ischemia,the bilateral vascular clips were removed.N.sativa(500 mg/kg)was given 2.5 hours after induction of reperfusion.The results showed that I/R injury has increased MDA and MPO activity,while a significant reduction in SOD activity and glutathione levels when compared to sham was observed.N.sativabefore I/R reversed MDA,MPO,SOD,and glutathione activity and I/R increase the serum levels of IL-1,IL-6,and TNF-α while treated withN.sativareduced the serum levels of these cytokines.The report shows the effectiveness ofN.sativain reversing the tissue damage[118].A study was conducted to validate the effect of TQ in polycystic ovary(PCO)rats both in vivo and in vitro.In vivo study,PCO rat model treated with TQ showed significant restoration of normal physio-molecular behavior of the ovary,like reduced cysts formation,increased ovulation rate,and normalization of key ovarian factors(like TNF-α stimulated gene/protein 6,hyaluronan, hyaluronan-binding protein 1,cycloocygenases 2, matrix metalloproteinases(membrane type 1-matrix metalloproteinase,matrix metalloproteinases 9 and 2)),tissue inhibitor of metalloproteinases 1 and 2,and gelatinases matrix metalloproteinases 9 and 2 activity during follicular maturation.In vitro study,NF-кB nuclear translocation,cycloocygenases 2,and reactive oxygen species expression were inhibited via TQ supplementation in RU486-treated KK1 cells.Due to the TQ modulatory effect on NF-кB signaling,elevates normal ovarian phenotype and physiological function in the PCO model,demonstrating its potential to cure PCO in rats[119].The study describes the effect of aqueous extract ofN.sativaon the endometrium of the female reproductive system.N.sativawas given at doses of 0.2/100 g showed an increase in uterine wet weight,with intense and persistent diffuse of endometrial hypertrophy and enhanced glandulogenesis when compared to the control group.The aqueous extract ofN.sativaacts via stimulating the endogenous release of estrogen and progesterone[120].

According to the report of Aqel and Shaheen,the volatile oil ofN.sativainhibited the spontaneous movement of uterine smooth muscles and also the contraction induced by oxytocin stimulation in rat and guinea pig[121].It may be due to blocking of calcium influx via voltage dependent Ca2+channel and receptor-operated Ca2+channel byN.sativa,thus proving the anti-oxytocic potential.When hexane extract ofN.sativawas given orally to experimental rats it prevented the pregnancy in experimental rats at a dose of 2 g/kg daily on day’s 1-10 postcoitum.The active hexane extract exhibited only mild uterotrophic activity when compared to ethinylestradiol,but was without any estrogenicity in the immature rat bioassay[122].In another study the postcoital anti-implantation activity of ethanolic extract ofN.sativaat three different doses(100,200 and 400 mg/kg)were studied.The results showed that the anti-implantation effect and fertility index showed the increased impairment of fertility at a higher dosage.The anti-fertility showed a significant difference in all doses.The data suggested thatN.sativacan be used as a postcoital contraceptive in female rats[123].

Dermatological and cosmetics

Trichophyton verrucosumisolated from cattle suffering from trichophytosis suspected animals indicated hair loss,scurf scales,crusts,and keratinization.On treatment with NSE in combination with enilconazole on the infected area showed a strong anti-fungal activity.This study has suggested the usage ofN.sativaas an adjunctive or alternative agent in the treatment of dermatophytosis[124].The effect of NSE and TQ on pigmentation activity was studied by isolating melanophores fromHemidactylus flaviviridis.They were assayed using mean melanospore size index and their response was recorded.The study suggested thatN.sativaand TQ mimic the action of acetylcholine in melanin diffusion leading to skin darkening via stimulation of cholinergic receptors of muscarinic nature in the melanophores ofHemidactylus flaviviridis.N.sativaand TQ have a novel melanogenic for its clinical application in skin disorders such as hypopigmentation or vitiligo[125].In vitro sun protective factor(SPF)for NSO was evaluated using optometrics,LLC,SPF-290S instruments.The protection factor was calculated by the wavelength range from 290-400 nm as per United States Food and Drug Administration standards.Since the SPF value ofN.sativawas more than 2 it can be formulated for sunscreen[126].

Currently,many researches are conducted on herbal products to use as an alternative medicine for several diseases due to their efficacy,safety and low cost.Among many medicinal plants,N.sativahas a broad spectrum in traditional folk medicines.N.sativaalso promotes the body with its natural reviving process and cures the disease.N.sativahas many positive effects when used as remedies and/or as an adjunct for various diseases integrating by different systems of the body.N.sativaseed contains wide-ranging of fixed oils,proteins,alkaloids,flavonoids,tannins,saponin,and essential oils,thus proving its multiple mechanisms of action behind its therapeutical potential[127].TQ is considered to be the most active constituent ofN.sativaand has reported exhibiting potent antioxidant,anti-inflammatory,analgesic,neuro-protective,hepatoprotective,nephroprotective,and anti-cancer properties[128].N.sativaalso has other constituents which also serve to enhance its medicinal property.Since the complex synergic activity of the constituents improves the state of cellular nutrition,and gradually improves vitality[129].The pharmacological perspective ofN.sativacan be because of conspicuous antioxidant properties[130].Several in vitro and in vivo studies are piloted inN.sativaand its constituents.But still,the mechanism of action is unclear in some perception.The plant composition in therapeutic and pharmacological potentials is very much essential for advancing the research on other beneficial and unknown features.So,N.sativaand the plant derived constituents can be used to treat several diseases[16].The consolidated pharmacological actions ofN.sativais given below(Figure 3).

Figure 3 Pharmacological potentials of N.sativa.N.sativa,Nigella sativa;iNOS,inducible nitric oxide synthase;COX-2,cyclooxygenase-2;GABAergic,gamma-aminobutyric acid-releasing;AchE,acetylcholinesterase;GST,glutathione-S-transferase;GSH-Px,glutathione peroxidase;AST,aminotransferase;ALT,alanine aminotransferase;LDH,lactate dehydrogenase;LDL,low-density lipoprotein;SPF,sun protective factor;FSH;follicle stimulating hormone;LH,leutinizing hormone.

Conclusion

The medicinal plantN.sativawas broadly used as folk medicines and it has been reported to have many therapeutical and pharmacological activities such as antioxidant,antimicrobial,anti-diabetic,anti-cancer,anti-inflammatory, immunomodulatory,anti-osteoporotic,anti-hyperlipidemia and protective effect against cardiovascular system,pulmonary system,pulmonary protective,gastrointestinal system,liver,kidney,skin,central nervous system and reproductive system.In future research,theN.sativaseeds and its bioactive compounds should be isolated and investigated using animal models and clinical trials to understand its novel molecular mechanism of action.It can be concluded thatN.sativais a valuable medicinal plant that can be developed into pharmaceutical and nutraceutical products to treat a variety of diseases.