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. 2014 Mar 11;52(4):1881–1888. doi: 10.1007/s13197-013-1238-x

Evaluation of antioxidant activities of bioactive compounds and various extracts obtained from saffron (Crocus sativus L.): a review

Somayeh Rahaiee 1, Sohrab Moini 1,✉, Maryam Hashemi 2, Seyed Abbas Shojaosadati 3,✉
PMCID: PMC4375186  PMID: 25829569

Abstract

Saffron (Crocus sativus L. stigma), the most valuable medicinal food product, belongs to the Iridaceae family which has been widely used as a coloring and flavoring agent. These properties are basically related to its crocins, picrocrocin and safranal contents which have all demonstrated health promoting properties. The present review article highlights the phytochemical constituents (phenolic and flavonoid compounds, degraded carotenoid compounds crocins and crocetin) that are important in antioxidant activity of saffron extracts. However, the synergistic effect of all the bioactive components presence in saffron gave a significant antioxidant activity similar to vegetables rich in carotenoids. Our study provides an updated overview focused on the antioxidant activity of saffron related to its bioactive compounds to design the different functional products in food, medicine and cosmetic industries.

Keywords: Saffron, Antioxidant activity, Bioactive compounds, Saffron extracts

Introduction

A wide range of factors such as psychological stress, toxins, industrial lifestyle, infections, drugs, smoking, and obesity can cause many people to develop abnormally high levels oxidative stress (Makhlouf et al. 2011). Oxidative stress is considered to play an important role in a variety of neurodegenerative disorders such as Alzhemer’s disease, Parkinson’s disease and etc. (Soeda et al. 2007). Reactive oxygen species (ROS) and free radical (FR) have been implicated in the etiology of various human diseases including inflammation, cardiovascular disease, cancer, diabetes through induction of lipid peroxidation in cellular membranes (Choil et al. 2006). In addition to the body’s natural antioxidant defense for removing FR and ROS, there would be need to provide the body with a constant supply of antioxidants through dietary supplementation. Natural products have long been used to prevent and treat many diseases including cancer and thus they are good candidates for the development of anti cancer drugs (Papandreou et al. 2011).

Saffron (Crocus Sativus Linnaeus) is a perennial herb and belongs to the Iridaceae family (Fig. 1; ISO 3632 2011) which has been cultivated in Iran, Greece, Morocco, India, Spain and Italy (Ebrahim-Habibi et al. 2010; Pintado et al. 2011; Modaghegh et al. 2008). Although the source of saffron is obscure, it is apparently originated from Asia Minor and Iran (Ahmad et al. 2011). It is estimated that saffron world production is about 205 t per year. Annual production of Iranian saffron is 80 % of the total which 85 % was obtained from Khorasan province, where has the best quality saffron of worldwide (Ahmad et al. 2011).

Fig. 1.

Fig. 1

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Diagram of the organography of Crocus sativus L. (ISO 3632; 2011)

Saffron is used as one of the most commonly spices with flavoring and coloring properties in bakery products and beverages at a level of 260 ± 1 ppm (Selim et al. 2000). Many studies have demonstrated various pharmacological effects of saffron and or its important compounds including antioxidant activity, anti tumor, memory and learning enhancing, treatment of hepatic disorders, anti inflammatory, anti depressant and insulin resistant reducing (Sanchez-Vioque et al. 2012; Makhlouf et al. 2011; Umigai et al. 2011; Ghadrdoost et al. 2011; Amin et al. 2011; Dhar et al. 2009; Nkhaei et al. 2008; Ochiai et al. 2007; Soeda et al. 2007; Abdullaev and Espinosa-Aguirre 2004). All these properties have been attributed to the stigmas, the sole commercially valuable part of the plant whereas other parts of the plant have been much less studied. It should be noted that several in vivo studies in animals have been indicated a very low or even non-existent toxicity of both saffron and its extracts. Other reports showed the toxic effect of 5 g and more amounts, with a lethal dose of approximately 20 g/kg which explains why toxicology researches currently consider it to be safe for human consumption (Bathaie and Mousavi 2010; Melnyk et al. 2010). The saffron use in food applications has been recently increased, despite its high price due to changes in consumer preference toward natural products (Amin et al. 2011).

Carotenoids are the red, orange and yellow tetraterpene pigments found in plants, algae and microorganisms, and they are thought to function as important antioxidants and functional ingredients in foods (Gharibzahedi et al. 2013). Interest in the impact of saffron carotenoids on human health is due to their high antioxidant capacity which was investigated by many researchers (Makhlouf et al. 2011; Asdaq and Inamdar 2010; Karimi et al. 2010; Bathaie and Mousavi 2010; Tavakkol-Afshari et al. 2008; Kanakis et al. 2007; Soeda et al. 2007; Magesh et al. 2006; Ochiai et al. 2004; Selim et al. 2000; Verma and Bordia 1998; Nair et al. 1995). One of hypothesis for the modes of anti carcinogenic and anti tumor actions of saffron has been proposed the inhibitory effect on free radical chain reactions (Makhlouf et al. 2011). Many studies have been shown the antioxidant and free- radical scavenging activities of saffron. Figure 2 demonstrates the percent of done studies on the antioxidant activity of saffron in different years.

Fig. 2.

Fig. 2

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Percent of literatures on the antioxidant activity of saffron in different years (obtained from Scopus database)

Therefore, this review paper will outline the current understanding of the antioxidant and free-radical scavenging activities of saffron capacities and various bioactive compounds commonly found in this golden spice.

Saffron chemistry

Crocus sativus L. stigma

The stigmas of the saffron flower contain many chemical substances. Ranges of all chemical constituents can vary greatly due to growing conditions and country of origin. Among the estimated more than 150 volatile and several non volatile compounds of saffron, approximately 40–50 constituents have already been identified (Bathaie and Mousavi 2010; Melnyk et al. 2010; Abdullaev and Espinosa-Aguirre 2004). Table 1 illustrates the chemical composition analyses of saffron stigma.

Table 1.

Analytical results of main components of saffron (Hosseinpour-Chermahini et al. 2010)

Type of component Size (w/w %)
Proteins 12
Fats 5
Minerals 5
Crude fibre 5
Carbohydrate (starch, reducing sugars, gums, pectin and dextrins) 63
moisture 10
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The obtained data showed that red stigmatic lobs of the C. sativus flower contain three main metabolites crocins, picrocrocin and safranal. The crocins include crocin 1–4 which are a group of water soluble and red colored carotenoids formed by esterification of crocetin (a dicarboxylic 20-carbon carotenoid, C20H24O4) with different glycosids. Unlike most carotenoids, crocins are soluble in water due to a saccharide link with glucose, gentibiose or neapolitanose. Among the four crocins, crocin 1 or α-crocin is the most abundant in saffron and has been extensively studied for its antioxidant activity by quenching free radicals and protecting cells and tissues against oxidation. Besides, other minor carotenoids are also present in saffron such as carotenes, lycopene, zeaxanthin, mangicrocin and xanthone-carotenoid glycosidic conjugate (Licon et al. 2012; Maggi et al. 2011; Bathaie and Mousavi 2010; Hadizedeh et al. 2010; Asai et al. 2005; Li et al. 1999). picrocrocin (C16H26O7) which is a colorless glycoside and product of zeaxanthin degradation and is the main substance responsible for the bitter taste of saffron. Picrocrocin is the second most abundant component (by weight) accounting for approximately 1 % to 13 % of saffron’s dry matter. Safranal (C10H14O) is a product of natural deglycosylation of picrocrocin and responsible for the individual odor of saffron. Safranal represents about 30–70 % of volatile components and 0.001–0.006 % of dry matter of saffron. safranal is present in low amount in fresh stigmas and after drying of the stigmas or the passage of time, it is produced in high amount from its precursor (Maggi et al. 2011; Melnyk et al. 2010; Jalali-Heravi et al. 2009; Abdullaev and Espinosa-Aguirre 2004).

Saffron extracts

Aqueous methanol, ethanol or water is commonly used for the extraction of many bioactive constituents. Crocin as a water-soluble carotenoid can be effectively isolated from saffron stigma using the solvent-assisted extraction methods (Fig. 3). Effect of the use of water and different organic solvents such as ethanol or methanol at various concentrations on the total polyphenol content and antioxidant activity was studied for saffron (Makhlouf et al. 2011; Karimi et al. 2010). Makhlouf et al. (2011) have been studied the different conditions for extraction of saffron components. The following extracts were obtained from saffron soaked and boiled and extracted with water and methanol 50 % (v/v) using maceration method. Karimi et al. 2010 prepared saffron extracts using three different solvents of ethanol 80 % (v/v), methanol 80 % (v/v) and boiling water by shaking conventional method. In another study, the extraction yield of saffron components with water and 80 % (v/v) ethanol using a following maceration step (5 days at 4 °C) was respectively increased to 71.0 % and 58.4 % (Amin et al. 2011).

Fig. 3.

Fig. 3

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Preparation stages of saffron extracts

The results showed that solvent with different polarity had significant effect on polyphenol content and antioxidant activity. A high correlation between polyphenol content and antioxidant activity of saffron extracts was observed (Makhlouf et al. 2011; Amin et al. 2011; Karimi et al. 2010; Chen et al. 2008). Change in solvent polarity alters its ability to dissolve a selected group of antioxidant compounds and influences the antioxidant activity estimation.

Antioxidant activity of bioactive compounds of saffron

Phenolic and flavonoid compounds

Several researches have shown that spices containing phenolic and flavonoid compounds indicated antioxidant activities, so they are frequently used as antioxidant food supplements (Rahaiee et al. 2012; Karimi et al. 2010; Martinez-Tome et al. 2001). The antioxidant property of C. sativus stigma could be credited to its phenolic content as well as to its active ingredients such as safranal, crocin, crocetin and carotene, all of which have been reported to have antioxidant properties (Karimi et al. 2010). Table 2 shows the antioxidative properties and scavenging activity of bioactive compounds of saffron.

Table 2.

Biological functions attributed to Crocus sativus L. (saffron)

Bioactive compounds Type of function Ref.
Phenolic content
 Safranal ABTS radical scavenging activities
 Crocin DPPH radical scavenging activities Amin et al. 2011
 Crocetin Exhibition of significant reducing power
 Carotene
 Crocetin Having potent antioxidant
Showing protective effects against hepatotoxicity Genotoxicity		 Papandreou et al. 2011
 Crocin Potential of hypolipidemic
Antioxidant activity		 Asdaq and Inamdar 2010
 Crocin
 Safranal		 Antioxidant activity Karimi et al. 2010
 Crocin Prevention of the formation of peroxidized lipids Partly restored superoxide dismutase (SOD) activity Bathaie and Mousavi 2010
 Crocin
 Crocetin		 Antioxidant
Free radical scavenging activity		 Tavakkol-Afshari et al. 2008
 Crocin Treating neurodegenerative damage Soeda et al. 2007
 Crocetin
 Safranal		 Antioxidative properties Kanakis et al. 2007
 Crocetin Inhibition of lipid peroxidation
Increasing antioxidant status		 Magesh et al. 2006
 Crocin Antioxidant effect Ochiai et al. 2004
 Crocetin Antioxidant activity Selim et al. 2000
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Makhlouf et al. (2011) found that the total dosage of polyphenols in saffron (16 mg gallic acid/l) is higher than that in white grape juice (6.285 mg gallic acid/l). Their study showed that saffron extract obtained from the C. sativus flower had high amounts of polyphenol components which can effectively reduce activity of free radicals and can provide a strong protection for the different organs (liver, kidneys, lung and heart) against some oxidative damages under a dose-dependent behaviour. It was demonstrated that saffron as an excellent antioxidant source has more antioxidant capacity compared to white grape (Makhlouf et al. 2011).

The high dose of saffron was found to be more in counteracting the manifestation of hyperlipidemia than high dose of crocin. These propose that apart from crocin of saffron, there are other components responsible for synergistic antihyperlipidemic and antioxidant potential of saffron. The powerful hyperlipidemic activities can be directly linked with the presence of flavonoids in saffron as it is known that flavonoids have powerful hypolipidemic properties (Asdaq and Inamdar 2010).

Crocin and crocetin

Crocin bleaching assay was suggested as a suitable method for screening radical scavenging activity and assessment of total antioxidant activity of plasma. This method designed according to important property of crocin carotenoid as a basic element for the antioxidant activity of saffron (Bathaie et al. 2011; Bathaie and Mousavi 2010). Several studies suggested that sugars attached to the crocetin moiety probably play a key role in biological effects of crocins (Papandreou et al. 2006; Abe et al. 1998; Escribano et al. 1996). To the best of our knowledge, little work has been done to investigate the structure- antioxidant relationships of crocins. Thus, the protective effects of crocetin, crocin and saffron extract related to their structures require further in vivo comparative examination. Also, it is suggested that some of the variations in antioxidant capacity might be attributed to genotypic and environmental differences within species, the part of the plant studied, the time of year, the samples were taken and the analytical methods used (Shan et al. 2005).

Asdaq and Inamdar (2010) evaluated the hypolipidemic and antioxidant potential of saffron and its carotenoid pigment, crocin, in heperlipidemic rats. This study suggested the possible role of other constituents apart from crocin for potent antioxidant action of saffron. As discussed earlier, crocin, dimethyl crocetin, safranal and flavonoids quench free radicals and have antioxidant effects (Asdaq and Inamdar 2010). Although the importance of such characteristics for the activity of saffron water soluble carotenoids is not yet clear, it is accepted that the mechanistic paths for scavenging of free radicals are similar to those of the most known non polar carotenoids (Pham et al. 2000). It is postulated when bioactive molecules crocin and crocetin absorbed into blood plasma, they may modulate intracellular oxidative stress by activation of body’s natural antioxidant enzymes (Ordoudi et al. 2009).

Ochiai et al. (2007) assessed the antioxidant activity of each crocin by monitoring the inhibition of coupled auto-oxidation of ß-carotene and linolenic acid emulsion in cell lysates. They showed that crocetin, a polyene di carboxylic acid consisting of seven conjugated double bonds and four side-chain methyl groups, had a key role in the antioxidant activity of carotenoids while, picrocrocin had no activity.

Depriving the Pc-12 cells (pheochtomocytoma cells) of serum/glucose caused peroxidation of their cell membrane lipids and decrease intercellular superoxide dismutase (SOD) activity. Ochiai et al. (2004) investigated crocin’s neuroprotective effect by focusing on its initial action on the cell membrane of serum/glucose–deprived Pc-12 cells. The results suggested that crocin’s antioxidant effect is stronger than that of α-tocopherol.

In a study by Soeda et al. (2007), crocin, as the unique water-soluble carotenoids in the nature, was found to become active in maintaining normal serum/glucose-deprived cells morphology than α-tocopherol. They concluded that crocin may have potential for treating neurodegenerative damage induced by oxidative stress. If antioxidant activity varies directly as the number of double bonds, therefore crocin which has seven double bonds in a molecule should be as more effective antioxidant than α-tocopherol (Bathaie and Mousavi 2010; Soeda et al. 2007). Assimopoulou et al. (2005) believed that the antioxidant activity of C. sativus stigma could be attributed to two bioactive compounds, crocin and safranal. Both contents were affected by nature of the solvent used. Crocin and safranal showed high radical scavenging activity, 50 % and 34 % for 500 ppm solution in methanol, respectively. Zheng et al. (2005) showed that administration of crocetin caused an increase the resistance of LDL to in vitro oxidation and decreased plasma levels of oxidized LDL, indicating that crocetin can exert an antioxidant effect in vivo.

Cancer continues to represent the largest cause of mortality in the world and needs to anticancer drugs with high efficacy and low toxicity. Since the relationship between saffron compounds and cancer is an important problem, comprehensive studies need to be conducted further along the following issues such as determination of the mechanism(s) involved in the therapeutic properties of saffron, study the mechanism(s) involved in saffron cancer chemoprevention, determine the biologically active components of saffron and human studies to define efficacy of saffron in cancer treatment and prevention (Hosseinpour-Chermahini et al. 2010). Among the different hypothesis underlying the modes of anticarcinogenic and anti tumor actions of saffron and its bioactive components, it is suggested that the inhibitory effect on free radical chain reactions may be related to antioxidant properties of saffron especially crocin compound which is regarded as the most promising anticancer compound in saffron (Melnyk et al. 2010; Hosseinpour-Chermahini et al. 2010). Thus, it is proposed that saffron and constituents, alone or in combination with other chemical substances, could attain particular relevance in future treatment of some cancers.

Antioxidant activity of aqueous and alcoholic extracts of saffron

Aqueous extract

The saffron was found to be superior to crocin, safranal etc. indicating the involvement of other potential constituents of saffron apart from crocin for its synergistic behavior of quenching the free radicals (Asdaq and Inamdar 2010). The antioxidant activity of saffron extract is dose dependent (Botsoglou et al. 2005). The antioxidant activity of different extracts of saffron is reported in Table 3.

Table 3.

Antioxidant activity of different extracts of saffron

Type of extract Observation Ref.
Ethanolic extract
Methanolic extract		 Having radical scavenging activity Montoro et al. 2012
Ethanolic extract
Aqueous extract		 The highest phenolic content in ehtanolic E.
The best antioxidant activity in ethanolic E.		 Amin et al. 2011
Aqueous extract Reduction of free radicals Makhlouf et al. 2011
Aqueous extract Having antioxidant effects in chronic stress Ghadrdoost et al. 2011
Boiling water
Ethanolic extract
Methanolic extract		 Higher reductive potential (FRAP assay) in Methanolic extract
Showing strong free radical scavenging activity		 Karimi et al. 2010
Methanolic extract
Water–methanol extract		 Hydrogen peroxide scavenging activity Bathaie and Mousavi 2010
Methanolic extract Remarkable intracellular antioxidant activity Ordoudi et al. 2009
Ethanolic extract Showing good antioxidant activity by in vitro antioxidant models (Anti-hemolysis, DPPH radical scavenging Lipid peroxidation assay and phosphor molybdenum) Chen et al. 2008
Aqueous extract Reducing lipid peroxidation products
Increasing antioxidant power		 Hosseinzadeh et al. 2005
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Martinez-Tome et al. (2001) reported that saffron extract had significant hydrogen peroxide scavenging activity and lipid peroxidation inhibitory effect. Ghadrdoost et al. (2011) demonstrated the protective effects of saffron aqueous extract and crocin against chronic stress induced impairment of learning and memory. This finding suggested antioxidant effects of these substances in chronic stress.

Alcoholic extract

Phenolic and flavonoid content analyses of saffron extracts (boiling water, ehanolic and methanolic) by Karimi et al. (2010) revealed that different solvents had different contents of total phenolics and flavonoids. Phenolic and flavonoid contents were markedly higher in the methanolic extract; however, the antioxidant activity was affected by the nature of solvent used. Methanolic extract showed a higher reductive potential (by FRAP assay) and a strong free radical scavenging activity (by DPPH radical scavenging assay) than the boiling water and ehanolic extracts. This might be due to the presence of higher total Phenolics and flavonoids which play a major role in antioxidant capacities. Gallic acid as the major phenolic and pyrogallol as the major flavonoid compound in methanolic extract were also identified (Karimi et al. 2010). The antioxidant potential of ethanolic extract of c. sativus was also analyzed by Chen et al. (2008) who confirmed the antioxidant activities of this alcoholic extract using in vitro antioxidant modes including anti-hemolysis, DPPH radical scavenging, lipid peroxidation assay and phosphor molybdenum method. They evidenced that crocins in saffron extract have potent antioxidant effects and these properties are influenced by the sugars attached to the crocetin moiety. It was shown that the antioxidant activity of both methanol extract and water–methanol extract of saffron was higher than tomatos and carrots (Papandreou et al. 2006; Assimopoulou et al. 2005).

Referring to Ordoudi et al. (2009), the results reinforced the perception of saffron bioactivity through antioxidant mechanism of action. They pointed out that saffron methanolic extracts exhibit a remarkable intracellular antioxidant activity by using a single-cell model system (human monocyte) as effectively as the selected phenolic antioxidants such as rosmarinic acid, curcumin and Trolox.

Conclusion

Saffron is a useful medicine plant with a very low toxicity and shows the beneficial effect in many diseases. It acts as a multi potential drug and effects on various systems simultaneously. Natural antioxidants are more ideal as food additives, not only for their free radical scavenging properties, but also on the belief that natural products are healthier and safer than synthetic ones.

Saffron extracts and its biological active compounds including crocin, crocetin, carotene and safranal have been shown both in vivo and in vitro antioxidant property. Many studies showed the Pharmacological and biological effects of saffron especially its alcoholic extracts due to its antioxidant activity. However, the synergistic effect of all the bioactive components gave saffron a significant antioxidant activity the same as vegetables rich in carotenoids. According to the results, it can be concluded that there are various causes for antioxidant activity of saffron including glucose sugars attached to crocetin, polyphenolic compounds and the presence of more double bonds. This increases its therapeutic importance and calls for more studies especially as a promising chemotherapeutic agent in cancer treatment in the future. This finding above makes the spice an excellent candidate for being a functional food.

Acknowledgement

This work was supported by grants from the University of Tehran and Agricultural biotechnology research institute of Iran (ABRII).

Contributor Information

Somayeh Rahaiee, Email: s_rahaiee@ut.ac.ir.

Sohrab Moini, Phone: +98-26-32248804, FAX: +98-26-32248804, Email: dr_smoini@yahoo.com.

Maryam Hashemi, Email: hashemim@abrii.ac.ir.

Seyed Abbas Shojaosadati, Phone: +98-21-82883341, FAX: +98-21-82883381, Email: shoja_sa@modares.ac.ir.

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