Exploring the Use of Iris Species: Antioxidant Properties, Phytochemistry, Medicinal and Industrial Applications

The genus Iris from the Iridaceae family consists of more than 262 recognized species. It is an ornamental and medicinal plant widely distributed in the Northern Hemisphere. Iris species convey a long history as valuable traditional drugs with a wide variety of applications in various cultures, having been recorded since medieval times. Currently, Iris spp. still find application in numerous fields, including cosmetics, pharmaceutics and the food industry. Moreover, many of their empirical uses have been validated by in vitro and in vivo studies, showing that Iris spp. exhibit potent antioxidant, anticancer, anti-inflammatory, hepatoprotective, neuroprotective and anti-microbial properties. Phytochemicals investigations have revealed that the plant extracts are rich in phenolic compounds, especially flavonoids and phenolic acids. As such, they constitute a promising lead for seeking new drugs with high susceptibilities towards various health issues, particularly oxidative-stress-related diseases such as cancers, neurodegenerative diseases, cardiovascular diseases, diabetes, etc. Herein, we present a literature review of the genus Iris intending to determine the plant’s chemical profile and establish a coherent overview of the biological applications of the plant extracts with reference to their traditional uses.


Introduction
For millennia, medicinal plants have long been recognized as a valuable wellspring of natural agents with high curative properties; they currently continue to be a precious resource for seeking new drug leads [1]. The dissemination of synthetic drugs has raised serious concerns regarding their quality, efficacy and safety [2]. In contrast, natural products are environmentally and biologically friendly since they are easily recognized by body cells, permitting their metabolism to be performed [3]. As a result, medicinal and aromatic plants that have historically been used by traditional practitioners (fortunetellers, midwives, herbalists) are gradually being exposed to scientific research to separate their active ingredients in order to use them in modern dispensing forms [4].
One such plant species is the Iris species (spp.) ( Figure 1) (with 389 accepted species in the world according to (http://www.theplantlist.org/tpl1.1/search?q=Iris; accessed on 25 August 2021), a popular plant commonly used in landscaping due to its wide showy and colored flowers [5]. The plant draws its name from the Greek goddess of rainbows, referring to the wide range of bloom colors featured in Iris species [6]. The use of Iris species can be traced back to medieval painters and manuscript illuminators, by whom the plant's flowers were used to obtain "Iris green" and "Iris blue" pigments [7]. Likewise, the rhizomes of the plant were blended with other herbs, such as hyssop (Hyssopus officinalis), and used to treat skin conditions, whereas, during the nineteenth century, they were utilized to disguise tobacco smell and reduce bad-breath odors [7]. Currently, Iris species are still finding application in numerous sectors, including cosmetics, pharmaceutics and the food industry. In Morocco, the rhizomes of Iris species, commonly known as Orris roots, are used as one of the many ingredients in Ras el hanout, a Moroccan spice blend [8]. Similarly, I. germanica L. rhizomes are peeled and used as a flavoring in ice cream, confectionery, baked products and alcoholic beverages [7,9]. In Southern Europe, Iris species are still grown for commercial purposes and are used in tooth powder, toothpaste and teething rings [10], while in the cosmetic field, some Iris spp., such as I. florentina L. and I. germanica L., are currently used in the manufacturing of high-priced luxury perfumes and lotions such as "Iris Ganach"©, Guerlain; "Extravagance d'Amarige"©, Givenchy; "Chanel 19"©; and "So pretty"©, Cartier [10][11][12][13].
This review focuses on the ethnobotanical uses, chemical constituents and pharmacological properties of extracts and compounds derived from Iris spp. This work could provide a scientific foundation and necessary information for further investigations.
As such, a scientific literature search regarding botany, geographical distribution, ethnobotanical uses, phytochemistry and biological activities of the genus Iris was performed using different electronic databases, such as PubMed, Elsevier, Research Gate and Google Scholar. Keywords and phrases such as "Genus Iris", "Iris uses", "Iris phytochemistry", "Iris essential oils" and "Iris pharmacological activities" were used in the search.

Botany (Taxonomy, Geographic Distribution and Edaphic Conditions)
The genus Iris (Table 1) is a well-reputed rhizomatous plant belonging to the Iridaceae, a family of herbaceous, perennial and bulbous plants [5]. This plant comprises over 260 species widely distributed in temperate regions across the Northern Hemisphere, occurring particularly across North America and Eurasia, with approximately four species in northern Africa [21,22]. Although numerous Iris species have been found to be growing in mesic or wetland environments, the majority of Iris species thrives in montane, desert, semi-desert, or dry and rocky habitats [22]. Therefore, Iris species can withstand a wide variety of harsh environments, from cold areas where the hard grounds freeze to subtropical climates [10]. In terms of edaphic conditions, several Iris spp., such as I. aucheri (Baker) Sealy and I. persica L., prefer relatively acid soil, whilst the majority grows in slightly acid-alkaline soil, such as I. danfordiae (Baker) Boiss [5,10]. Some other species favor sunny borders with well-drained soil and full shade, whereas others thrive in dappled shade [10]. The genus Iris is identified by the basal fan of unifacial leaves, colorful perianth of three horizontal sepals and three upright petals that are basally fused into the tube and style branches that are fused at the base [24]. They are petaloid and distally expand beyond the tiny flap-like, transverse stigma as a bifid crest; they also have three stamens that are opposite to the sepals and are petaloid in style [22,24].

Ethnobotanical Uses
Our literature review identified the ethnopharmacological uses of 25 Iris species which have been documented through ethnobotanical surveys with indigenous peoples worldwide ( Table 2). The variety of cultural backgrounds and geographical distribution of Iris species across the world has led to a diversity of know-how related to the preparation of remedies, used parts, administration modes and treated ailments. Aside from culinary purposes, the data collected from these studies revealed that Iris species are mainly applied orally (66%) or topically (31%) to treat and relieve a wide range of health conditions ( Figure 2). Flowers (24%) and rhizomes (20%) are the most frequently used parts in folk medicine, whereas decoction is the main method for the preparation of remedies (22%) (Figure 2). In the ayurvedic system, the local communities belonging to the Monpa tribe in India use I. clarkei Baker-based paste to treat muscle pain [25]. For that, they crush dried flowers, stems, roots and leaves together to make a powder and blend it with local millet wine to prepare the paste, which is then applied topically to relieve muscle pain [25]. In the Trans-Himalayan region of India, I. lactea Pall is locally known as "Dres-ma". The whole plant is dried and powdered and a decoction is made and consumed orally to increase appetite and treat stomach cramps, small and large intestinal obstruction and food-poisoning disorders [26]. Moreover, diverse ethnics groups in the same region use I. hookeriana Foster-based paste as an expectorant and to treat sore throats [27]. They grind the dried roots into a powder and blend it with ghee/butter to prepare an oral paste [27]. Furthermore, the native tribes in the Lahaul and Spiti valleys take 10 g of seed powder orally to eliminate stomach worms and prevent the burning sensation [28]. Native American Indians (Cherokee) drink the tea made from the rhizomes of Iris spp. for gastrointestinal, renal and bladder problems [7]. Cherokee Indians also utilize a paste made from crushed rhizomes of I. virginica L. as a skin ointment [7]. In China, various parts of I. dichotoma Pall., such as leaves, rhizomes and seeds, are believed to cure colds, coughs and liver diseases [29]. To relieve gum swelling and toothache, native herdsmen in China cut the root bark into smaller fragments and bite them between the teeth [30]. The native ranchers believe that the suitable period to collect the roots of this plant is on 5 May in the Chinese lunar calendar [30]. According to the latest Chinese Pharmacopoeia, the rhizomes of I. germanica L. and I. pseudacorus L. are used to treat constipation and stomachache, and as a diuretic and carminative [15]. Similarly, the rhizomes of I. tectorum Maxim are consumed orally to relieve sore throat, remove phlegm and for heat clearing [14].
In Turkey, the rhizomes, roots and flowers of I. persica L., I. germanica L. and I. caucasica Hoffm are consumed as a snack (either alone or with bread) [31][32][33][34]. In Italy, I. germanica L. rhizomes are used for respiratory diseases, to strengthen children's teeth, against chilblains and as a vomiting agent [35]. Further details about the ethnobotanical uses of Iris spp., mode of preparations, routes of administration and used parts are collected and listed in ( Table 2). The below figures are based on more than 40 ethnobotanical studies conducted worldwide.
To summarize, it is critical to protect and improve Iris' medical expertise. Additional research is needed to document uses relevant to undocumented species; in vivo and in vitro studies are also required to validate other ethnobotanical usages, shed light on potential toxicities and determine safe dosages.

Ethnoveterinary Uses
In developing countries, similar to other types of traditional knowledge, ethnoveterinary practices have been handed down verbally from one generation to another for ages [36,37]. They refer to a complex system of methods, skills, beliefs and practices used to prevent, cure and maintain animal health [37,38]. Several ethnoveterinary studies have stated that traditional knowledge relevant to ethnoveterinary practices is mainly held by elderly people, especially men, who are commonly the ones who look after animal herds [39,40]. However, because of the rapid technological, socioeconomic and environmental changes, the continued transmission is endangered. Indeed, a significant amount of veterinary knowledge remains unrecorded and may be doomed to extinction with the death of their practitioners [37]. Without question, allopathic drugs hold an important place in managing several diseases. However, their uses have been associated with many drawbacks, such as chemo-resistance in livestock and the high cost of veterinary drugs, including antiviral and cytostatic drugs [37,41].
According to ethnoveterinary surveys, livestock producers in India and Pakistan use the two species I. kashmiriana Baker and I. hookeriana Foster for animal healthcare (Table 2). In the Bandipora district of Jammu and Kashmir, Bhardwaj et al. [42] reported that rhizomes powder of I. kashmiriana Baker, locally known as "Mazarmund", water and raw sugar are mixed together to make semi-solid balls that are fed to cattle as a tonic for general body weakness. In Pahalgam and Sonmarg, India, I. kashmiriana Baker is called "Kabriposh" and indigenous people use the plant flowers as an antiseptic to treat wounded livestock [43]. In Pakistan, an ethnoveterinary study showed that the paste made from green leaves of I. hookeriana Foster is administered to sheep as a vermifuge [44].

Pharmaceutical Uses
Nowadays, a handful of market-available dietary supplements and pharmaceutical medicines is composed of Iris species. "Laktir"©, a medication in the form of coated tablets made from the dried extract of milk-white Iris, is extensively recommended as an anti-inflammatory agent to cure acute and chronic inflammatory disorders [72,73], to alleviate the detrimental side effects of chemotherapy and during radiation sickness [72]. I. Versicolor L. rhizomes are among the major components of Mastodynon (Bionorica SE©, Neumarkt, Germany), a complex drug used to treat mastopathy and to relieve premenstrual and menstrual disorders [13]. Kaliris EDAS-114©, homeopathic drops prepared from I. versicolor L., is widely prescribed for chronic pancreatitis, gastric ulcers and gastritis [72]. "Vitonk"©, a multivitamin product, is a prophylactic drug manufactured from I. lacteal Pall leaves whose use is recommended for cancer patients [13]. Similarly, I. versicolor L. roots have been reported to exhibit some health benefits; they act synergistically with other herbs, such as Gum Guggul (Commiphora Mukul), to support thyroid dysfunctions such as subclinical hypothyroidism and Hashimoto's disorder [74].

Potential Application in the Food Industry
In recent decades, because of the drawbacks linked to synthetic additives, the demand for new natural food additives with less harmful effects on human health has been intensified [23]. One such strong natural-source candidate with a broad spectrum of applications in the traditional cuisine of different countries worldwide is the genus Iris. Due to its pleasant, sweet flavor, it is used to aromatize soft beverages, candies, chewing gum and bread flour in several countries [8]. Recent studies have revealed that the isolated compounds and crude extracts of this plant possess significant antioxidant and antimicrobial properties, especially against food-poisoning bacteria and fungi [13,23]. All these properties support the potential use of Iris-based extracts to expand the shelf life of foodstuffs and as flavoring agents.
Hydroxybenzoic acid derivatives occur particularly in the rhizomes of several Iris spp., such as I. schachtii Markgr., I.germanica L., I. pseudacorus L., etc. [75][76][77][78]. Gallic acid, a trihydroxybenzoic acid with high antioxidant and anticancer properties, seems to be the most abundant monomer in the rhizomes of I. hungarica Waldst. & Kit and I. variegata L., where its content was estimated at 2.362 ± 0.076 and 3.729 ± 0.134 mg/g, respectively [75]. The aerial parts and rhizomes of I. schachtii Markgr have been found to contain syringic acid, a dimethoxybenzene and a gallic acid derivative, with high content, noticed in the rhizome aqueous extract (90 ± 4 µg/g) [76]. Vanillic acid, a mono hydroxybenzoic acid listed as an intermediate metabolite in the conversion of ferulic acid to vanillin, has been found in the leaves, rhizomes and roots of several Iris spp., including I. bungei Maxim., I. florentina L. and I. germanica L. [76,78].
Hydroxycinnamic acid derivatives, another important subclass of phenolic acids found in Iris spp., are distributed in the leaves, roots and rhizomes (Table 3). They have mainly been found in the plant rhizomes, except for p-coumaric acid (11) and caffeic acid (7), which occur particularly in Iris leaves [75][76][77][78][79]. These phenolic compounds may partially explain the extensive ethnomedicinal uses of Iris spp. in various cultures across the world. Likewise, they constitute a potential source of chemicals with high antioxidants, inflammatory, neuroprotective and hepatoprotective potencies.
Likewise, rhizomes and roots have been discovered to be rich in flavonols , primarily peltogynoids Irisoids (A-E), irisflavones (A-D) and quercetin diglycosides (95)(96)(97), bearing galactose, glucose and rhamnose as the sugar moiety [78,94,96]. Dihydroflavonols are only represented by songaricol (107), identified in the rhizomes and roots of I. songarica Schrenk [94]. It is worth noting that songaricol has been found to exhibit substantial antioxidant activity [94]. Another identified group of flavonoids with potential antioxidant and antimicrobial properties is flavanonols. A total of four flavanonols have been detected in the rhizomes of I. dichotoma Pall., I. tenuifolia Pall and I. tectorum Maxim [92,93,97].
On the other hand, literature data from previous studies showed that Iris spp. Eos may exhibit great variability in chemical composition depending on the growing chemotypes (genetic variation), geographic origin of the plant and phenological stages. For instance, the sesquiterpenes aristolone (40.26%), Cuparene (10.88%) and β-Gurjunene (10.88%) were identified as the major compounds of I. bulleyana Dykes rhizome essential oil of plants grown in China, whilst fatty acids were not detected [131].
Moreover, Al-Jaber [132] proved that the chemical composition of Iris essential oils varies significantly depending on the physiological stage, with monoterpenes dominating (40.93%) in the pre-flowering stage and aliphatic hydrocarbons prevailing in the fullblooming phase.
To sum up, the genus Iris has been demonstrated to be a rich source of essential oils, containing fatty acids as the major class and myristic acid as the most abundant monomer. These compounds are endowed with substantial health benefits, suggesting the possible use of the essential oils of this plant in the pharmaceutical, food and cosmetics fields.  Ukraine [11,133]  Abbreviations, GC-MS: gas chromatography coupled with mass spectrometry; GC-FID: gas chromatography with flame ionization detector; L: Leaves; Rh: Rhizomes.

Antioxidant Activity
Antioxidants are stable molecules that scavenge free radicals and maintain a lowered redox state inside cells to prevent or postpone cell damage [134]. The imbalance between free radicals and antioxidants leads to oxidative-stress-related diseases, such as diabetes, cancers, atherosclerosis, and inflammatory and neurodegenerative diseases [135]. Recently, several synthetic antioxidants, such as butylated hydroxytoluene and butylated hydroxyanisole, were discovered to be harmful to human health [135]. As such, the quest for effective, non-toxic, natural substances with potent antioxidative effects has recently intensified.
Studies have shown that there is a substantial relationship between chemical composition and antioxidant activity. In particular, the contents of polyphenols, flavonoids and saponins are responsible for the antioxidant properties. Polyphenolic compounds act as antiradical activity, reducing agents, and complexes of pro-oxidant metals and quenchers of singlet oxygen, promoting the natural antioxidative defense mechanisms and protecting enzyme activity [136]. The genus Iris has been proven to contain substantial amounts of phenolic compounds, particularly flavonoids and their derivatives. Therefore, various extracts of this plant have been evaluated for their antioxidant potency.
Mahdinezhad et al. [137] investigated the in vivo protective effects of I. germanica L. hydroalcoholic extract at doses of 100 and 200 mg/kg on the liver and pancreas of a streptozotocin-induced diabetic rat model for 4 weeks. Accordingly, the repeated oral administration of the extract lowered the high level of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) compared with diabetic control rats. The extract also improved the liver antioxidant capacity (increase in thiol groups). The protective effect was ascribed to the significant amounts of flavonoids and anthocyanins in the hydroalcoholic extract. The authors supported the use of the plant as a natural antioxidant source to preserve the human body from free-radical-related disorders, especially diabetes mellitus and hepatic injury [137].
The in vitro antioxidant activity of Iris has been shown to be significantly correlated with the total content of phenolic compounds. The antioxidant activity of petroleum ether, chloroform and methanol crude extracts of fresh I. suaveolens Boiss & Reut rhizomes was tested using the β-carotene-linoleic acid and CUPRAC techniques; quercetin and butylated hydroxytoluene (BHT) served as positive controls [138]. The results disclosed that both petroleum ether and chloroform extracts exhibited pronounced antioxidant potency. Thirteen phenolic and flavonoid compounds were isolated from the petroleum ether and chloroform extracts and were screened in vitro for their antioxidant effects. Coniferaldehyde, a phenolic compound obtained from the chloroform extract, displayed the greatest activity among all the investigated compounds at 25 and 50 mg/mL in both β-carotene-bleaching and CUPRAC systems [138].
Moreover, the aqueous and ethanol extracts of I. germanica L. were evaluated for their in vitro antioxidant activity using several testing systems, namely, free radical scavenging, reducing power, superoxide anion radical scavenging, metal chelating activities and hydrogen peroxide scavenging [139]. The results indicated that at concentrations of 15, 30 and 50 µg/mL both aqueous and ethanol fractions exhibited excellent antioxidant properties, displaying 95.9, 88.4 and 79.9% and 90.5, 78.0 and 65.3% inhibition of peroxidation of linoleic acid emulsion, respectively. At concentrations of 20, 40 and 60 µg/mL, both extracts showed remarkable reducing power, free radical scavenging, hydrogen peroxide scavenging, metal chelating and superoxide anion radical scavenging activities [139].
Similarly, the antioxidant activity of the ethanolic extracts I. germanica L. areal parts and rhizomes was assessed using free radical DPPH scavenging and β-carotene-linoleic acid assays [79]. The results showed that, in the DPPH system, the aerial part and rhizome extracts exhibited significant IC 50 values of 5.38 and 12.3 mg/mL, respectively, while at the concentration of 3.15 mg/mL, the total antioxidant activity of the extracts was 98.7% and 97.4%, respectively [79].
In a recent study, the antioxidant activity of the petroleum ether, ethyl acetate and methanol extracts of I. ensata leaves was analyzed using various antioxidant assays such as the DPPH radical scavenging assay and FRAP (ferric ion reducing assay) [140]. Accordingly, all the extracts exhibited pronounced antioxidant potential. In addition, the study reported that the IC 50 values decreased with the increase in polarity. In the ferric reducing assay, the IC 50 values of the three extracts were found to be 226.66, 188.94 and 124.63 µg/mL, respectively [140].
The genus Iris contains substantial amounts of glycosylated flavonoids and phenolic acids, which are, generally, water-soluble products and can be detected in great quantities in the bloodstream, thus exhibiting high oral bioavailability. Due to all these properties, polyphenols are involved in a wide range of biological effects, such as antibacterial, antiinflammatory, antiallergic, hepatoprotective, antiviral, antithrombotic, anticarcinogenic, cardioprotective and vasodilatory effects.

Anticancer Activity
Recently, the use of anticancer drugs has been hampered by the emergence of several impediments, with these mostly being the cellular resistance to chemotherapy drugs and toxicities [141]. Therefore, the global trend is being shifted toward medicinal plants and plant-based compounds owing to their accessibility, affordability and effectiveness [141]. Several Iris-based compounds have been isolated from various extracts and tested in vitro ( Table 6) for their cytotoxicity and chemopreventive activities (Figure 3). Irilone, iriflogenin, genistein and iris kashmirianin are only a few of the flavonoids isolated from I. germanica L. that have been shown to exert chemopreventive benefits by reducing cytochrome P450 1A activity and enhancing NAD(P)H: quinone reductase (QR)activity [16].
Similarly, Tantry et al. [143] studied the in vitro cytotoxicity activity of a new alkylated 1,4-benzoquinone derivative obtained from the chloroform extract of I. nepalensis rhizomes against various cancer cell lines using the MTT colorimetric assay. The compound revealed remarkable cytotoxicity against HCT116 (colon carcinoma), HL-60 (blood cancer) and ZR-75 (breast cancer), with IC 50 values of 10 ± 1.1002, 34 ± 1.1205 and 31 ± 1.1001, respectively. Likewise, the cytotoxicity potential of two flavonoids, 7-O-methylaromadendrin and tectorigenin, as well as four iridal-type triterpenes, iritectols A and B, isoiridogermanal and iridobelamal A, isolated from the rhizomes of I. tectorum Maxim were assessed against four cancer cell lines using the SRB method (sulphorhodamine B) [144]. The results indicated that iritectol B, isoiridogermanal and iridobelamal A displayed identical cytotoxicity against both MCF-7 and C32 cell lines, with IC 50 values for a range of 11 µM and 23 µM. Moreover, they found that iritectol B exhibited a dose-dependent apoptotic effect against COR-L23, while both 7-O-methylaromadendrin and tectorigenin flavonoids were discovered to be capable of triggering cell-cycle arrest at the S and G2/M phases, respectively ( Table 6). In vivo experiments based on animal models and molecular targets involved in the anticancer effects studies are mandatory to confirm the anticancer potential of Iris spp.

Neuroprotective Activity
The neuroprotective activity of Iris spp. has been shown to be related to the presence of flavonoid compounds, which, interestingly, prevent brain-related diseases due to their powerful antioxidant effect. The neuroprotective effect of the total content of flavonoids extracted from I. tenuifolia Pall was assessed on cultured cortical neurons under oxidative stress induced via H 2 O 2 exposure [151]. Pre-treatment with I. tenuifolia Pall flavonoids prevented H 2 O 2 -induced cell death in cortical neuronal cultures. The study reported that the mechanism underlying the neuroprotective effect was related to the activation of both ERK1/2 and was enacted by flavonoid-triggered Shp-2 pathways.
Similarly, the in vivo neuroprotective potential of I. tenuifolia Pall ethanolic extract was evaluated for the first time in a middle cerebral artery occlusion model (MCAO) using C57BL/6J mice [152]. Accordingly, the applications of I. tenuifolia Pall ethanolic extract one hour before or immediately after the surgery outstandingly decreased the infarct size. However, treatment with the same extract less than one hour after surgery did not show any protective effect. The reduction in infarct volume is likely attributable to the richness of I. tenuifolia Pall in flavonoid compounds, which acted as protective agents in the MCAO model due to their significant antioxidant potential. The other factor that might be involved in the protective effect is the activation of both ERK1/2 stimulated by I. tenuifolia Pall flavonoids. The study likewise reported an increase in interleukin-6 concentration in blood plasma. However, the mechanism via which interleukin-6 exerted its protective effects was not determined.
In a similar approach, the in vitro neuroprotective activity of three iridals, namely, Spirioiridotectal A, Spirioiridotectal Band and Spirioiridotectal F, isolated from the ethanolic extract of the rhizomes of I. tectorum Maxim was evaluated at the concentration of 10 µM against serum-deprivation-induced PC12 cell damage using the MTT method [153]. The results revealed that all the tested compounds exhibited moderate neuroprotective effects against serum-deprivation-induced PC12 cell damage. Despite some promising results in terms of neurological disease prevention, the neuroprotective activities of Iris species are still poorly investigated. In vitro and in vivo studies are still mandatory, especially against neurodegenerative diseases such as Alzheimer's disease.

Hepatoprotective Activity
The in vivo hepatoprotective activity of the methanolic extract of I. spuria rhizomes was evaluated against paracetamol-induced hepatotoxicity in Wistar rats at the two doses of 100 and 200 mg/kg [154]. The results revealed an increase in serum enzymes and bilirubin level as a sign of hepatic injury in intoxicated rats. Interestingly, the administration of paracetamol along with I. spuria L. methanolic extract was shown to exert a dose-dependent protective effect, bringing the levels of ALT, AST, ALP and total bilirubin to normal ranges as a consequence. Furthermore, the study reported that the methanolic extract restored the serum levels of albumin and glutathione (GSH) and prevented both elevated triglyceride and lipid peroxidation [154].
Likewise, the in vitro hepatoprotective potential of three iridal metabolites, iridojaponal A, B and C, isolated from the ethanolic extract of I. japonica whole plant was assessed against N-acetyl-p-aminophenol (APAP)-induced toxicity in HepG2 cells [155]. Accordingly, iridojaponal A and B exhibited moderate hepatoprotective effects, with cell survival rates of 55.27 and 56.45%, respectively, while the positive control displayed a cell survival rate of 59.28%.

Anthelmintic Activity
Standard anthelmintic drugs are widely utilized against internal parasites and encompass several classes, such as benzimidazoles and avermectins. They are classified based on their chemical structure and mode of action [156]. Although synthetic anthelmintics have effectively been applied to control helminth infections, their usage has lately been hampered by nematode resistance; they may also affect the host itself and remain as residues in edible tissue [156]. These drawbacks have prompted researchers to look for alternate control strategies, such as using traditional medicinal herbs.
Data have shown that I. hookeriana Linn and I. kashmiriana Linn exhibit significant in vitro and in vivo anthelmintic activities. To corroborate the ethnoveterinary use of I. kashmiriana Linn, Khan et al. [157] evaluated the in vitro anthelmintic activity of I. kashmiriana Linn aqueous and methanolic extracts against Haemonchus contortus nematodes using the motility inhibition test. The positive control was the standard treatment Levamisole 0.5 mg/mL, while the negative control was 0.95% (PBS solution). The worms were exposed to 50, 25 and 12.5 mg/mL crude extracts and their motility was examined 0, 1, 2, 5 and 8 h post-exposure. After 6 h of treatment, the authors observed that the aqueous extract of I. kashmiriana inhibited worm motility by 85.0% at 50 mg/mL, whereas the methanolic extract exhibited better anthelmintic activity, displaying a mean worm-motility inhibition of 100.0%. The anthelmintic effect was attributed to the presence of alcohol-soluble and water-soluble active molecules in the extracts.
Using the same method, Tariq et al. [158] tested the crude aqueous extract and crude ethanolic extract of I. hookeriana Linn rhizomes against Trichuris ovis worms to validate the ethnoveterinary uses of I. hookeriana Linn. They proved that both extracts had significant anthelmintic activity and the highest worm-motility inhibition was exhibited by the ethanolic extract (84.6%) at 25 mg/mL. Likewise, I. kashmiriana aqueous extract at 2 g/kg body weight exhibited a maximum (70.27%) egg-count reduction in sheep naturally infected with mixed gastrointestinal nematodes after 15 days of treatment [158]. In the same way, I. hookeriana ethanolic extract at 2 g/kg displayed a maximum (45.62%) egg-count reduction in sheep naturally infected with mixed gastrointestinal nematodes after 10 days of treatment. The authors of both studies supported the application of I. hookeriana and I. kashmiriana as natural veterinary agents to control sheep gastrointestinal nematode parasites [157,158].

Antibacterial Activity
The ethanol/water extracts (70/30, v/v) of I. haphylla L. rhizomes at the concentration of 1% were tested in vitro against standard Gram-positive and Gram-negative bacterium strains. The optimal activity was noticed against the Gram-positive strains, Basillus subtilis ATCC 6633 and Staphyloccocus aureus ATCC 25923, with diameters of growth inhibition of 16.00 and 15.60 nm, respectively. Meanwhile, Gram-negative strains were relatively resistant to the plant extracts [159].
The ethyl acetate fractions derived from 70% of ethanolic extract of I. unguicularis Poir rhizomes at concentrations of 25, 50 and 100 µg/mL were investigated for their antibacterial activity against two Gram-positive and five Gram-negative bacterium strains using the disk diffusion method [18]. The best antibacterial activity was observed against S. aureus (11-23 mm zone of inhibition) followed by B. subtilis (8-13 mm zone of inhibition). The lowest activity was noticed against M. Morganii [18]. The antibacterial activity of the methanolic extract of I. pseudopumila Tineo rhizomes was assessed against four Gramnegative and nine Gram-positive strains using the broth dilution method [160]. The extract exhibited prominent inhibition against all the bacterial strains with minimum inhibitory concentrations (MIC) ranging between 7.8 and 250 µg/mL. It is worth mentioning that the Gram-negative strains, especially E. coli and E. aerogenes, were more sensitive to the Iris species extract.

Antifungal Activity
The in vitro antifungal activity of I. unguicularis Poir methanolic extract was tested against the Aspergillus Niger 2CA936, Aspergillus flavus NRRL3357 and Candida albicans ATCC1024 fungal strains [161]. The results revealed that the methanolic extract exhibited potent antifungal properties, mainly against Aspergillus Niger 2CA936. I. unguicularis Poir antifungal activity was attributed to the lipophilic properties of the phenolic compounds. The essential oils of I. persica L. extracted from flowers, leaves and rhizomes were evaluated against three human pathogenic fungal strains, Candida albicans, Trichophyton mentagrophytes and Microsporum canis, using the broth microdilution assay. All the extracts exhibited moderate antifungal properties.The study also reported that the highest antifungal activity was detected for essential oils extracted from leaves and flowers.
Moreover, the antifungal activity of iridal, a triterpenoid compound isolated from the rhizomes of I. germanica L., was performed against Plasmodium falciparum chloroquineresistant and -sensitive strains. Iridal was less effective against both fungal strains, with minimal inhibitory concentration values exceeding 50 mg/mL from 24 to 48 h of incuba-tion [19]. Furthermore, the ethanolic extract of I. hungarica rhizomes was evaluated in vitro against Candida albicans ATCC 653/885 at the concentration of 1%. The fungal strain was interestingly sensitive to the ethanolic extract, with 16.30 nm as a diameter of growth inhibition [159].

Antiviral Activity
The aqueous and ethanolic extracts of I. sibirica L. were evaluated against herpes simplex virus type 1. Accordingly, the rhizome ethanolic extract was the most effective on the herpes simplex virus when compared with the aqueous extract [162].

Antidiabetic Activity
Standard antidiabetic drugs, especially α-amylase and α-glucosidase inhibitors, have recently been linked to a number of serious side effects in humans, including diarrhea, bloating and abdominal pain [163]. Thus, researchers have switched their attention to a plethora of medicinal plants that have been exploited by indigenous people worldwide, which has led to a rich know-how related to diabetes treatment. Researchers have lent credence to their ethnomedicinal uses and identified many bioactive compounds endowed with substantial antidiabetic activity, primarily flavonoids and phenolic acids [164].
Although there are more than 260 accepted species of the genus Iris worldwide, data have shown that the only Iris spp. that have been evaluated for their antidiabetic activity are I. germanica L. and I. ensata Thunb. In this sense, Mahdinezhad et al. [137] studied the hypoglycemic effect of the hydroalcoholic extract of I. germanica L. rhizomes on streptozotocin-induced diabetic rats. The repeated oral administration of the doses of 100 and 200 mg/kg for 4 weeks significantly decreased the levels of glucose, triglycerides and oxidative stress markers levels such as ALT (alanine aminotransferase), AST (aspartate aminotransferase) and ALP (alkaline phosphatase). The authors stated that the antihyperglycemic and antihypertriglyceridemic effects of I. germanica L. could be attributed to the abundance of phenolic constituents in the hydroalcoholic extract, especially anthocyanins.
Furthermore, Suresh et al. [165] used normal, glucose-loaded and streptozotocininduced diabetic rats to evaluate the hyperglycemic effect of I. Ensata Thunb dried root extract for 21 days. The authors reported that the oral administration of the extract reduced blood glucose in both normal and streptozotocin-diabetic rats. They associated the observed effect with the capacity of the extract to lower the intestinal uptake of glucose (digestiveenzyme inhibition), increase the glucose absorption at the tissue level (sensitize the cells) and enhance the activity of the β-cells of the pancreas.
On the other hand, the increase in blood glucose levels is mainly ascribed to the degradation of carbohydrates in the intestine, which is under the control of α-amylase, β-amylase and α-glucosidase [166]. Inhibiting or slowing down the activity of these key enzymes might be an effective therapeutic approach for preventing glucose from entering the bloodstream [163].
Therefore, Ibrahim et al. [167] identified eight known isoflavonoids, as well as two novel isoflavonoids, 8-hydroxyirilone 5-methyl ether and 8-hydroxyirilone, from the methanolic extract of I.germanica L. powdered rhizomes. Using acarbose as a reference, they assessed the in vitro α-amylase inhibitory potency of these compounds. They reported that, among all the tested components, 8-hydroxyirilone 5-methyl ether, 8-hydroxyirilone, irilone and irisolidone exhibited prominent α-amylase inhibitory capacity at the concentration of 250 µg/mL with inhibition rates of 66.1, 78.3, 67.3 and 70.1%, respectively. They indicated that the α-amylase inhibitory potency increased with the presence of C-7 hydroxyl and C-5 hydroxyl or with the methylation of the hydroxyl groups in the A and B rings of isoflavonoids.

Toxicity
No reports have been published regarding the toxicity nor the side effects of Iris species. The available data recommend I. versicolor L. root extract at the daily dose of 400-2400 mg [47].
Likewise, the use of this plant is strongly inadvisable under some health conditions such as pregnancy or breastfeeding, as well as stomach or intestinal disorders, such as ulcerative colitis, infections or Crohn's disease (https://www.rxlist.com/blue_flag/supplements.htm; accessed on 25 May 2021). Hence, in-depth toxicological studies are strongly required to assess the safe use of Iris species.

Conclusions and Perspectives
The genus Iris is an ornamental and medicinal plant widely distributed in the Northern Hemisphere. The genus Iris has long been used to treat and relieve a wide range of health conditions, including liver and spleen diseases, chronic pancreatitis, cancers, inflammation and bacterial and viral infections. Moreover, this plant is widely used in aromatherapy and in the industry of luxury perfumes due to its violet-like smell. For decades, Iris species have been the subject of numerous phytochemicals and biological studies, leading to the extraction and identification of various compounds belonging to several classes, such as flavonoids, phenolic acids, terpenes, fatty acids, aliphatic hydrocarbons and aldehydes.
On the other hand, several empirical uses of Iris spp. have been validated through in vitro and in vivo studies, demonstrating that the isolated compounds and crude extracts of this plant exhibit potent antioxidant, anticancer, hepatoprotective, neuroprotective, antidiabetic and antimicrobial properties. The powerful antioxidant and antimicrobial potencies of various extracts of this plant could support their potential use as natural antioxidants and antimicrobials agents against multiple pathogenic bacterial and fungal strains in foodstuffs and as good alternatives to synthetic additives.
More interestingly, the significant amounts of glycosylated flavonoids and phenolic acids in the plant extracts are generally water-soluble products and can be detected in great quantities in the bloodstream, thus exhibiting high oral bioavailability. The latter is a key parameter in drug development, as it quantifies the proportion of an absorbed active substance and its availability to produce pharmacological effects, rendering them potent candidates for the development of new drugs against oxidative-stress-related diseases, including diabetes, neurodegenerative diseases, cardiovascular diseases, etc. Despite the rich literature on the plant, the chemistry and biology of Iris spp. have yet to be thoroughly addressed.
Further studies regarding plant toxicity are mandatory to avoid any eventual hazardous effects on human health before proceeding with the elaboration of any pharmaceutical formulations, as the published in vivo and preclinical studies of different Iris extracts are extremely scarce. In-depth investigations are required to validate other traditional practices involving Iris spp.
Author Contributions: L.B. and S.K. designed the study, drafted the manuscript and collected and arranged the references; L.B. and C.F. analyzed the data, reviewed and edited the manuscript, supervised the final version of the paper. All authors have read and agreed to the published version of the manuscript.
Funding: This research received no external funding.