Flavonols and Flavones as Potential anti-Inflammatory, Antioxidant, and Antibacterial Compounds

Plant preparations have been used to treat various diseases and discussed for centuries. Research has advanced to discover and identify the plant components with beneficial effects and reveal their underlying mechanisms. Flavonoids are phytoconstituents with anti-inflammatory, antimutagenic, anticarcinogenic, and antimicrobial properties. Herein, we listed and contextualized various aspects of the protective effects of the flavonols quercetin, isoquercetin, kaempferol, and myricetin and the flavones luteolin, apigenin, 3′,4′-dihydroxyflavone, baicalein, scutellarein, lucenin-2, vicenin-2, diosmetin, nobiletin, tangeretin, and 5-O-methyl-scutellarein. We presented their structural characteristics and subclasses, importance, occurrence, and food sources. The bioactive compounds present in our diet, such as fruits and vegetables, may affect the health and disease state. Therefore, we discussed the role of these compounds in inflammation, oxidative mechanisms, and bacterial metabolism; moreover, we discussed their synergism with antibiotics for better disease outcomes. Indiscriminate use of antibiotics allows the emergence of multidrug-resistant bacterial strains; thus, bioactive compounds may be used for adjuvant treatment of infectious diseases caused by resistant and opportunistic bacteria via direct and indirect mechanisms. We also focused on the reported mechanisms and intracellular targets of flavonols and flavones, which support their therapeutic role in inflammatory and infectious diseases.


Introduction
Flavonoids are an essential and diversified class of secondary plant metabolites present in different concentrations in the leaves, flowers, roots, and fruits. Flavonoid concentrations are influenced by natural factors such as temperature, ultraviolet (UV) radiation, season, pollutant, drought, and salinity stress owing to their effect on plant metabolism (Li [1]).
Inflammation is a complex biological response to noxious stimuli. It protects organisms against pathogens, cell damage, and irritating chemicals. Moreover, it initiates the healing process, which involves different cells and chemical mediators [6,7]. Nevertheless, exacerbated and uncontrolled inflammation can also cause deleterious tissue damage. Flavonoids can alter cellular responses and production of chemical mediators related to inflammatory processes [8] caused by many agents, including natural toxins, pharmaceuticals, heavy metals, and environmental chemicals [9].
Oxidative stress is induced by mitochondrial dysfunction and inflammation [10,11]. Continuous oxidative stress generates reactive oxygen species (ROS) and reactive nitrogen species (RNS), causing lipid peroxidation, protein and nucleic acid oxidation, and even cell death [12]. These effects are dependent on redox balance because they act as damaging agents and physiological regulators of cell functions [13]. Plant extracts are valuable sources of bioactive compounds with antioxidant properties. For example, honey shows antioxidant and anti-inflammatory properties owing to its high flavonoid content [14][15][16]. Moreover, honey intake led to reduction in malondialdehyde and ROS levels generated by high physical activity in athletes and murine models [17][18][19][20]. A recent review discussed the role of consuming plant foods rich in antioxidants in protection against cognitive decline [12].
Similar to all secondary plant metabolites, plant defense mechanisms include flavonoid production; for example, a plant under attack by microorganisms may trigger the production of flavonoids having antimicrobial effects [21]. Herein, we discussed many aspects of the protective effects of flavonols and flavones in detail.

Flavonoids: Structural Characteristics and Subclasses
Flavonoids present a C6-C3-C6 carbon skeleton structure that consists of at least two aromatic rings, called A and B, linked by the three carbons chain that can form a heterocyclic ring containing oxygen (ring C) with ring A (Figure 1) [175]. Flavonoids are divided into flavones, flavonols, flava-nones, flavanols, isoflavones, leucoanthocyanidins, anthocyanidins, and chalcones [5] ( Figure 2). The subclasses are determined by minor structural variations, including the absence of ring C, position of the bond between ring B and ring C, degree of unsaturation, and ring C oxidation [22,23]. In addition, substitutions at these rings can lead to differences in hydrogenation, hydroxylation, methylation, malonylation, sulfation, and glycosylation, which determine different pharmacological activities [5]. Currently, there have been reported more than 9000 different flavonoids [24]. We focused on two groups: (a) flavones that have a double bond between C2 and C3 in the flavonoid skeleton, without substitution at C3 and oxidized at C4, and (b) flavonols that differ from flavones by a hydroxyl group at C3 (Figure 2).
2.1. Flavonols and Flavones: Importance, Occurrence, and Food Sources. Almost all natural flavonoids exist in plants in O-glycosides or C-glycosides forms. C-glycosides are sugar moieties combined directly to flavonoid backbone as C-C covalent bonds. O-glycosides are formed by attaching sugar to hydroxyl oxygen [25]. The most common sugars attached to these flavonoids are D-glucose and L-rhamnose, but at least eight monosaccharides or combinations thereof may bind to different hydroxyl groups of flavonoids, resulting in a large number of known glycosides [26,27]. They are widely present in plants and are potential metabolites involved in plant signaling and defense mechanism. Furthermore, they are vital ingredients in human diet and have significant health benefits [28][29][30]. Flavonols, which are primary flavonoids in nature, are found in various fruits and vegetables, such as apples, berries, grapes, tomatoes, and onions, and play a key role in attracting pollinators and seed disseminators [27,31].
Flavones and flavonols have antioxidant effects and are essential for protecting plants from UV radiation [32,33]. Quercetin (3,3 ′ ,4 ′ ,5,7-pentahydroxyflavone) is one of the most representative flavonoids, widely distributed in nature, and an abundant dietary flavonol, found in food, such as onions, peas, and apples [22,26]. Quercetin-3-O-rutinoside, known as rutin (3,3 ′ ,4 ′ ,5,7-pentahydroxyflavone-3-rhamnoglucoside) and also called vitamin P due to contribute to reducing the permeability of blood vessels, was introduced in the market and has been explored for important pharmacological effects [34]. Luteolin (3 ′ ,4 ′ ,5,7-tetrahydroxyflavone) and apigenin (4 ′ ,5,7-trihydroxyflavone) are the main flavones found in food, such as onions, celery, red pepper, and grapes, and have a wide range of effects in biological systems, including anti-inflammatory and anticarcinogenic [35]. Luteolin is found in cereals and herbs, and glycosylated luteolin in vegetables such as carrots and broccoli [31,36]. Luteolin inhibits cancer in head and neck squamous cell carcinoma (HNSCC) xenograft in a mouse model by inhibiting p300 lysine acetyltransferase activity via acting at multiple levels of gene expression, miRNA expression, and miRNA processing [37]. Luteolin effectively prevented pulmonary and hepatic fibrosis [38,39] and reduced cancer cell proliferation [40]. Both effects are due to the reduction in oxidative  Oxidative Medicine and Cellular Longevity stress and inflammatory responses [41]. Interestingly, luteolin can also play a protective role in inflammation and cancer by modulating the microbiota ( [42]. The flavonoid hesperidin possesses antioxidant and anti-inflammatory properties and plays a role in inhibition of tumor cell metastasis and angiogenesis [43]. Table 1 shows some flavones and flavonols aglycones and its natural sources and content (mg/ 100 g or mg/100 ml).

Inflammation.
Inflammation is a complex response initiated by an insult by chemical, physical, or biological agents, which leads to resolution and promotes tissue repair [54,55]. During the inflammatory process, immune system cells, such as neutrophils, monocytes, eosinophils, and lymphocytes, migrate to the lesion site [56,57]. The lipid mediators eicosanoids and cytokines produced during the inflammatory response cause vascular and cellular alterations such as vasodilation, an increase in vascular permeability and activation, adhesion, and cellular transmigration [54]. The increase in the concentrations of inflammatory cytokines, such as interleukin (IL)-1β and tumor necrosis factor (TNF)-α, is crucial for leukocyte migration to the lesion [58][59][60]. The production of mediators and increased vascular permeability generate an exudate rich in plasma proteins [61]. The production of inflammatory mediators leads to endothelial increased permeability and leukocyte activation and migration to the injury site, cellular, and tissue damage and triggers the characteristic symptoms of inflammation, known by the five cardinal signs: pain, redness, heat, edema, and loss of function [54,62]. When leukocytes reach the inflammation site, they are activated and respond to eliminate noxious stimuli, thus releasing the content of their toxic granules, which includes ROS, reactive nitrogen species (RNS), proteinase, cathepsin G, and elastase [61,63].
Inflammatory responses must be fine-tuned because if exacerbated, it will damage tissues and, if insufficient, will not cause complete elimination of pathogens. Exacerbation of immune response with uncontrolled inflammation results in poor outcomes [68]. Drugs or natural compounds with strong anti-inflammatory effects allow microbes to grow and decrease survival [69]. However, adequate antiinflammatory effects may prevent organ damage and improve host survival [69,70].

ROS and Oxidative
Stress. ROS are presented by one or more unpaired electrons produced in cells during mitochondrial oxidative metabolism and in cellular response to xenobiotics, cytokines, bacterial invasion, UV radiation, and pollutants, collectively known as oxidizing agents [71]; they promote cascade reactions and cause oxidative damage [71,72].
Normal metabolism generates reactive species that play a prominent role in physiological functions, including gene expression, cell signaling, and immune responses. Antioxidants neutralize reactive species in excess [71,72]  3 Oxidative Medicine and Cellular Longevity antioxidant is a molecule able to donate an electron to a rampaging free radical and neutralize it [73]. Some antioxidants can interact with other antioxidants regenerating their original properties called the "antioxidant network" [74]. They neutralize ROS by different mechanisms including directly scavenging free radicals, directly scavenging free radicals, and inhibiting lipid peroxidation between others [75]. Antioxidant system can be enzymatic or nonenzymatic. The nonenzymatic system blocks the initiation of oxidation. The enzymatic system comprises antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR). In the nonenzymatic antioxidant system, glutathione (GSH); ascorbate (AsA); numerous simple Nobiletin Grape fruits, teas (Camellia sinensis), herbal tea, and black tea n.d. [53] n.d.: not described; * mg/100 ml. 4 Oxidative Medicine and Cellular Longevity phenolic acids such as hydroxybenzoic acids, hydroxycinnamic acids, and different flavonoid compounds; carotenoids such as xanthophylls, beta-carotene, and α-carotene; vitamin E; and pigments such as chlorophylls, betaxanthins, betalains, and betacyanins such as amaranthine, isoamaranthine, betanidine, and isobetadine are the most noticeable components ( [76]). Antioxidants act by stabilizing ROS. An imbalance in ROS production and antioxidant activity in biological systems causes oxidative stress. As a result of this imbalance, cellular dysfunction may occur via modulating gene expression, protein stability, and membrane fluidity, leading to cell damage and death [72]. An increase in reactive species levels occurs with aging and the development of diseases, including cancer, Parkinson's disease, diabetes, and cardiovascular diseases [77]. ROS increase is linked to different diseases, reviewed by [78]. For instance, excessive ROS accumulation leads to glutathionylation modification and proteasomal degradation of MAP kinase phosphatase-1, which prevents insulin signaling. Also, ROS in excess inactivates AKT, inhibiting glucose uptake, which eventually causes type 2 diabetes. In addition, mitochondria-derived ROS elicit the production of NLRP3 and IL-1β, accounting for an inflammatory phenotype in atherosclerosis pathogenesis. Furthermore, phosphorylated tau protein and Aβ are risk factors in Alzheimer's disease, which originate from the downregulated Nrf2 and increased ROS burden. Finally, extracellular and intracellular ROS activate TGF-β signaling triggering the epithelial-mesenchymal transition of epithelial cells and survival of circulating tumor cells during cancer metastasis [78]. The search for new antioxidant compounds as therapeutic agents for diseases associated with imbalance in reactive species production and clearance is gaining attraction of researchers. Natural antioxidants, such as phenolic compounds (flavonoids and phenolic acids), have emerged as primary therapeutic agents as nonenzymatic antioxidants because they can sequester ROS [79]. They also act as antiinflammatory agents.

Limitations
Associated with the Use of Flavonoids. We consume fruits and vegetables that containing flavonoids, such as apples, berries, onions, and broccoli, on a daily basis [80,81]. However, their bioavailability can vary. A trial conducted with 97 humans showed that the plasma concentrations of total polyphenolic metabolites was 0-4 μmol/L after an intake of 50 mg aglycone equivalents, and the urinary excretion was 0.3-43% of the ingested dose [82]. Flavonoids are absorbed in the small intestine. Flavonoids that cannot be absorbed in the small intestine are degraded in the colon by microorganisms to phenolic acids [83].
The absorption of flavonoids from food was long considered to be very low, as they are present in foods linked to sugars such as β-glycosides (except catechins). Only aglycones forms were considered able to pass the gut wall, and no enzymes that can split these predominantly β-glycosidic bonds are secreted into the gut or present in the intestinal wall [84]. Unlike terpenes with high bioavailability, flavonoids can have low bioavailability due to the extensive first-pass effect and glucuronidation [85]. However, research on the mechanisms for aglycone transfer across the gut wall is lacking. In a study on ileostomized humans, the absorption of orally administered quercetin was 24%. These data show that humans absorb appreciable amounts of quercetin and that absorption of glycosides in the small intestine is possible. Humans consume, on average, between 0.45 and 1.17 mg of apigenin daily [86]. Apigenin when taken orally (whose bioavailability is about 30%) is systemically absorbed and recirculated by enterohepatic and local intestinal pathways [87].
Despite these concerns about bioavailability, which seems to be influenced by their chemical nature, flavonoids effectively prevent or improve diseases. There are three possible explanations for this: (1) act in the lumen or intestinal epithelium, (2) act in the peripheral tissues even at very low circulating levels, or (3) the microbial metabolites of flavonoids are also active in the peripheral tissues [88]. Flavonoid amount consumed through food is generally nontoxic. However, when consumed in large amount, they can show toxic effects. Administration of high doses (≥200 mg/kg) and prolonged use of luteolin may increase hepatic enzyme activity; however, no significant liver damage was observed, whereas low doses (≤50 mg/kg) are hepatoprotective [89].
The purification or production of bioactive compounds on a large scale is challenging. Although a specific bioactive compound has been identified and proven to be beneficial in treating inflammatory or infectious diseases, its purification or bioengineering remains a challenge. Moreover, where bioactive flavonols have been identified in natural food, the among ingested is difficult to determine because it varies from place to place depending on the environment in which the plant grows. Another crucial challenge is the development of optimum delivery formulations. Nevertheless, the flavonoid properties in biological systems are indisputable because they are bioactive compounds with the potential to treat or prevent diseases; therefore, future studies based on better delivery systems, such as nanoformulations, should be encouraged [90,91].

Flavonols and Inflammation.
The current strategies for treating inflammatory and infectious diseases often present side effects that markedly affect the quality of life of patients. In addition, the use of immunosuppressants may be essential to control the immune response in infectious diseases; however, it can increase the risk of spread of the infection and should be used with caution. Flavonols, such as quercetin, kaempferol, and myricetin, are the most common plant flavonoids. We discussed recent studies on anti-inflammatory effects of flavonols and listed them in Table 1. 2.6. Anti-Inflammatory Effects of Quercetin, Isoquercetin, Kaempferol, and Myricetin. First isolated in 1857 from oak (Quercus), quercetin is probably the most investigated flavonoid. It has strong and prolonged anti-inflammatory effects [75,92,93]. Quercetin acts through several mechanisms such as by inhibiting lipopolysaccharide (LPS)-induced TNF-α production in macrophages [94], inhibiting LPSinduced IL-8 in A549 lung carcinoma epithelial cells [95], and inhibiting the production of inflammatory mediator-5 Oxidative Medicine and Cellular Longevity producing enzymes such as COX and LOX [96,97]. Quercetin showed cardioprotective effect against isoproterenolinduced myocardial infarction in rats [98] (Figure 3). Additionally, it reduced levels of biomarkers of ischemic induction, such as creatine kinase myocardial band (CK-MB) and cardiac troponin I (cTnl), inhibited inflammatory cell infiltration into the myocardium, reduced the expression of TNF-α and IL-6, and inhibited upregulation of calpain genes 1 and 2 that protect the myocardium [98].
Oral administration of quercetin for the treatment of arthritis induced by inactive Mycobacterium butyricum in a rat model showed that quercetin could diminish inflammatory markers, such as IL-1β, C-reactive protein, and MPC-1 and restore the plasma antioxidant capacity. In addition, it inhibited 12/15-LOX activity in the lungs and liver and increased heme oxygenase-1 (HO-1) expression in joints and lungs [102]. Quercetin treatment decreased inflammation and attenuated pathological alterations in the pancreas and ileum of animals with acute necrotizing pancreatitis. This suggests that the flavonoid acts by partial inhibition of TLR4/MyD88/P38 and MAPK ERK, thus reducing inflammation and interrupting intestinal damage [103]. In a model of monohydrate crystal-induced arthritis fatty acids activates TLR2 inducing interleukin-1β production via the ASC/caspase 1 pathway [104]. In addition, quercetin inhibited the release of ROS, IL-1β, and IL-18, resulting in the protection of mitochondrial integrity in Caco-2 cells stimulated with Escherichia coli [105]. It also blocked the inflammasome NLRP3 activation in the rat kidneys [106,107].
Moreover, quercetin suppressed the CXCR4 upregulation in otitis media induced by non-typificable gramnegative bacillus Haemophilus influenzae (NTHi), both in vitro and in vivo [102]. It downregulated the TLR3/ Myd88 signaling by blocking NF-κB activation [108] and IKKα and p38 phosphorylation in mice inoculated with NTHi [102]. A recent study showed that isoquercetin treatment reversed weight loss, decreased glucose levels, and increased insulin production in streptozotocin-induced diabetic rats. Furthermore, isoquercetin reduced lipid peroxidation by increasing SOD and CAT activity, which improved the lipid profile in the diabetic group. In addition, this flavonol suppressed inflammatory genes that code for NF-κB, IL-1, IL-6, TLR, TNF-α, and COX-2 [109].
Kaempferol (from genus Kaempferia) is a flavonol abundantly present in various species and plant-derived foods; it has been shown to inhibit adipogenesis by blocking 3 T3-L1 differentiation and lipid accumulation in mature adipocytes. It modulated adipogenic gene expression and decreased lipid accumulation in mature adipocytes (CEBPA) via upregulating mRNA expression of Pnpla2 and Lipe [110]. Similarly, kaempferol treatment reduced lipid and protein accumulation along with the expression of the adipogenic markers PPARγ and C/EBPα in 3 T3-L1 cells [108]. These changes in adipocyte lipid metabolism affect the response of adipocytes to the production of inflammatory mediators.
First isolated from the bark of Myrica nagi, myricetin is a common flavonol occurring also in both free or glycosylated forms, with anti-inflammatory activity and cardioprotective effect, which prevents myocarditis by reducing apoptosis [111]. It markedly reduced cleaved caspase-3 and Bcl-2 levels and increased Bax levels in H9c2 cells. The flavonol decreased MCP-1 and IκBα phosphorylation levels by blocking the NF-κB activation in LPS-challenged H9c2 cells. Treatment with semisynthetic flavone derived from myricetin (the M10 compound) prevented chronic inflammation and ulcerative colitis induced by azoxymethane/dextran in colorectal tumorigenesis [24].

Flavones and Inflammation.
Flavones are characterized by the absence of a hydroxyl group at C3 when compared to flavonol that also exhibit structural variations of hydroxylation, O-methylation, C-methylation, or prenylation. This wide range of structural combinations results in several types of flavones, making them as an abundant subclass [112]. Herein, we discussed the most frequent flavones present in human diet, luteolin, and apigenin. Luteolin modulates the immune response more efficiently than other naturally derived compounds such as quercetin, genistein, and hesperidin [113] (Table 2). Besides, apigenin is also known for its antiinflammatory and antioxidant effects by inhibiting cytokine secretion and downregulating enzymes such as inducible nitric oxide synthase (iNOS) and COX-2 [114].  Oxidative Medicine and Cellular Longevity  Luteolin was first isolated in 1829 as the main yellowcolored compound extracted from Reseda luteola and structure confirmed by synthesis in 1900 [115]. Luteolin has antimicrobial, anti-inflammatory, chemopreventive, chemotherapeutic, cardioprotective, antidiabetic, neuroprotective, and antiallergic properties [116][117][118][119]. Moreover, consumption of luteolin-rich food reduces the risk of developing chronic diseases [120].
In addition, luteolin showed a cardioprotective effect in cardiomyocyte cultures and experimental type 1 diabetes. This flavone reduced inflammation by inhibiting the NF-κB signaling, cytokine production, and oxidative stress by blocking the activation of the nuclear factor 2 (NRF2) pathway. It has also been shown to prevent cardiac fibrosis, hypertrophy, and dysfunction in diabetic rats [122]. Moreover, luteolin inhibited the production of IL-8 and COX-2 and reduced the expression of iNOS and NO in colon HT-29 cells [123]. Hence, it has emerged as a potential candidate for the treatment of bowel disease because of its ability to modulate intracellular inflammatory signaling cascade. Inhibition of the JAK/STAT signaling pathway seems to be a key mechanism for anti-inflammatory action of luteolin in bowel diseases [124]. These results indicate that luteolin negatively modulates the main signaling pathways involved in the intestinal inflammation.
Luteolin inhibited the expression of TNF-α, IL-1β, and IL-6; suppressed IκBα and NF-κB p65 phosphorylation levels; and downregulated the expression of MMP-2 and MMP-9 in S. aureus-induced mastitis [125], showing its potential in reducing tissue damage and inflammation caused by S. aureus-induced mastitis. Moreover, luteolin showed effects in arthritis models through the involvement of metalloproteinase [113]. Luteolin plays a critical role as an anti-inflammatory agent by reducing the production of inflammatory mediators, inhibiting crucial inflammatory intracellular pathways, and blocking the translocation and activation of inflammatory transcription factors. Therefore, it is a potential agent for the treatment and prevention of inflammatory, infectious, and metabolic diseases.
Apigenin, a constituent from celery (Apium graveolens) also common in other food plants, such as parsley and chamomile [49], has therapeutic potential for various diseases (diabetes, cancer, and neurological and cardiovascular disorders) [126], considering its anti-inflammatory, antioxidant, and anticancer effects. The mechanism of action of apigenin involved the suppression of p65 phosphorylation in monocytes and macrophages and reduction in the expression of COX-2 and NO in rat macrophages [8]. Zhang et al. have reported that apigenin inhibited CD38, an inflammatory biomarker, reduced pro-inflammatory cytokines, and led to reduction in adipose tissue mass in a metabolic syndrome model [127]. Treatment with apigenin prevented pulmonary fibrosis and bleomycin-induced inflammation by decreasing TNF-α and TGF-β1 levels and attenuating the reduction in SOD activity [128]. An in vivo experiment on concanavalin A-stimulated rat splenocytes showed that apigenin prevented iNOS induction, downregulated COX-2 expression, and reduced pro-inflammatory cytokine production (TNF-α, IFN-γ, and IL-2). The flavone has also demonstrated an antiproliferative effect on lymphocytes [114].

Oxidative Medicine and Cellular Longevity
A recent study reviewed preclinical and clinical data on anti-inflammatory and analgesic properties of flavonoids; most data demonstrated the potential therapeutic role and efficacy of flavonoids in cardiovascular diseases, osteoarthritis, Parkinson's disease, colitis, cancer pain, arthritis, and neuropathic pain. Nevertheless, the mechanisms of action of flavonoids have yet to be fully elucidated. Considering the pharmacological activities of flavonoids, it would be promising to further develop delivery formulations containing flavonoids to treat inflammatory diseases and pain [139].
Different approaches to evaluate the anti-inflammatory activity of flavones described above, both in vitro and in vivo, showed a reduction in the production of pro-inflammatory mediator, and inhibition of activation of the signaling pathway and nuclear factor. Regardless of the chosen model of arthritis, colitis, or gastric ulcer, flavones inhibited different signaling pathways, production of mediators, transcription factor activation, and ROS production.

Antioxidant Effects of Flavonols and Flavones.
The balance between ROS production and antioxidant cell mechanisms regulates oxidative stress in cells and can lead to cell damage when unregulated. Antioxidant enzymes, SOD and CAT, can directly detoxify the generated ROS or facilitate antioxidant activity using GSH as a reducing agent [143]. CAT is responsible for peroxide decomposition in a cell [144].
The antioxidant activity of quercetin can be explained by its direct effect on GSH, signal transduction pathways, enzymatic activity, and ROS, which maintain oxidative balance. Quercetin regulates the levels of GSH, a hydrogen donor in decomposition of H 2 O 2 to H 2 O, by inducing its synthesis. By influencing the signal transduction pathways, the compound can inhibit the enzymatic activity of numerous enzymes associated with oxidative mechanisms, such as acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), and enhance the expression levels of antioxidant enzymes such as SOD, CAT, and GSH peroxidase. Finally, quercetin can remove ROS through various mechanisms involved in regulation of ROS production and modulation of antioxidant-related gene expression. Clinically, quercetin exhibits antitumor effect and is used for cancer and metastasis prevention [145]. It has also been shown to have therapeutic effects in cardiovascular diseases and significant antidepressant outcome [146]. Quercetin isolated from Flaveria bidentis (L.) inhibited ROS production in human mononuclear cells and polymorphic leukocytes more efficiently than that of vitamin C [147]. Quercetin reversed the oxidative damage induced by FeSO 4 ferrous ions in bovine sperm cells [148]. The flavonol also induced the expression of HO-1 and the subsequent end product, suppressing oxidative stress derived from NADPH oxidase [142]. Furthermore, quercetin decreased MDA levels in the myocardial tissue and activated SOD and GPx enzymes [98,142], demonstrating the antioxidant activity of quercetin in a rat model of myocardial infarction. In a similar study, treatment with quercetin decreased MDA levels in the myocardial tissue and further stimulated SOD and GPx enzymes [149].
Luteolin attenuated oxidative damage and reduced the generation of intracellular ROS by inducing the expression of the antioxidant enzyme HO-1, thus counteracting the damage induced by H 2 O 2 in rat cortical primary cell cultures [141]. Treatment with the flavone normalized AST and ALT levels. The protective role of luteolin against ROS and oxidative damage occurs by normal GSH level restoration and lipoperoxidation suppression in the plasma and liver tissues [140]. Moreover, the flavone blocked superoxide anion and ROS production in human neutrophils [150], strengthening the role of luteolin as an antioxidant agent. Chanput et al. evaluated the antioxidant activity of three classes of flavonoids: (i) flavonol: quercetin and myricetin; (ii) flavanone: eriodictyol and naringenin; and (iii) flavone: luteolin and apigenin [151]. The flavonol group (myricetin and quercetin) showed the best antioxidant activity among the three groups, which might be due to the presence of more hydroxyl groups in their structure and the presence of double bond between C2 and C3 of the aromatic ring that improve their antioxidant properties by the electron distribution pattern [152] compared to the other groups.

Antibacterial Activity of Flavones and Flavonols.
Several studies have supported the use of medicinal plants for their antimicrobial activities. Flavonoids act on different microorganisms. Herein, we discuss their effects on resistant and opportunistic bacteria. Antibiotic abuse may lead to the select bacterial resistance to current drugs. Medicinal plant extracts contain a variety of phytochemical substances that can act synergistically to inhibit the bacterial growth and prevent the development of drug resistance. Table 3 shows the flavonoids used for antibacterial purposes and their mechanisms of action.
The methanolic root extract from Garcinia macrophylla is an important medicinal herb used to treat diverse diseases and disorders in Traditional Chinese Medicine and showed antimicrobial activity against bacterial strains of Micrococcus luteus, E. faecalis, S. aureus, Staphylococcus epidermidis, Streptococcus uberis, P. aeruginosa, E. coli, and K. pneumoniae isolated from burn wounds, with the lowest minimum inhibitory concentration against S. epidermidis and the highest MIC value against E. coli [150]. The authors observed equivalent values for minimum bactericidal concentration (MBC). The antibacterial activity of Gentiana kurroo methanolic extract overcame the results presented by other comparable studies [155].  (i)Inhibition of IL-8 production, and COX-2 and iNOS expression (ii)Downregulation of the JAK/STAT signaling pathway [123] In vitro Primary cultures of rat cortical cells subjected to oxidative stress 3-30 μM (i)Attenuation of oxidative damage (ii)Inhibition of ROS generation (iii)Increase in bad phosphorylation in serine-112 (iv)Attenuation of induced caspase-3 activation (v)Upregulation of HO-1 expression (vi)Increase in the levels of ERK, p38, JNK, and AKT phosphorylation [141] 11 Oxidative Medicine and Cellular Longevity (i)Prevention of pulmonary fibrosis (ii)Prevention of bleomycin-induced inflammation by decreasing TNF-α and TGF-β1 levels (iii)Attenuation of reduced SOD activity (iv)Inhibition of leukocyte infiltration [128] 3′,4′ -dihydroxyflavone In vitro Murine BV2 microglial cells 0-10 μM (i)Inhibition of the production of NO and PGE2 chemokines (ii)Inhibition of pro-inflammatory cytokines such as TNFα, IL-1β, and IL-6 (iii)Inhibition of MAPK phosphorylation (iv)Inhibition of the NF-κB pathway [129] In vivo Male C57BL/6 mice 5 Mg/kg In vivo Colitis TNBS-induced (i)Showed protective effect by suppression of TNF-α, IL-6 and NF-κB [132] 12 Oxidative Medicine and Cellular Longevity The methanolic extract of B. purpurascens leaves, a medicinal herb containing diverse flavonoids (bergenin, catechin, naringenin, and myricetin), is used to treat wounds and bacterial infections. The extract exhibited strong antibacterial properties, particularly against S. aureus and Streptococcus spp. These results corroborate those of previous studies demonstrating the inhibition of growth of different bacteria by Bergenia spp. [156,157] and that silver-plated nanoparticles from Bergenia cilita crude extract had antimicrobial activity against several bacterial strains. Moreover, B. purpurascens extract reduced the mortality caused by infection of mice with S. aureus in vivo [158,159].
Microorganisms' growth and survival are associated with biofilm formation. Quorum sensing (QS) is an intercellular signaling or cell-cell communication process that is necessary for coordinating biofilm formation. The QS system is related to opportunism and resistance mechanisms developed by many bacterial strains. Blocking QS in bacteria is a far-reaching strategy for attenuating their virulence and making them more susceptible to elimination by the host immune system at lower doses of antibiotics. The modulatory effect of kaempferol-loaded chitosan nanoparticles on autoinducer (AI)-mediated anti-QS activity by Chromobacterium violaceum CV026 was used to evaluate the effect of this flavonol. Changes in bacterial cell density led to AI production via signaling molecules involved in communication among bacteria, composing QS, and managing growth. The results of anti-QS evaluation using the disk diffusion method showed that kaempferol produces a remarkable anti-QS effect; thus, its use against resistant bacteria is promising [158,159].
Baicalin, a flavonol extracted from the Chinese medicinal plant Scutellaria baicalensis, has also demonstrated in vitro and in vivo anti-QS activities [160]. Baicalin concentrations below the MIC (sub-MIC) inhibited Pseudomonas biofilm formation, which increased the bactericidal effects of various conventional antibiotics. MIC and broth microdilution methods were used to evaluate the MIC and MBC of baicalin and antibiotics, biofilm inhibition assay, biofilm dispersion evaluation, and combined antibiotic  13 Oxidative Medicine and Cellular Longevity use. Treatment with different concentrations of baicalin inhibited P. aeruginosa biofilm formation in a dosedependent manner. The lower number of viable cells and lower biomass indicated a synergistic effect compared to treatment with flavonol alone and antibiotics [161], demonstrating that baicalin co-treatment increased the antimicrobial activities of three different antibiotics.
Treatment with sub-MIC of baicalein reduced QS. It decreased most virulence factors, corroborating previously reported data and demonstrating that baicalin exerts anti-QS activity against P. aeruginosa PAO1 by inhibiting the QS gene expression and production of signaling molecules such as acyl-homoserine lactone (AHL). Treatment with a sub-MIC of baicalin also considerably increased the survival rate of C. elegans. Moreover, baicalin inhibited the motility and production of the QS molecule pyocyanin and significantly reduced the number of colony-forming units (CFUs) of P. aeruginosa in implants [160]. Previous studies have also demonstrated reduced bacterial growth in implants treated with baicalin and antibiotics, particularly levofloxacin, ampicillin/clavulanic acid, and ceftazidime [162]). In vivo experiments have shown a higher survival rate of worms in the baicalin-treated group, which indicates that treatment with baicalin reduced the pathogenicity of P. aeruginosa in C. elegans. In addition, baicalin treatment increased the clearance of P. aeruginosa from the implants of infected animals in a peritoneal implant infection model.
Quercetin, both free and chitosan-associated, and baicalein have shown a potential inhibitory effect on biofilm formation, QS formation, and cytotoxicity in free mammalian cells treated with chitosan-based nanocapsules [163]. A transformed E. coli Top 10 biosensor strain showed QS formation. These results are in line with the data of a previous study [164] that suggested that these phytochemicals may inhibit biofilm formation [163]. Indeed, baicalin inhibited P. aeruginosa PAO1 biofilm formation, whereas quercetin showed the highest anti-QS effect in PAO1 strain [165].
The flavonol rutin inhibited QS, biofilm formation, and virulence genes in pathogenic E. coli APEC-O78 in vitro. Bioluminescence assay demonstrated that AI-2 secretion and activity were significantly decreased after rutin treatment, resulting in marked inhibition of biofilm formation. The expression of APEC virulence genes (csgA, csgB, Flic, FyuA, IucD, LsrB, LsrK, Rpos, H-NS, Luxs, and Pf) decreased after treatment with rutin. These results highlight the role of rutin in controlling E. coli pathogenicity. Furthermore, flavonols reduced cellular damage in type II pneumocytes infected with pathogenic E. coli by interfering with the QS, thereby inducing a decrease in AI-2 production, biofilm formation, and virulence gene expression [162].
The efflux pump proteins encoded by genes involved in bacterial cellular functions may cause antibiotic resistance. Crude extracts and 3 ′ ,4 ′ ,7-trihydroxyflavone compound extracted from Myristica fragrans Houtt. (MSF) have shown in vitro antibacterial and antibiotic sensitizing activity against E. coli AG102, which is a multidrug-resistant (MDR) gram-negative enteric bacterium, and against P. stuartii strains. This study observed a synergistic effect of these substances with a PAβN efflux pump inhibitor. The antibacterial potential was improved by 73.37% with crude MFS against the tested MDR, whereas it was improved by 100% with 3′,4′,7-trihydroxyflavone. Therefore, both substances are considered substrates for efflux pumps because they have intracellular targets [166]. There was a synergistic action between the extracts and at least one of the six antibiotics tested against 50% of MDR bacterial strains. The compound 3 ′ ,4 ′ ,7-trihydroxyflavone improved the activity of 70% of the antibiotics tested. These results suggest that this  For example, pathway 1 is blocked by the following compounds: quercetin, luteolin, 3′,4′-dihydroxyflavone, baicalein, nobiletin, and tangeretin. Scutellarin, vicenin-2, apigenin, 5-O-methyl-scutellarin, and the substances are able to block pathway 1 and inhibit pathway 2. Only luteolin and lucenin-2 inhibit pathway 3. Furthermore, pathway 4 can only be inhibited by luteolin. Most of the compounds previously mentioned in this review block pathway 5, except nobiletin, tangeretin, vicenin-2, and lucenin-2. Furthermore, luteolin, scutellarin, vicenin-2, and lucenin-2 inhibited pathway 6. Only isoquercetin blocks pathway 7, and luteolin blocks pathway 8.

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Oxidative Medicine and Cellular Longevity compound may be considered as a potential efflux pump inhibitor because of its antibacterial effect and antibioticmodifying activity [166]. The effects of the hydroalcoholic extract of Glycyrrhiza glabra and its flavonoids were evaluated in an in vivo model of P. aeruginosa-induced pulmonary infection in rats. The study elucidated the mechanism of action, inhibition of biofilm formation, pathogenesis, and efflux activity of one of its pure glycyrrhizic acid components [167]. The findings were validated using molecular docking analysis using ExoS segmentation, which confirmed the potential of the herb G. glabra [153]. Isolated G. glabra glycosides altered membrane permeability and inhibited efflux activity and biofilm formation by P. aeruginosa [166]. These flavonoids isolated from licorice showed inhibitory effects on acute lipopolysaccharide-induced pneumonia [167]. These studies provide evidence of preclinical efficacy of G. glabra in P. aeruginosa pneumonia.
In addition to interfering with the QS and efflux pump, antimicrobials act by direct killing of microbial cells. Antibacterial activity was evaluated by MIC in DNA fragmentation test, caspase activity, lipid peroxidation, inner and external membrane permeabilization, membrane fluidity disturbance, and membrane damage. Isoquercitrin isolated from the perennial herb Aster yomena, generally found in Korea and Japan, inhibited E. coli growth or completely destroyed bacterial cells. Sub-MIC concentrations of isoquercitrin led to a rapid increase in DNA fragmentation and caspase activation, which are characteristics of apoptosis. The antibacterial activity of isoquercitrin in E. coli is attributed to membrane alterations, most likely caused by the induction of oxidative stress and apoptosis [167]. A double-blind, placebo-controlled clinical trial showed that propolis oral spray could be used to improve and immediately resolve the symptoms of uncomplicated upper respiratory tract bacterial infections, without any pharmacological treatment [168].
The antibacterial actions of flavonoids and flavonols range from QS disturbance and alterations in the efflux pump to direct induction of apoptosis (Table 4). Unfortunately, there are few studies detailing the mechanisms involved, but their microbicidal and synergistic effects with antibiotics should be further explored in the future.

Concluding Remarks
Conclusively, flavonoids are bioactive components found in fruits and vegetables being recognized as highly versatile substances with therapeutic applications, representing a vast source of pharmacological potential. Herein, we discuss the pivotal role of some flavonoids against inflammation and bacterial infection, which are responsible for millions of casualties every year and severely affect hospitalized and susceptible individuals. Currently available drugs used to treat of these individuals will probably no longer be effective shortly. Bioactive compounds derived from plants play a critical role in treating or preventing diseases either individually or synergistically with antibiotics. Furthermore, flavonoids play a critical role in modulating oxidative stress and treating inflammatory and infectious diseases, particularly those caused by resistant and opportunistic bacteria, through diverse mechanisms of action. Despite their low bioavailability, a proper delivery system such as nanoformulation may aid in their clinical applications. Nanotechnology offers multiple benefits in treating chronic human diseases by site-specific and target-oriented delivery of precise medicines, improving the efficacy of drugs and natural products [169,170]. For instance, phytochemical cyclodextrin complexes have the potential for transformation into drug delivery systems [171], and nanotechnology-based drug delivery for therapeutics are bioactive compounds from plant sources [172]. Besides, plant phenolic compounds have potential use as a natural feed additive [173], enhancing antioxidant status, immune function, and growth performance of farm animals [174]. We focused on the reported mechanisms and intracellular targets of flavonols and flavones ( Figure 6) to boost research on discovering bioactive compounds and their mechanisms of action, anticipating that they can be used as standard treatment for inflammatory and infectious diseases in the future.

Conflicts of Interest
The authors declare no conflicts of interest regarding the publication of this paper.

Authors' Contributions
Adriana R. Silva and Cassiano F. Gonçalves-de-Albuquerque contributed equally to this work.