Veronica Plants—Drifting from Farm to Traditional Healing, Food Application, and Phytopharmacology

The Veronica genus, with more than 200 species, belongs to the Plantaginaceae family and is distributed over most of the Northern Hemisphere and in many parts of Southern Hemisphere. These plants are traditionally used in medicine for wound healing, in the treatment of rheumatism, and in different human diseases. This paper reviews the chemical composition of some valuable Veronica species, the possibilities Veronica extracts have in food preservation and as food ingredients, and their functional properties. Veronica species represent a valuable source of biological active secondary metabolites, including iridoid glycosides and phenolic compounds. In particular, due to presence of these phytochemicals, Veronica species exhibit a wide spectrum of biological activities, including antimicrobial and antioxidant. In fact, some studies suggest that some Veronica extracts can inhibit foodborne pathogens, such as Listeria monocytogenes, but only a few of them were performed in food systems. Moreover, anticancer, anti-inflammatory, and other bioactivities were reported in vitro and in vivo. The bioactivity of Veronica plants was demonstrated, but further studies in food systems and in humans are required.


Introduction
The genus Veronica at present belongs to the family Plantaginaceae, while it was previously classified in the family Scrophulariaceae [1]. There are many suggestions (problems) related to the classification and rearrangement of this genus [2,3]. The family includes 120 plant genera with 7055 scientific plant names, of which 1614 are accepted species names [4]. The Plant List includes 1520 scientific plant names for the Veronica genus. Of these, 234 are accepted species names, 335 are synonyms, and 951 are unassessed. Thus, the total number of species belonging to the genus Veronica depends on synonym acceptance. These species are distributed over the Northern Hemisphere and into the Australasian region (Australia, New Zealand, New Guinea), with centers of diversity in western Asia and New Zealand [2]. Most of the species of Veronica occur in regions with a Mediterranean precipitation regime from the sea to high alpine elevations. Despite the importance in many habitats, aquatic plants of Veronica are mostly researched in modern biosystematic studies. The common member of the semi-aquatic plants in the Mediterranean region is Veronica sect. beccabunga [5]. Some other common species of Veronica genus are represented in Table 1. Traditional medicine is a sum of great knowledge about health, disease prevention, treatment, and physical and mental illnesses. There are different types of traditional medicine such as ancient Iranian medicine, traditional Chinese medicine, Ayurveda, traditional African medicine, acupuncture, and so on [7]. In 2019, the World Health Organization published a global report on traditional and complementary medicine and stated that 88% of its member states, corresponding to 170 member states, reported that their population uses traditional medicines to treat illnesses [8]. The wide diverse distribution of Veronica plants, from aquatic to dry steppe habitats and from sea-level to high alpine regions [9], could be related to the wide range of traditional uses within these cultures ( Table 1). As an example, Veronica peregrina L. is useful for treating hemorrhage, gastric ulcer, infections, and diseases related to macrophage-mediated inflammatory disorders, as illustrated in Korean traditional medicine [10]. Tea made from Veronica spicata L. is a well-known remedy in traditional medicine [11]. Veronica species have cytotoxic and anti-inflammatory activity. Veronica officinalis L. (common speedwell) is used for treating liver, eczema, ulceration, snake bites, wound healing, and skin lesions in Balkan traditional medicine [12]. Veronica species are used in traditional medicine for the treatment of rheumatism [13], hemoptysis, laryngopharyngitis, hernia [14], and lung and respiratory diseases (e.g., against cough or as an expectorant) [15]. They also have properties such as antiscorbutic and diuretic, as well as wound healing [16]. Three Veronica species, namely, V. officinalis, Veronica chamaedrys L., and Veronica herba DAC, are used in traditional Austrian herbal drugs [17]. V. officinalis is a popular medicinal plant, used as a commercial herbal product in many European countries [18]. There are around 79 Veronica species in Turkish flora, 26 of which are endemic. Different parts of Veronica species are used as a diuretic, for wound healing, and against rheumatic pains in Turkish folk medicine [19].
Thus, based on this knowledge, Veronica plants are potential sources of nutraceuticals and functional ingredients with a wide spectrum of bioactivities. In fact, Veronica species represent a valuable source of biological active compounds. Among others, the extracts of Veronica plants show antioxidant, antimicrobial, antifungal, anti-inflammatory, scolicidal, and anti-cancer activities, as well as inhibitory potential on acetylcholinesterase, tyrosinase, lipoxygenase, and xanthine oxidase [20]. Overall, these species might be considered good candidates for industrial or pharmacological applications. Therefore, this review covers information about the phytochemical composition of Veronica plants, mainly focused on phenolics and iridoids. Their biological properties are also detailed, as well as recent food applications.

Veronica Filiformis
A non-common flavone glycoside was isolated from the whole plant of V. filiformis and identified by means of 13 C NMR spectroscopy as isoscutellarein 4′-methyl ether 7-O-β-(6′′′-O-acetyl-2′′-O-allosylglucoside) (16) (Figure 3). This was the first report of 2-allosylglucose as a disaccharide unit of flavonoids [26]. Furthermore, the structure of other particular compounds in Veronica species is detailed in this section; in addition to the expected iridoid glucosides, Veronica species are sources of new phytochemicals.

Veronica linariifolia Pall. ex Link
A new flavonoid glycoside, linariifolioside (18) (Figure 3), was isolated from the alcohol extract of the dried whole herb.

Veronica Arvensis
The water extracts of this plant revealed the presence of cornoside aglycone and rengyolone (62). The iridoid glucoside, ajugol (63), and the phenylethanoid glucoside, cornoside (64) (Figure 11), were isolated from species of Veronica for the first time [39]. Other compounds are described in Table  2.

Veronica arvensis
The water extracts of this plant revealed the presence of cornoside aglycone and rengyolone (62). The iridoid glucoside, ajugol (63), and the phenylethanoid glucoside, cornoside (64) (Figure 11), were isolated from species of Veronica for the first time [39]. Other compounds are described in Table 2.

Veronica Derwentiana Andrews and Veronica Catarractae G. Forst.
Three unusual substituted benzoyl esters of aucubin were obtained from V. derwentiana and a chlorinated iridoid glycoside (catarractoside) (78) (Figure 13) from V. catarractae, in addition to other iridoids common to the genus. The chemical profile of V. perfoliata is similar to that of Northern Hemisphere species of Veronica because of the presence of characteristic 6-O-catalpol esters. The profile of V. derwentiana is unique, since 6-O-esters of aucubin rather than of catalpol dominate; however, the acyl groups are the same as those present in catalpol esters found in some other Veronica sections. V. catarractae also contains one of the catalpol esters characteristic of Veronica, in addition to three 6-O-rhamnopyranosyl substituted iridoid glycosides, one of which is 6-O-rhamnopyranosylcatalpol (79) (Figure 13) [41].

Veronica derwentiana Andrews and Veronica catarractae G. Forst.
Three unusual substituted benzoyl esters of aucubin were obtained from V. derwentiana and a chlorinated iridoid glycoside (catarractoside) (78) (Figure 13) from V. catarractae, in addition to other iridoids common to the genus. The chemical profile of V. perfoliata is similar to that of Northern Hemisphere species of Veronica because of the presence of characteristic 6-O-catalpol esters. The profile of V. derwentiana is unique, since 6-O-esters of aucubin rather than of catalpol dominate; however, the acyl groups are the same as those present in catalpol esters found in some other Veronica sections. V. catarractae also contains one of the catalpol esters characteristic of Veronica, in addition to three 6-O-rhamnopyranosyl substituted iridoid glycosides, one of which is 6-O-rhamnopyranosylcatalpol (79) (Figure 13) [41].

Veronica Rosea Desf.
The phytochemical study of butanolic extract of aerial parts of V. rosea led to the isolation and identification of four phytoconstituents: apigenin-7-O-β-glucopyranoside (114) (Figure 16

Antibacterial Activity
Although Veronica species are traditionally used for their antibacterial effect, there are only a few studies that showed its strong antibacterial effect. The antibacterial activity against Gram-positive and Gram-negative species depends on the extract type (solvent used, part extracted, species, etc.). As an example, the antimicrobial properties of aerial parts of V. spicata extracts with ethyl-acetate, methanol, or water using the diffusion method and microdilution method were examined. The bacterial strains used in study were found to be susceptible toward methanol and ethyl-acetate extracts, with minimum inhibitory concentration (MIC) values between 1.25 and 5 mg/mL using the microdilution method, while aqueous extracts were inactive. The extracts prepared from leaves showed larger zones of inhibition compared to flowers and steam extracts of V. spicata, indicating stronger antimicrobial activity [11,58]. In another work and using the microdilution method, Živković et al. (2014) investigated the antibacterial effect of the V. urticifolia methanol extract against the Gram-negative bacteria Escherichia coli, Enterococcus faecalis, and Pseudomonas aeruginosa, and Gram-positive bacteria Staphylococcus aureus, L. monocytogenes, and Bacillus cereus [59]. After the measurement of the minimum bactericidal concentration (MBC) and MIC values, it was found that the most sensitive germ was Staphylococcus aureus. The antistaphylococcal effect is due to a main phenolic compound acteoside, which could inhibit the incorporation of leucine and disturb protein synthesis. The same antistaphylococcal effects were found in other studies [20]. Other chemical compounds could be also responsible for this antibacterial activity, e.g., β-sitosterol, campesterol,

Antimicrobial Activities of Veronica Plants
Many plants were used since ancient times to treat infections caused by bacteria that are now resistant to antibiotics [56]. There are several traditional uses of the genus Veronica that could be related to antimicrobial properties. For example, these species are used as expectorants, restoratives, tonics, and for the treatment of influenza and other respiratory diseases in traditional Chinese medicine [57]. In Romanian medicine, aerial parts of Veronica plants are known for their wound-healing properties and are also used for the treatment of cough and catarrh [47]. Despite their widespread usage in folk medicine, there is a lack of information about the antimicrobial activity. Only a few studies confirmed that certain Veronica species showed noticeable bioactivity such as antibacterial, antifungal, and antiviral activity.

Antibacterial Activity
Although Veronica species are traditionally used for their antibacterial effect, there are only a few studies that showed its strong antibacterial effect. The antibacterial activity against Gram-positive and Gram-negative species depends on the extract type (solvent used, part extracted, species, etc.). As an example, the antimicrobial properties of aerial parts of V. spicata extracts with ethyl-acetate, methanol, or water using the diffusion method and microdilution method were examined. The bacterial strains used in study were found to be susceptible toward methanol and ethyl-acetate extracts, with minimum inhibitory concentration (MIC) values between 1.25 and 5 mg/mL using the microdilution method, while aqueous extracts were inactive. The extracts prepared from leaves showed larger zones of inhibition compared to flowers and steam extracts of V. spicata, indicating stronger antimicrobial activity [11,58]. In another work and using the microdilution method, Živković et al. (2014) investigated the antibacterial effect of the V. urticifolia methanol extract against the Gram-negative bacteria Escherichia coli, Enterococcus faecalis, and Pseudomonas aeruginosa, and Gram-positive bacteria Staphylococcus aureus, L. monocytogenes, and Bacillus cereus [59]. After the measurement of the minimum bactericidal concentration (MBC) and MIC values, it was found that the most sensitive germ was Staphylococcus aureus. The antistaphylococcal effect is due to a main phenolic compound acteoside, which could inhibit the incorporation of leucine and disturb protein synthesis. The same antistaphylococcal effects were found in other studies [20]. Other chemical compounds could be also responsible for this antibacterial activity, e.g., β-sitosterol, campesterol, stigmasterol, hispidulin, and flavonoids. These results are important for human health because S. aureus is a pathogen germ difficult to treat with the development of antibiotic resistance. For this reason, natural alternative therapies to solve this problem are useful. Other studies showed the antibacterial activity of methanol, ethanol, or aqueous extracts from Veronica urticifolia Jacq., Veronica orchidea Crantz, V. persica, and Veronica montana L. against Gram-positive and Gram-negative bacteria [20,47,57,59]. The summary of the antimicrobial activity of different Veronica species is represented in Table 3.
As commented before, not all results were positive. Dulger and Ugurlu (2005)  , and Kluyveromyces fragilis (ATCC 8608) using the disc diffusion method. In this case, V. lycica had weak antimicrobial effect against the tested microorganisms [60]. V. anagallis-aquatica was tested using the agar well diffusion assay against five bacterial and two yeast strains. None of the extracts of this Veronica species showed significant inhibition comparing to the positive control (gentamicin) [61].   [20] In some cases, the antimicrobial activity of the isolated compounds was determined. In a study conducted by Mocan and collaborators, V. officinalis, V. teucrium, and V. orchidea were selected from Romanian natural flora and investigated for their antioxidant and antimicrobial effects against anaerobic bacterial strains with emphasis on the isolated compounds, caffeic and chlorogenic acids. V. teucrium and V. orchidea presented a higher activity (MIC = 31.25 mg/mL and MBC = 62.5 mg/mL) than V. officinalis (MIC and MBC of 62.5 mg/mL), with the most sensitive strain being Peptostreptococcus anaerobius. All analyzed species contained both caffeic and chlorogenic acids, where the richest source of caffeic acid was V. officinalis and the highest amount of chlorogenic acid was found in V. teucrium [62].

Antifungal, Antiviral, and Antiparasitic Activity
Other studies demonstrated the antifungal effect of Veronica species [11,20,47]. Mocan and co-workers investigated the antifungal properties of V. persica against Aspergillus niger and Penicillium hirsutum using the Kirby-Bauer diffusimetric method. The antifungal effect was higher for A. niger than P. hirsutum and it is due to phenolic compounds [47]. Using the same method, antifungal effects of V. persica extract against Candida albicans and A. niger were demonstrated in another study [20]. The results showed that the highest antifungal effect for both fungal pathogens was obtained at a concentration 300 µg/mL of extract. Dunkic et al. demonstrated in another study the antifungal effect of methanol V. spicata extract at MIC values ranging from 1.25 mg/mL to 5 mg/mL. In addition, the methanol extract prepared only from V. spicata's leaves also had an effect against the dermatophyte Microsporum gypseum [11]. As a result of these positive data, Veronica plants can be considered good natural therapeutic alternatives for mild fungal infections.
A new biological property, i.e., antiviral activity (against herpes simplex viruses HSV1 and HSV2), of V. persica was demonstrated in a recent research conducted by Sharifi-Rad et al. (2018) [63]. In this study, the ethanol extract of V. persica was tested on Vero cells infected with both types of viruses. The stronger antiviral activity was found in the 80% methanol fraction of V. persica extract during and after infection of the cells with viruses, thus suggesting the interference of the extract with the intracellular entry of the virus and a possible inhibition of the viral intracellular replication of endogenous herpetic viruses. HSV1 was much more sensitive to the action of the methanolic fraction compared to HSV2. In addition, the 80% MeOH fraction of V. persica extract administered to the cells at the same time with aciclovir (antiviral drug) showed a synergistic effect in reducing plaque formation by herpetic viruses [63]. This antiviral action indicates the usefulness of the V. persica extract in combination with the antiviral medication (such as aciclovir) for decreasing the severity of symptomatic episodes of oral herpetic infection, which relapses when the immune system is weak. Moreover, in a recent investigation conducted by these authors, in vitro and in vivo susceptibility of Leishmania major to V. persica extract was evaluated. Antileishmanial activity of plant extract was investigated on cultured L. major promastigotes and in mice. In vitro tests showed a high and dose-dependent inhibitory activity of plant extract, which was able to reduce the survival time of promastigotes in a concentration-dependent manner; for example, the survival time of promastigotes decreased to 10% at 750 µg/mL after 72 h of exposure time. There was a significant influence of V. persica extracts in vivo on accelerating the healing process, as well as reducing the overall disease burden, in an animal model by inducing nitric oxide production in macrophage cells [64].
Antioxidant activities of Veronica species could be attributed to their content of iridoids and phenolic acids [65]. Moreover, acylated flavonoids and phenol glycosides from V. thymoides subsp. pseudocinerea exhibited potent radical scavenging activity against DPPH radicals [38]. The two phenylethanoid glycosides, turrilliosides A and B, isolated from Veronica turrilliana Stoj. & Stef. were found to be potent DPPH radical scavengers, approximately 1.6-fold better than the flavonoid quercetin [40]. In another work, the antioxidant capacity of V. persica phenolic-rich extracts was correlated with their total phenol content [56].

In Vivo Studies
The antioxidant activity of three Veronica species, V. teucrium, V. jacquinii, and V. urticifola, was examined in vivo in rats, and their effect on several hepatic antioxidant systems was tested, i.e., on the activity of glutathione peroxidase, glutathione reductase (GR), peroxidase (Px), catalase (CAT), and xanthine oxidase, as well as glutathione (GSH) content and level of thiobarbituric acid reactive substances (TBARS). Treatment with Veronica extracts (methanolic, aqueous-acetone, and water extracts) (100 mg/kg) inhibited CCl 4 -induced liver injury by decreasing TBARS level, increasing GSH content, and bringing the activities of antioxidative enzymes CAT, Px, and GR to control levels. The study suggested that the extracts analyzed could protect the liver cells from CCl 4 -induced liver damage by their antioxidative effect on hepatocytes [70]. Veronica ciliata Fisch. is also a considerable candidate for protecting liver injuries due to its antioxidant and anti-apoptosis properties [71].
The antioxidant activity of other Veronica extracts was associated with other bioactivities. As an example, Lu and co-authors reported that isolated compounds from V. ciliata showed anti-hepatocarcinoma activities against HepG2 liver hepatocellular carcinoma cells, which could be associated with its antioxidant activity [48]. Lee and co-workers evaluated the antioxidant activity, cytotoxicity, and collagen synthesis activity in vitro in order to test the anti-wrinkle effect of a formulated cream containing V. officinalis extract. Antioxidant evaluation was performed in fibroblast cells. The ethanolic extract showed good antioxidant activity against DPPH free radicals (103.50 µg/mL) and also exhibited a significant effect on collagen synthesis activity without cytotoxicity. In a placebo-controlled trial on women, the treatment with the formulated cream (Scoti-Speedwell) for 56 days significantly resulted in anti-wrinkle activity [72].

Anticancer Activities of Veronica Species
Plant-derived metabolites are beneficial sources of new anti-cancer drugs with reduced cytotoxicity and increased activity. More than 60% of today's drugs with proven anticancer properties originated from plants [73]. Extracts obtained from the aerial parts of various Veronica species are globally used as folk medicine for the therapy of cancer [16]. Despite this fact, only a few amounts of Veronica species were investigated for their cytotoxic and anticancer activities (as an example, see Table 4). With the exception of one in vivo confirmation, all of the investigations were performed in vitro against various cancer cell lines. Also, there is a lack of mechanistic studies for compounds isolated from Veronica species.

In Vitro Studies
Methanolic and water extracts of several Veronica species were tested against cancer cells in vitro. Harput and collaborators studied the cytotoxic activity of five Veronica species: V. cymbalaria, V. hederifolia, Veronica pectinata L., V. persica, and V. polita. Their methanolic extracts showed dose-dependent cytotoxicity against KB (human epidermoid carcinoma) and B16 (mouse melanoma) cells. Additional fractionation of investigated extracts pointed out that the active compounds occurred in the chloroform fraction, but not the water one. Moreover, KB cells were more sensitive to the CHCl 3 -soluble parts of the extracts compared to B16 cells. Except for V. cymbalaria, Veronica species exhibited similar activity against KB cells, while V. persica, V. polita, and V. pectinata demonstrated potent activity against B16 cells [74]. Saracoglu et al. (2011) evaluated the cytotoxicity of aqueous extracts (10-800 µL) of V. cuneifolia subsp. cuneifolia and V. cymbalaria using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay against various cell lines, Hep-2 (human epidermoid carcinoma), RD (human rhabdomyosarcoma), and L-20B (transgenic murine L-cells), as well as in non-cancerous Vero cells (African green monkey kidney cells). Both samples showed cytotoxic activity against used cell lines, but V. cuneifolia subsp. cuneifolia showed a stronger effect with IC 50 values ranging from 250.4 (for RD line) to 410.9 (for Vero cell line) µg/mL [75]. Moreover, a previous pharmacological investigation on edible V. americana (Raf.) Schwein species (American speedwell) showed that the methanolic extract of the aerial parts exhibited cytotoxic activity against colon (HF-6) and prostate (PC-3) human cancer cell lines with IC 50 values of 0.169 and 1.460 µg/mL, respectively [76].
Other studies focused on the elucidation of the active compounds. As an example, the authors of Reference [50] conducted bioassay-guided fractionation of V. americana methanolic extract, employing the cytotoxic activity against two previously mentioned cancer cell lines in order to determine active components. Compounds in this extract that demonstrated activity higher than camptothecin (used as the positive control) were 4β-hydroxy-6-O-(p-hydroxybenzoyl)-tetrahydrolinaride and 10-O-protocatechuyl catalpol. Their activity was higher against HF-6 cells with IC 50 values of 0.031 and 0.066 µM for americanoside and 10-O-protocatechuyl-catalpol, respectively. Also, according to the calculated selectivity index (SI), both compounds showed more selective cytotoxicity against applied cancer cell lines than against human normal MRC-5 (fetal lung fibroblast) cells. Yin and co-workers investigated anti-hepatocarcinoma activity of 95% ethanol extract of V. ciliata aerial parts and its fractions, using human hepatocellular carcinoma cells HepG2 and the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) test. They demonstrated that active compounds were concentrated in the ethyl acetate fraction of the extract [77]. Iridoid compounds veronicoside, catalposide, amphicoside, and verminoside strongly inhibited HepG2 cell proliferation in a dose-dependent manner. Their inhibition rates (with the exception of veronicoside) were significantly higher compared to that of 5-fluorouracil used as a positive control, i.e., IC 50 values ranged from 15.54 to 28.32 µg/mL, while the 5-fluorouracil IC 50 value was 29.62 µg/mL. In the later study conducted by the same group of authors [48], the anti-hepatocarcinoma activities of five other compounds isolated from V. ciliata were also determined in vitro against the HepG2 cell line using the Cell Counting Kit-8 (CCK-8) method. The results showed that the proliferation of HepG2 cells was notably inhibited by these five compounds in a dose-dependent manner, and their activities decreased in the following order: luteolin > verproside > catalposide > 3,4-dihydroxy benzoic acid > 4-hydroxybenzoic acid, with IC 50 values ranging from 102.36 to 444.76 µg/mL. The authors hypothesized that the exhibited activity of isolated compounds could be associated, at least in part, with their antioxidant activity. According to them, the higher cytotoxic potential of luteolin compared to analyzed iridoid glucosides and phenolic acids could be attributed to the higher number of phenolic groups in the molecule.
In a recent study, the cytotoxic properties of eight compounds isolated from V. sibirica were tested in vitro through the MTT assay against two cell lines: human neuroblastoma (SK-N-SH) and human hepatocellular carcinoma (BEL-7402). Compounds sibiriquinone A, cryptotanshinone, ferruginol, dihydrotanshinone I, tanshinone I, and tanshinone IIA showed beneficial inhibitory effects on the SK-N-SH cell growth, while compounds sibiriquinone A, cryptotanshinone, dihydrotanshinone I, and tanshinone IIA inhibited growth of the BEL-7402 cell line [42]. Moreover, the cytotoxic activity of iridoid compounds characteristic for Veronica species (aucubin, catalpol, and catalpol derivatives) was also determined against previously mentioned cell lines Hep-2, RD, and L-20B. Among tested compounds, verminoside showed very strong activity with IC 50 values of 128 µM, 70 µM, and 103 µM, respectively. The activities of amphicoside and veronicoside were lower, while verminoside, verproside, and 6-O-veratroylcatalposide showed cytostatic activity [78]. Moreover, the in vitro antitumor activity of diterpenes, chemical compounds from V. sibirica, was also shown using the SK-N-SH human neuroblastoma cell line and BEL-7402 human hepatoma cell line [42].

In Vivo Studies
Tumors and different types of cancers are difficult to treat, but classical conventional chemotherapy can be combined with cytotoxic natural remedies. Živković and co-workers evaluated the antitumor activity of V. urticifolia methanolic extract in vivo in animals with Ehrlich ascites carcinoma (EAC) [59]. It is an aggressive, fast-growing carcinoma initially described as a spontaneous murine mammary adenocarcinoma [80]. The antitumor properties of V. urticifolia methanolic extract were estimated via determination of the tumor cell count, ascites volume, and cell viability, and the results were compared with those achieved for positive control N-acetyl-l-cysteine. Pretreatment (2 mg/kg body weight) for seven days before EAC implantation showed a statistically significant decrease in tumor cell viability, while ascites volume and tumor cell count were reduced to some extent, but not statistically significant.
There are too few data about the in vivo antitumor effects of compounds isolated from Veronica species. Acteoside (phenylpropanoid compound dominant in V. urticifolia extract) applied intraperitoneally (i.p.) in a concentration of 50 mg/kg in C57BL/6 mice as pretreatment for 13 days before implantation of melanoma cells induced a statistically significant increase in survival rate in animals [81]. According to the aforementioned in vitro studies, other candidates for the development of effective anticancer therapeutic agents could be verminoside, 4β-hydroxy-6-O-(p-hydroxybenzoyl)-tetrahydrolinaride, and 10-O-protocatechuyl catalpol.

In Vitro Studies
The anti-inflammatory properties of several Veronica species were evaluated in vitro and in vivo ( Table 4). In particular, the anti-inflammatory effect of Veronica peregrina L. was demonstrated in a recent study [10]. Different concentrations of V. peregrina methanolic extracts were incubated with C57BL/6 mouse peritoneal macrophages and the release of pro-inflammatory mediators, cyclooxygenase-2 (COX 2 ) and nitric oxide (NO), mediated by lipopolysaccharides was assessed; these were reduced dependent on the concentration of the extract. The mechanism of anti-inflammatory action was also highlighted: the inhibition of inducible nitric oxide synthase (iNOS) enzyme and consecutive decrease of NO production [10]. This study revealed the usefulness of V. peregrina extract as synergistic natural therapy in inflammatory diseases mediated by activated mast cells, such as arthritis, obesity, and atherosclerosis [82]. A similar study conducted by Harput et al. demonstrated the same biological anti-inflammatory activity of a methanol extract of five Veronica species which inhibited NO release from activated macrophages [74].
In another study, aqueous-acetone extracts of V. teucrium, V. jacquinii, and V. urticifola were tested for this effect on calcium ionophore-stimulated platelets, which release pro-inflammatory enzymes 12-lipoxygenase (12-LOX) and cyclooxygenase-1 (COX-1) with tromboxane B 2 (TXB 2 ) and prostaglandin E2 (PGE 2 ). COX 1 and its metabolite PGE 2 were inhibited by V. urticifolia extract, while the 12-LOX enzyme was blocked only by the V. jacquinii extract (IC 50 = 1.072 mg/mL). This anti-inflammatory activity is correlated with the presence of the main chemical compounds genistein, baicalin, isoquercetin, and hyperoside, but the mechanism of action is not yet completely known [13]. The results of these studies are important for human health, because natural remedies can successfully complete anti-inflammatory medical treatments.
Extracts of V. chamaedrys and V. officinalis also revealed in vitro anti-inflammatory activity on peroxisome proliferator-activated receptors (PPARs) and on the pro-inflammatory mediators, interleukin-8 (IL-8) and E-selectin [17]. Among the active constituents, V. officinalis extract rich in iridoid glycosides (verproside and verminoside) inhibited pro-inflammatory mediators via the nuclear factor kappa B (NF-κB) signaling pathway in a human lung cell line [15].
In another study, the anti-inflammatory action of V. officinalis extract in allergic inflammatory lung diseases was evaluated using A549 human lung epithelial cells. The results showed that the V. officinalis extract (40 to 160 µg/mL) inhibited in a dose-dependent manner the release of the main pro-inflammatory mediators IL-6 and IL-8, prostaglandin E 2 , and eotaxin. The data of the study did not clarify the mode of action of V. officinalis extract, but it is supposed that there is a link between the most abundant and important active compounds, iridoid glycosides (verminoside and verproside), and the anti-inflammatory effect [15]. These positive molecular results are in contrast to other studies that showed that the oral bioavailability of verproside is very low [83]. Nonetheless, more studies are required to elucidate the metabolites formed and their bioactivity. Moreover, an alternative way to administer V. officinalis extracts could be the pulmonary route using a spray for inhalation with the direct release of active substances to the lung epithelium [83].

In Vivo Studies
Inflammatory bowel diseases (IBD) (Crohn s disease and ulcerative colitis) are a heterogeneous group of diseases with incomplete elucidate pathogenesis defined by inflammation, persistence, or recurrence in the gastrointestinal tract, different in terms of extension of lesions, symptoms, prognosis, and treatment [84]. The triggers of these diseases are complex and mediated by cytokines (interleukins, IL) [85] and signal transducer activator of transcription 3 (STAT3) [86,87], pro-inflammatory enzymes such as cyclooxygenase-2 (COX 2 ) [88], and non-specific pro-inflammatory mediators, e.g., thromboxanes, prostaglandins, leukotrienes, oxygen free radicals, and NO [89]. Understanding these mechanisms led to research of new therapies, including natural alternatives.
Thus, Akanda and co-workers investigated the anti-inflammatory property of V. polita species on experimental colitis induced in mice using dextran sulfate sodium (DSS) [90]. Forty mice divided into four groups were included in the study: control, mice treated with DSS, mice treated with DSS plus V. polita extract (200 mg/kg), and the last group received DSS associated with dexamethasone. The results of the study were surprising because, for the mice treated with V. polita associated with dexamethasone compared to those treated with DSS alone, the following data were obtained: pro-inflammatory cytokines (IL-1β, IL-6, tumor necrosis factor alpha (TNF-α)) were not activated and NO production in the intestine was reduced. In addition, COX-2, NF-κB, and the Janus kinase 2 (JAK2)/STAT3 signaling pathway were blocked in mice co-treated with DSS and V. polita, similarly to the group of mice co-treated with DSS and dexamethasone [90]. This can be attributed to the high content of total phenols and flavonoids of this species [46].
Kupeli et al. conducted a study concerning the antinociceptive and anti-inflammatory effect of the methanol and aqueous extracts of the species V. anagallis-aquatica, using a phenyl-p-benzoquinone (PBQ) writhing test and carrageenan-induced hind paw edema model in mice. Only the methanolic extract (500 mg/kg) showed significant antinociceptive and anti-inflammatory effects, which could be related to the presence of iridoid glucosides, catalposide and verproside. In addition, no toxic and adverse irritant gastric effects were revealed [16].
A new pharmaceutical formulation consisting of a microemulsion of V. persica was tested for anti-inflammatory effects on rat paw edema induced by two different substances-kaolin, stimulating pro-inflammatory cytokines, and dextran, increasing the release of histamine and consequently inducing vascular permeability. Inflammation induced by dextran was totally reduced following the co-administration of V. persica microemulsion compared to the kaolin inflammation. This effect was correlated with its high content of polyphenols and iridoids [65].

Other Properties
The neuroprotective and angiogenic effects of the methanolic and aqueous-acetone extracts of three Veronica species, V. teucrium, V. jacquinii, and V. urticifolia, were determined on human SH-SY5Y cell line neuroblastoma. The neurotoxicity was induced on the cells by oxidative and nitrosative stress using H 2 O 2 and nitroprusside sodium, respectively. The six extracts showed a moderate neuroprotective effect, and this effect was higher for the cytotoxicity induced by oxidative stress and using V. urticifolia extract. Moreover, these authors showed that Veronica extracts, especially the V. jacquinni methanol extract and V. teucrium aqueous-acetone extract, showed antiangiogenic properties by preventing the formation of tubular structures in a Matrigel assay. Although this effect was probably due to the content of acteoside and aucubine, the mechanisms of action are unclear [79].

Food Applications of Veronica Plants and Other Uses
The enzymatic activity in foodstuffs result in some chemical reactions leading to food spoilage, thus making foods inedible or decreasing their qualities [91,92]. In addition to being an important health issue, which may sometimes lead to death, it may also yield great economic losses [93]. The World Health Organization reported that unsafe food resulted in illnesses in approximately two billion people throughout the world, and some of the cases were actually fatal [94]. There are some other methods that could prevent food spoilage, such as thermal processing; however, this technique might also result in a decrease in the nutritional value and quality of the food while reducing vegetative microorganisms. Therefore, to prevent food-related diseases and even death, a preservative should be used and this preservative has to be appropriate for the health of consumers and should not yield toxic materials [95]. Food manufacturers also search for compounds that could meet the expectations of healthy food trends, and this is also an important issue since synthetic preservatives are known to yield unwanted health results [96]. Veronica species have widespread traditional use throughout the world and, in addition to this usage, stems and leaves of some of the species are consumed as food in certain regions. Moreover, according to the Food Additive Status List by the Food and Drug Administration (FDA), Veronica species are listed as substances in conjunction with flavors that can be used in alcoholic beverages only [97]. However, since some members of this genus are also consumed by humans and used in traditional medicine, we can consider that usage of this plant is relatively safe [57].
Thus, the use of Veronica species in food preservation could be plausible with no adverse effects since some of them were demonstrated to possess antimicrobial effects (see Section 4). Although not directed on their use in food matrices, these studies also proved that the species extracts possess antibacterial effects and, thus, this genus has a potential to be used in food preservation [11,47,[98][99][100]. In this sense, the antimicrobial effect of Veronica plants extracts against foodborne pathogenic and food contaminant bacteria was demonstrated by several authors, but with modest activity. As an example, the results of the experiment conducted by  on the antibacterial activity of V. officinalis, V. teucrium, and V. orchidea ethanolic extracts, tested using the microdilution assay, revealed that the most sensitive bacterial strains to V. officinalis were Listeria monocytogenes (ATCC 19114) and Listeria ivanovii (ATCC 19119), with the same values of minimum inhibitory (MIC) and minimum bactericidal concentration (MBC) of 7.81 mg/mL. In the case of V. teucrium antibacterial activity, the strains of Staphylococcus aureus (ATCC 49444), Bacillus cereus (ATCC 11778), and Enterococcus faecalis (ATCC 29212) showed equal values for MIC and MBC (7.81 mg/mL), being the most sensitive. The ethanolic extract of V. orchidea inhibited L. monocytogenes and L. ivanovii with MIC = 3.9 mg/mL and MBC = 7.81 mg/mL. Regarding the V. officinalis extract, Bacillus cereus, Pseudomonas aeruginosa (ATCC 27853), and Escherichia coli (ATCC 25922) were the most resistant species with MIC and MBC values higher than 15.62 mg/mL. According to , the activity of these extracts against Gram-positive bacteria like Listeria species and S. aureus could be related to their high β-sitosterol, campesterol, and stigmasterol content. Phenolic constituents such as apigenin, luteolin, and their glycosides could also contribute [11,20,47]. Another work also suggested that V. montana L. water extract and its main phenolic compound, protocatechuic acid, showed the highest antibacterial effect against S. aureus (ATCC 6538) (MIC and MBC, 7.5 mg/mL) but a poor effect against B. cereus (human isolate) (MIC = 22.5 mg/mL; MBC 45 mg/mL) [57].
The incorporation of these species in a food system was investigated recently. In this sense, the water extract of V. montana exerted antimicrobial effects against six pathogenic bacteria, but it was more effective against L. monocytogenes (MIC, 7.5 mg/L; MBC, 15.0 mg/mL). Its major compound, protocatechuic acid (15.7 mg/g), also showed antibacterial properties (MIC, 0.75 mg/L; MBC, 1.0 mg/mL) and it was evaluated after its incorporation in cream cheese, using this bacterium as a cheese contaminant. This compound preserved cream cheese by inhibiting the growth of L. monocytogenes at room temperature and in the refrigerator after three days of inoculation, without compromising the sensory properties. The compound was shown to alter the permeability of the bacterial cytoplasmic membrane, and this makes this plant species and its major component promising antibacterial food preservatives [57].
The antioxidant properties (see Section 5) can be also exploited to preserve food quality and reduce rancidity and off-flavors, but these studies were not carried out on food systems. In this context, further studies are required to establish their real applicability as natural preservatives instead of synthetic ones, including dosages, advantages, and disadvantages.
In another context, some wild edible plants are attracting attention as novel food ingredients for gourmet in agro-tourism, for example, for salads and refreshing candy products. Ricola ® (Laufen, Switzerland) sweets are made with V. officinalis in conjunction with other medicinal plants, used to refresh the mouth and throat. Blanco-Salas et al. (2019) suggested that V. anagallis-aquatica, among other wild plants in "Sierra Grande de Hornachos" (Spain), already entered the high Spanish hotel industry and small selected market niches, due to its sensory and nutritional characteristics [101]. As commented before, this is a natural source of iridoids and other phytochemicals.

Conclusions
More than 100 phytochemicals were identified in Veronica plants, which mainly belong to iridoid glycosides and phenolic compounds, particularly flavones and terpenoids. Veronica plants are described in traditional medicine for the treatment of many diseases, especially related to inflammatory disorders. In addition, they represent importance in cosmetic and food industries. The review of the literature highlights that Veronica plants have good antioxidant, anti-inflammatory, antimicrobial, and anticancer abilities, which are mainly related to the presence of iridoid glucosides and phenolic constituents. The antioxidant properties of Veronica species were determined through different in vitro and in vivo studies. In addition, Veronica plants showed interesting antimicrobial effects, and most studies focused on its effects against both Gram-positive and Gram-negative bacteria. This also included foodborne pathogens, such as L. monocytogenes. The anti-inflammatory studies agreed with the use of Veronica remedies for anti-inflammatory medical treatments. Many authors also hypothesized that the cytotoxic activity of Veronica plants is associated with their antioxidant and anti-inflammatory effects. However, studies proving the therapeutic effects of the Veronica genus in humans are scarce, with the exception of V. officinalis as an anti-wrinkle agent. Only few studies were conducted in vivo, while the molecular mechanisms of the pharmacotherapeutic effects remain still unknown. When looking at the food applications of Veronica plants, promissory reports showed that its incorporation into several food matrices, such as dairy products, results in an improvement of the shelf-life, through exerting antimicrobial and antioxidant effects. Thus, these plants may be conceived as upcoming and effective natural food preservatives.