Pharmacological, biological and phytochemical aspects of Thymus munbyanus Boiss. & Reut.: A review

Thymus munbyanus Boiss. & Reut., is a small shrub endemic to Morocco and Algeria, and it is commonly used to treat several diseases, including digestive, circulatory, genital, skin, urinary, nervous and respiratory diseases. The extracts of this thyme are rich in a wide variety of phenolic compounds such as polyphenols and volatile phenols and exhibit numerous biological activities. The present review summarizes the literature investigations reported on Thymus munbyanus concerning various pharmacological and biological properties as well as phytochemical aspects. This species revealed a richness in phenolic compounds in its volatile oils, including thymol and car-vacrol, as well as in its non - volatile extracts including phenolic acids phenolic acids (rosmarinic acid, caffeic acid, salvianolic acid, ferulic acid, etc.), flavonoids (luteolin, gallocatechin, quercetin, isorhamnetin, etc.). Moreover, powerful antioxidant and antimicrobial properties were reported for this Thymus species, which are attributed to its richness in bioactive antioxidants. Furthermore, this thyme was found to possess important nephropro-tective, hepatoprotective, and tumor cytotoxic properties. In conclusion, Thymus munbyanus is an important natural source of bioactive phenolic compounds that can be used for developing alternative natural drugs for the treatment and prevention of several pathologies.


Introduction
Countless Lamiaceae species are used as medicinal and aromatic plants all over the world.This herbal family is one of the most important angiosperm families that consists of 236 genera and more than 7000 species.It also contains numerous species with medicinal and economic importance around the world.Among those 236 genera, the genus Thymus is one of the most important and diversified genera, comprising over 300 species distributed throughout the world (1).The plants of this genus have been used since ancient times for their beneficial health properties, which are attributed to their richness in phytochemical components, especially essential oils and polyphenols.
Thymus species are widely used medicinal plants in folk medicine, food, and pharmaceutical industries due to their pharmacological features.Moreover, these species have been used in the treatment of several diseas-Thymus munbyanus Boiss.& Reut.(T.munbyanus), is a small shrub endemic to Morocco and Algeria and it is commonly used in North Africa and several other countries to treat several diseases.This species englobes four subspecies namely T. munbyanus subsp.munbyanus, T. munbyanus subsp.coloratus, T. munbyanus subsp.ciliates, and T. munbyanus subsp.abylaeus (4).Moreover, this thyme was reported in several studies for its interesting biological and pharmacological properties.
In the present review, we will summarize and discuss the studies reported on T. munbyanus, including phytochemical, pharmacological, and biological investigations.

Antioxidant activity
Thyme plants are an important source of natural antioxidants.Numerous Thymus species from different countries around the world are reported to exhibit powerful antioxidant effects (9)(10)(11).Likewise, T. munbyanus antioxidant potential was studied using different methods.The antioxidant activity of chloroform, ethyl acetate, and n-butanol extracts, as well as isolated compounds from T. munbyanus using total antioxidant capacity and free

Quercetin Luteolin
Fig. 1.Chemical structure of some abundant phenolic compounds in Thymus munbyanus radical scavenging activity assays were analyzed.The results of these studies showed that the different solvent extracts have an antioxidant effect comparable to that of the antioxidant standards butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT), whereas some of the isolated compounds, including methyl caffeate, tetrahydroxyflavanone, and pluchoic acid exerted a higher antioxidant effect than those of antioxidant standards which are known to be potent antioxidants (12,13).Moreover, the antioxidant potential of the essential oils of T. munbyanus has been evaluated in several studies.Benchabane et al.
(2014) evaluated the antioxidant activity of T. munbyanus essential oil using DPPH free radical and thiobarbituric acid reactive substances assays and reported that T. munbyanus essential oil exerted a powerful antioxidant activity, which was similar to that of BHT (14).Moreover, Bendif et al.
(2016) studied the antioxidant activity of essential oils from different parts (Leaves/stems and flower) of T. munbyanus using DPPH, ABTS, and FRAP in-vitro antioxidant tests.The results of this study showed that all the oils showed moderate antioxidant activity.Leaves with stems essential oils exhibited a considerably higher activity compared to flowers essential oils, but significantly lower than that exerted by the standard antioxidant Trolox (15).Furthermore, the powerful antioxidant potential reported for this thyme is generally attributed to the presence of powerful phenolic antioxidants in its volatile and non-volatile extracts in important amounts.

Antimicrobial activity
Thymus species essential oils exhibit a broad range of biological properties, including antibacterial and antifungal activities.Heni et al. studied the antibacterial activity of T. munbyanus essential oil against Listeria monocytogenes and Bacillus cereus, which are implicated in the contamination and poisoning of food.The results of this study demonstrated that the essential oil of this thyme exhibited a very good antibacterial activity against the two pathogenic strains.These pathogens displayed high sensitivity against the essential oil with inhibition diameters of 28.6 mm (L.monocytogenes) and 40 mm (B.cereus) and with a relatively low minimum inhibitory concentration (0.18 mg/ml) (16).Moreover, Bendifa et al. screened the antimicrobial activity of the essential oils of T. munbyanus using the disc diffusion method against four bacterial strains (Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and Enterococcus faecalis) and one yeast strain (Candida albicans).The results revealed that the essential oil of this thyme had a moderate antimicrobial effect on S. aureus, E. coli, and C. albicans with inhibition zone diameters in the range of 9 to 10 mm (15).Other studies investigated the antimicrobial properties of T. munbyanus essential oils against different microbial strains (17, 18) and showed that the essential oil of this thyme displays good antimicrobial effects.

Hepatoprotective activity
The liver is a vital organ with important metabolic functions, including body detoxification.However, hepatic dysfunction can cause serious complications.Liver disorders are associated with multiple factors such as biological, chemical, and drug overdose (19).Several plants were investigated for their hepatoprotective effects, including Thymus species (20).Furthermore, investigation was carried out to analyze the hepatoprotective effect of T. munbyanus against 2,4-dichlorophenoxyacetic acid (2,4-D) induced hepatic oxidative stress in albino Wistar rats by evaluating the biomarkers of hepatic function (alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, lactate dehydrogenase, and gamma-glutamyl transferase), glutathione (GSH) levels, protein, and albumin contents, lipid peroxidation marker (malondialdehyde MDA), enzymatic antioxidants (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione stransferase (GST) enzymes), and histopathological evaluation.This study revealed that the alterations induced by 2,4 -D at a dose of 5 mg/kg bw in the histological and biochemical parameters of the liver were significantly attenuated.It was reported that 2,4-D increased the biomarkers of hepatic function and MDA level significantly, decreased protein and albumin contents and hepatic GSH level, and the major enzymatic antioxidants, as well as the development of histological alterations of liver tissue with severe damages on liver cells.However, T. munbyanus extract attenuated all the alterations significantly in the histological and biochemical parameters of the liver ( 21).This study reported promising findings on the hepatoprotective potential of this Thymus species.However, testing the extracts of this thyme using other methods and different models is recommended to confirm its hepatoprotective effects.

Nephroprotective activity
The kidney is a vital organ that displays crucial functions, including the elimination of metabolic nitrogen waste, regulation of the pH, the fluid and electrolyte balance, production of hormones, and activation of vitamin D (22).However, renal dysfunction can lead to serious complications, including cardiovascular disease, hypertension, anaemia, etc. (23).Numerous medicinal plants were investigated for their nephroprotective effects (24).they can attenuate the biochemical, functional, and structural renal toxicities and represent effective nephroprotective alternatives (25).Likewise, Trea et al. tested the protective effects of T. munbyanus aqueous extract against 2,4-D induced nephrotoxicity at a dose of 5 mg/kg of body weight in male albino rats.This chemical compound causes oxidative stress and renal injury in the kidney, which increases the potential markers of renal filtration (urea and creatinine), malondialdehyde, and carbonyl protein levels, a decrease in uric acid, GSH, and antioxidant enzymes levels (SOD, GPx, and GST).The results showed that T. munbyanus extract supplementation for one month restored some blood parameters and alleviated the adverse cytotoxic effects of 2,4-D by increasing the levels of certain antioxidants, consequently attenuating the intensity of oxidative stress induced by 2,4-D, and attenuated the histological alterations observed in the kidneys (26).

Tumor Cytotoxicity and anti-proliferative activity
Bendif et al. investigated the tumor cytotoxicity of T. munbyanus flowers, leaves, and stems essential oils against three human tumor cell lines, namely human glioblastoma multiforme, human breast adenocarcinoma, and human malignant melanoma.The results of this study revealed that the essential oils exhibited a slight cytotoxic effect against all cell lines.Flowers essential oils showed the highest cytotoxic effect against malignant melanoma cell line (IC50 value = 46.95µg/ml), whereas the lowest effects were shown by stems and leaves essential oil against the same tumor cell line (15).Moreover, Tefiani et al. (2015) compared the anti-proliferative activity of the essential oils of T. munbyanus and an Apiaceae species (Ammoides pusilla) against human acute monocytic leukaemia cell line (THP-1).The results showed that T. munbyanus essential oil showed a significant (p˂0.05)anti-proliferative activity, in which after seven days of incubation at a concentration of 100 μg/mL of essential oils, the proliferation percentage of THP-1 cells was 13% for T. munbyanus comparing to 64% for A. pusilla (27).

Discussion
Thyme plants are known to have important amounts of phenolic compounds and exhibit strong antioxidant effects.Several studies showed that the species of this genus are rich in flavonoids, phenolic acids, and their derivatives (28,29).For example, Sarfaraz et al. (2021) carried out a chemical analysis of the methanolic extract polyphenolic composition of 11 Iranian Thymus species.The results of the analysis showed that these possess high amounts of polyphenols in which rosmarinic acid, salvianolic acid, and cinnamic acid are the major phenolic acids, and apigenin, epicatechin, and naringenin as the major flavonoids.Moreover, Boros et al. (2010) evaluated the chemical composition of five Hungarian Thymus species.
The results of the analysis of this study showed that the hydro-methanolic extracts of these species are rich in phenolic acids and flavonoids.Phenolic acids (rosmarinic acid, ferulic acid, caffeic acid, chlorogenic acid), flavanones (naringenin, eriodictyol and dihydroquercetin), flavones (apigenin), as well as flavonols (quercetin and rutin) were present in every examined Thymus species in which rosmarinic acid was the dominant compound (1.436 -83.49mg/g) (30).Likewise, T. munbyanus possess significant amounts of these polyphenols, including phenolic acids such as rosmarinic acid and different types of flavonoids.This suggests that the polyphenolic extracts of Thymus species, including T. munbyanus are an important source of these important metabolites.Furthermore, the chemical composition of the essential oils of T. munbyanus contains bioactive volatile phenols, namely thymol and carvacrol (7).Thymus species are a source of these volatile phenolic phytochemicals.Several studies investigated the chemical composition of several Thymus species essential oils and showed that thymol and carvacrol are abundant volatile compounds, including T. serpyllum and T. algeriensis (31), T. daenensis and T. kotschyanus (32), T. kotschyanus and T. vulgaris, etc. (33).
The extracts and essential oils of Thymus species, including T. munbyanus are a potent source of a wide variety of antioxidant phenolic compounds.Several thyme species demonstrated important antioxidant properties using diverse in-vitro (34,35) and in-vivo (36,37) methods.Thus, the potent antioxidant effects of the extracts of these thymes, including T. munbyanus can help in the protection against free radicals and their harmful effects and reduce the risk of occurrence of numerous pathologies.
T. munbyanus essential oils display great antimicrobial effects.This biological activity is commonly observed in thyme species.Several studies investigated the antimicrobial effects of Thymus plants and other Lamiaceae species.These studies showed that Thymus species display the most significant antimicrobial effects (38)(39)(40).In addition, these effects can be attributed to the richness of their essential oils in thymol and carvacrol.These two phenolic compounds exhibit powerful antimicrobial properties.Moreover, the most frequently reported mechanism of antibacterial effect of these compounds involves the disruption of the bacterial membrane causing bacterial lysis and leakage of intracellular contents (41).
The kidney and the liver are vital organs with indispensable functions in the body.Protecting these organs against harmful agents can prevent several diseases.The hepatoprotective and nephroprotective actions displayed by T. munbyanus show that this species can be efficient in the prevention of kidney and liver pathologies.Moreover, several thyme plants were also investigated for their hepatoprotective and nephroprotective effects (T.vulgaris (42), T. daenensis (43), etc.).These investigations showed that the species of this genus display important liver and kidney protective effects.Thus, thyme plants including T. munbyanus can be promising sources for developing more natural and safer hepatoprotective and nephroprotective alternative drugs.
Uncontrolled cell proliferation or cancer is a major health issue and it is the second leading cause of death worldwide.Moreover, cancer chemotherapy or radiation therapy are associated with a high mortality rate and toxicity (44).Therefore, developing newer and safer treatment methods, such as plant-based therapy can be a promising alternative method.The use of medicinal plants for the prevention and treatment of cancer is gaining more attention due to their richness in bioactive tumor cytotoxic constituents and fewer adverse effects (45).Several thyme plants were investigated for their anticancer properties particularly in in-vitro cellular models and revealed that they have a great potency as sources of anticancer agents (46).Hence, T. munbyanus extracts should be more investigated in in-vitro and in-vivo methods to confirm the beneficial effects of this species against cell proliferation.
Due to the beneficial effects of the different extracts of T. munbyanus on several pathologies and the richness of these extracts in a wide variety of bioactive compounds, this species can be considered as an important source of potent drugs in the future.However, more studies should be carried out, especially on the polyphenolic extracts of this species, to evaluate their bioactivity against several diseases such as diabetes, hyperlipidemia, atherosclerosis, etc.

Conclusion
This present study aims to review the phytochemical, pharmacological, and biological activities reported on T. munbyanus.This thyme showed that it is rich in a wide variety of volatile and non-volatile bioactive phenolic compounds, including rosmarinic acid, thymol, and carvacrol.The extracts of this thyme display significant antioxidant, antimicrobial, antitumoral, hepatoprotective, and nephroprotective properties.T. munbyanus can be used for developing alternative natural drugs for the treatment and prevention of several pathologies.