Send Orders of Reprints at reprints@benthamscience.

net
382 Current Analytical Chemistry, 2013, 9, 382-396

Overview on Mentha and Thymus Polyphenols

Olívia R. Pereira1,2 and Susana M. Cardoso1,3,*

1
CERNAS - Escola Superior Agrária, Instituto Politécnico de Coimbra, Bencanta, 3040-316 Coimbra, Portugal
2
Escola Superior de Saúde, Instituto Politécnico de Bragança, Av. D. Afonso V, 5300-121 Bragança, Portugal
3
CIMO - Escola Superior Agrária, Instituto Politécnico de Bragança, Campus de Santa Apolónia, 5301-854 Bragança,
Portugal

Abstract: Mentha and Thymus are important genera of the Lamiaceae family widely distributed in the entire World and
commonly used in traditional medicine. Indeed, many species of the two genera have been credited with a large list of
health-benefit effects, including antioxidant, anti-inflammatory, antimicrobial, analgesic, neuroprotective and anticarcino-
genic. In turn, these properties have been associated to the polyphenolic composition of the plants. The present review
summarizes the phenolic constituents found in Mentha and Thymus genera, as well as the main methods applied in their
extraction, purification and identification. Reported species of Mentha and Thymus usually comprise derivatives of caffeic
acid and distinct glycosidic forms of the flavonoids luteolin, apigenin, eriodictyol and naringenin. At present, the phenolic
composition of many relevant plants of Mentha and Thymus is still unknown and thus, more studies are required for the
adequate phenolic characterization of these two genera. In this context, the present implementation of faster and reliable
analytical methodologies, as e.g. the chromatographic techniques hyphenated with mass spectrometry, will surely be an
enormous tool in the upgrading of the missing information.

Keywords: Mentha, Thymus, Phenolic compounds, Phenolic acids, Flavonoids, HPLC, Mass spectrometry, NMR.

1. INTRODUCTION genera consists of eighteen species and includes eleven hy-

brids.
Lamiaceae is one important botanical family with about
230 genera, some of which have been used for centuries in Other important genus of Lamiaceae family is Thymus.
traditional medicine. One important Lamiaceae genus, the This genus comprises 300 to 400 endemic species, widely
Mentha, encloses species distributed over the worldwide, distributed throughout the world and particularly abundant in
with special abundance in temperate regions of Eurasia, Aus- West Mediterranean region [8]. Many Thymus plants appear
tralia and South Africa [1, 2]. In general, the plants of this as perennial and aromatic subshrubs or shrubs with quadran-
genus growth in distinct environments, although they are gular stem erect to prostrate. Usually, the plants have big
preferentially found in damp or wet places. Mentha plants clusters of flowers of diverse color (white, cream, pink or
are mainly perennial and grow 10 - 120 cm tall, with erect, violet) [9]. The leaves are simple, entire or sometimes
branched, four-sided or squared stems. They have white to toothed, frequently revolute, glabrous or hairy. Several spe-
purple flowers, small fruits containing a few number of cies of this genus are often cultivated for culinary purposes
seeds (one to four) and have leaves typically arranged in and to be used as medicinal plants. The latter usage is due to
opposite pairs, also showing a great color diversity [3-5]. several pharmacological of these plants, which are mostly
described for thyme and its wild form (Thymus vulgaris L.
Several mint species are of a great economic importance
and T. serpyllum, respectively).
and are used in food, flavour, cosmetic and pharmaceutical
industries mainly because of peculiar properties of their Previous studies on the chemical composition of Mentha
essential oils. The most used mints are peppermint (Mentha and Thymus have mainly dealt with their essential oils [10-

piperita) and M. spicata, commonly known as spearmint 13]. However in recent years, an increasing number of inves-
[6, 7]. Due to the frequent occurrence and to the vast tigations have focused on other secondary metabolites, in-
distribution of Mentha cultivation, there is a high polymor- cluding the polyphenols. In fact, phenolic compounds of
phism in this genus. More, hybridization in wild and culti- Mentha and Thymus plants have been associated to their
vated plants is emerging. These facts lead to the difficult and beneficial properties, supporting their ethnopharmacological
not consensual systematic classification of Mentha species. usage. These effects include the antioxidant capacity, that
Despite this, recent studies based in morphological, cyto- has been described for M. x piperita, M. “Native Wilmet”,
logical, and chemical markers
indicated that the Mentha M. dalmatica and M. spicata [1, 14, 15], the antitumorogenic
ability, that has been reported for M. spicata and M. piperita
L. species [16, 17], the neuroprotective action reported for
* Address correspondence to this author at the CERNAS - Escola Superior M.  piperita and M. aquatica species [18-20] and the anti-
Agrária, Instituto Politécnico de Coimbra, Bencanta, 3040-316 Coimbra, inflammatory activities showed for M. aquatica plant [21].
Portugal; Tel: +351 239 802940; Fax: +351 273 239 802979;
E-mail: scardoso@esac.pt In the case of Thymus genus, T. vulgaris L. species has been

1875-6727/13 $58.00+.00 © 2013 Bentham Science Publishers

Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 383

described to possess antitumorogenic [16, 22] and free radi- these genera will be described bellow in more detail. A re-
cal-scavenging [23] activities. This species together with T. sume of these phenolic compounds and the respective spe-
mastichina and T. capitata also have important antioxidant cies where they have been found are shown in Table 1.
activities [24-26], while T. broussonetii and T. pulegioides Moreover, the structure of these compounds is depicted in
have been shown to exert analgesic [27] and cardioprotective Fig. (1).
capacities [28], respectively. Note that Mentha and Thymus
Caffeic Acid and Derivatives
genus show many similarities regarding the type of polyphe-
nols in their composition, despite differences in their preva- Reported literature on Mentha and Thymus genera indi-
lence. As a result of that, several studies have simultane- cates that caffeic acid and its derivative compounds repre-
ously focused species of these two genera [29-31]. sents 15 - 60% and 60 - 80% of their total phenolic com-
pounds, respectively [34, 38-46]. The free form of this acid
The phenolic characterization of plants commonly en- is frequently detected, but most representative compounds
gage their extraction with water, methanol, ethanol or aque- are those commonly known as dimeric and trimeric forms of
ous mixtures and their further analysis by reversed phase the acid, as well as their derivatives.
high performance liquid chromatography (HPLC/UV/PDA)
[32] combined or, alternatively, hyphenated with electros- Caffeic acid (1) is vastly described in Mentha genus [1,
pray mass spectrometry (LC-MS). These techniques have 38-41, 47], although its relative abundance greatly varies in
allowed to perform the identification and quantification of between the reported studies. In general, the free form of the
phenolic compounds in many extracts of Mentha and Thy- acid appears as a minor phenolic component (approximately
mus genera plants. Nuclear magnetic resonance (NMR) ap- 1% of total phenolic content), as described by Dorman et al
pears as a powerful complementary technique for structural [1] for nine distinct Mentha species.
assignment [33], in particular of unknown or complex phe- As for Mentha species, caffeic acid in Thymus plants
nolic compounds. The present manuscript revises the main commonly appears in low relative quantities [43, 46]. Sev-
polyphenols described up to date in extracts of Mentha and eral authors have described caffeic acid in T. vulgaris L. [34,
Thymus plants and also examines the analytical methods 42, 43, 45, 46, 48-50] and reported data indicated a total of
used in their extraction, isolation and structural identifica- 0.1-0.48 mg/g of dry plant in this species with similar results
tion. for T. serpyllum [46, 49].
2. POLYPHENOLS OF MENTHA AND THYMUS Besides the free form of the acid, simple ester or gluco-
GENERA side derivatives of caffeic acid were also reported to occur as
Several studies have demonstrated that Mentha and Thy- minor phenolic components of Mentha and Thymus.
mus plants are rich in phenolic compounds, particularly in Namely, two esters of caffeic acid, the (Z,E)-[2-(3,5-
phenolic acids and flavonoids. As usually, the total polyphe- dihydroxyphenyl)ethenyl] 3-(3,4-dihydroxyphenyl)-2-
nolic content of these plants has been mainly estimated propenoate (2) and the (Z,E)-[2-(3,4-dihydroxyphenyl)-
through the use of Folin Ciocalteu method on enriched phe- ethenyl] 3-(3,4-dihydroxyphenyl)-2-propenoate (3), also
nolic extracts, which in turn have been obtained by water or, named nepetoidin A e B, respectively, have been reported in
more frequently, by ethanolic or methanolic aqueous solu- M. aquatica, M. longifolia and M. x villosa species [51]
tions. while chlorogenic acid (4) was reported in M.  piperita L.
[40]. For Thymus, caffeic acid ethyl ester (5) has been de-
The total polyphenolic content of M. pulegium and M. scribed for T. serpyllum [37] and hexoside forms of caffeic
viridis leaves and that of leaves and branches of M. ca-
acid (6) [35, 50], the compound chlorogenic acid [37, 50, 52]
nadensis has been previously established as approximately 8,
and dicaffeoylquinic acid (7) [50] have been described for T.
17 and 52 mg of gallic acid equivalent (GAE)/g of dry plant,
vulgaris L. Moreover these two latter compounds also have
respectively [34]. Moreover, Dorman et al [1] have reported
been reported to occur in T. webbianus [52].
a total phenolic content in the range of 47 – 77 mg of GAE/g
of dry plant, for the aerial parts of nine Mentha species. On the other hand, rosmarinic acid (8), the ester of caf-
Alternatively, values of aproximatly 20 – 46 mg of GAE/g feic acid with 3,4-dihydroxyphenyl lactic acid and com-
of dry plant have been described in aerial parts for the most monly known as a caffeic acid dimeric form, is the most
common Thymus species (T. vulgaris L.) [34-36] while a abundant phenolic in Mentha and Thymus genera. Its content
lower amount of total polyphenols (12 mg of GAE/g dry in M. x piperita L. species (the most studied Mentha species
plant) has been reported for T. serpyllum [37]. regarding phenolic compounds) was reported as being ap-
proximately 30% of the total phenolics [38-42] and even
2.1. Phenolic Acids higher in other Mentha species, such as M. aquatica, M. x
Phenolic acids occur in nature as hydroxycinnamic (C6- dalmatica, and M. canadensis L., where this phenolic was
C3) or hydroxybenzoic structures (C6-C1). Diversity in this reported to account for 3.1, 3.4 and 19.1 mg/g of dry plant
group is mainly due to the variation on the number and posi- (40, 46 and 90% of the total phenolics, respectively) [1, 34,
tions of hydroxyl functions in the phenolic ring. In the plant 41]. Rosmarinic acid has also been reported in M. spicata
kingdom, these compounds are often present in their free (1.1 mg/g of dry plant) and in M. longifolia, although no
form, as well as hexoside derivatives or as esters of benzoic content has been estimated for the latter species [1, 2, 41, 47,
or cynnamic acids. From all phenolic acids, Mentha and 53].
Thymus genera are particularly enriched in caffeic acid de- Rosmarinic acid is even more abundant in Thymus genus.
rivative compounds. These polyphenols, as well as other In T. vulgaris L. species this phenolic acid has been
hydroxycinnamic and hydroxybenzoic acids detected in described to account close to 70% of its total polyphenols,
384 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

O
HO
OH
X=
HO HO OH OH
O OH
HO OH O
O R HO
HO O
HO HO 4
1 R= H O O
5 R= CH 2 CH3 R1 2 R 1= H R 2= OH
6 R= Glc 3 R 1= OH R 2= H
R2 O HO

O R2
HO O O
O OH OH OH
O
HO O O O
HO R2 O OH R1
O O O
7 HO
HO
O O
HO 8 R 1= R2 = H
17 R1 = Glc
OH
HO O HO
R1 19 R1 = H R2 = CH3
9 R 1= H R2= X
10 R = R =1 X 2
OH
HO OH

HO OH
O OH HO OH
O CO2 ~ Mg 2 +
O CO2 - HO O OH
O CO2
O OH O CO2Na OH
O
HO 11 O
O
12
HO HO
R
OH
O O O HO OH
HO O
HO HO
O O HO
O
O R O
OH HO O
O
13 R= X O O
O 15 R= X R
OH 14 R= X O OH
O
O O
R
O HO
OH O
HO O R
HO O
OH
O O OH
HO
R OH
16 R= X 18 R= X
O
HO OH O
OH
HO
OH O
HO
20 R= X O O R
HO O
O
R OH HO
21 R= X
O
Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 385

O
O OH 23 R 1= R4= OH R2= R3 = H
R1
OH 24 R 1= R4= H R2 = R3 = OH
R4 25 R 1= R3= R4 = H R2 = OH
28 R 1= R2= R3 = OH R4 = H
R2 22 R 1= R2= H 29 R 1= R3= OCH3 R 2= OH R 4= H
26 R 1= R4= OCH 3 R2= OH R1 R3 30 R 1= R3= R4 = H R2 = OGlc
27 R 1= H R2= OH 31 R 1= R4= H R2 = OH R3 = OCH3
R2

32 R 4= OH R 1= R2 = R3 = R5 = H 46 R 1= OH R2 = CH 3 R3 = OCH3 R4 = R5 = H
33 R 1= R 3 = R5 = H R2 = Glc R4 = OH 47 R 1= OH R2 = CH 3 R3 = R4= OCH3 R 5 = H
34 R 1= R 3 = R5 = H R2 = Rut R4 = OH 48 R 1= OH R2 = R 5 = CH3 R3 = OCH3 R4 = H
35 R 1= R 3 = R5 = H R2 = GlcU R4 = OH R4 49 R 1= R 3= OCH3 R 2= R5 = CH3 R4 = H
36 R 1= R 3 = R5 = H R2 = GlcU CH 3 R 4 = OH O 50 R 1= R 3= R4 = OCH3 R2 = R5 = CH 3
37 R 1= R 3 = R5 = H R2 = GlcU R4= OGlcU R3 R5 51 R 1= R4 = OCH3 R2= CH3 R3= R5 = H
38 R 1= R5= H R2= Glc R3 = H R 4 = OGlc O O 52 R 1= OHR 2= CH3 R 3= R4 = R 5 = H
39 R 1= R 3 = R5 = H R2 = AcGlc R4 = OH R2 53 R 1= OCH3 R2 = R5 = CH3 R3 = R4 = H
40 R 1= R 3 = R5 = H R2 = GlcFer R4 = OH 54 R 1= OHR 2= R5 = CH3 R3 = R 4= H
41 R 1= R 2= R3 = R4= R5 = H 55 R 1= OCH3 R2 = CH3 R 3 = R4 = R 5= H
R1
42 R 2= Rut R 1= R3= R4 = R 5 = H 56 R 1= R 3 = OCH3 R2= CH3 R4 = R5 = H
43 R 2= Glc R1= R3= R4 = R 5 = H OH O 57 R 1= R 3 = OCH3 R2 = CH3 R4 = OH R5 = H
44 R 2= GlcU R1= R 3= R4 = R5 = H
45 R 1= R 3= Glc R 2= R4 = R 5= H

R2
58 R 1= R 3= H R2 = OH OH
O 59 R 1= Rut R 3= H R 2= OH
R3
60 R 1= Glc R 3= H R 2= OH
O O 61 R 1= GlcU R3 = H R 2= OH
HO O
R1 62 R 1= R 3= R2 = H
63 R 1= Glc R 2= R3= H
64 R 1= Rut R 2= R3= H 65
66 R 1= Rut R 2= OH R 3= CH 3 OH O
OH O

R2
OH
O
67 R 1= R3 = R4 = H R 2= OH R1 R3
68 R 1= R3 = H R2 = OH R4 = Rut HO O
69 R 1= Rut R 2= R3= R4 = H
O O R1
70 R 1= Rut R 2= R4= H R 3 =CH3
71 R 1= R3 = H R2 = OH R4 = Glc R4 OH
72 R 1= R3 = R4 = H R 2= OCH 3 O
OH O
73 R 1= R3 = H R2 = OCH3 R4 = Glc OH O 74 R 1= OH
75 R 1= H

GlcU- Glucuronide unit; Glc- Glucoside unit; Rut- Rutinoside unit; Ac- Acetyl unit; Fer- Feruloyl unit

Fig. (1). Chemical structures of polyphenols described in Mentha and Thymus plants. The reference numbers for the compound structures are
used throughout this paper.

Table 1. Phenolic Acids of Mentha and Thymus Genera

Compound Mentha species Thymus species

Caffeic acid derivatives

Caffeic acid (1) [34, 42, 43, 46, 48-
M. x piperita L. [1, 38-42, 58] T. vulgaris L.
50, 53]
M. aquatica [41] T. serpyllum [37, 46, 47, 49]
M. spicata [1, 47, 53]
M. canadensis L. [34]
M. x dalmatica [1]
T. quinquecostatus [54]
M. “Morocco” [41]
M. “Native Wilmet” [41]
M. arvensis [1, 41]
386 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

Table 1. contd…

Compound Mentha species Thymus species

Nepetoidin A (2) M. aquatica

M. longifolia [51]
Nepetoidin B (3)
M. x villosa

Chlorogenic acid (4) T. vulgaris L. [50]

M. x piperita L. [40] T. serpyllum [37]

T. webbianus [52]

Caffeic acid ethyl ester (5) T. serpyllum [37]

Caffeic acid glucoside (6) T. vulgaris L. [35, 50]

Dicaffeoylquinic acid (7) T. vulgaris L. [50]

T.webbianus [52]

Rosmarinic acid (8) [1, 2, 38-42, 58, [34, 35, 42-46, 49,
M. x piperita L. T. vulgaris L.
61-63] 53, 57, 58, 64]

M. aquatica [41] T. serpyllum [37, 46, 47, 49]

M. spicata [1, 41, 47, 53] T. sipyleus [55]

M. canadensis L. [34] T. quinquecostatus [54]

M. x dalmatica [1]

M. haplocalyx [1, 56]

M. “Morocco” [41]

M. “Native Wilmet” [41]

M. x verticillata [41]

M. arvensis [1, 41]

M. longifolia [2]

Lithospermic acid (9) M. x piperita L. [39] T. serpyllum [49]

M. haplocalyx [56]

Lithospermic acid B (10) M. haplocalyx [56]

Magnesium lithospermate B (11) M. haplocalyx [56]

Sodium lithospermate B (12) M. haplocalyx [56]

Cis salvianolic acid J (13) M. haplocalyx [56]

Salvianolic acid J (14) M. haplocalyx [56]

Didehydro-salvianolic acid (15) M. x piperita L. [2]

M. longifolia [2]

Salvianolic acid L (16) M. x piperita L. [2]

M. longifolia [2]

Rosmarinic acid glucoside (17) T. vulgaris L. [35]

3’-O-(8’’-Z-Caffeoyl)rosmarinic acid (18) T. vulgaris L. [57]
Methyl rosmarinate (19) T. vulgaris L. [49]
Salvianolic acid I (20) T. vulgaris L. [35]

Salvianolic acid K (21) T. vulgaris L. [35]

Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 387

Table 1. contd…

Compound Mentha species Thymus species

Other phenolic acids

Cinnamic acid (22) M. x piperita L. [6]
Gentisic acid (23) M. x piperita L. [42] T. vulgaris L. [42]
Protocatechuic acid (24) M. x piperita L. [42] T. vulgaris L. [42, 50]
T. vulgaris L. [42, 46, 50]
Hydroxybenzoic acid (25) M. x piperita L. [42]
T. serpyllum [37]
Ferulic acid (26) T. vulgaris L. [48, 50]
T. vulgaris L. [34, 42, 50]
p-Coumaric acid (27) T. webbianus [52]
T. serpyllum [37]
T. vulgaris L. [34, 43, 50]
Gallic acid (28)
T. webbianus [52]
Syringic acid (29) T. vulgaris L. [42]
Hydroxybenzoic acid-O-hexoside (30) T. vulgaris L. [50]
T. vulgaris L. [42, 50]
Vanillic acid (31) M. x piperita L. [42]
T. serpyllum [37]

representing approximately 3.4 to 22 mg/g of dry plant [34, much frequently detected in the latter genus. In more detail,
43, 49]. Besides this species, rosmarinic acid was also de- M. piperita L. has been described to contain minor amounts
tected as a major phenolic compound in other Thymus spe- of cinnamic acid (22) [6], gentisic acid (23), protocatechuic
cies, such as T. serpyllum, T. sipyleus and T. quinquecostatus acid (24) and p-hydroxybenzoic acid (25) [42]. Several hy-
var. japonica [37, 46, 49, 54, 55]. droxycinnamic and hydroxybenzoic acids were reported for
Thymus genus. Besides the caffeic acid, hydroxycinnamic
In addition to rosmarinic acid, other caffeic acid deriva-
acids previously detected in Thymus include ferulic acid (26)
tive compounds enclosed in the class of depsides (esters
[48, 50] and p-coumaric acid (27) [34, 37, 42, 50, 52]. More,
formed by the condensation of two or more molecules of
the hydroxybenzoic acids gallic acid (28) [34, 43, 50], gen-
phenolic carboxylic acids compounds), are relevant in Men-
tha and Thymus genera. Lithospermic acid (9), usually clas- tisic acid [42], protocatechuic acid [42, 50, 54], syringic acid
(29) [37, 42, 50], p- hydroxybenzoic acid [37, 42, 46], hy-
sified as a caffeic acid trimer, is a phenolic constituent of M.
droxybenzoic acid-O-hexoside [50] (30) and vanillic acid
x piperita L. and M. haplocalyx and its content in the first
(31) [37, 42, 50] have been detected in T. vulgaris L. while
species has been described to be approximately 3 mg/g of
some of those have also been described to occur in T. serpyl-
dry plant [39, 56]. Other caffeic acid derivatives, namely the
lum, T. quinquecostatus and T. webbianus.
lithospermic acid B (10), magnesium lithospermate B (11),
lithospermate B (12) and two salvianolic acid derivatives
2.2. Flavonoids
(cis salvianolic acid J (13), salvianolic acid J (14)) were only
detected in M. haplocalyx. Alternatively, the first species Mentha and Thymus species are rich in flavonoids. Men-
contained didehydro-salvianolic acid (15) and one tetrameric tha species are particularly abundant in flavanones (10-70%
form of caffeic acid (salvianolic acid L (16)) [2, 56]. of their total phenolics), although their flavones content is
In respect to Thymus genus, five derivatives of rosma- also relevant. The latter class of flavonoids is the most
common in Thymus genus. On the other hand, flavonols and
rinic acid namely, its glucoside (17), the 3’-O-(8’’-Z-
dihydroflavonols are only found as minor components in
Caffeoyl)rosmarinic acid (18), methyl rosmarinate (0.6 mg/g
these two plant genera. Table 2 resumes the main flavonoids
of dry plant) (19) and the two trimers of caffeic acid (salvi-
found in Mentha and Thymus genera and their respective
anolic acid I (20) and salvianolic acid K (21)) [35, 49, 57]
structure is shown in Fig. (1).
have been described to occur in T. vulgaris L.. Alternatively,
lithospermic acid has been described as accounting for ap-
Flavones
proximately 12 mg/g of dry plant in plant extract of T. ser-
pyllum [49]. Luteolin and its derivatives are the main flavones de-
scribed in Mentha and Thymus genera [1, 39-41, 49, 50, 58,
Other Phenolic Acids 59]. Luteolin (32) has been detected in M. piperita L., M.
Besides caffeic acid and their derivatives, other phenolic aquatica, M. longifolia, M. pulegium, M. arvensis, M. haplo-
acids have also been described in Mentha and Thymus plants calyx and M. spicata [1, 40, 58-60] and, as reported by
[1, 55] and, according to literature data, these compounds are Dorman et al. [1] for the latter three species, its content can
388 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

Table 2. Main Flavonoids in Mentha and Thymus Genera

Compound Mentha Species Thymus Species

Flavones

M. pulegium [60] T. vulgaris L. [44-46, 49, 50, 58]

M. x piperita L. [39, 40, 58, 60] T. serpyllum [37, 46, 49]

M. longifolia [59] T. sipyleus [55]

Luteolin (32) M. spicata [1] T. herba-barona [68]

M. haplocalyx [1] T. striatus [67]

M. arvensis [1, 41] T. webbianus [52]

M. aquatica [60]

M. x piperita L. [1, 41, 61] T. vulgaris L. [44, 46, 50, 65, 58, 64]

M. longifolia [59] T. serpyllum [37, 46]

M. aquatica [41] T. sipyleus [55]

M. spicata [1]

M. x dalmatica [1]
Luteolin-O-glucoside (33)
M. haplocalyx [1]

M. “Morocco” [41] T. webbianus [52]

M. “Native Wilmet” [41]

M. x verticillata [41]

M. arvensis [1, 41]

T. vulgaris L. [49, 50]

Luteolin-O-rutinoside (34) M. x piperita L. [2, 39, 58, 61-63]
T. serpyllum [49]

M. x piperita L. [2, 39, 58] T. vulgaris L. [35, 49, 50, 57, 64]

Luteolin-O-glucuronide (35) T. serpyllum [37, 49]

M. longifolia [2]
T. sipyleus [55]

M. x piperita L. [2]
Luteolin-O-glucuronide-methyl (36)
M. longifolia [2]

M. x piperita L [2]
Luteolin-O-diglucuronide (37)
M. longifolia [2]

Luteolin-O-diglucoside (38) T. vulgaris L. [64]

Luteolin-acetyl-O-glycoside (39) T. vulgaris L. [64]

Luteolin-7-O-(6”-feruloyl)-ß-glucopyranoside
T. sipyleus [55]
(40)

M. x piperita L. [40] T. vulgaris L. [44, 46, 50]

M. spicata [1] T. serpyllum [37, 46]

M. pulegium [60] T. herba-barona [68]

Apigenin (41)
M. arvensis [1, 41] T. striatus [67]

M. aquatica [60] T.webbianus [52]

M. x piperita L. [60]
Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 389

Table 2. contd…

Compound Mentha Species Thymus Species

[1, 39, 41, 58, 61-

M. x piperita L. T. vulgaris L. [50]
63]

M. aquatica [41]

M. spicata [1]

M. x dalmatica [1]
Apigenin-7-O-rutinoside (42)
M. haplocalyx [1]

M. “Morocco” [41]

M. “Native Wilmet” [41]

M. x verticillata [41]

M. arvensis [1, 41]

T. vulgaris L. [43, 46, 50]

Apigenin-7-O-glucoside (43) T. serpyllum [46]

T. webbianus [52]

T. vulgaris L. [35, 58, 64]

Apigenin-7-O-glucuronide (44)
T. serpyllum [37]

T. vulgaris L. [58]
Apigenin-6,8-di-C-glucoside (45)
T. webbianus [52]

M. spicata [66] T. herba-barona [68]

Thymusin (46)
M. x piperita L. [60, 66] T. striatus [67]

M. spicata [60, 66]

M. x piperita L. [60, 66]

Thymonin (47) M. suaveolens [60] T. striatus [67]

M. pulegium [60]

M. longifolia [60]

M. citrata [66]

Pebrellin (48) M. aquatica [66] T. striatus [67]

M. x piperita L. [60, 61, 66]

M. citrata [66]

Gardenin B (49) M. aquatica [66] T. striatus [67]

M. x piperita L. [60, 61, 66]

M. spicata
Desmethylnobiletin (50) [60, 66] T. striatus [66, 67]
M. x piperita L.

T. vulgaris L. [71]
Cirsilineol (51) M. spicata [66]
T. herba-barona [68]

M. x piperita L. [60]
Sorbifolin (52) T. herba-barona [68]
M. pulegium [60]

M. citrata [60, 66]

Salvigenin (53) M. aquatica [66] T. striatus [67]

M. x piperita L. [60, 66]

390 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

Table 2. contd…

Compound Mentha Species Thymus Species

M. x piperita L. [60, 66]

Ladanein (54) T. striatus [67]
M. pulegium [60]

T. serpyllum [37]

Cirsimaritin (55) T. herba-barona [68]

T. vulgaris L. [50]

T. striatus [67]
Xanthomicrol (56) M. x piperita L. [60]
T. herba-barona [68]

Sideritoflavone (57) M. spicata [66] T. herba-barona [68]

Flavanones

M. x piperita L. [1, 39] T. vulgaris L. [44, 49, 57]

M. “Morocco” [41] T. herba-barona [68]

Eridioctyol (58)
T. serpyllum [37, 49]
M. “Native Wilmet” [41]
T. webbianus [52]

[1, 2, 38, 39, 58,

M. x piperita L. T. vulgaris L. [46, 49, 65]
61, 62, 72]

M. aquatica [41]

M. spicata [1]

M. x dalmatica [1]
Eriocitrin (59)
M. haplocalyx [1]
T. serpyllum [46, 49]
M. “Morocco” [41]

M. “Native Wilmet” [1, 41]

M. x verticillata [41]

M. arvensis var. japanensis [1, 41]

Eridioctyol-O-glucoside (60) M. x piperita L. [39, 61, 63] T. vulgaris L. [35, 49]

T. vulgaris L. [35, 64]

Eridioctyol-O-glucuronide (61)
T. serpyllum [37]

M. x piperita L. [39, 40] T. vulgaris L. [49]

Naringenin (62) T. herba-barona [68]

M. aquatica [19]
T. webbianus [52]

M. x piperita L. [38, 39]

M. haplocalyx [1]
Naringenin-7-O-glucoside (63) T. vulgaris L. [49]
M. x verticillata [41]

M. arvensis [1, 41]

Naringenin-7-O-rutinoside (64) M. x piperita L. [39, 62, 63, 72] T. vulgaris L. [49]

Prenylnaringenin (65) T. serpyllum [37]

M. x piperita L. [39, 58, 61-63, 72]

Hesperidin (66) T. vulgaris L. [49]
M. longifolia [59]
Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 391

Table 2. contd…

Compound Mentha Species Thymus Species

Flavonols and dihydroflavonols

T. vulgaris L. [46, 50]

Quercetin (67) M. x piperita L. [40]
T. serpyllum [46]

Rutin (68) M. x piperita L. [40] T. vulgaris L. [50]

Kaempferol-7-O-rutinoside (69) M. x piperita L. [72]

4’-methoxykaempferol-7-O-rutinoside (70) M. x piperita L. [72]

Quercetin-3-O-hexoside (71) T. vulgaris L. [50]

Isorhamnetin (72) T. vulgaris L. [50]

Isorhamnetin-O-glucoside (73) T. vulgaris L. [50]

T. vulgaris L. [57]
Taxifolin (74)
T. quinquecostatus [54]

Aromadendrin (75) T. quinquecostatus [54]

account for up to 0.12 mg/g of dry plant. Moreover, glycosi- apigenin-7-O-rutinoside (42), was detected in several Men-
dic derivatives of luteolin are often described as major phe- tha species [1, 39, 41, 58, 61-63] with a concentration of
nolic compounds in Mentha. In this sense, Dorman et al. [1] approximately 0.8 mg/g of dry plant in M. x piperita L. [39,
reported the occurrence of approximately 0.1 to 3 mg of 41]. Apigenin has also been reported in Thymus, including T.
luteolin-O-glucoside (33)/g of dry plant [1, 41, 61], while vulgaris L. [44, 46, 50], T. serpyllum [37, 46] and T. webbi-
luteolin-O-rutinoside (34) has been shown to account close anus [52]. These three Thymus species were also described
to 8 mg/g of dry M. x piperita L. [2, 39, 58, 61-63]. Besides to contain mono-O glycosidic forms of apigenin, namely
those compounds, glucuronide luteolin derivatives, namely apigenin-7-O-glucoside (43) [43, 46, 50, 52] and apigenin-7-
the luteolin-O-glucuronide (35), luteolin-O-glucuronide- O-glucuronide (44) [35, 37, 58, 64]. Moreover, apigenin 7-
methyl (36) and luteolin-O-diglucuronide (37) have been O-rutinoside has been described to occur in T. vulgaris L.
[50], while a di-C-glycosidic derivative, the apigenin-6,8-di-
detected in M. piperita L. and in M. longifolia [2, 39, 58]. In
C-glucoside (45), has been detected in the latter species and
a similar way, luteolin has been reported to occur in several
in T. webbianus [52], although no quantification data has
Thymus species, including T. vulgaris L. [44-46, 49, 50, 58], been delivered.
T. serpyllum [37, 49] and T. webbianus [52] and its amount
was established as 0.6 mg/g of dry plant for T. vulgaris L. or Besides the above compounds, Mentha and Thymus
even higher for T. serpyllum (1.5 mg/g of dry plant) [49]. plants were shown to contain several OMe flavones. Thy-
Additionally, luteolin-O-glucoside, luteolin-O-diglucoside musin (46), thymonin (47), pebrellin (48) and gardenin B
(38) or luteolin-acetyl-O-glycoside (39) have been detected (49) are the most cited for Mentha species. They have been
in T. vulgaris L. [43, 44, 46, 50, 52, 55, 58, 64, 65] and the detected in M. spicata, M. x piperita L., M. aquatica, M.
first compound also in wild thyme (T. serpyllum) [37, 46]. citrata, M. longifolia, M. suaveolens and M. pulegium [59-
61, 66]. Some of these OMe flavones and others (com-
Besides these species, T. sipyleus has been described to con-
pounds (46) to (57) in Table 2) have been found in T. stria-
tain luteolin-O-glucoside and luteolin-7-O-(6”-feruloyl)-ß-
tus [67], T. herba-barona [68] and also in other Thymus spe-
glucopyranoside (40) [55]. Moreover, luteolin-O-rutinoside cies [37, 45, 50, 69-71].
is a luteolin glycosidic derivative commonly found in Thy-
mus. Its content in T. serpyllum and T. vulgaris L. has been Flavanones
estimated to be approximately 1.5 and 1.3 mg/g of dry plant,
respectively [49, 50]. On the other hand, glucuronide deriva- As previously mentioned, Mentha species are rich in
tives of luteolin seem to be much abundant in Thymus than flavanones and, according to the reported data in this genus,
in Mentha species [35, 37, 49, 50, 55, 57]. In particular, a compounds of this class enclose mainly derivatives of eri-
odictyol, naringenin and hesperitin, which frequently appear
high concentration of luteolin-O-glucuronide has been found
as O-glucoside derivatives. In this context, eridioctyol (58)
in T. vulgaris L. and T. serpyllum (close to 8 and 14 mg/g of
was reported in M. x piperita L. at concentrations in the
dry plant, respectively) [49].
range of 0.1 - 0.5 mg/g of dry plant and in M. “Native Wil-
Apigenin and its derivatives can also be found in Mentha met” with a concentration of 0.05 mg/g dry plant [1, 39]. On
and Thymus genera. In particular, variable concentrations of the other hand, eriodictyol-7-O-rutinoside (eriocitrin) (59),
apigenin (41) has been described in plant extracts of M. spi- which is the most abundant flavavone in Mentha plants, was
cata (0.01 mg/g of dry plant) and M. arvensis (0.03 mg/g of detected by distinct authors in M.x piperita L. [1, 2, 38, 39,
dry plant) [1, 41]. Moreover, its diglycosidic derivative, the 58, 61, 62, 72] and its content in that species was established
392 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

as approximately 13 - 19 mg/g of dry plant [1, 39]. Other 3. METHODS OF EXTRACTION AND PURIFICA-
studies focusing M. “Native Wilmet”, M. x dalmatica and M. TION
spicata concluded that these species also have considerable
3.1. Sample Preparation
amounts of eriocitrin, although its levels were much lower
than that reported for M. x piperita L. (7.4 mg/g dry plant in As for the majority of reported work on natural products,
M. “Native Wilmet” and 2.5 mg/g dry plant in M. x dal- those focusing on Mentha and Thymus phenolics have entail
matica and M. spicata [1]). regular practices to allow the improvement of the resulting
According to literature data, flavanones in Thymus repre- analytical data. Indeed, in the sample preparation step, the
sent approximately 3-9% of their total polyphenolics [44, 46, material has been commonly dried [34, 42, 43, 47, 48, 57,
49], which mostly appear as eriodictyol and as its deriva- 58], lyophilized [34, 64] or frozen ideally at -80ºC [45], thus
tives. In fact, eriodictyol has been described to occur in T. minoring the instability of polyphenols and the action of
vulgaris L. (1.5 mg/ g of dry plant) [44, 49, 57], in T. serpyl- several degradative enzymes. In addition, grounding is a
lum [37, 49], in T. webbianus [52] and in T. herba-barona well establish procedure before the extraction step [2, 34, 42,
[68], and eriodictyol-O-glucoside (60) and eriodictyol-7-O- 48, 50, 53, 61, 63], as the particle size reducement of the
rutinoside were both found in T. vulgaris L. (0.6 mg/g and plants increases the yield of extraction. Note that authors
1.2 mg/ g of dry plant, respectively). The latter has also been occasionally defatted the plant material with apolar solvents
described for T. serpyllum (1.2 mg/ g of dry plant) [46, 49]. (e.g. n-hexane) [44, 62, 71], for prevention of high levels of
Note that eriodictyol-O-glucuronide (61) has been found in lipophilic compounds in the extracts which can interfere in
thyme and wild thyme (Table 2), but no quantitative infor- the polyphenols analysis. Some authors have also performed
mation has been delivered on that compound [35, 37, 64]. acidic or enzymatic hydrolysis. These procedures have been
done before or simultaneously to the extraction procedure,
Naringenin (62) has been detected as a minor phenolic when only aglycones were intended to characterize [36, 40,
component in M. x pipperita L. [40] and in M. aquatica [19] 44, 48].
while its 7-O-glucoside (63) was found in M. x piperita L.
(1.0 mg/g of dry plant) [38, 39] and in M. arvensis (0.1 mg/g 3.2. Extraction
of dry plant), among others [1, 41]. The major naringenin
derivative described in this genus is narirutin (naringenin-7- Polyphenols have been mainly obtained by solvent ex-
O-rutinoside) (64) [39, 62, 63, 72]. traction. Aqueous mixtures of methanol or ethanol are the
most used ones, though pure water is also often applied. In
Despite less frequently described, naringenin derivatives fact, as the majority of polyphenolics in Mentha and Thymus
have also been reported in Thymus. Naringenin and prenyl- occur in glycosidic forms, aqueous mixtures are preferred.
naringenin (65) were the only naringenin derivatives de- Distinct species of these two genera have been extracted
tected in T. webbianus [52] and in T. serpyllum [37] respec- with hydromethanolic solutions of 60 - 80% (v/v) to obtain
tively, while significant amounts of this aglycone, of narin- phenolic acids [34, 42, 53], as well as this group combined
genin-7-O-glucoside and of naringenin-7-O-rutinoside have with flavones, flavanones and flavonols [2, 35-37, 40, 43,
been reported for T. vulgaris L. (approximately 0.4, 0.6 and 50, 53, 54, 58, 61, 64, 72, 73]. In a similar way, hydroetha-
0.3 mg/ g of dry plant, respectively [49]). nolic solutions of 80% (v/v) have been preferentially used
To our knowledge, hesperitin aglycone has not been de- for extracting phenolic acids or flavonoids in Mentha [40,
tected so far in Mentha, but its 7-O-rutinoside derivative, the 53, 58, 61, 72] or Thymus [53, 57] species. According to the
hesperidin (66), has been found in significant amounts (1.7 study of Reichling et al. [58], this mixture was much effi-
mg/g dry plant) in M. x piperita L. and also detected in M. cient than that of water:ethanol (80:20) in recovering rosma-
longifolia [58, 59, 61, 63, 72]. In Thymus plants, hesperidin rinic acid from four Mentha species, including M. x pip-
has been described in T. vulgaris L., accounting for 1 mg/g perita L. and T. vulgaris L.. Besides the above solvents, wa-
of dry plant [49]. ter [39, 46, 62], acetone [45, 56, 71], aqueous acetone mix-
tures [56] and diethyl ether [68-70] have been used.
Flavonols and Dihydroflavonols Authors have applied different techniques in the extrac-
Despite less mentioned, flavonols have been reported to tion process of Mentha or Thymus polyphenols. Stirring [35,
be minor phenolic components of some Mentha and Thymus 61], homogenization using a tissue homogenizer [43, 45,
species. In particular, M. x pipperita L. has been described to 50], maceration [40], sonication [47, 53, 72, 73] were the
contain quercetin (67), rutin (68), kaempferol-7-O-rutinoside most frequently applied. Commonly, these techniques have
(69) and its 4’-methoxy derivative (70) [40, 72]. Quercetin been performed at room temperature [43, 50, 56, 58], to
has also been described to occur in several Thymus species, minimize degradation of phenolic compounds [74]. Water
including the T. vulgaris L. [46, 50]. This latter species also extraction is the main exception, since authors frequently
contains two quercetin glycosides (quercetin-3-O-hexoside have applied boiling or refluxing [39, 46, 62].
(71) and rutin), the methylated flavonol isorhamnetin (72),
its glucoside (73) [50] and the dihydroflavonol taxifolin (74) 3.3. Clean-up and Fractionation
[57]. The latter phenolic has been also detected in T. quin- The main extraction process can be followed by addi-
quecostatus, together with aromadendrin (dihydro- tional purification of the enriched phenolic extracts. This
kaempferol) (75) [50, 54] and, besides these two dihydrofla- practice allows obtaining a cleaner sample for characteriza-
vonols, others have been described to occur in several spe- tion or to be used in biological assays. Reports on Mentha
cies of Thymus [70]. and Thymus genera have applied liquid-liquid extraction [55]
Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 393

and, most commonly, solid phase extraction on C18 car- Krzyzanowska and colleagues [2] for structural determina-
tridges or column chromatography on Sephadex LH-20. The tion of phenolic acids, flavones and flavanones in two spe-
two latter usually enclose sequential solubilisation with dis- cies of Mentha [2, 73]. Since these procedures entails a long
tinct solvents, according to the nature of compounds that are time of analysis, the present implementation of faster and
intended to separate [39, 42, 44, 56, 57, 62, 68, 75, 76]. reliable analytical methodologies, as e.g. the chroma-
tographic techniques hyphenated with mass spectrometry
4. ANALYTICAL TECNHIQUES appears as a good alternative. On-line LC–MS/MS analysis
As known, HPLC is the main technique applied in the has been used in the identification of phenolic acids and dis-
analysis of plant phenolics, since it allows a rapid qualitative tinct classes of flavonoids in M. pipperita L. [72], as well as
and quantitative screening [76]. In fact, the HPLC analysis in several Thymus species [35, 37, 50, 70], between others.
of the Mentha and Thymus plant extracts have been essen- In the majority of these studies, mass spectrometry analysis
tially carried out in C18 reversed-phase columns. Addition- was performed using electrospray ionization (ESI), a soft
ally, in order to control the reproducibility of the method, the mode of ionization that is suitable for structural characteriza-
temperature of the column is usually maintained constant tion of a high number of polar biomolecules, in particular the
(20-30ºC). phenolic compounds [35, 43, 72]. Moreover, the mass spec-
trometry analysis has been mainly carried out in the negative
Other important feature for achieving a good separation ion mode, due to its high sensitivity in detecting distinct
of phenolic constituents and consequently, high accuracy in classes of phenolic compounds [79].
the method, is the choice of the mobile phase. Distinct com-
binations of mobile and stationary phases provide diferent Besides mass spectrometry, NMR spectroscopy has
separation, since this is based on polarity differences among played an important role in structural analysis of polyphe-
polyphenols [77]. For e.g., studies on Mentha and Thymus nols in Mentha [39, 56, 60, 62, 66] and Thymus [54, 55, 57,
species applied acetonitrile/water or methanol/water combi- 68, 71] plants, coupled with other techniques as LC or MS.
nations in an isocratic mode for fractionation of phenolic The main drawback in NMR is its low sensibility when
acids [42]. However, in general, Mentha and Thymus poly- compared to MS and thus, there is the need of getting higher
phenols have been preferentially analysed in a binary system amounts of sample for analysis [80]. In this sense, when us-
of solvents, such as acetonitrile/water [37, 39, 43, 45, 48, 49, ing NMR technique, samples need to be obtained by prepa-
58, 61, 63], methanol/water [34, 35, 40, 42, 61, 73] or wa- rative chromatography, as done for Mentha [51, 62] and
ter/water-acetonitrile [2]. Note that acidified water (0.1% to Thymus [57]. A good alternative is the coupling of HPLC
5% of formic acid or other acids such as phosphoric acid or with RMN techniques (LC-NMR) that actually appears as
acetic acid) is preferentially used, as this procedure impairs the most powerful method for the separation and structural
analytes ionization and thus allows a better resolution and determination of organic compounds. Regardless of its effi-
superior reproducibility of the retention times, as well as the ciency for identification of on the nature of the polyphenol
minimization of peak tailing [76-78]. skeletons and on their substitution patterns, the method is not
widely used at present due the high entailed costs [77].
As commonly, the HPLC separation of Mentha and Thy-
mus polyphenols has been achieved at constant flow rates of 5. CONCLUDING REMARKS
approximately 1mL/min and their identification and quanti-
fication has been frequently done by comparison of the re- Mentha and Thymus species have been mostly studied for
tention times and integrated peak areas of the separated their oil composition. However, the close association of
compounds, to those of the corresponding reference com- some of their beneficial properties with their content in phe-
pound [34, 38-40, 43, 45, 46]. This information has also nolic compounds has encouraged the search on these latter
been combined to spectral information gathered by photodi- metabolites. So far, the reported data allows us to conclude
ode array detector (PDA) [1, 2, 34, 37, 40, 43, 52, 69, 70]. that plants of Mentha and Thymus genera are mostly rich in
Spectral data in those studies has been obtained in the range caffeic acid derivatives. Thymus plants are also particularly
of 200 to 450 nm, while the chromatograms have been plot- rich in glycosidic forms of the flavones luteolin and apigenin
ted according to their maximum absorbance peaks at 280 nm while glycosidic derivatives of eriodictyol and naringenin
for flavanones and hydroxybenzoic acids, at 320-330 nm for are particularly abundant in Mentha species. On the contrary,
hydroxycinnamic acids and flavones and at 350-370 nm for flavonols and dihydroflavonol are less described in these
flavonols [34, 40, 45, 48]. Alternatively, in case of exclusive genera. Despite the considerable number of studies focusing
usage of UV-Vis detector, the polyphenolic profiles were on Mentha and Thymus polyphenols, it must be noted that to
only recorded at a wavelength of 280 nm [36, 39, 42, 49, 54, date, only one third of Mentha species have been studied and
63]. a diminutive number of Thymus (less than 5% of total known
species) have been investigated. Thus besides summarizing
However, due to commercial unavailability of many
the existing information on this theme, we hope that the pre-
phenolic plant constituents, fine analytical techniques have
sent manuscript also encourages the search of the missing
also been implemented in order to improve the phenolic
data on these genera.
characterization. In this field, mass spectrometry has been
playing a crucial role, as its coupling to chromatographic
CONFLICT OF INTEREST
analysis allowed an increment on the sensitivity and selectiv-
ity of the method. HPLC fractionation combined with elec- The author(s) confirm that this article content has no con-
trospray ionization-MS/MS analyses have been used e.g. by flict of interest.
394 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

ACKNOWLEDGEMENTS Neurochemical Properties of Mint Extracts. Phytotherapy

Research, 2010, 24(6), 869-874.
The authors acknowledge the financial support provided [19] Jaeger, A. K.; Almqvist, J. P.; Vangsoe, S. A. K.; Stafford, G. I.;
by the FCT to CERNAS (project PEst-OE/AGR/UI0681/ Adsersen, A.; Van Staden, J. Compounds from Mentha aquatica
with affinity to the GABA-benzodiazepine receptor. South African
2011) and to CIMO (PEst-OE/AGR/UI0690/2011). Olívia R. Journal of Botany, 2007, 73(4), 518-521.
Pereira was supported by a PhD grant (SFRH/PROTEC/ [20] Olsen, H. T.; Stafford, G. I.; van Staden, J.; Christensen, S. B.;
49600/2009). Jaeger, A. K. Isolation of the MAO-inhibitor naringenin from
Mentha aquatica L. Journal of Ethnopharmacology, 2008, 117(3),
500-502.
REFERENCES [21] Conforti, F.; Sosa, S.; Marrelli, M.; Menichini, F.; Statti, G. A.;
[1] Dorman, H. J. D.; Kosar, M.; Kahlos, K.; Holm, Y.; Hiltunen, R. Uzunov, D.; Tubaro, A.; Menichini, F.; Della Loggia, R. In vivo
Antioxidant properties and composition of aqueous extracts from anti-inflammatory and in vitro antioxidant activities of
Mentha species, hybrids, varieties, and cultivars. Journal of Mediterranean dietary plants. Journal of Ethnopharmacology,
Agricultural and Food Chemistry, 2003, 51(16), 4563-4569. 2008, 116(1), 144-151.
[2] Krzyzanowska, J.; Janda, B.; Pecio, L.; Stochmal, A.; Oleszek, W.; [22] Caillet, S.; Lessard, S.; Lamoureux, G.; Lacroix, M. Umu test
Czubacka, A.; Przybys, M.; Doroszewska, T. Determination of applied for screening natural antimutagenic agents. Food Chem.,
Polyphenols in Mentha longifolia and M. piperita Field-Grown and 2011, 124(4), 1699-1707.
In Vitro Plant Samples Using UPLC-TQ-MS. Journal of Aoac [23] Amarowicz, R.; Zegarska, Z.; Rafalowski, R.; Pegg, R. B.;
International, 2011, 94(1), 43-50. Karamac, M.; Kosinska, A. Antioxidant activity and free radical-
[3] Brickell, C.; Trevor, C. The American Horticultural Society: scavenging capacity of ethanolic extracts of thyme, oregano, and
Encyclopedia of Plants & Flowers. USA: DK Inc.: New York, marjoram. European Journal of Lipid Science and Technology,
2002; p p. 605. 2009, 111(11), 1111-1117.
[4] Aflatuni, A.; Uusitalo, J.; Ek, S.; Hohtola, A. Variation in the [24] Gião, M. S.; Gonzalez-Sanjose, M. L.; Rivero-Perez, M. D.;
amount of yield and in the extract composition between Pereira, C. I.; Pintado, M. E.; Malcata, F. X. Infusions of
conventionally produced and micropropagated peppermint and Portuguese medicinal plants: Dependence of final antioxidant
spearmint. J. Essent. Oil Res., 2005, 17(1), 66-70. capacity and phenol content on extraction features. J. Sci. Food
[5] Brickell, C.; Zuk, J. D. The American Horticultural Society: A-Z Agric., 2007, 87(14), 2638-2647.
Encyclopedia of Garden Plants. USA: DK Publishing: New York, [25] Albano, S. M.; Miguel, M. G. Biological activities of extracts of
NY, 1997; p p. 668. plants grown in Portugal. Industrial Crops and Products, 2011,
[6] Rita, P.; Animesh, D. K. An update overview on peppermint 33(2), 338-343.
(Mentha piperita L.). International Research Journal of Pharmacy, [26] Bueyuekbalci, A.; El, S. N. Determination of In vitro antidiabetic
2011, 8 (2), 1-10. effects, antioxidant activities and phenol contents of some herbal
[7] Hayes, J. R.; Stavanja, M. S.; Lawrence, B. M. Mint. The Genus teas. Plant Food Hum. Nutr., 2008, 63(1), 27-33.
Mentha. CRC Press Taylor and Francis: Boca Raton, 2007. [27] Elhabazi, K.; Ouacherif, A.; Laroubi, A.; Aboufatima, R.; Abbad,
[8] Reddy, M. V. B.; Angers, P.; Gosselin, A.; Arul, J. A.; Benharref, A.; Zyad, A.; Chait, A.; Dalal, A. Analgesic activity
Characterization and use of essential oil from Thymus vulgaris of three thyme species, Thymus satureioides, Thymus maroccanus
against Botrytis cinerea and Rhizopus stolonifer in strawberry and Thymus leptobotrys. Afr. J. Microbiol. Res., 2008, 2(10), 262-
fruits. Phytochemistry, 1998, 47(8), 1515-1520. 267.
[9] Elisabeth Stahl-Biskup, F. S. Thyme, the genus Thymus. Taylor and [28] Grande, S.; Bogani, P.; De Saizieu, A.; Schueler, G.; Galli, C.;
Francis: London, 2002; p 317. Visioli, F. Vasomodulating potential of Mediterranean wild plant
[10] Omidbaigi, R.; Sefidkon, F.; Hejazi, M. Essential oil composition extracts. Journal of Agricultural and Food Chemistry, 2004,
of Thymus citriodorus L. cultivated in Iran. Flavour Frag. J., 2005, 52(16), 5021-5026.
20(2), 237-238. [29] Cakilcioglu, U.; Khatun, S.; Turkoglu, I.; Hayta, S.
[11] Horvath, G.; Szabo, L. G.; Hethelyi, E.; Lemberkovics, E. Essential Ethnopharmacological survey of medicinal plants in Maden
oil composition of three cultivated Thymus chemotypes from (Elazig-Turkey). Journal of Ethnopharmacology, 2011, 137(1),
Hungary. J. Essent. Oil Res., 2006, 18(3), 315-317. 469-486.
[12] Figueiredo, A. C.; Barroso, J. G.; Pedro, L. G.; Salgueiro, L.; [30] Fernandes, A. S. F.; Barros, L.; Carvalho, A. M.; Ferreira, I.
Miguel, M. G.; Faleiro, M. L. Portuguese Thymbra and Thymus Lipophilic and hydrophilic antioxidants, lipid peroxidation
Species Volatiles: Chemical Composition and Biological inhibition and radical scavenging activity of two Lamiaceae food
Activities. Curr. Pharm. Design, 2008, 14(29), 3120-3140. plants. European Journal of Lipid Science and Technology, 2010,
[13] Mata, A. T.; Proenca, C.; Ferreira, A. R.; Serralheiro, M. L. M.; 112(10), 1115-1121.
Nogueira, J. M. F.; Araujo, M. E. M. Antioxidant and [31] Berdowska, I.; Marcinkowska, A.; Zielinski, B.; Fecka, I.; Banas,
antiacetylcholinesterase activities of five plants used as Portuguese T. The effect of selected herb extracts on superoxide dismutase
food spices. Food Chem., 2007, 103(3), 778-786. activity in Jurkat cells. Adv. Clin. Exp. Med., 2007, 16(3), 361-364.
[14] Lopez, V.; Akerreta, S.; Casanova, E.; Maria Garcia-Mina, J.; [32] Robards, K. Strategies for the determination of bioactive phenols in
Yolanda Cavero, R.; Isabel Calvo, M. In vitro antioxidant and anti- plants, fruit and vegetables. Journal of Chromatography A, 2003,
rhizopus activities of lamiaceae herbal extracts. Plant Foods for 1000(1-2), 657-691.
Human Nutrition, 2007, 62(4), 151-155. [33] Carrasco-Pancorbo, A.; Cerretani, L.; Bendini, A.; Segura-
[15] Arumugam, P.; Ramamurthy, P.; Santhiya, S. T.; Ramesh, A. Carretero, A.; Gallina-Toschi, T.; Fernandez-Gutierrez, A.
Antioxidant activity measured in different solvent fractions Analytical determination of polyphenols in olive oils. Journal of
obtained from Mentha spicata Linn.: An analysis by ABTS(.+) Separation Science, 2005, 28(9-10), 837-858.
decolorization assay. Asia Pacific Journal of Clinical Nutrition, [34] Shan, B.; Cai, Y. Z.; Sun, M.; Corke, H. Antioxidant capacity of 26
2006, 15(1), 119-124. spice extracts and characterization of their phenolic constituents.
[16] Yi, W. G.; Wetzstein, H. Y. Anti-tumorigenic activity of five Journal of Agricultural and Food Chemistry, 2005, 53(20), 7749-
culinary and medicinal herbs grown under greenhouse conditions 7759.
and their combination effects. Journal of the Science of Food and [35] Nagy, T. O.; Solar, S.; Sontag, G.; Koenig, J. Identification of
Agriculture, 2011, 91(10), 1849-1854. phenolic components in dried spices and influence of irradiation.
[17] Arumugam, P.; Ramamurthy, P.; Ramesh, A. Antioxidant and Food Chem., 2011, 128(2), 530-534.
Cytotoxic Activities of Lipophilic and Hydrophilic Fractions of [36] Proestos, C.; Chorianopoulos, N.; Nychas, G. J. E.; Komaitis, M.
Mentha Spicata L. (Lamiaceae). Int. J. Food Prop., 2010, 13(1), RP-HPLC analysis of the phenolic compounds of plant extracts.
23-31. Investigation of their antioxidant capacity and antimicrobial
[18] Lopez, V.; Martin, S.; Pilar Gomez-Serranillos, M.; Emilia activity. Journal of Agricultural and Food Chemistry, 2005, 53(4),
Carretero, M.; Jager, A. K.; Isabel Calvo, M. Neuroprotective and 1190-1195.
[37] Miron, T. L.; Plaza, M.; Bahrim, G.; Ibanez, E.; Herrero, M.
Chemical composition of bioactive pressurized extracts of
Overview on Mentha and Thymus Polyphenols Current Analytical Chemistry, 2013, Vol. 9, No. 3 395

Romanian aromatic plants. Journal of Chromatography A, 2011, [57] Dapkevicius, A.; van Beek, T. A.; Lelyveld, G. P.; van Veldhuizen,
1218(30), 4918-4927. A.; de Groot, A.; Linssen, J. P. H.; Venskutonis, R. Isolation and
[38] Dorman, H. J. D.; Kosar, M.; Baser, K. H. C.; Hiltunen, R. structure elucidation of radical scavengers from Thymus vulgaris
Phenolic Profile and Antioxidant Evaluation of Mentha x piperita leaves. Journal of Natural Products, 2002, 65(6), 892-896.
L., (Peppermint) Extracts. Nat. Prod. Commun., 2009, 4(4), 535- [58] Reichling, J.; Nolkemper, S.; Stintzing, F. C.; Schnitzler, P. Impact
542. of Ethanolic Lamiaceae Extracts on Herpesvirus Infectivity in Cell
[39] Fecka, I.; Turek, S. Determination of water-soluble polyphenolic Culture. Forschende Komplementarmedizin, 2008, 15(6), 313-320.
compounds in commercial herbal teas from Lamiaceae: [59] Ghoulami, S.; Idrissi, A.; Fkih-Tetouani, S. Phytochemical study of
Peppermint, melissa, and sage. Journal of Agricultural and Food Mentha longifolia of Morocco. Fitoterapia, 2001, 72(5), 596-598.
Chemistry, 2007, 55(26), 10908-10917. [60] Zaidi, F.; Voirin, B.; Jay, M.; Viricel, M. R. Free flavonoid
[40] Misan, A. C.; Mimica-Dukic, N. M.; Mandic, A. I.; Sakac, M. B.; aglycones from leaves of Mentha pulegium and Mentha suaveolens
Milovanovic, I. L.; Sedej, I. J. Development of a rapid resolution (Labiatae). Phytochemistry, 1998, 48(6), 991-994.
HPLC method for the separation and determination of 17 phenolic [61] Areias, F. M.; Valentao, P.; Andrade, P. B.; Ferreres, F.; Seabra, R.
compounds in crude plant extracts. Cent. Eur. J. Chem, 2011, 9(1), M. Phenolic fingerprint of peppermint leaves. Food Chem., 2001,
133-142. 73(3), 307-311.
[41] Kosar, M.; Dorman, H. J. D.; Baser, K. H. C.; Hiltunen, R. [62] Inoue, T.; Sugimoto, Y.; Masuda, H.; Kamei, C. Antiallergic effect
Screening of free radical scavenging compounds in water extracts of flavonoid glycosides obtained from Mentha piperita L.
of Mentha samples using a postcolumn derivatization method. Biological & Pharmaceutical Bulletin, 2002, 25(2), 256-259.
Journal of Agricultural and Food Chemistry, 2004, 52(16), 5004- [63] Guedon, D. J.; Pasquier, B. P. Analysis and distribution of
5010. flavonoid glycosides and rosmarinic acid in 40 Mentha x piperita
[42] Zgorka, G.; Glowniak, K. Variation of free phenolic acids in clones. Journal of Agricultural and Food Chemistry, 1994, 42(3),
medicinal plants belonging to the Lamiaceae family. Journal of 679-684.
Pharmaceutical and Biomedical Analysis, 2001, 26(1), 79-87. [64] Justesen, U. Negative atmospheric pressure chemical ionisation
[43] Hossain, M. B.; Patras, A.; Barry-Ryan, C.; Martin-Diana, A. B.; low-energy collision activation mass spectrometry for the
Brunton, N. P. Application of principal component and hierarchical characterisation of flavonoids in extracts of fresh herbs. Journal of
cluster analysis to classify different spices based on in vitro Chromatography A, 2000, 902(2), 369-379.
antioxidant activity and individual polyphenolic antioxidant [65] Wang, M.; Li, J.; Ho, G. S.; Peng, X.; Ho, C. T. Isolation and
compounds. J. Funct. Food., 2011, 3(3), 179-189. identification of antioxidative flavonoid glycosides from thyme
[44] Kosar, M.; Dorman, H. J. D.; Hiltunen, R. Effect of an acid (Thymus vulgaris L.). Journal of Food Lipids, 1998, 5(4), 313-321.
treatment on the phytochemical and antioxidant characteristics of [66] Voirin, B.; Bayet, C.; Faure, O.; Jullien, F. Free flavonoid
extracts from selected Lamiaceae species. Food Chem., 2005, aglycones as markers of parentage in Mentha aquatica, M. citrata,
91(3), 525-533. M. spicata and M. x piperita. Phytochemistry, 1999, 50(7), 1189-
[45] Zheng, W.; Wang, S. Y. Antioxidant activity and phenolic 1193.
compounds in selected herbs. Journal of Agricultural and Food [67] Marin, P. D.; Grayer, R. J.; Kite, G. C.; Veljic, M. External
Chemistry, 2001, 49(11), 5165-5170. flavones from Thymus striatus Vahl (Lamiaceae). Biochemical
[46] Kulisic, T.; Dragovic-Uzelac, V.; Milos, M. Antioxidant activity of Systematics and Ecology, 2005, 33(11), 1179-1182.
aqueous tea infusions prepared from oregano, thyme and wild [68] Corticchiato, M.; Bernardini, A.; Costa, J.; Bayet, C.; Saunois, A.;
thyme. Food Technology and Biotechnology, 2006, 44(4), 485-492. Voirin, B. Free flavonoid aglycones from Thymus-herba-barona
[47] Janicsak, G.; Mathe, I.; Miklossy-Vari, V.; Blunden, G. and its monoterpenoid chemotypes. Phytochemistry, 1995, 40(1),
Comparative studies of the rosmarinic and caffeic acid contents of 115-120.
Lamiaceae species. Biochemical Systematics and Ecology, 1999, [69] Marin, P. D.; Grayer, R. J.; Kite, G. C.; Matevski, V. External leaf
27(7), 733-738. flavonoids of Thymus species from Macedonia. Biochemical
[48] Wojdylo, A.; Oszmianski, J.; Czemerys, R. Antioxidant activity Systematics and Ecology, 2003, 31(11), 1291-1307.
and phenolic compounds in 32 selected herbs. Food Chem., 2007, [70] Horwath, A. B.; Grayer, R. J.; Keith-Lucas, D. M.; Simmonds, M.
105(3), 940-949. S. J. Chemical characterisation of wild populations of Thymus
[49] Fecka, I.; Turek, S. Determination of polyphenolic compounds in from different climatic regions in southeast Spain. Biochemical
commercial herbal drugs and spices from Lamiaceae: thyme, wild Systematics and Ecology, 2008, 36(2), 117-133.
thyme and sweet marjoram by chromatographic techniques. Food [71] Miura, K.; Nakatani, N. Antioxidative activity of flavonoids from
Chem., 2008, 108(3), 1039-1053. thyme (Thymus-vulgaris L). Agricultural and Biological
[50] Hossain, M. B.; Rai, D. K.; Brunton, N. P.; Martin-Diana, A. B.; Chemistry, 1989, 53(11), 3043-3045.
Barry-Ryan, C. Characterization of Phenolic Composition in [72] Dolzhenko, Y.; Bertea, C. M.; Occhipinti, A.; Bossi, S.; Maffei, M.
Lamiaceae Spices by LC-ESI-MS/MS. Journal of Agricultural and E. UV-B modulates the interplay between terpenoids and
Food Chemistry, 2010, 58(19), 10576-10581. flavonoids in peppermint (Mentha x piperita L.). J. Photochem.
[51] Grayer, R. J.; Eckert, M. R.; Veitch, N. C.; Kite, G. C.; Marin, P. Photobiol. B-Biol., 2010, 100(2), 67-75.
D.; Kokubun, T.; Simmonds, M. S. J.; Paton, A. J. The [73] Oinonen, P. P.; Jokela, J. K.; Hatakka, A. I.; Vuorela, P. M.
chemotaxonomic significance of two bioactive caffeic acid esters, Linarin, a selective acetylcholinesterase inhibitor from Mentha
nepetoidins A and B, in the Lamiaceae. Phytochemistry, 2003, arvensis. Fitoterapia, 2006, 77(6), 429-434.
64(2), 519-528. [74] Akowuah, G. A.; Zhari, I. Effect of Extraction Temperature on
[52] Blazquez, M. A.; Manez, S.; Zafrapolo, M. C. Further flavonoids Stability of Major Polyphenols and Antioxidant Activity of
and other phenolics of Thymus-webbianus rouy. Z. Naturforsch. Orthosiphon stamineus Leaf. Journal of Herbs, Spices & Medicinal
(C), 1994, 49(9-10), 687-688. Plants, 2010, 16(3-4), 160-166.
[53] Wang, H.; Provan, G. J.; Helliwell, K. Determination of rosmarinic [75] Ozgen, U.; Mavi, A.; Terzi, Z.; Yildirim, A.; Coskun, M.;
acid and caffeic acid in aromatic herbs by HPLC. Food Chem., Houghton, P. J. Antioxidant properties of some medicinal
2004, 87(2), 307-311. Lamiaceae (Labiatae) species. Pharmaceutical Biology, 2006,
[54] Lee, I. C.; Bae, J. S.; Kim, T.; Kwon, O. J.; Kim, T. H. 44(2), 107-112.
Polyphenolic Constituents from the Aerial Parts of Thymus [76] Dai, J.; Mumper, R. J. Plant Phenolics: Extraction, Analysis and
quinquecostatus var. japonica Collected on Ulleung Island. J. Their Antioxidant and Anticancer Properties. Molecules, 2010,
Korean Soc. Appl. Biol. Chem., 2011, 54(5), 811-816. 15(10), 7313-7352.
[55] Ozgen, U.; Mavi, A.; Terzi, Z.; Kazaz, C.; Asci, A.; Kaya, Y.; [77] Stalikas, C. D. Extraction, separation, and detection methods for
Secen, H. Relationship Between Chemical Structure and phenolic acids and flavonoids. Journal of Separation Science,
Antioxidant Activity of Luteolin and Its Glycosides Isolated from 2007, 30(18), 3268-3295.
Thymus sipyleus subsp sipyleus var. sipyleus. Rec. Nat. Prod., [78] Mahugo Santana, C.; Sosa Ferrera, Z.; Esther Torres PadrÃn, M.;
2011, 5(1), 12-21. Juan Santana RodrÃ-guez, J. Methodologies for the Extraction of
[56] She, G. M.; Xu, C.; Liu, B.; Shi, R. B. Polyphenolic Acids from Phenolic Compounds from Environmental Samples: New
Mint (the Aerial of Mentha haplocalyx Briq.) with DPPH Radical Approaches. Molecules, 2009, 14(1), 298-320.
Scavenging Activity. J. Food Sci., 2010, 75(4), C359-C362.
396 Current Analytical Chemistry, 2013, Vol. 9, No. 3 Pereira and Cardoso

[79] Cuyckens, H.; Claeys, M. Mass spectrometry in the structural [80] Maul, R.; Schebb, N.; Kulling, S. Application of LC and GC
analysis of flavonoids. Journal of Mass Spectrometry, 2004, 39(4), hyphenated with mass spectrometry as tool for characterization of
1-15. unknown derivatives of isoflavonoids. Analytical and Bioanalytical
Chemistry, 2008, 391(1), 239-250.

Received: February 16, 2011 Revised: March 11, 2011 Accepted: March 12, 2011