Comparative Study of The Chemical Composition and Pharmacological Activity of Three Chemotypes of Peppermint

Strelyaeva A.V.¹, Kartashova N.V.¹, Karamova K.E.¹, Vaskova L.B.¹,

Fedorova L.V.¹, Bobkova N.V¹. Bondar A.A.¹, Davosyr D.P.²,

Lazareva Y.B.¹, Kuznetsov R.M.¹

¹Department of Pharmacy, Faculty of Pharmaceutical Natural Science, I.M. Sechenov First Moscow State Medical University (Sechenov University), 119991, Trubetskaya St. 8, bldg. 2, Moscow,

Russian Federation, Russia.

²Department of Chemistry, Faculty of Medicine and Biology, Pirogov Russian National Research Medical University, 117997, Ostrovitanova 1, Moscow, Russian Federation, Russia.

*Corresponding Author E-mail: kkristina_02@mail.ru

ABSTRACT:

The aim of the work was to study the composition of the alcohol extract, antioxidant activity and pharmacological properties of peppermint leaves of three forms - menthol, linalool and carvone. Materials and methods: Chromatography-mass-spectrometric analysis was performed on an Agilent Technologies device. For the model evaluation of antioxidant activity, the aerometric method for measuring the total antioxidant activity was used, implemented in the TsvetYauza-01-AA device. Tonic activity was studied in terms of the duration of swimming of laboratory animal (male mice) with a load of 7% of the animal's body weight until complete fatigue - "The mouse forced swim test". The discussion of the results. When studying the chemical composition of medicinal plant raw materials of peppermint leaves of menthol, linalool, carvone types, more than 50 compounds were identified. A comparative analysis showed that the content of menthol in peppermint leaves of the menthol type can exceed the content of menthol in other chemotypes by ten or more times. At the same time, the tonic effect of an aqueous extract from peppermint leaves is mostly associated with the menthol. It should be taken into account when using this type of raw material in the manufacturing. Findings. The raw material of peppermint leaves may have a different degree of pharmacological activity depending on the chemotype, which should be taken into consideration when using this type of raw material.

KEYWORDS: Peppermint of menthol, Linalool, Carvone types, Tonic activity.

INTRODUCTION:

Peppermint (Mentha piperita L.) belongs to the Lamiaceae family, is a hybrid of water mint and spearmint (Mentha aquatica x Mentha spicata) and is a cultivated perennial herbaceous plant 30–100 cm high¹. There is a lot of drugs used both externally and internally as anti-inflammatory, antispasmodic, choleretic medicine, including peppermint essential oil or menthol². Peppermint essential oil pharmacological activity study proved antimicrobial, antispasmodic, tonic and analgesic effect.^3,4,5,6,7

At the moment, new drugs and compositions are offered that contain both peppermint leaves and essential oil. ^8,9,10 The effect of peppermint on the central nervous system is especially noted. Several authors note both sedative and tonic effects of peppermint essential oil. ^11,12,13

However, the plant species Mentha piperita L. has many chemotype forms. For example, morphotypes such as menthol, linalool, and carvone.^14,15 The chemical composition of raw materials and the quantitative content of essential oil can vary greatly, not only depending on the chemotype, but also on the variety and method of plant introduction.^14,16,17

The aim of the work was to study the component composition of the alcohol extract, antioxidant activity and pharmacological properties of three introduced forms - menthol, linalool and carvone - of the species M. piperita L., cultivated in the Main Botanical Garden named after. N.V. Tsitsina RAS.

MATERIALS AND METHODS:

Chromato-mass spectrometric analysis was performed on an Agilent Technologies instrument consisting of a 7890 gas chromatograph (HP-5 column, 50 m x 320 µm x 1.05 µm) and a 5975 C mass selective detector with a quadrupole mass analyzer; chromatogram of samples - by total ion current; software - ChemStation E 02.00^18-22. The identification of the component composition (qualitative analysis) was carried out using the NIST-05 library of complete mass spectra and the corresponding values of the Kovacs chromatographic indices. The relative content (%) of the components of the mixture (quantitative analysis) was determined by calculating the ratio of the areas of chromatographic peaks (simple normalization method).

For the model evaluation of antioxidant activity, the amperometric method for measuring the total content of antioxidants, implemented in the TsvetYauza-01-AA device was used . For research, samples of dried plants were crushed to a particle size of 1–2 mm, and a sample of 1.0 g was poured with 96% ethanol, distilled water, or their mixture with a volume ratio of 70:30. The volume of the extractant was 100 ml, the temperature was 200C and 950C for water; 200C and 750- for ethanol; 200C and 850 - for a mixture of ethanol-water. The extraction was carried out for 10 minutes without temperature control, and the resulting solutions were quickly filtered through a blue-ribbon paper filter. The tonic activity of an aqueous extract from dried peppermint leaves of different chemotypes was studied in terms of the duration of swimming of animals with a load of 7% of the animal's body weight until complete fatigue - "The mouse forced swim test".

RESULTS AND DISCUSSION:

More than 50 components were identified in alcohol extracts obtained from dried peppermint leaves. The alcoholic extract of peppermint of menthol, linalool, and carvone chemotypes contains predominantly compounds belonging to the biosynthetic types of thuyane (α-thuyene, sabinene, 5-isopropyl-2-methyl-bicyclo[3,1]hexane-2- ol); fenhana (α-fenchen); pinane (α- and β-pinene, trans-pinocarveol, myrtenal and myrtenyl acetate); acyclic monoterpenes and monoterpenoids (linalool, β-myrcene, ocimenes (trans-3,7-dimethyl-1,3,6-octatriene and 2,6-dimethyl-2,4,6-octatriene), citral); methylated cyclohexane (2,5,5,8a-tetramethyl-3,4,4a,5,6,8a-hexahydro-2H-chromene); jasmone; menthane (menthol and menthyl acetate (a set of stereoisomers), mentofuran, menthone (a set of stereoisomers); -iso-propyl-1-methyl-2-cyclohexen-1-ol, 4-isopropenyl-1-methyl-2-cyclohexen-1-ol, isopulegol, isopulegylacetate, isopulegon, 4-terpineol, α-terpineol, pulegone, isopulegylacetate (set of stereoisomers), thymol, carvone, piperitone and piperitone oxide). At the same time, among the major components typical of peppermint essential oil of the menthol chemotype, is first of all menthol (52.55%) and menthyl acetate, mentofuran, menthone. In the essential oil of peppermint, linalool / carvone chemotypes, menthol is 2.31% / 4.96% and menthyl acetate 0.42% / 0.25%, and the product of cyclization - mentofuran - is not formed, and menthone in linalool mint is 8.15%, and in carvone type mint 7.92%. A sample of alcoholic extract of peppermint linalool chemotype is represented by the major monoterpenoid linalool, the content of which reaches 24.7%. Whereas the alcohol extract of peppermint of the menthol chemotype contains linalool in an amount of 0.69%, in carvone type mint it is 1.2%.

Sesquiterpenes and sesquiterpenoids in peppermint menthol linalool and carvone chemotypes are represented by compounds belonging to biosynthetic types of eleman (β- and τ-elemen); acyclic sesquiterpenoids (nero-lidol); biogenetic tree of germacran and biosynthetic types of cadinan (germacren D, germacren-D-4-ol, α-copaene, 1-isopropyl-4,7-dimethyl-1,2,4a,5,6,8a-hexahydronaphthalene and 1 -isopropyl-7-methyl-4-methylene-1,2,3,4,4a,5,6,8a-octahydronaphthalene (set of stereoisomers), 4-isopropyl-1,6-dimethyl-1,2,3,4 ,4a,7,8,8a-octahydronaphthalene-1-ol, δ-cadinene, cadina-1,3,5-triene); eudesman (α-selinene); bourbonan (β-bourbonene); aromadendran (aromadendren, α-guryu-nene, spatulenol, 1,1,4,7-tetramethyldecahydro-1H-cyclopropa[e]azulen-4-ol (set of stereo-isomers)); guayana (τ-gurjunen); cubeban (β-cubeben) and biogenetic tree of humulan (α-caryophyllene, β-caryophyllene and β-caryophyllene oxide). At the same time, in the alcohol extract of mint of both menthol and linalool and carvone chemotypes, β-caryophyllene acts as a major component (3.82%; 5.41% and 17.55%, respectively). For peppermint carvone type, this element can be considered marker along with carvone.

It is noteworthy that diterpene components - are present in mint of linalool and carvone chemtypes in a significant amount, while the menthol type contains them in amounts close to trace ones (Table 1).

It should be noted that in the alcohol extract of mint of both menthol, linalool, and carvone types, a wide range of components were identified that do not belong to the classes of terpenes and terpenoids and are contained in both mint morphotypes in comparable amounts. Those, for example, phytol, ethyllinolenate, ethyllineate (Table 1).

With the difference in the qualitative composition of the major components of the alcohol extract of mint belonging to the same species, there is an obvious difference in menthol, linalool and carvone chemotypes. This is an important factor that is not taken into account in the manufacture process of drugs, as well as mono raw materials, dietary supplements.

Table 1: Comparative results on the content of components Mentha piperita L., three chemotypes.

No.	Compound	Structural formula of a chemical substance	RT, Min	Menthol-chemotype	Linalool-chemotype	Carvone-chemotype
No.	Compound	Structural formula of a chemical substance	RT, Min	%	%	%
1	α-Tuyene, С10H16		23.66	0.05	0.09	0.15
2	α-Pinene, С10H16 β-Pinene, С10H16		24.05	0.09	0.24	0.29
3	β-Myrcene, С10H16		25.70	0.08	2.17	0.12
4	α-Terpinene, С10H16		27.06	0.09	0.06	0.11
5	Limonene, С10H16		27.53	0.75	0.65	0.55
6	β-Phellandrene, С10H16		27.68	0.10	0.06	0.22
7	1,8-Cineole, С10H18O		27.77	3.29	3.25	4.97
8	τ-Terpinene, С10H16		28.48	0.13	0.2	0.10
9	Cis-β-Terpineol (I,II) (total stereoisomes)		28.93 30.06	3.25	0.3	3.20
10	Linalool, С10H18O		29.60	0.69	24.70	1.2
11	Fenchon C10H16O		29.91	0.05	СК	СК
12	Isopulegol, С10H18O		31.76	0.14	0.11	0.16
13	Menton (I, II) (sum of stereoisomers) С10H18O		32.09 (I) 32.44 (II)	18.65	8.15	7.92
14	Carvone C10H14O		32.64	1.2	0.2	12.89
15	Menthol (I,II,III)		32.35	52.55	2.31	4.96
16	4-Terpineol, С10H18O		32.85	0.08	0.04	0.03
17	Kumaran №109771		33.43	0.10	0.1	0.30
18	Mirtenal, С10H14O		33.62	0.02	-	0.06
19	Piperitone С10H16O		35.32	0.79	0.21	0.34
20	Thymol, С10H14O		35.77	0.17	0.08	0.06
21	Myrtenyl acetate, С12H18O2		37.22	0.04	0.03	0.05
22	α-Copaen, С15H24		39.12	0.17	0.21	0.34
23	Aromadendren С15H24		40.22	0.12	0.91	0.89
24	α-caryophyllene, С15H24		41.59	0.30	0.21	1.12
25	δ-Kadinen, С15H24		42.98	0.12	-	0.24
26	4-Terpineol, С10H18O		32.85	0.08	0.04	0.03

The carvone chemotype of peppermint has the highest antioxidant activity (Table 2). Alcohol extraction at high heating shows the highest antioxidant activity, since this extraction extracts the maximum amount of flavonoid aglycones and other compounds.

Table 2: Temperature regimes of extraction of dried mint samples of menthol, linalool, carvone morphotypes for the determination of antioxidant activity by the amperometry method

Extraction Mode		Chemothype
		Linalool	menthol	Carvone
		Concentration of antioxidants, mg/g
1	Water, 20ºС	20.82±0,11	31.54±0,21	34,66±0,08
2	Water, 95ºС	30.70±0,08	40.25±0,19	45,25±0,21
3	Ethanol, 96%, 20ºС	5.38±0,07	8.62±0,09	10,22±0,16
4	Ethanol, 96%, 75ºС	11.52±0,10	17.50±0,11	23,21±0,09
5	Ethanol, 70%, 20ºС	22.71±0,14	29.44±0,20	32,43±0,18
6	Ethanol, 70%, 85ºС	39.53±0,18	45.00±0,09	46,33±0,10

The content of oxidizable substances (GF X1V) was studied by the method of permanganatometric titration. The amount of oxidizable substances turned out to be maximum in the mint sample of carvone type 10.24±0.21. Peppermint leaves of menthol and linalool types contain approximately the same number of oxidizable substances -7.00±0.18 and 8.28±0.07, respectively

In a comparative analysis of the tonic activity of an aqueous extract from dried leaves of peppermint of the menthol, linalool and carvone types, the maximum effect of an aqueous extract from the leaves of peppermint of the menthol type was found. Apparently, the effect on the central nervous system is exerted primarily by essential oil, namely, menthol and its derivatives. Aqueous extracts from linalool and carvone type peppermint samples also show pharmacological activity compared to control group (Table 3).

CONCLUSION:

More than 50 compounds of the medicinal plant raw materials of peppermint leaves of menthol, linalool, carvone types, were identified by chromatography-mass-spectrometry, that make up the essential oil. Comparative analysis showed that the content of menthol in peppermint leaves of the menthol type can exceed the content of menthol in other chemotypes by ten or more times. At the same time, the tonic effect of an aqueous extract from peppermint leaves is largely associated with the content of menthol. It should be taken into account when using this type of raw material in the manufacture of preparations and monotherapy. The raw material of peppermint leaves can have a different degree of pharmacological activity depending on the chemotype, which should be taken into consideration when using this type of raw material.

Table 3: Results of the experiment "The mouse forced swim test"

Groups of animals (n=40)	Duration of swimming at the beginning of the experiment			Duration of swimming at the end of the experiment
Groups of animals (n=40)	Primary (min)	Repeat (min)	Repeat %	Primary (min)	Repeat (min)	Repeat %
Control group	13,7±0,5	13,0±0,9	94,6±4,8	13,7±0,3	13,0±0,4	95,0±3,0
Aqueous extract from menthol-type mint leaves	13,6±0,2	12,5±0,3	92,0±2,1	13,3±0,5	16,0±0,6	120,3±4,8
Mint linalool type	13,5±0,2	12,4±0,2	92,5±2,4	13,3±0,5	15,2±0,3	114,8±4,3
Mint carvone type	13,7±0,2	12,6±0,2	92,0±1,6	13,3±0,4	15,3±0,3	115,1±3,4

Peppermint leaves of the carvone chemotype have the highest antioxidant activity compared to other chemotypes, which makes it possible to use this type of raw material as an antioxidant agent.

ACKNOWLEDGMENTS:

The study was supported by the Sechenov University.

CONFLICTS OF INTEREST:

The authors declare no conflicts of interest.

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Received on 18.02.2023 Modified on 27.09.2023

Research J. Pharm. and Tech 2024; 17(2):625-630.

DOI: 10.52711/0974-360X.2024.00097