Thyme-Eco extract is produced from the leaves and flowers of Thymus vulgaris, proceeding from organic crop.

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1 Thyme-Eco BOTANY Thymus vulgaris L., common name: thyme, is a perennial aromatic sub-shrub, member of the Labiatae (Lamiaceae) family. This plant grows between 10 and 40 cm in height. The stems are woody and grayish. The leaves are opposite, grayish-green, entire, linear or elliptic, up to 15 mm long, tomentose beneath. The flowers are small, bilabiate, pale-purple or white, arranged in terminal inflorescences that may be dense or loose. Flowers appear since the beginning of summer until the end of autumn. The fruit is an ovoid smooth achene. Thyme is native to the western Mediterranean area, especially the south of Italy, wherefrom it spread to almost every region. In the wild, it grows forming dry shrubberies, on rocky well-drained soils and sunny areas, up to 2500 m altitude. This plant is cultivated in almost every country, as an aromatic plant for culinary uses (especially in the south of France, Spain, Morocco and North America). Thyme-Eco extract is produced from the leaves and flowers of Thymus vulgaris, proceeding from organic crop. CHEMISTRY According to the relation with its properties and applications, the main components in thyme are essential oil ( %) and polyphenols (flavonoids and tannins) (Cañigueral S & Vanaclocha B, 2000). Essential oil It is mainly composed of the monoterpene phenols thymol and carvacrol, adding not less than 40% of the whole according to the European Pharmacopeia. A small amount of these phenols occur as glycosides. In the essential oil, other monoterpenes (pcymene, γ-terpinene) and several sesquiterpenes (βcariophyllene). The quantitative composition can differ according to the origin and time of harvest, being recognized six different chemotypes. The most common ones containing thymol, carvacol or linalol as the most abundant component. However, there are thyme varieties that contain other components Fig.1 Thymol V 02-11/

2 (geraniol as the most abundant component (Alonso J, 2004) (Cañigueral S & Vanaclocha B, 2000). Flavonoids Mainly luteol and apigenol heterosides with smaller proportions of methoxylated flavones: cosmosiin, thymonine, isothymonine, 8-dimethyl-thymonine, timusin, naringenin, eriodictiol, cirsimaritin, xanthomicrol, 5-desmethylnobiletin, 5-desmethylsinensetin, sideritoflavone, cirsilineol and 8-methoxycirsilineol. Flavanones, flavonols and luteolin heterosides have also been identified (Alonso J, 2004). Other active principles Tannins (7-10%), serpyllin (bitter principle), acid and neutral saponins, labiatic, oleanolic and ursolic acids (1.5%), phenylcarboxhylic acids (chlorogenic and caffeic), rosmarinic acid (<1%), lithospermic acid, resin (Alonso J, 2004). TRADITIONAL USES Thyme is prepared as infusion to treat spasmodic cough, laryngitis, bronchitis and urinary infections. It is also used as a decongestant, as a cholagoge, to reduce flatulence and to fight parasites. External uses of thyme include preparations to wash skin wounds or infections. Experience indicates that thyme has stronger antiseptic effects than hydrogen peroxide or phenol. Thyme macerated in oil and applied as poultices has analgesic effects on rheumatism, sprains or stiff neck. Scrubbing with thyme macerated in alcohol has the same effects. Thymol is extensively used in the cosmetic industry to prepare mouth wash and tooth paste. COSMETIC PROPERTIES Antimicrobial activity Thyme essence especially the phenolic components thymol and carvacol show antibacterial activity against Gram-positive and Gram-negative bacteria, mainly due to their effects on the bacterial membrane. Since thymol and carvacol are eliminated through the respiratory tract, these compounds have respiratory antiseptic action. Because of its antibacterial activity, thyme is also useful as an antiseptic for the urinary tract, mouth and skin wounds. Furthermore, thymol and carvacol are antimycotic agents, effective against Candida albicans. Thyme water extract showed significant in vitro inhibitory effects on the growth of Helicobacter pylori and its powerful urease activity (Cañigueral S & Vanaclocha B, 2000). Sebum excess promotes the growth of certain microorganisms on the skin and the scalp. Such microbial flora imbalance is one of the factors causing dandruff and acne. Thyme antimicrobial activity is of great use to formulate cosmetic products aimed at the regulation of sebum hypersecretion and treatment of related disorders. Therefore, thyme-eco extract is recommended to formulate cosmetic products with purifying and antiseptic activity. Anti-inflammatory activity In vivo tests of thyme ethanolic extract on rats showed anti-inflammatory and analgesic activities. These activities could be related to carvacol and thymol, which showed inhibitory effects on the enzyme cyclooxygenase in animal models, as well as inhibitory effects on the complement and on nitric oxide synthesis (Alonso J, 2004). Carvacol has inhibitory action on prostaglandin synthesis. This action supports the use of thyme in ointments and other preparations to treat muscle and joint pain. The rosmarinic acid in this extract also has anti-inflammatory activity. Topical applications of thyme essential oil are rubefacient and generate analgesia, beneficial in cases for bruises or sprains (Cañigueral S & Vanaclocha B, 2000). V 02-11/

3 A study carried out on isolated rat sciatic nerve demonstrated thymol analgesic effects. After 180 minutes application of concentrations ranging from mm, neuronal excitability was modified (Alonso J, 2004). Anti-inflammatory action has been reported for the phenol compounds in thyme extracts (García Bacallao L et al, 2002). Thus, thyme-eco extract is recommended to formulate cosmetic products with anti-irritant activity. Antioxidant activity The thymol and carvacol, present in thyme essence, as well as the flavonoids and other polyphenols are considered to be involved in the antioxidant activity (Cañigueral S & Vanaclocha B, 2000). Rosmarinic acid, hydroxycinnamic derivatives and flavonoid compounds showed important in vitro antioxidant activity by inhibiting iron-induced superoxide anion formation and lipid peroxidation in microsomal and mitochondrial systems. Furthermore, the thymol present in the essential oil showed in vitro antioxidant activity by neutralizing the DPPH (diphenyl-picrylhydrazyl) radical. Aged rats, which had been fed on a diet including thyme since young age, showed high proportions of antioxidant enzymes such as superoxide dismutase in liver and heart, as compared with a control group (Alonso J, 2004). Therefore, thyme-eco extract is recommended to formulate cosmetic products aimed at the protection of skin and hair integrity against oxidative processes. Finally, we would like to mention that the reference publication Plants in cosmetics. Vol. I (Council of Europe, 1994), which includes a monograph on the glycolic extract of Thymus vulgaris flowered herb. The following cosmetic effects and recommended uses appear: tonic, deodorant, purifying, astringent, fragance for use in deodorants, purifying products for greasy/acneic skin, stimulant baths. other possible effects: antiseptic (strong), stimulant, balsamic BIBLIOGRAPHIC EFFICACY TEST There are many papers with in vitro tests demonstrating the antimicrobial activity for extracts and the essential oil of Thymus vulgaris. Panizzi L et al (1993) carried out a study on the essential oil obtained from 4 species of Labiatae family. They analysed the chemical composition and evaluated the antimicrobial activity. 1. Experimental method The flowering tops of leafy twigs from Thymus vulgaris L., Rosmarinus officinalis L., Satureja montana L., y Calamintha nepeta (L) Savi were collected and dried. To obtain the essential oil, a 200g sample was subjected to steam distillation for 2 hours according to the Italian Pharmacopoeia (Farmacopea Ufficiale, 1985). Later, the composition of the essential oils was determined by gas chromatography analysys. Once obtained and analyzed the essential oils, the antimicrobial capacity was evaluated. For this investigation, MIC (Minimum Inhibitory Concentration) and MBC (Minimum Bactericidal Concentration) for the following microorganisms were calculated: V 02-11/

4 Staphylococcus aureus ATCC Escherichia coli ATCC Pseudomonas aeruginosa ATCC Bacillus subtilis BGA Saccharomyces cerevisiae ISS MC 40B Candida albicans (aislada en el laboratorio) These microorganisms were selected as representative of Gram positive, Gram negative bacteria and fungi. Each microorganism was cultured with different concentrations of each essential oil. Tween 80 was used to facilitate the incorporation of the essential oil to the aqueous culture medum. The inoculum was 5x10 6 cells, incubated 48 hours at 35ºC. 2. Results and conclusions Table 1 shows the results of the analysys composition of the essential oils obtained from the 4 species of Labiatae. Compound Satureja Rosmarinus Thymus Calamintha montana officinalis vulgaris nepeta α-pinene Canphene β-pinene Myrcene d-limonene ,8-Cineol α-phellandrene 1.41 α-terpinene 1.21 γ-terpinene p- Cimene Menthone 9.82 Isomenthone 0.19 Camphor Linalool Bornyl acetate β-caryophyllene Menthol 4.82 Pulegone 46.0 α-terpineol Piperitone oxide 2.29 Piperitenone 2.00 Piperitenone oxide 2.53 Borneol Verbenone 0.42 Carvone 0.41 Geraniol Thymol Carvacrol Table 1. Composition (%) of essential oils. Except for Rosemary, all the essences examined exhibit a composition similar to those commonly reported. Regarding Thyme, because of its essential oil composition, this plant belongs to the chemotype rich in phenolic compounds (thymol, carvacrol, γ-terpinene y p-cimene). V 02-11/

5 S. aureus E.coli P.aeruginosa B.subtilis S.cerevisiae C.albicans Satureja montana MIC 5 40 No MBC 5 40 Activity Calamintha nepeta MIC MBC Thymus vulgaris MIC 5 2 No MBC 5 2 Activity Rosmarinus MIC No officinalis MBC Activity Table 2. MIC and MBC of essential oils (values in μg/ml of liquid culture medium) In table 2 appear the results of the antimicrobial activity for the 4 species studied. All 4 essential oils show a wide spectrum of action. Moreover, there is a total coincidence between MIC and MBC values. This means that the inhibition of growth observed is always due to a biocidal effect. If we compare the efficacy of the 4 essences, Rosmarinus officinalis would be the less active. By other side, which offers the bests results is Thymus vulgaris one, being the stronger inhibitor in 4 out of 6 microorganisms, specially against E.coli and C.albicans. These results show the antimicrobial capacity that thyme extracts can have, specially because of its composition in essential oil. COSMETIC APPLICATIONS Action Active Cosmetic Application Antimicrobial Essential oil -Purifying -Antiseptic -Anti-seborrhea Anti-inflammatory Antioxidant Phenol compounds Essential oil Phenol compounds Essential oil -Anti-irritant -Anti-ageing -Photo-protector -Hair color protection RECOMMENDED DOSE The recommended dose is between 0.1% and 3.0%. V 02-11/

6 BIBLIOGRAPHY Alonso, J. Tratado de Fitofármacos y Nutracéuticos. Barcelona: Corpus, 2004, p: (633.8 ALO). Cañigueral Folcarà S & Vanaclocha Vanaclocha B. Usos terapéuticos del tomillo. Revista de Fitoterapia, 2000; 1 (1): 5-14 (ref. 4464). Garcia Bacallao, Rojo Domínguez DM, García Gómez LV, Hernández Ángel M. Plantas con propiedades antiinflamatorias. Rev. Cubana Invest. Biomed., 2002; 21(3): [scielo.sld.cu/pdf/ibi/v21n3/ibi12302.pdf]. Panizzi L, Flamini G, Cioni PL, Morelli I. Composition and antimicrobial properties of essential oils of four Mediterranean Lamiaceae. Journal of Ethnopharmacology, 1993; 39: 167:170. Websites: [accessed December 2007]. V 02-11/