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Pdf 521.14 K Journal of Medicinal Plants and By-products Original Article Genetic Variation of Shoot Yield, Essential Oil and Yield Components in Four Thyme Species Majid Dashti1, Ali Ashraf Jafari2, Narjes Azizi1*, Seid Ali Mazloom Maghaddam3 and Abdoul Karim Negari4 1Khorasan Razavi Agricultural and Natural Resources Research and Education Center, Agricultural Research, Education and Extension Organization (AREEO), Mashhad, Iran 2Research Institute of Forest and Rangeland, Agricultural Research and Education Organization, (AREEO), Tehran, Iran 3University of Tehran, Tehran, Iran 4Khorasan Razavi Agricultural and Natural Resources Research and Education Center, Agricultural Research, Education and Extension Organization (AREEO), Mashhad, Iran Article History: Received: 10 October 2020/Accepted in revised form: 16 January 2021 © 2012 Iranian Society of Medicinal Plants. All rights reserved. Abstract Thymus daenensis Celak, T. kotschyanus, T. transcaspicus and T. vulgaris are aromatic and medicinal species which due to hybridization within species and between species has high morphological diversity. A study was carried out in order to evaluate yield and yield components in 10 ecotypes of Thymus species collected from Isfahan, Markazi, Qazvin and West Azerbaijan provinces. This experiment was performed in a randomized complete block design with three replications at Research Center for Agriculture and Natural Resources Research and Education of Razavi Khorasan province. Plant height, shoot yield, 1000-seed weight, essential oils (EO) content, EO yield, number of nodes per stem, internode length and days to flowering stage were measured in all ecotypes. T. vulgaris was considered as the control. The results of analysis of variance showed that there were significant differences (p≤0.05) for days to 50 % flowering, number of nodes, internode length, plant height, stem weight, seed weight, EO content and yield among species and ecotypes within Thymus species. The highest EO content and yield among species were observed by 2.45% and 1.28 g.plant-1 in T. vulgaris, and 2.03% and 0.84 g. plant-1 in T. daenensis, respectively. The highest EO content was observed among ecotypes at 1.84% in T. kotschyanus, ecotype K54 and 2.32% in T. daenensis, ecotype D49. The strong and ** significant (P ≤ 0.01) positive correlation of EO yield was observed with EO content (r =0.88 ), seed weight per plant ** * ** ** (r=0.78 ), plant height (r=0.70 ), shoot yield (r=0.80 ) and internode length (r=0.89 ). In the principal component analysis in thyme species and ecotypes, the eigenvalues obtained from the first three main components, explain 53%, 15%, 11%, and a total of 80% of the total variance of the variables. It was concluded that the ecotypes of D49 and D72, in T. daenensis having higher EO yield were recommended for breeding improved varieties. Keywords: Thyme, Yield, Essential oil, Correlation analysis, Heritability Introduction Medicinal plants are very important in modern civilization in order to obtain natural active substances, known as secondary metabolites. Nowadays, the uncontrolled harvesting leads to the extinction of ecotype and species [1]. The genus Thymus is a well-known aromatic perennial herb widely used in folk medicine, food preservatives and pharmaceutical preparations originated from Mediterranean region belongs to the Lamiaceae family. This genus is consisting of about 215 species of herbaceous perennials and small shrubs in the world and 14 species in Iran. They are distributed in Iranian flora consist of T. daenensis, T. carmanicus, T. fallax, T. persicus, T. trautvetteri, T. migricus, T. kotschyanus, T. pubesens, T. nummularius, T. transcaspicus, T. eriocalyx, T. caucasicus, T. transcaucasicus, and T. fedtschenkoi [2]. Thymol and carvacrol are the major compounds in most of the Thymus essential oil. The therapeutic potential of thyme rests on contents of thymol, carvacrol, flavonoids, eugenol, aliphatic phenols as well as luteolin, saponins, and tetra methoxylated flavones. The EO of Thyme has antibacterial, antiseptic, antifungal, anti-parasitic and antioxidant activity [3,4]. *Corresponding Author: [email protected] The terpene phenols thymol and carvacrol represent the most important compounds in the genus, followed by linalool, p- cymene, γ-terpinene, borneol, terpinen-4-ol and 1, 8-cineole [5]. In traditional medicine, the leaves and flowering parts of Thymus species are widely used as tonic, herbal tea, flavoring agents (condiment and spice), antiseptic, antitussive and carminative as well as treating colds [6,7]. Based on the results of different studies, EO content (%, v/w) in cultivated samples of thyme varied between 0.32-0.75% and 0.62–1.05% within two examined lineages, whereas the herbs from the natural habitat contained 0.49–1.29% [8]. Twenty-four components in total were identified representing more than 85–98% of the oil composition, with thymol (33.9%–70.3%), carvacrol (4.0%–24.8%), γ-terpinene (3.9%–10.4%) and p-cymene (4.8%–8.6%) in Thymus daenensis. Thymol (35.5%–44.4%), carvacrol (4.4%–16.1%), γ-terpinene (10.5%–11.9%) and p-cymene (8.5%–16.1%) in Thymus vulgaris as major constitutes. The EO and their chemical compositions of Thymus species are strongly affected by environmental conditions and agronomic management practices. Cultivation of thyme, especially T. daenensis proved to be superior in both oil content and quality, in terms of, the phenols carvacrol and thymol [9]. The EO of leaves in Thymus transcaucasicus Ronniger was distinguished by high amounts of thymol (35.83%), α-terpineol (12.38%), geraniol (10.79%) and p-cymene (5.98%). Thirty-six constituents representing 99.75% of the flower oil were identified of which thymol (62.92%), p-cymene (5.78%), geraniol (5.58%) and carvacrol (4.14%) were major components. The main components of the root oil were thymol (60.61 %), p-cymene (9.32%), carvacrol (4.61%), pentacosane (3.17%) and 1.8- cineole (3.08%). Thymol (46.07%) and geraniol (9.20%) predominated in the stem oil [10]. Drought and salinity stress influenced to increase the amount of phenol contain and consequently increases its antioxidant properties and it makes it superior to Thyme [11]. This is an evidence that the synthesis of EO in aromatic plants can be influenced by genetics, stage of development and environmental factors such as temperature, nutrition and photoperiod [12]. Previous researches have shown that the EO yield was high in the full flowering stage, the sum of the two compounds is thymol and carvacrol. In another study concluded that suitable time of harvesting of the EO was at flowering stage in summer [13]. The study of the phenological and morphological characteristics of the medicinal plants has leads to a better understanding of the extent and manner of growth and development in different species and ecotypes. Maximum growth rates in any particular stage of fruit growth and secondary metabolites are not similar in early maturing to late maturing of ecotypes or cultivars in the whole growth stages. The amount of active substances could be done based on this feature [14]. This research was aimed to study of genetic variation and relationship of shoot yield, EO content and yield in four thyme species. Material and Methods Field experiments were carried out in a randomized complete block design with three replications at Razavi Khorasan Agricultural and Natural Resources Research and Education Center, Mashhad-Iran. The experimental farm located at latitude 36° 13' N, longitude 59° 40' E, and at an altitude of 980 m above the sea level with semi-arid climate, mean annual precipitation of 254 mm, mean annual temperature of 14.5 °C and the average temperature in winter and summer seasons of -1.8 °C and 33 °C, respectively. The analysis of soil chemical and physical properties are presented in Table 1. Treatments were consisting of 4 species and 10 ecotypes. The study ran for two years in 2018 and 2019. Seeds were provided by national project leader in Research Institute of forests and rangelands (RIFR). The origin of seeds is presented in Table 2. In order to implement the project, seed samples of the plants were first seeded in greenhouse in winter and then transplanted to the farm in early spring. They were planted in the plots of 2x5 meter with 4 rows for each ecotype in late March 2018. Irrigation was performed weekly in a dripping manner. The fertilization was done according to the recommended values using 50 kg of urea fertilizer + 150 kg of triple super phosphate per hectare. Weeds was controlled mechanical by 2-3 times during growth period. At 50% of flowering stage in the first and second year, plant height, number of nodes per stem, internode length, flowers weight, leaves and stem weight was measured in five plants. Plants in each plot were harvested for biomass production. The fresh and dry weight was measured for each sample. Biomass and grain yield and EO content were measured in all thyme species and ecotypes. EO were extracted by hydro distillation method using 30-gram sample and the Clevenger-type apparatus. Statistical analysis, including correlation, stepwise regression and PCA analysis were performed using SPSS24 and Minitab software. Table 1 Physical and chemical properties of the field soil (%) Soil depth FC Organic (Cm) WP TNV SP Clay Silt Sand N Carbon 0-30 8.4 16.4 0.45 17 34 23 36 41 0.092 30-60 9.3 17.8 0.19 23 39 23 34 43 0.038 (ppm) Soil depth EC pH (Cm) Cu Zn Mn Fe K P (ds.m-1) 0-30 0.96 0.56 5.40 3.76 219 10 1.63 8.0 30-60 0.54 0.12 3.32 5.86 69 4.4 1.44 8.0 Table 2 List of used Thymus species and ecotypes received from the gene bank resources in Iran. Ecotype Species Name Gene bank code Origin Treatment code D49 1149 Isfahan 1 T.
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