Changes of German Chamomile Essential Oil Content in Response to Different Plant Densities and Nitrogen Rates

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1 Changes of German Chamomile Essential Oil Content in Response to Different Plant Densities and Nitrogen Rates 1* A. Khourgami, 2 G. Sharafi, 3 S. Ahmadi and 4 Z. Bitarafan 1 Assistant Professor of Agronomy, Khorramabad Branch, Islamic Azad University, Khorramabad, Iran *(corresponding author) 2 Master of Agronomy, Khorramabad Branch, Islamic Azad University, Khorramabad, Iran 3 Instructor, Razi Herbal Medicine Research Center, Lorestan University of Medical Science, Khorramabad, Iran 4 Department of Agronomy, Science and Research Branch, Islamic Azad University, Tehran, Iran Abstract- A field study was conducted in Khorramabad, Iran during 2010 spring growing season with the objective to determine the effect of plant density and nitrogen application on German chamomile (Matricaria chamomilla L.) essential oil percent and components (Bisabolone, Caryophyllene, Chamazulene, Azulene, Carene, Eucalyptol, α- cubene and Spathulenol). The experiment were laid out in a three-replicatedrandomized complete block design with three levels of plant density (D: D 1 = 28.6, D 2 = 40 and D 3 = 66.7 p m -2 ) and three levels of nitrogen (N: N 1 = 0, N 2 = 100 and N 3 = 200 kg N ha -1 ). The study indicated that the essential oil percent influenced by interaction of plant density and nitrogen and the highest rate of it obtained in 40 p m -2 and 100 kg N ha -1 by average of 0.2%. Among assessed essential oil components, the highest Bisabolone, Chamazulene and Eucalyptol were obtained on D 3 N 2, D 2 N 3 and D 1 N 3, respectively and Caryophyllene, Carene, α- cubene and Spathulenol were not affected by treatments. Key words: German chamomile; Matricaria chamomilla L.; Plant density; Nitrogen; Essential oil, Bisabolone; Chamazulene. I. INTRODUCTION The medicinal plant of chamomile is widely used throughout the world. Its primary uses are as a sedative, anxiolytic, antispasmodic, and as a treatment for mild skin irritation and inflammation. Also it has widespread use as a home remedy [1]. There are numerous kinds of chamomile. Two most popular are Roman chamomile and German chamomile, both are from the Compositae family. German chamomile (Matricaria recutita L. (syn. M. chamomilla L., Chamomilla recutita L. Rauschert)) is more widely cultivated than Roman chamomile (Chamaemelum nobile (L.) All. (syn. Anthemis nobilis L.) [2,3]. The biological activity of chamomile is mainly due to the phenolic compounds, primarily the flavonoids apigenin, quercetin, patuletin, luteolin and their glucosides, but also to the principal components of the essential oil extracted from the flowers like α-bisabolol and its oxides and azulenes, including camazulene [4]. Costescu et al. (2008) revealed in a study the main sesquiterpenes in the chamomile essential oil. Camazulene (19.9%), α- bisabolol (20.9%), A and B bisabolol-oxides (21.6% and 1.2%, respectively) and β-farnesen (3.1%) were the major components. In lower concentrations were identified α- and β-caryophyllene, caryophylleneoxide and spathulenol, and also some monoterpenes like β-phellandrene (0.8%), limonene (0.8%), β- ocymene (0.4%) and γ-terpinen (0.2%) [5]. Essential oils of M. recutita L. cultivated in Estonia were isolated in another study by Orav et al. (2001) and thirty-seven components were identified. The main components were bisabolol oxide A (20 33%) and B (8 12%), bisabolon oxide A (7 14%), (E)-farnesene (4 13%), α- bisabolol (8 14%), chamazulene (5 7%), and en-yne-dicycloether (17 22%) [6]. Another investigation in Estonia indicated that the main constituents of the essential oils were as follows: bisabolol oxide A (39.4%), bisabolone oxide A (13.9%), (Z)-en-yne-dicycloether (11.5%), bisabolol oxide B (9.9%), α-bisabolol (5.6%), and chamazulene (4.7%) [7]. An Iranian study in Isfahan indicated essential oil components of German chamomile isolated by hydro-distillation of the aerial parts of the plant. Sixty-three components were characterized, representing 86.21% of the total oil components 19

2 detected. α-bisabolol oxide A (25.01%) and α- bisabolol oxide B (9.43%) were the major constituents of the oil [8]. Chamomile growth as well as composition and quality of essential oil are influenced by many factors like genotype, environment and agronomic practices. Fertilizing with nitrogen due to its multi-dimensional effects on the growth and development and plant density are two key points in chamomile production. It is necessary to apply sufficient amount of nitrogen to achieve optimum yield and high-quality product. However over-fertilization and insufficient fertilization applications lead to economic losses and discharge of excessive amount of nitrogen in the nitrate form through washing [9]. Also plant density is very important to facilitate aeration and light penetration into plant canopy for optimizing rate of photosynthesis [10, 11, 12]. Optimum plant density increase the canopy closure rate, leaf area index, environmental factors use efficiency, yield components and ultimately yield [13]. Salamon et al. (1992) suggested and 5-10 cm space between rows and plants, respectively in chamomile production [14]. Little is known about the effects of N fertilizer and plant density on essential oil components of chamomile, therefore the main objective of this study was assessing the effects of plant density and nitrogen application on some essential oil components of German chamomile (matricaria chamomilla L.) in order to determine the most appropriate plant density and nitrogen fertilizer rate. II. MATERIALS AND METHODS To assess the effect of plant density and nitrogen application on German chamomile (matricaria chamomilla L.) essential oil percent and components, an experiment was conducted in 2010 spring growing season at Research Farm of Khorramabad Branch of Islamic Azad University, Khorramabad, Iran ( E, N; 1350 m a.s.l). Average annual rainfall and temperature in this region are mm and 17 C, respectively. The soil type where the experiment took place was a silty clay loam soil. The soil characteristics were determined before experiment for fertilizer suggestions, which is presented in table 1 (table 1). Characteristic Depth (cm) 0-30 Clay 32 Silt 50 Sand 18 Available N (ppm) 14 Available K (ppm) 360 Available P (ppm) 28.2 Fe (ppm) 5.8 Mn (ppm) 8.6 Zn (ppm) 0.38 Cu (ppm) 0.48 TNV 16.7 Organic Carbon 1.36 ph 7.6 EC (ds m 1 ) 0.45 Soil texture SCL Table1. Soil characteristics of the experimental site in Khorramabad, Iran The experimental design was a factorial arrangement in the form of randomized complete block design with three replications. Treatments were included two agents: three levels of plant density (D: D 1 = 28.6, D 2 = 40 and D 3 = 66.7 p m -2 ) and three levels of nitrogen (N: N 1 = 0, N 2 = 100 and N 3 = 200 kg N ha -1 ). N was used in the form of urea and each N treatment applied in two stages: 1/2 pre-plant and 1/2 in stemming stage. Each experimental plot extended in 6 2 m 2 area with 6 and 2 m distance between blocks and plots, respectively to prevent treatments mingling. P was applied at a rate of 75 kg P 2 O 5 ha 1 pre-plant in the form of ammonium phosphate and incorporated in the soil before sowing. Seeds mixed by appropriate rate of sand, were planted on 12 Mar with fixed intra-row distance of 10 cm and 15, 25 and 35 cm inter-row distances to achieve the plant densities of 28.6, 40 and 66.7 p m -2, respectively. Irrigation was done each 3 days which was done using handy sprinkler at early stages to prevent seeds diffusion. The plants were thinned after complete emergence in the 3-4 leaf stage. At flowering stage, 4 middle rows were used for flower sampling (2 Jul. 2010). Essential oil of flowers extracted using Hydrodistillation method and its components (Bisabolone, Caryophyllene, Chamazulene, Azulene, Carene, Eucalyptol, α- cubene and Spathulenol) were determined. Analyses performed using the MSTATC software. A factorial analysis of variance (ANOVA) was performed for all parameters. In addition the 20

3 Duncan s Multiple Range Test (DMRT) (P = 0.05) was used to conduct mean comparison of treatments. III. RESULTS AND DISCUSSION The results of analysis of variance revealed that the simple effect of plant density was significant on Bisabolone, Chamazulene and Eucalyptol at P = 0.05, 0.01 and 0.01, respectively (Table 2). The highest rate of Bisabolone, Chamazulene and Eucalyptol were obtained in D 3, D 2 and D 1 by average of 8.49, 4.26 and 0.63%, respectively, although the difference among D 1 and D 3 in production of highest rate of Bisabolone was negligible (Fig. 1, 2 & 3). Also the simple effect of nitrogen was significant on Bisabolone, Chamazulene and Azulene all at P = 0.01 (Table 2). Bisabolone, Chamazulene and Azulene increased by nitrogen application as the highest rate of them obtained in N 2, N 3 and N 2 by average of 9.14, 4.34 and 0.53%, respectively (Fig. 1, 2 & 4). Franz (1983) reported chamomile essential oil increases by N application up to a definite level [15]. The interaction effect of D N were statistically significant on essential oil percent, Bisabolone and Chamazulene at P = 0.01 and on Eucalyptol at P = 0.05 (Table 2). Study the interaction effect of D N showed that the highest essential oil percent, Bisabolone, Chamazulene and Eucalyptol were obtained on D 2 N 2 (0.2%), D 3 N 2 (9.62%), D 2 N 3 (7.45%) and D 1 N 3 (0.75%), respectively and the lowest rates of them were obtained on D 2 N 3 (0.061%), D 2 N 1 (4.96%), D 3 N 2 (2.10%) and D 3 N 1 (0.26%), respectively (Fig. 5, 6,7 & 8). S.O.V. D. F. Essen tial oil Bisabo lone Caryoph yllene Chamaz ulene Azul ene Replic 2 ation Plant 2 ns * ns ** ns density Nitrog 2 ns ** ns ** ** en Plant 4 ** ** ns ** ns density nitroge n Error 16 Total C.V *, ** significant at 5 and 1% respectively, ns: not significant S.O.V. D.F. Caren e Eucalypto l α- cuben e Spathuleno l Replicatio 2 n Plant 2 ns ** ns ns density Nitrogen 2 ns ns ns ns Plant 4 ns * ns ns density nitrogen Error 16 Total C.V *, ** significant at 5 and 1% respectively, ns: not significant Table 2- Analysis of variance for assessed traits Caryophyllene, Carene, α- cubene and Spathulenol were not statistically affected by any of treatments in this study (Table 2). The German chamomile should be harvested when most of flowers have been grown. Early or late harvest would reduce the quality of effective materials. The flowers have maximum amounts of essential oils when ray florets are in mood; afterwards, the amount of the essential oils decreases [16]. Balak et al. (1999) reported the highest chamomile yield and growth indices obtained by 120 kg N ha -1 [17]. Hornok (1992) reported high levels of N fertilizer reduce the flower yield [18]. Rahmati et al. (2009) suggested the intra-row space of 20 cm and inter-row space of 25 cm to increase the dry flower yield [19]. Fig. 1. Simple effects of D and N on Bisabolone 21

4 Fig. 2. Simple effects of D and N on Chamazulene Fig. 5. Interaction effect of D and N on essential oil Fig. 3. Simple effect of N on Eucalyptol Fig. 6. Interaction effect of D and N on Bisabolone Fig. 4. Simple effect of D on Azulene Fig. 7. Interaction effect of D and N on Chamazulene 22

5 professionals. London: Pharmaceutical Press. ix: 296. [3] Blumenthal, M The complete German Commission E monographs: therapeutic guide to herbal medicines. Austin: American Botanical Council. [4] Hadaruga, N. G., D. I. Hadaruga, C. Tatu, A. Gruia, C. Costescu, A. X. Lupea Multivariate analysis (PCA) in Compositae biocompounds class. Journal of Agroalimentary Processes and Technologies. 15: Fig. 8. Interaction effect of D and N on Eucalyptol IV. CONCLUSIONS Choosing the most appropriate plant density and nitrogen fertilizer rate are two important agronomic practices in crop production management as they strongly affected different aspects of the crop. Essential oil content which is the main objective in production of medicinal plants could be changed in different plant densities and nitrogen rates. This study provides new information about the effect of different plant densities and nitrogen rates on essential oil percent and components of German chamomile. In this study essential oil percent influenced by interaction of plant density and nitrogen and the highest rate of it obtained by 40 p m -2 and 100 kg N ha -1. Among assessed essential oil components, the highest Bisabolone, Chamazulene and Eucalyptol were obtained on D 3 N 2, D 2 N 3 and D 1 N 3, respectively and Caryophyllene, Carene, α- cubene and Spathulenol were not affected by treatments. According our objective in production of specific component or components, the appropriate plant density and nitrogen rate should be suggested. REFERENCES [1] Gardiner, P Chamomile (Matricaria recutita, Anthemis nobilis). Retrieved from: and [2] Newall, C. A., L. A. Anderson, J. D. Phillipson Herbal medicines: a guide for health-care [5] Costescu, C. I., N. G. Hadaruga, A. Rivis, D. I. Hadaruga, A. X. Lupea, D. Parvu Antioxidant activity evaluation of some Matricaria chamomilla L. extracts. Journal of Agroalimentary Processes and Technologies. 14: [6] Orav, A., T. Kailas, K. Ivask Volatile constituents of Matricaria recutita L. from Estonia. Proc. Estonian Acad. Sci. Chem. 50: [7] Raal, A., H. Kaur, A. Orav, E. Arak, T. Kailas, M. Muurisepp Content and composition of essential oils in some Asteraceae species. Proceedings of the Estonian Academy of Sciences. 60: [8] Shams Ardakani, M., A. Ghannadi, A. Rahimzadeh Volatile Constituents of Matricaria chamomilla L. from Isfahan, Iran. Iranian J. Pharma. Sci. 2: [9] Grant, C Enhancing nitrogen use efficiency in dry land cropping systems on the Northern Great Plains. 18 th World Congress of Soil Science. Philadelphia, USA. [10] Khan, R. U., A. Ahad, A. Rashid, A. Khan Chickpea production as influenced by row spacing under rainfed conditions of Dera Ismail Khan. J. Biol. Sci. 3: [11] Azizi, K., D. Kahrizi Effect of nitrogen levels, plant density and climate on yield quantity and quality in cumin (Cuminum cyminum L.) under the conditions of Iran. Asian J. Plant Sci., 7(8): [12] Kahrizi, D., A. H. Salmanian, A. R. Zebarjadi Effect of cultivar and density of cultured 23

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