This art icle was downloaded by: [ Ege Universit esi Rekt orlugu] On: 28 January 2014, At : 04: 03 Publisher: Taylor & Francis I nform a Lt d Regist ered in England and Wales Regist ered Num ber: 1072954 Regist ered office: Mort im er House, 37- 41 Mort im er St reet , London W1T 3JH, UK Journal of Essential Oil Bearing Plants Publicat ion det ails, including inst ruct ions f or aut hors and subscript ion inf ormat ion: ht t p: / / www. t andf online. com/ loi/ t eop20 Essential Oil in Taurus Cedar (Cedrus libani A. Rich) Seeds a Nebi Bilir & Ayse Bet ul Avci a b Forest ry Facult y, Suleyman Demirel Universit y, Ispart a, TR-32260, Turkey b Odemis Vocat ional School of Ege Universit y, Odemis, Izmir, TR-35760, Turkey Published online: 25 Oct 2013. To cite this article: Nebi Bilir & Ayse Bet ul Avci (2013) Essent ial Oil in Taurus Cedar (Cedrus libani A. Rich) Seeds, Journal of Essent ial Oil Bearing Plant s, 16: 4, 538-544, DOI: 10. 1080/ 0972060X. 2013. 813249 To link to this article: ht t p: / / dx. doi. org/ 10. 1080/ 0972060X. 2013. 813249 PLEASE SCROLL DOWN FOR ARTI CLE Taylor & Francis m akes every effort t o ensure t he accuracy of all t he inform at ion ( t he “ Cont ent ” ) cont ained in t he publicat ions on our plat form . However, Taylor & Francis, our agent s, and our licensors m ake no represent at ions or warrant ies what soever as t o t he accuracy, com plet eness, or suit abilit y for any purpose of t he Cont ent . Any opinions and views expressed in t his publicat ion are t he opinions and views of t he aut hors, and are not t he views of or endorsed by Taylor & Francis. The accuracy of t he Cont ent should not be relied upon and should be independent ly verified wit h prim ary sources of inform at ion. Taylor and Francis shall not be liable for any losses, act ions, claim s, proceedings, dem ands, cost s, expenses, dam ages, and ot her liabilit ies what soever or howsoever caused arising direct ly or indirect ly in connect ion wit h, in relat ion t o or arising out of t he use of t he Cont ent . This art icle m ay be used for research, t eaching, and privat e st udy purposes. Any subst ant ial or syst em at ic reproduct ion, redist ribut ion, reselling, loan, sub- licensing, syst em at ic supply, or dist ribut ion in any form t o anyone is expressly forbidden. Term s & Condit ions of access and use can be found at ht t p: / / www.t andfonline.com / page/ t erm s- and- condit ions TEOP 16 (4) 2013 pp 538 - 544 538 ISSN Print: 0972-060X ISSN Online: 0976-5026 Essential Oil in Taurus Cedar (Cedrus libani A. Rich) Seeds Nebi Bilir 1,* and Ayse Betul Avci 2 1 Downloaded by [Ege Universitesi Rektorlugu] at 04:03 28 January 2014 2 Forestry Faculty, Suleyman Demirel University, Isparta, TR-32260, Turkey Odemis Vocational School of Ege University, Odemis, Izmir, TR-35760, Turkey Received 16 October 2012; accepted in revised form 24 January 2013 Abstract: This study was carried out to determine potential of essential oil isolated from seeds of Taurus cedar (Cedrus libani A. Rich), occupied mainly in Turkey. Rate and components of essential oil were studied on the seeds sampled from 25 open pollinated families of each three natural populations of the species in 2009. Heritability and variations within population and among populations were estimated for rate and components of essential oil. Besides, the rate and components of essential oil were compared for altitude, aspect, age and diameter at breast height. Average of essential oil rate was 1.82 %, while there were large differences among populations and within population for the rate. Totally, 137 components were determined, while α-pinene (27.08 %) and β-pinene (20.35 %) were major components in the oil. Significant differences (p<0.05) were found among populations, and generally age, altitude, and diameter classes for the rate and components. Heritability was very low (0.27) for the oil rate, while it was very high (>0.78) for the rate of major components. Results of the study showed importance of selected material for quality and quantity of essential oil. Key words: Cedrus libani, constituent, essential oil, heritability, ratio, seed. Introduction The main commercial product is wood in forestry as known, while there are many commercial products obtained from plant species in forest area. Extraction of essential oil is one of the most important forest products because of its commercial value and using in many industries like medicinal, cosmetics and pesticide industries. For instance, Turkey acquired 19 million USD by essential oil export in 2007 and 25 million USD in 2008 7, while commercial potential of whole the world was about 2.2 billion USD in 2006 6. There are many environmental and genetic factors on quality and quantity of essential oil. However, while genetic studies are very limited, there are many studies on using, components and rate of essential oils in different plant species. *Corresponding author (Nebi Bilir) E-mail: < nebibilir@sdu.edu.tr; nebilir@hotmail.com > Estimation of heritability is one of the most important genetic parameters, used different purposes in essential oil 3,13. Farah et. al. 15 reported that differences could be for rate of essential oil among populations in Myrtus communis. Avci and Bayram 2 emphasized importance of harvesting period and time on essential oil rate in M. communis. Large individual differences were reported for essential oil rate in Eucalyptus camaldulensis by Avci and Bilir 3. The essential oil is isolated from different part of plants. Seed is not an important commercial part of Taurus cedar (Cedrus libani A. Rich). Isolated essential oil from the seeds could be less damage to forests than that of wood or other parts of plants. The purposes of the study are to determine © 2013, Har Krishan Bhalla & Sons Nebi Bilir et al., / TEOP 16 (4) 2013 538 - 544 potential of an alternative forest product in the species for contribution to local and national economy, to estimate heritability for rate and major components and to compare the families and populations for the rate and components. Downloaded by [Ege Universitesi Rektorlugu] at 04:03 28 January 2014 Materials and methods Seed material Seeds were extracted from cones of 25 open pollinated families of each three natural populations (Table 1) sampled from Isparta province in southern part of Turkey at the end of 2009. Essential oil isolation Taurus cedar essential oil was extracted by hydrodistillation method for 3 h using Clevenger type apparatus 25 10 g of the air-dried Taurus cedar seed samples was used to extract essential oil and replicated four times in each family. The volatile oils were stored in dark glass bottles at 4°C until analysis 8. GS-MS analysis of essential oil The Taurus cedar seed extract was analyzed by gas chromatography-mass spectrometry (GS-MS) system in Marmara Research Center of The Scientific and Technological Research Council of Turkey. The GC-MS analyses were performed using a Perkin Elmer GC-MS Auto system XL MS were Turbo Mass. Volatile compounds were separated on Thermo 5MS column (30 m × 0.25 mm- 0.25 µm) using helium gas as the carrier gas which was split 1 mL/min. Oven temperature was programmed from 50°C for 2 minutes, ramp 50 to 240°C at 5°C/min, then hold at 240°C for 5 minutes. The temperature of injector and detector was 200°C and inlet temperature was 175°C. Mass spectra were taken on 70 eV, scan mode was full scan and range 50-450 in 200°C ion temperature. Evaluation process was made using 539 the library “Wiley, NIST and Tutor”. Data analysis The statistical analysis was carried out by SPSS statistical package 23 according to following ANOVA model was used for the analysis: where Y ij is the observation from the i th population of the jth family, ì is overall mean, Pi is the effect of ith population, F (P)j(i) is effect of the jth family in the ith population, and eij is random error. Heritability (h2; the fraction of the variance which is genetic among genotypes) for the essential oil rate and two major components was estimated as 5: where σ2f is the variance among families, σ2e is the variance within family. Variance components, expressed as coefficient of variation among families (CVf) and within family (CVe), were estimated as: CV f = 100σ f / x and CVe = 100σ e / x where x is overall character mean. Results Essential oil rate Average of essential oil rate was 1.82 % in pooled populations, while it was 1.66 %, 1.71 % and 2.09 % in the populations. It varied between 0.73 % and 3.20 % among families (Table 2). As presented in Table 2 avearge essential oil yield was 2.3 ml/tree ranged from 0.1 ml to 7.9 ml. While average of the essential oil yield was the highest (2.90 %) in first population, it was the lowest (1.20 %) in second population (Table 2). Table 1. Some details of studied populations Population No 1 2 3 Location Egirdir Senirkent Sarkikaraagac Latitude (N) Longitude (E) Altitude (m) 37°44’ 38°05’ 37°53’ 30°49’ 30°42’ 31°17’ 1565 1490 1620 Nebi Bilir et al., / TEOP 16 (4) 2013 538 - 544 540 Table 2. Average and range of essential oil rate and yield in the populations Population 1 (25)* Population 2 (25) Population 3 (25) Total Essential oil rate (%) 1.66(0.73-3.20)** 2.09(1.03-3.07) 1.71 (0.87-3.07) 1.82(0.73-3.20) Essential oil yield (ml) 2.90(0.7-7.10) 1.20 (0.2-5.8) 2.70(0.1-7.9) 2.30(0.1-7.9) Downloaded by [Ege Universitesi Rektorlugu] at 04:03 28 January 2014 *; Number of studied genotypes in the populations. **; ranges among families in the parenthesis. There were significant (p<0.05) differences among populations for rate and components according to results of analysis of variance. There were also significant differences (p 0.05) in age, diameter, and altitude for essential rate, while aspect has no significant effect (p>0.05) on that. Negative and significant (p£ 0.05; r=-0.298) correlation was estimated between essential oil rate and altitude. Essential oil components 137 components representing 99 % of the total oil were identified in pooled populations. Only 25 of total components were in all the populations, while 28, 20 and 38 of total components were only in first, second and third population, respectively. Beside, 60 components were identi- fied only in an each family. Major components of pooled populations were α-pinene (29.5 %) and β-pinene (21%). The other major components were 1-hexen-3-yne (5.78 %) and Bicyclo[2.2.1] heptan-2-ol (6.08 %) based on rate and identified family numbers. Examples of GS-MS spectrum of one genotype from each population were given in Fig.1. The ten highest components were presented in together with rate and retention indices in Table 3. There were significant (p<0.05) differences among populations, ages, diameters, and altitude for rates of α-pinene and β-Pinene according to results of analysis of variance. Altitude, diameter and age were significantly (p<0.05) effective on the rates of the α-Pinene and β-pinene based on results of correlation analysis. Population 1 Population 2 Population 3 Fig. 1. Examples of GS-MS spectrum of a genotype from each population Nebi Bilir et al., / TEOP 16 (4) 2013 538 - 544 541 Table 3. The highest percentage and most abundant components in populations Downloaded by [Ege Universitesi Rektorlugu] at 04:03 28 January 2014 Components α-Pinene 1-Hexen-3-yne β-Pinene β-Myrcene Cyclohexene D-Limonene β-Phellandrene Bicyclo[3,1,1]heptan-3-ol trans-Pinocarveol Isopinocarveol Bicyclo[3,1,1]hept-3-en-2-ol 2(10)-Pinen-3-one Borneol 3-Cyclohexen-1-ol 3-Cyclohexene-1-methanol p-menth-1-en-8-ol Bicyclo[3,1,1]hept-2-ene2-carboxaldehyde (1R)-(-)-Myrtenal 3-Caren-10-al Bicyclo[3,1,1]hept-3-en-2-one 3-Diazo-1,7,7-trimethylbicyclo [2,2,1]heptan-2-one trans-2-Caren-4-ol Bicyclo[2,2,1]heptan-2-ol Bornyl acetate Isobornyl propionate RI* Population 1 Rates (%) Population Population Pooled 2 3 populations 932 974 975 989 1027 1028 1028 1141 1141 1141 1146 1163 1168 1178 1193 1193 1197 28.74 (19)** 7.89 (10) 22.27 (15) 1.27 (3) 1.08 (1) 2.81 (2) 4.02 (7) 5.56 (4) 3.81 (2) 3.87 (8) 5.78(17) 2.78 (3) 2.65 (4) 2.93 (10) 4.38(11) 27.40 (17) 4.267 (6) 17.76 (15) 2.576 (1) 8.41 (3) 2.382 (1) 19.97 (1) 5.95 (8) 5.35 (2) 2.17 (6) 3.49 (22) 1.68 (6) 2.24 (7) 3.57 (4) 2.64 (9) 6.45 (6) 33.47 (24) 5.184 (3) 22.82 (23) 1.042 (12) 1.407 (8) 3.1 (4) 3.72 (1) 4.20 (7) 3.85 (7) 0.89 (3) 1.96 (12) 4.00 (14) 1.47 (6) 2.28 (15) 5.54 (2) 1.80 (16) 3.47 (9) 29.3 (60) 5.78 (19) 20.9 (53) 1.63 (16) 3.63 (12) 2.76 (7) 11.84 (7) 4.72 (22) 4.92 (13) 2.92 (11) 2.91 (22) 3.09 (53) 2.00 (15) 2.39 (26) 4.55 (6) 2.45 (35) 4.77 (26) 1197 1197 1211 1211 3.34 (8) 5.77 (5) 1.56 (10) 2.94 (7) 6.31 (3) 2.07 (6) 5.67 (3) 3.18 (4) 2.34 (8) 2.32 (1) 3.98 (18) 4.74 (7) 4.05 (13) 1.99 (17) 1276 1285 1286 1286 1.91 (5) 7.69 (1) 1.87 (11) 1.57 (3) 6.68 (10) 6.36 (2) 2.05 (13) 2.05 (3) 3.88 (17) 4.93 (2) 2.49 (5) 1.84 (11) 6.08 (28) 5.65 (4) 2.14 (29) *; Retention Indice; **: Number of identified family of the components Heritability and variation Heritability was very low for essential oil rate in each population and pooled populations, while it was very high for major components (Table 4). Variation within family was higher than that of among families for essential oil rates in the populations, while it was opposite for the components (Table 4). It could be said that, while rate was under environmental effect, genetic factors was very effective on components based on estimated heritability and coefficients of variations. Discussion Average of essential oil rate was 1.82 %. It ranged from 1.66 % to 2.09 % in the populations, and from 0.73 % to 3.2 % in the families. The large differences were also found in the essential oil production per tree (2.3 ml/tree). The results showed importance of production material in the essential oil. Essential oil rate was reported 1.72 % varied between 0.80 % and 2.75 % in seeds of six populations of Cedrus libani 10. High rate and rich components in essential oil were found in Nebi Bilir et al., / TEOP 16 (4) 2013 538 - 544 542 Table 4. Heritability (h2), coefficient of variation among families (CVf) and within family (CVe) Rate/Components Downloaded by [Ege Universitesi Rektorlugu] at 04:03 28 January 2014 Rate α-Pinene β-Pinene h2 CVf CVe 0.27 0.99 0.99 0.205 0.435 0.449 0.356 0.025 0.029 different parts of C. libani 10,17,18,19,20. The rate was 1.82 % in needles of C. atlantica 12. The rate of essential oil could change according to part of the plants. For instance, rates of essential oil were 0.21 % in cones, 0.41 % in cones, and 0.68 % in wood of C. libani 21. It could be said that essential oil rate was higher in seeds than that of other parts of plants in C.libani. Besides, many environmental and genetic factors were reported on essential oil rate 1,16,24. These results emphasized importance of part and location of plant material in essential oil production. Total number of essential oil components was 137 in pooled populations. It was 73, 64, and 76 in the populations. But, only 25 components of total components were identified in all three populations, while 28, 20, and 38 components were identified in only first, second, and third populations, respectively. Besides, each 61 components were identified in only each family/ tree. These results showed importance of seed source to obtain high components. The major two components were α-Pinene (29.54 %) and βpinene (21.02 %) in pooled populations. The number of components could change according to part of C. libani 17,18,19,20. Lozzio et. al. 21 also reported 28 components in needles, 18 components in cones, 12 components in wood of C. libani. Baser and Demircakmak 4 reported 37 components representing 90-93 % of total essential oil in stem and root of C. libani. The variations emphasize the part and location of plant material in isolation of essential oil. Derwich et. al. 12 identified 31 components, in which major component is, α-pinen (14.85 %) in C. atlantica. Rates of α-pinen were 51 % in cones, 2.2 % in needles and 0 % in wood in C. atlantica 12. Dayisoylu and Alma 11 reported 68.19 % αpinene and 11.91 % β-pinene in cones of Abies cilicica subsp. cilicica. New studies were conducted on utilization of essential oil in seeds of the species 10,14,22. Heritability, which is the fraction of the genetic variance among genotypes was very low 0.27 for essential oil rate, while it was very high (0.9) for the components. The results showed that genetic factors were more effective than environment on components. It could be used for selection of seed sources for quality of essential oil. Heritability was 0.71 in rate of essential oil in Eucalyptus camaldulensis 3. It was 0.5 for 1,8-cineole of Eucalyptus camaldulensis 13. The genetic parameters were used in increasing of quality and quantity of essential oil 9. Conclusions Rate and components are identified quality and quantity of essential oil, while they change among populations and within population. They should be investigated in large populations at family level to select the sources of essential oil. The studied part, seed is not an important commercial material of plants. So, it could be a new material to obtain a new alternative product from the species. Using of essential oil isolated from seed should be investigated in the species. Vegetative propagation should be considered especially for private plantation to obtain essential oil. Seed sources should be selected/established to isolated essential oil in the species. Quality classification should be prepared based on quantity of components in essential oil isolated from seeds of the species. Acknowledgements The authors thank to the Scientific and Technological Research Council of Turkey (TUBITAK) for the financial support (TOVAG110O474). 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