SOLVENT-FREE MICROWAVE EXTRACTION AND MICROWAVE-ASSISTED HYDRODISTILLATION OF ESSENTIAL OILS FROM SPICES

A THESIS SUBMITTED TO THE GRADUATE SCHOOL OF NATURAL AND APPLIED SCIENCES OF MIDDLE EAST TECHNICAL UNIVERSITY

BY

BESTE BAYRAMOĞLU

IN PARTIAL FULLFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE IN FOOD ENGINEERING

SEPTEMBER 2007

Approval of the thesis: SOLVENT-FREE MICROWAVE EXTRACTION AND MICROWAVEASSISTED HYDRODISTILLATION OF ESSENTIAL OILS FROM SPICES

submitted by BESTE BAYRAMOĞLU in partial fulfillment of the requirements for the degree of Master of Science in Food Engineering Department, Middle East Technical University by, Prof. Dr. Canan Özgen ___________ Dean, Graduate School of Natural and Applied Sciences Prof. Dr. B. Zümrüt Ögel Head of Department, Food Engineering Assoc. Prof. Dr. Serpil Şahin Supervisor, Food Engineering Dept., METU Assoc. Prof. Dr. S. Gülüm Şümnü Co-Supervisor, Food Engineering Dept., METU ___________ ___________ ___________

Examining Committee Members: Prof. Dr. Ferhunde Us Food Engineering Dept., HÜ Assoc. Prof. Dr. Serpil Şahin Food Engineering Dept., METU Assoc. Prof. Dr. S. Gülüm Şümnü Food Engineering Dept., METU Prof. Dr. Alev Bayındırlı Food Engineering Dept., METU Ins. Dr. İlkay Şensoy Food Engineering Dept., METU _____________________ _____________________ _____________________ _____________________ _____________________

Date: 05/09/2007

I hereby declare that all information in this document has been obtained and presented in accordance with academic rules and ethical conduct. I also declare that, as required by these rules and conduct, I have fully cited and referenced all material and results that are not original to this work.

Name, Last name: Beste Bayramoğlu Signature :

iii

) were chosen in this study since they have high antimicrobial and antioxidant effects and are widely grown and consumed in Turkey. and other quality parameters of iv . composition. Gülüm Şümnü September 2007. Prof.. Beste M.S. Dr.). S.) and rosemary (Rosmarinus officinalis L. 130 pages The undesirable effects of conventional methods generated the need for economical and safe techniques in the extraction of essential oils. which are thought to overcome this problem. The effects of microwave power and extraction time on the yield. Serpil Şahin Co-Supervisor: Assoc. Prof. Dr. Oregano (Origanum vulgare L. Department of Food Engineering Supervisor: Assoc. The objectives of this study were to examine the applicability of SFME in the extraction of essential oils from oregano and laurel. laurel (Laurus nobilis L. and MAHD in the extraction of rosemary essential oil.ABSTRACT SOLVENT-FREE MICROWAVE EXTRACTION AND MICROWAVEASSISTED HYDRODISTILLATION OF ESSENTIAL OILS FROM SPICES Bayramoğlu. Microwave-assisted hydrodistillation (MAHD) and solvent-free microwave extraction (SFME) are recently developed techniques.

. v . Rosmarinus officinalis L. Hydrodistillation was performed as control. no significant differences were obtained in yields (about 0. Main aroma compounds were 1. Main aroma compound was 1.8-cineole (430-500 mg 1. Microwave-assisted hydrodistillation (MAHD). Process time was reduced by 55-60%. Origanum vulgare L.022 mL oil/g laurel) obtained by SFME and hydrodistillation.8-cineole/mL oil). The process time was reduced by about 65%. SFME offered significantly higher essential oil yields (0. In the case of rosemary.054 mL oil/g oregano) from oregano as compared to hydrodistillation (0.048 mL oil/g oregano). Keywords: Solvent-free microwave extraction (SFME).the extracts were also investigated. Conventional process time was reduced by 80%.. Main aroma compound was thymol (650-750 mg thymol/mL oil).8-cineole (630-730 mg 1.8-cineole/mL oil) and camphor (150-210 mg camphor/mL oil). Laurus nobilis L. For laurel. no significant differences were obtained in yields (about 0.026 mL oil/g rosemary) obtained by MAHD at 622 W and hydrodistillation.

Dr. Mikrodalga yardımlı hidrodistilasyon (MYH) ve çözücüsüz mikrodalga ekstraksiyonu (ÇME). mikrodalga gücünün ve işlem süresinin uçucu yağ verimi ve kalitesi üzerindeki etkisinin vi . defne (Laurus nobilis L. Gıda Mühendisliği Bölümü Tez Yöneticisi: Doç. Gülüm Şümnü Eylül 2007. Bu çalışmada. biberiye uçucu yağı eldesinde kullanılabilirliğinin araştırılmasıdır. Bu çalışmanın amacı.). 130 sayfa Bitkilerden esansiyel yağ eldesi için kullanılan geleneksel yöntemlerin birtakım dezavantajlarının olması. S. bitkilerden esansiyel yağ eldesinde daha güvenilir ve ekonomik yöntemlerin arayışına yönlendirmiştir.) ve biberiye (Rosmarinus officinalis L. araştırmacıları. Türkiye’de çokça yetişmeleri ve tüketilmeleri nedeniyle dağ kekiği (Origanum vulgare L. Serpil Şahin Yardımcı Tez Yöneticisi: Doç. bu dezavantajların etkisini büyük oranda azaltacağı tahmin edilen. oldukça yüksek antimikrobiyal ve antioksidan özelliklere sahip olmaları.ÖZ ÇÖZÜCÜSÜZ MİKRODALGA EKSTRAKSİYONU VE MİKRODALGA YARDIMLI HİDRODİSTİLASYON YÖNTEMLERİ İLE BİTKİLERDEN ESANSİYEL YAĞLARIN ELDE EDİLMESİ Bayramoğlu. ÇME yönteminin kekik ve defne uçucu yağları eldesinde. Dr. Aynı zamanda. Beste Yüksek Lisans.) seçilmiştir. son yıllarda geliştirilmiş yöntemlerdir. MYH yönteminin ise.

Laurus nobilis L. Rosmarinus officinalis L. ÇME yöntemi (0. Origanum vulgare L. Hidrodistilasyon ile ulaşılan işlem süresi % 80 oranında düşürülmüştür. Mikrodalga yardımlı hidrodistilasyon (MYH).8-sineol’den (630-730 mg 1. Kekiğin ana maddesinin timol (650-750 mg timol/mL yağ) olduğu görülmüştür.incelenmesi amaçlanmaktadır.. Kekikte. Biberiyenin ana bileşenlerinin 1.. Defne yağının oldukça büyük bir kısmının 1. Anahtar kelimeler: Çözücüsüz mikrodalga ekstraksiyonu (ÇME).054 mL yağ/g kekik) ile hidrodistilasyona (0. ÇME ve hidrodistilasyon yöntemleri ile elde edilen verimlerde (yaklaşık olarak 0.8-sineol (430-500 mg 1. Defnede. vii .8-sineol/mL yağ) oluştuğu anlaşılmıştır. İşlem süresi % 55-60 oranında kısalmıştır.026 mL yağ/g biberiye) arasında istatistiksel olarak önemli bir fark gözlenmemiştir. Hidrodistilasyon yöntemi kontrol amaçlı kullanılmıştır.048 mL yağ/g kekik) nazaran istatistiksel olarak belirgin derecede fazla uçucu yağı verimi elde edilmiştir.022 mL yağ/g defne) istatistiksel olarak önemli bir fark gözlenmemiştir. Biberiyede ise. İşlem süresinin % 65 oranında azaldığı görülmüştür. MYH yöntemiyle ulaşılan verim ile hidrodistilasyonda elde edilen verim (yaklaşık olarak 0.8-sineol/mL yağ) ve kamfor (150-210 mg kamfor/mL yağ) olduğu belirlenmiştir.

To My Family viii .

Nadide Seyhun and Özge Şakıyan Demirkol are gratefully acknowledged. I would also like to express my gratitude to Fedot Sleptsov. criticism. TUBITAK (The Scientific & Technological Research Council of Turkey) is greatly acknowledged for supporting this project (TOVAG 1040265). Özlem Erçin. ix . Gülüm Şümnü for their guidance. Gül Çalışır and Ezgi Erol for their motivation and moral support they have provided throughout the study. Serpil Şahin and co-supervisor Assoc. The motivation and guidance of the other research group members. My special thanks go to Mecit Oztop for his suggestions. advice. I wish to thank my friends. S. Prof. Finally. SadıkFiliz-Ali Bayramoğlu for their great moral support and belief in me. Tunca Doğan and Elif Özlem Karabacak for helping me in coping with the picking of oregano leaves. Prof. encouragements and insight throughout the research. comments and solid helps during the study. İlkem Demirkesen. Dr. I would like to dedicate this work to my family members. Dr.ACKNOWLEDGEMENTS I would like to express my deepest gratitude to my supervisor Assoc. Mecnun Mert.

.................................25 1........ Specific Gravity....... Refractive Index............................ Hydrodistillation...............................................iv ÖZ....................................................................) ....................2.1.........................................................27 1.....................7....................2.....................14 1.......7....................................... Microwave-assisted Hydrodistillation (MAHD)......................................... Conventional Methods for the Extraction of Essential Oils.......... Composition......................................2.....................28 2.........31 2...........30 2......................................... Optical Rotation............34 x .............................................9 1..............................27 1.................................... Materials.......31 2.................30 2..1..................................2.................25 1......................33 2..........................................................................................xiii LIST OF FIGURES.ix TABLE OF CONTENTS.........................2..............4.......................3...26 1... Quality Parameters of Essential Oils...........................................................2.........2....................................... Solvent-free Microwave Extraction (SFME).................................7.........xvi CHAPTER 1......................1..7.........26 1................TABLE OF CONTENTS ABSTRACT............. Objectives of the Study....7......2..........11 1....... Oregano (Origanum vulgare L........................ Yield............................................1...........................................7................................7.........................6......4................................................) .......................33 2..........................................................................6.................5................................... Methods......................... MATERIALS AND METHODS.....25 1............................19 1.. Rosemary (Rosmarinus officinalis L.5............................8......................... INTRODUCTION............4...............1 1..............................1 1...................... Essential Oils..........)............... Analysis.....17 1............................................. Novel Technologies and Extraction of Essential Oils Using Microwaves...................................... Solubility in Alcohol.............................3..........x LIST OF TABLES. Laurel (Laurus nobilis L............................vi ACKNOWLEDGMENTS.3.............

... Refractive index..1.................................2.........................1..2....................................... Yield....................61 3................ Composition.........................3.) ... Effect of extraction on quality parameters of oregano essential oil......61 3................5.........3..................) ..1....2................................... RESULTS AND DISCUSSION..........2......2..........59 3.... Measurements of Solubility in Alcohol.......................2................2.....6...... Particle Size Distribution............................1............2......1...........3.........2.... Other parameters............1......................2........................................73 3.............3....78 3..........60 3.....4...........3.........................................3...... GC/GC-MS Analysis......2.................55 3...............................1...2.....2.....2...............42 3......... Particle Size Distribution of Oregano Leaves. Refractive index..2.77 3...... Specific gravity...43 3.2....41 3.............. Particle Size Distribution of Rosemary Leaves....45 3............2....................4..........1............1.....2.........2.......................2.....77 3......1............2.1............. IMPI-2L Test.....................3.......2...........................43 3......2. Specific Gravity Measurements........1.........4. Laurel (Lauris nobilis L.........60 3.2.......55 3.............................2.......... Rosemary (Rosmarinus officinalis L.... Composition. Particle Size Distribution of laurel leaves...4........2............................ Yield............4..................63 3.42 3................78 xi ....................................... Refractive Index Measurements. Oregano (Origanum vulgare L......1........2............2.......1...........34 2..... Other parameters...........2.....3.................3............1........40 2.......................5. Specific gravity.1...4...3...............2...........4...) .......... Effect of extraction on quality parameters of laurel essential oil..................40 2....................2.36 2...... Effect of extraction on quality parameters of rosemary essential oil..... Yield...........................2..............42 3.........73 3... Solubility in ethanol..........1...........35 2.....................2..............2..75 3..3............. Statistical Analysis..3.......3..................................2.....40 2..1...........2...57 3............

...2.................. ANOVA AND TUKEY’S PAIRWISE COMPARISON TEST TABLES.............................3.......................93 4.........................................2...................................................... Refractive index........2.....3.......3............ Other parameters.............................3.......................... Specific gravity.3.... Composition........................80 3.......2................3....................................91 3.................. CONCLUSION..........................2...............1............................98 APPENDICES A.91 3....3..96 REFERENCES..124 xii ...............3.....2.......

........................47 Table 2 Compounds present in the essential oil of Laurus nobilis L........124 Table A................................. . (Box-Cox transformed).... obtained by different methods and their concentrations..............125 Table A..... ...........................82 Table A..3 ANOVA results for monoterpene hydrocarbon concentrations obtained in the extraction of Origanum vulgare L.........................................66 Table 3 Compounds present in the essential oil of Rosmarinus officinalis L...................................125 Table A............1 ANOVA results for yield values obtained for Origanum vulgare L......... obtained by different methods and their concentrations........ obtained by different methods and their concentrations..........124 Table A.....6 ANOVA results for specific gravity values obtained in the extraction of Origanum vulgare L................2 Tukey’s pairwise comparison test results for yield values obtained in the extraction of Origanum vulgare L...5 ANOVA results for sesquiterpene concentrations obtained in the extraction of Origanum vulgare L.........4 ANOVA results for oxygenated compound concentrations obtained in the extraction of Origanum vulgare L..125 xiii ....... ...................... ............................................. ....LIST OF TABLES TABLES Table 1 Compounds present in the essential oil of Origanum vulgare L.................124 Table A.................

..........................127 Table A...10 ANOVA results for monoterpene hydrocarbon concentrations obtained in the extraction of Laurus nobilis L... ........................125 Table A...................126 Table A..........................13 Tukey’s pairwise comparison test results for sesquiterpene concentrations obtained in the extraction of Laurus nobilis L........... .. ...................14 ANOVA results for specific gravity values obtained in the extraction of Laurus nobilis L................126 Table A..........................................................................17 Tukey’s pairwise comparison test results for yield values obtained in the extraction of Rosmarinus officinalis L.............................................. .....................127 Table A.......................9 ANOVA results for yield values obtained for Laurus nobilis L........................................................ .......................................127 Table A...16 ANOVA results for yield values obtained for Rosmarinus officinalis L...7 ANOVA results for refractive index values obtained in the extraction of Origanum vulgare L...Table A............................................................ .......127 Table A...........126 Table A..128 Table A................. ........ ..12 ANOVA results for sesquiterpene concentrations obtained in the extraction of Laurus nobilis L...........15 ANOVA results for refractive index values obtained in the extraction of Laurus nobilis L.. ... .........8 ANOVA results for solubility in ethanol values obtained in the extraction of Origanum vulgare L.......... .128 xiv ....................126 Table A......11 ANOVA results for oxygenated compound concentrations obtained in the extraction of Laurus nobilis L................................

................ ....128 Table A...23 Tukey’s pairwise comparison test results for sesquiterpene concentrations obtained in the extraction of Rosmarinus officinalis L..........130 xv ........129 Table A.................................................................18 ANOVA results for monoterpene hydrocarbon concentrations obtained in the extraction of Rosmarinus officinalis L.21 Tukey’s pairwise comparison test results for oxygenated compound concentrations obtained in the extraction of Rosmarinus officinalis L.... ...............................20 ANOVA results for oxygenated compound concentrations obtained in the extraction of Rosmarinus officinalis L.....25 ANOVA results for refractive index values obtained in the extraction of Rosmarinus officinalis L....129 Table A..130 Table A................ ................ .........................................129 Table A...........................Table A.... ...129 Table A..........19 Tukey’s pairwise comparison test results for monoterpene hydrocarbon concentrations obtained in the extraction of Rosmarinus officinalis L............... ...........22 ANOVA results for sesquiterpene concentrations obtained in the extraction of Rosmarinus officinalis L...........24 ANOVA results for specific gravity values obtained in the extraction of Rosmarinus officinalis L........ ..........................................................................128 Table A............... ..

................................................................................................................................... essential oil..............................................46 xvi .........11 Figure 4 Structures of the major components of Laurus nobilis L.....45 Figure 10 Total ion chromatogram (obtained by GC-MS analysis) of the Origanum vulgare L............5 Figure 3 Structures of the major components of Origanum vulgare L............................................LIST OF FIGURES FIGURES Figure 1 Developmental pattern of essential oil glands............................................................................... leaves...............43 Figure 9 Variation of essential oil yield of Origanum vulgare L....................................21 Figure 7 Schematic diagram of the SFME & MAHD processes................................13 Figure 5 Structures of the major compounds of Rosmarinus officinalis L............. essential oil extracted by SFME at 100% power level.......................................................... during hydrodistillation and solvent-free microwave extraction (SFME) at different power levels........................................ essential oil................................................................ essential oil...................32 Figure 8 Particle size distribution of Origanum vulgare L.........................3 Figure 2 Major constituents of essential oils..16 Figure 6 Schematic diagram of a SFE system..................

...................55 Figure 17 Variation of specific gravity of the Origanum vulgare L............52 Figure 15 Variation of component classes with time in the essential oil of Origanum vulgare L....60 xvii ......... obtained by SFME at 100% power level........................... essential oil extracted by different methods..... obtained by SFME at 80% power level..........................56 Figure 18 Specific gravity values of Origanum vulgare L............ essential oil with time obtained by different methods.... obtained by hydrodistillation................. essential oil extracted by different methods............. obtained by SFME at 40% power level.............51 Figure 14 Variation of component classes with time in the essential oil of Origanum vulgare L...........................49 Figure 12 Variation of component classes with time in the essential oil of Origanum vulgare L...............59 Figure 21 Solubility in ethanol values of Origanum vulgare L...............................Figure 11 Variation of component classes with time in the essential oil of Origanum vulgare L.. essential oil extracted by different methods.............................. obtained by SFME at 60% power level.................... with respect to different extraction methods.........53 Figure 16 Variation of compound classes in the essential oil of Origanum vulgare L....57 Figure 19 Variation of refractive index of the Origanum vulgare L.................. essential oil with time obtained by different methods.....50 Figure 13 Variation of component classes with time in the essential oil of Origanum vulgare L...................58 Figure 20 Refractive index values of Origanum vulgare L....................................................

....................................... with respect to different extraction methods........... obtained SFME-40% power level................................ obtained SFME-100% power level.....................73 Figure 30 Variation of specific gravity of the Laurus nobilis L..... essential oil extracted by different methods....................... during hydrodistillation and solvent-free microwave extraction (SFME) at different power levels............................................. leaves... obtained hydrodistillation...........................................................74 Figure 31 Specific gravity values of Laurus nobilis L.........75 xviii .........................................62 Figure 24 Total ion chromatogram (obtained by GC-MS analysis) of the Laurus nobilis L.......................72 Figure 29 Variation of sesquiterpenes in the essential oil of Laurus nobilis L.................... essential oil extracted by SFME at 100% power level...............................69 Figure 27 Variation of component classes with time in the essential oil of Laurus nobilis L..........70 Figure 28 Variation of monoterpene hydrocarbons and oxygenated compounds in the essential oil of Laurus nobilis L.......................... with respect to different extraction methods..............61 Figure 23 Variation of essential oil yield of Laurus nobilis L.. essential oil with time obtained by different methods....................................Figure 22 Particle size distribution of Laurus nobilis L............68 Figure 26 Variation of component classes with time in the essential oil of Laurus nobilis L............................................................................65 Figure 25 Variation of component classes with time in the essential oil of Laurus nobilis L.

... essential oil extracted by MAHD at 100% power level.................. obtained MAHD-40% power level.................... leaves...........89 Figure 41 Variation of specifiv gravity of the Rosmarinus officinalis L..... with respect to different extraction methods..............................87 Figure 39 Variation of component classes with time in the essential oil of Rosmarinus officinalis L............Figure 32 Variation of refractive index of the Laurus nobilis L...............78 Figure 35 Variation of essential oil yield of Rosmarinus officinalis L...........77 Figure 34 Particle size distribution of Rosmarinus officinalis L................... obtained hydrodistillation........76 Figure 33 Refractive index values of Laurus nobilis L................ essential oil with time obtained by different methods......88 Figure 40 Variation of compound classes in the essential oil of Rosmarinus officinalis L..... during hydrodistillation and microwave-assisted hydrodistillation (MAHD) at different power levels............................................... essential oil with time obtained by different methods.79 Figure 36 Total ion chromatogram (obtained by GC-MS analysis) of the Rosmarinus officinalis L......92 xix ................................. essential oil extracted by different methods.......... obtained MAHD-100% power level.............................86 Figure 38 Variation of component classes with time in the essential oil of Rosmarinus officinalis L.....................................................81 Figure 37 Variation of component classes with time in the essential oil of Rosmarinus officinalis L.......................

......94 Figure 44 Refractive index values of Rosmarinus officinalis L..........93 Figure 43 Variation of refractive index of the Rosmarinus officinalis L......95 xx ............................................ essential oil with time obtained by different methods..............................Figure 42 Specific gravity values of Rosmarinus officinalis L........................... essential oil extracted by different methods.. essential oil extracted by different methods...................................

twigs.1. Arnald de Villanova. essential oils were being made by pharmacies and their pharmacological effects were described in pharmacopoeias.. Brunschwig and Reiff. Quinta essentia. their use does not appear to have been widespread in Europe until the 16th century (Crosthwaite. 2001). India and Persia (Guenther. However. 1948). rosemary. Guenther (1948) gave the names of two Strassburg physicians. juniper wood. spike (lavender). Spices have been used for their perfume. 1948) more than 2000 years ago and it was improved in the 9th century by the Arabs (Bauer et al. 1948). 1998). buds. Essential Oils Essential oils are defined as “aromatic oily liquids” which can be obtained by a number of techniques from the leaves. only oil of turpentine among the known essential oils was mentioned by Greek and Roman historians (Guenther. Guenther (1948) also called essential oils as “volatile or ethereal oils” and stated that the effective component of a drug.. (2001) states that by the 13th century. fruits and roots of the plant materials (Burt. The first authentic description of the distillation of real essential oils is generally ascribed to the Catalan physician.CHAPTER 1 INTRODUCTION 1. barks. woods. who may be said to have introduced the art of distillation into recognized European therapy (Guenther. seeds. who published separately in that century on the distillation and use of essential oils and mentioned the oils of turpentine. 1 . flavour and preservative properties since antiquity (Bauer et al. 2004). Bauer et al. herbs. 2001). Distillation was first used as a method of producing essential oils in Egypt. However. which was named by Paracelsus von Hohenheim (Swiss reformer of medicine in the 16th century) is thought to be the source of the term “essential oil”. flowers.

perfumes (fragrances and aftershaves) and pharmaceuticals (for their functional properties). soaps. although it is likely to have been used by the native Australians before that (Burt. either extracted from plant material or synthetically manufactured. Aromatherapy is known to constitute slightly more than 2% of the total market (Burt. pharmaceuticals. 2004). Joseph Du Chesne stated. 2004). the preparation of essential oils was well known in the 17th century and pharmacies generally stocked 15–20 different oils (Guenther. mace.1993). Van de Braak and Leijten (1999) claimed an estimated 3000 essential oils are known. Individual components of essential oils are also used as food flavourings. Today. toilet products. and also in insecticides. essential oils are used in a wide variety of consumer goods such as detergents. nutmeg. 2004). of which about 300 are commercially important adding that the greatest use of essential oils in the European Union (EU) is in food (as flavourings). soft and hard drinks. 2004). As the French physician. The essential oils in aromatic herbs are known to be largely located within the glandular structures that develop on the surface of leaves and other organs of the plants (Gaspar and Leeke. The first experimental measurement of the bactericidal properties of the vapours of essential oils is said to have been carried out by De la Croix in 1881 (Burt. Guenther (1948) stated that in the 19th and 20th centuries the use of essential oils in medicine gradually became secondary to their use for flavour and aroma. confectionary food products.clove. cosmetics. Bosabilidis and Tsekos (1984) stated that the peltate hairs appear to contain most of the oil and is henceforth called ‘the glands’. 1948). The use of tea tree oil for medical purposes has been documented since the colonisation of Australia at the end of the 18th century. The world consumption of essential oil and perfumes is increasing very fast (Singh.1 illustrates the development of these 2 . perfumes. However. Fig. anise and cinnamon.

etc. rates of plant metabolism. phenols. 2004).) are the principal odor carriers. ketones. lactones.etc (Singh. the oxygenated compounds (alcohols. Major constituents of most of the essential oils are hydrocarbons (terpenes. The foot and stalk cell remain unicellular throughout the subsequent development of the gland while the mother cell of the head further divides to give rise to 8 or 12 head cells (HC) by the end of the development (Gaspar and Leeke. unsaturated 3 . etc. contribute in some degree to the total odor and flavor of the oil (Guenther.). esters. aldehydes. Jerkovic et al. sesquiterpenes. esters. acids. Developmental pattern of essential oil glands (Gaspar and Leeke. The lower cell corresponds to the foot cell (FC) while the upper daughter cell re-divides to yield the stalk cell (SC) and the mother head cell (MHC). lactones. 1993) as shown in Fig 2. too. 1948). The pattern is as follows: Each gland originates from a single protodermal cell that undergoes division and derives two unequally sized cells. Fig 1. alcohols. (2001) claimed that climatic factors. aldehydes. ethers. On the other side.glands. ketones. The oxygenated compounds are more stable against oxidizing and more soluble in dilute alcohol as compared to other constituents. differentiation and secretory activity of glandular hairs affect synthesis and secretion of essentail oils. 2004). although the terpenes and sesquiterpenes. Among these. phenols.

1989) and Satureja douglasii (Peer and Langenheim. 1988. 2004)... 1948).constituents like monoterpenes and sesquiterpenes have tendency to oxidize or resinify easily under the influence of air and light (Singh. 1993) or under improper storing conditions which means spoilage of odor and flavor. Circella et al. 1993). 2002). There are also several reports in the literature indicating that the essential oil composition of Origanum species varies with the season or day length (Putievsky et al.. Ioannidis et al. 1997). 4 . exerts similar effects in Thymus vulgaris (Tanaka et al. thermal stability are of paramount importance in technology development of oxygenated compounds (Singh. 1999.. (1999) showed that enantiomers of essential oil components exhibited antimicrobial activity to different extents. The knowledge of individual constituents and their physical characteristics such as boiling point. Mentha spicata (Karousou et al.. 2000).. UV-B radiation has been shown to affect content and composition in Ocimum basilicum (Johnson et al. The strongest antimicrobial activity is generally attained in the essential oils produced from herbs harvested during or immediately after the flowering stage (McGimpsey et al. vapour-pressure-temperature relationship. acting through phytochrome photoreceptors. 1999). Essential oil composition within many species of a plant can significantly differ depending on developmental and environmental factors (Johnson et al. For example. 1994.. The composition of essential oils from different parts of the same plant can also differ widely (Burt. Marino et al. 1998) and Mentha piperita (Maffei and Scannerini. 1998). 1997.. LisBalchin et al. 2004).. 1995.. and lowering of the solubility in alcohol (Guenther. Kokkini et al. and visible radiation.

Major constituents of essential oils 5 .ESSENTIAL OILS Terpenes Esters Ethers Ketones Aldehydes Hydrocarbons Lactones Phenols Acids Alcohols 5 Benzoic Acetic Salicylic Cinnamic Cineole Anethole Safrole Cymene Stroene Cumarin Eugenol Thymol Carvacrol Limonene Cedrene Camphene Pinene Camphore Carvone Menthone Irone Methylnonyl Pulegone Ferrectone Thujone Vanillin Citral Citronelial Benzaldehyde Cinnamldehyde Cinnamic Aldehyde Benzoic Cinnamic Myristic Isovaleric Geraniol Menthol Citroneliol Linalool Terpineol Borneol Fig.2.

2000. 2004). Stecchini et al. turmeric (Negi et al. Smith-Palmer et al. clove and thyme (Farag et al. The antimicrobial activities of the essential oils of a variety of plant materials against many food borne pathogens have been tested up to now. 1989) and Listeria monocytogenes (Smith-Palmer et al. Prudent et al. oregano and thyme oils were found to be effective at levels of 5–20 µL g-1 in inhibiting L. 1999.. Farag et al.b. 1998a. 1995.). Skandamis and Nychas. 1998). coli (Burt and Reinders. aureus (Hammer et al.. Cosentino (1999) stated that the phenolic components are mainly responsible for the antibacterial properties of essential oils. 1999.... As for fish dishes. 1995).. the activity of rosemary oil against Escherichia coli was proved (Farag et al. Pintore et al. Sage..Until the early 1990s. 1999.. oregano oil was found to be 6 . 2002).. 2003) essential oils are among the ones having antibacterial properties. 1999. Smith-Palmer et al. 2003.. Rosemary essential oil has also been shown to possess antimicrobial activity against Salmonella typhimurium ( Hammer et al. Bacillus cereus (Chaibi et al... 1999) and S..... Hammer et al. 1984. Hammer et al. a fair number of trials have been carried out with essential oils in foods (Burt.. Lemay et al.. 1999). very few studies of the activity of essential oils in foods had been published (Board and Gould. Gill et al. as well. Prudent et al... Smith-Palmer et al. 1998.. Hammer et al. lemongrass (Hammer et al....1998. typhimurium (Hammer et al.. Certain oils possess stronger antimicrobial properties than others for meat applications (Burt.. Since then. mint and sage oils were less effective or ineffective (Shelef et al. 1995).. In addition. 1991). 2001) whilst mustard. 1992.1998. clove. 1997). S. oregano essential oil is active against some food borne pathogens such as E. For instance. 2002). A. 2002. cilantro. sometimes causing a marked initial reduction in the number of recoverable cells (Aureli et al. 1999). Hammer et al. Staphylococcus aureus (Pintore et al. 1999. monocytogenes. 1993... 2004). 2002. Tassou et al. Hao et al. Tsigarida et al. hydrophila and natural spoilage flora in meat products..1989. Eugenol and coriander. applying the same concentrations.1989. 1999) and tea bush (Bassole et al.

Wan et al. 2002). 2000).. Bendini et al. 2000.1–10 µL g-1 in washing water (Singh et al. All essential oils (thyme.1998). Shelef et al. Carvacrol and cinnamaldehyde were very effective in reducing the viable count of the natural flora on kiwifruit when used at 0. The 7 . thyme and basil (Lee and Shibamoto.15 µL g-1 in dipping solution. and rosemary and cinnamon (Dang et al. cereus in rice was ineffective. The antioxidant properties of 98 individual terpenoids and phenol derivatives present as components of essential oils were compared by studying their ability to inhibit the formation of linoleic acid peroxides (Ruberto and Baratta.. Mint oil inhibited the growth of yoghurt starter culture species at 0. Faleiro et al. whereas carvacrol at 0. Ouattara et al.. 2001) were found similar to those of synthetic antioxidants.. Thyme oil also showed antibacterial activity againts Pseudomonas putida on cooked shrimps and against natural flora both on surface and in flesh of the whole Asian sea bass (Harpaz et al. 1999).. 2002). 1995.15– 0.. 1995). enteritidis in low fat yoghurt and cucumber salad (Tassou et al. Sage oil at 0. 1984).75 µL g-1 was very effective in extending the lag phase and reducing the final population compared to control (Ultee et al. 2001). Many other studies were made searching for the antibacterial effects of various essential oils and their components on rice and fruits. The antioxidant activities of essential oils from savory (Gulluce et al. 2005.oregano) and their components (basil methyl chavicol. as well (Capecka et al. 2002.. Speaking of dairy products.5 µL g-1 when used against B. 2003. but less effective on honeydew melon (Roller and Seedhar.05–5 µL g-1 but cinnamon. mint oil at 5–20 µL g-1 was found to be effective against S. 2003).more effective than mint oil (Tassou et al.. cardamom and clove oils were much more effective (Bayoumi..2–0.. cinnamaldehyde. 1992).. thymol) that have been tested on vegetables appeared effective against the natural spoilage flora and food borne pathogens at levels of 0. cloves and nutmeg (Dorman et al.. 2002). Koutsoumanis et al. 2005... Essential oils have been shown to possess antioxidant properties.. 2000).

The United States Food and Drug Administration (FDA) has classified the substances as generally recognised as safe (GRAS) or as approved food additives. essential oils have been demonstrated to exhibit antifungal (Voda et al. In addition to their antibacterial and antioxidant properties. 2007. 1992.. Viuda-Martos et al... 2005). 1995). which are considered to present no risk to the health of the consumer and include amongst others carvacrol. New methods are being developed for making food safe and ‘green’. 2003. citral. Burt (2004) stated that a number of essential oil components have been registered by the European Commission for use as flavourings in foodstuffs. thymol. There is an increasing demand for safer and more natural food.. Karpouhtsis et al. 2004). 2000. 2004). 2003. 1988) ... insecticidal (Konstantopoulou et al. Pessoa et al. 2000). Akgül and Kıvanç. In other countries and if added to food for a purpose other than flavouring. Expensive safety and metabolic studies would be necessary for the approval of an essential constituent as a food additive (Burt. carvone. α-terpinolene.antioxidant activities of eugenol. Estragole and methyl eugenol were deleted from the list in 2001 due to their being genotoxic (Commission Decision of 23 January.. Cyclic monoterpene hydrocarbons with two double chains (γ. these compounds may be treated as new food additives. eugenol. owing to their components. Burt. too.and α-terpinenes. limonene. 1998) properties. Smid and Gorris (1999) suggested the use of a whole spice or herb or a whole essential oil as an ingredient than to use individual 8 . antimycotic (Mari et al. and sabinene) were shown to have very high activity (80-98%) (Ruberto and Baratta. carvacrol. Daferera et al. p-cymene. 2000). 2002). menthol and thymol.. cinnamaldehyde. One such possibility is the use of essential oils as antimicrobial and antioxidant additives. 2002. antiviral (Bishop. and methoxyphenol were similar to that of α-tocopherol. antiparasitic (Pandey et al. The presence of these compounds in essential oils determines their antioxidant properties (Misharina and Polshkov.

essential oil components in those countries for the sake of economical considerations. The use of essential oils in consumer goods is expected to increase in the future due to the rise of ‘green consumerism’, which stimulates the use and development of products derived from plants (Tuley de Silva, 1996). Bassett et al. (1990) stated that this applies to the food and cosmetic sectors as well as medicinal products. International standardisation of the composition of commercially available essential oils would be necessary for reliable applications (Carson and Riley, 2001).

1.2. Oregano (Origanum vulgare L.)

Origanum is used throughout the world as a very popular spice, under the name ‘oregano’. Despite the fact that the name 'oregano' is given to many species of various genera, most oregano spice products originate from species of the genus Origanum (Veres et al., 2003) belonging to the family, Lamiaceae (Souza et al., 2007). The genus Origanum is an annual, perennial and shrubby herb that is native to the Mediterranean, EuroSiberian and Irano-Siberian regions (Sahin et al., 2004). Both morphologically and chemically the genus Origanum is very diverse and many transitional forms occur worldwide (Veres et al., 2003). Ietswaart (1980), based on morphological criteria, recognised 3 groups, 10 sections, 38 species, 6 subspecies and 17 hybrids within the genus. Skoula et al. (1999) stated that the members of the genus are mainly distributed along the Mediterranean region while 75% of them are restricted to the East Mediterranean. 16 species are considered as endemic for the flora of Turkey (Sahin et al., 2004). Origanum species grow abundantly on stony slopes and in rocky mountain areas at the altitudes of 0–4000 m. Origanum plants belonging to different 9

species and ecotypes (biotypes) are widely used in agriculture and the pharmaceutical and cosmetic industries as a culinary herb, flavouring substances of food products, alcoholic beverages and perfumery for their spicy fragrance (Sahin et al., 2004). The essential oil of origanum has been shown to possess antimicrobial, antioxidant, and antimutagenic activities (Baratta et al., 1998; Kanazawa et al., 1995; Dapkevicius et al., 1998 ). Origanum vulgare L. is the most variable species of the genus and the only one commonly known as `oregano' in most European countries (D’Antuono et al., 2000). Bertelli et al. (2003) stated that it is the most important species in Europe and is widely distributed in Europe and Asia up to a height of 2000 m. In the literature, major components of the Origanum vulgare L. essential oil are specified as carvacrol, thymol, γterpinene, p-cymene with approximate compositions of trace-80%, trace64%, 2-52% and trace-52%, respectively (Lawrence, 1984; Prudent et al., 1995; Charai et al., 1996; Sivropoulou et al., 1996; Kokkini et al., 1997; Russo et al., 1998; Daferera et al., 2000; Demetzos and Perdetzoglou, 2001; Marino et al., 2001). Moreover, these compounds are among the ones responsible for the antibacterial activity of the oil (Burt, 2004). Thymol and carvacrol had been proved to be active against E. coli, S. typhimurium, S. aureus, L. monocytogenes and B. cereus (Lambert et al., 2001; Pol and Smid, 1999; Cosentino et al.,1999; Kim et al., 1995). Due to their phenolic nature, thymol and carvacrol exhibits antioxidant properties (Faleiro et al., 2005), as well. The structures of the main constituents of Origanum vulgare L. essential oil are shown in Fig 3.

10

CH3 OH

CH3

CH3

CH3

OH H3C CH3 H3C CH3 H3C CH3 H3C CH3

Carvacrol

Thymol

p-Cymene

γ−Terpinene

Fig. 3. Structures of the major components of Origanum vulgare L. essential oil

1.3. Laurel (Laurus nobilis L.)

Laurus nobilis L., commonly known as laurel, bay or sweet bay, is an evergreen tree or shrub belonging to the family Lauraceae and it is native to the Mediterranean region and Turkey (Kosar et al., 2005a). The species has been esteemed since ancient times. It was dedicated to Apollo, the ancient Greek god of light, and was a symbol of peace and victory used to make wreaths for emperors, generals, and poets (Conforti et al, 2006). Currently, the plant is cultivated in many Mediterranean countries (Kosar et al., 2005a) and the leaves are harvested principally in Turkey from wild growing plants (Akgül and Kıvanç, 1989). In 2000, Turkey exported 3600 tons of L. nobilis leaves with U.S. $7 500 000 income (Kiliç et al., 2004). Laurel leaf, traditionally, has been used as herbal medicine to promote perspiration, to treat rheumatism, earaches, indigestion, sprains (Fang et al., 2005) and dermatitis (Kiliç et al., 2004). However, with respect to its dosage, care has to be taken because of its allergic effect (Pino et al., 1993). Recent research showed that it can be used in treating diabetes and preventing migraine (Duke, 1997).

11

and it was affirmed that the composition of laurel leaf oil depended upon the geographical origin of the leaf (Bouzouita et al. 12 . 2001). 2001). It is also known that benzene compounds such as eugenol. 1993. 1987) has reviewed the compounds identified by different authors... aromatic-eucalyptaceous odor and a fresh. 2002). and Lawrence (1978. It is known that the essential oil is used widely in the perfume and soap industries (Kosar et al. France. and confectionery (Diaz-Maroto et al. delicate spicy. 1989). Dried laurel leaves and their essential oil possess a strong. 2005a) as well as in drugs (Kilic et al. The essential oil of laurel leaves has a freshgreen. present in percentages ranging between 1% and 12%.. meats. and elemicin. 1980... 1. 1992. Pino et al. (1993). 2004). spicy aroma. 1983. Biondi et al.) has been used as a spice since antiquity. and several monoterpene hydrocarbons such as β-pinene and sabinene (Fig... α-terpinyl acetate. soups. 2001). strong. 2002). sweet-spicy..4) (Diaz-Maroto et al. camphoraceous.. Generally. The essential oil of laurel leaf has been the subject of several studies.eucalyptaceous taste (Braun et al. Italy and Spain) was reported by Pino et al. sauces. and they are therefore widely used as flavor enhancers for foods such as fish (Bouzouita et al. are responsible for the spicy aroma of laurel leaves and are extremely important factors determining the sensory quality of the leaves (Borges et al. 2004). primarily because of its essential oil content (Akgül and Kıvanç. with percentages ranging between 30% and 56%. followed by linalool. The soaps are known to be good for acne and have an antidandruff activity (Kiliç et al. Another comparative study of chemical composition of laurel leaf from different geographical origins (Albania. methyl eugenol.. 1993).Laurel leaf (Laurus nobilis L.8cineole was found to be the major component of essential oil of laurel leaves. 1981.

2003. 1995).. aureus. Typhimurium and L. antibacterial. 2000). essential oil Several studies have evaluated the potential role of laurel essential oil as an antimicrobial agent (Bouzouita et al. typhimurium.. S. 13 . cereus (Cosentino et al. 2003. For instance.. S. 2004). 2004).. S. coli. 2005). 4. and inhibitory effects on alcohol absorption (Yoshikawa et al. Monocytogenes (Kim et al..1999) and eugenol was active against E. Simic et al. 2005) up to now... Sesquiterpene lactones identified in laurel leaves were found to have different pharmacological properties including inhibitory effects on NO (nitric oxide) production (antiinflammatory) (Matsuda et al.CH3 CH3 OH CH3 CH3 CH3 H3C H3C H2C CH3 CH3 O CH2 H3C O CH3 O H3C CH3 H3C CH3 1. and also the antioxidant properties of some leaves extracts (Simic et al. α-terpineol was found to be active against E. Structures of the major components of Laurus nobilis L. L. These properties are other factors for laurel to be used in the food industry as a food preservative (Kilic et al. coli.8-cineole linalool alpha-terpinyl acetate beta-pinene sabinene Fig. and antiinflammatory (Fang et al. Skerget et al.. Laurel leaves are also known to have pharmacological activities such as antifungal. antidiabetes. monocytogenes and B.... among the constituents of the essential oil of Laurus nobilis L. 2000).

5 m (Atti-Santos et al. which means dew. it is an aromatic plant with an intense pleasant smell... 2006). Egypt. which comprises up to 200 genera and about 3. Rosmarinus officinalis L.. and a long flowering season extending from April to August (Lo Presti et al. Additionally. Rosemary (Rosmarinus officinalis L. It is used in cosmetics. with a range of varieties and forms (Atti-Santos et al.. It belongs to the Lamiaceae family. is used fresh. France... Greece.. Spain. humus-poor soil. is one of the most spread species of the genus (Pintore et al. 2002) The main producers of rosemary oil are Turkey.) Rosmarinus officinalis L.. arid. Portugal and North Africa (Atti-Santos et al. 2002). dried or as the essential oil (Bauer et al. 2005). Usually the plant is clonally propagated because of the poor germinability of its seeds and the genetic diversity of the seedlings (Flamini et al.500 species. Italy. 2002) and the only one that grows naturally in the Mediterranean regions among the other species (Angioni et al. The name Rosmarinus comes from Latin ros-roris.4. Japan. grows in every soil type. 1948). 1990). the leaves are used as a tonic 14 .1.. as there are several species within the Genus Rosmarinus. 2005).. 2005). Because of its rusticity. dark green lavender-like leaves. hepatoprotective and antitumorigenic activity and for flavoring food (Ramírez et al. The botany of rosemary is rather complex. Rosmarinus officinalis L. Rosmarinus officinalis L. but prefers a sandy. herbal soft drinks. The leaves are used in the preparation of alcoholic beverages (vermouth). is a small evergreen shrub which grows wild in most Mediterranean countries. calcareous. 2005). reaching a height of 1. and cooked foods and sauces (Flamini et al.. Dalmatia. in traditional medicine for its choreretics. and some of the European Union countries are the principal importers (Flamini et al. while the United States. 2002). 2004). it was also called ‘antos’ by the ancient Greeks. which is ‘the flower’ for excellence or ‘libanotis’ for its smell of incense (Guenther.

. the oil is used in perfumery and as a component of disinfectants and insecticides as well (Lo Presti et al. Yugoslavia. 1995). The chemical composition of the oil was reported for the first time by Chalchat et al. and peripheral vascular disorders in folk medicine (Lo Presti et al. (1992). however the highest quality essential oil is obtained from the leaves (Lo Presti et al. France) (Lawrence. Greece. Tunisia. borneol. The essential oil is used as a seasoning for foodstuffs such as meat dishes. 2005). 2005). asthenia. The structures of the main constituents are shown in Figure 5. collected from spring to late autumn (Flamini et al. Turkey. Rezzoug et al. 1998) and oils with approximately equal ratios (20–30%) of 1. α-pinene and camphor (oils from France. who identified 48 constituents. Spain. salami. Elamrani et al. Greece. climatological environment. Bulgaria) (Lawrence. part and age of the plant. 1997.8-cineole (eucalyptol).8-cineole (oils from Morocco. and isolation method. camphor. twigs. (2000) stated that many factors can influence the essential oil yield of rosemary. comprising leaves.for blood circulation and the nervous system. specifying those factors as heredity.. A large amount of research has been carried out on this plant in terms of chemical composition and the variability of the qualitative and quantitative composition of the essential oil was also attributed to these factors (Lo Presti et al.. 2005).8-cineole. Besides.. 2002). 2005). and sauces.. 2005). myrcene.. and flowers. Italy. for chronic weakness.. 15 . Italy. α-pinene. (2002) claimed that two major types of rosemary oil can be distinguished with respect to these main constituents: oils with over 40% of 1. and p-cymene (Katerinopoulos et al. Since it is characterized by unique aromatic characteristics and balsamic properties. The major components of the oil were reported as 1. several studies investigated the composition of rosemary essential oil. Pintore et al. The plant parts used for essential oil production are generally the flowering aerial tops.

.. 2000). Caccioni and Guizzardi. As mentioned before. the oil exhibited insecticide properties and in vitro antifungal activity against Ascosphaera apis (Larràn et al.8-cineole. essential oil Rosmarinus officinalis L.. Rosemary essential oil exhibited good microbicidal activity against mycetes and Gram-positive and Gram-negative bacteria and the main active components were reported as 1. rosemary 16 . and pinenes (Hethelyi et al. 1997). In addition.. 5. camphor. Structures of the major compounds of Rosmarinus officinalis L. 1994. 1989.. Perrucci et al. 1993.8-cineole Fig. 2001). Biavati et al. 1994..CH3 CH3 CH3 OH H3C CH3 OH CH3 H3C CH3 alpha-pinene p-cymene borneol H3C CH2 CH3 CH2 H3C H3C CH3 O CH2 H3C CH3 camphor myrcene 1. Panizzi et al. has also been shown to exhibit antibacterial activity (Valero and Salmerón.. 2003) and to contain several antioxidants (Bicchi et al.

.5. Rosemary was considered as both lipid antioxidant and metal chelator (Nozaki. It is extensively used for wide variety of oil bearing plant materials such as seed. 1. It is the only spice commercially available for use as an antioxidant in Europe and the United States (Yanishlieva et al. root and wood having high boiling 17 .) including carnosol. 1989). Steam distillation is known to be commercially the most common method for the extraction of essential oils. due to the presence of phenolic diterpenes such as rosmarinic acid (Flamini et al. Thorsen and Hildebrandt (2003) stated that the quality as antioxidant and the price of commercial rosemary extract was highly correlated to the content of primarily carnosic acid and secondly to the total content of phenolic diterpenes (rosmanol... Before such substances can be analyzed. the final product (extract) was not considered as essential oils because of the presence of non-volatile constituents. A range of commercial products containing extracts of rosemary are available. etc.extracts show antioxidative properties. some of them are combined with tocopherols (Yanishlieva et al. steam distillation (SD) and simultaneous distillation-extraction (SDE). others are oil soluble. 2006). epirosmanol.. and in order to exploit the synergistic effect.. These conventional methods include hydrodistillation (HD). 2006). 2002). Conventional Methods for the Extraction of Essential Oils Essential oils are complex mixtures of volatile substances generally present at low concentrations. 1997). Various different methods have been used for that purpose up to now. they have to be extracted from the plant matrix (Deng et al. Although soxhlet extraction (SoE) and solvent extraction (SE) have been used to extract neutraceuticals from plant materials. Rosemary extracts were found also to scavenge superoxide radicals (Basaga et al. 2006). some of the products are water dispersible.

The resulting vapour. (2005) stated that hydrodistillation achieves component isolation according to their degree of hydrosolubility rather than to their boiling points. this method is believed to protect the oils extracted to a certain degree. It is suitable for plants which can easily agglutinate and form lumps with live steam (Singh. heating and saturating it with water. 1993). The mixture of water vapor and essential oil vapors is condensed in the condenser and finally collected in a receiver. In hydrodistillation. since the surrounding water acts as a barrier to prevent them from overheating (Sovová and Aleksovski. 2006). otherwise low oil yields may result (Singh. introduced in 1964 by 18 . which supports the charge of the plant material.constituents (Singh. 2006). simultaneous distillation–extraction. Temperature of volatilization is lowered by injection of steam into a charge. Lo Presti et al. the plant material is completely immersed in boiling water (Sovová and Aleksovski. Hydrodistillation with a modified Clevenger apparatus is another widely used conventional method to obtain the essential oil from plant materials. passes out at the top and is conveyed to a condenser (Sovová and Aleksovski. high-pressure steam is passed in at the base of a still into the space beneath a perforated grid. In steam distillation. it is advantageous in seperating heat sensitive materials. 1993). The water is boiled by application of heat and diffusion of essential oils and hot water through the plant membranes occurs (Guenther. Additionally. contrary to the steam distillation. 1948). Among the several techniques that have been developed to isolate volatile compounds. 1993). Steam distillation is used to seperate mixtures at a temperature lower than the normal boiling points of their constituents. The components are isolated in accordance with their boiling points. It is always desirable to use low pressure steam for thermally sensitive oil. Therefore. The steam passes through the plant. even distribution of plant materials in the still is necessary. In order to reduce the channeling. 2006). a mixture of steam and essential oil vapors.

The method has been successfully applied in the extraction of essential oils (Godefroot et al.. Quality of essential oils is of paramount importance in the food industry. aroma compounds (Blanch et al. the yield and quality of essential oils and their isolates are significantly governed by the way they are extracted and processed (Singh. The extracts obtained by simultaneous distillation-extraction are free from non-volatile materials (Teixeira et al. Moreover. 1999. 1998) from numerous matrices. Barták et al.. On the other hand. 1981. Losses of some volatile 19 .Likens and Nickerson. 1996) and other volatile products (Careri et al. this technique allows to carry out the analysis without a sample clean-up step. 2001).. Eikani et al. 2005).. 2004a. Novel Technologies and Extraction of Essential Oils Using Microwaves Improvement in production technology is an essential element to improve the overall yield and quality of the product. Essential oils used in drug and pharmaceutical industries are acceptable only if they pass pharmaceutical tests. 1993). properties like odour and taste become necessary in the case of perfumery and flavory industries and hence they have to pass organoleptic testings. It is known that conventional methods used for the extraction of essential oils and extracts from plant materials have some disadvantages mainly concerned with the quality of the final product. 2007). as well. given its particular characteristics. This technique combines steam distillation together with continuous extraction with a solvent or a mixture of solvents (Chaintreau. Ramos et al. 2000.. Evidently.6. 2007). 1. Stashenko et al... is one of most widely employed (Teixeira et al.. This one-step extraction technique is less time consuming and allows a greater reduction of solvent volumes due to the continuous recycling..

time and energy. Carbon dioxide is generally used as a solvent in supercritical fluid extraction. 2004) and flavors (Giannuzzo et al. low extraction efficiency. Solvent is easily recovered from phases for reuse by making small variations in temperature or pressure or both. degradation of unsaturated or ester compounds through thermal or hydrolytic effects and toxic solvent residue in the extract may be encountered using these extraction methods (Ferhat et al.. Its critical conditions are 31° C. Novel technologies developed for obtaining neutraceuticals from plant materials include ultrasound-assited extraction (UAE) (Vinatoru. Sonsuzer et al.. whereas.. Moreover. which use less solvent. and solvent-free microwave extraction (SFME) (Lucchesi et al. Smith. These shortcomings have led the researchers to develop new technologies. soluble components are extracted from the sample mixture. transport properties (viscosity.47 g/cm3 which permits extraction at around 40° C. 1993). Recently developed techniques such as microwave-assisted hydrodistillation (MAHD) (Stashenko et al. accelerated solvent extraction (ASE) (Kaufmann & Christen. Each method have its own advantages and disadvantages. 2002.compounds. depending upon the nature of plant materials (Singh. When the supercritical fluid and sample mixture are brought into contact.8 bar and 0.. raffinate and extract phases are separated. 2002). 72. and microwave-assisted extraction (MAE) (Kaufmann & Christen. 2003). essential oils (Coelho et al. 2007). especially lipids (Bernardo-Gil et al. these extraction procedures are time-consuming.. 2001). Wang 20 . diffusivity) and compressibility are like those of gases. 2002). 2004b). 2001). supercritical fluid extraction (SFE) (Lang & Wai.. Density and solvent power of supercritical fluids are like those of liquids. Supercritical fluid extration has been used to extract plant materials. Supercritical fluid extraction (Fig. Once the extraction is completed.. 2002).6) is analogous to extraction and distillation because both the solubility and volatility effects are utilized. 2004a) are among the novel technologies developed for the extraction of essential oils. 2003.

Fig.K.3 to 300 GHz. 6. flavors and natural aromas than conventional steam distillation. microwave heating is more effective than conventional heating methods.896 GHz in U. Owing to this combination of short wavelength and high frequency. Microwave energy is also characterized by short wavelengths. Polar molecules absorb microwave energy more easily causing 21 .45 GHz. Microwaves are electromagnetic radiations with a frequency from 0. while industrial microwaves operate occasionally at 0. Domestic microwave ovens operate at 2.915 GHz in the Europe & USA and at 0. there has been widespread interest in the application of microwave heating to extraction processes. Schematic diagram of a SFE system Recently.and Weller (2006) stated that supercritical fluid extraction can achieve higher yield and quality of essential oils.

food and plant materials.. The closed microwave-assisted extraction system is generally used for the extraction under high temperatures. Because water within the plant material absorbs microwave energy. Microwave-assisted extraction is one of the new methods which makes use of microwave heating for the extraction of neutraceuticals. efficiently and homogeneously. 2006). 2002). and focused microwave ovens at atmospheric pressure (Kaufmann and Christen. faster energy transfer. the focused microwave-assisted extraction system is operated at a maximum temperature determined by the boiling point of the solvent at atmospheric pressure. 2006). Much attention has been given to the application of microwave dielectric heating in analytical chemistry owing to its accompanying advantages such as more effective heating. If a dry biomaterial is re- 22 . Microwave heating is also more effective because heating proceeds from inside to outside. essential oils. simplified manipulation. unlike conventional heating systems where heat flows from outside to inside (Singh. faster response to process heating control. There are two types of commercially available microwaveassisted extraction systems: closed extraction vessels under controlled pressure and temperature. animal tissues.resonance and thus generating direct frictional heat. The technique offers a rapid delivery of energy to a total volume of solid plant matrix and/or solvent with subsequent heating of the solid matrix and/or solvent. elimination of process steps and higher purity of the final product (Ferhat et al. 1993). All the applications have shown that microwave-assisted solvent extraction is a good alternative to conventional techniques for such materials (Ferhat et al. dioxins and other organic compounds have been extracted efficiently from a variety of matrices such as soils. phenols. Up to now. pesticides.. cell disruption is promoted by internal superheating. faster start-up. sediments.. reduced analysis time. In contrast. equipment size and energy consumption. 2001). selective heating. aromas. improving the recovery of neutraceuticals (Kaufmann et al. which facilitates desorption of chemicals from the matrix.

Therefore.. there are reported studies which used microwave-assisted hydrodistillation for the extraction of essential oils from Anethum graveolens L. Coriandum sativum L. this method can be used to extract thermo-sensitive compounds such as essential oils (Wang and Weller. (Kosar et al. the matrix itself can interact with microwaves and hence facilitate the heating process. Brown (Stashenko et al. Thus. Heat is produced by microwave energy so the sample reaches its boiling point very rapidly. The microwave heating leads to the expansion and rupture of cell walls and is followed by the liberation of chemicals into the solvent (Spar Eskilsson & Bjorklund. 2005b). In this method. Microwaves interact selectively with the free water molecules present in the gland and vascular systems. 2005a). In the literature.E. there are some drawbacks of the method such as additional filtration or centrifugation necessary to remove the solid residue and poor efficiency of microwaves when the target compounds or the solvents are non-polar (Wang and Weller. with subsequent rupture of their walls.. 2000). leading to very short extraction or distillation times. In this case.. (2004b) developed microwave-assisted hydrodistillation for the extraction of essential oils in plant materials.. and the temperature increases rapidly near or above the boiling point of water. 2006).. this leads to localized heating. 2006). Stashenko et al. 2004a). The main benefits of microwave-assisted extraction technique are reduction of processing time and solvent used.) N. allowing the essential oil to flow towards free water (Benkaci-Ali et al.. It is also possible to achieve distillation with the water inside the material. such systems undergo a dramatic expansion. the surrounding solvent can have a low dielectric constant and thus remains cold during extraction. However. (Kosar et al. 2006). Lippia alba (Mill. the dry plant material and substantial amount of water placed in a Clevenger apparatus was heated inside a microwave oven (which has a hole on the top) for a short period of time to take out the essential oil. Lavandula angustifolia 23 . Laurus nobilis L.hydrated before extraction. Salvia triloba L.

2006). “microwave-assisted steam distillation” (MASD) (Chemat et al. (2004a. ternatum (Velen. which is connected to a Clevenger apparatus outside the microwave oven. platytaenium Boiss. H. and thyme). Lucchesi et al.. This process frees essential oil which is drawn by the water inside the plant material by azeotropic distillation. The essential oil is collected directly and dried without any solvent extraction step. Solvent free microwave extraction has been used to obtain essential oils from different raw materials. 2005). which uses microwave absorption solid medium such as carbonyl iron powders rather than water.. More recently. They called this method as “solvent-free microwave extraction”. (2004a) developed a new technique combining microwave heating and dry distillation at atmospheric pressure for isolation and concentration of the essential oils in fresh plant materials. H. and star anise) and cardamom seed. More techniques other than the stated ones above have been developed for the extraction of essential oils since the development of solvent-free microwave extraction. garden mint.. subsp. The internal heating of the water within the plant material distends it and makes the glands and oleiferous receptacles burst. The vapour then passes through a condenser outside the microwave cavity where it is condensed. sphondylium L.) Brummit (Ozek et al. 2006) in 24 . The method involves placing the fresh or rehydrated (by soaking in water for some time and then draining off the excess water) dry plant material in a microwave reactor. cumin. Heracleum crenatifolium Boiss.. These include “improved solvent-free microwave extraction” (ISFME) (Wang et al. 2004b. The distillate is collected continuously in the receiving flask. The inventors of the technique claimed that solvent-free microwave extraction is neither a modified microwave-assisted extraction nor a modified hydrodistillation which uses a large quantity of water. without adding any solvent or water.. 2006b)..(Iriti et al. three spices (ajowan. Lucchesi et al. 2007) analysed the essential oils of three aromatic herbs (basil.

Yield Essential oil yield can be defined as the ratio of amount of essential oil extracted to the amount of plant material used. Composition Composition of essential oils is of paramount importance in determining their quality.2. so they might decompose easily. the quality criteria change with respect to the final use of the product in the target industry (food. Moreover. To speak in general.).1. cosmetic. and “microwave-assisted simultaneous distillation-solvent extraction” (MWSDE) (Ferhat et al. In addition to their contribution 25 . Today. researchers and industries are working on obtaining high quality essential oils by improving the processing technologies. 2007). etc. All of them involve the combination of microwave heating with their corresponding extraction techniques.7.7. for this reason. thus possessing the aforementioned advantages of microwave extraction.7. Quality Parameters of Essential Oils 1. Due to the fact that whole essential oils with high yields are favored..which steam is produced by heating water with microwave radiation. 1. However. pharmaceutical. essential oils rich in oxygenated constituents are favored rather than rich in terpene constituents since oxygenated compounds are primarily responsible for the flavor and aroma of the essential oils. terpenes are very sensitive compounds to heat and light. 1. yield is among the most important quality parameters.

1.3. pycnometers are used for specific gravity measurements since they offer the most convenient results (Guenther.4. Among the physicochemical properties. Specific Gravity Specific gravity is another important criterion of the quality and purity of an essential oil. The index of refraction is related to the physical structure of the medium through which light is passing. 1948). the index of refraction is a characteristic of substances that can be employed in identifying unknowns.7. Values for essential oils vary between the limits of 0.696 and 1. Refractive index measurements should be made at constant temperature. 26 .188 at 15°C. the greater the amount of dispersion.to the aroma and flavor of the essential oil. which increase the brilliance of a material. For this reason. The higher the refractive index. some oxygenated compounds are known to possess high antioxidant and antibacterial activities. Refractive Index Refractive index can be defined as the ratio of the velocity of light in a vacuum to its velocity in a transparent specimen. It can be defined as the ratio of the weight of a given volume of essential oil at 15-25° C to the weight of an equal volume of distilled water at the same temperature.7. specific gravity has been reported most frequently in the literature. as well. Generally. 1.

Care should be taken for the temperature of the environment while making the measurements since solubility is affected by temperature (Guenther. in general. Optical rotation readings are usually taken at room temperature since the variation of optical rotation with temperature for most of the essential oils is very small (Guenther. Optical Rotation Most essential oils when placed in a beam of polarized light possess the property of rotating the plane of polarization to the right (dextrorotatory) or to the left (laevorotatory).90% . 1948).70% 80% .60% . oils rich in oxygenated constituents are more readily soluble in dilute alcohol than oils rich in terpenes.1. Solubility in Alcohol Since most essential oils are only slightly soluble in water and are miscible with absolute alcohol.5. It is known that. 1.7. Alcohols having strengths of 50% . 1948).6. The extent of the optical activity of an oil is determined by a polarimeter and is measured in degrees of rotation. 27 . Both the degree of rotation and its direction are important as criteria of purity.95% and occasionally 65% and 75% are used in determining the solubilities of essential oils.7. The determination of solubility of essential oils in dilute alcohol is a convenient way of evaluation of the quality of the oil. it is possible to determine the number of volumes of dilute alcohol required for the complete solubility of unit volume of oil.

. three endemic Turkish Heraclum species (Ozek et al.. dill seed. Laurus nobilis L. 2004b. 28 . 2004a.. Objectives of the Study It is known that conventional methods used for the extraction of essential oils from plant materials posses some disadvantages like losses of some volatile compounds. garden mint. 2005). cumin.. Brown (Stashenko et al. Although microwave-assisted hydrodistillation of rosemary has been studied. Oregano (Origanum vulgare L. essential oils of these plant materials are known to have high antimicrobial and antioxidant effects. cosmetic and pharmaceutical industries.1. variation of essential oil constituents during extraction and the effects of different microwave power levels have not been investigated.) are generally used in food. low extraction efficiency.. 2005a. there are no reports on the solvent-free microwave extraction of essential oils from oregano or laurel.8. However. coriander seed (Kosar et al. degradation of unsaturated or ester compounds through thermal or hydrolytic effects and presence of toxic solvent residue in the extract.) and rosemary (Rosmarinus officinalis L. laurel (Laurus nobilis L.).E. 2007) and MAHD for the extraction essential oils from Lippia alba (Mill. basil. 2005b) and black cumin seeds (Benkaci-Ali et al.. Additionally. In the literature. Microwave-assisted hydrodistillation (MAHD) and solvent-free microwave extraction (SFME) are two recently developed techniques. Therefore. usage of a safe and economical technique for the extraction of valuable essential oils from these plant materials is extremely important. These spices are widely grown and consumed in Turkey. star anise.. 2004b). there are reports indicating the use of SFME for the extraction of essential oils of ajowan. which are thought to overcome these disadvantages. Salvia triloba L. rosemary (Lo Presti et al.. 2005).) N. thyme and cardamom (Lucchesi et al. 2007).

laurel and rosemary. refractive index and solubility of essential oils in alcohol were also examined. it was aimed to compare these methods with the conventional hydrodistillation regarding to the yield and composition of the extracts.Moreover. there are quite few reports on the quantitative analysis of essential oil constituents of oregano. laurel and rosemary to investigate the effects of microwave power and extraction time on the yield and composition of the final product. 29 . The objectives of the present study are to examine the applicability of SFME and MAHD methods in the extraction of essential oils from oregano. In addition. Quality parameters such as specific gravity.

valencene (Fluka.. o-cimene. terpinolene. and Merck (Darmstadt. Germany). methyleugenol. Authentic standard materials were used for the qualitative and quantitative analysis of the essential oils.. thymol. 1. verbenone (Fluka. Steinheim. and Rosmarinus officinalis L. limonene caryophyllene (Fluka. β- cuminaldehyde.8-cineole. γ-terpinene. Bellefonte. Japan). Merck (Darmstadt. Laurus nobilis L. p-cymene. The other standard materials were βeudesmol. p-cymene (Fluka. bornyl acetate. Turkey). α-humulene. Manisa. Materials The dried Origanum vulgare L. isopulegol. βmyrcene. β-myrcene. thymol. β-pinene. Germany) and camphene (Supelco. copaene. 30 . Israel). PA. A. The mixture of α-pinene. fenchol. respectively. hexane.CHAPTER 2 MATERIALS AND METHODS 2. Germany). 4-terpineol. sodium sulfate anhydrous and ethanol were purchased from Fluka (Buchs.S. α-terpinene. camphor. (Fluka. (Fluka. USA). borneol. Switzerland). carvacrol and β-caryophyllene was purchased from Supelco (Bellefonte. Germany). eugenol. γ-terpinene. were obtained from Kutas (Kutas Tarim Urunleri Dis Tic. myrtenal. Nonane (internal standard). α-pinene. 4-terpineol. 3-carene. Switzerland). Riedel-de Haën (Seelze.. 1-octen-3-ol. San. linalool (Sigma-Aldrich. US). α-terpineol. USA) as a custom mix.1. Germany). PA. methyl jasmonate. Spain).

and Laurus nobilis L.2. Methods For the extraction of essential oils of Origanum vulgare L.2. 150 g of dried sample was soaked in 700 mL distilled water for 1 31 . four power levels. solvent-free microwave extraction (SFME) technique was performed. 50 g of dried Origanum vulgare L. 60% (373 W) and 40% (249 W) were studied for Origanum vulgare L. Therefore. 100% (622W). In the case of Laurus nobilis L.. After the Clevenger apparatus was placed into the oven. was soaked in 700 mL distilled water at room temperature for 1 h in order to hydrate the external layers of the plant material and the excess water was drained off. For SFME. 80% (498 W). However. Solvent-free Microwave Extraction (SFME) The microwave oven used for SFME was White-Westinghouse (Pittsburg.1. In SFME. 2. it was observed that SFME was unsuccessful in dry rosemary leaves since the plant material did not absorb enough water in the water soaking step prior to extraction due to its arid nature and burning was observed even for short extraction times. USA) operating at 2450 MHz with a maximum delivered power of 622 W (measured using IMPI-2L test (Buffler. The dimensions of the interior cavity are 29 х 37 х 40 cm..2. MAHD was performed rather than SFME for the extraction of the essential oil from Rosmarinus officinalis L. Microwave oven was modified by drilling a hole at the top. to study the effect of microwave power. Flat bottom flask having a capacity of 1000 mL was placed in the oven and connected to Clevenger apparatus through the hole. the hole around the neck of the flask was covered with polytetrafloraethylene (teflon) to prevent leakage of microwave. 1993)). and two power levels (100% and 40%) were studied for Laurus nobilis L.

dehydrated with anhydrous sodium sulfate.h at room temperature. The moistened plant material was placed in flatbottom flask connected to the Clevenger apparatus. Schematic diagram of the process is illustrated in Figure 7. 7. The process was continued until no more essential oil was obtained. capped under nitrogen and kept at 4°C until being analyzed. The essential oil was collected in amber colored vials. experiments were replicated twice. Schematic diagram of the SFME & MAHD processes 32 . During the process. For each condition. Cooler Essential Oil Aqueous Phase Microwave Oven Plant Material Fig. the vapor passes through the condenser outside the microwave cavity where it is condensed. Experiments were performed starting from the beginning each time to collect the essential oil samples during extraction since taking continuous data was not possible.

The process was continued until no more essential oil was obtained. For each condition. The essential oil was collected in amber colored vials. Solid:water ratios were 1:10 for Origanum vulgare L. A heater (Termal Laboratory Equipments. The procedure is the same as in SFME except the saoking step of plant material in water for 1 h. 2. dehydrated with anhydrous sodium sulfate. 33 .Turkey) with a maximum power of 200 W was used for this purpose.2.3. To investigate the effect of microwave power. experiments were conducted twice. capped under nitrogen and kept at 4°C until being analyzed.2. capped under nitrogen and kept at 4°C until being analyzed. Istanbul. and Laurus nobilis L. experiments were conducted twice. dehydrated with anhydrous sodium sulfate. Microwave-assisted Hydrodistillation (MAHD) The same experimental set-up used in SFME was utilized for the MAHD process (Fig. dried plant materials were hydrodistilled using a Clevenger apparatus. The essential oil was collected in amber colored vials. and 1:7 for Rosmarinus officinalis L. 350 mL of distilled water was added to 50 g of Rosmarinus officinalis L. For each condition. Experiments were performed starting from the beginning each time to collect the essential oil samples during extraction since taking continuous data was not possible.7). Instead.2. Hydrodistillation In conventional hydrodistillation. two power levels (100% and 40%) were studied for Rosmarinus officinalis L.2.

2.2.4. Analysis

2.2.4.1. Particle Size Distribution

Particle size distribution was obtained by performing a sieve analysis. A set of test sieves (Endecotts Ltd., England) was placed on an automatic shaker (Endecotts Octagon 200 Test Sieve Shaker, England). The sieve having the largest diameter of openings (8 mm) was placed at the top and the one having the smallest diameter of openings (425 micron) was placed at the bottom. The sizes of the openings of the other sieves were as follows: 4 mm, 2 mm, 1 mm, 850 micron. A pan was located under the last sieve for the particles passing the finest sieve. 50 g of Origanum vulgare L., 100 g of Laurus nobilis L. and 100 g of Rosmarinus officinalis L. leaves were seperately placed on the top sieve and the set was shaken for 30 min. Once the process was finished, the particles retained on each sieve were removed and weighed. Mass fractions of the retained particles were calculated. Particle size analysis was performed using the differential analysis method. Since the particle size ranges differ from sieve to sieve, a plot of particle size range versus W i/(Dp(i+1) – Dpi) was drawn, where W i is the mass fraction and (Dp(i+1) – Dpi) is the particle size range in sieve ‘i’ (McCabe et al., 1993; Sahin and Sumnu, 2006). Sauter mean diameter ( Ds ) of the plant materials was calculated using the following formula:

Ds =

1 ∑ (Wi D pi )

(2.1)

34

where D pi is the average particle diameter described as the following formula:

D pi =

D pi + D p (i +1) 2

(2.2)

2.2.4.2. IMPI-2L Test

The maximum power of the microwave oven was determined using the IMPI (International Microwave Power Institute) 2L test (Buffler, 1993). The oven was operated at 100% power for 5 min with a load of 2000 ± 5 g water placed in two 1-L Pyrex beakers. The oven cavity was wiped with a wet cloth. The subsequent procedure is as follows: Two beakers of 1-L capacity, each containing 1000 g of water, were placed in the center of the oven, side by side and touching each other in the width dimension of the cavity. Initial temperature of the water in the beakers was 20°C ± 2° C. The oven was operated for 2 min 2 s at 100% power level. Final temperatures were measured immediately after the oven was turned off. 3 replications were made and the oven cavity was wiped with a wet cloth each time. The power was calculated using the following formula:

P(W ) =

70 (∆T1 + ∆T2 ) 2

(2.3)

where ∆Τ1 and ∆Τ2 are the temperature rises of the water in the two beakers calculated by subtracting the initial water temperature from final temperature (Buffler, 1993).

35

2.2.4.3. GC/GC-MS Analysis

The essential oils obtained were analyzed by gas chromatography (Agilent Technologies 6890N Network GC System, USA) and gas chromatography coupled to mass spectrometry (Agilent Technologies 6890N Network GC System coupled to Agilent Technologies 5973 Network Mass Selective Detector, USA). In order to perform quantitative analysis with FID at the same time with the component characterization of MSD, a two hole ferrule was used in which two columns were placed. By this way, injection of the sample from one injection block was distributed equally into two columns. The capillary columns used for both of the analysis were HP-5MS (30m × 0.25mm × 0.25µm) with a 5% phenyl methyl siloxane stationary phase. The samples were injected to the columns with an Agilent Tecnologies 7683B Series Injector (Thailand). The components of the essential oils were identified by comparison of their retention times with those of available authentic standards and with library matching of their mass spectra (NIST98, Wiley7n, Flavor2). The data were analyzed by a software program, MSD ChemStation. GC-MS conditions for Origanum vulgare L. were as follows: carrier gas, He; flow rate, 1 mL/min; splitless; injection volume 1 µL; injection temperature 250°C; oven temperature program, from 50°C to 150°C with 3°C/min, holding at 150°C for 10 min, to 250°C with 10°C/min; MSD transfer line temperature, 250°C; MSD quadrupole temperature, 150°C; ionization temperature, 230°C; ionization mode, electronic impact at 70 eV. Solvent delay was for 4 min. The GC analysis was performed with the following conditions: flow rate, 0.6 mL/min; FID temperature, 250°C; makeup gas type, He with a make-up flow rate of 45 mL/ min. GC-MS conditions for Laurus nobilis L. were as follows: carrier gas, He; flow rate, 1.2 mL/min; splitless; injection volume 1 µL; injection

36

holding at 50°C for 2 min. USA) and internal standard (nonane) were prepared in 7 concentrations and injected into the columns with the GC method explained above. For the quantitation of the other constituents of oregano essential oil. MSD quadrupole temperature. β-myrcene. rising to 225°C with 3°C/min. make-up gas type. injection temperature 250°C. Supelco. GC-MS conditions for Rosmarinus officinalis L.5 min. 260°C. 0. ionization temperature. flow rate. make-up gas type. calibration solutions composed of the custom mix (α-pinene. They claimed that the response of FID to a large number of compounds within a component class is very similar and highly constant. (2000) was used. 230°C.668 ppm). p-cymene. He. 230°C. Solvent delay was for 4.8 mL/min. β-caryophyllene. MSD quadrupole temperature. α-terpinene. carvacrol.5 min. 250°C. 230°C.8 mL/min. oven temperature program.4 mL/min.temperature 250°C. oven temperature program. MSD transfer line temperature. 150°C. therefore. FID temperature. electronic impact at 70 eV. holding at 60°C for 5 min. A calibration curve (r2 ≥ 0.998) was obtained for each component in the solution. thymol. 275°C. ionization mode. 0. 0. ionization temperature. 1-octen-3-ol. Solvent delay was for 4. electronic impact at 70 eV. He with a makeup flow rate of 45 mL/ min. were as follows: carrier gas. γ-terpinene. 4-terpineol. The GC analysis was performed with the following conditions: flow rate. The GC analysis was performed with the following conditions: flow rate. MSD transfer line temperature. and rising to 210°C with 2°C/min. injection volume 1 µL. essential oil. the approach of Schoenmakers et al. which was used for the quantitation of the corresponding components in the oregano essential oil. He with a make-up flow rate of 45 mL/ min. For the quantitative analysis of Origanum vulgare L. The concentration of the internal standard in each solution was constant (229. FID temperature. ionization mode. splitless. 150°C. the response factor of a component within a class can be used for the quantitation of the other 37 .

respectively. alcohols and sesquiterpenes in the essential oil of Origanum vulgare L. ethers and sesquiterpenes in the essential oil of Laurus nobilis L.6). respectively.26). 4-terpineol (~1.4) where fi is the relative response factor of a target component.24). phenols. were quantified using the relative response factors of β-pinene (~1. alcohols. essential oil. cuminal. respectively. eugenol (~1.43). esters. bornyl acetate. aldehydes. Ai and As are the integrated peak area of a target component and internal standard. myrtenal (~1. Following this approach.26).855 ppm). camphor (~1.53) and βcaryophyllene (~1. which was used for the quantitation of the corresponding components in the laurel essential oil. 1-octen-3-ol (~1. 1. methyl eugenol. phenols. linalool.components in the same class. monoterpene hydrocarbons. A calibration curve (r2 ≥ 0. β-eudesmol (external standards) and nonane (internal standard) were prepared in 7 concentrations and injected into the columns with the corresponding GC method explained above. 3-octanone (ketone) was quantified by assuming its relative response factor as 1.26).8-cineole. bornyl acetate (~1. were quantified using the relative response factors of α-terpinene (~1. The concentration of the internal standard in each solution was constant (238. W i and Ws are the mass of a target component and internal standard.48).46) and β-caryophyllene (~1. camphor .998) was obtained for each component in the solution. ketones.23). γ-terpinene. αpinene. For the quantitation of 38 . 1. α-terpineol.8-cineole (~1. β-caryophyllene. βpinene. calibration solutions composed of eugenol. The relative response factor is described as follows: fi = AsWi AiWs (2. For the quantitative analysis of Laurus nobilis L. Following the same approach stated above. myrtenal. p-cymene. respectively. thymol (~1.34).18). monoterpene hydrocarbons. 4-terpineol.

fenchol (external standards) and nonane (internal standard) were prepared in 7 concentrations and injected into the columns with the corresponding GC method explained above.935). sesquiterpenes in the essential oil of Rosmarinus officinalis L. essential oil. camphor (~1. The samples were injected with their corresponding GC methods. ketones. the relative response factor of camphor was used since Zhu et al. 4-terpineol (~1.lactones the relative response factor of β-caryophyllene (sesquiterpene) was used. For the quantitative analysis of Rosmarinus officinalis L. bornyl acetate. For the quantitation of aldehydes. carvacrol. (2005) claimed that the relative response factors of aldehydes were close to those of ketones. β-pinene. bornyl acetate (~1. 4-terpineol. essential oil.943). camphor . p-cymene. 5 µL of oil was diluted with 1445 µL hexane and 50 µL of 1 % (v/v) nonane solution was added to the sample. which was used for the quantitation of the corresponding components in the rosemary essential oil. For the analysis of Laurus nobilis L.855 ppm). monoterpene ethers and hydrocarbons. methyl jasmonate. β-caryophyllene.068) and β-caryophyllene (~0. 1. were quantified using the relative response factors of β-pinene (~0. 39 .073).8-cineole (~1. carvacrol (~1. Finally. calibration solutions composed of γ-terpinene. 1.8-cineole.092).245). essential oil was diluted with 1545 µL hexane and 10 µL of 5 % (v/v) internal standard (nonane) solution was added to the sample in order to minimize the injection errors. Following the same approach stated above. essential oil. The concentration of the internal standard in each solution was constant (238. A calibration curve (r2 ≥ 0. respectively. phenols. for the the analysis of Rosmarinus officinalis L. 5 µL of Origanum vulgare L.059).998) was obtained for each component in the solution. methyl eugenol. verbenone. alcohols. 5 µL of oil was diluted with 1445 µL hexane and 50 µL of 1 % (v/v) nonane solution was added to the sample. esters.

2.4. and Rosmarinus officinalis L. Germany) at 22 ± 2° C.5. dilution process was continued this time with 0. The dilution procedure is continued until a clear mixture was obtained.4. Laurus nobilis L. The measurements were made at 22-23° C.4. were calculated by dividing the weight of 10 µL essential oil to that of 10 µL distilled water. Three replications were made.2. No turbidity or opalescence was observed during the process. The volume of alcohol (V) used to obtain a completely clear solution was recorded.4. The solution was throughly shaken each time 0. The results were expressed as: 40 .2.2. Weight measurements were made in triplicate using a highly sensitive balance (Denver Instrument.2. Measurements of Solubility in Alcohol Solubility of the essential oil in 85% (v/v) ethanol was determined by using the method of Institute of Turkish Standards (TS 780).1 mL of 85% ethanol and throughly shaken each time.6. Specific Gravity Measurements Specific gravities of the essential oils of Origanum vulgare L..5 mL ethanol was added. 2. 1 mL of oil was diluted with 0. RFM 330 refractometer (England). Measurement temperatures were 25 ± 2° C.5 mL of ethanol until 20 times of the volume of previously added alcohol was attained. Refractive Index Measurements Refractive index measurements were made in triplicate using the Bellingham Stanley Ltd. Once the clear solution was obtained.

2. Whenever significance was obtained. chemical classes of compounds and physical properties) were statistically evaluated by analysis of variance (ANOVA).1 volume of essential oil soluble in V volumes or more of 85% ethanol.5. 2. Normality assumption was checked by Anderson-Darling test. Once the normality or constant variance assumption was not satisfied for residuals. Box-Cox transformation was performed. 41 . Statistical Analysis The results (essential oil yields. The assumptions (normality and constant variance of the residuals) of ANOVA were checked. Tukey test (p≤0.05) was performed using MINITAB for Windows (Version 13).

The effects of microwave power and time on yield. the particle size distribution results of oregano. refractive index and solubility in alcohol were given and discussed. composition and the physical properties of the essential oils were also discussed.5 µm. 42 . Sauter mean diameter ( Ds ) of the leaves was calculated as 1727. Particle Size Distribution of Oregano Leaves Particle size distribution of oregano leaves is given in Figure 8.1. Additionally.CHAPTER 3 RESULTS AND DISCUSSION In this section.1. MAHD and hydrodistillation) of essential oils from these plant materials in terms of yield. composition and physical properties such as specific gravity.) 3. 3.1. the results obtained in the extraction (by SFME. Oregano (Origanum vulgare L. laurel and rosemary leaves were given.

The yield was 0. Particle size distribution of Origanum vulgare L.048 mL oil/g oregano in hydrodistillation method.054. The maximum yields obtained were found as 0. 60% (373 W) and 40% (249 W) power levels.1. 0. 80% (498 W).Fig.1.049 mL oil/g oregano. 0. for 100% (622 W). Effect of extraction on quality parameters of oregano essential oil 3.1. 8. respectively.052 and 0.053.05) except for the yields obtained in hydrodistillation and SFME working at 40% power level (Table 43 .2. The results were found to be significantly different (p≤0. The amount of water absorbed by dry oregano leaves was about 290 % of its initial weight when soaked in water for 1 h. leaves 3.2. Yield Effects of microwave power level and duration of the SFME process on the essential oil yield of oregano were investigated.

This is doubtlessly due to the rapid generation of heat inside the moistened oregano with the absorption of microwave energy and the subsequent formation of a higher pressure gradient inside the plant material when subjected to higher microwave power levels. 2007). the initial extraction rate increased with increase in microwave power. which causes a rise in the extraction rate (Ferhat et al. The essential oils must have been thrown out of the glands quicker due to this increased pressure gradient. Another observation was that the time to reach the maximum yield was significantly higher in hydrodistillation (3 h) than in SFME and decreased as the microwave power increased (35 min for 622 W. 44 . The decrease in processing time with an increase in power can be explained by the same reason.. The reason for the 80% reduction in the time of the extraction process in SFME method than in hydrodistillation is again the high pressure gradient formed inside the plant material. as compared to conventional hydrodistillation.A. it can be concluded that SFME method offered higher yield essential oil from Origanum vulgare L. Lucchesi et al. Therefore. As it is clearly seen from the graph. The decrease in the yield with lower microwave power and hydrodistillation process can be explained by the loss of some of the volatile compounds due to longer processing time..1-2). 2006. as microwave power increases the pressure gradient increases. Figure 9 shows the variation of essential oil yield with time in SFME and hydrodistillation processes. 50 min for 249 W).

It was also found that the essential oil was composed mainly of oxygenated compounds (80-85 45 . carvacrol (40-60 mg/mL oil).1. the composition of the essential oils obtained by both of the methods are almost the same. γ-terpinene (35-50 mg/mL oil). were determined as thymol (650-750 mg/mL oil) followed by p-cymene (60-85 mg/mL oil). Composition The total ion chromatogram of the origanum essential oil is given in Figure 10. SFME-40% power levelbc. β-myrcene (~15 mg/mL oil) and α-terpinene (10-15 mg/mL oil). □. during hydrodistillation and solvent-free microwave extraction (SFME) at different power levels (■. SFME-60% power levelac. hydrodistillationb) 3.Fig. 9. ♦.2. The composition of the essential oil of Origanum vulgare L. SFME-80% power levela. ●. As it is observed from the table. obtained by SFME and conventional hydrodistillation methods are given in Table 1. SFME-100% power levela. Variation of essential oil yield of Origanum vulgare L. ▲. The main components of the essential oil of Origanum vulgare L.2.

essential oil extracted by SFME at 100% power level (1: α-thujene.10. 17: carvacrol. 10: p-cymene. Fig. 13: terpinolene. 4: 1-octen-3-ol.6 % of it. 14: borneol. 9: α-terpinene. 15: 4-terpineol. 7: α-phellandrene. 11: β-phellandrene. 2004a). 3: camphene. 20: α-humulene. Total ion chromatogram (obtained by GC-MS analysis) of the Origanum vulgare L. 16: thymol. since oxygenated compounds are the ones responsible for the characteristic aroma of essential oils.. This situation explains the pungent odour of Origanum vulgare L. 2: α-pinene.%) while monoterpene hydrocarbons and sesquiterpenes constituted 1316 % and 1-1. 22: β-bisabolene) 46 . 6: β-myrcene. monoterpenes contribute to their fragrance to a very small extent (Lucchesi et al. 12: γterpinene. 18: β-caryophyllene. respectively.

901 144.328 RT 2: Retention time in min on HP-5MS column obtained by FID 98.234 98.858 59.493 1.617 23.369 684.941 1.897 979.715 272.502 815.737 4.655 16.489 8.837 54.249 64.914 29.968 3.068 11.690 4.353 758.751 11.248 19.272 23.426 5.541 36.835 8.662 157.277 16.175 3.903 15.970 2.728 47.334 9.946 54.523 26.405 2.692 42.499 36.973 35.270 764.228 23.585 99.416 4.120 944.480 832.996 1.419 24 (min) (mg/ml) 9.527 14.521 25.372 2.058 3.857 14.883 1.230 46.895 3.351 18 (min) (mg/ml) 8.767 40.405 84.149 13.122 147.542 27.192 3.999 2.706 (mg/ml) 8.115 5.310 6.184 8.771 2.498 34.500 8.729 15.783 8.729 13.539 14.012 2.264 2.058 14.215 2.093 2.327 132.628 4.790 11.812 810.592 3.940 581.039 45 (min) (mg/ml) 5.205 32.082 15.878 44.793 59.922 1.915 4.160 7.007 10.199 60.077 7.440 63.543 13.623 166.204 34.754 4.368 13.751 13.744 10.171 1.476 543.090 8.277 3.711 16 16.681 28.048 732.012 12.116 6.039 1.253 31.519 98.753 28.060 99.798 60 (min) (mg/ml) 4.029 68.834 702.225 10.511 11.038 3.332 10.193 5.791 4.373 3.395 24 (min) (mg/ml) 1.180 1.939 14.463 44.566 3.782 639.832 1.519 20.845 60 (min) (mg/ml) 4.934 5.017 44.581 1.645 5.203 70.565 25.450 2.92 29.916 1.992 6.095 2.676 83.212 78.076 208.271 11.105 65.299 1.510 90 (min) (mg/ml) 2.780 10.346 6.566 3.844 18.390 1.493 1.164 678.778 3.977 98.485 1.583 41.559 1.624 2.775 4.318 163.108 7.286 12.250 51.123 10.608 9.931 8.723 390.099 1.477 8.776 16.155 1.579 13.684 1.555 10.867 18.437 427.139 1.148 97.273 2.915 1.574 151.875 20.724 4.781 5.273 SFME-40% power level 8 (min) (mg/ml) 9.381 97.515 1.699 102.482 2.262 1.959 16.090 2.281 1.550 76.691 33.487 23.338 125.300 16.937 753.336 2.210 2.795 29.724 27.388 8.052 4.225 126.956 3.421 10.274 893.997 10.286 46.273 7. Compounds present in the essential oil of Origanum vulgare L.57 11.946 27.251 3.226 20 (min) (mg/ml) 2.010 67.275 12.477 1.242 SFME-100% power level 5 (min) (mg/ml) 6.945 10.713 180 (min) (mg/ml) 3.711 66.756 2.909 6.214 37.317 778.489 11.147 21.064 26.848 60 (min) (mg/ml) 2.499 98.212 2.797 6.093 1.291 12.189 49.369 304.361 96.608 1.800 3.599 .704 10.677 59.835 97.555 310.248 8.406 7.460 6.59 30.114 15.831 33. 99.569 98.201 34.221 14.212 17.949 8.395 48.116 20.167 4.225 5.652 552.488 2.153 526.059 13.544 146.671 715.116 77.336 1.041 1.342 1.602 3.260 1.825 15.747 13.328 8.944 135.145 450.466 19.590 2.129 1.104 1.795 10.416 879.409 9.249 1.242 19.313 3.751 2.308 11.572 12.519 11.200 17.910 8. obtained by different methods and their concentrations Hydrodistillation 46 (min) No 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 Compounds α-thujene α-pinene camphene 1-Octen-3-ol 3-Octanone β-myrcene α-phellandrene 3-Carene α-terpinene p-cymene β-phellandrene γ-terpinene Terpinolene borneol 4-terpineol thymol carvacrol β-caryophyllene aromadendrene α-humulene ledene β-bisabolene RT 1 (min) RT 2 (min) 7.680 1.015 3.067 6.481 2.712 1.631 2.368 2.924 19.206 1.377 29.872 7.533 6.013 2.234 10.263 9.038 16.293 1.853 47 % of total monoterpene hydrocarbons oxygenated compounds sesquiterpenes RT 1: Retention time in min on HP-5MS column obtained by MSD.944 SFME-80% power level 6 (min) (mg/ml) 8.921 6.549 24.310 21.363 1.643 1.759 350.318 13.661 6.100 820.047 1.451 SFME-60% power level 6 (min) (mg/ml) 10.302 37.666 676.897 53.658 5.426 7.172 860.721 651.846 12.634 1.788 1.624 132.385 30.489 77.407 6.173 6.089 14.98 32.617 12.997 13.661 10.669 1.714 3.494 18.481 6.003 1.897 4.794 127.375 14.400 2.559 37.160 4.917 97.877 11.248 15.553 2.161 98.288 2.398 1.383 28.166 97.858 4.005 9.178 1.627 466.911 1.586 6.844 1.027 24.Table 1.837 7.766 672.653 642.973 2.

Monoterpene hydrocarbons being the most volatile substances were preferably extracted at the beginning of the process whereas higher molecular weight compounds were extracted later. monoterpene hydrocarbons are known to be very unstable against heat and light. 48 . In addition. It was observed that monoterpene hydrocarbons decreased while sesquiterpenes increased with increase in time (Figure 11-15). They need time to reach their maximum levels in the essential oil. The decrease in monoterpene hydrocarbon concentration during the later stages of extraction may be due to the enhanced leaching of substances which were more difficult to be extracted (oxygenated compounds and sesquiterpenes). The increase in sesquiterpenes during the extraction period is probably due to their high molecular weights. so the decrese in their amount may also be explained by their probable degradation due to the thermal or hydrolytic effects.A general trend was obtained in the variation of the amounts of monoterpene hydrocarbons and sesquiterpenes with the time of extraction in both SFME and hydrodistillation methods. low volatilities and lower solubilities in water than the other constituents.

900 800 Concentration (mg/mL).. 700 600 500 400 300 200 100 0 46 90 Time (min) 180 Fig. sesquiterpenes) 49 . obtained by hydrodistillation ( .. oxygenated compounds..11. monoterpene hydrocarbons.. Variation of component classes with time in the essential oil of Origanum vulgare L. ... .

.. .. Variation of component classes with time in the essential oil of Origanum vulgare L. oxygenated compounds. 800 600 400 200 0 5 20 Time (min) 45 Fig. . sesquiterpenes) 50 .. monoterpene hydrocarbons. 12.. obtained by SFME at 100% power level ( .1200 1000 Concentration (mg/mL)..

. . sesquiterpenes) Concentration (mg/mL). oxygenated compounds. obtained by SFME at 80% power level ( . 13...1000 900 800 700 600 500 400 300 200 100 0 6 18 Time (min) 60 Fig.. 51 . Variation of component classes with time in the essential oil of Origanum vulgare L... monoterpene hydrocarbons. .

. sesquiterpenes) 52 ..900 800 Concentration (mg/mL).. oxygenated compounds... . Variation of component classes with time in the essential oil of Origanum vulgare L. 14.. 700 600 500 400 300 200 100 0 6 24 Time (min) 60 Fig. obtained by SFME at 60% power level ( . . monoterpene hydrocarbons.

15. obtained by SFME at 40% power level ( . oxygenated compounds.. 800 700 600 500 400 300 200 100 0 8 24 Time (min) 60 Fig. while the rest consisted of alcohols and ketones. their concentrations increased to some extent. their concentrations seemed to decrease slightly with time in SFME working at 100% power level as the concentrations of sesquiterpenes increased. 95% of the oxygenated compounds in the essential oil of Origanum vulgare L... monoterpene hydrocarbons. approximately. the extraction of oxygenated compounds were very fast in SFME at 100% power level because of pressure build-up inside the plant material.1000 900 Concentration (mg/mL). Generally. . However. Therefore. were made up of phenols (thymol and carvacrol). sesquiterpenes) It is hard to say that a perfect trend was obtained in the variation of the amounts of oxygenated compounds with time. . In general. Variation of component classes with time in the essential oil of Origanum vulgare L. It was found that.. The increase in the amount of oxygenated compounds with time is due to the increase in the 53 . an increase with time in the amount of phenols and a decrease in the amount of alcohols and ketones were observed (Table 1)...

. These two monoterpene hydrocarbons are known to be the precursors of thymol and carvacrol in the species of Origanum and Thymus (Burt. It was also found out that. oxygenated compounds and sesquiterpenes with the method of extraction. 2004). it can be concluded that SFME is a good alternative for the extraction of essential oil from Origanum vulgare L. but decreased the process time substantially. Therefore. This shows that SFME did not change the quality of the essential oil of Origanum vulgare L.05) (Table A.3-5). Therefore.amounts of phenols (thymol and carvacrol) since they are the main components. about 70% of the monoterpene hydrocarbons in the oregano essential oil were made up of p-cymene and γ-terpinene. 54 . Statistically. no significant difference in their corresponding concentrations was observed between the methods and power levels studied (p≤0. Figure 16 shows the variation of monoterpene hydrocarbons. as stated before.. the increase in the amounts of thymol and carvacrol with time might be due to the decrease in the amounts of pcymene and γ-terpinene resulting from a couple of reactions (aromatization of γ-terpinene to p-cymene followed by hydroxylation of pcymene (Nhu-Trang et al. That would also explain the decrease in the amount of monoterpene hydrocarbons with time since they consist mostly of pcymene and γ-terpinene. 2006)) activated under the thermal or hydrolytic effects.

monoterpene hydrocarbons decreased 55 . hydrodistillation180 min. with respect to different extraction methods ( . . Specific gravity Variation of specific gravity values of oreganum essential oil with time obtained by SFME at different power levels and conventional hydrodistillation is shown in Figure 17.1. This is probably caused by the fact that as extraction proceeded in both of the methods.1. as it can be seen from the figure.Fig. . . SFME-40% power level-60 min) (* means bars with different letters within each compound class are significantly different (p≤0.2. a slight increase with time was observed in both of the methods.05)) 3.16. SFME-100% power level-45 min.3. SFME-60% power level-60 min.3.2. Other parameters 3. Although most of the values practically maintained constant during the extraction process. Variation of compound classes in the essential oil of Origanum vulgare L. . SFME-80% power level60 min.1.

with time and sesquiterpenes together with oxygenated compounds increased to some extent. 0. Variation of specific gravity of the Origanum vulgare L. These results are in good agreement with those of Gamarra et al. Fig. Since oxygenated compounds and sesquiterpenes have higher molecular weights than the monoterpene hydrocarbons. 60% and 40% power levels were found as 0. ●. Statistically. specific gravity of the oil increased as they were gradually being extracted.921. 0. SFME at 100%. SFME-60% powera. 17.871. 56 . SFME-40% powera. ▲. hydrodistillationa ) Figure 18 shows the specific gravity values of oregano essential oil obtained by hydrodistillation and SFME at different power levels. SFME-100% powera.911 and 0. no significant difference was found between these values (p≤0. □.05) (Table A. The mean values for the specific gravities of the oregano oil extracted by hydrodistillation. respectively. essential oil with time obtained by different methods (■.6). (2006) who worked on lemon essential oil.897.

Lower values at the first stages of extraction is probably due to the higher amounts of monoterpene hydrocarbons and lower amounts of oxygenated compounds and sesquiterpenes compared to the last stages (Gamarra et al. Refractive index Variation of refractive index values of oregano essential oil with time obtained by SFME at different power levels and conventional hydrodistillation is shown in Figure 19. Like the specific gravity values. Specific gravity values of Origanum vulgare L.1.2. This is also in good agreement with the statement of Castilho et al. 18. They noted that refractive indices of natural fats and oils are related to their average degree of unsaturation in an almost linear way. a slight increase with time was observed in both of the methods although most of the values maintained practically constant. and tend to increase as the number of double bonds increase. essential oil extracted by different methods 3. It is known that as 57 .2.Fig. (2005).3. 2006).

19. 1. ▲.495. essential oil with time obtained by different methods (■. hydrodistillationa ) Figure 20 shows the refractive index values of oregano essential oil obtained by hydrodistillation and SFME at different power levels.512 and 1.05) (Table A. The mean values for the refractive index of the oregano oil extracted by hydrodistillation. refractive index values are expected to increase with the amount of sesquiterpenes.494. no significant difference was found between these values (p≤0. SFME at 100%.7). 60% and 40% power levels were found as 1. Variation of refractive index of the Origanum vulgare L. □. SFME-40% powera. respectively. the number of double bonds increases in terpenes.509. it can be concluded that they are more unsaturated than monoterpenes. Statistically. 58 . Fig.chain length increases. Since sesquiterpenes (15 carbon atoms) are bigger molecules than monoterpenes and their oxygenated forms (10 carbon atoms). ●. Therefore. SFME-60% powera. SFME-100% powera. 1.

respectively. 1.5 to 2 volumes. 60% and 40% power levels were found to be soluble in 1.3 to 1.20.7 volumes of ethanol. essential oil extracted by different methods 3.05) (Table A.2. Refractive index values of Origanum vulgare L. 1 volume of oregano essential oil extracted by hydrodistillation. The difference among the solubility of oregano essential oil obtained using hydrodistillation or SFME at different power levels were found to be insignificant (p≤0.2 to 1. 1. Solubility in ethanol Figure 21 shows the values of solubility of oreganum essential oil in ethanol extracted by different methods. SFME at 100%.6 volumes and 1.3.4 volumes.3 to 1.1. 59 .8).3.Fig.

4 µm.2.21. 60 . Particle Size Distribution of laurel leaves Particle size distribution of laurel leaves is given in Figure 22.2.) 3.Fig. essential oil extracted by different methods 3. Solubility in ethanol values of Origanum vulgare L.1. Sauter mean diameter ( Ds ) of the leaves was calculated as 2103. Laurel (Lauris nobilis L.

The amount of water absorbed by dry laurel leaves was about 135 % of its initial weight when soaked in water for 1 h. Effect of extraction on quality parameters of laurel essential oil 3. The variation of essential oil yield (mL oil/g laurel) with time during hydrodistillation and SFME processes is given in Figure 23.0235 and 0.2.1. leaves 3. it was decided to work with only two power levels (100% and 40%) for laurel. 22.2. respectively. Particle size distribution of Laurus nobilis L.2. In hydrodistillation method. The maximum yields obtained in SFME for 100% and 40% power levels were 0.022 mL oil/g laurel.2. Yield Finding out that there was no significant difference in the yields of essential oil of oregano extracted by different power levels.Fig.022 mL 61 . the maximum yield was found to be 0.

It was found that the time needed for the complete extraction of essential oil of laurel at 100% power was 85 min. SFME-40% powera.oil/g laurel. while it was 130 min in 40% 62 . The essential oils must have been thrown out of the glands quicker due to this high pressure gradient. ●. 23. no significant difference was obtained between these values (p≤0. Fig. hydrodistillationa) As in the case of oregano.05) (Table A. SFME-100% powera. Statistically.9). □. during hydrodistillation and solvent-free microwave extraction (SFME) at different power levels (■. as discussed before. Variation of essential oil yield of Laurus nobilis L. This is due to the higher pressure gradient formed inside the plant material when it was subjected to higher microwave power levels. the initial extraction rate increased with the increase in microwave power.

power.2. respectively. were determined as 1. Due to the lack of some of the authentic compound standards. The main components of the essential oil of Laurus nobilis L. sabinene (45-55 mg/mL oil). α-pinene (30-40 mg/mL oil). obtained by SFME and conventional hydrodistillation methods are given in Table 2. Composition Total ion chromatogram of the oil obtained by GC-MS analysis is given in Figure 24.8-cineole (630-730 mg/mL oil) followed by α-terpinyl acetate (90-115 mg/mL oil). This observation verifies the expectation for longer extraction times with lower microwave powers. Laurus nobilis L. It was also found that the essential oil was composed mainly of oxygenated compounds (≥75 %) while monoterpene hydrocarbons and sesquiterpenes constituted about ≥15 % and ≥0. The oxygenated compounds detected in laurel essential oil were found to be mainly 63 . essential oil owes its pungent odor to these high amount of oxygenated compounds as in the case of oregano.The composition of the essential oil of Laurus nobilis L. the process time was 195 min. the extraction time seems to be reduced by 55-60% in the case of SFME. It was observed that monoterpene hydrocarbons increased. In the case of hydrodistillation. A general trend was obtained in the variation of the amounts of monoterpene hydrocarbons and oxygenated compounds with the time of extraction in both SFME and hydrodistillation methods.2. No sesquiterpenes were detected in the essential oil extracted by hydrodistillation while they were detected only in the final stages of extraction in SFME (Figure 25-27). while oxygenated compounds decreased with time slightly. 3. which was discussed before. Doubtlessly. The composition of the essential oils obtained by both of the methods were found to be almost the same. only about 90% of the essential oil could be characterized. Therefore.2.15 % of it. 4-terpineol (35-40 mg/mL oil) and β-pinene (~30 mg/mL oil).

8-cineole (~ 70%). 4-terpineol. lactones (dehydrocostuslactone. the decrease with time in the amount of oxygenated components must be primarily due to the decrease in the amount of ethers. The rest of the oxygenated constituents of the oil were determined as alcohols (linalool. borneol. especially 1. eremanthin) and phenols (eugenol. 64 . cuminal). pinocarvone carvone). especially of 1. esters (bornyl acetate. α-terpineol. pinocarveol/trans-pinocarveol. trans/cis-methyl isoeugenol) (Table 2). ketones (camphor. αterpinyl acetate).8cineole. only ethers and ketones showed a reduction in their amounts as the process continued. Since ethers constituted the main components of the oxygenated compounds present in the laurel essential oil.composed of ethers (70-75%). aldehydes (myrtenal. Among these. They might have gone under some degradation reactions caused by high temperature and/or hydrolytic effects. β-eudesmol). cumic alcohol. although it was very small. sabina ketone.

35: erementhin(?) 65 . 21: myrtenal. 5: β-pinene. 19: 4terpineol.8-cineole. 24: bornyl acetate. 14: pinocarveol/trans-pinocarveol. 4: sabinene. 8: p-cymene.Fig. 28: eugenol. essential oil extracted by SFME at 100% power level (1: α-thujene. 24. 17: pinocarvone. Total ion chromatogram (obtained by GC-MS analysis) of the Laurus nobilis L. 11: γ-terpinene. 7: αterpinene. 34: dehydrocostuslactone(?). 30: methyl eugenol. 10: 1. 27: α-terpinyl acetate. 2: α-pinene. 3: camphene. 12: terpinolene. 13: linalool.

812 2.802 24.189 24.256 33.943 39.288 6.245 42.918 53.276 12.981 1.163 24.027 11.328 874.042 1.084 2.623 4.473 8.165 2.078 7.405 SFME-40% power level 25 (min) (mg/ml) 2.160 38.482 878.503 3.025 5.243 2.170 5.633 1.680 23.258 11.935 33.080 5.094 4.238 5.729 1.469 11.909 12. obtained by different methods and their concentrations Hydrodistillation 60 (min) No 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 Compounds alpha-thujene alpha-pinene camphene sabinene beta-pinene alpha-phellandrene alpha-terpinene p-cymene limonene 1.729 16.301 30.348 13.977 1.339 2.869 40.850 10.582 20.187 2.878 22.364 10 (min) (mg/ml) 2.494 3.932 6.296 4.103 2.023 20.401 8.060 12.464 2.431 3.273 9.000 3.748 4.916 3.604 6.413 731.719 RT 2 (min) 9.305 1.234 4.788 1.354 1.981 2.259 1.900 4.233 18.754 66.165 653.552 3.742 2.969 13.212 0.758 7.027 12.342 3.720 8.186 12.868 14.500 3.171 11.405 23.953 4.536 1.684 2.153 7 (min) (mg/ml) 1.158 35.984 24.795 14.885 2.184 9.349 8.012 195 (min) (mg/ml) 3.882 1.222 10.994 13.266 9.355 23.852 10.447 19.582 4.874 47.517 26.170 5.485 11.237 8.300 10.301 2.665 32.642 8.517 1.579 36.321 66 .181 2.697 9.537 7.483 5.148 10.152 10.197 26.004 9.301 843.953 19.051 15.547 43.495 1.476 19.560 4.639 115.716 1.004 9.915 2.401 8.328 4.284 4.899 12.168 23.487 9.889 2.399 9.627 16.448 38.496 20.180 3.946 18.403 3.228 8.809 36.928 28.263 3.828 0.050 10.289 38.691 11.460 1.Table 2.882 10.246 12.084 85 (min) (mg/ml) 3.310 2.072 1.120 18.8-cineole gamma-terpinene terpinolene linalool pinocarveol/trans-pinocarveol camphor sabina ketone pinocarvone borneol 4-terpineol alpha-terpineol myrtenal cuminal carvone bornyl acetate cumic alcohol pseudolimonene alpha-terpinyl acetate eugenol RT 1 (min) 6.321 17.050 1.024 107.106 1.535 1011.580 15.023 9.649 25.225 2.365 2.430 23.425 933.708 12.886 3.899 3.770 9.273 2.287 6.090 18.875 7.333 1.622 7.517 130 (min) (mg/ml) 2.676 36.571 2.179 10.636 35.521 11.397 3.769 1.830 2.133 13.628 2.946 3.364 12.674 6.966 31.986 9.504 2.840 19.176 5.325 15.300 8.728 68.790 3.460 1.201 3.441 1.957 36.917 41.930 1.877 28.603 2.088 4.894 45.320 6.461 40.840 75 (min) (mg/ml) 2.225 30.748 8.478 12.136 90.207 20.776 2.491 SFME-100% power level 15 (min) (mg/ml) 1.680 8.555 20.356 5.219 10.107 (mg/ml) 2.462 9.915 12.507 17.732 853.702 8.370 64.488 3.013 4. Compounds present in the essential oil of Laurus nobilis L.429 46.693 30.030 31.036 1.116 22.622 2.591 11.048 8.470 7.207 2.401 2.837 7.473 2.091 3.756 25.908 2.656 31.043 6.300 16.057 2.924 41.446 33.534 7.273 9.557 28.561 3.118 1.831 12.571 26.172 11.285 25.786 630.182 4.439 8.459 13.688 5.346 1.032 32.

930 14.771 1000.752 33.746 1067.836 0.153 3.597 5.382 47.087 91.078 39.080 92.000 6.082 5.214 39.223 39.000 10.933 3.364 5.550 302.696 64.380 1079.000 12.907 212.996 3. RT 2: Retention time in min on HP-5MS column obtained by FID 67 .685 413.030 RT 1: Retention time in min on HP-5MS column obtained by MSD.749 408.162 1132.283 1.793 44.Table 2.028 261.687 947.000 9.228 3.579 1025.012 3.350 206.258 93.101 0.387 0.972 91.583 0.905 1038.757 2.924 0.075 4.264 373.990 38.150 0.860 286.052 88.000 1.000 1.043 0.316 95.864 34.893 1.665 1086.550 0.838 5.690 53.212 289.689 859.429 64.631 4. (continued) 29 30 31 32 33 34 35 elemene(?) methyl eugenol beta-caryophyllene trans/cis-methyl isoeugenol beta-eudesmol dehydrocostuslactone(?) eremanthin(?) % of total monoterpene hydrocarbons oxygenated compounds sesquiterpenes 32.054 93.000 8.826 88.795 92.314 14.523 70.238 69.794 0.

. 25. Variation of component classes with time in the essential oil of Laurus nobilis L....1200 Concentration (mg/mL). .. sesquiterpenes) 68 . monoterpene hydrocarbons. obtained hydrodistillation ( ... . oxygenated compounds. 1000 800 600 400 200 0 60 75 Time (min) 195 Fig.

oxygenated compounds. ..1200 Concentration (mg/mL). 26... sesquiterpenes) 69 . Variation of component classes with time in the essential oil of Laurus nobilis L. monoterpene hydrocarbons. .. obtained SFME-100% power level ( . 1000 800 600 400 200 0 7 15 Time (min) 85 Fig....

. they were detected only in the final stages of SFME. They need time to reach their maximum levels in the essential oil. have lower volatilities. higher molecular weights and thus lower solubilities in water than the other constituents. sesquiterpenes) Sesquiterpenes. monoterpene hydrocarbons. No significant difference in the amount of monoterpene hydrocarbons and oxygenated compounds was observed between the methods and power levels studied (p≤0. Variation of component classes with time in the essential oil of Laurus nobilis L.. no sesquiterpenes were extracted in the hydrodistillation method while they were obtained only in the later stages of SFME (Table 2). As it is known.05) (Table A.1200 1000 Concentration (mg/mL). obtained SFME-40% power level ( . 27. oxygenated compounds. However. Therefore. This is probably because sesquiterpenes are high molecular weight compounds with low polarities.. .. Figure 28 and figure 29 show the variation of monoterpene hydrocarbons...10-11). . 800 600 400 200 0 10 25 130 Time (min) Fig. in 70 . oxygenated compounds and sesquiterpenes detected in the laurel essential oil with respect to different methods of extraction.

Therefore. they could be extracted in SFME probably because of the high pressure build-up inside the glands. thus their presence in foods is not so advantageous as compared to the presence of other constituents which possess antimicrobial and antioxidant effects. the extraction of sesquiterpenes could not be achieved in hydrodistillation most probably because of their high molecular weight nature since it is hard to extract high molecular weight compounds. the SFME process can be stopped in the later stages whenever a product free of sesquiterpenes is preferred. isolation of compounds is achieved according to their degree of polarity (which determines their degree of solubility in water) rather than their boiling points. 71 . Sesquiterpenes are generally known to have anti-inflammatory and antiallergenic effects. which induced the secretion of compounds with the rapid rupture of glands. However. Therefore. This would be advantageous also in shortening the process time.hydrodistillation.

SFME-100% power level. hydrodistillation. with respect to different extraction methods ( .Fig. . SFME-40% power level) 72 . . Variation of monoterpene hydrocarbons and oxygenated compounds in the essential oil of Laurus nobilis L. 28.

2. Other parameters 3. 3. hydrodistillation.12-13) It can be concluded that SFME is a good alternative for the extraction of essential oils from Laurus nobilis L. 29.3.2. Variation of sesquiterpenes in the essential oil of Laurus nobilis L. SFME-100% power level.2. . SFME-40% power level) (Table A. Although the values practically maintained constant 73 .2.Fig. . Specific gravity Variation of specific gravity values of laurel essential oil with time obtained by SFME at different power levels and conventional hydrodistillation is shown in Figure 30.1. since it provides essential oils of almost the same quality with conventional hydrodistillation while reducing the time of the process drastically. with respect to different extraction methods ( .3.

Variation of specific gravity of the Laurus nobilis L. 0. 30. ●. Specific gravity 74 . the specific gravity of the oil obtained by hydrodistillation was slightly lower. SFME40% powera.during the extraction process. SFME at 100% power level and at 40% power level were found as 0.856. no significant difference was determined between these values (p≤0. Fig. a slight increase with time in the first stages of extraction was observed in both of the methods. respectively.05) (Table A. hydrodistillationa ) Figure 31 shows the specific gravity values of laurel essential oil obtained by hydrodistillation and SFME at different power levels. □. Since no sesquiterpenes were extracted in hydrodistillation method.14). SFME-100% powera. This is probably due to the fact that compounds were gradually extracted as the process continued. The mean values for the specific gravities of the laurel oil extracted by hydrodistillation.867 and 0.861. Statistically. essential oil with time obtained by different methods (■.

essential oil extracted by different methods 3.2. As time increased. correspondingly increasing the refractive index 75 . Refractive index Variation of refractive index values of laurel essential oil with time obtained by SFME at different power levels and conventional hydrodistillation is shown in Figure 32. This is probably due to the fact that compounds were gradually extracted as the process continued. the extraction of unsaturated compounds increased. Fig.31.2.values of the laurel essential oil was similar to the values given in literature (Castilho et al.. Like the specific gravity values. 2005).2. a slight increase with time in the first stages of extraction was observed in both of the methods although most of the values maintained practically constant. Specific gravity values of Laurus nobilis L.3.

32. respectively.464.15). hydrodistillationa ) Figure 33 shows the refractive index values of laurel essential oil obtained by hydrodistillation and SFME at different power levels. SFME at 100% power level and at 40% power level were found as 1.values.05) (Table A. □. no significant difference was found between these values (p≤0. Statistically. SFME-100% powera. The mean values for the refractive index of the laurel oil extracted by hydrodistillation.465 and 1. ●.465. 76 . Variation of refractive index of the Laurus nobilis L. The refractive index of the essential oil obtained by hydrodistillation was lower since no sesquiterpenes were extracted in hydrodistillation method. 1. SFME40% powera. essential oil with time obtained by different methods (■. Fig.

Particle Size Distribution of Rosemary Leaves Particle size distribution of rosemary leaves is given in Figure 34. Rosemary (Rosmarinus officinalis L. Sauter mean diameter ( Ds ) of the leaves was calculated as 1392. essential oil extracted by different methods 3.3.Fig.33.3. 77 .) 3.1.4 µm. Refractive index values of Laurus nobilis L.

Particle size distribution of Rosmarinus officinalis L. it was decided to perform microwave-assisted hydrodistillation (MAHD) rather than SFME in rosemary.1. Yield It was found that solvent-free microwave extraction (SFME) method was unsuccessful in dry rosemary since the plant material did not absorb much water in the preliminary water soaking step due to its nature and burning was observed even for short extraction times.2. Effect of extraction on quality parameters of rosemary essential oil 3.3.2. leaves 3. Figure 35 shows the variation of oil yield with processing time during the conventional hydrodistillation and MAHD methods. Therefore. 34.3.Fig. 78 .

The maximum essential oil yields obtained in MAHD performed at 100% and 40% power levels were found as 0. hydrodistillationa) 79 . During the longer waiting periods at 40% power level.026 mL oil/g rosemary and 0. The reason for lower essential oil yield in MAHD at 40% power level might be explained as follows: the magnetron shuts itself in definite periods when operated at lower power levels than 100%.16-17). Fig. ●.021 mL oil/g rosemary. Therefore. respectively. □. Variation of essential oil yield of Rosmarinus officinalis L. 35. The maximum yield obtained in hydrodistillation was 0. during hydrodistillation and microwave-assisted hydrodistillation (MAHD) at different power levels (■. There was no significant difference between the yields obtained with MAHD at 100% power level and hydrodistillation.026 mL oil/g rosemary. some of the more volatile constituents of the oil might evaporate and be lost during these waiting periods. MAHD-40% powerb. while the yield obtained at 40% power level was found to be significantly different (p≤0. condenser which is made of glass may also be heated by conduction.05) (Table A. MAHD-100% powera.

Therefore. Approximately. β-pinene (~33 mg/mL oil).5-0. borneol (35-51 mg/mL oil). Composition Total ion chromatogram of the oil obtained by GC-MS analysis is given in Figure 36. respectively. It was also found that the essential oil was composed mainly of oxygenated compounds (77-82%) while monoterpene hydrocarbons and sesquiterpenes constituted 17-22 % and 0. α-pinene (65-85 mg/mL oil). essential oil owes its pungent odor to these high amount of oxygenated compounds as in the case of oregano and laurel.3. The reason was the formation of a higher pressure gradient with higher power level as discussed before. the extraction time was reduced by about 65% by using microwave. The time needed for the complete extraction of rosemary essential oil was found as 75 min for MAHD at 100% power level and 110 min for 40% power level. In hydrodistillation. were determined as 1.7% of it. The composition of the essential oil of Rosmarinus officinalis L. the duration of the process was found to be 210 min. It is clear that Rosmarinus officinalis L. 98% of the oil could be characterized.8-cineole (430-500 mg/mL oil) followed by camphor (150-210 mg/mL oil). 80 .Higher initial extraction rate at higher power level was observed as in the case of oregano and laurel.2. qualitatively. obtained by MAHD and conventional hydrodistillation methods are given in Table 3. whereas some quantitative differences were observed. The composition of the essential oils obtained by both of the methods were found. α-terpineol (35-55 mg/mL oil). 3.2. almost the same. The main components of the essential oil of Rosmarinus officinalis L. and camphene (20-25 mg/mL oil).

9: β-myrcene. 12: α-terpinene. Total ion chromatogram (obtained by GC-MS analysis) of the Rosmarinus officinalis L. 38: α-humulene. 7: β-pinene. 32: citronellol(?). 36: methyl eugenol. 4: camphene. 27: 4-terpineol. 13: pcymene. 36. 29: αterpineol. 15: 1.8-cineole. 35: carvacrol. 33: bornyl acetate. 2: α-thujene. 3: α-pinene. 8: 1-octen-3-ol. 39: methyl jasmonate) 81 . 23: isopulegol.Fig. 19: linalool. 17: γ-terpinene. essential oil extracted by MAHD at 100% power level (1: tricyclene. 22: camphor. 10: α-phellandrene. 37: β-caryophyllene. 31: verbenone. 18: terpinolene. 20: fenchol. 26: borneol.

886 210 (min) (mg/ml) 0.081 13.841 79.489 53.332 14.090 0.444 0.832 40.877 21.181 0.949 17.404 27.324 9.297 0.219 5.372 6.223 18.907 28.300 0.777 21.174 0.661 4.260 3.754 4.014 12.438 1.523 0.826 450.003 18 (min) (mg/ml) 1.342 0.497 0.844 4.556 4.532 1.696 2.981 78.324 100.892 19.437 10.275 16.104 75 (min) (mg/ml) 0.330 37.252 24.272 1.170 0.248 1.831 6.416 2.898 0.188 5.248 41.787 14.868 84.647 1.875 18.909 19.529 3.059 0.672 11.145 41.344 28.118 0.683 441.209 14.329 8.568 0.081 41.073 8.345 13.003 32.536 16.549 13.108 1.581 1.659 0.143 25.401 (mg/ml) 0.917 39.266 2.352 2.435 6.394 173.401 13.727 35.297 149.449 2.118 9.436 157.096 16.990 0.000 4.955 22.010 15.975 16.206 0.887 0.622 1.151 436.760 16.936 8.949 8.817 0.682 0.591 139.712 25.200 82 82 . Compounds present in the essential oil of Rosmarinus officinalis L.045 0.953 0.080 18.644 13.710 25.251 1.534 14.425 125 (min) (mg/ml) 0.488 7.442 0.922 66.385 3.799 0.718 85.277 7.329 8.611 MAHD-40% power level 35 (min) (mg/ml) 0.651 15.869 MAHD-100% power level 10 (min) (mg/ml) 1.869 9.399 17.224 33.281 32.764 2.138 538.279 10.727 0.119 11.592 1.520 1.583 1.161 1.875 6.427 21.549 1.213 17.837 0.030 0.319 5.059 7.234 18.386 1.826 15.810 15.696 153.299 21.847 1.095 8.466 1.648 12.427 1.423 11.583 1.502 17.752 12.579 1.813 0.743 16.160 11.000 3.629 15.226 150.539 RT 2 (min) 13.695 30.438 461.234 1.797 3.529 2.003 1.782 54.477 15.074 1.447 12.131 461.903 24.528 23.269 0.957 4.763 13.589 17.643 14.843 0.330 24.363 12.460 0.532 0.352 5.295 19.764 39.352 18.177 0.284 1.049 0.270 461.046 14.214 21.101 9.464 0.454 409.407 20.643 0.000 2.837 25.582 10.413 15.323 24.582 3.146 18.889 14.542 55.435 51.291 16.236 18.784 26.378 11.593 8.632 0.723 17.350 8.235 3.693 1.711 75 (min) (mg/ml) 1.910 8.057 0.294 1.425 25.484 7.724 38.939 18.Table 3.991 120.406 9.140 24.020 11.8-cineole cis-Ocimene gamma-terpinene terpinolene linalool fenchol campholaldehyde (?) camphor isopulegol pinocamphone/isopinocamphone pinocarvone/trans-pinocarvone borneol 4-terpineol p-cymene-8-ol (?) alpha-terpineol RT 1 (min) 7.263 0.874 9.728 13.509 0.153 0.649 0.521 1.385 1.820 0.654 498.211 0.970 10.332 11.502 14.208 31.903 0.176 10.605 11.810 3.356 25 (min) (mg/ml) 1.859 1.110 0.457 2.912 0.537 10.049 1.771 21.586 167.857 26.592 15. obtained by different methods and their concentrations Hydrodistillation 45 (min) No 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 Compounds tricyclene alpha-thujene alpha-pinene camphene verbenene (?) sabinene (?) beta-pinene 1-octen-3-ol beta-myrcene alpha-phellandrene 3-carene alpha-terpinene p-cymene limonene 1.176 25.843 0.570 4.481 1.613 190.420 23.928 0.846 11.316 25.756 1.893 36.697 1.726 208.742 4.750 1.625 20.786 0.637 12.

937 26.846105154 3.534 43.651675182 0.5 28.916 29.513157198 1.13280399 83 RT 1: Retention time in min on HP-5MS column obtained by MSD.197 4.617 1.534682 705.0215 0.89157564 194.169433005 1.94080399 2.0395 4.418 21.204 0.241 1.4509655 5.794 30.1115 1.6080648 775.458 6.991 1.261 0.068 34.964 29.5775 4.108 24.9385 0.8137516 4.9125319 750.496 25.665915351 1.824459259 3.005 2.2705 0.2115 1.413007986 0.5698686 2.494566 2.68546349 202.256734558 1.076087555 96.7685 2.7125 0.1772676 5.37 0.302 2.414703386 0.0785 0.8945 0.578129826 3.6185 1.626 2.341967 7.6799879 696.05299893 249.9865 1.192 0.609 1.78032623 133.850271214 2.9340372 4.3365 0.035 0.76336118 206.895 % of total monoterpene hydrocarbons oxygenated compounds sesquiterpenes 99.863 1.447175182 97.886 38.7955 0.975 1.38255075 121.812 24.543471239 3.339 35.6685 0.8755 0.195 36.199 2.886 3. RT 2: Retention time in min on HP-5MS column obtained by FID 83 .814 20.9846789 708.5032591 179.1685 4.133 2.515769374 99.61412913 176.643 1.5537594 871.7267823 647.315 26.273 0.0725 3.849657198 97.744771214 98.994083584 4.7865 2.68692941 1.837933005 99.401 2.112471649 1.552 29.312 23.923 0.153087555 0.9988948 743.480769374 1.352007986 97.006 0.195 3.0443739 4.49176509 159.389 2.978 4.3394915 4.253 0.436203386 97.6717458 736.939 0.2765 0.478 30. (continued) 30 31 32 33 34 35 36 37 38 39 myrtenol (?) verbenone citronellol (?) bornyl acetate thymol carvacrol methyl eugenol beta-caryophyllene alpha-humulene methyl jasmonate 19.Table 3.

A general trend was obtained in the variation of the amounts of monoterpene hydrocarbons and sesquiterpenes with the time of extraction in MAHD and hydrodistillation. It was observed that sesquiterpenes increased with increase in time (Table 3). This is probably due to their high molecular weights, low volatilities and lower solubilities in water than the other constituents. They need time to reach their maximum levels in the essential oil. In the case of oxygenated compounds, an increase in their concentrations with time was observed in MAHD, while the opposite trend was observed in conventional hydrodistillation (Figure 37-39). The oxygenated compounds detected in rosemary essential oil were found to be mainly composed of ethers (~ 60%), ketones (22-24 %), alcohols (15-17 %). Ethers were found to be constituted almost entirely of 1,8-cineole. The other oxygenated constituents of the oil were determined as aldehydes, esters, and phenols (Table 3). Since the oxygenated compounds were mainly composed of ethers, ketones and alcohols, the slight increase in their concentrations in MAHD can be attributed to the increase in the amount of the constituents stated above, especially 1,8-cineole. It is interesting to observe that 1,8-cineole concentration (thus, the oxygenated compound concentration) decreased with time in laurel essential oil while its concentration (and the oxygenated compound concentration) increased with time in rosemary oil which was extracted by MAHD method. In rosemary leaves, 1,8-cineole extraction was probably more difficult than its extraction from laurel leaves because of its plant matrix characteristics. Therefore, a couple of minutes may have not been enough for the entire extraction of 1,8-cineole from rosemary leaves as it was in the case of laurel leaves in which the going on process probably caused decomposition of the compound as noted before. In contrast to the situation in MAHD, as mentioned earlier, the oxygenated compounds in rosemary essential oil seemed to slightly decrease with time in

84

hydrodistillation. This is again caused by the decrease in the amounts of ethers, specifically 1,8-cineole, since other constituents of the oxygenated compounds increase with time. The opposite trends observed in both of the methods for oxygenated compounds might be due to the differences in extraction mechanisms of these methods and/or the noise factors involved in conventional hydrodistillation method, such as light. The system in MAHD does not contact with light since it is placed inside the microwave oven while the system in hydrodistillation does. Therefore, contact of the system with light might be among the reasons for the decomposition of 1,8-cineole with time in hydrodistillation. In the overall, monoterpene hydrocarbons decreased with time in MAHD, while they slightly increased with time in hydrodistillation. The concentration of monoterpene hydrocarbons seems to reduce with respect to time in the case of MAHD which may be due to the extraction of substances which were more difficult to be isolated (oxygenated compounds and sesquiterpenes). This dilution effect was not observed in the case of hydrodistillation since the concentration of oxygenated compounds decreased during extraction.

85

800 700 Concentration (mg/mL)........ 600 500 400 300 200 100 0 10 25 Time (min) 75

Fig. 37. Variation of component classes with time in the essential oil of Rosmarinus officinalis L. obtained by MAHD at 100% power level ( , monoterpene hydrocarbons; , oxygenated compounds; , sesquiterpenes)

86

. obtained by MAHD at 40% power level ( ... Variation of component classes with time in the essential oil of Rosmarinus officinalis L. monoterpene hydrocarbons.. 800 700 600 500 400 300 200 100 0 18 35 Time (min) 125 Fig.1000 900 Concentration (mg/mL).. sesquiterpenes) 87 .. . .. oxygenated compounds. 38.

. In all compound classes.... This is probably due to the small differences in the extraction mechanisms of the methods..20-21). In conventional hydrodistillation. . additionally. Therefore. extraction is achieved by dissolving the essential oil constituents in water and subsequent volatilization of the mixture. direct volatilization of the essential oil constituents from olifereous glands might occur due to the high pressure gradients formed inside the plant material. The essential oil extracted by MAHD was found to contain significantly higher amounts of oxygenated compounds than the one obtained by hydrodistillation (Table A. oxygenated compounds. .800 700 Concentration (mg/mL). Variation of component classes with time in the essential oil of Rosmarinus officinalis L. significant differences were observed between the methods and power levels studied (p≤0. monoterpene hydrocarbons. 600 500 400 300 200 100 0 45 75 Time (min) 210 Fig.. while in MAHD. oxygenated compounds and sesquiterpenes detected in the rosemary essential oil with respect to the method of extraction. sesquiterpenes) Figure 40 shows the variation of monoterpene hydrocarbons. obtained by hydrodistillation ( .. it may not be wrong to say that hydrodistillation was incapable of the entire extraction of 88 .05) (Table A. 39.18-23).

2006a) and it is known that as time increases the possibility of occurence of these reactions increases.05) (Table A. hydrodistillation.oxygenated compounds in rosemary leaves. . 40. . Variation of compound classes in the essential oil of Rosmarinus officinalis L. Oxygenated compounds in essential oils might be the products of either oxidation or hydrolysis (Wang et al. The reason for significantly higher amounts of oxygenated compounds obtained in MAHD at 40% power level than at 100% power level might be the longer extraction times involved in 40% power level. Fig.18-19). MAHD-40% power level) Significantly lower amounts of monoterpene hydrocarbons were detected in the rosemary essential oil obtained by MAHD than in the one obtained by hydrodistillation (p≤0. MAHD-100% power level. with respect to different extraction methods ( . This might be due to the diminution of thermal or hydrolytic effects because of significantly shorter 89 .

Thus. the possibility of degradation of monoterpene hydrocarbons increased which might be the reason of significantly lower amounts of monoterpene hydrocarbons at 40% power level than at 100% power level. so it is highly possible that they have gone under some degradation reactions. monoterpene hydrocarbons are unstable against thermal effects. isolation of compounds is achieved according to their degree of polarity (which determines their degree of solubility in water) rather than their boiling points.extraction times in MAHD than in conventional hydrodistillation. the entire extraction of sesquiterpenes might have not been achieved in hydrodistillation most propably bacause they could not totally dissolve in water. This is probably because sesquiterpenes are high molecular compounds with low polarities. This is probably due to the longer 90 . As mentioned before. as the time of extraction increased in MAHD. This might be explained as follows: degradation of monoterpene hydrocarbons may have become dominant over the decomposition of oxygenated compounds in MAHD since the extraction time was fairly shorter than hydrodistillation. Therefore. In contrast to this. in hydrodistillation. it is known that too long extraction times may also cause decomposition of the other constituents of essential oils such as oxygenated compounds. significantly higher amounts of sesquiterpenes were detected in the essential oil of rosemary extracted with MAHD than the one extracted with conventional hydrodistillation (Table A. significantly lower amounts of monoterpene hydrocarbons were detected in the oil extracted by MAHD at 40% power level than the one extracted at 100% power level although the time of extraction was longer in the former. Therefore. As stated before. the increase in the concentration of monoterpene hydrocarbons in hydrodistillation may be caused by the decomposition of some of the oxygenated compounds with the effect of long processing times. Lastly.22-23). However. the oil extracted with MAHD at 40% power level was found to contain significantly higher amounts of sesquiterpenes than the one extracted at 100% power level. Additionally.

a compound may be chemically linked to a substrate. a slight increase with time. so its extraction rate would depend on the decomposition rate of the chemical bond (Simard et al. Therefore. was observed. monoterpene hydrocarbons decreased with time and sesquiterpenes together with oxygenated compounds increased. 1988). 91 . Since oxygenated compounds and especially sesquiterpenes have higher molecular weights than the monoterpene hydrocarbons. It can be concluded that MAHD is a good alternative for the extraction of essential oils from Rosmarinus officinalis L..3.3.3. further extraction of sesquiterpenes might have been achieved with time. Specific gravity Variation of specific gravity values of rosemary essential oil with time obtained by MAHD at different power levels and conventional hydrodistillation is shown in Figure 41. specific gravity of the oil increased as they were gradually being extracted. since it provides essential oils of higher quality (higher oxygenated compounds. Other parameters 3.2.2. in some cases.3. In addition. lower monterpene hydrocarbons) compared to conventional hydrodistillation while reducing the time of the process drastically.1. This is probably caused by the fact that as MAHD proceeded. Although the values practically maintained constant during the extraction process. especially in MAHD. 3.extraction times involved in 40% power level.

Fig. hydrodistillationa ) Figure 42 shows the specific gravity values of rosemary essential oil obtained by hydrodistillation and MAHD at different power levels. The mean values for the specific gravities of the rosemary oil extracted by hydrodistillation. Statistically. 2005).05) (Table A. 41.24). 0. no significant difference was found between these values (p≤0.. □. respectively. The values were found to be in good agreement with the ones for the essential oil of Rosmarinus officinalis L. ●.879. and 0. Variation of specific gravity of the Rosmarinus officinalis L. MAHD-100% powera. MAHD at 100% power level and at 40% power level were found as 0. essential oil with time obtained by different methods (■.876.875. in literature (Atti-Santos et al. MAHD-40% powera. 92 .

essential oil extracted by different methods 3. Lower values at the first stages of extraction is probably due to the higher amounts of monoterpene hydrocarbons and lower amounts of oxygenated compounds and sesquiterpenes compared to the last stages (Gamarra et al.2.Fig. a slight increase with time was observed in both of the methods although the values can be assumed to be maintained practically constant.2. 2006). Refractive index Variation of refractive index values of rosemary essential oil with time obtained by MAHD at different power levels and conventional hydrodistillation is shown in Figure 43. Specific gravity values of Rosmarinus officinalis L. Like the specific gravity values. 42.3. 93 .3.

Variation of refractive index of the Rosmarinus officinalis L. in literature (Atti-Santos et al.05) (Table A. The mean values for the refractive index of the rosemary oil extracted by hydrodistillation. ●. 1. no significant difference was found between these values (p≤0.465. 2005).. MAHD-40% power. Statistically.Fig. MAHD at 100% power level and at 40% power level were found as 1. hydrodistillation ) Figure 44 shows the refractive index values of rosemary essential oil obtained by hydrodistillation and MAHD at different power levels. The values were found to be in good agreement with the ones for the essential oil of Rosmarinus officinalis L. □.25). MAHD-100% power.466. 94 . 43. respectively. essential oil with time obtained by different methods (■.465 and 1.

Refractive index values of Rosmarinus officinalis L.Fig. 44. essential oil extracted by different methods 95 .

The compositions of the essential oils in both of the methods were found to be almost the same which means SFME did not affect quality of the final product. Compositions of the laurel essential oils obtained by both of the methods were found to be the same except for sesquiterpenes. SFME offered significantly higher essential oil yields as compared to conventional hydrodistillation while reducing the conventional process time drastically. The main aroma compound in the essential oil of Origanum vulgare L. No sesquiterpenes were extracted in hydrodistillation. It can be concluded that SFME is a good alternative for the extraction of essential oils from Laurus nobilis L. were compared with the conventional hydrodistillation in terms of process time. This would be advantageous also in shortening the process time. Therefore. it can be concluded that SFME. was found to be 1. the SFME can be stopped in the later stages. In the case of Laurus nobilis L. can be a good alternative for the extraction of essential oils from Origanum vulgare L.CHAPTER 4 CONCLUSION SFME of dried Origanum vulgare L. since it provided essential oils of almost the same quality with 96 . and MAHD of dried Rosmarinus officinalis L. composition and other quality parameters of the essential oils. Since the presence of sesquiterpenes is not very advantageous as compared to the presence of other constituents which possess antimicrobial and antioxidant effects. was found to be thymol. For Origanum vulgare L. no significant differences were obtained in the essential oil yields obtained by SFME and hydrodistillation. and Laurus nobilis L.. which is a simple and time-saving method.. yield. Main constituent of the essential oil of Laurus nobilis L.8cineole.

Significantly higher amounts of oxygenated compounds and sesquiterpenes and significantly lower amounts of monoterpene hydrocarbons were detected in the essential oils extracted by MAHD than by hydrodistillation. Therefore. MAHD was performed instead for the extraction of essential oil from Rosmarinus officinalis L. No significant differences were obtained in the essential oil yields obtained by MAHD at higher power level and conventional hydrodistillation and the process time was found to be reduced substantially. Rosemary essential oil was mainly composed of 1. which means that MAHD offered higher quality essential oils than conventional hydrodistillation. since it provides essential oils of higher quality compared to conventional hydrodistillation and reduces the process time significantly.conventional hydrodistillation while reducing the time of the process substantially.8-cineole and camphor. MAHD is a good alternative for the extraction of essential oils from Rosmarinus officinalis L. 97 . Finding out that SFME was unsuccessful on dry rosemary leaves due to their arid nature.

Cabras. antimicrobial and antifungal activity investigation of the essential oil of Rosmarinus officinalis L. Aureli. A. (2005). P. Chemical composition.. Rossato. D. Physico chemical evaluation of Rosmarinus officinalis L. DM. Inhibitory effects of selected Turkish spices and oregano components on some foodborne fungi. essential oils. M. (1998). M. Rech. 10: 618-627. 48 (6): 1035-1039. Barra. M. Chemical composition. Dorman. G. M. and coriander essential oils. Rota.... oregano. Brazilian Archives Of Biology And Technology. Deans. Arlorio. sage... Serafini. (1989). Kıvanç. S. and antioxidative activity of laurel.. Journal of Essential. Dessi..REFERENCES Akgül. SG. A. Pauletti. Pansera.. P. GF. rosemary. 98 . Akgül. V. MT. Journal of Essential Oil Research. Kıvanç. Baratta... (1988). Moyne.. HJD. Antimicrobial activity of some plant essential oils against Listeria monocytogenes. plant genetic differences. Agostini. Costantini.. AC. A. 1:277280. P. Chemical Composition and antimicrobial effect of Turkish laurel leaf oil. 6: 263– 268... Angioni. MR.. A. Coroneo. E. (1992). antimicrobial. Barile.. LA.... F. JD. Zolea. LD. Ruberto. Coisson. Journal of Agricultural and Food Chemistry 52: 3530-3535. JC. Atti-Santos. Journal of Food Protection. S... 55 (5): 344–348. A. (2004). Oil Research. Cereti.. Biondi. International Journal of Food Microbiology.

. 99 . (1992).. Garbe. IHN. D. Traore. Bassett. Bendini. Bayoumi.. Bauer. Basaga. Garbe. Ouattara. G. SA. Bacteriostatic effect of some spices and their utilization in the manufacture of yogurt. Lercker. Surburg. H. K.. H.. D.) leaves Italian.. Antioxidative and free radical scavenging properties of rosemary extract. Toschi. Common Fragrance and Flavor Materials: Preparation. 30:105–108. K. 14 (1): 1724. Pannowitz. Properties and Uses. P.. 62: 209–212. S. ZI. (1990). (2002). H. Nebie. Tekkaya C. CAT. Bauer.. A. Medical Journal of Australia. Journal of Chromatography A 867 (1-2): 281-287. 21– 26. Properties and Uses. L. (2000) . IB.. Frnkova. (1990). Common Fragrance and Flavor Materials: Preparation. Food Science and Technology. (2001). Kabore. Ouattara. Cap... Chemie Mikrobiologie Technologie der Lebensmittel 14. Barnetson. Journal of Food Science . New York: Verlag. 153: 455–458...Bartak. RSC. P. Phytochemistry. F. TG. (2003). R. A comparitive study of tea-tree oil versus benzoylperoxide in the treatment of acne. (1997). Acikel. Weinheim : Wiley-VCH.Determination of phenols using simultaneous steam distillation-extraction.. Surburg.. Antioxidant activity of oregano (Origanum Vulgare L. Chemical composition and antibacterial activities of the essential oils of Lippia chevalieri and Lippia multiflora from Burkina Faso. AS. DL. Bassole.

Piattelli. Bernardo-Gil. G. Meklati. Antimicrobial activity and chemical composition of essential oils from Sicilian aromatic plants. Ruberto. seeds. Cardoso.104: 402–409. 64 (3-4): 227-231. Bicchi. P. A. D. M..Benkaci-Ali.. M. Biondi. Bertelli. Cianci. Chromatographia.. Mattarelli. Santos. BY. Benkaci-Ali.Baaliouamer. Phytochemical Analysis 11 (4): 236242. (2007). 100 . Chemat..55. Italy. European Journal of Lipid Science and Technology. Miglietta F. F. B.. 8:331. pp 54. D. Grenha. (1997)... Brandi.. Plessi... (2003).Baaliouamer.. F. Biavati. Chemical composition of seed essential oils from Algerian Nigella sativa extracted by microwave and hydrodistillation. Riolo Terme (RA).. M. Flavour Fragrance Journal. Presented at the Meeting on Atti Giornata Tecnico-Scientifica: “Piante aromatiche e da condimento”. Determination of phenolic diterpene antioxidants in rosemary (Rosmarinus officinalis L. BY. C.. P. Kinetic study of microwave extraction of essential oil of Nigella sativa L. Binello. Supercritical fluid extraction and characterization of oil from hazelnut. C. (2006). A. F. P. (2000). J. April 11.337. Marotti. 22:148-153.. (2002). A. (1993)... G. Effect of microwaves on volatile compounds in origanum. M. Flavour and Fragrance Journal .) with different methods of extraction and analysis.. Rubiolo. . J. Composizione chimica ed attivita` antimicrobica degli oli essenziali estratti da piante aromatiche da condimento. P.Geraci. G. Meklati.. Lebensmittel Wissenschaft und Technologie Food Science and Technology 36: 555-560.

Lognay. Food Chemistry 56 (4): 439-444. Kachouri...13:95-97. N.. R. Board. Food Preservatives. M. N. (1996). (2003). Flavour Fragrance Journal .. (Lauraceae). Zghoulli. Annals of Botany. G. GC. Gould. NJR and Gould GW (Ed. Blanch. 36:494. FJ. JM. P. F.). Bouzouita. 101 . Chaabouni. Braun... NA. Pino. Bosabalidis AM. Antimicrobial activity of essential oils from Tunisian aromatic plants. M. Herraiz.. Glandular Hair Formation in Origanum. MM. Chaabouni. (1991). 13 : 116-117.. Thonart. Journal of Essential Oil Research. Isolation and chemical characterization of laurel leaf oil.. B. P. (2001). (1995). In Gould.. S.. Hammerschmidt.. Reglero... Boutekedjiret C. Future prospects.18: 380–383. GW. Tsekos I. γTerpinyl acetate. Journal of Essential Oil Research.. Hamdi.. (2003). M.496. Meier. GP. J. Glasgow: Blackie Borges. 53 (4): 559–563. Nahrung-Food.. A new natural component from the essential leaf oil of Laurus nobilis L. Marlier.. (1992)... Belabbes. Journal of Essential Oil Research. 7:641– 644.. E. A. Antiviral activity of the essential oil of Melaleuca alternifolia (Maiden and Betche) Cheel (tea tree) against tobacco mosaic virus. Chemical composition of Laurus nobilis oil from Tunisia. 18 (6): 481-484 Bouzouita. Rapid extraction of wine aroma compounds using a new simultaneous distillation solvent extraction device.. MM. Nafti.. (267–284) . M. Sanchez. Kohlenberg. Bessiere. Bentahar.Bishop.(2001). Extraction of rosemary essential oil by steam distillation and hydrodistillation. (1984). RG. CD. Flavour and Fragrance Journal. F.

Internatıonal Journal of Food Microbology 94 (3): 223-253. G. Microwave cooking and processing: Engineering Fundamentals for the Food Scientist. M.. (2005). DRL. M. (2003).. (1994). 36 (3): 162–167. M. C. efficacy and provenance of tea tree (Melaleuca alternifolia) oil. A. (1999). Antibacterial activity of selected plant essential oils against Escherichia coli O157:H7.. 93 (2):223-226. (2005). CF. A new multivariate approach for the optimisation of the simultaneous distillation-extraction technique for free fatty acids using a face centred cube experimental design: application to Parmigiano-Reggiano cheese.. Mangia. Riley. Mori. Contact Dermatitis.. Inhibition of germination and growth of fruit and vegetable postharvest pathogenic fungi by essential oil components. New York: Avi Book. Castilho. Guizzardi. Careri. SA. TV. Capecka E. 102 . Burt. Portugal. Burt. (1993). Journal of Essential Oil Research. Antioxidant activity of fresh and dry herbs of some Lamiaceae species. JAOCS. Partidário. A.Buffler. Caccioni.... Mareczek A. Safety. (2001).. Letters in Applied Microbiology . MC. (2004). Rodrigues. 6:173-176. Reinders. Essential Oils: their antibacterial properties and potential applications in foods-a review. Characterization of laurel fruit oil from Madeira Island. RD. S. Musci.. Carson.. Analytica Chimica Acta. Leja M. 82 (12): 863-868. PC. Food Chemistry. 386 (1-2): 169-180. Costa. 45: 65– 67. A.

Busta. Charai. BB... 103 . F. F.... Franz C. and O. (1995). S. Iluiles essentielles de romarin (Rosmarinus officinalis): Comparison de la composition cbimique d’bulles du Maroc et d’amres provenances... G. 93:549-555 . Food Microbiology. RL. M. Chemat.. Journal of Essential Oil Research. Microwave accelerated steam distillation of essential oil from lavender: A rapid. Rivista Italia. Chemical composition and antimicrobial activities of two aromatic plants: Origanum majorana L. (1997). Resch H.Influence of day length and leaf insertion on the composition of marjoram essential oil. (1992).. A. Boucetta. Flavour and Fragrance Journal. Mendes. J. 16 (2): 136-148. (2003). Palavra.. R. JAP. Visinoni. FF. 14: 161–174. A.. Colnaghi.. A. compactum Benth. M. Favretto. Circella G. Analytica Chimica Acta. Lucchesi. AP. Michet. Chalchat. Chaintreau. 10: 371–374. (2006).. Smadja. L.. Flavour Fragrance Journal. Mosaddak. Supercritical carbon dioxide extraction of Foeniculum vulgare volatile oil. AMF. (1996). Flavour and Fragrance Journal. 18:316–319.. ME..Chaibi. Belasri. Garry. M. Pereira. clean and environmentally friendly approach. 8:657– 664. Chabart P. Faid.. Ababouch. LH. Simultaneous distillation-extraction: from birth to maturity-review.. (2001). 555 (1):157-160. Inhibition of germination and vegetative growth of Bacillus cereus T and Clostridium botulinum 62A spores by essential oils. Coelho.. EPPOS. Novak J.. K.

. International Federation of Essential Oils and Aroma Trades. Statti. K. M. piperitum (Ucria) Coutinho Seeds. Variability of essential oil content and composition of Origanum vulgare L. 104 . Arzedi. Tuberoso. LF. 21st International Conference on Essential Oils and Aroma’s. 86:471-478. Heat-Treatment of pollens ... CIG. Takacsova. MN. London : IFEAT. Conforti. (2000).Galletti. Pisano.. D. Leaves and Foeniculum vulgare subsp. P. Bouseta. A. F. 130– 135. (1998). D. Vanhavre.. F. 48:2576–2581.. S. (2000).. Bocchini. Crosthwaite. V. In vitro antimicrobial activity and chemical composition of Sardinian Thymus essential oils. B. Polissiou. Comparative Chemical Composition and Antioxidant Activities of Wild and Cultivated Laurus nobilis L. Mascia. Satta... Northern Italy). UK trade within the flavour and fragrance industry. M. . Kristianova.. DV. Dang. F. DJ. MG. 29. GC. (2001). S.Collin. Bodart.. Uzunov. Journal of Agricultural and Food Chemistry... D`Antuono. T. E. Nahrung Food. Journal of Agricultural and Food Chemistry 43 (2): 444-448. GC-MS analysis of essential oils from some Greek aromatic plants and their fungitoxicity on Penicillium digitatum. G. BN.impact on theır volatile flavor constituents. (1995).. Populations from a North Mediterranean Area (Liguria Region. Biological & Pharmaceutical Bulletin. Antioxidant activity of essential oils from various spices. 45 (1): 64–66. Daferera. Ziogas. (2006).. Annals of Botany. Menichini. Nguyen. E... 29 (10): 2056-2064 Cosentino. Letters in Applied Microbiology.. (1999). Palmas.

Rowshanzamir. Mirza. 77: 140-146.. Figueiredo. 15: 12–16. N. (2005). (2002) . Antioxidant activity of extracts obtained by different isolation procedures from some aromatic herbs grown in Lithuania. Composition and antimicrobial studies of the oils of Origanum calcaratum Juss. Journal of Essential Oil Research. Zhang.) . JG. Li. A. 50 (16): 4520-4524. DK.Effect of drying method on the volatiles in bay leaf (Laurus nobilis L. Journal of the Agriculture and Food Chemistry. Demetzos.. Flavour and Fragrance Journal . C. R. (2000). Deng. Xu. scabrum Boiss. 105 . (2006). MD.... Perdetzoglou. F.. 20 (6):555-558. Diaz-Maroto. (2001). Flavour And Fragrance Journal.. Barroso. Deans. M.. X.. Cabezudo. Golmohammad. Dorman. Perez-Coello.. Venskutonis. 13:460– 462. Duke. Eikani. Analytica Chimica Acta. Rapid determination of essential oil compounds in Artemisia Selengensis Turcz by gas chromatography-mass spectrometry with microwave distillation and simultaneous solid-phase microextraction. N. Van Beek.New York... 556: 289-294. SG. et Heldr..Dapkevicius. JA.Journal Of Agricultural and Food Chemistry. JPH. Linssen. C. Yao. MH. S. and O. TA. In vitro evaluation of antioxidant activity of essential oils and their components. X. AC. HJD. MS. NY:Rodale Press. The geen pharmacy: New discoveries in herbal remedies for common diseases and conditions from the world’s foremost authority on healing herbs. (1998). (1997). MC. from Greece. Recovery of water-soluble constituents of rose oil using simultaneous distillation-extraction.

. Fang. RS. Flamini. S. L... and chemical composition of their essential oils Journal of Agricultural and Food Chemistry. (2005). Ferhat. A. Miguel G. Chemat. Ho.. FM. Barroso... B.. Faleiro. Ceccarini. Elbaroty. BY. (2002). Antibacterial and Antioxidant Activities of Essential Oils Isolated from Thymbra capitata L.Elamrani.. J. 106 . (2000). L. F. (2006). MA. Rosen. SM. 1112: 121-126.. 12 (4): 487495. Pedro. Meklati. Main agronomic-productive characteristics of two ecotypes of Rosmarinus officinalis L. GSA. Zrira.. PL. MA.Chemat... Morelli. LG. KY. Venancio. 50 (12): 3512-3517. F. Journal of Chromatography A.. I. (2007). Journal of Food Protection. Gosslau. S. Cioni. CT. Gomes. (1989).. 65 (34): 217-222. F... Macchia. M. An improved microwave Clevenger apparatus for distillation of essential oils from orange peel. Chen. Meklati... ZY.. M. G. Journal of Essential Oil Research. BY. Rapid extraction of volatile compounds using a new simultaneous microwave distillation: Solvent extraction device.. Benjilali. Tigrine-Kordjani. Food Chemistry. JG.. 53 (21): 8162-8168... Daw. A. Berrada. Journal of the Agriculture and Food Chemistry. Smadja... Sang. Chromatographia. Figueiredo. RT. C. L. Costa. Teixeira. (2005) Isolation and identification of cytotoxic compounds from Bay leaf (Laurus nobilis) . A study of Moroccan rosemary oils. F.. (Cav. Hewedi. Antimicrobial activity of some Egyptian spice essential oils. A. Farag. 52 (9): 665– 667.. N. 93 (3): 497-501.. S.) and Origanum vulgare L. Ferhat. Chemat.

AN. Daferera.. Influence on the quality of essential lemon (Citrus aurantifolia) oil by distillation process. Supercritical fluid extraction of naringin from the peel of citrus paradise. Verzele. Sakanaka.. N. Journal of Essential Oil Research. Antimicrobial activity of essential oils and other plant extracts. Virens (Hoffm. Leeke. Nazareno. KA. TV. D. P. Journal of Applied Microbiology. Sokmen. and Sahin. CF. Guenther... LS. International Journal of Food Microbiology. et Link) letswaart: Content... AO.Gamarra. M. 51(14) : 3958–3965. Journal of the .. Phytochemical Analysis. Riley. H. Holley. 16 (2): 82-84. (2004).. (2003). New Method For Quantitative Essentıal Oil Analysis. F. Russo. Brazilian Journal Of Chemical Engineering. Delaquis.. 14:221–223. ssp. and antioxidant activities of the essential oil and methanol extracts of herbal parts and callus cultures of Satureja hortensis L.. FMC. EB.. M. Evaluation of antilisterial action of cilantro oil on vacuum packed ham. 86: 985– 990. (1981). New York: Van Nostrand Co. P. M. Agar. Godefroot. Essential Oil from Origanum vulgare L. HJ. Agricultural and. Cabral. F. Gulluce.. Giannuzzo. G. G. antifungal. (2006). Tambourgi.. Sokmen. (2003). Composition and Distribution Within the Bracts. Kartal. M.HT. Food Chemistry. RA. The Essential Oils. E. 1948. Journal Of Chromatography.. Boggetti. 23 (1): 147-151 Gaspar. A. Hammer. & Mishima.. 73: 83– 92. Ozkan. (1999). 203: 325- 107 .. MA.. Gill. Sandra.. FA. P. (2002). Carson. In vitro antibacterial. 335. Polissiou.. M.

Hao. Ankara: ITS. UV-B is required for normal development of oil glands in Ocimum basilicum L. Iriti. M. Glatman.. YY. (2003). (1998a). 15: 367–378. YY. Visinoni. Doyle. FA. MP.. 28:1-2. (1969). F. Smadja. TS 780: Solubility of essential oils in ethanol. Gelman. (1998b). Faoro. RE. Food Microbiology.. Annals of Botany. Efficacy of plant extracts in inhibiting Aeromonas hydrophila and Listeria monocytogenes in refrigerated cooked poultry. Effects of herbal essential oils used to extend the shelf life of freshwaterreared Asian sea bass fish (Lates calcarifer). MP.. 66 (3): 410–417. V. I. Netherlands: Leiden University Press. 108 . JM (1980). Ietswaart. Hethelyi. Histo-cytochemistry and scanning electron microscopy of lavender glandular trichomes following conventional and microwave-assisted hydrodistillation of essential oils: a comparative study.. Leiden. E. Drabkin. Flavour And Fragrance Journal. 90: 1–8. 61 (3): 307–312. (sweet basil). Bonner.. (2002). Brackett. Ioannidis. Colnaghi. A taxonomic revision of genus Origanum.. Chemat. 21 (4): 704-712. Doyle. (2006). F. Koczka. Johnson. S. (1989). Harpaz.Hao. Institute of Turkish Standards. Inhibition of Listeria monocytogenes and Aeromonas hydrophila by plant extracts in refrigerated cooked beef. L. L. CB.. Journal of Food Protection. Brackett.. Journal of Food Protection.. Phytochemical and antimicrobial analysis of essential oils. A. Tetenyi. P. D. J.. Herba Hungaria. RE.. G.

. (1999). Samejima.. Hirtum.The impact of both the season of collection and dryingon the volatile constituents of Origanum vulgare L. International Journal of Food Science and Technology 36 (6): 649-654. I. Johnson. 43: 404-409. Kawasaki. G. (1998). Naxakis. Mastelic. S. HE. K.. Substantial UV-Bmediated induction of essential oils in sweet basil (Ocimum basilicum L. 31 (1): 111-122 . P. (2001). Karousou. ssp. Journal of Agricultural and Food Chemistry 46. Danno. Kokkini. Afratis. N. S. (2005). Pardali. Specific desmutagens (antimutagens) in oregano against a dietary carcinogen. Pagona. (2004). Roditakis... H.. G. K. Effects of UV-B radiation on Mentha spicata essential oils. Pearson. Stratigakis. Milos.... A. Phytochemistry. J. M. Populational and Ontogenic Variation in the Volatile Oil Content and Composition of Individuals of Origanum vulgare subsp. Skoula. N. H. hirtum grown wild in Croatia. 49: 2273–2277. Assessed by GC Headspace Analysis and by SPME Sampling of Individual Oil Glands. Feggou. (1998). S. Kirby.Journal of Chemical Ecology.. Ashida.Jerkovic. Mavragani-Tsipidou. G. Journal of the Agriculture and Food Chemistry. J. Manetas. Grammatikopoulos. E. Scouras. Seasonal. Kazantzis. are galangin and quercetin. G.. M. Kokkini.. 22(3): 228-232. CB. CB. Johnson. Insecticidal and genotoxic activities of oregano essential oils. Composition and insect attracting activity of the essential oil of Rosmarinus officinalis .. 109 . Phytochemistry. Karpouhtsis. Katerinopoulos... (1995). E. I. 51: 507–510. Y. Kanazawa.). Lanaras T.. A. ZG. Trp-P-2. R. 1111– 1115.. Phytcochemical Analysis.

. Volatile Constituents and key odorants in leaves. P. Experientia. (2001). Lanaras T. 110 . Pharmaceutical Biology. MR.Kaufmann. H... Insecticidal effects of essential oils: A study of the effects of essential oils extracted from eleven Greek aromatic plants on Drosophila auraria.. Autumn essential oils of Greek oregano. Vassilopoulou. 327–331. (1992). Konstantopoulou. A. (2004). and Fruits of Laurus nobilis L. Goger. 13:105–113. Flowers. (1995).. ZG. P. 2839– 2845. M.... Phytochemical Analysis. JL. (2002). Journal of Agricultural and Food Chemistry.. CI. H.. (2005a).. T. Journal of Agricultural and Food Chemistry.. Kim. Comparison of microwave-assisted hydrodistillation and hydrodistillation methods for the analysis of volatile secondary metabolites . J. Antibacterial activity of some essential oil components against five foodborne pathogens. P. 52 (6): 16011606. Wei.. B. Kollmannsberger.. (1997). KHC. Marshall. 44: 883–886.. Veuthey. Ozek. B. Scouras.. Parameters affecting microwave-assisted extraction of withanolides.. Recent extraction techniques for natural products: Microwave-assisted extraction and pressurized solvent extraction. Kokkini S. Kosar. Kilic. M. 48 (6): 616– 619... Phytochemical Analysis. Baser. Dardioti A. Karousou R. 43. Kaufmann.. Mavragani-Tsipidou.. S. Phytochemistry. Nitz. Krigas N. 12. F. Kurkcuoglu. 43 (6): 491-495. Christen. Buds. Hafizoglu. Christen. L. I.

5 (4) : 29-33. Nychas.. Q. International Journal of Food Microbiology. KHC. Coote. M.... Progress in essential oils: Laurel leaf oil. (1983). 6 (2) : 60-61. Progress in essential oils: Laurel leaf oil. Re. GJE. Urrutia. Koutsoumanis. Ringuelet. Journal of Applied Microbiology.M.Kosar. Z. MI. T. Lawrence. Perfume and Flavour. S. GJE. MS.. K. by using microwave-assisted hydrodistillation. Kurkcuoglu. Skandamis. (2001).. (1999). Henning.. thymol and carvacrol. Ozek.Wai. 13:122-124. A simple method to obtain essential oils from Salvia Triloba L. 2 (7) : 46-48. (2001). Perfume and Flavour. CP. EL. A predictive model for the non-thermal inactivation of Salmonella enteritidis in a food model system supplemented with a natural antimicrobial. Tunalier. Progress in essential oils: Laurel leaf oil... M. Lambropoulou. BM. Lawrence. RJW.. Zeitschrift Fur Naturforschung C-A Journal Of Biosciences . Talanta. Baser. Lawrence. Perfume and Flavour. BM. P. Cerimele.. Lambert. In vitro fungistatic effect of essential oils against Ascosphaera apis. Perfume and Flavour. (2005b). JA. A study of the minimum inhibitory concentration and mode of action of oregano essential oil. and Laurus nobilis L. 8 (1) : 62-65. Lawrence. Carranza. MR... 49: 63– 74. BM. (1981). 111 . (2001).53: 771–782. BM. Lang. PN. (1980). C.60 (5-6): 501-504. Journal of Essential Oil Research.. (1978). K.. Supercritical fluid extraction in herbal and natural product studies—A practical review. 453–462. 91. Larran. Progress in essential oils: Laurel leaf ( or Mediterranean Bay) oil. Nychas..

Lawrence.Ragusa. Perfume and Flavour.. 78: 217– 226 Lis-Balchin. Trozzi. S. (1999). N. Hart. BM. Perfume and Flavour. 20(1) : 47. L. Progress in essential oils: Rosemary oil.. F.Lawrence. RJ.. P. Choquette. Determination of antioxidant potential of volatile extracts isolated from various herbs and spices Journal of the . Deans. Lawrence... A comparison between different techniques for the isolation of rosemary essential oil. Perfume and Flavour. S. Mondello. 28 (3): 273-280. Visinoni. M. Antimicrobial effect of natural preservatives in a cooked and acidified chicken meat model. Dugo. Garie´py. Rodrigue. Journal of Separation Science. Lawrence.. A. 50 (15): 4947–4952.. J. (2002). Saucier. Ochoka.. 11: 393– 397.. Delaquis. C. Differences in bioactivity between the enantiomers of α-pinene. 12(4) : 71.. Perfumer & Flavorist. Lemay. PJ. BM.M.. Progress in essential oils: Rosemary oil. M. L.. Food Chemistry... Dugo. Shibamoto. A. Journal of Essential Oil Research. Progress in essential oils: Laurel leaf oil.. (2005). Fazio. G. 22(5) : 71. Asztemborska. MJ. (1984).. 112 . Lee. (1995). T. 9:41– 51. KG. Lo Presti. BM. (1987)... (2002). International Journal of Food Microbiology. Agricultural and. The botanical and chemical aspects of oregano. (1997). BM. SG.

Impedance measurements to study the antimicrobial activity of essential oils from Lamiacea and Compositae. (2001). Lucchesi. Journal of Applied Microbiology 94: 761– 766.Antimicrobial activity of the essential oils of Thymus vulgaris L. Solvent free microwave extraction of Elletaria cardamomum L. S. (1999). 67: 187– 195. M. G. J. 113 .Lucchesi.. P.. Journal of Food Engineering. 19 (2): 134-138 Lucchesi.. Bersani. Scannerini. 79 (3): 1079-1086. 12: 523–529... M. J.). Pratella. Bersani. ME. S.. (2003). 62(9):1017– 1023. Journal of Essential Oil Research. M. C. (2004b).. Marino. (2000). 1043 (2): 323-327. Smadja. Bradshaw. Mari.. Marino. G. Comi. Solvent-free microwave extraction of essential oil from aromatic herbs: comparison with conventional hydro-distillation. An original solvent free microwave extraction of essential oils from spices . Journal of Chromatography A. Smadja. J. (2004a).Smadja.: A multivariate study of a new technique for the extraction of essential oil.. Flavour And Fragrance Journal.. Chemat F. (2007). Non-conventional methods for the control of post-harvest pear diseases. measured using a bioimpedometric method. Bertolini. M. C.. GC.. Chemat. International Journal of Food Microbiology. Maffei.. ME. ME. UV-B effect on photomorphogenesis and essential oil composition in peppermint (Mentha piperita L. Comi. Journal of Food Protection. F.

47: 4297– 4300. McCabe.Beauregard. P. JC. JA. (1994). H.Toguchia. Douglas. Unit Operations of Chemical Engineering. Deuterium/hydrogen ratio analysis of thymol. GK. Nat. H. DA. Antibacterial activity of turmeric oil: a byproduct from curcumin. Life Science.. KK.. Seasonal variation in essential oil yield and composition from naturalized Thymus vulgaris L. Nozaki. 31:27–31 114 . 1132 (1-2): 219-227. Antioxidant properties of essential oils: Autoxidation of essential oils from laurel and fennel and of their mixtures with essential oil from coriander.T. Jagan RM.. Applied Biochemistry and Microbiology 41 (6): 610-618.. carvacrol. H.. Nhu-Trang.. Grenier-Loustalot... JL. in New Zealand. Inhibitory effects of sesquiterpenes from Bay leaf on nitric oxide production in lipopolysaccharide-activated macrophages: structure requirement and role of heat shock protein induction. gamma-terpinene and p-cymene in thyme. Polshkov. savory and oregano essential oils by gas chromatography-pyrolysis-isotope ratio mass spectrometry. Misharina.. Smith. Casabianca. Negi. WL. MH. Yoshikawa.... 66:2151-2157. Journal of Chromatography A. Singapore: McGraw-Hill. TT.Matsuda. NB. Food Ind. Ueda. Kagerura. (1993). Flavour and Fragrance Journal. TA. Perry. Journal of Agricultural and Food Chemistry . (2005).. McGimpsey. L. Jayaprakasha. MF. M. Morikawa.. (1999). AN. PS. K (1989). I.. (Japan). Antioxidant activity of rosemary. T. 9: 347– 352. (2000). Harriot. (2006).. Van Klink.. Sakariah.

I. Panizzi L.. F. Baser. SF. Singh. Comparison of the essential oils of three endemic Turkish Heracleum species obtained by different isolation techniques... (2002). Mehrotra. Flamini G. I. Flamini. S. G. Morais.. Morelli. Biochemical Systematics and Ecology. S. M. Veterinary Parasitology 109 (1– 2): 59–63. SM. Pl. Anthelmintic activity of essential oil of Ocimum gratissimum Linn.17: 605-610. (2000). Mancianti.... (2005). G. CML. Ozek. WA. KHC.. Pandey. Lacroix. Bevilaqua..). International Journal of Food Microbiology. HN. 60 (2): 184-187. Morelli.... JHS... Pessoa. Cioni.. Pl. and eugenol against Haemonchus contortus. Peer.Ouattara... Kumar. Kalra.. Sabato. Combined effect of antimicrobial coating and gamma irradiation on shelf life extension of precooked shrimp (Penaeus spp.Composition and Antimicrobial Properties of Essentıal Oils Of 4 Mediterranean Lamiaceae Journal of Ethnopharmacology. Luciano. Cioni.. (1993) . JH. (1994). S. (1998). LM. Langenheim. 39 (3): 167-170. Essential oil compounds as potent source of nematicidal compounds. Influence of phytochrome on leaf monoterpene variation in Satureja douglasii. A. G. 115 . Journal of Phytopathology 148 (7–8): 501–502. Tandon. Ozek.. (2001).. T. Journal of Essential Oil Research. R. In-vitro antifungal activity of essential oils against some isolates of Microsporum-canis and Microsporum-gypseum Planta Medica.. N. 26: 25–34 Perrucci. 68:1– 9. Macchioni. B.

Putievsky E. Prudent.. Nahrung-Food. selection and conservation of oregano species in Israel...... Perineau.. GM. (2002). P. Combined action of nisin and carvacrol on Bacillus cereus and Listeria monocytogenes..)-. Putievsky.Journal of Pharmaceutical and Biomedical Analysis. U. J. 15–19. . Cultivation. Flavour and Fragrance Journal. (1997). Senorans. Journal of the Science of Food and Agriculture 43 (3): 225–228. Padulosi (ed. G. 29:166– 170. (1993). Bessiere. E. Cerri. P. Dudai. Pol. Chessa. MR. Borges. oils from Sardinia and Corsica.. (1988). 41 (5): 1606-1613 . 116 . M. N.. (103– 109).. FJ.. Bradesi. Casanova. Baccou.. JM.. Ramirez. J.).Pino. R. In S.. (2006). Isolation of functional ingredients from rosemary by preparative-supercritical fluid chromatography (Prep-SFC) . Journal of Essential Oil Research.evaluation of its bacterioatatic and fungistatic properties.. D. Santoyo. Rongal. Ibanez. The chemical composition of laurel leaf oil from various origins. U. EJ. Usai. F. Ravid. G. Reglero.. E. S. IE... Boatto. Dudai. (1995). (1999). 17.. 37 (6): 592-595.Rome: IPGRI. Phenological and seasonal influences on essential oil of a cultivated clone of Origanum vulgare L. Juliano. M. N. C. Letters in Applied Microbiology. G. Pintore. F. Tomi.. Proceedings of the IPGRI International Workshop on Oregano. Michel.. Ravid. 7: 165– 173. Garcia-Risco. Chemical composition and antimicrobial activity of Rosmarinus officinalis L.. JC. Smid. Analysis of the essential oil of wild oregano from Martinique (Coleus aromaticus Benth. P. E.

. Bocchini P. Hernandez. Gonzalez. Food Control. LM. Antioxidant activity of selected essential oil components in two lipid model systems. (1998). Agar. Sokmen. 15: 549-557. Essential oil chemical composition of wild populations of Italian oregano spice (Origanum vulgare ssp. H. Letters in Applied Microbiology. Food Chemistry. S. Sumnu.Ramos. hirtum (Link) Ietswaart): A preliminary evaluation of their use in chemotaxonomy by cluster analysis. Carvacrol and cinnamic acid inhibit microbial growth in fresh-cut melon and kiwifruit at 4 oC and 8 oC. D. Roller. (1998). MJ.. Journal Of Agricultural And Food Chemistry.. Allaf. 46 (9): 3741-3746. MW. Baratta..... Inflorescences.. (2000). 1. Louka. Boutekedjiret. M.. (2002). K. 69 (1): 167– 174. Selective extraction of polychlorinated biphenyls from dairy products using steam distillation solvent extraction at normal pressure.. Study of a new extraction process: controlled instantaneous decompression. vulgare in the Eastern Anatolia region of Turkey. M. (1998). Galletti GC. SA. Ruberto. .. Analytica Chimica Acta 376 (3): 313-323. Gulluce.. C. Tabera. P. Sahin S . G. M... Carnacini A. L.. N. Rezzoug.. Sokmen. Seedhar. F. Ozer. Flavour and Fragrance Journal 13 (4): 251-258. Sahin. (2006). 35. Application to the extraction of essential oil from rosemary leaves .. Springer 117 . Polissiou. G... G. M. Daferera. Physical Properties of Foods. Baghdadi. Russo M. 390– 394. (2004). Biological activities of the essential oils and methanol extract of Origanum vulgare ssp. A. J.

LA. MA. 8 (4): 561-573. Singh. GJ. Comparison of comprehensive two-dimensional gas chromatography and gas chromatography – mass spectrometry for the characterization of complex hydrocarbon mixtures.The case of ThujaOccidentalis L. PJ. Preliminary assay on the antioxidative activity of Laurus nobilis extracts. Ristic.. KP. (2003) . M. RL. Shelef. (1993).. Journal of Chromatography A.. T. Marin. Journal of Food Science. Kovacevic. JM. Phytotherapy Research. Genuit. S. 18: 713–717 Singh. J. Jyothi. Vukojevic.. Simic.. (1988). Efficacy of chlorine dioxide. Collin. 892: 29-46. D. 118 . Hachey. RK. PD. Journal of Wood Chemistry and Technology.. The variations of essential oil composition during the extraction process . Growth of enteropathogenic and spoilage bacteria in sage-containing broth and foods. N..Schoenmakers... N. (1984). Kundakovic... J. Bhunia.. Singh. AK. Simard. (2002). JLMM.. Simic A. 35. Sokovic. Stroshine. 720– 729. EK. S. Blomberg. Lebensmittelwissenchaften und Technologien.. 38(2): 83-89. Challenges and opportunities in essential oil processing industries. M. Bulgarelli. Oomen . Van Velzen. Grujic-Jovanovic. and Abies-Balsamea (L) Mill.. 49 (3): 737–740. ozone and thyme essential oil or a sequential washing in killing Escherichia coli O157:H7 on lettuce and baby carrots. W.... The chemical composition of some Lauraceae essential oils and their antifungal activities . G. Fitoterapia .74:613–616. (2000). Research & Industry. (2004)..

. 975:31–46. S. flavones and flavonols in some plant materials and their antioxidant activities. J. proanthocyanidins. L. In: Rahman.Sivropoulou. Fyfe. Stewart. Journal Of Supercritical Fluids. Nikolaou. (1996). Knez. Johnson. Hadolin M. 89:191–198 Skoula. 91: 1011 – 1022. P. Papanikolaou. EJ. Gotsiou.. Effect of oregano essential oil on microbiological and physico-chemical attributes of minced meat stored in air and modified atmospheres. G. Sahin.. (2002).. Skerget M. 119 . Skandamis. Smith. Food Chemistry. A. GJE.). Yilmaz. M. S. (1999). A chemosystematic investigation on the mono.. Gorris. Smith-Palmer.and sesquiterpenoids in the genus Origanum (Labiatae). Extractions with superheated water. A. 30 (2): 189-199. Antimicrobial properties of plant essential oils and essences against five important food-borne pathogens... Sonsuzer. Letters in Food Microbiology. Simonic. M. (Ed. MS. AR. (1998).. CB. Smid. Kokkini. (1999). (2005) Phenols.. E. New York: Marcel Dekker. 44: 1202–1205.. 26: 118–122. Handbook of Food Preservation. Hras. (2001). (2004) Optimization of supercritical CO2 extraction of Thymbra spicata oil.. C. Phytochemistry. (285–308). Journal of Agricultural and Food Chemistry. Z. Natural antimicrobials for food preservation.. 52: 649-657. PN.. Naxakis... LGM. Kotnik P. L. S. Journal of Applied Microbiology. Nychas. RM. Antimicrobial and cytotoxic activities of Origanum essential oils. Journal of Chromatography A..

. (2004a). TLM. 18: 409-413.. BE. Mathematical model for hydrodistillation of essential oils. Stamford. SA. EE. Sovová. Stashenko. Lima.o R. Effectiveness of Origanum vulgare L. Phytochromemediated production of monoterpenes in thyme seedlings.. Sarais. 881-889. 21 (6): 120 . Analysis of volatile secondary metabolites from Columbian Xylopia aromatica (Lamarck) by different extraction and headspace methods and gas chromatograhpy. Effect of essential oils on Aeromonas hydrophila in a culture medium and in cooked pork. Shigemot. Tanaka. Journal of Chromatography A. P. Tabata.) NE brown. Aleksovski. JR. Spar Eskilsson. 1025 (1): 93-103.. (2000).. Flavour And Fragrance Journal. Journal of Chromatography A. Comparison of different extraction methods for the analysis of volatile secondary metabolites of Lippia alba (Mill.. Yamaura. Journal of Food Protection. Giavedoni.. H.... ML. (2006). 56 (5): 406– 409. Jaramillo. E. EL. I. Journal of Chromatography A. Stecchini. Jaramillo.. EO. Food Control.Souza. 28: 2955–2957.. Trajano. Bjorklund. Analytical-scale microwaveassisted extraction. and evaluation of its in vitro antioxidant activity. S. 1025 (1): 105-113. essential oil to inhibit the growth of food spoiling yeasts. 902: 227–250. T. JR.. (1989). Martinez. grown in Colombia. BE. (1993). M. VN. Phytochemistry. Stashenko. (2007). Martinez. K. (2004b). EE.

Slump. EH. Simultaneous distillation-extraction of high-value volatile compounds from Cistus ladanifer L. Quantitative determination of phenolic diterpenes in rosemary extracts aspects of accurate quantification. (1996). Journal of Applied Microbiology. MA. 89: 901–909. Drosinos. Santos. Teixeira. L. 85: (1-2): 73-81.. GJE. E. G. Skandamis. A. C. M. Hildebrandt KS.. P. Journal of Food Protection 63 (5). Nychas. vacuum and modified atmosphere packaging conditions with or without the presence of oregano essential oil at 5 oC.. Valero. (2003). Journal of Chromatography A. 121 . Analytica Chimica Acta 584 (2): 439-446. K. GJE. MC.. Nychas. Ultee. RA.... (1995).. EJ. Salmeron. 995 (1-2): 119-125. (2007). International Journal of Food Microbiology. Alves. Tuley de Silva. S. 78: 593– 600. Antibacterial activity of 11 essential oils against Bacillus cereus in tyndallized carrot broth . 620–624. Thorsen.. Vienna: United Nations Industrial Development Organization Tsigarida.. Antimicrobial activity of carvacrol toward Bacillus cereus on rice.Tassou. Smid... (2000). (2003). A Manual on the Essential Oil Industry. Journal of Applied Bacteriology... A. Behaviour of Listeria monocytogenes and autochthonous flora on meat stored under aerobic. (2000). Effects of essential oil from mint (Mentha piperita) on Salmonella enteritidis and Listeria monocytogenes in model food systems at 4 oC and 10 oC. A. Steging. Mendes.

Wang. The effect of essential oils of basil on the growth of Aeromonas hydrophila and Pseudomonas fluorescens. 8: 303–313. 27 (1): 91-101. Ding.. L. É.... K. (1999).Van de Braak. Antifungal activities of thyme. vulgare ssp. Hajdύ.. X. (2007). TC. 84.. Dobos. Zhou. Voda.. (1998). MJ.. (2001). J.. Wilcock. E. 57(1-2): 95-98. Szabó. HQ.. B.. Journal of Food Safety. Li.. Coventry. (2003). 24 (5): 649-652. M. Ruiz-Navajas. Varga. I. Á. Centre for the Promotion of Imports from Developing Countries . J. and O.. 10 (1): 76-84 . Zhang. ZS. An overview of the ultrasonically assisted extraction of bioactive principles from herbs. Wan. Investigation of the Composition and Stability of the Essential Oils of Origanum vulgare ssp. Journal of Molecular Modeling. (2003). Németh. K.Rotterdam: CBI . GCJJ. Fast determination of essential oil from dried menthol mint and orange peel by solvent free microwave extraction using carbonyl iron powder as the microwave absorption medium.. Chromatographia. Essential Oils and Oleoresins: A Survey in the Netherlands and other Major Markets in the European Union. Boh. Fernández-López. A.. Ultrasonics Sonochemistry.. Veres. JA. ZM. Leijten. (2006a). J. M. 122 . Chinese Journal of Chemistry. Feng. clove and oregano essential oils. A quantitative structure–antifungal activity relationship study of oxygenated aromatic essential oil compounds using data structuring and PLS regression analysis.. K.. Vrtacnik. . M. Viuda-Martos. Pérez-Álvarez.. vulgare L. Vinatoru. SAAJ. Journal of Applied Microbiology. 152–158. Wang. Máthé. Y. hirtum (link) letswaart.. L.

. 123 . X.. GW. T. 8. L. European Journal of Lipid Science And Technology. Y. XL.. Wu. 1102 (12): 11-17. Marinova.. Quantitative determination of compounds in tobacco essential oils by comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry. Lu. 2002/113/EC: Official Journal L49 20/02/2002.. H.. Ding.. 1086 (12): 107-114. SK. L. (2000). He.. J. Yanishlieva. Liu... HQ.... Commission Decision of 23 January 2002 amending Commission Decision 1999/217/EC as regards the register of flavouring substances used in or on foodstuffs. Morikawa. J.. 108 (9): 776-793. Alcohol absorption inhibitors from bay leaf (Laurus nobilis): Structure-requirements of sesquiterpenes for the activity Bioorganic and Medicinal Chemistry. (2005). T. (2006). Kawahara. Xing.. Shimoda. NV. Liu. CY. Uemura. Journal of Chromatography A.. Zhu. Wang.. L. Improved solvent-free microwave extraction of essential oil from dried Cuminum cyminum L.. & Matsuda. Journal Of Chromatography A. Natural antioxidants from herbs and spices.. 2071-2077.. ZH. and Zanthoxylum bungeanum Maxim.. HJ. ZM. Kong. Wang. H. Recent advances in extraction of nutraceuticals from plants Trends In Food Science Technology 17 (6): 300-312. HW.. Yoshikawa. H. CL. Xu.. LJ. E. Li. Dong. Pokorny.Wang. Su. Zhou. Li.. L. Wang. (2006b). Zeng. H. (2006). M:.. Zhang. Y. Weller. TC. X.

APPENDIX A ANOVA AND TUKEY’S PAIRWISE COMPARISON TEST TABLES Table A.1.08 P 0. Hydrodistillation SFME-100%p SFME-40%p SFME-60%p SFME-80%p 533215 2772467 -748369 1490883 186557 2425809 413386 2652638 -2401211 -161958 -1466285 772968 -1239455 999797 -184700 2054552 42129 2281382 -892797 1346456 SFME-100%p SFME-40%p SFME-60%p Table A.006 Table A.899E+11 7. (Box-Cox transformed) Analysis of Variance for yield Source DF SS MS method 4 4.3. ANOVA results for monoterpene hydrocarbon concentrations obtained in the extraction of Origanum vulgare L.798E+10 Total 9 4. Tukey’s pairwise comparison test results for yield values obtained in the extraction of Origanum vulgare L. ANOVA results for yield values obtained for Origanum vulgare L.781E+12 F 14.44 0.2.391E+12 1. Analysis of Variance for monoterpene hydrocarbons Source DF SS MS F P method 4 4387 1097 0.098E+12 Error 5 3.776 Error 5 12427 2485 Total 9 16815 124 .

5.3 51. ANOVA results for specific gravity values obtained in the extraction of Origanum vulgare L.5 Total 9 425.29 0.566 Error 5 257.874 Error 5 28495 5699 Total 9 35072 Table A.01442 0.546 Table A.00180 Total 11 0.6.68 Error 8 0.00137 0. ANOVA results for refractive index values obtained in the extraction of Origanum vulgare L.00412 0.1 Table A. ANOVA results for sesquiterpene concentrations obtained in the extraction of Origanum vulgare L.0013746 P 0.0000722 Total 11 0.4.76 Error 8 0. ANOVA results for oxygenated compound concentrations obtained in the extraction of Origanum vulgare L.0 0.01854 P 0.062 125 . Analysis of Variance for sesquiterpenes Source DF SS MS F P method 4 167. Analysis of Variance for oxygenated compounds Source DF SS MS F P method 4 6578 1644 0. Analysis of Variance for refractive index Source DF SS MS F method 3 0.0007970 0.Table A.0002657 3.7.0005776 0.8 42.82 0. Analysis of Variance for specific gravity Source DF SS MS F method 3 0.

Table A.8. ANOVA results for solubility in ethanol values obtained in the extraction of Origanum vulgare L. Analysis of Variance for solubility in ethanol Source DF SS MS F P method 3 0.1441 0.0480 0.97 0.457 Error 7 0.3450 0.0493 Total 10 0.4891

Table A.9. ANOVA results for yield values obtained for Laurus nobilis L. Analysis of Variance for yield Source DF SS MS method 2 0.0000033 0.0000016 Error 3 0.0000008 0.0000003 Total 5 0.0000041

F 5.86

P 0.092

Table A.10. ANOVA results for monoterpene hydrocarbon concentrations obtained in the extraction of Laurus nobilis L. Analysis of Variance for monoterpene hydrocarbons Source DF SS MS F P method 2 124 62 0.22 0.818 Error 3 861 287 Total 5 984

Table A.11. ANOVA results for oxygenated compound concentrations obtained in the extraction of Laurus nobilis L. Analysis of Variance for oxygenated compounds Source DF SS MS F P method 2 23391 11695 1.19 0.417 Error 3 29552 9851 Total 5 52943

126

Table A.12. ANOVA results for sesquiterpene concentrations obtained in the extraction of Laurus nobilis L. Analysis of Variance for sesquiterpenes Source DF SS MS F method 2 37.840 18.920 24.67 Error 3 2.301 0.767 Total 5 40.141

P 0.014

Table A.13. Tukey’s pairwise comparison test results for sesquiterpene concentrations obtained in the extraction of Laurus nobilis L. Hydrodistillation SFME-100%p SFME-40%p -6.1149 1.2048 -9.7720 -2.4523 -7.3169 0.0027 SFME-100%p

Table A.14. ANOVA results for specific gravity values obtained in the extraction of Laurus nobilis L. Analysis of Variance for specific gravity Source DF SS MS F method 2 0.000170 0.000085 0.21 Error 6 0.002390 0.000398 Total 8 0.002560

P 0.814

Table A.15. ANOVA results for refractive index values obtained in the extraction of Laurus nobilis L. Analysis of Variance for refractive index Source DF SS MS method 2 0.0000042 0.0000021 Error 6 0.0000845 0.0000141 Total 8 0.0000887

F 0.15

P 0.864

127

Table A.16. ANOVA results for yield values obtained for Rosmarinus officinalis L. Analysis of Variance for yield Source DF SS MS method 2 0.0000303 0.0000152 Error 3 0.0000025 0.0000008 Total 5 0.0000328

F 18.20

P 0.021

Table A.17. Tukey’s pairwise comparison test results for yield values obtained in the extraction of Rosmarinus officinalis L. Hydrodistillation MAHD-100%p MAHD-40%p -0.0033149 0.0043149 0.0011851 0.0088149 0.0006851 0.0083149 MAHD-100%p

Table A.18. ANOVA results for monoterpene hydrocarbon concentrations obtained in the extraction of Rosmarinus officinalis L. Analysis of Variance for monoterpene hydrocarbons Source DF SS MS F P method 2 548.98 274.49 93.96 0.002 Error 3 8.76 2.92 Total 5 557.74

Table A.19. Tukey’s pairwise comparison test results for monoterpene hydrocarbon concentrations obtained in the extraction of Rosmarinus officinalis L. Hydrodistillation MAHD-100%p MAHD-40%p 0.393 14.678 15.838 30.123 8.303 22.588 MAHD-100%p

128

2826 MAHD-100%p 129 . Tukey’s pairwise comparison test results for sesquiterpene concentrations obtained in the extraction of Rosmarinus officinalis L.005 Table A.0672 P 0.3 60.90 -160.29 0.3 Table A.22.0 252.7225 -1. Hydrodistillation MAHD-100%p MAHD-40%p -70.5116 -2.01 -198.69 -133.0 15162.21.4287 49.00 Error 3 0.2099 0.0700 Total 5 7.80 -6.29 -95.1 Total 5 30504.49 MAHD-100%p Table A.8573 3.000 Error 3 180. Analysis of Variance for oxygenated compounds Source DF SS MS F P method 2 30324. ANOVA results for oxygenated compound concentrations obtained in the extraction of Rosmarinus officinalis L.23. Analysis of Variance for sesquiterpenes Source DF SS MS F method 2 6. Hydrodistillation MAHD-100%p MAHD-40%p -2. Tukey’s pairwise comparison test results for oxygenated compound concentrations obtained in the extraction of Rosmarinus officinalis L.3344 -0.4935 -0.1236 -3.Table A. ANOVA results for sesquiterpene concentrations obtained in the extraction of Rosmarinus officinalis L.20.

0000221 0. Analysis of Variance for specific gravity Source DF SS MS method 2 0.0000019 0.0000010 0.24.09 Error 6 0.0000648 0.0000590 Total 8 0.0003764 F 0.0000667 P 0. ANOVA results for specific gravity values obtained in the extraction of Rosmarinus officinalis L.Table A.0000108 Total 8 0.25.0003543 0.0000111 Error 6 0. Analysis of Variance for refractive index Source DF SS MS F method 2 0.915 130 .834 Table A.19 P 0. ANOVA results for refractive index values obtained in the extraction of Rosmarinus officinalis L.

Sign up to vote on this title
UsefulNot useful