Extraction of aromatic substances and essential oils from flowers of michelia alba

VIETNAM NATIONAL UNIVERSITY OF AGRICULTURE FACULTY OF BIOTECHNOLOGY GRADUATION THESIS Extraction of aromatic substances and essential oils from flowers of Michelia Alba Hanoi, 2022 VIETNAM NATIONAL UNIVERSITY OF AGRICULTURE FACULTY OF BIOTECHNOLOGY -  - GRADUATION THESIS TOPIC: Extraction of aromatic substances and essential oils from flowers of Michelia Alba Student : Dao Trung Nghia Class : K63CNSHE Major : Biotechnology Instructor : PhD Phi Thi Cam Mien HANOI, 2022 GUARANTEE I hereby certify that the research for this thesis is mine The information gathered is accurate and has never been utilized or published in any papers, theses, or other academic works I hereby certify that the information used to support this thesis was obtained from the sources and properly cited I bear full responsibility for these reassurances Hanoi, December 2022 Student Dao Trung Nghia ACKNOWLEDGEMENTS In order to complete this graduation thesis, in addition to my constant efforts, I have received a lot of attention and help from groups and individuals inside and outside the school First and foremost, I would like to offer my deep gratitude to PhD Phi Thi Cam Mien and teachers from Faculty of Biotechnology for imparting me critical and valuable knowledge during the study and training process at the Vietnam National University of Agriculture I would like to express my respect and appreciation to my supervisors Assoc Prof Nguyen Duc Bach and the teachers at the Institute for Microalgae and Pharmacosmetics (IMPC) for having enthusiastically guided, advised and supported me throughout the process of learning and study, as well as Institute for Microalgae and Pharmacosmetics (IMPC) for creating favourable conditions to help me finished my thesis Finally, I would like to send my sincerest thanks to my family and loved ones, who have supported and assisted me during practice time Due to limited time and knowledge, the thesis inevitably has certain limitations and shortcomings I would like to thank and appreciate the contributions from teachers, lecturers and students I sincerely thank! Hanoi, December 2022 Student Dao Trung Nghia CONTENTS Guarantee Acknowledgements CONTENTS LIST OF ABREVIATIONS LIST OF TABLES LIST OF FIGURES CHAPTER Introduction 10 1.1 Problem statement 10 1.2 Introduction 11 1.3 Aim, objectives and meaning of research 11 1.3.1 Aim of the research 11 1.3.2 Objectives 12 1.3.3 Meaning of the topic 12 1.3.4 New point of research 12 CHAPTER OVERVIEWS OF LITERATURE 13 2.1 General introduction to Michelia alba 13 2.1.1 Taxonomic classification and nomenclature 13 2.1.2 Botanical characteristics 13 2.1.3 Distribution and ecology 16 2.1.4 Phytochemistry 17 2.1.5 Traditional uses and potential application of Michelia alba 19 2.1.6 Bioactivities extracts of M alba 21 2.2 Introduction to essential oils 24 2.2.1 The concept of essential oils 24 2.2.2 Distribution of essential oils in nature 24 2.2.3 Physical properties of essential oils 25 2.2.4 The process of accumulating essential oils 26 2.2.5 Principles of essential oil production 27 2.2.6 Principle of essential oil extraction 27 2.2.7 Essential oil extraction methods 28 2.3 Introduction to perfumes and fragrances 34 2.3.1 Odor and odor detection threshold 34 2.3.2 Fragrances 35 2.3.3 Perfume production process 35 2.3.4 The technology of making perfume 36 CHAPTER MATERIALS AND METHODS 38 3.1 Time and place of study 38 3.2 Research subject and materials 38 3.3 Research content 38 3.4 Equipments and chemicals 38 3.5 Research methods 39 3.5.1 The process of extracting essential oils and investigating influencing factors 39 3.5.2 Comparative of M alba essential oils obtained from water distillation, steam distillation and water-steam entrainment distillation methods 40 3.5.3 Comparing two methods of separating flower scent 42 3.5.4 Studying on extraction of M alba essential oil with organic solvents (hexane, petroleum ether) 44 CHAPTER RESULTS AND DISCUSSION 46 4.1 Extraction results of M alba flower essential oil by distillation method 46 4.1.1 Finding the maximum amount of M alba essential oils obtained from water distillation, steam distillation and water-steam entrainment distillation methods 46 4.1.2 Comparative of M alba essential oils obtained from water distillation, steam distillation and water-steam entrainment distillation methods 47 4.2 Extraction results of M alba flower essential oil by separating flower scent method 49 4.2.1 Finding the maximum amount of M alba essential oils obtained from two methods of separating scent 49 4.2.2 Comparative of M alba essential oils obtained from cold and hot separating methods 51 4.3 Extraction results of M alba flower essential oil by using organic solvents (hexane, petroleum ether) 51 4.3.1 Finding the maximum amount of M alba essential oils by using hexane solvent 51 4.3.2 Comparative of M alba essential oils obtained from methods of using hexane and petroleum ether 53 4.4 Analyzing of M alba essential oil products obtained from various methods 54 CHAPTER CONCLUSIONS AND SUGGESTIONS 56 5.1 Conclusions 56 5.2 Suggestions 56 REFERENCES 57 LIST OF ABREVIATIONS M alba Michelia alba E coli Escherichia coli V parahaemolyticus Vibrio parahaemolyticus V cholerae Vibrio cholerae Flavobacterium spp Flavobacterium species T.cruzi Trypanosoma cruzi P falciparum Plasmodium falciparum A flavus Aspergillus flavus Na2SO4 Natri sulfat LIST OF TABLES Table 2.1 Classification and nomenclature of Michelia alba 13 Table 2.2 Morphological features of Michelia alba’s leaves, stems, flowers, fruits, and seeds 15 Table 3.1 The investigation of the effect of the amount of ingredients added into 200ml of palm stearin on the amount of essential oil obtained in cold separating method 42 Table 3.2 The investigation of the effect of the amount of ingredients added into 400ml of palm stearin on the amount of essential oil obtained in hot separating method 43 Table 3.3 The investigation of the effect of the amount of ingredients added into 1000ml of hexane solvent on the amount of essential oil obtained in extracting with hexane 44 Table 3.4 The investigation of the effect of the amount of ingredients added into 1000ml of petroleum ether solvent on the amount of essential oil obtained in extracting with petroleum ether 45 Table 4.1 The amount of essential oil obtain from water distillation method 46 Table 4.2 The amount of essential oil obtain from steam distillation method 47 Table 4.3 The amount of essential oil obtain from water-steam entrainment distillation method 47 Table 4.4 The optimal amount of essential oils obtained in cold separating method 49 Table 4.5 The optimal amount of essential oils obtained in hot separating method 50 Table 4.6 The optimal amount of essential oils obtained in using hexane solvent method 52 Table 4.7 The optimal amount of essential oils obtained in using petroleum ether solvent method 53 Table 4.8 The results of the experiment with hexane and petroleum ether 54 Table 4.9 Different physical states of essential oil products of three main methods 55 Amount of Volume of Time extraction The amount of flowers used (g) distilled water (hour) essential oil used (ml) 500 1000 obtain (%) 0.132 Table 4.2 The amount of essential oil obtain from steam distillation method 4.1.1.3 Experiment 3: Extracting M alba essential oil from water-steam entrainment distillation method and finding the amount of essential oil obtained After the extraction is complete, I obtain an amount of M alba essential oil by the water-steam entrainment distillation method with the ratio: 500g flowers/ 1000ml distilled water for hours below: Amount of Volume of Time extraction The amount of flowers used (g) distilled water (hour) essential oil used (ml) 500 1000 obtain (%) 0.189 Table 4.3 The amount of essential oil obtain from water-steam entrainment distillation method 4.1.2 Comparative of M alba essential oils obtained from water distillation, steam distillation and water-steam entrainment distillation methods The results of comparison of essential oils obtained from distillation methods are shown in Figure 4.1 below: 47 Figure 4.1 The maximum amount of M alba essential oils obtained by water distillation method, steam distillation and water-steam entrainment distillation According to figure 4.1, the maximum amount of essential oil obtained by water distillation method is 0.244%, steam distillation is 0.132% and water-steam entrainment distillation is 0.189% Accordingly, the amount of essential oil obtained by distillation from water will be the most optimal According to research documents, the extraction of essential oils by steam distillation method will give more concentrate and be applied more However, through experiments, it was shown that the content of essential oil from M Alba flowers obtained by water distillation method was higher than that of the two methods above For the method of distillation from water, steam and water-steam entrainment, the obtained essential oil has a dark yellow color and a characteristic odor of M alba flowers 48 4.2 Extraction results of M alba flower essential oil by separating flower scent method 4.2.1 Finding the maximum amount of M alba essential oils obtained from two methods of separating scent 4.2.1.1 Experiment 4: Extracting M alba essential oil from cold separating method After the extraction is complete, I obtain an amount of M alba essential oil by cold separating method with the ratio: 1.000g, 1.500g, 2.000g, 2.500g, 3.000g, and 3.500g/200 ml palm stearin below: Materials M alba flowers Volume of flowers used (g) Amount of palm stearin used (ml) Optimum essential oil obtained (%) 1.000 0.3524 1.500 0.3787 2.000 200 ml 0.3701 2.500 0.3688 3.000 0.3579 3.500 0.3115 Table 4.4 The optimal amount of essential oils obtained in cold separating method According to Table 4.4, the optimal amount of essential oil when giving the flower/palm oil ratios: 1000, 1500, 2000, 2500, 3000, 3500 (g) with 200ml of palm stearin, respectively: 0.3524, 0.3787, 0.3701, 0.3688, 0.3579, 0.3115 (%) The best amount of essential oil is 0.3787% when using ratio 1500g M alba flowers with 200ml of palm stearin This proves that not more ingredients will 49 give more essential oil, but to get the most optimal amount of essential oil depends on the amount of palm stearin that we use 4.2.1.2 Experiment 5: Extracting M alba essential oil from hot separating method After the extraction is complete, I obtain an amount of M alba essential oil by hot separating method with the ratio: 300g, 350g, 400g, 450g, 500g and 550g flower/400 ml palm oil below: Materials M alba flowers Amount of flowers used (g) Volume of palm stearin used (ml) Optimum essential oil obtained (%) 300 0.2676 350 0.2878 400 0.3015 400 450 0.3525 500 0.2723 550 0.2714 Table 4.5 The optimal amount of essential oils obtained in hot separating method According to Table 4.5, the optimal amount of essential oil when giving the flower/palm oil ratios: 300g, 350g, 400g, 450g, 500g, 550g with 400ml of palm oil, respectively: 0.2676, 0.2878, 0.3015, 0.3525, 0.2723, 0.2714 (%) The best amount of essential oil is 0.3525% when using ratio 450g M alba flowers with 400ml of palm 50 4.2.2 Comparative of M alba essential oils obtained from cold and hot separating methods Through the above results, we can see that the hot separation method gives more productivity This is probably due to the use of heat in the extraction process Furthermore, the product from both methods contains palm wax and palm oil, which will be extracted with alcohol Palm wax can be removed by filtration or centrifugation but the oil is difficult to remove Thus, the amount obtained is quite different 4.3 Extraction results of M alba flower essential oil by using organic solvents (hexane, petroleum ether) 4.3.1 Finding the maximum amount of M alba essential oils by using hexane solvent 4.3.1.1 Experiment 6: Extracting M alba essential oil by hexane After the extraction is complete, I obtain an amount of M alba essential oil by using organic solvents with the ratios: 100g, 150g, 200g, 250g, 300g and 350g flowers with 1000ml of hexane below: 51 Materials M alba flowers Amount of flowers used (g) Volume of hexane used (ml) Optimum essential oil obtained (%) 100 0.4667 150 0.6563 200 0.5004 1000 250 0.6625 300 0.4773 350 0.4712 Table 4.6 The optimal amount of essential oils obtained in using hexane solvent method According to Table 4.6, the optimal amount of essential oil when giving the flower/hexane ratios: 100g, 150g, 200g, 250g, 300g, 350g with 1000ml of hexane, respectively: 0.4667, 0.6563, 0.5004, 0.6625, 0.4773, 0.4712 (%) The best amount of essential oil is 0.6871% when using ratio 250g M alba flowers with 1000ml of hexane 4.3.1.2 Experiment 7: Extracting M alba essential oil by petroleum ether After the extraction is complete, I obtain an amount of M alba essential oil by using organic solvents with the ratios: 100g, 150g, 200g, 250g, 300g and 350g flowers with 1000ml of petroleum ether below: 52 Materials Amount of flowers used (g) M alba flowers Volume of petroleum ether used (ml) Optimum essential oil obtained (%) 100 0.4783 150 0.4924 200 0.5720 250 1000 0.5389 300 0.5112 350 0.4982 Table 4.7 The optimal amount of essential oils obtained in using petroleum ether solvent method According to Table 4.7, the optimal amount of essential oil when giving the flower ratios: 100g, 150g, 200g, 250g, 300g, 350g with 1000ml of petroleum ether, respectively: 0.4783, 0.4924, 0.5720, 0.5389, 0.5112, 0.4982 (%) The best amount of essential oil is 0.5720% when using ratio 200g M alba flowers with 1000ml of petroleum ether 4.3.2 Comparative of M alba essential oils obtained from methods of using hexane and petroleum ether After the experiment, the results of the experiment with hexane and petroleum ether are shown in the following table: 53 Table 4.8 The results of the experiment with hexane and petroleum ether Corresponding (%) Saturation point Concrete Absolute Amount of essential oi obtain (%) Hexane 250g flowers/ 0.0852 0.0487 0.6871 0.0752 0.0479 0.5720 1000ml hexane Petroleum 200g flowers/ ether 1000ml petroleum ether According to experimental evaluation and obtained results, the method of extracting M alba essential oil by petroleum ether solvent is more effective The hexane extraction product contains nine compounds and its main components are phenylethyl alcohol (39.27%) and indole (27.35%) The etherextracted product consisted of six compounds in which phenylethyl alcohol (35.06%) and linalool (36.14%) were the major constituents 4.4 Analyzing of M alba essential oil products obtained from various methods Solvent extraction is considered to be the most suitable method to extract essential oils from aromatic plants to avoid thermal decomposition of aromatic compounds However, this can only be done under safe conditions Steam distillation can be used in the extraction of essential oils, although some compounds can be lost to heat, and the resulting floral scents may cause 54 staining of the essential oils An interesting method is scent separation, especially hot incense separation, which gives high yields and smells most similar to fresh flowers Physical state Methods Distillation The oil is dark yellow and the scent of flowers rises - It is reddish brown in color and resembles solid wax Fragrance separation The smell of the liquid The liquid smell of the cold method is more like fresh flowers than the smell of the (hot/cold) hot method Solvents - Brown and wax-like semi-solid with a smell similar to fresh flowers but more pungent Extraction with ether oil showed a stronger scent than hexane extraction Table 4.9 Different physical states of essential oil products of three main methods Methods, such as hot separation scent, hexane solvent and ether oil yield high quality extracts from flowers because their constituents, linalool and indole, are the main constituents of the flowers used in many perfumes 55 CHAPTER CONCLUSIONS AND SUGGESTIONS 5.1 Conclusions The findings of this study prove and give conclusions as following: - For the method of distillation from water, steam and water-steam entrainment, the obtained essential oil has a dark yellow color and a characteristic odor of M alba flowers - Percent yield of finished products by cold and hot separation, hexane, petroleum ether at the saturation point respectively 0.3787, 0.3525, 0.6871 and 0.5720 (%) - The smell of M alba is obtained from the aromatic separation method more than other methods 5.2 Suggestions Based on the obtained conclusions, we recommend that: - Conducting a survey to collect feedback from users who tried a new Vietnamese dry perfume product on sensory criteria, hair care effectiveness and anti-dandruff effectiveness - Identifying chemical substances in M alba essential oil obtained by different methods 56 REFERENCES Vietnamese references: Phùng Thị Ái Hữu, 2012 Nghiên cứu chiết tách xác định thành phần hoá học tinh dầu sả chanh Cẩm Lệ - Đà Nẵng, Trường Đại học sư phạm – Đại học 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