Tumor cell-inhibiting composition

TECHNICAL FIELD

The present invention relates to a composition plant extract from a plant raw material based on a specific ratio. The composition has the efficiency of inhibiting the growth of tumor cells.

PRIOR ART

In the recent ten years, cancer ranks number one among the top ten causes of death, in which the lung cancer, hepatic carcinoma and intestinal cancer rank top three from both of male and female aspects. From female aspect, the fourth and fifth mortalities are breast cancer and pancreatic cancer, and from male aspect, the fourth and fifth ones are oral cancer and esophageal cancer. All of the domestic and overseas doctors and research scholars are devoted to the research of avoiding or reducing the generation or proliferation of tumor cell.

Pulmonary fibrosis refers to the pulmonary alveoli is being replaced by fibrous matter due to the effect of the pulmonary issue being subjected to phlogosis, so that the soft pulmonary issue becomes hard and loses its elasticity, gradually, the lungs would lose the ability of contraction, relaxation, and gas exchange. The patients would have the symptoms of brachypnea, chest tightness or dry cough, etc., severely, may not breathe and cause respiratory failure eventually.

Nowadays, the aforementioned diseases all have medicine and method to carry out a treatment or an inhibition. However, these medicines mostly are chemical-synthesized or radiotherapy, hence, during the treatment, many patients would have side effects, and the side effects would make the patients much more uncomfortable comparing to the illness from the diseases. This is a huge deficiency.

Taiwan cypress (Chamaecyparis formosensis) is one of the evergreen arbors in Conifers Cupressaceae, it is an important species among the needle-leaved trees. Taiwan cypress has some characteristics such as a scent without pungent smell, the wooden quality of hardness, the abilities not to be rotten or damaged by worms. Furthermore, the researches indicate that Taiwan cypress has the superior inhibitory function to typhoid, Escherichia coli, Achorion and Staphylococcus auresus, and Taiwan cypress also has an anti-inflammatory function.

Stout camphor tree (Cinnamomum micranthun) belongs to Lauraceous plants. Its appearance is tall, straight and delicate, hence, it is a perfect species for landscape design. Its wooden quality is superior and has a unique scent, which would also be the excellent material for wooden art craft and furniture. Stout camphor tree contains triterpenoids, superoxide dismutase, adenoside, polysaccharide, multiple proteins, and multiple vitamins, etc. It has the functions of anti-tumor, enhancing the immunity, reducing blood sugar and cholesterol and the like. Most of the related researches about the anti-cancer focus on the stout camphor tree. There is no research revealing that the stout camphor tree itself or its extract has specific function of anti-tumor.

Taiwan incense cedar (Calocedrus formosana) belongs to a Taiwan specific species, an evergreen big arbor of Cupressaceae Taiwan incense cedar. It has special scent, and its texture is compact and delicate. The research discloses that its extracts have the activities of anti-fungus, anti-oxidation and anti-inflammation. However, the researches related to the components of Taiwan incense cedar mainly are the extracts from branches and leaves, and less is about the extracts from wooden core. There is no research revealing that the extract of the Taiwan incense cedar has specific function of anti-cancer.

The recent researches discover that many plants (as aforementioned) have the functions of anti-inflammation or anti-oxidation, especially the distillated extracts of the plants would have the function to improve the cell membrane permeability for the nutrition and the oxygen needed by the cells getting into the cells and exhausting the waste produced by the cells out of the cells, so that the cell metabolism can be enhanced. Therefore, the cell may healthily be activated, and all the tissues, organs and immune system may function normally to provide human body the immunity, resistance and the ability of self-healing.

Prior art disclosed a method for preparing a plant extract liquid and its composition solution, wherein comprises shaving the plant raw material into pieces, and then putting the pieces in the main furnace of a steam kiln, and lighting up the wood to have the fractional distillation temperature to be set up. When each of the plant raw materials reach its steam reaction time, the steam extracts of each plants can be obtained via cooling and separation. The extract made by utilizing the method in prior art, no matter the oil layer or the water layer (purification), is different from the oil and the purification made by a distillation method carried out by a regular machine, which is characterized in that the extracts (oil or purification) made by the method would contain much richer compound from the distillation method carried out by the regular machine.

In addition, although these plants have the functions of anti-oxidation, anti-inflammation and the like individually, the recent researches know nothing about what treatment for diseases that a single plant can provide. Furthermore, a single plant offers a poor efficiency on the tumor inhibition and cannot be widely applied. These are the two deficiencies.

The applicants were aware of the deficiency of inhibition techniques, so that they invented a way to inhibit, reduce or ease the symptoms of the disease utilizing composite natural materials for improving the aforementioned conventional deficiency.

SUMMARY OF THE INVENTION

The main purpose of the present invention is providing a composition of natural plant extract (TGLON composition). The composition has the function of adjusting or facilitating the cell metabolism, and has the function of inhibiting the growth of tumor cells.

For achieving the aforementioned purpose, the characteristic of the present invention is utilizing the theory of mixing the plants grown from low, middle and high altitudes in Taiwan to design the composition of The Greatest Love of Nature composite plant extract (TGLON composition). Utilizing the following plant extracts with the specific ratio to combine a tumor cell-inhibited composition, wherein its components comprise: Taiwan cypress, stout camphor tree, Taiwan incense cedar, Cinnamomum camphora, Cymbopogon nardus, Litsea cubeba, Cinnamomum zeylanicum, Taiwan Cunninghamia lanceolatavar, Melaleuca alternifolia, Eucalyptus robusta smith, Cryptomeria japonica and Acacia confuse extracts.

The other characteristic of the present invention is the disclosed plant extract is the purification made by utilizing the method of prior art. The characteristic of the purification made by utilizing the method in prior art is having richer compound components comparing to a regular purification.

After a serial dilution of the TGLON composition, incubating the breast cancer cells (MCF-7), hepatic carcinoma cells (HepG2) and lung fibrous cells (MRC-5) for 72 hours, and then utilizing ELISA Reader (540 nm) to analyze the survival rate of these cell strains. As a result, when the composition is diluted by 40 folds, it has the inhibitory efficiency of cell growth above 50%.

DRAWINGS

FIG. 1 is the diagram disclosing the relative survival percentage of the composition of the present invention to the breast cancer cell strain.

FIG. 2 is the diagram disclosing the relative inhibitory percentage of the composition of the present invention to the breast cancer cell strain.

FIG. 3 is the diagram disclosing the relative survival percentage of the composition of the present invention to the lung fibrous cell strain.

FIG. 4 is the linear regression curve diagram disclosing the inhibitory percentage of the composition of the present invention to the lung fibrous cell strain.

FIG. 5 is the diagram disclosing the relative survival percentage of the composition of the present invention to the hepatic carcinoma cell strain.

FIG. 6 is the linear regression curve diagram disclosing the inhibitory percentage of the composition of the present invention to the hepatic carcinoma cell strain.

FIG. 7 is the analysis chart of the components of the composition of the present invention.

FIG. 8 is the analysis chart of the Taiwan cypress purification of the composition of the present invention.

FIG. 9 is the analysis chart of the stout camphor tree purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 10 is the analysis chart of the Taiwan incense cedar purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 11 is the analysis chart of the Cinnamomum camphora purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 12 is the analysis chart of the Cymbopogon nardus purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 13 is the analysis chart of the Litsea cubeba purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 14 is the analysis chart of the Cinnamomum zeylanicum purification of the composition of the present invention.

FIG. 15 is the analysis chart of the Taiwan Cunninghamia lanceolatavar purification of the composition of the present invention.

FIG. 16 is the analysis chart of the Melaleuca alternifolia purification of the composition of the present invention.

FIG. 17 is the analysis chart of the Eucalyptus robusta smith purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 18 is the analysis chart of the Cryptomeria japonica purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

FIG. 19 is the analysis chart of the Acacia confuse purification of the composition of the present invention, wherein the component less than 0.05% is not being labelled.

EMBODIMENT

For a better knowing and understanding of the tumor cell-inhibited composition of the present invention at the examiner's convenience, the present invention is illustrated in details by the examples with figures hereafter.

One of the tumor cell-inhibited composition (TGLON composition) of the present invention comprising the plant extract selected from the following group: Taiwan cypress, stout camphor tree, Taiwan incense cedar, Cinnamomum camphora, Cymbopogon nardus, Litsea cubeba, Cinnamomum zeylanicum, Taiwan Cunninghamia lanceolatavar, Melaleuca alternifolia, Eucalyptus robusta smith, Cryptomeria japonica and Acacia confuse. The present invention utilized the separation techniques to extract the effective component from the plant raw material, wherein the separation techniques are: distillation method, solvent extraction method, cold pressed method, supercritical carbon dioxide extraction and absorption method, etc. The TGLON composition itself has the function of adjusting or facilitating the cell metabolism, and has the function of inhibiting the growth of tumor cells.

For further illustrations, the plant extract of the present invention is the purification made by distillation method. For another further illustration, the purification is the purification made by the method disclosed in prior art, which is characterized in having richer compound components comparing to a regular purification.

The present invention also utilized the theory of mixing the plants grown from low, middle and high altitudes in Taiwan to be provided with a full efficiency in treatment, and further have the purification of the aforementioned plants mixed to become the tumor cell-inhibited composition. The weight percentage of the mixed composition is: 40%-50% of the Taiwan cypress purification, 10%-20% of the stout camphor tree purification, 10%-20% of the Taiwan incense cedar purification, 8%-10% of the Cinnamomum camphora purification, 8%-10% of the Cymbopogon nardus purification, 6%-8% of the Litsea cubeba purification, 6%-8% of the Cinnamomum zeylanicum purification, 4%-6% of the Taiwan Cunninghamia lanceolatavar purification, 2%-4% of the Melaleuca alternifolia purification, 2%-4% of the Eucalyptus robusta smith purification, 2%-4% of the Cryptomeria japonica purification and 2%-4% of the Acacia confuse purification.

Furthermore, utilizing the tumor cell-inhibited composition to carry out the survival experiments to the tumor cells based on the breast cancer cells (MCF-7), lung fibrous cells (MIRC-5) and hepatic carcinoma cell (HepG2).

Example 1

In the present example, the tumor cell-inhibited compositions are diluted by different multiples, and then are added into breast cancer cell strain (MCF-7) to be cultured. Furthermore, utilizing MTT colormetric method to test the composition inhibiting the growth of the breast cancer cell strain. The applicants observed whether the composition has the inhibitory efficiency to the growth of the breast cancer cell strain and its inhibitory concentration.

Firstly, culture the well-growing breast cancer cell strain into 96-well microplate, about 3,000 cells per well, then put the plate into the 5% of CO.sub.2, 37.quadrature. incubator overnight, then add the tumor cell-inhibited composition. Before adding the composition, filter the composition by 0.22 .mu.m filter to remove of bacteria. Then serial dilute the composition by distill water with two folds, completely shock the diluted composition into 96-well microplate, and the final concentration would be 40960.times., 20480.times., 10240.times., 5120.times., 2560.times., 1280.times., 640.times., 320.times., 160.times., 80.times., 40.times., 20.times. and 10.times.. Incubating them in the 5% of CO.sub.2, 37.degree. C. incubator for 72 hours, after 72 hours adding 1/10 of the total volume of MTT (5 mg/mL) solution into each well. Incubating them in the 5% of CO.sub.2, 37.degree. C. incubator for 4 hours, and then remove the mixed liquid in each well. Adding 100 .mu.l DMSO into each well to dissolve blue-purple crystal (formazan) in the cells, and furthermore place the 96-well microplate in the ELISA reader to read and analyze the data.

Please referring to Table 1 and FIGS. 1 and 2, the composition has the significant inhibitory efficiency to the growth of the breast cancer cell. When the dilution multiple is 80 folds, its inhibitory efficiency may achieve to 50% and more. When the dilution multiple is 10 folds to 40 folds, the inhibitory efficiency composition to the breast cancer cells may achieve to 98% and more.

Please further referring to FIG. 2, based on the data of the concentration of the composition (Y axis) and its corresponding inhibitory percentage (X axis), analyzing the two-dimensional linear regression and we may obtain a regression formula: Y=160.02-1.8222X+0.0043X.sup.2. If X=50 in this regression formula and we can get that Y=80, the value means the semi-inhibition concentration (50% Inhibition Concentration, IC.sub.50) of the composition to the breast cancer cell strain with 80 folds of diluted concentration.

TABLE-US-00001 TABLE 1 The growth inhibitory test of the composition of the present invention to the breast cancer cell strain (MCF-7) Concentration of Relative survival Relative the composition O.D. value percentage (%) inhibitory (Fold X) (mean .+-. SD) (mean .+-. SD) percentage (%) Control 1.188 .+-. 0.065 100.0 .+-. 5.5 0 40960X 1.195 .+-. 0.062 100.5 .+-. 5.3 0 20480X 1.162 .+-. 0.082 97.7 .+-. 6.9 2.3 10240X 1.134 .+-. 0.064 95.4 .+-. 5.4 4.6 5120X 1.242 .+-. 0.064 104.5 .+-. 5.4 0 2560X 1.295 .+-. 0.021 108.9 .+-. 1.8 0 1280X 1.234 .+-. 0.063 103.9 .+-. 5.3 0 640X 1.342 .+-. 0.030 113.0 .+-. 2.5 0 320X 1.283 .+-. 0.079 107.9 .+-. 6.6 0 160X 1.275 .+-. 0.074 107.3 .+-. 6.2 0 80X 0.579 .+-. 0.039 50.2 .+-. 3.3 49.8 40X 0.027 .+-. 0.004 2.3 .+-. 0.3 97.7 20X 0.043 .+-. 0.004 3.6 .+-. 0.3 96.4 10X 0.024 .+-. 0.005 2.0 .+-. 0.4 98.0

Example 2

In the present example, the tumor cell-inhibited compositions are diluted by different multiples, and then are added into lung fibrous cell strain (MRC-5) to be cultured. Furthermore, utilizing MTT colormetric method to test the composition inhibiting the growth of the lung fibrous cell strain. The applicants observed whether the composition has the inhibitory efficiency to the growth of the lung fibrous cell strain and its inhibitory concentration.

Firstly, culture the well-growing lung fibrous cell strain into 96-well microplate, about 3,000 cells per well, then put the plate into the 5% of CO.sub.2, 37.degree. C. incubator overnight, then add the tumor cell-inhibited composition. Before adding the composition, filter the composition by 0.22 .mu.m filter to remove of bacteria. Then serial dilute the composition by distill water with two folds, completely shock the diluted composition into 96-well microplate, and the final concentration would be 40960.times., 20480.times., 10240.times., 5120.times., 2560.times., 1280.times., 640.times., 320.times., 160.times., 80.times., 40.times., 20.times. and 10.times.. Incubating them in the 5% of CO.sub.2, 37.degree. C. incubator for 72 hours, after 72 hours adding 1/10 of the total volume of MTT (5 mg/mL) solution into each well. Incubating them in the 5% of CO.sub.2, 37.degree. C. incubator for 4 hours, and then remove the mixed liquid in each well. Adding 100 .mu.l DMSO into each well to dissolve blue-purple crystal (formazan) in the cells, and furthermore place the 96-well microplate in the ELISA reader to read and analyze the data.

Please referring to Table 2 and FIGS. 3 and 4, the composition has the significant inhibitory efficiency to the growth of the lung fibrous cell. When the dilution multiple is 80 folds, its inhibitory efficiency may achieve to 50% and more. When the dilution multiple is 10 folds to 40 folds, the inhibitory efficiency composition to the lung fibrous cells may achieve to 97% and more.

Please further referring to FIG. 4, based on the data of the concentration of the composition (Y axis) and its corresponding inhibitory percentage (X axis), analyzing the two-dimensional linear regression and we may obtain a regression formula: Y=83.595-0.787X+0.0008X.sup.2. If X=50 in this regression formula and we can get that Y=46, the value means the semi-inhibition concentration (50% Inhibition Concentration, IC.sub.50) of the composition to the breast cancer cell strain with 46 folds of diluted concentration.

TABLE-US-00002 TABLE 2 The growth inhibitory test of the composition of the present invention to lung fibrous cell strain (MRC-5) Concentration of Relative survival Relative the composition O.D. value percentage (%) inhibitory (Fold X) (mean .+-. SD) (mean .+-. SD) percentage (%) Control 1.177 .+-. 0.084 100.0 .+-. 7.1 0 40960X 1.134 .+-. 0.088 96.3 .+-. 7.5 3.7 20480X 1.157 .+-. 0.046 98.3 .+-. 3.9 1.7 10240X 1.171 .+-. 0.037 99.5 .+-. 3.1 0.5 5120X 1.216 .+-. 0.034 103.3 .+-. 2.9 0 2560X 1.104 .+-. 0.011 93.8 .+-. 0.9 6.2 1280X 1.189 .+-. 0.078 101.0 .+-. 6.7 0 640X 1.167 .+-. 0.046 99.1 .+-. 3.9 0.9 320X 1.153 .+-. 0.082 98.0 .+-. 7.0 2 160X 1.168 .+-. 0.036 99.2 .+-. 3.1 0.7 80X 1.124 .+-. 0.081 95.4 .+-. 6.9 4.6 40X 0.482 .+-. 0.012 40.9 .+-. 1.0 59.1 20X 0.032 .+-. 0.004 2.7 .+-. 0.3 97.3 10X 0.030 .+-. 0.004 2.5 .+-. 0.3 97.5

Example 3

In the present example, the tumor cell-inhibited compositions are diluted by different multiples, and then are added into hepatic carcinoma cell strain (HepG2) to be cultured. Furthermore, utilizing MTT colormetric method to test the composition inhibiting the growth of the hepatic carcinoma cell strain. The applicants observed whether the composition has the inhibitory efficiency to the growth of the lung hepatic carcinoma cell strain and its inhibitory concentration.

Firstly, culture the well-growing hepatic carcinoma cell strain into 96-well microplate, about 3,000 cells per well, then put the plate into the 5% of CO.sub.2, 37.degree. C. incubator overnight, then add the tumor cell-inhibited composition. Before adding the composition, filter the composition by 0.22 .mu.m filter to remove of bacteria. Then serial dilute the composition by distill water with two folds, completely shock the diluted composition into 96-well microplate, and the final concentration would be 40960.times., 20480.times., 10240.times., 5120.times., 2560.times., 1280.times., 640.times., 320.times., 160.times., 80.times., 40.times., 20.times. and 10.times.. Incubating them in the 5% of CO.sub.2, 37.degree. C. incubator for 72 hours, after 72 hours adding 1/10 of the total volume of MTT (5 mg/mL) solution into each well. Incubating them in the 5% of CO.sub.2, 37.degree. C. incubator for 4 hours, and then remove the mixed liquid in each well. Adding 100 .mu.l DMSO into each well to dissolve blue-purple crystal (formazan) in the cells, and furthermore place the 96-well microplate in the ELISA reader to read and analyze the data.

Please referring to Table 3 and FIGS. 5 and 6, the composition has the significant inhibitory efficiency to the growth of the hepatic carcinoma cell. When the dilution multiple is 160 folds, its inhibitory efficiency may achieve to 50% and more. When the dilution multiple is 10 folds to 40 folds, the inhibitory efficiency composition to the lung fibrous cells may achieve to 97% and more.

Please further referring to FIG. 6, based on the data of the concentration of the composition (Y axis) and its corresponding inhibitory percentage (X axis), analyzing the two-dimensional linear regression and we may obtain a regression formula: Y=0.0367X.sup.2-10.128X+66.72. If X=50 in this regression formula and we can get that Y=253, the value means the semi-inhibition concentration (50% Inhibition Concentration, IC.sub.50) of the composition to the hepatic carcinoma cell strain with 253 folds of diluted concentration.

TABLE-US-00003 TABLE 3 The growth inhibitory test of the composition of the present invention to the hepatic carcinoma cell strain (HepG2) Concentration of Relative survival Relative the composition O.D. value percentage (%) inhibitory (Fold X) (mean .+-. SD) (mean .+-. SD) percentage (%) Control 1.206 .+-. 0.027 100.0 .+-. 2.3 -- 40960X 1.199 .+-. 0.055 99.4 .+-. 4.6 0.6 20480X 1.276 .+-. 0.037 105.8 .+-. 3.1 -- 10240X 1.164 .+-. 0.023 96.5 .+-. 1.9 3.5 5120X 1.173 .+-. 0.023 97.2 .+-. 1.9 2.8 2560X 1.184 .+-. 0.028 98.2 .+-. 2.3 1.8 1280X 1.131 .+-. 0.039 93.8 .+-. 3.2 6.2 640X 1.163 .+-. 0.055 96.4 .+-. 4.5 3.6 320X 0.767 .+-. 0.036 63.5 .+-. 3.0 36.5 160X 0.342 .+-. 0.020 28.3 .+-. 1.7 71.7 80X 0.194 .+-. 0.019 16.0 .+-. 1.6 84.0 40X 0.017 .+-. 0.002 1.4 .+-. 0.2 98.6 20X 0.026 .+-. 0.002 2.2 .+-. 0.2 97.8 10X 0.051 .+-. 0.005 4.2 .+-. 0.4 95.8

Example 4

Please refers to Table 4 and FIG. 7, the composition can be confirmed that it has the function of inhibiting the growth of the tumor cells based on the aforementioned examples. Furthermore, the inventors have the tumor cell-inhibited composition analyzed by GC/MS, wherein the contained chemical components comprise: 1,8-Cineole, Camphor, Borneol, Terpinen-4-ol, .alpha.-Terpineol, (-)-Myrtenol, cis-Myrtanol, .alpha.-Pinene, Camphene, Sabinene, Myrcene, 1,4-Cineole, .alpha.-Terpinene, Cymene, Limonene, .gamma.-Terpinene, .alpha.-Terinolen, .alpha.-Fenchol, Citronellal, Citronellol, Nerol, Safrole, .alpha.-Terpinyl acetate, .beta.-Elemene, .alpha.-Cedrene, .beta.-Cedrene, .alpha.-Amorphene, .beta.-Selinene, .gamma.-2-Cadinene, .alpha.-Muurolene, .gamma.-Cadinene, .delta.-Cadinene, .beta.-Cadinene, .alpha.-Elemol, Cedrol, .alpha.-Cubebene, .gamma.-Eudesmol, .beta.-Eudesmol, tau-Muurolol.

TABLE-US-00004 TABLE 4 Peak Compound name 1 .alpha.-Pinene 2 Camphene 3 Sabinene 4 Myrcene 5 1,4-Cineole 6 .alpha.-Terpinene 7 Cymene 8 Limonene 9 1,8-Cineole 10 .gamma.-Terpinene 11 .alpha.-Terpinene 12 .alpha.-Fenchol 13 Camphor 14 Citronellal 15 Borneol 16 Terpinen-4-ol 17 .alpha.-Terpineol 18 (-)-Myrtenol 19 Citronellol 20 Nerol 21 cis-Myrtanol 22 Safrole 23 .alpha.-Terpinyl acetate 24 .beta.-Elemene 25 .alpha.-Cedrene 26 .beta.-Cedrene 27 .alpha.-Amorphene 28 .beta.-Selinene 29 .gamma.-2-Cadinene 30 .alpha.-Muurolene 31 .gamma.-Cadinene 32 .delta.-Cadiene 33 .beta.-Cedrene 34 .alpha.-Elemol 35 Cedrol 36 .alpha.-Cubebene 37 .gamma.-Eudesmol 38 .beta.-Eudesmol 39 tau-Muurolol

Please referring to FIGS. 8 to 19 and Table 5 to 16, the inventors further utilize GC/MS to analyze the components of the tumor cell-inhibited composition in each plant individually. The component less than 0.05% is not being labelled in the figures.

Please refer to Table 5 and FIG. 8, for chemical components contained in Taiwan cypress.

TABLE-US-00005 TABLE 5 Chemical Peak Compound name formula 1 .alpha.-Pinene C10H16 2 Camphene C10H16 3 Limonene C10H16 4 Terpinolene C10H16 5 (1R)-(+)-norinone C9H140 6 Camphor C10H16O 7 Borneol C10H18O 8 .alpha.-Terpineol C10H18O 9 (-)-Myrtenol C10H16O 10 Trans-Myrtanol C10H18O 11 cis-terpin hydrate C10H20O2 12 .alpha.-Terpinyl C12H20O2 acetate 13 .alpha.-Terpinyl formate C11H18O2 14 .alpha.-Cedrene C15H24 15 .gamma.-Cadinene C15H24 16 .beta.-Selinene C15H24 17 .alpha.-Muurolene C15H24 18 Cadinene C15H24 19 .delta.-Cadinene C15H24 20 .alpha.-Elemol C15H26O 21 Cedrol C15H26O 22 epi-.alpha.-Muurolene C15H26O 23 .alpha.-Cadinol C15H26O

Please refer to Table 6 and FIG. 9, for chemical components contained in stout camphor tree.

TABLE-US-00006 TABLE 6 1 Myrcene C10H16 2 1,4-Cineole C10H18O 3 .alpha.-Terpinene C10H16 4 p-Cymene C10H14 5 Limonene C10H16 6 Cineole:Eucalyptol C10H18O 7 .gamma.-Terpinene C10H16 8 .alpha.-Terpinolene C10H16 9 1-Terpineol C10H16 10 4-Terpineol C10H16 11 Safrole C10H10O2 12 3-Hydroxy-5-methyl-2- C9H15NO5 nitrocyclohexyl acetate 13 Myristicin C11H12O3 14 .delta.-Cadinene C15H24

Please refer to Table 7 and FIG. 10, for chemical components contained in Taiwan incense cedar.

TABLE-US-00007 TABLE 7 Chemical Peak Compound name formula 1 .alpha.-Pinene C10H16 2 .alpha.-Fenchene C10H16 3 p-Cymene C10H14 4 Limonene C10H16 5 Camphor C10H16O 6 4-Tepineol C10H18O 7 Terpineol Isomer C10H18O 8 .alpha.-Terpineol C10H18O 9 (-)-Myrtenol C10H16O 10 Verbenone C10H14O 11 Trans-Myrtanol C10H18O 12 Nopol* C11H18O 13 .beta.-Elemene C15H24 14 Myrtensaeure C10H14O2 15 .alpha.-Muurolene C15H24 16 .gamma.-Cadinene C15H24 17 .delta.-Cadinene C15H24 18 .alpha.-Cadinol C15H26O 19 3(10)-Caren-4-ol. C14H20O3 acetoacetic acid ester* 20 C12H20O2* C12H20O2 21 C12H20O2* C12H20O2 22 Oct-7-enol C10H20O 23 Piperitone* C10H16O 24 C10H20O2* C10H20O2 25 C10H20O2* C10H20O2 26 cis-terpin hydrate C10H20O2 27 C10H20O2* C11H20O2 28 C10H16* C10H16O *similarity scores were less than 95% among data base.

Please refer to Table 8 and FIG. 11, for chemical components contained in Cinnamomum camphora.

TABLE-US-00008 TABLE 8 Chemical Peak Compound name formula 1 .alpha.-(+)-Pinene C10H16 2 Camphene C10H16 3 .beta.-Pinene C10H16 4 p-cymene C10H14 5 Limonene C10H16 6 Cineole:Eucalyptol C10H18O 7 Linalool C10H18O 8 Camphor C10H16O 9 4-Terpineol C10H18O 10 .alpha.-Terpineol C10H18O 11 L-Citronellol C10H20O 12 Safrole C10H20O2 13 .alpha.-Copaene C15H24 14 (E)-.beta.-Farnesene C15H24 15 Nerolidol C15H26O 16 (-)-Guaiol C15H26O

Please refer to Table 9 and FIG. 12, for chemical components contained in Cymbopogon nardus.

TABLE-US-00009 TABLE 9 Chemical Peak Compound name formula 1 .alpha.-(+)-Pinene C10H16 2 Camphene C10H16 3 .beta.-Myrcene C10H16 4 p-Cymene C10H14 5 Limonene C10H16 6 Cineole:Eucalyptol* C10H18O 7 L-Fenchone C10H16O 8 Terpinolene C10H16 9 Linalool C10H18O 10 (+)-Fenchol C10H18O 11 Camphor C10H16O 12 (-)-Isopulegol* C10H18O 13 .beta.-Citronellal C10H18O 14 Borneol C10H18O 15 .alpha.-Terpineol C10H18O 16 .gamma.-Terpineol C10H18O 17 L-Citronellol C10H20O 18 .beta.-Geraniol C10H18O 19 Terpin Hydrate C10H20O2 20 para-menthane-3,8-diol C10H20O2 21 Citronellyl acetate C12H22O2 22 Eugenol C10H12O3 23 Geranyl acetate C15H24 24 .beta.-Elemene C15H24 25 .alpha.-Cedrene C15H24 26 .beta.-Cedrene C15H24 27 GERMACRENE D C15H24 28 .alpha.-Muurolene C15H24 29 .gamma.-Cadinene C15H24 30 .delta.-Cadinene C15H24 31 .alpha.-Elemol C15H26O 32 Cedrol C15H26O 33 .gamma.-Eudesmol C15H26O 34 tau-Cadinol C15H26O 35 tau-Muurolo* C15H26O *similarity scores were less than 95% among data base.

Please refer to Table 10 and FIG. 13, for chemical components contained in Litsea cabeba.

TABLE-US-00010 TABLE 10 Chemical Peak Compound name formula 1 .alpha.-Pinene C10H16 2 Sabinene C10H16 3 .beta.-Pinene C10H16 4 Cineole:Eucalyptol C10H18O 5 Camphor C10H16O 6 Terpinene-4-ol C10H18O 7 .alpha.-Terpineol C10H18O 8 Myrtenol C10H16O 9 .beta.-Citral C10H16O 10 (-)-trans-Myrtanol C10H18O 11 .alpha.-Citral C10H16O 12 Safrole C10H10O2 13 .alpha.-Terpinyl acetate C12H20O2 14 .beta.-Elemene C15H24 15 Caryophylene C15H24 16 .alpha.-Muurolene C15H24 17 .gamma.-Cadinene C15H24 18 .delta.-Cadinene C15H24 19 .alpha.-Elemol C15H26O 20 Cubenol* C15H26O 21 .gamma.-Eudesmol C15H26O 22 tau-Cadinol C15H26O 23 .beta.-Eudesmol C15H26O 24 tau-Muurolol* C15H26O *similarity scores were less than 95% among data base.

Please refer to Table 11 and FIG. 14, for chemical components contained in Cinnamomum zeylanicum.

TABLE-US-00011 TABLE 11 Chemical Peak Compound name formula 1 (+)-Fenchol C10H18O 2 Camphor C10H16O 3 Borneol C10H18O 4 .alpha.-Terpineol C10HI18O 5 (-)-Myrtenol C10H16O 6 (-)-trans-Myrtanol C10H18O 7 Cinnamaldehyde C10H16O 8 .beta.-Elemene C15H24 9 Cinnamyl acetate C11H12O2 10 Cadinene isomer C15H24 11 .beta.-Selinene C25H24 12 .alpha.-Muurolene C15H24 13 .gamma.-Cadinene C15H24 14 .delta.-Cadinene C15H24 15 .alpha.-Elemol C15H26O 16 tau-Cadinol C15H26O 17 .alpha.-Cadinol C15H26O

Please refer to Table 12 and FIG. 15, for chemical components contained in Taiwan Cunninghamia lanceolatavar.

TABLE-US-00012 TABLE 12 Peak Compound name 1 .alpha.-Pinene 2 Camphene 3 Myrcene 4 Limonene 5 .alpha.-Terpinolen 6 .alpha.-Fenchol 7 Camphor 8 Borneol 9 .alpha.-Terpineol 10 .beta.-Elemene 11 .alpha.-Cedrene 12 Caryophyllene 13 .beta.-Cedrene 14 Cedrol

Please refer to Table 13 and FIG. 16, for chemical components contained in Melaleuca alternifolia.

TABLE-US-00013 TABLE 13 Chemical Peak Compound name formula 1 .alpha.-Pinene C10H16 2 Sabinene C10H16 3 .alpha.-Terpinene C10H16 4 p-Cymene C10H24 5 Limonene C10H16 6 1,8-Cineole C10H18O 7 .gamma.-Terpinene C10H16 8 Terpinolene C10H16 9 Terpinen-4-ol C10H18O 10 .alpha.-Terpineol C10H18O 11 Aromadendrene C15H24 12 Ledene C15H24 13 .delta.-Cadinene C15H24 14 Globulol C15H26O 15 Viridiflorol C15H26O

Please refer to Table 14 and FIG. 17, for chemical components contained in Eucalyptus robusta smith.

TABLE-US-00014 TABLE 14 Chemical Peak Compound name formula 1 .alpha.-Pinene C10H16 2 p-Cymene C10H14 3 Limonene C10H16 4 Cineole C10H18O 5 (-)-Borneol C10H18O 6 .alpha.-Terpineol C10H18O 7 (-)-Myrtenol C10H16O 8 trans-Myrtanol C10H18O 9 .beta.-Elemene C15H24 10 .beta.-Panasinsene C15H24 11 .alpha.-Cedrene C15H24 12 Caryophylene C15H24 13 .delta.-Cadinene C15H24 14 .alpha.-Elemol C15H26O 15 .alpha.-Cadinol* C15H26O *similarity scores were less than 95% among data base.

Please refer to Table 15 and FIG. 18, for chemical components contained in Cryptomeria japonica.

TABLE-US-00015 TABLE 15 Chemical Peak Compound name formula 1 .alpha.-Pinene C10H16 2 Sabinene C10H16 3 .delta.-3-Carene C10H16 4 Limonene C10H16 5 Cineole:Eucalyptol C10H18O 6 .alpha.-Cubebene C15H24 7 Caryophyllene C15H24 8 N/A* C15H24 9 .alpha.-Humulene C15H24 10 N/A* C15H24 11 .gamma.-Muurolene* C15H24 12 .alpha.-Muurolene C15H24 13 (-)-Cedreanol* C15H26O 14 .delta.-Cadinene C15H24 15 (+)-Epizonarene C15H24 16 4.10-dimethyl-7-isopropyl- C15H24 bicyclo(4.4.0)-1,4-decadiene 17 .alpha.-Elemol C15H26O 18 N/A* C15H24 19 N/A* C15H24 20 .gamma.-Eudesmol C15H26O 21 tau-Cadinol* 22 .delta.-Cadinol C15H26O 23 .beta.-Eudesmol C15H26O 24 10-epi-.gamma.-Eudesmol C15H26O 25 N/A* C15H24 26 Kaur-16-ene C20H32 *similarity scores were less than 95% among data base.

Please refer to Table 16 and FIG. 19, for chemical components contained in Acacia confuse.

TABLE-US-00016 TABLE 16 Peak Compound name Chemical formula 1 L-Fenchone C10H16O 2 (1R)-(+)-Norinone C9H14O 3 Camphene C10H16O 4 .alpha.-Terpineol C10H18O 5 (-)-Myrtenol C10H16O 6 (-)-cis-Myrtanol C10H18O 7 cis-Terpin hydrate* C10H20O2 *similarity scores were less than 95% among data base.

The aforementioned detailed illustrations are the specific explanations for the workable examples of the present invention. However, these examples are not used as a limitation to the scope of claims of the present invention. The equal application or modification which falls in the scope of the present invention is included in the scope of the present application.