Agilawood essential oil as well as preparation method and application thereof

Abstract

The invention relates to the technical field of plant essential oil extraction, and particularly discloses a preparation method of agilawood essential oil, which comprises the following steps: adding agilawood powder into a citric acid solution with the concentration of 0.5-1%, homogenizing under high pressure, and then washing and filtering to obtain pretreated agilawood powder; the pretreated agilawood powder is added into a compound enzyme solution, the compound enzyme solution is prepared from, by mass, 1-2.5 g / L of cellulase, 1.5-3 g / L of xylanase, 6-12 g / L of hydroxylated lecithin and 5-10 g / L of sodium chloride, and the pH value of the compound enzyme solution is 4-5.5; treating at 40-60 DEG C for 1-4 hours to obtain a treated mixed solution; and performing steam distillation extraction on the treated mixed solution to obtain the agilawood essential oil. The extraction of effective components such as sesquiterpenoids and chromone compounds in the agilawood is improved through two-stage pretreatment, the steam distillation extraction time is shortened, the effect and yield of steam distillation extraction are improved, and the extracted agilawood essential oil is high in sesquiterpenoids and chromone substance content and high in quality. The quality and the biological activity of the agilawood essential oil are improved.

Description

Technical Field

[0001] The present invention relates to the technical field of plant essential oil extraction, and particularly to an agarwood essential oil, a preparation method thereof and an application thereof. Background Art

[0002] Agarwood essential oil is a natural aromatic substance extracted from plants of the genus Aquilaria, and has unique aroma characteristics and biopharmaceutical activities such as anti-inflammatory, sedative, antioxidant and other effects, and is widely used in cosmetics such as high-end perfumes, medicine and health care fields. However, the extraction efficiency, component retention rate and production cost of agarwood essential oil are still the key problems restricting industrialization.

[0003] At present, the main extraction methods of agarwood essential oil include supercritical CO 2 extraction, subcritical low-temperature extraction, solvent extraction, steam distillation and other methods. Supercritical CO 2 extraction method and subcritical low-temperature extraction method have the advantages of complete component retention, no solvent residue, environmental protection, etc., and the yield is relatively high, but the equipment is expensive, the process is complex, the energy consumption is high, and it is difficult to produce on a large scale; the solvent extraction method has a relatively high yield, but the solvent residue needs subsequent purification, the environmental pollution risk is high, and the cost is relatively high. The equipment cost and operation cost of steam distillation method for extracting agarwood essential oil are relatively low, the technology is mature and easy to operate, does not involve organic solvents, and can effectively extract light and medium volatile compounds in agarwood, and has unique flavor characteristics. These components may be the most important aroma sources in agarwood essential oil. Therefore, the mainstream method in the current market is to extract agarwood essential oil by steam distillation method. However, the yield of steam distillation for extracting essential oil is relatively low, and usually only volatile components can be obtained, while characteristic components such as chromone compounds are lost. Compounds such as 2-(2-phenylethyl)chromone and 2-[2-(4-methoxyphenyl)ethyl]chromone are characteristic components of agarwood essential oil, and have anti-inflammatory, antibacterial, anti-inflammatory, antioxidant, anti-diabetic and anti-tumor activities, and are of great significance in the field of drug research and development and medical applications.

[0004] At present, the prior art discloses that enzymatic hydrolysis for pretreatment can improve the yield of effective essential oil, but it will reduce the content of fragrance characteristic components such as agaroaromadendrene and calamenene in agarwood essential oil, and reduce the quality of agarwood essential oil. For example, the literature (Li Mingyue, Shen Huajie, etc., Study on the Auxiliary Extraction of Agarwood Essential Oil by Pectinase Pretreatment, Journal of Forestry Engineering, 2017, 2(6): 55-59) discloses that pectinase pretreatment and ethanol extraction are used to obtain agarwood essential oil, which can improve the yield of essential oil, but the main components of the extracted essential oil are chromone components, and there are very few aroma characteristic components. Chinese Patent with Publication No. CN 118374316 B discloses a method for extracting agarwood essential oil and agarwood oil by hierarchical pretreatment. Agarwood powder, laccase, and dimethylaniline hydrochloride are first pretreated, and then steam distillation is used to obtain agarwood essential oil. The residue of the extracted essential oil is put into a eutectic solvent-water mixed solution, reacted for a period of time, and then extracted with an organic solvent to obtain agarwood oil. The method in this patent can improve the yield of agarwood essential oil extracted by steam distillation and the content of sesquiterpene compounds, but the content of chromone compounds in the extracted agarwood essential oil is extremely low, and the efficacy and biological activity of agarwood essential oil are limited, which will limit the further application of agarwood essential oil. Summary of the Invention

[0005] In view of the above deficiencies, the present invention provides a method for preparing agarwood essential oil. By performing multi-stage pretreatment on agarwood, the efficiency and yield of extracting essential oil by steam distillation are improved, and the quality of agarwood essential oil is enhanced. The specific technical solutions are as follows:

[0006] A method for preparing agarwood essential oil, comprising the following steps:

[0007] (1) Dry and crush agarwood to obtain agarwood powder;

[0008] (2) Add the agarwood powder to a citric acid solution with a concentration of 0.5% - 1%, perform high-pressure homogenization, and then wash and filter to obtain pretreated agarwood powder;

[0009] (3) Add the pretreated agarwood powder to a composite enzyme solution, where the composite enzyme solution includes the following raw materials by mass fraction: cellulase 1 - 2.5 g / L, xylanase 1.5 - 3 g / L, hydroxylated lecithin 6 - 12 g / L, sodium chloride 5 - 10 g / L, and the pH is 4 - 5.5; treat at 40 - 60 °C for 1 - 4 h to obtain a treatment mixture;

[0010] (4) Perform steam distillation extraction on the treatment mixture obtained in step (3) to obtain agarwood essential oil.

[0011] In the method for preparing agarwood essential oil of the present invention, first, agarwood powder is pretreated by high-pressure homogenization with a citric acid solution, which plays a certain role in breaking the cell wall structure of agarwood, improving the treatment effect of subsequent biological enzymes and the extraction efficiency of essential oil. During this step, an appropriate citric acid concentration needs to be controlled to avoid damaging the effective components in agarwood. Combining homogenization treatment, high-pressure homogenization can shorten the pretreatment time and improve the treatment effect; then, it is treated with a composite enzyme solution. The composite enzyme hydrolysis further opens the dissolution channels of agarwood essential oil and promotes the leaching of sesquiterpenoids and chromone components. The inventor found in the research that hydroxylated lecithin can not only protect the components of agarwood essential oil and is beneficial to the extraction of effective components of agarwood essential oil by steam distillation, but also improve the hydrolysis effect of the composite enzyme. Analyzing the reason, it may be that hydroxylated lecithin can promote the binding of cellulase and xylanase to the relevant sites of agarwood powder, improving the hydrolysis effect; sodium chloride can maintain the enzyme activity, and adding a certain amount of sodium chloride is beneficial to the steam distillation extraction of chromone compounds. Through the combined action of the components of the composite enzyme solution, the components of the agarwood essential oil extracted by steam distillation are enriched, and the quality and biological activity of the agarwood essential oil are improved.

[0012] Preferably, in the above method for preparing agarwood essential oil, in the step (2), the ratio of agarwood powder to the citric acid solution is 1 g: 4 - 6 ml, the high-pressure homogenization temperature is 20 - 35 °C, the pressure is 20 - 30 MPa, and the high-pressure homogenization is carried out for 3 - 5 min.

[0013] Preferably, in the above method for preparing agarwood essential oil, the ratio of the agarwood powder to the composite enzyme solution is 1 g: 4.5 - 7 ml, the activity of the cellulase is 4 - 6*10 4 U / g, and the activity of the xylanase is 1 - 2*10 4 U / g.

[0014] Preferably, in the above method for preparing agarwood essential oil, the composite enzyme solution comprises the following raw materials by mass fraction: 1.8 g / L of cellulase, 2.5 g / L of xylanase, 10 g / L of hydroxylated lecithin, and 8 g / L of sodium chloride.

[0015] Preferably, in the above method for preparing agarwood essential oil, in the step (4), the steam distillation is carried out for 5 - 8 h.

[0016] Preferably, in the above method for preparing agarwood essential oil, in the step (4), after steam distillation, oil-water separation is carried out, and then refining and purification are carried out to obtain agarwood essential oil. The refining and purification are carried out using an independently developed agarwood essential oil refining device (patent application number 202111566667.4), and the refining and purification temperature is 130 - 150 °C.

[0017] Preferably, in the method for preparing the agarwood essential oil, in the step (1), the agarwood is dried at 40-50 °C for 12-24 h, and after being pulverized, the powder has a mesh number of 60-80 meshes.

[0018] On the other hand, the present invention also provides an agarwood essential oil prepared by the above preparation method.

[0019] On the other hand, the present invention also provides the application of the above agarwood essential oil in the preparation of antioxidant cosmetics or skin care products or antibacterial drugs.

[0020] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0021] 1. In the method for preparing the agarwood essential oil of the present invention, through two-stage pretreatment, the leaching of effective components such as sesquiterpenoid compounds and chromone compounds in agarwood is improved, the steam distillation extraction time is shortened, the effect and yield of steam distillation extraction are improved, the contents of sesquiterpenoid and chromone substances in the extracted agarwood essential oil are high, the biological activity and fragrance persistence of the agarwood essential oil are enhanced, and the quality of the agarwood essential oil is improved.

[0022] 2. In the method for preparing the agarwood essential oil of the present invention, the extraction method is relatively simple, the extraction time is short, no organic solvent is required, it is safe, green, and environmentally friendly, and it has high antioxidant and antibacterial activities, further improving the application value of the agarwood essential oil, and is beneficial to the application of the agarwood essential oil in antioxidant cosmetics or skin care products or antibacterial drugs. Description of the Drawings

[0023] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following will briefly introduce the drawings required for description in the embodiments. Obviously, the drawings in the following description are some embodiments of the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0024] Figure 1 It is the GC-MS diagram of the agarwood essential oil prepared in Example 1 of the present invention;

[0025] Figure 2 It is the GC-MS diagram of the agarwood essential oil prepared in Comparative Example 1 of the present invention;

[0026] Figure 3 It is the scavenging effect of the agarwood essential oil in Test Example 1 of the present invention on DPPH free radicals;

[0027] Figure 4 It is the scavenging effect of the agarwood essential oil in Test Example 1 of the present invention on ABTS free radicals; Detailed Embodiments

[0028] The following is a detailed description of the specific embodiments of the present invention, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments. Unless otherwise defined, all the professional terms used hereinafter have the same meaning as commonly understood by those skilled in the art. The professional terms used herein are only for the purpose of describing specific embodiments and are not intended to limit the protection scope of the present invention. Unless otherwise specifically stated, various raw materials, reagents, instruments, and equipment used in the present invention can be obtained through market purchase or can be prepared by existing methods.

[0029] Example 1

[0030] A method for preparing agarwood essential oil, comprising the following steps:

[0031] (1) Dry agarwood at 50 °C for 24 h, pulverize it and then sieve it to obtain agarwood powder with a mesh size of 60 - 80 meshes;

[0032] (2) Add the agarwood powder to a citric acid solution with a mass percentage concentration of 0.8%, and the ratio of agarwood powder to citric acid solution is 1 g:5 ml. Homogenize it for 3 min under the process conditions of 30 °C and 25 MPa, then filter it, wash it with distilled water until neutral, and dry it to obtain pretreated agarwood powder;

[0033] (3) Add the pretreated agarwood powder to a complex enzyme solution, and the ratio of material to liquid is 1 g:5 ml. The complex enzyme solution includes the following raw materials by mass fraction: cellulase 1.8 g / L, xylanase 2.5 g / L, hydroxylated lecithin 10 g / L, sodium chloride 8 g / L, pH is 4.8, the activity of cellulase is 5*10 4 U / g, the activity of xylanase is 1*10 4 U / g, and the specific preparation method is: add cellulase, xylanase, hydroxylated lecithin and sodium chloride to a buffer solution with a pH of 4.8 and stir to mix evenly. The buffer solution is a citric acid - sodium citrate buffer solution; stir and process it at 50 °C for 2 h, and the stirring speed is 400 r / min to obtain a processed mixture;

[0034] (4) Perform steam distillation on the processed mixture in step (3) for 6 h, separate the oil - water mixture with a separating funnel, collect the essential oil, and then carry out refining and purification using a self - developed agarwood essential oil refining device (patent application number 202111566667.4). The refining and purification temperature is 130 °C to obtain agarwood essential oil.

[0035] Example 2

[0036] A method for preparing agarwood essential oil, comprising the following steps:

[0037] (1) Dry agarwood at 50 °C for 24 h, pulverize it and then sieve it to obtain agarwood powder with a mesh size of 60 - 80 meshes;

[0038] (2) Add the agarwood powder into a citric acid solution with a mass percentage concentration of 0.5%, and the ratio of the agarwood powder to the citric acid solution is 1 g:5 ml. Homogenize it for 3 min under the process conditions of 30 °C and 30 MPa, then filter, wash it with distilled water until neutral, and dry to obtain the pretreated agarwood powder;

[0039] (3) Add the pretreated agarwood powder into the complex enzyme solution, and the material-liquid ratio is 1 g:5 ml. The complex enzyme solution includes the following raw materials by mass fraction: cellulase 2.2 g / L, xylanase 1.8 g / L, hydroxylated lecithin 8 g / L, sodium chloride 8 g / L, with a pH of 4.8. The activity of cellulase is 5*10 4 U / g, and the activity of xylanase is 1*10 4 U / g. The specific preparation method is as follows: Add cellulase, xylanase, hydroxylated lecithin and sodium chloride into a buffer solution with a pH of 4.8 and stir well to obtain it. The buffer solution is a citric acid-sodium citrate buffer solution; Stir and process it at 50 °C for 2 h, and the stirring speed is 400 r / min to obtain the treated mixture;

[0040] (4) Perform steam distillation on the treated mixture in step (3) for 6 h, collect the essential oil, and then refine and purify it using the self-developed agarwood essential oil refining device (patent application number 202111566667.4). The refining and purification temperature is 130 °C to obtain the agarwood essential oil.

[0041] Comparative Example 1 Conventional steam distillation method

[0042] A method for preparing agarwood essential oil, comprising the following steps:

[0043] (1) Dry the agarwood at 50 °C for 24 h, crush it and then sieve it to obtain agarwood powder with a mesh size of 60-80 meshes;

[0044] (2) Add the agarwood powder into a citric acid solution with a mass percentage concentration of 0.8%, stir and process it at 40 °C for 3 h, then filter, wash it with distilled water until neutral, and dry to obtain the pretreated agarwood powder;

[0045] (3) Perform steam distillation on the pretreated mixture in step (2) for 24 h, separate the oil-water mixture using a separating funnel, collect the essential oil, and then refine and purify it using the self-developed agarwood essential oil refining device. The refining and purification temperature is 130 °C to obtain the agarwood essential oil.

[0046] Comparative Example 2

[0047] This comparative example is different from Example 1 in that the complex enzyme solution comprises the following raw materials by mass fraction: cellulase 2.3 g / L, hydroxylated lecithin 10 g / L, sodium chloride 8 g / L, and the other steps and parameters are the same as those in Example 1.

[0048] Comparative Example 3

[0049] This comparative example is different from Example 1 in that the complex enzyme solution comprises the following raw materials by mass fraction: cellulase 1.8 g / L, xylanase 2.5 g / L, sodium chloride 8 g / L, and the other steps and parameters are the same as those in Example 1.

[0050] Comparative Example 4

[0051] This comparative example is different from Example 1 in that the complex enzyme solution comprises the following raw materials by mass fraction: cellulase 1.8 g / L, xylanase 2.5 g / L, hydroxylated lecithin 10 g / L, and the other steps and parameters are the same as those in Example 1.

[0052] The agarwood essential oil was analyzed by gas chromatography-mass spectrometry (GC-MS). The specific results are as Figures 1-2 shown in Tables 1 and 2.

[0053] Sample pretreatment: Weigh an appropriate amount of agarwood essential oil sample, dissolve it with n-hexane, filter it through a 0.22 μm filter membrane, and then use it for GC-MS analysis; the test instrument is Agilent 7820A-5977E, and the test conditions are as follows:

[0054] Gas chromatography conditions: An HP-5MS 5% Phenyl Methyl siloxane (30 m * 0.25 mm * 0.25 μm) elastic quartz capillary column was used; the temperature programming was as follows: the column temperature was 40 °C, and it was heated to 300 °C at a rate of 5 °C / min and held for 10 min; the vaporization chamber temperature was 250 °C; the carrier gas was high-purity He (99.999%); the column head pressure was 43 kPa, and the flow rate of the carrier gas (He) was 1.0 ml / min; the injection volume was 1.0 μL, and the injection was made without splitting, and the solvent delay time was 6.0 min;

[0055] Mass spectrometry conditions: Electron impact (EI) ion source, electron energy 70 eV, interface temperature 280 °C, ion source temperature 230 °C, quadrupole temperature 150 °C, the tuning method was standard tuning, the electron multiplier voltage was 1516 kV, and the mass scanning range was 40 - 550 m / z.

[0056] As can be seen from Table 1, the characteristic components in Example 1 are agarospirol, widdrol / valencene, (2-(2-phenylethyl)chromone), and 2-[2-(4-methoxyphenyl)ethyl]chromone. The components of the agarwood essential oil are rich. For the aroma evaluation of the agarwood essential oils in Example 1 and Comparative Example 1, the evaluation result is that the fragrance of Example 1 is better and the fragrance retention is longer, and the quality of the agarwood essential oil is improved.

[0057] Table 1 Comparison of the Contents of the Main Characteristic Components of Aquilaria sinensis Essential Oil in Example 1 and Example 2

[0058]

[0059] As can be seen from Table 2, the preparation method of the Aquilaria sinensis essential oil of the present invention can improve the yield of the Aquilaria sinensis essential oil extracted by steam distillation, increase the contents of sesquiterpenoids and chromones in the Aquilaria sinensis essential oil, and improve the quality of the Aquilaria sinensis essential oil.

[0060] Table 2 Essential Oil Extraction Rates, Sesquiterpenoid and Chromone Contents of Each Treatment Group

[0061] Group Extraction rate (%) Sesquiterpenoids (%) Chromones (%) Example 1 0.75 63.51 24.66 Example 2 0.69 61.36 23.75 Comparative Example 1 0.35 80.62 Not detected Comparative Example 2 0.58 61.45 20.83 Comparative Example 3 0.64 58.34 22.59 Comparative Example 4 0.68 60.71 21.47

[0062] Experimental Example 1 Determination of the Antioxidant Activity of Aquilaria sinensis Essential Oil

[0063] The antioxidant activities of the Aquilaria sinensis essential oils prepared in Example 1 and Comparative Example 1 were determined as follows:

[0064] Determination of the scavenging ability of Aquilaria sinensis essential oil on DPPH free radicals:

[0065] Test method: After mixing the Aquilaria sinensis essential oil solutions of each concentration with 0.1 mmol·L -1 DPPH ethanol solution in a ratio of 1:3, place it in the dark at room temperature for 30 min, measure the absorbance at a wavelength of 517 nm, and record it as A 1 , use absolute ethanol to replace DPPH, and measure the absorbance in the same way, and record it as A 2 ; use the sample solvent to replace the sample, and measure the absorbance in the same way, and record it as A 0 . Use ascorbic acid as a positive control and measure it according to the same method. Calculate the scavenging rate according to the following formula:

[0066] Scavenging rate = [(A 0 - A 1 + A 2 ) / A 0 × 100%.

[0067] Determination of the scavenging ability of Aquilaria sinensis essential oil on ABTS free radicals: Take 7 mmol·L -1 of ABTS and 2.45 mmol·L -1 of potassium persulfate solution and mix them evenly in equal volumes, place them in the dark for 16 h, and dilute them with distilled water to an absorbance of 0.7 ± 0.02 before use to obtain the ABTS working solution. Mix the Aquilaria sinensis essential oil solutions of each concentration with the ABTS working solution in a ratio of 1:3, measure the absorbance at a wavelength of 734 nm after reacting for 6 min, and record it as A 1 , use the solvent to replace the sample, and measure the absorbance in the same way, and record it as A 0; Deionized water was used instead of ABTS, and the absorbance was measured in the same way, denoted as A 2 . Ascorbic acid was used as a positive control to measure the scavenging rates, and the scavenging rate calculation formula was the same as that for DPPH

[0068] The results are as Figure 3 and Figure 4 shown. It can be seen from the results that within the measured concentration range, the antioxidant activity increases with the increase in the concentration of agarwood essential oil. The antioxidant activity of the agarwood essential oil in Example 1 is significantly higher than that in Comparative Example 1, indicating that the preparation method of the agarwood essential oil of the present invention can enhance the antioxidant activity of the agarwood essential oil

[0069] The foregoing description of specific exemplary embodiments of the present invention is for purposes of illustration and exemplification. These descriptions are not intended to limit the invention to the precise forms disclosed, and obviously, many modifications and variations are possible in light of the above teachings. The purpose of selecting and describing the exemplary embodiments is to explain the specific principles of the invention and its practical applications, so that those skilled in the art can implement and utilize the various different exemplary embodiments of the invention, as well as various different selections and modifications. The scope of the invention is intended to be defined by the claims and their equivalents

Claims

1. A method for preparing agarwood essential oil, characterized in that: The following steps are involved: (1) drying and crushing agarwood to obtain agarwood powder; (2) adding the agarwood powder to a citric acid solution having a mass percentage concentration of 0.5% to 1%, homogenizing under high pressure, and then washing and filtering to obtain the pretreated agarwood powder; (3) adding the pretreated agarwood powder to a complex enzyme solution, wherein the complex enzyme solution comprises the following raw materials by mass fraction: 1-2.5 g / L of cellulase, 1.5-3 g / L of xylanase, 6-12 g / L of hydroxylated lecithin, 5-10 g / L of sodium chloride, and a pH of 4-5.5; treating at 40-60° C. for 1-4 h to obtain a treated mixed solution; (4) subjecting the treated mixed solution obtained in step (3) to steam distillation extraction to obtain agarwood essential oil.

2. The method for preparing agarwood essential oil according to claim 1, characterized in that: In the step (2), the ratio of agarwood powder to citric acid solution is 1g:4-6ml, the high-pressure homogenization temperature is 20-35°C, the pressure is 20-30MPa, and the high-pressure homogenization is performed for 3-5min.

3. The method for preparing agarwood essential oil according to claim 1, characterized in that: The ratio of agarwood powder to complex enzyme solution is 1g:4.5-7ml, and the activity of cellulase is 4-6*10 4 U / g, the xylanase activity is 1 to 2*10 4 U / g.

4. The method for preparing agarwood essential oil according to claim 3, characterized in that: The complex enzyme solution comprises the following raw materials by mass fraction: 1.8 g / L of cellulase, 2.5 g / L of xylanase, 10 g / L of hydroxylated lecithin, and 8 g / L of sodium chloride.

5. The method for preparing agarwood essential oil according to claim 1, characterized in that: In the step (4), the steam distillation is carried out for 5 to 8 hours.

6. The method for preparing agarwood essential oil according to claim 1, characterized in that: In the step (4), steam distillation is performed, followed by oil-water separation, and then refining and purification to obtain agarwood essential oil, and the refining and purification temperature is 130-150°C.

7. The method for preparing agarwood essential oil according to claim 1, characterized in that: In the step (1), the agarwood is dried at 40-50°C for 12-24 hours and crushed into powder with a mesh size of 60-80.

8. An agarwood essential oil, characterized in that: The agarwood essential oil is prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the agarwood essential oil as claimed in claim 8 in the preparation of antioxidant cosmetics or skin care products or antibacterial drugs.

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