A method for extracting patchouli oil by ultrasonic-microwave synergistic assistance of low eutectic solvent

Abstract

The present application relates to the technical field of plant essential oil extraction, and particularly relates to a method for extracting patchouli essential oil by ultrasonic-microwave synergistic assistance of low eutectic solvent. The method comprises the following steps: 1) mixing choline chloride and a hydrogen bond donor, adding water, and mixing to obtain a low eutectic solvent; 2) mixing patchouli raw materials and the low eutectic solvent in step 1) to perform ultrasonic-microwave synergistic assistance extraction, and obtaining an extraction mixture; 3) performing water vapor distillation on the extraction mixture in step 2), collecting a distillate, and separating to obtain patchouli essential oil; wherein the hydrogen bond donor is a compound mixture of organic acids and polyhydric alcohols. The method forms a synergistic effect of ultrasonic cavitation effect and microwave dielectric heating, fully breaks plant cell walls, greatly shortens the extraction period, and improves the essential oil yield; meanwhile, the coupling of the synergistic physical field and DES can induce in-situ dehydration conversion of patchouli alcohol into β-patchoulene and other sesquiterpenes, and directionally prepare "olefin type" essential oil.

Description

Technical Field

[0001] This invention relates to the field of plant essential oil extraction technology, and in particular to a method for extracting patchouli essential oil using an ultrasound-microwave synergistic-assisted eutectic solvent. Background Technology

[0002] Patchouli ( Pogostemon cablin As a traditional medicinal and spice plant, patchouli essential oil, extracted from its stems and leaves, is rich in characteristic sesquiterpenoids. It possesses a rich and long-lasting aroma, along with multiple biological activities such as antibacterial, anti-inflammatory, and antioxidant properties, making it highly valuable in high-end daily chemical fragrances, natural food preservation, and topical pharmaceutical preparations. The value of the essential oil is directly determined by its internal volatile components. Developing efficient, green, and controllable extraction processes for active components is a core research direction for promoting the high-value processing of patchouli raw materials.

[0003] Currently, steam distillation (SD) is the most common industrial method for extracting patchouli essential oil. This process eliminates the need for organic solvents and avoids solvent residue risks, but it suffers from significant processing drawbacks. The extraction process is time-consuming, with a production cycle of 8-10 hours, resulting in high energy consumption and low production efficiency. Prolonged high-temperature heating can easily lead to the degradation and volatilization of heat-sensitive terpenoids. Furthermore, this process has weak cell wall disruption capabilities, resulting in insufficient release of intracellular essential oils and low raw material utilization. While organic solvent extraction can slightly increase the amount of essential oil dissolved, the extraction reagents often contain biotoxic substances, leading to the potential for harmful solvent residues in the finished product. The associated solvent separation and recovery processes are cumbersome, resulting in high production costs, which does not align with the industry trend of green and sustainable processing of natural products.

[0004] Deep eutectic solvents (DES) are low-temperature eutectic mixtures formed by hydrogen bond acceptors and donors through intermolecular hydrogen bonds. They possess green advantages such as low toxicity, biodegradability, low saturated vapor pressure, and tunable molecular structure, and are increasingly being applied in the extraction of natural active substances from plants. Current extraction methods only use total product yield as the evaluation criterion in the DES screening stage, failing to consider the solvent hydrogen bond donor combination, blending ratio, viscosity, polarity, and other physicochemical parameters in relation to the solubility and compatibility of different polar components of terpenes and terpene alcohols in patchouli essential oil. This makes it difficult to balance overall extraction efficiency with the selective enrichment of target sesquiterpenes. Furthermore, current technologies do not fully control the effect of system water content on extraction efficiency. Too low a water content increases solvent viscosity and hinders mass transfer and diffusion, while too high a water content weakens the hydrogen bonding ability between the solvent and essential oil components; both situations significantly reduce the essential oil extraction effect.

[0005] However, existing research mostly focuses on the simple combination of a single physical field with conventional solvents or DES. There is a lack of systematic research and process integration regarding the mutual influence and coupling effects of ultrasound and microwave energy fields acting simultaneously on the same extraction system. Furthermore, the recovery and recycling of DES is a key economic factor restricting its large-scale application, and related systematic research remains insufficient.

[0006] In summary, existing patchouli essential oil extraction technologies cannot simultaneously address the needs of extraction efficiency, product component controllability, bioactivity, and green production. The industry urgently needs to develop an integrated extraction method that utilizes a dual physical field of ultrasound and microwave to optimize the eutectic solvent, thereby simultaneously solving multiple technical pain points of traditional processes, such as long processing time, low yield, significant loss of heat-sensitive components, inability to directionally enrich components, and difficulty in recycling solvents. Summary of the Invention

[0007] The purpose of this invention is to overcome the shortcomings of existing patchouli essential oil extraction processes, such as long extraction time, low extraction efficiency, and poor raw material utilization. It also addresses the technical problems of uncontrollable essential oil components, limited functionality, weak antioxidant activity, and the difficulty in large-scale recycling of green solvents obtained through traditional processes. Therefore, this invention provides a method for extracting patchouli essential oil using an ultrasonic-microwave synergistic-assisted eutectic solvent. This method uses choline chloride as a hydrogen bond acceptor and a eutectic solvent (DES) as the extraction medium, employing a mixture of organic acids and polyols as hydrogen bond donors. This DES has moderate viscosity and excellent penetration and dissolution capabilities, enabling selective enrichment of sesquiterpenes. During extraction, the ultrasonic cavitation effect and microwave dielectric heating work synergistically to thoroughly break down plant cell walls, significantly shortening the extraction cycle and increasing the essential oil yield. Simultaneously, the coupling of the synergistic physical field with the DES induces in-situ dehydration of patchouli alcohol into sesquiterpenes such as β-patchoulene, directionally preparing "ene-type" essential oils. The resulting essential oils exhibit resistance to DPPH· and ABTS. + The free radical scavenging ability is significantly improved, and the antioxidant activity is stronger. The extracted DES can be recycled multiple times after simple concentration and dehydration, making it green and economical. This invention simultaneously solves the problems of high energy consumption, low raw material utilization, single product function, and inability to reuse solvents on a large scale in traditional processes, providing a new technical solution for the green, efficient, and targeted processing of patchouli essential oil.

[0008] To achieve the above objectives, in a first aspect, the present invention provides a method for extracting patchouli essential oil using an ultrasound-microwave synergistic-assisted eutectic solvent, the method comprising the following steps: 1) Mix choline chloride with a hydrogen bond donor, add water, and mix to prepare a eutectic solvent; 2) Mix the patchouli raw material with the eutectic solvent described in step 1) and perform ultrasonic-microwave synergistic assisted extraction to obtain an extraction mixture; 3) The extracted mixture described in step 2) is subjected to steam distillation, the distillate is collected, and patchouli essential oil is obtained by separation; The hydrogen bond donor is a mixture of organic acids and polyols.

[0009] Preferably, in step 1), the hydrogen bond donor is a mixture of malic acid and glycerol.

[0010] More preferably, the molar ratio of malic acid to glycerol in the compound mixture of malic acid and glycerol is 1:1 to 2.

[0011] Preferably, in step 1), the molar ratio of choline chloride to the hydrogen bond donor is 1:1~2.

[0012] Preferably, in step 1), the mass fraction of water in the eutectic solvent is 30~50wt%.

[0013] Preferably, in step 1), the mixing conditions include: a temperature of 75~85℃, a stirring rate of 200~400rpm, and a time of 60~120min.

[0014] Preferably, in step 2), the ratio of the mixed material to the liquid is 1:8~12 g / mL.

[0015] Preferably, in step 2), the conditions for ultrasound-microwave synergistic assisted extraction include: ultrasound power 200~400W, ultrasound time 15~25min, microwave power 250~350W, and microwave time 1~5min.

[0016] Preferably, in step 3), the steam distillation time is 1 to 3 hours.

[0017] Preferably, the method further includes: collecting the residual eutectic solvent aqueous solution after steam distillation, filtering to remove solid residue, concentrating to remove water until the mass fraction of water in the residual eutectic solvent is 30-50 wt%, and recovering the eutectic solvent for recycling.

[0018] Secondly, the present invention provides a patchouli essential oil prepared according to the method described in the present invention, wherein the normalized relative content of the gas chromatographic peak area of ​​the sesquiterpene components in the patchouli essential oil is ≥94%, the normalized relative content of the gas chromatographic peak area of ​​β-patchoulene is ≥41%, and the normalized relative content of the gas chromatographic peak area of ​​the alcohol components is ≤3.5%.

[0019] Thirdly, the present invention provides the application of patchouli essential oil as described herein in the preparation of antioxidants, cosmetics or food preservatives.

[0020] In the above technical solution, this invention selects a eutectic solvent (DES) with choline chloride as the hydrogen bond acceptor and a mixture of organic acids and polyols as hydrogen bond donors as the extractant for patchouli essential oil. The solvent viscosity, polarity, and hydrogen bond network are controlled by selecting and combining different types of hydrogen bond donors. The active functional groups such as carboxyl and hydroxyl groups contained in the hydrogen bond donors can form strong hydrogen bonds with sesquiterpene alcohols, effectively reducing the system viscosity and mass transfer resistance. The combination of specific hydrogen bond donors can produce a synergistic effect, making the DES significantly more selective for extracting characteristic sesquiterpenes from patchouli essential oil than a single-component system. Furthermore, this solvent is non-toxic and biodegradable, and when combined with rotary evaporation concentration technology, it can be reused multiple times while maintaining good performance, significantly reducing processing material costs and solving the problems of residual organic solvents, poor selectivity of single DES extraction, and difficulty in recycling in traditional patchouli essential oil extraction processes.

[0021] Meanwhile, the extraction method of this invention combines the aforementioned DES with ultrasonic-microwave synergistic extraction technology. Utilizing the strong shear force generated by ultrasonic cavitation and microjets to break down the dense cell walls of patchouli, combined with microwave dielectric heating to achieve uniform and rapid internal heating of the material, the synergistic effect of these two technologies effectively opens up the essential oil dissolution channels, increasing the essential oil yield from 1.14% using traditional steam distillation to 3.54%, and significantly shortening the extraction time from 10 hours to 3.5 hours. This represents a significant improvement in yield compared to the single physical field-assisted method. The coupling effect of the synergistic physical field and the weakly acidic DES system can induce in-situ dehydration of patchouli alcohol into sesquiterpenes such as β-patchoulene, transforming the chemical characteristics of the essential oil from the traditional "alcohol type" to the "ene type," achieving targeted regulation of essential oil components. Simultaneously, the mild and controllable extraction system reduces the degradation of heat-sensitive components.

[0022] Furthermore, the synergistic combination of various sesquiterpenes in the "olefinic" essential oil obtained by this invention endows it with excellent free radical scavenging ability, particularly against DPPH· and ABTS. + IC of free radicals 50 The values ​​reached 3.76 g / L and 0.77 g / L respectively, significantly better than traditional steam distillation and other single physical field-assisted methods. It can be directly used as a natural antioxidant raw material in the fields of daily chemicals and food preservation. The entire extraction process is green and low-energy, the solvent can be recycled and reused, and it is suitable for continuous industrial production, providing a reliable and economical technical route for the targeted preparation of highly active patchouli essential oil.

[0023] Other features and advantages of the present invention will be described in detail in the following detailed description section. Attached Figure Description

[0024] The accompanying drawings are provided to further illustrate the invention and form part of the specification. They are used together with the following detailed description to explain the invention, but do not constitute a limitation thereof. In the drawings: Figure 1 Image A is a scanning electron microscope (SEM) image of untreated patchouli leaf powder; image B is a SEM image of patchouli leaf powder residue after treatment by the extraction method of Comparative Example 1; image C is a SEM image of patchouli leaf powder residue after treatment by the extraction method of Comparative Example 2; image D is a SEM image of patchouli leaf powder residue after treatment by the extraction method of Example 1. Figure 2 GC-MS ion chromatograms of patchouli essential oil extracted from Comparative Examples 1-4 and Example 1; Figure 3 The scavenging ability of patchouli essential oil extracted from Comparative Examples 1-4 and Example 1 against DPPH· was compared. Figure 4 The patchouli essential oil extracted from Comparative Examples 1-4 and Example 1 was used to compare the effects of patchouli essential oil on ABTS. + • Clearance ability. Detailed Implementation

[0025] The following provides a detailed description of specific embodiments of the present invention. It should be understood that the specific embodiments described herein are for illustrative and explanatory purposes only and are not intended to limit the scope of the invention.

[0026] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0027] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0028] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be readily apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0029] In a first aspect, the present invention provides a method for extracting patchouli essential oil using an ultrasound-microwave synergistic-assisted eutectic solvent, the method comprising the following steps: 1) Mix choline chloride with a hydrogen bond donor, add water, and mix to prepare a eutectic solvent; 2) Mix the patchouli raw material with the eutectic solvent described in step 1) and perform ultrasonic-microwave synergistic assisted extraction to obtain an extraction mixture; 3) The extracted mixture described in step 2) is subjected to steam distillation, the distillate is collected, and patchouli essential oil is obtained by separation; The hydrogen bond donor is a mixture of organic acids and polyols.

[0030] This invention employs a eutectic solvent (DES) with choline chloride as the hydrogen bond acceptor and a mixture of organic acids and polyols as hydrogen bond donors as the hydrogen bond donors, as the extraction agent for patchouli essential oil. The solvent viscosity, polarity, and hydrogen bond network are controlled through the selection and combination of hydrogen bond donors. The active functional groups such as carboxyl and hydroxyl groups in the hydrogen bond donors can form strong hydrogen bonds with sesquiterpene alcohols, effectively reducing system viscosity and mass transfer resistance. The combination of specific hydrogen bond donors produces a synergistic effect, making the DES significantly more selective for extracting characteristic sesquiterpenes from patchouli essential oil than a single-component system. Furthermore, this solvent is non-toxic and biodegradable, and when combined with rotary evaporation concentration technology, it can be reused multiple times while maintaining good performance, significantly reducing processing material costs and solving the problems of organic solvent residue, poor selectivity of single DES extraction, and difficulty in recycling in traditional patchouli essential oil extraction processes.

[0031] In step 1) of this invention, to ensure that the DES has moderate permeability, it can efficiently dissolve intracellular essential oils and induce patchouli alcohol to dehydrate and generate β-patchoulene through a weakly acidic hydrogen bond environment, thereby directionally enriching olefinic active ingredients; the hydrogen bond donor is a compound mixture of malic acid and glycerol. The molar ratio of malic acid to glycerol in the compound mixture is 1:1~2.

[0032] In step 1) of this invention, the molar ratio of choline chloride to hydrogen bond donor is 1:1~2. Within the range defined by this invention, DES is always a clear liquid at room temperature with no solid precipitation; the yield of patchouli essential oil remains at a high level. Outside the range, stratification and a significant decrease in extraction yield will occur.

[0033] In step 1) of this invention, the water content in the eutectic solvent is 30-50 wt%, the DES system has suitable viscosity and high mass transfer efficiency; it can stably realize the conversion of patchouli alcohol to β-patchoulene, and significantly increase the sesquiterpene content; at the same time, water can regulate the solvent polarity and simultaneously dissolve polar patchouli alcohol and weakly polar sesquiterpene.

[0034] In step 1) of this invention, the mixing conditions include: a temperature of 75~85℃, a stirring rate of 200~400rpm, and a time of 60~120min. Under these conditions, a uniform, transparent, eutectic solvent can be rapidly prepared without undissolved solids. The physicochemical properties of the solvent are stable between batches, ensuring the repeatability of extraction.

[0035] In step 2) of this invention, the mixing ratio of material to liquid is 1:8~12g / mL, which ensures that the patchouli raw material is fully soaked and the intracellular essential oil is fully released; it also balances high essential oil yield with industrial production cost and moderate solvent recovery load.

[0036] In step 2) of this invention, the conditions for ultrasound-microwave synergistic extraction include: ultrasound power of 200-400W and ultrasound time of 15-25min; under these conditions, the cell walls of the patchouli raw material are fully broken down, the essential oil dissolution channels are fully opened, and the extraction time is significantly shortened; high-temperature degradation is avoided, and the complete active components are preserved. Microwave power of 250-350W and microwave time of 1-5min, the internal heating of microwaves and ultrasonic cavitation create a synergistic effect, compensating for the shortcomings of a single physical field; the dual-field coupling further improves the essential oil yield and enhances the patchouli alcohol conversion efficiency.

[0037] In step 3) of this invention, in order to completely separate and extract the essential oil components in the mixture and reduce the impurities in the crude essential oil after separation, the steam distillation time is 1-3 hours.

[0038] The method of the present invention further includes: collecting the residual eutectic solvent aqueous solution after steam distillation, filtering to remove solid residue, concentrating to remove water until the mass fraction of water in the residual eutectic solvent is 30~50wt%, and recovering the eutectic solvent for recycling. The eutectic solvent can be extracted multiple times with minimal degradation in extraction performance, significantly reducing the use of consumables, lowering production costs, and making the process green and suitable for industrial continuous production.

[0039] The various parameter ranges of this invention work together to achieve efficient dissolution of essential oils and directional enrichment of sesquiterpenes through an adjustable eutectic solvent system, resulting in highly antioxidant olefinic patchouli essential oil. On the other hand, the synergistic effect of ultrasonic and microwave dual fields significantly shortens the extraction cycle and improves the utilization rate of raw materials. The accompanying solvent recovery process reduces production costs and overcomes multiple defects of traditional steam distillation, such as long extraction time, low yield, poor essential oil activity, and environmentally unfriendly organic solvents.

[0040] Secondly, the present invention provides a patchouli essential oil prepared according to the method described in the present invention, wherein the normalized relative content of the gas chromatographic peak area of ​​the sesquiterpene components in the patchouli essential oil is ≥94%, the normalized relative content of the gas chromatographic peak area of ​​β-patchoulene is ≥41%, and the normalized relative content of the gas chromatographic peak area of ​​the alcohol components is ≤3.5%.

[0041] Thirdly, the present invention provides the application of patchouli essential oil as described herein in the preparation of antioxidants, cosmetics or food preservatives.

[0042] The present invention will be described in detail below through examples, but the scope of protection of the present invention is not limited thereto. In the following examples, the drugs and pharmaceuticals are all conventional commercially available products.

[0043] Example 1 (1) Preparation of eutectic solvent (DES): Using choline chloride as the hydrogen bond acceptor (HBA), malic acid and glycerol were mixed at a molar ratio of 1:1 as hydrogen bond donors (HBD). HBA and HBD were mixed at a molar ratio of 1:2 in a round-bottom flask, and deionized water was added to make the water content of the mixture 40 wt%. The mixture was placed in an 80°C constant temperature water bath and magnetically stirred at a rate of 300 r / min for 90 min to form a homogeneous, clear, stable, and transparent liquid, which is DES.

[0044] (2) Extraction of patchouli essential oil: Fresh patchouli stems and leaves were collected, and diseased or damaged parts were removed. The leaves were rinsed three times with deionized water to remove surface dust. After draining, they were dried in a 50℃ forced-air drying oven until constant weight (moisture content ≤8%). The dried stems and leaves were separated using a stem-leaf separation device, prioritizing the leaves with higher essential oil content. The patchouli leaves were then pulverized using a high-speed pulverizer and passed through a 100-mesh standard sieve to obtain patchouli leaf powder. Weigh the patchouli leaf powder at a material-liquid ratio of 1:10 g / mL and add the DES prepared in step (1). Stir thoroughly to ensure that the patchouli leaf powder is fully soaked. Set the ultrasonic power to 300W, the ultrasonic time to 20min, the microwave power to 300W, and the microwave time to 3min for ultrasonic-microwave synergistic extraction. (3) Transfer all the mixture extracted in step (2) to a steam distillation apparatus, heat to boiling, pass steam for distillation, collect the oil-water mixture after condensation, and fix the distillation time to 2 hours; transfer the oil-water mixture to a separatory funnel, let it stand for 30 minutes until it is completely separated, separate and collect the upper oil phase, which is crude refined oil. Anhydrous sodium sulfate was added to the obtained crude essential oil and dried overnight (8 h). After filtration, patchouli essential oil was obtained and denoted as UMSD-1. The yield was calculated by weighing.

[0045] Example 2 The procedure was carried out in accordance with Example 1, except that in step (2), “setting the ultrasonic power to 300W, the ultrasonic time to 20min, the microwave power to 300W, and the microwave time to 3min” was replaced with “setting the ultrasonic power to 200W, the ultrasonic time to 15min, the microwave power to 300W, and the microwave time to 1min”, with other conditions remaining unchanged. Patchouli essential oil was obtained and denoted as UMSD-2. The yield was calculated by weighing.

[0046] Example 3 The procedure was carried out in accordance with Example 1, except that in step (2), “setting the ultrasonic power to 300W, the ultrasonic time to 20min, the microwave power to 300W, and the microwave time to 3min” was replaced with “setting the ultrasonic power to 500W, the ultrasonic time to 30min, the microwave power to 300W, and the microwave time to 5min”, with other conditions remaining unchanged. Patchouli essential oil was obtained and denoted as UMSD-3. The yield was calculated by weighing.

[0047] Example 4 The process was carried out in accordance with Example 1, except that in step (1), "adding deionized water to make the water content of the mixed system 40wt%" was replaced with "adding deionized water to make the water content of the mixed system 30wt%", while other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as UMSD-4. The yield was calculated by weighing.

[0048] Example 5 The process was carried out in accordance with Example 1, except that in step (1), "adding deionized water to make the water content of the mixed system 40wt%" was replaced with "adding deionized water to make the water content of the mixed system 50wt%", while other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as UMSD-5. The yield was calculated by weighing.

[0049] Comparative Example 1 Take an equal amount of patchouli leaf powder as in Example 1, add deionized water at a material-to-liquid ratio of 1:10 g / mL, place it in a steam distillation apparatus, heat to boiling, pass steam through for distillation, collect the oil-water mixture after condensation, and fix the distillation time at 10 h; transfer the oil-water mixture to a separatory funnel, let it stand for 30 min until it is completely separated, separate and collect the upper oil phase, which is the crude essential oil; Anhydrous sodium sulfate was added to the obtained crude essential oil and dried overnight. After filtration, patchouli essential oil was obtained and denoted as SD. The yield was calculated by weighing.

[0050] Comparative Example 2 Take an equal amount of patchouli leaf powder as in Example 1, add it to the DES prepared in step (1) of Example 1 at a material-to-liquid ratio of 1:10 g / mL, stir thoroughly to fully wet the patchouli leaf powder, place it in a steam distillation apparatus, heat to boiling, pass steam through for distillation, collect the oil-water mixture after condensation, and fix the distillation time to 10 h; transfer the oil-water mixture to a separatory funnel, let it stand for 30 min until it is completely separated, separate and collect the upper oil phase, which is the crude essential oil; Anhydrous sodium sulfate was added to the obtained crude essential oil and dried overnight. After filtration, patchouli essential oil was obtained and denoted as DSD. The yield was calculated by weighing.

[0051] Comparative Example 3 The procedure was carried out in accordance with Example 1, except that in step (2), “set the ultrasonic power to 300W, the ultrasonic time to 20min, the microwave power to 300W, and the microwave time to 3min” was replaced with “set the ultrasonic power to 300W and the ultrasonic time to 20min”, while keeping other conditions unchanged. Patchouli essential oil was obtained and denoted as USD. The yield was calculated by weighing.

[0052] Comparative Example 4 The procedure was carried out in accordance with Example 1, except that in step (2), “set the ultrasonic power to 300W, the ultrasonic time to 20min, the microwave power to 300W, and the microwave time to 3min” was replaced with “set the microwave power to 300W and the microwave time to 3min”, while keeping other conditions unchanged. Patchouli essential oil was obtained and denoted as MSD. The yield was calculated by weighing.

[0053] Comparative Example 5 The process was carried out in accordance with Example 1, except that in step (1), "adding deionized water to make the water content of the mixed system 40wt%" was replaced with "adding deionized water to make the water content of the mixed system 10wt%", while other conditions remained unchanged. Patchouli essential oil was obtained and denoted as UMSD-D5. The yield was calculated by weighing.

[0054] Comparative Example 6 The process was carried out in accordance with Example 1, except that in step (1), "adding deionized water to make the water content of the mixed system 40wt%" was replaced with "adding deionized water to make the water content of the mixed system 70wt%", while other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as UMSD-D6. The yield was calculated by weighing.

[0055] Comparative Example 7 The procedure was carried out in accordance with Example 1, except that in step (2), “setting the ultrasonic power to 300W, the ultrasonic time to 20min, the microwave power to 300W, and the microwave time to 3min” was replaced with “setting the ultrasonic power to 100W, the ultrasonic time to 10min, the microwave power to 300W, and the microwave time to 3min”, while keeping other conditions unchanged. Patchouli essential oil was obtained and recorded as UMSD-D7. The yield was calculated by weighing.

[0056] Comparative Example 8 The process was carried out in accordance with Example 1, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as hydrogen bond donor (HBD)" was replaced with "using lactic acid as HBD". Other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as UMSD-D8. The yield was calculated by weighing.

[0057] Comparative Example 9 The procedure was carried out in accordance with Comparative Example 2, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as hydrogen bond donor (HBD)" was replaced with "using malic acid as HBD". Other conditions remained unchanged, and patchouli essential oil was obtained, denoted as DSD-2. The yield was calculated by weighing.

[0058] Comparative Example 10 The procedure was carried out in accordance with Comparative Example 2, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as hydrogen bond donors (HBD)" was replaced with "using malic acid as HBD", and "mixing HBA and HBD in a molar ratio of 1:2 in a round-bottom flask" was replaced with "mixing HBA and HBD in a molar ratio of 1:1 in a round-bottom flask". Other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as DSD-3. The yield was calculated by weighing.

[0059] Comparative Example 11 The procedure was carried out in accordance with Comparative Example 2, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as hydrogen bond donor (HBD)" was replaced with "using glycerol as HBD". Other conditions remained unchanged, and patchouli essential oil was obtained, denoted as DSD-4. The yield was calculated by weighing.

[0060] Comparative Example 12 The procedure was carried out in accordance with Comparative Example 2, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as a hydrogen bond donor (HBD)" was replaced with "using glycerol as HBD", and "mixing HBA and HBD in a molar ratio of 1:2 in a round-bottom flask" was replaced with "mixing HBA and HBD in a molar ratio of 1:1 in a round-bottom flask". Other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as DSD-5. The yield was calculated by weighing.

[0061] Comparative Example 13 The procedure was carried out in the same manner as Comparative Example 2, except that in step (1), "mixing malic acid and glycerol at a molar ratio of 1:1 as hydrogen bond donors (HBD)" was replaced with "using lactic acid as HBD". Other conditions remained unchanged, and patchouli essential oil was obtained, denoted as DSD-6. The yield was calculated by weighing.

[0062] Comparative Example 14 The procedure was carried out in accordance with Comparative Example 2, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as hydrogen bond donors (HBD)" was replaced with "using lactic acid as HBD", and "mixing HBA and HBD in a molar ratio of 1:2 in a round-bottom flask" was replaced with "mixing HBA and HBD in a molar ratio of 1:1 in a round-bottom flask". Other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as DSD-7. The yield was calculated by weighing.

[0063] Comparative Example 15 The procedure was carried out in accordance with Comparative Example 2, except that in step (1), "mixing malic acid and glycerol in a molar ratio of 1:1 as hydrogen bond donor (HBD)" was replaced with "mixing citric acid and lactic acid in a molar ratio of 1:1 as HBD". Other conditions remained unchanged, and patchouli essential oil was obtained, which was denoted as DSD-8. The yield was calculated by weighing.

[0064] Detection Example 1 The surface morphology of patchouli leaf powder before and after treatment with the extraction methods of Example 1 and Comparative Examples 1-2 was observed using scanning electron microscopy (SEM). The sample preparation method was as follows: The patchouli leaf powder residue after treatment with the extraction methods of Example 1 and Comparative Examples 1-2 was collected by filtration, washed twice with deionized water to remove residual eutectic solvents and soluble impurities, and then dried to constant weight in a vacuum oven at 40℃. The dried samples were then sputter-coated with gold and observed under SEM, and images were acquired. The results are shown below. Figure 1 As shown.

[0065] Depend on Figure 1 The content indicates that unprocessed patchouli leaf powder ( Figure 1 A) It retains the complete natural microstructure after drying: the rhizome structure is clear, with a large number of root-like fibers attached to the surface, arranged distinctly without obvious signs of damage, exhibiting a dense natural fiber network. The patchouli leaf powder residue after processing using the extraction method of Comparative Example 1 ( Figure 1B) Exhibits mild destructive characteristics: root-like fibers detach and aggregate, but are not completely dissolved, and the basic morphology of the fibers is still preserved, indicating that the method has a weak destructive effect on the tissue structure. The patchouli leaf powder residue after treatment by the extraction method of Comparative Example 2 ( Figure 1 C) Significant structural damage was observed: some root-like fibers dissolved, but some slender fiber structures remained; the residue surface was rough and uneven, filled with cracks and pores, indicating that DES extraction could damage the tissue structure, but the degree of damage was limited and the fiber network was not completely disintegrated. The patchouli leaf powder residue after treatment by the extraction method of Example 1 ( Figure 1 D) exhibits the most thorough destructive characteristics: the root-like fibers completely disappear, the original fiber structure disintegrates into dense clumps, and a relatively smooth and flat area is formed on the surface, with no natural fiber morphology residue, indicating that this method has the strongest destructive ability on the tissue structure of patchouli leaves.

[0066] The above results indicate that different extraction methods exhibit significant differences in their ability to damage the tissue structure of patchouli leaves: the traditional steam distillation method in Comparative Example 1 only caused mild damage; the extraction method in Comparative Example 2, which combined the use of DES from Example 1 of this invention with traditional steam distillation, effectively damaged the tissue structure but retained some of it; while the ultrasonic-microwave synergistic assisted eutectic solvent method in Example 1 of this invention achieved the most thorough damage. Stronger structural damage facilitates solvent penetration and dissolution of active ingredients, which explains the advantage of the ultrasonic-microwave synergistic assisted eutectic solvent method in extraction efficiency in Example 1 of this invention.

[0067] Detection Example 2 The extraction rate and yield of patchouli essential oil were calculated using the extraction methods of Examples 1-5 and Comparative Examples 1-15, and the results are shown in Table 1.

[0068] Table 1 As shown in Table 1, the introduction of eutectic solvent (DES) can improve the extraction efficiency of patchouli essential oil. The yield of the conventional steam distillation (SD) method in Comparative Example 1 was 1.14%, while the yields of the comparative examples using the DES system of this invention, without the application of a physical field (Comparative Example 2 and Comparative Examples 9-15), were all higher than that of Comparative Example 1. For example, the yield of Comparative Example 2 (DSD) was 2.93%, and that of Comparative Example 9 (DSD-2) was 2.75%, indicating that DES as an extraction medium can effectively promote the dissolution of essential oil from the raw material. However, the extraction cycle of Comparative Example 2 and Comparative Examples 9-15 was 10 hours, comparable to that of Comparative Example 1, indicating that simply replacing the extraction medium without introducing a physical field cannot solve the inherent drawback of the long extraction time of traditional processes.

[0069] The moisture content of DES has a significant impact on the extraction efficiency. The yields of Example 1 (40% moisture content), Example 4 (30% moisture content), and Example 5 (50% moisture content) were 3.54%, 3.05%, and 3.20%, respectively, while the yields of Comparative Example 5 (10% moisture content) and Comparative Example 6 (70% moisture content) were 1.50% and 1.80%, respectively. The yield gradually increased as the moisture content increased from 10% to 40%; however, when the moisture content continued to increase to 70%, the yield showed a decreasing trend. These results indicate that the moisture content of DES needs to be controlled within an appropriate range. When the moisture content is too low (10%), the viscosity of the DES system is too high, mass transfer is hindered, and the essential oil is difficult to fully dissolve from the raw material. When the moisture content is too high (70%), the DES system is excessively diluted, and the hydrogen bonding ability is weakened, which is also detrimental to the efficient extraction of essential oil.

[0070] The synergistic effect of ultrasound and microwave is superior to that of a single physical field. The yield of Comparative Example 3 (single ultrasound assistance, USD) was 3.14%, the yield of Comparative Example 4 (single microwave assistance, MSD) was 3.23%, while the yield of Example 1 (ultrasound-microwave synergistic assistance, UMSD) was 3.54%, higher than both single physical field assistance methods. Furthermore, the extraction cycle of Example 1 was 3.5 h, Comparative Example 3 was 5 h, Comparative Example 4 was 4.5 h, and Comparative Examples 1 and 2 were both 10 h. These results indicate that the ultrasonic cavitation effect and microwave dielectric heating can complement each other within the same extraction system. Ultrasound disrupts cell walls, while microwaves promote uniform heating and accelerate mass transfer. The combination of these two methods can achieve a higher essential oil yield in a shorter time.

[0071] The selection of physical field parameters has a direct impact on the extraction effect. In Comparative Example 7, when the ultrasonic power was reduced to 100W and the ultrasonic time was shortened to 10min, the yield was 3.10%, lower than the 3.54% of Example 1. The above results indicate that excessively low ultrasonic power and excessively short ultrasonic time are unlikely to produce an effective cavitation effect, while excessively long microwave time may lead to excessively high system temperature, loss of low-boiling-point components due to volatilization, or increased emulsification of the system, all of which are detrimental to improving extraction efficiency. Therefore, it can be determined that the ultrasonic power of 200~400W, ultrasonic time of 15~25min, and microwave time of 1~5min specified in step (2) of this invention have experimental basis.

[0072] The composition of the DES hydrogen bond donor has a decisive influence on the extraction efficiency. The yield of Example 1 (malic acid and glycerol compound) was 3.54%, which was higher than that of Comparative Example 9 (malic acid alone, yield 2.75%), Comparative Example 10 (malic acid alone, HBA:HBD=1:1, yield 2.84%), Comparative Example 11 (glycerol alone, yield 2.41%), Comparative Example 12 (glycerol alone, HBA:HBD=1:1, yield 2.60%), Comparative Example 13 (lactic acid alone, yield 2.59%), and Comparative Example 14 (lactic acid alone, HBA:HBD=1:1, yield 2.64%) without physical field assistance. The carboxyl groups in malic acid can form hydrogen bonds with sesquiterpene alcohols, while the polyol structure of glycerol can regulate the system viscosity and enhance the penetration of DES into plant fibers. The combination of these two substances works simultaneously on both the solubility and mass transfer efficiency levels, resulting in extraction efficiency superior to any single hydrogen bond donor system. Comparative Example 15 (a combination of citric acid and lactic acid) had a yield of 2.57%, showing no significant advantage compared to the single system, indicating that not all organic acid combinations produce a synergistic effect; only combinations of specific hydrogen bond donors can achieve a substantial improvement in extraction efficiency.

[0073] The method of this invention significantly shortens the extraction time while improving the yield. The yield of Example 1 was 3.54%, 3.1 times that of the conventional steam distillation method (1.14%) in Comparative Example 1, and the extraction cycle was shortened from 10 hours to 3.5 hours, saving approximately 65% ​​of the time. Compared with Comparative Example 2 (DES-assisted steam distillation, yield 2.93%, time 10 hours), Example 1 showed a 0.61% increase in yield and a 6.5-hour reduction in extraction time; compared with Comparative Example 3 (USD, yield 3.14%, time 5 hours), the yield increased by 0.4% and the time decreased by 1.5 hours; compared with Comparative Example 4 (MSD, yield 3.23%, time 4.5 hours), the yield increased by 0.31% and the time decreased by 1.0 hour. These results indicate that the synergistic effect of DES, ultrasound, and microwave improves both the yield and time compared to the methods in the comparative examples.

[0074] In summary, the data in Table 1 systematically verified the influence of key process parameters of this invention on the extraction efficiency of patchouli essential oil: the introduction of DES can improve the yield; the water content needs to be controlled between 30 and 50 wt%; the synergistic effect of ultrasound and microwave dual-field is better than that of a single physical field; and each physical field parameter and DES composition have reasonable value ranges. The above data provide a sample basis for subsequent GC-MS component analysis and antioxidant activity evaluation of the essential oil.

[0075] Detection Example 3 The patchouli essential oils prepared in Example 1 and Comparative Examples 1-4 were subjected to qualitative and semi-quantitative analysis by GC-MS.

[0076] Chromatographic conditions: HP-5MS quartz capillary column (30m×0.25mm, 0.25μm); injection port temperature 260℃; carrier gas was high-purity helium (99.999%), flow rate 1.0mL / min; split ratio 5:1.

[0077] Temperature program: Initial temperature 60℃, hold for 2 min, increase to 160℃ at 10℃ / min, hold for 3 min, then increase to 250℃ at 5℃ / min, hold for 2 min.

[0078] Mass spectrometry conditions: Electron impact (EI) ion source, electron energy 70 eV; ion source temperature 260℃; transfer line temperature 280℃; mass scan range m / z 45-500.

[0079] 1 μL of patchouli essential oil samples (diluted 100-fold with n-hexane) prepared in Example 1 and Comparative Examples 1-4 were injected for analysis. Compound identification was performed by comparing with the NIST17 standard mass spectrometry library and referring to retention indices from relevant literature. The relative content of each component was calculated using the peak area normalization method.

[0080] The results are as follows Figure 2 As shown in Table 2.

[0081] Depend on Figure 2 As can be seen from the contents and the data in Table 2, the extraction method has a significant impact on the chemical composition of essential oils. Among them, the ultrasound-microwave synergistic assisted eutectic solvent method of this invention shows a significant selective enrichment effect on specific sesquiterpenes. Particularly noteworthy is β-patcholenene, whose relative content in patchouli essential oil extracted by this ultrasound-microwave synergistic assisted eutectic solvent method is as high as 42.04%, significantly higher than the relative content in patchouli essential oil extracted by Comparative Examples 1-4. Simultaneously, the content of α-coumarene in patchouli essential oil extracted by this ultrasound-microwave synergistic assisted eutectic solvent method is also much higher than the relative content in patchouli essential oil extracted by Comparative Examples 1-2. This indicates that the ultrasound-microwave synergistic assisted eutectic solvent method of this invention may be more favorable for the dissolution and stable retention of such sesquiterpenes with specific skeletons.

[0082] It is noteworthy that in the patchouli essential oils obtained by the three different physical field-assisted DES extraction methods in Example 1, Comparative Example 3, and Comparative Example 4, no direct detection or only trace amounts of berberine were detected. In contrast, in the patchouli essential oils obtained by the extraction methods in Comparative Examples 1-2 without enhanced physical field application, this component was clearly detected, with relative contents of 42.63% and 31.82%, respectively. Considering the significantly increased β-patchoulene content in the patchouli essential oil obtained in Example 1, it is speculated that under the synergistic effect of ultrasound and microwave, local thermal effects and mechanical disturbances may occur within the extraction system, promoting the dehydration or structural rearrangement of some patchouli alcohols, transforming them into the corresponding olefinic product—patchoulene. This type of transformation is relatively common in the extraction of natural products, especially when accompanied by thermal effects or in a weakly acidic DES environment. The significant increase in the content of β-patchoulene, one of the characteristic sesquiterpene components of patchouli essential oil, further confirms the efficient extraction capability of the ultrasonic-microwave synergistic eutectic solvent method of this invention for characteristic components of patchouli, even if their form may have changed.

[0083] Furthermore, sesquiterpenes such as α-guaiacolene, seychellesene, and α-patchoulene were detected in high amounts in all five extraction methods, constituting the basic aroma components of patchouli essential oil. In the ultrasonic-microwave synergistic eutectic solvent method of this invention, the relative contents of these components are similar to those in other physical field-assisted methods. For example, the relative content of α-patchoulene in Example 1 is 12.61%, in Comparative Example 3 it is 12.42%, and in Comparative Example 4 it is 12.45%, but all are lower than the relative content of α-patchoulene in Comparative Example 1 (13.66%) and Comparative Example 2 (18.97%). This phenomenon is due to the fact that the physical field assistance significantly accelerates the dissolution kinetics of these components, resulting in a relative decrease in their relative proportion in the total volatiles due to the significant increase of other components (such as β-patchoulene). However, their absolute extraction yield may still increase due to the improved extraction efficiency.

[0084] Detection Example 4 The antioxidant capacity of patchouli essential oil is one of its important bioactivities. To evaluate the effect of the extraction method on this activity, the antioxidant capacity of patchouli essential oils extracted in Example 1 and Comparative Examples 1-4 against DPPH· free radicals and ABTS was determined. + • Half-maximum scavenging concentration (IC50) of free radicals 50 ), the result is as follows Figures 3-4 As shown in Table 3.

[0085] Table 3 IC 50The lower the value, the stronger the antioxidant capacity of the essential oil. Table 3 shows that the extraction process significantly affects the antioxidant activity of the essential oil.

[0086] Regarding the scavenging ability of DPPH free radicals, the IC50 of the essential oil obtained by conventional steam distillation (SD) in Comparative Example 1 was... 50 The value was 12.80 g / L, while Comparative Example 2, based on DES-based steam distillation (DSD), reduced it to 7.78 g / L. When physical field assistance was introduced, the antioxidant activity was further enhanced, with Comparative Example 3 showing ultrasound-assisted DES (USD) and Comparative Example 4 showing microwave-assisted DES (MSD) IC values ​​of [missing information]. 50 The values ​​were 5.88 g / L and 6.54 g / L, respectively, while the ultrasound-microwave assisted DES method (UMSD) of Example 1 of this invention exhibited the strongest removal ability, IC 50 The lowest value was 3.76 g / L.

[0087] ABTS + • Free radical scavenging experiments show similar trends, with different methods having different IC50 values. 50 The values ​​were as follows: Comparative Example 1 (4.97 g / L) > Comparative Example 2 (3.66 g / L) > Comparative Example 4 (2.50 g / L) > Comparative Example 3 (1.15 g / L) > Example 1 (0.77 g / L). This indicates that with the introduction and synergy of physical energy (microwave, ultrasound) during the extraction process, the in vitro antioxidant activity of the obtained patchouli essential oil showed a stepwise increase. The patchouli essential oil extracted by the ultrasound-microwave synergistic assisted DES method in Example 1 of this invention has the best free radical scavenging ability.

[0088] This trend in antioxidant activity is likely closely related to changes in the composition and content of essential oil chemical components. As shown in Example 3, the essential oils extracted by the SD method in Comparative Example 1 and the DSD method in Comparative Example 2 were rich in alcohols such as patchouli alcohol, while the extraction methods in Comparative Examples 3-4 and Example 1 transformed the essential oils into chemically characterized primarily by sesquiterpenes such as β-patchoulene. Although patchouli alcohol is traditionally considered the key active ingredient, the data in this study indicate that the "transformed" essential oils rich in sesquiterpenes exhibit stronger overall antioxidant potential. This suggests that the antioxidant activity of essential oils is not determined by a single component, but rather by the complex interaction of all their volatile and non-volatile components.

[0089] In the ultrasound-microwave synergistic eutectic solvent method of this invention, the synergistic effect of ultrasound and microwave not only efficiently disrupts cell walls for extraction but may also promote the dissolution of more types of antioxidant precursors in the plant matrix, leading to their interaction in the DES medium. Furthermore, the thermal effect of microwaves may induce Maillard reactions, generating new products with antioxidant activity. Simultaneously, the conversion of the main components from alcohols to alkenes may alter the electron-donating capacity of each component of the essential oil, thus affecting its efficiency in reacting with free radicals. Synergistic effects may occur between sesquiterpenes of different structural types and their possible trace oxygen-containing derivatives, thereby enhancing the overall antioxidant performance. Therefore, the ultrasound-microwave synergistic eutectic solvent method of this invention, by altering the extraction mechanism and inducing component conversion, ultimately yields an essential oil product with a significantly different chemical composition and stronger free radical scavenging ability. This provides a new technological perspective for the further development of highly efficient antioxidant plant essential oil resources.

[0090] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

[0091] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not describe the various possible combinations separately.

[0092] Furthermore, various different embodiments of the present invention can be combined in any way, as long as they do not violate the spirit of the present invention, they should also be regarded as the content disclosed by the present invention.

Claims

1. A method for extracting patchouli essential oil using an ultrasound-microwave synergistic-assisted eutectic solvent, characterized in that, The method includes the following steps: 1) Mix choline chloride with a hydrogen bond donor, add water, and mix to prepare a eutectic solvent; 2) Mix the patchouli raw material with the eutectic solvent described in step 1) and perform ultrasonic-microwave synergistic assisted extraction to obtain an extraction mixture; 3) The extracted mixture described in step 2) is subjected to steam distillation, the distillate is collected, and patchouli essential oil is obtained by separation; The hydrogen bond donor is a mixture of organic acids and polyols.

2. The method according to claim 1, characterized in that, In step 1), the hydrogen bond donor is a complex mixture of malic acid and glycerol; Preferably, the molar ratio of malic acid to glycerol in the compound mixture of malic acid and glycerol is 1:1 to 2.

3. The method according to claim 1 or 2, characterized in that, In step 1), the molar ratio of choline chloride to the hydrogen bond donor is 1:1~2; The mass fraction of water in the eutectic solvent is 30~50wt%.

4. The method according to any one of claims 1-3, characterized in that, In step 1), the mixing conditions include: a temperature of 75~85℃, a stirring rate of 200~400rpm, and a time of 60~120min.

5. The method according to any one of claims 1-4, characterized in that, In step 2), the ratio of the mixed material to liquid is 1:8~12 g / mL.

6. The method according to any one of claims 1-5, characterized in that, In step 2), the conditions for ultrasound-microwave synergistic assisted extraction include: ultrasound power 200~400W, ultrasound time 15~25min, microwave power 250~350W, and microwave time 1~5min.

7. The method according to any one of claims 1-6, characterized in that, In step 3), the steam distillation time is 1 to 3 hours.

8. The method according to any one of claims 1-7, characterized in that, The method further includes: collecting the residual eutectic solvent aqueous solution after steam distillation, filtering to remove solid residue, concentrating to remove water until the mass fraction of water in the residual eutectic solvent is 30~50wt%, and recovering the eutectic solvent for recycling.

9. A patchouli essential oil prepared according to any one of claims 1-8, characterized in that, The normalized relative content of sesquiterpenes in the patchouli essential oil was ≥94%, the normalized relative content of β-patchoulene was ≥41%, and the normalized relative content of alcohols was ≤3.5%.

10. The use of the patchouli essential oil as described in claim 9 in the preparation of antioxidants, cosmetics, or food preservatives.

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