WO2020135896A1 - 一种具有抗菌性能精油气味去除方法 - Google Patents

一种具有抗菌性能精油气味去除方法 Download PDF

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WO2020135896A1
WO2020135896A1 PCT/CN2020/077362 CN2020077362W WO2020135896A1 WO 2020135896 A1 WO2020135896 A1 WO 2020135896A1 CN 2020077362 W CN2020077362 W CN 2020077362W WO 2020135896 A1 WO2020135896 A1 WO 2020135896A1
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essential oil
yeast
oil
antibacterial properties
fermentation
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PCT/CN2020/077362
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English (en)
French (fr)
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巫世贵
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珠海市自然之旅生物技术有限公司
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L5/00—Preparation or treatment of foods or foodstuffs, in general; Food or foodstuffs obtained thereby; Materials therefor
    • A23L5/20—Removal of unwanted matter, e.g. deodorisation or detoxification
    • A23L5/28—Removal of unwanted matter, e.g. deodorisation or detoxification using microorganisms
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
    • A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
    • A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
    • A23L3/3463—Organic compounds; Microorganisms; Enzymes
    • A23L3/3472—Compounds of undetermined constitution obtained from animals or plants

Definitions

  • the invention relates to a method for effectively removing the flavor of essential oil.
  • Preservatives are more and more popular among consumers; on the other hand, the main reason for limiting the development of plant-based preservatives is that plant extracts will contain some special odors. If added to foods, some foods with relatively light taste In other words, it will greatly destroy the taste of food itself, and even make consumers feel unpleasant. In view of these circumstances, the practice is generally to suppress the odor of these plant extracts by adding fragrance, but after being added to food, the strong smell of plant extracts will still appear during the tasting process, which also greatly limits plant extracts. As a major obstacle to the application of preservatives in the food field. In view of this situation, at present, there is no way to solve this drawback.
  • the purpose of the present invention is to solve the problem that the prior art cannot effectively remove the taste of essential oils, and to provide a method for removing the smell of essential oils with antibacterial properties.
  • a method for removing odor of essential oil with antibacterial properties specifically includes the following steps: processing of essential oil with antibacterial properties using yeast fermentation broth.
  • the viable bacterial count of the yeast in the yeast fermentation broth is 10 7 cfu/mL to 10 9 cfu/mL.
  • the yeast in the yeast fermentation broth is one or more of Saccharomyces cerevisiae, Saccharomyces cerevisiae, Saccharomyces cerevisiae, 18 degree wine yeast, Angel yeast and white wine special yeast.
  • the essential oil with antibacterial properties is a single essential oil or a compound essential oil.
  • the single essential oil is cinnamon oil, clove essential oil, rosemary essential oil, garlic oil, broad scented oil, nutmeg oil, star anise oil, bergamot essential oil, Nanfeng tangerine essential oil, sphaeranthus essential oil or Shenghong thistle essential oil.
  • the compound essential oil is composed of two or more of cinnamon oil, clove essential oil, rosemary essential oil, broad holly oil, nutmeg oil and star anise oil.
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Fermentation add the essential oil with antibacterial properties to the yeast fermentation broth, mix and ferment it to obtain the essential oil fermentation broth; the volume ratio of the essential oil with antibacterial properties to the yeast fermentation broth is 6-10:1;
  • step (3) Merging: combining the essential oil I and the essential oil II obtained in step (3), that is, an essential oil with antibacterial properties after odor removal is obtained.
  • the edible yeast selected in the present invention effectively removes (or reduces) unpleasant odors in essential oils with antibacterial properties, does not produce additional harmful substances to essential oils with antibacterial properties, and at the same time can greatly retain essential oils with antibacterial properties Antibacterial activity.
  • the odor-removing essential oil obtained by the invention is used as an antiseptic agent.
  • This embodiment is a method for removing odor of essential oil with antibacterial properties, which specifically includes the following steps:
  • the yeast fermentation broth is used to treat the essential oil with antibacterial properties to successfully produce fermentation and effectively remove (or reduce) the Unpleasant odor in essential oils with antibacterial properties.
  • the embodiment selects the yeast fermentation broth to treat essential oils with antibacterial properties, which not only effectively removes (or reduces) unpleasant odors in essential oils with antibacterial properties, but also greatly retains the antibacterial activity of essential oils with antibacterial properties.
  • the difference between this embodiment and the first embodiment is that the viable bacterial count of yeast in the yeast fermentation broth is 10 7 cfu/mL to 10 9 cfu/mL. Others are the same as the first embodiment.
  • the yeast in the yeast fermentation broth is Saccharomyces pombe, Saccharomyces cerevisiae, Saccharomyces cerevisiae, 18 degree wine yeast, Angel yeast One or several of the yeasts for white wine. Others are the same as the specific embodiments one or two.
  • Saccharomyces pombe lipolytica described in this embodiment is produced by the Guangdong Institute of Microbiology, and the deposit number is ATCC18942.
  • the acid-reducing yeast, saccharomyces cerevisiae, 18-degree wine yeast and white wine special yeast described in this embodiment are produced by Shandong Yantai Dibos Yeast Co., Ltd.
  • Angel yeast described in this embodiment is produced by Angel Yeast Co., Ltd.
  • the essential oil with antibacterial properties is a single essential oil or a compound essential oil. Others are the same as the specific embodiments one to three.
  • the single essential oil is cinnamon oil, clove essential oil, rosemary essential oil, garlic oil, broad husk oil, nutmeg oil, star anise oil, Litsea Cubeba essential oil, Nan Rich tangerine essential oil, fluffy essential oil or Shenghong thistle essential oil. Others are the same as the fourth embodiment.
  • the compound essential oil is composed of two or more of cinnamon oil, clove essential oil, rosemary essential oil, broad holly oil, nutmeg oil and star anise oil. to make. Others are the same as the fourth embodiment.
  • Fermentation add the essential oil with antibacterial properties to the yeast fermentation broth, mix and ferment it to obtain the essential oil fermentation broth; the volume ratio of the essential oil with antibacterial properties to the yeast fermentation broth is 6-10:1;
  • step (3) Merging: combining the essential oil I and the essential oil II obtained in step (3), that is, an essential oil with antibacterial properties after odor removal is obtained.
  • yeast fermentation broth described in step (1) is obtained as follows:
  • bacteria activation put yeast into the activation liquid to activate, to obtain the yeast activation liquid containing the activated yeast; the mass fraction of white granulated sugar in the activation liquid is 5-15%;
  • step (2) the essential oil fermentation broth is inactivated at 0°C for 24 hours, then subjected to 250W to 300W ultrasonic treatment for 15min to 60min, and finally 12000 rpm Centrifuge for 10 to 25 minutes and collect to obtain supernatant and precipitate.
  • step (3) the essential oil fermentation broth is inactivated at 0°C for 24 hours, then subjected to 250W to 300W ultrasonic treatment for 15min to 60min, and finally 12000 rpm Centrifuge for 10 to 25 minutes and collect to obtain supernatant and precipitate.
  • step (3) the essential oil fermentation broth is inactivated at 0°C for 24 hours, then subjected to 250W to 300W ultrasonic treatment for 15min to 60min, and finally 12000 rpm Centrifuge for 10 to 25 minutes and collect to obtain supernatant and precipitate.
  • step (3) the supernatant obtained in step (2) is sequentially concentrated and concentrated until there is no water to obtain a supernatant concentrate, which is used
  • the organic solvent extracts the supernatant concentrate to obtain an organic solvent extract, and the organic solvent extract is concentrated again until all organic solvents are separated by rotary evaporation to obtain essential oil I;
  • the organic solvent is selected from ethyl acetate and ethanol , Ether and cyclohexane;
  • the volume ratio of the supernatant concentrate to the organic solvent is 1 to 3: 3 to 6;
  • the precipitate obtained in step (2) is extracted with a solvent to obtain a solvent extract, and the solvent extract is concentrated until all solvents are separated by rotary evaporation to obtain essential oil II; all solvents are selected from ethyl acetate, ethanol, ether and cyclo Hexane; the volume ratio of the mass of the precipitate to the solvent is 1 g to 4 g: 2 mL to 6 mL.
  • the water temperature of the rotary evaporator until all the organic solvents described in this embodiment are separated by rotary evaporation is 45 to 60° C., and the rotation speed is 45 r/min.
  • the water temperature of the rotary evaporator until all the solvents described in this embodiment are separated by rotary evaporation is 45 to 60° C., and the rotation speed is 45 r/min.
  • Bacterial species activation add the activation liquid to the pre-fermentation tank, and then add yeast for activation to obtain a yeast activation liquid containing the activated yeast; the mass fraction of white granulated sugar in the activation liquid is 5-15%;
  • Enrichment fermentation put the yeast activation solution and yeast medium containing the activated yeast into the main fermentation tank for shaker fermentation, the fermentation temperature is 24 ⁇ 35 °C, the shaker speed is 150r/min ⁇ 300r/min; Light conditions: dark 6h-8h, light 16h-18h, light intensity 2000lux-2500lux, until yeast live bacteria count is 10 7 cfu/mL ⁇ 10 9 cfu/mL, to obtain yeast fermentation broth;
  • Deodorization fermentation add the essential oil with antibacterial properties to the main fermentation tank equipped with yeast fermentation broth, after mixing, adjust the shaker speed to 150r/min ⁇ 300r/min, the temperature to 30 ⁇ 65°C, and then The shaker speed is 150r/min ⁇ 300r/min and the temperature is 30 ⁇ 65°C, the fermentation time is 72h ⁇ 120h, and the essential oil fermentation broth is obtained; the volume ratio of the antibacterial property essential oil and yeast fermentation broth is 6 ⁇ 10:1 ;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, then subjected to ultrasonic treatment at 250W to 300W for 15min to 60min, and finally centrifuged at 12000 rpm for 10min to 25min using a high-speed centrifuge, collected, and the supernatant is obtained Liquid and precipitation;
  • the water temperature of the rotary evaporator is 45-60°C and the rotation speed 45r/min ⁇ 65r/min to obtain essential oil II; all solvents are selected from ethyl acetate, ethanol, ether and cyclohexane; the volume ratio of the mass of the precipitate to the solvent is 1g ⁇ 4g: 2mL ⁇ 6mL;
  • step 6 Merging: The essential oil I and the essential oil II obtained in step 6 are combined, that is, an essential oil with antibacterial properties after odor removal is obtained.
  • the content of the present invention is not limited to the content of the above-mentioned embodiments, and the combination of one or several specific embodiments can also achieve the purpose of the invention.
  • a method for removing odor of essential oil with antibacterial properties specifically including the following steps: processing yeast essential oil with antibacterial properties; the essential oil with antibacterial properties is cinnamon oil;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 200g;
  • Bacterial species activation add 100g of the activation solution to the pre-fermentation tank, and then separately add S. cerevisiae, Saccharomyces cerevisiae and Angel yeast to activate, to obtain S. cerevisiae activation solution and S. cerevisiae activation Liquid and Angel yeast activation liquid; the mass fraction of white granulated sugar in the activation liquid is 5%;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast culture medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 30°C, shaker speed of 250r / min; light conditions: dark 7h, light 17h, light intensity 2300lux, to a total viable cell count of 10 8 cfu / far mL, resulting in the yeast fermentation broth; laminating said polyethylene lipolytica toruloides activation solution: The volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation add 300g of cinnamon oil to the main fermentation tank containing yeast fermentation broth in step 3, after mixing, adjust the shaker speed to 300r/min, the temperature to 35°C, and then shake the speed at Fermentation time at 300r/min and temperature of 35°C for 72h to obtain essential oil fermentation broth;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, and then subjected to 300W ultrasonic treatment for 25min, and finally centrifuged at 12000 rpm for 10min using a high-speed centrifuge and collected to obtain the supernatant and precipitate;
  • the water temperature of the rotary evaporator is 50°C and the rotation speed is 45r/ min to obtain essential oil II; all solvents are ethyl acetate; the volume ratio of the mass of the precipitate to the solvent is 1 g: 3 mL;
  • test bacteria were: Staphylococcus aureus, Escherichia coli, Candida albicans, Pseudomonas aeruginosa and Aspergillus niger.
  • the minimum inhibitory concentration MIC of cinnamon oil and the deodorized cinnamon oil obtained in Example 1 was determined by the liquid medium dilution method.
  • the specific method is as follows:
  • Example 2 Take 1g of the deodorized cinnamon oil obtained in Example 1 and dissolve it in 9mL of DMSO in advance to obtain a sample solution with a concentration of 1/10. Take a sterilized test tube, draw 1mL of the sample solution with a concentration of 1/10, and then add 9mL of nutrient meat Soup medium to obtain a sample solution with a concentration of 1/100. Pipette 0.5mL, 1mL and 2mL of sample solution with a concentration of 1/100 into 9.5mL nutrient broth medium, 9mL nutrient broth medium and 8mL The nutrient broth culture medium is formulated into sample liquids containing the detection liquid at concentrations of 0.5 ⁇ , 1 ⁇ , and 2 ⁇ , respectively, to obtain the sample liquid of the experimental group;
  • cinnamon oil was used instead of the deodorized cinnamon oil obtained in Example 1 to obtain a control sample.
  • Minimum bacteriostatic concentration test take 0.1 mL of each of the above bacterial suspensions and add 0.5 ⁇ , 1 ⁇ and 2 ⁇ concentration test tubes respectively, and set up three parallel test groups, in which the observation of bacteria is incubated at 37°C for 24h, the fungal Observation was placed at 28°C for 48h; after the end of the cultivation, 1mL of the test tube was pipetted into the plate and poured into an appropriate amount of medium. Bacterial observation was placed at 37°C for 24h and fungal observation was placed at 28°C for 48h; observation of plate colony growth.
  • a method for removing odors of essential oils with antibacterial properties specifically including the following steps: using yeast fermentation broth to process essential oils with antibacterial properties; the essential oils with antibacterial properties are compound essential oils; the compound essential oils are composed of cinnamon oil, clove essential oil and rosemary The essential oil is mixed, and the volume ratio of cinnamon oil: clove essential oil: rosemary essential oil in the compound essential oil is 5:3:2;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 150g;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast culture medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 30°C, shaker Rotation speed is 200r/min; light conditions: dark 7h, light 17h, light intensity 2000lux, until the total number of viable bacteria is 10 8 cfu/mL, the yeast fermentation broth is obtained;
  • the volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation add 250g of compound essential oil to the main fermentation tank containing yeast fermentation broth in step 3, after mixing, adjust the shaker speed to 300r/min, the temperature to 35°C, and then shake the speed
  • the fermentation time is 300r/min and the temperature is 35°C at 96h to obtain the fermentation broth of essential oil;
  • the essential oil fermentation broth is inactivated at 0°C for 24h, then treated with 300W ultrasonic wave for 45min, and finally centrifuged at 12000 rpm for 10min using a high-speed centrifuge, collected to obtain supernatant and precipitate;
  • the water temperature of the rotary evaporator is 50°C and the rotation speed is 45r/ min to obtain essential oil II; all solvents are ethyl acetate; the volume ratio of the mass of the precipitate to the solvent is 1 g: 3 mL;
  • test bacteria were: Staphylococcus aureus, Escherichia coli, Candida albicans, Pseudomonas aeruginosa and Aspergillus niger.
  • the minimum inhibitory concentration MIC of the compounded essential oil and the deodorized compounded essential oil obtained in Example 2 was determined by the liquid medium dilution method. The specific method is as follows:
  • Example 2 Take 1g of the deodorized compound oil obtained in Example 2 and dissolve it in 9mL of DMSO in advance to obtain a sample solution with a concentration of 1/10. Take a sterilized test tube and draw 1mL of the sample solution with a concentration of 1/10, then add 9mL of nutrition Broth culture medium to obtain a sample solution with a concentration of 1/100. Pipette 0.5mL, 1mL and 2mL of sample solution with a concentration of 1/100 into 9.5mL nutrient broth medium, 9mL nutrient broth medium and 8mL of nutrient broth culture medium was prepared into sample solutions containing the test solution at concentrations of 0.5 ⁇ , 1 ⁇ , and 2 ⁇ , respectively, to obtain the sample solution of the experimental group;
  • the compounded essential oil is used in place of the deodorized compounded essential oil obtained in Example 2 for operation to obtain a control sample.
  • Minimum bacteriostatic concentration test take 0.1 mL of each of the above bacterial suspensions and add 0.5 ⁇ , 1 ⁇ and 2 ⁇ concentration test tubes respectively, and set up three parallel test groups, in which the observation of bacteria is incubated at 37°C for 24h, the fungal Observation was placed at 28°C for 48h; after the end of the cultivation, 1mL of the test tube was pipetted into the plate and poured into an appropriate amount of medium. Bacterial observation was placed at 37°C for 24h and fungal observation was placed at 28°C for 48h; observation of plate colony growth.
  • a method for removing odor of essential oil with antibacterial properties specifically including the following steps: processing yeast essential oil with antibacterial properties; the essential oil with antibacterial properties is garlic oil;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast culture medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1500g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 150g;
  • Bacterial species activation add 200g of activation solution to the pre-fermentation tank, and then separately add Polysaccharopolysaccharomyces cerevisiae, Saccharomyces cerevisiae and Angel yeast to activate respectively, to obtain polysaccharolysis Saccharomyces cerevisiae activation liquid and Saccharomyces cerevisiae activation Liquid and Angel yeast activation liquid; the mass fraction of white granulated sugar in the activation liquid is 5%;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast culture medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 30°C, shaker Rotation speed is 200r/min; light conditions: dark 6h, light 18h, light intensity 2000lux, until the total number of viable bacteria is 10 8 cfu/mL, the yeast fermentation broth is obtained;
  • the volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation add 100 g of garlic oil to the main fermentation tank containing yeast fermentation broth in step 3, after mixing, adjust the shaker speed to 250 r/min, the temperature to 30°C, and then shake the shaker speed It is 250r/min and the fermentation time is 96h at a temperature of 30°C to obtain the fermentation broth of essential oil;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, then subjected to 250W ultrasonic treatment for 30min, and finally centrifuged at 12000 rpm for 10min using a high-speed centrifuge and collected to obtain the supernatant and precipitate;
  • the water temperature of the rotary evaporator is 60°C and the rotation speed is 45r/ min to obtain essential oil II; all solvents are ethanol; the volume ratio of the precipitated mass to the solvent is 1g: 5mL;
  • a method for removing odor of essential oil with antibacterial properties specifically including the following steps: processing an essential oil with antibacterial properties using yeast fermentation broth; the essential oil with antibacterial properties is clove oil;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 200g;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast culture medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 30°C, shaker speed of 250r / min; light conditions: dark 8h, light 16h, light intensity 2500lux, to a total viable cell count of 10 8 cfu / far mL, resulting in the yeast fermentation broth; laminating said polyethylene lipolytica toruloides activation solution: The volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation Add 150g clove oil to the main fermentation tank containing yeast fermentation broth in step 3, after mixing, adjust the shaker speed to 300r/min, the temperature to 35°C, and then the shaker speed to Fermentation time at 300r/min and temperature of 35°C for 72h to obtain essential oil fermentation broth;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, and then subjected to 300W ultrasonic treatment for 35min, and finally centrifuged at 12000 rpm for 10min using a high-speed centrifuge and collected to obtain the supernatant and precipitate;
  • the water temperature of the rotary evaporator is 55°C and the rotation speed is 45r/ min to obtain essential oil II; all solvents are ethanol; the volume ratio of the mass of the precipitate to the solvent is 2g: 5mL;
  • a method for removing odors of essential oils with antibacterial properties specifically including the following steps: using yeast fermentation broth to process essential oils with antibacterial properties;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 200g;
  • Bacterial species activation add 100g of the activation solution to the pre-fermentation tank, and then separately add S. cerevisiae, Saccharomyces cerevisiae and Angel yeast to activate, to obtain S. cerevisiae activation solution and S. cerevisiae activation Liquid and Angel yeast activation liquid; the mass fraction of white granulated sugar in the activation liquid is 5%;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast culture medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 30°C, shaker speed of 250r / min; light conditions: dark 7h, light 17h, light intensity 2300lux, to a total viable cell count of 10 8 cfu / far mL, resulting in the yeast fermentation broth; laminating said polyethylene lipolytica toruloides activation solution: The volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation add 100g of Guanghuo fragrant oil to the main fermentation tank containing yeast fermentation broth in step 3, after mixing, adjust the shaker speed to 200r/min, the temperature to 35°C, and then shake the speed
  • the fermentation time is 200r/min and the temperature is 35°C at 72h, and the essential oil fermentation broth is obtained;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, then treated with 250W ultrasonic wave for 45min, and finally centrifuged at 12000 rpm for 10min using a high-speed centrifuge, collected to obtain the supernatant and precipitate;
  • the water temperature of the rotary evaporator is 50°C and the rotation speed is 45r/ min to obtain essential oil II; all solvents are ether; the volume ratio of the mass of the precipitate to the solvent is 1 g: 3 mL;
  • a method for removing odor of essential oil with antibacterial properties specifically including the following steps: processing yeast essential oil with antibacterial properties; the essential oil with antibacterial properties is nutmeg oil;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 300g;
  • Bacterial species activation add 100g of the activation solution to the pre-fermentation tank, and then separately add S. cerevisiae, Saccharomyces cerevisiae and Angel yeast to activate, to obtain S. cerevisiae activation solution and S. cerevisiae activation Liquid and Angel yeast activation liquid; the mass fraction of white granulated sugar in the activation liquid is 5%;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 28°C, shaker speed of 200r / min; light conditions: dark 8h, light 16h, light intensity 2500lux, to a total viable cell count of 10 8 cfu / far mL, resulting in the yeast fermentation broth; laminating said polyethylene lipolytica toruloides activation solution: The volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation Add 300g of nutmeg oil to the main fermentation tank containing yeast fermentation broth in step 3. After mixing, adjust the shaker speed to 300r/min, the temperature to 35°C, and then shake the speed The fermentation time is 300r/min and the temperature is 35°C at 72h, and the essential oil fermentation broth is obtained;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, and then subjected to 300W ultrasonic treatment for 35min, and finally centrifuged at 12000 rpm for 10min using a high-speed centrifuge and collected to obtain the supernatant and precipitate;
  • the water temperature of the rotary evaporator is 60°C and the rotation speed is 65r/ min to obtain essential oil II; all solvents are ethanol; the volume ratio of the mass of the precipitate to the solvent is 2g: 3mL;
  • a method for removing odors of essential oils with antibacterial properties specifically including the following steps: processing essential oils with antibacterial properties using yeast fermentation broth; the essential oils with antibacterial properties are star anise oil;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 200g;
  • Bacterial species activation add 100g of the activation solution to the pre-fermentation tank, and then separately add S. cerevisiae, Saccharomyces cerevisiae and Angel yeast to activate, to obtain S. cerevisiae activation solution and S. cerevisiae activation Liquid and Angel yeast activation liquid; the mass fraction of white granulated sugar in the activation liquid is 5%;
  • Enrichment fermentation put the polylipopolysaccharopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast medium into the main fermentation tank for shaker fermentation, the fermentation temperature is 28°C, shaker speed of 200r / min; light conditions: dark 6h, light 18h, light intensity 2000lux, to a total viable cell count of 10 8 cfu / far mL, resulting in the yeast fermentation broth; laminating said polyethylene lipolytica toruloides activation solution: The volume ratio of Saccharomyces cerevisiae activation solution: Angel yeast activation solution is 1:1:1; and the total added amount of S. cerevisiae activation solution, Saccharomyces cerevisiae activation solution and Angel yeast activation solution is 10%;
  • Deodorization fermentation add 300g star anise oil to the main fermentation tank containing yeast fermentation broth in step 3, after mixing, adjust the shaker speed to 300r/min, the temperature to 35°C, and then shake the speed
  • the fermentation time is 300r/min and the temperature is 35°C at 48h, and the essential oil fermentation broth is obtained;
  • the essential oil fermentation broth is first inactivated at 0°C for 24h, then subjected to 200W ultrasonic treatment for 15min, and finally centrifuged at 10,000 rpm for 10min using a high-speed centrifuge and collected to obtain the supernatant and precipitate;
  • the water temperature of the rotary evaporator is 60°C and the rotation speed is 60 r/ min to obtain essential oil II; all solvents are ethanol; the volume ratio of the mass of the precipitate to the solvent is 2g: 3mL;
  • step 4 the fermentation time is 120 h at a shaker rotation speed of 300 r/min and a temperature of 35° C. An essential oil fermentation broth is obtained. Others are the same as in the first embodiment.
  • test bacteria were: Staphylococcus aureus, Escherichia coli, Candida albicans, Pseudomonas aeruginosa and Aspergillus niger.
  • the minimum inhibitory concentration MIC of cinnamon oil and the deodorized cinnamon oil obtained in Example 1 was determined by the liquid medium dilution method.
  • the specific method is as follows:
  • Example 2 Take 1g of the deodorized cinnamon oil obtained in Example 1 and dissolve it in 9mL of DMSO in advance to obtain a sample solution with a concentration of 1/10. Take a sterilized test tube, draw 1mL of the sample solution with a concentration of 1/10, and then add 9mL of nutrient meat Soup medium to obtain a sample solution with a concentration of 1/100. Pipette 0.5mL, 1mL and 2mL of sample solution with a concentration of 1/100 into 9.5mL nutrient broth medium, 9mL nutrient broth medium and 8mL The nutrient broth culture medium is formulated into sample liquids containing the detection liquid at concentrations of 0.5 ⁇ , 1 ⁇ , and 2 ⁇ , respectively, to obtain the sample liquid of the experimental group;
  • Example 8 the deodorized cinnamon oil obtained in Example 8 was used instead of the deodorized cinnamon oil obtained in Example 1 to obtain a control sample.
  • Minimum bacteriostatic concentration test take 0.1 mL of each of the above bacterial suspensions and add 0.5 ⁇ , 1 ⁇ and 2 ⁇ concentration test tubes respectively, and set up three parallel test groups, in which the observation of bacteria is incubated at 37°C for 24h, the fungal Observation was placed at 28°C for 48h; after the end of the cultivation, 1mL of the test tube was pipetted into the plate and poured into an appropriate amount of medium. Bacterial observation was placed at 37°C for 24h and fungal observation was placed at 28°C for 48h; observation of plate colony growth.
  • step 3 the polylipopolysaccharomyces cerevisiae activation solution, Angel yeast activation solution and yeast medium are put into the main fermentation tank for shaker fermentation, the fermentation temperature is 30°C shaking speed of 250r / min; light conditions: dark 7H, 17H light, light intensity 2300lux, to a total viable cell count of 10 8 cfu / far mL, resulting in the yeast fermentation broth; laminating said polyethylene lipolytica toruloides
  • the volume ratio of the activation solution: Angel yeast activation solution is 1:1; and the total added amount of the polylipopolysaccharomyces cerevisiae activation solution and Angel yeast activation liquid is 10%.
  • Others are the same as in the first embodiment.
  • test bacteria were: Staphylococcus aureus, Escherichia coli, Candida albicans, Pseudomonas aeruginosa and Aspergillus niger.
  • the minimum inhibitory concentration MIC of cinnamon oil and the deodorized cinnamon oil obtained in Example 1 was determined by the liquid medium dilution method.
  • the specific method is as follows:
  • Example 2 Take 1g of the deodorized cinnamon oil obtained in Example 1 and dissolve it in 9mL of DMSO in advance to obtain a sample solution with a concentration of 1/10. Take a sterilized test tube, draw 1mL of the sample solution with a concentration of 1/10, and then add 9mL of nutrient meat Soup medium to obtain a sample solution with a concentration of 1/100. Pipette 0.5mL, 1mL and 2mL of sample solution with a concentration of 1/100 into 9.5mL nutrient broth medium, 9mL nutrient broth medium and 8mL The nutrient broth culture medium is formulated into sample liquids containing the detection liquid at concentrations of 0.5 ⁇ , 1 ⁇ , and 2 ⁇ , respectively, to obtain the sample liquid of the experimental group;
  • Example 9 the deodorized cinnamon oil obtained in Example 9 was used instead of the deodorized cinnamon oil obtained in Example 1 to obtain a control sample.
  • Minimum bacteriostatic concentration test take 0.1 mL of each of the above bacterial suspensions and add 0.5 ⁇ , 1 ⁇ and 2 ⁇ concentration test tubes respectively, and set up three parallel test groups, in which the observation of bacteria is incubated at 37°C for 24h, the fungal Observation was placed at 28°C for 48h; after the end of the cultivation, 1mL of the test tube was pipetted into the plate and poured into an appropriate amount of medium. Bacterial observation was placed at 37°C for 24h and fungal observation was placed at 28°C for 48h; observation of plate colony growth.
  • Example 10 Comparison of fermentation of different yeasts on the deodorization of cinnamon oil:
  • a method for removing odor of essential oil with antibacterial properties specifically including the following steps: processing yeast essential oil with antibacterial properties; the essential oil with antibacterial properties is cinnamon oil;
  • yeast fermentation broth to treat essential oils with antibacterial properties includes the following steps:
  • Configure yeast medium add white granulated sugar, isomaltose oligosaccharide, polydextrose, peptone and 1000g of sterile water to the inoculum fermentation tank. After completely dissolved, transfer the inoculum fermentation tank to an autoclave at the temperature Sterilize at 121°C for 20 min, cool to room temperature to obtain an initial mixture, and sterilize glucose at 115°C for 30 min, then add to the initial mixture, dissolve and mix to obtain a yeast medium; the yeast medium The mass ratio of medium white sugar: glucose: isomaltose: polydextrose: peptone is 4:3:1:1:1, and the total mass of white sugar, glucose, isomaltose, polydextrose and peptone is 200g;
  • Bacterial species activation Add 6 parts of 100g activation solution to the pre-fermentation tank, and then add P. lipolytica Saccharomyces cerevisiae, Saccharomyces cerevisiae, Angel yeast, white wine special yeast, 18 degree wine yeast and acid-lowering yeast The activation is performed to obtain the polylipopolysaccharomyces cerevisiae activation solution, the Saccharomyces cerevisiae activation solution, the Angel yeast activation solution, the white wine special yeast activation solution, the 18-degree wine yeast activation solution and the acid-lowering yeast activation solution; The mass fraction of white granulated sugar is 5%;
  • Enrichment fermentation the polylipopolysaccharomyces cerevisiae activation solution, Saccharomyces cerevisiae activation solution, Angel yeast activation solution, white wine special yeast activation solution, 18 degree wine yeast activation solution and acid-lowering yeast activation solution are used as yeast activation
  • the liquid was put into the main fermentation tank with yeast culture medium for shaker fermentation, the fermentation temperature was 30°C, the shaker speed was 250r/min; light conditions: dark 8h, light 16h, light intensity 2000lux, to total live bacteria Up to 10 8 cfu/mL, obtain the polylipopolysaccharomyces cerevisiae fermentation broth, Saccharomyces cerevisiae fermentation broth, Angel yeast fermentation broth, white wine special yeast fermentation broth, 18 degree wine yeast fermentation broth and acid-lowering yeast fermentation broth ; The addition amount of the yeast activation solution is 10%;
  • Deodorization fermentation Add 6 parts of cinnamon oil to the fermentation liquid containing Saccharomyces cerevisiae, Saccharomyces cerevisiae fermentation liquid, Angel yeast fermentation liquid, white wine special yeast fermentation liquid, 18 degree wine yeast
  • 100g of cinnamon oil per serving After mixing, adjust the shaker speed to 300r/min, the temperature to 35°C, and then shaker speed to 300r/min and temperature
  • the fermentation time at 35°C is 72h, and 6 kinds of essential oil fermentation broth are obtained;
  • Extract the precipitate in the extraction tank with a solvent to obtain a solvent extract Use a rotary evaporator to concentrate the solvent extract until all solvents are separated by rotary evaporation.
  • the water temperature of the rotary evaporator is 50°C and the rotation speed is 45r/min to obtain essential oil II.
  • All solvents are ethyl acetate; the volume ratio of the mass of the precipitate to the solvent is 1g: 3mL;
  • step 6 The six groups of essential oil I and essential oil II obtained in step 6 are combined respectively to obtain cinnamon oil after deodorization of Polysaccharopolysaccharomyces pombe, yeast oil after deodorization of Saccharomyces cerevisiae, cinnamon oil after deodorization of Angel yeast, Cinnamon oil after deodorization of yeast for white wine, cinnamon oil after deodorization of 18 degree wine yeast and cinnamon oil after deodorization of acid-lowering yeast.
  • cinnamon oil after deodorization of Saccharomyces cerevisiae cinnamon oil after deodorization of Angel Yeast
  • cinnamon oil after deodorization of yeast for white wine after deodorization of cinnamon oil for yeast for white wine
  • deodorization of 18 degree wine yeast Comparison of odor change of cinnamon oil and cinnamon oil after deodorization:
  • Cinnamon oil cinnamon oil after deodorization of Polysaccharopolysaccharomyces cerevisiae, cinnamon oil after deodorization of Saccharomyces cerevisiae, cinnamon oil after deodorization of Angel yeast, cinnamon oil after deodorization of white wine special yeast, 18 degree wine yeast Comparison of antibacterial activity of cinnamon oil and acid-reducing yeast after deodorization on the Escherichia coli, Pseudomonas aeruginosa and Aspergillus niger;
  • the tested bacteria were: Escherichia coli, Pseudomonas aeruginosa and Aspergillus niger.
  • Cinnamon oil cinnamon oil after deodorization of Polysaccharomyces pombe, Saccharomyces cerevisiae after deodorization, cinnamon oil after deodorization of Saccharomyces cerevisiae, cinnamon oil after deodorization of Angel yeast, cinnamon oil after deodorization of white wine special yeast,
  • the minimum inhibitory concentration MIC of cinnamon oil after deodorization of 18-degree wine yeast and cinnamon oil after deodorization of acid-reducing yeast is as follows:
  • the sample solution containing the detection solution namely, the cinnamon oil sample solution, the cinnamon oil sample solution after deodorization of Polysaccharopolysaccharomyces pombe, the cinnamon oil sample solution after deodorization of Saccharomyces cerevisiae, and the cinnamon oil sample solution after deodorization of Angel yeast, White wine special yeast cinnamon oil sample liquid, 18 degree wine yeast cinnamon oil sample liquid and acid-reducing yeast cinnamon oil sample liquid;
  • Minimum bacteriostatic concentration test Take 0.1mL of each of the above bacterial suspensions into a 1 ⁇ concentration test tube, and set up three parallel test groups, in which the observation of bacteria is incubated at 37°C for 24h, and the observation of fungi is incubated at 28°C for 48h After the cultivation is completed, pipette 1mL into the plate, pour the appropriate amount of medium, observe the bacteria at 37 °C for 24h, and observe the fungi at 28 °C for 48h; observe the growth of the colony on the plate.
  • Cinnamon oil cinnamon oil after deodorization of Polysaccharopolysaccharomyces cerevisiae, cinnamon oil after deodorization of Saccharomyces cerevisiae, cinnamon oil after deodorization of Angel Yeast, cinnamon oil after deodorization of white wine special yeast, after deodorization of 18 degree wine yeast
  • Table 15 The bacteriostatic activity comparison results of cinnamon oil and acid-reducing yeast on the Escherichia coli, Pseudomonas aeruginosa and Aspergillus niger are shown in Table 15:
  • Example 11 Comparison of fermentation of deodorization of cinnamon oil by different organic reagents:
  • step 6 ethyl acetate, ethanol, ether, cyclohexane, benzene and acetone are used as organic solvents. Others are the same as in the first embodiment.
  • the tested bacteria were: Escherichia coli, Pseudomonas aeruginosa and Aspergillus niger.
  • the minimum bacteriostatic concentration MIC of ethyl acetate essential oil I, ethanol essential oil I, diethyl ether essential oil I, cyclohexane essential oil I, benzene essential oil I, acetone essential oil I and the essential oil I obtained in Example 1 was determined by the liquid medium dilution method. Methods as below:
  • Minimum bacteriostatic concentration test Take 0.1mL of each of the above bacterial suspensions into a 1 ⁇ concentration test tube, and set up three parallel test groups, in which the observation of bacteria is incubated at 37°C for 24h, and the observation of fungi is incubated at 28°C for 48h After the cultivation is completed, pipette 1mL into the plate, pour the appropriate amount of medium, observe the bacteria at 37 °C for 24h, and observe the fungi at 28 °C for 48h; observe the growth of the colony on the plate.

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Abstract

一种具有抗菌性能精油气味去除方法，利用酵母发酵液处理具有抗菌性能精油。所述方法的优点是有效去除或减轻具有抗菌性能精油中不愉悦的气味，对具有抗菌性能精油不会产生额外的有害物质添加，同时又能极大的保留具有抗菌性能精油的抑菌活性。

Description

一种具有抗菌性能精油气味去除方法 技术领域
本发明涉及一种有效去掉精油味道的方法。
背景技术
国际上，食品添加剂的发展趋向是往天然、营养方面发展。科学工作者对化学合成的、在自然界中不存在的食品添加剂持慎重的科学态度，消费者则对非天然食品添加剂非常忌讳。特别地在食品防腐剂的使用上，很多食品厂为了达到长时间的食品防腐效果，超量添加现象严重。因此，从动植物中寻找，提取天然食品防腐剂并应用到当前食品行业甚至是化妆品行业的研究成为主流。我国地域辽阔，植物资源十分丰富，从植物中提取防腐剂有广阔的基础及应用前景。
目前，我国针对植物源防腐剂已经筛选出很多有应用前景的种类，例如丁香、大蒜、紫苏、肉桂、小茴香、洋葱、八角等。这些植物提取物一般显示出有很好的抑菌广谱性，完全可以作为防腐剂应用到食品和化妆品中。而且随着国家对药食同源的相关制定和规范，使得这些植物提取物能够更好地应用在食品领域中。
虽然有如此一个很好的应用前景，但是研究了这么多年，植物源防腐剂一直长期处于原地踏步状态。这个中原因有两个，一方面是过往国家相关政策对防腐剂的使用管理相对宽松，而像人工合成防腐剂苯甲酸钠、山梨酸钾的成本很低，防腐效果好的背景下，植物源防腐剂举步维艰。在国外，苯甲酸钠因为对人体健康原因早已被禁止使用，在国内，还可以正常使用，但随着人们对健康的要求越来越高，对食品添加剂的关注度也比以往要多，植物源防腐剂得到越来越多消费者的欢迎；另一方面限制植物源防腐剂发展的原因主要是植物提取物中会含有一些特殊的气味，如果添加到食品中，对于一些自身味道相对轻的食品来说，它会极大破坏食品自身的味道，甚至让消费者有不愉悦的感受。针对这些情况，做法一般是通过香清的添加来抑制这些植物提取物的气味，但是添加到食品中后，植物提取物浓烈的气味还是会在品尝过程中显现出来，这也是大大限制植物提取物作为防腐剂应用到食品领域的一大障碍。针对这一情况，目前还没有找到能解决这一弊端的方法。
目前植物源防腐剂还没有很好地在国内应用起来，皆因它在味道上有一定的局限性限制它在食品领域中的应用。通过添加香精，的确可以使得植物提取物固有的气味给掩盖掉，但是在应用到例如蛋糕、布丁、面包等味道本来就相对轻的食品时，消费者在品尝这些食品时植物提取物的气味就会明显体会到，也就是说香精和浓烈的植物气味完成辨别出来。
发明内容
本发明的目的是要解决现有技术无法有效去掉精油味道的问题，而提供一种具有抗菌性能精油气味去除方法。
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油。
所述所述酵母发酵液中酵母的活菌数为10 7cfu/mL～10 9cfu/mL。
所述所述酵母发酵液中的酵母为聚解脂复膜孢酵母、降酸酵母、酿酒酵母、18度葡萄酒酵母、安琪酵母和白葡萄酒专用酵母中的一种或其中几种。
所述具有抗菌性能精油为单一精油或复配精油。
所述单一精油为肉桂油、丁香精油、迷迭香精油、大蒜油、广霍香油、肉豆蔻油、八角茴香油、山苍子精油、南丰蜜桔精油、飞蓬精油或胜红蓟精油。
所述复配精油由肉桂油、丁香精油、迷迭香精油、广霍香油、肉豆蔻油和八角茴香油中两种或两种以上混合而成。
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
(1)、发酵：将具有抗菌性能精油加入酵母发酵液中，混匀后发酵处理，得精油发酵液；所述具有抗菌性能精油与酵母发酵液的体积比为6～10∶1；
(2)、分离：对精油发酵液依次进行灭活处理、超声波处理和离心处理，得到上清液和沉淀；
(3)、萃取：对步骤(2)得到的上清液依次进行浓缩、萃取和再次浓缩，得到精油I；对步骤(2)得到的沉淀依次进行萃取和浓缩，得到精油II；
(4)、合并：合并步骤(3)得到的精油I和精油II，即得到去除气味后具有抗菌性能精油。
本发明优点：本发明选择食用酵母有效去除(或减轻)具有抗菌性能精油中不愉悦的气味，对具有抗菌性能精油不会产生额外的有害物质添加，同时又能极大的保留具有抗菌性能精油的抑菌活性。
本发明得到的去除气味后具有抗菌性能精油作为防腐剂使用。
具体实施方式
具体实施方式一：
本实施方式是一种具有抗菌性能精油气味去除方法，具体包括以下步骤：
利用酵母发酵液处理具有抗菌性能精油。
现有的抗菌精油去除或减轻气味，直接采用有机溶剂的方法，而本实施方式采用酵母发酵液，对具有抗菌性能精油不会产生额外的有害物质添加，最大限度的保留了天然提取物这个概念。
具有抗菌性能精油采用菌种发酵去除气味时，菌种的活性容易被抑制，而无法产生发酵；因此本实施方式选择酵母发酵液处理具有抗菌性能精油，成功产生发酵，有效去除(或减轻)具有抗菌性能精油中不愉悦的气味。
具有抗菌性能精油采用菌种发酵时，涉及到其中的官能团转化，需要平衡保留抗菌活性和去除/减轻气味这2者的关系，很容易产生去除气味的同时抗菌活性也被削弱的问题；而本实施方式选择酵母发酵液处理具有抗菌性能精油，不仅有效去除(或减轻)具有抗菌性能精油中不愉悦的气味，同时又能极大的保留具有抗菌性能精油的抑菌活性。
具体实施方式二：
本实施方式与具体实施方式一的不同点是：所述所述酵母发酵液中酵母的活菌数为10 7cfu/mL～10 9cfu/mL。其他与具体实施方式一相同。
具体实施方式三：
本实施方式与具体实施方式一或二之一不同点是：所述所述酵母发酵液中的酵母为聚解脂复膜孢酵母、降酸酵母、酿酒酵母、18度葡萄酒酵母、安琪酵母和白葡萄酒专用酵母中的一种或其中几种。其他与具体实施方式一或二相同。
本实施方式所述的聚解脂复膜孢酵母是广东微生物研究所生产的，保藏编号：ATCC18942。
本实施方式所述的降酸酵母、酿酒酵母、18度葡萄酒酵母和白葡萄酒专用酵母是山东烟台帝伯仕酵母有限公司生产的。
本实施方式所述的安琪酵母是安琪酵母股份有限公司生产的。
具体实施方式四：
本实施方式与具体实施方式一至三之一不同点是：所述具有抗菌性能精油为单一精油或复配精油。其他与具体实施方式一至三相同。
具体实施方式五：
本实施方式与具体实施方式四的不同点是：所述单一精油为肉桂油、丁香精油、迷迭香精油、大蒜油、广霍香油、肉豆蔻油、八角茴香油、山苍子精油、南丰蜜桔精油、飞蓬精油或胜红蓟精油。其他与具体实施方式四相同。
具体实施方式六：
本实施方式与具体实施方式四的不同点是：所述复配精油由肉桂油、丁香精油、迷迭香精油、广霍香油、肉豆蔻油和八角茴香油中两种或两种以上混合而成。其他与具体实施方式四相同。
具体实施方式七：
本实施方式与具体实施方式一至六之一不同点是：所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
(1)、发酵：将具有抗菌性能精油加入酵母发酵液中，混匀后发酵处理，得精油发酵液；所述具有抗菌性能精油与酵母发酵液的体积比为6～10∶1；
(2)、分离：对精油发酵液依次进行灭活处理、超声波处理和离心处理，得到上清液和沉淀；
(3)、萃取：对步骤(2)得到的上清液依次进行浓缩、萃取和再次浓缩，得到精油I；对步骤(2)得到的沉淀依次进行萃取和浓缩，得到精油II；
(4)、合并：合并步骤(3)得到的精油I和精油II，即得到去除气味后具有抗菌性能精油。
其他与具体实施方式一至六之相同。
具体实施方式八：
本实施方式与具体实施方式七的不同点是：步骤(1)中所述的酵母发酵液是按以下步骤获得：
①、配置酵母培养基：将复合营养物加入无菌水中，复合营养物完全溶解后转移至高压灭菌锅中，在温度为115℃～121℃下灭菌20min～30min，冷却至室温后，得到酵母培养基；所述复合营养物与无菌水的质量比为3∶10～90；所述复合营养物由白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨组成，且复合营养物中按质量比白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨＝4∶3∶1∶1∶1；
②、菌种活化：将酵母放入活化液中活化，得到含活化后酵母的酵母活化液；所述活化液中白砂糖的质量分数为5～15％；
③、发酵：将含活化后酵母的酵母活化液放入酵母培养基进行摇床发酵，发酵温度为24～35℃，摇床转速为150r/min～300r/min；光照条件：黑暗6h～8h，光照16h～18h，光照强度为2000lux～2500lux，至酵母的活菌数为10 7cfu/mL～10 9cfu/mL为止，得到酵母发酵液。
其他与具体实施方式七相同。
具体实施方式九：
本实施方式与具体实施方式七或八之一不同点是：步骤(1)中所述发酵处理具体操作过程如下：
将摇床转速调整至150r/min～300r/min，温度调整至30～65℃，然后在摇床转速为150r/min～300r/min和温度为30～65℃下发酵时间为72h～120h。
其他与具体实施方式七或八相同。
具体实施方式十：
本实施方式与具体实施方式七至九之一不同点是：步骤(2)中将精油发酵液在0℃下灭活处理24h，然后经250W～300W超声波处理15min～60min，最后12000转/min离心处理10min～25min，收集，得到上清液和沉淀。其他与具体实施方式七至九相同。
具体实施方式十一：
本实施方式与具体实施方式七至十之一不同点是：于步骤(3)中对步骤(2)得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，得到精油I；所述有机溶剂选自乙酸乙酯、乙醇、乙醚和环己烷；所述上清液浓缩液与有机溶剂的体积比为1～3∶3～6；
对步骤(2)得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，得到精油II；所有溶剂选自乙酸乙酯、乙醇、乙醚和环己烷；所述沉淀的质量与溶剂的体积比为1g～4g∶2mL～6mL。
其他与具体实施方式七至十相同。
本实施方式所述的有机溶剂全部旋转蒸发分离为止中旋转蒸发仪水温为45～60℃，转速45r/min。
本实施方式所述的溶剂全部旋转蒸发分离为止中旋转蒸发仪水温为45～60℃，转速45r/min。
具体实施方式十二：
本实施方式与具体实施方式七至十一之一不同点是：所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将复合营养物和无菌水加入种菌发酵槽中，复合营养物完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为115℃～121℃下灭菌20min～30min，冷却至室温后，得到酵母培养基；所述复合营养物与无菌水的质量比为3∶10～90；所述复合 营养物由白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨组成，且复合营养物中按质量比白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨＝4∶3∶1∶1∶1；
2、菌种活化：将活化液加入前期发酵槽中，然后加入酵母进行活化，得到含活化后酵母的酵母活化液；所述活化液中白砂糖的质量分数为5～15％；
3、富集发酵：将含活化后酵母的酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为24～35℃，摇床转速为150r/min～300r/min；光照条件：黑暗6h～8h，光照16h～18h，光照强度为2000lux～2500lux，至酵母的活菌数为10 7cfu/mL～10 9cfu/mL为止，得到酵母发酵液；
4、除味发酵：将具有抗菌性能精油加入装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至150r/min～300r/min，温度调整至30～65℃，然后在摇床转速为150r/min～300r/min和温度为30～65℃下发酵时间为72h～120h，得精油发酵液；所述具有抗菌性能精油与酵母发酵液的体积比为6～10∶1；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经250W～300W超声波处理15min～60min，最后采用高速离心机以12000转/min离心处理10min～25min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为45～60℃，转速45r/min，得到精油I；所述有机溶剂选自乙酸乙酯、乙醇、乙醚和环己烷；所述上清液浓缩液与有机溶剂的体积比为1～3∶3～6；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为45～60℃，转速45r/min～65r/min得到精油II；所有溶剂选自乙酸乙酯、乙醇、乙醚和环己烷；所述沉淀的质量与溶剂的体积比为1g～4g∶2mL～6mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去除气味后具有抗菌性能精油。
本发明内容不仅限于上述各实施方式的内容，其中一个或几个具体实施方式的组合同样也可以实现发明的目的。
采用下述试验验证本发明效果
实施例1：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有 抗菌性能精油；所述具有抗菌性能精油为肉桂油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为200g；
2、菌种活化：将100g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为250r/min；光照条件：黑暗7h，光照17h，光照强度为2300lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将300g肉桂油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至300r/min，温度调整至35℃，然后在摇床转速为300r/min和温度为35℃下发酵时间为72h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经300W超声波处理25min，最后采用高速离心机以12000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min，得到精油I；所述有机溶剂由乙酸乙酯和乙醇按体积比1∶1混合而成；所述上清液浓缩液与有机溶剂的体积比为1∶3；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min得到精油II；所有溶剂为乙酸乙酯；所述沉淀的质量与溶剂的体积比为1g∶3mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后肉桂油。
(一)肉桂油与实施例1得到的去味后肉桂油进行气味的变化比较：
感官评价描述数据参照表1。
表1
Figure PCTCN2020077362-appb-000001
肉桂油与实施例1得到的去味后肉桂油的气味评价结果如表2所示。
表2
Figure PCTCN2020077362-appb-000002
(二)肉桂油与实施例1得到的去味后肉桂油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比；
证明实验效果的实验方案和实验结果数据
以肉桂油与实施例1得到的去味后肉桂油进行抑菌效果的定量试验
供试菌种为：金黄色葡萄球菌、大肠杆菌、白色念珠菌、铜绿假单胞杆菌和黑曲霉。
采用液体培养基稀释法测定肉桂油和实施例1得到的去味后肉桂油的最低抑菌浓度MIC，具体方法如下：
(1)培养基的制备：营养肉汤培养基；
(2)菌悬液的制备：取活化后的金黄色葡萄球菌、活化后的大肠杆菌、活化后的铜绿假单胞杆菌于灭菌蒸馏水中分别制成1×10 8cfu/ml的菌悬液；取活化后的白色念珠菌和活化后的黑曲霉于灭菌蒸馏水中分别制成1×10 7cfu/ml的菌悬液；
(3)含待检测液试管的制备：以实施例1得到的去味后肉桂油为实验组，以肉桂油为对照组，
取1g实施例1得到的去味后肉桂油预先溶于9mL DMSO中，得到浓度为1/10的样液，取灭菌试管，吸取1mL浓度为1/10的样液，再加入9mL营养肉汤培养基，得到浓度为1/100的样液，分别吸取0.5mL、1mL和2mL浓度为1/100的样液依次加入9.5mL 的营养肉汤培养基、9mL的营养肉汤培养基和8mL的营养肉汤培养基中，配制成浓度分别为0.5‰、1‰和2‰的含检测液的样液，得到实验组样液；
按照上述过程，采用肉桂油代替实施例1得到的去味后肉桂油进行操作，得到对照组样液。
(4)最低抑菌浓度试验：取上述各菌悬液0.1mL分别加入0.5‰、1‰和2‰浓度试管，设置三组平行试验组，其中细菌的观察置于37℃培养24h，真菌的观察置于28℃培养48h；培养结束后试管分别吸取1mL于平板中，倒入适量培养基，细菌观察置于37℃培养24h，真菌的观察置于28℃培养48h；观察平板菌落生长情况。
肉桂油与实施例1得到的去味后肉桂油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比结果如表3和表4所述：
表3
Figure PCTCN2020077362-appb-000003
“+”代表有生长“-”代表无生长
表4
Figure PCTCN2020077362-appb-000004
“+”代表有生长“-”代表无生长
实施例2：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有 抗菌性能精油；所述具有抗菌性能精油为复配精油；所述复配精油由肉桂油、丁香精油和迷迭香精油混合而成，且复配精油中肉桂油∶丁香精油∶迷迭香精油的体积比为5∶3∶2；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为150g；
2、菌种活化：将300g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为200r/min；光照条件：黑暗7h，光照17h，光照强度为2000lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将250g复配精油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至300r/min，温度调整至35℃，然后在摇床转速为300r/min和温度为35℃下发酵时间为96h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经300W超声波处理45min，最后采用高速离心机以12000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min，得到精油I；所述有机溶剂由乙酸乙酯和环己烷按体积比2∶1混合而成；所述上清液浓缩液与有机溶剂的体积比为1∶4；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转 速45r/min得到精油II；所有溶剂为乙酸乙酯；所述沉淀的质量与溶剂的体积比为1g∶3mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后复配精油。
(一)复配精油与实施例2得到的去味后复配精油进行气味的变化比较：
感官评价描述数据参照表5。
表5
Figure PCTCN2020077362-appb-000005
复配精油与实施例2得到的去味后复配精油的气味评价结果如表6所示。
表6
Figure PCTCN2020077362-appb-000006
(二)复配精油与实施例2得到的去味后复配精油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比；
证明实验效果的实验方案和实验结果数据
以复配精油与实施例2得到的去味后复配精油进行抑菌效果的定量试验
供试菌种为：金黄色葡萄球菌、大肠杆菌、白色念珠菌、铜绿假单胞杆菌和黑曲霉。
采用液体培养基稀释法测定复配精油与实施例2得到的去味后复配精油的最低抑菌浓度MIC，具体方法如下：
(1)培养基的制备：营养肉汤培养基；
(2)菌悬液的制备：取活化后的金黄色葡萄球菌、活化后的大肠杆菌、活化后的铜绿假单胞杆菌于灭菌蒸馏水中分别制成1×10 8cfu/ml的菌悬液；取活化后的白色念珠菌和活化后的黑曲霉于灭菌蒸馏水中分别制成1×10 7cfu/ml的菌悬液；
(3)含待检测液试管的制备：以实施例2得到的去味后复配精油为实验组，以复配精油为对照组，
取1g实施例2得到的去味后复配精油预先溶于9mL DMSO中，得到浓度为1/10的样液，取灭菌试管，吸取1mL浓度为1/10的样液，再加入9mL营养肉汤培养基，得到 浓度为1/100的样液，分别吸取0.5mL、1mL和2mL浓度为1/100的样液依次加入9.5mL的营养肉汤培养基、9mL的营养肉汤培养基和8mL的营养肉汤培养基中，配制成浓度分别为0.5‰、1‰和2‰的含检测液的样液，得到实验组样液；
按照上述过程，采用复配精油代替实施例2得到的去味后复配精油进行操作，得到对照组样液。
(4)最低抑菌浓度试验：取上述各菌悬液0.1mL分别加入0.5‰、1‰和2‰浓度试管，设置三组平行试验组，其中细菌的观察置于37℃培养24h，真菌的观察置于28℃培养48h；培养结束后试管分别吸取1mL于平板中，倒入适量培养基，细菌观察置于37℃培养24h，真菌的观察置于28℃培养48h；观察平板菌落生长情况。
以复配精油与实施例2得到的去味后复配精油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比结果如表7和表8所示：
表7
Figure PCTCN2020077362-appb-000007
“+”代表有生长“-”代表无生长
表8
Figure PCTCN2020077362-appb-000008
“+”代表有生长“-”代表无生长
实施例3：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油；所述具有抗菌性能精油为大蒜油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1500g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为150g；
2、菌种活化：将200g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为200r/min；光照条件：黑暗6h，光照18h，光照强度为2000lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将100 g大蒜油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至250r/min，温度调整至30℃，然后在摇床转速为250r/min和温度为30℃下发酵时间为96h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经250W超声波处理30min，最后采用高速离心机以12000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为60℃，转速45r/min，得到精油I；所述有机溶剂为乙醇；所述上清液浓缩液与有机溶剂的体积比为1∶5；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为60℃，转 速45r/min得到精油II；所有溶剂为乙醇；所述沉淀的质量与溶剂的体积比为1g∶5mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后大蒜油。
实施例4：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油；所述具有抗菌性能精油为丁香油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为200g；
2、菌种活化：将300g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为250r/min；光照条件：黑暗8h，光照16h，光照强度为2500lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将150g丁香油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至300r/min，温度调整至35℃，然后在摇床转速为300r/min和温度为35℃下发酵时间为72h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经300W超声波处理35min，最后采用高速离心机以12000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为55℃，转速45r/min，得到精油I；所述有机溶剂由乙醚与乙醇按体积比3∶2混 合而成；所述上清液浓缩液与有机溶剂的体积比为1∶4；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为55℃，转速45r/min得到精油II；所有溶剂为乙醇；所述沉淀的质量与溶剂的体积比为2g∶5mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后丁香油。
实施例5：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油；所述具有抗菌性能精油为广霍香油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为200g；
2、菌种活化：将100g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为250r/min；光照条件：黑暗7h，光照17h，光照强度为2300lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将100g广霍香油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至200r/min，温度调整至35℃，然后在摇床转速为200r/min和温度为35℃下发酵时间为72h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经250W超声波处理45min，最后采用高速离心机以12000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓 缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min，得到精油I；所述有机溶剂由乙醚与环己烷按体积比3∶2混合而成；所述上清液浓缩液与有机溶剂的体积比为1∶3；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min得到精油II；所有溶剂为乙醚；所述沉淀的质量与溶剂的体积比为1g∶3mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后广霍香油。
实施例6：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油；所述具有抗菌性能精油为肉豆蔻油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为300g；
2、菌种活化：将100g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为28℃，摇床转速为200r/min；光照条件：黑暗8h，光照16h，光照强度为2500lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将300g肉豆蔻油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至300r/min，温度调整至35℃，然后在摇床转速为300r/min和温度为35℃下发酵时间为72h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经300W超声波处理35min，最后采用高速离心机以12000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为60℃，转速65r/min，得到精油I；所述有机溶剂由乙醇与环己烷按体积比1∶1混合而成；所述上清液浓缩液与有机溶剂的体积比为1∶3；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为60℃，转速65r/min得到精油II；所有溶剂为乙醇；所述沉淀的质量与溶剂的体积比为2g∶3mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后肉豆蔻油。
实施例7：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油；所述具有抗菌性能精油为八角茴香油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为200g；
2、菌种活化：将100g活化液加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母和安琪酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为28℃，摇床转速为200r/min；光照条件：黑暗6h，光照18h，光照强度为2000lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶酿酒酵母活化液∶安琪酵母活化液体积比为1∶1∶1；且聚解脂复膜孢酵母活化液、酿酒酵母活化液和安琪酵母活化液体的总添加量为10％；
4、除味发酵：将300g八角茴香油加入步骤3中装有酵母发酵液的主发酵槽中，混匀后将摇床转速调整至300r/min，温度调整至35℃，然后在摇床转速为300r/min和温度为35℃下发酵时间为48h，得到精油发酵液；
5、分离：对精油发酵液先在0℃下灭活处理24h，然后经200W超声波处理15min，最后采用高速离心机以10000转/min离心处理10min，收集，得到上清液和沉淀；
6、萃取：对步骤5得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为60℃，转速60r/min，得到精油I；所述有机溶剂由乙醇与乙酸乙酯按体积比3∶1混合而成；所述上清液浓缩液与有机溶剂的体积比为1∶5；
在萃取罐中对步骤5得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为60℃，转速60r/min得到精油II；所有溶剂为乙醇；所述沉淀的质量与溶剂的体积比为2g∶3mL；
7、合并：合并步骤6得到的精油I和精油II，即得到去味后八角茴香油。
实施例8：
本实施例与实施例1不同点是：步骤4中在摇床转速为300r/min和温度为35℃下发酵时间为120h，得精油发酵液。其他与实施例1相同。
实施例1得到的去味后肉桂油与实施例8得到的去味后肉桂油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比；
证明实验效果的实验方案和实验结果数据
以实施例1得到的去味后肉桂油与实施例8得到的去味后肉桂油进行抑菌效果的定量试验
供试菌种为：金黄色葡萄球菌、大肠杆菌、白色念珠菌、铜绿假单胞杆菌和黑曲霉。
采用液体培养基稀释法测定肉桂油和实施例1得到的去味后肉桂油的最低抑菌浓度MIC，具体方法如下：
(1)培养基的制备：营养肉汤培养基；
(2)菌悬液的制备：取活化后的金黄色葡萄球菌、活化后的大肠杆菌、活化后的铜绿假单胞杆菌于灭菌蒸馏水中分别制成1×10 8cfu/ml的菌悬液；取活化后的白色念珠菌和活化后的黑曲霉于灭菌蒸馏水中分别制成1×10 7cfu/ml的菌悬液；
(3)含待检测液试管的制备：以实施例1得到的去味后肉桂油为实验组，以实施例 8得到的去味后肉桂油为对照组，
取1g实施例1得到的去味后肉桂油预先溶于9mL DMSO中，得到浓度为1/10的样液，取灭菌试管，吸取1mL浓度为1/10的样液，再加入9mL营养肉汤培养基，得到浓度为1/100的样液，分别吸取0.5mL、1mL和2mL浓度为1/100的样液依次加入9.5mL的营养肉汤培养基、9mL的营养肉汤培养基和8mL的营养肉汤培养基中，配制成浓度分别为0.5‰、1‰和2‰的含检测液的样液，得到实验组样液；
按照上述过程，采用实施例8得到的去味后肉桂油代替实施例1得到的去味后肉桂油进行操作，得到对照组样液。
(4)最低抑菌浓度试验：取上述各菌悬液0.1mL分别加入0.5‰、1‰和2‰浓度试管，设置三组平行试验组，其中细菌的观察置于37℃培养24h，真菌的观察置于28℃培养48h；培养结束后试管分别吸取1mL于平板中，倒入适量培养基，细菌观察置于37℃培养24h，真菌的观察置于28℃培养48h；观察平板菌落生长情况。
实施例1得到的去味后肉桂油与实施例8得到的去味后肉桂油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比结果如表9和表10所示：
表9
Figure PCTCN2020077362-appb-000009
“+”代表有生长“-”代表无生长
表10
Figure PCTCN2020077362-appb-000010
Figure PCTCN2020077362-appb-000011
“+”代表有生长“-”代表无生长
实施例9：
本实施例与实施例1不同点是：步骤3中将聚解脂复膜孢酵母活化液、安琪酵母活化液和酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为250r/min；光照条件：黑暗7h，光照17h，光照强度为2300lux，至总活菌数为10 8cfu/mL为止，得到酵母发酵液；所述聚解脂复膜孢酵母活化液∶安琪酵母活化液体积比为1∶1；且聚解脂复膜孢酵母活化液和安琪酵母活化液体的总添加量为10％。其他与实施例1相同。
实施例1得到的去味后肉桂油与实施例8得到的去味后肉桂油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比；
证明实验效果的实验方案和实验结果数据
以实施例1得到的去味后肉桂油与实施例9得到的去味后肉桂油进行抑菌效果的定量试验
供试菌种为：金黄色葡萄球菌、大肠杆菌、白色念珠菌、铜绿假单胞杆菌和黑曲霉。
采用液体培养基稀释法测定肉桂油和实施例1得到的去味后肉桂油的最低抑菌浓度MIC，具体方法如下：
(1)培养基的制备：营养肉汤培养基；
(2)菌悬液的制备：取活化后的金黄色葡萄球菌、活化后的大肠杆菌、活化后的铜绿假单胞杆菌于灭菌蒸馏水中分别制成1×10 8cfu/ml的菌悬液；取活化后的白色念珠菌和活化后的黑曲霉于灭菌蒸馏水中分别制成1×10 7cfu/ml的菌悬液；
(3)含待检测液试管的制备：以实施例1得到的去味后肉桂油为实验组，以实施例9得到的去味后肉桂油为对照组，
取1g实施例1得到的去味后肉桂油预先溶于9mL DMSO中，得到浓度为1/10的样液，取灭菌试管，吸取1mL浓度为1/10的样液，再加入9mL营养肉汤培养基，得到浓度为1/100的样液，分别吸取0.5mL、1mL和2mL浓度为1/100的样液依次加入9.5mL的营养肉汤培养基、9mL的营养肉汤培养基和8mL的营养肉汤培养基中，配制成浓度分别为0.5‰、1‰和2‰的含检测液的样液，得到实验组样液；
按照上述过程，采用实施例9得到的去味后肉桂油代替实施例1得到的去味后肉桂油进行操作，得到对照组样液。
(4)最低抑菌浓度试验：取上述各菌悬液0.1mL分别加入0.5‰、1‰和2‰浓度试管，设置三组平行试验组，其中细菌的观察置于37℃培养24h，真菌的观察置于28℃培养48h；培养结束后试管分别吸取1mL于平板中，倒入适量培养基，细菌观察置于37℃培养24h，真菌的观察置于28℃培养48h；观察平板菌落生长情况。
实施例1得到的去味后肉桂油与实施例9得到的去味后肉桂油对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌、黑曲霉和白色念珠菌的抑菌活性对比结果如表11和表12：
表11
Figure PCTCN2020077362-appb-000012
“+”代表有生长“-”代表无生长
表12
Figure PCTCN2020077362-appb-000013
“+”代表有生长“-”代表无生长
实施例10：不同酵母对肉桂油祛味发酵比较：
一种具有抗菌性能精油气味去除方法，具体包括以下步骤：利用酵母发酵液处理具有抗菌性能精油；所述具有抗菌性能精油为肉桂油；
所述利用酵母发酵液处理具有抗菌性能精油包含以下步骤：
1、配置酵母培养基：将白砂糖、低聚异麦芽糖、聚葡萄糖、蛋白胨和1000g无菌水加入种菌发酵槽中，完全溶解后将种菌发酵槽转移至高压灭菌锅中，在温度为121℃下灭 菌20min，冷却至室温后，得到初混物，葡萄糖在温度为115℃下灭菌30min，然后加入初混物中，溶解混匀后得到酵母培养基；所述酵母培养基中白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨的质量比为4∶3∶1∶1∶1，且白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨的总质量为200g；
2、菌种活化：将6份100g活化液分别加入前期发酵槽中，然后分别加入聚解脂复膜孢酵母、酿酒酵母、安琪酵母、白葡萄酒专用酵母、18度葡萄酒酵母和降酸酵母进行活化，得到聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液、白葡萄酒专用酵母活化液、18度葡萄酒酵母活化液和降酸酵母活化液；所述活化液中白砂糖的质量分数为5％；
3、富集发酵：将聚解脂复膜孢酵母活化液、酿酒酵母活化液、安琪酵母活化液、白葡萄酒专用酵母活化液、18度葡萄酒酵母活化液和降酸酵母活化液作为酵母活化液，分别与酵母培养基放入主发酵槽中进行摇床发酵，发酵温度为30℃，摇床转速为250r/min；光照条件：黑暗8h，光照16h，光照强度为2000lux，至总活菌数为10 8cfu/mL为止，得到聚解脂复膜孢酵母发酵液、酿酒酵母发酵液、安琪酵母发酵液、白葡萄酒专用酵母发酵液、18度葡萄酒酵母发酵液和降酸酵母发酵液；所述酵母活化液的添加量为10％；
4、除味发酵：将6份肉桂油分别加入步骤3中装有聚解脂复膜孢酵母发酵液、酿酒酵母发酵液、安琪酵母发酵液、白葡萄酒专用酵母发酵液、18度葡萄酒酵母发酵液和降酸酵母发酵液的主发酵槽中，每份肉桂油100g，混匀后将摇床转速调整至300r/min，温度调整至35℃，然后在摇床转速为300r/min和温度为35℃下发酵时间为72h，得到6种精油发酵液；
5、分离：对6种精油发酵液分别先在0℃下灭活处理24h，然后经300W超声波处理35min，最后采用高速离心机以12000转/min离心处理10min，收集，得到6组上清液和沉淀；
6、萃取：依次取步骤5中6组上清液和沉淀中一组进行以下操作
对上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，在萃取罐中利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，采用旋转蒸发仪对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min，得到精油I；所述有机溶剂由乙酸乙酯、甲醇和乙醚混合而成，且有机溶剂中乙酸乙酯∶甲醇∶乙醚的体积比为5∶2∶3；所述上清液浓缩液与有机溶剂的体积比为1∶3；
在萃取罐中对沉淀采用溶剂进行萃取，得到溶剂萃取液，采用旋转蒸发仪对溶剂萃取 液进行浓缩，至溶剂全部旋转蒸发分离为止，旋转蒸发仪水温为50℃，转速45r/min得到精油II；所有溶剂为乙酸乙酯；所述沉淀的质量与溶剂的体积比为1g∶3mL；
7、合并：分别合并步骤6得到的6组精油I和精油II，即得到聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油。
(一)聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油进行气味的变化比较：
感官评价描述数据参照表13。
表13
Figure PCTCN2020077362-appb-000014
聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油的气味评价结果如表14所示。
表14
Figure PCTCN2020077362-appb-000015
(二)肉桂油、聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油对大肠埃希氏菌、铜绿假单胞菌和黑曲霉的抑菌活性对比；
证明实验效果的实验方案和实验结果数据
以肉桂油、聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油进行抑菌效果的定量试验。
供试菌种为：大肠杆菌、铜绿假单胞杆菌和黑曲霉。
采用液体培养基稀释法测定肉桂油、聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油的最低抑菌浓度MIC，具体方法如下：
(1)培养基的制备：营养肉汤培养基；
(2)菌悬液的制备：取活化后的大肠杆菌和活化后的铜绿假单胞杆菌于灭菌蒸馏水中分别制成1×10 8cfu/ml的菌悬液；取活化后的黑曲霉于灭菌蒸馏水中分别制成1×10 7cfu/ml的菌悬液；
(3)含待检测液试管的制备：以实施例1得到的去味后肉桂油为实验组，以肉桂油为对照组，
分别取1g肉桂油、1g聚解脂复膜孢酵母去味后肉桂油、1g酿酒酵母去味后肉桂油、1g安琪酵母去味后肉桂油、1g白葡萄酒专用酵母去味后肉桂油、1g 18度葡萄酒酵母去味后肉桂油和1g降酸酵母去味后肉桂油预先溶于9mL DMSO中，得到浓度为1/10的样液，取灭菌试管，吸取1mL浓度为1/10的样液，再加入9mL营养肉汤培养基，得到浓度为1/100的样液，分别吸取1mL浓度为1/100的样液加入9mL的营养肉汤培养基中，配制成浓度为1‰的含检测液的样液，即得到肉桂油样液、聚解脂复膜孢酵母去味后肉桂油样液、酿酒酵母去味后肉桂油样液、安琪酵母去味后肉桂油样液、白葡萄酒专用酵母去味后肉桂油样液、18度葡萄酒酵母去味后肉桂油样液和降酸酵母去味后肉桂油样液；
(4)最低抑菌浓度试验：取上述各菌悬液0.1mL加入1‰浓度试管，设置三组平行试验组，其中细菌的观察置于37℃培养24h，真菌的观察置于28℃培养48h；培养结束后试管分别吸取1mL于平板中，倒入适量培养基，细菌观察置于37℃培养24h，真菌的观察置于28℃培养48h；观察平板菌落生长情况。
肉桂油、聚解脂复膜孢酵母去味后肉桂油、酿酒酵母去味后肉桂油、安琪酵母去味后肉桂油、白葡萄酒专用酵母去味后肉桂油、18度葡萄酒酵母去味后肉桂油和降酸酵母去味后肉桂油对大肠埃希氏菌、铜绿假单胞菌和黑曲霉的抑菌活性对比结果如表15所示：
表15
Figure PCTCN2020077362-appb-000016
Figure PCTCN2020077362-appb-000017
“+”代表有生长“-”代表无生长
由表15结果在平衡酵母对肉桂味祛除和保留肉桂油原有抑菌活性效能考虑，得出聚解脂复膜孢酵母、安琪酵母和酿酒酵母相对适合应用于肉桂油祛味应用上。
实施例11：不同有机试剂对肉桂油祛味发酵比较：
本实施例与实施例1的不同点是：步骤6中分别采用乙酸乙酯、乙醇、乙醚、环己烷、苯和丙酮作为有机溶剂。其他与实施例1相同。
采用乙酸乙酯精油I、乙醇精油I、乙醚精油I、环己烷精油I、苯精油I、丙酮精油I和实施例1得到的精油I的对大肠埃希氏菌、铜绿假单胞菌和黑曲霉的抑菌活性对比；
证明实验效果的实验方案和实验结果数据
以乙酸乙酯精油I、乙醇精油I、乙醚精油I、环己烷精油I、苯精油I、丙酮精油I和实施例1得到的精油I进行抑菌效果的定量试验。
供试菌种为：大肠杆菌、铜绿假单胞杆菌和黑曲霉。
采用液体培养基稀释法测定乙酸乙酯精油I、乙醇精油I、乙醚精油I、环己烷精油I、苯精油I、丙酮精油I和实施例1得到的精油I的最低抑菌浓度MIC，具体方法如下：
(1)培养基的制备：营养肉汤培养基；
(2)菌悬液的制备：取活化后的大肠杆菌和活化后的铜绿假单胞杆菌于灭菌蒸馏水中分别制成1×10 8cfu/ml的菌悬液；取活化后的黑曲霉于灭菌蒸馏水中分别制成1×10 7cfu/ml的菌悬液；
(3)含待检测液试管的制备：以实施例1得到的去味后肉桂油为实验组，以肉桂油 为对照组，
分别取1g乙酸乙酯精油I、1g乙醇精油I、1g乙醚精油I、1g环己烷精油I、1g苯精油I、1g丙酮精油I和1g实施例1得到的精油I预先溶于9mL DMSO中，得到浓度为1/10的样液，取灭菌试管，吸取1mL浓度为1/10的样液，再加入9mL营养肉汤培养基，得到浓度为1/100的样液，分别吸取1mL浓度为1/100的样液加入9mL的营养肉汤培养基中，配制成浓度为1‰的含检测液的样液，即得到乙酸乙酯精油I样液、乙醇精油I样液、乙醚精油I样液、环己烷精油I样液、苯精油I样液、丙酮精油I样液和实施例1得到的精油I样液；
(4)最低抑菌浓度试验：取上述各菌悬液0.1mL加入1‰浓度试管，设置三组平行试验组，其中细菌的观察置于37℃培养24h，真菌的观察置于28℃培养48h；培养结束后试管分别吸取1mL于平板中，倒入适量培养基，细菌观察置于37℃培养24h，真菌的观察置于28℃培养48h；观察平板菌落生长情况。
乙酸乙酯精油I、乙醇精油I、乙醚精油I、环己烷精油I、苯精油I、丙酮精油I和实施例1得到的精油I的抑菌活性对比结果如表16所示：
表16
Figure PCTCN2020077362-appb-000018
由表16结果来看，不同有机试剂对处理后的肉桂油萃取物的活性有一定影响，最后选用乙酸乙酯作为有机溶剂，或选择乙酸乙酯与乙醇的混合物作为有机溶剂。

Claims (10)

  1. 一种具有抗菌性能精油气味去除方法，其特征在于利用酵母发酵液处理具有抗菌性能精油。
  2. 根据权利要求1所述的一种具有抗菌性能精油气味去除方法，其特征在于所述所述酵母发酵液中酵母的活菌数为10 7cfu/mL～10 9cfu/mL。
  3. 根据权利要求2所述的一种具有抗菌性能精油气味去除方法，其特征在于所述所述酵母发酵液中的酵母为聚解脂复膜孢酵母、降酸酵母、酿酒酵母、18度葡萄酒酵母、安琪酵母和白葡萄酒专用酵母中的一种或其中几种。
  4. 根据权利要求1所述的一种具有抗菌性能精油气味去除方法，其特征在于所述具有抗菌性能精油为单一精油或复配精油。
  5. 根据权利要求4所述的一种具有抗菌性能精油气味去除方法，其特征在于所述单一精油为肉桂油、丁香精油、迷迭香精油、大蒜油、广霍香油、肉豆蔻油、八角茴香油、山苍子精油、南丰蜜桔精油、飞蓬精油或胜红蓟精油；所述复配精油由肉桂油、丁香精油、迷迭香精油、广霍香油、肉豆蔻油和八角茴香油中的两种或两种以上混合而成。
  6. 根据权利要求1-5中任一项所述的一种具有抗菌性能精油气味去除方法，其特征在于包含以下步骤：
    (1)、发酵：将具有抗菌性能精油加入酵母发酵液中，混匀后发酵处理，得精油发酵液；所述具有抗菌性能精油与酵母发酵液的体积比为6～10∶1；
    (2)、分离：对精油发酵液依次进行灭活处理、超声波处理和离心处理，得到上清液和沉淀；
    (3)、萃取：对步骤(2)得到的上清液依次进行浓缩、萃取和再次浓缩，得到精油I；对步骤(2)得到的沉淀依次进行萃取和浓缩，得到精油II；
    (4)、合并：合并步骤(3)得到的精油I和精油II，即得到去除气味后具有抗菌性能精油。
  7. 根据权利要求6所述的一种具有抗菌性能精油气味去除方法，其特征在于步骤(1)中所述的酵母发酵液是按以下步骤获得：
    ①、配置酵母培养基：将复合营养物加入无菌水中，复合营养物完全溶解后转移至高压灭菌锅中，在温度为115℃～121℃下灭菌20min～30min，冷却至室温后，得到酵母培养基；所述复合营养物与无菌水的质量比为3∶10～90；所述复合营养物由白砂糖、葡萄糖、低聚异麦芽糖、聚葡萄糖和蛋白胨组成，且复合营养物中按质量比白砂糖∶葡萄糖∶低聚异麦芽糖∶聚葡萄糖∶蛋白胨＝4∶3∶1∶1∶1；
    ②、菌种活化：将酵母放入活化液中活化，得到含活化后酵母的酵母活化液；所述活化液中白砂糖的质量分数为5～15％；
    ③、发酵：将含活化后酵母的酵母活化液放入酵母培养基进行摇床发酵，发酵温度为24～35℃，摇床转速为150r/min～300r/min；光照条件：黑暗6h～8h，光照16h～18h，光照强度为2000lux～2500lux，至酵母的活菌数为10 7cfu/mL～10 9cfu/mL为止，得到酵母发酵液。
  8. 根据权利要求6所述的一种具有抗菌性能精油气味去除方法，其特征在于步骤(1)中所述发酵处理具体操作过程如下：
    将摇床转速调整至150r/min～300r/min，温度调整至30～65℃，然后在摇床转速为150r/min～300r/min和温度为30～65℃下发酵时间为72h～120h。
  9. 根据权利要求6所述的一种具有抗菌性能精油气味去除方法，其特征在于步骤(2)中将精油发酵液在0℃下灭活处理24h，然后经250W～300W超声波处理15min～60min，最后12000转/min离心处理10min～25min，收集，得到上清液和沉淀。
  10. 根据权利要求6所述的一种具有抗菌性能精油气味去除方法，其特征在于步骤(3)中对步骤(2)得到的上清液依次进行浓缩，浓缩至无水分为止，得到上清液浓缩液，利用有机溶剂对上清液浓缩液进行萃取，得到有机溶剂萃取物，对有机溶剂萃取物进行再次浓缩，至有机溶剂全部旋转蒸发分离为止，得到精油I；所述有机溶剂选自乙酸乙酯、乙醇、乙醚和环己烷；所述上清液浓缩液与有机溶剂的体积比为1～3∶3～6；
    对步骤(2)得到的沉淀采用溶剂进行萃取，得到溶剂萃取液，对溶剂萃取液进行浓缩，至溶剂全部旋转蒸发分离为止，得到精油II；所有溶剂选自乙酸乙酯、乙醇、乙醚和环己烷；所述沉淀的质量与溶剂的体积比为1g～4g∶2mL～6mL。
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