CN109077943B - Preparation method and application of octacosanol-containing liposome - Google Patents
Preparation method and application of octacosanol-containing liposome Download PDFInfo
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- CN109077943B CN109077943B CN201811056577.9A CN201811056577A CN109077943B CN 109077943 B CN109077943 B CN 109077943B CN 201811056577 A CN201811056577 A CN 201811056577A CN 109077943 B CN109077943 B CN 109077943B
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- octacosanol
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- taurine
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- dichloromethane
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- CNNRPFQICPFDPO-UHFFFAOYSA-N octacosan-1-ol Chemical compound CCCCCCCCCCCCCCCCCCCCCCCCCCCCO CNNRPFQICPFDPO-UHFFFAOYSA-N 0.000 title claims abstract description 114
- 239000002502 liposome Substances 0.000 title claims abstract description 69
- 229960002666 1-octacosanol Drugs 0.000 title claims abstract description 64
- 238000002360 preparation method Methods 0.000 title claims abstract description 11
- XOAAWQZATWQOTB-UHFFFAOYSA-N taurine Chemical compound NCCS(O)(=O)=O XOAAWQZATWQOTB-UHFFFAOYSA-N 0.000 claims abstract description 71
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 claims abstract description 42
- 229960003080 taurine Drugs 0.000 claims abstract description 42
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims abstract description 33
- 239000000243 solution Substances 0.000 claims abstract description 33
- 239000007864 aqueous solution Substances 0.000 claims abstract description 30
- 235000012000 cholesterol Nutrition 0.000 claims abstract description 21
- YDVNLQGCLLPHAH-UHFFFAOYSA-N dichloromethane;hydrate Chemical compound O.ClCCl YDVNLQGCLLPHAH-UHFFFAOYSA-N 0.000 claims abstract description 17
- 239000000839 emulsion Substances 0.000 claims abstract description 17
- 239000002245 particle Substances 0.000 claims abstract description 15
- -1 taurine octacosanol ester Chemical class 0.000 claims abstract description 14
- 238000004945 emulsification Methods 0.000 claims abstract description 11
- 238000004108 freeze drying Methods 0.000 claims abstract description 11
- 230000000887 hydrating effect Effects 0.000 claims abstract description 10
- 235000013361 beverage Nutrition 0.000 claims abstract description 6
- 239000002537 cosmetic Substances 0.000 claims abstract description 6
- 238000002390 rotary evaporation Methods 0.000 claims abstract description 6
- 235000013305 food Nutrition 0.000 claims abstract description 5
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 30
- 238000002156 mixing Methods 0.000 claims description 18
- 238000000034 method Methods 0.000 claims description 12
- 230000000694 effects Effects 0.000 claims description 11
- 102000005262 Sulfatase Human genes 0.000 claims description 9
- 239000000843 powder Substances 0.000 claims description 9
- 108060007951 sulfatase Proteins 0.000 claims description 9
- 238000006243 chemical reaction Methods 0.000 claims description 8
- 102000004169 proteins and genes Human genes 0.000 claims description 6
- 108090000623 proteins and genes Proteins 0.000 claims description 6
- 239000004372 Polyvinyl alcohol Substances 0.000 claims description 5
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 5
- 238000003756 stirring Methods 0.000 claims description 5
- 230000036571 hydration Effects 0.000 claims description 4
- 238000006703 hydration reaction Methods 0.000 claims description 4
- 230000035484 reaction time Effects 0.000 claims description 3
- 239000000654 additive Substances 0.000 claims description 2
- 230000000996 additive effect Effects 0.000 claims description 2
- 239000003054 catalyst Substances 0.000 claims description 2
- 238000007036 catalytic synthesis reaction Methods 0.000 claims description 2
- 230000002194 synthesizing effect Effects 0.000 claims description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 11
- 150000002191 fatty alcohols Chemical class 0.000 abstract description 4
- 239000000796 flavoring agent Substances 0.000 abstract description 3
- 235000019634 flavors Nutrition 0.000 abstract description 3
- 238000010521 absorption reaction Methods 0.000 abstract description 2
- 230000009286 beneficial effect Effects 0.000 abstract description 2
- 230000015572 biosynthetic process Effects 0.000 abstract description 2
- 230000004087 circulation Effects 0.000 abstract description 2
- 230000000295 complement effect Effects 0.000 abstract description 2
- 230000002195 synergetic effect Effects 0.000 abstract description 2
- 238000003786 synthesis reaction Methods 0.000 abstract description 2
- 239000012467 final product Substances 0.000 abstract 1
- 230000000052 comparative effect Effects 0.000 description 7
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 241000282414 Homo sapiens Species 0.000 description 3
- 150000001413 amino acids Chemical class 0.000 description 3
- 230000001804 emulsifying effect Effects 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 2
- 238000005034 decoration Methods 0.000 description 2
- 230000002708 enhancing effect Effects 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 235000016709 nutrition Nutrition 0.000 description 2
- 230000001766 physiological effect Effects 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 229910052717 sulfur Inorganic materials 0.000 description 2
- 239000011593 sulfur Substances 0.000 description 2
- 238000002834 transmittance Methods 0.000 description 2
- 208000037147 Hypercalcaemia Diseases 0.000 description 1
- 208000035150 Hypercholesterolemia Diseases 0.000 description 1
- 208000031226 Hyperlipidaemia Diseases 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 208000000112 Myalgia Diseases 0.000 description 1
- 208000001132 Osteoporosis Diseases 0.000 description 1
- 206010037660 Pyrexia Diseases 0.000 description 1
- WBWWGRHZICKQGZ-UHFFFAOYSA-N Taurocholic acid Natural products OC1CC2CC(O)CCC2(C)C2C1C1CCC(C(CCC(=O)NCCS(O)(=O)=O)C)C1(C)C(O)C2 WBWWGRHZICKQGZ-UHFFFAOYSA-N 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 229940088623 biologically active substance Drugs 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 230000004641 brain development Effects 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000001125 extrusion Methods 0.000 description 1
- 230000004438 eyesight Effects 0.000 description 1
- 239000002778 food additive Substances 0.000 description 1
- 235000013373 food additive Nutrition 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 210000000232 gallbladder Anatomy 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 230000000148 hypercalcaemia Effects 0.000 description 1
- 208000030915 hypercalcemia disease Diseases 0.000 description 1
- 208000020346 hyperlipoproteinemia Diseases 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000037323 metabolic rate Effects 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 208000013465 muscle pain Diseases 0.000 description 1
- 230000010016 myocardial function Effects 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 230000009329 sexual behaviour Effects 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
- 230000008326 skin blood flow Effects 0.000 description 1
- 208000017520 skin disease Diseases 0.000 description 1
- 238000002798 spectrophotometry method Methods 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000035488 systolic blood pressure Effects 0.000 description 1
- WBWWGRHZICKQGZ-GIHLXUJPSA-N taurocholic acid Chemical compound C([C@@H]1C[C@H]2O)[C@@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@@H]([C@@H](CCC(=O)NCCS(O)(=O)=O)C)[C@@]2(C)[C@H](O)C1 WBWWGRHZICKQGZ-GIHLXUJPSA-N 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/46—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing sulfur
- A61K8/466—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing sulfur containing sulfonic acid derivatives; Salts
-
- 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
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
-
- 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
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/175—Amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/02—Cosmetics or similar toiletry preparations characterised by special physical form
- A61K8/14—Liposomes; Vesicles
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/33—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing oxygen
- A61K8/34—Alcohols
- A61K8/342—Alcohols having more than seven atoms in an unbroken chain
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/10—General cosmetic use
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
- A61K2800/59—Mixtures
- A61K2800/592—Mixtures of compounds complementing their respective functions
Abstract
The invention discloses a preparation method of octacosanol-containing liposome, which comprises the steps of adding octacosanol and taurine into a water-dichloromethane emulsion to catalytically synthesize a taurine octacosanol ester solution; adding a cholesterol aqueous solution into the taurine octacosanol ester solution obtained in the step one for ultrasonic emulsification, and then performing rotary evaporation at room temperature to remove dichloromethane to obtain a liposome solution; hydrating the liposome solution, preparing octacosanol-containing liposome with uniform particle size by a liposome extruder, and freeze-drying to obtain the final product. The octacosanol-containing liposome is prepared by utilizing octacosanol and taurine through synthesis, so that the problem of poor water solubility of higher fatty alcohol is solved, the problem of no flavor and appearance is caused when the octacosanol-containing liposome is added into food, cosmetics and beverages, and the homogeneity is good; meanwhile, taurine and octacosanol have complementary and synergistic effects on organism regulation, and are beneficial to absorption and internal circulation.
Description
Technical Field
The invention relates to a preparation method and application of octacosanol-containing liposome, belonging to the field of biological engineering.
Background
Taurine (Taurine), also known as taurocholic acid, is a sulfur-containing non-protein amino acid with a chemical name of 2-aminoethanesulfonic acid, is a sulfur-containing amino acid with multiple physiological functions existing in a human body, has the effects of promoting brain development, enhancing eyesight, resisting inflammation, relieving fever, reducing blood pressure, reducing blood sugar, strengthening liver, benefiting gallbladder and the like, and is one of the most important amino acids in the human body. Taurine has no toxic side effect, so developed countries pay attention to the research and application of taurine. In recent years, as the physiological effects and nutritional values of taurine have been studied in depth, the taurine has been widely used as a nutritional health product and a food additive in large quantities abroad, and it is reported that the taurine is the most important consumer countries in the united states and japan, and the annual consumption amounts are ten thousand tons and more than five thousand tons, respectively.
Octacosanol is a biologically active substance whose major physiological effects include: 1) improving endurance, energy and physical strength; 2) shortening the reaction time of the muscle nerve and improving the reaction sensitivity; 3) promoting hormone function, and relieving muscle pain; 4) improving myocardial function, and reducing systolic blood pressure; 5) improving basal metabolic rate of organism. Octacosanol can be used for treating hypercalcemia osteoporosis, hypercholesterolemia and hyperlipoproteinemia, and stimulating sexual behavior of animals and human beings, and cosmetic containing octacosanol can promote skin blood circulation and activate cells, and has antiinflammatory and skin disease preventing and treating effects. And the octacosanol has extremely high safety and small dosage. And the octacosanol and the taurine are jointly acted on the body, do not generate any toxicity or mutation-causing effect and can jointly play the health care effect.
The addition of higher fatty alcohols to beverages can affect their flavor and appearance due to their poor water solubility. Therefore, appropriate modification is required to increase the solubility.
Disclosure of Invention
The invention aims to solve the technical problem of providing a preparation method of octacosanol-containing liposome, which is used for enhancing the solubility of octacosanol and achieving the health-care effect by combining with taurine.
The invention also aims to solve the technical problem of providing the application of the octacosanol-containing liposome.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows:
a preparation method of octacosanol-containing liposome comprises the following steps:
the method comprises the following steps: adding octacosanol and taurine into a water-dichloromethane emulsion, and catalytically synthesizing a taurine octacosanol ester (TO) solution by taking sulfatase as a catalyst;
step two: adding a cholesterol aqueous solution into the taurine octacosanol ester (TO) solution obtained in the step one for ultrasonic emulsification, and then performing rotary evaporation at room temperature TO remove dichloromethane TO obtain a liposome solution;
step three: hydrating the liposome solution, preparing octacosanol-containing liposome (TO liposome) with uniform particle size by a liposome extruder, and freeze-drying TO obtain powder.
Further, in step one, the water-dichloromethane emulsion is prepared by the following method: mixing an aqueous solution containing 1-2 wt% (preferably 1.5 wt%) of polyvinyl alcohol with dichloromethane at a volume ratio of 1: 1-2: 1 (preferably 1:1), and stirring at a high speed of 8000-10000 r/min to obtain a water-dichloromethane emulsion.
The mass ratio of the octacosanol to the taurine is 3: 1-4: 1 (preferably 3.3:1), and the total added mass of the octacosanol and the taurine is 5-10 mg/ml (preferably 8.6mg/ml) of water-dichloromethane emulsion.
The specific activity of the sulfatase is 1000-5000U/mg protein, and the addition amount of the sulfatase is 20-50 mg/ml (preferably 50mg/ml) of water-dichloromethane emulsion.
The reaction temperature of catalytic synthesis is 30-40 ℃ (preferably 37 ℃), the reaction time is 4-8 h (preferably 6h), standing and layering are carried out after the reaction is finished, and the lower layer is a taurine octacosanol ester solution (TO solution), wherein the structural formula of the taurine octacosanol ester is as follows:
in the second step, the concentration of the cholesterol aqueous solution is 5-10 mg/ml (preferably 8mg/ml), and the volume ratio of the cholesterol aqueous solution to the taurine octacosanol ester solution is 1: 4-1: 2 (preferably 1: 2.5); the ultrasonic emulsification power is 200-300W (preferably 250W), and the emulsification time is 10-30 min (preferably 20 min).
In the third step, the hydration method comprises the following steps: mixing and hydrating the liposome solution and 20-40 g/L (preferably 30g/L) of citric acid aqueous solution for 2-4 h (preferably 3h), wherein the mixing volume ratio of the liposome solution to the citric acid aqueous solution is 5: 1-2: 1 (preferably 3: 1); the freeze-drying temperature is-50 to-55 ℃ (preferably-50 ℃), the vacuum degree is 0.006 to 0.012mBar (preferably 0.008mBar), and the freeze-drying time is 72 to 96 hours (preferably 80 hours).
The octacosanol-containing liposome (TO liposome) prepared by the preparation method is also within the protection scope of the invention, and the particle size of the liposome is 100-400 nm, preferably 200 nm.
The invention also claims application of the octacosanol-containing liposome (TO liposome) as an additive TO food, beverage or cosmetics, wherein the liposome is prepared into 100-500 mg/L (preferably 200mg/L) aqueous solution TO be added TO the food, the cosmetic or the beverage.
Has the advantages that:
the octacosanol-containing liposome is prepared by utilizing octacosanol and taurine through synthesis, so that the problem of poor water solubility of higher fatty alcohol is solved, the problem of no flavor and appearance is caused when the octacosanol-containing liposome is added into food, cosmetics and beverages, and the homogeneity is good; meanwhile, taurine and octacosanol have complementary and synergistic effects on organism regulation, and are beneficial to absorption and internal circulation.
Detailed Description
The invention will be better understood from the following examples.
Example 1
1) Mixing 5ml of aqueous solution containing 1.5 wt% of polyvinyl alcohol (purchased from Allantin reagent Co., Ltd., product No. P119363) with 5ml of dichloromethane, and stirring at a high speed of 8000r/min to obtain water-dichloromethane emulsion;
2) adding 66mg of octacosanol and 20mg of taurine into the prepared water-dichloromethane emulsion, adding 500mg of sulfatase (the specific activity of the enzyme is 5000U/mg of protein), reacting at 37 ℃ for 6 hours, standing for layering after the reaction is finished, and taking the lower layer TO obtain a taurine octacosanol ester solution (TO solution);
3) adding 40mg of cholesterol into 2ml of water TO prepare a cholesterol aqueous solution, then mixing the cholesterol aqueous solution with a TO solution, carrying out ultrasonic emulsification with the power of 250W, emulsifying for 20min, carrying out rotary evaporation at room temperature TO remove dichloromethane, then adding 0.6ml of 30g/L citric acid aqueous solution, uniformly mixing, hydrating for 3h, then preparing a TO liposome with the average particle size of 200nm by using a liposome extruder, finally putting the TO liposome into a freeze dryer, and freeze-drying for 80h at the temperature of-50 ℃ and 0.008mBar TO obtain the TO liposome freeze-dried powder.
Example 2
1) Mixing 10ml of aqueous solution containing 1.0 wt% of polyvinyl alcohol (purchased from Allantin reagent Co., Ltd., product No. P119363) with 5ml of dichloromethane, and stirring at high speed of 10000r/min to obtain water-dichloromethane emulsion;
2) adding 60mg of octacosanol and 15mg of taurine into the prepared water-dichloromethane emulsion, adding 300mg of sulfatase (the specific activity of the enzyme is 1000U/mg of protein), reacting for 4 hours at 30 ℃, standing for layering after the reaction is finished, and taking the lower layer TO obtain a taurine octacosanol ester solution (TO solution);
3) adding 25mg of cholesterol into 2.5ml of water TO prepare a cholesterol aqueous solution, then mixing the cholesterol aqueous solution with a TO solution, carrying out ultrasonic emulsification at the power of 200W for 30min, carrying out rotary evaporation at room temperature TO remove dichloromethane, then adding 0.5ml of 20g/L citric acid aqueous solution, uniformly mixing, hydrating for 2h, then preparing a TO liposome with the average particle size of 100nm by using a liposome extruder, finally putting the TO liposome into a freeze dryer, and freeze-drying for 72h at the temperature of-55 ℃ and 0.006mBar TO obtain the TO liposome freeze-dried powder.
Example 3
1) Mixing 5ml of aqueous solution containing 2.0 wt% polyvinyl alcohol (available from Allantin reagent Co., Ltd., product No. P119363) with 5ml of methylene chloride, and stirring at 9000r/min to obtain water-methylene chloride emulsion;
2) adding 75mg of octacosanol and 25mg of taurine into the prepared water-dichloromethane emulsion, adding 400mg of sulfatase (the specific activity of the enzyme is 4000U/mg of protein), reacting for 8 hours at 40 ℃, standing for layering after the reaction is finished, and taking the lower layer TO obtain a taurine octacosanol ester solution (TO solution);
3) adding 50mg of cholesterol into 1.25ml of water TO prepare a cholesterol aqueous solution, then mixing the cholesterol aqueous solution with a TO solution, carrying out ultrasonic emulsification at the power of 300W for 10min, carrying out rotary evaporation at room temperature TO remove dichloromethane, then adding 0.625ml of 40g/L citric acid aqueous solution, uniformly mixing, hydrating for 4h, then preparing TO liposome with the average particle size of 400nm by using a liposome extruder, finally putting the TO liposome into a freeze dryer, and freeze-drying for 96h at the temperature of-52 ℃ and 0.012mBar TO obtain the TO liposome freeze-dried powder.
Comparative example 1
Adding 66mg of octacosanol and 40mg of cholesterol into 2ml of water, carrying out ultrasonic emulsification at the power of 250W, emulsifying for 20min, adding 0.6ml of 30g/L citric acid aqueous solution, mixing uniformly, hydrating for 3h, preparing liposome with the average particle size of 200nm by using a liposome extruder, finally putting into a freeze dryer, and freeze-drying for 80h at the temperature of-50 ℃ under the condition of 0.008mBar to obtain the octacosanol cholesterol freeze-dried powder.
Comparative example 2
Adding 20mg of taurine and 40mg of cholesterol into 2ml of water, carrying out ultrasonic emulsification at the power of 250W for 20min, adding 0.6ml of 30g/L citric acid aqueous solution, mixing uniformly, hydrating for 3h, preparing liposome with the average particle size of 200nm by using a liposome extruder, finally putting the liposome into a freeze dryer, and freeze-drying for 80h at the temperature of 50 ℃ below zero and 0.008mBar to obtain the taurine-cholesterol freeze-dried powder.
Comparative example 3
Adding 66mg of octacosanol, 20mg of taurine and 40mg of cholesterol into 2ml of water, carrying out ultrasonic emulsification at the power of 250W, emulsifying for 20min, adding 0.6ml of 30g/L citric acid aqueous solution, mixing uniformly, hydrating for 3h, preparing liposome with the average particle size of 200nm by using a liposome extruder, finally putting the liposome into a freeze dryer, and freeze-drying at the temperature of-50 ℃ for 80h by 0.008mBar to obtain the liposome freeze-dried powder containing taurine and octacosanol ester.
The liposome freeze-dried powders prepared in examples 1 to 3 and comparative examples 1 to 3 are respectively prepared into 200mg/L aqueous solutions, and then particle size and polydispersity tests and clarity tests are carried out, and the results are shown in Table 1.
Particle size and polydispersity measurements among these, the particle size and polydispersity of the solute in the solution was determined using Zetasizer Nano S90.
The clarity test adopts a spectrophotometry method, uses distilled water as a reference, and uses a 1cm cuvette to measure the light transmittance at the wavelength of 680nm, thereby representing the clarity of the sample.
TABLE 1
| Particle size (nm) | Polydispersity | Light transmittance (%) | |
| Example 1 | 202±8 | 0.112±0.019 | 99.89±0.08 |
| Example 2 | 105±12 | 0.221±0.034 | 95.97±0.15 |
| Example 3 | 403±32 | 0.249±0.025 | 94.58±1.95 |
| Comparative example 1 | 502±332 | 0.674±0.125 | 58.23±12.43 |
| Comparative example 2 | 492±323 | 0.853±0.048 | 45.68±14.68 |
| Comparative example 3 | 574±394 | 0.838±0.084 | 37.56±17.94 |
As can be seen from the data in Table 1, octacosanol and taurine are adopted TO synthesize a TO solution, and then the TO solution is ultrasonically emulsified with cholesterol, and then the octacosanol-containing liposome (TO liposome) obtained through hydration and liposome extrusion can effectively solve the problem of poor water solubility of higher fatty alcohol and has remarkable progress.
The invention provides a preparation method of octacosanol-containing liposome, and a thought and a method for application thereof, and a method and a way for realizing the technical scheme are many, the above description is only a preferred embodiment of the invention, and it should be noted that, for a person skilled in the art, a plurality of improvements and decorations can be made without departing from the principle of the invention, and the improvements and decorations are also regarded as the protection scope of the invention. All the components not specified in the present embodiment can be realized by the prior art.
Claims (8)
1. A preparation method of octacosanol-containing liposome is characterized by comprising the following steps:
the method comprises the following steps: adding octacosanol and taurine into a water-dichloromethane emulsion, and catalytically synthesizing a taurine octacosanol ester solution by taking sulfatase as a catalyst;
step two: adding a cholesterol aqueous solution into the taurine octacosanol ester solution obtained in the step one for ultrasonic emulsification, and then performing rotary evaporation at room temperature to remove dichloromethane to obtain a liposome solution;
step three: hydrating the liposome solution, preparing octacosanol-containing liposomes with uniform particle size by a liposome extruder, and freeze-drying to obtain powder;
in the first step, the weight ratio of octacosanol to taurine is 3: 1-4: 1, and the total added weight of octacosanol and taurine is 5-10 mg/ml of water-dichloromethane emulsion.
2. The method for preparing octacosanol-containing liposomes according to claim 1, wherein in the first step, the water-dichloromethane emulsion is prepared by the following method: mixing an aqueous solution containing 1-2 wt% of polyvinyl alcohol with dichloromethane according to a volume ratio of 1: 1-2: 1, and stirring at a high speed of 8000-10000 r/min to prepare a water-dichloromethane emulsion.
3. The method as claimed in claim 1, wherein in the step one, the specific activity of the sulfatase is 1000-5000U/mg protein, and the amount of the added sulfatase is 20-50 mg/ml water-dichloromethane emulsion.
4. The preparation method of octacosanol-containing liposomes according to claim 1, wherein in the first step, the reaction temperature of catalytic synthesis is 30-40 ℃, the reaction time is 4-8 h, and after the reaction is finished, the liposomes are allowed to stand for layering, and the lower layer is the taurine octacosanol ester solution.
5. The method for preparing octacosanol-containing liposomes according to claim 1, wherein in the second step, the concentration of the cholesterol aqueous solution is 5 to 10mg/ml, and the volume ratio of the cholesterol aqueous solution to the octacosanol taurine solution is 1:4 to 1:2.
6. The method for preparing octacosanol-containing liposome according to claim 1, wherein the hydration method in the third step is as follows: mixing the liposome solution and 20-40 g/L citric acid aqueous solution for hydration for 2-4 h, wherein the mixing volume ratio of the liposome solution to the citric acid aqueous solution is 5: 1-2: 1.
7. The octacosanol-containing liposome prepared by the preparation method of any one of claims 1 to 6, wherein the liposome has a particle size of 100 to 400 nm.
8. Use of the octacosanol-containing liposomes of claim 7 as an additive in food, beverages or cosmetics.
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