CN108030088A - A kind of preparation method of protein modified phytosterol liposome powder - Google Patents

A kind of preparation method of protein modified phytosterol liposome powder Download PDF

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CN108030088A
CN108030088A CN201711020091.5A CN201711020091A CN108030088A CN 108030088 A CN108030088 A CN 108030088A CN 201711020091 A CN201711020091 A CN 201711020091A CN 108030088 A CN108030088 A CN 108030088A
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phytosterol
liposome
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protein modified
lecithin
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CN108030088B (en
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王春维
祝爱侠
程铭
焦文佳
夏廉臣
王晓晖
陈浩
王铕
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Wuhan Polytechnic University
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    • A61K31/575—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of three or more carbon atoms, e.g. cholane, cholestane, ergosterol, sitosterol
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Abstract

The invention discloses a kind of preparation method of protein modified phytosterol liposome powder, solves the problems, such as phytosterol poorly water-soluble, the blood fat reducing function of phytosterol needs to be further improved.Phytosterol and lecithin are added in absolute ethyl alcohol by this method, dissolving are stirred at room temperature, up to organic phase；Soybean protein isolate is soluble in water, obtains water phase, and organic phase is slowly injected into water mutually under the conditions of high speed dispersion and high speed dispersion mixes, dry after high-pressure homogeneous, rotary evaporation removes ethanol to obtain through protein modified phytosterol liposome powder.Present invention process is simple, it is time-consuming few, selected materials safety environmental protection, gained liposome stability is good, and embedding rate is high, significantly improve the water solubility of phytosterol, solubility is good after liposome drying, and solubility is high, can preserve for a long time, it can be widely applied in food and medicine, to reduce the generation of hyperlipemia.

Description

A kind of preparation method of protein modified phytosterol liposome powder
Technical field
The present invention relates to a kind of food additives field, specifically a kind of protein modified phytosterol liposome powder Preparation method.
Background technology
Phytosterol, is the functional lipids with class cholesterol structure being present in plant, is steroid in plant The general designation of alcohol compound.Phytosterol is white crystalline powder or particle at normal temperatures, odorless, tasteless, fusing point up to 130~ 170 DEG C, not soluble in water, strong acid and highly basic dissolve in a variety of organic solvents, such as acetone, absolute ethyl alcohol, ether, benzene, chloroform, stone Oily ether etc..
Nineteen fifty-three, Pollak discoveries can treat high-cholesterol disease by taking in phytosterol, plant opened from this The research that sterol is applied in norcholesterol.The discoveries such as Lees daily intake content of phytosterol and reach 3g with regard to that can reach best drop Cholesterol effect, can reduce about 12% cholesterol, and is tested and confirmed by follow-up clinical.Plant steroid after emulsification More preferably, Richard has found that 1g phytosterols its blood fat reducing function after the emulsification of 300mg lecithin is swum for 1g to alcohol norcholesterol effect From as many as 3 times of phytosterol, 36.7% can reach.
At present, the plant sterol product occurred both at home and abroad is varied, it is contemplated that natural phytosterin dissolubility is poor, fusing point The problems such as height, poor taste, often to its it is modified be then added in food, but modifying process often take, effort, and conversion ratio compared with Low, production cost is high.Also have and improve phytosterol in water by preparing the means such as o/w lotions, nanoparticle, cyclodextrin embedding Dissolubility, but still it is difficult to reach higher embedding rate and load rate, solubility is not high in water, and stability, solubility are poor, add There are security risk for some synthetic emulsifiers added.
United States Patent (USP) 7,678,405 discloses the preparation method of a plant sterols o/w lotions, with xanthans, carragheen, The edible oil such as the compound emulsifying agents such as sucrose ester, agar and polyglycerol ester, addition corn oil, soybean oil, rapeseed oil is cooked oil phase preparation Phytosterol o/w lotions, selected phytosterol exist with esterified form mostly, comprise only free phytosterol on a small quantity.The U.S. Patent 6,113, the 972 invention preparation method of one plant sterols-albumen composition, using albumen as wall material, and is aided with single sweet The emulsifying agent such as ester and lecithin, phytosterol is dissolved in oil, prepares stable phytosterol-albumen microcapsules.Chinese patent Phytosterol is dissolved in oil phase by 101991160A, in a heated condition with sucrose fatty ester, Glycerin, mixed triester with caprylic acid capric acid and poly- sweet Oil and fat acid esters etc. emulsifies phytosterol, prepares phytosterol water-based beverage.The above method needs addition oil in preparation process Phytosterol meltage is mutually improved, but the Excess free enthalpy of oil will also result in lipids contents rise, can not reach preferable lipid-loweringing Effect.
Liposome is as a kind of pharmaceutical carrier, available for embedding various water-soluble or oil soluble materials.In liposome preparation During, often adding cholesterol improves phosphatide membrane structure, improves liposome stability.Phytosterol is similar to cholesterol structure, Have been found to may replace preparation of the cholesterol for liposome, and cholesterol in human body content can be reduced.Liposome can be made Into two kinds of forms of lotion and powder, for lotion, powder is not only more convenient storage and transport, but also from addition object shape State (liquid or solid-state) influences, can be freely in various aqueous foods, in the non-aqueous food such as cake, candy, biscuit and oatmeal It can also be added in product, therefore prepare powder there are more preferable market prospects.Liposome drying need to be added big frequently with freeze-drying Freeze drying protectant is measured, to improve the solubility of liposome, significantly reduces drugloading rate.
With 3 kinds of common water-solubility protein (soybean protein isolate, Whey Protein Concentrate and junket in Chinese patent 105410934A Protein acid sodium) it is emulsifying agent, phytosterol is dissolved in n-hexane, water-solubility protein-plant steroid is prepared by emulsifying evaporation Alcohol nano particle.This method is easy to operate, improves the solubility of nano particle.But the technical solution is dredged with water-solubility protein The water base direct embedded plant sterol of group, in the drying process since unfreezing can destroy nanoparticle structure, embedding rate reduces；Together When nanoparticle in phytosterol solubility it is still relatively low by water-solubility protein solubility limit, solubility in water.Peng Jie is used The anti-solvent precipitation method prepare phytosterol nanoparticle, are received respectively with soybean 7 S globulin and casein embedded plant sterol, gained Phytosterol solubility reaches 1.17mg/mL to grain of rice solubility in below 1mg/mL, sucrose ester embedding gained nanoparticle, but Particle diameter is more than 2 μm；Wen Juan embeds phytosterol using hydroxypropyl cyclodextrin, the solubility of phytosterol in inclusion compound It can reach 0.466mg/mL.Phytosterol solubility is had a great influence by coating wall material, therefore selects suitable emulsifying agent or wall material It is good to obtaining a kind of solubility, it is particularly important that solubility high phytosterol preparation.
The content of the invention
The purpose of the present invention is in order to solve the above-mentioned technical problem, there is provided a kind of technique is simple, derives from a wealth of sources, production cost The preparation method of protein modified phytosterol liposome powder low, control is easy, the product cut size of production is small, solubility is good, Solubility is high, has excellent lipid-lowering effect.
Technical solution includes the following steps：
(1) lecithin and phytosterol are according to 2~6:The ratio of 1 (w/w) is added in absolute ethyl alcohol, makes phytosterol dense Spend for 0.5%~4% (w/v), dissolving is stirred at room temperature, prepare organic phase；
(2) soybean protein isolate is added in deionized water, it is 0.5%~4% (w/ to make soybean protein isolate concentration V), water phase is prepared after dissolving aquation is stirred at room temperature；
(3) according to volume ratio 1：Organic phase is slowly injected into water phase by 1~10 ratio, and high speed dispersion mixes；Then Repeatedly it is high-pressure homogeneous after obtain protein modified phytosterol liposome；
(4) phytosterol liposome rotary evaporation obtained by step (3) is removed into ethanol, freeze-drying or spray drying, obtained To freeze-dried powder or spray powder.
The phytosterol is at least one of cupreol, stigmasterol, campesterol or brassicasterol.
The lecithin is soybean lecithin or egg yolk lecithin.
Water phase is made in the adjusting pH to 7.0 after dissolving is stirred at room temperature, 4 DEG C of aquations overnight.
The high speed dispersion time described in step (3) is 1~5min, and rotating speed is 8000~15000r/min.
High-pressure homogeneous pressure described in step (3) is 40~100MPa, and homogenization cycles are 2~8 times.
It is that 38~45 DEG C of rotary evaporations remove under 0.08~0.13MPa vacuums that ethanol described in step (4), which removes mode, Go.
Inventor just improves the problems such as phytosterol is water-soluble and conducts in-depth research, and has carried out following improvement, and (1) is adopted With the compounding of phytosterol and lecithin, using the emulsification of lecithin, make phytosterol nanosizing, solve it in water not The problem of molten；(2) phytosterol can effectively reduce T-CHOL and low density lipoprotein cholesterol content in blood fat, and rise is highly dense The effect of lipoprotein is spent, but it is invalid to triglycerides, and the lecithin arranged in pairs or groups is effective to triglycerides, has blood fat reducing function, So as to fulfill invented liposomes to triglycerides, regulation and control effect of T-CHOL and low density lipoprotein cholesterol；(3) first will Phytosterol and lecithin are combined into the less liposome of particle diameter, are re-introduced into soybean protein isolate, using albumen and lecithin it Between existing hydrophobic effect and electrostatic interaction, be combined with each other, (not direct to be combined with phytosterol), so as to further improve plant The stability of sterol liposome, moreover it is possible to greatly improve product solubility.Wherein, in order to ensure embedding of the lecithin to phytosterol Effect and load rate, lecithin are 2~6 with phytosterol by weight ratio:1 (w/w), the excessive load rate that can reduce phytosterol, The too low embedding effect that can influence phytosterol, liposomal particle size are bigger than normal.The organic solvent is absolute ethyl alcohol, organic phase and water The mixing ratio of phase is 1：1~10, proportion of ethanol too plantlet sterol dissolving it is insufficient, easily separated out in dispersion process, it is excessive then Easily cause a large amount of albuminous degenerations, influence to embed effect.
Gained beneficial effects of the present invention are as follows：
The present invention is using natural phytosterin and soybean lecithin as raw material, using absolute ethyl alcohol as solvent, soybean protein isolate For protective agent, protein modified phytosterol liposome is prepared using alcohol injection, not only increases the dissolubility of phytosterol, Also arrange in pairs or groups triglyceride reducing effect of lecithin, realize more preferable lipid-lowering effect, and improve the stability of liposome with again Dispersiveness.Selected raw material sources extensively, securely and reliably, are suitable for industrialized production and application.Products obtained therefrom particle diameter is small, dissolubility Good, embedding rate height, can be added in the industries such as food, medicine as additive, be particularly suitable for people with hyperlipidemia, have extensive Application prospect.
Embodiment
Embodiment 1
(1) lecithin and phytosterol are pressed 4:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 1% (w/v), is stirred at room temperature dissolving；
(2) soybean protein isolate is added in deionized water in 1% (w/v) ratio, is stirred at room temperature dissolving, adjust pH to 7.0,4 DEG C of aquations are stayed overnight；
(3) according to volume ratio 1：The phytosterol that 4 ratio obtains step (1)-lecithin ethanol solution is slowly injected into In the protein solution that step (2) obtains, and in 10000r/min high speed dispersions 5min；Then high-pressure homogeneous 8 times under 80MPa To protein modified phytosterol liposome；
(4) by phytosterol liposome obtained by step (3) under 0.11MPa vacuums, 40 DEG C of rotary evaporations remove ethanol, Freeze-drying, obtains freeze-dried powder.
Embodiment 2
(1) lecithin and phytosterol are pressed 3:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 2% (w/v), is stirred at room temperature dissolving；
(2) soybean protein isolate is added in deionized water in 1% (w/v) ratio, is stirred at room temperature dissolving, adjust pH to 7.0,4 DEG C of aquations are stayed overnight；
(3) according to volume ratio 1：The phytosterol that 4 ratio obtains step (1)-lecithin ethanol solution is slowly injected into In the protein solution that step (2) obtains, and in 10000r/min high speed dispersions 3min；Then high-pressure homogeneous 4 times under 90MPa To protein modified phytosterol liposome；
(4) by phytosterol liposome obtained by step (3) under 0.13MPa vacuums, 40 DEG C of rotary evaporations remove ethanol, Freeze-drying, obtains freeze-dried powder.
Embodiment 3
(1) lecithin and phytosterol are pressed 4:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 4% (w/v), is stirred at room temperature dissolving；
(2) soybean protein isolate is added in deionized water in 0.5% (w/v) ratio, dissolving is stirred at room temperature, adjust pH Stayed overnight to 7.0,4 DEG C of aquations；
(3) according to volume ratio 1：The phytosterol that 4 ratio obtains step (1)-lecithin ethanol solution is slowly injected into In the protein solution that step (2) obtains, and in 8000r/min high speed dispersions 5min；Then high-pressure homogeneous 4 times under 80MPa To protein modified phytosterol liposome；
(4) by phytosterol liposome obtained by step (3) under 0.11MPa vacuums, 45 DEG C of rotary evaporations remove ethanol, Freeze-drying, obtains freeze-dried powder.
Embodiment 4
(1) lecithin and phytosterol are pressed 2:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 2% (w/v), is stirred at room temperature dissolving；
(2) soybean protein isolate is added in deionized water in 1% (w/v) ratio, is stirred at room temperature dissolving, adjust pH to 7.0,4 DEG C of aquations are stayed overnight；
(3) according to volume ratio 1：The phytosterol that 2 ratio obtains step (1)-lecithin ethanol solution is slowly injected into In the protein solution that step (2) obtains, and in 10000r/min high speed dispersions 5min；Then high-pressure homogeneous 6 times under 90MPa To protein modified phytosterol liposome；
(4) by phytosterol liposome obtained by step (3) under 0.11MPa vacuums, 40 DEG C of rotary evaporations remove ethanol, Freeze-drying, obtains freeze-dried powder.
Embodiment 5
(1) lecithin and phytosterol are pressed 6:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 2% (w/v), is stirred at room temperature dissolving；
(2) soybean protein isolate is added in deionized water in 0.75% (w/v) ratio, dissolving is stirred at room temperature, adjusted PH to 7.0,4 DEG C of aquations are stayed overnight；
(3) according to volume ratio 1：The phytosterol that 10 ratio obtains step (1)-lecithin ethanol solution is slowly injected into In the protein solution that step (2) obtains, and in 15000r/min high speed dispersions 1min；Then high-pressure homogeneous 6 times under 100MPa Obtain protein modified phytosterol liposome；
(4) by phytosterol liposome obtained by step (3) under 0.11MPa vacuums, 38 DEG C of rotary evaporations remove ethanol, In 70 DEG C of inlet temperature, 55 DEG C of outlet temperature, is spray-dried under 0.4MPa vacuums, obtains spray powder.
Embodiment 6
(1) lecithin and phytosterol are pressed 2:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 1% (w/v), is stirred at room temperature dissolving；
(2) soybean protein isolate is added in deionized water in 4% (w/v) ratio, is stirred at room temperature dissolving, adjust pH to 7.0,4 DEG C of aquations are stayed overnight；
(3) according to volume ratio 1：The phytosterol that 4 ratio obtains step (1)-lecithin ethanol solution is slowly injected into In the protein solution that step (2) obtains, and in 10000r/min high speed dispersions 3min；Then high-pressure homogeneous 8 times under 40MPa To protein modified phytosterol liposome；
(4) by phytosterol liposome obtained by step (3) under 0.08MPa vacuums, 40 DEG C of rotary evaporations remove ethanol, In 70 DEG C of inlet temperature, 55 DEG C of outlet temperature, is spray-dried under 0.4MPa vacuums, obtains spray powder.
Comparative example 7
(1) lecithin and phytosterol are pressed 4:1 (w/w) ratio is added in absolute ethyl alcohol, makes the phytosterol concentration be 1% (w/v), is stirred at room temperature dissolving；
(2) according to volume ratio 1：Phytosterol-lecithin ethanol solution is slowly injected into deionized water by 4 ratio, and In 10000r/min high speed dispersions 3min；Then phytosterol liposome is obtained high-pressure homogeneous 6 times under 80MPa；
(3) by phytosterol liposome obtained by step (2) under 0.09MPa vacuums, 40 DEG C of rotary evaporations remove ethanol, Freeze-drying, obtains freeze-dried powder.
Comparative example 8
(1) phytosterol is pressed 5:1 (w/w) ratio is added in n-hexane, and 45 DEG C of stirring and dissolvings, it is molten to obtain phytosterol Liquid；
(2) soybean protein isolate is added in deionized water in 1% (w/v) ratio, is stirred at room temperature dissolving, adjust pH to 7.0,4 DEG C of aquations are stayed overnight；
(3) according to volume ratio 1：Phytosterol solution is slowly injected into protein solution by 10 ratio, and in 10000r/ Min high speed dispersions 3min；Then phytosterol nano particle is obtained high-pressure homogeneous 6 times under 80MPa；
(4) by phytosterol nano particle obtained by step (3) under 0.09MPa vacuums, 40 DEG C of rotary evaporations remove just Hexane, freeze-drying, obtains freeze-dried powder.
Malvern ParticleSizer measures front and rear particle diameter, pdI and the current potential size of phytosterol liposome drying：Liposome dilutes Particle diameter is measured after 200 times, current potential is measured after 20 times of dilution；Dried powder is dissolved with 1mg/ml, is measured current potential, is surveyed after 10 times of dilution Determine particle diameter, the results are shown in Table 1：
Physicochemical property characterizes before and after the protein modified phytosterol liposome drying of table 1
Gas phase analysis content of phytosterol：The liposome of preparation is centrifuged into 20min in 8000g, 200 μ l of supernatant is taken, adds It is internal standard to enter 200 μ l1mg/ml5 α-cholestane, adds 10ml1mol/l KOH ethanol solutions, and 80 DEG C of saponification 1h, add 5ml water, 10ml n-hexane extractions, take supernatant to wash 3 times, nitrogen is blown, BSTFA70 DEG C of derivative 1h, gas phase analysis content of phytosterol.It is dry Liposome powder is dissolved with 20mg/ml afterwards, and 8000g centrifugation 20min, take supernatant to measure content of phytosterol in the same way. The front and rear embedding rate of protein modified phytosterol liposome drying and load rate are as shown in table 2：
The liposome embedded rate of 2 phytosterol of table and load rate
Conclusion：Phytosterol liposomal particle size described in comparative example 7 is 72.71nm, and pdI 0.155, current potential is -34.8, fat Plastid particle diameter distribution is uniform, and stability is good；After liposome is freeze-dried, solubility is poor, and particle diameter has bright in more than 2000nm Aobvious insoluble matter.Protein modified phytosterol liposome described in embodiment has a little increase compared with proliposome particle diameter, but stability with Dispersiveness does not change；Liposome solubility is good after freeze-dried or spray drying, phytosterol bag in section Example Bury rate and still can reach more than 85%.Illustrate that lecithin works well phytosterol inclusion；Soybean protein isolate can effectively improve The solubility and stability of phytosterol liposome, improve the water solubility of phytosterol.8 the method for comparative example is separated with soybean Albumen prepares phytosterol nanoparticle, and products obtained therefrom solubility is good, but the solubility of phytosterol is still relatively low；Through protein modified plant Thing sterol liposome solves the problems, such as this, and not only solubility is changed the liposome after freeze-dried or spray drying Kind, the solubility of products obtained therefrom phytosterol is in more than 1.5mg/mL.

Claims (7)

1. a kind of preparation method of protein modified phytosterol liposome powder, it is characterised in that comprise the following steps that：
(1) lecithin and phytosterol are according to 2~6:The ratio of 1 (w/w) is added in absolute ethyl alcohol, makes the phytosterol concentration be 0.5%~4% (w/v), is stirred at room temperature dissolving, prepares organic phase；
(2) soybean protein isolate is added in deionized water, soybean protein isolate is 0.5%~4% (w/v), is stirred at room temperature Water phase is prepared after dissolving, aquation；
(3) according to volume ratio 1：Organic phase is slowly injected into water phase by 1~10 ratio, and high speed dispersion mixes；Then it is multiple Protein modified phytosterol liposome is obtained after high-pressure homogeneous；
(4) phytosterol liposome rotary evaporation obtained by step (3) is removed into ethanol, freeze-drying or spray drying, is frozen Dry powder or spray powder.
2. the preparation method of protein modified phytosterol liposome powder according to claim 1, it is characterised in that described Phytosterol at least one of for cupreol, stigmasterol, campesterol or brassicasterol.
3. the preparation method of protein modified phytosterol liposome powder according to claim 1, it is characterised in that described Lecithin is soybean lecithin or egg yolk lecithin.
4. the preparation method of protein modified phytosterol liposome powder according to claim 1, it is characterised in that step (2) in, water phase is made in the adjusting pH to 7.0 after dissolving is stirred at room temperature, 4 DEG C of aquations overnight.
5. the preparation method of protein modified phytosterol liposome powder according to claim 1, it is characterised in that step (3) high speed dispersion time described in is 1~5min, and rotating speed is 8000~15000r/min.
6. the preparation method of protein modified phytosterol liposome powder according to claim 1 or 5, it is characterised in that High-pressure homogeneous pressure described in step (3) is 40~100MPa, and homogenization cycles are 2~8 times.
7. according to the preparation method of the protein modified phytosterol liposome powder of claim 1-5 any one of them, its feature It is, it is under 0.08~0.13MPa vacuums that ethanol described in step (4), which removes mode, and 38~45 DEG C of rotary evaporations remove.
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CN110250518A (en) * 2019-07-31 2019-09-20 江南大学 A kind of preparation method of water-soluble plant sterols nano particle
CN110946285A (en) * 2019-11-29 2020-04-03 暨南大学 Preparation method of water-in-oil Pickering emulsion based on phytosterol stabilization
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CN112956680A (en) * 2021-02-10 2021-06-15 青岛农业大学 Plant oil body-imitated core-shell liposome and preparation method thereof
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