CN105925627B - Microbial oil and preparation method thereof - Google Patents

Microbial oil and preparation method thereof Download PDF

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Publication number
CN105925627B
CN105925627B CN201610271571.8A CN201610271571A CN105925627B CN 105925627 B CN105925627 B CN 105925627B CN 201610271571 A CN201610271571 A CN 201610271571A CN 105925627 B CN105925627 B CN 105925627B
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content
oil
microbial oil
acid
miscella
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CN105925627A (en
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汪志明
周强
陆姝欢
肖敏
田勇
李翔宇
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Limited By Share Ltd Biotechnology (wuhan) Co Ltd
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Limited By Share Ltd Biotechnology (wuhan) Co Ltd
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    • C—CHEMISTRY; METALLURGY
    • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
    • C11B3/00—Refining fats or fatty oils
    • C11B3/003—Refining fats or fatty oils by enzymes or microorganisms, living or dead
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6436—Fatty acid esters
    • C12P7/6445—Glycerides
    • C12P7/6463—Glycerides obtained from glyceride producing microorganisms, e.g. single cell oil
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/152—Milk preparations; Milk powder or milk powder preparations containing additives
    • A23C9/1528—Fatty acids; Mono- or diglycerides; Petroleum jelly; Paraffine; Phospholipids; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
    • A23C9/00—Milk preparations; Milk powder or milk powder preparations
    • A23C9/16—Agglomerating or granulating milk powder; Making instant milk powder; Products obtained thereby
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23D—EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS, COOKING OILS
    • A23D9/00—Other edible oils or fats, e.g. shortenings, cooking oils
    • A23D9/007—Other edible oils or fats, e.g. shortenings, cooking oils characterised by ingredients other than fatty acid triglycerides
    • A23D9/013—Other fatty acid esters, e.g. phosphatides
    • 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
    • A23L33/115—Fatty acids or derivatives thereof; Fats or oils
    • C—CHEMISTRY; METALLURGY
    • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
    • C11B1/00—Production of fats or fatty oils from raw materials
    • C11B1/10—Production of fats or fatty oils from raw materials by extracting
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6409—Fatty acids
    • C12P7/6418—Fatty acids by hydrolysis of fatty acid esters
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6409—Fatty acids
    • C12P7/6427—Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6436—Fatty acid esters
    • C12P7/6445—Glycerides
    • C12P7/6472—Glycerides containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone
    • 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

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Abstract

The present invention relates to a kind of microbial oil and preparation method thereof, the content of polyunsaturated fatty acid is greater than 30wt%, and content of triglyceride is less than 90wt%, and diacylglycerol content is not less than 5wt% and is not higher than 20 wt%.It is prepared the following steps are included: being fermented to obtain the fermentation liquid rich in PUFA microbial oil using oleaginous microorganism；The thallus for being rich in PUFA microbial oil is collected, obtains miscella after extraction filtering；Lipase is added in miscella and water is digested, obtains microbial oil after purification, or adds the mixture containing diglyceride into miscella, removes solvent after mixing and obtains microbial oil.It contains suitable diglyceride, is conducive to microbial oil and forms stable emulsion.During preparing microcapsules, microbial oil can be made preferably to be embedded, and then the surface oil content of microcapsules can be reduced, improved the oxidation resistance of microcapsules, and can moderately extend the shelf life of microcapsules, be conducive to subsequent further production and utilization.

Description

Microbial oil and preparation method thereof
Technical field
The present invention relates to microbial oils and preparation method thereof.
Background technique
Polyunsaturated fatty acid (polyunsaturated fatty acid, PUFA) refers to and contains two or more The fatty acid of double bond generally contains 18 ~ 22 carbon atoms.Industrialized PUFA is produced by unicellular microorganisms such as fungi, algae mostly It is raw, therefore also referred to as microbial oil.
Polyunsaturated fatty acid is broadly divided into two series of ω -3 and ω -6 because of its design feature.ω -3 series includes 18 Carbon trienic acid (being commonly called as alpha-linolenic acid, ALA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA).ω -6 series includes Octadecadienoic acid (being commonly called as linoleic acid, LA), octatecatrienoic acid (being commonly called as gamma-Linolenic acid, GLA), eicosatetraenoic acid (are commonly called as Arachidonic acid).Polyunsaturated fatty acid is the main component of human body cell membrane phospholipid, has decisive shadow to cell membrane function It rings.Some specific polyunsaturated fatty acids such as arachidonic acid and DHA are two kinds of main how unsaturateds in brain and retina Fatty acid influences significantly especially for fetus and infant, and insufficiency of intake may cause brain function and optic nerve development obstacle.
The microbial oil that industrialized production obtains mainly exists in the form of glyceride.Glyceride is glycerol and fatty acid Compound made of esterification, according to the difference of the extent of reaction, be divided into monoglyceride (monoglyceride, MG), diglyceride (diacylglycerol, DG), triglycerides (sweet three ester, TG).Wherein, triglycerides (TG) is formed by 3 molecules of fatty acids and 1 molecule glycerine esterification, is The principal mode that grease stores in the main source and nature different kind organism body of energy i (in vivo).Diglyceride (DG) is by 2 The product that molecules of fatty acids and 1 molecule glycerine esterification obtain, the natural component and grease for being grease are in human body metabolism Between product.Meanwhile the intermediate material of diglyceride or intracellular lipositol signaling pathways.
The microbial oil that industrialized production obtains all is much the functional or very strong grease of specific aim, is generally used for masses The additive or nutrition fortifier of the consumer goods such as dairy products, it is seldom directly edible.Since it is rich in polyunsaturated fatty acid, hold very much Easily be oxidized and flavor caused to deteriorate, thus its be used as food additives or nutrition fortifier when, it usually needs first carry out micro- Capsule embedding treatment.It is microcapsule embedded that mainly microbial oil core material is mixed with suitable material and water, is sheared, homogeneous, emulsification Afterwards, wall material (such as maltodextrin etc.) is added while spray drying to be embedded, makes grease by tight in wall material. Such microcapsule product can not only prevent grease to be oxidized, but also can improve the flavor and taste of product.Under normal conditions, grease Emulsifiability it is stronger, then embed that effect is better, the microcapsules flavor and stability produced are also better.
The patent application of Publication No. CN1662642A discloses a kind of micro- life containing at least 40% polyunsaturated fatty acid Object is oily, and the content of triglyceride in the microbial oil is greater than 90%.The microbial oil have the following deficiencies: due to triglycerides without Hydrophilic radical, no emulsifiability, therefore, the emulsifiability of the microbial oil are poor.It is micro- during the production of subsequent microcapsules Bio oil cannot form good embedding, and finally obtained microcapsule product, surface oil content is higher, be unfavorable for subsequent into one The production and application of step.
Therefore it provides a kind of improved microbial oil is actually necessary.
Summary of the invention
The first technical problem to be solved by the present invention be to provide it is a kind of with good emulsifiability, conducive to the micro- of embedding Bio oil.
The second technical problem to be solved by the present invention is to provide a kind of method for preparing mentioned microorganism oil.
The third technical problem to be solved by the present invention is to provide a kind of low with good embedding effect, surface oil content Microcapsules.
In order to solve the above-mentioned technical problem, the technical solution adopted by the present invention is that:
There is provided a kind of microbial oil, it is characterised in that: the content of its polyunsaturated fatty acid is greater than 30wt%, triglycerides Content is less than 90wt%, and diacylglycerol content is not less than 5wt% and is not higher than 20 wt %.
In above scheme, the content of diglyceride is not less than 10wt%, the content of the triglycerides in the microbial oil Not less than 75 wt %.
In above scheme, the polyunsaturated fatty acid is arachidonic acid, docosahexaenoic acid or 20 light dydrocarbons Olefin(e) acid.In above scheme, the microbial oil is crude oil.
A kind of preparation method of microbial oil is provided, comprising the following steps:
(1) it ferments to obtain the fermentation liquid rich in PUFA microbial oil using oleaginous microorganism；
(2) it collects the thallus for being rich in PUFA microbial oil and obtains miscella after extraction filtering；
(3) lipase is added in miscella and water is digested, obtain microbial oil after purification.
In above scheme, in the step (3), enzymatic hydrolysis parameter includes: to be stirred to react 0.5 ~ 2 under lipase optimum temperature Hour, fatty enzyme dosage is the 0.25wt% ~ 2wt% for mixing oil quality, and water consumption is the 15wt%-30 wt% for mixing oil quality.
In above scheme, the specific steps purified in the step (3) include: to stand miscella, to oily phase and water phase After layering, water layer is removed, is filtered to remove lipase, evaporation removing solvent removes free fatty acid by molecular distillation equipment, obtains To microorganism crude oil.
In above scheme, the preparation method of the microbial oil further includes carrying out essence to microbial oil after step (3) System.
The preparation method of another microbial oil is provided, comprising the following steps:
(1) it ferments to obtain the fermentation liquid rich in PUFA microbial oil using oleaginous microorganism；
(2) thallus for being rich in PUFA microbial oil is collected, obtains miscella after extraction filtering；
(3) mixture containing diglyceride is added into miscella, is removed solvent after mixing and is obtained microorganism Oil.
In above scheme, the preparation method of the microbial oil refines microbial oil after being included in step (3).
There is provided a kind of microcapsules, it is characterised in that: the microcapsules include the mentioned microorganism oil and package institute as core material State the wall material of microbial oil.
Microbial oil of the invention has the following beneficial effects:
Contain suitable diglyceride in microbial oil, since dialycerides have preferable emulsifiability, is conducive to Microbial oil forms stable emulsion.During preparing microcapsules, microbial oil can be made preferably to be embedded, Jin Erke The surface oil content for reducing microcapsules, improves the oxidation resistance of microcapsules, and can moderately extend the shelf life of microcapsules, favorably In subsequent further production and utilization.
Specific embodiment
The microbial oil production of the invention of following embodiment more detailed description and application method.
Embodiment one
Be to set out strain with Mortierella alpina, the present invention is described in detail contain arachidonic microbial oil production and Using.
1. fermentation
The culture medium solution that preparation glucose content is 0.03g/mL, yeast powder content is 0.02g/mL is in 500ml shaking flask In, more bottles can be prepared, appropriate Mortierella alpina mycelia and spore are accessed after sterilizing, is placed in 28 DEG C of constant-temperature tables, 150rpm, 2 days After merge shaking flask, move into it is sterilized, be contained with the culture that glucose content is 0.03g/mL, yeast powder content is 0.02g/mL The 1m of based sols3In fermentor (first class seed pot), it is continually fed into filtrated air, is kept for 28 ± 2 DEG C of cultivation temperature.First order seed After tank culture 2 days, by whole culture solutions move into it is sterilized, be contained with that glucose content is 0.03g/mL, yeast powder content is The 10m of the culture medium solution of 0.02g/mL3In fermentor (secondary seed tank), it is continually fed into filtrated air, keeps cultivation temperature 28±2℃.After secondary seed tank culture 1 day, by whole culture solutions move into it is sterilized, be contained with glucose content be 0.05g/ The 45m of mL, the culture medium solution that yeast powder content is 0.02g/mL3In fermentor, it is continually fed into filtrated air, keeps culture temperature 28 ± 2 DEG C of degree, the sterile dextrose of total amount about 0.02 ~ 0.05g/mL culture medium solution is added according to glucose consumption rate in batches Solution can get tunning after being further cultured for 7 days, wherein biomass content 32g/L, total oil content 51.9wt% in thallus butt, Arachidonic acid content 50.4wt% in total oil.
2. preparation is rich in arachidonic microbial oil
Following different process means can be used and prepare microbial oil of the invention.
Means one
Tunning is realized into separation of solid and liquid by centrifugation or filters pressing mode, collects wet thallus, using crushing crusher machine, then It is dried by fluidized drying tower, obtains dry mycelium.Dry mycelium is mixed to immersion extraction, filtering with organic solvent such as butane or hexane After obtain miscella.
Commercially available lipase is added into miscella and water carries out enzymolysis processing, which has glycerol ester hydrolysis Function, following embodiments are identical with this.The enzymolysis processing parameter includes: that fatty enzyme dosage is microorganism miscella weight The 0.25wt% of amount；Water consumption is the 15wt% of microorganism miscella weight, about 37 DEG C of reaction temperature, is stirred to react time 0.5h. Miscella is purified after enzyme digestion reaction, the specific steps of purifying include: to stand miscella, are mutually divided with water phase to oily After layer, water layer is removed, is filtered to remove enzyme, evaporation removing solvent removes free fatty acid by molecular distillation equipment, obtains micro- life Object crude oil.The crude oil is measured with following index: content of polyunsaturated fatty acid 61.5wt%, TG content 88.7wt%, DG content 5.5wt%。
Further, mentioned microorganism crude oil is refined, the step of purification includes: to pass through the microorganism crude oil 5wt% titanium dioxide is used in 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation again after filtering Silicon and 3wt% activity carbon decoloring, after filtering again, direct steam is deodorized under conditions of 200 DEG C, adds Vc palmitate and Ve As antioxidant, obtain containing arachidonic microorganism refined oil.After measured, polyunsaturated fatty acid in the refined oil It is 6.3 wt % that 62.0 wt %, TG content of total content, which is 89.8 wt %, DG contents,.
Further measure the part physical and chemical index of the microorganism refined oil: 1.1 wt% of unsaponifiable matter, 0.01 wt% of moisture, 0.01 wt% of insoluble impurities, dissolvent residual < 1.0 mg/kg, 0.1 mgKOH/g of acid value, 0.03 meq/kg of peroxide value, instead 0.06 wt% of formula fatty acid, aflatoxin B1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means two
The process means and means one are essentially identical, the difference is that enzymolysis processing technological parameter: fatty enzyme dosage is The 0.5wt% of microorganism miscella weight, water are the 20wt%, reaction time 1h of microorganism miscella weight.What is obtained is micro- In biological crude oil, content of polyunsaturated fatty acid 61.7wt%, TG content 87.0wt%, DG content 7.2wt%.
This microorganism crude oil is refined using with identical process for refining in means one, obtained microorganism refined oil In, polyunsaturated fatty acid total content 61.5wt%, TG content 88.4wt %, DG content 8.8wt %, other physical and chemical indexes and hand The physical and chemical index that section one obtains is close.
Means three
The process means and means one are essentially identical, the difference is that enzymolysis processing technological parameter: fatty enzyme dosage is The 1wt% of microorganism miscella weight, water are the 20wt%, reaction time 1.5h of microorganism miscella weight.Obtained microorganism In crude oil, content of polyunsaturated fatty acid 60.0wt%, TG content 84.0wt%, DG content 10.5wt%.
This microorganism crude oil is refined using with identical process for refining in means one, obtained microorganism refined oil In, polyunsaturated fatty acid total content 61wt%, TG content be 85.3wt %, DG content be 11.4wt %, other physical and chemical indexes with The physical and chemical index that means one obtain is close.
Means four
The process means and means one are essentially identical, the difference is that enzymolysis processing technological parameter: fatty enzyme dosage is The 1wt% of microorganism miscella weight, water are the 25wt%, reaction time 2h of microorganism miscella weight.Obtained microorganism is thick In oil, content of polyunsaturated fatty acid 65.0wt%, TG content 78.4wt%, DG content 13.7wt%.
This microorganism crude oil is refined using with identical process for refining in means one, obtained microorganism refined oil In, polyunsaturated fatty acid total content is 63.8wt%, and TG content is 80.9wt%, and DG content is 15.1wt%, other physical and chemical indexes The physical and chemical index obtained with means one is close.
Means five
The process means and means one are essentially identical, the difference is that enzymolysis processing technological parameter: fatty enzyme dosage is The 2wt% of microorganism miscella weight, water are the 30wt%, reaction time 2h of microorganism miscella weight.Obtained microorganism is thick In oil, content of polyunsaturated fatty acid 57.9wt%, TG content 75.3wt%, DG content 17.8wt%.
This microorganism crude oil is refined using with identical process for refining in means one, obtained microorganism refined oil In, polyunsaturated fatty acid total content 60wt%, TG content is 77.2wt%, and DG content is 19.1wt%, other physical and chemical indexes and hand The physical and chemical index that section one obtains is close.
Means six
Tunning is realized into separation of solid and liquid by centrifugation or filters pressing mode, collects wet thallus, using crushing crusher machine, then It is dried by fluidized drying tower, obtains dry mycelium.Dry mycelium is mixed to immersion extraction, filtering with organic solvent such as butane or hexane After obtain miscella.
The mixture containing diglyceride, such as fatty acid list diacylglycerol or its analog, the mixing are added into miscella Diacylglycerol content is 31.4wt% in object, and the specific gravity for adding mixture is the 11.5wt% of total miscella.To miscella precipitation, mix Microorganism crude oil is obtained after closing uniformly.The microorganism crude oil has following index feature: content of polyunsaturated fatty acid 38.0wt%, TG content 86.7wt%, DG content 5.1wt%.
Further, mentioned microorganism crude oil is refined, the step of purification includes: to pass through the microorganism crude oil 5wt% titanium dioxide is used in 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation again after filtering Silicon and 3wt% activity carbon decoloring, after filtering, direct steam is deodorized under conditions of 200 DEG C, adds Vc palmitate and Ve conduct Antioxidant is obtained containing arachidonic microorganism refined oil.After measured, polyunsaturated fatty acid always contains in the refined oil It is 6.4 wt % that amount, which is 89.2 wt %, DG contents up to 37.0 wt %, TG contents,.
Further measure the part physical and chemical index of the microorganism refined oil: 1.0 wt% of unsaponifiable matter, 0.01 wt% of moisture, 0.01 wt% of insoluble impurities, dissolvent residual < 1.0 mg/kg, 0.1 mgKOH/g of acid value, 0.03 meq/kg of peroxide value, instead 0.04 wt% of formula fatty acid, aflatoxin B1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means seven
The process means and means six are essentially identical, the difference is that: containing in diglyceride mixt, diglyceride contains Amount is 50.8wt%, and the specific gravity of the mixture of addition is the 17.5wt% of total miscella.In obtained microorganism crude oil, how unsaturated Content of fatty acid 41.4wt%, TG content 81.2wt%, DG content 10.4wt%.
Further, this microorganism crude oil is refined using process for refining identical with means six, what is obtained is micro- In biological refined oil, polyunsaturated fatty acid total content 40.4wt %, TG content is 83.6wt %, and DG content is 12.2wt %, The physical and chemical index that other physical and chemical indexes and means six obtain is close.
Means eight
The process means and means six are essentially identical, the difference is that: containing in diglyceride mixt, diglyceride contains Amount is 72.2wt%, and adding proportion is the 22.6wt% of the total miscella of Zhan.In obtained microorganism crude oil, polyunsaturated fatty acid contains Measure 55.7wt%, TG content 75.6wt%, DG content 17.8wt%.
Further, this microorganism crude oil is refined using process for refining identical with means six, what is obtained is micro- In biological refined oil, polyunsaturated fatty acid total content 55.9wt %, TG content is 77.0wt %, and DG content is 19.0wt %, The physical and chemical index that other physical and chemical indexes and means six obtain is close.
It is above-mentioned to prepare microcapsules containing arachidonic microbial oil
Respectively using the essence of arachidonic acid made from commercially available arachidonic oil, above-mentioned means one, means two, means three Liquefaction carries out ingredient according to following list of ingredients:
Title Ratio (wt%) in feed liquid
Arachidonic acid oil 12.5
Maltodextrin 32.5
Casein sodium 4
Sodium ascorbate 1
Pure water 50
After above-mentioned feed liquid is sheared 10min under the revolving speed of 8000rpm, homogeneous is carried out at 40MPa, obtains emulsion. Emulsion is subjected to press spray drying, spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Atomisation pressure
220℃ 90℃ 300g/min 3000m3/h 20Mpa
The surface oil content result for measuring each microcapsule product is as follows:
Used arachidonic acid oil Surface oil content (wt%)
Peanut on Sale tetraenoic acid is oily (3.8 wt % of diacylglycerol content) 0.35
Arachidonic acid refined oil made from means one 0.20
Arachidonic acid refined oil made from means two 0.19
Arachidonic acid refined oil made from means three 0.17
Respectively using arachidonic acid refined oil made from Peanut on Sale tetraenoic acid oil, above-mentioned means four, means five, according to Following list of ingredients carries out ingredient:
Title Ratio (wt%) in feed liquid
Arachidonic acid oil 11
Maltodextrin 14.5
Converted starch 28
Sodium ascorbate 1.5
Pure water 45
After above-mentioned feed liquid is sheared 15min under the revolving speed of 9000rpm, homogeneous is carried out at 45MPa, obtains emulsion. This emulsion is subjected to atomizer drying, spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Rotary speed
200℃ 80℃ 330g/min 3500m3/h 3500rpm
The surface oil content result for measuring microcapsules is as follows:
Used arachidonic acid oil Surface oil content (wt%)
Peanut on Sale tetraene acid lipid (diacylglycerol content 4wt %) 0.45
Arachidonic acid refined oil made from means four 0.29
Arachidonic acid refined oil made from means five 0.27
Respectively using arachidonic acid treating made from Peanut on Sale tetraenoic acid oil, above-mentioned means six, means seven, means eight Oil carries out ingredient according to following list of ingredients:
Title Ratio (wt%) in feed liquid
Arachidonic acid oil 13
Maltodextrin 25
Casein sodium 4
Sodium ascorbate 3
Pure water 55
After above-mentioned feed liquid is sheared 15min under the revolving speed of 10000rpm, homogeneous is carried out at 50MPa, obtains emulsion. This emulsion is subjected to spray granulating and drying, it is necessary first to put into 15kg maltodextrin and do bottom material, spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Rotary speed
200℃ 80℃ 330g/min 3500m3/h 3500rpm
The surface oil content result for measuring microcapsules is as follows:
Used arachidonic acid oil Surface oil content (wt%)
Peanut on Sale tetraene acid lipid (diacylglycerol content 3.5wt %) 0.31
Arachidonic acid refined oil made from means six 0.19
Arachidonic acid refined oil made from means seven 0.16
Arachidonic acid refined oil made from means eight 0.12
It is above-mentioned to prepare milk powder containing arachidonic microbial oil
Respectively using arachidonic acid refined oil made from Peanut on Sale tetraenoic acid oil, above-mentioned means one to eight, according to such as Lower list of ingredients carries out ingredient:
Title Ratio (wt%) in feed liquid
Arachidonic acid oil 0.2
Fresh milk 80
Whey powder 13
Lactose 1
Vegetable oil 5.8
After feeding intake according to above-mentioned formula rate, after shearing 10min under the revolving speed of 5000rpm, carried out at 20MPa equal Matter obtains emulsion.Then this emulsion is concentrated into moisture content 50% by three-level falling film type vacuum concentrator, it is most laggard Row press spray is dry, and spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Atomisation pressure
190℃ 75℃ 400g/min 3800m3/h 15MPa
The surface oil content result for measuring milk powder is as follows:
Used arachidonic acid oil Surface oil content (wt%)
Peanut on Sale tetraene acid lipid (4 wt % of dialycerides content) 0.62
Arachidonic acid refined oil made from means one 0.44
Arachidonic acid refined oil made from means two 0.42
Arachidonic acid refined oil made from means three 0.39
Arachidonic acid refined oil made from means four 0.37
Arachidonic acid refined oil made from means five 0.35
Arachidonic acid refined oil made from means six 0.45
Arachidonic acid refined oil made from means seven 0.43
Arachidonic acid refined oil made from means eight 0.31
The surface oil content of microcapsules is to characterize an important indicator of microcapsules quality, indicates the grease not being embedded In the ratio of surface of microcapsule.The surface oil content of microcapsules is higher, shows that more multi-surface grease can be oxidized, then the matter of product It is poorer to measure.Being compared by above data can be seen that under same process conditions, by of the invention containing arachidonic Microcapsules obtained by microbial oil and milk powder, surface oil content are lower.This is primarily due to: microbial oil of the invention contains There is more diglyceride, it can help microbial oil to form more stable emulsion, wrap microbial oil preferably Bury, to reduce the surface oil content of microcapsules and milk powder, improve the oxidation resistance of microcapsules and milk powder, extend microcapsules and The shelf life of milk powder.
Embodiment two
It is the strain that sets out with schizochytrium limacinum, the present invention is described in detail contains the production of the microbial oil of docosahexaenoic acid And application.
1. fermentation
The culture medium solution of glucose content 0.04g/mL, yeast extract content 0.02g/mL are prepared in 1000ml shaking flask In, more bottles can be prepared, appropriate refrigeration schizochytrium limacinum liquid is accessed after sterilizing, is placed in 28 DEG C of constant-temperature tables, 180rpm is activated.2 Second level expansion shaking flask is accessed after it to be cultivated, and shaking flask is merged after 2 days, moves into sterilized, glucose containing 5wt% and 2wt% ferment The 1m of female medicinal extract3In fermentor (first class seed pot), it is continually fed into filtrated air, is kept for 29 ± 1 DEG C of cultivation temperature.First order seed After tank culture 2 days, by whole culture solutions move into it is sterilized, containing glucose content be 0.03g/mL and yeast extract content The 10m of 0.02g/mL3In fermentor (secondary seed tank), it is continually fed into filtrated air, is kept for 29 ± 1 DEG C of cultivation temperature.Second level After seed tank culture 1 day, moves whole culture solutions and enter sterilized, 0.05g/mL containing glucose content and yeast extract content The 45m of 0.02g/mL3In fermentor, it is continually fed into filtrated air, is kept for 29 ± 1 DEG C of cultivation temperature, according to glucose consumption speed Degree batch adds the sterile dextrose solution of total about 0.02 ~ 0.04g/mL, can get tunning after being further cultured for 5 days, wherein giving birth to Object amount 89.7g/L, total oil content 38.5g/L, 51.0 wt % of docosahexaenoic acid content in total oil.
2. the microbial oil that preparation is rich in docosahexaenoic acid
Means one
Tunning is realized into separation of solid and liquid by centrifugation or filters pressing mode, collects wet thallus, using crushing crusher machine, then It is dried by fluidized drying tower, obtains dry mycelium.Dry mycelium is mixed to immersion extraction, filtering with organic solvent such as butane or hexane After obtain miscella.
Commercially available lipase is added into miscella and water carries out enzymolysis processing, which has glycerol ester hydrolysis Function.The enzymolysis processing parameter includes: the 0.25wt% that fatty enzyme dosage is microorganism miscella weight；Water consumption is The 30wt% of microorganism miscella weight, is stirred to react time 0.5h by about 37 DEG C of reaction temperature.To mixing after enzyme digestion reaction Oil is purified, and the specific steps of purifying include: to stand miscella, mutually and after water phase layering, is removed water layer after oily, is crossed and filter out Enzyme is removed, evaporation removing solvent removes free fatty acid by molecular distillation equipment, obtains microorganism crude oil.Measure crude oil tool There is following index: content of polyunsaturated fatty acid 64.0wt%, TG content 86.0wt%, DG content 9.8wt%.
Further, mentioned microorganism crude oil is refined, the step of purification includes: to pass through the microorganism crude oil 5wt% titanium dioxide is used in 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation again after filtering Silicon and 3wt% activity carbon decoloring, after filtering again, direct steam is deodorized under conditions of 200 DEG C, adds Vc palmitate and Ve As antioxidant, the microorganism refined oil containing docosahexaenoic acid is obtained.After measured, how unsaturated rouge in the refined oil Fat acid total content reaches 64.0wt %, and TG content is 88.7wt %, and DG content is 11.5wt %.
Further measure the part physical and chemical index of the microorganism refined oil: unsaponifiable matter 1.0wt%, moisture 0.01wt%, no Solubility impurity 0.01wt%, dissolvent residual < 1.0mg/kg, acid value 0.1mgKOH/g, peroxide value 0.03meq/kg, trans fats Sour 0.06wt%, aflatoxin B1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means two
Tunning is realized into separation of solid and liquid by centrifugation or filters pressing mode, collects wet thallus, using crushing crusher machine, then It is dried by fluidized drying tower, obtains dry mycelium.Dry mycelium is mixed to immersion extraction, filtering with organic solvent such as butane or hexane After obtain miscella.
The mixture containing diglyceride, such as fatty acid list diacylglycerol or its analog, the mixing are added into miscella Diacylglycerol content is 54.9wt% in object, and the specific gravity for adding mixture is the 16.2wt% of total miscella.To miscella precipitation, mix Microorganism crude oil is obtained after closing uniformly.The microorganism crude oil has following index feature: content of polyunsaturated fatty acid 47.2wt%, TG content 82.1wt%, DG content 10.4wt%.
Further, mentioned microorganism crude oil is refined, the step of purification includes: to pass through the microorganism crude oil 5wt% titanium dioxide is used in 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation again after filtering Silicon and 3wt% activity carbon decoloring, after filtering again, direct steam is deodorized under conditions of 200 DEG C, adds Vc palmitate and Ve As antioxidant, the microorganism refined oil containing docosahexaenoic acid is obtained.After measured, how unsaturated rouge in the refined oil Fat acid total content reaches 48.1wt %, and TG content is 84.8wt %, and DG content is 12.0wt %.Other physical and chemical indexes and means one The physical and chemical index of acquisition is close.
3. application
The above-mentioned microbial oil containing docosahexaenoic acid prepares microcapsules
Using docosahexaenoic acid oil made from commercially available docosahexaenoic acid oil, above-mentioned means one, means two, press Ingredient is carried out according to following list of ingredients:
Title Ratio (wt%) in solid content
Docosahexaenoic acid grease 12.5
Maltodextrin 20
Converted starch 15
Sodium ascorbate 2.5
Pure water 50
After above-mentioned feed liquid is sheared 10min under the revolving speed of 8000rpm, homogeneous is carried out at 40MPa, obtains emulsion. This emulsion is subjected to press spray drying, spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Atomisation pressure
220℃ 90℃ 300g/min 3000m3/h 20Mpa
The surface oil content result for measuring microcapsules is as follows:
Used docosahexaenoic acid grease Surface oil content (wt%)
Commercially available docosahexaenoic acid grease (diacylglycerol content 4wt%) 0.85
Docosahexaenoic acid grease made from means one 0.52
Docosahexaenoic acid grease made from means two 0.50
The above-mentioned microbial oil containing docosahexaenoic acid prepares milk powder
Using docosahexaenoic acid oil made from commercially available docosahexaenoic acid oil, above-mentioned means one, means two, press Ingredient is carried out according to following list of ingredients:
Title Ratio (wt%) in feed liquid
Docosahexaenoic acid grease 0.2
Fresh milk 80
Whey powder 11
Lactose 3
Vegetable oil 5.8
After feeding intake according to above-mentioned formula rate, after shearing 10min under the revolving speed of 5000rpm, carried out at 20MPa equal Matter obtains emulsion.Then this emulsion is concentrated into moisture content 50% by three-level falling film type vacuum concentrator, it is most laggard Row press spray is dry, and spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Atomisation pressure
190℃ 75℃ 400g/min 3800m3/h 15Mpa
The surface oil content result for measuring milk powder is as follows:
Used docosahexaenoic acid grease Surface oil content (wt%)
Commercially available docosahexaenoic acid grease (dialycerides content 4wt %) 0.68
Docosahexaenoic acid grease made from means one 0.42
Docosahexaenoic acid grease made from means two 0.39
It can be seen that under same process conditions by the above experimental data, two dodecahexaenes contained by of the invention Microcapsules and milk powder obtained by the microbial oil of acid, surface oil content are lower.This is primarily due to: microorganism of the invention Oil contains more diglyceride, it can help microbial oil to form more stable emulsion, make microbial oil preferably It is embedded, to reduce the surface oil content of microcapsules and milk powder, improves the oxidation resistance of microcapsules and milk powder, extend micro- glue The shelf life of capsule and milk powder.
Embodiment three
It is the strain that sets out with Pythium ultimum, the production and application of the microbial oil containing eicosapentaenoic acid is described in detail.
1. fermentation
The culture medium solution that preparation cane sugar content is 0.05g/mL and yeast powder content is 0.005g/mL is in 1000ml shaking flask In, more bottles can be prepared, appropriate Pythium ultimum is accessed after sterilizing, is placed in 28 DEG C of constant-temperature tables, 180rpm is activated.It is followed by within 2 days Enter second level expansion shaking flask to be cultivated, shaking flask is merged after 2 days, moving into sterilized, cane sugar content is 0.05g/mL and yeast powder Content is the 1m of 0.005g/mL3In fermentor (first class seed pot), it is continually fed into filtrated air, keeps cultivation temperature 28 ± 1 ℃.After first class seed pot culture 2 days, moving whole culture solutions to enter sterilized, cane sugar content is 0.05g/mL and yeast powder content For the 10m of 0.005g/mL3In fermentor (secondary seed tank), it is continually fed into filtrated air, is kept for 28 ± 1 DEG C of cultivation temperature.Two Grade is after seed tank culture 1 day, moves whole culture solutions and enters that sterilized, cane sugar content is 0.05g/mL and yeast powder content is The 45m of 0.005g/mL3In fermentor, it is continually fed into filtrated air, is kept for 28 ± 1 DEG C of cultivation temperature, according to sugar consumption speed Batch adds the sterilized sugar solution of total about 0.02 ~ 0.04g/mL, can get tunning after being further cultured for 5 days, wherein 20 carbon Pentaene acid content 207.8mg/L.
2. the microbial oil that preparation is rich in eicosapentaenoic acid
Means one
Tunning is realized into separation of solid and liquid by centrifugation or filters pressing mode, collects wet thallus, using crushing crusher machine, then It is dried by fluidized drying tower, obtains dry mycelium.Dry mycelium is mixed to immersion extraction, filtering with organic solvent such as butane or hexane After obtain miscella.
Commercially available lipase is added into miscella and water carries out enzymolysis processing, which has glycerol ester hydrolysis Function.The enzymolysis processing parameter includes: the 0.25wt% that fatty enzyme dosage is microorganism miscella weight；Water consumption is The 30wt% of microorganism miscella weight, is stirred to react time 0.5h by about 37 DEG C of reaction temperature.To mixing after enzyme digestion reaction Oil is purified, and the specific steps of purifying include: to stand miscella, mutually and after water phase layering, is removed water layer after oily, is crossed and filter out Enzyme is removed, evaporation removing solvent removes free fatty acid by molecular distillation equipment, obtains microorganism crude oil.The crude oil has such as Lower index feature: content of polyunsaturated fatty acid 59.8wt%, TG content 85.1wt%, DG content 8.5wt%.
Further, mentioned microorganism crude oil is refined, the step of purification includes: to pass through the microorganism crude oil 5wt% titanium dioxide is used in 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation again after filtering Silicon and 3wt% activity carbon decoloring, after filtering again, direct steam is deodorized under conditions of 200 DEG C, adds Vc palmitate and Ve As antioxidant, the microorganism refined oil containing eicosapentaenoic acid is obtained.After measured, polyunsaturated fat in the refined oil Sour total content reaches 59.4wt %, and TG content is 87.5wt %, and DG content is 10.4wt %.
Further measure the part physical and chemical index of the microorganism refined oil: unsaponifiable matter 0.8wt%, moisture 0.01wt%, no Solubility impurity 0.01wt%, dissolvent residual < 1.0mg/kg, acid value 0.1mgKOH/g, peroxide value 0.03meq/kg, trans fats Sour 0.06wt%, aflatoxin B1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means two
Tunning is realized into separation of solid and liquid by centrifugation or filters pressing mode, collects wet thallus, using crushing crusher machine, then It is dried by fluidized drying tower, obtains dry mycelium.Dry mycelium is mixed to immersion extraction, filtering with organic solvent such as butane or hexane After obtain miscella.
The mixture containing diglyceride, such as fatty acid list diacylglycerol or its analog, the mixing are added into miscella Diacylglycerol content is 56.1wt% in object, and the specific gravity of the mixture of addition is the 15.0wt% of total miscella.To miscella precipitation, Microorganism crude oil is obtained after mixing.The crude oil has following index feature: content of polyunsaturated fatty acid 45.0wt%, TG Content 82.0wt%, DG content 9.9wt%.
Further, mentioned microorganism crude oil is refined, the step of purification includes: to pass through the microorganism crude oil 5wt% titanium dioxide is used in 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation again after filtering Silicon and 3wt% activity carbon decoloring, after filtering again, direct steam is deodorized under conditions of 200 DEG C, adds Vc palmitate and Ve As antioxidant, the microorganism refined oil containing eicosapentaenoic acid is obtained.After measured, polyunsaturated fat in the refined oil Sour total content reaches 44.9wt %, and TG content is 84.3wt %, and DG content is 11.9wt %.Other physical and chemical indexes are obtained with means one The physical and chemical index obtained is close.
3. application
The above-mentioned microbial oil containing eicosapentaenoic acid prepares microcapsules
Using eicosapentaenoic acid oil made from commercially available eicosapentaenoic acid oil, above-mentioned means one, means two, according to such as Lower list of ingredients carries out ingredient:
Title Ratio (wt%) in solid content
Eicosapentaenoic acid lipid 14
Maltodextrin 32
Casein sodium 2.5
Sodium ascorbate 1.5
Pure water 50
After above-mentioned feed liquid is sheared 10min under the revolving speed of 8000rpm, homogeneous is carried out at 40MPa, obtains emulsion. This emulsion is spray-dried, spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Atomisation pressure
220℃ 90℃ 300g/min 3000m3/h 20Mpa
The surface oil content result for measuring microcapsules is as follows:
Used eicosapentaenoic acid lipid Surface oil content (wt%)
Commercially available eicosapentaenoic acid lipid (diacylglycerol content 4wt%) 0.79
Eicosapentaenoic acid lipid made from means one 0.51
Eicosapentaenoic acid lipid made from means two 0.47
The above-mentioned microbial oil containing eicosapentaenoic acid prepares milk powder
Using eicosapentaenoic acid oil made from commercially available eicosapentaenoic acid oil, above-mentioned means one, means two, according to such as Lower list of ingredients carries out ingredient:
Title Ratio (wt%) in feed liquid
Eicosapentaenoic acid lipid 0.1
Fresh milk 80
Whey powder 12
Lactose 2
Vegetable oil 5.9
After feeding intake according to above-mentioned formula rate, after shearing 10min under the revolving speed of 5000rpm, carried out at 20MPa equal Matter obtains emulsion.Then this emulsion is concentrated into moisture content 50% by three-level falling film type vacuum concentrator, it is most laggard Row press spray is dry, and spray drying parameters are as follows:
Inlet air temperature Leaving air temp Charging rate Intake volume Atomisation pressure
190℃ 75℃ 400g/min 3800m3/h 15Mpa
The surface oil content result for measuring milk powder is as follows:
Used eicosapentaenoic acid lipid Surface oil content (wt%)
Commercially available eicosapentaenoic acid lipid (dialycerides content 4wt %) 0.72
Eicosapentaenoic acid lipid made from means one 0.48
Eicosapentaenoic acid lipid made from means two 0.44
It can be seen that under same process conditions by above example and experimental data, by eicosapentaenoic of the invention Acid
Microcapsules obtained by microbial oil and milk powder, surface oil content are lower.This is primarily due to: of the invention is micro- Bio oil contains more diglyceride, it can help microbial oil to form more stable emulsion, makes microbial oil more It is embedded well, to reduce the surface oil content of microcapsules and milk powder, improves the oxidation resistance of microcapsules and milk powder, extend The shelf life of microcapsules and milk powder.

Claims (5)

1. a kind of preparation method of microbial oil, it is characterised in that: the following steps are included:
(1) it ferments to obtain the fermentation liquid rich in PUFA microbial oil using oleaginous microorganism；
(2) thallus for being rich in PUFA microbial oil is collected, obtains miscella after extraction filtering；
(3) mixture containing diglyceride is added into miscella, is removed solvent after mixing and is obtained microbial oil；
In the mixture containing diglyceride added in the step (3), the content of diglyceride is 72.2%, addition mixing The specific gravity of object is the 22.6% of total miscella；
The content of polyunsaturated fatty acid is greater than 30wt% in the microbial oil, and content of triglyceride is less than 90wt%, glycerol Two ester contents are not less than 5wt% and are not higher than 20wt%.
2. the preparation method of microbial oil according to claim 1, it is characterised in that: the content of the diglyceride is not low It is not less than 75wt% in the content of 10wt%, the triglycerides.
3. the preparation method of microbial oil according to claim 1 or 2, it is characterised in that: the polyunsaturated fatty acid For arachidonic acid, docosahexaenoic acid or eicosapentaenoic acid.
4. the preparation method of microbial oil according to claim 1 or 2, it is characterised in that: the microbial oil is crude oil.
5. the preparation method of microbial oil according to claim 1, it is characterised in that: to microorganism after step (3) Oil is refined.
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