CN105925627A - Microbial oil and preparation method thereof - Google Patents

Microbial oil and preparation method thereof Download PDF

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Publication number
CN105925627A
CN105925627A CN201610271571.8A CN201610271571A CN105925627A CN 105925627 A CN105925627 A CN 105925627A CN 201610271571 A CN201610271571 A CN 201610271571A CN 105925627 A CN105925627 A CN 105925627A
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oil
content
microbial oil
acid
microbial
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CN105925627B (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 invention relates to microbial oil and a preparation method thereof. The microbial oil has unsaturated fatty acid content greater than 30wt%, triglyceride content less than 90wt% and diglyceride content greater than or equal to 5wt% and less than or equal to 20wt%. The preparation method comprises 1, carrying out oleaginous microorganism fermentation to obtain PUFA microbial oil-rich fermentation broth, 2, collecting the PUFA microbial oil-rich microorganisms and carrying out extraction and filtration to obtain mixed oil, and 3, adding lipase and water into the mixed oil, carrying out enzymolysis and carrying out purification to obtain microbial oil, or adding a diglyceride-containing mixture into the mixed oil, carrying out uniform mixing and removing a solvent so that microbial oil is obtained. The microbial oil contains an appropriate amount of diglyceride and is conducive to formation of a stable emulsion from the microbial oil. In microcapsule preparation, microbial oil can be well coated so that microcapsule surface oil content is reduced, a microcapsule antioxidation capability is improved, a microcapsule shelf life is properly prolonged and later production and use are promoted.

Description

Microbial oil and preparation method thereof
Technical field
The present invention relates to microbial oil and preparation method thereof.
Background technology
Polyunsaturated fatty acid (polyunsaturated fatty acid, PUFA) refers to containing two or more The aliphatic acid of double bond, typically containing 18 ~ 22 carbon atoms.Industrialized PUFA is produced by the unicellular microorganism such as fungi, algae mostly Raw, therefore also referred to as microbial oil.
Polyunsaturated fatty acid is broadly divided into ω-3 and-6 two series of ω because of its design feature.ω-3 series includes 18 Carbon trienic acid (being commonly called as alpha-linolenic acid, ALA), eicosapentaenoic acid (EPA), DHA (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 Ring.Some specific polyunsaturated fatty acids such as arachidonic acid and DHA be in brain and retina two kinds main the most unsaturated Aliphatic acid, affects 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 is mainly presented in glyceride.Glyceride is glycerine and aliphatic acid Esterification compound, according to the difference of the extent of reaction, be divided into monoglyceride (monoglyceride, MG), diglyceride (diacylglycerol, DG), triglycerides (sweet three esters, TG).Wherein, triglycerides (TG) is formed by 3 molecules of fatty acids and 1 molecule glycerine esterification, is The main source of energy i (in vivo), is also the principal mode that grease stores in nature different kind organism body.Diglyceride (DG) is by 2 The product that molecules of fatty acids obtains with 1 molecule glycerine esterification, is the natural component of grease, and also grease is in human body metabolism Between product.Meanwhile, diglyceride or the intermediate material of intracellular lipositol signaling pathways.
The microbial oil that industrialized production obtains is the most all functional or that specific aim is the strongest grease, is generally used for masses The additive of the consumer goods such as dairy products or nutrition fortifier, the most edible.Owing to it is rich in polyunsaturated fatty acid, hold very much The most oxidized and cause local flavor to deteriorate, therefore it is when as food additives or nutrition fortifier, it usually needs first carry out micro- Capsule embedding treatment.Microcapsule embedded mainly microbial oil core mixed with suitable material and water, shear, homogeneous, emulsification After, while being spray-dried, add wall material (such as maltodextrin etc.) embed, make grease by tight in wall material. Such microcapsule product had both been possible to prevent grease oxidized, can improve again local flavor and the mouthfeel of product.Under normal circumstances, grease Emulsifiability the strongest, then embedding effect the best, the microcapsules local flavor produced and stability are the best.
The patent application of Publication No. CN1662642A discloses a kind of micro-life containing at least 40% polyunsaturated fatty acid Thing oil, the content of triglyceride in this microbial oil is more than 90%.There is following defect in this microbial oil: due to triglycerides without Hydrophilic radical, without emulsifiability, therefore, the emulsifiability of this microbial oil is poor.During follow-up microcapsules produce, micro- Bio oil can not form good embedding, the microcapsule product finally given, and its surface oil content is higher, be unfavorable for follow-up enter one The production of step and application.
Therefore it provides the microbial oil of a kind of improvement is the most necessary.
Summary of the invention
One of the technical problem to be solved is to provide and a kind of has good emulsifiability, is beneficial to the micro-of embedding Bio oil.
The two of the technical problem to be solved are to provide a kind of method preparing mentioned microorganism oil.
The three of the technical problem to be solved are to provide one to have, and well to embed effect, surface oil content low Microcapsules.
In order to solve above-mentioned technical problem, the technical solution used in the present invention is:
A kind of microbial oil is provided, it is characterised in that: the content of its polyunsaturated fatty acid is more than 30wt%, content of triglyceride Less than 90wt%, diacylglycerol content is not less than 5wt% and not higher than 20 wt %.
In such scheme, in described microbial oil, the content of diglyceride is not less than 10wt%, the content of described triglycerides It is not less than 75 wt %.
In such scheme, described polyunsaturated fatty acid is arachidonic acid, DHA or 20 carbon five Olefin(e) acid.In such scheme, described microbial oil is crude oil.
The preparation method of a kind of microbial oil is provided, comprises the following steps:
(1) oleaginous microorganism fermentation is utilized to obtain the zymotic fluid rich in PUFA microbial oil；
(2) collect the thalline rich in PUFA microbial oil, after extraction is filtered, obtain miscella；
(3) in miscella, add lipase and water carries out enzymolysis, obtain microbial oil after purification.
In such scheme, in described step (3), enzymolysis parameter includes: stir reaction 0.5 ~ 2 under lipase optimum temperature Hour, lipase consumption is the 0.25wt% ~ 2wt% of miscella quality, and water consumption is the 15wt%-30 wt% of miscella quality.
In such scheme, the concrete steps purified in described step (3) include: stood by miscella, treat oil phase and aqueous phase After layering, remove water layer, be filtered to remove lipase, evaporate desolvation, remove free fatty by molecular distillation equipment, To microorganism crude oil.
In such scheme, the preparation method of described microbial oil carries out essence to microbial oil after being additionally included in step (3) System.
The preparation method of another microbial oil is provided, comprises the following steps:
(1) oleaginous microorganism fermentation is utilized to obtain the zymotic fluid rich in PUFA microbial oil；
(2) collecting the thalline rich in PUFA microbial oil, extraction obtains miscella after filtering；
(3) in miscella, add the mixture containing diglyceride, remove solvent after mixing and obtain microbial oil.
In such scheme, microbial oil is refined after being included in step (3) by the preparation method of described microbial oil.
A kind of microcapsules are provided, it is characterised in that: these microcapsules include the mentioned microorganism oil as core and parcel institute State the wall material of microbial oil.
The microbial oil of the present invention has the advantages that
Containing appropriate diglyceride in microbial oil, owing to dialycerides has preferable emulsifiability, it is conducive to micro-life Thing oil forms stable emulsion.During preparing microcapsules, microbial oil can be made preferably to be embedded, and then can reduce The surface oil content of microcapsules, improve microcapsules oxidation resistance, and can appropriateness extend microcapsules shelf life, be conducive to after Continuous further production and utilization.
Detailed description of the invention
The microbial oil of following example more detailed description present invention produces and application process.
Embodiment one
With Mortierella alpina for the bacterial classification that sets out, describe in detail the present invention contain arachidonic microbial oil production and should With.
1. fermentation
Preparation glucose content is 0.03g/mL, dusty yeast content be the culture medium solution of 0.02g/mL in 500ml shaking flask, can Prepare many bottles, after sterilizing, access appropriate Mortierella alpina mycelia and spore, be placed in 28 DEG C of constant-temperature tables, 150rpm, merge after 2 days Shaking flask, move into sterilized, be contained with glucose content be 0.03g/mL, dusty yeast content be the culture medium solution of 0.02g/mL 1m3In fermentation tank (first class seed pot), it is continually fed into filtrated air, keeps cultivation temperature 28 ± 2 DEG C.First class seed pot is cultivated After 2 days, by whole nutrient solutions move into sterilized, be contained with glucose content be 0.03g/mL, dusty yeast content be 0.02g/ The 10m of the culture medium solution of mL3In fermentation tank (secondary seed tank), it is continually fed into filtrated air, keeps cultivation temperature 28 ± 2 ℃.Secondary seed tank was cultivated after 1 day, whole nutrient solutions are moved into sterilized, to be contained with glucose content be 0.05g/mL, ferment Female powder content is the 45m of the culture medium solution of 0.02g/mL3In fermentation tank, it is continually fed into filtrated air, keeps cultivation temperature 28 ± 2 DEG C, the sterile dextrose in batches adding total amount about 0.02 ~ 0.05g/mL culture medium solution according to glucose consumption rate is molten Liquid, can obtain tunning after being further cultured for 7 days, wherein total oil content 51.9wt% in biomass content 32g/L, thalline butt, total Arachidonic acid content 50.4wt% in oil.
2. preparation is rich in arachidonic microbial oil
Following different process means can be used to prepare the microbial oil of the present invention.
Means one
Tunning is realized separation of solid and liquid by centrifugal or press filtration mode, collects wet thallus, utilize pulverizer to crush, then pass through Fluidized drying tower is dried, and obtains dry mycelium.Dry mycelium is mixed with organic solvent such as butane or hexane immersion extraction, after filtration To miscella.
Adding commercially available lipase in miscella and water carries out enzymolysis processing, this commercially available lipase has glyceride hydrolysis Function, following embodiment is all identical with this.Described enzymolysis processing parameter includes: lipase consumption is microorganism miscella weight The 0.25wt% of amount；Water consumption is the 15wt% of microorganism miscella weight, reaction temperature about 37 DEG C, stirs reaction time 0.5h. Miscella is purified after terminating by enzyme digestion reaction, and the concrete steps of purifying include: is stood by miscella, treats that oil phase divides with aqueous phase After Ceng, remove water layer, be filtered to remove enzyme, evaporate desolvation, remove free fatty by molecular distillation equipment, obtain micro-life Thing crude oil.Record this crude oil and there is following index: content of polyunsaturated fatty acid 61.5wt%, TG content 88.7wt%, DG content 5.5wt%。
Further, refining mentioned microorganism crude oil, refined step includes: by this microorganism crude oil warp 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation, use 5wt% titanium dioxide after filtration again Silicon and 3wt% activated carbon decolorizing, after again filtering, live (open) steam deodorization under conditions of 200 DEG C, add Vc palmitate and Ve As antioxidant, obtain containing arachidonic microorganism refined oil.After measured, polyunsaturated fatty acid in this refined oil Total content 62.0 wt %, TG content is 89.8 wt %, and DG content is 6.3 wt %.
The part physical and chemical index of this microorganism refined oil of mensuration further: unsaponifiable matter 1.1 wt%, moisture 0.01 wt%, Insoluble impurities 0.01 wt%, and dissolvent residual < 1.0 mg/kg, acid value 0.1 mgKOH/g, peroxide value 0.03 meq/kg, instead Formula aliphatic acid 0.06 wt%, AFB1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means two
This process means is essentially identical with means one, and difference is enzymolysis processing technological parameter: lipase consumption is micro-life The 0.5wt% of thing miscella weight, the water yield is the 20wt% of microorganism miscella weight, and the reaction time is 1h.The microorganism obtained In crude oil, content of polyunsaturated fatty acid 61.7wt%, TG content 87.0wt%, DG content 7.2wt%.
Use the process for refining identical with means one that this microorganism crude oil is refined, the microorganism refined oil obtained In, polyunsaturated fatty acid total content 61.5wt%, TG content 88.4wt %, DG content 8.8wt %, other physical and chemical indexs and hand The physical and chemical index that section one obtains is close.
Means three
This process means is essentially identical with means one, and difference is enzymolysis processing technological parameter: lipase consumption is micro-life The 1wt% of thing miscella weight, the water yield is the 20wt% of microorganism miscella weight, and the reaction time is 1.5h.The microorganism crude oil obtained In, content of polyunsaturated fatty acid 60.0wt%, TG content 84.0wt%, DG content 10.5wt%.
Use the process for refining identical with means one that this microorganism crude oil is refined, the microorganism refined oil obtained In, polyunsaturated fatty acid total content 61wt%, TG content is 85.3wt %, and DG content is 11.4wt %, other physical and chemical indexs with The physical and chemical index that means one obtain is close.
Means four
This process means is essentially identical with means one, and difference is enzymolysis processing technological parameter: lipase consumption is micro-life The 1wt% of thing miscella weight, the water yield is the 25wt% of microorganism miscella weight, and the reaction time is 2h.In the microorganism crude oil obtained, Content of polyunsaturated fatty acid 65.0wt%, TG content 78.4wt%, DG content 13.7wt%.
Use the process for refining identical with means one that this microorganism crude oil is refined, the microorganism refined oil obtained In, polyunsaturated fatty acid total content be 63.8wt%, TG content be 80.9wt%, DG content be 15.1wt%, other physical and chemical indexs Close with the physical and chemical index that means one obtain.
Means five
This process means is essentially identical with means one, and difference is enzymolysis processing technological parameter: lipase consumption is micro-life The 2wt% of thing miscella weight, the water yield is the 30wt% of microorganism miscella weight, and the reaction time is 2h.In the microorganism crude oil obtained, Content of polyunsaturated fatty acid 57.9wt%, TG content 75.3wt%, DG content 17.8wt%.
Use the process for refining identical with means one that this microorganism crude oil is refined, the microorganism refined oil obtained In, polyunsaturated fatty acid total content 60wt%, TG content be 77.2wt%, DG content be 19.1wt%, other physical and chemical indexs and hand The physical and chemical index that section one obtains is close.
Means six
Tunning is realized separation of solid and liquid by centrifugal or press filtration mode, collects wet thallus, utilize pulverizer to crush, then pass through Fluidized drying tower is dried, and obtains dry mycelium.Dry mycelium is mixed with organic solvent such as butane or hexane immersion extraction, after filtration To miscella.
The mixture containing diglyceride, such as aliphatic acid list diacylglycerol or its analog, this mixing is added in miscella In thing, diacylglycerol content is 31.4wt%, adds the 11.5wt% that proportion is total miscella of mixture.To miscella precipitation, mixed Microorganism crude oil is obtained after closing uniformly.This microorganism crude oil has a following index feature: content of polyunsaturated fatty acid 38.0wt%, TG content 86.7wt%, DG content 5.1wt%.
Further, refining mentioned microorganism crude oil, refined step includes: by this microorganism crude oil warp 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation, use 5wt% titanium dioxide after filtration again Silicon and 3wt% activated carbon decolorizing, after filtration, live (open) steam deodorization under conditions of 200 DEG C, add Vc palmitate and Ve conduct Antioxidant, obtains containing arachidonic microorganism refined oil.After measured, in this refined oil, polyunsaturated fatty acid always contains Amount reaches 37.0 wt %, and TG content is 89.2 wt %, and DG content is 6.4 wt %.
The part physical and chemical index of this microorganism refined oil of mensuration further: unsaponifiable matter 1.0 wt%, moisture 0.01 wt%, Insoluble impurities 0.01 wt%, and dissolvent residual < 1.0 mg/kg, acid value 0.1 mgKOH/g, peroxide value 0.03 meq/kg, instead Formula aliphatic acid 0.04 wt%, AFB1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means seven
This process means is essentially identical with means six, and difference is: containing in diglyceride mixt, diacylglycerol content is 50.8wt%, the proportion of the mixture of interpolation is the 17.5wt% of total miscella.In the microorganism crude oil obtained, polyunsaturated fat Acid content 41.4wt%, TG content 81.2wt%, DG content 10.4wt%.
Further, using the process for refining identical with means six to refine this microorganism crude oil, obtain is micro- In biological fine liquefaction, 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 indexs obtain with means six is close.
Means eight
This process means is essentially identical with means six, and difference is: containing in diglyceride mixt, diacylglycerol content is 72.2wt%, adding proportion is to account for the 22.6wt% of total miscella.In the microorganism crude oil obtained, content of polyunsaturated fatty acid 55.7wt%, TG content 75.6wt%, DG content 17.8wt%.
Further, using the process for refining identical with means six to refine this microorganism crude oil, obtain is micro- In biological fine liquefaction, 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 indexs obtain with means six is close.
Above-mentioned prepare microcapsules containing arachidonic microbial oil
Use the arachidonic acid treating that commercially available arachidonic oil, above-mentioned means one, means two, means three prepare respectively Oil, carries out dispensing 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
By above-mentioned feed liquid after the rotating speed down cut 10min of 8000rpm, under 40MPa, carry out homogeneous, obtain emulsion.By breast Change liquid carries out press spray and is dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Atomisation pressure
220℃ 90℃ 300g/min 3000m3/h 20Mpa
The surface oil content result recording each microcapsule product is as follows:
The arachidonic acid oil used Surface oil content (wt%)
Peanut on Sale tetraenoic acid oil (diacylglycerol content 3.8 wt %) 0.35
The arachidonic acid refined oil that means one prepare 0.20
The arachidonic acid refined oil that means two prepare 0.19
The arachidonic acid refined oil that means three prepare 0.17
Use the arachidonic acid refined oil that Peanut on Sale tetraenoic acid means four oily, above-mentioned, means five prepare respectively, according to as follows List of ingredients carries out dispensing:
Title Ratio (wt%) in feed liquid
Arachidonic acid oil 11
Maltodextrin 14.5
Converted starch 28
Sodium ascorbate 1.5
Pure water 45
By above-mentioned feed liquid after the rotating speed down cut 15min of 9000rpm, under 45MPa, carry out homogeneous, obtain emulsion.By this Emulsion is centrifuged formula and is spray-dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Rotary speed
200℃ 80℃ 330g/min 3500m3/h 3500rpm
The surface oil content result recording microcapsules is as follows:
The arachidonic acid oil used Surface oil content (wt%)
Peanut on Sale tetraenoic acid grease (diacylglycerol content 4wt %) 0.45
The arachidonic acid refined oil that means four prepare 0.29
The arachidonic acid refined oil that means five prepare 0.27
Use the arachidonic acid refined oil that Peanut on Sale tetraenoic acid means six oily, above-mentioned, means seven, means eight prepare respectively, Dispensing is carried out 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
By above-mentioned feed liquid after the rotating speed down cut 15min of 10000rpm, under 50MPa, carry out homogeneous, obtain emulsion.By this Emulsion carries out spray granulating and drying, it is necessary first to putting into 15kg maltodextrin and do bed material, spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Rotary speed
200℃ 80℃ 330g/min 3500m3/h 3500rpm
The surface oil content result recording microcapsules is as follows:
The arachidonic acid oil used Surface oil content (wt%)
Peanut on Sale tetraenoic acid grease (diacylglycerol content 3.5wt %) 0.31
The arachidonic acid refined oil that means six prepare 0.19
The arachidonic acid refined oil that means seven prepare 0.16
The arachidonic acid refined oil that means eight prepare 0.12
Above-mentioned prepare milk powder containing arachidonic microbial oil
Use the arachidonic acid refined oil that Peanut on Sale tetraenoic acid means oily, above-mentioned to eight prepare respectively, according to joining as follows Material table carries out dispensing:
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 the rotating speed down cut 10min of 5000rpm, under 20MPa, carry out homogeneous, To emulsion.It is then passed through three grades of falling film type vacuum concentrators and this emulsion is concentrated into moisture 50%, finally carry out pressure Formula is spray-dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Atomisation pressure
190℃ 75℃ 400g/min 3800m3/h 15MPa
The surface oil content result recording milk powder is as follows:
The arachidonic acid oil used Surface oil content (wt%)
Peanut on Sale tetraenoic acid grease (dialycerides content 4 wt %) 0.62
The arachidonic acid refined oil that means one prepare 0.44
The arachidonic acid refined oil that means two prepare 0.42
The arachidonic acid refined oil that means three prepare 0.39
The arachidonic acid refined oil that means four prepare 0.37
The arachidonic acid refined oil that means five prepare 0.35
The arachidonic acid refined oil that means six prepare 0.45
The arachidonic acid refined oil that means seven prepare 0.43
The arachidonic acid refined oil that means eight prepare 0.31
The surface oil content of microcapsules is the important indicator characterizing microcapsules quality, and it represents that the grease not being embedded is micro- The ratio of capsule surface.The surface oil content of microcapsules is the highest, shows that more multi-surface grease can be oxidized, then the quality of product is just The poorest.Contrasted it can be seen that under equal process conditions by data above, by the present invention containing arachidonic micro-life Microcapsules obtained by thing oil and milk powder, its surface oil content is lower.This is primarily due to: the microbial oil of the present invention contains relatively Many diglycerides, it can help microbial oil to form more stable emulsion, make microbial grease preferably be embedded, from And reduce the surface oil content of microcapsules and milk powder, improve microcapsules and the oxidation resistance of milk powder, extend microcapsules and milk powder Shelf life.
Embodiment two
With schizochytrium limacinum for the bacterial classification that sets out, describe the production of the microbial oil that the present invention contains DHA in detail and answer With.
1. fermentation
Preparation glucose content 0.04g/mL, yeast extract content 0.02g/mL culture medium solution in 1000ml shaking flask, can Preparing many bottles, access appropriate refrigeration schizochytrium limacinum liquid, be placed in 28 DEG C of constant-temperature tables after sterilizing, 180rpm activates.Within 2 days, it is followed by Enter two grades of expansion shaking flasks to cultivate, merge shaking flask after 2 days, move into sterilized, containing 5wt% glucose and 2wt% yeast extract 1m3In fermentation tank (first class seed pot), it is continually fed into filtrated air, keeps cultivation temperature 29 ± 1 DEG C.First class seed pot is cultivated After 2 days, by whole nutrient solutions move into sterilized, be 0.03g/mL containing glucose content and yeast extract content 0.02g/mL 10m3In fermentation tank (secondary seed tank), it is continually fed into filtrated air, keeps cultivation temperature 29 ± 1 DEG C.Secondary seed tank cultivates 1 After it, move whole nutrient solution and enter sterilized, containing glucose content 0.05g/mL and the 45m of yeast extract content 0.02g/mL3 In fermentation tank, it is continually fed into filtrated air, keeps cultivation temperature 29 ± 1 DEG C, add the most about according to glucose consumption rate batch The sterile dextrose solution of 0.02 ~ 0.04g/mL, can obtain tunning after being further cultured for 5 days, wherein biomass 89.7g/L, total DHA content 51.0 wt % in oil content 38.5g/L, total oil.
2. preparation is rich in the microbial oil of DHA
Means one
Tunning is realized separation of solid and liquid by centrifugal or press filtration mode, collects wet thallus, utilize pulverizer to crush, then pass through Fluidized drying tower is dried, and obtains dry mycelium.Dry mycelium is mixed with organic solvent such as butane or hexane immersion extraction, after filtration To miscella.
Adding commercially available lipase in miscella and water carries out enzymolysis processing, this commercially available lipase has glyceride hydrolysis Function.Described enzymolysis processing parameter includes: lipase consumption is the 0.25wt% of microorganism miscella weight；Water consumption is The 30wt% of microorganism miscella weight, reaction temperature about 37 DEG C, stirs reaction time 0.5h.Enzyme digestion reaction terminate after to mixing Oil is purified, and the concrete steps of purifying include: stood by miscella, after oil phase is layered with aqueous phase, removes water layer, crosses and filter Remove enzyme, evaporate desolvation, remove free fatty by molecular distillation equipment, obtain microorganism crude oil.Record this crude oil tool There is a following index: content of polyunsaturated fatty acid 64.0wt%, TG content 86.0wt%, DG content 9.8wt%.
Further, refining mentioned microorganism crude oil, refined step includes: by this microorganism crude oil warp 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation, use 5wt% titanium dioxide after filtration again Silicon and 3wt% activated carbon decolorizing, after again filtering, live (open) steam deodorization under conditions of 200 DEG C, add Vc palmitate and Ve As antioxidant, obtain the microorganism refined oil containing DHA.After measured, many unsaturated lipids in this refined oil It is 88.7wt % that fat acid total content reaches 64.0wt %, TG content, and DG content is 11.5wt %.
The part physical and chemical index of this microorganism refined oil of mensuration further: 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 Acid 0.06wt%, AFB1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means two
Tunning is realized separation of solid and liquid by centrifugal or press filtration mode, collects wet thallus, utilize pulverizer to crush, then pass through Fluidized drying tower is dried, and obtains dry mycelium.Dry mycelium is mixed with organic solvent such as butane or hexane immersion extraction, after filtration To miscella.
The mixture containing diglyceride, such as aliphatic acid list diacylglycerol or its analog, this mixing is added in miscella In thing, diacylglycerol content is 54.9wt%, adds the 16.2wt% that proportion is total miscella of mixture.To miscella precipitation, mixed Microorganism crude oil is obtained after closing uniformly.This microorganism crude oil has a following index feature: content of polyunsaturated fatty acid 47.2wt%, TG content 82.1wt%, DG content 10.4wt%.
Further, refining mentioned microorganism crude oil, refined step includes: by this microorganism crude oil warp 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation, use 5wt% titanium dioxide after filtration again Silicon and 3wt% activated carbon decolorizing, after again filtering, live (open) steam deodorization under conditions of 200 DEG C, add Vc palmitate and Ve As antioxidant, obtain the microorganism refined oil containing DHA.After measured, many unsaturated lipids in this refined oil It is 84.8wt % that fat acid total content reaches 48.1wt %, TG content, and DG content is 12.0wt %.Other physical and chemical indexs and means one The physical and chemical index obtained is close.
3. application
The above-mentioned microbial oil containing DHA prepares microcapsules
Use the DHA oil that commercially available DHA means one oily, above-mentioned, means two prepare, according to such as Lower list of ingredients carries out dispensing:
Title Ratio (wt%) in solid content
Docosahexaenoic acid grease 12.5
Maltodextrin 20
Converted starch 15
Sodium ascorbate 2.5
Pure water 50
By above-mentioned feed liquid after the rotating speed down cut 10min of 8000rpm, under 40MPa, carry out homogeneous, obtain emulsion.By this Emulsion carries out press spray and is dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Atomisation pressure
220℃ 90℃ 300g/min 3000m3/h 20Mpa
The surface oil content result recording microcapsules is as follows:
The docosahexaenoic acid grease used Surface oil content (wt%)
Commercially available docosahexaenoic acid grease (diacylglycerol content 4wt%) 0.85
The docosahexaenoic acid grease that means one prepare 0.52
The docosahexaenoic acid grease that means two prepare 0.50
The above-mentioned microbial oil containing DHA prepares milk powder
Use the DHA oil that commercially available DHA means one oily, above-mentioned, means two prepare, according to such as Lower list of ingredients carries out dispensing:
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 the rotating speed down cut 10min of 5000rpm, under 20MPa, carry out homogeneous, To emulsion.It is then passed through three grades of falling film type vacuum concentrators and this emulsion is concentrated into moisture 50%, finally carry out pressure Formula is spray-dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Atomisation pressure
190℃ 75℃ 400g/min 3800m3/h 15Mpa
The surface oil content result recording milk powder is as follows:
The docosahexaenoic acid grease used Surface oil content (wt%)
Commercially available docosahexaenoic acid grease (dialycerides content 4wt %) 0.68
The docosahexaenoic acid grease that means one prepare 0.42
The docosahexaenoic acid grease that means two prepare 0.39
By above experimental data it can be seen that under equal process conditions, by the present invention containing DHA Microcapsules obtained by microbial oil and milk powder, its surface oil content is lower.This is primarily due to: the microbial oil of the present invention contains Having more diglyceride, it can help microbial oil to form more stable emulsion, makes microbial grease preferably be wrapped Bury, thus reduce the surface oil content of microcapsules and milk powder, improve microcapsules and the oxidation resistance of milk powder, extend microcapsules and The shelf life of milk powder.
Embodiment three
With Pythium ultimum for the bacterial classification that sets out, describe production and the application of the microbial oil containing eicosapentaenoic acid in detail.
1. fermentation
Preparation cane sugar content is 0.05g/mL and culture medium solution that dusty yeast content is 0.005g/mL is in 1000ml shaking flask, Can prepare many bottles, access appropriate Pythium ultimum after sterilizing, be placed in 28 DEG C of constant-temperature tables, 180rpm activates.Two are accessed after 2 days Level expands shaking flask and cultivates, and merges shaking flask after 2 days, and moving into sterilized, cane sugar content is 0.05g/mL and dusty yeast content 1m for 0.005g/mL3In fermentation tank (first class seed pot), it is continually fed into filtrated air, keeps cultivation temperature 28 ± 1 DEG C.One Level seed tank culture after 2 days, move whole nutrient solution enter sterilized, cane sugar content is 0.05g/mL and dusty yeast content is The 10m of 0.005g/mL3In fermentation tank (secondary seed tank), it is continually fed into filtrated air, keeps cultivation temperature 28 ± 1 DEG C.Two grades After seed tank culture 1 day, move whole nutrient solution enter sterilized, cane sugar content is 0.05g/mL and dusty yeast content is The 45m of 0.005g/mL3In fermentation tank, it is continually fed into filtrated air, keeps cultivation temperature 28 ± 1 DEG C, according to sugar consumption speed Batch adds the sterilized sugar solution of total about 0.02 ~ 0.04g/mL, can obtain tunning, wherein 20 carbon after being further cultured for 5 days Pentaene acid content 207.8mg/L.
2. preparation is rich in the microbial oil of eicosapentaenoic acid
Means one
Tunning is realized separation of solid and liquid by centrifugal or press filtration mode, collects wet thallus, utilize pulverizer to crush, then pass through Fluidized drying tower is dried, and obtains dry mycelium.Dry mycelium is mixed with organic solvent such as butane or hexane immersion extraction, after filtration To miscella.
Adding commercially available lipase in miscella and water carries out enzymolysis processing, this commercially available lipase has glyceride hydrolysis Function.Described enzymolysis processing parameter includes: lipase consumption is the 0.25wt% of microorganism miscella weight；Water consumption is The 30wt% of microorganism miscella weight, reaction temperature about 37 DEG C, stirs reaction time 0.5h.Enzyme digestion reaction terminate after to mixing Oil is purified, and the concrete steps of purifying include: stood by miscella, after oil phase is layered with aqueous phase, removes water layer, crosses and filter Remove enzyme, evaporate desolvation, remove free fatty by molecular distillation equipment, obtain microorganism crude oil.This crude oil have as Lower index feature: content of polyunsaturated fatty acid 59.8wt%, TG content 85.1wt%, DG content 8.5wt%.
Further, refining mentioned microorganism crude oil, refined step includes: by this microorganism crude oil warp 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation, use 5wt% titanium dioxide after filtration again Silicon and 3wt% activated carbon decolorizing, after again filtering, live (open) steam deodorization under conditions of 200 DEG C, add Vc palmitate and Ve As antioxidant, obtain the microorganism refined oil containing eicosapentaenoic acid.After measured, polyunsaturated fat in this refined oil It is 87.5wt % that acid total content reaches 59.4wt %, TG content, and DG content is 10.4wt %.
The part physical and chemical index of this microorganism refined oil of mensuration further: 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 Acid 0.06wt%, AFB1< 5.0 μ g/kg, total arsenic (in terms of As) < 0.1mg/kg, lead < 0.1 mg/kg.
Means two
Tunning is realized separation of solid and liquid by centrifugal or press filtration mode, collects wet thallus, utilize pulverizer to crush, then pass through Fluidized drying tower is dried, and obtains dry mycelium.Dry mycelium is mixed with organic solvent such as butane or hexane immersion extraction, after filtration To miscella.
The mixture containing diglyceride, such as aliphatic acid list diacylglycerol or its analog, this mixing is added in miscella In thing, diacylglycerol content is 56.1wt%, and the proportion of the mixture of interpolation is the 15.0wt% of total miscella.To miscella precipitation, Microorganism crude oil is obtained after mixing.This crude oil has a following index feature: content of polyunsaturated fatty acid 45.0wt%, TG Content 82.0wt%, DG content 9.9wt%.
Further, refining mentioned microorganism crude oil, refined step includes: by this microorganism crude oil warp 2.5wt% citric acid and 5wt% hot water degumming, excess sodium hydroxide solution depickling, sedimentation separation, use 5wt% titanium dioxide after filtration again Silicon and 3wt% activated carbon decolorizing, after again filtering, live (open) steam deodorization under conditions of 200 DEG C, add Vc palmitate and Ve As antioxidant, obtain the microorganism refined oil containing eicosapentaenoic acid.After measured, polyunsaturated fat in this refined oil It is 84.3wt % that acid total content reaches 44.9wt %, TG content, and DG content is 11.9wt %.Other physical and chemical indexs obtain with means one The physical and chemical index obtained is close.
3. application
The above-mentioned microbial oil containing eicosapentaenoic acid prepares microcapsules
Use the eicosapentaenoic acid oil that commercially available eicosapentaenoic acid means one oily, above-mentioned, means two prepare, according to joining as follows Material table carries out dispensing:
Title Ratio (wt%) in solid content
Eicosapentaenoic acid lipid 14
Maltodextrin 32
Casein sodium 2.5
Sodium ascorbate 1.5
Pure water 50
By above-mentioned feed liquid after the rotating speed down cut 10min of 8000rpm, under 40MPa, carry out homogeneous, obtain emulsion.By this Emulsion is spray-dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Atomisation pressure
220℃ 90℃ 300g/min 3000m3/h 20Mpa
The surface oil content result recording microcapsules is as follows:
The eicosapentaenoic acid lipid used Surface oil content (wt%)
Commercially available eicosapentaenoic acid lipid (diacylglycerol content 4wt%) 0.79
The eicosapentaenoic acid lipid that means one prepare 0.51
The eicosapentaenoic acid lipid that means two prepare 0.47
The above-mentioned microbial oil containing eicosapentaenoic acid prepares milk powder
Use the eicosapentaenoic acid oil that commercially available eicosapentaenoic acid means one oily, above-mentioned, means two prepare, according to joining as follows Material table carries out dispensing:
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 the rotating speed down cut 10min of 5000rpm, under 20MPa, carry out homogeneous, To emulsion.It is then passed through three grades of falling film type vacuum concentrators and this emulsion is concentrated into moisture 50%, finally carry out pressure Formula is spray-dried, and spray drying parameters is as follows:
EAT Leaving air temp Charging rate Intake volume Atomisation pressure
190℃ 75℃ 400g/min 3800m3/h 15Mpa
The surface oil content result recording milk powder is as follows:
The eicosapentaenoic acid lipid used Surface oil content (wt%)
Commercially available eicosapentaenoic acid lipid (dialycerides content 4wt %) 0.72
The eicosapentaenoic acid lipid that means one prepare 0.48
The eicosapentaenoic acid lipid that means two prepare 0.44
By above example and experimental data it can be seen that under equal process conditions, by the eicosapentaenoic acid of the present invention
Microcapsules obtained by microbial oil and milk powder, its surface oil content is lower.This is primarily due to: the microorganism of the present invention Oil is containing more diglyceride, and it can help microbial oil to form more stable emulsion, make microbial grease preferably It is embedded, thus reduces the surface oil content of microcapsules and milk powder, improve microcapsules and the oxidation resistance of milk powder, extend micro-glue Capsule and the shelf life of milk powder.

Claims (9)

1. microbial oil, it is characterised in that: the content of its polyunsaturated fatty acid is more than 30wt%, and content of triglyceride is less than 90wt%, diacylglycerol content is not less than 5wt% and not higher than 20 wt %.
Microbial oil the most according to claim 1, it is characterised in that: the content of described diglyceride is not less than 10wt%, The content of described triglycerides is not less than 75 wt %.
Microbial oil the most according to claim 1 and 2, it is characterised in that: described polyunsaturated fatty acid is arachidonic Acid, DHA or eicosapentaenoic acid.
Microbial oil the most according to claim 1 and 2, it is characterised in that: described microbial oil is crude oil.
5. the preparation method of the microbial oil described in claim 1, it is characterised in that: comprise the following steps:
(1) oleaginous microorganism fermentation is utilized to obtain the zymotic fluid rich in PUFA microbial oil；
(2) collecting the thalline rich in PUFA microbial oil, extraction obtains miscella after filtering；
(3) in miscella, add the mixture containing diglyceride, remove solvent after mixing and obtain microbial oil.
The preparation method of microbial oil the most according to claim 5, it is characterised in that: step (3) is added containing sweet In the mixture of oil diester, the content of diglyceride is 31.4%-72.2%, and the proportion adding mixture is total miscella 11.5%-22.6%。
The preparation method of microbial oil the most according to claim 5, it is characterised in that: to microorganism after step (3) Oil refines.
8. microcapsules, it is characterised in that: these microcapsules include the arbitrary described microbial oil of claim 1-4.
9. milk powder, it is characterised in that: this milk powder includes the arbitrary described microbial oil of claim 1-4.
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