WO2021056357A1

WO2021056357A1 - Method for preparing astaxanthin oil by cold pressing haematococcus pluvialis

Info

Publication number: WO2021056357A1
Application number: PCT/CN2019/108299
Authority: WO
Inventors: 张吉和; 沈家会; 王兴勇; 张勇
Original assignee: 云南爱尔康生物技术有限公司
Priority date: 2019-09-26
Filing date: 2019-09-26
Publication date: 2021-04-01

Abstract

A method for preparing natural Haematococcus pluvialis astaxanthin oil without any addition, particularly relating to a cold pressing method after pretreatment of Haematococcus pluvialis. The method sequentially comprises the following steps of: A) cleaning; B) wet grinding; C) homogenizing and wall breaking; D) spraying and drying; E) conditioning and granulating; F) cold pressing; and G) settling and fine filtering. A citrate buffer solution is applied to conditioning and granulating of wall-broken Haematococcus pluvialis powder, the complexation, sterilization, and color protection effects are obvious, a cold pressing oil path is smooth without material leakage and slippage, the comprehensive recovery rate and oil yield of astaxanthin are high, and the astaxanthin oil satisfies all quality requirements for edible oil and fat.

Description

Method for preparing astaxanthin oil by cold pressing Haematococcus pluvialis

Technical field

The invention relates to a method for preparing astaxanthin oil after pretreatment of Haematococcus pluvialis, in particular to a method for preparing astaxanthin oil by cold pressing after pretreatment of Haematococcus pluvialis.

Background technique

The freshwater green flagellate algae Haematococcus pluvialis can accumulate 6% of astaxanthin, which makes it the best natural resource for the production of astaxanthin. In natural and cultivated environments, this kind of algae exists in three forms: 1) Motile, bare biflagellate state; 2) Immobile, naked indeterminate population state; 3) Immobile, thick-walled static spores status. In the spore stage of its life cycle, it accumulates astaxanthin in the form of 3S, 3′S isomers in the cytoplasm around the nucleus, and is connected to C16:0, C18:0, C16:0, C18:0, and C16:0, C18:0, in the form of mono- or di-ester C18:1, C18:2 fatty acids.

The most suitable growth temperature for this kind of algae is 15°C to 20°C. It is dominant in the logarithmic growth phase of the batch culture stage, but it can form static spores that synthesize astaxanthin in the stable growth stage. The changes in cell morphology-rapidly dividing, shear-sensitive flagellar cells to very strong, immobile static spores-mean that they generally favor a two-stage batch growth process. In this process, the astaxanthin-free flagellar cells are synthesized preferentially to obtain the maximum biomass, and then the cells are forced to reduce the formation of flagella and accumulate astaxanthin to the greatest extent.

Haematococcus pluvialis can grow in photosynthesis autotrophically, concurrently or heterotrophically, using organic carbon sources such as acetate. Heterotrophic growth cells grow slowly and produce only a small amount of astaxanthin in dark conditions. In the concurrent growth, the addition of acetate and pyruvate can promote cell growth and the formation of astaxanthin.

A large number of "stress" factors, such as nutrient element limitation (especially limiting nitrogen sources), strong light, high temperature or the addition of sodium chloride, promote the synthesis of astaxanthin.

At present, the fully enclosed photobioreactor has completed the production process of its large-scale chemical plant. Strong light is given to the bubble column reactor, flat plate reactor, tubular bioreactor, water ditch or tubular photobioreactor filled with nutrients. And reduce the source of nitrogen and phosphorus, can accumulate a large amount of astaxanthin. The entire growth cycle may take several weeks. Finally, the product is obtained through enzymatic hydrolysis, mechanical crushing, fixation and spray drying. The static spores rich in astaxanthin can be used for pigment accumulation in aquatic animals or as nutrients for humans.

Astaxanthin, also known as astaxanthin and astaxanthin, is a keto carotenoid. It is a terpene unsaturated compound. Its chemical name is 3,3-dihydroxy-4,4-diketo-β, β-carotene has a molecular formula of C ₄₀ H ₅₂ O ₄ and a relative molecular mass of 596.86. There are two forms of astaxanthin in free state and esterified state. The esterified astaxanthin is divided into astaxanthin monoester and astaxanthin diester according to the number of hydroxyl groups bound by its fatty acid. Haematococcus pluvialis shrimp 75% of astaxanthin is astaxanthin monoester and 25% is astaxanthin diester.

Visible light has little effect on astaxanthin, while ultraviolet light has a great destructive effect on astaxanthin. Temperatures below 70°C have little effect on astaxanthin. Astaxanthin is destroyed by heat above 70°C; when the pH is in the range of 4-11, shrimp Astaxanthin is relatively stable. When pH<3 or pH>13, astaxanthin begins to degrade; metal ions such as calcium, sodium, magnesium, potassium, and zinc have basically no effect on astaxanthin. Ferrous ion, iron ion, and copper ion have no effect on astaxanthin. It has obvious destructive effect.

Astaxanthin is by far the strongest antioxidant found in nature by humans. Its antioxidant effect is mainly due to the relatively active electronic effects of the conjugated double bond chain in the molecular structure of astaxanthin and its unsaturated ketone and hydroxyl groups. Attract unpaired electrons of free radicals or provide electrons to free radicals, so as to achieve the purpose of scavenging free radicals.

As a fat-soluble pigment, astaxanthin not only has a bright red color, but also has strong antioxidant properties, and is widely used in food, health products, feed additives, and cosmetics industries.

The large-scale cultivation of Haematococcus pluvialis accumulates astaxanthin, which is mainly achieved through metabolic regulation. Common metabolic regulation methods include stress conditions such as strong light, high temperature, nutrient (nitrogen and phosphorus) starvation, and high salt. These conditions are also the basic conditions for the accumulation of fat in single-celled microalgae. Therefore, Haematococcus pluvialis accumulates astaxanthin, at the same time accumulates oil, extracts astaxanthin, and extracts oil at the same time, it is difficult to separate the two.

Common methods for extracting astaxanthin from Haematococcus pluvialis are: oil dissolution method, solvent extraction method, supercritical extraction method, subcritical extraction method, water media method and other methods.

The astaxanthin of Haematococcus pluvialis is extracted by the oil dissolution method, and astaxanthin is extracted from edible oils, mainly vegetable oils, such as soybean oil, sunflower oil, and linseed oil. A large amount of added edible oil cannot be concentrated, which brings new safety and health risks, and reduces the active components of the extracted astaxanthin oil, restricts its application, and increases the difficulty of subsequent purification and processing.

Solvent extraction methods commonly used extraction solvents include acetone, methanol, ethanol, isopropanol, petroleum ether, hexane, ethyl acetate, dichloromethane, acetonitrile and other single solvents or their mixed solvents. Most of these organic solvents have a certain degree of toxicity. Although they can be volatilized by vacuum distillation, the extracted astaxanthin oil inevitably has residues, and the extracted astaxanthin oil should be refined to meet the food requirements, which increases the safety and health of the astaxanthin oil. risk.

The supercritical extraction method uses CO ₂ supercritical fluid, adding ethanol, edible vegetable oil, etc. as entrainers, and extracts astaxanthin oil above 30MPa and above 60°C. The purity of astaxanthin oil has been improved, and solvent residues have been reduced. However, the extracted astaxanthin oil still contains foreign entrainer residues. The specific flavor of astaxanthin oil is missing, and the initial investment in equipment is large, the production technology requirements are high, and the industrial scale production is still There are certain difficulties.

The subcritical extraction method uses subcritical fluids such as butane and dimethyl ether, adding ethanol and edible vegetable oil as entrainers, and extracts astaxanthin oil below 2MPa and below 60°C. Compared with the supercritical extraction method, the initial investment in equipment and the requirements of production technology, industrial-scale production has made significant improvements, but its essential problems still exist, and they are only alleviated to a certain extent.

The water media method uses water as the medium to transfer the astaxanthin in Haematococcus pluvialis into the water phase, demulsification, extraction, and desolvation to produce astaxanthin oil, which improves the extraction conditions and reduces the water-soluble impurities of astaxanthin oil. However, the residual organic solvent is not improved compared with the solvent extraction method.

Other methods for extracting astaxanthin from Haematococcus pluvialis are mainly to strengthen the pretreatment methods of Haematococcus pluvialis wall breaking, such as hot acid, high-pressure homogenization, bead milling, microwave, enzymatic hydrolysis, etc., to obtain the product astaxanthin oil. Have a substantial effect.

Chinese Invention Patent CN 102492544 B The present invention provides a dry method for preparing DHA microalgae oil, which is carried out according to the following steps: a) Broken the microalgae fermentation broth to obtain the broken fermentation broth; b) The broken fermentation broth is treated with a flocculant to flocculate and aggregate the broken mycelium and grease. After solid-liquid separation, a separated solid is obtained; c) The separated solid is granulated, After drying and crushing, it is extracted with an organic solvent or physically pressed to obtain DHA microalgae oil. The process flow is: Chrysanthemum spp.-wall breaking-flocculation-solid-liquid separation-granulation-drying-organic solvent extraction / physical pressing-DHA algae oil. Obviously, organic solvent extraction is prone to produce organic solvent residues, which brings safety and health risks; physical pressing has two processes, hot pressing and cold pressing. The high temperature conditions of hot pressing are likely to cause degradation of the active ingredients of microalgae and introduce unsafe hidden dangers. The solid state of Chrysomyces sclerophylla was dried at 60°C to a moisture content of less than 5%, crushed into pellets, and cold pressed, without the support of the examples.

Chinese invention patent application 104263770 A discloses a semi-continuous stage fermentation and flocculation plate-and-frame dry method for preparing DHA from Schizochytrium sp., which is carried out according to the following steps: a) Using Schizochytrium as the strain, adopts semi-continuous, staged Formula fermentation method, prepare the Schizochytrium fermented broth; b) Adjust the prepared DHA fermentation broth to pH 5-6, temperature 25-30°C, stirring speed 100-120r/min, and then add flocculant for bacterial growth Flocculation reaction; c) The flocculated Schizochytrium fermented liquid is subjected to plate and frame filtration, the filter cake is crushed and granulated, and fast flashed to obtain DHA microalgae powder; d) DHA microalgae powder is extracted by puffing and pressing to extract DHA-rich oil. The technological process is: Schizochytrium algae-preparation-flocculation-filtration-crushing-granulation-flashing-expansion-squeezing-DHA algae oil. Puffing and pressing are generally squeezing and puffing and then pressing. The extrusion process is a high temperature and high pressure process, and the oil is prone to produce harmful substances such as trans fatty acids and grease polymers.

The traditional oil production process of oil crops includes hot pressing and solvent extraction. Because the macromolecular nutrients (protein, starch, dietary fiber, etc.) in the cake after the above process is used to make oil, there are different degrees of denaturation or the cake contains organic Solvents are only used for feed processing or fertilizers without high value-added utilization, causing serious waste of resources. With the improvement of people's awareness of health and comprehensive utilization of resources, they have begun to explore high-tech for the simultaneous production of high-quality vegetable oils and low-denaturing nutrients in cakes to achieve the comprehensive utilization of oil crops. Technical personnel at home and abroad have proposed cold pressing technology to solve the above problems, and carried out a lot of research around this technology, including cold pressing oil and low-denaturing protein technology, enzymatic oil and protein hydrolysate technology, enzymatic method Some research results have been successfully applied and promoted in the research of pretreatment cold pressing oil production technology and other researches, resulting in greater economic, social and ecological benefits.

Cold pressing oil production technology is a kind of oil and cakes that are directly pressed by a low-temperature oil press at room temperature to 65°C without rolling embryos or steamed to obtain oils and cakes with no change in molecular structure and nutritional value. Oil technology. The mechanical principle is that due to the propelling action of the rotating screw shaft in the press chamber, the press material is continuously pushed forward. Due to the shortening of the screw pitch on the screw shaft and the increase of the root circle diameter, as well as the reduction of the inner diameter of the press chamber. Small, so that the volume of the squeezing chamber is continuously reduced, which has a squeezing effect on the squeezing material. After the squeezed material is compressed, the oil is squeezed out from the gap of the squeezing cage, and at the same time, the squeezed material is pressed into cakes and discharged from the end of the pressing chamber.

The cold-pressing method of oil production is a physical method, which is pressurized without heating, and has no effect on oils and nutrients. At the same time, in addition to the general characteristics of ordinary oil-making processes, this process can also improve the quality of oils, avoid the production of harmful substances such as trans fatty acids and oil polymers due to high-temperature processing, and retain the active substances in the oil. Taking cold-pressed peanut oil as an example, cold-pressed oil can avoid the residual problems of acids, alkalis, heavy metals and other harmful substances caused by the addition of chemical additives during the refining process, while shortening the processing technology and saving 1/3 of the production. Cost, reducing project investment costs, and enhancing the market competitiveness of the product; in addition, the nutritional value of peanut meal after pressing has been improved, the nutrients such as protein and dietary fiber have not been denatured, and the active substances can be preserved, ensuring the meal’s quality Development and utilization value. Therefore, the technology is suitable for the production and processing of high-quality oils and macromolecular nutrients produced by the simultaneous pressing of oil crops.

Chinese invention patent application CN 104130854 A discloses a method of cold pressing oil, which includes the following steps: 1) cleaning the oil raw material; 2) peeling the shell and separating the shell and kernel; 3) rolling the embryo to obtain the embryo; 4) removing the embryo Soften; 5) Cold press the raw material; 6) Precipitate the raw material, filter the oil, and collect the extract to obtain crude oil; collect the precipitate to obtain the filter cake; 7) Adjust the acid value of the crude oil to pH ≤ 3; 8 ) Centrifuge the crude oil obtained in step (7), and stop the centrifugation after reaching the target; 9) deodorize, and obtain product oil after passing the inspection. The material blank is softened for 30 minutes, the discharging temperature is ≤60°C, and the moisture content in the material blank is 6%-10% when discharging.

Chinese invention patent application CN 10962820 A discloses an ancient physical cold-pressed oil process, which includes the following steps: Step 1, pretreatment: screening the raw materials to remove impurities, peeling, and crushing; Step 2, after pretreatment The raw materials are softened by warm water spray; the partially softened raw materials are fried in an iron pan; the third step is to mix the fried raw materials and the un-fried raw materials in a certain proportion, and spray again with warm water to soften them, and then Press at low temperature to obtain crude oil; step 4, add citral and glucanase in a certain proportion to the crude oil; step 5, at the end of the reaction, heat up to 130°C to inactivate the enzyme and remove the gum; sixth In step 5, the raw oil obtained in step 5 is subjected to filtration and degumming treatment; in step 7, the raw oil tank after filtration and degumming is installed in an iron tank.

The oil cold pressing technology is mainly for traditional oils, such as soybean, peanut, sesame, rapeseed, cottonseed, sunflower seed, linseed, hemp seed, castor bean, corn germ, wheat germ, rice bran, tea seed, tung seed, coconut, Palm fruit, Chinese tallow seeds and other herbal oils and woody oils. Haematococcus pluvialis is a single-celled microalgae and is rich in natural astaxanthin. Using cold pressing technology to prepare astaxanthin oil from Haematococcus pluvialis is still a technical gap.

Summary of the invention

In view of the deficiencies or deficiencies of the prior art, the problem to be solved by the present invention is to provide a method for preparing natural Haematococcus pluvialis astaxanthin oil without any addition, especially when it involves the pretreatment of Haematococcus pluvialis. The method of producing astaxanthin oil by cold pressing method.

To solve this problem, in the first aspect of the present invention, a method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis is provided, which includes the following steps: A) washing, and natural sedimentation of the harvested Haematococcus pluvialis liquid, Remove the supernatant and collect the underflow of Haematococcus pluvialis; B) Grind, grind the underflow of Haematococcus pluvialis to obtain ground Haematococcus pluvialis algae slurry; C) homogeneously break the wall, and grind the red pluvialis The algae slurry is homogeneously broken to the point where no intact cells can be seen to obtain a homogeneous broken-wall Haematococcus pluvialis algae slurry; D) Spray drying, the homogeneous broken-wall Haematococcus pluvialis algae slurry is spray-dried, and the dried rain is collected Haematococcus pluvialis powder; E) tempering and granulation, spraying buffer solution into dry Haematococcus pluvialis powder, adjusting the moisture content of Haematococcus pluvialis powder, mixing, shaking granulation to obtain Haematococcus pluvialis pellets F) cold pressing, cold-pressing Haematococcus pluvialis pellets to obtain cold-pressed astaxanthin crude oil; and G) sedimentation fine filtration, where the cold-pressed astaxanthin crude oil is sedimented and filtered to obtain astaxanthin oil.

Preferably, the pH of the buffer solution sprayed in step E) conditioning granulation is>7.

Preferably, the buffer solution sprayed in step E) conditioning and granulation is a citrate buffer solution.

Preferably, the concentration of the substance in the buffer solution sprayed in step E) conditioning granulation is 0.2 mol/L to 0.4 mol/L.

Preferably, the water content of the pellets of Haematococcus pluvialis prepared in step E) tempering granulation is 6.0%-8.0%.

Preferably, the particle size of the pellets of Haematococcus pluvialis prepared in step E) tempering granulation is 5 mesh to 20 mesh.

Preferably, the cold pressing temperature in step F) is 50°C-60°C.

In the second aspect of the present invention, there is also provided astaxanthin oil prepared by the method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to the first aspect.

The beneficial effects of the present invention are embodied in:

1) Combining cell crushing technology and oil cold pressing technology to prepare natural Haematococcus pluvialis astaxanthin oil without any additions, using citrate buffer solution to refine and granulate the broken Haematococcus pluvialis powder , The complexation, sterilization, and color protection effects are remarkable, the comprehensive recovery rate of astaxanthin is as high as 98.0%, and the astaxanthin oil fully meets all the quality requirements of edible oils and fats.

2) Combine the oil hydration and degumming technology with the oil softening and conditioning technology. It is easy to operate without heating at room temperature and pressure. The weakly alkaline citric acid buffer solution with pH 7-9 wets Haematococcus pluvialis algae powder and releases it by breaking the wall. Haematococcus pluvialis single-cell protein, phospholipids, etc. absorb water, soften and swell, gel solidify, and have obvious oil separation effect. The cold-pressed oil circuit of Haematococcus pluvialis is improved, and the oil output rate is as high as 80.0%.

3) Citrate buffer solution is composed of citric acid and sodium citrate, adding salt, adding osmotic capacity, the oil separation of Haematococcus pluvialis powder is further improved, the oil yield is increased by 1% to 2%, the amount of suitable substances Concentration, the cold pressing performance of Haematococcus pluvialis has been further improved, and the oil yield can also be increased by 1% to 2%, reaching a cumulative amount of 82% to 84%.

4) Control the moisture, particle size and cold pressing temperature of Haematococcus pluvialis pellets. The cold pressing of Haematococcus pluvialis is smooth, without material leakage, no slippage, smooth cake and oil output, and production efficiency meets the design requirements.

Description of the drawings

Figure 1 is a process flow diagram of preparing astaxanthin oil by cold pressing of Haematococcus pluvialis.

detailed description

It should be understood that different applications of the method disclosed herein can be changed according to specific needs in the field. It should also be understood that the terms used herein are only for describing specific embodiments of the present invention, and are not intended to be limiting.

The present invention provides a method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis. The method includes the following steps: A) washing; B) grinding; C) homogeneous wall breaking; D) spray drying; E) quenching and tempering Granules; F) cold pressing; G) sedimentation fine filtration.

The Haematococcus pluvialis used in the method of the present invention can be cultured using existing methods known in the art, and there is no particular limitation on this.

In some embodiments, in step A) cleaning, the natural sedimentation time of the harvested Haematococcus pluvialis liquid is subject to clear stratification, usually 0.5 hour to 5.0 hours, such as 0.5 hour, 1.0 hour , 1.5 hours, 2.0 hours, 2.5 hours, 3.0 hours, 3.5 hours, 4.0 hours, 4.5 hours, 5.0 hours, or the range and value between any of the above points, preferably 1.0 to 3.0 hours, more preferably 1.5 hours to 2.5 Hours, most preferably 2.0 hours.

In some embodiments, the supernatant includes a Haematococcus pluvialis culture solution, and autolyzed cells floating therein.

In some embodiments, pure water can be added to the underflow of Haematococcus pluvialis obtained after washing for slurry adjustment, wherein the amount of pure water added is suitable for uniform slurry adjustment, followed by natural sedimentation to remove the upper The clear liquid collects the underflow of Haematococcus pluvialis. The time for natural settlement can be as described above, subject to clear stratification, usually 0.5 hours to 5.0 hours, such as 0.5 hours, 1.0 hours, 1.5 hours, 2.0 hours, 2.5 hours, 3.0 hours, 3.5 hours, 4.0 Hours, 4.5 hours, 5.0 hours, or the range and value between any of the above points, preferably 1.0 to 3.0 hours, more preferably 1.5 hours to 2.5 hours, and most preferably 2.0 hours. The above steps of adding pure water, performing slurry adjustment, natural sedimentation, removing the supernatant, and collecting the underflow of Haematococcus pluvialis can be repeated many times, for example, once, twice, 3 times, 4 times, 5 times or more. Waited many times.

In some embodiments, in step B) grinding, conventional methods in the art can be used to grind using a common grinder in the art, for example, wet grinding, or colloid milling, for example, to obtain ground Haematococcus pluvialis Algae pulp. The grinding time is usually 0.5 hour to 2.0 hours, such as 0.5 hour, 1.0 hour, 1.5 hour, 2.0 hour, or the range and value between any of the above points. In some embodiments, the obtained ground Haematococcus pluvialis algae slurry can be returned to a grinding machine such as a colloid mill for grinding, and the grinding step can be repeated multiple times, for example, 1 time, 2 times, 3 times. , 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 11 times, 12 times or more, etc.

In some embodiments, in step C) homogeneous wall breaking, conventional methods in the art can be used to homogenize the wall using a common homogenizer in the art, for example, a high-pressure homogenizer is used to homogenize the wall to obtain Homogeneous broken wall Haematococcus pluvialis algae slurry. The solid content can be adjusted before homogeneous wall breaking, for example, in the range of 20% to 30%, such as 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29 %, 30%, or the range and value between any of the above points. The homogeneous wall breaking time is usually 2.0 hours to 6.0 hours, such as 2.0 hours, 2.5 hours, 3.0 hours, 3.5 hours, 4.0 hours, 4.5 hours, 5.0 hours, 5.5 hours, 6.0 hours, or the range between any of the above Numerical value. The pressure used in homogeneous wall breaking is, for example, 80-120 MPa, such as 80 MPa, 85 MPa, 90 MPa, 95 MPa, 100 MPa, 105 MPa, 110 MPa, 115 MPa, 120 MPa, or a range and value between any of the above points.

In some embodiments, the obtained homogeneous wall-broken Haematococcus pluvialis algae slurry can be returned to a homogenizer such as a high-pressure homogenizer for homogeneous wall breaking, and the homogeneous wall breaking step can be repeated multiple times It is performed, for example, once, twice, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 11 times, 12 times or more. In the process of homogeneous wall breaking, microscopic examination can be performed until intact cells are not visible in the end.

In some embodiments, in step D) spray drying, conventional methods in the art can be used to spray drying using a spray drying system commonly used in the art to collect dried Haematococcus pluvialis powder. When spray drying is performed, the inlet air temperature is, for example, in the range of 160°C to 180°C, such as 160°C, 165°C, 170°C, 175°C, 180°C, or a range and value between any of the above points. The temperature of the outlet air is in the range of, for example, 70°C to 90°C, such as 70°C, 75°C, 80°C, 85°C, 90°C, or a range and value between any of the above points.

In some embodiments, in step E) conditioning and granulation, a conventional method in the art may be used to spray a buffer solution using a spray system commonly used in the art. The buffer solution has a pH>7, for example, a pH of 7-9, for example a pH of 7, 8, or 9.

In some embodiments, the buffer solution may be a common buffer solution in the art, for example, a citrate buffer solution. The citrate buffer solution is usually prepared by dissolving citric acid, sodium citrate and/or sodium chloride in water.

In some embodiments, the amount of the substance in the buffer solution is 0.2 mol/L to 0.4 mol/L, such as 0.2 mol/L, 0.25 mol/L, 0.3 mol/L, 0.35 mol/L, 0.4 mol/L Wait. For example, in the citrate buffer solution, the total amount of citric acid, sodium citrate and sodium chloride is 0.2mol/L～0.4mol/L, such as 0.2mol/L, 0.25mol/L, 0.3mol/L L, 0.35mol/L, 0.4mol/L, etc.

In some embodiments, in step E) tempering and granulation, a conventional method in the art can be used for granulation using a granulator commonly used in the art.

In some embodiments, the prepared Haematococcus pluvialis pellets have a moisture content of 6.0% to 8.0%, such as 6.0%, 6.5%, 7.0%, 7.5%, 8.0%, or a range between any of the above points And value.

In some embodiments, the particle size of the prepared Haematococcus pluvialis pellets ranges from 5 mesh to 20 mesh, such as 5 mesh, 10 mesh, 15 or 20 mesh, or a range and value between any of the above points.

In some embodiments, in step F) cold pressing, a conventional method in the art may be used to perform cold pressing using a cold press commonly used in the art. For example, a twin-screw cold press is used for cold pressing to obtain astaxanthin crude oil produced by cold pressing. The cold pressing temperature is, for example, 40°C to 70°C, preferably 45°C to 65°C, more preferably 50°C to 60°C, such as 50°C, 51°C, 52°C, 53°C, 54°C, 55°C, 56°C, 57°C, 58°C, 59°C or 60°C, or a range and value between any of the above points.

In some embodiments, in step G) sedimentation and fine filtration, the cold-pressed astaxanthin-producing crude oil can be left to settle for 7 to 10 days, for example, 7 days, 8 days, 9 days or 10 days, or it can be determined by techniques in the art The personnel routinely determine the settlement time.

In some embodiments, in step G) sedimentation and fine filtration, conventional methods in the art can be used to perform fine filtration using common equipment in the art, such as pressure filtration with 0.2 μm filter membrane plates and frames to obtain astaxanthin oil.

Example

The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to the present invention will be described in further detail below in conjunction with certain embodiments of the present invention. These embodiments are conducive to a better understanding of the present invention, but do not limit the present invention. .

Example 1

Harvest 100t of Haematococcus pluvialis liquid, settling naturally for 2 hours, collect the underflow, remove the floating autolysed cells and culture fluid in the upper layer, add pure water to the underflow to adjust the slurry, settle naturally for 2 hours, collect the underflow, and repeat twice. The underflow of Haematococcus pluvialis is transferred to the colloid mill for grinding, and the ground algae slurry is returned to the colloid mill for repeated grinding, repeated 8 times. Grind Haematococcus pluvialis algae slurry to adjust the solid content to 25%, transfer it to a high-pressure homogenizer to homogenize the wall, homogenizing pressure 100MPa, and return the homogenized algae slurry to the high-pressure homogenizer to repeat the homogenization, microscopic inspection, until no Intact cells are seen, homogenize 5 times. The broken wall Haematococcus pluvialis algae slurry is transferred to the spray drying system for spray drying, the inlet temperature is 170℃～175℃, the outlet temperature is 80℃～85℃, and 100kg of dried haematococcus pluvialis powder is collected, containing 3.5 astaxanthin. %, crude fat 32.2%. Take all the dried Haematococcus pluvialis powder, put it into the tank mixer in batches, spray citrate buffer solution, the citrate buffer solution is made of 0.1moL/L sodium citrate (Na ₃ C ₆ H ₅ O ₇ · 2H ₂ O) solution, add 0.1moL/L citric acid (C ₆ H ₈ O ₇ ·H ₂ O) solution to adjust pH 7.0 to make, mix well, adjust the moisture content of Haematococcus pluvialis powder, and knead it by hand. The pellets are loosened and then dispersed. The pellets of Haematococcus pluvialis are prepared by swing granulation with a 5-mesh standard sieve. The pellets are tested and the water content is 6.0%. The pellets of Haematococcus pluvialis are put into the hopper of the twin-screw cold press, the temperature of the press chamber is set to 50 ℃, and the astaxanthin crude oil is obtained by cold pressing. The obtained cold-pressed crude oil produced astaxanthin was packed in a stainless steel barrel, covered with a lid, covered with a PE bag, tightened, and allowed to settle naturally for 7 days. Filtered with a 0.2 μm filter membrane frame to obtain 25.76 kg of astaxanthin oil, containing 10.71% of astaxanthin, and 74.14 kg of Haematococcus pluvialis cake, containing 0.90% of astaxanthin. The mass percentage of astaxanthin oil in the crude fat is the oil yield, and the oil yield is 80.0%. The mass percentage of astaxanthin in astaxanthin oil and cakes accounts for the mass percentage of astaxanthin in Haematococcus pluvialis powder. The comprehensive recovery rate is 98.0%. Take the astaxanthin oil, send it for inspection, batch number LZY2019001, the conclusion is qualified.

Example 2

Harvest 100t of Haematococcus pluvialis liquid, settling naturally for 2 hours, collect the underflow, remove the floating autolysed cells and culture fluid in the upper layer, add pure water to the underflow to adjust the slurry, settle naturally for 2 hours, collect the underflow, and repeat twice. The underflow of Haematococcus pluvialis is transferred to the colloid mill for grinding, and the ground algae slurry is returned to the colloid mill for repeated grinding, repeated 8 times. Grind Haematococcus pluvialis algae slurry to adjust the solid content to 25%, transfer it to a high-pressure homogenizer to homogenize the wall, homogenizing pressure 120MPa, and return the homogenized algae slurry to the high-pressure homogenizer to repeat the homogenization, microscopic inspection, until no Intact cells are seen, homogenize 4 times. The broken wall Haematococcus pluvialis algae slurry is transferred to the spray drying system for spray drying, the inlet temperature is 170℃～175℃, the outlet temperature is 80℃～85℃, and 120kg of dried haematococcus pluvialis powder containing astaxanthin 3.3 is collected. %, crude fat 33.2%. Take all the dried Haematococcus pluvialis powder, put it into the tank mixer in batches, spray citrate buffer solution, the citrate buffer solution is made of 0.1moL/L sodium citrate (Na ₃ C ₆ H ₅ O ₇ · 2H ₂ O) solution, add 0.1moL/L citric acid (C ₆ H ₈ O ₇ ·H ₂ O) solution to adjust pH 9.0 to make, mix well, adjust the water content of Haematococcus pluvialis powder, and knead it by hand The pellets are loosened when loosened. The pellets of Haematococcus pluvialis are prepared by swing granulation with a 20-mesh standard sieve. The pellets are inspected and have a water content of 8.0% and oil stains on the surface. The pellets of Haematococcus pluvialis are put into the hopper of the twin-screw cold press, and the temperature of the press chamber is set to 60°C, and the astaxanthin crude oil is obtained by cold pressing. The obtained cold-pressed crude oil produced astaxanthin is packed in a stainless steel barrel, covered with a lid, covered with a PE bag, tightened, and allowed to settle naturally for 10 days. Filter with a 0.2 μm filter membrane frame to obtain 31.91 kg of astaxanthin oil, containing 9.75% of astaxanthin, and 88.0 kg of Haematococcus pluvialis cake, containing 0.88% of astaxanthin. The mass percentage of astaxanthin oil in crude fat is the oil yield, and the oil yield is 80.1%. The mass percentage of astaxanthin in astaxanthin oil and cakes accounts for the mass percentage of astaxanthin in Haematococcus pluvialis powder. The comprehensive recovery rate is 98.2%. Take the astaxanthin oil, send it for inspection, batch number LZY2019002, the conclusion is qualified.

Example 3

Harvest 100t of Haematococcus pluvialis liquid, settling naturally for 2 hours, collect the underflow, remove the floating autolysed cells and culture fluid in the upper layer, add pure water to the underflow to adjust the slurry, settle naturally for 2 hours, collect the underflow, and repeat twice. The underflow of Haematococcus pluvialis is transferred to the colloid mill for grinding, and the ground algae slurry is returned to the colloid mill for repeated grinding, repeated 8 times. Grind Haematococcus pluvialis algae slurry to adjust the solid content to 25%, transfer it to a high-pressure homogenizer to homogenize the wall, homogenizing pressure 110MPa, and return the homogenized algae slurry to the high-pressure homogenizer to repeat the homogenization, microscopic inspection, until no Intact cells are seen, homogenize 5 times. The broken wall Haematococcus pluvialis algae slurry is transferred to the spray drying system for spray drying, the inlet temperature is 170℃～175℃, the outlet temperature is 80℃～85℃, and 110kg of dried Haematococcus pluvialis powder is collected, containing 2.88 astaxanthin %, crude fat 35.2%. Take all the dry Haematococcus pluvialis powder and put it into the tank mixer in batches, spray citrate buffer solution, the citrate buffer solution is made up of 0.1moL/L sodium citrate (Na ₃ C ₆ H ₅ O ₇ · 2H ₂ O) and 10g/L sodium chloride solution, add 0.1moL/L citric acid (C ₆ H ₈ O ₇ ·H ₂ O) solution to adjust pH to 8.5, mix and adjust Haematococcus pluvialis powder The water content is kneaded into a dough by hand, and it is loosened when loosened. The pellets of Haematococcus pluvialis are obtained by swinging and granulating with a 20-mesh standard sieve. The pellets are inspected and have a water content of 7.0%, and oil stains can be seen on the surface. The pellets of Haematococcus pluvialis are put into the hopper of the twin-screw cold press, the temperature of the press chamber is set to 55 ℃, and the astaxanthin crude oil is obtained by cold pressing. The obtained cold-pressed crude oil produced astaxanthin is packed in a stainless steel barrel, covered with a lid, a PE bag, tightened, and allowed to settle naturally for 8 days. Filtered with a 0.2 μm filter membrane frame to obtain 28.51 kg of astaxanthin oil, containing 6.50% astaxanthin, and 71.25 kg of Haematococcus pluvialis cake, containing 0.83% astaxanthin. The mass percentage of astaxanthin oil in the crude fat is the oil yield, and the oil yield is 81.0%. The mass percentage of astaxanthin in astaxanthin oil and cakes accounts for the mass percentage of astaxanthin in Haematococcus pluvialis powder. The comprehensive recovery rate is 98.1%.

Example 4

Harvest 100t of Haematococcus pluvialis liquid, settling naturally for 2 hours, collect the underflow, remove the floating autolysed cells and culture fluid in the upper layer, add pure water to the underflow to adjust the slurry, settle naturally for 2 hours, collect the underflow, and repeat twice. The underflow of Haematococcus pluvialis is transferred to the colloid mill for grinding, and the ground algae slurry is returned to the colloid mill for repeated grinding, repeated 8 times. Grind Haematococcus pluvialis algae slurry to adjust the solid content to 25%, transfer it to a high-pressure homogenizer to homogenize the wall, homogenizing pressure 100MPa, and return the homogenized algae slurry to the high-pressure homogenizer to repeat the homogenization, microscopic inspection, until no Intact cells are seen, homogenize 5 times. The broken wall Haematococcus pluvialis algae slurry is transferred to the spray drying system for spray drying, the inlet temperature is 170℃～175℃, the outlet temperature is 80℃～85℃, and 130kg of dried Haematococcus pluvialis powder is collected, containing 3.25 astaxanthin. %, crude fat 36.5%. Take all the dried Haematococcus pluvialis powder, put it into the tank mixer in batches, spray citrate buffer solution, the citrate buffer solution is made of 0.1moL/L sodium citrate (Na ₃ C ₆ H ₅ O ₇ · 2H ₂ O) and 10g/L sodium chloride solution, add 0.1moL/L citric acid (C ₆ H ₈ O ₇ ·H ₂ O) solution to adjust pH to 8.5, mix and adjust Haematococcus pluvialis powder The water content is kneaded into a dough by hand, and it is loosened when loosened. The pellets of Haematococcus pluvialis are obtained by swinging and granulating with a 20-mesh standard sieve. The pellets are inspected and have a water content of 7.0%, and oil stains can be seen on the surface. The pellets of Haematococcus pluvialis are put into the hopper of the twin-screw cold press, the temperature of the press chamber is set to 55 ℃, and the astaxanthin crude oil is obtained by cold pressing. The obtained cold-pressed crude oil produced astaxanthin was packed in a stainless steel barrel, covered with a lid, covered with a PE bag, tightened, and allowed to settle naturally for 9 days. Filter with 0.2 μm filter membrane frame to obtain 38.9 kg of astaxanthin oil, containing 8.74% astaxanthin, and 91.1 kg of Haematococcus pluvialis cake, containing 0.82% of astaxanthin. The mass percentage of astaxanthin oil in the crude fat is the oil yield, and the oil yield is 82.0%. The mass percentage of astaxanthin in astaxanthin oil and cakes accounts for the mass percentage of astaxanthin in Haematococcus pluvialis powder. The comprehensive recovery rate is 98.2%.

Example 5

Harvest 100t of Haematococcus pluvialis liquid, settling naturally for 2 hours, collect the underflow, remove the floating autolysed cells and culture fluid in the upper layer, add pure water to the underflow to adjust the slurry, settle naturally for 2 hours, collect the underflow, and repeat twice. The underflow of Haematococcus pluvialis is transferred to the colloid mill for grinding, and the ground algae slurry is returned to the colloid mill for repeated grinding, repeated 8 times. Grind Haematococcus pluvialis algae slurry to adjust the solid content to 25%, transfer it to a high-pressure homogenizer to homogenize the wall, homogenizing pressure 100MPa, and return the homogenized algae slurry to the high-pressure homogenizer to repeat the homogenization, microscopic inspection, until no Intact cells are seen, homogenize 5 times. The broken wall Haematococcus pluvialis algae slurry is transferred to the spray drying system for spray drying, the inlet temperature is 170℃～175℃, the outlet temperature is 80℃～85℃, and 94.5kg of dried haematococcus pluvialis powder is collected, containing astaxanthin 3.08%, crude fat 32.5%. Take all the dried Haematococcus pluvialis powder, put it into the tank mixer in batches, spray citrate buffer solution, the citrate buffer solution is made of 0.2moL/L sodium citrate (Na ₃ C ₆ H ₅ O ₇ · 2H ₂ O) and 10g/L sodium chloride solution, add 0.2moL/L citric acid (C ₆ H ₈ O ₇ ·H ₂ O) solution to adjust pH to 8.5, mix and adjust Haematococcus pluvialis powder The water content is kneaded into a dough by hand, and it is loosened when loosened. The pellets of Haematococcus pluvialis are obtained by swinging and granulating with a 20-mesh standard sieve. The pellets are inspected and have a water content of 7.0%, and oil stains can be seen on the surface. The pellets of Haematococcus pluvialis are put into the hopper of the twin-screw cold press, the temperature of the press chamber is set to 55 ℃, and the astaxanthin crude oil is obtained by cold pressing. The obtained cold-pressed crude oil produced astaxanthin was packed in a stainless steel barrel, covered with a lid, covered with a PE bag, tightened, and allowed to settle naturally for 9 days. Filter with 0.2 μm filter membrane frame to obtain 25.18 kg of astaxanthin oil, containing 9.28% astaxanthin, and 66.30 kg of Haematococcus pluvialis cake, containing 0.77% of astaxanthin. The mass percentage of astaxanthin oil in the crude fat is the oil yield, and the oil yield is 82.0%. The mass percentage of astaxanthin in astaxanthin oil and cakes accounts for the mass percentage of astaxanthin in Haematococcus pluvialis powder. The comprehensive recovery rate is 98.0%.

Example 6

Harvest 100t of Haematococcus pluvialis liquid, settling naturally for 2 hours, collect the underflow, remove the floating autolysed cells and culture fluid in the upper layer, add pure water to the underflow to adjust the slurry, settle naturally for 2 hours, collect the underflow, and repeat twice. The underflow of Haematococcus pluvialis is transferred to the colloid mill for grinding, and the ground algae slurry is returned to the colloid mill for repeated grinding, repeated 8 times. Grind Haematococcus pluvialis algae slurry to adjust the solid content to 25%, transfer it to a high-pressure homogenizer to homogenize the wall, homogenizing pressure 100MPa, and return the homogenized algae slurry to the high-pressure homogenizer to repeat the homogenization, microscopic inspection, until no Intact cells are seen, homogenize 5 times. The broken wall Haematococcus pluvialis algae slurry is transferred to the spray drying system for spray drying, the inlet air temperature is 170℃～175℃, the outlet temperature is 80℃～85℃, and 109kg of dried haematococcus pluvialis powder is collected, containing 3.18 astaxanthin %, crude fat 33.5%. Take all the dry Haematococcus pluvialis powder, put it into the tank mixer in batches, spray citrate buffer solution, the citrate buffer solution is made of 0.4moL/L sodium citrate (Na ₃ C ₆ H ₅ O ₇ · 2H ₂ O) and 10g/L sodium chloride solution, add 0.4moL/L citric acid (C ₆ H ₈ O ₇ ·H ₂ O) solution to adjust pH to 8.5, mix well, adjust Haematococcus pluvialis powder The water content is kneaded into a dough by hand, and it is loosened when loosened. The pellets of Haematococcus pluvialis are obtained by swinging and granulating with a 20-mesh standard sieve. The pellets are inspected and have a water content of 7.0%, and oil stains can be seen on the surface. The pellets of Haematococcus pluvialis are put into the hopper of the twin-screw cold press, the temperature of the press chamber is set to 55 ℃, and the astaxanthin crude oil is obtained by cold pressing. The obtained cold-pressed crude oil produced astaxanthin is packed in a stainless steel barrel, covered with a lid, a PE bag, tightened, and allowed to settle naturally for 8 days. Filter with 0.2 μm filter membrane frame to obtain 30.66 kg of astaxanthin oil, containing 9.33% astaxanthin, and 78.4 kg of Haematococcus pluvialis cake, containing 0.69% of astaxanthin. The mass percentage of astaxanthin oil in the crude fat is the oil yield, and the oil yield is 84.0%. The mass percentage of astaxanthin in astaxanthin oil and cakes accounts for the mass percentage of astaxanthin input into Haematococcus pluvialis powder. The comprehensive recovery rate is 98.1%.

Although the present invention has been described in detail with reference to certain embodiments of the present invention, these embodiments can be modified or changed without departing from the scope of the present invention. The protection scope of the appended claims should not be limited to the description of the embodiments contained herein, but cover all embodiments falling within the literal or equivalent meaning of the claims.

Claims

The method for preparing astaxanthin oil by cold pressing Haematococcus pluvialis includes the following steps:

A) Cleaning, natural sedimentation of the harvested Haematococcus pluvialis liquid, removing the supernatant, and collecting the underflow of Haematococcus pluvialis;

B) Grinding, grinding the Haematococcus pluvialis underflow to obtain a slurry of ground Haematococcus pluvialis;

C) Homogeneous wall breaking: the ground Haematococcus pluvialis algae slurry is homogenized and broken until no intact cells are visible, to obtain homogeneous wall-broken Haematococcus pluvialis algal slurry;

D) Spray drying, spray drying the homogeneous broken-wall Haematococcus pluvialis algae slurry, and collect dried Haematococcus pluvialis powder;

E) Conditioning and granulating, spraying a buffer solution into the dry Haematococcus pluvialis powder, adjust the moisture content of the Haematococcus pluvialis powder, mix, shake and granulate to obtain Haematococcus pluvialis pellets;

F) Cold pressing, cold pressing Haematococcus pluvialis pellets to obtain cold pressing crude astaxanthin production; and

G) Sedimentation fine filtration, sedimentation of the cold-pressed crude oil produced astaxanthin, and pressure filtration to obtain astaxanthin oil.
The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to claim 1, wherein the pH of the buffer solution sprayed in the step E) conditioning granulation is>7.
The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to claim 2, wherein the buffer solution sprayed in step E) conditioning and granulation is a citrate buffer solution.
The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to any one of claims 1 to 3, wherein the amount and concentration of the substance in the buffer solution sprayed in the step E) tempering granulation is 0.2mol/L～0.4mol/L.
The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to any one of claims 1 to 4, wherein the pellets of Haematococcus pluvialis obtained in step E) tempering granulation The moisture content is 6.0% to 8.0%.
The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to any one of claims 1 to 5, wherein the pellets of Haematococcus pluvialis obtained in step E) tempering granulation The particle size is 5 mesh to 20 mesh.
The method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to any one of claims 1 to 6, wherein the cold pressing temperature in step F) is 50°C-60°C.
Astaxanthin oil prepared by the method for preparing astaxanthin oil by cold pressing of Haematococcus pluvialis according to any one of claims 1-7.