A kind of preparation method of high-content astaxanthin oil

Abstract

The invention discloses a method for preparing high-content astaxanthin oil, and belongs to the technical field of astaxanthin. The method comprises the following steps: (1) spray drying the fermentation broth of Yarrowia lipolytica, wherein the inlet air temperature of the spray drying is 150-160°C and the outlet air temperature is 60-70°C, to obtain a fermentation powder; (2) using a roller press to break the fermentation powder under a roller pressure of 80-120MPa to obtain a sheet material; (3) crushing the sheet material into small sheet materials with a diameter of less than 0.7cm, and then performing supercritical extraction to obtain a high-content astaxanthin oil. The astaxanthin oil prepared by the method of the invention has a high content, a low water content, a high yield of astaxanthin, and is suitable for industrial large-scale production.

Description

Technical Field

[0001] The invention relates to a method for preparing high-content astaxanthin oil, and belongs to the technical field of astaxanthin. Background Art

[0002] Astaxanthin is a ketocarotene and an unsaturated terpene compound. It has 11 conjugated double bonds in its molecular structure and both unsaturated keto and hydroxyl groups at the tail end. Its special molecular structure makes it highly active and can attract free radicals or supply electrons to free radicals to eliminate them, thus showing excellent antioxidant properties and enhancing cell regeneration ability. It is the carotenoid with the strongest antioxidant capacity discovered so far.

[0003] With the discovery of astaxanthin's antioxidant capacity and other biological activities, astaxanthin has gradually been used in products such as animal feed, health foods, cosmetics and medical preparations. At present, the production of astaxanthin mainly includes chemical synthesis, natural extraction and microbial fermentation. Chemically synthesized astaxanthin is not only expensive, but also has significant differences from natural astaxanthin in terms of molecular structure, biological function, application effect and biosafety performance. The natural extraction method mainly extracts astaxanthin from Haematococcus pluvialis, but Haematococcus pluvialis has a long growth time, strict culture conditions, requires light, and limited production sites. In addition, astaxanthin exists in thick-walled spores, with low extraction rate and poor continuity, which is not conducive to large-scale cultivation. The microbial fermentation method mainly uses strains such as Phaffia rhodozyma and Yarrowia lipolytica to ferment and produce astaxanthin. The astaxanthin content in natural Phaffia rhodozyma is low, and its growth is easily affected by fermentation conditions such as temperature, dissolved oxygen, carbon and nitrogen sources, and pH value. It has high technical requirements and complex production processes. Yarrowia lipolytica is an unconventional oleaginous yeast with rich acetyl CoA content. Its own lipid accumulation is conducive to the storage of hydrophobic carotenoids. It can accumulate oils that account for more than 80% of its dry weight, making it an ideal cell factory for heterologous synthesis of carotenoids. In short, heterologous synthesis of astaxanthin using Yarrowia lipolytica has a good market prospect.

[0004] In the existing astaxanthin extraction and purification process, the methods are different, but many problems are also highlighted. Chinese patent CN111875528A discloses a method for preparing astaxanthin oil from Haematococcus pluvialis, in particular, a method for preparing astaxanthin oil by using Haematococcus pluvialis as a raw material, extracting and purifying with alcohol, wherein the method uses a dilute acid solution to break the wall, and stirs and extracts with alcohol, and obtains astaxanthin oil after vacuum distillation. In the process of preparing astaxanthin oil, the use of a dilute acid solution is prone to cause environmental pollution, and the use of a large amount of alcohol will result in excessively high costs and residual organic solvents in the product, affecting product quality. Chinese patent CN116041238A discloses a method for extracting astaxanthin oil and astaxanthin from bacterial cells, wherein wet bacterial residue is treated with lytic enzyme to break the wall and then dried into dry bacterial residue, and solvents one, two and three are used to extract astaxanthin in sequence, wherein solvent one is a mixed solution of alkanes and alcohols, solvent two is a mixed solution of dichloromethane and chloroform, and solvent three is ketones, alcohols or mixtures thereof, and the extraction process uses a large number of different types of organic solvents, which can cause solvent residues, and the toxicity is relatively large, polluting the ecological environment. Chinese patent CN117945969A discloses a subcritical extraction method for astaxanthin in red Phaffia yeast, wherein n-butane is used as a solvent, and astaxanthin is extracted from bacterial cells by a subcritical extraction method after mixing with fermentation liquid, but the fermentation liquid directly mixed with n-butane reduces the content of astaxanthin, the water content is increased, and n-butane has certain toxicity, and long-term contact or inhalation of n-butane may cause damage to human health, especially to the central nervous system and respiratory system. Chinese patent CN116396199A discloses an extraction and purification process for producing astaxanthin by fermentation, but the process is very complicated, cumbersome, and costly, which is not conducive to industrial production. In addition, due to the thick yeast cell wall, some processes use a high-pressure homogenizer to break the yeast cell wall to dissolve astaxanthin, but the high-pressure homogenizer has high energy consumption, is easily blocked, and has high equipment maintenance costs, which is not suitable for large-scale production. Summary of the invention

[0005] In view of the deficiencies in the prior art, the purpose of the present invention is to provide a method for preparing high-content astaxanthin oil. The prepared astaxanthin oil has high content, low water content, high astaxanthin yield, and is suitable for industrial large-scale production.

[0006] In order to solve the above technical problems, the technical solution provided by the present invention is:

[0007] A method for preparing high-content astaxanthin oil comprises the following steps:

[0008] (1) spray drying the fermentation liquid of Yarrowia lipolytica to obtain a fermentation powder;

[0009] (2) Using a roller press to break the cell wall of the fermentation powder to obtain a flake material;

[0010] (3) After the flake material is crushed into small flakes, supercritical extraction is performed to obtain high-content astaxanthin oil.

[0011] Furthermore, in step (1), the fermentation broth of Yarrowia lipolytica is prepared according to patent CN118441007A.

[0012] Furthermore, in step (1), the air inlet temperature of the spray drying is 150-160° C., and the air outlet temperature is 60-70° C. By using spray drying, the obtained fermentation powder has a low water content, which is conducive to subsequent supercritical extraction to obtain high-content, low-impurity astaxanthin oil.

[0013] Furthermore, in step (2), the roller pressure of the roller press is 80-120MPa. The roller press is used to break the cell wall, which is simple to operate, has low energy consumption, and can reduce production costs. However, the high-pressure homogenization breaking method used in the prior art can only process liquids, and is prone to clogging when the liquid is too concentrated. In addition, the obtained material has a high water content. When the astaxanthin oil is subsequently extracted by supercritical extraction, only anhydrous ethanol can be used as an entrainer, which will result in a decrease in the content of the extracted astaxanthin oil and anhydrous ethanol residue.

[0014] Furthermore, in step (2), the specific operation of the roller press for cell wall breaking is: the fermentation powder is rolled back and forth 20-30 times by the roller press to obtain a compact sheet material.

[0015] Furthermore, in step (2), the roller press is purchased from Wuxi Tianxinda Technology Co., Ltd.

[0016] Furthermore, in step (3), the diameter of the small flake material is less than 0.7 cm. The material state selected for supercritical extraction is preferably solid. When the solid material is extracted, astaxanthin oil can be extracted without using anhydrous ethanol as a carrier. When the solid is crushed into powder, although the contact area of ​​the extraction is large, it is easy to cause the supercritical extraction system to be blocked because the gas is in a high-pressure state and will carry the powder out of the reactor, causing the pipeline to be blocked. The present invention crushes the material into small flakes to increase the contact area, ensure the extraction effect, and prevent the extraction system from being blocked.

[0017] Furthermore, in step (3), the extraction pressure of the supercritical extraction is 70-80 MPa, the extraction temperature is 50-65° C., and the carbon dioxide flow rate is 30 L / min. When the extraction pressure is low, the solubility of the supercritical fluid is poor, the astaxanthin extraction is incomplete, and the extraction efficiency is low. When the extraction pressure is high, the solubility of the supercritical fluid is strong, most of the components in the material are dissolved, the astaxanthin content of the high-content astaxanthin oil is low, and energy consumption is increased, and the production cost is high.

[0018] Furthermore, in step (3), the supercritical CO 2 The extraction equipment was purchased from Nantong Ruizhi Supercritical Technology Development Co., Ltd.

[0019] Furthermore, in step (3), the content of astaxanthin in the high-content astaxanthin oil is 10-20%, and the yield of astaxanthin is greater than 95%. The present invention obtains the high-content astaxanthin oil by extracting the fermentation broth of Yarrowia solani, and the high-content astaxanthin oil has a high astaxanthin content, a high yield, and low loss.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] (1) The present invention does not use any organic solvents in the process of preparing high-content astaxanthin oil, thus avoiding the problem of residual and toxic organic solvents, reducing environmental pollution pressure, and having high ecological and environmental protection.

[0022] (2) Compared with the prior art which uses high-pressure homogenization to break the cell wall, the present invention uses a roller press to break the cell wall, which takes less time, is relatively simple in daily operation and maintenance, has low energy consumption, and greatly saves production costs. In addition, the astaxanthin oil obtained after subsequent supercritical extraction has low water content and high purity.

[0023] (3) The present invention has a simple fermentation broth treatment process and a simple operation method. The obtained astaxanthin oil has a high content and a low water content, and the astaxanthin yield is high, which is suitable for industrial large-scale production. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 This is the preparation process of high-content astaxanthin oil. DETAILED DESCRIPTION

[0025] Example 1

[0026] A method for preparing high-content astaxanthin oil comprises the following steps:

[0027] (1) The fermentation liquid of Yarrowia lipolytica was spray-dried with an inlet air temperature of 155°C and an outlet air temperature of 65°C to obtain 3.5 kg of fermentation powder;

[0028] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 93 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0029] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 70 MPa, an extraction temperature of 55°C, and a carbon dioxide flow rate of 30 L / min, to obtain 428.9 g of high-content astaxanthin oil.

[0030] Example 2

[0031] A method for preparing high-content astaxanthin oil comprises the following steps:

[0032] (1) The fermentation liquid of Yarrowia lipolytica was spray-dried with an inlet air temperature of 150°C and an outlet air temperature of 70°C to obtain 3.5 kg of fermentation powder;

[0033] (2) The fermentation powder is subjected to cell wall breaking by a roller press, the roller pressure of the roller press is 80 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0034] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 70 MPa, an extraction temperature of 55°C, and a carbon dioxide flow rate of 30 L / min, to obtain 461.1 g of high-content astaxanthin oil.

[0035] Example 3

[0036] A method for preparing high-content astaxanthin oil comprises the following steps:

[0037] (1) The fermentation liquid of Yarrowia lipolytica was spray-dried with an inlet air temperature of 160°C and an outlet air temperature of 60°C to obtain 3.5 kg of fermentation powder;

[0038] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 117 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0039] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 75 MPa, an extraction temperature of 55°C, and a carbon dioxide flow rate of 30 L / min, to obtain 363.2 g of high-content astaxanthin oil.

[0040] Example 4

[0041] A method for preparing high-content astaxanthin oil comprises the following steps:

[0042] (1) The fermentation broth of Yarrowia lipolytica was spray-dried at an air inlet temperature of 155°C and an air outlet temperature of 65°C to obtain 3.5 kg of fermentation powder;

[0043] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 117 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0044] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 80 MPa, an extraction temperature of 50°C, and a carbon dioxide flow rate of 30 L / min, to obtain 345.5 g of high-content astaxanthin oil.

[0045] Example 5

[0046] A method for preparing high-content astaxanthin oil comprises the following steps:

[0047] (1) The fermentation broth of Yarrowia lipolytica was spray-dried at an air inlet temperature of 155°C and an air outlet temperature of 65°C to obtain 3.5 kg of fermentation powder;

[0048] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 117 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0049] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 75 MPa, an extraction temperature of 60°C, and a carbon dioxide flow rate of 30 L / min, to obtain 354.9 g of high-content astaxanthin oil.

[0050] Example 6

[0051] A method for preparing high-content astaxanthin oil comprises the following steps:

[0052] (1) The fermentation broth of Yarrowia lipolytica was spray-dried at an air inlet temperature of 155°C and an air outlet temperature of 65°C to obtain 3.5 kg of fermentation powder;

[0053] (2) The fermentation powder is subjected to cell wall breaking by a roller press, the roller pressure of the roller press is 120 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0054] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 75 MPa, an extraction temperature of 65°C, and a carbon dioxide flow rate of 30 L / min, to obtain 362.0 g of high-content astaxanthin oil.

[0055] Comparative Example 1

[0056] A method for preparing astaxanthin oil comprises the following steps:

[0057] (1) spray drying the fermentation broth of Yarrowia lipolytica to obtain 3.5 kg of fermentation powder;

[0058] (2) The fermentation powder is subjected to cell wall breaking by a roller press, the roller pressure of the roller press is 75 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0059] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 70 MPa, an extraction temperature of 55°C, and a carbon dioxide flow rate of 30 L / min to obtain 553.2 g of astaxanthin oil.

[0060] Comparative Example 2

[0061] A method for preparing astaxanthin oil comprises the following steps:

[0062] (1) spray drying the fermentation broth of Yarrowia lipolytica to obtain 3.5 kg of fermentation powder;

[0063] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 93 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0064] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 60 MPa, an extraction temperature of 55°C, and a carbon dioxide flow rate of 30 L / min to obtain 585.1 g of astaxanthin oil.

[0065] Comparative Example 3

[0066] A method for preparing astaxanthin oil comprises the following steps:

[0067] (1) spray drying the fermentation broth of Yarrowia lipolytica to obtain 3.5 kg of fermentation powder;

[0068] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 93 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0069] (3) After the flake material was crushed into small flakes, 3 kg of the small flakes were taken for supercritical extraction at an extraction pressure of 70 MPa, an extraction temperature of 45°C, and a carbon dioxide flow rate of 30 L / min to obtain 567.3 g of astaxanthin oil.

[0070] Comparative Example 4

[0071] A method for preparing astaxanthin oil comprises the following steps:

[0072] (1) subjecting the fermentation broth of Yarrowia lipolytica to high-pressure homogenization, cell wall breaking, centrifugation, and filtration to obtain wet bacterial residue;

[0073] (2) The wet bacterial residue was subjected to supercritical extraction using anhydrous ethanol as the entrainer. The extraction pressure was 70 MPa, the extraction temperature was 55°C, and the carbon dioxide flow rate was 30 L / min, yielding 623.2 g of high-content astaxanthin oil.

[0074] Comparative Example 5

[0075] A method for preparing astaxanthin oil comprises the following steps:

[0076] (1) centrifuging and filtering the fermentation broth of Yarrowia lipolytica to obtain wet bacterial residue;

[0077] (2) The wet bacterial residue is subjected to cell wall breaking by a roller press, and the roller pressure of the roller press is 93 MPa. The fermentation powder is rolled back and forth 25 times by the roller press to obtain the cell wall-broken material;

[0078] (3) 3 kg of the material after cell wall breaking was subjected to supercritical extraction, using anhydrous ethanol as the entrainer, the extraction pressure was 70 MPa, the extraction temperature was 55 °C, and the carbon dioxide flow rate was 30 L / min, to obtain 607.9 g of high-content astaxanthin oil.

[0079] Comparative Example 6

[0080] A method for preparing high-content astaxanthin oil comprises the following steps:

[0081] (1) The fermentation liquid of Yarrowia lipolytica was spray-dried with an inlet air temperature of 155°C and an outlet air temperature of 65°C to obtain 3.5 kg of fermentation powder;

[0082] (2) The fermentation powder is subjected to cell wall disruption by a roller press, the roller pressure of the roller press is 93 MPa, and the fermentation powder is rolled back and forth 25 times by the roller press to obtain a flake material;

[0083] (3) After the flake material is crushed into powder material, 3 kg of the powder material is taken for supercritical extraction at an extraction pressure of 70 MPa, an extraction temperature of 55°C, and a carbon dioxide flow rate of 30 L / min.

[0084] Experimental Example 1

[0085] The astaxanthin content and astaxanthin yield in the high-content astaxanthin oil of Examples 1-6 were tested and calculated, and the calculation results are listed in Table 1.

[0086] Table 1. Comparison of astaxanthin oil

[0087]

[0088] As can be seen from Table 1, the high-content astaxanthin oil prepared by the method of the present invention has an astaxanthin content higher than 11% and an astaxanthin yield higher than 96%, indicating that the astaxanthin content in the astaxanthin oil prepared by the method of the present invention is high and the astaxanthin yield is high.

[0089] Experimental Example 2

[0090] The astaxanthin content and astaxanthin yield of the astaxanthin oils prepared under different experimental parameters in Example 1 and Comparative Examples 1-3 were tested and calculated, and the calculation results are listed in Table 2.

[0091] Table 2. Comparison of astaxanthin oil obtained under different parameters

[0092]

[0093] As can be seen from Table 2, Comparative Example 1 has a lower roller pressure than Example 1, a low astaxanthin content in astaxanthin oil, and a low astaxanthin yield. Comparative Example 2 has a lower extraction pressure than Example 2, a low astaxanthin content in astaxanthin oil, and a low astaxanthin yield. Comparative Example 3 has a lower extraction temperature than Example 1, a low astaxanthin content in astaxanthin oil, and a low astaxanthin yield. It can be seen that Comparative Examples 1-3 are not conducive to the dissolution of astaxanthin under the conditions of inappropriate roller pressure, extraction pressure or extraction temperature of the roller press, but are conducive to the dissolution of other oil components, resulting in an increase in the quality of astaxanthin oil and a decrease in the astaxanthin content in astaxanthin oil. It is shown that the astaxanthin content in the astaxanthin oil prepared under the experimental parameters of the present invention is high, and the astaxanthin yield is high.

[0094] Experimental Example 3

[0095] The astaxanthin content and astaxanthin yield in the astaxanthin oil of Example 1 and Comparative Example 4 were tested and calculated, and the calculation results are listed in Table 3.

[0096] Table 3. Comparison of astaxanthin oil

[0097]

[0098] As can be seen from Table 3, Example 1 uses roller press to break the wall, and the astaxanthin content in the prepared astaxanthin oil is 12.2%, and the astaxanthin yield is 96.5%; Comparative Example 4 uses high-pressure homogenization to break the wall, and the astaxanthin content in the prepared astaxanthin oil is 2.4%, and the astaxanthin yield is 38.4%; Comparative Example 5 obtains wet bacteria residue after centrifugation and filtration of the fermented liquid, and then prepares the astaxanthin oil with wet bacteria residue, and its astaxanthin content is 1.7%, and the astaxanthin yield is 26.5%; Comparative Example 6 crushes the sheet material into powdered material and then performs supercritical extraction, which will block the equipment and cannot obtain astaxanthin oil. It can be seen that the present invention uses spray drying, and then uses roller press to break the wall, and the water content of the astaxanthin oil obtained after subsequent supercritical extraction is low and the purity is high. At the same time, it is found in the experiment that the use of roller press to break the wall is less time-consuming than using a high-pressure homogenizer to break the wall, and the daily operation and maintenance of the roller press are relatively simple, low energy consumption, and greatly save costs.

[0099] The testing method in the specific implementation manner of the present invention is as follows:

[0100] (1) Testing of astaxanthin content

[0101] The test method is: accurately weigh 6 mg of high-content astaxanthin oil into a 25 mL small beaker, add 5 mL of dimethyl sulfoxide solution to dissolve it, transfer it to a 10 mL volumetric flask, and make up the volume. After dilution, the astaxanthin content in the solution is determined by ultraviolet / visible spectrophotometry (UV method, detection wavelength 478nm), and the absorbance value of the measurement is maintained between 0.3-0.8, and the dimethyl sulfoxide solution is used as a blank control.

Claims

1. A method for preparing high-content astaxanthin oil, characterized in that: The following steps are involved: (1) spray drying the fermentation liquid of Yarrowia lipolytica to obtain a fermentation powder; (2) Using a roller press to break the cell wall of the fermentation powder to obtain a flake material; (3) After the flake material is crushed into small flakes, supercritical extraction is performed to obtain high-content astaxanthin oil; In step (2), the roller pressure of the roller press is 80-120 MPa; the specific operation of the roller press for cell wall breaking is: rolling the fermentation powder back and forth 20-30 times with the roller press to obtain a compact sheet material; in step (3), the diameter of the small sheet material is less than 0.7 cm.

2. The method for preparing high-content astaxanthin oil according to claim 1, characterized in that: In step (1), the inlet air temperature of the spray drying is 150-160°C, and the outlet air temperature is 60-70°C.

3. The method for preparing high-content astaxanthin oil according to claim 1, characterized in that: In step (3), the extraction pressure of the supercritical extraction is 70-80 MPa, the extraction temperature is 50-65° C., and the carbon dioxide flow rate is 30 L / min.

4. The method for preparing high-content astaxanthin oil according to claim 1, characterized in that: In step (3), the content of astaxanthin in the high-content astaxanthin oil is 10-20%, and the yield of astaxanthin is >95%.

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