Phaffia rhodozyma astaxanthin high-efficiency extraction process

Abstract

This invention discloses a high-efficiency extraction process for astaxanthin from Pharfoays redis, belonging to the field of natural active ingredient extraction technology. The process comprises two core steps: acid-heat-high pressure homogenization and ethanol extraction. Pharfoays redis cells are dispersed in a 3.0–4.0 mol / L lactic acid solution, homogenized at 70–80 MPa for 40–50 min after high-speed shearing, and then extracted with anhydrous ethanol at a liquid-to-solid ratio of 15–25 mL / g at 70–80℃ for 30–45 min. Under optimal parameters, the astaxanthin extraction yield reaches 12.14 mg / g, with an astaxanthin purity ≥5.5 mg / g. At a concentration of 105 mg / L, the scavenging rate of DPPH and superoxide anion free radicals is ≥83%. This process achieves synergistic effects of chemical hydrolysis and physical disruption, offering advantages such as high extraction efficiency, good product activity, process stability, and no toxic reagents added. It is suitable for large-scale production, and the product can be applied in the food, health product, and aquaculture industries.

Description

Technical Field

[0001] This invention relates to the field of natural active ingredient extraction technology, specifically to a highly efficient extraction process for astaxanthin from Pharfogel's red yeast, which is particularly suitable for the efficient preparation of astaxanthin in large-scale production. The product can be applied in the fields of food, health products, and aquaculture. Background Technology

[0002] Astaxanthin is a potent natural antioxidant with various biological activities, including lowering blood lipids, anti-inflammation, anti-oxidation, and enhancing immunity. It has broad application prospects in food, health products, and animal feed. Red Pharf yeast is an important microbial source of astaxanthin, with abundant intracellular astaxanthin content. However, the yeast cell wall, composed of β-glucan and other components, has a dense structure that severely hinders the release of astaxanthin, leading to problems such as low extraction efficiency, low product purity, and easy loss of activity in traditional extraction processes.

[0003] Currently, the cell wall disruption methods for astaxanthin from *Phaeff's red yeast* mainly include single processes such as acid-heating, high-pressure homogenization, and ultrasonic disruption. While acid-heating can hydrolyze cell wall polysaccharides, the strong acid and high temperature easily lead to astaxanthin degradation. High-pressure homogenization has high disruption efficiency but consumes a lot of energy and easily causes cell fragments to aggregate and adsorb astaxanthin. Ultrasonic disruption is suitable for small-scale laboratory extraction, but suffers from low efficiency and high cost in large-scale production. Furthermore, existing extraction processes lack systematic optimization of ethanol extraction parameters, resulting in problems such as large solvent consumption, long extraction times, and low recovery rates of target components. In addition, some processes use extraction solvents with high toxicity, limiting the application range of the product.

[0004] Therefore, developing a process for extracting astaxanthin from *Phaves rubra* that balances crushing efficiency and product stability, is simple, energy-efficient, and suitable for large-scale production, has significant practical importance and application value. This invention establishes a highly efficient integrated extraction process by integrating the synergistic advantages of acid heat treatment and high-pressure homogenization, combined with systematic optimization of ethanol extraction parameters, effectively addressing the shortcomings of existing technologies. Summary of the Invention

[0005] To address the technical problems in existing Rhodopsin astaxanthin extraction processes, such as low cell wall disruption efficiency, significant astaxanthin loss, and unreasonable extraction parameters, this invention provides a high-efficiency extraction process for Rhodopsin astaxanthin. This process combines acid-heat-high pressure homogenization with optimized ethanol extraction to achieve efficient release and enrichment of astaxanthin, offering advantages such as process stability, high extraction efficiency, and good product activity.

[0006] The technical solution provided by this invention is as follows:

[0007] A highly efficient extraction process for astaxanthin from Phaefuer's yeast includes the following steps:

[0008] (1) Cell wall breaking treatment: The dry powder of Pharbitis rubescens cells was dispersed in lactic acid solution by acid-heat-high pressure homogenization mixing method. After high-speed shearing dispersion, it was placed in a high-pressure homogenizer for homogenization treatment to obtain cell wall breaking bacterial solution.

[0009] (2) Ethanol extraction: Centrifuge the cell wall-breaking bacterial solution obtained in step (1) to remove the supernatant, add anhydrous ethanol to the precipitate and shake to extract, centrifuge to collect the supernatant, repeat the extraction until the bacterial dry powder turns white, combine the supernatants to obtain astaxanthin extract.

[0010] Further, the concentration of the lactic acid solution in step (1) is 3.0~4.0 mol / L, preferably 3.65 mol / L; the liquid-to-solid ratio is 15~25 mL / g, preferably 20 mL / g.

[0011] Further, the conditions for high-speed shear dispersion in step (1) are: rotation speed 6800 rpm, time 2 min. Further, the conditions for high-pressure homogenization in step (1) are: pressure 70~80 MPa, preferably 76.3 MPa; time 40~50 min, preferably 45 min; and the system temperature is controlled at 30~40℃ during homogenization.

[0012] Further, the concentration of anhydrous ethanol in step (2) is 80%~100%, preferably 100%; the liquid-to-solid ratio is 15~25 mL / g, preferably 18.1 mL / g.

[0013] Further, the conditions for the oscillation extraction in step (2) are: water bath temperature 70~80℃, preferably 77℃; extraction time 30~45 min, preferably 36.7 min; oscillation speed 600 rpm.

[0014] Furthermore, the centrifugation conditions described in step (2) are: 10,000 rpm and centrifugation time of 15 min.

[0015] The present invention also protects the astaxanthin extract prepared by the above process, characterized in that the astaxanthin purity in the extract is ≥5.5 mg / g, the total carotenoid content is ≥12.0 mg / g, and the antioxidant activity is excellent, with a DPPH free radical scavenging rate of ≥86% and a superoxide anion free radical scavenging rate of ≥83% at a concentration of 105 mg / L.

[0016] Beneficial effects

[0017] The efficient extraction process of astaxanthin from Pharfovia rubescens of this invention has the following significant advantages:

[0018] 1. Significantly Improved Extraction Efficiency: The acid-heat-high pressure homogenization hybrid method achieves a synergistic effect between chemical hydrolysis and physical disruption. Lactic acid pretreatment gently hydrolyzes β-glucan in the cell wall, weakening the cell wall structure and reducing the energy consumption of subsequent high-pressure homogenization. The strong shear force and cavitation effect generated by high-pressure homogenization efficiently disrupt the weakened cell wall, significantly improving astaxanthin release efficiency. Experimental results show that the astaxanthin extraction yield of the hybrid method of this invention reaches 12.14 mg / g, which is about 150% higher than that of the single acid-heat method (4.85 mg / g) and about 53% higher than that of the single high-pressure homogenization method (7.92 mg / g).

[0019] 2. System Optimization of Process Parameters: Key process parameters were systematically optimized using response surface methodology, establishing a reliable quadratic multinomial regression model to ensure process stability and reproducibility. The regression model for the acid-heat-high pressure homogenization mixing method showed R² = 0.9923, with an adjusted R² = 0.9824; the regression model for the ethanol extraction method showed R² = 0.9414, with an adjusted R² = 0.8661. The models exhibited good fit, with small deviations between predicted and actual values.

[0020] 3. Sufficient retention of product activity: The process conditions are mild and controllable. The system temperature is controlled at 30~40℃ during cell wall disruption and at 77℃ during extraction, effectively avoiding damage to the astaxanthin structure caused by high temperatures. No toxic or harmful reagents are added throughout the process, ensuring high product safety. The prepared astaxanthin extract exhibits excellent antioxidant activity, achieving a DPPH free radical scavenging rate of 86.7±0.9%, a superoxide anion free radical scavenging rate of 83.0±1.1%, and a hydroxyl free radical scavenging rate of 82.4±1.0% at a concentration of 105 mg / L.

[0021] 4. Accurate and reliable quantitative analysis: HPLC is used for specific quantification of astaxanthin in the extract, effectively separating astaxanthin from other co-extracted carotenoids (such as β-carotene and lutein), resulting in more accurate and reliable results. The HPLC standard curve equation is y=13521x-29921, showing good linearity, and the pure astaxanthin content is 5.59 mg / g.

[0022] 5. Simple process suitable for scale-up: The process steps are simple and easy to operate. The equipment used are all commonly used in the food industry, making it suitable for large-scale industrial production. Ethanol, as the extraction solvent, has advantages such as strong solubility, low toxicity, and easy recovery, meeting the requirements for the preparation of food-grade products. The prepared astaxanthin extract can be widely used in the fields of food additives, health product raw materials, or functional feed additives for aquaculture. Attached Figure Description

[0023] Figure 1 This is a schematic diagram illustrating the effect of different lactic acid concentrations on astaxanthin extraction yield in Example 1 of the present invention;

[0024] Figure 2 This is a schematic diagram illustrating the effect of different liquid-to-solid ratios on astaxanthin extraction yield in Example 1 of the present invention;

[0025] Figure 3 This is a schematic diagram illustrating the effect of different homogenization pressures on astaxanthin extraction yield in Example 1 of the present invention;

[0026] Figure 4 This is a schematic diagram illustrating the effect of different homogenization times on astaxanthin extraction yield in Example 1 of the present invention;

[0027] Figure 5 This is a diagram showing the optimization results of the hybrid method response surface methodology in Embodiment 1 of the present invention.

[0028] Figure 6 This is a schematic diagram illustrating the effect of different ethanol concentrations on astaxanthin extraction yield in Example 2 of the present invention;

[0029] Figure 7 This is a schematic diagram illustrating the effect of different liquid-to-solid ratios on astaxanthin extraction yield in Example 2 of the present invention;

[0030] Figure 8 This is a schematic diagram illustrating the effect of different water bath temperatures on astaxanthin extraction yield in Example 2 of the present invention;

[0031] Figure 9 This is a schematic diagram illustrating the effect of different water bath times on astaxanthin extraction in Example 2 of the present invention;

[0032] Figure 10 This is a graph showing the response surface optimization results of the ethanol extraction method in Example 2 of the present invention;

[0033] Figure 11 This is the HPLC chromatogram of the astaxanthin standard in Example 3 of the present invention;

[0034] Figure 12 This is the HPLC chromatogram of the astaxanthin extract in Example 3 of the present invention. Detailed Implementation

[0035] Example 1: Optimization of cell wall disruption process and comparative experiment between mixing method and single cell wall disruption method

[0036] 1. Experimental materials: Pharrellis redis cell powder (purchased from Wanfang Dongxun Company), lactic acid, anhydrous ethanol and other reagents were all analytical grade.

[0037] 2. Experimental Methods:

[0038] 2.1 Single-factor experiment: Accurately weigh 1.0 g of Pharrellis redis cell powder (purchased from Wanfang Dongxun Company), and uniformly disperse it in lactic acid solution. Use a high-speed shear disperser at 6800 rpm for 2 min to fully disperse the cells. Place the uniformly dispersed bacterial solution in a microfluidic high-pressure homogenizer for homogenization. After centrifugation to remove the supernatant, add 20 mL of anhydrous ethanol and extract by shaking at 600 rpm in a 50℃ magnetically stirred water bath for 36 min. After centrifugation, take the supernatant and repeat the extraction until the yeast powder turns white. Combine the supernatants and measure the absorbance at a wavelength of 470 nm. Calculate the astaxanthin extraction amount according to formula (1).

[0039]

[0040] Where ODmax is the absorbance value at the maximum absorption wavelength of astaxanthin; V is the volume of organic solvent used for extraction (mL); D is the dilution factor; a is the conversion factor between astaxanthin concentration and ODmax; and W is the mass of Pharfia redis yeast (g).

[0041] The factors considered and the results are as follows:

[0042] (1) Effect of lactic acid concentration: With a fixed liquid-to-solid ratio of 20 mL / g, a homogenization pressure of 80 MPa, and a homogenization time of 40 min, the effects of lactic acid concentrations of 1, 3, 5, 7, and 9 mol / L on astaxanthin extraction were investigated. The results showed that ( Figure 1 Within the lactic acid concentration range of 1–5 mol / L, the astaxanthin extraction yield gradually increased with increasing concentration; the extraction yield reached its peak at a concentration of 5 mol / L; and decreased continuously when the concentration continued to increase to 7–9 mol / L. A suitable lactic acid concentration can gently hydrolyze the polysaccharide components in the cell wall of *Phaves rubrum*, weakening the cell wall's structural integrity; however, excessive acidification can lead to excessive cell wall contraction or denaturation, thus hindering the release of contents.

[0043] (2) Effect of liquid-to-solid ratio: With a fixed lactic acid concentration of 5 mol / L, homogenization pressure of 80 MPa, and homogenization time of 40 min, the effects of liquid-to-solid ratios of 10, 15, 20, 25, and 30 mL / g on astaxanthin extraction were investigated. The results showed that ( Figure 2 When the liquid-to-solid ratio is in the range of 10-20 mL / g, the extraction yield gradually increases with the increase of the liquid-to-solid ratio, reaching a maximum at 20 mL / g; when it is further increased to 25-30 mL / g, the extraction yield decreases significantly. A liquid-to-solid ratio of 20 mL / g provides the optimal solvent environment for astaxanthin dissolution, ensuring sufficient solvent while maintaining a high extraction concentration gradient.

[0044] (3) Effect of homogenization pressure: With a fixed lactic acid concentration of 5 mol / L, a liquid-to-solid ratio of 20 mL / g, and a homogenization time of 40 min, the effects of homogenization pressures of 40, 60, 80, 100, and 120 MPa on astaxanthin extraction were investigated. The results showed that ( Figure 3 Within a pressure range of 40–80 MPa, the extraction yield gradually increases with increasing pressure, reaching a peak at 80 MPa; when the pressure increases to 100–120 MPa, the extraction yield continuously decreases. 80 MPa effectively balances shear force and cell disruption efficiency, reducing cell debris aggregation while breaking down cell walls.

[0045] (4) Effect of homogenization time: With a fixed lactic acid concentration of 5 mol / L, a liquid-to-solid ratio of 20 mL / g, and a homogenization pressure of 80 MPa, the effects of homogenization times of 20, 30, 40, 50, and 60 min on astaxanthin extraction were investigated. The results showed that ( Figure 4 The homogenization time was in the range of 20-50 min, and the extraction yield gradually increased with the extension of time, reaching a peak at 50 min; when extended to 60 min, the extraction yield decreased instead. The 50 min homogenization time ensured that the high-pressure shear force acted fully on the cells, ensuring that the cell walls were completely broken.

[0046] 2.2 Response Surface Optimization Experiment

[0047] Based on the single-factor experiments, a three-factor, three-level response surface optimization experiment was conducted using a Box-Behnken design (BBD). Homogenization pressure (A), homogenization time (B), and lactic acid concentration (C) were used as independent variables, and astaxanthin extraction amount (Y) was used as the response value. The factor levels are shown in Table 1.

[0048] Table 1. Factors and Levels in Hybrid Response Surface Design

[0049]

[0050] A total of 15 experiments were conducted. The experimental protocols and results are shown in Table 2.

[0051] Table 2. Hybrid Response Surface Methodology Experimental Design and Results

[0052]

[0053] Through model optimization, the optimal process conditions for the mixed method were obtained as follows: homogenization pressure 76.3 MPa, homogenization time 45 min, and lactic acid concentration 3.65 mol / L. Under these conditions, the model predicted that the astaxanthin extraction yield was 11.21 mg / g.

[0054] 2.3 Verification Experiment

[0055] Three parallel validation experiments were conducted under the optimized process conditions. The actual average astaxanthin extraction yield was 10.87 mg / g, which was very close to the predicted value, thus verifying the accuracy and repeatability of the model and optimization results.

[0056] 2.4 Comparison with single cell wall disruption method

[0057] Using the same ethanol extraction conditions, the extraction effects of the mixed method (lactic acid concentration 3.65 mol / L, homogenization pressure 76.3 MPa, homogenization time 45 min), acid-heat method (lactic acid concentration 7 mol / L, liquid-to-solid ratio 20 mL / g, ultrasonication at 50℃ for 36 min), and high-pressure homogenization method (homogenization pressure 108 MPa, homogenization time 40 min, ultrasonic temperature 36℃) of this invention were compared. The results showed that the astaxanthin extraction yield of the mixed method (10.87 mg / g) was significantly better than that of the acid-heat method (4.85 mg / g) and the high-pressure homogenization method (7.92 mg / g), representing an increase of approximately 124% compared to the acid-heat method alone and approximately 37% compared to the high-pressure homogenization method alone.

[0058] Example 2: Optimization of Ethanol Extraction Process

[0059] 1. Experimental materials: Pharrellis redis cell powder (purchased from Wanfang Dongxun Company), lactic acid, anhydrous ethanol and other reagents were all analytical grade.

[0060] 2. Experimental methods: The cell wall disruption process optimized in Example 1 was used to treat Pharfovia rubescens cells. After centrifugation to remove the supernatant, a single-factor experiment on ethanol extraction was conducted to investigate the effects of ethanol concentration, liquid-to-solid ratio, water bath temperature, and water bath time on the astaxanthin extraction yield.

[0061] 2.1 Single-factor experiment:

[0062] (1) Effect of ethanol concentration: With a fixed liquid-to-solid ratio of 20 mL / g, a water bath temperature of 50℃, and a water bath time of 36 min, the effects of ethanol concentrations of 20%, 40%, 60%, 80%, and 100% on astaxanthin extraction were investigated. The results showed that ( Figure 6 With ethanol concentrations ranging from 20% to 100%, the extraction yield continuously increased with increasing concentration, showing no significant decreasing trend, reaching its highest value at 100% ethanol. As a fat-soluble substance, astaxanthin's solubility significantly increased with increasing ethanol concentration.

[0063] (2) Effect of liquid-to-solid ratio: With a fixed ethanol concentration of 100%, water bath temperature of 50℃, and water bath time of 36 min, the effects of liquid-to-solid ratios of 10, 15, 20, 25, and 30 mL / g on astaxanthin extraction were investigated. The results showed that ( Figure 7When the liquid-to-solid ratio is in the range of 10-25 mL / g, the extraction yield gradually increases with the increase of the liquid-to-solid ratio, reaching a peak at 25 mL / g; when it continues to increase to 30 mL / g, the extraction yield decreases instead.

[0064] (3) Effect of water bath temperature: With a fixed ethanol concentration of 100%, a liquid-to-solid ratio of 20 mL / g, and a water bath time of 36 min, the effects of water bath temperatures of 30, 40, 50, 60, 70, and 80℃ on the extraction yield of astaxanthin were investigated. The results showed that ( Figure 8 When the water bath temperature is in the range of 30~40℃, the extraction yield is basically the same; when the temperature is in the range of 40~70℃, the extraction yield gradually increases with the increase of temperature.

[0065] (4) Effect of water bath time: With a fixed ethanol concentration of 100%, a liquid-to-solid ratio of 20 mL / g, and a water bath temperature of 70℃, the effects of water bath times of 10, 20, 30, 40, 50, and 60 min on the astaxanthin extraction yield were investigated. The results showed that ( Figure 9 When the water bath time is in the range of 10 to 50 minutes, the extraction amount gradually increases with the extension of time, reaching a peak at 50 minutes; when the time is extended to 60 minutes, the extraction amount decreases instead.

[0066] 2.2 Response Surface Optimization Experiment

[0067] Based on the single-factor experiments, a three-factor, three-level response surface optimization experiment was conducted using a Box-Behnken design. The liquid-to-solid ratio (A), water bath time (B), and water bath temperature (C) were used as independent variables, and the astaxanthin extraction amount (Y) was used as the response value. The factor levels are shown in Table 4.

[0068] Table 4. Response surface design factors and levels for ethanol extraction method

[0069]

[0070] A total of 17 experiments were conducted, and the experimental protocols and results are shown in Table 5.

[0071] Table 5 Response surface methodology and results of ethanol extraction method

[0072]

[0073] Through model optimization, the optimal process conditions for ethanol extraction were obtained as follows: liquid-to-solid ratio of 18.1 mL / g, water bath time of 36.7 min, and water bath temperature of 77℃. Under these conditions, the model predicted an astaxanthin extraction yield of 12.92 mg / g.

[0074] 2.3 Verification Experiment

[0075] Three parallel validation experiments were conducted under the optimized optimal process conditions. The actual average astaxanthin extraction yield was 12.14 ± 0.04 mg / g, which was very close to the predicted value, thus verifying the accuracy and repeatability of the model and optimization results.

[0076] Example 3 Purity determination of astaxanthin extract

[0077] 3.1 Plotting the Astaxanthin Standard Curve

[0078] Accurately weigh 2.5 mg of astaxanthin standard (Sigma-Aldrich, purity ≥97%), dissolve it in chromatographic grade methanol-water solution (9:1 v / v), transfer to a 25 mL amber volumetric flask, and dilute to volume to prepare a stock solution with a concentration of 100 μg / mL. Store in a sealed container protected from light at -20°C. Before use, serially dilute the stock solution with the same solvent to prepare standard working solutions with concentrations of 1, 2, 3, 4, and 5 μg / mL. Filter the solutions through a 0.22 μm organic phase filter membrane before injection analysis.

[0079] High performance liquid chromatography (HPLC) conditions: Elite C18 column (250 mm × 4.6 mm × 5 μm); UV detector; detection wavelength 470 nm; injection volume 10 μL; flow rate 0.7 mL / min; run time 10 min; column temperature room temperature; mobile phase methanol:water = 9:1.

[0080] 3.2 Determination of astaxanthin content in samples

[0081] The astaxanthin extract prepared in Example 2 was concentrated by rotary evaporation, redissolved in methanol-water solution (9:1, v / v), filtered through a 0.22 μm organic phase filter membrane, and then analyzed by HPLC. The astaxanthin content in the sample was calculated by substituting the peak area into the standard curve (4). Three replicates were set for each sample, and the results are expressed as mean ± standard deviation.

[0082] The results showed that the astaxanthin content in the extract was 5.59 mg / g. For comparison, the total carotenoid content of the same extract was determined using UV-Vis spectrophotometry at 470 nm, and was 12.14 mg / g. The HPLC chromatogram showed (…). Figure 11-12 Astaxanthin exhibits symmetrical peak shape and stable retention time, and can be effectively separated from other carotenoids such as β-carotene and lutein, resulting in more specific and accurate measurement results.

Claims

1. A high-efficiency extraction process for astaxanthin from Phaefuer's yeast, characterized in that, The core processes include cell wall disruption and ethanol extraction, and the specific steps are as follows: (1) Cell wall breaking treatment: The acid-heat-high pressure homogenization mixing method was adopted. The dry powder of Pharrellis redis cells was accurately weighed, dispersed in lactic acid solution, and after high-speed shearing treatment, it was placed in a high-pressure homogenizer for homogenization and cell wall breaking to obtain cell wall broken bacterial solution. (2) Ethanol extraction: Centrifuge the cell wall-breaking bacterial solution from step (1) to remove the supernatant, add anhydrous ethanol, and extract by shaking under constant temperature water bath conditions. Centrifuge to collect the supernatant, and repeat the extraction until the bacterial dry powder turns white. Combine the supernatants to obtain astaxanthin extract.

2. The high-efficiency extraction process for astaxanthin from Phaefuer's yeast according to claim 1, characterized in that, The key parameters for cell wall disruption by mixing in step (1) are: lactic acid concentration 3.0~4.0 mol / L, liquid-to-solid ratio 15~25 mL / g, high-speed shearing speed 6800 rpm, shearing time 2 min, homogenization pressure 70~80 MPa, and homogenization time 40~50 min.

3. The high-efficiency extraction process for astaxanthin from Phaefuer's yeast according to claim 1 or 2, characterized in that, The optimal combination of parameters for cell wall disruption by mixing in step (1) is: lactic acid concentration 3.65 mol / L, liquid-to-solid ratio 20 mL / g, homogenization pressure 76.3 MPa, and homogenization time 45 min.

4. The high-efficiency extraction process for astaxanthin from Phaefuer's yeast according to claim 1, characterized in that, The key parameters for ethanol extraction in step (2) are: ethanol concentration 80%~100%, liquid-to-solid ratio 15~25 mL / g, water bath temperature 70~80℃, water bath time 30~45 min, and oscillation speed 600 rmp.

5. The high-efficiency extraction process for astaxanthin from Phaefuer's yeast according to claim 1 or 4, characterized in that, The optimal combination of parameters for ethanol extraction in step (2) is: ethanol concentration 100%, liquid-to-solid ratio 18.1 mL / g, water bath temperature 77℃, and water bath time 36.7 min.

6. The high-efficiency extraction process for astaxanthin from Phaefuer's yeast according to claim 1, characterized in that, After high-speed shearing in step (1), the bacterial solution should be kept in a uniformly dispersed state without obvious bacterial aggregation; during high-pressure homogenization, the system temperature should be controlled at 30~40℃.

7. The high-efficiency extraction process for astaxanthin from Phaefuer's yeast according to claim 1, characterized in that, In step (2), the centrifugation conditions are 10,000 rpm and the centrifugation time is 15 min. During the extraction process, it is necessary to replenish the amount of ethanol lost due to evaporation to maintain a stable liquid-to-solid ratio.

8. An astaxanthin extract prepared by the process described in any one of claims 1 to 7, characterized in that, The extract contains astaxanthin with a purity of ≥5.5 mg / g, total carotenoid content of ≥12.0 mg / g, and exhibits a DPPH free radical scavenging rate of ≥86% and a superoxide anion free radical scavenging rate of ≥83% at a concentration of 105 mg / L.

9. The astaxanthin extract according to claim 8, characterized in that, This extract can be used in the fields of food additives, health product raw materials, or functional feed additives for aquaculture.

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