Method for extracting anthocyanin from pitaya peel

Abstract

The invention belongs to the technical field of natural product extraction, and particularly relates to a method for efficiently extracting anthocyanin from pitaya peel by combining enzymolysis, a deep eutectic solvent and ultrasonic waves. The method is technically characterized in that cell walls are destroyed through enzymolysis pretreatment, the anthocyanin release efficiency is improved, a choline chloride-lactic acid eutectic solvent is adopted as an extraction medium, and pitaya peel anthocyanin is obtained through ultrasonic-assisted extraction. The method not only effectively improves the extraction rate of the anthocyanin in the pitaya peel and enhances the comprehensive utilization of the pitaya, but also has remarkable green and environment-friendly advantages and wide industrial application prospects.

Description

Technical Field

[0001] This invention relates to the field of natural product extraction technology. More specifically, this invention relates to a method for extracting anthocyanins from dragon fruit peel. Background Technology

[0002] Dragon fruit peel, a byproduct of dragon fruit processing, is rich in functional pigments such as betaine and anthocyanins, and has high application value. However, due to imperfect extraction processes, the comprehensive utilization rate of dragon fruit peel is currently low, leading to resource waste and potential environmental burden. Anthocyanins, as a natural pigment, possess significant antioxidant, anti-inflammatory, and antibacterial bioactivities and are widely used in the food, pharmaceutical, and cosmetic industries. However, traditional anthocyanin extraction methods mainly rely on organic solvents, which not only consume large quantities and are difficult to recover, but also easily lead to environmental pollution and adversely affect the stability and activity of anthocyanins. Therefore, developing green, efficient, and environmentally friendly extraction technologies is of great significance for realizing the industrial production of anthocyanins from dragon fruit peel. Summary of the Invention

[0003] Another objective of this invention is to provide a method for extracting anthocyanins from dragon fruit peel, which involves enzymatic pretreatment to disrupt cell walls and improve anthocyanin release efficiency, using choline chloride-lactic acid eutectic solvent as the extraction medium, and combining ultrasonic-assisted extraction to obtain anthocyanins from dragon fruit peel.

[0004] To achieve these objectives and other advantages according to the present invention, a method for extracting anthocyanins from dragon fruit peel is provided, comprising the following steps: Step 1: Soak the pretreated dragon fruit peel powder in a cellulase solution. During the soaking process of 0-20 minutes, the soaking temperature is at room temperature. Stir at 200 rpm for 10 seconds every 5 minutes. During the soaking process of 20-60 minutes, first add 0.5% (by volume) of calcium chloride solution to the cellulase solution. The mass fraction of calcium chloride solution is 0.5%. Then, raise the temperature to 40℃ at a rate of 0.8℃ / min and stir at 50 rpm. After soaking, filter to obtain the enzyme extract. The ratio of cellulase solution to dry dragon fruit powder is 10 mL: 1 g. The cellulase solution includes: 1.5% cellulase, 0.5% pomegranate peel extract, 0.2% L-lysine hydrochloride, and the remainder is water. Step 2: Mix choline chloride and lactic acid in a 1:1 molar ratio, stir and heat to 50°C to obtain a eutectic solvent. Add the enzyme extract to the eutectic solvent and extract by ultrasonication. Filter, concentrate and dry to obtain anthocyanin powder. The volume ratio of the eutectic solvent to the enzyme extract is 1:10. The ultrasonic extraction conditions are: ultrasonic power 450W, temperature 38-40°C and time 50 minutes.

[0005] Preferably, the preparation method of pomegranate peel extract is as follows: dried pomegranate peel is pulverized to 60 mesh, and 70% ethanol solution is added at a material-to-liquid ratio of 1g:10mL and refluxed for 1 hour. The filtrate is concentrated under reduced pressure at 50°C to obtain a concentrate. Water with a volume twice that of the concentrate is added to the concentrate, and the mixture is allowed to stand at 4°C for 24 hours. After centrifugation, the supernatant is retained, concentrated, and dried to obtain pomegranate peel extract.

[0006] Preferably, the dragon fruit peel powder is pretreated before being added to the cellulase solution: First, soak the dragon fruit peel powder in a 0.1 mol / L NaCl solution until it is moistened to a water content of 60%. Place the moistened dragon fruit peel powder in -20℃ freezer for 2 hours, then thaw. After thawing, the mixture was kept under a vacuum of -0.08 MPa for 5 minutes to obtain pretreated dragon fruit peel powder.

[0007] Preferably, the freezing method is as follows: freezing at -20°C for 2 hours and thawing: Before freezing, spray the moistened dragon fruit peel powder with a 0.6% sorbitol aqueous solution. The amount of sorbitol aqueous solution sprayed is 3 mL per 100g of moistened dragon fruit peel powder. Then, cool down to -5℃ at 1℃ / min and keep it at 20 minutes to form initial ice crystal nuclei. Then, cool down to -20℃ at 0.5℃ / min and keep it frozen for 2 hours. Thawing was performed using 40kHz ultrasound with a power of 120W in a 4℃ water bath.

[0008] Preferably, before adding the enzyme extract to the eutectic solvent, the enzyme extract is refrigerated at 4°C for 12 hours. After refrigeration, the temperature is increased to 25°C at 0.5°C / min. During the heating process, nitrogen gas is introduced into the enzyme extract at a flow rate of 0.05 L / (min·L), and the extract is allowed to stand at 25°C for 30 minutes. After adding the eutectic solvent, stir at 200 rpm for 30 seconds, and then perform ultrasonic extraction.

[0009] Preferably, before refrigeration, citric acid is added to adjust the pH of the enzyme extract to 4.8-5.2, and nitrogen gas is introduced for 5 minutes every 3 hours during refrigeration at a flow rate of 0.1 L / (min·L).

[0010] Preferably, the citric acid is used before pH adjustment: Disodium hydrogen phosphate and L-aspartic acid were dissolved in deionized water at a molar ratio of 1:1 to prepare a mixed solution with a mass concentration of 5%. The solution was stirred at 50-55℃ for 60-65 minutes to obtain a composite aqueous solution. Before adjusting the pH with citric acid, place the enzyme extract at 25-28℃ and add the compound aqueous solution, wherein the final volume concentration of the compound aqueous solution in the enzyme extract is 0.03%.

[0011] Preferably, during the soaking process of 0-20 min, cationic guar gum nanoparticles with a particle size of 5-10 μm are sprayed into the cellulase solution after stirring every 5 min, with the addition amount being 0.02% of the volume of the cellulase solution. During the heating process of 20-60 min of soaking, the dissolved oxygen concentration in the solution is monitored in real time using a dissolved oxygen sensor. When the dissolved oxygen concentration exceeds 5 mg / L, nitrogen gas is introduced to replace it, maintaining the dissolved oxygen concentration between 2-4 mg / L.

[0012] The present invention has at least the following beneficial effects: This invention achieves high yield through multi-step optimization. In the pretreatment stage, NaCl soaking, sorbitol-assisted freezing-ultrasonic thawing, and vacuum treatment effectively break down dragon fruit peel cells. During enzymatic hydrolysis, pomegranate peel extract and L-lysine hydrochloride are added to the cellulase solution, combined with staged stirring, temperature increase, and dissolved oxygen control to promote anthocyanin release and reduce oxidation. After the enzyme extract is adjusted with a composite aqueous solution, pH controlled, and stabilized by refrigeration and nitrogen purging, it is then extracted with a eutectic solvent using ultrasound. Each step synergistically improves extraction efficiency and stability.

[0013] Other advantages, objectives and features of the present invention will become apparent in part from the following description, and in part from those skilled in the art through study and practice of the invention. Attached Figure Description

[0014] Figure 1 This is a standard curve of anthocyanins in one of the technical solutions of the present invention. Detailed Implementation

[0015] The present invention will be further described in detail below with reference to embodiments, so that those skilled in the art can implement it based on the description.

[0016] <Example 1> A method for extracting anthocyanins from dragon fruit peel includes the following steps: Step 1: Add the pretreated dragon fruit peel powder (the preparation method of dragon fruit peel powder is: cut dragon fruit peel into pieces, dry at 65℃, take it out, grind it into powder and sieve it to obtain dragon fruit peel powder) to the cellulase solution for soaking. During the soaking process of 0-20min, the soaking temperature is room temperature, and the mixture is stirred at 200rpm for 10s every 5min. When soaking for 20-60min, first add 0.5% of the total volume of cellulase calcium chloride solution to the cellulase solution. The mass fraction of calcium chloride solution is 0.5%. Then, raise the temperature to 40℃ at a rate of 0.8℃ / min and stir at 50rpm. After soaking, filter to obtain the enzyme extract. The ratio of cellulase solution to dry dragon fruit powder is 10mL:1g. The cellulase solution includes: 1.5% cellulase, 0.5% pomegranate peel extract, 0.2% L-lysine hydrochloride, and the balance is water. Step 2: Mix choline chloride and lactic acid in a 1:1 molar ratio, stir and heat to 50°C to obtain a eutectic solvent. Add the enzyme extract to the eutectic solvent for ultrasonic extraction, filter, concentrate (rotary evaporation concentration is carried out at a low temperature of 40-50°C), and dry to obtain anthocyanin powder. The ratio of eutectic solvent to enzyme extract is 1:10 (mL / mL). The ultrasonic extraction conditions are: ultrasonic power 450W, temperature 40°C, and time 50 minutes.

[0017] Preparation method of pomegranate peel extract: Pulverize dried pomegranate peel to 60 mesh, add 70% ethanol solution at a material-to-liquid ratio of 1:10 g / mL and reflux for 1 hour. Concentrate the filtrate under reduced pressure at 50℃ to obtain a concentrated solution. Add water twice the volume of the concentrated solution to the concentrated solution, let it stand at 4℃ for 24 hours, centrifuge, retain the supernatant, concentrate and dry to obtain pomegranate peel extract.

[0018] Pre-treat the dragon fruit peel powder before adding it to the cellulase solution: First, soak the dragon fruit peel powder in a 0.1 mol / L NaCl solution until it is moistened to a water content of 60%. Place the moistened dragon fruit peel powder in -20℃ freezer for 2 hours, then thaw. After thawing, the mixture was kept under a vacuum of -0.08 MPa for 5 minutes to obtain pretreated dragon fruit peel powder.

[0019] Freeze at -20℃ for 2 hours and thaw as follows: Before freezing, spray the moistened dragon fruit peel powder with a 0.6% sorbitol aqueous solution. The amount of sorbitol aqueous solution sprayed is 3 mL per 100g of moistened dragon fruit peel powder. Then, cool down to -5℃ at 1℃ / min and keep it at 20 minutes to form initial ice crystal nuclei. Then, cool down to -20℃ at 0.5℃ / min and keep it frozen for 2 hours. Thawing was performed using 40kHz ultrasound with a power of 120W in a 4℃ water bath.

[0020] Before adding the enzyme extract to the eutectic solvent, the enzyme extract was refrigerated at 4°C for 12 hours. After refrigeration, the temperature was increased to 25°C at 0.5°C / min. During the heating process, nitrogen gas was introduced into the enzyme extract at a flow rate of 0.05 L / (min·L), and the mixture was allowed to stand at 25°C for 30 minutes. After adding the eutectic solvent, stir at 200 rpm for 30 seconds, and then perform ultrasonic extraction.

[0021] Before refrigeration, add citric acid to adjust the pH of the enzyme extract to 4.8-5.2. During refrigeration, purge with nitrogen for 5 minutes every 3 hours at a flow rate of 0.1 L / (min·L).

[0022] Before adjusting pH with citric acid: Disodium hydrogen phosphate and L-aspartic acid were dissolved in deionized water at a molar ratio of 1:1 to prepare a mixed solution with a mass concentration of 5%. The solution was stirred at 50-55℃ for 60-65 minutes to obtain a composite aqueous solution. Before adjusting the pH with citric acid, place the enzyme extract at 25-28℃ and add the compound aqueous solution, wherein the final volume concentration of the compound aqueous solution in the enzyme extract is 0.03%.

[0023] During the soaking process from 0 to 20 minutes, cationic guar gum nanoparticles with a particle size of 5 to 10 μm were sprayed into the cellulase solution after stirring every 5 minutes. The amount added was 0.02% of the volume of the cellulase solution. During the heating process from 20 to 60 minutes of soaking, the dissolved oxygen concentration in the solution was monitored in real time using a dissolved oxygen sensor. When the dissolved oxygen concentration exceeded 5 mg / L, nitrogen gas was introduced to replace it and maintain the dissolved oxygen concentration between 2 and 4 mg / L.

[0024] <Example 2> Anthocyanins were extracted using the method described in Example 1, with the following difference: dried pomegranate peel was pulverized to 60 mesh, added to 70% ethanol solution at a material-to-liquid ratio of 1:10 g / mL, and refluxed for 1 hour. The filtrate was concentrated under reduced pressure at 50°C to obtain a concentrated solution, which is the pomegranate peel extract. This example simplifies the preparation process of the pomegranate peel extract by omitting the step of adding water to the concentrated solution and centrifuging, and directly using the concentrated solution after ethanol reflux as the extract. This may result in more residual impurities in the extract, weakening its auxiliary stabilizing effect on anthocyanins, making anthocyanins more susceptible to environmental influences during the extraction process.

[0025] <Example 3> Anthocyanins were extracted using the method described in Example 1, except that no pretreatment or thawing was performed on the dragon fruit peel powder before adding it to the cellulase solution. Specifically, the following steps were not performed in this example: the dragon fruit peel powder was first soaked in a 0.1 mol / L NaCl solution until the water content reached 60%; the moistened dragon fruit peel powder was then frozen at -20°C for 2 hours and thawed; after thawing, it was placed under a vacuum of -0.08 MPa for 5 minutes to obtain pretreated dragon fruit peel powder.

[0026] <Example 4> Anthocyanins were extracted using the method described in Example 1, except that the moistened dragon fruit peel powder was frozen at -20°C for 2 hours and then thawed by placing it directly at room temperature.

[0027] <Example 5> Anthocyanins were extracted using the method described in Example 1, with the following difference: no pretreatment was performed before adding the enzyme extract to the eutectic solvent. Specifically, the following steps were not performed in this example: the enzyme extract was first refrigerated at 4°C for 12 hours, and after refrigeration, the temperature was increased to 25°C at a rate of 0.5°C / min. During the heating process, nitrogen gas was introduced at a flow rate of 0.05 L / min·L, and the mixture was allowed to stand at 25°C for 30 minutes. After adding the eutectic solvent, the mixture was stirred at 200 rpm for 30 seconds, followed by ultrasonic extraction. Before refrigeration, citric acid was added to adjust the pH of the enzyme extract to 4.8–5.2. During refrigeration, nitrogen gas was introduced for 5 minutes every 3 hours at a flow rate of 0.1 L / (min·L), and the pretreatment before citric acid pH adjustment was also performed.

[0028] <Example 6> Anthocyanins were extracted using the method described in Example 1, except that the following steps were not performed before refrigeration: citric acid was added to adjust the pH of the enzyme extract to 4.8-5.2; nitrogen gas was introduced for 5 minutes every 3 hours during refrigeration at a flow rate of 0.1 L / (min·L); and the pH adjustment was not performed before citric acid treatment.

[0029] <Example 7> Anthocyanins were extracted using the method described in Example 1, except that the following steps were not performed before adjusting the pH with citric acid: disodium hydrogen phosphate and L-aspartic acid were dissolved in deionized water at a 1:1 molar ratio to prepare a 5% (w / w) mixed solution, which was then stirred at 50-55°C for 60-65 minutes to obtain a composite aqueous solution; before adjusting the pH with citric acid, the enzyme extract was placed at 25-28°C, and the composite aqueous solution was added, wherein the final volume concentration of the composite aqueous solution in the enzyme extract was 0.03%.

[0030] <Example 8> Anthocyanins were extracted using the method described in Example 1, except that the following steps were not performed in this example: During the soaking process from 0 to 20 minutes, cationic guar gum nanoparticles with a particle size of 5-10 μm were sprayed into the cellulase solution after stirring every 5 minutes, with the addition amount being 0.02% of the volume of the cellulase solution; During the heating process from 20 to 60 minutes of soaking, the dissolved oxygen concentration in the solution was monitored in real time using a dissolved oxygen sensor; When the dissolved oxygen concentration exceeded 5 mg / L, nitrogen gas was introduced to replace it, maintaining the dissolved oxygen concentration between 2 and 4 mg / L.

[0031] <Example 9> Dragon fruit peel is made into dry powder (the preparation method of dragon fruit peel powder is: cut dragon fruit peel into pieces, dry at 65℃, take it out, grind it into powder and sieve it to obtain dragon fruit peel powder). Prepare a 1.5% cellulase solution. Add the dried dragon fruit powder to the enzyme solution at a material-to-liquid ratio of 1:10 (g / mL). Soak at 40°C for 1 hour. Filter the enzyme hydrolysate using a vacuum filtration device to obtain the enzyme extract. Choline chloride and lactic acid are mixed in a 1:1 molar ratio, stirred and heated to 50°C to form a uniform and transparent eutectic solvent. The enzyme extract was added to a eutectic solvent at a material-to-liquid ratio of 1:10 (g / mL) and extracted using an ultrasonic device at 40°C, with an ultrasonic power of 450W and an extraction time of 50 minutes. The extract was then filtered through a vacuum filtration device to obtain the extract. The extract was concentrated by rotary evaporation (rotary evaporation concentration was carried out at a low temperature of 40-50℃) and dried to obtain anthocyanin powder.

[0032] <Experimental Verification> 1. Method for determining anthocyanin content A stock solution of 1000 mg / mL cyanidin-3-glucose standard was precisely prepared. The stock solution was then diluted to obtain standard solutions with concentrations of 200 mg / mL, 400 mg / mL, 600 mg / mL, and 800 mg / mL. 200 μL of each of these standard solutions was added to a 96-well plate, and the absorbance was measured at 530 nm.

[0033] Plot anthocyanin standard curve, with the mass concentration of the standard on the x-axis and the absorbance on the y-axis, such as... Figure 1 As shown.

[0034] The anthocyanin extract from dragon fruit peel was concentrated by rotary evaporation, and its absorbance was measured using the same method. Based on the aforementioned standard curve, the mass concentration of anthocyanins in the sample (mg / mL) was calculated. The anthocyanin yield (mg / g dry sample) was calculated using the following formula: In the formula, Y: anthocyanin yield, mg / g; C: the mass concentration of anthocyanins in the extract, mg / mL; N: the dilution factor of the extract; V: the total volume of the extract, mL; M: the mass of the sample powder, g.

[0035] 2. Single-factor experiment Based on Example 9, single-factor experiments were conducted to investigate the effects of cellulase concentration, ultrasonic conditions, eutectic solvent ratio, and extraction time on anthocyanin extraction yield, as detailed below: With a fixed ultrasonic power of 550W, an ultrasonic time of 40 min, a choline chloride:lactic acid molar ratio of 1:1, and an extraction temperature of 50℃, cellulase concentrations were set at 0.3%, 0.5%, 1%, 1.5%, 2%, and 2.5%. The results showed that anthocyanin yield increased significantly with increasing enzyme concentration, reaching a maximum at 1.5%, after which the yield slightly decreased. This indicates that an appropriate enzyme concentration helps to disrupt cell walls and promote anthocyanin release, but excessively high enzyme concentrations may lead to interference between enzyme molecules or increased system viscosity, thereby inhibiting effective enzymatic hydrolysis and reducing extraction efficiency.

[0036] With a fixed cellulase concentration of 1%, an ultrasonic treatment time of 40 min, a choline chloride:lactic acid molar ratio of 1:1, and an extraction temperature of 50℃, the ultrasonic powers were 150 W, 250 W, 350 W, 450 W, 550 W, and 650 W. The results showed that the anthocyanin yield initially increased and then decreased with increasing ultrasonic power. The yield peaked at 450 W and then slightly declined. This indicates that appropriate ultrasonic intensity can accelerate solute diffusion and improve cell wall disruption efficiency, but excessively high power may cause local temperature rise or ultrasonic cavitation, damaging the anthocyanin structure and leading to its degradation, thus affecting the yield.

[0037] The cellulase concentration was fixed at 1%, the ultrasonic power was 550W, the choline chloride:lactic acid molar ratio was 1:1, the extraction temperature was 50℃, and the ultrasonic time was 10 min, 20 min, 30 min, 40 min, 50 min, and 60 min, respectively.

[0038] As the sonication time increased, the anthocyanin yield gradually increased, reaching its peak at 50 minutes. This indicates that a longer sonication time helps to effectively break down cell structures and promote anthocyanin release. However, excessively long treatment times may lead to increased energy consumption, heat accumulation, or pigment degradation, so they should not be extended indefinitely.

[0039] The cellulase concentration was fixed at 1%, the ultrasonic power was 550W, the ultrasonic time was 40min, the extraction temperature was 50℃, and the molar ratio of choline chloride to lactate was 10:1, 5:1, 1:1, 1:5, 1:10, and 1:15, respectively.

[0040] The results showed that different molar ratios of eutectic solvents significantly affected the extraction efficiency of anthocyanins. With changes in the molar ratio of choline chloride to lactic acid, the anthocyanin yield initially increased and then decreased, with the highest yield observed at a 1:1 molar ratio. This is likely because the eutectic solvent system formed at this ratio possesses an optimal hydrogen bond network structure and polar environment, enabling more effective dissolution of anthocyanin molecules. When the molar ratio deviated from 1:1, the ratio of hydrogen bond donors to acceptors in the system became unbalanced, reducing the affinity and stability for the target substance, leading to decreased solubility and consequently affecting the extraction efficiency. Therefore, a 1:1 molar ratio of choline chloride to lactic acid in the eutectic solvent was the optimal condition for this experiment.

[0041] The cellulase concentration was fixed at 1%, the ultrasonic power was 550W, the ultrasonic time was 40min, the choline chloride:lactic acid molar ratio was 1:1, and the extraction temperatures were 10℃, 20℃, 30℃, 40℃, 50℃, and 60℃.

[0042] Increased temperature enhances the diffusion rate and solubility of anthocyanins, leading to a higher yield, which reaches its maximum at 40°C. Further increases in temperature result in a decrease in yield, possibly due to structural instability and accelerated thermal degradation of anthocyanins at high temperatures, thus affecting the final yield. Therefore, extraction should be controlled under suitable temperature conditions to ensure the stability of the active ingredients.

[0043] 3. Anthocyanin yield The yields of anthocyanins obtained in Examples 1 and 3-9 were detected and calculated, as shown in Table 1. Table 1 Anthocyanin yield Yield (mg / g) Example 1 18.60 Example 3 14.70 Example 4 15.81 Example 5 13.55 Example 6 15.23 Example 7 16.10 Example 8 15.43 Example 9 7.36 Analysis of Table 1 shows that: Example 1 achieved a high yield (18.60 mg / g) through multi-step optimization. In the pretreatment stage, NaCl soaking, sorbitol-assisted freezing-ultrasonic thawing, and vacuum treatment effectively disrupted dragon fruit peel cells. During enzymatic hydrolysis, pomegranate peel extract and L-lysine hydrochloride were added to the cellulase solution. Combined with staged stirring, temperature increase, and dissolved oxygen control, anthocyanin release was promoted and oxidation was reduced. After the enzyme extract was adjusted with a composite aqueous solution, pH controlled, and stabilized by refrigeration and nitrogen purging, it was then extracted with a eutectic solvent using ultrasound. Each step synergistically improved extraction efficiency and stability. In Example 3, no pretreatment was performed on the dragon fruit peel powder (no NaCl soaking, freezing and thawing, or vacuum treatment). The cell structure remained intact, making it difficult for cellulase to penetrate into the cell interior and exert its function. Anthocyanin release was also hindered. Therefore, the yield (14.70 mg / g) was significantly lower than that in Example 1, demonstrating the key role of pretreatment in cell disruption and promoting component dissolution. In Example 4, the thawing method was changed to room temperature natural thawing, replacing the ultrasound-assisted 4°C water bath thawing in Example 1. During natural thawing, the melting rate of ice crystals inside and outside the cells was uneven, resulting in weaker cell disruption and a reduced amount of anthocyanins dissolved from the cells, leading to a lower yield than in Example 1. This indicates that the optimized thawing method can enhance the cell disruption effect.

[0044] Example 5 omitted all pretreatments before adding the enzyme extract to the eutectic solvent (no addition of composite aqueous solution, pH adjustment, refrigeration, and nitrogen purging). Anthocyanins in the enzyme extract were prone to degradation due to the lack of pH buffer and antioxidant protection. At the same time, the unstable solution state affected the efficiency of subsequent ultrasonic extraction, resulting in a low yield (13.55 mg / g), highlighting the importance of enzyme extract pretreatment for stability.

[0045] In Example 6, the pH of the enzyme extract was not adjusted before refrigeration, and nitrogen gas was not introduced during refrigeration, nor was a compound aqueous solution added before pH adjustment. This resulted in unstable pH of the enzyme extract and a lack of nitrogen gas to isolate oxygen. Anthocyanins were easily degraded by acid-base environment and oxidation during refrigeration, and the yield (15.23 mg / g) was lower than that in Example 1.

[0046] In Example 7, the combined aqueous solution of disodium hydrogen phosphate and L-aspartic acid was not added before pH adjustment. This resulted in the loss of the buffering and synergistic stabilizing effect of the solution on the enzyme extract environment. The stability of anthocyanins in the enzyme extract decreased, and some components were degraded in subsequent processing. The yield (16.10 mg / g) was lower than that in Example 1, indicating that the combined aqueous solution can help maintain the stability of the system.

[0047] In Example 8, no cationic guar gum nanoparticles were added during the soaking process, and the dissolved oxygen concentration was not monitored or adjusted. The adsorption and solubilizing effects of the cationic guar gum nanoparticles were absent, and the potentially excessively high dissolved oxygen concentration accelerated anthocyanin oxidation, resulting in a lower yield than in Example 1. This demonstrates the positive impact of nanoparticle assistance and dissolved oxygen control on extraction.

[0048] Example 9 employed the most basic extraction process, without any pretreatment, the enzymatic hydrolysate containing only cellulase (without auxiliary components), and without enzymatic extract stabilization treatment or dissolved oxygen control. Insufficient cell disruption, low enzymatic hydrolysis efficiency, and easy degradation of anthocyanins resulted in the lowest yield due to the lack of optimization at each stage. A comparison with Example 1 demonstrates that multi-step optimization is crucial for improving anthocyanin yield.

[0049] Although embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for the present invention. For those skilled in the art, other modifications can be easily made. Therefore, without departing from the general concept defined by the claims and their equivalents, the present invention is not limited to the specific details and embodiments shown and described herein.

Claims

1. A method for extracting anthocyanins from pitaya peel, characterized in that, The method comprises the following steps: Step 1: The pretreated pitaya peel powder is soaked in a cellulase solution, and during the soaking process for 0-20 min, the temperature is room temperature, and every 5 min, 10 s of stirring at 200 rpm is performed; during the soaking process for 20-60 min, 0.5% of calcium chloride solution in the total volume of the cellulase solution is added first, the mass fraction of the calcium chloride solution is 0.5%, then the temperature is increased to 40 DEG C at a rate of 0.8 DEG C / min, and 50 rpm stirring is performed, after the soaking is completed, filtration is performed, and an enzyme extraction solution is obtained, wherein the dosage ratio of the cellulase solution to the dry pitaya powder is 10 mL:1 g, the cellulase solution comprises 1.5% of cellulase, 0.5% of pomegranate peel extract, and 0.2% of L-lysine hydrochloride, and the rest is water; Step 2: Choline chloride and lactic acid are mixed at a molar ratio of 1:1, stirring is performed, and heating is performed to 50 DEG C to obtain a deep eutectic solvent, the enzyme extraction solution is ultrasonically extracted in the deep eutectic solvent, filtration, concentration and drying are performed, and a cyanidin powder is obtained, wherein the volume dosage ratio of the deep eutectic solvent to the enzyme extraction solution is 1:10, and the ultrasonic extraction conditions are as follows: an ultrasonic power of 450 W, a temperature of 38-40 DEG C, and a time of 50 min.

2. The method of claim 1, wherein the method is characterized by, The preparation method of the pomegranate peel extract is as follows: dry pomegranate peel is crushed to 60 meshes, 70% ethanol solution is added at a material-liquid ratio of 1 g:10 mL for refluxing for 1 h, the filtrate is concentrated under reduced pressure at 50 DEG C to obtain a concentrated solution, water with a volume of 2 times that of the concentrated solution is added to the concentrated solution, the solution is statically placed at 4 DEG C for 24 h, centrifugation is performed, the supernatant is concentrated and dried to obtain the pomegranate peel extract.

3. The method of claim 1, wherein the method is characterized by, The pitaya peel powder is pretreated before being added into the cellulase solution: The pitaya peel powder is first soaked in a 0.1 mol / L NaCl solution until the moisture content is 60%; The soaked pitaya peel powder is frozen at-20 DEG C for 2 h, and then thawed; After thawing, the pitaya peel powder is placed in a vacuum environment with a vacuum degree of-0.08 MPa for 5 min to obtain the pretreated pitaya peel powder.

4. The method of claim 3, wherein the method is characterized by, The method for freezing at-20 DEG C for 2 h and thawing is as follows: Before freezing, 3 mL of a 0.6% sorbitol aqueous solution is sprayed on every 100 g of the soaked pitaya peel powder, the temperature is then decreased to-5 DEG C at a rate of 1 DEG C / min for 20 min to form initial ice crystal nuclei, and then the temperature is decreased to-20 DEG C at a rate of 0.5 DEG C / min for 2 h of freezing; During thawing, 120 W of ultrasonic power is used for assistance in a 4 DEG C water bath.

5. The method of claim 1, wherein the method is characterized by, Before the enzyme extraction solution is added into the deep eutectic solvent, the enzyme extraction solution is placed in a 4 DEG C refrigerator for 12 h, after the refrigeration is completed, the temperature is increased to 25 DEG C at a rate of 0.5 DEG C / min, during the temperature increasing process, nitrogen gas is introduced into the enzyme extraction solution at a flow rate of 0.05 L / (min·L), and the enzyme extraction solution is statically placed at 25 DEG C for 30 min; After the deep eutectic solvent is added, 30 s of stirring at 200 rpm is performed, and then ultrasonic extraction is performed.

6. The method of claim 5, wherein the extraction of the anthocyanins from the pitaya peel is performed by the steps of: Before refrigeration, add citric acid to adjust the pH of the enzyme extract to 4.8-5.2, and during the refrigeration process, pass nitrogen gas for 5 minutes every 3 hours, with a nitrogen flow rate of 0.1 L / (min·L).

7. The method of claim 6, wherein the method is characterized by, Before adjusting the pH with citric acid: Dissolve disodium hydrogen phosphate and L-aspartic acid in deionized water at a molar ratio of 1:1 to prepare a mixed solution with a mass concentration of 5%, and stir at 50-55°C for 60-65 minutes to obtain a composite aqueous solution; Before adjusting the pH with citric acid, first place the enzyme extract at 25-28°C, and add the composite aqueous solution, wherein the final volume concentration of the composite aqueous solution in the enzyme extract is 0.03%.

8. The method of claim 1, wherein the method is characterized by, During the soaking process for 0-20 min, after stirring every 5 min, spray cationic guar gum nanoparticles with a particle size of 5-10 μm into the cellulase solution, with an addition amount of 0.02% of the volume of the cellulase solution, and during the warming process for 20-60 min, use a dissolved oxygen sensor to monitor the dissolved oxygen concentration in the solution in real time, and when the dissolved oxygen concentration exceeds 5 mg / L, pass nitrogen gas for replacement, maintaining the dissolved oxygen concentration at 2-4 mg / L.

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