A method for ultrasonic-assisted propylene carbonate extraction of coarse cereal protein

Abstract

The application provides a method for ultrasonic-assisted propylene carbonate extraction of coarse cereal protein. The application uses a mixed solution of low-toxic, green and degradable propylene carbonate and water as an extraction agent, and adopts ultrasonic-assisted extraction of coarse cereal protein. The propylene carbonate can be recovered by adding sodium chloride to separate phases, and used for extraction of new coarse cereal protein. The water phase is freeze-dried to obtain protein, and the protein extraction rate of the propylene carbonate aqueous solution is higher than that of pure water. The method is successfully used for green and efficient extraction of coarse cereal protein.

Description

Technical Field

[0001] This invention belongs to the technical field of plant protein extraction, and specifically discloses a method for extracting protein from miscellaneous grains using ultrasound-assisted propylene carbonate. Background Technology

[0002] Coarse grains possess various bioactive properties, such as antioxidant, anti-obesity, and anti-diabetic effects, and offer good health benefits for cardiovascular diseases. Sorghum protein, as an important plant protein resource, has a balanced amino acid composition and high content. Due to its unique nutritional characteristics and functional activities, sorghum protein shows promising application prospects in the development of functional foods and plant-based products.

[0003] Among existing protein extraction methods, aqueous extraction is inefficient due to the strong hydrophobicity and weak hydration of proteins. Alkaline extraction has a higher extraction rate, but it relies on high concentrations of acid and alkali, resulting in problems such as high solvent toxicity, difficult separation, high energy consumption, and high costs for waste treatment. Enzymatic hydrolysis offers mild extraction conditions, but enzyme procurement is expensive, and the enzymatic reaction is time-consuming. These shortcomings limit the in-depth development and utilization of coarse grain proteins. Summary of the Invention

[0004] To address the problems of high solvent toxicity, high reagent cost, difficult separation, high energy consumption, and difficulty in solvent recovery and reuse in existing methods for extracting protein from miscellaneous grains, this invention proposes a method for extracting protein from miscellaneous grains using ultrasound-assisted propylene carbonate.

[0005] The above-mentioned method for ultrasound-assisted extraction of protein from grains using propylene carbonate includes the following steps: S1, Preparation of the extractant solution; Prepare an extraction solution by mixing water and propylene carbonate in a preset volume ratio; S2, extraction and recycling of protein from whole grains; Weigh the mixed grain powder, add the extractant solution, perform ultrasonic-assisted extraction, collect the supernatant after centrifugation, add sodium chloride to the supernatant, vortex and separate the two phases. The upper layer is the aqueous phase and the lower layer is the propylene carbonate phase. The mixed grain protein in the extractant solution is recovered to the upper aqueous phase. After freeze-drying, the protein solid powder can be obtained. S3, recycling and reuse of propylene carbonate; The propylene carbonate phase obtained in step S2 is recovered, and new grain protein is extracted again following steps S1 and S2.

[0006] In step S1, the volume ratio of water to propylene carbonate is 1:1 to 9:1.

[0007] In step S2, the grain powder is sorghum powder.

[0008] In step S2, the ratio of grain powder to extractant solution is 1:10 to 1:50 g / mL.

[0009] In step S2, the ultrasonic time is 0~60 min and the ultrasonic temperature is 20~100 °C.

[0010] In step S2, the mass concentration of sodium chloride is 40~200 mg / mL.

[0011] In step S2, the centrifugal force is 9000 r / min, the centrifugation time is 10 min, and the vortexing time is 1 min.

[0012] Preferably, the volume ratio of water to propylene carbonate is 5:1, the ratio of grain powder to extractant solution is 1:20 g / mL, the ultrasonic time is 30 min, the ultrasonic temperature is 60°C, and the mass concentration of sodium chloride is 160 mg / mL.

[0013] Compared with the prior art, the present invention has the following beneficial effects.

[0014] 1. This invention uses inexpensive, low-toxicity, and environmentally friendly propylene carbonate and water to prepare an extraction solution as an extractant. This is the first report of using propylene carbonate as an extractant to achieve green and efficient protein extraction.

[0015] 2. This invention avoids the use of strong acids, strong alkalis, and enzymes, thus solving the problems of environmental pollution, high cost, and high energy consumption in existing extraction methods.

[0016] 3. Compared with pure water, the propylene carbonate aqueous solution prepared by this invention has a higher protein extraction rate.

[0017] 4. This invention separates the propylene carbonate phase from the aqueous phase using sodium chloride-induced phase separation, enabling the recovery of propylene carbonate for reuse in new extractions, and yielding protein from the aqueous phase. The recovery of the extractant and protein eliminates the need for distillation or column chromatography, reducing energy consumption, shortening extraction time, and achieving sustainable extraction.

[0018] 5. Propylene carbonate can be degraded into non-toxic metabolites, which, compared with traditional solvents, are better able to meet the safety requirements of subsequent protein food processing. Attached Figure Description

[0019] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0020] Figure 1 This is a flowchart of a method for ultrasound-assisted extraction of protein from grains using propylene carbonate.

[0021] Figure 2 This describes the effect of the extractant volume ratio on the protein extraction rate.

[0022] Figure 3 This relates to the effect of the material-to-liquid ratio on the protein extraction rate.

[0023] Figure 4 This describes the effect of ultrasound time on protein extraction rate.

[0024] Figure 5 This describes the effect of ultrasonic temperature on protein extraction rate.

[0025] Figure 6 This describes the effect of sodium chloride concentration on the recovery rate of propylene carbonate.

[0026] Figure 7 This relates to the effect of the type of extractant on the protein extraction rate. Detailed Implementation

[0027] Propylene carbonate is a high-boiling-point, low-melting-point, low-viscosity, and low-toxicity polar solvent, possessing both biodegradability and economic viability in production. This solvent can enhance protein extraction efficiency through mechanisms such as disrupting cell structure, promoting protein dissolution, and regulating extract affinity. Furthermore, solvent recovery can be achieved through salt separation at room temperature, eliminating the need for high-temperature distillation and significantly reducing energy consumption. In addition, ultrasound-assisted extraction technology is simple to operate, operates at lower temperatures, and causes minimal damage to heat-sensitive active substances. Ultrasonic cavitation can break down cell walls, enhancing protein mass transfer and dissolution, and improving the extraction efficiency of the extractant. Based on these theories, this invention combines propylene carbonate, ultrasound-assisted extraction, and salt-phase solvent recycling to design and develop a simple, green, and sustainable method for the efficient extraction and separation of proteins from miscellaneous grains.

[0028] To better explain and facilitate understanding of the present invention, a detailed description of the invention is provided below with reference to the accompanying drawings and specific embodiments. Different lowercase letters in the figures indicate significant differences at the 95% confidence level. Unless otherwise specified, the techniques used in the embodiments are conventional methods well-known to those skilled in the art, and the reagents used are commercially available products.

[0029] Example 1 Optimization of the extractant A. Extractant volume ratio (1) Experimental method: Water and propylene carbonate were prepared into extraction solutions (1:1, 3:1, 5:1, 7:1, 9:1) according to a certain volume ratio. 0.1 g of sorghum powder was weighed into a centrifuge tube, 2 mL of extraction solution was added, and the centrifuge tube was placed in an ultrasonic cleaner. Ultrasonic extraction was performed at 60 ℃ for 30 min, followed by centrifugation at 9000 r / min for 10 min. The supernatant was collected into the centrifuge tube. Sodium chloride was added to adjust the sodium chloride concentration to 160 mg / mL. After vortexing for 1 min, the two phases were separated. The upper aqueous phase containing sorghum protein and the lower propylene carbonate phase were collected. Figure 1 As shown in the figure. Subsequently, the protein content in the supernatant was determined using the Braford Protein Assay Kit.

[0030] (2) Experimental results: such as Figure 2 As shown.

[0031] (3) Results analysis: As the volume ratio of the extractant increases, the extraction rate shows a trend of first increasing and then decreasing. When the volume ratio of the extractant is 5:1, the extraction rate reaches its maximum value. P <0.05). When the propylene carbonate content is too low, the interaction with proteins is not significant, and the extraction rate cannot be effectively improved. When the propylene carbonate content is too high, the liquid surface tension and viscosity are high, and the ultrasonic cavitation effect is weakened. Therefore, an extractant volume ratio of 5:1 is selected.

[0032] B. Material-to-liquid ratio (1) Experimental method: The method is the same as that in Example 1A, except that different volumes of extractant solution are used, i.e., material-liquid ratio (1:10, 1:20, 1:30, 1:40, 1:50 g / mL) to extract sorghum protein.

[0033] (2) Experimental results: such as Figure 3 As shown.

[0034] (3) Results Analysis: As the solid-liquid ratio increased, the extraction rate showed a trend of first increasing and then slowly decreasing. The extraction rate reached its maximum value when the solid-liquid ratio was 1:20 g / mL (P<0.05). A low solid-liquid ratio would lead to insufficient liquid volume, affecting protein dissolution. When the solid-liquid ratio was too high, the ultrasonic energy received by the unit volume of extractant decreased, the cavitation effect weakened, protein release was hindered, and the extraction rate also decreased. Therefore, a solid-liquid ratio of 1:20 g / mL was selected.

[0035] Example 2: Optimization of Extraction Conditions A. Ultrasound time (1) Experimental method: The method is the same as that in Example 1A, except that different ultrasonic times (0, 15, 30, 45, 60 min) are used to extract sorghum protein.

[0036] (2) Experimental results: such as Figure 4 As shown.

[0037] (3) Results Analysis: With the increase of ultrasonic time, the extraction rate showed a trend of first increasing and then decreasing, reaching its maximum value when the ultrasonic time was 30 min (P < 0.05). Too short an ultrasonic time resulted in insufficient ultrasonic mechanical effect. Too long an ultrasonic time may damage the protein structure, leading to a decrease in extraction rate, and also increases energy consumption, which is inconsistent with the principles of green and economical methods. Therefore, an ultrasonic time of 30 min was selected.

[0038] B. Ultrasonic temperature (1) Experimental method: The method is the same as that in Example 1A, except that different ultrasonic temperatures (20, 40, 60, 80, 100 ℃) are used to extract sorghum protein.

[0039] (2) Experimental results: such as Figure 5 As shown.

[0040] (3) Results Analysis: As the ultrasonic temperature increased, the extraction rate showed a trend of first increasing and then decreasing, reaching its maximum value at an ultrasonic temperature of 60 ℃ (P<0.05). At lower ultrasonic temperatures, the interaction between solvent molecules and protein molecules was weaker, resulting in a lower protein extraction rate. At excessively high ultrasonic temperatures, the synergistic effect between solvent molecules and protein molecules was disrupted, and volatilization may alter the volume ratio of the solvent system, affecting the polarity of the solvent and the protein's solubility. Therefore, an ultrasonic temperature of 60 ℃ was selected.

[0041] Example 3: Recovery of propylene carbonate (1) Experimental method: The method is the same as that in Example 1A, except that different sodium chloride concentrations (40, 80, 120, 160, 200 mg / mL) are used to recover propylene carbonate.

[0042] (2) Experimental results: such as Figure 6 As shown.

[0043] (3) Results Analysis: With the increase of sodium chloride concentration, the recovery rate of propylene carbonate showed a trend of first increasing and then leveling off. The recovery rate reached its maximum when the sodium chloride concentration was 160 mg / mL (P<0.05). Further addition of sodium chloride did not significantly increase the recovery rate. When sodium chloride was insufficient, the solubility of propylene carbonate in water remained relatively high, resulting in a smaller volume of propylene carbonate collected. Therefore, a sodium chloride concentration of 160 mg / mL was selected for the separation and recovery operation.

[0044] Example 4: Comparison of Extractant Types (1) Experimental method: The method is the same as that in Example 1A, except that different extractants (water and aqueous solution of propylene carbonate) are used to extract sorghum protein.

[0045] (2) Experimental results: such as Figure 7 As shown.

[0046] (3) Results analysis: The extraction efficiency of protein by aqueous propylene carbonate was significantly higher than that of pure water extraction (P<0.05).

[0047] Comparison with existing extraction methods 1. The propylene carbonate extractant designed in this invention has lower toxicity and is more environmentally friendly compared to existing acid-base extractants.

[0048] 2. Compared with enzymatic hydrolysis, the ultrasonic-assisted propylene carbonate extraction method designed in this invention can reduce extraction costs and shorten extraction time.

[0049] 3. The propylene carbonate extractant designed in this invention can be separated by a simple salt-induced phase, avoiding the problems of high energy consumption and long separation time of traditional separation methods such as distillation and column chromatography.

[0050] 4. Compared with pure water, propylene carbonate aqueous solution has a higher extraction rate of mixed grain protein.

[0051] In summary, this invention utilizes ultrasound-assisted extraction of propylene carbonate from sorghum protein and achieves the recovery of both sorghum protein and propylene carbonate. The innovation of this invention lies in designing a mixed solution of propylene carbonate and water as the extraction agent, and using ultrasound-assisted extraction of sorghum protein. Propylene carbonate can be recovered by adding sodium chloride for phase separation and used to extract new sorghum protein. The protein can be obtained by freeze-drying the aqueous phase; the extraction rate of protein from the propylene carbonate aqueous solution is higher than that from pure water. This method solves the problems of high solvent toxicity, high reagent cost, long extraction time, difficult separation, high energy consumption, and difficulty in solvent recovery and reuse in current protein extraction methods.

[0052] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for ultrasound-assisted extraction of protein from grains using propylene carbonate, characterized in that, Includes the following steps: S1, Preparation of the extractant solution; Prepare an extraction solution by mixing water and propylene carbonate in a preset volume ratio; S2, extraction and recycling of protein from whole grains; Weigh the mixed grain powder, add the extractant solution, perform ultrasonic-assisted extraction, collect the supernatant after centrifugation, add sodium chloride to the supernatant, vortex and separate the two phases. The upper layer is the aqueous phase and the lower layer is the propylene carbonate phase. The mixed grain protein in the extractant solution is recovered to the upper aqueous phase. After freeze-drying, the protein solid powder can be obtained. S3, recycling and reuse of propylene carbonate; The propylene carbonate phase obtained in step S2 is recovered, and new grain protein is extracted again following steps S1 and S2.

2. The method for extracting protein from grains using ultrasound-assisted propylene carbonate according to claim 1, characterized in that, In step S1, the volume ratio of water to propylene carbonate is 1:1 to 9:

1.

3. The method for extracting protein from grains using ultrasound-assisted propylene carbonate according to claim 1, characterized in that, In step S2, the grain powder is sorghum powder.

4. The method for extracting protein from grains using ultrasound-assisted propylene carbonate according to claim 1, characterized in that, In step S2, the ratio of grain powder to extractant solution is 1:10 to 1:50 g / mL.

5. The method for extracting protein from grains using ultrasound-assisted propylene carbonate according to claim 1, characterized in that, In step S2, the ultrasonic time is 0~60 min and the ultrasonic temperature is 20~100 °C.

6. The method for extracting protein from grains using ultrasound-assisted propylene carbonate according to claim 1, characterized in that, In step S2, the mass concentration of sodium chloride is 40~200 mg / mL.

7. The method for ultrasound-assisted extraction of protein from grains using propylene carbonate according to claim 1, characterized in that, In step S2, the centrifugal force is 9000 r / min, the centrifugation time is 10 min, and the vortexing time is 1 min.

8. The method for ultrasound-assisted extraction of protein from grains using propylene carbonate according to any one of claims 1-7, characterized in that, The volume ratio of water to propylene carbonate was 5:1, the ratio of grain powder to extractant solution was 1:20 g / mL, the ultrasonic time was 30 min, the ultrasonic temperature was 60°C, and the mass concentration of sodium chloride was 160 mg / mL.

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