Ultrasonic extraction method of Maotai-flavor liquor vinasse polysaccharide

Abstract

The invention discloses an ultrasonic extraction method of Maotai-flavor liquor vinasse polysaccharide, and relates to the technical field of comprehensive utilization of liquor brewing waste materials, the ultrasonic extraction method comprises the following steps: S1, raw material pretreatment: drying liquor vinasse; s2, ultrasonic water extraction: putting the dried Baijiu vinasse into an extraction tank, adding purified water as an extraction solvent, carrying out constant-temperature ultrasonic extraction, filtering, and removing insoluble impurities to obtain a primary extracting solution; s3, concentration: putting the primary extracting solution into a rotary evaporator, carrying out vacuum concentration, and concentrating the primary extracting solution to obtain a concentrated extracting solution; s4, alcohol precipitation: adding absolute ethyl alcohol into the concentrated extracting solution, uniformly mixing, standing overnight, centrifugally separating the precipitate after the polysaccharide is completely precipitated, discarding the supernate, and collecting the polysaccharide precipitate; s5, purification: deproteinization is performed by adopting an enzyme method in combination with a Sevage method. According to the method, the Maotai-flavor liquor vinasse polysaccharide can be efficiently extracted, and the biological activity of the polysaccharide is kept in the extraction process.

Description

Technical Field

[0001] The invention relates to the technical field of comprehensive utilization of liquor brewing waste, and more specifically to the technical field of an ultrasonic extraction method for polysaccharides from sauce-flavor liquor lees. Background Art

[0002] Liquor lees are the main by-product of traditional solid liquor production. In recent years, with the continuous improvement of people's living standards, the sauce-flavor liquor industry has developed rapidly, producing a large amount of lees. If it is not used reasonably and effectively, it is easy to cause resource waste and environmental pollution. Liquor lees contain high levels of crude starch, protein, cellulose, hemicellulose, rich amino acids, mineral elements and other nutrients, so they are widely used in the production of animal feed, liquor lees, organic acids, edible fungi cultivation, etc. However, the high added value and deep utilization of liquor lees need to be further developed, among which active polysaccharides are a good research direction.

[0003] Polysaccharides extracted from brewing raw materials have good antioxidant activity and have become one of the hot spots in future food research and development. The biological activity of polysaccharides is related to their structural characteristics such as structure, molecular weight and monosaccharide composition, and different extraction methods have an impact on the chemical structure, monosaccharide composition and molecular weight of polysaccharides. There is an urgent need to solve these problems in this field to increase the high added value and deep utilization of Maotai-flavor liquor lees and reduce resource waste and environmental pollution. Summary of the invention

[0004] The purpose of the present invention is to solve the above technical problems and provide a method for ultrasonic extraction of polysaccharides from lees of Maotai-flavor liquor.

[0005] In order to achieve the above-mentioned purpose, the present invention specifically adopts the following technical solutions:

[0006] The present invention provides a method for ultrasonically extracting polysaccharides from Maotai-flavor liquor lees, comprising the following steps:

[0007] S1. Raw material pretreatment: drying the liquor lees to control the water content at 8-14%;

[0008] S2, ultrasonic water extraction: placing the dried liquor lees in an extraction tank, adding purified water as an extraction solvent, performing constant temperature ultrasonic extraction, filtering, and removing insoluble impurities to obtain a preliminary extract;

[0009] S3. Concentration: placing the preliminary extract in a rotary evaporator for vacuum concentration to concentrate the preliminary extract to obtain a concentrated extract.

[0010] S4, alcohol precipitation: add anhydrous ethanol to the concentrated extract, mix well, let stand overnight, wait for the polysaccharide to precipitate completely, centrifuge to separate the precipitate, discard the supernatant, and collect the polysaccharide precipitate;

[0011] S5. Purification: Deproteinization was performed by combining enzyme method with Sevage method.

[0012] In one embodiment, in step S2, the dried liquor lees are placed in an extraction tank, pure water is added as an extraction solvent at a solid-liquid ratio of 1:30, constant temperature ultrasonic extraction is performed at a certain time and power, and the insoluble impurities are removed to obtain a preliminary extract.

[0013] In one embodiment, in step S3, the preliminary extract is placed in a rotary evaporator and vacuum concentrated to concentrate the preliminary extract to 1 / 5 of its original volume to obtain a concentrated extract.

[0014] In one embodiment, in step S4, 4 times the volume of anhydrous ethanol is added to the concentrated extract, mixed evenly, and allowed to stand overnight until the polysaccharide is completely precipitated, the precipitate is separated by centrifugation, the supernatant is discarded, and the polysaccharide precipitate is collected.

[0015] In one embodiment, in step S5, the specific steps of deproteinizing by using an enzyme method combined with a Sevage method are as follows:

[0016] S51, preparing papain and polysaccharide, mixing and dissolving;

[0017] S52, add Sevage reagent, put it into a shaker and shake vigorously, centrifuge, take the supernatant, and repeat several times until no protein suspension is produced;

[0018] S53, transferring the supernatant to a rotary evaporator to remove the organic phase by evaporation under reduced pressure, and freeze-drying and storing for later use;

[0019] S54, dissolving the deproteinized polysaccharide in distilled water, purifying it with a DEAE-Sepharose Fast F l ow ion exchange column, using distilled water as an eluent for elution, collecting the eluent, concentrating it under reduced pressure, and then freeze-drying it to obtain a water-soluble neutral polysaccharide from liquor lees.

[0020] In one embodiment, in step S51, the papain mass:polysaccharide mass=1:10 is mixed and dissolved, enzymolysis is performed in a 55° C. water bath for 2.5 h, and the papain is inactivated by heating in boiling water for 10 min.

[0021] In one embodiment, in step S52, 1 / 5 volume of Sevage reagent (chloroform: n-butanol = 4:1) is added, the mixture is shaken on a shaker for 30 minutes, centrifuged at 4000 r / min for 5 minutes, the supernatant is taken, and the process is repeated multiple times until no protein suspension is produced.

[0022] In one embodiment, in step S54, the deproteinized polysaccharide is dissolved in distilled water and purified using a DEAE-Sepharose Fast F l ow ion exchange column (30×5 cm) with distilled water as the eluent at a flow rate of 3 mL / min for 5 h. The collected eluate is concentrated under reduced pressure at 60°C and then freeze-dried to obtain white wine lees water-soluble neutral polysaccharide (VP).

[0023] The beneficial effects of the present invention are as follows:

[0024] The invention provides a process for extracting polysaccharides from lees of Maotai-flavor liquor, which can efficiently extract polysaccharides from Maotai-flavor liquor lees and maintain the biological activity of polysaccharides during the extraction process. The process is simple to operate, highly feasible, and suitable for industrial production, providing a new idea for the treatment of Maotai-flavor liquor. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for use in the embodiments. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0026] Figure 1 This is a graph showing the effect of different ultrasonic times on polysaccharide extraction rates according to the method of the present invention;

[0027] Figure 2 This is a graph showing the effect of different material-liquid ratios on polysaccharide extraction rates according to the method of the present invention;

[0028] Figure 3 This is a graph showing the effect of different ultrasonic powers on polysaccharide extraction rates according to the method of the present invention;

[0029] Figure 4 The method of the present invention is a three-factor interaction response surface and contour map of material-liquid ratio and ultrasonic time (a), material-liquid ratio and power (b), and ultrasonic time and power (c);

[0030] Figure 5 This is a molecular weight distribution diagram of polysaccharides in liquor waste by the method of the present invention;

[0031] Figure 6 The ABTS free radical scavenging rate of polysaccharide at different concentrations according to the method of the present invention is Figure 6 (a) DPPH free radical scavenging rate Figure 6 (b) Hydroxyl radical scavenging rate Figure 6 (c) Fe 2+ Chelating ability Figure 6 (d);

[0032] Figure 7 This is a graph showing the inhibitory activity of water-soluble neutral polysaccharide (VP) on α-glucosidase. DETAILED DESCRIPTION

[0033] In order to make the technical problems, technical solutions and technical effects of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Generally, the components of the embodiments of the present invention described and shown in the drawings here can be arranged and designed in various different configurations.

[0034] Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the invention claimed for protection, but merely represents selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0035] An ultrasonic extraction process for highly efficient and biologically active Maotai-flavor liquor lees polysaccharide comprises the following steps:

[0036] S1. Raw material pretreatment: Dry the liquor lees to control the water content at 8-14%.

[0037] S2. Ultrasonic water extraction: Place the dried liquor lees in an extraction tank, add pure water as the extraction solvent at a solid-liquid ratio of 1:30, perform constant temperature ultrasonic extraction at a certain time and power, filter, remove insoluble impurities, and obtain a preliminary extract.

[0038] S3. Concentration: Place the preliminary extract in a rotary evaporator and perform vacuum concentration to concentrate the extract to 1 / 5 of the original volume to obtain a concentrated extract.

[0039] S4, alcohol precipitation: add 4 times the volume of anhydrous ethanol to the concentrated extract, mix well, let stand overnight, wait for the polysaccharide to precipitate completely, centrifuge and separate the precipitate, discard the supernatant, and collect the polysaccharide precipitate.

[0040] S5, purification: deproteinization by enzyme method combined with Sevage method. First, the mixture was dissolved according to the mass of papain: mass of polysaccharide = 1:10, enzymolysis was carried out in a water bath at 55°C for 2.5h, and the papain was inactivated by heating in boiling water for 10min. Then, 1 / 5 volume of Sevage reagent (chloroform: n-butanol = 4:1) was added, shaken vigorously for 30min, centrifuged (4000r / min, 5min), and the supernatant was taken. The supernatant was transferred to a rotary evaporator and evaporated under reduced pressure to remove the organic phase, and freeze-dried and stored for later use. The deproteinized polysaccharide was dissolved in distilled water and purified using a DEAE-Sepharose Fast F l ow ion exchange column (30×5cm). Distilled water was used as the eluent, and the eluent was eluted at a flow rate of 3mL / min for 5h. The collected eluent was concentrated under reduced pressure at 60°C, and then freeze-dried to obtain a sauce-flavored liquor lees polysaccharide with good biological activity.

[0041] Example 1

[0042] This embodiment provides an ultrasonic extraction process for polysaccharides from Maotai-flavor liquor lees that is efficient and has good biological activity, comprising the following steps:

[0043] S1. Raw material pretreatment: drying the liquor lees to control the water content at 10%;

[0044] S1. Ultrasonic water extraction: Place the dried liquor lees in an extraction tank, add pure water as the extraction solvent at a solid-liquid ratio of 1:40, process under ultrasonic extraction conditions of 400W for 30 minutes, filter, remove insoluble impurities, and obtain a preliminary extract;

[0045] S1. Concentration: placing the preliminary extract in a rotary evaporator and performing vacuum concentration to concentrate the extract to 1 / 5 of the original volume to obtain a concentrated extract;

[0046] S4, alcohol precipitation: add 4 times the volume of anhydrous ethanol to the concentrated extract, mix well, let stand overnight, wait for the polysaccharide to precipitate completely, centrifuge and separate the precipitate, discard the supernatant, and collect the polysaccharide precipitate;

[0047] S5, purification: deproteinization by enzyme method combined with Sevage method. First, the mixture was dissolved according to the mass of papain: mass of polysaccharide = 1:10, enzymolysis was performed in a water bath at 55°C for 2.5h, and the papain was inactivated by heating in boiling water for 10min. Then, 1 / 5 volume of Sevage reagent (chloroform: n-butanol = 4:1) was added, shaken vigorously for 30min, centrifuged (4000r / min, 5min), and the supernatant was taken. The supernatant was transferred to a rotary evaporator and evaporated under reduced pressure to remove the organic phase, and freeze-dried and stored for later use. The deproteinized polysaccharide was dissolved in distilled water and purified using a DEAE-Sepharose Fast F l ow ion exchange column (30×5cm). Distilled water was used as the eluent, and the eluent was eluted at a flow rate of 3mL / min for 5h. The collected eluent was concentrated under reduced pressure at 60°C, and then freeze-dried to obtain a Maotai-flavor liquor lees polysaccharide with good biological activity.

[0048] The polysaccharide extraction rate of this embodiment is 5.29%, the ABTS free radical scavenging rate is 83.33%, and the α-glucosidase inhibition activity is 92.33%. It is a white wine lees polysaccharide with excellent antioxidant and hypoglycemic activities.

[0049] Example 2

[0050] This embodiment provides an ultrasonic extraction process for polysaccharides from Maotai-flavor liquor lees that is efficient and has good biological activity, comprising the following steps:

[0051] S1. Raw material pretreatment: drying the liquor lees to control the water content at 10%;

[0052] S2, ultrasonic water extraction: the dried liquor lees are placed in an extraction tank, pure water is added as an extraction solvent at a solid-liquid ratio of 1:40, and the mixture is treated under ultrasonic extraction conditions of 500W for 20 minutes, filtered, and insoluble impurities are removed to obtain a preliminary extract;

[0053] S3, concentration: placing the preliminary extract in a rotary evaporator and performing vacuum concentration to concentrate the extract to 1 / 5 of the original volume to obtain a concentrated extract;

[0054] S4, alcohol precipitation: add 4 times the volume of anhydrous ethanol to the concentrated extract, mix well, let stand overnight, wait for the polysaccharide to precipitate completely, centrifuge and separate the precipitate, discard the supernatant, and collect the polysaccharide precipitate;

[0055] S5, purification: deproteinization by enzyme method combined with Sevage method. First, the mixture was dissolved according to the mass of papain: mass of polysaccharide = 1:10, enzymolysis was performed in a water bath at 55°C for 2.5h, and the papain was inactivated by heating in boiling water for 10min. Then, 1 / 5 volume of Sevage reagent (chloroform: n-butanol = 4:1) was added, shaken vigorously for 30min, centrifuged (4000r / min, 5min), and the supernatant was taken. The supernatant was transferred to a rotary evaporator and evaporated under reduced pressure to remove the organic phase, and freeze-dried and stored for later use. The deproteinized polysaccharide was dissolved in distilled water and purified using a DEAE-Sepharose Fast F l ow ion exchange column (30×5cm). Distilled water was used as the eluent, and the eluent was eluted at a flow rate of 3mL / min for 5h. The collected eluent was concentrated under reduced pressure at 60°C, and then freeze-dried to obtain a Maotai-flavor liquor lees polysaccharide with good biological activity.

[0056] The polysaccharide extraction rate of this embodiment is 4.41%, the ABTS free radical scavenging rate is 81.83%, and the α-glucosidase inhibition activity is 88.99%. It is a white wine lees polysaccharide with excellent antioxidant and hypoglycemic activities.

[0057] Example 3

[0058] This embodiment provides an ultrasonic extraction process for polysaccharides from Maotai-flavor liquor lees that is efficient and has good biological activity, comprising the following steps:

[0059] S1. Raw material pretreatment: drying the liquor lees to control the water content at 10%;

[0060] S2, ultrasonic water extraction: the dried liquor lees are placed in an extraction tank, pure water is added as an extraction solvent at a solid-liquid ratio of 1:50, and the mixture is treated under ultrasonic extraction conditions of 400W for 20 minutes, filtered, and insoluble impurities are removed to obtain a preliminary extract;

[0061] S3, concentration: placing the preliminary extract in a rotary evaporator and performing vacuum concentration to concentrate the extract to 1 / 5 of the original volume to obtain a concentrated extract;

[0062] S4, alcohol precipitation: add 4 times the volume of anhydrous ethanol to the concentrated extract, mix well, let stand overnight, wait for the polysaccharide to precipitate completely, centrifuge and separate the precipitate, discard the supernatant, and collect the polysaccharide precipitate;

[0063] S5, purification: deproteinization by enzyme method combined with Sevage method. First, the mixture was dissolved according to the mass of papain: mass of polysaccharide = 1:10, enzymolysis was performed in a water bath at 55°C for 2.5h, and the papain was inactivated by heating in boiling water for 10min. Then, 1 / 5 volume of Sevage reagent (chloroform: n-butanol = 4:1) was added, shaken vigorously for 30min, centrifuged (4000r / min, 5min), and the supernatant was taken. The supernatant was transferred to a rotary evaporator and evaporated under reduced pressure to remove the organic phase, and freeze-dried and stored for later use. The deproteinized polysaccharide was dissolved in distilled water and purified using a DEAE-Sepharose Fast F l ow ion exchange column (30×5cm). Distilled water was used as the eluent, and the eluent was eluted at a flow rate of 3mL / min for 5h. The collected eluent was concentrated under reduced pressure at 60°C, and then freeze-dried to obtain a Maotai-flavor liquor lees polysaccharide with good biological activity.

[0064] The polysaccharide extraction rate of this embodiment is 4.63%, the ABTS free radical scavenging rate is 80.78%, and the α-glucosidase inhibition activity is 85.65%, which is a white wine lees with excellent antioxidant and hypoglycemic activities.

[0065] The following is an experiment on the method of this embodiment, and the experimental results are obtained and analyzed.

[0066] 1. Single factor experimental results

[0067] 1.1. Effect of ultrasonic time on polysaccharide extraction rate:

[0068] Depend on Figure 1 It can be seen that the method of Example 1 shows an upward trend in the extraction rate of polysaccharides as the ultrasonic time is prolonged, and reaches a peak at 30 minutes. After that, the extraction rate of polysaccharides decreases instead. It may be that the ultrasonic time is too long, which destroys the structure of polysaccharides and degrades them, which leads to a decrease in the extraction rate. Therefore, the optimal ultrasonic extraction time is 30 minutes.

[0069] 1.2. Effect of solid-liquid ratio on polysaccharide extraction rate:

[0070] Depend on Figure 2 It is known that the method of Example 1 shows an upward trend in the extraction rate of polysaccharides with the increase of the solid-liquid ratio. Before the solid-liquid ratio reaches 1:40, the polysaccharide yield increases significantly. After the solid-liquid ratio reaches 1:40, the increase trend of the polysaccharide extraction rate slows down. In addition, too large a solid-liquid ratio will reduce the efficiency of the ultrasonic medium in absorbing ultra-energy and increase energy consumption. Therefore, 1:40 is selected as the optimal solid-liquid ratio.

[0071] 1.3. Effect of ultrasonic power on polysaccharide extraction rate:

[0072] Depend on Figure 3It is known that the extraction rate of polysaccharides in Example 1 gradually increases with the increase of ultrasonic power, and reaches a maximum value at 400 W, and then the extraction rate decreases. It may be that the ultrasonic power is too large, causing the degradation of polysaccharides in the liquor waste, resulting in a decrease in the extraction rate of polysaccharides in the liquor waste.

[18] Therefore, the optimal ultrasonic extraction power of liquor lees should be 400W.

[0073] 2. Response surface experiment results

[0074] 2.1. Box-Behnken test design results:

[0075] According to the results of the single factor experiment, the response surface experiment was designed using the Box-Behnken principle. The experimental results are shown in Table 1.

[0076] Table 1 Response surface experimental design results

[0077]

[0078]

[0079] According to the analysis of the test results, the coded regression equation obtained by fitting the three-factor results is:

[0080] Y=0.053+0.002A-0.003B+0.00006C-0.004AB+0.001AC-0.002BC-0.011A 2 -0

[0081] .004B 2 -0.009C 2 .

[0082] The results of variance analysis are shown in Table 2. The significance of the effects of various combinations and factors in the regression equation on the response value is judged by the P value. P less than 0.01 can be considered extremely significant, and P less than 0.05 can be considered significant. The results show that the model P value <0.0001<0.01, indicating that the regression model is extremely significant and has a high statistical significance. The P value of the solid-liquid ratio is <0.05, and the solid-liquid ratio has a significant effect on the extraction rate. The P value of time is <0.01, indicating that the extraction time has a very significant effect on the extraction rate.

[0083] Table 2 Box-Behnken experimental analysis results

[0084]

[0085] 2.2. Determination of the best extraction process:

[0086] Design-Expert 8.0.6 analysis software was used to analyze the influencing factors of polysaccharide extraction from liquor waste. Combined with the response surface diagram and contour diagram, the effects of the three factors on the polysaccharide extraction rate can be intuitively reflected.

[0087] Combined with Table 2 Figure 4 (a) It can be seen that the contour map is elliptical, and the P value of AB is less than 0.01, indicating that the interaction between the solid-liquid ratio and time has a significant effect on the polysaccharide yield. It can be seen from the figure that the response value increases with the increase of the solid-liquid ratio. When the solid-liquid ratio reaches 1:40, the response value reaches the maximum, and then the response value slowly decreases with the increase of the solid-liquid ratio. When the solid-liquid ratio is 1:40, the response value slowly increases with the increase of time, and then decreases with the increase of ultrasonic time. It may be due to the degradation of polysaccharides caused by too long ultrasonic time, which leads to a rapid decrease in the response value.

[19] .

[0088] Depend on Figure 4 (b) Combined with Table 2, it can be seen that the contour map is circular, and the P value of AC is greater than 0.05, indicating that the interaction between the solid-liquid ratio and the power has no significant effect on the polysaccharide yield. When the solid-liquid ratio is 1:40, the response value first increases with the increase of power, reaches the maximum when the power reaches 400W, and then decreases with the continued increase of power. It may be due to the fact that excessive power causes an increase in bubbles in the solvent during the cavitation process, thereby reducing the efficiency of ultrasonic energy transmission to the ultrasonic medium, and excessive ultrasonic power will lead to the degradation of polysaccharides. ] , which leads to a decrease in the polysaccharide extraction rate.

[0089] Depend on Figure 4 (c) Combined with Table 3, it can be seen that the P value of BC is greater than 0.05, and the interaction between power and time has no significant effect on the polysaccharide yield. If the ultrasonic time is too long or the power is too high, the response value will decrease.

[0090] In summary, the optimal process for ultrasonic extraction of polysaccharides from liquor waste is a solid-liquid ratio of 1:40, a time of 30 min, and a power of 400 W. Under these conditions, the maximum response value predicted by the model is 5.28667%. The results of the validation experiment show that the average polysaccharide extraction rate is 5.16% ± 0.41% (n = 3). This is very close to the predicted value, indicating that the model is effective in optimizing the extraction process.

[0091] 2.3 Monosaccharide composition of liquor lees:

[0092] The monosaccharide composition of polysaccharides is shown in Table 3. Liquor waste polysaccharides are mainly composed of Man (16.65±0.72), Gal (5.49±0.30) and Glc (68.95±0.43).

[0093] Table 3 Monosaccharide composition of polysaccharides from liquor waste

[0094]

[0095]

[0096] 2.4. Molecular weight parameters of polysaccharides from liquor waste:

[0097] The molecular weight distribution of polysaccharides in liquor waste is Figure 5 As shown in the figure, the polysaccharide of liquor lees has four symmetrical peaks, indicating that the polysaccharide of liquor lees extracted by ultrasonic extraction is composed of four polysaccharide components with different molecular weights, and the molecular weight distribution of polysaccharides is 1.14×10 4 ~1.02×10 6 Da. According to their peak retention time, the four components were named P1, P2, P3 and P4. Among the four components, P1 has the largest molecular weight [1.02×10 6 (±0.0.27%)Da], followed by P2 [8.52×10 4 (±0.26%)Da], and the Mw values ​​of P3 and P4 were lower, which were 2.45×10 4 (±0.556%), 1.14×10 4 (±1.49%) Da. Polysaccharides are generally polydisperse compounds with a wide molecular weight distribution. As shown in Table 4, the polydispersity coefficients of P1, P2, P3 and P4 are 1.55, 1.30, 1.04 and 1.20, respectively, indicating that the four components are polysaccharides with a wide distribution.

[0098] Table 3 Molecular weight and conformation parameters of polysaccharides from liquor waste

[0099]

[0100]

[0101] 2.5. Effect of polysaccharides on antioxidant activity in vitro:

[0102] Figure 6 (a) reflects the scavenging ability of white wine lees polysaccharides of different concentrations on ABTS. The scavenging ability of polysaccharides on ABTS is dependent on the concentration. With the increase of concentration, the scavenging ability of polysaccharides on ABTS increases. When the concentration of polysaccharides is 0.1-2 mg / mL, the scavenging ability of polysaccharides on ABTS shows a relatively obvious linear relationship with the concentration. When the concentration of polysaccharides is 5 mg / mL, the scavenging rate of ABTS is 92.23%±0.22%, indicating that white wine lees polysaccharides have good scavenging activity on ABTS free radicals.

[0103] Figure 6(b) shows the scavenging ability of white wine waste polysaccharides of different concentrations on DPPH. The scavenging ability of polysaccharides on DPPH is dependent on the concentration. With the increase of concentration, the scavenging ability of polysaccharides on DPPH increases. When the concentration of polysaccharides is 5 mg / mL, its scavenging ability on DPPH is 43.32%±1.29%.

[0104] Figure 6 (c) shows that the scavenging ability of white wine waste polysaccharides at different concentrations on hydroxyl radicals gradually increases with the increase of concentration, showing concentration dependence. When the concentration of polysaccharides is 5 mg / mL, the scavenging ability of hydroxyl radicals can reach 51.55%±1.03%.

[0105] Figure 6 (d) The effect of different concentrations of liquor waste polysaccharides on Fe 3+ With the increase of polysaccharide concentration, the reduction capacity of polysaccharide on Fe 3+ The reducing ability of the sample solution increased, showing an obvious linear relationship. When the polysaccharide concentration was 5 mg / mL, its absorbance was 0.35. The greater the absorbance of the sample solution, the stronger the antioxidant capacity of the sample.

[0106] Some researchers have studied the antioxidant activity of cornus seed polysaccharides. When the concentration of cornus seed polysaccharides is 5 mg / mL, the scavenging capacity of ABTS, DPPH and hydroxyl radicals is 61.68%, 68.61% and 72.24% respectively. It has good antioxidant properties and is a potential natural antioxidant. Polysaccharides with lower molecular weight have stronger antioxidant capacity, and ultrasound can properly degrade high molecular weight polysaccharides and enhance their antioxidant activity. Ultrasonic-extracted white wine lees polysaccharides have significant antioxidant properties. When the concentration is 5 mg / mL, the scavenging rate of ABTS free radicals is significantly higher than that of cornus seed polysaccharides, reaching 92.23% ± 0.22%. It is expected to be used in the development of natural antioxidants.

[0107] 2.6. Glucosidase inhibitory activity:

[0108] α-glucosidase inhibitors can reduce the release of glucose from starch and delay sugar absorption by inhibiting the activity of sugar hydrolases in the small intestine. Therefore, natural α-glucosidase inhibitors have attracted great attention as oral hypoglycemic drugs for diabetic patients. The inhibitory activity of polysaccharides on α-glucosidase was determined. The results are as follows Figure 7 The α-glucosidase inhibitory activity was concentration-dependent and had a good linear relationship when the concentration was 0.1-5 mg / mL. When the concentration reached 5 mg / mL, the inhibition rate reached 88.99%±3.34%.

Claims

1. A method for ultrasonic extraction of polysaccharides from Maotai-flavor liquor lees, characterized in that: The steps include: S1. Raw material pretreatment: drying the liquor lees to control the water content at 8-14%; S2, ultrasonic water extraction: placing the dried liquor lees in an extraction tank, adding purified water as an extraction solvent, performing constant temperature ultrasonic extraction, filtering, and removing insoluble impurities to obtain a preliminary extract; S3, concentration: placing the preliminary extract in a rotary evaporator and performing vacuum concentration to concentrate the preliminary extract to obtain a concentrated extract; S4, alcohol precipitation: add anhydrous ethanol to the concentrated extract, mix well, let stand overnight, wait for the polysaccharide to precipitate completely, centrifuge to separate the precipitate, discard the supernatant, and collect the polysaccharide precipitate; S5. Purification: Deproteinization was performed by combining enzyme method with Sevage method.

2. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 1, characterized in that: In step S2, the dried liquor lees are placed in an extraction tank, pure water is added as an extraction solvent at a solid-liquid ratio of 1:30, constant temperature ultrasonic extraction is performed according to time and power, and the insoluble impurities are removed by filtering to obtain a preliminary extract.

3. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 1, characterized in that: In step S3, the preliminary extract is placed in a rotary evaporator and vacuum concentrated to 1 / 5 of the original volume to obtain a concentrated extract.

4. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 1, characterized in that: In step S4, 4 times the volume of anhydrous ethanol is added to the concentrated extract, mixed evenly, and allowed to stand overnight until the polysaccharide is completely precipitated, the precipitate is separated by centrifugation, the supernatant is discarded, and the polysaccharide precipitate is collected.

5. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 1, characterized in that: In step S5, the specific steps of deproteinizing by using an enzyme method combined with a Sevage method are as follows: S51, preparing papain and polysaccharide, mixing and dissolving; S52, add Sevage reagent, put it into a shaker and shake vigorously, centrifuge, take the supernatant, and repeat several times until no protein suspension is produced; S53, transferring the supernatant to a rotary evaporator to remove the organic phase by evaporation under reduced pressure, and freeze-drying and storing for later use; S54. Dissolve the deproteinized polysaccharide in distilled water, purify it with a DEAE-Sepharose Fast Flow ion exchange column, use distilled water as an eluent for elution, collect the eluent, concentrate it under reduced pressure, and then freeze-dry it to obtain a water-soluble neutral polysaccharide from liquor lees.

6. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 5, characterized in that: In step S51, the mixture is dissolved in a ratio of papain mass to polysaccharide mass = 1:10, enzymolysis is carried out in a water bath at 55° C. for 2.5 h, and the papain is inactivated by heating in boiling water for 10 min.

7. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 5, characterized in that: In step S52, 1 / 5 volume of Sevage reagent was added and the mixture was shaken on a shaker for 30 min, centrifuged at 4000 r / min for 5 min, and the supernatant was taken. The process was repeated several times until no protein suspension was produced.

8. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 7, characterized in that: The components of the Sevage reagent include chloroform and n-butanol, and the ratio of chloroform to n-butanol is 4:

1.

9. The ultrasonic extraction method of polysaccharides from Maotai-flavor liquor lees according to claim 5, characterized in that: In step S54, the deproteinized polysaccharide is dissolved in distilled water and purified using a DEAE-Sepharose Fast Flow ion exchange column with distilled water as the eluent at a flow rate of 3 mL / min for 5 hours. The collected eluate is concentrated under reduced pressure at 60° C. and then freeze-dried to obtain white wine lees water-soluble neutral polysaccharide.

Images