CN113087815A - Application of beautiful millettia root polysaccharide or modified polysaccharide thereof in delaying grease oxidation - Google Patents
Application of beautiful millettia root polysaccharide or modified polysaccharide thereof in delaying grease oxidation Download PDFInfo
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Abstract
The invention discloses application of millettia speciosa champ polysaccharide or modified polysaccharide thereof in delaying grease oxidation. According to the invention, the millettia speciosa champ polysaccharide is added into the grease to effectively delay the oxidation of the grease, but the emulsification stability of the millettia speciosa champ polysaccharide is relatively poor, so that the millettia speciosa champ polysaccharide is modified by adopting a strategy of grafting hydrophobic groups, the emulsification performance of the modified millettia speciosa champ polysaccharide can be obviously improved, and the emulsion added with the modified millettia speciosa champ polysaccharide can keep the same lipid oxidation rate as the emulsion added with the millettia speciosa champ polysaccharide, because more polysaccharides of the modified millettia speciosa champ polysaccharide are adsorbed to an oil-water interface, a compact and viscoelastic interface layer is formed, a physical shielding effect can be achieved, and further the. Therefore, the beautiful millettia root polysaccharide and the modified polysaccharide thereof can be used for delaying the oxidation of grease.
Description
Technical Field
The invention relates to the application field of polysaccharides, in particular to application of beautiful millettia root polysaccharides or modified polysaccharides thereof in delaying grease oxidation.
Background
Millettia speciosa is the main variety of south Chinese medicines, belongs to the family of Leguminosae, and is the plant Millettia dielsiana, which is mainly distributed in south China and some countries in southeast Asia. Radix millettiae speciosae is a Chinese medicinal material used as both medicine and food, and has the effects of treating arthritis and bronchitis, protecting liver, strengthening tendons and bones, tonifying deficiency, moistening lung and the like.
Recently, beautiful millettia root polysaccharide is found to have certain emulsifying capacity, but the stable emulsion of beautiful millettia root polysaccharide is unstable, and serious delamination phenomenon appears immediately after preparation. Further, the effect of the modified polysaccharide on oxidation of fats and oils in an emulsion has not been clarified yet. Therefore, in order to broaden the application of beautiful millettia root polysaccharide (MSCP) in the food industry, particularly in the field of milky foods, it is necessary to modify beautiful millettia root polysaccharide to improve its emulsifying properties and to explore the change in oxidation of fats and oils before and after modification.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provide the application of the millettia speciosa champ polysaccharide and/or the modified polysaccharide thereof in delaying the oxidation of grease.
The purpose of the invention is realized by the following technical scheme:
application of beautiful millettia root polysaccharide and/or modified polysaccharide thereof in delaying grease oxidation.
The beautiful millettia root polysaccharide can be prepared according to examples 1-3 in Chinese patent (patent number is 201911238835X, named beautiful millettia root polysaccharide, a preparation method and application thereof in the antibacterial aspect).
The beautiful millettia root modified polysaccharide is acetylated beautiful millettia root modified polysaccharide, namely the beautiful millettia root modified polysaccharide which is acetylated by an acetic anhydride method and the like; preferably prepared by the following method:
dissolving the beautiful millettia root polysaccharide in water, adjusting the pH value to 8.0-9.0, then dropwise adding acetic anhydride, dropwise adding an alkaline solution to maintain the pH value of the solution at 8.0-8.5, adjusting the pH value to 7.0 after the reaction is finished, dialyzing, and freeze-drying to obtain the modified beautiful millettia root polysaccharide.
The dosage of the beautiful millettia root polysaccharide is calculated according to the proportion of 30 +/-5 mg of beautiful millettia root polysaccharide per milliliter (mL) of water.
The pH value is preferably adjusted to 8.0-8.5; more preferably, the pH is adjusted to 8.0.
The water is preferably distilled water.
The amount of the acetic anhydride is calculated according to the proportion of 60-300 mg of beautiful millettia root polysaccharide per milliliter (mL) of acetic anhydride.
The alkaline solution is NaOH solution; NaOH solution having a concentration of 5mol/L is preferred.
The reaction time is 3-5 h; preferably 4 hours.
The pH value is adjusted to 7.0 by adopting HCl solution; preferably, the adjustment is carried out using a 5mol/L HCl solution.
The dialysis is carried out by adopting a dialysis bag with the molecular weight cutoff of 3500 Da; preferably, the dialysis is carried out for more than 48h by using a dialysis bag with the molecular weight cut-off of 3500 Da.
The application of the beautiful millettia root polysaccharide and/or the modified polysaccharide thereof in delaying the oxidation of the grease is that the beautiful millettia root polysaccharide and/or the beautiful millettia root modified polysaccharide are added into the grease and are uniformly mixed (emulsion is formed) so as to delay the oxidation of the grease.
The oil is edible oil or industrial oil; preferably soybean oil or bulk oil, etc.
The uniform mixing can be carried out in a homogenizing mode; preferably, a high-speed homogenizer is adopted for homogenization, and then a micro-jet device is adopted for homogenization.
The conditions for homogenizing in the high-speed homogenizer are preferably as follows: homogenizing at 20000rpm for 3-5 min.
The pressure for homogenizing the micro-fluidic device is preferably as follows: 75 MPa.
The application of the beautiful millettia root polysaccharide and/or the modified polysaccharide thereof in preparing the emulsifier.
The application of the beautiful millettia root polysaccharide and/or the modified polysaccharide thereof in oil-in-water emulsion.
Compared with the prior art, the invention has the following advantages and effects:
(1) the millettia speciosa champ polysaccharide is modified by adopting a grafting hydrophobic group strategy, the emulsifying property of the modified millettia speciosa champ polysaccharide is obviously improved, and the modified millettia speciosa champ polysaccharide can be used for stabilizing an oil-in-water emulsion, so that the modified millettia speciosa champ polysaccharide can be used for developing an emulsifier.
(2) The particle size of the modified beautiful millettia root polysaccharide stable emulsion is smaller than that of the beautiful millettia root polysaccharide stable emulsion, and the formed smaller liquid drops enlarge the contact specific surface area, generally accelerate the oxidation of grease, but the experimental result shows that the emulsion prepared by the modified beautiful millettia root polysaccharide still keeps the same lipid oxidation rate as the emulsion prepared by the beautiful millettia root polysaccharide, because the modified beautiful millettia root polysaccharide has more polysaccharides adsorbed to an oil-water interface, a compact and viscoelastic interface layer is formed, and the interface layer can play a physical shielding role for some oxidation promoters, so the beautiful millettia root polysaccharide or the modified beautiful millettia root polysaccharide can effectively delay the oxidation rate of the lipid.
Drawings
FIG. 1 is a photograph of emulsions prepared from Millettia speciosa MSCP, AC-MSCP1, AC-MSCP2, AC-MSCP3 and gum arabic GA (in the figure, A: MSCP; B: AC-MSCP 1; C: AC-MSCP 2; D: AC-MSCP 3; E: gum arabic GA).
FIG. 2 is a graph of the particle size distribution of emulsions prepared with Millettia speciosa polysaccharide MSCP, modified Millettia speciosa polysaccharide AC-MSCP, and gum arabic GA.
Figure 3 is a graph of the Mean particle size (Mean droplet size) of emulsions prepared with beautiful millettia root polysaccharide MSCP, modified beautiful millettia root polysaccharide AC-MSCP and gum arabic GA.
Figure 4 is a graph of the change in mean particle size of emulsions prepared with beautiful millettia root polysaccharide MSCP, modified beautiful millettia root polysaccharide AC-MSCP and gum arabic GA during storage.
FIG. 5 is a graph of the scavenging of DPPH and Hydroxyl (Hydroxyl) radicals by Millettia speciosa polysaccharide MSCP, modified Millettia speciosa polysaccharide AC-MSCP; wherein a is the clearance rate of DPPH; b is the clearance rate of hydroxyl free radical.
FIG. 6 is a graph of the change in Lipid hydroperoxides (Lipid hydroperoxides) and Malondialdehyde (MDA) during storage; wherein a is the change condition of the grease hydroperoxide; b is the change of malondialdehyde.
Fig. 7 is a graph of the results of a rheological dynamic frequency sweep.
Detailed Description
The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto. Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated. The test methods in the following examples, in which specific experimental conditions are not specified, are generally performed according to conventional experimental conditions or according to the experimental conditions recommended by the manufacturer. Unless otherwise specified, reagents and starting materials for use in the present invention are commercially available.
The pretreatment method of the AB-8 macroporous resin, the determination method of the content of the modified beautiful millettia root polysaccharide and the preparation method of the beautiful millettia root polysaccharide used in the embodiment of the invention can be carried out by referring to Chinese patent (with the patent number of 201911238835X, the name of beautiful millettia root polysaccharide and the preparation method and the application thereof in the antibacterial aspect), and concretely comprises the following steps:
1. the AB-8 macroporous resin (purchased from Shanghai leaf Biotechnology Co., Ltd.) used in the embodiment of the invention is pretreated firstly, and the specific steps are as follows:
(1) the column (column height 90cm, internal diameter 5cm) was cleaned before packing to prevent contamination of the resin by harmful substances and to drain the water in the column.
(2) Adding ethanol or methanol which is 0.4-0.5 times of the volume of the filled resin into the adsorption column, putting new resin into the column (AB-8 is filled into the chromatographic column and has the height of 65cm) to ensure that the liquid level is about 0.3m higher than the resin layer, and soaking for 24 hours.
(3) 2BV (BV means column volume) of ethanol or methanol is used for soaking for 4-5 hours, and the flow rate is 2 BV/h.
(4) Passing through the resin layer with ethanol or methanol at a flow rate of 2BV/h until the effluent is not cloudy, and washing with water at the same flow rate.
(5) And (3) passing a HCl solution with the mass fraction of 2BV and the mass fraction of 5% through the resin layer at the flow rate of 4-6 BV/h, soaking for 2-4 hours, and then washing with water at the same flow rate until the pH of the effluent is neutral.
(6) And (3) passing a 2BV NaOH solution with the mass fraction of 2% through the resin layer at the flow rate of 4-6 BV/h, soaking for 2-4 hours, and then washing with water at the same flow rate until the pH of the effluent is neutral.
2. The method for determining the content of the modified beautiful millettia root polysaccharide by adopting the phenol-sulfuric acid method in the embodiment of the invention comprises the following steps:
drawing a standard curve: weighing 10mg of glucose dried to constant weight, diluting the glucose to a constant volume with distilled water in a 100mL brown volumetric flask to prepare a standard glucose solution with the concentration of 0.1mg/mL, respectively sucking 0, 0.2, 0.4, 0.6, 0.8, 1.0, 1.2, 1.4 and 1.6mL of the standard glucose solution in a test tube, supplementing the standard glucose solution to 2mL with distilled water, adding 1mL of phenol with the mass fraction of 5% and 5mL of concentrated sulfuric acid (the mass fraction is 95.5%), shaking the glucose solution uniformly, placing the glucose solution at room temperature for 20min, and measuring the absorbance value at 490nm to obtain a regression equation: y is 0.0119x-0.0023 (R)2＝0.9905)。
Preparing a sample to be detected into 1mg/mL, and measuring according to the same method, namely adding 1mL of phenol with the mass fraction of 5% and 5mL of concentrated sulfuric acid, shaking uniformly, placing at room temperature for 20min, measuring the absorbance value at 490nm, substituting the absorbance value into a regression equation, and calculating to obtain the content of the sample to be detected.
Example 1: preparation of modified beautiful millettia root polysaccharide
(1) Preparing beautiful millettia root polysaccharide:
cleaning root blocks of beautiful millettia root (produced in Zhaoqing city, Guangdong province, China), cutting into slices, drying (drying for 15 hours at 60 ℃), crushing, and sieving with a 100-mesh sieve to obtain the root block powder of the beautiful millettia root; extracting radix millettiae speciosae block powder by a hot water reflux method under the conditions of the temperature of 80 ℃, the material-liquid ratio of 1: 10, extracting for 1 h; centrifuging after leaching, taking supernatant, repeatedly extracting for two times according to the conditions, combining the supernatants, and concentrating under reduced pressure at 55 ℃ to 1/4 volumes to obtain a radix millettiae speciosae polysaccharide crude extract;
secondly, in order to remove pigments and proteins, 300mL of the crude extract of the beautiful millettia root polysaccharide is added into an AB-8 macroporous resin column (the column height is 65cm, the inner diameter is 5cm), the mixture is kept stand and adsorbed for 4 hours, then the mixture is eluted by distilled water, the eluent is collected, and the reduced pressure concentration is carried out at 55 ℃ until the volume is 100 mL; removing residual protein by using a Sevag method, adding 1/5 volumes of Sevag reagent (chloroform: n-butyl alcohol is 4: 1, v/v) into the concentrated solution, oscillating for 20min in a shaking table (180rpm), centrifuging for 4min at 4000r/min, taking the supernatant, and repeating the operation for 5 times (namely adding 1/5 volumes of Sevag reagent into the obtained supernatant, oscillating, centrifuging, taking the supernatant, and repeating the operation for 5 times); adding 4 times volume of anhydrous ethanol into the supernatant, precipitating with ethanol, standing in a refrigerator at 4 deg.C for 12h, centrifuging at 4000r/min for 4min, collecting precipitate, adding water (30ml) to redissolve the precipitate, dialyzing with flowing water (3500 Da dialysis bag) for 48h to remove small molecular impurities, and lyophilizing to obtain Millettia speciosa MSCP-1;
(2) preparing modified beautiful millettia root polysaccharide: dissolving 300mg of beautiful millettia root polysaccharide MSCP-1 in 10mL of distilled water, stirring until the MSCP-1 is completely dissolved, adjusting the pH to 8.0, dropwise adding 1mL of acetic anhydride into the polysaccharide solution, and simultaneously dropwise adding NaOH (5M) to maintain the pH of the solution at 8.0-8.5 all the time, wherein the whole reaction lasts for 4 hours; after the reaction was completed, the pH was adjusted back to 7.0 with HCl (5M), dialyzed in distilled water for 48 hours (3500 Da dialysis bag), and lyophilized to obtain modified beautiful millettia root polysaccharide AC-MSCP 1.
Example 2: preparation of modified beautiful millettia root polysaccharide
(1) Preparing beautiful millettia root polysaccharide:
cleaning root blocks of beautiful millettia root (produced in Zhaoqing city, Guangdong province, China), cutting into slices, drying (drying for 15 hours at 60 ℃), crushing, and sieving with a 100-mesh sieve to obtain the root block powder of the beautiful millettia root; extracting radix millettiae speciosae block powder by a hot water reflux method under the conditions that the temperature is 90 ℃, the ratio of material to liquid is 1: 20, extracting for 2 h; centrifuging after leaching, taking supernatant, repeatedly extracting for two times according to the conditions, combining the supernatants, and concentrating under reduced pressure at 65 ℃ to 1/4 volumes to obtain a radix millettiae speciosae polysaccharide crude extract;
secondly, in order to remove pigments and proteins, 300mL of the crude extract of the beautiful millettia root polysaccharide is added into an AB-8 macroporous resin column (the column height is 65cm, the inner diameter is 5cm), the mixture is kept stand and adsorbed for 4 hours, then the mixture is eluted by distilled water, the eluent is collected, and the mixture is decompressed and concentrated to 100mL at 65 ℃; removing residual protein by Sevag method, adding 1/5 volume Sevag reagent (chloroform: n-butanol is 4: 1, v/v) into the concentrated solution, shaking in shaking table (180rpm) for 30min, centrifuging at 5000r/min for 5min, collecting supernatant, and repeating the operation for 5 times; adding 4 times volume of anhydrous ethanol into the supernatant, precipitating with ethanol, standing in a refrigerator at 4 deg.C for 12h, centrifuging at 5000r/min for 5min to obtain precipitate, adding water (30ml) to redissolve the precipitate, dialyzing with flowing water (3500 Da dialysis bag) for 48h to remove small molecular impurities, and lyophilizing to obtain Millettia speciosa MSCP-2;
(2) preparing modified beautiful millettia root polysaccharide: dissolving 300mg of beautiful millettia root polysaccharide MSCP-2 in 10mL of distilled water, stirring until the solution is completely dissolved, adjusting the pH to 8.0, dropwise adding 3mL of acetic anhydride into the polysaccharide solution, and simultaneously dropwise adding NaOH (5M) to maintain the pH of the solution at 8.0-8.5 all the time, wherein the whole reaction lasts for 4 hours; after the reaction was completed, the pH was adjusted back to 7.0 with HCl (5M), dialyzed in distilled water for 48 hours (3500 Da dialysis bag), and lyophilized to obtain modified beautiful millettia root polysaccharide AC-MSCP 2.
Example 3: preparation of modified beautiful millettia root polysaccharide
(1) Preparing beautiful millettia root polysaccharide:
cleaning root blocks of beautiful millettia root (produced in Zhaoqing city, Guangdong province, China), cutting into slices, drying (drying for 15 hours at 60 ℃), crushing, and sieving with a 100-mesh sieve to obtain the root block powder of the beautiful millettia root; extracting radix millettiae speciosae block powder by a hot water reflux method under the conditions that the temperature is 100 ℃, the ratio of material to liquid is 1: 30, the extraction time is 3 hours; centrifuging after leaching, taking supernatant, repeatedly extracting for two times according to the conditions, combining the supernatants, and concentrating under reduced pressure at 75 ℃ to 1/4 volumes to obtain a radix millettiae speciosae polysaccharide crude extract;
secondly, in order to remove pigments and proteins, 300mL of the crude extract of the beautiful millettia root polysaccharide is added into an AB-8 macroporous resin column (the column height is 65cm, the inner diameter is 5cm), the mixture is kept stand and adsorbed for 4 hours, then the mixture is eluted by distilled water, the eluent is collected, and the mixture is decompressed and concentrated to 100mL at the temperature of 75 ℃; removing residual protein by Sevag method, adding 1/5 volume Sevag reagent (chloroform: n-butanol 4: 1, v/v) into the concentrated solution, shaking in shaking table (180rpm) for 40min, centrifuging at 6000r/min for 6min, collecting supernatant, and repeating the operation for 5 times; adding 4 times volume of anhydrous ethanol into the supernatant, precipitating with ethanol, standing in a refrigerator at 4 deg.C for 12h, centrifuging at 6000r/min for 6min to obtain precipitate, adding water (30ml) to redissolve the precipitate, dialyzing with flowing water (3500 Da dialysis bag) for 48h to remove small molecular impurities, and lyophilizing to obtain Millettia speciosa MSCP-3;
(2) preparing modified beautiful millettia root polysaccharide: dissolving 300mg of beautiful millettia root polysaccharide MSCP-3 in 10mL of distilled water, stirring until the MSCP-3 is completely dissolved, adjusting the pH to 8.0, dropwise adding 5mL of acetic anhydride into the polysaccharide solution, and simultaneously dropwise adding NaOH (5M) to keep the pH of the solution at 8.0-8.5 all the time, wherein the whole reaction lasts for 4 hours; after the reaction was completed, the pH was adjusted back to 7.0 with HCl (5M), dialyzed in distilled water for 48 hours (3500 Da dialysis bag), and lyophilized to obtain modified beautiful millettia root polysaccharide AC-MSCP 3.
Example 4: determination of substitution degree of modified beautiful millettia root polysaccharide
Respectively dissolving 20mg of the AC-MSCP1, the AC-MSCP2 and the AC-MSCP3 prepared in the examples 1-3 in 10ml of NaOH (0.01M) solution under stirring to obtain a modified beautiful millettia root polysaccharide mixture; the mixture was then heated to 50 ℃ for 2 hours and the excess NaOH was titrated with 0.01M HCl, with phenolphthalein as an indicator. The results were compared with the unmodified Millettia speciosa MSCP prepared in examples 1-3, respectively, as a blank control. The degree of substitution is calculated according to the following equation:
in the formula, V1 represents the volume of HCl required for titration of the blank, V2 represents the volume of HCl required for titration of the sample, M represents the molecular weight of HCl, and W represents the mass of the sample.
And (4) analyzing results: the degrees of substitution of AC-MSCP1, AC-MSCP2 and AC-MSCP3 were 0.13, 0.43 and 0.70, respectively.
Example 5: molecular weight determination of beautiful millettia root polysaccharide and modified beautiful millettia root polysaccharide
The molecular weights of the Millettia speciosa MSCP-1 prepared in example 1 and the modified Millettia speciosa AC-MSCP1, AC-MSCP2, AC-MSCP3 prepared in examples 1 to 3 were determined by means of a Gel Permeation Chromatography (GPC) system equipped with an Agilent 1260 parallax detector, G3000PWXL(7.8x300 i.d.,5 μm) and G5000PWXL(7.8x300 i.d.,10 μm). The sample amount was 20. mu.L, the mobile phase was 0.02mol/L potassium dihydrogen phosphate, and the flow rate was 0.6 mL/min. Pullulan series dextran (6, 10, 21.7, 48.8, 113, 210, 366, 805, 2500kDa (all available fromShanghai Boo Biotechnology Co., Ltd.) a calibration curve was prepared for the standard. The regression equation is: LogMolWt 1270-2-2.73V3+0.0815V4-0.000968V5(R20.999); in the formula, MolWt represents a molecular weight, and V represents a volume.
And (4) analyzing results: the weight average molecular weights of Millettia speciosa polysaccharide MSCP-1 and modified Millettia speciosa polysaccharides AC-MSCP1, AC-MSCP2 and AC-MSCP3 determined by the above method are 1.56 × 104Da，0.90×104Da，1.46×104Da，1.88×104Da。
Example 6: determination of interfacial tension of beautiful millettia root polysaccharide and modified beautiful millettia root polysaccharide
And (3) measuring the interfacial tension of the modified beautiful millettia root polysaccharide on an oil-water interface by using a surface contact angle measuring instrument. Commercially available soybean oil (a brand of goldfish) was used as the oil phase. MSCP-1 and AC-MSCP1, AC-MSCP2, AC-MSCP3 solutions (2%, w/w) (soybean oil was contained in a glass container and the sample solution was dropped into the soybean oil) were each filled into a syringe and pressurized to produce one drop of solution (8 μ l) in a tube containing soybean oil. The form of the liquid drop is recorded by a camera, the contour of the liquid drop is fitted by a Young-Laplace equation, and the interface tension is automatically calculated by an instrument. Each sample was repeated 10 times.
And (4) analyzing results: the interfacial tensions of Millettia speciosa MSCP-1 and the modified Millettia speciosa AC-MSCP1, AC-MSCP2 and AC-MSCP3 determined by the above method were 11.68mN/m, 8.82mN/m, 6.78mN/m and 6.17mN/m, respectively. It can be seen that the interfacial tension of the modified beautiful millettia root polysaccharide is gradually reduced.
Example 7: determination of emulsification performance of modified beautiful millettia root polysaccharide
(1) The determination method comprises the following steps:
millettia speciosa polysaccharide MSCP-1 (MSCP for short) and modified Millettia speciosa polysaccharide AC-MSCP1, AC-MSCP2, AC-MSCP3 and acacia GA are respectively dissolved in distilled water to prepare continuous phases with the concentration of 2% (w/w). The oil phase was commercially available soybean oil. Mixing and homogenizing the continuous phase and the oil phase at 20000rpm for 3min by using a high-speed homogenizer to obtain a coarse emulsion (the oil phase accounts for 5 wt% of the mixture after mixing the continuous phase and the oil phase); the resulting crude emulsion was then homogenized twice at 75MPa using Microfluidics International corporation, Newton, MA to give the final emulsion, which was photographed after 12h standing and the particle size distribution and average particle size of the emulsion were measured using a laser particle size analyzer during fresh or storage.
(2) And (4) analyzing results:
the formed emulsion is shown in figure 1, obvious layering phenomenon occurs rapidly after the preparation of the MSCP stable emulsion is finished, and the modified beautiful millettia root polysaccharide AC-MSCP1, AC-MSCP2 and AC-MSCP3 stable emulsion is not layered, which shows that the layering stability of the emulsion can be effectively improved through hydrophobic modification.
The particle size distribution and the average particle size are shown in figures 2 and 3, the MSCP stable emulsion has a wide range of droplet particle size distribution (0.5-100 um), and the average particle size is 15.51 +/-1.09 um. In contrast, the droplet size distribution of the modified Millettia speciosa polysaccharides AC-MSCP1, AC-MSCP2, AC-MSCP3 stabilized emulsions shifted to a lower particle size range, where the average droplet size of the AC-MSCP1, AC-MSCP2 and AC-MSCP3 stabilized emulsions were 1.57. + -. 0.02. mu.m, 0.24. + -. 0.01. mu.m and 0.31. + -. 0.01. mu.m, respectively. The particle size of the modified beautiful millettia root polysaccharide stable emulsion is smaller than that of the beautiful millettia root polysaccharide stable emulsion, which shows that the hydrophobic modification can effectively improve the emulsifying capacity of the beautiful millettia root polysaccharide. Furthermore, the average droplet size of the AC-MSCP2 and AC-MSCP3 stabilized emulsions was smaller than that of the GA stabilized emulsions (0.71 ± 0.03um), indicating that the emulsifying properties of AC-MSCP2 and AC-MSCP3 are superior to those of commercial polysaccharide emulsifiers.
The change in the average particle size of the emulsions prepared with beautiful millettia root polysaccharide, modified beautiful millettia root polysaccharide and gum arabic GA during storage is shown in fig. 4, and the average particle size of the MSCP-stabilized emulsion after storage for 30 days is significantly increased from 15.51 ± 1.09um to 30.30 ± 2.39um, which is nearly 2-fold greater than the initial particle size. In contrast, the average droplet sizes of the AC-MSCP1, AC-MSCP2, and AC-MSCP3 stabilized emulsions gradually increased from 1.57. + -. 0.02. mu.m, 0.24. + -. 0.01. mu.m, and 0.31. + -. 0.01. mu.m, to 1.82. + -. 0.09. mu.m, 1.10. + -. 0.01. mu.m, and 0.50. + -. 0.08. mu.m, respectively. The result shows that the hydrophobic modification can effectively improve the emulsification stability of the beautiful millettia root polysaccharide.
Example 8: determination of antioxidant Activity
(1) Determination of DPPH radical scavenging Capacity: 150uL of bovine Millettia speciosa polysaccharide MSCP-1 (MSCP for short), and modified bovine Millettia speciosa polysaccharide AC-MSCP1, AC-MSCP2 and AC-MSCP3 solutions (0.25-2mg/mL) are mixed with 50uL of 1, 1-diphenyl-2-trinitrophenylhydrazine (DPPH) solution (1mM), and the reaction is kept in the dark for 30 min. The absorbance at 517nm was recorded using a microplate reader. The background group was composed of an equal amount of absolute ethanol instead of DPPH solution, and the blank group was composed of an equal amount of distilled water instead of sample solution.
(2) Determination of hydroxyl radical scavenging Capacity: accordingly, 1ml salicylic acid (9mM) and 1ml distilled water were added to 1ml ferrous sulfate (9 mM). Then, 1mL of a sample solution to be tested (0.25-2mg/mL) and 1mL of H were added to the mixture2O2(8.8 mM). After incubation at 37 ℃ for 30min, the absorbance at 510nm was measured. The blank control group was prepared by replacing the sample solution with equal amount of distilled water, and replacing the sample solution with equal amount of distilled water2O2As a background group. Clearance (scraping activity) was calculated according to the following equation:
clearance rate ═ 1- (A)1-A2)/A0)×100；
In the formula, a0, a1 and a2 represent a blank control group, a sample group and a background group, respectively.
(3) And (4) analyzing results: as shown in fig. 5a, DPPH radical clearance for all samples gradually increased with increasing concentration. At a concentration of 2mg/mL, the DPPH radical clearance rates for MSCP, AC-MSCP1, AC-MSCP2 and AC-MSCP3 were 93.57% + -3.03, 76.25% + -1.25, 84.17% + -1.91 and 86.25% + -3.54, respectively. Compared with beautiful millettia root polysaccharide, the acetylated beautiful millettia root polysaccharide has reduced DPPH free radical scavenging activity. Similarly, at any concentration, the scavenging rate of hydroxyl radicals of the beautiful millettia root polysaccharide is obviously higher than that of the modified beautiful millettia root polysaccharide (figure 5b), which indicates that the modification method adopted has certain damage to the antioxidant activity of the beautiful millettia root polysaccharide.
Example 9: measurement of oxidation products of fats and oils
(1) The contents of hydroperoxide and malondialdehyde in the emulsions prepared in example 7 (i.e. the emulsions stabilized with millettia speciosa MSCP-1 (MSCP for short), and the emulsions stabilized with modified millettia speciosa AC-MSCP1, AC-MSCP2 and AC-MSCP 3) were determined, using hydroperoxide and malondialdehyde in bulk oil (commercially available soybean oil-goldfish brand) as controls, wherein the determination was as follows:
measurement of hydroperoxide: to 1.5mL of mixture a (isooctane: 2-propanol ═ 3:1, v/v) was added 0.5mL of the emulsion sample, which was then vortexed 3 times (20 s each). After centrifugation at 3000g for 5min, 0.2mL of supernatant was collected and 2.8mL of mixture B was added (methanol: 1-butanol ═ 2:1, v/v). Then, 15uL of ammonium thiocyanate solution (3.94M) and 15uL of ferrous ions (ferrous ions were obtained by mixing equal volumes of 0.288M ferrous sulfate solution and 0.264M barium chloride solution and centrifuging at 5000r/mim for 3min, supernatant). The resulting mixture was incubated at room temperature for 20 minutes and then the absorbance was measured at 510 nm. Hydrogen peroxide solutions (15.625, 31.25, 62.5, 125umol/L) were prepared as standards and measured according to the same method as above, and a standard curve was plotted, the regression equation being: y is 0.0105x +0.0366 (R)20.998, y denotes absorbance value and x denotes hydrogen peroxide concentration).
Measurement of malondialdehyde: 0.2mL of the sample emulsion was mixed with 1.8mL of distilled water and 4mL of thiobarbituric acid (TBA) (0.375%, w/v), and then the mixture was put into boiling water for 15min, centrifuged at 10000rpm for 5min, and then the supernatant was taken and absorbance was measured at 532 nm. Preparing malonaldehyde diacetal with a series of concentrations (0.1, 0.2, 0.4, 0.6, 0.8, 1ug/mL) as a standard substance, measuring according to the same method, drawing a standard curve, and obtaining a regression equation: y 0.4106x +0.0552 (R)20.999, y denotes absorbance value and x denotes malondialdehyde concentration).
(2) And (4) analyzing results: as shown in FIG. 6a, the hydroperoxide concentration of the Bulk oil (Bulk oil) increased significantly from 0.0575. + -. 0.004mmol/kg of oil to 0.366. + -. 0.038mmol/kg of oil after 30d of storage. However, the hydroperoxide (LH) concentration in the stable emulsion of beautiful millettia root polysaccharide or modified beautiful millettia root polysaccharide increases only slightly during storage. Furthermore, no significant difference was found in the hydroperoxide concentration in the emulsions prepared with beautiful millettia root polysaccharide MSCP or modified beautiful millettia root polysaccharides AC-MSCP1, AC-MSCP2, AC-MSCP3 after 30d storage (p < 0.05). Similarly, the stable emulsions of Millettia speciosa MSCP or modified Millettia speciosa AC-MSCP1, AC-MSCP2 and AC-MSCP3 have a malondialdehyde content significantly lower than that of bulk oil after storage for 30 days (FIG. 6 b). The results show that the beautiful millettia root polysaccharide MSCP or the modified beautiful millettia root polysaccharide AC-MSCP1, AC-MSCP2 and AC-MSCP3 can effectively delay the oxidation rate of lipid. In addition, although acetylation causes the oxidation resistance of the antioxidant to be reduced (fig. 5), the emulsion prepared from the modified beautiful millettia root polysaccharide still maintains the same lipid oxidation rate as the emulsion prepared from the beautiful millettia root polysaccharide, because more modified polysaccharides are adsorbed to an oil-water interface after modification, a denser and viscoelastic interface layer is formed, and the interface layer can play a physical shielding role for some oxidation promoters, so that the oxidation rate of the lipid can be effectively delayed.
Example 10: determination of rheological Properties
The interfacial rheological behaviour of the fresh emulsion was determined at 25 ℃ using a rheometer MCR 102(Anton Paar, Austria). The diameter of the parallel plate is 40mm, and the gap is 1 mm. Samples, namely, the millettia speciosa MSCP-1 (MSCP for short) stabilized emulsion prepared as in example 7, and the modified millettia speciosa AC-MSCP1, AC-MSCP2 and AC-MSCP3 stabilized emulsion, were placed on a rheometer platform and dynamic frequency sweeps were recorded recording the change in elastic (G') and viscous (G ") moduli over the frequency range 0.1 to 10rad/s, all measurements being taken in the linear viscoelastic region (stress fixed at 0.5%).
And (4) analyzing results: as shown in fig. 7, the elastic modulus (G') was higher than the viscous modulus (G ") in all the emulsions over the entire frequency range, indicating that the prepared emulsions exhibited a distinct elastic behavior. Meanwhile, due to the interaction between the hydrophobic acetyl groups in the modified beautiful millettia root polysaccharide chain and oil drops, G 'and G' of the emulsion prepared by the modified beautiful millettia root polysaccharide are higher than those of the unmodified beautiful millettia root polysaccharide. Furthermore, as the DS value (degree of substitution) increased, G' and G "increased significantly (i.e., as the degree of substitution increased, the viscoelasticity of the AC-MSCP1, AC-MSCP2, and AC-MSCP3 stabilized emulsions also increased gradually), indicating that an increase in the degree of acetylation helped to form a more elastic and viscous interface. These results indicate that acetylation modification has a positive effect on the rheological behaviour of the interfacial layer formed by the modified beautiful millettia root polysaccharide. That is, the grafted acetyl groups help to build an interface layer with strong viscoelasticity. The interface layer with strong viscoelasticity can play a physical shielding role on the oxidation promoting agent, and further can delay the oxidation of grease.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (10)
1. The application of beautiful millettia root polysaccharide and/or modified polysaccharide thereof in delaying grease oxidation is characterized in that: the beautiful millettia root modified polysaccharide is acetylated beautiful millettia root modified polysaccharide.
2. The use of claim 1, wherein the beautiful millettia root modified polysaccharide is prepared by the following method:
dissolving the beautiful millettia root polysaccharide in water, adjusting the pH value to 8.0-9.0, then dropwise adding acetic anhydride, dropwise adding an alkaline solution to maintain the pH value of the solution at 8.0-8.5, adjusting the pH value to 7.0 after the reaction is finished, dialyzing, and freeze-drying to obtain the modified beautiful millettia root polysaccharide.
3. Use according to claim 2, characterized in that:
the dosage of the acetic anhydride is calculated according to 60-300 mg of beautiful millettia root polysaccharide per ml of acetic anhydride.
4. Use according to claim 2, characterized in that:
and the pH value is adjusted to 8.0-8.5.
5. Use according to claim 2, characterized in that:
the alkaline solution is NaOH solution;
the pH value is adjusted to 7.0 by adopting HCl solution.
6. Use according to claim 2, characterized in that:
the reaction time is 3-5 h;
the dialysis is carried out by adopting a dialysis bag with the molecular weight cutoff of 3500 Da.
7. Use according to claim 1, characterized in that: the beautiful millettia root polysaccharide and/or the beautiful millettia root modified polysaccharide are/is added into the grease and evenly mixed to delay the oxidation of the grease.
8. Use according to claim 7, characterized in that: the oil is edible oil or industrial oil.
9. Use of the beautiful millettia root polysaccharide and/or the modified polysaccharide thereof described in claim 1 for the preparation of an emulsifier.
10. Use of the beautiful millettia root polysaccharide and/or the modified polysaccharide thereof described in claim 1 in an oil-in-water emulsion.
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| ZHI HUANG等: "Effect of acetylation modification on the emulsifying and antioxidant properties of polysaccharide from Millettia speciosa Champ", 《FOOD HYDROCOLLOIDS》 * |
| 李俊俊: "结构修饰对沙蒿多糖乳化性的影响", 《中国优秀博硕士学位论文全文数据库（硕士）工程科技Ⅰ辑》 * |
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