阅读说明：本技术 一种pH响应缓释的大豆蛋白纤维亚铁复合物的制备方法及其复合物 (Preparation method of pH response slow-release soybean protein fiber ferrous complex and complex thereof ) 是由 杨平 于 2021-07-19 设计创作，主要内容包括：本发明公开了一种pH响应缓释的大豆蛋白纤维亚铁复合物的制备方法及其复合物,通过大豆蛋白纤维溶液、亚铁溶液以及水溶性抗坏血酸进行制备,其中,大豆纤维形成的空间网络结构,对亚铁具有良好的黏附性,减少无机铁强化剂中亚铁易被氧化的问题,使亚铁缓慢释放出来,同时蛋白纤维隔离保护亚铁营养素,降低亚铁对胃部的刺激,作为载体用于亚铁营养素缓释,实现最优输送,提高其生物利用率,由于该制备方法中的材料及过程均未使用有毒有害物质,未引入有毒有害的化学物质,这对于亚铁大豆蛋白复合物后续的推广应用,安全可靠性强。所述复合物,采用上述方法制备获得,具有pH敏感性,而且具有稳定性好,释放速度缓慢以及高生物利用率等优点。(The invention discloses a preparation method of a pH response slow-release soybean protein fiber ferrous complex and the complex, wherein the preparation method is carried out by a soybean protein fiber solution, a ferrous solution and water-soluble ascorbic acid, wherein a spatial network structure formed by soybean fibers has good adhesion to ferrous, the problem that ferrous in an inorganic iron reinforcer is easy to oxidize is solved, the ferrous is slowly released, meanwhile, the protein fibers isolate and protect ferrous nutrient, the stimulation of ferrous to the stomach is reduced, the protein fibers are used as a carrier for the slow release of the ferrous nutrient, the optimal delivery is realized, the bioavailability is improved, toxic and harmful chemical substances are not introduced due to the fact that toxic and harmful substances are not used in the materials and the process in the preparation method, and the subsequent popularization and application of the ferrous soybean protein complex are high in safety and reliability. The compound prepared by the method has pH sensitivity, and has the advantages of good stability, slow release speed, high bioavailability and the like.)

1. A preparation method of a pH response slow-release soybean protein fiber ferrous complex is characterized by comprising the following steps:

1) preparing a soybean protein fiber solution with the mass concentration of 1-2% for later use;

2) preparing a ferrous solution with the concentration of 0.5-1 mol/L for later use;

3) and adding a spare ferrous solution into the soybean protein fiber solution, stirring for 2-4 hours at room temperature by using a magnetic stirrer under the conditions of light protection and nitrogen protection, and freeze-drying to obtain the pH response slow-release soybean protein fiber ferrous compound.

2. The method for preparing the ferrous soy protein fiber complex with slow release of pH response according to claim 1, wherein the soy protein fiber solution prepared in step 1) is specifically:

1.1) dissolving the isolated soy protein in distilled water, stirring at room temperature, and then hydrating overnight under the refrigeration condition of 0-4 ℃;

1.2) adjusting the pH value of the soybean protein isolate solution after the overnight hydration to 1.5-2.4, then carrying out the overnight hydration under the refrigeration condition of 0-4 ℃, centrifuging, and taking the supernatant;

1.3) diluting the supernatant by using distilled water until the mass fraction is 1.5-2%, dialyzing to remove salt, and filtering by using a filter membrane to obtain filtered supernatant;

and 1.4) placing the supernatant into a small bottle, sealing, heating in a water bath for 20-48 h at the temperature of 60-80 ℃, and cooling to obtain the soybean protein fiber solution.

3. The method for preparing the pH-responsive slow-release ferrous soy protein fiber complex as claimed in claim 2, wherein in step 1.1), the mass ratio of the soy protein isolate to the distilled water is 1: 9.

4. The method for preparing the pH-response slow-release ferrous soy protein fiber complex as claimed in claim 2, wherein in step 1.2), the pH value of the solution of the soy protein isolate after being hydrated overnight is adjusted to 1.5-2.4 by adding dilute hydrochloric acid or dilute acetic acid.

5. The method for preparing the pH-responsive slow-release ferrous soy protein fiber complex as claimed in claim 2, wherein in step 1.2), the centrifugation speed is 15000rpm/min and the centrifugation time is 15 min.

6. The method for preparing the pH-responsive slow-release ferrous soy protein fiber complex as claimed in claim 2, wherein in step 1.3), the dialysis desalination is specifically: dialyzing for 2-4 h by adopting a 6-8 kDa dialysis bag.

7. The method for preparing the pH-responsive slow-release ferrous soy protein fiber complex as claimed in claim 2, wherein in step 2), the ferrous solution is prepared by: weighing ferrous sulfate or ferrous chloride, grinding, dissolving in distilled water, preparing into a solution with the concentration of 0.5-1 mol/L, and carrying out ultrasonic treatment for 15-20 min at 200-300W in the dark for later use.

8. The method for preparing the pH-response slow-release ferrous soy protein fiber compound as claimed in claim 1, wherein in the step 3), the mass ratio of the soy protein fiber solution to the ferrous solution is 1: 3-7.

9. The method for preparing the pH-response slow-release ferrous soy protein fiber complex as claimed in claim 1, wherein in step 3), the ferrous solution is added to the soy protein fiber solution and at the same time, water-soluble ascorbic acid with a mass concentration of 1-2% is added.

10. A pH response slow-release soybean protein fiber ferrous complex, which is characterized by being prepared by any one preparation method of claims 1-8.

Technical Field

The invention relates to the technical field of iron-fortified nutritional foods, in particular to a preparation method of a pH response slow-release soybean protein fiber ferrous complex and the complex thereof.

Background

Iron, the trace element with the highest content in human body, participates in oxygen transportation and storage, maintains hematopoietic function, enhances immune function, and participates in metabolism. The iron in food exists mainly in the form of trivalent iron, and is not easy to be absorbed by human body. Iron deficiency nutrition is still a global health concern, iron deficiency is expressed by the following factors, namely women, pregnant women and children as high-incidence people: 1. anemia; 2. influence on growth and development of children: the children often have physical development retardation and a reduced health level due to iron deficiency; 3. influence on development of motor ability: the children with iron deficiency have weak muscles, reduced motor ability and easy fatigue; 4. affecting immune function, and increasing susceptibility of children suffering from iron deficiency.

The iron-enriched food is an effective measure for preventing iron deficiency and iron-deficiency anemia in high risk groups. Among the nutrient supplements, inorganic iron supplements, organic iron supplements, and nano-iron supplements are commonly used.

The inorganic iron enhancer is low in cost and easy to obtain, but is extremely easy to oxidize due to the strong rust taste and ferrous ions, has gastrointestinal irritation and low absorptivity, has obvious influence on the sensory characteristics of food, is easily highly unstable due to the action of chemical substances, light, oxygen, heat and the like in the food processing process and the storage process, and limits the wide popularization of the inorganic iron enhancer in fortified food.

The organic iron enhancer has high bioavailability, but is unstable in the environment and is easy to dissociate into inorganic iron salts, so that the bioavailability is reduced. In addition, the research shows that the iron-supplementing agent does not show obvious advantages compared with an inorganic iron-supplementing agent, when the iron-deficiency anemia of a patient with gastrotomy is treated, due to the strong acid environment of the stomach, the chelated mineral substances are completely or partially dissociated into inorganic substances in the gastrointestinal tract, the absorption capacity of the body to the chelated mineral substances is reduced, for example, glycine chelated iron has no better effect than ferrous sulfate, and the bioavailability of the glycine chelated iron is not improved.

The nano iron enhancer is prepared by taking iron as a core material and taking a natural or synthetic high molecular substance as a wall material to prepare a microcapsule by means of physical, chemical and biological means, and wrapping ferrous iron, so that the effects of increasing the stability and slowly releasing the iron are achieved. However, the nano capsule of the nano iron enhancer is complex to manufacture, and the diffusion rate of the core material is influenced by the thickness of the wall material. The nano ferrous iron with the thin wall material is rapidly disintegrated and has high diffusion rate, the phenomenon of 'peak to valley' of iron concentration caused by the sudden release of the ferrous iron is easily caused, and the characteristics of slow release, enteric solubility and the like are poor; the nano ferrous iron of the multilayer wall material is difficult to control the releasable limited thickness, and a large part of the wall material does not belong to the range of food additives. Moreover, the microcapsules adopt a high-temperature ring section in a spray drying process flow to accelerate the oxidation speed of ferrous ions and reduce the nutritional value of the microcapsules.

Therefore, how to develop a novel sustained-release ferrous complex for application in iron-fortified food becomes a problem to be solved urgently.

Disclosure of Invention

In view of the above, the invention provides a preparation method of a pH response slow-release soybean protein fiber ferrous complex and a complex thereof, so as to solve the problems of poor stability, difficult absorption, high release speed and the like of the conventional iron fortifier, and the conventional iron fortifier cannot be well applied to iron fortified food.

In one aspect, the invention provides a preparation method of a pH response slow-release soybean protein fiber ferrous complex, which comprises the following steps:

1) preparing a soybean protein fiber solution with the mass concentration of 1-2% for later use;

2) preparing a ferrous solution with the concentration of 0.5-1 mol/L for later use;

3) and adding a spare ferrous solution into the soybean protein fiber solution, stirring for 2-4 hours at room temperature by using a magnetic stirrer under the conditions of light protection and nitrogen protection, and freeze-drying to obtain the pH response slow-release soybean protein fiber ferrous compound.

Preferably, the preparation of the soy protein fiber solution in step 1) specifically comprises:

1.1) dissolving the isolated soy protein in distilled water, stirring at room temperature, and then hydrating overnight under the refrigeration condition of 0-4 ℃;

1.2) adjusting the pH value of the soybean protein isolate solution after the overnight hydration to 1.5-2.4, then carrying out the overnight hydration under the refrigeration condition of 0-4 ℃, centrifuging, and taking the supernatant;

1.3) diluting the supernatant by using distilled water until the mass fraction is 1.5-2%, dialyzing to remove salt, and filtering by using a filter membrane to obtain filtered supernatant;

and 1.4) placing the supernatant into a small bottle, sealing, heating in a water bath for 20-48 h at the temperature of 60-80 ℃, and cooling to obtain the soybean protein fiber solution.

Further preferably, in the step 1.1), the mass ratio of the soy protein isolate to the distilled water is 1: 9.

Preferably, in the step 1.2), the pH of the soy protein isolate solution after the overnight hydration is adjusted to 1.5-2.4 by adding dilute hydrochloric acid or dilute acetic acid.

Further preferably, in step 1.2), the speed of centrifugation is 15000rpm/min and the centrifugation time is 15 min.

Further preferably, in step 1.3), the dialysis desalination specifically comprises: dialyzing for 2-4 h by adopting a 6-8 kDa dialysis bag.

Further preferably, in step 2), the preparing a ferrous solution specifically includes: weighing ferrous sulfate or ferrous chloride, grinding, dissolving in distilled water, preparing into a solution with the concentration of 1mol/L, and carrying out ultrasonic treatment for 15-20 min at 200-300W in a dark place for later use.

Further preferably, in the step 3), the mass ratio of the soybean protein fiber solution to the ferrous solution is 1: 3-7.

Preferably, in the step 3), the spare ferrous solution is added into the soybean protein fiber solution, and simultaneously, water-soluble ascorbic acid with the mass concentration of 1-2% is added.

On the other hand, the invention also provides a pH response slow-release soybean protein fiber ferrous complex, which is prepared by any one preparation method of claims 1-8.

The preparation method of the soybean protein fiber ferrous complex with pH response slow release provided by the invention is characterized in that the soybean protein fiber solution, the ferrous solution and the water-soluble ascorbic acid are used for preparing the soybean protein fiber ferrous complex, wherein a spatial network structure formed by soybean fibers has good adhesion to ferrous, the problem that ferrous in an inorganic iron enhancer is easily oxidized is solved, the ferrous is slowly released, the phenomenon that peaks and troughs appear due to the sudden release of the ferrous is avoided, meanwhile, the protein fibers isolate and protect the ferrous nutrient, the stimulation of the ferrous to the stomach is reduced, the protein fiber ferrous complex is used for slow release of the ferrous nutrient as a carrier, and the protein fiber ferrous complex is stable in the stomach, realizes the optimal delivery and improves the bioavailability of the protein fiber ferrous complex. As no toxic and harmful substance is used in the material and the process of the preparation method, and no toxic and harmful chemical substance is introduced, the preparation method is safer and more reliable for subsequent popularization and application of the ferrous soy protein compound, especially for infant iron fortified food.

The pH response slow-release soybean protein fiber ferrous complex provided by the invention is prepared by the method, and has the advantages of single substance of a preparation system, pH sensitivity, good stability, slow release speed, high bioavailability and the like.

It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the disclosure.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the invention and together with the description, serve to explain the principles of the invention.

In order to more clearly illustrate the embodiments or technical solutions in the prior art of the present invention, the drawings used in the description of the embodiments or prior art will be briefly described below, and it is obvious for those skilled in the art that other drawings can be obtained based on these drawings without creative efforts.

FIG. 1 is an electron microscope picture of a ferrous complex in a soybean protein fiber;

FIG. 2 is a photograph comparing ferrous complexes of soy protein fibers with pH;

FIG. 3 is a bar graph of ferrous retention of a ferrous complex of soy protein fiber as a function of pH;

FIG. 4 is a bar graph of ferrous retention of ferrous complexes of soy protein fiber in gastric fluid;

FIG. 5 is a bar graph of the ferrous retention rate of a ferrous soy protein fiber complex in intestinal fluid;

FIG. 6 is a photograph comparing thermal stability of a ferrous soy protein fiber complex in an environment of 4-90 deg.C;

FIG. 7 is a histogram of the ferrous retention rate of the soy protein fiber ferrous complex in an environment of 4-90 ℃.

Detailed Description

The present invention is further illustrated by the following examples, which are not intended to limit the scope of the invention.

In order to provide a nutrition enhancer suitable for being applied to iron-fortified food, considering that protein fiber is degradable in a living body and active substances can be slowly released from the interior of gel, the embodiment tries to use soybean protein fiber as a carrier of ferrous ions to achieve the purposes of slow release and the like, and provides a preparation method of a pH response slow release soybean protein fiber ferrous complex, which comprises the following specific preparation steps:

1) preparing a soybean protein fiber solution: when the pH value in the environment is far away from the isoelectric point and the ionic strength is low, the charge quantity on the surfaces of protein molecules is large, and strong electrostatic repulsion is generated among the molecules, so that the electrostatic repulsion is balanced with the hydrophobic interaction among the molecules, and the hydrophilic gel with a certain network structure is formed. Dissolving the isolated soy protein in distilled water, wherein the mass ratio of the isolated soy protein to the distilled water is 1: and 9, stirring at room temperature for 0.5-1 h, and then hydrating at 4 ℃ under a refrigeration condition overnight. The pH of the solution was adjusted to 2.0 using dilute hydrochloric or acetic acid solution and hydrated overnight at 4 ℃. The protein solution was centrifuged at 15000rpm/min for 15min and the supernatant was taken. Diluting the protein supernatant with distilled water (with the preset pH value of 2.0) to the mass concentration of 1.5-2% for later use, dialyzing to remove salt (dialyzing for 2-4 h by a 6-8 kDa dialysis bag), and filtering by a 0.45 micrometer filter membrane. Putting the protein solution into a small bottle with a sealing cover, heating in water bath at 60-80 ℃ for 20-48 h, immediately cooling the heated protein solution to 4 ℃ to stop fibrosis, and storing at 4 ℃ for later use.

2) Preparing a ferrous solution: accurately weighing ferrous sulfate or ferrous chloride solid, grinding the ferrous sulfate or ferrous chloride solid, dissolving the ground ferrous sulfate or ferrous chloride solid in distilled water to prepare a solution with the concentration of 1mol/L, and carrying out ultrasonic treatment for 15-20 min at 200-300W in a dark place

3) Preparing a soybean protein fiber ferrous complex: adjusting the mass concentration of a protein solution of the protein fiber to be 1-2%, adding ferrous sulfate or ferrous chloride according to the mass ratio of 1: 3-7, then adding a proper amount of water-soluble ascorbic acid with the mass concentration of 0.01-0.1%, stirring for 2-4 h at room temperature by using a magnetic stirrer under the conditions of light shielding and nitrogen protection, and freeze-drying to obtain the pH response slow-release soybean protein fiber ferrous complex.

The water-soluble ascorbic acid is added mainly for preventing ferrous ions in ferrous sulfate or ferrous chloride from being oxidized, and is not used as a core component, and the specific addition amount can be determined according to circumstances as long as the content in the final product does not exceed the national standard requirement.

Fig. 1 is an electron microscope picture of the ferrous complex in the soybean protein fiber.

Example 1

PH responsiveness detection of soy protein fiber ferrous complex

Adjusting the pH value of the soybean protein fiber ferrous complex prepared in the embodiment to 1.5, 2, 3, 4, 5, 6, 7, 8 and 9 by using 0.1mol/L NaOH under the condition of keeping out of the sun, reacting for 30min at room temperature, measuring the ferrous content in the sample by using an o-phenanthroline colorimetric method, and calculating the ferrous retention rate.

Wherein the content of the first and second substances,

referring to fig. 2 and fig. 3, as the pH value of the protein fiber ferrous iron complex increases, the retention rate of ferrous iron gradually decreases, the retention rate of ferrous iron at pH 1.5 is 92.99%, the retention rate decreases to 70.55% at pH 4, the retention rate decreases to 17.32% at pH 7, and the retention rate decreases to 10.09% at pH 9.

Example 2

In vitro digestive stability of soybean protein fiber ferrous complex

(1) Simulating stability in gastric fluid. Preparing gastric juice (preparing dilute HCl solution by using 100mL of distilled water and 23.4mL of concentrated hydrochloric acid, adding 80mL of distilled water into 1.64mL of dilute hydrochloric acid, uniformly mixing, weighing 1g of pepsin, adding distilled water, and fixing the volume in a 100mL volumetric flask, wherein the pH value in the solution is about 2.0). Adding the soybean protein fiber-ferrous complex into the simulated gastric juice to ensure that the concentration of ferrous in the solution is 1mg/mL, digesting for 2h (37 ℃, magnetically stirring), taking the dialysate every 0.5h, and calculating the retention rate of ferrous in the complex, wherein the specific result is shown in figure 4.

(2) Simulating stability in intestinal fluid. Preparing intestinal juice (accurately weighing 0.68g potassium dihydrogen phosphate dissolved in 25mL water, adding 0.2mol/L sodium hydroxide solution 7.7mL and water 50mL, adding trypsin 1.0g, stirring to dissolve completely, and adding 0.2mol/L NaHCO₃Adjust the pH of the sample to 5.0 and hold the volume in a 100mL volumetric flask. ) Adding 1mL of simulated intestinal fluid into the soybean protein fiber ferrous complex digested in simulated gastric juice for 2h, digesting for 8h under the conditions of magnetic stirring and light protection at 37 ℃, and taking dialysate every 1 h. Transferring the solution after digestion into a dialysis bag, dialyzing at 37 ℃ for 3h, and calculating the retention rate of ferrous iron in the compound, wherein the specific result is shown in figure 5.

When simulated gastric fluid digestion is carried out, the retention rate of the ferrous iron is 92.35% in 0.5h and 81.36% in 2h, and the retention rate of the ferrous iron in the compound does not decrease remarkably along with the extension of digestion time; when intestinal juice digestion is simulated, the retention rate of the compound ferrous iron is 82.35% in 1h, 74.72% in 2h, 69.21% in 3h, 51.36% in 4h, 43.96% in 5h, 35.84% in 6h, 28.45% in 7h and 15.51% in 8h, and the retention rate of the ferrous iron gradually decreases along with the increase of the intestinal juice digestion time. As can be seen, the protein fiber ferrous complex is relatively stable in gastric juice, and the protein is slowly degraded in intestinal juice to release ferrous ions.

Example 3

Thermal stability of soybean protein fiber ferrous complex

The heat stability of the soybean protein fiber ferrous iron compound is detected within the temperature range of 4-90 ℃, the retention rate of ferrous iron in the compound does not change significantly (p is greater than 0.05) along with the rise of the temperature, and the retention rates of ferrous iron within the temperature range of 4-90 ℃ are all above 83%, and the method is shown in fig. 6 and fig. 7.

Other embodiments of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. This application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the invention being indicated by the following claims.

It is to be understood that the present invention is not limited to what has been described above, and that various modifications and changes may be made without departing from the scope thereof. The scope of the invention is limited only by the appended claims.