CN116813943B

CN116813943B - A method for preparing starch nanospheres with uniform size

Info

Publication number: CN116813943B
Application number: CN202310317987.9A
Authority: CN
Inventors: 徐恩波; 姚思羽; 朱青青; 程焕; 陈健乐; 刘东红
Original assignee: Zhejiang University ZJU
Current assignee: Zhejiang University ZJU
Priority date: 2023-03-29
Filing date: 2023-03-29
Publication date: 2025-01-03
Anticipated expiration: 2043-03-29
Also published as: CN116813943A

Abstract

The present invention relates to a method for preparing starch nanospheres of uniform size, comprising the following steps: (1) gelatinizing a starch suspension and subjecting it to microfluidization to form a starch cluster solution, wherein the starch suspension is water or a phosphate buffer solution of pH 2-8, and the starch content in the starch suspension is 1-20wt%; (2) subjecting the starch cluster solution to alcohol precipitation, removing soluble molecules by centrifugation, taking the precipitate for freeze-drying, grinding and sieving, and obtaining starch nanospheres; wherein the alcohol precipitation treatment comprises: dripping anhydrous ethanol into the starch cluster solution at a uniform speed of 5-30 mL/min, and stirring is maintained during the alcohol precipitation process, the stirring speed is 200-1500 r/min, and the volume ratio of anhydrous ethanol to the starch cluster solution is 0.5:1-10:1. The present invention utilizes microfluidization to induce activation of starch hydroxyl groups, and promotes starch chain crosslinking by alcohol precipitation treatment, so as to prepare starch nanospheres with smaller size, better uniformity and better dispersibility.

Description

Preparation method of starch nanospheres with uniform size

Technical Field

The invention relates to the technical field of starch nanospheres, in particular to a method for preparing starch nanospheres with uniform size.

Background

Nano-starches are of interest because of their high biocompatibility, low sensitization, small size and large specific surface area. In recent years, nano starch has been prepared in various forms such as spheres, rods, polyhedrons, flakes, etc. to satisfy the fields of medicine, cosmetics, foods, etc. Wherein the starch nanospheres are uniformly stressed and have the most stable structure, which is beneficial to enhancing the membrane performance and delivering the drug application. However, since starch is a polymer, the multi-stage structure is complex (from basic glucose units, starch short chains, starch long straight chains/branched chains and various space structures formed by the starch long straight chains/branched chains, such as starch clusters and starch clusters, to growth rings), the staggered rigid structure (starch crystallization area) and flexible structure (starch amorphous area) of the starch sugar chains exist, the starch nanosphere structure is difficult to be accurately regulated and designed, and the problems of different sizes, changeable forms, uneven surfaces, easy aggregation and the like of the nanoscale starch spheres exist.

There are various methods for preparing nano starch, such as enzymolysis, acidolysis, nano precipitation, etc. The enzymolysis method and the acidolysis method are based on a top-down strategy to prepare the nano-sized starch, and have the defects that the shape of the degradation of the original micro-sized starch particles is difficult to control, and the particle size distribution of the product is wider. The nano precipitation method is a green, efficient and controllable method, and ethanol is used as an antisolvent to prepare the starch nanospheres. However, nano-sized starch prepared by ethanol sedimentation is more physically screened by a similar compatibility principle, so that the uniformity of the product is poor, the yield is low, and meanwhile, the problem of easy aggregation of the product limits the advantages of large specific surface area and nano-sized effect. Based on this, many studies have been made to enhance repulsive force between nano-particles by adding chemical surfactants to assist in preparing and modifying nano-starch, thereby improving retention of the balling morphology of nano-starch and dispersibility in a liquid system. However, after the chemical surfactant is blended into the system, the chemical surfactant is not easy to remove in the nano starch product, and the use safety is not enough.

Thus, there is a need for a method of preparing starch nanospheres of uniform size.

Disclosure of Invention

The invention provides a preparation method of starch nanospheres with uniform size, which utilizes the mechanical force of microjet to activate the hydroxyl groups of starch chains to form small molecular starch clusters, and the starch nanospheres with uniform size are formed by crosslinking under the action of alcohol precipitation.

In order to realize the above summary, the technical scheme of the application is as follows, a preparation method of starch nanospheres with uniform size comprises the following steps:

(1) Dispersing starch in water or phosphate buffer solution with pH of 2-8 to obtain starch suspension with mass fraction of 1-20wt%;

(2) Carrying out alcohol precipitation treatment on the starch cluster solution, removing soluble molecules through centrifugation, taking precipitate, carrying out freeze-drying treatment, grinding and sieving to obtain starch nanospheres;

The alcohol precipitation treatment is that absolute ethyl alcohol is dripped into the starch cluster solution at a constant speed of 5-30mL/min, stirring is kept in the alcohol precipitation process, the stirring speed is 200-1500r/min, and the volume ratio of the absolute ethyl alcohol to the starch cluster solution is 0.5:1-10:1.

Microfluidics is an emerging physical method, and relates to mechanical actions such as high-pressure effect, high-speed impact, high-frequency vibration, instantaneous pressure drop, strong shearing and cavitation effects, and the like, which is green and efficient compared with a chemical method. So far, research into microfluidics has focused mainly on structural and functional modification of starch granules. The invention creatively applies the microjet to the preparation of the ultra-uniform starch nanospheres, after the fully gelatinized starch solution is injected into a microjet pipeline, the microjet is divided into a plurality of liquid drops, and the micro-molecular starch clusters formed by the intense collision of high-speed impact, strong shearing, instantaneous pressure drop, cavitation effect and other comprehensive acting forces in a microjet reactor are regulated and controlled by the strength of a designed force field, the hydroxyl groups are activated by intense collision induction, and then the uniform starch nanosphere particles are obtained by alcohol precipitation treatment. In the process of adding ethanol, water and ethanol molecules form interface reactions such as turbulence and the like to promote starch chains to further collide and crosslink to form new bonds, so that more uniform starch nanospheres are formed, the starch nanospheres are separated out in the ethanol solution, and therefore, soluble small molecules are removed, and the separation and purification effects are achieved.

Further, the starch of the starch suspension is one or more of rice starch, corn starch, wheat starch, sweet potato starch, tapioca starch, sweet potato starch and potato starch.

Further, the micro-jet treatment adopts a micro-jet reactor, and the micro-jet reactor structure is a Y-shaped or Z-shaped micro-jet reactor. The smaller the channel size of the micro-jet reactor is, the stronger the mechanical shearing force applied to the starch particles is, the channel size is proper within 75-200 mu m, and the starch nanospheres can be prepared.

Further, the condition of the micro-jet treatment is that the micro-jet pressure is 20-200MPa, the jet speed of the solution in the micro-jet reactor is 20-150mL/min, the solution temperature is 3-30 ℃, and the repeated jet is carried out for 1-20 times.

Further, the centrifugation condition is that the centrifugation rotating speed is 4000-12000r/min, and the centrifugation time is 5-15min.

Further, the condition of the freeze-drying treatment is that the freeze-drying treatment is carried out under the vacuum drying at the temperature of-60 ℃.

Further, the conditions of the pulverizing and sieving were that the powder was pulverized with a pulverizer at 15000rpm for 90 seconds and then passed through a 200 mesh sieve.

The average grain diameter of the prepared starch nanospheres is 50.6-115.5nm.

The method has the beneficial effects that the starch nanospheres with small size and good uniformity and dispersibility are prepared by utilizing the micro-jet to induce the activation of starch hydroxyl groups and promoting the crosslinking of starch chains through alcohol precipitation treatment. The invention provides a green and efficient physical field processing technology for preparing starch nanospheres, which has the advantages of good dispersion performance, uniform surface and the like, is a good material for enhancing film performance and delivering medicines, and expands the application of the starch nanospheres in the fields of medicines, cosmetics, foods and the like.

Drawings

FIG. 1 is a process flow diagram of a starch nanosphere of uniform size prepared in accordance with the present invention;

FIG. 2 (a) is the particle size distribution chart of example 1, (b) is the particle size distribution chart of example 2, (c) is the particle size distribution chart of example 3, (d) is the particle size distribution chart of example 4, (e) is the particle size distribution chart of example 5, (f) is the particle size distribution chart of comparative example 1, (g) is the particle size distribution chart of comparative example 2, (h) is the particle size distribution chart of comparative example 3, (i) is the particle size distribution chart of comparative example 5, (j) is the particle size distribution chart of comparative example 6, and (k) is the particle size distribution chart of comparative example 7;

fig. 3 (a) is a Scanning Electron Microscope (SEM) image of example 1, (b) is a SEM image of example 2, (c) is a SEM image of example 3, (d) is a SEM image of example 4, (e) is a SEM image of example 5, (f) is a SEM image of comparative example 1, (g) is a SEM image of comparative example 2, (h) is a SEM image of comparative example 3, (i) is a SEM image of comparative example 5, (j) is a SEM image of comparative example 6, and (k) is a SEM image of comparative example 7;

Fig. 4 (a) is a Transmission Electron Microscope (TEM) image of example 1, (b) is a TEM image of example 2, (c) is a TEM image of example 3, (d) is a TEM image of example 4, (e) is a TEM image of example 5, (f) is a TEM image of comparative example 1, (g) is a TEM image of comparative example 2, (h) is a TEM image of comparative example 3, (i) is a TEM image of comparative example 5, (j) is a TEM image of comparative example 6, and (k) is a TEM image of comparative example 7;

FIG. 5 (a) is a Scanning Electron Microscope (SEM) image of the nano-starch prepared in comparative example 4 without alcohol precipitation treatment, FIG. b is a partial enlarged view of FIG. a, FIG. c is ¹ H-NMR image, and FIG. d is FTIR image;

Fig. 6 (a) is an iodine binding force (ISI) image of example 1, (b) is an ISI image of example 2, (c) is an ISI image of example 3, (d) is an ISI image of example 4, (e) is an ISI image of example 5, (f) is an ISI image of comparative example 1, (g) is an ISI image of comparative example 2, (h) is an ISI image of comparative example 3, (i) is an ISI image of comparative example 5, (j) is an ISI image of comparative example 6, and (k) is an ISI image of comparative example 7;

FIG. 7 (a) is the starch chain length profile (CLD) of example 1, (b) is the starch chain length profile of example 2, (c) is the starch chain length profile of example 3, (d) is the starch chain length profile of example 4, (e) is the starch chain length profile of example 5, (f) is the starch chain length profile of comparative example 1, (g) is the starch chain length profile of comparative example 2, (h) is the starch chain length profile of comparative example 3, (i) is the starch chain length profile of comparative example 5, (j) is the starch chain length profile of comparative example 6, and (k) is the starch chain length profile of comparative example 7;

FIG. 8 (a) is ¹³ C-NMM (carbon spectrum nuclear magnetic resonance) of example 1, (b) is ¹³ C-NMM of example 2, (C) is ¹³ C-NMM of example 3, (d) is ¹³ C-NMM of comparative example 1, (e) is ¹³ C-NMM of comparative example 2, and (f) is ¹³ C-NMM of comparative example 3;

FIG. 9 (a) shows the hydrogen nuclear magnetic resonance chart (¹ H-NMM) of example 1, (b) shows the ¹ H-NMM of example 2, (c) shows the ¹ H-NMM of example 3, (d) shows the ¹ H-NMM of comparative example 1, (e) shows the ¹ H-NMM of comparative example 2, and (f) shows the ¹ H-NMM of comparative example 3;

Fig. 10 (a) shows a Y-shape of the cavity structure of the micro-jet, and (b) shows a Z-shape of the cavity structure of the micro-jet.

Detailed Description

In order to further describe the technical means and effects adopted by the present invention for achieving the intended purpose, the following detailed description will refer to the specific implementation, structure, characteristics and effects according to the present invention with reference to the accompanying drawings and preferred embodiments.

Example 1

A preparation method of starch nanospheres with uniform size is shown in figure 1, and comprises the following steps:

(1) A gelatinized starch suspension was prepared by dispersing corn starch in water to prepare a suspension, the starch content of the starch suspension being 1wt% and gelatinizing in a boiling bath for 30min.

(2) And (3) carrying out microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Y-shaped single channel (shown in figure 10 a), the size of the channel in the cavity is 75 mu m, the injection speed of the solution in the microjet cavity is 20mL/min, the set pressure is 20MPa, the treatment times are 1, and the sample is kept constant temperature through 3 ℃ circulating water cooling water.

(3) And (3) alcohol precipitation treatment, namely dripping the solution prepared in the step (2) into 0.5 times of absolute ethyl alcohol at a rate of 5mL/min, and keeping a stirring speed of 200rpm in the process.

(4) Centrifuging, namely centrifuging the solution prepared in the step (3) for 5min at the speed of 4000rpm, removing the supernatant and retaining the precipitate.

(5) Vacuum freeze-drying, namely vacuum drying the precipitate obtained in the step (4) at the temperature of-60 ℃, grinding the powder and sieving the powder with a 200-mesh sieve.

The morphology and structure of the prepared samples were characterized as follows:

(1) Particle size measurement as shown in fig. 2a, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 a), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 a) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 a), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 a).

(4) Structure measurement, solid nuclear magnetic resonance carbon spectrum (¹³ C-NMR) was measured on the sample powder obtained in step (5) (fig. 8 a), and nuclear magnetic resonance hydrogen spectrum (¹ H-NMR) was measured on the sample powder obtained in step (5) (fig. 9 a).

The starch nanospheres prepared in this example have a particle size of 81.7nm, and form a uniform and super-spherical morphology from TEM and SEM images. From ISI, CLD, ¹³ C-NMR and ¹ H-NMR, starch nanospheres were formed by crosslinking of starch clusters.

Example 2

(1) A gelatinized starch suspension was prepared by dispersing corn starch in phosphate buffer (pH 2) with a starch ratio of 10wt% and gelatinizing in a boiling bath for 30min.

(2) And (3) carrying out microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Y-shaped multi-channel (shown in figure 10 a), the size of the channel in the cavity is 75 mu m, the injection speed of the solution in the microjet cavity is 75mL/min, the set pressure is 100MPa, the treatment times are 10 times, and the sample is kept at a constant temperature through 15 ℃ circulating water cooling water.

(3) And (3) alcohol precipitation treatment, namely dripping the solution prepared in the step (2) into 5 times of absolute ethyl alcohol at a rate of 15mL/min, and keeping a stirring speed of 1000rpm in the process.

(4) Centrifuging, namely centrifuging the solution prepared in the step (3) for 10min at a speed of 8000rpm, removing supernatant and retaining precipitate.

(5) Vacuum freeze-drying, namely vacuum drying the precipitate obtained in the step (4) at the temperature of-60 ℃, grinding and sieving, grinding for 90s by adopting a grinder at 15000rpm, and then sieving by adopting a 200-mesh sieve to obtain the starch nanospheres.

(1) Particle size measurement as shown in fig. 2b, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 b), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 b) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 b), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 b).

(4) Structure measurement, solid nuclear magnetic resonance carbon spectrum (¹³ C-NMR) was measured on the sample powder obtained in step (5) (fig. 8 b), and nuclear magnetic resonance hydrogen spectrum (¹ H-NMR) was measured on the sample powder obtained in step (5) (fig. 9 b).

The starch nanospheres prepared in this example have a particle size of 67.1nm, and form a uniform and super-spherical morphology from TEM and SEM images. From ISI, CLD, ¹³ C-NMR and ¹ H-NMR, starch nanospheres were formed by crosslinking of starch clusters.

Example 3

(1) A gelatinized starch suspension was prepared by dispersing corn starch in phosphate buffer (pH 8) with a starch content of 20wt% in the starch suspension and gelatinizing in a boiling bath for 30min.

(2) And (3) microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Z-shaped single channel (figure 10 b), the size of the channel in the cavity is 200 mu m, the injection speed of the solution in the microjet cavity is 150mL/min, the set pressure is 200MPa, the treatment times are 20 times, and the sample is kept constant temperature by 30 ℃ circulating water cooling water.

(3) And (3) alcohol precipitation treatment, namely dripping the solution prepared in the step (2) into 10 times of absolute ethyl alcohol at a rate of 30mL/min, and keeping a stirring speed of 1500rpm during the process.

(4) Centrifuging, namely centrifuging the solution prepared in the step (3) for 15min at a speed of 12000rpm, removing supernatant and retaining precipitate.

(1) Particle size measurement as shown in fig. 2c, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 c), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 c) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 c), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 c).

(4) Structure measurement, solid nuclear magnetic resonance carbon spectrum (¹³ C-NMR) was measured on the sample powder obtained in step (5) (fig. 8C), and nuclear magnetic resonance hydrogen spectrum (¹ H-NMR) was measured on the sample powder obtained in step (5) (fig. 9C).

The starch nanospheres prepared in this example have a particle size of 105.8nm, and form a uniform and super-spherical morphology from TEM and SEM images. From ISI, CLD, ¹³ C-NMR and ¹ H-NMR, starch nanospheres were formed by crosslinking of starch clusters.

Example 4

(1) A gelatinized starch suspension was prepared by dispersing rice starch and wheat starch (mass ratio 1:1) in phosphate buffer (pH 2) with a starch content of 10wt% and gelatinizing in a boiling bath for 30min.

(1) Particle size measurement as shown in fig. 2d, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 d), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 d) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 d), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 d).

The starch nanospheres prepared in this example have a particle size of 50.6nm, and form a uniform and super-spherical morphology from TEM and SEM images. From ISI and CLD, starch nanospheres are formed by cross-linking of starch clusters.

Example 5

(1) A gelatinized starch suspension is prepared, wherein sweet potato starch, tapioca starch, sweet potato starch and potato starch (the mass ratio is 1:1:1) are mixed, and then the starch is dispersed in a phosphate buffer solution (pH 8) starch suspension, the starch accounts for 20wt%, and the mixture is gelatinized for 30min in a boiling bath.

(1) Particle size measurement as shown in fig. 2e, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 e), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) after the suspension droplets are dried (figure 4 e).

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 e), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 e).

The starch nanospheres prepared in this example have a particle size of 115.5nm, and form a uniform and super-spherical morphology from TEM and SEM images. From ISI and CLD, starch nanospheres are formed by cross-linking of starch clusters.

Comparative example 1

(1) A gelatinized starch suspension was prepared by dispersing corn starch in phosphate buffer (pH 2) with a starch content of 10wt% in the starch suspension and gelatinizing in a boiling bath for 30min.

(2) And (3) microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Y-shaped multi-channel (figure 10 a), the size of the channel in the cavity is 75 mu m, the injection speed of the solution in the microjet cavity is 75mL/min, the set pressure is 100MPa, the treatment times are 25 times, and the sample is kept constant temperature by 15 ℃ circulating water cooling water.

(1) Particle size measurement as shown in fig. 2f, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 f), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 f) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 f), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 f).

(4) Structure measurement, solid nuclear magnetic resonance carbon spectrum (¹³ C-NMR) was measured on the sample powder obtained in step (5) (fig. 8 d), and nuclear magnetic resonance hydrogen spectrum (¹ H-NMR) was measured on the sample powder obtained in step (5) (fig. 9 d).

The nano particles prepared in the comparative example have the particle size of 690.2nm, and form super-spherical morphology from TEM and SEM images, but the inter-particle adhesion is serious and difficult to disperse. From ISI, CLD, ¹³ C-NMR and ¹ H-NMR, starch cluster cross-linking still occurred in the nanoparticles. In comparative example 2, it is known that the number of micro-jet treatments is important for forming starch nanospheres with good uniformity and dispersibility, and that the excessive number of micro-jet treatments leads to excessive activation of starch chains and confusion of crosslinking, resulting in aggregation of starch nanospheres and adverse dispersion of starch nanospheres.

Comparative example 2

(2) And (3) carrying out microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Y-shaped multi-channel (shown in figure 10 a), the size of the channel in the cavity is 75 mu m, the injection speed of the solution in the microjet cavity is 75mL/min, the set pressure is 10MPa, the treatment times are 10 times, and the sample is kept constant temperature by 15 ℃ circulating water cooling water.

(1) Particle size measurement As shown in FIG. 2g, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of the nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, carrying out metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 g), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 g) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 g), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 g).

(4) Structure measurement, solid nuclear magnetic resonance carbon spectrum (¹³ C-NMR) of the sample powder obtained in step (5) (fig. 8 e), and nuclear magnetic resonance hydrogen spectrum (¹ H-NMR) of the sample powder obtained in step (5) (fig. 9 e).

The particle size of the nano particles prepared in the comparative example is 810.6nm, and the spherical morphology boundary is unclear, so that the particles are seriously adhered and difficult to disperse from TEM and SEM images. From ISI, CLD, ¹³ C-NMR and ¹ H-NMR, starch cluster cross-linking still occurred in the nanoparticles. Comparative examples 1, 2 and 3 show that the low pressure treatment of the microfluidic treatment, too low pressure and too weak mechanical force, cannot shear starch chains to form starch nanospheres, and is detrimental to the formation and dispersion of starch nanospheres.

Comparative example 3

(1) A gelatinized starch suspension was prepared by dispersing corn starch in phosphate buffer (pH 2) to prepare a 10wt% suspension and gelatinizing in a boiling bath for 30min.

(2) And (3) carrying out microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Y-shaped multi-channel (shown in figure 10 a), the size of the channel in the cavity is 75 mu m, the injection speed of the solution in the microjet cavity is 75mL/min, the set pressure is 250MPa, the treatment times are 10 times, and the sample is kept at a constant temperature through 15 ℃ circulating water cooling water.

(1) Particle size measurement as shown in fig. 2h, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample by using a Scanning Electron Microscope (SEM) under the acceleration voltage of 3KV (figure 3 h), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets by using a Transmission Electron Microscope (TEM) under the acceleration voltage of 80KV after the suspension droplets are dried (figure 4 h).

(3) Chain length distribution measurement, namely, measuring binding force (ISI) between the sample powder obtained in the step (5) and iodine (figure 6 h), and measuring Chain Length Distribution (CLD) of starch by size exclusion chromatography after enzymolysis of the sample powder obtained in the step (5) (figure 7 h).

(4) Structure measurement, solid nuclear magnetic resonance carbon spectrum (¹³ C-NMR) was measured on the sample powder obtained in step (5) (fig. 8 f), and nuclear magnetic resonance hydrogen spectrum (¹ H-NMR) was measured on the sample powder obtained in step (5) (fig. 9 f).

The nano particles prepared in this comparative example have a particle size of 750.9nm, and from the TEM and SEM images, the conditions formed starch nano sphere particles, but the inter-particle adhesion was severe and difficult to disperse. From ISI, CLD, ¹³ C-NMR and ¹ H-NMR, starch cluster cross-linking still occurred in the nanoparticles. Comparative example 2 shows that the high pressure treatment of the microfluidic treatment, too high pressure, results in an increase in van der Waals and electrostatic attraction forces on the surface of the starch nanospheres, and the formation of secondary aggregated particles is detrimental to the dispersion of the starch nanospheres.

Comparative example 4

(1) A gelatinized starch suspension was prepared by dispersing corn starch in phosphate buffer (ph 5.6) with a starch content of 3wt% in the starch suspension and gelatinizing in a boiling bath for 30min.

(2) And (3) microjet treatment, namely injecting the solution treated in the step (1) into microjet equipment, wherein the cavity is made of diamond material, the cavity structure is a Y-shaped single channel (shown in figure 10 a), the size of the channel in the cavity is 75 mu m, the injection speed of the solution in the microjet cavity is 75mL/min, the set pressure is 100MPa, the treatment times are 1, and the sample is kept constant temperature through 4 ℃ circulating water cooling water.

(3) Dialyzing the solution prepared in the step (2) for 2 days under the condition of 3500Da molecular, and removing the soluble micromolecules and phosphate. Samples were obtained that were microjet treated without alcohol precipitation.

(4) Vacuum freeze-drying, namely vacuum drying the sample obtained in the step (3) at the temperature of-60 ℃, grinding the sample into powder and sieving the powder through a 200-mesh sieve.

(1) Morphology observation the sample powder obtained in step (4) was uniformly coated on a conductive adhesive and subjected to a metal spraying treatment, and the surface morphology of the sample was observed under an acceleration voltage of 3KV by means of a Scanning Electron Microscope (SEM) (FIGS. 5 a-b).

(2) Structure measurement, namely, nuclear magnetic resonance hydrogen spectrogram (¹ H-NMR) of the sample powder obtained in the step (5) (fig. 5 c), and Fourier infrared spectrogram (FTIR) of the sample powder obtained in the step (5) (fig. 5 d).

The nanoparticles prepared in this comparative example had no super-spherical structure and had non-uniform particle size distribution as seen from SEM images. The cross-linking of the starch clusters is not evident from FTIR and ¹ H-NMR. Comparing examples 1,2 and 3, it is known that the alcohol precipitation treatment is an essential step for forming starch nanospheres after the micro-jet treatment.

Comparative example 5

(1) A gelatinized starch suspension was prepared by dispersing corn starch in phosphate buffer (ph 5.6) with a starch content of 10wt% in the starch suspension and gelatinizing in a boiling bath for 30min.

(3) And (3) alcohol precipitation treatment, namely dripping the solution prepared in the step (2) into 15 times of absolute ethyl alcohol at a rate of 15mL/min, and keeping a stirring speed of 1000rpm in the process.

Characterization of morphology and structure of the prepared samples:

(1) Particle size measurement as shown in fig. 2i, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, carrying out metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 i), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 i) after the suspension droplets are dried.

(3) Chain length distribution measurement, namely, measuring binding force (ISI) between the sample powder obtained in the step (5) and iodine (figure 6 i), and measuring Chain Length Distribution (CLD) of starch by size exclusion chromatography after enzymolysis of the sample powder obtained in the step (5) (figure 7 i).

The finished product prepared in the comparative example has the particle size of 1548.9nm, is micron-sized particles from SEM and TEM images, and cannot obtain starch nanospheres. From ISI and CLD, starch cluster cross-linking still occurs. In comparative examples 1,2 and 3, it is known that the volume ratio of ethanol to starch cluster solution with an excessively high ratio is unfavorable for the formation of small-particle nano-starch, and the collision degree of starch clusters is aggravated by the excessively high ratio of ethanol to water, so that the cross-linking between starch clusters is disordered, and the small-particle nano-starch is difficult to be orderly arranged.

Comparative example 6

(3) And (3) alcohol precipitation treatment, namely dripping the solution prepared in the step (2) into 5 times of absolute ethyl alcohol at a rate of 40mL/min, and keeping a stirring speed of 1000rpm in the process.

(1) Particle size measurement as shown in fig. 2j, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 j), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) after the suspension droplets are dried (figure 4 j).

(3) Chain length distribution measurement, namely, measuring binding force (ISI) between the sample powder obtained in the step (5) and iodine (figure 6 j), and measuring Chain Length Distribution (CLD) of starch by size exclusion chromatography after enzymolysis of the sample powder obtained in the step (5) (figure 7 j).

The nano starch prepared in the comparative example has the particle size of 785.6nm, and the nano starch forms spherical nano particles with different sizes from the SEM and TEM images. From ISI and CLD, starch cluster cross-linking still occurs. Comparative examples 1, 2 and 3 show that too high a speed of ethanol pouring allows local starch clusters to crosslink too fast to precipitate as soon as starch nanospheres are ordered, resulting in non-uniform finished product size and poor dispersion of the formed particles.

Comparative example 7

(3) And (3) alcohol precipitation treatment, namely dripping the solution prepared in the step (2) into 5 times of absolute ethyl alcohol at a rate of 15mL/min, and keeping a stirring speed of 5000rpm during the process.

(1) Particle size measurement as shown in fig. 2k, a nano starch suspension was prepared, and after dispersion by a homogenizer, the particle size distribution of nano starch was measured by a nano particle sizer (DLS).

(2) Morphology observation, namely uniformly coating the sample powder obtained in the step (5) on a conductive adhesive, performing metal spraying treatment, observing the surface morphology of the sample under an acceleration voltage of 3KV by using a Scanning Electron Microscope (SEM) (figure 3 k), configuring the sample powder obtained in the step (5) into suspension droplets on a carbon-coated copper grid, and observing the suspension droplets under an acceleration voltage of 80KV by using a Transmission Electron Microscope (TEM) (figure 4 k) after the suspension droplets are dried.

(3) Chain length distribution measurement the binding force (ISI) with iodine was measured on the sample powder obtained in step (5) (fig. 6 k), and the Chain Length Distribution (CLD) of starch was measured by size exclusion chromatography after enzymatic hydrolysis of the sample powder obtained in step (5) (fig. 7 k).

The nano starch prepared in the comparative example has the particle size of 394.6nm, and the nano starch forms spherical nano particles with different sizes from the SEM and TEM images. From ISI and CLD, starch cluster cross-linking still occurs. Comparative examples 1, 2 and 3 show that too fast an alcohol precipitation stirring speed allows local starch clusters to crosslink too fast to precipitate as soon as starch nanospheres are ordered, thus resulting in uneven finished product size and poor dispersion of the formed particles.

From the above examples and comparative examples, it can be seen that the alcohol precipitation reaction is particularly critical for the post-treatment of starch microfluidics. The starch grain crystal structure after the thermal gelatinization collapses, starch chains are unfolded, starch solution passes through a dynamic micro-jet physical field (mechanical actions such as high pressure effect, high-speed impact, high-frequency vibration, instantaneous pressure drop, strong shearing and cavitation effect are involved), starch macromolecules are violently collided with a micro-jet cavity, hydroxyl groups on starch chain links are activated, starch main chains and side chains are broken, starch small cluster units with different sizes of activated hydroxyl groups are formed, and starch small clusters with different chain length distributions can be designed by changing the pressure and the treatment times of micro-jet treatment conditions. In the nano alcohol precipitation process, because of unbalanced interactions such as tension change, flow, diffusion and the like generated between ethanol and water interfaces of a system, oscillation and reaction occur among small starch clusters, the action among hydroxyl groups of different chain segments is activated to form new C-O-C bonds, and the crosslinking reaction between the starch clusters and the inside of the clusters is promoted, so that starch nano ball particles with smooth surfaces are orderly arranged. In the whole system, the alcohol precipitation reaction not only provides the condition of collision and crosslinking of starch clusters, but also increases the saturation of the system to play a role in separation and purification, so that the nano starch with uniform particle size and good dispersibility is obtained. In the process, the ratio of ethanol to water, the speed of pouring the ethanol into the starch solution and the stirring speed of the starch solution all influence the final sample effect, the collision degree of starch clusters is aggravated by the excessively high ratio of the ethanol to the water, the cross-linking between the starch clusters is disordered, small-particle nano starch is difficult to be orderly arranged, the system saturation of the excessively low ratio of the ethanol to the water is insufficient, the cross-linking between the starch clusters is insufficient, nano starch particles are difficult to separate out from the system, and a finished product is difficult to obtain. The speed of pouring the ethanol into the starch solution and the stirring speed of the starch solution relate to mass transfer and heat transfer of a system, local starch clusters are crosslinked too fast to be separated out as soon as starch nanospheres are ordered, so that the finished product is uneven in size, and the too slow mass transfer and heat transfer speed influences the crosslinking reaction efficiency among the starch clusters and is also unfavorable for the formation of uniform starch nanospheres.

The present invention is not limited in any way by the above-described preferred embodiments, but is not limited to the above-described preferred embodiments, and any person skilled in the art will appreciate that the present invention can be embodied in the form of a program for carrying out the method of the present invention, while the above disclosure is directed to equivalent embodiments capable of being modified or altered in some ways, it is apparent that any modifications, equivalent variations and alterations made to the above embodiments according to the technical principles of the present invention fall within the scope of the present invention.

Claims

1. A method for preparing starch nanospheres of uniform size, characterized in that it comprises the following steps:

(1) dispersing starch in water or a phosphate buffer solution with a pH of 2-8 to obtain a starch suspension with a mass fraction of 1-20 wt%; gelatinizing the starch suspension and subjecting it to microfluidization to form a starch cluster solution; the microfluidization treatment uses a microfluidizer, the microfluidizer structure is a "Y" type or "Z" type microfluidizer, and the channel size is 75-200 μm; the conditions of the microfluidization treatment are: the microfluidizer pressure is 20-200 MPa, the solution injection rate in the microfluidizer is 20-150 mL/min, the solution temperature is 3-30°C, and the injection is repeated 1-20 times;

(2) The starch cluster solution is subjected to alcohol precipitation treatment, the soluble molecules are removed by centrifugation, the precipitate is freeze-dried, ground and sieved to obtain starch nanospheres;

The alcohol precipitation treatment is as follows: anhydrous ethanol is uniformly dripped into the starch cluster solution at a speed of 5-30 mL/min, stirring is maintained during the alcohol precipitation process, the stirring speed is 200-1500 r/min, and the volume ratio of anhydrous ethanol to starch cluster solution is 0.5:1-10:1.

2. preparation method according to claim 1, is characterized in that the starch of described starch suspension is one or more in rice starch, corn starch, wheat starch, sweet potato starch, tapioca starch, sweet potato starch, potato starch.

3. The preparation method according to claim 1 is characterized in that the centrifugal conditions are: the centrifugal speed is 4000-12000 r/min, and the centrifugal time is 5-15 min.

4. The preparation method according to claim 1, characterized in that the freeze-drying treatment is carried out under vacuum at -60°C.

5. The preparation method according to claim 1, characterized in that the conditions for grinding and screening are: grinding with a grinder at 15000 rpm for 90 seconds and then passing through a 200 mesh sieve.