A 3D printing ink for food, its preparation method and application

Abstract

The present invention discloses a food 3D printing ink and its preparation method and application. The preparation method includes: subjecting a pea protein dispersion to high-pressure homogenization and ultrasonic treatment in sequence to obtain a modified pea protein; fully mixing and reacting sodium alginate, the modified pea protein and water to obtain a modified pea protein / sodium alginate complex; and mixing medium-chain triglycerides with a solution containing the modified pea protein / sodium alginate complex and subjecting the mixture to shear treatment to obtain a food 3D printing ink. Compared with the 3D printing ink prepared from natural pea protein, the food 3D printing ink prepared by the present invention has a smaller particle size, a more uniform and compact microstructure, a stronger storage modulus and apparent viscosity. At the same time, the 3D printing model prepared has a smooth and regular shape, a stable three-dimensional structure and fine resolution, and can maintain the stability of the structure within 24 hours.

Description

Technical Field

[0001] The present invention belongs to the technical field of food 3D printing inks, and particularly relates to a food 3D printing ink, a preparation method thereof, and an application thereof. Background Art

[0002] 3D printing can rapidly and precisely construct three-dimensional objects through digital control technology, and has attracted wide attention due to its advantages of high cost-effectiveness, customizable product shape, and rapid product creation. In recent years, 3D printing technology has rapidly emerged in the food field because of its potential to manufacture foods that simultaneously meet morphological requirements and nutritional characteristics. Existing research has applied 3D printing technology to various food designs, such as 3D printing of cereal-based foods using pea protein, wheat, corn starch, and potato starch, and 3D printing of gel-based foods using lemon juice as a printing material. The properties of 3D printing ink are the basis of 3D printing. To achieve food 3D printing, food materials with high support and molding capabilities are required as inks.

[0003] Plant proteins have attracted the interest of many consumers due to their high yield, low price, sustainability, and health benefits, especially as substitutes for animal proteins in various foods. Among them, pea protein has broad prospects as a 3D printing food raw material due to its high nutritional value, low production cost, low allergenicity, non-GMO status, and wide applicability. However, compared with most animal proteins, pea protein has lower solubility and poor emulsifying ability, resulting in unsatisfactory performance of 3D printing inks prepared from it. Physical modification is a feasible method to improve the functional properties of poorly soluble proteins. Compared with other methods, physical modification has the advantages of low cost, short time consumption, no toxic side effects, little damage to protein nutrition, and no introduction of other impurities. However, physical modification has the defect of incomplete modification in improving the solubility, foaming property, and emulsifying property of poorly soluble proteins. Single use of a certain physical modification method cannot guarantee that the modified protein reaches the required function.

[0004] Currently, food materials for extrusion-based 3D printing are mainly limited to dough, starch, gels, hydrocolloids, high internal phase emulsions, etc. Among these food-grade printing inks, the application of emulsion systems in 3D printing is restricted mainly because most emulsions are prone to deformation, difficult to maintain the designed shape, and prone to phase separation during the extrusion process of 3D printing. In the emulsion system, the melting behavior and plasticization of the emulsion can be regulated by changing the emulsifying properties, composition, microstructure, and composite ratio of the emulsifier, which provides a new means for designing emulsion-based 3D printing inks. However, due to the poor properties of commercial plant proteins, inks based on plant proteins usually rely on the addition of high-concentration proteins (greater than 5 wt%) and the induction of gelling agents (such as transglutaminase and glucono-d-lactone), which inevitably leads to an increase in production costs and process complexity and cannot meet the diverse needs of 3D printed foods. Therefore, how to improve the properties of plant proteins and the functions of emulsifiers so that they can be used as emulsions for 3D printing materials remains a technical gap today. Summary of the Invention

[0005] The main object of the present invention is to provide a food 3D printing ink, its preparation method and application to overcome the deficiencies of the prior art.

[0006] To achieve the foregoing invention object, the technical solutions adopted by the present invention include:

[0007] An embodiment of the present invention provides a preparation method of a food 3D printing ink, which includes:

[0008] Perform high-pressure homogenization and ultrasonic treatment on the pea protein dispersion in sequence to obtain modified pea protein;

[0009] Fully mix and react sodium alginate, the modified pea protein and water to obtain a modified pea protein / sodium alginate complex;

[0010] And, mix the edible oil with the solution containing the modified pea protein / sodium alginate complex and perform shear treatment to obtain a food 3D printing ink.

[0011] An embodiment of the present invention also provides the food 3D printing ink prepared by the foregoing preparation method, and the content of the modified pea protein / sodium alginate complex in the food 3D printing ink is 0.3 wt% - 1.0 wt%.

[0012] An embodiment of the present invention also provides a preparation method of a modified pea protein / sodium alginate complex, which includes:

[0013] Perform high-pressure homogenization and ultrasonic treatment on the pea protein dispersion in sequence to obtain modified pea protein;

[0014] And, sodium alginate and the modified pea protein are fully mixed and reacted with water to obtain a modified pea protein / sodium alginate composite.

[0015] The embodiment of the present invention also provides a 3D printing method, which includes:

[0016] Providing the aforementioned food 3D printing ink;

[0017] And, using an extrusion 3D printing method with a nozzle to print the food 3D printing ink to obtain a food 3D printed part.

[0018] The embodiment of the present invention also provides a modified pea protein / sodium alginate composite prepared by the aforementioned preparation method.

[0019] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0020] (1) The present invention prepares a modified pea protein / sodium alginate (denoted as HU-PPI-SA) composite by simple physical mixing. As the concentration of sodium alginate (denoted as SA) increases, the particle size of the composite significantly increases, presenting a uniform granular structure under a microscope.

[0021] (2) In the HU-PPI-SA composite of the present invention, the modified pea protein (denoted as HU-PPI) and SA are mainly combined through electrostatic interaction and hydrogen bond; a higher concentration of SA (HU-PPI:SA concentration ratio ≤ 2:1) will change the secondary structure of HU-PPI, making it looser.

[0022] (3) The food 3D printing ink prepared by the present invention has a smaller particle size, a more uniform and compact microstructure, a stronger storage modulus and apparent viscosity, and can be used for 3D printing.

[0023] (4) The 3D printed model obtained by using the food 3D printing ink in the present invention has a smooth and regular shape, a stable three-dimensional structure, a fine resolution, and can maintain the structural stability within 24 hours. Description of the Drawings

[0024] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only some embodiments recorded in the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0025] Figures 1a - 1b It is the particle size, polydispersity index PDI and ζ-potential diagram of HU-PPI and HU-PPI-SA composites in Example 1 of the present invention;

[0026] Figures 2a - 2f These are SEM images of the HU-PPI and HU-PPI-SA complexes in Example 1 of the present invention (in the figure, 10:1 - 1:1 are the complexes formed when the mass ratio of HU-PPI to SA is 10:1 to 1:1).

[0027] Figures 3a - 3b These are the Fourier transform infrared spectra and circular dichroism spectra of the HU-PPI and HU-PPI-SA complexes in Example 1 of the present invention.

[0028] Figure 4 These are the apparent images of the high internal phase Pickering emulsions (HIPPEs) stabilized by the commercial pea protein / sodium alginate (denoted as C-PPI-SA) mixture and the HU-PPI-SA complex in Example 2 of the present invention.

[0029] Figures 5a - 5b These are the average particle size images of the HIPPEs stabilized by the C-PPI-SA mixture and the HU-PPI-SA complex in Example 2 of the present invention.

[0030] Figures 6a - 6b These are the optical microscope images of the HIPPEs stabilized by the C-PPI-SA mixture and the HU-PPI-SA complex in Example 2 of the present invention.

[0031] Figure 7 These are the CLSM images of the HIPPEs stabilized by the C-PPI-SA mixture and the HU-PPI-SA complex in Example 2 of the present invention.

[0032] Figures 8a - 8b These are the storage modulus and loss modulus images of the HIPPEs stabilized by the C-PPI-SA mixture and the HU-PPI-SA complex in Example 2 of the present invention.

[0033] Figures 8c - 8d These are the apparent viscosity images of the HIPPEs stabilized by the C-PPI-SA mixture and the HU-PPI-SA complex in Example 2 of the present invention.

[0034] Figure 9 These are the schematic diagrams of the mechanism for the synergistic stabilization of HIPPEs by commercial and modified PPI and SA in Example 2 of the present invention (HPH-US represents high-pressure homogenization first and then ultrasonic treatment).

[0035] Figure 10 These are the line extrusion morphology images of the HIPPEs in Example 2 of the present invention (the mass ratio of PPI to SA is 4:1).

[0036] Figure 11is a 3D printed morphology diagram of HIPPEs in Example 2 of the present invention (the ratio in the figure is the mass ratio of PPI to SA);

[0037] Figures 12a - 12b This is a storage stability diagram of the high internal phase emulsion 3D printed model in Example 2 of the present invention (the ratio in the figure is the mass ratio of PPI to SA). DETAILED DESCRIPTION

[0038] In view of the defects of the prior art, the inventor of this case has proposed the technical solution of the present invention after long-term research and extensive practice. The technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0039] Specifically, as one aspect of the technical solution of the present invention, a method for preparing a food 3D printing ink involves:

[0040] The pea protein dispersion is subjected to high pressure homogenization and ultrasonic treatment in sequence to obtain modified pea protein;

[0041] The sodium alginate, the modified pea protein and water are fully mixed and reacted to obtain a modified pea protein / sodium alginate complex;

[0042] And, medium-chain triglycerides are mixed with a solution containing the modified pea protein / sodium alginate complex and subjected to shearing treatment to prepare food 3D printing ink.

[0043] In some preferred embodiments, the content of pea protein in the pea protein dispersion is 0.5-1.5 wt %.

[0044] In some preferred embodiments, the pressure used in the high-pressure homogenization treatment is 30-70 MPa, and the number of cycles is 3-5 times.

[0045] In some preferred embodiments, the ultrasonic treatment uses a power of 200-600 W and a treatment time of 15-25 min.

[0046] In some more specific embodiments, the preparation method specifically comprises:

[0047] Disperse 1.0 wt% of pea protein (PPI) powder in distilled water and continuously stir it for 3 h at 25 °C using a magnetic stirrer. Then, adjust the pH value of the sample solution to 7.0 ± 0.2 using 2.0 M NaOH, hydrate it overnight at 4 °C, and process the PPI solution three cycles at 50 MPa using a high-pressure homogenizer to obtain high-pressure homogenized PPI labeled as H-PPI;

[0048] Use an ultrasonic cell disruptor to process the H-PPI solution for 20 min at 20 kHz and 400 W ultrasonic power, with the ultrasonic probe immersed about 2 cm below the PPI solution to prepare a modified pea protein solution.

[0049] In some preferred embodiments, the preparation method specifically includes: fully stirring and mixing a dispersion containing modified pea protein with a sodium alginate solution to react to obtain a modified pea protein / sodium alginate complex; the modified pea protein and sodium alginate are mainly combined through electrostatic interaction and hydrogen bonding; wherein, the mass ratio of the modified pea protein to sodium alginate is 8:1 - 2:1.

[0050] Furthermore, the mass ratio of the modified pea protein to sodium alginate is 6:1 - 2:1.

[0051] In some preferred embodiments, the preparation method specifically includes: using a high-speed shear dispersion device and under the condition of a rotation speed of 3000 - 6000 rpm, dropwise add edible oil to the modified pea protein / sodium alginate complex solution and mix evenly, and then perform high-speed shearing for 100 - 200 s under the condition of a rotation speed of 8000 - 12000 rpm to prepare a food 3D printing ink.

[0052] Furthermore, the volume ratio of the edible oil to the modified pea protein / sodium alginate complex solution is 3:1 - 5:1.

[0053] Even further, the volume ratio of the edible oil to the modified pea protein / sodium alginate complex solution is 3:1, 4:1 or 5:1.

[0054] Furthermore, the edible oil includes any one or a combination of two or more of medium-chain triglycerides, corn oil, and algal oil, and is not limited thereto.

[0055] Furthermore, the content of the modified pea protein / sodium alginate complex in the modified pea protein / sodium alginate complex solution is 1.0 wt% - 3.0 wt%.

[0056] In some more specific embodiments, the preparation method specifically includes:

[0057] The modified pea protein (denoted as HU-PPI) solution was freeze-dried and then sealed and stored at -20 °C for later use;

[0058] Weigh 2 g of the HU-PPI freeze-dried sample and redissolve it in 100 mL of distilled water. Stir magnetically at 600 rpm for half an hour and then hydrate overnight at 4 °C. Weigh 2 g of sodium alginate (SA) and dissolve it in 100 mL of distilled water. Stir overnight at 600 rpm to fully dissolve it;

[0059] Dilute the SA aqueous solution to a certain concentration and mix it with the 2 wt% HU-PPI dispersion in equal volumes. Stir magnetically for 30 min to mix the two evenly to obtain the modified pea protein / sodium alginate complex (HU-PPI-SA complex).

[0060] Another aspect of the embodiments of the present invention also provides a food 3D printing ink prepared by the aforementioned preparation method, and the content of the modified pea protein / sodium alginate complex in the food 3D printing ink is 0.3 wt% - 1.0 wt%.

[0061] Another aspect of the embodiments of the present invention also provides a 3D printing method, which includes:

[0062] Providing the aforementioned food 3D printing ink;

[0063] And, using the nozzle extrusion 3D printing method to print the food 3D printing ink to obtain a food 3D printed part.

[0064] Another aspect of the embodiments of the present invention also provides a preparation method of a modified pea protein / sodium alginate complex, which includes:

[0065] Subjecting the pea protein dispersion to high-pressure homogenization and ultrasonic treatment in sequence to obtain modified pea protein;

[0066] And, fully mixing and reacting sodium alginate, the modified pea protein and water to obtain the modified pea protein / sodium alginate complex.

[0067] In some preferred embodiments, the content of pea protein in the pea protein dispersion is 0.5 - 1.5 wt%.

[0068] In some preferred embodiments, the pressure used for the high-pressure homogenization treatment is 30 - 70 MPa, and the number of cycles is 3 - 5 times.

[0069] In some preferred embodiments, the power used for the ultrasonic treatment is 200 - 600 W, and the treatment time is 15 - 25 min.

[0070] In some preferred embodiments, the preparation method specifically includes: fully stirring and mixing a dispersion containing modified pea protein with a sodium alginate solution to obtain a modified pea protein / sodium alginate complex; the modified pea protein and sodium alginate are mainly combined through electrostatic interaction and hydrogen bonding; wherein, the mass ratio of the modified pea protein to sodium alginate is 8:1 - 2:1.

[0071] Further, the mass ratio of the modified pea protein to sodium alginate is 6:1 - 2:1.

[0072] Another aspect of the embodiments of the present invention also provides a modified pea protein / sodium alginate complex prepared by the foregoing preparation method.

[0073] The technical solutions of the present invention will be further described in detail below in conjunction with several preferred embodiments and the accompanying drawings. These embodiments are implemented on the premise of the technical solutions of the present invention, and detailed implementation manners and specific operation processes are given. However, the protection scope of the present invention is not limited to the following embodiments.

[0074] The experimental materials used in the following examples can be obtained from conventional biochemical reagent companies without special instructions.

[0075] The specific materials and reagents are shown in Table 1:

[0076] Table 1

[0077]

[0078] The specific instruments and equipment are shown in Table 2:

[0079] Table 2

[0080]

[0081] Example 1

[0082] Disperse 1.0 wt% of commercial pea protein (C-PPI) powder in distilled water and continuously stir with a magnetic stirrer at 25 °C for 3 h. Then, adjust the pH value of the sample solution to 7.0 ± 0.2 with 2.0 M NaOH and hydrate overnight at 4 °C. Treat the PPI solution with a high-pressure homogenizer at 50 MPa for three cycles to obtain high-pressure homogenized PPI labeled as H-PPI;

[0083] Treat the H-PPI solution with an ultrasonic cell disruptor at 20 kHz and 400 W ultrasonic power for 20 min, with the ultrasonic probe immersed about 2 cm below the PPI solution to obtain a modified pea protein (denoted as HU-PPI) solution;

[0084] The modified pea protein (denoted as HU-PPI) solution was freeze-dried and stored sealed at -20 °C for later use;

[0085] Weighed 2 g of the HU-PPI freeze-dried sample and redissolved it in 100 mL of distilled water. After magnetic stirring at 600 rpm for half an hour, it was hydrated overnight at 4 °C. Weighed 2 g of the polysaccharide sodium alginate (SA) and dissolved it in 100 mL of distilled water, and stirred it overnight at 600 rpm to fully dissolve it;

[0086] The SA aqueous solution was diluted to a certain concentration and then mixed with an equal volume of a 2 wt% HU-PPI dispersion. Magnetic stirring was carried out for 30 min to make the two mix evenly, obtaining the modified pea protein / sodium alginate complex (HU-PPI-SA complex). The pH values of these complexes were measured to be between 7.0 and 6.5.

[0087] Performance characterization:

[0088] 1. Particle size and ζ-potential of the HU-PPI-SA complex

[0089] Test method: A nano laser particle size analyzer was used to measure the average particle size and particle size distribution of the sample. The sample was measured after equilibrating at 25 °C for 2 minutes, and the measurement was repeated three times. A dynamic light scattering analyzer was used to measure the ζ-potential of HU-PPI and the HU-PPI-SA complex. The sample was diluted with distilled water, and the experiment was repeated 3 times at 25 °C.

[0090] Figures 1a - 1b The ratio in the figure is the mass ratio of PPI to SA, and different lowercase letters indicate significant differences (p < 0.05); First, the effect of SA concentration on the particle size of the HU-PPI-SA complex particles was explored. The HU-PPI formed by the synergistic treatment of high-pressure homogenization and ultrasonic treatment formed nanoparticles with an average particle size of 227.6 ± 9.2 nm ( Figure 1a ), with the increase of the added SA concentration, the particle size first increased slowly. When the concentration ratio of HU-PPI-SA was 3:1, the particle size was 457.4 ± 42.4 nm, and then it increased extremely rapidly to 841.3 ± 201.2 nm (concentration ratio of 2:1), 1063.2 ± 546.1 nm (concentration ratio of 1:1). This indicates that there is a certain interaction between HU-PPI and SA, which can combine to form composite particles and increase the particle size. Existing studies have shown that PPI and SA form electrostatic complexes at a pH value near 6.5.

[0091] In addition, HU-PPI is negatively charged under neutral pH conditions, and the anionic polysaccharide SA is also negatively charged. After adding SA, the absolute value of the ζ-potential of the system increased slightly ( Figure 1b)。Interestingly, the ζ-potential of the complex remained around -30 mV in the SA concentration range of proteoglycan ratios from 8:1 to 3:1, but increased significantly to -34.7 ± 0.3 mV when the concentration ratio reached 2:1, and reached -46.8 ± 1.0 mV at a HU-PPI-SA concentration of 1:1. This may be because when the SA concentration is low, there is an electrostatic interaction between the negatively charged SA and the positively charged patches on the surface of PPI molecules, forming a soluble complex. As this interaction increases, the particle size of the corresponding complex also gradually increases. When the SA concentration increases to more than 0.5 wt% (concentration ratios of 2:1 and 1:1), the positive charges on the surface of HU-PPI molecules that can react with SA are already saturated, and the excess SA present in the system significantly reduces the ζ-potential. These results indicate that electrostatic interaction plays an important role in the formation of the complex.

[0092] 2. Microstructure of HU-PPI-SA Complex

[0093] Test method: The freeze-dried sample was adhered to a cylindrical aluminum sample stage using a double-sided carbon tape, and then gold was sprayed on the surface of the sample. The surface morphology of the protein was observed using a scanning electron microscope at an acceleration voltage of 5 kV. The prepared complex was freeze-dried and the microstructure was observed using a scanning electron microscope (SEM).

[0094] The micro-morphology of the HU-PPI-SA complex was observed by scanning electron microscope (SEM) ( Figures 2a - 2f , the ratio in the figure is the mass ratio of PPI to SA). The PPI modified by combined high-pressure homogenization and ultrasonic treatment is uniformly distributed spherical small particles. After adding sodium alginate, the particles seem to become compact and smooth; as the SA concentration increases, the particles gradually increase; when the concentration ratio of HU-PPI to SA is 2:1, there are some voids between the particles, and these voids become more obvious at 1:1. This is consistent with the results of the particle size. The increasing particles indicate that more SA is complexed with HU-PPI, and the voids appearing in the samples with high SA concentration indicate that SA is in excess and fills between the proteins.

[0095] 3. Interaction Analysis of HU-PPI-SA Complex

[0096] Test method: Fourier transform infrared spectroscopy (FTIR): Take 2 mg of the freeze-dried complex and mix it with 198 mg of KBr, grind it evenly and press it into a thin slice, and measure the wave number range of 4000 - 400 cm -1 , scan 64 times, and set the resolution to 4 cm -1 ;

[0097] Circular dichroism (CD): After diluting the HU-PPI and HU-PPI-SA composite solutions to a protein concentration of 0.2 mg / mL, circular dichroism analysis was performed.

[0098] Figures 3a - 3b In the ratio, it is the mass ratio of PPI to SA. FTIR spectroscopy is a common method to reveal the molecular interactions between protein and polysaccharide functional groups. As Figure 3a shown: In the spectrum of HU-PPI, the prominent characteristic peak at 1655 cm -1 corresponds to the C=O stretching vibration (amide I band), and the prominent characteristic peak at 1543 cm -1 corresponds to the N-H bending vibration (amide II band). After complexing with SA, these two characteristic peaks still exist and do not shift, indicating that the interaction between SA and HU-PPI is not related to the amide group (Zhan et al. 2019). In addition, the sample has a broad absorption peak in the range of 3600 - 3000 cm -1 , which is related to the stretching vibration of O-H. Here, the absorption peak of HU-PPI at 3292 cm -1 gradually shifts towards a longer wavelength after adding SA. When HU-PPI and SA are complexed in a 1:1 ratio, the peak position redshifts to 3422 cm -1 , indicating that hydrogen bonds play an important role in the binding of HU-PPI and SA.

[0099] Far-ultraviolet CD spectra were used to analyze the changes in the secondary structure of HU-PPI during different complexation processes. As Figure 3b depicts the CD spectra of HU-PPI and HU-PPI-SA complexes, and the secondary structure content of the protein was obtained by online analysis using DichroWeb (see Table 3). The influence of lower concentrations of SA on the secondary structure of HU-PPI is not obvious (8:1 and 4:1), and the CD curves of the samples basically overlap with that of HU-PPI, which is consistent with previous studies. The simple physical mixing with the polysaccharide sample did not change the secondary structure of soy protein isolate. When the SA concentration increased to a HU-PPI:SA ratio of 2:1, the curve deviated significantly, the maximum absorption peak redshifted and the intensity decreased. At this time, compared with HU-PPI, the α-helix content of the complex decreased significantly, and the random coil content increased significantly. It has been reported in the literature that ordered secondary structures (such as α-helix and β-sheet) are usually hidden in the polypeptide chain. Therefore, the reduction of these structures confirms that the attachment of anionic polysaccharides changes the spatial conformation of the protein, resulting in a looser protein structure. The increase in the random coil content further confirms the unfolding of the protein molecular structure. This may be because there is an excess of SA in the system, forming large aggregates with HU-PPI.

[0100] Table 3 Secondary structure contents (%) of HU-PPI and HU-PPI-SA complexes

[0101]

[0102] *Note: Different lowercase letters indicate significant differences in the contents of the same type of secondary structure among different samples (p < 0.05). The ratios in the table are the mass ratios of PPI to SA

[0103] In summary, the present application prepared the HU-PPI-SA complex by simple physical mixing. As the concentration of SA increased, the particle size of the complex increased significantly, presenting a uniformly sized granular structure under the microscope, confirming the formation of the complex; the main binding between HU-PPI and SA was through electrostatic interaction and hydrogen bond; a higher concentration of SA (HU-PPI∶SA concentration ratio ≤ 2∶1) would change the secondary structure of HU-PPI, making it more loose

[0104] Example 2

[0105] High internal phase emulsions were prepared using commercial pea protein PPI (C-PPI), HU-PPI modified by combined high-pressure homogenization and ultrasonication, C-PPI-SA mixture, and HU-PPI-SA complex. Using a high-speed shear disperser, at a rotation speed of 6000 rpm, 12 mL of medium-chain triglyceride (MCT) was slowly added dropwise to 4 mL of a C-PPI or C-PPI-SA solution with a protein concentration of 1.0 wt%. After mixing evenly, the rotation speed was adjusted to 12000 rpm and high-speed shearing was carried out for 150 s to prepare high internal phase Pickering emulsions (HIPPEs) with an oil phase volume fraction of 75%, namely food 3D printing ink

[0106] Performance characterization:

[0107] 1. Morphology, structure, and rheological properties of HIPPEs stabilized by different substances

[0108] The inventors of this case explored the ability of the HU-PPI-SA complex to stabilize high internal phase emulsions. In addition, to verify the necessity of modifying C-PPI, HIPPEs stabilized by unmodified C-PPI-SA mixture were prepared for control experiments

[0109] (1) Apparent morphology and microstructure of HIPPEs

[0110] First, 1 wt% of commercial PPI (C-PPI) and HU-PPI were used to attempt to prepare high internal phase emulsions with a MCT volume fraction of 75%Figure 4 , the PPI concentration in the figure was fixed at 1%, and the ratio in the figure was the mass ratio of C-PPI to SA. It was found that demulsification and inversion occurred during shearing, indicating that neither single 1 wt% C-PPI nor HU-PPI could stabilize the high internal phase emulsion. Then, C-PPI-SA was used to attempt to prepare high internal phase emulsions under the condition of the proteoglycan concentration ratio from 10:1 to 1:1. It should be noted that C-PPI has poor solubility, and the insoluble protein aggregates exist in the form of precipitation and are difficult to complex with SA, so it is represented by the C-PPI-SA mixture; while the modified HU-PPI exists as soluble small aggregates, and the unfolding of the protein makes it easy to interact with SA, which is represented by the HU-PPI-SA complex. As Figure 4 shown, the C-PPI-SA mixture can stabilize high internal phase emulsions under the condition of the concentration ratio of 5:1 - 1:1, and the samples with the ratio of 4:1 - 1:1 can be self-supporting when inverted; the HU-PPI-SA complex can form HIPPEs under the condition of the concentration ratio of 8:1 - 2:1, and the samples with the ratio of 6:1 - 2:1 can be self-supporting when inverted. The prepared HIPPEs showed a uniform milky white color and could maintain the emulsion stability without stratification for a long time at 4 °C. This shows that, on the one hand, the addition of polysaccharides improves the emulsifying property of C-PPI. However, when the SA content is too high (1:1), the emulsifying property of the HU-PPI-SA complex decreases, which may be because large-sized aggregates are formed, which is not conducive to the stability of the emulsion ( Figure 1a ). On the other hand, the emulsifying property of PPI can also be improved by the combined modification of high-pressure homogenization and ultrasonic treatment, enabling it to stabilize high internal phase emulsions at a lower SA concentration.

[0111] The average particle size of HIPPEs was measured in the experiment ( Figures 5a - 5b , different lowercase letters indicate significant differences (p < 0.05), and the ratio in the figure is the mass ratio of PPI to SA). Compared with the HIPPEs stabilized by the C-PPI-SA mixture (group C), the particle size of the HIPPEs stabilized by the HU-PPI-SA complex (group HU) decreased significantly. Taking the sample with a ratio of 4:1 as an example, the particle size of group HU was about half of that of group C. In addition, the SA concentration also had a significant impact on the particle size of HIPPEs. With the increase of the SA concentration, the particle sizes of both groups of HIPPEs first decreased and then increased. A smaller droplet size is beneficial to improving the stability of HIPPEs. Therefore, when the ratio of HU-PPI:SA is 4:1, the obtained HIPPEs have the best stability.

[0112] Optical microscopy and confocal laser scanning microscopy (CLSM) were used to observe the microstructure of HIPPEs ( Figures 6a - 6bC represents the emulsion stabilized by the C-PPI-SA mixture, and HU represents the emulsion stabilized by the HU-PPI-SA complex (the ratio is the mass ratio of PPI to SA). The HIPPEs stabilized by the C-PPI-SA mixture exhibit a non-uniform droplet morphology under an optical microscope. The particle size of the large droplets exceeds 100 μm, and the change in SA concentration has little effect on its droplet morphology. This may be because the commercial PPI has poor solubility and exists in the form of a suspension itself, and cannot fully react with SA. The HIPPEs stabilized by the HU-PPI-SA complex have dense and uniformly distributed oil droplets, and most of the droplet sizes are less than 50 μm. With the increase in SA concentration, the particle size of the HU group decreases significantly. When the proteoglycan concentration ratio is 2:1, the particle size increases slightly. Due to the existence of smaller droplets and a more compact overall droplet packing, the energy requirement for the emulsion to start flowing will increase. Therefore, the HIPPEs stabilized by the HU-PPI-SA complex have stronger stability.

[0113] As Figure 7 (As shown in the figure, C represents the emulsion stabilized by the C-PPI-SA mixture, and HU represents the emulsion stabilized by the HU-PPI-SA complex (the ratio is the mass ratio of PPI to SA)), the oil-water interface morphology can be observed more clearly under CLSM. The variation law of the HIPPEs droplet size is consistent with the results observed under an optical microscope. The sample emulsion droplets of HU 8:1 are relatively dispersed, and a large amount of protein exists in the dispersed phase. With the increase in SA concentration, the emulsion droplets become denser, and more protein exists at the oil-water interface, improving the stability of the emulsion. The HIPPEs stabilized by the C-PPI-SA mixture have dispersed and non-uniform droplet sizes, and there are many proteins in the continuous phase that have not adsorbed to the oil-water interface, so the system stability is poor.

[0114] (2) Rheological properties of HIPPEs

[0115] Rheological properties are used to evaluate the printing suitability of emulsion samples. The apparent viscosity is related to the extrusion difficulty, while the storage modulus and loss modulus reflect the self-supporting performance of 3D printing materials.

[0116] As Figure 8a and Figure 8b shown, in the entire angular frequency change range (increasing from 0.1 to 100 rad / s), HIPPEs mainly exhibit elastic properties (G′ > G″), indicating that the particles are effectively adsorbed at the oil-water interface to form an elastic gel-like network structure. For the emulsion stabilized by C-PPI-SA, the effect of SA on the elastic modulus of the emulsion does not seem to be obvious ( Figure 8a) This may be because the solubility of C-PPI is low, its dispersibility is poor, and it is unevenly distributed in the system. After adding SA, SA interacts with the uneven PPI particles to form uneven polymers. The loose and sparse complexes between droplets are not conducive to the formation of particle bridges and network structures, resulting in the transformation of the rheological properties of the sample from a solid-like state to a liquid-like state, and G′ and G″ may decrease. On the contrary, with the increase in the proportion of SA in the sample, the elastic modulus of HU-PPI-SA-stabilized HIPPEs increases significantly ( Figure 8b ). This is because HU-PPI can form a uniform protein dispersion and form a uniform complex with SA. As the concentration of SA increases in HU-PPI-SA-stabilized HIPPEs, the droplet size decreases, and the emulsion droplets are packed in a denser manner. The emulsion droplets are more closely distributed, more resistant to deformation, and the fluidity of the emulsion droplets decreases, thus effectively enhancing the gel-like structure of the emulsion. In addition, a high content of SA has more active sites to form strong interactions (including hydrogen bonds and electrostatic interactions) with HU-PPI, which can also strengthen the gel network structure. This gel-like structure can improve the shape stability of HIPPEs and prevent premature collapse of the structure during 3D printing. On the other hand, the large difference between G′ and G″ can enable the high internal phase emulsion to flow stably during the extrusion process.

[0117] The apparent viscosity of HIPPEs gradually decreases as the shear rate increases from 0.1 to 100 s -1 (In the figure, for data clarity, the abscissa intercepts the part from 0.1 - 10 s -1 ) ( Figures 8c - 8d ), showing shear-thinning behavior and belonging to a pseudoplastic fluid. The influence of SA on the apparent viscosity of HIPPEs shows different trends. For C-PPI-SA-stabilized HIPPEs, the initial apparent viscosity is the largest when the C-PPI∶SA concentration ratio is 5∶1 and the smallest when it is 1∶1 (Figure 3-8c); on the contrary, with the increase in the concentration of SA, the initial apparent viscosity of HU-PPI-SA-stabilized HIPPEs gradually increases (Figure 3-8d). This may be because sodium alginate displaces the aqueous solution in the continuous phase, increasing the soluble solid content of the system. However, although the polysaccharide SA itself has a thickening effect, the uneven polymers formed by the mixture of C-PPI and SA may lead to the loosening of the system structure and a decrease in the overall viscosity.

[0118] 2. Mechanisms of stabilizing HIPPEs with different substances

[0119] Based on the above experimental results, the possible mechanisms of stabilizing HIPPEs with C-PPI-SA mixtures and HU-PPI-SA complexes were speculated.

[0120] The most widely used method in the production of commercial PPI is spray drying. However, globulins are prone to denaturation under high-temperature conditions during spray drying, forming insoluble protein aggregates, which greatly limits their application in the food industry. In this application, C-PPI exists in water as a large amount of insoluble large aggregates and a small amount of soluble small aggregates. After combined treatment of high-pressure homogenization and ultrasound, the insoluble protein aggregates dissociate to form soluble aggregates with relatively uniform sizes, and the protein structure unfolds to a certain extent. In addition, 1 wt% of PPI is not sufficient to stabilize high internal phase emulsions, and adding an appropriate proportion of polysaccharide SA can promote the formation of high internal phase Pickering emulsions.

[0121] As Figure 9 shown, C-PPI exists as aggregates of different sizes. Among them, the smaller soluble aggregates can bind to the added SA, while a large number of insoluble large aggregates precipitate at the bottom of the aqueous solution and it is difficult to interact with SA. Therefore, more SA exists in the aqueous solution in a free form. When the mixture of C-PPI and SA stabilizes oil droplets, it mainly relies on the adsorption of C-PPI at the oil-water interface and the thickening effect of SA in the continuous phase, while the adsorption of SA at the interface is less. Due to the different sizes of C-PPI aggregate particles, the particle size distribution of the stabilized oil droplets is also relatively wide. The modified HU-PPI is uniformly dispersed in the aqueous solution as soluble aggregates. Due to its reduced particle size and unfolded protein structure, the negatively charged SA can undergo electrostatic interactions with the positively charged groups on the surface of HU-PPI, and can also form a HU-PPI-SA complex through hydrogen bonding and hydrophobic interactions. When the HU-PPI-SA complex stabilizes oil droplets, it mainly relies on the adsorption of protein-polysaccharide composite particles at the oil-water interface and the thickening effect of SA. Compared with C-PPI, the average particle size of the soluble aggregates of HU-PPI is smaller, the particle size distribution is concentrated, and it can still maintain a uniform particle size after being combined with polysaccharides. Therefore, the stabilized oil droplets also have a relatively uniform particle size distribution, the voids between droplets are smaller, and a denser structure can be formed, improving the stability, storage modulus and viscosity.

[0122] It should be noted that the concentration of SA has an important impact on the formation and stability of HIPPEs. The C-PPI-SA mixture can stabilize HIPPEs when the proteoglycan concentration ratio is 5:1 - 1:1, while the HU-PPI-SA complex can stabilize HIPPEs when the proteoglycan concentration ratio is 8:1 - 2:1. On the one hand, compared with HU-PPI, due to the difference in aggregate size, the gaps between the O / W droplets formed by C-PPI are larger. Therefore, a higher concentration of SA is required to fill and thicken between the droplets to form a stable high internal phase emulsion. In addition, compared with the adsorption of the HU-PPI-SA complex at the oil-water interface, the interfacial adsorption ability mainly composed of C-PPI and supplemented by a small amount of C-PPI-SA is weaker. Therefore, more SA is needed to form a spatial barrier in the aqueous phase system to stabilize the oil droplets. On the other hand, when HU-PPI:SA is 8:1, both HU-PPI and the HU-PPI-SA complex exist in the system, and they jointly stabilize the oil-water interface; as the concentration of SA increases, the proportion of the HU-PPI-SA complex in the system also increases; when the SA concentration increases to 0.5 wt%, that is, when HU-PPI:SA decreases to 2:1, the proteins in the system basically exist in the form of the HU-PPI-SA complex, and SA is in excess. A part of the excess SA can thicken between the droplets to further stabilize the high internal phase emulsion, while the other part will form larger protein-polysaccharide aggregates with HU-PPI; when the SA concentration further increases to 1 wt%, the average particle size of the HU-PPI-SA complex aggregates reaches 1 μm. At this time, the too large particles are not conducive to the formation and stability of the emulsion, which explains why the high internal phase emulsion cannot be stabilized when HU-PPI:SA is 1:1. Most of the proteins in C-PPI exist as insoluble aggregates and are difficult to interact with SA. Even when the SA concentration reaches 1 wt%, large protein-polysaccharide aggregates will not be formed, and HIPPEs can still be formed. However, the poor rheological properties at this time also indicate that SA in the system is in excess and is not conducive to the stability of HIPPEs.

[0123] 3. 3D printing properties of HIPPEs

[0124] (1) Line morphology of HIPPEs

[0125] Before being used for food 3D printing, the extrusion line morphology of the HIPPEs emulsion was observed. As Figure 10As shown, two groups of samples with a proteoglycan concentration of 4:1 were selected. After being loaded into a syringe, different English letters and symbols were manually extruded. First, observe the extruded letter "NWAFU": After the 4:1 sample in group C was extruded, the gaps between the strokes merged and converged, making the letter difficult to distinguish; while after the 4:1 sample in group HU was extruded, the original line shape was maintained and the letter was clearly distinguishable. Then, a larger letter "W" and symbol "#" were extruded to observe the side shape of the line: The side of the line of the 4:1 sample in group C collapsed, and the intersection of the "plus" sign merged into a point, and oil droplets were observed to precipitate; the line of the 4:1 sample in group HU maintained a smooth three-dimensional structure, and an obvious two-layer building effect was observed at the intersection of the "plus" sign, with a clear boundary between the layers and no oil separation phenomenon. Therefore, HU-PPI-SA-stabilized HIPPEs have the potential for food 3D printing.

[0126] (2) 3D printing performance of HIPPEs

[0127] To verify the feasibility of the prepared HIPPEs for food 3D printing, the HIPPEs were printed into three different three-dimensional patterns. The results showed that HU-PPI-SA complex-stabilized HIPPEs have the potential for 3D printing and can successfully print out pentagram, four-leaf clover, and snowflake patterns; while the HIPPEs stabilized by the C-PPI-SA mixture could not maintain stability after being extruded through the nozzle, internal oil droplets began to precipitate, the lines merged, and the structure collapsed, and the printing of simple 3D patterns could not be completed ( Figure 10 ).

[0128] Ideal HIPPEs for 3D printing should be able to maintain the printed shape with high fidelity, be smoothly extruded during the printing process without compression deformation, and be able to adhere to the previously printed deposition layer, which is closely related to the rheological properties of HIPPEs. The storage modulus G′ is used to reflect the mechanical strength of the material and is crucial for the support and structure maintenance of the previously printed deposition layer. A higher G′ means an improvement in printing accuracy and deposition layer resolution, which is beneficial to the stability of the structure and shape retention. In addition, appropriate shear thinning behavior is beneficial to the extrusion of the material from a fine-diameter printing nozzle and the deposition during 3D printing. This requires the sample viscosity to be high enough to prevent convergence with the previously deposited layer, and the viscosity should not be too high to allow HIPPEs to be easily extruded through a small nozzle.

[0129] As Figure 11As shown, the integrity of the printed shape and the clarity of the printed layers reflect the printability of HIPPEs. For the 8:1 samples in the HU group, the bottom of the printed pentagrams and snowflakes of HIPPEs had a tendency to spread outwards, the internal lines of the four-leaf clovers were somewhat collapsed, and there was some wire drawing at the tight lines. In contrast, the samples in the HU group of 4:1 and the HU group of 2:1 had continuous filaments, showing a smooth surface, clear edges and precise geometries, indicating that the HIPPEs ink had good printing stability. Combining with the rheological properties of HIPPEs, mainly the HU-PPI-SA complex endows HIPPEs with excellent mechanical strength, viscoelasticity and self-supporting properties, making the stable HIPPEs perform well in 3D printing. In addition, the increase in SA concentration led to an increase in the G′ and viscosity of HIPPEs, and the printing resolution of 3D printing was also better. Therefore, the HIPPEs stabilized by the HU-PPI-SA complex can produce 3D printed foods with simple structures to meet the nutritional needs and sensory experiences of consumers.

[0130] To evaluate the stability of the 3D printed sample structure, the printed model was stored at 4 °C. As Figure 12a shown, after printing for 24 h, the sample structure was basically maintained intact, and there seemed to be a small amount of oil layer precipitated at the bottom. As Figure 12b shown, after printing for 72 h, the sample structure collapsed to varying degrees, the side edges became blurred, the bottom area expanded, and the height decreased slightly. This may be because cohesion began to occur between adjacent deposited layers, and partial rupture of the bottom emulsion led to the precipitation of the internal oil phase. Among them, the samples in the HU group of 4:1 and the HU group of 2:1 had relatively distinct edges and corners and had strong resistance to structural collapse, which was related to their strong rheological properties.

[0131] In summary, an appropriate ratio of PPI-SA can stabilize high internal phase emulsions (oil phase volume fraction of 75%) at a low protein concentration of 1.0 wt%. Among them, the HU-PPI treated by high-pressure homogenization and ultrasound combined has better emulsifying properties and can stabilize HIPPEs in the presence of a lower concentration of SA. Moreover, the HIPPEs stabilized by the HU-PPI-SA complex has a smaller particle size, a more uniform and compact microstructure, a stronger storage modulus and apparent viscosity compared with those stabilized by the C-PPI-SA mixture. It is more suitable as an ink for food 3D printing. The 3D printing model based on the HIPPEs stabilized by the HU-PPI-SA complex has a smooth and regular shape, a stable three-dimensional structure, and fine resolution, and can maintain the structural stability within 24 h, indicating that the HIPPEs prepared in this application can be used as an ink for food 3D printing.

[0132] In addition, the inventors of this case also referred to the foregoing embodiments and conducted tests with other raw materials, process operations, and process conditions described in this specification, and all obtained relatively ideal results.

[0133] It should be understood that the technical solution of the present invention is not limited to the limitations of the above specific embodiments. Any technical deformation made according to the technical solution of the present invention without departing from the gist of the present invention and the scope protected by the claims falls within the protection scope of the present invention.

Claims

1. A preparation method of a 3D printing ink for food, characterized in that, Comprising: Subjecting a pea protein dispersion to high-pressure homogenization and ultrasonic treatment in sequence to obtain a modified pea protein; wherein, the content of pea protein in the pea protein dispersion is 0.5 - 1.5 wt%; the pressure used for the high-pressure homogenization treatment is 30 - 70 MPa, and the number of cycles is 3 - 5 times; the power used for the ultrasonic treatment is 200 - 600 W, and the treatment time is 15 - 25 min; Fully mixing and reacting sodium alginate, the modified pea protein and water to obtain a modified pea protein / sodium alginate complex; wherein, the mass ratio of the modified pea protein to sodium alginate is 8:1 - 2:1; And, mixing medium-chain triglycerides with a solution containing the modified pea protein / sodium alginate complex and subjecting it to shearing treatment to obtain a food 3D printing ink, wherein, the volume ratio of the medium-chain triglycerides to the modified pea protein / sodium alginate complex solution is 3:1 - 5:1, and the content of the modified pea protein / sodium alginate complex in the food 3D printing ink is 0.3 wt% - 1.0 wt%.

2. The preparation method according to claim 1, wherein Specifically including: Fully stirring and mixing a dispersion containing a modified pea protein with a sodium alginate solution to react to obtain a modified pea protein / sodium alginate complex; the modified pea protein and sodium alginate are mainly combined through electrostatic interaction and hydrogen bond; wherein, the mass ratio of the modified pea protein to sodium alginate is 6:1 - 2:

1.

3. The preparation method according to claim 1, wherein Specifically including: Using a high-speed shear dispersion device and under the condition of a rotation speed of 3000 - 6000 rpm, dropping medium-chain triglycerides into a modified pea protein / sodium alginate complex solution and mixing evenly, and then performing high-speed shearing for 100 - 200 s under the condition of a rotation speed of 8000 - 12000 rpm to obtain a food 3D printing ink; Wherein, the content of the modified pea protein / sodium alginate complex in the modified pea protein / sodium alginate complex solution is 1.0 wt% - 3.0 wt%.

4. A food 3D printing ink prepared by the preparation method according to any one of claims 1 - 3.

5. A 3D printing method, characterized in that, Comprising: Providing the food 3D printing ink according to claim 4; And, using a nozzle extrusion 3D printing method to print the food 3D printing ink to obtain a food 3D printed part.

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