Full-nutrient fresh algae gel for 3D printing and preparation method of full-nutrient fresh algae gel

Abstract

The invention provides a preparation method of full-nutrient fresh algae gel for 3D printing, which comprises the following steps: mixing and dissolving starch, a rheological property changing agent, nutritive salt required by microalgae growth and boiling water, adding microalgae cells, and uniformly stirring to form bio-ink for 3D printing. The full-nutrition fresh algae gel not only can support efficient culture of microalgae, but also can be directly used as a full-nutrition fresh algae gel functional food after being subjected to microalgae culture due to the full-food-grade raw materials of the full-nutrition fresh algae gel.

Description

Technical Field

[0001] This invention relates to the field of microalgae cultivation technology, and more specifically to a 3D-printed fully nutritious fresh algae gel and its preparation method. Background Technology

[0002] Microalgae are typically cultured in liquid form, and after cultivation, they need to be collected through centrifugation, filtration, flocculation, and drying for further applications. These collection methods inevitably lead to the loss or decomposition of microalgal nutrients, including sulfonic acid polysaccharides and lipids. Furthermore, the novel sodium alginate-based hydrogel scaffold for microalgae cultivation requires post-cultivation hydrogel scaffold disintegration, microalgae collection, and nutrient extraction, increasing production steps and costs.

[0003] In the food industry, edible microalgae are primarily used in the form of algae powder. This involves drying the microalgae into powder or flakes. While this form is easy to store and transport, traditional drying methods can lead to nutrient loss, deterioration of aroma and flavor, and low efficiency. Furthermore, microalgae extracts suffer from incomplete extraction due to the complex cell structure and cell walls that hinder extraction. Traditional extraction methods are also inefficient and energy-intensive.

[0004] Fresh microalgae, without drying or other processing, retains its rich vitamins and minerals to the greatest extent possible. For example, antioxidant vitamins such as vitamin C and vitamin E in fresh microalgae maintain high activity without high-temperature drying. Vitamin C has antioxidant properties, promotes collagen synthesis, enhances immunity, and aids in iron absorption. Regarding minerals, calcium, iron, and zinc in fresh microalgae exist in their natural bound forms, making them easier for the body to absorb.

[0005] Many microalgae are rich in high-quality protein, such as Chlorella and Spirulina. In their fresh state, the tertiary and quaternary structures of these proteins remain intact, making them easier for human digestive enzymes to break down into amino acids, which are then absorbed by the body to synthesize the proteins it needs. However, during the drying process of microalgae powder, the proteins may denature. Although their amino acid composition remains unchanged, this may affect the efficiency with which digestive enzymes bind to them, thus impacting the efficiency of protein digestion and absorption.

[0006] However, since edible microalgae are generally cultured in liquid form, they require collection after cultivation through centrifugation, filtration, flocculation, and drying for subsequent applications. These collection methods inevitably lead to the loss or decomposition of microalgal nutrients, including sulfonic acid polysaccharides and lipids. Furthermore, the use of novel sodium alginate-based hydrogel scaffolds for microalgae cultivation requires post-cultivation dissociation of the hydrogel scaffold, collection of the microalgae, and nutrient extraction, increasing production steps and costs. Moreover, even if the sodium alginate-based hydrogel scaffold is used entirely for food or pharmaceutical purposes, excessive sodium alginate intake can cause gastrointestinal discomfort, thus limiting its application in these fields. Summary of the Invention

[0007] In view of this, and to address the problems of the prior art, the present invention aims to provide a 3D-printed fully nutritious fresh algae gel and its preparation method. Specifically, it includes the efficient cultivation of microalgae in a gel state, and its direct use as an easily swallowed, fully nutritious fresh algae gel food after microalgae cultivation.

[0008] To achieve the above objectives, this invention provides a 3D-printed fully nutritious fresh algae gel and its preparation method, the method comprising the following steps:

[0009] Step 1: Dissolve BG11 powder in water, add microalgae to obtain microalgae culture medium, centrifuge a portion of the microalgae culture medium, and collect the precipitate for later use.

[0010] Step 2: Mix and dissolve starch, rheology modifier, solid culture medium required for microalgae growth and boiling water, then transfer the mixture to a cold water bath and stir to cool to room temperature; add the microalgae precipitate collected in Step 1 and stir evenly, then refine to form bio-ink;

[0011] Step 3: 3D print the bio-ink to prepare a starch-based hydrogel scaffold.

[0012] Step 4: Add the nutrients required by the microalgae to the hydrogel scaffold and culture the microalgae starch-based hydrogel in a high humidity environment (>70%);

[0013] In one embodiment of the present invention, the microalgae cells in step 1 are OD cells. 680 The precipitate formed after centrifugation of microalgae culture medium with a concentration of 0.4-3.0.

[0014] In one embodiment of the present invention, the rheological property modifier in step 2 is selected from carrageenan, pectin or xanthan gum;

[0015] In one embodiment of the present invention, the solid culture medium in step 2 is an inorganic salt;

[0016] In one embodiment of the present invention, in step 2, 25-50 mg of the starch is dissolved per milliliter of water, 10-20 mg of the rheological modifier and 1-10 mg of nutrients required for microalgae growth are added, and 0.2-2 ml of microalgae culture medium is used.

[0017] In one embodiment of the present invention, the rheological property modifier mentioned in step 2 is selected from food-grade thickeners, such as colloids (pectin, xanthan gum, carrageenan) and modified starches (acetic acid starch, sodium starch phosphate, phosphorylated distarch phosphate).

[0018] In one embodiment of the present invention, the printing parameters in step 3 are: layer height: 0.27mm; number of layers: 12; printing fill, printing outline, and starting point are random; model: quadrilateral; speed: 8mm / s. -1 Broken wire elevation: 1mm; Air pressure: 0.3MPa; Early wire exit: 380ms; Early wire closing: 0.2mm; Filling mode: linear filling; Filling method: linear filling; Filling offset: 0.7mm; Filling spacing: 1.4mm; Rotation angle: 90°; Number of rotations: 2.

[0019] In one embodiment of the present invention, the cultivation conditions in step 4 include: a temperature of 22-25°C, a humidity of 70-90%, and a light intensity of 20-50 μmol photons / m². -2 s -1 .

[0020] In one embodiment of the present invention, the nutritionally complete fresh algae gel prepared according to the above method is directly used as a nutritionally complete food.

[0021] Compared to existing technologies, this invention has the following advantages: This invention uses specific substances to form bio-ink and then prints it to create a fully nutritious fresh algae gel. Because this fully nutritious fresh algae gel uses only easily swallowed and digestible food-grade raw materials, it can be directly used as a complete nutritional food after cultivation. This fully nutritious fresh algae gel can retain the proteins, lipids, and carbohydrates in microalgae to the greatest extent possible. Attached Figure Description

[0022] Figure 1 Fluorescence microscope image of the 3D-printed fully nutrient-rich fresh algae gel prepared in Example 1;

[0023] Figure 2 Fluorescence microscope image of the 3D-printed fully nutrient-rich fresh algae gel prepared in Example 1;

[0024] Figure 3 A bright-field optical microscope for a 3D-printed, fully nutrient-rich fresh algae gel prepared in Example 1;

[0025] Figure 4 A comparison diagram of the activity of the 3D-printed fully nutritious fresh algae gel prepared in Example 1;

[0026] Figure 5 The scaffold morphology of the 3D-printed fully nutritious fresh algae gel prepared in Example 1. Detailed Implementation

[0027] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0028] It should be noted that, unless otherwise defined, all terms (including technical and scientific terms) used in the embodiments of this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. It should also be understood that terms such as those defined in a common dictionary should be interpreted as having a meaning consistent with their meaning in the context of the relevant art, and not as being interpreted in an idealized or highly formalized sense, unless expressly defined herein.

[0029] This invention provides a 3D-printed fully nutritious fresh algae gel and its preparation method, comprising:

[0030] (1) Microalgae culture medium for forming Chlorella CP;

[0031] The microalgae culture medium for *Chlorella pyrenoidosa* (CP) was prepared by adding BG11 powder (Qingdao High-tech Industrial Park Haibo Biotechnology Co., Ltd.) at a concentration of 1.7 g / L. -1 The concentration is obtained by dissolving in water and then sterilizing it.

[0032] (2) Formation of bio-ink;

[0033] Add 30g starch, 9g carrageenan, and 2.04g BG11 powder to 600ml of boiling water and mix until the starch and carrageenan are completely dissolved. Then transfer the mixture to a cold water bath and stir to cool to room temperature. Next, at OD... 680 =0.8, take 600 ml of microalgae culture medium from (1), centrifuge at 5000 rpm for 3 minutes, then collect the precipitate, add the precipitate to an aqueous solution of starch and carrageenan, and let the mixture stand for 10 minutes.

[0034] (3) 3D printing;

[0035] The bio-ink is transferred into a syringe and loaded into a 3D printer. (SR, Regenovo, Hangzhou, China) A fully nutrient-rich fresh algae gel with a mesh structure was formed. The printer software allowed for editing of the microalgae gel scaffold structure, and the diameter of the monofilaments could be adjusted by switching needles. In summary, the entire process required a sterile environment to ensure the success of 3D bioprinting. The printing parameters were: layer height: 0.27 mm; number of layers: 12; randomized printing infill, printing outline, and starting point; model: quadrilateral; printing speed: 8 mm / s. -1 Broken wire elevation: 1mm; Air pressure: 0.3MPa; Early wire exit: 380ms; Early wire closing: 0.2mm; Filling mode: linear filling; Filling method: linear filling; Filling offset: 0.7mm; Filling spacing: 1.4mm; Rotation angle: 90°; Number of rotations: 2.

[0036] (4) Cultivation

[0037] The 3D-printed all-nutrient fresh algae gel (5cm×5cm×1cm) was placed directly onto a blank culture dish (90mm in diameter) and placed in an incubator for static cultivation at 23℃, 70% relative humidity, and 20μmol photonsm. -2 s -1 .

[0038] Example 1: Performance Modulation of Hydrogel Scaffolds

[0039] 3D-printable hydrogel scaffold structures can be prepared using food-grade starch and carrageenan. The overall performance of the hydrogel scaffold can be optimized by adjusting the ratio of starch to carrageenan. Using too little or too much starch and carrageenan can result in hydrogel scaffold materials that are either too soft (fluid) or too hard (granular), as shown in the following figures. Figure 1 The results showed that the optimal concentration was 25-50 mg of starch dissolved per milliliter of water, with 10-20 mg of carrageenan added accordingly.

[0040] Example 2: Cultivation of microalgae in gel

[0041] A piece of fully nutritious fresh algae gel (5cm×5cm×1cm) on day 0 contained a small number of live microalgae cells and was pale green in color. (See attached image.) Figure 2 After seven days of cultivation, the microalgal gel turned dark green, indicating the enrichment of chlorophyll in the microalgae. Fluorescence microscopy ( Figure 2 This demonstrates that after 7 days of cultivation, the biomass of microalgal cells in the gel was significantly increased. (Bright-field optical microscopy) Figure 3 This demonstrates that after 7 days of cultivation, the appearance and activity of the microalgae in the gel remained unchanged.

[0042] Detection of the activity of microalgal starch hydrogel in Example 3

[0043] In traditional liquid microalgae culture, after cultivation, the microalgae need to be concentrated using techniques such as centrifugation to extract nutrients. However, microalgae are highly susceptible to death in high-concentration solutions, leading to nutrient loss. Figure 4 As shown, the activity of the initial algal solution was over 90% (initial). However, microalgae in high-concentration liquid algal solutions (100g L) showed significantly reduced activity. -1 After being kept in a liquid state for 7 days, the activity of microalgae decreased to about 45%. However, after being kept in a starch-based gel for 7 days (solid state), the activity of microalgae did not change significantly, indicating that microalgae starch hydrogels can maximize the preservation of microalgae activity and nutrients in their cells.

[0044] Detection Example 4

[0045] The optimized hydrogel scaffold structure in Example 1 can be fabricated using 3D printing, combining multiple 3D models to obtain various hydrogel scaffolds with special morphologies. Figure 5 These hydrogel scaffolds enable efficient gel-state cultivation of microalgae. Furthermore, the aforementioned various uniquely morphological, fully nutritious fresh algae gels, after 3D printing, offer convenient packaging and consumer acceptability, avoiding nutrient loss caused by traditional liquid cultivation, collection, and drying processes. They can be directly used in functional foods made from fully nutritious fresh algae gels.

[0046] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for preparing a 3D-printed, fully nutritious fresh algae gel, characterized in that, The method includes: Step 1: Dissolve BG11 powder in water, add microalgae to obtain microalgae culture medium, centrifuge a portion of the microalgae culture medium, and collect the precipitate for later use. Step 2: Mix and dissolve starch, rheology modifier, solid culture medium required for microalgae growth and boiling water, then transfer the mixture to a cold water bath and stir to cool to room temperature; add the microalgae precipitate collected in Step 1 and stir evenly, then refine to form bio-ink; Step 3: 3D print the bio-ink to prepare a starch-based hydrogel scaffold. Step 4: Add the nutrients required by the microalgae to the hydrogel scaffold and culture the microalgae starch-based hydrogel in a high humidity environment (>70%).

2. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The microalgal cells mentioned in step 1 are OD 680 The precipitate formed after centrifugation of microalgae culture medium with a concentration of 0.4-3.

0.

3. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The rheological property modifier mentioned in step 2 is selected from carrageenan, pectin, and xanthan gum.

4. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The solid culture medium mentioned in step 2 is an inorganic salt.

5. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: In step 2, each milliliter of water corresponds to the dissolution of 25-50 mg of the starch, the addition of 10-20 mg of the rheological modifier and 1-10 mg of nutrients required for microalgae growth, and the use of 0.2-2 ml of microalgae culture medium.

6. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The rheological modifier mentioned in step 2 is selected from food-grade thickeners and modified starch.

7. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The rheological property modifier mentioned in step 2 is selected from one or more of pectin, xanthan gum, carrageenan, starch acetate, sodium starch phosphate, and phosphorylated distarch phosphate.

8. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The printing parameters mentioned in step 3 are: layer height: 0.27mm; number of layers: 12; printing fill, printing outline, and starting point are random; model: quadrilateral; Speed: 8mm / s -1 Broken wire elevation: 1mm; air pressure 0.3MPa; Early fiber emergence: 380ms; Pre-closing wire: 0.2mm; Fill mode: Linear fill; Fill method: Linear fill; Fill offset: 0.7mm; Fill spacing: 1.4mm; Rotation angle: 90°; Number of rotations:

2.

9. The method for preparing 3D-printed fully nutritious fresh algae gel as described in claim 1, characterized in that: The cultivation conditions in step 4 include: temperature of 22-25℃, humidity of 70-90%, and light intensity of 20-50 μmol photons / m². -2 s -1 .

10. The 3D-printed fully nutritious fresh algae gel prepared by the method according to any one of claims 1-8, characterized in that: The fully nutritious fresh algae gel prepared according to the above method can be directly used as a fully nutritious food.

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