Bioink for 3D printing, the preparation method and usage

Abstract

The present invention relates to a bioink for 3D printing, the preparation method, and the usage. Such bioink is a gel made of -zein, porogen by 0-10% of the weight of zein, ethanol and water. The preparation method consists of: 10-50% zein is dissolved into an aqueous solution containing 40-90% (v/v) of ethanol, then, the porogen by 0-10% of the weight of zein is added, and then, this solution is allowed to stand at 5-95 C. for 1-10 days, or stirred for 30 min-24 hours, and thus, the bioink for 3D printing can be obtained. The conditions applied to prepare such bioink are mild and the method adopted is easy to operate, in addition, the said bioink for 3D printing has good mechanical properties and biocompatibility, which can be applied in the field of biomedicine for preparing tissue engineered substitutes and hemostatic materials by 3D printing at room temperature.

Claims

1. A bioink for 3D printing, wherein the bioink is a shear-thinning gel consisting of -zein, ethanol, water, and porogen.

2. The bioink for 3D printing according to claim 1, wherein the gel contains -zein in an amount of 10-50% by weight, and porogen in an amount of 0-10% of the weight of zein, and a remainder of the gel is made of the ethanol and the water; a volume ratio of ethanol to water is 40-90:10-60; and the porogen is a water-soluble salt selected from the group consisting of: mannitol, ammonium bicarbonate, sodium chloride, sodium tartrate, sodium citrate, etc., sugar particles, paraffin particles and ice particles.

3. The bioink for 3D printing according to claim 2, wherein the gel includes the -zein in an amount of 30-50% by weight.

4. The bioink for 3D printing according to claim 2, wherein the volume ratio of ethanol to water is 60-85:15-40.

5. The bioink for 3D printing according to claim 1, wherein the bioink can be printed at room temperature.

6. The bioink for 3D printing according to claim 1, wherein a zero-shear viscosity of the bioink is 30-5000 Pa.Math.s.

7. A preparation method of the bioink for 3D printing according to claim 1, the method comprising: dissolving 10-50% -zein into a mixed solution of ethanol and water; allowing the mixed solution of ethanol and water with the 10-50% -zein dissolved therein to stand at 5-95 C. for 1-10 days, or stirring the mixed solution of ethanol and water with the zein 10-50% -zein dissolved therein for 30 min-24 hours to form the gel; and adding porogen in an amount of 0-10% of the weight of -zein to the gel, wherein, a volume ratio of ethanol to water is 40-90:10-60.

8. The preparation method of the bioink for 3D printing according to claim 7, wherein the mixed solution of ethanol and water with the 10-50% -zein dissolved therein is allowed to stand at 10 C.-50 C. for 1 to 3 days.

9. The bioink for 3D printing according to claim 1, wherein the bioink is used to prepare biomedical materials.

10. The bioink for 3D printing according to claim 9, wherein the biomedical materials are tissue engineered substitutes or hemostatic materials with tubular, membraniform, or 3D scaffold structures.

11. The bioink for 3D printing according to claim 10, wherein the tissue engineered substitutes consist of nerve conduit, artificial windpipe, artificial urethra, artificial blood vessel, artificial skin, hemostatic membrane materials, artificial bone, artificial cartilage and artificial ear.

12. The bioink for 3D printing according to claim 9, wherein the biomedical materials are materials used to stop bleeding resulting from a deep penetrating wound with irregular wound directly by means of injection or smearing, or to stop bleeding.

13. The preparation method of the bioink for 3D printing according to claim 7, wherein dissolving the 0-50% zein into the mixed solution of ethanol and water includes dissolving -zein in an amount of 10-50% by weight, and porogen in an amount of 0-10% of the weight of zein, with a remainder made of the ethanol and the water measured; and the porogen is a water-soluble salt selected from the group consisting of: mannitol, ammonium bicarbonate, sodium chloride, sodium tartrate, sodium citrate, etc., sugar particles, paraffin particles and ice particles.

14. The bioink for 3D printing according to claim 1, wherein the bioink is prepared by a method comprising: (a) dissolving 10-50% -zein into a mixed solution of ethanol and water with a volume ratio of 40-90:10-60; (b) allowing the mixed solution to stand at 5-95 C. for 1-10 days, or stirring the mixed solution for 30 min-24 hours to form the gel; and (c) adding porogen in an amount of 0-10% of the weight of -zein.

15. The bioink for 3D printing according to claim 14, wherein the mixed solution of ethanol and water with the 10-50% -zein dissolved therein is allowed to stand at 10 C.-50 C. for 1 to 3 days.

16. The bioink for 3D printing according to claim 1, wherein the bioink is configured for use in preparing biomedical materials selected from: tissue engineered substitutes, hemostatic materials, nerve conduits, artificial blood vessels, and artificial skin.

17. A method of treating a wound, comprising applying the bioink of claim 1 to a deep penetrating wound with irregular shape by injection or smearing.

Description

BRIEF DESCRIPTION OF THE FIGURES

(1) FIG. 1: The gelatinous bioink of -zein for 3D printing prepared in Embodiment 2.

(2) FIG. 2: The tubular tissue engineered product produced by 3D printing with the bioink of -zein prepared in Embodiment 3.

(3) FIG. 3: The mesh-type hemostatic membrane material produced by 3D printing with the bioink of -zein prepared in Embodiment 4, wherein, the part in black color indicates the pore structure of the mesh.

(4) FIG. 4: The diagram of hemostatic effect of the said hemostatic membrane material of Embodiment 4 on dermatorrhagia.

(5) FIG. 5: The 3D products in different shapes produced by 3D printing with the bioink of -zein prepared in Embodiment 7.

(6) FIG. 6: The tissue engineered artificial ear containing cells produced by 3D printing with the bioink of -zein prepared in Embodiment 8. Wherein, the artificial ear is in gel state when it is just printed out (left picture), and the picture on the right shows the artificial ear after solidifying.

(7) FIG. 7: The diagram of hemostatic effect of the said porous hemostatic material in Embodiment 7 on the model of penetrating injury in collaboration with the gel material of -zein prepared in Embodiment 9.

DESCRIPTION OF THE INVENTION

(8) The text below will describe the present invention in detail in conjunction with the following embodiments, which can only be used to further illustrate the present invention rather than being understood as imposing restrictions on the scope of protection of the present invention. For those skilled in the art can make various variations or modifications to the present invention after reading the content of the present invention, and the equivalent forms of which also shall fall within the scope of the attached claims of the present application.

Embodiment 1

(9) A bioink for 3D printing, which is made from the material according to following parts by mass: 10 g of -zein.

(10) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(11) , -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 40%, and then, the milky white solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is kept at 10 C. for 10 days for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 28 Pa.Math.s.

Embodiment 2

(12) A bioink for 3D printing, which is made from the material according to following parts by mass: 20 g of -zein.

(13) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(14) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 65%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is kept at 20 C. for one days for completing reaction, and thus, the bioink for 3D printing can be obtained, and it is gelatinous as shown in FIG. 1. Wherein, the zero-shear viscosity of the bioink is about 120 Pa.Math.s.

Embodiment 3

(15) A bioink for 3D printing, which is made from the material according to following parts by mass: 30 g of -zein.

(16) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(17) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 70%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is kept at 30 C. for two days for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 389 Pa.Math.s.

(18) The bioink prepared in this Embodiment is mixed with mannitol particles, which is used as porogen (accounts for 6% of the mass of zein, w/w), and then, loaded into the print nozzle. After that, the tubular products are printed through the print needle with a diameter of 0.5 mm, and under the conditions that the temperature is at 10 C., the printing speed is 8 mm/s and the printing pressure is 150 kPa. Then, the porous tubular product would be obtained after removing porogen by means of the water-boiling method. As shown in FIG. 2, the product can be used as a tissue engineered nerve conduit or for other purposes.

Embodiment 4

(19) A bioink for 3D printing, which is made from the material according to following parts by mass: 40 g of -zein.

(20) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(21) The -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 80%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is kept at 40 C. for 4 days for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 1832 Pa.Math.s. The mesh-type membrane product can be printed at the following conditions: the diameter of printing needle is 0.33 mm, the temperature is at 20 C., the printing speed is 8 mm/s and the printing pressure is 80 kPa. As shown in FIG. 3, the product can be used as a hemostatic membrane material. In addition, FIG. 4 is the diagram of hemostatic effect of the hemostatic membrane material on dermatorrhagia occurred on the body of rat.

Embodiment 5

(22) A bioink for 3D printing, which is made from the material according to following parts by mass: 50 g of -zein.

(23) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(24) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 85%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is kept at 50 C. for 8 days for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 1623 Pa.Math.s.

Embodiment 6

(25) A bioink for 3D printing, which is made from the material according to following parts by mass: 20 g of -zein.

(26) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(27) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 90%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is stirred at 10 C. for 12 hours for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 55 Pa.Math.s.

Embodiment 7

(28) A bioink for 3D printing, which is made from the material according to following parts by mass: 30 g of -zein.

(29) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(30) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 65%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is stirred at 20 C. for 30 min for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 344 Pa.Math.s. The tissue engineered substitutes can be printed at the following conditions: the diameter of printing needle is 0.5 mm, the temperature is at 50 C., the printing speed is 8 mm/s and the printing pressure is 200 kPa. FIG. 5 shows the 3D hemostatic materials with various sizes printed by the bioink of this embodiment. It can be seen that the bioink made from zein is suitable for printing 3D scaffold products. Wherein, the compressive strength of scaffold is 3.341.25 MPa, the compressive modulus is 63.9319.22 MPa; in addition, the bending strength is 5.301.32 MPa, the bending modulus is 52.0711.07 MPa; the tensile strength is 0.110.02 MPa, and the tensile modulus is 2.631.25 MPa.

Embodiment 8

(31) A bioink for 3D printing, which is made from the material according to following parts by mass: 30 g of -zein.

(32) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(33) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 85%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is stirred at 30 C. for 6 hours for completing reaction, and thus, the bioink for 3D printing can be obtained, wherein, the zero-shear viscosity of the bioink is about 551 Pa.Math.s.

(34) Then, the aforesaid bioink for 3D printing is loaded in the first print nozzle, and the alginate sodium mixed with cells (cell density is 1*106/mL) is loaded in the second print nozzle. After that, the tissue engineered organs can be printed at the following conditions: the diameter of printing needle is 0.33 mm, the temperature is at 37 C., the printing speed is 8 mm/s and the printing pressure is 80 kPa. FIG. 6 shows the printed artificial ear. Furthermore, such bioink be used alone or mixed with cells or other bioink to print out tissue engineered substitutes in irregular shapes by means of 3D printing.

Embodiment 9

(35) A bioink for 3D printing, which is made from the material according to following parts by mass: 50 g of -zein.

(36) The preparation method of the bioink for 3D printing in this embodiment consists of following steps:

(37) -zein is dissolved into 100 mL of the ethanol-aqueous solution, in which the volume fraction of ethanol is 90%, and then, the transparent solution is obtained after ultrasonic treatment at room temperature for 10 min. After that, this solution is stirred at 50 C. for 4 hours for completing reaction, and thus, the gel bioink can be obtained, wherein, the zero-shear viscosity of the bioink is about 3476 Pa.Math.s. Furthermore, the product can be used to stop bleeding resulted from deep penetrating wound with irregular wound directly by means of injection or smearing, or to stop bleeding in collaboration with the 3D printing products (FIG. 7).