# US 11,439,727 B2 · 3D printable bio gel and method of use

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- Number: US 11,439,727 B2
- Document ID: US11439727B2
- Status: Granted
- Kind: B2
- Application number: 16/304,212
- Filed: 2017-05-25
- Published: 2020-10-08
- Granted: 2022-09-13
- Earliest claimed: 2016-05-26
- Assignees: TDBT IP Inc.
- Applicants: Tdbt IP Inc.
- Inventors: Daniel L. Cohen; Chris Pelletier
- Web page: https://patentgrep.com/patent/US11439727B2

## CPC

- A61L 27/24: Collagen
- A61L 27/3804: {characterised by specific cells or progenitors thereof, e.g. fibroblasts, connective tissue cells, kidney cells}
- A61L 27/3808: {Endothelial cells}
- A61L 27/3813: {Epithelial cells, e.g. keratinocytes, urothelial cells}
- A61L 27/3817: {Cartilage-forming cells, e.g. pre-chondrocytes}
- A61L 27/3821: {Bone-forming cells, e.g. osteoblasts, osteocytes, osteoprogenitor cells}
- A61L 27/3826: {Muscle cells, e.g. smooth muscle cells}
- A61L 27/383: {Nerve cells, e.g. dendritic cells, Schwann cells}
- A61L 27/3834: {Cells able to produce different cell types, e.g. hematopoietic stem cells, mesenchymal stem cells, marrow stromal cells, embryonic stem cells}
- A61L 27/52: Hydrogels or hydrocolloids
- A61L 27/54: Biologically active materials, e.g. therapeutic substances {(A61L27/227 takes precedence)}
- B33Y 10/00: Processes of additive manufacturing
- B33Y 70/00: Materials specially adapted for additive manufacturing
- A61L: METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES (preservation of bodies or disinfecting characterised by the agents employed A01N; preserving, e.g. sterilising, food or foodstuffs A23; preparations for medical, dental or toiletry purposes A61K)
- B33Y: ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING

## Abstract

Collagen compositions, methods for preparing those collagen compositions, and 3D constructs formed from those collagen compositions are provided. In particular, methods of isolating collagen that exhibits an enhanced rate of gelling, such collagen compositions, and 3D constructs formed from such collagen compositions are provided.

## Claims

### Claim 1

A bio gel composition comprising:
collagen material in an amount greater than 5 mg/mL of the total bio gel composition;
a weak acid solution;
one or more cross-linking agents selected from ribose, riboflavin, and combinations thereof; and
optionally living cells;
wherein the bio gel composition:
undergoes gelation in less than about 3 min at a temperature of about 25° C. to about 37° C.;
is liquid under shear stress of about 15 Pa to about 100 Pa; and
remains solid during gelation under hydrostatic pressure of about 30 Pa to about 120 Pa; and
wherein the bio gel composition is obtained by a method comprising:
providing a first receptacle having the collagen in the weak acid solution;
providing a second receptacle having a carrier; and
contacting the collagen material in the weak acid solution of the first receptacle with the carrier of the second receptacle to obtain the bio gel composition, wherein the contacting comprises stirring the collagen material in the weak acid solution and the carrier to form the bio gel composition until the bio gel composition has a homogeneity of about 50% to about 99.9%; and
wherein the collagen material is obtained by a method comprising:
(a) processing a collagen-based biomaterial to obtain biomaterial particles;
(b) contacting the biomaterial particles with a weak acid solution to obtain a collagen-containing solution;
(c) segregating the collagen-containing solution to obtain an extracted collagen solution;
(d) contacting the extracted collagen solution with a salt solution to obtain a collagen precipitate;
(e) re-suspending the collagen precipitate in a weak acid solution to obtain a re-suspended collagen solution; and performing viral and prion inactivation of the re-suspended collagen solution;
(f) desalting the re-suspended collagen solution; and
(g) drying the re-suspended collagen solution to obtain the collagen material.

### Claim 2 (depends on 1)

The composition of claim 1, wherein the collagen material comprises type I collagen, wherein the type I collagen is harvested from mammalian sources selected from bovine, porcine, murine, or combinations thereof.

### Claim 3 (depends on 1)

The composition of claim 1, wherein the collagen material is present in an amount of about 5 mg/mL to about 200 mg/mL.

### Claim 4 (depends on 1)

The composition of claim 1, wherein the weak acid solution comprises weak acid in an amount of about 0.01% to about 3% of the weak acid solution.

### Claim 5 (depends on 1)

The composition of claim 1, wherein the weak acid solution comprises formic acid, propanoic acid, acetic acid, citric acid, butanoic acid, salicylic acid, gluconic acid, heptonic acid, carbonic acid, hydrofluoric acid, nitrous acid, hypochlorous acid, or combinations thereof.

### Claim 6 (depends on 1)

The composition of claim 1, wherein the bio gel composition sags less than about 1% to about 30% of its height under gravity over a period of up to about 60 s to about 120 s after gelation.

### Claim 7 (depends on 1)

The composition of claim 1, wherein the bio gel composition increases storage modulus up to about 5000% after gelation.

### Claim 8 (depends on 1)

The bio gel composition of claim 1, wherein the contacting comprises stirring the collagen material in the weak acid solution and the carrier to form the bio gel composition until the bio gel composition has a homogeneity of about 99.9%.

### Claim 9 (depends on 1)

The composition of claim 1, wherein:
the method of obtaining the collagen material comprises contacting the biomaterial particles with the weak acid solution over a period of 24 h to 2 weeks; and
the segregating comprises centrifuging the collagen-containing solution.

### Claim 10 (depends on 1)

The composition of claim 1, wherein contacting the extracted collagen solution with the salt solution is repeated until at least 95% pure collagen precipitate is obtained.

### Claim 11 (depends on 1)

The composition of claim 1, wherein the method of obtaining the collagen material further comprises adjusting the pH of the re-suspended collagen solution to a pH of 2.8 to 4.5.

### Claim 12 (depends on 1)

The composition of claim 1, wherein:
the desalting comprises dialysis against a 0.01% to 20% weak acid solution; and
the drying is carried out by supercritical fluid drying, cyclic pressure drying, inert medium drying, spray drying, fluidized bed drying, lyophilization, or dehydration.

### Claim 13 (depends on 1)

The composition of claim 1, wherein the method of obtaining the collagen material further comprises, after (g):
(h) sterilizing the collagen material using irradiation, incubation with peracid, or supercritical fluid treatment.

### Claim 14 (depends on 1)

The composition of claim 1, wherein the contacting comprises stirring the collagen material in the weak acid solution and the carrier to form the bio gel composition until the homogeneity of the bio gel composition is about 90%.

### Claim 15 (depends on 1)

The composition of claim 1, wherein the collagen material is present in an amount of about 50 mg/mL to about 200 mg/mL of the total bio gel composition.

### Claim 16 (depends on 1)

The composition of claim 1, wherein the collagen material is present in an amount of about 60 mg/mL to about 200 mg/mL of the total bio gel composition.

### Claim 17 (depends on 1)

The composition of claim 1, wherein the collagen material is present in an amount of about 100 mg/mL to about 200 mg/mL of the total bio gel composition.

### Claim 18 (depends on 1)

A method for fabricating the bio gel composition of claim 1 as a three-dimensional structure, comprising the steps of:
maintaining the bio gel composition at a temperature of about 0° C. to about 12° C.;
depositing the bio gel composition to form a three-dimensional structure, wherein the three-dimensional structure undergoes gelation; and
curing the three-dimensional structure.

### Claim 19 (depends on 18)

The method of claim 18, wherein the three-dimensional structure undergoes gelation in less than about 3 min at a temperature of about 25° C. to about 37° C.

### Claim 20 (depends on 18)

The method of claim 18, wherein the curing is performed at a temperature from about 34° C. to about 37° C. in a buffer solution.