WO2025259801A2 - Deposition-based additive manufacturing (am) of whipped silk creams - Google Patents

Abstract

A printhead may include an extrusion nozzle configured to extrude a biopolymer-based cellular biomaterial and a ventilation system, the ventilation system positioned adjacent to the extrusion nozzle, wherein an airflow from the ventilation system is directed to the biopolymer-based cellular biomaterial extruded from the extrusion nozzle to form an extruded biopolymer-based cellular biomaterial; and wherein the printhead is configured to store the biopolymer-based cellular biomaterial under a storage pressure.

Description

DEPOSITION-BASED ADDITIVE MANUFACTURING (AM) OF WHIPPED SILK CREAMS

CLAIM TO PRIORITY

[0001] This application claims priority to and the benefit of U.S. Provisional Application No. 63/658,741 (T002789 PROV) filed in the U.S. Patent and Trademark Office on June 11, 2024, and U.S. Provisional Application No. 63/658,745 (T002833 PROV) filed on June 11, 2024. The foregoing patent applications are hereby incorporated by reference in their entireties for all purposes.

BACKGROUND

[0002] Polymeric foams, or cellular or expanded polymers (e.g., polyurethane, polyethylene), are an integral part of our daily lives. Their unique traits have made them desirable across various industries. At present, these materials are commonplace due to their unmatched properties, affordable manufacturing costs, and flexibility. Key features of polymeric foams include their lightweight nature, excellent thermal insulation, high strength-to-weight ratio, ease of molding, impact strength, and low dielectric constant. However, these materials also present challenges such as their persistence in the environment, microplastic generation upon degradation, inclusion of toxic additives, non-biodegradability, and resource-intensive production.

[0003] Conversely, there's a growing trend towards using biomaterials with additive manufacturing techniques. This approach allows for the creation and customization of multi-functional materials. For instance, it enables the design of components with specific structures and properties that are not only fully compatible with biological systems and scalable, but also recyclable, sustainable, and decomposable.

[0004] A need exists for improved materials and methods.

SUMMARY

[0005] In some aspects, the techniques described herein relate to a printhead of a three-dimensional printer including: an extrusion nozzle configured to extrude a biopolymer-based cellular biomaterial; a ventilation system, the ventilation system positioned adjacent to the extrusion nozzle, wherein an airflow from the ventilation system is directed to the biopolymer-based cellular biomaterial extruded from the extrusion nozzle to form an extruded biopolymer-based cellular biomaterial; and wherein the printhead is configured to store the biopolymer-based cellular biomaterial under a storage pressure.

[0006] In some aspects, the techniques described herein relate to a process for generating a biopolymer-based cellular biomaterial, the process including: flowing a biopolymer-based cellular biomaterial under pressure through a printhead of a three-dimensional printer, the biopolymer-based cellular biomaterial including silk fibroin in an amount by weight of between 1% and 40%, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1% and 10%, at least one polysaccharide in an amount by weight of between 10% and 30%, and at least one plasticizer in an amount by weight of between 20% and 75%; extruding the biopolymer-based cellular biomaterial through an extrusion nozzle of the printhead to form an extruded biopolymer-based cellular biomaterial, the extruding in accordance with a volumetric extrusion rate (Q) defined by Q = co x h x v (where co = extrusion width, h = layer height, v = velocity of extrusion); and drying the extruded biopolymer-based cellular biomaterial exiting the extrusion nozzle.

[0007] In some aspects, the techniques described herein relate to a method of making a biopolymerbased cellular biomaterial, the method including: whipping a liquid composition for a predetermined whipping time at a predetermined speed to form a biopolymer-based cellular biomaterial, the liquid composition including silk fibroin, at least one surfactant, at least one polysaccharide, and at least one plasticizer; pressurizing a volume of the biopolymer-based cellular biomaterial; and concurrently extruding and drying the biopolymer-based cellular biomaterial to form an extruded, dried biopolymer-based cellular biomaterial.

[0008] These and other systems, methods, objects, features, and advantages of the present disclosure will be apparent to those skilled in the art from the following detailed description of the preferred embodiment and the drawings.

[0009] All documents mentioned herein are hereby incorporated in their entirety by reference. References to items in the singular should be understood to include items in the plural, and vice versa, unless explicitly stated otherwise or clear from the text. Grammatical conjunctions are intended to express any and all disjunctive and conjunctive combinations of conjoined clauses, sentences, words, and the like, unless otherwise stated or clear from the context.

BRIEF DESCRIPTION OF THE FIGURES

[0010] The disclosure and the following detailed description of certain embodiments thereof may be understood by reference to the following figures:

[0011] Fig. 1 A depicts an embodiment of a printhead, in accordance with aspects of the present disclosure.

[0012] Fig. IB depicts an object printed by an embodiment of the printhead.

[0013] Fig. 1C depicts a representation of the extrusion and ventilation system.

[0014] Fig. 2 depicts the impact of nozzle geometries on material extrusion (left) and the nozzles used (right).

[0015] Fig. 3 depicts an oscillatory amplitude sweep at 1 rad/s experiment conducted at 20 °C. DETAILED DESCRIPTION

[0016] Before the present disclosure is described in further detail, it is to be understood that the disclosure is not limited to the particular embodiments described. It is also understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting. The scope of the present disclosure will be limited only by the claims. As used herein, the singular forms "a", "an", and "the" include plural embodiments unless the context clearly dictates otherwise.

[0017] In this application, unless otherwise clear from context, (i) the term “a” may be understood to mean “at least one”; (ii) the term “or” may be understood to mean “and/or”; (iii) the terms “comprising” and “including” may be understood to encompass itemized components or steps whether presented by themselves or together with one or more additional components or steps; and (iv) the terms “about” and “approximately” are used as equivalents and may be understood to permit standard variation as would be understood by those of ordinary skill in the art; and (v) where ranges are provided, endpoints are included.

[0018] Approximately: as used herein, the term “approximately” or “about,” as applied to one or more values of interest, refers to a value that is similar to a stated reference value. In certain embodiments, the term “approximately” or “about” refers to a range of values that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value).

[0019] Composition: as used herein, may be used to refer to a discrete physical entity that comprises one or more specified components. In general, unless otherwise specified, a composition may be of any form - e.g., gas, gel, liquid, solid, etc. In some embodiments, “composition” may refer to a combination of two or more entities for use in a single embodiment or as part of the same article. It is not required in all embodiments that the combination of entities result in physical admixture, that is, combination as separate co-entities of each of the components of the composition is possible; however many practitioners in the field may find it advantageous to prepare a composition that is an admixture of two or more of the ingredients in a pharmaceutically acceptable carrier, diluent, or excipient, making it possible to administer the component ingredients of the combination at the same time.

[0020] Improve, increase, or reduce: as used herein or grammatical equivalents thereof, indicate values that are relative to a baseline measurement, such as a measurement in a similar composition made according to previously known methods. [0021] Substantially: as used herein, the term “substantially” refers to the qualitative condition of exhibiting total or near-total extent or degree of a characteristic or property of interest. One of ordinary skill in the biological arts will understand that biological and chemical phenomena rarely, if ever, go to completion and/or proceed to completeness or achieve or avoid an absolute result. The term “substantially” is therefore used herein to capture the potential lack of completeness inherent in many biological and chemical phenomena.

[0022] It should be apparent to those skilled in the art that many additional modifications beside those already described are possible without departing from the inventive concepts. In interpreting this disclosure, all terms should be interpreted in the broadest possible manner consistent with the context. Variations of the term "comprising" should be interpreted as referring to elements, components, or steps in a non-exclusive manner, so the referenced elements, components, or steps may be combined with other elements, components, or steps that are not expressly referenced. Embodiments referenced as "comprising" certain elements are also contemplated as "consisting essentially of" and "consisting of" those elements. When two or more ranges for a particular value are recited, this disclosure contemplates all combinations of the upper and lower bounds of those ranges that are not explicitly recited. For example, recitation of a value of between 1 and 10 or between 2 and 9 also contemplates a value of between 1 and 9 or between 2 and 10. When a value is given in an amount by weight, the value may be given in a dry solids basis, excluding the weight of water or other solvents within the composition.

[0023] The disclosure herein describes a novel process for additive manufacturing (AM) of silkbased foam, known elsewhere and herein alternatively as a whipped silk cream or a biopolymerbased cellular biomaterial. It should be appreciated that the term “foam” may be used in a different context elsewhere (e.g., where noted in the Examples), but the usage herein refers primarily to printing of a whipped silk cream as described in the Examples and in PCT International Application Number PCT/US2025/026472, filed April 25, 2025, which is incorporated by reference in its entirety herein for all purposes. A person of skill in the art would understand that where a foam is described herein, the foam may be the same as the whipped silk cream of PCT/US2025/026472 or a variation thereof. In some cases, the foam may have the same composition as the whipped silk cream. In some cases, the foam may have an adjusted composition. A skilled artisan would appreciate how to vary a whipped silk cream’ s composition to make it suitable for use with the claimed invention. For example, a skilled artisan, having the benefit of this disclosure and given a specific context, would understand that certain applications may require a different polysaccharide or combination of polysaccharides in the whipped silk cream composition depending on the ultimate material properties desired. [0024] The process is divided into two main parts: the first focuses on the printed material (silk foam), and the second on the additive manufacturing (AM) method described herein. This is the basis of our research at the intersection of natural biomaterials and additive manufacturing, leading to the development and optimization of a silk-based cellular biomaterial and a manufacturing technique specifically designed for such complex materials.

[0025] Silk foam, as described herein and by U.S. Provisional Patent Applications Nos. 63/658,741 and 63/658,745 which are hereby incorporated by reference in their entireties for all purposes, possesses intriguing structural and physicochemical properties. The compositions described herein are based on the use of an aqueous silk fibroin (SF) solution or SF powder in conjunction with glycerol, xanthan gum from Xanthomonas Campestris, and/or alginic acid sodium salt from brown algae. As an added benefit, silk fibroin is renowned for its capacity to stabilize biological molecules and colorimetric sensors. The potential to 3D print foam structures with high porosity and surface area is likely to augment the sensitivity and versatility of other sensor technologies, particularly in gas sensing applications.

[0026] To achieve a foam-like composition, the time and speed must be optimized to incorporate a sufficient quantity of air. The inventors discovered that in some cases, a range of 4 minutes to 6 minutes at a speed of 500 rpm - 600 rpm (250 revolutions - 300 revolutions) may be ideal depending on the concentrations of the ingredients, although a skilled artisan will appreciate that a combination of higher or lower times and speeds may achieve similar effects.

[0027] One embodiment of the printhead disclosed herein is shown in Fig. 1 A. An object printed by the printhead is shown in Fig. IB. At a high level, a printhead consisting of a ventilation system, a screw head, and a nozzle is attached to a syringe, stepper motor, and transmission mounted on an axis. The nozzle may be modular and may have a range of geometries, opening sizes and shapes, and volumes. An automatic level sensor may be used to correct the printer’s level in the Z direction. In some cases, the level sensor may be the BL Touch. A skilled artisan will appreciate that certain parts or the arrangement of parts may be modified to achieve similar results.

[0028] A zoomed in view of one embodiment of the extrusion and ventilation system is shown in Fig. 1C. In some cases, there may be one or more fans attached to the printhead at an angle to the extrusion nozzle. In the case of more than one fan attached to the printhead, fans may be arranged at angles with an axis of symmetry down the longitudinal axis of the printhead, ensuring even application of airflow to the material being extruded. The at least one fan may be directed towards the biopolymer-based cellular biomaterial exiting the extrusion nozzle to enable layer-by-layer drying of the extruded biopolymer-based cellular biomaterial. The fan may move, rotate, or be fixed. An airflow from the fan may be directed at an angle with respect to a direction of extrusion. The angle may be between 35° and 65°. The angle may be at least 40°, at least 42°, at least 45°, or at least 47°. The fan may have a fan speed of between 3000 bpm and 4000 bpm. The fan speed may be at least 3100 bpm, at least 3200 bpm, at least 3500 bpm, or at least 3800 bpm. These fan speeds combined with the fan angles were found to enable layer-by-layer drying of the printed object by evenly drying the material as extruded. Even drying reduces shrinkage of extruded materials normally observed in water-based materials. The ventilation system may be configured to deliver an airflow at a predetermined fluid velocity. The predetermined fluid velocity may be between 4 m/s and 10 m/s. The predetermined fluid velocity may be at least 5 m/s, at least 7 m/s, or at least 9 m/s. The predetermined fluid velocity may be at most 10 m/s, at most 8 m/s, or at most 6 m/s.

[0029] The ventilation system may be configured to deliver an airflow at a variable fluid velocity. The variable fluid velocity may vary with at least one of of a change in distance from the ventilation system to a surface of the extruded biopolymer-based cellular biomaterial, an extrusion rate from the extrusion nozzle, a layer height, a length of path, an extrusion width, or a composition of the biopolymer-based cellular biomaterial. The ventilation system may be arranged at a distance of between 60 mm and 80 mm from a tip of the extrusion nozzle. The distance from a tip of the extrusion nozzle may be at least 65 mm, at least 70 mm, or at least 75 mm. The ventilation system may be arranged at a distance of between 60 mm and 100 mm from the extruded biopolymer-based cellular biomaterial. The distance from the extruded biopolymer-based cellular biomaterial may be at least 65 mm, at least 80 mm, or at least 95 mm.

[0030] The nozzle may be fed by a vessel which stores material to be extruded under a storage pressure. The vessel may include a syringe that is removable and/or refillable. The vessel may be configured to store a volume of the biopolymer-based cellular biomaterial under the storage pressure. The storage pressure may be adjustable or fixed. The volume may be between 100 mL and 400 mL. The volume may be at least 150 mL, at least 250 mL, or at least 300 mL. The volume of storage may be adjustable or fixed. The vessel may have more than one chamber. The volume of storage may be divisible among the more than one chamber of the vessel. In some aspects, the storage pressure may be greater than atmospheric pressure. The storage pressure may be between 0.25 MPa and 25 MPa. In some cases, the storage pressure may be at least 0.5 MPa, including but not limited to, at least 1 MPa, at least 5 MPa, at least 10 MPa, or at least 20 MPa.

[0031] The printhead described herein was developed as an improvement to a traditional Cartesian printer, with the motherboard replaced and the standard fused filament fabrication extrusion kit replaced with the mechanical extrusion system described herein.

[0032] Furthermore, additive manufacturing of this material is complicated by various factors that are addressed by specific features of the printhead and/or method. For example, the compressibility of the silk foam material makes it difficult to handle in a pressurized environment. In another example, the instability of the high polydispersity biomaterial due to coalescence of the material during storage, loading and printing can be induced by shear.

[0033] The inventors discovered a threshold of pressure at which certain materials would flow. Further, it was discovered that adding surfactant helped in stabilizing the foam.

[0034] The inventors discovered that a surfactant may be used to fine tune the stability and flow behavior of the cream during extrusion, however the surfactant must be carefully chosen. In some cases, the surfactant is a monoglyceride. In some cases, the surfactant is glycerol monostearate. The inventors unexpectedly discovered that some common surfactants, such as sodium dodecyl sulfate (SDS), do not generate stable foams. The absence of an adequate surfactant made the material more compressible which translates into the need to increase the pressure drastically during printing making the material difficult to control. In some cases, the biopolymer-based cellular biomaterial and/or the surfactant can be free of anionic surfactant, free of sodium sulfate surfactant, and/or free of sodium dodecyl sulfate. A proper surfactant augmented the shear resistance of the material during extrusion and facilitated the preservation of the foam structure, including by thickening interfacial films.

[0035] In yet another example, shrinkage during solidification of the material occurs due to evaporation of water. Modifications to the nozzle profile improved extrusion overall, and the addition of a built-in drying fan permits the homogenous layer-by-layer solidification (facilitated by evaporation of the solvent) of the material and reduces shrinkage drastically.

[0036] Without the above-described factors being properly addressed by the inventors, the material collapses faster, meaning the material’s mechanical properties would change either during storage or during printing. Additionally, not adding a proper surfactant makes the material more compressible which translates to increasing the pressure drastically during printing - negatively impacting the ability to control the material.

[0037] Surprisingly, this printhead overcomes the aforementioned issues due to the arrangement of components and selection of settings described herein, discovered after extensive experimentation. Thanks to its versatility in printing complex structures on demand, this biopolymer-based cellular biomaterial-based additive manufacturing is widely applicable across various sectors including biomedical, engineering, manufacturing, aerospace, automotive, and architecture.

[0038] The present disclosure provides a printhead for a three-dimensional printer. The printhead includes an extrusion nozzle and a ventilation system. The extrusion nozzle is configured to extrude a biopolymer-based cellular biomaterial. The ventilation system is positioned adjacent to the extrusion nozzle. Airflow from the ventilation system is directed to the biopolymer-based cellular biomaterial extruded from the extrusion nozzle. The printhead is configured to store the biopolymerbased cellular biomaterial under a storage pressure.

[0039] In some aspects, the printhead may include a mechanical extrusion system. The mechanical extrusion system may include a bipolar stepper motor with a gearbox and a rear axle configured to operate at a torque of between 1 N-m and 4 N-m, including at least 1.5 N-m. In some cases, the torque may be at least 1.5 N-m, at least 2 N-m, or at least 3 N-m. In some cases, the torque may be at most 4 N-m, at most 3.5 N-m, at most 2.5 N-m, or at most 2 N-m. The bipolar stepper motor may enable both clockwise and counterclockwise rotation of a belt and a threaded rod, which guides a piston and facilitates the biopolymer-based cellular biomaterial extrusion. When powered, the motor enables both clockwise and counterclockwise rotation of a belt and threaded rod, guiding the piston and facilitating material extrusion.

[0040] In some aspects, the biopolymer-based cellular biomaterial may have a firmness of between 100 N/m² and 400 N/m² and/or a density of between 0.08 g/cm³ and 0.25 g/cm³. The firmness may be at least 150 N/m², at least 250 N/m², or at least 350 N/m². The firmness may be at most 400 N/m², at most 300 N/m², or at most 200 N/m². The density may be at least 0. 12 g/cm³, at least 0. 16 g/cm³, or at least 0.20 g/cm³. The density may be at most 0.25 g/cm³, at most 0.21 g/cm³, at most 0.17 g/cm³, or at most 0.13 g/cm³.

[0041] In other aspects, the printhead may extrude the biopolymer-based cellular biomaterial in accordance with a volumetric extrusion rate defined as Q = co x h x v, where co = extrusion width, h = layer height, and v = velocity of extrusion. The extrusion rate may be between 125 mm/min and 350 mm/min. The extrusion rate may be at least 265 mm/min, at least 290 mm/min, or at least 310 mm/min. The extrusion rate may be at most 310 mm/min, at most 250 mm/min, at most 200 mm/min, or at most 150 mm/min. The extrusion rate may be further defined by a rate of volume change of the extruded biopolymer-based cellular material. The extrusion rate may vary up to 45% of an original value of the extrusion rate, including up to 75%. The extrusion rate may vary up to 65%, up to 60%, up to 55%, up to 48%, or up to 44%.

[0042] In some aspects, drying may include directing an airflow from a ventilation system adjacent to the extrusion nozzle towards the extruded biopolymer-based cellular biomaterial exiting the extrusion nozzle. Drying may also include any variations discussed herein.

[0043] The present disclosure provides a method of making a biopolymer-based cellular biomaterial. The method includes whipping a liquid composition for a predetermined whipping time at a predetermined speed to form a biopolymer-based cellular biomaterial. The liquid composition includes silk fibroin, at least one polysaccharide, and at least one plasticizer. The method further includes pressurizing a volume of the biopolymer-based cellular biomaterial and concurrently extruding and drying the biopolymer-based cellular biomaterial to form an extruded, dried biopolymer-based cellular biomaterial.

[0044] The present disclosure provides a process for generating a biopolymer-based cellular biomaterial. The process includes flowing a biopolymer-based cellular biomaterial including silk fibroin in an amount by weight of between 1% and 40%, optionally as a powder, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1% and 10%, at least one polysaccharide in an amount by weight of between 10% and 30%, and plasticizer in an amount by weight of between 20% and 75% under pressure through a printhead of a three-dimensional printer.

[0045] The process then includes extruding the biopolymer-based cellular biomaterial through an extrusion nozzle of the printhead to form an extruded biopolymer-based cellular biomaterial. The extrusion is in accordance with an extrusion rate defined as Q = co x h x v, where co = extrusion width, h = layer height, and v = velocity of extrusion. The extrusion rate may also correspond to the amount of material extruded along the tool path and/or the deposited volume per unit of path. The process further includes drying the extruded biopolymer-based cellular biomaterial exiting the extrusion nozzle.

[0046] In the compositions disclosed herein, SF contributes to the water content of the whipped cream, while polysaccharides influence the final density. Without wishing to be bound by any particular theory, it is believed that polysaccharides reinforce the bubbles’ shells. Unexpectedly, glycerol is essential for plasticizing the foams and achieving the target properties, despite being detrimental to foam stability. Glycerol monostearate is used as an emulsifier, a stabilizer, and a surfactant. Without wishing to be bound by any particular theory, glycerol monostearate is believed to reduce surface tension by absorbing at the air- liquid interface as well as enhance cream stability by creating thicker interfacial films. These factors, among others, collectively affect the viscoelastic properties of the creams. Impressively, the inventors have carefully optimized their ranges to enable the storage of a foam volume of between 100 mL and 200 mL under pressure and uniform printing without defects.

[0047] In some aspects, the predetermined whipping length of time may be at least 8.5 minutes or between 5 minutes and 30 minutes. The predetermined whipping length of time may be at least 5 minutes, at least 6 minutes, at least 7 minutes, at least 8 minutes, at least 9 minutes, or at least 10 minutes. The predetermined whipping length of time may be at most 30 minutes, at most 25 minutes, at most 20 minutes, at most 15 minutes, or at most 10 minutes. The predetermined speed may be at least 185 rpm or between 160 rpm and 200 rpm. The predetermined speed may be at least 160 rpm, at least 170 rpm, at least 175 rpm, or at least 180 rpm. The predetermined speed may be at most 200 rpm, at most 190 rpm, or at most 180 rpm. [0048] In other aspects, the at least one plasticizer may be glycerol, di(ethylene glycol), a tri(ethylene glycol), a di(propylene glycol), a tri(propylene glycol), triethanolamine, a vegetable oil, or combinations thereof. The at least one polysaccharide may be xanthan gum, an alginate, a high molecular weight sugar, a cellulose derivate, or combinations thereof. The liquid composition may comprise silk fibroin, glycerol, sodium alginate, and xanthan gum.

[0049] In other aspects, the biopolymer-based cellular biomaterial may further include sodium alginate in an amount by weight of between 1 % and 20% and xanthan gum in an amount by weight of between 1% and 20%. Sodium alginate may be present in at least 1%, at least 5%, at least 9%, or at least 14% by weight. Sodium alginate may be present in at most 20%, at most 16%, at most 12%, or at most 8% by weight. Xanthan gum may be present in at least 1%, at least 5%, at least 9%, or at least 14% by weight. Xanthan gum may be present in at most 20%, at most 16%, at most 12%, or at most 8% by weight.

[0050] In some aspects, the biopolymer-based cellular biomaterial of printhead, process, or method described herein may include silk fibroin. The biopolymer-based cellular biomaterial may include silk fibroin, at least one plasticizer, at least one surfactant, and at least one polysaccharide. The at least one polysaccharide may be xanthan gum, an alginate, a high molecular weight sugar, a cellulose derivative, or a combination thereof. The alginate may be alginic acid sodium salt.

[0051] The at least one plasticizer may be of a glycerol, a di(ethylene glycol), a tri(ethylene glycol), a di(propylene glycol), a tri(propylene glycol), triethanolamine, a vegetable oil, or combinations thereof. The plasticizer may have at least one -OH substituent. The plasticizer may be present in the biopolymer-based cellular biomaterial in an amount by weight of between 20.0% and 75.0%. The plasticizer may be present in at least 30%, at least 40%, at least 50%, or at least 60%. The plasticizer may be present in at most 70%, at most 65%, at most 55%, or at most 45%.

[0052] The biopolymer-based cellular biomaterial may include silk fibroin in an amount by weight of between 1% and 40%, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1% and 10%, between 1% and 20% sodium alginate, and between 1% and 20% xanthan gum.

[0053] Silk fibroin may be present in an amount by weight of between 10% and 30% or between 15% and 25%. The silk fibroin may be present in an amount by weight of at least 15%, at least 17%, at least 19%, at least 21%, or at least 23%. The silk fibroin may be present in an amount by weight of at most 24%, at most 22%, at most 20%, at most 18%, or at most 16%.

[0054] The surfactant may be a monoglyercide. The surfactant may be glycerol monostearate. The surfactant may be present in an amount by weight of at least 1%, at least 3%, at least 5%, at least 7%, or at least 9%. The surfactant may be present in an amount by weight of at most 10%, at most 8%, at most 6%, at most 4%, or at most 2%.

[0055] The polysaccharide may be present in an amount by weight of between 0.1% and 30%. The polysaccharide may be present in an amount by weight of at least 15%, at least 17%, at least 19%, at least 21%, or at least 23%. The polysaccharide may be present in amount by weight of at most 24%, at most 21%, at most 18%, at most 12%, or at most 6%. The polysaccharide may be sodium alginate, xanthan gum, or a combination of the two.

[0056] The silk fibroin and the at least one polysaccharide (e.g., the xanthan gum, the alginate, or a mixture thereof) may be present in the biopolymer-based cellular biomaterial in a weight ratio of between 1 :4 and 20: 1 or between 1 :2 and 10: 1. The weight ratio of silk fibroin to xanthan gum may be at least 1 :4, at least 1 :3, or at least 1 :2. The weight ratio of silk fibroin to xanthan gum may be at most 20:1, at most 19:1, al most 18: 1, at most 16: 1, at most 15: 1, al most 14: 1, al most 12: 1, al most 11 :1, or at most 10:1.

[0057] The biopolymer-based cellular biomaterial may further include at least one of a sensing agent, a therapeutically active agent, a colorant, or an aroma-providing compound.

[0058] In some aspects, the biopolymer-based cellular biomaterial of printhead, process, or method described herein may have a regular or an irregular porosity. The biopolymer-based cellular biomaterial may have a firmness of at least 100 N/m², at least 200 N/m², at least 250 N/m², at least 300 N/m², or at least 400 N/m². The biopolymer-based cellular biomaterial may have a density of at least 0.08 g/cm³, at least 0. 1 g/cm³, at least 0.12 g/cm³, at least 0.2 g/cm³, or at least 0.25 g/cm³. [0059] In other aspects, the biopolymer-based cellular biomaterial of the printhead, process, or method described herein may be extruded in a single path.

[0060] As used herein, "silk fibroin" refers to silk fibroin protein whether produced by silkworm, spider, or other insect, or otherwise generated (Lucas et al., Adv. Protein Chem., 13: 107-242 (1958)). Any type of silk fibroin can be used in different embodiments described herein. Silk fibroin produced by silkworms, such as Bombyx mori, is the most common and represents an earth-friendly, renewable resource. For instance, silk fibroin used in a silk film may be attained by extracting sericin from the cocoons of B. mori. Organic silkworm cocoons are also commercially available. There are many different silks, however, including spider silk (e.g., obtained from Nephila clavipes), transgenic silks, genetically engineered silks, such as silks from bacteria, yeast, mammalian cells, transgenic animals, or transgenic plants, and variants thereof, that can be used. See, e.g., WO 97/08315 and U.S. Pat. No. 5,245,012, each of which is incorporated herein by reference in their entireties. [0061] According to various embodiments, a variety of functionalizing agents may be used with the silk-containing embodiments described herein (e.g., silk membrane, silk composition, silk articles, silk matrix, silk foam, silk microsphere, liquid composition, whipped silk cream, silk meringue, compressed silk meringue, hot-pressed silk meringue, silk leather, silk powder, silk toner, edible silkbased films, etc.). It should be understood that the examples herein may recite one or a few silkcontaining embodiments but are applicable to any silk-containing embodiment, as applicable. In some embodiments, a functionalizing agent may be any compound or molecule that facilitates the attachment to and/or development (e.g., growth) of one or more endothelial cells on a silk membrane. In some embodiments, a functionalizing agent may be any compound or molecule that facilitates the attachment and/or development (e.g., growth) of one or more megakaryocytes and/or hematopoietic progenitor cells on a silk matrix and/or silk membrane. In some embodiments, a functionalizing agent may be or comprise an agent suitable for facilitating the production of one or more of white blood cells and red blood cells.

[0062] In some embodiments, a functionalizing agent may be or comprise a cell attachment mediator and/or an extracellular matrix protein, for example: collagen (e.g., collagen type I, collagen type III, collagen type IV or collagen type VI), elastin, fibronectin, vitronectin, laminin, fibrinogen, von Willebrand factor, proteoglycans, decorin, perlecan, nidogen, hyaluronan, and/or peptides containing known integrin binding domains e.g. “RGD” integrin binding sequence, or variations thereof, that are known to affect cellular attachment.

[0063] In some embodiments, a functionalizing agent may be any soluble molecule produced by endothelial cells. Non-limiting examples include fibroblast growth factor-1 (FGF1 ) and vascular endothelial growth factors (VEGF).

[0064] According to some embodiments, a plurality of functionalizing agents may be used. For example, in some embodiments wherein production of platelets is desired, provided compositions may comprise the use of laminin, fibronectin and/or fibrinogen, and type IV collagen in order to facilitate the attachment and growth of endothelial cells on a silk membrane (e.g., a porous silk membrane) and/or attachment of megakaryocytes to a silk matrix.

[0065] In some embodiments, a functionalizing agent may be embedded or otherwise associated with a silk membrane and/or silk matrix such that at least a portion of the functionalizing agent is surrounded by a silk membrane and/or silk matrix as contrasted to a functionalizing agent simply being positioned along the surface of a silk membrane and/or silk matrix. In some embodiments, a functionalizing agent is distributed along and/or incorporated in substantially the entire surface area of a silk membrane/silk wall. In some embodiments, a functionalizing agent is distributed and/or incorporated only at one or more discrete portions of a silk membrane/wall and/or silk matrix. In some embodiments, a functionalizing agent is distributed in and/or along at least one of the lumenfacing side of a silk wall and the matrix-facing side of a silk wall.

[0066] According to various embodiments, any application-appropriate amount of one or more functionalizing agents may be used. In some embodiments, the amount of an individual functionalizing agent may be between about 1 pg/ml and 1,000 pg/ml (e.g., between about 2 and 1,000, 5 and 1,000, 10 and 1,000, 10 and 500, 10 and 100 pg/ml). In some embodiments, the amount of an individual functionalizing agent may be at least 1 pg/ml (e.g., at least 5, 10, 15, 20 25, 50, 100, 200, 300 400, 500, 600, 700, 800, or 900 pg/ml ). In some embodiments, the amount of an individual functionalizing agent is at most 1,000 pg/ml (e.g., 900, 800, 700, 600, 500, 400, 300 200, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, or 5 pg/ml).

[0067] In some aspects, the composition comprises one or more sensing agents, such as a sensing dye. The sensing agents/sensing dyes are environmentally sensitive and produce a measurable response to one or more environmental factors. In some aspects, the environmentally- sensitive agent or dye may be present in the composition in an effective amount to alter the composition from a first chemical-physical state to a second chemical-physical state in response to an environmental parameter (e.g., a change in pH, light intensity or exposure, temperature, pressure or strain, voltage, physiological parameter of a subject, and/or concentration of chemical species in the surrounding environment) or an externally applied stimulus (e.g., optical interrogation, acoustic interrogation, and/or applied heat). In some cases, the sensing dye is present to provide one optical appearance under one given set of environmental conditions and a second, different optical appearance under a different given set of environmental conditions. Suitable concentrations for the sensing agents described herein can be the concentrations for the colorants and additives described elsewhere herein. A person having ordinary skill in the chemical sensing arts can determine a concentration that is appropriate for use in a sensing application of the inks described herein.

[0068] In some aspects, the first and second chemical-physical state may be a physical property of the composition, such as mechanical property, a chemical property, an acoustical property, an electrical property, a magnetic property, an optical property, a thermal property, a radiological property, or an organoleptic property. Exemplary sensing dyes or agents include, but are not limited to, a pH sensitive agent, a thermal sensitive agent, a pressure or strain sensitive agent, a light sensitive agent, or a potentiometric agent.

[0069] Exemplary pH sensitive dyes or agents include, but are not limited to, cresol red, methyl violet, crystal violet, ethyl violet, malachite green, methyl green, 2-(p- dimethylaminophenylazo)pyridine, paramethyl red, metanil yellow, 4-phenylazodiphenylamine, thymol blue, metacresol purple, orange IV, 4-o-Tolylazo-o-toluindine, quinaldine red, 2,4- dinitrophenol, erythrosine disodium salt, benzopurpurine 4B, N,N-dimethyl-p-(m-tolylazo) aniline, p-dimethylaminoazobenene, 4,4’-bis(2-amino-l-naphthylazo)-2,2’-stilbenedisulfonic acid, tetrabromophenolphthalein ethyl ester, bromophenol blue, Congo red, methyl orange, ethyl orange, 4-(4-dimethylamino-l-naphylazo)-3-methoxybenesulfonic acid, bromocresol green, resazurin, 4- phenylazo-l-napthylamine, ethyl red 2-(l-dimethylaminophenyazo) pyridine, 4-(p-ethoxyphenylazo)- m-phenylene-diamine monohydrochloride, resorcin blue, alizarin red S, methyl red, propyl red, bromocresol purple, chlorophenol red, p-nitrophenol, alizarin, 2-(2,4-dinitrophenylazo)-l-napthol-

3.6-disulfonic acid, bromothymol blue, 6, 8-dinitro-lH-quinazoline-2, 4-dione, brilliant yellow, phenol red, neutral red, m-nitrophenol, cresol red, turmeric, metacresol purple, 4,4'-bis(3-amino-l- naphthylazo)-2,2'-stilbenedisulfonic acid, thymol blue, p-naphtholbenzein, phenolphthalein, o- cresolphthalein, ethyl bis(2,4-dimethylphenyl) ethanoate, thymolphthalein, nitrazine yellow, alizarin yellow R, alizarin, p-(2,4-dihydroxyphenylazo) benzenesulfonic acid, 5,5'-indigodisulfonic acid,

2.4.6-trinitrotoluene, 1,3,5-trinitrobenezne, and clayton yellow.

[0070] Exemplary light responsive dyes or agents include, but are not limited to, photochromic compounds or agents, such as triarylmethanes, stilbenes, azastilbenes, nitrones, fulgides, spiropyrans, napthopyrans, spiro-oxazines, quinones, derivatives, and combinations thereof.

[0071] Exemplary potentiometric dyes include, but are not limited to, substituted amiononaphthylehenylpridinium (ANEP) dyes, such as di-4-ANEPPS, di-8-ANEPPS, and N-(4- Sulfobutyl)-4-(6-(4-(Dibutylamino)phenyl)hexatrienyl)Pyridinium (RH237).

[0072] Exemplary temperature sensitive dyes or agents include, but are not limited to, thermochromic compounds or agents, such as thermochromic liquid crystals, leuco dyes, fluoran dyes, octadecylphosphonic acid.

[0073] Exemplary pressure or strain sensitive dyes or agents include, but are not limited to, spiropyran compounds and agents.

[0074] Exemplary chemi-sensitive dyes or agents include, but are not limited to, antibodies such as immunoglobulin G (IgG) which may change color from blue to red in response to bacterial contamination.

[0075] In some aspects, the compositions comprise one or more additive, dopant, or biologically active agent suitable for a desired intended purpose. In some aspects, the additive or dopant may be present in the composition in an amount effective to impart an optical or organoleptic property to the composition. Exemplary additives or dopants that impart optical or organoleptic properties include, but are not limited to, dyes/pigments, flavorants, aroma compounds, granular or fibrous fillers. [0076] Additionally or alternatively, the additive, dopant, or biologically active agent may be present in the composition in an amount effective to "functionalize" the composition to impart a desired mechanical property or added functionality to the composition. Exemplary additive, dopants, or biologically active agent that impart the desired mechanical property or added functionality include, but are not limited to: environmentally sensitive/sensing dyes; active biomolecules; conductive or metallic particles; micro and nanofibers (e.g., silk nanofibers for reinforcement, carbon nanofibers); nanotubes; inorganic particles (e.g., hydroxyapatite, tricalcium phosphate, bioglasses); drugs (e.g., antibiotics, small molecules or low molecular weight organic compounds); proteins and fragments or complexes thereof (e.g., enzymes, antigens, antibodies and antigen-binding fragments thereof); DNA/RNA (e.g., siRNA, miRNA, mRNA); cells and fractions thereof (viruses and viral particles; prokaryotic cells such as bacteria; eukaryotic cells such as mammalian cells and plant cells; fungi).

[0077] In some aspects, the additive or dopant comprises a flavoring agent or flavorant.

[0078] Exemplary flavorants include ester flavorants, amino acid flavorants, nucleic acid flavorants, organic acid flavorants, and inorganic acid flavorants, such as, but not limited to, diacetyl, acetyl propionyl, acetoin, isoamyl acetate, benzaldehyde, cinnamaldehyde, ethyl propionate, methyl anthranilate, limonene, ethyl decadienoate, allyl hexanoate, ethyl maltol, ethylvanillin, methyl salicylate, manzanate, glutamic acid salts, glycine salts, guanylic acids salts, inosinic acid salts, acetic acid, ascorbic acid, citric acid, fumaric acid, lactic acid, malic acid, phosphoric acid, tartaric acid, derivatives, and mixtures thereof.

[0079] In some aspects, the additive or dopant comprises an aroma compound. Exemplary aroma compounds include ester aroma compounds, terpene aroma compounds, cyclic terpenes, and aromatic aroma compounds, such as, but not limited to, geranyl acetate, methyl formate, methyl acetate, methyl propionate, methyl butyrate, ethyl acetate, ethyl butyrate, isoamyl acetate, pentyl butyrate, pentyl pentanoate, octyl acetate, benzyl acetate, methyl anthranilate, myrcene, geraniol, nerol, citral, citronellal, cironellol, linalool, nerolidol, limonene, camphor, menthol, carone, terpineol, alpha-ionone, thujone, eucalyptol, benzaldehyde, eugenol, cinnamaldehyde, ethyl maltol, vanillin, anisole, anethole, estragole, thymol.

[0080] In some aspects, the additive or dopant comprises a colorant, such as a dye or pigment. In some aspects, the dye or pigment imparts a color or grayscale to the composition. The colorant can be different than the sensing agents and/or sensing dyes below. Any organic and/or inorganic pigments and dyes can be included in the inks. Exemplary pigments suitable for use in the present disclosure include International Color Index or C.I. Pigment Black Numbers 1 , 7, 1 1 and 31, C.I. Pigment Blue Numbers 15, 15:1, 15:2, 15:3, 15:4, 15:6, 16, 27, 29, 61 and 62, C.I. Pigment Green Numbers 7, 17, 18 and 36, C.I. Pigment Orange Numbers 5, 13, 16, 34 and 36, C.I. Pigment Violet Numbers 3, 19, 23 and 27, C.I. Pigment Red Numbers 3, 17, 22, 23, 48:1, 48:2, 57:1 , 81 :1 , 81 :2, 81 :3, 81:5, 101, 114, 122, 144, 146, 170, 176, 179, 181, 185, 188, 202, 206, 207, 210 and 249, C.I. Pigment Yellow Numbers 1, 2, 3, 12, 13, 14, 17, 42, 65, 73, 74, 75, 83, 30, 93, 109, 1 10, 128, 138, 139, 147, 142, 151, 154 and 180, D&C Red No. 7, D&C Red No. 6 and D&C Red No. 34, carbon black pigment (such as Regal 330, Cabot Corporation), quinacridone pigments (Quinacridone Magenta (228-0122), available from Sun Chemical Corporation, Fort Lee, N.J.), diarylide yellow pigment (such as AAOT Yellow (274- 1788) available from Sun Chemical Corporation); and phthalocyanine blue pigment (such as Blue 15:3 (294-1298) available from Sun Chemical Corporation). The classes of dyes suitable for use in present invention can be selected from acid dyes, natural dyes, direct dyes (either cationic or anionic), basic dyes, and reactive dyes. The acid dyes, also regarded as anionic dyes, are soluble in water and mainly insoluble in organic solvents and are selected, from yellow acid dyes, orange acid dyes, red acid dyes, violet acid dyes, blue acid dyes, green acid dyes, and black acid dyes. European Patent 0745651, incorporated herein by reference, describes a number of acid dyes that are suitable for use in the present disclosure. Exemplary yellow acid dyes include Acid Yellow 1 International Color Index or C.I. 10316); Acid Yellow 7 (C.I. 56295); Acid Yellow 17 (C.I. 18965); Acid Yellow 23 (C.I. 19140); Acid Yellow 29 (C.I. 18900); Acid Yellow 36 (C.I. 13065); Acid Yellow 42 (C.I. 22910); Acid Yellow 73 (C.I. 45350); Acid Yellow 99 (C.I. 13908); Acid Yellow 194; and Food Yellow 3 (C.I. 15985). Exemplary orange acid dyes include Acid Orange 1 (C.I. 13090/1); Acid Orange 10 (C.I. 16230); Acid Orange 20 (C.I. 14603); Acid Orange 76 (C.I. 18870); Acid Orange 142; Food Orange 2 (C.I. 15980); and Orange B. [0081] Exemplary red acid dyes include Acid Red 1 (C.I. 18050); Acid Red 4 (C.I. 14710); Acid Red 18 (C.I. 16255); Acid Red 26 (C.I. 16150); Acid Red 27 (C.I. 16185); Acid Red 51 (C.I. 45430, available from BASF Corporation, Mt. Olive, N.J.); Acid Red 52 (C.I. 45100); Acid Red 73 (C.I. 27290); Acid Red 87 (C.I. 45380); Acid Red 94 (C.I. 45440) Acid Red 194; and Food Red 1 (C.I. 14700). Exemplary violet acid dyes include Acid Violet 7 (C.I. 18055); and Acid Violet 49 (C.I. 42640). Exemplary blue acid dyes include Acid Blue 1 (C.I. 42045); Acid Blue 9 (C.I. 42090); Acid Blue 22 (C.I. 42755); Acid Blue 74 (C.I. 73015); Acid Blue 93 (C.I. 42780); and Acid Blue 158A (C.I. 15050). Exemplary green acid dyes include Acid Green 1 (C.I. 10028); Acid Green 3 (C.I. 42085); Acid Green 5 (C.I. 42095); Acid Green 26 (C.I. 44025); and Food Green 3 (C.I. 42053). Exemplary black acid dyes include Acid Black 1 (C.I. 20470); Acid Black 194 (Basantol® X80, available from BASF Corporation, an azo/1 :2 CR-complex.

[0082] Exemplary direct dyes for use in the present disclosure include Direct Blue 86 (C.I. 74180); Direct Blue 199; Direct Black 168; Direct Red 253; and Direct Yellow 107/132 (C.I. Not Assigned). [0083] Exemplary natural dyes for use in the present disclosure include Alkanet (C.I.

75520,75530); Annatto (C.I. 75120); Carotene (C.I. 75130); Chestnut; Cochineal (C.I.75470); Cutch (C.I. 75250, 75260); Divi-Divi; Fustic (C.L 75240); Brazilin (C.I. 75280); Logwood (C.I. 75200); Osage Orange (C.I. 75660); Paprika; Quercitron (C.I. 75720); Saffron (C.I. 75100) ; Sandal Wood (C.I. 75510, 75540, 75550, 75560); Sumac; and Turmeric (C.I. 75300). Exemplary reactive dyes for use in the present disclosure include Reactive Yellow 37 (monoazo dye); Reactive Black 31 (diazo dye); Reactive Blue 77 (phthalo cyanine dye) and Reactive Red 180 and Reactive Red 108 dyes. Suitable also are the colorants described in The Printing Ink Manual (5th ed., Leach et al. eds.

(2007), pages 289-299). Other organic and inorganic pigments and dyes and combinations thereof can be used to achieve the colors desired.

[0084] In addition to or in place of visible colorants, compositions provided herein can contain ETV fluorophores that are excited in the ETV range and emit light at a higher wavelength (typically 400 nm and above). Examples of ETV fluorophores include but are not limited to materials from the coumarin, benzoxazole, rhodamine, napthalimide, perylene, benzanthrones, benzoxanthones or benzothia-xanthones families. The addition of a UV fluorophore (such as an optical brightener for instance) can help maintain maximum visible light transmission. The amount of colorant, when present, generally is between 0.05% to 5% or between 0.1% and 1% based on the weight of the composition.

[0085] For non- white compositions, the amount of pigment/dye generally is present in an amount of from at or about 0.1 wt% to at or about 20 wt% based on the weight of the composition. In some applications, a non-white ink can include 15 wt% or less pigment/dye, or 10 wt% or less pigment/dye or 5 wt% pigment/dye, or 1 wt% pigment/dye based on the weight of the composition. In some applications, a non-white ink can include 1 wt% to 10 wt%, or 5 wt% to 15 wt%, or 10 wt% to 20 wt% pigment/dye based on the weight of the composition. In some applications, a non-white ink can contain an amount of dye/pigment that is 1 wt%, 2 wt%, 3 wt%, 4 wt%, 5%, 6 wt%, 7 wt%, 8 wt%, 9 wt%, 10 wt%, 11 wt%, 12 wt%, 13 wt%, 14 wt%, 15%, 16 wt%, 17 wt%, 18 wt%, 19 wt% or 20 wt% based on the weight of the composition.

[0086] For white compositions, the amount of white pigment generally is present in an amount of from at or about 1 wt% to at or about 60 wt% based on the weight of the composition. In some applications, greater than 60 wt% white pigment can be present. Preferred white pigments include titanium dioxide (anatase and rutile), zinc oxide, lithopone (calcined coprecipitate of barium sulfate and zinc sulfide), zinc sulfide, blanc fixe and alumina hydrate and combinations thereof, although any of these can be combined with calcium carbonate. In some applications, a white ink can include 60 wt% or less white pigment, 55 wt% or less white pigment, 50 wt% white pigment, 45 wt% white pigment, 40 wt% white pigment, 35 wt% white pigment, 30 wt% white pigment, 25 wt% white pigment, 20 wt% white pigment, 15 wt% white pigment, or 10 wt% white pigment, based on the weight of the composition. In some applications, a white ink can include 5 wt% to 60 wt%, 5 wt% to 55 wt%, 10 wt% to 50 wt%, 10 wt% to 25 wt%, 25 wt% to 50 wt%, 5 wt% to 15 wt%, or 40 wt% to 60 wt% white pigment based on the weight of the composition. In some applications, a non-white ink can an amount of dye/pigment that is 5%, 6 wt%, 7 wt%, 8 wt%, 9 wt%, 10 wt%, 11 wt%, 12 wt%, 13 wt%, 14 wt%, 15%, 16 wt%, 17 wt%, 18 wt%, 19 wt%, 20 wt%, 21 wt%, 22 wt%, 23 wt%, 24 wt%, 25%, 26 wt%, 27 wt%, 28 wt%, 29 wt%, 30 wt%, 31 wt%, 32 wt%, 33 wt%, 34 wt%, 35%, 36 wt%, 37 wt%, 38 wt%, 39 wt%, 40 wt%, 41 wt%, 42 wt%, 43 wt%, 44 wt%, 45%, 46 wt%, 47 wt%, 48 wt%, 49 wt%, 50 wt%, 51 wt%, 52 wt%, 53 wt%, 54 wt%, 55%, 56 wt%, 57 wt%, 58 wt%, 59 wt% or 60 wt% based on the weight of the composition.

[0087] In some aspects, the additive or dopant comprises a conductive additive. Exemplary conductive additives include, but are not limited to graphite, graphite powder, carbon nanotubes, and metallic particles or nanoparticles, such as gold nanoparticles. In some aspects, the conductive additive is biocompatible and non-toxic.

[0088] In some aspects, the additive is a biologically active agent. The term “biologically active agent” as used herein refers to any molecule which exerts at least one biological effect in vivo. For example, the biologically active agent can be a therapeutic agent to treat or prevent a disease state or condition in a subject. Biologically active agents include, without limitation, organic molecules, inorganic materials, proteins, peptides, nucleic acids (e.g., genes, gene fragments, gene regulatory sequences, and antisense molecules), nucleoproteins, polysaccharides, glycoproteins, and lipoproteins. Classes of biologically active compounds that can be incorporated into the composition provided herein include, without limitation, anticancer agents, antibiotics, analgesics, antiinflammatory agents, immunosuppressants, enzyme inhibitors, antihistamines, anti-convulsants, hormones, muscle relaxants, antispasmodics, ophthalmic agents, prostaglandins, anti-depressants, anti-psychotic substances, trophic factors, osteoinductive proteins, growth factors, and vaccines. [0089] The term “active agent” may also be used herein to refer to a biological sample (e.g., a sample of tissue or fluid, such as for instance blood) or a component thereof, and/or to a biologically active entity or compound, and/or to a structurally or functionally labile entity.

[0090] Exemplary active agents include, but are not limited to, therapeutic agents, diagnostic agents (e.g., contrast agents), and any combinations thereof. In some embodiments, the active agent present in a silk matrix (e.g., a silk microsphere), composition, or the like can include a labile active agent, e.g., an agent that can undergo chemical, physical, or biological change, degradation and/or deactivation after exposure to a specified condition, e.g., high temperatures, high humidity, light exposure, and any combinations thereof. In some embodiments, the active agent present in the silk matrix (e.g., a silk microsphere), composition, or the like can include a temperature-sensitive active agent, e.g., an active agent that will lose at least about 30% or more, of its original activity or bioactivity, upon exposure to a temperature of at least about 10 °C. or above, including at least about 15 °C. or above, at least about room temperature or above, or at least about body temperature (e.g., about 37 °C.) or above.

[0091] The active agent can be generally present in the silk matrix (e.g., a silk microsphere), composition, or the like in an amount of about 0.01% (w/w) to about 70% (w/w), about 0.1% (w/w) to about 50% (w/w), or about 1% (w/w) to about 30% (w/w). The active agent can be present on a surface of the silk matrix (e.g., a silk microsphere), composition, or the like and/or encapsulated and dispersed in the silk matrix (e.g., a silk microsphere), composition, or the like homogeneously, heterogeneously, or in a gradient. In some embodiments, the active agent can be added into the silk solution, which is then subjected to the methods described herein for preparing a silk matrix (e.g., a silk microsphere), composition, or the like. In some embodiments, the active agent can be coated on a surface of the silk matrix (e.g., a silk microsphere), composition, or the like. In some embodiments, the active agent can be loaded in a silk matrix (e.g., a silk microsphere), composition, or the like by incubating the silk microsphere in a solution of the active agent for a period of time, during which an amount of the active agent can diffuse into the silk matrix (e.g., a silk microsphere), composition, or the like, and thus distribute within the silk matrix (e.g., a silk microsphere), composition, or the like. [0092] In some aspects, the additive is a therapeutic agent. As used herein, the term “therapeutic agent” means a molecule, group of molecules, complex or substance administered to an organism for diagnostic, therapeutic, preventative medical, or veterinary purposes. As used herein, the term “therapeutic agent” includes a “drug” or a “vaccine.” This term includes externally and internally administered topical, localized and systemic human and animal pharmaceuticals, treatments, remedies, nutraceuticals, cosmeceuticals, biologicals, devices, diagnostics and contraceptives, including preparations useful in clinical and veterinary screening, prevention, prophylaxis, healing, wellness, detection, imaging, diagnosis, therapy, surgery, monitoring, cosmetics, prosthetics, forensics and the like. This term can also be used in reference to agriceutical, workplace, military, industrial and environmental therapeutics or remedies comprising selected molecules or selected nucleic acid sequences capable of recognizing cellular receptors, membrane receptors, hormone receptors, therapeutic receptors, microbes, viruses or selected targets comprising or capable of contacting plants, animals and/or humans. This term can also specifically include nucleic acids and compounds comprising nucleic acids that produce a therapeutic effect, for example deoxyribonucleic acid (DNA), ribonucleic acid (RNA), nucleic acid analogues (e.g., locked nucleic acid (LNA), peptide nucleic acid (PNA), xeno nucleic acid (XNA)), or mixtures or combinations thereof, including, for example, DNA nanoplexes, siRNA, microRNA, shRNA, aptamers, ribozymes, decoy nucleic acids, antisense nucleic acids, RNA activators, and the like. Generally, any therapeutic agent can be included in the composition provided herein.

[0093] The term “therapeutic agent” also includes an agent that is capable of providing a local or systemic biological, physiological, or therapeutic effect in the biological system to which it is applied. For example, the therapeutic agent can act to control infection or inflammation, enhance cell growth and tissue regeneration, control tumor growth, act as an analgesic, promote anti-cell attachment, and enhance bone growth, among other functions. Other suitable therapeutic agents can include anti-viral agents, hormones, antibodies, or therapeutic proteins. Other therapeutic agents include prodrugs, which are agents that are not biologically active when administered but upon administration to a subject are converted to biologically active agents through metabolism or some other mechanism. Additionally, a silk-based drug delivery composition can contain one therapeutic agent or combinations of two or more therapeutic agents.

[0094] A therapeutic agent can include a wide variety of different compounds, including chemical compounds and mixtures of chemical compounds, e.g., small organic or inorganic molecules; saccharides; oligosaccharides; polysaccharides; biological macromolecules, e.g., peptides, proteins, and peptide analogs and derivatives; peptidomimetics; antibodies and antigen binding fragments thereof; nucleic acids; nucleic acid analogs and derivatives; an extract made from biological materials such as bacteria, plants, fungi, or animal cells; animal tissues; naturally occurring or synthetic compositions; and any combinations thereof. In some aspects, the therapeutic agent is a small molecule.

[0095] The term “bioactivity,” as used herein in reference to an active agent, generally refers to the ability of an active agent to interact with a biological target and/or to produce an effect on a biological target. For example, bioactivity can include, without limitation, elicitation of a stimulatory, inhibitory, regulatory, toxic or lethal response in a biological target. The biological target can be a molecule or a cell. For example, a bioactivity can refer to the ability of an active agent to modulate the effect/activity of an enzyme, block a receptor, stimulate a receptor, modulate the expression level of one or more genes, modulate cell proliferation, modulate cell division, modulate cell morphology, or any combination thereof. In some instances, a bioactivity can refer to the ability of a compound to produce a toxic effect in a cell. Exemplary cellular responses include, but are not limited to, lysis, apoptosis, growth inhibition, and growth promotion; production, secretion, and surface expression of a protein or other molecule of interest by the cell; membrane surface molecule activation including receptor activation; transmembrane ion transports; transcriptional regulations; changes in viability of the cell; changes in cell morphology; changes in presence or expression of an intracellular component of the cell; changes in gene expression or transcripts; changes in the activity of an enzyme produced within the cell; and changes in the presence or expression of a ligand and/or receptor (e.g., protein expression and/or binding activity). Methods for assaying different cellular responses are well known to one of skill in the art, e.g., western blot for determining changes in presence or expression of an endogenous protein of the cell, or microscopy for monitoring the cell morphology in response to the active agent, or FISH and/or qPCR for the detection and quantification of changes in nucleic acids. Bioactivity can be determined in some embodiments, for example, by assaying a cellular response.

[0096] In reference to an antibody, the term “bioactivity” includes, but is not limited to, epitope or antigen binding affinity, the in vivo and/or in vitro stability of the antibody, the immunogenic properties of the antibody, e.g., when administered to a human subject, and/or the ability to neutralize or antagonize the bioactivity of a target molecule in vivo or in vitro. The aforementioned properties or characteristics can be observed or measured using art-recognized techniques including, but not limited to, scintillation proximity assays, ELISA, ORIGEN immunoassay (IGEN), fluorescence quenching, fluorescence ELISA, competitive ELISA, SPR analysis including, but not limited to, SPR analysis using a BIAcore biosensor, in vitro and in vivo neutralization assays (see, for example, International Publication No. WO 2006/062685), receptor binding, and immunohistochemistry with tissue sections from different sources including human, primate, or any other source as needed. In reference to an immunogen, the “bioactivity” includes immunogenicity, the definition of which is discussed in detail later. In reference to a virus, the “bioactivity” includes infectivity, the definition of which is discussed in detail later. In reference to a contrast agent, e.g., a dye, the “bioactivity” refers to the ability of a contrast agent when administered to a subject to enhance the contrast of structures or fluids within the subject's body. The bioactivity of a contrast agent also includes, but is not limited to, its ability to interact with a biological environment and/or influence the response of another molecule under certain conditions.

[0097] As used herein, the term “small molecule” can refer to compounds that are “natural productlike,” however, the term “small molecule” is not limited to “natural product-like” compounds. Rather, a small molecule is typically characterized in that it contains several carbon — carbon bonds and has a molecular weight of less than 5000 Daltons (5 kDa), preferably less than 3 kDa, still more preferably less than 2 kDa, and most preferably less than 1 kDa. In some cases, it is preferred that a small molecule has a molecular weight equal to or less than 700 Daltons.

[0098] Exemplary therapeutic agents include, but are not limited to, those found in Harrison’s Principles of Internal Medicine, 13th Edition, Eds. T.R. Harrison et al. McGraw-Hill N.Y., NY; Physicians’ Desk Reference, 50th Edition, 1997, Oradell New Jersey, Medical Economics Co.; Pharmacological Basis of Therapeutics, 8th Edition, Goodman and Gilman, 1990; United States Pharmacopeia, The National Formulary, ETSP XII NF XVII, 1990, the complete contents of all of which are incorporated herein by reference.

[0099] Therapeutic agents include the herein disclosed categories and specific examples. It is not intended that the category be limited by the specific examples. Those of ordinary skill in the art will recognize also numerous other compounds that fall within the categories and that are useful according to the present disclosure. Examples include a radiosensitizer, a steroid, a xanthine, a beta- 2-agonist bronchodilator, an anti-inflammatory agent, an analgesic agent, a calcium antagonist, an angiotensin-converting enzyme inhibitors, a beta-blocker, a centrally active alpha- agonist, an alpha - 1 -antagonist, an anticholinergic/antispasmodic agent, a vasopressin analogue, an antiarrhythmic agent, an antiparkinsonian agent, an antiangina/antihypertensive agent, an anticoagulant agent, an antiplatelet agent, a sedative, an anxiolytic agent, a peptidic agent, a biopolymeric agent, an antineoplastic agent, a laxative, an antidiarrheal agent, an antimicrobial agent, an antifungal agent, a vaccine, a protein, or a nucleic acid. In a further aspect, the pharmaceutically active agent can be coumarin, albumin, steroids such as betamethasone, dexamethasone, methylprednisolone, prednisolone, prednisone, triamcinolone, budesonide, hydrocortisone, and pharmaceutically acceptable hydrocortisone derivatives; xanthines such as theophylline and doxophylline; beta-2- agonist bronchodilators such as salbutamol, fenterol, clenbuterol, bambuterol, and salmeterol; antiinflammatory agents, including antiasthmatic anti-inflammatory agents, antiarthritis antiinflammatory agents, and non-steroidal anti-inflammatory agents, examples of which include but are not limited to sulfides, mesalamine, budesonide, salazopyrin, diclofenac, pharmaceutically acceptable diclofenac salts, nimesulide, naproxen, acetaminophen, ibuprofen, ketoprofen and piroxicam; analgesic agents such as salicylates; calcium channel blockers such as nifedipine, amlodipine, and nicardipine; angiotensin converting enzyme inhibitors such as captopril, benazepril hydrochloride, fosinopril sodium, trandolapril, ramipril, lisinopril, enalapril, quinapril hydrochloride, and moexipril hydrochloride; beta-blockers (i.e., beta adrenergic blocking agents) such as sotalol hydrochloride, timolol maleate, esmolol hydrochloride, carteolol, propanolol hydrochloride, betaxolol hydrochloride, penbutolol sulfate, metoprolol tartrate, metoprolol succinate, acebutolol hydrochloride, atenolol, pindolol, and bisoprolol fumarate; centrally active alpha-2-agonists such as clonidine; alpha- 1 -antagonists such as doxazosin and prazosin; anticholinergic/antispasmodic agents such as dicyclomine hydrochloride, scopolamine hydrobromide, glycopyrrolate, clidinium bromide, flavoxate, and oxybutynin; vasopressin analogues such as vasopressin and desmopressin; antiarrhythmic agents such as quinidine, lidocaine, tocainide hydrochloride, mexiletine hydrochloride, digoxin, verapamil hydrochloride, propafenone hydrochloride, flecainide acetate, procainamide hydrochloride, moricizine hydrochloride, and disopyramide phosphate; antiparkinsonian agents, such as dopamine, L-Dopa/Carbidopa, selegiline, dihydroergocryptine, pergolide, lisuride, apomorphine, and bromocriptine; antiangina agents and antihypertensive agents such as isosorbide mononitrate, isosorbide dinitrate, propranolol, atenolol and verapamil; anticoagulant and antiplatelet agents such as coumadin, warfarin, acetylsalicylic acid, and ticlopidine; sedatives such as benzodiazepines and barbiturates; anxiolytic agents such as lorazepam, bromazepam, and diazepam; peptidic and biopolymeric agents such as calcitonin, leuprolide and other LHRH agonists, hirudin, cyclosporin, insulin, somatostatin, protirelin, interferon, desmopressin, somatotropin, thymopentin, pidotimod, erythropoietin, interleukins, melatonin, granulocyte/macrophage-CSF, and heparin; antineoplastic agents such as etoposide, etoposide phosphate, cyclophosphamide, methotrexate, 5 -fluorouracil, vincristine, doxorubicin, cisplatin, hydroxyurea, leucovorin calcium, tamoxifen, flutamide, asparaginase, altretamine, mitotane, and procarbazine hydrochloride; laxatives such as senna concentrate, casanthranol, bisacodyl, and sodium picosulphate; antidiarrheal agents such as difenoxin hydrochloride, loperamide hydrochloride, furazolidone, diphenoxylate hydrochloride, and microorganisms; vaccines such as bacterial and viral vaccines; antimicrobial agents such as penicillins, cephalosporins, and macrolides, antifungal agents such as imidazolic and triazolic derivatives; and nucleic acids such as DNA sequences encoding for biological proteins, and antisense oligonucleotides.

[0100] Anti-cancer agents include alkylating agents, platinum agents, antimetabolites, topoisomerase inhibitors, antitumor antibiotics, antimitotic agents, aromatase inhibitors, thymidylate synthase inhibitors, DNA antagonists, farnesyltransferase inhibitors, pump inhibitors, histone acetyltransferase inhibitors, metalloproteinase inhibitors, ribonucleoside reductase inhibitors, TNF alpha agonists/antagonists, endothelin A receptor antagonists, retinoic acid receptor agonists, immuno-modulators, hormonal and antihormonal agents, photodynamic agents, and tyrosine kinase inhibitors.

[0101] Antibiotics include aminoglycosides (e.g., gentamicin, tobramycin, netilmicin, streptomycin, amikacin, neomycin), bacitracin, carbapenems (e.g., imipenem/cilastatin), cephalosporins, colistin, methenamine, monobactams (e.g., aztreonam), penicillins (e.g., penicillin G, penicillin V, methicillin, nafcillin, oxacillin, cioxacillin, dicloxacillin, ampicillin, amoxicillin, carbenicillin, ticarcillin, piperacillin, mezlocillin, azlocillin), polymyxin B, quinolones, and vancomycin; and bacteriostatic agents such as chloramphenicol, clindamycin, macrolides (e.g., erythromycin, azithromycin, clarithromycin), lincomycin, nitrofurantoin, sulfonamides, tetracyclines (e.g., tetracycline, doxycycline, minocycline, demeclocy cline), and trimethoprim. Also included are metronidazole, fluoroquinolones, and rifampin. [0102] Enzyme inhibitors are substances which inhibit an enzymatic reaction. Examples of enzyme inhibitors include edrophonium chloride, N-methylphysostigmine, neostigmine bromide, physostigmine sulfate, tacrine, 1 -hydroxymaleate, iodotubercidin, p-bromotetranisole, 10-(alpha- diethyl aminopropionyl) -phenothiazine hydrochloride, calmidazolium chloride, hemicholinium-3,3,5- dinitrocatechol, diacylglycerol kinase inhibitor I, diacylglycerol kinase inhibitor II, 3- phenylpropargylamine, N°-monomethyl-L-arginine acetate, carbidopa, 3 -hydroxybenzylhydrazine, hydralazine, clorgyline, deprenyl, hydroxylamine, iproniazid phosphate, 6-MeO-tetrahydro-9H- pyrido-indole, nialamide, pargyline, quinacrine, semicarbazide, tranylcypromine, N,N- diethylaminoethyl-2,2-diphenylvalerate hydrochloride, 3-isobutyl-l-methylxanthine, papaverine, indomethacin, 2-cyclooctyl-2-hydroxyethylamine hydrochloride, 2,3-dichloro-a-methylbenzylamine (DCMB), 8,9-dichloro-2,3,4,5-tetrahydro-lH-2-benzazepine hydrochloride, p-aminoglutethimide, p- aminoglutethimide tartrate, 3-iodotyrosine, alpha-methyltyrosine, acetazolamide, dichlorphenamide, 6-hydroxy-2-benzothiazolesulfonamide, and allopurinol.

[0103] Antihistamines include pyrilamine, chlorpheniramine, and tetrahydrozoline, among others. [0104] Anti-inflammatory agents include corticosteroids, nonsteroidal anti-inflammatory drugs (e.g., aspirin, phenylbutazone, indomethacin, sulindac, tolmetin, ibuprofen, piroxicam, and fenamates), acetaminophen, phenacetin, gold salts, chloroquine, D-Penicillamine, methotrexate colchicine, allopurinol, probenecid, and sulfinpyrazone.

[0105] Muscle relaxants include mephenesin, methocarbamol, cyclobenzaprine hydrochloride, trihexyphenidyl hydrochloride, levodopa/carbidopa, and biperiden.

[0106] Anti-spasmodics include atropine, scopolamine, oxyphenonium, and papaverine.

[0107] Analgesics include aspirin, phenylbutazone, indomethacin, sulindac, tolmetin, ibuprofen, piroxicam, fenamates, acetaminophen, phenacetin, morphine sulfate, codeine sulfate, meperidine, nalorphine, opioids (e.g., codeine sulfate, fentanyl citrate, hydrocodone bitartrate, loperamide, morphine sulfate, noscapine, norcodeine, normorphine, thebaine, nor-binaltorphimine, buprenorphine, chlomaltrexamine, funaltrexamine, nalbuphine, nalorphine, naloxone, naloxonazine, naltrexone, and naltrindole), procaine, lidocaine, tetracaine and dibucaine. Ophthalmic agents include sodium fluorescein, rose bengal, methacholine, adrenaline, cocaine, atropine, alphachymotrypsin, hyaluronidase, betaxolol, pilocarpine, timolol, timolol salts, and combinations thereof. [0108] Prostaglandins are art recognized and are a class of naturally occurring chemically related long -chain hydroxy fatty acids that have a variety of biological effects.

[0109] Anti-depressants are substances capable of preventing or relieving depression.

[0110] Examples of anti-depressants include imipramine, amitriptyline, nortriptyline, protriptyline, desipramine, amoxapine, doxepin, maprotiline, tranylcypromine, phenelzine, and isocarboxazid. [0111] Trophic factors are factors whose continued presence improves the viability or longevity of a cell trophic factors include, without limitation, platelet-derived growth factor (PDGP), neutrophilactivating protein, monocyte chemoattractant protein, macrophage- inflammatory protein, platelet factor, platelet basic protein, and melanoma growth stimulating activity; epidermal growth factor, transforming growth factor (alpha), fibroblast growth factor, platelet- derived endothelial cell growth factor, insulin-like growth factor, glial derived growth neurotrophic factor, ciliary neurotrophic factor, nerve growth factor, bone growth/cartilage-inducing factor (alpha and beta), bone morphogenetic proteins, interleukins (e.g., interleukin inhibitors or interleukin receptors, including interleukin 1 through interleukin 10), interferons (e.g., interferon alpha, beta and gamma), hematopoietic factors, including erythropoietin, granulocyte colony stimulating factor, macrophage colony stimulating factor and granulocyte-macrophage colony stimulating factor; tumor necrosis factors, and transforming growth factors (beta), including beta-1, beta-2, beta-3, inhibin, and activin. [0112] Hormones include estrogens (e.g., estradiol, estrone, estriol, diethylstilbestrol, quinestrol, chlorotrianisene, ethinyl estradiol, mestranol), anti-estrogens (e.g., clomiphene, tamoxifen), progestins (e.g., medroxyprogesterone, norethindrone, hydroxyprogesterone, norgestrel), antiprogestin (mifepristone), androgens (e.g., testosterone cypionate, fluoxymesterone, danazol, testolactone), anti-androgens (e.g., cyproterone acetate, flutamide), thyroid hormones (e.g., triiodothyronne, thyroxine, propylthiouracil, methimazole, and iodixode), and pituitary hormones (e.g., corticotropin, somatotropin, oxytocin, and vasopressin). Hormones are commonly employed in hormone replacement therapy and/or for purposes of birth control. Steroid hormones, such as prednisone, are also used as immunosuppressants and anti-inflammatories. In some aspects, the additive is an agent that stimulates tissue formation, and/or healing and regrowth of natural tissues, and any combinations thereof. Agents that increase formation of new tissues and/or stimulates healing or regrowth of native tissue at the site of injection can include, but are not limited to, fibroblast growth factor (FGF), transforming growth factor- beta (TGF-beta, platelet-derived growth factor (PDGF), epidermal growth factors (EGFs), connective tissue activated peptides (CTAPs), osteogenic factors including bone morphogenic proteins, heparin, angiotensin II (A-II) and fragments thereof, insulin-like growth factors, tumor necrosis factors, interleukins, colony stimulating factors, erythropoietin, nerve growth factors, interferons, biologically active analogs, fragments, and derivatives of such growth factors, and any combinations thereof.

[0113] In some aspects, the silk composition can further comprise at least one additional material for soft tissue augmentation, e.g., dermal filler materials, including, but not limited to, poly(methyl methacrylate) microspheres, hydroxyapatite, poly(L-lactic acid), collagen, elastin, and glycosaminoglycans, hyaluronic acid, commercial dermal filler products such as BOTOX® (from Allergan), DYSPORT®, COSMODERM®, EVOLENCE®, RADIESSE®,RESTYLANE®, JUVEDERM® (from Allergan), SCULPTRA®, PERLANE®, and CAPTIQEIE®, and any combinations thereof.

[0114] In some aspects, the additive is a wound healing agent. As used herein, a “wound healing agent" is a compound or composition that actively promotes wound healing process.

[0115] Exemplary wound healing agents include, but are not limited to dexpanthenol; growth factors; enzymes; hormones; povidon-iodide; fatty acids; anti-inflammatory agents; antibiotics; antimicrobials; antiseptics; cytokines; thrombin; angalgesics; opioids; aminoxyls; furoxans; nitrosothiols; nitrates and anthocyanins; nucleosides, such as adenosine; and nucleotides, such as adenosine diphosphate (ADP) and adenosine triphosphate (ATP); neurotransmitter/neuromodulators, such as acetylcholine and 5 -hydroxy tryptamine (serotonin/5-HT); histamine and catecholamines, such as adrenalin and noradrenalin; lipid molecules, such as 5 -sphingosine- 1 -phosphate and lysophosphatidic acid; amino acids, such as arginine and lysine; peptides such as the bradykinins, substance P and calcium gene-related peptide (CGRP); nitric oxide; and any combinations thereof. [0116] In certain aspects, the active agents provided herein are immunogens. In one aspect, the immunogen is a vaccine. Most vaccines are sensitive to environmental conditions under which they are stored and/or transported. For example, freezing may increase reactogenicity (e.g., capability of causing an immunological reaction) and/or loss of potency for some vaccines (e.g., HepB, and DTaP/IPV/FQB), or cause hairline cracks in the container, leading to contamination. Further, some vaccines (e.g., BCG, Varicella, and MMR) are sensitive to heat. Many vaccines (e.g., BCG, MMR, Varicella, Meningococcal C Conjugate, and most DTaP-containing vaccines) are light sensitive. See, e.g., Galazka et al., Thermostability of vaccines, in Global Programme for Vaccines & Immunization (World Health Organization, Geneva, 1998); Peetermans et al., Stability of freeze-dried rubella virus vaccine (Cendehill strain) at various temperatures, J. Biological Standardization 179 (1973). Thus, the compositions and methods provided herein also provide for stabilization of vaccines regardless of the cold chain and/or other environmental conditions.

[0117] In some aspects, the additive is a cell, e.g., a biological cell. Cells useful for incorporation into the composition can come from any source, e.g., mammalian, insect, plant, etc. In some aspects, the cell can be a human, rat or mouse cell. In general, cells to be used with the compositions provided herein can be any types of cells. In general, the cells should be viable when encapsulated within compositions. In some aspects, cells that can be used with the composition include, but are not limited to, mammalian cells (e.g. human cells, primate cells, mammalian cells, rodent cells, etc.), avian cells, fish cells, insect cells, plant cells, fungal cells, spore cells, bacterial cells, and hybrid cells. In some aspects, exemplary cells that can be used with the compositions include platelets, activated platelets, stem cells, totipotent cells, pluripotent cells, and/or embryonic stem cells. In some aspects, exemplary cells that can be encapsulated within compositions include, but are not limited to, primary cells and/or cell lines from any tissue. For example, cardiomyocytes, myocytes, hepatocytes, keratinocytes, melanocytes, neurons, astrocytes, embryonic stem cells, adult stem cells, hematopoietic stem cells, hematopoietic cells (e.g. monocytes, neutrophils, macrophages, etc.), ameloblasts, fibroblasts, chondrocytes, osteoblasts, osteoclasts, neurons, sperm cells, egg cells, liver cells, epithelial cells from lung, epithelial cells from gut, epithelial cells from intestine, liver, epithelial cells from skin, etc., and/or hybrids thereof, can be included in the silk/platelet compositions disclosed herein. Those skilled in the art will recognize that the cells listed herein represent an exemplary, not comprehensive, list of cells. Cells can be obtained from donors (allogenic) or from recipients (autologous). Cells can be obtained, as a non-limiting example, by biopsy or other surgical means known to those skilled in the art.

[0118] In some aspects, the cell can be a genetically modified cell. A cell can be genetically modified to express and secrete a desired compound, e.g. a bioactive agent, a growth factor, differentiation factor, cytokines, and the like. Methods of genetically modifying cells for expressing and secreting compounds of interest are known in the art and easily adaptable by one of skill in the art.

[0119] Differentiated cells that have been reprogrammed into stem cells can also be used.

[0120] For example, human skin cells reprogrammed into embryonic stem cells by the transduction of Oct3/4, Sox2, c-Myc and Klf4 (Junying Yu, et. ah, Science, 2007, 318, 1917-1920 and Takahashi K. et. al, Cell, 2007, 131 , 1-12).

[0121] Unless otherwise specified or indicated by context, the terms “a”, “an”, and “the” mean “one or more.” For example, “a molecule” should be interpreted to mean “one or more molecules”.

[0122] As used herein, “about”, “approximately”, “substantially”, and “significantly” will be understood by persons of ordinary skill in the art and will vary to some extent on the context in which they are used. If there are uses of the term which are not clear to persons of ordinary skill in the art given the context in which it is used, “about” and “approximately” will mean plus or minus <10% of the particular term and “substantially” and “significantly” will mean plus or minus >10% of the particular term.

[0123] As used herein, the terms “include” and “including” have the same meaning as the terms “comprise” and “comprising.” The terms “comprise” and “comprising” should be interpreted as being “open” transitional terms that permit the inclusion of additional components further to those components recited in the claims. The terms “consist” and “consisting of” should be interpreted as being “closed” transitional terms that do not permit the inclusion of additional components other than the components recited in the claims. The term “consisting essentially of’ should be interpreted to be partially closed and allowing the inclusion only of additional components that do not fundamentally alter the nature of the claimed subject matter.

[0124] All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., “such as”) provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0125] All references, including publications, patent applications, and patents, cited herein are hereby incorporated by reference to the same extent as if each reference were individually and specifically indicated to be incorporated by reference and were set forth in its entirety herein.

[0126] Preferred aspects of this invention are described herein, including the best mode known to the inventors for carrying out the invention. Variations of those preferred aspects may become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventors expect a person having ordinary skill in the art to employ such variations as appropriate, and the inventors intend for the invention to be practiced otherwise than as specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the abovedescribed elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

[0127] While the invention has been illustrated and described in detail in the foregoing drawings and description, the same is to be considered as illustrative and not restrictive in character, it being understood that only illustrative embodiments thereof have been shown and described and that all changes and modifications that come within the spirit of the invention are desired to be protected. For example, any of the features or functions of any of the embodiments disclosed herein may be incorporated into any of the other embodiments disclosed herein.

[0128] The following examples illustrate some embodiments and aspects of the invention. It will be apparent to those skilled in the relevant art that various modifications, additions, substitutions, and the like can be performed without altering the spirit or scope of the invention, and such modifications and variations are encompassed within the scope of the invention as defined in the claims which follow. The following examples do not in any way limit the invention.

[0129] EXAMPLES

[0130] Example 1: Nozzle Optimization [0131] The impact of nozzle geometries on material extrusion was investigated as commercially available nozzles were insufficient. Data is shown in Fig. 2 (left) and the nozzles used are shown in Fig. 2 (right). When extruding at a rate of 5 mm/min, extrusion began around 23 kPa for the 1.8 mm nozzle compared to around 30 kPa for the 2.0 mm nozzle. The maximum extrusion pressure for the 1.8 mm nozzle was 28.39 kPa, compared to 32.36 kPa for the 2.0 mm nozzle. The nozzles were developed to facilitate extrusion of the biopolymer-based cellular biomaterial, ensuring the correct amount of shear while preventing clogging. A radius of between 1 mm and 3 mm and a length of between 10 mm and 20 mm was found to be sufficient for some use cases.

[0132] Example 2: Method Development

[0133] A unique computer aided design (CAD) method was developed using Grasshopper 3D (Rhinoceros plug-in) environment along with custom Python components. To manage the compressible foam and its shear-thinning behavior, we developed a slicing algorithm that facilitates single-path printing and enables non-linear compression of the foam during printing. Consequently, the extrusion rate or flow rate (E) is modulated during printing to obtain a 35% reduction. The algorithm also pre-compresses the foam to initiate the material’s flow (transition from solid phase to liquid phase). The algorithm also allows the printing speed to be varied to match the compression state of the foam. We have therefore calculated that a progressive feedrate (volumetric measure of the amount of material extruded in function of time) of 150 mm/min (about 48%) to 310 mm/min (100%) is sufficient for some use cases.

[0134] Example 3: Material Development

[0135] A silk foam with a firmness of around 250 N/m² (within a range of 100 N/m² -400 N/m²) and a density of 0.14 g/cm³ (within a range of 0.26 g/cm³ - 0.47 g/cm³) has been determined to be ideal for certain applications. Experimentation indicates that a foam consisting of 19 + 2% silk fibroin, 57 ± 6% glycerol, 19 ± 2% alginate, xanthan gum, or a combination of both, and 5 + 1% glycerol monostearate works well with the deposition system described herein. Furthermore, we have observed that the inclusion of 20% silk fibroin powder enhances foam stability during printing due to its surfactant properties which is also improved by the addition of glycerol monostearate.

[0136] Through thorough evaluation, the optimal relationship between the physical properties of the cream (e.g., density and firmness), its rheology (e.g., viscosity), and its printability (e.g., resolution of print, printing speed) was identified. Rheological behavior of the material was also studied and characterized (Fig. 3) as a shear-thinning material, specifically a pseudoplastic fluid. Depending on compositions, the yield stress may vary between 30 Pa and 125 Pa. The maximum pressure needed for extrusion may be between 20 kPa to 40 kPa. [0137] EQUIVALENTS AND SCOPE

[0138] The recitation of a listing of elements in any definition of a variable herein includes definitions of that variable as any single element or combinations (or subcombinations) of listed elements. The recitation of an embodiment herein includes that embodiment as any single embodiment or in combination with any other embodiments or portions thereof. Those skilled in the art will recognize or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein.

[0139] In addition to the features described above and elsewhere herein, the present disclosure also includes the following clauses:

[0140] Clause 1. A printhead of a three-dimensional printer comprising: an extrusion nozzle configured to extrude a biopolymer-based cellular biomaterial; a ventilation system, the ventilation system positioned adjacent to the extrusion nozzle, wherein an airflow from the ventilation system is directed to the biopolymer-based cellular biomaterial extruded from the extrusion nozzle to form an extruded biopolymer-based cellular biomaterial; and wherein the printhead is configured to store the biopolymer-based cellular biomaterial under a storage pressure.

[0141] Clause 2. The printhead of clause 1, wherein the storage pressure is greater than atmospheric pressure.

[0142] Clause 3. The printhead of clause 1, wherein the storage pressure is between 0.25 MPa and 25 MPa, including at least 0.5 MPa, at least 1 MPa, at least 5 MPa, at least 10 MPa, or at least 20 MPa.

[0143] Clause 4. The printhead of clause 1 , wherein the biopolymer-based cellular biomaterial has a firmness of between 100 N/m² and 400 N/m², including at least 250 N/m², and a density of between 0.08 g/cm³ and 0.25 g/cm³, including at least 0.12 g/cm³.

[0144] Clause 5. The printhead of clause 1, wherein the printhead comprises a mechanical extrusion system.

[0145] Clause 6. The printhead of clause 5, wherein the mechanical extrusion system comprises a bipolar stepper motor with a gearbox and rear axle configured to operate at a torque of between 1 N-m and 4 N-m, including at least 1.5 N-m, at least 2 N-m, or at least 3 N-m.

[0146] Clause 7. The printhead of clause 6, wherein the bipolar stepper motor enables both clockwise and counterclockwise rotation of a belt and a threaded rod, guiding a piston and facilitating the biopolymer-based cellular biomaterial extrusion.

[0147] Clause 8. The printhead of clause 1, further comprising a vessel for delivering the biopolymer-based cellular biomaterial to the extrusion nozzle. [0148] Clause 9. The printhead of clause 8, wherein the vessel comprises a removable, refillable syringe.

[0149] Clause 10. The printhead of clause 8, wherein the vessel is configured to store a volume of the biopolymer-based cellular biomaterial under the storage pressure.

[0150] Clause 11. The printhead of clause 10, wherein the volume is between 100 mL and 400 mL, including at least 100 mL, at least 200 mL, or at least 300 mL, and at most 400 mL, at most 350 mL, at most 250 mL, and at most 150 mL.

[0151] Clause 12. The printhead of clause 1, wherein the ventilation system comprises at least one fan.

[0152] Clause 13. The printhead of clause 12, wherein the at least one fan is directed towards the biopolymer-based cellular biomaterial exiting the extrusion nozzle to enable layer-by-layer drying of the extruded biopolymer-based cellular biomaterial.

[0153] Clause 14. The printhead of clause 12, wherein an airflow from the at least one fan is directed at an angle with respect to a direction of extrusion.

[0154] Clause 15. The printhead of clause 14, wherein the angle is between 35° and 65°, including at least 40°, at least 42°, at least 45°, or at least 47°.

[0155] Clause 16. The printhead of clause 12, wherein a fan speed of the at least one fan is between 3000 bpm and 4000 bpm, including at least 3100 bpm, at least 3200 bpm, at least 3500 bpm, or at least 3800 bpm.

[0156] Clause 17. The printhead of clause 1, wherein the ventilation system is configured to deliver an airflow at a predetermined fluid velocity.

[0157] Clause 18. The printhead of clause 17, wherein the predetermined fluid velocity is between 4 m/s and 10 m/s, including at least 5 m/s, at least 7 m/s, or at least 9 m/s.

[0158] Clause 19. The printhead of clause 1, wherein the ventilation system is configured to deliver an airflow at a variable fluid velocity.

[0159] Clause 20. The printhead of clause 19, wherein the variable fluid velocity varies with at least one of a change in distance from the ventilation system to a surface of the extruded biopolymerbased cellular biomaterial, an extrusion rate from the extrusion nozzle, a layer height, a length of path, an extrusion width, or a composition of the biopolymer- based cellular biomaterial.

[0160] Clause 21 . The printhead of clause 1, wherein the ventilation system is arranged at a distance of between 60 mm and 80 mm from a tip of the extrusion nozzle, including at least 65 mm, a least 70 mm, or at least 75 mm. [0161] Clause 22. The printhead of clause 1, wherein the ventilation system is arranged at a distance of between 60 mm and 100 mm from the extruded biopolymer-based cellular biomaterial, including at least 65 mm, at least 80 mm, or at least 95 mm.

[0162] Clause 23. The printhead of clause 1, wherein the printhead extrudes the biopolymer-based cellular biomaterial in accordance with an extrusion rate defined by Q= co x h x v (where co = extrusion width, h = layer height, v = velocity of extrusion).

[0163] Clause 24. The printhead of clause 23, wherein the extrusion rate is between 125 mm/min and 350 mm/min, including at least 265 mm/min, at least 290 mm/min, at least 310 mm/min, and at least 330 mm/min, or at most 310 mm/min, at most 250 mm/min, at most 200 mm/min, or at most 150 mm/min.

[0164] Clause 25. The printhead of clause 23, wherein the extrusion rate is further defined by a rate of volume change of the extruded biopolymer-based cellular biomaterial.

[0165] Clause 26. The printhead of clause 23, wherein the extrusion rate varies by up to 45% of an original value of the extrusion rate, including up to 75%, up to 65%, up to 60%, up to 55%, up to 48%, or up to 44%.

[0166] Clause 27. A process for generating a biopolymer-based cellular biomaterial, the process comprising: flowing a biopolymer-based cellular biomaterial under pressure through a printhead of a three-dimensional printer, the biopolymer-based cellular biomaterial comprising silk fibroin in an amount by weight of between 1% and 40%, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1 % and 10% , at least one polysaccharide in an amount by weight of between 10% and 30%, and at least one plasticizer in an amount by weight of between 20% and 75%; extruding the biopolymer-based cellular biomaterial through an extrusion nozzle of the printhead to form an extruded biopolymer-based cellular biomaterial, the extruding in accordance with a volumetric extrusion rate (Q) defined by Q = co x h x v (where co = extrusion width, h = layer height, v = velocity of extrusion); and drying the extruded biopolymer-based cellular biomaterial exiting the extrusion nozzle.

[0167] Clause 28. The process of clause 27, wherein drying comprises directing an airflow from a ventilation system adjacent to the extrusion nozzle towards the extruded biopolymer-based cellular biomaterial exiting the extrusion nozzle.

[0168] Clause 29. The process of clause 27, wherein the biopolymer-based cellular biomaterial further comprises 10% sodium alginate and 10% xanthan gum.

[0169] Clause 30. The process of clause 27, wherein the plasticizer is glycerol.

[0170] Clause 31. A method of making a biopolymer-based cellular biomaterial, the method comprising: whipping a liquid composition for a predetermined whipping time at a predetermined speed to form a biopolymer-based cellular biomaterial, the liquid composition comprising silk fibroin, at least one surfactant, at least one polysaccharide, and at least one plasticizer; pressurizing a volume of the biopolymer-based cellular biomaterial; and concurrently extruding and drying the biopolymer-based cellular biomaterial to form an extruded, dried biopolymer-based cellular biomaterial.

[0171] Clause 32. The method of clause 31, wherein the predetermined whipping time is at least 8.5 minutes.

[0172] Clause 33. The method of clause 32, wherein the predetermined speed is at least 185 rpm.

[0173] Clause 34. The method of clause 31, wherein the predetermined whipping time is between 5 minutes and 30 minutes, including but not limited to, at least 5 minutes, at least 6 minutes, at least 7 minutes, at least 8 minutes, at least 9 minutes, or at least 10 minutes and at most 30 minutes, at most 25 minutes, at most 20 minutes, at most 15 minutes, or at most 10 minutes.

[0174] Clause 35. The method of clause 31, wherein the predetermined speed is between 160 rpm and 200 rpm, including but not limited to, at least 160 rpm, at least 170 rpm, at least 175 rpm, or at least 185 rpm, and at most 200 rpm, at most 190 rpm, or at most 180 rpm.

[0175] Clause 36. The method of clause 31, wherein the at least one plasticizer is selected from the group consisting of a di(ethylene glycol), a tri(ethylene glycol), a di(propylene glycol), a tri(propylene glycol), triethanolamine, a vegetable oil, and combinations thereof.

[0176] Clause 37. The method of clause 31, wherein the at least one polysaccharide is selected from the group consisting of xanthan gum, an alginate, a high molecular weight sugar, a cellulose derivative, or a combination thereof.

[0177] Clause 38. The method of clause 31, wherein the at least one surfactant is a monoglyceride. [0178] Clause 39. The method of clause 31, wherein the at least one surfactant is glycerol monostearate.

[0179] Clause 40. The method of clause 31, wherein the liquid composition comprises silk fibroin, glycerol, sodium alginate, glycerol monostearate, and xanthan gum.

[0180] Clause 41. The printhead, process, or method of any one of the preceding clauses wherein the biopolymer-based cellular biomaterial comprises silk fibroin.

[0181] Clause 42. The printhead, process, or method of any one of the preceding clauses, wherein the biopolymer-based cellular biomaterial comprises silk fibroin, at least one plasticizer, at least one surfactant, and at least one polysaccharide.

[0182] Clause 43. The printhead, process, or method of clause 42, wherein the at least one polysaccharide is xanthan gum, an alginate, a high molecular weight sugar, a cellulose derivative, or a combination thereof. [0183] Clause 44. The printhead, process, or method of the immediately preceding clause, wherein the alginate is alginic acid sodium salt.

[0184] Clause 45. The printhead, process, or method of clause 42, wherein the at least one plasticizer is selected from the group consisting of a glycerol, a di(ethylene glycol), a tri(ethylene glycol), a di(propylene glycol), a tri(propylene glycol), triethanolamine, a vegetable oil, and combinations thereof.

[0185] Clause 46. The printhead, process, or method of clause 40, wherein the at least one plasticizer comprises at least one -OH substituent.

[0186] Clause 47. The printhead, process, or method of clause 42, wherein the at least one surfactant is a monoglyceride.

[0187] Clause 48. The printhead, process, or method of clause 42, wherein the at least one surfactant is glycerol monostearate.

[0188] Clause 49. The printhead, process, or method of clause 42, wherein the at least one plasticizer is present in the biopolymer-based cellular biomaterial in an amount by weight of between 20.0% and 75.0%, including but not limited to at least 30%, at least 40%, at least 50%, or at least 60%, or at most 70%, at most 65%, at most 55%, or at most 45%.

[0189] Clause 50 The printhead, process, or method of any one of the preceding clauses, wherein the biopolymer-based cellular biomaterial has an irregular porosity.

[0190] Clause 51. The printhead, process, or method of any one of clauses 1 to 49, wherein the biopolymer-based cellular biomaterial has a regular porosity.

[0191] Clause 52. The printhead, process, or method of any one of the preceding clauses, wherein the biopolymer-based cellular biomaterial has a firmness of at least 100 N/m², at least 200 N/m², at least 250 N/m², at least 300 N/m², or at least 400 N/m².

[0192] Clause 53. The printhead, process, or method of any of the preceding clauses, wherein the biopolymer-based cellular biomaterial has a density of at least 0.08 g/cm³, at least 0.1 g/cm³, at least 0.12 g/cm³, at least 0.2 g/cm³, or at least 0.25 g/cm³.

[0193] Clause 54. The printhead, process, or method of any one of the preceding clauses, wherein the biopolymer-based cellular biomaterial comprises silk fibroin in an amount by weight of between 1% and 40%, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1% and 10%, sodium alginate in an amount by weight of between 1% and 20%, and xanthan gum in an amount by weight of between 1% and 20%.

[0194] Clause 55. The printhead, process, or method of any one of the preceding clauses, wherein the silk fibroin is present in the biopolymer-based cellular biomaterial in an amount by weight of between 10% and 30% or between 15% and 25%, including but not limited to, at least 15%, at least 17%, at least 19%, at least 21%, or at least 23% and at most 24%, at most 22%, at most 20%, at most 18%, or at most 16%.

[0195] Clause 56. The printhead, process, or method of clause 42, wherein the at least one polysaccharide is present in the biopolymer-based cellular biomaterial in an amount by weight of between 0.1% and 30.0%, including but not limited to, at least 0.1%, at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, or at least 25%, and at most 30%, at most 24%, at most 21%, at most 18%, at most 12%, or at most 6%.

[0196] Clause 57. The printhead, process, or method of clause 42, wherein the silk fibroin and the at least one polysaccharide are present in the biopolymer-based cellular biomaterial in a weight ratio of between 1 :4 and 20: 1 or between 1 :2 and 10: 1 , including but not limited to, at least 1 :4, at least 1:3, or at least 1:2, and at most 20:1, at most 19:1, at most 18:1, at most 16:1, at most 15: 1, at most 14: 1, at most 12: 1, at most 11: 1, or at most 10:1.

[0197] Clause 58. The printhead, process, or method of any one of the preceding clauses, wherein the biopolymer-based cellular biomaterial further comprises at least one of a sensing agent, a therapeutically active agent, a colorant, or an aroma-providing compound.

[0198] Clause 59. The printhead, process, or method of any of the preceding clauses, wherein the biopolymer-based cellular biomaterial is extruded in a single path.

[0199] Clause 60. The printhead, process, or method of clause 40, wherein the surfactant is a monoglyceride.

[0200] Clause 61. The printhead, process, or method of clause 40, wherein the surfactant is glycerol monostearate.

[0201] Clause 62. The printhead, process, or method of clause 40, wherein the surfactant is present by weight in an amount between 1% and 10%, including but not limited to, at least 1%, at least 3%, at least 5%, at least 7%, or at least 9%, and at most 10%, at most 8%, at most 6%, at most 4%, or at most 2%.

[0202] The scope of the present invention is not intended to be limited to the above Description, but rather is as set forth in the following claims:

Claims

CLAIMS What is claimed is:

1. A printhead of a three-dimensional printer comprising: an extrusion nozzle configured to extrude a biopolymer-based cellular biomaterial; a ventilation system, the ventilation system positioned adjacent to the extrusion nozzle, wherein an airflow from the ventilation system is directed to the biopolymer-based cellular biomaterial extruded from the extrusion nozzle to form an extruded biopolymer-based cellular biomaterial; and wherein the printhead is configured to store the biopolymer-based cellular biomaterial under a storage pressure.

2. The printhead of claim 1 , wherein the biopolymer-based cellular biomaterial has a firmness of between 100 N/m² and 400 N/m², including at least 250 N/m², and a density of between 0.08 g/cm³ and 0.25 g/cm³, including at least 0.12 g/cm³.

3. The printhead of claim 1 , wherein the printhead comprises a mechanical extrusion system.

4. The printhead of claim 3, wherein the mechanical extrusion system comprises a bipolar stepper motor with a gearbox and rear axle configured to operate at a torque of between 1 N-m and 4 N-m, including at least 1.5 N-m, at least 2 N-m, or at least 3 N-m.

5. The printhead of claim 4, wherein the bipolar stepper motor enables both clockwise and counterclockwise rotation of a belt and a threaded rod, guiding a piston and facilitating the biopolymer-based cellular biomaterial extrusion.

6. The printhead of claim 1 , further comprising a vessel for delivering the biopolymer-based cellular biomaterial to the extrusion nozzle.

7. The printhead of claim 1 , wherein the ventilation system comprises at least one fan.

8. The printhead of claim 7, wherein the at least one fan is directed towards the biopolymerbased cellular biomaterial exiting the extrusion nozzle to enable layer-by-layer drying of the extruded biopolymer-based cellular biomaterial, wherein the angle is between 35° and 65°, including at least 40°, at least 42°, at least 45°, or at least 47°.

9. The printhead of claim 7, wherein an airflow from the at least one fan is directed at an angle with respect to a direction of extrusion

10. The printhead of claim 1, wherein the ventilation system is configured to deliver an airflow at a predetermined fluid velocity.

1 1. The printhead of claim 1 , wherein the ventilation system is arranged at a distance of between 60 mm and 80 mm from a tip of the extrusion nozzle, including at least 65 mm, at least 70 mm, or at least 75 mm.

12. The printhead of claim 1, wherein the biopolymer-based cellular biomaterial comprises silk fibroin in an amount by weight of between 1% and 40%, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1% and 10%, sodium alginate in an amount by weight of between

1 % and 20%, and xanthan gum in an amount by weight of between 1 % and 20%.

13. The printhead of claim 1 , wherein the silk fibroin is present in the biopolymer-based cellular biomaterial in an amount by weight of between 10% and 30% or between 15% and 25%, including but not limited to, at least 15%, at least 17%, at least 19%, at least 21%, or at least 23% and at most 24%, at most 22%, at most 20%, at most 18%, or at most 16%.

14. The printhead of claim 1 , wherein the at least one polysaccharide is present in the biopolymer-based cellular biomaterial in an amount by weight of between 0.1% and 30.0%, including but not limited to, at least 0.1%, at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, or at least 25%, and at most 30%, at most 24%, at most 21%, at most 18%, at most 12%, or at most 6%.

15. The printhead of claim 1 , wherein the at least one polysaccharide is xanthan gum, an alginate, a high molecular weight sugar, a cellulose derivative, or a combination thereof.

16. The printhead of claim 1 , wherein the surfactant is present by weight in an amount between 1% and 10%, including but not limited to, at least 1%, at least 3%, at least 5%, at least 7%, or at least 9%, and at most 10%, at most 8%, at most 6%, at most 4%, or at most 2%.

17. The printhead of claim 1, wherein the surfactant is a monoglyceride.

18. The printhead of claim 1, wherein the surfactant is a glycerol monostearate.

19. A process for generating a biopolymer-based cellular biomaterial, the process comprising: flowing a biopolymer-based cellular biomaterial under pressure through a printhead of a three-dimensional printer, the biopolymer-based cellular biomaterial comprising silk fibroin in an amount by weight of between 1% and 40%, a surfactant (e.g., glycerol monostearate) in an amount by weight of between 1 % and 10% , at least one polysaccharide in an amount by weight of between 10% and 30%, and at least one plasticizer in an amount by weight of between 20% and 75%; extruding the biopolymer-based cellular biomaterial through an extrusion nozzle of the printhead to form an extruded biopolymer-based cellular biomaterial, the extruding in accordance with a volumetric extrusion rate (Q) defined by Q = ® x h x v (where co = extrusion width, h = layer height, v = velocity of extrusion); and drying the extruded biopolymer-based cellular biomaterial exiting the extrusion nozzle.

20. A method of making a biopolymer-based cellular biomaterial, the method comprising: whipping a liquid composition for a predetermined whipping time at a predetermined speed to form a biopolymer-based cellular biomaterial, the liquid composition comprising silk fibroin, at least one surfactant, at least one polysaccharide, and at least one plasticizer; pressurizing a volume of the biopolymer-based cellular biomaterial; and concurrently extruding and drying the biopolymer-based cellular biomaterial to form an extruded, dried biopolymer-based cellular biomaterial.