The use of three-dimensional printers in postbiotic encapsulation

Abstract

The present invention relates to the encapsulation of postbiotics by three-dimensional printing in order to prepare them in a form suitable for the field of use. The method developed by the invention is essentially based on the deposition of a composition comprising at least one postbiotic material in a texture, shape and size determined according to the field of use by a three-dimensional printing method. Embodiments of the invention realized with various three-dimensional printing techniques are described. Within the scope of the invention, thanks to the use of three-dimensional printing techniques, encapsulated postbiotics whose macro- and micro-structural properties can be easily adapted to the site of use can be produced and the postbiotics can be delivered to the site of use without being affected by temperature, pH and enzymes, paving the way for high utilization.

Description

[0001] THE USE OF THREE-DIMENSIONAL PRINTERS IN POSTBIOTIC ENCAPSULATION

[0002] Technical Field

[0003] This invention relates to the encapsulation of postbiotics by three-dimensional printing in order to prepare them in a suitable form for the field of use.

[0004] Prior Art

[0005] In "Prebiotics and postbiotics synergistic delivery microcapsules from microfluidics for treating colitis" (Yang, K., Wang, X., Huang, R., Wang, H., Lan, P., & Zhao, Y. (2022). Advanced Science, 9(16), 2104089.), for the encapsulation of prebiotics and postbiotics, the encapsulation of a mixture containing indole-3 -propionic acid, postbiotics and prebiotics including sodium alginate, resistant starch and chitosan by microfluidic electrospray technique is described and demonstrated.

[0006] In the document entitled "The Effect of Encapsulating a Prebiotic-Based Biopolymer Delivery System for Enhanced Probiotic Survival" (Kistaubayeva, A., Abdulzhanova, M., Zhantlessova, S., Savitskaya, I., Karpenyuk, T., Goncharova, A., & Sinyavskiy, Y. (2023). Polymers, 15(7), 1752.), a solution based on the use of biopolymers for the encapsulation of prebiotics is described and its contribution to probiotic efficacy is demonstrated.

[0007] Document numbered EP3881835A1 discloses the encapsulation of bioactive materials, including prebiotic or probiotic materials, by various techniques including inkjet and three- dimensional printing.

[0008] Document numbered WO2023147518A1 discloses three-dimensional printing systems and methods for the production of foodstuffs. This utilizes the deposition of a bioink containing a biopolymer in forms including aqueous solution or aqueous suspension. It is also indicated that the bioink may contain a bioactive material or probiotic in an encapsulated state. The fact that the document, and in particular the claims, do not refer to a specific foodstuff indicates that the document relates to a food supplement containing a bioactive material, including a probiotic. Aims of the Invention

[0009] The object of the present invention is to develop a method for encapsulation of postbiotics in a form suitable for ensuring adequate access to the field of use.

[0010] Detailed Description of the Invention

[0011] With the inventive postbiotic encapsulation method, basically, a composition containing at least one postbiotic material is deposited in a texture, shape and size determined according to the field of use with a three-dimensional printing (additive manufacturing) technique. Within the scope of the invention the composition comprising the postbiotic material is deposited using, the selective laser sintering (SLS) technique in which the composition in powder form is fused by melting, the layered bulk modeling (FDM, fused deposition modeling) technique in which the composition in liquid form or melted during application is extruded, the binder jetting (BJ) technique in which a binder is applied to the powdered composition, or in the preferred embodiment of the invention, the ink jetting technique in which the liquid composition is deposited in the form of droplets.

[0012] An embodiment of the invention comprises the steps of mixing at least one postbiotic material and at least one binder in solid form at the temperature of use, depositing the resulting mixture in layers, laser sintering each deposited layer prior to deposition of the next layer.

[0013] Another embodiment of the invention comprises the process steps of mixing at least one postbiotic material and at least one binder in solid form at the use temperature at a temperature above the glass transition temperature or melting temperature of the binder, depositing the resulting mixture in layers, cooling each deposited layer to the use temperature or lower prior to deposition of the next layer.

[0014] The mixture can be deposited directly after preparation, or it can be cooled and stored as granules or filaments and then reheated. A further embodiment of the invention comprises the process steps of depositing at least one postbiotic material in powder form in layers, spraying at least one binder in liquid form onto each deposited layer prior to deposition of the next layer, solidifying the binder sprayed onto each deposited layer prior to deposition of the next layer.

[0015] The binder may consist of a mixture of liquid and solid components. In this case, the liquid components are removed to solidify the binder. This can be achieved by applying heat and / or low pressure to evaporate the liquid components.

[0016] The binder can also be a solid material at the use temperature. In this case, the binder is heated before spraying and cooled to solidify it. The binder can be cooled by spraying a fluid, such as air, on it to remove heat or by waiting.

[0017] A preferred embodiment of the invention comprises the steps of, mixing at least one postbiotic material and at least one binder in liquid form, depositing the resulting mixture in droplets in the form of layers, drying and solidifying each deposited layer prior to the deposition of the next layer.

[0018] The drying of the layers can be achieved by applying heat and / or low pressure to evaporate the liquid components.

[0019] The use temperature refers to the temperature to which the encapsulated postbiotics obtained by a method according to the invention will be exposed during consumption by a user. Said use temperature is selected to be at or above body temperature.

[0020] Postbiotics are composed of metabolites of probiotic organisms. These metabolites may be secreted by probiotic organisms, lysates or mixtures thereof. The beneficial effects of probiotic organisms can also be provided by postbiotics.

[0021] According to the invention, the postbiotic material can be encapsulated by depositing it with specific textures, shapes and sizes. These textures, shapes and sizes are determined by the selected postbiotic material and the site of use. For example, a postbiotic material targeted to act in a certain part of the intestine can be encapsulated with the texture, shape and dimensions determined by considering the conditions it will be exposed to until it reaches there.

[0022] Thanks to the use of three-dimensional printing techniques within the scope of the invention, encapsulated postbiotics whose macro- and micro-structural properties can be easily tailored to the area of use can be produced. By predetermining the form of encapsulation, large-scale production can be realized without the need for skilled personnel input.

[0023] Binders can consist of digestible biopolymers.

[0024] Additives such as preservatives, flavorings and colorants can also be added to the postbiotic material and / or binder.

[0025] The inventive method prevents the loss of probiotics and postbiotics during freeze or spray drying. Furthermore, thanks to the encapsulation suitable for the site of use, postbiotics can be delivered to the site of use without being affected by temperature, pH and enzymes and high utilization is achieved.

[0026] In an exemplary embodiment of the invention, chitosan is used for the encapsulation of the postbiotic material. Accordingly, chitosan nanoparticles are mixed with the postbiotic material as a binder, followed by encapsulation by ionic gelation of chitosan. The postbiotic material can be obtained from lactic acid bacteria.

[0027] In order to demonstrate the effectiveness of the invention, encapsulated postbiotics were produced according to the exemplary embodiment described above and experiments demonstrating postbiotic efficacy were also performed.

[0028] Accordingly, firstly, chitosan nanoparticle solution was obtained by stirring 300 mg of chitosan powder in 2% acetic acid aqueous solution at 2000 rpm for 8 hours at room temperature. On the other hand, inactivation of a Lactiplantibacillus plantarum F2 bacterial culture as a postbiotic source and extraction of postbiotic material was performed. Postbiotic extracts can be obtained by inactivation and extraction by centrifugation of the bacterial culture and removal of the supernatant. Polysorbate 20 at the rate of 0.25 wt% per unit volume was added to the obtained extracts as a surfactant and mixed at 2000 rpm for 3 to 5 minutes. Then chitosan solution was added and mixed within 30 minutes. Then, 0.2 wt% sodium tri-polyphosphate was added to the medium at a rate of 1 mL / h and mixed. Sodium tri-polyphosphate provides the stabilization of chitosan by ionic gelation through the formation of physical cross-links between chitosan nanoparticles. The added sodium triphosphate was mixed at 2000 rpm for 40 minutes. The resulting mixture was fed into a three- dimensional printer to produce encapsulated postbiotic samples. To demonstrate the efficacy of encapsulated postbiotics, the samples were encapsulated in film form. They were then incubated in phosphated saline at 37 °C for up to 72 hours. For this, a well diffusion test was performed using Listeria monocytogenes ATCC 7644 pathogen as indicator. The test results are shown in Table 1.

[0029] Table 1 Efficacy change of film-encapsulated postbiotics according to the invention measured by well diffusion test.

[0030] As shown in Table 1, when the encapsulated postbiotics according to the invention are examined under conditions corresponding to human body temperature and over a period of time selected to cover the digestion process, the release continues and shows activity up to 12 hours. Although the method according to the invention has been described in the exemplary embodiment and in the relevant experiments using chitosan, acetic acid for solubilization of chitosan, polysorbate 20 and sodium tri-polyphosphate, the method according to the field of use can also be applied with other substances that can be safely used in food and medicines for human or other animals. Similarly, other postbiotics can be encapsulated according to the invention instead of lactic acid bacteria extracts.

Claims

CLAIMS1. A method that enables the encapsulation of postbiotics in a form suitable for ensuring adequate access to the field of use; characterized by the deposition of a composition containing at least one postbiotic material with a three-dimensional printing technique in a texture, shape and size determined according to the field of use.

2. A method according to claim 1, characterized in that it comprises the process steps of mixing at least one postbiotic material and at least one binder in liquid form, depositing the resulting mixture in droplets, forming layers, drying and solidifying each deposited layer before depositing the next layer.

3. A method according to claim 2, characterized in that the liquid components are evaporated by applying heat and / or low pressure to dry the layers.

4. A method according to claim 1, characterized in that it comprises the process steps of mixing at least one postbiotic material and at least one binder in solid form at the use temperature, depositing the resulting mixture to form layers, laser sintering each deposited layer prior to deposition of the next layer.

5. A method according to claim 1, characterized in that it comprises the process steps of mixing at least one postbiotic material and at least one binder in solid form at the use temperature at a temperature above the glass transition temperature or melting temperature of the binder, depositing the resulting mixture to form layers, cooling each deposited layer to the use temperature or lower prior to deposition of the next layer.

6. A method according to claim 5, characterized in that the mixture is directly deposited after preparation.

7. A method according to claim 5, characterized in that, the mixture is deposited by first cooling the mixture after preparation into granules or filaments and then reheating.

8. A method according to claim 1, characterized in that it comprises the process steps of - depositing at least one postbiotic material in powder form in layers, spraying at least one binder in liquid form onto each deposited layer prior to deposition of the next layer, solidifying the binder sprayed onto each deposited layer prior to deposition of the next layer.

9. A method according to claim 8 wherein the binder comprises a mixture of liquid and solid components, characterized in that the liquid components are evaporated by applying heat and / or low pressure to solidify the binder.

10. A method according to claim 8 wherein the binder is solid at the use temperature, characterized in that the binder is cooled to solidify.

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