Compositions based on polyhydroxyalkanoate(s) for the production of 3D printing filaments, formulations comprising said compositions, process for preparing said formulations and corresponding consumable filament for 3D printing.

Abstract

The invention relates to formulations and compositions of consumable filaments for 3D printing comprising at least polyhydroxyalkanoate. The invention also relates to a method for preparing such formulations and compositions. The invention further relates to consumable filaments for 3D printing comprising such formulations.

Description

Title of the invention: Compositions based on polyhydroxyalkanoate(s) for the production of 3D printing filaments, formulations comprising said compositions, process for preparing said formulations and corresponding consumable filament for 3D printing. Technical field

[0001] The present invention belongs to the technical field of 3D printing, and more precisely to formulations and compositions usable for 3D printing filaments.

[0002] More particularly, the invention relates to formulations and compositions of consumable filaments for 3D printing comprising at least one polyhydroxyalkanoate. The invention also relates to a method for preparing such formulations and compositions. The invention also relates to consumable filaments for 3D printing made according to these formulations. Prior art

[0003] Polyhydroxyalkanoates (PHAs) are naturally occurring thermoplastic polyesters that can be used as replacements for conventional plastics in various application areas, such as consumer packaging, disposable diaper liners and trash bags, food and medical products, etc. Initial efforts have focused on their use for molding or thermoforming applications, particularly for consumer packaging items such as bottles, cosmetic containers, pens, and golf tees.

[0004] One of the most useful properties of PHAs, which easily distinguishes them from petrochemically derived polymers, is their natural origin and their biodegradability, for example in domestic or industrial compost, in fresh water or in sea water, and more generally in any environment in which a living and active ecosystem is present.

[0005] The biodegradation rate of these PHAs will depend on different factors. The presence of microorganisms, temperature, pH, molecular weight and crystallinity are the main ones.

[0006] PHAs are synthesized naturally by certain microorganisms when the natural environment is unfavorable. They are reserve molecules allowing the microorganism to survive.

[0007] The production of these PHAs by biotechnological means notably uses biofermenters which allow perfect control of the production conditions of this biopolymer.

[0008] The chemical structure of a PHA is a polyester characterized by the length of the side chain in number of carbons. There are therefore different chemical structures of PHA: - PHA salts, which comprise 3 to 5 carbons; - PHA mcl, which comprise 6 to 14 carbons; and - PHAs Here, which include a number greater than 14 carbons.

[0009] There are currently formulations for the manufacture of 3D filaments based on biopolymer compositions containing PHAs. These known biopolymer compositions for 3D filaments mainly contain PHBs (PHA salt having 4 carbons) used in small quantities, in a mixture with other polymers which are not biopolymers (mainly polylactic acid (PLA)) or additives (mainly fillers and fibers). These prior art formulations include, for example, compositions consisting of 20% PHA for 80% PLA.

[0010] Against all expectations, the inventors of the present invention have succeeded in developing 3D filaments composed mainly of PHA biopolymers, which is particularly interesting since these biopolymers are much more compostable and biodegradable than PLA for example. PHAs are in fact biodegradable in soil, fresh water, sea water, domestic compost and industrial compost, while PLA is degradable only in industrial compost. These PHAs can be integrated either alone in formulation in the manufacture of these filaments, or with other polymers distinct from PHAs, the PHAs remaining in majority quantities, and where appropriate with fibers and / or fillers, also in minority proportions. Statement of the invention

[0011] The invention therefore consists, on the one hand, of formulations using PHA compositions as a base, or in other words as a raw material, for the manufacture of filaments for 3D printing. These formulations comprise such PHA compositions as the major ingredient, that is to say that these compositions are present in the formulations of the invention in a content greater than 50% by weight, relative to the total weight of the formulations, up to 100% by weight, relative to the total weight of the formulations, preferably a content greater than 60% by weight, relative to the total weight of said formulation, more preferably a content greater than 70% by weight, relative to the total weight of said formulation. Thus, such formulations comprise at least one PHA as the major ingredient. For the purposes of the invention, it will therefore be understood that a formulation comprises a composition as introduced previously, with or without additives. In a case where no additive is added to the formulation, it will correspond in every respect to the composition it comprises.

[0012] A 3D printing technique that implements these formulations is, for example, 3D printing by fused deposition of material, also called FFF 3D printing (Fused Filament Fabrication), which consists of depositing a molten material, such as plastic, in successive layers via an extruder which moves and constructs a physical object following a design defined by a computer-aided design file.

[0013] The invention also consists, on the other hand, of compositions consisting of PHA, intended to be integrated in a majority manner in the manufacture of formulations for filaments for 3D printing. Such compositions may comprise either a single type of PHA, or a mixture of several types of PHA according to predetermined ratios, or a single type of PHA and at least one other polymer and / or biopolymer and / or other compound, distinct from the PHAs, or a mixture of several types of PHA and at least one other polymer and / or biopolymer and / or other compound, distinct from the PHAs.For the purposes of the invention, it will therefore be understood that a composition comprises in all possible cases PHA, either in the form of a single type of PHA, or in the form of a mixture of at least two types of PHA, and that it may additionally comprise one or more other polymers, whether biosourced or not, or even one or more other compounds, but in any case always distinct from PHAs.

[0014] Thus, a first object according to the invention relates to a composition intended for the design of consumable filaments for 3D printing, which mainly comprises at least one polyhydroxyalkanoate (PHA).

[0015] The invention is therefore based on a completely new and original approach to manufacturing filaments for 3D printing. Such compositions according to the invention comprising PHAs as the majority ingredients, biopolymers of natural origin and biodegradable, are advantageous from an ecological and economic point of view.

[0016] According to one embodiment, the at least one PHA is chosen from the group comprising: PHA salt, PHA mcl, PHA Ici, and their combination.

[0017] The possible use of these different types of PHA makes it possible to modulate the physical, chemical and mechanical parameters, among others, of the compositions with regard to the various additive manufacturing systems in which they can be used, the various printable objects, as well as the application associated with these printed objects.

[0018] According to one embodiment, the at least one PHA is derived from bacteria chosen from the group comprising bacteria of strain CNCM 1-5658, bacteria of strain CNCM 1-5079, and their combination.

[0019] The PHAs integrated into the compositions according to the invention are therefore obtained in a conventional manner by biosynthesis with a microorganism, which does not cause any particular difficulty in their preparation.

[0020] A second subject matter according to the invention relates to a raw material formulation for the design of consumable filaments for 3D printing, which comprises a composition as defined above according to a content greater than 50% by weight, relative to the total weight of the formulation, up to 100% by weight, relative to the total weight of the formulation, preferably a content greater than 60% by weight, relative to the total weight of the formulation, more preferably a content greater than 70% by weight, relative to the total weight of the formulation.

[0021] The compositions previously introduced can be integrated into the production of formulations for 3D printing. Such formulations have, among other things, the same advantages as those of the compositions they comprise.

[0022] According to one embodiment, such a formulation further comprises at least one additional polymer, distinct from a PHA.

[0023] Such an additional polymer also makes it possible to modulate the physical, chemical and mechanical properties, among others, of the formulations according to the invention. The presence of such compounds may, for example, have an impact on the physical or chemical behavior of the PHA(s) present in the compositions according to the invention, themselves present in the formulations according to the invention, in particular during the heating of these PHA(s) during the 3D printing extrusion process: the crystallization temperature of the PHA(s) may, for example, be modified by the presence of such additional polymers.

[0024] According to one embodiment, such a formulation further comprises at least one additive.

[0025] According to one embodiment, this at least one additive is chosen from fibers, fillers and their combination.

[0026] It is therefore advantageously possible to add different types of additives to modulate the formulations and compositions according to the invention, so that they are more suited to 3D filament extrusion.

[0027] According to one embodiment, such a formulation comprises a content of 0.01% to 49% by weight of fibers, relative to the total weight of the formulation, preferably a content of 0.1% to 5% by weight, relative to the total weight of the formulation, more preferably a content of 0.3% to 3% by weight, relative to the total weight of the formulation, and / or a content of 0.01% to 49% by weight of fillers, relative to the total weight of the formulation, preferably a content of 1% to 30% by weight, relative to the total weight of the formulation, more preferably a content of 10% to 20% by weight, relative to the total weight of the formulation.

[0028] These ranges of acceptable values ​​for the integration of additives in the formulations according to the invention make it possible to further adapt the formulations and compositions of the invention according to the 3D printing environment and the properties sought for the printed objects.

[0029] A third object according to the invention relates to a process for preparing a formulation as defined above, comprising the following steps: -culture of PHA-producing bacteria; -concentration of cellular biomass created by the culture of bacteria; -drying of cellular biomass; -extraction of PHA from biomass; and -PHA purification.

[0030] This formulation comprises a content greater than 50% by weight of a composition as defined above, relative to the total weight of the formulation, up to 100% by weight, relative to the total weight of the formulation, preferably a content greater than 60% by weight, relative to the total weight of the formulation, more preferably a content greater than 70% by weight, relative to the total weight of the formulation.

[0031] This process, which makes it possible to prepare a formulation according to the invention, implements conventional steps for obtaining PHA, by the microbiological route, which do not require equipment that is difficult to obtain, and does not present any particular difficulty.

[0032] According to one embodiment, the at least one PHA is chosen from the group comprising: PHA salt, PHA mcl, PHA Ici, and their combination.

[0033] According to one embodiment, the formulation prepared according to this method further comprises at least one additional polymer, distinct from a PHA.

[0034] According to one embodiment, such a formulation further comprises at least one additive, the latter being chosen from fibers and fillers.

[0035] A fourth object according to the invention relates to a consumable filament for 3D printing comprising a composition as defined previously or a formulation as also defined previously. Detailed description of the invention

[0036] Formulations for the manufacture of filaments for 3D printing can thus be prepared, mainly comprising biopolymers, and in particular PHAs. Such formulations can comprise a composition comprising for example PHBHV alone, where PHBHV corresponds to salt PHAs with 4 or 5 carbons, or a composition comprising for example a mixture of PHBHV and PHO, where PHO corresponds to mcl PHAs with 8 carbons. In all cases, such formulations may further comprise one or more additional polymers and / or biopolymers distinct from the PHAs, as well as one or more fillers and / or fibers. Formulations according to the invention

[0037] A formulation for 3D filaments according to the invention can thus consist either of only a composition comprising only one or more PHAs, or of a composition comprising one or more PHAs in majority quantities and further comprising one or more polymers distinct from the PHAs, or even one or more other compounds, always distinct from the PHAs, or of one or other of these two types of compositions in the presence also of one or more fillers and / or fibers, the latter being present in the formulation in minority quantities.

[0038] The formulations in accordance with the invention can thus comprise at least one fiber.

[0039] By "fiber" is meant an object of length L and diameter D such that L is much greater than D, D being the diameter of the circle in which the section of the fiber is inscribed. In particular, the ratio L / D (or form factor) is chosen in the range from 3.5 to 2500, in particular from 5 to 500, and more particularly from 5 to 150. The fibers usable in the formulations of the invention can be fibers of synthetic or natural, mineral or organic origin. They can be short or long, unitary or organized, for example braided, hollow or solid. Their section can be of any shape and in particular of circular or polygonal section (square, hexagonal or octagonal) depending on the specific application envisaged. Their shape can be linear, curved, sinusoidal or curled. In particular, their ends are blunt and / or polished to avoid injury.They can be fruit pomace, for example apple pomace, or even cellulose. The latter has the advantage of resisting high temperatures.

[0040] When present, the fibers may be present in the formulations according to the invention in a content ranging from 0.01% to 49% by weight, relative to the total weight of the formulation, in particular from 0.1% to 5% by weight, and more particularly from 0.3% to 3% by weight.

[0041] The formulations in accordance with the invention may also comprise at least one filler. The fillers may be chosen from those well known to those skilled in the art and commonly used in formulations for the manufacture of 3D filaments. These fillers may be inert or confer activities on the composition organic. These can include oyster shell powder, calcined kaolin, or calcium carbonate.

[0042] When present, the fillers may be present in the formulations according to the invention in a content ranging from 0.01% to 49% by weight, relative to the total weight of the formulation, in particular from 1% to 30% by weight, and more particularly from 10% to 20% by weight.

[0043] A person skilled in the art is able to choose, from among all of these possible additives, both the nature and the quantity of those which will be added to the formulations, so that the latter retain all of their properties. Of course, a person skilled in the art will take care to choose the possible additional additives and / or their quantity in such a way that the advantageous properties of the formulations according to the invention are not, or not substantially, altered by the envisaged addition.

[0044] In particular, the fillers and fibers used in the invention are compounds whose purpose is to improve the processability of the formulations used in the manufacture of filaments for 3D printing. They are integrated into these formulations to act on predetermined characteristics of the filaments that one seeks to obtain, that is to say for example on the hardness of these filaments, their melting temperature, their crystallization temperature or even their glass transition. These fillers and fibers are used in the formulations of the invention for the mechanical effects that they induce on the filaments produced, but also for other possible effects, such as aesthetic effects.In this sense, it is possible to integrate into these formulations fillers which are colorants: such coloring compounds can be used in the invention both for the mechanical properties which they induce in the filaments and for the aesthetic coloring effect which they provide. They can also be used solely for the coloring effect.

[0045] The fillers and fibers used in the invention may also be nucleating agents (fillers), which, when incorporated into the formulations of the invention, allow the filaments produced to harden / set more quickly, because they act on the crystallization temperature of the PHA(s) present in these formulations. This additive therefore allows the filaments produced not to stick and to be able to be wound on themselves to form filament reels.

[0046] The fillers and fibers according to the invention can also confer biological properties to the filaments produced. For example, the integration of copper in the formulations of the invention can confer antibacterial properties to the filaments.

[0047] The formulations according to the invention can also confer different mechanical properties within the same reel of filaments, by integrating different fillers and / or fibers each conferring a distinct mechanical property. (for example, both magnetizing property, antibacterial property and action on the crystallization temperature of the PHA(s) in the formulation composition).

[0048] The integration into the formulations according to the invention of different fillers and / or fibers can also generate a synergistic effect on the final properties of the filaments produced. Indeed, the mixture of distinct fillers and / or fibers within the same formulation can improve each of the effects provided by these fillers and / or fibers compared to these effects present in isolation in separate formulations. Such a mixture of fillers and / or fibers can also provide a formulation with a new effect on the filaments produced, which results from the simultaneous action of these fillers and / or fibers. Compositions according to the invention

[0049] A PHA composition that can be integrated into a formulation for 3D filaments according to the invention can therefore comprise either a single type of PHA (for example PHA salt, such as PHBHV or PHB), or a mixture of at least two types of PHA (for example a PHA scl / PHA mcl mixture, such as PHBHV / PHO, or even two types of PHA salt such as PHBHV / PHB).

[0050] The PHA scl / PHA mcl ratio, for example PHBHV / PHO, in the compositions according to the invention can range from 50 / 50 up to 99.9 / 0.1, preferably from 70 / 30 to 99 / 1, more preferably from 90 / 10 to 95 / 5.

[0051] The PHBHV / PHB ratio in the compositions according to the invention can range from 50 / 50 up to 99.9 / 0.1, preferably from 60 / 40 to 99 / 1, more preferably from 80 / 20 to 95 / 5.

[0052] Naturally, the limits of these intervals are not imperative limits, and the person skilled in the art will be able to adjust the preferred values ​​around 80 / 20, 90 / 10 or 95 / 5.

[0053] For the record, as indicated further in the present application, it is also possible to provide a single type of PHA, for example PHBHV alone, therefore at a content of 100%.

[0054] These PHA compositions according to the invention may also further comprise at least one additional polymer or other additional compound, distinct from the PHAs and present in minor proportions in the compositions. These additional polymers and / or molecules may also be biopolymers, although distinct from the PHAs. They may in particular be chosen from poly(butyl succinate) (PBS), polycaprolactone (PCL), poly(butylene succinate-co-butylene adipate) (PBSA), polytrimethylene terephthalate (PTT), polylactic acid (PLA), starch, polybutylene adipate terephthalate (PBAT), polyurethane (PU), polysaccharides (PS), or even exopolysaccharides (EPS), this list not being exhaustive.

[0055] Such formulations and compositions according to the invention have never been produced until today, because the PHA mainly developed, PHB, is a brittle biopolymer and incompatible alone for this application, hence the preparation and use in the prior art of a formulation containing a low proportion of PHB.

[0056] It is indeed common for high molecular mass polymers to undergo the phenomenon of "cold crystallization", relating, when cooling the latter, to the fact that they are unable to crystallize because their macromolecular chains are too long. They therefore solidify in a glassy state. When they are reheated, the chains of these polymers acquire enough energy for them to start moving, thus allowing them to acquire a position allowing crystallization. This is then an imperfect crystallization, which results in reduced mechanical properties and risks of deformation of the printed part, due to post-crystallization which occurs over time.

[0057] Therefore, according to the invention, when PHB alone is used in the PHA compositions, such compositions may also comprise at least one other type of PHA which is not PHB, and / or additional polymers, such as biopolymers distinct from PHAs. Also, a solution to counter this brittle effect of 3D filaments, which does not allow filament reels to be produced, due to the presence of PHB in large quantities in the compositions of the invention, the final formulations which comprise such compositions may also comprise fillers and / or fibres.

[0058] Indeed, in the manufacture of 3D filaments, the PHAs of the compositions according to the invention, which are amorphous compounds, are melted at a certain temperature to form a resin, then cooled to allow their crystallization. For example, the crystallization temperature of the PHA salt PHBHV is around 60°C. If it is used alone in the compositions, its crystallization will only be complete when the filament produced has reached this temperature of around 60°C, which means that the filament produced can shrink a long time after its production. Therefore, when PHBHV is used as the sole PHA in the compositions according to the invention, it is advantageous, or even necessary, to add another polymer distinct from the PHAs, and / or at least one filler or fiber to the final formulation, such as a nucleating agent (filler), which makes it possible to make the crystallization temperature of the PHBHV negative, and therefore its crystallization immediate.The produced filament will not retract once its production is finalized at the printer output.

[0059] This type of parameter (crystallization temperature, melting temperature, glass transition, etc.) can be measured, for example, using the differential scanning calorimetry technique. scanning (Differential Scanning Calorimetry, DSC), which measures the temperatures and heat flows associated with the thermal transitions of a material. Obtaining polyhydroxyalkanoate biopolymers

[0060] The present invention therefore does not expressly cover the strains capable of producing the biopolymers, but the use of the polyhydroxyalkanoates secreted by them for the production of formulations and compositions for the manufacture of threads for 3D printing.

[0061] Obtaining PHA salt is obtained in a known manner by biosynthesis with a microorganism of the genus halomonas by a fermentation process, and in particular the microorganism registered in the National Collection of Culture of Microorganisms under the Budapest Treaty under number CNCM 1-5079, deposit made in the name of the company Polymaris Biotechnology, located in Brest - France.

[0062] Obtaining PHA mcl is also obtained in a known manner by biosynthesis with a microorganism of the genus pseudomonas by a fermentation process, and in particular, the microorganism registered in the National Collection of Culture of Microorganisms under the Budapest Treaty under number CNCM 1-5658 deposit made in the name of the company Polymaris Biotechnology, located in Brest - France.

[0063] The fermentation to produce the PHA is carried out under conditions of agitation and aeration of the medium at a temperature between 20°C and 37°C, preferably at 30°C, the medium having a pH between 6.5 and 9, preferably close to 7.5, and being adjusted if necessary during the fermentation. The duration of the fermentation is between 24 and 120 hours, preferably between 36 and 72 hours. According to one embodiment of the invention, mineral salts are provided during the fermentation of the bacteria. By way of illustration and not limitation, mention may be made, among the mineral salts, of those capable of providing the culture medium with Na+, K+, NH4+, Ca2+, Mg2+, PO43-, SO42-, Cl-, CO3- ions or essential elements such as Cu, Mn, Fe and Zn.

[0064] The methods of isolation and purification of PHA are obtained by conventional means well known to those skilled in the art, such as centrifugation, filtration, ultrafiltration, extraction, evaporation and dialysis. Structure of PHA salt

[0065] The PHA salt which can be integrated into the compositions according to the invention consists of a repetition of polymeric units of formula A1 and polymeric units of formula B1, such as: Formula Al:-[-O-CH(Rl)-(CH2)n-CO-]- Formula Bl:-[-O-CH(R2)-(CH2)m-CO-]-in which: RI and R2, different from each other, represent a group chosen from linear or branched alkyl (C0-C2), and linear or branched alkenyl (C0-C2), said alkyl or alkenyl group being optionally substituted by 1 to 4 groups, identical or different, chosen from halogen (F, Cl, Br, I), -CN, -OH, -OR, -NH2, -NHR, C02H, -CO2R in which R represents a linear or branched alkyl group (C1-C6); n and m, identical or different, represent an integer chosen from 1 or 2.

[0066] In the case of P3HBHV: Rl= Cl, R2 =C2, with n=m=l. Structure of the PHA mcl

[0067] The PHA mcl which can be integrated into the compositions according to the invention consists of a repetition of polymeric units of formula Al', of polymeric units of formula Bl' and of polymeric units of formula Cl such that: Formula Al':-[-O-CH(Rl)-(CH2)n-CO-]- Formula Bl':-[-O-CH(R2)-(CH2)m-CO-]- Formula Cl:-[-O-CH(R3)-(CH2)p-CO-]- in which: RI, R2 and R3, different from each other, represent a group chosen from linear or branched (C3-C11) alkyl, and linear or branched (C3-C11) alkenyl, said alkyl or alkenyl group being optionally substituted by 1 to 4 groups, identical or different, chosen from halogen (F, Cl, Br, I), -CN, -OH, -OR, -NH2, -NHR, CO2H, -CO2R in which R represents a linear or branched (C1-C6) alkyl group; n, m and p, identical or different, represent an integer chosen from 1 or 2.

[0068] In the case of PHO: RI = C3, R2 = C5, with n=m=l, Cl not existing.

[0069] Preparation and isolation of polyhydroxyalkanoates

[0070] Method for culturing the bacteria registered under deposit number CNCM 1-5658

[0071] For the production of the PHA biopolymer, all operations are carried out according to good microbiology practices under sterile conditions. The different media are also sterilized either by heat treatment (121°C for 20 minutes) or by 0.2 pm filtration.

[0072] A 2 mL cryotube of the CNCM 1-5658 strain is inoculated into an Erlenmeyer flask containing 50 mL of medium for the culture of marine microorganisms (marine broth medium E2216, Difco, France). The culture is incubated at 30°C with shaking for 12 to 18 hours.

[0073] The culture obtained is then inoculated into an Erlenmeyer flask containing 1.45 L of mineral medium suitable for its growth (Ml medium) at 30°C with stirring for 8 hours.

[0074] Finally, the culture is inoculated into a fermenter containing 13.5 L of mineral culture medium adapted to its growth and the biosynthesis of the biopolymer (M2 medium) containing dextrose (10 g / L). The culture is maintained at 30°C with shaking for at least 30 hours.

[0075] During the first 28 hours of culture, a solution of octanoic acid buffered to pH 6 with NaOH ION is regularly added.

[0076] The pH is regulated at 7.2 by 10 N NaOH or 2 N H2SO4.

[0077] Oxygenation and agitation of the culture are also regulated to allow cell growth and accumulation of the biopolymer.

[0078] At the end of fermentation, the cellular biomass containing the biopolymers is concentrated by centrifugation.

[0079] This biomass is then dried in order to eliminate all the water and until a constant mass is obtained.

[0080] The composition of the medium Ml is detailed in Table 1 below.

[0081] [Tables 1] Elements g / kg in H2O g / kg in H2O NaNH4HPO44H2O 3.5 KH2PO4 3.7 k2hpo4 7.5 MgSO4 0.25 NaCl 20 Solution No. IA 40 Sodium Citrate 250 Yeast Extract 25 Solution No. 2 1.3 FeSO4.7H2O 2.78 CaCl2.2H2O 1.47 MnCl2.4H2O 1.98 CoC12.6H2O 2.38 ZnSO4.7H2O 0.29 CuC12.2H2O 0.17

[0082] The composition of the M2 medium is detailed in Table 2 below.

[0083] [Tables2] Elements g / kg in H2O g / kg in H2O NaNH4HPO44H2O 3.5 KH2PO4 3.7 k2hpo4 7.5 MgSO4 0.25 NaCl 20 Solution No. IA 40 Sodium citrate 250 Yeast extract 25 Solution No. 2 1.3 FeSO4.7H2O 2.78 CaCl2.2H2O 1.47 MnCl2.4H2O 1.98 CoC12.6H2O 2.38 ZnSO4.7H2O 0.29 CuC12.2H2O 0.17

[0084] Method of culturing the bacteria registered under deposit number CNCM 1-5079

[0085] For the production of the biopolymer, all operations are carried out according to good microbiology practices under sterile conditions. The different media are also sterilized either by heat treatment (121°C for 20 minutes) or by 0.2 pm filtration.

[0086] The M3 medium is composed of 0.5 g / L of NH4C1, 0.5 g / L of MgSO-7H2O, 0.1 g / L of CaC12-2 H2O, 0.04 g / L of KH2PO4, 0.12 g / L of K2HPO4, 20 g / L of NaCl and 6 g / L of HEPES. A trace element solution was also prepared. It consists of 7.3 g / L of Na2-EDTA, 2.5 g / L of FeSO4-7H2O, 0.02 g / L of MnC12-4H2O, 0.242 g / L of Na2MoO4-2H2O, 0.038 g / L of CuC12-2H2O and is added after autoclaving to the culture medium.

[0087] A 2 mL cryotube of the CNCM 1-5079 strain is inoculated into an Erlenmeyer flask containing 50 mL of medium for the culture of marine microorganisms (marine broth medium E2216, Difco, France). The culture is incubated at 30°C with shaking for 12 to 18 hours.

[0088] The culture obtained is then inoculated into an Erlenmeyer flask containing 1.45 L of mineral medium suitable for its growth (M3 medium) at 30°C with stirring for 8 hours.

[0089] Finally, the culture is inoculated into a fermenter containing 13.5 L of mineral culture medium adapted to its growth and the biosynthesis of the biopolymer (M2 medium + glycerol). The culture is maintained at 30°C with shaking for at least 30 hours.

[0090] During cultivation, glycerol and valeric acid are added to allow the accumulation of the desired PHA salt.

[0091] The pH is regulated at 7.2 by 10 N NaOH or 2 N H2SO4.

[0092] Oxygenation and agitation of the culture are also regulated to allow cell growth and accumulation of the biopolymer.

[0093] At the end of fermentation, the cellular biomass containing the biopolymers is concentrated by centrifugation.

[0094] This biomass is then dried in order to eliminate all the water and until a constant mass is obtained.

[0095] Purification of polyhydroxyalkanoate salt and mcl biopolymers

[0096] The extraction of the PHA biopolymer is carried out by adding dichloromethane at a rate of 10% of dry biomass per volume of solvent at room temperature and with stirring for 18 hours.

[0097] The biomass is removed by centrifugation (8000 rpm X 20 min) and the supernatant containing the biopolymer is recovered.

[0098] The latter can be filtered in order to improve the quality of the final biopolymer.

[0099] It is concentrated by removing dichloromethane using a rotary evaporator.

[0100] The concentrated biopolymer in solution in dichloromethane is finally precipitated by adding a cold ethanol solution (10:1) then dried in a ventilated oven at 40°C until complete evaporation of the solvent.

[0101] Physicochemical characterization of the obtained polyhydroxyalkanoate biopolymer

[0102] The molecular weight of the PHA obtained is determined by size exclusion chromatography with a refractometer (RI) for detection. It is determined by comparing the retention time of the polymer with the retention time of polystyrene standards via a calibration line.

[0103] The analysis makes it possible to determine the average molecular weight (Mw in Dalton), the number average molecular weight (Mn in Dalton) in PS (polystyrene) equivalent, as well as the polydispersity index.

[0104] The composition of the biopolymer is determined by gas chromatography equipped with a flame ionization detector (FID) and confirmed by 1H, 13C, HMBC and HMQC NMR analysis.

[0105] A copolymer consisting of 96% by weight of poly(hydroxy-3-octanoate) and 4% by weight of poly(hydroxy-3-hexanoate) for the PHA mcl was thus obtained, and a copolymer of 80% of poly-3-hydroxybutyrate and 20% of poly-3-hydroxyvalerate for the PHA salt. Examples

[0106] Filament extrusion tests with formulations comprising a PHA composition and a filler, which is a nucleating agent

[0107] On the one hand, the formation of a coil of filaments by filament extrusion was tested using formulations comprising mainly a composition consisting of PHA, and in particular PHA salt, in a mixture with a nucleating agent. Such a nucleating agent, by virtue of its nucleation effects, participates in the control, or even the improvement, of the structural and mechanical properties of the PHA filaments.

[0108] Three different formulations were prepared, according to the procedure described in the detailed description above, intended to be tested on a laboratory extrusion line for 3D printing. In particular, an attempt was made to form filament reels to be integrated into 3D printing machines whose extruded rods had a diameter of 1.75 mm for each of these three compositions.

[0109] The first formulation tested (Fl) comprises 97% of a PHA salt composition, which is PHBHV, as well as 3% of cellulose fiber of 30 μm particles. The PHBHV is composed of 80% by weight of poly-3-hydroxyburyrate and 20% by weight of poly-3-hydroxyvalerate.

[0110] The second formulation tested (F2) comprises 97% of a PHBHV composition, as well as 3% of a nucleating agent which is boron nitride.

[0111] The third formulation tested (F3) comprises 97% of a PHBHV composition, as well as 3% of boron nitride and 3% of a colorant (here, MMbleu). Test conditions

[0112] A small diameter extruder (15 mm - 20D) was used, consisting of five heating zones. The die outlet cavity was 3 mm, and cylinder pulling was carried out in order to achieve a rod with a diameter of 1.75 mm.

[0113] The extruder was equipped with a gear pump and gear head for a regular flow rate. It was mounted on a lifting device to regulate the cooling of the extrudate and a thermo-regulated, hot or cold, closed-loop overflow water cooling tank.

[0114] The line was equipped with a three-position constant torque winder, in order to provide low winding tensions.

[0115] A position to have little tension was used for the test, according to the following parameters: spooling 40 rpm, station furthest from the tank 50 rpm.

[0116] The steaming was done in a turbo oven at 50°C overnight (minimum 8 hours; maximum 12 hours).

[0117] During filament extrusion, the material cooling tank was at a temperature of 8°C, and was increased to 50°C.

[0118]

[0119] Results The results of the extrusion tests on the three formulations are grouped in the Table 3 below. [Tables3] Formulations Temperature of the extruder heating zones (°C) Pressure (bars) and screw rotation speed (rpm) Drawing speed (m. min ') Cooling tank temperature (°C) Results Fl: test 1 From 150 to 175 / / 8 -2mm filament -Sticky material Fl: test i2 From 150 to 175 20 43 / 50 -1.7mm filament -Material less sticky than test 1 Fl: test i3 From 150 to 175 20 38 4.3 50 -1.75mm filament -Obtaining a filament spool -But material that remains sticky F2 From 150 to 175 19 38 4.3 50 -1.75mm filament -Obtaining a reel of filament -Material less sticky than Fl test 3 F3 From 150 to 175 11 38 4.3 50 -1.75 mm filament -Obtaining a reel of filament

[0120] It is clear from the results of this table that it was necessary to conduct several tests under different conditions (screw rotation speed, pressure, cooling tank temperature, etc.) to successfully obtain a filament spool from formulation F1. However, the filament material of this spool remains sticky, and it is necessary to run water over the extruder rollers in order to limit the adhesion of the filament to these rollers. This spool thus obtained is therefore difficult to unwind. It has therefore been determined that formulation F1 is not perfectly suited for filament extrusion.

[0121] The experimental conditions of test number 3 of Fl could be reused to test formulation F2. It was also possible to obtain a filament reel whose material is less sticky than that of the filament of test 3 of Fl. It is therefore no longer necessary to run water over the extruder rollers. It was therefore determined that this formulation F2 behaves well in filament extrusion.

[0122] The same experimental conditions were again applied for the extrusion test of formulation F3. A reel could also be obtained. The filament of the reel is slightly whitened in places (adhesion to the rollers, inhomogeneous dispersion of the dye), and the circulation of water on the rollers was reinstated compared to the test of formulation F2. Despite everything, it was determined that formulation F3 is suitable for filament extrusion.

[0123] Filament extrusion tests with formulations comprising only PHA compositions, as well as with formulations comprising PHA compositions in the presence of fillers

[0124] The formation of filament coils by filament extrusion was also tested using formulations still comprising mainly PHA compositions, and in particular compositions consisting of a mixture of PHA salt and PHA mcl, as well as using such compositions mixed with fillers.

[0125] Initially, different compositions were prepared in batch mode in order to determine the optimal parameters for obtaining filaments suitable for 3D printing.

[0126] The conditions of such filament extrusion tests are detailed in Table 4 below.

[0127] [Tables4] Compositions Extruder temperature (°C) Screw rotation speed (rpm) PHBHV (mass%) PHO (mass%) Test 1 168 50 95 5 Test 2 168 50 90 10 Test 3 168 50 85 15 Test 1' 178 50 90 10 Test 2' 178 50 85 15 Test 3' 178 50 70 30

[0128] Based on the results of these compositions tested in filament extrusion, which results also took into account the crystallization kinetics of the extruded product, four different formulations were prepared, always according to the operating method described in the detailed description above, intended to be tested on a laboratory extrusion line for 3D printing.

[0129] The first formulation tested (Fl') comprises 100% of a composition consisting of 95% PHBHV and 5% PHA mcl, which is PHO. The PHO is composed of 96% by weight of poly-3-hydroxyoctanoate and 4% by weight of poly-3-hydroxyhexanoate.

[0130] The second formulation tested (F2') comprises 80% of a composition consisting of 76% PHBHV and 4% PHO, as well as 20% oyster shell powder (ostrecal 15).

[0131] The third formulation tested (F3') comprises 80% of a composition consisting of 76% PHBHV and 4% PHO, as well as 20% calcined kaolin (opacilite).

[0132] The fourth formulation tested (F4) comprises 80% of a composition consisting of 76% PHBHV and 4% PHO, as well as 20% calcium carbonate.

[0133] The results of the extrusion tests on these four formulations, on a 3DEvo Advanced 1.0 single-screw spinning extrusion line, are grouped in Table 5 below.

[0134] [Tables5] Formulations Temperature of the extruder heating zones (°C) Screw rotation speed (rpm) Results Fl' From 155 to 175 200 Filaments suitable for 3D printing obtained F2 ' From 155 to 175 250 Filaments suitable for 3D printing obtained F3' From 155 to 175 250 Filaments suitable for 3D printing obtained F4 From 155 to 175 250 Filaments suitable for 3D printing obtained

[0135] These extrusion tests show that the four formulations Fl' to F4 behave well in filament extrusion. They were then able to be integrated into 3D printing tests, in a p-Delta printer, EmotionTech, Titan Extruder 1.75 mm, Hotends E3D V6, Nozzle 0.5 and 0.8 mm, and allowed the manufacture of 3D objects, such as pots, containers, or even more complex objects, such as boat figurines.

Claims

Claims

1. Composition intended for the design of consumable filaments for 3D printing, characterized in that it consists of at least two different types of polyhydroxyalkanoate (PHA).

2. Composition according to claim 1, characterized in that said at least two different types of PHA are chosen from the group comprising PHA salt, PHA mcl, and their combination.

3. Composition according to claim 2, characterized in that it consists of a mixture of two types of PHA salt and PHA mcl according to a PHA scl / PHA mcl ratio which can range from 50 / 50 up to 99.9 / 0.1, preferably from 70 / 30 to 99 / 1, more preferably from 90 / 10 to 95 / 5.

4. Composition according to any one of claims 1 to 3, characterized in that said at least two different types of PHA are derived from bacteria chosen from the group comprising bacteria of strain CNCM 1-5658, bacteria of strain CNCM 1-5079, and their combination.

5. Raw material formulation for the design of consumable filaments for 3D printing, characterized in that it comprises a composition according to claims 1 to 4 in a content greater than 50% by weight, relative to the total weight of said formulation, up to 100% by weight, relative to the total weight of said formulation, preferably a content greater than 60% by weight, relative to the total weight of said formulation, more preferably a content greater than 70% by weight, relative to the total weight of said formulation.

6. Formulation according to claim 5, characterized in that it further comprises at least one additional polymer, distinct from a PHA.

7. Formulation according to claim 5 or 6, characterized in that it further comprises at least one additive.

8. Formulation according to claim 7, characterized in that said at least one additive is chosen from fibers, fillers and their combination.

9. Formulation according to claim 7 or 8, characterized in that it comprises a content of 0.01% to 49% by weight of said fibers, relative to the total weight of the formulation, preferably a content of 0.1% to 5% by weight, relative to the total weight of said formulation, more preferably a content of 0.3% to 3% by weight, relative to the total weight of said formulation, and / or a content of 0.01% to 49% by weight of said fillers, relative to the total weight of the formulation, preferably a content of 1% to 30% by weight, relative to the total weight of said formulation, more preferably a content of 10% to 20% by weight, relative to the total weight of said formulation.

10. A method for preparing a formulation according to claims 5 to 9 comprising the following steps: -cultivation of at least two bacteria respectively producing two different types of PHA; -concentration of at least two cellular biomasses respectively created by said bacterial cultures; -drying of said at least two cellular biomasses; -extraction of said at least two different types of PHA respectively from said at least two biomasses; -purification of said at least two different types of PHA;characterized in that said formulation comprises a content greater than 50% by weight of a composition according to claims 1 to 4, relative to the total weight of the formulation, up to 100% by weight, relative to the total weight of the formulation, preferably a content greater than 60% by weight, relative to the total weight of said formulation, more preferably a content greater than 70% by weight, relative to the total weight of said formulation.;

11. A method according to claim 10, characterized in that said at least two different types of PHA are selected from the group comprising PHA salt, PHA mcl, and combinations thereof.

12. Method according to claim 10 or 11, characterized in that said formulation further comprises at least one additional polymer, distinct from a PHA.

13. Method according to any one of claims 10 to 12, characterized in that said formulation further comprises at least one additive, said additive being chosen from fibers and fillers.

14. A consumable filament for 3D printing comprising a composition according to claims 1 to 4 or a formulation according to claims 5 to 9.