Method for marking material for authentication and / or traceability of said material

Abstract

The invention relates to a method for marking materials, for example for the purpose of authentication and / or traceability of said materials, in particular batch-produced materials. The labeling method comprises:-a step of supplying a labeling composition selected from molecular tags, and-a step of incorporating the labeling composition into the material. And, according to the invention, said marking composition comprises particles which, after incorporation into said material, advantageously remain intact, comprising:-a homogeneous porous matrix comprising a plurality of pores; -an information carrier added to the homogeneous porous matrix wherein the information carrier consists of a polymer comprising an information sequence; and-a particle surface having a coating comprising at least one fluidizing agent, suitable for promoting the homogeneous dispersion of said particles within said material.

Description

Technical Field

[0001] The present invention relates to the technical field of labelling materials using molecular tags, for example for the purpose of authentication and / or traceability of said materials. Background Art

[0002] Marking methods are positioned in the context of two main applications to give a material or finished product a competitive advantage, namely:

[0003] - authentication, in particular to prevent forgery, which is a cause of commercial losses, and

[0004] - Traceability, as brands and retailers are increasingly under pressure from consumers to create transparency across their entire supply and production chains.

[0005] Typical marking methods are based on traditional barcodes, QR codes (Quick Response Codes) or even RFID (Radio Frequency Identification) chips, which are authentication and batch tracking systems positioned on the packaging or in the product surface and have fast reading systems.

[0006] However, these “surface” tags cannot be applied to certain materials or in situations where the code needs to be invisible to the naked eye.

[0007] Molecular tags address these shortcomings through their nanoscale footprint and difficulty in counterfeiting.

[0008] In fact, "molecular tags" are constituent elements of the molecular composition of the material into which they are incorporated. They are an integral part of the mass of material of interest.

[0009] A molecular tag is therefore a true molecular marker, making it possible to trace a batch of material and follow its evolution over time.

[0010] This authentication technology also provides added value to materials by collecting data from tagged materials and tracking them as they are incorporated into finished products.

[0011] Molecular tagging therefore offers innovative solutions for physically marking, tracking and authenticating materials, from the producer of the products made from them to the retailer, creating transparency throughout the entire supply chain (linear or circular).

[0012] However, current molecular tags are not entirely satisfactory.

[0013] For example, WO2013143014 discloses encapsulating informative DNA within non-porous silica nanoparticles, suitable for invisible security marking or information as part of a product. Specifically, the DNA is packaged between a silica core and a silica shell.

[0014] However, silica nanoparticles have non-negligible solubility in water at alkaline pH values ​​(above pH 9), so the silica shell may be unstable in alkaline materials and the dispersibility of colloidal silica nanoparticles is affected.

[0015] In particular, DNA can also be protected by the nanoparticle's outer layer consisting of covalently linked silicones.

[0016] However, to release the DNA, the siloxane covalent bonds must be broken using hydrofluoric acid, a relatively dangerous acid to handle.

[0017] Therefore, there is a need for a molecular tag that can be well dispersed (preferably uniformly dispersed) in the body of a target material without changing its properties.

[0018] Furthermore, preferably, the information support (information carrier) needs to be protected in a material and then be recovered (recovered, extracted, restored) together with the associated information sequence.

[0019] The process of extracting information from products refined with the target material must be simple and secure enough to be transposed to an analytical laboratory. Summary of the Invention

[0020] In order to remedy the above-mentioned disadvantages of the prior art, the present invention proposes a method for marking materials, for example for the purpose of authentication and / or traceability of said materials, in particular batch-produced materials.

[0021] The marking method includes:

[0022] - a step of supplying a labeling composition selected from molecular tags, and

[0023] - A step of incorporating said marking composition into said material.

[0024] Furthermore, according to the present invention, the marking composition comprises particles which advantageously remain intact after incorporation into the material, the particles comprising:

[0025] - a homogeneous porous matrix containing a plurality of pores,

[0026] - an information carrier added to said homogeneous porous matrix, and

[0027] - The surface of the particles has a coating comprising at least one fluidizing agent, suitable for promoting a homogeneous dispersion of said particles within said material.

[0028] Preferably, the combination of information carrier, homogeneous porous matrix and fluidizing agent is selected so as to achieve self-association. It therefore does not require chemically reactive substances to form covalent bonds, thereby promoting their dissociation.

[0029] This labeling method is particularly interesting because the particles of the labeling composition disperse particularly well in the bulk of the target material without altering its properties. This optimized dispersion is particularly interesting for increasing the sensitivity when recovering molecular tags from the material. Furthermore, the fluidizing agent advantageously improves the stability of the information carrier and the homogeneous porous matrix.

[0030] Furthermore, the information carrier is advantageously well protected in the material and can then be recovered together with the associated information sequence.

[0031] Preferably, the information carrier is stabilized by association with a porous matrix and a fluidizing agent, which results in its protection within the material and subsequent recovery together with the associated information sequence.

[0032] The use of a homogeneous porous matrix has a clear advantage because it can load more information carriers for particles of comparable composition and size.

[0033] The information carrier is added to a porous material and surface modified with a fluidizing agent to ensure the stability of the encoded data and its applicability to a range of materials.

[0034] Finally, preferably in another advantageous aspect, the particles are stable in the material, but if exposed to the environment, they are degradable. Furthermore, the decomposition products advantageously produce molecules that are environmentally friendly and safe for human health.

[0035] Further non-limiting and advantageous features of the process according to the invention (considered individually or in all technically possible combinations) are as follows:

[0036] a homogeneous porous matrix consisting of at least 90% w / w of a salt, preferably an inorganic salt, consisting of an ionic assembly of positively charged cations and negatively charged anions or of a metal oxide, wherein the homogeneous porous matrix is ​​insoluble in the material;

[0037] The particles are decomposable by metal chelating agents or by specific pH conditions (e.g., acidic or alkaline solutions), preferably mildly acidic conditions, even more preferably pH 2 to 5, even more preferably aqueous solutions of citric acid, acetic acid, or hydrochloric acid;

[0038] - the particles have a size in the range of 200 nm to 100 μm, preferably 500 nm to 10 μm, more preferably 750 nm to 3 μm;

[0039] - the particles have a pore size in the range of 2 nm to 500 nm, preferably 2 to 200 nm, more preferably in the range of 10 nm to 100 nm;

[0040] - the information carrier is contained in the pores of the homogeneous porous matrix and / or adsorbed on the surface of the homogeneous porous matrix;

[0041] - the information sequence of the information carrier is obtained by transcoding a system of ordered bits, the ordered bit system being obtained by encoding a set of original data;

[0042] the information carrier is selected from nucleic acids and nucleic acid analogs, preferably DNA and more preferably single-stranded DNA or double-stranded DNA; the information carrier advantageously consists of a nucleic acid fragment, said nucleic acid fragment preferably having a size of less than 500 bp, preferably in the range of 50 to 250 bp;

[0043] - the information carrier is associated with said homogeneous porous matrix, preferably adsorbed and / or contained by said homogeneous porous matrix;

[0044] - the fluidizing agent is selected from the group consisting of surface treatment agents for inorganic fillers (see KATZ, Harry S. et MILESKI, JV (ed.). Handbook of fillers for plastics. Springer Science & Business Media, 1987; for a review of inorganic fillers such as precipitated calcium carbonate, barite, kaolin, talc, silica and carbon black), which include saturated fatty acids; unsaturated fatty acids; phospholipids; lipid mixtures; long alkyl chain surfactants; amphiphilic polymers; resin acids; waxes; hydrophilic polymers; peptides and proteins; polysaccharides or other inorganic salts;

[0045] the fluidizing agent is selected from fluidizing agents that improve dispersion in hydrophobic materials, preferably amphiphilic molecules or surfactants, such as hydrophilic saturated fatty acids, advantageously having a carbon chain length of C15 to C20, such as palmitic acid and stearic acid; unsaturated fatty acids, advantageously having a carbon chain length of C15 to C20, such as oleic acid and linoleic acid; phospholipids, such as phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine; lipid mixtures, preferably of biological origin or biodegradable, such as lecithin; organic amphiphilic molecules with a lipophilic part, such as phosphonates, glycols, alcohols, phenates, sulfonsates, salicylates, succinic anhydride; waxes, such as beeswax; resin acids, such as alicyclic carboxylic acids and rosin acids and their salts, esters and ethers; bifunctional polymers with a hydrophobic part, such as or some filler coupling agents, such as titanate (LICA ) or zirconate

[0046] - fluidizing agents selected from fluidizing agents that improve dispersion in hydrophilic materials, for example hydrophilic polymers, such as polycarboxylic acids, fused phosphoric acid, polyacrylates, polyamines, polyacrylamides, poly-aminoacyls, polystyrene, polyethylene glycol (PEG), polyvinylpyrrolidone (PVP), polymaleic acid (PMA), polyepoxysuccinic acid (PESA), polysaccharides (dextran, alginate, chitosan, chondroitin, cellulose, pectin, carboxymethyl inulin, etc.), peptides (such as polyaspartic acid, polyglutamic acid), proteins, lignin, preferably biogenic or biodegradable polymers; other inorganic salts (such as calcium phosphate, hydroxyapatite);

[0047] - the material is selected from liquid, semi-solid or solid materials, such as plastics, thermosetting materials, varnishes, rubbers, coatings, oils, lubricants, pharmaceuticals, cosmetics, foods;

[0048] - The incorporation step consists of incorporating the marking composition into said material during its manufacture.

[0049] The present invention also relates to a marking composition for marking a material, for example for the purpose of authentication and / or traceability of said material, in particular of batch-produced material, wherein said marking composition comprises particles which advantageously remain intact after incorporation into said material, said particles comprising:

[0050] - a homogeneous porous matrix containing a plurality of pores,

[0051] - an information carrier added to said homogeneous porous matrix, wherein said information carrier consists of a polymer comprising an information sequence, and

[0052] - The surface of the particles has a coating comprising at least one fluidizing agent, suitable for promoting a homogeneous dispersion of said particles within said material.

[0053] The present invention also relates to materials comprising the marking composition of the present invention.

[0054] The material can be:

[0055] - hydrophobic materials (e.g. plastics, oils, waxes, varnishes, lubricants) or

[0056] - Hydrophilic materials (such as ink, paint, water-based thermosetting materials, milk, cement).

[0057] The different features, variants and embodiments of the invention can be combined with one another in various combinations without being mutually incompatible or exclusive. BRIEF DESCRIPTION OF THE DRAWINGS

[0058] Figure 1 Provided are optical and fluorescence microscopy (x100) images of molecular tags obtained using a Nikon H600L fluorescence microscope: CaCO3 porous microparticles (vehicle), loaded with 10 mg / g of double-stranded 70 bp DNA labeled with green fluorescent Atto488 (information carrier), and 0.3% w / w sodium polyacrylate (fluidizer). A / Optical micrograph (differential interference contrast) of a 6 μm molecular tag. B / Green fluorescence micrograph of the same 6 μm molecular tag. The green fluorescence is due to the Atto488-labeled DNA. C / Optical micrograph (differential interference contrast) of a 2.5 μm molecular tag. D / Green fluorescence micrograph of the same 2.5 μm molecular tag.

[0059] Figure 2 Demonstration of changes in the physicochemical properties of molecular tags in the presence or absence of a fluidizing agent is provided. Particle size (size in μm) and polydispersity measurements in water (volume frequency in %) were obtained using a Malvern Mastersizer 3000 equipped with a Hydro2000S modular system.

[0060] A: Single 2.5 μm homogeneous porous calcium carbonate support;

[0061] B: 2.5 μm hydrophilic molecular tags fully dispersed in water (fluidizer: sodium polyacrylate 0.5% w / w).

[0062] Figure 3 The protective effect of the carrier+fluidizing agent combination on the chemical reactivity of DNA towards reactive genotoxic molecules such as isothiocyanates (FITC: fluorescein isothiocyanate or RITC: rhodamine isothiocyanate) is shown.

[0063] Left: Graph of relative DNA fluorescein fluorescence after 48 h of reaction with FITC (fluorescence in arbitrary units).

[0064] 1: Water; 2: 80bp DNA without FITC (negative control); 3: 80bp DNA with FITC (positive control); 4: 80bp DNA loaded into a carrier at 10 mg / g, with FITC added; 5: 80bp DNA loaded into a carrier at 3.5 mg / g, conjugated with the fluidizing agent sodium polyacrylate (0.5% w / w), with FITC added.

[0065] Right: 4% low melting point agarose electrophoresis gel of the same DNA fragments.

[0066] A / shows fluorescein fluorescence; B / shows DNA fragments with BET staining.

[0067] Only when loaded into a carrier associated with a fluidizing agent can DNA be protected from the effects of isothiocyanates. The CaCO3 carrier alone is not sufficient to protect DNA.

[0068] Figure 4 A 4% agarose plate is provided after electrophoresis of DNA fragments generated by PCR from DNA extracted from thermoplastic material. The thermoplastic material is labeled with a hydrophobic molecular tag (homogeneous porous CaCO3 carrier, loaded with 70 bp DNA at 10 mg / g and covered with 2% w / w stearic acid).

[0069] The 70 bp DNA information carrier extracted from the solidified and molded polymer was amplified with 2 x 35 bp extension primers (2 x [20 bp primer for DNA amplification + 15 bp for incorporation into plasmid]) to amplify a 100 bp DNA oligomer.

[0070] 1: Negative control (untreated water); 2: Negative control (untreated digestion buffer); 3: Negative control (extracted polymer PDLG 7507); 4: negative control (extracted polymer PDLG5010); 5: extracted polymer

[0071] PDLG 7507, labeled with 2ppm molecular tag; 6: extracted polymer PDLG 5010, labeled with 1 ppm molecular tag. 7: positive control (PCR of 1 ng DNA), L: untreated reference TriDye TM Ultra-low range DNA ladder from NEB. DETAILED DESCRIPTION

[0072] In addition, various other features of the present invention will become apparent from the following description.

[0073] The present invention relates to a method for marking a material using a marking composition, for example for the purpose of authentication and / or traceability of said material, in particular batch-produced material.

[0074] As used herein, "material" or "object" refers to a material to be labeled (or subsequently labeled) using a labeling composition comprising at least a molecular tag.

[0075] The material may be an element, component, part or substance that constitutes or can be used to make something.

[0076] The marking compositions are particularly useful in authentication methods that allow differentiation and / or identification of materials.

[0077] Therefore, verification methods are suitable for distinguishing genuine materials from fake ones.

[0078] In addition, verification methods apply to the identification of materials (e.g. by batch type, manufacturing location, manufacturing time).

[0079] Generally, the marking method of the present invention is intended to confer a competitive advantage on the marked material, namely:

[0080] - authentication, indicating the origin of the material and preventing, inter alia, counterfeiting, and

[0081] - Traceability, which enables manufacturers and trading partners, as well as consumers and regulators, to identify, authenticate and track goods throughout the supply chain.

[0082] According to the present invention, the marking method comprises the following steps:

[0083] - a step of supplying a labeling composition selected from molecular tags, and

[0084] - A step of incorporating said marking composition into said material.

[0085] According to the invention, starting from a material containing a marking composition, said material can be subjected to a recovery step in order to recover the information carrier.

[0086] Preferably, the recycling step is performed on the material refined product to extract the information carrier.

[0087] As used herein, a "molecular tag" advantageously relates to a chemical compound suitable for identifying and / or authenticating a material of interest.

[0088] Molecular tags are constituent elements of the molecular composition of the material into which they are incorporated. They are an integral part of the body of material of interest.

[0089] In other words, the present invention also relates to the use of the marking composition for marking a material, for example for the purpose of authentication and / or traceability of the material, in particular batch-produced material.

[0090] Overview of Marking Compositions

[0091] In the present invention, the labeling composition comprises particles composed of molecular tags, which particles comprise a combination of at least three components:

[0092] - a homogeneous porous matrix containing a plurality of pores,

[0093] - an information carrier added to said homogeneous porous matrix, wherein said information carrier consists of a polymer comprising an information sequence, and

[0094] - The surface of the particles has a coating comprising at least one fluidizing agent, suitable for promoting a homogeneous dispersion of said particles within said material.

[0095] Typically and preferably, the combination of information carrier, homogeneous porous matrix and fluidizing agent is chosen to obtain self-association. Thus, no chemically reactive species are required to form covalent bonds, thereby promoting their dissociation.

[0096] In particular, "self-association" refers to the ability of chemical entities to interact naturally and spontaneously without the need for covalent chemical bonds or the presence of specific chemical reactants. It is a phenomenon in which chemical entities interact due to intermolecular forces (such as van der Waals forces, hydrogen bonds, or ionic interactions) without forming permanent chemical bonds.

[0097] This self-association occurs when molecules or compounds have functional groups or features that allow them to be attracted to each other. These interactions are reversible, meaning that the entities involved can subsequently dissociate without the need for complex chemical reactions.

[0098] In the present invention, the selected components, namely the information carrier, the homogeneous porous matrix and the fluidizing agent, are designed to interact spontaneously without the need for reactive chemicals to form covalent bonds. Because these interactions are not permanent, this simplifies the subsequent dissociation process, which is useful for the recovery of the information carrier.

[0099] Preferably, the marking composition comprises a "matrix-fluidizer" couple suitable for the information carrier and the target material.

[0100] As used herein, the term "particle" or "microparticle" refers to the smallest entity in a marking composition that can be identified as a microparticle by its appearance.

[0101] The particles advantageously have a size ranging from 200 nm to 100 μm, preferably from 500 nm to 10 μm, more preferably from 750 nm to 3 μm.

[0102] Preferably, the size is within any combination selected from the following values: 0.5μm, 1μm, 1.5μm, 2μm, 2.5μm, 3μm, 3.5μm, 4μm, 4.5μm, 5μm, 5.5μm, 6μm, 6.5μm, 7μm, 7.5μm, 8μm, 8.5μm, 9μm, 9.5μm, 10μm.

[0103] The term "size" advantageously refers to a physical characteristic dimension of the particle in connection with the method of measurement (or dimensional analysis by an appropriate technique), for example by laser diffraction or sieving or image analysis.

[0104] The size of the solid particles advantageously corresponds to their "equivalent spherical diameter" or "equivalent spherical diameter", that is to say advantageously the volume diameter (d v ), which is defined as the diameter of a perfect sphere having the same volume as the solid particle under analysis.

[0105] The particle size, in particular the particle size distribution, of these particles is advantageously determined by D 50 or d 50 The value definition of , also known as the "median size" or "median diameter".

[0106] D 50 represents the particle size for which 50% of the volume (or mass) has a smaller (or larger) particle size; in other words, D 50 It is the diameter corresponding to 50% of the cumulative frequency of number, mass or volume.

[0107] Thus, as used herein, "median size" with respect to a particle advantageously refers to the 50th percentile particle diameter in its volume size distribution (volume median particle diameter, D 50 ) (e.g., measured by a particle size analyzer based on laser diffraction / scattering spectroscopy).

[0108] Particle size distribution can be measured by laser diffraction using a Mastersizer 2000 or 3000 equipped with a Hydro 2000S system.

[0109] In general, the particles, preferably in combination with a fluidizing agent, advantageously form a protective agent for the information carrier.

[0110] As used herein, the term "protective agent" refers to a molecule capable of preventing information carriers from damage caused by environmental factors (such as UV light, ionizing radiation and genotoxic chemicals) or by naturally occurring agents (such as reactive oxygen species, reactive nitrogen species, reactive carbonyl species, lipid peroxidation products and alkylating agents).

[0111] The particles of the marking composition advantageously remain intact after incorporation into the material of interest.

[0112] As used herein, the term "remains intact" means that the particles are contained within the material of interest and the information carrier remains attached / associated to the homogeneous porous matrix.

[0113] The terms "remain intact" also include at least a portion of particles, wherein the homogeneous porous matrix is ​​fractionated or divided due to the marking process, but wherein the information carrier remains attached / associated to the homogeneous porous matrix.

[0114] In general, the concentration of information carrier in the particles is advantageously between 0.1 and 500 mg, preferably between 10 and 500 mg, of information carrier per g of homogeneous porous matrix.

[0115] Preferably, the concentration is within the range of any combination selected from the following values: 10 mg.g -1 , 50mg.g -1 , 100mg.g -1 , 150mg.g -1 , 200mg.g -1 , 250mg.g -1 , 300mg.g -1 , 350mg.g -1 , 400mg.g -1 , 450mg.g -1 , 500mg.g -1 .

[0116] Homogeneous porous matrix

[0117] The homogeneous porous matrix advantageously forms a "support", ie a compound that can be loaded with the information carrier and allows its incorporation into the target material.

[0118] The homogeneous porous matrix is ​​advantageously intended to carry the information carrier by confining it.

[0119] As used herein, "homogeneous" or "homogeneous" specifically means that the matrix does not contain any continuous layers (eg, does not contain a core-shell).

[0120] As used herein, "porous" means that the substrate is a solid having pores (ie, cavities, channels, or gaps) that are deeper than they are wide. The term "porous" also means that the substrate can adsorb the information carrier described herein.

[0121] Generally, the term "porous" as used herein also follows the IUPAC recommendations (Rouquérol, 1994) and refers to:

[0122] - for mesoporous particles, the free diameter of the pores is in the range of 2 to 50 nm, or

[0123] - For macroporous particles, the free diameter of the pores has a width greater than 50 nm.

[0124] The "free diameter" or "pore size" of a pore refers to the pore diameter measured in the dry state using measurement techniques (such as determination of nitrogen adsorption isotherms) and analytical methods (such as density functional theory (DFT) or the method of Barrett, Joyner and Halenda (BJH method)), which is used to extract the pore size distribution from the experimental isotherm based on the Kelvin model of pore filling.

[0125] For evaluation of larger pores, mercury intrusion porosimetry (pore sizes from 3.2 nm to greater than 400 μm) and capillary flow porometry (pore sizes from 13 nm to 500 μm) can be used.

[0126] Preferably, the homogeneous porous matrix of the present invention has:

[0127] - About 1 to about 1500m 2 / g maximum surface area, and / or

[0128] - a pore diameter ranging from 2 to 500 nm, preferably from 2 to 200 nm, more preferably in the range of 10 nm to 100 nm.

[0129] Preferably, the maximum surface area is within a range selected from any combination of the following values: 1, 10, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500 m 2 / g.

[0130] Preferably, the pore size is within a range selected from any combination of the following values: 2 nm, 10 nm, 20 nm, 30 nm, 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 90 nm, 100 nm.

[0131] Advantageously, the "surface area" is obtained by deriving the surface area from physical adsorption isotherm data applying the Brunauer-Emmett-Teller (BET) method. According to ISO 9277-1995, the specific surface area is measured by the BET method using nitrogen measurement.

[0132] Porosity can be further revealed and visualized in transmission electron microscopy (TEM) images. TEM images taken in brightfield imaging mode show variations in contrast between light and dark regions within a particle, with the brighter areas representing areas where electrons have less interaction with the material. These areas correspond to pores in the material.

[0133] More preferably, the size of the pores is compatible with the size of the information carrier.

[0134] In a preferred embodiment, the homogeneous porous matrix is ​​composed of at least 90% w / w of a salt, preferably an inorganic salt, consisting of:

[0135] - an ionic combination of a positively charged cation and a negatively charged anion, or

[0136] -Metal oxides, such as ZrO2.

[0137] As used herein, a mineral compound (salt, metal oxide) is considered "insoluble" if less than 1 gram of the mineral compound can be completely dissolved in 1 L of the target solvent. A table of insoluble salts in water can be found in Techniques de l'Ingénieur "Solubilité dans l'eau des composésminéraux", Réf: K590 v1 (1992) https: / / doi.org / 10.51257 / a-v1-k590: based on JH Perry, Chemical Engineers Handbook. For insoluble compounds, solubility is expressed in milligrams of anhydrous substance per liter of solvent (usually water) (which varies with temperature). For example, at 18°C, only 11.2 mg of CaCO3 can be dissolved in 1 L of water.

[0138] Anions are advantageously selected from carbonate, phosphate, sulfate, borate, silicate (e.g. CO3 2- PO4 3- 、SO42- , BO3 3- 、SiO3 2- etc.) or a combination thereof.

[0139] The cation is advantageously selected from calcium, magnesium, barium, aluminum, zinc, strontium, manganese (e.g., Ca 2+ Mg 2+ 、Ba 2+ 、Al 3+ 、Zn 2+ 、Sr2 + 、Mn 2+ etc.) or a combination thereof.

[0140] Preferably, the particles consist essentially (preferably more than 90% w / w) of calcium carbonate, calcium phosphate, barium sulfate, barium phosphate, barium carbonate, borosilicate, calcium silicate, manganese carbonate or combinations thereof.

[0141] The calcium carbonate is preferably mesoporous calcium carbonate, ie the chemical element CaCO 3 ,XH 2 O, which is crystallized in the form of particles and exhibits a structural characteristic organized in a network of channels with variable pore sizes.

[0142] Porous calcium carbonate particles are disclosed, for example, in:

[0143] -Volodkin DV,Petrov AI,Prevot M,Sukhorukov GB.Matrix polyelectrolytemicrocapsules: new system for macromolecule encapsulation.Langmuir.2004Apr 13;20(8):3398-406.doi:10.1021 / la036177z.PMID:15875874;

[0144] -Volodkin DV, Larionova NI, Sukhorukov GB.

[0145] -Anna Vikulina,Joseph Webster,Denis Voronin,Evgenii Ivanov,RawilFakhrullin,Vladimir Vinokurov,Dmitry Volodkin,Mesoporous additive-freevaterite CaCO3 crystals of untypical sizes:From submicron to Giant, Materials & Design, Volume 197, 2021, 109220, https: / / doi.org / 10.1016 / j.matdes.2020.109220;

[0146] -Ferreira AM, Vikulina AS, Volodkin D. CaCO3 crystals as versatile carriers for controlled delivery of antimicrobials.

[0147] The homogeneous porous matrix may also be selected from biological sources and homogeneous porous matrices of biological origin.

[0148] This homogeneous porous matrix is, for example, selected from coccoliths (coccoliths).

[0149] Coccoliths are produced by microalgae (e.g. Emiliania huxleyi); they exhibit suitable properties (e.g., 3 μm size, 4 nm pore diameter, and 19 nm 2 .g -1 specific surface area).

[0150] Cobblestones are for example published in:

[0151] -Jakob,I.,Chairopoulou,MA, M., Posten, C., & Teipel, U. (2017). Biogenic calcite particles from microalgae-Coccoliths as a potential raw material. Engineering in life sciences, 17(6), 605-612. DOI: 10.1002 / elsc.201600183, PMID: 28701909, or

[0152] -Lomora, M., Shumate, D., Rahman, AA, & Pandit, A. (2019). Therapeutic applications of phytoplankton, with an emphasis on diatoms and coccolithophores. Advanced Therapeutics, 2(2), 1800099; https: / / doi.org / 10.1002 / adtp.201800099.

[0153] Advantageously, the homogeneous porous matrix is ​​insoluble in said material (the matrix is ​​unable to form such a solution in said material).

[0154] The homogeneous porous matrix advantageously integrates other organic and / or inorganic chemical components.

[0155] The organic and / or inorganic chemical components may advantageously influence the properties of the particles, such as their size, their structure and their properties.

[0156] The organic and / or inorganic chemical components are, for example, selected from short polymers, fatty acids, short organic molecules such as ethylene glycol, or other inorganic salts.

[0157] The particles are advantageously decomposed by conditions which are inert with respect to the stability of the information carrier, for example:

[0158] - metal chelators, such as EDTA (ethylenediaminetetraacetic acid), EGTA (ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid), NTA (N,N-bis(carboxymethyl)glycine), DTPA (diethylenetriaminepentaacetic acid) or the natural and biodegradable L-glutamic acid diacetic acid (GLDA) and / or

[0159] - Specific pH conditions (such as acidic aqueous solutions), advantageously short exposure times (preferably less than 1 hour).

[0160] - Combination of mild acidic conditions and chelating agents.

[0161] Citric acid is a natural molecule derived from lemons that acts as both a calcium chelator and a weak acid. Therefore, the particles can be easily and quickly dissolved using a citric acid solution.

[0162] As used herein, the term "decompose" or "decomposition" advantageously refers to the dissolution (in a soluble salt or gas) of a homogeneous porous matrix using a metal chelating agent and / or using acidic pH conditions to release the information carrier.

[0163] According to IUPAC (Muller, P." Glossary of terms used in physical organic chemistry (IUPAC Recommendations 1994, Pure and Applied Chemistry, vol. 66, no. 5, 1994, pp. 1077-1184. https: / / doi.org / 10.1351 / pac199466051077), "chelate" refers to the formation or existence of two or more separate coordinate bonds between a polydentate (multiply bonded) ligand and a single central atom.

[0164] Typically, these ligands are organic compounds and are referred to as chelants (chelating agents), chelators, chelating agents, or sequestering agents.

[0165] Metal chelators are compounds that react with metal ions to form stable, water-soluble metal complexes. These agents rearrange the metal's chemical composition and increase its overall stability and potential for binding to other substances.

[0166] Preferably, the particles are decomposable by mildly acidic conditions, ie, pH 2 to 5, such as aqueous solutions of citric, acetic or hydrochloric acid.

[0167] Preferably, the pH is within a range selected from any combination of the following values: 2, 3, 4, 5.

[0168] DNA is easily depurinated (i.e., purine bases are lost from the DNA) at pH values ​​below 5. Therefore, chelating agents are preferred.

[0169] Acidic conditions are preferably used for a very short extraction period (preferably less than 1 hour) to facilitate the decomposition process caused by the chelating agent.

[0170] Thus, advantageously, particle decomposition does not require the handling of acids (eg, hydrofluoric acid, nitric acid, chlorosulfuric acid, perchloric acid, trifluoromethanesulfonic acid, fluoroantimonic acid) that are highly harmful to human health.

[0171] For example, porous calcium carbonate particles can be dissolved using:

[0172] -100 mM EDTA solution, pH 5, or

[0173] - an aqueous acetic acid solution having a pH of 3,

[0174] - 1% (w / v) citric acid aqueous solution with pH 2, due to Ca 2+ and CO3 2- Ions dissociate, which can immediately dissolve particles.

[0175] In particular, LDHs (layered double hydroxides) are excluded from the present invention since they do not form a homogeneous porous matrix.

[0176] In fact, LDH is not a porous particle. The layered structure of LDH creates interlayer spaces between layers, and these interlayer spaces can accommodate anions and water molecules. However, these spaces are not pores within the meaning of the present invention.

[0177] Information carrier

[0178] An information carrier is added to the homogeneous porous matrix, wherein the information carrier consists of a polymer comprising an information sequence.

[0179] The information carrier can be loaded onto and / or into the homogeneous porous matrix by adsorption or coprecipitation.

[0180] In other words, the information carrier is advantageously contained within the pores of the homogeneous porous substrate and / or on the surface of the homogeneous porous substrate.

[0181] In other words still, the information carrier is advantageously at least partially or completely contained within the pores of the homogeneous porous matrix.

[0182] The information sequence of the information carrier advantageously results from a transcoding of an ordered bit system resulting from an encoding of a set of original data.

[0183] As used herein, "transcoding" advantageously refers to the use of a code to represent information to ensure the integrity of the information (error detection and correction) and to guarantee the security of the information (encryption / ciphering). Therefore, such a code preferably has an error correction system.

[0184] The encryption method is disclosed, for example, in:

[0185] -Erlich et al., DNA Fountain enables a robust and efficient storagearchitecture,Science 355,950-954(2017),

[0186] -Cheng Kai Lim, Saurabh Nirantar, Wen Shan Yew, Chueh Loo Poh, Novel Modalities in DNA Data Storage, Trends in Biotechnology, Volume 39, Issue 10, 2021, Pages 990-1003. https: / / doi.org / 10.1016 / j.tibtech.2020.12.008, or

[0187] -Ceze, L., Nivala, J. & Strauss, K. Molecular digital data storage using DNA. Nat Rev Genet 20, 456-466 (2019). https: / / doi.org / 10.1038 / s41576-019-0125-3.

[0188] As used herein, the term "cryptographic method" refers to the process of converting data into an ordered binary code and then translating it into a chemical structure that will become the carrier of the information.

[0189] Nucleic acids (such as DNA), including nucleic acid analogs, can store digital information by designing a sequence of nucleotide bases that encodes the zeros and ones of the digital information. A variety of techniques and encoding schemes for representing digital information using nucleotide bases are known to those skilled in the art.

[0190] Advantages of using nucleic acids and / or the like (rather than another storage medium) to store digital information include information density and permanence.

[0191] In the order-checking and synthesis of nucleic acids and / or analogs, substitution, insertion and deletion errors may occur, and therefore it is necessary to protect the data from these errors. In addition, environmental factors such as reactive oxygen species or UV radiation may also alter the nucleotide sequence. Therefore, encoding and error correction codes (such as Huffman codes or Reed-Solomon codes) known to those skilled in the art can be used to overcome these shortcomings.

[0192] According to the encryption method, the ordered binary coded data is converted into a nucleic acid base sequence to obtain an information sequence.

[0193] Finally, the encrypted data can be decrypted by determining the nucleic acid base sequence of the information sequence.

[0194] As used herein, the term "encrypted data" advantageously refers to any data element (e.g., text, image, file) that can be encrypted or encoded in an information sequence and that can be deciphered from an information carrier recovered from a product and decoded or decrypted according to a selected method. The decoded or decrypted data can then be used to verify the nature of the material, or to authenticate the material, or to exclude counterfeit items.

[0195] Therefore, in a preferred embodiment, the information carrier is selected from nucleic acids and nucleic acid analogs, preferably DNA, and more preferably single-stranded DNA or double-stranded DNA.

[0196] As used herein, the term "nucleic acid" refers to an oligomer composed of nucleotides, which can be deoxyribonucleotides or ribonucleotides. These compounds can be natural or synthetically produced deoxyribonucleotides or ribonucleotides. Synthetically produced nucleic acids can have naturally occurring sequences or non-natural sequences. The terms "ribonucleic acid" and "RNA" refer to oligomers composed of ribonucleotides. The terms "deoxyribonucleic acid" and "DNA" refer to oligomers composed of deoxyribonucleotides.

[0197] The terms "nucleic acid," "polynucleotide," "target polynucleotide," and "target nucleic acid" are used interchangeably.

[0198] As used herein, the term "nucleic acid analogue" refers to a nucleic acid that can include one or more analogues (e.g., altered backbones, sugars, or nucleobases). Some non-limiting examples of analogues include: 5-bromouracil, peptide nucleic acids, heterologous nucleic acids, morpholino compounds, locked nucleic acids, diol nucleic acids, threose nucleic acids, dideoxynucleotides, cordycepin, 7-deaza-GTP, fluorophores (e.g., rhodamine or fluorescein linked to a sugar), thiol-containing nucleotides, biotin-linked nucleotides, fluorescent base analogues, CpG islands, methyl-7-guanosine, methylated nucleotides, inosine, thiouridine, pseudouridine, dihydrouridine, steviol glycosides, and tetanoside.

[0199] The information carrier preferably consists of a nucleic acid fragment, the size of which is preferably less than 500 bp, preferably in the range of 50 to 250 bp.

[0200] Sequencing is the primary form of information retrieval, or "reading," and its advancements have driven the feasibility of DNA data storage. The main methods for reading DNA include sequencing by synthesis (SBS) and third-generation sequencing methods involving single-molecule sequencing via nanopores or enzymatic well-based reactions.

[0201] Preferably, the information sequence comprises an identifiable sequence, also called a "detectable sequence", such as a nucleic acid or nucleic acid analog sequence, which can be detected and sequenced by specific techniques (such as hybridization, PCR technology, sequencing, capture, electrochemical detection).

[0202] Fluidizing agent

[0203] As mentioned above, the particles have a particle surface having a coating containing at least one fluidizing agent, suitable for promoting uniform dispersion (or homogeneous dispersion) of the particles in the material. In the present invention, "uniform dispersion" also includes "homogeneous dispersion" or "random dispersion", advantageously without particle aggregation.

[0204] "Uniformly dispersed" advantageously means that the particles are uniformly or at least approximately uniformly spaced or distributed within the material.

[0205] As used herein, a dispersion is defined as a system in which distributed particles of one material are dispersed in a continuous phase of another material. The two phases can be the same or different states of matter.

[0206] Dispersions are classified in several different ways, including the size of the particles relative to the particles of the continuous phase, whether sedimentation occurs, and the presence of Brownian motion.

[0207] Generally, dispersions of particles large enough to settle are called suspensions, while dispersions of smaller particles are called colloids and solutions. See "Terminology of polymers and polymerization processes indispersed systems (IUPAC Recommendations 2011)". Pure and Applied Chemistry. 83(12): 2229-2259 and Compendium of Polymer Terminology and Nomenclature (IUPAC Recommendations 2008) (2nd ed.). RSC Publ. p. 464 for IUPAC definitions.

[0208] As used herein, coating advantageously consists in the surface modification of the particles, ie by applying a chemical substance, preferably a surface coating, which changes the physicochemical properties of the surface, forming surface-treated particles.

[0209] Surface coating advantageously covers the surface of the particles with an adhesion layer, ie the fluidizing agent.

[0210] As used herein, a fluidizing agent refers to a particulate surface treatment agent selected to alter the handling characteristics of particles in a target material.

[0211] In the context of the present invention, the term "fluidizing agent" advantageously refers to a functionalizing agent that modifies the physicochemical properties of the particles in order to improve their incorporation into the target material.

[0212] The "fluidizing agent" is advantageously selected from crystallization inhibitors, dispersants, surface treatment agents, agents for enhancing the fluidity of microparticle dispersions, or combinations thereof.

[0213] Advantageously, the at least one fluidizing agent is selected in direct dependence on the properties of the target material, which can in particular be:

[0214] - hydrophobic materials (oils, plastics, waxes, hydrophobic polymers) or

[0215] - Aqueous media or hydrophilic materials (paints, thermosetting prepolymers, inks, milk, cement).

[0216] For example, to incorporate molecular tags into hydrophobic plastics such as PVC (polyvinyl chloride), a fluidizing agent is selected from fatty acids, such as stearic acid, to improve the dispersibility of the particles.

[0217] In contrast, for incorporating molecular tags into thermosetting prepolymers consisting of aqueous latex solutions, the fluidizing agent was selected from sodium polyacrylate, which improves the dispersion of particles in such materials.

[0218] Preferably, the present invention excludes fluidizing agents that form a covalently bonded layer (which makes it impossible to use chelating agents such as organosilanes or siloxanes to break down the particles).

[0219] In modifying the surface properties of the particles, the fluidizing agent advantageously acts on at least one particle property, and thus can be measured, namely:

[0220] - changing the surface charge (zeta potential) and thus the stability behavior and / or the interaction with other components of the material,

[0221] - changing the aggregation / sedimentation properties of the particles (alone in the solvent, or in interaction with other particles of the material),

[0222] - If the target material is a liquid, change the particle dispersibility. As used herein, particle dispersibility corresponds to the ease with which a powder can be dispersed into a liquid to achieve spatial uniformity and a desired particle size (see ISO / TS 22107:2021),

[0223] - if the target material is solid (no particle aggregation), changing the homogeneity of the particle dispersion. According to the invention, the physicochemical properties of the molecular tag can be adapted to the material to be labeled:

[0224] - Hydrophilic molecular tags can be generated using hydrophilic fluidizing agents, preferably for incorporation into hydrophilic materials, or

[0225] - Hydrophobic fluidizing agents can be used to generate hydrophobic molecular tags, preferably for incorporation into hydrophobic materials.

[0226] Advantageously, the fluidizing agent imparts properties to the particles that improve their dispersibility in the solvent or environment.

[0227] The zeta potential of a particle typically has a value in the range of -100 to +100 mV. The magnitude of the zeta potential is indicative of the colloidal stability of the solution. The default zeta potential analysis limits are +150 to -150 mV, respectively. See Tech Note: Zeta potential quality report for the Zetasizer Nano.

[0228] For example, the zeta potential of porous calcium carbonate microparticles can be changed from a negative zeta potential to a positive zeta potential by coating with a positive polymer.

[0229] The zeta potential of porous CaCO3 microparticles was measured to be -12.2 mV in water at a fixed pH. Poly(allylamine hydrochloride) (PAH, Mw ~70 kDa) fluidizer adsorbed onto the particle surface was sufficient to achieve a zeta potential value of +10 mV (see Volodkin DV et al., Matrix polyelectrolyte microcapsules: new system for macromolecule encapsulation. Langmuir. 2004 Apr 13; 20(8): 3398-406. doi: 10.1021 / 1a036177z. PMID: 15875874).

[0230] In another example, surface modification of nanoscale precipitated calcium carbonate nanoparticles (nanoPCC) by fluidizing agents such as alginate, pectin, acrylamide (A-PAM), or nanofibrillated cellulose (NFC) results in a charge reversal, manifested by a change in the sign of the zeta potential from positive to negative.

[0231] The addition of NFC to the nanoPCC dispersion induces a charge reversal, with the zeta potential shifting from approximately +10 mV to −20 mV at the fixed pH of the experiment.

[0232] The stability of the saturated dispersion was determined by measuring the change in turbidity, which is the measurement of light transmitted through the sample using a Turbiscan Ma 2000 instrument.

[0233] In the unmodified case, nanoPCC dispersions were unstable, as evidenced by a rapid increase in light transmittance (over 40% within a few minutes) due to the settling of flocculated particles. Pectin-modified nanoPCC and NFC-modified nanoPCC dispersions were stable, and no change in turbidity over time was observed.

[0234] The use of A-PAM and alginate also improved the stability of nanoPCC particles; no phase separation was detected, but a slight increase in light transmittance was observed (Δ turbidity ~8%).

[0235] In contrast, the addition of 1 mL of a 1% hydrophobic agent, such as alkenyl succinic anhydride (ASA), to the dispersion of unmodified nanoPCC particles caused the formed mixture to aggregate and phase separation was observed, as evidenced by a rapid increase in Δ turbidity (%), reaching 80% within 120 min.

[0236] This fluidizing agent ASA with a branched isoalkenyl chain (C14 to C22) is widely used, inter alia, in the surface sizing of paper, paperboard and cardboard and in the hydrophobization of cellulose fibers. This fluidizing agent destabilizes the particles in aqueous solvents but enhances their dispersibility in hydrophobic materials such as organic solvents or plastics (see T et al. Tailoring Surface Properties of Paper Using Nanosized Precipitated Calcium Carbonate Particles. ACS Appl. Mater. Interfaces 2011, 3, 9, 3725-3731. https: / / doi.org / 10.1021 / am200913t).

[0237] The effect of the fluidizing agent on the particles can be measured by a variety of techniques well known in the art depending on the parameter of interest.

[0238] For surface charge, the electrophoretic mobility of the sample under investigation can be advantageously measured by a ZetaSizer Nano ZS (Malvern, UK) operating at a wavelength of 633 nm. This device measures the electrophoretic mobility of the particles and converts it to zeta potential (ZP) using the von Smoluchowski equation. The results are expressed as the average of at least three independent measurements.

[0239] For the hydrodynamic radius, the particle average hydrodynamic diameter Dh of the particles is advantageously determined by DLS using a Zetasizer 3000 (Malvern, UK) equipped with a 10 mW He-Ne laser (633 nm) as a light source. The measurement is performed at a scattering angle of 90° and the reported results are the average of three independent measurements.

[0240] For the aggregation / sedimentation properties, an optical analyzer is advantageously used to study the suspension behavior and determine the sedimentation velocity (Turbiscan MA 2000, Formulaction, Toulouse, France). This instrument detects and measures changes in concentrated and opaque suspensions. The Turbiscan scans the entire sample vertically in steps using a pulsed near-infrared light source (λ = 850 nm) and converts the macroscopic aspects of the mixture into two graphs (see Mengual O, Meunier G, Cayré I, Puech K, Snabre P. TURBISCAN MA 2000: multiple light scattering measurement for concentrated emulsion and suspension instability analysis. Talanta. 1999 Sep 13; 50(2): 445-56. doi: 10.1016 / s0039-9140(99)00129-0. PMID: 18967735). Transmission and backscatter data are acquired from a transmission detector, which receives light that passes through the sample (at 0° to the incident beam), and a backscatter detector, which receives light backscattered by the sample at 135° to the incident beam. The stability of the dispersion is determined by measuring the change in turbidity, i.e., the light transmitted through the sample. The Turbiscan gives the backscattered light flux (Δ turbidity in % relative to an external standard) as a function of sample height. Turbiscan data can be reported as the change in transmitted light over time, measured from the middle of the test tube. Turbidity measurements are started immediately after dispersion preparation.

[0241] Turbiscan is also used to examine the dispersion of particles in liquids. The stability of the particle dispersion is evaluated in terms of the Turbiscan Stability Index (TSI) value.

[0242] Regarding examining the uniformity of particle dispersion in a solid material, the particles can be directly observed using a scanning electron microscope (SEM) or an atomic force microscope (AFM).

[0243] Regarding the recrystallization properties, advantageously depending on the size of the unstable porous support, the stability of the unstable porous support can be directly observed by microscopy techniques such as optical microscopy or scanning electron microscopy.

[0244] Regarding the surface area of ​​the microparticles, an advantageous standard practice is to derive the surface area from the physical adsorption isotherm data using the Brunauer-Emmett-Teller (BET) method. The specific surface area is measured by the BET method using nitrogen measurements according to ISO 9277-1995.

[0245] As regards the surface energy, the contact angle is advantageously measured by means of a video optical contact angle tester (CA; OCA25, Eastern-Dataphy, Germany).

[0246] In practice, the fluidizing agent advantageously comprises a wide variety of molecules, including coatings with organic molecules (such as fatty acids, polymers, resins) and / or coatings with inorganic salts.

[0247] In a preferred embodiment, the at least one fluidizing agent is selected from surface treatment agents for inorganic fillers based on hydrophilic materials or hydrophobic materials.

[0248] Fluidizing agents that impart hydrophobic properties to particles, thereby improving their dispersion in hydrophobic solvents or environments, are usually amphiphilic molecules or surfactants, such as:

[0249] - saturated fatty acids, advantageously with a carbon chain length of C15 to C20, such as palmitic acid and stearic acid,

[0250] - unsaturated fatty acids, advantageously with a carbon chain length of C15 to C20, such as oleic acid and linoleic acid,

[0251] - phospholipids, such as phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine,

[0252] - a lipid mixture, preferably of biological origin or biodegradable, such as lecithin;

[0253] - organic amphiphilic molecules with a lipophilic moiety, such as phosphonates, diols, alcohols, phenates, sulfonates, salicylates, succinic anhydride;

[0254] - waxes, such as beeswax, shellac, vegetable waxes;

[0255] - resin acids, such as alicyclic carboxylic acids and abietic acids, and their salts, esters and ethers;

[0256] - Bifunctional polymers with hydrophobic parts, e.g. or some filler coupling agents, such as titanate (LICA ) or zirconate

[0257] Fluidizing agents that improve or impart dispersion of particles in hydrophilic materials include:

[0258] hydrophilic polymers, for example polycarboxylic acids, molten phosphoric acid, polyacrylates, polyamines, polyacrylamides, polyaminoacyl groups, polystyrene, polyethylene glycol (PEG), polyvinylpyrrolidone (PVP), polymaleic acid (PMA), polyepoxysuccinic acid (PESA), polysaccharides (dextran, alginate, chitosan, chondroitin, cellulose, pectin, carboxymethyl inulin, etc.), peptides (for example polyaspartic acid, polyglutamic acid), proteins, lignin, preferably bioderived or biodegradable polymers (see Vroman I et al., Biodegradable Polymers. Materials (Basel). 2009; 2(2): 307-344. Published 2009 Apr. doi: 10.3390 / ma2020307),

[0259] - Other inorganic salts, such as calcium phosphate, hydroxyapatite.

[0260] As used herein, "biodegradable polymer" refers to a polymer that is susceptible to degradation by biological activity, with degradation being accompanied by a decrease in its molar mass, according to IUPAC Recommendations 2003.

[0261] For example, the particles may be treated or coated with a hydrophobic agent (such as, for example, an aliphatic carboxylic acid) or a surfactant.

[0262] Suitable fatty acids are, for example, C15 to C28 fatty acids, such as stearic acid, palmitic acid, myristic acid, lauric acid or mixtures thereof.

[0263] The particles may also be treated or coated with a hydrophilic polymer to render them cationic or anionic, for example with polyacrylates or polyaspartic acid (anionic) or polyarginine, polylysine (cationic).

[0264] Synthesis of particles of the labeling composition

[0265] As shown in the examples, the information carrier and the fluidizing agent are advantageously incorporated simultaneously from solution onto the homogeneous porous matrix.

[0266] For example, information carrier and fluidizing agent can advantageously be incorporated simultaneously by direct adsorption of macromolecules from solution onto a preformed homogeneous porous matrix (physisorption).

[0267] In an alternative embodiment, depending on the nature of the "fluidizing agent", the information carrier and the fluidizing agent can also be added sequentially to the porous carrier.

[0268] For example, in the case of fatty acids such as stearic acid, the information carrier and the fluidizing agent are added successively by direct adsorption.

[0269] Typically, the combination of information carrier, homogeneous porous matrix and fluidizing agent is advantageously self-associating and therefore does not require chemically reactive substances to form covalent bonds, thereby facilitating their dissociation.

[0270] Material

[0271] The present invention also relates to a material comprising the marking composition according to the invention.

[0272] In general, the material is advantageously chosen from liquid, semisolid or solid materials, such as plastics, thermosetting materials, varnishes, rubbers, lubricants, coatings, oils, pharmaceuticals, cosmetics, foodstuffs.

[0273] The material may be any solid traceable item, such as an electronic device, an item of clothing, paper, fiber or fabric, or any other item of commerce, or cash or valuables, whether in storage or in transit.

[0274] Furthermore, the commercial item to be labeled using the molecular tag may be a liquid such as ink, oil, dye, or spray.

[0275] As a commercial item, the product may be a commodity item such as paper, metal, wood, plastic, rubber, or powder.

[0276] Furthermore, the material may be a medicine or a food.

[0277] For example, the material is a viscous water-based prepolymer that can be irreversibly cured by heating, resulting in the formation of a thermoset material (“Cured Thermoset Polymers”, see PAC, 2004, 76, 889. (page 898 for Definition of Terms for Polymer Reactions and Functional Polymeric Materials (IUPAC Recommendations 2003))).

[0278] Conversely, the target material can be a thermoplastic. Unlike thermoset polymers, thermoplastics can be transformed without undergoing a chemical reaction. Thermal degradation does not occur, and the polymer's molecular structure remains unchanged during the transformation. Thermoplastics are advantageously defined in "ISO 7792-1:2012, Plastics — Thermoplastic polyester (TP) molding and extrusion materials — Part 1: Nomenclature system and basis for specifications."

[0279] In a preferred embodiment, the incorporation step advantageously consists of incorporating the marking composition into the material during the preparation of said material.

[0280] In other words, the present invention also relates to a method for the preparation of a material comprising the step of incorporating said marking composition into said material during its preparation.

[0281] Preferably, the material is a batch generated material. As used herein, "batch generated" advantageously refers to a manufacturing method in which a material is manufactured in specific groups or quantities over a period of time.

[0282] Batch materials are advantageously passed through a series of steps in a larger manufacturing process to produce the final desired material.

[0283] Typically, the concentration of the marking composition is advantageously between 0.1, preferably 0.5 and 150 mg of marking composition per L or kg of material (0.1 to 150 ppm, 1 ppm = 1 mg / kg).

[0284] Preferably, the concentration is within a range selected from any combination of the following values: 0.1, 0.5, 1, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150 mg of marking composition per L or kg of material.

[0285] Typically, the particles of the marking composition are advantageously intact within the material of interest.

[0286] Extraction and reading of information carriers

[0287] The material is advantageously processed to extract the particles and the information carrier is then extracted from said extracted particles.

[0288] Advantageously, the information sequence is retrieved from said material by means of an "information sequence extraction method", ie a method for obtaining an information sequence from a material.

[0289] Thus, the particles can be treated with the previously disclosed decomposition conditions (metal chelating agents or acidic pH conditions) to specifically release the information carrier.

[0290] For example, the calcium carbonate homogeneous porous matrix can be treated with an acidic solution (pH 2, preferably citric acid) or an EDTA solution, or a combination thereof, to release the information carrier.

[0291] Thus, in general, the extraction process is advantageously carried out by dissolving the homogeneous porous matrix in an aqueous solution of, for example, a complexing agent, followed by extraction and purification steps to recover the information carrier.

[0292] To extract the information carrier from the molecular tag containing the hydrophobic fluidizing agent embedded in the hydrophobic material, the extraction is advantageously performed by combining a hydrophobic solvent (e.g. toluene, chloroform, dichloromethane, ethyl acetate) with an aqueous chelating agent decomposition solution. The information carrier will be in the aqueous fraction.

[0293] Once the vector is solubilized, the extracted information carrier can be purified and amplified.

[0294] Finally, the reading step corresponds to sequencing the information carrier and decoding the information sequence by means of the chosen encoding method.

[0295] If the material being labeled is a solid, the particles are first extracted by dissolving or melting the material in a suitable solvent. The choice of solvent depends largely on the material of the product. Suitable solvents can be easily determined by those skilled in the art, including organic solvents and aqueous (acidic or basic) solvents.

[0296] Typically, an aqueous chelating agent solution or an aqueous acid solution is advantageously used to dissolve the particles.

[0297] Preferably, the organic solvent containing the particles can be brought into contact directly with the decomposition solution and then extracted (the information carrier is in the aqueous phase). Alternatively, an intermediate step can be performed to separate the particles from the organic solvent, for example by centrifugation or filtration.

[0298] Various other modifications may be made to the invention within the scope of the appended claims.

[0299] Example

[0300] Example 1: Synthesis of size-controlled mesoporous vaterite microparticles (support).

[0301] In a typical experiment, 0.33 M Na2CO3 solution (50 mL) was quickly poured into an equal volume of 0.33 M CaCl2 solution (50 mL) at 20°C and after vigorous controlled stirring (650 rpm) with a digital electronic overhead stirrer for 30 seconds, the white precipitate was filtered off on a Satorlon polyamide filter (pore size 0.2 μm).

[0302] The precipitate was thoroughly washed twice with pure water to remove excess ions and then washed twice with 100% ethanol to improve the dispersion characteristics.

[0303] The resulting powder was dried for about 12 hours at 55° C. This procedure produced highly homogeneous spherical CaCO 3 microparticles with an average diameter of 6 μm.

[0304] The size of the microparticles can be varied by changing parameters such as salt concentration, stirring speed or temperature.

[0305] In another typical experiment, 0.6 M Na2CO3 solution (50 mL) was quickly poured into an equal volume of 0.6 M CaCl2 solution (50 mL) at 20° C. After vigorous controlled stirring (650 rpm) using a digital electronic overhead stirrer for 30 seconds, the white precipitate was filtered off on a Satorlon polyamide filter (pore size 0.2 μm).

[0306] Applying the same filtration and drying procedure, highly homogeneous spherical CaCO3 particles with an average diameter of 2.5 μm were obtained.

[0307] Example 2: Generation and synthesis of information carrier DNA sequence.

[0308] Computer-generated short naked coding DNA sequences (usually 10 bp to 210 bp) are used to introduce digital information consisting of alphanumeric characters.

[0309] For example, digital information can be encoded into nucleobases (naked coding sequences) using the Reed-Solomon error correction algorithm. This format allows the storage of digital information in polynucleotide molecules.

[0310] On both sides of the naked coding sequence, 2 x 20 bp orthogonal (non-interacting) polymerase chain reaction primer binding regions are generated for amplifying DNA sequences. The naked coding sequence plus the 2 x 20 bp primer binding regions on both sides provide the information carrier sequence.

[0311] In a specific embodiment aimed at stabilizing DNA molecules, a DNA strand can be generated that is complementary to the sequence of the information carrier carrying the digital information.

[0312] Finally, double-stranded DNA information carriers (ranging from 50 bp to 250 bp) were chemically synthesized.

[0313] Example 3: Loading information carrier and hydrophilic fluidizing agent (surface functionalizing agent), synthesis of hydrophilic molecular tag:

[0314] Double-stranded nucleotide information carriers and fluidizing agents were incorporated by direct adsorption of macromolecules from solution onto preformed CaCO3 microspheres (physisorption).

[0315] In a typical experiment, DNA at a concentration of 15 μM and polyacrylic acid sodium salt (fluidizing agent) at 0.5% (w / w) relative to the mass of CaCO 3 microparticles were dissolved in loading buffer (KCl 0.5N, pH 6.5).

[0316] Then, 50 mg of CaCO 3 microparticles were resuspended in a 2 ml Eppendorf tube containing 500 μL of loading buffer, vortexed vigorously and incubated under stirring for at least 4 hours.

[0317] It is worth noting that the pH of the loading buffer was chosen to favor the interaction of the particles with negatively charged macromolecules, as the zeta potential of the microparticles changes from positive to negative between pH 8.0 and 9.0, which can be enhanced by excess Ca 2+ Therefore, at pH 6.5, the microparticles have a positive overall net charge.

[0318] After incubation, the microparticles were centrifuged to remove the loading buffer and washed once (5 min) with clean KCl 0.5N pH 6.5 buffer to remove unadsorbed macromolecules. The microparticles were washed twice with 70% ethanol solution and dried at 55°C in a fume hood.

[0319] DNA loading can be determined indirectly by measuring the disappearance of DNA absorbance (at 260 nm) in loading buffer and wash buffer.

[0320] In a typical experiment, 6 μm molecular tags were loaded at concentrations ranging from 0.1 to 2 μg DNA per mg inorganic carrier.

[0321] In another typical experiment, 2.5 μm molecular tags were loaded at a concentration range of 2 to 15 μg DNA per mg inorganic support (depending on the DNA loading solution concentration).

[0322] Example 4: Sequential loading of information carrier and then hydrophobic fluidizing agent, hydrophobic molecular tag synthesis :

[0323] In a typical experiment, DNA was dissolved in loading buffer (KCl 0.5N, pH 6.5) at a concentration of 15 μM.

[0324] Then, 50 mg of CaCO 3 microparticles were resuspended in a 2 ml Eppendorf tube containing 500 μL of loading buffer, vortexed vigorously and incubated under stirring for at least 4 hours.

[0325] After incubation, the microparticles were centrifuged to remove the loading buffer and washed once (5 min) with clean KCl 0.5N pH 6.5 buffer to remove unadsorbed macromolecules. The microparticles were washed twice with 70% ethanol solution and dried at 55°C in a fume hood.

[0326] DNA loading can be determined indirectly by measuring the disappearance of DNA absorbance (at 260 nm) in loading buffer and wash buffer.

[0327] In a typical experiment, 6 μM molecular tags were loaded at concentrations ranging from 0.5 to 10 μg DNA per mg inorganic carrier.

[0328] In another typical experiment, 2.5 μm molecular tags were loaded at a concentration range of 2 to 15 μg DNA per mg inorganic support (depending on the DNA loading solution concentration).

[0329] Stearic acid fluidizer was dissolved in ethanol / water solution (2 / 1, v / v) at 1 mg / mL at 60°C.

[0330] Once dried, the DNA-loaded microparticles were mixed with a portion of the stearic acid solution. The amount of stearic acid added was 2% (w / w), e.g., for 50 mg of DNA-loaded microparticles, 1 mg of stearic acid was added.

[0331] Once mixed with the stearic acid solution, the microparticles were incubated in a sealed tube (to prevent evaporation of ethanol and water) at 58°C with vigorous stirring for 4 hours.

[0332] After incubation, the microparticles were centrifuged to remove the stearic acid loading buffer and washed once (5 min) with clean ethanol / water 2 / 1 (v / v) solution to remove unadsorbed molecules. The microparticles were washed twice with 70% ethanol solution and dried at 55°C in a fume hood.

[0333] It can be observed that the DNA is not released during this process because the colorful fluorescent DNA remains well confined within the microparticles.

[0334] Example 5: Particle size analysis:

[0335] The size of the particles can be measured directly from DIC (differential interference contrast) or fluorescence microscopy images. Images were acquired on a Nikon H600L fluorescence microscope (Japan) using NIS Elements software version 4.40 equipped with an ANDOR 5.5Neo camera, a Spectra X-ray engine computer-controlled source Lumencor, and fluorescence filters (DAPI, FITC, RITC) - see Figure 1 .

[0336] Manual measurement of the average size of 200 microparticles gave corresponding sizes of 6.28 ± 0.59 μm for microparticles made with a 0.3 M salt concentration and 2.32 ± 0.32 μm for microparticles made with a 0.6 M salt concentration.

[0337] Example 6: Effect of fluidizing agent on compatibility, polydispersity and aggregation tendency of microparticles in aqueous medium (Figure 2)：

[0338] The hydrodynamic volume and diameter of microparticles diluted in distilled water can be measured using a Malvern Mastersizer 3000 (particle size distribution analyzer) particle size analyzer equipped with a Hydro 2000S system for measuring the size of microparticles diluted in a continuous stream of water.

[0339] First of all, we must point out that the hydrophobic microparticles tested (stearic acid fluidizer) are not compatible with these measurements because they are immiscible with water, whether empty or loaded with DNA. In fact, they float on water.

[0340] On the other hand, this type of measurement highlights the dispersing properties of the hydrophilic fluidizer sodium polyacrylate.

[0341] While microparticles without a fluidizer (calcium carbonate carrier alone) tended to form aggregates of varying sizes (6.5 μm or 90 μm for 2.5 μm particles), the addition of 0.5% w / w hydrophilic sodium polyacrylate fluidizer inhibited aggregate formation and produced a single population of non-aggregated microparticles with a hydrodynamic diameter of 2.96 μm.

[0342] The effect of the fluidizing agent (0.5% w / w sodium polyacrylate) on the sedimentation properties of the support was also confirmed by Turbiscan measurements at 10 mg / mL in water. While the bare support settled in water within 1 hour, the support modified with the hydrophilic fluidizing agent settled much more slowly (4 hours). In contrast, the support modified with the hydrophobic fluidizing agent (stearic acid 2% w / w) was no longer miscible with water and tended to float on the water surface.

[0343] Example 7: Direct release and amplification of nucleotide information carriers from microparticles:

[0344] One of the particular advantages of calcium carbonate particles is that they can be easily broken down and removed using an acidic pH (<5) and / or by adding chelating agents such as EDTA.

[0345] Therefore, in a typical experiment, microparticles were disaggregated using "disaggregation buffer" (EDTA 100 mM, pH 5).

[0346] The disintegration of the particles was almost instantaneous, as evidenced by the appearance of CO2 bubbles as the particles dissociated.

[0347] When all the carriers have decomposed, the opaque white solution becomes clear. If the particles are in an alkaline buffer (pH > 8), first, the pH of the solution must be lowered to below 7 before adding the chelating agent. A more concentrated chelating agent solution (500 mM EDTA or 200 mM EGTA, pH 5 solution) can be used. In addition, for relatively large amounts of carriers, the molar number of the chelating agent needs to be adjusted to achieve a balance between the chelating agent and the Ca 2+ The ions reach equimolarity.

[0348] After vigorous stirring for 5 min, the informative nucleotide vector can be easily purified by commercial DNA purification kits (NEB PCR Clean-up or Macherey-Nagel NucleoSpin Gel and PCR Clean-up) and then amplified by PCR.

[0349] Another option for releasing the information carrier is to use a 1% citric acid (w / v) solution at pH 2. Using citric acid (C6H8O7) to dissolve calcium carbonate (CaCO3) requires sufficient acid to react with the calcium carbonate and convert it into calcium citrate (Ca3(C6H5O7)2), carbon dioxide (CO2) and water (H2O) according to the following chemical equation:

[0350] 3CaCO3+2C6H8O7→Ca3(C6H5O7)2+3CO2+3H2O

[0351] The resulting calcium citrate (Ca3(C6H5O7)2) is a non-toxic calcium salt of citric acid used as a food additive (E333).

[0352] This purification step is necessary for the chelation of Mg 2+ ions to inhibit the PCR reaction, it is necessary to eliminate EDTA, EGTA or citric acid chelating agents.

[0353] Finally, to extract the digital information, the amplified DNA fragments simply need to be sequenced using a polynucleotide sequencer.

[0354] For hydrophobic molecular tags such as stearic acid-coated carriers, equal volumes of chloroform are added to the decomposition buffer and carrier. After vigorous stirring and separation of the two immiscible solvents (chloroform and aqueous decomposition solution), the information carrier is present in the aqueous phase and is ready for analysis.

[0355] Example 8: Demonstration of the effect of a combination of a homogeneous porous carrier and a fluidizing agent on DNA protection from chemically reactive genotoxic substances The protective effect of molecules (isothiocyanates) Figure 3 :

[0356] First, to study the protection and chemical reactivity in aqueous environments, double-stranded DNA (70-mer or 80-mer) modified at the 5' end with a free amino (amine) functional group (amino-DNA) was customized. Primary amines are highly reactive with isothiocyanates, molecules that react with 5' amino groups or the amino functional groups of the nitrogenous bases adenine and guanine. Fluorescent isothiocyanates (FITC (fluorescein isothiocyanate) and RITC (rhodamine isothiocyanate)) were then selected to quantify the reactivity of the molecules. Therefore, if the DNA is not well protected, it will fluoresce. In one specific example, three different amino-DNA samples were reacted with 100 equivalents of FITC or RITC for 48 hours. The first sample is a single amino-DNA, the second sample is the amino-DNA (per g carrier load 10mg amino-DNA) loaded in a 2.5 μm homogeneous porous calcium carbonate carrier, and the third sample is the amino-DNA loaded in a 2.5 μm homogeneous porous calcium carbonate carrier (per g carrier load 3.5mg amino-DNA) and functionalized with 0.5% (w / w) hydrophilic sodium polyacrylate fluidizer. In short, the DNA amount in sodium carbonate / sodium bicarbonate buffer (100 μL, pH 10) is adjusted to 25 μg. Then, 100 equivalents of RITC or FITC diluted in DMF (10 μL) are added, shaken vigorously, and incubated for 48 hours under stirring in the dark. The reaction is terminated by adding 100 μL Tris HCl buffer (500mM, pH 8) (quencher). The pH of the solution is then adjusted to 5 with diluted HCl solution. Finally, the sample is diluted to 500 μL with "decomposition buffer" (EDTA 100mM pH 5). The 500 μL sample solution must be clear, indicating that the microparticles are well dissolved. Otherwise, a small amount of acetic acid or dilute HCl must be added. Once the particles are dissolved, 500 μL of DNA sample is purified using a PD MiniTRAP G25 desalting column (Cytiva) equilibrated with water at pH 7 according to the manufacturer's instructions. The purified DNA is then lyophilized and reconcentrated in 100 mM Tris HCl buffer at pH 8. The concentration of each sample is adjusted to the same concentration and then eluted using a Fisherbrand TMThe analysis was performed using a horizontal mini gel electrophoresis system in a 4% agarose electrophoresis gel for 45 min. Fluorescein and rhodamine fluorescence can be observed before BET staining using a BioRad Gel Doc XR+ imager and Image Lab software. Once the gel was made, the DNA was repurified using a Macherey Nagel (NucleoSpin Gel and PCR Clean-up) purification kit, the concentration was readjusted to the same concentration, and the fluorescence could then be measured using an EnSpire multimode plate reader (Perkin) microplate reader. All experiments performed gave the same results: DNA alone and DNA loaded in a homogeneous porous calcium carbonate support reacted with isothiocyanates (RITC or FITC) in the same manner. Therefore, the homogeneous porous support alone was unable to protect the DNA from the effects of such small reactive molecules. In contrast, when 0.5% (w / w) of sodium polyacrylate fluidizing agent was added, more than 80% of the DNA was protected and did not react with the fluorescent molecules. The synergistic tryptic association of three entities (information carrier+carrier+fluidizing agent) forms a molecular tag in which more than 80% of the DNA that can provide useful information is protected.

[0357] Example 9: Incorporation of molecular tags into thermosetting polymers:

[0358] The basic formulation of the thermosetting material was kindly provided by an industrial partner. While the detailed composition cannot be disclosed for proprietary reasons, it is important to note that the pH of this material is alkaline (pH>9). In a typical experiment, a preloaded 2.5 μm hydrophilic molecular tag (a homogeneous porous calcium carbonate support loaded with 70 bp double-stranded DNA at 10 mg / g and functionalized with 0.5% (w / w) hydrophilic sodium polyacrylate fluidizer) was diluted directly into an alkaline aqueous thermosetting prepolymer at a selected dilution factor (e.g., 50 mg molecular tag per L prepolymer). The aqueous mixture was then vigorously stirred to facilitate particle dispersion and homogenization.

[0359] Subsequently, curing (polymerization and cross-linking of the prepolymer) of the thermosetting prepolymer containing the molecular tag can be achieved by heat treatment (at 100°C for 3 hours), resulting in the formation of a "tagged" thermosetting material.

[0360] The resulting polymerized insoluble thermosetting material is a solid. Typically, 250 μl of heated thermosetting prepolymer gives 50 mg of solid thermosetting material.

[0361] Example 10: Extraction and amplification from cured thermoset materials:

[0362] The polymerized thermoset material can be treated to extract the information nucleotide carrier.

[0363] Briefly, 50 mg of solid was pulverized into micron particles using a cryogenic grinder (CryoMill, RETSH) to facilitate the contact between the “lysis buffer” (EDTA 100 mM, pH 5) and the molecular tag.

[0364] For the same reason, solid thermosetting microparticles (50 mg) were "pre-swelled" in 600 μL of chloroform (or toluene) for 1 hour.

[0365] Then, add 200 μL of "lysis buffer" and vigorously stir the sample for at least one hour to disintegrate the calcium carbonate support. To increase the yield of DNA purification from highly diluted DNA samples, add Sigma's poly(A) at 1 μg / mL to the "lysis buffer" during the extraction process. Poly(A) serves as a support for quantitative precipitation or purification of DNA and RNA.

[0366] After decantation, in the case of chloroform, the nucleotide information carrier is in the upper aqueous phase, while the hydrophobic components that can interfere with the PCR reaction are in the lower phase.

[0367] In order to eliminate EDTA, which would also inhibit the PCR reaction, the aqueous phase containing the message carrier was purified using a classic DNA purification kit (NEB PCR Clean up) before allowing the PCR reaction to amplify the message.

[0368] The purified DNA can then be sequenced.

[0369] Example 11: Incorporation of molecular tags into biodegradable thermoplastic materials such as polylactic acid (PLA) or polycaprolactone Polyester (PCL) or DL-lactide and glycolide copolymer (PDLG) or L-lactide and ε-caprolactone copolymer (PLC)):

[0370] GMP grade thermoplastic polymers and copolymers were purchased from Corbion In a typical experiment, PDLG 7507 or PDLG 5010 (a 75:25 molar ratio of DL-lactide and glycolide copolymer) particles were completely dissolved in chloroform at a concentration of 10 wt% (10 g copolymer diluted in 100 mL chloroform) by stirring at 40°C for 60 minutes. 1 mg of pre-loaded hydrophobic molecular tag (10 μg information DNA per mg of 2.5 μm CaCO3 support, 2% w / w stearic acid fluidizer) was diluted in 1 mL of ethanol, vortexed, and then directly incorporated into the 10% w / v copolymer / chloroform solution under vigorous mixing (10 min). After this stage, the prepared mixture was poured into a clean Teflon dish and the chloroform was allowed to evaporate at ambient temperature and pressure in a chemical fume hood.

[0371] Finally, the obtained solid film was dried in an oven at 60 °C for 48 h.

[0372] First, the final concentration of the molecular tag was 100 ppm (100 mg in 1 kg), and the concentration of the DNA carrier providing useful information (10 μg in 10 g of copolymer) corresponded to 1 ppm.

[0373] To verify whether this holds true at higher dilutions, we tested molecular tag concentrations of 2 ppm, 1 ppm, and 0.1 ppm (20, 10, and 1 ppb DNA, respectively, relative to the target material).

[0374] We also compared hydrophilic molecular tags (fluidizer: sodium polyacrylate) with hydrophobic molecular tags (fluidizer: stearic acid). Microscopic observation within molded thermoplastic polymers showed that the hydrophilic tags tended to form small clusters of 3 to 5 particles, while the hydrophobic tags were well dispersed (no aggregates).

[0375] Example 12: Extraction and amplification of information carriers from thermoplastic materials Figure 4 :

[0376] Solid thermoplastic materials (dried polymers or copolymers) can be processed to extract information nucleotide carriers. Briefly, 100 mg of solid was cut and placed in a bottle. 500 μL of chloroform was then added to the bottle, and the polymer was redissolved under vigorous stirring. Subsequently, 500 μL of "decomposition buffer" (EDTA 100 mM, pH 5, supplemented with 1 μg of polyA carrier) was added, and the bottle was stirred for 1 hour. After decantation, the nucleotide information carrier was in the upper aqueous phase, so 300 μL of the aqueous phase was removed using a sterile syringe, and the DNA was subsequently purified using the Macherey-Nagel NucleoSpin Gel and PCR Clean-up kit.

[0377] In 96-well Applied PCR was performed on a thermal cycler for 25 cycles (98°C for 10 s; 52°C for 30 s; 72°C for 10 s). Briefly, 5 μL of purified DNA sample was added to 25 μL of HF buffer (New England Biolabs), 15 μL of PCR-grade water (Roche), and 5 μL of primer mix (forward and reverse 35 bp primers, 10 μM each). High-fidelity PCR master mix ( High-Fidelity PCR Master Mix).

[0378] Once amplified, use DNA fragments were visualized by electrophoresis on a 4% agarose gel for 45 min using a horizontal mini gel electrophoresis system and by BET staining using a BioRad Gel Doc XR+ imager and Image Lab software.

[0379] All experiments were valid regardless of the type of thermoplastic tested (PLA, PLC, PDLG & PCL). It should be noted that a negative control was performed with unlabeled thermoplastic to ensure that the extracted DNA was not due to contamination within the laboratory. The purified DNA can then be sequenced.

[0380] Example 13: Incorporation of molecular tags and information carrier extracts into commercially available nail polish:

[0381] In a typical experiment, pre-loaded 2.5 μm hydrophilic molecular tags (homogeneous porous calcium carbonate supports loaded with 70 bp double-stranded DNA at 10 mg / g and functionalized with 0.5% (w / w) hydrophilic sodium polyacrylate fluidizer) were diluted into the same unloaded support microparticles at a selected dilution (e.g., 1% w / w). This mixture was then directly incorporated into 5 mL of commercially available nail polish (Yves Rocher's "Go Green") at a selected dilution (e.g., 5 to 0.1 ppm of nail polish). ) and stirred vigorously to facilitate the dispersion of molecular tags. Then, nail polish was applied on the ceramic ball (MP 1 / 4 inch ceramic balls) and dried in an oven overnight. Specifically, after drying (evaporation of the solvent in the nail polish, such as ethyl acetate), the dry mass accounts for 30% of the deposited mass of the varnish (50 mg).

[0382] Next, for extraction, the colored ceramic spheres were placed directly into ethyl acetate solvent (400 μL) and stirred, resulting in decolorization of the ceramic spheres. Then, 400 μL of "decomposition buffer" (100 mM EDTA, pH 5, supplemented with 1 μg of Poly(A)) was added and the sample was vigorously stirred for at least one hour to decompose the calcium carbonate carrier.

[0383] After decantation assisted by rapid centrifugation (1 minute at 10,000 rpm), the nucleotide information carrier is in the lower aqueous phase, while the hydrophobic components that can interfere with the PCR reaction are in the organic ethyl acetate upper phase. Finally, before performing the PCR reaction that allows the amplification of the information, the information nucleotide carrier is purified from the aqueous lower phase using a classic DNA purification kit (NEB PCR Cleanup).

[0384] This example was reproduced and confirmed using molecular tags diluted to 0.1 ppm (1 ppb DNA).

[0385] Example 14: DNA sequencing and digital data reading:

[0386] The amplified and purified polynucleotide information carrier can be sequenced using a polynucleotide sequencer.

[0387] Finally, the digital information can be decrypted by using the computational algorithm selected in Example 2 to read and re-extract the alphanumeric data from the nucleotide sequence.

Claims

1. A method for marking a material, for example for the purpose of authentication and / or traceability of the material, in particular of a batch-produced material, in, Marking methods include: - a step of supplying a labeling composition selected from molecular tags, and - a step of incorporating said marking composition into said material, Characterized in that the marking composition comprises particles which advantageously remain intact after incorporation into the material, said particles comprising: - a homogeneous porous matrix containing a plurality of pores, - an information carrier added to said homogeneous porous matrix, wherein said information carrier consists of a polymer comprising an information sequence, and - The surface of the particles has a coating comprising at least one fluidizing agent, suitable for promoting a homogeneous dispersion of said particles within said material.

2. The marking method according to claim 1, characterized in that The homogeneous porous matrix consists of at least 90% w / w of a salt, preferably an inorganic salt, consisting of an ionic combination of a positively charged cation and a negatively charged anion or a metal oxide, wherein the homogeneous porous matrix is ​​insoluble in the material.

3. The marking method according to any one of claims 1 or 2, characterized in that The particles are decomposed by metal chelators or by specific pH conditions, preferably mildly acidic conditions, even more preferably pH 2 to 5, even more preferably aqueous solutions of citric acid, acetic acid or hydrochloric acid.

4. The marking method according to any one of claims 1 to 3, characterized in that The particles have a size in the range of 200 nm to 100 μm, preferably 500 nm to 10 μm, more preferably 750 nm to 3 μm.

5. The marking method according to any one of claims 1 to 4, characterized in that The particles have a pore size in the range of 2 nm to 500 nm, preferably in the range of 2 to 200 nm, more preferably in the range of 10 nm to 100 nm.

6. The marking method according to any one of claims 1 to 5, characterized in that The information carrier is contained within the pores of the homogeneous porous matrix and / or adsorbed on the surface of the homogeneous porous matrix.

7. The marking method according to any one of claims 1 to 6, characterized in that The information sequence of the information carrier is obtained by transcoding an ordered bit system, and the ordered bit system is obtained by encoding a group of original data.

8. The marking method according to any one of claims 1 to 7, characterized in that The information carrier is selected from nucleic acids and nucleic acid analogs, preferably DNA, and more preferably single-stranded DNA or double-stranded DNA.

9. The marking method according to claim 8, characterized in that The information carrier consists of a nucleic acid fragment which preferably has a size of less than 500 bp, preferably in the range of 50 to 250 bp.

10. The marking method according to any one of claims 1 to 9, characterized in that The information carrier is associated with the homogeneous porous matrix, preferably adsorbed by the homogeneous porous matrix and / or contained by the homogeneous porous matrix.

11. The marking method according to any one of claims 1 to 10, characterized in that: The fluidizing agent is selected from the surface treatment agents of inorganic fillers, including saturated fatty acids, unsaturated fatty acids, phospholipids, lipid mixtures, organic amphiphilic molecules with lipophilic parts, amphiphilic polymers, resin acids, waxes, hydrophilic polymers, peptides and proteins, polysaccharides, inorganic salts, more preferably, the fluidizing agent is selected from fluidizing agents that improve dispersibility in hydrophobic materials or hydrophilic materials.

12. The marking method according to any one of claims 1 to 11, characterized in that The material is selected from liquid, semi-solid or solid materials, such as plastics, thermosetting materials, varnishes, rubbers, coatings, oils, lubricants, pharmaceuticals, cosmetics, foods.

13. The marking method according to any one of claims 1 to 12, characterized in that The incorporating step consists of incorporating the marking composition into the material during the preparation of the material.

14. A marking composition for marking a material, for example for the purpose of authentication and / or traceability of said material, in particular of batch-produced material, in, The marking composition comprises particles which advantageously remain intact after incorporation into the material, the particles comprising: - a homogeneous porous matrix containing a plurality of pores, - an information carrier added to said homogeneous porous matrix, wherein said information carrier consists of a polymer comprising an information sequence, and - The surface of the particles has a coating comprising at least one fluidizing agent, suitable for promoting a homogeneous dispersion of said particles within said material.

15. A material comprising the marking composition according to claim 14 or a marking composition obtained by the marking method according to any one of claims 1 to 13.

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