FR3118627A1 - Labeling of semi-crystalline nano- and/or micro-fibrillated cellulose with chitosan - Google Patents

Abstract

The invention relates to a process for labeling semi-crystalline nano- and/or micro-fibrillated cellulose (NMFC) which consists in adding nanoparticles of chitosan functionalized with a benzaldehyde derivative to a liquid dispersion of semi-crystalline NMFC. The invention also relates to a method for detecting this labeled semi-crystalline NMFC, as well as a cellulose-based substrate coated with a layer of said labeled semi-crystalline NMFC. Figure for abstract: Fig. 1.

Description

Labeling of semi-crystalline nano- and/or micro-fibrillated cellulose with chitosan

The present invention relates to a method for labeling semi-crystalline nano- and/or micro-fibrillated cellulose, a method for detecting this labeled cellulose, and a substrate coated with this labeled cellulose.

For several years, scientists and industrialists have been developing materials derived from plant or animal biomass, or even bio-inspired materials, as alternatives to materials derived from petrochemicals. The field of packaging, and especially packaging in the food industry, is very illustrative of this tendency to abandon materials of fossil origin in favor of materials based on biopolymers. Indeed, packaging for the packaging and preservation of goods (for example raw materials or finished products) are not only subject to health regulations but are also the source of economic and environmental problems which oblige their manufacturers to eco-responsible approach by designing materials that are biodegradable, recyclable and carbon neutral.

Cellulose is the most abundantly produced renewable organic material on the planet, which makes it a raw material of choice for the design of new biodegradable materials. Cellulose is a fibrous substance, rigid and insoluble in water. It is the main component of plant cell walls.

Cellulose has the following chemical structure:

It is known to transform cellulose fibers by chemical treatment, followed by acid hydrolysis, so as to obtain cellulose nanocrystals (whose acronym is “NCC”), or in other words nanocrystalline cellulose or cellulose nanocrystalline.

Cellulose films can be transformed by mechanical treatment, followed by defibrillation, so as to obtain nano- and/or micro-fibrillated cellulose (whose acronym is "NMFC") semi-crystalline, or in other words semi-crystalline cellulose nano- and/or micro-fibrils. In the remainder of the text, nano- and/or micro-fibrillated cellulose will be designated more simply by its acronym “NMFC”.

NCC and semi-crystalline NMFC can be used in many and various applications due to their very advantageous mechanical properties (in particular as a reinforcing agent) and barriers, for example for food packaging.

More specifically, the semi-crystalline NMFC is particularly advantageous, because it makes it possible to provide, on the one hand, an impermeability property, in particular to water, thanks to its crystalline part, and, on the other hand, a barrier property. to oxygen thanks to its amorphous part. These barrier properties are all the more interesting when the amorphous part has a low free volume fraction and a high cohesive density.

These properties of semi-crystalline NMFC are used in particular in the food and medical fields for antibacterial packaging, but also in electronic devices. For example, certain papers and/or films, in particular cellulose-based films, dedicated to the field of food, can be covered with a thin layer of semi-crystalline NMFC to ensure a barrier effect against the diffusion of oxygen.

However, this thin layer of semi-crystalline NMFC on the substrate on which it is deposited is not always persistent and can therefore slowly disappear over time. However, when the substrate is cellulose in nature, because this thin layer of semi-crystalline NMFC is composed of the same chemical elements (namely carbon and oxygen) as the cellulose-based substrate, the semi-crystalline NMFC is not therefore not distinguishable from the substrate, both chemically and in terms of the size of the cellulose fibers. Moreover, because semi-crystalline NMFC is sometimes only nanometer in size compared to cellulose, this also does not make it distinguishable by microscopic observation, especially by scanning electron microscopy or transition electron microscopy.

This is why there is a real need to have means for evaluating the persistence of a thin layer of semi-crystalline NMFC when it is deposited on a substrate, in particular a cellulose-based substrate. These means may consist of labeling the semi-crystalline NMFC which must be homogeneous and stable over time.

In this respect, patent FR 3 041 353 B1 describes a process for labeling semi-crystalline NMFC with metallic nanoparticles which consists of:
- add to a dispersion of semi-crystalline NMFC a metallic salt (for example HAuCl ₄ or AgNO ₃ ),
- stir the mixture until the metallic salt dissolves,
- add a reducing agent to the mixture thus obtained,
- Subjecting said mixture to microwave irradiation.

However, this marking method is not fully satisfactory for the following reasons:
- it is complex due to its multiple stages (namely dissolving a metal salt, adding a reducing agent, then microwave irradiation) and the different constituents used (namely a metal salt and a reducing agent) to allow the complexation of metal ions and the formation of bonds of the cellulose-OM type (M being a metal, for example Au or Ag);
- the constituents used during the process are not all derived from renewable natural resources;
- the marking means consisting of a metal salt, for example a silver or gold salt, is expensive;
- The detection of this marking means requires analysis under the microscope, spectrometry or even spectroscopy and therefore cannot be done immediately with a simple detection device.

Patent FR 3 053 055 B1 partly overcomes the drawbacks of the labeling process described in patent FR 3 041 353 B1 by proposing another process for labeling semi-crystalline NMFC which consists of:
- adding a chromophore organic molecule to a dispersion of semi-crystalline NMFC,
- subjecting the mixture thus obtained to microwave irradiation under conditions conducive to the attachment of the chromophore organic molecule to the semi-crystalline NMFC.

Depending on the chromophore organic molecule used, the semi-crystalline NMFC thus labeled can easily be detected either visually in visible light, or optically by fluorescence by exposing the labeled semi-crystalline NMFC to a wavelength corresponding to the maximum d absorption of the chromophore molecule to obtain maximum fluorescence intensity.

However, some of the chromophore molecules described in patent FR 3 053 055 B1, such as Propyl Astra Blue Iodide, IR-676 iodide, Indocyanine Green and Rhodamine 6G perchlorate, have the disadvantage of being photosensitive. Thus, the semi-crystalline NMFC labeling process described in this French patent proves not always to be stable over time.

The inventors of the present invention have sought to overcome these drawbacks detailed above of the known processes for labeling semi-crystalline NMFC by developing a new labeling process which, in addition to being perfectly stable over time, has the advantages of being simple, rapid, inexpensive to implement with biosourced molecules, as well as effective with a homogeneous distribution of the labeling means in the semi-crystalline NMFC.

The present invention thus has as a first object a process for labeling semi-crystalline NMFC which is characterized in that it consists in adding to a liquid dispersion of semi-crystalline NMFC nanoparticles of chitosan which has been functionalized by reaction of its groups primary amines with a benzaldehyde derivative which has a hydroxyl group in the ortho position to the aldehyde function.

Chitosan, like chitin from which it is produced by chemical deacetylation in an alkaline or enzymatic medium, are linear copolymers of random distribution of D-glucosamine linked in ß-(1-4) (deacetylated unit) and of N-acetyl-D-glucosamine (acetylated unit). Chitosan and chitin differ in their degree of acetylation.

Chitosan has the following chemical structure:

Chitin has the following chemical structure:

Chitosan and chitin thus have chemical structures very close to that of cellulose, which was detailed above.

Chitin is the component of the cell walls of fungi and algae, the exoskeletons of arthropods (crustaceans) and insects or even the endoskeletons of cephalopods (e.g. squid). Chitin is the most abundant biopolymer after cellulose. Chitosan is produced mainly from crustacean waste and the industrial processing of mushrooms.

Chitosan thus has the advantage of being a polymer obtained from a very abundant biopolymer.

Due to the chemical structural analogy between semi-crystalline NMFC and chitosan, during the process according to the invention, a chemical affinity by single bonds (of the Van der Waals type, hydrogen or electrostatic bridges) is created between these two polymers so that the chitosan is permanently attached to the semi-crystalline NMFC, and this in a perfectly homogeneous manner.

Thus, due to its bioavailability and its chemical affinity with semi-crystalline NMFC, chitosan constitutes a means of labeling of choice for semi-crystalline NMFC. The labeling process according to the invention has the advantage of not modifying either chemically or structurally the semi-crystalline NMFC.

In the context of the present invention, it is essential that the chitosan be functionalized by reaction of its primary amine groups with a benzaldehyde derivative which has a hydroxyl group in the ortho position with respect to the aldehyde function. Indeed, thanks to this hydroxyl group in the ortho position, a fluorescence maximum can be observed in an appropriate spectral window (for example in the visible) which corresponds to the absorption maximum of the benzaldehyde derivative when it is irradiated at a length given excitation wave. This fluorescence maximum induced by the functionalization of chitosan with the benzaldehyde derivative is more intense and out of step with the respective fluorescence emissions of cellulose and chitosan which are close to each other due to the similarity of their chemical structures.

This functionalization of chitosan with a benzaldehyde derivative as described above is thus essential in order to be able to optically detect the semi-crystalline NMFC labeled with chitosan nanoparticles, and thus to be able to easily distinguish it from the cellulose-based substrate on which it is filed. Indeed, the semi-crystalline NMFC thus labeled with the functionalized chitosan can be detected optically by measuring fluorescence or fluorescent revelation.

The semi-crystalline NMFC used during the labeling process according to the invention may have been obtained by mechanical treatment of films of any type of cellulose. This means that the cellulose can come from any source known to those skilled in the art. In other words, any source of cellulose raw material can be used. The raw material can come from any plant material that contains cellulose. The plant material can be wood. The wood can come from softwood trees like spruce, pine, fir, larch, Douglas fir or hemlock, or from hardwood trees like birch, aspen, alder, eucalyptus or acacia, or a mixture of softwoods and hardwoods. Plant material can also come from agricultural residues, grasses or other plant material such as straw, leaves, bark, seeds, husks, flowers, cottonseed vegetables or fruits, corn , wheat, oats, rye, barley, rice, flax, hemp, jute, ramie, bagasse, bamboo or reed.

The semi-crystalline NMFC can have fiber lengths of between 0.5 μm and 100 μm, preferably between 1 μm and 50 μm, more preferably between 5 μm and 10 μm. These fibers can have a diameter of between 1 nm and 100 nm, preferably between 5 nm and 50 nm, more preferably between 10 nm and 30 nm.

The semi-crystalline NMFC can be chosen from the semi-crystalline NMFC marketed by the company INNVENTIA or by the company FPInnovations.

The liquid dispersion of semi-crystalline NMFC may have been prepared, prior to the addition of functionalized chitosan nanoparticles, by diluting, preferably with stirring, semi-crystalline NMFC in a liquid. Preferably, the liquid is water. Semi-crystalline NMFC is indeed perfectly dispersible in water.

Besides water, the liquid can be chosen from solvents which are miscible with water. For example, it can be ethanol, acetone or tetrahydrofuran. Very advantageously, the liquid is water so that the marking process according to the invention is perfectly ecological.

The mass content of semi-crystalline NMFC in the liquid dispersion can be between 0.1% and 99%, preferably between 0.5% and 10%, more preferably between 0.5% and 5%, relative to the total mass of the liquid dispersion. This liquid dispersion therefore contains the semi-crystalline NMFC, as well as the functionalized chitosan nanoparticles, the technical characteristics of which and the method of their addition to the said liquid dispersion are detailed just below. Depending on the application of the labeling method according to the invention referred to, it is entirely within the reach of those skilled in the art to choose the appropriate mass content of semi-crystalline NMFC. Indeed, depending on the applications envisaged, it may be necessary for the mass content of semi-crystalline NMFC to be high, for example of the order of 50%, whereas for other applications, a much lower mass content is required. , for example of the order of 0.5%.

In the context of the present invention, the term “chitosan nanoparticles” means chitosan nanoparticles whose size is between approximately 1 nm and 1000 nm.

The chitosan nanoparticles are advantageously chitosan nanoparticles whose CAS number is 148411-57-8, the average molar mass is 5000 g/mol and which are for example supplied by the company SIGMA ALDRICH. The CAS number is the unique registration number of this polymer with the Chemical Abstracts Service database (the acronym of which is “CAS”).

Preferably, the benzaldehyde derivative is chosen from salicylaldehyde, 2,4-dihydroxybenzaldehyde, 5-iodosalicylaldehyde, 3-bromo-5-chlorosalicylaldehyde, 2-hydroxy-5-nitrobenzaldehyde, 3-tert-butyl- 2-hydroxybenzaldehyde, ethyl 3-formyl-4-hydroxybenzoate and 2-hydroxy-3-methoxybenzaldehyde.

Salicylaldehyde has the following chemical formula:

2,4-dihydroxybenzaldehyde has the following chemical formula:

5-iodosalicylaldehyde has the following chemical formula:

3-bromo-5-chlorosalicylaldehyde has the following chemical formula:

2-Hydroxy-5-nitrobenzaldehyde has the following chemical formula:

3-tert-Butyl-2-hydroxybenzaldehyde has the following chemical formula:

Ethyl 3-formyl-4-hydroxybenzoate has the following chemical formula:

2-Hydroxy-3-methoxybenzaldehyde has the following chemical formula:

Most preferably, the benzaldehyde derivative is ethyl 3-formyl-4-hydroxybenzoate.

The chitosan may have been wet-functionalized in ethanol with an ethanol-soluble benzaldehyde derivative or alternatively using a supercritical fluid (e.g. supercritical carbon dioxide) with a supercritical fluid-soluble benzaldehyde derivative .

The functionalization of chitosan by wet process can be carried out in the following way:
- chitosan nanoparticles (for example 8 mmol) are dispersed under strong magnetic stirring in ethanol (for example in 30 mL);
- a benzaldehyde derivative having a hydroxyl group in the ortho position relative to the aldehyde function and which is soluble in ethanol (for example 4 mmol or 0.5 molar equivalent) is dissolved beforehand in ethanol (for example in 20 mL) then is added to the ethanol solution containing the chitosan nanoparticles;
- the reaction mixture is stirred at ambient temperature (namely at approximately 20° C.) so as to obtain nanoparticles of chitosan functionalized with the benzaldehyde derivative.

The stirring time of the mixture can be between 30 minutes and 24 hours.

The functionalization of chitosan by wet process in ethanol is perfectly within the reach of those skilled in the art who will know how to implement the different reaction parameters.

At the end of the functionalization reaction, the ethanol containing the dispersed functionalized chitosan nanoparticles can be added to the liquid dispersion of semi-crystalline NMFC which has been described above.

Or else, in another embodiment of the invention, at the end of the functionalization reaction, the functionalized chitosan nanoparticles are centrifuged (for example for 5 minutes at a speed of 7000 rpm), then optionally dispersed in water (preferably a water bath subjected to ultrasound to clean them) the volume of which is adjusted to obtain the desired concentration of functionalized chitosan nanoparticles.

The functionalization of chitosan using a supercritical fluid can be carried out in the following way:
- Chitosan nanoparticles are reacted with a benzaldehyde derivative having a hydroxyl group in the ortho position relative to the aldehyde function and which is soluble in a supercritical fluid under the conditions of said supercritical fluid. Advantageously, the chitosan/benzaldehyde derivative molar ratio is 2/1. If the supercritical fluid is supercritical carbon dioxide, the conditions are: a pressure comprised between 200 bar and 300 bar, a temperature comprised between 80° C. and 130° C. and a reaction time comprised between 1 hour and 8 hours.
- Then, a slow decompression down to atmospheric pressure and ambient temperature of the reaction medium is carried out.
- Functionalized chitosan nanoparticles are recovered.

At the end of the functionalization by wet process or by using a supercritical fluid, the nanoparticles of functionalized chitosan can:
- be dispersed in an aqueous medium or ethanol, and this dispersion of functionalized chitosan nanoparticles is added to the liquid dispersion of semi-crystalline NMFC, or
- be directly added to the liquid dispersion of semi-crystalline NMFC.

The mass content of functionalized chitosan nanoparticles may be between 0.01% and 30%, preferably between 0.01% and 10%, more preferably between 0.01% and 5%, relative to the total mass of the liquid dispersion. As explained above, this liquid dispersion therefore contains the semi-crystalline NMFC and the functionalized chitosan nanoparticles. Depending on the application of the labeling method according to the invention referred to, it is entirely within the reach of those skilled in the art to choose the appropriate mass content of functionalized chitosan nanoparticles. Indeed, depending on the applications envisaged, it may be necessary for the mass content of functionalized chitosan nanoparticles to be high, for example of the order of 30%, whereas for other applications, a much lower mass content is required. , for example of the order of 0.01%.

The semi-crystalline NMFC labeled with the labeling method according to the invention can be detected by optical identification of the presence of functionalized chitosan by fluorescence measurement or by fluorescent revelation. The fluorescence measurement can be carried out by microscopy or spectroscopy.

This is why the invention also relates to a method for detecting semi-crystalline NMFC labeled according to the labeling method as described above in a sample to be analyzed, which is characterized in that it consists of:
- irradiate at an excitation wavelength corresponding to the maximum absorption of functionalized chitosan a sample to be analyzed;
- detecting the re-emitted light (corresponding to a maximum fluorescence intensity) at the emission wavelength of the functionalized chitosan;
- measuring or controlling said re-emitted light in order to identify the presence of said functionalized chitosan so as to conclude whether the sample to be analyzed contains said labeled semi-crystalline NMFC.

A person skilled in the art is able to adapt the means for detecting the labeled semi-crystalline NMFC obtained according to the labeling method of the invention, in particular with regard to the nature of the benzaldehyde derivative used to functionalize the chitosan.

Different light sources for the excitation of the functionalized chitosan can be used depending on the benzaldehyde derivative used for the functionalization, for example an ultraviolet lamp, such as a mercury vapor lamp emitting at a wavelength of 365 nm.

The device used for the fluorescence spectra can be, for example, a FLUOROLOG FL3-2iHR from the HORIBA brand.

The semi-crystalline NMFC labeled with the labeling method according to the invention can be used in a wide variety of applications.

One of these major applications is the use of semi-crystalline NMFC as a coating for a substrate, due to its oxygen and water barrier properties which have been mentioned above.

This is why the invention also relates to a substrate coated with at least one layer of semi-crystalline NMFC marked according to the marking method described above. The substrate can be an article from various technical fields, for example food, medicine or electronics.

Preferably, the substrate is based on cellulose. Indeed, as explained above, the labeling process according to the invention then presents all its interest because it is easy to distinguish the semi-crystalline labeled NMFC from the cellulose of the substrate. The substrate cellulose can be any type of cellulose that has been described above.

For example, the substrate can be a film or a paper.

In a preferred embodiment of the invention, the substrate is a film or a paper for packaging in the food industry or in the medical field.

The techniques for coating a labeled NMFC-semi-crystalline substrate are well within the reach of those skilled in the art. The fact that the semi-crystalline NMFC is labeled with the labeling method according to the invention has no impact on the choice of the coating technique. It may for example be stick coating (whose English name is "bar-coating"), screen printing, centrifugal coating (whose English name is "spin-coating"), inkjet or roll-to-roll printing. The coating technique by bar-coating is particularly preferred.

Due to its nanometric or micrometric size, the semi-crystalline NMFC marked according to the marking method according to the invention has a very good affinity for cellulose during the implementation of the coating on the substrate.

The thickness of the layer of semi-crystalline NMFC labeled according to the labeling process according to the invention can be between 1 μm and 500 μm, preferably between 10 μm and 100 μm, more preferentially between 50 μm and 100 μm.

Thus, to return to the detection method described above, the sample to be analyzed may be a sample of a substrate as described above, in particular a cellulose-based substrate, and for which it is desired to ensure that it is indeed coated with at least one layer of semi-crystalline NMFC marked according to the marking method according to the invention. The identification of the functionalized chitosan during the detection process makes it possible to conclude that the substrate is indeed coated with labeled semi-crystalline NMFC.

The invention will be better understood with the aid of the detailed description which is given below with reference to the appended drawing representing, by way of non-limiting example, emission and excitation spectra which have been obtained from labeled semi-crystalline NMFC according to the invention and unlabeled semi-crystalline NMFC.

There represents the emission spectra of different samples of labeled or unlabeled semi-crystalline NMFC, before or after abrasion.

There represents the excitation spectra of different samples of labeled or unlabeled semi-crystalline NMFC, before or after abrasion.

EXPERIMENTAL PART :

Functionalization of nanoparticles of chitosan :

Chitosan nanoparticles (CAS number 148411-57-8, average molar mass of 5000 g/mol provided by the company SIGMA ALDRICH) were functionalized by wet process according to the following protocol:
- 1.424 g (8 mmol) of chitosan nanoparticles were dispersed under strong magnetic stirring in 30 mL of ethanol;
- 0.776 g (4 mmol) of ethyl 3-formyl-4-hydroxybenzoate were dissolved beforehand in 20 mL of ethanol and then were added to the ethanol solution containing the chitosan nanoparticles;
- the reaction mixture was stirred at ambient temperature (namely at approximately 20° C.) so as to obtain nanoparticles of chitosan functionalized with ethyl 3-formyl-4-hydroxybenzoate dispersed in 50 mL of ethanol.

Labeling of semi-crystalline NMFC with the nanoparticles of chitosan functionalized:

20 mL of an aqueous dispersion of semi-crystalline NMFC whose mass content was 2% by mass relative to the mass of said aqueous dispersion were withdrawn and diluted in 30 mL of deionized water so as to obtain an aqueous dispersion of semi-crystalline NMFC whose mass content was 0 .8%.

0.45 mL of the dispersion of functionalized chitosan nanoparticles in ethanol was added with vigorous stirring (100 rpm) to the aqueous dispersion of semi-crystalline NMFC.

The dispersion thus obtained was kept under stirring (100 revolutions/minute) for 1 hour at ambient temperature so as to obtain a liquid dispersion containing, in mass percentages expressed relative to the total mass of said dispersion: 0.8% of NMFC semi -crystalline and 0.04% of functionalized chitosan nanoparticles and in which the semi-crystalline NMFC was labeled with the functionalized chitosan.

Coating a substrate with a semi-crystalline NMFC layer

The semi-crystalline labeled NMFC thus obtained was deposited by coating with a stick (whose English name is "bar-coating") on a cellulose substrate so as to obtain a coating of semi-crystalline labeled NMFC with a thickness of 20 μm. The deposition was carried out on aluminum TABER plates to then carry out the mechanical stresses. The substrate thus coated with labeled semi-crystalline NMFC was dried for one hour at room temperature (approximately 20° C.).

This substrate coated with labeled semi-crystalline NMFC constituted a ^first sample.

Furthermore, a 2 ^nd sample was prepared identically to the 1 ^st sample except that the semi-crystalline NMFC was not labelled.

Testing abrasion:

Abrasion tests were carried out on the 1 ^st and 2 ^nd samples in order to simulate real conditions of use (and therefore of wear) of the substrate coated with a layer of semi-crystalline NMFC.

These abrasion tests were carried out in accordance with NF EN ISO 7784-1 7784-2 standards with a TABER device. It was more precisely a Rotary TABER, Rotary Platform Abraser from the company TABER INDUSTRIES.

These tests were carried out in controlled environments (temperature of approximately 21°C +/- 3°C, ambient humidity, zero particulate background noise).

The 1 ^st and 2 ^nd samples were thus abraded so as to obtain respectively a 3 ^rd sample and a 4 ^th sample.

Specifically, the 3 ^rd sample contained the abraded labeled semi-crystalline NMFC fraction collected as a residual powder and the aerosolized labeled semi-crystalline NMFC fraction collected on polycarbonate membranes in a collection cassette.

The ^4th sample contained the abraded unlabeled semi-crystalline NMFC fraction collected as a residual powder and the aerosolized unlabeled semi-crystalline NMFC fraction collected on polycarbonate membranes in a collection cassette.

Samples 1 to 4 were irradiated at an excitation wavelength of 317 nm with a mercury vapor lamp and the variation of fluorescence intensity as a function of emission wavelength was analyzed. .

On the thus shown are the fluorescence emission spectra of samples 1 to 4 at this excitation wavelength of 317 nm.

It is noted that the fluorescence emission maxima agree between the samples of semi-crystalline NMFC (labeled or not) and the samples of semi-crystalline NMFC (labeled or not) abraded. Indeed, the emission maximum is located at an emission wavelength of 450 nm for samples n°2 and n°4 of unlabeled semi-crystalline NMFC and at an emission wavelength of 500 nm for samples #1 and #3 of labeled semi-crystalline NMFC.

In addition, it is noted that the labeling of the semi-crystalline NMFC according to the labeling method according to the invention, namely with a functionalized chitosan as described above, had the effect of shifting the emission maximum to a emission wavelength changing from 450 nm to 500 nm. This testifies to the effectiveness of the labeling process according to the invention for labeling the semi-crystalline NMFC and distinguishing it unambiguously from a cellulose-based substrate on which it can be deposited.

The differences in fluorescence emission intensity are explained by the difference in the amount of material present in the samples. Indeed, the quantities of samples n°3 and n°4 containing the semi-crystalline NMFC (labeled or not) abraded are less important than those of the samples n°1 and n°2 containing the semi-crystalline NMFC (labeled or No).

Next, the emission wavelength was set at 444 nm and the excitation wavelength was cycled through to establish the excitation spectra for each of samples 1 through 4.

On the Shown are the fluorescence excitation spectra of samples 1-4 at an emission wavelength of 444 nm.

As with the emission spectra, it is noted that the fluorescence excitation maxima agree between the samples of semi-crystalline NMFC (labeled or not) and the samples of semi-crystalline NMFC (labeled or not) abraded. Indeed, the excitation maximum is located at an excitation wavelength of 320 nm for samples n°2 and n°4 of unlabeled semi-crystalline NMFC and at an excitation wavelength of 370 nm for samples #1 and #3 of labeled semi-crystalline NMFC.

As for the emission spectra, it is noted that the labeling of the semi-crystalline NMFC according to the labeling method according to the invention had the effect of shifting the excitation maximum to an excitation wavelength from 320 nm to 370 nm. This testifies to the effectiveness of the labeling process according to the invention for labeling the semi-crystalline NMFC and distinguishing it unambiguously from a cellulose-based substrate on which it can be deposited.

As explained above for the emission spectra, the differences in fluorescence excitation intensity are explained by the difference in the amount of material present in the samples.

Claims (12)

Process for labeling semi-crystalline nano- and/or micro-fibrillated cellulose (hereinafter abbreviated "NMFC"), characterized in that it consists in adding to a liquid dispersion of semi-crystalline NMFC nanoparticles of chitosan which has been functionalized by reaction of its primary amine groups with a benzaldehyde derivative which has a hydroxyl group in the ortho position with respect to the aldehyde function. Labeling process according to Claim 1, characterized in that the semi-crystalline NMFC has fiber lengths of between 0.5 µm and 100 µm and a diameter of between 1 nm and 100 nm. Marking process according to claim 1 or 2, characterized in that the mass content of semi-crystalline NMFC in the liquid dispersion is between 0.1% and 99%, preferably between 0.5% and 10%, more preferably between 0.5% and 5%, relative to the total mass of the liquid dispersion. Labeling process according to any one of Claims 1 to 3, characterized in that the benzaldehyde derivative is chosen from salicylaldehyde, 2,4-dihydroxybenzaldehyde, 5-iodosalicylaldehyde, 3-bromo-5-chlorosalicylaldehyde, 2-hydroxy-5-nitrobenzaldehyde, 3-tert-butyl-2-hydroxybenzaldehyde, ethyl 3-formyl-4-hydroxybenzoate and 2-hydroxy-3-methoxybenzaldehyde. Labeling process according to any one of Claims 1 to 4, characterized in that the chitosan has been functionalised by the wet route in ethanol with a derivative of benzaldehyde soluble in ethanol or using a supercritical fluid with a derivative of benzaldehyde soluble in supercritical fluid. Labeling process according to any one of Claims 1 to 5, characterized in that the mass content of functionalized chitosan nanoparticles is between 0.01% and 30%, preferably between 0.01% and 10%, more preferably between 0.01% and 5%, relative to the total mass of the liquid dispersion. Substrate, characterized in that it is coated with at least one layer of semi-crystalline NMFC labeled according to the labeling method according to any one of Claims 1 to 6. Substrate according to Claim 7, characterized in that the substrate is based on cellulose. Substrate according to Claim 7 or 8, characterized in that the substrate is a film or a paper. Substrate according to any one of Claims 7 to 9, characterized in that the thickness of the layer of semi-crystalline NMFC marked according to the marking method according to any one of Claims 1 to 6 is between 1 µm and 500 μm, preferably between 10 μm and 100 μm, more preferably between 50 μm and 100 μm. Method for detecting semi-crystalline NMFC labeled according to the labeling method according to any one of Claims 1 to 6 in a sample to be analyzed, characterized in that it consists in:
- irradiate at an excitation wavelength corresponding to the maximum absorption of the functionalized chitosan the sample to be analyzed;
- detect the re-emitted light at the emission wavelength of the functionalized chitosan;
- measuring or controlling said re-emitted light in order to identify the presence of said functionalized chitosan so as to conclude whether the sample to be analyzed contains said labeled semi-crystalline NMFC. Detection method according to Claim 11, characterized in that the sample to be analyzed is a sample of a substrate according to any one of Claims 7 to 10.