Hydrogel-based decontamination of aqueous samples containing nanoparticles

Abstract

The present invention relates to the use of a supramolecular system in order to remove particles from a liquid medium containing same. According to the invention, the supramolecular system includes at least one molecule having a low molecular weight and/or an organic compound from living organisms, preferably from jellyfish, said compound being selected from among collagen, a polysaccharide, a proteoglycan or a mixture of two of said organic compounds, and said molecule having a low molecular weight and formula (I) as defined herein. The invention also relates to a method for removing particles from a liquid medium containing same. The invention is particularly suitable for use in water decontamination and biotechnology.

Claims

1. A process for subtracting nanoparticles from a liquid medium containing the same, comprising a step (a) of adding a supramolecular system to said liquid medium at a temperature of between 2 and 95 C., wherein the supramolecular system comprises: at least one low-molecular-weight molecule, wherein said low-molecular-weight molecule being of formula (I), said supramolecular system forming a gel on contact with the liquid medium, said gel capturing the nanoparticles contained in said liquid medium, and a step (b) of separating said liquid medium and said gel having captured said nanoparticles, wherein said low-molecular-weight molecule being of formula (I) below: ##STR00024## wherein: X represents an oxygen atom; B represents a natural or unnatural puric or pyrimidic base, optionally substituted with an R.sub.3 group as defined below; substituents L.sub.1 and L.sub.2: are identical or different and represent: (i) a hydrogen atom, (ii) a hydroxyl group, (iii) a heteroaryl group comprising 1 to 4 nitrogen atoms, unsubstituted or substituted with a saturated or unsaturated, linear or branched C.sub.2-C.sub.30 hydrocarbon-based chain, (iv) a group chosen from an oxycarbonyl group OC(O), a thiocarbamate group OC(S)NH, a carbonate group OC(O)O, a carbamate group OC(O)NH, an ether group O, a phosphate group or a phosphonate group, knowing that said L.sub.1 group is substituted with an R.sub.1 group and said L.sub.2 group is substituted with an R.sub.2 group, where R.sub.1 and R.sub.2, which may be identical or different, represent: a linear or branched, saturated or unsaturated, partially fluorinated or perfluorinated, C.sub.2-C.sub.30, hydrocarbon-based chain, a C.sub.2-C.sub.30 acyl radical, or an acylglycerol group, or form a ketal group of formula (II) below: ##STR00025## formula (II) in which K.sub.1 and K.sub.2 are identical or different and represent a saturated or unsaturated C.sub.1-C.sub.19 hydrocarbon-based chain, R.sub.3 and R.sub.3 represent, independently of one another: (i) a hydroxyl, amino, phosphate, phosphonate, phosphocholine, O-alkyl phosphocholine, thiophosphate or phosphonium group, (ii) a linear or branched C.sub.2-C.sub.30 alkyl chain optionally substituted with at least one hydroxyl group, (iii) a (CH.sub.2).sub.nVR.sub.8 group, wherein V represents an O, S or NH group, R.sub.8 represents a C.sub.2-C.sub.30 alkyl, and n is an integer from 1 to 50, (iv) a VC(O)R.sub.8 group, wherein V represents an O, S or NH group, and R.sub.8 represents a C.sub.2-C.sub.30 alkyl, or (v) a heteroaryl group containing from 1 to 4 nitrogen atoms, said heteroaryl group being unsubstituted or substituted with a C.sub.2-C.sub.30 alkyl, or with a (CH.sub.2).sub.mO(CH.sub.2).sub.pR.sub.9 group, or with a (CH.sub.2).sub.0-1YC(O)R group, or with a monosaccharide or polysaccharide, or with a group: ##STR00026## or with a group: ##STR00027## wherein: m is an integer from 1 to 6, p is an integer from 0 to 10 and R.sub.9 represents a C.sub.1 to C.sub.10 alkyl group, or a cyclic ketal group containing 5 to 7 carbon atoms, said cyclic ketal group being unsubstituted or substituted with at least one linear or branched C.sub.2-C.sub.30 alkyl, a sterol group, a diacyl glycerol, a hydrofluorocarbon-based chain or at least one monosaccharide or polysaccharide, Y is an oxygen atom, an NH group or a sulfur atom, and R is a hydrocarbon-based chain or a fluorocarbon-based chain, R is a hydrocarbon-based chain.

2. The process according to claim 1, also comprising, before step (b), when the temperature of the liquid medium obtained in step (a) is below 50 C., the following intermediate steps: (a1) heating the medium obtained in step (a) to a temperature of between 50 and 95 C., and (a2) cooling the medium obtained in step (a1) to a temperature of between 2 and 50 C.

3. The process according to claim 1, wherein the concentration of supramolecular system used is between 0.001 mg.Math.ml.sup.1 and 100 mg.Math.ml.sup.1 of aqueous medium.

4. The process according to claim 1, wherein the nanoparticles have a size of between 5 nm and 100 nm.

5. The process according to claim 1, wherein the liquid medium is chosen from the group comprising water, an organic solvent and a polyphase medium.

6. The process according to claim 1, wherein said low-molecular-weight molecule is an amphiphilic fluorinated glycosyl nucleoside.

7. The process according to claim 1, wherein X represents an oxygen atom, and/or B represents a thymine, adenine, guanine, cytosine, 6-methoxypurine or hypoxanthine or an unnatural puric or pyrimidic base, which can encompass the base: ##STR00028## wherein: R.sub.3 is as defined below, and/or L.sub.1 represents a hydroxyl group, L.sub.2 represents a hydrogen atom, or L.sub.1 and L.sub.2 together form a ketal group of formula (II) below: ##STR00029## wherein K.sub.1 and K.sub.2 are identical or different and represent a saturated or unsaturated C.sub.1-C.sub.19 hydrocarbon-based chain and/or R.sub.3 and R.sub.3 represent: (i) a hydroxyl, amino, phosphate, phosphonate, phosphocholine, O-alkyl phosphocholine, thiophosphate or phosphonium group, (ii) a linear or branched C.sub.2-C.sub.30 alkyl chain, optionally substituted with at least one hydroxyl group, (iii) a (CH.sub.2).sub.nVR.sub.8 group, wherein V represents an O, S or NH group, R.sub.8 represents a C.sub.2-C.sub.30 alkyl, and n is an integer from 1 to 50, (iv) a VC(O)R.sub.8 group, wherein V represents an O, S or NH group, and R.sub.8 represents a C.sub.2-C.sub.30 alkyl, or (v) a heteroaryl group containing from 1 to 4 nitrogen atoms, said heteroaryl group being unsubstituted or substituted with a C.sub.2-C.sub.30 alkyl, or with a (CH.sub.2).sub.mO(CH.sub.2).sub.pR.sub.9 group, or with a (CH.sub.2).sub.0-1YC(O)R group, or with a monosaccharide or polysaccharide, or with a group: ##STR00030## or with a group: ##STR00031## wherein: m is an integer from 1 to 6, p is an integer from 0 to 10 and R.sub.9 represents a C.sub.1 to C.sub.10 alkyl group, or a cyclic ketal group containing 5 to 7 carbon atoms, said cyclic ketal group being unsubstituted or substituted with at least one linear or branched C.sub.2-C.sub.30 alkyl, a sterol group, a diacyl glycerol, a hydrofluorocarbon-based chain or at least one monosaccharide or polysaccharide, Y is an oxygen atom, an NH group or a sulfur atom, and R is a hydrocarbon-based chain or a fluorocarbon-based chain, R is a hydrocarbon-based chain.

8. The process according to claim 1, wherein: X represents an oxygen atom, B represents an unnatural pyrimidic base substituted with a heteroaryl group containing three nitrogen atoms, said heteroaryl group being substituted with a group: ##STR00032## substituents L.sub.1 and L.sub.2 are identical or different and represent: (i) a hydrogen atom, (ii) a hydroxyl group, R.sub.3 represents a heteroaryl group comprising three nitrogen atoms, said heteroaryl group being substituted with a group: ##STR00033## wherein: R is a hydrocarbon-based chain; Y is an oxygen atom, an NH group or a sulfur atom, and R is a hydrocarbon-based chain or a fluorocarbon-based chain.

9. The process according to claim 1, wherein X represents an oxygen atom; B represents a puric or pyrimidic base; substituents L.sub.1 and L.sub.2 form a ketal group of formula (II) below: ##STR00034## formula (II) in which K.sub.1 and K.sub.2 are identical or different and represent a saturated or unsaturated C.sub.1-C.sub.19 hydrocarbon-based chain, R.sub.3 represents a phosphocholine group.

10. The process according to claim 1, wherein said low-molecular-weight molecule is chosen from: the group defined by formula (III) below, wherein R is a hydrocarbon-based chain: ##STR00035## the group defined by formula (IV) below, wherein Y is an oxygen atom, an NH group or a sulfur atom, and R is a hydrocarbon-based chain or a fluorocarbon-based chain: ##STR00036## the group defined by formula (V) below, wherein n is an integer between 0 and 19: ##STR00037## or the group defined by formula (VI) below, wherein n is an integer between 0 and 19: ##STR00038##

11. The process according to claim 1, wherein said low-molecular-weight molecule is chosen from the group comprising: 5-(4-((2H,2H,3H,3H-perfluoroundecanamide)methyl)-1H-1,2,3-triazol-1-yl)-N3-(1-((-D-glucopyranoside)-1H-1,2,3,-triazol-4-yl)methyl)thymidine, 5-(4-((1H, 1H,2H, 2H-perfluoroundecanamide)methyl)-1H-1,2,3-triazol-1-yl)-N3-(1-((-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methyl)thymidine, 5-(4-((oleamide)methyl)-1H-1,2,3-triazol-1-yl)-N3-(1-((-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methyl)thymidine, 5-(4-((stearamide)methyl)-1H-1,2,3-triazol-1-yl)-N3-(1-((-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methyl)thymidine, 5-(4-((octadecyloxy)methyl)-1H-1,2,3-triazol-1-yl)-N3-(1-((-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methyl)thymidine, and 5-(4-((cholesteryloxy)methyl)-1H-1,2,3-triazol-1-yl)-N3-(1-((-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methyl)thymidine, 2,3-O-18-pentatriacontanylidenuridine-5-phosphocholine, 2,3-O-18-pentatriacontanylidenadenosine-5-phosphocholine, and a mixture of at least two of these compounds.

Description

BRIEF DESCRIPTION OF THE FIGURES

(1) FIG. 1 represents a microtube comprising a hydrogel and a supernatant of a solution of encapsulated QDs. Represented in 1A is the microtube in the normal position. Represented in 1B is the same microtube as 1A in the upside down position. Represented in 1C is the hydrogel and the supernatant without UV. Represented in 1D is the same microtube as 1C under UV at .sub.max=312 nm. SU means supernatant, RF means that a red fluorescence is observed, and NF means that no fluorescence is observed.

(2) FIG. 2 represents a fluorescence spectrum of a solution of encapsulated QDs in water (a) and of the supernatant after the formation of the hydrogel according to the protocol described in this example (b). Represented along the x-axis is the wavelength in nanometers (NM) and represented along the y-axis is the intensity in arbitrary units (I).

(3) FIG. 3 represents photos taken under a microscope of FGNs without (3A) or with (3B) encapsulated QDs.

(4) FIG. 4 represents a microtube (a) of a solution of AuNPs covered with L-lysine in water, (b) of AuNPs covered with L-lysine 48 hours after the formation of the hydrogel of FGNs and (c) of the colorless liquid supernatant (I) and of the gel of FGNs and of AuNPs covered with L-lysine of ruby red color (II).

EXAMPLES

(5) The purpose of the experiments presented below is to demonstrate that the supramolecular system used according to the invention makes it possible to decontaminate a liquid medium contaminated with particles.

Example 1: Decontamination of a Liquid Medium Containing QDs

(6) In this example, FGNs (amphiphilic fluorinated glycosyl nucleosides) were used as supramolecular system comprising low-molecular-weight molecules for decontaminating a liquid medium comprising encapsulated quantum dots (QDs).

(7) Material

(8) Amphiphilic Fluorinated Glycosyl Nucleosides (FGNs):

(9) 5-(4-))2H,2H,3H,3H-Perfluoroundecanamide)methyl)-1H-1,2,3-triazol-1-yl)N3-)1-((b-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methylthymidine was synthesized according to the protocol described in the document Godeau et al. (Godeau et al., Tetrahedron Lett., 2010, vol. 51, p. 1012-1015 [2]).

(10) Quantum Dots (QDs):

(11) The QDs were obtained from the company Evident Technologies (catalog reference: ED-C11-TOL-0620). These QDs are at a concentration of 10 mg.Math.ml.sup.1 in toluene (24.4 nM). Their core is composed of CdSe and their shell of ZnS. These QDs are encapsulated in trioctylphosphine oxide (TOPO).

(12) The QDs have unique optical properties. Indeed, QDs of the same material, but having different sizes, can emit light of various colors. The physical reason is the quantum confinement effect.

(13) DOPC:

(14) The (2-{[(2S)-2,3-bis[(9Z)-octadec-9-enoyloxy]propylphosphonato]oxy}ethyl)trimethylazanium) was obtained from the company Genzyme Pharmaceuticals (catalog reference: LP-R4-070).

(15) DOTAU:

(16) The (N-[5-(2,3-dioleoyl)uridine]-N,N,N-trimethylammonium tosylate) was synthesized according to the protocol described in the document Chabaud et al. (Chabaud et al., Bioconjugate Chem., 2006, vol. 17, p. 466-472 [3]).

(17) Process for Encapsulating QDs in DOTAU/DOPC Micelles

(18) 200 l of QDs encapsulated in TOPO (10 mg.Math.ml.sup.1 in toluene) and 100 ml of chloroform were poured into an RB flask. The solvents were evaporated off in a rotary evaporator at 30 C. and then the QDs were dried under a high vacuum using a vacuum pump for 3 to 4 hours.

(19) The QDs were then suspended in 35 ml of chloroform with 5 ml of phospholipids (2.5 ml DOTU and 2.5 ml DOPC, at a concentration of 10 mg.Math.ml.sup.1 in chloroform).

(20) 400 l of the solution obtained were then distributed into 100 test tubes (400 l per tube). After complete evaporation of the chloroform from each test tube, the residues were heated to 80 C. and 1 ml of water was added to 50 of these test tubes in order to obtain a clear suspension containing DOTU/DOPC micelles. The content of each of these 50 test tubes was transferred into another of the other 50 test tubes in order to obtain an optically clear suspension containing DOTU/DOPC micelles. This transfer was carried out so as not to have a total volume greater than 50 ml.

(21) The phospholipid-encapsulated QDs were then transferred into 2000 l microtubes. Any aggregates of QDs encapsulated in the phospholipids were removed by centrifugation at 40000 rpm at 20-25 C. for twice 15 minutes.

(22) In order to separate the QD solution from the residual aggregates, the supernatant was collected and transferred into a Vivaspin (cut-off threshold Mw of 30 kDa).

(23) The total volume of the QD solution was reduced by 20 to 25 ml by centrifugation at 4000 rpm at 20-25 C. for 15 minutes. All the QD solution was then harvested in a flask and stored in the dark at 4 C.

(24) The size of the encapsulated QDs obtained was less than or equal to 20 nm (measured on a Zetasizer apparatus by means of DLS experiments) (Binil Itty Ipe et al., ChemPhysChem, 2006, 7, 1112-1118 [4]).

(25) Their concentration was 17 microg.Math.ml.sup.1 in water (measured by fluorescence spectroscopy). The fluorescence spectra were recorded on an LS 55 spectrofluorimeter (Perkin Elmer) equipped with a xenon flashlamp. The data processing was carried out with the SigmaPlot 11 program.

(26) Experimental Section: Formation of a Gel for Decontaminating a Solution Containing QDs

(27) 0.1 mg of FGNs in solution at 0.1% (i.e. 1.0 mg.Math.ml.sup.1) was mixed with 1 ml of QDs in solution in water at a concentration of 17 g.Math.ml.sup.1 in a 2 ml microtube, at 20-25 C.

(28) A gel was prepared by heating the solution to 80 C. in a water bath with constant agitation until a visually clear solution was obtained. The agitation time in this experiment was 3 to 4 minutes. The solution was then left in the dark for 48 hours in order for it to be able to stabilize.

(29) After 48 hours, a gel was formed with a liquid supernatant. The presence of the gel and of the supernatant was confirmed by turning the microtube upside down: the gel is maintained in the bottom of the tube, while the supernatant flows (see FIGS. 1A and 1B).

(30) The liquid supernatant was separated from the gel. An observation of the liquid supernatant under UV (.sub.max=312 nm) then confirmed that the encapsulated QDs were all captured by the gel. Indeed, a red fluorescence was observed for the gel, whereas no fluorescence was observed for the supernatant (FIGS. 1C and 1D).

(31) Furthermore, FIG. 2 shows the fluorescence spectrum of a solution of encapsulated QDs in water (a) and of the supernatant after the formation of the gel according to the protocol described in this example (b). The capture of the encapsulated QDs by the FGN used was observed under a microscope (FIGS. 3A and 3B).

(32) Conclusion

(33) The FGNs (amphiphilic fluorinated glycosyl nucleosides) make it possible to form a gel on contact with the liquid medium which makes it possible to decontaminate a liquid medium contaminated with particles having a size of less than 20 nm (encapsulated QDs).

Example 2: Decontamination of a Liquid Medium Containing Gold Nanoparticles

(34) In this example, FGNs (amphiphilic fluorinated glycosyl nucleosides) were used as supramolecular system comprising low-molecular-weight molecules for decontaminating a liquid medium comprising gold nanoparticles (AuNPs).

(35) Material

(36) Amphiphilic Fluorinated Glycosyl Nucleosides (FGNs):

(37) 5-(4-))2H,2H,3H,3H-perfluoroundecanamide)methyl)-1H-1,2,3-triazol-1-yl)N3-)1-((b-D-glucopyranoside)-1H-1,2,3-triazol-4-yl)methyl)thymidine was synthesized according to the protocol described in the document Godeau et al. (Godeau et al., Tetrahedron Lett., 2010, vol. 51, p. 1012-1015 [2]).

(38) Chloroauric acid (HAuCl.sub.4):

(39) The chloroauric acid was obtained from the company Alfa Aesar (ref No. 36400).

(40) Process for Preparing Gold Nanoparticles Covered with L-Lysine

(41) This process was carried out according to the protocol described in Selvakannan et al. (Selvakannan et al., Langmuir, 2003, vol. 19, p. 3545-3549 [5]).

(42) 100 ml of aqueous solution of chloroauric acid (HAuCl.sub.4) at a concentration of 10.sup.4 M were reduced by adding 0.01 g of sodium borohydride (NaBH.sub.4) at 20-25 C. so as to produce colloidal gold particles. This procedure results in a change of color of the solution from pale yellow to ruby red. This indicates the formation of gold nanoparticles.

(43) The colloidal gold particles were covered by adding 10 ml of an aqueous solution of lysine at 10.sup.3 M to 90 ml of colloidal gold particle solution. The mixture is put aside overnight. This will provide the L-lysine with water-soluble gold-capped nanoparticles. The solution obtained was left to stand overnight in order to form the L-lysine-covered AuNPs.

(44) The mean diameter of the AuNPs not covered with L-lysine was 6.5 nm0.7 nm.

(45) The concentration of the L-lysine-covered AuNPs was 10.sup.4 M.

(46) Experimental Section: Formation of a Gel for Decontaminating a Solution Containing L-Lysine-Covered AuNPs

(47) 0.1 mg of FGNs in solution at 0.1% (i.e. 1.0 mg.Math.ml.sup.1) was mixed with 2 ml of L-lysine-covered AuNPs in solution in water at a concentration of 10.sup.4 M in a 2 ml microtube, at 20-25 C.

(48) A gel was prepared by heating the solution to 80 C. in a water bath with constant agitation until a visually clear solution was obtained. The agitation time in this experiment was 3-4 minutes. The solution was then left in the dark for 48 hours in order for it to be able to stabilize.

(49) After 48 hours, a gel was formed with a liquid supernatant. The presence of the gel and of the supernatant was confirmed by turning the microtube upside down: the gel is maintained in the bottom of the tube, while the supernatant flows in the same way as in example 1.

(50) The liquid supernatant was separated from the gel. A ruby red color was observed in the gel, whereas the supernatant was colorless, thereby confirming that all the L-lysine-covered AuNPs were captured in the FGN gel.

(51) Furthermore, FIG. 4 shows the fluorescence spectrum (a) of a solution of L-lysine-covered AuNPs in water, (b) of the L-lysine-covered AuNPs 48 hours after the formation of the FGN gel, and (c) of the colorless liquid supernatant (I) and of the ruby red-colored gel of FGNs and of L-lysine covered AuNPs.

(52) Conclusion

(53) The FGNs (amphiphilic fluorinated glycosyl nucleosides) make it possible to form a gel on contact with the liquid medium which makes it possible to decontaminate a liquid medium contaminated with L-lysine-covered gold particles having a size of less than approximately 6.5 nm.

Example 3: Decontamination of a Liquid Medium Containing QDs

(54) In this example, natural polymers were used as supramolecular system comprising a polymer for decontaminating a liquid medium comprising encapsulated quantum dots (QDs).

(55) Material

(56) Jellyfish:

(57) Mnemiopsis Leidyi ctenophore jellyfish (Agassiz, 1865) weighing 0.5 gram and 1 centimeter in diameter were harvested from the Etang de Berre [Berre lake] (France).

(58) These jellyfish consist of glycosaminoglycans (GAGs) and/or of mucopolysaccharides.

(59) Scyphozoa Cnidaria:

(60) Scyphozoa Cnidaria, Aurelia, weighing 200 g and 10 centimeters in diameter were harvested from the Etang de Berre [Berre lake] (France).

(61) Encapsulated (DOPC/DOTU) Quantum Dots (QDs):

(62) The encapsulated QDs were obtained in the same way as in example 1 above.

(63) The size of the encapsulated QDs obtained is less than or equal to 20 nm (measured on a Zetasizer apparatus by means of DLS experiments) (Binil Itty Ipe et al. [4]).

(64) Their concentration was 17 microg.Math.ml.sup.1 in water (measured by fluorescence spectroscopy). The fluorescence spectra were recorded on an LS 55 spectrofluorimeter (Perkin Elmer) equipped with a xenon flashlamp. The data processing was carried out with the SigmaPlot 11 program.

(65) Experimental Section: Formation of a Gel for Decontaminating a Solution Containing Encapsulated QDs

(66) About ten Mnemiopsis jellyfish were brought into contact with 1 ml of QDs in solution in water at a concentration of 17 g.Math.ml.sup.1 in a 5 ml glass tube, at 20-25 C.

(67) The mixture was moderately agitated for a few seconds and left to stand for 2 to 3 days.

(68) The glass tube containing the above reaction mixture was examined under UV light immediately after mixing. It was observed that, under UV, the entire reaction mixture was fluorescent red, with the exception of the jellyfish portions. The jellyfish portion appeared as black marks and no fluorescence was observed. This means that the encapsulated QDs were absorbed only on the surface of the jellyfish and are not capable of penetrating into the jellyfish.

(69) After 24 hours, the same observations as above were observed. The jellyfish were still alive.

(70) After 48 hours, all the jellyfish were dead and divided into small soft pieces. The entire solution was fluorescent red under UV.

(71) After 72 hours, the entire solution became transparent with some soft pieces precipitated at the bottom of the glass tube or stuck to the surface of the tube. The liquid supernatant was carefully removed and was observed under UV. Surprisingly, no fluorescence was observed. Virtually all the encapsulated QDs were absorbed by the dead jellyfish. When this reaction mixture is filtered through a strip of Whatman paper under gravity, a filtrate with no QD is obtained.

(72) Experiments on Aurelia aurita Scyphozoa Cnidaria (Linnaueus, 1758) weighing 200 g and 10 centimeters in diameter also give the same results.