DEVICE FOR MAINTAINING METAL HOMEOSTASIS, AND USES THEREOF

Abstract

The present invention relates to the field of medical devices, more particularly to devices for extracting metals from an organism. The use of these devices makes it possible, for example, to prevent and/or treat pathologies linked to dysregulation of metal homeostasis in the organism, for example neurological diseases.

Claims

1. A device for maintaining metal homeostasis for therapeutic purposes, characterized in that it comprises a means for extracting metal cations, said means being in particular selected from: an implant comprising at least one chelator, or a perfusion fluid containing at least one chelator, said perfusion fluid being contained in a dialysis system.

2. The device for maintaining the metal homeostasis as claimed in claim 1, characterized in that the chelator is capable of complexing metal cations, and characterized in that the complexing constant log(K.sub.Cl) of said chelator for at least one of said metal cations is greater than 10, and preferably greater than or equal to 15, and in particular said cations are selected from cations of the metals Cu, Fe, Zn, Hg, Cd, Pb, Mn, Mg, Ca, Gd and Al, and more particularly Cu, Fe and Zn.

3. The device for maintaining the metal homeostasis as claimed in claim 1, characterized in that it contains trace elements selected from Calcium, Magnesium, Iron, Copper, Zinc, or Manganese.

4. The device for maintaining metal homeostasis as claimed in claim 1, characterized in that said means makes it possible to extract metal cations from a biological fluid, an organ or a tissue, in particular when the content of said metal cations is less than 1 ppm, in particular 0.1 ppm, 0.01 ppm and is preferably less than 1 ppb.

5. The device for maintaining metal homeostasis as claimed in claim 1, characterized in that said means makes it possible to extract an amount of metal cations representing at least 1% of its mass, and preferably more than 10% of its mass.

6. The device for maintaining metal homeostasis as claimed in claim 1, characterized in that it comprises a dialysis system comprising: a. a porous dialysis membrane, and b. a reservoir containing perfusion fluid, and in that the perfusion fluid is selected from: a colloidal suspension of nanoparticles having a mean diameter greater than the pores of said porous dialysis membrane, said nanoparticles comprising as active principle at least one chelator, or a colloidal suspension of polymers whose mean diameter is greater than the pores of said porous dialysis membrane, said polymers being grafted to an active principle which is at least one chelator, a solution of chelating molecules.

7. The device for maintaining metal homeostasis as claimed in claim 6, characterized in that the colloidal suspension contains more than 1% by mass of nanoparticles or polymers, preferably more than 10% by mass.

8. The device for maintaining metal homeostasis as claimed in any claim 6, characterized in that said nanoparticles are polysiloxane-based nanoparticles having a mean diameter greater than 3 nm, preferably less than 50 nm.

9. The device for maintaining metal homeostasis as claimed in claim 6, characterized in that said nanoparticles comprise: a. polysiloxanes, with a silicon mass ratio of at least 8% of the total mass of the nanoparticle, preferably between 8% and 50% of the total mass of the nanoparticle, b. chelators, preferably in a proportion between 5 and 1000, and preferably between 5 and 100 per nanoparticle, c. if need be, metallic elements, for example in a proportion between 5 and 100, preferably between 5 and 20 per nanoparticle, said metallic elements being complexed to the chelators.

10. The device for maintaining metal homeostasis as claimed in claim 6, characterized in that said nanoparticles are of the following formula (I):
Si.sub.n [O].sub.m[OH].sub.o [Ch.sub.1].sub.a [Ch.sub.2].sub.b [Ch.sub.3].sub.c [M.sup.y+].sub.d[D.sup.2+].sub.e [Gf].sub.f(I) wherein: n is between 20 and 50,000 preferably between 50 and 1000. m is greater than n and less than 4 n o is between 0 and 2 n Ch.sub.1, Ch.sub.2 and Ch.sub.3 are chelators, identical or different, linked to the Si of the polysiloxanes by a covalent SiC bond; a, b and c are integers between 0 and n and a+b+c is less than or equal to n, preferably a+b+c is between 5 and 100, for example between 5 and 20, M.sup.y+ and D.sup.2+ are metal cations, identical or different, with y and z=1 to 6; d and e are integers between 0 and a+b+c, and d+e is less than or equal to a+b+c, Gf are targeting grafts, identical or different, each linked to Si by an SiC bond and derived from the grafting of a targeting molecule allowing the targeting of nanoparticles to biological tissues of interest, for example to tumor tissues, f is an integer between 0 and n.

11. The device for maintaining metal homeostasis as claimed in claim 6, characterized in that the chelators are obtained by grafting onto the nanoparticles or onto the polymer one of the following complexant molecules or derivatives thereof: DOTA, DTPA, EDTA, EGTA, BAPTA, NOTA, DOTAGA, DFO, DOTAM, NOTAM, DOTP, NOTP, TETA, TETAM, TETP and DTPABA, or mixtures thereof.

12. The device for maintaining metal homeostasis as claimed in claim 6, characterized in that said nanoparticles are polysiloxane-based nanoparticles with a mean diameter between 3 and 50 nm, comprising the chelator obtained by grafting DOTA, DOTAGA or DTPA onto the nanoparticles.

13. The device for maintaining metal homeostasis as claimed in claim 6, characterized in that said nanoparticles are polysiloxane-based nanoparticles with a mean size greater than 20 kDa and less than 1 MDa, comprising the chelator obtained by grafting DOTA, DOTAGA or DTPA onto the nanoparticles.

14. The device for maintaining metal homeostasis as claimed in claim 1, characterized in that said device comprises means allowing it to be brought into contact through a dialysis membrane or to be implanted within: a biological fluid, such as blood, cerebrospinal fluid, synovial fluid or peritoneal fluid, or an organ, such as the brain, liver, pancreas, intestines or lungs, or a tissue, such as the peritoneum or tumor tissue.

15. A colloidal suspension of nanoparticles comprising an active principle or of polymers grafted to an active principle, for use for therapeutic purposes, characterized in that it is contained in a device for maintaining metal homeostasis comprising a porous dialysis membrane, and in that the mean diameter of said nanoparticles or of said grafted polymers is greater than the pores of the porous dialysis membrane of said device.

16. A polysiloxane-based nanoparticle having a diameter greater than 3 nm, preferably less than 50 nm, for use for therapeutic purposes in a device for maintaining metal homeostasis, said nanoparticle comprising as active principle at least one chelator capable of complexing said metal cations, and characterized in that its complexing constant log(K.sub.Cl) for at least one of said metal cations is greater than 10, and preferably greater than or equal to 15.

17. A polymer, for use for therapeutic purposes in a device for maintaining metal homeostasis, said polymer being grafted to at least one chelator capable of complexing said metal cations, and characterized in that its complexing constant log(K.sub.Cl) for at least one of said metal cations is greater than 10, and preferably greater than or equal to 15.

18. The nanoparticle or colloidal suspension or polymer for use as claimed in claim 15, for use: in maintaining metal homeostasis, or in the treatment of neurological diseases or brain degeneration, such as Parkinson's disease, Alzheimer's disease, NBIA, Wilson's disease, or Huntington's disease, or in the treatment of autism, or in the treatment of type II diabetes or cardiovascular disease, or in the treatment of tumors.

Description

FIGURES

[0126] FIG. 1 shows the image obtained at the end of the perfusion of the MnCl.sub.2 solution. This is a coronal section at the microdialysis membrane (black dot). The highlight around the membrane corresponds to the presence of Mn.sup.2+ (positive MRI contrast agent).

[0127] FIG. 2 shows the image obtained at the end of the perfusion with the nanoparticle suspension. This is a coronal section at the microdialysis membrane (black dot). The highlight around the membrane corresponds to the presence of Mn.sup.2+ (positive MRI contrast agent).

[0128] FIG. 3 shows the image corresponding to the difference between the two previous images (shown in FIGS. 1 & 2) and highlighting the decrease in tissue concentration in Mn.sup.2+ (highlighted at the microdialysis probe).

[0129] FIG. 4 shows the image obtained at the end of the perfusion with the MnCl.sub.2 solution. This is a coronal section at the microdialysis membrane (black dot). The highlight around the membrane corresponds to the presence of Mn.sup.2+ (positive MRI contrast agent).

[0130] FIG. 5 shows the image obtained at the end of the perfusion with saline. This is a coronal section at the microdialysis membrane (black dot). The highlight around the membrane corresponds to the presence of Mn.sup.2+ (positive MRI contrast agent).

[0131] FIG. 6 shows the image corresponding to the difference between the two previous images (shown in FIGS. 4 & 5) and highlighting the absence of decrease in tissue concentration in Mn.sup.2+ (almost no highlighting at the microdialysis probe).

[0132] FIG. 7 shows the MRI image of solutions 1, 2, 3, 4 and 5.

[0133] FIG. 8: Hydrodynamic diameter of the nanoparticles obtained in Example 7.

[0134] FIG. 9: Hydrodynamic diameter of the nanoparticles obtained in Example 8.

EXAMPLES

Example 1: Extraction of Manganese Ions from Rodent Brain

[0135] The study was conducted on male Wistar rats (weight: 250 g).

[0136] On Day 0, for the insertion of the microdialysis cannula, the animal is placed under gas anesthesia (2.5% isoflurane under O.sub.2/N.sub.2 (80:20)) with the use of a heating mat used during the procedure and the recovery phase. Prior to incision of the skin to clear the skull, local anesthesia with subcutaneous injection of lidocaine (Xylovet 21.33 mg/mL) is performed (4 mg/kg diluted in 0.9% NaCl with an injected volume of 10 L/g). After incision of the skin, the skull is cleared in order to position a micro drill (diameter <1 mm) for skull piercing. The insertion of the probe is done under stereotaxy. The dialysis cannula (diameter <500 m) is gently inserted into the brain at the desired position and depth. After positioning the cannula, a fast-setting fixing resin is applied and screwed onto the animal's skull. The skin is then sewn back together to close the wound. Before the animal wakes up, an analgesic (Buprecare) is administered subcutaneously. Administration of the analgesic is repeated at intervals of 8 to 12 hours for 2 days following the insertion of the microdialysis probe. In order to limit dehydration of the animal, a subcutaneous injection of 0.9% NaCl (about 0.5 mL for mice, 5 mL for rats) is carried out at the beginning of the procedure. To prevent dry eye, an ophthalmological ointment (Liposic) is applied at the beginning of the procedure.

[0137] The MRI spectroscopy and imaging protocol is performed on Day 3. The protocol is performed on animals under gas anesthesia (2.5% isoflurane under O.sub.2/N.sub.2 (80:20)) with the use of a heating mat used during the procedure and the recovery phase and with breath control during NMR acquisitions. Before positioning the animal in the MRI (Bruker Biospin 4.7 Tesla), the microdialysis probe (2 mm long membrane, 6 kDa cut-off, CMA Microdialysis AB, Kista, Sweden) is inserted into the microdialysis cannula. An MRI surface antenna (Doty Scientific, 8 mm diameter, used for transmission and reception, is positioned on the skull of the animal vertically to the microdialysis probe. The MRI acquisitions (T1-weighted Flash sequence, echo time 2 ms, repetition time 150 ms, coronal sections, section thickness 1 mm, acquisition time 3 minutes) are performed continuously during the perfusion of the microdialysis probe.

[0138] Results

Example 1A

[0139] The microdialysis probe is perfused with a 1 mM MnCl.sub.2 solution in saline at a flow rate of 10 L/min for 30 minutes. The microdialysis probe is then perfused with a suspension of polysiloxane nanoparticles with free DOTAGA on their surface (28.1 mg diluted in 1 mL saline+100 L of NaOH and HCl to equilibrium at pH 7=or 28.1 mg in total volume of 1100 L) at a flow rate of 10 L/min for 30 minutes. The polysiloxane nanoparticles used consist of a polysiloxane matrix to which are grafted cyclic chelators of DOTAGA. These nanoparticles have a hydrodynamic diameter of 11.56.3 nm. This size prevents their passage through the dialysis membrane, whose pore diameter is 2 to 3 nm.

[0140] The image obtained at the end of the perfusion of the MnCl.sub.2 solution is shown in FIG. 1, and the image obtained at the end of the perfusion with the nanoparticle suspension is shown in FIG. 2. FIG. 3 shows the image corresponding to the difference between the two previous images and highlighting the decrease in tissue Mn.sup.2+ concentration (highlighted at the microdialysis probe).

Example 1B

[0141] The microdialysis probe is perfused with a 1 mM MnCl.sub.2 solution in saline at a flow rate of 10 L/min for 30 minutes. The microdialysis probe is then perfused with saline at 10 L/min for 30 minutes. The image obtained at the end of perfusion of the MnCl.sub.2 solution is shown in FIG. 4, and the image obtained at the end of perfusion with saline is shown in FIG. 5. FIG. 6 shows the image corresponding to the difference between the two previous images and showing the absence of a decrease in tissue Mn.sup.2+ concentration (almost no highlighting at the microdialysis probe).

CONCLUSIONS

[0142] MRI allows the objectification of tissue concentration variations in Mn.sup.2+ cation (paramagnetic MRI contrast agent). The presence of chelating nanoparticles in the perfusate results in a significant decrease in intensity in MRI sections due to a decrease in local tissue Mn.sup.2+ concentration. This decrease in intensity is not observed in the absence of chelating nanoparticles.

Example 2: Extraction of Intra-Tissue Gadolinium by Perfusion of Nanoparticle Solution

[0143] The study was conducted on male Wistar rats (weight: 250 g).

[0144] On Day 0, for the insertion of the microdialysis cannula, the animal is placed under gas anesthesia (2.5% isoflurane under O.sub.2/N.sub.2 (80:20)) with the use of a heating mat used during the procedure and the recovery phase. Prior to incision of the skin to clear the skull, local anesthesia with subcutaneous injection of lidocaine (Xylovet 21.33 mg/mL) is performed (4 mg/kg diluted in 0.9% NaCl with an injected volume of 10 L/g). After incision of the skin, the skull is cleared in order to position a micro drill (diameter <1 mm) for the drilling of the skull. The insertion of the probe is done under stereotaxy. The dialysis cannula (diameter <500 m) is gently inserted into the brain at the desired position and depth. After positioning the cannula, a fast-setting fixing resin is applied and screwed onto the animal's skull. The skin is then sewn back together to close the wound. Before the animal wakes up, an analgesic (Buprecare) is administered subcutaneously. Administration of the analgesic is repeated at intervals of 8 to 12 hours for 2 days following the insertion of the microdialysis probe. In order to limit dehydration of the animal, a subcutaneous injection of 0.9% NaCl (about 0.5 mL for mice, 5 mL for rats) is carried out at the beginning of the procedure. To prevent dry eye, an ophthalmological ointment (Liposic) is applied at the beginning of the procedure.

[0145] The microdialysis perfusion protocol is performed on Day 3. The protocol is performed on animals under gas anesthesia (2.5% isoflurane under O.sub.2/N.sub.2 (80:20)) with the use of a heating mat used during the procedure and the recovery phase and with control of the respiratory frequency. The microdialysis probe (2 mm long membrane, 6 kDa cut-off, CMA Microdialysis AB, Kista, Sweden) is inserted into the microdialysis cannula and the perfusion is performed at a flow rate of 10 L/min.

[0146] Perfusion is carried out over 30 minutes with a perfusate consisting of saline supplemented with 1 mM GdCl3 (solution 1). The dialysate is collected at the end of microdialysis (solution 2).

[0147] The microdialysis probe is then perfused with a nanoparticle suspension VL29-5 (28.1 mg diluted in 1 mL saline+100 L NaOH and HCl to equilibrate at pH 7=or 28.1 mg in a total volume of 1100 L) for 30 minutes (solution 3). The dialysate is collected at the end of microdialysis (solution 4). The nanoparticles used are identical to those in Example 1, i.e. they have a hydrodynamic diameter of 11.56.3 nm. This size prevents their passage through the dialysis membrane, which has a pore diameter of 2 to 3 nm.

[0148] These 4 solutions (as well as a saline solution 5) are imaged in a 4.7 Tesla MRI with a T1-weighted gradient echo sequence (repetition time 40 ms, echo time 2.6 ms, tilt angle 80).

[0149] The images of the 5 tubes are shown in FIG. 7.

[0150] Results

[0151] The results in FIG. 7 thus highlight the increase in intensity of solution 4 compared to solution 5, which clearly shows the uptake and chelation of tissue Gd during the passage of the nanoparticle solution through the microdialysis probe.

Example 3: Synthesis of Chitosane-DTPA-BA

[0152] The chitosan used has a mean molecular weight of 200 kDa. DTPA-BA (diethylenetriaminepentaacetic dianhydride) was supplied by Chematech, Dijon, France and used as such. The VIVAFLOW cassettes were purchased from Sartorius and used as is. The perfusion fluid was purchased from Phymep (Perfusion Fluid CNS Sterile, item number P000151) and used as is.

[0153] A mass of 0.5 g of chitosan was weighed and inserted into a 500 mL container. A volume of 250 mL of distilled water was added and the solution was stirred. Using a pH meter and a 50% acetic acid solution, the pH was adjusted to 4.00.1. The solution was stirred for 24 h. At 24 h the pH was again adjusted to 4.00.1. This procedure was repeated until all the chitosan was completely dissolved. A mass of 5.36 g of DTPA-BA was weighed and added to the resulting solution. The solution was stirred for 48 h. At 48 h the solution was purified using a Vivaflow cassette with a 100 kDa cut-off until a purification rate of at least 100,000 was achieved. Again using a Vivaflow cassette, the solvent is replaced by the CNS perfusion fluid at the same concentration.

Example 4: Synthesis of Chitosan-DFO

[0154] The chitosan used has a mean molecular weight of 200 kDa. p-NCS-Bz-DFO (N1-hydroxy-N1-(5-(4-(hydroxy(5-(3-(4-isothiocyanatophenyl)thioureido)pentyl)amino)-4-oxobutanamido)pentyl)-N4-(5-(N-hydroxyacetamido)pentyl)succinamide) was purchased from Chematech Mdt and used as is. VIVAFLOW cassettes were purchased from Sartorius and used as is. The perfusion fluid was purchased from Phymep (Perfusion Fluid CNS Sterile, item number P000151) and used as is.

[0155] A mass of 0.5 g of chitosan was weighed and placed in a 500 mL container. A volume of 250 mL of distilled water was added and the solution was stirred. Using a pH meter and a 50% acetic acid solution, the pH was adjusted to 4.00.1. The solution was stirred for 24 h. At 24 h the pH was again adjusted to 4.00.1. This procedure was repeated until all the chitosan was completely dissolved. A 500 mg mass of p-NCS-Bz-DFO was weighed and added to the resulting solution. The solution was stirred for 48 h. At 48 h the solution was purified using a Vivaflow cassette with a 100 kDa cut-off until a purification level of at least 100,000 was reached. Again using a Vivaflow cassette, the solvent is replaced by the CNS perfusion fluid at the same concentration.

Example 5: MetalSorb Purification and Conditioning

[0156] The polyacrylamide polymer containing dithiocarbamate functions, Metalsorb FZ, was supplied by SNF, France and used as is. VIVAFLOW cassettes were purchased from Sartorius and used as is. The perfusion fluid was purchased from Phymep (Perfusion Fluid CNS Sterile, item number P000151) and used as is.

[0157] A volume of 50 mL of Metalsorb 20% w/w was measured and placed in a 250 mL container. A volume of 150 mL of water was added and the solution was stirred for two hours. At 2 h, the solution was purified using a Vivaflow cassette with a 100 kDa cut-off until a purification rate of at least 100,000 was achieved. Again using a Vivaflow cassette, the solvent was replaced by CNS perfusion fluid at equal concentration.

Example 6: Use of Materials Obtained in Examples 3, 4 and 5

[0158] The materials obtained in Examples 3, 4 and 5 above may be advantageously used as a means for extracting metal cations according to the present invention. The solutions can be used directly or by adapting the formulation to form a perfusion fluid, or the polymers can be extracted and consolidated to form a macroscopic solid which can be implanted.

Example 7: Synthesis of Polysiloxane-EDTA Nanoparticles

[0159] Polysiloxane particles comprising EDTA (ethylenediaminetetraacetate) type chelates Si@EDTA are obtained by mixing three silane precursors: (i) TEOS (tetraethylorthosilicate ((Si(OC.sub.2H.sub.5).sub.4, 98%Sigma-Aldrich Chemicals, France)), (ii) APTES (3(aminopropyl)triethoxy silane (H.sub.2N(CH.sub.2).sub.3Si(OC.sub.2H.sub.5).sub.3, 99%Sigma-Aldrich Chemicals, France)) and (iii) Si-EDTA (N-(trimethoxysilylpropyl)ethylenediaminetriacetic acid, trisodium salt (N-[3-trimethoxysilylpropyl]ethylenediamine triacetic acid trisodium salt at 45% in water, ABCR, Germany)). The 3 precursors are placed in DEG (diethylene glycol-DEG, 99% SDS Carlo Erba (France)) with a molar ratio 2:1:3 (TEOS/APTES/Si-EDTA). The mixture is kept under stirring at room temperature for 30 minutes before adding a 3 times higher volume of water and a new stirring phase of 17 hours at the same temperature. The temperature is then raised to 80 C. and stirring is maintained for 6 hours (the pH is adjusted to a value of 7.4 after two hours of heating). The heating is then switched off and the solution is kept under stirring for 17 hours. The solution is then purified by tangential filtration. The nanoparticles have a hydrodynamic diameter of 219 nm in dynamic light scattering (DLS) using a PCS-based Malvern Zeta Sizer Nano-S particle size analyzer (FIG. 8).

Example 8: Synthesis of Polysiloxane-DTPA Nanoparticles

[0160] For nanoparticles comprising chelates of the DTPA (diethylenetriaminepentaacetic acid) type, a preliminary step is necessary to graft the chelate onto a silane. The silane comprising DTPA is obtained by reacting a derivative of DTPA: DTPA-BA (diethylenetriaminepentaaceticacid dianhydride CheMatech, Dijon, France) with APTES in DEG in a ratio of 1:1 DTPA-BA/APTES. The solution is left under stirring for 24 hours. Then TEOS is added with a 3:1:1 TEOS/APTES/DTPA-BA ratio. After 1 hour under stirring in DEG, water is added (10 times the volume of DEG used). The solution is then stirred for 24 hours at room temperature, heated to 50 C. and stirred again for 24 hours. Finally, the solution is cooled to room temperature and left to stir for 72 hours. The nanoparticles are then purified by tangential filtration and the pH is raised to 7.4. The nanoparticles have a hydrodynamic diameter of 73 nm in DLS, evaluated using a PCS-based Malvern Zeta Sizer Nano-S particle size analyzer, with a second population at 207 nm (FIG. 9).

Example 9: Comparison of Microdialysis Flow Rates on Metal Extraction

[0161] The extractability of a perfusion fluid comprising a chelator in a microdialysis device from an aqueous solution comprising several metal cations was evaluated in this example.

[0162] Several flow rates (1, 2 and 5 L/min) were tested using the same perfusion fluid (polysiloxane-EDTA nanoparticles whose synthesis is described in Example 8 at an EDTA concentration of 15 mM dispersed in water). The microdialysis membrane (63 Microdialysis Catheter, M Dialysis AB, Sweden) had a cut-off of 20 kDa. The solution used to test the chelating ability of the perfusion fluid was an aqueous solution comprising Al(III), Cd(II), Zn(II), Cu(II) and Pb(II) ions each at a concentration of 100 ppb. The pH of the solution was adjusted to 7.4 and HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, Sigma-Aldrich Chemicals (France) was added as a buffer at a concentration of 1.2 g/L. The total volume of the solution is 600 mL. Microdialysis extraction took 40 min at flow rates of 2 and 5 L/min. The sample at 1 L/min was obtained in 100 minutes. These samples were analyzed by ICP/MS and the amounts of each metal were reported in Table 2. This experiment was replicated 4 times and shows a better extraction for each of the metals using the perfusion fluid based on chelating nanoparticles compared to a conventional microdialysis use where the perfusion fluid initially contains only water (H.sub.2O) under all conditions tested. Metal uptake at concentrations above their diffusion concentration in the medium to be purified was observed in the case of the use of a perfusion fluid comprising a chelator. Chelation is particularly effective for aluminum because of its smaller size, which allows faster diffusion through the membrane. A flow rate of 2 L/min appears to be a good compromise between efficient extraction and sample amount and has been selected for Examples 10 and 11.

TABLE-US-00002 TABLE 2 Concentration of extracted metals in the perfusion fluid by comparing water and polysiloxane-EDTA nanoparticles (15 mM) at different flow rates. Perfusion Flow rate Al Cu Cd Pb Fluid (L .Math. min.sup.1) (ppb) (ppb) (ppb) (ppb) H.sub.2O 5 15 32 24 33 H.sub.2O 2 32 55 29 83 H.sub.2O 1 72 55 61 86 Polysiloxane - 5 55 51 50 51 EDTA Polysiloxane - 2 436 85 105 95 EDTA Polysiloxane - 1 292 106 67 109 EDTA

Example 10: Comparison of Perfusion Fluids Based on Polysiloxane-DTPA and Polysiloxane-EDTA Nanoparticles

[0163] The relative efficiency of the nanoparticles obtained in Examples 7 and 8 was compared using the same metal mixture as described in Example 9 with a microdialysis flow rate of 2 L/min.sup.1 and a sample collection time of 40 min with a microdialysis membrane having a cut-off of 20 kDa. Table 3 summarizes the results obtained using 3 different perfusion fluids: (i) water, (ii) polysiloxane-EDTA nanoparticles and (iii) polysiloxane-DTPA nanoparticles at a chelator concentration of 15 mM. DTPA-based nanoparticles have a very high aluminum extraction capacity due to the very high affinity of the chelator for this species. The presence of aluminum seems to saturate the surface chelators reducing the efficiency of the fluid for other metals. The polysiloxane-DTPA nanoparticles make it possible to obtain a very specific perfusion fluid for the extraction of aluminum.

TABLE-US-00003 TABLE 3 Concentration of extracted metals in the perfusion fluid comparing water and polysiloxane-EDTA and polysiloxane- DTPA nanoparticles (15 mM) at a flow rate of 2 L/min.sup.1. Perfusion Flow rate Al Cu Cd Pb Fluid (L .Math. min.sup.1) (ppb) (ppb) (ppb) (ppb) H.sub.2O 2 32 55 29 83 Polysiloxane - 2 436 85 105 95 EDTA Polysiloxane - 2 702 56 78 84 DTPA

Example 11: Use of Polysiloxane-EDTA Nanoparticles as Perfusion Fluid for Cerebrospinal Fluid (CSF)

[0164] In order to model the CSF, a solution composed of NaCl (147 mM), KCl (2.7 mM), CaCl.sub.2) (1.2 mM) and MgCl.sub.2 (0.85 mM) was synthesized. The metal extraction was carried out with this solution in order to check that the extraction power was not diminished by the different ions that could interfere. A solution similar to the one in Example 9 was made up (i.e. 600 mL of reconstituted CSF containing 100 ppb of each of the ions Al(III), Cd(II), Zn(II), Cu(II) and Pb(II)). The microdialysis membrane (63 Microdialysis Catheter, M Dialysis AB, Sweden) used has a cut-off of 20 kDa and the flow rate was set at 2 L/min.sup.1 with a collection time of 40 min. The analysis of the extracted metal amounts was performed by ICP/MS. The perfusion fluid consisted of either reconstituted CSF or the polysiloxane-EDTA nanoparticles whose synthesis is described in Example 7 dispersed in the reconstituted CSF. The results of the extraction are given in Table 4. It can be noted that the perfusion fluid containing only the CSF has a very low extraction capacity. The addition of the nanoparticles to the perfusion fluid significantly increases the metal extraction regardless of the metal. In these conditions, a metal extraction factor of more than 5 for lead, more than 7 for copper, more than 25 for cadmium and more than 125 for aluminum is gained.

TABLE-US-00004 TABLE 4 Concentration of extracted metals in a CSF solution comparing CSF and polysiloxane-EDTA nanoparticles (10 mM) as perfusion fluid at a flow rate of 2 L/min.sup.1. CSF perfusion Flow rate Al Cu Cd Pb fluid (L/min.sup.1) (ppb) (ppb) (ppb) (ppb) CSF 2 5 8 4 21 Polysiloxane- 2 643 63 112 116 EDTA