Micron-scale biocompatible materials that can bind to negatively charged biomembrane bearing species

Abstract

The invention relates to biocompatible micron-scale spherical or non-spherical materials that can selectively separate bacterial cells from blood and various other liquids, and the synthesis methods thereof.

Description

[0001] MICRON-SCALE BIOCOMPATIBLE MATERIALS THAT CAN BIND TO

[0002] NEGATIVELY CHARGED BIOMEMBRANE BEARING SPECIES

[0003] Technical Field of the Invention

[0004] The invention relates to biocompatible micron-scale spherical or non-spherical materials capable of selectively separating bacterial cells from various liquids. The invention could potentially be used for the separation and determination of bacterial contamination in pharmaceutical products, food products, and wastewater discharges. The invention has application areas such as separating bacteria from the blood of bacteremia patients, separating bacteria from environmental and food samples, and separating species with membrane properties similar to bacteria from similar matrices.

[0005] State of the Art of the Invention (Prior Art)

[0006] Bacteremia is the presence of bacteria in the blood, which should normally be sterile. If bacteremia cannot be diagnosed and treated quickly, this may cause sepsis. Sepsis can be defined as the body's immune response to bacterial infection being very severe and the condition damaging healthy tissue. Sepsis may result in septic shock, organ failure, and death. According to CDC data, 1.7 million patients get sepsis in the USA every year, of which approximately 270000 die from sepsis. 30% of patients who die in hospital intensive care units die as a result of sepsis. 15-30% of blood infections worldwide result in death. With the current methods, the clinical diagnosis of bacteremia takes 2-4 days, and its treatment takes 14 days.

[0007] In the state of the art, there are materials developed for the separation of bacterial cells in the blood.

[0008] Bradley D. Smith et al. described the development of synthetic zinc(II)-dipicolylamine (ZnDPA) receptors as selective targeting agents for anionic membranes in cell culture and living experimental subjects. ZmBDPA systems have been known to target bacterial membranes since at least 2006. The preparation of micron-scale support materials carrying ZmBDPA complexes and the separation of negatively charged membrane-carrying species (liposomes, bacterial cells, viruses, yeast cells, cancer cells) from liquids are not mentioned in this document.

[0009] In patent document US 2014 / 0212335 Al, nanoparticles are derivatized to bind to bacterial contaminants such as Gram-positive bacteria, Gram-negative bacteria, and endotoxins. The nanoparticles comprise a core containing a magnetic material and ligands bound to the core. Ligands that bind to bacterial contaminants include, for example, a metal ion to form a bis- Zn-DPA or bis-Cu-DPA, e.g., bis(dipicolylamine) (“DPA”) coordinated with Zn2+or Cu2+. Nanoparticles allow the separation of bacterial contaminants when added to liquids such as blood, e.g., whole, or diluted blood, buffer solutions, albumin solutions, beverages for human and / or animal consumption, e.g., drinking water, liquid drugs for humans and / or animals, or other liquids. The ZmBDPA complex used in the mentioned invention has a low affinity for negatively charged membranes. It also requires a microfluidic device since nanoparticles are used. Therefore, it is not suitable for applications requiring high flow rates and high volumes, e.g., whole blood filtration. Some studies have shown that dendrimers, dyes, and liposomes carrying ZmBDPA can selectively bind to bacterial cells in vivo and in vitro [1-4],

[0010] The patent document WO 2015 / 164198 Al provides a method for removing a significant amount of bacteria (e.g., Gram-negative bacteria and Gram-positive bacteria, including bacteria that have no or low affinity for heparan sulfate) from whole blood, serum, or plasma using an adsorption medium. The main invention in this document is that heparin sulfate can bind some bacterial strains. Although it is not clear what the substrate used is, it may be a synthetic polymer. It is stated that the substrates used are micron-sized. Whole blood filtration requires micron-scale particles, and this is a known condition.

[0011] None of the known systems in the state of the art is suitable for the separation of negatively charged membrane carrying species (liposomes, bacterial cells, yeast cells, free circulating cancer cells, and some virus particles) from liquids of various volumes (from milliliters to liters) flowing at high flow rates and for complete (i.e., high volume) blood filtration. Brief Description and Objects of the Invention

[0012] The present invention is related to micron-scale spherical and nonspherical materials derivatized with ligands that can selectively bind to bacterial cells, that satisfy the aforementioned requirements, that eliminate all the disadvantages of the prior art and put forward some additional advantages, and that can selectively remove bacterial cells from various liquids, and is also related to the synthesis of said materials.

[0013] With this invention, it is possible to separate negatively charged membrane-carrying species (liposomes, bacterial cells, yeast cells, free circulating cancer cells, and some virus particles) from liquids (blood, diluted blood, food samples, drug samples, waste waters, etc.) of various volumes (from milliliters to liters) flowing at any flow rate above the diffusion limit or a high flow rate of 0.0001 mL / min to 100000 mL / min by derivatizing micron-scale, spherical or non-spherical and biocompatible materials with ZmBDPA ligands. The same separation process with the materials of the invention can also be performed with still liquids.

[0014] The primary object of the invention is the selective separation of bacterial cells from various liquids. For this purpose, micron-scale spherical materials derivatized with ligands that can selectively bind to bacterial cells were obtained. It is argued that the micron-scale (50-300 pm) particle size will also allow the flowing liquids to pass through a filtration column consisting of these particles. In this way, bacterial cells can be selectively separated from various liquids, such as blood.

[0015] The advantages of the micron-scale spherical material synthesized by the invention are listed below: i. Suitable for the separation of bacterial cells from high volume liquids. ii. Suitable for large-scale production. iii. Suitable for the separation of bacterial cells from various liquids, including blood. iv. In addition to bacterial cells, they can be used for the separation of circulating tumor cells (i.e., cancer cells circulating in the blood), yeasts, and viral particles (e.g., influenza and SARS-CoV-2) from liquid biological samples, such as blood, and from environmental samples.

[0016] With this invention, it is possible to selectively separate bacteria and species with similar membrane properties from liquids. By immobilizing ZmBDPA complexes, which have the capability of selectively binding to bacteria (and other species with similar membrane properties), onto low porosity microparticles (in this case cellulose microparticles) it is possible to separate said species from liquids flowing at a certain rate. In this way, by using whole blood filtration, the concentration of bacteria in the blood of bacteremia patients can be reduced and the patients can be cured. In the same way, bacteria in analytical blood samples can be separated using such particles and bacteremia diagnosis can be facilitated. The materials of the invention can also be used both for the large-scale separation of bacteria from similarly flowing environmental and food samples and for the separation of bacteria from analytical-scale samples. During the invention studies, these materials have been shown to bind to bacteria (1-1.5 micrometers in size) and negatively charged liposomes (150 nm in size). Such materials have the potential to bind to some other species with negatively charged lipid membranes, such as some viruses (e.g., influenza viruses, coronaviruses), some yeast species, and cancer cells circulating in the blood. Therefore, the present invention has many potential applications.

[0017] Prototype materials developed for this purpose were obtained by appropriately derivatizing spherical regenerated cellulose microspheres (CM, diameter 50-75 pm) known to be biocompatible and conjugating them with zinc(II) bisdipicolylamine (ZmBDPA) complexes that are known to bind selectively to negatively charged phosphate amphiphiles (e.g., phosphatidylglycerol, Lipid A, lipoteichoic acid) on the surfaces of bacterial cells. The materials obtained by three different chemical approaches are BDPA-CM-I, BDPA-CM-II, and BDPA-CM-III.

[0018] With the invention, a ligand that can bind to a broad spectrum of bacterial cells is conjugated to the micron-scale particles, and these derivatized particles bind to negatively charged liposomes and bacterial cells. Scaled production of spherical and nonspherical microparticles derivatized with ligands capable of selectively binding to bacterial cells is possible and such particles can be used for the removal of bacterial cells from scaled (i.e., large volume) liquids. In addition, it will be possible to diagnose bacteremia rapidly with cartridges, columns, or kits containing these materials and to perform bacteremia treatment with full blood filtration. With the resulting prototype material, in addition to bacteria, it will be possible for cancer cells, some yeast cells, and some viral particles circulating in the blood to be separated from blood and other biological liquids. With these particles, it is possible to separate the relevant species not only from still liquids but also from flowing liquids.

[0019] With the invention, conjugates of micron-scale particles with ZmBDPA complexes were synthesized, and these functional microparticles were shown to bind to negatively charged liposomes and bacterial cells. In addition, these materials are expected to bind to species bearing negatively charged phosphate lipids similar to bacterial cells such as other microbial pathogens, viruses, and free circulating cancer cells in the blood.

[0020] It will be possible to quickly determine blood infections and to quickly separate the bacteria in blood with the materials of the invention. In addition, a large number of applications requiring bacterial separation will be possible with these materials. Species such as yeast and viral particles with lipid membranes similar to bacteria can also be separated from biological and other liquids with these materials.

[0021] Products such as cartridges, syringe filters, kits, and other possible devices or apparatus containing the materials synthesized by the invention will be obtained and it will be possible to selectively separate bacteria, viruses, yeast cells, and cancer cells circulating in the blood from various liquids.

[0022] Definitions of Figures Describing the Invention

[0023] The figures and related descriptions required to better understand the subject of the invention are as follows.

[0024] Figure 1: Preparation of CM-epoxy, CM-amine, and BDPA-CM-I.

[0025] Figure 2: Preparation of CM-bisaldehyde and BDPA-CM-II Figure 3: Preparation of BDPA-CM-III. Figure 4: Normalized fluorescence data after incubation of liposomal dispersions with CMs (Liposome A = neutral liposomes, Liposome B = negatively charged liposomes).

[0026] Figure 5: Bright field micrographs (lOx) for CM-amine (A) and BDPA-CM-I (C) and fluorescence micrographs (lOx, TX-2 filter) for CM-amine (B) and BDPA-CM-I (D) after treatment with negatively charged liposomes and washing.

[0027] Figure 6: Bright field micrographs (lOx) for CM-bisaldehyde (A) and BDPA-CM-II (C) and fluorescence micrographs (lOx, TX-2 filter) for CM-bisaldehyde (B) and BDPA-CM-II (D) after treatment with negatively charged liposomes and washing.

[0028] Figure 7: Bright field micrographs (lOx) for CM-epoxy (A) and BDPA-CM-III (C) and fluorescence micrographs (lOx, TX-2 filter) for CM-epoxy (B) and BDPA-CM-III (D) after treatment with negatively charged liposomes and washing.

[0029] Figure 8: Bright field micrographs (lOx) for CM-amine (A) and BDPA-CM-I (C) and fluorescence micrographs (lOx, GFP filter) for CM-amine (B) and BDPA-CM-I (D) after treatment with GFP expressing Escherichia coli cells and washing.

[0030] Figure 9: Bright field micrographs (lOx) for CM-bisaldehyde (A) and BDPA-CM-II (C) and fluorescence micrographs (lOx, GFP filter) for CM-bisaldehyde (B) and BDPA-CM-II (D) after treatment with GFP expressing Escherichia coli cells and washing.

[0031] Figure 10: Bright field micrographs (lOx) for CM-epoxy (A) and BDPA-CM-III (C) and fluorescence micrographs (lOx, GFP filter) for CM-epoxy (B) and BDPA-CM-III (D) after treatment with GFP expressing Escherichia coli cells and washing.

[0032] Detailed Description of the Invention

[0033] In this detailed description, biocompatible micron-scale spherical or non-spherical materials capable of selectively separating bacterial cells from various liquids, and their method of preparation and characterization are described only for a better understanding of the subject matter and without any limiting effect.

[0034] The material that can bind to negatively charged biomembrane bearing species, is characterized in that it contains microparticles obtained from 20-1000 pm particle-sized materials with a porosity of less than 500 nm, and structures selected from the following groups is bound to the surface of these microparticles: • Zinc(II) bisdipicolylamine (ZmBDPA) complexes or complexes of other transition group metal cations with the BDPA ligand

[0035] • Complexes of zinc (II) and other transition group metal cations with DPA, cyclene, or imidazole (bis, tris, tetra, or poly) ligands

[0036] • Complexes obtained with ligand structures containing nitrogen, phosphorus, sulphur or oxygen, or mixtures thereof, known to form complexes with zinc (II) and other transition group metals

[0037] • Cationic groups of amine, polyamine, ammonium, polyammonium, guanidinium, or polyguanidinium,

[0038] • Peptides, proteins, antibiotics, or antibodies.

[0039] The invention relates to biocompatible cellulose microspheres having low porosity (less than 500 nm) decorated with ZmBDPA complexes capable of selectively separating species bearing cell membranes displaying negatively charged lipids from various liquids. Species displaying negatively charged lipids on their cell membranes can be liposomes, bacteria, viruses, yeast cells, and cancer cells that circulate in the blood. Liquids from which bacteria and species with similar membrane properties can be separated include blood, plasma, food, environmental samples, and pharmaceutical products and formulations.

[0040] Apart from cellulose decorated with ZmBDPA complexes, species bearing cell membranes displaying negatively charged lipids can be separated from liquids by using microspheres manufactured from other materials with low porosity, such as glass, synthetic and natural polymers. The species can be separated from liquids which such materials can be liposomes, bacteria, viruses, yeast cells, and cancer cells that circulate in the blood.

[0041] The object of the invention is to separate bacterial cells from liquids using micron-scale spherical materials derivatized with ligands (ZmBDPA complexes) that can selectively bind to bacterial cells (and species with membrane structures carrying similar negatively charged phosphate amphiphiles). The ligands bound to the particles provide selective binding. The micron-scale (50-300 pm) particle size allows the flowing liquids to pass through a filtration column of these particles. In this way, bacterial cells can be selectively separated from various liquids, such as blood. Such materials have the potential to bind to some other species with negatively charged lipid membranes, such as some viruses (e.g., influenza viruses, coronaviruses), some yeast species, and cancer cells circulating in the blood.

[0042] In the structure of the ZmBDPA complex ligand, instead of Zn2+(zinc ion) as the metal cation, there may be Cu2+(copper ion) or other transition metals with stable oxidation numbers of +2 and +3.

[0043] In the ligand structure, instead of BDPA, there may be ligand structures containing DPA, cyclene, bis- or tetraimidazole, or nitrogen, phosphorus, sulfur, or oxygen that can form complexes with transition elements with stable oxidation numbers of +2 and +3.

[0044] The microsphere-bound recognition group (i.e., the ZmBDPA ligand in the prototype material) may additionally be cationic groups (amine, ammonium, guanidinium, etc.), antibodies, peptides, antibiotics (e.g., vancomycin), or carbohydrates that can bind to bacterial cells and species with similar membrane structures.

[0045] Prototype materials developed for this purpose were obtained by appropriately derivatizing regenerated spherical cellulose microspheres (CM, diameter 50-75 pm) known to be biocompatible and conjugating them with zinc(II) bisdipicolylamine (ZmBDPA) complexes known to bind selectively to negatively charged phosphate amphiphiles (e.g., phosphatidylglycerol, Lipid A, lipoteichoic acid) on the surfaces of bacterial cells. The materials obtained by three different chemical approaches are BDPA-CM-I, BDPA-CM-II, and BDPA-CM-III. BDPA-CM-I was obtained by conjugation of BDPA-Acid and CM- Amine (Figure 1). BDPA-CM-II was obtained from the reaction of CM-Bisaldehyde with BDPA- Amine (Figure 2). BDPA-CM-III was obtained from the reaction of CM-Epoxy with BDPA-Amine (Figure 3). These materials are characterized by FTIR and elemental analysis.

[0046] Procedures and Characterization Data for BDPA-CM-I, BDPA-CM-II, and BDPA-CM- III

[0047] 1. Preparation of BDPA-CM-I

[0048] CM-Epoxy: 0.5 mL of epichlorohydrin and 1.5 mL of 2.5 M aqueous NaOH solution were added to 500 mg cellulose microspheres. The reaction was allowed to proceed for 2.5 hours at 40°C in a shaking incubator at 250 rpm. Following the reaction, the microspheres were filtered and washed with deionized water and acetone. After air drying for 24 hours, CM- epoxy was lyophilized. The oxirane concentration of CM-epoxy was determined by the thiosulfate titration method. Functional groups in the spheres were determined by FTIR. FTIR (KBr) v 1580, 1237, 1022, 896, 796.7 cm'1.

[0049] CM-Amine: 1 mL of 25% NFL OH solution was added to 100 mg CM-epoxy in the test tube and the reaction was continued at room temperature for 16 hours in a shaking incubator at 150 rpm. Following the reaction, the microspheres were filtered and washed with deionized water and acetone. After air drying for 24 hours, CM-amine was lyophilized. The lyophilized CM- amine spheres were examined by CHNS analysis and the amine group concentration in the spheres was in the range of 257-421 pmol / g. Functional groups in the spheres were determined by FTIR. FTIR (KBr) v 1427, 1050 cm'1.

[0050] BDPA-CM-I: To 25 mg BDPA-acid in 2.5 mL DMF (dimethylformamide) was added 7.19 mg (0.039 mmol) of PfpOH (pentafluorophenol), 7.47 mg (0.039 mmol) of EDC (l-ethyl-3- (3-dimethylaminopropyl)carbodiimide), and 1 mg (0.007 mmol) of DMAP (p-N,N- dimethylaminopyridine) at 0°C, respectively. After the reaction was continued at 0°C for 10 minutes, it was brought to room temperature and continued at this temperature for 16 hours. 5.05 mg (6.81 pL, 0.039 mmol) of DIPEA (ethyldiisopropylamine) and 35 mg (containing 343 pmol / g of amine) of CM-amine were added to the reaction in 0.5 mL DMF, respectively. The reaction was continued for 16 hours at room temperature and under a nitrogen atmosphere. After the solvent was removed under low pressure, BDPA-CM-I spheres were washed with 3x25 mL deionized water and 1x25 mL ethanol. Lyophilized BDPA-CM-I spheres were examined by CHNS analysis and the BDPA ligand concentration in the spheres was found to be 0.123 pmol / mg. Functional groups in the spheres were determined by FTIR. FTIR (KBr) v 1651, 1061, 894, 766 cm'1.

[0051] 2. Preparation of BDPA-CM-II

[0052] CM-Bisaldehyde: 1.5 mL 0.20 M NalOi (in water) was added to 488.1 mg cellulose microspheres, and the reaction was continued for 24 hours at room temperature in a shaking incubator at 250 rpm in the dark. After the reaction, the microspheres were filtered and washed five times with deionized water. CM-Bisaldehyde spheres were stored at 4°C until use. An analytical sample was lyophilized and the content of aldehyde by titration was found in the range of 5.79-12.87 mmol / g. Functional groups in the spheres were determined by FTIR. FTIR (KBr) v 1681, 894 cm'1. BDPA-CM-II: 100 mg (0.2 mmol) of BDPA-amine in 5 mL MeOH (methanol) was added to 100 mg (1.3 mmol aldehyde functional group) CM-bisaldehyde and the reaction was continued for 16 hours at room temperature in a shaking incubator at 250 rpm. 38.6 mg (1 mmol) of NaBEL (sodium borohydride) was added to the reaction and the reaction was continued with 250 rpm agitation at room temperature for 4 hours. After filtration, the spheres were washed with 1x5 mL of 1 M NaOH and 3x25 mL of deionized water, and 1x25 mL of ethanol. The BDPA-CM-II obtained was lyophilized and the BDPA ligand concentration in the spheres was found to be 0.070 pmol / mg by CHNS analysis. Functional groups in the spheres were determined by FTIR. FTIR (KBr) v 1592, 1156, 1026, 894, 798 cm'1.

[0053] 3. Preparation of BDPA-CM-III

[0054] CM-Epoxy BDPA-Amine BDPA-CM-III

[0055] BDPA-CM-III: 50 mg (83.8 pmol) BDPA-amine was added to 100 mg (83.8 pmol epoxy group containing) CM-Epoxy in 1 mL of acetic acid and the reaction was boiled using a reflux condenser under nitrogen atmosphere for 16 hours. The reaction was washed with 20 mL of ethanol, 10 mL of 0.1 M HC1, 10 mL of 0.1 M NaOH, and 10 mL of deionized water. The BDPA-CM-III obtained was lyophilized and the BDPA ligand concentration in the spheres was found to be 0.020 pmol / mg by CHNS analysis. Functional groups in the spheres were determined by FTIR. FTIR (KBr) v 1596, 1157, 1023, 796 cm'1.

[0056] Two different studies were conducted to determine the affinity of BDPA-CM-I, BDPA-CM- II, and BDPA-CM-III to negatively charged membranes. In the first study, the binding of these conjugates and controls (CM-Amine for BDPA-CM-I, CM-Bisaldehyde for BDPA-CM- II, CM-Epoxy for BDPA-CM-III) to negatively charged liposomes was examined. In liposome studies, neutral liposomes were made from phosphatidylcholine (68%), cholesterol (30%), and DiO fluorescent dye (2%), and negatively charged liposomes were made from phosphatidylglycerol (10%) which is a negatively charged lipid, phosphatidylcholine (58%), cholesterol (30%), and DiD fluorescent (red) dye (2%). The diameters of the liposomes used are between 140-180 nm on average. It was observed that BDPA-CMs and their respective controls did not significantly bind to neutral liposomes, negatively charged liposomes were bound in significant amounts by BDPA-CM-I, BDPA-CM-II, and BDPA-CM-III, but not in significant amounts by their associated controls (Figure 4, Liposome A = neutral liposomes, Liposome B = negatively charged liposomes). Fluorescence microscopy has also shown that BDPA-CM-I, BDPA-CM-II, and BDPA-CM-III bind to negatively charged liposomes (Figures 5, 6, and 7). It can be calculated that BDPA-CM-I and BDPA-CM-III particles with an average diameter of 62 pm each bind an average of 69000 liposomes per microsphere. BDPA-CM-II binds to a slightly lesser amount of negatively charged liposomes. In the second study conducted to determine the affinity of BDPA-CM-I, BDPA-CM-II, and BDPA-CM-III to negatively charged membranes, the binding of these conjugates and their controls to GFP (green fluorescent protein) expressing Escherichia coli bacterial cells was examined. CMs were incubated with bacterial dispersions, washed, and examined with bright field and fluorescence microscopy. In this study, it was observed that BDPA-CMs were able to bind to bacterial cells much better than their controls (Figure 8, 9, 10).

[0057] These studies have shown that cellulose microspheres with diameters of 50-75 pm, derivatized with a ligand that can bind to bacterial cells, can bind to negatively charged liposomes and bacterial cells. Some potential application areas of the materials subject to this this invention are listed below:

[0058] • Preconcentration of bacterial cells by separating them from blood for rapid diagnosis of bacteremia that may be defined as bacteria in the blood or blood infection

[0059] • Whole blood filtration with columns containing these materials for the treatment of bacteremia

[0060] • Separation of bacterial cells from environmental, food, and drug samples.

[0061] In addition to these potential applications, such materials can be used to separate cancer cells circulating in the blood, yeasts, and viral particles (e.g., influenza and SARS-CoV-2), all of which have negatively charged cell membranes, from liquid biological and environmental samples.

[0062] In addition to cellulose, which has been used in the preparation of the prototype material, other polymeric materials can be used as the support material. In addition to the ZmBDPA complexes used in the prototype material, complexes of other transition group metal cations with BDPA or ligands with other structures, cationic groups, peptides, antibiotics, and antibodies can be used as affinity ligands.

[0063] References

[0064] 1. Rice, D.; Plaunt, A. J.; Turkyilmaz, S.; Smith, M.; Wang, Y.; Rusckowski, M.; Smith B.D. Evaluation of [11 Iln]-Labeled Zinc-Dipicolylamine Tracers for SPECT Imaging of Bacterial Infection. Mol. Imaging Biol. 2015, 77, 204-213

[0065] 2. Turkyilmaz, S.; Rice, D.R.; Palumbo, R.; Smith B.D. Selective recognition of anionic cell membranes using targeted liposomes coated with zinc(II)-bis(dipicolylamine) affinity units. Org. Biomol. Chem. 2014, 12, 5645-5655

[0066] 3. Xiao, S.; Abu-Esba, L.; Turkyilmaz, S.; White, A.G.; Smith B.D. Multivalent Dendritic Molecules as Broad Spectrum Bacteria Agglutination Agents. Theranostics 2013, 3 (9), 658-666

[0067] 4. Xiao, S.; Turkyilmaz, S.; Smith, B.D. Convenient Synthesis of Multivalent Zinc(II)- Dipicolylamine Complexes as Potential Cell Targeting Agents. Tetrahedron Let. 2013, 54, 861-864

Claims

CLAIMS1. A material that can bind to negatively charged biomembrane bearing species, comprising microparticles obtained from a material with a porosity of less than 500 nm and are in 20-1000 pm particle size range, wherein the structure selected from the following group is bound to the surface of these microparticles:• Zinc(II) bisdipicolylamine (ZmBDPA) complexes or complexes of other transition group metal cations with BDPA ligands• Complexes of zinc (II) and other transition group metal cations with DPA, cyclene, or imidazole (bis, tris, tetra, or poly) ligands• Complexes obtained with ligand structures containing nitrogen, phosphorus, sulphur or oxygen or mixtures thereof known to form complexes with zinc (II) and other transition group metal cations• Cationic groups like amines, polyamines, ammoniums, polyammoniums, guanidiniums, or polyguanidiniums,• Peptides, proteins, antibiotics, or antibodies.

2. A material according to claim 1, wherein the material has a spherical or non-spherical shape.

3. A material according to claim 1, wherein the microparticles are microparticles obtained from cellulose.

4. A material according to claim 1, wherein the microparticles are microparticles obtained from glass, synthetic and natural polymers, metals, or composites of these materials.

5. A material according to claim 1, which are used in the separation of the species with cell membranes displaying negatively charged lipids from liquids with any flow rate above the diffusion limit or 0.0001 mL / min and under 100000 mL / min or from still liquids.

6. A material according to claim 5, wherein the liquid is blood, plasma, food, environmental samples, wastewater discharges, pharmaceutical products, or pharmaceutical formulations.

7. A material according to claim 1, wherein the species with cell membranes displaying negatively charged lipids are bacteria, viruses, yeast cells or cancer cells circulating in the blood.

8. An apparatus comprising the material of any one of the preceding claims.

9. A device comprising the material of any one of the preceding claims.

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