CN109174033B - Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof - Google Patents

Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof Download PDF

Info

Publication number
CN109174033B
CN109174033B CN201810981030.3A CN201810981030A CN109174033B CN 109174033 B CN109174033 B CN 109174033B CN 201810981030 A CN201810981030 A CN 201810981030A CN 109174033 B CN109174033 B CN 109174033B
Authority
CN
China
Prior art keywords
lead
blood
erythrocytes
ion scavenger
solution
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active
Application number
CN201810981030.3A
Other languages
Chinese (zh)
Other versions
CN109174033A (en
Inventor
万密密
毛春
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Nanjing Normal University
Original Assignee
Nanjing Normal University
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Nanjing Normal University filed Critical Nanjing Normal University
Priority to CN201810981030.3A priority Critical patent/CN109174033B/en
Publication of CN109174033A publication Critical patent/CN109174033A/en
Application granted granted Critical
Publication of CN109174033B publication Critical patent/CN109174033B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/02—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material
    • B01J20/06—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material comprising oxides or hydroxides of metals not provided for in group B01J20/04
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
    • B01D15/08—Selective adsorption, e.g. chromatography
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/02—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material
    • B01J20/10—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material comprising silica or silicate
    • B01J20/103—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material comprising silica or silicate comprising silica
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
    • B01J20/24—Naturally occurring macromolecular compounds, e.g. humic acids or their derivatives
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
    • B01J20/26—Synthetic macromolecular compounds
    • B01J20/262—Synthetic macromolecular compounds obtained otherwise than by reactions only involving carbon to carbon unsaturated bonds, e.g. obtained by polycondensation
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
    • B01J20/28002—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their physical properties
    • B01J20/28009—Magnetic properties
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2220/00—Aspects relating to sorbent materials
    • B01J2220/40—Aspects relating to the composition of sorbent or filter aid materials
    • B01J2220/46—Materials comprising a mixture of inorganic and organic materials
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2220/00—Aspects relating to sorbent materials
    • B01J2220/40—Aspects relating to the composition of sorbent or filter aid materials
    • B01J2220/48—Sorbents characterised by the starting material used for their preparation
    • B01J2220/4806—Sorbents characterised by the starting material used for their preparation the starting material being of inorganic character
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2220/00—Aspects relating to sorbent materials
    • B01J2220/40—Aspects relating to the composition of sorbent or filter aid materials
    • B01J2220/48—Sorbents characterised by the starting material used for their preparation
    • B01J2220/4812—Sorbents characterised by the starting material used for their preparation the starting material being of organic character
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2220/00—Aspects relating to sorbent materials
    • B01J2220/40—Aspects relating to the composition of sorbent or filter aid materials
    • B01J2220/48—Sorbents characterised by the starting material used for their preparation
    • B01J2220/4812—Sorbents characterised by the starting material used for their preparation the starting material being of organic character
    • B01J2220/4825—Polysaccharides or cellulose materials, e.g. starch, chitin, sawdust, wood, straw, cotton

Landscapes

  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Organic Chemistry (AREA)
  • Inorganic Chemistry (AREA)
  • External Artificial Organs (AREA)
  • Solid-Sorbent Or Filter-Aiding Compositions (AREA)

Abstract

The invention discloses a blood lead ion scavenger capable of safely entering and exiting red blood cells, and a preparation method and application thereof. Meanwhile, the blood lead ion scavenger has a novel blood lead removal mechanism, namely the blood lead ion scavenger freely enters and exits erythrocytes, the lead-containing hemoglobin is captured through the complexation of amino groups in the mesoporous silicon dioxide structure on lead ions, and the lead-containing hemoglobin is fixed through mesoporous channels, so that the purpose of removing the lead ions in the erythrocytes is achieved, and the lead ions in the blood are efficiently removed.

Description

Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof
Technical Field
The invention belongs to a novel biological material, and particularly relates to a blood lead ion remover capable of safely entering and exiting red blood cells, and a preparation method and application thereof.
Background
The health of human body is greatly threatened by the exposure of lead ions, and the lead ions can cause serious toxic effects and diseases (such as heart diseases, kidney diseases and the like). When lead ions enter the body, they are mainly deposited in the bones, soft tissues and blood. Based on the biomedical level of today, it is very difficult to remove lead ions deposited in human bone and soft tissues. Therefore, the removal of trace lead ions (blood lead) from blood is of great significance. However, more than 95% of the blood lead is located in erythrocytes and binds to hemoglobin, which makes removal of blood lead a great difficulty. At present, the clinical treatment method for the blood lead poisoning can be divided into the administration of lead-expelling medicines and blood perfusion; the method for treating mild blood lead poisoning comprises administering small molecule lead expelling medicine such as disodium edetate calcium, dimercaprol, and dimercaptosuccinic acid, and has long treatment period (40-60 days, great side effects and poor treatment effect); the treatment method of severe blood lead poisoning is blood perfusion, wherein blood perfusion is to use a blood perfusion device containing resin or activated carbon with adsorption function to clean blood and adsorb toxic substances in blood serum. Because the adsorbent in the blood perfusion device has larger size and is fixed in the blood perfusion device, the adsorbent cannot enter red blood cells to remove lead ions in the red blood cells, so the blood perfusion device generally removes toxic substances in blood serum and has poor effect on removing the lead ions in the red blood cells. Researchers have proposed alternative strategies with emphasis on the development of blood adsorbents, however, they have introduced sonication to increase the removal efficiency, which impairs the structure of the erythrocyte membrane from practical biomedical applications. There have also been proposed strategies for removing lead ions from plasma, which attempt to achieve removal of lead ions from erythrocytes by means of the dynamic equilibrium between erythrocytes and lead ions in plasma, but the stability of the binding of blood lead to haemoglobin leads to equilibration times which are usually quite long (several months) making this method difficult to apply practically.
In summary, the main problem with current lead removal strategies is the failure of researchers to correctly understand the presence of lead. To date, there is no blood lead scavenger that safely enters and exits red blood cells and competes for adsorption of hemoglobin-complexed lead ions, and the current research and patents only mention blood bulk lead ion removal, confounding the two actual existing states of lead ions, and thus they cannot really be used to remove hemoglobin-complexed lead ions in red blood cells in amounts as high as 95%. Therefore, there is a need for a new safe and effective blood lead removal strategy, which includes designing an adsorbent that can safely enter red blood cells and effectively capture blood lead and hemoglobin, and then safely leave red blood cells and blood.
Disclosure of Invention
The purpose of the invention is as follows: aiming at the problems in the prior art, the invention provides a preparation method of a magnetic ferroferric oxide/aminated mesoporous silica composite material and a blood lead ion scavenger. The blood lead ion scavenger can be really used for removing the lead ions which are complexed with the hemoglobin and have the content of 95 percent in the red blood cells, can safely enter the red blood cells, effectively captures the blood lead and the hemoglobin polluted by the blood lead, and then safely leaves the red blood cells and the blood.
The invention also provides a blood lead ion scavenger capable of safely entering and exiting red blood cells and an application thereof.
The blood lead ion scavenger has a novel blood lead removal mechanism, namely, the blood lead ion scavenger freely enters and exits red blood cells through a magnetic ferroferric oxide/rich aminated mesoporous silica composite material, the lead-containing hemoglobin is captured through the complexation of amino groups in a mesoporous silica structure to lead ions, and the lead-containing hemoglobin is fixed through mesoporous channels, so that the purpose of removing the lead ions in the red blood cells is achieved, and the lead ions in the blood are efficiently removed.
The technical scheme is as follows: in order to achieve the above object, the present invention provides a method for preparing a blood lead ion scavenger capable of safely accessing erythrocytes, comprising the steps of:
(1) weighing ferric trichloride hexahydrate, dissolving the ferric trichloride hexahydrate in glycol solution, adding sodium acetate and sodium citrate trihydrate or polyethylene glycol for continuous reaction to obtain Fe3O4A solution;
(2) fe prepared in the step (1)3O4Uniformly mixing the solution and hydrazine hydrate, carrying out ultrasonic treatment, adding tetraethyl orthosilicate (TEOS), magnetically collecting the obtained product, re-dispersing the product into deionized water containing hydrazine hydrate and hexadecyl trimethyl ammonium bromide (CTAB), carrying out ultrasonic treatment again, and adding tetraethyl orthosilicate for reaction;
(3) and (3) drying and calcining the product obtained by the final reaction in the step (2) to remove the template agent hexadecyl trimethyl ammonium bromide, collecting the obtained product, dispersing the product into deionized water containing hydrazine hydrate and an amine-rich organic matter template agent, adding tetraethyl orthosilicate for reaction after ultrasonic treatment, and continuing the reaction to obtain the product, namely the magnetic ferroferric oxide/amine-rich mesoporous silica composite material blood lead ion scavenger.
Preferably, the reaction temperature in the step (1) is 150-220 ℃, and the reaction time is 10-24 h.
Wherein, the Fe in the step (1)3O4Has a size of 50-300 nm.
Preferably, the reaction temperature of adding tetraethyl orthosilicate in each step in the step (2) is 60-90 ℃, and the reaction time is 1-5 h.
Preferably, the molar ratio of the tetraethyl orthosilicate to the amine-rich organic template in the step (3) is 5: 1-50: 1.
preferably, the amine-rich organic template in step (3) is one or more of hyperbranched polyamide, polylysine and chitosan.
When the amine-rich organic matter template agent is polylysine, the processes of the steps (2) and (3) are as follows: dissolving sodium lauroyl sarcosinate in citric acid/sodium citrate (0.1M, pH 5.2) buffer solution, and adding epsilon-polylysine into the solution to generate polylysine/sodium lauroyl sarcosinate composite micelles; fe prepared in the step (1)3Adding O4 into citric acid/sodium citrate buffer solution, and adding polylysine/sodium lauroylsarcosine composite micelle solution; sequentially adding tetraethyl orthosilicate and APTES (3-aminopropyltriethoxysilane), continuously stirring, transferring the product into a high-temperature reaction kettle, and reacting for 8 hours at the temperature of 80 ℃ to obtain the magnetic ferroferric oxide/polylysine mesoporous silica composite blood lead ion scavenger.
The magnetic ferroferric oxide/aminated mesoporous silica composite blood lead ion scavenger prepared by the preparation method of the blood lead ion scavenger capable of safely entering and exiting red blood cells is provided.
Preferably, the composite material is a biocompatible material, the hemolysis rate is 0.5-5%, the activated partial thromboplastin time is 12-30 s, the plasma prothrombin time is 8-20 s, and the thrombin time is 8-20 s.
The magnetic ferroferric oxide/aminated mesoporous silica composite blood lead ion scavenger prepared by the preparation method of the blood lead ion scavenger capable of safely entering and exiting red blood cells is applied to adsorption removal of lead ions in the red blood cells.
The mechanism process of adsorbing and removing the lead ions in the red blood cells is that the composite material freely enters and exits the red blood cells, the lead-containing hemoglobin is captured through the complexation of the amino group in the mesoporous silicon dioxide structure on the lead ions, and the lead-containing hemoglobin is fixed through the mesoporous pore channel, so that the lead ions in the red blood cells are removed.
Has the advantages that: compared with the prior art, the invention has the following advantages:
(1) the invention utilizes the self-assembly performance of the amine-rich organic matter, namely, the amine-rich organic matter is used as a template agent in the material synthesis process, the processes of material synthesis, functional group modification and the like are realized through a 'one-step method', the forming process of the ferroferric oxide nuclear outer mesoporous molecular sieve is fused with the organic functional group modification process, and the aminated magnetic ferroferric oxide/aminated mesoporous silicon dioxide composite material is obtained.
(2) The prepared aminated magnetic ferroferric oxide/aminated mesoporous silica composite material has a regular mesoporous structure and highly dispersed organic functional groups, can freely enter and exit red blood cells, captures the lead-containing hemoglobin through the complexation of amino groups in the mesoporous silica structure on lead ions, and fixes the lead-containing hemoglobin through mesoporous channels, thereby achieving the purpose of removing the lead ions in the red blood cells. Overcomes the defects of unclear blood lead removal principle and low efficiency in current clinic and research.
Drawings
FIG. 1 shows Fe (a) prepared in example 13O4NPs, (b) transmission electron micrographs of MMS/P NPs;
FIG. 2 is a scanning electron micrograph of (a) MMS/P NPs and an elemental spectrum analysis chart of (b) MMS/P NPs;
FIG. 3 shows (a) Fe3O4The infrared spectra of NPs and (b) MMS/P NPs;
FIG. 4 is a graph showing nitrogen adsorption desorption isotherms and pore size distribution (internal adsorption) of MMS/P NPs before (a) and after (b) template removal;
FIG. 5 is a graph showing the in vitro clotting time of MMS/P NPs before and after anticoagulation treatment;
FIG. 6 is a graph showing the hemolysis rate of MMS/P NPs before and after loading with heparin;
FIG. 7 is a graph showing a comparison of the relative amounts of common ions in blood before and after adsorption of MMS/P NPs;
FIG. 8 is a TEM image of MMS/H NPs prepared in example 2;
FIG. 9 shows (a) a nitrogen adsorption/desorption curve of MMS/H NPs; (b) a schematic diagram of pore size distribution;
FIG. 10 is a schematic diagram showing the adsorption effect of MMS/H NPs on lead ions in real blood (rabbit blood and human blood);
FIG. 11 is a schematic representation of the process of the MMS/H NPs entering and exiting erythrocytes: (a) a single pure red blood cell, (b) a MMS/H NPs entering the red blood cell, (c) a TEM image of the red blood cell after separation of the MMS/H NPs from the red blood cell; (d-f) an enlarged view of erythrocytes after MMS/P NPs have entered erythrocytes;
FIG. 12 is a process of applying MMS/H NPs to lead toxicity model pig in vitro lead removal (i) intravenous intubation and (ii, iii) extracorporeal circulation;
fig. 13 is a routine blood test of lead poisoning model pigs before and after surgery (blood lead removal).
Detailed Description
The invention is further illustrated by the following figures and examples.
Example 1
1) Preparation of magnetic ferroferric oxide (Fe)3O4NPs)
Ferric chloride hexahydrate (1.35g,5mmol) was weighed out and dissolved well in 30mL of ethylene glycol solution. Sodium acetate (NaAc,3.6g) and polyethylene glycol-2000 (PEG-20001.0 g) were added with stirring. Stirring for 30min, transferring the solution into a high-temperature reaction kettle, and reacting at 200 ℃ for 72 h. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃.
2) Preparation of magnetic ferroferric oxide/rich aminated mesoporous silica composite material (MMS/P NPs) by using polylysine as template agent
100mg of sodium lauroyl sarcosinate was dissolved at room temperature in 10mL of citric acid/sodium citrate buffer (0.1M, pH 5.2), and 150. mu.L of ε -polylysine (20 wt%) was added to the solution. At this time, the solution immediately became an emulsion, and polylysine/sodium lauroyl sarcosinate complex micelles were formed.
150mg of Fe prepared above was added at room temperature3O4NPs were added to 50mL of citric acid/sodium citrate buffer solution (0.1M, pH 5.2) and 10mL of polylysine/sodium lauroyl sarcosinate complex micelle solution was added. Stirring for 20min, sequentially adding 500 μ L TEOS and 50 μ L APTES, stirring at room temperature for 2 hr, and transferring the product to high temperature reaction kettle to react at 80 deg.C for 8 hr. And after natural cooling, sequentially washing with deionized water and an ethanol solution for multiple times, and carrying out vacuum drying at 60 ℃ to obtain the magnetic ferroferric oxide/polylysine mesoporous silica composite material blood lead ion scavenger.
The size and morphology of the nanoparticles in the composite synthesis process were characterized by transmission electron microscopy, as shown in FIG. 1, Fe3O4NPs are uniformly dispersed, the size is about 120nm, and Fe is obtained under the general condition3O4The size of the NPs is about 50-300 nm. The size of the obtained MMS/P NPs is about 170nm, and the size of the obtained MMS/P NPs is about 50-500nm in general. Further, a scanning electron microscope image is adopted to represent the size and the morphology of the composite material, and as shown in fig. 2, the energy spectrum analysis result shows that the contents of iron (Fe), oxygen (O), silicon (Si), carbon (C) and nitrogen (N) elements are 51.66%, 25.20%, 4.65%, 17.23% and 1.25% in sequence. The above results all confirm the successful construction of MMS/P NPs.
The infrared spectrogram is used for characterizing the synthesis process of MMS/P NPs, as shown in FIG. 3. Before modification, as in FIG. (a), at 587cm-1Has obvious characteristic absorption peak corresponding to Fe-O peak. As shown in FIG. b, the reaction was further progressed at 1608cm-1And 1409cm-1A distinct characteristic absorption peak appears indicating the presence of an amide bond. This is achieved byOutside, 3000-3500 cm-1A broad peak corresponding to amino group (-NH)2-NH-). The presence of the amide and amino groups reflects that epsilon-polylysine is retained during the reaction. Furthermore, at 1094cm-1And 802cm-1And characteristic peaks are also simultaneously shown and respectively correspond to Si-O-Si and Si-O peaks, thereby proving the successful construction of the silicon dioxide shell layer.
The nitrogen adsorption and desorption curve is often used to reflect the pore structure of the mesoporous material. As shown in FIG. 4, the pore size of MMS/P NPs is about 5.1 nm.
As can be seen by comparing the coagulation time before and after anticoagulation treatment as shown in FIG. 5, compared with the blank group, due to the fact that a large number of amino groups in the material can be combined with protein with negative points on the surface, the time of MMS/P NPs APTT, PT and TT without anticoagulation treatment is shortened. After anticoagulation treatment, the APTT, PT and TT time is recovered to a normal level, which proves that the modified material can not influence the blood coagulation system. As shown in fig. 6, loading of heparin was effective in reducing the rate of hemolysis. The composite material is a biocompatible material, the hemolysis rate is 0.5-5%, the activated partial thromboplastin time is 12-30 s, the plasma prothrombin time is 8-20 s, and the thrombin time is 8-20 s.
3) Researching the adsorption behavior of the magnetic ferroferric oxide/aminated mesoporous silica composite material prepared in the step 2) on the blood lead:
putting a quantitative magnetic ferroferric oxide/rich aminated mesoporous silica composite material into a centrifuge tube, adding 3mL of leaded blood (0.6ppm), placing the centrifuge tube on a constant-temperature oscillator, oscillating at 37 ℃ for a fixed time, taking 2mL of supernatant, digesting the solution by a high-temperature digestion method, cooling and fixing the volume to 5 mL.
In order to know the adsorption condition of MMS/P NPs on lead in real blood, the real blood is adopted for adsorption test. As shown in fig. 7, the adsorption efficiency reached 52.64% after 30min of adsorption.
Example 2
1) Preparation of magnetic ferroferric oxide
Method for synthesizing ferroferric oxide nano particles (Fe) by adopting hydrothermal method3O4NPs). The specific method comprises the following steps: ferric trichloride hexahydrate (1.35g,5mmol) was weighed and dissolved well in 40mL of ethylene glycol solution. Sodium acetate (NaAc,1.8g) and sodium citrate trihydrate (Na) were added with stirring3Cit 0.2 g). Stirring is continued for 1h, and the solution is transferred to a high-temperature reaction kettle to react for 20h at 200 ℃. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃.
2) Preparation of magnetic ferroferric oxide/rich aminated mesoporous silicon dioxide composite material (MMS/H NPs) by using hyperbranched Polyamide (PAMAM) as template agent
Mixing Fe3O4NPs solution (3mg/mL,50mL) was mixed well with 9mL hydrazine hydrate and sonicated for 30 min. The mixture was transferred to a three-necked flask and 70mL of deionized water was added. 90mg TEOS were then added and stirring continued for 2h at 90 ℃. The resulting product was collected magnetically and redispersed in 150mL of deionized water containing 2.1mL hydrazine hydrate and 450mg CTAB. After another 30min of sonication, 0.6mL TEOS was added and stirred at 90 ℃ for 2 h. The obtained product is naturally cooled, washed by deionized water and ethanol solution for a plurality of times in sequence, and dried in vacuum at 60 ℃. And calcining the dried product at 550 ℃ for 5h in an air atmosphere to remove the CTAB template agent. The product obtained (100mg) was collected and dispersed in 150mL of deionized water containing 2.1mL of hydrazine hydrate and 450mg (0.45mmol) of PAMAM. Sonication for 30min, 0.6mL (2.88mmol) TEOS was added and stirring continued at 90 deg.C for 2 h. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃. And finally, soaking the product in 10mL of heparin solution (10mg/mL) for 12h, and performing vacuum drying at 60 ℃ to obtain the magnetic ferroferric oxide/hyperbranched polyamide mesoporous silica composite material blood lead ion scavenger.
As shown in FIG. 8, the MMS/H NPs have a particle size of about 300 nm. As shown in FIG. 9, the pore channel structure of the material is characterized by a nitrogen adsorption and desorption curve, and the BET surface area is about 248m2·g-1The pore size is about 24 nm.
3) Researching the adsorption behavior of the magnetic ferroferric oxide/aminated mesoporous silica composite material prepared in the step 2) on the blood lead:
putting a quantitative magnetic ferroferric oxide/rich aminated mesoporous silica composite material into a centrifuge tube, adding 3mL of leaded blood (0.6ppm), placing the centrifuge tube on a constant-temperature oscillator, oscillating at 37 ℃ for a fixed time, taking 2mL of supernatant, digesting the solution by a high-temperature digestion method, cooling and fixing the volume to 5 mL.
As shown in FIG. 10, the adsorption efficiency was 53% when the adsorbent addition amount reached 5mg/mL, and the adsorption efficiency was increased to 78% when the adsorbent addition amount was increased to 10 mg/mL. As shown in FIG. 11, MMS/H NPs can enter into erythrocytes, and can be separated by magnetic separation without affecting the morphology of erythrocytes.
4) Blood lead removal experiment for lead poisoning model pig by using blood lead removal instrument
A lead poisoning model of about 0.5ppm was established for pigs by the food poisoning method. The porcine extracorporeal circulation pathway (fig. 12) was then established as follows: pigs were anesthetized by injection of 3% sodium pentobarbital (1ml/kg) in the peritoneal cavity and maintained under anesthesia during the blood lead cleaning, with continuous injections of 0.9% NaCl and 2.5% sodium pentobarbital. The vein-calming blood vessel is connected with the blood lead cleaning device and returns arterial blood after passing through the magnetic separation device. The blood flow rate is 25ml/min, the circulation time is 50min, the amount of the adsorbent is 1g, the blood volume of the lead poisoning model pig is about 3L, and the volume ratio of the adsorbent to the blood can be adjusted between 1:10 and 10:1 g/L. The wound was sutured after surgery, and the blood routine and blood lead removal efficiency of lead poisoning model pigs before and after surgery were examined. Experimental results show that when the blood lead content of pig is 500ppb, the removal efficiency of blood lead can reach 75% after a cleaning process, and the removal rate of blood lead can be continuously improved through secondary treatment (the removal rate is calculated by respectively extracting blood before and after operation, and respectively marking the concentration before and after operation as C through inductive coupling plasma spectrometer detection0，C1By the formula E ═ C0-C1)/C0The removal efficiency of blood lead was calculated at 100%. In the formula, E is the blood lead removal efficiency (%), C0Blood lead concentration (ppb), C, before surgery1Post-operative blood lead concentration (ppb)). The adsorbent has good separation effect, shows high removal efficiency of the adsorbent and the blood lead removing instrument, and is cleaned simultaneouslyThe blood before and after the removal of lead ions shows that the blood inflammation is reduced (figure 13), which shows the safety and high efficiency of the blood adsorbent and the blood lead removal instrument in the process of removing the blood lead.
Example 3
1) Preparation of magnetic ferroferric oxide
Method for synthesizing ferroferric oxide nano particles (Fe) by adopting hydrothermal method3O4NPs). The specific method comprises the following steps: ferric trichloride hexahydrate (1.35g,5mmol) was weighed and dissolved well in 40mL of ethylene glycol solution. Sodium acetate (NaAc,1.8g) and sodium citrate trihydrate (Na) were added with stirring3Cit 0.2 g). Stirring is continued for 1h, and the solution is transferred to a high-temperature reaction kettle to react for 20h at 200 ℃. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃.
2) Preparation of magnetic ferroferric oxide/aminated mesoporous silica composite material by using chitosan as template agent
Mixing Fe3O4NPs solution (3mg/mL,50mL) was mixed well with 9mL hydrazine hydrate and sonicated for 30 min. The mixture was transferred to a three-necked flask and 70mL of deionized water was added. 90mg TEOS were then added and stirring continued for 2h at 90 ℃. The resulting product was collected magnetically and redispersed in 150mL of deionized water containing 2.1mL hydrazine hydrate and 450mg CTAB. After another 30min of sonication, 0.6mL TEOS was added and stirred at 90 ℃ for 2 h. The obtained product is naturally cooled, washed by deionized water and ethanol solution for a plurality of times in sequence, and dried in vacuum at 60 ℃. And calcining the dried product at 550 ℃ for 5h in an air atmosphere to remove the CTAB template agent. The resulting product (100mg) was collected and dispersed in 150mL of deionized water containing 2.1mL of hydrazine hydrate and 3.4g (0.0576mmol) of chitosan. Sonication for 30min, 0.6mL (2.88mmol) of TEOS was added, followed by stirring at 90 ℃ for 2 h. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃. And finally, soaking the product in 10mL of heparin solution (10mg/mL) for 12h, and performing vacuum drying at 60 ℃ to obtain the magnetic ferroferric oxide/chitosan mesoporous silica composite material blood lead ion scavenger.
Example 4
1) Preparation of magnetic ferroferric oxide
Method for synthesizing ferroferric oxide nano particles (Fe) by adopting hydrothermal method3O4NPs). The specific method comprises the following steps: ferric trichloride hexahydrate (1.35g,5mmol) was weighed and dissolved well in 40mL of ethylene glycol solution. Sodium acetate (NaAc,1.8g) and sodium citrate trihydrate (Na) were added with stirring3Cit 0.2 g). Stirring is continued for 1h, and the solution is transferred to a high-temperature reaction kettle to react for 24h at the temperature of 150 ℃. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃.
2) Preparation of magnetic ferroferric oxide/rich aminated mesoporous silica composite material (MMS/H NPs) by using hyperbranched polyamide as template agent
Mixing Fe3O4NPs solution (3mg/mL,50mL) was mixed well with 9mL hydrazine hydrate and sonicated for 30 min. The mixture was transferred to a three-necked flask and 70mL of deionized water was added. 90mg TEOS were then added and stirring continued for 2h at 90 ℃. The resulting product was collected magnetically and redispersed in 150mL of deionized water containing 2.1mL hydrazine hydrate and 450mg CTAB. After another 30min of sonication, 0.6mL TEOS was added and stirred at 60 ℃ for 5 h. The obtained product is naturally cooled, washed by deionized water and ethanol solution for a plurality of times in sequence, and dried in vacuum at 60 ℃. And calcining the dried product at 550 ℃ for 5h in an air atmosphere to remove the CTAB template agent. The product obtained (100mg) was collected and dispersed in 150mL of deionized water containing 2.1mL of hydrazine hydrate and 576 mg (0.576mmol) of PAMAM. Sonication for 30min, 0.6mL (2.88mmol) TEOS was added and stirring continued at 90 deg.C for 2 h. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃. And finally, soaking the product in 10mL of heparin solution (10mg/mL) for 12h, and performing vacuum drying at 60 ℃ to obtain the magnetic ferroferric oxide/hyperbranched polyamide mesoporous silica composite material blood lead ion scavenger.
Example 5
1) Preparation of magnetic ferroferric oxide
Method for synthesizing ferroferric oxide nano particles (Fe) by adopting hydrothermal method3O4NPs). The specific method comprises the following steps: ferric trichloride hexahydrate (1.35g,5mmol) was weighed and dissolved well in 40mL of ethylene glycol solution. Adding acetic acid under stirringSodium (NaAc,1.8g) and sodium citrate trihydrate (Na)3Cit 0.2 g). Stirring is continued for 1h, and the solution is transferred to a high-temperature reaction kettle to react for 10h at 220 ℃. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃.
2) Preparation of magnetic ferroferric oxide/aminated mesoporous silica composite material by using hyperbranched polyamide as template agent
Mixing Fe3O4NPs solution (3mg/mL,50mL) was mixed well with 9mL hydrazine hydrate and sonicated for 30 min. The mixture was transferred to a three-necked flask and 70mL of deionized water was added. 90mg TEOS were then added and stirring continued for 2h at 90 ℃. The resulting product was collected magnetically and redispersed in 150mL of deionized water containing 2.1mL hydrazine hydrate and 450mg CTAB. After another 30min of sonication, 0.6mL TEOS was added and stirred at 90 ℃ for 1 h. The obtained product is naturally cooled, washed by deionized water and ethanol solution for a plurality of times in sequence, and dried in vacuum at 60 ℃. And calcining the dried product at 550 ℃ for 5h in an air atmosphere to remove the CTAB template agent. The product obtained (100mg) was collected and dispersed in 150mL of deionized water containing 2.1mL of hydrazine hydrate and 450mg of PAMAM. Sonication was carried out for 30min, 0.6mL TEOS was added, and stirring was continued at 90 ℃ for 2 h. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃. And finally, soaking the product in 10mL of heparin solution (10mg/mL) for 12h, and performing vacuum drying at 60 ℃ to obtain the magnetic ferroferric oxide/hyperbranched polyamide mesoporous silica composite material blood lead ion scavenger.
Example 6
1) Preparation of magnetic ferroferric oxide
Method for synthesizing ferroferric oxide nano particles (Fe) by adopting hydrothermal method3O4NPs). The specific method comprises the following steps: ferric trichloride hexahydrate (1.35g,5mmol) was weighed and dissolved well in 40mL of ethylene glycol solution. Sodium acetate (NaAc,1.8g) and sodium citrate trihydrate (Na) were added with stirring3Cit 0.2 g). Stirring is continued for 1h, and the solution is transferred to a high-temperature reaction kettle to react for 20h at 200 ℃. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃.
2) Using hyperbranched polyamide as a moldPreparation of magnetic ferroferric oxide/rich aminated mesoporous silica composite material of plate agent Fe3O4NPs solution (3mg/mL,50mL) was mixed well with 9mL hydrazine hydrate and sonicated for 30 min. The mixture was transferred to a three-necked flask and 70mL of deionized water was added. 90mg TEOS were then added and stirring continued for 2h at 90 ℃. The resulting product was collected magnetically and redispersed in 150mL of deionized water containing 2.1mL hydrazine hydrate and 450mg CTAB. After another 30min of sonication, 0.6mL TEOS was added and stirred at 90 ℃ for 2 h. The obtained product is naturally cooled, washed by deionized water and ethanol solution for a plurality of times in sequence, and dried in vacuum at 60 ℃. And calcining the dried product at 550 ℃ for 5h in an air atmosphere to remove the CTAB template agent. The product obtained (100mg) was collected and dispersed in 150mL of deionized water containing 2.1mL of hydrazine hydrate and 57.6mg (0.0576mmol) of PAMAM. Sonication for 30min, 0.6mL (2.88mmol) TEOS was added and stirring continued at 90 deg.C for 2 h. After natural cooling, washing with deionized water and ethanol solution for multiple times in sequence, and vacuum drying at 60 ℃. And finally, soaking the product in 10mL of heparin solution (10mg/mL) for 12h, and performing vacuum drying at 60 ℃ to obtain the magnetic ferroferric oxide/hyperbranched polyamide mesoporous silica composite material blood lead ion scavenger.

Claims (6)

1. The application of a blood lead ion scavenger capable of safely entering and exiting erythrocytes in the preparation of a reagent for adsorbing and removing lead ions in erythrocytes is characterized in that the blood lead ion scavenger capable of safely entering and exiting erythrocytes is a magnetic ferroferric oxide/aminated mesoporous silica composite material blood lead ion scavenger, the process of adsorbing and removing lead ions in erythrocytes is that the composite material freely enters and exits erythrocytes, lead-containing hemoglobin is captured through the complexation of amino groups in a mesoporous silica structure on the lead ions, and the lead-containing hemoglobin is fixed through a mesoporous channel, so that the lead ions in the erythrocytes are removed;
the preparation method comprises the following steps:
(1) weighing ferric trichloride hexahydrate, dissolving the ferric trichloride hexahydrate in glycol solution, adding sodium acetate and sodium citrate trihydrate, or continuing to react with polyethylene glycol to obtain Fe3O4A solution;
(2) fe prepared in the step (1)3O4Uniformly mixing the solution and hydrazine hydrate, carrying out ultrasonic treatment, adding tetraethyl orthosilicate, collecting the obtained product, re-dispersing the product into deionized water containing hydrazine hydrate and hexadecyl trimethyl ammonium bromide, carrying out ultrasonic treatment again, and adding tetraethyl orthosilicate for reaction;
(3) drying and calcining the product obtained by the final reaction in the step (2) to remove hexadecyl trimethyl ammonium bromide, collecting the obtained product, dispersing the product into deionized water containing hydrazine hydrate and an amine-rich organic matter template agent, adding tetraethyl orthosilicate for reaction after ultrasonic treatment, and continuing the reaction to obtain a product, namely the magnetic ferroferric oxide/amine-rich mesoporous silica composite material blood lead ion scavenger; the amine-rich organic template agent is hyperbranched polyamide or chitosan.
2. The method as claimed in claim 1, wherein the reaction temperature in step (1) is 150 ℃ to 220 ℃ and the reaction time is 10-24 h.
3. Use according to claim 1, wherein step (1) is Fe3O4Has a size of 50-300 nm.
4. The use of claim 1, wherein the reaction temperature of each addition of tetraethyl orthosilicate in steps (2) and (3) is 60-90 ℃ and the reaction time is 1-5 h.
5. The use according to claim 1, wherein the molar ratio of tetraethyl orthosilicate to the amine-rich organic templating agent in step (3) is 5: 1-50: 1.
6. The application of a blood lead ion scavenger capable of safely entering and exiting erythrocytes in the preparation of a reagent for adsorbing and removing lead ions in erythrocytes is characterized in that the blood lead ion scavenger capable of safely entering and exiting erythrocytes is a magnetic ferroferric oxide/aminated mesoporous silica composite material blood lead ion scavenger, the process of adsorbing and removing lead ions in erythrocytes is that the composite material freely enters and exits erythrocytes, lead-containing hemoglobin is captured through the complexation of amino groups in a mesoporous silica structure on the lead ions, and the lead-containing hemoglobin is fixed through a mesoporous channel, so that the lead ions in the erythrocytes are removed;
the preparation method comprises the following steps:
(1) weighing ferric trichloride hexahydrate, dissolving the ferric trichloride hexahydrate in glycol solution, adding sodium acetate and sodium citrate trihydrate, or continuing to react with polyethylene glycol to obtain Fe3O4A solution;
(2) dissolving sodium lauroyl sarcosine in citric acid/sodium citrate buffer solution, and adding an amine-rich organic matter template epsilon-polylysine into the solution to generate a polylysine/sodium lauroyl sarcosine composite micelle; the Fe prepared in the step (1) is added3O4Adding the mixture into citric acid/sodium citrate buffer solution, and adding polylysine/sodium lauroyl sarcosine composite micelle solution; and sequentially adding tetraethyl orthosilicate and APTES, continuously stirring, transferring the product into a high-temperature reaction kettle for reaction, and obtaining the magnetic ferroferric oxide/polylysine mesoporous silica composite material blood lead ion scavenger.
CN201810981030.3A 2018-08-27 2018-08-27 Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof Active CN109174033B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201810981030.3A CN109174033B (en) 2018-08-27 2018-08-27 Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201810981030.3A CN109174033B (en) 2018-08-27 2018-08-27 Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof

Publications (2)

Publication Number Publication Date
CN109174033A CN109174033A (en) 2019-01-11
CN109174033B true CN109174033B (en) 2021-07-27

Family

ID=64916222

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201810981030.3A Active CN109174033B (en) 2018-08-27 2018-08-27 Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof

Country Status (1)

Country Link
CN (1) CN109174033B (en)

Families Citing this family (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110251479B (en) * 2019-06-06 2021-10-19 南京师范大学 Erythrocyte membrane-wrapped bionic blood hexavalent chromium reduction remover/magnetic nano motor and preparation method and application thereof
CN110755690A (en) * 2019-11-21 2020-02-07 南京鼓楼医院 Preparation method of mesoporous silica in-situ doped acrylic resin bone cement composite material for enhancing sustained release capacity of drugs
CN113680333A (en) * 2021-09-09 2021-11-23 商洛学院 Magnetic hyperbranched adsorbent for removing heavy metal ions and preparation method thereof

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101193962A (en) * 2005-01-14 2008-06-04 卡尔斯特里姆保健公司 Amine polymer-modified nanoparticulate carriers
CN102786061A (en) * 2012-07-20 2012-11-21 东华大学 Preparation method of hollow mesoporous silica nanoparticle
CN105964216A (en) * 2016-05-12 2016-09-28 西安建筑科技大学 Preparation and application of amino modified Fe3O4@SiO2@mSiO2 composite particles with mesoporous structure
CN106977746A (en) * 2017-03-28 2017-07-25 南京师范大学 Ultrabranching polyamide or derivatives thereof mesoporous molecular sieve composite material, its preparation method and application
CN107790075A (en) * 2016-09-06 2018-03-13 南京理工大学 A kind of magnetic mesoporous SiO of nucleocapsid shell structure2The preparation method of nano-particle

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101193962A (en) * 2005-01-14 2008-06-04 卡尔斯特里姆保健公司 Amine polymer-modified nanoparticulate carriers
CN102786061A (en) * 2012-07-20 2012-11-21 东华大学 Preparation method of hollow mesoporous silica nanoparticle
CN105964216A (en) * 2016-05-12 2016-09-28 西安建筑科技大学 Preparation and application of amino modified Fe3O4@SiO2@mSiO2 composite particles with mesoporous structure
CN107790075A (en) * 2016-09-06 2018-03-13 南京理工大学 A kind of magnetic mesoporous SiO of nucleocapsid shell structure2The preparation method of nano-particle
CN106977746A (en) * 2017-03-28 2017-07-25 南京师范大学 Ultrabranching polyamide or derivatives thereof mesoporous molecular sieve composite material, its preparation method and application

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
Preparation and Properties of Janus Heparin-Loaded Ammoniated-Hollow Mesoporous Silica Nanomotors;Sisheng Hu et al.;《The Journal of Physical Chemistry C》;20180413;第122卷;第9681页左栏第5-6段、右栏第1-2段，9683页右栏第3段及Figure 2 *

Also Published As

Publication number Publication date
CN109174033A (en) 2019-01-11

Similar Documents

Publication Publication Date Title
CN109174033B (en) Blood lead ion scavenger capable of safely entering and exiting red blood cells and preparation method and application thereof
AU702073B2 (en) Pharmaceutical composition containing selected lanthanum carbonate hydrates
EP0600347B1 (en) Process for the selective elemination of anorganic phosphate from liquids by adsorptionmaterials modified with polymuclear metal oxide hydroxides
CN105195111B (en) Preparation method of polyethyleneimine-modified shitosan coated magnetic nano-cellulose adsorbent and products thereof
US7135156B2 (en) Method for processing a zirconium oxide composition in crystalline form
EP1745793A1 (en) Adsorbent for oral administration, therapeutic or preventive agent for kidney disease, and therapeutic or preventive agent for liver disease
CN104984736A (en) Blood heavy metal ion adsorbent, preparation method thereof and blood perfusion device
CN107486156B (en) A kind of method of magnetic nanoparticle adsorbing separation bovine serum albumin(BSA)
CN110251479B (en) Erythrocyte membrane-wrapped bionic blood hexavalent chromium reduction remover/magnetic nano motor and preparation method and application thereof
CN109621912A (en) A kind of coating method of blood perfusion acticarbon
CN104402984B (en) A kind of preparation method of the iron protein succinylate of high amount of iron load
CN110665465A (en) Magnetic covalent organic framework material for glycopeptide enrichment and preparation method and application thereof
Zhang et al. Treatment of hyperphosphatemia based on specific interactions between phosphorus and Zr (iv) active centers of nano-MOFs
CN108079974A (en) A kind of preparation method and adsorbent equipment of western blot polymeric sorbent
CN109172908B (en) Blood lead removing instrument for removing blood lead in vitro
Duan et al. A Ce-MOF@ polydopamine composite nanozyme as an efficient scavenger for reactive oxygen species and iron in thalassemia disease therapy
CN106963735A (en) A kind of threonine chelated iron nano liposome and its application
JP6200950B2 (en) Heat-resistant nanoparticle preparation and related methods
Wadsworth et al. A study of the coagulation of the blood; the chemical reactions underlying the process
CH638401A5 (en) Process for the preparation of haemoglobin preparations having an increased oxygen delivery
CN109913200A (en) A kind of nano material and preparation method thereof with magnetism and fluorescent dual function
RU2516961C1 (en) Magnetically controlled sorbent for endo- and exotoxin elimination in humans
CN111892670A (en) Modified cation exchange resin and preparation method and application thereof
CN1431044A (en) Adsorbent based on montmorillonite for purifying blood and its preparing method
CN104689698B (en) A kind of method and apparatus of use bodied ferric sulfate absorption cleaning hydrogen sulfide

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant