WO2018143787A1 - Physiologically active substance carrier - Google Patents

Abstract

The present invention relates to a physiologically active substance carrier, comprising: a physiologically active substance; and porous silica particles supporting the physiologically active substance and having a plurality of pores with a diameter of 5-100 nm, wherein the porous silica particles have particular physical properties, can deliver all various drugs by a supported amount in a sustained manner, and can be parenterally administered.

Description

Bioactive substance carrier

The present invention relates to a bioactive substance carrier.

Drug delivery system refers to a medical technology that can efficiently deliver the required amount of drugs, such as proteins, nucleic acids, or other small molecules by minimizing the side effects and maximizing efficacy and effects of existing drugs. This technology, which saves the cost and time required for the development of new drugs, has recently become one of the cutting-edge technologies that create new added value in the pharmaceutical industry, combined with nanotechnology. The company has invested in the development of drug delivery systems along with the development of new drugs, mainly by companies and companies.

To date, viral genes, recombinant proteins, liposomes, cationic polymers, and various types of nanoparticles and nanomaterials have been used for drug delivery into animal cells. However, many cationic liposomes and cationic polymers have been found to be unsuitable because of their high toxicity to cells for clinical applications. In addition, a method of chemically modifying the main chain of the nucleic acid has been attempted for stable cell membrane penetration of the nucleic acid. However, this method is not suitable for clinical applications because it is expensive, time consuming, and requires labor intensive processes. As a significant attempt, drug delivery systems (DDS) have been developed that utilize various types of nanoparticles, including quantum dots, magnetic particles, or gold nanoparticles. For example, there has been a related study such as "image diagnosis drug carrier using porous silicon particles and its manufacturing method (Korean Patent Publication No. 2010-0117433)". However, these particles have a disadvantage in that they are toxic to cells, have a structure which is not easy to introduce biopolymers such as nucleic acids, and have low efficiency of introduction into cells.

Efficient delivery systems are needed for the study of the function of bioactive substances in cells or for intracellular delivery. However, the development of a universal delivery system capable of delivering a wide range of bioactive substances, a system capable of accommodating and delivering a large amount of drugs, and a system for releasing drugs in a sustained manner is still under development.

It is an object of the present invention to provide a general-purpose bioactive substance carrier.

It is an object of the present invention to provide a bioactive substance carrier capable of sustained release of various bioactive substances.

1. bioactive substances; And porous silica particles supporting the bioactive material and having a plurality of pores having a diameter of 5 nm to 100 nm.

The porous silica particles have a physiologically active substance carrier having t of 24 or more, wherein a ratio of absorbance of Equation 1 is 1/2:

[Equation 1]

A _t / A ₀

Wherein A ₀ is the absorbance of the porous silica particles measured by placing 5 ml of the 1 mg / ml suspension of the porous silica particles into a cylindrical permeable membrane having pores having a diameter of 50 kDa,

Outside the permeable membrane is in contact with the permeable membrane, 15ml of the same solvent as the suspension is located, the inside and the outside of the permeable membrane is stirred 60 rpm at 37 ℃ horizontal,

The pH of the suspension is 7.4,

A _t is the absorbance of the porous silica particles measured after t hours have elapsed since the measurement of A ₀ ).

2. The above 1, wherein the suspension is at least one bioactive substance carrier selected from the group consisting of PBS (phosphate buffered saline) and SBF (simulated body fluid).

3. In the above 1, wherein A _t of Equation 1 is measured in an environment in which the solvent outside the permeable membrane is replaced every predetermined period of time.

4. according to the above 1, wherein t is a biologically active substance carrier of 24 to 120.

5. In the above 1, wherein the porous silica particles are biodegradable bioactive material carrier.

6. In the above 1, wherein the porous silica particles, the ratio of the absorbance of Equation 1 is 1/5 t is a bioactive material carrier of 70 to 120.

7. In the above 1, wherein the porous silica particles, the ratio of the absorbance of Equation 1 is 1/20 t is a bioactive material carrier of 130 to 220.

8. In the above 1, wherein the ratio of the absorbance and t of the physiologically active substance carrier of Pearson correlation coefficient of 0.8 or more.

9. In the above 1, wherein the pore diameter is 7nm to 30nm bioactive material carrier.

10. In the above 1, wherein the porous silica particles are spherical bioactive material carrier.

11. In the above 1, wherein the porous silica particles have a mean diameter of 150nm to 1000nm bioactive material carrier.

12. In the above 1, wherein the porous silica particles have a BET surface area of 200m ² / g to 700m ² / g bioactive material carrier.

13. In the above 1, wherein the porous silica particles BET surface area 300m ² / g to 450m ² / g bioactive material carrier.

14. In the above 1, wherein the porous silica particles volume of 0.7ml to 2.2ml bioactive material carrier.

15. The bioactive substance carrier according to 1 above, wherein the porous silica particles have a volume per gram of 1.0ml to 2.0ml.

16. The bioactive material carrier according to 1 above, wherein the porous silica particles have a hydrophilic substituent or a hydrophobic substituent on an outer surface or inside a pore.

17. The physiologically active substance carrier according to 1 above, wherein the porous silica particles have a positive or negative charge at a neutral pH at an external surface or inside a pore.

18. The bioactive material carrier according to 1 above, wherein the bioactive material is poorly soluble and the porous silica particles have a hydrophobic substituent on an outer surface or inside a pore.

19. The bioactive material carrier according to 1 above, wherein the bioactive material is poorly soluble, and the porous silica particles have a hydrophobic substituent inside the pores and a hydrophilic substituent on the outer surface.

20. The bioactive material carrier according to 1 above, wherein the bioactive material is negatively charged at neutral pH, and the silica particles are positively charged at neutral pH at the outer surface or the inside of the pore.

21. The bioactive material carrier according to 1 above, wherein the bioactive material is positively charged at neutral pH, and the silica particles are negatively charged at neutral pH at the outer surface or inside the pore.

22. bioactive substances; And spherical porous silica particles carrying the physiologically active substance and having a particle diameter of 150 nm to 500 nm and a plurality of pores having a diameter of 7 nm to 30 nm.

The porous silica particles are physiologically active substance carrier having t of 24 to 120, where the ratio of absorbance of Equation 1 is 1/2:

[Equation 2]

A _t / A ₀

Wherein A ₀ is the absorbance of the porous silica particles measured by placing 5 ml of the 1 mg / ml suspension of the porous silica particles into a cylindrical permeable membrane having pores having a diameter of 50 kDa,

Outside the permeable membrane is in contact with the permeable membrane, 15ml of the same solvent as the suspension is located, the inside and the outside of the permeable membrane is stirred 60 rpm at 37 ℃ horizontal,

The suspension is PBS or SBF, pH is 7.4,

A _t is the absorbance of the porous silica particles measured after t hours have elapsed since the measurement of A ₀ ).

23. The bioactive material carrier according to the above 22, wherein the porous silica particles have a BET surface area of 300 m ² / g to 450 m ² / g and a volume per g of 1.0 ml to 2.0 ml.

24. The bioactive material carrier according to the above 22, wherein the bioactive material is poorly soluble and the porous silica particles have a hydrophobic substituent on an outer surface or inside a pore.

25. The bioactive material carrier according to the above 22, wherein the bioactive material is poorly soluble and the porous silica particles have a hydrophobic substituent inside the pores and a hydrophilic substituent on the outer surface.

26. The bioactive material carrier according to the above 22, wherein the bioactive material is negatively charged at neutral pH, and the silica particles are positively charged at neutral pH at the outer surface or inside the pore.

27. The physiologically active material carrier according to the above 22, wherein the bioactive material is positively charged at neutral pH, and the silica particles are negatively charged at neutral pH at the outer surface or inside the pore.

28. A method of preparing a bioactive substance carrier comprising contacting porous silica particles with a bioactive substance in a solvent.

29. The above-mentioned 28, wherein the solvent is water, chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride, 1,2-di Bromoethane, acetone, methyl isobutyl ketone, cyclohexanone, benzene, toluene, xylene, N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide, N-methylpi At least one member selected from the group consisting of rolidone, methanol, ethanol, propanol, butanol, PBS, SBF, borate-buffered saline and tris-buffered saline.

31. The method of 28 above, wherein the weight ratio of the porous silica particles and the bioactive material is 1: 0.05 to 0.8.

31. A method of delivering a physiologically active substance comprising parenterally administering the drug carrier of any one of 1 to 27 above to an individual.

32. The method of above 31, wherein the parenteral administration is intraorbital, intraocular, infusion, intraarterial, intraarticular, intracardiac, dermal, intramuscular, intraperitoneal, intrapulmonary, intramedullary, intrasternal, vertebral, intrauterine , Intravenous, subarachnoid, subcapsular, subcutaneous, transmucosal or intranasal administration.

In the bioactive substance carrier of the present invention, porous silica particles carrying a bioactive substance may be slowly degraded in vivo to deliver the drug in a sustained manner.

In the bioactive substance carrier of the present invention, the porous silica particles carrying the bioactive substance are completely decomposed in vivo, thereby completely delivering the supported bioactive substance to the living body.

The bioactive substance carrier of the present invention can be parenterally administered.

The bioactive substance carrier of the present invention can deliver various drugs in a sustained release.

1 is a micrograph of porous silica particles according to an embodiment of the present invention.

2 is a micrograph of porous silica particles according to an embodiment of the present invention.

Figure 3 is a micrograph of the small pore particles during the manufacturing process of the porous silica particles according to an embodiment of the present invention.

Figure 4 is a micrograph of the small pore particles according to an embodiment of the present invention.

Figure 5 is a micrograph of the pore diameter of the porous silica particles according to an embodiment of the present invention.

DDV (Degradable Delivery Vehicle) is the particle of the embodiment, the number in parenthesis means the diameter of the particle, the number of subscripts means the pore diameter. For example, DDV 200 ₁₀ refers to a particle of an embodiment having a particle diameter of 200 nm and a pore diameter of 10 nm.

Figure 6 is a micrograph to confirm the biodegradability of the porous silica particles according to an embodiment of the present invention.

7 is a tube having a cylindrical permeable membrane according to one example.

8 is a result of decreasing absorbance over time of porous silica particles according to an embodiment of the present invention.

9 is a result of decreasing absorbance for each particle diameter over time of the porous silica particles according to an embodiment of the present invention.

10 is a result of decreasing absorbance for each pore diameter of porous silica particles according to an embodiment of the present invention over time.

FIG. 11 is a result of decreasing absorbance for each pH of the environment over time of porous silica particles according to one embodiment of the present invention.

12 is a result of decreasing absorbance over time of the porous silica particles according to an embodiment of the present invention.

13 to 17 is a degree of release over time of the bioactive material supported on the porous silica particles according to an embodiment of the present invention.

18 is a tube for confirming the release of a bioactive material according to one example.

19 to 25 are the degree of release over time of the bioactive material supported on the porous silica particles according to an embodiment of the present invention.

FIG. 26 is a photograph of a Cas9 protein supported on porous silica particles according to an embodiment of the present invention and transferred into cells. FIG.

FIG. 27 is a micrograph showing the release of siRNA in mice by supporting siRNA on porous silica particles according to an embodiment of the present invention. FIG.

Bioactive substance carrier of the present invention is a bioactive substance; And porous silica particles supporting the bioactive material and having a plurality of pores having a diameter of 5 nm to 100 nm.

Bioactive substances

A bioactive substance is a bioactive substance / biofunction modulator that is supported on porous silica particles and can be delivered to an individual and exhibit activity. The bioactive substance has direct or indirect, therapeutic, physiological and / or pharmacological effects on human or animal organisms. It can be a therapeutically active agent that can provide.

The therapeutically active agent may be, for example, a general medicine, drug, prodrug or target group, or a drug or prodrug comprising the target group.

Therapeutic active agents include, for example, cardiovascular drugs, in particular antihypertensive agents (eg calcium channel blockers, or calcium antagonists) and antiarrhythmic agents; Congestive heart failure drugs; Muscle contractors; Vasodilators; ACE inhibitors; diuretic; Deoxidation dehydratase inhibitors; Cardiac glycosides; Phosphodiesterase inhibitors; Blockers; β blockers; Sodium channel blockers; Potassium channel blockers; β-adrenergic agonists; Platelet inhibitors; Angiotensin II antagonists; Anticoagulants; Thrombolytics; Bleeding drugs; Anemia treatments; Thrombin inhibitors; Antiparasitic agents; Antibacterial agents; Anti-inflammatory agents, in particular nonsteroidal anti-inflammatory agents (NSAIDs), more particularly COX-2 inhibitors; Steroidal anti-inflammatory agents; Prophylactic anti-inflammatory agents; Anti-glaucoma; Mast cell stabilizer; Acid aids; Drugs affecting the respiratory system; Allergic rhinitis; Alpha-adrenergic antagonists; Corticosteroids; Chronic obstructive pulmonary disease; Xanthine-oxidase inhibitors; Anti-arthritis agents; Gout treatments; Potent drugs and potent drug antagonists; Anti-TB drugs; Antifungal agents; Antiprotozoa; helminthic; Antiviral agents, in particular against respiratory antiviral agents, herpes, cytomegalovirus, human immunodeficiency virus and hepatitis infections; Leukemia and Kaposi's sarcoma therapy; Opioids including pain management agents, in particular anesthetics and analgesics, opioid receptor agonists, opioid receptor partial agonists, opioid antagonists, opioid receptor mixed agonists-antagonists; Neuroleptics; Sympathetic nervous system; Adrenaline antagonists; Drugs affecting neurotransmitter absorption and release; Anticholinergic agents; Anti-hemorrhagic agents; Prophylactic or therapeutic agents for radiation or chemotherapy effects; Adipogenic agents; Fat reducing agents; Anti-obesity agents such as lipase inhibitors; Sympathetic stimulants; Gastric ulcer and inflammation therapeutic agents such as proton pump inhibitors; Prostaglandins; VEGF inhibitors; Antihyperlipidemic agents, in particular statins; Drugs that affect the central nervous system (CNS), such as antipsychotic, antiepileptic and antiseizure agents (anticonvulsants), psychoactive agents, stimulants, anti-anxiety and hypnotics; Antidepressants; Anti-Parkinson's; Hormones such as sexual hormones and fragments thereof; Growth hormone antagonists; Gonadotropin releasing hormone and analogs thereof; Steroid hormones and antagonists thereof; Selective esterogen modulators; Growth factor; Antidiabetic agents such as insulin, insulin fragments, insulin analogues, glucagon-like peptides and hypoglycemic agents; H1, H2, H3 and H4 antihistamines; Peptides, proteins, polypeptides, nucleic acids and oligonucleotide drugs; Analogs, fragments and variants such as natural proteins, polypeptides, oligonucleotides and nucleic acids; Drugs used to treat migraine headaches; Asthma medicine; Cholinergic antagonists; Glucocorticoids; Androgen; Anti-androgens; Inhibitors of adrenocorticoid biosynthesis; Therapeutic agents for osteoporosis, such as biphosphonates; Antithyroid agents; Sunscreens, sunscreens and filters; Cytokine antagonists; Antitumor agents; Anti-alzheimer's agents; HMGCoA reductase inhibitors; Fibrate; Cholesterol absorption inhibitors; HDL cholesterol synergists; Triglyceride reducing agents; Anti-aging or anti-wrinkle agents; Precursor molecules for the development of hormones; Proteins such as collagen and elastin, antibacterial agents; Anti-acne medications; Antioxidants; Hair treatments and skin lightening agents; Sunscreens, sunscreens and filters; Variants of human apolipoproteins; Precursor molecules for the development of hormones; Proteins and peptides thereof; amino acid; Plant extracts such as grape seed extract; DHEA; Isoflavones; Nutrients including vitamins, phytosterols and iridoid glycosides, sesquiterpene lactones, terpenes, phenol glycosides, triterpenes, hydrquinone derivatives, phenylalkanones; Antioxidants such as retinoids, including retinol and other retinic acids and coenzyme Q10; Omega-3-fatty acid; Glucosamine; Nucleic acids, oligonucleotides, antisense drugs; enzyme; Coenzyme; Cytokine analogs; Cytokine agonists; Cytokine antagonists; Immunoglobulins; Antibodies; Antibody medicines; Gene therapy agents; Lipoprotein; Erythropoietin; vaccine; Allergy / asthma, arthritis, cancer, diabetes, growth disorders, cardiovascular disease, inflammation, immune disorders, baldness, pain, eye disease, epilepsy, gynecological disorders, CNS disease, viral infections, bacterial infections, parasitic infections, Gl disease, obesity and Small molecule therapeutic agents for the treatment or prevention of human and animal diseases such as blood diseases, but are not limited thereto.

The therapeutically active agent is for example erythropoietine (EPO). Cytokines such as thrombopoietine, interleukin (including IL-1 to IL-17), insulin, insulin-like growth factors (including IGF-1 and IGF-2), epidermal growth factors factor (EGF)), transforming growth factor (including TGF-alpha and TGF-beta), human growth hormone, transferrine, low density lipoprotein, high density lipoprotein (high density) lipoprotein, leptine, VEGF, PDGF, ciliary neurotrophic factor, prolactine, adrenocorticotropic hormone (ACTH), calcitonine, human chorionic gonadotropin (chrorionic gonadotropin), cortisol, estradiol, follicle stimulating hormone (FSH), thyroid-stimulating hormone (TSH), luteinizing hormone (LH) ), Progesterone one), testosterone (testosterone), toxins including lysine (ricine) and the like may be additional active agents.

The therapeutically active agent can be selected from the group of drugs for the treatment of oncological diseases and cellular or tissue alterations. Suitable therapeutic agents include alkyl sulfonates, for example busulfan, improsulfan, piposulfane, benzodepa, carboquone, metredepa. ), Alkylating agents such as arizidine, such as uredepa; Ethyleneimine such as altretamine, triethylene melamine, triethylene phosphoramide, triethylene thiophosphoramide, trimethylolmelamine and Methylmelamine; Chlorambucil, chlornaphazine, cyclophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydro So-called nitrogen mustards such as chloride (mechlorethaminoxide hydrochloride), melphalan, nomobichin, phenesterine, prednimustine, trofosfamide, uracil mustard nitrogen mustard; Nitroso urea compounds such as carmustine, chlorozotocin, potenmustine, lomustine, nimustine, and ranimustine compound); Dacarbazine, mannomustine, mitobranitol, mitoractol; Pipobroman; Sorafenib; An antineoplastic agent, including doxorubicin and cis-platimum and derivatives thereof and the like and any combination and / or derivatives of the foregoing.

Therapeutic actives include alaccinomycin, actinomycin, anthracycin, anthraceycin, azaserrin, bleomycin, cuctinomycin, carubicin, carubicin, Carzinophilin, chromomycin, ductinomycin, daunorbicin, 6-diazo-5-oxon-1-norycin (6-diazo-5-oxn- 1-norieucin, doxorubicin, epirubicin, mitomycin, mycophenolsaure, mogalumycin, olivomycin, peplomycin, peplomycin, Plicamycin, porfiromycin, poromycin, puromycin, streptonigrin, streptozocin, tubercidine, ubenimex, ubenimex, genostatin (zinostatin), zorubicin, aminoglycoside or It may be selected from Lien (polyene) or macrolide antibiotics (macrolid-antibiotics), and anti-viral agents and anti-bacterial agents such as the group including any combinations thereof and / or a derivative thereof.

Therapeutic actives include endostatin, angiostatin, interferon, platelet factor 4 (PF4), thrombospondin, transforming growth factor beta, metal Roperotinase-1. Tissue inhibitor of the metalloproteinases -1, -2, and -3 (TIMP-1, -2 and -3), TNP-470, marimastat, neovastat ( neovastat), BMS-275291, COL-3, AG3340, thalidomide, squalamin, combrestastatin, SU5416, SU6668, IFN- [alpha], EMD121974, CAI, IL-12 And radio-sensitizer drugs such as IM-862, steroidal or nonsteroidal anti-inflammatory drugs, or agents relating to angiogenesis, and combinations and / or derivatives thereof. have.

The therapeutically active agent may be selected from the group comprising nucleic acids, wherein the term nucleic acid is wherein at least two nucleotides are covalently linked to each other, for example to provide gene therapeutic or antisense effects. Oligonucleotides that are present. The nucleic acid preferably comprises phosphodiester bonds and also includes analogs with different backbones. Analogs include, for example, phosphoramide phosphorothioate, phosphorodithioate, O-methylphosphoroamidit-compound, and peptide-nucleic acid backbones. (peptide-nukleic acid-backbone) and skeletons thereof, and the like. Other analogues are those having an ionic backbone, having a nonionic backbone, and having a non-ribose-backbone. Nucleic acids having one or more carbocyclic sugars may be suitable as nucleic acids for use in the present invention. In addition to the selection of nucleic acids and nucleic acid analogs known in the art, any combination of naturally occurring nucleic acids and nucleic acid analogs or mixtures of nucleic acids and analogs may be used.

Therapeutic active agents are for example, everolimus, tacrolimus, sirolimus, mycophenololate-mofetil, rapamycin, paclitaxel ), Anti-mobility such as actinomycine D, angiopeptin, batimastate, estradiol, VEGF, statin and the like and derivatives and analogs thereof -migratory, anti-proliferative or immuno-suppresive, anti-inflammatory or re-endotheliating agents.

Therapeutic active agents include opioid receptor agonists and antagonists, compounds exhibiting agonist / antagonistic activity and compounds exhibiting partial action, such as morphine, depomorphine, etropin, diacetyl morphine, hydromorphine, oxymorphone, levorpa Knoll, methadone, levomethadyl, meperidine, fentanyl, serpentanyl, alfentanil, codeine, hydrocodone, oxycodone, thebaine, desormorphine, nicomorphine, dipropanoylmorphine, benzylmorphine, ethylmorphine, pettidine , Methadone, tramadol, dextrosepropoxyphene; Naloxone and naltrexone; Buprenorphine, nalbuphine, butorpanol, pentazocin and ethyl ketocyclylacin.

Therapeutic active agents and combinations thereof include heparin, synthetic heparin analogs (eg fondaparinux), hirudin, antithrombin III, drotrecogin alpha; Alteplase, plasmin, lysokinase, factor VIIa, prourokinase, urokinase, anistreplase, streptokinase fibrinolytics such as streptokinase; Platelet aggregation inhibitors such as acetylsalicylic acid (aspirine), ticlopidine, clopidogrel, abciximab, dextran and the like; Alclometasone, amcinonide, augmented betamethasone, beclomethasone, betamethasone, budesonide, cordesonide, clobetasol ( clobetasol, clocortolone, desonide, desoximetasone, dexamethasone, sexamethasone, fluocinolone, fluocinonide, fluandrenolide flurandrenolide, flunisolide, fluticasone, halcinonide, halobetasol, hydrocortisone, methylprednisolone, momethasone, Corticosteroids such as prednicarbate, prednisone, prednisolone, triamcinolone, and the like; Diclofenac, diflunisal, etodolac, fenoprofen, flurbiprofen, ibuprofen, indofetacin, indothathacin, ketoprofen (ketoprofen), ketorolac, meclofenamate, mefenamic acid, meloxacam, nabumetone, naproxen, oxaprozin, So-called non-steroidal anti-inflammatory drugs, such as pyroxicam, salsalate, sulindac, tolmetin, celecoxib and rofecoxib -inflammatory drugs) (NSAIDs); Cytostatic agents such as alkaloide and podophyllum toxin such as vinblastine and vincristine; Cytotoxic antibiotics such as daunorbicin, doxorubicin and other anthracycline and related substances, bleomycin, mitomycin, and the like; Antimetabolites such as folic acid analogs, purine analogs or pyrimidine analogs; Paclitaxel, docetaxel, sirolimus; Platinum compounds such as carboplatin, cisplatin or oxaliplatin; Amsacrin, irinotecan, iritincan, imatinib, topotecan, interferon-alpha 2a, interferon-alpha 2b, hydroxycarbide Miltefosine, pentostatin, porfimer, aldesleukin, bexaroten, tretinoin; Antiandrogens and antiestrogens; Quinarine type antiarrhythmic, quinidine, disopyramid, azmaline, prasmalium bitartrate, detajmium bitartrate, and the like Antiarrhythmics, which are type I antiarrhythmics; For example, lidocaine type antiarrhythmic agents, such as lidocaine, mexiletin, phenytoin, tocainid, etc .; For example, type Ic antiarrhythmic agents, such as propafenon and flecainid (acetate); Class II antiarrhythmic beta-receptor blockers such as metoprolol, esmolol, propranolol, atenolol, oxprenolol, and the like; Type III antiarrhythmic agents such as amiodarone and sotalol; Type IV antiarrhythmics such as diltiazem, verapami, gallopamil, and the like; Antiarrhythmic agents other than adenosine, orciprenaline, and ipratropium bromide; Agents that stimulate angiogenesis in myocardium such as vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), non-viral DNA, viral DNA, and endothelial growth factor ; FGF-1, FGF-2, VEGF, TGF; Antibiotics, monoclonal antibodies, anticalin; Stem cells, endothelial progenitor cells (EPC); Digitalis glycosides such as acetyl digoxin / methyldigoxin, digitoxin, digoxin, and the like; Cardiac glycosides such as ouabain and proscillaridin; Antihypertensive agents such as methyldopa, CNS active antiadrenergic substances which are imidazoline receptor agonists; Dihydropyridine-type calcium channel blockers such as nifedipine and nitrendipine; ACE inhibitors; Quinaprilate, cilazapril, moexipril, trandolapril, spirapril, imidaapril; Angiotensin II antagonists; Candesartancilexetil, valsartan, telmisartan, olmesartanmedoxomil, eprosartan; Peripherally active alpha-receptor blockers such as prazosin, urapidil, doxazosin, doazzosin, bunazosin, terazosin and indoramin ; Vasodilatators such as dihydralazine, diisopropylamine dichloraetate, minoxidil, nitroprusside sodium, and the like; Indapamide, co-dergocrine mesylate, dihydroergotoxin methanesulfonate, cicletanin, bosetan, fludrocortisone ( antihypertensive agents other than fludrocortisone); Phosphodiesterase inhibitors such as milrinon and enoximon, and especially dobutamine, ephinephrine, etilefrine, norfenefrine Adrenergic and dopaminergic substances such as norepinephrine, oxilofrine, dopamine, midodrine, poledrine, ameziniummetil, etc. and antihypertensive agents such as dopaminergic substances); Partial adrenoceptor agonists such as dihydroergotamine; Fibronectin, polylysine, ethylene vinyl acetate, TGFβ, PDGF, VEGF, bFGF, TNFα, NGF, GM-CSF, IGF-a, IL-1, IL-8, IL- 6, inflammatory cytokine such as growth hormone; Moreover, adhesive substances, such as cyanoacrylate, beryllium, and silica; And growth factors such as erythropoetin, corticotropin, gonadotropin, somatotropin, thyrotrophin, desmopressin, and ter Terlipressin, cytosine (pxytocin), cetrorelix, corticorelin, corticorelin, leuprorelin, triptorelin, gonadorelin, ganadorelin hormones such as (ganirelix), buserelin, buserelin, nafarelin, goserelin, and also regulatory peptides such as somatostatin and octreotid; Bone and cartilage stimulating peptides, recombinant human BMP-2 (rhBMP-2), bisphophonates (e.g. risedronate, pamidronate, ibandronate ), Recombinant BMPs such as zoledronic acid, clodronic acid, etidronic acid, alendronic acid, and tiludronic acid, disodium fluoro Bone morphogenetic proteins (BMPs), which are fluorides such as phosphite and sodium fluoride; Calcitonin, dihydrotachystyrol; Epidermal growth factor (EGF), platelet-derived growth factor (PDGF), fibrobast growth factor (FGFs), transforming growth factor (b) -b (TGFs-b)), transforming growth factors-a (TGFs-a), erythropoietin (EPO), insulin-like growth factor-I IGF-I)), insulin-like growth factor-II (IGF-II), interleukin-1 (IL-1), interleukin-2 (IL 2)), interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-a (TNF-a) Tumor necrosis factor-b (TNF-b), interferon-g (INF-g), colony stimulating factors (CSFs); Monocyte chemotactic protein, fibroblast stimulating factor 1, histamine, fibrin or fibrinogen, endothelin-1, angiotensin II, collagen, bro Bromocriptine, metysergide, methotrexate, carbon tetrachloride, thioacetamide and ethanol; In addition, β-lactamase-sensitive penicillin such as silver (ion), titanium dioxide, in particular benzyl penicillin (penicillin G), phenoxymethylphenicillin (penicillin V) and the like; Β-lactamase-resistent penicillin such as, for example, amoxicillin, ampicillin, and bacampicillin; Acylaminopenicillins such as mezlocillin and piperacillin; Cefazoline, cefuroxim, cefoxitin, cefotiam, cefaclor, cefadroxil, cefaroxin, cefalexin, loracarbef Carboxy penicillins such as cefixim, cefuroximaxetil, ceftibuten and cefpodoximproxetil; Aztreonam, ertapenem, meropenem; Β-lactamase inhibitors such as sulbactam and sulfamicillintosylate; Tetratracycline, such as doxycycline, minocycline, tetracycline, chlorotetracycline, oxytetracycline, etc .; Gentamicin, neomycin (neomycin), streptomycin, streptomycin, tobramycin, amikacin, netilmicin, paromomycin, parmymytin, framyceetin ), Aminoglycosides such as spectinomycin; Macrolide antibodies such as azithromycin, clarithromycin, erythromycin, erythromycin, roxithromycin, spiramycin, josamycin, and the like; Limcosamides such as clindamycin and lincomycin; For example ciprofloxacin, ofloxacin, moxifloxacin, norfloxacin, gatifloxacin, enoxacin, fleroxacin, levroxacin, levofloxacin gyrase inhibitors such as fluoroquinolone which is levofloxacin; Quinolones such as pipemidic acid; Sulfonamide, trimethoprim, sulfadiazine, sulffalene; Glycopeptide antibiotics such as vancomycin and teicoplanin; Polypeptide antibiotics such as polymycin, for example, colistin, polymyxin-b, for example, nitroimidazole derivatives such as metronidazole and tinidazole; Aminoquinolones such as chloroquine, mefloquine and hydroxychloroquine; Biguanids such as proguanil; Quinine alkaloids such as pyrimethamine and diaminopyrimidine; Amphenicol, such as chloramphenicol; Rifabutin, dapson, fusidic acid, fosfomycin, nifuratel, telithromycin, fusafungin, pentamidine diise Pentamidine diisethionate, rifampicin, taurolidin, atovaquon, linezolid; Acyclovir (aciclovir), ganciclovir, famciclovir, foscarnet, inosine- (dimefranol-4-acetamidobenzoate) (ionsine- (dimepranol- 4-acetanidobenzoate)), virus statics such as valganciclovir, valaciclovir, cidofovir, brivudin, etc .; Antiretroviral active substances such as lamivudine, zalcitabine, didanosine, zidanovudin, zidovudin, tenofovir, stavudin, and avacavir antiretroviral active ingredients (nucleoside analog reverse-transcriptase inhibitors and derivatives); Non-nucleoside analog reverse-transcriptase inhibitors; Amprenavir, indinavir, saquinavir, lopinavir, ritonavir, nelfinavir; Amantadine, ribavirine, zanamivir, oseltamivir or lamivudine, and any combinations and mixtures thereof.

Therapeutic active agents are, for example, alprazolam, amoxapine, betazepam, bromazepam, clolazepine, clovazam, clotiazepam, diazepam, lorazepam, flunitrazepam, flulazepam, lomezepam, Medazepam, nitrazepam, oxazepam, temazepam, maprotilin, myanserine, noritilline, risperidone, sertraline, trazodone, baloperidol, trimipramine malate fluoxetine, ondansetron, midazolam, chlor Promazine, haloperidol, triazolam, clozapine, fluoropromazine, flufenazine decanoate, fluanison, perfenazine, pimozide, prochlorperazine, sulfide, thiolidazine, paroxtin, Antidepressants, antipsychotics or anti-anxiety agents including cytaprolam, bupuropion, phenelzin, olanzapine, divalprox sodium and venlafaxine.

Therapeutic active agents are for example opioid receptor agonists and antagonists, compounds exhibiting mixed action / antagonist activity and compounds exhibiting partial action, such as morphine, depomorphine, etropin, diacetyl morphine, hydromorphine, oxymorphone , Levofanol, methadone, levomethadyl, meperidine, fentanyl, serpentanyl, alfentanil, codeine, hydrocodone, oxycodone, thebaine, desormorphine, nicomorphine, dipropanoylmorphine, benzylmorphine, ethylmorphine , Petidine, methadone, tramadol, dextrosepropoxyphene; Naloxone and naltrexone; Buprenorphine, nalbuphine, butorpanol, pentazocin and ethyl ketocyclylacin.

The therapeutically active agent can be, for example, tricyclic compounds including azothiopine, amitriptyline, pamotidine, promethazine, paroxatin, oxcabazapine and merthazapine.

Therapeutic active agents include, for example, antiacetic agents, including acetohexamide, chlorpropamide, glybenclide, glyclazide, glyphide, metformin, tolazamide, glyberid, glymepyride and tolbutamide It may be diabetes.

Therapeutic active agents are, for example, beclamid, carbamazepine, gafapentin, tiagabine, vigabatrin, topiramate, clonazepam, etotoin, metoin, methsimid, methylphenobabitone, ox Carbazepine, paramethadione, phenacemide, phenobarbitone, phenyloin, fenximide, primidone, sultiamine, phenytoin sodium, nitrofrantoin monohydrate, gabapentin, lamotrigine, zonisamide, ethoximide And valproic acid.

Therapeutic active agents include, for example, zolpidem tartrate, amylobarbitone, barbitone, butobabitone, pentobarbitone, brotizolam, carbromal, chlordiazepoxide, chlormethiazole, ethinamate, Hypnotics / sedatives and / or muscle relaxants, including meprobamate, metaquaalum, cyclobenzaprene, cyclobenzaprene, tizanidine, bacclifen, butalbital, zodiaclone, atraccurium, tubocurin and phenobarbital Can be.

Therapeutic active agents are, for example, amphotericin, butoconazole nitrate, clotrimazole, echonazol nitrate, fluconazole, flucitocin, griseoflubin, itraconazole, ketoconazole, myconazole, natamycin, nystatin, Sulfonazole nitrate, terconazole, thioconazole and undecenoic acid; Benzidazole, Clioquinol, Decoquinate, Diiodohydroxyquinoline, Diloxanide furoate, Dinitolamide, Furzolidone, Metronidazole, Nimorazol, Nitrofurazone, Ornidazole, Terbinafine, Clotritri Antifungal, antiprotozoal or antiparasitic agents, including mazol, chloroquine, mefloquine, itraconazole, pyrimethamine, prazicuantel, quinacrine, mebendazole and tinidazole.

Therapeutic active agents are, for example, antidepressants that can be used, for example, candesartan, hydralazine, clonidine, triamterin, felodipine, cappibrozil, fenofibrate, nifedical, prazosin, mecamylamine, doxazosin, dobutamine, and cilexetil Hypertension or heart treatment.

The therapeutically active agent may be, for example, an antimigraine agent comprising dihydroergotamine mesylate, ergotamine tartrate, methisurged malate, pizotifen malate and sumatrippan succinate.

The therapeutically active agent can be an antimuscarinic agent, including, for example, atropine, benzhexol, biferdene, etopropazine, hydroxyamine, mefenzolate bromide, oxybutynin, oxyphencyclimine and trophamide have.

Therapeutic active agents are, for example, aminoglutetimides, amsacrine, azathioprine, busulfan, chlorambucil, cyclosporin, dacarbazine, estramastine, etoposide, romastin, melphalan, mercaptopurine Antineoplastic agents (or immunosuppressive agents), including, metoclexate, mitomycin, mitotans, mitoxanthrone, procarbazine, tamoxifen citrate, testosteroltone, tacrolimus, mercaptopurine and sirolimus Can be.

Therapeutic active agents are for example bromocriptine mesylate, levodopa, tolcapone, lopinitrol, bromocriptine, hypoglycemic agents such as sulfonylurea biguanides, alpha-glucosidase inhibitors, Anti-Parkinson's agents, including thiazolidinediones, cabergoline, carbidopa and lisuride malate.

The therapeutically active agent may be an antithyroid agent, including, for example, carbazole and propithiouracil.

The therapeutically active agent can be, for example, a cardiac muscle contractor including amlinone, milnonone, digitoxine, enoxymon, lanatoside C and medigoxin.

The therapeutically active agent can be hypolipidemia or hyperlipidemia, including, for example, fenofibrate, clofibrate, probucol, egestimib and tocetrapib.

The therapeutically active agent can be an anti-inflammatory agent, including, for example, meoxycham, triamcinolone, chromoline, nedocromyl, hydroxychloroquine, montelukast, giluton, zapyrucast and meloxycamp.

Therapeutic active agents are, for example, pesofenadin, chloral hydrate, hydroxyzine, promethazine, cetyrazine, cimetidine, cclizin, meclizin, dimenhydrinate, loratabin, nizatabin and It may be an antihistamine including promethazine.

The therapeutically active agent may be an anti-ulcer agent including, for example, omeprazole, lansoprazole, pantoprazole and ranitidine.

The therapeutically active agent may be a diuretic including, for example, hydrochlorothiazide, amylolide, acetazolamide, furosemide, and torsemide.

The therapeutically active agent is, for example, a second occurrence, such as retinol, retinal, tretanoin (retinoic acid, retin-A), isotretinoin and alitretinoin, a second occurring retinoid, etretinate and its metabolite acitretin. Developmental retinoids; Retinoids, including third generation retinoids such as tazarotene, bexarotene, and adapalene.

The therapeutically active agent can be, for example, statins including atorvastatin, fluvastatin, lovastatin, nystatin, roschvastatin, pravastatin, olistat and simvastatin and / or derivatives thereof.

The therapeutically active agent may be, for example, a stimulant including amphetamine, phentermine, tyramine, eiffelrin, metaramimin, phenylephrine, dexamphetamine, dexfenfluramine, fenfluramine, nicotine, caffeine and marginol.

The therapeutically active agent may be a vasodilator including, for example, carvedilol, terazosin, pentolamin and menthol.

The therapeutically active agent can be, for example, an anti-alzheimer's agent including levetiracetam, levetiracetam and donepezil.

The therapeutically active agent may be, for example, an ACE inhibitor including benzapril, enalapril, ramipril, fosinopril sodium, ricinopril, minoxidil, isosorbide, lamprill and quinapril.

The therapeutically active agent may be, for example, a beta adrenergic receptor antagonist, including atenolol, timolol, pindolol, pronanol hydrochloride, bisprolol, esmolol, metoprolol succinate, metoprolol and metoprolol tartrate.

The therapeutically active agent may be, for example, angiotensin II antagonist, including rozatan.

The therapeutically active agent may be a platelet inhibitor, including, for example, Absiksimab, clopidrogel, tyropiban and aspirin.

Therapeutic active agents are, for example, tramadol, tramadol hydrochloride, allopurinol, calcitriol, cilostazol, soltalol, urasodiol bromperidol, dropperidol, flufenticsol decanoate, albuterol, albuterol Alcohols or phenols including sulfate, carisoprodol, clobetasol, rofinirol, labetalol and metocarbamol.

The therapeutically active agent can be, for example, ketones or esters including amioderon, fluticasone, spironolactone, prednisone, triazonedon, desoxymethasone, methyl prednisone, benzonatate nabumethone and buspyrone .

The therapeutically active agent may be an antiemetic agent, including, for example, metoclopramide.

The therapeutically active agent may be, for example, an eye treatment comprising dorzolamide, brimonidine, olopatadine, cyclopenttolate, pilocarpine and ecothioate.

The therapeutically active agent may be an anticoagulant or antithrombotic agent, including, for example, warfarin, enoxaparin and repyrudine.

The therapeutically active agent may be, for example, a gout treatment comprising probenesin and sulfinpyrazone.

The therapeutically active agent may be, for example, a COPD or asthma treatment comprising ypratropium.

The therapeutically active agent can be, for example, a treatment for osteoporosis, including raloxifene, pamidronate and risedronate.

The therapeutically active agent can be, for example, a cosmetic peptide comprising acetyl hexapeptide-3, acetyl hexapeptide-8, acetyl octapeptide and l-carnosine.

Therapeutic active agents include, for example, vaccines comprising toxoids (inactivated toxic compounds); Proteins, protein subunits and polypeptides; Polynucleotides such as DNA and RNA; Conjugates; Vaccines comprising saponins, virosomes, murine and organic adjuvants such as jostaxax.

Therapeutic active agents include, for example, coenzyme Q10 (or ubiquinone), ubiquinol or resveratrol; carotenoids such as α, β or γ-carotene, lycopene, lutein, zeaxanthin and astaxanthin; Phytonutrients such as lycopene, lutein and cyaxanthin; Omega-3 fatty acids, including linoleic acid, conjugated linoleic acid, docosahexaenoic acid (DHA) and ericosapentaenoic acid (EPA) and their glycerol-esters; Vitamin D (D2, D3 and derivatives thereof), vitamin E (α, β, γ, δ-tocopherol, or α, β, γ, δ-tocotrienol), vitamin A (retinol, retinal, retinoic acid and derivatives) Fat soluble vitamins including vitamin K (K1, K2, K3 and derivatives thereof), capric / caprylic triglycerides, folic acid, iron, niacin, glyceryl linoleate, omega 6 fatty acids, vitamin F, selenium, cyanocobalamin , Aloe vera, beta glucan, bisabool, camellia thea (green tea) extract, capric / caprylic triglyceride, gotu cola extract, cetearyl olivate, chlorophyll, orange oil, cocoyl proline, dicapryl ether , Disodium lauriminodipropionate tocopheryl phosphate (vitamin E phosphate), glycerin, glyceryl oleate, licorice extract, witch hazel extract, lactic acid, lecithin, lutein, macadamia seed oil, chamomile (chamomile) Extract, Evening Primrose Oil, Olive Leaf Extract, Rice Bran Oil, Avocado Oil, Sewage Extract, Pomegranate Sterol, Resveratrol, Rose Oil, Sandalwood Oil, Titanium Dioxide, Folic Acid, Glycerin, Glyceryl Linoleate (Omega 6 (Fat Vitamin F)), Vitamin A palmitate, grapeseed oil, halobetaazole, adenosine, adenosine triphosphate, alpha hydroxy acid, allantoin, hyaluronic acid and derivatives, isoleutrol, tranexamic acid, glycolic acid, arginine, ascorbyl glucosamine, ascorbyl Palmitate, salicylic acid, carnosic acid, alpha lipoic acid, gamma linolenic acid (GLA), panthenol, retinyl propionate, retinyl palmitate, furfuryl adenine, retinaldehyde, gripeptide, idebenone, dimethylaminoethanol (DMAE) , Niacinamide, beta-glucan, palmitoyl pentapeptide-4, palmitoyl oligopeptide / tetrapeptide-7, etocin, cerami , Phenylalanine, glucuronolactone, L-carnitine, hydroxyapatite, palmitoyl tripeptide-3, phospholine, zinc oxide, α-bisabolol, eugenol, silybin, soy isoflavone, catalpol, Pseudoguaianolides from rosenica acid, rosemarinic acid, rosemanol, silasilates, for example salicylic acid, salginine and salicylic acid, talaxasterol, α-lactocerol, isolaxetrol, taraxacoside, sere It may be a nutritional or cosmetic active agent, including mead, arbutin, gingerol, shogaol, hypericin, elastin, collagen and peptides thereof.

Porous silica particles

Silica particles according to the present invention (Porous Silica Particle, pSP) is a particle of silica (SiO ₂ ) material, and has a particle size of nano size.

Silica nanoparticles according to the present invention is a porous particle, having nano-sized pores.

Porous silica particles according to the present invention may carry a bioactive material on its surface and / or pores.

Porous silica particles according to the present invention is a biodegradable particle, it is possible to release the bioactive material as it is biodegraded in the body when administered to the body carrying a bioactive material.

As the porous silica particles are biodegraded, the bioactive material is released, and the porous silica particles according to the present invention may be slowly decomposed in the body so that the supported bioactive material may have sustained release. For example, t which becomes the ratio of the absorbance of following formula (1) 1/2 is 24 or more.

[Equation 1]

A _t / A ₀

Wherein A ₀ is the absorbance of the porous silica particles measured by placing 5 ml of the 1 mg / ml suspension of the porous silica particles into a cylindrical permeable membrane having pores having a diameter of 50 kDa,

Outside the permeable membrane is in contact with the permeable membrane, 15ml of the same solvent as the suspension is located, the inside and the outside of the permeable membrane is stirred 60 rpm at 37 ℃ horizontal,

The pH of the suspension is 7.4,

A _t is the absorbance of the porous silica particles measured after t hours have elapsed since the measurement of A ₀ ).

Equation 1 means that the rate at which the porous silica particles are degraded in an environment similar to the body.

Absorbance A ₀ , A _t in Equation 1 may be measured by putting porous silica particles and a suspension in a cylindrical permeable membrane and putting the same suspension outside the permeable membrane, as illustrated in FIG. 7.

The porous silica particles of the present invention are biodegradable, and can be slowly decomposed in suspension, 50 kDa in diameter corresponds to about 5 nm, and biodegradable porous silica particles can pass through a permeable membrane of 50 kDa in diameter, and a cylindrical permeable membrane is 60 rpm horizontal. Under stirring, the suspension can be mixed evenly and the degraded porous silica particles can come out of the permeable membrane.

The absorbance in Equation 1 may be measured, for example, in an environment in which the suspension outside the permeable membrane is replaced with a new suspension. The suspension may be one that is constantly replaced, one that may be replaced every period, and the period may be periodic or irregular. For example, within the range of 1 hour to 1 week, 1 hour interval, 2 hours interval, 3 hours interval, 6 hours interval, 12 hours interval, 24 hours interval, 2 days interval, 3 days interval, 4 days interval, 7 It may be replaced at day intervals, but is not limited thereto.

The ratio of the absorbance to 1/2 means that after t hours the absorbance is half of the initial absorbance, which means that approximately half of the porous silica particles are decomposed.

The suspension may be a buffer solution, for example, at least one selected from the group consisting of phosphate buffered saline (PBS) and simulated body fluid (SBF), and more specifically, PBS.

T in which the ratio of absorbance of Equation 1 according to the present invention is 1/2 is 24 or more, for example, t may be 24 to 120, for example, within the range of 24 to 96, 24 to 72, 30 to 70, 40 to 70, 50 to 65 and the like, but is not limited thereto.

The porous silica particles according to the present invention may be, for example, 70 to 140, where t is the ratio of absorbance of Equation 1 to 1/5, for example, 80 to 140, 80 to 120, and 80 to 80 within the above range. 110, 70 to 140, 70 to 120, 70 to 110 and the like, but is not limited thereto.

The porous silica particles according to the present invention may be, for example, 130 to 220 in which the ratio of the absorbance of Equation 1 is 1/20, for example, 130 to 200, 140 to 200, 140 to 140 within the above range. 180, 150 to 180, and the like, but is not limited thereto.

The porous silica particles according to the present invention may have a measured absorbance of 0.01 or less, for example, 250 or more, for example, 300 or more, 350 or more, 400 or more, 500 or more, 1000 or more, and the upper limit thereof is 2000. It may be, but is not limited thereto.

In the porous silica particles according to the present invention, the ratio of the absorbance of Formula 1 and t have a high positive correlation, for example, the Pearson correlation coefficient may be 0.8 or more, for example, 0.9 or more and 0.95 or more. have.

T in Equation 1 means how fast the porous silica particles decompose in an environment similar to the body, for example, the surface area, particle diameter, pore diameter, surface and / or inside the pores of the porous silica particles. It can be controlled by controlling the substituent, the degree of compactness of the surface, and the like.

For example, the surface area of the particles can be increased to reduce t, or the surface area can be reduced to increase t. The surface area can be adjusted by adjusting the diameter of the particles and the diameter of the pores. In addition, by placing substituents on the surface and / or within the pores, t can be increased by reducing the direct exposure of porous silica particles to the environment (such as solvents). In addition, by supporting the bioactive material on the porous silica particles and increasing the affinity between the bioactive material and the porous silica particles, it is possible to increase the t by reducing the direct exposure of the porous silica silica particles to the environment. In addition, the surface may be made more densely at the time of preparation of the particles to increase t. In the above, various examples of adjusting t in Equation 1 have been described, but are not limited thereto.

Porous silica particles according to the present invention may be, for example, spherical particles, but is not limited thereto.

Porous silica particles according to the present invention may have an average diameter of, for example, 150nm to 1000nm, for example within the above range, for example 150nm to 800nm, 150nm to 500nm, 150nm to 400nm, 150nm to 300nm, 150nm to 200nm It may be, but is not limited thereto.

The porous silica particles according to the present invention may have an average pore diameter of, for example, 1 nm to 100 nm, for example, within the above range, for example, 5 nm to 100 nm, 7 nm to 100 nm, 7 nm to 50 nm, 10 nm to 50 nm, 10 nm to It may be 30nm, 7nm to 30nm, but is not limited thereto. Having a large diameter as described above can carry a large amount of bioactive material, it is possible to carry a large size of the bioactive material.

The porous silica particles according to the present invention may have a BET surface area of, for example, 200 m ² / g to 700 m ² / g. For example, within the range of 200m ² / g to ^{700m 2 / g, 200m 2 /} g to ^{650m 2 / g, 250m 2 /} g to ^{650m 2 / g, 300m 2 /} g to 700m ² / g, 300m ² / g to 650m ² / g, 300m ² / g to 600m ² / g, 300m ² / g to 550m ² / g, 300m ² / g to 500m ² / g, 300m ² / g to 450m ² / g, etc. It is not limited to this.

Silica nanoparticles according to the present invention may have a volume per g, for example, 0.7 ml to 2.2 ml. For example, within the above range may be 0.7ml to 2.0ml, 0.8ml to 2.2ml, 0,8ml to 2.0ml, 0.9ml to 2.0ml, 1.0ml to 2.0ml and the like, but is not limited thereto. If the volume per gram is too small, the rate of decomposition may be too high, and excessively large particles may be difficult to manufacture or may not have an intact shape.

Porous silica particles according to the present invention may have hydrophilic substituents and / or hydrophobic substituents on the outer surface and / or inside the pores. For example, only hydrophilic substituents may exist on both the surface and inside of the pores, or only hydrophobic substituents may exist, hydrophilic substituents may exist on the surface or inside of the pores, hydrophobic substituents may exist on the surface, hydrophilic substituents on the surface, and hydrophobic substituents inside the pores. It may be present and vice versa.

The release of the bioactive material supported on the porous silica particles is mainly performed by the decomposition of the nanoparticles, and the interaction of the porous silica particles with respect to the bioactive material release environment is controlled by the control of the substituents. The rate of degradation may be controlled to control the release rate of the bioactive material. In addition, the bioactive material may be diffused and released from the nanoparticles, and the binding force of the bioactive material to the nanoparticles may be controlled by controlling the substituents. Active substance release can be controlled.

In addition, hydrophobic substituents are present inside the pores to enhance binding to poorly soluble (hydrophobic) bioactive substances, and the surface of the particles may be treated with hydrophilic substituents in view of ease of use and formulation. Do.

Hydrophilic substituents are, for example, hydroxyl groups, carboxy groups, amino groups, carbonyl groups, sulfhydryl groups, phosphate groups, thiol groups, ammonium groups, ester groups, imide groups, thiimide groups, keto groups, ether groups, indene groups, sulfonyl groups, polyethylene Glycol groups and the like, and the hydrophobic substituent is, for example, substituted or unsubstituted C1 to C30 alkyl group, substituted or unsubstituted C3 to C30 cycloalkyl group, substituted or unsubstituted C6 to C30 aryl group, substituted Or an unsubstituted C2 to C30 heteroaryl group, a halogen group, a C1 to C30 ester group, a halogen-containing group, or the like.

In addition, the porous silica particles according to the present invention may be one in which the outer surface and / or the inside of the pores are positively and / or negatively charged. For example, both the surface and the inside of the pore may be positively charged, or may be negatively charged, only the surface or the inside of the pore may be positively charged, or may be negatively charged, the surface may be positively charged, and the interior of the pore may be negatively charged. The reverse is also possible.

The charging may be, for example, by the presence of a cationic substituent or an anionic substituent.

The cationic substituent may be, for example, an amino group or other nitrogen-containing group as the basic group, and the anionic substituent may be, for example, a carboxy group (-COOH), sulfonic acid group (-SO ₃ H), thiol group (- SH) and the like, but is not limited thereto.

Likewise, the interaction of the porous silica particles with respect to the physiologically active substance releasing environment is controlled by the charging of the substituent so that the decomposition rate of the nanoparticles can be controlled to regulate the physiologically active substance release rate, and also, the physiological activity The material may be diffused from the nanoparticles and released, and the binding force of the bioactive material to the nanoparticles may be controlled by controlling the substituents, thereby controlling the release of the bioactive material.

In addition, the porous silica particles according to the present invention may be carried on the surface and / or the pores in addition to the support of the bioactive material, the transfer of the bioactive material to the target cell, the support of the material for other purposes, or other additional substituents. Substituents for such may be present, and may further include antibodies, ligands, cell permeable peptides, or aptamers bound thereto.

Substituents, charges, binders and the like within the aforementioned surfaces and / or pores may be added, for example, by surface modification.

Surface modification can be carried out, for example, by reacting a compound having a substituent to be introduced with the particles, which may be, for example, an alkoxysilane having a C1 to C10 alkoxy group, but is not limited thereto. The alkoxysilane has one or more alkoxy groups, and may have, for example, 1 to 3, and there may be a substituent to be introduced into a site where the alkoxy group is not bonded or a substituent substituted therewith.

Preparation of Porous Silica Particles

For example, the porous silica particles of the present invention may be prepared through a small pore particle preparation and a pore expansion process, and may be manufactured through a calcination process, a surface modification process, and the like, as necessary. If both the calcination and surface modification process has gone through may be surface modified after calcination.

The small pore particles may be, for example, particles having an average pore diameter of 1 nm to 5 nm.

Small pore particles can be obtained by adding a surfactant and a silica precursor to a solvent, stirring and homogenizing.

The solvent may be water and / or an organic solvent, and the organic solvent may be, for example, ethers such as 1,4-dioxane (particularly cyclic ethers); Halogenated hydrocarbons such as chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride and 1,2-dibromoethane; Acetone, methyl isobutyl ketone, γ-butyrolactone, 1,3-dimethyl-imidazolidinone, methyl ethyl ketone, cyclohexanone, cyclopentanone, 4-hydroxy-4-methyl-2-pentanone, etc. Ketones; Carbon-based aromatics such as benzene, toluene, xylene and tetramethylbenzene; Alkyl amides such as N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide and N-methylpyrrolidone; Alcohols such as methanol, ethanol, propanol and butanol; Ethylene glycol monoethyl ether, ethylene glycol monomethyl ether, ethylene glycol monobutyl ether, diethylene glycol monoethyl ether, diethylene glycol monomethyl ether, diethylene glycol monobutyl ether, propylene glycol monomethyl ether, propylene glycol monoethyl ether Glycol ethers (cellosolve) such as dipropylene glycol diethyl ether and triethylene glycol monoethyl ether; Dimethylacetamide (DMAc), N, N-diethylacetamide, dimethylformamide (DMF), diethylformamide (DEF), N, N-dimethylacetamide (DMAc), N-methylpyrrolidone ( NMP), N-ethylpyrrolidone (NEP), 1,3-dimethyl-2-imidazolidinone, N, N-dimethylmethoxyacetamide, dimethyl sulfoxide, pyridine, dimethyl sulfone, hexamethylphosphoamide , Tetramethylurea, N-methylcarrolactam, tetrahydrofuran, m-dioxane, P-dioxane, 1,2-dimethoxyethane and the like can be used. Specifically, alcohol, more specifically methanol can be used. May be, but is not limited thereto.

When using a mixed solvent of water and an organic solvent, the ratio may be, for example, water and an organic solvent in a volume ratio of 1: 0.7 to 1.5, for example, 1: 1: 0.8 to 1.3, but is not limited thereto.

The surfactant may be, for example, cetyltrimethylammonium bromide (CTAB), hexadecyltrimethylammonium bromide (TMABr), hexadecyltrimethylpyridinium chloride (TMPrCl), tetramethylammonium chloride (TMACl), and the like, and specifically, CTAB may be used.

The surfactant may be added in an amount of, for example, 1 g to 10 g, for example, 1 g to 8 g, 2 g to 8 g, 3 g to 8 g, etc., per liter of solvent, but is not limited thereto.

The silica precursor may be added after stirring with the addition of a surfactant to the solvent. The silica precursor may be, for example, tetramethyl orthosilicate (TMOS), but is not limited thereto.

The stirring may be performed, for example, for 10 minutes to 30 minutes, but is not limited thereto.

The silica precursor may be added, for example, 0.5 ml to 5 ml per liter of solvent, for example, 0.5 ml to 4 ml, 0.5 ml to 3 ml, 0.5 ml to 2 ml, 1 ml to 2 ml, etc. within the above range, but is not limited thereto. It is not.

If necessary, sodium hydroxide may be further used as a catalyst, which may be added with stirring after adding the surfactant to the solvent and before adding the silica precursor.

Sodium hydroxide may be, for example, 0.5 ml to 8 ml per liter of solvent, for example, 0.5 ml to 5 ml, 0.5 ml to 4 ml, 1 ml to 4 ml, 1 ml to 3 ml, 2 ml to 3 ml, etc., based on 1 M aqueous sodium hydroxide solution. However, the present invention is not limited thereto.

After addition of the silica precursor, the solution can be reacted with stirring. The stirring may be performed for example, for 2 hours to 15 hours, for example, within the above range, for example, 3 hours to 15 hours, 4 hours to 15 hours, 4 hours to 13 hours, 5 hours to 12 hours, 6 hours to 12 hours. , 6 hours to 10 hours, but is not limited thereto. If the stirring time (reaction time) is too short, nucleation may be insufficient.

After the agitation, the solution may be aged. Aging may be performed for example, for 8 hours to 24 hours, for example, within the range of 8 hours to 20 hours, 8 hours to 18 hours, 8 hours to 16 hours, 8 hours to 14 hours, 10 hours to 16 hours. , 10 hours to 14 hours, but is not limited thereto.

Thereafter, the reaction product may be washed and dried to obtain porous silica particles.

If desired, separation of unreacted material may be preceded before washing.

Separation of the unreacted material can be carried out by separating the supernatant, for example by centrifugation.

Centrifugation can be carried out, for example, at 6,000 to 10,000 rpm, the time being for example 3 to 60 minutes, for example 3 to 30 minutes, 3 to 30 minutes, 5 minutes to within the above range. It may be performed in 30 minutes, but is not limited thereto.

The washing may be performed with water and / or an organic solvent, and in particular, since the substances that can be dissolved for each solvent may be different, water and an organic solvent may be used once or several times, and water or an organic solvent may be used only once or several times. Can be washed The number of times may be, for example, two or more, ten or less, for example, three or more and ten or less, four or more and eight or less, four or more and six or less.

The organic solvent is, for example, ethers such as 1,4-dioxane (particularly cyclic ethers); Halogenated hydrocarbons such as chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride and 1,2-dibromoethane; Acetone, methyl isobutyl ketone, γ-butyrolactone, 1,3-dimethyl-imidazolidinone, methyl ethyl ketone, cyclohexanone, cyclopentanone, 4-hydroxy-4-methyl-2-pentanone, etc. Ketones; Carbon-based aromatics such as benzene, toluene, xylene and tetramethylbenzene; Alkyl amides such as N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide and N-methylpyrrolidone; Alcohols such as methanol, ethanol, propanol and butanol; Ethylene glycol monoethyl ether, ethylene glycol monomethyl ether, ethylene glycol monobutyl ether, diethylene glycol monoethyl ether, diethylene glycol monomethyl ether, diethylene glycol monobutyl ether, propylene glycol monomethyl ether, propylene glycol monoethyl ether Glycol ethers (cellosolve) such as dipropylene glycol diethyl ether and triethylene glycol monoethyl ether; Dimethylacetamide (DMAc), N, N-diethylacetamide, dimethylformamide (DMF), diethylformamide (DEF), N, N-dimethylacetamide (DMAc), N-methylpyrrolidone ( NMP), N-ethylpyrrolidone (NEP), 1,3-dimethyl-2-imidazolidinone, N, N-dimethylmethoxyacetamide, dimethyl sulfoxide, pyridine, dimethyl sulfone, hexamethylphosphoamide , Tetramethylurea, N-methylcarrolactam, tetrahydrofuran, m-dioxane, P-dioxane, 1,2-dimethoxyethane and the like can be used, and specifically, alcohol, more specifically ethanol can be used. May be, but is not limited thereto.

Washing may be carried out under centrifugation, for example at 6,000 to 10,000 rpm, the time being for example 3 to 60 minutes, for example 3 to 30 minutes, 3 minutes within this range. To 30 minutes, 5 minutes to 30 minutes, etc., but is not limited thereto.

Washing may be performed by filtering out particles with a filter without centrifugation. The filter may have pores less than or equal to the diameter of the porous silica particles. If the reaction solution is filtered through such a filter, only particles remain on the filter, and water and / or an organic solvent can be poured over the filter and washed.

When washing, water and an organic solvent may be used one or several times, and may be washed once or several times with water or an organic solvent alone. The number of times may be, for example, two or more, ten or less, for example, three or more and ten or less, four or more and eight or less, four or more and six or less.

Drying may be performed, for example, at 20 ° C. to 100 ° C., but is not limited thereto, and may be performed in a vacuum state.

Thereafter, the pores of the obtained porous silica particles are expanded.

Pore expansion can be performed using pore expanding agents.

For example, the pore swelling agent may be trimethylbenzene, triethylbenzene, tripropylbenzene, tributylbenzene, tripentylbenzene, trihexylbenzene, toluene, benzene, and the like, and specifically, trimethylbenzene may be used, but is not limited thereto. It doesn't happen.

In addition, the pore swelling agent may use, for example, N, N-dimethylhexadecylamine (N, N-dimethylhexadecylamine, DMHA), but is not limited thereto.

Pore expansion can be carried out, for example, by mixing porous silica particles in a solvent with a pore swelling agent and heating to react.

The solvent may be, for example, water and / or an organic solvent, and the organic solvent may be, for example, ethers such as 1,4-dioxane (particularly cyclic ethers); Halogenated hydrocarbons such as chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride and 1,2-dibromoethane; Ketones such as acetone, methyl isobutyl ketone and cyclohexanone; Carbon-based aromatics such as benzene, toluene and xylene; Alkyl amides such as N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide and N-methylpyrrolidone; Alcohols such as methanol, ethanol, propanol and butanol; Or the like, and specifically, alcohol, more specifically ethanol, may be used, but is not limited thereto.

The porous silica particles are, for example, 10 g to 200 g per liter of solvent, for example, 10 g to 150 g, 10 g to 100 g, 30 g to 100 g, 40 g to 100 g, 50 g to 100 g, 50 g to 80 g, 60 g to 80 g, etc., within the above range. It may be added in a ratio, but is not limited thereto.

The porous silica particles may be evenly dispersed in a solvent, for example, the porous silica particles may be added to the solvent and ultrasonically dispersed. In the case of using a mixed solvent, the second solvent may be added after the porous silica particles are dispersed in the first solvent.

The pore swelling agent is, for example, 10 to 200 parts by volume, 100 to 150 parts by volume, 10 to 100 parts by volume, 10 to 80 parts by volume, 30 to 80 parts by volume, 30 to 70 parts by volume based on 100 parts by volume of solvent. It may be added in a ratio such as volume parts, but is not limited thereto.

The reaction can be carried out, for example, at 120 ° C to 180 ° C. For example, within the range of 120 ℃ to 170 ℃, 120 ℃ to 160 ℃, 120 ℃ to 150 ℃, 130 ℃ to 180 ℃, 130 ℃ to 170 ℃, 130 ℃ to 160 ℃, 130 ℃ to 150 ℃ It may be performed, but is not limited thereto.

The reaction can be carried out, for example, for 24 hours to 96 hours. For example, within the range of 30 hours to 96 hours, 30 hours to 96 hours, 30 hours to 80 hours, 30 hours to 72 hours, 24 hours to 80 hours, 24 hours to 72 hours, 36 hours to 96 hours, 36 36 hours to 80 hours, 36 hours to 72 hours, 36 hours to 66 hours, 36 hours to 60 hours, 48 hours to 96 hours, 48 hours to 88 hours, 48 hours to 80 hours, 48 hours to 72 hours, etc. It is not limited to this.

The time and temperature can be adjusted within the ranges exemplified above so that the reaction can be carried out sufficiently without excess. For example, when the reaction temperature is lowered, the reaction time may be increased, or when the reaction temperature is lowered, the reaction time may be shortened. If the reaction is not sufficient, the expansion of the pores may not be sufficient, and if the reaction proceeds excessively, the particles may collapse due to the expansion of the pores.

The reaction can be carried out, for example, by gradually raising the temperature. Specifically, it may be carried out by gradually increasing the temperature at a rate of 0.5 ℃ / min to 15 ℃ / min from the room temperature to the temperature, for example, 1 ℃ / min to 15 ℃ / min, 3 ℃ / min within the above range To 15 ° C./minute, 3 ° C./minute to 12 ° C./minute, 3 ° C./minute to 10 ° C./minute, and the like, but are not limited thereto.

After the reaction, the reaction liquid can be cooled slowly, for example, it can be cooled by gradually reducing the temperature. Specifically, it may be carried out by gradually decreasing the temperature at a rate of 0.5 ℃ / min to 20 ℃ / min from the temperature to room temperature, for example, from 1 ℃ / min to 20 ℃ / min, 3 ℃ / min to within the above range 20 ° C./minute, 3 ° C./minute to 12 ° C./minute, 3 ° C./minute to 10 ° C./minute, and the like, but is not limited thereto.

After cooling, the reaction product can be washed and dried to obtain porous silica particles with expanded pores.

If desired, separation of unreacted material may be preceded before washing.

Separation of the unreacted material can be carried out by separating the supernatant, for example by centrifugation.

Centrifugation can be carried out, for example, at 6,000 to 10,000 rpm, the time being for example 3 to 60 minutes, for example 3 to 30 minutes, 3 to 30 minutes, 5 minutes to within the above range. It may be performed in 30 minutes, but is not limited thereto.

The washing may be performed with water and / or an organic solvent, and in particular, since the substances that can be dissolved for each solvent may be different, water and an organic solvent may be used once or several times, and water or an organic solvent may be used only once or several times. Can be washed The number of times may be, for example, two or more times, ten times or less, for example, three times, four times, five times, six times, seven times, eight times, and the like.

The organic solvent is, for example, ethers such as 1,4-dioxane (particularly cyclic ethers); Halogenated hydrocarbons such as chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride and 1,2-dibromoethane; Ketones such as acetone, methyl isobutyl ketone and cyclohexanone; Carbon-based aromatics such as benzene, toluene and xylene; Alkyl amides such as N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide and N-methylpyrrolidone; Alcohols such as methanol, ethanol, propanol and butanol; Or the like, and specifically, alcohol, more specifically ethanol, may be used, but is not limited thereto.

Washing may be carried out under centrifugation, for example at 6,000 to 10,000 rpm, the time being for example 3 to 60 minutes, for example 3 to 30 minutes, 3 minutes within this range. To 30 minutes, 5 minutes to 30 minutes, etc., but is not limited thereto.

Washing may be performed by filtering out particles with a filter without centrifugation. The filter may have pores less than or equal to the diameter of the porous silica particles. If the reaction solution is filtered through such a filter, only particles remain on the filter, and water and / or an organic solvent can be poured over the filter and washed.

When washing, water and an organic solvent may be used one or several times, and may be washed once or several times with water or an organic solvent alone. The number of times may be, for example, two or more, ten or less, for example, three or more and ten or less, four or more and eight or less, four or more and six or less.

Drying may be performed, for example, at 20 ° C. to 100 ° C., but is not limited thereto, and may be performed in a vacuum state.

The particles obtained can then be calcined.

Calcination is a process of heating the particles to have a more dense structure on the surface and the inside, and removing the organic substances filling the pores, for example, 3 to 8 hours at 400 ℃ to 700 ℃, specifically 500 to 600 It may be performed at 4 ℃ to 5 hours, but is not limited thereto.

Thereafter, the obtained porous silica particles may be surface modified.

Surface modification can be performed inside the surface and / or pores. The particle surface and the inside of the pore may be surface-modified identically or may be surface-modified differently.

Surface modification can cause the particles to charge or to have hydrophilic and / or hydrophobic properties. Surface modification can be carried out, for example, by reacting a compound having substituents such as hydrophilic, hydrophobic, cationic, anionic and the like to be introduced with the particles, and the compound can be, for example, an alkoxysilane having a C1 to C10 alkoxy group. However, it is not limited thereto. The alkoxysilane has one or more alkoxy groups, and may have, for example, 1 to 3, and there may be a substituent to be introduced into a site where the alkoxy group is not bonded or a substituent substituted therewith.

When the alkoxysilane reacts with the porous silicon particles, a covalent bond is formed between the silicon atom and the oxygen atom so that the alkoxysilane can be bonded to the surface and / or inside the pores of the porous silicon particle, and the alkoxysilane has a substituent to be introduced. The substituents may be introduced into the surface and / or the pores of the porous silicon particles.

The reaction may be carried out by reacting porous silica particles dispersed in a solvent with an alkoxysilane.

The solvent may be water and / or an organic solvent, and the organic solvent may be, for example, ethers such as 1,4-dioxane (particularly cyclic ethers); Halogenated hydrocarbons such as chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride and 1,2-dibromoethane; Acetone, methyl isobutyl ketone, γ-butyrolactone, 1,3-dimethyl-imidazolidinone, methyl ethyl ketone, cyclohexanone, cyclopentanone, 4-hydroxy-4-methyl-2-pentanone, etc. Ketones; Carbon-based aromatics such as benzene, toluene, xylene and tetramethylbenzene; Alkyl amides such as N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide and N-methylpyrrolidone; Alcohols such as methanol, ethanol, propanol and butanol; Ethylene glycol monoethyl ether, ethylene glycol monomethyl ether, ethylene glycol monobutyl ether, diethylene glycol monoethyl ether, diethylene glycol monomethyl ether, diethylene glycol monobutyl ether, propylene glycol monomethyl ether, propylene glycol monoethyl ether Glycol ethers (cellosolve) such as dipropylene glycol diethyl ether and triethylene glycol monoethyl ether; Dimethylacetamide (DMAc), N, N-diethylacetamide, dimethylformamide (DMF), diethylformamide (DEF), N, N-dimethylacetamide (DMAc), N-methylpyrrolidone ( NMP), N-ethylpyrrolidone (NEP), 1,3-dimethyl-2-imidazolidinone, N, N-dimethylmethoxyacetamide, dimethyl sulfoxide, pyridine, dimethyl sulfone, hexamethylphosphoamide , Tetramethylurea, N-methyl carrolactam, tetrahydrofuran, m-dioxane, P-dioxane, 1,2-dimethoxyethane and the like can be used. Specifically, toluene can be used, but is not limited thereto. It doesn't happen.

Charging to a positive charge can be carried out by reacting with an alkoxysilane having a basic group such as a nitrogen-containing group such as an amino group or an aminoalkyl group. Specifically, N- [3- (Trimethoxysilyl) propyl] ethylenediamine, N1- (3-Trimethoxysilylpropyl) diethylenetriamine, (3-Aminopropyl) trimethoxysilane, N- [3- (Trimethoxysilyl) propyl] aniline, Trimethoxy [3- (methylamino) propyl] silane, 3- (2-Aminoethylamino) propyldimethoxymethylsilane, etc. may be used, but is not limited thereto.

Charging to the negative charge can be carried out by reacting with an alkoxysilane having an acidic group such as a carboxyl group, a sulfonic acid group, a thiol group, and the like. Specifically, (3-Mercaptopropyl) trimethoxysilane may be used, but is not limited thereto.

Hydrophilic properties include hydrophilic groups such as hydroxy groups, carboxy groups, amino groups, carbonyl groups, sulfhydryl groups, phosphate groups, thiol groups, ammonium groups, ester groups, imide groups, thiimide groups, keto groups, ether groups, indene groups, sulfonyl groups And an alkoxysilane having a polyethylene glycol group or the like. Specifically, N- [3- (Trimethoxysilyl) propyl] ethylenediamine, N1- (3-Trimethoxysilylpropyl) diethylenetriamine, (3-Aminopropyl) trimethoxysilane, (3-Mercaptopropyl) trimethoxysilane, Trimethoxy [3- (methylamino) propyl] silane, 3- (2-Aminoethylamino) propyldimethoxymethylsilane may be used, but is not limited thereto.

Hydrophobic properties include hydrophobic substituents such as substituted or unsubstituted C1 to C30 alkyl groups, substituted or unsubstituted C3 to C30 cycloalkyl groups, substituted or unsubstituted C6 to C30 aryl groups, substituted or unsubstituted C2 It can be made to react with the alkoxysilane which has a heteroaryl group of C30-C30, a halogen group, an ester group of C1-C30, a halogen containing group, etc. Specifically, Trimethoxy (octadecyl) silane, Trimethoxy-n-octylsilane, Trimethoxy (propyl) silane, Isobutyl (trimethoxy) silane, Trimethoxy (7-octen-1-yl) silane, Trimethoxy (3,3,3-trifluoropropyl) Silane, Trimethoxy (2-phenylethyl) silane, Vinyltrimethoxysilane, Cyanomethyl, 3- (trimethoxysilyl) propyl] trithiocarbonate, (3-Bromopropyl) trimethoxysilane, etc. may be used, but is not limited thereto.

In addition, hydrophobic substituents are present inside the pores to enhance the bonding ability with poorly water-soluble (hydrophobic) bioactive substances through surface modification, and the surface of the particles has hydrophilic substituents in terms of ease of use and formulation. May be treated, and a substituent may be present on the surface to bind other substances.

Surface modification may also be carried out in combination. For example, two or more surface modifications may be performed on the outer surface or inside the pores. As a specific example, it is possible to change the positively charged particles to have different surface properties by binding a compound containing a carboxyl group to an amide bond with silica particles into which amino groups are introduced, but are not limited thereto.

The reaction of the porous silica particles with alkoxysilanes can be carried out, for example, under heating.

The heating is performed at 80 ° C to 180 ° C, for example, at 80 ° C to 160 ° C, 80 ° C to 150 ° C, 100 ° C to 160 ° C, 100 ° C to 150 ° C, 110 ° C to 150 ° C, and the like within the above range. It may be, but is not limited thereto.

The reaction of the porous silica particles with the alkoxysilane is for example 4 to 20 hours, for example 4 to 18 hours, 4 to 16 hours, 6 to 18 hours, 6 to 16 hours, 8 hours to 18 hours, 8 hours to 16 hours, 8 hours to 14 hours, 10 hours to 14 hours, etc., but is not limited thereto.

The reaction temperature, time, and the amount of the compound used for surface modification may be selected according to the degree of surface modification, and the porous silica particles may be changed depending on the hydrophilicity, hydrophobicity, and charge of the bioactive material. By controlling the degree of hydrophilicity, hydrophobicity, and charge, the rate of release of the bioactive substance can be controlled. For example, if the bioactive material has a strong negative charge at neutral pH, the reaction temperature can be increased or the reaction time can be increased, and the compound throughput can be increased, so that the porous silica particles have a strong positive charge. However, the present invention is not limited thereto.

In addition, the porous silica particles according to an embodiment of the present invention may be produced by, for example, the preparation of particles of small pores, pore expansion, surface modification, internal pore modification process.

The small pore particle preparation and pore expansion process may be based on the above-described process, and the washing and drying process may be performed after the small pore particle production and after the pore expansion process.

If desired, separation of unreacted material may be preceded before washing.

Separation of the unreacted material can be carried out by separating the supernatant, for example by centrifugation.

Centrifugation can be carried out, for example, at 6,000 to 10,000 rpm, and the time is, for example, 3 to 60 minutes, specifically, 3 to 30 minutes, 3 to 30 minutes, 5 minutes to within the above range. It may be performed in 30 minutes, but is not limited thereto.

The washing after the preparation of the particles of the small pores may be performed by a method / condition within the ranges exemplified above, but is not limited thereto.

Washing after pore expansion can be carried out in more relaxed conditions than previously illustrated. For example, washing may be performed within three times, but is not limited thereto.

Surface modification and internal pore modification may be by the processes as described above, respectively, the process may be carried out in the order of surface modification and internal pore modification, and the washing process of the particles may be further performed between the two processes. have.

When the washing is performed in a more relaxed condition after the preparation of the small pores and the expansion of the pores, the reaction solution such as the surfactant used in the preparation of the particles and the expansion of the pores is filled in the pores so that the inside of the pores is not modified during surface modification. Only the surface can be modified. Then, washing the particles may remove the reaction solution in the pores.

Particle washing between surface modification and internal pore reforming can be done with water and / or an organic solvent, and in particular, different solvents can be used to dissolve the water and the organic solvent once or several times. Alternatively, the organic solvent alone may be washed once or several times. The number of times may be, for example, two or more, ten or less, specifically, three or more and ten or less, four or more and eight or less, four or more and six or less.

Washing may be carried out under centrifugation, for example at 6,000 to 10,000 rpm, the time being for example 3 to 60 minutes, specifically 3 to 30 minutes, 3 minutes within this range. To 30 minutes, 5 minutes to 30 minutes, etc., but is not limited thereto.

Washing may be performed by filtering out particles with a filter without centrifugation. The filter may have pores less than or equal to the diameter of the porous silica particles. If the reaction solution is filtered through such a filter, only particles remain on the filter, and water and / or an organic solvent can be poured over the filter and washed.

When washing, water and an organic solvent may be used one or several times, and may be washed once or several times with water or an organic solvent alone. The number of times may be, for example, two or more, ten or less, specifically, three or more and ten or less, four or more and eight or less, four or more and six or less.

Drying may be performed, for example, at 20 ° C. to 100 ° C., but is not limited thereto, and may be performed in a vacuum state.

Of bioactive substances Support

The bioactive material may be supported on the surface of the porous silica particles and / or inside the pores.

Support may be carried out, for example, by mixing porous silica particles and a bioactive material in a solvent.

The solvent may be water and / or an organic solvent, and the organic solvent may be, for example, ethers such as 1,4-dioxane (particularly cyclic ethers); Halogenated hydrocarbons such as chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride and 1,2-dibromoethane; Ketones such as acetone, methyl isobutyl ketone and cyclohexanone; Carbon-based aromatics such as benzene, toluene and xylene; Alkyl amides such as N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide and N-methylpyrrolidone; Alcohols such as methanol, ethanol, propanol and butanol; Etc. can be used.

In addition, PBS (phosphate buffered saline solution), SBF (Simulated Body Fluid), Borate-buffered saline, Tris-buffered saline may be used as a solvent.

The ratio of the porous silica particles and the bioactive material is not particularly limited. For example, the weight ratio is 1: 0.05 to 0.8, for example, 1: 0.05 to 0.7, 1: 0.05 to 0.6, 1: 0.1 to 0.8 within the above range. , 1: 0.1 to 0.6, 1: 0.2 to 0.8, 1: 0.2 to 0.6, and the like.

Release of bioactive substances

The bioactive material supported on the porous silica particles may be gradually released over an extended time. Such slow release may be continuous or discontinuous, linear or nonlinear, and may vary due to the nature of the porous silica particles and / or their interaction with the bioactive material.

The bioactive material supported on the porous silica particles is released as the porous silica particles are biodegraded, and the porous silica particles according to the present invention may be slowly decomposed to release the supported bioactive materials in a sustained manner. This may be controlled by, for example, adjusting the surface area, particle diameter, pore diameter, substituents in the surface and / or pores, degree of compactness of the porous silica particles, and the like, but are not limited thereto.

In addition, the bioactive material supported on the porous silica particles may be released while being diffused from the porous silica particles, which is affected by the relationship between the porous silica particles, the bioactive material and the bioactive material emitting environment. By controlling this it is possible to control the release of bioactive substances. For example, it can be controlled by strengthening or weakening the binding strength of the porous silica particles with the bioactive material by surface modification.

More specifically, if the supported bioactive material is poorly water-soluble (hydrophobic), the surface and / or the inside of the pores may have hydrophobic substituents, thereby increasing the bonding strength between the porous silica particles and the bioactive material, Thereby, the bioactive substance can be released in a sustained manner. This may be, for example, the surface-modified porous silica particles with an alkoxysilane having a hydrophobic substituent.

As used herein, "poorly soluble" means to be insoluble (practically insoluble) or only slightly soluble (with respect to water), which means "Pharmaceutical Science" 18 ^th Edition ( USP, Remington, Mack Publishing Company).

The poorly water-soluble bioactive substance may be, for example, water solubility at less than 10 g / L, specifically less than 5 g / L, more specifically less than 1 g / L at 1 atmosphere and 25 ° C., but is not limited thereto.

When the supported bioactive material is water-soluble (hydrophilic), the surface and / or the inside of the pore may have a hydrophilic substituent, thereby increasing the binding force between the porous silica particles and the bioactive material, whereby the bioactive material is sustained. May be released. This may be, for example, the surface-modified porous silica particles with an alkoxysilane having a hydrophilic substituent.

For example, the water-soluble bioactive substance may have a water solubility of 10 g / L or more at 1 atmosphere and 25 ° C., but is not limited thereto.

When the supported bioactive material is charged, the surface of the particle and / or the inside of the pore may be charged with the opposite charge, thereby increasing the binding force between the porous silica particles and the bioactive material, whereby the bioactive material is This can be released slowly. This may be, for example, the surface-modified porous silica particles with an alkoxysilane having an acidic group or a basic group.

Specifically, if the bioactive material is positively charged at neutral pH, the surface of the particles and / or the inside of the pores may be negatively charged at the neutral pH, whereby the binding force between the porous silica particles and the bioactive material is increased. Increasingly, bioactives may be released in a sustained manner. For example, the porous silica particles may be surface-modified with an alkoxysilane having an acidic group such as a carboxyl group (-COOH) and a sulfonic acid group (-SO ₃ H).

In addition, if the bioactive material is negatively charged at neutral pH, the surface of the particles and / or the inside of the pores may be positively charged, thereby increasing the binding force between the porous silica particles and the bioactive material, thereby increasing the bioactive material. This can be released slowly. For example, the porous silica particles may be surface-modified with an alkoxysilane having a basic group such as an amino group or another nitrogen-containing group.

The bioactive material may be released for a period of, for example, 7 days to 1 year or more, depending on the type of treatment required, the release environment, and the porous silica particles used.

In addition, since the porous silica particles according to the present invention may be 100% decomposed as biodegradable, the bioactive material supported thereon may be released 100%.

Of bioactive carriers Formulation  And administration

Bioactive substance carriers of the invention may be formulated for delivery via any route of administration. "Route of administration" can refer to any route of administration known in the art including, but not limited to, aerosol, nasal, oral, transmucosal, transdermal, parenteral or intestine.

The porous silica particles according to the present invention are biodegradable and can be 100% decomposed, and thus can be parenterally administered because of excellent stability in the body, and thus can be formulated into preparations for parenteral administration.

"Parenteral" means orbital, intraocular, intravenous, intraarterial, intraarticular, intracardiac, dermal, intramuscular, intraperitoneal, intrapulmonary, spinal cord, intrasternal, intravertebral, intrauterine, intravenous, subarachnoid, It refers to a route of administration generally associated with injection, including subcapsular, subcutaneous, transmucosal, or synapse. Via the parenteral route, the carrier may be in the form of a solution or suspension, for infusion or for injection or lyophilized form. Via the parenteral route, the carrier may be in the form of a solution or suspension for infusion or for injection. Via the enteric route, bioactive carriers may be in the form of tablets, gel capsules, sugar coated tablets, syrups, suspensions, solutions, powders, granules, emulsions, microspheres or nanospheres or lipid vesicles or polymer vesicles that allow controlled release. Can be. Typically, the carrier is administered by injection, either intravenously or intraperitoneally. Methods for these administrations are known to those skilled in the art.

The bioactive substance carrier according to the present invention may also contain any pharmaceutically acceptable carrier. As used herein, a "pharmaceutically acceptable carrier" is a pharmaceutical composition involved in the transport or transport of a compound of interest from one tissue, organ or part of the body to another tissue, organ or part of the body. It refers to acceptable materials, compositions or vehicles. For example, the carrier may be a liquid or solid filler, diluent, excipient, solvent or encapsulating material or combinations thereof. Each component of the carrier must be "pharmaceutically acceptable", ie compatible with the other ingredients of the formulation. It should also be suitable for use when in contact with any tissue or organ to which it can be contacted, which should not involve the risk of any other complications that are too great than toxic, irritant, allergic reactions, immunogenicity or its therapeutic advantages. do.

Bioactive carriers according to the invention can also be encapsulated, purified or prepared in emulsions or syrups for oral administration. Pharmaceutically acceptable solid or liquid carriers may be added to enhance or stabilize the composition, or to facilitate the preparation of the composition. Liquid carriers include syrup, peanut oil, olive oil, glycerin, saline, alcohol and water. Solid carriers include starch, lactose, calcium sulfate, dihydrate, white earth, magnesium stearate or stearic acid, talc, pectin, acacia, agar or gelatin. The carrier may also include sustained release materials such as glyceryl monostearate or glyceryl distearate, alone or in combination with waxes.

Bioactive carriers can be milled, mixed, granulated and compressed if necessary for tablet form; Or prepared according to conventional pharmaceutical techniques involving grinding, mixing and filling for hard gelatin capsule forms. When a liquid carrier is used, the preparation will be in the form of a syrup, elixir, emulsion or aqueous or non-aqueous suspension. Such liquid formulations may be administered orally directly or filled into soft gelatin capsules.

The bioactive substance carrier according to the present invention may be delivered in a therapeutically effective amount. The exact therapeutically effective amount is that amount of the composition that produces the most effective result for therapeutic efficacy in a given subject. This amount includes the characteristics of the therapeutic compound (including activity, pharmacokinetics, pharmacodynamics and bioactivity), the physiological condition of the subject (age, sex, disease type and stage, general physical health, response to a given dosage and type of medicament) ), The nature of the pharmaceutically acceptable carrier or carriers in the formulation, and the route of administration, and will depend upon a number of factors. One skilled in the clinical and pharmacological arts will determine the therapeutically effective amount through routine experimentation, for example by monitoring the subject's response to administration of the compound and adjusting the dosage accordingly. For further guidance, see Remington: The Science and Practice of Pharmacy (Gennaro ed. 20th edition, Williams & Wilkins PA, USA) (2000).

The subject to which the drug delivery agent of the present invention is administered may be a mammal, including a human, specifically a human.

Prior to administration to the subject, the formulation may be added to the formulation. Liquid formulations may be preferred. For example, these formulations may include oils, polymers, vitamins, carbohydrates, amino acids, salts, buffers, albumin, surfactants, extenders or combinations thereof.

Carbohydrate formulations include sugars or sugar alcohols such as monosaccharides, disaccharides or polysaccharides or water soluble glucans. Sugars or glucans are fructose, dextrose, lactose, glucose, mannose, sorbose, xylose, maltose, sucrose, dextran, pullulan, dextrin, alpha and beta cyclodextrin, soluble starch, hydroxyethyl starch and Carboxymethylcellulose or mixtures thereof. "Sugar alcohol" is defined as a C4 to C8 hydrocarbon with --OH group and includes galactitol, inositol, mannitol, xylitol, sorbitol, glycerol and arabitol. These sugars or sugar alcohols mentioned above may be used individually or in combination. There is no fixed limit to the amount used if the sugar or sugar alcohol is soluble in the aqueous formulation. In one embodiment, the sugar or sugar alcohol concentration is 1.0 w / v% to 7.0 w / v, more preferably 2.0 to 6.0 w / v%.

Amino acid formulations include the sympathetic (L) forms of carnitine, arginine and betaine; However, other amino acids may be added.

In some embodiments, the polymer as a formulation comprises polyvinylpyrrolidone (PVP) having an average molecular weight of 2,000 to 3,000, or polyethylene glycol (PEG) having an average molecular weight of 3,000 to 5,000.

It is also desirable to use a buffer in the composition to minimize pH change in solution prior to lyophilization or after reconstitution. Most of the physiological buffers can be used, including but not limited to citrate, phosphate, succinate and glutamate buffers or mixtures thereof. In some embodiments, the concentration is 0.01-0.3 moles. Surfactants that can be added to the formulation are shown in European Patents 270,799 and 268,110.

In addition, the carriers can be chemically modified, for example, by covalent conjugation to the polymer to increase their circulating half-life. Preferred polymers and methods for attaching them to peptides are described in US Pat. No. 4,766,106; 4,179,337; No. 4,495,285; And 4,609,546, all of which are incorporated by reference in their entirety. Preferred polymers are polyoxyethylated polyols and polyethylene glycols (PEG). PEG is soluble in water at room temperature, and in some embodiments, the average molecular weight is 500-40,000, 2000-20,000, or 3,000-12,000. In some embodiments, PEG has at least one hydroxy group, such as a terminal hydroxy group. The hydroxyl group can be activated to react with the free amino group on the inhibitor. However, the type and amount of reactor can be varied to achieve the covalently conjugated PEG / antibody of the present invention.

Water soluble polyoxyethylated polyols are also useful in the present invention. They include polyoxyethylated sorbitol, polyoxyethylated glucose, polyoxyethylated glycerol (POG) and the like. POG is preferred. One reason is that the glycerol backbones of polyoxyethylated glycerol are mono-, di-, triglycerides of the same backbone of natural origin, for example in animals and humans. Thus, this branch is not necessarily regarded as a foreign agent in the body. POG is in the same molecular weight range as PEG. The structure for POG is described in Knauf et al., 1988, J. Bio. Chem. 263: 15064-15070, a discussion of POG / IL C 2 conjugates is found in US Pat. No. 4,766,106, both of which are incorporated herein by reference in their entirety.

After the liquid bioactive carrier is prepared, it can be lyophilized to prevent degradation and preserve sterility. Methods of lyophilizing liquid compositions are known to those skilled in the art. Immediately before use, the carrier may be reconstituted with a sterile diluent (eg, Ringer's solution, distilled water, or sterile saline) which may include additional components. Upon reconstitution, the carrier is administered to the subject using the methods known to those skilled in the art.

Use of Bioactive Substance Carriers

As described above, the bioactive substance carrier of the present invention includes a drug and porous silica particles, and the present invention provides the use of the porous silica particles described above in preparing a bioactive substance carrier.

As described above, the porous silica particles according to the present invention are biodegradable and can be slowly decomposed in vivo, and can release the supported bioactive substances in a sustained manner, and thus can be used in the preparation of sustained-release bioactive substance carriers. Can be.

The detailed physical properties, specifications, surface modifications, and the like may be within the ranges exemplified above, and may be manufactured by the methods / conditions within the ranges exemplified above.

Example

1. Preparation of Porous Silica Particles

(1) Preparation of Porous Silica Particles

One) Small pore  Preparation of Particles

Into a 2 L round bottom flask was placed 960 mL of distilled water (DW) and 810 mL of MeOH. 7.88 g of CTAB was added to the flask, followed by rapid addition of 4.52 mL of 1 M NaOH while stirring. After stirring for 10 minutes to add a uniform mixture, 2.6 mL of TMOS was added. After stirring for 6 hours to mix uniformly, it was aged for 24 hours.

The reaction solution was then centrifuged at 8000 rpm for 10 minutes at 25 ° C. to remove the supernatant, centrifuged at 8000 rpm for 10 minutes at 25 ° C. and washed five times with alternating ethanol and distilled water.

Thereafter, the resultant was dried in an oven at 70 ° C. to obtain 1.5 g of powdery small pore porous silica particles (pore average diameter of 2 nm and particle diameter of 200 nm).

2) pore expansion

1.5 g of small pore porous silica particle powder was added to 10 ml of ethanol for ultrasonic dispersion, and 10 ml of water and 10 ml of TMB (trimethyl benzene) were added for ultrasonic dispersion.

Thereafter, the dispersion was placed in an autoclave and reacted at 160 ° C. for 48 hours.

The reaction was carried out starting at 25 ° C. and warming up at a rate of 10 ° C./min, then slowly cooling at a rate of 1-10 ° C./min in the autoclave.

The cooled reaction solution was centrifuged at 8000 rpm for 10 minutes at 25 ° C to remove the supernatant, and centrifuged at 8000 rpm for 10 minutes at 25 ° C, and washed five times with alternating ethanol and distilled water.

After drying in an oven 70 ℃ to obtain a powdery porous silica particles (pore diameter 10 ~ 15nm, particle diameter 200nm).

3) calcination

The porous silica particles prepared in 2) were put in a glass vial, heated at 550 ° C. for 5 hours, and cooled slowly to room temperature after completion of the reaction to prepare particles.

(2) Preparation of Porous Silica Particles

Porous silica particles were prepared in the same manner as in 1. (1), except that the reaction conditions at the time of pore expansion were changed to 140 ° C. and 72 hours.

(3) Preparation of Porous Silica Particles (10L Scale)

Porous silica particles were prepared in the same manner as in Example 1. (1), except that a 5-fold large container was used and each material was used in a 5-fold volume.

(4) Preparation of Porous Silica Particles (Particle Diameter 300nm)

Porous silica particles were prepared in the same manner as in (1), except that 920 ml of distilled water and 850 ml of methanol were used to prepare the small pore particles.

(5) Preparation of Porous Silica Particles (Particle Size 500nm)

Porous silica particles were prepared in the same manner as in (1), except that 800 ml of distilled water, 1010 ml of methanol, and 10.6 g of CTAB were used to prepare the small pore particles.

(6) Preparation of Porous Silica Particles (Particle Diameter 1000nm)

Porous silica particles were prepared in the same manner as in (1), except that 620 ml of distilled water, 1380 ml of methanol, and 7.88 g of CTAB were used to prepare the small pore particles.

(7) Preparation of Porous Silica Particles (Porosity diameter  4nm)

Porous silica particles were prepared in the same manner as in (1), except that 2.5 mL of TMB was used for pore expansion.

(8) Preparation of Porous Silica Particles (Porosity diameter  7nm)

Porous silica particles were prepared in the same manner as in (1), except that 4.5 mL of TMB was used for pore expansion.

(9) Preparation of Porous Silica Particles (Porosity diameter  17nm)

Porous silica particles were prepared in the same manner as in (1), except that 11 mL of TMB was used for pore expansion.

(10) Preparation of Porous Silica Particles (Porosity diameter  23 nm)

Porous silica particles were prepared in the same manner as in (1), except that 12.5 mL of TMB was used for pore expansion.

(11) Preparation of Porous Silica Particles ( Dual reforming )

One) Small pore  Preparation of Particles

Example (1) Small pore particles were prepared in the same manner as in 1).

2) pore expansion

In the same manner as in Example (1) 2), the small pore particles were reacted with TMB, cooled and centrifuged to remove the supernatant. Thereafter, centrifuged under the same conditions as in Example (1) 2), washed three times with alternating ethanol and distilled water, and then dried under the same conditions as in Example (1) 2) to form porous silica particles (pore diameter 10) 15 nm, particle diameter 200 nm).

3) surface modification

After dispersing 0.8 g to 1 g of porous silica particles having expanded pores in 50 mL of toluene, 5 mL of (3-aminopropyl) triethoxysilane was added thereto, followed by heating at reflux at 120 ° C for 12 hours. The procedure was followed by the washing and drying procedures described above, followed by 1 mL of threethylene glycol (PEG3, 2- [2- (2-methoxyethoxy) ethoxy] acetic acid) and 100 mg of EDC (1-Ethyl-3- ( 3-dimethylaminopropyl) carbodiimide) and 200 mg of N-Hydroxysuccinimide (NHS) were dispersed in 30 mL PBS and allowed to react for 12 hours while stirring at room temperature. The product is then washed and dried.

The reaction solution of the previous step remains inside the pore, so that the inside of the pore is not modified.

4) pore inside washing

800 mg of surface modified particle powder was dissolved in 40 ml of 2M HCl / ethanol and refluxed under vigorous stirring for 12 hours.

Thereafter, the cooled reaction solution was centrifuged at 8000 rpm for 10 minutes to remove the supernatant, centrifuged at 8000 rpm for 10 minutes at 25 ° C, and washed five times with alternating ethanol and distilled water.

After drying in an oven 70 ℃ to obtain a powdery porous silica particles.

5) Pore internal reforming

(1) A profile group was introduced into the pores in the same manner as in Example 2. (2) 1).

② An octyl group was introduced into the pores in the same manner as in Example 2. (2) 2) described later.

2. Surface modification

(1) Approach with positive charge

1) 300nm particle size

Example 1. The porous silica particles of (4) were reacted with (3-Aminopropyl) triethoxysilane (APTES) to charge with a positive charge.

Specifically, 100 mg of porous silica particles were dispersed in a 10 mL toluene in a 100 mL round bottom flask with a bath sonicator. Then 1 mL of APTES was added and stirred at 400 rpm and stirred at 130 ° C. for 12 hours.

After the reaction was slowly cooled to room temperature, the supernatant was removed by centrifugation at 8000 rpm for 10 minutes, centrifuged at 8000 rpm for 10 minutes at 25 ° C and washed five times with alternating ethanol and distilled water.

After drying in an oven at 70 ℃ to obtain a porous porous silica particles having an amino group on the surface and inside the pores.

2) particle size 200nm

① Example 1. The porous silica particles of (1) were charged with positive charge by reacting with (3-Aminopropyl) triethoxysilane (APTES), except that 0.4 ml of APTES was added and the reaction time was 3 hours. 2. (1) It was modified similarly to the method of 1).

② Example 1. The porous silica particles of (9) were charged with positive charge by reacting with (3-Aminopropyl) triethoxysilane (APTES), and the other methods were modified in the same manner as in 2. (1) 1). .

③ Example 1. The porous silica particles of (10) were charged with positive charge by reacting with (3-Aminopropyl) triethoxysilane (APTES), and were modified in the same manner as in 2. (1) 1).

(2) Introduction of hydrophobic groups

1) Profiler

1. The porous silica particles of (1) were reacted with Trimethoxy (propyl) silane to introduce propyl groups on the surface and inside of the pores, except that 0.35ml of Trimethoxy (propyl) silane was added instead of APTES and reacted for 12 hours. Modification was carried out in the same manner as 2. (1).

2) Octyl

1.The porous silica particles of (1) were reacted with Trimethoxy-n-octylsilane to introduce propyl groups on the surface and inside of pores, except that 0.5ml of Trimethoxy-n-octylsilane was added instead of APTES and reacted for 12 hours. Modification was carried out in the same manner as 2. (1).

(3) Approach with negative charge

1) carboxyl group

1. The porous silica particles of (1) were reacted with succinic anhydride and charged negatively,

Dimethyl sulfoxide (DMSO) was used instead of toluene, 80 mg of succinic anhydride was added instead of APTES, and the mixture was stirred at room temperature for 24 hours. (1) Modified in the same manner as in 1).

2) Thiol group

It was modified in the same manner as in Example 2. (1) 1), except that 1.1 mL of MPTES was used instead of APTES.

3) Sulfonic acid group

(3) 100 mg of porous silica nanoparticles of 2) were dispersed in 1 mL of 1 M aqueous sulfuric acid solution and 20 mL of 30% hydrogen peroxide solution, stirred at room temperature to induce an oxidation reaction to oxidize thiol groups with sulfonic acid groups. After the same washing and drying as in Example 2 (1) 1).

3. Bioactive substance loading

(One) Doxorubicin ( Doxorubicin )

Example 1. Doxorubicin was loaded into the porous silica particles of (1).

Specifically, 5 mg of porous silica particle powder and 2 mg of doxorubicin were mixed under distilled water, and then allowed to stand at room temperature for 1 hour.

(2) Irinotecan ( Irinotecan )

Negatively charged Example 2. (3) 5 mg of the porous silica particle powder of 1) was dispersed in 1 mL of 1 × PBS, 2 mg of irinotecan was added and dispersed for 15 minutes, and then allowed to stand at room temperature for 1 hour.

(3) Sorafenib ( sorafenib )

Example 1. (11) 5) Sorafenib was loaded into the porous silica particles of ①.

Specifically, 5 mg of porous silica particle powder and 2 mg of sorafenib were mixed in 1 ml of deionized water / ethanol in a 5: 5 mixing ratio (volume ratio), and then incubated at room temperature for 1 hour. Then washed three times with 1 ml of deionized water.

(4) Retinoic acid ( Retinoic  acid)

Example 2. (1) 1 ml of retinoic acid solution (50 mM ethanol) was added to 100 µg of the porous silica particle powder of ① and left to stand at room temperature for 4 hours, followed by washing three times with 1 ml of ethanol.

(5) p53 Peptide

As the porous silica particles, the particles of Example 1. (11) 5) ① were used.

The p53 peptide used mimics a portion of the p53 protein sequence involved in apoptosis. The mimicked sequence relates to the sequence of the hydrophobic secondary helix structure where the p53 protein binds to the hMDM2 protein. Thus, the p53 peptide acts as an antagonist of the hMDM2 protein.

The amino acid sequence of the p53 peptide (Cal. m.w. 2596.78, found by MALDI-TOF 2597.92) is shown in Formula 1 (N terminus-> C terminus).

[Formula 1]

Z-Gly-Gly-Qln-Ser-Qln-Qln-Thr-Phe-Y-Asn-Leu-Trp-Arg-Leu-Leu-X-Qln-Asn-NH2

(X is a non-natural amino acid with an azide functional group introduced, 2-amino-5-azido-pentanoic acid; Y is a non-natural amino acid with an alkyne functional group introduced, and the side chain of D-Lys ) Introduced 4-pentynoic acid;

X and Y are linked together to form a triazole functional group via an azide-alkyne cycloaddition, or click reaction;

Z is 5 (6) -carboxyfluorescein (FAM).

1.3 mg (500 nmole) of p53 peptide was dissolved in 100 μl of DMSO, and 5 mL of an aqueous solution of 5 mg of porous silica particle powder was mixed in a 15 mL conical tube, followed by incubation at room temperature for 12 hours.

Porous silica particles loaded with p53 peptide were purified by centrifugation (9289 rcf, 8500 rpm, 20 minutes, 15 mL conical tube) and washing with water three times.

(6) siRNA

21 base pair duplex siRNAs targeting Green Fluorescence Protein (GFP) were purchased and synthesized by Bionic, Inc. (SEQ ID NO: sense; 5'-GGCUACGUCCAGGAGCGCACC-3 '(SEQ ID NO: 1), antisense; 5' UGCGCUCCUGGACGUAGCCUU-3 '(SEQ ID NO: 2)).

Example 2. (1) 10 μg of porous silica particles of 2) and 50 pmol of siRNA were mixed under 1 × PBS conditions and allowed to be loaded at room temperature for 30 minutes.

(7) Plasmid  DNA

A 6.7k base pair of plasmid DNA (SEQ ID NO: 5) prepared to express GFP with pcDNA3.3 backbone was produced from bacteria and used after purification.

Example 2. (1) 10 μg of porous silica particles of 3) and 0.25 μg of plasmid DNA were mixed under 1 × PBS and loaded at room temperature for 30 minutes.

(8) linear DNA

Forward primer-CMV promotor-eGFP cDNA-Reverse primer was used to obtain a linear DNA (SEQ ID NO: 6) of 1.9k base pair obtained by amplification by PCR.

Example 2 (1) 2) ③ 12.5 μg of porous silica particles and 0.25 μg of linear DNA were mixed under 1 × PBS conditions and loaded at room temperature for 30 minutes.

(9) protein

1) BSA

100 µg of porous silica particle powder of Example 2. (1) 2) ② and 10 µg of BSA (sigma Aldrich, A6003) were mixed in 200 µl of 1 x PBS, and then incubated at room temperature for 1 hour.

2) IgG

100 μg of the porous silica particle powder of Example 2. (1) 2) ② and 10 μg of anti-twist IgG (Santacruz, sc-81417) were mixed in 200 μl of 1 × PBS, and then incubated at room temperature for 1 hour.

3) RNase  A

100 μg of the porous silica particle powder of Example 1. (9) and 10 μg of RNase A (Sigma-aldrich, R6513) were mixed in 200 μl of 1 × PBS, and then incubated at room temperature for 1 hour.

4) Cas9

40 μg of the porous silica particle powder of Example 2. (1) 2) ①, 4 μg of Cas9 protein (SEQ ID NO: 3) and 2.25 μg of guide RNA (SEQ ID NO: 4) were mixed in 10 μl of 1 × PBS, and then stored at room temperature. Incubate for 1 hour.

Experimental Example

1. Confirm particle formation and pore expansion

Example 1. The microporous particles of the particles of (1) to (3) and the prepared porous silica particles were observed under a microscope to determine whether the small pore particles were formed uniformly or the pores were sufficiently expanded to form the porous silica particles uniformly. It was confirmed whether or not (Figs. 1 to 4).

1 is a photograph of the porous silica particles of 1. (1), Figure 2 is a photograph of the porous silica particles of 1. (2) it can be seen that evenly formed spherical porous silica particles with sufficiently expanded pores,

Figure 3 is a photograph of the small pore particles of 1. (1), Figure 4 is a comparison photograph of the small pore particles of 1. (1) and 1. (3), confirming that the spherical small pore particles are evenly generated. Can be.

2. BET surface area and pore volume calculation

The surface area and pore volume of the small pore particles of Example 1. (1) and the porous silica particles of Examples 1. (1), (7), (8) and (10) were calculated. The surface area was calculated by the Brunauer-Emmett-Teller (BET) method, and the pore size distribution was calculated by the Barrett-Joyner-Halenda (BJH) method.

The micrographs of the particles are shown in FIG. 5, and the calculation results are shown in Table 1 below.

3. Confirmation of Biodegradability of Porous Silica Particles

In order to confirm the biodegradability of the porous silica particles of Example 1 (1), the degree of biodegradation at 37 ° C. and SBF (pH 7.4) was observed under a microscope at 0 hours, 120 hours, and 360 hours, which is shown in FIG. 6. .

Referring to this, it can be seen that porous silica particles are biodegraded and nearly decomposed after 360 hours.

4. of porous silica particles Absorbance ratio  Measure

The absorbance ratio according to Equation 1 according to time was measured.

[Equation 1]

A _t / A ₀

Wherein A ₀ is the absorbance of the porous silica particles measured by placing 5 ml of the 1 mg / ml suspension of the porous silica particles into a cylindrical permeable membrane having pores having a diameter of 50 kDa,

Outside the permeable membrane is in contact with the permeable membrane, 15ml of the same solvent as the suspension is located, the inside and the outside of the permeable membrane is stirred 60 rpm at 37 ℃ horizontal,

A _t is the absorbance of the porous silica particles measured after t hours have elapsed since the measurement of A ₀ ).

Specifically, 5 mg of porous silica particle powder was dissolved in 5 ml of SBF (pH 7.4). Thereafter, 5 ml of the porous silica particle solution was placed in a permeable membrane having pores having a diameter of 50 kDa shown in FIG. 7. 15 ml of SBF was added to the outer membrane, and the SBF of the outer membrane was changed every 12 hours. The decomposition of the porous silica particles was performed at 37 ° C. with 60 rpm horizontal stirring.

The absorbance was then measured by UV-vis spectroscopy and analyzed at λ = 640 nm.

(1) absorbance ratio measurement

The absorbance ratio of the porous silica particles of Example 1 (1) was measured according to the above method, and the results are shown in FIG. 8.

Referring to this, it can be seen that t, which has an absorbance ratio of 1/2, decomposes very slowly in about 58 hours.

(2) By particle size

Example 1. The absorbance of the porous silica particles of (1), (5) and (6) was measured according to Equation 1, and the results are shown in FIG. 9 (using SBF as a suspension and a solvent).

Referring to this, it can be seen that t decreases with increasing particle diameter.

(2) pore average By diameter

Example 1. The absorbance of the porous silica particles of (1) and (9), and the microporous porous silica particles of Example 1. (1) as a control was measured according to Equation 1, and the results are shown in FIG. (SBF was used as the suspension and the solvent).

Referring to this, it can be seen that the porous silica particles of the example have a significantly larger t than the control.

(3) by pH

The absorbance for each pH of the porous silica particles of Example 1. (4) was measured. Absorbance was measured in SBF and in Tris at pH 2, 5, and 7.4 and the results are shown in FIG. 11.

Referring to this, although there is a difference in t for each pH, t, which is a ratio of absorbance 1/2, was 24 or more.

(4) Daejeon

The absorbance of the porous silica particles of Example 2. (1) 1) was measured, and the results are shown in FIG. 12 (Tris (pH 7.4) was used as the suspension and the solvent).

Referring to this, t, which has a ratio of absorbance 1/2 of a positively charged particle, was 24 or more.

5. Release of bioactive substances

(One) Doxorubicin

0.5 mg of porous silica particles loaded with doxorubicin (0.1 mg) were dispersed in PBS. The solution is maintained in a dynamic condition of horizontal stirring at 37 ° C. at 200 rpm. At each time point, the porous silica solution loaded with doxorubicin was allowed to settle using a centrifuge, and the absorbance (λab = 480 nm) of the supernatant was measured to determine the amount of doxorubicin released. The results are shown in FIG. 13.

Referring to this, doxorubicin is loaded with a relatively weak binding force with the particle surface, it can be seen that because of the high solubility of doxorubicin is released in relatively fast release, the physiologically active substance was released continuously over 70 hours have.

(2) Irinotecan

1 mg of porous silica particles loaded with irinotecan (0.2 mg) were dispersed in 1 mL of human plasma. The solution is maintained in a dynamic condition of horizontal stirring at 37 ° C. at 200 rpm. At each time point, the porous silica solution loaded with irinotecan was allowed to settle using a centrifuge, and the absorbance (λab = 255 or 278 nm) of the supernatant was measured to determine the amount of irinotecan released. The results are shown in FIG. 14.

Referring to this, it can be seen that about 50% of irinotecan was released after 5.5 hours, and the physiologically active substance was continuously released until 120 hours or more.

(3) Sorafenib

1 mg of porous silica particles loaded with sorafenib (0.1 mg) were each dispersed in 10 mL of 1 × PBS. Dynamic conditions were maintained with the solution horizontally agitated at 37 ° C. at 200 rpm. At each time point, the porous silica solution loaded with sorafenib was allowed to settle using a centrifuge and the absorbance (λab = 270 nm) of the supernatant was measured to determine the amount of sorafenib released. The results are shown in FIG. 15.

Referring to this, it can be seen that sorafenib, a poorly soluble bioactive substance, is released very slowly by interaction with porous silica particles having a hydrophobic substituent.

(4) Retinoic acid

0.1 mg of particles loaded with retinoic acid was placed in a PBS (pH 7.4) solution containing 5% ethanol and maintained at 37 ° C. with horizontal stirring. Every 24 hours, the solution containing the particles was centrifuged to measure the absorbance of the supernatant at a wavelength of 350 nm to determine the amount of retinoic acid released. The results are shown in FIG. 16.

Referring to this, it can be seen that retinoic acid having a negative charge is released very slowly by interaction with the positively charged porous silica particles, and is almost 100% released in about 10 days.

(5) p53 Peptide

5 mg of p53 peptide loaded particles were added to 5 mL of 1x PBS containing 10% FBS, or 5 mL of 1x PBS and rotated at 37 ° C. at 20 rpm to maintain a dynamic environment. The fluorescence intensity of 5 (6) -carboxyfluorescein (FAM), a fluorescent label bound to p53 peptide from the supernatant by centrifugation at 8500 rpm at each time point, was measured (Absorbance: 480 nm, Emmision: 520 nm).

The results are shown in FIG.

Referring to this, the p53 peptide is loaded with the binding force through the hydrophobic effect (hydrophobic effect) inside the porous silica particles can be seen that the release in the PBS solution. However, when a protein such as FBS (fetal bovine serum) is present in the solution, the p53 peptide can be dissolved in the solution while binding to the hydrophobic segment of the FBS protein and released out of the porous silica particles. have. Or, as the p53 peptide loaded inside the particle is released to the outside of the particle, FBS protein will be introduced into the particle.

(6) siRNA

10 [mu] l of porous silica particles loaded with Cy5-siRNA were resuspended in SBF (pH 7.4, 37 [deg.] C.) and placed in a permeable membrane (tube in FIG. 18) with a pore diameter of 20 kDa.

Thereafter, the permeation tube was soaked in 1.5 ml of SBF.

Release of siRNA was carried out at 37 ° C. with 60 rpm horizontal stirring.

Release solvents were recovered at 0.5, 1, 2, 4, 8, 12, 24 hours elapsed time before 24 hours, thereafter at 24 hour intervals, 0.5 ml of released solvent was recovered for fluorescence measurements and SBF was added.

The fluorescence intensity of Cy5-siRNA was measured at 670 nm wavelength (λex = 647 nm) to measure the degree of emission of siRNA, and the results are shown in FIG. 19.

Referring to this, it can be seen that the siRNA 50% release time is about 48 hours.

(7) Plasmid  DNA

Porous silica particles loaded with Plasmid DNA (1 μg of psdmid DNA, 50 μg of porous silica particles) were resuspended in PBS (pH 7.4, 37 ° C.), and a permeable membrane having a pore diameter of 20 kDa (the same tube as the tube of FIG. 18). Put in.

Thereafter, the permeation tube was soaked in 1.5 ml PBS.

Release of Plasmid DNA was performed at 37 ° C. with 60 rpm horizontal stirring.

The release solvent was recovered at the time of 0.5, 1, 2, 4, 8, 12, 24 hours before 24 hours, thereafter, at 24 hours intervals, 0.5 ml of the release solvent was recovered for the Hoechst-binding assay. Equivalent amount of PBS was added.

The fluorescence intensity of Hoechst 33342 was measured at 460 nm wavelength (λex = 360 nm) to measure the degree of emission of plasmid DNA, and the results are shown in FIGS. 20 and 21.

Referring to this, it can be seen that the release time of 50% of the plasmid DNA is about 24 hours.

(8) linear DNA

Porous silica particles loaded with linear DNA (3 μg linear DNA, 100 μg porous silica particles) were resuspended in PBS (pH 7.4, 37 ° C.) and a permeable membrane having a pore diameter of 20 kDa (the same tube as the tube of FIG. 18). Put in.

Thereafter, the permeation tube was soaked in 1.5 ml PBS.

Release of Plasmid DNA was performed at 37 ° C. with 60 rpm horizontal stirring.

The release solvent was recovered at the time of 0.5, 1, 2, 3, 4, 6, 12, and 24 hours before 24 hours, and thereafter, at 24 hours, 0.5 ml of the release solvent was collected for the Hoechst-binding assay. Recovered and added an equal amount of PBS.

The fluorescence intensity of Hoechst 33342 was measured at 460 nm wavelength (λex = 360 nm) to measure the degree of emission of plasmid DNA, and the results are shown in FIG. 22.

Referring to this, it can be seen that the release time of 50% of linear DNA is about 24 hours.

(9) protein

1) BSA

100 µg of porous silica particles loaded with BSA labeled with Fluorescein fluorescence were resuspended in 200 µl of SBF (pH 7.4) or PBS (pH 7.4).

Release of BSA was carried out at 37 ° C. with 60 rpm horizontal stirring.

At 6 h, 12 h, 24 h, 48 h, 96 h, 144 h and 240 h points, 200 μl of releasing solvent was recovered for fluorescence measurements and an equivalent amount of SBF or PBS was added.

Fluorescence intensity of Fluorescein fluorescence-labeled BSA was measured at 517 nm wavelength (λex = 492 nm) to measure the degree of emission of BSA, and the results are shown in FIG. 23.

Referring to this, it can be seen that BSA is released in both SBF and PBS in a sustained manner, and is almost 100% released over 250 hours.

2) IgG

100 μg of porous silica particles loaded with Fluorescein fluorescence labeled IgG were resuspended in 200 μl of SBF (pH 7.4) or PBS (pH 7.4),

Release of IgG was performed with 60 rpm horizontal stirring at 37 ° C.

At 6 h, 12 h, 24 h, 48 h, 96 h, 144 h and 240 h points, 200 μl of releasing solvent was recovered for fluorescence measurements and an equivalent amount of SBF or PBS was added.

Fluorescence intensity of Fluorescein fluorescence-labeled IgG was measured at 517 nm wavelength (λex = 492 nm) to measure the degree of emission of BSA, and the results are shown in FIG. 24.

Referring to this, it can be seen that IgG is released slowly in both SBF and PBS, and is almost 100% released over 250 hours.

3) RNase  A

100 μg of porous silica particles loaded with Fluorescein fluorescence labeled RNase A were resuspended in 200 μl of SBF (pH 7.4) or PBS (pH 7.4),

Release of RNase A was carried out at 37 ° C. with 60 rpm horizontal stirring.

At 6 h, 12 h, 24 h, 48 h, 96 h, 144 h and 240 h points, 200 μl of releasing solvent was recovered for fluorescence measurements and an equivalent amount of SBF or PBS was added.

Fluorescence intensity of Fluorescein fluorescence-labeled RNase A was measured at 517 nm wavelength (λex = 492 nm) to measure the degree of emission of BSA, and the results are shown in FIG. 25.

Referring to this, it can be seen that RNaseA is released in both SBF and PBS in a sustained manner and is almost 100% released over 250 hours.

4) Cas9

40 μg of porous silica particles loaded with Cas9 protein / guide RNA complex were suspended in PBS (pH 7.4).

Subsequently, the porous silica particles were treated in a serum-free medium in a slide glass on which 50,000 NIH 3T3 cells, known as mouse fibroblasts, were laid and incubated at 5% CO ₂ at 37 ° C.

At 1, 3, 6, and 24 hours, the medium was removed, washed with 1 × PBS solution, and incubated with 4% paraformaldehyde for 15 minutes to fix cells.

After washing with PBS and incubated for 1 hour in blocking buffer (1x PBS, 5% normal goat serum, 0.3% triton X-100).

After washing with PBS, His tag antibody (Santa Cruz, sc-8036) was incubated for 16 hours.

After washing with PBS again, Alexa Fluor 488-linked anti mouse secondary antibody (Abcam, ab150113) was incubated for 2 hours.

After washing with PBS, the slide glass was treated with DAPI (dye staining cell nuclei) to stain the nuclei of cells. Since the distribution of protein in the cell was confirmed using a fluorescence microscope, the results are shown in FIG.

In Figure 26, DAPI is a reagent for staining the nucleus, which is shown in blue in the fluorescence microscope image, and shows the location of the cell nucleus. Alexa Fluor 488 is a fluorescent dye labeled with Cas9 protein, which appears green in the fluorescence microscopy image and shows the location of the intracellular Cas9 protein. When the silica particles loaded with Cas 9 protein labeled Alexa Fluor488 were treated with DAPI staining and stained with DAPI, the fluorescence microscopy image confirmed whether the Cas 9 protein was introduced into the cell by the silica particles and the position of the nucleus.

Referring to this, Cas9 protein introduced into the cell is mainly observed in the cytoplasmic part 3 hours after the introduction, it can be seen that observed in the nucleus after 24 hours. Since the used silica particle itself is almost impossible to enter into the cell nucleus, it can be seen that the Cas9 protein is released from the silica particle after 24 hours in the cell and enters the nucleus known as an intracellular organelle, where the Cas9 protein is accumulated.

6. Delivery of bioactive substances

To verify the possible role of the carrier in the siRNA delivery studies at the animal level, the degree of tumor suppression according to the release of bioactive substances in mice (mouse) was confirmed.

Balb / c nude males (5 weeks old) were purchased from Orient Bio, Inc., and 3 million HeLa cells (cervical cancer cells) were dispersed in sterile 1x PBS to grow subcutaneous Xenograft tumors in mice, 70 mm ^When solidified tumors of ^three sizes were identified, PBS, FITC-porous silica particles (porous silica particles of Example 2. (1) 2) ②, FITC-porous silica particles loaded with Cy5-siRNA (Example 2) (1) 2) the porous silica particles of ②) were injected intramuscularly into the mouse tumor, respectively, and the fluorescence intensity and distribution were measured immediately before, immediately after, and 48 hours after the administration of the FOBI Fluorescence in vivo imaging system (Neo science, Korea). Observed through.

The FITC label was prepared by dispersing 50 mg of silica particles in 1 mL dimethyl sulfoxide (DMSO) and adding 25 μg (10 μl) of FITC-NHS (N-hydroxycuccinimide) solution (2.5 mg / mL) and blocking the light with aluminum foil. After reacting for 18 hours at room temperature, the reaction product was purified by centrifugation (8500 rpm, 10 minutes), and the supernatant was discarded. The supernatant was collected and dispersed evenly in ethanol, and this was repeated 3-4 times with ethanol-distilled water. Purification was performed until no FITC color was seen.

The results are shown in FIG. 27.

In FIG. 27, the control is PBS alone administration, cy5-siRNA is cy5-siRNA administration alone, FITC-DDV is FITC-only porous silica particles alone, the complex is cy5-siRNA loaded and FITC-labeled porous silica particles are administered. As shown, it can be seen that the siRNA delivered to the body by loading the particles have a longer duration of activity and stay longer at the injected site, showing strong fluorescence even after 48 hours.

Claims (32)

Bioactive substances; And porous silica particles supporting the bioactive material and having a plurality of pores having a diameter of 5 nm to 100 nm. The porous silica particles have a physiologically active substance carrier having t of 24 or more, wherein a ratio of absorbance of Equation 1 is 1/2: [Equation 1] A _t / A ₀ Wherein A ₀ is the absorbance of the porous silica particles measured by placing 5 ml of the 1 mg / ml suspension of the porous silica particles into a cylindrical permeable membrane having pores having a diameter of 50 kDa, Outside the permeable membrane is in contact with the permeable membrane, 15ml of the same solvent as the suspension is located, the inside and the outside of the permeable membrane is stirred 60 rpm at 37 ℃ horizontal, The pH of the suspension is 7.4, A _t is the absorbance of the porous silica particles measured after t hours have elapsed since the measurement of A ₀ ). The bioactive substance carrier according to claim 1, wherein the suspension is at least one selected from the group consisting of phosphate buffered saline (PBS) and simulated body fluid (SBF). The bioactive substance carrier according to claim 1, wherein A _t in Equation 1 is measured in an environment in which a solvent outside the permeable membrane is replaced every predetermined period. The bioactive substance carrier according to claim 1, wherein t is 24 to 120. The bioactive substance carrier of claim 1, wherein the porous silica particles are biodegradable. The physiologically active substance carrier according to claim 1, wherein the porous silica particles have a t of 70 to 120 such that the absorbance ratio of Equation 1 is 1/5. The physiologically active substance carrier according to claim 1, wherein the porous silica particles have a t of 130 to 220 such that the absorbance ratio of Equation 1 is 1/20. The physiologically active substance carrier according to claim 1, wherein the ratio of absorbance and t of the formula (1) has a Pearson correlation coefficient of 0.8 or more. The bioactive substance carrier according to claim 1, wherein the pore diameter is 7 nm to 30 nm. The bioactive substance carrier of claim 1, wherein the porous silica particles are spherical. The bioactive substance carrier according to claim 1, wherein the porous silica particles have an average diameter of 150 nm to 1000 nm. The bioactive substance carrier according to claim 1, wherein the porous silica particles have a BET surface area of 200 m ² / g to 700 m ² / g. The bioactive substance carrier of claim 1, wherein the porous silica particles have a BET surface area of 300 m ² / g to 450 m ² / g. The bioactive substance carrier according to claim 1, wherein the porous silica particles have a volume of 0.7 ml to 2.2 ml per gram. The bioactive substance carrier according to claim 1, wherein the porous silica particles have a volume of 1.0 ml to 2.0 ml per gram. The bioactive substance carrier according to claim 1, wherein the porous silica particles have a hydrophilic substituent or a hydrophobic substituent on an outer surface or inside a pore. The bioactive substance carrier according to claim 1, wherein the porous silica particles have a positive charge or a negative charge at a neutral pH at an outer surface or a pore inside. The bioactive material carrier of claim 1, wherein the bioactive material is poorly soluble, and the porous silica particles have a hydrophobic substituent on an outer surface or inside a pore. The bioactive substance carrier according to claim 1, wherein the bioactive substance is poorly soluble, and the porous silica particles have a hydrophobic substituent in the pores and a hydrophilic substituent in the outer surface. The bioactive substance carrier of claim 1, wherein the bioactive substance is negatively charged at neutral pH, and the silica particles are positively charged at neutral pH at the outer surface or the inside of the pore. The bioactive substance carrier of claim 1, wherein the bioactive substance is positively charged at neutral pH, and the silica particles are negatively charged at neutral pH at the outer surface or the inside of the pore. Bioactive substances; And spherical porous silica particles carrying the physiologically active substance and having a particle diameter of 150 nm to 500 nm and a plurality of pores having a diameter of 7 nm to 30 nm. The porous silica particles are physiologically active substance carrier having t of 24 to 120, where the ratio of absorbance of Equation 1 is 1/2: [Equation 2] A _t / A ₀ Wherein A ₀ is the absorbance of the porous silica particles measured by placing 5 ml of the 1 mg / ml suspension of the porous silica particles into a cylindrical permeable membrane having pores having a diameter of 50 kDa, Outside the permeable membrane is in contact with the permeable membrane, 15ml of the same solvent as the suspension is located, the inside and the outside of the permeable membrane is stirred 60 rpm at 37 ℃ horizontal, The suspension is PBS or SBF, pH is 7.4, A _t is the absorbance of the porous silica particles measured after t hours have elapsed since the measurement of A ₀ ). The bioactive substance carrier according to claim 22, wherein the porous silica particles have a BET surface area of 300 m ² / g to 450 m ² / g, and a volume per g of 1.0 ml to 2.0 ml. The bioactive material carrier of claim 22, wherein the bioactive material is poorly soluble and the porous silica particles have a hydrophobic substituent on an outer surface or inside a pore. The bioactive substance carrier according to claim 22, wherein the bioactive substance is poorly soluble, and the porous silica particles have a hydrophobic substituent inside the pores and a hydrophilic substituent on the outer surface. The bioactive substance carrier according to claim 22, wherein the bioactive substance is negatively charged at neutral pH, and the silica particles are positively charged at neutral pH at the outer surface or the inside of the pore. The bioactive carrier according to claim 22, wherein the bioactive material is positively charged at neutral pH, and the silica particles are negatively charged at neutral pH at the outer surface or the inside of the pore. A method for producing a bioactive substance carrier comprising contacting porous silica particles with a bioactive substance in a solvent. The method of claim 28, wherein the solvent is water, chloroform, methylene chloride, carbon tetrachloride, 1,2-dichloroethane, dichloroethylene, trichloroethylene, perchloroethylene, dichloropropane, amyl chloride, 1,2-dibromo Carbon, acetone, methyl isobutyl ketone, cyclohexanone, benzene, toluene, xylene, N, N-dimethylformamide, N, N-dibutylformamide, N, N-dimethylacetamide, N-methylpyrrolidone At least one selected from the group consisting of methanol, ethanol, propanol, butanol, PBS, SBF, borate-buffered saline and tris-buffered saline. The method of claim 28, wherein the weight ratio of porous silica particles to bioactive material is 1: 0.05 to 0.8. A method of delivering a physiologically active substance comprising parenterally administering the drug delivery agent of any one of claims 1 to 27 to an individual. The method of claim 31, wherein the parenteral administration is intraorbital, intraocular, infusion, intraarterial, intraarticular, intracardiac, intradermal, intramuscular, intraperitoneal, intrapulmonary, spinal cord, intrasternal, intravertebral, intrauterine, intravenous Intranasal, subarachnoid, subcapsular, subcutaneous, transmucosal or intranasal administration.

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