CA2083027C - Formation of protein microparticles by antisolvent precipitation - Google Patents
Formation of protein microparticles by antisolvent precipitation Download PDFInfo
- Publication number
- CA2083027C CA2083027C CA002083027A CA2083027A CA2083027C CA 2083027 C CA2083027 C CA 2083027C CA 002083027 A CA002083027 A CA 002083027A CA 2083027 A CA2083027 A CA 2083027A CA 2083027 C CA2083027 C CA 2083027C
- Authority
- CA
- Canada
- Prior art keywords
- protein
- solution
- solvent
- particles
- supercritical
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0065—Oxidoreductases (1.) acting on hydrogen peroxide as acceptor (1.11)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
- A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
- A61K9/1682—Processes
- A61K9/1688—Processes resulting in pure drug agglomerate optionally containing up to 5% of excipient
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/30—Extraction; Separation; Purification by precipitation
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/575—Hormones
- C07K14/62—Insulins
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/665—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans derived from pro-opiomelanocortin, pro-enkephalin or pro-dynorphin
- C07K14/695—Corticotropin [ACTH]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/98—Preparation of granular or free-flowing enzyme compositions
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/141—Feedstock
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/54—Improvements relating to the production of bulk chemicals using solvents, e.g. supercritical solvents or ionic liquids
Abstract
Microparticles of a material are formed by bringing a supercritical antisolvent gas into contact with a solution of the material in a solvent at a controlled rate operable to expand the solution and precipitate the material. The small particulate material formed in accordance with the invention is particularly useful in the preparation of devices that provide control of the rate of drug release.
Description
FORUIATION Of PROTEIN MICROPARTTCLES BY ANTISOLVENT
fRECIPIfATION
'1'he present invention relates to a method of forming protein microparticles using gas antisolvent precipitation and to composition of such proteins.
Conventional means of administering drugs te.g., pills and tablets) provide a single burst or peak of drug in the blood. This initial spike is followed by a decay in blood concentration. Because every drug has a range of concentration below which its therapeutic effect is limited, and above which toxic side effects occur, it is desirable to release the drug at a controlled rate and minimize fluctuations. In controlled release, this is achieved by incorporating a rate-limiting step into the design of the delivery system. Among the many types of controlled release systems there are bioerodible polymer microspheres in the range of 1 to 50 micrometers (um) . Such small mi.crospheres can be injected subcutaneously or intramuscularly.
Bioerodible polymers are materials that are degraded by body fluids to non-toxic products. The polymer particles contain the drug of interest in dispersed form. Drug release occurs partly as a result of polymer degradation inside the body.
Systems aimed at providing spatial or temporal control of drug release in the body are referred ~~~~~z~
to generically as controlled drug delivery devices.
Controlled release of proteins, such as therapeutic enzymes, requires the formation of small particles which can be uniformly dispersed in the polymer matrix. Techniques to produce protein particles include spray drying, lyophilization, milling, grinding, and protein micronisation, WO/90/132. Only the last method leads to small particles.
Jean W. Tom and Pablo G. Debenedetti, "Formation of Bioerodible Microspheres and Microparticles by Rapid Expansion of Supercritical Solutions", Department of Chemical Engineering, Princeton University, 1991, disclose a process to made biocompatible and bioerodible polymer microspheres, mainly polyhydroxy acids including poly(L-lactic acid) (L-PLA), poly(D,L-lactic acid), (DL-PLA) and poly(glycolic acid) (PGA).
Microparticles and microspheres of these polymers were made with the goal of being used for controlled delivery of pharmaceuticals.
Nucleation of poly(L-lactic acid) from C02 and C02-acetone mixtures produced micropa:rticles and microspheres ranging from 4 to 25 micrometers (um). Microspheres (2-20 um) were also obtained using chlorotrifluoromethane as a solvent.
The technique to produce the microspheres and microparticles used by Tom and Debenedetti involved applying rapid expansion of _2_ supercritical solutions. This was known from Matson et al., "Expansion of Supercritical Fluid Solutions: Solute Formation of Powders, Thin Films and Fibers". Ind. Eng. Chem. Res. 26, 2298-2306 (1987). In the process of rapid expansion of supercritical solutions, a nonvolatile solute is dissolved in a supercritical fluid. The resulting solution is highly compressible in the vicinity of the solvents critical point. Nucleation of the solute is triggered mechanically by reducing the solution's density through a rapid expansion, thereby reducing its dissolving capacity; Kumar et al., "Modeling the Solubility of Solids in Supercritical Fluids with Density as the Z5 Independent Variable", J. Supercrit, Fluids.
1988, l, 15-22. The combination of a rapidly propagating mechanical perturbation arid high supersaturation ratios leads to uniform conditions within the carrier fluid and hence, in principle, to narrow particle size distributions into small particles.
Chang et al., "Solvent Expansion axed Solute Solubility Predictions in Gas-Expanded Liquids", AIChE Journal, 36, No. 6, 939-942 (1990) disclose gas antisolvent addition for liquid phase precipitation of solids. See also Uallagher et al., "Gas (Gas Anti-Solvent) Recrystallization: A
New Process to Recrystallize Compounds Insoluble in Supercritical Fluids'°, Am. Chew.. Soc. Symp.
Ser., No. 406 (1989).
Chang et al, disclose recrystallization of acetaccu.no_pluen Lrom butanol and /~-carotene from toluene using C02. 'the C02 was charged at the top of the column or reservoir containing the solution to be gas expanded.
The present invention relates to a method of forming microparticles of a material, particularly protein microparticles, from a solution by antisolvent recrystallization using a supercritical gas. Tn accordance with the present invention, a supercritical antisolvent gas is brought into contact with a solution of the material in a solvent at a controlled rate operable to expand the solution and precipitate the material.
The present invention is particularly useful for the formation of protein microparticles, wherein the protein is a hydrophobic enzyme, (i.e., one that can be dissolved in a predominantly non-aqueous solvent).
Preferred proteins are selected from the group consisting of insulin, catalase, adrenocorticotrophin hormone and peroxidase.
Preferred solvents fo:r the protein are non-aqueous solvents selected from the group consisting of ethanol, dimethylsulfoxide, tetrahydrofuran, acetic acid, formamide, dimethylformamide, ethylene glycol, liquid polyethylene glycol, and dimethylanine. Preferred antisolvent gases are selected from the group consisting of carbon dioxide, ethane, ethylene, sulfur hexafluoride, 2~8j~~r nitrous oxide, chlorotrifluoromethane and monofluoromethane.
A preferred method of the present invention comprises passing the solution of a soluble material in a solvent through a continuum of supercritical antisolvent gas and precipitating the soluble material. This can be conducted by passing the solution through the continuum of gas phase in the form oL droplets, which can be sprayed through the gas phase. The plurality of droplets can be passed cocurrently or countercurrently with respect to a stream of antisolvent gas. Most preferably, the droplets are passed cocurrently. Alternatively, the solution can be passed through the continuum of supercritical antisolvent gas in the farm of a thin film or a plurality of fine streams.
In accordance with the method of the present invention, particularly by the use of a continuous phase of supercritical antisolvent gas, the liquid solution rapidly expands, causing the soluble material to precipitate out very rapidly.
This results in extremely uniform and fine particle sizes.
In accordance with the present invention, the soluble material, preferably protein, can be made having a particle size wherein the precipitated material has a particle size of less than 10, preferably less than 5, more preferably less than 3, and most preferably less ~~~~~?'~
than 1 micrometer in equivalent diameter.
Equivalent diameter is defined as the diameter of a sphere having the same volume as the particle would have an equivalent diameter as indicated.
Where the soluble material is glabular, such as globular protein, the equivalent diameter is preferably Less than 3 micrometers, and more preferably less than 1 micrometer. Wherein the soluble material precipitates out in a needle-like configuration, the average diameter of the needle is preferably less than 3 and more preferably less than 1 micrometer, and the length of the particle is less than 5 and preferably less than 3 micrometers. Typically, the particles of the present invention have a uniform and relatively narrow particle distribution.
The microparticles formed according to the present invention have a chemically uniform composition. Where more than one particle is made the blend is a controlled blend of two or more, preferably two or three different materials in controlled amounts and sizes.
'Phe small particulate material of the present invention is particularly useful in the preparation of devices that provide control of the rate of drug release. Such compositions comprise a bioerodible polymer and a plurality of very small active ingredients, such as proteins. The small size of the particles is important to insure a uniform dispersion within the polymeric matrix.
It is important that the polymer particles be 2~a~~r small (<50 pm) so that they can be injected. 'Che method of the present invention and proteins described in the present invention are therefore particularly useful in such formulations.
Formulations of particular importance are those made using a bioerodible polymeric matrix and at least one protein having an equivalent diameter or less than about 3 micrometers. Preferred polymers are polyhydroxy acids such as poly(L-lactic acid), poly(D,L-lactic acid) and poly(glycolic acid). The preferred composition comprises from 0.1 to 50 weight percent of protein and a corresponding amount of polymer matrix.
BR:Ci:F DI?:SCRIP'f I: ON OF THE DFtAwING5 Figure 1 is a schematic drawing of an experimental apparatus for the gas antisolvent recrystallization and liquid expansion useful in the present invention. In Figure 1 "V°' represents valves, "PI" represents pressure sensors, "TI"
represents temperature sensors. Back pressure regulators, rotameters filters, check valves, metering' valves, shut-off valves, rupture discs, and heat exchangers are depicted with conventional symbols.
Figure 2 is a microphotograph magnified 8,000 times showing catalase particles made in Example 1 the particles collected on the glass slides upside.
~~~~~1~,~
Figure 3 is a microphotograph of the same material as in Figure 2 having a magnification of 15,000.
Figure 4 is a microphotograph of the same material as in Figure 3 where the particles are collected on a filter and magnified 10,000 times.
Figure 5 is a microphotograph as recited in Figure 4 magnified 15,000 times.
Figure 6 is a microphotograph of the insulin partip cles made according to Example 2 magnified 10,000 times and collected an a filter.
Figure 7 is a microphotograph of insulin particles made in Example 2 magnified 5,500 times and collected on a filter.
Descri~i ion 1.5 Referring to Figure 1, a solution of dissolved mate-rial, preferably a protein solution, from solution source 14 (in which the solution can be prepared and/or stored) is fed to crystallizer l0. 'fhe solution is fed through suitable flow metering means, such as rotameter 30 and high pressure liquid pump 32. The pressure can be controlled using a back-pressure regulator 34. The solution can be brought to the desired temperature by a suitable heating means, such as coils 36, which are kept at the desired temperature by circulating air.
Heating can be provided by strip heaters and forced circulation of air.
The pressurized solution is fed to crystallizer chamber 20. Preferably, it is injected into the top of the crystallizer thrQUgh a laser-drilled platinum disc 53 to produce a fine spray of solution. droplets in the _g_ ~~1~3~~~~
crystallizes chamber 20. Disc 53 thus will have at least one orifice to produce a fine spray. Typical the orifice is from 5 to 50 and preferably from 10 to 30 and most preferably from 15 to 20 micrometers in diame--ter. Preferably the solution takes the form of a plu-rality of droplets having a diameter of from 10 to 500 Vim, at least one continuous fine stream having a diame-ter of less than 1 millimeter, or a thin film having a thickness of less than 1 millimeter.
An antisolvent gas is fed from antisolvent gas tank 18 to an antisolvent gas compression pump 40. The gas pressure can be controlled by a gas back-pressure regu-lator 42. The temperature of the antisolvent in tank 18 will range of from 10 to 40°C, preferably from 20 to 30'C, such as liquid carbon dioxide at 25°C. The anti-solvent is cooled in heat exchanger 44 with solvent pump 40 bringing the liquid antisolvent to supercriti-cal pressure. Excess solvent in the gas stream can recycle back to the liquid inlet side of the pump 40.
Typical conditions at the outlet of pump 40 are from to 45°C and more preferably from 30 to 40°C, and 60 to 200 atmosphere pressure, more preferably 100 to 150 atmosphere pressure. The compressed antisolvent is fed through suitable micrometering means, such as micro-25 metering valve 46. Optionally, there can be additional thermostating means such as coils 48 between solvent pump 40 and crystallizes 10. The supercritical anti solvent gas then is fed to crystallizes chamber 20 at a controlled rate so as to contain a continuum of super critical gas.
In crystallizes chamber 20 the supercritical anti-solvent gas dissolves in the protein solution at a con-trolled rate depending on the stream or droplet geome-_g_ ~~~~~% ~~
try, temperature and concentration. As solution eXpaIldS, the soluble material precipitates out. Use of a continuum of. supercritical fluid and passage of a fine stream, film, or droplets through the supercriti-cal gas results in rapid expansion of the liquid solu-tion and precipitation of the dissolved materialas extremely fine particles less than 10 micrometers in equivalent diameter and preferably less than 5, more preferably less than 3, and most preferably less than 1 micrometer. in diameter, particularly for soluble mate-rial which precipitates out in a globular shape. Nee-dle-like precipitates have a diameter of less than about 3 and more preferably less than 1 micrometer with a length of less than 5, and preferably less than 3 micrometers. The rapid precipitation results in a nar-row particle distribution as exemplified and shown in Figures 2 to 7.
The depleted solution and spent supercritical anti-solvent gas are fed to depressurization tank 22, to be brought back to ambient conditions. The precipitated crystals are collected from the crystallizer 10 at crystal collection port 26. The crystals can be col-lected by any suitable means, such as on a filter and/or a glass plate.
A fluid mixture of spent solvent and supercritical fluid can be collected from the bottom of the crystal-lizes chamber 20 through line 50. The fluid mixture passes through valve V7 to collection tank 52 through valve V4 to depressurization tank 22 where the mixture is depressurized and expanded to atmospheric pressure.
The system should be sized to handle pressures of up to 6000 psi, and preferably in the range of from atmo-~~8~~~?'~
spheric pressure to 6,000 psi from temperatures ranging from 20°C to 60°C and preferably 30°C to 50°C.
Different flow patterns can be used in crystallizer 1Ø The direction of antisolvent supercritical gas flow in the crystallizes (upward or downward) can be determined by the valves before and after the crystal-lizes. For upward flow in the crystallizes, valve V2 and valve V3 are open, and valve V1 and valve V4 are closed. Where the antisolvent fluid is to flow down-ward the valves are reversed. The flow of protein solution can be cocurrent or countercurrent to the flow of antisolvent fluid. In the preferred embodiment with continuous operation, protein solution is pumped into the cxystallizer by high pressure liquid pump 32 and its instantaneous flow rate is measured by the liquid rotameter 30. The pressure is controlled using a back-pressure regulator 34 and pressurized protein solution is injected into the top of the crystallizes. The antisolvent gas is also injected into the top of the crystallizes for cocurrent flow of both the supercriti-cal solvent fluid continuum through the crystallizes and the protein solution through the crystallizes, both from top to bottom. 'fhe crystallizes can be operated in batch, semi-batch or continuous operation. The solution of soluble material can be passed through cocurrently or countercurrently in relation to a con-tinuum stream of antisolvent supercritical gas.
Suitable soluble material are protein, particularly hydrophobic enzymes such as insulin, catalase, adreno-corticotrophin hormone, and peroxidase. The method has applicability, however, to virtually any protein and is not dependent on chemical structure or biological activity.
2i~83=~~' Useful solutions for the protein comprise at least one non-aqueous solvent suchs as ethanol, formamide, dimethylsulfoxide, tetrahydrofuran, acetic acid, dimethylformamide, ethylene glycol, liquid polyethylene glycol and dimethylaniline.
Supercritical gases which can be used include (with an indication of its critical temperatures (°C) and critical pressures (atm)} include ethane (32.2°C, 48.1 atm), ethylene (9.21°C, 39.7 atm); sulfur hexafluoride (45.5°C, 37.1 atm), nitrous oxide (36.5°G, 71.7 atm) chlorotrifluoromethane (28°C, 38.7 atm), and mono fluoromethane (44.5°C, 58 atm). A solution of water and ethanol has been used. However, the presence of water in such solutions has been found to lower the production of small particle protein.
In accordance with the present invention there is obtained a protein composition having protein particles wherein substantially all of the protein particles are artificially isolated and have an equivalent diameter of less than 5, more preferably less than 3, and most preferably less than 1 micrometer. The protein compo-sition has a narrow particle distribution shown in lr'ig-ures 2-7. These proteins have uniform or controlled chemical compositions. Therefore, samples of a compo-sition consisting essentially of a desired protein can be isolated and made.
The isolated proteins of the present invention can be used to make temporal drug release compositions.
Such compositions can comprise a bioerodible polymeric matrix and at least one protein having an equivalent diameter of less than 3 micrometers. Preferred poly-mers are polyhydroxy acids such as those selected from the group consisting of poly(L-lactic acid), poly(D,L-2~~~L~r lactic acid) and poly(glycolic acid). The composition can comprise for 0.1 to 50 weight percent of the pro-tein.
Preferably the drug release compositions contain a polymer matrix having a continuum of bioerodible poly-mer matrix with the protein particles dispersed there-with. Such particles can be made by means known in the art as discussed above.
Following are several Examples which illustrate the nature of the invention and 'the manner of carrying it out.
Referring to Figure 1, liquid carbon dioxide in sol-vent tank 18 was compressed by high pressure liquid pump 40 which was an American Lewa Plunger Metering Pump, Model EL-1; rated at 6,000 psi and 2 gallons per hour. The pressure was controlled by a back-pressure regulator 42 which was a Tescom, Model 54-2100 Series, rated at 6,000 psi. The compressed carbon dioxide was introduced into a see-through crystallizer 20 which was a Jerguson Gauge, Model 19T40, 316 stainless steel 5,000 psi, 1.3 centimeter by 1.3 centimeter, 31.8 cen timeter long, 50 cubic centimeter through mic.rometering valve 46 which was an Autoclave Engineering Micrometer ing Valve 60VRMM:. The pressurized carbon dioxide was preheated in coiled tubes 48.
The pressure in the crystalli~er was indicated by a crystallizer pressure gauge 54 which was a Bourdon Gauge, Omega Model PGJ-45B-5000, rated at 5000 psi, and controlled by back-pressure regulator 59 which was a Tescom 26 -7.700 Series, rated at 6,000 psi.
The protein solution from protein solution tank 14 was pumped in continuous operation by a high pressure pump which was a Milton Roy LDC Duplex Metering Pump.
The instantaneous flow was measured by liquid rotameter 30 which was a Fischer and Porter; Model 10A6132, 0-14 cubic centimeters per minute of water flow. The pres sure of the protein solution was controlled using a bank-pressure regulator 34 which was a Tescom; 26-1700 Series, 10,000 psi rated regulator. The protein solu tion was preheated in coiled tubes 36.
The pressurized protein solution was injected into the top of the crystallizes 10 through a laser-drilled platinum disc 53, Ted Pella; 3 mm OD x 0.24 mm thick;
micrometers in diameter.
15 At the bottom of the crystallizes the protein parti-cles were precipitated and deposited on an inclined glass slide after crystallization. The plane of the glass slide 55 was at a l0° angle to the direction of the protein solution flow. Additionally, a filter 57, 20 Mott Metallurgical? 316 Stainless Steel 1.6 centimeters in diameter, 0.5-2 micrometer pore size was located below the glass slide to collect all the protein parti cles. A thermocouple 56, Omega Engineering Type J, was placed in the middle of the crystallizes to monitor the temperature.
Protein particles collected on the glass slides were examined through a Carl Zeiss Universal Optical Micro-scope and a Scanning Electron Microscopy JEOL JSM-840A, with samples coated with gold-palladium. The particles on the microfilter were also examined with the Scanning Electron Microscope.
~. r5 The fluid mixture of carbon dioxide, ethanol and water coming out of the crystallizer was depressurized and expanded to atmospheric pressure by passing through a cylindrical depressurizing tank 22, Swagelok, 150 ml, 5,000 psi and back-pressure regulator 5g, Tescam, 26°
1700 Series, rated at 6,000 psi.
The instantaneous and total flow rates of solute free C02 gas were measured with rotameter 60 (Fischer and Porter; Model 10A4555, 0-3.35 SCFM AIR and dry test meter 62, American Meter: Model DTM200A, respectively.
During the experiment the normal flow rates of pro-tein solution and antisolvent gas were 0.35 cm3/min and 35g/min, respectively, and typical operating time was 4 hours for continuous operation. The whole system was enclosed in an air chamber where temperature was con-trolled using a PID temperature controller, Omega Engi-neering Model C~I9000, and strip heaters.
To measure the expansion behavior of C02-ethanol solution, 20 mls of ethanol solution was preloaded into the crystallizer and pressure was increased by 200 psi increments through valves V2 and V7. Gas solvent was then circulated through valve V8, crystallizer 20 and valve V5 using a high pressure compressor (Haskell;
Dauble Acting Single Stage Model ACD-62) with closed valves V6 and Vii until the system reached the equilib rium state and the liquid level remained constant.
Example 1 Catalase particles, Figures 2-5, were made having an equivalent diameter of less than 1 Im.
A solution of 20 mg catalase (from bovine liver) [Sigma Chemicals C-40] in 200 ml of 90% ethanol (Pharmco Products Co., 200 proof) and 10~ water (deionized through a reverse osmosis apparatus, Hydro Picosystem) was used. The pH of the solution was adjusted to 3.22 with hydrochloric acid.
Liquid carbon dioxide (MG industriest Hone-dry grade, >93.8~) was compressed by a high pressure liquid pump 40. The delivery pressure (1600 psi) was con-trolled by a back-pressure regulator 42. The pressur-ized liquid was pre-heated to a supercritical tempera-ture (35°C) and flowed through coiled tube 48 before entering the crystallizes 10. The system was enclosed and thermostating was achieved by circulating.hot air under temperature control (Omega Engineering Model CN9000), with heating provided by strip heaters. The see-through crystallizes chamber 20 was kept at 35°C.
The supercritical fluid was fed to the crystallizes through a micrometering valve.46, with valves V1 and V4 open; and V2 and V3 closed. The pressure inside the crystallizes was kept at 1300 psi by back-pressure reg-ulator 59. The instantaneous and total flow rates of supercritical fluid were measured with rotameter 60 and dry test meter 62, respectively. The flow rate of the supercritical fluid was 33 g/min.
The liquid solution containing the enzyme was pres-surized and circulated by liquid pump 32 and back-pres-sure regulator control (1430-2530 psi). The solution circulated through coil 36 and was preheated to 35°C.
It entered the top of the crystallizes through a laser--drilled platinum disc (Ted Pella: 3 mm OD x 0.24 mm thicke 20 ~Cm), and emerged as very small. droplets. The liquid flow rate was 0.35 cc/min. The liquid and supercritical streams circulated cocurrently downwards.
~083f~'l The fluid mixture of carbon dioxide, ethanol and water exiting the crystallizes was depressurized and expanded to atmospheric pressure by flowing through cylindrical depressurizing tank 22, and back-pressure regulator 58e The supercritical fluid expanded and eventually dis-solved most of the liquid solvent, causing the enzyme particles to precipitate. The particles were collected on an inclined glass slide located at the bottom of the crystallizes, forming an angle of approximately 10° to the direction of the protein solution's flow. Parti-cles were also collected on a filter (Mott Metallurgi-cal; 316 Stainless Steel, 1.6 cm diameter, 0.5 ~.m pore size). The carbon dioxide outlet was located approxi-mately 8 cm above the filter. The duration of the experiment was 260 minutes.
Figures 2 and 3 are particles collected on the glass slide's up side (facing the nozzle) . Figures 4 and 5 are particles collected on the filter.
Example 2 The pH of a solution of 20 mg zinc insulin [Miles;
low endotoxin 86--003J in 200 ml of 90% ethanol (Pharmco Products Co., 200 proof)-10% water (deionized through a reverse osmosis apparatus, Hydro Picosystem) was adjusted to 2.56 with hydrochloric acid.
Liquid carbon dioxide (MG industries; Bone-dry grade,>99.8%) was compressed by a high pressure liquid pump 40. The delivery pressure (2000 psi) was con-trolled by a back-pressure regulator 42. The pressur-ized liquid was pre-heated to a supercritical tempera-ture (35°C) as it flowed through coiled tube 48 before entering the crystallizes 10. The system was enclosed, and thermostatting was achieved by circulating hot air under temperature control (Omega Engineering Model CN~000), with heating provided by strip heaters. The see-through crystallizes chamber 20 was kept at 35°C.
The supercritical fluid was fed to the crystallizes through a micrometering valve 46, with valves 1 and 4 openp and 2 and 3 closed. The pressure inside the crystallizes was kept at 1300 psi by back-pressure reg-ulator 59. The instantaneous and total flow rates of supercritical fluid were measured with a rotameter 60 and dry test meter 62, respectively. The flow rate of the supercritical fluid was 35.6 g/min.
The liquid solution containing the enzyme was pres-surized and circulated by liquid pump 32 under back-pressure regulator 34 control (1450 psi). The solution circulated through coil 36 and was preheated to 35°C.
It entered the top of the crystallizes through a laser-drilled platinum disc (Ted Pella: 3 mm OD x 0.24 mm thick) 20 um), and emerged as very small droplets. The liquid flow rate was 0.39 cc/min. The liquid and supercritical streams circulated cocurrently downwards.
The fluid mixture of carbon dioxide and water exit-ing the crystallizes was depressurized and expanded to atmospheric pressure by flowing through cylindrical depressurizing tank 22 and a back-pressure regulator 58.
The supercritical fh::id expanded and eventually dis-solved most of the liquid solvent, causing the enzyme particles to precipitate. The particles were collected on an inclined glass slide located at the bottom of the crystallizes, forming an angle of approximately 10° to the direction of the protein solution's flow. Parti-cles were also collected on a filter (Mott Metallurgi-cal; 316 Stainless Steel, 1.6 cm diameter, 0.5 /gym pore size). The carbon dioxide outlet was located approxi-mately 60 mm below the filter.
The duration of the experiment was 296 minutes for carbon dioxide input, arid 237 minutes for liquid input, followed by 17 minutes of liquid solution flow without dissolved enzyme.
Figures 6 illustrates particles collected on the filter which were needlelike having a diameter of less than 1 ~Cm and being less than 3 Vim. Figure 7 illus-trates particles collected which were globular, having an equivalent diameter of less than about 1 ~.m.
6~lhile exemplary embodiments of this invention have been described, the true scope of the invention is determined fram the following claims.
fRECIPIfATION
'1'he present invention relates to a method of forming protein microparticles using gas antisolvent precipitation and to composition of such proteins.
Conventional means of administering drugs te.g., pills and tablets) provide a single burst or peak of drug in the blood. This initial spike is followed by a decay in blood concentration. Because every drug has a range of concentration below which its therapeutic effect is limited, and above which toxic side effects occur, it is desirable to release the drug at a controlled rate and minimize fluctuations. In controlled release, this is achieved by incorporating a rate-limiting step into the design of the delivery system. Among the many types of controlled release systems there are bioerodible polymer microspheres in the range of 1 to 50 micrometers (um) . Such small mi.crospheres can be injected subcutaneously or intramuscularly.
Bioerodible polymers are materials that are degraded by body fluids to non-toxic products. The polymer particles contain the drug of interest in dispersed form. Drug release occurs partly as a result of polymer degradation inside the body.
Systems aimed at providing spatial or temporal control of drug release in the body are referred ~~~~~z~
to generically as controlled drug delivery devices.
Controlled release of proteins, such as therapeutic enzymes, requires the formation of small particles which can be uniformly dispersed in the polymer matrix. Techniques to produce protein particles include spray drying, lyophilization, milling, grinding, and protein micronisation, WO/90/132. Only the last method leads to small particles.
Jean W. Tom and Pablo G. Debenedetti, "Formation of Bioerodible Microspheres and Microparticles by Rapid Expansion of Supercritical Solutions", Department of Chemical Engineering, Princeton University, 1991, disclose a process to made biocompatible and bioerodible polymer microspheres, mainly polyhydroxy acids including poly(L-lactic acid) (L-PLA), poly(D,L-lactic acid), (DL-PLA) and poly(glycolic acid) (PGA).
Microparticles and microspheres of these polymers were made with the goal of being used for controlled delivery of pharmaceuticals.
Nucleation of poly(L-lactic acid) from C02 and C02-acetone mixtures produced micropa:rticles and microspheres ranging from 4 to 25 micrometers (um). Microspheres (2-20 um) were also obtained using chlorotrifluoromethane as a solvent.
The technique to produce the microspheres and microparticles used by Tom and Debenedetti involved applying rapid expansion of _2_ supercritical solutions. This was known from Matson et al., "Expansion of Supercritical Fluid Solutions: Solute Formation of Powders, Thin Films and Fibers". Ind. Eng. Chem. Res. 26, 2298-2306 (1987). In the process of rapid expansion of supercritical solutions, a nonvolatile solute is dissolved in a supercritical fluid. The resulting solution is highly compressible in the vicinity of the solvents critical point. Nucleation of the solute is triggered mechanically by reducing the solution's density through a rapid expansion, thereby reducing its dissolving capacity; Kumar et al., "Modeling the Solubility of Solids in Supercritical Fluids with Density as the Z5 Independent Variable", J. Supercrit, Fluids.
1988, l, 15-22. The combination of a rapidly propagating mechanical perturbation arid high supersaturation ratios leads to uniform conditions within the carrier fluid and hence, in principle, to narrow particle size distributions into small particles.
Chang et al., "Solvent Expansion axed Solute Solubility Predictions in Gas-Expanded Liquids", AIChE Journal, 36, No. 6, 939-942 (1990) disclose gas antisolvent addition for liquid phase precipitation of solids. See also Uallagher et al., "Gas (Gas Anti-Solvent) Recrystallization: A
New Process to Recrystallize Compounds Insoluble in Supercritical Fluids'°, Am. Chew.. Soc. Symp.
Ser., No. 406 (1989).
Chang et al, disclose recrystallization of acetaccu.no_pluen Lrom butanol and /~-carotene from toluene using C02. 'the C02 was charged at the top of the column or reservoir containing the solution to be gas expanded.
The present invention relates to a method of forming microparticles of a material, particularly protein microparticles, from a solution by antisolvent recrystallization using a supercritical gas. Tn accordance with the present invention, a supercritical antisolvent gas is brought into contact with a solution of the material in a solvent at a controlled rate operable to expand the solution and precipitate the material.
The present invention is particularly useful for the formation of protein microparticles, wherein the protein is a hydrophobic enzyme, (i.e., one that can be dissolved in a predominantly non-aqueous solvent).
Preferred proteins are selected from the group consisting of insulin, catalase, adrenocorticotrophin hormone and peroxidase.
Preferred solvents fo:r the protein are non-aqueous solvents selected from the group consisting of ethanol, dimethylsulfoxide, tetrahydrofuran, acetic acid, formamide, dimethylformamide, ethylene glycol, liquid polyethylene glycol, and dimethylanine. Preferred antisolvent gases are selected from the group consisting of carbon dioxide, ethane, ethylene, sulfur hexafluoride, 2~8j~~r nitrous oxide, chlorotrifluoromethane and monofluoromethane.
A preferred method of the present invention comprises passing the solution of a soluble material in a solvent through a continuum of supercritical antisolvent gas and precipitating the soluble material. This can be conducted by passing the solution through the continuum of gas phase in the form oL droplets, which can be sprayed through the gas phase. The plurality of droplets can be passed cocurrently or countercurrently with respect to a stream of antisolvent gas. Most preferably, the droplets are passed cocurrently. Alternatively, the solution can be passed through the continuum of supercritical antisolvent gas in the farm of a thin film or a plurality of fine streams.
In accordance with the method of the present invention, particularly by the use of a continuous phase of supercritical antisolvent gas, the liquid solution rapidly expands, causing the soluble material to precipitate out very rapidly.
This results in extremely uniform and fine particle sizes.
In accordance with the present invention, the soluble material, preferably protein, can be made having a particle size wherein the precipitated material has a particle size of less than 10, preferably less than 5, more preferably less than 3, and most preferably less ~~~~~?'~
than 1 micrometer in equivalent diameter.
Equivalent diameter is defined as the diameter of a sphere having the same volume as the particle would have an equivalent diameter as indicated.
Where the soluble material is glabular, such as globular protein, the equivalent diameter is preferably Less than 3 micrometers, and more preferably less than 1 micrometer. Wherein the soluble material precipitates out in a needle-like configuration, the average diameter of the needle is preferably less than 3 and more preferably less than 1 micrometer, and the length of the particle is less than 5 and preferably less than 3 micrometers. Typically, the particles of the present invention have a uniform and relatively narrow particle distribution.
The microparticles formed according to the present invention have a chemically uniform composition. Where more than one particle is made the blend is a controlled blend of two or more, preferably two or three different materials in controlled amounts and sizes.
'Phe small particulate material of the present invention is particularly useful in the preparation of devices that provide control of the rate of drug release. Such compositions comprise a bioerodible polymer and a plurality of very small active ingredients, such as proteins. The small size of the particles is important to insure a uniform dispersion within the polymeric matrix.
It is important that the polymer particles be 2~a~~r small (<50 pm) so that they can be injected. 'Che method of the present invention and proteins described in the present invention are therefore particularly useful in such formulations.
Formulations of particular importance are those made using a bioerodible polymeric matrix and at least one protein having an equivalent diameter or less than about 3 micrometers. Preferred polymers are polyhydroxy acids such as poly(L-lactic acid), poly(D,L-lactic acid) and poly(glycolic acid). The preferred composition comprises from 0.1 to 50 weight percent of protein and a corresponding amount of polymer matrix.
BR:Ci:F DI?:SCRIP'f I: ON OF THE DFtAwING5 Figure 1 is a schematic drawing of an experimental apparatus for the gas antisolvent recrystallization and liquid expansion useful in the present invention. In Figure 1 "V°' represents valves, "PI" represents pressure sensors, "TI"
represents temperature sensors. Back pressure regulators, rotameters filters, check valves, metering' valves, shut-off valves, rupture discs, and heat exchangers are depicted with conventional symbols.
Figure 2 is a microphotograph magnified 8,000 times showing catalase particles made in Example 1 the particles collected on the glass slides upside.
~~~~~1~,~
Figure 3 is a microphotograph of the same material as in Figure 2 having a magnification of 15,000.
Figure 4 is a microphotograph of the same material as in Figure 3 where the particles are collected on a filter and magnified 10,000 times.
Figure 5 is a microphotograph as recited in Figure 4 magnified 15,000 times.
Figure 6 is a microphotograph of the insulin partip cles made according to Example 2 magnified 10,000 times and collected an a filter.
Figure 7 is a microphotograph of insulin particles made in Example 2 magnified 5,500 times and collected on a filter.
Descri~i ion 1.5 Referring to Figure 1, a solution of dissolved mate-rial, preferably a protein solution, from solution source 14 (in which the solution can be prepared and/or stored) is fed to crystallizer l0. 'fhe solution is fed through suitable flow metering means, such as rotameter 30 and high pressure liquid pump 32. The pressure can be controlled using a back-pressure regulator 34. The solution can be brought to the desired temperature by a suitable heating means, such as coils 36, which are kept at the desired temperature by circulating air.
Heating can be provided by strip heaters and forced circulation of air.
The pressurized solution is fed to crystallizer chamber 20. Preferably, it is injected into the top of the crystallizer thrQUgh a laser-drilled platinum disc 53 to produce a fine spray of solution. droplets in the _g_ ~~1~3~~~~
crystallizes chamber 20. Disc 53 thus will have at least one orifice to produce a fine spray. Typical the orifice is from 5 to 50 and preferably from 10 to 30 and most preferably from 15 to 20 micrometers in diame--ter. Preferably the solution takes the form of a plu-rality of droplets having a diameter of from 10 to 500 Vim, at least one continuous fine stream having a diame-ter of less than 1 millimeter, or a thin film having a thickness of less than 1 millimeter.
An antisolvent gas is fed from antisolvent gas tank 18 to an antisolvent gas compression pump 40. The gas pressure can be controlled by a gas back-pressure regu-lator 42. The temperature of the antisolvent in tank 18 will range of from 10 to 40°C, preferably from 20 to 30'C, such as liquid carbon dioxide at 25°C. The anti-solvent is cooled in heat exchanger 44 with solvent pump 40 bringing the liquid antisolvent to supercriti-cal pressure. Excess solvent in the gas stream can recycle back to the liquid inlet side of the pump 40.
Typical conditions at the outlet of pump 40 are from to 45°C and more preferably from 30 to 40°C, and 60 to 200 atmosphere pressure, more preferably 100 to 150 atmosphere pressure. The compressed antisolvent is fed through suitable micrometering means, such as micro-25 metering valve 46. Optionally, there can be additional thermostating means such as coils 48 between solvent pump 40 and crystallizes 10. The supercritical anti solvent gas then is fed to crystallizes chamber 20 at a controlled rate so as to contain a continuum of super critical gas.
In crystallizes chamber 20 the supercritical anti-solvent gas dissolves in the protein solution at a con-trolled rate depending on the stream or droplet geome-_g_ ~~~~~% ~~
try, temperature and concentration. As solution eXpaIldS, the soluble material precipitates out. Use of a continuum of. supercritical fluid and passage of a fine stream, film, or droplets through the supercriti-cal gas results in rapid expansion of the liquid solu-tion and precipitation of the dissolved materialas extremely fine particles less than 10 micrometers in equivalent diameter and preferably less than 5, more preferably less than 3, and most preferably less than 1 micrometer. in diameter, particularly for soluble mate-rial which precipitates out in a globular shape. Nee-dle-like precipitates have a diameter of less than about 3 and more preferably less than 1 micrometer with a length of less than 5, and preferably less than 3 micrometers. The rapid precipitation results in a nar-row particle distribution as exemplified and shown in Figures 2 to 7.
The depleted solution and spent supercritical anti-solvent gas are fed to depressurization tank 22, to be brought back to ambient conditions. The precipitated crystals are collected from the crystallizer 10 at crystal collection port 26. The crystals can be col-lected by any suitable means, such as on a filter and/or a glass plate.
A fluid mixture of spent solvent and supercritical fluid can be collected from the bottom of the crystal-lizes chamber 20 through line 50. The fluid mixture passes through valve V7 to collection tank 52 through valve V4 to depressurization tank 22 where the mixture is depressurized and expanded to atmospheric pressure.
The system should be sized to handle pressures of up to 6000 psi, and preferably in the range of from atmo-~~8~~~?'~
spheric pressure to 6,000 psi from temperatures ranging from 20°C to 60°C and preferably 30°C to 50°C.
Different flow patterns can be used in crystallizer 1Ø The direction of antisolvent supercritical gas flow in the crystallizes (upward or downward) can be determined by the valves before and after the crystal-lizes. For upward flow in the crystallizes, valve V2 and valve V3 are open, and valve V1 and valve V4 are closed. Where the antisolvent fluid is to flow down-ward the valves are reversed. The flow of protein solution can be cocurrent or countercurrent to the flow of antisolvent fluid. In the preferred embodiment with continuous operation, protein solution is pumped into the cxystallizer by high pressure liquid pump 32 and its instantaneous flow rate is measured by the liquid rotameter 30. The pressure is controlled using a back-pressure regulator 34 and pressurized protein solution is injected into the top of the crystallizes. The antisolvent gas is also injected into the top of the crystallizes for cocurrent flow of both the supercriti-cal solvent fluid continuum through the crystallizes and the protein solution through the crystallizes, both from top to bottom. 'fhe crystallizes can be operated in batch, semi-batch or continuous operation. The solution of soluble material can be passed through cocurrently or countercurrently in relation to a con-tinuum stream of antisolvent supercritical gas.
Suitable soluble material are protein, particularly hydrophobic enzymes such as insulin, catalase, adreno-corticotrophin hormone, and peroxidase. The method has applicability, however, to virtually any protein and is not dependent on chemical structure or biological activity.
2i~83=~~' Useful solutions for the protein comprise at least one non-aqueous solvent suchs as ethanol, formamide, dimethylsulfoxide, tetrahydrofuran, acetic acid, dimethylformamide, ethylene glycol, liquid polyethylene glycol and dimethylaniline.
Supercritical gases which can be used include (with an indication of its critical temperatures (°C) and critical pressures (atm)} include ethane (32.2°C, 48.1 atm), ethylene (9.21°C, 39.7 atm); sulfur hexafluoride (45.5°C, 37.1 atm), nitrous oxide (36.5°G, 71.7 atm) chlorotrifluoromethane (28°C, 38.7 atm), and mono fluoromethane (44.5°C, 58 atm). A solution of water and ethanol has been used. However, the presence of water in such solutions has been found to lower the production of small particle protein.
In accordance with the present invention there is obtained a protein composition having protein particles wherein substantially all of the protein particles are artificially isolated and have an equivalent diameter of less than 5, more preferably less than 3, and most preferably less than 1 micrometer. The protein compo-sition has a narrow particle distribution shown in lr'ig-ures 2-7. These proteins have uniform or controlled chemical compositions. Therefore, samples of a compo-sition consisting essentially of a desired protein can be isolated and made.
The isolated proteins of the present invention can be used to make temporal drug release compositions.
Such compositions can comprise a bioerodible polymeric matrix and at least one protein having an equivalent diameter of less than 3 micrometers. Preferred poly-mers are polyhydroxy acids such as those selected from the group consisting of poly(L-lactic acid), poly(D,L-2~~~L~r lactic acid) and poly(glycolic acid). The composition can comprise for 0.1 to 50 weight percent of the pro-tein.
Preferably the drug release compositions contain a polymer matrix having a continuum of bioerodible poly-mer matrix with the protein particles dispersed there-with. Such particles can be made by means known in the art as discussed above.
Following are several Examples which illustrate the nature of the invention and 'the manner of carrying it out.
Referring to Figure 1, liquid carbon dioxide in sol-vent tank 18 was compressed by high pressure liquid pump 40 which was an American Lewa Plunger Metering Pump, Model EL-1; rated at 6,000 psi and 2 gallons per hour. The pressure was controlled by a back-pressure regulator 42 which was a Tescom, Model 54-2100 Series, rated at 6,000 psi. The compressed carbon dioxide was introduced into a see-through crystallizer 20 which was a Jerguson Gauge, Model 19T40, 316 stainless steel 5,000 psi, 1.3 centimeter by 1.3 centimeter, 31.8 cen timeter long, 50 cubic centimeter through mic.rometering valve 46 which was an Autoclave Engineering Micrometer ing Valve 60VRMM:. The pressurized carbon dioxide was preheated in coiled tubes 48.
The pressure in the crystalli~er was indicated by a crystallizer pressure gauge 54 which was a Bourdon Gauge, Omega Model PGJ-45B-5000, rated at 5000 psi, and controlled by back-pressure regulator 59 which was a Tescom 26 -7.700 Series, rated at 6,000 psi.
The protein solution from protein solution tank 14 was pumped in continuous operation by a high pressure pump which was a Milton Roy LDC Duplex Metering Pump.
The instantaneous flow was measured by liquid rotameter 30 which was a Fischer and Porter; Model 10A6132, 0-14 cubic centimeters per minute of water flow. The pres sure of the protein solution was controlled using a bank-pressure regulator 34 which was a Tescom; 26-1700 Series, 10,000 psi rated regulator. The protein solu tion was preheated in coiled tubes 36.
The pressurized protein solution was injected into the top of the crystallizes 10 through a laser-drilled platinum disc 53, Ted Pella; 3 mm OD x 0.24 mm thick;
micrometers in diameter.
15 At the bottom of the crystallizes the protein parti-cles were precipitated and deposited on an inclined glass slide after crystallization. The plane of the glass slide 55 was at a l0° angle to the direction of the protein solution flow. Additionally, a filter 57, 20 Mott Metallurgical? 316 Stainless Steel 1.6 centimeters in diameter, 0.5-2 micrometer pore size was located below the glass slide to collect all the protein parti cles. A thermocouple 56, Omega Engineering Type J, was placed in the middle of the crystallizes to monitor the temperature.
Protein particles collected on the glass slides were examined through a Carl Zeiss Universal Optical Micro-scope and a Scanning Electron Microscopy JEOL JSM-840A, with samples coated with gold-palladium. The particles on the microfilter were also examined with the Scanning Electron Microscope.
~. r5 The fluid mixture of carbon dioxide, ethanol and water coming out of the crystallizer was depressurized and expanded to atmospheric pressure by passing through a cylindrical depressurizing tank 22, Swagelok, 150 ml, 5,000 psi and back-pressure regulator 5g, Tescam, 26°
1700 Series, rated at 6,000 psi.
The instantaneous and total flow rates of solute free C02 gas were measured with rotameter 60 (Fischer and Porter; Model 10A4555, 0-3.35 SCFM AIR and dry test meter 62, American Meter: Model DTM200A, respectively.
During the experiment the normal flow rates of pro-tein solution and antisolvent gas were 0.35 cm3/min and 35g/min, respectively, and typical operating time was 4 hours for continuous operation. The whole system was enclosed in an air chamber where temperature was con-trolled using a PID temperature controller, Omega Engi-neering Model C~I9000, and strip heaters.
To measure the expansion behavior of C02-ethanol solution, 20 mls of ethanol solution was preloaded into the crystallizer and pressure was increased by 200 psi increments through valves V2 and V7. Gas solvent was then circulated through valve V8, crystallizer 20 and valve V5 using a high pressure compressor (Haskell;
Dauble Acting Single Stage Model ACD-62) with closed valves V6 and Vii until the system reached the equilib rium state and the liquid level remained constant.
Example 1 Catalase particles, Figures 2-5, were made having an equivalent diameter of less than 1 Im.
A solution of 20 mg catalase (from bovine liver) [Sigma Chemicals C-40] in 200 ml of 90% ethanol (Pharmco Products Co., 200 proof) and 10~ water (deionized through a reverse osmosis apparatus, Hydro Picosystem) was used. The pH of the solution was adjusted to 3.22 with hydrochloric acid.
Liquid carbon dioxide (MG industriest Hone-dry grade, >93.8~) was compressed by a high pressure liquid pump 40. The delivery pressure (1600 psi) was con-trolled by a back-pressure regulator 42. The pressur-ized liquid was pre-heated to a supercritical tempera-ture (35°C) and flowed through coiled tube 48 before entering the crystallizes 10. The system was enclosed and thermostating was achieved by circulating.hot air under temperature control (Omega Engineering Model CN9000), with heating provided by strip heaters. The see-through crystallizes chamber 20 was kept at 35°C.
The supercritical fluid was fed to the crystallizes through a micrometering valve.46, with valves V1 and V4 open; and V2 and V3 closed. The pressure inside the crystallizes was kept at 1300 psi by back-pressure reg-ulator 59. The instantaneous and total flow rates of supercritical fluid were measured with rotameter 60 and dry test meter 62, respectively. The flow rate of the supercritical fluid was 33 g/min.
The liquid solution containing the enzyme was pres-surized and circulated by liquid pump 32 and back-pres-sure regulator control (1430-2530 psi). The solution circulated through coil 36 and was preheated to 35°C.
It entered the top of the crystallizes through a laser--drilled platinum disc (Ted Pella: 3 mm OD x 0.24 mm thicke 20 ~Cm), and emerged as very small. droplets. The liquid flow rate was 0.35 cc/min. The liquid and supercritical streams circulated cocurrently downwards.
~083f~'l The fluid mixture of carbon dioxide, ethanol and water exiting the crystallizes was depressurized and expanded to atmospheric pressure by flowing through cylindrical depressurizing tank 22, and back-pressure regulator 58e The supercritical fluid expanded and eventually dis-solved most of the liquid solvent, causing the enzyme particles to precipitate. The particles were collected on an inclined glass slide located at the bottom of the crystallizes, forming an angle of approximately 10° to the direction of the protein solution's flow. Parti-cles were also collected on a filter (Mott Metallurgi-cal; 316 Stainless Steel, 1.6 cm diameter, 0.5 ~.m pore size). The carbon dioxide outlet was located approxi-mately 8 cm above the filter. The duration of the experiment was 260 minutes.
Figures 2 and 3 are particles collected on the glass slide's up side (facing the nozzle) . Figures 4 and 5 are particles collected on the filter.
Example 2 The pH of a solution of 20 mg zinc insulin [Miles;
low endotoxin 86--003J in 200 ml of 90% ethanol (Pharmco Products Co., 200 proof)-10% water (deionized through a reverse osmosis apparatus, Hydro Picosystem) was adjusted to 2.56 with hydrochloric acid.
Liquid carbon dioxide (MG industries; Bone-dry grade,>99.8%) was compressed by a high pressure liquid pump 40. The delivery pressure (2000 psi) was con-trolled by a back-pressure regulator 42. The pressur-ized liquid was pre-heated to a supercritical tempera-ture (35°C) as it flowed through coiled tube 48 before entering the crystallizes 10. The system was enclosed, and thermostatting was achieved by circulating hot air under temperature control (Omega Engineering Model CN~000), with heating provided by strip heaters. The see-through crystallizes chamber 20 was kept at 35°C.
The supercritical fluid was fed to the crystallizes through a micrometering valve 46, with valves 1 and 4 openp and 2 and 3 closed. The pressure inside the crystallizes was kept at 1300 psi by back-pressure reg-ulator 59. The instantaneous and total flow rates of supercritical fluid were measured with a rotameter 60 and dry test meter 62, respectively. The flow rate of the supercritical fluid was 35.6 g/min.
The liquid solution containing the enzyme was pres-surized and circulated by liquid pump 32 under back-pressure regulator 34 control (1450 psi). The solution circulated through coil 36 and was preheated to 35°C.
It entered the top of the crystallizes through a laser-drilled platinum disc (Ted Pella: 3 mm OD x 0.24 mm thick) 20 um), and emerged as very small droplets. The liquid flow rate was 0.39 cc/min. The liquid and supercritical streams circulated cocurrently downwards.
The fluid mixture of carbon dioxide and water exit-ing the crystallizes was depressurized and expanded to atmospheric pressure by flowing through cylindrical depressurizing tank 22 and a back-pressure regulator 58.
The supercritical fh::id expanded and eventually dis-solved most of the liquid solvent, causing the enzyme particles to precipitate. The particles were collected on an inclined glass slide located at the bottom of the crystallizes, forming an angle of approximately 10° to the direction of the protein solution's flow. Parti-cles were also collected on a filter (Mott Metallurgi-cal; 316 Stainless Steel, 1.6 cm diameter, 0.5 /gym pore size). The carbon dioxide outlet was located approxi-mately 60 mm below the filter.
The duration of the experiment was 296 minutes for carbon dioxide input, arid 237 minutes for liquid input, followed by 17 minutes of liquid solution flow without dissolved enzyme.
Figures 6 illustrates particles collected on the filter which were needlelike having a diameter of less than 1 ~Cm and being less than 3 Vim. Figure 7 illus-trates particles collected which were globular, having an equivalent diameter of less than about 1 ~.m.
6~lhile exemplary embodiments of this invention have been described, the true scope of the invention is determined fram the following claims.
Claims (9)
1. A method of forming dry protein particles having an equivalent diameter of less than 5 µm, which comprises bringing droplets of a liquid solution of a protein in a solvent for said protein, the droplets having diameters of from 10 µm to 500 µm, into contact at a controlled rate with an excess of a supercritical antisolvent having low solvent power for said protein but operable to dissolve in and expand said liquid solution and precipitate the dry protein particles.
2. The method according to claim 1, wherein the supercritical antisolvent is carbon dioxide, ethane, ethylene, sulfur hexafluoride, nitrous oxide, chlorotrifluoromethane, monofluoromethane, or a mixture thereof.
3. The method according to claim 2, wherein the supercritical antisolvent is carbon dioxide.
4. The method according to claim 1, wherein the solvent of the protein is ethanol, dimethylsulfoxide, tetrahydrofuran, acetic acid, formamide, NN-dimethy-formamide, ethylene glycol, liquid polyethylene glycol, or N,N-dimethylaniline.
5. The method according to claim 4, wherein the solvent for the protein is aqueous ethanol.
6. The method according to claim 1, wherein the solution of the protein is sprayed into a continuum of said supercritical antisolvent.
7. The method according to claim 1, wherein the protein has low water solubility.
8. The method according to claim 1, wherein the protein is insulin, catalase, adrenocorticotrophin hormone, or peroxidase.
9. A method forming dry protein particles having an equivalent diameter of less than 5 µm, which comprises bringing droplets of a liquid solution of a protein in a solvent for said protein, the droplets having diameters of from 10 µm to 500 µm, into contact at a controlled rate without change in pressure with an excess of a supercritical antisolvent having low solvent power for said protein but operable to dissolve in and expand said liquid solution and precipitate the dry protein particles.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US07/792,292 US6063910A (en) | 1991-11-14 | 1991-11-14 | Preparation of protein microparticles by supercritical fluid precipitation |
| US07/792,292 | 1991-11-14 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2083027A1 CA2083027A1 (en) | 1993-05-15 |
| CA2083027C true CA2083027C (en) | 2004-01-20 |
Family
ID=25156397
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002083027A Expired - Fee Related CA2083027C (en) | 1991-11-14 | 1992-11-16 | Formation of protein microparticles by antisolvent precipitation |
Country Status (9)
| Country | Link |
|---|---|
| US (1) | US6063910A (en) |
| EP (1) | EP0542314B9 (en) |
| JP (1) | JP3342899B2 (en) |
| AT (1) | ATE167870T1 (en) |
| AU (1) | AU668367B2 (en) |
| CA (1) | CA2083027C (en) |
| DE (1) | DE69226064T3 (en) |
| DK (1) | DK0542314T4 (en) |
| ES (1) | ES2118778T5 (en) |
Families Citing this family (142)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| SI9400079B (en) * | 1994-02-15 | 2003-02-28 | Dr. Weidner Eckhard, Dipl. Ing. | Method and device for extraction and fractionation of small particles from solutions saturated with gas |
| GB9413202D0 (en) * | 1994-06-30 | 1994-08-24 | Univ Bradford | Method and apparatus for the formation of particles |
| US6428771B1 (en) * | 1995-05-15 | 2002-08-06 | Pharmaceutical Discovery Corporation | Method for drug delivery to the pulmonary system |
| EP0855906B1 (en) * | 1995-10-17 | 2008-02-20 | Jagotec AG | Insoluble drug delivery |
| US5874029A (en) * | 1996-10-09 | 1999-02-23 | The University Of Kansas | Methods for particle micronization and nanonization by recrystallization from organic solutions sprayed into a compressed antisolvent |
| US5833891A (en) * | 1996-10-09 | 1998-11-10 | The University Of Kansas | Methods for a particle precipitation and coating using near-critical and supercritical antisolvents |
| FR2753639B1 (en) * | 1996-09-25 | 1998-12-11 | PROCESS FOR THE PREPARATION OF MICROCAPSULES OF ACTIVE MATERIALS COATED WITH A POLYMER AND NOVEL MICROCAPSULES OBTAINED IN PARTICULAR BY THE PROCESS | |
| US5766637A (en) * | 1996-10-08 | 1998-06-16 | University Of Delaware | Microencapsulation process using supercritical fluids |
| DE19711393C1 (en) * | 1997-03-19 | 1998-08-13 | Fraunhofer Ges Forschung | Process for microencapsulating particles |
| US6541606B2 (en) * | 1997-12-31 | 2003-04-01 | Altus Biologics Inc. | Stabilized protein crystals formulations containing them and methods of making them |
| GB9806462D0 (en) * | 1998-03-26 | 1998-05-27 | Glaxo Group Ltd | Improved compositions for inhalation |
| GB9810559D0 (en) * | 1998-05-15 | 1998-07-15 | Bradford Particle Design Ltd | Method and apparatus for particle formation |
| US6451349B1 (en) | 1998-08-19 | 2002-09-17 | Quadrant Healthcare (Uk) Limited | Spray-drying process for the preparation of microparticles |
| GB9814172D0 (en) * | 1998-06-30 | 1998-08-26 | Andaris Ltd | Formulation for inhalation |
| US6051694A (en) * | 1998-09-17 | 2000-04-18 | Castor; Trevor Percival | Method for size reduction of proteins |
| SE9804001D0 (en) † | 1998-11-23 | 1998-11-23 | Astra Ab | New process |
| ATE555773T1 (en) | 1999-06-09 | 2012-05-15 | Sievers Robert E | SUPERCRITICAL FLUID-ASSISTED NEBULIZATION AND BUBBLES DRYING |
| EP2280020B1 (en) * | 1999-06-29 | 2016-02-17 | MannKind Corporation | Pharmaceutical formulations comprising a peptide complexed with a diketopiperazine |
| US9006175B2 (en) * | 1999-06-29 | 2015-04-14 | Mannkind Corporation | Potentiation of glucose elimination |
| GB9915975D0 (en) | 1999-07-07 | 1999-09-08 | Bradford Particle Design Ltd | Method for the formation of particles |
| US6680284B1 (en) | 1999-07-13 | 2004-01-20 | Degussa Ag | Method for producing powdery particle-reduced formulations with the aid of compressed gases |
| FR2803538B1 (en) * | 1999-12-15 | 2002-06-07 | Separex Sa | METHOD AND DEVICE FOR CAPTURING FINE PARTICLES BY PERCOLATION IN A BED OF GRANULES |
| FR2802445B1 (en) * | 1999-12-15 | 2002-02-15 | Separex Sa | METHOD AND DEVICE FOR CAPTURING FINE PARTICLES BY TRAPPING WITHIN A SOLID MIXTURE OF THE CARBON SNOW TYPE |
| AU2729101A (en) | 1999-12-21 | 2001-07-03 | Rxkinetix, Inc. | Particulate drug-containing products and method of manufacture |
| US6761909B1 (en) * | 1999-12-21 | 2004-07-13 | Rxkinetix, Inc. | Particulate insulin-containing products and method of manufacture |
| CH694686A5 (en) * | 2000-03-04 | 2005-06-15 | Eco2 Sa | Product micronization of pharmaceutical substances. |
| US6620351B2 (en) | 2000-05-24 | 2003-09-16 | Auburn University | Method of forming nanoparticles and microparticles of controllable size using supercritical fluids with enhanced mass transfer |
| US6821413B1 (en) * | 2000-08-31 | 2004-11-23 | Fluidphase Technologies, Inc. | Method and apparatus for continuous separation and reaction using supercritical fluid |
| US6562952B1 (en) * | 2000-10-31 | 2003-05-13 | The University Of Kansas | Precipitation of proteins from organic solutions |
| GB0027357D0 (en) | 2000-11-09 | 2000-12-27 | Bradford Particle Design Plc | Particle formation methods and their products |
| AUPR197000A0 (en) * | 2000-12-08 | 2001-01-11 | Unisearch Limited | Synthesis of small particles |
| US20050048126A1 (en) * | 2000-12-22 | 2005-03-03 | Barrett Rabinow | Formulation to render an antimicrobial drug potent against organisms normally considered to be resistant to the drug |
| US6977085B2 (en) | 2000-12-22 | 2005-12-20 | Baxter International Inc. | Method for preparing submicron suspensions with polymorph control |
| US6951656B2 (en) * | 2000-12-22 | 2005-10-04 | Baxter International Inc. | Microprecipitation method for preparing submicron suspensions |
| US8067032B2 (en) | 2000-12-22 | 2011-11-29 | Baxter International Inc. | Method for preparing submicron particles of antineoplastic agents |
| US9700866B2 (en) * | 2000-12-22 | 2017-07-11 | Baxter International Inc. | Surfactant systems for delivery of organic compounds |
| US7193084B2 (en) * | 2000-12-22 | 2007-03-20 | Baxter International Inc. | Polymorphic form of itraconazole |
| US20040256749A1 (en) * | 2000-12-22 | 2004-12-23 | Mahesh Chaubal | Process for production of essentially solvent-free small particles |
| US6884436B2 (en) | 2000-12-22 | 2005-04-26 | Baxter International Inc. | Method for preparing submicron particle suspensions |
| US20030096013A1 (en) * | 2000-12-22 | 2003-05-22 | Jane Werling | Preparation of submicron sized particles with polymorph control |
| US7037528B2 (en) | 2000-12-22 | 2006-05-02 | Baxter International Inc. | Microprecipitation method for preparing submicron suspensions |
| US20040052854A1 (en) * | 2000-12-26 | 2004-03-18 | Tomohiro Yoshinari | Porous substances and methods for producing the same |
| EP2325205A3 (en) * | 2000-12-28 | 2011-10-12 | Altus Pharmaceuticals Inc. | Crystals of whole antibodies and fragments thereof and methods for making and using them |
| US7708915B2 (en) * | 2004-05-06 | 2010-05-04 | Castor Trevor P | Polymer microspheres/nanospheres and encapsulating therapeutic proteins therein |
| PT1363726E (en) * | 2001-02-26 | 2004-11-30 | Dompe Spa | APPARATUS AND METHOD FOR THE FORMATION OF MICROMETRIC AND SUBMICROMETRIC PARTICLES |
| US6610223B2 (en) | 2001-03-30 | 2003-08-26 | Picoliter Inc. | Focused acoustic energy in the generation of solid particles |
| US6869551B2 (en) | 2001-03-30 | 2005-03-22 | Picoliter Inc. | Precipitation of solid particles from droplets formed using focused acoustic energy |
| US6596206B2 (en) | 2001-03-30 | 2003-07-22 | Picoliter Inc. | Generation of pharmaceutical agent particles using focused acoustic energy |
| WO2003000014A2 (en) | 2001-06-21 | 2003-01-03 | Altus Biologics, Inc. | Spherical protein particles and methods of making and using them |
| GB0208742D0 (en) | 2002-04-17 | 2002-05-29 | Bradford Particle Design Ltd | Particulate materials |
| GB0117696D0 (en) * | 2001-07-20 | 2001-09-12 | Bradford Particle Design Plc | Particle information |
| US20080026068A1 (en) * | 2001-08-16 | 2008-01-31 | Baxter Healthcare S.A. | Pulmonary delivery of spherical insulin microparticles |
| US20060003012A9 (en) * | 2001-09-26 | 2006-01-05 | Sean Brynjelsen | Preparation of submicron solid particle suspensions by sonication of multiphase systems |
| IL160570A0 (en) * | 2001-09-26 | 2004-07-25 | Baxter Int | Preparation of submicron sized nanoparticles via dispersion and solvent or liquid phase removal |
| CN1273113C (en) * | 2001-10-10 | 2006-09-06 | 贝林格尔·英格海姆药物公司 | Powder processing with pressurized gaseous fluids |
| US7112340B2 (en) * | 2001-10-19 | 2006-09-26 | Baxter International Inc. | Compositions of and method for preparing stable particles in a frozen aqueous matrix |
| US7250152B2 (en) * | 2001-10-22 | 2007-07-31 | Dompé Pha.R.Ma S.P.A. | Supercritical fluids processing: preparation of protein microparticles and their stablilisation |
| EP1461020A4 (en) * | 2001-12-07 | 2008-10-29 | Map Pharmaceuticals Inc | Synthesis of small particles |
| EP1494732B1 (en) | 2002-03-20 | 2008-01-30 | MannKind Corporation | Inhalation apparatus |
| AU2003221888B2 (en) * | 2002-04-11 | 2008-11-06 | Medimmune, Llc | Preservation of bioactive materials by spray drying |
| US7582284B2 (en) * | 2002-04-17 | 2009-09-01 | Nektar Therapeutics | Particulate materials |
| GB0216562D0 (en) | 2002-04-25 | 2002-08-28 | Bradford Particle Design Ltd | Particulate materials |
| US9339459B2 (en) | 2003-04-24 | 2016-05-17 | Nektar Therapeutics | Particulate materials |
| AU2004237131B2 (en) * | 2003-05-08 | 2009-09-10 | Nektar Therapeutics | Particulate materials |
| US7767118B2 (en) | 2003-06-03 | 2010-08-03 | Ferro Corporation | Nanoparticles from supercritical fluid antisolvent process using particle growth and agglomeration retardants |
| US8075919B2 (en) * | 2003-07-18 | 2011-12-13 | Baxter International Inc. | Methods for fabrication, uses and compositions of small spherical particles prepared by controlled phase separation |
| US20070092452A1 (en) * | 2003-07-18 | 2007-04-26 | Julia Rashba-Step | Methods for fabrication, uses, compositions of inhalable spherical particles |
| US20050142205A1 (en) * | 2003-07-18 | 2005-06-30 | Julia Rashba-Step | Methods for encapsulating small spherical particles prepared by controlled phase separation |
| EP1646354A4 (en) * | 2003-07-22 | 2010-03-17 | Baxter Int | Small spherical particles of low molecular weight organic molecules and methods of preparation and use thereof |
| AU2004272077A1 (en) * | 2003-09-10 | 2005-03-24 | Map Pharmaceuticals, Inc. | Aerosol formulations for delivery of dihydroergotamine to the systemic circulation via pulmonary inhalation |
| WO2005042219A1 (en) * | 2003-10-24 | 2005-05-12 | Ferro Corporation | Method of forming particles |
| US7338171B2 (en) * | 2003-10-27 | 2008-03-04 | Jen-Chuen Hsieh | Method and apparatus for visual drive control |
| ES2584867T3 (en) * | 2004-01-12 | 2016-09-29 | Mannkind Corporation | A method that reduces serum proinsulin levels in type 2 diabetics |
| WO2005077337A2 (en) * | 2004-02-05 | 2005-08-25 | Baxter International Inc. | Dispersions prepared by use of self-stabilizing agents |
| US20080085298A1 (en) * | 2004-03-12 | 2008-04-10 | Biodel, Inc. | Rapid Mucosal Gel or Film Insulin Compositions |
| EP2319500B1 (en) * | 2004-03-12 | 2012-10-24 | Biodel, Inc. | Rapid acting drug delivery compositions |
| US20080090753A1 (en) * | 2004-03-12 | 2008-04-17 | Biodel, Inc. | Rapid Acting Injectable Insulin Compositions |
| US20080096800A1 (en) * | 2004-03-12 | 2008-04-24 | Biodel, Inc. | Rapid mucosal gel or film insulin compositions |
| EP2335689A1 (en) | 2004-05-12 | 2011-06-22 | Baxter International Inc. | Method of manufacturing nucleic acid micropheres |
| WO2005112885A2 (en) * | 2004-05-12 | 2005-12-01 | Baxter International Inc. | Oligonucleotide-containing microspheres, their use for the manufacture of a medicament for treating diabetes type 1 |
| US8728525B2 (en) * | 2004-05-12 | 2014-05-20 | Baxter International Inc. | Protein microspheres retaining pharmacokinetic and pharmacodynamic properties |
| EP1753404A1 (en) | 2004-05-12 | 2007-02-21 | Baxter International Inc. | Microspheres comprising protein and showing injectability at high concentrations of said agent |
| DK1786784T3 (en) | 2004-08-20 | 2011-02-14 | Mannkind Corp | Catalysis of diketopiperazine synthesis |
| PL2322180T3 (en) | 2004-08-23 | 2015-10-30 | Mannkind Corp | Diketopiperazine salts for drug delivery |
| CN101188996B (en) * | 2005-04-27 | 2013-03-27 | 巴克斯特国际公司 | Surface-modified microparticles and methods of forming and using the same |
| EP1731219B1 (en) * | 2005-06-11 | 2007-05-23 | TuTech Innovation GmbH | Process for encapsulating organic substances in particle form by spraying an inert, supercritical carrier gas together with the coating material for the organic substance into a high pressure fluidised bed contained in an autoclave |
| US20060280787A1 (en) * | 2005-06-14 | 2006-12-14 | Baxter International Inc. | Pharmaceutical formulation of the tubulin inhibitor indibulin for oral administration with improved pharmacokinetic properties, and process for the manufacture thereof |
| JP2008543842A (en) * | 2005-06-14 | 2008-12-04 | バクスター インターナショナル インコーポレイテッド | Pharmaceutical formulations for minimizing drug-drug interactions |
| CN104324366B (en) | 2005-09-14 | 2016-10-05 | 曼金德公司 | Method for preparation of drug based on improving the active agent affinity to crystalline microparticle surfaces |
| US7713929B2 (en) | 2006-04-12 | 2010-05-11 | Biodel Inc. | Rapid acting and long acting insulin combination formulations |
| US8084420B2 (en) * | 2005-09-29 | 2011-12-27 | Biodel Inc. | Rapid acting and long acting insulin combination formulations |
| WO2007041481A1 (en) * | 2005-09-29 | 2007-04-12 | Biodel, Inc. | Rapid acting and prolonged acting insulin preparations |
| EP1954245A2 (en) * | 2005-11-15 | 2008-08-13 | Baxter International Inc. | Compositions of lipoxygenase inhibitors |
| JP5457680B2 (en) * | 2006-01-24 | 2014-04-02 | アンサン バイオファーマ，インコーポレイテッド | Polymer microsphere preparation technology |
| MX360812B (en) | 2006-02-22 | 2018-11-16 | Mannkind Corp | A method for improving the pharmaceutic properties of microparticles comprising diketopiperazine and an active agent. |
| AU2007238114B2 (en) * | 2006-04-12 | 2010-10-14 | Biodel, Inc. | Rapid acting and long acting insulin combination formulations |
| DE102006020148A1 (en) * | 2006-05-02 | 2007-11-08 | Universität Bremen | Genetically engineered perlucin and its use for the production of biomaterials |
| US20070281031A1 (en) * | 2006-06-01 | 2007-12-06 | Guohan Yang | Microparticles and methods for production thereof |
| CA2657517C (en) | 2006-08-04 | 2016-11-01 | Baxter International Inc. | Microsphere-based composition for preventing and/or reversing new-onset autoimmune diabetes |
| JP2010505882A (en) * | 2006-10-06 | 2010-02-25 | バクスター・インターナショナル・インコーポレイテッド | Microencapsules containing surface modified microparticles and methods for their formation and use |
| US20090152176A1 (en) * | 2006-12-23 | 2009-06-18 | Baxter International Inc. | Magnetic separation of fine particles from compositions |
| HUE026884T2 (en) | 2007-02-11 | 2016-08-29 | Map Pharmaceuticals Inc | Method of therapeutic administration of dhe to enable rapid relief of migraine while minimizing side effect profile |
| WO2008124522A2 (en) * | 2007-04-04 | 2008-10-16 | Biodel, Inc. | Amylin formulations |
| US8426467B2 (en) * | 2007-05-22 | 2013-04-23 | Baxter International Inc. | Colored esmolol concentrate |
| US20080293814A1 (en) * | 2007-05-22 | 2008-11-27 | Deepak Tiwari | Concentrate esmolol |
| US8722736B2 (en) * | 2007-05-22 | 2014-05-13 | Baxter International Inc. | Multi-dose concentrate esmolol with benzyl alcohol |
| US8785396B2 (en) | 2007-10-24 | 2014-07-22 | Mannkind Corporation | Method and composition for treating migraines |
| US20090175840A1 (en) * | 2008-01-04 | 2009-07-09 | Biodel, Inc. | Insulin formulations for insulin release as a function of tissue glucose levels |
| US8485180B2 (en) | 2008-06-13 | 2013-07-16 | Mannkind Corporation | Dry powder drug delivery system |
| DE202009018480U1 (en) | 2008-06-13 | 2012-01-26 | Mannkind Corp. | Dry powder inhaler and drug delivery system |
| MX2010014240A (en) | 2008-06-20 | 2011-03-25 | Mankind Corp | An interactive apparatus and method for real-time profiling of inhalation efforts. |
| TWI532497B (en) | 2008-08-11 | 2016-05-11 | 曼凱公司 | Use of ultrarapid acting insulin |
| US8323615B2 (en) * | 2008-08-20 | 2012-12-04 | Baxter International Inc. | Methods of processing multi-phasic dispersions |
| US8367427B2 (en) * | 2008-08-20 | 2013-02-05 | Baxter International Inc. | Methods of processing compositions containing microparticles |
| US8323685B2 (en) | 2008-08-20 | 2012-12-04 | Baxter International Inc. | Methods of processing compositions containing microparticles |
| US20100047292A1 (en) * | 2008-08-20 | 2010-02-25 | Baxter International Inc. | Methods of processing microparticles and compositions produced thereby |
| WO2010065387A1 (en) * | 2008-12-01 | 2010-06-10 | Innovative Technologies, Inc. | Improved preparation of organic compounds for enhanced reactivity |
| US8314106B2 (en) | 2008-12-29 | 2012-11-20 | Mannkind Corporation | Substituted diketopiperazine analogs for use as drug delivery agents |
| US9060927B2 (en) * | 2009-03-03 | 2015-06-23 | Biodel Inc. | Insulin formulations for rapid uptake |
| US8538707B2 (en) | 2009-03-11 | 2013-09-17 | Mannkind Corporation | Apparatus, system and method for measuring resistance of an inhaler |
| KR101875969B1 (en) | 2009-06-12 | 2018-07-06 | 맨카인드 코포레이션 | Diketopiperazine microparticles with defined specific surface areas |
| CA2778698A1 (en) | 2009-11-03 | 2011-05-12 | Mannkind Corporation | An apparatus and method for simulating inhalation efforts |
| RU2571331C1 (en) | 2010-06-21 | 2015-12-20 | Маннкайнд Корпорейшн | Systems and methods for dry powder drug delivery |
| US9138705B2 (en) | 2010-11-22 | 2015-09-22 | Hong Kong Polytechnic University | Method for precipitating a solute from a solution |
| US9808030B2 (en) | 2011-02-11 | 2017-11-07 | Grain Processing Corporation | Salt composition |
| BR122020008875B8 (en) | 2011-04-01 | 2022-12-06 | Mannkind Corp | BLISTER PACKAGING AND METHOD OF MANUFACTURING A BLISTER PACKAGE |
| WO2012174472A1 (en) | 2011-06-17 | 2012-12-20 | Mannkind Corporation | High capacity diketopiperazine microparticles |
| CA2852536A1 (en) | 2011-10-24 | 2013-05-02 | Mannkind Corporation | Methods and compositions for treating pain |
| CN105688188A (en) | 2011-11-18 | 2016-06-22 | 瑞泽恩制药公司 | Polymer protein microparticles |
| US20140099380A1 (en) | 2012-06-28 | 2014-04-10 | Ansun Biopharma, Inc. | Microparticle Formulations for Delivery to the Lower and Central Respiratory Tract and Methods of Manufacture |
| KR102264177B1 (en) | 2012-07-12 | 2021-06-11 | 맨카인드 코포레이션 | Dry powder drug delivery systems and methods |
| US10159644B2 (en) | 2012-10-26 | 2018-12-25 | Mannkind Corporation | Inhalable vaccine compositions and methods |
| BR112015023168B1 (en) | 2013-03-15 | 2021-08-10 | Mannkind Corporation | COMPOSITION OF 3,6-BIS(N-FUMARYL-4-AMINOBUTYL)-2,5-CRYSTALLINE DICETOPIPERAZINE, METHOD OF PRODUCTION OF 3,6-BIS(N-FUMARYL-4-AMINOBUTYL)-2,5-DICETOPIPERAZINE PARTICLES AND USE OF A CRYSTALLINE DICETOPIPERAZINE COMPOSITION |
| CN114848614A (en) | 2013-07-18 | 2022-08-05 | 曼金德公司 | Heat stable dry powder pharmaceutical compositions and methods |
| EP3030294B1 (en) | 2013-08-05 | 2020-10-07 | MannKind Corporation | Insufflation apparatus |
| US10307464B2 (en) | 2014-03-28 | 2019-06-04 | Mannkind Corporation | Use of ultrarapid acting insulin |
| GB201409451D0 (en) | 2014-05-28 | 2014-07-09 | Ipabc Ltd | Antimicrobial preparations, methods for preparing the same and uses thereof to combat microorganisms |
| US10561806B2 (en) | 2014-10-02 | 2020-02-18 | Mannkind Corporation | Mouthpiece cover for an inhaler |
| CN108310797A (en) * | 2018-04-09 | 2018-07-24 | 南通市华安超临界萃取有限公司 | A kind of supercritical fluid recrystallization device |
| KR20220104797A (en) | 2019-11-25 | 2022-07-26 | 리제너론 파마슈티칼스 인코포레이티드 | Sustained release formulation using non-aqueous emulsion |
| TW202237070A (en) | 2020-11-25 | 2022-10-01 | 美商再生元醫藥公司 | Sustained release formulations using non-aqueous membrane emulsification |
| WO2022133135A1 (en) | 2020-12-17 | 2022-06-23 | Regeneron Pharmaceuticals, Inc. | Fabrication of protein-encapsulating microgels |
Family Cites Families (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3773919A (en) * | 1969-10-23 | 1973-11-20 | Du Pont | Polylactide-drug mixtures |
| DE2837115A1 (en) * | 1978-08-25 | 1980-03-06 | Bayer Ag | NEW METHOD FOR THE PRODUCTION OF STERILE SOLIDS |
| KR890002631B1 (en) * | 1984-10-04 | 1989-07-21 | 몬산토 캄파니 | Composition of prolonged release of biologically active somatotropin |
| US5169968A (en) * | 1985-09-10 | 1992-12-08 | Vitamins, Inc. | Mass separation of liquid or soluble components from solid materials utilizing supercritical fluids |
| US4985270A (en) * | 1988-01-26 | 1991-01-15 | The Nutrasweet Company | Cream substitute ingredient and food products |
| DE3744329A1 (en) * | 1987-12-28 | 1989-07-06 | Schwarz Pharma Gmbh | METHOD FOR THE PRODUCTION OF AT LEAST ONE ACTIVE AGENT AND A TRAITER COMPRISING PREPARATION |
| GB8802223D0 (en) * | 1988-02-02 | 1988-03-02 | St Ivel Ltd | Low fat spread |
| JP2843857B2 (en) * | 1988-10-05 | 1999-01-06 | ファルマシア・アンド・アップジョン・カンパニー | Finely divided solid crystalline powder via precipitation in antisolvent |
| US5639441A (en) * | 1992-03-06 | 1997-06-17 | Board Of Regents Of University Of Colorado | Methods for fine particle formation |
| US5301664A (en) * | 1992-03-06 | 1994-04-12 | Sievers Robert E | Methods and apparatus for drug delivery using supercritical solutions |
| GB9313642D0 (en) * | 1993-07-01 | 1993-08-18 | Glaxo Group Ltd | Method and apparatus for the formation of particles |
| US5904935A (en) * | 1995-06-07 | 1999-05-18 | Alza Corporation | Peptide/protein suspending formulations |
| US5814678A (en) * | 1996-02-02 | 1998-09-29 | University Technology Corp. | Chemical reactions in water-in-carbon dioxide microemulsions and control thereof |
-
1991
- 1991-11-14 US US07/792,292 patent/US6063910A/en not_active Expired - Fee Related
-
1992
- 1992-11-11 AU AU28310/92A patent/AU668367B2/en not_active Ceased
- 1992-11-12 JP JP32738992A patent/JP3342899B2/en not_active Expired - Fee Related
- 1992-11-13 AT AT92119498T patent/ATE167870T1/en not_active IP Right Cessation
- 1992-11-13 ES ES92119498T patent/ES2118778T5/en not_active Expired - Lifetime
- 1992-11-13 EP EP92119498A patent/EP0542314B9/en not_active Expired - Lifetime
- 1992-11-13 DK DK92119498T patent/DK0542314T4/en active
- 1992-11-13 DE DE69226064T patent/DE69226064T3/en not_active Expired - Fee Related
- 1992-11-16 CA CA002083027A patent/CA2083027C/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| AU668367B2 (en) | 1996-05-02 |
| EP0542314B2 (en) | 2005-01-05 |
| ES2118778T3 (en) | 1998-10-01 |
| DE69226064D1 (en) | 1998-08-06 |
| DE69226064T2 (en) | 1998-10-29 |
| ATE167870T1 (en) | 1998-07-15 |
| EP0542314A1 (en) | 1993-05-19 |
| JPH05293361A (en) | 1993-11-09 |
| CA2083027A1 (en) | 1993-05-15 |
| EP0542314B9 (en) | 2005-05-04 |
| EP0542314B1 (en) | 1998-07-01 |
| DK0542314T4 (en) | 2005-05-17 |
| DE69226064T3 (en) | 2005-07-07 |
| ES2118778T5 (en) | 2005-07-16 |
| AU2831092A (en) | 1993-05-20 |
| JP3342899B2 (en) | 2002-11-11 |
| DK0542314T3 (en) | 1999-04-12 |
| US6063910A (en) | 2000-05-16 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CA2083027C (en) | Formation of protein microparticles by antisolvent precipitation | |
| Debenedetti et al. | Application of supercritical fluids for the production of sustained delivery devices | |
| Benedetti et al. | Production of micronic particles of biocompatible polymer using supercritical carbon dioxide | |
| Tom et al. | Formation of bioerodible polymeric microspheres and microparticles by rapid expansion of supercritical solutions | |
| US6541035B1 (en) | Nanosheres comprising a biocompatible polysaccharide | |
| Kompella et al. | Preparation of drug delivery systems using supercritical fluid technology | |
| US5851453A (en) | Method and apparatus for the formation of particles | |
| Thiering et al. | The influence of operating conditions on the dense gas precipitation of model proteins | |
| JP4374249B2 (en) | Processing method with supercritical fluid: Preparation of protein microparticles and their stabilization | |
| AU2005209541B2 (en) | A method for producing fine particles using method of rapid expansion into poor solvent from supercritical fluid | |
| AU3720999A (en) | Methods and apparatus for particle formation | |
| KR20090077066A (en) | Particle formation | |
| Knutson et al. | Preparation of microparticulates using supercritical fluids | |
| Hong et al. | Precipitation of microparticulate organic pigment powders by a supercritical antisolvent process | |
| US7449136B2 (en) | Method and apparatus for producing composite particles using supercritical fluid as plasticizing and extracting agent | |
| US8142814B2 (en) | Method and apparatus for supercritical fluid assisted particle production | |
| EP1499296B1 (en) | Formulation of fine particles using liquefied or dense gases | |
| WO2004108265A2 (en) | Nanoparticles from supercritical fluid antisolvent process using particle growth and agglomeration retardants | |
| CA2466525C (en) | Synthesis of small particles | |
| US20030013634A1 (en) | Synthesis of small particles | |
| US20070267768A1 (en) | Production of Porous Materials by Supercritical Fluid Processing | |
| Bustami et al. | Recent applications of supercritical fluid technology to pharmaceutical powder systems | |
| AU2002221320B2 (en) | Synthesis of small particles | |
| Benita | Recent advances and industrial applications of microencapsulation | |
| Tech | REPORT DOCUMENTATION PAGE* v |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| EEER | Examination request | ||
| MKLA | Lapsed |