USOO6596206B2 (12) United States Patent (10) Patent No.: US 6,596,206 B2 Lee (45) Date of Patent: *Jul. 22, 2003

(54) GENERATION OF PHARMACEUTICAL FOREIGN PATENT DOCUMENTS

AENESSING FOCUSED WO WO OO/37169O542314 6/20007/1998 (75) Inventor: partson-Hui Lee, Mountain View, W W s: 2: OTHER PUBLICATIONS (73) Assignee: Picoliter Inc., Sunnyvale, CA (US) Debenedetti et al. (1993), “Application of Supercritical Fluids for the Production of Sustained Delivery Devices.” (*) Notice: Subject to any disclaimer, the term of this Journal of Controlled Release 24:27–44. patent is extended or adjusted under 35 Tom et al. (1991), “Formation of Bioerodible Polymeric U.S.C. 154(b) by 14 days. Microspheres and Microparticles by Rapid Expansion of Supercritical Solutions,” Biotechnol. Prog. 7(5):403-411. This patent is Subject to a terminal dis- Chattopadhyay et al. (2001), “Production of Griseofulvin claimer. Nanoparticles Using Supercritical CO2 Antisolvent with Enhanced Mass Transfer.” International Journal of Phar (21) Appl. No.: 09/823,899 maceutics 228:19-31. (22) Filed: Mar 30, 2001 Jung et al. (2001), “Particle Design Using Supercritical e a Vs Fluids: Literature and Patent Survey,” Journal of Supercriti (65) Prior Publication Data cal Fluids 20:179-219. Primary Examiner Mary Lynn Theisen US 2002/0142049 A1 Oct. 3, 2002 (74) Attorney, Agent, or Firm-Dianne E. Reed; Reed & (51) Int. Cl." ...... B29C 9/00 Eberle LLP (52) U.S. Cl...... 264/9; 264/5; 2647. (57) ABSTRACT (58) Field of Search ...... 264/9, 7, 5; 425/6, A method and device for generating pharmaceutical agent 425/10 particles using focused acoustic energy are provided. A Solution of the pharmaceutical agent is provided in a Solvent, (56) References Cited which may be an aqueous fluid, a nonaqueous fluid, or a Supercritical fluid. Focused acoustic energy is used to eject U.S. PATENT DOCUMENTS a droplet of the solution, which is then directed into or 4,308,547 A 12/1981 Lovelady et al...... 346/140 R through an antisolvent that upon admixture with the Solution 5,041,849 A 8/1991 Ouate et al...... 346/140 R droplet causes the pharmaceutical agent in the droplet to 5,874,029 A 2/1999 Subramaniam et al...... 264/12 precipitate. In a preferred embodiment, the Solvent is an 2002/OOOO681 A1 1/2002 Gupta et al...... 264/9 aqueous or organic liquid, and the antisolvent is a Super 2002/0073989 A1 6/2002 Hadimioglu ...... 128/200.14 critical fluid. 2002/0073990 A1 6/2002 Noolandi et al...... 128/200.16 2002/0077369 A1 6/2002 Noolandi et al...... 514/958 109 Claims, 1 Drawing Sheet

U.S. Patent Jul. 22, 2003 US 6,596,206 B2

US 6,596.206 B2 1 2 GENERATION OF PHARMACEUTICAL Supercritical fluid technology has Solved Some of these AGENT PARTICLES USING FOCUSED problems. One method for using this relatively new tech ACOUSTIC ENERGY nology is called the rapid expansion of Supercritical Solu tions or “RESS" method. See Tom et al. (1991) Biotechnol. s Prog. 7(5):403-411. In the RESS method, the solute of TECHNICAL FIELD interest, e.g., a pharmaceutical agent, is first Solubilized in a This invention relates generally to the generation of Supercritical fluid, i.e., a fluid at a temperature and preSSure pharmaceutical agent particles, and more particularly relates greater than its critical temperature (T) and critical pressure to the use of focused acoustic energy in generating Solid (P) Generally, the Supercritical fluid is carbon dioxide, particles comprised of a pharmaceutical agent. although other fluids are available. The solution is then BACKGROUND rapidly passed through a nozzle that is connected to a relatively low-pressure medium. The Sudden depressuriza Rapid and efficient production of particles, particularly tion of the Solution as it passes into the relatively low Small and/or Substantially uniform particles, is needed in a preSSure medium causes the Supercritical fluid to expand, variety of industries. Among other advantages, Small, Sub i.e., the density of the Supercritical fluid decreases, reducing Stantially uniform particles possess favorable flow charac 15 the ability of the Supercritical fluid to solubilize the thera teristics and exhibit little variation in interparticle behavior. peutic agent. As a direct consequence of the reduced In the pharmaceutical industry, for example, the particle size Solubility, a SuperSaturated Solution develops, which, in turn, of a therapeutic agent can affect the dissolution rate, bio causes the Solute agent to precipitate or crystallize out in availability and overall stability of the agent in a formula very Small particles. tion. Precise control of the particle Size of therapeutic agents A variation of this idea is to prepare a Solution of a is particularly important for Sustained release applications, therapeutic drug in a conventional Solvent, and then Spray where the rate of drug release is related to the Size of a the Solution through a nozzle into a Supercritical fluid that particle containing the drug. In addition, pulmonary delivery acts as an anti-Solvent. When the two fluids make contact, a of a therapeutic agent requires specifically sized particles, rapid Volume expansion occurs, reducing Solvent density generally on the order of about 1 um to about 7 lum. Particles 25 and Solvent capacity, in turn increasing SuperSaturation, that are too large may be deposited within the throat, while Solute nucleation and particle formulation. This method is particles that are too small will be exhaled. Thus, the ability commonly referred to as gas anti-Solvent recrystallization or to produce Small, uniform particles of a therapeutic agent is “GAS.” See, for example, Debenedetti et al. (1993) J. critically important in the development of particulate phar Control. Release 24:27-44 and PCT WOOO/37169 to Mer maceutical products. rifield. This proceSS has been applied to various proteins to Various approaches for attaining Small and uniform par produce particle sizes of about 5 um. See European Patent ticles have been used. Conventional comminution No. O 542 314. techniques, e.g., crushing, grinding and milling, rely on Although use of Supercritical fluid technology offers the mechanical forces to break apart relatively large particles capability of producing relatively Small particles of uniform into Smaller particles. Air-jet mills and other mills, available 35 size, it is not without drawbacks. One problem associated from, for example, DT Industries, Bristol, Pa., under the with these Supercritical methods is the reliance on nozzles tradename STOKES(R), are commonly used by the pharma and tubes for delivering the solutions. Nozzles are known to ceutical industry to decrease the particle Size of a bulk wear down over time, altering the geometry of the equip therapeutic agent into a range Suitable for pharmaceutical ment and affecting the size of the droplets formed. In applications. One drawback to Such mechanical comminu 40 addition, nozzles may become blocked during use, when, for tion techniques, however, is that Some therapeutic agents, example, particles agglomerate upon rapid expansion within particularly proteins and other therapeutic biomolecules, are the nozzle bore. In addition, nozzles and associated compo damaged during the process. Another drawback of mechani nents require cleaning and may contaminate Solutions when cal comminution is the wide distribution of particle sizes not properly maintained. produced by these techniques. Among other problems, large 45 Furthermore, the droplet sizes of the solutions (both variations in the size of particles limit the ability to produce Supercritical and conventional Solutions) produced by meth Sustained-release formulations and waste large amounts of ods relying on nozzles are relatively varied. As a result there therapeutic agents intended, for example, for inhalation. will be a large variance of the Surface tension between Although Sieving a comminuted therapeutic agent through droplets of different sizes. At the sizes required for Super an appropriate mesh Screen provides a more narrow particle 50 critical methods, the differences in Surface tension between Size distribution, large quantities of particles not having the droplets causes large variations in crystallization kinetics desired size are wasted and the potential for contamination and growth. These differences result in differently sized is increased, as the therapeutic agent must contact additional particles. Although U.S. Pat. No. 5,874,029 to Subramaniam Surfaces. et al. discusses methods for producing Small-sized droplets Other techniques for producing pharmaceutical particles 55 using nozzles, the methods still suffer from the inability to include conventional recrystallization methods. In Such effectively and consistently produce droplets of uniform methods, the therapeutic agent is initially dissolved in a SZC. Suitable Solvent. In one approach, the temperature of the Thus, there is a need in the art for an improved particle Solution is changed So that the Solubility of the Solute is formation technique wherein particle formation is highly decreased. In another approach, a Second Solvent, an 60 reproducible, controllable and predictable, and Substantially “antisolvent,” is added so that the solubility of the solute is uniform particle size can be achieved. An ideal method decreased. In both approaches, the Solute precipitates or would minimize or eliminate contact of the particle-forming crystallizes out of the solution due to reduced solubility in fluid(s) with Surfaces of process equipment or contaminants the altered Solution. These methods, however, often require adsorbed thereon. The present invention addresses the afore toxic Solvents, result in wet particles (that require further 65 mentioned need in the art by using focused acoustic energy processing, e.g., drying), and may produce particles having to eject particle-forming droplets from a pharmaceutical variable sizes. agent Solution. US 6,596.206 B2 3 4 SUMMARY OF THE INVENTION droplet therefrom; and, optionally, a means for positioning Accordingly, it is a primary object of the invention to the ejector in acoustic coupling relationship to the reservoir. address the aforementioned need in the art by providing a BRIEF DESCRIPTION OF THE DRAWINGS novel method and device for generating pharmaceutical FIGS. 1A and 1B, collectively referred to as FIG. 1, agent particles using focused acoustic ejection technology. Schematically illustrate in Simplified cross-sectional view a Additional objects, advantages and novel features of the focused acoustic energy device useful in conjunction with invention will be set forth in part in the description which preparing pharmaceutical agent particles according to the follows, and in part will become apparent to those skilled in method of the invention. the art upon examination of the following, or may be learned 1O by practice of the invention. DETAILED DESCRIPTION OF THE In one aspect, then, the invention provides a method and INVENTION device for generating Solid particles of pharmaceutical Before describing the present invention in detail, it is to agents using focused acoustic energy. A Solution of the be understood that unless otherwise indicated this invention pharmaceutical agent is provided in a Solvent, which may be 15 is not limited to Specific pharmaceutical agents, fluids, an aqueous fluid, a nonaqueous fluid, or a Supercritical fluid. acoustic ejection devices, or the like, as Such may vary. It is Focused acoustic energy is used to eject a droplet of the also to be understood that the terminology used herein is for Solution, which is then directed into or through an antisol the purpose of describing particular embodiments only, and vent that upon admixture with the Solution droplet causes the is not intended to be limiting. pharmaceutical agent in the droplet to precipitate. The Solid It must be noted that, as used in this specification and the particle that results is then collected. In a preferred appended claims, the singular forms “a,” “an” and “the” embodiment, the Solvent is an aqueous or organic liquid, and include plural referents unless the context clearly dictates the antisolvent is a Supercritical fluid. It will be appreciated otherwise. Thus, for example, the term “a solvent” is that the pharmaceutical agent must be leSS Soluble in the intended to mean a single Solvent or a mixture of a Solvent antisolvent than in the solvent, and substantially inert in both 25 with one or more coSolvents, “a pharmaceutical agent' the Solvent and antisolvent. refers to a single pharmaceutical agent as well as to a Generally, the solution and the antisolvent will both be mixture of different pharmaceutical agent, “an antisolvent' present in the reservoir, with the reservoir being covered or includes one antisolvent or a mixture of different otherwise enclosed So as to provide the “contained Space.” antisolvents, “a reservoir is intended to mean one or more The particles resulting from the ejected droplets are col reservoirs, and the like. lected on a Surface within the contained Space, typically on In describing and claiming the present invention, the a Surface within the reservoir enclosure. following terminology will be used in accordance with the With Supercritical antisolvents, expansion of the solution definitions set out below. droplet upon ejection into a lower pressure Supercritical The terms "acoustic coupling” and "acoustically coupled' medium causes rapid depressurization of the droplet, Super 35 used herein refer to a State wherein a first entity is placed in Saturation thereof, and precipitation of Virtually direct or indirect contact with another entity So as to allow contaminant-free particles, ideally in crystalline form. acoustic radiation to be transferred between the objects The method is advantageous in a number of respects. For without Substantial loSS of acoustic energy. When two enti example, the method: 40 ties are indirectly acoustically coupled, an "acoustic cou can be used to prepare very Small particles, on the order pling medium' is needed to provide an intermediary through of microns or even nanometers in diameter; which acoustic radiation may be transmitted. Thus, an ejector may be acoustically coupled to a fluid, e.g., by gives rise to particles of Substantially uniform size, i.e., immersing the ejector in the fluid or by interposing an having a narrow particle size distribution; acoustic coupling medium between the ejector and the fluid can be used to prepare different crystal Structures of a 45 to transfer acoustic radiation generated by the ejector Single molecular entity (i.e., by Selection of a proper through the acoustic coupling medium and into the fluid. Solvent and/or Solvent-coSolvent combination); The term “fluid” as used herein refers to matter that is is highly reproducible, controllable and predictable; nonsolid or at least partially gaseous and/or liquid. A fluid can be readily Scaled up, but is also quite effective with may contain a Solid that is minimally, partially or fully very Small quantities of both pharmaceutical agents and 50 Solvated, dispersed or Suspended. Examples of fluids fluids, making it ideal for manufacturing particles of include, without limitation, aqueous liquids (including water rare and/or expensive drugs, per se and buffered water), nonaqueous liquids Such as is a single-step process, in contrast to the many multi-step organic Solvents and lipidic liquids, Supercritical fluids, processes of the prior art, and 55 gases, and the like. is Suitable for use with a wide range of pharmaceutical The term “supercritical fluid” refers to a fluid at or above agents and excipients. both its critical pressure P and critical temperature T. The In another aspect, the invention provides a device for molar volume and Solubilizing capacity of a Supercritical carrying out the aforementioned method. The device com fluid can be Substantially altered by varying the temperature prises: a reservoir containing a Solution of the pharmaceu 60 and/or pressure of the fluid. Fluids that are slightly below tical agent in a Solvent; an antisolvent in a contained space their actual critical temperature and pressure can also exhibit in fluid communication with the Solution in the reservoir Such characteristics, and the term "Supercritical fluid” is Such that droplets ejected from the Solution are directed into intended to encompass Such fluids as well. the antisolvent; an acoustic ejector comprising an acoustic The term “solvent” refers to a fluid that is capable of at radiation generator for generating acoustic radiation and a 65 least partially dissolving a Solute of interest. focusing means for focusing the acoustic radiation at a focal The term “antisolvent” refers to a fluid that when mixed point within the Solution in the reservoir So as to eject a with a solvent in which a solute is dissolved, reduces the US 6,596.206 B2 S 6 capacity of the Solvent to dissolve the Solute. Thus, when an agents, including antineoplastic drugs, anticholinergics, antisolvent is admixed with a Solution of a Solute in a anticonvulsants, antidepressants, antidiabetic agents, antidi solvent, the solubility of the solute can be reduced to the arrheals, antihelminthics; antihistamines, antihyperlipi point at which it precipitates out of Solution. Gases, and demic agents, antihypertensive agents, anti-infective agents particularly compressed gases, can act as antisolvents, Such as antibiotics and antiviral agents, antiinflammatory although the preferred antisolvents herein are in a Super agents, antimigraine preparations, antinauseants, antiparkin critical fluid state. The antisolvent must be sufficiently Sonism drugs, antipruritics, antipsychotics, antipyretics, miscible with the Solvent that Solute precipitation does in antispasmodics, antitubercular agents, antiulcer agents, fact result. It will be appreciated that miscibility can be anxiolytics, appetite SuppreSSants, attention deficit disorder controlled by varying one or more parameters within the (ADD) and attention deficit hyperactivity disorder (ADHD) Solvent/antisolvent System, e.g., the Solvent and antisolvent drugs, cardiovascular preparations including calcium chan System may be maintained at a Sufficiently low temperature nel blockers, CNS agents; beta-blockers and antiarrhythmic so that the two fluids are not particularly miscible (for agents, central nervous System Stimulants, cough and cold Storage, for example), and the temperature may then be preparations, including decongestants, diuretics; genetic raised So that the two fluids are miscible and particle 15 materials; herbal remedies, hormonolytics, hypnotics; formation can occur. hypoglycemic agents, immunosuppressive agents, leukot The term “focusing means' as used herein refers to a riene inhibitors, mitotic inhibitors, muscle relaxants, nar device that causes acoustic waves to converge at a focal cotic antagonists, nicotine; nutritional agents, Such as point by an action analogous to that of an optical lens. A Vitamins, essential amino acids and fatty acids, ophthalmic focusing means may be as Simple as a Solid member having drugS Such as antiglaucoma agents, parasympatholytics; a curved Surface, or it may include complex Structures Such psychoStimulants, Sedatives, Steroids, Sympathomimetics, as those found in Fresnel lenses, which employ diffraction in tranquilizers, and vasodilators including general coronary, order to direct acoustic radiation. peripheral and cerebral. The terms “particle' and “solid particle' are used inter The pharmaceutical agent may also be a biomolecule, 25 e.g., a molecular moiety Selected from the group consisting changeably herein to refer to Solid particles or particles that of DNA, RNA, antisense oligonucleotides, peptidyl drugs, include Solid matter. Generally, the average Size of the i.e., peptides, polypeptides and proteins (including fluores particles prepared using the method of the invention is in the cent proteins), ribosomes and enzyme cofactorS Such as range of approximately 0.1 nm to about 5 um in diameter, biotin. Biomolecules (as well as other agents) may be more typically in the range of approximately 5 nm to about radioactively tagged or otherwise labeled for diagnostic 2.5 um. A “droplet” is distinguishable from a particle in that purposes, as will be discussed in further detail below. droplets are nonsolid. Suitable pharmacologically active peptides will generally The terms “pharmaceutical agent,” “active agent' and although not necessarily have a molecular weight of at least “drug” are used interchangeably herein to refer to a chemical 300 Da, and preferably at least 800 Da. Examples of such material or compound which, when administered to an 35 peptides which may be substantially stable in the extended organism (human or animal) induces a desired pharmaco release formulations over the intended period of release, and logical effect, including therapeutic effects, prophylactic which may therefore be used in the compositions of this effects, and diagnostic effects. invention, are oxytocin, Vasopressin, adrenocorticotropic By “pharmaceutically acceptable carrier' is meant a mate hormone (ACTH), epidermal growth factor (EGF), rial or materials that are Suitable for drug administration and 40 prolactin, luteinizing hormone, follicle Stimulating not biologically or otherwise undesirable, i.e., that may be hormone, luliberin or luteinizing hormone releasing hor administered to an individual along with an active agent mone (LHRH), insulin, Somatostatin, glucagon, interferon, without causing any undesirable biological effects or inter gastrin, tetragastrin, pentagastrin, urogastrone, Secretin, acting in a deleterious manner with any of the other com calcitonin, enkephalins, endorphins, kyotorphin, taftsin, ponents of the pharmaceutical formulation in which it is 45 thymopoietin, thymosin, thymoStimulin, thymic humoral contained. factor, Serum thymic factor, tumour necrosis factor, colony Similarly, a “pharmacologically acceptable' Salt, ester or Stimulating factors, motilin, bombesin, dinorphin, other derivative of an active agent as provided herein is a neurotensin, cerulein, bradykinin, urokinase, kallikrein, Sub Salt, ester or other derivative that is not biologically or stance P analogues and antagonists, angiotensin II, nerve otherwise undesirable. 50 growth factor, blood coagulation factors VII and IX, The term “reservoir” as used herein refers a receptacle or lysozyme chloride, re nin, bradykinin, tyrocidin, chamber for holding or containing a fluid. Thus, a fluid in a gramicidines, growth hormones, melanocyte Stimulating reservoir necessarily has a free Surface, i.e., a Surface that hormone, thyroid hormone releasing hormone, thyroid allows a droplet to be ejected therefrom. Stimulating hormone, parathyroid hormone, pancreozymin, “Optional' or “optionally” means that the Subsequently 55 cholecystokinin, human placental lactogen, human chori described circumstance may or may not occur, So that the onic gonadotropin, protein Synthesis Stimulating peptide, description includes instances where the circumstance gastric inhibitory peptide, vasoactive intestinal peptide, occurs and instances where it does not. platelet derived growth factor, growth hormone releasing The pharmaceutical agent may be any known or hereafter factor, bone morphogenic protein, and Synthetic analogues discovered pharmacologically active agent, and may be a 60 and modifications and pharmacologically active fragments compound that occurs in nature, a chemically modified thereof. Peptidyl drugs also include Synthetic analogs of naturally occurring compound, or a compound that is chemi LHRH, e.g., buSerelin, deslorelin, fertirelin, goserelin, cally Synthesized. The agent will typically be chosen from histrelin, leu prolide (leu prorelin), lutrelin, nafarelin, the generally recognized classes of pharmacologically active tryptorelin, and pharmacologically active Salts thereof. agents, including, but not necessarily limited to, the follow 65 Those skilled in the art will recognize that since the ing: analgesic agents, anesthetic agents, antiarthritic agents; invention is used in the manufacture of particles and respiratory drugs, including antiasthmatic agents, anticancer powders, the primary pharmaceutical agent candidates will US 6,596.206 B2 7 8 be those that are Suitable for administration of particulate Other additives commonly included in particulate phar dosage forms, e.g., using inhalation therapy and, for maceuticals include diluents, Stabilizers, Surfactants, example, a dry powder inhaler. Delivery of pharmaceutical lubricants, etc., and may be incorporated into the pharma particles via the respiratory System is of increasing interest ceutical particles prepared herein in an analogous manner. in the pharmaceutical field, particularly for those active For controlled release particles, from which the drug agents that are problematic when administered orally, e.g., releases gradually in a controlled manner, a biodegradable cause gastrointestinal distreSS and/or have variable rates of polymer may be incorporated into the Solid particles pre absorption and metabolism. pared according to the invention as described above for Pharmaceutical agents that are known candidates for carriers and other additives. The biodegradable polymer administration using dry powder inhalation therapy include 1O may be used in any convenient amount relative to the the following: peptidyl drugs, as described above; pharmaceutical agent, and the weight ratio of the biodegrad analgesics, e.g., codeine, dihydromorphine, ergotamine, fen able polymer to the pharmaceutical agent can vary from tanyl and morphine, anginal preparations, e.g., diltiazem; about 0.1 to 1 to about 100,000 to 1 depending upon the anti-allergy agents, e.g., cromoglycates Such as cromolyn application. Most controlled release applications, however, Sodium, ketotifen or nedocromil, antihistamines, e.g., meth 15 will involve a weight ratio of from about 10 to 1 to about 100 apyrilene; and respiratory drugs, i.e., pharmaceutical agents to 1. Any biodegradable polymer may be used that may be that are used in the treatment of respiratory diseases Such as co-dissolved in the organic Solvent along with the pharma asthma, bronchitis, emphysema and cystic fibrosis. Respi ceutical agent. Examples of Such biodegradable polymers ratory drugs include: anti-inflammatory corticosteroids, e.g., include those prepared by polymerization of at least one of flunisolide, flunisolide he mihydrate, budesonide, the following: an O-hydroxycarboxylic acid; a cyclic diester beclomethasone, beclomethasone monopropionate, beclom of an O-hydroxycarboxylic acid; a dioxanone; a lactone; a ethasone dipropionate, dexamethasone, dexamethasone cyclic carbonate; a cyclic oxalate; an epoxide, a glycol, and Sodium phosphate, fluticasone, and triamcinolone acetonide; an anhydride. Preferred biodegradable polymers are poly antituSSives, e.g., noScapine, bronchodilators, including B (ortho esters), poly (e-caprolactone), and poly (lactide-co adrenergic agonists, anticholinergic agents, and Xanthine 25 glycolide). The biodegradable polymers may be a derivatives, e.g. albuterol, bitolterol, clenbuterol, ephedrine, homopolymer or a copolymer of two or more different adrenaline, fenote rol, for mote rol, isoprenaline, monomers. Preferred homopolymers include poly(lactic isoproterenol, isoproterenol Sulfate, levalbuterol (i.e., homo acid), polylactide, poly(glycolic acid), polyglycolide and chiral (R)-albuterol), metaprote renol, metaproterenol poly(ethylene glycol). Sulfate, phenylephrine, pirbuterol, pirbuterol acetate, Depending on the hydrophilicity of the pharmaceutical procaterol, reproterol, rimiterol, Salmeterol, Salmeterol agent, Suitable Solvents for the pharmaceutical agent will be Xinotoate, terbutaline, and terbutaline Sulfate; either hydrophilic or lipophilic. Hydrophilic agents will of anticholinergics, e.g., ipratropium bromide, atropine and course dissolve in aqueous Solvents and other hydrophilic OXitropium; and Xanthines, e.g. aminophylline, choline solvents, while hydrophobic agents will dissolve in theophyllinate, lysine theophyllinate or theophylline. 35 lipophilic, nonaqueous Solvents, the latter including, for The pharmaceutical agent may also be a diagnostic agent, example, many organic Solvents and lipidic fluids. Super insofar as pulmonary administration of fine particles for critical fluids may also serve as Solvents for the pharmaceu diagnosis is well known. These diagnostic particles include tical agent, but will generally be used as antisolvents, i.e., as MRI contrast agents and radio-labeled compounds, known precipitating fluids. The Solution of the pharmaceutical to be useful for diagnosis of pulmonary abnormalities, 40 agent in the Solvent will generally although not necessarily including abnormalities of function and Structure, be Saturated, Such that SuperSaturation results when a Solu blockages, tumors, and the like. Examples of Such diagnos tion droplet contacts and admixes with the antisolvent. tic particles include, but are not limited to, Na-Fe Aqueous Solvents comprise water, and may additionally diethylenetriamine pentaacetic acid (DTPA), NaCr(DTPA), include other components that are Soluble or miscible in the NaGe(DTPA), gadolinium(III) contrast agents, radioactive 45 water, which components may be useful or desired for rhenium-containing or phosphorus-containing Salts, 2.2,6, particular applications. Thus, aqueous Solvents herein may 6,-tetramethyl-1-piperidinoxyl (TEMPO) spin-label agents include, but not limited to, water, a water-ethanol or water and other contrast aids. isopropanol admixture, ammonia water, a buffered aqueous It will be appreciated by those skilled in the art of medium, acidified water, basified water, and the like. pharmaceutical formulation and drug delivery that particu 50 Examples of organic Solvents include, without limitation: late dosage forms Such as those prepared herein will typi hydrocarbons, including aliphatic alkanes Such as hexane, cally include components other than the active agent. In heptane, decalin, octane, etc., cyclic alkanes Such as order to incorporate these additional components, they are cyclohexane, and aromatic hydrocarbons Such as benzene, added to the agent-containing Solvent prior to droplet ejec cumene, pyridine, pseudocumene, cymene, Styrene, toluene, tion into the antisolvent fluid. For example, a dry powder 55 Xylenes, tetrahydronaphthalene and mesitylene; halogenated composition for pulmonary administration will typically compounds Such as carbon tetrachloride and chlorinated, include a carrier Such as a mono-, di- or polysaccharide, fluorinated and brominated hydrocarbons Such as including dextrose (anhydrous and the monohydrate; also chloroform, bromoform, methyl chloroform, chlorobenzene, referred to as glucose and glucose monohydrate), lactitol, o-dichlorobenzene, chloroethane, 1,1-dichloroethane, 1,2- lactose, maltitol, maltose, Sucrose, fructose, galactose, 60 dichloroethane, tetrachloroethane, epichlorohydrin, trichlo mannitol, D-mannose, meleZitose, myoinositol, palatinite, roethylene and tetrachloroethylene, etherS Such as diethyl raffinose, Sorbitol, Sorbose, Stachyose, trehalose, Xylitol, ether, diisopropyl ether, diisobutyl ether, diglyme, 1,4- cellulosic polymerS Such as hydroxypropyl methylcellulose, dioxane, 1,3-dioxolane, dimethoxymethane, furan and tet cyclodextrins Such as 2-hydroxypropyl-3-cyclodextrin, or rahydrofuran, aldehydes Such as methyl formate, ethyl for the like, and the selected carrier would be dissolved in the 65 mate and furfural, ketones Such as acetone, disobutyl Solvent along with the pharmaceutical agent prior to droplet ketone, cyclohexanone, methyl ethyl ketone, N-methyl-2- ejection and particle formation. pyrrollidone and isophorone, amides Such as dimethyl for US 6,596.206 B2 10 mamide and dimethyl acetamide, alcohols Such as ethanol, or Supercritical processing. These fluids include, for isopropanol, n-propanol, t-butyl alcohol, cyclohexanol, example, carbon dioxide, water, ammonia, nitrogen, nitrous 1-hexanol, 1-octanol and trifluoroethanol; polyhydric alco oxide, methane, ethane, ethylene, propane, butane, hols Such as 1,3-propanediol, glycerol, ethylene glycol, n-pentane, benzene, methanol, ethanol, isopropanol, propylene glycol, and low molecular weight (typically less isobutanol, halocarbons (including monofluoromethane, than 400) polyethylene glycol, amines, including cyclic trifluorome thane, chloro trifluorome thane, amines Such as pyridine, piperidine, 2-methylpyridine, monofluoromethane, hexafluoroethane 1,1-difluoroethylene morpholine, etc., and mono-, di- and tri-Substituted amines and 1,2-difluoroethylene), toluene, pyridine, cyclohexane, Such as trimethylamine, dimethylamine, methylamine, m-creSol, decalin, cyclohexanol, O-Xylene, tetralin, aniline, triethylamine, diethylamine, ethylamine, n-butylamine, acetylene, chlorotrifluorosilane, Xenon, Sulfur hexafluoride, t-butylamine, trie thanolamine, diethanolamine and propane, and others. CoSolvents or other modifiers may be ethanolamine, and amine-Substituted hydrocarbons Such as added to the Supercritical fluid to change its intrinsic prop ethylene diamine, diethylene triamine; carboxylic acids Such erties in or around the critical point. Suitable coSolvents and as acetic acid, trifluoroacetic acid and formic acid; esters modifiers are known in the art and include, for example, Such as ethyl acetate, isopentyl acetate, propylacetate, etc.; 15 methanol, ethanol, isopropanol and acetone. lactams Such as caprolactam, nitriles Such as acetonitrile, For many pharmaceutical agents, it is desirable to use an propane nitrile and adiponitrile, organic nitrates Such as antisolvent fluid that permits processing at relatively mild nitrobenzene, nitroethane and nitromethane, and Sulfides temperatures. This is particularly important for processing Such as carbon disulfide. peptidyl drugs, which are Susceptible to a loSS of biological The Solvent may also be a lipidic material including, but activity when Subjected either to very low temperatures or to not limited to, the following: phospholipids Such as phos very high temperatures. For processing of proteins and phorylated diacylglycerides, and particularly phospholipids polypeptides, then, the antisolvent fluid should preferably Selected from the group consisting of diacyl have a critical temperature of from about 0°C. to about 50 phosphatidylcholines, diacyl phosphatidylethanolamines, C. Included in this category of antisolvent fluids are carbon diacyl phosphatidylserines, diacyl phosphatidylinositols, 25 dioxide, nitrogen, nitrous oxide, ethane, ethylene, diacyl phosphatidylglycerols, diacyl phosphatidic acids, and chlorotrifluoromethane, monofluoromethane, acetylene, 1,1- mixtures thereof, wherein each acyl group contains about 10 difluoroethylene, hexafluoroethane, chlorotrifluorosilane, to about 22 carbon atoms and is Saturated or unsaturated; and Xenon. A particularly preferred antisolvent fluid is fatty acids Such as isoValeric acid, Valeric acid, caproic acid, carbon dioxide because it is readily available, non-toxic (it enanthic acid, caprylic acid, pelargonic acid, capric acid, has "GRAS, or “generally regarded as Safe,” status), non lauric acid, myristic acid, palmitic acid, Stearic acid, flammable, relatively low cost, has low chemical reactivity, arachidic acid, behenic acid, lignoceric acid, oleic acid, and has a critical temperature of 31.3 C. and a critical linoleic acid, linolenic acid, and arachidonic acid; lower pressure of 72.9 atm (1072 psi), which permits processing fatty acid esters comprising esters of the foregoing fatty under relatively mild conditions. Another preferred Super acids, wherein the carboxylic acid group of the fatty acid is 35 critical fluid is nitrogen. replaced with an ester moiety -(CO)-OR wherein R is a After preparation, the drug-containing particles are C-C alkyl moiety optionally Substituted with one or two packaged, as is or in dosage forms, preferably dosage forms hydroxyl groups, fatty alcohols corresponding to the afore containing a unit dose of the active agent. For pulmonary mentioned fatty acids, wherein the carboxylic acid group of administration, the powderS may be incorporated into a the fatty acid is replaced by a -CH-OH group; glycolipids 40 Suitable inhaler. For oral delivery pharmaceutical powders Such as cerebroside and gangliosides, oils, including animal may be dissolved in water prior to administration, although oils. Such as cod liver oil and, menhaden oil, and vegetable capsules are preferred for oral administration. Suitable cap oils. Such as babasSu oil, castor oil, corn oil, cotton Seed oil, Sule materials may be either hard or Soft, and as will be linseed oil, mustard oil, olive oil, palm oil, palm kernel oil, appreciated by those skilled in the art, typically comprise a peanut oil, poppy Seed oil, rapeseed oil, Safflower oil, Sesame 45 tasteless, easily administered and water Soluble compound oil, Soybean oil, Sunflower Seed oil, tung oil or wheat germ Such as gelatin, Starch or a cellulosic material. See Reming oil, and waxes, i.e., higher fatty acid esters, including animal ton. The Science and Practice of Pharmacy, Nineteenth waxes. Such as beeswax and shellac, mineral waxes Such as Edition (Easton, Pa.; Mack Publishing Co., 1995), which montan, petroleum waxes. Such as microcrystalline wax and describes materials and methods for preparing encapsulated paraffin, and vegetable waxes Such as carnauba wax. 50 pharmaceuticals. Each capsule will typically contain a thera In a preferred embodiment, a Supercritical fluid is used as peutically effective dose of the active agent. Alternatively, the antisolvent, or precipitating fluid, effective to cause the the dosage forms may contain less than a therapeutically pharmaceutical agent to precipitate upon admixture with the effective dose in which case administration of two or more pharmaceutical agent-Solvent Solution. Although it is pref dosage forms would be required to achieve the therapeuti erable that the agent be insoluble in the antisolvent, it is 55 cally effective dose. Sufficient for the purpose of the present invention that the It will also be appreciated by those of skill in the art that agent be leSS Soluble in the antisolvent than in the Solvent. particulate pharmaceutical preparations may be manufac Thus, it is possible that the antisolvent fluid may be capable tured in other ways, and administered using any convenient of dissolving Some amount of the pharmaceutical agent, method. For example, douche powders are typically dis although in a preferred embodiment, the antisolvent fluid is 60 Solved in water prior to use as antiseptics or as cleansing Substantially incapable of dissolving a significant portion of agents for a body cavity, typically for Vaginal the pharmaceutical agent. Also, the antisolvent fluid is at administration, but also for nasal, otic or ophthalmic use. least partially miscible with the organic Solvent Such that the The powderS may also be used as “dusting powders, i.e., antisolvent fluid is capable of penetrating into the pharma locally applied preparations that are intended to have no ceutical agent-Solvent Solution Sufficiently to cause the 65 Systemic action, used, for example, as lubricants, desired precipitation of the pharmaceutical agent. The anti protectives, absorbents, antiseptics, antipruritics, antibrom Solvent fluid may comprise any Suitable fluid for near critical hidrosis agents, astringens and antiperSpirants. Powders may US 6,596.206 B2 11 12 also be introduced into the body directly, without a carrier The device also includes an acoustic ejector 33 comprised liquid, as “insufflations, in which case an insufflator of an acoustic radiation generator 35 for generating acoustic (powder blower) or pressure aerosol device is used to radiation and a focusing means 37 for focusing the acoustic introduce the powder into body cavities Such as the ears, radiation at a focal point within the upper fluid layer or the nose and throat. It may be necessary to dilute the pharma lower fluid layer, but is preferably just below the interface ceutical powder with an inactive Solid carrier, also in powder therebetween. As shown in FIG. 1, the focusing means 37 form, to produce what is commonly called a “trituration.” may comprise a Single Solid piece having a concave Surface In preparing Solid particles according to the invention, 39 for focusing acoustic radiation, but the focusing means focused acoustic energy is used Substantially as described in may be constructed in other ways as discussed below. The detail in co-pending patent applications U.S. Ser. No. acoustic ejector 33 is thus adapted to generate and focus 09/669,996 (“Acoustic Ejection of Fluids From a Plurality acoustic radiation So as to eject a droplet of fluid from the of Reservoirs”), inventors Ellson and Foote, and U.S. Ser. Solution into the antisolvent when acoustically coupled to No. 09/669,194 (“Method and Device for Generating Drop the reservoirs. The acoustic radiation generator 35 and the lets of Immiscible Fluids”), inventors Ellson, Mutz and focusing means 37 may function as a Single unit controlled Foote, both filed on Sep. 25, 2000 and assigned to Picoliter 15 by a Single controller, or they may be independently Inc. (Sunnyvale, Calif.). FIG. 1 illustrates a focused acoustic controlled, depending on the desired performance of the ejection device that can be used in accordance with the foregoing method. The device is shown in Simplified croSS device. Typically, Single ejector designs are preferred over Sectional view, wherein, as with all figures referenced multiple ejector designs because accuracy of droplet Veloc herein, like parts are referenced by like numerals, the figure ity (i.e., both the speed and direction of the ejected droplet) is not necessarily to Scale, and certain dimensions may be and consistency in droplet size and Velocity are more easily exaggerated for clarity of presentation. The device 11 may achieved with a single ejector. include a Single reservoir, two reservoirs, or a plurality of AS will be appreciated by those skilled in the art, any of reservoirs. For Simplicity, the device is illustrated as con a variety of focusing means may be employed in conjunction taining two reservoirs, with a first reservoir indicated at 13 with the present invention. For example, one or more curved and a second reservoir indicated at 15. Each reservoir 25 Surfaces may be used to direct acoustic radiation to a focal contains a Solution of the pharmaceutical agent provided in point near a fluid Surface. One Such technique is described a Solvent, with the antisolvent in a contained space generally in U.S. Pat. No. 4,308,547 to Lovelady et al. Focusing above and in fluid communication with the Solution in the means with a curved Surface have been incorporated into the reservoir Such that droplets ejected from the Solution are construction of commercially available acoustic transducers directed into the antisolvent. The Solvent may comprise a such as those manufactured by Panametrics Inc. (Waltham, mixture of Solvents, and the antisolvent, Similarly, may Mass.). In addition, Fresnel lenses are known in the art for comprise a mixture of antisolvents. Reservoirs containing directing acoustic energy at a predetermined focal distance both the Solution and the antisolvent are illustrated in the from an object plane. See, e.g., U.S. Pat. No. 5,041,849 to figure, with reservoir 13 containing the pharmaceutical Quate et al. Fresnel lenses may have a radial phase profile agent-solvent solution as a lower fluid 10 and the antisol 35 that diffracts a Substantial portion of acoustic energy into a vent as an upper fluid 12, and, analogously, reservoir 15 predetermined diffraction order at diffraction angles that containing the pharmaceutical agent Solution as a lower fluid vary radially with respect to the lens. The diffraction angles 14 and the antisolvent as an upper fluid 16. Generally, with should be selected to focus the acoustic energy within the liquid antisolvents, the upper fluid layer comprised of the diffraction order on a desired object plane. antisolvent will have a thickness (i.e., height) that is less 40 There are also a number of ways to acoustically couple the than about 10% of the thickness of the lower fluid layer ejector 33 to an individual reservoir and thus to the fluid comprised of the Solvent, typically in the range of about therein. One Such approach is through direct contact as is 0.1% to 5% of the thickness of the lower fluid. It must be described, for example, in U.S. Pat. No. 4,308,547 to emphasized, however, that the two-phase System is illus Lovelady et al., wherein a focusing means constructed from trated and described merely for Simplicity, and each reser 45 a hemispherical crystal having Segmented electrodes is voir may contain three or more fluids, and the invention is Submerged in a liquid to be ejected. The aforementioned not limited in this regard. For example, a third fluid may be patent further discloses that the focusing means may be incorporated So as to physically Segregate the Solvent and positioned at or below the surface of the liquid. However, antisolvent. Alternatively, the Solvent and antisolvent may this approach for acoustically coupling the focusing means be maintained in a Substantially Segregated State by main 50 to a fluid is undesirable when the ejector is used to eject taining the Solvent, the antisolvent, or both, at a predeter different fluids in a plurality of containers or reservoirs, as mined temperature and pressure. repeated cleaning of the focusing means would be required With liquid antisolvents, upper fluid layers 12 and 16 will in order to avoid cross-contamination; contamination of any have fluid surfaces respectively indicated at 17 and 19. As Sort must, of course, be avoided in the preparation of shown, the reservoirs are of Substantially identical construc 55 pharmaceuticals. The cleaning process would necessarily tion So as to be Substantially acoustically indistinguishable, lengthen the transition time between each droplet ejection but identical construction is not a requirement. The reser event. In addition, in Such a method, fluid would adhere to voirs are shown as Separate removable components but may, the ejector as it is removed from each container, wasting if desired, be fixed within a plate or other substrate. For material that may be costly or rare. example, the reservoirs may be individual wells in a well 60 Thus, a preferred approach, when a multiple reservoir plate. Each of the reservoirs 13 and 15 is preferably axially System is employed, is to acoustically couple the ejector to Symmetric as shown, having vertical walls 21 and 23 extend the reservoirs and reservoir fluids without contacting any ing upward from circular reservoir bases 25 and 27 and portion of the ejector, e.g., the focusing means, with any of terminating at openingS 29 and 31, respectively, although the fluids to be ejected. To this end, an ejector positioning other reservoir shapes may be used. The material and 65 means may be provided for positioning the ejector in con thickness of each reservoir base should be Such that acoustic trolled and repeatable acoustic coupling with each of the radiation may be transmitted therethrough. fluids in the reservoirs to eject droplets therefrom without US 6,596.206 B2 13 14 Submerging the ejector therein. This typically involves direct In addition, the material used in the construction of reser or indirect contact between the ejector and the external voirs must be compatible with the fluids contained therein. Surface of each reservoir. When direct contact is used in Thus, if it is intended that the reservoirs or wells contain a order to acoustically couple the ejector to each reservoir, it particular organic Solvent, polymers that dissolve or Swell in is preferred that the direct contact is wholly conformal to that solvent would be unsuitable for use in forming the ensure efficient acoustic energy transfer. That is, the ejector reservoirs or well plates. For water-based fluids, a number of and the reservoir should have corresponding Surfaces materials are Suitable for the construction of reservoirs and adapted for mating contact. Thus, if acoustic coupling is include, but are not limited to, ceramics Such as Silicon oxide achieved between the ejector and reservoir through the and aluminum oxide, metals Such as StainleSS Steel and focusing means, it is desirable for the reservoir to have an 1O platinum, and polymerS Such as polyester and polytetrafluo outside Surface that corresponds to the Surface profile of the roethylene. Many well plates are commercially available focusing means. Without conformal contact, efficiency and and may contain, for example, 96, 384 or 1536 wells per accuracy of acoustic energy transfer may be compromised. well plate. Manufactures of suitable well plates for use in the In addition, Since many focusing means have a curved inventive device include Corning Inc. (Corning, N.Y.) and Surface, the direct contact approach may necessitate the use 15 Greiner America, Inc. (Lake Mary, Fla.). However, the of reservoirs having a Specially formed inverse Surface. availability of such commercially available well plates does Optimally, acoustic coupling is achieved between the not preclude manufacture and use of custom-made well ejector and the reservoir through indirect contact, as illus plates containing at least about 10,000 wells, or as many as trated in FIG. 1A. In the figure, an acoustic coupling 100,000 wells or more. In addition, for multiple reservoir medium 41 is placed between the ejector 33 and the base 25 Systems, it is preferable that the center of each reservoir is of reservoir 13, with the ejector and reservoir located at a located not more than about 1 centimeter, preferably not predetermined distance from each other. The acoustic cou more than about 1 millimeter and optimally not more than pling medium may be an acoustic coupling fluid, preferably about 0.5 millimeter from another reservoir center. In order an acoustically homogeneous material in conformal contact to ensure the accuracy of fluid ejection, it is important to with both the acoustic focusing means 37 and each reservoir. 25 determine the location and the orientation of the fluid In addition, it is important to ensure that the fluid medium Surface from which a droplet is to be ejected with respect to is Substantially free of material having different acoustic the ejector. Otherwise, ejected droplets may be improperly properties than the fluid medium itself. AS Shown, the sized or travel in an improper trajectory. Thus, another reservoir 13 is acoustically coupled to the acoustic focusing embodiment of the invention relates to a method for deter means 37 Such that an acoustic wave is generated by the mining the height of a fluid Surface in a reservoir between acoustic radiation generator and directed by the focusing ejection events. The method involves activating the acoustic means 37 into the acoustic coupling medium 41, which then generator to produce a detection acoustic wave that travels transmits the acoustic radiation into the reservoir. The acous to the fluid surface and is reflected thereby as a reflected tic ejector 33 is positionable by means of ejector positioning acoustic wave. Parameters of the reflected acoustic radiation means 43, shown below reservoir 13, in order to achieve 35 are then analyzed in order to assess the Spatial relationship acoustic coupling between the ejector and the reservoir between the acoustic radiation generator and the fluid Sur through acoustic coupling medium 41. Once the ejector, the face. Such an analysis will involve the determination of the reservoir and the Substrate are in proper alignment, the distance between the acoustic radiation generator and the acoustic radiation generator 35 is activated to produce fluid Surface and/or the orientation of the fluid Surface in acoustic radiation that is directed by the focusing means 37 40 relationship to the acoustic radiation generator. to a focal point within solution 10 in the first reservoir. As More particularly, the acoustic radiation generator may a result, droplet 47 is ejected from the solution, into the activated So as to generate low energy acoustic radiation that antisolvent 12, and optionally onto a site on the underside is insufficiently energetic to eject a droplet from the fluid Surface 51 of the enclosure 45. Surface in the reservoir. This is typically done by using an Then, as shown in FIG. 1B, the ejector 33 has been 45 extremely short pulse (on the order of tens of nanoseconds) repositioned below reservoir 15 and in acoustically coupled relative to that normally required for droplet ejection (on the relationship thereto by Virtue of acoustic coupling medium order of microSeconds). By determining the time it takes for 41. Once properly aligned as shown in FIG. 1B, the acoustic the acoustic radiation to be reflected by the fluid surface radiation generator 35 of ejector 33 is activated to produce back to the acoustic radiation generator and then correlating acoustic radiation that is then directed by focusing means 37 50 that time with the speed of Sound in the fluid, the distance to a focal point within the solution in reservoir 15, thereby and thus the fluid height-may be calculated. Of course, ejecting droplet 55 into the antisolvent 16 and optionally care must be taken in order to ensure that acoustic radiation onto the underside 53 of enclosure 49. AS discussed above, reflected by the interface between the reservoir base and the a single reservoir may be used or two or more reservoirs may fluid is discounted. It will be appreciated by those of be used, wherein the well(s) may be removable or contained 55 ordinary skill in the art of acoustic microScopy that Such a within a well plate. Similarly, a Single ejector can be used, method employs conventional or modified Sonar techniques. as shown in the figures, or a plurality of ejectors can be used Once the analysis has been performed, an ejection acous with each ejector positioned So as to eject a droplet from a tic wave having a focal point near the fluid Surface is different locus within a single reservoir (if a single reservoir generated in order to eject at least one droplet of the fluid, is used) or from each of a plurality of reservoirs. Also, unless 60 wherein the optimum intensity and directionality of the it is intended that the ejector is to be submerged in the fluids ejection acoustic wave is determined using the aforemen to be ejected, the reservoirs or well plates must have acoustic tioned analysis optionally in combination with additional transmission properties Sufficient to allow acoustic radiation data. The “optimum” intensity and directionality are gener from the ejector to be conveyed to the surfaces of the fluids ally Selected to produce droplets of consistent size and to be ejected. Typically, this involves providing reservoir or 65 Velocity. For example, the desired intensity and direction well bases that are Sufficiently thin to allow acoustic radia ality of the ejection acoustic wave may be determined by tion to travel therethrough without unacceptable dissipation. using not only the Spatial relationship assessed as above, but US 6,596.206 B2 15 16 also geometric data associated with the reservoir, fluid I claim: property data associated with the fluid to be ejected, and/or 1. A method for generating a Solid particle containing a by using historical droplet ejection data associated with the pharmaceutical agent, the method comprising: ejection Sequence. In addition, the data may show the need providing a reservoir containing a Solution of the phar to reposition the ejector So as to reposition the acoustic maceutical agent in a Solvent; radiation generator with respect to the fluid Surface, in order providing an antisolvent in a contained Space in fluid to ensure that the focal point of the ejection acoustic wave communication with the Solution in the reservoir, is near the fluid Surface, where desired. For example, if wherein the antisolvent is Selected Such that the phar analysis reveals that the acoustic radiation generator is maceutical agent is leSS Soluble in the antisolvent than positioned Such that the ejection acoustic wave cannot be in the Solvent, and focused near the fluid Surface, the acoustic radiation gen applying focused acoustic energy to the reservoir So as to erator is repositioned using vertical, horizontal and/or rota produce a droplet of the Solution in the antisolvent in tional movement to allow appropriate focusing of the ejec the contained Space, whereby admixture of the Solution droplet and the antisolvent results in the precipitation of tion acoustic wave. the pharmaceutical agent, forming a Solid particle. The device may include other components that enhance 15 2. The method of claim 1, wherein focused acoustic performance. For example, the device may further comprise energy is applied to the reservoir at a plurality of loci So as cooling means for lowering the temperature of a particle to eject a plurality of droplets, whereby a plurality of Solid collection means (e.g., an upper Surface within the contained particles is provided. enclosure) positioned above the reservoir. The device may 3. The method of claim 1, wherein the solvent is acqueous. also comprise a heating means for maintaining the fluid in 4. The method of claim 1, wherein the Solvent is non the reservoir at a constant temperature, and which, in aqueous. combination with a pressurizing means, for maintaining the 5. The method of claim 4, wherein the solvent is organic. antisolvent in a Supercritical State. Design and construction 6. The method of claim 1, wherein the solvent is a of Such temperature maintaining means and preSSurizing Supercritical fluid. 25 7. The method of claim 1, wherein the antisolvent is a means are known to one of ordinary skill in the art. For many Supercritical fluid. biomolecular applications, it is generally desired that the 8. The method of claim 4, wherein the antisolvent is a fluid containing the biomolecule is kept at a constant tem Supercritical fluid. perature without deviating more than about 1 C. or 2 C. 9. The method of claim 5, wherein the antisolvent is a therefrom. In addition, for a biomolecular fluid that is Supercritical fluid. particularly heat sensitive, it is preferred that the fluid be 10. The method of claim 1, wherein the antisolvent is kept at a temperature that does not exceed about 10 C. gaseous. above the melting point of the fluid, preferably at a tem 11. The method of claim 1, wherein the precipitation of perature that does not exceed about 5 C. above the melting the pharmaceutical agent comprises crystallization thereof. point of the fluid. Thus, for example, when the biomolecule 12. The method of claim 1, wherein the Solution and the containing fluid is acqueous, it may be optimal to keep the 35 antisolvent are both present in the reservoir. fluid at about 4 C. during ejection. 13. The method of claim 12, wherein a third fluid is present as a Segregating layer between the Solution and the The device of the invention enables ejection of droplets at antisolvent. a rate of at least about 1,000,000 droplets per minute from 14. The method of claim 12, wherein the Solvent and the same reservoir, and at a rate of at least about 100,000 40 antisolvent are maintained in a Substantially Segregated State drops per minute from different reservoirs. In addition, by maintaining the Solvent, the antisolvent, or both, at a current positioning technology allows for the ejector posi predetermined temperature and pressure. tioning means to move from one reservoir to another quickly 15. The method of any one of claims 7, 8, 9, 12, 13 and and in a controlled manner, thereby allowing fast and 14, wherein the antisolvent is Selected from the group controlled ejection of different fluids. That is, current com 45 consisting of carbon dioxide, water, ammonia, nitrogen, mercially available technology allows the ejector to be nitrous oxide, methane, ethane, ethylene, propane, butane, moved from one reservoir to another, with repeatable and n-pentane, benzene, methanol, ethanol, isopropanol, controlled acoustic coupling at each reservoir, in less than isobutanol, monofluorome thane, trifluorome thane, about 0.1 Second for high performance positioning means chlorot rifluorome thane, monofluorome thane, and in less than about 1 Second for ordinary positioning 50 he Xafluoroethane, 1,1-difluoroethylene, 1,2- means. A custom designed System will allow the ejector to difluoroethylene, toluene, pyridine, cyclohexane, m-creSol, be moved from one reservoir to another with repeatable and decalin, cyclohexanol, O-Xylene, tetralin, aniline, acetylene, controlled acoustic coupling in less than about 0.001 Second. chlorotrifluorosilane, Xenon, Sulfur hexafluoride, propane, In order to provide a custom designed System, it is important and combinations thereof. to keep in mind that there are two basic kinds of motion: 55 16. The method of claim 15, wherein the antisolvent is pulse and continuous. Pulse motion involves the discrete carbon dioxide. Steps of moving an ejector into position, emitting acoustic 17. The method of claim 1, wherein the solution droplet energy, and moving the ejector to the next position; again, is ejected onto a Surface of a SubStrate having the antisolvent using a high performance positioning means with Such a thereon. method allows repeatable and controlled acoustic coupling 60 18. The method of claim 1, wherein the Solution is a at each reservoir in less than 0.1 Second. A continuous Saturated Solution. motion design, on the other hand, moves the ejector and the 19. The method of claim 1, wherein the pharmaceutical reservoirs continuously, although not at the same Speed, and agent is hydrophilic, the Solvent is hydrophilic, and the provides for ejection during movement. Since the pulse antisolvent is lipophilic. width is very short, this type of process enables over 10 Hz 65 20. The method of claim 19, wherein the pharmaceutical reservoir transitions, and even over 1000 Hz reservoir tran agent is hydrophilic, the Solvent is acqueous, and the anti Sitions. Solvent is lipidic. US 6,596.206 B2 17 18 21. The method of claim 1, wherein the pharmaceutical 42. The method of claim 41, wherein the diagnostic agent agent is lipophilic, the Solvent is lipophilic, and the antisol is a radio-labeled compound. vent is hydrophilic. 43. The method of claim 1, wherein the Solution further 22. The method of claim 12, wherein the Solution is a comprises at least one pharmaceutical excipient. lower layer having a first thickness and the antisolvent is an 44. The method of claim 1, wherein the Solution further upper layer having a Second thickness less than about 10% comprises a biodegradable polymer. of the first thickness. 45. The method of claim 1, wherein the reservoir is 23. The method of claim 22, wherein the second thickness is in the range of about 0.1% to 5% of the first thickness. adapted to contain no more than about 100 nanoliters of 24. The method of claim 22, wherein the upper layer is a fluid. molecular monolayer. 1O 46. The method of claim 45, wherein the reservoir is 25. The method of claim 22, wherein the upper layer is a adapted to contain no more than about 10 nanoliters of fluid. molecular bilayer. 47. A method for generating a plurality of particles 26. The method of claim 1, wherein the size of the particle containing a pharmaceutical agent, the method comprising: is in the range of approximately 0.1 nm to about 5 um. providing a plurality of reservoirs each containing a 27. The method of claim 26, wherein the size of the 15 Solution of the pharmaceutical agent in a Solvent; particle is in the range of approximately 5 nm to about 2.5 providing an antisolvent in a contained Space in fluid plm. communication with the Solution in each reservoir, 28. The method of claim 1, wherein the pharmaceutical wherein the antisolvent is Selected Such that the phar agent is Selected from the group consisting of analgesic maceutical agent is leSS Soluble in the antisolvent than agents, anesthetic agents, antiarthritic agents, respiratory in the Solvent, and drugs, anticancer agents, anticholinergics, anticonvulsants; applying focused acoustic energy to each reservoir So as antidepressants, antidiabetic agents, antidiarrheals; antihel to produce droplets of the Solution into the antisolvent minthics, antihistamines, antihyperlipidemic agents, antihy in the contained space, whereby admixture of the pertensive agents, anti-infective agents, antiinflammatory Solution droplets and the antisolvent results in the agents, antimigraine preparations, antinauseants, antine 25 precipitation of the pharmaceutical agent, forming Solid oplastic agents, antiparkinsonism drugs, antipruritics, antip particles. Sychotics, antipyretics; antispasmodics, antitubercular 48. The method of claim 47, wherein the focused acoustic agents, antiulcer agents, antiviral agents, anxiolytics, appe energy is applied to each reservoir Simultaneously using a tite Suppressants; attention deficit disorder (ADD) and atten plurality of acoustic ejection devices. tion deficit hyperactivity disorder (ADHD) drugs; cardio 49. The method of claim 47, wherein the focused acoustic vascular preparations, central nervous System Stimulants, energy is applied to each reservoir in Succession using a cough and cold preparations, hormonolytics, hypnotics, Single acoustic ejection device. hypoglycemic agents, immunOSuppressive agents, leukot 50. A device for making solid particles of a pharmaceu riene inhibitors, mitotic inhibitors, muscle relaxants, nar tical agent, comprising: cotic antagonists, nicotine, nutritional agents, ophthalmic 35 drugs, parasympatholytics, psychoStimulants, Sedatives, Ste a reservoir containing a Solution of the pharmaceutical roids, Sympathomimetics, tranquilizers, vasodilators, and agent in a Solvent; combinations thereof. an antisolvent in a contained Space in fluid communica 29. The method of claim 1, wherein the pharmaceutical tion with the solution in the reservoir such that droplets agent is a biomolecule Selected from the group consisting of 40 ejected from the Solution are directed into the DNA, RNA, antisense oligonucleotides, peptides, proteins, antisolvent, wherein the antisolvent is Selected Such ribosomes and enzyme cofactors. that the pharmaceutical agent is less Soluble in the 30. The method of claim 29, wherein the pharmaceutical antisolvent than in the Solvent; agent is DNA. an acoustic ejector comprising an acoustic radiation gen 31. The method of claim 29, wherein the pharmaceutical 45 erator for generating acoustic radiation and a focusing agent is RNA. means for focusing the acoustic radiation at a focal 32. The method of claim 29, wherein the pharmaceutical point within the Solution in the reservoir So as to eject agent is an antisense oligonucleotide. a droplet therefrom; and 33. The method of claim 29, wherein the pharmaceutical a means for positioning the ejector in acoustic coupling agent is a peptide or protein. 50 relationship to the reservoir. 34. The method of claim 29, wherein the pharmaceutical 51. The device of claim 50, comprising a single acoustic agent is a protein. ejector. 35. The method of claim 34, wherein the protein is a 52. The device of claim 50, comprising a plurality of fluorescent protein. acoustic ejectors positioned to direct focused acoustic 36. The method of claim 29, wherein the pharmaceutical 55 energy to a plurality of loci within the Solution So as to eject agent is a ribosome. a plurality of droplets, whereby a plurality of Solid particles 37. The method of claim 29, wherein the pharmaceutical is provided. agent is an enzyme cofactor. 53. The device of claim 50, wherein the reservoir is 38. The method of claim 29, wherein the pharmaceutical adapted to contain no more than about 100 nanoliters of agent is a respiratory drug. 60 fluid. 39. The method of claim 38, wherein the respiratory drug 54. The device of claim 50, wherein the reservoir is is Selected from the group consisting of anti-inflammatory adapted to contain no more than about 10 nanoliters of fluid. corticosteroids, bronchodilators, and mixtures thereof. 55. The device of claim 50, further comprising a means 40. The method of claim 1, wherein the pharmaceutical for maintaining the Solvent in the reservoir at a constant agent is a diagnostic agent. 65 temperature. 41. The method of claim 40, wherein the diagnostic agent 56. The device of claim 50, wherein the acoustic coupling is an MRI contrast agent. relationship between the ejector and the Solution in the US 6,596.206 B2 19 20 reservoir is established by providing an acoustically con agents, anesthetic agents, antiarthritic agents, respiratory ductive medium between the ejector and the reservoir. drugs, anticancer agents, anticholinergics, anticonvulsants; 57. The device of claim 50, wherein acoustic coupling antidepressants, antidiabetic agents, antidiarrheals, antihel between the ejector and the fluid in each reservoir is estab minthics, antihistamines, antihyperlipidemic agents, antihy lished at a predetermined distance between the ejector and pertensive agents, anti-infective agents, antiinflammatory each reservoir. agents, antimigraine preparations, antinauseants, antine 58. The device of claim 50, wherein the solvent is oplastic agents, antiparkinsonism drugs, antipruritics, antip aqueous. Sychotics, antipyretics; antispasmodics, antitubercular 59. The device of claim 50, wherein the solvent is agents, antiulcer agents, antiviral agents, anxiolytics, appe non-aqueous. tite Suppressants; attention deficit disorder (ADD) and atten 60. The device of claim 59, wherein the solvent is organic. tion deficit hyperactivity disorder (ADHD) drugs; cardio 61. The device of claim 50, wherein the solvent is a vascular preparations, central nervous System Stimulants, Supercritical fluid. cough and cold preparations, hormonolytics, hypnotics, 62. The device of claim 50, wherein the antisolvent is a hypoglycemic agents, immunosuppressive agents, leukot Supercritical fluid. 15 riene inhibitors, mitotic inhibitors, muscle relaxants, nar 63. The device of claim 59, wherein the antisolvent is a cotic antagonists, nicotine, nutritional agents, ophthalmic Supercritical fluid. drugs, parasympatholytics, psychoStimulants, Sedatives, Ste 64. The device of claim 60, wherein the antisolvent is a roids, Sympathomimetics, tranquilizers, vasodilators, and Supercritical fluid. combinations thereof. 65. The device of claim 50, wherein the antisolvent is 80. The device of claim 50, wherein the pharmaceutical gaseous. agent is a biomolecule Selected from the group consisting of 66. The device of claim 50, wherein the Solution and the DNA, RNA, antisense oligonucleotides, peptides, proteins, antisolvent are both present in the reservoir. ribosomes and enzyme cofactors. 67. The device of claim 66, wherein a third fluid is present 81. The device of claim 80, wherein the pharmaceutical as a Segregating layer between the Solution and the antisol 25 agent is DNA. Vent. 82. The device of claim 80, wherein the pharmaceutical 68. The device of claim 66, wherein the solvent and agent is RNA. antisolvent are maintained in a Substantially Segregated State 83. The device of claim 80, wherein the pharmaceutical by maintaining the Solvent, the antisolvent, or both, at a agent is an antisense oligonucleotide. predetermined temperature and pressure. 84. The device of claim 80, wherein the pharmaceutical 69. The device of any one of claims 62, 63, 64, 66, 67 and agent is a peptide or protein. 68, wherein the antisolvent is selected from the group 85. The device of claim 80, wherein the pharmaceutical consisting of carbon dioxide, Water, ammonia, nitrogen, agent is a protein. nitrous oxide, methane, ethane, ethylene, propane, butane, 86. The device of claim 85, wherein the protein is a n-pentane, benzene, methanol, ethanol, isopropanol, 35 fluorescent protein. isobutanol, monofluorome thane, trifluorome thane, 87. The device of claim 80, wherein the pharmaceutical chlorot rifluorome thane, monofluorome thane, agent is a ribosome. he Xafluoroethane, 1,1-difluoroethylene, 1,2- 88. The device of claim 80, wherein the pharmaceutical difluoroethylene, toluene, pyridine, cyclohexane, m-creSol, agent is an enzyme cofactor. decalin, cyclohexanol, O-Xylene, tetralin, aniline, acetylene, 40 89. The device of claim 80, wherein the pharmaceutical chlorotrifluorosilane, Xenon, Sulfur hexafluoride, propane, agent is a respiratory drug. and combinations thereof. 90. The device of claim 89, wherein the respiratory drug 70. The device of claim 69, wherein the antisolvent is is Selected from the group consisting of anti-inflammatory carbon dioxide. corticosteroids, bronchodilators, and mixtures thereof. 71. The device of claim 50, wherein the Solution is a 45 91. The device of claim 80, wherein the pharmaceutical Saturated Solution. agent is a diagnostic agent. 72. The device of claim 50, wherein the pharmaceutical 92. The device of claim 50, wherein the diagnostic agent agent is hydrophilic, the Solvent is hydrophilic, and the is an MRI contrast agent. antisolvent is lipophilic. 93. The device of claim 50, wherein the diagnostic agent 73. The device of claim 72, wherein the pharmaceutical 50 is a radio-labeled compound. agent is hydrophilic, the Solvent is acqueous, and the anti 94. The device of claim 50, wherein the Solution further Solvent is lipidic. comprises at least one pharmaceutical excipient. 74. The device of claim 50, wherein the pharmaceutical 95. The device of claim 50, wherein the solution further agent is lipophilic, the Solvent is lipophilic, and the antisol comprises a biodegradable polymer. vent is hydrophilic. 55 96. A device for making Solid particles of a pharmaceu 75. The device of claim 66, wherein the Solution is a lower tical agent, comprising: layer having a first thickness and the antisolvent is an upper a plurality of reservoirs each containing a Solution of the layer having a second thickness less than about 10% of the pharmaceutical agent in a Solvent; first thickness. an antisolvent in a contained Space above and in fluid 76. The device of claim 75, wherein the second thickness 60 communication with the Solution in each reservoir Such is in the range of about 0.1% to 5% of the first thickness. that droplets ejected from the Solution are directed into 77. The device of claim 75, wherein the upper layer is a the antisolvent, wherein the antisolvent is Selected Such molecular monolayer. that the pharmaceutical agent is leSS Soluble therein 78. The device of claim 75, wherein the upper layer is a than in the Solvent; molecular bilayer. 65 an acoustic ejector comprising an acoustic radiation gen 79. The device of claim 50, wherein the pharmaceutical erator for generating acoustic radiation and a focusing agent is Selected from the group consisting of analgesic means for focusing the acoustic radiation at a focal US 6,596.206 B2 21 22 point within the Solution in the reservoir So as to eject 104. The device of claim 103, comprising in the range of a droplet therefrom; and about 100,000 to about 4,000,000 reservoirs. a means for positioning the ejector in acoustic coupling 105. The device of claim 96, wherein each reservoir is relationship to the reservoir. adapted to contain no more than about 100 nanoliters of 97. The device of claim 96, comprising a single acoustic fluid. ejector. 106. The device of claim 96, wherein each reservoir is 98. The device of claim 96, comprising a plurality of adapted to contain no more than about 10 nanoliters of fluid. acoustic ejectors positioned to direct focused acoustic 107. The device of claim 96, further comprising means for energy to each reservoir So as to eject a plurality of droplets maintaining the Solvent in each reservoir at a constant therefrom. temperature. 99. The device of claim 96, wherein each of the reservoirs 108. The device of claim 96, wherein the acoustic cou is removable from the device. pling relationship between the ejector and the fluid in each 100. The device of claim 96, wherein the reservoirs are reservoir is established by providing an acoustically con individual wells in a well plate. ductive medium between the ejector and each reservoir. 101. The device of claim 96, wherein the reservoirs are 15 109. The device of claim 96, wherein acoustic coupling Substantially acoustically indistinguishable. between the ejector and the fluid in each reservoir is estab 102. The device of claim 96, comprising at least about lished at a predetermined distance between the ejector and 10,000 reservoirs. each reservoir. 103. The device of claim 102, comprising at least about 100,000 reservoirs.