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|HAO HA TA MARIAUS009730892B2 TAI MAI MULTE DETALIAN MATURE (12 ) United States Patent ( 10 ) Patent No. : US 9 , 730 ,892 B2 Schutt et al. ( 45 ) Date of Patent : Aug. 15 , 2017

(54 ) METHOD FOR FORMULATING LARGE ( 56 ) References Cited DIAMETER SYNTHETIC MEMBRANE VESICLES U .S . PATENT DOCUMENTS 2 ,044 , 296 A 6 /1936 Hardgrove ( 71 ) Applicant: Pacira Pharmaceuticals , Inc ., San 2 ,824 ,807 A 2 / 1958 Laster et al . Diego , CA (US ) ( Continued ) (72 ) Inventors : Ernest George Schutt , San Diego , CA (US ) ; Ronald Warren McGuire , FOREIGN PATENT DOCUMENTS Murrieta , CA (US ) ; Peter Andrew AU 2002301268 B2 2 /2003 Walters, San Diego , CA (US ) ; AU 2006200044 Al 2 / 2006 Kathleen D . A . Los, San Diego , CA CA 933796 9 / 1973 ?? 960811 1 / 1975 (US ) CA 2 564 120 A1 8 / 1998 CA 2464668 5 / 2003 (73 ) Assignee : Pacira Pharmaceuticals , Inc ., San CH 519936 7 / 1971 Diego , CA (US ) CH 587023 7 / 1974 CN 101396345 5 / 2009 DE 1199191 8 / 1965 ( * ) Notice : Subject to any disclaimer, the term of this DE 51254 10 / 1966 patent is extended or adjusted under 35 DE 56223 5 / 1967 U . S . C . 154 ( b ) by 0 days . ( Continued ) ( 21 ) Appl. No . : 15 / 249 , 195 OTHER PUBLICATIONS Aug . 26 , 2016 (22 ) Filed : Atiemo -Obeng et al. “ Rotor - Stator Mixing Devices ” , Handbook of (65 ) Prior Publication Data Industrial Mixing Science and Practice , 2004 , 479 -505 , Eds. Paul, US 2016 /0361260 A1 Dec. 15 , 2016 et al. , Hoboken , NJ: John Wiley and Sons, Inc. Related U . S. Application Data (Continued ) (63 ) Continuation of application No. 13 / 786 ,378 , filed on Primary Examiner — Monzer R Chorbaji Mar . 5 , 2013 , which is a continuation of application ( 74 ) Attorney , Agent, or Firm — Knobbe Martens Olson No . 13 /083 ,485 , filed on Apr. 8 , 2011. & Bear LLP (Continued ) (57 ) ABSTRACT (51 ) Int. Ci. The present invention generally relates to the field of phar B01D 11 /04 ( 2006 . 01 ) maceutical sciences . More specifically , the present invention BOID 11 /02 ( 2006 .01 ) includes apparatus and devices for the preparation of phar maceutical formulations containing large diameter synthetic ( Continued ) membrane vesicles , such as multivesicular liposomes, meth (52 ) U . S . CI. ods for preparing such formulations , and the use of specific CPC ...... A61K 9 / 1271 ( 2013 . 01 ) ; A61K 9 / 127 formulations for therapeutic treatment of subjects in need ( 2013. 01 ) ; A61K 9 /1277 ( 2013. 01 ) ; thereof. Formation and use of the pharmaceutical formula (Continued ) tions containing large diameter synthetic membrane vesicles (58 ) Field of Classification Search produced by using the apparatus and devices for therapeutic CPC A61L 2 / 022; B01D 39/ 00 ; CO2F 1 / 00 ; A61K treatment of subjects in need thereof is also contemplated . 9 /4833 (Continued ) 14 Claims, 19 Drawing Sheets

100 5 P - 10 22 . - 10 22 1 00 64 N - - 56 - 60 64 . S . - 66 102 65. L 19 12 65 Log-- 12 02 - Logo { 2K13 el * Gas Supply - Gas Supply B 420 * * * 35 A Gas Supply -- 25- - 7 55 105 85 23. - M 34 9,5 85 75 110 97 96 97 96 50 . HB 50 Die , 115 d 115 45 l --- 777 - 125 / 150 150 -- 132 4h 4 T 130 - 70 120 US 9 ,730 , 892 B2 Page 2

Related U . S . Application Data 3 , 738 , 938 A 6 / 1973 Barrett 3 , 741 , 273 A 6 / 1973 Meade 3 ,746 ,657 A 7 / 1973 Miller et al. (60 ) Provisional application No . 61/ 322, 814 , filed on Apr. 3 ,749 ,671 A 7 / 1973 Gedge 9 , 2010 . 3 , 751, 398 A 8 / 1973 Dahl 3 , 756 , 831 A 9 / 1973 Wingerd et al. (51 ) Int. CI. 3 ,759 ,450 A 9 / 1973 Fram et al . A61K 9 / 127 3 , 761, 288 A 9 / 1973 Glicksman et al . ( 2006 . 01 ) 3 ,764 ,346 A 10 / 1973 Noznick et al. BOID 1/ 16 ( 2006 .01 ) 3 ,770 , 207 A 11/ 1973 Muller et al. A61K 31 /4458 ( 2006 .01 ) 3 ,776 ,688 A 12 / 1973 Damgaard - Iversen et al . BOIF 3 / 08 ( 2006 .01 ) 3 , 784 ,648 A 1 / 1974 Bergmeister et al. B01F 5 / 10 ( 2006 .01 ) 3 , 793 , 465 A 2 / 1974 Bohren 3 , 803 , 111 A 4 / 1974 Munro et al. BOIF 7700 ( 2006 .01 ) 3 , 803 , 723 A 4 / 1974 Lamm et al. A61K 9 / 48 ( 2006 .01 ) 3 , 805 , 869 A 4 / 1974 Winter et al. A61M 11/ 00 ( 2006 .01 ) 3 , 821, 434 A 6 / 1974 Houghton - Larsen et al. BOIJ 13 / 04 ( 2006 .01 ) 3 , 830 , 750 A 8 / 1974 Wellman BOIJ 13 / 12 ( 2006 . 01) 3 , 840 , 996 A 10 / 1974 Grindstaff et al . 3 , 843 , 380 A 10 / 1974 Beyn A61K 31/ 445 ( 2006 . 01 ) 3 , 845 , 801 A 11/ 1974 Kuchar BOIF 3 / 04 ( 2006 . 01 ) 3 , 858 , 854 A 1 / 1975 Win et al . BOIF 15 / 06 ( 2006 . 01 ) 3 , 883 , 494 A 5 / 1975 Winter et al. 3 , 885 , 918 A 5 / 1975 Isahaya (52 ) U . S . CI. 3 , 920 , 187 A 11/ 1975 Willis CPC ...... A61K 9 /4833 ( 2013. 01 ) ; A61K 31/ 445 3 , 923 , 248 A 12 / 1975 Creswell ( 2013 .01 ); A61K 31/ 4458 (2013 . 01 ) ; A61M 3 , 923 ,680 A 12 / 1975 Roeder et al. 11/ 00 (2013 .01 ) ; BOID 1 / 16 (2013 .01 ) ; BOIF 3 , 932 ,587 A 1 / 1976 Grantham et al. 3 /04049 ( 2013 .01 ) ; B01F 3 /0807 ( 2013 .01 ) ; 3 , 949 ,094 A 4 / 1976 Johnson et al. 3 , 956 , 009 A 5 / 1976 Lundquist , Jr. et al. B01F 3 /088 ( 2013 .01 ) ; B01F 5 / 104 ( 2013 .01 ) ; 3 , 956 ,521 A 5 / 1976 Pisecky et al. 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50 25HzProcess 30HzProcess met35HzProcess mmBatchLotA *BatchLotB 40 Hours FIG.12

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100

70 LowOsmolality-NoHeatTreatment HeatTreated30sec. 60 -NoHeatTreatment 50 Hours FIG.15 mug 40

0 8 8 8 8 8 g o )mL /ng ( Bupivacaine US 9 , 730 ,892 B2

METHOD FOR FORMULATING LARGE ments , a diameter of the fluid contacting chamber is larger DIAMETER SYNTHETIC MEMBRANE than a diameter of the first fluid conduit. In some embodi VESICLES ments , the fluid contacting chamber conically narrows in diameter from the top of the fluid contacting chamber to the INCORPORATION BY REFERENCE TO ANY 5 exit orfice of the fluid contacting chamber . In some embodi PRIORITY APPLICATIONS ments , the fluid contacting chamber conically narrows in diameter from a point below the top of the fluid contacting Any and all priority claims identified in the Application chamber to the exit orfice of the fluid contacting chamber. Data Sheet, or any correction thereto , are hereby incorpo Some embodiments provide a process for preparing drop rated by reference under 37 CFR 1 . 57 . 10 lets using an atomizing nozzle as disclosed herein compris ing applying a first liquid to the first fluid conduit , applying FIELD OF THE INVENTION a second liquid to the second fluid conduit , applying a gas to the third fluid conduit , wherein the gas exiting the third The present invention generally relates to the field of fluid conduit exit orifice impinges the liquid exiting the at pharmaceutical sciences. More specifically , the present 15 least one exit orifice of the fluid contacting chamber, pro invention relates to pharmaceutical formulations containing viding atomized droplets , wherein the droplets have an large diameter synthetic membrane vesicles , such as multi average diameter from about 100 nm to about 300 uM . In vesicular liposomes (MVL ) , methods for preparing such some embodiments , the first liquid is an emulsion comprised formulations, and the use of specific formulations for thera of a first aqueous phase , and a first organic phase comprising peutic treatment of subjects in need thereof. 20 a first organic solvent. In some embodiments , the first organic solvent is chloroform or methylene chloride . In BACKGROUND some embodiments , the first organic phase further comprises at least one amphipathic lipid and at least one neutral lipid . The following includes information that may be useful in In some embodiments , the at least one amphipathic lipid is understanding the present embodiments . It is not an admis - 25 selected from the group consisting of phosphatidylcholines , sion that any of the information provided herein is prior art, phosphatidylserines, phosphatidylethanolamines , phospha or relevant, to the presently described or claimed embodi - tidylinositols , sphingomyelin , soybean lecithin (soya leci ments , or that any publication or document that is specifi- thin ) , egg lecithin , lysophosphatidylcholines , lysophospha cally or implicitly referenced is prior art . tidylethanolamines , phosphatidylglycerols , Large scale methods of manufacturing large diameter 30 phosphatidylserines, phosphatidylinositols , phosphatidic synthetic membrane vesicles, such as multivesicular lipo - acids, cardiolipins , acyl trimethylammonium propane , dia somes , often require large amounts of solvents , time sensi cyldimethylammonium propane , stearylamine, and ethyl tive steps and concentration adjustment of the final product phosphatidylcholine . In some embodiments , the at least one under sterile conditions. In addition , current methods of amphipathic lipid is selected from the group consisting of manufacturing large diameter synthetic membrane vesicles , 35 1 , 2 - dioleoyl- sn - glycero - 3 -phosphocholine , 1 , 2 - dilauroyl such as multivesicular liposomes on a commercial scale , sn - glycero - 3 -phosphocholine , 1 , 2 -dimyristoyl - sn - glycero require significant commitments in manufacturing space , 3 -phosphocholine , 1 , 2 -dipalmitoyl - sn - glycero - 3 -phospho cost, and time . As such , developing stable multivesicular choline , 1 , 2 -di stearoyl- sn - glycero - 3 -phosphocholine , 1 , 2 liposome formulations containing a therapeutic agent in a diarachidoyl - sn - glycero - 3 -phosphocholine , 1 , 2 - dibehenoyl cost effective and timely manner remains an ongoing chal- 40 sn - glycero - 3 -phosphocholine , 1 , 2 - dipalmitoleoyl- sn lenge. glycero - 3 -phosphocholine , 1, 2 -dieicosenoyl - sn glycero - 3 - phosphocholine , 1 , 2 - dierucoyl- sn - glycero - 3 SUMMARY phosphocholine , 1 , 2 - dipalmitoyl- sn - glycero - 3 -phospho glycerol, 1 , 2 -dioleoyl - sn - glycero - 3 -phosphoglycerol . In Some embodiments provide an atomizing nozzle appara - 45 some embodiments , the at least one neutral lipid is selected tus , comprising a first fluid conduit and a second fluid from the group consisting of glycerol esters , glycol esters , conduit each having at least one entrance orifice and at least tocopherol esters , sterol esters , hydrocarbons and squalenes. one exit orifice, a fluid contacting chamber having a top In some embodiments , the at least one neutral lipid is comprising at least one entrance orifice and having a bottom selected from the group consisting of triolein , tripalmitolein , comprising at least one exit orifice and connecting to the at 50 trimyristolein , trilinolein , tributyrin , tricaprylin , tricaproin , least one exit orifice of the first fluid conduit , a third liquid and tricaprin . In some embodiments , the first aqueous phase channel, wherein the third fluid conduit annularly surrounds further comprises a therapeutic agent. In some embodi a portion of the fluid contacting chamber . In some embodi m ents , the therapeutic agent is bupivacaine . In some ments, the fluid contacting chamber connects to the at least embodiments , the second liquid applied to the second fluid one exit orifice of the second fluid conduit. In some embodi- 55 conduit is a second aqueous phase . In some embodiments , ments, the at least one exit orifice of the fluid contacting the droplet comprises a first component core and a second chamber and the at least one exit orifice of the third fluid aqueous phase shell. In some embodiments , the gas is conduit are flush . In some embodiments , the at least one exit nitrogen . In some embodiments, the droplets have an aver orifice of the fluid contacting chamber is recessed within the age diameter from about 20 uM to about 60 UM . In some at least one exit orifice of the third fluid conduit . In some 60 embodiments , the droplets have an average diameter from embodiments , the at least one exit orifice of the fluid about 35 uM to about 45 uM . contacting chamber extends beyond the at least one exit S ome embodiments provide atomized droplet comprising orifice of the third fluid conduit. In some embodiments , the an emulsion core , wherein the emulsion core comprises i ) a first fluid conduit and the second fluid conduit are co -axial first aqueous phase ; and ii ) a first organic phase comprising for a first portion of the first fluid conduit length . In some 65 a first organic solvent; and an aqueous phase shell , wherein embodiments , the second fluid conduit annularly surrounds said atomized droplet is made by a process comprising a second portion of the first fluid conduit . In some embodi- combining a first component, an aqueous phase , and a gas US 9 ,730 ,892 B2 using an atomizing nozzle as disclosed and described here - In some embodiments , the first organic phase further com rin , said process comprising applying a first component to prises at least one amphipathic lipid and at least one neutral the first fluid conduit , applying an aqueous phase to the lipid . second fluid conduit , and applying a gas to the third fluid Some embodiments provide an evaporation apparatus, conduit , wherein the gas exiting the third fluid conduit exit 5 comprising at least one atomizing nozzle apparatus of Claim orifice impinges the liquid exiting the at least one exit orifice 1 and means for evaporating an organic solvent. of the fluid contacting chamber, providing atomized drop Some embodiments provide an evaporation apparatus , lets, wherein the droplets have an average diameter from comprising a solvent removal vessel having a top , a bottom about 100 nm to about 300 uM . and a circular wall , at least one atomizing nozzle connected Some embodiments provide atomizing nozzle apparatus , 10 to the circular wall , a carrier gas entrance orifice connected comprising an first fluid conduit , a second fluid conduit and to the circular wall , a solvent removal gas exit orifice a third fluid conduit each having at least one entrance orifice centrally connected to the top , and a product exit orifice and at least one exit orifice , a first fluid contacting chamber connected to the bottom of the vessel. In some embodi having a top comprising at least one entrance orifice and ments, at least part of the solvent removal vessel is jacketed . having a bottom comprising at least one exit orifice and 15 In some embodiments , the atomizing nozzle is mounted to connecting to the at least one exit orifice of the first fluid and extending through the top of the solvent removal vessel. conduit , wherein the second fluid conduit annularly sur - In some embodiments , the top of the solvent removal vessel rounds a portion of the first fluid contacting chamber, a comprises a lid . In some embodiments , the apparatus further second fluid contacting chamber having a top comprising at comprises a rinse nozzle mounted to and extending through least one entrance orifice and having a bottom comprising at 20 the top of the solvent removal vessel . In some embodiments , least one exit orifice and connecting to the at least one exit the circular wall has a central axis and the solvent removal orifice of the first fluid contacting chamber , wherein the third gas exit orifice further comprises a tube extending into the fluid conduit annularly surrounds a portion of the second solvent removal vessel residing along the central axis . In fluid contacting chamber, and a fourth fluid conduit , wherein some embodiments , the atomizing nozzle is angled at least the fourth fluid conduit annularly surrounds a portion of the 25 5 degrees measured off the central axis of the wall and in a second fluid contacting chamber . In some embodiments , the plane parallel to the wall nearest to it . In some embodiments , first fluid contacting chamber connects to the at least one exit the carrier gas entrance orifice is combined with the atom orifice of the second fluid conduit . In some embodiments , izing nozzle . In some embodiments , the solvent removal gas the at least one exit orifice of the second fluid contacting exit orifice further comprises a tube extending into the chamber and the at least one exit orifice of the fourth fluid 30 solvent removal vessel , wherein the tube is fitted with a conduit are flush . In some embodiments , the at least one exit narrowing cone and an annular ring . In some embodiments , orifice of the second fluid contacting chamber is recessed the tube extends from about 1/ 3 to about 45 of the way into within the at least one exit orifice of the fourth fluid conduit . the solvent removal vessel . In some embodiments , the tube In some embodiments , the at least one exit orifice of the extends about 2 / 3 of the way into the solvent removal vessel . second fluid contacting chamber extends beyond the at least 35 In some embodiments , the bottom tip of the narrowing cone one exit orifice of the fourth fluid conduit . In some embodi of the solvent removal gas exit orifice diameter is from about ments , the first fluid conduit and the second fluid conduit are 1/ 1000 to about 1 /5 of a diameter of the inside of the solvent co - axial for a first portion of the first fluid conduits length . removal vessel . In some embodiments , the solvent removal In some embodiments , the second fluid conduit annularly gas exit orifice diameter is less than 1/ 10 of a diameter of the surrounds a second portion of the first fluid conduit . In some 40 inside of the solvent removal vessel. In some embodiments , embodiments , the first fluid contacting chamber conically the at least one atomizing nozzle is an atomizing nozzle narrows in diameter from the top of the fluid contacting apparatus as disclosed and described herein . In some chamber to the exit orfice of the fluid contacting chamber. In embodiments , the ratio of the inside diameter of the solvent some embodiments , the second fluid conduit and the third removal vessel to the diameter of the bottom tip of the fluid conduit are co - axial for a first portion of the second 45 narrowing cone of the solvent removal gas exit orifice is fluid conduits length . In some embodiments , the third fluid between approximately 5 : 1 and 100 : 1 . In some embodi conduit annularly surrounds a second portion of the second ments , the ratio of the inside diameter of the solvent removal fluid conduit . In some embodiments , the first fluid contact - vessel to the diameter of the bottom tip of the narrowing ing chamber conically narrows in diameter from the top of cone of the solvent removal gas exit orifice is between the fluid contacting chamber to the exit orfice of the fluid 50 approximately 20 : 1 and 60 : 1 . contacting chamber . Some embodiments provide a process for preparing large Some embodiments provide a process for preparing drop - diameter synthetic membrane vesicles using an evaporation lets using an atomizing nozzle apparatus as disclosed and apparatus as disclosed and described herein , comprising described herein , comprising applying a first liquid to the introducing large diameter synthetic membrane vesicles first fluid conduit , applying a second liquid to the second 55 pre -droplets to the solvent removal vessel, wherein the large fluid conduit , applying a third liquid to the third fluid diameter synthetic membrane vesicles pre- droplets comprise conduit, applying a gas to the fourth fluid conduit , wherein a first component core and an aqueous phase shell , applying the gas exiting the fourth fluid conduit exit orifice impinges a carrier gas in a tangential direction to the circular wall the liquid exiting the at least one exit orifice of the second through the carrier gas entrance orifice , and removing a fluid contacting chamber, providing atomized droplets , 60 solvent removal gas through the solvent removal gas exit wherein the droplets have an average diameter from about orifice to provide the large diameter synthetic membrane 100 nm to about 300 uM . In some embodiments , the first vesicles. In some embodiments , the first component core liquid is an emulsion comprised of i ) a first aqueous phase , comprises a first aqueous phase and a first organic phase . In and ii ) a first organic phase comprising a first organic some embodiments, the first organic phase comprises a solvent, the second liquid is a second aqueous phase , and the 65 continuous first organic solvent. In some embodiments , the third liquid is a second organic phase . In some embodiments , first organic solvent is chloroform or methylene chloride . In the first organic solvent is chloroform or methylene chloride . some embodiments , the first organic phase further comprises US 9 , 730 , 892 B2 at least one amphipathic lipid and at least one neutral lipid . ment large diameter synthetic membrane vesicles to an In some embodiments , the first component core is first outlet line ; contacting the pre - temperature treatment large aqueous phase droplets as a suspension in a first organic diameter synthetic membrane vesicles with a hot solution in phase . In some embodiments , the first aqueous phase drop the outlet line , wherein the hot solution has a temperature lets have an average diameter of from about 10 nm to about 5 ranging from about 30° C . to about 100° C . to provide 10 um , about 100 nm to about 5 um , or about 500 nm to post - temperature treatment large diameter synthetic mem about 2 um . In some embodiments , the first aqueous phase brane vesicles ; transferring the post - temperature treatment droplets have an average diameter of about 1 um . In some large diameter synthetic membrane vesicles to a continuous embodiments , the carrier gas comprises nitrogen . In some flow particle -concentration system or continuous phase embodiments , the carrier gas comprises nitrogen and water 10 exchange system ; cooling the post - temperature treatment vapor. In some embodiments, the solvent removal gas com large diameter synthetic membrane vesicles to a second prises nitrogen and organic solvent. In some embodiments , temperature to provide the large diameter synthetic mem the carrier gas and the solvent removal gas travel in a vortex brane vesicles; and isolating the large diameter synthetic largein the diameter solvent syntheticremoval membranevessel. In somevesicles embodiments are multivesicu , the - 15 membrane vesicles . In some embodiments , the first compo lar liposomes having a structure including multiple non nent core comprises a first aqueous phase and a first organic concentric chambers and comprising at least one amphip - phase . In some embodiments , the first organic phase com athic lipid and at least one neutral lipid . In some prises a continuous first organic solvent. In some embodi embodiments , the at least one amphipathic lipid is selected ments , the first organic solvent is chloroform or methylene from the group consisting of phosphatidylcholines, phos - 20 chloride . In some embodiments , the first organic phase phatidylserines , phosphatidylethanolamines, phosphati further comprises at least one amphipathic lipid and at least dylinositols , sphingomyelin , soybean lecithin ( soya leci one neutral lipid . In some embodiments , the first component thin ), egg lecithin , lysophosphatidylcholines, core is a suspension of first aqueous phase droplets in a first lysophosphatidylethanolamines , phosphatidylglycerols , organic phase . In some embodiments , the first aqueous phosphatidylserines , phosphatidylinositols , phosphatidic 25 phase droplets have an average diameter of from about 10 acids , cardiolipins, acyl trimethylammonium propane, dia - nm to about 10 um , about 100 nm to about 5 um , or about cyldimethylammonium propane , stearylamine , and ethyl 500 nm to about 2 um . In some embodiments , the first phosphatidylcholine . In some embodiments , the at least one aqueous phase droplets have an average diameter of about 1 amphipathic lipid is selected from the group consisting of u m . In some embodiments , the carrier gas comprises nitro 1, 2 -dioleoyl - sn - glycero - 3 - phosphocholine , 1 , 2 - dilauroyl- 30 gen . In some embodiments , the solvent removal gas com sn - glycero - 3 - phosphocholine , 1 , 2 -dimyristoyl - sn - glycero - prises nitrogen and organic solvent. In some embodiments , 3 -phosphocholine , 1 , 2 -dipalmitoyl -sn - glycero - 3 -phospho - the carrier gas and the solvent removal gas travel in a vortex choline, 1 , 2 -distearoyl - sn -glycero - 3 -phosphocholine , 1 , 2 in the solvent removal vessel . In some embodiments , the diarachidoyl- sn - glycero -3 -phosphocholine , 1, 2 -dibehenoyl - large diameter synthetic membrane vesicles are multivesicu sn - glycero - 3 -phosphocholine , 1 , 2 - dipalmitoleoyl- sn - 35 lar liposomes having a structure including multiple non glycero - 3 -phosphocholine , 1, 2 -dieicosenoyl - sn - concentric chambers and comprising at least one amphip glycero - 3 - phosphocholine , 1 , 2 - dierucoyl - sn - glycero - 3 - athic lipid and at least one neutral lipid . In some phosphocholine , 1 , 2 - dipalmitoyl- sn - glycero - 3 - phospho - embodiments , the at least one amphipathic lipid is selected glycerol, and 1 , 2 -dioleoyl - sn - glycero - 3 - phosphoglycerol. In from the group consisting of phosphatidylcholines, phos some embodiments , the at least one neutral lipid is selected 40 phatidylserines , phosphatidylethanolamines, phosphati from the group consisting of glycerol esters, glycol esters , dylinositols , sphingomyelin , soybean lecithin ( soya leci tocopherol esters , sterol esters , hydrocarbons and squalenes. thin ) , egg lecithin , lysophosphatidylcholines, In some embodiments , the at least one neutral lipid is lysophosphatidylethanolamines , phosphatidylglycerols , selected from the group consisting of triolein , tripalmitolein , phosphatidylserines, phosphatidylinositols , phosphatidic trimyristolein , trilinolein , tributyrin , tricaprylin , tricaproin , 45 acids , cardiolipins, acyl trimethylammonium propane , dia and tricaprin . In some embodiments , the multivesicular cyldimethylammonium propane, stearylamine , and ethyl liposomes further comprises a therapeutic agent. In some phosphatidylcholine . In some embodiments , the at least one embodiments , the therapeutic agent is bupivacaine . In some amphipathic lipid is selected from the group consisting of embodiments , the multivesicular liposomes comprises an 1 , 2 - dioleoyl- sn - glycero - 3 -phosphocholine , 1 , 2 - dilauroyl outer surface layer whose composition is different than the 50 sn - glycero - 3 - phosphocholine, 1 , 2 - dimyristoyl- sn - glycero composition of the internal structure . 3 -phosphocholine , 1 , 2 - dipalmitoyl- sn - glycero - 3 - phospho Some embodiments provide an evaporation apparatus, choline , 1, 2 - distearoyl- sn - glycero - 3 -phosphocholine , 1, 2 comprising at least one atomizing nozzle apparatus as dis diarachidoyl- sn - glycero - 3 -phosphocholine , 1 , 2 -dibehenoyl closed and described herein and means for removing an sn - glycero - 3 -phosphocholine , 1 , 2 - dipalmitoleoyl- sn organic solvent from a droplet. 55 glycero - 3 - phosphocholine , 1 , 2 - dieicosenoyl- sn Some embodiments provide a process for preparing large glycero - 3 - phosphocholine, 1, 2 - dierucoyl- sn - glycero - 3 diameter synthetic membrane vesicles using the evaporation phosphocholine, 1 , 2 -dipalmitoyl - sn -glycero - 3 -phospho apparatus as disclosed and described herein , comprising glycerol, and 1 , 2 - dioleoyl- sn - glycero - 3 - phosphoglycerol. In introducing large diameter synthetic membrane vesicles some embodiments, the at least one neutral lipid is selected pre -droplets to the solvent removal vessel, wherein the large 60 from the group consisting of glycerol esters, glycol esters, diameter synthetic membrane vesicles pre - droplets comprise tocopherol esters , sterol esters , hydrocarbons and squalenes . a first component core and an aqueous phase shell ; applying In some embodiments , the at least one neutral lipid is a carrier gas in a tangental direction to the circular wall selected from the group consisting of triolein , tripalmitolein , through the carrier gas entrance orifice ; removing a solvent trimyristolein , trilinolein , tributyrin , tricaprylin , tricaproin , removal gas through the solvent removal gas exit orifice to 65 and tricaprin . In some embodiments , the multivesicular provide pre - temperature treatment large diameter synthetic liposomes further comprises a therapeutic agent. In some membrane vesicles; introducing the pre- temperature treat embodiments , the therapeutic agent is bupivacaine . US 9 , 730 , 892 B2 Some embodiments provide a composition comprising rotation axis of the rotor and is in fluid communication with multivesicular liposomes having a structure including mul - the rotor. In some embodiments , a continuous phase tiple non - concentric chambers and comprising at least one entrance line is connected to the recirculation loop upstream amphipathic lipid and at least one neutral lipid , wherein said of the mixer and downstream of the heat exchanger and an multivesicular liposomes are made by a process comprising 5 outlet line is connected to the recirculation loop downstream removing organic solvent from multivesicular liposomes of the mixer and upstream of the heat exchanger. In some pre -droplets using an evaporation apparatus as disclosed and embodiments , the continuous - flow emulsification system described herein , said process comprising introducing mul - further comprises an emulsion , which emulsion recirculates tivesicular liposomes pre - droplets pre - droplets to the solvent through the recirculation line back to the mixer an average removal vessel , wherein the large diameter synthetic mem - 10 of at least 5 or more times . In some embodiments, the brane vesicles pre - droplets comprise a first component core continuous - flow emulsification system further comprises and an aqueous phase shell ; applying a carrier gas in a emulsion droplets produced in the mixer , which are on tangental direction to the circular wall through the carrier average less than 10 microns in diameter. gas entrance orifice ; and removing a solvent removal gas Some embodiments provide a continuous processing sys through the solvent removal gas exit orifice to provide the 15 tem , comprising one or more concentrator units , each unit large diameter synthetic membrane vesicles . comprising a retentate vessel ; a particle suspension inlet Some embodiments provide a composition comprising line , connected to the retentate vessel; a first outlet line , large diameter synthetic membrane vesicles made by a connecting the retentate vessel and a particle concentrating process disclosed and described herein . In some embodi device ; a pump located along the first outlet line between the ments, the large diameter synthetic membrane vesicles are 20 retentate vessel and the particle concentrating device ; and a multivesicular liposomes having a structure including mul- second outlet line, leading to another concentrator unit or the tiple non -concentric chambers and comprising at least one final product collection vessel; and means for removing or amphipathic lipid and at least one neutral lipid . In some exchanging solvent. In some embodiments, the means for embodiments , the at least one amphipathic lipid is selected removing or exchanging solvent are each independently from the group consisting of phosphatidylcholines , phos - 25 selected from the group consisting of a tangential flow phatidylserines , phosphatidylethanolamines , phosphati - filtration unit, a hydro -cyclone unit , and a centrifugal sepa dylinositols , sphingomyelin , soybean lecithin ( soya leci - rator. In some embodiments , the continuous processing thin ) , egg l ecithin , lysophosphatidylcholines, system further comprises a new suspending medium inlet lysophosphatidylethanolamines , phosphatidylglycerols , line connected to the retentate vessel. In some embodiments , phosphatidylserines , phosphatidylinositols, phosphatidic 30 the means for removing or exchanging solvent are each a acids , cardiolipins, acyl trimethylammonium propane, dia tangential flow filtration unit . In some embodiments , the cyldimethylammonium propane , stearylamine , and ethyl means for removing or exchanging solvent are each a phosphatidylcholine . In some embodiments, the at least one centrifugal separator. In some embodiments , the system amphipathic lipid is selected from the group consisting of comprises at least one tangential flow filtration unit and at 1 , 2 -dioleoyl - sn - glycero - 3 -phosphocholine , 1 , 2 - dilauroyl- 35 least one centrifugal separator. In some embodiments, the sn - glycero - 3 - phosphocholine , 1 , 2 -dimyristoyl - sn - glycero - continuous processing system is a continuous - flow particle 3 - phosphocholine , 1, 2 -dipalmitoyl - sn - glycero - 3 -phospho concentration system or continuous phase exchange system . choline , 1 , 2 - distearoyl- sn - glycero - 3 - phosphocholine, 1 , 2 Some embodiments provide a process for making multi diarachidoyl- sn - glycero - 3 -phosphocholine , 1 , 2 -dibehenoyl vesicular liposomes using the atomizing nozzle apparatus as sn - glycero - 3 - phosphocholine , 1 , 2 - dipalmitoleoyl- sn - 40 disclosed and described herein , comprising applying a first glycero - 3 -phosphocholine , 1 , 2 - dieicosenoyl- sn - glycero - 3 - liquid to the first fluid conduit , wherein the first liquid phosphocholine , 1 , 2 -dierucoyl - sn - glycero - 3 - comprises an organic solvent, applying a second liquid to the phosphocholine , 1, 2 - dipalmitoyl- sn - glycero -3 second fluid conduit , applying a pressurized gas to the third phosphoglycerol, and 1 , 2 - dioleoyl- sn - glycero - 3 - fluid conduit to provide atomized droplets , wherein the phosphoglycerol. In some embodiments , the at least one 45 pressurized gas exiting the third fluid conduit exit orifice neutral lipid is selected from the group consisting of glycerol impinges the liquid exiting the fluid contacting chamber exit esters , glycol esters , tocopherol esters , sterol esters , hydro - orifice , and removing the organic solvent from the atomized carbons and squalenes . In some embodiments , the at least droplets , wherein less than 4000 ppm of the organic solvent one neutral lipid is selected from the group consisting of remains in the atomized droplets . In some embodiments , the triolein , tripalmitolein , trimyristolein , trilinolein , tributyrin , 50 first liquid is an emulsion comprised of a discontinuous tricaprylin , tricaproin , and tricaprin . In some embodiments, aqueous phase , and a continuous organic phase comprising the multivesicular liposomes further comprises a therapeutic the organic solvent. In some embodiments , the organic agent. In some embodiments , the therapeutic agent is bupi - solvent is methylene chloride . In some embodiments , the vacaine . continuous organic phase further comprises a therapeutic Some embodiments provide a continuous - flow emulsifi - 55 agent. In some embodiments , the therapeutic agent is bupi cation system , comprising a mixer , comprised of a rotor and vacaine or a salt thereof. In some embodiments, the second a stator ; a recirculation loop , comprised of one or more liquid applied to the second fluid conduit is an aqueous recirculation lines ; a heat exchanger ; one or more outlet solution . In some embodiments , the aqueous solution further lines; one or more continuous phase inlet lines ; and a comprises dextrose and lysine . In some embodiments , the discontinuous phase inlet line , wherein the heat exchanger 60 gas is a sterilized gas . In some embodiments, the gas is and the mixer are connected together in the recirculation nitrogen . In some embodiments, the process further com loop by one or more recirculation lines ; further wherein the prises introducing atomized droplets to an evaporation appa one or more outlet lines and one or more continuous phase ratus as disclosed and described herein ; introducing a pres inlet lines are connected to the recirculation loop ; further surized carrier gas tangentially to the circular wall into the wherein the end of the discontinuous phase inlet line is 65 solvent removal vessel through the carrier gas entrance located within approximately 1 /3rd of a rotor diameter from orifice ; removing a solvent removal gas wherein the solvent the rotor and approximately 1/ 3rd of a rotor diameter of the removal gas removes greater than 90 % of the organic US 9 ,730 ,892 B2 10 solvent in the atomized droplets resulting in formation of BRIEF DESCRIPTION OF THE DRAWINGS multivesicular liposomes . In some embodiments , the carrier gas is heated and humidified . In some embodiments , the These and other objects and features of the instant process further comprises spraying a wall rinse solution into embodiment will become more fully apparent in the follow the solvent removal vessel using a rinse nozzle , wherein the 5 ing descriptions and appended claims taken in conjunction wall rinse solution prevents build -up of particles in the with the following drawings where like references numbers evaporation apparatus . In some embodiments , the atomized indicate identical or functionally similar elements . droplets contain organic solvent in the range of from about FIG . 1A is a schematic diagram of significant components 400 ppm to about 3500 ppm . used in one of the instant systems for manufacturing syn Some embodiments provide a process for making an thetic membrane vesicles . emulsion using an emulsification system as disclosed and FIG . 1B is a schematic diagram of significant components described herein , comprising feeding an organic discontinu - used in another system for manufacturing synthetic mem ous phase into the emulsification system through the dis - brane vesicles. continuous phase inlet line and feeding an aqueous continu - 1 FIG . 1C is a schematic of one embodiment of a continu ous phase into the emulsification system through one or ous heat treatment system including a temperature con more continuous phase inlet lines. trolled tank and a holding coil tubing used in a manufac Some embodiments provide a process for making an turing system such as the manufacturing system of FIG . 1A emulsion using the emulsification system as disclosed and or FIG . 1B . described herein , comprising feeding an aqueous discon - 20 FIG . 2 is a schematic of one embodiment of an emulsi tinuous phase into the emulsification system through the fication system used in themanufacturing system of FIG . 1A discontinuous phase inlet line and feeding an organic con - or FIG . 1B . tinuous phase fed into the emulsification system through the FIG . 3A is a schematic view ofone embodiment of a three one or more continuous phase inlet lines. In some embodi- channel atomizing nozzle used in a manufacturing system , ments , the organic continuous phase is comprised of an 25 such as the manufacturing system of FIG . 1A of FIG . 1B . organic solvent and a neutral lipid . In some embodiments , FIG . 3B is a cross - sectional view of the atomizing nozzle the organic solvent is methylene chloride . In some embodi- of FIG . 3A , taken along line B of FIG . 3A . ments, the aqueous discontinuous phase is comprised of an FIG . 3C is an expanded view of the cylindrical tip of the acid and a therapeutic agent. In some embodiments, the acid is phosphoric acid . In some embodiments , the therapeutic 30 atomizing31 nozzle of FIG . 3A , shown as position C in FIG . agent is bupivacaine . In some embodiments , a portion of the emulsion is fed through one or more outlet lines to the inner FIG . 3D is an expanded view of an atomized droplet fluid conduit of an atomizing nozzle as disclosed and produced by the atomizing nozzle of FIG . 3A , shown as described herein . In some embodiments , a portion of the position D in FIG . 3A . emulsion is fed through one or more outlet lines to an 35 FIG . 3E is a schematic view of one embodiment of a four evaporation apparatus as disclosed and described herein . channel atomizing nozzle used in a manufacturing system , Some embodiments provide a process for preparing large such as the manufacturing system of FIG . 1A or FIG . 1B . diameter synthetic membrane vesicles using an evaporation FIG . 3F is a cross - sectional view of the atomizing nozzle apparatus as disclosed and described herein . of FIG . 3E , taken along line F of FIG . 3E . Some embodiments provide a large diameter synthetic 40 FIG . 3G is an expanded view of the cylindrical tip of the membrane vesicles made by a process as disclosed and atomizing nozzle of FIG . 3E , shown as position G in FIG . described herein . 3E . Some embodiments produce a plurality ofMVL particles FIG . 3H is an expanded view of an atomized droplet made by using the process utilizing the atomizing nozzle produced by the atomizing nozzle of FIG . 3E , shown as disclosed herein . 45 position H in FIG . 3E . Some embodiments produce a plurality of MVL particles FIG . 31 is a schematic view of another embodiment of a made by using the process utilizing the emulsification sys four channel atomizing nozzle used in a manufacturing tem and the atomizing nozzle disclosed herein . system , such as the manufacturing system of FIG . 1A or Some embodiments produce a plurality of MVL particles FIG . 1B . made by using the process utilizing the evaporation appa - 50 FIG . 3J is a cross - sectional view of the atomizing nozzle ratus and the atomizing nozzle disclosed herein . of FIG . 31, taken along line J of FIG . 31. Some embodiments produce a plurality of MVL particles FIG . 3K is an expanded view of the cylindrical tip of the made by using the process utilizing the emulsification sys - atomizing nozzle of FIG . 31, shown as position K in FIG . 31. tem , the evaporation apparatus and the atomizing nozzle FIG . 3L is an expanded view of an atomized droplet disclosed herein . 55 produced by the atomizing nozzle of FIG . 31, shown as Some embodiments produce a plurality ofMVL particles position L in FIG . 31 . made by using the process utilizing the evaporation appa FIG . 4A is a schematic view of one embodiment of an ratus, the atomizing nozzle , and particle concentration sys - atomizing nozzle used in a manufacturing system , such as tem disclosed herein . the manufacturing system of FIG . 1A or FIG . 1B . Some embodiments produce a plurality of MVL particles 60 FIG . 4B is a cross -sectional view of the atomizing nozzle made by using the process utilizing the emulsification sys - of FIG . 4A , taken along line B of FIG . 4A . tem , the evaporation apparatus , the atomizing nozzle , and FIG . 4C is an expanded view of the cylindrical tip of the particle concentration system disclosed herein . atomizing nozzle of FIG . 4A , shown as position C in FIG . Some embodiments produce a plurality of MVL particles 4A . made by any of the above product- by - process embodiments , 65 FIG . 4D is an expanded view of an atomized droplet wherein the MVL contains bupivacaine or a pharmaceuti - produced by the atomizing nozzle of FIG . 4A , shown as cally acceptable salt thereof. position D in FIG . 4A . US 9 , 730 , 892 B2 FIG . 5 is a detailed view of one embodiment of an place (CIP ) and sterile - in - place (SIP ) protocols . Allowing atomizing nozzle used in a manufacturing system , such as for greatly reduced utility requirements to implement CIP the manufacturing system of FIG . 1A or FIG . 1B . and SIP protocols . FIG . 6 is an exploded view of the individual components The devices described in FIGS . 1A and 1B are particularly of the atomizing nozzle of FIG . 5 . 5 well suited for making multivesicular liposomes (MVL ) . FIG . 7 is a schematic of one embodiment of a solvent Multivesicular liposomes (MVL ), first reported by Kim , et removal vessel used for evaporating solvent from atomized al. (Biochim , Biophys. Acta , 728 :339 - 348 , 1983 ), are particles in a manufacturing system such as the manufac uniquely different from other lipid -based drug delivery turing system of FIG . 1A or FIG . 1B . systems such as unilamellar (Huang , Biochemistry , 8 : 334 FIG . 8 is a schematic of one embodiment of a particle 10 352 , 1969; Kim , et al. , Biochim . Biophys. Acta , 646 : 1 - 10 , concentration system used in a manufacturing system such 1981 ) and multilamellar (Bangham , et al. , J Mol. Bio . , as the manufacturing system of FIG . 1A or FIG . 1B . 13 : 238 - 252 , 1965 ) liposomes. For example , multivesicular FIG . 9A and FIG . 9B provide cross -sectional views of a liposomes made by the processes described herein typically droplet, a more detailed view of a droplet , and a large can have diameters ranging from about 10 to 100 um , and diameter synthetic membrane vesicle particle . In some 15 more typically ranging from about 20 to 55 um . In contrast , embodiments , the large diameter synthetic membrane multilamellar liposomes usually have diameters of from 0 . 2 vesicle particle can be a MVL particle . The large diameter to 5 um and unilamellar liposomes usually have diameters of synthetic membrane vesicle particle can be formed by from 0 .02 to 0 . 5 um . removal of the organic solvent from the emulsion droplet. Additionally , multivesicular liposomes (MVL ) contain FIG . 10 is a schematic of one embodiment of a particle 20 multiple aqueous chambers per particle and the multiple concentration system including a plurality of filtration units aqueous chambers are non -concentric . In contrast unilamel and at least one centrifuge unit used in a manufacturing lar liposomes ( also known as unilamellar vesicles ) and system such as the manufacturing system of FIG . 1A or FIG . multilamellar liposomes ( also known as multilamellar 1B . vesicles ) contain a single chamber per particle . Further , FIG . 11 is a schematic of one embodiment of a particle 25 neutral lipids are necessary to form multivesicular liposomes concentration system including a plurality of centrifuge (MVL ) . In contrast unilamellar liposomes ( also known as units used in a manufacturing system such as the manufac unilamellar vesicles ) and multilamellar liposomes (also turing system of FIG . 1A or FIG . 1B . known as multilamellar vesicles) do not require inclusion of FIG . 12 is a graph of an in vitro release study of neutral lipids to form . multivesicular liposome formulations . 30 Multivesicular liposomes (MVL ) are entirely distinct FIG . 13 is a graph of an in vivo PK study of multivesicular from unilamellar liposomes and multilamellar liposomes . liposome formulations. The structural and functional characteristics of multivesicu FIG . 14 is a graph of an accelerated stability profile study l ar liposomes are not directly predictable from current of a multivesicular formulation made with the instant manu knowledge of unilamellar liposomes and multilamellar lipo facturing system with and without heat treatment. The low 35 somes . As described in the book edited by Jean R . Philippot osmolality is more stable than the sample without heat and Francis Schuber (Liposomes as Tools in Basic Research treatment and the heat treated sample is most stable . and Industry , CRC press , Boca Raton , Fla ., 1995 , page 19 ) , FIG . 15 is a graph of an in - vivo release profile study of a Multivesicular liposomes (MVL ) are bounded by an external multivesicular formulation made with and without heat bilayer membrane shell, but have a very distinctive internal treatment. 40 morphology , which may arise as a result of the special Large scale methods of manufacturing large diameter method employed in the manufacture . Topologically , mul synthetic membrane vesicles, such as multivesicular lipo - tivesicular liposomes (MVL ) are defined as liposomes con somes, often require large amounts of solvents , time sensi- taining multiple non -concentric chambers within each lipo tive steps and concentration adjustment of the final product some particle , resembling a “ foam - like " matrix ; whereas under sterile conditions , for example , as described in WO99 / 45 multilamellar vesicles contain multiple concentric chambers 25319 . In addition , current methods of manufacturing large within each liposome particle , resembling the “ layers of an diameter synthetic membrane vesicles, such as multivesicu onion ” . lar liposomes on a commercial scale , require significant The presence of internal membranes distributed as a commitments in manufacturing space , cost, and time. As network throughout multivesicular liposomes (MVL ) may such , developing stable multivesicular liposome formula - 50 serve to confer increased mechanical strength to the vesicle , tions containing a therapeutic agent in a cost effective and while still maintaining a high volume: lipid ratio compared timely manner remains an ongoing challenge . with multilamellar vesicles . The multivesicular nature of In large scale manufacturing of large diameter synthetic multivesicular liposomes MVL( ) also indicates that, unlike membrane vesicles, the present embodiments require less for unilamellar liposomes, a single breach in the external water for injection (WFI ) , space , time and energy to produce 55 membrane of a multivesicular liposomes MVL( ) will not an equivalent amount of the large diameter synthetic mem - result in total release of the internal aqueous contents . Thus , brane vesicles than under previously described large scale both structurally and functionally the multivesicular lipo manufacturing conditions. As a result, waste disposal is somes (MVL ) are unusual, novel and distinct from all other reduced and overall costs are reduced . For example , the types of liposomes . As a result , the functional properties of present embodiments provide systems that can be housed in 60 multivesicular liposomes (MVL ) are not predictable based a one story room ( e . g . 5x5 meter room ) that previously on the prior art related to conventional liposomes such as described systems would require a multistory room with at unilamellar liposomes and multilamellar liposomes. least a ten fold increase in the area of the room . Additionally , In some embodiments, the large diameter synthetic mem the present embodiments provide systems that are particu - brane vesicles are multivesicular liposomes . In some larly well adapted to continuous processing for more rapid 65 embodiments , the multivesicular liposomes further com production of large diameter synthetic membrane vesicles prise bupivaciane, DEPC , DPPG , and tricaprylin . In some and allowing for more efficient implementation of clean - in - embodiments , the multivesicular liposomes further com US 9 , 730 , 892 B2 13 14 prise bupivacaine phosphate , DEPC , DPPG , and tricaprylin . claims, taken in conjunction with the accompanying draw In some embodiments , the multivesicular liposomes further ings. It will be understood these drawings depict only certain comprise bupivacaine , DEPC , DPPG , tricaprylin and cho - embodiments in accordance with the disclosure and , there lesterol. In some embodiments , the multivesicular liposomes fore , are not to be considered limiting of its scope ; the further comprise bupivacaine phosphate , DEPC , DPPG , 5 disclosure will be described with additional specificity and tricaprylin and cholesterol. detail through use of the accompanying drawings . An appa In some embodiments , the multivesicular liposomes fur ratus , system or method according to some of the described ther comprise bupivaciane, dextrose , L - Lysine, DEPC , embodiments can have several aspects , no single one of DPPG , and tricaprylin . In some embodiments , the multive sicular liposomes further comprise bupivacaine phosphate , which necessarily is solely responsible for the desirable dextrose , L -Lysine , DEPC , DPPG , and tricaprylin . In some 10 attributes of the apparatus, system or method . After consid embodiments, the multivesicular liposomes further com ering this discussion , and particularly after reading the prise bupivacaine , dextrose , L - Lysine , DEPC , DPPG , tri section entitled “ Detailed Description of the Preferred caprylin and cholesterol . In some embodiments , the multi Embodiment” one will understand how illustrated features vesicular liposomes further comprise bupivacaine serve to explain certain principles of the present disclosure. phosphate , dextrose , L -Lysine , DEPC , DPPG , tricaprylin 15 In the following detailed description , only certain exem and cholesterol. plary embodiments have been shown and described , simply In some embodiments , the multivesicular liposomes fur by way of illustration . As those skilled in the art would ther comprise bupivaciane , dextrose , DEPC , DPPG , and realize , the described embodiments may be modified in tricaprylin . In some embodiments, the multivesicular lipo - various different ways , all without departing from the spirit somes further comprise bupivacaine phosphate , dextrose , 20 or scope of the present disclosure . Accordingly , the drawings DEPC , DPPG , and tricaprylin . In some embodiments , the and description are to be regarded as illustrative in nature multivesicular liposomes further comprise bupivacaine, and not restrictive . In addition , when an element is referred dextrose , DEPC , DPPG , tricaprylin and cholesterol. In some to as being “ on ” another element , it can be directly on the embodiments , the multivesicular liposomes further com another element or be indirectly on the another element with prise bupivacaine phosphate , dextrose , DEPC , DPPG , tri - 25 one or more intervening elements interposed therebetween . caprylin and cholesterol. In some embodiments , the multivesicular liposomes fur Also , when an element is referred to as being “ connected to ” ther comprise bupivaciane , L - Lysine, DEPC , DPPG , and another element, it can be directly connected to the another tricaprylin . In some embodiments, the multivesicular lipo element or be indirectly connected to the another element somes further comprise bupivacaine phosphate , L -Lysine , with one or more intervening elements interposed therebe DEPC , DPPG , and tricaprylin . In some embodiments , the 30 tween . Hereinafter , like reference numerals refer to like multivesicular liposomes further comprise bupivacaine , elements . Since the disclosure may be modified in various L - Lysine , DEPC , DPPG , tricaprylin and cholesterol. In ways and have various embodiments , the disclosure will be some embodiments , the multivesicular liposomes further described in detail with reference to the drawings . However, comprise bupivacaine phosphate , L -Lysine , DEPC , DPPG it should be understood that the disclosure is not limited to tricaprylin and cholesterol. 35 a specific embodiment but includes all changes and equiva In some embodiments , the multivesicular liposomes fur lent arrangements and substitutions included in the spirit and ther comprise bupivacaine, morphine , cytarabine , or their scope of the disclosure . In the following description , if the pharmaceutically acceptable salts as the therapeutic agent. detailed description of the already known structure and In some embodiments , the multivesicular liposomes further operation may confuse the subject matter of the present comprise bupivacaine phosphate , morphine sulfate , or cyt - 40 disclosure , the detailed description thereof will be omitted . arabine HCl. While such terms as “ first, " " second ,” etc ., may be used In another embodiment, any one of the above described to describe various components, such components must not embodiments can be used alone or in combination with any be limited to the above terms. The above terms are used only one or more of the above described embodiments . For to distinguish one component from another. Terms used in example , any above described atomizing nozzle , evapora - 45 the following description are to describe specific embodi tion apparatus, continuous - flow emulsification system , con - ments and is not intended to limit the disclosure . The tinuous- flow diafiltration system , continuous- flow diafiltra expression of singularity includes plurality meaning unless tion further comprising one or more centrifuges, continuous the singularity expression is explicitly different in context. It flow centrifuge system , or continuous processing system can should be understood that the terms " comprising, ” “ having, " be used alone or in combination . Thus , an evaporation 50 “ including ,” and “ containing” are to indicate features , num apparatus can be used in conjunction with a three - fluid bers , steps, operations , elements , parts, and / or combinations atomizing nozzle . This evaporation system /atomizing nozzle but not to exclude one or more features, numbers , steps, can be used with a continuous - flow emulsification system , operations, elements , parts, and / or combinations or addi as depicted in FIGS. 1A , 1B , and 1C . The three - fluid tional possibilities . atomizing nozzle /evaporation apparatus combination can be 55 Some embodiments provide continuous processes for used in conjunction with a continuous - flow system , as making multivesicular liposomes . Prior methods of making depicted in FIGS. 8 , 10 , and 11 . Any of these combinations multivesicular liposomes required batch processing . This can be used to make multivesicular liposomes . In particular batch processing required removal of the solvent from the any of the combinations can be used to make multivesicular droplets of first emulsion surrounded by a second aqueous liposomes containing bupivacaine or its salts as the thera - 60 phase by contacting the suspension of first emulsion droplets peutic agent. in a continuous aqueous phase with a discontinuous gas phase by sparging (bubbling ) gas through the aqueous phase DETAILED DESCRIPTION OF THE or blowing gas over a flask containing continuous aqueous PREFERRED EMBODIMENT phase . This batch processing takes tens of minutes to remove 65 the solvent . Features of the present disclosure will becomemore fully It was surprisingly discovered that forming a first emul apparent from the following description and appended sion surrounded by an aqueous shell in the form of a droplet US 9 ,730 ,892 B2 15 and contacting it with a continuous gas phase, reduces the peutic agent ; preparing a droplet ( s ) by mixing said first time needed to remove the organic solvent to a few seconds component and a second aqueous phase , said emulsion and possibly a fraction of a second . (much less than the tens droplet( s ) comprising an aqueous phase ; preparing a multi of minutes stated above for batch processing ) . vesicular liposome particle by removing the organic solvent This is due to the tremendous gas contacting surface area 5 from the emulsion droplet , wherein the removing comprises of the atomized droplets ; and themuch faster diffusion of the contacting the emulsion droplet with a gas ; and preparing a solvent in gasses versus water ; and the very short distances multivesicular liposome composition by collecting the mul that the solvent has to diffuse through the aqueous phase to tivesicular liposome particles, wherein the multivesicular reach the gas ( now only microns instead of the distanced liposome composition is suspended in a continuous aqueous between sparging bubbles . ) 10 phase . This extremely fast solvent removal enables the continu - Some embodiments relate to a process for preparing a ous processing. The solvent is removed in less than the time multivesicular liposome composition comprising preparing that it takes for the atomized droplets to reach the bottom of a first component by mixing a first aqueous phase and an the solvent removal vessel ( a few seconds at most ). organic phase , said organic phase comprising an organic Definitions 15 solvent, at least one amphipathic lipid and at least one As used herein , abbreviations are defined as follows : neutral lipid , wherein the first component comprises a thera aq . Aqueous peutic agent , preparing an emulsion of first component CIP Clean -in -place processing droplets , a second component droplet , by mixing said first ° C . Temperature in degrees Celsius component and a second aqueous phase , said second com d10 the diameter where 10 mass - % (volume % ) (of the 20 ponent droplet comprising an aqueous phase , wherein the particles ) of the particles have a smaller equivalent diam - second component droplet is prepared using a device as eter, and the other 90 mass - % ( volume % ) have a larger described herein , preparing a multivesicular liposome par equivalent diameter in um ticle by removing the organic solvent from the second d50 the diameter where 50 mass - % ( volume % ) ( of the component droplet , wherein the removing comprises con particles ) of the particles have a smaller equivalent diam - 25 tacting the second component droplet with a gas, and eter , and the other 50 mass - % (volume % ) have a larger preparing a multivesicular liposome composition by collect equivalent diameter in um ing the multivesicular liposome particles, wherein the mul d90 the diameter where 90 mass - % (volume % ) (of the tivesicular liposome composition is suspended in a continu particles ) of the particles have a smaller equivalent diam - ous aqueous phase . eter , and the other 100 mass - % (volume % ) have a larger 30 As used herein the term “ amphipathic lipid ” refers to a equivalent diameter in um substance including a hydrophilic region and a hydrophobic DCM methylene chloride region , such as phospholipids. Amphipathic lipids can be g Gram ( s ) zwitterionic phospholipids , zwitterionic lipids , lipids having h Hour (hours ) a net negative charge , and lipids having a net positive mL Milliliter( s ) 35 charge . For example , amphipathetic lipids include , but are mg Milligram (s ) not limited to , phosphatidylcholines, phosphatidylserines , mOsm /kg Osmolality per kilogram phosphatidylethanolamines , phosphatidylinositols , sphingo pH measure of the acidity or alkalinity of a liquid using a pH myelin , soybean lecithin ( soya lecithin ), egg lecithin , lyso meter or pH indicator phosphatidylcholines, lysophosphatidylethanolamines , PPV Packed particle volume 40 phosphatidylglycerols , phosphatidylserines , phosphati PSD Particle size distribution dylinositols , phosphatidic acids , cardiolipins , acyl trimeth RT, rt Room temperature ylammonium propane , diacyldimethylammonium propane , SIP Sterile - in - place processing stearylamine , ethyl phosphatidylcholine and the like . Phos Tert, t tertiary pholipids used in the methods described herein can be of a uL Microliter ( s ) 45 single class or a mixture of classes . Some embodiments ug Microgram ( s ) include crude preparations of phospholipids, such as soy WFI Water for injection bean lecithin (soya lecithin ) and egg lecithin . Soya lecithin Some embodiments relate to a process for preparing a is a combination predominantly of naturally - occurring phos large diameter synthetic membrane vesicle composition pholipids; phosphatidylcholine (PC ), phosphatidyletha comprising preparing a first component by mixing a first 50 nolamine ( PE ) and phosphatidylinositol (PI ) . Examples of aqueous phase and an organic phase , said organic phase phosphatidylcholines and phosphatidylglycerols include , comprising an organic solvent, at least one amphipathic lipid but are not limited to , 1 , 2 - dioleoyl- sn - glycero - 3 - phospho and at least one neutral lipid ; preparing a droplet (s ) by choline, 1, 2 -dilauroyl - sn - glycero -3 -phosphocholine , 1, 2 mixing said first component and a second aqueous phase , dimyristoyl- sn - glycero - 3 -phosphocholine , 1 ,2 -dipalmitoyl said droplet( s ) comprising an aqueous phase ; preparing a 55 sn - glycero - 3 -phosphocholine , 1 , 2 - distearoyl- sn large diameter synthetic membrane vesicle by removing the glycero - 3 - phosphocholine, 1 , 2 - diarachidoyl- sn - glycero - 3 organic solvent from the droplet ( s ) , wherein the removing phosphocholine, 1 , 2 - dibehenoyl -sn - glycero - 3 -phosphocho comprises contacting the droplet ( s ) with a gas; and collect- line, 1 ,2 -dipalmitoleoyl - sn - glycero - 3 -phosphocholine , 1, 2 ing the large diameter synthetic membrane vesicle particles , dieicosenoyl- sn - glycero - 3 -phosphocholine , 1, 2 -dierucoyl wherein the large diameter synthetic membrane vesicle is 60 sn - glycero - 3 -phosphocholine , 1 , 2 - dipalmitoyl- sn - glycero suspended in a continuous aqueous phase . 3 - phosphoglycerol, 1 , 2 - dioleoyl- sn - glycero - 3 Some embodiments relate to a process for preparing a phosphoglycerol, and mixtures thereof. multivesicular liposome composition comprising preparing As used herein the term “ neutral lipid ” refers to oils , a first component by mixing a first aqueous phase and an waxes or fatty acid esters that lack a charged or hydrophilic organic phase , said organic phase comprising an organic 65 head group . Neutral lipids include but are not limited to , solvent, at least one amphipathic lipid and at least one glycerol esters , glycol esters, tocopherol esters , sterol esters , neutral lipid , wherein the first component comprises a thera - hydrocarbons and squalenes . US 9 ,730 , 892 B2 17 18 As used herein the terms “ glycerol esters , " " triglycer tered and does not abrogate the biological activity and ides, ” and “ triacylglycerols” refer to triesters formed from properties of the compound . In some embodiments , the salt glycerol and fatty acids. Glycerol esters include but are not is an acid addition salt of the compound . Pharmaceutical limited to , triolein , tripalmitolein , trimyristolein , trilinolein , salts can be obtained by reacting a compound with inorganic tributyrin , tricaprylin , tricaproin , and tricaprin . 5 acids such as hydrohalic acid ( e . g . , hydrochloric acid or As used herein the term " organic solvent” refers to ethers , hydrobromic acid ) , sulfuric acid , nitric acid , phosphoric acid esters, halogenated ethers, aromatic or aliphatic hydrocar and the like . Pharmaceutical salts can also be obtained by bons , aromatic or aliphatic halohydrocarbons , or Freons . reacting a compound with an organic acid such as aliphatic Organic solvents include but are not limited to , diethyl ether , or aromatic carboxylic or sulfonic acids, for example acetic , tert- butylmethyl ether, tetrahydrofuran , sevoflurane, desflu - 10 succinic , lactic , malic , tartaric , citric , ascorbic , nicotinic , rane , isoflurane , enflurane , halothane , chloroform , dichlo - methanesulfonic, ethanesulfonic , p -toluensulfonic , salicylic romethane , ethyl acetate , hexane, hexanes, cyclohexane , or naphthalenesulfonic acid . Pharmaceutical salts can also pentane , cyclopentane , petroleum ether, toluene , and any be obtained by reacting a compound with a base to form a combinations thereof. salt such as an ammonium salt , an alkali metal salt, such as As used herein the term " aqueous phase ” refers to any 15 a sodium or a potassium salt , an alkaline earth metal salt, solution or mixture having water as the major component such as a calcium or a magnesium salt , a salt of organic The aqueous phase can include constituents , such as pH bases such as dicyclohexylamine, N -methyl - D - glucamine , buffering agents , salts , osmotic agents , simple sugars , amino tris (hydroxymethyl )methylamine , C . - C , alkylamine , cyclo acids , electrolytes , preservatives , other water soluble excipi- hexylamine , triethanolamine , ethylenediamine , and salts ents and the like . Aqueous phase constituents can include, 20 with amino acids such as arginine , lysine , and the like . In but are not limited to , sodium chloride , hydrochloric acid , some embodiments , the therapeutic agent can have a low phosphoric acid , lysine, dextrose , glucose and the like . aqueous solubility in neutral form . In some embodiments , As used herein the term “ fluid ” refers to a substance that the pharmaceutically acceptable salt of a therapeutic agent has the ability to flow . Fluids can be a gas, a liquid , a liquid can have higher aqueous solubility in comparison to a with a substance ( s ) suspended throughout the liquid , an 25 therapeutic agent in neutral form . emulsion , a vapor, or a gas / vapor mixture . Many and varied therapeutic agents can be incorporated As used herein the term “ therapeutic agent” and “ drug” by encapsulation within the synthetic membrane vesicles . A refers to a chemical compound , mixtures of chemical com - non - limiting list of therapeutic agent classes include , but are pounds, or biological molecules , such as biological macro - not limited to , antianginas, antiarrhythmics , antiasthmatic molecules or peptides that may have therapeutic properties . 30 agents , antibiotics , antidiabetics , antifungals , antihista The therapeutic agent can be purified , substantially purified mines , antihypertensives, antiparasitics, antineoplastics , or partially purified . In some embodiments, the therapeutic antiobesity agents , antiviral agents , otologicals, cardiac gly agent can be selected from the group including antianginas, cosides , hormones, immunomodulators , monoclonal anti antiarrhythmics, antiasthmatic agents , antibiotics , antidi bodies , neurotransmitters , sedatives, vaccines, vasopressors , abetics , antifungals , antihistamines, antihypertensives , anti- 35 anesthetics, amide anaesthetics , corticosteroids, tricyclic parasitics , antineoplastics , antitumor drugs, antivirals , car - antidepressants , tetracyclic antidepressants , selective sero diac glycosides , hormones , immunomodulators , monoclonal tonin reuptake inhibitors, steroid receptor modulators , antip antibodies , neurotransmitters , nucleic acids, proteins , radio sychotic drugs , antiprotozoal drugs, opioids , antiprolifera contrast agents , radionuclides, sedatives , analgesics , ste - tive agents , salicylanilides , antihelminthic drugs, vinca roids , tranquilizers , vaccines, vasopressors, anesthetics , pep - 40 alkaloids, anti - inflammatory agents , antidepressants , prosta tides and the like . Any pharmaceutically acceptable salt of a glandins, phosphodiesterase IV inhibitors ; retinoids, ste particular therapeutic agent is also envisioned as being roids , 3 - adrenergic receptor ligands, anti -mitotic agents , useful in the present embodiments . In some embodiments , microtubule inhibitors ,microtubule - stabilizing agents , sero the therapeutic agent can be introduced in either an aqueous tonin norepinephrine reuptake inhibitors, noradrenaline or a solvent phase , depending on their solubility in these 45 reuptake inhibitors , non - steroidal immunophilin - dependent phases . In a typical embodiment, the therapeutic agent can immunosuppressants , non -steroidal immunophilin - depen be selected from the group including semisynthetic amino dent immunosuppressant enhancers ; antimalarial agents , glycoside antibiotics such as amikacin ; antidiabetics ; pep - analgesics, immunosuppressants , expectorants , sulfa drugs, tides such as insulin ; antitumor drugs such as paclitaxel; cardiovascular drugs, central nervous system ( CNS ) depres antineoplastics including cytarabine, 5 - fluorouracil and 50 sants , H2- blockers , anti - platelet drugs , anticonvulsants , floxuridine ; alkaloid opiate analgesics including morphine alpha blockers, beta - blockers, cholinesterase inhibitors, cal and hydromorphine ; local anesthetics including bupiva cium channel blockers , H1- receptor antagonists , and pro caine ; synthetic anti- inflammatory adrenocortical steroids teinaceous materials . The therapeutic agents listed herein including dexamethasone ; antimetabolites including metho - can be used in the preparation of medicaments for the trexate ; glycopeptide antibiotics including bleomycin ; vin - 55 treatment of a disease for which the therapeutic agent is caleukoblastines and stathmokinetic oncolytic agents known to those of skill in the art to be effective . Therapeutic including vincristine and vinblastine ; hormones , plasma agents , and diseases for which the therapeutic agent is proteins , cytokines, growth factors, DNA and RNA from a effective, can be identified by reference to , for example , The variety of organisms, and antisense oligonucleotides . In Physician ' s Desk Reference , which is incorporated herein some embodiments, the therapeutic agent can be an amide 60 by reference in its entirety . anesthetic . Amide anesthetics include, but are not limited to , Examples of proteinaceous materials that can be incor bupivacaine , mepivacaine , ropivacaine, lidocaine, pyrro - porated into the synthetic membrane vesicles , include but caine , prilocaine, their stereoisomers , and combinations are not limited to , DNA , RNA , proteins of various types , thereof, and pharmaceutically acceptable salts thereof. protein hormones produced by recombinant DNA technol As used herein the term “ pharmaceutically acceptable 65 ogy effective in humans, hematopoietic growth factors , salt ” refers to a salt of a compound that does not cause monokines, lymphokines, tumor necrosis factor , inhibin , significant irritation to an organism to which it is adminis - tumor growth factor alpha and beta , Mullerian inhibitory US 9 , 730 ,892 B2 19 20 substance , nerve growth factor, fibroblast growth factor, moxastine , p -methyldiphenhydramine , pemirolast , pheni platelet - derived growth factor, pituitary and hypophyseal ramine, phenyltoloxamine, resporal , rondec, semprex - d , set hormones including LH and other releasing hormones. astine, sominex , talastine , terfenadine, thenyldiamine, thi Examples of antiarrhythmics, include but are not limited azinamium , and triprolidine . to , quinidine, procainamide , disopyramide, ajmaline , lido - 5 Examples of antihypertensives , include but are not limited caine , tocainide, mexiletine, flecainide, propafenone , mori - to , bumetanide , ethacrynic acid , furosemide , torsemidet , cizine, propranolol, esmolol, timolol , metoprolol, atenolol, epitizide , hydrochlorothiazide, chlorothiazide, bendroflu amiodarone , sotalol, ibutilide , dofetilide, verapamil, dilti - methiazide, indapamide , chlorthalidone , metolazone , azem , and digoxin . amiloride , triamterene , spironolactone , atenolol, metoprolol, Examples of antiasthmatic agents , include but are not 10 nadolol, oxprenolol , pindolol, propranolol, timolol , doxa limited to , salbutamol, levalbuterol, terbutaline , bitolterol, zosin , phentolamine , indoramin , phenoxybenzamine, pra epinephrine , ipratropium bromide , salmeterol, formoterol, zosin , terazosin , tolazoline, bucindolol , carvedilol , labetalol , bambuterol, and albuterol. clonidine, methyldopa , guanfacine, amlodipine , felodipine , Examples of antibiotics, include but are not limited to , isradipine , lercanidipine, nicardipine, nifedipine, nimo amikacin , gentamicin , kanamycin , neomycin , netilmicin , 15 dipine , nitrendipine , diltiazem , verapamil, captopril , enal streptomycin , tobramycin , paromomycin , geldanamycin , april , fosinopril , lisinopril , perindopril, quinapril , ramipril , herbimycin , loracarbef, ertapenem , doripenem , imipenem , trandolapril , benazepril, candesartan , eprosartan , irbesartan , meropenem , cefadroxil , cefazolin , cefalotin , cefalexin , cefa - losartan , olmesartan , telmisartan , valsartan , eplerenone , clor, cefamandole, cefoxitin , cefprozil , cefuroxime, spironolactone , sodium nitroprusside , clonidine , guanabenz , cefixime, cefdinir , cefditoren , cefoperazone , cefotaxime, 20 methyldopa , moxonidine, guanethidine, and reserpine . cefpodoxime, ceftazidime, ceftibuten , ceftizoxime, ceftriax Examples of antiparasitics , include but are not limited to , one, cefepime, ceftobiprole , teicoplanin , vancomycin , mebendazole , pyrantel pamoate , thiabendazole , diethycar azithromycin , clarithromycin , dirithromycin , erythromycin , bazine , niclosamide , praziquantel, rifampin , amphotericin roxithromycin , troleandomycin , telithromycin , spectinomy - B , and melarsoprol . cin , aztreonam , amoxicillin , ampicillin , azlocillin , carbeni- 25 Examples of antineoplastics , include but are not limited cillin , cloxacillin , dicloxacillin , flucloxacillin , mezlocillin , to , aclarubicin , altretamine , aminopterin , amrubicin , azaciti meticillin , nafcillin , oxacillin , penicillin , piperacillin , ticar - dine, azathioprine , belotecan , busulfan , camptothecin , cillin , bacitracin , colistin , polymyxin b , ciprofloxacin , capecitabine , carboplatin , carmofur , carmustine , chlorambu enoxacin , gatifloxacin , levofloxacin , lomefloxacin , moxi- cil, cisplatin , cladribine , clofarabine , cyclophosphamide , floxacin , norfloxacin , ofloxacin , trovafloxacin , grepafloxa - 30 cytarabine , daunorubicin , decitabine , doxorubicin , epirubi cin , sparfloxacin , mafenide , prontosil , sulfacetamide, sul- cin , etoposide, floxuridine , fludarabine, 5 - fluorouracil , fluo famethizole , sulfanilimide , sulfasalazine , sulfisoxazole , rouracil, gemcitabine , idarubicin , ifosfamide , irinotecan , trimethoprim , trimethoprim - sulfamethoxazole , demeclocy - mechlorethamine , melphalan , mercaptopurine, methotrex cline , doxycycline , minocycline , oxytetracycline , tetracy ate , mitoxantrone, nedaplatin , oxaliplatin , pemetrexed , pen cline , arsphenamine , chloramphenicol, clindamycin , linco - 35 tostatin , pirarubicin , pixantrone , procarbazine , pyrimeth mycin , ethambutol, fosfomycin , fusidic acid , furazolidone , amine raltitrexed , rubitecan , satraplatin , streptozocin , isoniazid , linezolid , metronidazole , mupirocin , nitrofuran - thioguanine, triplatin tetranitrate , teniposide , topotecan , toin , platensimycin , pyrazinamide , quinupristin , rifampin , tegafur, trimethoprim , uramustine , valrubicin , vinblastine , thiamphenicol , and tinidazole . vincristine , vindesine , vinflunine , vinorelbine , and zorubi Examples of antidiabetics, include but are not limited to , 40 cin . tolbutamide , acetohexamide, tolazamide , chlorpropamide , Examples of antiviral agents , include but are not limited glipizide , glyburide , glimepiride, gliclazide , repaglinide, to , abacavir , aciclovir, acyclovir , adefovir , amantadine , nateglinide , metformin , rosiglitazone, pioglitazone , troglita - amprenavir, arbidol, atazanavir , atripla , boceprevir , cidofo zone , miglitol, acarbose , exenatide , liraglutide , taspoglatide , vir , combivir , darunavir , delavirdine , didanosine , edoxudine , vildagliptin , sitagliptin , GLP - 1 , and analog to GLP - 1 . 45 efavirenz, emtricitabine , enfuvirtide , entecavir , famciclovir , Examples of antifungals , include but are not limited to , fomivirsen , fosamprenavir , foscarnet , fosfonet , ganciclovir , natamycin , rimocidin , filipin , nystatin , amphotericin B , can ibacitabine , imunovir, idoxuridine , imiquimod , indinavir , dicin , miconazole , ketoconazole , clotrimazole , econazole , inosine , lamivudine , lopinavir , loviride , maraviroc , moroxy bifonazole , butoconazole , fenticonazole , isoconazole , oxi- dine , nelfinavir , nevirapine , nezavir , oseltamivir , penciclo conazole , sertaconazole, sulconazole , tioconazole , flucon - 50 vir , peramivir , pleconaril , raltegravir , ribavirin , rimantadine, azole , itraconazole, isavuconazole , ravuconazole , posacon - ritonavir , saquinavir , stavudine , tenofovir , tenofovir diso azole , voriconazole , terconazole , abafungin , terbinafine , proxil , tipranavir , trifluridine, trizivir, tromantadine, valaci amorolfine, naftifine, butenafine , anidulafungin , caspo - clovir , valganciclovir , vicriviroc , vidarabine , viramidine , fungin , micafungin , ciclopirox , tolnaftate , undecylenic acid , zalcitabine , zanamivir , and zidovudine. 5 - fluorocytosine , and griseofulvin . 55 Examples of otologicals , include but are not limited to , Examples of antihistamines , include but are not limited to , betamethasone, chloramphenicol, chlorhexidine , clioquinol, aceprometazine , alimemazine, astemizole , azatadine , dexamethasone, gentamicin , hydrocortisone , lidocaine , azelastine , benadryl, bepotastine , bisulepine , brompheni- miconazole , neomycin , nitrofural, polymyxin b , predniso ramine , chlorcyclizine , chloropyramine , chlorothen , chlo - lone, rifamycin , and tetracycline . rphenamine , cinnarizine , clemastine , clemizole , clobenze - 60 Examples of cardiac glycosides , include but are not pam , clobenztropine , clocinizine, cyclizine, cyproheptadine, limited to , digitoxin , digoxin , and deslanoside . dacemazine , dexbrompheniramine , dexchlorpheniramine , Examples of hormones , include but are not limited to , diphenhydramine , doxylamine , drixoral, ebastine , adiponectin , adrenocorticotropic hormone , aldosterone , embramine , emedastine , epinastine , etymemazine, fex - androstenedione , angiotensinogen , angiotensin , antidiuretic ofenadine , homochlorcyclizine, hydroxyzine , iproheptine , 65 hormone , antimullerian hormone , atrial -natriuretic peptide , isopromethazine , ketotifen , levocabastine , mebhydrolin , brain natriuretic peptide , 25 - hydroxyvitamin D2, calcitonin , mepyramine, methafurylene, methapyrilene, methdilazine , 1 ,25 - dihydroxyvitamin D3, cholecystokinin , corticotropin US 9 ,730 ,892 B2 22 releasing hormone , cortisol, dehydroepiandrosterone, dihy - mitumomab , morolimumab , motavizumab , muromonab , drotestosterone , dopamine , endothelin , enkephalin , epineph - stamulumab , nacolomab tafenatox , naptumomab estafena rine , erythropoietin , estradiol, estriol, estrone , follicle tox , natalizumab , nebacumab , necitumumab , nerelimomab , stimulating hormone, gastrin , ghrelin , glucagon , nimotuzumab , nofetumomab merpentan , ocrelizumab , odu gonadotropin - releasing hormone , growth hormone , growth 5 limomab , ofatumumab , omalizumab , oportuzumab mona hormone -releasing hormone, histamine , human chorionic tox , oregovomab , otelixizumab , pagibaximab , palivizumab , gonadotropin , human placental lactogen , inhibin , insulin , panitumumab , panobacumab , pascolizumab , pemtumomab , insulin - like growth factor, leptin , leukotrienes, lipotropin , luteinizing hormone, melanocyte stimulating hormone , pertuzumab , pexelizumab , pintumomab , priliximab , pritu melatonin , neuropeptide y , norepinephrine , orexin , oxyto - 10 mumab , rafivirumab , ramucirumab , ranibizumab , raxibacu cin , pancreatic polypeptide , parathyroid hormone , proges mab , regavirumab , reslizumab , rilonacept, rilotumumab , terone , prolactin , prolactin releasing hormone , prostacyclin , rituximab , robatumumab , rovelizumab , rozrolimupab , rupli prostaglandins , relaxin , renin , secretin , serotonin , soma zumab , satumomab , sevirumab , sibrotuzumab , siltuximab , tostatin , testosterone , thrombopoietin , thromboxane , thy siplizumab , solanezumab , sonepcizumab , sontuzumab , sta roid -stimulating hormone , thyrotropin - releasing hormonemone ., 15 mulumabm , sulesomab , tacatuzumab tetraxetan , tadocizumab , thyroxine , and triiodothyronine . talizumab , tanezumab , taplitumomab paptox , tefibazumab , Examples of immunomodulators , include but are not telimomab aritox , tenatumomab , teneliximab , teplizumab , limited to , abatacept, abetimus , adalimumab , afelimomab , ticilimumab , tigatuzumab , tocilizumab , toralizumab , tositu aflibercept, afutuzumab , alefacept, anakinra , aselizumab , momab , trastuzumab , tremelimumab , tucotuzumab celmo atlizumab , atorolimumab , azathioprine, basiliximab , belata - 20 leukin , tuvirumab , urtoxazumab , ustekinumab , vapaliximab , cept, belimumab , bertilimumab , cedelizumab , clenoliximab , vedolizumab , veltuzumab , vepalimomab , visilizumab , certolizumab pegol, ciclosporin , daclizumab , deforolimus, volociximab , votumumab , zalutumumab , zanolimumab , dorlimomab aritox , dorlixizumab , efalizumab , erlizumab , ziralimumab , and zolimomab aritox . elsilimomab , etanercept, everolimus, faralimomab , fontoli Examples of neurotransmitters, include but are not limited zumab , galiximab , gantenerumab , gavilimomab golimumab , 25 to , acetylcholine , adenosine , adenosine - 5 ' - triphosphate , gomiliximab , gusperimus, infliximab , inolimomab , ipilim - aspartate , norepinephrine, dopamine , glycine, serotonin , umab keliximab , lebrilizumab , leflunomide, lenalidomide, melatonin , histamine, glutamate , gamma aminobutyric acid , lerdelimumab , lumiliximab , maslimomab , mepolizumab , and guanosine - 5 - triphosphate . metelimumab , methotrexate , morolimumab , muromonab - Examples of sedatives , include but are not limited to , cd3 , mycophenolic acid , natalizumab , nerelimomab , ocreli - 30 alprazolam , amobarbital, carisoprodol, chlordiazepoxide , zumab , odulimomab , omalizumab , otelixizumab , pascoli clomethiazole , clonazepam , diazepam , diphenhydramine , zumab , pexelizumab , pimecrolimus, reslizumab , rilonacept, estazolam , eszopiclone , ethchlorvynol, flunitrazepam , rovelizumab , ruplizumab , siplizumab , sirolimus , tacrolimus, gamma -hydroxybutyrate , glutethimide , ketamine , loraze talizumab , telimomab aritox , temsirolimus , teneliximab , pam , methaqualone , methyprylon , midazolam , nitrazepam , teplizumab , teriflunomide, thalidomide, tocilizumab , torali - 35 oxazepam , pentobarbital, phenobarbitoltriazolam , ramelt zumab , tremelimumab , ustekinumab , vapaliximab , vepali - eon , secobarbital, temazepam , thalidomide , zaleplon , zolpi momab , visilizumab , zanolimumab , ziralimumab , zoli - dem , and zopiclone . momab aritox , zotarolimus , and tetrachlorodecaoxide . Examples of vaccines , include but are not limited to , Examples of monoclonal antibodies , include but are not measles vaccine , mumps vaccine, rubella vaccine , varicella limited to , abagovomab , abatacept, abciximab , adalimumab , 40 vaccine , inactivated polio vaccine , inactivated influenza adecatumumab , aflibercept, afutuzumab , alacizumab pegol, vaccine , influenza a virus subtype H1N1 vaccine , diphtheria alemtuzumab , altumomab , afelimomab , anatumomab toxoid vaccine , tetanus toxoid vaccine , haemophilus influ mafenatox , anrukinzumab , apolizumab , arcitumomab , aseli - enzae type B vaccine , hepatitis B vaccine , hepatitis A zumab , atlizumab , atorolimumab , bapineuzumab , basilix - vaccine, and pneumoccocal conjugate vaccine . imab , bavituximab , bectumomab , belatacept, belimumab , 45 Examples of vasopressors , include but are not limited to , bertilimumab , besilesomab , bevacizumab , biciromab bral - epinephrine , phenylephrine , dobutamine, isoproterenol, nor lobarbital, bivatuzumab mertansine , blinatumomab , briaki epinephrine, aceprometazine , alimemazine , astemizole , aza numab , canakinumab , cantuzumab mertansine , capromab tadine , azelastine, benadryl, bepotastine , bisulepine , bro pendetide , catumaxomab , cedelizumab , certolizumab pegol, mpheniramine , chlorcyclizine , chloropyramine , chlorothen , cetuximab , citatuzumab bogatox , cixutumumab , clenolix - 50 chlorphenamine , cinnarizine, clemastine , clemizole , cloben imab , golimumab , ustekinumab , conatumumab , dacetu - zepam , clobenztropine , clocinizine , cyclizine , cyprohepta zumab , dacliximab , daclizumab , denosumab , detumomab , dine, dacemazine , dexbrompheniramine , dexchlorpheni dorlimomab aritox , dorlixizumab , ecromeximab , eculi ramine , diphenhydramine , doxylamine , drixoral , ebastine, zumab , edobacomab , edrecolomab , efalizumab , efungumab , embramine, emedastine , epinastine , etymemazine, fex elsilimomab , enlimomab pegol, epitumomab cituxetan , 55 ofenadine , homochlorcyclizine, hydroxyzine , iproheptine, epratuzumab , erlizumab , ertumaxomab , etanercept, etaraci isopromethazine, ketotifen , levocabastine, mebhydrolin , zumab , exbivirumab , fanolesomab , faralimomab , felvi mepyramine , methafurylene, methapyrilene , methdilazine , zumab , fezakinumab , figitumumab , fontolizumab , fora moxastine , p -methyldiphenhydramine , pemirolast , pheni virumab , galiximab , gantenerumab , gavilimomab , ramine , phenyltoloxamine, resporal, rondec, semprex -d , set gemtuzumab ozogamicin , golimumab , gomiliximab , ibali - 60 astine , sominex , talastine , terfenadine , thenyldiamine , thi zumab , ibritumomab tiuxetan , igovomab , imciromab , inflix - azinamium , and triprolidine . imab , intetumumab , inolimomab , inotuzumab ozogamicin , Examples of anesthetics, include but are not limited to , ibalizumab , ipilimumab , iratumumab , keliximab , labetu propofol, etomidate , methohexital and sodium thiopental , zumab , lemalesomab , lebrilizumab , lerdelimumab , lexatu - midazolam , diazepam , and ketamine, benzocaine , chloro mumab , libivirumab , lintuzumab , lucatumumab , lumilix - 65 procaine, cocaine, cyclomethycaine, dimethocaine , imab , mapatumumab , maslimomab , matuzumab , propoxycaine , procaine , proparacaine, tetracaine , articaine , mepolizumab , metelimumab , milatuzumab , minretumomab , bupivacaine, carticaine , dibucaine , etidocaine , levobupiva US 9 , 730 , 892 B2 23 24 caine , lidocaine , mepivacaine , piperocaine , prilocaine , ropi- taxel, pemetrexed , pentostatin , pirarubicin , pixantrone , pro vacaine , trimecaine , saxitoxin , and tetrodotoxin . carbazine, pyrimethamine raltitrexed , rubitecan , satraplatin , Examples of amide anesthetics, include but are not lim sirolimus, streptozocin , thioguanine, triplatin tetranitrate , ited to , articaine, bupivacaine , carticaine , dibucaine , etido teniposide , topotecan , tegafur, trimethoprim , uramustine, caine , levobupivacaine, lidocaine , mepivacaine , pipero - 5 valrubicin , vinblastine, vincristine , vindesine, vinflunine , caine , prilocaine , ropivacaine , and trimecaine . vinorelbine , and zorubicin . Examples of corticosteroids, include but are not limited Examples of salicylanilides , include but are not limited to , to , hydrocortisone , hydrocortisone acetate , cortisone acetate , niclosamide , oxyclozanide , and rafoxanide . tixocortol pivalate, prednisolone , methylprednisolone , pred Examples of antihelminthic drugs , include but are not nisone , triamcinolone acetonide , triamcinolone alcohol, 10 limited to , abamectin , albendazole , diethylcarbamazine , mometasone , amcinonide , budesonide , desonide , fluocino - mebendazole , niclosamide , ivermectin , suramin , thiabenda nide , fluocinolone acetonide, halcinonide , betamethasone , zole , pyrantel pamoate , levamisole , praziquantel , triclaben betamethasone sodium phosphate , dexamethasone , dexam - dazole , flubendazole , fenbendazole , emodepside , and ethasone sodium phosphate , fluocortolone, hydrocortisone - monepantel . 17 - butyrate , hydrocortisone - 17 - valerate , aclometasone 15 Examples of vinca alkaloids, include but are not limited dipropionate , betamethasone valerate , betamethasone dipro - to , vinblastine , vincristine , vindesine and vinorelbine. pionate , prednicarbate , clobetasone - 17 -butyrate , clobetasol- Examples of anti - inflammatory agents , include but are not 17 - propionate , fluocortolone caproate , fluocortolone piva - limited to , phenylbutazone , mofebutazone , oxyphenbuta late , and fluprednidene acetate . zone , clofezone , kebuzone , indometacin , sulindac , tolmetin , Examples of tricyclic antidepressants , include but are not 20 zomepirac , diclofenac , alclofenac, bumadizone, etodolac , limited to , amitriptyline , butriptyline, clomipramine , dosu - lonazolac , fentiazac , acemetacin , difenpiramide , oxameta lepin , doxepin , imipramine, lofepramine , trimipramine , cin , proglumetacin , ketorolac , aceclofenac , bufexamac , desipramine, nortriptyline, and protriptyline . piroxicam , tenoxicam , droxicam , lornoxicam , meloxicam , Examples of tetracyclic antidepressants , include but are ibuprofen , naproxen , ketoprofen , fenoprofen , fenbufen , not limited to , amoxapine, maprotiline , mianserin , mirtazap - 25 benoxaprofen , suprofen , pirprofen , flurbiprofen , indoprofen , ine , and setiptiline . tiaprofenic acid , oxaprozin , ibuproxam , dexibuprofen , Examples of selective serotonin reuptake inhibitors , flunoxaprofen , alminoprofen , dexketoprofen , mefenamic include but are not limited to , citalopram , dapoxetine, esci acid , tolfenamic acid , flufenamic acid , meclofenamic acid , talopram , fluoxetine, fluvoxamine , paroxetine , sertraline , celecoxib , rofecoxib , valdecoxib , parecoxib , etoricoxib , vilazodone, and zimelidine . 30 lumiracoxib , nabumetone, niflumic acid , azapropazone , glu Examples of antipsychotic drugs, include but are not cosamine, benzydamine , glucosaminoglycan polysulfate , limited to , haloperidol, droperidol, chlorpromazine , flu proquazone , orgotein , nimesulide , feprazone, diacerein , phenazine, perphenazine , prochlorperazine , thioridazine, tri morniflumate , tenidap , oxaceprol, and chondroitin sulfate . fluoperazine, mesoridazine, periciazine , promazine , triflu Examples of cancers that can be treated with an anticancer promazine , levomepromazine , promethazine , pimozide , 35 agent include , but are not limited to , head and neck cancer, chlorprothixene , flupenthixol, thiothixene , zuclopenthixol, breast cancer , colorectal cancer, gastric cancer, hepatic can clozapine, olanzapine, risperidone, quetiapine, ziprasidone , cer , bladder cancer, cervical cancer, endometrial cancer, lung amisulpride, asenapine , paliperidone , aripiprazole , and cancer (non -small cell ), ovarian cancer, pancreatic cancer , bifeprunox . prostate cancer; choriocarcinoma (lung cancer ); hairy cell Examples of antiprotozoal drugs , include but are not 40 leukemia , chronic lymphotic leukemia , acute lymphocytic limited to , eflornithine , furazolidone , melarsoprol, metron - leukemia (breast & bladder ), acute myelogenous leukemia , idazole, ornidazole , paromomycin sulfate , pentamidine , Hodgkin ' s lymphoma, non -Hodgkin ' s lymphoma ( osteo pyrimethamine , and tinidazole . genic sarcoma , adult soft tissue sarcoma ) , meningeal leuke Examples of opioids , include but are not limited to , mia , multiple myeloma, chronic myelogenous leukemia , endorphins , enkephalins , dynorphins , endomorphins , 45 erythroleukemia , and T - cell lymphoma. codeine , morphine , thebaine, oripavine , diacetylmorphine , Examples of inflammatory and autoimmune diseases that dihydrocodeine , hydrocodone , hydromorphone , nicomor - can be treated with an inflammatory agent include , but are phine, oxycodone, oxymorphone , fentanyl, alphamethylfen not limited to , B cell disorders, T cell disorders , rheumatoid tanyl, alfentanil, sufentanil , remifentanil , carfentanyl , ohm - arthritis (RA ) , systemic lupus erythematosus ( SLE ) , efentanyl, pethidine , ketobemidone, allylprodine , prodine , 50 Sjogren ' s syndrome, immune thrombocytopenic purpura propoxyphene , dextropropoxyphene, dextromoramide , bez - ( ITP ) , multiple sclerosis MS( ) , myasthenia Gravis (MG ) , itramide, piritramide , methadone, dipipanone , levomethadyl Graves disease , psoriasis , Hashimoto ' s disease , immune acetate , loperamide , diphenoxylate , dezocine , pentazocine , thrombocytopenic purpura , scleroderma , and inflamatory phenazocine, buprenorphine , dihydroetorphine, etorphine , bowel disease ( e. g . Crohn 's disease and ulcerative colitis ). butorphanol, nalbuphine, levorphanol, levomethorphan , 55 In some embodiments, the therapeutic agent can be used lefetamine , meptazinol, tilidine , tramadol , tapentadol, singly or in combination with the limitation that the amount nalmefene , naloxone , and naltrexone . of the physiologically active substance in the pharmaceutical Examples of antiproliferative agents , include but are not composition be sufficient to enable the diagnosis of , pro limited to , aclarubicin , altretamine , aminopterin , amrubicin , phylaxis against, or treatment of an undesired condition in a azacitidine , azathioprine , belotecan , busulfan , camptothecin , 60 living being . In some embodiments , the pharmaceutical capecitabine , carboplatin , carmofur, carmustine , chlorambu - compositions can be administered to a living being by any cil , cisplatin , cladribine , clofarabine, cyclophosphamide , desired route , for example, intramuscular, intra articular, cytarabine, daunorubicin , decitabine, docetaxel, doxorubi- epidural , intraperitoneal, subcutaneous, intra lymphatic , cin , epirubicin , etoposide, floxuridine, fludarabine, 5 - fluo oral , submucosal, transdermal , rectal , vaginal , intranasal , rouracil , fluorouracil, gemcitabine , idarubicin , ifosfamide, 65 intraocular , and by implantation under different kinds of irinotecan , mechlorethamine , melphalan , mercaptopurine , epithelia , including the bronchial epithelia , the gastrointes methotrexate , mitoxantrone , nedaplatin , oxaliplatin , pacli - tinal epithelia , the urogenital epithelia , and the various US 9 ,730 ,892 B2 25 26 mucous membranes of the body. Generally , the dosage will disclosed and described herein , said second component vary with the age , condition , sex and extent of the undesired comprising an aqueous phase , removing the organic solvent condition in the patient , and can be determined by one from the second component to form a composition of large skilled in the art . In some embodiments , the dosage range diameter synthetic membrane vesicle particles, wherein the appropriate for human use includes a range of from 0 . 1 to 5 removing can be accomplished by using a solvent removal 6 ,000 mg of the therapeutic agent per square meter of chamber as disclosed and described herein , and optionally surface area . Alternate dosage range can be based on weight filtering the composition by particle concentration . Such instead of surface area . In one embodiment, a human dosage of bupivacaine can be 50 - 1 ,000 mg, 100 - 600 mg 100 - 350 steps may be combined with other steps . In some embodi mg. For example , the human dosage of bupivacaine can be 10 ments the lipid phase can include cholesterol. In some approximately 300 mg . embodiments , the organic solvent can be a volatile water Methods of Preparation immiscible or sparingly miscible solvent. In some embodi Some embodiments relate to a process for preparing a ments , the first component can be a first emulsion . In some large diameter synthetic membrane vesicle ( s ) composition embodiments , the second component can be a second emul comprising the steps of, forming a first component by 15 sion . In some embodiments, the second component can be a mixing a first aqueous phase and an organic phase , said droplet. In some embodiments , the large diameter synthetic organic phase comprising an organic solvent, at least one membrane vesicle ( s ) can be multivesicular liposomes . amphipathic lipid , and at least one neutral lipid , encapsu Some embodiments relate to a process for preparing a lating said first component in a second aqueous phase to multivesicular liposome composition comprising the steps provide a second component using an atomizing nozzle as 20 of, forming a first component by mixing a first aqueous disclosed and described herein , said second component phase and an organic phase , said organic phase comprising comprising an aqueous phase , removing the organic solvent a volatile water - immiscible or sparingly miscible solvent, at from the second component to form a composition of large least one amphipathic lipid , and at least one neutral lipid , diameter synthetic membrane vesicle particles , wherein the encapsulating said first component in a second aqueous removing can be accomplished by contacting the second 25 phase to provide a second component, said second compo component with a gas , optionally heating and optionally nent comprising an aqueous phase , removing the volatile filtering the composition by particle concentration . Such water - immiscible or sparingly miscible solvent from the steps may be combined with other steps. In some embodi second component to form a composition ofMVL particles , ments the lipid phase can include cholesterol. In some wherein the removing can be accomplished by contacting embodiments , the organic solvent can be a volatile water - 30 the second component with a gas , and optionally filtering the immiscible or sparingly miscible solvent . In some embodi- multivesicular liposome composition by particle concentra ments, the first component can be a first emulsion . In some tion . Such steps may be combined with other steps . In some embodiments , the second component can be a second emul- embodiments the lipid phase can include cholesterol. sion . In some embodiments , the second component can be a First Component droplet . In some embodiments , the large diameter synthetic 35 In embodiments that include a first component, the first membrane vesicle (s ) can be multivesicular liposomes. component can be formed by mixing two phases, such as an Some embodiments relate to a process for preparing a organic phase and a first aqueous phase . In some embodi large diameter synthetic membrane vesicle ( s ) composition ments , a therapeutic agent can be added to the organic phase . comprising the steps of, forming a first component by In some embodiments , a therapeutic agent can be added to mixing a first aqueous phase and an organic phase , said 40 the first aqueous phase . In some embodiments , a therapeutic organic phase comprising an organic solvent , at least one agent can be added to both the organic phase and the first amphipathic lipid , and at least one neutral lipid , encapsu - aqueous phase . In some embodiments , the organic phase can lating said first component in a second aqueous phase to include at least one amphipathic lipid , at least one neutral provide a second component, said second component com lipid , and an organic solvent. In some embodiments, the prising an aqueous phase , removing the organic solvent from 45 therapeutic agent can be in the form of a pharmaceutically the second component to form a composition of large acceptable salt . diameter synthetic membrane vesicle particles , wherein the In some embodiments , the mixing of the two phases can removing can be accomplished by using a solvent removal be accomplished using ultrasound . In some embodiments , chamber as disclosed and described herein , and optionally the mixing of the two phases can be accomplished using filtering the composition by particle concentration . Such 50 high pressure emulsification . Such emulsification utilizes an steps may be combined with other steps . In some embodi- atomizing nozzle as disclosed and described herein . In some ments the lipid phase can include cholesterol. In some embodiments , the mixing of the two phases can be accom embodiments , the organic solvent can be a volatile water - plished using mechanical processes including using high immiscible or sparingly miscible solvent. In some embodi shear type devices, rotor/ stator and homogenizers, shear ments , the first component can be a first emulsion . In some 55 type mixer , static mixer , impeller , porous pipe , any of the embodiments , the second component can be a second emul- disclosed mechanical processes optionally in combination sion . In some embodiments , the second component can be a with a heat exchanger , or other processes known to produce droplet. In some embodiments , the large diameter synthetic water - in -oil emulsions. In some embodiments, the mixing of membrane vesicle ( s ) can be multivesicular liposomes. the two phases can be accomplished using a combination of Some embodiments relate to a process for preparing a 60 ultrasound and high pressure emulsification . In some large diameter synthetic membrane vesicle ( s ) composition embodiments , the mixing of the two phases can be accom comprising the steps of, forming a first component by plished using a combination of mechanical processes per mixing a first aqueous phase and an organic phase , said formed by a device selected from the group consisting of organic phase comprising an organic solvent , at least one high - shear type devices , rotor/ stator mixers and homogeniz amphipathic lipid , and at least one neutral lipid , encapsu - 65 ers , shear- type mixer, static mixer, impeller, porous pipe , lating said first component in a second aqueous phase to high energy vibration , injection into a high velocity liquid provide a second component using an atomizing nozzle as stream such as in an aspirator, and the like . US 9 , 730 , 892 B2 27 28 In some embodiments , the first component can comprise 100 mM to about 300 mM . For example , the concentration particles having an average diameter in the range from about of phosphoric acid can be 200 mM . 0 . 1 um to about 100 um . For example , the particles can have In some embodiments , the organic phase can include an an average diameter of at least about 0 . 1 um , 1 um , 5 um , 10 amphipathic lipid . In some embodiments , the amphipathic um , 15 um , 20 um , 50 um , or 100 um , or a diameter within 5 lipid can be selected from the group consisting of soya a range defined by any of two of the preceding values. In lecithin , 1 , 2 - dioleoyl- sn - glycero - 3 - phosphocholine , 1 , 2 - di some embodiments , the particles can have an average diam lauroyl- sn - glycero - 3 -phosphocholine , 1, 2 -dimyristoyl - sn eter in the range from about 0 . 2 um to about 100 um , from glycero - 3 -phosphocholine , 1 , 2 - dipalmitoyl- sn about 0 . 2 um to about 50 um , from about 0 . 5 um to about 30 glycero - 3 -phosphocholine , 1 , 2 -distearoyl - sn - glycero - 3 um , from about 0 . 5 um to about 10 um , from about 10 um 10 to about 50 um , from about 15 um to about 45 um , or from phosphocholine , 1, 2 -diarachidoyl - sn -glycero - 3 -phospho about 20 um to about 40 um . In a typical embodiment, the choline , 1, 2 -dibehenoyl - sn - glycero - 3 -phosphocholine , 1, 2 particles can have an average diameter in the range of from dipalmitoleoyl- sn - glycero - 3 - phosphocholine, 1 , 2 about 0 . 5 um to about 3 um . dieicosenoyl- sn - glycero -3 - phosphocholine , 1, 2 -dierucoyl ente in be formed 15 sn - glycero - 3 - phosphocholine, 1 , 2 - dipalmitoyl- sn - glycero at a temperature in the range from about - 5° C . to about 990 3 -phosphoglycerol , 1, 2 -dioleoyl - sn -glycero - 3 C ., from about 0° C . to about 60° C . , from about 2° C . to phosphoglycerol, and mixture thereof. In a typical about 40° C . , from about 4° C . to about 20° C . , from about embodiment , the amphipathic lipid can be selected from the 5° C . to about 50° C ., from about 10° C . to about 40° C ., or group consisting of 1 , 2 - dierucoyl -sn - glycero - 3 - phospho from about 15° C . to about 35° C ., or from about 10° C . to 20 choline (DEPC ) , 1 ,2 -dipalmitoyl - sn -glycero - 3 -phospho about 20° C . glycerol ( DPPG ) and mixtures thereof. For example , the In some embodiments , the organic solvent can be selected amphipathic lipid can be a mixture of 1 , 2 - dierucoyl- sn from the group consisting of diethyl ether, tert -butylmethyl glycero - 3 - phosphocholine ( DEPC ) and 1 , 2 - dipalmitoyl- sn ether, tetrahydrofuran , sevoflurane, desflurane , isoflurane , glycero - 3 - phosphoglycerol (DPPG ) . In some embodiments and enflurane. In some embodiments , the organic solvent 25 the ratio of 1 , 2 - dierucoyl- sn -glycero - 3 - phosphocholine can be selected from the group consisting of halothane , (DEPC ) to 1 , 2 -dipalmitoyl - sn - glycero - 3 - phosphoglycerol chloroform , and dichloromethane . In some embodiments , (DPPG ) can be in the range from about 100 : 1 to about 1 : 10 , the organic solvent can be selected from the group consisting from about 50 : 1 to about 1 :1 , from about 25 : 1 to about 2: 1 , of ethyl acetate , hexane , hexanes, cyclohexane , pentane , from about 15 : 1 to about 10 : 1 , from about 10 : 1 to about cyclopentane , petroleum ether, and toluene. In some 30 30 : 1 , or from about 15: 1 to about 20 : 1 . For example , the embodiments , the organic solvent can be selected from the ratio of 1 , 2 - dierucoyl- sn - glycero - 3 -phosphocholine (DEPC ) group consisting of freons, chlorofluorocarbons (CFCs ) and to 1 , 2 - dipalmitoyl- sn - glycero - 3 - phosphoglycerol (DPPG ) hydrochlorofluorocarbons (HCFCs ) with boiling points can be about 16 . 8 : 1 . above 15º C . In a typical embodiment, the organic solvent In some embodiments , the organic phase can include a can be selected from the group consisting of chloroform , and 35 neutral lipid . In some embodiments , the neutral lipid can be dichloromethane . For example, the organic solvent can be selected from the group consisting of triolein , tripalmitolein , dichloromethane . trimyristolein trilinolein , tributyrin , tricaprylin , tricaproin , In some embodiments , the first aqueous phase can be and tricaprin , and mixture thereof . In a typical embodiment , selected from the group consisting ofwater solutions includ the neutral lipid can be selected from the group consisting of ing one or more components selected from the group con - 40 tricaprylin , tricaproin , and tricaprin , and mixtures thereof. sisting of a therapeutic agent, dextrose , lysine , dextrose ! For example, the neutral lipid can be tricaprylin . In some lysine , sodium chloride , hydrochloric acid , phosphoric acid , embodiments the ratio of amphipathic lipid to neutral lipid an osmotic pressure adjusting agent such as a sugar, dex - can be in the range from about 50 : 1 to about 1 : 5 , from about trose , sucrose , trehalose , fructose, sorbitan , glycerol, or 25 : 1 to about 1 : 2 , from about 15 : 1 to about 1 : 1 , from about manitol, a therapeutic agent solubility enhancer, and pH 45 10 : 1 to about 2 : 1 or from about 6 : 1 to about 3 : 1 . For modifying agents such as sodium hydroxide , arginine , his - example , the ratio of amphipathic lipid to neutral lipid can tidine, sodium borate , acids, bases, (hydroxymethyl ) amin - be about 4 . 4 : 1 . omethane , or Good ’s buffers . In some embodiments , the In some embodiments , the organic phase can include concentration of any one component in the aqueous phase , cholesterol. In some embodiments the ratio of amphipathic other than the therapeutic agent , can be in the range from 50 lipid to cholesterol can be in the range from about 50 : 1 to about 1 uM to about 1 M , from about 1 mM to about 500 about 1 : 10 , from about 25 : 1 to about 1 : 5 , from about 10 : 1 to mM , from about 10 mM to about 400 mM , from about 100 about 1 : 2 , from about 5 : 1 to about 1 : 1 or from about 3 : 1 to mM to about 300 mM . In a typical embodiment, any one about 1 . 5 : 1 . For example , the ratio of amphipathic lipid to therapeutic agent component if used in the first aqueous cholesterol can be about 1 . 8 : 1. In some embodiments the phase can be in the range from about 1 mM to about 1 M . 55 ratio of cholesterol to neutral lipid can be in the range from For example , the therapeutic agent component can be about 50 : 1 to about 1 : 10 , from about 25 : 1 to about 1 : 5 , from approximately 200 mM . In a typical embodiment, the con - about 10 : 1 to about 1 : 2 , from about 5 : 1 to about 1 : 1 or from centration of any one component in the first aqueous phase about 3 : 1 to about 2 : 1 . For example , the ratio of cholesterol can be in the range from about 100 mM to about 300 mm . to neutral lipid can be about 2 . 4 : 1 . In a typical embodiment, For example , the concentration can be 200 mM . In some 60 any one therapeutic agent component if used in the organic embodiments , the first aqueous phase can include phos - phase can be in the range from about 1 mM to about 1 M . phoric acid as a component. In some embodiments , the For example , the therapeutic agent component can be concentration of phosphoric acid can be in the range from approximately 200 mM . about 1 uM to about 1 M , from about 10 uM to about 750 In some embodiments , the aqueous phase or organic mM , from about 1 mM to about 500 mM or from about 10 65 phase can comprise the therapeutic agent . In a typical mM to about 250 mM . In a typical embodiment, the con - embodiment, the therapeutic agent can be selected from the centration of phosphoric acid can be in the range from about group including semisynthetic aminoglycoside antibiotics US 9 , 730 , 892 B2 29 30 such as amikacin ; antidiabetics ; peptides such as insulin ; ous phase can be in the range of from about 2 : 1 to 1 : 2 . For antitumor drugs such as paclitaxel; antineoplastics including example , the volume : volume ratio of first component to cytarabine, 5 - fluorouracil and floxuridine ; alkaloid opiate second aqueous phase can be about 1 : 1 . In some embodi analgesics including morphine and hydromorphine ; local ments , the first component can be an emulsion . anesthetics including bupivacaine ; synthetic anti- inflamma - 5 In some embodiments , the second aqueous phase can be tory adrenocortical steroids including dexamethasone ; anti - selected from the group consisting of water solutions of, a metabolites including methotrexate ; glycopeptide antibiot- therapeutic agent, or pharmaceutically acceptable salt ics including bleomycin ; vincaleukoblastines and thereof, dextrose , lysine, dextrose /lysine , sodium chloride , stathmokinetic oncolytic agents including vincristine and hydrochloric acid , phosphoric acid , an osmotic pressure vinblastine ; hormones , plasma proteins, cytokines , growth 10 adjusting agent such as a sugar, dextrose , sucrose , trehalose , factors , DNA and RNA from a variety of organisms, and fructose , sorbitan , glycerol, or manitol , a therapeutic agent antisense oligonucleotides . In some embodiments , the thera - solubility enhancer , pH modifying agents such as sodium peutic agent can be an amide anesthetic . In some embodi- hydroxide , arginine , histidine , sodium borate , acids , bases , ments, the therapeutic agent can be selected from the group (hydroxymethyl ) aminomethane , or Good ’ s buffers, and consisting of bupivacaine , mepivacaine , ropivacaine, lido - 15 mixtures thereof . In some embodiments , the concentration caine , pyrrocaine , prilocaine , their stereoisomers , and com - of any one component in the second aqueous phase can be binations thereof. In a typical embodiment, the therapeutic in the range from about 1 uM to about 1 M , from about 10 agent can be bupivacaine or a therapeutically acceptable salt uM to about 750 mM , from about 100 uM to about 500 mM , thereof. For example, bupivacaine can be a free base . In 1 mM to about 250 mM , from about 10 mM to about 150 some embodiments , the aqueous phase can comprise an acid 20 mM , from about 50 mM to about 125 mM , from about 100 in sufficient quantity to maintain the bupivacaine or a mM to about 200 mM , from about 100 mM to about 500 therapeutically or pharmaceutically acceptable salt thereof mM , or from about 200 mM to about 400 mM . In a typical in the first aqueous phase . embodiment, the concentration of any one component in the In some embodiments , the first component can be formed second aqueous phase can be in the range from about 50 mM by mixing two phases. In some embodiments , the first 25 to about 270 mM . component can be an emulsion . In some embodiments, the In some embodiments , the second aqueous phase can be first component can be in the form of droplets . In some an aqueous dextrose / lysine solution . In some embodiments , embodiments , the first component droplets can be formed by the concentration of dextrose can be in the range from about mixing two phases, such as an organic phase and a first 1 mM to about 500 mM from about 10 mM to about 300 aqueous phase where the speed of mixing can control the 30 mM , from about 25 mM to about 200 mM or from about 50 size of the first component droplets . In some embodiments , mM to about 150 mM . In a typical embodiment, the con the size of the first component droplets can have an average centration of dextrose can be in the range from about 60 mM diameter of at least about 0 . 1 um , 1 um , 5 um , 10 um , 15 um , to about 90 mM . For example , the concentration of dextrose 20 um , 50 um , or 1000 um , or a diameter within a range can be about 80 mM . In some embodiments , the concentra defined by any of two of the preceding values . In some 35 tion of lysine can be in the range from about 1 mM to about embodiments , the average first component droplet size can 750 mM , from about 10 mM to about 500 mM , from about be preferably between 0 . 5 um and 2 um . For example , the 50 mM to about 400 mM or from about 100 mM to about first component droplet size can be approximately 1 um . In 200 mM . In a typical embodiment, the concentration of some embodiments , the first component droplet can be an lysine can be in the range from about 150 mM to about 250 emulsion droplet . 40 mM . For example , the concentration of lysine can be about Second Component Droplet 200 mM . In some embodiments , the first component can then be In some embodiments, a first component can be mixed combined with a second aqueous phase to provide a second with a second organic phase comprising one or more second component droplet. The second component droplet can be amphipathic lipids prior to combination with a second formed by combining the first component with the second 45 aqueous phase to provide a second component . In some aqueous phase using a three - fluid nozzle to form a first emembodiments , the second component can be in the form of component/ second aqueous phase mixture . In some embodi droplets . For example , the second organic phase can include ments , the first fluid applied to the nozzle is a first liquid , the same or different amphipathic lipids than in the first made up of a first component, the second fluid applied to the component. The second component droplet can be formed nozzle is a second liquid made up of a second aqueous 50 by combining the first component with the second aqueous phase , and the third fluid can be considered to be a gas (or phase using a three fluid nozzle with an additional inlet for a gas/ vapor mixture ) , such as nitrogen gas ( or a nitrogen the second organic phase . The second organic phase can gas /aqueous vapor mixture ) or air scrubbed of CO2, or air alternatively be pumped into the conduit feeding first com free of, or substantially free of CO2. In some embodiments , ponent to the three fluid nozzle with an optional static mixer contacting the first component/ second aqueous phase mix - 55 in the conduit between the point of second organic phase ture with the third fluid creates the second component addition and the three fluid nozzle . Some active agents can droplets as the gas acts to shear the first emulstion /second interact with the lipids ( e . g . peptides or proteins, have seen aqueous phase mixture into droplets . In some embodiments , this already , especially the charged lipids like PG , e . g . the volume: volume ratio of first component to second aque - DPPG ). For this and other reasons e . g . biocompatibility , ous phase can be in the range of from about 1 : 1000 to about 60 storage stability , vibration stability or release rate modifi 1000 : 1 , in the range of from about 1 :500 to about 500 : 1 , in cation , it may be desirable to have the outside layer of the range of from about 1 : 50 to about 50 : 1 , in the range of phospholipids composition different from the inside cham from about 1: 10 to about 5 : 1, or in the range of from about ber wall composition ( e .g . PG only on the outside where it 1 : 5 to about 5 : 1 . In a typical embodiment, the volume: gives charge stability to theMVL but not present during high volume ratio of first component to second aqueous phase can 65 shear mixing with the active agent first aqueous ). In some be in the range of from about 1 : 3 to about 3 : 1 . Alternately, embodiments , the outside layer of phospholipid is put on the the volume: volume ratio of first component to second aque - particle when the first component contacts the second aque US 9 , 730 , 892 B2 31 32 ous phase , inside the nozzle and is subsequently atomized medium with a new suspending medium , performed in one and it comes from the phospholipids left dissolved in the step or incrementally . These two processes are closely solvent after the first emulsion is made . Phospholipids with related as exchanging the suspending medium can be the desired outside composition , dissolved in solvent, can be accomplished by concentrating the suspension and adding injected into the first fluid conduit , just before the 3 fluid 5 new suspending medium . These terms relate to concentrat nozzle , and just before an optional static or other mixer so ing the particle suspension and exchanging the suspending that these lipids are present and dissolved in the solvent medium to be done separately or simultaneously . In some embodiments, the large diameter synthetic membrane phase of the first component . These phospholipids would vesicles can be multivesicular liposomes. only contact active containing first component droplets for+ 10 In one embodiment , the continuous aqueous solution can seconds under mild shear condition and would thus not 10 be modified / exchanged by diafiltration or cross - flow filtra interact appreciably with the outer layer lipids. Some of tion . The diafiltration can achieve several objectives, includ these added lipids may get incorporated between the cham ing : exchanging an aqueous solution by an isotonic solution , bers of the forming MVL which the 4 fluid nozzle avoids . concentrating of the large diameter synthetic membrane In some embodiments , the one or more second amphipup -- 15 vesicles particles, removing unencapsulated drug, and athic lipids in the second organic phase can be selected from removing of residual organic solvent. In some embodiments , the group consisting of soya lecithin , 1 , 2 -dioleoyl - sn - the large diameter synthetic membrane vesicles can be glycero - 3 - phosphocholine , 1 , 2 - dilauroyl- sn - glycero - 3 - multivesicular liposomes. phosphocholine, 1, 2 -dimyristoyl - sn - glycero - 3 -phosphocho - Particle concentration , such as through diafiltration , can line , 1, 2 -dipalmitoyl - sn -glycero - 3- phosphocholine , 1 ,2 - 20 be a method employed for the filtration , purification , and distearoyl- sn - glycero - 3 -phosphocholine , 1 , 2 -diarachidoyl separation of the large diameter synthetic membrane sn - glycero - 3 - phosphocholine , 1 , 2 - dibehenoyl- sn vesicles particles from complex mixtures by virtue of the glycero - 3 -phosphocholine , 1, 2 -dipalmitoleoyl - sn -glycero physical characteristics of the large diameter synthetic mem 3 -phosphocholine , 1, 2 -dieicosenoyl - sn - glycero - 3 -phospho - brane vesicles particles. For example , the most common choline, 1 , 2 -dierucoyl - sn - glycero - 3 - phosphocholine , 1 , 2 - di- 25 physical characteristic used is size . This filtration can palmitoyl -sn - glycero - 3 -phosphoglycerol , 1 , 2 - dioleoyl- sn - involve cross - flow filtration , as opposed to dead - end filtra glycero - 3 -phosphoglycerol , and mixture thereof. In a typical tion . In cross - flow filtration , a suspension can be circulated embodiment, the amphipathic lipid can be selected from the under pressure and in contact with a filter , so that a permeate group consisting of 1 , 2 - dierucoyl- sn - glycero - 3 -phospho - ( the material which passes through the filter leaves the choline (DEPC ) , 1 , 2 -dipalmitoyl - sn - glycero - 3 - phospho - 30 system , and a retentate ( the material which does not pass glycerol ( DPPG ) and mixtures thereof. through the filter) is left behind and exits the filter housing Solvent Removal through a port different than the permeate and can be In some embodiments , the organic solvent, such as meth - recirculated through the filter . The suspension then becomes ylene chloride , can be nearly or completely removed from a concentrated in material that does not pass through the filter. droplet by further contacting the droplet with a gas ( or a gas 35 In processes in which a first solution is to be exchanged for with some degree of aqueous phase vapor ), such as nitrogen a second solution , the second solution is introduced on the gas, or air scrubbed of CO2, or air substantially free of CO2 retentate side of the filter , until the permeate gradually in a solvent removal chamber. As described below , in some consists of the second solution . By this time, the first embodiments , the gas can have 50 % to 100 % relative solution has been flushed from the suspension . An additional humidity . For example , the humidity can be 100 % . In some 40 consequence of suspending medium exchange is to further embodiments , the droplet can remain suspended in the gas remove any residual organic solvent, such as methylene within the solvent removal chamber until nearly all of the chloride which has a small but appreciable water solubility organic solvent is removed thereby creating a large diameter and is thus removed in the permeate stream . Likewise the synthetic membrane vesicle particle . In some embodiments , therapeutic agent released during the large diameter syn the solvent removal can be done in a solvent removal 45 thetic membrane vesicles formation can also be removed in chamber as discussed herein . The large diameter synthetic the permeate . In some embodiments, the large diameter membrane vesicle particle can then be collected along with synthetic membrane vesicles can be multivesicular lipo multiple other large diameter synthetic membrane vesicle somes . particles , forming large diameter synthetic membrane In some embodiments , aseptic methods can be included vesicles suspended in a continuous aqueous phase . In some 50 with any of the described methods and apparatus described embodiments , the large diameter synthetic membrane herein . vesicles suspended in a continuous aqueous phase can Although sterilization of a final filled container as a undergo further processing . In some embodiments , the large dosage form is often a preferred process in pharmaceutical diameter synthetic membrane vesicles can be multivesicular manufacture for the assurance of minimal risk of microbial liposomes . 55 contamination in a lot, the large diameter synthetic mem Particle Concentration brane vesicles manufactured in the present processes can be Optionally , the large diameter synthetic membrane susceptible to unacceptable damage when subjected to some vesicles suspended in the continuous aqueous solution can terminal sterilization techniques, such as autoclaving and be processed to modify / exchange the continuous aqueous gamma irradiation . In the absence of validated , non - dam solution via a particle concentration system . In this docu - 60 aging terminal sterilization , some embodiments of the ment the term concentration unit, concentration apparatus , invention utilize aseptic techniques, in which the product is concentration system , particle - concentration system , par prepared according to a carefully designed series of aseptic ticle - concentrating device , and particle concentrator are steps . These are designed to prevent the introduction of meant to encompass units and processes that remove some viable microorganisms into components, where sterile , or of the particle suspending medium of a particle suspension 65 once an intermediate process has rendered the bulk product and therefore concentrating the particle concentration as or its components free from viable microorganisms. Prod well as encompassing the exchange of the suspending ucts defined as aseptically processed can consist of compo US 9 , 730 , 892 B2 33 34 nents that have been sterilized by aseptic means. For components ( e . g . valves , steam lines , condensate drains) to example , bulk products which are filterable liquids, can be facilitate sterilization by steaming - in - place. sterilized by filtration . Final empty container components Sterility testing of product lots can be carried out directly can be sterilized by heat ; dry heat for glass vials and after the lot is manufactured as a final product quality control autoclaving for rubber seal components . The requirements 5 test . Testing can be done in accordance with various proce for properly designed , validated and maintained filling and dures found in the U . S . Pharmacopeia ( U . S . P .) and FDA processing facilities are directed to : an air environment free regulations. from viable microorganisms, and designed to permit effec Sterile filtration of all fluids which enter the manufactur tive maintenance of air supply units ; training of personnel ing system is essential for an aseptic process , as envisioned who are adequately equipped and gowned . Available pub - 10 in certain embodiments of the present application . Rating of lished standards for controlled work areas include: Federal pore sizes of filter membranes is by a nominal rating Standard No. 209B , Clean Room and Work Station Require - reflecting the capacity of the membrane to retain microor ments for a Controlled Environment, Apr. 24 , 1973 ; NASA ganisms of size represented by specified trains , not by Standard for Clean Room and Work Stations for Microbially determination of an average pore size and statement of Controlled Environment, publication NHB5340 . 2 , August 15 distribution of sizes. Sterilizing filter membranes include 1967 ; and Contamination Control of Aerospace Facilities , those capable of retaining 100 % of a culture of 10 ' organ U . S . Air Force , T . 0 . 00 - 25 - 203 , Dec . 1 , 1972 , change 1 , isms of a strain of Pseudomonas diminua (ATTC 19146 ) per Oct. 1 , 1974 . In some embodiments, the large diameter square cm of membrane surface under a pressure of not less synthetic membrane vesicles can be multivesicular lipo - than 30 psi . Such filter membranes can be nominally rated somes . 20 0 . 22 um or 0 . 2 um , depending on the manufacturer . Bacterial In aseptic processing , one of the most important labora - filter membranes capable of retaining only larger microor tory controls is the establishment of an environmental moni ganisms ( including Serratia Marcescens (ATTC 14756 ) ) are toring program . Samples can be collected from areas in labeled with a nominal rating of 0 .45 um . Filter membranes which components and product are exposed to the environ used in the present processes can be of the 0 . 2 um type , and ment, including mixing rooms and component preparation 25 can be used in all lines feeding from liquid solution and gas areas . Microbiological quality of aseptic processing areas is storage tanks to vessels and transfer lines used in the monitored to determine whether or not aseptic conditions are methods disclosed herein . maintained during filling and closing activities . Routine In some embodiments the process apparatus is sterilized sampling and testing of the room air , floors , walls , and before use and isolated from the environment by the use of equipment surfaces is carried out. This program establishes 30 sterilizing filters on all apparatus inputs and outputs where the effectiveness of cleaning and sanitizing equipment and the apparatus includes the sterilized product vessel. In this product contact surfaces, and ensures that potential contami embodiment sterile product can be produced in an aseptic nants are held to an acceptable level . The disinfectants are fashion , in a non sterile environment. checked to assure that their efficacy against normal micro - Description of the Instant Devices and their Methods of Use bial flora is maintained . Sampling schedules , including 35 The present embodiments will now be described in more locations and frequency of sampling , are maintained . Pas - detail in terms of features and operations with reference to sive air samplers such as settling plates ( Petri dishes) are the accompanying drawings. employed as well . FIG . 1A Aseptic assembly operations can be validated by the use An embodiment of the present application is a continu of a microbiological growth nutrient medium to simulate 40 ous- flow system for manufacturing pharmaceutical formu sterile product filling operations, known as " sterile media lations . An example of such a continuous - flow system is fills . " The nutrient medium can be manipulated and exposed presented in FIG . 1A . FIG . 1A is a schematic diagram of the to the operators , equipment , surfaces and environmental significant components and sub - systems used in a system for conditions to closely simulate the same exposure which the manufacturing pharmaceutical formulations . Each of the product itself will undergo . The sealed drug product con - 45 components and sub -systems can be included in , and oper tainers filled with the media can then be incubated to detect ated as, a part of the larger manufacturing system or may , in microbiological growth and the results are assessed to the alternative , be run autonomously in a continuous fashion determine the probability that any given unit of drug product to accomplish the objective of each sub -system as described may become contaminated during actual filling and closing herein . Additionally , each sub -system may be run in a operations . Media filling, in conjunction with comprehen - 50 continuous - flow manner using batch inputs and producing a sive environmental monitoring can be particularly valuable batch output. in validating the aseptic processing of sterile solutions, In one embodiment , the manufacturing system 100 is suspensions , and powders . Filling liquid media , as part of comprised of a tank 5 , which can hold a first fluid , the first validating the processing of powders , may necessitate use of fluid can be pumped by a positive -displacement pump 2 equipment and / or processing steps that would otherwise not 55 through a hydrophilic sterilizing filter 15 and into a high be attendant to routine powder operations . shear mixer 25 . In some embodiments , the first fluid can be In some embodiments , clean - in - place (CIP ) and sterilize - an aqueous solution . In some embodiments , the first fluid in - place (SIP ) procedures can be utilized by methods gen can be a first liquid . In some embodiments , the first liquid erally known in the art . Some embodiments include moni- can be an aqueous solution . Similarly , tank 10 can feed a toring of the temperature at the steam traps . According to 60 second fluid , via a positive - displacement pump 12 , through this procedure , as steam is admitted into the vessels and fill a hydrophobic sterilizing filter 20 into the high shear mixer lines to effect sterilization , the temperature at the outlet 25 . In some embodiments , the second fluid can be a second points is monitored until bacterial kill is assured . At this liquid . In some embodiments , the second liquid can be an point, the seals can be closed , and the system can be organic solvent. The high shear mixer 25 , a heat exchanger sterilized for further use . Subsequently, the equipment can 65 30 , and associated inlet line 96 and outlet line 97 comprise be used aseptically in a non - sterile room environment. The one embodiment of a first sub - system . In some embodi systems described herein , may also comprise additional ments , the first sub -system can be a first emulsification US 9 ,730 ,892 B2 35 36 sub - system . In some embodiments , the high shear mixer 25 one or more atomizing nozzles , used in parallel to evaporate can create a first dispersion of aqueous particles ( the “ dis the solvent from the atomized droplets . In some embodi continuous phase ” ) suspended in an organic continuous ments , the evaporation sub - system can be comprised of a phase . In some embodiments , the first dispersion can be a plurality of solvent removal vessels , such as solvent removal first component. In some embodiments , a majority of the 5 vessel 50 , each with one or more atomizing nozzles , such as first component can then be fed from the high shear mixer a tomizing nozzle 75 , used in parallel to evaporate solvent 25 into the heat exchanger 30 , and the cooled first compo from the atomized droplets . In some embodiments, the nent can flow back to the high shear mixer 25 . In one solvent removal vessel can have additional multiple atom embodiment, however, a portion of the emulsion leaving the izing nozzles (not shown ) such as 75 . high shear mixer 25 can be forced towards a nozzle 75 , for 10 In some embodiments , a portion of the gas passing example by the addition of additional organic solvent and through the heater /humidifier 90 can be directed towards a aqueous solution to the first emulsification sub - system . gas inlet 110 located in the lid of the solvent removal vessel In some embodiments , the emulsion leaving the high 50 . In some embodiments , the gas inlet 110 allows gas to shear mixer 25 can be transferred to an evaporation appa - enter the solvent removal vessel 50 and circulate in the upper ratus or evaporation sub -system . In one embodiment the 15 portion of the vessel acting to prevent particle buildup on the evaporation apparatus, or evaporation sub -system , is com lid . prised of a solvent removal vessel 50 , the atomizing nozzle In some embodiments , a two - fluid rinse nozzle 105 can be 75 , a gas inlet 115 , and a gas outlet 80 . In some embodi placed in and through the lid of the solvent removal vessel ments , the nozzle 75 acts to spray atomized droplets of the 50 . In some embodiments , the rinse nozzle 105 can spray the first component into the solvent removal vessel 50 . In some 20 wall of solvent removal vessel 50 . The rinse nozzle 105 , embodiments , the nozzle 75 can combine the first compo - which receives a buffer solution from a buffer solution tank nent with a fluid from a tank 60 and spray the first compo - 66 , through a pump 64 and a sterilizing filter 62 , can spray nent/ liquid mixture into the solvent removal vessel 50 as atomized tank wall rinse solution particles into the solvent atomized droplets . In some embodiments , the fluid from the removal vessel 50 . The buffer solution can be atomized by tank 60 is a liquid . In some embodiments , the liquid from the 25 gas traveling into the nozzle 105 through a pressure regu tank 60 is a buffer solution . In some embodiments , the buffer lator 21 and a sterilizing gas filtration system 85 . Spraying solution can be fed by a positive - displacement pump 22 atomized wall rinse solution particles into the solvent from the tank 60 through a hydrophilic sterile filter 65 into removal vessel 50 can act to prevent the large diameter the nozzle 75 . Gas from the gas supply can be passed synthetic membrane vesicles droplets ( FIG . 7 , component through a pressure regulator 11 and a sterilizing gas filtration 30 7380 ) from sticking to the walls of the solvent removal system 35 before entering the nozzle 75 . In some embodi- vessel 50 by rinsing or flushing particles from the internal ments , the gas acts to atomize the first component/ liquid surfaces of the vessel 50 , and out a drain port 130 on the mixture as it exits the nozzle 75 into the solvent removal bottom of the vessel 50 . vessel 50 . Removal of the solvent from the atomized droplets , In some embodiments , a carrier gas can be supplied to the 35 affords large diameter synthetic membrane vesicles coated solvent removal vessel 50 from the gas supply and enter the in a buffer solution shell as droplets . In some embodiments , solvent removal vessel through the gas inlet 115 . The carrier the large diameter synthetic membrane vesicles can be gas can be, for example , nitrogen gas ( or a nitrogen gas / multivesicular liposomes . These droplets then can collect on aqueous vapor mixture ) or air scrubbed of CO , . The pres - the bottom of the solvent removal vessel 50 to form a sure of the carrier gas can be regulated using a pressure 40 suspension of large diameter synthetic membrane vesicles in regulator 31 . In some embodiments , the carrier gas first a buffer solution . This large diameter synthetic membrane passes through a heater /humidifier 90 and then through a gas vesicles suspension can subsequently be pumped by a pump filtration system 45 before entering the solvent removal 125 through a solvent removal vessel outlet line 120 to an vessel 50 through the gas inlet 115 . In some embodiments , optional sub - system 70 . In some embodiments , the large the carrier gas circulates through the solvent removal vessel 45 diameter synthetic membrane vesicles suspension can be 50 , and creates an intense gas vortex , which can facilitate pumped by a pump 125 through a continuous heat treatment solvent evaporation from the atomized droplets as they are system 150 before entering the option sub -system 70 . In sprayed into the solvent removal vessel 50 from the nozzle some embodiments , the buffer solution can be exchanged 75 . The gas vortex can substantially prevent the atomized for, for example , a saline solution . In some embodiments , droplets from exiting the solvent removal chamber with the 50 the sub - system 70 can be a single or series of concentration carrier gas through gas outlet 80 . With evaporation of the units running in batch or continuous mode . In a typical solvent, the atomized droplets become droplets of large embodiment, the concentration units can be run in continu diameter synthetic membrane vesicles within an aqueous ous mode . In a typical embodiment, large diameter synthetic phase . In some embodiments , the atomized droplets can be membrane vesicles can be concentrated and the resulting first component/ liquid mixture droplets . In some embodi- 55 suspension collected by the process . In some embodiments , ments , the large diameter synthetic membrane vesicles can the large diameter synthetic membrane vesicles can be be within an aqueous phase . In some embodiments , the large multivesicular liposomes . diameter synthetic membrane vesicles can be multivesicular In some embodiments, one or more components of the liposomes . In some embodiments , the carrier gas and evapo - manufacturing system 100 can be omitted from the system . rated solvent can then be removed from the solvent removal 60 FIG . 1B vessel 50 through the gas outlet 80 , after which they can then In some embodiments , the manufacturing system 100 pass through a filtration system 55 comprised of a pre - filter further comprises a mass flow controller 13 , a mass flow and a sterilizing barrier filter, before being removed as waste controller 23 , a mass flow controller 33 , a rotometer flow 95 . In some embodiments, the prefilter (not shown ) can be indicator 57 , a heater 93 , a steam generator 40 , and a a high efficiency cyclone separator. 65 metering pump 6 as shown in FIG . 1B . Each of the com In one embodiment, the evaporation sub -system is com - ponents and sub - systems can be included in , and operated prised of a plurality of solvent removal vessels , each with as , a part of the larger manufacturing system or may, in the US 9 ,730 ,892 B2 37 38 alternative , be run autonomously in a continuous fashion to the solution first passing through a manual valve 162 and a accomplish the objective of each sub - system as described sterilizing hydrophilic filter 163 , at a rate of 1 . 4 L per 1 L herein . Additionally , each sub -system may be run in a suspension / dextrose mixture added to the retentate vessel continuous- flow manner using batch inputs and producing a 168 . In some embodiments , the solution flows through the batch output. 5 line 164 at a rate of 578 ml/min . In some embodiments , the In some embodiments , the mass flow controllers 13 , 23 , tank 160 is temperature controlled . In some embodiments , and 33 measure , indicate , and control the gas flow supply to the temperature controlled tank is pressurized . In some their associated apparatus . In some embodiments , the rotom - embodiments , the tank 160 is pressurized with the nitrogen eter flow indicator 57 measures and indicates the gas flow to source 157 through a line 158 and a manual valve 159 . In the lid protection nozzle (FIG . 1A 110 7430 . In some 10 some embodiments , the solution is a saline solution . In some embodiments , the steam generator 40 allows precise humidi- embodiments , the saline solution is a cold saline solution . In fication of the carrier gas . In some embodiments , the meter some embodiments , the retentate vessel 168 is temperature ing pump 6 allows precise control of the water vapor controlled or cooled . generated by the steam generator 40 . In some embodiments , a portion of the suspension /dex FIG . 1C 15 trose mixture in the retentate vessel 168 can be pumped by One embodiment of the present application includes a a pump 171 through a cross - flow (tangential - flow ) filtration continuous heat treatment system . FIG . 1C provides a sche module 167 . In some embodiments , the permeate can be matic of one example of a continuous heat treatment system drawn off through a permeate line 173 (passing through a 150. In some embodiments, the large diameter synthetic sterilizing hydrophilic filter 172 and a manual valve 174 ) , membrane vesicles suspension can be pumped from the 20 wherein for each volume of suspension / dextrose mixture solvent removal vessel ( FIG . 1A , component 50 ) flowing added to the retentate vessel 168 , a volume can be removed through a feed line 120 (also seen in FIG . 1A , component and discarded . In some embodiments , the retentate from the 120 ) , to the continuous heat treatment system 150 before filtration module 167 can be circulated back into the reten entering a subsystem ( FIG . 1A , component 70 ) . tate vessel 168 via a retentate line 166 . In some embodi In some embodiments , the continuous heat treatment 25 ments , for each volume of suspension / dextrose mixture system comprises a temperature controlled tank 151 , option - added to the retentate vessel 168 , a volume of the liquid can ally a temperature control jacket (not shown ), a pressurized be removed from the retentate vessel 168 through a feed line tank 160, a holding coil tubing 156 , and a nitrogen source 169 and a metering pump 170 to be further processed by a 157 as shown in FIG . 1C . Each of the components and subsystem ( as seen in FIG . 1A , component 70 ; systems of sub - systems can be included in , and operated as, a part of the 30 FIG . 8 , FIG . 10 and FIG . 11 ) . In some embodiments , the larger manufacturing system or may , in the alternative , be suspension can flow through the pump 170 at a rate of 490 run autonomously in a continuous fashion to accomplish the ml/min . objective of each sub - system as described herein . Addition - Filters 154 , 163 , and 172 are sterilizing hydrophilic filters . ally , each sub -system may be run in a continuous- flow Filters , 161 and 176 are sterilizing hydrophobic gas vent manner using batch inputs and producing a batch output. 35 filters used in the vessels and fed by gas lines 165 and 175 , In some embodiments , a portion of the large diameter respectively . synthetic membrane vesicles suspension can be pumped by FIG . 2 a pump ( FIG . 1A , component 125 ) through the feed line 120 One embodiment of the present application includes an into a mixing vessel 180 , for example an in - line static mixer. emulsification system , the process of using the system , and In some embodiments , the large diameter synthetic mem - 40 the large diameter synthetic membrane vesicles products brane vesicles suspension , flowing through the line 120 , can made by the process . In some embodiments , the large come into contact with a solution from the tank 151 before diameter synthetic membrane vesicles can be multivesicular flowing through a line 155 into the mixing vessel 180 . In one liposomes . FIG . 2 provides a schematic of one example of embodiment, the suspension feeding the mixing vessel 180 a continuous - flow emulsification system 210 . The emulsifi through the line 120 , flows at 165 ml/min and the solution 45 cation system 210 includes a high shear mixer 2130 and a through the line 155 flows at 247 . 5 ml/min , In some embodi- heat exchanger 2170 . In some embodiments , the high shear ments, the solution can be fed through a line 152 by a pump mixer 2130 can be used in preparing a first component. The 153, the solution first passing through a hydrophilic sterile high shear mixer 2130 used in one embodiment is the Ross filter 154 at a rate of 1 . 5 L per 1 L of the large diameter HSM - 703XS - 20 Sanitary Inline High Shear Mixer made by synthetic membrane vesicles suspension added to the mixing 50 the Charles Ross & Son Company of Hauppauge , N . Y . In vessel 180 . In some embodiments , the tank 151 is tempera some embodiments , the head of the high shear mixer 2130 ture controlled . In some embodiments , the solution from the can be connected to an aqueous inlet line 2120 . In some tank 151 is a dextrose solution . In some embodiments, the embodiments , the head of the high shear mixer 2130 can be dextrose solution is heated to about 98° C . in the temperature fed an aqueous solution through the aqueous inlet line 2120 . controlled tank 151 . In some embodiments, the suspension / 55 In some embodiments , the head of the high shearmixer 2130 dextrose mixture flows through the holding coil tubing 156 . can be connected to a recirculation line 2125 . In some In some embodiments , the holding coil tubing 156 holds the embodiments , the high shear mixer 2130 can feed a recir suspension /dextrose mixture for a specified treatment time. culated emulsion through the recirculation line 2125 . The In some embodiments, the treatment time is between 10 recirculated emulsion contains already - sheared aqueous seconds and 30 seconds. In some embodiments, the suspen - 60 droplets dispersed in a continuous organic solution . In some sion / dextrose mixture is continuously heated for 30 seconds embodiments , the aqueous solution can be stored in a tank to a temperature at or above 60 C . In some embodiments, the (FIG . 1A , component 10 ) and pass through a liquid steril holding coil tubing 156 has a volume of 206 ml. ization filter (FIG . 1A , component 20 ) attached to the In some embodiments , the suspension /dextrose mixture aqueous inlet line 2120 before entering the high shear mixer can leave the holding coil tubing 156 to enter a retentate 65 2130 at various volumetric flow rates . High shear mixers are vessel 168 . In some embodiments , a solution can be fed from available to deliver volumetric flow rates between 1 the tank 160 into the retentate vessel 168 through a line 164, mL /minute and 4 ,000 mL/ minute , between 10 mL /minute US 9 ,730 ,892 B2 39 40 and 1 , 000 mL /minute , between 10 mL /minute and 100 to drive a high volumetric flow rate through the heat mL /minute , and preferably between 15 mL/ minute and 50 exchanger and around the recirculation loop , back to the mL /minute . In some embodiments , the aqueous inlet line high shear mixer head 2130 . 2120 can be configured to run co - axially through a portion Attached to the recirculation line 2125 is an organic of the recirculation line 2125 . The range of volumetric flow 5 solution inlet line 2160 , which can be used to replenish the rates above is appropriate when contemplating the use of organic solution in the emulsification system 210 according one , or optionally multiple , atomizing nozzles . to the volumetric amount of organic solution fed in the first component to the nozzle feed line 2180 . In some embodi In some embodiments , the aqueous solution can be ments, the organic solution can be stored in a tank (not injected through the center of a stator inside of the head of 10 shown) and flows, at a volumetric flow rate between 1 the high shear mixer 2130 into the center of a spinning rotor mL /minute and 4 , 000 mL /minute , between 10 mL /minute (not shown ) also inside the high shear mixer head 2130 . As and 1 , 000 mL /minute , between 10 mL /minute and 100 the aqueous phase passes between the rotor and the stator , mL/ minute , and preferably between 20 mL /minute and 50 inside the high shear mixer head 2130 , it is sheared by the mL /minute . The range of volumetric flow rates above is rotor teeth and stator teeth into aqueous droplets .· The 15 appropriate when contemplating the use of one, or option sheared aqueous droplets become part of a flowing stream of ally multiple , atomizing nozzles . In some embodiments, the the recirculated emulsion which itself can be fed back into organic solution flows through a liquid sterilization filter the high shear mixer through the recirculation line 2125 . The (shown in FIG . 1A and FIG . 1B , component 15 ) attached to combined aqueous solution and recirculated emulsion can the organic inlet line 2160 before entering the recirculation pass through blade shear gaps (not shown ) in the high shear 20 line 2125 and the high shear mixer 2130 . In one embodi mixer head 2130 , as it travels from the center to the outside ment, the volumetric flow rate of the organic solution added of the rotor. In one embodiment, on average , aqueous through the inlet line 2160 can be equal to the volumetric droplets are recirculated approximately 100 times through flow rate of aqueous solution added to the high shear mixer the emulsification system 210 , and through that process , the 2130 through the aqueous inlet line 2120 . That is , in one aqueous droplets are sheared roughly 1 , 300 times by the 25 embodiment, the volumetric flow rates of the added liquids blade shear gaps ( not shown ) . are at a 1 : 1 ratio . At steady - state , because these are incom Exiting the high shear mixer head 2130 through an exit pressible fluids and the piping described herein is non line 2150 is the resultant emulsion , with aqueous droplets expandable piping , the flow rate of the first component dispersed in the organic continuous phase , traveling at a traveling to the nozzle through the nozzle feed line 2180 is volumetric flow rate between 1 L /minute and 500 L /minute , 30 equal to the sum of the flow rates of the organic solution between 5 L /minute and 100 L /minute , and preferably being replenished to the emulsification system 210 through between 10 L /minute and 50 L /minute . The major portion of the organic phase inlet line 2160 and the aqueous solution the emulsion exiting the high shear mixer 2130 , approxi- being replenished to the emulsification system 210 through mately between 50 % and 99 . 99 % , between 70 % and 99 . 9 % , the aqueous inlet line 2120 . and preferably between 80 % and 99 . 9 % , is passed through 35 In one embodiment, the emulsification system is used the heat exchanger 2170 to cool the emulsion which has autonomously to make MVL where the first component is been heated by the mechanical shearing process of the high collected in a container and further processed to make the shear mixer 2130 . Additionally , a smaller portion of the second emulsion . The second emulsion is then sparged and recirculating emulsion , approximately between 1 mL /min - filtered in a batch mode as described in PCT Patent Publi ute and 8 , 000 mL/ minute , between 10 mL /minute and 2 ,000 40 cation No . W099 / 13865 to S . Kim et al. , incorporated herein mL /minute , between 20 mL/ minute and 200 mL /minute , and by reference . preferably between 40 mL /minute and 100 mL /minute , In one embodiment, the emulsification system is travels from the high shear mixer 2130 through a nozzle feed employed autonomously to produce common emulsions of line 2180 to a nozzle for use in making the first component/ triglycerides ( vegetable oils ) and surfactants ( Tween - 20 , buffer atomized droplets . The range of volumetric flow rates 45 Pluronic F - 66 , egg lecithin ) and water with common addi above is appropriate when contemplating the use of one , or t ives ( scents , flavors ) . In an additional embodiment, a cos optionally multiple , atomizing nozzles . In some embodi metic lotion is made with a triglyceride as the discontinuous ments , the heat exchanger 2170 can be comprised of a series phase (oil in water emulsion ) . In still other embodiments , the of coils surrounded by circulating solution . In some embodi- emulsification system produces ointments , creams and ments, the circulating solution can be at a temperature 50 salves, with the triglyceride as the continuous phase . Addi ranging from about - 5° C . to about 30° C . In some embodi- tionally , a triglyceride emulsion in water with egg yolk as ments, the circulating solution can be a glycol/ water solu - the surfactant is used in the emulsification system to produce tion . In some embodiments , the glycol/ water solution can be a type of edible mayonnaise . at a temperature ranging from about 0° C . to about 10° C . In FIG . 3A - 6 some embodiments , the heat exchanger 2170 is comprised 55 One embodiment of the present application includes an of a series of coils surrounded by circulating 5° C . glycol) atomizing nozzle , the process of using the nozzle , and the water solution . In some embodiments , the circulating glycol) large diameter synthetic membrane vesicles made by the water solution can be fed to the heat exchanger 2170 through process . In some embodiments , the large diameter synthetic an inlet line 2110 and an outlet line 2105 . In some embodi- membrane vesicles can be multivesicular liposomes . ments , the emulsion travels through the heat exchanger 60 Examples of the instant atomizing nozzles , the processes for coils , being cooled by the glycol/ water solution , and exits using them , as well as the MVL products , are presented in the heat exchanger through a heat exchanger outlet 2175 . In FIG . 3A - 6 and described herein . some embodiments , the heat exchanger outlet 2175 can FIG . 3A carry the cooled emulsion back through the recirculation line FIG . 3A is a schematic partial view of an atomizing 2125 for additional mixing with added aqueous phase and 65 nozzle 310 , providing a cross- sectional view of the lower added organic solution . In some embodiments , the high portion of the atomizing nozzle including a fluid contacting shear mixer 2130 can act as a high volume centrifugal pump chamber 3125 . The fluid contacting chamber 3125 is coni US 9 ,730 ,892 B2 41 cally tapered from the bottom of an inner fluid conduit 3165 be smaller than the ratio of the diameters of the first fluid in to a cylindrical tip 3145 . The fluid contacting chamber 3125 inner fluid conduit 3165 to the second fluid in the outer fluid is annularly surrounded by an annular gas chamber ( a third conduit 3123 . fluid conduit ) 3135 , which narrows to form an annular gas FIG . 3D shows an expanded cross -sectional view of the orifice 3150 . The bottom portion of the cylindrical tip 3145 5 first fluid and second fluid in a droplet 3155 . In some is annularly and concentrically surrounded by the annular embodiments , the ratio of the diameters of the first fluid to gas orifice 3150 . the second fluid in a droplet 3155 can be the same as the ratio In some embodiments , a first fluid 3115 can travel at a of the diameters of the first fluid in the inner fluid conduit volumetric flow rate between 2 mL /minute and 1 ,000 3165 to the second fluid in the outer fluid conduit 3123 . In some embodiments , the ratio of the diameters of the first mL /minute , between 10 mL /minute and 500 mL /minute , fluid to the second fluid in a droplet 3155 can be larger than between 20 mL /minute and 100 mL /minute , and preferably the ratio of the diameters of the first fluid in the inner fluid between 50 mL /minute and 100 mL /minute through an inner conduit 3165 to the second fluid in the outer fluid conduit fluid conduit (central needle ) 3165 . In a typical embodiment, 3123 . In some embodiments , the ratio of the diameters of the the first fluid 3115 is an emulsion . In some embodiments , the 15 first fluid to the second fluid in a droplet 3155 can be smaller first fluid exits the bottom 3163 of the inner fluid conduit than the ratio of the diameters of the first fluid in the inner 3165 and enters the fluid contacting chamber 3125 , where - fluid conduit 3165 to the second fluid in the outer fluid upon the first fluid 3115 comes in physical communication conduit 3123 . with a second fluid 3120 . In a typical embodiment, the The nozzle 310 comprises a gas input channel 3130 which second fluid 3120 is a buffer solution . In some embodiments , 20 supplies a gas 3140 . In some embodiments , the volumetric the second fluid 3120 travels at a volumetric flow rate flow rate of the gas can be between 30 L /minute and 1 , 000 between 2 mL /minute and 1 , 000 mL /minute , between 10 L /minute , between 30 L /minute and 500 L /minute , between mL /minute and 500 mL /minute , between 20 mL /minute and 20 L /minute and 200 L /minute , and preferably between 25 100 mL/ minute , and preferably between 50 mL /minute and L /minute and 100 L /minute and at a pressure between 5 psig 100 mL /minute , through the outer fluid conduit 3123 , which 25 and 1, 000 psig , between 10 psig and 250 psig , between 20 annularly surrounds the inner fluid conduit 3165 ( as seen psig and 150 psig , and preferably between 40 psig and 120 from the downward perspective view B and displayed in psig to an annular gas chamber 3135 . The above atomizing FIG . 3B ) , to reach the fluid contacting chamber 3125 . The nozzle flow rates are for a single atomizing nozzle . Prior to above atomizing nozzle flow rates are for a single atomizing reaching nozzle 310 , the gas 3140 can be passed through a nozzle . In some embodiments , the first fluid 3115 forms a 30 gas filtration system ( not shown ) comprising a hydrophobic cylindrical core traveling through the fluid contacting cham - gas sterilizer ( as seen in FIG . 1A and FIG . 1b , component ber 3125 , now being in physical communication with , and 35 ) . The wall of the fluid contacting chamber 3125 provides annularly surrounded by , the second fluid 3120 . In some a barrier between the annular gas chamber 3135 and the fluid embodiments , the first fluid and the second fluid are immis - contacting chamber 3125 such that there is no physical cible . In some embodiments , the first fluid and second fluid 35 contact between the fluids in the fluid contacting chamber can be sparingly miscible . In some embodiments , the immis - 3125 and the gas 3140 in the annular gas chamber 3135 . In cibility between the second fluid 3120 and the first fluid some embodiments , the outlet of the cylindrical tip 3145 and 3115 , and the velocity at which the second fluid 3120 and the the outlet of the annular gas orifice 3150 can be flush with first fluid 3115 , causes the second fluid 3120 to form a sheath each other such that second fluid 3120 does not come into around the first fluid 3115 core . In a typical embodiment , the 40 contact with the gas 3140 prior to the gas 3140 exiting the first fluid 3115 is an emulsion and the second fluid 3120 is atomizing nozzle 310 . In other embodiments , the cylindrical a buffer solution . tip 3145 extends lower than the annular gas orifice 3150 . In In some embodiments , the diameter of the fluid contacting still other embodiments , the cylindrical tip 3145 is slightly chamber 3125 can be conically narrowed as it joins the recessed within the annular gas orifice 3150 , such that the cylindrical tip 3145 . As the first fluid 3115 travels through 45 gas 3140 comes into physical communication with the the narrowing fluid contacting chamber 3125 and into the second fluid 3120 prior to the second fluid fully exiting from cylindrical tip 3145 , the diameter of the first fluid 3115 core the nozzle 310 . is correspondingly decreased . Likewise, the second fluid In some embodiments , as the gas 3140 passes through the 3120 can be constricted to create a thinner concentric annular gas orifice 3150 and exits the atomizing nozzle 310 , annular sheath around the first fluid 3115 core . As a result of 50 it comes in physical communication with the second fluid decreasing the diameter of the fluid contacting chamber 3120 and acts to shear the stream of the second fluid 3120 3125 and passing the fluids through the cylindrical tip 3145 , and the first fluid 3115 into multiple atomized droplets 3155 the velocities of the first fluid 3115 and second fluid 3120 are made of a first fluid 3115 core and a second fluid 3120 shell . increased . In a typical embodiment, the first fluid 3115 is an This configuration of the three fluid nozzle can be best emulsion and the second fluid 3120 is a buffer solution . 55 achieved when the cross sectional areas of the inner fluid FIG . 3C shows an expanded cross -sectional view of the conduit 3165 and the second fluid path within the fluid first fluid and second fluid in the cylindrical tip 3145 . In contacting chamber 3125 are chosen such that the velocities some embodiments , the ratio of the diameters of the first of the first fluid 3115 and the second fluid 3120 are approxi fluid to the second fluid in the nozzle tip can be the same as mately equal at the exit of the inner conduit 3163 . the ratio of the diameters of the first fluid in inner fluid 60 In some embodiments , the atomizing nozzle 310 can be conduit 3165 to the second fluid in the outer fluid conduit configured to produce very high yield large diameter syn 3123 . In some embodiments , the ratio of the diameters of the thetic membrane vesicles ( that is , vesicles that encapsulate first fluid to the second fluid in the nozzle tip can be larger a high amount of a therapeutic agent ) . In some embodi than the ratio of the diameters of the first fluid in the inner ments , the large diameter synthetic membrane vesicles can fluid conduit 3165 to the second fluid in the outer fluid 65 be unilamilar vesicles or multilamilar vesicles or polymer conduit 3123 . In some embodiments, the ratio of the diam - spheres encasing a liquid comprising a therapeutic agent . In eters of the first fluid to the second fluid in the nozzle tip can some embodiments , the large diameter synthetic membrane US 9 , 730 , 892 B2 43 44 vesicles can be multivesicular liposomes. In some embodi with a high transition temperature as they are deposited from ments , the first fluid 3115 can be an aqueous ( or similar an evaporating solvent solution and do not have to be formed hydrophilic liquid ) phase containing the therapeutic agent in aqueous solutions as per conventional liposome pro and the second fluid 3120 can be an organic phase compris cesses . ing phospholipids or other encasing material e . g . a polymer 5 When the first core fluid 3115 is the first component or PLGA ( poly ( lactic - co - glycolic acid ) , biocompatible and lacking any lipids that could interact with the active agent in - vivo degradable polymer ) or a wax . In some embodi- ( e . g . charged lipid like DPPG ) and 3170 is volatile solvent ments , the gas 3140 combines with an aqueous phase and an could be same or different that used in first component ) and organic phase to afford a droplet of aqueous phase inside a 3120 is a suspending buffer , MVLs with different lipids on droplet of organic phase, which can , for example , be used to 10 the outside. These outside lipids ( from 3170 ) can be highly form unilamilar vesicles or multilamilar vesicles . In some charged for stability or long chain saturated for mechanical embodiments , the size of the inner aqueous phase droplet strength . The lipids can be put on the MVL as 1 or 2 bilayers can have an average diameter of at least about 0 . 5 um , 1 um , or, if the lipid concentration in 3170 is high , be put on as a 5 um , 10 um , 15 um , 20 um , or 50 um , or a diameter within thick mechanically strong many multiple -bilayers (not pre a range defined by any of two of the preceding values. In 15 viously done ) to increase stability of the MVL and provide some embodiments , the organic solvent can be removed by longer slower active agent release . If polymers ( e . g . PLGA ) using a normal spray drying system . In some embodiments , were also added to 3120 ( w /wo lipids ) , a polymer skeleton the organic solvent can be removed by using solvent could be formed surrounding the MVL , further stabilizing it removal chamber such as the one described in FIG . 7 . physically and chemically . If the polymer layer were strong In some embodiments , the nozzle 310 can comprise an 20 enough . it would allow the MVLs to be lyophilized and be additional outer fluid conduit ( not shown ), which annularly stable for very long times at room temperature . surrounds the inner conduit 3165 to provide a four fluid FIG . 3E is a schematic partial cross -sectional view of a atomizing nozzle as shown in FIGS . 3E - 3L . In some four fluid atomizing nozzle 320 , comprising a first inner embodiments , the conduit which annularly surrounds the fluid conduit ( central needle ) 3165 , a second inner conduit inner conduit 3165 can include an aqueous phase suitable as 25 3175 , an outer conduit 3123 , a gas input channel 3130 , an a suspending medium . In some embodiments, the nozzle can annular gas chamber ( a fourth fluid conduit ) 3135 , a first be operated to produce an emulsion where an aqueous drug fluid contacting chamber 3125 , a second fluid contacting core can be surrounded by phospholipids in a volatile chamber 3137 , and an annular gas orifice 3150 . The first solvent that in turn is surrounded by another aqueous phase , fluid contacting chamber 3137 is conically tapered from the which can , for example , be used to form unilamilar vesicles 30 bottom of the first inner fluid conduit 3165 and the exit orfice or multilamilar vesicles . In some embodiments , the large 3147 of the second inner fluid conduit 3175 . The second diameter synthetic membrane vesicles will initially have a fluid contacting chamber 3137 is conically tapered from the hydrophobic outer surface . exit orfice 3147 of the second inner fluid conduit 3175 to a In some embodiments , the nozzle 310 can omit the cylindrical tip 3145 . In some embodiments , the exit orfice conduit 3123 in order to provide a two fluid nozzle ( not 35 3147 of the second inner fluid conduit 3175 can be between shown ). In some embodiments , the two fluids are a liquid the exit orfice of the first inner fluid conduit 3163 and the and a gas. In some embodiments , the two fluid nozzle ( not cylindrical tip 3145 . shown ) can be used to spray a first component into the In some embodiments , the second fluid contacting cham instant solvent removal chamber such as the one described ber, allows for the first and second fluids to contact the third in FIG . 7 . In some embodiments , the organic solvent can be 40 fluid 3120 for a shorter duration than the nozzle described in removed by contacting the first component with an inert gas FIG . 31. In some embodiments , the four fluid atomizing to in order to create the MVL . In some embodiments excess nozzle 320 depicted in FIG . 3E can handle flow rates higher phospholipids can be used in the first component to create than the one fluid contacting chamber nozzle described in the MVL having a coating of phospholipids which initially FIG . 31 through FIG . 3L due to lower flow instabilities. In has a hydrophic surface and can partially shed to afford the 45 some embodiments , the first and second fluid contacting MVL with a hydrophilic surface. chambers are hydraulic extrusion cones . FIG . 3E through FIG . 3L The second fluid contacting chamber 3137 is annularly Liposomes with high incorporation percentages ( espe - surrounded by the annular gas chamber 3135 , which nar cially important for an expensive active agents ) can be made rows to form an annular gas orifice 3150 . The exit orfice with a four fluid atomizing nozzle 320 as depicted in FIG . 50 portion of the cylindrical tip 3145 is annularly and concen 3E through FIG . 3L . In some embodiments , a first fluid 3115 trically surrounded by the annular gas orifice 3150 . can be a fluid comprising an aqueous active agent, while a In some embodiments , a first fluid 3115 can travel at a second fluid 3170 can be a volatile solvent comprising a volumetric flow rate between 2 mL /minute and 1 ,000 lipid solution , wherein the second fluid 3170 forms a mem - mL/ minute , between 10 mL /minute and 500 mL /minute , brane while a third fluid 3120 can be a suspending buffer. 55 between 10 mL /minute and 100 mL /minute , and preferably Without the suspending buffer outer layer, the lipids would between 50 mL /minute and 100 mL /minute through the first for with their hydrophobic regions facing out towards the inner fluid conduit 3165 . In some embodiments , the first hydrophobic gas . This four fluid nozzle assembles the outer fluid exits the exit orfice 3163 of the first inner fluid conduit membrane in a way to prevent aggregation . In such embodi - 3165 and enters the first fluid contacting chamber 3125 , ments, the incorporation of the active agent is high because 60 whereupon the first fluid 3115 comes in physical commu the active agent fluid can be surrounded by the lipid layer nication with the second fluid 3170 . In some embodiments , forming solvent 3170 . Small concentrations of lipids in 3170 the second fluid 3170 travels at a volumetric flow rate can provide at least one bilayer. In some embodiments , between 2 mL /minute and 1 , 000 mL /minute , between 10 higher concentrations of lipid in the second fluid can provide mL /minute and 500 mL /minute , between 10 mL /minute and liposomes comprising thick large numbers of bilayers. In 65 100 mL /minute , and preferably between 50 mL /minute and some embodiments , structurally strong and stable liposomes 100 mL /minute , through a second inner fluid conduit 3175 . can be made using tight packing saturated phospholipids In a typical embodiment, the first fluid 3115 is an emulsion US 9 ,730 ,892 B2 45 46 without any lipids that could interact with the active agent of polymer or phospholipid or combination thereof consti ( e . g . no charged lipid such as DPPG ) In another embodi- tute further embodiments as disclosed herein . ment, the second fluid 3170 is a volatile solvent comprising The four fluid atomizing nozzle 320 comprises a gas input a lipid solution ( could be the same or different than that used channel 3130 which supplies a gas 3140 . In some embodi in the first component) . ments , the volumetric flow rate of the gas can be between 30 In some embodiments , the first fluid 3115 travels through L /minute and 1 . 000 L /minute , between 30 L /minute and 500 the first fluid contacting chamber 3125 , surrounded by the L /minute , between 20 L /minute and 200 L /minute , and second fluid 3170 . In some embodiments , the first fluid 3115 and the second fluid 3170 continue through the first fluid preferably between 25 L /minute and 100 L /minute and at a contacting chamber 3125 into the second fluid contacting 10 pressure between 5 psig and 1 ,000 psig , between 10 psig and chamber 3137 . In some embodiments, the second fluid 3170 250 psig , between 20 psig and 150 psig , and preferably forms a sheath around the first fluid 3115 core . between 50 psig and 120 psig to the annular gas chamber In some embodiments , the first and second fluids can 3135 . The above atomizing nozzle flow rates are for a single travel at a volumetric flow rate between 2 mL/ minute and atomizing nozzle . Prior to reaching the nozzle 320 , the gas 1 . 000 mL /minute between 10 mL /minute and 500 mL /min - 15 3140 can be passed through a gas filtration system (not ute , between 10 mL /minute and 100 mL /minute , and pref shown ) comprising a hydrophobic gas sterilizer ( as seen in erably between 50 mL /minute and 100 mL /minute through FIG . 1A and FIG . 1B , component 35 ) . In some embodi the second inner conduit 3175 . In some embodiments , the ments , the outlet of the cylindrical tip 3145 and the outlet of first and second fluids exits the exit orfice 3147 the second the exit orfice 3147 of the first second contacting chamber inner conduit 3175 and enter the second fluid contacting 20 3137 have the same diameter . In some embodiments , the chamber 3137 , whereupon the first and second fluids come diameter of the outlet of the cylindrical tip 3145 is larger in physical communication with a third fluid 3120 . In a than the diameter of the outlet of the exit orfice 3147 of the typical embodiment, the third fluid 3120 is a buffer solution . second fluid contacting chamber 3137 . In some embodi In some embodiments , the third fluid 3120 travels at a ments , the diameter of the outlet of the cylindrical tip 3145 volumetric flow rate between 2 mL /minute and 1 , 000 25 is smaller than the diameter of the exit orfice 3147 of the mL /minute , between 10 mL /minute and 500 mL /minute , second fluid contacting chamber 3137 . between 10 mL /minute and 100 mL /minute , and preferably FIG . 3G shows an expanded cross - sectional view of the between 50 mL /minute and 100 mL /minute , through an first fluid , second fluid and third fluid in the cylindrical tip outer fluid conduit 3123 , which annularly surrounds the first 3145 . inner fluid conduit 3165 and the second inner conduit 3175 30 FIG . 3H shows an expanded cross - sectional view of the ( as seen from the downward perspective view F and dis first fluid , second fluid and third fluid in a droplet 3255 played in FIG . 3F ) . In some embodiments , the first , second FIG . 31 and third fluids exit the exit orfice 3145 of the second fluid FIG . 31 is a schematic partial cross -sectional view of a contacting chamber 3137 . In some embodiments , the third four fluid atomizing nozzle 320 , comprising a first inner fluid 3120 forms a sheath around the first and second fluids. 35 fluid conduit 3165 , a second inner conduit 3175 , an outer In some embodiments, the diameter of the first fluid conduit 3123 , a fluid contacting chamber 3125 , a gas input contacting chamber 3125 can be conically narrowed as it channel 3130 , an annular gas chamber ( a fourth fluid con approaches the exit orfice 3163 of the first inner fluid conduit duit ) 3135 , and an annular gas orifice 3150 . In some embodi 3165 . As the first fluid 3115 travels through the narrowing ments , the fluid contacting chamber 3125 is conically first fluid contacting chamber 3125 and toward the exit 40 tapered from the exit orfice 3173 of an outer fluid conduit orfice 3147 of the second fluid conduit 3175 , the diameter of 3123 to a cylindrical tip 3145 . the first fluid 3115 core is correspondingly decreased . Like - The fluid contacting chamber 3125 is annularly sur wise , the second fluid 3170 can be constricted to create a rounded by an annular gas chamber 3135 , which narrows to thinner concentric annular sheath around the first fluid 3115 form an annular gas orifice 3150 . The bottom portion of the core . As a result of decreasing the diameter of the first fluid 45 cylindrical tip 3145 is annularly and concentrically sur contacting chamber 3125 and passing the fluids through the rounded by the annular gas orifice 3150 . exit orfice 3147 of the second fluid conduit 3175 , the In some embodiments , a first fluid 3115 can travel at a velocities of the first fluid 3115 and the second fluid 3170 are volumetric flow rate between 2 mL / minute and 1 , 000 increased . mL /minute , between 10 mL /minute and 500 mL /minute , In some embodiments , the diameter of the second fluid 50 between 10 mL /minute and 100 mL /minute , and preferably contacting chamber 3125 can be conically narrowed as it between 50 mL/ minute and 100 mL /minute through the first joins the cylindrical tip 3145 . As the first fluid 3115 core and inner fluid conduit 3165 . In a typical embodiment, the first second fluid 3120 travel through the narrowing second fluid fluid 3115 is an emulsion . In some embodiments , the first contacting chamber 3125 into the cylindrical tip 3145 , the fluid exits the bottom 3163 of the first inner fluid conduit diameter of the first fluid 3115 core , the second fluid 3120 55 3165 and enters the fluid contacting chamber 3125 , where the third fluid 3170 can be further correspondingly upon the first fluid 3115 comes in physical communication decreased . Likewise , the third fluid 3170 can be constricted with a second fluid 3170 . In some embodiments , a second to create a thinner concentric annular sheath around the first fluid 3170 exits the bottom 3173 of the second inner conduit fluid 3115 core and the second fluid 3120 sheath . In some 3175 and enters the fluid contacting chamber 3125 , where embodiments , the multi vesicular liposomes are lipid coated 60 upon the second fluid 3170 comes in physical communica with lipids on the outside that are different than the lipids on tion with the first fluid 3115 and a third fluid 3170 . In a the inside . In some embodiments, the four fluid nozzle typical embodiment, the first fluid 3115 in the first inner fluid illustrated in FIG . 3E and FIG . 31 could be used to manu - conduit 3165 is annularly surrounded by the second fluid facture multi vesicular liposomes coated with an additional 3170 in the second inner conduit 3175 which is annularly layer of polymer PLGA or a highly charged phospholipid 65 surrounded by the third fluid 3120 in the outer conduit 3123 different from the phospholipid of the internal membrane . (as seen from the downward perspective view J and dis These multi vesicular liposomes coated with an extra layer played in FIG . 3J) . US 9 , 730 , 892 B2 48 In some embodiments , the first fluid 3115 travels through their diameter is correspondingly decreased along the axis of the first fluid contacting chamber 3125 , surrounded by the travel. Likewise , the second fluid 4120 is constricted to second fluid 3170 which is surrounded by the third fluid create a thinner shell around the first fluid droplets 4157 . As 3120 . In some embodiments , the second fluid 3170 forms a a result of decreasing the diameter of the fluid contacting sheath around the first fluid 3115 and the third fluid 3120 5 chamber 4125 and passing the solutions through the cylin forms a sheath around the first and second fluids. drical tip 4145 , the velocities of the first fluid droplets 4157 In some embodiments , the diameter of the fluid contacting and second fluid 4120 are increased . In a typical embodi chamber 3125 can be conically narrowed as it joins the ment, the first fluid 4115 is an emulsion and the second fluid cylindrical tip 3145 . In some embodiments , as the first fluid 4120 is a buffer solution . 3115 travels through the narrowing first fluid contacting 10 FIG . 4C shows an expanded cross - sectional view of the chamber 3125 and toward the cylindrical tip 3145 , the first fluid droplets 4157 and second fluid 4120 in the diameter of the first fluid 3115 core is correspondingly cylindrical tip 4145 . decreased . In some embodiments, the second fluid 3170 can FIG . 4D shows an expanded cross - sectional view of the be constricted to create a thinner concentric annular sheath first fluid and second fluid in a droplet 4155 . In some around the first fluid 3115 core . In some embodiments , the 15 embodiments , the ratio of the diameters of the first fluid to third fluid 3120 can be constricted to create a thinner the second fluid in a droplet 4155 can be the same as the ratio concentric annular sheath around the first fluid 3115 core of the diameters of the first fluid in the inner fluid conduit and the concentric annular second fluid 3170 . As a result of 4165 to the second fluid in the outer fluid conduit 4123 . In decreasing the diameter of the first fluid contacting chamber some embodiments , the ratio of the diameters of the first 3125 and passing the fluids through the cylindrical tip 3145 , 20 fluid to the second fluid in a droplet 4155 can be the larger the velocities of the first fluid 3115 , second fluid 3170 and than the ratio of the diameters of the first fluid in the inner third fluid 3120 can be increased . fluid conduit 4165 to the second fluid in the outer fluid The four fluid atomizing nozzle 320 comprises a gas input conduit 4123 . In some embodiments , the ratio of the diam channel 3130 which supplies a gas 3140 . In some embodi- eters of the first fluid to the second fluid in a droplet 4155 can ments , the volumetric flow rate of the gas can be between 30 25 be the smaller than the ratio of the diameters of the first fluid L /minute and 1, 000 L / minute , between 30 L /minute and 500 in the inner fluid conduit 4165 to the second fluid in the outer L /minute , between 20 L /minute and 200 L /minute , and fluid conduit 4123 . preferably between 25 L /minute and 100 L /minute and at a The nozzle 410 comprises a gas input channel 4130 which pressure between 5 psig and 1 , 000 psig , between 10 psig and supplies a gas 4140 to an annular gas chamber ( a third fluid 250 psig , between 20 psig and 150 psig , and preferably 30 conduit ) 4135 . Prior to reaching the nozzle 410 , the gas 4140 between 40 psig and 120 psig to the annular gas chamber can be passed through a gas filtration system (not shown ) 3135 . The above atomizing nozzle flow rates are for a single comprising a hydrophobic gas sterilizer (FIG . 1A and FIG . atomizing nozzle . Prior to reaching the nozzle 320 , the gas 1B , component 35 ) . The wall of the fluid contacting chamber 3140 can be passed through a gas filtration system (not 4125 provides a barrier between the annular gas chamber shown ) comprising a hydrophobic gas sterilizer ( as seen in 35 4135 and the fluid contacting chamber 4125 such that there FIG . 1A and FIG . 1B , component 35 ) . is no physical contact between the fluids in the fluid con FIG . 3K shows an expanded cross -sectional view of the tacting chamber 4125 and the gas 4140 in the annular gas first fluid , second fluid and third fluid in the cylindrical tip chamber 4135 . In some embodiments , the outlet of the 3145 . cylindrical tip 4145 and the outlet of the annular gas orifice FIG . 3L shows an expanded cross -sectional view of the 40 4150 can be flush with each other such that the second fluid first fluid , second fluid and third fluid in a droplet 3255 . 4120 does not come into contact with the gas 4140 prior to FIG . 4A the gas 4140 exiting the atomizing nozzle 410 . In other FIG . 4A depicts an atomizing nozzle 410 where the first embodiments , the cylindrical tip 4145 may extend below the fluid 4115 can break down into large droplets in the fluid end of the annular gas orifice 4150 . In still other embodi contacting chamber 4125 prior to reaching the cylindrical tip 45 ments , the cylindrical tip 4145 may be slightly recessed 4145 . within the annular gas orifice 4150 , such that the gas 4140 In some embodiments , a first fluid 4115 exits a inner fluid comes into physical communication with the second fluid conduit 4165 through the exit orfice 4163 of the inner fluid 4120 prior to the second fluid fully exiting from the nozzle conduit 4165 and enters the fluid contacting chamber 4125 , 410 . In a typical embodiment, the first fluid 4115 is an whereupon the first fluid 4115 comes in physical commu - 50 emulsion and the second fluid 4120 is a buffer solution . nication with a second fluid 4120 . The second fluid 4120 In some embodiments , as the gas 4140 passes through the travels through an outer fluid conduit 4123 , which annularly annular gas orifice 4150 and exits the atomizing nozzle 410 , surrounds the inner fluid conduit 4165 (as seen from the it comes in physical communication with the second fluid downward perspective view B and displayed in FIG . 4B ), to 4120 and acts to shear the stream of the second fluid 4120 reach the fluid contacting chamber 4125 . The first fluid 4115 55 and the first fluid 4115 droplets into atomized droplets 4155 forms a plurality of first fluid droplets 4157 traveling made of a first fluid 4115 core and a second fluid 4120 shell . through the fluid contacting chamber 4125 , the first fluid In this embodiment the first fluid core stream breaks up due droplets 4157 now being in physical communication with , to the mismatch of velocities between the first fluid 4115 and and surrounded by , the second fluid 4120 . In some embodi - the second fluid 4120 at the exit orfice 4163 of the inner fluid ments , the first and second fluid are immiscible . In a typical 60 conduit 4165 . In a typical embodiment, the first fluid 4115 embodiment, the first fluid 4115 is an emulsion and the is an emulsion and the second fluid 4120 is a buffer solution . second fluid 4120 is a buffer solution . FIG . 5 In some embodiments , the diameter of the fluid contacting FIG . 5 is a more detailed view of one embodiment of an chamber 4125 is conically narrowed as it joins the cylindri - atomizing nozzle 505 . The nozzle depicted in FIG . 5 is cal tip 4145 . As the first fluid 4115 travels through the 65 derived from Part No . 1 / 8JJN -SS + SUJIA -SS manufactured narrowing fluid contacting chamber 4125 and into the cylin - by Spraying Systems Co . of Wheaton , Ill. , as modified in drical tip 4145 , the first fluid droplets 4157 are squeezed and FIG . 5 and described herein . The atomizing nozzle lower US 9 ,730 ,892 B2 49 50 section 510 of FIG . 5 provides three fluid input channels for holes in the gas restrictor 5182 , which annularly surrounding the introduction of three fluids into the nozzle 505 . A first the liquid fluid cap 5180 . The presence of the gas restrictor fluid input channel is the central longitudinal channel 5114 , 5182 creates a lower annular gas chamber 5136 between the which extends substantially downwardly through the atom - gas restrictor 5182 and the gas cap 5160 . The gas restrictor izing nozzle lower section 510 and extends into a fluid 5 5182 can minimize the turbulence possibly created as a gas contacting chamber 5125 . In some embodiments , a second travels through the annular gas chamber 5135 to the annular fluid input channel is a buffer input channel 5128 and a third gas orifice 5150 . The nozzle lower section 510 can be fitted fluid input channel is a gas input channel 5130 . to an orifice housing 5185 which acts to keep the gas cap The central longitudinal channel 5114 is a channel which 5160 and liquid fluid cap 5180 securely fitted to the hori may be of circular cross -section . The central longitudinal 10 zontal channel casing 5175 . In some embodiments , the channel 5114 may begin as an aperture 5111 in a line coupler orifice housing 5185 can be threaded . 5113 . The line coupler 5113 is a means for attaching a supply In some embodiments, the first fluid 5115 can enter the line to the nozzle 505 for the introduction of the first fluid longitudinal channel 5114 of the nozzle lower section 510 5115 into the nozzle lower section 510 . A polymeric sealing from a supply line ( not shown ) attached to the line coupler washer 5112 is retained in contact with the line coupler 5113 15 5113 . The first fluid travels through the longitudinal channel and interposed between a coupler nut 5116 of the line 5114 until it reaches the fluid contacting chamber 5125 coupler 5113 and a head (not shown ) of a inner fluid conduit whereupon it comes in physical contact with the second fluid 5165 . 5120 in the fluid contacting chamber 5125 . In a typical The inner fluid conduit 5165 can provide a continuation of embodiment, the first fluid 5115 is an emulsion and the the central longitudinal channel 5114 , with a narrowed 20 second fluid 5120 is a buffer solution . central bore 5164 extending through a removable housing In some embodiments , the second fluid 5120 enters the member 5170 into the horizontal channel casing 5175 , and nozzle lower section 510 through an input aperture 5127 and into the liquid fluid cap 5180 , ending in the fluid contacting travels through the input channel 5128 . In some embodi chamber 5125 of the liquid fluid cap 5180 . In some embodi - ments , the second fluid 5120 then passes through the internal ments , the housing member 5170 can be threaded . The 25 aperture 5126 leading to the top of the liquid fluid cap 5180 housing member 5170 can be fitted on the top with a housing and travels through the annular outer fluid conduit 5123 , and cap 5172 and housing gasket member 5171 to provide travels around and surrounds the inner fluid conduit 5165 at secure fastening and fluid seal . The housing member 5170 a lower section to reach the fluid contacting chamber 5125 . extends downwardly into a top portion of the horizontal In some embodiments , upon exiting the exit orfice 5163 of channel casing 5175 , wherein it can be tightened to form a 30 the inner fluid conduit 5165 or the central longitudinal secure connection with the horizontal channel casing 5175 , channel 5114 and entering the fluid contacting chamber for example using internal screw threading . 5125 , the first fluid 5115 can form a cylindrical core trav In addition to the central longitudinal channel 5114 , the eling through the fluid contacting chamber 5125 . In some horizontal channel casing 5175 can be comprised of the gas embodiments , the first fluid 5115 cylindrical core can be input channel 5130 and the buffer input channel 5128 . The 35 surrounded annularly by the second fluid 5120 . In some gas input channel 5130 can have an external aperture 5131 embodiments , the immiscibility between the second fluid to which a gas supply line (not shown ) can be to be attached . 5120 and the first fluid 5115 , and because of the high The gas input channel 5130 can have an internal aperture velocity at which the second fluid 5120 and first fluid 5115 5132 which leads to an annular gas chamber ( third fluid are traveling , causes the second fluid 5120 to form a sheath conduit ) 5135 , the annular gas chamber 5135 situated to 40 around the first fluid 5115 core . The first fluid 5115 and annularly surround the liquid fluid cap 5180 . The liquid fluid second fluid 5120 can travel through the fluid contacting cap 5180 can be detachably and securely fitted into the chamber 5125 into the cylindrical tip 5145 of the outer horizontal channel casing 5175 such that fluid contents of conduit 5123. In a typical embodiment , the first fluid 5115 the liquid fluid cap 5180 and the fluid contacting chamber is an emulsion and the second fluid 5120 is a buffer solution . 5125 cannot physically communicate with the fluid contents 45 The cross - sectional diameter of the fluid contacting cham of the annular gas chamber 5135 inside the nozzle . The b er 5125 decreases as it conically narrows to join the buffer input channel 5128 can have an external aperture cylindrical tip 5145 . As the first fluid 5115 travels through 5127 to which a fluid supply line ( not shown ) can be the narrowing fluid contacting chamber 5125 and into the attached . The buffer input channel 5128 can have an internal cylindrical tip 5145 the diameter of the first fluid 5115 core aperture 5126 which leads to the top opening of the liquid 50 is decreased . Likewise , the second fluid 5120 can be con fluid cap 5180 . stricted to create a thinner concentric annular sheath around In some embodiments , the liquid fluid cap 5180 can be the first component 5115 core . In some embodiments , as the cylindrically shaped in a top portion and conically tapered in solutions pass through the narrowing cylindrical tip , the a exit orfice portion . In some embodiments , the liquid fluid velocities of the first fluid 5115 and the second fluid 5120 cap 5180 can be cylindrically tapered in both a top portion 55 can be increased . In some embodiments , the threads on the and a bottom portion . In some embodiments , the fluid inner fluid conduit 5165 and the housing member 5170 contacting chamber 5125 and the outer fluid conduit 5123 allow the outlet tip of the inner fluid conduit 5165 to be form the interior wall of the liquid fluid cap 5180 . The fluid adjusted vertically within the conical portion of the fluid contacting chamber 5125 is conically tapered from the exit contacting chamber 5125 within the liquid fluid cap 5180 . In orfice of an annular outer fluid conduit 5123 to a cylindrical 60 some embodiments, the housing member 5170 can be tip 5145 . The exit orfice portion of the cylindrical tip 5145 adjusted so that the velocities of the second fluid 5120 and can be annularly and concentrically surrounded by an annu - the first fluid 5115 can be made approximately equal at the lar gas orifice 5150 . The annular gas orifice 5150 can be point where the first fluid 5115 exits the inner fluid conduit created by placement of the cylindrical tip 5145 in the center 5165 . This allows optimal operation as shown in FIG . 3A . In of a cylindrical opening in a gas cap 5160 . The liquid fluid 65 some embodiments , the housing member 5170 can be cap 5180 can have a gas restrictor 5182 attached to the adjusted so that the velocities of the second fluid 5120 and exterior of the liquid fluid cap 5180 with several circular the first fluid 5115 can bemade uequal at the point where the US 9 ,730 , 892 B2 52 first fluid 5115 exits the inner fluid conduit 5165 . This allows In some embodiments , location 5130 can be a gas input optimal operation as shown in FIG . 4A . In a typical embodi channel; location 5132 can be an internal gas chamber ment, the first fluid 5115 is an emulsion and the second fluid aperture ; location 5128 can be a fluid channel; location 5126 5120 is a buffer solution . can be an internal fluid cap aperture ; and location 5150 can In some embodiments , a gas 5140 can be first passed 5 be an annular gas orifice . through a gas filtration system ( not shown ) comprising a FIG . 7 hydrophobic gas sterilizer ( not shown ). In some embodi- An embodiment of the present application is a system for ments , the gas 5140 can enter the gas input channel 5130 manufacturing formulations including an evaporation appa through the external gas input aperture 5131 and then passes ratus, or evaporation sub - system , the process of using the through the internal gas chamber aperture 5132 to reach the 10 system , and the large diameter synthetic membrane vesicles annular gas chamber 5135 . In some embodiments , the liquid products made by the process . An example of an evaporation fluid cap 5180 can be detachably and securely fitted into the apparatus is the solvent removal vessel presented in FIG . 7 horizontal channel casing 5175 such that the gas 5140 and described herein . FIG . 7 shows a schematic of a solvent cannot physically communicate with the first fluid 5115 or removal vessel 710 . In some embodiments , atomized drop the second fluid 5120 inside the nozzle lower section 510 . 15 lets 7155 can be sprayed into the solvent removal vessel 710 The gas 5140 travels through the holes in the gas restrictor in order to remove solvent from atomized droplets 7155 . In 5182 to reach the lower annular gas chamber 5136 . some embodiments , the solvent removal vessel 710 can As the gas 5140 passes through the annular gas orifice include a three - fluid atomizing nozzle 7510 , described 5150 and exits the atomizing nozzle lower section 510 , it above , attached to and extending through a lid 7220 of the comes in physical communication with the second fluid 20 solvent removal vessel 710 . In some embodiments , the 5120 and acts to shear the stream of the second fluid 5120 cylindrical tip and annular gas orifice of the atomizing and first fluid 5115 into atomized droplets 5155 made of a nozzle 7510 can be configured to be substantially flush with first fluid 5115 core and a second fluid 5120 shell. In a the inside edge of the lid 7220 . In some embodiments , the typical embodiment, the first fluid 5115 is an emulsion and cylindrical tip and annular gas orifice of the atomizing the second fluid 5120 is a buffer solution . In a typical 25 nozzle 7510 can be configured to extend beyond the inside embodiment, the first fluid 5115 core is an emulsion and the edge of the lid 7220 . second fluid 5120 shell is a buffer solution . Alternatively , in some embodiments , the solvent removal Some embodiments provide an atomizing nozzle appara - vessel 710 can include a three - fluid atomizing nozzle 7510 , tus , comprising three channels , each having at least one described above , attached to and extending through a wall entrance orifice and one exit orifice, the channels comprise 30 7350 of the solvent removal vessel 710 . In some embodi of an inner fluid conduit and an outer fluid conduit, wherein ments , the cylindrical tip and annular gas orifice of the the exit orifice for the inner fluid conduit is of a diameter atomizing nozzle 7510 can be configured to be substantially smaller than , and is located centrally in , the outer fluid flush with the inside edge of the wall 7350 . In some conduit and is directed towards the outer fluid conduit exit embodiments , the cylindrical tip and annular gas orifice of orifice , a gas channel, wherein a pressurized gas exiting the 35 the atomizing nozzle 7510 can be configured to extend gas channel exit orifice impinges a liquid exiting the exit beyond the inside edge of the wall 7350 . orifice of the outer fluid conduit. Additional embodiments In some embodiments , the vessel 710 also includes a gas include the processes for using such a device and the MVL outlet tube 7340 , which can pass through the center of the lid products made by the same. In some embodiments , the gas 7220 , and extend into a solvent removal chamber 7230 . In channel exit orifice can annularly surround the exit orifice of 40 some embodiments , the gas outlet tube 7340 can extend the outer fluid conduit . In some embodiments , the gas down from the lid from about 15 to about 19 inches and can channel exit orifice and the outer fluid conduit exit orifice be about 1 inch in diameter . In some embodiments , the gas can be flush . In some embodiments , the outer fluid conduit outlet tube 7340 can extend down from the lid from about 20 exit orifice can be recessed within the gas channel exit cm to about 60 cm and can be from about 1 cm to about 4 orifice . In some embodiments , the outer fluid conduit exit 45 cm in diameter. In some embodiments , a conical fitting 7300 orifice can extend beyond the gas channel exit orifice . In can be attached to the end of the gas outlet tube 7340 . In some embodiments , the exit orifices of all three channels can some embodiments, the conical fitting 7300 can be used to be coaxial. In some embodiments , the exit orifice of the narrow the diameter of the gas outlet tube 7340 at a gas inner fluid conduit can be more than two outer fluid conduit outlet 7310 . In some embodiments , the gas outlet 7310 can exit orifice diameters away from the outer fluid conduit exit 50 be approximately 0 . 5 inches in diameter . In some embodi orifice . ments , the gas outlet 7310 can be from about 0 . 5 cm to about In some embodiments , the multi vesicular liposome prod 3 cm in diameter . In some embodiments , a vortex stabilizer uct produced by the atomizing nozzle in FIGS . 3 - 6 can be disk or ring 7360 can be attached to the conical fitting 7300 . directly collected and purified . Alternatively , the MULTI In some embodiments , a gas inlet pipe 7290 can be VESICULAR LIPOSOME product can be processed by an 55 connected to the wall 7350 of the solvent removal vessel 710 evaporation tower or the evaporation apparatus of FIG . 7 ( or can be creating a gas inlet 7280 . In some embodiments, the as part of FIGS. 1A - 1C ) before being processed . solvent removal vessel 710 has a bottom 7250 , which can FIG . 6 include a product exit orifice 7260 . In some embodiments , FIG . 6 is an exploded schematic 605 of the individual the exit orifice 7260 can be in the center of the domed components of the device of FIG . 5 . 60 bottom 7250 . In some embodiments , a product outlet pipe In some embodiments , component 5113 is a line coupler ; 7270 can be attached to the exit orifice 7260 . In some component 5165 is an inner fluid conduit ; component 5172 embodiments , the solvent removal vessel 710 , includes a is a housing cap ; component 5170 is a housing member ; temperature control jacket ( not shown ) . In some embodi component 5171 is a housing gasket member ; component m ents , temperature controlling fluid can be circulated 5180 is a liquid fluid cap ; component 5145 is a cylindrical 65 through a temperature control jacket (not shown ) of the tip ; component 5160 is a gas cap ; component 5182 is a gas solvent removal vessel 710 , wherein the temperature control restrictor; and component 5185 is an orifice housing . jacket can surround the wall 7350 and the bottom 7250 of US 9 , 730 , 892 B2 53 54 the solvent removal vessel 710 for temperature regulation in outlet pipe 7340 can be equipped with a sterile barrier the solvent removal chamber 7230 . In some embodiments, filtration system (not shown ) outside of solvent removal the bottom 7250 of the solvent removal vessel 710 could be vessel 710 . The filtration system in one embodiment can be of a shape such as a dome, a cone or an angled flat bottom . comprised of filter ( e . g . a course filter or conventional In some embodiments , a carrier gas 7370 can be first 5 cyclone separator ) and a HEPA filter or other sterilizing gas heated and humidified and then passed through a gas filtra - filter. tion system ( not shown ) comprising a hydrophobic gas In some embodiments, a portion of the atomized droplets sterilizer (not shown ) . In some embodiments , the carrier gas 7155 , traveling in the slow gas rotation 7240 , can reach the 7370 can be supplied to the solvent removal vessel 710 intense gas vortex 7245 , where they can be kicked outwards through the gas inlet pipe 7290 . In some embodiments , the 10 again by the intense centrifugal forces within the intense gas carrier gas 7370 can pass through the gas inlet 7280 and vortex 7245 , and thus are not removed through the gas outlet enter a gas rotation jet 7285 . In some embodiments , the gas 7310 . For example , some of the atomized droplets 7155 rotation jet 7285 directs the flowing carrier gas 7370 inside kicked outwards by the gas vortex 7245 can travel again the solvent removal chamber 7230 such that the carrier gas through the slow gas rotation 7240 and some of the atomized 7370 exits the gas rotation jet 7285 (and enters the solvent 15 droplets begin to fall towards the domed bottom 7250 . In removal chamber 7230 ) horizontally and in a direction one embodiment, very few of the atomized droplets 7155 tangential to the solvent removal vessel wall 5350. In some escape with the carrier gas 7370 through the gas outlet 7310 . embodiments , substantially all the carrier gas 7370 entering In some embodiments , removal of substantially all the the solvent removal chamber 7230 through the gas rotation solvent from the atomized droplets 7155 can afford large jet 7285 can exit the solvent removal chamber 7230 through 20 diameter synthetic membrane vesicles droplets 7380 still the gas outlet 7310 and gas outlet pipe 7340 . coated in a fluid shell . In some embodiments , droplets 7380 In some embodiments, the carrier gas 7370 supplied to permanently fall out of the gas vortex primarily to the domed solvent removal chamber 7230 can be injected tangential to bottom 7250 of the solvent removal vessel 710 . In some the wall 7350 , causing the carrier gas 7370 to first travel embodiments , the plurality of large diameter synthetic mem slowly clockwise (as viewed from above ) around the solvent 25 brane vesicles droplets 7380 form a suspension 7390 of large removal vessel 710 near the wall 7350 , and forming a slow diameter synthetic membrane vesicles particles in a solution gas rotation 7240 . In some embodiments , the solvent at the domed bottom 7250 . In a typical embodiment, the removal chamber 7230 can be pressured to approximately 1 large diameter synthetic membrane vesicles are multivesicu psig , creating a pressure differential between the solvent lar liposomes . removal chamber 7230 and the gas outlet pipe 7340 . In some 30 The solvent removal vessel 710 can optionally be embodiments , the carrier gas 7370 traveling in the solvent equipped with a two - fluid rinse nozzle 7400 in the lid 7220 removal chamber 7230 can be pulled inwards, as it circu - of the solvent removal vessel 710 . In some embodiments , lates, towards the gas outlet 7310 . In some embodiments, the the two -fluid rinse nozzle 7400 can be positioned in the lid change in angular momentum on the carrier gas 7370 causes 7220 at 90 degrees clockwise from the atomizing nozzle the carrier gas 7370 to accelerate as it moves closer to the 35 7510 . In some embodiments , the two - fluid rinse nozzle 7400 gas outlet 7310 . In some embodiments, the acceleration of can be positioned in the lid 7220 at 180 degrees clockwise the carrier gas 7370 near the gas outlet 7310 can be from the atomizing nozzle 7510 . The rinse nozzle 7400 can sufficiently strong to create an intense gas vortex 7245 be fed a wall rinse solution 7410 and a filtered gas 7420 in underneath the vortex stabilizer 7360 and gas outlet 7310 . In order to spray atomized wall rinse solution droplets 7415 some embodiments , the carrier gas 7370 can be pushed out 40 into the solvent removal vessel 710 for the purpose of of the solvent removal chamber 7230 through the gas outlet rinsing any wayward atomized emulsion droplets from the 7310 into the gas outlet pipe 7340 for disposal after spinning wall 7350 and rinsing the large diameter synthetic mem through the gas vortex 7245 . brane vesicles suspension 7390 to the product exit orifice In some embodiments , the three - fluid atomizing nozzle 7260. In a typical embodiment, the large diameter synthetic 7510 can be supplied with a first fluid 7115 , a second fluid 45 membrane vesicles are multivesicular liposomes . 7120 , and a third fluid 7140 , as described above . In a typical In some embodiments , solvent removal vessel 710 can be embodiment , the first fluid can be a first component, the optionally equipped with a lid -protecting gas inlet 7430 in second fluid can be a buffer solution , and the third fluid can the lid 7220 . In some embodiments , solvent removal vessel be a gas. In some embodiments , the resultant atomized 710 can be optionally equipped with a lid -protecting gas droplets 7155 , comprised of a first fluid core and a second 50 inlet 7430 in the lid 7220 on a side opposite the atomizing fluid shell , can be sprayed into the solvent removal chamber nozzle 7510 . In some embodiments , humidified , sterilized , 7230 . In some embodiments, the atomized droplets 7155 and filtered gas 7440 can be streamed into the solvent come into contact with the carrier gas 7370 being circulated removal chamber 7230 in a direction tangential to the in the slow gas rotation 7240 as the atomized droplets 7155 solvent removal vessel wall 7350 through the lid -protecting can travel down through the solvent removal chamber 7230 . 55 gas inlet 7430 , near the top of the solvent removal chamber In some embodiments , the atomized droplets 7155 can be 7230 . In some embodiments , gas 7440 can travel circularly , picked up by , and incorporated in , the slow gas rotation 7240 as a lid protection jet, around the top of solvent removal and begin to circulate through the solvent removal chamber chamber 7230 and , in essence , acts a gas cushion above the 7230 , and begin to settle toward the domed bottom 7250 . gas vortex 7245 and slow gas rotation 7240 to prevent In some embodiments , the solvent can be substantially 60 droplet buildup on the lid 7220 . In some embodiments , the evaporated from the core of the atomized droplets 7155 as gas streamed through the lid -protecting gas inlet can be , for the atomized droplets 7155 circulate along with the carrier example , nitrogen gas (or a nitrogen gas/ aqueous vapor gas 7370 in the slow gas rotation 7240 . In some embodi- mixture ) or air scrubbed of CO2. In some embodiments , the ments , the evaporated solvent can be removed from the carrier gas can be nitrogen gas ( or a nitrogen gas / aqueous solvent removal chamber 7230 along with the circulating 65 vapor mixture ) or air scrubbed of CO2. carrier gas 7370 through the gas outlet 7310 and into the gas In some embodiments , the vortex stabilizer 7360 can be outlet pipe 7340 for disposal. In some embodiments , the gas a metal disk or ring extending radially outward from the US 9 ,730 ,892 B2 55 56 conical fitting 7300 and can be fitted flush with the gas outlet nozzle which is not located on the central axis of the circular 7310 at the end of the conical fitting 7300 . In some embodi wall , a carrier gas entrance orifice tangential to the circular ments, the diameter of the vortex stabilizer 7360 is twice to wall, a carrier gas exit orifice located on the central axis of six times the diameter of the gas outlet 7310 . In some the circular wall and directed along the axis with a diameter embodiments , the vortex stabilizer 7360 acts to ensure 5 of less than 1 /5 of the circular wall diameter , and a product helical stability and integrity of the intense gas vortex 7245 exit orifice in the bottom of the vessel. Additional embodi by protecting the tip of the intense gas vortex 7245 from the ments include the processes for using such a device and the turbulence caused by the spray emanating from the atom - MVL products made by the same. In some embodiments , at izing nozzle 7510 . In some embodiments , the stability of the least part of the vessel can be jacketed . In some embodi gas vortex 7245 can be maintained by placement of the 10 ments , the atomizing nozzle can be mounted to and extend atomizing nozzle 7510 at a distance between 1/ 4 lid radius ing through the lid of the solvent removal vessel. In some from the wall 7350 and 1/ 4 lid radius from the gas outlet pipe embodiments , the apparatus further comprises a rinse nozzle 7340 . In some embodiments , the atomizing nozzle 7510 can mounted to and extending through the lid of the solvent be positioned at a distance of 711 the radius of the lid 7220 removal vessel. In some embodiments , the atomizing nozzle from the gas outlet pipe 7340 . Strategic placement of the 15 can be used to cause the gas within the vessel to rotate . In atomizing nozzle 7510 minimizes the impact on the intense some embodiments , the atomizing nozzle can be angled at gas vortex 7245 of spraying the atomized droplets 7155 into least 5 degrees measured off the central axis of the wall and the solvent removal chamber 7230 and also minimizes the in a plane parallel to the wall nearest to it . In some number of the atomized droplets 7155 that impinge on the embodiments , the gas entrance orifice can be combined with wall 7350 . In some embodiments, the rinse nozzle 7400 20 the atomizing nozzle . In some embodiments , the carrier gas placement is not critical as long as a substantial quantity of exit orifice can comprise a tube extending approximately 2/ 3 rinse solution droplets 7415 enter the slow gas rotation 7240 of the way into the solvent removal vessel. In some embodi within the solvent removal vessel 7240 and deposit on the ments , the tube can be fitted with a narrowing cone and an vessel walls 7350 and bottom 7250 . In some embodiments , optional annular ring . In some embodiments , the gas exit the rinse nozzle 7400 can also be of a one fluid ( liquid ) 25 orifice diameter can be less than a 1/ 10 diameter of the design fed by pressurized rinse solution . In some embodi - diameter of the inside of the tank . In some embodiments , the ments , the nozzle can be 180 degrees from the three fluid atomizing nozzle can be a nozzle as disclosed herein . In nozzle to suppress off access gas rotation . some embodiments , the ratio of the inside diameter of the In some embodiments , the suspension 7390 collected at solvent removal vessel to the diameter of the carrier gas exit the domed bottom 7250 of the solvent removal vessel 710 30 orifice can be between approximately 5 : 1 and 100 : 1 . In some can be drained and optionally pumped by a pump (not embodiments , the ratio of the inside diameter of the solvent shown ) from the domed bottom 7250 through the product removal vessel to the diameter of the carrier gas exit orifice exit orifice 7260 into the product outlet pipe 7270 to can be between approximately 20 : 1 and 60 : 1. In some optionally be further processed . In some embodiments , the embodiments , the solvent removal vessel is comprised of suspension 7390 can be further processed by a buffer 35 two , three , or four atomizing nozzles which are used to spray exchange through a series of diafilters . the atomized droplets into the solvent removal vessel. In some embodiments , the solvent removal vessel of FIG . FIG . 8 7 can be a component of FIGS . 1A and B ( component 50 ). In some embodiments , the large diameter synthetic mem In some embodiments , the atomizing nozzle 7510 can brane vesicles suspension resulting from the spray evapo receive fluid from the line 2180 of FIG . 2 . In some embodi - 40 ration process may optionally undergo a filtration and / or ments , the large diameter synthetic membrane vesicles sus concentration process in a continuous - flow particle - concen pension 7390 can exit the solvent removal vessel of FIG . 7 tration system to concentrate the large diameter synthetic through the product exit orifice 7260 and can enter the membrane vesicles and remove the buffer solution . One systems of FIG . 1C (through line 120 ) , FIG . 8 ( through line embodiment is a continuous - flow particle - concentration 8120 ) , FIG . 10 (through line 0120 ) and /or FIG . 11 ( through 45 unit , as well as the process to concentrate and /or filter the line 1120 ) . Alternatively, the large diameter synthetic mem - large diameter synthetic membrane vesicles particles so brane vesicles suspension 7390 can be collected directly produced . In a typical embodiment, the large diameter from the product exit orifice 7260. synthetic membrane vesicles are multivesicular liposomes . In some embodiments , the apparatus is configured where Other embodiments of particle - concentration systems the gas inlet can be situated in the chamber to cause the gas 50 include the use of one or more hydro - cyclones or one or to rotate within the chamber around an axis of the chamber more centrifuges . An example of a hydro -cyclone is the and a small diameter gas exit orifice situated on that same Minicyclone or Minicyclone Array available from ChemIn axis and adapted to form a small diameter , in relation to the dustrial Systems, Inc . of Cedarburg , Wis . An example of a tank diameter , intense rapidly rotating gas vortex capable of disk centrifuge is Model No . Pathfinder SC1 -06 - 177 manu substantially preventing atomized droplets from exiting 55 factured by GEA Westfalia Separator of Oelde , Germany . through the gas exit. In this document the term concentration unit , concentra In some embodiments , the apparatus is configured where tion apparatus , concentration system , particle - concentration the gas inlet, tank dimensions and gas exit diameter and system , particle -concentrating device , and particle concen position are together adapted to retain the atomized droplets trator are meant to encompass units and processes that in the circulating gas stream rather that force them to a wall. 60 remove some or all of the particle suspending medium of a In some embodiments , the atomized droplets can build up in particle suspension and therefore concentrate the particles. the circulating gas stream until the rate of settling to the Furthermore , the definition of these terms encompasses the liquid exit equals the rate of generation by the atomizing exchange of the suspending medium with a new suspending nozzle . medium , performed in one step or incrementally . These two Some embodiments provide an evaporation apparatus , 65 processes are closely related as exchanging the suspending comprising a sealed solvent removal vessel, comprised of a medium can be accomplished by concentrating the suspen lid , a bottom , and a circular wall , at least one atomizing sion and adding new suspending medium . These terms relate US 9 , 730 , 892 B2 57 58 to concentrating the particle suspension and exchanging the of 1 .33 (large diameter synthetic membrane vesicles con suspending medium done separately or simultaneously . centration is increased by 33 % ) . În a typical embodiment, An embodiment of the present application is a system for the large diameter synthetic membrane vesicles are multi manufacturing formulations including a continuous - flow vesicular liposomes. diafiltration system , the process of using the system , and the 5 In some embodiments , similar filtration can occur in the large diameter synthetic membrane vesicles made by the second retentate vessel 8200 as described above for the first process . An example of a continuous- flow diafiltration sys retentate vessel 8100 . In some embodiments , the large tem is presented in FIG . 8 and described herein . FIG . 8 is a diameter synthetic membrane vesicles suspension can enter schematic of one example of a continuous - flow diafiltration the second retentate vessel 8200 at a rate of 0 .75 L per 1 L system 810 . In some embodiments , the permeate discarded 10 added to the first retentate vessel 8100 . In some embodi in the steps described herein contains a buffer solution in ments , a solution can be fed into the second retentate vessel which large diameter synthetic membrane vesicles can be 8200 through a line 8132 , the solution first passing through suspended as the large diameter synthetic membrane a manual valve 8204 and a sterilizing hydrophilic filter 8172 , vesicles suspension exits the solvent removal vessel. In a at a rate of 2 L per 0 .75 L concentrated large diameter typical embodiment, the large diameter synthetic membrane 15 synthetic membrane vesicles suspension added to the second vesicles are multivesicular liposomes . retentate vessel 8200 . In some embodiments , a portion of the In some embodiments, the large diameter synthetic mem - large diameter synthetic membrane vesicles suspension in brane vesicles suspension can be pumped from the solvent the second retentate vessel 8200 can be pumped by pump removal vessel , such as the example vessel depicted in FIG . 8112 through a cross - flow ( tangential- flow ) filtration mod 7 and described above , to the diafiltration system 810 20 ule 8152 . In some embodiments , the permeate can be drawn through a particle suspension inlet line 8120 (also seen in off through a permeate line 8162 (passing through a steril FIG . 1A , component 120 and in FIG . 7 , component 7270 ) , izing hydrophilic filter 8192 and a manual valve 8206 ) , reaching a first retentate vessel 8100 . In some embodiments , wherein for each 0 . 75 L of large diameter synthetic mem for each 1 L of large diameter synthetic membrane vesicles brane vesicles suspension added to the second retentate suspension fed to the first retentate vessel 8100 , a 2 L 25 vessel 8200 , 2 . 25 L of permeate can be removed and solution can be fed into the first retentate vessel 8100 discarded . In some embodiments , the retentate from the through a line 8130 , the solution first passing through a filtration module 8152 can be circulated back into the second manual valve 8212 and a sterilizing hydrophilic filter 8170 . retentate vessel 8200 via a retentate line 8142 . In some In some embodiments , the large diameter synthetic mem - embodiments , for each 0 .75 L of large diameter synthetic brane vesicles are multivesicular liposomes and the solution 30 membrane vesicles suspension added to the second retentate is a saline solution . vessel 8200 , a 0 . 50 L flow of the concentrated large diameter In some embodiments , a portion of the large diameter synthetic membrane vesicles suspension can be removed synthetic membrane vesicles suspension in the first retentate from the second retentate vessel 8200 through a feed line vessel 8100 can be pumped by pump 8110 through a 8124 and a metering pump 8125 to be further filtered in a cross - flow ( tangential- flow ) filtration module 8150 . For 35 final retentate vessel 8300 . In some embodiments , the large example , this can be a hollow fiber type module . In one diameter synthetic membrane vesicles suspension exiting embodiment, the hollow fiber filter used is Model No. the second retentate vessel 8200 can now be concentrated CFP - 2 - E -8A ( 0 . 2 micron ) manufactured by Amersham Bio - 200 % , whereas the buffer solution concentration in the large sciences of Westborough , Mass . In some embodiments , the diameter synthetic membrane vesicles suspension can be pore size of the cross - flow ( tangential- flow ) filtration mod - 40 roughly 9 . 1 % with respect to the large diameter synthetic ules 8150 , 8152 and 8154 , can be chosen to retain the large membrane vesicles suspension in the inlet line 8120 . diameter synthetic membrane vesicles while allowing the In some embodiments , similar filtration can occur in the suspending medium to pass through the filter membranes as final retentate vessel 8300 as described above for the first permeate . The pumps used in the diafiltration system can be retentate vessel 8100 and second retentate vessel 8200 . In of various types , such as peristaltic or rotary lobe positive 45 some embodiments , the concentrated large diameter syn displacement pumps. In some embodiments , cross - flow thetic membrane vesicles suspension traveling through the recirculation pumps 8110 , 8112 and 8114 operate with at feed line 8124 can enter the final product vessel 8300 where least twice the permeate flow rate of their associated cross - it is further filtered and concentrated to contain all the large flow filter module and preferably 3 times , 5 times or 10 times diameter synthetic membrane vesicles entering through the the permeate flow rates . In some embodiments , the permeate 50 inlet line 8120 , in the final product vessel 8300 . In some can be drawn off through a permeate line 8160 (passing embodiments , a solution can be fed into the final retentate through a sterilizing hydrophilic filter 8190 and a manual vessel 8300 through a line 8132 , the solution first passing valve 8202 ), wherein for each 1 L of large diameter synthetic through a manual valve 8208 and a sterilizing hydrophilic membrane vesicles suspension added to the first retentate filter 8174 , at a rate of 1 .75 L per 0 . 50 L concentrated large vessel 8100 , 2 . 25 L of permeate can be removed and 55 diameter synthetic membrane vesicles suspension added to discarded . In some embodiments , the retentate from the the final retentate vessel 8300 . In some embodiments , the filtration module 8150 can be circulated back into the first large diameter synthetic membrane vesicles suspension can retentate vessel 8100 via a retentate line 8140 . In some be pumped through the pump 8114 to the cross - flow filtra embodiments , for each 1 L of large diameter synthetic tion module 8154 , wherein the permeate can be drawn off to membrane vesicles suspension added to the first retentate 60 be discarded through a permeate line 8164 ( passing through vessel 8100 , a 0 .75 L flow of the concentrated large diameter a sterilizing hydrophilic filter 8194 and a manual valve synthetic membrane vesicles suspension can be removed 8210 ) at a rate of 2 . 25 L per 0 . 5 L of large diameter synthetic from the first retentate vessel 8100 through a feed line 8122 membrane vesicles suspension fed to the final product vessel and a metering pump 8123 to be further filtered in a second 8300 . In some embodiments , in the final product vessel retentate vessel 8200 . In some embodiments , the large 65 8300 , the large diameter synthetic membrane vesicles sus diameter synthetic membrane vesicles suspension exiting pension can contain a buffer concentration . In some embodi the first retentate vessel 8100 can be concentrated by a factor ments , the buffer concentration can be as low as 2 % . US 9 ,730 , 892 B2 59 60 Filters 8170 , 8190 , 8172 , 8192 , 8174 , and 8194 are approximately 39 . 7 um . In a typical embodiment , the large sterilizing hydrophilic filters . Filters 8180 , 8182 , and 8184 diameter synthetic membrane vesicles droplet 930 can be a are sterilizing hydrophobic gas vent filters used in the multivesicular liposomes droplet . In this embodiment, the retentate vessels and fed by gas lines 8131, 8133, and 8135 , first component core then is approximately 31. 5 um and the respectively . In some embodiments, the systems depicted in 5 dextrose /lysine shell 924 can be approximately 4 . 1 um thick . FIG . 1A through FIG . 8 can be operated in a sterile (aseptic ) When the organic solvent is removed from this atomized fashion ). For example , the addition of appropriate steam droplet 922 , the resultant MVL 930 is 25 um in diameter ( the lines , condensate drain lines and valves , can allow the dextrose /lysine shell 924 is omitted for clarity ) . In some system to be sterilized . In some embodiments , all inputs and embodiments , chambers 936 of the MVL 930 can have the outputs are equipped with sterile barrier filters . In a typical 10 same volume as the suspension of droplets in the continuous embodiment , the large diameter synthetic membrane phase of the first component. vesicles are multivesicular liposomes . FIG . 10 is a schematic of one example of a continuous In some embodiments , a continuous - flow particle concen - flow diafiltration system 1010 comprising a continuous flow tration system can be configured where a set of cascading centrifuge . The large diameter synthetic membrane vesicles particle suspension concentrators can be in series together 15 suspension can be pumped from a solvent removal vessel to adapted to remove and / or replace the suspending medium of the continuous - flow diafiltration system 1010 . a particle suspension in a continuous fashion . In some In some embodiments , the continuous- flow diafiltration embodiments , the cascading particle suspension concentra - system 1010 can include a solvent removal vessel as tors can be cross flow filters (FIG . 8 , components 8150 , depicted in FIG . 7 and as in component 70 of FIG . 1A and 8152 , 8154 ) and the suspending medium can be replaced by 20 FIG . 1B . The large diameter synthetic membrane vesicles diafiltration . In some embodiments, the cascading particle suspension can travel to the diafiltration system 1010 suspension concentrators can be hydro - cyclones or disk through a particle suspension inlet line 10120 ( also seen in centrifuge units . FIG . 7 , component 7270 ) , reaching a first retentate vessel FIG . 9 10100 . In some embodiments , for each 1 L of large diameter FIG . 9A provides cross -sectional views of an atomized 25 synthetic membrane vesicles suspension fed to the first droplet 902 , and a large diameter synthetic membrane retentate vessel 10100 , a 2 L solution can be fed into the first vesicles particle 912 . In a typical embodiment, the large retentate vessel 10100 through a line 10130 , the solution first diameter synthetic membrane vesicles particle 912 can be passing through a manual valve 10600 and a sterilizing formed by removal of an organic solvent from the atomized hydrophilic filter 10170 . In some embodiments , the large droplet 902 . In a typical embodiment , the atomized droplet 30 diameter synthetic membrane vesicles are multivesicular 902 can comprise a first component core and a buffer liposomes and the solution is a saline solution . solution shell 904 . In some embodiments , first component In some embodiments , a portion of the large diameter core can comprise a continuous phase 908 and a suspension synthetic membrane vesicles suspension in the first retentate of droplets 906 in the continuous phase 908 . In some vessel 10100 can be pumped by pump 10110 through a embodiments , the droplets 906 can be aqueous phase drop - 35 cross - flow ( tangential- flow ) filtration module 10150 . For lets and can be surrounded by a continuous phase 908 that example , this can be a hollow fiber type module . In one can be an organic solvent. In some embodiments , the aque - embodiment, the hollow fiber filter used is Model No . ous phase droplets can be from about 10 nm to about 10u in CFP - 2 - E - 10A ( 0 . 2 micron ) manufactured by Amersham diameter. In some embodiments , the aqueous phase droplets Biosciences of Westborough , Mass . In some embodiments , can be from about 500 nm to about 5u in diameter. In a 40 the pore size of the cross - flow ( tangential- flow ) filtration typical embodiment, the aqueous phase droplets can be from modules 10150 , 10152 and 10154 , can be chosen to retain about 100 nm to about 2u in diameter. For example the the large diameter synthetic membrane vesicles while allow average diameter of the aqueous phase droplets can be about ing the suspending medium to pass through the filter mem lu in diameter. In some embodiments , the organic solvent branes as permeate . The pumps used in the diafiltration can then be removed in the solvent removal chamber to 45 system can be of various types, such as peristaltic or rotary provide droplets of large diameter synthetic membrane lobe positive displacement pumps . In some embodiments , vesicles 912 within a shell of dextrose / lysine 914 . In a cross - flow recirculation pumps 10110 , 10112 and 10114 typical embodiment, the large diameter synthetic membrane operate with at least twice the permeate flow rate of their vesicles droplet 912 are a multivesicular liposomes droplet . associated cross - flow filter module and preferably 3 times, 5 Multivesicular liposomes MVL( ) are uniquely different 50 times or 10 times the permeate flow rates . In some embodi from other lipid -based drug delivery systems. Topologically, ments , the permeate can be drawn off through a permeate MVL are defined as liposomes containing multiple non - line 10160 (passing through a sterilizing hydrophilic filter concentric chambers 916 within each droplet 912 , resem - 10190 and a manual valve 10602 ) , wherein for each 1 L of bling a “ foam - like ” matrix . The chambers 916 of the MVL large diameter synthetic membrane vesicles suspension can have the same volume as the first component particles , 55 added to the first retentate vessel 10100 , 2 . 25 L of permeate ( e . g . lu ) as shown in FIG . 9A . The presence of internal can be removed and discarded . In some embodiments , the membranes distributed as a network throughout the MVL retentate from the filtration module 10150 can be circulated may serve to confer increased mechanical strength to the back into the first retentate vessel 10100 via a retentate line vesicle , while still maintaining a high volume: lipid ratio . 10140 . In some embodiments, for each 1 L of large diameter Thus, both structurally and functionally the MVL are 60 synthetic membrane vesicles suspension added to the first unusual, novel and distinct from all other types of lipo - retentate vessel 10100 , a 0 .75 L flow of the concentrated somes . large diameter synthetic membrane vesicles suspension can In one embodiment, a large diameter synthetic membrane be removed from the first retentate vessel 10100 through a vesicles droplet 930 made by the instant processes can be , as feed line 10122 and a metering pump 10123 to be further depicted in FIG . 9B , for example , an atomized droplet 922 65 filtered in a second retentate vessel 10200 . In some embodi containing equal volumes of dextrose / lysine 924 and first ments , the large diameter synthetic membrane vesicles sus component, where the diameter of the atomized droplet is pension exiting the first retentate vessel 10100 can be US 9 ,730 ,892 B2 62 concentrated by a factor of 1 .33 ( large diameter synthetic contain a buffer concentration . In some embodiments , the membrane vesicles concentration is increased by 33 % ). In a buffer concentration can be as low as 2 % . typical embodiment, the large diameter synthetic membrane In some embodiments , the large diameter synthetic mem vesicles are multivesicular liposomes. brane vesicles suspension traveling through a feed line In some embodiments , similar filtration can occur in the 5 10126 , can be metered by a pump 10127 and can enter a second retentate vessel 10200 as described above for the first continuous flow centrifuge module 10400 wherein the retentate vessel 10100 . In some embodiments , the large supernantant can be pumped to be discarded through a pump diameter synthetic membrane vesicles suspension can enter 10116 through a permeate line 10166 ( passing through a the second retentate vessel 10200 at a rate of 0 . 75 L per 1 L sterilizing hydrophilic filter 10196 ) . In a typical embodi added to the first retentate vessel 10100 . In some embodi- 10 ment , the large diameter synthetic membrane vesicles are ments , a solution can be fed into the second retentate vessel multivesicular liposomes. 10200 through a line 10132 , the solution first passing In some embodiments , the large diameter synthetic mem through a manual valve 10204 and a sterilizing hydrophilic brane vesicles suspension traveling through a feed line filter 10172 , at a rate of 2 L per 0 . 75 L concentrated large 10128 , can be metered by a pump 10129 and can enter a final diameter synthetic membrane vesicles suspension added to 15 retentate vessel 10500 . In a typical embodiment, the large the second retentate vessel 10200 . In some embodiments , a diameter synthetic membrane vesicles are multivesicular portion of the large diameter synthetic membrane vesicles liposomes . suspension in the second retentate vessel 10200 can be Filters 10170 , 10190 , 10172 , 10192, 10174 , 10194, and pumped by pump 10112 through a cross - flow ( tangential - 10196 are sterilizing hydrophilic filters. Filters 10180 , flow ) filtration module 10152. In some embodiments, the 20 10182 , 10184 and 10186 are sterilizing hydrophobic gas permeate can be drawn off through a permeate line 10162 vent filters used in the retentate vessels and fed by gas lines ( passing through a sterilizing hydrophilic filter 10192 and a 10131 , 10133 , 10135 and 10137 , respectively . manual valve 10606 ) , wherein for each 0 . 75 L of large FIG . 11 is a schematic of one example of a continuous diameter synthetic membrane vesicles suspension added to flow centrifuge system 1110 comprising a plurality of con the second retentate vessel 10200 , 2 . 25 L of permeate can be 25 tinuous flow centrifuges. The large diameter synthetic mem removed and discarded . In some embodiments , the retentate brane vesicles suspension can be pumped from a solvent from the filtration module 10152 can be circulated back into removal vessel to the continuous - flow centrifuge system the second retentate vessel 10200 via a retentate line 10142 . 1110 . In some embodiments , for each 0 .75 L of large diameter I n some embodiments , the continuous - flow centrifuge synthetic membrane vesicles suspension added to the second 30 system 1110 can include the solvent removal vessel as retentate vessel 10200 , a 0 . 50 L flow of the concentrated depicted in FIG . 7 . The large diameter synthetic membrane large diameter synthetic membrane vesicles suspension can vesicles suspension can travel to the centrifuge system 1110 be removed from the second retentate vessel 10200 through through a particle suspension inlet line 11120 (also seen in a feed line 10124 and a metering pump 10125 to be further FIG . 7 , component 7270 , and as represented by components filtered in a third retentate vessel 10300 . In some embodi- 35 70 in FIG . la and FIG . 1B ) , reaching a first retentate vessel ments , the large diameter synthetic membrane vesicles sus - 11100 . In some embodiments , a large diameter synthetic pension exiting the second retentate vessel 10200 can now membrane vesicles suspension can be fed to the first reten be concentrated 200 % , whereas the buffer solution concen - tate vessel 11100 , and a solution can be fed into the first tration in the large diameter synthetic membrane vesicles retentate vessel 11100 through a line 11130 , the solution first suspension is roughly 9 . 1 % with respect to the large diam - 40 passing through a manual valve 11600 and a sterilizing eter synthetic membrane vesicles suspension in the inlet line hydrophilic filter 11170 . The large diameter synthetic mem 10120 . brane vesicles suspension in the first retentate vessel 11100 In some embodiments , similar filtration can occur in the can be pumped by a pump 11220 to a first centrifuge module third retentate vessel 10300 as described above for the first 11150 . In some embodiments , the permeate can be drawn off retentate vessel 10100 and second retentate vessel 10200 . In 45 from the first centrifuge 11150 through a permeate line some embodiments , the concentrated large diameter syn - 11160 (passing through a sterilizing hydrophilic filter 11190 thetic membrane vesicles suspension traveling through the being pumped by a pump 11110 ), and discarded . In some feed line 10124 can enter the third product vessel 10300 embodiments , large diameter synthetic membrane vesicles where it is further filtered and concentrated to contain all the suspension can exit the first centrifuge module 11150 large diameter synthetic membrane vesicles entering 50 through a feed line 11122 , metered by a pump 11123 , to be through the inlet line 10120 , in the third product vessel further processed in a second retentate vessel 11200 . In some 10300 . In some embodiments , a solution can be fed into the embodiments , the large diameter synthetic membrane third retentate vessel 10300 through a line 10132 , the vesicles suspension flowing in feed line 11122 can be solution first passing through a manual valve 102010 and a concentrated by at least 33 % . In a typical embodiment, the sterilizing hydrophilic filter 10174 , at a rate of 1 .75 L per 55 large diameter synthetic membrane vesicles are multivesicu 0 .50 L concentrated large diameter synthetic membrane lar liposomes and the solution is a saline solution . vesicles suspension added to the third retentate vessel In some embodiments , the concentrated large diameter 10300 . In some embodiments , the large diameter synthetic Synthetic membrane vesicles suspension can enter the sec membrane vesicles suspension can be pumped through the ond retentate vessel 11200 for further processing , and a pump 10114 to the cross - flow filtration module 10154 , 60 solution can be fed through a line 11132 , the solution first wherein the permeate can be drawn off to be discarded passing through a manual valve 11604 and a sterilizing through a permeate line 10164 (passing through a sterilizing hydrophilic filter 11172 . The large diameter synthetic mem hydrophilic filter 10194 and a manual valve 10610 ) at a rate brane vesicles suspension from the second retentate vessel of 2 .25 L per 0 . 5 L of large diameter synthetic membrane 11200 can be pumped by a pump 11240 to a second vesicles suspension fed to the third product vessel 10300 . In 65 centrifuge module 11152. In some embodiments , the per some embodiments , in the third product vessel 10300 , the meate can be drawn off from the second centrifuge 11152 large diameter synthetic membrane vesicles suspension can through a permeate line 11162 (passing through a sterilizing US 9 , 730 , 892 B2 63 64 hydrophilic filter 11192 being pumped by a pump 11120 ) , In some embodiments , the multivesicular liposomes fur and discarded . In some embodiments , large diameter syn ther comprise bupivaciane , dextrose , DEPC , DPPG , and thetic membrane vesicles suspension can exit the second tricaprylin . In some embodiments , the multivesicular lipo centrifuge module 11152 through a feed line 11124 , metered somes further comprise bupivacaine phosphate , dextrose , by a pump 11125 , to be further processed in a third retentate 5 DEPC , DPPG , and tricaprylin . In some embodiments, the vessel 11300 . In some embodiments , the large diameter multivesicular liposomes further comprise bupivacaine , synthetic membrane vesicles suspension flowing in feed line dextrose , DEPC , DPPG , tricaprylin and cholesterol. In some 11125 can be concentrated by at least 33 % . In a typical embodiments , the multivesicular liposomes further com embodiment, the large diameter synthetic membrane prise bupivacaine phosphate , dextrose , DEPC , DPPG , tri vesicles are multivesicular liposomes and the solution is a 10 caprylin and cholesterol. saline solution . In some embodiments , the multivesicular liposomes fur In some embodiments , the large diameter synthetic mem - ther comprise bupivaciane , L -Lysine , DEPC , DPPG , and brane vesicles suspension traveling through the feed line tricaprylin . In some embodiments , the multivesicular lipo 11124 , can be metered by a pump 11125 and can enter the somes further comprise bupivacaine phosphate , L -Lysine , third retentate vessel 11300 where it can be further pro - 15 DEPC , DPPG , and tricaprylin . In some embodiments , the cessed . In some embodiments , a solution can be fed into the multivesicular liposomes further comprise bupivacaine , third retentate vessel 11300 through a line 11134 , the solu - L - Lysine , DEPC , DPPG , tricaprylin and cholesterol. In tion first passing through a manual valve 11608 and a some embodiments , the multivesicular liposomes further sterilizing hydrophilic filter 11174 . In some embodiments, comprise bupivacaine phosphate , L -Lysine , DEPC , DPPG , the large diameter synthetic membrane vesicles suspension 20 tricaprylin and cholesterol. can be pumped by a pump 11260 to a third centrifuge In some embodiments , the multivesicular liposomes fur module 11154 . In some embodiments , the permeate can be ther comprise bupivacaine , morphine , cytarabine, or their drawn off from the centrifuge through a permeate line 11164 pharmaceutically acceptable salts as the therapeutic agent. (passing through a sterilizing hydrophilic filter 11194 being In some embodiments , the multivesicular liposomes further pumped by pump 11130 ) , and discarded . In a typical 25 comprise bupivacaine phosphate , morphine sulfate , or cyt embodiment, the large diameter synthetic membrane arabine HCI. vesicles are multivesicular liposomes . In another embodiment, any one of the above described In some embodiments , the concentrated large diameter embodiments can be used alone or in combination with any synthetic membrane vesicles suspension traveling through one or more of the above described embodiments. For feed line 11166 , can be metered by pump 11127 and can 30 example , any above described atomizing nozzle, evapora enter a final retentate vessel 11400 . In a typical embodiment, tion apparatus , continuous - flow emulsification system , con the large diameter synthetic membrane vesicles are multi tinuous- flow diafiltration system , continuous - flow diafiltra vesicular liposomess . tion further comprising one ormore centrifuges, continuous Filters 11170 , 11190 , 11172 , 11192 , 11174 , and 11194 are flow centrifuge system , or continuous processing system can sterilizing hydrophilic filters . Filters 11180 , 11182 , 11184 35 be used alone or in combination . Thus , an evaporation and 11186 are sterilizing hydrophobic gas vent filters used in apparatus can be used in conjunction with a three - fluid the retentate vessels and fed by gas lines 11131, 11133 , atomizing nozzle . This evaporation system /atomizing nozzle 11135 and 11137 , respectively. In a typical embodiment, the can be used with a continuous - flow emulsification system , large diameter synthetic membrane vesicles are multivesicu - as depicted in FIGS. 1A , 1B , and 1C . The three -fluid lar liposomes . 40 atomizing nozzle / evaporation apparatus combination can be The systems depicted in FIG . 1A through FIG . 11 can be used in conjunction with a continuous - flow system , as operated in a sterile (aseptic ) fashion . With the addition of depicted in FIGS. 8 , 10 , and 11 . Any of these combinations appropriate steam lines, condensate drain lines and valves , can be used to make multivesicular liposomes . In particular the system can be sterilized . All inputs and outputs are any of the combinations can be used to make multivesicular equipped with sterile barrier filters . 45 liposomes containing bupivacaine or its salts as the thera In some embodiments, the large diameter synthetic mem - peutic agent. brane vesicles are multivesicular liposomes . In some The following examples are meant to further illustrate the embodiments , the multivesicular liposomes further com - embodiments , they are not meant to be limiting in any way . prise bupivaciane , DEPC , DPPG , and tricaprylin . In some embodiments , the multivesicular liposomes further com - 50 EXAMPLE 1 prise bupivacaine phosphate , DEPC , DPPG , and tricaprylin . In some embodiments , the multivesicular liposomes further The following is an example utilizing the process param comprise bupivacaine , DEPC , DPPG , tricaprylin and cho - eters and steps of the devices depicted in the Figures. The lesterol. In some embodiments, themultivesicular liposomes three fluids applied to the atomizing nozzle ( FIG . 1A and further comprise bupivacaine phosphate, DEPC , DPPG , 55 FIG . 1B , component 75 ; FIG . 3A , component 310 ; FIG . 7 , tricaprylin and cholesterol. component 7510 ) as part of the process of forming multi In some embodiments , the multivesicular liposomes fur- vesicular liposomes have the following compositions per ther comprise bupivaciane , dextrose , L - Lysine , DEPC , liter . DPPG , and tricaprylin . In some embodiments , the multive The first fluid (FIG . 3A -3L , component 3115 ; FIG . 5 , sicular liposomes further comprise bupivacaine phosphate , 60 component 5115 ; FIG . 7 , component 7115 ) was a first liquid dextrose , L - Lysine , DEPC , DPPG , and tricaprylin . In some made up of the first component, the first component having embodiments , the multivesicular liposomes further com - two components : an organic phase and a firstaqueous phase prise bupivacaine , dextrose , L - Lysine , DEPC , DPPG , tri - which are emulsified with equal volumes. The organic phase caprylin and cholesterol. In some embodiments , the multi - was composed of 1 , 2 - dierucoyl- sn - glycero - 3 -phosphocho vesicular liposomes further comprise bupivacaine 65 line ( 17 . 78 g ) , 1 , 2 - dipalmitoyl- sn - glycero - 3 - phosphoglyc phosphate , dextrose , L - Lysine , DEPC , DPPG , tricaprylin e rol ( 1 .056 g ) , cholesterol (10 . 34 g ) , tricaprylin (4 .32 g ) , and cholesterol. water ( 0 .70 g ) and methylene chloride ( quantity sufficient to US 9 ,730 ,892 B2 65 66 make 1 L total volume of the organic phase. The first 1A AND FIG . 1B , component 50 ; FIG . 7 component 710 ) aqueous phase was composed of 0 . 2 molar (200 mM and were each centered 10 . 2 cm from the vessel wall ( FIG . phosphoric acid and bupivacaine (40 g ) and water ( quantity 7 , component 7350 ) . The lid protection gas inlet (FIG . 7 , sufficient to make 1 L total volume of the first aqueous component 7430 ) had an inside diameter of 0 . 95 cm , extends phase ) . 5 through the lid of the vessel ( FIG . 7 , component 7220 ) , and The second fluid (FIG . 3A - 3L , component 3120 ; FIG . 5 , faced parallel to the inside of the lid in a direction to give component 5120 ; FIG . 7 , component 7120 ) was a second clockwise (viewed down from above ) rotation of the gas. liquid made up of a second aqueous phase composed of Starting with the three - fluid atomizing nozzle ( FIG . 1A and L -lysine (monohydrate ) ( 16 . 8 g ), dextrose ( 13 . 25 g ) , and FIG . 1B , component 75 ; FIG . 3A , component 310 ; FIG . 7 , water (quantity sufficient to make 1 L total volume of the 10 component 7510 ) as zero degrees, the main carrier gas inlet second fluid . line was located 52 degrees clockwise , the rinse nozzle was The third fluid (FIG . 3A -3L , component 3140; FIG . 5 , 90 degrees clockwise , and the lid protection gas inlet was component 5140 ; FIG . 7 , component 7140 ) was nitrogen gas 135 degrees clockwise with its outlet approximately 180 containing water vapor ( 100 % relative humidity at 42° C . degrees clockwise . and 25 psig ). 15 The sides ( FIG . 7, component 7350 ) and bottom (FIG . 7 , Preparation of Solvent Evaporation Chamber component 7250 ) of the solvent removal vessel were jack Nitrogen was supplied to the solvent removal vessel (FIG . eted at 24 . 1° C . by connection to a circulating bath . The 1A AND FIG . 1B , component 50 ; FIG . 7 component 710 ) temperature in the jacket was adjusted to approximately through two solvent removal vessel gas inlet lines: FIG . 1A match the steady state gas exit temperature . Such a match components 115 and 110 ; FIG . 7 components 7280 and 20 prevented evaporation and drying of the multivesicular 7430 ) the main carrier gas inlet line (component 115 of FIG . liposomes on the wall or condensation of water that could 1A AND FIG . 1B ; component 7280 of FIG . 7 ) , or main rupture , through osmotic shock , the multivesicular lipo rotation jet, was tangential to vessel wall and approximately somes that are being formed in the vessel. 40 % up from vessel bottom , causing clockwise rotation Preparation of a First Component viewed from above ; and the lid protection gas inlet (com - 25 The recirculation loop ( FIG . 2 , component 2125 ) con ponent 110 of FIG . 1A AND FIG . 1B ; component 7430 of nected to the high - shear mixer (FIG . 1A AND FIG . 1B , FIG . 7 ), or lid protection jet, at the corner of lid ( component component 25 , FIG . 2 , component 2130 ) (Ross model 7220 of FIG . 7 ) and vessel wall ( component 7350 of FIG . 7 ) , HSM -703XS - 20 Sanitary Inline High Shear Mixer equipped tangential to the wall and moving in the same rotational with a 3 " diameter X - 5 Series rotor / stator for operation to direction ( component 7240 of FIG . 7 ) . The nitrogen entering 30 14 , 400 rpm . (11 , 300 feet /min . tip speed ) with gap ring # 3 ) these inlet lines was humidified to 100 % relative humidity at was primed with methylene chloride to ensure that all air 42° C . and at 25 psig ( pounds per square inch gauge ). The was removed from the high - shear mixer. The jacket of the main rotation jet supplies 335 L /min at 42° C . of humidified heat exchanger (FIG . 1A AND FIG . 1B , component 30 ; FIG . nitrogen while the lid protection jet flow was 25 L / min at 42° 2 component 2170 ) was supplied with 5° C . coolant ( water + C . of humidified nitrogen ( the lid protection jet keeps 35 50 % ethylene glycol) (FIG . 1A , components 96 and 97; FIG . deposit buildup off the lid ) . The nitrogen was humidified 2 , components 2110 and 2105 ). The mixer seal lubricant prior to entering the solvent removal vessel by passing it tank (FIG . 1A AND FIG . 1B , component 10 ) , filled with through a heated tube - in -shell heat exchanger ( component water, was also cooled with 5° C . coolant (water + 50 % 90 , FIG . 1A AND FIG . 1B ) that was coated with water ethylene glycol ) . The mixer was started at a setting of 30 Hz (humidification water ) followed by an excess - liquid water 40 ( approx. 7 , 200 rpm ) causing a flow around the mixer loop removal chamber component 45 , FIG . 1A AND FIG . 1B ) estimated at 21 , 000 mL /min and with an internal volume of and liquid bleed . The nitrogen was humidified to reduce 280 mL . Thus, fluid in the loop went through the mixer evaporation of water from the spray which raised the osmo - blades and heat exchanger an average of every 0 . 8 seconds . lality of the suspending buffer. After the high shear mixer was primed with methylene In this example , the solvent removal vessel had a volume 45 chloride , the organic phase and first aqueous phase peristal of approximately 138 liters , the inside diameter was 56 cm , tic pumps ( components 12 and 2 , respectively, of FIG . 1A the walls were 52 cm high and the dome at the bottom was AND FIG . 1B ) were concurrently started . The organic phase 10 cm deep . Thus the inside height of the solvent removal was pumped at 33 mL /min and the first aqueous phase was vessel was 62 cm from the lid to the bottom of the domed also pumped at 33 mL /min . The organic phase and first bottom . The gas outlet tube ( component 80 , FIG . 1A AND 50 aqueous phase entered the high - shear mixer starting the FIG . 1B ; component 7340 , FIG . 7 ) ( including the conical formation of the first component. As the first component fitting FIG . 7 , component 300 and vortex stabilizer FIG . 7 , circulated around the high shear mixer recirculation loop , a component 7360 ) extends 42 . 5 cm into the solvent removal small fraction of the flow was forced through the first vessel down from lid . The diameter of the gas outlet tube component exit line (FIG . 2 , component 2180 ; FIG . 5 , was 2 . 3 cm ( inside diameter ) and the conical fitting tapers 20 55 component 5114 ) to the three - fluid atomizing nozzle ( FIG . degrees to a 1 . 5 cm inside diameter for the gas outlet orifice 1A AND FIG . 1B , component 75 ; FIG . 3A - 3L , component ( FIG . 7 , component 7310 ). The vortex stabilizer attached to 310 ; FIG . 5 , component 505 ; FIG . 7 , component 7510 ) at 66 the end of the conical fitting had a diameter of 2 . 5 cm . The mL /min ( total of the two flow rates ). Consequently, the main rotation (carrier ) gas inlet was tangential to the wall priming methylene chloride was soon flushed from the and 37 cm down from the lid and had an inside diameter of 60 mixer loop by this flow ( 4 .2 min per loop volume of flush ) . 1 . 9 cm . Concurrent with the starting of the organic phase and first The three - fluid atomizing nozzle ( FIG . 1A and FIG . 1B , aqueous phase peristaltic pumps , the second aqueous phase component 75 ; FIG . 3A , component 310 ; FIG . 7 , component ( connected to the three - fluid atomizing nozzle ) and wall 7510 ), the rinse nozzle (FIG . 1A AND FIG . 1B , component rinse ( to the rinse nozzle ( FIG . 1A and FIG . 1B , component 105 ; FIG . 7 , component 7400 ) , and lid protection gas inlet 65 105 ; FIG . 7 , component 7400 ) ) peristaltic pumps ( compo (FIG . 1A , component 110 ; FIG . 7 , component 7340 ) all nents 22 and 64 , respectively , of FIG . 1A and FIG . 1B ) were extend through the lid of the solvent removal vessel (FIG . started and pumped their respective components at 66 US 9 , 730 ,892 B2 68 mL /min each . The wall rinse solution was 33 . 5 g of dextrose EXAMPLE 2 per liter ofwater . Nitrogen at 60 psig (room temperature , not humidified ) was supplied to these two nozzles (FIG . 1A and Preparation of First Component FIG . 1B , components 75 and 105 ; FIG . 7 , components 7510 The recirculation loop connected to the high - shear mixer and 7400 ). The second aqueous phase flowed through the 5 ( FIG . 1A and FIG . 1B , component 25 ; FIG . 2 , component three fluid nozzle at 66 mL /min . The three - fluid atomizing 2130 ) (Ross Model HSM - 703XS - 20 Sanitary Inline High nozzle had a nitrogen flow rate of 51 L /min @ 1 atm and the Shear Mixer equipped with a 3 " diameter X - 5 Series rotor / wall rinse nozzle (manufactured by GEA Process Engineer stator for operation to 14 ,400 rpm . ( 11, 300 feet /min . tip ing of Columbia , Md .) had a flow rate of 66 L /min @ 1 atm . speed ) with gap ring # 3 ) was primed with methylene chlo After exiting the three - fluid atomizing nozzle , the formed 10 ride to ensure that all air was removed from the high -shear atomized emulsion droplets came into contact with the mixer. The jacket of the heat exchanger ( FIG . 1A and FIG . carrier gas (nitrogen ) in the solvent removal chamber ( FIG . 1B , component 30 ; FIG . 2 , component 2170 ) was supplied 7 , component 7230 ) of the solvent removal vessel (FIG . 1A with 5° C . coolant (water + 50 % ethylene glycol) . The mixer and FIG . 1B , component 50 ; (FIG . 7 , component 710 ). The 15 seal lubricant tank , filled with water , was also cooled with 5° carrier gas rotated inside the chamber (FIG . 7 , component C . coolant (water + 50 % ethylene glycol) . The high - shear 7240 ) as a small, rapidly rotating , intense gas vortex ( FIG . mixer was started at a setting of 25 Hz (6 , 000 rpm ), 30 Hz 7 , component 7245 ) formed at the exit orifice ( FIG . 7, approx . ( 7 ,200 rpm ) or 35 Hz (8 ,400 rpm ). component 7310 ). This allowed the droplets to contact the After the high shear mixer was primed with methylene gas for an extended period of time in order to effectuate 20 chloride, the organic phase and first aqueous phase peristal methylene chloride evaporation and removal. After removal tic pumps (FIG . 1A and FIG . 1B , components 12 and 2 , of the methylene chloride , the formed multivesicular lipo - respectively ) were concurrently started . The organic phase was pumped at 33 mL /min and the first aqueous phase was some suspension droplets ( FIG . 7 , component 7380 ) were also pumped at 33 mL /min . Entry of the organic phase and collected as a suspension of multivesicular liposomes ( FIG . first aqueous phase begin formation of the first component. 7 , component 7390 ) at the bottom of an evaporation vessel 25 As the first component circulates around the high shear ( FIG . 1A and FIG . 1B , component 50 ; FIG . 7 , component mixer recirculation loop (FIG . 2 , component 2125 ) , a small 7250 ) . The multivesicular liposomes were collected in the fraction of the flow was forced through the first component solvent removal vessel and then drained out of the drain port exit line ( FIG . 2 , component 2180 ) to the three - fluid atom ( FIG . 1A and FIG . 1B , component 130 ; FIG . 7 , component izing nozzle ( FIG . 1A and FIG . 1B , component 75 ; FIG . 3A ; 7270 ) through a peristaltic positive displacement pump 30 FIG . 4A ; FIG . 7 7510 ) at 66 mL /min ( total of the two flow rates) , the temperature of the emulsion being forced through (FIG . 1A and FIG . 1B , component 125 ), set to pump slightly the first component exit line (FIG . 2 , component 2180 ) was faster ( approximately 200 mL /min ) than the suspension from 16 .5 to 21. 1° C . The flow of the organic phase and first drains out of the bottom (approximately 165 mL / min ), aqueous rapidly flushed the priming methylene chloride therefore the exit stream of multivesicular liposome suspen - 35 from the mixer loop ( 4 . 2 min per loop volume of flush ) . sion were periodically interrupted by small segments of chamber gas. This pump rate prevented any appreciable TABLE 1 venting of solvent vapors into the room and protects the multivesicular liposome suspension from exposure to high Organic Phase Components (per 2 L Total Volume) velocity gas streams or foaming . 40 cholesterol (Nippon ) 20 . 8 g At system equilibrium , the nitrogen exiting the solvent DEPC (Nippon ) 36 .0 g DPPG ( Lipoid ) 1 .89 g removal vessel through the gas outlet ( FIG . 1A and FIG . 1B , tricaprylin (NOF ) 8 .81 g component 80 ; FIG . 7 , component 7310 ) was at a tempera Water for injection 0 .49 mL ture of approximately 21. 5° C . The solvent removal vessel Methylene chloride (EMD ) 2 ,569 g jacket was cooled to approximately 24 .1° C . The tempera - 45 ture of the first component (part of which travels to the The components of the organic phase are shown in Table three - fluid atomizing nozzle ) leaving the heat - exchanger 1 . was 15 . 3° C . After traveling through heat exchanger, and while traveling back to the high -shear mixer through the TABLE 2 recirculation line, the temperature of the first component was 30 approximately 14 .3° C . First Aqueous Phase Components (per 2 L Total Volume ) At equilibrium the system then ran continuously making Bupivacaine (BASF ) 80 g multivesicular liposomes for as long as the feed solutions 0. 20M H3PO4 (2 L , Mallinckrodt) 200 mM and nitrogen were supplied . Five hundred mL samples of sa multivesicular liposome suspension were taken from the 55 The components of the first aqueous phase are shown in suspension outlet ( FIG . 7 , component 7260 ) in the solvent Table 2 . removal vessel and diafiltered (using a hollow fiber filter, Model No. CFP - 2 - E -8A from Amersham Biosciences of TABLE 3 Westborough , Mass. ) with four volumes of normal saline on 60 a small scale batch diafiltration system ( FIG . 1A and FIG . Particle Sizing of Diluted Emulsion Samples by laser light scatter analysis . 1B , component 70 ) . Optionally , settling of the multivesicu PSD ( um ) ( volume based ) lar liposomes and excess liquid could be decanted to obtain the final multivesicular liposomes in a chosen aqueous Batch dio dso d90 Span solution at a chosen MVL concentration . Full continuous 65 Ross 30 Hz 0 . 5 0. 9 11 . 4 1 . 0 operation can be obtained by connecting the solvent removal Ross 35 Hz 0 . 5 0 .. 8 11 . 3 11 . 0 vessel outlet pump to the apparatus of FIG . 8 . US 9 , 730 , 892 B2 69 70 TABLE 3 -continued approximately 15 mg of bupivacaine per mL in the final multivesicular liposomes suspension . Particle Sizing of Diluted Emulsion Samples by laser light scatter analysis .

PSD ( um ) ( volume based ) TABLE 5 BatchBatch dio d 50 d 9o Span Wall Rinse Solution (per 5 L Total Volume) Ross 25 Hz 0 . 6 1 . 1 1 . 7 1 . 0 50 % Dextrose soln ( B Braun ) 335 mL Batch Lot D 0 . 855 1 . 151 1 . 506 0 . 566 Deionized water ( to final volume) 5 L Batch Lot E 0 .720 1 . 110 1 .530 0 . 730 As can be seen in FIG . 12 , controlled release of bupiva The emulsion samples were diluted and analyzed using a caine from MVL particles was examined in vitro at 37° C . light scattering device (Horiba Instruments La -910 ) and the in 0 . 5 % ovine serum albumin dissolved in 50 mM phosphate resultsre are shown in Table 1 . buffered saline (pH7 ) showing a similar release of bupiva The second aqueous phase ( connected to the three - fluid 15 caine to that in Batch Lot A and B made by the process atomizing nozzle ) and wall rinse ( to the rinse nozzle ( FIG . disclosed in patent WO 99 /25319 . 1A and FIG . 1B , component 105 ; FIG . 7 , component 7400 ) ) As can be seen in FIG . 13 , the PK profile in rats of the peristaltic pumps (components 22 and 64 , respectively , of continuous process samples shows a similar sustained FIG . 1A and FIG . 1B ) were started when the organic phase release profile of bupivacaine to that in Batch Lot C made by and first aqueous phase peristaltic pumps were started . The 20 the batch process disclosed in patent WO 99 / 25319 . wall rinse solution having 33 . 5 g of dextrose per liter of water was introduced to the evaporation chamber at a flow TABLE 6A rate of 66 mL /min . Nitrogen at 60 psig ( room temperature , Final Material Properties not humidified ) was supplied to the atomizing nozzle . The second aqueous phase was introduced to the three fluid 25 Total Bupi? Free nozzle at a flow rate of 66 mL /min . The three - fluid atom (mg % Bupi % PSDili (um ) pH izing nozzle has a nitrogen flow rate of 51 L /min @ 1 atm and the wall rinse nozzle (manufactured by GEA Process Batch mL ) PPVii (mg / mL ) Free d10 050 090 Int Ext Engineering of Columbia , Md. ) has a flow rate of 66 30 Hz 14 . 9 41 0 . 79 3 . 2 18. 0 44 . 7 102. 8 5 . 7 7 . 0 L /min @ 1 atm . The emulsion , second aqueous phase and Batch nitrogen gas were combined using the three fluid nozzle to 35 Hz 13. 5 38 0 .81 3 . 7 14 . 2 36 . 3 83. 2 6 .0 7. 3 afford atomized droplets ( FIG . 3 , component 3155 ; FIG . 7 , Batch component 7155 ) which traveled in the evaporation chamber Batch25 Hz 12 .8 38 0 .83 4 .0 14832 . .4 69. 4 5. 9 7 .4 until the majority of the methylene chloride was removed 25 from the atomized droplets . Removal of the methylene chloride produced multivesicular liposomes (FIG . 7 , com ponent 7380 ) which form a suspension (FIG . 7 , component TABLE 6B 7390 ) at the bottom of the evaporation chamber (FIG . 7 , Final Material Properties (cont ' d ) component 7250 ) . 40 Total Lipids Batch Choliv DEPC DPPGvi TCvii TABLE 4 30 Hz 3 . 24 5. 98 0 . 23 1 . 50 Second Aqueous Phase Components (per 5 L Total Volume) Batch 5. 98 1. 50 35 Hz 3 . 70 6 . 72 0 . 31 11 .61 L - Lysine monohydrate 84 g 45 Batch 50 % Dextrose soln ( B Braun ) 132. 5 mL 25 Hz 3 . 30 6 . 15 0 . 22 1 . 51 Deionized water ( to final volume) 5 L Batch The Components of the Second Aqueous Phase are Shown i ) bupivacaine; ii ) packed particle volume; iii ) particle in Table 4 . 50 size distribution by mass ; After system equilibrated ( 10 minutes) , 500 mL samples iv ) cholesterol; v ) 1 , 2 -dierucoyl - sn -glycero - 3 -phospho of multivesicular liposomes suspension were collected . The choline ; Ross RPM was then changed and allowed to equilibrate for vi) 1 ,2 -dipalmitoyl - sn - glycero -3 -phosphoglycerol ; vii ) 10 minutes before collecting the next 500 mL sample . This tricaprylin was repeated for the last RPM . Each 500 mL batch was 55 diafiltered in a batch mode (using a hollow fiber filter, Model EXAMPLE 3 No. CFP - 6 - D - 9A from Amersham Biosciences of Westbor ough , Mass . ), for four volumes with normal saline on a small Heat Treatment of MVL Suspension scale batch diafiltration system (FIG . 1 , component 70 ) . The The system of FIG . 1B was used with the humidified recirculation pump was set to 5 ,700 ml/min , while saline 60 rotation gas (N2 ) supplied by combination electric heater addition pump was set to 410 mL /min . The permeate valve and tube- in shell heat exchanger as described for FIG . 1A , was adjusted to keep the liquid volume in the diafiltration component 90 . The system was equilibrated for 10 minutes system constant at 1, 000 and thus was also 410 mL /min , and a 1 , 000 ml sample ofMVL suspension , exiting the drain since the minimum working volume of this system was port ( FIG . 1B , component 130 ) of the solvent removal vessel approximately 550 mL . In the present example , the pro - 65 50 , was collected . The MVL sample was divided into two cessed multivesicular liposomes suspension was allowed to samples of 500 mL each . The first 500 mL MVL sample was settle at 5° C . and then supernatant was decanted to achieve heat treated as follows . The heat treatment was performed by US 9 ,730 , 892 B2 rapidly adding 750 mL of 100° C . dextrose solution to the bution of the MVLs and had only small effects on the first sample to raise the mixture temperature up to approxi- bupivacaine (active agent) content and lipid composition of mamately 63° C . After 30 seconds, 1 ,750 mL of + 5° C . saline the particlesparticles . was rapidly added to lower the temperature of themixture to near room temperature ( 35° C . or below ) . The sample 5 The heat treatment did have a surprising effect on the volume was now 3 , 000 mL . The second 500 mL multive accelerated stability (30° C . stability ) of these MVL suspen sicular liposomes sample was not heat treated . The second sions , as can be seen in FIG . 14 . Accelerated stability at 30° sample was diluted with the same volumes of dextrose C . was greatly improved with heat treatment. A lower slope solution ( 750 mL) and saline ( 1 ,750 mL ) as the first sample means less bupivacaine release and longer stability . Both but the solutions were at room temperature . samples were usable products when stored at + 5° C . but the Each sample was diafiltered batch wise , with 4 volumes heat treated sample was projected to have a much longer of saline . These 3 ,000 ml samples were each concentrated to shelf life . 1 liter and then diafiltered in a batch mode (using a hollow As can be seen in FIG . 15 , The PK profile in rats fiber filter , Model No . CFP - 6 - D - 9A from Amersham Bio (measured substantially as per patent number WO sciences of Westborough , Mass. ), for four volumes with 15 02 /096368 ) also shows an improvementwith heat treatment . normal saline on a small scale batch diafiltration system Both samples have acceptable PK profiles. The heat treated (FIG . 1B , component 70 ). The recirculation pump ( FIG . 8 , sample gives longer lasting sustained release with higher component 8110 , 8112 , and 8114 ) was set to 5 , 700 mL /min , serum values at 48 and 72 hours . The not heat treated sample while saline addition pump ( FIG . 8 , component 8170 , 8172 , was essentially zero at 72 hours . The heat treatment surpris and 8174 ) was set to 410 mL /min . The permeate valve (FIG . 20 ingly improved the accelerated stability of the MVLs and 8 , component 8202 , 8206 , and 8210 ) was adjusted to 410 also improved their in -vivo release profile . mL /min to keep the liquid volume in the system constant at 1 ,000 mL . Since the minimum working volume of this system was approximately 550 ml, which was too large to EXAMPLE 4 allow these small samples to be concentrated to the target 25 bupivacaine concentration of 15 mg/ mL , they were allowed Effect of Lowered 1st Aqueous Osmolality to settle at + 5° C . and the supernatant was decanted to The osmolality of the 1st aqueous solution in the previous achieve approximately 15 mg of bupivacaine per ml in the examples was significantly above 300 mOsm /kg . If there final MVL suspensions. was no appreciable loss of bupivacaine or phosphoric acid or The analytical results for the samples were as follows : 30 water transport across the forming phospholipid membranes during the MVL production process , the internal chambers TABLE 7 of the resultantMVLs will be filled with an aqueous solution with an osmolality at or near that of the final saline ( 300 dio doo dooppvbupi/ ml mOsm /kg ) storage suspending medium . If , on the other Not heat treated 14. 9 528 . 107. 7 56 % 16. 84 35 hand , the osmolality of the 1st aqueous and resultant MVLS Heat treated 18 . 2 51 . 3 95 . 9 55 % 18 was lower than that of saline, the MVLs will shrink slightly as the saline draws water out of the internal chambers . This will compress the phospholipids making up the MVL mem TABLE 8 branes and make them more stable and less bupivacaine permeable . Chol DEPC D PPG Tricap Sample Total mg/ml Both the lipid - solvent (LC ) and the bupivacaine -acid (1st aqueous) were made up at 1/ 2 the concentration of normal Not heat treated 4 .97 8 . 50 0 . 39 2 . 11 ( see below for formulations ) . This gave a 1st aqueous Heat treated 7 .07 12 .35 0 .58 3 .00 solution with an osmolality of 189 mOsm /kg which was expected to cause the MVLs to shrink and compress their As can be seen in the Tables 7 and 8 above , the heat membranes when diafiltered into normal saline of 300 treatment did not significantly effect the particle size distri mOsm /kg . TABLE 9 FIG . 2 # s FIG . 1B # s (but with no sterile filters ) 2160 12 LC , Solvent- lipids pump 33 ml/min 2120 2 1st aqueous, acid Bupi pump 33 ml/min 22 Dextrose - lysine pump, 3 fluid noz 66 ml/min 64 Dextrose only pump , wall rinse noz 66 ml/min 120 Output rate out of spray chamber 165 ml/min 115 & 110 Temperature of water humidified N2 44 . 6 C . feeding rotation jets Temperature of N2 exiting electric heater 46 C . 90 Flow rate of humidification water into 9 ml/min ue$ steam generator Flow to both rotation jets (N2 before 400 L /min steam ) Flow to top rotation jet 15 L /min q Chamber Jacket supply temperature 25 C . Chamber exit temperature 21 . 9 C . 2150 & 2180 heat exchanger emulsion inlet supplied to 18 . 8 C . 3 fluid nozzle US 9 ,730 , 892 B2 73 74 TABLE 9 -continued FIG . 2 # s FIG . 1B # s (but with no sterile filters ) 2175 heat exchanger emulsion outlet 17 .2 C . 2110 Heat exchanger coolant supply 4 C . 11 3 Fluid nozzle N2 pressure 60 psig Wall rinse nozzle pressure 60 psig 3 fluid nozzle N2 flow 54 L /min Wall rinse nozzle N2 flow 53 L /min Ross Mixer always with gap ring # 3 Hz on Run at 30 VFD resulting in this blade rotation rate 7 ,200 RPM 2160 10 Solvent solution (LC ) , 4 liters 4 liters cholesterol Nippon ) 20 .8 g DEPC (Nippon ) 36 . 0 g DPPG (Lipoid ) 1 .89 g tricaprylin (NOF ) 8 .81 g WFI 1 . 4 ml MeCl ( EMD ) 5 ,138 g 2120 1st aqueous soln . Acid & Bupi, 2 liters 2 liters (emulsified by Ross, fed to rotor center ) Dissolve Bupivacaine base (BASF ) 40 g in 2 liters of 0 . 112M H3PO4 (Mallinckrodt ) 112 mm Osmolality 189 mOsm 60 Dextrose / lysine solution 5 liters (fed to sheath of 3 fluid nozzle ) L - Lysine monohydrate 84 g 50 % Dextrose soln ( B Braun ) 105 g DI water to final volume of 5 L Osmolality 145 mOsm 66 Dextrose wall rinse solution 5 liters 50 % Dextrose soln ( B Braun ) 329 g DI water to final volume of 5 L Osmolality 148 mOsm

Since only 1/ 2 the lipids and 1/ 2 the previous examples EXAMPLE 5 bupivacaine are processed per minute but the Dextrose Solvent Exchange /Concentration Lysine ( second aqueous ) and Dextrose only (wall rinse ) FIGS . 8 , 10 and 11 depict example continuous buffer solutions are pumped at the normal rate , the concentration of 3535 exchange and MVL concentration systems ( seen in FIG . 1A bupivacaine MVLs out of the spray chamber ( FIG . 1 , and FIG . 1B , component 70 ) which are fed a MVL suspen component 50 ) will be 1 /2 of that seen in the previous sion from a solvent removal vessel ( seen in FIG . 1A and examples. A larger 1 , 000 mL sample was therefore taken for FIG . 1B , component 50 ) by a solvent line (seen in FIG . 1A concentration and diafiltrationdiafiltration as inin thethe previous example . and FIG . 1B , component 120 ) . This sample was not heat treated . These systems take the continuous flow of MVL suspen sion produced in a solvent removal vessel ( seen in FIG . 1A TABLE 10 and FIG . 1B , component 50 ) and exchange the suspending buffer for another suspending medium ( e . g . normal saline ) Analysis pf the final decanted product MVLs are as follows: and at the same time optionally concentrate the MVL 45 suspension . In most cases it is desired to produce slightly Bupi over concentrated MVL suspensions in a retentate vessel mg/ ml % free PPV % d10d50 D90 (seen in FIG . 8 , component 8300 ; FIG . 10 , component 15 .21 1 . 1 % 71 % 9 . 3 22. 5 55 . 5 10500 ; and FIG . 11 , component 11400 ). The lot is analyzed and sterilely diluted to exact concentration before filling into vials aseptically. The final concentration of the MVL is The accelerated stability ( 30 C ) plot for this low osmo - 50 obtained by processing in two or more stages connected in lality sample can be seen in FIG . 14 . The low osmolality series . Additionally , processing results in any percent of sample was significantly more stable than the not heat original suspending buffer removal . treated sample but less stable than the heat treated sample . These systems include medium supply source , such as a As seen in FIG . 15 , the Rat in - vivo release profile of the tank and a particle concentrating device . The particle con low osmolality sample can be the most desirable as it has the 55 centrating device is a hollow fiber tangential flow filter ( e . g . lowest initial peak and the longest duration ; highest blood Model No . CFP - 6 - D - 9A from Amersham Biosciences of concentrations at 72 and 96 hours . In this process , starting Westborough , Mass. ) or a continuous or semi- continuous with a 1st aqueous solution with an osmolality lower than the centrifuge (Centritech Lab Ill or CARR ViaFuge Pilot from Pneumatic Scale Angelus Corp . , Clearwater, Fla . ) or any final MVL suspending solution was unexpectedly found to other device that separates the MVLs from the suspending both increase MVL storage stability , reduce the initial in - 60 medium . vivo release peak and prolong the duration of bupivacaine The following constraints apply to analogous components delivery. of all the steady state stages of the continuous buffer As can be seen from 1 Table 10 , this example produced a exchange and MVL concentration systems: smaller particle size d50 than the previous examples . As can In a continuousmanner , the first stage of FIG . 8 consisting be seen from FIG . 14 , the low osmolality sample , which was 65 of components 8120 , 8100 , 8122 , 8123 , 8110 , 8190 , 8202 . not heat treated , has a much better accelerated stability than 8160 , 8150 , 8140 , 8180 , 8131 , 8130 , 8212 and 8170 will the not heat treated sample of the previous example . exchange the initial suspending buffer with normal saline . US 9 ,730 ,892 B2 75 76 In reference to FIG . 8 , at steady state where a constant The third stage in FIG . 8 contains the final retentate volume is in the tank , the input volume flows ; MVL sus vessel, 8300 , and is large enough to hold one lot of product, pension in 8120 and saline in 8130 must equal the volume out flows; permeate 8160 and MVL suspension transferred e . g . 40 liters . As shown , the volume in final retentate vessel to the next stage by metering pump 8123 . This is done by 8300 remains constant but the concentration is continuously controlling / fine adjusting any of the above 4 flow rates to rising as there is no MVL out flow , until it reaches the keep the volume in the tank constant, e . g . saline fine desired final concentration of MVLs. By choosing appro adjustment valve 8212 . priate flow rates in pipes 8124 , 8208 and 8164 this stage is At steady state the mass or number ofMVLs is conserved and thus the input rate of the number of MVLs, concentra alternatively run with a constant MVL concentration and tion times the flow rate , from pipe 8120 must equal theMVL 10 rising volume. outflow from 8122 , again number concentration times flow When starting these systems, the systems are either rate . This means that if the volume outflow rate in 8122 is started with each tank filled to the appropriate volume with lower than the input volume flow rate in 8120 , the concen tration of MVLs in the tank 8100 and pipe 8122 rises until saline or they are started empty and filled with the MVL the MVL in matches the MVL out. This gives a MVL suspension input and saline input but only start the output concentration factor, output MVL concentration divided by 15 pump, e . g . 8123 , and hollow fiber recirculation pump , e . g . the input MVL concentration equal to the volume flow in MVL input pipe, 8120 , divided by the volume flow in the 8110 when the tank for that stage fills to its desired volume. output pipe , 8122 . Likewise at the end of a lot , the MVL input line is switched The original buffer is diluted by saline at each stage . This to saline which moves all MVLs to the product tank , or the dilution factor is the flow rate of buffer in to the tank , 8100 , 20saline and MVL inputs and the hollow fiber recirculation divided by the total of the volume flow rate ofbuffer in pipe 8120 and the volume flow rate of saline in pipe 8212 . The pump is stopped to allow each tank to empty into the next MVLs take up appreciable volume and so the flow rate of until the lot is again all in the product vessel . buffer into the tank , 8100 , is the volume flow rate in pipe Flow rate and additional parameters for the system of 8120 times one minus the volume percentage of that sus FIG . 8 using the hollow fiber cartridge and filtration rates pension that is MVLS, (PPV % , Packed Particle Volume and MVL output rate of the 25 Hz Process, 30 Hz Process , percentage) . or 35 Hz Process of Example 1 is given below : TABLE 11 ( flow rates are for fluid traveling through the components from FIG . 8 ) concentration factor of 100 % in 1st 2 stages (buffer exchange only ) MVL MVL suspension suspension in Saline in out permeate Stage (mL / min ) (mL /min ) (mL / min ) (mL / min ) 8120 165 8130 410 8122 165 8160 410 8122 165 8132 410 8124 165 8162 410 WNE 8124 165 8134 245 8164 410 Lysine Net tank volume part conc . dilution % Lysine % Stage change out PPV % factor left exchanged System 810 Input, 100 % 20 . 0 % 100 % 8120 > > > 8100 0 100 % 20 . 0 % 0 . 2435 24 . 4 % 75 . 6 % 8200 0 100 % 20 . 0 % 0 .2435 5 . 9 % 94 . 1 % WNA 8300 rising rising 0 . 3501 2 . 1 % 97 . 9 %

Adhering to these constraints , the first 2 stages of FIG . 8 , The flow rates in Table 11 result in buffer exchange only . containing tanks 8100 and 8200 will reach steady state . The . The MVL PPV remains at 20 % but the original buffer concentration is reduced by 99 . 7 % with its concentration in smaller the tanks, the faster the equilibration time and the the product being 2 . 1 % of original. For comparison , a 4 lower the total holdup in the system . For the systems volume batch diafiltration exchange as used in the examples , described in Examples 2 and 3 , a tank volume of from 0 . 25 reduces the buffer concentration to 1. 8 % or 98 .2 % liter to 10 liters and preferably 0 .75 liters is appropriate . exchanged . TABLE 12 ( flow rates are for fluid traveling through the components from FIG . 8 ) A total concentration factor of 200 % in 1st 2 stages and buffer exchange MVL MVL suspension suspension in Saline in out permeate Stage (mL / min ) (mL / min ) (mL / min ) (mL /min ) ?? 8120 165 8130 369 8122 124 8160 410

? 8122 124 8132 396 8124 110 8162 410 ???? 8124 110 8134 300 0 81648164 410 US 9 , 730 ,892 B2 77 TABLE 12 - continued ( flow rates are for fluid traveling through the components from FIG . 8 ) A total concentration factor of 200 % in 1st 2 stages and buffer exchange Stage Lysine Net tank volume concentration dilution % Lysine % Stage change factor PPV % factor left exchanged System 810 Input , 100 % 20 . 0 % 100 % 8120 > > > 8100 0 0 133133 % 26 . 6 % 0 .2635 26 . 3 % 73 . 7 % ?? 8200 0 150 % 40 . 0 % 0 . 1870 4 . 9 % 95 . 1 % 830008300 O rising rising 0 .1798 0 . 9 % 99 . 1 %

The flow rates in Table 12 result in buffer 99. 1 % 15 .including the tanks (seen in FIG . 10, components 10100 , exchanged down to 0 . 9 % of the original concentration . 10200 and 10300 ) run at both constant volume and MVL TABLE 13 ( flow rates are for fluid traveling through the components from FIG . 11 ) concentration factor of 300 % in 3 stages and buffer exchange Product is collected in Tank 11400 with volume rising at 114 . 3 mL/ min MVL suspension MVL Supernatant Saline suspension out out Stage (mL /min ) (mL /min ) (mL / min ) (ml /min ) 11120 165 11130 949 11122 114 . 3 11160 1 , 000 ????? 11122 114 11132 1000 11124 114 . 3 11162 1 ,000 11124 114 11134 1000 11166 114 . 3 11164 1 , 000 Stage Lysine Net tank volume concentration dilution % Lysine % Stage change factor PPV % factor left exchanged System 1110 100 % 20 . 0 % 100 % Input, 11120 > > > 11100 O 144 % 28 . 9 % 0 . 1221 12 . 2 % 87 . 8 % ????? 11200 O 144 % 41 . 7 % 0 . 0752 0 . 9 % 99 . 1 % 11301130 0 144 % 60 . 1 % 0 .0625 0 . 1 % 99 . 9 %

The flow rates in Table 13 result in a concentration factor 40 concentration while the final tank (seen in FIG . 10 , compo of 300 % with the same input conditions used for Tables 11 nent 10500 ) collects the concentrated buffer exchanged and 12 . MVL suspension Systems can be assembled with any combination of FIG . 11 depicts a system for continuous buffer exchange hollow fiber cartridges and centrifuges. They can have two and MVL concentration including only centrifuges. Using or three or more stages . With more stages the buffer is 45 typical process rates for the ViaFuge Pilot and the same feed exchanged with less saline but there is more equipment to conditions as the hollow fiber system , provides a system that keep aseptic . exchanges 99 . 9 % of the buffer while concentrating the MVL suspension by a factor of 3 when using the parameters from The semi- continuous centrifuges above, Centritech Lab Table 13 . The saline inputs should only flow when the input III or CARR ViaFuge Pilot, are operated in an aseptic 50. MVL stream is flowing . fashion . The semi- continuous centrifuges are capable of Any tanks connected to these centrifuges must be large very high particle concentration factors e. g. 100 to 1 and can enough to accommodate the intermittent nature of the cen trifuge input and output. also discharge MVL concentrate up to 80 % PPV % . The While the present invention has been described with Centritech Lab III is semi- continuous as it has a constant 55 reference to the specific embodiments thereof, it should be feed with an intermittent concentrate discharge , every 10 understood by those skilled in the art that various changes seconds to 2 minutes . The ViaFuge Pilot has a constant feed may be made and equivalents may be substituted without departing from the true spirit and scope of the invention . In which is interrupted every 2 to 10 minutes by a rapid addition , many modifications may be made to adapt a discharge cycle . 60 particular situation , material, composition of matter, pro FIG . 10 depicts continuous buffer exchange and MVL cess , process step or steps, to the objective , spirit and scope concentration systems where the hollow fiber filters operate of the present invention . All such modifications are intended at lower particle concentrations , a condition where they have to be within the scope of the claims appended hereto . higher permeate rates . The centrifuge ( seen in FIG . 10 , What is claimed is : component 10400 ), e . g . CARR ViaFuge Pilot, is used to do 65 1 . A composition comprising a suspension of multivesicu a final concentration into the final product vessel (seen in lar liposomes in a suspending medium , said multivesicular FIG . 10 , component 10500 ) . In this system all three stages liposomes having a structure including multiple non - con US 9 ,730 ,892 B2 79 80 centric chambers and comprising at least one amphipathic a first solution inlet line supplying a second suspend lipid , at least one neutral lipid and a therapeutic agent: ing medium ; wherein said multivesicular liposomes are made by a a first sterilizing hydrophilic filter, connecting the process comprising removing greater than 90 % of an first retentate vessel and the first solution inlet organic solvent from multivesicular liposome pre -drop - 5 line ; lets using an evaporation apparatus comprising : a first pump located along the first particle suspen a solvent removal vessel having a top , a bottom and a sion outlet line between the first retentate vessel circular wall ; and the first hollow fiber tangential flow filter ; a rinse nozzle mounted to and extending through the a first retentate line between the first hollow fiber top of the solvent removal vessel; 1010 tangential flow filter and the first retentate vessel ; a carrier gas entrance orifice ; a second particle suspension outlet line , leading to a a solvent removal gas exit orifice centrally connected to second concentrator unit , the top and further comprising a gas outlet tube said second concentrator unit comprising : extending into the solvent removal vessel , wherein a second retentate vessel; the gas outlet tube is fitted with a narrowing cone and 15 a second particle suspension inlet line , connected to the narrowing cone includes a tip through which gas the second retentate vessel and the second particle residing in the solvent removal vessel enters the suspension outlet line ; narrowing cone ; a third particle suspension outlet line, connecting the a product exit orifice connected to the bottom of the second retentate vessel and a second hollow fiber vessel; and 20 tangential flow filter; a product outlet pipe connected to the product exit a second solution inlet line supplying a third sus orifice ; pending medium ; at least one atomizing nozzle comprising : a second sterilizing hydrophilic filter, connecting the a first fluid conduit and a second fluid conduit each second retentate vessel and the second solution having at least one entrance orifice and at least one 25 inlet line ; and exit orifice ; a fluid contacting chamber having a a second pump located along the third particle sus top comprising at least one entrance orifice and pension outlet line between the second retentate having a bottom comprising at least one exit vessel and the second hollow fiber tangential flow orifice and said at least one entrance orifice con filter ; necting to the at least one exit orifice of the first 30 a second retentate line between the second hollow fluid conduit and the at least one exit orifice of the fiber tangential flow filter and the second retentate second fluid conduit ; a third fluid conduit , wherein vessel; and the third fluid conduit annularly surrounds a por optionally a fourth particle suspension outlet line . tion of the fluid contacting chamber; 2 . The composition of claim 1 , wherein the therapeutic wherein the organic solvent removal process comprises : 35 agent is bupivacaine . introducing multivesicular liposome pre -droplets to the 3 . The composition of claim 1 , wherein the continuous solvent removal vessel by the at least one atomizing processing system further comprises the fourth particle nozzle , wherein the multivesicular liposome pre - suspension outlet line and a third concentrator unit , said droplets comprise a first component core and an third concentrator unit comprising : aqueous phase shell, wherein the first component 40 a third retentate vessel; core comprises a suspension of first aqueous phase a third particle suspension inlet line , connected to the third droplets suspended in a first organic phase ; retentate vessel and the fourth particle suspension out applying a carrier gas in a tangental direction to the let line ; circular wall through the carrier gas entrance orifice ; a fifth particle suspension outlet line, connecting the third removing a solvent removal gas through the solvent 45 retentate vessel and a third hollow fiber tangential flow removal gas exit orifice to provide multivesicular filter; liposomes in a first suspending medium and remov a third solution inlet line supplying a fourth suspending ing the formed multivesicular liposomes suspended medium ; in the first suspending medium through the product a third sterilizing hydrophilic filter, connecting the third exit orifice through a product outlet pipe; and 50 retentate vessel and the third solution inlet line ; exchanging the first suspending medium by introducing a third pump located along the fifth particle suspension the multivesicular liposomes in the first suspending outlet line between the third retentate vessel and the medium into a continuous processing system to form third hollow fiber tangential flow filter ; a multivesicular liposomes suspension in the sus a third retentate line between the third hollow fiber pending medium , wherein the continuous processing 55 tangential flow filter and the third retentate vessel; and system , comprises : optionally a sixth particle suspension outlet line . a first concentrator unit comprising : 4 . The composition of claim 3 , wherein the continuous a first retentate vessel; processing system further comprises the sixth particle sus a first particle suspension inlet line , connected to the pension outlet line and a fourth concentrator unit, said fourth first retentate vessel and the product outlet pipe , 60 concentrator unit comprising : wherein the process comprises applying the sus a fourth retentate vessel ; pension of multivesicular liposomes in the first a fourth particle suspension inlet line , connected to the suspending medium from the product outlet pipe fourth retentate vessel and the sixth particle suspension to the first particle suspension inlet line ; outlet line ; a first particle suspension outlet line, connecting the 65 a seventh particle suspension outlet line , connecting the first retentate vessel and a first hollow fiber tan fourth retentate vessel and a fourth hollow fiber tan gential flow filter ; gential flow filter; US 9 , 730 ,892 B2 82 a fourth solution inlet line supplying a fifth suspending pension outlet line and a final product vessel connected to medium ; the third retentate vessel through the sixth particle suspen a fourth sterilizing hydrophilic filter , connecting the sion outlet line . 9 . The composition of claim 3 , wherein the third retentate fourth retentate vessel and the fourth solution inlet line ; 5 vessel is a final product vessel . a fourth pump located along the seventh particle suspen - 5 10 . The composition of claim 3 , wherein the therapeutic sion outlet line between the fourth retentate vessel and agent is bupivacaine . the fourth hollow fiber tangential flow filter; 11 . The composition of claim 1 , wherein the continuous a fourth retentate line between the fourth hollow fiber processing system further comprises the fourth particle tangential flow filter and the fourth retentate vessel ; and suspension outlet line and a final product vessel connected optionally an eighth particle suspension outlet line . to the second retentate vessel through the fourth particle 5 . The composition of claim 4 , wherein the continuous suspension outlet line . processing system further comprises the eighth particle 12 . The composition of claim 1, wherein the second suspension outlet line and a final product vessel connected retentate vessel is a final product vessel. to the fourth retentate vessel through the eighth particle 15 13. The composition of claim 1, wherein the removing of suspension outlet line . greater than 90 % of the organic solvent from the multive 6 . The composition of claim 4 , wherein the fourth reten - sicular liposome pre -droplets occurs in a few seconds. tate vessel is a final product vessel. 14 . The composition of claim 1 , wherein the removing of 7 . The composition of claim 4 , wherein the therapeutic greater than 90 % of the organic solvent from the multive agent is bupivacaine . 20 sicular liposome pre -droplets occurs in a fraction of a 8. The composition of claim 3, wherein the continuous second . processing system further comprises the sixth particle sus * * * *