Name: manufacture of concentrated, lipid-based oxygen microbubble emulsions by high shear homogenization and serial concentration
Uploaded: 2014-05-26T14:00:00
Description: Watch this Scientific Journal Video about Manufacture of Concentrated, Lipid-based Oxygen Microbubble Emulsions by High Shear Homogenization and Serial Concentration at JoVE.com

Finansiering: US Army Medical Research &amp; Materiel Kommando (USAMRMC) og forvaltes af Telemedicin &amp; Advanced Technology Research Center. Shunxi Ji bidrog ændringen af ​​sprøjter, som beskrevet her.

1.. System Set-up Systemet består af en bedrift og koncentrere tanken (HCT), forsynet med en enkelt fase mixer, en in-line homogenisator med høj forskydning, en rulle-pumpe til at flytte væske mellem HCT og homogenisatoren, og en varmeveksler (figur 1). <ol><li> Placer en steriliseret, bred åbning 4 L samling glasbeholder udstyret med 2 basishavne og 3 sideporte beneath enkelt fase mixer. Sænk mikserhovedet til mundingen af ​​beholderen og sikre en gastæt monteri...

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De vigtigste skridt til at skabe koncentrerede, stærkt iltet LOMs omfatter: 1) at sikre, at headspace i HCT forbliver fuldt iltet; 2) at sikre, at renheden af ​​lipid hjælpestofferne er optimal (herunder opbevaring og brug af GMP produkter); 3) at sikre, at de pulveriserede lipider blandes fuldstændigt med den vandige fase forud for priming af systemet; og 4) meget opmærksom på stigningen i fraktionen gas i HCT at sikre, at volumen fraktion af gas ikke overstiger 70%. Fremgangsmåde...

Mikrobobler bestående af protein, polymer og lipid skaller er blevet udviklet som vektorer for drug delivery, genterapi, og ultralyd-kontrastmidler 1-5. Fordi disse terapeutiske anvendelser kræver intravaskulære mikrobobler vedholdenhed er sådanne mikrobobler almindeligvis fyldt med inerte, høje molekylvægt gasser såsom perfluorcarboner 6, der har lav opløselighed i blod og stabilisere boblen 3,4. For nylig har lipid-baserede ilt mikrobobler (LOMs) er designet til at levere terapeutiske doser af ilt, der kan bevare slutorganet ilttilførsel og forhindre hæmodynamisk ustabilitet i perioder med luftvejsobstruktion eller hypoxæmi 7. Emulsioner designet til intravenøs gas levering kræver forskellige design funktioner end dem, der anvendes til ultralyd kontraststoffer eller målrettede drug delivery. Først, fordi kroppen forbruger store mængder ilt gas (~ 200 ml / min), LOMs skal produceres oginjiceret på en stor skala. Dette kræver, at fremstillingsprocessen være effektiv. For det andet bør fremstillingsprocessen være lukket kredsløb for at undgå kontaminering nitrogen gennem eksponering LOMs (som skal fyldes med 100% oxygen) til den omgivende luft. For det tredje, fordi formålet med LOMs er intravenøs gas levering, gas fraktion af LOMs skal maksimeres, anerkender de begrænsninger, som emulsion viskositet 7. Endelig, som med enhver intravenøs injektion, præcis kontrol over partikelstørrelsesfordeling er afgørende for at undgå mikrovaskulære obstruktion 8. Der er flere etablerede metoder til mikrobobler fremstillingen. Lydbehandling udnytter høj intensitet, lavfrekvente ultralyd påføres luft-væske-grænsefladen af en emulsion, der omfatter et overfladeaktivt middel, såsom et amfipatisk phospholipid, i nærvær af en gas headspace at fremstille mikrobobler 7,9. Denne proces kan styres ved at variere ultralyd frekvens, effekt og puls varighed, og den resulterende størrelse fordelingen kan skræddersyes til at producere mikrobobler af en bestemt størrelse distribution, selvom lydbehandling er sjældent anvendes til fremstilling af klinisk anvendte mikrobobler. Sammenlægning er intens mekanisk omrøring af et overfladeaktivt middel og gas i et lukket system, som også er vanskelig at skalere op til at rumme store mængder 2. Droplet-baserede MicroFluidics giver præcis styring af mikroboblestørrelse fordeling 10-13. Selvom traditionelt svært at skalere op, multi-kanal, high-speed microfluidics er blevet beskrevet som øger mikrobobler produktionseffektivitet 13. Mikrobobler fremstillet ved hjælp af en af disse metoder kan kræve post-fremstilling størrelse reduktion processer, såsom centrifugal fraktionering 14,15 og mikrobobler flotation 16,17. En anden fast metode til fremstilling af meget stabile mikrobobler er shear homogenizning 6, som kan resultere i en stabiliserende sekskantet phospholipid mønster på mikroboble overflade 18. Med udgangspunkt i dette koncept beskriver vi inkorporeringen af en in-line høj forskydning homogeniseringsapparat at skabe selvsamlende LOMs 19. I denne proces udnytter homogenisatoren hurtigt roterende knive tæt på dobbelt finmasket emulsor skærme, skabe høj mekanisk og hydraulisk forskydning til oprettelse af mikrobobler. Serial koncentration af fedtemulsionen via dette system giver en mere koncentreret gasfraktion, som kan yderligere koncentreres ved centrifugering.

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	<tbody>
		<tr height="42">
			<td height="42" style="height:43px;width:239px;">Name of Material/ Equipment</td>
			<td style="width:141px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:405px;">Comments/Description</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)</td>
			<td style="width:141px;">Avanti Polar Lipids</td>
			<td style="width:119px;">770365</td>
			<td>Alternate product: non-GMP from NOF America (Coatsome MC-8080)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">Cholesterol</td>
			<td style="width:141px;">Sigma Aldrich</td>
			<td style="width:119px;">C75209</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">Plasma-Lyte A</td>
			<td style="width:141px;">VWR</td>
			<td style="width:119px;">80089-818</td>
			<td>Alternatively can use NaCl</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">Glass collection vessel</td>
			<td style="width:141px;">Specialty Glass, Inc.</td>
			<td style="width:119px;">Custom</td>
			<td>Contact: Pam Zurbrick - 281-595-2210</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">Gas composition (oxygen) monitor</td>
			<td style="width:141px;">Precision Medical</td>
			<td style="width:119px;">PM5900L</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">Sarns 8000 roller pump</td>
			<td style="width:141px;">Calicut Medical</td>
			<td style="width:119px;">16407</td>
			<td>Part of a modular perfusion system</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">BIOtherm Heat Exchanger</td>
			<td style="width:141px;">Medtronic</td>
			<td style="width:119px;">ECMOtherm-II</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">Verso laboratory in-line mixer</td>
			<td style="width:141px;">Silverson Machines, Inc</td>
			<td style="width:119px;">TH-IL-102-VERSO</td>
			<td>Use multistage workheads and front-end extension with T piece</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">T-piece for Silverson Verso inlet port</td>
			<td style="width:141px;">Process Innovations</td>
			<td style="width:119px;">Custom</td>
			<td>Contact: Brian Leavitt - 508-423-2266</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">L5M-A laboratory mixer</td>
			<td style="width:141px;">Silverson Machines, Inc</td>
			<td style="width:119px;">NC0136483</td>
			<td>Use mesh emulsor screen (fine)</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">Rochester-Ochsner toothed forceps</td>
			<td style="width:141px;">Fisher Scientific</td>
			<td style="width:119px;">13-812-18</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">140 mL syringe</td>
			<td style="width:141px;">Kendall Healthcare Monoject&#160;</td>
			<td style="width:119px;">8881114030</td>
			<td>Ensure there is a luer lock.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:239px;">IX71 Inverted light microscope</td>
			<td style="width:141px;">Olympus</td>
			<td style="width:119px;">IX71</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:239px;">Retiga-2000R microscope camera</td>
			<td style="width:141px;">QImaging</td>
			<td style="width:119px;">RET-2000R-F-M-12</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:239px;">Accusizer 780A Autodilution</td>
			<td style="width:141px;">PSS-NICOMP Particle Sizing Systems</td>
			<td style="width:119px;">Out of production</td>
			<td></td>
		</tr>
	</tbody>
</table>

manufacture of concentrated, lipid-based oxygen microbubble emulsions by high shear homogenization and serial concentration

Gasfyldte mikrobobler er udviklet som ultralyd kontrast og drug delivery agenter. Mikrobobler kan fremstilles ved behandling af overfladeaktive midler ved hjælp af sonikering, mekanisk omrøring, mikrofluide anordninger eller homogenisering. For nylig har lipid-baserede ilt mikrobobler (LOMs) er designet til at levere ilt intravenøst ​​i løbet af ulykkestilfælde, vende livstruende hypoxæmi og forhindre efterfølgende orgel skade, hjertestop og død. Vi præsenterer metoder til opskaleret produktion af stærkt iltet mikrobobler ved hjælp af et lukket kredsløb high-shear homogenisator. Processen kan producere 2 L af koncentrerede LOMs (90 volumen%) i 90 minutter. Resulterende bobler har en gennemsnitlig diameter på ~ 2 um og en rheologisk profil i overensstemmelse med den af ​​blod, når den fortyndes til 60 volumen%. Denne teknik producerer LOMs i høj kapacitet og med høj ilt renhed, hvilket tyder på, at denne teknik kan være nyttige for translationelle forskningslaboratorier.

High shear homogenisering muliggør en effektiv (dvs. inden for en eftermiddag) produktion af tilstrækkelige LOMs for en dyr undersøgelse og ikke kræver teknisk ekspertise. Når dygtige, kan op til 2 L koncentrerede LOMs blive fremstillet i 90 min. Mikrobobler størrelse og morfologi blev vurderet ved lysmikroskopi og lys dæmpning. Når en 10 pi prøve af LOMs blev visualiseret blev sfæriske LOMs bemærkes, såvel som en relativ mangel på lipid debris (figur 3A).</stro...

Watch this Scientific Journal Video about Manufacture of Concentrated, Lipid-based Oxygen Microbubble Emulsions by High Shear Homogenization and Serial Concentration at JoVE.com

Manufacture of Concentrated, Lipid-based Oxygen Microbubble Emulsions by High Shear Homogenization and Serial Concentration

Vi beskriver metoder til fremstilling af store mængder lipid-baserede ilt mikrobobler (LOMs) beregnet til intravenøs ilttilførsel ved hjælp af high-shear homogenisering og serienummer koncentration.

Fremstilling af koncentreret, Lipid-baserede Oxygen Microbubble Emulsioner af High Shear Homogenisering og Serial Koncentration

Gas-filled microbubbles have been developed as ultrasound contrast and drug delivery agents. Microbubbles can be produced by processing surfactants using ...

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