Methods for making ultrasound contrast agents
Provided herein are improved methods for preparing phospholipid formulations including phospholipid UCA formulations.
1. A method of ultrasound contrast imaging a subject comprising
(a) combining one or more phospholipids and a non-aqueous solvent, in a methanol and toluene free and methyl t-butyl ether free condition, to form a phospholipid solution having calcium and/or magnesium divalent metal cation present at a total divalent metal cation concentration of less than 0.7 ppm,
(b) combining the phospholipid solution with an aqueous solution to form a phospholipid suspension,
(c) activating the phospholipid suspension with a perfluorocarbon gas to form lipid-encapsulated gas microspheres,
(d) administering the lipid-encapsulated gas microspheres to a subject, and
(e) obtaining an ultrasound image of the subject.
2. The method of claim 1 , wherein the one or more phospholipids comprise (a) DPPC and MPEG-5000-DPPE or (b) DPPA, DPPC and MPEG-5000-DPPE.
3. The method of claim 1 , wherein the one or more phospholipids comprise DPPA, DPPC and MPEG-5000-DPPE.
4. The method of claim 2 , wherein DPPC has a combined calcium and magnesium concentration of less than 90 ppm, and MPEG5000-DPPE has a combined calcium and magnesium concentration of less than 115 ppm.
5. The method of claim 1 , wherein the non-aqueous solvent comprises propylene glycol.
6. The method of claim 1 , wherein the non-aqueous solvent has a combined calcium and magnesium concentration of less than 0.7 ppm.
7. The method of claim 1 , wherein the non-aqueous solvent comprises propylene glycol and glycerol.
8. The method of claim 1 , wherein the phospholipid solution has no detectable phospholipid precipitate.
9. The method of claim 1 , wherein the one or more phospholipids comprise DPPA, DPPC and MPEG-5000-DPPE in molar percentages of 5-15%, 77-90% and 5-15%, respectively.
10. The method of claim 1 , wherein the one or more phospholipids comprise DPPA, DPPC and MPEG-5000-DPPE in a mole % ratio of 10 to 82 to 8 (10:82:8).
11. The method of claim 1 , wherein the perfluorocarbon gas is perfluoropropane gas.
12. The method of claim 1 , wherein the one or more phospholipids comprise DPPC and MPEG-5000-DPPE, the phospholipid solution comprises DPPC, MPEG5000-DPPE and propylene glycol, the aqueous solution comprises water, propylene glycol and glycerol, and the phospholipid suspension comprises DPPC, MPEG5000-DPPE, propylene glycol, water and glycerol.
13. The method of claim 1 , wherein the aqueous solution further comprises one or more salts.
14. The method of claim 1 , wherein the aqueous solution further comprises one or more buffers.
15. A method of ultrasound contrast imaging a subject comprising
(a) selecting an MPEG5000-DPPE having calcium and/or magnesium divalent metal cation present at a total divalent metal cation concentration of less than 115 parts per million (ppm),
(b) combining the MPEG-5000-DPPE with one or more phospholipids and a non-aqueous solvent, in a methanol and toluene free and methyl t-butyl ether free condition, to form a phospholipid solution having calcium and/or magnesium divalent metal cation present at a total divalent metal cation concentration of less than 0.7 ppm,
(c) combining the phospholipid solution with an aqueous solution to form a phospholipid suspension,
(d) activating the phospholipid suspension with a perfluorocarbon gas to form lipid-encapsulated gas microspheres,
(e) administering the lipid-encapsulated gas microspheres to a subject, and
(f) obtaining an ultrasound image of the subject.
16. The method of claim 15 , wherein the MPEG-5000-DPPE with the one or more phospholipids comprise (a) DPPC and MPEG-5000-DPPE or (b) DPPA, DPPC and MPEG-5000-DPPE.
17. The method of claim 15 , wherein the MPEG-5000-DPPE with the one or more phospholipids comprise DPPA, DPPC and MPEG-5000-DPPE.
18. The method of claim 16 , wherein DPPC has a combined calcium and magnesium concentration of less than 90 ppm, and MPEG5000-DPPE has a combined calcium and magnesium concentration of less than 115 ppm.
19. The method of claim 15 , wherein the non-aqueous solvent comprises propylene glycol.
20. The method of claim 15 , wherein the non-aqueous solvent has a combined calcium and magnesium concentration of less than 0.7 ppm.
21. The method of claim 15 , wherein the non-aqueous solvent comprises propylene glycol and glycerol.
22. The method of claim 15 , wherein the phospholipid solution has no detectable phospholipid precipitate.
23. The method of claim 15 , wherein the MPEG-5000-DPPE with the one or more phospholipids comprise DPPA, DPPC and MPEG-5000-DPPE in molar percentages of 5-15%, 77-90% and 5-15%, respectively.
24. The method of claim 15 , wherein the MPEG-5000-DPPE with the one or more phospholipids comprise DPPA, DPPC and MPEG-5000-DPPE in a mole % ratio of 10 to 82 to 8 (10:82:8).
25. The method of claim 15 , wherein the perfluorocarbon gas is perfluoropropane gas.
26. The method of claim 15 , wherein the MPEG-5000-DPPE with the one or more phospholipids comprise DPPC and MPEG-5000-DPPE, the phospholipid solution comprises DPPC, MPEG5000-DPPE and propylene glycol, the aqueous solution comprises water, propylene glycol and glycerol, and the phospholipid suspension comprises DPPC, MPEG5000-DPPE, propylene glycol, water and glycerol.
27. The method of claim 15 , wherein the aqueous solution further comprises one or more salts.
28. The method of claim 15 , wherein the aqueous solution further comprises one or more buffers.