Determination of hydroperoxides in liposomes by the modified IDF and the modified tiron methods.

Two new methods, the modified International Dairy Federation and the modified Tiron methods, were developed for the determination of hydroperoxides in liposomes. Interferences of alpha-tocoferol and egg-phosphatidylcholine (EPC) required a solid-phase extraction before the analysis to eliminate alpha-tocoferol and a fluid-fluid extraction based on the solvent triangle of Bligh and Dyer to separate EPC. The developed color, using thiocyanate and Tiron, respectively, as complex-formers for the generated ferri-ions, was measured spectrophotometrically. Both techniques showed good reproducibility and high sensitivity. Peroxide contents of 0.04 and 0.07% (g/g) in EPC samples were easily determined.

Neurotoxicological assessment after intracisternal injection of liposomal bupivacaine in rabbits.

UNLABELLED: Experiments were performed on rabbits randomly assigned to intracisternally receive 0.3 mL of plain bupivacaine 5 mg/mL, liposomal bupivacaine 5 mg/mL, bupivacaine-free liposomes, or isotonic phosphate-buffered saline. Mechanical ventilation was initiated or intravenous dopamine was infused when respiratory depression or hypotension occurred. Seven days after the injection, the whole spinal cord was removed and histopathologic characteristics were studied on transverse sections. All preparations were devoid of phosphatidylcholine hydrolysis or oxidation compounds. Solutions without bupivacaine produced transient irritative signs that required sedation in most rabbits. Despite the similar duration of respiratory depression in groups receiving liposomal or plain bupivacaine, liposomes produced significantly prolonged motor blockade (126 vs 70 min). Correction of hypotension after plain bupivacaine required a longer dopamine infusion and larger doses than after liposomal bupivacaine (28 vs 18 min and 74 vs 47 mg). Necrosis was observed in the cervical area of two rabbits (one in the liposomal bupivacaine group and another in the phosphate buffer group). No demyelinated areas were noted in spinal cord examinations. We conclude that liposomal bupivacaine leads to a less severe sympathetic block and to a prolonged motor block, whereas histologic changes are not significantly different among groups. IMPLICATIONS: Multilamellar liposomes containing bupivacaine administered intracisternally to rabbits produce spinal cord histopathologic changes not significantly different from those observed with plain bupivacaine. Sustained release of bupivacaine from liposomes is suggested by the prolonged motor blockade and the reduced severity of arterial hypotension. Use of these liposomes could prolong the local anesthetic effects of bupivacaine.

Scintigraphic visualization of intrathecal liposome biodistribution.

BACKGROUND: Liposomes containing local anaesthetics have been administered intrathecally and in the epidural space. Poor attention has been given to the pharmacokinetics of liposomes as drug carriers. Therefore, we observed the biodistribution of liposomes after intrathecal injection in rats by scintigraphic imaging during 24 h. METHODS: We administered 99mTc-labeled multilamellar (MLV) and small unilamellar vesicles (SUV) of defined size and volume dispersities into the cerebrospinal fluid at the lumbar level. Those vesicles were free of contamination by radiolabeled colloids as visualized by light and electron microscopy and of neurotoxic products from phosphatidylcholine hydrolysis and peroxidation, both during the preparation process and after 24 h incubation in cerebrospinal fluid at 37 degrees C in vitro. RESULTS: SUV immediately diffused from the lumbar site of injection to the head and were cleared between 1 and 24 h after injection. MLV were cleared more slowly from the spinal space and appeared in the head region 1 h after injection where they accumulated up to 24 h. These differences were explained in terms of vesicle sizes and volumes. SUV with 0.05 micron diameters were rapidly absorbed into the blood through the arachnoid granulations. In contrast, particles larger than the upper size limit of the arachnoid granulations permeability (+/- 8 microns) could accumulate in the head with a slow elimination rate. CONCLUSION: This difference in clearance from the intrathecal space outlines the importance of defining the size of the liposomes, the distribution of a tracer or a drug inside the liposomal preparation, the chemical stability and the absence of toxic degradation products of liposome formulations before clinical use.

99mTechnetium-stannous oxinate as marker of liposome formulations.

Liposomes associated with tin(II) dioxinate were prepared from egg yolk phosphatidylcholine and cholesterol as sterile and pyrogen-free multilamellar or unilamellar vesicles. Complexing of liposomal tin(II) dioxinate with 99mTc attained 98% of the added radioactivity. Thirty percent 99mTc were released during 24-h incubation in biological fluids. The absence of tin colloids seen by electron microscopy and the stability of liposomal phospholipid and tin(II) dioxinate during 72-h incubation at 37 degrees C in plasma and cerebrospinal fluid would allow safe and reliable scintigraphic liposome pharmacokinetic studies.

Biodistribution of liposomes after extradural administration in rodents.

We have mapped over 24 h the biodistribution of 99mTc-labelled multilamellar and small unilamellar liposomes in rabbits and rats by scintigraphic imaging after extradural injection. Multilamellar vesicles formed a depot at the site of injection; small unilamellar vesicles spread immediately along the extradural space and entered the systemic compartment 30 min after injection. Well-delineated liver and kidney labellings were seen after 24 h. The use of 3H-cholesterol-labelled small unilamellar vesicles suggested hepatic capture of intact liposomes with sizes averaging 0.05 microns drained unmodified into the systemic circulation through the extradural lymphatics. These results have led to the selection of multilamellar vesicles (0.1-15 microns size range) for clinical trials using liposome-associated local anaesthetics.