Reaction #1121558

Reaction equation

doxorubicin HCl

→

doxorubicin

Solvents

desferal

Conditions

Workup

1. 1
   workup.ADDITIONwas added to I ml of the liposome dispersion after the liposomes
2. 2
   Filtrationfiltrated on a Sephadex G-50 column in Step B
3. 3
   Othershorter than less than 15 minutes
4. 4
   workup.ADDITIONA column containing 1-2 g
5. 5
   Otherdry weight of the resin
6. 6
   Concentrationthe phospholipids and doxorubicin concentration
7. 7
   ConcentrationPC concentration
8. 8
   workup.ADDITIONwere added to the assay mixture, in order
9. 9
   Otherto avoid partitioning of doxorubicin into the organic phase of the assay

Procedure

1 ml of a 10 mg/ml solution of doxorubicin HCl dissolved in saline-desferal was added to I ml of the liposome dispersion after the liposomes were gel filtrated on a Sephadex G-50 column in Step B. The mixture was incubated at room temperature for approximately 24 hours. The incubation was shorter for certain types of kinetic experiments. At the end of incubation period, the mixture was passed through a Dowex 50 WX-4 (Serva) column to adsorb the free, unincorporated drug. As small amount as 60 mg of the resin was able to adsorb as much as 1 mg of free doxorubicin in a time shorter than less than 15 minutes. Liposome incorporated doxorubicin did not adsorb to the Dowex resin at all and remained in the liposomes. A column containing 1-2 g dry weight of the resin was sufficient to adsorb all unincorporated drug. To determine the ratio of incorporated drug to liposomal lipid, the phospholipids and doxorubicin concentration were determined. PC concentration was determined according to Anal. Biochem., 104: 10-14 (1980) modified so that 20 ul of 10M HCl were added to the assay mixture, in order to avoid partitioning of doxorubicin into the organic phase of the assay. EPC was used for a standard curve.

Source