The level of uptake and retention of amino-containing drugs in large unilamellar vesicles (LUVs) following uptake in response to a transmembrane pH gradient (DeltapH) can vary dramatically depending on the drug. For example, the anticancer drugs doxorubicin and epirubicin can be readily retained, whereas the anticancer drug vincristine and the antibiotic ciprofloxacin tend to leak out rapidly. In this investigation, we examine the influence of the hydrophobicity of the entrapped amines (that induce the DeltapH) and the anionic lipid content of the LUV on drug retention. It is shown that entrapment of increasingly hydrophobic monoamines (methylamine to amylamine) all lead to an induced DeltapH of 3 units and essentially complete drug uptake under the conditions employed, but do not lead to improved retention of vincristine and ciprofloxacin. However, significantly improved retention could be achieved by substitution of the anionic lipid distearoylphosphatidylglycerol (DSPG) for distearoylphosphatidylcholine (DSPC) in the LUV bilayer. Further, it is shown that if the induced DeltapH is reduced to 1.4 units (driven by entrapped diamine) nearly 100% accumulation of doxorubicin and epirubicin could be achieved, whereas only 25% loading for vincristine and ciprofloxacin was observed. Taken together these results provide methodology for improving (weak base) drug retention in liposomes and indicate that drugs that can partition into the lipid bilayer exhibit improved uptake and retention characteristics.