A novel family of cationic steroids has been synthesized and found to display potential both in non-viral gene delivery and as antimicrobials. The compounds were formed by amide couplings of the polyamines spermine, spermidine, and putrescine with cholenic and cholanic acid derivatives and span a hydrophobicity range of 0.64 to 5.63 logD (octanol-water distribution coefficients). On liposomal formulation with the neutral helper lipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 5β-cholanyl-spermide-3α-ol and 8-cholenyl-spermide-3α,12α-diol were the most successful as gene delivery vectors achieving transfection efficiencies comparable to those found with Lipofectamine2000™ in bovine aortic endothelial cells. The toxicity to the cells was reduced by nearly half that of Lipofectamine2000™ for both of these compounds. For the more hydrophilic 5β-cholanyl-spermide-3α,7α,12α-triol, transfection efficiency was enhanced by over 50% through incorporating 20-30mol% bis-substituted amide in the liposome formulation without increasing the toxicity. Evaluation of antibacterial activity against gram positive bacteria (B. Subtilis ATCC 6051) revealed MIC₅₀ (concentration required to inhibit the growth of 50% of the organism) of less than 25 mg/L for the majority of the compounds. When tested against gram negative bacteria (E. Coli MG 1655), the MIC₅₀ values of 5β-cholanyl-spermide-3α-ol and 8-cholenyl-spermide-3α,12α-diol were less than 25 mg/ml and the effective therapeutic indexes (ratio of hemolytic activity over MIC₅₀) were 37 and 53, respectively.