Several neutral glycosphingolipids were hydrogenated and subjected to trifluoroacetylation in trifluoroacetic acid/trifluoroacetic acid anhydride under conditions leading to complete exchange of the N-acetyl groups of GalNAc for N-trifluoroacetyl. The derivatized glycosphingolipids were analyzed for binding by P-fimbriated uropathogenic Escherichia coli, recognizing the globo series of glycolipids (carrying Galα1-4Gal). Using E. coli it was shown that a GalNCO-CF3 next to the minimum binding epitope Galα1-4Gal did not substantially influence the binding, as did not a trifluoro acetyl group on the ceramide. Exchange of N-acetyl of GalNAc in the receptor active gangliotetraosylceramide, Galβ1-3GalNAcβ1-4Galβ1-4Glcβ1-1Cer, for N-trifluoroacetyl, did not change the binding of two out of the three strains tested of the bacterium Neisseria subflava. Discussion concerning the binding epitopes of the bacterial adhesins to carbohydrates is based on these results.