Aminolevulinic acid (ALA), ALA methylester (ALA-Me) and ALA hexylester (ALA-Hex) were topically applied for 5 and 20 hr, respectively, on normal skin of mice. The distribution of protoporphyrin IX (PpIX) induced in 7 different tissues by these drugs was determined either by spectrofluorometric measurements with an optical fibre probe or by chemical extraction of PpIX from the tissues. The results from these 2 types of measurements were compared. Both methods showed that ALA and the esters induced similar amounts of PpIX at the skin spot where they were applied and that the esters produced much less PpIX at remote skin spots (i.e., spots outside the location where the drugs were applied) than ALA did, notably after 20 hr application. After 20 hr of drug application ALA produced much more PpIX in liver, intestine and lungs than the esters did. In contrast with the direct fluorescence measurements, the extraction method showed detectable amounts of PpIX in liver, intestine and lung after application of the esters, notably of ALA-Me. The discrepancy is probably related to the fact that the pigmented tissues absorb light and, therefore, the direct fluorescence readings are misleading. Notably in the liver, which contains high concentration of light-absorbing pigments, very weak direct fluorescence was seen. In no case there was any accumulation of PpIX in muscle tissue nor in brain. The esters seem to penetrate less into the circulation than ALA, and PpIX formed by them in the skin is faster cleared than PpIX formed from ALA. This is also true after oral and i.p. administration of the drugs. © 2002 Wiley-Liss, Inc.