The particle-induced X-ray emission (PIXE) of thick biomineral targets provides pertinent surface analysis, but if good reference materials are missing then complementary approaches are required to handle the matrix effects. This is illustrated by our results from qualitative and semiquantitative analysis of biornaterials and calcified tissues in which PIXE usually detected up to 20 elements with Z > 14 per sample, many at trace levels. Relative concentrations allow the classification of dental composites according to the mean Z and by multivariate statistics. In femur bones from streptozotocin-induced diabetic rats, trace element changes showed high individual variability but correlated to each other, and multivariate statistics improved discrimination of abnormal pathology. Changes on the in vitro demineralization of dental enamel suggested that a dissolution of Ca compounds in the outermost layer results in the uncovering of deeper layers containing higher trace element levels. Thus, in spite of significant limitations, standardless PIXE analysis of thick biomineral samples together with proper additional procedures can provide relevant information in biomedical research.