Early steps of ribosomal maturation have been studied by analysis of nucleolar extracts using different extraction procedures. Early 45‐S nucleolar RNA is found to be associated with slowly sedimenting elements, distinct from previously described 80‐S and 55‐S nucleolar preribosomes. This early 45‐S RNA has been shown to be of preribosomal type according to the following criteria. (a) When hybridized with nucleolar DNA, competition with rRNA can be observed; (b) its biosynthesis is sensitive to low doses of actinomycin D; (c) it is methylated at an early stage; (d) it contains no linked poly(A) segments. This early 45‐S RNA seems to be specifically linked with proteins. The protein content of these early RNA‐protein complexes is significantly lower than that for 80‐S preribosomes. 45‐S RNA sensitivity to nucleolytic activities that can take place in the course of preribosome extraction has been found to be higher in these RNA‐protein complexes than in 80‐S preribosomes. Identical results were obtained when different mammalian cell species were studied (rat, hamster, or HeLa cells).