Abstract Acetylation in fibrous state of holocellulose, its alkali-refined product, rayon pulp and Bemberg rayon was undertaken to disclose the effect of pentosan and the structure of holocellulose. 1) Acetylation of air-dried holocellulose is roughly controlled by surface resistance regardless of various ratios depending on conditions of acetylation and degree of refining. The rate of pretreated sample seems to be determined by internal diffusion. 2) The birefringence decreases at first abruptly and then increases and finally decreases again. The final higher birefringence compared to acetic acid content may be ascribed to acetolysis. Definite amorphous portion is acetylated easily at the initial stage and lost, so that residual crystallinity, and also birefringence, increase. 3) By alkali-refining, acetylation is retarded and initial abrupt change, which is caused by the amorphous portion is diffused. There is optimum point in the concentration of refining alkali, probably relating to hornification, loss of amorphous portion, transformation of crystal form and hindrance by residual pentosan layer. The refining temperature affects the acetylation slightly. 4) Pentosan in product is not reduced apparently, probably due to oxymethylfurfural yielding and poor solubility of acetate. Also pentosan may not be entirely accessible at initial stage.