Biomasses, both residual and dedicated grown energy crops, are suitable substrates for the digestion in agricultural biogas plants. Among energy crops, sorghum represents an interesting feedstock for methane production. The aim of this study was to investigate the effects of particle size on anaerobic digestion process with and without sodium hydroxide pretreatment of ensiled sorghum forage. Ensiled sorghum forage, used for animal feed, was collected from a farm near Cremona (Lombardy region, Italy). After collection, it was dried and ground using a cutting mill (Retsch) with a 2, 1, 0.5 and 0.25 mm screens. Mean diameters of fractions were 994, 471, 269 and 169 μm. Sodium hydroxide pretreatment tests were conducted by soaking samples (471and 169 μm particle sizes) in a NaOH solution at 55°C for 12 h, at different NaOH dosages (4 and 10 g NaOH/100gTS). Biochemical methane potential (BMP) tests were performed under mesophilic conditions (35°C), using plasma flasks (V = 0.5 L) closed with rubber septa. The substrate to inoculum ratio was between 1 and 1.1 gVS/gVS. To evaluate the effect of particle sizes on anaerobic digestion kinetics, the first order kinetic constants were calculated for each BMP test, by using least-squares fit of methane production data during time (t). To explain the results, chemical composition analysis and Infra-Red Spectroscopy analysis were performed. Infra-Red Spectroscopy was performed to study the variations of sample structure introduced by milling, and particularly the cristallinity of samples. A reduction of particle sizes, within the range studied, didn’t improve significantly the maximum methane production (from 288±4 NmLCH4/gVS to 298±4 NmLCH4/gVS) and the anaerobic digestion kinetic (0.11 d-1) of ensiled sorghum forage. No significant variation of volatile solids (VS), total organic carbon (TOC), total nitrogen (TKN) and protein content (TKN*6.25) were observed. No significant variation of Klason lignin content was also observed. Infra-Red Spectroscopy analysis revealed that milling didn’t change the composition of sorghum and the internal structure of cellulose. The lignin to cellulose ratio (ratio of absorbance between peaks at 1510 cm-1 and 898 cm-1) and the cellulose cristallinity index (ratio of absorbance between peaks at 1315 cm-1 and 2900 cm-1) were the same for all fractions. Moreover, particle sizes (of 471and 169 μm) had no significant influence in terms of maximum methane production after a sodium hydroxide pre-treatment at 4% NaOH dosage (299±4 and 302±2 NmLCH4/gVS, respectively) and at 10 % NaOH dosage (328±5 and 327±3 NmLCH4/gVS, respectively). No significant influence of the particle sizes on anaerobic digestion kinetic was also observed (kinetic constants were 0.14 d-1 and 0.16 d-1 at 4 % and 10% NaOH dosage, for both particle sizes).