A series of acylated chitosan oligosaccharides (LCSOs) were synthesized by reacting chitosan oligosaccharide (CSO) with lauroyl chloride in methane sulfonic acid. The chemical structures of LCSOs were characterized by Fourier transform infrared. Differential scanning calorimetry (DSC) showed that LCSOs had two phase transitions during heating. Thermoplastic chitosan oligosaccharide (LCSO-4) has distinct melting temperature (Tm) at 64 °C and 110°C. X-ray diffraction (XRD) analyses indicated that the crystal structure of CSO was changed through acylation and had created new crystal domains of lauroyl side chains. New strong diffraction peaks were observed around 2θ values of 20.0°, 21.5° and 26.3° for thermoplastic acylated chitosan oligosaccharide (LCSO-4). The melting and crystallization properties of thermoplastic acylated chitosan oligosaccharide were observed by polarized optical microscope (POM).