ABSTRACT Blend nanofibres composed of synthetic polymers with biological macromolecules, such as natural biopolymers, and 3D structures created via electrospinning technique, have a high potential for modification to promote cell growth and function. In this study, 5 and 7 wt% of glucosamine sulfate (GAS), as one of the key components in the extracellular matrix (ECM) of natural cartilage tissue, were added to poly (3-hydroxybutyrate)-chitosan (PHB-CS)/functionalized multiwalled carbon nanotubes (f-MWCNTs) solution (100:20:1) for production of electrospun scaffolds. Prepared fibrous scaffolds are characterized by SEM, FTIR, XRD, TGA, and DSC. Tensile tests are used to study their mechanical properties and their hydrophilicity is also assessed. subsequently, the drug release profile of the scaffolds, adipose stem cell proliferation, cell viability, and the differentiation of adipose stem cells to chondrocyte cells were evaluated for further cartilage tissue engineering application. Our results showed that the addition GAS has a positive effect on the hydrophilicity of the fibers and the initiation of chondrogenic differentiation. Without, effects on tensile strength of the scaffolds by adding 5%. So, that PHB-CS/f-MWCNTs scaffold containing GAS has more acceptable properties than the PHB or PHB-CS for cartilage tissue engineering.