Display omitted •The Carbon Black was characterized by MEV, MET, EDX, DRX and RAMAN.•An ultrathin CB film was applied in the development of an electrochemical sensor.•AA, DA, UA and PAR were simultaneously determined using a CB/GCE electrode.•The new DPV method was used in the determination of these analytes in biological fluids. A novel material for the simultaneous electrochemical determination of ascorbic acid (AA), dopamine (DA), uric acid (UA) and paracetamol (PAR) using a composite based on carbon black film at glassy carbon electrode (CB/GC) has been evaluated. The morphology, structure and electrochemical performance of the composite electrodes were characterized by transmission electron microscopy, scanning electron microscopy, Raman spectra and cyclic voltammetry. The transmission electron microscopy and scanning electron microscopy images showed good distribution of the carbon black with a nanoscale particle size. The electrochemical measurements demonstrated the high-performance electrocatalytic of CB/GC electrode. Differential pulse voltammetry was applied to simultaneously detect AA, DA, UA and PAR levels in biological samples.