A droplet-based electrochemical sensor for direct measurement of D-glucosamine was developed using carbon paste electrodes (CPEs) modified with gold nanoparticles (AuNPs) and polyaniline (PANI). Central composition design (CCD) was employed as a powerful method for optimization of parameters for electrode fabrication. The optimized amounts of AuNPs and PANI obtained from the response surface were determined as 300 and 3000mgL(-1), respectively. Coupled with a droplet microfluidic system, the analysis of glucosamine was performed in a high-throughput manner with a sample throughput of at least 60 samples h(-1). In addition, the adsorption of the analyte on the electrode surface was prevented due to compartmentalization in droplets. Linearity of the proposed system was found to be in the range of 0.5-5mM with a sensitivity of 7.42×10(-3)Amol(-1)Lcm(-2) and limits of detection and quantitation of 0.45 and 1.45mM, respectively. High intraday and interday (evaluated among 3 days) precisions for the detection of 50 droplets containing glucosamine were obtained with relative standard deviation less than 3%. The system was successfully used to determine the amounts of glucosamine in supplementary products with error percentage and relative standard deviation less than 3%. In addition, the amounts of glucosamine measured using the developed sensor were in good agreement with those obtained from a CE method. These indicate high accuracy and precision of the proposed system.