Excessive phosphate could cause not only eutrophication of environment water, but also human diseases, such as hyperphosphatemia. Hence study of analytical technology of phosphate becomes important. This report based on silanized multi-walled carbon nanotubes (MWCNTs-SH) combined with Au nanoparticles (AuNPs) was presented for the quick and cost-effective determination of inorganic phosphate in freshwater. The MWCNTs-SH could realize approximately 2-fold increase in catalytic response current of molybdophosphate complex, in comparison with pristine MWCNTs. That could be interpreted that nanotube structures with nano-defects and conical shaped emission centers formed on the outer of wall could increase effective surface area of electrode and surface potential of electric double layer. Square wave voltammetry (SWV) was used to quantify the concentration of phosphate, and a sequential injection (SI) system was employed to realize automated solution handling and on-line monitoring in the design drawing. The sensor presented short response time of 8 s to phosphate with linear range from 0.5 to 15 mg L−1 and a detection limit of 0.3 mg L−1. Importantly, the favorable recovery rates of 106.6% ± 10.33% were achieved by detecting real water, and the data were verified by the standard spectrophotometry, showing that the work provided an efficient and promising strategy for phosphate monitoring in the environment filed.