Unexpected benefits to the catalytic performance of materials often originate from the presence of surface defects. Here, novel D-penicillamine modified molybdenum oxide nanodots, with abundant oxygen vacancy defects, were fabrication by a mild, simple, and cost-effective method. Ultraviolet–visible (UV–Vis) absorption spectra analysis showed that the nanodots had peroxidase-like and catalase-like activities. The reactive oxygen species were probed by electronic paramagnetic resonance technique and spectroscopic methods, demonstrating that the nanodots also had oxidase-like activity. Interestingly, the peroxidase-like activity of nanodots was synergistically enhanced in the presence of ferrous ions or ferric ions. Remarkably, less than nanomolar levels of ferrous ions were required to display this phenomenon, meaning Fenton reagent acted as leverage. Based on this, a sensitive colorimetric and fluorescent dual-mode sensor for alendronate sodium was developed. The linear ranges for colorimetric and fluorescence analysis were 0.2–2.5 and 0.2–2.0 μM, with detection limits of 31.21 and 71.84 nM, correspondingly. The method has a simple large-scale material preparation process with higher sensitivity and shorter reaction time, which can inspire and enlighten the design of nanozyme sensors.