As a chemical agent, malachite green is used as a fungicide in aquatic products because of its good bactericidal properties, low price and a small amount are imperceptible. However, because its metabolite in the body——the recessive malachite green has stability and highly toxic chemical properties. Malachite green has been classified as a banned drug in China. This experiment discusses how to use three-dimensional fluorescence spectroscopy to detect recessive malachite green in aquatic products, and use three-dimensional fluorescence spectroscopy to explore the three-dimensional characteristics of recessive malachite green, and further analyze the relationship between fluorescence intensity and pH change. Finally, fluorescence spectroscopy is carried out for fish samples to discuss the feasibility of three-dimensional fluorescence spectroscopy. The accumulation data and foundation of recessive malachite green in aquatic products are detected by three-dimensional fluorescence spectrometry.The experimental results show thatthe highest peak of the recessive malachite green ethanol solution was obtained at the excitation wavelength of 324 nm/the emission wavelength of 355 nm. The highest peak of recessive malachite green methanol solution was obtained at the excitation wavelength of 320 nm/the emission wavelength of 355 nm. The experimental results show that the fluorescence peak strength is strongest in the pH 7.0 environment. According to the experience of fish sample, we can find that the relationship between the concentration of recessive malachite green and the fluorescence intensity showed a good linear relationship. The recovery rate was 96.65%, which proved that the experimental results have good stability. Therefore, the recessive malachite green in aquatic products can be accurately and quickly detected by three-dimensional fluorescence spectroscopy technology.