Preparation of marine active substances by bioprocessing technologies is a hot research topic in recent years. During the bioprocessing, it is of great significance to achieve rapid detection of the target products, while the traditional detection methods are faced with many limitations, so there is an urgent need for novel and simple analytical methods. In this study, phosphatidylserine (PS) was chosen as the model target, a colorimetric aptamer-based biosensor (aptasensor) was developed for the rapid detection of PS in bioprocessing system. The positively-charged chitosan oligosaccharides and negatively-charged aptamers firstly bound through electrostatic adsorption. In the presence of PS, the aptamers specifically bound with the PS, causing the release of chitosan oligosaccharides, which in turn adsorbed onto the negatively-charged gold nanoparticles to form nanozymes with high peroxide catalytic activity towards 3,3',5,5'-tetramethyl benzidine (TMB), producing colorimetric signals which were positively correlated with the concentration of PS. The absorbance values were collected and processed into rate of change with comparison to that of the negative samples without PS, and the rapid quantification was thus achieved. The concentration of aptamers, the concentration of chitosan oligosaccharides, and the conditions of nanozyme catalysis reaction were optimized in detail. Under optimal conditions, the developed aptasensor exhibited a good linear detection range (0.50-50.00 nmol/L) and high sensitivity, with a limit of detection (LOD) as low as 93.84 pmol/L. Moreover, the aptasensor showed high selectivity, accuracy, and repeatability. Recovery rates of 95.63%-110.76% and RSD less than 9.00% were obtained, when analyzing the PS in the actual bioprocessing samples. Only 40 min was needed for the entire analysis. In short, an aptasensor was successfully developed for highly sensitive and rapid detection of PS. The study can provide new ideas and research basis for the rapid detection method of PS as well as other biological processed products.