To investigate the diurnal variation in phytoplankton biomass and its regulating factors during the diurnal cycle, we conducted in situ observations in June 2018 at three buoy stations, including Douwei Buoy Station, Minjiang Estuary Buoy Station, and Huangqi Buoy Station on the western side of the Taiwan Strait. The calibration of buoy sensor data, including temperature, salinity, dissolved oxygen, pH, chlorophyll, and phycoerythrin, was conducted simultaneously. In addition, water sampling was conducted to measure chlorophyll a and phycoerythrin concentrations at hourly time intervals. The results showed that the 24 h cumulative chlorophyll a concentration order for the buoys was Minjiang Estuary (10.280 μg/L) > Huangqi (7.411 μg/L) > Douwei (4.124 μg/L). The Minjiang Estuary had a lower nighttime biomass proportion than Douwei and Huangqi. The diurnal variation in phytoplankton was jointly regulated by water masses, tides, and light. There were three response patterns, including the “light trumps tidal influences” pattern at Douwei, the “Low-tide, High-biomass” pattern at Minjiang Estuary, and the “High-tide, High-biomass” pattern at Huangqi. The prediction of algal blooms and hypoxia using buoy monitoring needs to be based on seasonal water mass background and tidal influence.