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Makeup water constitutes a key component in the water management of microalgal cultivation systems. However, the effect of makeup water addition on microalgal growth remains largely unexplored. This study compared two deionized water addition intervals (1 day and 4 days) for their effect on the growth of <i>Scenedesmus dimorphus</i> (<i>S. dimorphus</i> hereafter) in 2000 mL Pyrex bottles under controlled conditions. Cell counts and dry algal biomass (<i>DAB</i>) were measured to characterize the microalgal growth rate. Water addition intervals impacted algal cell counts but had little effect on <i>DAB</i>. Adding makeup water every day resulted in a higher growth rate (8.80 ± 1.46 × 10⁵ cells mL⁻¹ day⁻¹; <i>p</i> = 0.22, though) and an earlier occurrence of the peak cell count (day 9) than adding it every 4 days (6.95 ± 1.68 × 10⁵ cells mL⁻¹ day⁻¹ and day 12, respectively). It is speculated that water loss over an extended period and the following makeup water addition posed stress on <i>S. dimorphus</i>. Surpassing the peak cell count, <i>S. dimorphus</i> continued to grow in <i>DAB</i>, resulting in an increased cell weight as a response to nutrient starvation. Optical density at 670 nm (OD₆₇₀) was also measured. Its correlation with <i>DAB</i> was found to be affected by water addition intervals (R² = 0.955 for 1 day and 0.794 for 4 days), possibly due to a water loss-induced change in chlorophyll <i>a</i> content. This study is expected to facilitate the makeup water management of photobioreactor and open pond cultivation systems.