We examined the within-day variability in seawater optical properties and biogeochemical constituents for a high-latitude location in the Ross Sea, Antarctica, during development of the annual spring phytoplankton bloom. Measurements of particulate organic carbon concentration (POC), chlorophyll-a concentration (Chl), and particle size distribution were conducted at 4-6 hour intervals in parallel with determinations of the spectral absorption and attenuation coefficients of particles, and the spectral remote-sensing reflectance of the surface ocean (Rrs). Surface POC and Chl exhibited more than a twofold variation throughout the day in the continuous presence of natural light. A minimum occurred near local noon coinciding with peak solar irradiance, a maximum in the evening, and a subsequent decrease throughout the night-time hours. These patterns were accompanied by large changes in the magnitude and spectral shape of Rrs, including the blue-to-green spectral band ratios used in ocean colour algorithms for estimating POC and Chl. The variability in Rrs could not be explained by changes in solar zenith angle, but was consistent with observations of within-day variations in spectral absorption and scattering by particles which were influenced by changes in the particle concentration and size distribution. The accuracy of an empirical ocean colour algorithm for estimating POC from Rrs was unaffected by within-day variability, implying that short-term variations in surface POC can be potentially monitored by multiple within-day measurements of Rrs, through means of in situ and remote sensing observations if available. Our findings also suggest that within-day changes in POC can be significant compared with the variability observed on meso-scale spatial scales, potentially confounding the interpretation of remote-sensing data obtained from temporal and spatial compositing of images measured at different times within a single day. © 2014 © Taylor & Francis.