This study aims to establish quantitative relationships between dry biomass, photosynthetic pigment concentrations, and optical density in cultures of lower phototrophs. The red marine microalga Porphyridium purpureum (Bory) Drew et Ross and the cyanoprokaryote Arthrospira (Spirulina) platensis North were used as model organisms and were cultivated in open ponds under natural illumination. In the experiments, dry biomass was measured together with optical density at 545, 620, 680, and 750 nm. Pigment concentrations (chlorophyll a, B-phycoerythrin, and C-phycocyanin) were determined using standard biochemical assays and by deconvolution of true absorption spectra with Gaussian curve fitting. Linear correlation coefficients between dry biomass and optical density at 750 nm (D₇₅₀) were determined, and their values were similar for both species. Pigment concentrations calculated by biochemical and mathematical methods showed linear correlations when standard extinction coefficients were applied. For Porphyridium purpureum, the slopes of the linear correlations were 0.96 for chlorophyll a and 0.99 for B-phycoerythrin. For Arthrospira platensis, the corresponding values were 1.46 and 1.3 for chlorophyll a and C-phycocyanin, respectively. An alternative approach was developed based on culture optical density ΔD_λ and an empirically determined coefficient. When the optical path length was doubled, the ratio of pigment concentration to ΔD_λ decreased twofold for Porphyridium purpureum, whereas for Arthrospira platensis the coefficients differed by a factor of 1.8–1.9. The resulting equations for Porphyridium purpureum and Arthrospira platensis enable rapid estimation of chlorophyll and phycobiliprotein content in microalgal biomass.