Experimental dataset on the characterization of a new photo-calorespirometry setup

This dataset complements the main article "Direct Real-Time Determination of Photosynthetic Efficiency by Photo-Calorespirometry Using Synechocystis sp. PCC 6803 as Model Cyanobacterium" by providing detailed experimental data, calibration procedures, and validation measurements for the photo-calorespirometry (Photo-CR) method. The data ensure precise quantification of light energy input ( $P_I$ ), thermal signals, and photosynthetic performance, and validate control systems essential for accurate heat-based assessment of photosynthesis in phototrophic microorganisms. Light delivery was optimized and in a dual-ampoule calorimetric setup trough calibrated LED-light guide assemblies. including distance-dependent attenuation profiles, peak $P_I$ determinations (20 mW), and corrections for thermal asymmetry. A photosynthetically inactive reference system was established by formaldehyde fixation of Synechocystis cells, preserving morphology and pigment content. Physiological and optical validations included Coulter counter cell-size analysis, pigment quantification, and absorption spectroscopy to confirm spectral similarity between fixed and living cells. Control datasets include thermal profiles of living versus dead cells, medium-only baselines, and light-independent heat generation. Additional measurements capture temporal variations in photosynthetic performance during multi-step light ramp experiments, normalized to biomass via Monod-based growth modeling. Furthermore, a correlation dataset from 18 independent samples links cell number, optical density (OD₇₅₀), and cell dry weight, enabling conversion between these measures. Collectively, these datasets provide a comprehensive reference for calibrating and validating Photo-CR experiments, supporting reproducible measurement of photosynthetic energy conversion in cyanobacteria. They are intended for reuse in methodological replication, adaptation to other phototrophic microorganisms, optimization of light-dependent calorimetric studies, and the development of standardized reference controls for heat-based photosynthetic efficiency assessment.