This dataset comprises samples collected in the Eastern Gotland Basin (Central Baltic Sea) during the EMB295 CenBASE research cruise conducted from June 30th to July 19th 2022. Sea surface microlayer (SML) samples were collected using established methods, i.e. the glass plate and the Garrett screen. Reference samples from the underlying water (ULW) were collected at approximately 1 m depth using a manually deployed Niskin bottle. To characterize the biochemical composition of the SML and the ULW, water samples were collected twice daily for the analysis of total organic carbon (TOC), total amino acids (TAA), total combined carbohydrates (TCCHO), and surfactants (determined using phase-sensitive AC voltammetry). TOC was measured using a TOC analyser. High-performance anion-exchange chromatography coupled to pulsed amperometric detection (HPAEC-PAD) was performed for carbohydrate analysis. Amino acids were determined using a high-performance liquid chromatography (HPLC). Chlorophyll a samples were taken from the Niskin bottles at 1m depth (ULW) and analysed using a fluorometer from Turner Designs (Turner TD 10-AU005). Abundance and community composition of nano- and picophytoplankton (20µm (POC>20µm), particulate amino acids >20µm (PAA>20µm), and particulate combined carbohydrates >20µm (PCCHO>20µm). POC was measured using an Euro EA elemental analyser. To determine the composition of the microphytoplankton community (>20 µm), samples were preserved with Lugol's solution and analyzed according to the Utermöhl method. Sampling stations were subsequently classified based on the abundance of microphytoplankton (>20 µm) and nano/picophytoplankton (≤20 µm). Hydrographic parameters, including salinity and temperature, were obtained from CTD measurements at a depth of 3 m to characterize the prevailing hydrological conditions.