Sandy beaches are dynamic ecosystems supporting diverse life forms and remain less explored in terms of biodiversity compared to other coastal systems. Environmental DNA (eDNA) coupled with sequencing technology has emerged as a non-invasive method for assessing biodiversity, particularly in understudied habitats such as sandy beaches. This study employed eDNA metabarcoding on sediments from four Uruguayan beaches (two urban and two rural) using 18S and COI markers to assess the eukaryotic diversity of eDNA found in sandy beaches. Sediments were collected from the intertidal zone, and DNA was extracted. PCR products were then obtained and subsequently sequenced using high-throughput methods. Taxonomic diversity was analyzed at the family level, categorizing molecular operational taxonomic units (MOTUs) into ecological groups (e.g. algae, fungi, benthos). The 18S marker identified 67% of MOTUs, outperforming the COI marker, particularly for algae, fungi, and microbenthos. Algae had the highest diversity (in terms of the number of MOTUs), followed by microbenthos and fungi, while parasites and zooplankton had fewer MOTUs detected. Beaches showed no clear eDNA diversity differences, although site-specific variations in MOTUs abundance were noted. Some taxa, such as harmful algae and pathogens, have both ecological and health significance. These results demonstrate the utility of eDNA in revealing the diversity of sandy beach DNA and detecting taxa of ecological concern. Expanding spatial and taxonomic coverage, increasing the number of samples, and refining methodologies could further enhance our understanding of biodiversity patterns in these ecosystems.