Environmental DNA (eDNA) metabarcoding is a valuable tool for assessing aquatic biodiversity, but the high cost and complexity of DNA extraction pose challenges for widespread adoption, especially in developing countries. This study presents a cost-effective eDNA extraction method using a guanidine hydrochloride (GuHCl) buffer, proteinase-K digestion, and isopropanol precipitation to improve the detection of fish communities. Comparison with the Qiagen DNeasy Blood & Tissue Kit using MiFish universal primers showed that the GuHCl protocol detected more fish species in freshwater samples, with comparable performance in relative read abundance metrics. However, the GuHCl method exhibited higher PCR inhibition in brackish samples, likely due to salinity and natural inhibitors. The results suggest that the GuHCl-based method is a viable alternative, offering enhanced sensitivity for low-abundance species in freshwater samples and cost savings. This protocol facilitates large-scale eDNA metabarcoding for ecological studies and conservation management efforts.•The GuHCl protocol identified a greater diversity of fish species in freshwater samples than the Qiagen kit, but detected fewer species in brackish water samples.•Both extraction methods demonstrated robust positive correlations in metrics of relative read abundance.