Mycotoxins, toxic secondary metabolites from certain fungi, present significant health risks to humans and animals. Although their effects on living organisms have been widely studied, their presence in postmortem tissues, especially the brain, is underexplored in forensic analysis. Understanding the quantitative assessment of mycotoxin levels in postmortem brain tissue is vital due to the brain's critical role and its susceptibility to mycotoxin induced damage. Objective: This study aimed to develop an analytical method for quantitatively evaluating mycotoxin levels in postmortem brain tissue and investigating their forensic implications. Method: An observational cross-sectional design was used to analyze postmortem brain tissue samples from forensic autopsies for mycotoxin content via high-performance liquid chromatography coupled with mass spectrometry (HPLC MS). Standardized protocols and quality control measures ensured accurate and reliable mycotoxin measurements. Results: Detectable levels of aflatoxins, ochratoxins, and trichothecenes were found in postmortem brain tissue samples. Mycotoxin levels varied across demographic groups and brain regions, with higher levels observed in individuals diagnosed with neurological disorders compared to healthy controls. Conclusion: This study highlights mycotoxin exposure's prevalence and potential forensic implications in postmortem brain tissue, underscoring the need for further research on mycotoxin-induced neurotoxicity and standardized analytical protocols