Background and Aim:Aflatoxins are secondary metabolites of a group of filamentous fungi that are produced in most foods, including cereals, dairy products, nuts, and spices, under favorable environmental conditions. The aim of this study was to determine and compare the amount of aflatoxin B1 in bulk and packaged black pepper in Sanandaj city. Materials and Methods:The present study was a cross-sectional analysis on 15 samples of bulk black pepper sold in the traditional market of Sanandaj city and 5 samples of this product in packaged form belonging to 5 reputable brands, which were collected and tested in a simple random manner. The ELISA (Enzyme-linked immunosorbent assay, ELISA) method was used to analyze the samples. Data analysis was performed using SPSS software and ANOVA statistical test. Results:Aflatoxin B1 was found in all samples and its concentration was determined in the range of 0.003 to 1.958 μg/kg. The mean concentration of aflatoxin B1 in bulk powdered samples was significantly higher than that in whole bulk samples, but there was no significant difference between the mean concentration of aflatoxin B1 in bulk and packaged black pepper. However, the concentration of aflatoxin B1 in none of the samples exceeded the permissible limit of the European :union: standard and the Iranian national standard (5 μg/kg). Conclusion: Given the presence of aflatoxin B1 in black pepper samples and the relatively high consumption of this spice, it is recommended to regularly monitor the processing and storage of this product. Keywords:Aflatoxin B1, Aspergillus fungus, Piper nigrum, ELISA