Abstract: To study the effect of carbonaceous matter on gold recovery in bio-pretreatment followed by a cyanide leaching process, the carbonaceous matter was characterized in high-sulfur Carlin-type gold flotation concentrate by gas chromatography-mass spectrometry (GC-MS), Raman spectroscopy, Preg-robbing index tests, and gold loading tests. The results indicate the presence of kerogen in the concentrate, which interferes with the gold extraction due to its strong gold adsorption. Both the Raman and PRI tests show a strong preg-robbing capacity of the carbonaceous matter in the samples. In addition, activated carbon in the cyanidation process of the bio-oxidized product could decrease by approximately 10.14% the adsorption of exposed gold by the carbonaceous matter. A final gold recovery rate is 80.17%.