During the Tianmushan tunneling, some of the working staffs suffer from acute poisoning symptoms, such as bloodshot puffy eyes and red swollen skins followed by exfoliation. In order to find the causes of the acute poisoning, the hazardous substance that results in the acute poisoning is identified by monitoring the dust concentration variation in the tunnel and analyzing the chemical components of the dust. The detection results show that: 1) The dust concentration in the working zone can be lowered below exposure threshold, that is 1 mg/m3, after 10 min of ventilation with an air volume 2 274 m3/min after blasting. 2) The dust concentration in the working zone is 56 times exposure threshold during downthehole (DTH) drilling, which would causes acute poisoning symptoms of working staffs. 3) The concentrations of vanadium, nickel, manganese and chromium in the dust and those of arsenic, copper, chromium, nickel and vanadium in surrounding rocks are high, and the concentrations of vanadium and arsenic in section that acute poisoning of working staffs occurs are relatively high as well. The detection results coincide with the symptoms of vanadium and arsenic poisoning, which shows that the main causes of acute poisoning of working staffs are dust containing vanadium and arsenic.