Nigerian sprint hurdler Samuel Okon has tested positive for the banned stimulant methylhexaneamine, Games Federation chief Mike Fennell said on Tuesday.
Compatriot and women's 100 metres gold medallist Osayemi Oludamola tested positive for the same substance and was suspended on Monday pending the analysis of her B sample.
"We have had approximately 1,200 tests up to last night and I again regret to inform that we have had a second anti-doping violation," Fennell told a news conference.
"This is also a Nigerian athlete, Samuel Okon, in the 110m hurdles and for the same substance, methylhexaneamine," he added.
Okon, who finished sixth in the sprint hurdles final last Friday, had waived the right for his B sample to be tested, Fennell said, although the Federation were waiting for personal confirmation of this.
"I heard about the second test and as the chef de mission of the Nigerian contingent, I am really concerned," Elias Usman Gora said.
Methylhexaneamine, a stimulant commonly used in nasal decongestants, was added to the banned list of the World Anti Doping Agency (WADA) for this year.
Media reports in Nigeria said Oludamola blamed a toothache remedy for the traces of the banned substance being found in her sample but Fennell suggested nutritional supplements might be to blame.
"The same substance is coming up and at this stage I cannot speak very definitively as to where it is coming from but it appears to us that it may be coming from the use of supplements," he said.
The result of Oludamola's B sample is expected to be known on Wednesday when she could be stripped of the gold medal she won when Australian Sally Pearson was disqualified for a false start.
Natasha Mayers, who crossed the line third in the blue riband sprint to take St Vincent and the Grenadines' first athletics medal, will be elevated to gold medallist if Oludamola is stripped of the title.
Mayers herself served a two-year doping ban after testing positive for testosterone in 2005.