New psychoactive substances (NPS) are emerging illegal substances or synthetic drugs that pose public health threats worldwide. This study was aimed at reporting the clinical characteristics of NPS and classical illicit substances used by patients who presented to the emergency room. We conducted a retrospective cohort study on patients with suspected illicit substance use who visited the emergency department (ED) with the suspicion of illicit substance use. We divided the patients into 4 groups based on the NPS testing results: NPS positive, NPS negative, NPS combined with classical illicit drugs (INPS), and subjects with negative testing results. The majority of patients in all groups were male. The NPS users were significantly younger than those with negative results on toxic testing (26.4 vs 37.5,P= .005 < 0.05). The heart rate of NPS users was significantly faster than that of the group with negative results of toxic testing (111.1 vs 93.5 beats per minute,P= .046). The heartbeats of INPS group were also significantly faster than those with a negative result in toxicology screen (119.6 vs 93.5 beats per minute,P= .024). Those who used classical illicit drugs combined with NPS had significantly higher palpitation than those with negative results of toxic testing (27.3% vs 3.1%,P= .017). Patients who were highly suspicious of NPS use were younger, had tachycardia, felt palpitations, and had fair oxygen saturation compared to patients who were negative for urine toxicity screening.