Based on the Raman spectrocopy of the nanostructure-based SERS chips, the trace amount of methamphetamine and its primary metabolite, i.e., amphetamine, were successfully identified from human urine. The SERS substrates were self-assembled vertically-aligned silver nanorods prepared by the magnetron sputtering technique. The matrix spikes of the human urine specimens were prepared to simulate those of the methamphetamine abusers. With the acidulation treatments to the specimen samples, the organic urea-based byproducts were clearly separated. The remaining dissolved metham-phetamine/amphetamine in the urine specimens could therefore be enhanced for the SERS analyses, without any interference from the urea. The performance of the proposed acidulation treatments toward the SERS detections of the matrix-spike human urine specimens yielded the detection limit at 50 ng/mL. (C) 2016 Elsevier B.V. All rights reserved.