In addition to N-nitrosoproline (NPRO), N-nitrosamino acids of a new type containing sulfur were found as major N-nitroso compounds in the urine of seven healthy subjects. These compounds were detected using a gas chromatograph connected with a Thermal Energy Analyzer (GC-TEA), and they were identified as N-nitroso-L-thiazolidine-4-carboxylic acid (N-nitroso-L-thioproline, NTPRO) and the cis and trans isomers of N-nitroso-L-methylthiazolidine-4-carboylic acid (N-nitroso-L-methylthioproline, NMTPRO) by using gas chromatography-mass spectrometry (GC-MS). The amounts of NPRO, NTPRO and NMTPRO found in 100 mL samples of human urine were 65-471 ng, 67-2 250 ng and 275-5 825 ng, respectively. L-Thioproline was nitrosated 20-60 times faster than L-proline by nitrite over the whole range of acidic pH. The precursors of these new-N-nitrosamino acids containing sulfur may be formed by the reaction of L-cysteine with formaldehyde and acetaldehyde in the human body. These sulfur-containing N-nitroso compounds, NTPRO and NMTPRO, in addition to NPRO, could be useful as probes for investigation the dynamics of nitrosation in the human body.