The adulteration of urine samples is an ongoing problem in forensic drug-testing laboratories, even in the military where the practice of observed collections is performed. These adulterants are used to produce a false-negative result when samples are analyzed for drugs of abuse. It has been reported that papain, a cysteine protease, could be successfully used as a urine adulterant, altering the concentration of 11-nor-Delta9-tetrahydrocannabinol-9- carboxylic acid (THCCOOH) in urine samples. The current study analyzes the effects of latex papain (Sigma, 10 mg/mL) and Lawry's Adolph's Meat Tenderizer (papain is an active ingredient, 10 mg/mL) on immunoassays (FPIA, EMIT, KIMS) and gas chromatography-mass spectrometry (GC-MS) analysis for biological samples. The samples were analyzed initially between 2 and 4 h and then at 1-, 3-, 7-, and 10-day time intervals after the addition of papain. A decrease in response averaged over the course of the study was observed with FPIA (Abbott, 22%) and EMIT (Syva) Dade Behring, 26%, Microgenics, 10%) screening assays by the addition of latex papain to the samples. An increase in response was found using the KIMS (Roche) assay (156% increase). In addition, the GC-MS results (27% decrease) demonstrate that papain affects both the screening and confirmation assays. The addition of meat tenderizer caused decrease in the FPIA (Abbott, 11%) screening assay and GC-MS results (22%) similar to the latex papain while having varied results on the other screening assays. This study confirms papain could be a potential problem for urine drug-testing programs.