The accuracy of a drug test hinges on the integrity of the collected urine sample. Testing facilities use a multi-layered verification process to ensure the specimen is valid, meaning it is fresh, undiluted, and has not been substituted or tampered with. This process detects changes that occur naturally when a sample is collected hours or days prior, which defines “old” urine in this context. Laboratories use physical and chemical checks to look for markers inconsistent with a newly voided, natural human sample. The full validity assessment involves instant physical checks followed by quantitative laboratory analysis of specific chemical components.
Initial Checks for Sample Validity
The first line of defense against sample substitution or adulteration occurs immediately upon collection with a temperature check. A newly voided urine specimen should fall within a narrow range, typically between 90°F and 100°F (32°C and 38°C), when measured within four minutes of collection. A temperature outside this range strongly indicates the sample has been substituted, perhaps with pre-collected, “old” urine that has cooled, or with synthetic urine.
A collector also performs a visual inspection to note any abnormal physical characteristics that might suggest tampering. This includes checking for unusual color or clarity, such as a sample that is too clear or has an unnatural hue. The presence of excessive foam, foreign materials, or an unusual odor can also indicate that an adulterant has been added to interfere with the drug assay. These initial observations are logged on the chain-of-custody form.
Detecting Chemical Degradation
A definitive way a laboratory identifies an old or improperly stored sample is by detecting chemical changes that occur outside the body. When urine is left at room temperature, it begins to undergo measurable biological and chemical degradation. This decomposition is primarily driven by bacterial action on the urea naturally present in the specimen.
Bacteria break down urea into ammonia, causing a significant shift in the sample’s pH level. Human urine typically has a pH between 4.5 and 8.0, but as ammonia accumulates, the urine becomes alkaline, with the pH rising to 9.0 or higher within a few days at room temperature. A moderately abnormal pH (9.0 to 11.0) is reported as “invalid.” A severely abnormal pH (over 11.0) is reported as “adulterated” because it is inconsistent with any human physiological state.
The drug metabolites the test is designed to detect can degrade or change structure over time, especially in a non-refrigerated environment. This chemical breakdown can make the quantification of target substances difficult or inaccurate, potentially causing a false-negative result. Increased storage temperatures accelerate the degradation of analytes and the rise in pH, providing a time-dependent chemical signature of a non-fresh sample.
Specific Gravity and Creatinine Measurement
Two objective and regulated measures for specimen validity are the quantitative analysis of creatinine and specific gravity. Creatinine is a metabolic waste product generated by muscle tissue and excreted in urine at a relatively constant rate. Federal guidelines establish a required reference range for a valid sample, typically requiring creatinine concentrations to be above 20 mg/dL.
A sample with a creatinine concentration below 20 mg/dL but above 2 mg/dL is reported as “dilute.” A concentration below 2 mg/dL is reported as “substituted,” meaning it is physiologically impossible to be natural human urine. Specific gravity (SG) measures the concentration of dissolved solids, comparing the density of the urine to the density of water. A low SG reading indicates that the urine is excessively watery.
The normal range for specific gravity is between 1.003 and 1.030. A reading below 1.003 indicates dilution, and a reading below 1.001 is physiologically impossible for human urine. These two markers confirm the sample is natural and concentrated enough to provide an accurate drug test result. While these measurements primarily detect dilution, an old or substituted sample often fails these concentration checks.
Consequences of a Compromised Sample
When a urine specimen fails any validity check—due to abnormal temperature, pH, creatinine, or specific gravity—it is reported by the laboratory as “invalid,” “adulterated,” or “substituted.” An “invalid” result means the lab was unable to obtain a clear positive or negative result, often due to an unidentified interfering substance. An “adulterated” result means a substance was found that is not a normal constituent of urine, or an endogenous substance is present at an abnormal concentration.
In a regulated testing environment, a failed integrity check is a serious procedural outcome. A result reported as “substituted” or “adulterated” is treated as a refusal to test, which often carries the same consequence as a confirmed positive drug test for employment purposes. An “invalid” or “dilute” result usually requires the person to provide a new sample immediately, often under direct observation.