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Published:
Journal of Analytical Toxicology,
ISSN 0146-4760,
Volume 24,
Number 8, November/December,
pp. 708-714
Investigation of
Nitrite Adulteration on the Immunoassay and GC–MS Analysis of Cannabinoids
in Urine Specimens
Jane S.C.
Tsai1, Mahmoud. A. ElSohly2, Shiow-Fen Tsai1, Timothy P. Murphy2, Barbara Twarowska1,
and
Salvatore J. Salamone1
1Roche Diagnostics Corporation, 9115 Hague Road, Indianapolis, Indiana 46250 and
2ElSohly Laboratories, Inc., 5 Industrial Park Drive, Oxford, Mississippi 38655
Nitrite ion has been identified as the active ingredient of two commercial adulterants
that could cause discrepant results between the immunoassay screening and gas
chromatographic–mass spectrometric (GC–MS) confirmation of 11-nor-D9-tetrahydrocannabinol-9-carboxylic
acid (THCCOOH) in urine. Procedures to chemically convert the nitrite ion at
the beginning of sample preparation for GC–MS analysis may not overcome
all nitrite adulteration cases because portions of the THCCOOH might have been
lost between the time of sample collection and the time of analysis. This study
was conducted to further investigate the influence of both urine sample matrix
and the duration of nitrite exposure on nitrite interference of THCCOOH detection.
Forty clinical “THC-positive samples” that had been screened and confirmed
positive for the presence of THCCOOH were spiked with 0.15M or 0.3M of nitrite.
The levels of THCCOOH at various time intervals after nitrite spiking were monitored
by instrument-based cannabinoids immunoassays (Syva EMIT d.a.u. and/or Roche
Abuscreen ONLINE assays) and by an onsite THC immunoassay (Roche ONTRAK TESTSTIK).
Results from this report demonstrate that the two outstanding “urine specimen
factors” that dictated the effectiveness of the nitrite adulteration were
urinary pH and the original drug concentration before nitrite spiking. Significant
decreases in the immunoassay results could be observed within 4 h of nitrite
treatment in the majority of samples with acidic urinary pH values. Regardless
of their original concentration of THCCOOH (GC–MS ranging from 33 to 488
ng/mL), all of the 20 samples that had acidic pH values gave negative immunoassay
results 1 day after nitrite adulteration. In contrast, the immunoassay results
of samples with neutral or basic pH values were less affected by nitrite exposure
in the same studies. Approximately two-thirds of the samples with pH values
greater than 7.0 remained immunoassay-positive 3 days after nitrite spiking.
Nevertheless, some of the adulterated urine that showed no change in immunoassay
results might exhibit significant decrease in GC–MS recoveries even with
bisulfite treatment, collaborating with the observations that a portion of samples
screened positive with THC immunoassay in the laboratory could fail to confirm
with GC–MS analysis. The decrease or loss of immunoassay detectable cannabinoid
cross-reactives in acidic “THC-positive samples” can be attenuated
by chemically increasing the pH value of the samples to the basic pH range. Reproduction
of editorial content of this journal is prohibited without publisher’s
permission.
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