More on Interference of N-Acetylcysteine in Measurement of Acetaminophen

Dept. of Clin. Biochem., Hadassah Hosp., Hebrew Univ.-Hadassah Med. School, P. O. Box 12000, Jerusalem, IL-91120 Israel
^a Author for correspondence. Fax 972-2-6435778; e-mail mayer{at}hadassah.org.il.

To the Editor:

Acetaminophen (ACET) is a common analgesic and antipyretic drug that in excessive concentrations may cause acute liver damage and acute renal failure (1). Although the normal elimination of ACET seems to involve conjugation of a toxic alkylating metabolite with liver glutathione to yield a detoxified conjugation product, excessive doses of ACET deplete liver glutathione stores, causing accumulation of the toxic metabolite N-acetyl-imidoquinone (1)(2)(3). Acetylcysteine (NAC, N-acetyl-3-mercaptoalanine) is the drug of choice for the treatment of an ACET overdose. Although the mechanism responsible for the ability of NAC to serve as an effective antidote in vivo has not been fully elucidated, a major route of detoxification seems to depend on the ability of NAC to serve as a potent sulfhydryl donor that restores depleted hepatic reduced glutathione (3).

A 28-year-old woman was recently admitted to the emergency room of our hospital after a confirmed (suicidal) ACET overdose. One hour after treatment with NAC was initiated (150 mg/kg of body weight, infused i.v. for 15 min), a blood sample was withdrawn for ACET assay in our laboratory. We routinely assay ACET by an enzymatic assay (GDS enzymatic acetaminophen, GDS Diagnostics) based on the hydrolysis of ACET by acylamidase to yield p-aminophenol and acetate. The p-aminophenol is measured colorimetrically by its conversion to indophenol in the presence of o-cresol, using periodate as a catalyst. Unexpectedly, no ACET was found in the serum despite the patient's ingestion of a toxic dose of the drug. A similar enzymatic assay (acetaminophen assay, cat. no. 503–10, Diagnostic Chemical) that uses acyl amidohydrolase to cleave the amide bond in ACET but in which the formed p-aminophenol reacts with 8-hydroxyquinoline in the presence of manganese ions to form the colored compound 5-(4-iminophenol)-8-quinolone, also failed to detect ACET in the sample.

In contrast to the two enzyme-based assays that failed to demonstrate ACET intoxication, the TDx assay (Abbott Laboratories) confirmed high, toxic concentrations of the drug in the same blood sample. The TDx assay is based on the competition between ACET in the sample and tracer-labeled ACET for a specific antibody and measures the change in the polarization of fluorescent radiation emitted by the fluorescein-labeled tracer.

These conflicting results and a recent report on false negative results for urinary ACET screening in the presence of NAC (4) raised the possibility that NAC interferes with the enzymatic assay of ACET. We studied this possibility by performing the enzymatic assay on controls containing increasing concentrations of ACET in the presence of different therapeutic concentrations of NAC (Fig. 1 A). Evidently, NAC concentrations 100 mg/L cause a marked negative bias, reaching total suppression of the reaction in the presence of 10 g/L NAC. In contrast, similar concentrations of NAC have no effect on the ACET TDx immunoassay (Fig. 1B ). According to the TDx method manual, concentrations of up to 1000 mg/L NAC cause only a minimal bias of <1 mg/L ACET in this assay.

View larger version (15K): [in this window] [in a new window]   Figure 1. Comparison of enzymatic and TDx assays for ACET determination. (A) Enzymatic assays; (B) TDx assay.

Plasma concentrations of NAC after administration of the common initial dose of 150 mg/kg of body weight over a period of 15 min have been reported to range from 304 to 875 mg/L (5). NAC concentrations in this range, and certainly the higher concentrations of blood NAC reached after larger dosages during infusion, markedly impede ACET determination by the enzymatic assay (Fig. 1A ).

That the drug of choice for treating ACET intoxication produces a very large negative bias in a commonly used enzymatic-colorimetric assay of ACET is worth noting. On the basis of the present observation, we recommend that patients' blood should be sampled for ACET before the initiation of NAC treatment. Alternatively, enzyme-based colorimetric methods should be replaced by immunoassays or HPLC assays of ACET in which NAC causes no interference.

References

1. Rumack BH, Peterson RG. Acetaminophen overdose: incidence, diagnosis, and management in 416 patients. Pediatrics 1978;62:898-903. [Abstract/Free Full Text]
2. Forrest JAH, Clements JA, Prescott LF. Clinical pharmacokinetics of paracetamol. Clin Pharmacokinet 1982;7:93-107. [Web of Science][Medline] [Order article via Infotrieve]
3. Lauterburg BH, Corcoran GB, Mitchell JR. Mechanism of action of N-acetylcysteine in the protection against the hepatotoxicity of acetaminophen in rats in vivo. J Clin Invest 1983;71:980-991.
4. Davey L, Naidoo D. Urinary screen for acetaminophen (paracetamol) in the presence of N-acetylcysteine. Clin Chem 1993;39:2348-2349. [Medline] [Order article via Infotrieve]
5. Prescott LF, Donovan JW, Jarvie DR, Proudfoot AT. The disposition and kinetics of intravenous N-acetylcysteine in patients with paracetamol overdosage. Eur J Clin Pharmacol 1989;37:501-506. [Web of Science][Medline] [Order article via Infotrieve]