Role of liquid chromatography–high-resolution mass spectrometry (LC-HR/MS) in clinical toxicology

Abstract

Background. Gas chromatography (GC) and liquid chromatography (LC) coupled with mass spectrometry (MS) are widely used to confirm drug screening results and for urine screening in presumed intoxicated patients. These techniques are better suited to targeted analysis than to general unknown screening and, due to the complexity of testing, results are seldom available rapidly enough to contribute to the immediate care of the patient. High resolution (HR)/MS with time-of-flight (TOF) or orbitrap instruments offer potential advantages in clinical toxicology. Comparison of GC-MS, LC-MS/MS and LC-HR/MS. For unknown analyses, GC-MS and LC-MS/MS require comparison of full-scan spectra against preestablished libraries. Operation in full-scan mode greatly reduces sensitivity and some drugs present in low but significant concentrations may be missed. Selected ion monitoring (SIM) in GC/MS and selected reaction monitoring (SRM) in LC-MS/MS, where only targeted ions are monitored, increase sensitivity but require prior knowledge of what compound is to be measured. LC-HR/MS offers mass assignment with an accuracy of 0.001 atomic mass units (amu) compared with 1 amu in conventional MS. Tentative identification is thus directed to a very limited set of compounds (or even one unique compound) based on the exact molecular formula rather than a fragmentation pattern, since HR/MS can discriminate between compounds with the same nominal molecular mass. LC-MS/MS has clear advantages over GC/MS in ease and speed of sample preparation and the opportunities for its automation. LC-HR/MS is more suitable to clinical toxicology because the drugs present in a sample are rarely known a priori, and tentative identifications of unknowns can be made without the availability of a reference standard or a library spectrum. Blood can be used in preference to urine which is more relevant to the patient's current clinical situation. Methods. A literature search was conducted using PUBMED for clinical toxicology, adulterants in illicit drugs and herbal supplements, and case reports using LC-TOF/MS and LC-HR/MS. Only 42 papers in English were identified in these searches. LC-HR/MS in clinical toxicology. LC-HR/MS has been used to detect designer drugs, doping agents, (neurosteroids) and adulterants such as levamisole, a veterinary antihelmitic found in street cocaine, and pharmaceuticals in herbal medications marketed to contain only natural ingredients. LC-HR/MS has proved useful for cases where existing tests were unable to identify the cause of the intoxication. One patient suffered a drug-induced seizure which was originally thought to be caused by an herbal medication, but diphenhydramine was determined to be the culprit. In another, 5-oxoproline was identified as the cause of metabolic acidosis seen in chronic acetaminophen (paracetamol) use. LC-HR/MS has successfully identified medications that were mislabeled or misrepresented street drugs. In one case, medications sold as diazepam were determined to be glyburide instead. The identification of novel designer amines, stimulants found in “bath salts”, and synthetic cannabinoids are well suited to LC-HR/MS. Dozens or even hundreds of possible compounds cannot realistically be tested on an individual basis by targeted LC-MS/MS or GC/MS analysis. Conclusions. LC-HR/MS offers unique opportunities for time-sensitive clinical analysis of blood samples from intoxicated patients and for comprehensive screening in a wide range of situations and materials. While the identification is not as definitive as that obtained by conventional fragmentation MS, the presumptive identification can be confirmed later with standards and spectral library matches. Optimum utilization of the presumptive diagnosis requires close collaboration between the laboratory analysts and their clinical counterparts.