Author ORCID Identifier



Date of Graduation


Document Type


Degree Type



Eberly College of Arts and Sciences


Forensic and Investigative Science

Committee Chair

Glen P. Jackson

Committee Co-Chair

Luis Arroyo

Committee Member

Patrick C. Callery


Designer drugs and novel psychoactive substances (NPSs) are continuously entering the black market and causing serious health problems among users. NPSs present an especially difficult challenge to forensic chemists and toxicologists because they are not well characterized and often involve families of structurally related analogs. In crime laboratories, mass spectrometry is commonly used as a part of an analytical scheme to characterize and identify unknown analytes. Towards this end, tandem mass spectra of questioned analytes are often compared to reference spectra of standards that are either collected contemporaneously with the analyte or collected previously and stored in a database. However, new designer drugs are often not present in reference libraries and standards are often not available. Therefore, the ability to interpret fragment ion spectra and extrapolate understanding to different structures is an important supporting endeavor of forensic science.

In this work, multistage mass spectrometry (MSn) and isotope labeling enabled the interpretation of the fragmentation behavior of the synthetic cannabinoid 5F-APP-PINACA, also known as PX-2. Although reference mass spectra for PX-2 are available through databases such as NIST, the identities of specific peaks and how these peaks are formed are not widely known. Previous work on the tandem mass spectrometry of PX-2 suggested multiple explanations for the fragment ion at m/z 251, including the addition of water via an ion-neutral reaction and phenyl or hydroxyl group transfers to the nitrogen in the 2-position of the indazole core.

The major finding of the fragmentation behavior of PX-2 is that unintentional ion-molecule reactions can occur within certain ion trapping mass spectrometers that cause the addition of water (18 Da) to certain major fragment ions. In some cases, the adducts can become dominant in the product ion spectra and therefore confound spectral interpretation. The enhanced reactivity for indazole-containing NPSs like PX-2 seems to stem from the production of a ketene (R-C=C=O) functional group on the indazole ring. PX-1, which is structurally very similar to PX-2, contains an indole group instead of the indazole group and does not undergo such adduction with residual water. In a positive light, the adduction of residual water could be used in future studies to help identify ketene-like intermediates on indazole cores of PINACA-type NPSs. However, in terms of immediate impact on the criminal justice system, analysts should be aware that indazole-containing NPSs are prone to reactions with water during tandem mass analysis, and abundant product ions can be formed that contain atoms (H and O) that were not present in the isolated precursor ions. If the pressures of residual gases like water and methanol are not monitored or controlled, they could dramatically influence the relative ion abundances of adduct ions and therefore affect the reliability of tandem mass analysis of indazole-containing NPSs such as the PINACA, FUBINACA and CHMINACA analogs.

Embargo Reason

Publication Pending

Available for download on Saturday, July 27, 2024