HPLC-MS-MS Determination of ZCZ-011, A Novel Pharmacological Tool for Investigation of the Cannabinoid Receptor in Mouse Brain Using Clean Screen FASt™ Column Extraction

Justin L Poklis, Deborah J Clay, Bogna M Ignatowska-Jankowska, Chiara Zanato, Ruth A Ross, Iain R Greig, Rehab A Abdullah, Mohammed A Mustafa, Aron H Lichtman, Alphonse Poklis

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A high-performance liquid chromatography tandem mass spectrometry method was developed for the detection and quantification of 6-methyl-3-(2-nitro-1-(thiophen-2-yl)propyl)-2-phenyl-1H-indole (ZCZ-011) using 2-phenylindole as the internal standard (ISTD). ZCZ-011 was synthesized as a possible positive allosteric modulator with the CB1 cannabinoid receptor. The analytical method employs a rapid extraction technique using Clean Screen FASt™ columns with a Positive Pressure Manifold. FASt™ columns were originally developed for urine drug analysis but we have successfully adapted them to the extraction of brain tissue. Chromatographic separation was performed on a Restek Allure Biphenyl 5 µ, 100 × 3.2 mm column (Bellefonte, PA). The mobile phase consisted of 1:9 deionized water with 10 mmol ammonium acetate and 0.1% formic acid-methanol. The following transition ions (m/z) were monitored for ZCZ-011: 363 > 207 and 363 > 110 and for the ISTD: 194 > 165 and 194 > 89. The FASt™ columns lowered and stabilized the ion suppression over the linear range of the assay (40-4,000 ng/g). The method was evaluated for recovery, ion suppression, accuracy/bias, intraday and interday precision, bench-top stability, freeze-thaw and post-preparative stability. The method was successfully applied to brain tissue from C57BL/6J mice that received intraperitoneal (i.p.) injections with 40 mg/kg of ZCZ-011 or vehicle.

Original languageEnglish
Pages (from-to)353-359
Number of pages7
JournalJournal of analytical toxicology
Issue number5
Early online date3 Mar 2015
Publication statusPublished - 1 Jun 2015


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