title: Determination of colchicine and O-demethylated metabolites in decomposed skeletal tissues by microwave assisted extraction, microplate solid phase extraction and ultra-high performance liquid chromatography (MAE-MPSPE-UHPLC).
abstract: Microwave assisted extraction (MAE) followed by microplate solid phase extraction (MPSPE) coupled with ultra high performance liquid chromatography (UHPLC) for the semi-quantitative determination of colchicine, 3-demethyl colchicine and 2-demethyl colchicine in postmortem rat bone is described. Rats (n=4) received 50mg/kg colchicine (i.p), and euthanized by CO2 asphyxiation. Remains decomposed to skeleton outdoors and vertebral bones were collected cleaned, and ground to a fine powder. Powdered bone underwent MAE using methanol in a closed microwave system, followed by MPSPE and analysis using UHPLC-PDA. MAE analyte stability was assessed and found to be stable for at least 60 min irradiation time. The majority (>95%) of each analyte was recovered after 15 min. The MPSPE-UHPLC method was linear between 10 and 2,000 ng/mL, with coefficients of variation <20% in triplicate analysis, with a limit of detection of 10 ng/mL for each of the three analytes. Following MAE for 30 min (80°C, 1200W), MPSPE-UHPLC analysis of vertebral bone of colchicine-exposed rats detected colchicine (1.8-4.1 μg/g), 3-demethyl colchicine (0.77-1.8 μg/g) and 2-demethyl colchicine (0.43-0.80 μg/g) in all samples assayed.
authors.affiliation: Department of Forensic Science, Laurentian University, 935 Ramsey Lake Rd, Sudbury, ON, Canada P3E 2C6. Electronic address: jwatterson@laurentian.ca.
authors.forename: J H
authors.initials: JH
authors.lastname: Watterson
authors.affiliation: Department of Forensic Science, Laurentian University, 935 Ramsey Lake Rd, Sudbury, ON, Canada P3E 2C6.
authors.forename: A B
authors.initials: AB
authors.lastname: Imfeld
authors.affiliation: Department of Forensic Science, Laurentian University, 935 Ramsey Lake Rd, Sudbury, ON, Canada P3E 2C6.
authors.forename: H C
authors.initials: HC
authors.lastname: Cornthwaite
mesh_terms: D000818:Animals; D001842:Bone and Bones / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003078:Colchicine / Q000031:analogs & derivatives* / Q000032:analysis* / Q000737:chemistry / Q000493:pharmacokinetics; D008872:Microwaves; D051381:Rats; D017208:Rats, Wistar; D052616:Solid Phase Extraction / Q000379:methods*
chemical_list: D003078:Colchicine
keywords: Bone; Colchicine; Forensic toxicology; Metabolites; Microwave assisted extraction; UHPLC