Influence of dose-death interval on colchicine and metabolite distribution in decomposed skeletal tissues

Abstract

The semi-quantitative analysis of decomposed bone of rats exposed to colchicine and euthanized following different time intervals postexposure (i.e., dose-death interval, DDI) is described. Rats received colchicine (50 mg/kg, i.p.) and were euthanized 30 min (DDI1; n = 4), 60 min (DDI2; n = 4), or 180 min (DDI3; n = 4) postdose. Drug-free animals (n = 3) served as negative controls. Perimortem heart plasma was collected. Remains were decomposed to skeleton outdoors and then collected and sorted (skull, vertebrae, rib, pelvis, femur, tibia). Bones were dried, pulverized, and prepared by microwave-assisted extraction and microplate solid-phase extraction (MAE-MPSPE), followed by analysis for colchicine, 3-demethylcolchicine (3DMC), and 2-demethylcolchicine (2DMC) by ultra-high-performance liquid chromatography with photodiode array detection (UHPLC-PDA) at 350 nm. Bone type was a main effect (Kruskall-Wallis, p < 0.05) with respect to drug level (expressed as mass-normalized response ratio, RR/m) for each analyte, at each DDI. For all samples, DDI was a main effect (Kruskall-Wallis, p < 0.05) with respect to analyte level, and the ratio of analyte levels (RR_3DMC/RR_COLCH, RR_2DMC/RR_COLCH, and RR_2DMC/RR_3DMC). Bone COLCH levels varied by 19-fold, 12-fold, and 60-fold across all bone types in the DDI1, DDI2, and DDI3 groups, respectively. Bone 3DMC levels varied by 12-fold, 11-fold and 17-fold across all bone types in the DDI1, DDI2, and DDI3 groups, respectively. Bone 2DMC levels varied by 20-fold, 14-fold, and 14-fold across all bone types in the DDI1, DDI2, and DDI3 groups, respectively. Values of RR_3DMC/RR_COLCH varied by 16-fold, 5-fold, and 5-fold across all bone types in the DDI1, DDI2, and DDI3 groups, respectively. Values of RR_2DMC/RR_COLCH varied by 10-fold, 6-fold, and 12-fold across all bone types in the DDI1, DDI2, and DDI3 groups, respectively. Values of RR_2DMC/RR_3DMC varied by 3-fold, 5-fold, and 2-fold across all bone types in the DDI1, DDI2, and DDI3 groups, respectively. Measured analyte levels in bone correlated poorly with corresponding levels in blood (r = −0.65–+0.31). Measured values of RR_2DMC/RR_COLCH and RR_2DMC/RR_3DMC in bone also correlated poorly with corresponding values in blood. Measured values of RR_3DMC/RR_COLCH were well correlated with corresponding blood levels for all bone types except skull (r = 0.91–0.97).