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  • Title: Development of a UHPLC-MS/MS method for the measurement of chlortetracycline degradation in swine manure.
    Author: Shelver WL, Varel VH.
    Journal: Anal Bioanal Chem; 2012 Feb; 402(5):1931-9. PubMed ID: 22207281.
    Abstract:
    An ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed capable of simultaneously measuring chlortetracycline (CTC), epi-chlortetracycline, and isochlortetracycline (ICTC), as well as other structurally related tetracyclines in swine manure. A simple sample preparation was used consisting of extraction, dilution, centrifugation, and ultrafiltration. The concentrations of analyte were calculated using d(6)-tetracycline as an internal standard in the matrix-matched standard curve. A solvent gradient resolved the compounds in 3.5 min with an additional 1.5 min of re-equilibration allowing the analyses of a large number of samples in a short period of time. MS/MS was used as the detection method giving analyte confirmation in addition to a large dynamic range and low detection limit. The UHPLC-MS/MS method successfully resolved multiple degradation products of CTC from the complex manure matrix. The method detection limits ranged from 1.9 pg/μL for CTC to 7.3 pg/μL for ICTC, and the calibration curve was linear from 1 to 10,000 pg/μL. The method was tested by measuring CTC and its degradation products as a function of time in incurred swine manure that had been incubated at three different temperatures (22 °C, 38 °C, and 55 °C). CTC concentration at 22 °C decreased 44% after 25 days; greater percentage decreases were observed when the manure was stored at elevated temperatures (96% and 98% for 38 °C and 55 °C, respectively). The concentration of the microbiologically inactivate isomer, ICTC, increased over the incubation period. At 22 °C, ICTC continued to increase through 25 days of incubation; at 38 °C, ICTC concentration plateaued on day 14 while at 55 °C ICTC concentration plateaued on day 7, with concentration increases of 198%, 374%, and 282% for 22 °C, 38 °C, and 55 °C, respectively.
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