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Column-switching technique for the sensitive determination of ertapenem in human cerebrospinal fluid using liquid chromatography and ultraviolet absorbance detection
Ist Teil von
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2004-01, Vol.799 (1), p.9-14
Ort / Verlag
Amsterdam: Elsevier B.V
Erscheinungsjahr
2004
Quelle
Elsevier ScienceDirect Journals
Beschreibungen/Notizen
A sensitive reversed-phase high-performance liquid chromatographic (RP-HPLC) assay with on-line extraction was developed for quantifying ertapenem in human cerebrospinal fluid (CSF). This assay is at least five times more sensitive than previously published ertapenem methods with a lower limit of quantitation at 0.025
μg/ml. In this assay, a CSF sample is extracted on-line using a RP extraction column and an aqueous acidic mobile phase (0.1% formic acid) to wash away polar endogenous materials, while ertapenem is retained on the column. Ertapenem is then back-flushed off the extraction column and directed to a RP analytical column using an acidic mobile phase with an organic modifier (acetonitrile/0.1% formic acid, 15:85 (v/v)) and detected using UV absorbance. The acidic mobile phase provided a sharper chromatographic peak and on-line extraction allowed large injection volumes (≥150
μl) of buffered CSF to be injected without compromising column integrity. These assay conditions were necessary to quantify ertapenem at levels expected to be found in human CSF (<0.05
μg/ml). The method was successfully validated and implemented for a clinical study: intraday precision and accuracy of the CSF assay for calibration standards (0.025–10
μg/ml) and quality control samples (0.1, 0.5, and 2.5
μg/ml) were <6.2% coefficient of variation and 96.8–104.0% of nominal concentration, respectively.