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M. Saleh, F. Jehl, A. Sauer, G. Prevost, C. Speeg-Schatz, T. Bourcier; High-Performance Liquid Chromatographic Assay for Linezolid in Serum and Vitreous Humor. Invest. Ophthalmol. Vis. Sci. 2008;49(13):2503. doi: https://doi.org/.
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Linezolid represents the first new class (oxazolidinone) of antimicrobial agent to be introduced into infectious disease practice in over two decades. It can be administered orally with 100% bioavailability and produces peak serum levels of approximately 18 mg/L after repeated doses of 600 mg. Thus, linezolid could be a promising drug in the management of bacterial endophthalmitis. As further pharmacokinetic investigations are needed, we developed an assay for quantifying linezolid in vitreous humor.
An aliquot of either serum or vitreous humor (0.5 mL) was mixed with an equal volume of acetonitrile and centrifuged 10 min at 3000 g. 20µL of the supernatant was injected into the column. Chromatography was performed on a high speed analytical column (150*4.6 mm I.D.) packed with 5µm particles (XL-ODS, Beckmann). The mobile phase consisted of 25 mmol/L ammonium acetate -acetonitrile (76:24, v/v) adjusted to pH 5 with glacial acetic acid. The flow-rate was set at 1.0mL/min and the eluent was monitored at 251 nm.
The quantification limit was 0.1 mg/L in vitreous humor. The linearity study was carried out with concentrations ranging from 0.2 to 20 mg/L. The regression analysis (SigmaPlot) revealed that the method is linear in both serum and vitreous (r=0.99, r=0.97). The extraction recovery from serum and vitreous spiked with linezolid was close to 100%. Concerning the intra-assay precision in serum, coefficients of variation reached 6.08 % (0.5mg/L), 7.73% (5mg/L) and 7.59% (20mg/L). The values for inter-assay precision respectively were 4.78 %, 7.91% and 7.92%. Accuracy for targeted values were below 15%. At last, the assay stability showed that linezolid was steady in vitreous for at least 4 weeks at 4°C and 4 weeks at -80°C.
HPLC methods for linezolid were developed in a variety of matrices (plasma, serum, urine, microdialysate) but none of them refers to eye tissues. Herein we developed the first accurate, rapid, and sensitive HPLC method for the measurement of linezolid in vitreous humor.
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