Clin Chem Lab Med 2010;48(3):391–398  2010 by Walter de Gruyter • Berlin • New York. DOI 10.1515/CCLM.2010.078 2010/359 Article in press - uncorrected proof Development and validation of a high performance liquid chromatography method to determine linezolid concentrations in pig pulmonary tissue Laura Guerrero 1 , Pilar Martı ´nez-Olondris 2 , Montserrat Rigol 3 , Mariano Esperatti 2 , Cristina Esquinas 2 , Ne ´stor Luque 2 , Raquel Pin ˜er 2 , Antoni Torres 2 and Dolors Soy 1, * 1 Pharmacy Service, Hospital Clı ´nic de Barcelona, Institut d’Investigacions Biome `diques August Pi i Sunyer (IDIBAPS), Universidad de Barcelona, Spain 2 Pneumology Service, Institut Clı ´nic del To `rax, Hospital Clı ´nic de Barcelona, Institut d’Investigacions Biome `diques August Pi i Sunyer (IDIBAPS), Universidad de Barcelona, Spain 3 Cardiology Service, Institut Clı ´nic del To `rax, Hospital Clı ´nic de Barcelona, Institut d’Investigacions Biome `diques August Pi i Sunyer (IDIBAPS), Universidad de Barcelona, Spain Abstract Background: Linezolid is the first synthetic compound of a new group of antimicrobials, the oxazolidinones, which inhibit protein synthesis. It shows a broad spectrum of activ- ity against Gram positive organisms. With respect to its phar- macokinetics, linezolid shows a relatively high volume of distribution and good penetration into inflammatory fluids, bone, fat and muscle. Methods: A reversed-phase isocratic high-performance liq- uid chromatographic method for linezolid analysis in piglet pulmonary tissue is described. Tissue samples and controls were prepared in 1=TBE (1 M Tris, 0.9 M boric acid, 0.01 M EDTA). The mobile phase consisted of 20% ultrafiltered water and 80% of (A) 15 mM potassium monohydrogen phosphate buffer (pHs5) with (B) acetonitrile (80%/20%; v/v). Samples were homogenized and precipitated with HClO 4 3% (1/1, v/v). The injection volume was 100 mL. Ofloxacin was used as an internal standard. Results: The assay was linear over a linezolid concentration range: 1.6–100 mg/mL. The method provided good valida- tion data (ns15): inaccuracy (3.6%), intra and inter-day var- iability (4.2% and 5.2%, respectively), recovery (91.8%), limit of detection (0.8 mg/mL) and quantitation (1.6 mg/mL) and acceptable stability within 24 h in the auto-sampler. *Corresponding author: Dolors Soy, PharmD, PhD, Pharmacy Service, Hospital Clı ´nic de Barcelona, C/Villarroel, 170 – 08036, Barcelona, Spain Phone: q34 93 2275479, Fax: q34 93 2275457, E-mail: dsoy@clinic.ub.es Received July 13, 2009; accepted November 11, 2009; previously published online February 1, 2010 Conclusions: The method offers a fast and simple approach to determine linezolid in pulmonary tissue which could be of use in pharmacokinetic studies. Clin Chem Lab Med 2010;48:391–8. Keywords: analytics; chromatography; high pressure liquid; linezolid; tissue; validation. Introduction Linezolid (LNZ) (1) is the first synthetic compound of a new group of antimicrobial drugs, the oxazolidinones (Figure 1) which inhibit protein synthesis early in translation. It shows a broad spectrum of activity against Gram positive organ- isms including methicillin resistant Staphylococcus aureus (MRSA), penicillin resistant pneumococci and vancomycin resistant Enterococcus faecalis and Enterococcus faecium. After a dose of 600 mg twice a day, the maximum serum concentration is ;20 mg/L. With respect to its pharmaco- kinetics (PK), linezolid shows a relatively high volume of distribution (40–50 L) and good penetration into inflamma- tory fluids, bone, fat and muscle (2). It is well known that antibiotic concentrations at the site of the infection differ greatly from those in plasma. Drug penetration varies depending on the drug, the tissue involved and the infection. Also, protein binding, physical-chemical drug properties, lipid solubility, etc. have some influence on the amount of drug that is able to reach peripheral tissue/ compartments. Data on drug penetration for the latest antimicrobial drugs, such as ertapenem, tigecycline, teli- thromycin or linezolid have recently been published (3–6). Thus, in localized infections including lung infection, men- ingitis, endocarditis or osteomyelitis, it is extremely valuable to know what fraction of the free drug will be able to cross membranes and barriers and reach the site of the infection. Hence, many reports in the literature have been performed on drug penetration into different tissues. Examples include drug penetration into the epithelial lining fluid (ELF) (7–10) and lung tissue (11). High performance liquid chromatography (HPLC) can be considered an adequate technique (12) for determining line- zolid in pulmonary tissue. The sensitivity and precision of HPLC and its applicability to a wide variety of compounds has led to its use in clinical laboratories for monitoring of a variety of therapeutic agents in hospital settings, and phar- macokinetic and metabolism studies (13, 14). Brought to you by | Universitat de Barcelona Authenticated Download Date | 4/27/18 7:06 PM