Available online at www.sciencedirect.com Journal of Pharmaceutical and Biomedical Analysis 46 (2008) 723–727 Simultaneous determination of rifampicin and levofloxacin concentrations in catheter segments from a mouse model of a device-related infection by liquid chromatography/electrospray ionization tandem mass spectrometry Donghui Bao a,∗ , Thanh-Thai Truong a , Paul J. Renick a , Mark E. Pulse b , William J. Weiss a a Cumbre Pharmaceuticals Inc., 1502 Viceroy Drive, Dallas, TX 75235, USA b UNT Health Science Center, 3500 Camp Bowie Boulevard, Fort Worth, TX 76107, USA Received 20 August 2007; received in revised form 4 November 2007; accepted 20 November 2007 Available online 26 November 2007 Abstract The aim of this study was to develop a specific and sensitive liquid chromatography mass spectrometry (LC/MS) method for the determination of rifampicin and levofloxacin concentrations from infected tissues within teflon catheter segments which were subcutaneously implanted in mice. A solid-phase extraction procedure was used to extract analytes from tissue homogenates of the catheter segments and reverse-phase HPLC combined with positive electrospray ionization mass spectrometry was used for analyte separation and quantification. The assay was found to be linear over the concentration range of 0.02–2 g/g for rifampicin and levofloxacin in tissues and provided good validation data for accuracy and precision. The intra-day accuracy as determined by the relative error was -1.3% for levofloxacin and 6.1% for rifampicin, and precision was evaluated by R.S.D.s with a maximum of 5.1% for levofloxacin and 8.1% for rifampicin. The inter-day accuracy was -3.3% for levofloxacin and -4.6% for rifampicin, and precision was 8.6% for levofloxacin and 7.1% for rifampicin. The assay uses less tissue than previously described methods and can be applied to determine the penetration of rifampicin and the fluoroquinolone in catheter segments from a mouse model of a device-related infection. Finally, the HPLC–MS assay should be applicable to studies of rifamycin + quinolone combination therapies in other animal models of bacterial infection. © 2007 Elsevier B.V. All rights reserved. Keywords: Levofloxacin; Rifampicin; LC/MS; Device-related infections; Catheter 1. Introduction Rifampicin is a semi-synthetic rifamycin analog that is active against gram-positive bacteria and some gram-negative bacteria by inhibition of bacterial RNA polymerase [1–4]. It possesses excellent activity against slow-growing bacteria and penetrates into structured bacterial communities on the sur- faces of implanted devices known as biofilms [5–8]. Clinically, rifampicin is used mainly in antibiotic combination thera- pies, because high-level resistance develops rapidly when the agent is administered in monotherapy [9]. Fluoroquinolones have been frequently used in combination with rifampicin for the treatment of device-related infections [10], and effi- ∗ Corresponding author. Current address: 303-A College Road East, Princeton, NJ 08540, USA. Tel.: +1 609 613 4100; fax: +1 609 613 4150. E-mail address: dbao@pharmasset.com (D. Bao). cacy has been demonstrated in some clinical studies [11,12]. Levofloxacin is a second-generation fluoroquinolone that has improved antimicrobial and pharmacokinetic properties. The plasma elimination half-life for levofloxacin ranges from 6 to 8 h, which is complementary to the 3–5 h half-life for rifampicin, making this combination promising for the treatment of infec- tions of indwelling medical devices. In order to correlate efficacy with the pharmacokinetics of rifampicin and fluoroquinolones in an animal model, a sensitive and selective assay for the determination of drug concentra- tions is required, preferably within the same assay. Traditional approaches to monitor plasma drug concentrations are less predictive for device-related infections, where the infections are mainly within extravascular tissue sites. Measuring the concentrations of antimicrobial agents at the site(s) of infec- tion is an important parameter in the systematic evaluation of anti-infective agents for the treatment of device-related infections. There are many assay methods for rifampicin and 0731-7085/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.jpba.2007.11.023