Journal of Chromatography B, 799 (2004) 337–341 Short communication Validation of a simplified method for determination of cimetidine in human plasma and urine by liquid chromatography with ultraviolet detection Tahira Iqbal, Chetan S. Karyekar, Minori Kinjo, Gilbert C. Ngan, Thomas C. Dowling ∗ Renal Clinical Pharmacology Lab, Department of Pharmacy Practice and Science, University of Maryland, School of Pharmacy, 100 N. Penn St., AHB Rm. 540-D, Baltimore, MD 21201, USA Received 24 June 2003; received in revised form 14 October 2003; accepted 20 October 2003 Abstract A HPLC method was developed for determination of cimetidine in human plasma and urine. Plasma samples were alkalinized followed by liquid extraction with water-saturated ethyl acetate then evaporated under nitrogen. The extracts were reconstituted in mobile phase and injected onto a C 18 reversed-phase column; UV detection was set at 228 nm. Urine samples were diluted with an internal standard/mobile phase mixture (1:9) prior to injection. The lower limit of quantification in plasma and urine were 100 ng/ml and 10 g/ml, respectively; intra- and inter-day coefficients of variation were ≤4.2%. Advantages of this validated assay include a readily available internal standard, simplified plasma extraction and urine dilution methods, and applicability to clinical studies investigating the renal handling of cimetidine. © 2003 Elsevier B.V. All rights reserved. Keyword: Cimetidine 1. Introduction Cimetidine (N ′′ -cyano-N-methyl-N ′ -[2-[[(5-methyl-1H- imidazol-4-yl) methyl]thio]ethyl]-guanidine) is a histamine H(2)-receptor antagonists that is used widely to treat gastric and duodenal ulcers [1,2]. Cimetidine is excreted predom- inantly unchanged by the kidneys (approximately 70%) and undergoes extensive tubular secretion with renal clear- ance values approximately four-fold greater than creatinine clearance [3,4]. Cimetidine has also been identified as a substrate for P-glycoprotein (P-GP), an MDR-encoded membrane transporter that is expressed in normal tissues including kidney proximal tubules [5–7]. Thus, evaluation of renal P-GP probe compounds, such as cimetidine, is critical in order to identify potential renal drug interac- tions, prevent drug toxicity, and optimize drug therapy in patients. Several HPLC methods for the determination of cime- tidine in human plasma and urine have been reported. ∗ Corresponding author. Tel.: +1-410-706-6590; fax: +1-410-706-6580. E-mail address: tdowling@rx.umaryland.edu (T.C. Dowling). Most methods utilized either solid-phase extraction or liquid-phase extraction techniques. Limitations of these methods include the requirement to extract large volumes of plasma (0.5–1.0 ml) [8–10], low or inconsistent recov- ery in plasma [11,12], use of an internal standard that is either not commercially available [10,13] requires an addi- tional protonation step using hydrochloric acid [14], lack of urine analysis capabilities [12,14,15] or requirement of solid-phase or liquid-liquid extraction methods for urine analysis [1,8,16,17]. This report describes a validated HPLC method for determining cimetidine concentrations in plasma that incorporates a simplified procedure for urine sample analysis. 2. Experimental methods 2.1. Reagents and chemicals Cimetidine, famotidine, heptanesulfonic acid, sodium acetate and sodium carbonate were purchased from Sigma (St. Louis, MO, USA). The purity of cimetidine and famo- tidine standards was ≥99.0%. HPLC grade acetonitrile, 1570-0232/$ – see front matter © 2003 Elsevier B.V. All rights reserved. doi:10.1016/j.jchromb.2003.10.030