Analytica Chimica Acta 569 (2006) 250–259 Simultaneous determination of flufenamic and meclofenamic acids in human urine samples by second-order multivariate parallel factor analysis (PARAFAC) calibration of micellar-enhanced excitation–emission fluorescence data A. Mu ˜ noz de la Pe ˜ na a,∗ , N. Mora D´ ıez a , D. Bohoyo Gil a , A.C. Olivieri b , G.M. Escandar b a Departamento de Qu´ ımica Anal´ ıtica, Facultad de Ciencias, Universidad de Extremadura, 06071 Badajoz, Spain b Departamento de Qu´ ımica Anal´ ıtica, Facultad de Ciencias Bioqu´ ımicas y Farmac´ euticas, Universidad Nacional de Rosario, Suipacha 531, Rosario S2002LRK, Argentina Received 7 November 2005; received in revised form 14 February 2006; accepted 17 March 2006 Available online 29 March 2006 Abstract A new spectrofluorimetric method has been developed for the quantitative determination of the anti-inflammatory flufenamic and meclofenamic acids in urine samples. The method is based on second-order multivariate calibration applying parallel factor analysis (PARAFAC). The fluorescence excitation–emission matrices (EEMs) of these compounds are highly overlapped, precluding analyte direct determination without performing a previous separation procedure. The use of a micellar medium allowed for both fluorescence enhancement and analyte discrimination from the biological background. The calibration solutions were prepared in water, with concentrations in the ranges from 0.15 to 4.00 g ml -1 for flufenamic acid and from 0.25 to 6.00 g ml -1 for meclofenamic acid. The use of the second-order calibration method, in the standard addition mode, was required to avoid internal filter interference effects from the urine background. The measured EEMs of the two analytes as analytical signals allowed their simultaneous determination in human urine samples, in the therapeutic ranges from ca. 40 to 150 mg l -1 . The proposed procedure was validated by comparing the obtained results with a capillary electrophoresis method, with satisfactory results for the assayed method. © 2006 Elsevier B.V. All rights reserved. Keywords: Second-order multivariate calibration; PARAFAC; Excitation–emission matrices; Flufenamic acid; Meclofenamic acid; Urine; Micellar media 1. Introduction Flufenamic [N-(,,-trifluoro-m-tolyl) anthranilic] and meclofenamic [N-(2,6-dichloro-m-tolyl) anthranilic] acids are potent analgesic and anti-inflammatory drugs. They have been used in the treatment of arthritis and other illnesses related to muscular-skeletal problems [1]. Some cases of nephrotoxicity have been reported, making the monitoring of these drugs in biological fluids of pharmacological and medical interest [2]. The fluorescent properties of these anthranilic acid deriva- tives have been widely exploited for analytical purposes. Mehta and Schulman [3], suggested that their native fluorescence in organic solvents, e.g. dioxane and chloroform, could be useful ∗ Corresponding author. Tel.: +34 924289378; fax: +34 924289375. E-mail address: arsenio@unex.es (A. Mu ˜ noz de la Pe ˜ na). for their determination. Miller et al. [4] reported that flufenamic and mefenamic acids showed no significant fluorescence at room temperature in acidic, neutral or alkaline ethanol solution, but were strongly fluorescent at low temperature. Dell et al. [5] indicated that solvents with a very low dielectric constant were required to obtain fluorescent solutions. Other spectrofluorimet- ric methods reported for the determination of these compounds involve the use of different fluorogenic agents [6–8]. The application of luminescent techniques to the analysis of drugs in biological fluids is advantageous because of the high sensitivity that can be achieved. However, the selectivity is often reduced by extensive spectral overlap or in the presence of matrix interferences. In this context, multivariate calibration methods are helpful in circumventing the selectivity problems. Recently, a new method for the determination of flufenamic, mefenamic and meclofenamic acids in mixtures by fluorescence emission spectroscopy, using a partial least-squares (PLS) first 0003-2670/$ – see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.aca.2006.03.077