Current Pharmaceutical Analysis, 2012, 8, 00-00 1 1573-4129/12 $58.00+.00 © 2012 Bentham Science Publishers Development and Validation of UHPLC/ESI-Q-TOF-MS Method for Terbutaline Estimations in Experimental Rodents: Stability Effects and Plasma Pharmacokinetics Md. Faiyazuddin 1,2, *, Niyaz Ahmad 1 , Zeenat Iqbal 1 , Sushma Talegaonkar 1 , Aseem Bhatnagar 3 , Roop Krishen Khar 1 , Farhan Jalees Ahmad 1 1 Department of Pharmaceutics, Faculty of Pharmacy, Hamdard University, New Delhi-110062, India; 2 Department of Pharmaceutics, Faculty of Pharmacy, Integral University, Lucknow, Uttar Pradesh, India; 3 Department of Nuclear Medicine, Institute of Nuclear Medicine & Allied Sciences (INMAS), Brig. S. K. Mazumdar Road, Delhi-110054, India Abstract: An ultra high performance liquid chromatography-electrospray ionization-tandem mass spectrometric method (UHPLC/ESI-Q-TOF-MS) for the analysis of terbutaline (TB) in Wistar rat plasma has been developed and validated. The chromatographic separation was achieved on a Waters ACQUITY UPLC TM BEH C18 (100.0 mm2.1 mm; 1.7 m) col- umn using isocratic mobile phase, consisting of 2 mM ammonium acetate and acetonitrile (90: 10; v/v), at a flow rate of 0.25 mL min -1 . The transitions occurred at m/z 226.19152.12 for TB, and m/z 260.34183.11 for the internal standard. The recovery of the analytes from Wistar rat plasma was optimized using liquid- liquid extraction technique (LLE) in ethyl acetate. The total run time was 3.0 min and the elution of TB occurred at 1.85±0.05 min. The linear dynamic range was established over the concentration range 1–1000 ng mL -1 (r 2 ; 0.9938±0.0005) for TB. The intra-assay and inter-assay accuracy in terms of % CV was in between 1.8–3.5. The lower limit of quantitation (LLOQ) for TB was 1.0 ng mL -1 . Analytes were stable under various conditions (in autosampler, during freeze–thaw, at room temperature, and under deep- freeze conditions). The developed method was successfully applied for pharmacokinetic profiling in rodents. Keywords: Asthma, Bronchodilator, Ex-vivo Stability, In-vitro assay, Matrix effect, Pharmacokinetics, Terbutaline; UHPLC/ESI-Q-TOF-MS, Validation. 1. INTRODUCTION Terbutaline (TB), an amphiphilic  -[(tert-butylamino) methyl]-3,5-dihydroxy- benzyl alcohol is a synthetic  2 - adrenoceptor ( 2 AR) stimulant that is widely used to treat bronchial asthma (BA), chronic obstructive pulmonary dis- eases (COPD) and related disorders [1, 2]. There has been a plethora of analytical methods reported for the quantitative estimation of TB in bulk, pharmaceutical formulations and biomatrices mainly based on high performance-liquid chro- matography (HPLC) [3-11], liquid chromatography-tandem mass spectrometry (LC/MS/MS) [12, 13] and gas chroma- tography-mass spectrometry (GC-MS) [14, 15]. These pub- lished methods either involve cumbersome sample prepara- tion steps or are insufficiently sensitive or precise to perform TB analysis. A validated HPLC method for the determina- tion of TB and its possible degradation products has been reported in literature [16], utilizing Hypersil 100 C 18 , 1504.6 mm (5 m) column, found inadequate sensitivity of the method due to high limit of quantitation (LOQ; 0.11 g mL -1 ) and limit of detection (LOD; 0.37 g mL -1 ) values. In our previous study, a stability-indicating high performance- thin layer chromatography (HPTLC) has been developed and validated for the direct estimation of TB and its potential degradants in bulk and submicronized dry powder inhalers *Address correspondence to this author at the Department of Pharmaceutics, Faculty of Pharmacy, Hamdard University, New Delhi-110062, India; Tel: +91-7668963486; E-mail: md.faiyazuddin2008@gmail.com [17]. The low value of LOQ (28.35 ng spot –1 ) and LOD (9.41 ng spot –1 ) indicated adequate assay sensitivity of our reported method. Recently, a chiral liquid chromatography– tandem mass spectrometric method (LC–MS/MS) for the simultaneous analysis of bambuterol and its active metabo- lite terbutaline in Wistar rat plasma has been reported [18]. In this method, the chromatographic separation was achieved on a teicoplanin-containing Chirobiotic T chiral (250 mm4.6 mm; 5 m) column at a flow-rate of 0.4 mL min -1 , however, and has the disadvantage of being time-consuming because of longer retention time (R T >15 min) and lengthy processing steps. All these studies have further emphasized the need to perform rapid and sensitive quality-control analysis of TB with shorter retention time and improved ef- fects. An approach to chiral separation and analysis of TB enantiomers by chiral complexation and electrospray high- field asymmetric waveform ion mobility spectrometry cou- pled to mass spectrometry (ESI-FAIMS-MS) has been also reported in literature [19]. The limit of detection is 0.10% ()-terbutaline in a sample of (+)-terbutaline, which is the limiting factor of the method. In a recent study, TB sulfocon- jugates were examined in urinary samples by LC/ESI- MS/MS on Orbitrap mass spectrometer [20]. However, this method was not sensitive enough to detect TB in urine on single dose administration. Therefore a hyphenated chroma- tographical technique with advanced feature is desired to compensate the aforesaid loopholes. However, UHPLC is a novel chromatographic technique utilizing high linear veloci- ties, which is based on concept using columns with smaller