RESEARCH ARTICLE An LC–MS/MS method for the quantification of diclofenac sodium in dairy cow plasma and its application in pharmacokinetics studies Ya‐Jun Yang | Xi‐Wang Liu | Xiao‐Jun Kong | Zhe Qin | Shi‐Hong Li | Zeng‐Hua Jiao | Jian‐Yong Li Key Laboratory of New Animal Drug Project of Gansu Province, Key Laboratory of Veterinary Pharmaceutical Development of Ministry of Agriculture, Lanzhou Institute of Husbandry and Pharmaceutical Sciences of Chinese Academy of Agricultural Sciences, Lanzhou, China Correspondence Prof. Dr. Jian‐Yong Li, Key Laboratory of New Animal Drug Project of Gansu Province, Key Laboratory of Veterinary Pharmaceutical Development of Ministry of Agriculture, Lanzhou Institute of Husbandry and Pharmaceutical Sciences of Chinese Academy of Agricultural Sciences, No. 335, Jiangouyan, Qilihe district, Lanzhou, Gansu province, 730050, China. Email: lijy1971@126.com Funding information Science‐technology innovation engineering of CAAS, Grant/Award Number: CAAS‐ASTIP‐ 2014‐LIHPS‐2 Abstract An LC–MS/MS method with internal standard tolfenamic acid for determining diclofenac sodium (DCF) in dairy cow plasma was developed and validated. Samples were processed with protein precipitation by cold formic acid–acetonitrile. Determi- nation of DCF was performed using LC–ESI + –MS/MS with the matrix‐matched cali- bration curve. The results showed that the method was sensitive (LOD 2 ng mL -1 , LOQ 5 ng mL -1 ), accurate (97.60 ± 5.64%), precise (<10%) and linear in the range of 5–10,000 ng mL -1 . A single intravenous (i.v.) or intramuscular (i.m.) administration of 5% diclofenac sodium injection at a dose of 2.2 mg kg -1 was performed in six healthy dairy cows according to a two‐period crossover design. The main pharmaco- kinetic (PK) parameters after a single i.v. administration were as follows: t 1/2β , 4.52 ± 1.71 h; AUC, 77.79 ± 16.76 h μg mL -1 ; mean residence time, 5.16 ± 1.11 h. The main PK parameters after a single i.m. administration were as follows: T max , 2.38 ± 1.19 h; C max , 7.46 ± 1.85 μg mL -1 ; t 1/2β , 9.46 ± 2.86 h; AUC 67.57 ± 13.07 h μg mL -1 . The absolute bioavailability was 87.37 ± 5.96%. The results showed that the diclofenac sodium injection had PK characteristics of rapid absorption and slow elimination, and high peak concentration and bioavailability in dairy cows, and that the recommended clinical dosage of diclofenac sodium injection is 2.2 mg kg -1 . KEYWORDS bioavailability, dairy cows, diclofenac sodium, LC–MS/MS, pharmacokinetics 1 | INTRODUCTION Diclofenac sodium (DCF), sodium 2‐[(2, 6‐dichlorophenyl)amino] phenylacetate (CAS no. 15307‐79‐6), is a nonsteroidal anti‐ inflammatory drug (NSAID) and, more specifically, a phenyl acetic acid derivative (Figure 1). DCF shows preferential inhibition of the cyclooxygenase‐2 (COX‐2) enzyme (Blain et al., 2002; Viegas, Manso, Corvo, Marques, & Cabrita, 2011). DCF has been used in human medicine for many years (Goa & Chrisp, 1992; Todd & Sorkin, 1988). It has been proven to be equally effective when compared with other commonly used NSAIDs. For the long‐term symptomatic treatment of rheumatoid arthritis, osteoarthri- tis, ankylosing spondylitis and acute gouty arthritis, DCF is adminis- tered through different routes and may be used for up to 12 weeks. DCF is used for the alleviation of post‐operative pain and dysmenor- rhoea on a short‐term basis as it possesses potent analgesic effects. DCF has been authorized for treatment in cattle and swine as an anti‐inflammatory agent by the European Medicines Agency (EMEA, n.d.). It was reported that a single intramuscular (i.m.) dose of DCF was equally effective as three intravenous (i.v.) injections of flunixin Received: 28 November 2018 Revised: 26 January 2019 Accepted: 22 February 2019 DOI: 10.1002/bmc.4520 Biomedical Chromatography. 2019;33:e4520. https://doi.org/10.1002/bmc.4520 © 2019 John Wiley & Sons, Ltd. wileyonlinelibrary.com/journal/bmc 1 of 8