Feasibility of Trypsin Digestion as a Sample Preparation for Daptomycin Quantification in Murine Skeletal Muscles

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  • Sakai Yusuke
    The United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University
  • Murakami Eriko
    Gifu Pharmaceutical University
  • Kato Hideo
    Department of Pharmacy, Aichi Medical University School of Hospital
  • Ohyama Kaname
    Department of Pharmacy Practice, Graduate School of Biomedical Sciences, Nagasaki University
  • Esaka Yukihiro
    The United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University Gifu Pharmaceutical University
  • Yamamoto Takuhei
    Gifu Pharmaceutical University
  • Hagihara Mao
    Department of Pharmacy, Aichi Medical University School of Hospital
  • Mikamo Hiroshige
    Department of Infection Control and Prevention, Aichi Medical University School of Hospital
  • Uno Bunji
    The United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University Gifu Pharmaceutical University

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<p>It is important to evaluate the amount of daptomycin (DAP) distributed to skeletal muscles to elucidate the mechanisms related to penetration and side effects, such as myopathies. However, no attempt has been made to measure DAP concentrations in skeletal muscles. The study’s aim to investigate the feasibility of trypsin digestion, as a muscle sample preparation technique for the determination of DAP in murine skeletal muscle, was evaluated in conjunction with a conventional HPLC-UV analysis. Compared with trypsin digestion, DAP was less recovered from spiked skeletal muscle by the conventional extraction, including homogenization, centrifugation, and filtration, because of its incorporation into the muscle protein. On the other hand, a sample preparation technique involving enzymatic digestion employing trypsin fully recovered DAP from the spiked skeletal muscle. Based on the spike recovery assay results, we proposed an efficient muscle sample preparation method involving trypsin digestion. HPLC analysis in conjunction with the sample preparation method has successfully determined DAP concentrations of skeletal muscles collected from mice administrated subcutaneously with DAP. The proposed method is suitable for application to investigations that include animal experiments on drug migration into muscle and mechanism underlying skeletal muscle injury as a side reaction, such as myopathies, of DAP therapy.</p>

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