Analytical and Bioanalytical Chemistry

, Volume 409, Issue 4, pp 1059–1065 | Cite as

The use of a temperature-responsive column for the direct analysis of drugs in serum by two-dimensional heart-cutting liquid chromatography

  • Toshiyasu Mikuma
  • Ryo Uchida
  • Mizuki Kajiya
  • Yuki Hiruta
  • Hideko Kanazawa
Research Paper


A novel pretreatment method, which was performed using a two-dimensional high-performance liquid chromatography (2D-HPLC) system, was proposed for the direct analysis of drugs in human serum. A temperature-responsive column was used as a pretreatment column. The stationary phase of the temperature-responsive column exhibits temperature-regulated hydrophilic/hydrophobic characteristics. Controlling the ionic strength of the eluent enables human serum albumin (HSA) to pass through the column without retention. When serum samples containing barbiturates or benzodiazepines were injected into the temperature-responsive column using 10 mM of ammonium acetate (pH 6.5) as the mobile phase and in the temperature range of 10–40 °C, HSA was eluted from the column near the dead time, followed by the individual drugs. When the column temperature was changed, the retention times of the drugs were altered owing to surface property changes within the pretreatment column. These closely eluted compounds were subsequently introduced into the analytical column using a column-switching valve, with a minimal gap time to avoid foreign substance contamination. This new 2D-HPLC method afforded high-quality chromatograms of multiple drugs without unwanted peaks from foreign substances. The present technique could be an attractive choice in selecting the analytical method for drug analysis.


Two-dimensional liquid chromatography Temperature-responsive chromatography Drugs Serum Heart-cutting 



The authors appreciate Hitachi High-Technologies Corporation for providing valuable information. This study was supported in part by a Grant-in-Aid for Scientific Research (B) (Grant No. 21390012 and 25293009) from the Ministry of Education, Culture, Sports, Science and Technology, Japan.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Supplementary material

216_2016_24_MOESM1_ESM.pdf (219 kb)
ESM 1 (PDF 218 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Toshiyasu Mikuma
    • 1
    • 2
  • Ryo Uchida
    • 1
  • Mizuki Kajiya
    • 1
  • Yuki Hiruta
    • 1
  • Hideko Kanazawa
    • 1
  1. 1.Graduate School of Pharmaceutical SciencesKeio UniversityTokyoJapan
  2. 2.Forensic Science LaboratoryChiba Prefectural Police H.Q.Chiba-cityJapan

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