A “Two-in-One” Tandem Immunoaffinity Column for the Sensitive and Selective Purification and Determination of Trace/Ultra-Trace Olaquindox and Its Major Metabolite in Fish Tissues by LC–MS/MS

  • Jie Xie
  • Weijie Zeng
  • Xiaoyun Gong
  • Rui Zhai
  • Zejian Huang
  • Meiying Liu
  • Guoqing Shi
  • You Jiang
  • Xinhua DaiEmail author
  • Xiang FangEmail author


A “two-in-one” bilayer immunoaffinity column (IAC) coupled in tandem was prepared for the highly selective purification of olaquindox (OLA) and its major metabolite, 3-methyl-quinoxaline-2-carboxylic acid (MQCA), and their determinations were carried out with the developed high-performance liquid chromatography–tandem mass spectrometry (LC–MS/MS) method. The prepared highly selective two-in-one IAC was used for fish tissue sample clean-up after acidolysis. Then, the purified samples were analyzed by LC–MS/MS in ESI+ and multiple reaction monitoring mode. The calibration curves showed excellent linearity, with a correlation coefficient (R2) greater than 0.999. Under the optimal conditions, the average recoveries at the spiked levels of 1.0, 5.0, and 10.0 μg/kg were 81.2–94.6% with intra- and inter-day relative standard deviations (RSD; intra-day, n = 5; inter-day, n = 3) that were less than 8.1% and 11.7%, respectively. The limits of detection (LODs) were 0.01 and 0.3 μg kg−1, and the limits of quantifications (LOQs) were 0.04 and 0.5 μg kg−1 for OLA and MQCA, respectively. The validated method was successfully applied to the qualitative and quantitative detection of real samples collected from several local markets.


Olaquindox 3-methyl-quinoxaline-2-carboxylic acid Immunoaffinity columns LC–MS/MS Fish 



We sincerely thank Prof. Bei Xu, Prof. Dazhou Chen, Shiying Chu, Chang Li, and Yuting An for their help with experiments.

Funding Information

This study was supported financially by grants from the National Key R&D Program of China (2016YFF0200504, 2016YFF0200502, 2018YFF0212503), the National Natural Science Foundation of China (21575132, 21605135), and Research Project of National Institute of Metrology (AKYCX1904).

Compliance with Ethical Standards

Conflict of Interest

Jie Xie declares that she has no conflict of interest. Weijie Zeng declares that he has no conflict of interest. Xiaoyun Gong declares that he has no conflict of interest. Rui Zhai declares that she has no conflict of interest. Zejian Huang declares that he has no conflict of interest. Meiying Liu declares that she has no conflict of interest. Guoqing Shi declares that he has no conflict of interest. You Jiang declares that he has no conflict of interest. Xinhua Dai declares that she has no conflict of interest. Xiang Fang declares that he has no conflict of interest.

Ethical Approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Informed Consent

Not applicable.


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Jie Xie
    • 1
  • Weijie Zeng
    • 1
    • 2
  • Xiaoyun Gong
    • 1
  • Rui Zhai
    • 1
  • Zejian Huang
    • 1
  • Meiying Liu
    • 1
  • Guoqing Shi
    • 2
  • You Jiang
    • 1
  • Xinhua Dai
    • 1
    Email author
  • Xiang Fang
    • 1
    Email author
  1. 1.Mass Spectrometry Engineering Technology Research Center, Center for Advanced Measurement ScienceNational Institute of MetrologyBeijingPeople’s Republic of China
  2. 2.School of Chemistry and Biological EngineeringUniversity of Science and Technology BeijingBeijingPeople’s Republic of China

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