Sensitive Determination of Four Polypeptide Antibiotic Residues in Milk Powder by High Performance Liquid Chromatography–Electrospray Tandem Mass Spectrometry

  • Tong Liu
  • Chuanbin Zhang
  • Feng ZhangEmail author
  • Bo Nie
  • Fei Yuan
  • Hailan Huang
  • Hongna Li


Polypeptide antibiotics abuse can lead to antibiotic residues in food products and have unwanted effects on human health. A selective, accurate, and sensitive analytical method for the simultaneous determination of four polypeptide antibiotics (gramicidin S, bacitracin, polymyxin B, and polymyxin E) in infant formula powder was developed using solid-phase extraction (SPE) combined with high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS). The samples were extracted with acidified methanol, deproteinized with acetonitrile, and degreased with n-hexane before SPE. After cleanup with 0.1% formic acid and methanol (3:1, v/v) in Oasis HLB cartridges, the extracts were analyzed by HPLC–MS/MS with electrospray ionization (ESI) source and time-scheduled multiple reaction monitoring (MRM). Linearity was assessed by using matrix-matched standard calibration and good determination coefficients (r2 > 0.995) were obtained. The average recoveries for blank sample at three spiked concentration levels were in the range of 82.8–101.2%. The limits of detection (LODs) and limits of quantitation (LOQs) of all analytes were in the range of 5–15 μg kg−1 and 20–50 μg kg−1, respectively. The intra-day and inter-day precisions were lower than 10%. The results of method validation demonstrated that the developed method is accurate and reliable, and it can be applied for screening and quantitation of target polypeptide antibiotics in food.

Graphic Abstract


Polypeptide antibiotics HPLC–MS/MS Milk powder Gramicidin S 



The authors wish to acknowledge the financial support given by the project of National Key Research and Development Program of China and Public Welfare Fund of Inspection and Quarantine of China.


This study was funded by the project of National Key Research and Development Program of China and Public Welfare Fund of Inspection and Quarantine of China (Grant Numbers 2018YFC1603606, 201510038, 2016YFD0401103, and 2016YFF0203903).

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Ethical Approval

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

Supplementary material

10337_2019_3777_MOESM1_ESM.docx (161 kb)
Supplementary material 1 (DOCX 161 kb)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Institute of Food SafetyChinese Academy of Inspection and QuarantineBeijingChina
  2. 2.School of Chemistry and Chemical EngineeringQingdao UniversityQingdaoChina
  3. 3.Key Laboratory of Chinese Internal Medicine of Ministry of Education and Beijing, Dongzhimen HospitalBeijing University of Chinese MedicineBeijingChina

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