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Second-tier test for quantification of underivatized amino acids in dry blood spot for metabolic diseases in newborn screening

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Abstract

The quantitative analysis of amino acids (AAs) in single dry blood spot (DBS) samples is an important issue for metabolic diseases as a second-tier test in newborn screening. An analytical method for quantifying underivatized AAs in DBS was developed by using liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS). The sample preparation in this method is simple and ion-pairing agent is not used in the mobile phase that could avoid ion suppression, which happens in mass spectrometry and avoids damage to the column. Through chromatographic separation, some isomeric compounds could be identified and quantified, which cannot be solved through only appropriate multiple reactions monitoring transitions by MS/MS. The concentrations of the different AAs were determined using non-deuterated internal standard. All calibration curves showed excellent linearity within test ranges. For most of the amino acids the accuracy of extraction recovery was between 85.3 and 115 %, and the precision of relative standard deviation was <7.0 %. The 35 AAs could be identified in DBS specimens by the developed LC–MS/MS method in 17–19 min, and eventually 24 AAs in DBS were quantified. The results of the present study prove that this method as a second-tier test in newborn screening for metabolic diseases could be performed by the quantification of free AAs in DBS using the LC–MS/MS method. The assay has advantages of high sensitive, specific, and inexpensive merits because non-deuterated internal standard and acetic acid instead of ion-pairing agent in mobile phase are used in this protocol.

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (No. 81073040) and Natural Science Foundation of Jilin (No. 201215094).

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Authors and Affiliations

  1. Chang Chun Institute of Applied Chemistry, Chinese Academy of Sciences, 130022, Changchun, People’s Republic of China

    Chunyan Wang, Hongbin Zhu, Fengrui Song, Zhiqiang Liu & Shuying Liu

  2. Graduate School of the Chinese Academy of Sciences, 100039, Beijing, People’s Republic of China

    Chunyan Wang & Hongbin Zhu

  3. Jilin Center of Newborn Screening, Jilin Maternal and Child Health Hospital, 130061, Changchun, People’s Republic of China

    Wenyan Zhang

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  1. Chunyan Wang
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  2. Hongbin Zhu
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  3. Wenyan Zhang
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  4. Fengrui Song
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  5. Zhiqiang Liu
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  6. Shuying Liu
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Correspondence to Fengrui Song.

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Wang, C., Zhu, H., Zhang, W. et al. Second-tier test for quantification of underivatized amino acids in dry blood spot for metabolic diseases in newborn screening. Amino Acids 44, 661–671 (2013). https://doi.org/10.1007/s00726-012-1389-5

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  • DOI: https://doi.org/10.1007/s00726-012-1389-5

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