Microchimica Acta

, 185:562 | Cite as

A capillary column packed with a zirconium(IV)-based organic framework for enrichment of endogenous phosphopeptides

  • Haizhu Lin
  • Hemei Chen
  • Xi Shao
  • Chunhui DengEmail author
Original Paper


A zirconium(IV)-based metal organic framework (Zr-MOF) was deposited on polydopamine-coated silica microspheres to form microspheres of type SiO2@PDA@Zr-MOF. These were packed into capillary columns for enrichment of phosphopeptides. The column was off-line coupled to both matrix-assisted laser desorption/ionization time of flight mass spectrometry and LC-ESI-MS/MS. The method has a detection limit as low as 4 fmol of β-casein digest and a selectivity as high as 1:1000 (molar ratio of β-casein and BSA digest). It was applied to the analysis of human saliva. In total, 240 endogenous phosphopeptides were identified in only 25 μL human saliva.

Graphical abstract

A zirconium-based metal organic framework (Zr-MOF) was modified outside of polydopamine-coated silica microspheres to form microspheres named SiO2@PDA@Zr-MOF. Then they were packed in capillary columns for selective enrichment of phosphopeptides via interaction between Zr-O clusters and phosphate groups. The pre-concentration resulted in a better detection of phosphopeptides by mass spectrometry. Tris: Tris(hydroxymethyl)aminomethane; DMF: Dimethyl Formamide; Zr-MOF: Zirconium(IV)-organic framework; MOAC: Metal oxide affinity chromatography.


MOF Human saliva Mass spectrometry Metal oxide affinity chromatography Phosphopeptide On-column enrichment MALDI-TOF MS Phosphoproteome Polydopamine Size exclusion effect 



This work was financially supported by National Key R&D Program of China (2018YFA0507501) and the National Natural Science Foundation of China (21425518).

Compliance with ethical standards

The author(s) declare that they have no competing interests.

Supplementary material

604_2018_3109_MOESM1_ESM.docx (1.2 mb)
ESM 1 (DOCX 1.20 MB)
604_2018_3109_MOESM2_ESM.xlsx (25 kb)
ESM 2 (XLSX 25 kb)


  1. 1.
    Lundby A, Secher A, Lage K, Nordsborg NB, Dmytriyev A, Lundby C, Olsen JV (2012) Quantitative maps of protein phosphorylation sites across 14 different rat organs and tissues. Nat Commun 3:876. CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Huttlin EL, Jedrychowski MP, Elias JE, Goswami T, Rad R, Beausoleil SA, Villen J, Haas W, Sowa ME, Gygi SP (2010) A tissue-specific atlas of mouse protein phosphorylation and expression. Cell 143(7):1174–1189. CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Aebersold R, Mann M (2003) Mass spectrometry-based proteomics. Nature 422:198–207. CrossRefGoogle Scholar
  4. 4.
    Leitner A, Sturm M, Lindner W (2011) Tools for analyzing the phosphoproteome and other phosphorylated biomolecules: a review. Anal Chim Acta 703(1):19–30. CrossRefGoogle Scholar
  5. 5.
    Hortin GL (2006) The MALDI-TOF mass spectrometric view of the plasma proteome and peptidome. Clin Chem 52(7):1223–1237. CrossRefGoogle Scholar
  6. 6.
    Wang ZG, Lv N, Bi WZ, Zhang JL, Ni JZ (2015) Development of the affinity materials for phosphorylated proteins/peptides enrichment in phosphoproteomics analysis. ACS Appl Mater Interfaces 7(16):8377–8392. CrossRefGoogle Scholar
  7. 7.
    Kailasa SK, Wu H-F (2014) Recent developments in nanoparticle-based MALDI mass spectrometric analysis of phosphoproteomes. Microchim Acta 181(9–10):853–864. CrossRefGoogle Scholar
  8. 8.
    Tan S, Wang J, Han Q, Liang Q, Ding M (2018) A porous graphene sorbent coated with titanium(IV)-functionalized polydopamine for selective lab-in-syringe extraction of phosphoproteins and phosphopeptides. Microchim Acta 185(7):316. CrossRefGoogle Scholar
  9. 9.
    Jiang J, Sun X, She X, Li J, Li Y, Deng C, Duan G (2018) Magnetic microspheres modified with Ti(IV) and Nb(V) for enrichment of phosphopeptides. Microchim Acta 185(6):309. CrossRefGoogle Scholar
  10. 10.
    Zhang L, Gan Y, Sun H, Yu B, Jin X, Zhang R, Zhang W, Zhang L (2016) Magnetic mesoporous carbon composites incorporating hydrophilic metallic nanoparticles for enrichment of phosphopeptides prior to their determination by MALDI-TOF mass spectrometry. Microchim Acta 184(2):547–555. CrossRefGoogle Scholar
  11. 11.
    Yan Y, Zheng Z, Deng C, Li Y, Zhang X, Yang P (2013) Hydrophilic polydopamine-coated graphene for metal ion immobilization as a novel immobilized metal ion affinity chromatography platform for phosphoproteome analysis. Anal Chem 85(18):8483–8487. CrossRefGoogle Scholar
  12. 12.
    Gu ZY, Chen YJ, Jiang JQ, Yan XP (2011) Metal-organic frameworks for efficient enrichment of peptides with simultaneous exclusion of proteins from complex biological samples. Chem Commun 47(16):4787–4789. CrossRefGoogle Scholar
  13. 13.
    Zhu X, Gu J, Yang J, Wang Z, Li Y, Zhao L, Zhao W, Shi J (2015) Zr-based metal–organic frameworks for specific and size-selective enrichment of phosphopeptides with simultaneous exclusion of proteins. J Mater Chem B 3(20):4242–4248. CrossRefGoogle Scholar
  14. 14.
    Peng J, Zhang H, Li X, Liu S, Zhao X, Wu J, Kang X, Qin H, Pan Z, Wu R (2016) Dual-metal centered zirconium-organic framework: a metal-affinity probe for highly specific interaction with Phosphopeptides. ACS Appl Mater Interfaces 8(51):35012–35020. CrossRefGoogle Scholar
  15. 15.
    Zhao M, Deng C, Zhang X (2014) The design and synthesis of a hydrophilic core-shell-shell structured magnetic metal-organic framework as a novel immobilized metal ion affinity platform for phosphoproteome research. Chem Commun 50(47):6228–6231. CrossRefGoogle Scholar
  16. 16.
    Liu Q, Sun N, Gao M, Deng C (2018) Magnetic binary metal–organic framework as a novel affinity probe for highly selective capture of endogenous Phosphopeptides. ACS Sustain Chem Eng 6(3):4382–4389. CrossRefGoogle Scholar
  17. 17.
    Zhou J, Liang Y, He X, Chen L, Zhang Y (2017) Dual-functionalized magnetic metal–organic framework for highly specific enrichment of Phosphopeptides. ACS Sustain Chem Eng 5(12):11413–11421. CrossRefGoogle Scholar
  18. 18.
    Yang X, Xia Y (2016) Urea-modified metal-organic framework of type MIL-101(Cr) for the preconcentration of phosphorylated peptides. Microchim Acta 183(7):2235–2240. CrossRefGoogle Scholar
  19. 19.
    Li D, Bie Z (2017) Metal–organic framework incorporated monolithic capillary for selective enrichment of phosphopeptides. RSC Adv 7(26):15894–15902. CrossRefGoogle Scholar
  20. 20.
    Li D, Yin D, Chen Y, Liu Z (2017) Coupling of metal-organic frameworks-containing monolithic capillary-based selective enrichment with matrix-assisted laser desorption ionization-time-of-flight mass spectrometry for efficient analysis of protein phosphorylation. J Chromatogr A 1498:56–63. CrossRefGoogle Scholar
  21. 21.
    Yang H, Deng C, Zhang X (2016) Preparation of Ti(4+)-immobilized modified silica capillary trapping column for on-line selective enrichment of phosphopeptides. Talanta 153:285–294. CrossRefGoogle Scholar
  22. 22.
    Choi H, Lee S, Jun CD, Park ZY (2011) Development of an off-line capillary column IMAC phosphopeptide enrichment method for label-free phosphorylation relative quantification. J Chromatogr B Anal Technol Biomed Life Sci 879(28):2991–2997. CrossRefGoogle Scholar
  23. 23.
    Richardson BM, Soderblom EJ, Thompson JW, Moseley MA (2013) Automated, reproducible, titania-based phosphopeptide enrichment strategy for label-free quantitative phosphoproteomics. J Biomol Tech 24(1):8–16. CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Ruprecht B, Koch H, Medard G, Mundt M, Kuster B, Lemeer S (2015) Comprehensive and reproducible phosphopeptide enrichment using iron immobilized metal ion affinity chromatography (Fe-IMAC) columns. Mol Cell Proteomics 14(1):205–215. CrossRefGoogle Scholar
  25. 25.
    Xie Y, Deng C (2017) Designed synthesis of a "one for two" hydrophilic magnetic amino-functionalized metal-organic framework for highly efficient enrichment of glycopeptides and phosphopeptides. Sci Rep 7(1):1162. CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Lin H, Shao X, Lu Y, Deng C (2018) Preparation of iminodiacetic acid functionalized silica capillary trap column for on-column selective enrichment of N-linked glycopeptides. Talanta 188:499–506. CrossRefGoogle Scholar
  27. 27.
    Vitorino R, Lobo MJ, Duarte JR, Ferrer-Correia AJ, Domingues PM, Amado FM (2005) The role of salivary peptides in dental caries. Biomed Chromatogr 19(3):214–222. CrossRefGoogle Scholar
  28. 28.
    Long X, Li J, Sheng D, Lian H (2016) Low-cost iron oxide magnetic nanoclusters affinity probe for the enrichment of endogenous phosphopeptides in human saliva. RSC Adv 6(98):96210–96222. CrossRefGoogle Scholar
  29. 29.
    Su J, He X, Chen L, Zhang Y (2017) Adenosine phosphate functionalized magnetic mesoporous graphene oxide nanocomposite for highly selective enrichment of Phosphopeptides. ACS Sustain Chem Eng 6(2):2188–2196. CrossRefGoogle Scholar
  30. 30.
    Sun N, Deng C, Li Y, Zhang X (2014) Size-exclusive magnetic graphene/mesoporous silica composites with titanium(IV)-immobilized pore walls for selective enrichment of endogenous phosphorylated peptides. ACS Appl Mater Interfaces 6(14):11799–11804. CrossRefGoogle Scholar
  31. 31.
    La Barbera G, Capriotti AL, Cavaliere C, Ferraris F, Montone CM, Piovesana S, Zenezini Chiozzi R, Lagana A (2018) Saliva as a source of new phosphopeptide biomarkers: development of a comprehensive analytical method based on shotgun peptidomics. Talanta 183:245–249. CrossRefGoogle Scholar
  32. 32.
    Lin H, Deng C (2016) Development of immobilized Sn(4+) affinity chromatography material for highly selective enrichment of phosphopeptides. Proteomics 16(21):2733–2741. CrossRefGoogle Scholar
  33. 33.
    Xiong Z, Zhang L, Fang C, Zhang Q, Ji Y, Zhang Z, Zhang W, Zou H (2014) Ti4+−immobilized multilayer polysaccharide coated magnetic nanoparticles for highly selective enrichment of phosphopeptides. J Mater Chem B 2(28):4473–4480. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Austria, ein Teil von Springer Nature 2018

Authors and Affiliations

  1. 1.The Fifth People’s Hospital of Shanghai, Fudan UniversityShanghaiChina
  2. 2.Institutes of Biomedical Sciences, Fudan UniversityShanghaiChina
  3. 3.Department of Chemistry, and Collaborative Innovation Center of Genetics and DevelopmentFudan UniversityShanghaiChina

Personalised recommendations