Peptide Enrichment DOI: 10.1002/anie.200603917 Selective Extraction of Peptides from Human Plasma by Highly Ordered Mesoporous Silica Particles for Peptidome Analysis** Ruijun Tian, He Zhang, Mingliang Ye, Xiaogang Jiang, Lianghai Hu, Xin Li, Xinhe Bao, and Hanfa Zou* The peptidome of human plasma has attracted increasing interest for its role in the elucidation of biological and pathological variation and discovery of useful biomarkers. [1] Recently, the development of mass spectrometry techniques has increased efforts to broadly characterize its peptide constituents. However, the complexity and high dynamic rangeofhumanplasmamakethecharacterizationachalleng- ing task. [2] Toward this end, two main methodologies are currently used to enrich peptides from plasma samples. Centrifugal ultrafiltration with accurate molecular weight (MW) cutoff is the most widely used method to extract peptides by removing proteins with larger MWs based on a size-exclusion mechanism. [3] However, as a result of the high protein content of human plasma (60–80 mgmL À1 ), the ultrafiltration time will increase sharply if a large amount of plasma sample is applied. Furthermore, other low-MW contaminants (for example, salts) will also be concentrated. Another option is to use adsorbents carrying charged or hydrophobic groups for peptide adsorption. [4] While the peptides are enriched, various proteins will be adsorbed equally. Therefore, it is necessary to develop a general method to overcome the limitations of the state-of-the-art methods. Silica particles with highly ordered mesostructures (for example, M41s [5] and SBA-15 [6] ) have been widely applied in the fields of separation and adsorption. [7] Theuniqueproper- tiesofthesesilicamaterialsincludehighin-poresurfaceareas, extremely narrow pore size distribution, perfectly adjustable pore size, and the presence of silanol groups and siloxane bridges,whichmakethemeffectiveforselectiveadsorptionof standard proteins with different MWs based on the size- exclusion mechanism. [8] In this study, attempts were made to usethesematerialswithcriticalporesizestoselectivelyenrich peptides from human plasma, but exclude other proteins by an accurate MW cutoff like that of centrifugal ultrafiltration. Three types of highly ordered mesoporous silica particles with different pore sizes were synthesized according to reported procedures. [5,6,9] A representative TEM image and theporesizedistributiondiagramforMCM-41silicaparticles (Figure1) demonstrate the existence of highly ordered cylindrical mesopores running along the same direction. This property is critical for selectively adsorbing peptides withaccurateMWcutoffandenhancedmasstransferforfast adsorption/desorption equilibration. The characteristics of the three kinds of mesoporous silica particles with similar mesostructuresarelistedinTable1.Allofthesilicaparticles Figure 1. Barrett–Joyner–Halenda (BJH) pore size distribution calcu- lated from the adsorption branch of the nitrogen adsorption/desorp- tion isotherm. Inset: TEM image of corresponding MCM-41 mesopo- rous silica particles. Table 1: Comparison of the material properties of three types of mesoporous silica particles and their adsorption efficiency for standard protein lysozyme. Silica particles Surface area [m 2 g À1 ] [a] Pore size [] [a] Amounts of lysozyme adsorbed [mgg À1 ] [b] MCM-41 871 20.5 63.6 C-SBA-15 645 84.5 191.3 L-SBA-15 639 120 420.8 [a] The surface area and pore size were determined by N 2 adsorption. [b] The amounts of lysozyme adsorbed on the silica particles were determined by UV adsorption at 280 nm (see the Supporting Informa- tion). [*] R. Tian, Prof. Dr. M. Ye, X. Jiang, L. Hu, X. Li, Prof. Dr. H. Zou National Chromatographic R&A Center Dalian Institute of Chemical Physics The Chinese Academy of Sciences Dalian 116023 (China) Fax: (+ 86)411-84379620 E-mail: hanfazou@dicp.ac.cn H. Zhang, Prof. Dr. X. Bao State Key Laboratory of Catalysis Dalian Institute of Chemical Physics (DICP) The Chinese Academy of Sciences Dalian 116023 (China) [**] This work was supported by the National Natural Sciences Foundation of China (No. 20327002), a China State Key Basic Research Program Grant (2005 CB522701), and the Knowledge Innovation Program of DICP to H.Z. Supporting information for this article is available on the WWW under http://www.angewandte.org or from the author. Communications 962 # 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2007, 46, 962 –965