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The reasonable design of metal-organic framework (MOF)-derived nanomaterial has important meaning in increasing the enrichment efficiency in the study of protein phosphorylation. In this work, a polyoxometalate (POM) functionalized magnetic MOF nanomaterial (Fe 3 O 4 @MIL-125-POM) was designed and fabricated. The nanomaterial with multi-affinity sites (unsaturated metal sites and metal oxide clusters) was used for the enrichment of phosphopeptides. Fe 3 O 4 @MIL-125-POM had high-efficient enrichment performance towards phosphopeptides (selectivity, a mass ratio of bovine serum albumin/α-casein/β-casein at 5000:1:1; sensitivity, 0.1 fmol; satisfactory repeatability, ten times). Furthermore, Fe 3 O 4 @MIL-125-POM was employed to enrich phosphopeptides from non-fat milk digests, saliva, serum, and A549 cell lysate. The enrichment results illustrated the great potential of Fe 3 O 4 @MIL-125-POM for efficient identification of low-abundance phosphopeptides., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

For mass spectrometry (MS)-based phosphoproteomics studies, sample pretreatment is an essential step for efficient identification of low-abundance phosphopeptides. Herein, a cobalt phthalocyanine-modified magnetic metal-organic framework (MOF) (Fe 3 O 4 @MIL-101-CoPc) was prepared and applied to enrich phosphopeptides before MS analysis. Fe 3 O 4 @MIL-101-CoPc exhibited an excellent magnetic response (74.98 emu g -1 ) and good hydrophilicity (7.75°), which were favorable for the enrichment. Fe 3 O 4 @MIL-101-CoPc showed good enrichment performance with high selectivity (1:1:5000), sensitivity (0.1 fmol), reusability (10 circles), and recovery (91.3%). Additionally, the Fe 3 O 4 @MIL-101-CoPc-based MS method was able to successfully detect 827 phosphopeptides from the A549 cell lysate, demonstrating a high enrichment efficiency (89.3%). This study promotes the application of postfunctionalized MOFs for phosphoproteomics analysis.

Polyoxometalate-based metal-organic frameworks (POMOFs) have become a promising affinity material for separation and enrichment. The analysis of protein phosphorylation represents a challenge for the development of efficient enrichment materials. Here, a novel zirconium-rich magnetic POMOF was successfully designed and prepared for the enrichment of phosphopeptides. The binding affinity of the nanomaterial partly came from Fe-O clusters in the MOF. The Lewis acid-base interactions between V-O clusters and zirconium ions in V 10 O 28 -Zr 4+ and phosphate groups in phosphopeptides further strengthened the enrichment ability. The zirconium-rich magnetic POMOF was employed to capture phosphopeptides from non-fat milk, human saliva, and serum. Additionally, 748 unique phosphopeptide peaks were detected from the tryptic digests of lung cancer A549 cell proteins with a high specificity (86.9 %). POMOFs will become an active competitor for the design of protein affinity materials and will provide a new approach for phosphopeptide analysis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Highly selective extraction of phosphopeptides is necessary before mass spectrometry (MS) analysis. Herein, zirconium phthalocyanine-modified magnetic nanoparticles were prepared through a simple method. The Fe-O groups on Fe 3 O 4 and the zirconium ions on phthalocyanine had a strong affinity for phosphopeptides based on immobilized metal ion affinity chromatography (IMAC). The enrichment platform exhibited low detection limit (0.01 fmol), high selectivity (α-/β-casein/bovine serum albumin, 1/1/5000), good reusability (10 circles), and recovery (91.1 ± 1.1%) toward phosphopeptides. Nonfat milk, human serum, saliva, and A549 cell lysate were employed as actual samples to assess the applicability of the enrichment protocol. Metallo-phthalocyanine will be a competitive compound for designing highly efficient adsorbents and offers a new approach to phosphopeptide analysis.

In this study, magnetic polyoxometalate-based metal-organic frameworks (Fe 3 O 4 -POMOFs) were designed and applied to the enrichment of phosphopeptides. Thanks to the abundant metal oxide and metal ion sites, the material had a strong affinity for phosphopeptides. Simultaneously, the high amount of amino and guanidyl groups provided hydrophilicity and positive charge for phosphopeptide capture. By coupling with MS detection, the established platform possessed good reusability, high sensitivity (0.01 fmol), and high selectivity (α-casein/β-casein/bovine serum albumin = 1:1:5000). Furthermore, the method was successfully used for the detection of phosphopeptides in nonfat milk, human serum, saliva, and A549 cell lysate, showing great potential for practical application.

A layer-by-layer (LbL) assembly strategy was developed to prepare multilayered double hydroxide/polyoxometalate shell-coated magnetic nanoparticles. The introduction of functional shells not only offered abundant affinity sites of metal oxide and metal ions but also increased the surface area for the contact with targets. By combining the enrichment strategies of immobilized metal ion affinity chromatography and metal oxide affinity chromatography, the nanomaterial can capture phosphopeptides via a synergistic effect. The method presented a low detection limit of 0.1 fmol in combination with matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) analysis. The nanomaterial showed satisfactory selectivity (1:1:5000 M ratio of α-/β-casein/bovine serum albumin), good recovery (92.07%), high adsorption capacity (117.6 mg g -1 ), and ten times reusability for capturing phosphopeptides., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Exfoliated magadiite nanosheets embedded with Fe 3 O 4 were constructed. Advantage was taken of the strong coordination between the silanol groups in magadiite nanosheets and the Gd 3+ ion to prepare the final adsorbent, Gd 3+ -immobilized magnetic magadiite nanosheets. The adsorbent with two-dimensional (2D) morphology offered high surface area and abundant Gd 3+ contents for phosphopeptides enrichment, on which Fe 3 O 4 with positive electricity incorporated the magnetic properties. Combining with matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI TOF-MS), the method showed low detection limit (0.05 fmol). The feasibility of using the 2D nanocomposite for phosphopeptides enrichment was demonstrated using mixtures of β-casein and bovine serum albumin (1:5000). The standard deviation of captured phosphopeptides in three repeated experiments were in the range 0.15-0.42 (< 0.5% RSD). Further evaluation revealed that the nanocomposite was capable of enriching phosphopeptides from non-fat milk, human saliva, and serum. A novel Gd 3+ -immobilized two-dimensional magnetic magadiite nanosheets-based enrichment platform was designed. The developed material was employed as the adsorbent for the selective enrichment of phosphopeptides by coupling with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The material was successfully applied to enrich phosphopetides from standard peptide mixtures, nonfat milk, human saliva, and serum., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Efficient enrichment and identification of phosphopeptides are of great significance in biological applications. Glycocyamine functionalized magnetic layered double hydroxides (Fe 3 O 4 @LDH@NH 2 -GAA) was fabricated through an easy process. The magnetic composite possessed high surface area, good biocompatibility, and fast magnetic response. Fe 3 O 4 @LDH@NH 2 -GAA, combining not only metal ions (Cu 2+ and Ga 3+ ) in LDH but also functional guanidyl groups in glycocyamine, offered multiple affinity sites for phosphopeptides enrichment. With these favorable characters, it exhibited high selectivity (β-casein: bovine serum albumin = 1:5000), low detection limit (0.1 fmol), satisfactory enrichment recovery (94.5%), high adsorption capacity (82.4 mg g -1 ), and good repetitiveness. Moreover, the efficient enrichment of phosphopeptides by Fe 3 O 4 @LDH@NH 2 -GAA from nonfat milk, human saliva, serum, and A549 cell lysates further confirmed its great potential for trace biological detection and proteomic analysis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Highly efficient enrichment of phosphopeptides in low abundance prior to mass spectrometry analysis is essential in phosphopeptidomics analysis. Herein, an affinity probe possessing both interactions of polyoxometalates (POMs) and hydrophilic interaction chromatography (HILIC) was facilely prepared. POMs and cysteamine hydrochloride-modified chitosan (CYECS) were assembled onto magnetic nanoparticles (MNPs) via a layer-by-layer approach. The developed MNPs-(POM/CYECS) material owned merits of large metal oxide surfaces, positive charge, and hydrophilicity as well as superparamagnetism. By combining with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), the prepared material was utilized for effective enrichment of phosphopeptides. High selectivity (mass ratio of β-casein and bovine serum albumin digests of 1:5000) and low detection limit (0.02 fmol) toward phosphopeptides were achieved. The recovery of phosphopeptides was measured to be 92.6%. The specific enrichment of phosphopeptides from complex samples (nonfat milk, human saliva, serum, and A549 cell lysate) with MNPs-(POM/CYECS) material further confirmed its bright prospects for isolation of phosphopeptides from biological samples., (Copyright © 2019 Elsevier B.V. All rights reserved.)

A method for the enrichment of phosphopeptides based on magnetic nanoparticles coated with polyoxometalate (POM) was developed. A layer-by-layer assembly technique was used to prepare magnetic nanoparticles (MNPs) coated with polyoxometalate and chitosan. The composites were used for phosphopeptide enrichment prior to analysis by matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS). The composites exhibited advantages of rapid magnetic separation, hydrophilicity, positive electricity, and a high selectivity for enrichment of phosphopeptides. β -Casein was selected as a model phosphorylated protein to assess the performance of the enrichment technique. After the enrichment step, the limit of detection was determined to be 0.02 fmol. The prepared POM-based MNPs thus exhibit significant potential in the detection of low-abundance phosphoproteins.

Dendrimers exhibit tunable terminal functionality and bio-friendly nature, making them of being promising materials for applications in the field of separation and enrichment. In this work, we prepared magnetic hydrazide-functionalized poly-amidoamine (PAMAM) dendrimer embedded with TiO 2 for the enrichment of phosphopeptides. The novel affinity probe possessed superparamagnetism, realizing its rapid separation from sample solution. Electrostatic attraction and hydrogen bonding existed between PAMAM and phosphopeptides while Lewis acid-base interaction was originated between TiO 2 and the targets. The combined synergistic strength of multiple binding interactions contributed to the highly selective enrichment of phosphopeptides. The specificity for the capture of phosphopeptides was reflected in quantities as low as 1:1000 mass ratio of phosphopeptides to non-phosphopeptides. The detection limit of β-casein digests was low to 0.4 fmol, indicating the high sensitivity of the developed method. Fifteen and four phosphopeptides could be selectively captured from non-fat milk digests and human serum samples, which further confirmed the great potential of the affinity probe in the extraction of low-abundance phosphopeptides from real complex biological samples., (Copyright © 2018 Elsevier B.V. All rights reserved.)

In this study, we designed a gadolinium-based immobilized metal ion affinity chromatography material for the selective enrichment of phosphopeptides. Gadolinium ion was immobilized on the surface of glutathione-coated magnetic nanoparticles through a facile and effective synthetic route. The adsorbent integrated the advantages of superparamagnetism of Fe 3 O 4 core, good biological compatibility of glutathione, and strong interaction between gadolinium ion and phosphopeptides. It was employed to enrich phosphopeptides from standard protein digests coupled with MALDI-TOF MS. Results demonstrated that the adsorbent possessed high selectivity for phosphopeptides, good reusability and reproducibility. Moreover, the material provided selective enrichment of phosphopeptides from real samples including non-fat milk digests and human serum. The developed method exhibited high sensitivity (detection limit of 10 fmol), showing great potential in the detection of low-abundance phosphopeptides in biological samples., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Selective and efficient preconcentration is indispensable for low concentration of phosphopeptides in phosphorylated protein-related samples prior to MS-based analysis. Herein, an on-chip system coupled magnetic SPE with MALDI-TOF MS was designed. A metal oxide affinity chromatography material, indium oxide, was coated on the surface of Fe 3 O 4 magnetic nanoparticles to prepare the adsorbent, spatially confined with an applied magnetic field. The adsorbent exhibited high selectivity for phosphopeptides in tryptic digests of the mixture of β-casein and BSA (1:1000) and the mixture of β-casein, BSA, and ovalbumin (1:100:100). Thanking to the enrichment ability and specificity for phosphopeptides with the adsorbent, the on-chip magnetic SPE-MALDI-TOF MS approach showed high sensitivity with a low detection limit of 4 fmol. In addition, the developed approach was used to analyze phosphopetides in non-fat milk digests and human serum successfully., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Cerium ions immobilized magnetic graphite nitride material have been prepared using L-Alanyl-L-Glutamine as the new chelator. The resulting Fe3O4/g-C3N4-L-Ala-L-Gln-Ce4+, as an immobilized metal ion affinity chromatography (IMAC) sorbent, was reusable. This is due to the strong coordination interaction between L-Alanyl-L-Glutamine and cerium ions. After a series of characterizations, the magnetic nanocomposite showed high surface area, good hydrophilicity, positive electricity, and magnetic response. Fe3O4/g-C3N4-L-Ala-L-Gln-Ce4+ had high sensitivity (0.1 fmol), selectivity (α-/β-casein/bovine serum albumin, 1:1:5000), and good recyclability (10 cycles). A total of 647 unique phosphopeptides mapped to 491 phosphoproteins were identified from A549 cell lysate by nano LC–MS analysis. [ABSTRACT FROM AUTHOR]

In the work, we developed polyoxometalate modified magnetic metal–organic framework and investigated its enrichment performance for phosphopeptides. The magnetic nanocomposite can interact with phosphopeptides through Zr−O clusters provided by the MOF shell and metal oxide originated from Zr-W-POM. MALDI-TOF MS and nano LC-MS were employed for phosphopeptide determinations. The magnetic nanocomposite had high sensitivity, good recyclability, and high selectivity. Non-fat milk, drinking milk, human saliva, and serum were selected as real samples for exploration the applicability of the magnetic nanocomposite. [Display omitted] • Synthesis of Fe 3 O 4 @PDA@MOF-POM through a simple post-modification method. • Application of Fe 3 O 4 @PDA@MOF-POM for phosphopeptide enrichment. • High enrichment specificity and selectivity of Fe 3 O 4 @PDA@MOF-POM. • High enrichment efficiency in non-fat milk, drinking milk, human serum, saliva. Herein, polyoxometalate modified magnetic metal–organic framework (Fe 3 O 4 @PDA@MOF-POM) was successfully prepared and then applied to the enrichment of phosphopeptides. Fe 3 O 4 @PDA@MOF-POM can bind with phosphopeptides through Zr O clusters provided by the MOF layer and metal oxide originated from Zr-W-POM. The magnetic nanomaterial had high sensitivity (0.1 fmol), good reusability (10 cycles), and high selectivity (α -casein: β -casein: bovine serum albumin = 1: 1: 5000). Coupling with MALDI-TOF MS determination, non-fat milk was selected to preliminarily explore the applicability of Fe 3 O 4 @PDA@MOF-POM. Furthermore, 241 phosphopeptides derived from 232 phosphoproteins and 99 phosphopeptides attributed to 89 phosphoproteins were identified in human serum and saliva by nano LC-MS. This work paves a new way for the recognition of phosphopeptides in bio-samples. [ABSTRACT FROM AUTHOR]

Exfoliated magadiite nanosheets embedded with Fe3O4 were constructed. Advantage was taken of the strong coordination between the silanol groups in magadiite nanosheets and the Gd3+ ion to prepare the final adsorbent, Gd3+-immobilized magnetic magadiite nanosheets. The adsorbent with two-dimensional (2D) morphology offered high surface area and abundant Gd3+ contents for phosphopeptides enrichment, on which Fe3O4 with positive electricity incorporated the magnetic properties. Combining with matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI TOF-MS), the method showed low detection limit (0.05 fmol). The feasibility of using the 2D nanocomposite for phosphopeptides enrichment was demonstrated using mixtures of β-casein and bovine serum albumin (1:5000). The standard deviation of captured phosphopeptides in three repeated experiments were in the range 0.15–0.42 (< 0.5% RSD). Further evaluation revealed that the nanocomposite was capable of enriching phosphopeptides from non-fat milk, human saliva, and serum. [ABSTRACT FROM AUTHOR]