Serological Exosome Metabolic Biopsy of Hepatocellular Carcinoma via Designed Core-Shell Nanoparticles.

Exosome metabolite-based liquid biopsy is a promising strategy for large-scale application in practical clinics toward precise medicine. Given the current challenges in successive isolation and analysis of exosomes and their metabolites in this field, we established a low-cost, high-throughput, and rapid platform for serological exosome metabolic biopsy of hepatocellular carcinoma (HCC) via designed core-shell nanoparticles. It starts with the efficient extraction of high-quality serum exosomes and exosome metabolic features, based on which significantly obvious sample clusters are observed by unsupervised cluster analysis. The following integration of feature selection and supervised machine learning enables the identification of six key metabolites and achieves high-performance prediction between HCC, liver cirrhosis, and healthy controls. Specifically, both sensitivity and accuracy achieve 100% among any pairwise intergroup discrimination in a blind test. The quality and reliability of six key metabolites are further evaluated and validated by using different machine learning algorithms and pathway exploration. Our platform contributes to the future growth of new liquid biopsy technologies for precision diagnosis and real-time monitoring of HCC, among other conditions.

Multichannel Nanogenerator-Driven Collaborative Metabolic Fingerprint Diagnostic Strategy for Early Screening and Risk Evaluation of Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD), along with its progressive forms nonalcoholic steatohepatitis (NASH) and NASH fibrosis, has emerged as a global health crisis. However, the absence of robust screening and risk evaluation tools contributes to the underdiagnosis of NAFLD. Herein, we reported a multichannel nanogenerator-assisted laser desorption/ionization mass spectrometry (LDI-MS) platform for early screening and risk evaluation of NAFLD. Specifically, titanium oxide nanosheets (TiNS) and covalent-organic framework nanosheets (COFNS) were employed as nanogenerators with excellent optical properties and exhibited efficient desorption/ionization during the LDI-MS process. Only ∼0.025 µL of serum without pretreatments and separation, serum metabolic fingerprints (SMFs) can be extracted within seconds. Notably, integrated SMFs from TiNS and COFNS significantly improved diagnostic performance and achieved the area under the curve (AUC) values of 1.000 with 100% sensitivity and 100% specificity for the validation sets of global diagnosis, early diagnosis, high-risk NASH, and NASH fibrosis evaluation. Additionally, four biomarker panels were identified, and their diagnostic AUC values were more than 0.944. Ultimately, key metabolic pathways indicating the change from simple NAFLD to high-risk NASH and NASH fibrosis were uncovered. This work provided a noninvasive and high-throughput screening and risk evaluation strategy for NAFLD healthcare management, thus contributing to the precise treatment of the NALFD.

High-Throughput Metabolic Pattern Screening Strategy for Early Colorectal and Gastric Cancers Based on Covalent Organic Frameworks-Assisted Laser Desorption/Ionization Mass Spectrometry.

Precise early diagnosis and staging are conducive to improving the prognosis of colorectal cancer (CRC) and gastric cancer (GC) patients. However, due to intrusive inspections and limited sensitivity, the prevailing diagnostic methods impede precisely large-scale screening. In this work, we reported a high-throughput serum metabolic patterns (SMP) screening strategy based on covalent organic frameworks-assisted laser desorption/ionization mass spectrometry (hf-COFsLDI-MS) for early diagnosis and staging of CRC and GC. Notably, 473 high-quality SMP were extracted without any tedious sample pretreatment and coupled with multiple machine learning algorithms; the area under the curve (AUC) value is 0.938 with 96.9% sensitivity for early CRC diagnosis, and the AUC value is 0.974 with 100% sensitivity for early GC diagnosis. Besides, the discrimination of CRC and GC is accomplished with an AUC value of 0.966 for the validation set. Also, the screened-out features were identified by MS/MS experiments, and 8 metabolites were identified as the biomarkers for CRC and GC. Finally, the corresponding disordered metabolic pathways were revealed, and the staging of CRC and GC was completed. This work provides an alternative high-throughput screening strategy for CRC and GC and highlights the potential of metabolic molecular diagnosis in clinical applications.

Designed Directional Growth of Ti-Metal-Organic Frameworks for Decoding Alzheimer's Disease-Specific Exosome Metabolites.

Exosomes, a growing focus for liquid biopsies, contain diverse molecular cargos. In particular, exosome metabolites with valuable information have exhibited great potential for improving the efficiency of liquid biopsies for addressing complex medical conditions. In this work, we design the directional growth of Ti-metal-organic frameworks on polar-functionalized magnetic particles. This design facilitates the rapid synergistic capture of exosomes with the assistance of an external magnetic field and additionally synergistically enhances the ionization of their metabolites during mass spectrometry detection. Benefiting from this dual synergistic effect, we identified three high-performance exosome metabolites through the differential comparison of a large number of serum samples from individuals with Alzheimer's disease (AD) and normal cognition. Notably, the accuracy of AD identification ranges from 93.18 to 100% using a single exosome metabolite and reaches a flawless 100% with three metabolites. These findings emphasize the transformative potential of this work to enhance the precision and reliability of AD diagnosis, ushering in a new era of improved diagnostic accuracy.

Straightforward Creation of Multishell Hollow Hybrids for an Integrated Metabolic Monitoring System in Disease Management.

Multidimensional metabolic analysis has become a new trend in establishing efficient disease monitoring systems, as the constraints associated with relying solely on a single dimension in refined monitoring are increasingly pronounced. Here, coordination polymers are employed as derivative precursors to create multishell hollow hybrids, developing an integrated metabolic monitoring system. Briefly, metabolic fingerprints are extracted from hundreds of serum samples and urine samples, encompassing not only membranous nephropathy but also related diseases, using high-throughput mass spectrometry. With optimized algorithm and initial feature selection, the established combined panel demonstrates enhanced accuracy in both subtype differentiation (over 98.1%) and prognostic monitoring (over 95.6%), even during double blind test. This surpasses the serum biomarker panel (≈90.7% for subtyping, ≈89.7% for prognosis) and urine biomarker panel (≈94.4% for subtyping, ≈76.5% for prognosis). Moreover, after attempting to further refine the marker panel, the blind test maintains equal sensitivity, specificity, and accuracy, showcasing a comprehensive improvement over the single-fluid approach. This underscores the remarkable effectiveness and superiority of the integrated strategy in discriminating between MN and other groups. This work has the potential to significantly advance diagnostic medicine, leading to the establishment of more effective strategies for patient management.

Hollow Core-Shell Metal Oxide Heterojunctions for the Urinary Metabolic Fingerprint-Based Noninvasive Diagnostic Strategy.

Urine is a preferred object for noninvasive diagnostic strategies. Urinary metabolic analysis is speculatively regarded as an ideal tool for screening diseases closely related to the genitourinary system in view of the intimate relationship between metabolomics and phenotype. Herein, we propose a urinary metabolic fingerprint-based noninvasive diagnostic strategy by designing hollow core-shell metal oxide heterojunctions (denoted as MOHs). With outstanding light absorption and electron-hole separation ability, MOHs aid in the extraction of high-performance urine metabolic fingerprints. Coupled with optimized machine learning algorithms, we establish a metabolic marker panel for accurate diagnosis of prostate cancer (PCa), which is the most common malignant tumor of the male genitourinary system, achieving accuracies of 84.72 and 83.33% in the discovery and validation sets, respectively. Furthermore, metabolite variations and related pathway analyses confirm the credibility and change correlation of key metabolic features in PCa. This work tends to advance the noninvasive diagnostic strategy toward clinical realities.

Resol/triblock copolymer composite-guided smart fabrication of carbonized mesopores for efficiently decoding exosomal glycans.

Soft-template carbonized mesopores were developed for the purpose of enriching urinary exosomal glycans through organic-organic self-assembly using block copolymers and resol precursors. With a high surface area of 229 m2 g-1, a small pore size of 3.1 nm, and a significant amount of carbon that specifically interacts with oligosaccharides in glycans, this carbonized mesopore material exhibits high selectivity and low limits of detection (5 ng µL-1) towards glycans. Our analysis of complex urine samples from healthy volunteers and bladder carcinoma patients successfully profiled 48 and 56 exosomal glycans, respectively, and 16 of them were significantly changed. Moreover, one upregulated bisecting N-acetylglucosamine (GlcNAc)-type glycan with core fucose, two upregulated and two downregulated terminal-sialylated glycans were revealed to be linked to bladder carcinoma. This approach is of significant importance for understanding diseases that arise from protein glycosylation mutations, and it may contribute to the development of novel diagnostic and therapeutic strategies for bladder carcinoma.

Advances in proteomics sample preparation and enrichment for phosphorylation and glycosylation analysis.

As the common and significant chemical modifications, post-translational modifications (PTMs) play a key role in the functional proteome. Affected by the signal interference, low concentration, and insufficient ionization efficiency of impurities, the direct detection of PTMs by mass spectrometry (MS) still faces many challenges. Therefore, sample preparation and enrichment are an indispensable link before MS analysis of PTMs in proteomics. The rapid development of functionalized materials with diverse morphologies and compositions provides an avenue for sample preparation and enrichment for PTMs analysis. In this review, we summarize recent advances in the application of novel functionalized materials in sample preparation for phosphoproteomes and glycoproteomes analysis. In addition, this review specifically discusses the design and preparation of functionalized materials based on different enrichment mechanisms, and proposes research directions and potential challenges for proteomic PTMs research.

Multiscale Element-Doped Nanowire Array-Coupled Machine Learning Reveals Metabolic Fingerprints of Nonreversible Liver Diseases.

Timely detection of nonreversible liver diseases contributes greatly to reasonable therapy and quality of life. Given the current situation, minimally invasive high-specificity molecular diagnosis based on body fluid can be a good choice. Herein, a mesoporous superstructure is designed using silicon atom-doped nanowire arrays to uniformly load Pt nanoparticles on the surface to produce a desirable ionization effect. We apply the multiscale element-doped nanowire arrays to efficiently assist extraction of high-quality metabolic fingerprints from only 35 nL of serum within seconds. Using different machine learning algorithms, we establish specific biomarker panels to distinguish different liver diseases from the healthy control, with more than 90% accuracy, sensitivity, and specificity. Moreover, from established biomarker panels, we further determine key metabolites of significant difference (p < 0.01) via group comparison to realize the discrimination of different liver diseases with 100% sensitivity. Our work confirms the design protocol of an advanced diagnosis tool and lays a robust foundation for metabolic molecular diagnosis in large-scale clinical application.

Diagnosing, Typing, and Staging of Renal Cell Carcinoma by Designer Matrix-Based Urinary Metabolic Analysis.

Molecular diagnosing, typing, and staging have been considered to be the ideal alternatives of imaging-based detection methods in clinics. Designer matrix-based analytical tools, with high speed, throughout, efficiency and low/noninvasiveness, have attracted much attention recently for in vitro metabolite detection. Herein, we develop an advanced metabolic analysis tool based on highly porous metal oxides derived from available metal-organic frameworks (MOFs), which elaborately inherit the morphology and porosity of MOFs and newly incorporate laser adsorption capacity of metal oxides. Through optimized conditions, direct high-quality fingerprinting spectra in 0.5 µL of urine are acquired. Using these fingerprinting spectra, we can discriminate the renal cell carcinoma (RCC) from healthy controls with higher than 0.99 of area under the curve (AUC) values (R2Y(cum) = 0.744, Q2 (cum) = 0.880), as well, from patients with other tumors (R2Y(cum) = 0.748, Q2(cum) = 0.871). We also realize the typing of three RCC subtypes, including clear cell RCC, chromophobe RCC (R2Y(cum) = 0.620, Q2(cum) = 0.656), and the staging of RCC (R2Y(cum) = 0.755, Q2(cum) = 0.857). Moreover, the tumor sizes (threshold value is 3 cm) can be remarkably recognized by this advanced metabolic analysis tool (R2Y(cum) = 0.710, Q2(cum) = 0.787). Our work brings a bright prospect for designer matrix-based analytical tools in disease diagnosis, typing and staging.

Tractable Method for Rapid Quality Assessment of Therapeutic Antibodies in Harvested Cell Culture Fluid based on FcγRIIIa-Immobilized Magnetic Microspheres.

FcγRIIIa-binding affinity is one of the key factors to ensure the efficacy of many antitumor therapeutic antibodies, which should be monitored along with the titer, protein aggregation, and other critical quality attributes. The conventional workflow for the quality assessment of therapeutic antibodies in harvested cell culture fluid (HCCF) is time-consuming and costly nevertheless. In this study, a tractable method was established for rapid quality assessment of a HCCF sample through differentially extracting IgG with different FcγRIIIa affinity levels using FcγRIIIa-immobilized magnetic microspheres, followed by size exclusion chromatography (SEC) to determine the amount and monomer percentage of IgGs in the preceding eluate. FcγRIIIa-immobilized magnetic microspheres with polydopamine (PDA) and hydrophilic dendrimer (PAMAM) coating (denoted as Fe3O4@PDA@PAMAM-FcγRIIIa) were synthesized for the first time as magnetic adsorbents. The PDA cladding endowed the composites with good chemical stability in acidic elution buffer, and the PAMAM dendrimer empowered the composites of high ligand immobilization capacity and hydrophilic surface. The labile FcγRIIIa was immobilized under mild conditions. By directly applying a simple magnetic solid phase extraction procedure to treat HCCF, favored IgG species with high FcγRIIIa affinity would be selectively captured by Fe3O4@PDA@PAMAM-FcγRIIIa composites for subsequent SEC analysis. The monomer peak area value in SEC, which was set as the read-out of the proposed method, correlated directly with the theoretical overall quality of standard-spiked HCCF samples.

In Vitro Diagnostic Examination and Prognosis Surveillance by Hierarchical Heterojunction-Assisted Metabolic Analysis.

High-throughput metabolic analysis based on laser desorption/ionization mass spectrometry exhibits broad prospects in the field of large-scale precise medicine, for which the assisted ionization ability of the matrix becomes a determining step. In this work, the gold-decorated hierarchical metal oxide heterojunctions (dubbed Au/HMOHs) are proposed as a matrix for extracting urine metabolic fingerprints (UMFs) of primary nephrotic syndrome (PNS). The hierarchical heterojunctions are simply derived from metal-organic framework (MOF)-on-MOF hybrids, and the native built-in electric field from heterojunctions plus the extra Au decoration provides remarkable ionization efficiency, attaining high-quality UMFs. These UMFs are employed to realize precise diagnosis, subtype classification, and effective prognosis evaluation of PNS by appropriate machine learning, all with 100% accurate ratios. Moreover, a high-confidence marker panel for PNS diagnosis is constructed. Interestingly, all panel metabolite markers present obviously uniform downregulation in PNS compared to healthy controls, shedding light on mechanism exploration and pathway analysis. This work drives the application of metabolomics toward precision medicine.

IL-37 blocks gouty inflammation by shaping macrophages into a non-inflammatory phagocytic phenotype.

OBJECTIVE: Interleukin (IL)-37 is a natural suppressor of inflammation. Macrophages play an important role in acute gout flare by dominating the inflammation and spontaneous relief. We have reported that IL-37 could limit runaway inflammation in gout. Here we focus on whether IL-37 inhibits gouty inflammation by altering macrophage functions, and how it does so. METHODS: Macrophage functions were evaluated in terms of phagocytosis, pyroptosis, polarization and metabolism. Phagocytosis and polarization of macrophages were detected by side scattering and double-labelling induced nitrogen monoxide synthase (iNOS)/arginase-1 (Arg-1) using flow cytometry, respectively. Transcription of pyroptosis-related molecules was detected by qPCR. Metabolomics was performed by liquid chromatograph mass spectrometer. Human IL-37 knock-in mice and a model with point mutation (S9A) at mouse Gsk3b locus were created by CRISPR/Cas-mediated genome engineering. MSU was injected into the paws and peritoneal cavity to model acute gout. Vernier calliper was used to measure the thickness of the paws. The mice paws and human synovium tissues or tophi were collected for pathological staining. Peritoneal fluid of mice was used to enrich macrophages to detect polarization. RESULTS: IL-37 promoted non-inflammatory phagocytic activity of macrophages by enhancing phagocytosis of MSU, reducing transcription of pyroptosis-related proteins and release of inflammatory cytokines, protecting mitochondrial function, and mediating metabolic reprogramming in MSU-treated THP-1 cells. These multifaceted roles of IL-37 were partly depended on the mediation of glycogen synthase kinase-3ß (GSK-3ß). CONCLUSIONS: Our study revealed that IL-37 could shape macrophages into a 'silent' non-inflammatory phagocytic fashion. IL-37 may become a potentially valuable treatment option for patients of chronic gout, especially for those with tophi.

Mesoporous NiO@ZnO nanofiber membranes via single-nozzle electrospinning for urine metabolism analysis of smokers.

An effective matrix is very important for impact laser desorption/ionization mass spectrometry (LDI-MS), and the physicochemical properties of the matrix nanostructures can impact the LDI-MS performance. In this study, a simple and efficient single-nozzle electrospinning strategy using polystyrene (PS) spheres and polyvinyl pyrrolidone (PVP) to construct a mesoporous NiO@ZnO nanofiber membrane was developed. Compared with the NiO and ZnO nanomaterials alone, the obtained NiO@ZnO nanofiber membrane was proven to be an efficient material as the matrix to increase the intensity of the mass spectrum speaks of small molecules. The NiO@ZnO nanofiber membrane was used as the matrix for the LDI-MS method for the urine metabolism analysis of smokers, which revealed differences in the metabolic and the possible metabolic markers of smokers through the statistical analysis of the urine samples of 27 smokers and 11 nonsmoker controls.

Magnetic porous carbon-dependent platform for the determination of N-glycans from urine exosomes.

A magnetic porous carbon-dependent platform is established to separate and determine N-glycans from urine exosomes of healthy people and patients with gastric cancer. The results of the comparison reveal that 6 N-glycans shared by the two groups are downregulated, most of which present core fucose or bisecting N-acetylglucosamine (GlcNAc) type. In addition, five shared N-glycans including two of sialic acid type are upregulated. These obvious differences indicate the close relationship between glycans and gastric cancer thus permitting early diagnosis. A magnetic porous carbon material (FeMPC) from MIL-101(Fe) was employed to separate and analyze N-glycans from urine exosomes of healthy people and patients with gastric cancer.

Boric-acid-modified Fe3O4@PDA@UiO-66 for enrichment and detection of glucose by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Herein, boric-acid-modified multifunctional Zr-based metal-organic frameworks (denoted as Fe3O4@PDA@B-UiO-66) were synthesized by hydrothermal reaction and surface modification. Compared to traditional matrix, Fe3O4@PDA@B-UiO-66 has the advantages of high ionization efficiency, high surface area, low matrix background, porous structure, and numerous boric-acid-active sites. By combining matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), Fe3O4@PDA@B-UiO-66 was used as an adsorbent and matrix for enrichment and detection of glucose, based on a specific reaction between boric acid and glucose. The limit of detection was 58.5 nM. The proposed method provides a simple and efficient approach for the sensitive and quantitative detection of glucose in complex samples based on MALDI-TOF MS. Design and synthesis of boric-acid-modified multifunctional magnetic metal-organic frameworks (designated as Fe3O4@PDA@B-UiO-66) applied as adsorbent and matrix for the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis of glucose in complex biosamples.

Recognition of urinary N-linked glycopeptides in kidney cancer patients by hydrophilic carbohydrate functionalized magnetic metal organic framework combined with LC-MS/MS.

A hydrophilic carbohydrate functionalized magnetic metal organic framework (Mag Zr-MOF@G6P) was synthesized via a facile one-step modification strategy for selective glycopeptide capture in virtue of hydrophilic interaction chromatography technique. The inherently hydrophilic Zr-MOF layer not only provides selective size-sieving pore structures but also offers large specific surface area to afford abundant affinity sites. Hydroxyl-rich glucose-6-phosphate was immobilized onto the Zr-MOF via a straightforward coordination manner to regulate its surface property, for the purpose of enhancing its hydrophilicity. Benefitting from the merits of Zr-MOF and glucose-6-phosphate, the as-designed composite exhibits good selectivity (the mass ratio of HRP digests to BSA digests was up to1:200) and low limit of detection (0.1 fmol µL-1) towards the recognition of glycopeptides from standard samples. More excitingly, glycopeptides in urine of healthy people and patients with kidney cancer were successfully enriched and identified by the combined liquid chromatography-mass spectrometry/mass spectrometry technology (LC-MS/MS). Further gene ontology analysis of molecular function and biological process revealed that 13 original glycoproteins of the identified glycopeptides from urine of patients significantly participate in diverse cancer-associated events, including collagen binding, immunoglobulin receptor binding, antigen binding, and complement activation process. Graphical abstract.

Dual metal cations coated magnetic mesoporous silica probe for highly selective capture of endogenous phosphopeptides in biological samples.

For the first time, dual metal ions (Ti4+-Zr4+) were successfully modified into the channel of magnetic mesoporous silica to obtain an affinity probe for highly selective capture of endogenous phosphopeptides in biological samples. The newly prepared Fe3O4@mSiO2@Ti4+-Zr4+ composites possessed the advantages of ordered mesoporous channels, superparamagnetism, and enhanced affinity properties of dual metal ions of Ti4+ and Zr4+. The phosphopeptide enrichment efficiency of the Fe3O4@mSiO2@Ti4+-Zr4+ composite was investigated, and the result indicated an ultrahigh size exclusive ability (weight ratio of ß-casein tryptic digests, BSA, and α-casein protein reached up to 1:1000:1000). Compared to magnetic affinity probes with single metal ions (Fe3O4@mSiO2@Ti4+, Fe3O4@mSiO2@Zr4+), the composite possessed stronger specificity, higher sensitivity, and better efficiency; and more importantly, it showed much enhanced enrichment ability towards both mono- and multi-phosphorylated peptides. Additionally, by utilizing the Fe3O4@mSiO2@Ti4+-Zr4+ affinity probe, a total number of 104 endogenous phosphopeptides including 95 mono-phosphopeptides and 9 multi-phosphopeptides were captured and identified from human saliva, indicating the great potential for the application of the novel probe for the peptidome analysis in the future. Graphic abstract.

Immobilization of titanium dioxide/ions on magnetic microspheres for enhanced recognition and extraction of mono- and multi-phosphopeptides.

The authors are presenting a novel strategy for global phosphoproteome recognition in practical samples. It integrates metal oxide affinity chromatography (MOAC) and immobilization metal ion affinity chromatography (IMAC). This resulted in a kind of titanium dioxide/ion-based multifunctional probe (dubbed T2M). The T2M combines the features of MOAC and IMAC including their recognition preferences towards mono- and multi-phosphorylated peptides. Hence, they exhibit an outstanding recognition capability towards global phosphoproteome, high sensitivity (the limit of detection of which is merely 10 fmol) and excellent specificity in MALDI-TOF MS detection. Their performance is further demonstrated by the identification of the phosphoproteome in non-fat milk and human saliva. By combining T2M with nano LC-MS/MS, remarkable results are obtained in the tryptic digestion of healthy eye lens and cataract lens phosphoproteomes. A total of 658 and 162 phosphopeptides, respectively, were identified. This indicates that phosphorylation and the appearance of cataract can be related to each other. Graphical abstract Schematic presentation of the preparation of titanium dioxide/ion-based multifunctional magnetic nanomaterials (T2M). The T2M based enrichment protocol exhibits outstanding recognition capability towards global phosphoproteome. This protocol shows great prospect for clarifying mechanism of phosphorylation-related diseases via further information acquisition.

Magnetite nanoparticles coated with mercaptosuccinic acid-modified mesoporous titania as a hydrophilic sorbent for glycopeptides and phosphopeptides prior to their quantitation by LC-MS/MS.

A hydrophilic material consisting of a magnetite core coated with mercaptosuccinic acid modified mesoporous titania (denoted as Fe3O4@mTiO2-MSA) has been fabricated. It is shown to be a viable sorbent for capturing glycopeptides and phosphopeptides. The sorbent combines the features of metal oxide-based affinity chromatography and of hydrophilic interaction liquid chromatography (HILIC) with the advantages of using mesoporous titania. The use of magnetic microspheres provides magnetic response and simplifies separation. Following elution with 10% ammonia, the peptides were submitted to LC-MS/MS analysis. The method enabled 327 phosphopeptides and 65 glycopeptides to be identified in three isolated replicates of merely 5 µL samples of human saliva. Among them, the phosphorylation sites and glycosylation sites were detected in 20 peptide segments. Graphical abstract Schematic presentation of preparation of novel hydrophilic magnetic mesoporous titania nanomaterials (Fe3O4@mTiO2-MSA). This specific sorbent exhibits highly selective and efficient simultaneous adsorption ability for both glycopeptides and phosphopeptides from biosamples by mass spectrometric analysis.