Suresh Kumar Kailasa

Quantum dots laser desorption/ionization MS: multifunctional CdSe quantum dots as the matrix, concentrating probes and acceleration for microwave enzymatic digestion for peptide analysis and high resolution detection of proteins in a linear MALDI-TOF MS

We report the first use of functionalized cadmium selinide quantum dots (CdSe QDs) with 11‐mercaptoundecanoic acid (MUA) as the matrix for the selective ionization of proteins with high resolution and rapid analysis of amino acids and peptides by using quantum dots laser desorption/ionization mass spectrometry (QDLDI‐MS). The mercaptocarboxylic groups of CdSe QDs have been known to be an effective affinity probe to interact with the biomolecules at low abundance level. Using these QDs as the matrix, sensitivity of the method was greatly enhanced and the LOQ of peptides was found to be 100u2005pM with RSD <10%. The QDLDI‐MS is capable for the selective ionization of insulin, lysozyme and myoglobin with high resolution, which is not observed with sinapic acid (SA) as the matrix. The QDLDI‐MS technique offers many advantages for the analysis of amino acids, peptides and proteins with regard to simplicity, rapidity, sensitivity and the mass spectra were generated in the presence of signal suppressors such as urea and Trition X‐100. In addition, the CdSe QDs have been successfully applied as preconcentrating probes for the analysis of digested peptides in lysozyme from chicken egg white by microwave‐assisted enzymatic digestion. This indicates that the QDs are able to absorb radiation from microwave and their ability to trap peptides from microwave‐digested lysozyme. These results demonstrate that the CdSe QDs are promising candidates for the selective ionization of the analytes with an accurate platform to the rapid screening of biomolecules.

Multifunctional ZrO2 nanoparticles and ZrO2-SiO2 nanorods for improved MALDI-MS analysis of cyclodextrins, peptides, and phosphoproteins

AbstractMultifunctional ZrO2 nanoparticles (NPs) and ZrO2-SiO2 nanorods (NRs) have been successfully applied as the matrices for cyclodextrins and as affinity probes for enrichment of peptides (leucine-enkephalin, methionine-enkephalin and thiopeptide), phosphopeptides (from tryptic digestion products of β-casein) and phosphoproteins from complex samples (urine and milk) in atmospheric pressure matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) and MALDI time-of-flight (TOF) MS. The results show that the ZrO2 NPs and ZrO2-SiO2 NRs can interact with target molecules (cyclodextrins, peptides, and proteins), and the signal intensities of the analytes were significantly improved in MALDI-MS. The maximum signal intensities of the peptides were obtained at pH 4.5 using ZrO2 NPs and ZrO2-SiO2 NRs as affinity probes. The limits of detection of the peptides were found to be 75-105xa0fmol for atmospheric pressure MALDI-MS and those of the cyclodextrins and β-casein were found to be 7.5-20 and 115-125xa0fmol, respectively, for MALDI-TOF-MS. In addition, these nanomaterials can be applied as the matrices for the analysis of cyclodextrins in urine samples by MALDI-TOF-MS. ZrO2 NPs and ZrO2-SiO2 NRs efficiently served as electrostatic probes for peptide mixtures and milk proteins because 2–11 times signal enhancement can be achieved compared with use of conventional organic matrices. Moreover, we have successfully demonstrated that the ZrO2 NPs can be effectively applied for enrichment of phosphopeptides from tryptic digestion of β-casein. Comparing ZrO2 NPs with ZrO2-SiO2 NRs, we found that ZrO2 NPs exhibited better affinity towards phosphopeptides than ZrO2-SiO2 NRs. Furthermore, the ZrO2 and ZrO2-SiO2 nanomaterials could be used to concentrate trace amounts of peptides/proteins from aqueous solutions without tedious washing procedures. This approach is a simple, straightforward, separation-and washing-free approach for MALDI-MS analysis of cyclodextrins, peptides, proteins, and tryptic digestion products of phosphoproteins.n FigureMultifunctional ZrO2 nanoparticles and ZrO2-SiO2 nanorods for improved MALDI-MS

High resolution detection of high mass proteins up to 80,000 Da via multifunctional CdS quantum dots in laser desorption/ionization mass spectrometry

CdS quantum dots (∼ 5 nm) are used as multifunctional nanoprobes as an effective matrix for large proteins, peptides and as affinity probes for the enrichment of tryptic digest proteins (lysozyme, myoglobin and cytochrome c) in laser desorption/ionization time-of-flight mass spectrometry (LDI-TOF MS). The use of CdS quantum dots (CdS QDs) as the matrix allows acquisition of high resolution LDI mass spectra for large proteins (5000-80,000 Da). The enhancement of mass resolution is especially notable for large proteins such as BSA, HSA and transferrin (34-49 times) when compared with those obtained by using SA as the matrix. This technique demonstrates the potentiality of LDI-TOF-MS as an appropriate analytical tool for the analysis of high-molecular-weight biomolecules with high mass resolution. In addition, CdS QDs are also used as matrices for background-free detection of small biomolecules (peptides) and as affinity probes for the enrichment of tryptic digest proteins in LDI-TOF-MS.

Surface modified silver selinide nanoparticles as extracting probes to improve peptide/protein detection via nanoparticles-based liquid phase microextraction coupled with MALDI mass spectrometry.

We report the first use of functionalized Ag(2)Se nanoparticles (NPs) as effective extracting probes for NPs-based liquid-phase microextraction (NPs-LPME) to analyze hydrophobic peptides and proteins from biological samples (urine and plasma) and soybean in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Surface modified functional groups such as octadecanethiol (ODT) and 11-mercaptoundecanoic acid (MUA) on Ag(2)Se NPs were found to play an important role for efficient extraction of peptides and proteins from test samples through hydrophobic interactions. The peptides can be efficiently extracted using functionalized Ag(2)Se NPs as extracting probes in the presence of high concentration of matrix interferences such as 4M urea, 0.5% Triton X-100 and 3% NaCl. Ag(2)Se@ODT NPs have shown better extraction efficiency and detection sensitivity for peptides than Ag(2)Se@MUA NPs, bare Ag(2)Se NPs and conventional MALDI-MS. The LODs are 20-68 nM for valinomycin and 100-180 nM for gramicidin D using Ag(2)Se@ODT NPs-LPME in the MALDI-MS. The current approach is highly sensitive and the target analytes can be effectively isolated without sample loss and efficiently analyzed in MALDI-MS.

Nanoparticle-single drop microextraction as multifunctional and sensitive nanoprobes: Binary matrix approach for gold nanoparticles modified with (4-mercaptophenyliminomethyl)-2-methoxyphenol for peptide and protein analysis in MALDI-TOF MS

For the first time, we demonstrated that the nanoparticle-single drop microextraction (NP-SDME) technique can be applied as multifunctional nanoprobes to serve as the binary matrix, affinity and desalting probes for sensitive detection of peptides and proteins in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). This approach introduced a simple method to synthesis gold nanoparticles modified with (4-mercaptophenyliminomethyl)-2-methoxyphenol in toluene to serve as extraction phases in the single drop microextraction (SDME) technique. We successfully applied this multifunctional nanoprobe to extract an array of peptides and proteins at low concentration (fmol range) in MALDI-MS. In addition, for the first time, we demonstrated that the two matrix system (binary matrix) by simply mixing the organic matrix with the Au NP-SDME microdroplets, can offer high sensitivity detection of peptides and proteins. Signal intensity can be significantly enhanced up to 35-fold for a hydrophobic peptide (gramicidin D) comparing to that of the traditional organic matrix. Finally, current approach could effectively analyze milk proteins from milk samples even under high salt conditions (5M NaCl). We believe that the interplay of nanoparticles with SDME as multifunctional probes for MALDI-MS analysis of peptides and proteins could be a powerful tool for future proteomic studies.

Surface modified BaTiO3 nanoparticles as the matrix for phospholipids and as extracting probes for LLME of hydrophobic proteins in Escherichia coli by MALDI–MS

In this paper, we report the dual function of 12-hydroxy octadecanoic acid (HOA)-modified barium titanate nanoparticles (BaTiO3 NPs) as the matrix for phospholipids (PLs) and as hydrophobic affinity probes for liquid-liquid microextraction (LLME) of hydrophobic proteins in Escherichia coli prior to their identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-MS). FT-IR, SEM and TEM were used for the characterization of the HOA-modified BaTiO3 NPs. The surface modified BaTiO3 NPs acted as multifunctional probes (as extracting probes and as the matrix) for the analysis of PLs by MALDI-MS. Compared to 2,5-dihydroxybenzoic acid (2,5-DHB), the HOA-modified BaTiO3 NPs provided good PLs mass spectra with similar or improved signal-to-noise (S/N) ratio, which demonstrated the potentiality of HOA-modified BaTiO3 NPs as a PLs purpose matrix. This method was found to be linear in concentration ranges of 1.0-5.0 μM and 1.0-10.0 μM for L-A-phosphatidyl-l-serine (PS) and L-A-phsophatidic acid sodium (PA) with correlation coefficient (R(2)) values from 0.9905 to 0.9987. The detection limits were 0.20-0.35 μM and 0.25-0.40 μM for PS and PA, respectively. We also demonstrated the HOA-modified BaTiO3 NPs as extracting and as preconcentrating probes for the LLME of hydrophobic proteins in E. coli prior to their identification by MALDI-MS. Thus, the surface modified BaTiO3 NPs-assisted LLME coupled with MALDI-MS provides a simple methodology for the efficient extraction and determination of hydrophobic molecules in biological samples.

Functionalized quantum dots with dopamine dithiocarbamate as the matrix for the quantification of efavirenz in human plasma and as affinity probes for rapid identification of microwave tryptic digested proteins in MALDI-TOF-MS☆☆☆

Functionalized quantum dots with dopamine dithiocarbamate (QDs-DDTC) were utilized for the first time as an efficient material for the quantification of efavirenz in human plasma of HIV infected patients and rapid identification of microwave tryptic digest proteins (cytochrome c, lysozyme and BSA) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The synthesized QDs-DDTC was characterized by using spectroscopic (UV-visible, FT-IR and (1)H NMR) and microscopic (SEM and TEM) techniques. Functionalized QDs-DDTC exhibited a high desorption/ionization efficiency for the rapid quantification of small molecules (efavirenz, tobramycin and aspartame) at low-mass region. QDs-DDTC has well ability to trap target species, and capable to transfer laser energy for efficient desorption/ionization of analytes with background-free detection. The use of QDs-DDTC as a matrix provided good linearity for the quantification of small molecules (R(2)=~0.9983), with good reproducibility (RSD<10%), in the analysis of efavirenz in the plasma of HIV infected patients by the standard addition method. We also demonstrated that the use of functionalized QDs-DDTC as affinity probes for the rapid identification of microwave tryptic digested proteins (cytochrome c, lysozyme and BSA) by MALDI-TOF-MS. QDs-DDTC-based MALDI-TOF-MS approach provides simplicity, rapidity, accuracy, and precision for the determination of efavirenz in human plasma of HIV infected patients and rapid identification of microwave tryptic digested proteins. This new material presents a marked advance in the development of matrix-free mass spectrometric methods for the rapid and precise quantitative determination of a variety of molecules. This article is part of a Special Issue entitled: Proteomics: The clinical link.

Cysteine‐capped ZnSe quantum dots as affinity and accelerating probes for microwave enzymatic digestion of proteins via direct matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometric analysis

Fluorescent semiconductor quantum dots (QDs) exhibit great potential and capability for many biological and biochemical applications. We report a simple strategy for the synthesis of aqueous stable ZnSe QDs by using cysteine as the capping agent (ZnSe-Cys QDs). The ZnSe QDs can act as affinity probes to enrich peptides and proteins via direct matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) analysis. This nanoprobe could significantly enhance protein signals (insulin, ubiquitin, cytochrome c, myoglobin and lysozyme) in MALDI-TOFMS by 2.5-12 times compared with the traditional method. Additionally, the ZnSe-Cys QDs can be applied as heat absorbers (as accelerating probes) to speed up microwave-assisted enzymatic digestion reactions and also as affinity probes to enrich lysozyme-digested products in MALDI-TOFMS. Furthermore, after the enrichment experiments, the solutions of ZnSe-Cys QDs mixed with proteins can be directly deposited onto the MALDI plates for rapid analysis. This approach shows a simple, rapid, efficient and straightforward method for direct analysis of proteins or peptides by MALDI-TOFMS without the requirement for further time-consuming separation processes, tedious washing steps or laborious purification procedures. The present study has demonstrated that ZnSe-Cys QDs are reliable and potential materials for rapid, selective separation and enrichment of proteins as well as accelerating probes for microwave-digested reactions for proteins than the regular MALDI-MS tools. Additionally, we also believe that this work may also inspire investigations for applications of QDs in the field of MALDI-MS for proteomics.

Semiconductor cadmium sulphide nanoparticles as matrices for peptides and as co‐matrices for the analysis of large proteins in matrix‐assisted laser desorption/ionization reflectron and linear time‐of‐flight mass spectrometry

The use of semiconductor cadmium sulphide nanoparticles (CdS NPs) capped with 4-aminothiophenol (ATP) and 11-mercaptoundecanoic acid (MUA) is described for the first time as matrices and as co-matrices for the analysis of peptides and proteins in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). UV-visible spectroscopy, Fourier transform infrared (FT-IR) spectroscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were applied for the characterization of functionalized CdS NPs. The synthesized CdS-ATP and CdS-MUA NPs exhibit uniform size distribution with diameter of 15-25 nm and 20-30 nm, respectively. The -NH(2) (ATP) and -COOH (MUA) groups modified on the surfaces of CdS NPs provide ionizable moieties for efficient transfer of protons during the desorption/ionization of analytes. The functionalized CdS NPs have desirable properties for the analysis of peptides in reflectron MALDI-TOF-MS with suppressed background noise and increased mass resolution (4-13-fold) in linear MALDI-TOF-MS. The application of CdS-MUA NPs and SA as the co-matrices in MALDI-MS is demonstrated for the analysis of hydrophobic proteins from soybean.

Rapid enrichment of phosphopeptides by BaTiO3 nanoparticles after microwave-assisted tryptic digest of phosphoproteins, and their identification by MALDI-MS

AbstractWe show that BaTiO3 nanoparticles (NPs) can be used as a novel substrate for the rapid enrichment of phosphopeptides from microwave tryptic digests of α-casein and non-fat milk prior to their identification by MALDI-MS. Protein digestion is achieved by microwave tryptic digest for 50xa0s, and the resulting phosphopeptides can be effectively adsorbed on the surfaces of the NPs. The phosphopeptides were selectively detected via MALDI-MS. Digestion, enrichment and detection are accomplished within ∼60xa0min. The method was applied to the indentification of 24 phosphopeptides from α-casein and of 21 phosphopeptides (of the α-casein type) from nonfat milk.n FigureBaTiO3 NPs as affinity probes for the rapid analysis of phosphopeptides by MALDI MS