Phospholipids are key components of cellular membrane and signaling. Among cellular phospholipids, phosphoinositides, phosphorylated derivatives of phosphatidylinositol are important as a participant in essential metabolic processes in animals. However, due to its low abundance in cells and tissues, it is difficult to identify the composition of phosphoinositides. Recent advances in mass spectrometric techniques, combined with established separation methods, have allowed the rapid and sensitive detection and quantification of a variety of lipid species including phosphoinositides. In this mini review, we briefly introduce progress in profiling of cellular phosphoinositides using mass spectrometry. We also summarize current progress of matrices development for the analysis of cellular phospholipids using matrix-assisted laser desorption/ionization mass spectrometry. The phosphoinositides profiling and phospholipids imaging will help us to understand how they function in a biological system and will provide a powerful tool for elucidating the mechanism of diseases such as diabetes, cancer and neurodegenerative diseases. The investigation of cellular phospholipids including phosphoinositides using electrospray ionization mass spectrometry and matrix-assisted laser desorption/ionization mass spectrometry will suggest new insights on human diseases, and on clinical application through drug development of lipid related diseases.
Animals ; Humans ; Mass Spectrometry/*methods ; Phosphatidylinositols/*metabolism ; Phospholipids/*metabolism ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

The amino acid sequence of the fusion protein FP3 was measured by two types of LC-MS/MS and its primary structure was confirmed. After reduction and alkylation, the protein was digested with trypsin and glycosyl groups in glycopeptide were removed by PNGase F. The mixed peptides were separated by LC, then Q-TOF and Ion trap tandem mass spectrometry were used to measure b, y fragment ions of each peptide to analyze the amino acid sequence of fusion protein FP3. Seventy-six percent of full amino acid sequence of the fusion protein FP3 was measured by LC-ESI-Q-TOF with the remaining 24% completed by LC-ESI-Trap. As LC-MS and tandem mass spectrometry are rapid, sensitive, accurate to measure the protein amino acid sequence, they are important approach to structure analysis and identification of recombinant protein.
Amino Acid Sequence ; Chromatography, High Pressure Liquid ; Molecular Sequence Data ; Peptide Mapping ; Recombinant Fusion Proteins ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Tandem Mass Spectrometry ; Vascular Endothelial Growth Factor A ; antagonists & inhibitors ; chemistry

The alkaloids in Zanthoxylum nitidum were identified by HPLC-DAD/ESI-Q-TOF-MS. Separation was performed on a Hanbon C18 column with acetonitrile (with 0.1% formic acid) and water(with 0.1% formic acid) as mobile phase. Based on the high-resolution mass information, MS/MS fragmentation behaviors and chemical components from literatures, 48 components were identified or tentatively characterized including 6 new compounds. This work could be useful for the quality control and further studies of the plant.
Alkaloids ; chemistry ; Chromatography, High Pressure Liquid ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Zanthoxylum ; chemistry

Stem cell factor is an important hematopoietic growth factor. In this study, the human stem cell factor was produced by recombinant E. coli, and the structure and biological activity of the recombinant stem cell factor(rhSCF) was studied. It was indicated that the rhSCF was a uncovalent dimer in phosphate buffer,and had the correct mass spectra, mass peptides spectra, composition of amino acid, N-terminal sequernce, C-terminal sequence and intrachain disulfide linkages, rhSCF alone or synergy with rhG-CSF could mobilze hematopoietic progenitors to blood in monkey.
Animals ; Cell Differentiation ; drug effects ; Cells, Cultured ; Chromatography, High Pressure Liquid ; Haplorhini ; Hematopoietic Stem Cells ; cytology ; drug effects ; Humans ; Recombinant Proteins ; chemistry ; genetics ; metabolism ; pharmacology ; Sequence Analysis, Protein ; Spectrometry, Mass, Fast Atom Bombardment ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Stem Cell Factor ; chemistry ; genetics ; metabolism ; pharmacology

Biomarkers, Tumor ; metabolism ; Diagnosis, Differential ; Diagnostic Imaging ; Humans ; Mass Spectrometry ; methods ; Neoplasms ; diagnosis ; drug therapy ; metabolism ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

By investigating the interaction between components from Flos Genkwa (FG) and Radix et Rhizoma Glycyrrhizae (RRG) and the dissolution profile of toxic components in co-decoction, the characteristics and possible mechanism of incompatibility were revealed. Ultra-high performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) and ultra-high performance liquid chromatography triple-quadrupole mass spectrometry (UPLC-TQ/MS) were used to analyze multi-components in different herb extractions prepared by different ratios of FG/FG processed by vinegar (FGV) and RRG, which reflect the interaction and characteristics of multiple components in incompatibility combinations. The results showed that the components dissolution was influenced by compatibility ratio with certain regularity. Whether FG processed by vinegar or not, with the increase of RRG in co-decoction, the dissolution of diterpenes, especially for yuanhuacine, yuanhuadine and yuanhuajine, the toxic ingredients of FG, increased significantly. From these results, the material basis and one possible mechanism of incompatibility between FG and RRG is the increasing dissolution of diterpenes, toxic components of FG in co-decoction process, which caused by interaction between multi-components in these two herbs.
Acetic Acid ; chemistry ; Chromatography, High Pressure Liquid ; Daphne ; chemistry ; Diterpenes ; analysis ; Drug Incompatibility ; Flowers ; chemistry ; Glycyrrhiza uralensis ; chemistry ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry ; Rhizome ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Tandem Mass Spectrometry ; Terpenes ; analysis

The aim is to determine the primary structure of a new hirudin and reteplase fusion protein (HV12p-rPA) by LC-ESI-MS/MS spectrometry. The molecular weight of the hirudin and reteplase fusion protein (HV12p-rPA) was measured by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The HV12p-rPA was digested with trypsin and chymotrypsin separately and the peptides in the digest mixtures were identified by LC-ESI-MS/MS. The molecular weight of HV12p-rPA was 41,472 Da, which was in accordance with the theoretical value. The peptide fragments of HV12p-rPA digested with trypsin were identified by LC-ESI-MS/MS spectrometry and the results indicated that the fusion protein contained r-PA. Then, the peptides of HV12p-rPA digested with chymotrypsin were identified by the same method. The results indicated that the fusion protein contained HV12p and the linker-containing peptide, DEGGGSY. MASDF and LDWIRDNMRP were identified as the N-terminal and C-terminal containing peptides in the chymotryptic digest mixture of the fusion protein. All of the Xcorr values exceeded 1.5, some of which were above 3.0, showing that the results were correct and credible and a sequence coverage of 85% was achieved. HPLC/MS analysis coupled with uncompleted digestion indicated that all these peptides were arranged with the correct order as expected. Thus, sequence of the fusion protein was confirmed and it was consistent with our design in upstream construction.
Amino Acid Sequence ; Chromatography, Liquid ; methods ; Chymotrypsin ; chemistry ; Fibrinolytic Agents ; analysis ; chemistry ; Hirudins ; analysis ; chemistry ; Molecular Sequence Data ; Molecular Weight ; Peptide Fragments ; Recombinant Fusion Proteins ; analysis ; chemistry ; Recombinant Proteins ; analysis ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; methods ; Tandem Mass Spectrometry ; methods ; Tissue Plasminogen Activator ; analysis ; chemistry ; Trypsin ; chemistry

A novel rapid method for detection of the illicit beta2-agonist additives in health foods and traditional Chinese patent medicines was developed with the desorption corona beam ionization mass spectrometry (DCBI-MS) technique. The DCBI conditions including temperature and sample volume were optimized according to the resulting mass spectra intensity. Matrix effect on 9 beta2-agonists additives was not significant in the proposed rapid determination procedure. All of the 9 target molecules were detected within 1 min. Quantification was achieved based on the typical fragment ion in MS2 spectra of each analyte. The method showed good linear coefficients in the range of 1-100 mg x L(-1) for all analytes. The relative deviation values were between 14.29% and 25.13%. Ten claimed antitussive and antiasthmatic health foods and traditional Chinese patent medicines from local pharmacies were analyzed. All of them were negative with the proposed DCBI-MS method. Without tedious sample pretreatments, the developed DCBI-MS is simple, rapid and sensitive for rapid qualification and semi-quantification of the illicit beta2-agonist additives in health foods and traditional Chinese patent medicines.
Adrenergic beta-2 Receptor Agonists ; analysis ; Drugs, Chinese Herbal ; chemistry ; Food, Organic ; analysis ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Substance Abuse Detection ; methods ; Tandem Mass Spectrometry

To identify the structure of three related substances in potassium sodium dehydroandrographolide succinate (PSDS), an HPLC preparation method was used to separate the impurities. These main impurities were identified using LC-ESI/TOFMS, LC-ESI/MSn, NMR, UV and IR. One of the main impurities was a hydrolyzed and oxidized product of PSDS, which has not been reported previouely. The other two impurities were hydrolyzed products of PSDS after losing different succinic acids. The results indicate that PSDS can be easily hydrolyzed and oxidized. It should be stored at cool and dry places.
Andrographis ; chemistry ; Antiviral Agents ; chemistry ; isolation & purification ; Chromatography, High Pressure Liquid ; Diterpenes ; chemistry ; isolation & purification ; Drug Contamination ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Molecular Structure ; Plants, Medicinal ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

To evaluate the reagent 2-methoxy-4,5-dihydro-1H-imidazole used for isotopic labeling in quantitative proteomics, we synthesized 2-methoxy-4,5-dihydro-1H-imidazole and its tetradeuterated analog in three steps. Prior to tryptic cleavage, bovine serum albumin (BSA) was reduced and alkylated. Tryptic peptides were derivatized with an equal volume of either DO or D4 and D4-derivatized peptides were mixed with at variable ratio (from 10:1 to 1:5) prior to MS and MS/MS analysis. We used matrix assisted laser desorption/ionization-mass spectrometry (MALDI-MS) and Electro spray ionization-mass spectrometry (ESI-MS) to evaluate the quantitative capability of labeling. The specificity of the reagent is excellent: only lysine side chains were modified among tryptic peptides. MALDI and ESI ionization modes not only could achieve the quantification of differentially expressed proteins but also facilitate the de novo sequencing. This side-chain modification can be used for quantitative analysis with proteomic strategies involving liquid chromatography. Reverse phase liquid chromatography (RPLC) kept a good resolution, and the introduction of D atoms did not introduce a variation of retention time between heavy and light peptides in RPLC.
Imidazoles ; chemistry ; Isotope Labeling ; methods ; Lysine ; chemistry ; Peptides ; analysis ; chemistry ; Proteomics ; methods ; Serum Albumin, Bovine ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; methods