ESI-MS Method Research Articles

Sustainable and Stability Indicating Liquid Chromatography–Mass Spectrometry Method for the Quantitation of Delafloxacin in Pharmaceutical Formulations

ABSTRACTA sensitive, precise, accurate, and green analytical HPLC–ESI–MS method for the quantification of delafloxacin and its degradation products in pharmaceutical dosage forms has been optimized and validated. The best separation was achieved with isocratic elution, the mobile phase is composed of a mixture of 0.1% trifluoroacetic acid in water and acetonitrile 65:35 (v/v), the flow rate is 0.5 mL min−1, and the column used is Kinetex Core‐Shell C18 (250 × 4.6 mm, 5 µm). Forced degradation studies were performed to prove that the method indicates stability. The pharmaceutical substance is prone to oxidative (H2O2 3%), acidic (HCl 0.1 M), and basic (0.1 M) conditions. Delafloxacin proved to be susceptible to acidic (HCl 0.1 M), basic (0.1 M), and oxidative (H2O2 3%) conditions. The proposition of the structures of degradation products has been based on the MS spectrum and the known reactivity of delafloxacin in the oxidative medium. The validation of the analytical method was carried out following the International Conference on Harmonization guidelines. The method was validated in terms of specificity, precision, linearity, and accuracy. The limits of detection and quantification of delafloxacin are, respectively, 0.005 and 0.017 µg mL−1.

Analysis of the Polyphenolic Composition of Vaccinium L. Extracts and Their Protective Effect on Red Blood Cell Membranes

The blueberry fruit of the genus Vaccinium, including high blueberry, low blueberry, and wild bilberry, is consumed for its flavor and medicinal properties. The purpose of the experiments was to investigate the protective effect and mechanism of the interaction of blueberry fruit polyphenol extracts with the erythrocytes and their membranes. The content of polyphenolic compounds in the extracts was determined using the chromatographic UPLC–ESI–MS method. The effects of the extracts on red blood cell shape changes, hemolysis and osmotic resistance were examined. Changes in the order of packing and fluidity of the erythrocyte membrane and the lipid membrane model caused by the extracts were identified using fluorimetric methods. Erythrocyte membrane oxidation was induced by two agents: AAPH compound and UVC radiation. The results show that the tested extracts are a rich source of low molecular weight polyphenols that bind to the polar groups of the erythrocyte membrane, changing the properties of its hydrophilic area. However, they practically do not penetrate the hydrophobic part of the membrane and do not damage its structure. Research results suggest that the components of the extracts can defend the organism against oxidative stress if they are delivered to the organism in the form of dietary supplements.

Cation–π interaction of thallium (I) with [7]helicene: an experimental and theoretical study

Electrospray ionisation mass spectrometry was used to investigate the cation–π interaction of Tl+ with [7]helicene. Further, applying quantum mechanical calculations in the frame of the density functional theory, applying B3LYP functional with Def2TZVP basis set, the most probable structure of the [7]helicene–Tl+ complex was derived. In the resulting complex, the ‘central’ cation Tl+ is bounded between two terminal benzene rings of the parent [7]helicene ligand via cation–π interaction. In such conformation, the [7]helicene creates a tweezer for Tl+ ion. The interaction energy, E(int), of this cation–π complex was calculated to be –160 kJ/mol, which is consistent with the formation of the considered cationic complex species. Other structures were also found and generally lie energetically about 19–24 kJ/mol above the tweezer complex. HIGHLIGHTS Cation Tl+–[7]helicene complex has been observed using the ESI-MS method The structure of the resulting complex has been determined by applying quantum chemical DFT calculations Helicene functions as a chiral molecular tweezer for Tl+.

Molecular structure and spectroscopic studies of the product of acidic degradation of salinomycin and its potassium salt

Salinomycin (SAL), a monocarboxylic polyether antibiotic isolated from Streptomyces albus is widely used in veterinary medicine as a growth promoter and as a supplement in poultry feed to control infection with coccidia. SAL exhibits also potent activity against the proliferation of various cancer cells, including those that display multidrug resistance (MDR), and can precisely kill cancer stem cells (CSCs). The compound 2 – a product of the acidic degradation of SAL has been isolated and structurally characterized by X-ray, FT-IR, NMR, and DFT methods. Additionally, to check the ionophoretic properties of this compound its potassium salt (2-K) has been also obtained and the crystal structures of these two compounds as well as their structures in solution were studied in detail. It was found that the structures of 2 and 2-K salt were different in the solid and in the solution. Using ESI MS method it was demonstrated that that compound 2 was also able to form 1:1 and 2:1 complexes with Na+, K+, and Rb+ cations. For the first time we have demonstrated that the product of salinomycin degradation still shows ionophoretic properties.

New Insights into the Structure of Kappa/Beta-Carrageenan: A Novel Potential Inhibitor of HIV-1.

New insights into the structure of the hybrid κ/β-carrageenan (κ/β-CRG) of the red alga Tichocarpus crinitus have been obtained. Carrageenan oligosaccharides were prepared through the chemical and enzymatic depolymerization of κ/β-CRG with κ-carrageenase and its the enzyme-resistant fraction. The composition and distribution of the repetition units of κ/β- CRG were investigated by using the negative ion tandem MALDI-TOFMS and ESIMS method, which made it possible to prove and characterize the hybrid structure of this polysaccharide. An analysis revealed the blockwise distribution of the long β-blocks along the polysaccharide chain, with the inclusion of κ/β, μ/ν-blocks and some ι-blocks. Furthermore, the desulfated κ/β-CRG was shown to contain of –G–D– repeating units up to 3.5 kDa. Previous studies have demonstrated that CRGs suppress the replication of several viruses. Here, we established that κ/β-CRG and its oligosaccharides significantly inhibit the transduction efficiency of replication-defective lentiviral particles pseudotyped with the envelope proteins of three different viruses. We found that the polysaccharide and its oligosaccharides strongly reduced the transduction efficiency of lentiviral particles pseudotyped with GP160—the envelope protein of the human immunodeficiency virus HIV-1—when added to T-lymphocyte Jurkat cells. The CRG oligosaccharides displayed significantly higher antiviral activity.

Mass Spectrometric Investigation of Organo-Functionalized Magnetic Nanoparticles Binding Properties toward Chalcones.

Chalcones are naturally occurring compounds exhibiting multiple biological functions related to their structure. The investigation of complexes formed by chalcones, namely 2′,4′-dihydroxy-2-methoxychalcone (DH-2-MC) and 2′,4′-dihydroxy-3-methoxychalcone (DH-3-MC), with organo-functionalized Fe3O4 magnetic nanoparticles using mass spectrometric techniques is reported. The magnetic nanoparticles were obtained by the silanization of Fe3O4 particles with 3-aminopropyltrimethosysilane, which were subsequently reacted with 3-hydroxybenzaldehyde (3-HBA) or 2-pyridinecarboxaldehyde (2-PCA), resulting in the formation of Schiff base derivatives. The formation of their complexes with chalcones was studied using electrospray (ESI) and flowing atmosphere-pressure afterglow (FAPA) mass spectrometric (MS) ionization techniques. The functional nanoparticles which were synthesized using 3-hydroxybenzaldehyde displayed higher affinity towards examined chalcones than their counterparts obtained using 2-pyridinecarboxaldehyde, which has been proved by both ESI and FAPA techniques. For the examined chalcones, two calibration curves were obtained using the ESI-MS method, which allowed for the quantitative analysis of the performed adsorption processes. The presence of Cu(II) ions in the system significantly hindered the formation of material–chalcone complexes, which was proved by the ESI and FAPA techniques. These results indicate that both mass spectrometric techniques used in our study possess a large potential for the investigation of the binding properties of various functional nanoparticles.

Review of Adulterated Herbal Products and Supplements and Methods of Analysis

Herbal products and supplements are widely used for primary health care. Many reports of drugs added to these products, such as weight-loss supplements, analgesic herbal products, and erectile dysfunction supplements. This study aims to find out products that are often mixed with drugs and effective methods used for their analysis. The method used in this research is a systematic review. The resources were retrieved through searching the keywords related to the world adulterated herbal products and supplements in the available databases and search engines including Science Direct, PubMed, Springerlink, and Google Scholar. Based on PRISMA flow diagram, a systematic review showed that sibutramine HCl, sildenafil citrate, dexamethasone, and sodium diclofenac are most commonly found in the herbal and supplement products and the ESI-MS method is an effective method for their analysis.

Phospholipid and n-alkane composition, anti-α-glucosidase and anti-cyclooxygenase activities of milk thistle oil

Although milk thistle oil has been authorized as a new edible oil in many countries, there is a lack of information about its chemical and biological properties. Thus, this study aimed to characterize the phospholipid and n-alkane composition as well as the α-glucosidase and cyclooxygenase inhibitory effects of milk thistle oil. The phospholipid composition of milk thistle seed oil was determined by liquid chromatography coupled to mass spectrometry HPLC–ESI–MS method and five glycerophospholipid classes were detected: phosphatidylinositol (35.8 ± 2.31%), phosphatidylcholine (29.2 ± 1.43%), phosphatidylethanolamine (15.7 ± 1.75%), phosphatidylglycerol (12.4 ± 1.61%) and phosphatidic acid (6.9 ± 0.82%). Twenty-three phospholipid molecular species were characterized, containing mainly palmitic, oleic and linoleic acids. Using gas chromatography coupled to mass spectrometry, seventeen n-alkanes were detected ranging from C14 to C35 carbon atoms. Nonacosane (13.79 ± 1.15%) was the predominant compound followed by heptacosane (9.52 ± 0.87%) and hexacosane (9.33 ± 1.21%). The carbon preference index of long-chain n-alkanes (CPI25–35) exhibited mainly odd/even predominance. The phospholipid and n-alkane composition of milk thistle oil is quite different from that of other oilseeds, indicating the possible use of those compounds as a means of determining oil authenticity. The total phenolic content of the tested oil was 10.28 ± 1.2 mg GAE/ g of oil. Furthermore, the tested oil showed moderate anti-cyclooxygenase1 capacity (IC50 = 42.3 ± 0.8 μg/mL) in comparison with the positive control aspirin. However, the inhibitory effect of milk thistle oil against α-glucosidase (IC50 = 28.4 ± 0.54 μg/mL) was similar to that of the positive control acarbose, which reveals the tested oil as a promising source of natural α-glucosidase inhibitors, proves protection against type 2 diabetes. Milk thistle oil has an excellent potential for application in the food and pharmaceutical industries.

Enhancement in MS‐based peptide detection by microfluidic free‐flow zone electrophoresis

Matrix components are known to significantly alter the ionization of a target analyte in ESI-based measurements particularly when working with complex biological samples. This issue however may be alleviated by extracting the analyte of interest from the original sample into a relatively simple matrix compatible with ESI mass-spectrometric analysis. In this article, we report a microfluidic device that enables such extraction of small peptide molecules into an ESI-compatible solvent stream significantly improving both the sensitivity and reproducibility of the measurements. The reported device realizes this analyte extraction capability based on the free-flow zone electrophoretic fractionation process using a set of internal electrodes placed across the width of the analysis channel. Employing lateral electric fields and separation distances of 75 V/cm and 600µm, respectively, efficient extraction of the model peptide human angiotensin II was demonstrated allowing a reduction in its detection limit by one to three orders of magnitude using the ESI-MS method. The noted result was obtained in our experiments both for a relatively simple specimen comprising DNA strands and angiotensin II as well as for human serum samples spiked with the same model peptide.

Analysis of condensed tannins in Populus spp. using reversed phase UPLC-PDA-(-)esi-MS following thiolytic depolymerisation.

Condensed tannins (CTs) are proanthocyanidin heteropolymers that are widely distributed among plants. Their biochemical properties are determined by molecular structure (e.g. polymer size, hydroxylation, stereochemistry). In Populus, genetically and environmentally-determined CT concentrations have been related to ecological effects, while the potential role of CT molecular structure has received little attention. Evaluate CT polymerisation, major constituent monomers, stereochemistry and overall content in Populus tremuloides foliage using ultra-high-performance liquid chromatography with photodiode array and mass spectrometry (UPLC-PDA-(-)esi-MS) detection following thiolytic depolymerisation of the CTs. CTs were extracted from dried foliage of six P. tremuloides genotypes into methanol and thiolytically depolymerised into constituent monomers. Calibration standards were prepared by thiolysis of CT mixtures isolated from P. tremuloides foliage on Sephadex LH-20, followed by preparative high-performance liquid chromatography (HPLC). Populus tremuloides CTs contained predominantly repeating subunits of three putative stereoisomers each of catechin and gallocatechin. Linear calibrations for standards of these subunits and their thioethers (purities 44-87%, UPLC-(-)esi-MS) were generally stable over the course of 10months. CT polymer size, hydroxylation, stereochemistry and concentrations differed among genotypes. This thiolysis-UPLC-PDA-(-)esiMS method was optimised for analysis of CT polymer size, hydroxylation, stereochemistry, and total concentration in Populus foliage. It revealed significant variation in each of these properties among P. tremuloides genotypes, and will facilitate evaluation of how environmental factors affect CT molecular structures.

ESI-MS measurements for the equilibrium constants of copper(II)-insulin complexes

Trace elements regulate many biological reactions in the body. Copper(II) is known as one of trace elements and capable of binding to proteins. Insulin is a blood glucose-lowering peptide hormone and it is secreted by the pancreatic β-cells. In this study, Cu(II)-insulin complexes were investigated by using ESI-MS method. Insulin molecule gives ESI-MS peaks at +4, +5, +6 and +7 charged states. Cu(II)-insulin complexes can be monitored and quantified on the ESI-MS spectra as the shifted peaks according to insulin peaks. The solutions of Cu(II)-insulin complexes at different pHs and mole ratios of Cu(II) ions to insulin molecule were measured on the ESI-MS. The highest complex formation ratio for Cu(II)-insulin were found at pH 7. The multiple bindings of Cu(II) ions to insulin molecule was observed. The formation equilibrium constants of Cu(II)-insulin complexes were calculated as Kf1: 3.34 × 104, Kf2: 2.99 × 104, Kf3: 7.00 × 103 and Kf4:2.86 × 103. The specific binding property of Cu(II) ions was controlled by using different spray ion sources including electrospray and nano-electrospray. The binding property of Cu(II) also investigated by MS/MS fragmentation. It was concluded from the ESI-MS measurements that Cu(II) ion has a high affinity to insulin molecules to form stable complexes.

Quantifying Na(I)-insulin and K(I)-insulin non-covalent complexes by ESI–MS method and calculation of their equilibrium constants

In this study, the dissociation and formation equilibrium constants of Na(I)-insulin and K(I)-insulin complexes have been calculated after the quantifying them on ESI mass spectrometer. The ESI–MS spectra of the complexes were measured by using the solvents as 50% MeOH in water and 100% water. The effect of pH on the Na(I)-insulin and K(I)-insulin complex formation were examined. Serial binding of Na(I) and K(I) ions to the insulin molecule were observed in the ESI–MS measurements. The first formation equilibrium constants were calculated as Kf1: 5.48×103 1/M for Na(I)-insulin complex and Kf1: 4.87×103 1/M for K(I)-insulin in water. The binding capability of Na(I) ions to insulin molecule is higher than the capability of K(I) ions. In case of a comparison together with Ca(II)-insulin and Mg(II)-insulin, the formation equilibrium constants (Kf1) are in order of Ca(II)-insulin>Mg(II)-insulin>Na(I)-insulin>K(I)-insulin in water. The results showed that Na(I) and K(I) ions are involved in the formation of the non-covalent complexes with insulin molecule, since high extracellular and intracellular concentrations of them in the body.

Development and validation of a specific and sensitive HPLC-ESI-MS method for quantification of lysophosphatidylinositols and evaluation of their levels in mice tissues

Increasing evidence suggests that lysophosphatidylinositols (LPIs), a subspecies of lysophospholipids, are important endogenous mediators. Although LPIs long remained among the less studied lysophospholipids, the identification of GPR55 as their molecular target sparked a renewed interest in the study of these bioactive lipids. Furthermore, increasing evidence points towards a role for LPIs in cancer development. However, a better understanding of the role and functions of LPIs in physiology and disease requires methods that allow for the quantification of LPI levels in cells and tissues. Because dedicated efficient methods for quantifying LPIs were missing, we decided to develop and validate an HPLC–ESI–MS method for the quantification of LPI species from tissues. LPIs are extracted from tissues by liquid/liquid extraction, pre-purified by solid-phase extraction, and finally analyzed by HPLC-ESI-MS. We determined the method’s specificity and selectivity, we established calibration curves, determined the carry over (< 2%), LOD and LLOQ (between 0.116–7.82 and 4.62–92.5pmol on column, respectively), linearity (0.988<R2<0.997), repeatability (CV<20%), accuracy (> 80%), intermediate precision (CV<20%) as well as the recovery from tissues. We then applied the method to determine the relative abundance of the LPI species in 15 different mouse tissues. Finally, we quantified the absolute LPI levels in six different mouse tissues. We found that while 18:0 LPI represents more than 60% of all the LPI species in the periphery (e.g. liver, gastrointestinal tract, lungs, spleen) it is much less abundant in the central nervous system where the levels of 20:4 LPI are significantly higher. Thus this validated HPLC-ESI-MS method for quantifying LPIs represents a powerful tool that will facilitate the comprehension of the pathophysiological roles of LPIs.

Development and Validation of a Rapid and Simple UPLC-ESI-MS Method for Pharmacokinetics and Tissue Distribution of Astragaloside III in Rats.

A rapid and simple ultra-performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS) method for the determination of astragaloside III was developed and used in a pharmacokinetic and tissue distribution study in rats following the oral administration 95% ethanol extraction of Zhenqi Fuzheng capsules. Although astragaloside III and astragaloside IV have the same molecular weight and very similar structures, they were successfully separated using this method. Quantification was performed using low-energy collision tandem mass spectrometry (CID-MS-MS) with the multiple reaction monitoring scan mode of the following precursor ion → product ion atm/z807.61→335.22 for astragaloside III and atm/z633.18→331.18 for the internal standard (hesperidin). Both astragaloside III and astragaloside IV in rat plasma were best fit to a two-compartment model. The tissue distribution study showed the overall trend of disposition of astragaloside III were C thymus > C spleen > C stomach > C liver > C heart > C kidney > C lung > C testicle The high levels of astragaloside III in thymus and spleen indicated an accumulation in organs involved in immune responses and showed that these organs are major target sites in vivo The results in the article will provide valuable information for use in clinical applications of astragaloside III.

Chemical composition and biological activity of the plum seed extract

The aim of this paper was to estimate the biological activity of the plum seed extract and to define the chemical composition by using the ESI-MS method. During the investigation of the antioxidant activity, the extract showed a better ability to inhibit DPPH radicals compared with amygdalin standard. The results of the antimicrobial study indicate that the extract has a greater effect on Gram-negative bacteria compared with amygdalin. Gram-positive bacteria and fungi remained resistant in both cases. Unlike amygdalin, the extract showed a greater antimicrobial activity against the same microbiological strains. Amygdalin was isolated from the extract before the investigation of antiproliferative activity against MDA-MB-361, MDA-MB-453, HeLa and LS174 cell lines using MTT assay. Unlike the MDA-MB-453 cell line, the activity of standard and isolated amygdalin was not observed at the concentrations lower than 400 μg cm-3 on MDA-MB-361 cell line. The isolated amygdalin has not shown the activity in other cell lines. The antiproliferative activity of amygdalin standard was also noticed on HeLa and LS-174 cell lines at the concentrations of 333.27 and 333.17 μg cm-3, respectively.

Investigation of the structural characterization and reversible oxygenation mechanism of oxygenated cobalt–3-(4-thiazolyl)-l-alanine and cobalt–histidine complexes using electrospray tandem mass spectrometry

A new ESI-MS method was developed to identify oxygenated species in Co–Tza formed by the uptake of dioxygen. The structures and fragmentation mechanisms of Co–Tza and Co–His were deduced.

The signal response linearity in phthalates determination using ESI-MS method with the Loop injection technique

The possibility of using ESI-MS technique for the quantitative determination of six most commonly used phthalates (DMP, DnBP, BzBP, DEHP, DiNP, DiDP) was tested. The test was performed by using Loop technique for recording chromatogram obtained after the injection of different phthalates concentrations. The investigation was performed by determining the linear relationship between the concentration of phthalates and the peak areas in the chromatogram of mass range which corresponds to the molecular ion and ion adduct with sodium. The obtained results showed that there is linearity between the concentration of phthalates and peak areas in the chromatogram of monitoring molecular and adduct ions with sodium in case of DiNP (r > 0.999) and DMP (r > 0.999 for the molecular ion; r > 0.989 for the ion adduct with sodium). The results obtained for the molecular ion in DiDP case showed a better linear relationship than the ion adduct with sodium, while the results obtained for BzBP showed opposite values. The same correlation coefficients for both monitoring ions, molecular ion and ion adduct with sodium were obtained for DEHP (r = 0.98371 and 0.98962, respectively). The correlation coefficients DnBP showed lower values than other linear coefficients (r = 0.96034 and 0.94669, respectively). Although the signals of adduct ions with sodium are steady and intense, they are not reliable for quantification since the source of sodium ions is not defined and therefore, the actual amount of sodium in the sample may vary.

The qualitative analysis of the green tea extract using ESI-MS method

The aim of this paper was the identification of the bioactive compounds in the green tea extract enriched with quercetin. The extraction was performed under reflux for 60 min using absolute ethanol at the solid-to-liquid ratio of 1:18 (m/v). The mass spectrometry method with an electrospray interface was applied and tuned with the quercetin standard. Mass spectra were recorded in negative and positive ionization modes with the ion source voltage of 4.95 kV, the source current of 2.61 μA, the capillary temperature of 275 °C, the capillary voltage of 50 V, the sheath gas pressure of 137 KPa and auxiliary gas pressure of 13.9 KPa. In addition to quercetin, the presence of gallic acid, (-)-gallocatechin, 3-O-caffeoylquinic acid, theobromine, 5-O-galloylquinic acid, (+)-catechin, caffeine, epicatechin-3-gallate, (-)-epigallocatechin-3-gallate, 4-p-coumaroylquinic acid, quercetin-3-O-rutinoside, quercetin-3-galactoside, kampferol-3-O-glucoside, apigenin glycoside, theaflavin-3,3'-digallate, thafla­vin-3-gallate and the kaempferol-rhamnose-hexose-rhamnose conjugate was confirmed based on fragmentation patterns available in literature.

ESI-MS Study of Ionization Pathways and Cation-Receptor Properties of the Iron(II) Mono- and Bis-Clathrochelates

Ionization pathways of iron(II) di- and hexachloromonoclathrochelates, bis-clathrochelates and their oximehydrazonate, dimethyl- and diamine-containing macrobicyclic analogs, and the cation-receptor properties of such cage compounds towards alkali metal cations (as individual ions or their mixtures) as well as the competitive complexation in the presence of 18-crown-6 were studied by ESI-MS method. Ionization of iron(II) mono- and bis-clathrochelates under the conditions of ESI-MS experiments was found to proceed via different pathways. The macrobicyclic iron(II) oximehydrazonate as an ionic associate of the clathrochelate cation with BF 4 – anion undergoes the ionization by its heterolytic dissociation. The main pathways of ionization of the dioximate cage and bis-cage intracomplexes are substantially affected by ribbed substituents in their chelate fragments. The methyland amine-containing macrobicyclic complexes easily form anion-radical species by one-electron oxidation of their polyazomethine cage frameworks, whereas in the case of iron(II) dihalogenoclathrochelates the most intensive peaks correspond to their ionic associates with monocharged alkali metal cations. ESI-MS positive-mode spectrum of the iron(II) hexachloroclathrochelate contains no detectable peaks of the cationic species, which resulted either from its oxidation to the corresponding cation-radical or from its ionic associates with alkali metal cations. In the negative mode, an intensive peak assigned to the anion-radical product of one-electron reduction was observed. As ESI mass spectrometry allows maintaining precise concentrations of the complex under study and that of “cationization agents” (i.e., the compounds that promote formation of cationic species of the analyte and have been used in the case of low-ionizable complexes), the alkali metal tetraphenylborates were applied as such agents to study cation-receptor properties of the iron(II) mono- and bis-clathrochelates with ribbed substituents of the different nature towards these metal ions. Iron(II) monoclathrochelates form two types of ionic associates with such cations with both the 1:1 and 2:1 stoichiometries, which are more and less intensive in their ESI-MSs, respectively, whereas ESI-MSs of bis-cage complexes contain the intensive peaks of only 1:1 ionic associates with high affinity towards Cs + cation.

Analysis of the monosaccharide composition of water-soluble polysaccharides from Sargassum fusiforme by high performance liquid chromatography/electrospray ionisation mass spectrometry

Sargassum fusiforme (hijiki) is the well-known edible algae, whose polysaccharides have been proved to possess interesting bioactivities like antitumor, antioxidant, antimicrobial and immunomodulatory activities. A facile and sensitive method based on high-performance liquid chromatography method of pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP) coupled with electrospray ionisation mass spectrometry (HPLC/ESI–MS) has been established for the analysis of the monosaccharide composition of polysaccharides in S. fusiforme. Monosaccharides have been converted into PMP-labelled derivatives with aqueous ammonia as a catalyst at 70°C for 30min. The optimisation of the pre-column derivatization process was studied. The LODs of the monosaccharides were in the range from 0.01 to 0.02nmol. PMP-labelled mixture of monosaccharides has been well separated by a reverse-phase HPLC and detected by on-line ESI–MS method under optimised conditions. The mobile phase of elution system was chosen as acetonitrile (solvent A) and 20mM aqueous ammonium acetate (solvent B) (pH 3.0) with Zorbax XDB-C18 column at 30°C for the separation of the monosaccharide derivatives. Identification of the monosaccharides composition was carried out by analysis with mass spectral behaviour and chromatography characteristics of 1-phenyl-3-methyl-5-pyrazolone (PMP) labelled monosaccharides. All PMP-labelled derivatives display high chemical stabilities, whose regular MS fragmentation is specific for reducing labelled sugars. The result showed that the S. fusiforme polysaccharide consisted of mannose, glucose, galactose, xylose, fucose and glucuronic acid or galacturonic acid, or both uronic acids.