Studies on the non-covalent complexes between oleanolic acid and cyclodextrins using electrospray ionization tandem mass spectrometry

Non-covalent inclusion complexes formed between an anti-inflammatory drug, oleanolic acid (OA), and alpha-, beta- and gamma-cyclodextrins (CDs) were investigated by means of solubility studies and electrospray ionization tandem mass spectrometry (ESI-MSn). The order of calculated association constants (K-1:1) of complexes between OA and different CDs in solution is in good agreement with the order of their relative peak intensities and the relative CID energies of the complexes under the same ESI-MSn conditions. These results indicate a direct correlation between the behaviors of solution- and gas-phase complexes. ESI-MS can thus be used to evaluate solution-phase non-covalent complexes successfully. The experimental results show that the most stable 1:1 inclusion complexes between three CDs and OA can be formed, but 2:1 CD-OA complexes can be formed with beta- and gamma-CDs. Multi-component complexes of alpha-CD-OA-beta-CD (1:1:1), alpha-CD-OA-gamma-CD (1:1:1) and beta-CD-OA-gamma-CD (1:1:1) were found in equimolar CD mixtures with excess OA. The formation of 2:1 and multi-component 1:1:1 non-covalent CD-OA complexes indicates that beta- and gamma-CD are able to form sandwich-type inclusion non-covalent complexes with OA. The above results can be partly supported by the relative sizes of OA and CD cavities by molecular modeling calculations.

Analysis of flavonoid constituents from leaves of Acanthopanax senticosus Harms by electrospray tandem mass spectrometry

Three known flavonoids, quercetin, quercitrin (quercetin-3-0-rhamnoside) and rutin (quercetin-3-0-rutinoside), have been identified for the first time in the leaves of Acanthopanax senticosus Harms by using electrospray tandem mass spectrometry techniques (ESI-MSn). The flavonoid hyperin (quercetin-3-0-beta-galactoside), already known to be present, was also investigated. The diagnostic fragment ions of the aglycone quercetin were obtained in the ESI-MSn experiments, and a fragmentation mechanism proposed.

Rapid identification of saponins in plant extracts by electrospray ionization multi-stage tandem mass spectrometry and liquid chromatography/tandem mass spectrometry

Electrospray ionization multi-stage tandem mass spectrometry (ESI-MSn) and liquid chromatography coupled with on-line mass spectrometry (LC/MS/MS) were applied to characterize saponins in crude extracts from Panax ginseng. The MSn data of the [M - H](-) ions of saponins can provide structural information on the sugar sequences of the saccharide chains and on the sapogins of saponins. By ESI-MSn, non-isomeric saponins and isomeric saponins with different aglycones can be determined rapidly in plant extracts. LC/MS/MS is a good complementary analytical tool for determination of isomeric saponins. These approaches constitute powerful analytical tools far rapid screening and structural assignment of saponins in plant extracts. Copyright (C) 2000 John Wiley & Sons, Ltd.

Gaseous dipeptide complexes with metal ions by electrospray ionization and tandem mass spectrometry

Electrospray ionization (ESI) and tandem mass spectrometry have been used to investigate the gas-phase interactions of five metal ions and seven dipeptides. For silver ion, two complexes ([M+Ag](+) and [2M+Ag](+)) were obtained as well as the one complex ([2M+Met-H](+)) for transition-metal ions. Upon collision activation, there is an obvious difference in MS/MS data between metal ion complex and the protonated molecule. The fragment pathway of each complex is related to the structures of dipeptide and the nature of metal ion which suggest that there are several interaction between the metal ions and dipeptides in gas phase.

Studies on negative electrospray ionization mass spectrometry on saponins

Two kinds of saponins have been investigated by negative electrospray ionization (NESI) mass spectrometry. Under ESI conditions, the [M - H](-) ions of saponins were observed which provide the molecular weights of saponins. The fragment pathways of [M - H](-) ions of these two saponins depend on their structures. For steroidic saponins, [M - H](-) ion only produces the fragment ions by the losses of sugar units. For oleanolic saponins, [M - H](-) ion yields the cross-ring ions as well as the fragment ions by the losses of sugar units. Moreover, the abundance of the former is higher than that of the latter. The characteristic fragments are used to provide the sequence and some linkage information of sugar moieties of saponins. Especially, their fragment difference strongly depends on the linkage between the aglycone and the sugar moieties.

Investigation of metalloporphyrins and their imidazole complexes using electrospray ionization mass spectrometry

Three compounds of metalloporphyrins were studied using electrospray ionization mass spectrometry. The bonding power between substitutional phenyl and porphyrin cycle and the coordinate conditions of metalloporphyrins with imidazole were discussed. The experimental result indicated that the bonding power between substitutional phenyl and porphyrin cycle in metalloporphyrins became weak from Mn, Fe to Co. The complexes abundances formed by metallophorphyrin with imidazole were stronger with the increase of the ligand concentration. At the same ligand concentration, the abundance of the complexes was intensified gradually and the stability of the ligands was become stronger from Mn, Fe to Co.

Metal ion adducts in the structural analysis of ginsenosides by electrospray ionization with multi-stage mass spectrometry

The effect of metal (Li+, Na+, K+, Ag+) cationization on collision-induced dissociation of ginsenosides was investigated by electrospray ionization mass spectrometry combined with multi-stage mass spectrometry (ESI-MSn). The fragments of sodiated and lithiated molecules give valuable structural information regarding the nature of the aglycone and the sequence and linkage information of sugar moieties. However, the number and relative abundances of fragment ions from lithiated ginsenosides are significantly greater than for the sodiated species, The K+ adducts undergo glycosidic cleavages and very limited cross-ring reactions. The silver ion adducts fragment mainly through glycosidic cleavages. Copyright (C) 2001 John Wiley & Sons, Ltd.

Electrospray ionization mass spectrometry of cyclodextrin complexes with amino acids in incubated solutions and in eluates of gel permeation chromatography

1:1 complexes of beta-cyclodextrin (CD) with three amino acids (Gly, Phe and Trp) have been detected as ions in the gas phase using infusion positive and negative ion electrospray ionization mass spectrometry (ESI-MS). In contrast with the positive ion ESI mass spectra of simple aqueous solutions, the aggregates and adducts usually formed in the ESI process did not appear in the positive ion ESI spectra of solutions buffered with ammonium acetate (NH4Ac), even at higher analyte concentrations, These studies suggest that addition of buffer and/or use of a low analyte concentration should be used to overcome formation of aggregates and metal ion adducts in such mass spectrometry studies. Also, the deprotonated complexes are dissociated by collision induced dissociation (CID) to form an abundant product ion, the deprotonated CD, requiring transfer of a proton to the amino acid carboxyl group, To understand formation of complexes in the gas phase, gel permeation chromatography (GPC) was used to separate free amino acids (AAs) from complexes in an incubated solution. The ESI mass spectra of the GPC fractions show the presence of 1:1 complexes of both CD-aromatic amino acids and CD-aliphatic amino acids. Compared with CD-aliphatic amino acid complexes, CD-aromatic amino acid complexes appear to be destabilized in the gas phase, possibly because the hydrophobic interaction which binds the aromatic group of amino acids in the CD cavity in solution may become repulsive when solvent evaporates from the droplets during the electrospray process, whereas those complex ions formed as proton bound dimers are stabilized by electrostatic forces, the major binding force for such complexes in the gas phase. In addition, the GPC technique coupled with off-line ESI-MS can rapidly separate CD complexes by size, and provides some information on the character of the complexes in solution. (C) 1998 John Wiley & Sons, Ltd.

Studies on beta-cyclodextrin amino acids inclusion complexes by electrospray ionization mass spectrometry

The positive- and negative-ion electrospray ionization mass spectra of beta-cyclodextrin-amino acids complexes in NH4Ac buffer have been reported in this paper. Compared with positive-ion ESI mass spectra of beta-cyclodextrin-amino acids complexes under the same condition, negative-ion mass spectra obtained for inclusion complexes of beta-cyclodextrin (CD) with tyrosine, phenylalanine and tryptophan, respectively, were completely dominated by deprotonated complex ions and [CD-H](-) ion which is the only daughter ion in collision-induced dissociation (CID) experiment of deprotonated complexes, The results indicated that the charged position for protonated and deprotonated complexes is different from each other. In addition, two complex ions for the same complex have similarly relative dissociation energies, which are higher than that of [CD+NH4](+), indicating that complexes observed in gasphase are not electrostatic adducts at all but complexes formed by hydrogen bonds.

Rapid analysis of steroidal saponin mixture using electrospray ionization mass spectrometry combined with sequential tandem mass spectrometry

Using electrospray ionization mass spectrometry (MS) combined with sequential tandem MS(ESI-MSn), two major steroidal saponins extracted from Tribulus terrestris were studied, and considerable useful structural information was obtained. The structure of the proposed known steroidal saponin was verified, and the structure of the unknown saponin was investigated using MSn experiments. Some special fragment ions were also observed, and the corresponding fragmentation mechanisms were investigated which are characteristic for steroidal saponins and can give some information on the linkage position of some sugar groups in saponins. This methodology has been established as a powerful tool for the rapid, comparative analysis of mixtures such as crude plant extracts. (C) 1998 John Wiley & Sons, Ltd.

Determination of tetrodotoxin by liquid chromatography coupled with electrospray ion-trap mass spectrometry

Two methods for tetrodotoxin analysis using liquid chromatography coupled with electrospray iontrap mass spectrometry (LC-ESI-MS) have been established with C,, reversed phase column and hydrophilic interaction liquid chromatography (HILIC) column, respectively. Sensitivity and reproducibility of the methods were compared. The method using C-18 column in selected ion monitoring (SIM) mode had a detection limit (S/N = 3) of 120 pg, and a good linearity of the calibration curve was obtained for tetrodotoxin (r = 0. 9992). High reproducibility of the method was observed, with a relative standard deviation (RSD) below 10%. The method using HILIC column in SIM mode and selected reaction monitoring (SRM) mode had detection limits (S/N = 3) of 15 and 3.75 pg, respectively. Good linearity of the calibration curves was obtained for tetrodotoxin (r = 0. 9996 and 0. 9998 in SIM and SRM mode, respectively). T he reproducibility was high in SIM mode but relatively poor in SRM mode. Based on the results, the method using HILIC column in SIM mode was suggested for the analysis of tetrodotoxin with LC-MS system.

Determination of aliphatic amines from soil and wastewater of a paper mill by pre-column derivatization using HPLC and tandem mass spectrometry (HPLC-MS/MS)

A pre-column derivatization method for the sensitive determination of aliphatic amines using the labeling reagent 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) followed by HPLC with fluorescence detection and APCI/NIS identification in positive-ion mode has been developed. The chromophore of 2-(9-carbazole)-ethyl chloroformate (CEOC) reagent was replaced by the 1,2-benzo-3,4-dihydrocarbazole functional group, which resulted in a sensitive fluorescence derivatizing reagent, BCEOC, that could easily and quickly label amines. Derivatives were stable enough to be efficiently analyzed by HPLC and showed an intense protonated molecular ion corresponding m/z [M + H](+) with APCI/MS in positive-ion mode. The collision induced dissociation of the protonated molecular ion formed characteristic fragment ions at m/z 264.1, m/z 246.0 and m/z 218.1, corresponding to the cleavages of CH2CH2O-CO, CH2CH2-OCO, and N-CH2CH2O bonds. Studies on derivatization conditions demonstrated that excellent derivatization yields close to 100% were observed with a 3 to 4-fold molar reagent excess in acetonitrile solvent, in the presence of borate buffer (pH 9.0) at 40 degrees C for 10 min. In addition, the detection responses for BCEOC derivatives were compared with those obtained with CEOC and FMOC as labeling reagents. The ratios I-BCEOC/I-CEOC and I-BCEOC/I-FMOC were, respectively, 1.40-2.76 and 1.36-2.92 for fluorescence responses (here, I was the relative fluorescence intensity). Separation of the amine derivatives had been optimized on an Eclipse XDB-C-8 column. Detection limits calculated from an 0.10 pmol injection, at a signal-to-noise ratio of 3, were 18.65-38.82 fmol (injection volume 10 mu L for fluorescence detection. The relative standard deviations for intraday determination (n = 6) of standard amine derivatives (50 pmol) were 0.0063-0.037% for retention times and 3.36-6.93% for peak areas. The mean intra-and inter-assay precision for all amines were <5.4% and 5.8%, respectively. The recoveries of amines ranged from 96 to 113%. Excellent linear responses were observed with correlation coefficients of >0.9994. The established method provided a simple and highly sensitive technique for the quantitative analysis of trace amounts of aliphatic amines from biological and natural environmental samples.

Determination of free fatty acids in soil and bryophyte plants by precolumn derivatization via HPLC with fluorescence detection and tandem mass spectrometry (HPLC-MS/MS)

A sensitive method for the determination of 30 kinds of free fatty acids (FFAs, C-1-C-30) with 1-[2-(p-toluenesulfonate)-ethyl]-2-phenylimidazole-[4,5-f] 9,10-phenan- threne (TSPP) as labeling reagent and using high performance liquid chromatography with fluorescence detection and identification by online postcolumn mass spectrometry with atmospheric pressure chemical ionization (APCI) source in positive-ion mode (HPLC/MS/APCI) has been developed. TSPP could easily and quickly label FFAs in the presence of K2CO3 catalyst at 90 degrees C for 30 min in N,N-dimethylformamide (DMF) solvent, and maximal labeling yields close to 100% were observed with a 5-fold excess of molar reagent. Derivatives were stable enough to be efficiently analyzed by high performance liquid chromatography. TSPP was introduced into fatty acid molecules and effectively augmented MS ionization of fatty acid derivatives and led to regular MS and MS/MS information. The collision induced cleavage of protonated molecular ions formed specific fragment ions at m/z [MH](+)(molecular ion), m/z [M'+CH2CH2](+)(M' was molecular mass of the corresponding FFA) and m/z 295.0 (the, mass of protonated molecular core structure of TSPP). Fatty acid derivatives were separated on a reversed-phase Eclipse XDB-C-8 column (4.6 x 150 mm, 5 mu m, Agilent) with a good baseline resolution in combination with a gradient elution. Linear ranges of 30 FFAs are 2.441 x 10(-3) to 20 mu mol/L, detection limits are 3.24 similar to 36.97 fmol (injection volume 10 mu L, at a signal-to-noise ratio of 3, S/N 3:1). The mean interday precision ranged from 93.4 to 106.2% with the largest mean coefficients of variation (R.S.D.) < 7,5%. The mean intraday precision for all standards was < 6.4% of the expected concentration. Excellent linear responses were observed with correlation coefficients of > 0.9991. Good compositional data could be obtained from the analysis of extracted fatty acids from as little as 200 mg of bryophyte plant samples.Therefore, the facile TSPP derivatization coupled with HPLC/MS/APCI analysis allowed the development of a highly sensitive method for the quantitation of trace levels of short and long chain fatty acids from biological and natural environmental samples.

On-line concentration of proteins in pressurized capillary electrochromatography coupled with electrospray ionization-mass spectrometry

Pressurized capillary electrochromatography (pCEC) and electrospray ionization-mass spectrometry (ESI-MS) have been hyphenated for protein analysis. Taken cytochrome c, lysozyme, and insulin as samples, the limits of detection (LODs) for absolute concentrations are 10(-11) mol (signal-to-noise ratio S/N = 3) with relative standard deviations (RSDs) of retention time and peak area, respectively, of less than 1.7% and 4.8%. In order to improve the detection sensitivity, on-line concentration by field-enhanced sample-stacking effect and chromatographic zone-sharpening effect has been developed, and parameters affecting separation and detection, such as pH and electrolyte concentration in the mobile phase, separation voltage, as well as enrichment voltage and time, have been studied systematically. Under the optimized conditions, the LODs of the three proteins could be decreased up to 100-fold. In addition, the feasibility of such techniques has been further demonstrated by the analysis of modified insulins at a concentration of 20 mu g/mL.