Stereoselective Chemical Ionization Mass Spectrometry: Reactions of CH 3 OPOCH 3 + with Cyclic Vicinal Diols

A rapid and accurate method of quantifying positional isomeric mixtures of phosphorylated hexose and N-acetylhexosamine monosacchrides by using gas-phase ion/molecule reactions coupled with FT-ICR mass spectrometry is described. Trimethyl borate, the reagent gas, reacts readily with the singly charged negative ions of phosphorylated monosaccharides to form two stable product ions corresponding to the loss of one or two neutral molecules of methanol from the original adduct. Product distribution in the ion/molecule reaction spectra differs significantly for isomers phosphorylated in either the 1-or the 6-position. As a result, the percents of total ion current of these product ions for a mixture of the two isomers vary with its composition. In order to determine the percentage of each isomer in an unknown mixture, a multicomponent quantification method is utilized in which the percents of total ion current of the two product ions for each pure monosaccharide phosphate and the mixture are used in a two-equation, two-unknown system. The applicability of this method is demonstrated by successfully quantifying mock mixtures of four different isomeric pairs: Glucose-1-phosphate and glucose-6-phosphate; mannose-1-phosphate and mannose-6-phosphate; galactose-1-phosphate and galactose-6-phosphate; N-acetylglucosamine-1-phosphate and N-acetylglucosamine-6-phosphate. The effects of mixture concentrations and ion/molecule reaction conditions on the quantification are also discussed. Our results demonstrate that this assay is a fast, sensitive, and robust method to quantify isomeric mixtures of phosphorylated monosaccharides. . In particular, the phosphate linkage position of carbohydrates is important to regulate biological function. For example, the conversion between the 6-phosphate and the 1-phosphate isomers of several monosaccharides, which is regulated by the corresponding phosphate mutases, plays an important role in the glycosylation of glycoproteins [5,6]. Specifically, the deficiency of the enzyme phosphomannomutase, which catalyzes the conversion of mannose-6-phosphate to mannose-1-phosphate in the early step of biosynthesis of lipid-linked oligosaccharides, is responsible for a disease called CDG-1a [1], in which glycoproteins with truncated N-linked carbohydrates are formed.Despite their importance, analysis of phosphorylated carbohydrates by traditional methods has been problematic due to their structural variety and non-characteristic UV absorbance. As a result, traditional enzyme kinetic measurements of the phosphate mutases mentioned above require up to three coupling enzymes [7][8][9]. Recently, Turecek et al. [10] reported an affinity capture and elution/electrospray ionization mass spectrometry assay of phosphomannomutase and phosphomannose isomerase. In their method, the product isomer was subjected to another enzymatic reaction that altered its mass in order to be detected by mass spectrometry. Direct quantitative analysis of phosphorylated glucose has been reported using anion-exchange chromatography ...

Section: Differentiation Of Phosphorylated Isomers Through the Use Ofsupporting

confidence: 82%

mentioning

confidence: 99%

Investigation of ion/molecule reactions as a quantification method for phosphorylated positional isomers: An FT-ICR approach

Gao

Kim

Leavell

et al. 2003

“…This is likely due to the fact that the phosphate and the C(2)-OH are anti in the Man1P isomer and syn in the Glc1P and Gal1P isomers. Similar axial and equitoral positioning of hydroxyl groups have been found to be important in determining the stereochemistry of hexose isomers upon CID in the analysis of transition metal-ligand derivatized monosaccharides [34 -37] and in the stereochemical analysis of cyclic diols [20,21,38]. Thus, it appears that the stereochemistry of the hydroxyl group adjacent to the phosphate plays a role in the product ions observed upon subjecting the [HexP/TMSCl-HCl] Ϫ complexes to collisionally activated conditions.…”

Section: Stabilization Of Phosphates and Determination Of Phosphate Pmentioning

confidence: 69%

“…The analytical application of ion/molecule reactions has long been realized and is currently of much interest as is evidenced by two recent reviews [18,19]. Other recent examples include those by Kenttämaa and coworkers who utilized ion/molecule reactions to differentiate isomeric species such as diols [20,21] and steroids [22]. In other studies, ion/molecule reactions have been utilized to develop a potential gas phase sequencing method for peptides [23], to determine enantiomeric excess [24 -26], and to explore non-covalent interactions [27].…”

mentioning

confidence: 99%

Probing isomeric differences of phosphorylated carbohydrates through the use of ion/molecule reactions and FT-ICR MS

Leavell

Leary

2003

Through the use of ion/molecule reactions and tandem mass spectrometry, phosphate position is assigned in both phosphorylated monosaccharides and oligosaccharides. In previous work [1, 2] phosphate moieties of monosaccharides were stabilized under collisional activation, by first derivatizing the deprotonated monosaccharide with trimethyl borate through an ion/molecule reaction, and the phosphate position determined through marker ions generated in tandem mass spectra. In this work, the methodology is extended to larger phosphorylated oligomers employing chlorotrimethylsilane (TMSCl) as the ion/molecule reagent. Phosphorylated monosaccharides were first investigated to determine diagnostic ions for phosphate linkage in monomeric standards. It was observed that the diagnostic ions showed both linkage and some monosaccharide stereochemical information. Furthermore, it was observed that TMS addition stabilized the phosphate moiety under collisionally activated conditions. Upon identification of the diagnostic ions, the methodology was applied to lactose-1-phosphate. It was found that TMSCl, stabilized the phosphate moiety upon collisional activation, and furthermore, the phosphate linkage could be determined through tandem mass spectrometric analysis. As a further extrapolation to biologically relevant problems, the methodology was applied to a lipophosphoglycan analog from the protozoan parasite Leishmania. This sample contains bridging phosphates which were converted to terminal phosphates through collision induced dissociation. The sample was then analyzed in the same manner as lactose-1-phosphate, yielding phosphate linkage information and stereochemical information. This study showed that, using the developed methodology, phosphate linkage can be determined from both monosaccharides and larger oligosaccharides; furthermore it is applicable to samples in which the phosphates are either terminating or bridging. (J Am Soc Mass Spectrom 2003, 14, 323-331)

“…The manifold and ion source temperatures were maintained at 150°C and 70°C, respectively. The CH 3 P(O)OCH 3 ϩ (or CH 3 OP(O)OCH 3 ϩ ) ions were generated by 70 eV electron ionization (EI) of DMMP (Lancaster, Pelham, NH) [35] and trimethyl phosphate [36]. Ion/molecule reactions were performed by mass selecting the precursor ions in Q1, reacting these ions with the neutral reagent in Q2, and then mass analyzing the product ions in Q3.…”

Section: Methodsmentioning

confidence: 99%

Meerwein reaction of phosphonium ions with epoxides and thioepoxides in the gas phase

Meurer

Chen

Riter

et al. 2004

Phosphonium ions are shown to undergo a gas-phase Meerwein reaction in which epoxides (or thioepoxides) undergo three-to-five-membered ring expansion to yield dioxaphospholanium (or oxathiophospholanium) ion products. When the association reaction is followed by collision-induced dissociation (CID), the oxirane (or thiirane) is eliminated, making this ion molecule reaction/CID sequence a good method of net oxygen-by-sulfur replacement in the phosphonium ions. This replacement results in a characteristic mass shift of 16 units and provides evidence for the cyclic nature of the gas-phase Meerwein product ions, while improving selectivity for phosphonium ion detection. This reaction sequence also constitutes a gas-phase route to convert phosphonium ions into their sulfur analogs. Phosphonium and related ions are important targets since they are commonly and readily formed in mass spectrometric analysis upon dissociative electron ionization of organophosphorous esters. The Meerwein reaction should provide a new and very useful method of recognizing compounds that yield these ions, which includes a number of chemical warfare agents. The Meerwein reaction proceeds by phosphonium ion addition to the sulfur or oxygen center, followed by intramolecular nucleophilic attack with ring expansion to yield the 1,3,2-dioxaphospholanium or 1,3,2-oxathiophospholanium ion. Product ion structures were investigated by CID tandem mass spectrometry (MS 2 ) experiments and corroborated by DFT/HF calculations. (J Am Soc Mass Spectrom 2004, 15, 398 -405)