Adduct simplification in the analysis of cyanobacterial toxins by matrix‐assisted laser desorption/ionization mass spectrometry

Abstract

A novel method for simplifying adduct patterns to improve the detection and identification of peptide toxins using matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry is presented. Addition of 200 microM zinc sulfate heptahydrate (ZnSO(4) . 7H(2)O) to samples prior to spotting on the target enhances detection of the protonated molecule while suppressing competing adducts. This produces a highly simplified spectrum with the potential to enhance quantitative analysis, particularly for complex samples. The resulting improvement in total signal strength and reduction in the coefficient of variation (from 31.1% to 5.2% for microcystin-LR) further enhance the potential for sensitive and accurate quantitation. Other potential additives tested, including 18-crown-6 ether, alkali metal salts (lithium chloride, sodium chloride, potassium chloride), and other transition metal salts (silver chloride, silver nitrate, copper(II) nitrate, copper(II) sulfate, zinc acetate), were unable to achieve comparable results. Application of this technique to the analysis of several microcystins, potent peptide hepatotoxins from cyanobacteria, is illustrated.

Impairment of the mitochondrial oxidative phosphorylation system and oxidative stress in liver of crucian carp (Carassius auratus L.) exposed to microcystins

“…In recent decades, cyanobacterial blooms in water have generated great public concerns worldwide due to their ability to produce toxic substances known as cyanotoxins. Among these toxins, microcystins (MCs) are considered to be the most common and potent (Howard and Boyer, ). Many studies have confirmed the toxicity of MCs to different organisms including humans (Jochimsen et al, ; Chen et al, ; He et al, ; Rogers et al, ).…”

Direct Coupling of High-Performance Thin-Layer Chromatography with UV Spectroscopy and IR-MALDI Orthogonal TOF MS for the Analysis of Cyanobacterial Toxins

“…Furthermore, a significantly reduced sensitivity has been reported for UV-MALDI MS of HPTLC-separated oligosaccharides as a result of a high background level over the whole mass range as compared to IR-MALDI MS. 49 It is noteworthy that in the case of both analyte species, no fragment ions were observed under the mass spectrometric conditions used, and this contrasts to the results obtained previously using UV-MALDI MS where fragmentation of the Adda side chain has been observed. 38 This finding points to the mild desorption/ionization conditions of the IR-MALDI process employed in this study. Furthermore, the formation of dimeric gasphase ions points to the moderate excess of internal energy deposited in the analyte ions.…”

“…Photometric analysis yielded considerably reduced peak areas compared to UV absorption of the unstained bands at λ ) 232 nm (data not shown). 38 The 10-fold magnified region contains peaks assigned with numbers 5-10 representing gas-phase dimers. Protonated species with low abundance [2M + H] + (peak 5) and dimers of MC LR with 1 to as many as 5 sodium ions attached are present (peaks 6-10).…”

Enhanced Sample Preparation for Quantitation of Microcystins by Matrix‐Assisted Laser Desorption/Ionisation–Time of Flight Mass Spectrometry

“…All preparations used CHCA as a matrix and either a 384‐spot polished steel MALDI–TOF target or an anchorchip™ (600 µm; Bruker Daltonics, Bremen, Germany). Angiotensin I was utilised as an internal standard to enhance reproducibility and zinc sulphate heptahydrate was added to eliminate sodium and potassium adducts (Howard and Boyer, ). MC standard ([Dha 7 ] MC‐LR, MC‐YR or MC‐RR) were dissolved in 40% methanol and on the day of the analyses were mixed 9:1 with 500 µM angiotensin I/50 mM Zn 2+ (IS), to yield final concentrations of 50 µM and 5 mM, respectively.…”

“…Values expressed in this study were based on the [M + H] + monoisotopic peak and did not include any adduct peaks. Only the peak height was utilised in accordance with the findings of previous studies (Nelson et al , ; Howard and Boyer, ). The peak height ratio (PHR) was found by dividing the MC signal by that of the internal standard.…”