Improvement of chemical analysis of antibiotics: XXIII. Identification of residual tetracyclines in bovine tissues by electrospray high-performance liquid chromatography–tandem mass spectrometry

To reliably identify the residual tetracycline antibiotics (TCs), oxytetracycline (OTC), tetracycline, chlortetracycline (CTC) and doxycycline (DC), in bovine tissues, we have established a confirmation method using electrospray ionization liquid chromatography–tandem mass spectrometry (ESI LC–MS–MS) with daughter ion scan. All TCs gave [M+H−NH₃]⁺ and [M+H−NH₃−H₂O]⁺ as the product ions, except for DC when [M+H]⁺ was selected as the precursor ion. The combination of C₁₈ cartridge clean-up and the present ESI LC–MS–MS method can reliably identify TCs fortified at a concentration of 0.1 ppm in bovine tissues, including liver, kidney and muscle, and has been successfully applied to the identification of residual OTC in bovine liver and residual CTC in bovine muscle samples previously found at concentrations of 0.58 ppm and 0.38 ppm by LC, respectively.     

Simultaneous determination of 18 tetrahydrocorticosteroid sulfates in human urine by liquid chromatography/electrospray ionization-tandem mass spectrometry

A liquid chromatography (LC)/electrospray ionization (ESI)-mass spectrometry (MS) method for the direct determination of eighteen tetrahydrocorticosteroid sulfates in human urine has been developed. The analytes were 3- and 21-monosulfates and 3,21-disulfates of tetrahydrocortisol (THF), tetrahydrocortisone (THE), tetrahydro-11-deoxycortisol (THS), and their corresponding 5α-H stereoisomers. The mass spectrometric behavior of these sulfates in negative-ion ESI-MS/MS revealed the production of intense structure specific product ions within the same group of sulfates and permitted distinction between regioisomeric sulfates by collision-induced fragmentation with the MS/MS technique using a linear ion-trap instrument. For the quantitative analysis, selected reaction monitoring analysis in the negative-ion detection mode using triple-stage quadrupole mass spectrometer was performed by monitoring transitions from [M−H]⁻ to the most abundant product ion of each tetrahydrocorticosteroid sulfate. After addition of 3- and 21-monosulfates of [2,2,3β,4,4-d₅]-THF, -THE, and -THS as internal standards, urine sample was applied to a solid phase extraction using a lipophilic-weak anion exchange cartridge column, and then analyzed by LC/ESI-MS/MS. The method had satisfactory performance in terms of intra- and inter-assay precision (less than 9.7% and 9.6%, respectively), and accuracy (91.2–108.2%). The limit of quantification was lower than 2.5 ng/mL for all sulfates examined. We applied this method to determine the concentration of eighteen tetrahydrocorticosteroid sulfates in the urine of healthy subjects. Thus, we have developed a sensitive, precise and accurate assay for urinary tetrahydrocorticosteroid sulfates that should be useful for clinical and biological studies.     

Development of analytical methods for some penicillins in bovine milk by ion-paired chromatography and confirmation by thermospray mass spectrometry

Analytical methods for the determination of cloxacillin, ampicillin/hetacillin, and amoxicillin in bovine milk were developed. The methods involved ultrafiltration of milk diluted with methanol, acetonitrile, and water on a 10 000-dalton cut-off filter. Separation of penicillins from other milk components was accomplished by ion-paired chromatography using a microbore column. The penicillins were detected using ultraviolet photodiode array (UV-PDA) detection and confirmed by thermospray liquid chromatography—mass spectrometry (LC—MS). The thermospray spectra of these compounds exhibited [M + H]⁺ and [M + Na]⁺ ions along with several fragment ions. The limits of detection for these antibiotics were estimated to be 50 to 100 ppb for LC with UV-PDA detection and 100–200 ppb for thermospray LC—MS detection.     

Confirmation of spectinomycin in milk using ion-pair solid-phase extraction and liquid chromatography–electrospray ion trap mass spectrometry

A confirmation procedure is described for residues of spectinomycin in bovine milk. Spectinomycin is extracted from raw milk using ion-pair reversed-phase solid-phase extraction. The extracts are ion-pair chromatographed on a polymeric reversed-phase column and analyzed on a quadrupole ion trap mass spectrometer equipped with an electrospray interface. MS–MS data are acquired in the scan mode of product ions deriving from m/z 333, the protonated molecular ion. The estimated limit of confirmation is between 0.05 and 0.1 μg/ml. The procedure was validated with control milk, fortified milk (0.1–5.0 μg/ml), and milk from cows dosed with spectinomycin.     

The effect of silver and other metal ions on the in vitro growth of root-rotting Phytophthora and other fungal species

A range of metal ions and the oxoanion WO₄^2-were toxic to zoospores of Phytophthora nicotianae parasitica in the order: Ag⁺ > Cu⁺⁺ > WO₄^2-> Ni⁺ > Co⁺⁺ > Zn⁺. The LD₅₀ for Ag⁺, 0.11 μM (11.4 ppb), compared with 1.84 μm (117 ppb) for Cu⁺⁺. Silver was similarly toxic to a range of pathogens including Pythium aphanidermatum, Thielaviopsis basicola and Fusarium oxy-sporum f.spp. Most zoospores of Phytophthora spp. were killed by Ag⁺ in the range 46–460 nM (5–50 ppb), bursting at the higher concentrations. A small sub-population of most propagules exhibited greater tolerance to silver than the whole. In 0.93 μM (100 ppb) Ag⁺ 1.4% of P. nicotianae parasitica zoospores survived but were all killed at 500 ppb. A population of P. cryptogea (1.9%) surviving 0.47 μm (50 ppb) were killed at 0.93 μM (100 ppb). Zoospore cysts and germlings showed the same sensitivity to silver. Oospores were mostly killed over the range 0.23–0.93 μm (25–100 ppb) Ag⁺, some surviving up to the lethal concentration of 9.26 μM (1000 ppb). Mycelium of P. cryptogea was generally less sensitive, with some growth occurring at 9.26 μm (100 ppb). Zoosporangiogenesis was unaffected over the range 0.47–4.65 μm (50–500 ppb). Toxicity increased with increased pH over the range 5.0–6.5. Ionic silver was lost from solution during a microscope slide bioassay by binding to the glass surface. In the presence of chloride ions, colloidal AgCl formed which was equally toxic to P. cryptogea. Silver and AgCl were further lost from solution by colloidal agglomeration - Ostwald ripening - and by AgCl adsorption to glass. Silver, < 90 nM (10 ppb) Ag⁺ as AgNO₃ and particles of silver chloride were both strongly attractive to zoospores of P. cryptogea. Spores burst or failed to germinate on entering lethal concentrations. The results are discussed in the context of the use of silver salts to control Phytophthora root-rot pathogens and the importance of ion availability in in vitro toxicity assays.     

Improvement of chemical analysis of antibiotics: XXIII. Identification of residual tetracyclines in bovine tissues by electrospray high-performance liquid chromatography–tandem mass spectrometry

To reliably identify the residual tetracycline antibiotics (TCs), oxytetracycline (OTC), tetracycline, chlortetracycline (CTC) and doxycycline (DC), in bovine tissues, we have established a confirmation method using electrospray ionization liquid chromatography–tandem mass spectrometry (ESI LC–MS–MS) with daughter ion scan. All TCs gave [M+H−NH₃]⁺ and [M+H−NH₃−H₂O]⁺ as the product ions, except for DC when [M+H]⁺ was selected as the precursor ion. The combination of C₁₈ cartridge clean-up and the present ESI LC–MS–MS method can reliably identify TCs fortified at a concentration of 0.1 ppm in bovine tissues, including liver, kidney and muscle, and has been successfully applied to the identification of residual OTC in bovine liver and residual CTC in bovine muscle samples previously found at concentrations of 0.58 ppm and 0.38 ppm by LC, respectively.     

Quantitative determination of helicid in rat biosamples by liquid chromatography electrospray ionization mass spectrometry

A simple liquid chromatography electrospray ionization mass spectrometry (LC–ESI–MS) method with highly improved sensitivities for the determination of helicid in rat bile, urine, feces and most tissues was developed. The tissues and feces were firstly homogenized mechanically using deionized water as the media. Bile, urine, tissues and feces homogenates were extracted by liquid–liquid extraction with n-butyl alcohol for sample preparation. The subsequent analysis procedures were performed on a Shimadzu LCMS2010A system (electrospray ionization single quadrupole mass analyzer). A Luna C₁₈ column (150 mm × 2.00 mm, 5 μm) was used as the analytical column, while a mixture of acetonitrile and ammonium chloride water solution was used as the mobile phase. The proportions of mobile phase were changed timely according to gradient programs. Chlorinated adducts of molecular ions [M+Cl]^? at m/z 319.00 and 363.05 were used to quantify helicid and bergeninum (internal standard), respectively. The method was validated to be accurate, precise and rugged with good linearity. The proposed method was successfully applied to the preclinical tissue distribution and excretion studies of helicid in rats.     

Simultaneous quantitative determination of olmesartan and hydrochlorothiazide in human plasma and urine by liquid chromatography coupled to tandem mass spectrometry

A specific, sensitive and rapid method based on high performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) was developed for the simultaneous determination of olmesartan (OLM) and hydrochlorothiazide (HCTZ) in human plasma and urine. Solid-phase extraction (SPE) was used to isolate the analytes from biological matrices followed by injection of the extracts onto a C₁₈ column with isocratic elution. Detection was carried out on a triple quadrupole tandem mass spectrometer in multiple reaction monitoring (MRM) mode using negative electrospray ionization (ESI). The method was validated over the concentration range of 1.00–1000 ng/mL and 5.00–5000 ng/mL for OLM in human plasma and urine as well as 0.500–200 ng/mL and 25.0–25,000 ng/mL for HCTZ in human plasma and urine, respectively. Inter- and intra-run precision of OLM and HCTZ were less than 15% and the accuracy was within 85–115% for both plasma and urine. The average extraction recoveries were 96.6% and 92.7% for OLM, and 87.2% and 72.1% for HCTZ in human plasma and urine, respectively. The linearity, recovery, matrix effect and stability were validated for OLM/HCTZ in human plasma and urine.     

Analysis of sesterterpenoids from Aspergillus terreus using ESI‐QTOF and ESI‐IT

Introduction – Biosynthesis of terretonin was studied due to the interesting skeleton of this series of sesterterpenoids. Very recently, López‐Gresa reported two new sesterterpenoids (terretonins E and F) which are inhibitors of the mammalian mitochondrial respiratory chain. Mass spectrometry (MS), especially tandem mass spectrometry, has been one of the most important physicochemical methods for the identification of trace natural products due to it rapidity, sensitivity and low levels of sample consumption. The potential application prospect and unique skeleton prompted us to study structural characterisation using MS. Objective – To obtain sufficient information for rapid structural elucidation of this class of compounds using MS. Methodology – The elemental composition of the product ions was confirmed by low‐energy ESI‐CID‐QTOF‐MS/MS analyses. The fragmentation pathways were postulated on the basis of ESI‐QTOF‐MS/MS/MS and ESI‐IT‐MSⁿ spectra. Common features and major differences between ESI‐QTOF‐MS/MS and IT‐MSⁿ spectra were compared. For ESI‐QTOF‐MS/MS/MS experiments, capillary exit voltage was raised to induce in‐source dissociation. Ammonium acetate or acetic acid were added into solutions to improve the intensity of [M + H]⁺. The collision energy was optimised to achieve sufficient fragmentation. Some fragmentation pathways were unambiguously proposed by the variety of abundance of fragment ions at different collision energies even without MSⁿ spectra. Results – Fragmentation pathways of five representative sesterterpenoids were elucidated using ESI‐QTOF‐MS/MS/MS and ESI‐IT‐MSⁿ in both positive‐ and negative‐ion mode. The key group of characterising fragmentation profiles was ring B, and these fragmentation patterns are helpful to identify different types of sestertepenoids. Conclusion – Complementary information obtained from fragmentation experiments of [M + H]⁺ (or [M + NH₄]⁺) and [M ? H]^? precursor ions is especially valuable for rapid identification of this kind of sesterterpenoid.     

Multi-residue extraction–purification procedure for corticosteroids in biological samples for efficient control of their misuse in livestock production

A fast and efficient multi-residue extraction–purification procedure was developed for 12 corticosteroids in biological matrices (hair, urine and meat), in order to control their illegal use as growth promoters in cattle. Detection and identification of the analytes were achieved using a previously described LC–MS–MS method based on negative electrospray ionisation and a triple quadrupole analyser. The presented procedures included acid (hair) or enzymatic (urine and meat) hydrolysis, C₁₈ reversed-phase SPE, Na₂CO₃ liquid–liquid clean-up and SiOH normal-phase SPE. The detection limits of the developed methods were between 2.9 and 9.3 pg/mg (ppb) for hair samples and in the 40 – 70 pg/g (ppt) range for the urine or meat samples. The acid hydrolysis used for corticosteroid extraction in hair was optimised using an experimental design and response surface methodology. Achieved performances were linked to a physico–chemical approach based on the corticosteroids specific C₁₇ side-chain. This original multi-residue and multi-matrices analytical methodology will be used for further metabolism studies.     

Quantitative liquid chromatographic–tandem mass spectrometric determination of orlistat in plasma with a quadrupole ion trap

This report evaluates the use of a quadrupolar ion trap for quantitation in a bioanalytical laboratory. The evaluation was accomplished with the cross-validation of an LC–MS–MS quantitative method previously validated on a triple quadrupole mass spectrometer. The method was a multi-level determination of the anti-obesity drug, orlistat, in human plasma. The method has been refined previously on a triple quadrupole instrument to provide rapid sample throughput with robust reproducibility at sub-nanogram detection limits. Optimization of the method on the ion trap required improved chromatographic separation of orlistat from interfering plasma matrix components coextracted during the initial liquid–liquid extraction of plasma samples. The ion trap produces full-scan collision-induced dissociation mass spectra containing characteristic orlistat fragment ions that are useful for quantitation. Data collection on the ion trap required a precursor ion isolation width of 3.0 Da and optimal quantitative results were obtained when three fragment ions were monitored with a 1.8 Da window for each ion. Although a direct cross-validation between the ion trap and the tandem triple quadrupole mass spectrometer was not possible, quantitative results for orlistat comparable to those obtained from the triple quadrupole instrument were achieved by the ion trap with the modified method. The limit of quantitation for orlistat in plasma on the ion trap was 0.3 ng ml⁻¹ with a linear dynamic range of 0.3 to 10 ng ml⁻¹. Precision and accuracy varied from 4 to 15% over the quantitation range. The overall results provide an example of the utility of an ion trap in bioanalytical work.     

Determination of ZD9583, a thromboxane receptor antagonist, in human plasma and urine by liquid chromatography atmospheric pressure chemical ionization tandem mass spectrometry

A liquid chromatography–atmospheric pressure chemical ionization tandem mass spectrometry (LC–APCI-MS–MS) method is described for the determination of a thromboxane receptor antagonist (4Z)-6-((2S,4S,5R)-2-(1-(2-cyano-4-methylphenoxy)-1-methylethyl)-4-(3-pyridyl)-3-dioxan-5-yl)hex-4-enoic acid (ZD9583, I) in human plasma and urine. Proteins in plasma and urine samples are precipitated using acidified acetonitrile. The resulting supernatant is chromatographed on a C₈ reversed-phase chromatography column. Following the diversion of the solvent front from the mass spectrometer by a switching valve, the column eluate is passed on to the mass spectrometer via a heated nebulizer interface where the analyte is detected by multiple reaction monitoring (MRM). The method has a chromatographic run time of less than 2 min, a linear calibration curve with a range of 1–500 ng ml⁻¹ and intra- and inter-day precision estimates of less than 10% over the calibration range.     

Structural characterization of neutral glycosphingolipids using high-performance liquid chromatography-electrospray ionization mass spectrometry with a repeated high-speed polarity and MS<Superscript>n</Superscript> switching system

Four types of neutral glycosphingolipids (LacCer, Gb₃Cer, Gb₄Cer, and IV³αGalNAc-Gb₄Cer; 10 pmol each) were analyzed using high-performance liquid chromatography (HPLC)-electrospray ionization quadrupole ion trap time-of-flight (ESI-QIT-TOF) mass spectrometry (MS) with a repeated high-speed polarity and MSⁿ switching system. This system can provide six types of mass spectra, including positive and negative ion MS, MS², and MS³ spectra, within 1 s per cycle. Using HPLC with a normal-phase column, information on the molecular weights of major molecular species of four neutral glycosphingolipids was obtained by detecting [M+Na]⁺ in the positive ion mode mass spectra and [M?H]^? in the negative ion mode mass spectra. Sequences of glycosphingolipid oligosaccharide were obtained in the negative ion MS² spectra. In addition, information on the ceramide structures was clearly obtained in the negative ion MS³ mass spectra. GlcCer molecular species were analyzed by HPLC-ESI-QIT-TOF MS with a reversed-phase column using 1 pmole of GlcCer. The structures of the seven molecular species of GlcCer, namely, d18:1-C16:0, d18:1-C18:0, d18:1-C20:0, d18:1-C22:0, d18:1-C23:0, d18:1-C24:1, and d18:1-C24:0, were characterized using positive ion MS and negative ion MS, MS², and MS³. The established HPLC-ESI-QIT-TOF MS with MSⁿ switching and a normal phase column has been successfully applied to the structural characterization of LacCer and Gb₄Cer in a crude mixture prepared from human erythrocytes.     

Structure elucidation of ostreocin-A and ostreocin-E1, novel palytoxin analogs produced by the dinoflagellate Ostreopsis siamensis,using LC/Q-TOF MS

Palytoxin analogs are marine toxins with large complex polyol structures. A benthic dinoflagellate Ostreopsis siamensis produces more than ten palytoxins (ostreocins, OSTs). The limited sample availability of minor OSTs restricts the definition of their chemical structures. The present investigation characterizes structures of two minor OSTs, i.e., ostreocin-A (OSTA) and ostreocin-E1 (OSTE1), using ostreocin-D (OSTD) as a reference compound, by liquid chromatography/quadrupole-time-of-flight mass spectrometry. The molecular formulas of OSTA and OSTE1 were C₁₂₇H₂₁₉N₃O₅₄ and C₁₂₇H₂₁₇N₃O₅₂, respectively. Compared to OSTD, OSTA has an extra oxygen atom whereas OSTE1 lacks one oxygen atom and two hydrogen atoms. The MS/MS experiments (precursor ions: [M + H]⁺ and [M-H]^?) suggested a hydroxyl substitution at C82 in OSTA and alteration(s) between C53 and C100 in OSTE1. Further analysis of structural details in OSTE1 was performed through a pseudo-MS³ experiment (precursor ion: m/z 1432.748). Accordingly, the planar structures of OSTA and OSTE1 were assigned to 42,82-dihydroxy-3,26-didemethyl-19,44-dideoxypalytoxin and 42-hydroxy-3,26-didemethyl-19,44,73-trideoxypalytoxin-72-ene, respectively.

Abbreviations:CID: collision induced dissociation; HR-LC/MS/MS: high-resolution liquid chromatography/tandem mass spectrometry; LC/ESI/Q-TOF MS: liquid chromatography/quadrupole time-of-flight mass spectrometry equipped with an electrospray ionization source; NMR: nuclear magnetic resonance; OSTs: ostreocins; OSTA: ostreocin-A; OSTB: ostreocin-B; OSTD: ostreocin-D; OSTE1: ostreocin-E1; OVTX-a: ovatoxin-a; OVTXs: ovatoxins; PLTX: palytoxin     

A Novel Geometry Mass Spectrometer, the Q-TOF, for Low-Femtomole/Attomole-Range Biopolymer Sequencing

Ultra-high-sensitivity, biopolymer sequencing is a goal in many fields of molecular biology, and collisionally activated decomposition electrospray mass spectrometry (CAD ES MS/MS) using a triple quadrupole mass spectrometer has become a method of choice for work in the high- to mid-femtomole range. However, when the detection of ions becomes statistical, as it may in that range, the mass assignment of fragment ions is inaccurate and either sequencing becomes impossible or ambiguities result due, for example, to the closeness in amino acid residue masses (I/L, N or K/Q, E). Some ambiguities may be resolved by synthesizing possible sequences, but this is unsatisfactory. In considering the limitations of triple quadrupole MS/MS with respect to scanning ion detection, resolution, transmission, and mass accuracy, we reasoned that a novel geometry quadrupole orthogonal acceleration time-of-flight (Q-TOF) instrument would have special merit for ultra-high-sensitivity MS/MS sequencing, and suggested its construction for this purpose some three years ago. A prototype Q-TOF has now been built by Micromass [Morris et al. (1996), Rapid Commun. Mass Spectrom. 10, 889–896], and in the first research on the instrument, including MHC antigen and filarial nematode glycoprotein studies, we demonstrate low-femtomole- and attomole-range sequencing with mass accuracy of better than 0.1 Da throughout the daughter-ion spectrum, thus removing sequencing ambiguities in some of the most challenging work demanding the highest sensitivity.     

Design of a stable isotope dilution gas chromatography/mass spectrometric assay for cAMP: Comparison with standard protein-binding and radioimmunoassay methods

A stable isotope dilution assay is presented in which picomole quantities of cAMP can be determined with high precision and selectivity using gas chromatography and electron impact mass spectrometry with multiple ion detection techniques. Using synthetic [2,8-²H₂,6-¹⁵N]-cAMP as the internal standard, suitable specificity was obtained by monitoring the (MCH₃)⁺ fragment ions of the trimethylsilyl derivatives of cAMP and the internal standard at $\text{m}\text{z}$ 530 and $\text{m}\text{z}$ 533, respectively. The sensitivity of the assay as judged from the lower limit of detection of the mass spectrometer was 3.0 pmol. Rat liver and human urine cAMP levels were assayed using gas chromatography/mass spectrometry and were compared with levels determined by protein-binding assays and radioimmunoassays for the same samples. The intraassay coefficients of variation of the gas chromatography/mass spectrometry assay were 5.3% for the rat liver sample (cAMP level 832 pmol/g) and 6.0% for the urine sample (cAMP level 2.50 μmol/liter). Comparison of the levels of cAMP determined by the three assay methods showed correlation to within 10% variation.     

Stable isotope dilution quantification of mutagens in cooked foods by combined liquid chromatography-thermospray mass spectrometry

A method of general applicability for the detection and quantification of mutagens in cooked foods at the ppb level is presented. A minimal sample prefractionation is employed and [Me-²H₃-labeled analogs of the compounds of interest are added for identification and quantification of mutagens by accurate measurement of chromatographic retention (K′) in reverse-phase high-performance liquid chromatography (HPLC), and by measurement of the ratio of response of the protonated molecular ions of analyte and internal standard by directly coupled liquid chromatography-mass spectrometry (LC/MS). Initial application is demonstrated in the analysis of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quonoline (MeIQ) in broiled salmon. Measured levels of IQ and MeIQ in broiled salmon flesh were 0.3–1.8 ppb and 0.6–2.8 ppb, respectively, and for the skin of broiled salmon 1.1–1.7 ppb and 1.5–3.1 ppb, respectively. Results on cooked beef and sardine are also reported.     

Application of gas chromatography—mass spectrometry with selected ion monitoring to the urinanalysis of 4-pyridoxic acid

An analytical protocol has been developed for the analysis of urinary 4-pyridoxic acid (4-PA) by gas chromatography—mass spectrometry (GC—MS) for use in metabolic studies. Aliquots of urine were deproteinised and fractionated by isocratic reversed-phase high-performance liquid chromatography. The eluent fraction containing the 4-PA was collected, freeze-dried and silylated using N-methyl-N-(tert.-butyldimethylsilyl)trifluoroacetamide. Derivatisation produced the mono-tert.-butyldimethylsilyl derivative of 4-PA lactone. This derivative was readily amenable to GC—MS analysis in the electron ionisation (70 eV) mode, yielding a prominent fragment ion at m/z 222 ([M — 57]⁺; base peak). A heavy isotope-labelled derivative of pyridoxine [dideuteriated pyridoxine; 3-hydroxy-4-(hydroxymethyl)-5-[hydroxymethyl-²H₂]-2-methylpyridine] has been synthesised and is being employed to determine the kinetics of labelling of the body pools of vitamin B₆. Kinetic measurements are based on the determination of the relative proportions of metabolically produced deuterium-labelled and non-labelled 4-PA in urine, obtained from stable isotope ratios determined by low-resolution selected ion monitoring using a bench-top quadrupole GC—MS system.     

Emerging Lyngbya wollei toxins: A new high resolution mass spectrometry method to elucidate a potential environmental threat

Mass spectrometric methods for the quantitative and qualitative analyses of algal biotoxins are often complicated by co-eluting compounds that present analytically as interferences. This issue is particularly critical for organic polyamines, where co-eluting materials can suppress the formation of cations during electrospray ionization. Here we present an extraction procedure designed specifically to overcome matrix-derived ion suppression of algal toxins in samples of Lyngbya wollei, a filamentous benthic algae known to produce several saxitoxin analogues. Lyngbya wollei samples were collected from a large, persistent harmful algal bloom in Lake Wateree, SC. Six known Lyngbya wollei-specific toxins (LWT1–6) were successfully resolved and quantified against saxitoxin using hydrophilic interaction liquid chromatography coupled with triple quadrupole and quadrupole time-of-flight mass spectrometry. The parent ions [M²⁺ – H⁺]⁺ were observed for LWTs 1–6 and the [M]²⁺ ion was observed for LWT5. High resolution mass spectra and unique fragmentation ions were obtained for LWTs 1–6. A dilution factor of 50 resulted in a linear calibration of saxitoxin in the algae matrix. Ion suppression was resolved by sample dilution, which led to linear, positive correlations between peak area and mass of the extracted sample (R² > 0.96). Optimized sample extraction method and instrument parameters are presented.