Application of Capillary Zone Electrophoresis for the Analysis of Low Molecular Weight Organic Acids in Environmental Samples

Abstract

The applicability of a recently developed capillary zone electrophoretic (CZE) method for the determination of low molecular weight (LMW) organic acids in water was tested on five types of environmental samples (rainwater, water extract from peat, and soil water from two polluted sites). A full baseline resolved separation of fourteen commonly found LMW carboxylic acids in natural waters (malonic, oxalic, fumaric, maleic, formic, succinic, tartaric, glutaric, adipic, acetic, propionic, butyric, valeric and citric acids), was achieved within eight minutes. The limits of detection (2 X noise) were in the ranges 90–200 μg/l and 0.5 – 5 μg/l for hydrodynamic and electrokinetic injection, respectively. Two different CZE systems, a Waters Quanta 4000 and a Hewlett Packard HP ^3DCE system, were used and their performance compared.     

Determination of Carboxylic Acids from Plant Root Exudates by Ion Exclusion Chromatography with ESI-MS

Ion-exclusion chromatography (IEC) coupled with electrospray ionization quadrupole mass spectrometry (ESI-MS) has been used for the analysis of carboxylic acids. The use of ESI-MS provides increased specificity and sensitivity compared to existing detection methods. This paper applies IEC-ESI-MS to the analysis of carboxylic acids in commonly found root exudates and shows that the separation of nine carboxylic acids (pyruvic, oxalic, lactic, malonic, maleic, succinic, tartaric, aconitic and citric acids) can be achieved by IEC within 8 min. The ESI provided reasonable signals from negative ions, [M−H]⁻ in the negative ionization mode. Linear plots of peak area versus concentration were obtained in the range 50–25,000 μg L⁻¹ for MS detection under optimized MS conditions. The detection limits of target organic acids, based upon signal to noise ratio (S/N = 3), ranged from 10 to 30 μg L⁻¹. The reproducibility of peak areas was <2.5% (n = 5). The proposed method was used for the confirmation and quantification of carboxylic acids in nutrient solutions containing root exudates.     

Analysis of low-molecular-mass organic acids using capillary zone electrophoresis-electrospray ionization mass spectrometry

A capillary zone electrophoresis-electrospray ionization-mass spectrometry (CZE-ESI-MS) method was developed to facilitate identification and determination of eleven low-molecular-mass (LMW) organic acids (i.e. oxalic, lactic, malonic, maleic, citric, tartaric, adipic, glutaric, gluconic, isosaccharinic and succinic acid) in different sample matrices. This CZE method was adapted to suit MS conditions. Sheath liquid, sheath flow and MS parameters were optimized to achieve high mass spectrometric sensitivity. The CZE-ESI-MS procedure showed good sensitivity (limit of detection of < 0.05-0.7 mg/l for all acids), linearity (r2 = 0.9925-0.9998) and reproducibility (2.09-5.34% RSD). The applicability of the CZE-ESI-MS was demonstrated on LMW organic acids in an ale sample. In addition the (here presented) method also provided quantification of fumaric, galacturonic and 2-ketoglutaric acid with high sensitivity.     

Confirmation and determination of carboxylic acids in root exudates using LC-ESI-MS

Reversed-phase liquid chromatography with UV detection is of limited applicability in the separation and identification of carboxylic acids because of the column's poor separation efficiency and the non-selective nature of the UV detector. To address this issue, RP-LC with electrospray ionization mass spectrometry has been explored for the confirmation and determination of carboxylic acids in plant root exudates, with ESI-MS providing structural information, high selectivity, and high sensitivity. The separation of 10 carboxylic acids (pyruvic, lactic, malonic, maleic, fumaric, succinic, malic, tartaric, trans-aconitic, and citric acid) was performed on a C(18) column using an eluent containing 0.1% (v/v) acetic acid within 10 min, where the acidic eluent not only suppressed the ionization of the carboxylic acids to be retained on the column, but was also compatible with ESI-MS detection. In addition, an additional standard was used to overcome the matrix effect. The results showed that peak areas correlated linearly with the concentration of carboxylic acids over the range 0.05-10 mg/L. The detection limits of target acids (signal-to-noise S/N ratio of 3) ranged from 20 to 30 microg/L. Finally, the proposed method was used for the confirmation and determination of low-molecular-weight carboxylic acids in plant root exudates, and provided a simple analytical procedure, including sample processing, fast separation, and high specificity and sensitivity.     

胶束电动毛细管色谱-间接紫外检测法测定糖蜜酒精废液中的有机酸

建立了一种利用胶束电动毛细管色谱-间接紫外检测法同时测定丙二酸、甲酸、酒石酸、苹果酸、琥珀酸、戊二酸、乙酸、乳酸和谷氨酸的新方法。以7.5 mmol/L邻苯二甲酸氢钾-1.5 mmol/L十六烷基三甲基溴化铵(用0.1 mol/L氢氧化钠调pH至6.50)混合液作为电泳介质,检测波长为300 nm,参比波长为210 nm,未涂层弹性石英毛细管(50 μm i.d.×64 cm)为分离通道,在6 min内实现了9种酸的完全分离。9种有机酸的浓度与其峰面积在一定的范围呈良好的线性关系,检出限均低于1.5 mg/L,迁移时间和峰面积的日内、日间相对标准偏差均小于6%。该法用于糖蜜酒精废液中有机酸的测定,结果令人满意,9种有机酸的样品加标回收率均在93%以上。     

Analysis of low molecular mass organic acids in natural waters by ion exclusion chromatography tandem mass spectrometry

A sensitive and selective method for the analysis of aliphatic low molecular mass organic acids (LMMOAs) in natural waters is presented. The method is based on separation with ion exclusion chromatography and detection with electrospray ionization tandem mass spectrometry (LC-MS/MS). The extra selectivity gained by applying MS/MS allows for a minimum of sample preparation and the use of a sub-optimal mobile phase regarding chromatographic resolution. Instead the mobile phase, comprising aqueous formic acid with methanol as organic modifier, was mainly optimized for maximum sensitivity and long term MS stability. Detection limits for malonic, fumaric, maleic, succinic, citraconic, glutaric, malic, alpha-ketoglutaric, tartaric, shikimic, trans-aconitic, cis-aconitic, isocitric and citric acid were in the range 1-50 nM, while the detection limits for pyruvic, oxalic and lactic acid were around 250 nM for an injection volume of 100 microL. Due to their metal-chelating properties, these LMMOAs are all considered to affect the bioavailability of metals and to be involved in soil forming processes. It is thus of interest to be able to monitor their presence in natural waters, and the method developed within this work was successfully applied for the analysis of LMMOAs in soil solution and stream water samples.     

Artifactual Signal Splitting in the Capillary Electrophoresis Analysis of Organic Acids in Wine

Abstract

An artifactual signal splitting was observed for acetic acid during the capillary zone electrophoresis of wine acids. Further study revealed that other comparably weak acids, propionic and butyric, show a similar split. Stronger carboxylic acids, formic, tartaric, malic, citric, and lactic acids showed no artifactual signal splits under the analysis conditions. Succinic acid showed intermediate behavior. The signal splitting could be eliminated by reducing the sample size and/or by employing the background electrolyte as the diluant.     

Effect of stationary phase hydrophobicity and mobile phase composition on the separation of carboxylic acids in ion chromatography

In a previous work, we studied the retention behavior of monovalent and divalent carboxylic acids on a highly cross-linked polystryene-divinylbenzene anion-exchange column (IonPac AS4A-SC) using a carbonate-based buffer, and a retention model was applied to the chromatographic data obtained. In this work we characterized the retention of carboxylates (formic, acetic, propionic, lactic, pyruvic, oxalic, malonic, succinic, fumaric, maleic, tartaric, glutaric, adipic, malic, mucic, trans-beta-hydromuconic, trans,trans-muconic acids) on a column with higher hydrophilicity (IonPac AS11) according to analyte and stationary phase properties, using previously investigated eluent compositions and comparing the retention data obtained. Moreover, the effect of organic modifiers (CH3OH and CH3CN) in the eluent on the retention factors was also evaluated. The chromatographic data obtained on the IonPac AS11 column were fitted by the retention model and allowed one to obtain and to compare ion-specific selectivity constants (parameters of the model) with the ones obtained with the previous column.     

Development and optimization of a method for the analysis of low-molecular-mass organic acids in plants by capillary electrophoresis with indirect UV detection

Capillary zone electrophoresis was developed to analyze low-molecular-mass organic acids including oxalic, tartaric, formic, malic, citric, succinic, glutaric, acetic and lactic acid. The influences of some crucial parameters such as buffer concentration, pH value, surfactant concentration and separation temperature, on electrophoretic separation were investigated. Under the conditions of 15 mM phthalate containing 0.6 mM tetradecyltrimethylammonium bromide as the run buffer (pH 5.6); separation voltage -15 kV (-263.2 V/cm) and temperature 25 degrees C, a satisfactory separation of nine organic acids was accomplished within 7 min. The detection limits (S/N=3) ranged from 0.008 to 0.08 microg/ml and the quantification limits ranged from 0.01 to 0.1 microg/ml for electrokinetic injection. The method was successfully applied to analyze organic acids in different parts of Var splendens (an edible vegetable in China). The recoveries of organic acids in real samples ranged from 88 to 121%.     

离子排斥色谱法同时测定果汁中11种有机酸

 用离子排斥色谱法实现了对果汁中 11种有机酸 (草酸、柠檬酸、酒石酸、苹果酸、抗坏血酸、乳酸、琥珀酸、甲酸、乙酸、戊二酸、富马酸 )的分离测定。以 17mmol/L硫酸为淋洗液 ,样品在ICE ION 30 0离子排斥柱上分离后 ,用紫外检测器在 2 10nm处测定其中的有机酸。各组分质量浓度测定的相对标准偏差在 1.5 %～ 9.8% (n =10 )。     

Characterization of carboxylic acids in atmospheric aerosols using hydrophilic interaction liquid chromatography tandem mass spectrometry

A sensitive hydrophilic interaction liquid chromatography/electrospray ionization mass spectrometry (HILIC/ESI-MS/MS) method was developed for determination of selected aliphatic (i.e. malonic, succinic, glutaric, adipic, pimelic, suberic, azelaic, maleic, fumaric, glycolic and pyruvic acid), alicyclic (i.e. cis-pinonic and pinic acid) and aromatic (i.e. trimesic, phthalic acid and its isomers) carboxylic acids. Analytes were separated on an amide column using a gradient elution with a 10mM constant ionic strength mobile phase containing acetonitrile and aqueous ammonium acetate buffer (pH 5.0). The influence of the buffer type, pH, polar modifier and temperature on analyte retention under HILIC was studied. Static sonication-assisted solvent extraction was optimized for sample preparation prior to analysis. The recoveries obtained were higher than 90% for most analytes. The method was proven to be sensitive with limits of detection ranged from 0.03 to 16.0 μg/L in selected reaction monitoring mode (SRM). The repeatability and intermediate precision of the method, expressed as RSD (%) of the peak area ratio between analytes and their internal standards were generally lower than 5%. The method was successfully applied for determination of the studied acids in samples of ambient aerosol particles. A big advantage of the new method is also its ability to detect and separate the isobaric compounds of the selected carboxylic acids. Our results demonstrate that the method is specific and sensitive for the determination of a wider range of polar carboxylic acids at low concentrations in complex samples of aerosol particles.     

Determination of organic acids in alcoholic and nonalcoholic beverages by reversed-phase high-performance liquid chromatography

A procedure for the quantification of 9 organic acids, acetic, formic, citric, tartaric, lactic, malic, succinic, oxalic, and fumaric, in alcoholic and alcohol-free beverages by reversed-phase HPLC on a Pronto-SIL C18 AQ (300 × 3 mm) column (3 μm) with the mobile phase 5 mM Li₂SO₄ (pH 3.00, H₂SO₄) at the rate 0.5 mL/min and conductometry detection. The analytical ranges made 5–200 mg/L for tartaric, malic, lactic and acetic acids, 2–200 mg/L for the citric and fumaric, 10–400 mg/L for succinic, 15–400 for oxalic, and 20–200 for the formic acids, and so the detection limits: 1 mg/L for tartaric, formic, malic and fumaric, 2 mg/L for lactic, acetic and citric, 5 mg/L for succinic, and 10 mg/L for oxalic acids. The analysis of alcoholic beverages takes 30–40 min, and of non-alcoholic ones, 20–30 min; the standard deviation of the results of analyses does not exceed 5%.     

Simultaneous determination of dl-lactic acid and dl-3-hydroxybutyric acid enantiomers in saliva of diabetes mellitus patients by high-throughput LC–ESI-MS/MS

A simultaneous determination method for the enantiomers of chiral carboxylic acids by the combination of ultraperformance liquid chromatography and mass spectrometry (UPLC-MS/MS) has been developed. (S)(+)-1-(2-Pyrrolidinylmethyl)-pyrrolidine (S-PMP) was used as the derivatization reagent for the high-throughput determination of biological chiral carboxylic acids, i.e., lactic acid (LA) and 3-hydroxybutyric acid (HA). The S-PMP efficiently reacted with the carboxylic acids under mild conditions at room temperature in the presence of 2,2'-dipyridyl disulfide and triphenylphosphine. The resulting S-PMP derivatives were highly responsive in the electrospray ionization (ESI)-MS operating in the positive-ion mode and gave characteristic product ions during the MS/MS, which enabled the sensitive detection using selected reaction monitoring. The derivatization was effective for the enantiomeric separation of the chiral carboxylic acids, and the resolution values of DL-LA and DL-HA were 4.91 and 9.37, respectively. Furthermore, a rapid separation of the derivatives of DL-LA and DL-HA within 7?min was performed using the UPLC system. The limits of detection on the column were in the low femtogram range (5-12?fg). The proposed procedure was successfully applied for the determination of the D- and L-isomers of LA and HA in the saliva of diabetes mellitus (DM) patients and healthy volunteers. The D-LA in DM patients was clearly higher than that in normal subjects. The derivatization followed by UPLC-ESI-MS/MS enabled the enantiomeric separation and detection of trace amounts of LA and HA in human saliva with a simple pretreatment and small sample volume.     

Capillary electrophoresis for the analysis of short-chain organic acids in coffee

A simple and rapid capillary electrophoresis method for low-molecular mass carboxylic acids measurement in coffee has been optimised and validated. Regarding separation conditions, phosphate concentration in the background electrolyte, surfactant type [cetyltrimethylammonium bromide (CTAB), tetradecyltri methylammonium bromide (TTAB) and hexadimethrine bromide (HDB)], percentages of organic modifier and pH were assayed. The best conditions were: 500 mM phosphate buffer at pH 6.25 with CTAB 0.5 mM. The separation was carried out with an uncoated fused-silica capillary (57 cm x 50 microm i.d.) which was operated at -10 kV potential. Detection was performed at 200 nm. In such conditions 17 short-chain organic acids: oxalic, formic, fumaric, mesaconic, succinic, maleic, malic, isocitric, citric, acetic, citraconic, glycolic, propionic, lactic, furanoic, pyroglutamic, quinic acids plus nitrate were separated, identified and measured. Validation parameters of the method allow us to consider it lineal, accurate and precise and, therefore, reliable for its employment in food composition studies or for quality control. Results in coffees with different industrial treatment allow the detection of important differences in the organic acid profile.     

Polymeric sensors for determination of anions of organic acids

New polymeric electrochemical sensors based on various neutral and charged organic ionophores were suggested. The new sensors have a high sensitivity to anions of various organic acids (acetic, oxalic, lactic, malic, succinic, tartaric, and citric) in a wide concentration range. The selectivity and the detection limit of the developed sensors with respect to anions of the acids were studied.     

LC Determination of Trace Short-Chain Organic Acids in Wheat Root Exudates Under Aluminum Stress

A novel and reliable high-performance liquid chromatographic method for trace short-chain organic acids measurement in wheat root exudates under aluminum stress has been optimized and validated. The chromatographic separation of the short-chain organic acids (citric, oxalic, malonic, succinic, tartaric, malic, and acetic acids) was achieved with Bio-rad Aminex HPX-87H cation exchange resin column. These seven organic acids were identified and quantified in 25 min. Well-shaped peaks were obtained for trace organic acids using dilute sulfuric acid as mobile phase. Under optimum conditions, Bio-rad Aminex HPX-87H column showed distinct advantages of the ability to well separate different short-chain organic acids (especially for tartaric and malic acids, as well as malonic and acetic acids) in wheat root exudates under aluminum stress, and offered accurate and precise results for the analysis of these organic acids. This HPLC method can efficiently eliminate the aluminum’s interference and is quite suitable to the trace detection of trace organic acids in wheat root exudates under aluminum stress.

     

LC Determination of Trace Short-Chain Organic Acids in Wheat Root Exudates Under Aluminum Stress

A novel and reliable high-performance liquid chromatographic method for trace short-chain organic acids measurement in wheat root exudates under aluminum stress has been optimized and validated. The chromatographic separation of the short-chain organic acids (citric, oxalic, malonic, succinic, tartaric, malic, and acetic acids) was achieved with Bio-rad Aminex HPX-87H cation exchange resin column. These seven organic acids were identified and quantified in 25 min. Well-shaped peaks were obtained for trace organic acids using dilute sulfuric acid as mobile phase. Under optimum conditions, Bio-rad Aminex HPX-87H column showed distinct advantages of the ability to well separate different short-chain organic acids (especially for tartaric and malic acids, as well as malonic and acetic acids) in wheat root exudates under aluminum stress, and offered accurate and precise results for the analysis of these organic acids. This HPLC method can efficiently eliminate the aluminum’s interference and is quite suitable to the trace detection of trace organic acids in wheat root exudates under aluminum stress.     

Application of co-electroosmotic capillary electrophoresis for the determination of inorganic anions and carboxylic acids in soil and plant extract with direct UV detection

Summary Indirect UV detection in capillary zone electrophoresis (CZE) is frequently used for the determination of inorganic anions and carboxylic acids. However, there are few reports on direct UV detection of these solutes in real samples. This paper describes the use of direct UV detection of inorganic anions and organic acids in environmental samples using co-electroosmotic capillary zone electrophoresis (co-CZE) at 185 nm. The best separation and detection of the solutes was achieved using a fused silica capillary with an electrolyte containing 25 mM phosphate, 0.5 mM tetradecyltrimethylammonium bromide (TTAB) and 15% acetonitrile (v/v) at pH 6.0. Four common inorganic anions (Cl⁻, NO₂ ⁻, NO₃ ⁻ and SO₄ ²⁻) and 11 organic acids (oxalic, formic, fumaric, tartaric, malonic, malic, citric, succinic, maleic, acetic, and lactic acid), were determined simultaneously in 15 min. Linear calibration plots for the test solutes were obtained in the range 0.02–0.5 mM with detection limits ranging from 1–9 μM depending on the analyte. The proposed method was successfully used to determine inorganic anions and carboxylic acids in soil and plant tissue extracts with direct injection of the sample.     

Thermodynamic parameters for the protonation of carboxylic acids in aqueous tetraethylammonium iodide solutions

The protonation constants of 21 carboxylic acids (formic, acetic, propionic, benzoic, phenoxyacetic, salicylic, oxalic, malonic, succinic, itaconic, malic, tartaric, oxydiacetic, thiodiacetic, thiodipropionic, phthalic, maleic, citric, 1,2,3-tricarboxylic, 1,2,4-tricarboxylic and 1,2,4,5-tetracarboxylic), have been determined potentiometrically, by pH-metric measurements, at several temperatures and ionic strengths, 5T55°C, 0<11 mol-dm^–3, using tetraethylammonium iodide as background salt. General equations for the dependence on ionic strength of thermodynamic parameters have been found. The statistical significance of results and the possibility of using a simple model for the thermodynamics of carboxylic acids protonation, is discussed.     

In vitro and in vivo evaluation of a <Superscript>64</Superscript>Cu-labeled propylene amine oxime complex as a potential hypoxia imaging agent bearing two 3-nitrotriazole groups

The effect of coating with nine different carboxylic acids (glycolic, propionic, lactic, malic, tartaric, citric, mandelic, caproic and caprylic) on nanostructured magnetite (D ~ 10 nm) was studied by Raman and photoacoustic, magnetic and ⁵⁷Fe Mössbauer measurements. Mössbauer spectra of frozen suspensions showed dominantly magnetically split envelopes at lower temperatures, which were evaluated by hyperfine field distribution method. Mössbauer and Raman spectroscopy indicated similar variation of relative occurrence of magnetite and maghemite phases. These results are discussed on the basis of the hypothesis that different carboxylic acids can promote either the oxidation or reduction of iron oxide nanoparticles.