Separation and purification of two minor typical diarrhetic shellfish poisoning toxins from harmful marine microalgae via combined liquid chromatography with mass spectrometric detection

A novel method was developed for the purification of two typical diarrhetic shellfish poisoning toxins from toxin‐producing marine microalgae using macroporous resin, high‐speed countercurrent chromatography–mass spectrometry, and semipreparative high‐performance liquid chromatography–mass spectrometry. Analytical high‐performance liquid chromatography–mass spectrometry was used for identification and purity analysis of okadaic acid and dinophysistoxin‐1 because they exhibit no visible or ultraviolet absorption. First, four kinds of macroporous resins were investigated, and HP‐20 macroporous resin was selected for the preenrichment and cleanup of the two target toxins. Second, the resin‐purified sample was further purified using high‐speed countercurrent chromatography coupled with a mass spectrometer. The purities of the obtained okadaic acid and dinophysistoxin‐1 were 89.0 and 83.0%, respectively, as determined through analytical high‐performance liquid chromatography–mass spectrometry. Finally, further purification was carried out using semipreparative high‐performance liquid chromatography with mass spectrometry, and the purities of the final okadaic acid and dinophysistoxin‐1 products were both over 98.0% based on the analytical high‐performance liquid chromatography–mass spectrometry chromatograms and fraction spectra. This work demonstrates that the proposed purification process is a powerful method for the preparation of high‐purity okadaic acid and dinophysistoxin‐1 from toxin‐producing marine microalgae. Moreover, it is particularly important for the purification and preparation of minor toxins that exhibit no visible or ultraviolet absorption from harmful marine algae.     

Identification and quantitation of major phenolic compounds from penthorum chinense pursh. by HPLC with tandem mass spectrometry and HPLC with diode array detection

Penthorum chinense Pursh. is a traditional Chinese herbal medicine used for the treatment of various ailments specially related to liver. Gansu Granule, the medicine made from the extract of P. chinense, has been widely used in the clinical setting. But the information about its active ingredients is lacking. In this paper, the extract of P. chinense was analyzed by high performance liquid chromatography with electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry. Among the 27 compounds that were identified based on their mass spectrometry data, ten were reported for the first time from P. chinense. Chromatographic fingerprints generated by high‐performance liquid chromatography by analyzing 21 batches of P. chinense, displayed six common peaks. Finally, four major compounds were identified namely; gallic acid, brevifolin carboxylic acid, 2,6‐dihydroxyacetophenone‐4‐O‐β‐d‐ glucoside, and pinocembrin‐7‐O‐β‐d‐ glucoside. The average content of each compound was 24.58, 109.6, 15.52, and 18.81 mg/g, respectively. In addition, this study also suggests that the qualitative liquid chromatography with mass spectrometry and the quantitative high‐performance liquid chromatography analytical methods using monolithic columns are simple, rapid, accurate, and reproducible and have the potential to be used for the comprehensive quality control of P. chinense.     

Qualitative and quantitative analysis of major constituents from Dazhu Hongjingtian capsule by UPLC/Q‐TOF‐MS/MS combined with UPLC/QQQ‐MS/MS

In this work, a sensitive and efficient method was established and validated for qualitative and quantitative analysis of major bioactive constituents in Dazhu Hongjingtian capsule by liquid chromatography tandem mass spectrometry. A total of 32 compounds were tentatively identified using ultra‐performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry. Furthermore, 12 constituents, namely gallic acid, 3,4‐dihydroxybenzoic acid, salidroside, p‐ coumaric acid‐4‐O ‐β ‐d ‐glucopyranoside, bergeninum, 4‐hydroxybenzoic acid, 4‐hydroxyphenylacetic acid, syringate, 6′′‐O ‐galloylsalidroside, rhodiosin, rhodionin and kaempferol‐7‐O ‐α ‐l ‐rhamnoside, were simultaneously quantified by the developed ultra‐performance liquid chromatography coupled with a triple quadrupole mass spectrometry method in 9 min. All of them were analyzed on an Agilent ZorBax SB‐C₁₈ column (3.0 × 100 mm, 1.8 μm) with linear gradient elution of methanol–0.1% formic acid water. The proposed method was applied to analyze three batches of samples with acceptable linearity (R , 0.9979–0.9997), precision (RSD, 1.3–4.7%), repeatability (RSD, 1.7–4.9%), stability (RSD, 2.2–4.9%) and recovery (RSD, 0.6–4.4%) of the 12 compounds. As a result, the analytical method possessing high throughput and sensitivity is suitable for the quality control of Dazhu Hongjingtian capsule.     

Differentiation of Mint (Mentha haplocalyx Briq.) from different regions in China using gas and liquid chromatography

In this study, complex substances such as Mint (Mentha haplocalyx Briq.) samples from different growing regions in China were analyzed for phenolic compounds by high‐performance liquid chromatography with diode array detection and for the volatile aroma compounds by gas chromatography with mass spectrometry. Chemometrics methods, e.g. principal component analysis, back‐propagation artificial neural networks, and partial least squares discriminant analysis, were applied to resolve complex chromatographic profiles of Mint samples. A total of 49 aroma components and 23 phenolic compounds were identified in 79 Mint samples. Principal component analysis score plots from gas chromatography with mass spectrometry and high‐performance liquid chromatography with diode array detection data sets showed a clear distinction among Mint from three different regions in China. Classification results showed that satisfactory performance of prediction ability for back‐propagation artificial neural networks and partial least squares discriminant analysis. The major compounds that contributed to the discrimination were chlorogenic acid, unknown 3, kaempherol 7‐O‐rutinoside, salvianolic acid L, hesperidin, diosmetin, unknown 6 and pebrellin in Mint according to regression coefficients of the partial least squares discriminant analysis model. This study indicated that the proposed strategy could provide a simple and rapid technique to distinguish clearly complex profiles from samples such as Mint.     

Liquid chromatography/mass spectrometry for investigating the biochemical effects induced by aristolochic acid in rats: the plasma metabolome

In this study, high‐performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry in conjunction with chemometric methods including principal components analysis was used to investigate metabolic profiling in plasma samples from rats dosed with aristolochic acid (AA). Differentiating metabolites were identified by high‐resolution mass spectrometry and tandem mass spectrometry analyses, database searching and comparison with the analytical results of authentic standards. Several metabolites in plasma including a glucuronide conjugate, bile acids, lysophosphatidylcholines and fatty acids were detected in rats exposed to AA. To improve visualization, a z‐score plot and a cluster heat map were generated for the concentration fluctuations of the metabolites in different dosage groups. The results obtained from this study indicated that class‐specific metabolomic patterns were obviously differentiated for each metabolite in the different dosage groups of AA. Copyright © 2010 John Wiley & Sons, Ltd.     

Simultaneous determination of polysaccharides and 21 nucleosides and amino acids in different tissues of Salvia miltiorrhiza from different areas by UV–visible spectrophotometry and UHPLC with triple quadrupole MS/MS

Salvia miltiorrhiza, a traditional Chinese medicine, is a widely used herbal medicine to treat cardiovascular and cerebrovascular diseases. In this study, ultraviolet (UV)–visible spectrophotometry and ultra‐high performance liquid chromatography with triple quadrupole tandem mass spectrometry analytical methods were used for rapid quantification of polysaccharides and 21 nucleosides and amino acids in S. miltiorrhiza to determine 17 samples of different tissues from different areas. Based on the total contents, hierarchical clustering analysis and principal components analysis were performed to classify these samples. The established methods were validated with good linearity, precision, repeatability, stability, and recovery. Chemical analysis revealed a higher content of total analytes in the sample of inflorescence from Nanjing (34.17 mg/g), sample of root and rhizome from Shaanxi (34.13 mg/g) and sample of stem and leaf from Nanjing (31.14 mg/g), respectively, indicating that root and rhizome from Shaanxi and the aerial parts from Nanjing exhibited the highest quality due to their highest content. In addition, contents of nucleosides and amino acids in the aerial parts (14.67 mg/g) were much higher than that in roots and rhizomes (9.17 mg/g). This study suggested that UV–visible spectrophotometry and ultra‐high performance liquid chromatography with triple quadrupole tandem mass spectrometry are effective techniques to analyze polysaccharides, nucleosides, and amino acids in plants, and they provided valuable information for the development and utilization value of the aerial parts of S. miltiorrhiza. This analysis would also provide useful information for the quality control of S. miltiorrhiza.     

Combination of ultrasound‐assisted ethyl chloroformate derivatization and switchable solvent liquid‐phase microextraction for the sensitive determination of l‐methionine in human plasma by GC–MS

A green and fast analytical method for the determination of l ‐methionine in human plasma is presented in this study. Preconcentration of the analyte was carried out by switchable solvent liquid phase microextraction after ethyl chloroformate derivatization reaction. Instrumental detection of the analyte was performed by means of gas chromatography–mass spectrometry. N,N‐Dimethyl benzylamine was used in the synthesis of switchable solvent. Protonated N,N‐dimethyl benzylamine volume, volume/concentration of sodium hydroxide, and vortex period were meticulously fixed to their optimum values. Besides, ethyl chloroformate, pyridine, and ethanol volumes were optimized in order to get high derivatization yield. After the optimization studies, limit of detection and quantitation values were attained as 3.30 and 11.0 ng/g, respectively, by the developed switchable solvent liquid phase microextraction gas chromatography–mass spectrometry method that corresponding to 76.7‐folds enhancement in detection power of the gas chromatography–mass spectrometry system. Applicability and accuracy of the switchable solvent liquid phase microextraction–gas chromatography–mass spectrometry method were also checked by spiking experiments. Percent recovery results were ranged from 97.8 to 100.5% showing that human plasma samples could be analyzed for its l ‐methionine level by the proposed method.     

Analytical methodologies for the determination of ticagrelor

Ticagrelor is an orally administered platelet aggregation inhibitor with a cyclopentyl‐triazolopyrimidine structure; it is a selective reversible P2Y12 receptor antagonist, which prevents P2Y12‐mediated and ADP‐mediated platelet activation and aggregation. It is used to reduce the rate of cardiovascular death, myocardial infarction and stroke in patients with acute coronary syndrome or history of myocardial infarction. Several analytical methods have been published for the determination of ticagrelor in pharmaceuticals and biological materials by spectrophotometry, high‐performance liquid chromatography with ultraviolet detection and liquid chromatography coupled with tandem mass spectrometry. The purpose of the current review is to provide a systematic survey of the analytical techniques used for the determination of ticagrelor since its introduction in therapy until today.     

Cost effective,robust, and reliable coupled separation techniques for the identification and quantification of phospholipids in complex biological matrices: Application to insects

The quantification of phospholipid classes and the determination of their molecular structures are crucial in physiological and medical studies. This paper's target analytes are cell membrane phospholipids, which play an important role in the seasonal acclimation processes of poikilothermic organisms. We introduce a set of simple and cost‐effective analytical methods that enable efficient characterization and quantification of particular phospholipid classes and the identification and relative distribution of the individual phospholipid species. The analytical approach involves solid‐phase extraction and high‐performance thin‐layer chromatography, which facilitate the separation of particular lipid classes. The obtained fractions are further transesterified to fatty acid methyl esters and subjected to gas chromatography coupled to flame ionization detection, which enables the determination of the position of double bonds. Phospholipid species separation is achieved by high‐performance liquid chromatography with mass spectrometry, which gives information about the headgroup moiety and attached fatty acids. The total content of each phospholipids class is assessed by phosphorus determination by UV spectrophotometry. The simultaneous analysis of phosphorus, fatty acid residues, and phospholipid species provides detailed information about phospholipid composition. Evaluation of these coupled methods was achieved by application to an insect model, Pyrrhocoris apterus. High correlation was observed between fatty acid compositions as determined by gas chromatography and high‐performance liquid chromatography analysis.     

High‐speed counter‐current chromatography assisted preparative isolation of bioactive compounds from stem bark of Juglans mandshurica

High‐speed counter‐current chromatography was applied for the first time for the separation and purification of bioactive compounds contained in the stem bark of Juglans mandshurica Maxim. Silica gel column chromatography was first used to obtain three composition‐enriched target fractions from a crude J. mandshurica methanol extract. Three independent high‐speed counter‐current chromatography processes were then used to further isolate 13 bioactive compounds, namely, six galloyl glucose derivatives, three flavonones, three naphthoquinones, and ethyl gallate. The isolates were identified by ultrahigh‐performance liquid chromatography with tandem mass spectrometry, and ultraviolet and NMR spectroscopy, and compared with literature data. Their purities were determined to be >94.6% by ultrahigh performance liquid chromatography. Furthermore, based on the total phenolic content and results of a 2,2‐diphenyl‐1‐picrylhydrazyl test, the methanol extract and two of the three initial fractions were observed to be rich in phenolic compounds and exhibit good free radical scavenging abilities, while nine of the isolated compounds exhibited remarkable antioxidant activity, superior to that of butyrate hydroxy‐toluene and comparable to that of gallic acid. The results of this research confirm the effectiveness of high‐speed counter‐current chromatography for the separation of compounds contained in extremely complex samples, and provide a basis for the further utilization of J. mandshurica .     

In vitro screening and isolation of human aromatase inhibitors from Cicer arietinum by a novel continuous online method combining chromatographic techniques

Ultrafiltration liquid chromatography with mass spectrometry can efficiently and rapidly screen and identify ligands from the seeds of Cicer arietinum for human aromatase. Using this method, we identified 11 major compounds, including organic acids, organic acid glycosides, flavone glycosides, isoflavones, and isoflavone glycosides, as potent human aromatase inhibitors. A continuous online method, including pressurized liquid extraction, countercurrent chromatography, and preparative liquid chromatography, was developed for scaling up the production of these compounds with high purity and efficiency. The bioactivity of the separated compounds was assessed by an in vitro enzyme inhibition assay. This novel approach using a combination of ultrafiltration liquid chromatography with mass spectrometry and pressurized liquid extraction with countercurrent chromatography and preparative liquid chromatography as well as an in vitro enzyme inhibition assay could be applied to efficiently screen and isolate human aromatase inhibitors from complex samples and to the large‐scale production of functional food and nutraceutical ingredients.     

Isolation,purification, and identification of the main phenolic compounds from leaves of celery (Apium graveolens L. var. dulce Mill./Pers.)

We developed a simple and meaningful preparative method for the separation and purification of the main phenolic compounds from the leaves of celery (Apium graveolens L. var. dulce Mill./Pers.) and we established an accurate and specific analytical method for the identification of the main phenolic compounds from celery leaves. The crude extract from celery leaves was prefractioned by polyamide resin to enrich the phenolic compounds. They were then purified further by preparative high‐performance liquid chromatography, and seven main phenolic compounds were obtained: including chlorogenic acid, luteolin 7‐O‐β‐d‐ apiofuranosyl(1→2)‐β‐d‐ glucopyranoside, luteolin 7‐O‐β‐d‐ glucopyranoside, apiin, chrysoeriol 7‐O‐β‐d‐ apiofuranosyl(1→2)‐β‐d‐ glucopyranoside, luteolin 7‐O‐[β‐d‐ apiofuranosyl(1→2)‐(6′′‐O‐malonyl)]‐β‐d‐ glucopyranoside, and apigenin 7‐O‐[β‐d‐ apiofuranosyl(1→2)‐(6′′‐O‐malonyl)]‐β‐d‐ glucopyranoside. Their purities were measured by using high‐performance liquid chromatography, and their chemical structures were confirmed using UV spectrophotometry, ultra high performance liquid chromatography with quadrupole time‐of‐flight tandem mass spectrometry, and NMR spectroscopy. Our studies indicate that preparative high‐performance liquid chromatography combined with polyamide resin is a simple and meaningful preparative method for the separation and purification of phenolic compounds from the leaves of celery or other plants, and the use of UV spectrophotometry, ultra high performance liquid chromatography with quadrupole time‐of‐flight tandem mass spectrometry, and NMR spectroscopy is an accurate and specific analytical method for the identification of phenolic compounds.     

Integrating qualitative and quantitative characterization of traditional Chinese medicine injection by high‐performance liquid chromatography with diode array detection and tandem mass spectrometry

The present study aims to describe and exemplify an integrated strategy of the combination of qualitative and quantitative characterization of a multicomponent mixture for the quality control of traditional Chinese medicine injections with the example of Danhong injection (DHI). The standardized chemical profile of DHI has been established based on liquid chromatography with diode array detection. High‐performance liquid chromatography coupled with time‐of‐flight mass spectrometry and high‐performance liquid chromatography with electrospray multistage tandem ion‐trap mass spectrometry have been developed to identify the major constituents in DHI. The structures of 26 compounds including nucleotides, phenolic acids, and flavonoid glycosides were identified or tentatively characterized. Meanwhile, the simultaneous determination of seven marker constituents, including uridine, adenosine, danshensu, protocatechuic aldehyde, p‐coumaric acid, rosmarinic acid, and salvianolic acid B, in DHI was performed by multiwavelength detection based on high‐performance liquid chromatography with diode array detection. The integrated qualitative and quantitative characterization strategy provided an effective and reliable pattern for the comprehensive and systematic characterization of the complex traditional Chinese medicine system.     

Comprehensive identification of potential antioxidant components in the aerial parts of Polygonum chinense L. var. hispidum using ultra high performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry

The aerial parts of Polygonum chinense L. var. hispidum are one of the key herbs in Cantonese herbal tea, which is quite a common local beverage in LingNan area of China. Previous investigation has found that this herb possesses antioxidant activity and the ethyl acetate fraction of its ethanol extract shows the strongest antioxidant activity. However, little is known about its antioxidant chemical constituents. The aim of this research was to investigate the active constituents of this plant by identifying and characterizing the chemical profile in ethyl acetate fraction using ultra high performance liquid chromatography with quadrupole time‐of‐flight mass spectrometry, which can provide characteristic ultraviolet absorption, accurate molecular weight, and diagnostic tandem mass spectrometry fragment ions. As a result, 85 compounds were identified including 22 flavonoids, 12 ellagic acids, 34 ellagitannins, 16 phenolic acids, and one phenolic amide. All the phenolic compounds identified in this work, especially ethyl gallate, geraniin, chebulagic acid, and quercitrin with the higher peak areas in the ultra high performance liquid chromatography with mass spectrometry chemical profile of this plant, could be the bioactive principles responsible for the antioxidant activity. These findings in the present study could benefit further studies involving the functions and chemicals of this plant, and provide scientific evidence for usage of Cantonese herbal tea.     

Analysis of diphenylarsinic acid in human and environmental samples by HPLC–ICP‐MS

A simple, rapid and robust analytical method for determining diphenylarsinic acid in human and environmental samples was developed based on a combination of hydrophilic polymer‐based gel‐permeation high‐performance liquid chromatography (HPLC) and inductively coupled plasma mass spectrometry (ICP‐MS). Hair and nail samples were digested with alkali, and liberated diphenylarsinic acid (derivative) was extracted with diethyl ether, redissolved in water and injected for HPLC–ICP‐MS analysis. Human urine, groundwater and water extracts from soils were injected for HPLC–ICP‐MS directly after filtration. Using the method, diphenylarsinic acid in a solution was quantified in 7 min duration for an analysis with a detection limit of sub‐nanograms per milliliter. The method has been applied to groundwater arsenic pollution recently uncovered in Japan. Copyright © 2005 John Wiley & Sons, Ltd.     

Enantioselective analysis of bambuterol in human plasma using microwave‐assisted chiral derivatization coupled with ultra high performance liquid chromatography and tandem mass spectrometry

In this study, an enantioselective analytical method based on microwave‐assisted chiral derivatization coupled with ultra high performance liquid chromatography and tandem mass spectrometry was developed for the determination of bambuterol enantiomers in human plasma. The chiral derivatization reaction was greatly accelerated by microwave irradiation. Under the optimized conditions, both the derivatization time and separation time on column was only 3 min, and the lower limit of quantification was 2.5 pg/mL. The recoveries were in the range of 90.1–93.0% without significant matrix effect. Compared with the conventional heating chiral derivatization, microwave‐assisted chiral derivatization obtained higher chiral derivatization yields with much shorter time due to the effect of microwave irradiation. Furthermore, the racemization during the derivatization reaction was systematically investigated. The results showed the concentration of acetic acid and the reaction time had significant effects on the racemization, which could be well controlled during microwave‐assisted chiral derivatization for the short reaction time. Finally, this novel approach was demonstrated by determining bambuterol in human plasma of a clinical pharmacokinetic study in eight healthy volunteers. On the basis of the results, microwave‐assisted chiral derivatization coupled with ultra high performance liquid chromatography and tandem mass spectrometry as a simple and effective enantioselective analysis technique for the determination of chiral drugs in complex biological samples showed great promise.     

Simultaneous determination of selected estrogenic endocrine disrupting chemicals and bisphenol A residues in whole milk using fabric phase sorptive extraction coupled to HPLC‐UV detection and LC‐MS/MS

A simple and sensitive analytical methodology is developed for rapid screening and quantification of selected estrogenic endocrine disrupting chemicals and bisphenol A from intact milk using fabric phase sorptive extraction in combination with high‐performance liquid chromatography coupled to ultraviolet detection/tandem mass spectrometry. The new approach eliminates protein precipitation and defatting step from the sample preparation workflow. In addition, the error prone and time‐consuming solvent evaporation and sample reconstitution step used as the sample post‐treatment has been eliminated. Parameters with most significant impact on the extraction efficiency of fabric phase sorptive extraction including sorbent chemistry, sample volume, extraction time have been thoroughly studied and optimized. Separation of the selected estrogenic endocrine disrupting chemicals including α‐estradiol, hexestrol, estrone, 17α‐ethinyl estradiol, diethylstilboestrol, and bisphenol A were achieved using a Zorbax Extend‐C18 high‐performance liquid chromatography column (15 cm × 4.6 mm, 5 μm particle size). The limit of detection values obtained in fabric phase sorptive extraction with high‐performance liquid chromatography with ultraviolet detection ranged from 25.0 to 50.0 ng/mL. The method repeatability values were 3.6–13.9 (relative standard deviation, %) and intermediate precision values were 4.6–12.7 (relative standard deviation, %). The fabric phase sorptive extraction method was also coupled to liquid chromatography with tandem mass spectrometry for identifying each endocrine disrupting chemical at 10 ng/mL.     

Qualitative and quantitative determination of YiXinShu Tablet using ultra high performance liquid chromatography with Q Exactive hybrid quadrupole orbitrap high‐resolution accurate mass spectrometry

To clarify and quantify the chemical profile of YiXinShu Tablet rapidly, a feasible and accurate strategy was developed by applying ultra high performance liquid chromatography with Q Exactive hybrid quadrupole orbitrap high‐resolution accurate mass spectrometry. A total of 105 components were identified, including 25 phenanthraquinones, 11 lactones, 19 lignans, 24 acids, and 26 other compounds. Among them, 26 major compounds were unambiguously detected by comparing with reference standards. And 19 of these compounds in three batches of YiXinShu Tablet were selected for quantitative determination. (Z )‐Ligustilide, salvianic acid A, salvianolic acid A, salvianolic acid B, and rosmarinic acid were abundant in these three batches with contents over 1 mg/g. The established analysis methods were examined to be accurate and feasible. The results show that the ultra high performance liquid chromatography with Q Exactive hybrid quadrupole orbitrap high‐resolution accurate mass spectrometry method has a powerful qualitative ability and promising quantitative application.     

Combination of different liquid chromatography/mass spectrometry technologies for the identification of transformation products of rhodamine B in groundwater

Rhodamine B and its five de‐ethylated transformation products could be identified in a groundwater sample. Using high‐performance thin‐layer chromatography (HPTLC) six fluorescent zones were detected in the sample. In order to identify the compounds in the zones by exact mass mass spectrometry (MS) measurements and tandem mass spectrometry (MS/MS), they were extracted from the HPTLC plate for subsequent analysis by nano‐chip high‐performance liquid chromatography quadrupole‐time‐of‐flight mass spectrometry (nano‐chip HPLC/QTOFMS). In addition, chemical derivatisation experiments on HPTLC plates were applied to detect the presence of a primary amino group in the transformation products. From the combined analytical results it was possible to allocate rhodamine B and its five de‐ethylated transformation products to the six different HPTLC zones. The quantification of rhodamine B in different groundwater samples was carried out by a high‐performance liquid chromatography/triple quadrupole mass spectrometry (HPLC/MS/MS). The maximum detected concentration of rhodamine B was 83 µg L^?1. Copyright © 2010 John Wiley & Sons, Ltd.     

Simultaneous separation and determination of thallium in water samples by high‐performance liquid chromatography with inductively coupled plasma mass spectrometry

An analytical method for separation and determination of thallium species in water using high‐performance liquid chromatography with inductively coupled plasma mass spectrometry was developed. The composition and concentration of mobile phase, injection volume, and pH value were optimized respectively with an anion or cation exchange column. The results showed that Tl(I) and Tl(III) were effectively separated using anion exchange column Hamilton PRP‐X100, with the mobile phase consisting of 200 mmol/L ammonium acetate and 10 mmol/L diethylenetriaminepentaacetic acid (pH = 4.2). When using a Dionex cation exchange guard column, CS12A, 15 mmol/L HNO₃, and 3 mmol/L diethylenetriaminepentaacetic acid as the mobile phase, Tl(I) and Tl(III) could be effectively separated. The detection limits of the methods were 3–6 and 9–12 ng/L, respectively. In a solution containing Fe ions and oxalic acid, a significant quantity of Tl(I) was oxidized. Fe ions and oxalic acid in the water samples did not interfere with high‐performance liquid chromatography‐inductively coupled plasma mass spectrometry measurement results.