Application of central composite designs for optimisation of the chromatographic separation of monomethylarsonate and dimethylarsinate and of selenomethionine and selenite by ion-pair chromatography coupled with plasma mass spectrometric detection

Central composite designs (CCDs) were used in the study of the ion-pair chromatographic separation of arsenic and selenium species in tap water: monomethylarsonate, dimethylarsinate, selenomethionine and selenite. The ternary eluent consisted of tetrabutylammonium phosphate (TBA), sodium hydrogenphosphate (Na2HPO4) and 1% acetonitrile. CCD allowed the investigation of the influence of the eluent parameters, which varied from 0.5 to 4.2 mmol l-1 Na2HPO4, 0.5 to 4.2 mmol l-1 TBA and pH 4.9 to 8.2, on the capacity factors (k') of arsenic and selenium compounds. Furthermore, another mathematical model that permitted the variation of the chromatographic selectivity of species, computed from their retention data to be followed, was investigated. This showed the ability to locate the optimum conditions within the experimental design, so that arsenic and selenium species could be simultaneously quantified with good efficiency and resolution. A comparison between the predicted and the experimental response values was made in order to assess the prediction quality of the model. Response surfaces and isoresponse curves obtained from the mathematical models allowed the determination of the optimum chromatographic conditions and the robustness of the method. The predicted optimum zone allowing satisfactory determination of both arsenic and selenium compounds was pH 5.5-6.5, 2.5 mmol l-1 Na2HPO4 and 3.0-4.0 mmol l-1 TBA.     

Determination of 'arsenosugars' in algae with anion-exchange chromatography and an inductively coupled plasma mass spectrometer as element-specific detector

The retention behavior of four naturally occurring dimethylarsinoylribosides with -CH2-CHOH-CH2X (X = OH, HO3POCH2CHOHCH2OH, SO3H, OSO3H) as aglycones, of arsenous acid, arsenic acid, methylarsonic acid, and dimethylarsinic acid was investigated on a Hamilton PRP-X100 anion-exchange column with aqueous solutions of ammonium dihydrogen phosphate (20 mmol/L) in the pH range of 3.8-9.0 as mobile phase. A HP 4500 inductively coupled plasma mass spectrometer (ICP-MS) served as arsenic-specific detector. The influence of pH, temperature, and the concentration of methanol in the mobile phase on the retention times of these arsenic compounds was explored. An aqueous 20 mM ammonium dihydrogen phosphate solution at pH 5.6 at a column temperature of 40 degrees C was considered optimal as it allowed the separation of seven of the arsenic compounds within 16 min. Only arsenous acid and the ribose with the glycerol aglycone have overlapping signals with both migrating almost with the solvent front. At a concentration of 0.50 ng As mL(-1) the relative standard deviations (n = 3) of the signal areas of the eight arsenic compounds was in the range from 3.5 to 8.1%. The linear calibration curves (peak areas) from 0.5 to 10 ng/mL had correlation coefficients > 0.997. Extracts obtained from the brown algae Fucus spiralis and Halidrys siliquosa were chromatographed under the optimized conditions. Both species contained the sulfate riboside as the major arsenic compound (approximately 55% of total extractable arsenic) together with the sulfonate- and phosphate riboside. Arsenic acid was a significant constituent of Halidrys siliquosa (approximately 6.5%), but was not detected in Fucus spiralis.     

Online preconcentration of arsenic compounds by dynamic pH junction-capillary electrophoresis

An online preconcentration technique by dynamic pH junction was studied to improve the detection limit for anionic arsenic compounds by CE. The main target compound is roxarsone, or 3-nitro-4-hydroxyphenylarsonic acid, which is being used as an animal feed additive. The other inorganic and organoarsenic compounds studied are the possible biotransformation products of roxarsone. The arsenic species were separated by a dynamic pH junction in a fused-silica capillary using 15 mM phosphate buffer (pH 10.6) as the BGE and 15 mM acetic acid as the sample matrix. CE with UV detection was monitored at a wavelength of 192 nm. The influence of buffer pH and concentration on dynamic pH junction were investigated. The arsenic species focusing resulted in LOD improvement by a factor of 100-800. The combined use of C18 and anion exchange SPE and dynamic pH junction to CE analysis of chicken litter and soils helps to increase the detection sensitivity. Recoveries of spiked samples ranged between 70 and 72%.     

Retention study of inorganic and organic selenium compounds on a silica-based reversed phase column with mixed ion-pairing reagents

Summary Separation of seven inorganic and organic selenium compounds, namely selenic acid [Se(VI)], selenous acid [Se(IV)], trimethylselenonium iodide (TMSe⁺), selenocystine (SeCys), selenomethionine (SeMet), selenoethionine (Seet), and selenocystamine (SeCM), has been performed on a LiChrosorb C 18 column by using mixed ion-pair reagents; 1-butanesulfonic acid and tetramethylammonium hydroxide. Flame atomic absorption spectrometry (FAAS) was used as an element-specific detector. The retention behaviors of selenium compounds in terms of several chromatographic parameters, such as pH of the mobile phase, the concentrations of ion-pair reagents, and the content of organic modifier (methanol) were investigated. It was found that the separation of both inorganic and organic selenium compounds can be achieved within 12 min with a mobile phase of 10 mM 1-butanesulfonic acid −4 mM tetramethylammonium hydroxide −4 mM malonic acid −0.05% methanol adjusted to pH 4.5 at a flow rate of 1.0 mL min⁻¹. The results obtained in this study showed that the use of mixed ion-pair reagents is very useful to improve the separation of selenium compounds. The applicability of this technique for the speciation of selenium compounds in real samples was demonstrated by the determination of selenium compounds in a selenium nutritional supplement. The results were found to be in good agreement with those obtained by ion-exchange HPLC-ICP-MS.     

Determination of ‘arsenosugars’ in algae with anion-exchange chromatography and an inductively coupled plasma mass spectrometer as element-specific detector

The retention behavior of four naturally occurring dimethylarsinoylribosides with –CH₂-CHOH-CH₂X (X = OH, HO₃POCH₂CHOHCH₂OH, SO₃H, OSO₃H) as aglycones, of arsenous acid, arsenic acid, methylarsonic acid, and dimethylarsinic acid was investigated on a Hamilton PRP-X100 anion-exchange column with aqueous solutions of ammonium dihydrogen phosphate (20 mmol/L) in the pH range of 3.8–9.0 as mobile phase. A HP 4500 inductively coupled plasma mass spectrometer (ICP-MS) served as arsenic-specific detector. The influence of pH, temperature, and the concentration of methanol in the mobile phase on the retention times of these arsenic compounds was explored. An aqueous 20 mM ammonium dihydrogen phosphate solution at pH 5.6 at a column temperature of 40?°C was considered optimal as it allowed ¶the separation of seven of the arsenic compounds within 16 min. Only arsenous acid and the ribose with the glycerol aglycone have overlapping signals with both migrating almost with the solvent front. At a concentration of 0.50 ng As mL^–1 the relative standard deviations (n = 3) of the signal areas of the eight arsenic compounds was in the range from 3.5 to 8.1%. The linear calibration curves (peak areas) from 0.5 to 10 ng/mL had correlation coefficients ¶> 0.997. Extracts obtained from the brown algae Fucus spiralis and Halidrys siliquosa were chromatographed under the optimized conditions. Both species contained the sulfate riboside as the major arsenic compound (～55% of total extractable arsenic) together with the sulfonate- and phosphate riboside. Arsenic acid was a significant constituent of Halidrys siliquosa (～6.5%), but was not detected in Fucus spiralis.     

Determination of selenium compounds in urine by high-performance liquid chromatography--inductively coupled plasma mass spectrometry

Selenium species, selenite, selenate, selenomethionine (Semet), seneloethionine (Seet) and trimethylselenonium ion (TmSe) were separated in aqueous solution using a gel-permeation (polyvinyl alcohol-based resin) GS-220 column by eluting with 25 mM tetramethylammonium hydroxide and 25 mM malonic acid at pH 7.9. The GS-220 column coupled with inductively coupled plasma mass spectrometry was used for the separation, identification, and quantification of selenium compounds present in certified reference material (CRM) No. 18 human urine from the National Institute for Environmental Studies in Japan (NIES). Spiking of the authentic standard to the urine and use of a silica-based LC-SCX cation-exchange column validated the peak of selenium compounds. High concentrations of chloride and bromide in the urine eluted from the GS-220 column formed molecular ions 40Ar37Cl+ and 81Br1H+ in the plasma, and these molecular ions created additional peaks in the chromatograms when 77Se and 82Se isotopes were monitored respectively. Thus, both the isotopes were selected concurrently for signal monitoring to eliminate the interfering signals. On the LC-SCX column, chloride and bromide were eluted with selenate and complicated its determination, but the peak of TmSe was baseline separated from rest of the Se compounds. Two unknown Se compounds were detected in both the columns. An additional Se compound having the same retention time as that of Semet was detected on the LC-SCX column. Peaks of selenite, selenate, TmSe and unknown selenium compounds in the urine were baseline separated on the GS-220 column, and were free from interferences. Therefore, the GS-220 column was used for the determination of selenium compounds in NIES CRM No. 18. Unknown Se compounds were the predominant selenium species followed by selenite, TmSe and selenate. The estimated value of TmSe as Se, by the standard additions method using the GS-220 column, was 3.42 +/- 0.17 microg l(-1) and was in good agreement with the LC-SCX value [3.38 +/- 0.21 (n=5) microg l(-1)].     

The simultaneous microdetermination of carbon,hydrogen, arsenic,or selenium in organic compounds

Simple modifications have been described for the simultaneous microdetermination of carbon, hydrogen, and arsenic or selenium in organic compounds using the rapid straight empty tube combustion technique. A silica insertion tube placed in the combustion tube at the open end of the ignition capsule and cooled via external exposure of the combustion tube around it to a stream of air, was found to be suitable for the quantitative retention of arsenic trioxide produced during the combustion of organo-arsenic compounds. The same modification was proved to be suitable for complete trapping of selenium dioxide produced in the combustion of organo-selenium compounds. In the latter case, the insertion tube was packed at one end with a short silica wool plug and cooling it with air is unnecessary.Using these simple modifications, accurate and reproducible carbon, hydrogen, and arsenic or selenium results were obtained in the analysis of a wide range of organic compounds containing arsenic or selenium. The conventional gravimetric finish was employed for the evaluation of three elements.     

Speciation analysis of arsenic and selenium compounds in environmental and biological samples by ion chromatography-inductively coupled plasma dynamic reaction cell mass spectrometer

An inductively coupled plasma mass spectrometer (ICP-MS) was used as an ion chromatographic (IC) detector for the speciation analysis of arsenic and selenium. The arsenic and selenium species studied included arsenite [As(III)], arsenate [As(V)], monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), arsenobetaine (AsB), selenite [Se(IV)] and selenate [Se(VI)]. Gradient elution using (NH₄)₂CO₃ and methanol at pH 9 allowed the chromatographic separation of all species in less than 12 min. Effluents from the IC column were delivered to the nebulization system of ICP-DRC-MS for the determination of arsenic and selenium. The potentially interfering ³⁸Ar⁴⁰Ar⁺ and ⁴⁰Ar⁴⁰Ar⁺ at the selenium masses m/z 78 and 80 were reduced in intensity by approximately 3 orders of magnitude by using 0.6 mL min⁻¹ CH₄ as reactive cell gas in the DRC while an Rpq value of 0.3 was used. Meanwhile, arsenic was determined as the adduct ion ⁷⁵As¹²CHH⁺ at m/z 89, which is more sensitive than ⁷⁵As. The limits of detection for arsenic and selenium were in the range of 0.002–0.01 ng mL⁻¹ and 0.01–0.02 ng mL⁻¹, respectively, based on peak height. The relative standard deviation of the peak areas for five injections of 5 ng mL⁻¹ As and Se mixture was in the range of 2–4%. The concentrations of arsenic and selenium species have been determined in urine samples collected locally. The major As and Se species in urines were AsB, DMA and probably selenosugar at concentration of 20–40, 15–19 and 17–31 ng mL⁻¹, respectively. The recoveries were in the range of 94–105% for all the determinations. This method has also been applied to determine various arsenic compounds in two fish samples. In this study, a simple and rapid microwave-assisted extraction method was used for the extraction of arsenic compounds from fish. The arsenic species were quantitatively leached with an 80% v/v methanol solution in a focused microwave field during a period of 5 min.     

The retention behavior of arsenic compounds on PRP-X100 column under alkaline condition.

系统地研究了碱性条件下（ｐＨ８～１０．８）Ａｓ３＋,Ａｓ５＋,ＭＡ,ＤＭＡ和ＡＢ等砷化合物在ＰＲＰ－Ｘ１００阴离子交换柱上的保留行为。用火焰原子吸收光谱（ＦＡＡＳ）测定从ＨＰＬＣ分离的砷化合物,即通过一根１ｍ×０．２３ｍｍｉ．ｄ．不锈钢毛细管,将ＨＰＬＣ柱出口与ＦＡＡＳ的雾化器连接起来,采用乙炔／空气火焰,在１９３．７ｎｍ处测定。具体研究了两个流动相（２０ｍｍｏｌ／ＬＮＨ４ＨＣＯ３和２．５ｍｍｏｌ／Ｌ对－羟基苯甲酸－１．０ｍｍｏｌ／Ｌ苯甲酸水溶液）。     

Retention behavior of arsenobetaine, arsenocholine, trimethylarsine oxide and tetramethylarsonium iodide on a styrene-divinylbenzene column with benzenesulfonates as ion-pairing reagents

The pH-dependent retention behavior of arsenobetaine, arsenocholine, trimethylarsine oxide, tetramethylarsonium iodide (cationic arsenic compounds), arsenite, arsenate, methylarsonic acid, and dimethylarsinic acid (anionic arsenic compounds) was studied on a Hamilton PRP-1 reversed-phase column (250×4.1 mm I.D.) with 10 mM aqueous solutions of benzensulfonic acids (X-C₆H₄SO₃⁻; X=H, 4-HO, 3-CO₂H; 4-HO-3-HO₂C-C₆H₃SO₃⁻) as ion-pairing reagents in the pH range 2–5 using flame atomic absorption spectrometry as the arsenic-specific detector. The dependencies of the k′-values of the ‘cationic’ arsenic compounds was rationalized on the basis of the protonation/deprotonation behavior of the arsenic compounds and of the four benzenesulfonates. The results provided evidence for the formation of a cationic species from trimethylarsine oxide below pH 3. Benzenesulfonate is the most hydrophobic ion-pairing reagent causing strong retention of the cationic arsenic compounds and consequently impeding their rapid separation. With the less hydrophobic, substituted benzenesulfonates the cationic arsenic compounds had retention times not exceeding 6 min. At a flow-rate of 1.5 cm³ min⁻¹ 10 mM aqueous 3-carboxy-4-hydroxybenzenesulfonate solution adjusted to pH 3.5 allowed the separation of arsenate, methylarsonic acid, arsenobetaine, trimethylarsine oxide, the tetramethylarsonium ion, and arsenocholine within 3 min. Dimethylarsinic acid coelutes with arsenobetaine at pH 3.5, but can be separated from arsenobetaine with the same mobile phase at pH 2.5. At pH 2.5 the signals for trimethylarsine oxide, the tetramethylarsonium ion, and arsenocholine are too broad to be useful for quantification. Arsenite and methylarsonic acid cannot be separated under these conditions.     

Determination of inorganic oxyanions of As and Se by HPLC-ICPMS

A liquid chromatographic separation of inorganic oxyanions of As (As(V) and As(III)) and Se (Se(VI) and Se(IV)) using mixed ion-pairing reagents followed by ICPMS detection is described. The separation was accomplished in less than 4 min on Capcell C18 RP column using mixed ion-pairing modifier containing 5 mM of butane sulfonic acid (BSA), 2 mM malonic acid, 0.30 mM hexane sulfonic acid (HSA) and 0.5% methanol of pH 2.5. All four species were resolved with retention times of 2.4, 2.6, 3.0, and 3.1 min for Se(VI), As(V), As(III), and Se(IV), respectively. The detection limits were less than 0.08 and 0.77 μg l⁻¹ for arsenic and selenium species, respectively. The relative standard deviation of the proposed method for arsenic (at 2.5 μg l⁻¹) and selenium (at 10 μg l⁻¹) was less than 3.7 and 4.8%, respectively. The technique was used to determine inorganic oxyanions of As and Se in water samples (tap, well, and river) and extracts of coal fly ash and sediment. Low power microwave digestion was employed for extraction from fly ash and sediment samples.     

Reversed-Phase Separation of the Major Deoxyribonucleosides and Their Mononucleotides Using Tetrabutylammontum Hetaerons

Abstract

Simultaneous separation of the four major deoxyribonucleosides and their monophosphate nucleotides was achieved using tetrabutyl ammonium phosphate hetaerons with a reversed-phase (C8) packing material. Baseline resolution for all eight solutes was achieved within 48 minutes, using a 7.5% methanol mobile phase, 2.0 mM in TBA, buffered with 50 mM phosphate at pH 4.8. The effect of methanol and TBA concentrations upon the retention of neutral and anionic solutes was studied in detail. It was determined that changes in solute k' with increasing methanol could be explained by essentially independent phenomena. These are: 1) a decrease in the partition coefficient of the TBA cation with increasing organic concentration, resulting in lower surface charge densities, and 2) a decrease in the hydrophobic interactions of the solutes with the reversed-phase HPLC. The overall effect was a log-linear decrease in k' with increasing methanol concentration. An empirical equation was derived for the above model which was found to be helpful in determining the optimal separation conditions for the nucleosides and nucleotides.     

Separation and Detection of Arsenic and Selenium Species in Environmental Samples by HPLC-ICP-MS

A method is presented for arsenic speciation analysis of an oyster sample using ion chromatography coupled with an inductively coupled plasma mass spectrometry (ICP-MS) instrument. A strong anion exchange resin was employed with a step gradient elution of 0.1 mM/0.1 M K 2 SO 4 at pH 10.2. Arsenobetaine and dimethylarsinic acid were determined following extraction based on trypsin enzymolysis with 95-100% extraction efficiency. Limits of detection in the range 0.1-0.3 mg kg m 1 of arsenic were obtained for organic arsenic species. No inorganic arsenic was detected. Validation was performed using TORT-2 as a certified reference material. Although high performance liquid chromatography (HPLC) coupled to ICP-MS is an effective method for speciation analysis it is not always necessary to obtain such a detailed picture. A simple liquid chromatographic separation technique based upon mini-column technology is presented. It was developed to obtain a fast, efficient and reliable separation of inorganic from organic, i.e. assumed toxic from non-toxic, arsenic and selenium species suitable for use as an initial screening method for environmental analysis. Two types of strong anion exchange resin were tested. Excellent separation was obtained for both min-column resins and analysis times were within 7 min. Limits of detection obtained for inorganic arsenic, organic arsenic, selenomethionine, Se IV and Se VI were 1.6, 1.8, 66, 32 and 22 µg kg m 1 , respectively.     

离子色谱-双阳极电化学氢化物发生-原子荧光光谱法测定Ⅰ型牙髓失活材料中的砷形态

将自制的双阳极电化学氢化物发生器作为离子色谱与原子荧光光谱的联用接口,建立了离子色谱-双阳极电化学氢化物发生-原子荧光光谱法在线分析砷形态系统.最佳实验条件为淋洗液6.0 mmol/L NH4H2PO4(pH 6.20),电解液0.40 mol/L H2SO4,电解液流量为阳极4.0 mL/min,阴极 1.5 mL/min,电解电流密度0.50 A/cm.2,载气流量300 mL/min,屏蔽气流量500 mL/min,氢气流量80 mL/min.在优化的实验条件下,As(Ⅲ)、DMA、MMA的线性范围为5～200 μg/L、As(Ⅴ)的线性范围为10～200 μg/L,As(Ⅲ)、DMA、MMA和As(Ⅴ)检出限分别为3.04、4.27、3.97和9.30 μg/L(信噪比S/N=3).50 μg/L的As(Ⅲ)、DMA、MMA和As(Ⅴ)混合标准溶液平行进样7次,得到的色谱峰面积的相对标准偏差(RSD)分别为415%、3.08%、519%和3.62%.将该方法用于牙髓失活材料中的砷形态分析.     

Separation and determination of lignans from seeds of Schisandra species by micellar electrokinetic capillary chromatography using ionic liquid as modifier

In this paper, a micellar electrokinetic chromatographic (MEKC) method using ionic liquid as modifier for the quantification of the active components of lignans found in the medicinal herbs Schisandra species was developed for the first time. Preliminary investigations employing sodium dodecyl sulfate (SDS) as surfactant did not lead to the necessary resolution of the studied compounds, the addition of ionic liquid 1-butyl-3-methylimidazolium tetrafluoroborate (BMIM-BF4) to the SDS micellar system resulted in the complete separation of all the compounds. The effects on the separation by several parameters such as BMIM-BF4 and SDS concentration, applied voltage, background electrolyte pH and concentration, were evaluated. Under the optimal conditions (5 mM borate-5 mM phosphate buffer in the presence of 20 mM SDS and 10 mM BMIM-BF4, pH 9.2, applied voltage 25 kV and detection at 254 nm), the method successfully applied to the determination of lignans in extracts of Schisandra chinensis (Turcz.) Baill. and Schisandra henryi C.B. Clarke in less than 13 min. The separation mechanism was also discussed.     

Capillary electrophoretic separation of inorganic and organic arsenic compounds

Capillary zone electrophoresis was used to separate arsenite, arsenate, dimethylarsinic and diphenylarsinic acid, methanearsonic acid, phenyl- and p-aminophenyl arsonic acid, phenylarsineoxide and phenarsazinic acid. Anionic and uncharged species were separated in a fused silica capillary with on-column UV detection at 200 nm. A 15 mM phosphate solution adjusted to pH 6.5 containing 10 mM sodium dodecylsulfonate served as background electrolyte. The influence of pH and applied voltage on separation efficiency, as well as the feasibility of identification of arsenic compounds in spiked urine, were investigated. Received: 18 March 1998 / Revised: 25 May 1998 / Accepted: 30 May 1998     

Determination of selenoamino acids using two-dimensional ion-pair reversed phase chromatography with on-line detection by inductively coupled plasma mass spectrometry

Analytical methods for the speciation of nine selenium species (selenite, selenate, selenourea, trimethylselenonium ion, selenocystamine, selenocystine, selenocysteine, selenomethionine and selenoethionine) that are commonly encountered in biological and environmental samples were developed. Good separation was achieved by either a mixed ion-pair reversed phase chromatography (LiChrosorb RP 18, 2.5 mM 1-butanesulfonate-8 mM tetramethylammonium hydroxide-4 mM malonic acid-0.05% methanol, pH 4.5) or a conventional ion-pair reversed phase chromatography (Inertsil ODS, 10 mM tetraethylammonium hydroxide-4.5 mM malonic acid, pH 6.8) with on-line ICP-MS detection. Using a 20-μl sample loop, low detection limits around 1 ng ml⁻¹ expressed as Se were achieved for the examined selenium species. The methods were used for the determination of selenoamino acids in a selenium nutritional supplement. The developed methods were found to be rather robust. No alteration of the separation was observed when the protease enzymatic extracts were analyzed without dilution. Both water extracts and enzymatic extracts were chromatographed first with the mixed ion-pair reversed phase chromatographic system, then the major chromatographic peaks were collected and analyzed by the second ion-pair reversed phase chromatographic system for a further verification of their identity. Selenomethionine was found to be the major selenium species in the supplement. A major unknown species, probably Se-adenosylhomocysteine, could be determined in the extracts. A biological reference material, Dolt-2, was also examined for the selenoamino acids. Selenocystine and selenomethionine could be detected in its enzymatic extract, suggesting that Dolt-2 may be used as a reference material for the identification of selenoamino acids in biological and environmental samples. As selenoethionine does not occur naturally in the investigated samples, it is added as an internal standard in this study.     

Speciation analysis of arsenic and selenium compounds by capillary electrophoresis

High-performance capillary electrophoresis is applied to the separation of different inorganic and organic arsenic and selenium compounds. In comparison with UV-detection, an approach with conductivity detection is described expecting higher sensitivity and universality. In this case the capillary was statically modified with CTAB before the electromigration procedure. The separation was performed with an electrolyte system consisting of CHES and Triton X-100. Detection limits of 0.06 mg/L or lower were obtained for As(V) and Se(VI). Water samples of an arsenic-polluted tailing of tin mining processes were analysed for anions as well as arsenic and selenium species.