Determination of cobalamins using capillary electrophoresis inductively coupled plasma mass spectrometry

The determination of cobalamins using capillary electrophoresis inductively coupled plasma mass spectrometry (CE-ICP-MS) was investigated. Both capillary zone electrophoresis (CZE) and micellar electrokinetic chromatography (MEKC) modes of operation were studied. The optimal separation of four cobalamin species (cyanocobalamin, hydroxocobalamin, methylcobalamin, and 5′-deoxyadenosylcobalamin) and a potentially harmful corrinoid analogue (cobinamide dicyanide) was obtained using CZE at a pH of 2.5. Both 20 mM phosphate and 20 mM formate buffers were used with success, although the formate buffer provided improved resolution. The CZE-ICP-MS method was used to quantify cyanocobalamin in a vitamin supplement and the analytical results were in good agreement (±5%) with values obtained by ICP-MS for total Co levels. The solution detection limits for cobalamins using CZE-ICP-MS were approximately 50 ng/ml. MEKC was found to be useful for the screening of vitamin preparations because it provided a rapid means of distinguishing cyanocobalamin (the form most commonly used in vitamin preparations) from free cobalt. The separation of free cobalt and cyanocobalamin using MEKC was achieved in less than 10 min.     

Hyphenation of capillary electrophoresis to inductively coupled plasma mass spectrometry as an element-specific detection method for metal speciation

A stepwise development for the use of capillary electrophoresis and inductively coupled plasma mass spectrometry (ICP-MS) for speciation investigations is presented. The high resolution power of CE is used for the separation of metal species, whereas ICP-MS is taken for element-specific detection with low detection limits. This contribution starts with an off-line combination of both instruments. Separation and identification of species in model solutions and real samples are shown by scanning UV detection at the CE unit with subsequent metal quantification in peak related fractions, applying electrothermal vaporization ICP-MS. Finally, first separations are demonstrated, using the on-line hyphenation with a laboratory-made nebulizer. Here, standard solutions are separated and monitored by UV and ICP-MS. Stability of electrical current during nebulization was checked and a possibly interfering suction flow was estimated. After optimization sufficient electropherograms were obtained. Advantages and problems are discussed for both modes.     

Capillary electrophoresis inductively coupled plasma mass spectrometry

Chemical speciation (extraction of elemental information and identification of molecular environment for an analyte in a complex sample) has been a long sought after goal for analytical chemists. Recently, because of successful developments in more sensitive element-specific detectors and gentle separation schemes, which preserve the true chemical information in a real sample, routine speciation experiments are becoming a common occurrence in the scientific literature. For many reasons, the combination of capillary electrophoresis (for separation of different chemical species) with inductively coupled plasma mass spectrometry (for element and isotope specific detection) has emerged as the method of choice for these analyses. In this article the basic principles of capillary electrophoresis inductively coupled plasma mass spectrometry are discussed. Design consideration for instrument interface, anticipated difficulties with speciation experiments and applications for specific matrices and analytes are also presented in this article.     

Determination of monophosphate nucleotides by capillary electrophoresis inductively coupled plasma mass spectrometry

A preliminary study of a modified microconcentric nebulizer (CEI-100, CETAC) as the sample introduction device of capillary electrophoresis inductively coupled plasma mass spectrometry (CE-ICP-MS) for the determination of monophosphate nucleotides is described. The monophosphate nucleotides studied include adenosine 5'-monophosphate (AMP), guanosine 5'-monophosphate (GMP), uridine 5'-monophosphate (UMP) and inosine 5'-monophosphate (IMP). The species studied were well separated using a 70 cm length x 75 microm id fused silica capillary while the applied voltage was set at -22 kV and a 20 mmol l(-1) ammonium citrate/citric acid buffer (pH 4.0) containing 0.1% m/v cationic polymer (hexadimethrine bromide, Polybrene) was used as the electrophoretic buffer. The electroosmotic flow was reversed by flushing the fused silica capillary with 0.2% m/v Polybrene to accelerate separation. The detection limit of various species studied was in the range of 0.036-0.054 microg P ml(-1), which corresponded to the absolute detection limit of 1.1-1.6 pg P based on the injection volume of 30 nl. We determined the concentrations of nucleotides in two IG-enriched monosodium glutamates purchased from the local market. The recovery was in the range of 100-112% for various species, and the concentrations of IMP and GMP in these samples were in the range of 0.15-0.18% m/m.     

Interface for capillary electrophoresis coupled with inductively coupled plasma atomic emission spectrometry.

A modified concentric nebulizer was used as the interface to couple capillary electrophoresis (CE) to inductively coupled plasma atomic emission spectrometry (ICP-AES). The CE capillary replaces the central tube of the concentric nebulizer. The tip of the nebulizer tapers slowly to allow uncertainty in the position of the capillary. A platinum wire was inserted into the CE capillary to provide electrical connection to the CE power supply. pH changes inside the capillary due to electrolysis of the background buffer electrolyte was small and has minimal effects on the CE separation. The peak broadening effects due to the nebulizing gas flow, however, were significant. Resolution decreases quickly when the flow-rate of the carrier gas increases. Sample stacking technique was used to improve the resolution of species of opposite charge, e.g., Cr(VI) vs. Cr(III) ions. Detection limit of Cr based on peak area is approximately 10 ppb for the CE-ICP-AES system.     

Characterization of microconcentric nebulizer uptake rates for capillary electrophoresis inductively coupled plasma mass spectrometry

There is demonstrated interest in combining capillary electrophoresis (CE) with inductively coupled plasma mass spectrometry (ICP-MS) for speciation determinations. When self-aspirating nebulizers are used for this application, it is important to offset the suction effect to avoid degradation of the separation. In this study, sample uptake rates for three microconcentric nebulizers of the same model, in combination with a cyclonic spray chamber, were characterized and compared for future utilization in CE–ICP-MS interfaces. The specific model studied was a MicroMist with a nominal uptake rate of 100 μl/min at 1 l/min argon gas flow rate per the manufacturer's specifications. Sample uptake rates at various nebulizer gas flows were measured by aspirating water from a weighed container and calculating the uptake rate in microliter per minute. The nebulizers studied provided good reproducibility from day to day, but a comparison of the different nebulizers reflected a significant difference in performance. A characteristic observed during the study was that uptake rates decreased with increasing nebulizer gas flow. This can be used for sample introduction for CE–ICP-MS. Interestingly, very different performance was observed when comparing the three different nebulizers of the same model. Uptake rates showed strong dependence on argon gas flow rates and the dimensions of the sample uptake tubing.     

Separation of metalloporphyrins by capillary electrophoresis with UV detection and inductively coupled plasma mass spectrometric detection

Vitamin B12, cobalt protoporphyrin, manganese protoporphyrin, and zinc protoporphyrin were separated using capillary electrophoresis, and a comparison was made between detection with inductively coupled plasma mass spectrometry (ICP-MS) and UV detection. Absolute limits of detection were slightly better with ICP-MS detection than with UV detection, but for both methods absolute detection limits were in the picogram range. The migration times of the analytes decreased by several minutes when ICP MS detection was employed, and this phenomenon was believed to be a result of a "suction effect" that developed when the CE capillary was interfaced to the ICP-MS nebulizer. However, the resolution between species containing the same metal atom was not altered significantly, and the separation was completed in much less time relative to separations performed with UV detection.     

A new interface used to couple capillary electrophoresis with an inductively coupled plasma mass spectrometry for speciation analysis

In this work, a novel and high-efficiency interface has been developed in coupling CE with inductively coupled plasma MS (ICPMS). The interface completely avoids laminar flow in CE capillary caused by the suction of nebulizer, and can be easily and stably operated at room temperature with high analyte transport efficiency to ICPMS. The new interface has a liquid dead volume smaller than 5 nL, which was much smaller than those (65-2500 microL) reported previously for other interfaces. All above features led to a higher sensitivity and a better electrophoretic resolution for CE-ICPMS coupled with this new interface. With the help of this new interface, we have successfully separated and determined five species of arsenic, As(III), As(V), monomethylarsonic acid, dimethylarsinic acid and p-aminophenylarsonic acid using CE-ICPMS within 11 min with a detection limit of 0.046-0.075 ng/mL and an RSD of 2-6% (n=6).     

Quantification of phosphorus in DNA using capillary electrophoresis hyphenated with inductively coupled plasma mass spectrometry

We have analyzed phosphorus in an enzymatically digested DNA molecule using capillary electrophoresis (CE) hyphenated with inductively coupled plasma mass spectrometry (ICP-MS). The DNA concentration was quantified by the phosphorus value obtained in the CE-ICP-MS analysis. The CE-ICP-MS measurement, for which the interface device AIF-01 equipped three layered nebulizer was adopted, was achieved with limited μL/min nebulizing without loss of sample in the vaporizing chamber. The samples of nucleotides and free phosphate were separated well in the CE-ICP-MS measurement, and the calibration curve (0.1-10μg/mL) of the phosphorus showed a linear (R(2)=0.999) increase in intensity. After digestion of the 100-bp double-strand DNA sample to deoxyribonucleotide-5'-monophosphates (dNMPs) by phosphodiesterase-I, phosphorus was detected by CE-ICP-MS without further purification steps. In this study, we applied two calculation schemes of DNA analysis using a dNMP concentration obtained from CE-ICP-MS. Comparative CE-ICP-MS analysis with DNA digested to dNMPs showed that the assay gave an equal value obtained from the total DNA quantification using fluorescence detection. The detection limits of the DNA sample obtained from these species and phosphorus in nucleotides using CE-ICP-MS were 3.1-26ng/mL. These LOD values were equal to the conventional fluorescence determination of DNA.     

Identification of a metallothionein in Synechococcus by capillary electrophoresis hyphenated with inductively coupled plasma mass spectrometry.

A home-made system hyphenating capillary electrophoresis with an inductively coupled plasma mass spectrometer (CE-ICP-MS) for cadmium speciation of protein-binding and free cadmium ions in solution is presented. The CE-ICP-MS interface consisted of an acrylic block with an internal volume ca. 20 microL in which a platinum electrode, a capillary column, and a connection to an ICP nebulizer were inserted. A make-up electrolyte solution containing 50 mmol L(-1) Tris-HCl buffer solution (pH 9.0) was continuously flowed through the interface to the ICP nebulizer. The separation of free Cd ions, Cd-cysteine, and Cd bounded to metallothionein (MT) isoforms from rabbit liver was carried out by capillary electrophoresis, and the analytes were detected by ICP-MS. The feasibility to isolate metallothionein compounds extracted from the cyanobacterium Synechococcus PCC7942 was demonstrated. The Cd binding proteins were induced in Synechococcus PCC7942 and further analyzed by CE ICP-MS.     

Simultaneous detection of zinc dimethyldithiocarbamate and zinc ethylenebisdithiocarbamate in cabbage leaves by capillary electrophoresis with inductively coupled plasma mass spectrometry

We report a simple and highly sensitive method for the simultaneous detection of trace zinc dimethyldithiocarbamate and zinc ethylenebisdithiocarbamate by capillary electrophoresis with inductively coupled plasma mass spectrometry. Zinc dimethyldithiocarbamate and zinc ethylenebisdithiocarbamate were chelated with trans‐1,2‐diaminocyclohexane‐N,N,N′,N′‐tetraacetic acid to form a macromolecule complex. Then, these two compounds were separated by α‐cyclodextrin‐modified capillary electrophoresis within 12 min at a separation voltage of 15 kV and measured by inductively coupled plasma mass spectrometry. The developed method is sensitive with detection limit of 1.9 and 3.0 ng Zn/mL for zinc dimethyldithiocarbamate and zinc ethylenebisdithiocarbamate, respectively. By means of ultrasound‐assisted extraction methods, zinc dimethyldithiocarbamate and zinc ethylenebisdithiocarbamate spiked into cabbage leaves were successfully extracted and determined with a relative standard deviation (n  = 5) ≤ 6% and a recovery of 95–107%.     

Element-tagged immunoassay with inductively coupled plasma mass spectrometry for multianalyte detection

The multianalyte immunoassay approach is currently attracting increasing attention due to its high sample throughput, short assay time, low sample consumption and reduced overall cost per assay. This paper reviews progress in the field of multianalyte immunoassays using inductively coupled plasma mass spectrometry, as well as applications of this approach in different fields. Examples related to the combination of protein microarray technology with the multitag approach of the immunoassay ICP-MS method and to the use of ICP-MS in the field of imaging are described. A novel strategy that involves tagging antibodies for ICP-MS detection in sensitive multitag bioassays is also presented. Finally, the outlook for this promising technique is discussed.     

Investigation of a liposomal oxaliplatin drug formulation by capillary electrophoresis hyphenated to inductively coupled plasma mass spectrometry (CE-ICP-MS)

A capillary electrophoresis-inductively coupled plasma mass spectrometry (CE-ICP-MS) method was developed for separation of the free oxaliplatin drug substance from liposome-entrapped oxaliplatin. Simultaneous determination of phosphorous and platinum opened the possibility to simultaneously monitor the liposomes (phospholipids) and platinum-based drug. In order to suppress the interferences, argon gas was used as a collision gas in ICP-MS. A detection limit of 29 ng/mL of platinum and a precision of 2.9% (for 10 μg/mL of oxaliplatin standard) were obtained. Measurement of the total concentration of free and encapsulated oxaliplatin by CE-ICP-MS was compared with total determination by ICP-MS after microwave digestion and showed a good agreement. A liposomal formulation of oxaliplatin based on PEGylated liposomes was used as a model drug formulation. Studies of accelerated drug release induced by sonication and phospholipase A₂ catalyzed hydrolysis were performed. It was demonstrated that the CE-ICP-MS was an efficient in vitro characterization method in the development and quality assurance purposes of lipsome-based formulation of metallodrugs.     

A study of the interactions between carboplatin and blood plasma proteins using size exclusion chromatography coupled to inductively coupled plasma mass spectrometry

To study the carboplatin–protein interaction, a sensitive method using size exclusion chromatography coupled to inductively coupled plasma mass spectrometry (SEC–ICP–MS) was developed. The complexes formed between plasma proteins and carboplatin were monitored and identified with this method. Composite blood plasma samples from patients who were undergoing chemotherapy were analyzed, and carboplatin was found to bind plasma proteins. In addition, blank plasma samples were spiked with carboplatin and were analyzed as a time course study, and the results confirmed that carboplatin formed complexes with plasma proteins, primarily albumin and γ-globulin. To further substantiate the study, these two proteins were incubated with carboplatin. The binding between carboplatin and these proteins was then characterized qualitatively and quantitatively. In addition to a one-to-one binding of Pt to protein, protein aggregation was observed. The kinetics of the binding process of carboplatin to albumin and γ-globulin was also studied. The initial reaction rate constant of carboplatin binding to albumin was determined to be 0.74 M⁻¹ min⁻¹, while that for γ-globulin was 1.01 M⁻¹ min⁻¹, which are both lower than the rate constant of the cisplatin–albumin reaction previously reported.     

A new interface for combining capillary electrophoresis with inductively coupled plasma-mass spectrometry

This paper describes the development and design of a new, efficient, simple and robust interface for coupling capillary electrophoresis (CE) with inductively coupled plasma-mass spectrometry. The interface is based on a modified microconcentric nebulizer which permits a low flow rate of about 6 μL/min in the free aspiration mode. This interface construction provides an electrical connection for stable electrophoretic separations and adapts the flow rate of the electro-osmotic flow inside the CE capillary to the flow rate of the nebulizer for efficient transport of the analytes into the plasma. By optimization of the fluid mechanical properties the interface prevents the nebulizer from causing any laminar flow in the CE capillary and thus the high resolution power of CE can be preserved. Furthermore, this new device permits independent optimization of the nebulization from the CE whereby exact positioning of the CE capillary is not necessary, thus enabling fast exchange. A low dead volume spraychamber has been constructed which circumvents any band broadening of the sharp CE signals. Peak widths down to 3.5 s comparable to CE with UV detection are possible. Received: 5 February 1999 / Revised: 21 April 1999 / Accepted: 23 April 1999     

A new interface for combining capillary electrophoresis with inductively coupled plasma-mass spectrometry

This paper describes the development and design of a new, efficient, simple and robust interface for coupling capillary electrophoresis (CE) with inductively coupled plasma-mass spectrometry. The interface is based on a modified microconcentric nebulizer which permits a low flow rate of about 6 μL/min in the free aspiration mode. This interface construction provides an electrical connection for stable electrophoretic separations and adapts the flow rate of the electro-osmotic flow inside the CE capillary to the flow rate of the nebulizer for efficient transport of the analytes into the plasma. By optimization of the fluid mechanical properties the interface prevents the nebulizer from causing any laminar flow in the CE capillary and thus the high resolution power of CE can be preserved. Furthermore, this new device permits independent optimization of the nebulization from the CE whereby exact positioning of the CE capillary is not necessary, thus enabling fast exchange. A low dead volume spraychamber has been constructed which circumvents any band broadening of the sharp CE signals. Peak widths down to 3.5 s comparable to CE with UV detection are possible. Received: 5 February 1999 / Revised: 21 April 1999 / Accepted: 23 April 1999     

A simple and demountable capillary microflow nebulizer with a tapered tip for inductively coupled plasma mass spectrometry

A simple and demountable capillary microflow nebulizer (d-CMN) was developed for inductively coupled plasma mass spectrometry (ICP-MS). It consisted of a nebulizer body, a fused-silica capillary with a tapered tip and a polytetrafluoroethylene (PTFE) adapter. The gas orifice i.d., the solution capillary tip i.d. and its wall thickness were 200, 30, and 5 μm, respectively. The sensitivities, detection limits, precisions and the long-term stability with the d-CMN were evaluated. The experimental results indicated that its performances at low uptake rates were similar or better than those obtained with the conventional concentric nebulizer at 820 μL/min and the micromist nebulizer at 200 μL/min. The demountable construction of the d-CMN permitted that the blocked or broken solution capillary could be conveniently renewed. The low self-aspiration rate (4.77 μL/min) and the analytical characteristics comparable to commercial microflow nebulizers made the d-CMN a good choice for coupling capillary electrophoresis and microbore high-performance liquid chromatograph to ICP-MS. The proposed d-CMN was successfully applied in the iodine speciation by coupling microchip capillary electrophoresis (MCE) to ICP-MS. The absolute detection limits for iodide and iodate of 0.20 and 0.29 fg were achieved with satisfactory resolution.     

Determination of arsenic species in Solanum Lyratum Thunb using capillary electrophoresis with inductively coupled plasma mass spectrometry

A simple and highly efficient interface to couple capillary electrophoresis with inductively coupled plasma mass spectrometry by a microflow polyfluoroalkoxy nebulizer and a quadruple ion deflector was developed in this study. By using this interface, six arsenic species, including arsenite, arsenate, monomethylarsonic acid, dimethylarsinic acid, arsenobetaine, and arsenocholine, were baseline‐separated and determined in a single run within 11 min under the optimized separation conditions. The instrumental detection limit was in the range of 0.02–0.06 ng/mL for the six arsenic compounds. Repeatability expressed as the relative standard deviation (n = 5) of both migration time and peak area were better than 2.5 and 4.3% for six arsenic compounds. The proposed method, combined with a closed‐vessel microwave‐assisted extraction procedure, was successfully applied for the determination of arsenic species in the Solanum Lyratum Thunb samples from Anhui province in China with the relative standard deviations (n = 5) ≤4%, method detection limits of 0.2–0.6 ng As/g and a recovery of 98–104%. The experimental results showed that arsenobetaine was the main speciation of arsenic in the Solanum Lyratum Thunb samples from different provinces in China, with a concentration of 0.42–1.30 μg/g.     

Determination of sulfur-containing amino acids by capillary electrophoresis dynamic reaction cell inductively coupled plasma mass spectrometry

Capillary electrophoresis dynamic reaction cell™ inductively coupled plasma mass spectrometry (CE-DRC-ICP-MS) for the determination of sulfur-containing amino acids is described. The sulfur-containing amino acids studied include l-cysteine, l-cystine, dl-homocystine and l-methionine. The species studied were well separated using a 70 cm length×75 μm i.d. fused silica capillary while the applied voltage was set at +22 kV and a 10 mmol l⁻¹ disodium tetraborate buffer (pH 9.8) containing 0.1 mmol l⁻¹ EDTA and 0.5 mmol l⁻¹ Triton X-100 was used as the electrophoretic buffer. The sulfur-selective electropherogram was determined at m/z 48 as by using its reaction with O₂ in the reaction cell. The method avoided the effect of polyatomic isobaric interferences at m/z 32 caused by and on by detecting as the oxide ion at m/z 48, which is less interfered. The detection limit of various species studied was in the range of 0.047-0.058 μg S ml⁻¹, which corresponded to the absolute detection limit of 1.3-1.6 pg S based on the injection volume of 27 nl. We determined the concentrations of selected sulfur-containing amino acids in urine and nutritive complement samples. The recovery was in the range of 92-128% for various species.