Indirect UV Absorption Detection Research Articles

Development of a background electrolyte for the determination of inorganic cations in high ionic strength samples by capillary electrophoresis with indirect UV-absorption detection

In this study, a background electrolyte capable to separate and quantify inorganic cations in high ionic strength samples by UV-absorption indirect detection was designed. In this regard, the four most abundant monovalent and divalent cations in earth crust (K+, Na+, Ca+2, Mg+2) were selected as model compounds. A group of small carboxylic acids and, several toluidines and pyridines were evaluated as mild strength complexing agents and chromophoric probes, respectively. The optimized background electrolyte was composed of 200 mM 2,4,6-trimethylpyridine as the chromophoric probe, 250 mM lactic acid as the weak complexing agent and pH buffering reagent (adjusted to pH 4.5), and 5% v/v methanol as organic solvent modifier. Based on a minimum number of components, it provided outstanding separation performance in less than 4 min in a wide linear dynamic range (10 - 2500 µg·mL−1). Performances were contrasted against a reference method based on conductometric detection. Furthermore, studies of separation efficiency and peak shape were carried out at different analyte concentrations in high electric conductivity solutions. The herein developed method demonstrated exceptional features in terms of limits of detection (~10 µg·mL−1), resolution, speed of analysis, sensitivity and peak capacity in high electric conductivity samples. Moreover, the method was successfully applied to high ionic strength samples such as rock digest, sea water, soy sauce and isotonic drinks.

Quantitation of underivatized branched-chain amino acids in sport nutritional supplements by capillary electrophoresis with direct or indirect UV absorbance detection.

The branched-chain amino acids (BCAAs) including leucine (Leu), isoleucine (Ile) and valine (Val) play a pivotal role in the human body. Herein, we developed capillary electrophoresis (CE) coupled with conventional UV detector to quantify underivatized BCAAs in two kinds of sport nutritional supplements. For direct UV detection at 195 nm, the BCAAs (Leu, two enantiomers of Ile and Val) were separated in a background electrolyte (BGE) consisting of 40.0 mmol/L sodium tetraborate, and 40.0 mmol/L β-cyclodextrin (β-CD) at pH 10.2. In addition, the indirect UV detection at 264 nm was achieved in a BGE of 2.0 mmol/L Na2HPO4, 10.0 mmol/L p-aminosalicylic acid (PAS) as UV absorbing probe, and 40.0 mmol/L β-CD at pH 12.2. The β-CD significantly benefited the isomeric separation of Leu, L- and D-Ile. The optimal conditions allowed the LODs (limit of detections) of direct and indirect UV absorption detection to be tens μmol/L level, which was comparable to the reported CE inline derivatization method. The RSDs (relative standard deviations) of migration time and peak area were less than 0.91% and 3.66% (n = 6). Finally, CE with indirect UV detection method was applied for the quantitation of BCAAs in two commercial sport nutritional supplements, and good recovery and precision were obtained. Such simple CE method without tedious derivatization process is feasible of quality control and efficacy evaluation of the supplemental proteins.

Analysis of γ-hydroxy butyrate by combining capillary electrophoresis-indirect detection and wall dynamic coating: application to dried matrices.

γ-Hydroxybutyric acid (GHB) is a powerful central nervous system depressant, currently used in medicine for the treatment of narcolepsy and alcohol dependence. In recent years, it has gained popularity among illegal club drugs, mainly because of its euphoric effects as well as doping agent and date rape drug. The purpose of the present work was the development of a rapid analytical method for the analysis of GHB in innovative biological matrices, namely dried blood spots (DBSs) and dried urine spots (DUSs). The analytical method is based on capillary zone electrophoresis with indirect UV absorption detection at 210nm and capillary wall dynamic coating. The background electrolyte is composed of a phosphate buffer containing nicotinic acid (probe for detection) and cetyltrimethylammonium bromide (CTAB, reversal of electroosmosis in wall dynamic coating). The influence of probe and CTAB concentration, together with buffer pH, on migration time and signal response was investigated. Under the optimized conditions, analytical linearity and precision were satisfactory; absolute recovery values were also high (>90%); the use of dried matrices (DBSs and DUSs) was advantageous as an alternative matrix to classical ones. No interferences were found either from the most common exogenous or from endogenous compounds. This analytical approach can offer a rapid, precise and accurate method for GHB determination in innovative biological samples, which could be important for screening purposes in clinical and forensic toxicology. Graphical Abstract CE method, by combined indirect UV detection and dynamic coating, for GHB determination in DBSs and DUSs.

Capillary Zone Electrophoresis Method for the Determination of Inorganic Anions and Formic Acid in Honey

A capillary zone electrophoresis method for the determination of inorganic anions and formic acid in honey samples was developed for the first time. The complete separation of chloride, nitrate, sulfate, phosphate, and formic acid was achieved with a simple electrolyte composed by 2 mM potassium dichromate as the carrier solution and background absorbance provider and 0.05 mM tetraethylenepentamine (TEPA) as electro-osmotic flow suppressor (pH 4.00). Injection was performed hydrostatically by elevating the sample at 10 cm for 10 s. The running voltage was -27 kV at 25 degrees C. Indirect UV absorption detection was achieved at 254 nm. The detection limit was in the range between 0.03 and 20 mg/kg, and the quantification limits ranged from 1.52 to 20.6 mg/kg. The calibration graphs were linear in the concentration range from the quantification limit to at least 2.5 g/kg for chloride, 0.25 g/kg for nitrate, 0.75 g/kg for sulfate, 1.50 g/kg for phosphate, and 0.75 g/kg for formic acid. Precision data in the honey samples analyzed showed repeatability and reproducibility relative standard deviations lower than 1.4 and 2.4% for migration time and lower than 1.8 and 4.3% for anion content, respectively. Recoveries of anions in honey samples analyzed ranged from 94.4 to 99.8%. Ten honey samples were analyzed to test the proposed method. Mean contents of 260.5, 3.93, 60.5, 139.4, and 209.3 mg/kg were found, respectively, for chloride, nitrate, sulfate, phosphate, and formic acid in analyzed honeys. These results agreed with literature data.

Rapid capillary zone electrophoresis method for the determination of metal cations in beverages

A rapid and reliable capillary zone electrophoresis method for the determination of inorganic cations was developed. The complete separation of K +, Ba 2+, Ca 2+, Na +, Mg 2+, Mn 2+, Ni 2+, Cd 2+, Li + and Cu 2+ can be achieved in 4 min with a simple electrolyte composed by 10 mM imidazole as the carrier buffer and background absorbance provider and acetic acid as the complexing agent (pH 3.60). Injection was performed hydrostatically by elevating the sample at 10 cm for 30 s. The running voltage was +25 kV at room temperature. Indirect UV-absorption detection was achieved at 185 nm. The detection limit was in the range between 0.06 mg/l (Mg 2+) and 0.57 mg/l (K +) and the quantification limits ranged from 0.10 mg/l (Ni 2+) to 0.80 mg/l (Cu 2+). The calibration graphs were linear in the concentration range from the quantification limit till at least 1 g/l in K +, 10 mg/l in Ba 2+, Ca 2+, Mg 2+, Mn 2+, Ni 2+ and Cd 2+, 40 mg/l in Na + and 12 mg/l in Li + and Cu 2+. The repeatability, intraday and interday analysis were ≤1.55% and ≤3.64% for migration time and ≤3.38% and ≤3.63% for peak area. The method developed has been applied to several beverage samples with only a simple dilution and filtration treatment of the sample. The proposed method is simple, fast, cheap and it is achieved with common products in either laboratory. For these reasons, it is a very useful method for routine analysis.

Chloride present in biological samples as a tool for enhancement of sensitivity in capillary zone electrophoretic analysis of anionic trace analytes.

Effects originating from the variability of the sample matrix can be efficiently eliminated when the separation conditions are selected so that compounds of like charge with high concentration referred to as macrocomponents are embodied into the system of transient isotachophoresis. For stacking and separation of anionic trace analytes in biological samples, the presence of chloride is shown to be important to balance out effects of other macrocomponents that act against isotachophoretic stacking. Having acetoacetate, malate, citrate, and some drug metabolites in untreated human serum samples, the stacking mechanism of these compounds in an electrolyte system comprising 5 mM mandelic acid and epsilon -aminocaproic acid, pH 3.8, is explained. Analytes are monitored by indirect UV-absorption detection. Attention is paid to the minimum chloride concentration required with respect to the concentration ratio of phosphate (stacker) and lactate (destacker) present in the sample so as to ensure both stacking and separation of trace analytes. Insight into the separation process is given both with computer simulations and experiments. For selected analytes, the effect of chloride concentration on quantitative evaluation, sensitivity and limit of detection is demonstrated as well. Moreover, the applicability of the mobility window between phosphate and lactate for an additional group of metabolites is sketched.

Determination of γ-hydroxybutyric acid in human urine by capillary electrophoresis with indirect UV detection and confirmation with electrospray ionization ion-trap mass spectrometry

γ-Hydroxybutyric acid (GHB), a minor metabolite or precursor of γ-aminobutyric acid (GABA), acts as a neurotransmitter/neuromodulator via binding to GABA receptors and to specific presynaptic GHB receptors. Based upon the stimulatory effects, GHB is widely abused. Thus, there is great interest in monitoring GHB in body fluids and tissues. We have developed an assay for urinary GHB that is based upon liquid–liquid extraction and capillary zone electrophoresis (CZE) with indirect UV absorption detection. The background electrolyte is composed of 4 m M nicotinic acid (compound for indirect detection), 3 m M spermine (reversal of electroosmosis) and histidine (added to reach a pH of 6.2). Having a 50 μm I.D. capillary of 40 cm effective length, 1-octanesulfonic acid as internal standard, solute detection at 214 nm and a diluted urine with a conductivity of 2.4 mS/cm, GHB concentrations ≥2 μg/ml can be detected. Limit of detection (LOD) and limit of quantitation (LOQ) were determined to be dependent on urine concentration and varied between 2–24 and 5–60 μg/ml, respectively. Data obtained suggest that LOD and LOQ (both in μg/ml) can be estimated with the relationships 0.83 κ and 2.1 κ, respectively, where κ is the conductivity of the urine in mS/cm. The assay was successfully applied to urines collected after administration of 25 mg sodium GHB/kg body mass. Negative electrospray ionization ion-trap tandem mass spectrometry was used to confirm the presence of GHB in the urinary extract via selected reaction monitoring of the m/z 103.1→ m/z 85.1 precursor–product ion transition. Independent of urine concentration, this approach meets the urinary cut-off level of 10 μg/ml that is required for recognition of the presence of exogenous GHB. Furthermore, data obtained with injection of plain or diluted urine indicate that CZE could be used to rapidly recognize GHB amounts (in μg/ml) that are ≥ 4 κ.

Separation of alkali and alkaline earth metal and ammonium cations by capillary zone electrophoresis with indirect UV absorbance detection

Carrier electrolytes combining complexing effects of negatively charged counter ions and electroneutral 18-crown-6 were investigated in the separations of alkali and alkaline earth metal and ammonium cations by capillary zone electrophoresis with indirect UV absorption detection. The use of tartrate with 18-crown-6 at a 0.04 mol 1 −1 concentration permitted the complete resolution of these cations in one electrophoretic run. Benzimidazole served as a visualization co-ion for indirect detection at 254 nm. The separation of some alkali and alkaline earth metal and ammonium cations in rain, tap and mineral water samples illustrates the application potential of the proposed approach.

Speciation of aluminum using capillary zone electrophoresis with indirect UV detection

Capillary zone electrophoresis was evaluated as a method for the speciation of aluminum in aqueous solution using indirect UV absorption detection. Flu

The determination of lactate, acetate, chloride and phosphate in an intravenous solution by non-suppressed ion chromatography

The determination of acetate, lactate, chloride and phosphate in an intravenous solution is investigated using non-suppressed ion chromatography with indirect UV absorption detection. When phthalate eluents are used with low capacity anion-exchange columns, the above solute species cannot be resolved unless acetonitrile is added to the eluent. Optimum results are obtained with 0.3 mM phthalate (pH 6.0) containing 30% acetonitrile as eluent. The improved resolution with this eluent is attributed to the existence of a partial reversed-phase retention mechanism operating on the unfunctionalised portions of the styrene-divinylbenzene polymeric ion-exchange material.

Conductivity and indirect UV absorption detection of inorganic cations in non-suppressed ion chromatography using aromatic bases as eluents : I. Principles of operation

An equation is developed to describe the retention behaviour of inorganic cations in non-suppressed ion chromatography and it is shown that a linear relationship exists between the logarithm of the concentration of the eluting species in the eluent and the logarithm of the solute capacity factor. Moreover, the negative slope of this relationship is equal to the ratio of the charges on the solute and eluent ions, provided only one eluent species is responsible for elution. The conductivity change accompanying elution of a solute cation can be positive or negative, depending on whether the eluent cation has a high or low limiting equivalent ionic conductance. When an aqueous solution of an aromatic base is used as eluent, the direction of the conductivity signal when a solute cation elutes is dependent on the relative eluting roles of hydrogen ion and the protonated base cation. When both of these species contribute to the elution of a solute, detection sensitivity is decreased in comparison to the situation where only one species is responsible for elution. An equation for the conductivity change occurring on elution of a solute cation is presented. Indirect UV absorption detection is also possible when aromatic bases are used as eluents, provided the role of hydrogen ion as an eluting species is minimised. This can be achieved using an eluent comprising an aqueous solution of an aromatic base with p K a value in excess of 7.5, and by maintaining the eluent pH in the neutral to alkaline region. Benzylamine at pH 7.15 is shown to be a suitable eluent for indirect UV absorption and conductivity detection and using this eluent, detection limits in the low parts per billion range (for an injection volume of 10 μl) were obtained for lithium, sodium, ammonium and potassium, with both detection methods.

The occurrence and origin of system peaks in non-suppressed ion chromatography of inorganic anions with indirect ultraviolet absorption detection

Chromatograms obtained for inorganic anions using non-suppressed ion chromatography with indirect UV absorption detection show two extraneous peaks; a non-retained peak (called the injection peak) and a late eluting peak (called the system peak). Experimental studies are reported which show that the system peak is governed by the sample concentration, the injected volume of sample, the nature of the sample ion, the sample pH and the eluent concentration and pH. A mechanism for the generation of injection and system peaks is presented which proposes that the injection peak is due to eluent dilution effects combined with the displacement of adsorbed eluent anions by the injected sample. The system peak is attributed to adsorption and desorption of neutral eluent molecules from the column surface which are retained by a reversed-phase mechanism on the unfunctionalised portions of the anion-exchange column. Evidence is presented to support this proposal. System peaks may be eliminated using an eluent pH such that no neutral eluent molecules are present.