Radoslav Halko

Identification of Natural Dyestuff in Archeological Coptic Textiles by HPLC with Fluorescence Detection

Abstract High performance liquid chromatography with UV–Vis detection and post-column derivatization with fluorimetric detection was used for analysis of dyes used in Coptic textiles. To obtain the best separation of analyzed compounds composition of mobile phase was optimized. The best results were obtained for 25 mM phosphoric buffer at pH 2.5 with gradient change of methanol concentration (Table 1). As post-column reagents used to enhance fluorescence signal of analyzed compounds, solutions of Al(III), Ga(III), In(III), and Zn(II) were used. The highest increase of fluorescence signal was observed for Ga(III) and 10 mM solution of this ion was used for investigation of plant extracts and extracts from Coptic textiles.

Some theoretical and practical aspects in the separation of humic substances by combined liquid chromatography methods

Permanent need to understand nature, structure and properties of humic substances influences also separation methods that are in a wide scope used for fractionation, characterization and analysis of humic substances (HS). At the first glance techniques based on size-exclusion phenomena are the most useful and utilized for relating elution data to the molecular mass distribution of HS, however, with some limitations and exceptions, respectively, in the structural investigation of HS. The second most abundant separation mechanism is reversed-phase based on weak hydrophobic interactions beneficially combined with the step gradients inducing distinct features in rather featureless analytical signal of HS. Relatively great effort is invested to the developments of immobilized-metal affinity chromatography mimicking chelate-forming properties of HS as ligands in the environment. Surprisingly, relatively less attention is given to the ion-ion interactions based ion-exchange chromatography of HS. Chromatographic separation methods play also an important role in the examination of interactions of HS with pesticides. They allow us to determine binding constants and the other data necessary to predict the mobility of chemical pollutants in the environment. HS is frequently adversely acting in analytical procedures as interfering substance, so more detailed information is desired on manifestation of its numerous properties in analytical procedures. The article topic is covered by the review emphasizing advances in the field done in the period of last 10 years from 2000 till 2010.

Flow-injection sample preconcentration for ion-pair chromatography of trace metals in waters

Selected trace transition metal ions have been determined in an FIA/HPLC hyphenated system using on-line preconcentration on cellulose functionalised sorbent Cellex P. For HPLC separation ion-pair chromatography was employed with spectrophotometric detection at 510 nm using post-column derivatisation with PAR. Favourable kinetic conditions of sorption and elution as well as optimisation of hyphenated system allowed to obtain detection limits at sub-microgL(-1) level at 25 min preconcentration time. The developed method was employed for determination of Co(II), Ni(II), Cd(II) and Mn(II) in river water with reasonable agreement of obtained results with electrothermal AAS determination.

Reversed phase liquid chromatography trace analysis of pesticides in soil by on‐column sample pumping large volume injection and UV detection

The idea of utilization of one hydraulic line of a common commercial HPLC pump for direct on-column sample pumping injection of large sample volumes, 20 mL, was further investigated with the aim to develop multicomponent pesticides trace residues HPLC method in gram soil samples. Target pesticides group involve asulam, atrazine, 2,4-D, PCA, propazine, simazine, 4-chloro-2-methylphenoxyacetic acid, 2-(4-chloro-2-tolyloxy) propionic acid, chlortoluron, metoxuron, epoxiconazole. The results proved the applicability of this approach in experiments with mixtures of analytes at low ng/mL levels. Analysis of 20 mL of soil leachates and extracts of fortified soil samples containing these pesticides at the 10-50 ng/g level (in dry soil) revealed good figures of merit, also in the presence of large excess of humics. LODs achieved by detection at 220 nm evaluated from calibration runs of spiked soil extracts by Hubaux et al. method ranged from 5-12 ng per injected volume. For 20 mL large volume injection it represents 0.25-0.6 ng/mL of diluted soil extract, or 2.5-6 ng/mL of crude extract, or 6-5 ng/g dry soil. Recoveries of pesticides at concentration levels approaching half of maximum allowable concentration of pesticides in soil (100 ng/g) ranged from 85 to 98% with acceptable reproducibility, except asulam and metoxuron.

Study of high-performance liquid chromatographic separation of selected herbicides by hydro-methanolic and micellar liquid chromatography using Genapol X-080 non-ionic surfactant as mobile phase constituent

Abstract Chromatographic behaviour of six selected herbicides (chlortoluron, metoxuron, chloridazon, simazine, propazine and atrazine) was studied by reversed-phase (RP) high-performance liquid chromatography (HPLC) containing Genapol X-080 non-ionic surfactant as methanol/water mobile phase constituent. The concentration of methanol was changed from 50 to 0% (v/v) for constant 2% (v/v) concentration of the surfactant. The surfactant concentration in purely aqueous micellar mobile phase varied from 1 to 5% (v/v) what is approximately 360–1800 times above the CMC. Within this concentration range Genapol X-080 proves concentration dependent selectivity changes for chlortoluron/atrazine critical pair not occurred in hydro-methanolic mobile phases. Further studies revealed that this chromatographic system offers high compatibility with cloud-point extraction environmental sample pretreatment approaches using Genapol X-080 for the purpose, too.

Separation of mercury(II), methylmercury and phenylmercury by micellar high-performance liquid chromatography on short columns

Three environmentally and agrochemicaly important mercury species: methylmercury, phenylmercury and mercury(II) are separated within 4 minutes as bromocomplexes by micellar liquid chromatography using very short reversed-phase (RP) C18 columns (up to 30 mm). The micellar mobile phase containig 0.05M cetyltrimethylammonium bromide (CTMA+Br−), 1% (v/v) 2-propanol, 0.001M cyclohexylenediaminetetraacetic acid (DCTA) and sulfuric acid (pH 2) showed good selectivity in mixed reversed-phase and anion-exchange mode. The above mentioned separation order in which organomercurials are cluted far behind the void volume of the column, but before the mercury(II) peak is advantageous in all instances where mercury(II) is present in real samaples in great excess. Environmental and agrochemical samples contain humic material which does not interfere in this particular system. The low cost photometric detection at 500 nm after post-column derivatization by CTMA+Br− micellized dithizone is almost free from interferences and enables detection limits at the 1–3 ng level (e.g., 0.1 ppm Hg) for 20 μl samples.

Determination of Copper in Human Urine by Cloud Point Extraction and Flame Atomic Absorption Spectrometry

ABSTRACT This work reports novel cloud point extraction and preconcentration for the determination of copper in human urine by flame atomic absorption spectrometry. The method is based on the selective reaction of 2,9-dimethyl-1,10-phenanthroline (neocuproine) with Cu(I) to produce highly hydrophobic chelate that is extracted and preconcentrated using polyethylene glycol tert-octylphenyl ether (Triton X-114) as nonionic surfactant. Before the derivatization reaction, Cu(II) was reduced to Cu(I) by ascorbic acid. The effect of pH, concentration of the chelating agent and surfactant, equilibration temperature, and incubation time were investigated. Under optimum conditions, preconcentration of copper from 10 mL of urine sample in the presence of 0.10% (w/v) Triton X-114 and 0.1 mmol/L neocuproine at pH 5.9 permitted a limit of detection of 0.01 µg/L for copper in urine. The relative standard deviation was less than 3.5% for copper concentrations from 0.5 to 100.0 µg/L following cloud point extraction. The correlation coefficients of the calibration curves were higher than 0.999. The effects of matrix ions (cations and anions) on the recovery of copper were studied systematically. The use of flame atomic absorption spectrometry with certified reference material for the determination of copper in human urine yielded satisfactory results.

Fast ultra-high-performance liquid chromatography with diode array and mass spectrometry method for determination of tadalafil drug substance and its impurities

Tadalafil is used for the treatment of erectile dysfunction. Its related patents expired in 2016, and so related generic drug production is predicted to be increased. This work is focused on developing a fast ultra-high-performance liquid chromatography with diode array detection and/or mass spectrometry detection for the separation and determination of tadalafil and its impurities in pharmaceutical samples. A modern reversed-phase stationary phase with sub-2 μm particle size, Zorbax StableBond Rapid Resolution High Definition with octylsilane chemically bonded phase to totally porous silica particles, was used for the solving this problem. Column temperature was set at 40 ± 0.1°C. A mobile phase consisting of acetonitrile and aqueous solution of 0.1% (v/v) trifluoroacetic acid for diode array detection detection and 0.05% (v/v) formic acid, both running at a flow rate of 0.62 mL/min, were used to achieve the required separation of all components within a 5 min run. The limit of detection was 3.5 μg/L and the limit of quantification was 10.0 μg/L for the method for both UV and MS detectors. Accurate mass spectra of tadalafils related impurities are shown for advanced confirmation. The method is directly transferable to routine analysis of tadalafil in pharmaceutical and control laboratories.