Assay of naproxen in rat serum by high-performance thin-layer chromatography/densitometry

A simple, sensitive, and reproducible high-performance thin-layer chromatographic method using densitometry is presented for the determination of naproxen in rat serum. Only 0.1 mL serum was used for extraction. Separations were performed on 10 x 10 cm plates coated with silica gel, with toluene-ethyl acetate-acetic acid (82 + 15 + 3) as the mobile phase. Benzophenone was used as the internal standard. Quantification was performed by densitometry at 260 nm. The response response for naproxen was linear (r = 0.992) over the range 2-100 mg/L. Method validation demonstrated good recoveries (92-96%), sensitivity (limit of quantitation, 1 mg/L), repeatability of sample application (4%), repeatability of the method (8%), and intermediate precision (5%). The procedure was applied to the quantitation of naproxen in rat serum.     

Assay for dopamine beta-hydroxylase in human plasma and rat serum by high-performance liquid chromatography with fluorimetric detection

A highly sensitive assay for dopamine beta-hydroxylase activity in human plasma and rat serum by high-performance liquid chromatography with fluorimetric detection is described. Norepinephrine, formed enzymatically from the substrate dopamine, and epinephrine (internal standard), after clean-up with a cation-exchange cartridge (Toyopak IC-SP M), are converted into the corresponding fluorescent compounds by reaction with 1,2-diphenylethylenediamine. The derivatives are separated by reversed-phase chromatography on TSK gel ODS-80TM. The detection limit for norepinephrine formed enzymatically is 5 pmol per assay tube.     

Assay of human transthyretin-bound holo-retinol-binding protein with reversed-phase high-performance liquid chromatography

We describe a reversed-phase high-performance liquid chromatographic method for the determination of vitamin A-transporting (holo) transthyretin-bound (TTR) retinol-binding protein (RBP) concentrations in serum or plasma. Holo-TTR-RBP and free retinol derived primarily from free RBP are consistently observed with this chromatographic method. Holo-TTR-RBP concentrations determined by this method are highly correlated to holo-TTR-RBP concentrations measured by chromatography. This method has the advantage of using less expensive columns and having peak areas which are more proportional to their true concentrations in plasma, as determined by comparison to purified protein spectrophotometry and radial immunodiffusion. The percentage of RBP circulating as holo-TTR-RBP decreased significantly as the total concentration of RBP or retinol increased. Because purified holo-TTR-RBP did not dissociate under these chromatographic conditions, this suggests that more vitamin A circulates as holo-free RBP or free retinol in the blood of people with high serum RBP.     

Superiority of thin-layer chromatography over high-performance liquid chromatography in enantioseparation

JPC – Journal of Planar Chromatography – Modern TLC - The scientific development in the area of enantioseparation during the last few decades has centered on the production of new...     

Quantitative determination of miconzole in human serum by high-performance liquid chromatography

Summary A high-performance liqid chromatographic method is reported for the measurement of miconazole in systemic, fungal infectious patients. Pharmacokinetic data are presented for a single patient receiving miconazole therapy. Sample preparation involves protein precipitation by acetonitrile (1:1, vol/vol). Analyses are carried out on a reversed-phae chromatographic system using octadecylsilane stationary phase: a mobile phase consisting of 0.05 M acetate buffer (pH 7.4)acetonitrile (20:80, vol/vol) is used to elaute miconazole is quantified on the basis of ultraviolet absorption at 220 nm. The precision of the method ranged from 3.21% at 0.5 mg/L to 0.85% at 2.0 mg/L. The limit of quantification was established as 0.1 mg/L. Interference from other drugs that are co-adimistered such as amphotericin B, 5-fluorocytosine of ketoconazole and most other comonly encountered drugs was not observed.     

Determination of myoglobin in human serum by high-performance liquid chromatography with chemiluminescence detection

A highly sensitive method for the determination of myoglobin in serum is described, based on high-performance size-exclusion chromatography with chemiluminescence detection. Serum proteins are separated according to their molecular masses on columns packed with TSK-SW gel and those containing haem are detected selectively by post-column chemiluminescence reaction with luminol using a conventional fluorimetric detector. The method is rapid (30 min) and sufficiently sensitive for the diagnosis of myocardial infarction. The minimum detectable myoglobin concentration is 10 ng/ml.     

Rapid analysis of human serum albumin by high-performance liquid chromatography.

High-performance liquid chromatographic analysis of human serum albumin, using a column containing quaternized dimethyl-aminomethylstyrene-ethylene glycol dimethacrylate, was performed by isocratic elution. This column afforded resolution of albumin components, such as human mercaptalbumin and human nonmercaptalbumin. The method is an alternative to gradient chromatography, and allows rapid determination of the albumin components.     

Assay for tyrosine hydroxylase by high-performance liquid chromatography with fluorescence detection

A sensitive assay method for tyrosine hydroxylase in rat brain and adrenal medulla by high-performance liquid chromatography with fluorescence detection is described. L-DOPA formed enzymatically from the substrate L-tyrosine and alpha-methyldopa (internal standard), after clean-up with small cartridges of an activated alumina and a cation exchanger, Toyopak IC-SP M, are converted into the corresponding fluorescent compounds by reaction with 1,2-diphenylethylenediamine. The derivatives are separated by reversed-phase chromatography on TSK gel ODS-120T. The detection limit for L-DOPA formed enzymatically is 2 pmol per assay tube.     

Determination of ofloxacin in human serum by high-performance liquid chromatography with column switching.

The chromatographic behaviour of ofloxacin on various sorbents, including ODS, C8, C1, nitril, phenyl and tert,-butyl, as stationary phases was investigated and a high-performance liquid chromatography (HPLC) assay was developed for the determination of ofloxacin in serum. The serum samples were directly introduced onto an HPLC column after filtering through a Morcut II membrane filter to remove proteins. The filtrate was concentrated on a pre-column using a phenyl stationary phase and was then introduced to an analytical column with an ODS stationary phase by column switching. Ofloxacin and enoxacin as an internal standard were detected by ultraviolet absorbance at 300 nm. Determination was possible for ofloxacin over the concentration range 50-2000 ng/ml; the limit of detection was 20 ng/ml. The recovery of ofloxacin added to serum was 88.8-101.7% with a coefficient of variation of less than 5.2%. This method is applicable to pharmacokinetic studies of patients after treatment with ofloxacin.     

Simultaneous assay of olanzapine and fluoxetine in tablets by column high-performance liquid chromatography and high-performance thin-layer chromatography

This paper describes validated high-performance liquid chromatographic (LC) and high-performance thin-layer chromatographic (TLC) methods for the simultaneous estimation of olanzapine and fluoxetine in pure powder and tablet formulations. The LC separation was achieved on a Lichrospher 100 RP-180, C18 column (250 mm, 4.0 mm id, 5 microm) using 0.05 M potassium dihydrogen phosphate buffer (pH 5.6 adjusted with o-phosphoric acid)-acetonitrile (50 + 50, v/v) as the mobile phase at a flow rate of 1 mL/min and ambient temperature. The TLC separation was achieved on aluminum sheets coated with silica gel 60F254 using methanol-toluene (40 + 20, v/v) as the mobile phase. Quantitation was achieved by measuring ultraviolet absorption at 233 nm over the concentration range of 10-70 and 40-280 microg/mL with mean recovery of 99.54 +/- 0.89 and 99.73 +/- 0.58% for olanzapine and fluoxetine, respectively, by the LC method. Quantitation was achieved by measuring ultraviolet absorption at 233 nm over the concentration range of 100-800 and 400-3200 ng/spot with mean recovery of 101.53 +/- 0.06 and 101.45 +/- 0.35% for olanzapine and fluoxetine, respectively, by the TLC method with densitometry. These methods are simple, precise, and sensitive, and they are applicable for simultaneous determination of olanzapine and fluoxetine in tablet formulations.