A fast method using a new hydrophilic–lipophilic balanced sorbent in combination with ultra-high performance liquid chromatography for quantification of significant bioactive metabolites in wines

This manuscript describes the development and validation of an ultra-fast, efficient, and high throughput analytical method based on ultra-high performance liquid chromatography (UHPLC) equipped with a photodiode array (PDA) detection system, for the simultaneous analysis of fifteen bioactive metabolites: gallic acid, protocatechuic acid, (−)-catechin, gentisic acid, (−)-epicatechin, syringic acid, p-coumaric acid, ferulic acid, m-coumaric acid, rutin, trans-resveratrol, myricetin, quercetin, cinnamic acid and kaempferol, in wines. A 50-mm column packed with 1.7-μm particles operating at elevated pressure (UHPLC strategy) was selected to attain ultra-fast analysis and highly efficient separations. In order to reduce the complexity of wine extract and improve the recovery efficiency, a reverse-phase solid-phase extraction (SPE) procedure using as sorbent a new macroporous copolymer made from a balanced ratio of two monomers, the lipophilic divinylbenzene and the hydrophilic N-vinylpyrrolidone (Oasis™ HLB), was performed prior to UHPLC–PDA analysis. The calibration curves of bioactive metabolites showed good linearity within the established range. Limits of detection (LOD) and quantification (LOQ) ranged from 0.006 μg mL−1 to 0.58 μg mL−1, and from 0.019 μg mL−1 to 1.94 μg mL−1, for gallic and gentisic acids, respectively. The average recoveries ± SD for the three levels of concentration tested (n = 9) in red and white wines were, respectively, 89 ± 3% and 90 ± 2%. The repeatability expressed as relative standard deviation (RSD) was below 10% for all the metabolites assayed. The validated method was then applied to red and white wines from different geographical origins (Azores, Canary and Madeira Islands). The most abundant component in the analysed red wines was (−)-epicatechin followed by (−)-catechin and rutin, whereas in white wines syringic and p-coumaric acids were found the major phenolic metabolites. The method was completely validated, providing a sensitive analysis for bioactive phenolic metabolites detection and showing satisfactory data for all the parameters tested. Moreover, was revealed as an ultra-fast approach allowing the separation of the fifteen bioactive metabolites investigated with high resolution power within 5 min.

Determination of gatifloxacin in bulk and tablet preparations by high-performance liquid chromatography

A sensitive, precise, and specific high-performance liquid chromatographic (HPLC) method was developed for the assay of gatifloxacin (GATX) in raw material and tablets. The method validation parameters yielded good results and included the range, linearity, precision, accuracy, specificity, and recovery. It was also found that the excipients in the commercial tablet preparation did not interfere with the assay. The HPLC separation was carried out by reversed-phase chromatography on a C18 absorbosphere column (250 x 4.6 mm id, 5 pm particle size) with a mobile phase composed of acetic acid 50/o--acetonitrile-methanol (70 + 15 + 15, v/v/v) pumped isocratically at a flow rate of 1.0 mL/min. The effluent was monitored at 287 nm. The calibration graph for GATX was linear from 4.0 to 14.0 mu g/mL. The interday and intraday precisions (relative standard deviation) were less than 1.05%.

High-performance liquid chromatographic determination of amfepramone hydrochloride, mazindol, and diazepam in tablets

Simple and rapid procedures were developed for the quantification of amfepramone hydrochloride and diazepam and mazindol and diazepam in tablets using high performance liquid chromatography (HPLC) with UV detection. These techniques provided conditions for the separation of each active ingredient from the complex matrices of the dosage forms by dilution or extraction in methanol. Isocratic reversed phase chromatography was performed using acetonitrile, methanol, and aqueous 0,1% ammonium carbonate (70:10:20, v/v/v) as a mobile phase, Radial-Pak C-18 column (100 x 8 mm id, 4 mu m), a column temperature of 25+/-1 degrees C and detection at 255 nm. The calibration curves were linear over a wide concentration range (100-1000 mu g.mL(-1) to amfepramone hydrochloride and mazindol and 10-100 mu g.mL(-1) to diazepam) with good correlation factors of 0.9978, 0.9956 and 0.9997 for amfepramone hydrochloride, mazindol, and diazepam, respectively.Mean recoveries obtained from the two kinds of samples ranged from 83.2 to 102.5%, with coefficients of variation ranging from 1.0 to 6.1.These results demonstrated the efficiency of the proposed methods, as well as advantages such as simplicity and short duration of analysis.

Identification of oligomers in polyethyleneterephthalate bottles for mineral water and fruit juice - Development and validation of a high-performance liquid chromatographic method for the determination of first series cyclic trimer

Cyclic oligomers were identified in PET bottles used for mineral water and fruit juice using MS and H-1 and C-13 NMR: a first series cyclic trimer, a first series cyclic tetramer, a first series cyclic dimmer and a second series cyclic trimer. An analytical method to determine first series cyclic trimer in these bottles was developed and validated, using HPLC. The first series cyclic trimer levels were 316-462 mg/100 g of PET bottle. (c) 2005 Elsevier B.V. All rights reserved.

Analytical Validation of Quantitative High-Performance Liquid Chromatographic Methods in Pharmaceutical Analysis: A Practical Approach

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Development of a new high-performance liquid chromatographic method for the determination of ceftazidime

A rapid, accurate, and sensitive high-performance liquid chromatographic (HPLC) method was developed and validated for the determination of ceftazidime in pharmaceuticals. The method validation parameters yielded good results and included range, linearity, precision, accuracy, specificity, and recovery. The excipients in the commercial powder for injection did not interfere with the assay. Reversed-phase chromatography was used for the HPLC separation on a Waters C18 (WAT 054275; Milford, MA) column with methanol-water (70 + 30, v/v) as the mobile phase pumped isocratically at a flow rate of 1.0 mL/min. The effluent was monitored at 245 nm. The calibration graph for ceftazidime was linear from 50.0 to 300.0 mu g/mL. The values for interday and intraday precision (relative standard deviation) were < 1 %. The results obtained by the HPLC method were calculated statistically by analysis of variance. We concluded that the HPLC method is satisfactory for the determination of ceftazidime in the raw material and pharmaceuticals.

Performance characteristics of high performance liquid chromatography, first order derivative UV spectrophotometry and bioassay for fluconazole determination in capsules

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

High-performance liquid chromatographic separation and identification of polyphenolic compounds from the infusion of Davilla elliptica St. Hill

The isolation of polyphenolic compounds from an infusion of the Brazilian plant Davilla elliptica (Dilleniaceae), used as tea by virtue of its digestive properties, is described. An improved preparative HPLC method was used in order to isolate pure polyphenols from the complex mixture. Liquid-liquid extraction and solid-phase extraction were employed to minimise the interference of polymeric compounds and to provide an enriched fraction of the compounds of interest. The identification of the isolated compounds was performed using analytical HPLC as well as direct injection electrospray ionisation ion trap tandem mass spectrometry (ESI-IT-MS/MS). The high flavonoid content suggests that D. elliptica may be a promising source of compounds to produce natural phytomedicines. Copyright (C) 2007 John Wiley & Sons, Ltd.

Quantitation of malondialdehyde in gingival crevicular fluid by a high-performance liquid chromatography-based method

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Development and application of methods for determination of residual monomer in dental acrylic resins using high performance liquid chromatography

Two high-performance liquid chromatographic methods for determination of residual monomer in dental acrylic resins are described. Monomers were detected by their UV absorbance at 230 nm, on a Nucleosil((R)) C-18 (5 mu m particle size, 100 angstrom pore size, 15 x 0.46 cm i.d.) column. The separation was performed using acetonitrile-water (55:45 v/v) containing 0.01% triethylamine (TEA) for methyl methacrylate and butyl methacrylate, and acetonitrile-water (60:40 v/v) containing 0.01% TEA for isobutyl methacrylate and 1,6-hexanediol dimethacrylate as mobile phases, at a flow rate of 0.8 mL/min. Good linear relationships were obtained in the concentration range 5.0-80.0 mu g/mL for methyl methacrylate, 10.0-160.0 mu g/mL for butyl methacrylate, 50.0-500.0 mu g/mL for isobutyl methacrylate and 2.5-180.0 mu g/mL for 1,6-hexanediol dimethacrylate. Adequate assay for intra- and inter-day precision and accuracy was observed during the validation process. An extraction procedure to remove residual monomer from the acrylic resins was also established. Residual monomer was obtained from broken specimens of acrylic disks using methanol as extraction solvent for 2 h in an ice-bath. The developed methods and the extraction procedure were applied to dental acrylic resins, tested with or without post-polymerization treatments, and proved to be accurate and precise for the determination of residual monomer content of the materials evaluated. Copyright (c) 2005 John Wiley & Sons, Ltd.

Phenylalanine ammonia-lyase activity in soybean roots extract measured by reverse-phase high performance liquid chromatography

The high performance liquid chromatography (HPLC) technique was applied to measure phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) activity in soybean (Glycine max L. Merril cv. BR16) roots. t-Cinnamate, the catalytic product of the PAL reaction was quantified at 275 nm by isocratic elution with methanol:water through an ODS(M) column. Comparative experiments were carried out with 1.0 mM ferulic acid, an inducer of PAL activity. The results suggest that liquid chromatography is a rapid and sensitive method to analyze PAL activity in non-purified extract.

High-Performance Liquid Chromatographic Quantification of Flavonoids in Eriocaulaceae Species and Their Antimicrobial Activity

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Validated high-performance liquid chromatographic method for the standardisation of Ptychopetalum olacoides Benth., Olacaceae, commercial extracts

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)