Vendula Roblová

Monitoring of HPLC profiles of selected polyphenolic compounds in sea buckthorn (Hippophaë rhamnoides L.) plant parts during annual growth cycle and estimation of their antioxidant potential

AbstractPresented work summarizes the data about polyphenolic profiles in various plant parts (leaves, shoots, berries) of sea buckthorn (Hippophaë rhamnoides L.) during the annual growth cycle. A reversed-phase high performance liquid chromatography method (RP-HPLC) coupled with diode-array detection (DAD) was optimized for determination of catechin, epicatechin, gallic acid, p-coumaric acid, caffeic acid, ferulic acid, rutin (quercetin 3-rutinoside) and quercitrin (quercetin 3-rhamnoside). The content of these polyphenolic compounds was monitored in extracts of sea buckthorn plant samples from April to October. The total antioxidant activity was determined using scavenging of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonate) cation radical (ABTS·+) and 1,1-diphenyl-2-picrylhydrazyl radical (DPPH·). The total content of polyphenols was estimated by conventional spectrophotometric method using Folin-Ciocalteu reagent. The monitoring of temporal changes of selected polyphenolic compounds by RP-HPLC showed that catechin, epicatechin and gallic acid were the most abundant analytes in annual green shoots and leaves, and their content varied significantly during the studied period.

Determination of sterols using liquid chromatography with off-line surface-assisted laser desorption/ionization mass spectrometry

A new method, reversed phase liquid chromatography with off-line surface-assisted laser desorption/ionization mass spectrometry (RPLC-SALDI MS) for the determination of brassicasterol (BR), cholesterol (CH), stigmasterol (ST), campesterol (CA) and β-sitosterol (SI) in oil samples has been developed. The sample preparation consisted of alkaline saponification followed by extraction of the unsaponificable fraction with diethyl ether. The recovery of the sterols ranged from 91 to 95% with RSD less than 4%. Separation of the five major sterols on a C18 column using methanol-water gradient was achieved in about 10min. An on-line UV detector was employed for the initial sterol detection prior to effluent deposition using a laboratory-built spotter with 1:73 splitter. Off-line SALDI MS was then applied for mass determination/identification and quantification of the separated sterols. Ionization of the nonpolar analytes was achieved by silver ion cationization with silver nanoparticles used as the SALDI matrix providing limits of detection 12, 6 and 11fmol for CH, ST and SI, respectively. Because of the incorporated splitter, the effective limits of detection of the RPLC-SALDI MS analysis were 4, 3 and 4pmol (or 0.08, 0.06 and 0.08μg/mL) for CH, ST and SI, respectively. For quantification, 6-ketocholestanol (KE) was used as the internal standard. The method has been applied for the identification and quantification of sterols in olive, linseed and sunflower oil samples. The described off-line coupling of RPLC to SALDI MS represents an alternative to GC-MS for analysis of nonpolar compounds.

Capillary electrophoresis fingerprinting and spectrophotometric determination of antioxidant potential for classification of Mentha products

In this work aqueous infusions from ten Mentha herbal samples (four different Mentha species and six hybrids of Mentha x piperita) and 20 different peppermint teas were screened by capillary electrophoresis with UV detection. The fingerprint separation was accomplished in a 25 mM borate background electrolyte with 10% methanol at pH 9.3. The total polyphenolic content in the extracts was determined spectrophotometrically at 765 nm by a Folin-Ciocalteu phenol assay. Total antioxidant activity was determined by scavenging of 2,2-diphenyl-1-picrylhydrazyl radical at 515 nm. The peak areas of 12 dominant peaks from CE analysis, present in all samples, and the value of total polyphenolic content and total antioxidant activity obtained by spectrophotometry was combined into a single data matrix and principal component analysis was applied. The obtained principal component analysis model resulted in distinct clusters of Mentha and peppermint tea samples distinguishing the samples according to their potential protective antioxidant effect. Principal component analysis, using a non-targeted approach with no need for compound identification, was found as a new promising tool for the screening of herbal tea products.