Chromatographic separation of 10 triterpenoids (α-amyrin, β-amyrin, δ-amyrin, lupeol, lupenon, lupeol acetate, cycloartenol, cycloartenol acetate, ursolic acid, oleanolic acid) and 2 sterols (stigmasterol and β-sitosterol) was studied. The chromatographic techniques included silica gel and reversed-phase (C18 RP) thin-layer chromatography (TLC) and C18 RP high-performance liquid chromatography (HPLC) using UV and mass spectrometric (MS) detection with atmospheric pressure chemical ionization (APCI). The TLC separation of the isomeric triterpenols lupeol, α-amyrin, β-amyrin, and cycloartenol was achieved for the first time using C18 RP-HPTLC plates. Cycloartenol could be separated from related compounds only on C18 RP-TLC but not on the C18 RP-HPLC. δ-Amyrin isolated from extracted cuticular waxes of tomato fruit could be separated from other amyrins only by HPLC. Tandem mass spectrometry allowed discrimination between the isomers lupeol, α-amyrin, β-amyrin, δ-amyrin, cycloartenol and between lupeol acetate and cycloartenol acetate. The combination of 3 TLC methods and 2 HPLC methods enables qualitative determination of all 12 compounds and proves to be useful for the analysis of these compounds in plant extracts. It is recommended that TLC screening on silica gel and C18 RP are performed before HPLC analysis. CITATIONS: 26 (source: Scopus)
COBISS.SI-ID: 4217626
Nine shikonin pigments: shikonin (S), acetylshikonin (AS), propionylshikonin (PS), isobutyrylshikonin (IBS), tiglylshikonin (TS), 3,3-dimethylacrylshikonin (DAS), angelylshikonin (ANS), 2-methyl-n-butyrylshikonin (MBS) and isovalerylshikonin (IVS) were identified in the root epidermis of Echium italicum L. for the first time. A new thin-layer chromatographic method for the separation of enantiomers alkannin and shikonin proved only shikonin after saponification of the root extract, and was afterwards esterified with the corresponding acyl chloride to acquire seven standard compounds (all except ANS). The developed isocratic high-peformance liquid chromatographic (HPLC) methods with VIS and mass spectrometry detection, allowed for the first time simultaneous separation of all nine compounds with similar structures including positional and geometric isomers in a short time. Structures of the main five compounds (AS, IBS, ANS, MBS, IVS) isolated from the extract by a new semi-preparative HPLC on C18 have additionally been confirmed by 1H and 13C nuclear magnetic resonance spectra, which were reported for AS and MBS for the first time. CITATIONS: 13 (source: Scopus)
COBISS.SI-ID: 4134682
We report the optimization of a sensitive, selective and robust derivatization method using 4-dimethylaminocinnamaldehyde (DMACA) for densitometric determination of (+)-catechin and (-)-epicatechin. The separation of these compounds was achieved by thin-layer chromatography on cellulose plates developed with water. With DMACA in HCl, both compounds gave blue bands, while under the same conditions, vanillin produced a fast fading red coloration of bands. Quantitation at 655 nm showed that for both compounds the calibration curve was linear from 2 to 12 ng and polynomial from 2 to 30 ng, and the repeatability of chromatography of 20 ng was 3.5% (RSD, n=6). The visible limit of detection of both standards was 1 ng, but the densitometric limit of detection was lower (0.2 ng). The optimized DMACA reagent is superior to the more frequently used vanillin reagent and is applicable also for determination of mixtures containing other catechins ((-)-catechin, (-)-epicatechin gallate, (-)-epigallocatechin gallate, procyanidin A2, procyanidin B1 and procyanidin B2). CITATIONS: 14 (source: Scopus)
COBISS.SI-ID: 4134938
The main problem in the densitometric determination of carotenoids is their rapid degradation during and immediately after chromatography, respectively. In this study, we show that 15 ng of lutein, lycopene and β-carotene standards applied on C-18 RP high-performance thin-layer chromatography (HPTLC) plates pre-developed with dichloromethane-methanol 1:1 (v/v) remained stable for 1 h after the development of chromatogram using methanol-acetone 1:1 (v/v) with 0.1% of 2-tert-butylhydroquinone (TBHQ), which is a substantial improvement of their stability. An HPTLC quantification procedure for free lutein, with densitometry at 450 nm based on the developed method described above, was established and validated. Repeatabilities of the chromatography expressed by the relative standard deviation from 6 applications of lutein standard at 5, 15 and 25 ng were 3.41, 1.33 and 1.65%, respectively. The best fit calibration curve from 5 ng to 30 ng of lutein was polynomial. Limit of detection (1.5 ng) and limit of quantification (5 ng) were the best achieved so far. With these chromatographic conditions dietary carotenoids lutein esters, lycopene, free lutein and β-carotene from food supplements were also well separated and were identified by visible absorption spectra scanned in situ and by mass spectra. Some additional developing solvents with the same type of chromatographic layer are proposed for the fast separation of lutein esters from free lutein in food supplements. CITATIONS: 5 (source: Scopus)
COBISS.SI-ID: 4915226
Shikonin and its ester derivatives belong to a group of secondary metabolites with a wide array of beneficial effects on human health. However, shikonin is principally used in oil-based preparations due to the low solubility of the pigment in aqueous media, and the positive properties of shikonin are not exploited to their full potential. Such low aqueous solubility often results in poor bioavailability, makes shikonin undesirable for oral administration, and restricts its broadened use in the food and pharmaceutical industries. The purpose of this study was to enhance the aqueous solubility of shikonin by the addition of β-lactoglobulin and to characterize the macromolecule-ligand binding interaction by means of spectrophotometry, spectrofluorometry, high-performance liquid chromatography, and mass spectrometry. In the presence of β-lactoglobulin the solubility of shikonin is increased up to 181-fold. One shikonin molecule binds covalently to β-lactoglobulin via Cys121, whereas the remaining pigment molecules most probably bind to the protein via noncovalent interactions. CITATIONS: 1 (source: Scopus)
COBISS.SI-ID: 5114394