The binding of tannin fraction to protein fractions isolated from broad bean seeds was studied by precipitating potential and fluorescence quenching methods. The tannin fraction with high proanthocyanidins content was isolated from broad bean coats. Storage proteins of broad bean, 11S, 7S, and 2S, were isolated from broad bean cotyledons and purified. Gelatin, BSA, as well as broad bean 2S and 7S protein fractions exhibited similar shape of curves illustrating the effect of pH on protein precipitation and were precipitated by broad bean tannin fraction over the wide range of pH. For pea proteins isolate and 11S fraction of broad bean proteins, quite different effect of pH was observed. The relationships between the amount of proteinpolyphenols complex precipitated and the content of tannin fraction were linear and characterized by high squared correlation coefficients ranging from 0.9613 to 0.9938. Among the protein fractions isolated from broad bean seeds, the highest precipitating potential was noted for 11S fraction, followed by 7S and 2S fractions and amounted to 1.63, 1.34, and 0.87, respectively. To characterize the association of proteins with phenolic compounds yielding formation of soluble complexes, fluorescence spectroscopy was applied. Similarly, the most extensive fluorescence quenching was observed in the case of 11S protein fraction.
liquid chromatography with a diode array detection (SE-HPLC-DAD) in analysis of lignan macromolecule (LM) and phenolic compounds liberated from LM after chemical and enzymatic hydrolysis. MATERIAL AND METHODS MaterialGround, partially defatted flaxseeds were purchased from the "Ekoprodukt" company (Częstochowa, Poland). Preparation of crude extractThe material was defatted with hexane and then phenolic compounds were extracted using a dioxane:ethanol (1:1; v:v) mixture [Johnsson et al., 2000]. The extraction was carried out for 16 h, at 60°C with continuous shaking in a water bath. Then, solvent was evaporated using Büchi Rotavapor R-200 at 40°C. Amberlite column chromatographyThe flaxseed phenolic compounds extract was purified using column chromatography on Amberlite XAD-16 (34 mm i.d.; 170 mm length) [Srivastava et al., 2010]. A 5-g portion of the extract was suspended in 5 mL of distilled water and loaded on the column. Firstly, water soluble compounds, mainly sugars, were eluted using distilled water and discarded, then solvent was changed over to methanol which eluted phenolic compounds. The solvent was removed using Büchi Rotavapor R-200. Base hydrolysisThe purified extract was subjected to base hydrolysis. Briefly, the purified extract was suspended in 0.3 mol/L NaOH, SE-HPLC-DAD Analysis of Flaxseed Lignan Macromolecule and its HydrolysatesAgnieszka Kosińska, Anna Urbalewicz, Kamila Penkacik, Magdalena Karamać, Ryszard Amarowicz* Sciences, Olsztyn, ul. Tuwima 10, Poland Key words: flaxseed, phenolics, lignans, SE-HPLC-DAD, hydrolysis A lignan macromolecule (LM) was extracted from defatted flaxseeds using an ethanol-dioxan system (1:1, v/v) and purified using Amberlite column chromatography with water and methanol as mobile phases. The LM was subjected to chemical hydrolysis (base, acid, base & acid), as well as to enzymatic processing using pepsin, pancreatin, cellulase, and β-glucuronidase. Division of Food Sciences, Institute of Animal Reproduction and Food Research of the Polish Academy ofThe study revealed that lignan macromolecule in flaxseed was not homogenous. The chemical hydrolysis as well as enzymatic treatment using β-glucuronidase and cellulase released low molecular phenolic compounds from the lignan macromolecule. The liberation of secoisolariciresinol (SECO) and free phenolic acids (p-coumaric and ferulic acids) from flaxseed lignan macromolecule as a result of the base and acid hydrolyses was noted. The application of pepsin and pancreatin did not change the composition of the lignan macromolecule.
SummaryThe influence of copper ions on the regeneration of carrot (Daucus carota L.) androgenic embryos, accumulation of copper in rosettes, soluble esterbound phenolic acids and some parameters of oxidative stress were investigated. Two carrots: cv. Feria and 1014 breeding line were subjected to 1 μM, 10 μM and 100 μM Cu stress for 16 and 24 weeks. Under this stress, better growth, lower lipid peroxidation (TBARS level) and higher phenolic acid contents were observed in the cv. Feria. The rosettes of 1014 line accumulated less copper and produced smaller amount of TBARS after 24 weeks of incubation than after 16 weeks. Chlorogenic and caffeic acids were the main phenolic acids in both cultures. In the Feria rosettes the application of 10 μM Cu caused relatively high level of chlorogenic acid combined with low accumulation of copper in the tissues and unchanged levels of TBARS after both 16 and 24 weeks of incubation. On the other hand, despite the dose-dependent decline of chlorogenic acid in the rosettes of 1014 line, decrease in TBARS content was also observed after 24 weeks. The obtained results might suggest that the Feria carrot culture was able to develop more effective protection system/strategy against Cu excess in comparison to the 1014 line.
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