Eleonora Lampart-Szczapa

Antioxidant properties of lupin seed products

Antioxidant properties of lupin flours and hulls were examined. Chemical composition (protein, fat, fatty acids, tocopherols and tannin contents) was determined and radiation effects were estimated (1, 5 and 10 kGy). Antioxidant properties of the ethanol lupin extracts were examined using the Rancimat and Oxidograph tests. Alpha-, gamma- and delta-tocopherols were found in the lupin oil. Lupin tannins contents in the flours were a few times higher than in the hulls. Antioxidant activity was found both in the flours and in the hulls. Correlation between the antioxidant properties and the tocopherol and tannin contents (the natural lupin antioxidants) was not found. Increasing doses of irradiation lowered antioxidant effects of lupin extracts; however, the antioxidant activities of some samples were higher. The observed negative changes in the tocopherols contents were effects of the irradiation dose as well as storage time.

Composition of fractional and functional properties of dietary fiber of lupines (L. luteus and L. albus)

In this study the lupine raw materials (flour and hull) of L. luteus var. Juno and L. albus var. Wat were characterized with regard to the dietary fiber content (NDF) and its fractional composition. Functional properties, i.e. water holding capacity (WHC) and cation exchange capacity (CEC) of lupine raw material were determined, with respect to various conditions existing in each part of the human digestive tract (pH-value, time of passage). Experimental products (shortcakes, ginger breads, pancakes, minced meat and dumplings filled with meat) with addition of 5, 10 or 15% of lupine flour or shell were processed and sensory evaluation was performed according to the 5-point scale. The NDF content ranged from 75.7% to 78% in the hull of the Wat and Juno lupine vars. respectively, and 28.8% to 33.4% in the flour. Cellulose was predominant in the hulls NDF while in the flour hemicellulose was major fraction. WHC of samples depended mainly on pH-value and was higher in lupine hulls (up to 5.14 g/g dry matter (d.m.) than in the flours (up to 3.83 g/g d.m.). The CEC of lupine ranged from 0.260 to 0.750 mEq/g d.m. and from 0.330 to 0.870 mEq/g d.m. in flour of the Wat and Juno varieties. The CEC of hull was lower in the Wat var. (0.290 to 0.650 mEq/g d.m.) in comparison with the Juno variety (0.150 to 0.750 mEq/g d.m.) Sensory evaluation showed that 10% addition of flour or hull of lupine to experimental products enables preparation of good quality foodstuffs.

Release of flavonoids from lupin globulin proteins during digestion in a model system.

Lupin seed globulin proteins form complexes with flavonoids, predominantly apigenin C-glycosides. Enzymes typical for the gastrointestinal tract were used to hydrolyze lupin seed globulins. Release of native flavonoids as a result of the proteolysis reaction was observed. Different analytical methods such as size exclusion chromatography, HPLC-MS, and fluorescence spectroscopy (steady-state fluorescence, fluorescence anisotropy, fluorescence lifetimes) were used for a detailed characterization of this phenomenon. Flavonoids liberated from lupin globulin proteins as a result of pancreatin-catalyzed digestion were bound by γ-conglutin resistant to this enzyme. Two possible mechanisms of this interaction may be suggested: hydrogen bonding between oligosaccharide chains of glycoproteins and the sugar moieties of the flavonoid glycosides or electrostatic attraction between positively charged γ-conglutin and flavonoids partially ionized at pH 7.5.

Characterisation of different digestion susceptibility of lupin seed globulins

This study describes in vitro digestion of lupin seed globulins by pancreatin, trypsin and chymotrypsin. Lupin seed globulins turned out to be almost totally susceptible to chymotrypsin digestion. When panceratin or trypsin were used for digestion of lupin seed globulins, γ-conglutin appeared to be resistant to proteolysis. Different fluorescence spectroscopic methods such as fluorescence anisotropy, fluorescence lifetimes and fluorescence quenching measurements were used for detailed characterisation of this phenomenon. A potential reason for γ-conglutin insensitivity to digestion may be related to the fact that lysine, as well as arginine, are positively charged at cell physiological pH. Simultaneously, flavonoids at this pH are partially ionised, which may lead to the occurrence of ionic interactions between these molecules at pH 7.5. The confirmation of this explanation may be the fact that γ-conglutin and vitexin form a static complex, which was observed using fluorescence quenching measurements.

Changes in the protein complex of wheat dough affected by soybean 11 S globulin: Part 2—The interactions of soybean 11 S globulin with gluten proteins

Abstract The interactions between soy 11 S globulin and protein fractions of wheaten doughs were examined using a multistep extraction method followed by molecular sieving and characterisation of the fractions obtained. The soy 11 S globulin preparation was coupled with fluoresceine isothiocyanate to permit tracing of this protein in the complexes formed. In model experiments the average molecular weights of globulin/prolamine complexes were determined, the dispersion of light on the protein molecules being measured. The formation of high molecular weight complexes of soy 11 S globulin and prolamine was found. A shift between albumin/globulin and gluten fractions was also observed, which resulted in an increase in the contents of low molecular weight fractions dispersible in pyrophosphate buffer and acetic acid. Most of the investigations concerning the enrichment of bread with soybean proteins deal with technological problems. The documentation of the chemically induced changes in the dough-protein complex is rather limited and a number of questions are still awaiting answers. The data reported by Jakubczyk et al. (1973), Matthews (1972), Pollock & Geeds (1960 a, b ) do not explain the character of the interactions between soy proteins and gluten systems and give only inadequate information on the changes of molecular structure or aggregation/disaggregation phenomena occurring in the proteins of enriched dough. The results presented in our previous study (Lampart-Szczapa & Jankiewicz, 1982, encouraged us to continue the experiments to explain the role played by soybean 11 S globulin treated as a model soy protein in modifying the gluten matrix of wheat dough. In this paper the effects of soybean 11 S globulin on the fractional distribution of the dough-protein complex and average molecular weights of the fractions are presented.

Structure of γ-conglutin: insight into the quaternary structure of 7S basic globulins from legumes

γ-Conglutin from lupin seeds is an unusual 7S basic globulin protein. It is capable of reducing glycaemia in mammals, but the structural basis of this activity is not known. γ-Conglutin shares a high level of structural homology with glycoside hydrolase inhibitor proteins, although it lacks any kind of inhibitory activity against plant cell-wall degradation enzymes. In addition, γ-conglutin displays a less pronounced structural similarity to pepsin-like aspartic proteases, but it is proteolytically dysfunctional. Only one structural study of a legume 7S basic globulin, that isolated from soybean, has been reported to date. The quaternary assembly of soybean 7S basic globulin (Bg7S) is arranged as a cruciform-shaped tetramer comprised of two superposed dimers. Here, the crystal structure of γ-conglutin isolated from Lupinus angustifolius seeds (LangC) is presented. The polypeptide chain of LangC is post-translationally cleaved into α and β subunits but retains its covalent integrity owing to a disulfide bridge. The protomers of LangC undergo an intricate quaternary assembly, resulting in a ring-like hexamer with noncrystallographic D3 symmetry. The twofold-related dimers are similar to those in Bg7S but their assembly is different as a consequence of mutations in a β-strand that is involved in intermolecular β-sheet formation in γ-conglutin. Structural elucidation of γ-conglutin will help to explain its physiological role, especially in the evolutionary context, and will guide further research into the hypoglycaemic activity of this protein in humans, with potential consequences for novel antidiabetic therapies.

Digestion susceptibility of seed globulins isolated from different lupin species

The aim of this work was to study the processes taking place during the enzymatic hydrolysis of seed globulins isolated from narrow-leaf (var. Zeus and Bojar) and yellow (var. Lord and Parys) lupin seeds species cultivated in Poland. In lupin seed globulins hydrolysis studies, there were used enzymes typical for the human gastrointestinal tract, such as: pepsin, pancreatin, trypsin and chymotrypsin. The obtained hydrolysates were assessed based on the results of electrophoretic, immunoblotting, as well as chromatographic separations. The evaluation of lupin seed globulins digestion susceptibility was supported by the bioinformatics analyses. Analysis of hydrolysates showed that the proteins present in globulins are completely hydrolysed by: pepsin, double digestion model (pepsin followed by pancreatin) or chymotrypsin. The high specificity of pancreatin and trypsin action results in limited lupin globulins hydrolysis. Protein fraction resistant to the action of these enzymes was γ-conglutin which also retains its antigenic properties. Its insensitivity to the hydrolysis might be associated with the formation of complexes with flavonoids which were released from other protein connections during digestion, as well as with relatively low number of cleavage sites for trypsin. The analysis of the three-dimensional structure of γ-conglutin enabled a very accurate description of the amino acid residues localisation, at which trypsin hydrolysis should occur.

Proteomic analysis of Lupinus angustifolius (var. Zeus and Bojar) and Lupinus luteus (var. Lord and Parys) seed proteins and their hydrolysates

BACKGROUND Proteins enzymatic digestion is a very complex process, during which some components are degraded, whereas others remain in an unchanged form. Moreover, enzymatic hydrolysis is one of the most popular methods used to reduce the allergenicity of food proteins. In the present study, the efficiency of enzymatic hydrolysis of lupin seed proteins was assessed by proteomic analysis as performed by two-dimensional gel electrophoresis (2-DE) coupled with mass spectrometry identification. Two digestion systems were used: oriented digestion carried out by trypsin and model in vitro digestion mimicking the conditions present in the gastrointestinal tract. RESULTS The comparisons of 2-DE maps of proteins isolated form different lupin seed species revealed that the differences in proteins expression were observed mainly in the central parts of gels (i.e. in the molecular weight range from 20 to 70 kDa, and the pH range 5-7). In total, 27 differentially expressed proteins spots were successfully identified by mass spectrometry analysis. An important reduction in the number of proteins spots on 2-DE maps was observed when trypsin and the in vitro digestion model were applied. The protein spot insensitive to digestion in both hydrolysis systems was identified as β-conglutin. CONCLUSIONS The results of the present study provide insight into the nature of the digestion process that may take place after lupin seed protein intake and highlight the important fact that some of the proteins are insensitive to digestive enzyme activity. Moreover, evaluation of digestion activity of trypsin towards lupin seed proteins may be used for the development of specific processes with respect to hypoallergenic food production.

Immunoreactivity changes during lupin seed storage proteins digestion

Lupin seeds are already widely used as an ingredient in different food products. Their attractiveness is related mainly to their high protein content that is characterised by a favourable amino acid composition, as well as the desired technological properties. However, with the increase of lupin seeds usage in food manufacture, their potential allergic properties have been demonstrated. The aim of this work was to study the immunoreactivity changes taking place during the enzymatic hydrolysis of the major seed proteins of narrow-leafed (Lupinus angustifolius, varieties Zeus and Bojar) and yellow (L. luteus, var. Lord and Parys) lupin species. Two digestion systems were used, namely the in vitro model simulating digestion taking place in digestive track, and specific hydrolysis carried out by trypsin. The obtained hydrolysates were analysed by means of one-dimensional electrophoresis, and their immunoreactivity was assessed with the use of a sera pool from patients with lupin-specific IgE. An important reduction in allergenicity of lupin seed proteins was observed when trypsin digestion was applied. The digestion in the in vitro model revealed the possibility of formation of neoallergens which were identified on the basis of mass spectrometry results as β-conglutin fraction.