M. Muzquiz

Determination of phytate and lower inositol phosphates in Spanish legumes by HPLC methodology

Abstract The separation and quantitative determination of phytic acid (inositol hexaphosphate) and lower inositol phosphates (tri-, tetra- and penta-phosphates) was carried out using a HPLC method that included purification by anion-exchange chromatography, and analysis by ion-pair chromatography on a C18 reverse phase column. Samples of the most important legumes in the Mediterranean diet were analysed using this system. The different samples presented a variable content of phytic acid and different relative proportions of lower inositol phosphates.

Determination of caffeic and chlorogenic acids and their derivatives in different sunflower seeds

Five different genotypes (Tesoro, Marko, Clip, Vyp-70 and Nanta) of sunflower seeds were studied and the phenolic compositions of their kernel and hull were characterised by HPLC and MS techniques. The main phenolic compounds present in both the kernel and hull are chlorogenic acid, caffeic acid and two caffeoylquinic derivatives. Both derivatives were characterised by partial alkaline hydrolysis and LC/MS. The four phenolic compounds present in the kernel ranged from 94.6% (genotype Clip) to 99.3% (genotype Marko) of the total phenolic compounds of the whole seed. Therefore the dehulling of the seed scarcely improves its nutritional value. # 2000 Society of Chemical Industry

Heat and pressure treatments effects on peanut allergenicity.

Peanut allergy is recognized as one of the most severe food allergies. The aim of this study was to investigate the changes in IgE binding capacity of peanut proteins produced by thermal-processing methods, including autoclaving. Immunoreactivity to raw and thermally processed peanut extracts was evaluated by IgE immunoblot and skin prick test in patients with clinical allergy to peanut. Roasted peanut and autoclaved roasted peanut were selected for IgE ELISA experiments with individual sera, immunoblot experiments with antibodies against peanut allergens (Ara h 1, Ara h 2 and Ara h 3), digestion experiments, and circular dichroism spectroscopy. In vitro and in vivo experiments showed IgE immunoreactivity of roasted peanut proteins decreased significantly at extreme conditions of autoclaving. Circular dichroism experiments showed unfolding of proteins in autoclave treated samples, which makes them more susceptible to digestion. Autoclaving at 2.56atm, for 30min, produces a significant decrease of IgE-binding capacity of peanut allergens.

Evaluation of antinutritional factors of selected varieties of Phaseolus vulgaris

The antinutrient composition of 19 varieties of Phaseolus vulgaris (L) grown in different localities of Spain was determined. Moreover, eight of the 19 varieties were grown in two different environmental areas with the aim of studying the influence of variety and environment on antinutrients and the existence of relationships between some antinutritional factors. The content of sucrose and raffinose oligosaccharides was determined by HPLC. Stachyose was the predominant a-galactoside in all the samples. Raffinose and verbascose were also present in significant quantities. Capillary-gas chromatography of the bean samples revealed only one peak corresponding to soya- sapogenol B, which is common to both saponins I and V. The content of lectins (PHA) determined by ELISA methodology showed a high variation. Statistical analyses of data established that the compounds analysed were influenced by both variety and environment. The possibility of selecting some varieties to be used for large-scale cultivation in Spain is discussed. # 1999 Society of Chemical Industry

Lupins as a potential source of raffinose family oligosaccharides: Preparative method for their isolation and purification

Abstract Results of studies on the content of raffinose family oligosaccharides (RFOs) in plants of 11 species of the Leguminosae family are presented. It is clearly shown that plants of the Leguminosae family, especially of the lupin species, are the richest source of these sugars among the plants tested. A simple method of preparative isolation and purification of RFOs from bitter lupin extract was developed. This method seems to be useful for efficient isolation of oligosaccharides from plant material for further nutritional tests.

Effect of germination, under different environmental conditions, on saponins, phytic acid and tannins in lentils (Lens culinaris)

The effect of germination conditions on some antinutrients of Lens culinaris var Magda 20 seeds were studied. The seeds were germinated at 20°C under variable conditions of time, water and light. Quantitative analyses of the soyasapogenols, inositol phosphates and tannins were carried out by capillary gas chromatography, high-performance liquid chromatography and spectrophotometric techniques respectively. Germinated seeds at day 6 contained higher levels soyasapogenol B than the controls, whereas in general the tannin content was reduced. Total phytic acid amounts did not decrease after 3 days of germination but was greatly reduced after 6 days. This work shows that the optimal conditions to reduce some antinutritional factors (tannins and phytic acid) in lentils were 6 days of seed germination in dark and with alternate watering. Therefore, germination conditions offer a good opportunity to improve the nutritional quality of lentils.

Influence of thermal processing on IgE reactivity to lentil and chickpea proteins.

In the last years, legume proteins are gaining importance as food ingredients because of their nutraceutical properties. However, legumes are also considered relevant in the development of food allergies through ingestion. Peanuts and soybeans are important food allergens in Western countries, while lentil and chickpea allergy are more relevant in the Mediterranean area. Information about the effects of thermal-processing procedures at various temperatures and conditions is scarce; therefore, the effect of these procedures on legume allergenic properties is not defined so far. The SDS-PAGE and IgE-immunoblotting patterns of chickpeas and lentils were analyzed before and after boiling (up to 60 min) and autoclaving (1.2 and 2.6 atm, up to 30 min). The results indicated that some of these treatments reduce IgE binding to lentil and chickpea, the most important being harsh autoclaving. However, several extremely resistant immunoreactive proteins still remained in these legumes even after this extreme treatment.

Effect of soaking, cooking and germination on the oligosaccharide content of selected Nigerian legume seeds

The identity and quantity of and effect ofprocessing on raffinose oligosaccharides in raw,mature seeds of lima beans (Phaseolus lunatus),pigeon peas (Cajanus cajan), African yam beans(Sphenostylis sternocarpa) and jackbeans (Canavalia ensiformis) were investigated. Sucrose,raffinose, stachyose and verbascose were identified byHPLC in all the legume seeds. The totalα-galactoside contents of the seeds in decreasing order were African yam beans3.84 mg/100 mg; white lima beans 3.62 mg/100 mg; creampigeon peas 3.51 mg/100 mg; red lima beans3.37 mg/100 mg; jackbeans 2.83 mg/100 mg and brownpigeon peas 2.34 mg/100 mg. The predominantoligosaccharide was verbascose in pigeon peas andstachyose in the other three legumes. Cooking unsoakedseeds brought about a greater reduction in the totalα-galactoside content than soaking for ninehours. The removal of oligosaccharides was higher inlegumes cooked in alkaline solution than in water.Germination quantitatively reduced raffinose,stachyose and verbascose while sucrose was increasedin all seeds except red lima beans and jackbeans.

Bioactive compounds in legumes: pronutritive and antinutritive actions. Implications for nutrition and health

Legume seeds are employed as a protein source for animal and human nutrition not only for their nutritional value (high in protein, lipids and dietary fibre), but also their adaptability to marginal soils and climates. Human consumption of legumes has been increased in recent years, being regarded as beneficial food ingredients. Legume seeds contain a great number of compounds which qualify as bioactive compounds with significant potentials benefits to human health. These compounds vary considerably in their biochemistry and they can be proteins, glycosides, tannins, saponins, alkaloids, etc. Hence, methods for their extraction, determination and quantification are specific of each compound. They do not appear equally distributed in all legumes, and their physiological effects are diverse. Some of these compounds are important in plant defence mechanisms against predators or environmental conditions. Others are reserve compounds, accumulated in seeds as energy stores in readiness for germination. Processing generally improves the nutrient profile of legume seed by increasing in vitro digestibility of proteins and carbohydrates and at the same time there are reductions in some antinutritional compounds. Most antinutritional factors are heat-labile, such as protease inhibitors and lectins, so thermal treatment would remove any potential negative effects from consumption. On the other hand tannins, saponins and phytic acid are heat stable but can be reduced by dehulling, soaking, germination and/or fermentation. New directions in bioactive compounds research in the last decade have led to major developments in our understanding of their role in nutrition. The scientific interest in these compounds is now also turning to studies of their possible useful and beneficial applications as gut, metabolic and hormonal regulators and as probiotic/prebiotic agents.

Influence of Enzymatic Hydrolysis on the Allergenicity of Roasted Peanut Protein Extract

Background: Peanut allergy is recognized as one of the most severe food allergies. Some studies have investigated the effects of enzymatic treatments on the in vitro immunological reactivity of members of the Leguminosae family, such as the soybean, chickpea and lentil. Nevertheless, there are only a few studies carried out with sera from patients with a well-documented allergy. Methods: Roasted peanut protein extract was hydrolyzed by the sequential and individual action of 2 food-grade enzymes, an endoprotease (Alcalase) and an exoprotease (Flavourzyme). Immunoreactivity to roasted peanut extract and hydrolyzed samples was evaluated by means of IgE immunoblot, ELISA and 2-dimensional electrophoresis using sera from 5 patients with a clinical allergy to peanuts and anti-Ara h 1, anti-Ara h 2 and anti-Ara h 3 immunoblots. Results: Immunoblot and ELISA assays showed an important decrease of IgE reactivity and Ara h 1, Ara h 2 and Ara h 3 levels in the first 30 min of hydrolyzation with Alcalase. In contrast, individual treatment with Flavourzyme caused an increase in IgE reactivity detected by ELISA at 30 min and led to a 65% inhibition of IgE reactivity at the end of the assay (300 min). Ara h 1 and the basic subunit of Ara h 3 were still recognized after treatment with Flavourzyme for 300 min. Conclusion: Hydrolysis with the endoprotease Alcalase decreases IgE reactivity in the soluble protein fraction of roasted peanut better than hydrolysis with the exoprotease Flavourzyme.