Purification of free arginine from chickpea (Cicer arietinum) seeds

© 2015 Elsevier Ltd. All rights reserved. Chickpea is a grain legume widely consumed in the Mediterranean region and other parts of the world. Chickpea seeds are rich in proteins but they also contain a substantial amount of free amino acids, especially arginine. Hence chickpea may represent a useful source of free amino acids for nutritional or pharmaceutical purposes. Arginine is receiving great attention in recent years because it is the substrate for the synthesis of nitric oxide, an important signaling molecule involved in numerous physiological and pathological processes in mammals. In this work we describe a simple procedure for the purification of arginine from chickpea seeds, using nanofiltration technology and an ion-exchange resin, Amberlite IR-120. Arginine was finally purified by precipitation or crystallization, yielding preparations with purities of 91% and 100%, respectively. Chickpea may represent an affordable green source of arginine, and a useful alternative to production by fermentation or protein hydrolysis.This work was carried out with the financial support of Junta de Andalucía (Spain) to the Laboratory of Bioactive and Functional Components of Plant Products (Instituto de la Grasa, C.S.I.C.). Cristina Megias is recipient of a JAE-Doc (C.S.I.C.) contract from the “Junta para la Ampliación de Estudios” program (cofinanced by the European Social Fund). Isabel Cortés-Giraldo is recipient of a JAE-Pre (C.S.I.C) fellowship from the “Junta para la Ampliación de Estudios” program (cofinanced by the European Social Fund). Thanks are due to María Dolores García-Contreras for technical assistance and to Jose Julian Rios for HPLC-MS analyses of purified arginine.Peer Reviewe

Nutritional Characteristics of the Seed Protein in 23 Mediterranean Legumes

The search for new sources of plant protein for food and animal feed is driven by an increasing demand in developing countries and the interest in healthy alternatives to animal protein. Seeds from 23 different wild legumes belonging to tribes Gallegeae, Trifolieae, and Loteae were collected in southern Spain and their total amino acid composition was analyzed, by reverse phase-high performance liquid chromatography (RP-HPLC), in order to explore their nutritional value. Protein content in the seeds ranged from 15.5% in Tripodium tetraphyllum to 37.9% and 41.3% in Medicago minima and Medicago polymorpha, respectively. Species belonging to tribe Trifolieae, such as Melilotus elegans and Trifolium spp., showed the most equilibrated amino acid composition and the best theoretical nutritional values, although all species were deficient in sulfur amino acids. The amino acid composition of the seeds from some of these legumes was characterized by high levels of the anticancer non-proteic amino acid canavanine This amino acid was found free in the seeds from some of the species belonging to each of the three tribes included in the present work. Astragalus pelecinus in tribe Gallegea, Trifolium angustifolium in tribe Trifolieae, and Anthyllis vulneraria in tribe Loteae have 3.2%, 3.7%, and 7.2% canavanine, respectively. Seeds from Anthyllis vulneraria, Hymenocarpus lotoides, and Hymenocarpos cornicina have the highest contents in canavanine overall. In conclusion, the seeds from some of these legumes could be used for human consumption and for feeding animals because they contain protein of good nutritional quality. These plants could be useful in domestication and breeding programs for production of new varieties with improved nutritional and functional properties. In addition, some of these species may be of interest as a source of the bioactive compound canavanineinfo:eu-repo/semantics/publishedVersio

Purificación y caracterización parcial de proteínas de almacenamiento de semillas de Lupinus angustifolius

Lupinus angustifolius seed proteins have been purified by sequential dialysis and ion exchange chromatography, and their amino acid composition has been studied in order to determine their nutritional value as sources of essential amino acids. Albumins include a great variety of proteins. Globulins were resolved in α, β and δ conglutins. Conglutin α is the main protein in the seeds of L. angustifolius, representing 76.6% of the total. While lysine was found to be the limiting amino acid in L. Angustifolius seed proteins as a whole, tyrosine was the limiting amino acid in albumins, and methione and lysine were limiting in globulins. Lysine, methionine and histidine were limiting amino acids in α conglutin.Se ha realizado una purificación por diálisis secuencial y cromatografía de intercambio iónico de las proteínas de la semilla de L. angustifolius, y su composición en aminoácidos ha sido estudiada para determinar su valor nutricional como fuentes de aminoácidos esenciales. Las albúminas están compuestas por una compleja mezcla de proteínas. Las globulinas se resolvieron en conglutinas α, β y δ. La conglutina α es la proteína más abundante en las semillas de L. angustifolius representando el 76.6% del total. Las proteínas de la semilla de L. angustifolius son limitantes en lisina. Las albúminas son limitantes en tirosina y las globulinas en metionina y lisina. La α conglutina es limitante en lisina, metionina e histidina

Mejora de la extracción proteica de la harina de girasol mediante hidrólisis con alcalasa

Extraction of proteins from defatted sunflower meal has been improved by addition of the protease alcalase during alkaline extraction. This method offers several additional advantages as compared to the traditional alkaline extraction without alcalase, which is usually carried out after a sedimentation/flotation step to remove the lignocellulosic fraction. As compared to extraction without alcalase, addition of 0.1% (v/v) alcalase improved the yield of protein extraction from 57.5% to 87.4%, providing an extract that is 22% hydrolyzed. In addition, an increment of up to 4.5 times in protein solubility at low pH values is achieved, which correlates with the degree of hydrolysis. The extracts that were obtained in the presence of alcalase had a higher proline and glycine content, suggesting that the protease improves extraction of proline-rich and glycine-rich cell wall proteins that are part of the lignocellulosic fraction. These protein extracts can be directly dried without generation of wastewater, and the resulting fiber-rich material could be used for animal feeding.Se ha mejorado la extracción proteica de la harina desengrasa de girasol mediante la adición de la proteasa alcalasa durante la extracción alcalina. Este método ofrece varias ventajas adicionales en comparación con la extracción alcalina tradicional sin alcalasa, que se desarrolla normalmente mediante un proceso de flotación/sedimentación para retirar la fracción lignocelulósica. En comparación a la extracción sin alcalasa, la adicción de 0.1% (v/v) de alcalasa mejora los rendimientos de extracción proteica desde un 57.5% a un 87.4%, dando un extracto con un 22% de grado de hidrólisis. Además se obtiene un incremento de hasta 4.5 veces de la solubilidad proteica a bajos pHs, que se correlaciona con el grado de hidrólisis. Los extractos obtenidos con alcalasa tenían un mayor contenido de prolina y glicina, sugiriendo que la proteasa mejora la extracción de las proteínas ricas en prolina y glicina de la pared celular que forma parte de la fracción lignocelulósica. Este extracto proteico puede ser secado directamente sin generación de aguas residuales, y el material resultante rico en fibra podría ser usado para alimentación animal.This work was supported by grant AGL 2001-0526.Peer reviewe

Determination of -Cyano-L-alanine, -Glutamyl--cyano-L-alanine, and Common Free Amino Acids in Vicia sativa (Fabaceae) Seeds by Reversed-Phase High-Performance Liquid Chromatography

A method for determination of -cyano-L-alanine, -glutamyl--cyano-L-alanine and other free amino acids in Vicia sativa is presented. Seed extracts were derivatized by reaction with diethyl ethoxymethylenemalonate and analyzed by reverse-phase highperformance liquid chromatography. Calibration curves showed very good linearity of the response. The limit of detection and quantification was 0.15 and 0.50 M, respectively. The method has high intra-(RSD = 0.28-0.31%) and interrepeatability (RSD = 2.76-3.08%) and remarkable accuracy with a 99% recovery in spiked samples. The method is very easy to carry out and allows for ready analysis of large number of samples using very basic HPLC equipment because the derivatized samples are very stable and have very good chromatographic properties. The method has been applied to the determination of -glutamyl--cyano-L-alanine, -cyano-L-alanine, and common free amino acids in eight wild populations of V. sativa from southwestern Spain

Polyphenol composition and in vitro antiproliferative effect of corm, tepal and leaf from Crocus sativus L. on human colon adenocarcinoma cells (Caco-2)

25 Páginas, 3 Figuras, 3 TablasSaffron is cultivated for production of the saffron spice. Nevertheless, a huge amount of saffron by-products including corms, tepals and leaves with little or no commercial value are generated during the processing of the spice. This biomass contains bioactive compounds whose exploitation can increase the profitability and sustainability of this traditional crop. A significant amount of polyphenols, mainly glycosides of kaempferol, luteolin and quercetin, have been determined in tepals and leaves of saffron. Proliferation of Caco-2 cells was greatly inhibited by the tepal and leaf extracts (ED50 0.42 mg/ml), while the corm extract caused some signs of toxicity and completely abolished proliferation (ED50 0.05 mg/ml). To our knowledge, these are the first data reporting the inhibition of the proliferation of Caco-2 cells by extracts from tepals and leaves of saffron, and polyphenols could be responsible for this effect.This work has been supported by the grant RTA2013-00005-00-00 (INIA, Spain) and is part of a Short Term Scientific Mission (STSM) carried out in the Agricultural University of Athens (AUA) and supported by the grant COST ActionFA1101 (http://www.saffronomics.org). We are grateful to the European Social Fund, Fundación Parque Científico y Tecnológico de Castilla-La Mancha and Junta de Andalucía for additional financing.Peer reviewe

Estudio estructural y de biodisponibilidad de compuestos de reacción entre lisina y aldehídos procedentes de oxidación lipidica

X, 174 páginas, 25 figuras, 15 tablas. Memoria de la Tesis presentada por el Ldo. en Biología Julio Girón Calle para optar al grado de Doctor en Biología.La reacción de proteínas alimentarias con lípidos oxidados o sus productos de descomposición provoca pérdidas de valor nutricional en las mismas. Es de especial interés la reacción de varios aldehídos, que se forman como productos secundar ios de oxidación lipídica, con los grupos ϵ-amino protéicos, por afectar a un aminoácido esencial y a menudo limitante como es la lisina. El mismo tipo de reacciones puede sufrir la lisina como aminoácido libre cuando se añade a determinados alimentos para corregir deficiencias.Se ha realizado una evaluación nutricional de alimentos y proteínas en los que químicamente se había comprobado la alteración de los restos de lisina proteicos, al haber sido sometidos a la acción de lípidos oxidados. Sin embargo, se hace necesaria una caracterización precisa de las nuevas estructuras químicas que se forman en estos procesos, para así poder luego estudiar la relevancia nutricional que pudiera tener su formación en los alimentos. Es por tanto necesaria una aproximación desde la Química Biológica a este problema. Esto es lo que nos proponemos en el presente trabajo para el caso de la reacción de lisina con ciertos aldehídos que es conocido que se producen en la oxidación lipídica como productos secundarios.Dado lo complejo que es el aislamiento y caracterización a partir de los alimentos de los compuestos que nos interesan (que se pueden producir con rendimientos muy bajos) se recurre al estudio de las estructuras de interacción que se forman en sistemas más sencillos. Siguiendo esta línea, estudiamos como modelo de la interacción entre lisina y aldehídos procedentes de oxidación lipídica la reacción del éster fenilazofenacílico de la lisina con malondialdehido y propanal. Se obtiene un perfil completo de la reacción, y se caracterizan en lo posible los productos formados mediante las técnicas habituales para la elucidación de estructuras de compuestos orgánicos.En una segunda parte del trabajo se estudia en animales de experimentación en tránsito metabólico de dos clases de productos cuya formación comprobamos en la primera parte de la investigación. Se realizan para ello experimentos de seguimiento de la excreción e incorporación a diversos órganos y tejidos de estos productos marcados radiactivamente. Asimismo se estudia su incorporación a microsomas hepáticos y su ruptura por homogenados totales de hígado, riñón y mucosa intestinal.Peer reviewe

Producción de hidrolizados proteicos de Lupinus angustifolius con mejores propiedades funcionales

Protein hydrolysates were obtained from lupin flour and from the purified globulin a-conglutin, and their functional properties were studied. Hydrolysis with alcalase for 60 minutes yielded degrees of hydrolysis ranging from 4% to 11% for lupin flour, and from 4% to 13% for a-conglutin. Protein solubility, oil absorption, foam capacity and stability, emulsifying activity, and emulsion stability of hydrolysates with 6% degree of hydrolysis were determined and compared with the properties of the original flour. The protein hydrolysates showed better functional properties than the original proteins. Most impor tantly, the solubility of the a-conglutin and L. angustifolius flour hydrolysates was increased by 43% and 52%, respectively. Thus, lupin seed protein hydrolysates have improved functional properties and could be used in the elaboration of a variety of products such as breads, cakes, and salad dressings.Se obtuvieron hidrolizados proteicos de la harina del altramuz y de la globulina α-conglutina purificada y se estudiaron sus propiedades funcionales. La hidrólisis con alcalasa durante 60 minutos produjo hidrolizados con grados de hidrólisis entre el 4 % y el 11 % para la harina y entre el 4 % y el 13 % para la α-conglutina. Se estudió en un hidrolizado con un 6 % de grado de hidrólisis la solubilidad proteica, absorción de aceite, capacidad y estabilidad espumante y actividad y estabilidad emulsificante. Los hidrolizados proteicos mostraron mejores propiedades funcionales que las proteínas originales. Más aún, la solubilidad de los hidrolizados de α-conglutina y la harina se incrementó en un 43 % y 52 % respectivamente. Así pues, hidrolizados de proteínas de semilla de lupino presentan mejores propiedades funcionales y podrían usarse en la elaboración de productos como pan, dulces, salsas o cremas.This work was supported by research grant AGL2001-0526 (F.M.), AGL2002-02836 (J.G.C.) and a Ramón y Cajal grant (J.G.C.) from Spanish Ministerio de Ciencia y Tecnología.Peer reviewe

Effect of chickpea protein hydrolysates on cell proliferation and in vitro bioavailability

6 páginas, 4 figuras.Bioactive peptides in foods may have health-promoting properties, including effects on cell proliferation. The goal of this study was to determine whether physiological digestion of chickpea protein using pepsin and pancreatin could release this type of peptides. The THP-1 and Caco-2 cell lines were used as in vitro models to determine the effects of the hydrolysates on cell proliferation. Hydrolysates with a high degree of hydrolysis inhibited the growth of Caco-2 cells by up to 45%, suggesting that chickpea-derived peptides might inhibit the growth of tumors in the colon. Proliferation of THP-1 cells was inhibited by up to 78% by direct exposure to the hydrolysates. Nevertheless, bioavailability experiments using differentiated Caco-2 cells as a model of the intestinal barrier, in co-culture with THP-1 cells, showed that proliferation of the monocytic THP-1 cells was actually enhanced by up to 66%, which could represent an immunomodulatory activity.This work was supported by the Ministerio de Ciencia e Innovación of Spain through grants AGL2005-01120 (J.G.C.), and AGL2007-63580 (M.A.), which were partially supported with FEDER funds from the European Union.Peer reviewe