Bean Inhibitor Research Articles

PURIFICACIÓN Y CARACTERIZACIÓN DE INHIBIDORES DE PAPAÍNA PROVENIENTES DE SEMILLAS DE AMARANTO (Amaranthus caudatus) Y FRIJOL (Phaseolus vulgaris)

Los inhibidores de proteasas vegetales con frecuencia son proteínas de baja masa molar, presentes en tejidos de almacenamiento y aéreos de las plantas. Su expresión se induce en respuesta al daño por insectos y microorganismos patógenos y el contenido de estos inhibidores puede ser alto en órganos de cereales y leguminosas. Bajo la premisa de que la combinación de técnicas de purificación de proteínas de baja y alta resolución permite incrementar la actividad específica de inhibidores de proteasas presentes en tejidos vegetales, el objetivo de este estudio fue aislar, purificar y caracterizar bioquímicamente inhibidores de papaína a partir de semillas de amaranto (Amaranthus caudatus) y frijol (Phaseolus vulgaris). El diseño experimental fue completamente al azar para seleccionar las técnicas de baja resolución que incrementan significativamente (p≤0.05) el grado de purificación de los extractos tratados. En todos los estudios se hicieron tres repeticiones y las mediciones se realizaron por triplicado. A partir de harinas de semillas de amaranto y frijol se obtuvieron extractos proteicos crudos que después se ultrafiltraron en membranas de 10 kDa. Las fracciones de baja masa molar se purificaron por cromatografía de afinidad en papaína-glioxil-sepharosa 6B-CL. El extracto crudo de amaranto presentó actividad inhibidora de 1.177 ± 0.067 mU·mL-1 y el de frijol 1.556 ± 0.542 mU·mL-1. Los extractos con purificación parcial de amaranto y frijol se purificaron por cromatografía de afinidad en papaína-glioxil-sepharosa 6B-CL, hasta 137.5 y 79.1 veces, respectivamente. La masa molar estimada fue 7.50 kDa para el inhibidor de amaranto y 8.20 kDa para el de frijol. El inhibidor de amaranto mostró inhibición competitiva con Ki de 0.872 µM, mientras que la del frijol fue no competitiva y con Ki de 0.058 µM. Esto evidencia la presencia de inhibidores de cisteíno proteasas en las semillas de amaranto y frijol.

Inhibitory effect of aqueous bean extract, Phaseolus vulgaris (fabaceae), on α-amylase of the cabbage aphid, Brevicoryne brassicae

ABSTRACT Brevicoryne brassicae is one of the most important pests of wide range of cruciferous host plants. In this study, the inhibitory effects of aqueous extracts of five cultivars of common bean (Phaseolus vulgaris L.), that includes white beans, pinto, red, cowpea and keshavarzi on the activity of the α-amylase enzyme of B. brassicae was studied. Aphids were reared under laboratory conditions with 25 ± 1°C, 60 ± 5% RH and 16 L:8D h photoperiods. According to obtained results, the cytotoxic effect of extracts of the highest concentration of 0.05 mg ml−1 and within 72 hours had the best performance and mortality. Also, all bean extracts had an inhibitory effect on insect α-amylase enzyme and red bean had the highest inhibitory effect (IC50 = 0.223). The effects of the bean inhibitor on α-amylase of B. brassicae showed a dose-dependent manner of inhibition, e.g., less inhibition of enzyme activity (around 10%) with a lower dose (0.25 mg protein) and high inhibition of enzyme activity (around 90%) when a high dose of inhibitor was used (1 mg protein). Based on the data presented in this study, it could be said that the bean extract has good inhibitory activity on B. brassicae α-amylase.

Исследование in silico структуры альфа-амилазы колорадского жука и ее взаимодействия с ингибиторами

To develop effective and environmentally friendly methods for protecting crop plants from pests, it is necessary to know the structure of pests hydrolases and the mechanisms of their interaction with plant inhibitors. The aim of this work was a modeling of the spatial structure of Colorado potato beetle alpha-amylase, analysis of the effect of revealed structural features on interaction with protein inhibitors, as well as modeling of interaction with amylase inhibitors from various organisms.The spatial structure of the Colorado potato beetle alpha-amylase was obtained by the method of computer modeling using the IntFOLD and NOMAD-Ref services. Colorado potato beetle amylase differs in the structure and physicochemical properties of the active site in comparison with flour beetle amylase, but has the same conformation of the main chain. The possible effect of these differences on the interaction of the enzyme with plant inhibitors – a lectin-like bean inhibitor and a RATI inhibitor – was shown. The CABS-dock service was used to model the interaction of Colorado potato beetle amylase with polypeptide inhibitors of amylase from various plant species, bacteria, and actinomycetes. Among the structures obtained, the best values of complex free energy were possessed by the knottin-like inhibitor of amaranth and the purothionin-like inhibitor of oats; the least favorable is the binding with the inhibitors of the actinobacterium Thermopolyspora flexuosa and the aquatic plant Alternanthera sessilis. The model structure of a knottin-like inhibitor in a complex with Colorado potato beetle amylase, in comparison with the known structure of a complex with flour beetle amylase, has a similar chain folding, but significantly different conformation. The data obtained can be used to search for new effective pests’ amylase inhibitors and environmentally friendly methods of protecting potato plants from insect pests, including using the genetic transformation of plants.

Identification and characterization of hydrolytic enzymes from the midgut of Sunn Pest of wheat (Eurygaster integriceps)

To help in the development of Sunn Pest-resistant transgenic plants employing protease or α-amylase inhibitors, midgut hydrolytic enzymes of Sunn Pest (Eurygaster integriceps, Put.) (Heteroptera: Scutelleridae) were identified and characterized biochemically. We observed levels of very low proteolytic activity of trypsin (3 nmoles/min/mg), elastase (0.66 nmoles/min/mg) and leucine aminopeptidase-like (14.4 nmoles/min/mg) proteases, but no chymotrypsin and papain-like activity. Proteolytic activities were insensitive to inhibition by soybean trypsin inhibitor (SBTI) and aprotinin, but were inhibited to varying degrees (40–100%) by the synthetic protease inhibitors PMSF, TPCK, CdCl2 and CuCl2. Compared to proteolytic activity, significantly higher amylolytic activity (4.45 mmoles/min/mg) was observed. Αlpha-amylase activity was found to be resistant to inhibition by bean and chickpea α-amylase inhibitors, and only slightly inhibited by wheat (36.9 ± 1.8%) and maize (40.2 ± 2.8) α-amylase inhibitors. Here, we report on the biochemical properties of digestive enzymes from Sunn Pest midgut, and their inhibition patterns by several synthetic and natural hydrolytic enzyme inhibitors.

Isolation and partial sequence of a Kunitz-type elastase specific inhibitor from marama bean (Tylosema esculentum)

An isolation procedure utilizing ammonium sulfate fractionation and affinity chromatography was used to purify an elastase inhibitor present in large amounts in marama beans (Tylosema esculentum). The protein appeared to be heterogeneous due to carbohydrate differences, demonstrating two bands on SDS gels with molecular weights of 17.8 kDa and 20 kDa. Partial sequence, derived from mass spectrometry, indicated that the protein is a Kunitz-type inhibitor distinct from other known plant serine protease inhibitors. The marama bean inhibitor is specific for elastase, with very low Ki for both pancreatic and neutrophil elastase. The quantity of elastase inhibitor present in marama beans is many times greater than in soybean or any other bean or nut source reported to date. This raises the question of why a bean found in an arid corner of the Kalahari Desert would be so rich in a very potent elastase inhibitor.

Purification and characterization of two α-amylase inhibitors from seeds of tepary bean ( Phaseolus acutifolius A. Gray)

Two proteinaceous α-amylase inhibitors termed αAI-Pa1 and αAI-Pa2 were purified from seeds of a cultivated tepary bean ( Phaseolus acutifolius A. Gray, cv. PI311897). The two inhibitors differed in their specificity towards α-amylases of insect pests such as bruchids, although neither showed any inhibitory activity against α-amylases of mammalian, bacterial or fungal origin. αAI-Pa2 resembles two common bean inhibitors, αAI-1 and αAI-2, in several characteristics such as N-terminal amino acid sequences and oligomeric structure being composed of α and β subunits. In contrast αAI-Pa1 is composed of a single glycopolypeptide with a molecular mass of 35 kDa, and its N-terminal amino acid sequence resembled that of seed lectins in tepary bean and common bean. The information on the two tepary bean α-amylase inhibitors may be useful not only for providing insight into critical structure for the specificity towards different α-amylase enzymes but also for enhancing insect resistance in crops.

X-Ray study on an artificial mung bean inhibitor complex with bovine β-trypsin in neat cyclohexane

The active trypsin inhibiting component, SPC1, was obtained during the synthesis of a 22-residue peptide with three disulfide bridges according to the mimic mung bean Bowman-Birk type inhibitor. The K i value of SPC1 is 1.2×10 −7 M. In order to determine the topological structure of SPC1, X-ray diffraction studies were carried out on the complex of SPC1 with bovine β-trypsin. Only the binding loop of SPC1 resolved at 2.2 Å resolution due to conformational flexibility of the other residues [1]. The amino acid sequence was re-determined and electrospray mass spectroscopy was also performed to ensure that no cleaving occurred on SPC1 and the primary sequence of SPC1 is correct. Because the protein is more rigid in nonaqueous medium as has been proved by others [2], we treated the complex of SPC1 with neat cyclohexane and then subjected it to X-ray diffraction analysis, and the result showed that all the 22 residues of SPC1 were located in the electron density map. So the topological structure of SPC1 has been determined, suggesting that crystal treatment with cyclohexane may be used as a method to determine the conformation of the disordered regions in protein crystal structures.

Complete amino acid sequences of three proteinase inhibitors from white sword bean (Canavalia gladiata).

Three major serine proteinase inhibitors (SBI-1, -2, and -3) were purified from the seeds of white sword bean (Canavalia gladiata) by FPLC and reversed-phase HPLC. The sequences of these inhibitors were established by automatic Edman degradation and TOF-mass spectrometry. SBI-1, -2, and -3 consisted of 72, 73, and 75 amino acid residues, with molecular masses of 7806.5, 7919.8, and 8163.4, respectively. The sequences of SBI-1 and -2 coincided with those of CLT I and II [Terada et al. (1994) Biosci. Biotech. Biochem., 58, 376-379] except only N- or C-terminal amino acid residues. Analysis of the amino acid sequences showed that the active sites of the inhibitors contained a Lys21-Ser22 against trypsin and Leu48-Ser49 against chymotrypsin, respectively. Further, it became apparent that about seven disulfide bonds were present. These results suggest that sword bean inhibitors are members of the Bowman-Birk proteinase inhibitor family.

Crystal Structure of Mung Bean Inhibitor Lysine Active Fragment Complex with Bovine β-trypsin at 1.8Å Resolution

The crystal structure of the complex of mung bean inhibitor lysine active fragment with bovine β-trypsin has been determined by X-ray crystallographic analysis at a resolution of 1.8 Å. Refinement of the model of the complex converged at a final R value of 0.16. From the resulting electron density map, about one-third of the residues of the inhibitor were identified and two residues, at position P4 and P2′ respectively, were found to be inconsistent with the sequence reported previously. The peptide chain of the inhibitor at the trypsin active site turns back sharply at Pro23I and forms a 9-residue reactive loop, which interacts with trypsin in a similar manner to the other families of inhibitors, suggesting an important and common role of these regions in exhibiting inhibitory activity.

A plant-seed inhibitor of two classes of alpha-amylases: X-ray analysis of Tenebrio molitor larvae alpha-amylase in complex with the bean Phaseolus vulgaris inhibitor.

The alpha-amylase from Tenebrio molitor larvae (TMA) has been crystallized in complex with the alpha-amylase inhibitor (alpha-AI) from the bean Phaseolus vulgaris. A molecular-replacement solution of the structure was obtained using the refined pig pancreatic alpha-amylase (PPA) and alpha-AI atomic coordinates as starting models. The structural analysis showed that although TMA has the typical structure common to alpha-amylases, large deviations from the mammalian alpha-amylase models occur in the loops. Despite these differences in the interacting loops, the bean inhibitor is still able to inhibit both the insect and mammalian alpha-amylase.

Purification and Characterization of Protease Inhibitor from Rice Bean (Vigna umbellata T.) Seeds

A Protease inhibitor from seeds of rice bean has been purified to apparent homogeneity as judged by native-PAGE with about 29% recovery using ammonium sulfate fractionation, ion exchange chromatography on DEAE-cellulose, and gel filtration through Sephadex G-100. The purified preparation with molecular weight of 16.8 kD was found to be a monomer as revealed by SDS-PAGE under reducing and non-reducing conditions. The purified inhibitor in solution was stable upto 40°C. However, it lost its activity gradually and was completely inactive when heated at 100°C for 2 h and 125°C for 1 h. Heating at 125°C changed the conformation of the inhibitor as was evident from the altered UV spectrum compared to that of native. It had two pH optima at pH 6.0 and at 10.0 and was stable over a wide range of pH (pH 3.0 to 10.0). It lost its activity on exposure to 2-mercaptoethanol, indicating the role of S-S linkages in maintaining the three dimensional structure of the protein inhibitor. The inhibitor was completely inactive towards papain, while it inhibited pepsin only slightly. Trypsin and chymotrypsin were inhibited upto the extent of 60% and 30%, respectively. Trypsin inhibition was of non-competitive type with dissociation constant for the enzyme-inhibitor complex in the region of 2.07 mg ml-1. The rice bean inhibitor appears to be of Bowman-Birk type as it has molecular weight lower than that generally observed for Kunitz type inhibitors and seems to be double headed - a characteristic specific of Bowman - Birk type inhibitor.

Genetic engineering with α-amylase inhibitors makes seeds resistant to bruchids

AbstractSeeds of the common bean,Phaseolus vulgaris, contain two inhibitors of mammalian and insect α-amylases (αAls) that show specificity towards the amylases of different insect species. Expression in pea (Pisum sativum) and azuki bean (Vigna angularis) of a chimeric gene consisting of the cDNA of bean αAl-1 and a seed-specific promoter makes the seeds of these legumes resistant to three species of Old World bruchids whose amylases are inhibited by αAl-1. This was the first successful genetic engineering of insect resistance in seeds. To understand the basis of the specificity between amylases and inhibitors we cloned a second bean inhibitor (αAl-2) with different specificity, and we cloned the cDNA of the New World bruchid,Zabrotes subfasciatus. The amylase of this bruchid is inhibited by αAl-2, but not by αAl-1. Knowledge of the amino acid sequences and of the three-dimensional structure of the pancreatic α-amylase–αAl-1 complex allows us to predict the peptide domains and amino acids of the proteins that are important for protein–protein recognition and inhibition of enzyme activity.

Characterization of Helicoverpa armigera gut proteinases and their interaction with proteinase inhibitors using gel X-ray film contact print technique.

Since Helicoverpa armigera is a devastating pest, an attempt was made to separate its gut proteinases and assess their diversity. Gelatin coating present on the X-ray film was used as a substrate to detect electrophoretically separated proteinases of H. armigera gut extract on native polyacrylamide gel electrophoresis (PAGE), sodium dodecyl sulfate (SDS)-PAGE and isoelectric focusing gels. The method involves electrophoresis, followed by washing the gel with Triton X-100 in case of SDS-PAGE, equilibration of the gel in proteinase assay buffers, overlaying the gel on X-ray film followed by washing the film with hot water to remove hydrolyzed gelatin revealing bands of proteinase activity. Using this protocol, at least six different proteinase isoforms were detected in H. armigera gut contents while three isoproteinases were identified in a commercial bacterial proteinase preparation. Adoption of the technique facilitated characterization of the H. armigera gut proteinases (HGP) and provided an easy tool to study the properties of the individual proteinases without purification. The approximate molecular masses of HGP as determined by SDS-PAGE were: 172.9, 59.3, 54.9, 47.6, 44.1 and 41.6 kDa, and of bacterial proteinases: 180.7, 127.3 and 95.3 kDa. The isoelectric point (pI) values of HGP and bacterial proteinase were in the range of 5.1-7.1 and 3.5-7.7, respectively. Some of the HGP isoforms were found to be highly pH-sensitive and showed activity only at pH 10.0. The major HGPs were inhibited by phenylmethylsulfonyl fluoride but not by (4-amidinophenyl)-methanesulfonyl fluoride. Incubation of HGP-resolved electrophoretic gel strips in chickpea or winged bean proteinase inhibitor solution permitted identification of specific inhibitors of individual proteinases and revealed that the major HGPs were insensitive to chickpea inhibitors whereas winged bean inhibitors effectively inhibited all the HGPs. Our results suggest that considerable variability exists among the isoproteinases of H. armigera gut with respect to their pH optima and sensitivity towards chemical and plant proteinase inhibitors. Such diversity is of immense biological significance as it explains the polyphagous nature of the insect which imparts unique adaptability to it against the defensive proteinase inhibitors of its wide range of host plants.

α-Amylase Activities in Larval Midgut Extracts from Four Species of Lepidoptera (Tortricidae and Gelechiidae): Response to pH and to Inhibitors from Wheat, Barley, Kidney Bean, and Streptomyces

Levels of α-amylase activity between 0.6 and 5.0 µmoles maltose minute-1 gram insect-1 occurred in larval midgut extracts from 4 species of lepidopterous pests-light brown apple moth, Epiphyas postvittana (Walker), Planotortrix octo (Dugdale), and Ctenopseustis Obliquana (Walker) are pests of apple leaves and fruit, and potato tuber moth, Phthorimaea Operculella (Zeller), is a pest of potato tubers and foliage. Activity response curves described the optima for activity at a highly alkaline pH range (pH 10.5 in leafrollers and pH 9.5 in potato tuber moth). This result is consistent with the known high midgut pH in other species of Lepidoptera and with the pH optima of their digestive proteases. The 3 wheat derived α-amylase inhibitors inhibited enzyme activities of all 4 species of Lepidoptera, but the other 3 (barley tetramer, Streptomyces, and kidney bean inhibitors) had no effect. Wheat dimeric inhibitor (30 µM) completely inhibited α-amylase activity in potato tuber moth midgut extracts.

The influence of pea seed trypsin inhibitors on the in vitro digestibility of casein

AbstractThe influence of pea seed trypsin inhibitors on the digestion of casein has been compared with that of soya bean trypsin inhibitors, using an in vitro digestion system, based on porcine pepsin and pancreatic enzymes. The data obtained suggest that pea trypsin inhibitors are much less effective inhibitors of digestion than the two soya bean inhibitors tested. The different behaviour of the inhibitors in the system used is not due to a differential sensitivity to pepsin, but may, in part, reflect a differential modification of the inhibitors by pancreatic enzymes.

Effects of bean and wheat α‐amylase inhibitors on α‐amylase activity and growth of stored product insect pests*

AbstractInsect α‐amylase inhibiting and/or growth inhibiting activities of proteinaceous inhibitors from red kidney bean (Phaseolus vulgaris) and hard red winter wheat (Triticum aestivum) were examined. The bean inhibitor was most effective in vitro against α‐amylases from the red flour beetle (Tribolium castaneum) and the confused flour beetle (T. confusum), followed by those from the rice weevil (Sitophilus oryzae) and yellow mealworm (Tenebrio molitor). The insect enzymes were from two‐ to 50‐fold more susceptible than human salivary α‐amylase. When the inhibitors were added at a 1% level to a wheat flour plus germ diet, the growth of red flour beetle larvae was slowed relative to that of the control group of larvae, with the bean inhibitor being more effective than the wheat inhibitor. Development of both the red flour beetle and flat grain beetle (Cryptolestes pusillus) was delayed by 1% bean inhibitor, but development of the sawtoothed grain beetle (Oryzaephilus surinamensis) and lesser grain borer (Rhyzopertha dominica) was not affected by either the bean or wheat inhibitor at the 1% level. Rice weevil adults fed a diet containing 1% bean or wheat inhibitor exhibited more mortality than weevils fed the control diet. When the wheat amylase inhibitor was combined with a cysteine protease inhibitor, E‐64, and fed to red flour beetle larvae, a reduction in the growth rate and an increase in the time required for adult eclosion occurred relative to larvae fed either of the inhibitors separately. The bean inhibitor was just as effective alone as when it was combined with the protease inhibitor. These results demonstrate that plant inhibitors of insect digestive enzymes act as growth inhibitors of insects and possibly as plant defense proteins, and open the way to the use of the genes of these inhibitors for genetically improving the resistance of cereals to storage pests.

Difference in Digestive Proteolysis in the Stored Maize Beetles: Sitophilus zeamais (Coleoptera: Curculionidae) and Prostephanus truncatus (Coleoptera: Bostrichidae)

Digestive proteinases in two maize feeding, stored-product insects, Sitophilus zeamais Motschulsky and Prostephanus truncates (Horn), were identified by determining the pH for maximal hydrolysis of synthetic protease substrates and the effects of potential protease activators and inhibitors. A thiol-activated cysteine proteinase was identified in S. zeamais by Bz-Arg-NHNa (benzoyl-DL-arginine- β -naphthylamide) hydrolysis with maximal activity at pH 5.5. Substrate hydrolysis was activated by thiol compounds, inhibited by iodoacetamide, and was not affected by either E64 aprotinin or soy bean inhibitor. Bz-Arg-Np (benzoyl-DL-arginine- p -nitroanilide) was not hydrolyzed by S. zeamais . Trypsin was identified in P. truncates by maximal hydrolysis of Bz-Arg-NHNa and Bz-Arg-Np at pH 9.0 and 8.5, respectively. Hydrolysis of both substrates was inhibited by aprotinin, soy bean inhibitor, and tosyl-L-lysine chloromethyl ketone and was not affected by either E64 or iodoacetamide. Chymotrypsin was identified in both insects by the hydrolysis of Bz-Tyr-OEt (benzoyl-L-tyrosine ethyl ester); maximal activity occurred at pH 7.5 in S. zeamais and at pH 8.5 in P. truncates . Substrate hydrolysis was inhibited by soy bean inhibitor, aprotinin, and tosyl-L-phenylalanine-chloromethyl ketone.

The 0.25-nm X-ray structure of the Bowman-Birk-type inhibitor from mung bean in ternary complex with porcine trypsin.

The structure of the Bowman-Birk-type inhibitor from mung bean Phaseolus aureus has been determined in ternary complex with porcine trypsin. The complex formed crystals of the trigonal space group P3(1)21 which diffracted to a resolution of 250 pm. Each of the two mung bean protease reactive sites is bound to trypsin according to the standard mechanism for serine proteinase inhibition. The binding loops thereby adopt the canonical conformation for the standard mechanism; however, the sub-van der Waals contact between the active-site serine O gamma (195) and the P1 carbonyl carbon of both loops is significantly smaller (210 pm) than hitherto observed, with continuous electron density connecting the two atoms. The inhibitor is formed by two double-stranded antiparallel beta-sheets, which are connected into a moderately twisted beta-sheet by a network of hydrogen bonds involving main-chain atoms and two water molecules. All contacts with neighbors in the crystal lattice occur between trypsin molecules. This apparently gives rise to an unusual form of disorder where the complexes pack in two orientations Ta:MaMb:Tb and Tb:MbMa:Ta (Ta, Tb = trypsin, Ma = mung bean loop I, Mb = mung bean loop II), such that the asymmetric unit consists of the ternary complex in two orientations, each with half occupancy. This is nearly equivalent to an asymmetric unit which has one trypsin molecule with full occupancy and one mung bean inhibitor with half occupancy and a crystallographic twofold symmetry axis through its center. Because of the approximate twofold symmetry of the inhibitor itself, however, the electron density was interpretable for most of the inhibitor (17 residues at the termini were not resolved) and shows evidence of its double orientation.

Raw Soy and Purified Proteinase Inhibitors Induce the Appearance of Inhibitor-Resistant Trypsin and Chymotrypsin Activities in Wistar Rat Duodenal Juice

Rats were fed raw soybeans or purified soybean proteinase inhibitors by tube to see whether they were able to produce inhibitor-resistant trypsin, as previously demonstrated in humans. Their duodenal chyme contained only 20-50% of the enzymatic activities of animals fed bovine serum albumin (BSA) as test protein. However, both tryptic and chymotryptic activities had considerable resistance to low- and high-molecular-weight inhibitors of serine proteinases. In particular, the tryptic activity demonstrated a high degree of inhibitor resistance. Human alpha 1-antitrypsin and lima bean inhibitor in amounts that inhibited bovine serum albumin-induced trypsin completely caused only 2-12% inhibition of the raw soybean-induced tryptic activity. The inhibitor-resistant tryptic and chymotryptic activities after raw soybean instillation might be caused by the Bowman-Birk and Kunitz trypsin inhibitors. The physiologic significance of an inhibitor-resistant trypsin might be to assure activation of other pancreatic proenzymes. The results of the present rat experiments confirm the previous findings of inhibitor-resistant trypsin in humans.

The Nutritive Value of Ten Inbred Lines of Faba Beans (Vicia faba L.) in Relation to their Content of Antinutritional Constituents and Protein Quality

AbstractTen inbred lines of fabe beans (Vicia faba L.) selected according to their quality characters have been investigated for carbohydrates, proteins, and antinutritional compounds. Digestible energy, N‐balance trials with growing rats comprising determination of the protein digestibility and biological value were used as criteria in connection with comprehensive chemical‐biochemical analysis.The chemical composition of the ten lines showed a considerable diversity as cad the results from the rat crisis. However, the content of vicine and convicine in all or the investigated lines were below the level previously found to have effects on the nutritive value, Starch, protein and fibre were the quantitatively dominating seed constituents, and all showed great variation among the lines.The starch content was not correlated to the quality or nutritive value of the seed, whereas the protein content was negatively correlated to the biological value and net protein utilization. These correlations followed the content of essential amino acids, lysine, threonine and methionine in the faba bean proteins. The content of the sulfur‐containing ammo acids cysteine and methionine are especially dominant factors for protein quality. A simple method for total sulphur determination was found nut 10 be a sufficiently reliable technique for evaluating the content of methionine and cysteine in the faba beans.Tannin, insoluble‐ and total dietary fibre are phenolic aromatic compounds which were negatively correlated with the faba bean quality as expressed by digestible energy, the protein digestibility and the biological value of the faba beans. The results obtained have also revealed, that it is not sufficient to consider tannin as a group in relation to the faba bean quality. We need to separate and evaluate the different types, of phenolics in relation to the variations in quality of faba beans. Some of the low molecular weight (LMW) phenolics in the faba beans seem 10 be involved in inhibitory effects on hydrolase enzymes, chymotrypsin and trypsin. Trypsin and chymotrypsin from different animals were different in their sensitivity to faba bean inhibitors and additional experiments are required to reveal details about these effects. It has also been revealed that fructosans Lind LMW carbohydrates (oligosaccharides) are important in relation to the quality of faba beans.