Genotype × environment influence on cowpea (Vigna unguiculata (L) walp) antinutritional factors: 1 – trypsin inhibitors,tannins, phytic acid and haemagglutinin

The relative influence of genotype, environment and genotype×environment effects on four antinutritional factors (g kg⁻¹) of importance in cowpea were studied using 15 local and improved cowpea genotypes grown in 12 environments, comprising three locations over four seasons per location. The locations Ago‐Iwoye (6°58′N4°00′E), Mokwa (9°17′N5°04E) and Kano (12°00′N8°31′E) were representatives of the major agroecological zones where cowpeas are produced. Genotypes effects were strongest in controlling trypsin inhibitor content, while the environment was the major source of variation for tannins, haemagglutinin and phytic acid contents. Thus, the variability in the levels of these antinutritional factors in cowpea seeds depends largely on the environment where they are grown. This implies that a cowpea genotype grown and consumed safely in an environment can be poisonous when grown and consumed in another environment. Genotype×environment effects were significant for tannins, haemagglutinins and trypsin inhibitor contents. Correlation coefficients (pooled data) from the three locations indicated that trypsin inhibitor was positively correlated to phytic acid (r=0.59, 0.001<P<0.05) and haemagglutinins (r=0.64, 0.001<P<0.05) but negatively correlated to tannin contents ( r=−0.79, 0.001<P<0.05). © 1999 Society of Chemical Industry     

Immunoaffinity chromatography purification and characterisation of pea trypsin inhibitors

Trypsin inhibitors from pea (Pisum sativum (L) cultivar Progreta grown in Denmark) have been isolated and shown to consist of at least nine pea proteinase inhibitors (PPI) with inhibitor activity toward both trypsin and chymotrypsin. The isoelectric points of the inhibitors were in the range 4.9–7.8. From this PPI mixture at least four inhibitors were isolated by immunoaflinity chromatography on a column containing immobilised monoclonal antibody (mAb) with inhibitor specificity. The PPI isolated by immunoaffinity chromatography were further separated by HPLC, and subsequent SDS-PAGE analysis showed molecular weights for four of the PPI in the range 11.3–14.2 kD. Their pI were determined by isoelectric focusing, mAbs were used for immunochemical characterisation and their amino acid composition showed a high (14.7–21%) content of cysteine. Tryptophan was not present in any of the isolated PPI. The data now obtained support the resemblance of PPI with inhibitors of the Bowman-Birk class and the differences in immunochemical properties of the various PPI indicate that pea has at least two gene loci coding for the inhibitors.     

Studies on the biological responses of rats to seed trypsin inhibitors using near-isogenic lines of Pisum sativum L (pea)

In order to determine the true antinutritional status of pea seed trypsin inhibitor (TI) proteins, pea lines are being produced that are near-isogenic except for the genetic locus, Tri, containing the TI structural genes. These lines are based on selection from the progeny of a cross between lines showing quantitative variation in TI content, as well as TI isoform and gene polymorphisms that serve as markers. Chemical analyses revealed that the composition of seeds from lines of each near-isogenic pair was extremely similar, except for a more than five-fold difference in TI content. Such lines provide material that is superior to the diverse lines previously used for nutritional assessment of pea TI. The specific biological effects of pea TI were studied by including the near-isogenic lines in standardised rat diets. The results indicated that TI content was correlated with a significant negative effect on protein digestibility and biological value. The difference in TI content of the pea seeds was reflected in the relative activity of pancreatic chymotrypsin whereas the activities of trypsin, lipase and amylase were less clearly affected. © 1999 Society of Chemical Industry     

Effect of year‐to‐year variation and genotype on trypsin inhibitor level in common bean (Phaseolus vulgaris L) seeds

Changes in trypsin inhibitor (TI) level in common bean (Phaseolus vulgaris L) seeds were investigated in relation to protein content, year‐to‐year variation and genotype. Twenty‐one local populations from the Basilicata region (southern Italy) were tested over 3 years (1995–97). The populations were cultivated in the same environment in which they were traditionally cultivated and are currently grown. A wide variation of TI content, expressed as units of inhibitor per milligram of dry matter (TIU mg^?1 DM), was found within the populations of the collection (x? ± 2σ: 27.67 ± 2.72 in 1995, 25.31 ± 2.82 in 1996, and 23.39 ± 2.14 in 1997). It was found that 16 populations showed a decrease of TI levels from 1995 to 1997; one population showed the opposite trend, in two populations the TI level remained unchanged, and in another two it reached its maximum in the intermediate year. A t‐test showed that only the 1995–97 variation is highly significant (p < 0.02). The increase of TI expression might possibly be related to the drought stress suffered by plants during the vegetative growth stage: rainfall received in 1995 during the growing season was lower (171 mm) than in 1996 (477 mm) and 1997 (388 mm); however, other abiotic or biotic factors cannot be excluded. It is suggested that the extent of TI variation contains a genetic component. Copyright © 2003 Society of Chemical Industry     

The effect of pea (Pisum sativum L.)‐originated asparaginase on acrylamide formation in certain bread types

The aim of this study was to investigate the effect of pea (Pisum sativum L.)‐originated asparaginase on acrylamide formation in white wheat, wheat bran and whole‐grain wheat breads. Two‐day germinated pea flour was used at 0%, 1%, 3% and 5% levels for each bread type. Acrylamide analysis was performed with liquid chromatography–mass spectrometry. Besides, colour and sensory properties of the breads were evaluated to search out the effects of pea flour substitution on the consumer acceptance. Reduction of acrylamide in white wheat bread was not found significant and addition of pea flour decreased the acceptance. However, it was found that acrylamide level can be reduced by 57% and 68% with addition of 5% pea flour in wheat bran and whole‐grain breads, respectively, without any negative impact on colour and sensory properties.     

Effect of genotype and pretreatment of field peas (Pisum sativum) on their dehulling and cooking quality

Field peas (Pisum sativum), an important pulse crop in Australia, are consumed as human food either as whole seeds or as splits after decortication. The yield of splits is an important economic factor for processors and the cooking quality is important for consumers. Effects of genotype, other physical characteristics and pretreatment in various solutions on both dehulling and cooking quality were studied for 23 known genotypes and are market sample of unknown genotype. Large variations were found for most characteristics. Seed size was positively correlated with yield of splits while husk content and broken seeds were negatively correlated with yield. Preconditioning seeds in salt solutions improved yield of splits. Variation in cooking quality among genotypes was reduced following splitting and cooking time was reduced by decortication and splitting and by presoaking in salt solutions (10 or 20 g kg⁻¹), particularly with sodium tripolyphosphate. Loss of seed material into cooking water was correlated with cooking time. No characteristic was found that could be used to predict cooking time. © 1998 SCI.     

Relationship between physicochemical and cooking properties,and effects of cooking on antinutrients,of yellow field peas (Pisum sativum)

The relationship between the physicochemical and cooking properties of yellow peas was examined in this study. A positive correlation was found between seed weight and water hydration capacity. The Peleg model, which was modified, could be used to describe the water absorption characteristics of peas and could be used to predict the rate of water absorption in the initial water absorption period. Cooking time could be measured objectively using the Mattson cooker. Cooking time was found to decrease with an increase in water hydration capacity. Hardness of cooked peas, measured using a texture analyser, was negatively correlated with both seed weight and water hydration capacity. Seed coats had a significant effect on water hydration and cooking quality of peas. Peas with relatively thin seed coats exhibited higher water hydration capacity, shorter cooking times and softer texture after cooking. The effects of soaking and cooking on trypsin inhibitor activity (TIA) and oligosaccharide levels in peas were also studied. Cooking was more effective than soaking in reducing TIA levels and oligosaccharides (raffinose, stachyose and verbascose) in peas. Copyright © 2003 Society of Chemical Industry     

Effect of dry heat on the in vitro digestibility and trypsin inhibitor activity of chickpea flour

The effect of a dry heat treatment on trypsin inhibitors, protein quality and molecular weight of products obtained from enzymatic hydrolysis of chickpea flours was analysed in order to use chickpeas as food protein hydrolysates. Chickpea flour obtained from dehusked seeds was processed under different conditions, either by heating or enzymatic treatment. Heat treatment at 140 °C for varying times (1–24 h) inactivated trypsin inhibitors and facilitated enzymatic treatment but showed an unacceptable loss in the nutritional quality of the protein for heating times longer than 6 h. Enzymatic treatment with a commercial protease, Alcalase 0.6L, at pH 8 and 50 °C, increased the protein nutritional value of the chickpea by breaking the protein chains into shorter peptide chains more suitable to human nutrition.     

Biological response of broiler chickens fed peas (Pisum sativum L.) expressing the bean (Phaseolus vulgaris L.) α‐amylase inhibitor transgene

The nutritive value of transgenic peas expressing an α‐amylase inhibitor (α‐Ai1) was evaluated with broiler chickens. The effects of feeding transgenic peas on the development of visceral organs associated with digestion and nutrient absorption were also examined. The chemical composition of the conventional and the transgenic peas used in this study were similar. In the two feeding trials, that were conducted normal and transgenic peas were incorporated into a maize–soybean diet at concentrations up to 500 g kg^?1. The diets were balanced to contain similar levels of apparent metabolisable energy (AME) and amino acids. In the first trial, the birds were fed the diets from 3 to 17 days post‐hatching and with levels of transgenic peas at 250 g kg^?1 or greater there was a significant reduction in body weight but an increase in feed intake resulting in deceased feed conversion efficiency. In the second trial, in which the birds were fed diets containing 300 g kg^?1 transgenic peas until 40 days of age, growth performance was significantly reduced. It was also demonstrated that the ileal starch digestibility coefficient (0.80 vs 0.42) was significantly reduced in the birds fed transgenic peas. Determination of AME and ileal digestibility of amino acids in 5‐week‐old broilers demonstrated a significant reduction in AME (12.12 vs 5.08 MJ kg^?1 DM) in the birds fed the transgenic peas. The AME value recorded for transgenic peas reflected the lower starch digestibility of this line. Ileal digestion of protein and amino acids was unaffected by treatment. Expression of α‐Ai1 in peas did not appear to affect bird health or the utilisation of dietary protein. However, the significant reduction in ileal digestion of starch in transgenic peas does reduce the utility of this feedstuff in monogastric diets where efficient energy utilisation is required. Copyright © 2006 Society of Chemical Industry     

Effect of genotype and environment on the concentrations of starch and protein in, and the physicochemical properties of starch from, field pea and fababean

BACKGROUND: The effects of genotype and environment and their interaction on the concentrations of starch and protein in, and the amylose content and thermal and pasting properties of starch from, pea and fababean are not well known. RESULTS: Differences due to genotype were observed in the concentrations of starch and protein in pea and fababean, in the onset temperature (To) and peak temperature (Tp) of gelatinization of fababean starch, and in the pasting, trough, cooling and final viscosities of pea starch and fababean starch. Significant two‐way interactions (location × genotype) were observed for the concentration of starch in fababean and the amylose content, To, endothermic enthalpy of gelatinization (ΔH) and trough viscosity of fababean starch. Significant three‐way interactions (location × year × genotype) were observed for the concentration of starch in pea and the pasting, trough, cooling and final viscosities of pea starch. CONCLUSION: Differences observed in the concentrations of starch and protein in pea and fababean were sufficient to be of practical significance to end‐users, but the relatively small differences in amylose content and physicochemical properties of starch from pea and fababean were not. Copyright © 2011 Society of Chemical Industry     

大豆中植酸和胰蛋白酶抑制剂的抗营养和抗癌效应

大豆是重要的油料作物和植物蛋白来源,富含植物化学物质,如植酸和胰蛋白酶抑制荆等。传统营养理论认为这些植物化学物质是抗营养因子,然而近来研究显示这些化学成分具有天然抗癌作用。以植酸和STI为例主要介绍了它们的结构、分布、生理活性、抗营养效应及抗癌效应。     

Starch but not protein digestibility is altered in pigs fed transgenic peas containing α‐amylase inhibitor

Eighteen individually housed boars were randomly allocated to one of three dietary treatments, an experimental wheat diet containing 989.4 g kg^?1 of a basal wheat diet, or this experimental wheat diet with 500 g kg^?1 of the basal wheat diet replaced with 500 g kg^?1 of either transgenic or non‐transgenic peas. The transgenic peas expressed the bean (Phaseolus vulgaris L.) α‐amylase inhibitor 1 gene. Diets contained n‐hexatriacontane (0.2 g kg^?1) as an indigestible marker to allow the determination of nutrient digestibility at the terminal ileum. Pigs were offered 1.6 kg day^?1 for 15 days, after which they were anaesthetised, the ileal and faecal digesta collected and the pigs subsequently euthanased. The ileal dry matter and starch digestibilities of the experimental wheat, non‐transgenic and transgenic pea diets were 78.3, 74.2 and 45.8% and 95.9, 95.2 and 42.4%, respectively. The apparent nutrient digestibilities of the non‐transgenic and transgenic peas were determined by difference. The ileal dry matter digestibility was significantly reduced in the transgenic peas compared with the non‐transgenic peas (12.7 and 69.9%, respectively; P = 0.006), which was largely due to a reduced starch digestibility. The apparent crude protein digestibilities of the transgenic peas were similar to the non‐transgenic, being 79.7 and 78.5%, respectively. The amino acid digestibilities of the transgenic and non‐transgenic peas were also similar. Copyright © 2006 Society of Chemical Industry     

An Objective Dry Pea ‘Colour’ Scoring System for Commercial and Plant Breeding Applications

For human consumption, colour of dry peas is an important quality criterion, presently assessed subjectively. Samples which are dark and even in colour are preferred to those which are bleached and variable. The capacity of image analysis methods to provide an objective colour scoring system was investigated, and useful predictive relationships were found between subjective rankings for colour allotted to members of a large set of training samples, and ranking for median optical density, optical density proportional range and median seed size. Training samples were drawn at random from the pea sample population, and were not evenly distributed, so spacings between rankings were corrected by reference to the variance of the mean ranking for a sample. This correction allowed the predictive variables to explain 64% of the observed variation in the corrected rank. When the required precision of a derived scoring system was set to ±1·0 score points, it was found that the set of training samples could be covered by 6·95 score points, so a scoring calibration equation was developed in which increasing colour loss caused a higher score. The darkest sample in the training set was arbitrarily chosen to have a score of 5, to allow for the possibility of intrinsically darker, less bleached samples. This scoring system is at least twice as precise as the current subjective approach, and can be applied using any image analysis system that can be calibrated against cheap, highly consistent, photographic standard grey cards. The performance of the system may be improved further if chrominance variation can be taken into account. © 1997 SCI.     

The apparent ileal digestibility,determined with young broilers,of amino acids in near‐isogenic lines of peas (Pisum sativum L) differing in trypsin inhibitor activity

Two pairs of near‐isogenic lines of peas were developed to have high and low concentrations of seed trypsin inhibitor. HA5 and LA5 contained 8.73 ± 0.19 and 1.45 ± 0.19 trypsin inhibitor units per mg dry weight respectively, whereas HB5 and LB5 contained 7.40 ± 0.65 and 1.78 ± 0.15 trypsin inhibitor units per mg dry weight respectively. The four pea samples were included in semi‐synthetic diets and evaluated in a poultry metabolism experiment, using birds aged 3 weeks, to determine the coefficient of apparent ileal amino acid digestibility (CAID). There was a significant difference in CAID for the amino acids reported that correlated with pea genotype. Thus, for example, CAID for cystine was 0.738 ± 0.0196 and 0.812 ± 0.0128 for HA5 and LA5 respectively and 0.721 ± 0.0139 and 0.804 ± 0.0178 for HB5 and LB5 respectively. Corresponding data for methionine, the other main nutritionally deficient amino acid in peas, were 0.887 ± 0.0103, 0.930 ± 0.0089, 0.885 ± 0.0083 and 0.929 ± 0.0125 respectively. Other amino acids followed similar patterns. These data provide information on the nutritional consequences of variation in trypsin inhibitor content of pea seeds in near‐isogenic backgrounds. Plant breeding for low trypsin inhibitor activity in seeds accordingly offers potential for improving nutritional value. © 2003 Society of Chemical Industry     

Mathematical modelling of the heat inactivation of trypsin inhibitors in soymilk at 121–154 °C

Response surface methodology (RSM) was used to investigate the effects of processing temperature and time on the inactivation of trypsin inhibitors (TI) in soymilk. The factorial experimental design consisted of four levels of temperature and six levels of time in a temperature range of 121–154 °C and a time interval of 10–90 s. A quadratic polynomial equation, relating log(% TI retained) as a function of heating time and temperature, was satisfactorily fitted to the experimental data by least squares regression with r² (correlationcoefficient) = 0.959. Within the range of heating times investigated, TI in soymilk was satisfactorily destroyed to 10% retained at 143 and 154 °C with 62 and 29 s heating time respectively. © 2002 Society of Chemical Industry     

A Genetic Approach to Studying the Morphology,Structure and Function of Starch Granules using Pea as a Model

A series of near‐isogenic lines have been developed for mutations induced and identified at six independent loci controlling steps in starch biosynthesis of pea (Pisum sativum) seeds. The review describes the methodology associated with the development of these lines and with the characterisation of the starches produced, in particular the effects that the mutations have on the morphology, structure and properties of the starch granules. The shape of starch granules from lines containing mutations at either the r or rug5 loci, encoding a starch branching enzyme and a soluble starch synthase, differ significantly from granules found in the non‐mutant parental line. Mutations at each of the six loci result in characteristic changes in granular structure, which has been studied using polarised light microscopy, wide angle X‐ray diffraction, solid state NMR and differential scanning calorimetry. Relatively small but significant changes were found in the proportion of crystalline material within the granules and much greater differences in the spatial distribution of the crystals within the granules. In particular, granules from r and rug5 mutants do not show the characteristic ‘Maltese cross’ pattern when viewed under polarised light. Large differences were found between the mutants in the proportion of A‐ and B‐type crystallites within the granules, ranging from about 60% A‐type in rug3 mutants to 100% B‐type in r mutant lines. It is suggested that the pea lines are an invaluable resource for answering fundamental questions on the genetic control of starch granule structure and function.     

Preparation of trypsin‐immobilised chitosan beads and their application to the purification of soybean trypsin inhibitor

BACKGROUND: Trypsin inhibitors are among the most important antinutritional factors in legumes. Recent research has shown that soybean trypsin inhibitor (SBTI) exhibits multiple bioactivities, but very few studies on the purification of SBTI are available. Enzymes are commonly used as biospecific ligands in affinity purification of their substrates or inhibitors. The aim of the present study was to prepare trypsin (EC 3.4.21.4)‐immobilised chitosan beads and use them to purify trypsin inhibitor from soybean whey. RESULTS: Compared with free trypsin, the immobilised trypsin had higher thermal and pH stability. The adsorption ratio of SBTI from crude SBTI aqueous solution by trypsin‐immobilised chitosan beads was 33.3%. The purified SBTI obtained by affinity chromatography was characterised by sodium dodecyl sulfate polyacrylamide gel electrophoresis as a single polypeptide band with an M_r of 8.3 kDa belonging to the Bowman–Birk family. CONCLUSION: Trypsin‐immobilised chitosan beads were effectively used in the affinity separation of trypsin inhibitor from soybean seeds, thus indicating that immobilised trypsin may have practical application in the soybean‐processing industry. The results of this study provide a background for further investigation of potential applications of soybean bioactive constituents in the areas of agriculture and food. Copyright © 2008 Society of Chemical Industry     

Evaluation of the seeds of selected lines of three Lathyrus spp for β-N-oxalylamino-L- alanine (BOAA), tannins,trypsin inhibitor activity and certain in-vitro characteristics

Sixty eight lines comprising 36 Lathyrus sativus, 16 L cicera and 16 of L ochrus were analysed for crude protein (CP), the neurotoxin β-N-oxalylamino- L-alanine (BOAA), protein precipitable tannin (PPT), catechin equivalents (CE), and trypsin inhibitor activity (TIA). Randomly chosen lines from each of the species were also evaluated for other chemical and in-vitro characteristics including neutral detergent fibre (NDF), acid detergent fibre (ADF), in-vitro dry matter and in-vitro organic matter digestibilities (IVDMD, IVOMD), respectively. The species with the lowest and highest CP were L cicera(295.39 ± 18.69 g kg^?1 DM) and L sativus(324.99 ± 13.40 g kg^?1 DM), respectively. BOAA concentration, also, was least in L cicera (1.26 ± 0.18 g kg^?1 DM) and highest in L sativus(5.63 ± 0.43 g kg^?1 DM). There were higher intraspecies variations in BOAA levels among L sativus and L cicera than in L ochus genotypes. On average, BOAA levels in L ochrus and L sativus were about four to five times higher than those in L cicera. The PPT level in L sativus(4.54 ± 0.31 g kg^?1 DM) was similar to that in the other species. However, there were some lines without detectable PPT in all the species. CE, which detects simple flavonoids as well as condensed tannins, was no more than 6.10 g kg^?1DM in any accession, except in L ochrus in which CE ranged from 14.07 g kg^?1 to 28.50 g kg^?1 DM. There were high variabilities between and within species in CE values as shown by the high coefficients of variation. Mean TIA in all the L sativus lines (18.16 ± 2.38 g kg^?1 DM) was similar to that in L ochrus and about twice the levels in L cicera. The coefficients of variation ranged from 10.93% in L ochrus to 17.13% in L sativus. The ash, organic matter and NDF content of the lines did not differ significantly. However, the ADF, IVDMD and IVOMD differed significantly (P < 0.05) between the species. The IVDMD and IVOMD of L cicera (88.55 ± 0.80% and 85.53 ± 0.85%, respectively) were significantly lower (P < 0.05) than in the other species. Seed weight correlated significantly (P < 0.05) with CE and TIA in both L sativus and L ochrus while CP correlated significatly (P < 0.01) with TIA in both L sativus and L cicera with respective r values of 0.70 and ?0.75.