Trypsin inhibitor in moth bean: thermal stability and changes during germination and cooking

Trypsin inhibitor from moth bean was studied for thermal stability and changes during germination and cooking. The application of dry heat did not inactivate the inhibitor. However, autoclaving at 120°C at 15lbs pressure destroyed inhibitor activity completely. The extracted inhibitor lost 70% activity in 60 min when incubated at 100°C. Soaking of moth bean seeds for 8 h decreased trypsin inhibitor activity by 20%. The germination of seeds for 24 h resulted in 70% reduction in inhibitor activity. No activity was detected in 48 h germinated seeds. Germination (for 24 h) followed by cooking of moth bean seeds destroyed the trypsin inhibitor completely.     

The effect of slow-cooking on the trypsin inhibitor and hemagglutinating activities and in vitro digestibility of brown beans (Phaseolus vulgaris,var.Stella) and kidney beans (Phaseolus vulgaris,var.Montcalm)

Brown beans and kidney beans were subjected to two modes of cooking in a household slow cooker: (A), a fixed low setting for 10 h, and (B), a high setting for 2.5 h and a low setting for 7.5 h. Temperature changes in the beans were recorded. With treatment A over 90% of the hemagglutinating and trypsin inhibitor activities occurred after 6 h at which time the temperature had reached 80°C. With treatment B inactivation of these activities was almost complete at the end of 2 h when a maximum temperature of 100°C had been attained. The in vitro digestibility of the bean protein was considerably increased by either treatment. By way of contrast, only 20 min of heating was required to destroy these activities when the beans were brought to a boil in an open vessel.     

Comparative studies on the storage proteins and anti-nutritional factors from seeds ofPsophocarpus tetragonolobus (L.) DC from five South-East Asian countries

The seeds of 27 varieties ofP. tetragonolobus from six regions of South-East Asia have been examined by polyacrylamide gel electrophoresis. No marked variations in the electrophoretic patterns were found which could be exploited by plant breeders to improve nutritional quality with respect to sulfur containing amino acids. The amino acid compositions of varieties from the different regions showed little variation. Seed extracts of all 27 varieties ofP. tetragonolobus showed trypsin and chymotrypsin inhibitory activities, and hemagglutinating activities. The levels of trypsin inhibitory activities showed some genotypic differences varying from 22.2 to 42.5 mg trypsin inbibited g^?1 of seed meal. As a group the varieties from Malaysia showed the lowest levels of trypsin inhibitor. The chymotrypsin inhibitory activity also showed a similar variation (30.1–47.6 mg chymotrypsin inhibited g^?1 of seed meal). Seed extracts agglutinated type O⁺ and B⁺ human and rabbit erythrocytes with little difference in activity between varieties. Autoclaving winged bean seed meal prior to protein extraction inactivated the anti-nutritional factors and resulted in considerable reduction of protein solubility.     

Nutritional studies on rats and fish (carp Cyprinus carpio) fed diets containing unheated and heated Jatropha curcas meal of a non-toxic provenance

Unheated and heated (121 ^°C, 66% moisture; 15, 30 and 45 min) Jatropha meals of non-toxic provenance from Veracruz state in Mexico were evaluated using rats and fish. With rats, the weight gain was highest for the casein diet followed by heated (30 min; only this treatment was studied using rats) and unheated Jatropha meal containing diets. The protein efficiency ratio (PER) for unheated and heated Jatropha meal containing diets was 37 and 86%, respectively, of the casein diet. On the other hand, the body weight gain, PER and feed conversion ratio of fish were statistically similar for unheated and heated (15, 30 and 45 min) Jatropha meal containing diets fed for a period of 35 days. Although these parameters were statistically similar for the unheated and heated Jatropha meal containing diets, the body weight gain, PER and protein productive value were highest and the feed conversion ratio lowest with 15 min heated Jatropha meal, suggesting that the heat treatment for 15 min is optimal for the meal. Trypsin inhibitor and lectin activities decreased drastically (>83 and 99%, respectively) after 30 and 45 min of heat treatment and after 15 min, the residual lectin activity was negligible and the residual trypsin inhibitor activity was 34%. These results, together with the nutritional parameters investigated, imply that Jatropha trypsin inhibitors and lectins do not have any adverse effects on carp at least up to 35 days of feeding. The nutritional value of Jatropha meal of the non-toxic provenance is high, and potential exists for its incorporation into the diets of monogastrics, fish and possibly humans.     

Occurrence and Stabilities of Oat Trypsin and Chymotrypsin Inhibitors

Nine chymotrypsin and four trypsin inhibitors have been extracted and separated from ungerminated oat grains. The inhibitors fell into two groups, based on their heat and pH stabilities. Members of the most abundant group are labile and are inactivated at 80 °C or at pHs of 3·3 or lower. Members of the second group are stable and are resistant to boiling for 30 min. On germination, the labile inhibitors are inactivated after 2 days and the stable chymotrypsin inhibitors after 3 days. Most of the labile inhibitors from ungerminated grain are destroyed when incubated at 20 °C for 20 h but addition of PMSF, a serine protease inhibitor, prevented their inactivation. Labile inhibitors in extracts of ungerminated oats are inactivated on incubation with an extract prepared from germinated oats, but not in the presence of PMSF. Most oat chymotrypsin and trypsin inhibitors are heat labile and pH sensitive. These inhibitors are apparently inactivated by serine proteinase(s) already present in ungerminated grain.     

Assay of some nutritional and antinutritional factors in different cultivars of winged bean (Psophocarpus tetragonolobus (L.) DC) seeds

Seed of 34 cultivars of winged bean of different origin were evaluated for their nutritional composition and the antinutritional factor, the trypsin inhibitor. The values ranged from 14.1–17.6, 6.12–8.73, 21.94–34.89 and 30.86–39.05% for their fat, fibre, carbohydrate and protein contents, respectively. The protein content in defatted seed meal ranged from 33.43–47.25%, whereas the trypsin inhibitor activity (TIA) registered a wide variation from 63–123 mg g^?1 with seed coat and 51–98 mg g^?1 without seed coat. The strain selection-12 and NBRI selection contained the minimum TIA (63 mg g^?1), while the highest value (123 mg g^?1) was in variety V7, a 2-fold higher activity.     

Protein quality of chickpea (Cicer arietenum) with different grades ofCallosobruchus chinensis L. infestation

Dry heat treatments at 140 °C for 20 min or 160 °C for 10 min of peanut kernels significantly improved protein solubility, water and oil absorptions, foaming capacity and stability, and least gelation concentration of the defatted meal. However, heat treatments at 160 °C for 30 min or more than 30 min period resulted in significant decrease in these properties. Such heat treatments significantly decreased the level of albumin and globulin fraction with concomitant increase in glutelin fraction in the meal.     

Effect of some variables on the extractability of proteins,urease activity,free alpha-amino groups and soluble carbohydrates from soybean meals

The effect of several variables on the extraction of soybean meal protein has been investigated. Solubility studies of the meal show that globulin is the main protein component. Overnight suspension and a meal to solvent ratio 1:10 (w/v) were found to extract maximum amount of protein in distilled water adjusted to pH 6.4 (91.07%). More protein was extracted by diluted sodium hydroxide solutions (0.02 M) adjusted to various alkaline pH-values. Results with several other salts show that concentration as well as pH are very important factors in controlling the percentage of protein extracted. The optimum temperature of extraction was 70°C for 30 minutes, although over a temperature range of 60–75°C the extraction remains essentially constant. Destruction of urease activity was achieved by heating the suspended meal at 75°C for 45 Min. The kinetic inactivation of the enzyme was studied and the energy of inactivation was found to be 32.575 K cal. The level of free alpha amino groups in soybean meals ranged from 5.50 grams: 16 gram nitrogen in meal extracted with 0.1 N HCL adjusted to pH 5.0 to 1.71/100 grams protein (in meal extracted with distilled water.). Their level correlated significantly (p=0.01) upon heat treatment with the carbohydrate content of the corresponding meal. Contrary tothe protein solubility pattern, the -NH₂ groups decrease considerably upon longer exposure to temperatures between 60–75°C.     

Growth and Nutrient Accumulation of Winged Bean and Velvet Bean as Cover Crops in a Subtropical Region

We examined biomass dry matter and nutrient uptake of live plant parts, leaf area index, and litter of winged bean (Psophocarpus tetragonolobus) and velvet bean (Mucuna pruriens) 12, 18, 24 and 30 weeks after sowing (WAS). The two plants had similar leaf and stem+petiole biomasses. At 30 WAS winged bean had a significantly lower pod yield than velvet bean. Between 18 and 30 WAS, winged bean produced less litter than velvet bean due to differences in growth stages. The total mulch of live parts and litter of winged bean and velvet bean completely covered the ground by 18 and 12 WAS, respectively. Compared to velvet bean, the leaf and stem+petiole of winged bean had a significantly higher N concentration; significantly higher N uptake at 24 and 30 WAS; significantly lower C/N ratio; and significantly higher P, K and Mg concentrations. In winged bean, P uptake was significantly higher in the leaf at 30 WAS and in the stem+petiole at all harvesting times. The total biomass of the leaf, stem+petiole and litter of winged bean was 317–561 g DM m-², and their N content was 12.3–17.7 g m-². The total biomass of live parts and litter of winged bean might be sufficient to suppress weeds and increase soil N. Winged bean is an appropriate legume cover crop and green manure due to its longer growing period and superior ground-covering ability and high N input.     

Inactivation of trypsin inhibitors in sweet potato and taro tubers during processing

In order to understand the extent of elimination of trypsininhibitors during processing of sweet potato (Ipomoea batatas) andtaro (Colocasia esculenta) tubers, a detailed study was conductedusing tubers processed by oven drying, cooking, and microwavebaking. Between 80 and 90% trypsin inhibitor (TI) activity wasretained in sweet potato chips up to 2h at 70 ^°C. Among thefour cultivars of sweet potatoes, RS-III-2 trypsin inhibitors were more heatlabile. Heating at 100 ^°C led to rapid inactivation of TI of sweetpotatoes. Varietal differences in thermal stability were more pronouncedfor the trypsin inhibitors of taro than sweet potatoes. Taro inhibitors werealso more rapidly inactivated than sweet potato TI. Between 17 and31% TI activity was retained in cooked tuber pieces of sweet potatoes,while only 3–10% were retained in taro cultivars. Very effectiveinactivation of trypsin inhibitors of sweet potatoes and taro could beobtained through microwave baking. Flour prepared from taro was devoidof TI activity, while 5–12% TI activity was retained in the flour preparedfrom sweet potatoes. The study clearly established that among the fourtechniques used, microwave baking and flour preparation were the best methods to eliminate TI from sweet potatoes and taro.     

Effect of heat treatments on trypsin/chyomotrypsin inhibitor activity of red gram (Cajanus cajan L.)

The application of dry heat to the seeds and meal was not effective in inactivating the trypsin inhibitory activity (TIA) and chymotrypsin inhibitory activity (CIA). Soaking for 24 hours followed by cooking for 20 min was effective in destroying the TIA and CIA.     

Protein quality of raw and autoclaved cowpeas: Comparison between some insect resistant and susceptible varieties

A comparative study was conducted with four improved cowpea (2 pod bug resistant and 2 susceptible) varieties to assess the effect of autoclaving at 105°C under 15 psi for 30 min on their protein quality, as measured by the levels of some antinutritional factors and performance of rats fed the different cowpea diets. Also, the relationships between antinutrients in the cowpeas and protein quality indicators were examined. Results showed that autoclaving completely eliminated trypsin inhibitor, hemagglutinin and HCN, whereas it reduced significantly (p<0.01) the levels of phytic and tannic acids in the cowpeas. In both raw and autoclaved forms, the insect susceptible cowpeas were better (P<0.05) utilized than their resistant counterparts, a condition which was attributable to lower levels of antinutrients in the susceptible varieties. In the raw cowpeas, trypsin inhibitor, hemagglutinin and HCN were significant variables affecting cowpea protein utilization, while in autoclaved samples, tannic and phytic acids were important though nonsignificant (P>0.05) factors. It was concluded that autoclaving at the above temperature and time improved the protein quality of the insect susceptible more than the insect resistant cowpea varieties.     

Effect ofCallosobruchus chinensis (Bruchid) infestation on antinutritional factors in stored legumes

Effect of bruchid infestation on selected antinutritional factors, saponin, phytic acid and trypsin inhibitor activities, of three (mung bean, chickpea and pigeon pea) legumes stored for six months was studied. The three antinutritional factors, saponin, phytic acid and trypsin inhibitor activity (TIA), increased with increased storage period. Saponin, phytic acid and trypsin inhibitor activity values were significantly lower in control samples compared to stored infested samples. The difference between the contents of antinutritional factors were significant (p<0.05) during different storage periods.     

Soaking and cooking parameters of tepary beans: effects of cooking time and cooking temperature on hardness and activity of nutritional antagonists

Tepary samples were examined for patterns of hydration, dry matter losses during the processes of soaking and cooking, residual hardness in partially cooked samples and heat lability of endogenous proteinaceous antinutritional factors. At 24 °C, teparies imbibed water equivalent to their weight (100% hydration) in 4 h and continued to absorb water rapidly for an additional 4 h before reaching an equilibrium hydration. During the processes of soaking and cooking, materials leached from raw beans represented 7.3 and 13.5% of their dry weight, 4.3 and 12.4% of their protein content, 7.1 and 12.2% of their stored carbohydrate and 22.4 and 33.4% of their mineral levels, respectively. In samples prepared at different cooking times (60, 90, 120, 150, 180 min) and cooking temperatures (80, 85, 90, 95°C), longer times and higher temperatures resulted in greater reductions in residual bean hardness; interactive effects of time and temperature treatments were significant. Residual activity of trypsin and chymotrypsin inhibitors in partially-cooked samples appeared to be negligible. In addition, at least 80% of the original hemaglutinating activity of lectins in raw beans was lost during partial-cooking of samples under all cooking regimes.     

Supercritical carbon dioxide (SC-CO2) extraction of Nigella sativa L. oil using full factorial design

In this study, Nigella sativa L. oil was extracted using supercritical carbon dioxide with full factorial design to determine the best extraction condition (pressure, temperature and dynamic extraction time) for obtaining an extract with high yield, antioxidant activity and thymoquinone (TQ) quantity. The maximum thymoquinone content in the highest overall yield was achieved through SC-CO₂ extraction condition of 150 bar, 40 °C, 120 min with the value of 4.09 mg/ml. The highest SC-CO₂ extraction yield was 23.20% which obtained through extraction condition of 350 bar, 60 °C and 120 min. The extraction conducted at 350 bar, 50 °C, 60 min showed the lowest IC₅₀ value (highest antioxidant activity) of 2.59 mg/ml using DPPH radical scavenging activity method. Fatty acid composition of the extracted oil with highest radical scavenging activity was obtained by gas chromatographic analysis.     

Screening soybean (grain and vegetable) genotypes for nutrients and anti-nutritional factors

Fifty six genotypes of grain-type soybean and 17 genotypes of vegetable-type soybean collections were analyzed for protein and oil content, trypsin inhibitor, and lipoxygenase activities. The protein and oil content ranged from 36.9 to 47.9% and from 13.3 to 23.0% for different accessions in grain- and vegetable-type soybeans, respectively. Trypsin inhibitor and lipoxygenase activities ranged from 22.0 to 47.0 trypsin inhibitor units/mg meal and from 482 to 6265 lipoxygenase units/min/mg meal for grain- and vegetable-type soybeans, respectively. Significant correlations (r=–0.62 and –0.52,P<0.05) were found between protein and oil, and between protein and trypsin inhibitor. A significant positive correlation (r=0.42,P<0.05) was also calculated for oil and lipoxygenase activity. Several genotypes of soybean and vegetable soybean (plant introductions 423905, 417330, 417223, 171451, 200506, 200523, 417124, 227687, 203402, 445842, 203399, 423852, 416771, FC31927, Avoyelles, and Sooty) showed good nutritional potential and may be useful in a breeding program to improve the nutritional quality of soybean. Screening for essential amino acids, fatty acids, and trace minerals for selected genotypes is underway.     

Effects of heat treatments and germination on trypsin inhibitor activity and polyphenols in jack bean (Canavalia ensiformis L. DC)

The application of dry heat to seeds and meal was not effective in inactivating the TI and reducing the polyphenol content. Soaking for 24 h followed by cooking for 20 min, was equally effective in destroying the TI activity. Germination of jack bean seeds for 40 h decreased the levels of TI and polyphenols by 31% and 35%, respectively.     

An evaluation of the effect of Bacillus cells and Bacillus spores in association with cowpea granules as starter cultures for the fermentation of African oil bean (Pentaclethra macrophylla Bentham) to `ugba'

Studies on the improvement of the traditional production of `ugba',a protein-rich fermented African oil bean seed product, were undertaken,by developing starter cultures of Bacillus subtilis cells andspores in association with cowpea granules. The viability of the cellsin association remained stable at 94.5% for 6 months at 30 <sup>°</sup>C and for up to 10 months at 4 <sup>°</sup>C while the viability of the sporesin association remained stable at ca. 96% for up to 10 months at both4 and 30 <sup>°</sup>C. The starter cultures resulted in high increases inprotease activity from ca 2.8 mg N/min to about 51.6 ± 0.4 mg N/minin 48 h and a corresponding increase in amino-nitrogen content of ca2.0 ± 0.2 mg N 100 g dry matter (DM) to ca 18.5 ± 0.3 mg N/100g (DM) during the same period. Changes in the protease activity of thenatural process were gradual and increased from 3.0 mg N/min to 38.0 ±0.8 mg N/min after 5 days of fermentation. The maximum amino nitrogencontent of `ugba' produced by the starter cultures (18.5 ± 0.3 mgN/100 g DM) after 2 days was significantly (p < 0.05) higher than themaximum amino nitrogen content (12.5 ± 0.8 mg N/100 g DM), of `ugba'obtained by the natural process. `Ugba' produced by the starter cultureswere well accepted and compared favorably with the natural product.     

Effects of seed treatments and storage on the changes in lipids of pearl millet meal

Lipids in pearl millet meal showed a rapid hydrolytic decomposition during storage. The magnitude of such degradation was influenced significantly by the nature of the storage container used, the temperature and heat treatments given to the seeds. The hydrolytic breakdown of lipids was significantly low in the meals stored in polyethylene bags, plastic boxes and under refrigerated (5±2 °C) conditions. Hot water blanching at 98 °C for 10 sec and dry heating of seeds at 100 °C for 120 min were found to be most effective in minimising the undesirable changes in lipids of the meal during storage.     

Edible provenances of Jatropha curcas from Quintana Roo state of Mexico and effect of roasting on antinutrient and toxic factors in seeds

Seven seed samples of J. curcas, both in raw and roasted state, sold in some villages in Quintana Roo state, Mexico for human consumption were analyzed for physical characteristics, nutrients and antinutrients. The average seed weight varied from 0.53 to 0.74 g and kernel weight as proportion of raw seed weight was from 61 to 66%. The contents of crude protein, lipid and ash of kernels from raw seeds were 27–30%, 55–62% and 3.7–5.2% respectively. The levels of antinutrients in meal from the raw seeds were: trypsin inhibitor activity (14.6–28.7 mg trypsin inhibited/g), lectin (25.6–52.2 unit; one unit is the reverse of minimum amount of mg meal/ml assay which produced haemagglutination), saponins (1.9–2.3% as diosgenin equivalent) and phytate (8.4–10%). Phorbol esters in kernels from raw seeds were not detected in four samples and in other three samples it ranged from 0.01 to 0.02 mg/g as phorbol-12-myristate 13-acetate equivalent. Roasting of seeds inactivated almost 100% of trypsin inhibitor activity. Although lectin activity reduced on roasting, it was still present in high amounts. Saponins, phytate and phorbol esters were not affected by roasting.