A fast and cost-effective procedure for reliable measurement of trypsin inhibitor activity in soy and soy products.

Rapid and accurate measurement of trypsin inhibitor is critical for soy processors to assess the quality of soy meal. Currently, trypsin inhibitor activity is measured using the American Oil Chemists' Society (AOCS) and the American Association of Cereal Chemists International (AACCI) approved method. We have modified and improved the AACCI/AOCS approved method resulting in the elimination of several time-consuming steps and drastically reducing the assay volume. By employing our simplified procedure, we have measured trypsin inhibitor activity of several soy and soy products. A side-by side comparison of our simplified procedure with AOCS approved method revealed strikingly similar results indicating that several time-consuming and tedious steps associated with AACCI/AOCS approved methods can be eliminated without sacrificing the accuracy of the assay. Moreover, we demonstrate that our assay can also be carried out in 96-well microplates which will enable high-throughput screening of large number of soy meal samples.

Quality control of hospital preparations: Establishment of a simple and rapid method for quantifying ulinastatin in vaginal suppositories.

Ulinastatin vaginal suppositories, used to prevent threatened premature delivery, are frequently used in hospitals. However, there is no established method for quantifying ulinastatin contained in suppositories. Therefore, we investigated a simple and efficient method for quantifying ulinastatin contained in suppositories. Our analytical method involved removal of the base; optimising the enzyme inhibition reaction time and enzyme reaction time; and measuring the absorbance. The modified method was reproducible, operation time was significantly shortened, and cost was reduced to approximately 1/17 of that of the previously reported method. This simple and rapid quantitative method could contribute to the improvement of quality control of ulinastatin vaginal suppositories as an extemporaneous hospital preparation.

Total proteolytic activity and concentration of alpha-1 antitrypsin in meconium for assessment of the protease/antiprotease balance.

BACKGROUND: During intrauterine life, various proteolytic enzymes and their main inhibitor, alpha-1 antitrypsin, accumulate naturally in meconium. A protease/antiprotease balance is required to maintain the biological stability of the environment in which the fetus develops. METHODS: The pool of active proteases was determined using the EnzChek Protease Assay Kit. The concentration of alpha-1 antitrypsin in meconium was measured by enzyme-linked immunosorbent assay. Serial portions of meconium (n=80) were collected from healthy full-term neonates (n=19). RESULTS: Mean concentrations of active proteases and alpha-1 antitrypsin were 1.55 [standard deviation (SD) 1.3]mgg-1 (range 0.15-6.17) and 3.72 (SD 1.78)mgg-1 (range 0.76-8.55), respectively, with significant correlation (Rs=0.32, p=0.004). A significant increase in the concentration of active proteases was found between the first and last meconium portions (p<0.05). The proteases in the last meconium portions had a higher reaction velocity and affinity for the substrate than the proteases in the first meconium portions. The active protease:alpha-1 antitrypsin ratio was <0.5 in all first meconium portions, but was higher in the last meconium portions. CONCLUSIONS: Strong correlation between the concentrations of active proteases and alpha-1 antitrypsin in meconium may indicate their mutual interaction in the intrauterine environment. Alpha-1 antitrypsin maintains the protease/antiprotease balance during fetal development.

Chemical characterisation of kernels, kernel meals and oils from Jatropha cordata and Jatropha cardiophylla seeds.

BACKGROUND: Jatropha cordata and Jatropha cardiophylla are native to northwestern Mexico and are adapted to arid and semi-arid conditions (<500 mm of precipitation and temperatures from 8 to 45 °C). The aim of this study was to evaluate the chemical composition of J. cordata and J. cardiophylla kernels and oils as well as antinutrients in the defatted kernel meals of these species. RESULTS: Kernels of J. cordata and J. cardiophylla seeds analysed in this study were rich in crude protein (283 and 289 g kg(-1) respectively) and lipid (517 and 537 g kg(-1) respectively). The main fatty acids in J. cordata and J. cardiophylla oils were linoleic and oleic acids. High levels of trypsin inhibitor and phytates and low levels of saponins were present in the meals. The phorbol ester contents in J. cordata and J. cardiophylla kernel meals were 2.73 and 1.46 mg g(-1) respectively. CONCLUSION: For both J. cordata and J. cardiophylla it could be inferred that (a) the oil and kernel meal were toxic and the kernel meal could be used as livestock feed only after detoxification, (b) the oil could be used for non-alimentary purposes, i.e. biodiesel production, and (c) the seed or oil could be used for isolating various bioactive compounds for pharmaceutical and agricultural applications.

Optimization of process for reduction of antinutritional factors in edible cereal brans.

Reduction of various antinutritional factors in cereal brans by different treatments (microwave heating, dry heating and wet heating) were studied. There was significant difference (p ≤ 0.05) in reduction of antinutritional factors of treated cereal brans except for dry heating at low temperature. Microwave heating at 2450 MHz for 2.5 min resulted in 53.85%, 57.21%, 65.00% and 100% reduction in phytic acid, polyphenols, oxalates and saponins, respectively. Wet heating resulted in maximum reduction in trypsin inhibitor activity (83.07%) at 110 °C for 25 min. Processing treatment resulted in increase in bulk density and slight darkening of the brans. The most effective method of detoxifying most of the toxicants was microwave heating for 2.5 min, and therefore it could be exploited for making treated brans an ideal source for potential food application.

Experimental assessment of toxic phytochemicals in Jatropha curcas: oil, cake, bio-diesel and glycerol.

BACKGROUND: Jatropha curcas seed is a rich source of oil; however, it can not be utilised for nutritional purposes due to presence of toxic and anti-nutritive compounds. The main objective of the present study was to quantify the toxic phytochemicals present in Indian J. curcas (oil, cake, bio-diesel and glycerol). RESULTS: The amount of phorbol esters is greater in solvent extracted oil (2.8 g kg⁻¹) than in expeller oil (2.1 g kg⁻¹). Liquid chromatography-mass spectroscopy analysis of the purified compound from an active extract of oil confirmed the presence of phorbol esters. Similarly, the phorbol esters content is greater in solvent extracted cake (1.1 g kg⁻¹) than in cake after being expelled (0.8 g kg⁻¹). The phytate and trypsin inhibitory activity of the cake was found to be 98 g kg⁻¹ and 8347 TIU g⁻¹ of cake, respectively. Identification of curcin was achieved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the concentration of curcin was 0.95 g L⁻¹ of crude concentrate obtained from cake. CONCLUSION: Higher amounts of phorbol esters are present in oil than cake but bio-diesel and glycerol are free of phorbol esters. The other anti-nutritional components such as trypsin inhibitors, phytates and curcin are present in cake, so the cake should be detoxified before being used for animal feed.

Assessment of cyanogenic potential, nitrate and nitrite contents, and trypsin inhibitor activity of some Nigerian legumes.

Edible seeds of seven varieties of legumes commonly consumed by Nigerians in large quantities were evaluated for total protein, cyanogens, nitrate and nitrite contents, and trypsin inhibitor activity using chemical, biochemical, enzymatic, and spectrophotometric methods. All analyzed samples contained cyanogen and nitrate with levels ranging from 5.88 +/- 0.26 to 28.55 +/- 1.32 mg/100 g of DM and from 49.64 +/- 4.60 to 239.42 +/- 7.20 mg/100 g of DM, respectively. Only three of the varieties contained detectable levels of nitrite, which varied from 0.54 +/- 0.01 to 3.19 +/- 0.2 mg/100 g of DM. Trypsin inhibitor activity was detected in all of the samples, ranging from 7.75 to 100.75 micromol/min/mg of protein. Total protein content of the legumes ranged from 17.8 to 28% on a dry weight basis. There was a positive correlation (r = 0.71) between the cyanogenic potential and the nitrate content of the dry seeds. Processing reduced about 78.6-88.8% and 71.0-89.5% of total cyanogen and nitrate contents of the seeds, respectively. Following administration of 5.0-15 mg of NO3 to rats by stomach intubation, analysis of their 24, 48, and 72 h urine showed that only 40% of the administered nitrate appeared in the urine unmetabolized. Processing was shown to drastically reduce these antinutritional factors to very low levels. The health implications of these findings are discussed.

Underutilised legumes: potential sources for low-cost protein.

Seeds of 104 leguminous species belonging to 17 genera were analysed for their protein contents. The promising ones were investigated for fibre, carbohydrate, ash, oil, fatty acids, amino acid profile and trypsin inhibitor activity (TIA). The variation of fibre contents was 4.1-8.9%, carbohydrate 18.4-49.2%, ash 1.8-7.2%, TIA 48.7-87.5 mg/g, oil 1.3-19.8% and protein 11.0-51.6%. The protein content (41-45%) in Acacia mellifera (41.6%), Albizzia lebbek (43.6%), Bauhinia triandra (42.7%), Lathyrus odoratus (42.8%), Parkinsonia aculeata (41.6%), Psophocarpus tetragonolobus (41.9%), Sesbania paludosa (41.2%) and S. sesban (43.8%) was in close proximity to soybean (42.8%), whereas Bauhinia retusa (51.6%), B. variegata (46.5%), Delonix elata (48.7%) and Gliricidia maculata (46.3%) showed higher percentages of protein than soybean. The essential amino acid composition of some of the seed proteins was reasonably well balanced (lysine up to 7.6%). The seeds of Bauhinia retusa (18.6%), B. triandra (16.5%), B. variegata (17.3%), Gliricidia maculata (16.2%), Parkia biglandulosa (18.9%) and Psophocarpus tetragonolobus (19.8%) had a good amount of oil, comparable to soybean (18-22%). The fatty acid composition of some genera/species was quite promising with high amount of unsaturated fatty acids.

Assessment of postruminal amino acid digestibility of roasted and extruded whole soybeans with the precision-fed rooster assay.

The objectives of these studies were to predict the effects of roasting and extrusion temperatures of whole soybeans (SB) on intestinal protein digestibility in cattle. Intestinal digestibility was assessed with a two-stage in vitro or in situ ruminal incubation/precision-fed cecectomized rooster bioassay. In Exp. 1, whole SB (raw SB or SB roasted to 141, 149, or 157 degrees C exit temperature from a commercial roaster and steeped for 30 min) were incubated in strained ruminal fluid and McDougall's buffer (50:50) at 39 degrees C for 16 h. In Exp. 2, SB (ground raw SB or SB extruded at 116, 138, or 160 degrees C) were placed in polyester bags (20 x 30 cm) and suspended in the ventral rumen of steers for 16 h. Lyophilized residue of the in vitro or in situ incubations and samples of raw SB and most extensively heated SB (roasted SB at 157 degrees C or extruded SB at 160 degrees C) for each respective experiment were crop-intubated to cecectomized roosters. Total excreta were collected for 48 h after intubation and lyophilized, and amino acid (AA) concentrations were determined. In Exp. 1, total AA digestibility was 61.6 and 84.5% for unincubated whole raw SB and 157 degrees C roasted SB, respectively, and 66.2, 88.9, 91.3, and 91.6% for in vitro residues of whole raw SB and SB roasted at 141, 149, and 157 degrees C, respectively. Trypsin inhibitor (TI) activity was 20.09, 1.69, 1.54, and 1.84 mg/g fat-free DM for unincubated whole raw SB and 141, 149, and 157 degrees C roasted SB, respectively, and 30.84, 1.01, .90, and .26 mg/g fat-free DM for in vitro residues of whole raw SB, 141, 149, and 157 degrees C roasted SB, respectively. In Exp. 2, total AA digestibility was 68.5 and 87.7% for unincubated ground raw SB and 160 degrees C extruded SB, respectively, and 81.9, 91.3, 89.7, and 89.4% for in situ residues of ground raw SB and 116, 138, and 160 degrees C extruded SB, respectively. Trypsin inhibitor activity was 17.61, 4.89, 4.08, and 1.56 mg/g fat-free DM for unincubated ground raw SB, 116, 138, and 160 degrees C extruded SB, respectively, and 3.62, .59, .55, and .21 mg/g fat-free DM for incubated ground raw SB, 116, 138, and 160 degrees C extruded SB, respectively. Heat treatment by roasting and extrusion improved AA digestibilities of SB, but there were no differences detected among the roasting or extrusion temperatures. Ruminal fermentation did not eliminate the negative effects of TI activity on intestinal digestibility of AA in whole SB but did reduce TI activity in ground SB.

Chemical and nutritional assessment of sweetpotato proteins.

The solubility of sweetpotato nitrogen increased outside the pH range 3-6 for the pulp, and 2-5 for the peel of the two varieties namely, "Abees" and "Giza 69". The minimum nitrogen extractability occurred between the pH range 3-4, and 4-5 for the peel and pulp, respectively. The sweetpotato protein isolates were prepared separately from both peel and pulp of the two varieties. The former had the highest values of carbohydrates and ash while the latter had the highest values of protein and fat. The in-vitro digestibility of sweetpotato proteins and casein by pepsin-pancreatin was studied. Casein was more easily digested than the proteins of sweetpotatoes, whose digestibilities were higher in the pulp than in the peel of the two varieties. No trypsin inhibitor activity was detected in peel and pulp of both varieties under test. The effect of the variety of sweetpotatoes on the protein patterns was studied using the polyacrylamide gel electrophoresis and varietal specific patterns were obtained.