Absence Of Trypsin Inhibitor Research Articles

Physicochemical quality of brown soybean preserve in function of maceration time and ascorbic acid

Abstract The aim of the present study was to evaluate the physical and chemical quality of the brown soybean preserve in function of the ascorbic acid concentration and the maceration time, and also to evaluate the chemical composition, microbiological hazard, and acceptability of the best preserve, in addition verify its technological, nutritional, functional and sensory viability. The hardness of the grain ranged between 8.6 and 23N and was significantly affected by the maceration time. The ascorbic acid concentration influenced antioxidant activity (ranged between 4.3 and 183.8 mg100g-1) and total phenolic compounds (ranged between 176.4 and 466.2 mg100g-1). The best brown soybean preserve was obtained with a maceration time of 23 min and an ascorbic acid concentration of 0.15 g100g-1 . This shows high nutritional value and essential amino acid content, absence of trypsin inhibitor and microbiological risk, and also sensory acceptance. Therefore, the brown soybean preserve is viable, and its consumption is recommended.

Perfil de ácidos graxos e fatores antinutricionais de amêndoas de pequi crua e torrada

O pequi é um dos frutos mais abundantes no Cerrado, sendo sua polpa muito apreciada na culinária regional, porém, sua amêndoa (também comestível) é pouco aproveitada. O consumo da amêndoa crua pode acarretar em problemas digestivos, devido à possível presença de fatores antinutricionais, assim como a torra pode alterar, nutricionalmente, a amêndoa. Este trabalho objetivou verificar a composição centesimal, o perfil de ácidos graxos e os fatores antinutricionais, em amêndoas de pequi crua e torrada (a 270ºC, por 15 minutos), oriundas do Estado de Goiás e submetidas às análises de composição centesimal (umidade, proteínas, lipídeos, cinzas, carboidratos e valor energético), perfil de ácidos graxos e fatores antinutricionais (inibidores de tripsina, tanino e fitatos). A composição centesimal e os fatores antinutricionais, respectivamente para as amêndoas crua e torrada, apresentaram os seguintes teores: umidade: 25,40% e 1,70%; cinzas: 3,90% e 4,60%; proteínas: 13,40% e 14,70%; lipídeos: 24,70% e 25,70%; carboidratos: 32,50% e 53,30%; e valor energético: 406,20 kcal/100 g e 503,00 kcal/100 g, com ausência de inibidores de tripsina e taninos com 1,21% e 1,17% e fitatos com 2,64% e 1,86%. No perfil de ácidos graxos, respectivamente para as amêndoas crua e torrada, foram obtidos 86,90% e 84,61% de ácidos saturados e 10,57% e 10,40% de insaturados. A torra das amêndoas não influenciou, significativamente (p > 0,05), somente a variável proteína, interferindo, assim, nas características nutricionais e antinutricionais.

A Kunitz trypsin inhibitor from chickpea (<i>Cicer arietinum</i> L.) that exerts an antimicrobial effect on Fusarium oxysporum f.sp. ciceris

Fusarium oxysporum f.sp. ciceris (Foc) is one of the most important fungal pathogens of chickpea and is regarded as a constant threat in tropical and subtropical countries. In order to correlate Fusarium wilt resistance/susceptibility in Cicer arietinum to the presence or absence of trypsin inhibitor (TI) in the crude extract, trypsin inhibitory assay (TIA) and in vitro activity of TI against Foc were studied. In the present study, a 20 kDa trypsin inhibitor was purified from Fusarium wilt resistant cultivar (viz. JG 2001-12) by ammonium sulfate precipitation, dialysis and chromatographies with Sephadex G-100 and Diethyl aminoethyl cellulose (DEAE-cellulose-52) ion-exchange column. Results of pathogenecity assay were found to be in correlation to the trypsin inhibitor assay where the Fusarium wilt resistant cultivar showed high trypsin inhibitory activity (99%) in the presence of trypsin enzyme using both natural and synthetic substrates. Preliminary studies using crude extracts of JG 2001-12 showed a decrease in radial growth of Foc. A 45%-82% reduction in conidium germination at 20 μg&middotmL-1 Cicer arietinum trypsin inhibitor (CaTI) concentration was observed, thereby, indicating the use of CaTI in suppression of pathogen and in its deployment through transgenic plants for the management of Fusarium wilt.

New highly specific botulinum type C1 endopeptidase immunoassays utilising SNAP25 or Syntaxin substrates

Botulinum neurotoxins contain proteases that cleave specific intra-neural proteins essential for neurotransmitter release. Toxin types A, C1 and E intra-cellularly cleave SNAP25 and/or Syntaxin (type C1 only) resulting in a flaccid paralysis. Although highly sensitive, robust in vitro endopeptidase immunoassays have been developed for some serotypes, an endopeptidase immunoassay for type C1 has not previously been described. The current studies utilised solid phase synthesized SNAP25(137–206) peptide substrate, and a new specific antibody to the SNAP25(191–198) octapeptide epitope that becomes exposed following cleavage by type C1 toxin. The highly specific nature of the detecting antibody was illustrated by the failure of anti-SNAP25(191–198) to recognise the type A cleavage product which differs by just one amino acid residue. Conversely, anti-SNAP25(190–197), which recognises the type A cleavage product, fails to cross react with the type C1 toxin cleavage product. Utilising Syntaxin(232–266) peptide substrate, and a specific antibody to the cleavage product epitope, Syntaxin(254–261), it was also possible to develop an endopeptidase immunoassay. Assay sensitivities allowed the detection of less than 0.1 LD 50/ml (25 pg/ml) of type C1 haemagglutinin-complexed toxin. The assay failed to detect toxin serotypes A, B, D, E, F or G and therefore also provides an alternative highly specific in vitro identity test. In the absence of trypsin inhibitors, the assay is also capable of detecting 2 pg/ml of trypsin activity, or trypsin like contaminants. These new immunoassays will therefore provide highly specific tools for monitoring botulinum toxin light chain endopeptidase activity and serotype identity.

Inhibition of binding of an enzymatically stable thrombin inhibitor to lumenal proteases as an additional mechanism of intestinal absorption enhancement.

The objective of the study was to investigate the mechanisms behind increased bioavailability of an enzymatically stable thrombin inhibitor, inogatran, after coadministration with a trypsin inhibitor, aprotinin. Rat jejunum, ileum and colon segments were stripped and mounted in modified Ussing chambers, and the permeability to inogatran was determined both in the presence and absence of aprotinin. Inogatran and aprotinin were also coadministered intraduodenally to conscious rats. Competitive binding of inogatran to trypsin was studied using kinetic dialysis and was compared to aprotinin. The fraction of free (unbound) trypsin probe, in the absence of trypsin inhibitors was determined by performing experiments without pancreatine and without inhibitors, respectively. A 3-fold increased permeability to inogatran in the presence of aprotinin was seen in vitro, in some cases correlated with changed barrier properties of the intestinal segments. The in vitro results were well correlated with the in vivo results. There was a 5-fold increase in the bioavailability of inogatran in the presence of aprotinin. The binding of a trypsin probe was inhibited by both the presence of inogatran and aprotinin. Aprotinin showed a several fold higher displacement than inogatran. The results indicate both an effect of aprotinin on the epithelial membrane and an inhibition of binding of the thrombin inhibitor to trypsin or other serine proteases in the gut. The coadministration of aprotinin with enzymatically stable peptides, like thrombin inhibitors, may improve their absorption after oral administration. This suggests a new additional mechanism for intestinal absorption enhancement of peptide drugs.

The biosynthesis of insulin in rabbit islets of Langerhans

When isolated rabbit islets of Langerhans were incubated with 3H-labelled leucine for 2 h in the presence of trypsin inhibitor, label was predominantly incorporated into material which behaved like porcine proinsulin on gel electrophoresis. Trypsin treatment produced material which was apparently rabbit insulin. In the absence of trypsin inhibitor, incorporation of labelled leucine is mainly into rabbit insulin.

Effect of proteolytic enzyme inhibitors on steroidogenic action of corticotropin and its synthetic analogues.

The in vitro effect of Kunitz pancreatic trypsin inhibitor (KPTI) on the release of corticosterone from rabbit adrenals in the presence of three different preparations of corticotropins has been investigated. These preparations include highly purified porcine corticotropin, Pcorticotropin and Gly^a-*^ corticotropin. The effect of KPTI was found to be more marked in sustaining the action of corticotropin with shorter peptide chain than that with longer peptide chain. KPTI per se did not show any stimulatory effects on steroidogenesis. It may be concluded that KPTI protects shorter chain corticotropin against digestion by proteolytic enzymes. The above mentioned effect of KPTI was not demonstrated, however, with soybean trypsin inhibitor. (Endocrinology 90: 1652, 1972) T T HAS BEEN known that corticotropins are •*• inactivated by proteolytic enzymes. White and Gross (1) reported that plasmin inactivated corticotropins in blood. Other proteolytic enzymes, such as trypsin, pepsin and liver cathepsin have been also recognized as corticotropin inactivators (1). However, little is known as to whether corticotropins could be inactivated in the adrenal, during exerting their hormone action in adrenal. In the present report we employed proteolytic enzyme inhibitors to evaluate the possible influence of tissue proteolytic activities on the maintenance of steroidogenic action of corticotropins including synthetic analogues with shorter chain peptides. Our data suggest that the action of synthetic corticotropin analogue with 18 amino acids is markedly augmented by protecting against the degradation of the peptide. Materials and Methods A highly purified porcine corticotropin (HP corticotropin) with a potency of 120 U/mg was purchased from Calbiochem; synthetic P* corticotropin from Organon-Daiichi; Kunitz pancreatic trypsin inhibitor and soybean trypsin inhibitor from Worthington Biochemical Corp.; bovine serum albumin (Fr. V) from Armour Pharm. Co.; synthetic Gly-a-2 Received November 17, 1971, corticotropin was a gift from Dr. Otsuka of Shionogi Laboratory, Osaka. Adrenals were obtained from male rabbits weighing 2.0-2.5 kg. They were washed exhaustively by Krebs-Ringer bicarbonate buffer, pH 7.4 (KRB) and quadricepted after removal of thin fibrous capsules. Each adrenal preparation weighed 30-50 mg. Two pieces of quadricepted tissue from a pair of adrenal glands were placed in each incubation vessel containing 5 ml KRB with glucose (2 mg/ml) and bovine serum albumin (5 mg/ml) (G-KRB) in the presence or absence of trypsin inhibitors. The vessels were preincubated for 60 min in an atmosphere of 95% O2—5% CO2 at 37 C with shaking at 100 cycles/min. After preincubation, corticotropin preparations were added and incubation was continued for another 120 min under the same condition. The steroidogenic action was followed by measuring the amount of corticosterone released into the incubation medium. 0.5 ml aliquots of medium were removed at 0, 20, 40, 60 and 120 min for the assay of corticosterone which was performed by the method of Guillemin (2). Results Effect of KPTI on steroidogenic action of corticotropins The effect of three different corticotropin preparations on corticosterone production in rabbit adrenals were tested. As shown in Fig. 1A, HP corticotropin (0.04 y-g/ml), synthetic J3 NOTES AND COMMENTS 1653 FIG. 1. The effect of HP corticotropin, P' corticotropin and Gly-a-2 corticotropin on the corticosterone production in rabbit adrenals in the presence or absence of proteolytic enzyme inhibitors (KPTI or STI). Quadricepted adrenals were preincubated for 60 min with or without proteolytic enzyme inhibitors, each preparation of corticotropins was then added to incubation vessels as indicated with an arrow and incubation was continued for 120 min. Each value is the mean ± SE of four or five experiments. 12.0