Analysis of trypsin activity at β-casein layers formed on hydrophobic surfaces using a multiharmonic acoustic method

Proteolysis of milk proteins, such as caseins, caused by milk proteases, can change the organoleptic and nutritional characteristics of milk, and therefore it is essential to monitor this enzymatic activity....

Digestive Activity of Pancha Harithakadi Churna

The traditional systems of medicine are really effective but the problem with them is they lack in quality assurance. Standardization is the need of the hour in ayurvedic system of medicine. PanchaHarithakadi Churna (PHC) is a traditional polyherbal formulation which consists of five household ingredients used for indigestion. It is mainly used for Constipation and Bloating. Churna’s will play a major role in gastro intestinal problems and they have greater bioavailability because of smaller particle size. It consists of fine powder (sieve 100 size) of ginger rhizomes, fennel fruits, myrobalan fruits, senna leaflets and pink rock salt in equal proportions (1:1:1:1:1) are mixed well. PHC was formulated by standard procedures and evaluated by microscopic characterization, inorganic analysis and digestive studies. Microscopical characters indicate the presence of genuine crude drugs used in the formulation. Inorganic analysis shows the presence of calcium, magnesium, sodium, chloride and phosphate. The PHC showed pronounced amylolytic activity and trypsin activity whereas moderate lipolytic activity, proteolytic activity and pepsin activity, mild chymotrypsin activity in treating indigestion. In future we will carry out in vivo digestive studies.

Toxic Effects of Aflatoxin B1 in Chinese Sea Bass (Lateolabrax maculatus)

This study was performed to assess the effects of dietary aflatoxin B1 (AFB1) on the growth, antioxidant and immune response, digestive enzyme activities, and intestinal morphology of Lateolabrax maculatus during a 56-day feeding trial. Four diets were formulated including 0, 0.1, 0.5, and 1.0 mg/kg of AFB1. Each diet was randomly assigned to 3 fish tanks with 40 fish per tank. Results indicated that the fish’s final body weight, weight gain rate, specific growth rate, feed intake, condition factor, viscerosomatic index, hepatosomatic index, and intestinesomatic index decreased (p < 0.01) as dietary AFB1 increased. AFB1 levels in diets increased (p < 0.05) serum total antioxidant capacity (TAOC), superoxide (SOD), catalase, malondialdehyde (MDA), alkaline phosphatase (AKP), and lysozyme (LZM), and increased (p < 0.05) the TAOC, SOD, MDA, AKP, LZM, and immunoglobulin M in the livers of the fish. Dietary AFB1 decreased (p < 0.05) intestinal trypsin activity and induced intestinal injury. In summary, dietary AFB1 up to 1.0 mg/kg was toxic to L. maculatus as judged by reduced growth, enhanced antioxidant and immune response, decreased intestinal trypsin activity, and impaired intestinal morphology.

Culex quinquefasciatus Late Trypsin Biosynthesis Is Translationally Regulated by Trypsin Modulating Oostatic Factor

Trypsin is a serine protease that is synthesized by the gut epithelial cells of female mosquitoes; it is the enzyme that digests the blood meal. To study its molecular regulation, Culex quinquefasciatus late trypsin was purified by diethylaminoethyl (DEAE), affinity, and C18 reverse-phase high performance liquid chromatography (HPLC) steps, and the N-terminal amino acid sequence was determined for molecular cloning. Five overlapping segments of the late trypsin cDNA were amplified by PCR, cloned, and the full sequence (855 bp) was characterized. Three-dimensional models of the pro-trypsin and activated trypsin were built and compared with other trypsin models. Trypsin modulating oostatic factor (TMOF) concentrations in the hemolymph were determined by ELISA and compared with trypsin activity in the gut after the blood meal. The results showed that there was an increase in TMOF concentrations circulating in the hemolymph which has correlated to the reduction of trypsin activity in the mosquito gut. Northern blot analysis of the trypsin transcripts after the blood meal indicated that trypsin activity also followed the increase and decrease of the trypsin transcript. Injections of different amounts of TMOF (0.025 to 50 μg) decreased the amounts of trypsin in the gut. However, Northern blot analysis showed that TMOF injections did not cause a decrease in trypsin transcript abundance, indicating that TMOF probably affected trypsin translation.

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

This paper describes smart sampling paper to be used for bottom-up protein analysis. Four different manners to immobilize trypsin on cellulose were evaluated. Untreated paper, potassium-periodate-functionalized paper (with and without post-immobilization reduction) and 2-hydroxyethyl methacrylate (HEMA)/2-vinyl-4,4-dimethylazlactone (VDM)-functionalized paper were all used to immobilize trypsin. For the evaluation, Coomassie Brilliant Blue staining of proteins on paper and the BAEE trypsin activity assay needed to be modified. These methods allowed, together with data from mass spectrometric analysis of cytochrome C digestions, us to acquire fundamental insight into protein binding, and trypsin action and activity on paper. All functionalized discs bind more protein than the untreated discs. Protein binding to functionalized discs is based on both adsorption and covalent binding. Trypsin immobilized on potassium-periodate-functionalized discs exhibits the highest trypsin activity when using cytochrome C as substrate. It is proven that it is trypsin attached to paper (and not desorbed trypsin) which is responsible for the enzyme activity. The use of discs on complex biological samples shows that all functionalized discs are able to digest diluted serum; for the best-performing disc, HEMA-VDM functionalized, up to 200 high-confidence proteins are qualified, showing its potential.

Enzymatic activity in the shrimp Penaeus vannamei fed at different feeding frequencies

Correct management of the feeding regime in shrimp aquaculture has been beneficial. Still, when looking for improvement in shrimp performance, the results have been contradictory, and the limits between better growth and independent growth as a function of the feeding regime are not clear. In this study, trypsin and α-amylase activity, as well as an interpretation of the energy utilized for enzyme production, were evaluated in shrimp weighing 1 g. Four feeding groups were set to feed one, two, four, or eight times per day over a month, after which trypsin and α-amylase activities were evaluated during 29 h. Results indicated that the group fed once per day ingested 90% of the feed, whereas the other groups ingested 100%. The α-amylase was not consumed during the daytime in all groups, unlike trypsin. Total trypsin activity was not significantly different between feeding groups, but α-amylase was significantly different. Shrimp fed eight times had an elevated α-amylase activity level that was 2.6 times greater than those fed only once, and 0.8 and 0.5 times greater than those fed four and twice per day, respectively. Feeding more frequently generates a higher use of energy that may or may not be reflected in growth but could be essential for all the energy-dependent metabolic processes required by shrimp.

Colitis Linked to Endoplasmic Reticulum Stress Induces Trypsin Activity Affecting Epithelial Functions

Background and Aims Intestinal epithelial cells [IECs] from inflammatory bowel disease [IBD] patients exhibit an excessive induction of endoplasmic reticulum stress [ER stress] linked to altered intestinal barrier function and inflammation. Colonic tissues and the luminal content of IBD patients are also characterized by increased serine protease activity. The possible link between ER stress and serine protease activity in colitis-associated epithelial dysfunctions is unknown. We aimed to study the association between ER stress and serine protease activity in enterocytes and its impact on intestinal functions Methods The impact of ER stress induced by Thapsigargin on serine protease secretion was studied using either human intestinal cell lines or organoids. Moreover, treating human intestinal cells with protease-activated receptor antagonists allowed us to investigate ER stress-resulting molecular mechanisms that induce proteolytic activity and alter intestinal epithelial cell biology. Results Colonic biopsies from IBD patients exhibited increased epithelial trypsin-like activity associated with elevated ER stress. Induction of ER stress in human intestinal epithelial cells displayed enhanced apical trypsin-like activity. ER stress-induced increased trypsin activity destabilized intestinal barrier function by increasing permeability and by controlling inflammatory mediators such as C-X-C chemokine ligand 8 [CXCL8]. The deleterious impact of ER stress-associated trypsin activity was specifically dependent on the activation of protease-activated receptors 2 and 4. Conclusions Excessive ER stress in IECs caused an increased release of trypsin activity that, in turn, altered intestinal barrier function, promoting the development of inflammatory process.