Hydrolysis of Casein by Pepsin Immobilized on Heterofunctional Supports to Produce Antioxidant Peptides.

A study was carried out on the immobilization of pepsin in activated carbon functionalized by different techniques (glutaraldehyde, genipin, and metallization) aiming at its application in obtaining bioactive peptides through casein hydrolysis. Studies of the immobilized derivatives were carried out in addition to the evaluation of the antioxidant potential of the peptides. Among the pH range studied, pH 3.0 was selected due to the higher activity of the derivatives at this pH. The support modification by metallization was the method with the best results, providing a 121% increase in enzymatic activity compared to other immobilization methods. In addition, this derivative provided activity closer to the soluble enzyme activity (3.30 U) and better storage stability, and allows reuse for more than 8 cycles. In turn, the peptides from casein hydrolysis showed potential as antioxidant agents, with a DPPH radical scavenging activity higher than 70%, maximum protection against ß-carotene oxidation close to 70%, and a maximum reducing power of Fe(III) into Fe(II) of 400 uM by the FRAP assay. The results showed that the new techniques for modification of activated carbon can be a promising approach for pepsin immobilization.

Pepsin immobilization on activated carbon and functionalized with glutaraldehyde and genipin for the synthesis of antioxidant peptides of goat casein.

In this article, the synthesis of antioxidant peptides in the enzymatic hydrolysis of caprine casein was analyzed at three different time points (60 min, 90 min, and 120 min) using immobilized pepsin on activated and modified carbon (AC, ACF, ACG 50, ACG 100). The immobilization assays revealed a reduction in the biocatalysts' activity compared to the free enzyme. Among the modified ones, ACG 50 exhibited greater activity and better efficiency for reuse cycles, with superior values after 60 min and 90 min. Peptide synthesis was observed under all studied conditions. Analyses (DPPH, ß-carotene/linoleic acid, FRAP) confirmed the antioxidant potential of the peptides generated by the immobilized enzyme. However, the immobilized enzyme in ACG 50 and ACG 100, combined with longer hydrolysis times, allowed the formation of peptides with an antioxidant capacity greater than or equivalent to those generated by the free enzyme, despite reduced enzymatic activity.

Natural parasitism of the coffee leaf miner: climate factors, insecticide, and landscape affecting parasitoid diversity and their ecosystem services in coffee agroecosystems.

Climate factors, pesticides, and landscape in coffee agroecosystems directly affect the populations of the coffee leaf miner and its parasitoids. This study aimed to investigate the effects of climate factors, insecticide use, and landscape on natural parasitism, parasitoid diversity, and infestation of L. coffeella in coffee plantations in the Planalto region, Bahia, Brazil. Mined leaves were collected monthly in six coffee plantations with varying edge density, vegetation cover, landscape diversity in scales of 500 to 3000 m of radius, insecticide use, and climate factors. Closterocerus coffeellae, and Proacrias coffeae (Eulophidae) predominated in the pest's natural parasitism. Our record is the first for the occurrence of Stiropius reticulatus, Neochrysocharis sp. 1, Neochrysocharis sp. 2, and Zagrammosoma sp. in Bahia. Higher temperature and larger forest cover increased the coffee leaf miner infestation. Higher rainfall values, insecticide use, and landscape diversity decreased the pest infestations. Natural parasitism and species diversity are favoured by increase in temperature, forest cover, and edge density, while increase in rainfall, insecticide use, and landscape diversity lead them to decrease.The natural parasitism and diversity of parasitoid species of the coffee leaf miner have been enhancing in the areas with greater forest cover and edge density associated with low use of insecticides. The areas composed of different lands with annual croplands surrounding the coffee plantations showed less natural parasitism and parasitoid species diversity. The ecosystem services provided by C. coffeellae and P. coffeae in coffee crops areas require conservation and these species are potential bioproducts for applied biological control programmes.

Functionalized activated carbon as support for trypsin immobilization and its application in casein hydrolysis.

This study aimed to immobilize trypsin on activated carbon submitted to different surface modifications and its application in casein hydrolysis. With the aim of determining which support can promote better maintenance of the immobilized enzyme. Results showed that pH 5.0 was obtained as optimal for immobilization and pH 9.0 for the casein hydrolysis reaction for activated carbon and glutaraldehyde functionalized carbon. Among the supports used, activated carbon modified with iron ions in the presence of a chelating agent was the one that showed best results, under the conditions evaluated in this study. Presenting an immobilization yield of 95.15% and a hydrolytic activity of 4.11 U, same as soluble enzyme (3.76 U). This derivative kept its activity stable at temperatures above 40 °C for1 h and when stored for 30 days at 5 °C. Furthermore, it was effective for more than 6 reuse cycles (under the same conditions as the 1st cycle). In general, immobilization of trypsin on metallized activated carbon can be an alternative to biocatalysis, highlighting the advantages of protease immobilization.

The effects of thiamethoxam on coffee seedling morphophysiology and Neotropical leaf miner (Leucoptera coffeella) infestations.

BACKGROUND: Coffee (Coffea arabica L.) is one of the main commodities produced in Brazil. Insecticides like the (systemic) neonicotinoid thiamethoxam are widely used to suppress pest populations during coffee production, in particular the Neotropical leaf miner (Leucoptera coffeella Guérin-Mèneville & Perrottet, 1842) (Lepidoptera: Lyonetiidae). In addition to its efficacy against this pest species, thiamethoxam is also thought to be a bioactivator of plant metabolism, but has not yet been tested for such activity. Thus, the objectives of the present study were (1) to assess the concentration-response effects of thiamethoxam on the vegetative vigor of coffee seedlings (C. arabica 'Catuaí 144' cultivar) at different concentrations [2, 20, 40, 80 and 200 mg active ingredient (a.i.) kg-1 ] applied via soil drenching and (2) to evaluate if the plant response interferes with the effectiveness of thiamethoxam in controlling leaf miner populations. The morphophysiological traits of the coffee seedlings were evaluated 20, 40, 60 and 80 days after application, and leaf miner infestations were recorded starting 20 days after the insecticide application with the releasing of adults, and every 20 days afterwards. RESULTS: The results indicated that thiamethoxam has a deleterious effect on the morphophysiological traits of the plants compromising their development with increase in concentrations. However, leaf area exhibited a different pattern with a peak at 50 mg a.i. kg-1 consistent with thiamethoxam-induced hormesis (i.e. biphasic response with stimulatory effect at sublethal range of a toxic substance at the higher concentration). Nonetheless, such bioactivator effect did not affect thiamethoxan effectiveness against the leaf miner even at the lowest concentration tested. CONCLUSION: Thiamethoxan exhibited bioactivation effect on leaf at low concentration, but without compromising efficacy against leaf miner populations. Therefore, its proposed metabolism-boosting properties may encourage the unnecessary use of this insecticide, potentially leading to higher selection for insecticide resistance and an eventual decline in its effectiveness against the Neotropical leaf miner. © 2022 Society of Chemical Industry.

Pepsin immobilization: Influence of carbon support functionalization.

Among the matrices for enzyme immobilization, activated carbon has been standing out in immobilization processes due to its properties and to its characteristics that provide superficial modification by inserting new functional groups capable of binding the enzymes forming covalent bonds. In this study the effect of different modification methods of activated carbon (functionalization with genipin, metallization, metallization in the presence of chelating agent, and functionalization with glutaraldehyde) on efficiency of pepsin immobilization was evaluated. The effect of immobilization pH and the reaction medium on hydrolysis activity of bovine casein was also evaluated. The functionalization of activated carbon using iron ions allowed an immobilization capacity of 98.93 mg·g-1, with immobilization efficiency greater than 99%, and enzyme activity of 2.30 U, which was higher than the other modifications, and closer to the enzyme in the native form activity (3.32 U). In general, the carbon surface modifications were responsible for forming more stable bonds between support and enzyme, improving its proteolytic activity (from 1.84 to 2.30 U) when compared to traditional immobilization methods by adsorption and covalent binding using glutaraldehyde (from 1.04 to 1.1 U).

Hydrolysis of casein from different sources by immobilized trypsin on biochar: Effect of immobilization method.

The present study aimed to evaluate the effect of the immobilization method of trypsin on biochar on the hydrolysis of casein from different sources, when compared to the process using trypsin in native form, to obtain bioactive peptides. The modification of the surface of biochar with glutaraldehyde was effective, as shown by the results of FTIR assay and the texture profile of the materials. Both activated and functionalized biochar showed high immobilization efficiency (greater than 87%) and high binding capacity (greater than 91 mg/g). During hydrolysis, the biocatalyst obtained by enzyme immobilization on the functionalized biochar presented a higher hydrolysis capacity for the different caseins when compared to the enzyme immobilized by adsorption, with values of 3.05 and 2.73 U/mg for goat casein, 2.36 and 1.85 U/mg for bovine casein, and 2.60 and 2.37 U/mg for buffalo, casein, respectively, with 60 min of reaction. The results of inhibitory activity in this study ranged from 93.5% and 25.5% for trypsin in its free form and immobilized on functionalized activated carbon, respectively, under the same reaction conditions. The immobilization methods were efficient, presenting high immobilization capacity. The proteolytic activity of trypsin immobilized via covalent binding was higher when compared the immobilization by adsorption. Thus, the functionalized biochar has proven to be potential support for enzyme immobilization, and the biocatalyst can be reused for more than 4 cycles. Despite lower ACE inhibition values of hydrolyzed obtained with the immobilized enzymes compared to free enzymes, biocatalysts present advantage due to the possibility of reuse.

Area-Wide Survey of Chlorantraniliprole Resistance and Control Failure Likelihood of the Neotropical Coffee Leaf Miner Leucoptera coffeella (Lepidoptera: Lyonetiidae).

The Neotropical coffee leaf miner, Leucoptera coffeella (Guérin-Mèneville & Perrottet, 1842), is a key pest species of unshaded coffee plantations in Neotropical America, particularly in Brazil, where pest management involves intensive insecticide use. As a consequence, problems of resistance to conventional insecticides are frequent, and more recently developed insecticide molecules, such as diamide insecticides, are at risk of becoming ineffective. Thus, a survey of resistance to the diamide insecticide chlorantraniliprole was carried out in high-yield coffee-producing areas in the State of Bahia, Brazil. The likelihood of control failure with this insecticide was also assessed. Spatial dependence among the insect sampling sites was assessed and spatial mapping of chlorantraniliprole resistance and risk of control failure was carried out. The frequency of chlorantraniliprole resistant populations was high (34 out of 40 populations, or 85%), particularly in western Bahia, where 94% of the populations were resistant. Resistance levels ranged from low (<10-fold) to moderate (between 10- and 40-fold) with more serious instances occurring in western Bahia. This results in lower chlorantraniliprole efficacy among these populations, with a higher risk of control failure and exhibiting spatial dependence. These findings invite attention to problems with the intensive use of this relatively recent insecticide and demand management attention, but they suggest that local, farm-based management efforts are likely to be the most effective actions against resistance problems in this pest species.

Oyster Culinary-Medicinal Mushroom, Pleurotus ostreatus (Higher Basidiomycetes), Growth in Grain-Based Diet Improves Broiler Chicken Production.

Many alternative compounds have been tested to improve poultry performance but few of them have previously used mycelial-colonized substrate to partially replace standard diet in broiler chickens. The objective of this study was to evaluate broiler chicken production, health, and meat sensory characteristics, with partial replacement of the standard diet by Pleurotus ostreatus-colonized substrate. One hundred fifty 1-day-old male Cobb chicks were given standard diet partially replaced by 0, 5, 10, 100, or 200 g·kg⁻¹ of P. ostreatus-colonized substrate and randomly distributed into five treatments. Each treatment had three replicates, with 10 birds per replicate, totaling 30 birds. The replacement of the standard diet by 10 g·kg⁻¹ of colonized substrate increased (P≤0.05) chicken body mass up to 57% at 21 days, and up to 28% at 42 days. In general, partial replacement of standard diet by colonized substrate increased hematocrits and typical lymphocytes, and reduced low density lipoproteins. Also, it reduced chicken production period up to 21% and there is no meat taste alteration. The use of P. ostreatus-colonized substrate in chicken feeding is an alternative method to improve broiler chicken production.