Exploring metalloproteins found in the secretion of venomous species: Biological role and therapeutical applications.

Several species during evolution suffered random mutations in response to various environmental factors, which resulted in the formation of venom in phylogenetically distant species. The composition of the venom of most species is poorly known. Snake venom is well characterized while most species have poorly known composition. In contrast, snake venoms are well characterized which proteins and peptides are the main active and most abundant constituents. 42 protein families have been identified, including metalloproteins known as metalloproteinases. These macromolecules are enzymes with zinc in their active site derived from the disintegrin A and metalloproteinase (ADAM) cellular family and are categorized into three classes (PI, PII and PIII) according to their domain organization. The snake venom metalloproteinases (SVMP) are cytotoxic, neurotoxic, myotoxic and/or hematotoxic with a crucial role in the defense and restraint of prey. In this scenario envenoming represents a danger to human health and has been considered a neglected disease worldwide, particularly in tropical and subtropical countries. Nevertheless, recently advances in "omics" technologies have demonstrated interesting biological activities of SVMPs such as antimicrobial, anticancer, against cardiovascular diseases and nervous system disorders. Metalloproteins have the therapeutic potential to be converted into drugs as other components of the venom have undergone this process (e.g., captopril, tirefiban and eptifibatide). So, this chapter is focused on the metalloproteins found in the secretions of venomous species, highlight some aspects such as structure, biological activity, pharmacological therapeutic potential and on.

Application of experimental design as a statistical approach to recover bioactive peptides from different food sources.

Bioactive peptides (BAPs) derived from samples of animals and plants have been widely recommended and consumed for their beneficial properties to human health and to control several diseases. This work presents the applications of experimental designs (DoE) used to perform factor screening and/or optimization focused on finding the ideal hydrolysis condition to obtain BAPs with specific biological activities. The collection and discussion of articles revealed that Box Behnken Desing and Central Composite Design were the most used. The main parameters evaluated were pH, time, temperature and enzyme/substrate ratio. Among vegetable protein sources, soy was the most used in the generation of BAPs, and among animal proteins, milk and shrimp stood out as the most explored sources. The degree of hydrolysis and antioxidant activity were the most investigated responses in obtaining BAPs. This review brings new information that helps researchers apply these DoE to obtain high-quality BAPs with the desired biological activities.

Preliminary ionome of the parotoid gland secretion from Rhinella jimi toad.

The cururu toad (Rhinella jimi) is an anuran belonging to the fauna of the Brazilian northeast region, which releases a secretion with toxins from your parotoid glands. Although it has some information about secondary metabolites and proteins, the elemental composition of the released secretion is unknown. Therefore, this is the first report on the ionome of the secretion of the parotoid glands from R. jimi, investigating the influences of abiotic factors such as biome, seasonality, and gender. ICP-MS was used for measurements combined with principal component analysis (PCA). A screening of the secretion sample detected 68 elements which the total concentration of 18 elements was determined. PCA revealed that biome and seasonality factors have a greater influence on the ionomic profile of parotoid secretion. The presence of toxic metals in the secretion samples indicates that the R. jimi toad can be considered a potential bioindicator. These findings may contribute to understanding the metabolism, lifestyle, and interaction of the R. jimi toad with environmental factors as well as open new perspectives to investigate the relationships of the ionome with other biomolecules, for example, metalloproteins and their physiological functions.

Combining high intensity ultrasound and experimental design to improve carotenoid extraction efficiency from Buriti (Mauritia flexuosa).

Buriti (Mauritia flexuosa L.) is a significant source of carotenoids, but these compounds have been extracted using laborious and low-effective methods. The present work evaluated the high-intensity ultrasound combined with a chemometric approach to developing an optimal extraction method of carotenoids from buriti pulp. The multivariate optimization was carried out through two steps. First, a simplex-lattice mixture design was used to optimize the extractor solution finding higher extraction yield (903 ± 21 µg g-1) with the acetone:ethanol (75/25) mixture. After, sample mass (80 mg) and sonication time (30 min) were optimized applying central composite design (CCD) which provided a 14% improvement in the extraction method yield. So, the total carotenoid content (TCC) with optimal extraction conditions was 1026 ± 13 µg g-1which is almost twice the yield of methods known in the literature for buriti. The RP-HPLC-DAD analysis revealed that the carotenoids are gently extracted and ß-carotene is the major compound in the extracts. To confirm the accuracy, buriti samples spiked with ß-carotene standard and the developed method showed recovery >84% and precision <6.5%. Furthermore, the optimized ultrasound-assisted extraction (UAE) method was applied to other samples (tomato, guava, carrot, mango, acerola, papaya, and pumpkin) and presented a yield to 5.5-fold higher when compared to the reported methods indicating high robustness. Based on results, the UAE method developed has demonstrated feasibility and reliability for the study of carotenoids in buriti pulp as well as in other plant matrices with high biological relevance.

Trace element homeostasis in the neurological system after SARS-CoV-2 infection: Insight into potential biochemical mechanisms.

BACKGROUND: Several studies have suggested that COVID-19 is a systemic disease that can affect several organs, including the brain. In the brain, specifically, viral infection can cause dyshomeostasis of some trace elements that promote complex biochemical reactions in specialized neurological functions. OBJECTIVE: Understand the neurovirulence of SARS-CoV-2 and the relationship between trace elements and neurological disorders after infection, and provide new insights on the drug development for the treatment of SARS-CoV-2 infections. METHODS: The main databases were used to search studies published up September 2021, focusing on the role of trace elements during viral infection and on the correct functioning of the brain. RESULTS: The imbalance of important trace elements can accelerate SARS-CoV-2 neurovirulence and increase the neurotoxicity since many neurological processes can be associated with the homeostasis of metal and metalloproteins. Some studies involving animals and humans have suggested the synapse as a vulnerable region of the brain to neurological disorders after viral infection. Considering the combined evidence, some mechanisms have been suggested to understand the relationship between neurological disorders and imbalance of trace elements in the brain after viral infection. CONCLUSION: Trace elements play important roles in viral infections, such as helping to activate immune cells, produce antibodies, and inhibit virus replication. However, the relationship between trace elements and virus infections is complex since the specific functions of several elements remain largely undefined. Therefore, there is still a lot to be explored to understand the biochemical mechanisms involved between trace elements and viral infections, especially in the brain.

Multivariate optimization and comparison between conventional extraction (CE) and ultrasonic-assisted extraction (UAE) of carotenoid extraction from cashew apple.

Carotenoids are an essential component of cashew and can be used in pharmaceuticals, cosmetics, natural pigment, food additives, among other applications. The present work focuses on optimizing and comparing conventional and ultrasound-assisted extraction methods. Every optimization step took place with a 1:1 (w:w) mixture of yellow and red cashew apples lyophilized and ground in a cryogenic mill. A Simplex-centroid design was applied for both methods, and the solvents acetone, methanol, ethanol, and petroleum ether were evaluated. After choosing the extractor solvent, a central composite design was applied to optimize the sample mass (59-201 mg) and extraction time (6-34 min). The optimum conditions for the extractor solvent were 38% acetone, 30% ethanol, and 32% petroleum ether for CE and a mixture of 44% acetone and 56% methanol for UAE. The best experimental conditions for UAE were a sonication time of 19 min and a sample mass of 153 mg, while the CE was 23 min and 136 mg. Comparing red and yellow cashews, red cashews showed a higher carotenoid content in both methodologies. The UAE methodology was ca. 21% faster, presented a more straightforward composition of extracting solution, showed an average yield of superior carotenoid content in all samples compared to CE. Therefore, UAE has demonstrated a simple, efficient, fast, low-cost adjustment methodology and a reliable alternative for other applications involving these bioactive compounds in the studied or similar matrix.

Proteins from Rhinella jimi parotoid gland secretion: A comprehensive analytical approach.

Toad skin secretions are sources of complex mixtures of bioactive compounds, such as proteins and peptides. Rhinella jimi species is a common toad in the Brazilian northeast, considered by only a few known studies. The experimental design was applied to optimize the protein extraction method from R. jimi parotoid gland secretions. The optimum condition was using 100 mmol L-1 Tris-HCl buffer pH 7.2 under vortexing for 5 min. The FTIR analysis combined with PCA revealed high-protein purity of the extracts, confirming the success of the proposed extraction method. The total protein concentration by the Bradford method was 102.4 and 66.5 mg g-1 on toad poisons from Teresina and Picos, respectively. The comparative proteomic analysis using HPLC-SEC-DAD and 1D SDS-PAGE revealed significant differences in protein abundance. HMW biomolecules showed greater abundance in toads from Teresina, while LMW protein species were more abundant in toads from Picos. The significant difference in amphibian proteome can be attributed to the edaphoclimatic conditions of their habitat. The cytotoxicity of the protein extract from Teresina was higher on the tumor cell lines 4T1 and CT26.WT. These new findings are fundamental for future studies the on identity and biological activity of biomolecules from this noble sample.

Expanding information on the bioaccessibility and bioavailability of iron and zinc in biofortified cowpea seeds.

This work presents new findings on the nutritional quality of recently introduced biofortified and non-biofortified cowpea cultivars as well as some common beans. ICP-MS was used for the measurements. Biofortified cowpea cultivars showed high levels of Fe and Zn, greater than 60 and 40 mg kg-1 dry weight, respectively. The in vitro digestion protocol enabled simultaneous evaluation of bioaccessibility and bioavailability. Fe levels in cowpea cultivars were ca. 2.5-fold higher than in common beans. Cowpea seeds also had higher Zn levels, reaching 50.1% bioaccessibility and 44.2% bioavailability. Cooking improved the availability of micronutrients in bean seeds. The cooked biofortified Aracê cowpea showed a high Zn bioavailability above 60%. Consumption of 50 g of Aracê would contribute 27% and 48% of the Fe and Zn DRI for 1-3-year-old children. The new cowpea cultivars biofortified are a potential vehicle for improving the Fe and Zn status in groups in which the micronutrient deficiency is prevalent.

Application of multivariate optimization for the selective extraction of phenolic compounds in cashew nuts (Anacardium occidentale L.).

An extraction method based on a multivariate analytical approach was developed for enhancement of the phenolic compounds in cashew nut extracts. The different extractor solvents (acetone, water, ethanol, and methanol) and their binary, ternary, and quaternary combinations were evaluated using a simplex-centroid design and surface response methodology. The special cubic model exhibits no lack of fit and explains 89.2% of the variance. The total phenolic measurements by the Folin-Ciocalteu method revealed the highest values for ethanol (5.93 mg GAE g-1) and acetone-methanol-ethanol ternary mixture (5.92 mg GAE g-1) extracts. ESI (-)-Q/TOFMS analyses combined with PCA and HCA revealed the presence of fatty acids, phospholipids, and sugars in the ternary mixture cashew extract, while for the ethanol extract only phenolic compounds, such as anacardic acids and derivatives, were found. The proposed approach was adequate to reach the optimal extractor which ethanol, a low-toxicity solvent, enabled the selective extraction of a high content of phenolic compounds from cashew nuts.

Optimization of the protein extraction method of goat meat using factorial design and response surface methodology.

Protein extraction from goat meat was carried out based on the combination of response surface methodology and factorial design to optimize the variables: temperature (25-50 °C), extraction time (8-20 min), volume (3-10 mL) and extractor concentration (0.05-0.1 mol L-1). The proposed model did not present a lack of fit, explaining 96% of the total data variance (R2 = 0.96). The optimum extraction conditions were: 0.05 mol L-1 for extractor concentration, extraction time of 10 min, temperature of 44 °C and extractor volume of 3.5 mL. The protein content (19.3 g/100 g) obtained by the optimized method was higher than some results reported in the literature. HPLC-SEC-DAD analysis revealed that the extraction conditions used did not significantly modify the protein structure. The proposed method proves to be simple, fast, robust, cheap and adequate for native protein extraction, being a potential approach for proteomic research focusing in goat meat.

Evaluation of some effects on plant metabolism through proteins and enzymes in transgenic and non-transgenic soybeans after cultivation with silver nanoparticles.

To evaluate the effects of silver nanoparticles (AgNP) exposition, transgenic (through gene cp4EPSPS) and non-isogenic non-transgenic soybeans were cultivated in the presence or absence of AgNP or silver nitrate (AgNO3) at 50 mg/kg of silver. Physiological aspects of the plants including mass production and development of roots, proteomics such as protein amount and differential proteins, enzymes and lipid peroxidation were determined after exposition. The mass production of non-transgenic plants treated with AgNP or AgNO3 was decreased by 25 and 19%, respectively, on their mass based, while for transgenic soybean this effect was observed for AgNP cultivation only. Fifty-nine proteins were identified from the differentially abundant spots by two-dimensional difference gel electrophoresis and nano-electrospray ionization liquid chromatography coupled tandem mass spectrometry. Identified species as ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), ATP synthase, superoxide dismutase (SOD), related to plant metabolism were less abundant for the cultivation with either AgNP and AgNO3 than the control. Finally, this work demonstrated significant correlation as evidenced by changes in lipid peroxidation content and catalase activity, which were a result of exposure to either AgNP or AgNO3 cultivations. Further, necrotic areas in the basal part of the stems and damage or chlorotic areas were found in the leaves. SIGNIFICANCE: Once nanoparticles have been employed for several applications in recent years and they can be released in the environmental matrices, this study highlights proteomic and enzymatic alterations in transgenic and non-transgenic soybeans, an important crop, after cultivation with silver nanoparticles. Such strategy employing proteomic and enzymatic approaches to evaluate soybeans exposed to silver nanoparticles has not yet been reported. Therefore, the results obtained in this study can expand the information concerning the effects of silver nanoparticles in soybean plants.

Evaluation of proteome alterations induced by cadmium stress in sunflower (Helianthus annuus L.) cultures.

The present study evaluates, at a proteomic level, changes in protein abundance in sunflower leaves in the absence or presence (at 50 or 700mg) of cadmium (as CdCl2). At the end of the cultivation period (45 days), proteins are extracted from leaves with phenol, separated by two-dimensional difference gel electrophoresis (2-D DIGE), and excised from the gels. The differential protein abundances (for proteins differing by more than 1.8 fold, which corresponds to 90% variation) are characterized using nESI-LC-MS/MS. The protein content decreases by approximately 41% in plants treated with 700mg Cd compared with control plants. By comparing all groups of plants evaluated in this study (Control vs. Cd-lower, Control vs. Cd-higher and Cd-lower vs. Cd-higher), 39 proteins are found differential and 18 accurately identified; the control vs. Cd-higher treatment is that presenting the most differential proteins. From identified proteins, those involved in energy and disease/defense (including stress), are the ribulose bisphosphate carboxylase large chain, transketolase, and heat shock proteins are the most differential abundant proteins. Thus, at the present study, photosynthesis is the main process affected by Cd in sunflowers, although these plants are highly tolerant to Cd.