Trypsin Inhibitor Activity Research Articles

Novel Ag(I)‐based 1,2,4‐Oxadiazole Complexes: Synthesis, X‐Ray Crystal Structure, and Biological Evaluation as Anticancer Candidates

ABSTRACTThe 3,5‐diaryl‐1,2,4‐oxadiazole scaffolds 2 and 3 were synthesized and used as ligands to obtain three novel Ag(I) complexes 4–6. The structure of the Ag(I) complexes 4–6 has been confirmed by single crystal X‐ray diffraction. Complex 4 has the dinuclear formula [Ag(2)(NO3)]2. 5 and 6 are monomeric complexes having the formula [Ag(3)2(NO3)] and [Ag(3)2]ClO4, respectively. In vitro, trypsin, ALDH2, and iNOS inhibition activities were assessed for the free ligands and their Ag(I) complexes. Interestingly, Ag(I) complexes 4–6 revealed more prominent trypsin inhibitory activity than the ligands 2 and 3. The oxadiazole derivative 3 and its Ag(I) complex (6) showed significant ALDH2 inhibition (45% and 55%, respectively). Complex 6 (IC50 = 35.61 μM) surpassed its 1,2,4‐oxadiazole ligand 3 (IC50 = 88 μM) and evidenced the most prominent ALDH2 inhibitory activity. The oxadiazole derivative 3 and its corresponding Ag(I) complexes 5 and 6 showed significant iNOS inhibition (77.77%, 84.12%, and 84.15%, respectively). With IC50 values of 18.13, 18.15, and 13.96 μM, respectively, complex 6 is the most potent against iNOS, surpassing the reference standard.

The impact of milling degree on physicochemical, nutritional, rheological, textural properties, and in vitro digestibility of rice: From brown rice to white rice

The objective of this study was to characterize the rice milling fractions acquired at each stage of a commercial milling system. This characterization included an analysis of color, ash content, dietary fiber, mineral composition, as well as antinutritional compounds like phytic acid, trypsin inhibitor activity, and saponin. Additionally, we investigated in vitro starch and in vitro protein digestibility, along with pasting, cooking, and textural properties.Our findings revealed that milling improved the visual appeal of rice (e.g., volume expansion, weight gain, whiteness) and notably enhanced its starch digestibility. However, milling reduced dietary fiber, mineral content, antinutrients, cooking time, and texture characteristics such as hardness, chewiness, gumminess, springiness, and cohesiveness to varying degrees. Certain parameters, such as dietary fiber, exhibited a gradual change with the duration of milling, while others, such as mineral content and texture, showed significant variation at the initial stage of milling, particularly at the first mill.

Comparison of rice bran characteristics fermented by mono- or di-strain probiotics based on omics technology

Rice bran is a premium agricultural byproduct rich in nutrients and bioactive substances. The aim of this study was to compare the effects of fermentation with Bacillus subtilis, Lactobacillus plantarum, and a mixture of both on the bacterial communities and metabolic functions of rice bran based on omics techniques. The results showed that probiotic fermentation decreased crude fiber (CF) content, phytic acid, and trypsin inhibitor activity (TIA), and increased trichloroacetic acid soluble protein (TCA-SP), reducing sugar (RS), and total phenol (TP) in rice bran. Ten dominant genera with relative abundances greater than 1% were successfully identified, and probiotic fermentation significantly reduced the relative abundances of Enterobacter, Kasakonia, and Pantoea. A total of 63 differentially expressed metabolites (DEMs) were identified and further analysis revealed that probiotic fermentation increased essential amino acids, phenolic compounds, and organic acids in rice bran. Additionally, the di-strain probiotic fermentation of Bacillus subtilis and Lactobacillus plantarum facilitated the growth of beneficial microbiota (Limosilactobacillus, Prevotella and Dialister) and the production of bioactive substances (lactic acid, catechol, beta-tyrosine, etc.). In conclusion, co-fermentation of di-strain probiotics is a valuable method to produce functional compounds and enhance the health-promoting properties of rice bran.

Credible and Facile Fluorometric Detection of Soybean Trypsin Inhibitor Activity with a Water-Soluble Poly(diphenylacetylene) Derivative

Credible and Facile Fluorometric Detection of Soybean Trypsin Inhibitor Activity with a Water-Soluble Poly(diphenylacetylene) Derivative

Evaluation of the Effective Cowpea [Vigna unguiculata (L.) Walp] Sprouting or Roasting for Pig Feeding

Background: The study investigated the effective roasting or sprouting of cowpeas (Vigna unguiculata) based on effects on in vitro digestibility (IVDMD), supported by measurement of key chemical components and trypsin inhibitor activity (TIA). Methods: The in vitro dry matter digestibility (IVDMD) of raw and all processed cowpeas were evaluated using standard, 3-step (gastric-ileal-colon) simulation of porcine digestion, modified for micro (0.5 g) sample digestion. Standard methods were employed to track processing effects on Ash, CP, Fat, ADF, NDF and trypsin inhibitor activity (TIA) at key processing points. Result: Sprouting for 2 and 3 days significantly decreased (p less than 0.05) gastric-ileal IVDMD while increasing (p less than 0.05) colon IVDMD. Total (steps 1-3) IVDMD increased (p less than 0.05) in 2-day (0.911) and 4-day (0.902) sprouts. The 20-minute cowpea roasting to 105°C terminal grain temperature resulted in high (p less than 0.05) step 3 and total IVDMD coefficients. The lowest (p greater than 0.05) total IVDMD was recorded in 15-minute (0.883) (95°C terminal grain temperature) roasts. In both experiments 1 and 2, quantitative changes in IVDMD were consistent with the changes in the chemical components (ADF, NDF, fat, CP, minerals) and trypsin inhibitor activity. In conclusion, the compartmental and total IVDMD and quantitative change in chemical components and TIA indicated 4 days sprouting and 20-minute (105°C) roasting were respectively most effective for cowpea processing.

Reduction of antinutrients and off-flavour in kidney bean flour by acidic and alkaline reactive extrusion

The presence of antinutrients and undesirable flavours in kidney bean flour poses challenges to consumer acceptance. Although extrusion can mitigate antinutrients to some extent, its impact on reducing beany flavour in bean flour remains underexplored. This study investigated the effects of injecting acetic acid or sodium carbonate solutions at three concentration levels (0.05, 0.1, 0.15 mol/L), in conjunction with three temperature profiles (40/60/80/80/90, 40/60/80/90/110, 50/70/90/110/130 °C) and two feed moisture levels (25, 30 %), on the removal of antinutrients (condensed tannins, trypsin inhibitor activity, phytic acid, raffinose family oligosaccharides) and reduction of volatile compounds that contribute to beany flavour in whole kidney bean flour. The results showed that all concentrations of acetic acid and sodium carbonate solutions effectively reduced condensed tannins compared to water, especially at 130 °C extrusion temperature. Introducing acetic acid and sodium carbonate solutions at a concentration of 0.15 mol/L led to 72 and 90 % reduction of total raffinose family oligosaccharide content, respectively, in contrast to the 17 % reduction observed with water alone. The incorporation of sodium carbonate solution reduced the total volatile compounds by 45–58 % as compared with water (23–33 %) and acetic acid (11–27 %). This reduction was primarily due to the reduction of aldehydes, alcohols, and aromatic hydrocarbons. These results indicate that injecting sodium carbonate solution during extrusion can effectively reduce antinutrients and beany flavour compounds in kidney bean flour.

Pellet die thickness and a commercial throughput agent interacted to demonstrate that high frictional heat increased apparent ileal amino acid digestibility, but did not influence trypsin inhibitor activity or male broiler performance

Pellet die thickness and a commercial throughput agent interacted to demonstrate that high frictional heat increased apparent ileal amino acid digestibility, but did not influence trypsin inhibitor activity or male broiler performance

Recovery of Proteases and Protease Inhibitors from Ganoderma spp. Cultivated in Amazonian Lignocellulose Wastes.

Ganoderma spp. are a great source of bioactive molecules. The production and recovery of bioactive molecules vary according to strain, growth substrate, and extraction solution. Variations in protease and their inhibitors in basidiomata from a commercial strain (G. lingzhi) and an Amazonian isolate (Ganoderma sp.) cultivated in Amazonian lignocellulosic wastes and extracted with different solutions are plausible and were investigated in our study. Basidiomata from cultivation in substrates based on açaí seed, guaruba-cedro sawdust and three lots of marupá sawdust were submitted to extraction in water, Tris-HCl, and sodium phosphate. Protein content, proteases, and protease inhibitors were estimated through different assays. The samples were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Fourier transform infrared spectroscopy with attenuated total reflectance (FTIR-ATR). Tris-HCl provided higher protein extraction from Ganoderma sp. and higher caseinolytic, gelatinolytic, and fibrinolytic activity for G. lingzhi cultivated in açaí. Water extracts of Ganoderma sp., in general, exhibited higher trypsin and papain inhibitor activities compared to G. lingzhi. Extracts in Tris-HCl and sodium phosphate showed more intense protein bands in SDS-- PAGE, highlighting bands of molecular weights around 100, 50, and 30 kDa. FTIR spectra showed patterns for proteins in all extracts, with variation in transmittance according to substrate and extractor. Water extract from Amazonian Ganoderma sp. cultivated in marupá wastes are promising as a source of protease inhibitors, while the Tris-HCL extract of G. lingzhi from açaí cultivation stands out as a source of proteases with fibrinolytic, caseinolytic, and gelatinolytic activities.

Chemical Composition, Microbiological and Sensory Qualities of Banana Based Complementary Diets Fortified with African Breadfruit Seed and Carrot Flours

About the age of six months, an infant requires more nutrients and energy than those provided by the breast milk, and complementary diets are therefore introduced to meet those needs. The objective of this study was therefore to develop and evaluate complementary diets from cooking banana, African breadfruit seeds and carrot flour blends. Ratios of cooking banana, African breadfruit seed and carrot flour blends in 90:0:10 as Diet 1, 85:5:10 as Diet 2, 80:10:10 as Diet 3, 75:15:10 as Diet 4 and 70:20:10 as Diet 5, were used for formulation of complementary diets. The flour blends for diet production showed bulk density and gelatinization temperature decreased from 0.98 to 0.65 g/ml and 85.67 to 78.10oC respectively, while swelling and water absorption capacity increased from 268.33 to 295.00% and 208.33 to 243.33% respectively, with increase in African breadfruit seed flour in the composite blends. Phytate (1.25 – 11.03 mg/100g) and trypsin inhibitor activities (1.05 – 1.17 TIU/100g) increased with increasing African breadfruit flour supplementations. Proximate composition of diets showed increased protein (2.16 – 18.47%), fat (1.81 – 2.23%), ash (2.42 – 2.96 %) and decreased carbohydrate (73.14 – 89.80%). Iron and calcium content of Diets increased from 14.11 – 19.23mg/100g and 28.24 – 43.23 mg/100g, while magnesium decreased from 6.24 – 8.25mg/100g respectively with increasing African breadfruit flour. Vitamin A and C decreased from 13.09 to 9.16 and 0.98 – 0.75mg/100g respectively. Microbial quality of all Diets developed were within the acceptable limit of 103 cfu/g for both total bacterial (TBC) and total fungal count (TFC). Diet 3 (80:10:10) was most accepted in sensory qualities. Complementary diets produced with 80% cooking banana, 10% African breadfruit seed and 10% carrot flour blends could help fill the gaps, when transitioning from breast milk to family diet, to provide the essential nutrients and energy required for infant’s activity, optimum growth and development.

Anti-SARS-CoV-2 Viral Activity of Sweet Potato Trypsin Inhibitor via Downregulation of TMPRSS2 Activity and ACE2 Expression In Vitro and In Vivo.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic. Known as COVID-19, it has affected billions of people worldwide, claiming millions of lives and posing a continuing threat to humanity. This is considered one of the most extensive pandemics ever recorded in human history, causing significant losses to both life and economies globally. However, the available evidence is currently insufficient to establish the effectiveness and safety of antiviral drugs or vaccines. The entry of the virus into host cells involves binding to angiotensin-converting enzyme 2 (ACE2), a cell surface receptor, via its spike protein. Meanwhile, transmembrane protease serine 2 (TMPRSS2), a host surface protease, cleaves and activates the virus's S protein, thus promoting viral infection. Plant protease inhibitors play a crucial role in protecting plants against insects and/or microorganisms. The major storage proteins in sweet potato roots include sweet potato trypsin inhibitor (SWTI), which accounts for approximately 60% of the total water-soluble protein and has been found to possess a variety of health-promoting properties, including antioxidant, anti-inflammatory, ACE-inhibitory, and anticancer functions. Our study found that SWTI caused a significant reduction in the expression of the ACE2 and TMPRSS2 proteins, without any adverse effects on cells. Therefore, our findings suggest that the ACE2 and TMPRSS2 axis can be targeted via SWTI to potentially inhibit SARS-CoV-2 infection.

Correction: Park et al. Identification of a Novel KTi-1 Allele Associated with Reduced Trypsin Inhibitor Activity in Soybean Accessions. Agriculture 2023, 13, 2070

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Effect of variety, growing location and year on composition, certain antinutritional factors, and functionality of faba beans (Vicia faba) grown in Canada

AbstractBackground and ObjectivesFaba bean is a rich source of proteins, carbohydrates, vitamins, and minerals. It serves as a staple food in Asia, Africa, and the Mediterranean region. Faba bean production in Canada has recently increased due to the interest in sustainable plant‐based proteins. Identifying faba bean varieties that can grow across different environments and provide good quality, nutrition, and functionality is important. This study aimed to determine the effect of variety, growing location, and year on the composition, certain anti‐nutrients, and functionality of the faba bean varieties grown in Canada.FindingsVariety, growing location, and year had significant effects on the protein content, crude fat, ash, phytic acid, stachyose, verbascose, trypsin inhibitor activity, and functional properties of faba beans, including oil emulsion and water holding capacity. Starch content, total dietary fiber, and least gelling concentration were significantly affected by variety and growing location, whereas raffinose, oil absorption capacity, foaming capacity, and stability were significantly affected by variety and growing year. Significant interactions of variety, growing location, and year were observed for most characteristics. Environment played a greater role in affecting faba bean characteristics than variety except for starch content, total dietary fiber, oligosaccharides, and foaming stability.ConclusionsIt was found that varietal and environmental factors affected each trait differently. Selecting suitable varieties and growing conditions would improve the quality of faba bean.Significance and NoveltyInformation from this study will be useful to breeders, growers, and food manufacturers to improve the production and utilization of faba beans.

Ultrasound-assisted germination of red kidney beans: Enhancements in physicochemical and nutritional profiles

To improve the conventional germination process and improve the nutritional quality of red kidney beans, this study employed high-intensity ultrasound (HIU) supplemented with hydrogen peroxide as a pre-germination treatment. The results showed that the 350 W-10 min treatment yielded the highest germination rate (77.09%), with its sprout length 81.13% greater than that of the control group. The 350 W-10 min treatment increased total protein, soluble protein, and ash content, while simultaneously reducing the fat, starch, and soluble sugar content. The HIU treatment accelerated the accumulation of phenolic and flavonoid compounds, ascorbic acid, and γ-aminobutyric acid. The 350 W-10 min treatment also decreased the levels of phytic acid, trypsin inhibitor activity, and tannin by 42.71%, 65.58%, and 53.18%, respectively. Furthermore, ultrasonic cavitation enhanced antioxidative capacity and improved amino acid composition and protein digestibility. Consequently, HIU serves as a cost-efficient method to accelerate the germination process and enhance their nutritional composition.

Bowman-Birk Inhibitor Mutants of Soybean Generated by CRISPR-Cas9 Reveal Drastic Reductions in Trypsin and Chymotrypsin Inhibitor Activities.

Despite the high quality of soybean protein, raw soybeans and soybean meal cannot be directly included in animal feed mixtures due to the presence of Kunitz (KTi) and Bowman-Birk protease inhibitors (BBis), which reduces animal productivity. Heat treatment can substantially inactivate trypsin and chymotrypsin inhibitors (BBis), but such treatment is energy-intensive, adds expense, and negatively impacts the quality of seed proteins. As an alternative approach, we have employed CRISPR/Cas9 gene editing to create mutations in BBi genes to drastically lower the protease inhibitor content in soybean seed. Agrobacterium-mediated transformation was used to generate several stable transgenic soybean events. These independent CRISPR/Cas9 events were examined in comparison to wild-type plants using Sanger sequencing, proteomic analysis, trypsin/chymotrypsin inhibitor activity assays, and qRT-PCR. Collectively, our results demonstrate the creation of an allelic series of loss-of-function mutations affecting the major BBi gene in soybean. Mutations in two of the highly expressed seed-specific BBi genes lead to substantial reductions in both trypsin and chymotrypsin inhibitor activities.

Enzymatic and Algebraic Methodology to Determine the Contents of Kunitz and Bowman-Birk Inhibitors and Their Contributions to Total Trypsin or Chymotrypsin Inhibition in Soybeans.

Soybeans are the number one source of plant proteins for food and feed, but the natural presence of protein protease inhibitors (PIs), namely, the Kunitz trypsin inhibitor (KTI) and the Bowman-Birk inhibitor (BBI), exerts antinutritional effects. This communication describes a new methodology for simultaneously quantitating all parameters of PIs in soybeans. It consists of seven steps and featured enzymatically measuring trypsin and chymotrypsin inhibitory activities, respectively, and subsequently determining the contents of reactive KTI and BBI and the contributions of each toward total PI mass and total trypsin or chymotrypsin inhibition by solving a proposed system of linear equations with two variables (C = dB + eK and T = xB + yK). This enzymatic and algebraic (EA) methodology was based on differential inhibitions of KTI and BBI toward trypsin and chymotrypsin and validated by applications to a series of mixtures of purified KTI and BBI, two KTI-null and two conventional soybeans, and by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The EA methodology allowed calculations of PI composition and the contributions of individual inhibitors toward total inhibition with ease. It was first found that although BBI constituted only about 30% of the total PI mass in conventional raw soybeans, it contributed about 80% toward total chymotrypsin inhibitor activity and about 45% toward trypsin inhibitor activity. Therefore, BBI caused more total protease inhibitions than those of KTI. Furthermore, the so-called KTI-null soybean mutants still contained measurable KTI content and thus should be named KTI-low soybeans.

Development of nutri-rich powder for pregnant/lactating women

Pregnancy and lactation are the two states, which need increased nutritional demand like protein, calcium, iron etc. in women. The study is part of the effort to provide nutritional food that can be more cost effective for pregnant/lactating women. To cater the elevated demand of nutrition, moth bean and finger millet were selected as major ingredients, being richer sources of iron and calcium respectively. The processing steps like soaking and germination helped to lower anti-nutritional factors. The best germination time was optimized for both, moth bean and finger millet based upon reduction in the phytic acid and trypsin inhibitor activity and increase in amylase activity with optimum yield. The best germination period selected for moth bean and finger millet were 36 h at 30ºC and 24 h at 30ºC respectively with 80% RH. The formulation was optimized using Response Surface Methodology at five levels of three variables viz. moth bean flour, finger millet flour and barley malt extract. The optimized product was prepared by mixing germinated and roasted moth bean (21.98%) and finger millet (19.37%) flours with skim milk and barley malt extract (20%), followed by cooking (100ºC, 5 min.), vacuum drying (70ºC, 700 mm Hg), grinding and sieving. The developed nutri-rich food contained 16.74g protein, 5.70 mg iron, 284.49 mg calcium and 1.8 mg zinc, which can provide very good nutrition as per recommendations by ICMR for pregnant/lactating women. The product cost was around 122 INR per kg and it was shelf stable up to 180 days of ambient storage.

Effects of Soaking and Germination Treatments on the Nutritional, Anti-Nutritional, and Bioactive Characteristics of Adzuki Beans (Vigna angularis L.) and Lima Beans (Phaseolus lunatus L.).

Lima beans (Phaseolus lunatus) and adzuki beans (Vigna angularis) are some of the most nutritious underutilized pulses that are significant in being used as basic ingredients for the preparation of various food products. The present study aimed to determine the impact of soaking and germination on nutritional and bioactive components, in vitro protein digestibility, reducing power, metal chelating capacity, antioxidant activity, and anti-nutritional components of lima and adzuki beans. The findings showed that during the germination treatment, the in vitro protein digestibility of lima and adzuki beans increased by 14.75 and 10.98%, respectively. There was an increase in the antioxidant activity of lima beans by 33.48% and adzuki beans by 71.14% after 72 h of germination, respectively. The reducing power assay of lima and adzuki beans indicated an increase of 49.52 and 36.42%, respectively, during germination. Similarly, the flavonoid and metal chelating activity increased in lima and adzuki beans after 72 h of germination. In contrast, the anti-nutrients, such as phytic acid, tannin content, and trypsin inhibitor activity, decreased significantly p < 0.05 after 72 h of germination. These results are encouraging and allow for utilizing the flour obtained from the germinated beans in functional bakery products, which can contribute to eradicating protein deficiency among some population groups. At the same time, promoting soaking and germination of the beans as a way to enhance the nutritional quality and reduce anti-nutrients can contribute to the interest in these underutilized pulses. They could be seen as an additional tool to improve food security.

Chemical constituents of the essential oils from Uvariodendron mbagoi

Several Uvariodendron species are known for their scented leaves and stem bark, hence this study. The leaves, twigs, stem and root barks of the newly described Uvariodendron species, U. mbagoi Dagallier and Couvreur from Kwedijela forest (KF) and Kimboza Forest Reserve (KFR) collected in dry and wet seasons, were steam distilled to yield essential oils that was more abundant in the root bark (2.07-3.85%) than in the leaves, twigs and stem bark (0.09-0.71%). The essential oils from the root bark collected from KF in the wet season (2.07%) was slightly lower than that obtained from the plant part collected from the same locality during the dry season (3.85%). GC-MS analysis indicated that linalool was the most prominent constituent of the leaf (82.18%, KF) and (98.86%, KFR), twig (89.66%, KFR), and stem bark (97.97%, KFR) oil while methyl eugenol (41.08%, KFR) and elemicin (39.28%, KFR) were the major constituents of the root bark oil. The essential oil from the leaves, stem bark, twigs and root bark of U. mbagoi exhibited mild to moderate potency when screened for antioxidant, antibacterial, and protease (trypsin) inhibitory activities. KEY WORDS: Uvariodendron mbagoi, Annonaceae, Essential oil, Linalool, Elemicin, Methyl eugenol Bull. Chem. Soc. Ethiop. 2024, 38(3), 1027-1035. DOI: https://dx.doi.org/10.4314/bcse.v38i4.17

Effects of thermal processing on the nutritional and antinutritional properties of African yam bean (Sphenostylis stenocarpa) seed flours

The study was carried out to evaluate the effects of thermal processing treatments on nutrient and antinutrient contents of African yam bean seed flours. The African yam bean seeds were sorted, cleaned and divided into five equal lots of one kilogram each. Four lots were processed into boiled, blanched, roasted and autoclaved African yam bean flours, while the last lot was processed raw and used as control. The flour samples obtained were analysed for proximate, mineral, vitamin and antinutrient contents using standard methods. The proximate composition of the samples revealed that the flours had a range of 6.14-11.24% moisture, 8.18-14.37% crude protein, 3.06-4.61% fat, 2.04–3.32% ash, 3.18–3.56% crude fibre, 62.90–76.98% carbohydrate and 350.57–368.50 kJ/100g energy, respectively. The mineral composition of the samples showed that the flours contained 128.81–174.16 mg/100g calcium, 88.86–212.20 mg/100g, potassium, 134.71–166.77 mg/100g phosphorus, 89.17 – 122.76 mg/100g, magnesium, 14.28–18-11 mg/100g iron and 3.24–5.59 mg/100g zinc, respectively. The vitamin composition of the flours were 1.15–1.37 mg/100g ascorbic acid, 1.15–135 mg/100g thiamine, 1.19–1.55 mg/100g niacin, 1.34–1.85 mg/100g riboflavin, 1.09–1.29 mg/100g folic acid, 1.37–1.95 mg /100g vitamin A and 1.27–1.66 mg/100g vitamin E, respectively. The results showed that the roasted and autoclaved African yam bean flours generally had higher crude protein, fat, ash, crude fibre, mineral and vitamin contents than the boiled and blanched flour samples compared to the raw sample. The antinutrient composition of the flours also showed that the levels of trypsin inhibitor activity, tannin, phytate, oxalate, saponin and haemagglutinin of the samples were significantly (p&lt;0.05) reduced by boiling, autoclaving roasting and blanching treatments compared to the raw sample. However, the study revealed that the processed African yam bean flours have the potentials to be used as nutrient dense ingredients in the preparation of a wide range of food products than the raw sample especially in both underdeveloped and developing countries where the problems of protein-energy malnutrition and micronutrients deficiencies are prevalent.

Cutaneous glands of the striped toad, Rhinella crucifer (Wied-Neuwied, 1821) (Amphibia: Bufonidae): Histological study and bioactivities of glandular secretions

This study investigated the morphology of Rhinella crucifer cutaneous glands, as well as the protein/peptide profiles and bioactivities of body gland secretions (BGS) and parotoid macrogland secretions (PS). The parotoid as well as dorsal and ventral skin fragments of male and female individuals were processed for histological analysis. The protein and peptide profiles of male and female gland secretions were evaluated. Male secretions were also assessed for proteolytic, trypsin inhibiting, hemagglutinating, hemolytic, antimicrobial, and anticoagulant activities. The R. crucifer skin structure presented protuberances that are clearly visible and formed by the integument, which has cutaneous glands throughout the body. An average of 438 and 333 glands were identified in males in females, respectively. No significant differences were observed in the distribution of glands across the body as well as for area and perimeter of glands. Differences were observed in protein composition between the PS and BGS from males and females, and secretions from animals collected from undisturbed and anthropogenically disturbed areas. Proteins with similarities to catalase and elongation factor 1-alpha were detected in the PS. Zymography revealed proteolytic activity in both male BGS and PS. Male BGS showed antibacterial activity against Enterococcus faecalis and Escherichia coli and anticoagulant activity, being able to prolong prothrombin time by 6.34-fold and activated partial thromboplastin time by 2.17-fold. Finally, male PS and BGS caused a maximum hemolysis degree of 1.4%. The data showed that the cutaneous secretions of R. crucifer are potentially promising for biotechnological prospecting.