A Structural In Silico Analysis of the Immunogenicity of L-Asparaginase from Penicillium cerradense.

L-asparaginase is an essential drug used to treat acute lymphoid leukemia (ALL), a cancer of high prevalence in children. Several adverse reactions associated with L-asparaginase have been observed, mainly caused by immunogenicity and allergenicity. Some strategies have been adopted, such as searching for new microorganisms that produce the enzyme and applying protein engineering. Therefore, this work aimed to elucidate the molecular structure and predict the immunogenic profile of L-asparaginase from Penicillium cerradense, recently revealed as a new fungus of the genus Penicillium and producer of the enzyme, as a motivation to search for alternatives to bacterial L-asparaginase. In the evolutionary relationship, L-asparaginase from P. cerradense closely matches Aspergillus species. Using in silico tools, we characterized the enzyme as a protein fragment of 378 amino acids (39 kDa), including a signal peptide containing 17 amino acids, and the isoelectric point at 5.13. The oligomeric state was predicted to be a homotetramer. Also, this L-asparaginase presented a similar immunogenicity response (T- and B-cell epitopes) compared to Escherichia coli and Dickeya chrysanthemi enzymes. These results suggest a potentially useful L-asparaginase, with insights that can drive strategies to improve enzyme production.

Antiproliferative and Apoptotic Effects of Murici (Byrsonima crassifolia (L.) Kunth and verbascifolia (L.) DC) and Taperebá (Spondias mombin L.) Extracts in Human Prostate Cell Line (PC-3).

OBJECTIVE: The present study aimed to evaluate the antiproliferative and apoptotic effects of extracts obtained from the murici (Byrsonima crassifolia (L.) Kunth and verbascifolia (L.) DC) and taperebá (Spondias mombin L.) pulps, on cell proliferation, cell cycle and apoptosis on human prostate cell line (PC-3). METHODS: Four extract was produced from the pulps:  murici aqueous extract (MA), taperebá aqueous extract (TA), murici ethanolic extract (ME) and taperebá ethanolic extract (TE). In the present study, the analysis of cell viability, cell cycle and apoptosis analyze were performed using the MTT method and flow cytometry. RESULTS: The results showed that murici and taperebá extracts proved to be inhibitors of cell growth, modulation of cell cycle promoters and capable of enhancing the death in prostate carcinoma cells PC-3; suggesting a regulatory effect in prostate cell line, depending on type of extract and dosage used. CONCLUSION: These results open a series of perspectives on the use of these bioactive extracts in the prevention and treatment of prostate cancer.

Heterologous expression of fungal L-asparaginase: a systematic review.

Aim: To review the available literature about heterologous expression of fungal L-asparaginase (L-ASNase). Materials & methods: A search was conducted across PubMed, Science Direct, Scopus and Web of Science databases; 4172 citations were identified and seven articles were selected. Results: The results showed that heterologous expression of fungal L-ASNase was performed mostly in bacterial expression systems, except for a study that expressed L-ASNase in a yeast system. Only three publications reported the purification and characterization of the enzyme. Conclusion: The information reported in this systematic review can contribute significantly to the recognition of the importance of biotechnological techniques for L-ASNase production.

Asparaginase is a common treatment for the most common type of leukemia in children. These treatments generally use asparaginase sourced from bacteria. Some people can experience bad reactions to these treatments. One way that has been explored to avoid this is to use asparaginase sourced from fungi because they are more similar to humans. However, fungi produce less asparaginase than bacteria. This review looks into ways that the production of fungal asparaginases can be made more productive.

Food Service Kitchen Scraps as a Source of Bioactive Phytochemicals: Disposal Survey, Optimized Extraction, Metabolomic Screening and Chemometric Evaluation.

Untargeted metabolomics is a powerful tool with high resolution and the capability to characterize a wide range of bioactive natural products from fruit and vegetable by-products (FVB). Thus, this approach was applied in the study to evaluate the phenolic compounds (PC) by metabolomic screening in five FVB after optimizing their extraction. The total phenolic content and antioxidant activity analyses were able to select the best extractor (SM) and ultrasonication time (US) for each FVB; methanol was used as a control. Although ultrasonication yielded a lower number of PC identifications (84 PC), the US extract was the most efficient in total ionic abundance (+21% and +29% compared to the total PC and SM extracts, respectively). Ultrasonication also increased the phenolic acid (+38%) and flavonoid classes (+19%) extracted compared to SM, while the multivariate analyses showed the control as the most dissimilar sample. FVB extracted from the same parts of the vegetable/fruit showed similarities and papaya seed presented the most atypical profile. The application of the metabolomics approach increased the knowledge of the bioactive potential of the evaluated residues and possibilities of exploring and valorizing the generated extracts.

Probiotic fermented whey-milk beverages: Effect of different probiotic strains on the physicochemical characteristics, biological activity, and bioactive peptides.

The effect of probiotic strains (Lactobacillus acidophilus La-03 (La-03); Lactobacillus acidophilus La-05 (La-05); Bifidobacterium Bb-12 (Bb-12) or Lacticaseibacillus casei-01 (L. casei-01)) on the characteristics of fermented whey-milk beverages during storage (4 °C, 30 days) was evaluated. The products were assessed for biological and antioxidant activities, physicochemical characteristics, and bioactive peptides. Probiotic addition increased α-amylase and α-glucosidase inhibition and antioxidant activities, mainly at 15 days of storage. L. casei-01 showed higher metabolic activity (higher titratable acidity and lower pH values) and the presence of anti-hypertensive peptides, while La-5 and Bb-12 showed higher α-glucosidase inhibition, improvements in the high saturated hypercholesterolemic index, and peptides with ACE-inhibitory, antimicrobial, immunomodulatory, and antioxidant activities. Our findings suggest that probiotic fermented whey-milk beverages may exert antidiabetic and antioxidant properties, being suggested La-5 or Bb-12 as probiotics and 15 days of storage.

Evaluation of the antioxidant capacity, volatile composition and phenolic content of hybrid Vitis vinifera L. varieties sweet sapphire and sweet surprise.

Bioactive compounds were extracted using two different extraction solvents (acetone and water) from pulp and whole grape berries derived from hybrid Vitis vinifera L. varieties Sweet sapphire (SP) and Sweet surprise (SU) and were characterised based on a comprehensive metabolomic approach by chromatography coupled with mass spectrometry (UPLC-QTOF-MSE and GC-FID/MS). GC-FID/MS analysis was performed with two different extraction methods (solvent extraction method and solid-phase extraction). Anthocyanins were characterised and quantified by HPLC-UV. The antioxidant potential was assessed by different assays. SP acetone extract from grape skin had the highest mean to DPPH, FRAP, ORAC and phenolic content SP samples, also showed higher anthocyanin content. Globally, 87 phenolic compounds were identified. The relative quantification by UPLC-MSE showed flavonoids the most abundant class. Forty two compounds were found in the volatile fraction of SU, while only thirty one volatile compounds were found in the SP samples.

Antiproliferative Activity on Human Colon Adenocarcinoma Cells and In Vitro Antioxidant Effect of Anthocyanin-Rich Extracts from Peels of Species of the Myrtaceae Family.

There is a significant indication of the beneficial health effects of fruit rich diets. Fruits of native plant species have noticeably different phytochemicals and bioactive effects. The aim of this work was to characterize and compare the constituents of jabuticaba (Myrciaria jaboticaba, MJ), jamun-berry (Syzygium cumini, SC), and malay-apple (Syzygium malaccense, SM) extracts and their influence on antioxidant activity in vitro and antiproliferative effects on human colon adenocarcinoma cells. According to the results, dried peel powders (DP) have a high anthocyanin content, phenolic compounds, and antioxidant activity when compared to freeze dried extracts (FD). M. jaboticaba dried peel powder extract had a higher total anthocyanin and phenolic compounds content (802.90 ± 1.93 and 2152.92 ± 43.95 mg/100 g, respectively). A reduction in cell viability of HT-29 cells after treatment with M. jaboticaba extracts (DP-MJ and FD-MJ) was observed via MTT assay. Flow cytometry showed that the treatment with the anthocyanin-rich extracts from MJ, SC, and SM had an inhibitory impact on cell development due to G2/M arrest and caused a rise in apoptotic cells in relation to the control group. The findings of this study highlight the potential of peel powders from Myrtaceae fruits as an important source of natural antioxidants and a protective effect against colon adenocarcinoma.

Effect of grape juice, red wine and resveratrol solution on antioxidant, anti-inflammatory, hepactic function and lipid profile in rats feds with high-fat diet.

The objective was to investigate the role of consumption of grape juice (GJ), red wine (RW) or resveratrol solution (RS) on rats treated with a high-fat diet (HFD). Among the drinks offered, GJ had lower content of polyphenols and trans-resveratrol. Nevertheless, GJ showed similar content of anthocyanin and antioxidant activity to RW, although higher than RS. In rats treated with HFD, consumption of GJ presented best antioxidant and anti-inflammatory activity, reducing glutathione peroxidase and interleukin-6 serum levels. In addition, GJ promoted better levels of cholesterol and liver markers. On the other hand, RW aggravated the oxidizing effect of HFD, increasing catalase activity and interleukin-6 level. Already, RS showed no benefit in animals. Thus, GJ minimized the effects of HFD on oxidative stress and inflammation beyond promoted better levels of lipid profile and liver biomarkers. However, consumption of RS showed no benefit and RW revealed a pro-oxidant effect, as did HFD. [Figure: see text].

Antiproliferative and apoptotic effects of probiotic whey dairy beverages in human prostate cell lines.

The present study aimed to evaluate the antiproliferative and apoptotic effects of probiotic whey dairy beverages in human prostate cancer cell lines (PC-3 and DU-145). Five different whey beverages were manufactured: conventional whey beverage (without addition of probiotic strain, CTL); and whey beverages containing Lactobacillus acidophilus La-05, Lactobacillus acidophilus La-03, Lactobacillus casei-01, and Bifidobacterium animalis Bb-12. Cell viability was determined by MTT assay and cell cycle arrest, and apoptosis by flow cytometry. All the samples presented cytotoxic activities against both cell lines. A decrease in the percentage of PC-3 cells in G0/G1 and S, followed by an increase in G2/M phase were observed with L. casei-01, Bb-12 and La-05 beverages (50.0 and 100.0 µg/mL). The extracts of the whey beverages caused extensive apoptosis induction in both cells' lines, regardless of the probiotic strain. However, the whey beverage added with L. casei-01 might be a better candidate against prostate cancer cells.

Antiproliferative effect of guava fruit extracts in MDA-MB-435 and MCF-7 human breast cancer cell lines.

Breast cancer is the most frequent and lethal neoplastic disease among women worldwide. Psidium Guajava is a promising functional food against cancer, owing to a variety of bioactive compounds. This study aimed to evaluate the anticarcinogenic potential of Pedro Sato (PS), Hitigio (HI) and Tsumori (TS) guava cultivars fruit pulp extracts in MDA-MB-435 and MCF-7 human breast cancer cells. The antioxidant capacity of the extracts and their effect on cell viability, cell cycle and apoptosis were assessed. Additionally, the concentration of carotenoids, total phenolics, ascorbic acid and other physicochemical parameters were evaluated. PS pulp extract showed the highest in vitro antioxidative activity by all tested methods, as well as the highest content of lycopene and total phenolics, while TS pulp extract presented the highest concentration of ß-carotene. After 48 hours treatment, all guava cultivars' extracts caused reduction of MDA-MB-435 and MCF-7 cells viability, with PS and HI being the most effective extracts. All guava extracts caused MDA-MB-435 and MCF-7 cell count reduction in G0/G1 and G2/M phases and increased apoptosis. The present results strongly suggest that guava pulp exerts antiproliferative effect on breast adenocarcinoma cells.

Fruit-based drink sensory, physicochemical, and antioxidant properties in the Amazon region: Murici (Byrsonima crassifolia (L.) Kunth and verbascifolia (L.) DC) and tapereba (Spondia mombin).

Increased fruit consumption due its protective effect on the organism is accompanied by the development of the processing industry of these products. The aim of this work was to optimize fruit pulp-based beverage formulations from the murici and tapereba Amazon region, taking into account their sensory acceptance and antioxidant activity. Total soluble solid content, reducing sugar content, titratable acidity contents, pH, and ascorbic acid content were determined in pulps and formulations. The total content phenolic compounds and antioxidant activity were also evaluated. A 22 factorial experiment was formulated to optimize ingredients for the production of murici and tapereba fruit drinks. The murici pulp had higher acidity and higher ascorbic acid content. The analysis of phenolic compounds and antioxidant activity presented higher quantity in tapereba pulp. Tapereba-based beverages had better acceptance by the evaluated criteria. Fruit-based beverages murici and tapereba are a well-accepted product and have important nutritional characteristics.

Metabolite Profiling by UPLC-MSE, NMR, and Antioxidant Properties of Amazonian Fruits: Mamey Apple (Mammea Americana), Camapu (Physalis Angulata), and Uxi (Endopleura Uchi).

The metabolite profiling associated with the antioxidant potential of Amazonian fruits represents an important step to the bioactive compound's characterization due to the large biodiversity in this region. The comprehensive bioactive compounds profile and antioxidant capacities of mamey apple (Mammea americana), camapu (Physalis angulata), and uxi (Endopleura uchi) was determined for the first time. Bioactive compounds were characterized by ultra-performance liquid chromatography coupled to high resolution mass spectrometry (UPLC-MSE) in aqueous and ethanolic extracts. Globally, a total of 293 metabolites were tentatively identified in mamey apple, campau, and uxi extracts. The main classes of compounds in the three species were terpenoids (61), phenolic acids (58), and flavonoids (53). Ethanolic extracts of fruits showed higher antioxidant activity and total ion abundance of bioactive compounds than aqueous. Uxi had the highest values of phenolic content (701.84 mg GAE/100 g), ABTS (1602.7 µmol Trolox g-1), and ORAC (15.04 µmol Trolox g-1). Mamey apple had the highest results for DPPH (1168.42 µmol TE g-1) and FRAP (1381.13 µmol FSE g-1). Nuclear magnetic resonance (NMR) spectroscopy results showed that sugars and lipids were the substances with the highest amounts in mamey apple and camapu. Data referring to chemical characteristics and antioxidant capacity of these fruits can contribute to their economic exploitation.

Effects of cooking on the phytochemical profile of breadfruit as revealed by high-resolution UPLC-MSE.

BACKGROUND: This study evaluated the impact of cooking on the profile of bioactive compounds in unripe breadfruit. To this end, the accessibility of bioactive compounds by various solvents was assessed through total phenolic content and antioxidant capacity analyses. The most efficient solvent was applied to extract the metabolites, which were evaluated by ultra-performance liquid chromatography coupled with high-resolution quadrupole time-of-flight mass spectrometry in MSE mode. RESULTS: Cooked and raw breadfruit presented total phenolic content and antioxidant capacities in almost all extracts, and pure water proved to be the best extractor. Globally, 146 bioactive compounds have been identified for both raw and cooked fruits' aqueous extracts. Most of these compounds were stable to the heat treatment applied (121 °C/10 min). However, results revealed that 22 metabolites contributed to significantly distinguishing the raw from the cooked samples. Among those, 15 compounds, such as pyrogallol, 1-acetoxypinoresinol, and scopolin, evidenced higher relative abundance in the cooked extracts. On the other hand, only seven metabolites, such as 4-hydroxyhippuric acid, epicatechin, and leptodactylone, decreased post-heating. CONCLUSIONS: Cooking promoted little alteration in the bioactive compounds profile of immature breadfruit and thus appears to be an exploitation alternative for this perishable fruit, which seems to be a source of a large range of bioactive compounds. © 2019 Society of Chemical Industry.

Amazon Fruits Inhibit Growth and Promote Pro-apoptotic Effects on Human Ovarian Carcinoma Cell Lines.

Murici (Byrsonima crassifolia (L.) Kunth and B. verbascifolia (L.) DC.) and tapereba (Spondias mombin) are Amazonian fruits that contain bioactive compounds. Biochemical and molecular characterization of these fruits can reveal their potential use in preventing diseases, including cancer. The extracts were characterized regarding the presence and profile of carotenoids by high-performance liquid chromatography (HPLC), total phenolic content by the Folin-Ciocalteu assay, and antioxidant activity by antioxidant value 2,2-diphenyl-1-picrylhydrazyl (DPPH) content analysis, 22,20-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) content analysis, Ferric-Reducing Ability of Plasma (FRAP), and Oxygen Radical Absorbance Capacity (ORAC) analysis. The extracts of tapereba and murici studied were important sources of total carotenoids and lutein, respectively. The extracts were then tested for their effect on the viability of the A2780 ovarian cancer (OC) cell line and its cisplatin (CDDP)-resistant derived cell line, called ACRP, by using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. Their influence on cell cycle and apoptosis were analyzed by using flow cytometry. Murici and tapereba cell extracts exhibited a strong bioactivity by inhibiting A2780 and ACRP cell viability by 76.37% and 78.37%, respectively, besides modulating the cell cycle and inducing apoptotic cell death. Our results open new perspectives for the development of innovative therapeutic strategies using these Amazon fruit extracts to sensitize ovarian cancer cells to current chemotherapeutic options.

Nicotiana benthamiana is a suitable transient system for high-level expression of an active inhibitor of cotton boll weevil α-amylase.

BACKGROUND: Insect resistance in crops represents a main challenge for agriculture. Transgenic approaches based on proteins displaying insect resistance properties are widely used as efficient breeding strategies. To extend the spectrum of targeted pathogens and overtake the development of resistance, molecular evolution strategies have been used on genes encoding these proteins to generate thousands of variants with new or improved functions. The cotton boll weevil (Anthonomus grandis) is one of the major pests of cotton in the Americas. An α-amylase inhibitor (α-AIC3) variant previously developed via molecular evolution strategy showed inhibitory activity against A. grandis α-amylase (AGA). RESULTS: We produced in a few days considerable amounts of α-AIC3 using an optimised transient heterologous expression system in Nicotiana benthamiana. This high α-AIC3 accumulation allowed its structural and functional characterizations. We demonstrated via MALDI-TOF MS/MS technique that the protein was processed as expected. It could inhibit up to 100% of AGA biological activity whereas it did not act on α-amylase of two non-pathogenic insects. These data confirmed that N. benthamiana is a suitable and simple system for high-level production of biologically active α-AIC3. Based on other benefits such as economic, health and environmental that need to be considerate, our data suggested that α-AIC3 could be a very promising candidate for the production of transgenic crops resistant to cotton boll weevil without lethal effect on at least two non-pathogenic insects. CONCLUSIONS: We propose this expression system can be complementary to molecular evolution strategies to identify the most promising variants before starting long-lasting stable transgenic programs.

Effect of Roasting Levels and Drying Process of Coffea canephora on the Quality of Bioactive Compounds and Cytotoxicity.

Coffee is a popular drink consumed all over the world. Besides its long-recognized stimulant effect, it has important nutritional and health effects. However, the type of bean processing modifies the composition of brewed coffee and possibly its bioactivity. In this study, extracts obtained from green and roasted beans of Coffea canephora (Coffea canephora var. robusta) were submitted to spray- or freeze-drying and were tested for antiproliferative activity, using MTT assay, and their influence on the cell cycle and apoptosis by flow cytometry analysis. Moreover, colors and nutrient contents were measured to identify the changes due to the roasting process. The results obtained showed that extracts from green and light roasted beans exhibited strong bioactive capacity. Coffee extracts promoted a decrease in cell viability, modulated cell cycle and induced apoptosis in human prostate carcinoma cell line (DU-145). The level of roasting reduced this property, but the type of drying did not in all cases.

Açai (Euterpe oleracea Mart.) Seed Extract Induces Cell Cycle Arrest and Apoptosis in Human Lung Carcinoma Cells.

Açai fruit has been studied for its antioxidant properties, with positive feedback against many diseases, including cancer. Although açai seeds are not edible, their composition has been studied in order to find new applications and reduce garbage generation. This study aimed to evaluate the cytotoxic effects and impacts on the cell cycle and apoptosis of açai seed extract (ASE) on human lung carcinoma cell line (A549). Antioxidant activity of açai seed extract (ASE) was measured by DPPH assay, Trolox Equivalent Antioxidant Capacity (ABTS/TEAC), Ferric Reducing Ability (FRAP) and Oxygen radical absorbance capacity (ORAC) assays. Human lung carcinoma cell viability (A549) was monitored by MTT assay method and the effects on cell cycle and apoptosis were measured by flow cytometry. The results indicate high antioxidant activity in ASE and high values of total phenolic compounds (37.08 ± 8.56 g gallic acid/100 g). The MTT assay showed a maximum decrease (72.07%) in the viability of A549 cells after 48 h treatment with ASE (200 µg/mL). Flow cytometer analysis revealed that ASE increased the percentage of cells in G0/G1 phase and promoted a high increase of apoptotic cells when compared to the untreated cells. The present study suggests that ASE has a high antioxidant capacity and may have a protective effect against lung cancer.

Antioxidant Activity and Cytotoxicity Effect of Cocoa Beans Subjected to Different Processing Conditions in Human Lung Carcinoma Cells.

Lung cancer is a common malignancy in men and the second leading cause of cancer-related mortality in men in the western world. Phenolic cocoa ingredients have a strong antioxidative activity and the potential to have a protective effect against cancer. In the present study, we have evaluated the influence of cocoa beans subjected to different processing conditions on cell viability and apoptosis of human lung cancer cells (A549). We measured the viability of lung cells treated with cocoa beans, unroasted slates (US), roasted slates (RS), unroasted well fermented (UWF) cocoa, and roasted well fermented (RWF) cocoa for 24 h. Using an MTT assay, we observed a decrease in the viability of A549 cells after treatment with cocoa bean extracts. Flow cytometer analysis revealed that cocoa beans increased the percentage of cells in sub-G1 phase and promoted up to twofold increase of apoptotic cells when compared to the control group. Taken together, the present study suggests that cocoa beans may have a protective effect against lung cancer.