Vero Cell Cytotoxicity Research Articles

Vernonia amygdalina Leaf Extract Induces Apoptosis in HeLa Cells: A Metabolomics and Proteomics Study.

Medicinal plants produce various bioactive molecules with potential anti-cancer properties with favorable safety profiles. We aimed to investigate the comprehensive composition of Vernonia amygdalina leaf extract and its cytotoxic effects via apoptosis in HeLa cells. The metabolomics approach using LC-MS/MS was conducted to gather the metabolite profile of the extract. Proteomics was performed to understand the comprehensive mechanistic pathways of action. The apoptosis was visualized by cellular staining and the apoptotic proteins were evaluated. V. amygdalina leaf extract exhibited dose-dependent cytotoxic effects on both HeLa and Vero cells after 24 h of exposure in the MTT assay with the IC50 values of 0.767 ± 0.0334 and 4.043 ± 0.469 µg mL-1, respectively, which demonstrated a higher concentration required for Vero cell cytotoxicity. The metabolomic profile of 112 known metabolites specified that the majority of them were alkaloids, phenolic compounds, and steroids. Among these metabolites, deacetylvindoline and licochalcone B were suggested to implicate cytotoxicity. The cytotoxic pathways involved the response to stress and cell death which was similar to doxorubicin. The upstream regulatory proteins, phosphatase and tensin homolog deleted on chromosome ten (PTEN) and X-box binding protein 1 (XBP1), were significantly altered, supporting the regulation of apoptosis and cell death. The levels of apoptotic proteins, c-Jun N-terminal kinases (JNK), p53, and caspase-9 were significantly increased. The novel insights gained from the metabolomic profiling and proteomic pathway analysis of V. amygdalina leaf extract have identified crucial components related to apoptosis induction, highlighting its potential to develop future chemotherapy.

Synthesis, in vitro and in silico evaluation of gallamide and selenogallamide derivatives as inhibitors of the SARS-CoV-2 main protease.

The present work reports the inhibitory effect of amides derived from gallic acid (gallamides) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (Mpro), along with cytotoxicity evaluation and molecular docking studies. In addition to gallamides, other relevant compounds were also synthesized and evaluated against Mpro, making a total of 25 compounds. Eight compounds presented solubility issues during the inhibitory assay and one showed no inhibitory activity. Compounds 3a, 3b, and 3f showed the highest enzymatic inhibition with IC50 = 0.26 ± 0.19 µM, 0.80 ± 0.38 µM, and 2.87 ± 1.17 µM, respectively. Selenogallamide 6a exhibited IC50 values of 5.42 ± 2.89 µM and a comparison with its nonselenylated congener 3c shows that the insertion of the chalcogen moiety improved the inhibitory capacity of the compound by approximately 10 times. Regarding the cellular toxicity in THP-1 and Vero cells, compounds 3e and 3g, showed moderate cytotoxicity in Vero cells, while for THP-1 both were nontoxic, with CC50 > 150 µM. Derivative 3d showed moderate cytotoxicity against both cell lines, whereas 6d was moderatly toxic to THP-1. Other compounds analyzed do not induce substantial cellular toxicity at the concentrations tested. The molecular docking results for compounds 3a, 3b, and 3f show that hydrogen bonding interactions involving the hydroxyl groups (OH) of the gallate moiety are relevant, as well as the carbonyl group.

In Vitro Antiprotozoal Activity of Schinus molle Extract, Partitions, and Fractions against Trypanosoma cruzi.

Chagas disease, caused by the protozoan Trypanosoma cruzi, represents an important and worldwide public health issue, particularly in Latin America. Limitations of conventional treatment with benznidazole and nifurtimox underscore the urgent need for new therapeutic strategies for this disease. Schinus molle, a tree used in traditional medicine for various ailments, has demonstrated promising antiparasitic activity. The in vitro anti-T. cruzi activity of Schinus molle crude methanol extract, partitions, and fractions, as well as their cytotoxicity in Vero cells and Artemia salina, and hemolytic activity in human erythrocytes were assessed. Most of the extracts possessed anti-T. cruzi effects, with Sm-CF3 being the fraction with the highest activity (IC50 = 19 µg/mL; SI = 6.8). Gas chromatography-mass spectrometry analysis identified 20 compounds, with fatty acyls comprising the predominant chemical class (55%). We also identified the antiparasitic compounds cis-5,8,11,14,17-eicosapentaenoic acid and trans-Z-α-bisabolene epoxide, suggesting their potential contribution to the observed anti-T. cruzi activity. In conclusion, our findings support the therapeutic potential of S. molle as a source of novel antiparasitic agents against T. cruzi.

5-Fluoroindole Reduces the Bacterial Burden in a Murine Model of Mycobacterium tuberculosis Infection.

Tuberculosis is a disease caused by a single pathogen that leads to a death toll estimated to be more than a million per year. Mycobacterium tuberculosis (Mtb), which affects mainly the lungs, spreads by airborne transmission when infectious respiratory particles from an infected human enter the respiratory tract of another person. Despite diagnosis and treatment being well established, the rise of cases of patients infected with Mtb strains with multidrug resistance to the antibiotics used in the regimen against the disease is alarming. Indole used as a core molecule has been described as a promising structure to treat several diseases. 5-Fluoroindole (5-FI) compound, evaluated in the free base and in the hydrochloride (5-FI.HCl) forms, inhibited the growth of pan-sensitive Mtb H37Rv strain in the same range (4.7-29.1 μM) of clinical isolates that have resistance to at least two first-line drugs. Although 5-FI showed no cytotoxicity in Vero and HepG2 cells, high permeability (2.4.10-6 cm/s) in the PAMPA assay, and high metabolic stability (Clint 9.0 mL/min/kg) in rat liver microsomes, limited solubility at plasmatic and intestinal pH values prompted formation and employment of its salt form (5-FI.HCl). Although the 5-FI.HCl compound showed increased solubility at pH values of 7.4 and 9.1 and increased stability in aqueous solutions, data for intrinsic clearance (Clint = 48 mL/min/kg) and a half-life (t 1/2 = 12 min) showed decreased metabolic stability. As 5-FI.HCl showed both good absorption and ability to reach the systemic circulation of animals without the need to use vehicles containing cosolvents or surfactants, it was chosen to evaluate its effectiveness in the model of tuberculosis in mice. The in vivo results showed the concentration of the compound in plasma increasing within 30 min in the systemic circulation and the capacity of reducing the Mtb burden in the lungs at the concentration of 200 μmol/kg after 21 days of infection, with no toxicity in mice.

Novel 1,3-Indanedione-Thiazole Hybrids as Small-Molecule SARS-COV-2 Main Protease Inhibitors With Potential anti-Coronaviral Activity

Since arising in 2019, COVID 19 has been causing rapidly-increasing mortality and morbidity rates across the globe. Herein, novel 1,3-indanedione-thiazole hybrids were designed and synthesized as small-molecule SARS-COV-2 Main protease (Mpro) inhibitors with potential anti-covid activity. All target compounds were screened in vitro for their ability to inhibit SARS-COV-2 Mpro. Several compounds displayed potent SARS-COV-2 Mpro inhibition at one-digit IC50 values ranging from 4.3 to 9.9 µM, compared to ritonavir (IC50= 2.4 µM). Moreover, antiviral assay revealed the ability of compounds 12c, 12f, and 16a to significantly inhibit the replication of SARS-COV-2 in Vero cells at EC50 values of 7.79, 2.79 and 1.65 µM, respectively, relative to ritonavir (EC50 = 1.72 µM). Cytotoxicity assay was also conducted. None of the tested compounds exhibited significant cytotoxicity in Vero cells showing CC50 values from 171.77 to 299.96 µM and SI from 38.5 to 178.6 µM. In addition, a docking study revealed proper orientation and well-fitting of title compounds into the binding pocket of SARS-COV-2 Main protease.

The palm oil-based chlorophyll removal and the evaluation of antiaging properties on Centella asiatica ethanolic extract

Centella asiatica is an essential medicinal herb in numerous traditions. The high chlorophyll concentration of C. asiatica makes dosage form formulation challenging. On the other hand, palm oil is feasible to eliminate the chlorophyll from plant-based products. This research aim to analysis the phytochemical profile and in vitro anti-aging effects on chlorophyll removal of C. asiatica extract. The extract was soxhletated in 70% ethanol. The extract was partitioned with 50% ethanol and palm oil to create a dechlorophyllation extract. Asiaticoside was analyzed using TLC-densitometry. The cytoprotective effect of H2O2-induced oxidative stress in Vero cells was assessed using MTT assays. Flow cytometer analysis was used for quantifying the intracellular ROS. The senescence-associated-β-galactosidase assay was used to identify senescent cells. The efficiency of chlorophyll removal by palm oil was 90.94±0.62%. The dechlorophyllation extract (DE) showed a concentration of 1.58±0.02% for asiaticoside, but it was not found in the original extract (OE). DE significantly reduced H2O2-induced cytotoxicity in Vero cells compared to OE. Both DE and OE reduced intracellular ROS and senescent cells. Palm oil-based chlorophyll removal improved the phytochemical content and anti-aging effects of C. asiatica in ethanolic extract.

Comparative phytochemistry using UPLC-ESI-QTOF-MS phenolic compounds profile of the water and aqueous ethanol extracts of Tagetes minuta and their cytotoxicity

The complexity of secondary metabolites in different medicinal plants, including Tagetes minuta, defines the general properties and uses of the plant. Tagetes minuta is reportedly a valuable medicinal plant that has long been used as a spice herb, medicine, a natural herbicide for weeds, and pest control in agriculture. In this study, the secondary metabolite profiles of the water and aqueous ethanol extracts of T. minuta were profiled through identification and quantification using colorimetric assays and a Waters Synapt G2 Quadrupole time-of-flight (QTOF) mass spectrometer (MS) connected to a Waters Acquity ultra-performance liquid chromatograph (UPLC). The antioxidant capacity of the extracts was evaluated with 2,2-Azino-di-3-ethylbenzthiazoline sulfonate and ferric reducing antioxidant power, as well as their toxicity on HepG2 (cancerous) and Vero (non-cancerous) cell lines. The major classes of secondary metabolites identified and quantified were phenolic acids (benzoic acids and hydroxycinnamic acids), flavonoids (chalcones, flavonols, flavanols, and flavones), fatty acids, coumarins and furanocoumarins. The bioavailability of these secondary metabolites is influenced by the polarity of the solvent of extraction. Extracts from aqueous ethanol showed higher secondary metabolites, corresponding to antioxidant content and activity as well as cytotoxicity in HepG2 and Vero cells, compared to the water extract. This study presents a comprehensive phytochemical knowledge of secondary metabolites, primarily phenolic compounds, identified in T. minuta, which can pave the way for future studies on the pharmacological importance and standardisation of the medicinal uses of the plant.

Targeting the Schistosoma mansoni nutritional mechanisms to design new antischistosomal compounds

The chemical classes of semicarbazones, thiosemicarbazones, and hydrazones are present in various compounds, each demonstrating diverse biological activities. Extensive studies have revealed their potential as schistosomicidal agents. Thiosemicarbazones, in particular, have shown inhibitory effects on Schistosoma mansoni's cathepsin B1 enzyme (SmCB1), which plays a crucial role in hemoglobin degradation within the worm's gut and its nutrition processes. Consequently, SmCB1 has emerged as a promising target for novel schistosomiasis therapies. Moreover, chloroquinoline exhibits characteristics in its aromatic structure that hold promise for developing SmCB1 inhibitors, along with its interaction with hemoglobin's heme group, potentially synergizing against the parasite's gut. In this context, we report the synthesis of 22 hybrid analogs combining hydrazones and quinolines, evaluated against S. mansoni. Five of these hybrids demonstrated schistosomicidal activity in vitro, with GPQF-8Q10 being the most effective, causing worm mortality within 24 h at a concentration of 25 µM. GPQF-8Q8 proved to be the most promising in vivo, significantly reducing egg presence in feces (by 52.8%) and immature eggs in intestines (by 45.8%). These compounds exhibited low cytotoxicity in Vero cells and an in in vivo animal model (Caenorhabditis elegans), indicating a favorable selectivity index. This suggests their potential for the development of new schistosomiasis therapies. Further studies are needed to uncover specific target mechanisms, but these findings offer a promising starting point.

Evaluating Phytochemical Profiles, Cytotoxicity, Antiviral Activity, Antioxidant Potential, and Enzyme Inhibition of Vepris boiviniana Extracts.

In the present study, we performed comprehensive LC-MS chemical profiling and biological tests of Vepris boiviniana leaves and stem bark extracts of different polarities. In total, 60 bioactive compounds were tentatively identified in all extracts. The 80% ethanolic stem bark extract exhibited the highest activity in the ABTS assay, equal to 551.82 mg TE/g. The infusion extract of stem bark consistently demonstrated elevated antioxidant activity in all assays, with values ranging from 137.39 mg TE/g to 218.46 mg TE/g. Regarding the enzyme inhibitory assay, aqueous extracts from both bark and leaves exhibited substantial inhibition of AChE, with EC50 values of 2.41 mg GALAE/g and 2.25 mg GALAE/g, respectively. The 80% ethanolic leaf extract exhibited the lowest cytotoxicity in VERO cells (CC50: 613.27 µg/mL) and demonstrated selective cytotoxicity against cancer cells, particularly against H1HeLa cells, indicating potential therapeutic specificity. The 80% ethanolic bark extract exhibited elevated toxicity in VERO cells but had reduced anticancer selectivity. The n-hexane extracts, notably the leaves' n-hexane extract, displayed the highest toxicity towards non-cancerous cells with selectivity towards H1HeLa and RKO cells. In viral load assessment, all extracts reduced HHV-1 load by 0.14-0.54 log and HRV-14 viral load by 0.13-0.72 log, indicating limited antiviral activity. In conclusion, our research underscores the diverse bioactive properties of Vepris boiviniana extracts, exhibiting potent antioxidant, enzyme inhibitory, and cytotoxicity potential against cancer cells.

New insights into Bacillus cytotoxicus sources, screening, toxicity, and persistence in food production facilities

Bacillus cytotoxicus is a thermotolerant member of the Bacillus cereus group. It has been linked to rare, but at times fatal cases of diarrheal disease and might be missed at routine diagnostic screening temperatures commonly used for the B. cereus group. The pathogen is mostly found on dehydrated foods containing potato starch or insects. How it enters the food chain or whether it persists in food producing environments is largely unknown. Increased consumption of insects and convenience foods in Europe and the lack of information on the persistence of B. cytotoxicus in food environments and its virulence demand for further characterization. In this study, we aimed to obtain a better understanding of i) the food sources of B. cytotoxicus, ii) screening temperatures needed for its isolation from food matrices, iii) cytotoxicity of the organism, and iv) its ecological niche and potential epidemiological links. To this end, 112 food samples were collected, with a focus on foods exhibiting low water activity. The samples were screened for B. cytotoxicus at 42 °C and at 50 °C. Presumptive isolates were characterized by cytK-1 toxin gene PCR for differentiation of B. cytotoxicus from other B. cereus group members. Vero cell cytotoxicity assays were performed, and selected isolates were sequenced. Our results show that screening at 42 °C might be insufficient for detecting B. cytotoxicus in foods that harbor other less thermophilic Bacillus species. When screening at 50 °C, B. cytotoxicus was detected in 23% of the food samples (n = 26 isolates). The highest prevalence was detected in mashed potato products (82%) and potato flakes (67%). In contrast, a wide range of products not containing any potato ingredients did not yield B. cytotoxicus isolates. All B. cytotoxicus isolates exhibited either low or no detectable cytotoxicity. WGS analysis revealed that a highly toxic isolate is closely related to the French outbreak strain NVH 391-98. In addition, we could show that two isolates sampled 5 years apart from the same production facility only differed by seven SNPs, making it likely that B. cytotoxicus is able to persist in production facilities over a long time. Interestingly, the reoccurring strain possessed an additional plasmid and did not show cytotoxic potential when re-isolated after 5 years.

Apoptin NLS2 homodimerization strategy for improved antibacterial activity and bio-stability.

The emergence of antibiotic resistance prompts exploration of viable antimicrobial peptides (AMPs) designs. The present study explores the antimicrobial prospects of Apoptin nuclear localization sequence (NLS2)-derived peptide ANLP (PRPRTAKRRIRL). Further, we examined the utility of the NLS dimerization strategy for improvement in antimicrobial activity and sustained bio-stability of AMPs. Initially, the antimicrobial potential of ANLP using antimicrobial peptide databases was analyzed. Then, ANLP along with its two homodimer variants namely ANLP-K1 and ANLP-K2 were synthesized and evaluated for antimicrobial activity against Escherichia coli and Salmonella. Among three AMPs, ANLP-K2 showed efficient antibacterial activity with 12µM minimum inhibitory concentration (MIC). Slow degradation of ANLP-K1 (26.48%) and ANLP-K2 (13.21%) compared with linear ANLP (52.33%) at 480min in serum stability assay indicates improved bio-stability of dimeric peptides. The AMPs presented no cytotoxicity in Vero cells. Dye penetration assays confirmed the membrane interacting nature of AMPs. The zeta potential analysis reveals effective charge neutralization of both lipopolysaccharide (LPS) and bacterial cells by dimeric AMPs. The dimeric AMPs on scanning electron microscopy studies showed multiple pore formations on the bacterial surface. Collectively, proposed Lysine scaffold dimerization of Apoptin NLS2 strategy resulted in enhancing antibacterial activity, bio-stability, and could be effective in neutralizing the off-target effect of LPS. In conclusion, these results suggest that nuclear localization sequence with a modified dimeric approach could represent a rich source of template for designing future antimicrobial peptides.

The influence of exposure to various concentrations of five antimicrobial agents on intracellular cytotoxin B production in <i>Clostridioides difficile</i>

Background: Clostridioides difficile is an important cause of healthcare-associated diarrhea. Several anti- microbial agents are known to promote C. difficile infection (CDI). The impact of various concentrations of ampicillin (AMP), cefotaxime (CTX), clindamycin (CC), metronidazole (MTZ) and vancomycin (VAN) on intra-cellular cytotoxin B production was investigated in this study.
 Methodology: Six clinical strains of C. difficile were grown at minimum inhibitory concentration (MIC) and sub-MIC concentrations of these antibiotics. Inoculum standardization was performed by Miles and Misra method. Intracellular toxin B production was detected using Vero cell cytotoxicity assay in sonicated cultures on days 1, 2, 3, 4, 5 and 7 days of incubation.
 Results: There was a heterogeneous relationship between antibiotic exposure and the intra-cellular toxin production by the toxigenic strains. Clinical strains of C. difficile when exposed to MIC and sub-inhibitory concentrations of certain antibiotics produced high cytotoxin levels. All toxigenic isolates produced increased levels of cell-bound cytotoxin after exposure to antibiotics but there was no consistent pattern and the response to different doses varied considerably. Metronidazole was the most potent inducer of cell-bound cytotoxin followed by cefotaxime and clindamycin. Vancomycin induced the least amount of cytotoxin activity.
 Conclusion: The effects of sub-inhibitory concentration of antibiotic that predispose to C. difficile infection may partially suppress the normal gut flora, allowing colonization and growth of C. difficile, and may affect the level of toxin produced.

Screening antibiofilm activity of invasive plants growing at the Slope Merapi Mountain, Central Java, against Candida albicans

BackgroundCandida albicans causes high-mortality candidiasis. Antifungal drug resistance demands the development of virulence factor-targeting drugs, particularly antibiofilm. This study screened the effects of five invasive plants growing in Indonesia (Mimosa pudica, Lantana camara, Acacia mangium, Ageratina riparia, and Mikania micrantha) against C. albicans biofilms. Antifungal activity, antiphospholipase activity, biofilm morphology of C. albicans, and cytotoxic capacity were also evaluated.MethodsMaceration was used to extract the plants, and the most active extract inhibiting the biofilms was fractionated using liquid–liquid fractionation. Antibiofilm activity was determined by a colorimetric assay, MTT. Antifungal activity was tested using the broth microdilution method. A phospholipase assay was performed using the egg-yolk agar method. Influence on the C. albicans morphology was assessed using scanning electron microscopy (SEM). The cytotoxic effect was carried out against Vero and HeLa cell lines.ResultsM. pudica extracts showed the most potent antifungal efficacy with minimum inhibitory concentration (MIC) of 15.62 µg/mL and 7.81 µg/mL for aerial parts and roots, respectively. At high concentrations (500 µg/mL and 250 µg/mL), ethanol extract of M. pudica aerial parts strongly inhibited the phospholipase activity. Ethyl-acetate fraction of M. pudica aerial parts demonstrated the most potent antibiofilm activity against 24 h old biofilm of C. albicans with an inhibitory concentration (53.89%) of 62.5 µg/mL showed no cytotoxicity in both Vero and HeLa cells. This fraction affected the morphology of C. albicans and contained promising compounds for inhibiting the 24 h old biofilm of C. albicans.ConclusionsInvasive M. pudica plant inhibited the growth of planktonic C. albicans cells and its ethyl acetate fraction decreased the metabolic activity of C. albicans biofilms. This result demonstrates the potential of invasive M. pudica plant to reduce biofilm-associated candida infection.

In Vitro Antioxidant and Antitrypanosomal Activities of Extract and Fractions of Terminalia catappa.

Chagas disease is a severe infectious and parasitic disease caused by the protozoan Trypanosoma cruzi and considered a public health problem. Chemotherapeutics are still the main means of control and treatment of the disease, however with some limitations. As an alternative treatment, plants have been pointed out due to their proven pharmacological properties. Many studies carried out with Terminalia catappa have shown several biological activities, but its effect against T. cruzi is still unknown. The objective of this work is to evaluate the therapeutic potential of extracts and fractions obtained from T. catappa on the parasite T. cruzi, in addition to analyzing its antioxidant activity. T. catappa ethyl acetate fraction were produced and submitted the chemical characterization by Liquid Chromatography Coupled to Mass Spectrometry (LC-MS). From all T. catappa extracts and fractions evaluated, the ethyl acetate and the aqueous fraction displayed the best antioxidant activity by the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging method (IC50 of 7.77 ± 1.61 and 5.26 ± 1.26 µg/mL respectively), and by ferric ion reducing (FRAP) method (687.61 ± 0.26 and 1009.32 ± 0.13 µM of Trolox equivalent/mg extract, respectively). The ethyl acetate fraction showed remarkable T. cruzi inhibitory activity with IC50 of 8.86 ± 1.13, 24.91 ± 1.15 and 85.01 ± 1.21 µg/mL against epimastigotes, trypomastigotes and intracellular amastigotes, respectively, and showed no cytotoxicity for Vero cells (CC50 > 1000 µg/mL). The treatment of epimastigotes with the ethyl acetate fraction led to drastic ultrastructural changes such as the loss of cytoplasm organelles, cell disorganization, nucleus damage and the loss of integrity of the parasite. This effect could be due to secondary compounds present in this extract, such as luteolin, kaempferol, quercetin, ellagic acid and derivatives. The ethyl acetate fraction obtained from T. catappa leaves can be an effective alternative in the treatment and control of Chagas disease, and material for further investigations.

Lycopene from Red Guava (Psidium guajava L.): From Hepatoprotective Effect to Its Use as Promising Self-Emulsifying Drug Delivery System for Anti-Inflammatory and Antioxidant Applications.

Lycopene is a carotenoid with potential use in the treatment of chronic illnesses. Here, different formulations of lycopene were studied: lycopene-rich extract from red guava (LEG), purified lycopene from red guava (LPG) and a self-emulsifying drug delivery system loaded with LPG (nanoLPG). The effects of administering orally various doses of LEG to hypercholesterolemic hamsters were evaluated regarding the liver function of the animals. The cytotoxicity of LPG in Vero cells was analyzed by a crystal violet assay and by fluorescence microscopy. In addition, nanoLPG was employed in stability tests. LPG and nanoLPG were tested for their cytotoxic effect on human keratinocytes and antioxidant capacity on cells in an endothelial dysfunction model in an isolated rat aorta. Finally, the effect of different nanoLPG concentrations on the expression of immune-related genes (IL-10, TNF-α, COX-2 and IFN-γ) from peripheral blood mononuclear cells (PBMC) using real-time PCR was also analyzed. Results suggest that LEG, despite not being able to improve blood markers indicative of liver function in hypercholesterolemic hamsters, reduced hepatic degenerative changes. Additionally, LPG did not show cytotoxicity in Vero cells. In relation to nanoLPG, the effects produced by heat stress evaluated by Dynamics Light Scattering (DLS) and visually were loss of color, texture change and phase separation after 15 days without interfering with the droplet size, so the formulation proved to be efficient in stabilizing the encapsulated lycopene. Although LPG and nanoLPG showed moderate toxicity to keratinocytes, which may be related to cell lineage characteristics, both revealed potent antioxidant activity. LPG and nanoLPG showed vasoprotective effects in aortic preparations. The gene expression assay indicates that, although no significant differences were observed in the expression of IL-10 and TNF-α, the PBMCs treated with nanoLPG showed a reduction in transcriptional levels of IFN-γ and an increased expression of COX-2. Thus, the work adds evidence to the safety of the use of lycopene by humans and shows that tested formulations, mainly nanoLPG due to its stability, stand out as promising and biosafe products for the treatment of diseases that have oxidative stress and inflammation in their etiopathology.

Preclinical evaluation of antimalarial activity of CPF-1 formulation as an alternative choice for the treatment of malaria

BackgroundKheaw Hom remedy is a traditional Thai medicine used to treat fever. Some plants used in the Kheaw Hom remedy show promising in vitro antimalarial activity. This study prepared novel formulations of plants from the Kheaw Hom remedy and evaluated their antimalarial and toxicological activities.MethodsSeven new formulations were prepared by combining at least three herbs of six selected plants from the Kheaw Hom remedy, namely Mammea siamensis Kosterm., Mesua ferrea L., Dracaena loureiroi Gagnep., Pogostemon cablin (Blanco) Benth., Kaempferia galanga L, and Eupatorium stoechadosmum Hance. In vitro antimalarial activities of each formulation’s aqueous and ethanolic extracts were evaluated using the parasite lactate dehydrogenase (pLDH) assay. Cytotoxicity in Vero and HepG2 cells was assessed using the MTT assay. An extract with good antimalarial potency and selectivity index (SI) was selected for in vivo antimalarial activity using Peter’s 4-day suppressive test and acute oral toxicity test in mice. In addition, bioactive compounds were identified using Gas chromatography-mass spectrometry (GC-MS) analysis.ResultsAmong the seven new formulations, ethanolic extracts of CPF-1 (Formulation 1) showed the highest activity with an IC50 value of 1.32 ± 0.66 µg/ml, followed by ethanolic extracts of Formulation 4 and Formulation 6 with an IC50 value of 1.52 ± 0.28 µg/ml and 2.48 ± 0.34 µg/ml, respectively. The highest SI values were obtained for the ethanolic extract of CPF-1 that was selected to confirm its in vivo antimalarial activity and toxicity. The results demonstrated a significant dose-dependent reduction in parasitemia. Maximum suppressive effect of the extract (72.01%) was observed at the highest dose administered (600 mg/kg). No significant toxicity was observed after the administration of 2000 mg/kg. Using GC-MS analysis, the most abundant compound in the ethanolic extract of CPF-1 was ethyl p-methoxycinnamate (14.32%), followed by 2-propenoic acid, 3-phenyl-, ethyl ester, (E)- (2.50%), and pentadecane (1.85%).ConclusionThe ethanolic extract of CPF-1 showed promising in vitro and in vivo antimalarial efficacy, with no toxic effects at a dose of 2000 mg/kg, suggesting that the ethanolic extract of CPF-1 may serves as a new herbal formulation for the treatment of malaria. Additional research is required for safety and clinical pharmacology studies.

Synthetic Curcumin Analogues Present Antiflavivirus Activity In Vitro with Potential Multiflavivirus Activity from a Thiazolylhydrazone Moiety

Arboviral diseases caused by flaviviruses, such as dengue, are a continuing threat and major concern worldwide, with over three billion people estimated to be living with the risk of dengue virus (DENV) infections. There are thus far no antiviral drugs available for treatment, and limited or no vaccines are available. Curcumin and seven synthetic analogues were evaluated for their antiviral activity against dengue virus serotype 2, yellow fever virus and Zika virus, as well as for their cytotoxicity in Vero cells, both by employing MTT assays. Compounds 6 and 7, which present a thiazolylhydrazone moiety, showed moderate activity against all three flaviviruses, with selectivity index (SI) values up to 4.45. In addition, the envelope protein (E) was predicted as the potential target inhibited by both compounds, supported by molecular docking and dynamics simulation analysis. We hope that this data can contribute to the development of new curcumin antiviral analogues in the near future and can help in the search for new promising compounds as potential therapeutic agents to treat flaviviruses infections.

Non-Toxigenic Clostridioides difficile Strain E4 (NTCD-E4) Prevents Establishment of Primary C. difficile Infection by Epidemic PCR Ribotype 027 in an In Vitro Human Gut Model.

Clostridioides difficile infection (CDI) remains a significant healthcare burden. Non-toxigenic C. difficile (NTCD) strains have shown a benefit in preventing porcine enteritis and in human recurrent CDI. In this study, we evaluated the efficacy of metronidazole-resistant NTCD-E4 in preventing CDI facilitated by a range of antimicrobials in an in vitro human gut model. NTCD-E4 spores (at a dose of 107) were instilled 7 days before a clinical ribotype (RT) 027 (at the same dose) strain (210). In separate experiments, four different antimicrobials were used to perturb gut microbiotas; bacterial populations and cytotoxin production were determined using viable counting and Vero cell cytotoxicity, respectively. RT027 and NTCD-E4 proliferated in the in vitro model when inoculated singly, with RT027 demonstrating high-level cytotoxin (3-5-log10-relative units) production. In experiments where the gut model was pre-inoculated with NTCD-E4, RT027 was remained quiescent and failed to produce cytotoxins. NTCD-E4 showed mutations in hsmA and a gene homologous to CD196-1331, previously linked to medium-dependent metronidazole resistance, but lacked other metronidazole resistance determinants. This study showed that RT027 was unable to elicit simulated infection in the presence of NTCD-E4 following stimulation by four different antimicrobials. These data complement animal and clinical studies in suggesting NTCD offer prophylactic potential in the management of human CDI.

Mulberroside F from In Vitro Culture of Mulberry and the Potential Use of the Root Extracts in Cosmeceutical Applications

Mulberry (Morus spp.) is primarily used in sericulture, and its uses also extend to the food, pharmaceutical, and cosmetic industries. Mulberry extracts are rich in many bioactive compounds that exhibit a wide range of biological properties. Mulberroside F (Moracin M-6, 3'-di-O-β-D-glucopyranoside), one of the bioactive compounds found in mulberry, has previously been reported as a whitening agent by inhibiting melanin synthesis and exhibiting antioxidant effects. However, there is still limited information on the presence of this compound in plants cultured in vitro. In this study, the mulberroside F content, biochemical, and cytotoxic properties of the extracts from mulberry cultured in vitro were determined. The results revealed that both root and callus were found to be a potential source of mulberroside F. Furthermore, the mulberroside F content was positively correlated with the inhibitory effects on tyrosinase activity. Cell viability assay also revealed that crude extract of the mulberry root has no cytotoxicity in both human keratinocyte cell line (HaCaT) and Vero cells. Taken together, mulberry tissue culture represents a possible alternative and continuous production of mulberroside F, which could be further utilized in cosmeceutical applications.

The potent antioxidant effect of Neutrase-assisted hydrolysate from heat-resistant Pyropia yezoensis by molecular weight change

Increased oceanic temperature due to climatic change is gradually decreasing the production rates of Pyropia yezoensis in South Korea, which accounts for 50 % of the global supply. This study demonstrates the antioxidant property of a newly developed edible marine red algae, heat-resistant P. yezoensis (HrP). The water extract of HrP displayed a notably higher reactive oxygen species (ROS) scavenging effect on DPPH, alkyl, hydroxyl radicals, and H2O2 than the commercial P. yezoensis water extract. To increase the radical scavenging ability of HrP, it was enzymatically hydrolyzed using seven enzymes (Alcalase, Celluclast, Flavourzyme, Neutrase, Protamex, Ultraflo, and Viscozyme). These enzymes-assisted HrP hydrolysates (EHrPs) were evaluated for their free radical scavenging activity. Among them, Neutrase-assisted HrP hydrolysate (NHrP), containing the highest protein content, showed a significant protective effect against H2O2-induced cytotoxicity in Vero cells. The 10 kDa molecular weight cut-off fractionation of NHrP revealed that the filtered fraction (10B; ≤10 kDa) was superior to the unfiltered fraction (10A; >10 kDa) in antioxidant activity. The characterization of the three fractions indicated that the high radical scavenging function mainly depends on the level of proteins. Moreover, 10B treatment ameliorated H2O2-induced oxidative stress in Vero cells and zebrafish embryos (increased survival percentage, and decreased cell death and ROS formation). These data suggest that HrP might be a potential antioxidant for nutraceutical industries.