Acute Toxicity Assays with the Artemia salina Model: Assessment of Variables.

The use of the brine shrimp Artemia salina (Leach) in acute toxicity assays has great potential due to its simplicity, low cost and reproducibility. In the current study, some of the variables that can influence the reliability of the assay in terms of test organism survival, were evaluated as part of its implementation in our laboratory. The quality and type of water used, the buffer components and other parameters (salinity, pH and dissolved oxygen level), were all evaluated for optimisation purposes. DMSO (dimethyl sulphoxide) was used as the test substance in the toxicity assay, to evaluate the concentration limits as a solvent in sample preparation. Regarding the buffer salinity, pH and dissolved oxygen level, we found that a 25% to 30% deviation from the standard values did not affect the survival of the nauplii (the first-instar larval stage) under assay conditions. In summary, we corroborate the potential use of this model for the prediction of the toxic potential of substances, to inform future testing strategies.

Purification of andibenin and a chromanone analogue from rhizospheric Aspergillus flavus and soil-borne Penicillium notatum exhibiting cytotoxic and antibacterial properties.

The discovery of bioactive compounds from fungal natural sources holds immense potential for the development of novel therapeutics. The present study investigates the extracts of soil-borne Penicillium notatum and rhizosphere-inhabiting Aspergillus flavus for their antibacterial, antifungal, and cytotoxic potential. Additionally, two compounds were purified using chromatographic and spectroscopic techniques. The results demonstrated that the ethyl acetate fraction of A. flavus exhibited prominent cytotoxic activity against Artemia salina, whereas the ethyl acetate fraction of P. notatum displayed promising antibacterial potential. At dose concentrations of 10, 100, and 1000 µg mL-1, the ethyl acetate fraction of A. flavus showed mortality percentages of 7.6%, 66.4%, and 90%, respectively. The ethyl acetate fraction of P. notatum extract exhibited significant antibacterial activity, forming inhibition zones measuring 41, 38, 34, 34, and 30 mm against B. subtilis, S. flexneri, E. coli, K. pneumoniae, and S. aureus, respectively, at 1000 µg mL-1. At this concentration, inhibition zones of 28, 27, and 15 mm were recorded for P. vulgaris, S. typhi, and X. oryzae. Using bioassay-guided approach, one compound each was purified from the fungal extracts. The initial purification involved mass spectroscopic analysis, followed by structural elucidation using 500 MHz nuclear magnetic resonance (NMR) spectroscopy. Compound 1, derived from A. flavus, was identified as ethyl 2-hydroxy-5,6-dimethyl-4-oxocyclohex-2-ene-1-carboxylate, with a mass of 212, whereas compound 2, isolated from P. notatum, was identified as 3-amino-2-(cyclopenta-2,4-dien-1-ylamino)-8-methoxy-4H-chromen-4-one, with an exact mass of 270. Based on bioassay results, compound 1 was subjected to brine shrimp lethality assay and compound 2 was tested for its antibacterial potential. Compound 1 exhibited 30% lethality against brine shrimp larvae at a concentration of 100 µg mL-1, whereas at 1000 µg mL-1 the mortality increased to 70%. Compound 2 displayed notable antibacterial potential, forming inhibition zones of 30, 24, 19, and 12 mm against S. aureus, E. coli, B. subtilis, and S. flexneri, respectively. In comparison, the standard antibiotic tetracycline produced inhibition zones of 18, 18, 15, and 10 mm against the respective bacterial strains at the same concentration.

Assessment of the Artemia salina toxicity assay as a substitute of the mouse lethality assay in the determination of venom-induced toxicity and preclinical efficacy of antivenom.

Mice are routinely used in snake venom research but are costly and subject to pain and suffering. The crustacean Artemia salina could be an alternative to mice, but data to support its adoption in snake venom research is limited. The aim of the present study was to evaluate the suitability of A. salina as a surrogate of mice in assessing the toxicity of venoms and the preclinical efficacy of antivenoms. The toxicity of venoms from 22 snakes of medical importance in sub-Saharan Africa was evaluated in mice (intraperitoneally; i.p. and intravenously; i.v.) and in A. salina. Subsequently, the capacity of a commercial antivenom to neutralize the toxicity of these venoms in mice and A. salina was investigated. There was a positive correlation between the i.v. median lethal doses (LD50s) and the i.p. LD50s in mice (r = 0.804; p < 0.0001), a moderate correlation between the i.v. LD50s in mice and the median lethal concentrations (LC50s) in A. salina (r = 0.606; p = 0.003), and a moderate correlation between the i.p. LD50s in mice and the LC50s in A. salina (r = 0.426; p = 0.048). Moreover, there was a strong correlation between the i.p. median effective doses (ED50s) and the i.v. ED50s in mice (r = 0.941, p < 0.0001), between the i.p. ED50s in mice and the ED50s in A. salina (r = 0.818, p < 0.0001), and between the i.v. ED50s in mice and the ED50s in A. salina (r = 0.972, p < 0.0001). These findings present A. salina as a promising candidate for reducing reliance on mice in snake venom research. Future investigations should build upon these findings, addressing potential limitations and expanding the scope of A. salina in venom research and antivenom development.

Efficacy and safety of artificial tears containing Artemia salina extract with dinucleotides for dry eye.

CLINICAL RELEVANCE: This clinical trial was conducted as part of the marketing procedures for a medical device comprising artificial tears containing Artemia salina extract with dinucleotides. These molecules previously demonstrated secretagogue properties by enhancing the production of aqueous, mucinous, and lipidic components of the tears. BACKGROUND: After confirming the efficacy of artificial tears containing Artemia salina extract in an animal model, this study proceeded to evaluate their efficacy and safety on dry eye participants. METHODS: A randomised controlled clinical trial was performed on 36 dry eye participants (41.6 ± 20.6 years). Half of the participants were treated with saline solution as a placebo for four weeks, while the other half were treated with artificial tears containing Artemia salina, randomly assigned. After a wash-out period of two weeks, the treatments were crossed for another four weeks. Participants were assessed at baseline and after one week, two weeks, and four weeks. Efficacy variables were: eye dryness frequency (primary), eye comfort, visual satisfaction, tear secretion, tear break-up time, corneal staining, conjunctival staining, and conjunctival hyperaemia. Safety variables were: high- and low-contrast visual acuity, intraocular pressure, and eye fundus images analysis. RESULTS: Compared with the baseline, the saline solution showed no significant changes in any of the studied variables after four weeks of treatment (p ≥ 0.05). However, the topical instillation of the artificial tears with Artemia salina for four weeks significantly improved eye dryness frequency (p = 0.014) and corneal staining (p = 0.010). No systemic or ocular adverse events were reported during the clinical trial. CONCLUSION: The topical instillation of artificial tears containing Artemia salina in mild to moderate dry eye participants for four weeks slightly improved their symptoms related to eye dryness frequency and reduced corneal damage, with no undesirable side effects observed.

Serbian Artemisia species - chemical composition, acute toxicity and larvicidal activity of the essential oils.

The chemical composition and biological activities of the essential oils obtained from Serbian Artemisia species (A. alba, A. absinthium, A. annua, A. vulgaris, and A. scoparia) were analysed. The essential oil was obtained by merging several samples (same plant species, different localities) and the chemical composition was compared with pre-merging results. In the merged A. scoparia sample four components were not found in any pre-merging sample and one of those is present in the highest percentage (capillin 35.7%). The least toxic essential oil in Artemia salina test was A. annua, followed by A. alba (both showing medium toxicity), while A. absinthium, A. vulgaris, and A. scoparia showed strong toxicity. All tested samples showed activity against Drosophila melanogaster larvae in descending order ΣAS > ΣAN > ΣAV > ΣAB > ΣAA. The essential oil of A. scoparia has exceptional larvicidal activity (in concentrations of 2% and 1% causes complete mortality).

Eco-friendly green synthesis of AgNPs from Elaeocarpus serratus fruit extract: potential to antibacterial, antioxidant, cytotoxic effects of colon cancerous cells (HT-29) and its toxicity assessments of marine microcrustacean Artemia nauplii.

BACKGROUND: The present work demonstrated the green synthesis and characterization of silver nanoparticles (AgNPs) utilizing Elaeocarpus serratus fruit extract. The study examined the effectiveness of phytocompounds in fruit extract in reducing Ag+ to Ag° ions. METHODS: The water-soluble biobased substance production from silver ions to AgNPs in 45 min at room temperature. Surface plasmon resonance (SPR) peak was seen in the UV-visible absorption spectrum of the biologically altered response mixture. Examination with X-ray diffraction (XRD) showed that AgNPs are strong and have a face-centered cubic shape. Scanning electron microscope (SEM) investigation proved the production of AgNPs in a cuboidal shape. RESULTS: The AgNPs demonstrated remarkable antibacterial activity and a potent capacity to neutralize DPPH (2,2-Diphenyl-1-picrylhydrazyl) radicals. The highest growth inhibition was found for E. serratus against S. dysenteriae (18.5 ± 1.0 mm) and S. aureus (18 ± 1.2 mm). These nanoparticles exhibited robust antiradical efficacy even at low concentrations. The AgNPs additionally exhibited cytotoxic effects on (HT-29) human colon adenocarcinoma cancer cells. The MTT assay (3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) indicated an inhibitory concentration (IC50) value of 49.1 ± 2.33 µg/mL for AgNPs, contrasting with the untreated cells of the negative control. The biotoxicity assessment using A. salina displayed mortality rates ranging from 8 to 69.33%, attributable to the E. serratus synthesized AgNPs. CONCLUSIONS: In our results concluded that simply first-hand information on that E. serattus fruit extract synthesized AgNPs were efficiently synthesized without the addition of any hazardous substances, and that they may be a strong antibacterial, antioxidant, and potential cytotoxic effects for the treatment of colon carcinoma cell lines.

Chemical Composition of Piper nigrum L. Cultivar Guajarina Essential Oils and Their Biological Activity.

The essential oils and aroma derived from the leaves (L), stems (St), and spikes (s) of Piper nigrum L. cv. Guajarina were extracted; the essential oils were extracted using hydrodistillation (HD), and steam distillation (SD), and the aroma was obtained by simultaneous distillation and extraction (SDE). Chemical constituents were identified and quantified using GC/MS and GC-FID. Preliminary biological activity was assessed by determining the toxicity against Artemia salina Leach larvae, calculating mortality rates, and determining lethal concentration values (LC50). The predominant compounds in essential oil samples included α-pinene (0-5.6%), ß-pinene (0-22.7%), limonene (0-19.3%), 35 linalool (0-5.3%), δ-elemene (0-10.1%), ß-caryophyllene (0.5-21.9%), γ-elemene (7.5-33.9%), and curzerene (6.9-31.7%). Multivariate analysis, employing principal component analysis (PCA) and hierarchical cluster analysis (HCA), revealed three groups among the identified classes and two groups among individual compounds. The highest antioxidant activity was found for essential oils derived from the leaves (167.9 41 mg TE mL-1). Larvicidal potential against A. salina was observed in essential oils obtained from the leaves (LC50 6.40 µg mL-1) and spikes (LC50 6.44 µg mL-1). The in silico studies demonstrated that the main compounds can interact with acetylcholinesterase, thus showing the potential molecular interaction responsible for the toxicity of the essential oil in A. salina.

Naturally weathered polypropylene microplastic from environment and its toxic behaviour in Artemia salina.

The increasing use of polypropylene (PP) in consumer products leads to the microplastic (PP MPs) contamination of the aquatic ecosystems. Comprehensive toxicological studies of weathered/aged and new PP MPs with Artemia salina are a need of the hour. Our study explores the toxicological differences between naturally weathered (aged) and prepared new PP MPs on Artemia salina. Both the weathered and new PP MPs were prepared using controlled grinding and sieving at ≤ 125 µm. Artemia salina was treated with different concentrations (0.25, 0.5, and 1 mg/mL) of PP MP particles for up to 48 h. The uptake of weathered PP MP particles by Artemia salina was higher than the new PP MPs. The accumulation of PP MP particles was found in the intestine. There was increased oxidative stress recorded in the animal treated with the weathered PP MPs than the new PP MPs. Artemia salina treated with weathered PP MPs showed higher ROS generation and increased, activity of oxidative enzymes like LPO, SOD, and CAT. Collectively, our findings underscore the detrimental effects of weathered and prepared new PP MPs on Artemia salina, which is an ecologically significant species of zooplankton. There is an urgent need and effective measures required to address plastic disposal strategies in an environmentally safe manner.

Degradation of multiple PAHs and co-contaminants by microbial consortia and their toxicity assessment.

The anthropogenic activities toward meeting the energy requirements have resulted in an alarming rise in environmental pollution levels. Among pollutants, polycyclic aromatic hydrocarbons (PAHs) are the most predominant due to their persistent and toxic nature. Amidst the several pollutants depuration methods, bioremediation utilizing biodegradation is the most viable alternative. This study investigated the biodegradation efficacy using developed microbial consortium PBR-21 for 2-4 ringed PAHs named naphthalene (NAP), anthracene (ANT), fluorene (FLU), and pyrene (PYR). The removal efficiency was observed up to 100 ± 0.0%, 70.26 ± 4.2%, 64.23 ± 2.3%, and 61.50 ± 2.6%, respectively, for initial concentrations of 400 mg L-1 for NAP, ANT, FLU, and PYR respectively. Degradation followed first-order kinetics with rate constants of 0.39 d-1, 0.10 d-1, 0.08 d-1, and 0.07 d-1 and half-life [Formula: see text] of 1.8 h, 7.2 h, 8.5 h, and 10 h, respectively. The microbial consortia were found to be efficient towards the co-contaminants with 1 mM concentration. Toxicity examination indicated that microbial-treated PAHs resulted in lesser toxicity in aquatic crustaceans (Artemia salina) than untreated PAHs. Also, the study suggests that indigenous microbial consortia PBR-21 has the potential to be used in the bioremediation of PAH-contaminated environment.

Prepared microplastics interaction with Artemia salina under low pH conditions representing ocean acidification; a simulated environmental exposure.

Ocean acidification and microplastic pollution are two of the major ecological concerns. The distribution of large quantities of plastic debris and microplastics all across the oceans emphasises the need to determine the influence of microplastics in ocean acidification and to evaluate its concomitant toxicological effects on aquatic life forms. Studies on the combined impact of both the stressors are very limited, but much needed in the current scenario. Where most of the present-day research use purchased microplastics of defined size and morphology (microspheres, fibres, rods, etc.), the present study employs prepared "true to life microplastics" that resemble the environmental microplastic pollutants in morphology and size heterogeneity. The present study focusses on evaluating the fate and impact of oceanic microplastics on the physiology and development of Artemia salina (Brine shrimp), one among the most ecologically significant zooplankton species. Natural sea water was acidified by controlled perturbation of carbon dioxide using a valve system. The hatching rate of A. salina cysts receded significantly (p < 0.05) upon singular exposures to microplastics and low pH (7.80), whereas combined effect was insignificant. The reactive oxygen species (ROS) elevated as a result of individual exposures to microplastics and low pH. However, only in 0.5 mg mL-1 PE treatments at pH 7.80, an additive impact was reported for ROS activity (p < 0.05). The SOD activities increased significantly but it can be attributed as the individual responses towards exposure to both the stressors. A significant additive impact was not observed for SOD activity (p > 0.05). But during the development, significant morphological anomalies were observed. Changes in the appendages of nauplii and juveniles as a result of combined exposure to microplastics and low pH treatments are significant findings. Our observations suggest that coupled exposure to microplastics and low pH could induce significant oxidative stress in the marine zooplanktons and also adversely affect their normal development. Findings from the current study emphasise the need for further research to understand the coupled toxicological impacts of ocean acidification and predominant pollutants such as microplastics to other marine animals as well.

Sublethal Effects of Polystyrene Nanoplastics on the Embryonic Development of Artemia salina (Linnaeus, 1758).

Currents, wave motion, solar radiation, and abrasion are mechanisms responsible for the degradation of large plastic artifacts and contribute to the dispersion of micro and nanoplastics into aquatic ecosystems, which are, currently, the most dangerous threats due to their invisibility and persistence. The present work evaluated the possible lethal and sublethal effects of amino-modified polystyrene nanoplastics (nPS-NH2) with diameters of 50 nm and 100 nm on Artemia salina (A. salina), an organism at the base of the trophic chain of the aquatic system, using a widely used model for the analysis of embryotoxicity from environmental pollutants. For this purpose, after evaluating the biodistribution of nanoplastics in the body of the tested animals, several endpoints such as anomalies, apoptosis, and ROS production were assessed. In addition, particular attention was dedicated to evaluating the correlation between toxicity and the particle size tested. The results reported that, despite the absence of a lethal impact, several sublethal effects involving gut and body size malformations, as well as the enhancement of apoptosis and oxidative stress in relation to an increase in tested concentration and a decrease in nanoparticle size.

In vitro biofilm inhibition efficacy of Aerva lanata flower extract against Gram negative and Gram-positive biofilm forming bacteria and toxicity analysis using Artemia salina.

A biofilm consists of Gram positive and Gram-negative bacteria enclosed in a matrix. Industrial biofouling is caused by biofilms, which can exhibit antimicrobial resistance during infections. Many biofilm studies find that nearly all biofilm communities consist of Gram positive and Gram-negative bacteria. It is therefore necessary to better understand the conserved themes in biofilm formation to develop therapeutics based on biofilm formation. Plant extracts can effectively combat pathogenic bacterial biofilms. This study evaluated the antibacterial and antibiofilm activity of Aerva lanata flower extract against Staphylococcus aureus and Pseudomonas aeruginosa. Methanol extract of dried A. lanata flower was tested against S. aureus and P. aeruginosa to determine the antibacterial activity (10, 25, 50, 75, 100 µg/mL) resulted in a maximum of 0.5-1 log reduction and 2 log reduction in comparison to the control or untreated bacterial cells respectively. A. lanata showed maximum biofilm inhibition up to 1.5-fold and 1-fold against P. aeruginosa and S. aureus. Light microscopic analysis of biofilm treated with A. lanata extract showed efficient distortion of the biofilm matrix. Further, the in vivo analysis of A. lanata in the Artemia salina brine shrimp model showed >50% survival and thus proving the efficacy of A. lanata extract in rescuing the brine shrimps against P. aeruginosa and S. aureus infection.

Chemical Composition, Preliminary Toxicity, and Antioxidant Potential of Piper marginatum Sensu Lato Essential Oils and Molecular Modeling Study.

The essential oils (OEs) of the leaves, stems, and spikes of P. marginatum were obtained by hydrodistillation, steam distillation, and simultaneous extraction. The chemical constituents were identified and quantified by GC/MS and GC-FID. The preliminary biological activity was determined by assessing the toxicity of the samples to Artemia salina Leach larvae and calculating the mortality rate and lethal concentration (LC50). The antioxidant activity of the EOs was determined by the DPPH radical scavenging method. Molecular modeling was performed using molecular docking and molecular dynamics, with acetylcholinesterase being the molecular target. The OES yields ranged from 1.49% to 1.83%. The EOs and aromatic constituents of P. marginatum are characterized by the high contents of (E)-isoosmorhizole (19.4-32.9%), 2-methoxy-4,5-methylenedioxypropiophenone (9.0-19.9%), isoosmorhizole (1.6-24.5%), and 2-methoxy-4,5-methylenedioxypropiophenone isomer (1.6-14.3%). The antioxidant potential was significant in the OE of the leaves and stems of P. marginatum extracted by SD in November (84.9 ± 4.0 mg TE·mL-1) and the OEs of the leaves extracted by HD in March (126.8 ± 12.3 mg TE·mL-1). Regarding the preliminary toxicity, the OEs of Pm-SD-L-St-Nov and Pm-HD-L-St-Nov had mortality higher than 80% in concentrations of 25 µg·mL-1. This in silico study on essential oils elucidated the potential mechanism of interaction of the main compounds, which may serve as a basis for advances in this line of research.

Investigation of Metal Toxicity on Microalgae Phaeodactylum tricornutum, Hipersaline Zooplankter Artemia salina, and Jellyfish Aurelia aurita.

The escalating global anthropogenic activities associated with industrial development have led to the increased introduction of heavy metals (HMs) into marine environments through effluents. This study aimed to assess the toxicity of three HMs (Cr, Cu, and Cd) on organisms spanning different trophic levels: Phaeodactylum tricornutum (a primary producer), Artemia salina (a primary consumer), and Aurelia aurita (a secondary consumer). The EC50 values obtained revealed varying relative toxicities for the tested organisms. Phaeodactylum tricornutum exhibited the highest sensitivity to Cu, followed by Cd and Cr, while Artemia salina displayed the highest sensitivity to Cr, followed by Cu and Cd. A. aurita, on the other hand, demonstrated the highest sensitivity to Cu, followed by Cr and Cd. This experimental investigation further supported previous studies that have suggested A. aurita as a suitable model organism for ecotoxicity testing. Our experiments encompassed sublethal endpoints, such as pulsation frequency, acute effects, and mortality, highlighting different levels of sensitivity among the organisms.

Chitosan (CS)/Hydroxyapatite (HA)/Tricalcium Phosphate (ß-TCP)-Based Composites as a Potential Material for Pulp Tissue Regeneration.

This research focused on developing new materials for endodontic treatments to restore tissues affected by infectious or inflammatory processes. Three materials were studied, namely tricalcium phosphate ß-hydroxyapatite (ß-TCP), commercial and natural hydroxyapatite (HA), and chitosan (CS), in different proportions. The chemical characterization using infrared spectroscopy (FTIR) and X-ray diffraction (XRD) analysis confirmed the composition of the composite. Scanning electron microscopy (SEM) demonstrated that the design and origin of the HA, whether natural or commercial, did not affect the morphology of the composites. In vitro studies using Artemia salina (A. salina) indicated that all three experimental materials were biocompatible after 24 h, with no significant differences in mortality rate observed among the groups. The subdermal implantation of the materials in block form exhibited biocompatibility and biodegradability after 30 and 60 days, with the larger particles undergoing fragmentation and connective tissue formation consisting of collagen type III fibers, blood vessels, and inflammatory cells. The implanted material continued to undergo resorption during this process. The results obtained in this research contribute to developing endodontic technologies for tissue recovery and regeneration.

Progress and promise of alternative animal and non-animal methods in biomedical research.

Cell culture and invertebrate animal models reflect a significant evolution in scientific research by providing reliable evidence on the physiopathology of diseases, screening for new drugs, and toxicological tests while reducing the need for mammals. In this review, we discuss the progress and promise of alternative animal and non-animal methods in biomedical research, with a special focus on drug toxicity.

Understanding the Artemia Salina (Brine Shrimp) Test: Pharmacological Significance and Global Impact.

BACKGROUND: The microplate benchtop brine shrimp test (BST) has been widely used for screening and bio-guided isolation of many active compounds, including natural products. Although the interpretation given to the results appears dissimilar, our findings suggest a correlation between positive results with a specific mechanism of action. OBJECTIVE: This study aimed to evaluate drugs belonging to fifteen pharmacological categories having diverse mechanisms of action and carry out a bibliometric analysis of over 700 citations related to microwell BST. METHODS: Test compounds were evaluated in a serial dilution on the microwell BST using healthy nauplii of Artemia salina and after 24 hrs of exposition, the number of alive and dead nauplii was determined, and the LC50 was estimated. A metric study regarding the citations of the BST miniaturized method, sorted by type of documents cited, contributing country, and interpretation of results was conducted on 706 selected citations found in Google Scholar. RESULTS: Out of 206 drugs tested belonging to fifteen pharmacological categories, twenty-six showed LC50 values <100 µM, most of them belonging to the category of antineoplastic drugs; compounds with different therapeutical uses were found to be cytotoxic as well. A bibliometric analysis showed 706 documents citing the miniaturized BST; 78% of them belonged to academic laboratories from developing countries located on all continents, 63% interpreted their results as cytotoxic activity and 35% indicated general toxicity assessment. CONCLUSION: BST is a simple, affordable, benchtop assay, capable of detecting cytotoxic drugs with specific mechanisms of action, such as protein synthesis inhibition, antimitotic, DNA binding, topoisomerase I inhibitors, and caspases cascade interfering drugs. The microwell BST is a technique that is used worldwide for the bio-guided isolation of cytotoxic compounds from different sources.

Chemical profile, antioxidant potential and toxicity of Smilax brasiliensis Sprengel (Smilacaceae) stems.

Smilax brasiliensis Sprengel is a monocotyledon of the Smilacaceae family, native to the Brazilian Cerrado, popularly known as "salsaparrilha" or "japecanga". In this study, the ethanol extract (EE) and the hexane (HEXF), dichloromethane (DCMF), ethyl acetate (ACF), and hydroethanol (HEF) fractions of the stems were obtained. The chemical composition was determined, the contents of phenolic compounds and flavonoids were quantified, and the antioxidant potential and the cytotoxic effect on Artemia salina were evaluated. Fatty acid esters, hydrocarbons, and phytosterols were identified in the HEXF analyzed by gas chromatography - mass spectrometry (GC-MS). The EE and DCMF, ACF, and HEF were analyzed by liquid chromatography coupled to a diode array detector and mass spectrometer (LC-DAD-MS), and the identified constituents included glycosylated (rutin, 3-O-ß-galactopyranosyl quercetin, 3-O-ß-glucopyranosyl quercetin, O-deoxyhexosyl-hexosyl quercetin, O-deoxyhexosyl-hexosyl kaempferol, O-deoxyhexosyl-hexosyl O-methyl quercetin, and others), and non-glycosylated (quercetin) flavonoids, phenylpropanoids (3-O-E-caffeoyl quinic acid, 5-O-E-caffeoyl quinic acid, O-caffeoyl shikimic acid, and others), neolignan, steroidal saponin (dioscin), and N-feruloyltyramine. The EE, DCMF, and ACF showed high total contents of phenolic compounds (112.99, 175.71, and 524.02 µg of GAE/mg, respectively), and in the ACF and DCMF a great content of flavonoids was also quantified (50.08 and 31.49 µg of QE/mg, respectively). The EE, DCMF, ACF, and HEF exhibited great antioxidant potential by DPPH (IC50 1.71 - 32.83 µg/mL) and FRAP (IC50 0.63 - 6,71 µg/mL) assays. A maximum cytotoxic activity on A. salina of 60% was observed for the DCMF (LC50 = 856.17 µg/mL). This study contributes to the phytochemical study of S. brasiliensis since these compounds were identified for the first time in the stems of this species. The S. brasiliensis stems demonstrated to be a rich source of polyphenols compounds and exhibited high antioxidant potential without toxicity. Thus, extract and fractions obtained from the S. brasiliensis stems can be used in food supplements or as natural antioxidants in the food industry.

Innate Immune Response Assessment in Cyprinus carpio L. upon Experimental Administration with Artemia salina Bio-Encapsulated Aeromonas hydrophila Bacterin.

The present study aimed to analyze the enhancement of innate immune responses in juvenile-stage common carp (Cyprinus carpio L.), upon the administration of heat-killed Aeromonas hydrophila at a dosage of 1 × 107 CFU ml-1 through bio-encapsulation in the aquatic crustacean, Artemia salina. This work emphasizes the modulation of innate immune response when administered with the bio-encapsulated heat-killed antigen that acts as an inactivated vaccine against Motile Aeromonas Septicemia disease. Bio-encapsulated oral administration of antigens promotes innate immunity in juvenile-stage fishes. The optimization of effective bio-encapsulation of bacterin in Artemia salina nauplii was carried out and the best optimal conditions were chosen for immunization. The functional immune parameters such as myeloperoxidase, lysozyme, alkaline phosphatase, antiprotease and respiratory burst activity in serum, blood and intestinal tissue samples were analyzed along with blood differential leukocyte count and tissue histopathology studies. Both humoral and cellular immune responses analyzed were substantially induced or enhanced in the treatment groups in comparison with the control group. The results showed a significant variation in the bio-encapsulation group than the control group and also were comparable to the protection conferred with immersion route immunization under similar conditions. Thus, most of the innate non-specific immune responses are inducible, despite being constitutive of the fish immune system, to exhibit a basal level of protection and a road to better vaccination strategy in Cyprinus carpio L. aquaculture worldwide.

Investigating on the toxicity and bio-magnification potential of synthetic glitters on Artemia salina.

Our research aims to assess the toxic impacts of polyethylene terephthalate (PET) glitters on Artemia salina as a model zooplankton. The mortality rate was assessed using a Kaplan Maier plot as a function of various microplastic dosages. The ingestion of microplastics was confirmed by their presence in digestive tract and faecal matter. Gut wall damage was confirmed by dissolution of basal lamina walls and an increase in the secretory cells. A significant decrease in the activities of cholinesterase (ChE) and glutathione-S-transferase (GST) were noted. A decrease in catalase activity could be correlated to an increase in the generation of reactive oxygen species (ROS). Cysts incubated in presence of microplastics exhibited delay in their hatching into 'umbrella' and 'instar' stages. The data presented in the study would be useful for scientists working on discovering new sources of microplastics, related scientific evidences, image data and model of study.