Over 100 years of vitamin E: An overview from synthesis and formulation to application in animal nutrition.

The groundbreaking discovery of vitamin E by Evans and Bishop in 1922 was an important milestone in vitamin research, inspiring further investigation into its crucial role in both human and animal nutrition. Supplementing vitamin E has been proved to enhance multiple key physiological systems such as the reproductive, circulatory, nervous and muscular systems. As the main antioxidant in the blood and on a cellular level, vitamin E maintains the integrity of both cellular and vascular membranes and thus modulates the immune system. This overview showcases important and innovative routes for synthesizing vitamin E on a commercial scale, provides cutting-edge insights into formulation concepts for successful product form development and emphasizes the importance and future of vitamin E in healthy and sustainable animal nutrition.

Testing biological actions of medicinal plants from northern Vietnam on zebrafish embryos and larvae: Developmental, behavioral, and putative therapeutical effects.

Evaluating the risks and benefits of using traditional medicinal plants is of utmost importance for a huge fraction of the human population, in particular in Northern Vietnam. Zebrafish are increasingly used as a simple vertebrate model for testing toxic and physiological effects of compounds, especially on development. Here, we tested 12 ethanolic extracts from popular medicinal plants collected in northern Vietnam for their effects on zebrafish survival and development during the first 4 days after fertilization. We characterized more in detail their effects on epiboly, hatching, growth, necrosis, body curvature, angiogenesis, skeletal development and mostly increased movement behavior. Finally, we confirm the effect on epiboly caused by the Mahonia bealei extract by staining the actin filaments and performing whole genome gene expression analysis. Further, we show that this extract also inhibits cell migration of mouse embryo fibroblasts. Finally, we analyzed the chemical composition of the Mahonia bealei extract and test the effects of its major components. In conclusion, we show that traditional medicinal plant extracts are able to affect zebrafish early life stage development to various degrees. In addition, we show that an extract causing delay in epiboly also inhibits mammalian cell migration, suggesting that this effect may serve as a preliminary test for identifying extracts that inhibit cancer metastasis.

Penicillium digitatum as a Model Fungus for Detecting Antifungal Activity of Botanicals: An Evaluation on Vietnamese Medicinal Plant Extracts.

Medicinal plants play important roles in traditional medicine, and numerous compounds among them have been recognized for their antimicrobial activity. However, little is known about the potential of Vietnamese medicinal plants for antifungal activity. In this study, we examined the antagonistic activity of twelve medicinal plant species collected in Northern Vietnam against Penicillium digitatum, Aspergillus flavus, Aspergillus fumigatus, and Candida albicans. The results showed that the antifungal activities of the crude extracts from Mahonia bealei, Ficus semicordata, and Gnetum montanum were clearly detected with the citrus postharvest pathogen P. digitatum. These extracts could fully inhibit the growth of P. digitatum on the agar medium, and on the infected citrus fruits at concentrations of 300-1000 µg/mL. Meanwhile, the other tested fungi were less sensitive to the antagonistic activity of the plant extracts. In particular, we found that the ethanolic extract of M. bealei displayed a broad-spectrum antifungal activity against all four pathogenic fungi. Analysis of this crude extract by enrichment coupled with high-performance liquid chromatography revealed that berberine and palmatine are major metabolites. Additional inspections indicated berberine as the key compound responsible for the antifungal activity of the M. bealei ethanolic extract. Our study provides a better understanding of the potential of Vietnamese medicinal plant resources for combating fungal pathogens. This work also highlights that the citrus pathogen P. digitatum can be employed as a model fungus for screening the antifungal activity of botanicals.

Notes on Culturable Endophytic Microorganisms Isolated from 14 Medicinal Plants in Vietnam: A Diversity Analysis to Predict the Host-Microbe Correlations.

Endophytes can generate a cornucopia of marvelous bioactive secondary metabolites useful for mankind but their biodiversity and associations with host plants are still elusive. In this study, we explored the culturable endophytic microorganisms associated with 14 medicinal plants that are of high socio-economic value and/or reportedly endemic to northern Vietnam. Specifically, we isolated the endophytic microorganisms by applying surface sterilization methods and identified them based on morphological and rDNA sequence analyses. Agglomerative Hierarchical Clustering (AHC) and Principal Component Analysis (PCA) were used to analyze the correlations between the taxonomic affiliations of the culturable endophytes and the characteristics of their hosts. Most of the culturable endophytes obtained were bacteria (80), and few of those were actinomycetes (15) and fungi (8). Many of them are reported to be endophytes of medicinal plants for the first time. A number of plants (5) are also reported for the first time to contain microbial endophytes, while some plants with powerful pharmaceutical potential harbor unique endophytes. Furthermore, our results reveal a strikingly close relation between the compositions of bacterial and fungal isolates from plants having anti-bacterial activity and those from plants having anti-inflammatory activity, or between the compositions of the microbial endophytic isolates from plants having anti-cancer activity and those from plants having antioxidant activity. Altogether, the results provide new findings which can be inspiring for further in-depth studies to explore and exploit the relationships between medicinal plants and their associated endophytes in northern Vietnam and world-wide.

In vitro antiviral activities of ethanol and aqueous extracts of Vietnamese traditional medicinal plants against Porcine Epidemic Diarrhea virus: a coronavirus family member.

Porcine epidemic diarrhea virus (PEDV) causes diarrhea in pigs leading to severe illnesses and high mortality rates. The development of medicinal agents to treat PEDV infection is therefore crucial. In this study, antiviral activities against PEDV of ethanol and aqueous extracts of 17 Vietnamese traditional medicinal plants were evaluated using the cytopathic effect-based assay. The results showed that 14 out of 17 medicinal plants could inhibit the cytopathic effect of PEDV. The ethanol extract of Stixis scandens was identified as the most active extract with its MIC (minimum inhibitory concentration) being 0.15 µg/mL. Other plant extracts also displayed strong antiviral activity against PEDV, including Anisomeles indica, Pericampylus glaucus and Croton kongensis. The results demonstrate that certain medicinal plants have a high antiviral potential and may serve as a lead to develop novel pharmaceutical agents to cure PED as well as the diseases caused by other coronaviruses.

Toxicity and Anti-Proliferative Properties of Anisomeles indica Ethanol Extract on Cervical Cancer HeLa Cells and Zebrafish Embryos.

In this study, we showed that crude extract of Anisomeles indica (AI-EtE) expressed its toxicity to HeLa cells with an IC50 dose of 38.8 µg/mL and to zebrafish embryos with malformations, lethality and hatching inhibition at 72-hpf at doses higher than 75 µg/mL. More interestingly, flow cytometry revealed that AI-EtE significantly promoted the number of cells entering apoptotic. Accordingly, the transcript levels of BAX, CASPASE-8, and CASPASE-3 in the cells treated with AI-EtE at IC50 dose were 1.55-, 1.62-, and 2.45-fold higher than those in the control cells, respectively. Moreover, treatment with AI-EtE caused cell cycle arrest at the G1 phase in a p53-independent manner. Particularly, percentages of AI-EtE-treated cells in G1, S, G2/M were, respectively 85%, 6.7% and 6.4%; while percentages of control cells in G1, S, G2/M were 64%, 15% and 19%, respectively. Consistent with cell cycle arrest, the expressions of CDKN1A and CDNK2A in AI-EtE-treated cells were up-regulated 1.9- and 1.64-fold, respectively. Significantly, treatment with AI-EtE also decreased anchorage-independent growth of HeLa cells. In conclusion, we suggest that Anisomeles indica can be considered as a medicinal plant with a possible use against cervical cancer cells; however, the used dose should be carefully monitored, especially when applying to pregnant women.

Developmental toxicity of Clerodendrum cyrtophyllum turcz ethanol extract in zebrafish embryo.

ETHNOPHARMACOLOGICAL RELEVANCE: Clerodendrum cyrtophyllum Turcz has been used in traditional medicine for the treatment of various diseases. In spite of its therapeutic applications, research on its toxicity and teratogenicity is still limited. AIM OF THE STUDY: The study aimed to investigate the developmental toxicity of the ethanol extract of C. cyrtophyllum (EE) in zebrafish embryo model. MATERIAL AND METHODS: Major compounds from crude ethanol extract of Clerodendron cyrtophyllum Turcz leaves were determined using HPLC-DAD-Orbitrap-MS analysis. The developmental toxicity of EE were investigated using zebrafish embryo model. Zebrafish embryos at 6 h post-fertilization (hpf) were treated with EE at different concentrations. Egg coagulation, mortality, hatching, yolk sac edema, pericardial edema and teratogenicity were recorded each day for during a 5-day exposure. At time point 120 hpf, body length, pericardial area, heartbeat and yolk sac area were assessed. In order to elucidate molecular mechanisms for the developmental toxicity of EE, we further evaluated the effects of the EE on the expression of genes involved on signaling pathways affecting fish embryo's development such as heart development (gata5, myl7, myh6, has2, hand2, nkx 2.5), oxidative stress (cat, sod1, gpx4, gstp2), wnt pathway (ß-catenin, wnt3a, wnt5, wnt8a, wnt11), or cell apoptosis (p53, bax, bcl2, casp3, casp8, casp9, apaf-1, gadd45bb) using qRT-PCR analysis. RESULTS: Our results demonstrated that three major components including acteoside, cirsilineol and cirsilineol-4'-O-ß-D-glucopyranoside were identified from EE. EE exposure during 6-96 h post-fertilization (hpf) at doses ranging from 80 to 200 µg/mL increased embryo mortality and reduced hatching rate. EE exposure at 20 and 40 µg/mL until 72-120 hpf induced a series of malformations, including yolk sac edema, pericardial edema, spine deformation, shorter body length. Based on two prediction models using a teratogenic index (TI), a 25% lethality concentration (LD25) and the no observed-adverse-effect level (NOAEL), EE is considered as teratogenic for zebrafish embryos with TI (LC50/EC50) and LD25/NOAEC values at 96 hpf reaching 3.87 and 15.73 respectively. The mRNA expression levels of p53, casp8, bax/bcl2, gstp2, nkx2.5, wnt3a, wnt11, gadd45bb and gata5 were significantly upregulated by EE exposure at 20 and 40 µg/mL while the expression of wnt5, hand2 and bcl2 were downregulated. CONCLUSIONS: These results provide evidence for toxicity effects of EE to embryo stages and provide an insight into the potential toxicity mechanisms on embryonic development.

Anti-inflammatory properties of the ethanol extract from Clerodendrum cyrtophyllum Turcz based on in vitro and in vivo studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Clerodendrum cyrtophyllum Turcz, a plant belonging to the Verbenaceae family, has been used in traditional medicine for the treatment of various inflammatory diseases in many Asian countries. AIM OF THE STUDY: The study aimed to evaluate anti-inflammatory properties of the ethanol extract from Clerodendrum cyrthophyllum Turcz leaves (EE-CC) through in vitro and in vivo models. MATERIAL AND METHODS: Total phenolic and flavonoid contents in the extract were determined using colorimetric methods and HPTLC. In red blood cell membrane stabilization model, rat erythrocyte suspension was treated with crude ethanol extract at different concentrations, the hemoglobin content of the supernatant solution released by red blood hemolysis was estimated. We also evaluated the effects of the ethanol extract from this plant on the production of nitric oxide (NO), tumor necrosis factor alpha (TNF-α) in stimulated RAW 264.7 cells. In order to elucidate its anti-inflammatory molecular mechanisms, we further evaluated the effects of the EE-CC on the expression of the inflammatory genes in inflammation-induced zebrafish model by tail-cutting using qPCR analysis. RESULTS: Colorimetric methods and HPTLC revealed high phenolic and flavonoid contents in the extract. In the red blood cell membrane stabilization model, the amount of hemoglobin released by red blood hemolysis significantly decreased in the presence of EE-CC, demonstrating a strong membrane stabilizing activity. EE-CC did not cause any toxic effect on cell viability but strongly inhibited NO and TNF-ɑ release due to LPS induction. The association with EE-CC significantly reduced the expression of cox-2, pla2, c3a, il-1(il1fma), il-8 (cxcl8b.1), tnf-α, and nf-ƙb, while increased the expression of the anti-inflammatory cytokine il-10 gene in cut-tail induced inflammation of zebrafish model. CONCLUSIONS: Taken together, the results suggest that the raw ethanol extract from C. cyrtophyllum Turcz leaves presents potent anti-inflammatory activities and may be useful for the treatment of various inflammatory diseases.

Arsenic removal from groundwater using iron electrocoagulation: effect of charge dosage rate.

We demonstrate that electrocoagulation (EC) using iron electrodes can reduce arsenic below 10 µg/L in synthetic Bangladesh groundwater and in real groundwater from Bangladesh and Cambodia, while investigating the effect of operating parameters that are often overlooked, such as charge dosage rate. We measure arsenic removal performance over a larger range of current density than in any other single previous EC study (5000-fold: 0.02 - 100 mA/cm(2)) and over a wide range of charge dosage rates (0.060 - 18 Coulombs/L/min). We find that charge dosage rate has significant effects on both removal capacity (µg-As removed/Coulomb) and treatment time and is the appropriate parameter to maintain performance when scaling to different active areas and volumes. We estimate the operating costs of EC treatment in Bangladesh groundwater to be $0.22/m(3). Waste sludge (~80 - 120 mg/L), when tested with the Toxic Characteristic Leachate Protocol (TCLP), is characterized as non-hazardous. Although our focus is on developing a practical device, our results suggest that As[III] is mostly oxidized via a chemical pathway and does not rely on processes occurring at the anode. Supplementary materials are available for this article. Go to the publisher's online edition of Journal of Environmental Science and Health, Part A, to view the free supplemental file.

The application of reporter gene assays for the detection of endocrine disruptors in sport supplements.

The increasing availability and use of sports supplements is of concern as highlighted by a number of studies reporting endocrine disruptor contamination in such products. The health food supplement market, including sport supplements, is growing across the Developed World. Therefore, the need to ensure the quality and safety of sport supplements for the consumer is essential. The development and validation of two reporter gene assays coupled with solid phase sample preparation enabling the detection of estrogenic and androgenic constituents in sport supplements is reported. Both assays were shown to be of high sensitivity with the estrogen and androgen reporter gene assays having an EC(50) of 0.01 ng mL(-1) and 0.16 ng mL(-1) respectively. The developed assays were applied in a survey of 63 sport supplements samples obtained across the Island of Ireland with an additional seven reference samples previously investigated using LC-MS/MS. Androgen and estrogen bio-activity was found in 71% of the investigated samples. Bio-activity profiling was further broken down into agonists, partial agonists and antagonists. Supplements (13) with the strongest estrogenic bio-activity were chosen for further investigation. LC-MS/MS analysis of these samples determined the presence of phytoestrogens in seven of them. Supplements (38) with androgen bio-activity were also selected for further investigation. Androgen agonist bio-activity was detected in 12 supplements, antagonistic bio-activity was detected in 16 and partial antagonistic bio-activity was detected in 10. A further group of supplements (7) did not present androgenic bio-activity when tested alone but enhanced the androgenic agonist bio-activity of dihydrotestosterone when combined. The developed assays offer advantages in detection of known, unknown and low-level mixtures of endocrine disruptors over existing analytical screening techniques. For the detection and identification of constituent hormonally active compounds the combination of biological and physio-chemical techniques is optimal.

Leukemic cell xenograft in zebrafish embryo for investigating drug efficacy.

Zebrafish were proposed as an alternative to mammalian models to assess the efficacy and toxicity of antileukemic drugs. Due to the limited number of transgenic zebrafish leukemia models, we explored human leukemic cell xenograft in zebrafish embryos. Human leukemic cell lines and blast cells sorted from patients with acute myelogenous leukemia were injected 48 hours post-fertilization and remained in the circulation of zebrafish embryos for several days without affecting their development. Imatinib and oxaphorines did not demonstrate any toxicity on normal zebrafish embryos and decreased the leukemic burden in animals xenografted with sensitive leukemic cell lines. Two other molecules, all-trans retinoic acid and the translation inhibitor 4EGI-1, demonstrated teratogenic effects at concentrations shown to be efficient in vitro, which precluded investigation of their antileukemic activity in such models. Altogether, xenografted leukemic cells in zebrafish embryos are a pharmacologically relevant model for screening non-teratogenic drugs.

Expression of the zinc finger Egr1 gene during zebrafish embryonic development.

Egr1 is a highly conserved zinc finger protein which plays important roles in many aspects of vertebrate development and in the adult. The cDNA coding for zebrafish Egr1 was obtained and its expression pattern was examined during zebrafish embryogenesis using whole-mount in situ hybridization. Egr1 mRNA is first detected in adaxial cells in the presomitic mesoderm between 11 and 20 h post-fertilization (hpf), spanning the 4-24 somite stages. Later, Egr1 expression is observed only in specific brain areas, starting at 21 hpf and subsequently increasing in distinct domains of the central nervous system, e.g. in the telencephalon, diencephalon and hypothalamus. Between 24 and 48 hpf, Egr1 is expressed in specific domains of the hypothalamus, mesencephalon, tegmentum, pharynx, retina, otic vesicle and heart.