Purified Pyropia yezoensis Pigment Extract-Based Tandem Dye Synthesis.

Red phycoerythrin (R-PE) is a highly valuable protein found in an edible seaweed, Pyropia yezoensis. It is used extensively in biotechnological applications due to its strong fluorescence and stability in diverse environments. However, the current methods for extracting and purifying R-PE are costly and unsustainable. The aim of the present study was to enhance the financial viability of the process by improving the extraction and purification of R-PE from dried P. yezoensis and to further enhance R-PE value by incorporating it into a tandem dye for molecular biology applications. A combination of ultrafiltration, ion exchange chromatography, and gel filtration yielded concentrated (1 mg·mL-1) R-PE at 99% purity. Using purified PE and Cyanine5 (Cy5), an organic tandem dye, phycoerythrin-Cy5 (PE-Cy5), was subsequently established. In comparison to a commercially available tandem dye, PE-Cy5 exhibited 202.3% stronger fluorescence, rendering it suitable for imaging and analyzes that require high sensitivity, enhanced signal-to-noise ratio, broad dynamic range, or shorter exposure times to minimize potential damage to samples. The techno-economic analysis confirmed the financial feasibility of the innovative technique for the extraction and purification of R-PE and PE-Cy5 production.

A rapid and multi-endpoint ecotoxicological test using Mychonastes afer for efficient screening of metals and herbicides.

Microalgal growth-based tests are international standards for ecotoxicity assessment; however, their long exposure times, large sample volumes, and reliance on a single growth-endpoint make them inadequate for rapid toxicity screening. Here, we aimed to develop a rapid and simple ecotoxicological test using the fast-growing green alga Mychonastes afer, with multiple endpoints-growth, lipid content, and photosynthesis. We exposed M. afer to two metals-silver and copper-and two herbicides-atrazine and diuron-for 24 h and identified the most sensitive and reliable endpoints for each toxicant: the maximum electron transport rate (ETRmax) for Ag, Cu and atrazine, and the lipid content for diuron. Lipid content was found to be both a sensitive and reliable biomarker, meeting the effluent limit guidelines in both the Republic of Korea and the USA. The sensitivity of M. afer to Ag and atrazine also closely matched the HC5 values derived from the species sensitivity distribution approach, confirming its reliability for setting regulatory concentrations of these contaminants. Our calculated predicted no-effect concentration (PNEC) values were similar to established European Union PNECs for Ag, Cu, atrazine, and diuron, underlining the utility of these biological endpoints for ecological risk assessment and regulatory decision making. This method required lower sample volume (2 mL vs 100 mL) and exposure time (24 h vs 72-120 h) than conventional green algal tests, and eliminated the need for labour-intensive cell counting, expensive equipment, and chlorophyll fluorescence measurement expertise. Overall, this M. afer test can be a valuable tool for the rapid screening of wastewater for metals and herbicides, contributing to environmental protection and management practices.

Photosynthetic and biochemical traits change in the green-tide-forming macroalga Ulva pertusa during sporulation1.

The physiological and biochemical changes in the green macroalga Ulva pertusa during the progression of sporulation have been characterized. The transition from the vegetative to the sporulation stage was accompanied by an increase in chlorophyll a (Chl a), chlorophyll b (Chl b), and carotenoid content, as well as an increase in DPPH scavenging and responsiveness to diphenylamine. However, oxygen evolution and maximum electron transport rate decreased. The discrepancy between photosynthetic performance and pigment content might relate to the self-shading of spores within a sporangium. Spore-forming U. pertusa thalli were low-light-adapted, due to an increase in the number of photosynthetic units. Decreased electron transport during sporulation might trigger sporulation, as for some cyanobacteria and other Ulva spp., via oxidization of the plastoquinone pool and cyclic phosphorylation, thus producing ATP to generate carbon and nitrogen skeletons required for spores. It is thus concluded that carotenoids function both in spore initiation and/or maturation and in their photoprotection.

Rapid toxicity assessment of six antifouling booster biocides using a microplate-based chlorophyll fluorescence in Undaria pinnatifida gametophytes.

Biocides of antifouling agents can cause problems in marine ecosystems by damaging to non-target algal species. Aquatic bioassays are important means of assessing the quality of water containing mixtures of contaminants and of providing a safety standard for water management in an ecological context. In this study, a rapid, sensitive and inexpensive test method was developed using free-living male and female gametophytes of the brown macroalga Undaria pinnatifida. A conventional fluorometer was employed to evaluate the acute (48 h) toxic effects of six antifouling biocides: 4,5-Dichloro-2-octyl-isothiazolone (DCOIT), diuron, irgarol, medetomidine, tolylfluanid, zinc pyrithione (ZnPT). The decreasing toxicity in male and female gametophytes as estimated by EC50 (effective concentration at which 50% inhibition occurs) values was: diuron (0.037 and 0.128 mg l-1, respectively) > irgarol (0.096 and 0.172 mg l-1, respectively) > tolylfluanid (0.238 and 1.028 mg l-1, respectively) > DCOIT (1.015 and 0.890 mg l-1, respectively) > medetomidine (12.032 and 12.763 mg l-1, respectively). For ZnPT, 50% fluorescence inhibition of U. pinnatifida gametophytes occurred at concentrations above 0.4 mg l-1. The Undaria method is rapid, simple, practical, and cost-effective for the detection of photosynthesis-inhibiting biocides, thus making a useful tool for testing the toxicity of antifouling agents in marine environments.

The Anti-Inflammatory Effects of Angiogenin in an Endotoxin Induced Uveitis in Rats.

Angiogenin (ANG) is involved in the innate immune system and inflammatory disease. The aim of this study is to evaluate the anti-inflammatory effects of ANG in an endotoxin induced uveitis (EIU) rat model and the pathways involved. EIU rats were treated with balanced salt solution (BSS), a non-functional mutant ANG (mANG), or wild-type ANG (ANG). The integrity of the blood-aqueous barrier was evaluated by the infiltrating cell and protein concentrations in aqueous humor. Histopathology, Western blot, and real-time qRT-PCR of aqueous humor and ocular tissue were performed to analyze inflammatory cytokines and transcription factors. EIU treated with ANG had decreased inflammatory cells and protein concentrations in the anterior chamber. Compared to BSS and mANG, ANG treatment showed reduced expression of IL-1ß, IL-8, TNF-α, and Myd88, while the expression of IL-4 and IL-10 was increased. Western blot of ANG treatment showed decreased expression of IL-6, inducible nitric oxide synthase (iNOS), IL-1ß, TNF-α, and phosphorylated NF-κB and increased expression of IL-10. In conclusion, ANG seems to reduce effectively immune mediated inflammation in the EIU rat model by reducing the expression of proinflammatory cytokines, while increasing the expression of anti-inflammatory cytokines through pathways related to NF-κB. Therefore, ANG shows potential for effectively suppressing immune-inflammatory responses in vivo.

Towards a multi-bioassay-based index for toxicity assessment of fluvial waters.

Despite their proven reliability for revealing 'acceptable' degrees of toxicity in waste- and reclaimed waters, bioassays are rarely used to assess the toxicity of hazardous contaminants present in natural waters. In this study, we used organisms from different trophic levels to assess the toxicity of water samples collected from four different South Korean rivers. The main objective was to develop a multi-descriptor index of toxicity for undiluted river water. The responses of six test organisms (Aliivibrio fischeri, Pseudokirchneriella subcapitata, Heterocypris incongruens, Moina macrocopa, Danio rerio and Lemna minor) after laboratory exposure to water samples were considered for this index, as well as the frequency of teratologies in diatom assemblages. Each individual test was attributed a toxicity class and score (three levels; no toxicity = 0, low toxicity = 1, confirmed toxicity = 2) based on the organism's response after exposure and a total score was calculated. The proposed index also considers the number of test organisms that received the highest toxicity score (value = 2). An overall toxicity category was then attributed to the water sample based on those two metrics: A = no toxicity, B = slight toxicity, C = moderate toxicity; D = toxicity and E = high toxicity. The susceptibility of the test organisms varied greatly and the sensitivity of their response also differed among bioassays. The combined responses of organisms from different trophic levels and with different life strategies provided multi-level diagnostic information about the intensity and the nature of contamination.

Testing the toxicity of metals, phenol, effluents, and receiving waters by root elongation in Lactuca sativa L.

Phytotoxicity tests using higher plants are among the most simple, sensitive, and cost-effective of the methods available for ecotoxicity testing. In the present study, a hydroponic-based phytotoxicity test using seeds of Lactuca sativa was used to evaluate the water quality of receiving waters and effluents near two industrial sites (Soyo and Daejon) in Korea with respect to the toxicity of 10 metals (As, Cd, Cr, Cu, Fe, Pb, Mn, Hg, Ni, Zn) and phenol, and of the receiving waters and effluents themselves. First, the L. sativa hydroponic bioassay was used to determine whether the receiving water or effluents were toxic; then, the responsible toxicant was identified. The results obtained with the L. sativa bioassay ranked the EC50 toxicities of the investigated metal ions and phenol as: Cd > Ni > Cu > Zn > Hg > phenol > As > Mn > Cr > Pb > Fe. We found that Zn was the toxicant principally responsible for toxicity in Daejeon effluents. The Daejeon field effluent had a higher Zn concentration than permitted by the effluent discharge criteria of the Ministry of Environment of Korea. Our conclusion on the importance of Zn toxicity was supported by the results of the L. sativa hydroponic assay, which showed that the concentration of Zn required to inhibit root elongation in L. sativa by 50% (EC50) was higher in the Daejeon field effluent than that of pure Zn. More importantly, we proved that the L. sativa hydroponic test method can be applied not only as an alternative tool for determining whether a given waste is acceptable for discharge into public water bodies, but also as an alternative method for measuring the safety of aquatic environments using EC20 values, with respect to the water pollutants investigated (i.e., Cd, Cr, Cu, Pb, Mn, Hg, Ni, Zn, and phenol).

The sensitivity of an hydroponic lettuce root elongation bioassay to metals, phenol and wastewaters.

The root elongation bioassay is one of the most straightforward test methods used for environmental monitoring in terms of simplicity, rapidity and economy since it merely requires filter paper, distilled water and Petri dishes. However, filter paper as a support material is known to be problematic as it can reduce the sensitivity of the test. The newly developed hydroponic method reported here differs from the conventional root elongation method (US EPA filter paper method) in that no support material is used and the exposure time is shorter (48 h in this test versus 120 h in the US EPA test). For metals, the hydroponic test method was 3.3 (for Hg) to 57 (for Cu) times more sensitive than the US EPA method with the rank orders of sensitivity, estimated from EC50 values, being Cu≥Cd>Ni≥Zn≥Hg for the former and Hg≥Cu≥Ni≥Cd≥Zn for the latter methods. For phenol, the results did not differ significantly; EC50 values were 124 mg L(-1) and 108-180 mg L(-1) for the hydroponic and filter paper methods, respectively. Lettuce was less sensitive than daphnids to wastewaters, but the root elongation response appears to be wastewater-specific and is especially sensitive for detecting the presence of fluorine. The new hydroponic test thus provides many practical advantages, especially in terms of cost and time-effectiveness requiring only a well plate, a small volume of distilled water and short exposure period; furthermore, no specialist expertise is required. The method is simpler than the conventional EPA technique in not using filter paper which can influence the sensitivity of the test. Additionally, plant seeds have a long shelf-life and require little or no maintenance.

Phytochemical constituents from the florets of tiger grass Thysanolaena latifolia from Nepal.

Phytochemical investigation on the florets of Thysanolaena latifolia leads to the isolation of a new compound 6″-O-acetylorientin-2″-O-α-L-rhamnopyranoside (1), named amrisoside and other 34 known compounds. The chemical structures of the compounds were determined from the interpretation of spectroscopic data including NMR, MS, and IR. This is the first report of phytochemical constituents from the monotypic genus Thysanolaena.

Three New Isoprenylated Flavonoids from the Root Bark of Morus alba.

Phytochemical investigation of the root bark of Morus alba has led to the isolation and identification of three new isoprenylated flavonoids, namely sanggenon U (1), sanggenon V (2), and sanggenon W (3), along with four known isoprenylated flavonoids: euchrenone a7 (4), sanggenon J (5), kuwanon E (6), and kuwanon S (7). All compounds were isolated by repeated silica gel (SiO2), octadecyl SiO2 (ODS), and Sephadex LH-20 open column chromatography. The structure of the compounds were determined based on spectroscopic analyses, including nuclear magnetic resonance (NMR), mass spectrometry (MS), circular dichroism (CD), and infrared (IR). In addition, compounds 1-4 were isolated for the first time from the root bark of M. alba in this study.

Antidepressant-Like Effects of Sanggenon G, Isolated from the Root Bark of Morus alba, in Rats: Involvement of the Serotonergic System.

The root bark of Morus alba is commonly used as an alternative medicine due to its numerous health benefits in humans. However, the antidepressant effects of various active components from M. alba have not been fully elucidated. In this study, we aimed to determine whether sanggenon G, an active compound isolated from the root bark of M. alba, exhibited antidepressant-like activity in rats subjected to forced swim test (FST)-induced depression. Acute treatment of rats with sanggenon G (30 mg/kg, intraperitoneally (i.p.)) significantly reduced immobility time and increased swimming time without any significant change in climbing. Rats treated with sanggenon G also exhibited a decrease in the limbic hypothalamic-pituitary-adrenal (HPA) axis response to the FST, as indicated by attenuation of the corticosterone response and decreased c-Fos immunoreactivity in the hypothalamic paraventricular nucleus (PVN). In addition, the antidepressant-like effects of sanggenon G were significantly inhibited by WAY100635 (1 mg/kg, i.p.; a selective 5-hydroxytryptamine1A (5-HT1A) receptor antagonist), but not SCH23390 (0.05 mg/kg, i.p.; a dopamine D1 receptor antagonist). Our findings suggested that the antidepressant-like effects of sanggenon G were mediated by an interaction with the serotonergic system. Further studies are needed to evaluate the potential of sanggenon G as an alternative therapeutic approach for the treatment of depression.

Two new isoarylbenzofuran diglucosides from the root bark of Morus alba.

Two new arylbenzofuran diglucopyranosides, (2″R)-(-)-moracin-O-5',3″-ß-D-diglucopyranoside (1) and (2″R)-(-)-moracin-P-5',2″-ß-d-diglucopyranoside (2), along with known arylbenzofurans, moracin M 6-ß-D-glucopyranoside (3), and an isomeric mixture of R-(-)-moracin O (4) and R-(-)-moracin P (5), were isolated from the root bark of Morus alba L. The structure of the compounds was elucidated based on mass spectrometry, infrared, 1D and 2D nuclear magnetic resonance spectroscopic data.

Two New Cyototoxic Cardenolides from the Whole Plants of Adonis multiflora Nishikawa & Koki Ito.

A phytochemical investigation of the whole plants of Adonis multiflora Nishikawa & Koki Ito. resulted in the isolation and identification of two new cardenolides--adonioside A (1) and adonioside B (6)--as well as four known cardenolides: tupichinolide (2) oleandrine (3), cryptostigmin II (4), and cymarin (5). Their structures were elucidated on the basis of NMR, MS, and IR spectroscopic analyses. Compounds 1, 2, 5, and 6 showed significant cytotoxicity against six human cancer cell lines (HCT-116, HepG2, HeLa, SK-OV-3, and SK-MEL-5, and SK-BR-3).

Cynandione A from Cynanchum wilfordii attenuates the production of inflammatory mediators in LPS-induced BV-2 microglial cells via NF-κB inactivation.

Cynanchum wilfordii is one of most widely used medicinal plants in Oriental medicine for the treatment of various conditions. In the present study, we isolated cynandione A (CA) from an extract of Cynanchum wilfordii roots (CWE) and investigated the effects of CA on the expression of inducible nitric oxide synthase (iNOS) and pro-inflammatory cytokines in lipopolysaccharide (LPS)-induced BV-2 microglial cells. CWE and CA significantly decreased LPS-induced nitric oxide production and the expression of iNOS in a concentration-dependent manner, while they (CWE up to 500 µg/mL and CA up to 80 µM) did not exhibit cytotoxic activity. Results from reverse transcription-polymerase chain reaction (RT-PCR) analysis and enzyme-linked immunosorbent assay (ELISA) showed that CA significantly attenuated the expression of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and IL-1ß in LPS-stimulated BV-2 cells. Furthermore, CA inhibited the phosphorylation of inhibitor kappa B-alpha (IκB-α) and translocation of nuclear factor-kappa B (NF-κB) to the BV-2 cell nucleus, indicating that CWE and CA may have effective anti-inflammatory activities via NF-κB inactivation in stimulated microglial cells.

Flavonoids from Machilus japonica stems and their inhibitory effects on LDL oxidation.

Stems of Machilus japonica were extracted with 80% aqueous methanol (MeOH) and the concentrated extract was successively extracted with ethyl acetate (EtOAc), normal butanol (n-BuOH), and water. Six flavonoids were isolated from the EtOAc fraction: (+)-taxifolin, afzelin, (-)-epicatechin, 5,3'-di-O-methyl-(-)-epicatechin, 5,7,3'-tri-O-methyl-(-)-epicatechin, and 5,7-di-O-methyl-3',4'-methylenedioxyflavan-3-ol. The chemical structures were identified using spectroscopic data including NMR, mass spectrometry and infrared spectroscopy. This is the first report of isolation of these six compounds from M. japonica. The compounds were evaluated for their diphenyl picryl hydrazinyl scavenging activity and inhibitory effects on low-density lipoprotein oxidation. Compounds 1 and 3-6 exhibited DPPH antioxidant activity equivalent with that of ascorbic acid, with half maximal inhibitory concentration (IC50) values of 0.16, 0.21, 0.17, 0.15 and 0.07 mM, respectively. The activity of compound 1 was similar to the positive control butylated hydroxytoluene, which had an IC50 value of 1.9 µM, while compounds 3 and 5 showed little activity. Compounds 1, 3, and 5 exhibited LDL antioxidant activity with IC50 values of 2.8, 7.1, and 4.6 µM, respectively.

Antidepressant-like effects of the ethyl acetate soluble fraction of the root bark of Morus alba on the immobility behavior of rats in the forced swim test.

In this study, the antidepressant-like effects of Morus alba fractions in rats were investigated in the forced swim test (FST). Male Wistar rats (9-week-old) were administered orally the M. alba ethyl acetate (EtOAc 30 and 100 mg/kg) and M. alba n-butanol fractions (n-BuOH 30 and 100 mg/kg) every day for 7 consecutive days. On day 7, 1 h after the final administration of the fractions, the rats were exposed to the FST. M. alba EtOAc fraction at the dose of 100 mg/kg induced a decrease in immobility behavior (p < 0.01) with a concomitant increase in both climbing (p < 0.05) and swimming (p < 0.05) behaviors when compared with the control group, and M. alba EtOAc fraction at the dose of 100 mg/kg decreased the hypothalamic-pituitary-adrenal (HPA) axis response to the stress, as indicated by an attenuated corticosterone response and decreased c-fos immunoreactivity in the hippocampal and hypothalamic paraventricular nucleus (PVN) region. These findings demonstrated that M. alba EtOAc fraction have beneficial effects on depressive behaviors and restore both altered c-fos expression and HPA activity.

Extracellular Vimentin Alters Energy Metabolism And Induces Adipocyte Hypertrophy.

BACKGRUOUND: Previous studies have reported that oxidative stress contributes to obesity characterized by adipocyte hypertrophy. However, mechanism has not been studied extensively. In the current study, we evaluated role of extracellular vimentin secreted by oxidized low-density lipoprotein (oxLDL) in energy metabolism in adipocytes. METHODS: We treated 3T3-L1-derived adipocytes with oxLDL and measured vimentin which was secreted in the media. We evaluated changes in uptake of glucose and free fatty acid, expression of molecules functioning in energy metabolism, synthesis of adenosine triphosphate (ATP) and lactate, markers for endoplasmic reticulum (ER) stress and autophagy in adipocytes treated with recombinant vimentin. RESULTS: Adipocytes secreted vimentin in response to oxLDL. Microscopic evaluation revealed that vimentin treatment induced increase in adipocyte size and increase in sizes of intracellular lipid droplets with increased intracellular triglyceride. Adipocytes treated with vimentin showed increased uptake of glucose and free fatty acid with increased expression of plasma membrane glucose transporter type 1 (GLUT1), GLUT4, and CD36. Vimentin treatment increased transcription of GLUT1 and hypoxia-inducible factor 1α (Hif-1α) but decreased GLUT4 transcription. Adipose triglyceride lipase (ATGL), peroxisome proliferator-activated receptor γ (PPARγ), sterol regulatory element-binding protein 1 (SREBP1), diacylglycerol O-acyltransferase 1 (DGAT1) and 2 were decreased by vimentin treatment. Markers for ER stress were increased and autophagy was impaired in vimentin-treated adipocytes. No change was observed in synthesis of ATP and lactate in the adipocytes treated with vimentin. CONCLUSION: We concluded that extracellular vimentin regulates expression of molecules in energy metabolism and promotes adipocyte hypertrophy. Our results show that vimentin functions in the interplay between oxidative stress and metabolism, suggesting a mechanism by which adipocyte hypertrophy is induced in oxidative stress.

Assessing industrial wastewater effluent toxicity using boosting algorithms in machine learning: A case study on ecotoxicity prediction and control strategy development.

Trace heavy metals have a tendency to persist in the effluent of industrial wastewater treatment facilities, leading to toxic effects on downstream water bodies. Traditional assessment methods relied on animal testing, but ethical concerns have rendered them unacceptable. An alternative solution is to evaluate wastewater toxicity using trophic-level aquatic organisms as bioassays. However, these bioassay methods involve costly and time-consuming chemical and biological analytical experiments. In this study, an artificial intelligence-powered water quality assessment (AiWA) approach is proposed for predicting industrial effluent ecotoxicity to further enhance the quick and cost-effective ecotoxicity assessment process. Initially, 99 samples were collected from industrial wastewater treatment plants representing 21 different industries in the Republic of Korea. Fourteen parameters were measured, encompassing both physicochemical and ecotoxicological aspects. Boosting algorithms, especially extreme gradient boosting (XGBoost) and adaptive boosting (AdaBoost), were employed for model development. XGBoost outperformed AdaBoost in terms of model performance. Feature selection analysis revealed that conductivity, copper, lead, selenium, pH, and zinc concentrations were the most suitable inputs for training the boosting model. The innovated XGBoost-based AiWA model demonstrated significantly higher performance (i.e., up to 80%) compared to conventional models with an R2 value of exceeding 0.94 and root mean square error of 3.5 toxicity unit for predicting the integrated toxicity unit (ITU). Additionally, pH and conductivity emerged as crucial indicators for reflecting ecotoxicity levels. Specially, this case study indicated that non-toxic/directly dischargeable levels (TU ≤ 1) were achieved when the pH ranged from 6.8 to 8.4 and the conductivity remained below 1651 µS/cm. These findings are expected to facilitate rapid and cost-effective detection of heavy metal ecotoxicity in industrial wastewater effluents, aiding decision-making in wastewater management.

Genome editing in macroalgae: advances and challenges.

This minireview examines the current state and challenges of genome editing in macroalgae. Despite the ecological and economic significance of this group of organisms, genome editing has seen limited applications. While CRISPR functionality has been established in two brown (Ectocarpus species 7 and Saccharina japonica) and one green seaweed (Ulva prolifera), these studies are limited to proof-of-concept demonstrations. All studies also (co)-targeted ADENINE PHOSPHORIBOSYL TRANSFERASE to enrich for mutants, due to the relatively low editing efficiencies. To advance the field, there should be a focus on advancing auxiliary technologies, particularly stable transformation, so that novel editing reagents can be screened for their efficiency. More work is also needed on understanding DNA repair in these organisms, as this is tightly linked with the editing outcomes. Developing efficient genome editing tools for macroalgae will unlock the ability to characterize their genes, which is largely uncharted terrain. Moreover, given their economic importance, genome editing will also impact breeding campaigns to develop strains that have better yields, produce more commercially valuable compounds, and show improved resilience to the impacts of global change.