Enhancement of root abscisic acid mediated osmotic regulation by macroalgal compounds promotes adaptability of rice (Oryza sativa L.) in response to progressive metal ion mediated environmental stress.

Soil deterioration is a major cause of poor agricultural productivity, necessitating sufficient nutrient inputs like fertilizers and amendments for sustainable use. As one such strategy, the current study evaluates the potential of Sargassum wightii, a brown seaweed extract, as an osmopriming agent to improve seed germination, early establishment, and competent seedling performances in acidic soil. The elemental makeup of seaweed extract (BS) showed that it included major plant macro (Potassium, Nitrogen and Phosphorous), as well as micronutrients (Magnesium and Iron) and trace elements (Zinc, Copper, and Molybdenum). While seed germination was impacted by H+ ion toxicity, seeds primed with BS emerged earlier and showed a higher germination percentage (98.2%) and energy (92.4%). BS treatments enhanced seedling growth by 63% and had a positive effect on root growth (68.2%) as well as increases in root surface area (10%) and volume (67.01%). Stressed seedlings had 76.39% and 63.2% less carotenoid and chlorophyll, respectively. In seedlings treated with BS, an increase in protein and Total Soluble Sugars content of 14.56 and 7.19%, respectively, was seen. Fourier Transform-Infra Red analysis of postharvest soil indicated improved soil health with absorbance corresponding to enhanced soil water holding capacity and organic matter. Increased abscisic acid synthesis rate and associated antioxidant enzyme system (Malondialdehyde, Glutathione peroxidases and ascorbate peroxidase) activation, along with enhanced H+ adenosine triphosphate-ase and glutathione activities, help ameliorate and deport H+ ions from cells, scavenge Reactive Oxygen Species, thus protecting cells from injury. Seaweed extract successfully reduced H+-induced ion toxicities in rice by promoting their germination, physiological, metabolically, and growth parameters that could ultimately increase their productivity and yield in a sustainable and environmentally friendly manner.

Purification and Evaluation of N-benzyl Cinnamamide from Red Seaweed Gracilaria fisheri as an Inhibitor of Vibrio harveyi AI-2 Quorum Sensing.

Previously, we reported that the ethanol extract from red seaweed Gracilaria fisheri effectively decreased biofilm formation of Vibrio harveyi. In this study, the anti-biofilm active compounds in the ethanol extract were isolated and their structures identified. The anti-biofilm fractionation assay for minimum inhibitory concentration (MIC) produced two fractions which possessed maximal inhibitory activities toward the biofilm formation of V. harveyi strains 1114 and BAA 1116. Following chromatographic separation of the bioactive fractions, two pure compounds were isolated, and their structures were elucidated using FTIR, NMR, and HR-TOF-MS. The compounds were N-benzyl cinnamamide and α-resorcylic acid. The in vitro activity assay demonstrated that both compounds inhibited the biofilm formation of V. harveyi and possessed the anti-quorum sensing activity by interfering with the bioluminescence of the bacteria. However, the N-benzyl cinnamamide was more potent than α-resorcylic acid with a 10-fold lesser MIC. The present study reveals the beneficial property of the N-benzyl cinnamamide from the ethanol extract as a lead anti-microbial drug against V. harveyi.

Variation in microbial populations and antibiotic resistance genes in mariculture sediments in the present of the seaweed Ulva fasciata and under selective pressure of oxytetracycline.

The widely distributed seaweed Ulva fasciata has nutrient absorption abilities and can be used in the bioremediation of polluted maricultural environments. This study explored microbial community and antibiotic resistance gene (ARG) variation in mariculture sediments in response to different trace levels (10, 100, and 500 µg L-1) of oxytetracycline (OTC) and the presence of Ulva fasciata. The increase in OTC level promoted nutrient (NO3_-N and PO43--P) removal mainly due to Ulva fasciata adsorption. The abundances of the Euryarchaeota and Planctomycetes phyla in sediments were positively related to the increase in OTC stress, while a negative correlation occurred for the Proteobacteria phylum via metagenomic analysis. Compared with the control system, the increase rates of total ARGs were 3.90%, 7.36% and 13.42% at the OTC levels of 10, 100 and 500 µg L-1, respectively. OTC stress mainly favoured the collateral enrichment of non-corresponding polypeptide and MLS ARGs, mainly due to the enrichment of the phyla Planctomycetes and Euryarchaeota by the synergistic effect of OTC and nutrients. The results of quantitative PCR with tetracycline resistance genes (TRGs) (tetO, tetT, tetPB, tetW and otrA) and a horizontal transfer gene (intl1) demonstrated that all of genes had much higher gene numbers in sediments after 3 months of OTC stress than in those without OTC stress, which was strongly related to the variation in the phyla Bacteroidetes, Gemmatimonadetes and Acidobacteria. The significant correlation between intl1 and the target TRGs is indicative of the important role of the horizontal transfer of integron-resistant genes in the spread of TRGs.

Heavy metal resistance in algae and its application for metal nanoparticle synthesis.

The ungenerous release of metals from different industrial, agricultural, and anthropogenic sources has resulted in heavy metal pollution. Metals with a density larger than 5 g cm-3 have been termed as heavy metals and have been stated to be potentially toxic to human and animals. Algae are known to be pioneer organisms with the potential to grow under extreme conditions including heavy metal-polluted sites. They have evolved efficient defense strategies to combat the toxic effects exerted by heavy metal ions. Most of the algal strains are reported to accumulate elevated metal ion concentration in cellular organelles. With respect to that, this review focuses on understanding the various strategies used by algal system for heavy metal resistance. Additionally, the application of this metal resistance in biosynthesis of metal nanoparticles and metal oxide nanoparticles has been investigated in details. We thereby conclude that algae serve as an excellent system for understanding metal uptake and accumulation. This thereby assists in the design and development of low-cost approaches for large-scale synthesis of nanoparticles and bioremediation approach, providing ample opportunities for future work.

Phytoremediation potential of Ulva ohnoi (Chlorophyta): Influence of temperature and salinity on the uptake efficiency and toxicity of cadmium.

Ulva ohnoi is a green macroalga with fast growth and high rates of nitrogen and phosphorus absorption. Recently, this species has been recorded in several places with record green tide formation in some of them. Using molecular tools, we herein report the first occurrence of this species in Brazil and demonstrate its potential for phytoremediation in typical environmental concentrations of Cd (0.625-15 µg L-1). Similarly, the effects of physicochemical parameters (salinity and temperature) on the toxicity and uptake efficiency of this species were evaluated. Molecular analysis of two sequences (1141 bp) obtained corroborates another 34 sequences for U. ohnoi obtained from GenBank. The addition of Cd in the medium affected photosynthetic parameters and reduced growth rate. U. ohnoi showed resistance to Cd when cultivated at 18 °C, S15 and 18-25 °C, S35, at concentrations between 0.625 and 2.5 µg. L-1 of Cd; yet, positive growth rate was maintained. Dose-dependent accumulation was observed in all combinations of factors used with a maximum value of 4.20 µg Cd per gram of dry seaweed at 15 µg. L-1 of Cd at 18 °C and S35. Maximum value of the concentration factor was 81.3 ±â€¯1.1% of Cd added at the concentration of 0.625 µg. L-1 to S15 and 18 °C. Our results demonstrate the potential of using U. ohnoi in the phytoremediation of Cd in saltwater or brackish water.

Efficient enzymatic saccharification of macroalgal biomass using a specific thermostable GH 12 endoglucanase from Aspergillus terreus JL1.

Stranded green macroalgae represents an important and renewable biomass that remains under valorized despite the numerous environmental problems generated by their accumulation in coastal regions. This work describes the isolation of a filamentous thermophile fungus identified as Aspergillus terreus JL1 that produces an efficient cellulolytic activity for green macroalgae saccharification. The characterization of the endoglucanase activity obtained after submerged fermentation showed a differential induction depending on the carbon source used with a unique isoform released when Ulva lactuca was used as inducer. The crude extract obtained hydrolyzed efficiently the untreated algal biomass (70.5%) compared to other cellulolytic extracts. The unique endoglucanase released was then purified to homogeneity (Yield: 49.6%; Specific activity: 30.1 U/mg; Purification fold: 4.36) and characterized biochemically. Its peptidic sequence was then determined and showed its belonging to the GH12. The described enzyme represents a promising biotechnological tool for algal biomass conversion.

Indole Derivatives Isolated from Brown Alga Sargassum thunbergii Inhibit Adipogenesis through AMPK Activation in 3T3-L1 Preadipocytes.

Seaweed, a popular and abundant food ingredient mainly consumed in Asian countries, is a good source of bioactive compounds with anti-obesity effects. However, the anti-obesity effects of Sargassum thunbergii have not yet been established. In this study, we isolated six indole derivatives (STCs)-indole-2-carboxaldehyde (STC-1), indole-3-carboxaldehyde (STC-2), indole-4-carboxaldehyde (STC-3), indole-5-carboxaldehyde (STC-4), indole-6-carboxaldehyde (STC-5), and indole-7-carboxaldehyde (STC-6)-from S. thunbergii and evaluated their inhibitory effects on adipocyte differentiation in 3T3-L1 cells. We found that STC-1 and STC-5 resulted in non-toxic inhibition of the differentiation of 3T3-L1 adipocytes and thus selected these compounds for further study. STC-1 and STC-5 significantly inhibited lipid accumulation and downregulated the expression of peroxisome proliferator-activated receptor-γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein 1c (SREBP-1c) in a dose-dependent manner. The specific mechanism mediating the effects of STC-1 and STC-5 was shown to be AMP-activated protein kinase (AMPK) activation. Our results demonstrated the inhibitory effect of STC-1 and STC-5 on adipogenesis through the activation of the AMPK signal pathway. Together, these findings suggested that STC-1 and STC-5 may be effective candidates for the prevention of obesity or obesity-related diseases.

Bioaccumulation and toxicity studies of macroalgae (Charophyceae) treated with aluminium: Experimental studies in the context of lake restoration.

The objective of this study was to examine the impact of aluminium on the perennial macroalgae Chara hispida L. and its bioaccumulation capacities. Aluminium (Al) was introduced into the environment in the form of polyaluminium chloride, an agent utilized in the restoration of waterbodies. Research was conducted in an experimental setting using mesocosms (volume 0.8m3) placed in the littoral zone of a lake with C. hispida. Three doses of the coagulant were applied, each with a different volume: low - 6.1g Al m-3, medium - 12.2gm-3 and high - 24.5g Al m-3. A significant acidification of environment was determined, which would imply the presence of toxic Al3+ ions. It has been demonstrated that aluminium penetrates and accumulates in the cells of the charophyte. This caused damage to the thalli, which manifested itself in chloroses, necroses, flaking of the cortex cells and softening of the thallus, whose severity was proportionate to the dose of the coagulant. The first negative signs were observed after 24h. The study shows that C. hispida is a poor accumulator of aluminium (bioconcentration factor < 200), while bioaccumulation capacity was inhibited at the concentration of approx. 2.0mg Al g-1 d.w. Accumulation in the thalli of the charophytes accounted for 58% of variation following removal of aluminium from the environment. The results of the experiment demonstrate a negative impact of aluminium on charophytes at concentrations used in aggressive restoration of lakes.

Scalable and Cost-Effective Synthesis of Highly Efficient Fe2N-Based Oxygen Reduction Catalyst Derived from Seaweed Biomass.

A simple and scalable synthesis of a 3D Fe2N-based nanoaerogel is reported with superior oxygen reduction reaction activity from waste seaweed biomass, addressed the growing energy scarcity. The merits are due to the synergistic effect of the 3D porous hybrid aerogel support with excellent electrical conductivity, convenient mass transport and O2 adsorption, and core/shell structured Fe2N/N-doped amorphous carbon nanoparticles.

Biochemical responses of filamentous algae in different aquatic ecosystems in South East Turkey and associated water quality parameters.

To the best of our knowledge, any study about biochemical response of filamentous algae in the complex freshwater ecosystems has not been found in the literature. This study was designed to explore biochemical response of filamentous algae in different water bodies from May 2013 to October 2014, using multivariate approach in the South East of Turkey. Environmental variables were measured in situ: water temperature, oxygen concentration, saturation, conductivity, salinity, pH, redox potential, and total dissolved solid. Chemical variables of aqueous samples and biochemical compounds of filamentous algae were also measured. It was found that geographic position and anthropogenic activities had strong effect on physico-chemical variables of water bodies. Variation in environmental conditions caused change in algal biomass composition due to the different response of filamentous species, also indicated by FTIR analysis. Biochemical responses not only changed from species to species, but also varied for the same species at different sampling time and sampling stations. Multivariate analyses showed that heavy metals, nutrients, and water hardness were found as the important variables governing the temporal and spatial succession and biochemical compounds. Nutrients, especially nitrate, could stimulate pigment and total protein production, whereas high metal content had adverse effects. Amount of malondialdehyde (MDA), H2O2, total thiol groups, total phenolic compounds, proline, total carbohydrate, and metal bioaccumulation by filamentous algae could be closely related with heavy metals in the ecosystems. Significant increase in MDA, H2O2, total thiol group, total phenolic compounds, and proline productions by filamentous algae and chlorosis phenomenon seemed to be an important strategy for alleviating environmental factors-induced oxidative stress as biomarkers.

Cystophloroketals A-E, Unusual Phloroglucinol-Meroterpenoid Hybrids from the Brown Alga Cystoseira tamariscifolia.

Cystophloroketals A-E (1-5), five new phloroglucinol-meroditerpenoid hybrids, have been isolated together with their putative biosynthetic precursor, the monocyclic meroditerpenoid 6, from the Mediterranean brown alga Cystoseira tamariscifolia. They represent the first examples of meroditerpenoids linked to a phloroglucinol through a 2,7-dioxabicyclo[3.2.1]octane moiety. The chemical structures, including absolute configurations, were elucidated on the basis of extensive spectroscopic analysis (HR-ESIMS, 1D and 2D NMR, and ECD) and TDDFT ECD calculations. Compounds 1-6 were tested for their antifouling activity against several marine colonizing species (bacteria, fungi, invertebrates, micro- and macroalgae). Compound 6 showed high potency for the inhibition of macrofoulers (invertebrates and macroalgae), while cystophloroketals B (2) and D (4) displayed strong inhibition of the germination of the two macroalgae tested and moderate antimicrobial activities (bacteria, microalgae, and fungi).

Thermo-Acidic Pretreatment of Beach Macroalgae from Rügen to Optimize Biomethane Production--Double Benefit with Simultaneous Bioenergy Production and Improvement of Local Beach and Waste Management.

Eutrophication is a phenomenon which can rapidly generate masses of marine macroalgae, particularly in areas with high nutrient pollution. Washed ashore, this biomass impairs coastal tourism and negatively affects the coastal ecosystem. The present study evaluates the biochemical methane potential (BMP) of a macroalgae mix (Rügen-Mix, RM (RM = Rügen-Mix)) originating from Rügen, Germany. To improve biomethane recovery, thermo-acidic pretreatment was applied to the biomass prior to biomethanation to disintegrate the biomass macrostructure. Acid hydrolysis was successfully triggered with 0.2 M industry-grade HCl at 80 °C for a 2 h period, increasing biomethane recovery by +39%, with a maximum BMP of 121 mL·g(-1) volatile solids (VS). To reduce the necessity for input material, HCl was replaced by the acidic waste product flue gas condensate (FGC). Improved performance was achieved by showing an increase in biomethane recovery of +24% and a maximum BMP of 108 mL·g(-1) VS. Continuous anaerobic digestion trials of RM were conducted for three hydraulic retention times, showing the feasibility of monodigestion. The biomethane recovery was 60 mL and 65 mL·g(-1) VS·d(-1) for thermophilic and mesophilic operation, respectively. The quality of biomethanation performance aligned to the composition of the source material which exhibited a low carbon/nitrogen ratio and an increased concentration of sulfur compounds.

Accumulation of silver by Fucus spp. (Phaeophyceae) and its toxicity to Fucus ceranoides under different salinity regimes.

Metals constitute an important group of abiotic stressors that elicit stress responses in marine algae that include the production of reactive oxygen species (ROS). Silver (Ag) is a highly toxic metal to organisms but despite this there are relatively few studies on how it affects marine macroalgae (seaweeds). In a landmark study published in 1977 the first information was provided on the accumulation of Ag in Fucus spp. (Phaeophyceae) from the Looe estuary, located in south-west England, an area with a long history of mining activity. In the present study, the estuary has been re-visited and the patterns of Ag accumulation in two Fucus spp. and sediment re-examined after 35 years. We conclude that Ag concentrations in sediment and macroalgae from specific sites within the catchment remain high, but more generally sediment concentrations have declined by approximately 65 % and the dissolved, bioavailable fraction by 24 % over this period. In addition, from laboratory studies we provide data on the speciation and toxic effects of Ag under different salinity regimes in the euryhaline brown seaweed, Fucus ceranoides. From these exposure experiments, it was found that with increasing Ag concentrations growth was inhibited and lipid peroxidation associated with ROS production increased. The magnitude of the toxic effects was greater at a salinity of 10 than 28 psu which reflects the greater bioavailability of the toxic species of Ag (Ag(+) and AgCl(0)) at reduced salinities. These findings emphasise the importance of investigating the effects of metal pollution in conjunction with other, natural, environmental stressors such as salinity.

Synergistic effects of NaOCl and ultrasound combination on the reduction of Escherichia coli and Bacillus cereus in raw laver.

The current study investigated the synergistic effects of NaOCl (50-200 ppm)/ultrasound (37 kHz, 380 W for 5-100 min) combination on the reduction of Escherichia coli and Bacillus cereus in raw laver. The synergistic reductions were not dependent on concentrations of NaOCl and times of ultrasound. Synergistic reduction ranged from 0.1 to 0.5 log10 colony-forming units (CFU)/g and 0.1-1.1 log10 CFU/g, respectively, for E. coli and B. cereus, with the largest synergistic reduction in the combination of 200 ppm NaOCl and 60-min ultrasound. Moreover, significant differences of "L" (lightness), "a" (redness), and "b" (yellowness) were not observed in combined with 50-200 ppm NaOCl and 100-min ultrasound compared to those in raw laver treated by only 100-min ultrasound. The results in the current study indicate that the combined treatment of 200 ppm NaOCl and 60-min ultrasound could be regarded a potential optimum hurdle approach in the seaweed production, processing, and distribution process to enhance seaweed safety.

Seaweeds early development: detrimental effects of desiccation and attenuation by algal extracts.

The effects of desiccation on the early development stages of Mazzaella laminarioides, Scytosiphon lomentaria and Lessonia nigrescens, algal species with different patterns of distribution across the intertidal zone, were examined in the laboratory. In addition, the protective effect against desiccation was evaluated using algal extracts, including those from Porphyra columbina, a macroalga tolerant to desiccation that lives in the uppermost part of the intertidal zone. Our results showed that M. laminarioides displayed the highest resistance to daily desiccation, followed by S. lomentaria, whereas L. nigrescens was the most susceptible. Spores from L. nigrescens exposed to desiccation, although being able to germinate, ceased further post-germination development. In addition, our results showed that all species exposed to extracts from desiccated P. columbina successfully completed their development and strongly suggest the occurrence of compounds with protective properties that help in attenuating the stress caused by desiccation. Finally, our results indicate that the magnitude of the effects generated by desiccation on the early algal development is related to the position of the species in the intertidal zone, and that the protective effects of P. columbina extracts reveal an exceptional metabolism of this species under desiccation stress.

Interactions of silver nanoparticles with the marine macroalga, Ulva lactuca.

The marine macroalga, Ulva lactuca, has been exposed for 48 h to different concentrations of Ag added as either silver nanoparticles (AgNP) or aqueous metal (AgNO(3)) and the resulting toxicity, estimated from reductions in quenching of chlorophyll-a fluorescence, and accumulation of Ag measured. Aqueous Ag was toxic at available concentrations as low as about 2.5 µg l(-1) and exhibited considerable accumulation that could be defined by the Langmuir equation. AgNP were not phytotoxic to the macroalga at available Ag concentrations up to at least 15 µg l(-1) and metal measured in U. lactuca was attributed to a physical association of nanoparticles at the algal surface. At higher AgNP concentrations, a dose-response relationship was observed that was similar to that for aqueous Ag recorded at much lower concentrations. These findings suggest that AgNP are only indirectly toxic to marine algae through the dissolution of Ag(+) ions into bulk sea water, albeit at concentrations orders of magnitude greater than those predicted in the environment.

Inter-population comparisons of copper resistance and accumulation in the red seaweed, Gracilariopsis longissima.

Copper (Cu) resistance and accumulation of five populations of the red seaweed Gracilariopsis longissima collected from sites in south west England (Fal Estuary, Helford Estuary and Chesil Fleet) that differ in their degree of Cu contamination was assessed under controlled laboratory conditions, on two separate occasions (April and October). The effects of a range of Cu concentrations (0-250 µg l(-1)) on relative growth rates was the same for all populations with reductions observable at concentrations as low as 12 µg l(-1) and cessation of growth at 250 µg l(-1). There was no significant difference in the calculated EC(50) values for the April and October samples, with means of 31.1 and 25.8 µg l(-1), respectively. Over the range of concentrations used in this study, copper content increased linearly and the pattern of accumulation was the same for all populations at both time periods. From the linear regressions of the pooled data a concentration factor of 2.25 × 10(3) was calculated. These results imply that G. longissima has an innate tolerance to Cu and that populations have not evolved copper-tolerant ecotypes. In laboratory studies, accumulated Cu was released when transferred to 'clean' seawater with approximately 80% being lost after 8 days, with no significant difference between populations in their response. The results from a 30 days in situ transplantation experiment using two populations from the Fal Estuary provided further evidence for dynamic changes in Cu content in response to changes in Cu bioavailability. The findings in this study are discussed in the context of implications for seaweed biomonitoring.

Toxic effects of imidazolium ionic liquids on the green seaweed Ulva lactuca: oxidative stress and DNA damage.

The green credentials of ionic liquids (ILs) are being increasingly questioned due to the growing evidence of their toxicity to aquatic ecosystems, although the mechanisms of toxicity are unknown. This study provides insights into the mechanism of toxicity and biological effects of 1-alkyl-3-methylimidazolium bromide [C(n)mim]Br (n = 4 to 16) on the marine macroalga Ulva lactuca. The cell viability of this alga during IL exposure was found to be negatively correlated to the chain length of the alkyl group. The IL ([C(12)mim]Br) exposure triggers the generation of reactive oxygen species (ROS viz. O(2)(•-), H(2)O(2), and OH(•)), damage of the membrane and DNA, and inhibition of antioxidant systems in the alga. The enhanced production of ROS and lipid peroxidation in the alga subjected to LC(50) concentration for 4 days was largely attributed to lipoxygenase (LOX) activity coupled with the induction of two LOX isoforms (~80 kDa and ~55 kDa). Pretreatment of the algal thallus with enzyme inhibitors such as diphenylene iodonium, sodium azide, cantharidin, and oxadiazoloquinoxalin-1-one, prior to [C(12)mim]Br exposure showed the regulation of ROS by the activation of membrane bound NADPH-oxidase and cytochrome oxidase. The IL exposure resulted in the accumulation of n-3 and n-6 fatty acids at 0.5 LC(50) concentration indicating the induction of desaturase enzymes. Furthermore, antioxidant enzyme activities such as superoxide dismutase (SOD), ascorbate peroxidase (APX), and glutathione reductase (GR) were enhanced by 1.3-2.0-fold, while glutathione peroxidase (GSH-Px) diminished, together with a higher regeneration rate of reduced ascorbate and glutathione. The isoforms of antioxidant enzymes, namely, Mn-SOD (~85 kDa), APX (~125 and 45 kDa), and GR (~135 kDa) regulated differentially to IL exposure. The comet assay performed for the first time for seaweeds revealed the significant induction of DNA damage (>50-70% increase in % tail DNA over control) in alga exposed to ≥ LC(50) concentration.

Effect of organic complexation on copper accumulation and toxicity to the estuarine red macroalga Ceramium tenuicorne: a test of the free ion activity model.

Current water quality criteria (WQC) regulations on copper toxicity to biota are still based on total dissolved (<0.4 µm membrane filter) copper concentrations with a hardness modification for freshwaters. There are however ongoing efforts to incorporate metal speciation in WQC and toxicity regulations (such as the biotic ligand model-BLM) for copper and other metals. Here, we show that copper accumulation and growth inhibition of the Baltic macroalga Ceramium tenuicorne exposed to copper in artificial seawater at typical coastal and estuarine DOC concentrations (similar to 2-4 mg/L-C as fulvic acid) are better correlated to weakly complexed and total dissolved copper concentrations rather than the free copper concentration [Cu2+]. Our results using a combination of competitive ligand exchange-adsorptive cathodic stripping voltammetry (CLE-ACSV) measurements and model calculations (using visual MINTEQ incorporating the Stockholm Humic Model) show that copper accumulation in C. tenuicorne only correlates linearly well to [Cu2+] at relatively high [Cu2+] and in the absence of fulvic acid. Thus the FIAM fails to describe copper accumulation in C. tenuicorne at copper and DOC concentrations typical of most marine waters. These results seem to indicate that at ambient total dissolved copper concentration in coastal and estuarine waters, C. tenuicorne might be able to access a sizable fraction of organically complexed copper when free copper concentration to the cell membrane is diffusion limited.