Acute toxicity and biodegradation of endosulfan by the polychaeta Perinereis aibuhitensis in an indoor culture

The polychaete Perinereis aibuhitensis, a key species in estuarine ecosystems, can improve the culture condition of sediment. Endosulfan is an organochlorine pesticide used globally to control insects and mites; however, it is a source of pollution in aquaculture as a result of runoff or accidental release. In this study, we evaluated the toxicity of endosulfan to polychaeta and its ability to improve polluted sediment. Specifically, the effects of a series of endosulfan concentrations (0, 1.25, 2.5, 5, 10, 15, and 20 mg/L) were investigated, and the results indicated that the 24-h median lethal concentration (24-h LC₅₀) was 55.57 mg/L, while the 48-h median lethal concentration (48-h LC₅₀) was 15.56 mg/L, and the safe concentration was about 1.556 mg/L. In a 30-d exposure experiment, the animal specimen could decompose endosulfan effectively while improving endosulfan-polluted aquatic sediment.     

Acute Toxicity of Formalin to the Red-tide Dinoflageilate Prorocentrum minimum Schiller （Protozoa, Mastigophora）

The acute toxic levels of formalin to a marine red-tide dinoflagellate Prorocentrum minimum Schiller (Mastigophora, Dinoflagellida) were determined by linear regression analysis. The data obtained revealed that there is a close regression relationship between logarithmic concentrations of formalin and mortality probit scales of the organism when exposure times are 1, 2, 6, 12, 24 h. The median lethal concentrations (LC50 values) obtained from correlation analysis for these exposure times (with 95% confidence intervals), were 11.83, 6.76, 4.37, 4.27 and 3.98mgL^-1 respectively. A toxicity curve was obtained to connect the exposure time and LC50 value from the correlation between them. The results indicated that P. minimum could be killed or fixed by formalin in the concentration range of 4-12mgL^-1 within 1-24h.     

Tolerance of ciliated protozoan Paramecium bursaria （Protozoa,Ciliophora） to ammonia and nitrites

The tolerance to ammonia and nitrites in freshwater ciliate Paramecium bursaria was measured in a conventional open system. The ciliate was exposed to different concentrations of ammonia and nitrites for 2h and 12h in order to determine the lethal concentrations. Linear regression analysis revealed that the 2h-LC50 value for ammonia was 95.94 mg/L and for nitrite 27.35 mg/L using probit scale method (with 95% confidence intervals). There was a linear correlation between the mortality probit scale and logarithmic concentration of ammonia which fit by a regression equation y=7.32x-9.51 (R^2=0.98; y, mortality probit scale; x, logarithmic concentration of ammonia), by which 2 h-LC50 value for ammonia was found to be 95.50 mg/L. A linear correlation between mortality probit scales and logarithmic concentration of nitrite is also followed the regression equation y=2.86x 0.89 (R^2=0.95; y, mortality probit scale; x, logarithmic concentration of nitrite). The regression analysis of toxicity curves showed that the linear correlation between exposed time of ammonia-N LC50 valueand ammonia-N LC50 value followed the regression equation y=2 862.85e-0.08x (R^2=0.95; y, duration of exposure to LC50 value; x, LC50 value), and that between exposed time of nitrite-N LC50 value and nitrite-N LC50 value followed the regression equation y = 127.15e-0.13x (R^2=0.91; y, exposed time of LC50 value; x, LC50 value). The results demonstrate that the tolerance to ammonia in P. bursaria is considerably higher than that of the larvae or juveniles of some metozoa, e.g. cultured prawns and oysters. In addition, ciliates, as bacterial predators, are likely to play a positive role in maintaining and improving water quality in aquatic environments with high-level ammonium, such as sewage treatment systems.     

Acute toxicity of organochlorine insecticide endosulfan to the giant freshwater prawn Macrobrochium rosenbergii

Endosulfan, an organochlorine pesticide, is highly toxic and effective at controlling pests in agriculture, horticulture, and public health programs. In this study, static bioassays were used to evaluate the toxicity of endosulfan to freshwater prawns( Macrobrachium rosenbergii) of various lengths(1.5±0.03,4±0.08, and 7±0.06 cm). Additionally, the activities of peroxidase(POD), acid phosphatase(ACP),alkaline phosphatase, acetylcholinesterase(AChE), and Na + /K +-ATPase were analyzed to refl ect the effects of endosulfan exposure. The 96 h LC 50 of endosulfan for prawns 1.5, 4, and 7 cm long were 1.86, 4.53,and 6.09μg/L, respectively, improved tolerance to endosulfan with growth. The POD activities of test organisms exposed to low concentrations of endosulfan were inhibited, indicating the presence of oxygen damaged tissue. Moreover, a notable decrease in AChE activity was observed due to overstimulation of neurotransmission, which might result in abnormal behavior. The effect caused by endosulfan on phosphatase production in the hepatopancreas of prawns 1.5, 4, and 7 cm long was different because the ability of nonspecifi c immune regulation increased with growth. The 96 h LC 50 values obtained in this study could be used in the formulation of water-quality criteria in China. Moreover, the changes in enzymes activities of M. rosenbergii under stress of endosulfan could be applied in the establishment of early warning indicators for bio-safety.     

Laboratory study on the ecological impact of sophorolipid used for harmful algae elimination

We studied the role of sophorolipid in inhibiting harmful algae bloom (HAB). Different sophorolipid concentrations were tested on marine microalgae, zooplankton, fish, and bivalve (Mytilus edulis) in laboratory. The result shows that sophorolipid could inhibit the growth of algal species selectively. Among three algae species selected, Platymonas helgolandica var. tsingtaoensis was promoted with increasing sophorolipid concentration; Isochrysis galbana was inhibited seven days later in sophorolipid concentration below 40 mg/L; and Nitzschia closterium f. minutissima was inhibited obviously in only a high sophorolipid concentration over 20 mg/L. Therefore, sophorolipid in a low concentration at <20 mg/L could remove certain harmful algae species effectively without harming other non-harmful microalgae. For other animals, sophorolipid could inhibit the growth of ciliate Strombidium sp. by 50% at 20 mg/L sophorolipid concentration after 96 h. The concentration in 96-h LC₅₀ for Calanus sinicus, Neomysis awatschensis, Lateolabrax japonicus, and Paralichthys olivaceus was 15, 150, 60, and 110 mg/L, respectively. The 24 h LC₅₀ value for Artemia salina was 600 mg/L. The relative clearance rate of mussel Mytilus edulis decreased to 80%, 40%, and 20% of the control group after being exposed to 20, 50, and 100 mg/L sophorolipid for 24 h. Therefore, the toxicity for mitigation of harmful algae bloom at previously recommended concentration of 5–20 mg/L sophorolipid is low for most tested organisms in this reaserch.     

Toxic effects of zinc on four species of freshwater fish

The toxic effects of Zn²⁺ on Silver Carp (Hypophthalmichthyps molitrix C. et V.), Big Head Carp (Aristrichthys nobilis Richardson), Grass Carp (Ctenopharyngodon ilellus C. et V.), and Blunt Snout Bream (Megalobrama aimbly-cephala Yih) are studied. The test results are: (1) There are linear correlations between 24h LC₅₀ and 48h LC₅₀ of Zn²⁺ for Silver Carp fingerling and temperature. 24h LC₅₀, 48h LC₅₀ and 96h LC₅₀ of Zn²⁺ for fry of the four species are also determined; (2) There are logarithmic correlations between the growth rates of the fry and the concentrations of Zn²⁺ and between expansion of fish egg membranes after absorbing water and concentrations of Zn²⁺; (3) The tolerance of fry of the four species to Zn²⁺ is in the following order: Grass Carp>Silver Carp>Blunt Snout Bream>Big Head Carp; (4) The safe concentrations of Zn²⁺ are: Big Head Carp: 0.008 mg/L, Grass Carp: 0.046 mg/L, Blunt Snout Bream: 0.010 mg/L, Silver Carp: 0.012 mg/L, Silver Carp fingerling: 0.09 mg/L.     

Impact of brine acidification on hatchability,survival and reproduction of Artemia parthenogenetica and Artemia franciscana in salt ponds,Bohai Bay,China

We studied the effect of pH(pH 5, 6, 7 and 8) on the hatching percentage, survival and reproduction of Artemia strains in Bohai Bay salt ponds. Strains included parthenogenetic Artemia from Bohai Bay(BHB), Artemia franciscana from San Francisco Bay, and A. franciscana artifi cially produced in salt ponds in Vietnam. The latter was included as a potential inoculum for biological management of salt ponds. The hatching percentage of cysts after 24 h and the survival rate of the tested Artemia strains were signifi cantly reduced when exposed to a culture medium at pH 5 for 18 d( P 0.05). The tolerance of Artemia to 48 h acid exposure varied with developmental stage, increasing in the following order: juvenile, nauplii, pre-adult, with maximum tolerance in adults. All strains of Artemia tested could not reproduce at pH 5. At pH levels from pH 6–8, a higher pH generally resulted in a shorter brood interval and enhanced ovoviviparity. Hence, we suggest that brine acidifi cation has a negative impact on Artemia populations in the Bohai Bay saltworks. Inoculation of Artemia with either local parthenogenetic Artemia or exotic A. franciscana should be feasible at pH 7–8.     

Exploration of Antifouling Potential of the Brown Algae <Emphasis Type="Italic">Laminaria</Emphasis> ‘Sanhai’

Seaweeds are one of the largest producers of biomass in the marine environment. It has been well known that marine algae, especially brown algae was a rich source of biogenic compounds with antifouling potential that could be ideal alternatives of tributyltin (TBT). In this paper, antifouling potential of the brown algae Laminaria ‘sanhai’ was explored. Firstly, the dried alga was extracted and the antialgal and antilarval activities were investigated. The EC₅₀ and LC₅₀ values of crude extract of Laminaria ‘sanhai’ against diatom (Skeletonema costatum) and barnacle larval (Chthamalus challengeri) were 8.9 μg mL^?1 and 12.0 μg mL^?1 respectively. Then, guided by bioassay, the bioactive substances were isolated by liquid-liquid extraction. The antialgal and antilarval activities of isolated fraction were improved with the EC₅₀ value of 7.4 μg mL^?1 against S. costatum and LC₅₀ value of 9.7 μg mL^?1 against C. challengeri larvae. Identification by IR, Q-TOFMS and GC-MS of the isolated bioactive substances revealed the abundance of fatty acids. These fatty acids, most with 16, 18 or 20 carbon atoms, contained myristic, hexadecanoic, oleic, linolenic, arachidonic and eicosapentaenoic acids. The results indicated that both the crude extract and the isolated bioactive substances had high antialgal and antilarval activities with no highlighted cytotoxicity which made the brown algae Laminaria ‘sanhai’ a promising source of the environmentally friendly antifoulants.     

Antifouling potential of bacteria isolated from a marine biofilm

Marine microorganisms are a new source of natural antifouling compounds. In this study, two bacterial strains, Kytococcus sedentarius QDG-B506 and Bacillus cereus QDG-B509, were isolated from a marine biofilm and identified. The bacteria fermentation broth could exert inhibitory effects on the growth of Skeletonema costatum and barnacle larvae. A procedure was employed to extract and identify the antifouling compounds. Firstly, a toxicity test was conducted by graduated pH and liquid-liquid extraction to determine the optimal extraction conditions. The best extraction conditions were found to be pH 2 and 100% petroleum ether. The EC50 value of the crude extract of K. sedentarius against the test microalgae was 236.7 ± 14.08 μg mL-1, and that of B. cereus was 290.6 ± 27.11 μg mL-1. Secondly, HLB SPE columns were used to purify the two crude extracts. After purification, the antifouling activities of the two extracts significantly increased: the EC50 of the K. sedentarius extract against the test microalgae was 86.4 ± 3.71 μg mL-1, and that of B. cereus was 92.6 ± 1.47 μg mL-1. These results suggest that the metabolites produced by the two bacterial strains are with high antifouling activities and they should be fatty acid compounds. Lastly, GC-MS was used for the structural elucidation of the compounds. The results show that the antifouling compounds produced by the two bacterial strains are myristic, palmitic and octadecanoic acids.     

Biochemical response of the mussel Mytilus coruscus (Mytiloida: Mytilidae) exposed to in vivo sub-lethal copper concentrations

Many aquatic organisms are negatively affected by exposure to high copper concentrations.We investigated the biochemical response of the mussel Mytilus coruscus(Mytiloida:Mytilidae) to copper exposure.In vivo bioassays using M.coruscus and different copper concentrations were conducted.The activity of six biomarkers,namely superoxide dismutase(SOD),catalase(CAT),acid phosphatase(ACP),alkaline phosphatase(AKP),glutamic-oxaloacetic transaminase(GOT) and glutamic-pyruvic transaminase(GPT) were measured.Survival rates decreased with increased copper concentrations and exposure times.The LC50 values at 48,72,and 96 h exposure were 0.48,0.37,and 0.32 mg/L,respectively.Within digestive glands,CAT activity increased with increasing Cu concentrations.The activity of AKP showed no significant change,while the remaining four enzymes showed decreasing activity with increasing Cu concentrations.Within the gills,AKP activity increased when the Cu concentration was 0.05 mg/L,but showed no significant changes at higher concentrations.Activity of CAT and ACP within gills tended to decrease with increasing Cu concentration.The activity of SOD and GPT decreased at an exposure concentration of 0.2 mg/L.GOT activity within gills decreased at 0.1 mg/L and increased at an exposure concentration of 0.2 mg/L.Within the adductor muscle,AKP activity increased at 0.05 mg/L but did not change at higher exposure concentrations.ACP activity within adductor muscle tissue showed no change,while activities of CAT,GOT and GPT decreased with increasing Cu concentrations.SOD activity within the adductor muscle tissue significantly decreased at the 0.02,0.05 and 0.2 mg/L exposure concentrations.Our results show tissue specific differences for the six biomarkers in for M.coruscus.Our findings provide the basis for the establishment of reference activity levels against which biomarker changes can be estimated,and are essential preliminary steps in development of in vivo bioassays.     

Antialgal and antilarval activities of bioactive compounds extracted from the marine dinoflagellate <Emphasis Type="Italic">Amphidinium carterae</Emphasis>

With the global ban on the application of organotin-based marine coatings by the International Maritime Organization, the development of environmentally friendly, low-toxic and nontoxic antifouling compounds for marine industries has become an urgent need. Marine microorganisms have been considered as a potential source of natural antifoulants. In this study, the antifouling potential of marine dinoflagellate Amphidinium carterae, the toxic and red-tide microalgae, was investigated. We performed a series of operations to extract the bioactive substances from Amphidinium carterae and tested their antialgal and antilarval activities. The crude extract of Amphidinium carterae showed significant antialgal activity and the EC₅₀ value against Skeletonema costatum was 55.4 μg mL^?1. After purification, the isolated bioactive substances (the organic extract C) exhibited much higher antialgal and antilarval activities with EC₅₀ of 12.9 μg mL^?1 against Skeletonema costatum and LC₅₀ of 15.1 μg mL^?1 against Amphibalanus amphitrite larvae. Subsequently, IR, Q-TOFMS, and GC-MS were utilized for the structural elucidation of the bioactive compounds, and a series of unsaturated and saturated 16- to 22-carbon fatty acids were detected. The data suggested the bioactive compounds isolated from Amphidinium carterae exhibited a significant inhibiting effect against the diatom Skeletonema costatum and Amphibalanus amphitrite larvae, and could be substitutes for persistent, toxic antifouling compounds.     

Acute toxicities of potassium permanganate, formalin, and Lugol’s iodine solution to a marine ciliate, Pleuronema coronatum (ciliophora, scuticociliatida)

Acute toxicities of potassium permanganate, formalin, and Lugol’s iodine solution to a commonly occurred marine ciliate Pleuronema coronatum (Ciliophora, Scuticociliatida) were measured. Linear regression analysis of the results highlighted the close relationships between doses of the medicines and mortalities of the organisms, thus providing a capability to predict toxicity effects from the dose. Toxic effects of the medicines on the ciliates were described in the present paper, and the median lethal concentrations (LC₅₀ values) were given. Results of measurements indicated that 2 h-LC₅₀ and 12 h-LC₅₀ values of formalin on P. coronatum were 59.00×10⁻⁶ and 43.57×10⁻⁶, while those of Lugol’s solutions were 90.13 and 67.84×10⁻⁶ respectively. The tolerance of P. coronatum to formalin is apparently lower than that to Lugol’s iodine solution and potassium permanganate is a suitable medicine to kill ciliates in short time.     

An approach to determination of phenolic compounds in seawater using SPME-GC-MS based on SWCNTs coating

Phenolic compounds have become one kind of the important pollutants of the marine environment. Single-walled Carbon nanotubes, as one-dimensional nano materials, have light weight and perfect hexagonal structure of connections, with many unusual mechanical, chemical and electrical properties. In recent years, with the research of carbon nanotubes and other nano materials, the application prospect is also constantly discussed. In this paper, homemade single-walled carbon nanotubes(SWCNTs) coating was used for establishing an analytical approach to the determination of five kinds of phenolic compounds in seawater using SPME-GC-MS. Optimal conditions: After saturation was conducted with Na Cl, and p H was adjusted to 2.0 with H_2SO_4, the extract was immersed in a water bath at 40 for GC℃-MS determination through 40-min agitating extraction at 500 rmin~(-1) and 3-min desorption at 280℃. The liniearities ranged between 0.01-100 μg L~(-1), and the determination limits ranged between 1.5-10 ngL~(-1). The relative standard deviation(RSD, n = 5) was less than 6.5%. For the phenolic compounds obtained from the spiked recovery test for actual seawater samples, the rates of recovery were 87.5%-101.7%, and the RSDs were less than 8.8%, which met the requirements of determination. Due to its simplicity, high efficiency and low consumption, this approach is suitable for the analysis of trace amounts of phenolic compounds in marine waters.     

The effect of salinity on Fucus ceranoides (Ochrophyta,Phaeophyceae) in the Mondego River (Portugal)

Algae(and their extracts) are increasingly important for pharmaceutical applications due to the diversity of useful compounds they contain. The genus Fucus contains one of the most studied species,Fucus vesiculosus. The species F. ceranoides differs from the others of the genus by presenting longitudinal air-vesicles and a capacity to survive at low salinities. It is an alga that inhabits the Mondego River estuary(Portugal), at the southern limit of its distribution, and can serve as a role model to understand the effect of a salt gradient on the production of bioactive compounds. We assessed the phenolic content and antioxidant activity of different F. ceranoides extracts(e.g. methanolic, aqueous and polysaccharide) prepared from samples harvested from two different zones to evaluate if the adaptation of F. ceranoides to different salinity levels influenced its chemical composition. The antioxidant activity of the extracts was determined using 1,2-diphenyl-picrylhydrazyl(DPPH) and 2.2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)(ABTS)radicals. These assays demonstrated that the methanolic extract of lyophilized F. ceranoides that grew at low salinities was the most bioactive, i.e. DPPH(IC_(50)=50.39 μg/mL) and ABTS(TEAC=2.42). The total phenolic content(Folin-Ciocalteu method) and the methanolic extract of the lyophilized F. ceranoides collected from a low salinity habitat exhibited the highest phenolic content(PGE=49.48 μg/mg of lyophilized extract)amongst those sampled. Thin layer chromatography(TLC) and Fourier transform infrared spectroscopy(FTIR) were used for the identification of compounds in the extracts. This characterization allowed confirmation that the various extracts contained almost the same compounds but with notable quantitative differences. Based on these results, we conclude that there were differences in the quantity of the compounds due to the effect of salinity. The drying methods used were also found to have influenced the quality of the extracted compounds.     

CYTOTOXICITY AND GENOTOXICITY OF POLYETHYLENIMINE AND NICKEL CHLORIDE IN RED SEA BREAM（Pagrosomus major）FIN CELL LINE RSBF

A continuous marine fish cell line RSBF(i.e.Red Sea Bream Fin)was utilized to screen the cytotoxicity and genotoxicity of polyethylenimine(PEI)and nickel cholride(NiCl2)in this study on the deleterious effects of aquatic genotoxins on fish.At the 0.01 to 1μgml concentration tested,PEI had acute toxicity to the treated RSBF cells(IC50=1.12,0.92,0.88 and 0.64μg/ml PEI for time 0 h,24 h,48 h and 72 h after treatment,respectively)and markedly inhibited their proliferation in a dose-dependent manner.At the 0.001 to 5 μmol/L concentration tested,NiCl2 posed no acute toxicity but significantly stimulated their growth(107?214?of control).Random amplified polymorphic DNA(RAPD)technique was used to detect the genotoxic effects of PEI and NiCl2 by comparing the RAPD banding patterns of the control and treated cells.RAPD analysis indicated that at the concentrations tested,PEI was more genotoxic than NiCl2 to RSBF cells;that there was a slight dose-dependent response in the genotoxic effect of PEI bue not NiCl2;and that RAPD technique might provide a sensitive,non-specif-ic gentoxic endpoint.And the potent cytotoxicity and genotoxicity of PEI on fish cells showed that we should be cautious in utilizing it as gene vector in fish gene transfer and human gene therapy.     

Ontogenetic changes in digestive enzyme activities and the amino acid profile of starry flounder <Emphasis Type="Italic">Platichthys stellatus</Emphasis>

Ontogenetic changes in digestive enzyme activities and the amino acid (AA) profile of starry flounder, Platichthys stellatus, were investigated and limiting amino acids were estimated compared with the essential AA profile between larvae and live food to clarify starry flounder larval nutritional requirements. Larvae were collected at the egg stage and 0, 2, 4, 7, 12, 17, 24 days after hatching (DAH) for analysis. Larvae grew from 1.91 mm at hatching to 12.13 mm at 24 DAH. Trypsin and chymotrypsin activities changed slightly by 4 DAH and then increased significantly 4 DAH. Pepsin activity increased sharply beginning 17 DAH. Lipase activity increased significantly 4 DAH and increased progressively with larval growth. Amylase activity was also detected in newly hatched larvae and increased 7 DAH followed by a gradual decrease. High free amino acid (FAA) content was detected in starry flounder eggs (110.72 mg/g dry weight). Total FAA content dropped to 43.29 mg/g in 4-DAH larvae and then decreased gradually to 13.74 mg/g in 24-DAH larvae. Most FAAs (except lysine and methionine) decreased >50% in 4-DAH larvae compared with those in eggs and then decreased to the lowest values in 24-DAH larvae. Changes in the protein amino acid (PAA) profile were much milder than those observed for FAAs. Most PAAs increased gradually during larval development, except lysine and phenylalanine. The percentages of free threonine, valine, isoleucine, and leucine decreased until the end of the trial, whereas the protein forms of these four AAs followed the opposite trend. A comparison of the essential AA composition of live food (rotifers, Artemia nauplii, and Artemia metanauplii) and larvae suggested that methionine was potentially the first limiting AA. These results may help develop starry flounder larviculture methods by solving the AA imbalance in live food. Moreover, the increased digestive enzyme activities indicate the possibility of introducing artificial compound feed.     

Significance of different microalgal species for growth of moon jellyfish ephyrae, <Emphasis Type="Italic">Aurelia</Emphasis> sp.1

The scyphozoan Aurelia aurita (Linnaeus) sp. l., is a cosmopolitan species-complex which blooms seasonally in a variety of coastal and shelf sea environments around the world. The effects of different microalgal species on the growth of newly-released Aurelia sp.1 ephyrae were studied under laboratory conditions. We fed ephyrae with four different microalgal species (diatom, autotrophic dinoflagellate, heterotrophic dinoflagellate, and chlorophyta) plus Artemia nauplii for 12-24 d at 18°C. Results showed that the growth rate diverged significantly for Artemia nauplii compared to other food types. In addition, there was no significant variation between the growth rates for Skeletonema costatum and Prorocentrum donghaiense, and no significant variation was found in the growth rates for N. scintillans and P. subcordiformis. Artemia nauplii could support the energy requirement for the newly-released ephyrae to develop to meduase, and the ephyrae with Artemia nauplii showed a significant average growth rate of 25.85% d^?1. Newly-released ephyrae could grow slightly with some species of microalgae in the earliest development stage. Chain diatom Skeletonema costatum and autotrophic dinoflagellate Prorocentrum donghaiense, could not support the growth of the ephyrae, while heterotrophic dinoflagellate Noctiluca scintillans and chlorophyta Platymonas subcordiformis could support the growth of the ephyrae. However, none of the ephyrae fed with the tested phytoplankton could mature to medusae.     

Optimization of the microwave-assisted extraction of phlorotannins from Saccharina japonica Aresch and evaluation of the inhibitory effects of phlorotannin-containing extracts on HepG2 cancer cells

The use of a microwave-assisted extraction(MAE) method for the extraction of phlorotannins from Saccharina japonica Aresch(S.japonica) has been evaluated with particular emphasis on the influential parameters,including the ethanol concentration,solid/liquid ratio,extraction time,extraction temperature,and microwave power.The MAE procedure was optimized using single-factor design and orthogonal array design(OAD).The content of total phlorotannins in S.japonica was determined using a Folin-Ciocalteu(FC) assay.A maximum total phlorotannin content of 0.644 mg of phloroglucinol equivalent per gram of dry weight plant(mg PGE/g DW) was obtained using the optimized model,which included an ethanol concentration of 55%,solid/liquid ratio of 1:8,extraction time of 25 min,irradiation power of 400 W,and temperature of 60°C.Under similar conditions,the application of a conventional extraction method led to a lower phlorotannin yield of 0.585 mg PGE/g WD.These results demonstrated that the MAE approach provided better results for the extraction of phlorotannins from S.japonica and was a promising technique for the extraction of phenolic compounds from S.japonica and other materials.In addition,screening tests for the inhibitory activity showed that the phlorotannin-containing extracts significantly inhibited the growth of human hepatocellular carcinoma cells(HepG2) by inducing their apoptosis.The morphological changes that occurred during cell apoptosis were characterized using Hoechst33258 staining.     

Diel variation in metabolism and ammonia excretion of <Emphasis Type="Italic">Marphysa sanguinea</Emphasis> (Polychaeta: Eunicidae)

Polychaetes provide an excellent food resource for fish and represent the dominant zoobenthos in marine ecosystems. Diel variation in the rates of metabolism and ammonia-N excretion of Marphysa sanguinea were studied. The worms were grouped according to their wet body weight into small (S; 1.24±0.06 g), medium (M; 4.00±0.30 g), and large (L; 8.54±1.08 g) categories. Their weight-specific metabolic rates, based on aerobic respiration (R), were measured at 16°C (±0.2°C) and classed as either routine (R _R) or standard (R _S) rates. Both respiration types decreased with increasing body weight. Respiration was described by R = a W ^b, where b was -0.400 9 and -0.532 0 for R _R and R _S, respectively. Diurnal changes in R _S for each group was relatively flat, with a slightly increasing trend with time, but was relatively stable as a whole. R _R of the diurnal variation of worms was higher than R S, but both had similar overall trends. The peak values of specific dynamic action (SDA) (R _SDA) in the S, M, and L groups were 2.704, 1.149, and 0.682 mg/(g?h), respectively. The durations of SDA were 13, 6, and 6 h, respectively and the energy expenditures of SDA were 377.98, 117.34, and 74.94 J/g, respectively. These data indicate that the metabolic rates were higher in smaller individuals, which is advantageous for their rapid growth.     

Microanalysis and preliminary pharmacokinetic studies of a sulfated polysaccharide from <Emphasis Type="Italic">Laminaria japonica</Emphasis>

A rapid, sensitive and reproducible high performance liquid chromatography (HPLC) method with post-column fluorescence derivatization has been developed to determine the amount of low-molecular-weight sulfated polysaccharide (GFS) in vivo. The metabolism of GFS has been shown to fit a two component model following its administration by intravenous injection, and its pharmacokinetic parameters were determined to be as follows: half-time of distribution phase (t _1/2α)=11.24±2.93 min, half-time of elimination phase (t _1/2β)=98.20±25.78 min, maximum concentration (C _max)=110.53 μg/mL and peak time (T _max)=5 min. The pharmacokinetic behavior of GFS was also investigated following intragastric administration. However, the concentration of GFS found in serum was too low for detection, and GFS could only be detected for up to 2 h after intragastric administration (200 mg/kg body weight). Thus, the bioavailability of GFS was low following intragastric administration because of the metabolism of GFS. In conclusion, HPLC with postcolumn derivatization could be used for quantitative microanalysis and pharmacokinetic studies to determine the presence of polysaccharides in the serum following intravenous injection.