The Effects of Inorganic Phosphates on Trochophore Larvae of the Oyster,Crassostrea virginica

Summary

Trochophore larvae were exposed to monosodium orthophosphate (MSOP), sodium hexametaphosphate (SHMP), sodium tripolyphosphate (STPP), and tetrasodium pyrophosphate (TSPP) at concentrations of 15 ppm and 150 ppm phosphate for 24, 48, and 72 hr. All four compounds caused morphological developmental changes, increased mortality, and inhibition of shell growth at both 15 ppm and 150 ppm. The rank order of effectiveness of the compounds was determined from percentage abnormal larvae, percentage survival, and from shell measurements. The order was MSOP>SHMP>STPP≥TSPP for abnormality and mortality and TSPP>STPP>MSOP>SHMP for shell growth inhibition. The relative effect of the individual compounds in causing mortality varied with the time of exposure. Orthophosphate retarded growth rates of the two valves to different degrees.     

6种土壤微生物提取剂的比较

比较了6种土壤微生物提取剂（6-偏磷酸钠溶液、焦磷酸钠溶液、磷酸盐缓;中液、林格溶液、NaCl溶液和水）对土壤细菌、真菌和放线菌数量的影响。结果表明：0．1％的6-偏磷酸钠（HMP）和焦磷酸钠（PYS）溶液（w／v）提取的细菌、真菌、放线菌数量最多,磷酸缓冲液（pH值7．2）对土壤真菌提取效率与焦磷酸钠和6-偏磷酸钠溶液相似,而对细菌和放线菌的提取效率则低于焦磷酸钠和6-偏磷酸钠溶液,其余3种提取剂的提取效率相对较低。     

Preparation of phosphorus and carbohydrate microcapsules for manipulating dietary C : P ratio for aquatic suspension-feeders

SUMMARY 1. Dietary phosphorus can be limiting for aquatic animals such as suspension-feeders. However, our understanding has been limited by the difficulty of manipulating dietary P without altering other aspects of food quality. We microencapsulated various forms of bioavailable P with carbohydrate to manipulate dietary C : P ratio for suspension-feeders.
2. Calcium phosphate, sodium hexametaphosphate, sodium tripolyphosphate and tetrasodium pyrophosphate were each mixed with a concentrated solution of a carbohydrate base (either maltodextrin or potato starch) and microencapsulated using an interfacial polymerisation technique. Each of the 10 types of capsules produced had a particle size ideal for suspension-feeders (3–10 μm).
3. Leakage rates were low (<12% of capsule weight per day). Relative enzymatic breakdown in vitro by carbohydrases (amylase or cellulase) was similar among the 10 capsule types and was always at least 15 times the comparable leakage rate.
4. Release of dissolved P from enzyme-treated capsules varied depending on capsule P content. Liberation of P from capsules prepared from 20% w/w sodium hexametaphosphate in maltodextrin (molar C : P = 1.8) was three times greater than all other types, and this combination appears most suitable as a dietary supplement for zooplankton.
5. Although P content and capsule integrity were greatly influenced by choice of carbohydrate, choice of P compound, and the mixing ratio of the two, P-rich artificial microparticles can be produced that have low leakiness, high digestibility, and a physical size suitable for aquatic suspension-feeders. Therefore, microcapsules represent promising tools for manipulating dietary C : P for suspension feeders.     

The location of the polyphosphate-binding sites on cytochrome c measured by NMR paramagnetic difference spectroscopy.

Analyses of unimolecular electron self-exchange reactions provide a comparatively simple and direct approach to understanding biological electron transfer. Such studies are currently limited by a lack of well characterised aggregating systems. In the presence of sodium hexametaphosphate, cytochrome c forms stable protein aggregates as a result of binding hexametaphosphate at a single site on its surface (preceding paper in this issue of the journal). Here we report the location of the principal polyphosphate binding site on the surface of cytochrome c for both hexametaphosphate and a second polyphosphate, tripolyphosphate determined using 1H-NMR spectroscopy in conjunction with the relaxation probe potassium hexacyanochromium(III). Addition of either hexametaphosphate or tripolyphosphate to ferricytochrome c in the presence of the relaxation probe causes a decrease in intensity of several resonances in the paramagnetic difference spectrum, including Phe82 ortho/meta, Ile85 delta methyl and Ile9 gamma methyl. Together these effects put the site of polyphosphate binding close to lysines 13, 86, and 87. Additionally the effect of sodium tripolyphosphate and sodium trimetaphosphate on cytochrome c aggregation is described. The potential role of this site in anion-induced cytochrome c aggregation is discussed.     

Factors affecting sensitivity of Staphylococcus aureus 196E to polyphosphates.

The effect of polyphosphates (eight compounds) on growth of Staphylococcus aureus 196E in brain heart infusion broth was studied. The organism was sensitive (in decreasing order) to chain polyphosphates with 21, 3, 13, and 15 PO4 groups, and bactericidal effects were observed with 0.5% of these compounds. No inhibition was effected by PPi or a metaphosphate. The inhibitory effects were pH dependent, and bacterial sensitivity was highest at pH greater than 7.4. Initial populations affected the number of survivors. No growth was observed after 24 h at 35 degrees C when the initial cell population was less than 10(4) CFU/ml, and a 100- to 1,000-fold decline in cell numbers occurred when initial populations were higher than 10(4) CFU/ml. Sodium tripolyphosphate produced less inhibition after heat sterilization (15 min, 121 degrees C) than after filter sterilization, whereas sodium hexametaphosphate (n = 21) retained most of its antimicrobial activity after heat sterilization. Supplementation of broth with Mg2+ was effective in overcoming inhibition by 0.5% sodium tripolyphosphate, and an addition of 0.25 to 1.0 mM cation restored most of the growth. Inhibition was partially eliminated by Ca2+ and Fe2+, but not by Zn2+ or Mn2+.     

Factors affecting sensitivity of Staphylococcus aureus 196E to polyphosphates

The effect of polyphosphates (eight compounds) on growth of Staphylococcus aureus 196E in brain heart infusion broth was studied. The organism was sensitive (in decreasing order) to chain polyphosphates with 21, 3, 13, and 15 PO4 groups, and bactericidal effects were observed with 0.5% of these compounds. No inhibition was effected by PPi or a metaphosphate. The inhibitory effects were pH dependent, and bacterial sensitivity was highest at pH greater than 7.4. Initial populations affected the number of survivors. No growth was observed after 24 h at 35 degrees C when the initial cell population was less than 10(4) CFU/ml, and a 100- to 1,000-fold decline in cell numbers occurred when initial populations were higher than 10(4) CFU/ml. Sodium tripolyphosphate produced less inhibition after heat sterilization (15 min, 121 degrees C) than after filter sterilization, whereas sodium hexametaphosphate (n = 21) retained most of its antimicrobial activity after heat sterilization. Supplementation of broth with Mg2+ was effective in overcoming inhibition by 0.5% sodium tripolyphosphate, and an addition of 0.25 to 1.0 mM cation restored most of the growth. Inhibition was partially eliminated by Ca2+ and Fe2+, but not by Zn2+ or Mn2+.     

Synthesis of fluorescent organic phosphates and their equilibrium binding to bovine oxyhemoglobin.

Fluorescent organic phosphates, beta-naphthyl diphosphate, beta-naphthyl triphosphate, and beta-naphthyl tetraphosphate, were synthesized from beta-naphthyl monophosphate using Pi and N,N'-dicyclohexylcarbodiimide. These organic phosphates were interacted with bovine oxyhemoglobin, all in no buffer, 0.1 M NaCl, at 25 degrees and in the pH range 5.5-7.0. Equilibrium binding parameters were determined by measuring the fluorescence quenching upon their interaction. It is indicated that bovine oxyhemoglobin has more than one binding site, one of which is very strong. The strength of binding to the stronger site is in the order beta-naphthyl tetraphosphate greater than beta-naphthyl triphosphate greater than beta-naphthyl diphosphate. The logarithms of association constants of these phosphates depend linearly on the net charges of these phosphates at any pH. The results were accounted for by electrostatic effects using a simple charge model. In that model, the average positive net charges in oxyhemoglobin involved in the binding of beta-naphthyl phosphate are shown as a function of pH. It is shown that the binding of these fluorescent organic phosphates is prevented reversibly by the excess addition of nonfluorescent organic and iorganic phosphates, inositol hexaphosphate, tripolyphosphate, and pyrophosphate. Assuming competitive binding in a single strong site, the association constants of these nonfluorescent phosphates were also determined by measuring the recovery of the fluorescence intensity upon the release of fluorescent phosphates. At pH 6.18, the association constant of pyrophosphate is lower than that of tripolyphosphate by one order.     

The cis-trans isomerization of aromatic heptaenes catalyzed by sulfur and phosphorus compounds]

It was shown that sulfur and phosphorus compounds (sodium thiosulfate, sodium tripolyphosphate, sodium hexametaphosphate, monosodium phosphate) catalyze cis-trans isomerization of aromatic heptaens. Preparative method of levorin isomerezation at the presence of sodium thiosulfate was elaborated. The isolated product was a fully trans-isomer.     

Pyrophosphate and tripolyphosphate affect firefly luciferase luminescence because they act as substrates and not as allosteric effectors

The activating and stabilizing effects of inorganic pyrophosphate, tripolyphosphate and nucleoside triphosphates on firefly luciferase bioluminescence were studied. The results obtained show that those effects are a consequence of the luciferase-catalyzed splitting of dehydroluciferyl-adenylate, a powerful inhibitor formed as a side product in the course of the bioluminescence reaction. Inorganic pyrophosphate, tripolyphosphate, CTP and UTP antagonize the inhibitory effect of dehydroluciferyl-adenylate because they react with it giving rise to products that are, at least, less powerful inhibitors. Moreover, we demonstrate that the antagonizing effects depended on the rate of the splitting reactions being higher in the cases of inorganic pyrophosphate and tripolyphosphate and lower in the cases of CTP and UTP. In the case of inorganic pyrophosphate, the correlation between the rate of dehydroluciferyl-adenylate pyrophosphorolysis and the activating effect on bioluminescence only occurs for low concentrations because inorganic pyrophosphate is, simultaneously, an inhibitor of the bioluminescence reaction. Our results demonstrate that previous reports concerning the activating effects of several nucleotides (including some that do not react with dehydroluciferyl-adenylate) on bioluminescence were caused by the presence of inorganic pyrophosphate contamination in the preparations used.     

In vitro antifungal activity of the tea tree (Melaleuca alternifolia) essential oil and its major components against plant pathogens

AIMS: The aim of this study was to examine the effect of Melaleuca alternifolia essential oil (TTO) and its principal components on four cereal-pathogenic fungi. METHODS AND RESULTS: The antimycotic properties of TTO and of terpinen-4-ol, gamma-terpinen and 1,8-cineole (eucalyptol) were evaluated in vitro on Fusarium graminearum, Fusarium culmorum and Pyrenophora graminea. Moreover, barley leaves infected with Blumeria graminis were treated with whole TTO. All the tested fungi were susceptible to TTO and its components. CONCLUSIONS: TTO exerted a wide spectrum of antimycotic activity. Single TTO purified components were more active than the whole oil in reducing in vitro growth of fungal mycelium and, among the tested compounds, terpinen-4-ol was the most effective. SIGNIFICANCE AND IMPACT OF THE STUDY: TTO and its components can be considered potential alternative natural fungicides.     

<Emphasis Type="Italic">Penicillium verrucosum</Emphasis> as promising candidate for bioremediation of environment contaminated with synthetic detergent at high concentration

The aim of this study was to investigate the effect of synthetic detergent Merix (Henkel, Kru?evac, Serbia), and its particular components—ethoxylated oleyl-cetyl alcohol and sodium tripolyphosphate on the growth and metabolic activity of Penicillium verrucosum. During 19 days of fungal cultivation in Czapek-Dox liquid medium supplemented with or without 0.5% pollutants, the following parameters were observed: pH, the total biomass dry weight, the quantity of free and total organic acids and proteolytic activity. The detergent caused a slight stimulatory (2.41%) effect whereas sodium tripolyphosphate and ethoxylated oleyl-cetyl alcohol provided a slight inhibitory action (0.59 and 2.75%, respectively) on the fungal biomass. The pollutants decreased pH values of the media and the quantity of free organic acids. In contrast, they enhanced the quantity of total organic acids. Proteolytic activity remained nearly unchanged (95.8%) in the presence of detergent and reduced to 80.1% in sodium tripolyphosphate-supplemented medium. In contrast, the enzyme activity sharply increased (260.8%) with ethoxylated oleyl-cetyl alcohol. The obtained results indicate the potential of P. verrucosum in bioremediation of environment contaminated with synthetic detergent taken in high concentration.     

Influence of Mg2+ and inorganic phosphates on the assembly of tubulin depleted of its exchangeable guanine nucleotide.

After the removal of the exchangeable guanine nucleotides by chromatography on phenyl-Sepharose [Hanssens, I., Baert, J., and Van Cauwelaert, F. (1990) Biochemistry 29, 5160-5165] tubulin polymerizations with GTP, GDP, tripolyphosphate, pyrophosphate or orthophosphate as possible stimulants are compared. It is demonstrated that, besides GTP and pyrophosphate, also tripolyphosphate stimulates the assembly into microtubules at high concentrations (4.65 mM) of Mg2+. The influence of Mg2+ is more pronounced in combination with pyrophosphate and tripolyphosphate than with GTP. The microtubules assembled in combination with Mg2+ and tripolyphosphate or pyrophosphate are short, suggesting that especially the nucleation step of microtubule assembly is favoured.     

Formation of pyrophosphate,tripolyphosphate, and phosphorylimidazole with the thioester,N, S-diacetylcysteamine,as the condensing agent

Summary Reaction of 0.20M orthophosphate with 0.20M N,S-diacetylcysteamine in 0.40M imidazole at pH 7.0 or 8.0 under drying conditions at 50°C for 6 days yields pyrophosphate and tripolyphosphate in the presence and absence of 0.10M divalent metal ion. The efficiency of utilization of N,S-diacetylcysteamine in the formation of pyrophosphate linkages ranges from 3 – 8% under the above conditions. The thioester, N,S-diacetylcysteamine, and imidazole are required for phosphoanhydride formation.Reaction of 0.40M orthophosphate with 0.20M N, S-diacetylcysteamine in 0.40M imidazole at ambient temperature for 6 days yields phosphorylimidazole in the absence or presence of 0.05M MgCl₂. Phosphorylimidazole and pyrophosphate are formed in the presence of 0.05M CaCl₂; pyrophosphate and tripolyphosphate are formed with 0.15M CaCl₂. The efficiency of utilization of N,S-diacetylcysteamine in the formation of pyrophosphate linkages is roughly 7% at 6 days of reaction with 0.15M CaCl₂. The thioester, N,S-diacetylcysteamine and imidazole are required for the formation of phosphoanhydrides. The significance of these reactions to molecular evolution is discussed.Abbreviations P₁ orthophosphate - P₂ pyrophosphate - P₃ tripolyphosphate - ImP phosphorylimidazole - Ac-Csa(Ac) N, S-diacetylcysteamine - Im imidazole     

卵孢金孢藻对不同磷化合物的利用及生长响应

为了解水华蓝藻卵孢金孢藻对水体不同磷化合物的利用能力,试验以磷酸氢二钾为对照,通过室内模拟的方法探究不同磷基质条件下卵孢金孢藻的生长响应.结果表明: 卵孢金孢藻能直接利用三聚磷酸钠和十水焦磷酸钠,且对三聚磷酸钠有极高的利用率.15天后,三聚磷酸钠处理组的藻细胞生物量和叶绿素a浓度最高,分别达到(426.96±47.42) mg·L^-1和(1852.34±116.60) μg·L^-1.2-氨基乙基膦酸和五水β-甘油磷酸钠处理组生物量与对照无明显差异,可溶性无机磷变化特征响应了碱性磷酸酶活性大小,表明卵孢金孢藻能通过酶水解利用这两种有机磷.而整个培养过程中,草甘膦处理组可溶性无机磷浓度趋近于0 mg·L^-1,藻细胞生物量、比生长速率、叶绿素a浓度及光合活力均显著低于对照,表明草甘膦不仅不利于藻细胞摄取磷,还对其生长产生抑制作用.本研究结果为了解卵孢金孢藻向不同水生态系统的扩散机理提供了新的思路,对我国新型蓝藻水华的防治具有一定的理论参考价值.     

Influence of the interaction of temperature and water activity on the production of ochratoxin A and the growth of Aspergillus niger, Aspergillus carbonarius and Aspergillus ochraceus on coffee-based culture medium

In the present study, the effect of temperature and water activity on fungal growth and ochratoxin production on coffee-based medium was assessed. Optimal growth of three Aspergillus strains was observed in the same ecological conditions, namely 30 degrees C and 0.99 water activity. Maximal daily growth is 11.2, 6.92, and 7.22 mm/day for Aspergillus niger, Aspergillus carbonarius, and Aspergillus ochraceus, respectively. However, ecological conditions for optimal ochratoxin production vary according to the toxinogenic strain, with water activity as a limiting factor. Such an ochratoxin A production is inhibited at 42 degrees C and 0.75 water activity. Correspondence between laboratory tested water activity and that measured on a sun-dried ripe cherry batch shows that the first 5 days of drying are critical for fungal growth and ochratoxin A production. Accordingly, artificial drying of cherries at temperatures above 42 degrees C will impede not only fungal growth but also contamination with ochratoxin A.     

Effects of potassium sorbate and other antibotulinal agents on germination and outgrowth of Clostridium botulinum type E spores in microcultures.

The effects of potassium sorbate, sodium hypophosphite, sodium tripolyphosphate, sodium nitrite, and linoleic acid on the germination and outgrowth of Clostridium botulinum type E spores were studied in microcultures. At pH 5.8 to 6.0 in liver veal agar, the germination rate was decreased to nearly zero with 1.0, 1.5, or 2.0% sorbate. At pH 7.0 t 7.2, these levels of sorbate afforded germination and outgrowth of abnormally shaped cells that were defective in cell division. At the high pH range, 0.5 or 1.0% hypophosphite had effects similar to those of sorbate. The use of 0.05% sodium nitrite with sorbate enhanced the lysis of outgrowing cells at pH 7.2 or lower. Emergence and elongation were inhibited by 0.05% linoleic acid with or without 1.0% sorbate at pH 7.0 to 7.2. The addition of 0.5% tripolyphosphate to media containing 1.5% sorbate at pH 7.1 prevented normal cell growth to an extent greater than with sorbate alone.     

Chlorella virus RNA triphosphatase. Mutational analysis and mechanism of inhibition by tripolyphosphate

Chlorella virus RNA triphosphatase (cvRtp1) is the smallest member of a family of metal-dependent phosphohydrolases that includes the RNA triphosphatases of fungi, protozoa, poxviruses, and baculoviruses. The primary structure of cvRtp1 is more similar to that of the yeast RNA triphosphatase Cet1 than it is to the RNA triphosphatases of other DNA viruses. To evaluate the higher order structural similarities between cvRtp1 and the fungal enzymes, we performed an alanine scan of individual residues of cvRtp1 that were predicted, on the basis of the crystal structure of Cet1, to be located at or near the active site. Twelve residues (Glu(24), Glu(26), Asp(64), Arg(76), Lys(90), Glu(112), Arg(127), Lys(129), Arg(131), Asp(142), Glu(163), and Glu(165)) were deemed essential for catalysis by cvRtp1, insofar as their replacement by alanine reduced phosphohydrolase activity to <5% of the wild-type value. Structure-activity relationships were elucidated by introducing conservative substitutions at the essential positions. The mutational results suggest that the active site of cvRtp1 is likely to adopt a tunnel fold like that of Cet1 and that a similar constellation of side chains within the tunnel is responsible for metal binding and reaction chemistry. Nonetheless, there are several discordant mutational effects in cvRtp1 versus Cet1, which suggest that different members of the phosphohydrolase family vary in their reliance on certain residues within the active site tunnel. We found that tripolyphosphate and pyrophosphate were potent competitive inhibitors of cvRtp1 (K(i) = 0.6 microm tripolyphosphate and 2.4 microm pyrophosphate, respectively), whereas phosphate had little effect. cvRtp1 displayed a weak intrinsic tripolyphosphatase activity (3% of its ATPase activity) but was unable to hydrolyze pyrophosphate.     

Effects of growth, food intake, and dietary zinc on diadenosine tetraphosphate concentrations in rats

The nucleotide diadenosine tetraphosphate has been suggested to function as a signal molecule for the initiation of DNA replication. Previous studies have indicated that diadenosine tetraphosphate is synthesized by certain aminoacyl tRNA synthetases and that diversion of AMP from the amino acid-enzyme complex to ATP to form diadenosine tetraphosphate is facilitated by zinc ions. The growth retardation of zinc-deficient rats is associated with specific reduction in DNA replication and also with a potentially growth-limiting decrease in food intake. The possibility has been investigated that in zinc-deficient rats, lack of Zn(2+) restricts diadenosine tetraphosphate synthesis, resulting in a failure to synthesize DNA and in a reduction in growth. The results indicate that the depressed growth potential caused by the reduction in food intake associated with the deficiency was sufficient to lower diadenosine tetraphosphate concentrations significantly in the liver and spleen. However, there was no indication of a specific effect of zinc deficiency on diadenosine tetraphosphate values.     

Fungicidal activity of Ranunculus asiaticus and other weeds against Fusarium oxysporum f.sp. lycopersici

The effects of aqueous extracts of some common weed species against Fusarium oxysporum Schlecht f.sp. lycopersici (the causal agent of tomato wilt disease) were investigated under laboratory conditions. Anagallis foemina L., Cerastium dicotomum L., Falcaria vulgaris L., Ranunculus asiaticus L., Scorpiurus mur-icatus L. and Solanum nigrum L. extracts were the most toxic to the fungus. Further studies on buttercup (Ranunculus asiaticus L.) showed that fresh shoot extract of this species prevented growth of F. oxysporum when incorporated into agar medium. Extracts of different parts of the plant inhibited fungus growth and sporulation, but the fungitoxicity decreased with incubation period with only slight changes in the toxicity of fresh shoot extract. The shoot and fresh parts extracts were more toxic than root and dried tissue extracts. Addition of 0.5 ml fresh shoot or 1 ml fresh root extract to the growing medium significantly reduced fungal colony growth, and the effect was extract concentration dependent. Fresh shoot extract of R. asiaticus added to a liquid medium significantly reduced mycelial dry weight compared with the control, and incorporation of 0.1 g dried shoot or 0.2 g dried roots in the media strongly inhibited fungus growth. Results of a pot experiment showed no harmful effects of R. asiaticus extracts on tomato growth.     

Clotrimazole as a pharmaceutical: past,present and future.

Clotrimazole is a broad‐spectrum antimycotic drug mainly used for the treatment of Candida albicans and other fungal infections. A synthetic, azole antimycotic, clotrimazole is widely used as a topical treatment for tinea pedis (athlete's foot), as well as vulvovaginal and oropharyngeal candidiasis. It displays fungistatic antimycotic activity by targeting the biosynthesis of ergosterol, thereby inhibiting fungal growth. As well as its antimycotic activity, clotrimazole has become a drug of interest against several other diseases such as sickle cell disease, malaria and some cancers. It has also been combined with other molecules, such as the metals, to produce clotrimazole complexes that show improved pharmacological efficacy. Moreover, several new, modified‐release pharmaceutical formulations are also undergoing development. Clotrimazole is a very well‐tolerated product with few side effects, although there is some drug resistance appearing among immunocompromised patients. Here, we review the pharmaceutical chemistry, application and pharmacology of clotrimazole and discuss future prospects for its further development as a chemotherapeutic agent.