Polyphosphate Chain Length Research Articles

Physical and chemical changes to commercial sodium phosphate glasses after aging or dynamic vapor sorption

Commercial sodium phosphate glasses are used as reference materials for polyphosphate chain length analysis. Water can spontaneously hydrolytically degrade the characteristic polyphosphate P-O-P bond, and reduce the polyphosphate chain length. XRD, Raman spectroscopy, and SEM were used to characterize glasses stored for two weeks, six months and >10 years, and three commercial sodium phosphate glasses after controlled water vapor adsorption and desorption. Rapid water vapor absorption was measured above 50 % relative humidity. The chain length distribution for a dissolved commercial sodium phosphate glass with an average chain length of 45 PO3− units was analyzed with polyacrylamide gel electrophoresis. A distribution of linear polyphosphates, interrupted with a population spike of ∼13 PO3− units, trimetaphosphate rings, and orthophosphates were identified. 31P-MAS NMR suggested that the linear polyphosphate population spike could be the product of large ring polyphosphate scission upon dissolution. This study confirms the importance of dry storage for chain length reference phosphate glasses.

Short-chain polyphosphates: Extraction effects on migration and size estimation of Saccharomyces cerevisiae extracts with polyacrylamide gel electrophoresis.

Polyacrylamide gel electrophoresis is commonly used to characterize the chain length of polyphosphates (polyP), more generally called condensed phosphates. After separation, nonradioactive, optical polyP staining is limited to chain lengths greater than 15 monomers with toluidine blue or 4',6-diamidino-2-phenylindole. PolyP chain lengths longer than 62 monomers were correlated to the shortest DNA ladders. In this study, synthetic linear polyPs (Sigma-Aldrich "Type 45", estimated mean length of 45 monomers), trimetaphosphate (trimetaP: 3 ring), tripolyphosphate (tripolyP), pyrophosphate (PPi ), and inorganic orthophosphate (o-Pi ) were visualized after separation by an in situ hydrolytic degradation process to o-Pi that was subsequently stained with methyl green. Statistically insignificant migration reduction of synthetic short-chain polyP after perchloric acid or phenol-chloroform extraction was confirmed with the Friedman test. 31 P diffusion-ordered NMR spectroscopy confirmed that extraction also reduced PPi diffusivity by <10%. Linear regression between the Rf peak migration value and the logarithm of synthetic polyP molecular weights enabled estimation of extracted polyP chain lengths from 2 to 45 monomers. Linear polyP extracts from Saccharomyces cerevisiae grown in aerobic conditions were generally shorter than extracts cultured in anaerobic conditions. Extractions from both aerobic and anaerobic S. cerevisiae included tripolyP and o-Pi , but no PPi .

XANES Study of Tribofilm Formation With Low Phosphorus Additive Mixtures of Phosphonium Ionic Liquid and Borate Ester

The development of low phosphorus engine oils is important to minimize phosphorus-induced exhaust catalyst poisoning and resulting in harmful emissions. In this study, low phosphorus oil formulations were prepared by using an ashless additive mixture of borate ester (SB) with ionic liquid composed of a phosphonium cation and phosphate anion (P_DEHP) at 350 and 700 ppm phosphorus. Tribological properties of this binary additive system were evaluated using a reciprocating cylinder on a flat test configuration. Favorable interaction between P_DEHP and SB resulted in a significant reduction in friction coefficient and wear volume, in particular for P_DEHP(700P) + SB oil blend. Time-scale analysis of tribofilm formation was determined by running the tribological experiments for 5, 15, and 60 min duration. Electrical contact resistance (ECR) results revealed that the addition of P_DEHP at 350 ppm of phosphorus to SB at 500 ppm of boron can reduce the incubation time from 300 to 100 s for stable tribofilm formation. X-ray absorption near-edge spectroscopy (XANES) analysis of tribofilms indicates that the tribofilm mechanism for additive mixtures of P_DEHP and SB initially involves the formation of boron oxide-based films, which later interact with phosphorus to form boron phosphates in addition to iron phosphates. Incorporation of the high amount of boron phosphates in addition to boron oxide/acid and iron phosphates in the tribofilms contributed to the improved tribological performance of P_DEHP(700P) + SB oil. XANES results reveal that tribofilms formed due to the interaction of SB and P_DEHP evolve to a cross-linked structure, wherein the chain length of polyphosphates is increased with the increase in rubbing time.

Polyphosphates diminish solubility of a globular protein and thereby promote amyloid aggregation

Structural changes of globular proteins and their resultant amyloid aggregation have been associated with various human diseases, such as lysozyme amyloidosis and light-chain amyloidosis. Because many globular proteins can convert into amyloid fibrils in vitro, the mechanisms of amyloid fibril formation have been studied in various experimental systems, but several questions remain unresolved. Here, using several approaches, such as turbidimetry, fluorescence and CD spectroscopy, EM, and isothermal titration calorimetry, we examined the binding of polyphosphates to hen egg-white lysozyme under acidic conditions and observed polyphosphate-induced structural changes of the protein promoting its aggregation. Our data indicate that negatively charged polyphosphates bind to protein molecules with a net positive charge. The polyphosphate-bound, structurally destabilized protein molecules then start assembling into insoluble amorphous aggregates once they pass the solubility limit. We further show that the polyphosphates decrease the solubility limit of the protein and near this limit, the protein molecules are in a labile state and highly prone to converting into amyloid fibrils. Our results explain how polyphosphates affect amorphous aggregation of proteins, how amyloid formation is induced in the presence of polyphosphates, and how polyphosphate chain length is an important factor in amyloid formation.

Determination of individual chain length and chain-length distribution of polyphosphates in microalgae by 31P-DOSY-NMR

Due to its ecological and biotechnological relevance, polyphosphate in microalgae is currently the focus of intense research. Numerous biological functions are performed by or dependent on polyphosphate, and they depend, among other factors, on its chain length. Chain length determination is important for understanding polyphosphate metabolism and for maximizing intracellular polyphosphate abundance per unit weight of biomass. 31P-DOSY NMR virtually separates various polyphosphate polymers in a mixture based on different translational diffusion coefficients. The diffusion coefficient of a polyphosphate molecule correlates with its molecular weight, enabling determination of individual chain lengths. Moreover, the polydispersity index can also be uniquely determined by DOSY as a measure of the overall chain-length distribution of polyphosphates. By contrast, conventional 31P NMR is only able to estimate the average chain length of the entire polyphosphate pool. Therefore, DOSY provides the opportunity to deepen our insight into polyphosphate metabolism and dynamics in algal biomass.

Polyphosphate Chain Length Determination in the Range of Two to Several Hundred P-Subunits with a New Enzyme Assay and 31P NMR.

Currently, 31P NMR is the only analytical method that quantitatively determines the average chain length of long inorganic polyphosphate (>80 P-subunits). In this study, an enzyme assay is presented that determines the average chain length of polyphosphate in the range of two to several hundred P-subunits. In the enzyme assay, the average polyP chain length is calculated by dividing the total polyphosphate concentration by the concentration of the polyphosphate chains. The total polyphosphate is determined by enzymatic polyphosphate hydrolysis with Saccharomyces cerevisiae exopolyphosphatase 1 and S. cerevisiae inorganic pyrophosphatase 1, followed by colorimetric orthophosphate detection. Because the exopolyphosphatase leaves one pyrophosphate per polyphosphate chain, the polyphosphate chain concentration is assayed by coupling the enzymes exopolyphosphatase (polyP into pyrophosphate), ATP sulfurylase (pyrophosphate into ATP), hexokinase (ATP into glucose 6-phosphate), and glucose 6-phosphate dehydrogenase (glucose 6-phosphate into NADPH), followed by fluorometric NADPH detection. The ability of 31P NMR and the enzyme assay to size polyP was demonstrated with polyP lengths in the range from 2 to ca. 280 P-subunits (no polyP with a longer chain length was available). The small deviation between methods (-4 ± 4%) indicated that the new enzyme assay performed accurately. The limitations of 31P NMR (i.e., low throughput, high sample concentration, expensive instrument) are overcome by the enzyme assay that is presented here, which allows for high sample throughput and requires only a commonly available plate reader and micromole per liter concentrations of polyphosphate.

Minocycline-loaded calcium polyphosphate glass microspheres as a potential drug-delivery agent for the treatment of periodontitis.

Background: Periodontitis is an inflammatory disease with a bacterial etiology that affects the supporting structures of the teeth and is a major cause of tooth loss. The objective of this study was to investigate the drug loading and in vitro release of minocycline from novel calcium polyphosphate microspheres intended for use in treating periodontitis. Methods: Calcium polyphosphate coacervate, produced by a precipitation reaction of calcium chloride and sodium polyphosphate solutions, was loaded with minocycline and subsequently used to produce microspheres by an emulsion/solvent extraction technique. Microspheres classified by size were subjected to a 7-day elution in a Tris-buffer solution under dynamic conditions. The physicochemical characteristics of the drug-loaded microspheres were investigated using scanning electron microscopy, particle size analysis, Phosphorus-31 Nuclear Magnetic Resonance spectroscopy, and Inductively Coupled Plasma Optical Emission Spectroscopy. Drug loading and release were determined using ultraviolet -visible (UV/VIS) spectrophotometry. Results: Minocycline-loaded calcium polyphosphate microspheres of varying size were successfully produced, with small and large microspheres having volume mean diameters of 22 ± 1 µm and 193 ± 5 µm, respectively. Polyphosphate chain length and calcium to phosphorus mole ratio remained stable throughout microsphere production. Drug loading was 1.64 ± 0.16, 1.35 ± 0.55, and 0.84 ± 0.14 weight% for the coacervate and large and small microspheres, respectively, corresponding to mean encapsulation efficiencies of 81.7 ± 12.2 % and 50.9 ± 3.9 % for the large and small microspheres. Sustained drug release was observed in vitro over a clinically relevant 7-day period, with small and large microspheres exhibiting similar elution profiles. Antibiotic release generally followed microsphere degradation as measured by Ca and P ion release. Conclusions: This study demonstrated successful drug loading of calcium polyphosphate microspheres with minocycline. Furthermore, in vitro sustained release of minocycline over a 7-day period was observed, suggesting potential utility of this approach for treating periodontitis.

Phosphatase-Mediated Hydrolysis of Linear Polyphosphates.

Polyphosphates are a group of phosphorus (P) containing molecules that are produced by a wide range of microorganisms and human activities. Although polyphosphates are ubiquitous in aquatic environmentsand are of environmental significance, little is known about their transformation and cycling. This study characterized the polyphopshate-hydrolysis mechanisms of several representative phosphatase enzymes and evaluated the effects of polyphosphate chain length, light condition, and calcium (Ca2+). 31P nuclear magnetic resonance (NMR) spectroscopy was used to monitor the dynamic changes of P molecular configuration during polyphosphate hydrolysis and suggested a terminal-only degradation pathway by the enzymes. Such mechanism enabled the quantification of the hydrolysis rates by measuring orthophosphate production over time. At the same initial concentration of polyphosphate molecules, the hydrolysis rates were independent of chain length. The hydrolysis of polyphosphate was also unaffected by light condition, but was reduced by the presence of Ca2+. The released orthophosphates formed Ca-phosphate precipitates in the presence of Ca2+, likely in amorphous phases. Results from this study lay the foundation for better understanding the chemical processes governing polyphosphate transport and transformation in various environmental settings.

Tribological properties and surface interaction of novel water-soluble ionic liquid in water-glycol

Abstract Two kinds of novel water-soluble ILs were synthesized and applied as lubricant additives in water-glycol. ILs, as the multifunctional and high efficiency additive, could drastically improve antiwear, friction-reducing and extreme pressure properties of water-glycol. SEM and EDS results indicated that the sample lubricated by PEG600MO phosphate-ammonium IL (P1) acquires higher active element content in the area with heavy wear, while that lubricated by PEG600MO phosphite-ammonium IL (P2) is relatively homogeneous on the worn surface with heavy and mild wear. L-edge XANES results showed that the tribofilm is composed of polyphosphate, and the increase of the polyphosphate chain length could improve the tribological properties. Additionally, the additive concentration, the rubbing time and the load could change the chain length of polyphosphate.

Cell wall canals formed upon growth of Candida maltosa in the presence of hexadecane are associated with polyphosphates.

Canals are supramolecular complexes observed in the cell wall of Candida maltosa grown in the presence of hexadecane as a sole carbon source. Such structures were not observed in glucose-grown cells. Microscopic observations of cells stained with diaminobenzidine revealed the presence of oxidative enzymes in the canals. 4΄,6΄-diamino-2-phenylindole staining revealed that a substantial part of cellular polyphosphate was present in the cell wall of cells grown on hexadecane in condition of phosphate limitation. The content and chain length of polyphosphates were higher in hexadecane-grown cells than in glucose grown ones. The treatment of cells with yeast polyphosphatase PPX1 resulted in the decrease of the canal size. These data clearly indicated that polyphosphates are constituents of canals; they might play an important role in the canal structure and functioning.

Synergistic effect between organic borate esters and phosphorus-based additives on tribological performances as lubricant additives in mineral oil

Nowadays, looking for environment-friendly lubricant additives to replace phosphorus-based derivatives in automotive lubricating system is one of the main objectives. Organic borate ester is one kind of potential substitutes for phosphorus-based derivatives. In this paper, the interactions of organic borate esters and dioctylphosphite ester on tribological performances were studied. Results showed that they had a synergistic effect on antiwear performance and friction-reducing property as lubricant additives in mineral oil. The chemical nature of worn surface was explored by X-ray absorption near-edge structure spectroscopy. Borate ester could facilitate the decomposition of phosphorus-based additive and the formation of phosphate. What is more, boron-containing species could react with phosphate to form boron–phosphorus-based thin film on the surface. Boron mainly forms B2O3 on the surface. In the near surface of tribofilm, nitrogen-containing borate ester reduced the polyphosphate chain length due to the formation of ammonium cation. The polyphosphate chain length of the tribofilm generated from dioctylphosphite ester mixed with nitrogen-containing alkyl borate ester is shorter compared to that of the tribofilm generated from dioctylphosphite ester mixed with nitrogen-containing heterocyclic borate ester.

A novel route for preparing 5' cap mimics and capped RNAs: phosphate-modified cap analogues obtained via click chemistry.

The significant biological role of the mRNA 5' cap in translation initiation makes it an interesting subject for chemical modifications aimed at producing useful tools for the selective modulation of intercellular processes and development of novel therapeutic interventions. However, traditional approaches to the chemical synthesis of cap analogues are time-consuming and labour-intensive, which impedes the development of novel compounds and their applications. Here, we explore a different approach for synthesizing 5' cap mimics, making use of click chemistry (CuAAC) to combine two mononucleotide units and yield a novel class of dinucleotide cap analogues containing a triazole ring within the oligophosphate chain. As a result, we synthesized a library of 36 mRNA cap analogues differing in the location of the triazole ring, the polyphosphate chain length, and the type of linkers joining the phosphate and the triazole moieties. After biochemical evaluation, we identified two analogues that, when incorporated into mRNA, produced transcripts translated with efficiency similar to compounds unmodified in the oligophosphate bridge obtained by traditional synthesis. Moreover, we demonstrated that the triazole-modified cap structures can be generated at the RNA 5' end using two alternative capping strategies: either the typical co-transcriptional approach, or a new post-transcriptional approach based on CuAAC. Our findings open new possibilities for developing chemically modified mRNAs for research and therapeutic applications, including RNA-based vaccinations.

Degradation and hemostatic properties of polyphosphate coacervates

Sodium polyphosphate is a linear polymer formed from phosphate units linked together by sharing oxygen atoms. Addition of calcium to a solution of sodium polyphosphate results in phase separation and formation of a polyphosphate coacervate best described as a polymeric rich viscoelastic material. Polyphosphate coacervate is an interesting candidate as a biomaterial based on its ability to bind with different cations and to be loaded with drugs. Here, in vitro degradation and hemostatic properties of polyphosphate coacervates are comprehensively evaluated. We show that polyphosphate coacervates degrade and dissolve at a fast rate, losing half of their original mass in a week and transforming to mainly pyrophosphate after 4weeks. This burst dissolution phase happens earlier for the coacervate prepared from very short chain polyphosphate but overall using longer polyphosphate chains does not increase the coacervate longevity significantly. Substitution of Ca with Sr or Ba does not affect the hydrolysis of coacervates but slows down their dissolution into the media. In a whole blood clotting assay, coacervates profoundly decrease the clotting time especially when very long chain polyphosphates are used. While coacervate chain length and divalent cation type were found to significantly affect prothrombin time and thromboplastin time compared to the control, no discernible trends were observed. Platelets adhere in large numbers to coacervates, especially those containing long chain polyphosphate, but the cell morphology observed suggests that they might not to be fully activated. Overall, the long chain polyphosphate coacervate holds a great potential as a resorbable hemostatic agent. Statement of SignificanceDivalent cation additions to a sodium polyphosphate solution result in polyphosphate coacervates, or highly viscous gel-like materials, having great potential in bio-applications such as drug delivery and hemostasis. As these coacervates degrade in aqueous environments, we undertook a comprehensive evaluation to better understand the impact of polyphosphate chain length and divalent cation substitution on this hydrolytic response in order to better predict degradation behavior in the body. Furthermore, there is great interest in the role of polyphosphates in hemostasis following recent publications showing that platelets secrete polyphosphates upon thrombin stimulation. In this paper, we evaluate the hemostatic potential of polyphosphate coacervates as bulk constructs, demonstrating that indeed these materials hold great potential as a degradable hemostatic agent.

Rheology of polyphosphate coacervates

Polyphosphate is a highly water-soluble linear polyanion comprised of phosphate groups. A phase separation happens when divalent cations are added to polyphosphate solutions resulting in the formation of polyphosphate coacervates. Such coacervates could potentially be used as a glass precursor or in a variety of bioapplications including microencapsulation. In all of these applications, viscoelastic properties of polyphosphate coacervates directly affect their usage. Here, we show that these polyphosphate coacervates act as extremely viscous Newtonian liquids at low shear rates, with their viscosity highly dependent on the polyphosphate degree of polymerization (Dp) and type of divalent cations (Ca2+, Sr2+, or Ba2+) used for their preparation. For Ca2+-only polyphosphate coacervates, specific viscosity is directly related to Dp1.46 at 20 °C, similar to highly concentrated polymer solutions where chain entanglement is not important. For very long chain polyphosphate coacervates, however, elastic properties are dominant presumably caused by physical chain entanglement. Replacement of a fraction of Ca2+ with Sr2+ or Ba2+ results in coacervates having significantly more elastic characters and profoundly higher viscosity than their Ca2+-only counterparts. Overall, varying polyphosphate chain length and divalent cation type allows one to modify these materials for a desired application.

Polyphosphates and Polyphosphatase Activity in the Yeast Saccharomyces cerevisiae during Overexpression of the DDP1 Gene.

The effects of overexpression of yeast diphosphoinositol polyphosphate phosphohydrolase (DDP1) having endopolyphosphatase activity on inorganic polyphosphate metabolism in Saccharomyces cerevisiae were studied. The endopolyphosphatase activity in the transformed strain significantly increased compared to the parent strain. This activity was observed with polyphosphates of different chain length, being suppressed by 2mM tripolyphosphate or ATP. The content of acid-soluble and acid-insoluble polyphosphates under DDP1 overexpression decreased by 9 and 28%, respectively. The average chain length of salt-soluble and alkali-soluble fractions did not change in the overexpressing strain, and that of acid-soluble polyphosphate increased under phosphate excess. At the initial stage of polyphosphate recovery after phosphorus starvation, the chain length of the acid-soluble fraction in transformed cells was lower compared to the recipient strain. This observation suggests the complex nature of DDP1 involvement in the regulation of polyphosphate content and chain length in yeasts.

Mechanism of tribofilm formation with P and S containing ionic liquids

Mechanism of tribofilm formation with two ionic liquids (IL), choline bis(2-ethylhexyl)phosphate and choline dibutyldithiophosphate was studied. XANES analysis of tribofilms indicates that the underlying mechanism of tribofilm formation with ionic liquids is similar to that formed when ZDDP is used. The chain length of glassy polyphosphates with IL in base oil is longer in length in comparison to that formed with ZDDP under identical conditions indicating a higher level of networking. In fully formulated oils, Ca replaces Zn and Fe (in the case of ZDDP) or Fe (when IL׳s are used) as the primary cationic species present in the polyphosphate network. The sulfur is present in the form of sulfates of different cationic species including Fe and Ca.

Polyphosphate Chain Length Variations and Implications in Enhanced Biological Phosphorus Removal (EBPR) Systems

Polyphosphate Chain Length Variations and Implications in Enhanced Biological Phosphorus Removal (EBPR) Systems

Tribological properties of Mg/Al–CO3 layered double hydroxide as additive in base oil

The tribological performance of Mg/Al–CO3–layered double hydroxide (LDHs) nanoparticles in base oil was studied as a sole additive and in combination with zinc dialkyldithiophosphate (ZDDP) at 100°C. Results show that LDHs could improve antifriction and antiwear properties of base oil. The blend with LDHs alone shows better antifriction properties than that of the mixture of LDHs and ZDDP, but the addition of ZDDP helps to stabilise the friction coefficient. Surface analyses have been performed to study the morphology, nanoparticle distribution and chemical species in the tribofilm. The results show that tribofilms contain Mg, Al, C, O, Zn, P and S, and the structure of Mg/Al–CO3–LDHs changes under shearing stress and this process can help reduce friction coefficient. Thus, LDHs could be chemically incorporated into tribofilm to reduce the polyphosphate chain length originated from ZDDP decomposition resulting in a medium chain polyphosphate, which provides very good wear protection.

The importance of chain length for the polyphosphate enhancement of acidic potassium permanganate chemiluminescence

Sodium polyphosphate is commonly used to enhance chemiluminescence reactions with acidic potassium permanganate through a dual enhancement mechanism, but commercially available polyphosphates vary greatly in composition. We have examined the influence of polyphosphate composition and concentration on both the dual enhancement mechanism of chemiluminescence intensity and the stability of the reagent under analytically useful conditions. The average chain length (n) provides a convenient characterisation, but materials with similar values can exhibit markedly different distributions of phosphate oligomers. There is a minimum polyphosphate chain length (∼6) required for a large enhancement of the emission intensity, but no further advantage was obtained using polyphosphate materials with much longer average chain lengths. Providing there is a sufficient average chain length, the optimum concentration of polyphosphate is dependent on the analyte and in some cases, may be lower than the quantities previously used in routine detection. However, the concentration of polyphosphate should not be lowered in permanganate reagents that have been partially reduced to form high concentrations of the key manganese(III) co-reactant, as this intermediate needs to be stabilised to prevent formation of insoluble manganese(IV).

The effect of individual phosphate emulsifying salts and their selected binary mixtures on hardness of processed cheese spreads

The aim of this work was to observe the effects of emulsifying salts composed of trisodium citrate and sodium phosphates with different chain length (disodium phosphate (DSP), tetrasodium diphosphate (TSPP), pentasodium triphosphate (PSTP) and sodium salts of polyphosphates with 5 different mean length (n ≈ 5, 9, 13, 20, 28)) on hardness of processed cheese spreads. Hardness of processed cheese spreads with selected binary mixtures of the above mentioned salts were also studied. Measurements were performed after 2, 9 and 30 days of storage at 6 °C. Hardness of processed cheese increased with increase in chain length of individually used phosphates. Majority of applied binary mixtures of emulsifying salts had not significant influence on hardness charges in processed cheese spreads. On the other hand, a combination of phosphates salts (DSP with TSPP) was found, which had specific effect on hardness of processed cheese spreads. Textural properties of samples with trisodium citrate were similar compared to samples with DSP.