Presence Of CaCl2 Research Articles

Isolation and characterisation of micellar casein from buffalo milk using microfiltration technique with modified buffer composition

The objective of this work was to study the effect of di‐sodium phosphate (Na2HPO4) and calcium chloride (CaCl2) salt concentrations (10 to 90 mM) on the isolation of casein micelles from buffalo milk by microfiltration (0.20 μm) process. Destabilisation of buffalo micellar casein resulted in the presence of CaCl2. A complete isolation of caseins from buffalo skim milk was achieved using modified buffer containing 90 mM Na2HPO4 after 4th cycle of washing with 92% removal of whey proteins. Modification of casein micelles with Na2HPO4 resulted in 33% removal of calcium in the preparation of buffalo micellar casein. Based on transmission electron micrographs, no aggregation was recorded for buffalo micellar casein and size varied from 30 to 80 nm.

Solution and interfacial self-assembly of Bacillus subtilis bacterial lipoteichoic acid (LTA): nanoclustering, and effects of Ca2+ and temperature.

Lipoteichoic acid (LTA) is a major structural and functional molecule in the Gram-positive bacteria membrane. Knowledge of LTA adsorption at interfaces and its solution self-assembly is crucial to understanding its role in bacterial adhesion and colonisation, infections and inflammations. Here, we report the self-assembly behaviour of LTA extracted from Bacillus subtilis, a Gram-positive bacterium, in an aqueous solution using cryogenic transmission electron microscopy (Cryo-TEM) and small-angle neutron scattering (SANS) and its adsorption behaviour at the solid-liquid interface using atomic force microscopy (AFM) imaging and quartz crystal microbalance with dissipation monitoring (QCM-D). The Cryo-TEM results indicated the formation of spherical LTA micelles that decreased in size on addition of calcium chloride (CaCl2), attributed to charge neutralisation and possible formation of stable Ca2+-bridges between the phosphate groups on neighbouring LTA chains. Analysis of the SANS data from the polydisperse LTA aggregates in solution using the two Lorentzian model revealed the existence of two correlation lengths, which could respectively account for the presence of LTA micelle clusters and the local structure arising from LTA intra-molecular interactions. In the presence of CaCl2, the decrease in the correlation lengths of the clusters indicated possible disruption of H-bonding by Ca2+, leading to poorer water-LTA interactions. At higher temperatures, the correlation length corresponding to the clusters increased, indicating a temperature assisted growth caused by the fluidization of micellar core and dehydration of the polar LTA chains. AFM imaging showed that adsorption of LTA aggregates at the SiO2-water interface was significantly prompted by the addition of CaCl2, also confirmed by QCM-D measurements. These unprecedented nanoscopic structural details on the morphology of LTA aggregates in solution and at the solid-liquid interface add to our fundamental understanding of its self-assembly behaviour hitherto underexplored.

Redox behaviour of CaCl2 melts in presence of moisture as impurity. Part I: Cyclic voltammetry

• Redox behaviour of bare CaCl 2 melts -explored using cyclic voltammetry at 900 °C at inert tungsten electrode. • Effect of addition of CaCl 2 .2H 2 O and Ca(OH) 2 in CaCl 2 melt studied using cyclic voltammetry. • Peak current densities observed in bare melts enhanced with addition of CaCl 2 .2H 2 O or Ca(OH) 2 . • Complex reduction behaviour attributed to formation of CaOHCl in melt at high temperatures. • Peak analysis of cyclic voltammograms and semi-integral curves carried out; cathodic reduction mechanism proposed. Redox behaviour of CaCl 2 melt at 900 °C is well known; what is not completely established so far is how the voltammetric behaviour of melt would be in presence of moisture. CaCl 2 prepared from vacuum drying of CaCl 2 .2H 2 O retained some residual moisture that eventually led to formation of CaOHCl at 900 °C as a result of which two redox couples were observed in the cyclic voltammograms at tungsten electrode. In order to decipher whether the redox peaks were a reflection of formation of single electro-active species, CaCl 2 .2H 2 O and Ca(OH) 2 were added in pre-electrolysed CaCl 2 melts and those were again analysed by cyclic voltammetry. Enhancement of peak current density of redox peaks, which were observed in pre-electrolysed CaCl 2 melt, was seen. This was attributed to the increased concentration of CaOHCl in the melt in presence of CaCl 2 .2H 2 O and Ca(OH) 2 . Subsequent to voltammetric peak analysis and logarithmic analysis of semi-integral curves for all melts, a mechanism involving formation of ( CaOH ) 2 2 + adsorbed undergoing a one-step two-electron transfer process in the first redox step was proposed. Second redox step resulted due to adsorption of O 2 - on tungsten, and its peak current density also got enhanced due to addition of CaCl 2 .2H 2 O and Ca(OH) 2 in melt.

Effects of common ions on the flotation of fluorapatite and dolomite with oleate collector

In this study, the effects of common ions, especially Ca2+, on the reverse flotation of fluorapatite and dolomite in acidic conditions with NaOL collector were investigated using micro-flotation, zeta potential, contact angle, adsorption analysis, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy and density functional theory-based first principal calculations. It is found that the dolomite flotation recovery was enhanced and the fluorapatite flotation was depressed by the addition of CaCl2 at pH 5. For dolomite, in the presence of CaCl2, the solubility of metal ions was increased, especially for Ca2+. The Ca and Mg sites exposed on the dolomite surface, increased the NaOL adsorption, thus, enhancing the dolomite flotation. However, the formation of Ca(H2PO4)2 species on fluorapatite surface inhibit its flotation.

Design of Lipid-Based Nanocarriers via Cation Modulation of Ethanol-Interdigitated Lipid Membranes.

Short-chain alcohols (i.e., ethanol) can induce membrane interdigitation in saturated-chain phosphatidylcholines (PCs). In this process, alcohol molecules intercalate between phosphate heads, increasing lateral separation and favoring hydrophobic interactions between opposing acyl chains, which interpenetrate forming an interdigitated phase. Unraveling mechanisms underlying the interactions between ethanol and model lipid membranes has implications for cell biology, biochemistry, and for the formulation of lipid-based nanocarriers. However, investigations of ethanol-lipid membrane systems have been carried out in deionized water, which limits their applicability. Here, using a combination of small- and wide-angle X-ray scattering, small-angle neutron scattering, and all-atom molecular dynamics simulations, we analyzed the effect of varying CaCl2 and NaCl concentrations on ethanol-induced interdigitation. We observed that while ethanol addition leads to the interdigitation of bulk phase 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayers in the presence of CaCl2 and NaCl regardless of the salt concentration, the ethanol-induced interdigitation of vesicular DPPC depends on the choice of cation and its concentration. These findings unravel a key role for cations in the ethanol-induced interdigitation of lipid membranes in either bulk phase or vesicular form.

Long-term stability, high strength, and 3D printable alginate hydrogel for cartilage tissue engineering application

Cartilage damage is one of the main causes of disability, and 3D bioprinting technology can produce complex structures that are particularly suitable for constructing a customized and irregular tissue engineering scaffold for cartilage repair. Alginate is an attractive biomaterial for bioinks because of its good biological safety profile and fast ionic gelation. However, ionically crosslinked alginate hydrogels are recognized as lacking enough mechanical property and long-term stability due to ion exchange. Here, we developed a double crosslinked alginate (DC-Alg) hydrogel for 3D bioprinting, and human umbilical cord mesenchymal stem cells (huMSCs) could differentiate into chondrocytes on its printed 3D scaffold after 4 weeks’ culture. We performed sequential modification of alginate with L-cysteine and 5-norbornene-2-methylamine, and the DC-Alg hydrogels were obtained in the presence of CaCl2 and ultraviolet light with stronger mechanical properties than those of the single ionic crosslinked alginate hydrogels, which was similar to natural cartilage. They also had better stability and could be maintained in DMEM medium for over 1 month, as well good viability for huMSCs. Moreover, the DC-Alg as 3D printing inks demonstrated a better printing accuracy (∼200 µm). After 4 weeks culture of huMSCs in the 3D printed DC-Alg scaffolds, the expressions of chondrogenic genes such as aggrecan (agg), collagen II (col II), and SRY-box transcription factor 9 (sox-9) were obviously observed, indicating the differentiation of huMSCs into cartilage. Immumohistochemical staining analysis further exhibited cartilage tissue developed well in the 3D printed scaffolds. Our study is the first demonstration of DC-Alg in 3D printing for MSC differentiation into cartilage, which shows a potential application in cartilage defect repair.

Behaviour of flotation tailings from a rare earth element deposit at high salinity

Various rare earth element (REE) deposits hosted by carbonatite complexes have been identified in southern (Montviel, Niobec) and northern Quebec (Eldor deposit). During the winter in Quebec, the use of road salts to facilitate transportation on the mine site and/or avoid water freezing during mine operation may be necessary. The sources of salinity can be diverse on a mine site: process water, precipitation, alteration of minerals in the soil. Thus, tailings may come in contact with these salts and react. The purpose of the present study was to evaluate the impact of salinity on the behaviour of flotation tailings (Eldor deposit), i.e. the mobility of the elements contained in the tailings under these conditions and the environmental risks involved. For this purpose, leaching column tests were developed. The solutions were deionized water (CW column), NaCl (25 g/L; CS1 column) and CaCl2 (25 g/L; CS2 column). The leachate analysis revealed that the divalent cations (Ba, Cd, Mg, Mn, Sr, and Zn) are more mobile in the presence of CaCl2 (CaCl2 > NaCl > deionized water). The mobility of these elements appears to be governed by the competition with Ca2+ for tailings sorption sites. U and Sc are most mobile in the presence of salts regardless of the applied salt solution, i.e. CaCl2 = NaCl > deionized water. The formation of soluble chloride complexes with these elements could therefore be the cause of this phenomenon. For S, the leaching solution has no impact on its mobility. In conclusion, the presence of salts would tend to increase the mobility of divalent cations present in these residues and enhance their contamination potential. Modeling using PHREEQC software allowed comparison of these results with post-dismantling mineralogical characterization. Both methods showed: (i) total dissolution of fluorite [CaF2], galena [PbS], richterite [Na(CaNa(Mg,Fe2+)5[Si8O22](OH)] and Ba silicate; (ii) precipitation of iron oxides/hydroxides and silicate minerals. However, the modeling was unable to predict the behaviour of carbonate minerals. Further modeling tests involving kinetics should be considered in a future study.

Rheological behavior of nanocellulose gels at various calcium chloride concentrations

In this work, the effects of calcium chloride (CaCl2) concentration on the creep-recovery, linear and nonlinear rheological behavior of nanocellulose gels had been investigated to quantify gel properties. The absolute zeta potential of nanocellulose gels were decreased as the CaCl2 concentration increased, which was related to the electrostatic repulsion that origin from carboxyl group could be effectively screened with increasing CaCl2 concentration. Rheological measurements further confirmed this result for nanocellulose gels, which revealed that the increased modulus and viscoelastic properties were obtained in the presence of CaCl2. The rheological properties of nanocellulose gels were showed to depend on CaCl2 concentration. The enhanced gel network structure was related to the Ca2+ ions that promoted crosslink between nanocellulose by salt bridge. This work highlighted the potential of using electrostatic complexation between nanocellulose and Ca2+ ions to form gels, and demonstrated the tunability of the rheological behavior by adjusting the concentration of CaCl2.

Effect of Cations on the Oxidation and Atmospheric Corrosion of Iron Interfaces to Minerals.

Surface corrosion involves a series of redox reactions that are catalyzed by the presence of ions. On infrastructure surfaces and in complex and natural environments, iron surfaces readily undergo redox reactions, impacting chemical processes. In this study, the effect of how cations influence the formation of the mineral scale on iron surfaces and its connection to surface corrosion was investigated in CaCl2(aq) and NaCl(aq) electrolytes. Polarized modulated-infrared reflection absorption spectroscopy (PM-IRRAS) measurements were used to measure the oxidation and formation of carbonates at the air/electrolyte/iron interface, which confirmed that the iron surface oxidized faster in CaCl2(aq) than in NaCl(aq). PM-IRRAS, attenuated total reflectance-Fourier transformed infrared spectroscopy, and X-ray photoelectron spectroscopy show that after the adsorption of atmospheric O2 and CO2, calcium carbonate (CaCO3) in the form of calcite and aragonite was produced on iron in the presence of CaCl2(aq), whereas siderite (FeCO3) was produced on the surface of iron in the presence of NaCl(aq). However, in either solution without gradual O2 and CO2 exposure, a heterogeneous mixture of lepidocrocite (γ-FeOOH) and an iron hydroxy carbonate (Fex(OH)yCO3) was grown on the iron surface. In situ liquid AFM was used to measure the surface roughness in CaCl2(aq) and NaCl(aq), as an estimation of the corrosion rate. In CaCl2(aq), Fe was found to corrode faster than Fe in NaCl(aq) due to more ions at equimolar concentrations. Surface physical changes, as measured by ex situ AFM, confirmed the presence of a heterogeneous mixture of γ-FeOOH and an Fex(OH)yCO3 in the submerged region. This indicates that the cation does not affect the type of mineral grown on the Fe surface in the region completely submerged in the electrolyte. These results suggest that the cations play a unique role in the initial stages of corrosion at the interface region, influencing the uptake of atmospheric CO2 and mineral nucleation. The knowledge gained from these interfacial reactions are important for understanding the connection between surface corrosion, mineral grown, and CO2 capture for sequestration.

Identification of mitigation strategies to reduce acrylamide levels during the production of black olives

Californian-style processes are widely applied to the elaboration of black ripe olives in the table olive industry. During this procedure, a carcinogenic contaminant known as acrylamide is generated in thermal oxidation. The content of this compound can be modified according to the stage of processing. The present study evaluates the conditions pertaining to different phases of the elaboration process with the aim of identifying optimal conditions for the production of table olives with the lowest acrylamide concentration possible. ‘Hojiblanca’ variety olives were used. Olives were harvested at two different maturation stages, specifically, the green and yellow-green stages. Olives with yellow-green maturation indices had higher acrylamide concentrations than green table olives. Significant reductions in the contaminant were observed as storage time increased, with green olives stored for 21 months displaying the lowest acrylamide levels. Acrylamide was also decreased by spraying olives with water (18 % reduction) and washing them with water heated to 25 °C for 40 min (36 % reduction). Following treatment with lye during the packaging and preparation of olives, unpitted olives contained 12–31% more acrylamide than pitted olives and 42–62% more than sliced olives. Outcomes were the same for olives canned both with and without brine. The presence of CaCl2 and the addition of greater NaCl concentrations (from 2% to 4% w/v), increased the acrylamide content in all olive formats. This increase was not observed in olives not stored in liquid. Outcomes reported here are valuable for redesigning the elaboration process of industrial black ripe olives and allowing producers to manufacture better-quality products with significantly reduced acrylamide concentrations.

Controlling the release rate of topotecan from PLGA spheres and increasing its cytotoxicity towards glioblastoma cells by co-loading with calcium chloride

It has been suggested that local administration of topotecan (TT) could increase its efficacy in the treatment of glioblastoma. In this context, a PLGA implant model in the form of spheres with a porous core and stiff surface, loaded with TT and CaCl2 was developed. An array of formulations differing from each other by the type of PLGA used, the integrity of the surface, the concentrations of TT and CaCl2 added during the preparation, and the volume of water in the PLGA mix, was prepared, screened and explored by computerized multifactorial analysis. This analysis enabled the simultaneous identification of the most influential experimental factors on the experimental responses, which were pre-determined as the efficiency of TT loading and the TT % cumulative release at 14 days. The multifactorial analysis also revealed how the interactions among the experimental factors affect the performance of the various formulations. Thus, TT concentration and its factorial interaction with the concentration of CaCl2 added during the spheres’ preparation were identified as most prominent on the loading efficiency, while the surface integrity (intact or punctured) and CaCl2 amount in the spheres were identified as most prominent on the TT % cumulative release from the spheres. TT was found to be cytotoxic towards glioblastoma U87 MG cells, an activity which was enhanced, synergistically, in the presence of CaCl2 (the relative viability was reduced from 36 to 28% with combination indices of 1.0, 0.37, 0.13 and 0.06 for EC50, EC75, EC90 and EC95, respectively). Interestingly, dividing the TT dose into 3 equal portions, replenished daily to the incubation medium, increased TT cytotoxicity. The relative viability was then reduced from 35 to 7% and in the presence of CaCl2 – from 28 to 1.9%, suggesting that a local, slow input of TT could be effective in the treatment of glioblastoma by an adjacent TT implant. The increased effect of CaCl2 on cytotoxicity was also observed when it was co-loaded into the TT spheres. In that case, the cells’ viability was reduced from 72 to 27%. It is suggested that the PLGA spheres could be used for tunable local delivery of TT in post-resection adjuvant therapy of glioblastoma.

Roles of solution chemistry and reagent–reagent interaction on carboxymethylcellulose adsorption onto graphite and implications on its floatability

The influence of electrolytes (NaCl and CaCl2) and a copper sulphide collector (isopropyl ethyl thionocarbamate, IPETC) on adsorption of carboxymethylcellulose (CMC) onto a graphite surface were studied using adsorption tests, zeta potential measurements and floatability tests. Adsorption tests revealed negligible CMC adsorption in the absence of electrolytes and presence/absence of IPETC at lower CMC concentration of 0–100 mg/L, whereas CMC mild adsorption occurred at higher CMC concentration of 150–200 mg/L at pH 5.7 and 10. An increase in CMC adsorption was observed in the presence of electrolytes and their mixtures with IPETC. The CMC adsorption increased more in the presence of CaCl2 than NaCl, suggesting an effect of the cation valence. The addition of IPETC to NaCl solutions adversely affected the CMC adsorption and its effect was independent of the pH. Conversely, the addition of IPETC to CaCl2 solutions enhanced the CMC adsorption at pH 5.7, whereas it diminished at pH 10. The presence of electrolyte may induce the formation of complexes between CMC macromolecules with IPETC depending on the electrolyte type. Furthermore, CMC adsorption in presence of NaCl follows Langmuir isotherm whereas in presence of CaCl2 follows Freundlich isotherm. Moreover, the floatability tests indicated that the graphite floatability decreased more at pH 10 than at pH 5.7 in the presence of electrolytes. Only the floatability at pH 5.7 was comparable for both electrolytes. Finally, addition of IPETC to electrolyte solutions had a marked effect on graphite floatability. These findings have implications in copper sulphide flotation systems containing graphite as gangue in which IPETC is used as a collector and CMC as a depressant for graphite.

Producing cement clinker assemblages in the system: CaO-SiO2-Al2O3-SO3-CaCl2-MgO

The cement industry is carbon-intensive, and the valorisation of industrial side-streams/residuals for use as alternative raw materials can enable the cement industry to reduce its carbon footprint as well as promote resource efficiency. Apart from key clinker ingredients such as CaO, Al2O3, and SiO2, industrial residues can also contain MgO, CaCl2, and SO3. Therefore, this study investigates the formation of cement clinker assemblages in the system CaO-SiO2-Al2O3-SO3-CaCl2-MgO at temperatures ranging between 1100 and 1300 °C. The production of a clinker composed mainly of alinite and ye'elimite is first attempted; it is found that these phases cannot be simultaneously produced. Ternesite is also not compatible with alinite under the conditions studied. Wadalite is compatible with both ye'elimite and ternesite, while ternesite is also compatible with chlormayenite at 1150 °C. Additionally, the low-temperature formation of alite was also observed with the presence of CaCl2 in the raw-material mix.

Enantioselectivity-Switchable Organocatalytic [4 + 2]-Annulation to Access the Spirooxindole-Norcamphor Scaffold.

An organocatalyzed enantiodivergent synthesis of a multifunctionalized spirooxindole-norcamphor scaffold via a [4 + 2]-annulation between cyclicα, β-unsaturated ketone and methylene indolinones has been established. The presence of CaCl2 in DMF could reverse the enantioselectivity to facilely deliver the enantiomers of the corresponding spirooxindoles. Both enantiomers of the corresponding spirooxindoles were obtained in excellent yield and diastereo-/enantioselectivity by employing one single prolinosulfonamide catalyst.

Production, properties and some applications of protease from alkaliphilic Bacillus sp. EBTA6

Extracellular protease production by a novel strain, Bacillus sp. EBTA6, has been optimized by using central composite design of response surface methodology and properties and industrial applications of crude enzyme have been investigated. Three independent variables (temperature, pH and yeast extract concentration) chosen in the experimental design were significant terms and reduced cubic model fit with the design at p < 0.0001 level. The recommended temperature, pH and yeast extract concentration were 30 °C, 8, and 15 g/L, respectively. Crude enzyme displayed activity over a wide pH and temperature ranges having the optimum at 50–60 °C and pH 8. It was quite stable at high pH values and at 50 °C. Amongst the metal ions (Mg+, Cu2+, Ca2+, Zn2+, K2+, and Sn2+), Ca2+ enhanced the activity and the others either decreased or did not change it. The enzyme activity was reduced by phenyl-methyl-sulfonyl fluoride (PMSF), and ethylene diamine tetra acetic acid (EDTA). The results revealed that the protease was serine alkaline type. Tween 20 and Tween 80 did not inhibit the enzyme, however, sodium dodecyl sulfate (SDS), reduced it by 39%. It completely removed blood stain in 20 min and coagulated milk in the presence of CaCl2.

High-Temperature Corrosion Behavior of SUS310 Steels Embedded in Alkali Metal Chloride Salt Mixtures in Air

Higher steam temperature is one of the crucial requirements to increase the efficiency of waste-to-energy (WTE) plants. However, current steam temperature in the WTE plants, up to 450°C, is much lower than that in fossil-fuel power plants which are commonly used up to 650°C. Superheater tubes used in waste incineration plants are exposed in extremely corrosive environments, which contains Cl2, SO2, O2, H2O, CO2 etc. The combustion ash containing not only oxides but also chloride salts and sulfate accelerates the corrosion of alloys and coatings. This is one of the main reasons hindering increase of steam temperature in WTE plants. High-temperature corrosion in such harsh environments are so complicated, and it strongly depends on the plants because waste components varies in different areas. In order to understand a fundamental aspect of the corrosion behavior of heat resistant steels, the corrosion behavior of commercial steels as well as austenitic Fe-20Ni-25Cr based model alloys under an embedding in the simulated ashes at different temperatures lower than the melting temperature of the ashes was investigated.The commercial SUS310STB (base composition Fe-20Ni-25Cr-1Si), QSX5(Fe-20Ni-25Cr-1Mo-3Si), and model alloys of Fe-20Ni-25Cr with and without 2Mo or 2Si (in wt%) were used in this study. The specimens were embedded in the simulated ashes, which composed of CaO-4.5NaCl-4.5wt%KCl or CaO-3.1NaCl-3.96KCl-2.94CaCl2, in an Al2O3 crucible. Corrosion tests were conducted by a box furnace in air for up to 100 h at temperatures ranging from 400 to 580°C. Fig. 1 shows the corrosion mass loss of steels and model alloys as a function of corrosion temperature. In this figure, the results in the ash obtained from the actual plant are also plotted for comparison. Both of simulated ashes are less corrosive comparing to the actual ash and the corrosion mass loss of SUS310STB was much higher than that of QSX5. Based on the corrosion behavior of model alloys, higher corrosion resistance of QSX5 can be attributed to higher Si content in the alloy.The corrosion mass loss in the simulated ashes was low up to about 480°C and it stated to increase at 520°C in both of the simulated ashes, however, it was much lower in CaO-NaCl-KCl than in CaO-NaCl-KCl-CaCl2. The corrosion mass loss in CaO-NaCl-KCl was apparently increased at 580°C, which indicates that the presence of CaCl2 in ash accelerates corrosion. When steels were corroded at 580°C, the corrosion mass loss in CaO-NaCl-KCl-CaCl2 was significantly increased because the applied temperature is higher than the melting point of the ash. Cross-sectional observation of the steels after corrosion test at 580°C in CaO-NaCl-KCl revealed that the alloys formed a sponge-like very porous Fe-oxide scale. Above the sponge-like Fe-oxide scale, a thick CaCrO4 layer was developed. Moreover, K2CrO4 was identified in the corrosion products from EPMA and XRD analysis, suggesting that KCl reacted with a Cr2O3 scale, which resulted in a breakdown of the Cr2O3 scale and generation of chlorine. Because CaCrO4 is considered to be formed by a reaction of Cr-chloride vapor and CaO in the ash, higher evaporation of Cr-chloride might increase the metal consumption at higher temperatures. In the CaCl2 containing ash, the chlorine potential in the atmosphere increases due to oxidation of CaCl2, which might result in further increase in corrosion mass loss even though steels were exposed at the temperatures below melting temperature of the salt. Figure 1

A shell matrix protein of Pinctada mazatlanica produces nacre platelets in vitro

Nacre is the main component of the pearl oyster shells and it is synthesized by specialized soluble and insoluble shell matrix proteins. Insoluble proteins from the decalcification of the shell are the less studied proteins due to the technical problems to isolate them from the organic matrix. In this study, an insoluble shell matrix protein from Pinctada mazatlanica, pearlin (Pmaz-pearlin), was successfully cloned from the mantle tissue, and the native protein isolated from the shell was functionally characterized. The full coding sequence of Pmaz-pearlin mRNA consists of 423 base pairs, which encode to a 16.3 kDa pearlin. Analysis of the deduced amino acid sequence revealed that Pmaz-pearlin contained four acidic regions, an NG repeat domain, and Cys conserved residues, the latter potentially forms four disulfide bridges which might stabilize the protein structure. The isolated protein from the shell is a glycoprotein of ~ 16.74 kDa which can produce aragonite and calcite crystals in vitro. Our results show that Pmaz-pearlin is a well-conserved protein involved in nacre layer growth, which produces calcite crystals in the presence of CaCl2, aragonite crystal polymorphs with a hexagonal structure in the presence of MgCl2, and needle-like crystal structure polymorphs in the presence of CaCO3 The identity of the crystals was confirmed using RAMAN analyses.

Transcription Factor RUNX1 Regulates Factor FXIIIA Subunit (F13A1) Expression in Megakaryocytic Cells and Platelet F13A1 Expression is Downregulated in RUNX1 Haplodeficiency

Plasma Factor XIII, a transglutaminase, is a complex of two A (gene F13A1) chains with catalytic activity and two B chains (gene F13B). FXIII induces clot stabilization and retraction via cross-linking of fibrin monomers and other proteins. Factor XIII deficiency in plasma causes a major bleeding disorder with impaired clot retraction. FXIIIA is synthesized in megakaryocytes, and is abundant in platelets. It is expressed on activated platelets and cross-links proteins on platelet surface. Little is known regarding the regulation of F13A1 in megakaryocytes and platelets. RUNX1 is a major hematopoietic transcription factor and regulates expression of numerous genes in megakaryocytes and platelets. Patients with RUNX1 haplodeficiency (RHD) with heterozygous mutations have thrombocytopenia, platelet dysfunction with impaired responses to activation, and predisposition to myeloid malignancies. Platelet expression profiling using Affymetrix microarrays of a patient (P1) with a heterozygous RUNX1 mutation (c.969-323G&amp;gt;T) revealed decreased F13A expression, which encodes for subunit A of FXIII: fold-change 0.30; p=0.006; patient profiled twice 10 months apart and compared to 6 healthy subjects. These findings were confirmed by qPCR With RT-PCR platelet F13A mRNA expression was decreased to ~ 20% of mean expression in 4 subjects. Platelet F13A protein expression was decreased by ~50% as assessed by corrected total cellular fluorescence imaging. Platelet F13A mRNA expression was also decreased (to less than 20% of expression in 5 controls) in two siblings (P2 and P3 ages 8 and 3 years) from an unrelated family with a heterozygous RUNX1 mutation 508+1G&amp;gt;A). Plasma FXIII was normal in the patient P1. We studied the regulation of F13A by RUNX1 in human erythroleukemia (HEL) cells treated with phorbol myristate acetate (PMA) to induce megakaryocytic transformation. In silico analyses of the 5' upstream region of F13A showed 7 RUNX1 consensus sites within 545bp from ATG. Chromatin immunoprecipitation studies performed using HEL cells and RUNX1 antibody showed that that RUNX1 binds in the region encompassing site 4 and sites 5-7. In parallel, electrophoretic mobility shift assay (EMSA) studies showed binding of nuclear protein to labeled DNA probes with sites 1, 4, 5, 6, and 7. RUNX1 antibody produced a "supershift" with addition suggesting RUNX1 binding. Site-directed mutagenesis of the RUNX1 binding sites in F13A1 promoter decreased promoter activity in luciferase reporter studies in HEL cells, providing evidence that the binding sites are functional. Further, overexpression of RUNX1 in HEL cells increased F13A1 promoter activity and protein while siRNA RUNX1 knockdown reduced F13A1 protein. We assessed clot retraction over 90 min at 37oC in the patient P1 and his daughter (also with RUNX1 mutation) in citrated whole blood following addition of tissue factor (1 pM) and CaCl2 (1 mM), and in washed platelets resuspended in platelet poor plasma from a healthy subject supplemented with added fibrinogen (0.5 mg/mL) and stimulated with 0.5 U/mL thrombin in the presence of CaCl2. These preliminary studies suggested that clot retraction was slower in the patient P1 compared to the controls, particularly in studies with platelet suspensions. Conclusions: Overall, these studies provide the first evidence that hematopoietic transcription factor RUNX1 regulates expression of F13A1 in megakaryocytic cells and that platelet expression of F13A1 is decreased in platelets from patients with RUNX1 haplodeficiency. The decreased expression of F13A1 in RHD may contribute to the platelet dysfunction in RHD. These findings are particularly interesting because they reflect regulation of a coagulation protein (FXIIIA) by hematopoietic transcription factor RUNX1. Disclosures Lambert: Dova: Consultancy, Membership on an entity's Board of Directors or advisory committees; Principia: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Shionogi: Consultancy; Educational Concepts in Medicine: Consultancy; Octapharma: Consultancy, Research Funding; Bayer: Consultancy; Platelet Disorder Support Association (PDSA): Consultancy; 22qSociety: Consultancy; ITP Australia: Consultancy; CdLS Foundation: Consultancy; RDMD ITP study: Consultancy; Sysmex: Research Funding; AstraZeneca: Research Funding; Argenix: Consultancy; ClinGen: Honoraria.

Bio-inspired biopolymeric coacervation for entrapment and targeted release of anthocyanin

In order to provide stability of anthocyanin, a powerful antioxidant, encapsulant was made by utilizing two biopolymers that can be used further for the targeted release due to its intrinsic structure. Present study was carried out to encapsulate anthocyanin in coacervate employing pectin and chitosan. Entrapment efficiency of anthocyanin was evaluated in coacervates prepared in nine different combinations employing three concentration each of pectin (40, 50 and 60 mg mL−1) and chitosan (5, 6 and 7 mg mL−1) in presence of CaCl2. The developed formulation was characterised by FT-IR, XRD and SEM techniques, which clearly depicts ionic interaction between chitosan and pectin. Release pattern of anthocyanin from the coacervate followed Korsmeyer-Peppas model. The n-values calculated from the curves suggested a quasi-Fickian diffusion pattern pH 1.7 and 4.0. However, at pH 7.0, the behaviour of release was presumed to be an anomalous or case-II transport. In-vitro study revealed that bioaccessibility (%) of coacervated formulation (24.3 ± 0.5) was significantly higher than crude extract (12.2 ± 1.0) as well as purified anthocyanin crystals (18.9 ± 0.7). Better bioaccessibility of coacervated anthocyanin provided important information about its potential use in functional foods.

Co-immobilization of amylases in porous crosslinked gelatin matrices by different reticulations approaches

The aim of this study was to carry out the co-immobilization of α-amylase and glucoamylase in crosslinked gelatin porous supports. For this, two methods of co-immobilization were proposed based on the crosslinking with glutaraldehyde (Ggta) or CaCl2 in presence of alginate (Gcal). The supports characterization revealed a porous microstructure with good interaction between its components according to the FTIR analysis and thermal properties. Optimal pH and temperature of the Gcal co-immobilized enzymes were determined at 60 °C and pH 6.0, present an enzymatic activity of 120 μmol·mL·min−1. Moreover, both supports were reused for up to 8 hydrolysis cycles. In addition, co-immobilized enzymes were more efficient than free enzymes in starch saccharification of starch in the long term. These results reveal that the co-immobilization of amylases in gelatinous supports is a promising approach in enzymatic chain reactions.