scholarly journals Influence of Salts on the Adsorption of Lysozyme on a Mixed-Mode Resin

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Jannette Kreusser ◽  
Fabian Jirasek ◽  
Hans Hasse
Keyword(s):  
Sodium Chloride ◽  
Ionic Strength ◽  
Ammonium Sulfate ◽  
Protein Separation ◽  
Mixed Mode ◽  
Hydrophobic Interaction Chromatography ◽  
Exchange Chromatography ◽  
Separation And Purification ◽  
Adsorption Equilibria ◽  
Wide Range

Mixed-mode chromatography (MMC), which combines features of ion exchange chromatography (IEC) and hydrophobic interaction chromatography (HIC), is an interesting method for protein separation and purification. The design of MMC processes is challenging as adsorption equilibria are influenced by many parameters, including ionic strength and the presence of different salts in solution. Systematic studies on the influence of those parameters in MMC are rare. Therefore, in the present work, the influence of four salts, namely, sodium chloride, sodium sulfate, ammonium chloride, and ammonium sulfate, on the adsorption of lysozyme on the mixed-mode resin Toyopearl MX-Trp-650M at pH 7.0 and 25°C was studied systematically in equilibrium adsorption experiments for ionic strengths between 0 mM and 3000 mM. For all salts, a noticeable adsorption strength was observed over the entire range of studied ionic strengths. An exponential decay of the loading of the resin with increasing ionic strength was found until approx. 1000 mM. For higher ionic strengths, the loading was found to be practically independent of the ionic strength. At constant ionic strength, the highest lysozyme loadings were observed for ammonium sulfate, the lowest for sodium chloride. A mathematical model was developed that correctly describes the influence of the ionic strength as well as the influence of the studied salts. The model is the first that enables the prediction of adsorption isotherms of proteins on mixed-mode resins in a wide range of technically interesting conditions, accounting for the influence of the ionic strength and four salts of practical relevance.

scholarly journals Destruction of Polyelectrolyte Microcapsules Formed on CaCO3 Microparticles and the Release of a Protein Included by the Adsorption Method

Polymers ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 520
Author(s):  
Egor V. Musin ◽  
Aleksandr L. Kim ◽  
Sergey A. Tikhonenko
Keyword(s):  
Sodium Chloride ◽  
Ionic Strength ◽  
Ammonium Sulfate ◽  
Fluorescein Isothiocyanate ◽  
Total Protein Content ◽  
High Concentration ◽  
Adsorption Method ◽  
Polyelectrolyte Microcapsules ◽  
High Level ◽  
Increasing Temperature

The degradation of polyelectrolyte microcapsules formed on protein-free CaCO3 particles consisting of polyallylamine (PAH) and polystyrene sulfonate (PSS) and the resulting yield of protein in the presence of various salts of different concentrations, as well as at two pH values, was studied by fluorescence spectroscopy; the protein was incorporated into prepared microcapsules by adsorption. It was found that a high concentration of sodium chloride (2 M) leads to considerable dissociation of PAH, which is apparently due to the loosening of polyelectrolytes under the action of ionic strength. At the same time, 0.2 M sodium chloride and ammonium sulfate of the same ionic strength (0.1 M) exert less influence on the amount of dissociated polymer. In the case of ammonium sulfate (0.1 M), the effect is due to the competitive binding of sulfate anions to the amino groups of the polyelectrolyte. However, unlike microcapsules formed on CaCO3 particles containing protein, the dissociation of polyelectrolyte from microcapsules formed on protein-free particles increased with increasing temperature. Apparently, a similar effect is associated with the absence of a distinct shell, which was observed on microcapsules formed on protein-containing CaCO3 particles. The high level of the presence of Fluorescein isothiocyanate (FITC)-labeled Bovine Serum Albumin (BSA) in the supernatant is explained by the large amount of electrostatically bound protein and the absence of a shell that prevents the release of the protein from the microcapsules. In 2 M NaCl, during the observation period, the amount of the released protein did not exceed 70% of the total protein content in the capsules, in control samples, this value does not exceed 8%, which indicates the predominantly electrostatic nature of protein retention in capsules formed on protein-free CaCO3 particles. The increase in protein yield and peeling of PAH with increasing pH is explained by the proximity of pH 7 to the point of charge exchange of the amino group of polyelectrolyte, as a result of the dissociation of the microcapsule.

Review on the Application of Mixed-mode Chromatography for Separation of Structure Isoforms

2018 ◽  
Vol 20 (1) ◽  
pp. 56-60 ◽  
Author(s):  
Tsutomu Arakawa
Keyword(s):  
Ion Exchange ◽  
Mixed Mode ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Charged State ◽  
Partial Structure ◽  
Reverse Phase ◽  
Oligomer Formation ◽  
Structure Rearrangement ◽  
Disulfide Scrambling

Proteins often generate structure isoforms naturally or artificially due to, for example, different glycosylation, disulfide scrambling, partial structure rearrangement, oligomer formation or chemical modification. The isoform formations are normally accompanied by alterations in charged state or hydrophobicity. Thus, isoforms can be fractionated by reverse-phase, hydrophobic interaction or ion exchange chromatography. We have applied mixed-mode chromatography for fractionation of isoforms for several model proteins and observed that cation exchange Capto MMC and anion exchange Capto adhere columns are effective in separating conformational isoforms and self-associated oligomers.

scholarly journals Impact of the particle mixing state on the hygroscopicity of internally mixed sodium chloride-ammonium sulfate single droplets: A theoretical and experimental study

2021 ◽  
Author(s):  
Yeny A. Tobon ◽  
Danielle El Hajj ◽  
Samantha Seng ◽  
Ferdaous Bengrad ◽  
Myriam Moreau ◽  
...  
Keyword(s):  
Experimental Study ◽  
Sodium Chloride ◽  
Ammonium Sulfate ◽  
Atmospheric Transport ◽  
Sea Salt ◽  
Main Constituent ◽  
Sea Salt Aerosols ◽  
Particle Mixing ◽  
Secondary Aerosols ◽  
Single Droplets

Sodium chloride (NaCl) is the main constituent of sea-salt aerosols. During atmospheric transport, sea-salt aerosols can interact with gases and other particles including secondary aerosols containing ammonium sulfate ((NH4)2SO4). This...

scholarly journals Intracellular Ionic Strength Sensing Using NanoLuc

2021 ◽  
Vol 22 (2) ◽  
pp. 677
Author(s):  
Tausif Altamash ◽  
Wesam Ahmed ◽  
Saad Rasool ◽  
Kabir H. Biswas
Keyword(s):  
Thermal Stability ◽  
Phase Separation ◽  
Drug Discovery ◽  
Ionic Strength ◽  
Cellular Signaling ◽  
Live Cells ◽  
Protein Activity ◽  
Cellular Survival ◽  
Cellular Processes ◽  
Wide Range

Intracellular ionic strength regulates myriad cellular processes that are fundamental to cellular survival and proliferation, including protein activity, aggregation, phase separation, and cell volume. It could be altered by changes in the activity of cellular signaling pathways, such as those that impact the activity of membrane-localized ion channels or by alterations in the microenvironmental osmolarity. Therefore, there is a demand for the development of sensitive tools for real-time monitoring of intracellular ionic strength. Here, we developed a bioluminescence-based intracellular ionic strength sensing strategy using the Nano Luciferase (NanoLuc) protein that has gained tremendous utility due to its high, long-lived bioluminescence output and thermal stability. Biochemical experiments using a recombinantly purified protein showed that NanoLuc bioluminescence is dependent on the ionic strength of the reaction buffer for a wide range of ionic strength conditions. Importantly, the decrease in the NanoLuc activity observed at higher ionic strengths could be reversed by decreasing the ionic strength of the reaction, thus making it suitable for sensing intracellular ionic strength alterations. Finally, we used an mNeonGreen–NanoLuc fusion protein to successfully monitor ionic strength alterations in a ratiometric manner through independent fluorescence and bioluminescence measurements in cell lysates and live cells. We envisage that the biosensing strategy developed here for detecting alterations in intracellular ionic strength will be applicable in a wide range of experiments, including high throughput cellular signaling, ion channel functional genomics, and drug discovery.

scholarly journals A novel tandem differential mobility analyzer with organic vapor treatment of aerosol particles

2001 ◽  
Vol 1 (1) ◽  
pp. 51-60 ◽  
Author(s):  
J. Joutsensaari ◽  
P. Vaattovaara ◽  
M. Vesterinen ◽  
K. Hämeri ◽  
A. Laaksonen
Keyword(s):  
Water Vapor ◽  
Sodium Chloride ◽  
Citric Acid ◽  
Ammonium Sulfate ◽  
Adipic Acid ◽  
Aerosol Particles ◽  
Ethanol Vapor ◽  
Differential Mobility Analyzer ◽  
Differential Mobility ◽  
Organic Vapor

Abstract. A novel method to characterize the organic composition of aerosol particles has been developed. The method is based on organic vapor interaction with aerosol particles and it has been named an Organic Tandem Differential Mobility Analyzer (OTDMA). The OTDMA method has been tested for inorganic (sodium chloride and ammonium sulfate) and organic (citric acid and adipic acid) particles. Growth curves of the particles have been measured in ethanol vapor and as a comparison in water vapor as a function of saturation ratio. Measurements in water vapor show that sodium chloride and ammonium sulfate as well as citric acid particles grow at water saturation ratios (S) of 0.8 and above, whereas adipic acid particles do not grow at S <  0.96. For sodium chloride and ammonium sulfate particles, a deliquescence point is observed at S = 0.75 and S = 0.79, respectively. Citric acid particles grow monotonously with increasing saturation ratios already at low saturation ratios and no clear deliquescence point is found. For sodium chloride and ammonium sulfate particles, no growth can be seen in ethanol vapor at saturation ratios below 0.93. In contrast, for adipic acid particles, the deliquescence takes place at around S = 0.95 in the ethanol vapor. The recrystallization of adipic acid takes place at S < 0.4. Citric acid particles grow in ethanol vapor similarly as in water vapor; the particles grow monotonously with increasing saturation ratios and no stepwise deliquescence is observed. The results show that the working principles of the OTDMA are operational for single-component aerosols. Furthermore, the results indicate that the OTDMA method may prove useful in determining whether aerosol particles contain organic substances, especially if the OTDMA is operated in parallel with a hygroscopicity TDMA, as the growth of many substances is different in ethanol and water vapors.

Seed germination biology of sweet acacia (Vachellia farnesiana) and response of its seedlings to herbicides

Weed Science ◽  
2021 ◽  
pp. 1-19
Author(s):  
Bhagirath S. Chauhan ◽  
Shane Campbell ◽  
Victor J. Galea
Keyword(s):  
Sodium Chloride ◽  
Seed Germination ◽  
Development Stage ◽  
Hot Water ◽  
Control Treatment ◽  
Similar Species ◽  
Weed Species ◽  
Physical Dormancy ◽  
Seed Biology ◽  
Wide Range

Abstract Sweet acacia [Vachellia farnesiana (L.) Willd.]is a problematic thorny weed species in several parts of Australia. Knowledge of its seed biology could help to formulate weed management decisions for this and other similar species. Experiments were conducted to determine the effect of hot water (scarification), alternating temperatures, light, salt stress, and water stress on seed germination of two populations of V. farnesiana and to evaluate the response of its young seedlings (the most sensitive development stage) to commonly available POST herbicides in Australia. Both populations behaved similarly to all the environmental factors and herbicides; therefore, data were pooled over the populations. Seeds immersed in hot water at 90 C for 10 min provided the highest germination (88%), demonstrating physical dormancy in this species. Seeds germinated at a wide range of alternating day/night temperatures from 20/10 C (35%) to 35/25 C (90%) but no seeds germinated at 15/5 C. Germination was not affected by light, suggesting that seeds are nonphotoblastic and can germinate under a plant canopy or when buried in soil. Germination was not affected by sodium chloride concentrations up to 20 mM and about 50% of seeds could germinate at 160 mM sodium chloride, suggesting its high salt tolerance ability. Germination was only 13% at −0.2 MPa osmotic potential and no seeds germinated at −0.4 MPa, suggesting that V. farnesiana seeds may remain ungerminated until moisture conditions have become conducive for germination. A number of POST herbicides, including 2,4-D + picloram, glufosinate, paraquat and saflufenacil, provided >85% control of biomass of young seedlings compared with the nontreated control treatment. Knowledge gained from this study will help to predict the potential spread of V. farnesiana in other areas and help to integrate herbicide use with other management strategies.

Purification and Characterization of Laccase fromChaetomium thermophilium and Its Role in Humification

1998 ◽  
Vol 64 (9) ◽  
pp. 3175-3179 ◽  
Author(s):  
Benny Chefetz ◽  
Yona Chen ◽  
Yitzhak Hadar
Keyword(s):  
Municipal Solid Waste ◽  
Solid Waste ◽  
Laccase Activity ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Water Soluble ◽  
Type I ◽  
Light Spectrum ◽  
Solid Media ◽  
Wide Range

ABSTRACT Chaetomium thermophilium was isolated from composting municipal solid waste during the thermophilic stage of the process.C. thermophilium, a cellulolytic fungus, exhibited laccase activity when it was grown at 45°C both in solid media and in liquid media. Laccase activity reached a peak after 24 h in liquid shake culture. Laccase was purified by ultrafiltration, anion-exchange chromatography, and affinity chromatography. The purified enzyme was identified as a glycoprotein with a molecular mass of 77 kDa and an isoelectric point of 5.1. The laccase was stable for 1 h at 70°C and had half-lives of 24 and 12 h at 40 and 50°C, respectively. The enzyme was stable at pH 5 to 10, and the optimum pH for enzyme activity was 6. The purified laccase efficiently catalyzed a wide range of phenolic substrates but not tyrosine. The highest levels of affinity were the levels of affinity to syringaldazine and hydroxyquinone. The UV-visible light spectrum of the purified laccase had a peak at 604 nm (i.e., Cu type I), and the activity was strongly inhibited by Cu-chelating agents. When the hydrophobic acid fraction (the humic fraction of the water-soluble organic matter obtained from municipal solid waste compost) was added to a reaction assay mixture containing laccase and guaiacol, polymerization took place and a soluble polymer was formed. C. thermophilium laccase, which is produced during the thermophilic stage of composting, can remain active for a long period of time at high temperatures and alkaline pH values, and we suggest that this enzyme is involved in the humification process during composting.

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