Impact Of Protein Concentration Research Articles

The role of protein concentration in heat‐induced particulation of soy proteins at different pHs: Structure and functional properties

AbstractProtein particulation is a modification strategy for the optimization of the use of protein materials. The development and subtypes of particulate structures are largely dependent on the aggregated state of proteins after heat‐induced interactions, which is profoundly influenced by protein concentration (PC). In this work, the impact of PC below and above the critical gelation point, that is, 5% (w/v) and 10% (w/v), on the structure and functional properties of heat‐treated soy protein isolates (SPIs) at different pHs (2.0, 4.0, and 7.0), was investigated. The results showed that heat‐induced aggregation of SPIs was promoted by increasing the PC, leading to a β‐sheet‐dominated secondary structure. At pH 2.0 and 7.0, the 10% SPIs exhibited larger particle size and lower solubility, surface hydrophobicity index, and oil‐holding capacity compared to the 5% SPIs after heating. Furthermore, at neutral pH, the 10% SPI microgels had higher storage modulus (G′) and loss modulus (G″) than their acidifying counterparts, as well as the excellent emulsifying property for oil droplet stabilization. These findings would provide the theoretical basis for the structure modification and function improvement of plant proteins and, therefore, broaden the application of plant proteins in the food industry.

Surface and foaming properties of potato proteins: Impact of protein concentration, pH value and ionic strength

Using a multiscale approach, potato protein isolate was characterized holistically in terms of its solubility and surface charge, surface activity and surface dilatational properties as well foaming properties and bubble structural attributes. By means of varying protein concentration (0.1–10.0%), pH (3.0–10.0) and NaCl concentration (0–200 mM), it was aimed to establish a better mechanistic understanding of potato protein functionality. Overall, surface activity was found to be rather unaffected by a modification of protein concentration and pH, whereas pH had a clear effect on surface dilatational elasticity. Likewise, foam stability was affected by pH, yielding a maximum around the isoelectric point, which is due to an enhanced network formation at the air/water interface imparting an increased surface film stability. Coarsening exponents were found to reflect the results on foam stability. By comparison, NaCl presence clearly had an enhancing effect on surface activity and led to increased protein-protein interactions within the surface film. More compact surface films in turn resulted in a better foamability and foam stability as well as lower coarsening exponents. Comparison of findings to mechanisms known for β-lactoglobulin-stabilized surfaces revealed their principal applicability also to potato proteins’ functional properties. Overall, by means of the obtained knowledge, foam structures based on potato proteins can be controlled more specifically.

Pharmacokinetics, microscale distribution, and dosimetry of alpha-emitter-labeled anti-PD-L1 antibodies in an immune competent transgenic breast cancer model

BackgroundStudies combining immune checkpoint inhibitors with external beam radiation have shown a therapeutic advantage over each modality alone. The purpose of these works is to evaluate the potential of targeted delivery of high LET radiation to the tumor microenvironment via an immune checkpoint inhibitor.MethodsThe impact of protein concentration on the distribution of 111In-DTPA-anti-PD-L1-BC, an 111In-antibody conjugate targeted to PD-L1, was evaluated in an immunocompetent mouse model of breast cancer. 225Ac-DOTA-anti-PD-L1-BC was evaluated by both macroscale (ex vivo biodistribution) and microscale (alpha-camera images at a protein concentration determined by the 111In data.ResultsThe evaluation of 111In-DTPA-anti-PD-L1-BC at 1, 3, and 10 mg/kg highlighted the impact of protein concentration on the distribution of the labeled antibody, particularly in the blood, spleen, thymus, and tumor. Alpha-camera images for the microscale distribution of 225Ac-DOTA-anti-PD-L1-BC showed a uniform distribution in the liver while highly non-uniform distributions were obtained in the thymus, spleen, kidney, and tumor. At an antibody dose of 3 mg/kg, the liver was dose-limiting with an absorbed dose of 738 mGy/kBq; based upon blood activity concentration measurements, the marrow absorbed dose was 29 mGy/kBq.ConclusionsThese studies demonstrate that 225Ac-DOTA-anti-PD-L1-BC is capable of delivering high LET radiation to PD-L1 tumors. The use of a surrogate SPECT agent, 111In-DTPA-anti-PD-L1-BC, is beneficial in optimizing the dose delivered to the tumor sites. Furthermore, an accounting of the microscale distribution of the antibody in preclinical studies was essential to the proper interpretation of organ absorbed doses and their likely relation to biologic effect.

Abstract LB-183: The impact of protein concentration on the distribution of 111In-antibody conjugate for imaging of programmed death-ligand 1 (PD-L1)

Abstract Background: Programmed cell Death Ligand 1 (PD-L1) is part of an immune checkpoint system that is essential for preventing autoimmunity. Tumor cells have developed the ability to co-opt these immune checkpoints to suppress anti-tumor immunity. Recent approaches in immunotherapy targeting immune checkpoints have shown great promise in a variety of cancers. PD-L1 overexpression is associated with a poorer prognosis in a variety of cancers, but typically have a stronger response to anti-PD-L1 therapy. PD-L1 is a dynamic biomarker and the ability to image its dynamic nature could provide key information in identifying patients who will best respond to anti-PD-L1 treatment with limited adverse effects. Here we evaluated an 111In-antibody conjugate targeted to PD-L1 for SPECT imaging in an aggressive mouse melanoma model. In addition, we investigated the relationship between the spleen, which we have found to be a sink for anti-PD-L1 antibodies, and specific activity (activity/protein concentration) of the 111In-antibody conjugate. Methods: An anti-PD-L1 antibody was conjugated to DTPA for 111In-labeling. The biodistribution of the resulting conjugate, 111In-DTPA-anti-PD-L1, was performed over a range of specific activities in B16F10 tumor bearing mice. SPECT imaging was also performed with the 111In tracer. Results: 111In-DTPA-anti-PD-L1 (Kd = 0.6±0.1 nM) having a specific activity (SA) of 0.57 Ci/μmol demonstrated uptake in the B16F10 tumor (6.1±2.8%ID/g) at 24 hours, as well as high uptake in the spleen (32.8±5.1%ID/g). Decreasing the SA to 0.01 Ci/μmol blocked PD-L1-postive cells in the spleen (13.4±4.8%ID/g) forcing more of the tracer to remain in circulation, allowing for increase accumulation in the B16F10 tumor (16.8±6.2%ID/g) at 24 hours. The lower SA also demonstrated a reduction in tumor to blood ratios compared to the higher SA (2.2 vs. 4.1), but an increase was shown in the tumor to muscle ratio (27 vs. 4.6). Conclusion: 111In-DTPA-anti-PD-L1 is capable of identifying tumors that overexpresses PD-L1, and has the potential to help select patients that will best respond to anti-PD-L1 therapy. In addition, the further understanding of the spleen/tumor relationship could be utilized to tailor individual doses for anti-PD-L1 therapy and minimize adverse effects. Citation Format: Jessie Nedrow, Anders Josefsson, Sunju Park, Sagar Ranka, George Sgouros. The impact of protein concentration on the distribution of 111In-antibody conjugate for imaging of programmed death-ligand 1 (PD-L1). [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-183.

Sterile Filtration of Highly Concentrated Protein Formulations: Impact of Protein Concentration, Formulation Composition, and Filter Material

Differences in filtration behavior of concentrated protein formulations were observed during aseptic drug product manufacturing of biologics dependent on formulation composition. The present study investigates filtration forces of monoclonal antibody formulations in a small-scale set-up using polyvinylidene difluoride (PVDF) or polyethersulfone (PES) filters. Different factors like formulation composition and protein concentration related to differences in viscosity, as well as different filtration rates were evaluated. The present study showed that filtration behavior was influenced by the presence or absence of a surfactant in the formulation, which defines the interaction between filter membrane and surface active formulation components. This can lead to a change in filter resistance (PES filter) independent on the buffer system used. Filtration behavior was additionally defined by rheological non-Newtonian flow behavior. The data showed that high shear rates resulting from small pore sizes and filtration pressure up to 1.0 bar led to shear-thinning behavior for highly concentrated protein formulations. Differences in non-Newtonian behavior were attributed to ionic strength related to differences in repulsive and attractive interactions. The present study showed that the interplay of formulation composition, filter material, and filtration rate can explain differences in filtration behavior/filtration flux observed for highly concentrated protein formulations thus guiding filter selection.

The Impact of Protein Concentration on Mannitol and Sodium Chloride Crystallinity and Polymorphism Upon Lyophilization

The effect of protein concentration, in the range of 0–26 mg/mL for two Fc-fusion proteins, on the crystallinity and polymorphism of mannitol and sodium chloride in a lyophilized model formulation was examined. Mannitol hydrate levels were quantified based on moisture data and correlated to the X-ray diffraction peak area. In all formulation conditions, sodium chloride did not crystallize in samples with >44% total amorphous content. As protein concentration increased through the range of 1–5 mg/mL prior to lyophilization, β-mannitol decreased in amount, becoming undetectable at protein concentrations above 5 mg/mL. Conversely, δ-mannitol increased as a function of protein concentration, reaching a maximum level at ∼5 mg/mL protein. Above 10 mg/mL protein, mannitol crystallization was increasingly inhibited. Sucrose control vials showed higher levels of mannitol hydrate than either model protein. Both proteins behaved comparably with respect to mannitol crystallinity and polymorphism despite significant differences in molecular weight. Because of the differences between protein and sucrose control samples, protein concentration must be taken into consideration when assessing the lyophilization of mannitol containing solutions. © 2008 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci

Production of biological nanoparticles from bovine serum albumin for drug delivery

Bovine serum albumin (BSA) was used for generation of nanoparticles in a drug delivery system. The size of the fabricated nano-particles was measure by laser light scanning. Several process parameters were examined to achieve a suitable size of nanoparticle such as pH, temperature, BSA concentration, agitation speed, glutaraldehyde concentration, organic solvent adding rate and the ratio of BSA/organic solvent. The smallest size of nanoparticles achieved, was 101 nm and the largest size was 503 nm. The most effective parameters for the fabrication of the nanoparticles were the agitation speed and the media temperature. The minimum size of nanoparticles at the desired incubator of 4°C and constant agitation rate of 300 - 400 rpm was obtained. The impact of protein concentration and additional rate of organic solvent (i.e. ethanol) upon the particle size was investigated. The protein concentration of 5-40 mg.ml-1 was resulted; the main effect on the particle size and minimum mean size diameter gained was 30 mg.ml-1 protein concentration. The nanoparticle sample was purified with 50,000 g centrifuge then followed by dialysis, micro and ultrafiltration and then analyzed by SEM, PCS as well as SDS gel electrophoresis.

Microdialysis evaluation of the ocular pharmacokinetics of propranolol in the conscious rabbit.

This study was conducted to assess the effects of anesthesia and aqueous humor protein concentrations on ocular disposition of propranolol. Rabbits were anesthetized and a microdialysis probe was inserted into the anterior chamber of one eye; the contralateral eye served as a control. At timed intervals after probe placement, a 100-microl sample of aqueous humor was aspirated from each eye to determine protein concentration. In vitro protein binding parameters were used to simulate the impact of protein concentration on propranolol disposition. To assess the influence of anesthesia, probes were implanted in the anterior chamber of each eye. After >5-day stabilization, conscious and anesthetized rabbits (n = 3/group) received a 200-microg topical dose of [3H]DL-propranolol in each eye; propranolol was assayed in probe effluent. Changes in aqueous humor protein concentrations were observed following probe insertion. Simulations demonstrated that the unbound propranolol AUC (approximately 2.4-fold) in aqueous humor should be reduced due to protein influx. Intraocular propranolol exposure in anesthetized rabbits was approximately 8-fold higher than in conscious rabbits, and approximately 1.9-fold higher than in rabbits without a post-surgical recovery period. Anesthesia and time-dependent aqueous humor protein concentrations may alter ocular pharmacokinetics, and must be taken into account in the design of microdialysis experiments.