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RADA 16-I is an ionic self-assembling peptide that can form macroscopic scaffolds through β-sheet structures which are used in favor of cell growth and tissue engineering. This peptide has also the ability to stop bleeding effectively and quickly (∼20 seconds) when applied directly to the injuries. This study is focused on coagulation process, platelet aggregation, C3 and C4 concentrations, CBC counting, hemolysis, and white blood cell morphology tests to analyze hemocompatibility of RADA 16-I at different concentrations - 0.1, 0.2, 0.3 and 0.5%. According to the results, RADA 16-I hydrogel decreased the number of blood cells, slightly increased clot formation time and platelet aggregation, and yielded negligible hemolysis and only small changes in C3 and C4 concentrations and white blood cell morphology. All by all, the in vitro tests of hemocompatibility showed no perturbation in the blood composition when the peptides were in contact with the blood. The observed rapid hemostasis might be a result of increasing local concentrations of molecules involved in the formation of clot near the peptide hydrogel, thereby making a barrier which ended up with complete hemostasis. In conclusion, our experiments strongly supported further development of biomaterials based on RADA 16-I peptide.

This study investigated the effects of using a combination of substrates (fructose, propanol, citric acid, acetic acid, propionic acid and beef extract) on the synthesis of poly (3-hydroxybutyrate-co-3-hydroxyvalerate), which is also known as PHBV. The results of batch PHBV optimization by RSM, indicated that PHB was synthesized only when fructose was used as the sole carbon source. Fructose in combination with propanol and beef extract, exhibited better PHBV production performance, as compared to that of propionic acid which was used in previous studies. GC, (1)H and (13)C NMR spectroscopy revealed that the best copolymer composition was 54.1 mol% of 3-hydroxyvalerate (3HV) which was obtained in a basal medium combined with 0.3% (v/v) propanol and 0.0172% (w/v) beef extract. This is the first report on the production of PHBV with HV content of 54.1% by a wild-type Cupriavidus necator. The Fourier Transform Infrared Spectroscopy (FTIR) spectra of the PHAs indicated absorption bands at 1723, 1274, 1373, 1453, 2932cm(-1) corresponding to CO, CO stretching, CH3, -CH2 and -CH groups, respectively. This study showed that the recovered PHBV biopolymer from the optimized culture to have a melting temperature of 103.6°C, melting enthalpy of 55.67J/g and crystallization temperature of 76.1°C., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The low stability of recombinant human interferon-γ (rhIFN-γ) therapeutic protein imposes some restrictions in its medical applications. In the current study, the effect of oxygen tension on the stability of purified rhIFN-γ was investigated. The rhIFN-γ was purified (>99%) by a two-step chromatographic process. Storage vials were filled by purified formulated product under normal atmospheric oxygen and low oxygen tension conditions. At different time intervals, the amounts of rhIFN-γ covalent dimers and deamidated forms were analyzed using analytical high-performance liquid chromatography (HPLC; size exclusion and cation exchange) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) methods. To determine the biological activity of purified rhIFN-γ, an antiviral activity assay against vesicular stomatitis virus (VSV) was performed. Upon rhIFN-γ long-term storage in a low oxygen tension condition, the amounts of rhIFN-γ covalent dimers and deamidated forms and also the biological activity of rhIFN-γ changed a little. In contrast, by 9 months of storage of rhIFN-γ preparations under normal atmospheric condition, the amount of covalent dimers and deamidated forms increased with time and reached to approximately 3.5% and 11.5% of the initial amount, respectively. The antiviral specific activity of 9-month-old rhIFN-γ preparations decreased to 41% of the initial amount at normal storage condition, while no significant reduction was seen at the low oxygen tension condition. In conclusion, oxygen tension during storage could have a significant impact on rhIFN-γ stability and finally on the quality of pharmaceutical rhIFN-γ product.

Process development and optimization studies were performed in order to improve the purification process of (rhIFN-gamma). The objective was to generate material with higher purity and quantity. An in-process control screening was developed to obtain the optimal condition for column chromatographic purification by measuring LPS, nucleic acids, rhIFN- gamma, monomer and its covalent dimers. A new resin screening method was applied to select optimal resin for each of the chromatographic columns. The resulting process used Butyl and Q-Sepharose, refolding and SP-Sepharose for purification of IFN-gamma. Effects of different process conditions such as cell lysis, removal of impurity and oxygen concentration were evaluated. Removal of impurities was evaluated by washing of inclusion bodies with 1% Triton X-100 and 3M urea and different chromatography steps. The results reveal that Triton removed about 43% of the LPS but urea had no effect on removal of nucleic acids and LPS. Further analysis show that removal of impurities by column chromatography decreases aggregation and increases the process yield. Oxygen concentration was identified as parameter that could have a significant impact on covalent dimers formation, as an unacceptable pharmaceutical form of rhIFN-gamma. On the basis of small-scale studies, optimum operating conditions were chosen and the purification process was successfully scaled-up to a pilot scale process with step yield and product quality that were better than previous reports.