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Chitosan, being one of the most promising biological macromolecules, has an immense scope in agriculture to boost crop growth and defense responses. In this study, chitosan was exposed to gamma rays in order to obtain a low molecular weight derivative. Viscometric characterization showed a sharp decrease in molecular weight and FTIR based analysis confirmed retention of structural integrity of the polymer upon gamma irradiation. Assessments of various physiological and biochemical attributes were carried out on sugarcane plantlets that were subjected to progressive water deficit stress. The irradiated chitosan was found to differentially ameliorate water deficit stress tolerance against that of normal chitosan through positive modulation of various gas exchange parameters alongside significant improvement in relative tissue water content, SOD activity, soluble sugars and adenine energetics. Furthermore, application of irradiated chitosan significantly reduced cell membrane damage, lipid peroxidation, H2O2 and free-proline accumulations. This is the first report on the use of gamma irradiated chitosan to alleviate water deficit stress tolerance in sugarcane. Overall comparative assessments showed that differential plant responses were triggered upon foliar application of normal and gamma irradiated chitosan in sugarcane plants grown under water deficit stress conditions.

Solid-state is the preferred choice for storage of protein therapeutics to improve stability and preserve the biological activity by decreasing the physical and chemical degradation associated with liquid protein formulations. Lyophilization or freeze-drying is an effective drying method to overcome the instability problems of proteins. However, the processing steps (freezing, primary drying and secondary drying) involved in the lyophilization process can expose the proteins to various stress and harsh conditions, leading to denaturation, aggregation often a loss in activity of protein therapeutics. Stabilizers such as sugars and surfactants are often added to protect the proteins against physical stress associated with lyophilization process and storage conditions. Another way to curtail the degradation of proteins due to process related stress is by modification of the lyophilization process. Slow freezing, high nucleation temperature, decreasing the extent of supercooling, and annealing can minimize the formation of the interface (ice-water) by producing large ice crystals with less surface area, thereby preserving the native structure and stability of the proteins. Hence, a thorough understanding of formulation composition, lyophilization process parameters and the choice of analytical methods to characterize and monitor the protein instability is crucial for development of stable therapeutic protein products. This review provides an overview of various stress conditions that proteins might encounter during lyophilization process, mechanisms to improve the stability and analytical techniques to tackle the proteins instability during both freeze-drying and storage.

Research into stress and stability is essential during the development of therapeutic proteins to ensure quality and safety of the final medicine. Greater knowledge of the effects of stress on aggregation can help avoid undesirable conformational and colloidal instabilities. With this in mind we investigated five marketed therapeutic monoclonal antibodies (mAbs) namely bebacizumab (BVZ), cetuximab (CTX), infliximab (IFX), rituximab (RTX) and tratuzumab (TTZ) in their innovative medicines. These were submitted to different controlled stresses, to freeze/thaw cycles and used for long-term stability studies once the vials were opened. Aggregate formation was tracked by analyzing the mAbs chromatographic profiles by size-exclusion liquid chromatography coupled with diode array detection. Results indicated that the tendency to aggregate depends on the particular stress conditions and on the concentration and nature of the mAb, even though all share similar IgG1-structure. Fragmentation of the mAb produced by the stress was probably due to the rupture of cystines between the two heavy chains. Regarding stability study, BVZ, RTX and TTZ proved to be the most stable when stored at 4 °C and in freeze/thaw cycles with no tendency to form aggregates. INF tends to form aggregates at 0.5 mg/mL, while in CTX, the most unstable, degradation was detected.

Nature has selected osmolytes to protect intracellular macromolecules against denaturing stress conditions. These molecules are accumulated in the intracellular environment at considerably high concentrations. In general, osmolytes are known to stabilize proteins. Osmolytes are small naturally occurring compound that are utilized as chemical chaperones by changing the environmental condition in diseased cellular stress state. Osmolytes are present in different organisms, including microorganisms, animals and plants. In intracellular environment osmolytes are naturally accumulates at comparatively high concentrations while cells/tissues are exposed in stressful conditions. This is important because aggregation, misfolding and destabilization of the protein lead to several life-threatening neurodegenerative disorders. So, we have thrown light on some of the properties of the osmolyte which assist in protein folding, increases their stability and inhibit aggregation. Osmolytes are also known as chemical chaperones. Hence, it may be therapeutically used for several misfolding diseases. Intracellular osmolytes protects proteins against denaturing stresses and provides valuable insight into the fundamental problem of protein stabilization, a problem that plagues the pharmaceutical industry, biotechnology, and everyday research.