An Assessment of Biodegradability and Phytotoxicity of Natural Rubber in a Simulated Soil Condition via CO2 Evolution Measurement.

In this study, the biodegradation of various natural rubber (NR) samples, i.e., neat NR and NR filled with two different curative contents was investigated under a long-term simulated soil condition at a temperature of 25 ± 2 °C in accordance with ISO 17556. Natural clay loam soil, with a pH of 7.2 and a water holding capacity of 57.6%, was employed. Under controlled test condition both unvulcanized and vulcanized NR samples having low curative content, respectively designated as UNRL and VNRL, exhibited similar biodegradation behaviors to the neat NR. They showed fast biodegradation at the early stage, and their biodegradation rate did not significantly change throughout the test period (365 days). However, for the NR samples having high curative content, respectively called UNRH and VNRH for the unvulcanized and vulcanized samples, a biodegradation delay was observed within the first 130 days. Surprisingly, the UNRH showed a relatively high biodegradation rate after the induction period. At the end of the test, most of the rubber samples (the neat NR, UNRL, VNRL, and UNRH) showed a comparable degree of biodegradation, with a value ranging from 54-59%. The VNRH, on the other hand, showed the lowest degree of biodegradation (ca. 28%). The results indicate that the number of curatives does not significantly affect the biodegradability of unvulcanized NR in the long term, despite the fact that a high curative content might retard microorganism activity at the beginning of the biodegradation process. Apparently, crosslink density is one of the key factors governing the biodegradability of NR. The phytotoxicity of the soils after the biodegradation test was also assessed and represented in terms of seedling emergence, survival rate, and plant biomass for Sorghum bicolor. The values of seedling emergence (≥80%), survival rate (100%), and plant biomass of all soil samples were not statistically different from those of the blank soil, indicating the low phytotoxicity of the tested soils subjected to the biodegradation of the rubber samples. Taken as a whole, it can be concluded that the CO2 measurement technique is one of the most effective methods to assess the biodegradability of rubbers. The knowledge obtained from this study can also be applied to formulate more environmentally friendly rubber products.

Oxidized carbon nanoparticles as an effective protein antigen delivery system targeting the cell-mediated immune response.

Background: The demand for an effective vaccine delivery system that drives a suitable immune response is increasing. The oxidized carbon nanosphere (OCN), a negatively charged carbon nanoparticle, has the potential to fulfill this requirement because it can efficiently deliver macromolecules into cells and allows endosomal leakage. However, fundamental insights into how OCNs are taken up by antigen-presenting cells, and the intracellular behavior of delivered molecules is lacking. Furthermore, how immune responses are stimulated by OCN-mediated delivery has not been investigated. Purpose: In this study, the model protein antigen ovalbumin (OVA) was used to investigate the uptake mechanism and intracellular fate of OCN-mediated delivery of protein in macrophages. Moreover, the immune response triggered by OVA delivered by OCNs was characterized. Methods: Bone-marrow-derived macrophages (BMDMs) from mice were used to study antigen uptake and intracellular trafficking. Mice were immunized using OCN-OVA combined with known adjuvants, and the specific immune response was measured. Results: OCNs showed no cytotoxicity against BMDMs. OCN-mediated delivery of OVA into BMDMs was partially temperature independent process. Using specific inhibitors, it was revealed that intracellular delivery of OCN-OVA does not rely on phagocytosis or the clathrin- and lipid raft/caveolae-mediated pathways. Delivered OVA was found to colocalize with compartments containing MHC class I, but not with early endosomes, lysosomes, and autophagosomes. Immunization of OVA using OCNs in combination with the known adjuvant monophosphoryl lipid A specifically enhanced interferon gamma (IFNγ)- and granzyme B-producing cytotoxic T cells (CTLs). Conclusion: OCNs effectively delivered protein antigens into macrophages that localized with compartments containing MHC class I partially by the temperature independent, but not clathrin- and lipid raft/caveolae-mediated pathways. Increased CD8+ T-cell activity was induced by OCN-delivered antigens, suggesting antigen processing toward antigen presentation for CTLs. Taken together, OCNs are a potential protein antigen delivery system that stimulates the cell-mediated immune response.

Hydrophilic and Cell-Penetrable Pyrrolidinyl Peptide Nucleic Acid via Post-synthetic Modification with Hydrophilic Side Chains.

Peptide nucleic acid (PNA) is a nucleic acid mimic in which the deoxyribose-phosphate was replaced by a peptide-like backbone. The absence of negative charge in the PNA backbone leads to several unique behaviors including a stronger binding and salt independency of the PNA-DNA duplex stability. However, PNA possesses poor aqueous solubility and cannot directly penetrate cell membranes. These are major obstacles that limit in vivo applications of PNA. In previous strategies, the PNA can be conjugated to macromolecular carriers or modified with positively charged side chains such as guanidinium groups to improve the aqueous solubility and cell permeability. In general, a preformed modified PNA monomer was required. In this study, a new approach for post-synthetic modification of PNA backbone with one or more hydrophilic groups was proposed. The PNA used in this study was the conformationally constrained pyrrolidinyl PNA with prolyl-2-aminocyclopentanecarboxylic acid dipeptide backbone (acpcPNA) that shows several advantages over the conventional PNA. The aldehyde modifiers carrying different linkers (alkylene and oligo(ethylene glycol)) and end groups (-OH, -NH2, and guanidinium) were synthesized and attached to the backbone of modified acpcPNA by reductive alkylation. The hybrids between the modified acpcPNAs and DNA exhibited comparable or superior thermal stability with base-pairing specificity similar to those of unmodified acpcPNA. Moreover, the modified apcPNAs also showed the improvement of aqueous solubility (10-20 folds compared to unmodified PNA) and readily penetrate cell membranes without requiring any special delivery agents. This study not only demonstrates the practicality of the proposed post-synthetic modification approach for PNA modification, which could be readily applied to other systems, but also opens up opportunities for using pyrrolidinyl PNA in various applications such as intracellular RNA sensing, specific gene detection, and antisense and antigene therapy.

Penetration of Oxidized Carbon Nanospheres through Lipid Bilayer Membrane: Comparison to Graphene Oxide and Oxidized Carbon Nanotubes, and Effects of pH and Membrane Composition.

Here we show that the ability of oxidized carbon particles to penetrate phospholipid bilayer membrane varies with the particle shapes, chemical functionalities on the particle surface, lipid compositions of the membrane and pH conditions. Among the similar surface charged oxidized carbon particles of spherical (oxidized carbon nanosphere, OCS), tubular (oxidized carbon nanotube, OCT), and sheet (oxidized graphene sheet, OGSh) morphologies, OCS possesses the highest levels of adhesion to lipid bilayer membrane and penetration into the cell-sized liposome. OCS preferably binds better to the disordered lipid bilayer membrane (consisting of 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine) as compared to the ordered membrane (consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine and cholesterol). The process of OCS-induced leak on the membrane is pH responsive and most pronounced under an acidic condition. Covalently decorating the OCS's surface with poly(ethylene oxide) or (2-aminoethyl)trimethylammonium moieties decreases its ability to interact with the membrane. When used as carriers, OCSs can deliver curcumin into nucleus of A549 human lung cancer and human embryonic kidney cells, in contrast, curcumin molecules delivered by OCTs remain in the cytoplasm. OGShs cannot significantly enter cells and cannot induce noticeable cellular uptake of curcumin.