Synthesis, Characterization and Biological Applications of Phyto-assisted Chitosan Nanocomposites

Abstract

There is an increased demand for eco-friendly bio-degradable food packaging material to control the worldwide environmental crises arising due to petroleum derived plastic packaging. This study was designed in a contribution to make active chitosan films reinforced with two different types of nano fillers (cellulose nanocrystals and nano keratin) to increase the mechanical and barrier properties of chitosan films. Three ethanolic plant extracts (Cinnamomum tamala, Amomum subulatum and Mentha piperita) and their major polyphenols (quercetin, rutin and vanillic acid) were incorporated as additives to augment the antioxidant and antimicrobial properties of prepared films. Moreover, combinations of two nano fillers (synergistic effect of hybrid system) with plant extracts were evaluated for their effect on film properties. Tensile testing, thermo-gravimetric analysis, differential scanning calorimetry, dynamic mechanical analysis, water vapor permeability and opacity were used to evaluate the mechanical and barrier properties. Biodegradability of films was compared both in soil and lysozyme. The in vitro hemo-compatibility of films was checked on red blood cells. Acute oral toxicity effects of prepared films were analyzed in vivo. Baby carrot packaging was done with synthesized films and the effect of packaging on storage quality was assessed. Screening of plants resulted in selection of C. tamala, A. subulatum and M. piperita for incorporation in chitosan films due to their highest total phenolic content, 70-80% free radical scavenging and moderate antimicrobial activities Among polyphenols, quercetin, rutin and vanillic acid were selected due to their highest amount in the above extracts. Docking analysis predicted that selected polyphenols were one of the major reasons for the antimicrobial effect of these extracts via inhibition of specific enzymes. Chitosan films were synthesized successfully via solvent casting and results showed that extract incorporated films reinforced with both nano fillers (CNC & NK) exhibited 59% increase in tensile strength of chitosan films. However, an increase of 63% was seen in tensile strength of CNC reinforced polyphenol composites. Extract incorporated films depicted excellent barrier properties (42% decrease in water vapour permeability), a two-fold increase in opacity, an enhanced thermal stability and glass transition temperature as compared to control chitosan films. Polyphenol composites displayed only 22% decrease in WVP and 38% increase in opacity. Extract incorporated Abstract x composites were found to be more thermally stable as compared to polyphenol composites. Analysis of both extract and polyphenol incorporated films showed higher polyphenol content as compared to control which indicates correct incorporation of extract/polyphenol in the synthesized films. The films having higher TPC were found to have higher antioxidant activity. Synthesized films showed 70% free radical scavenging however the antimicrobial activity was found to be moderate. Among all combinations of nanocomposites films, M. piperita and quercetin loaded nanocomposite films displayed best DPPH scavenging and provided least weight loss of baby carrots while determining the food quality. Biodegradability results showed that synthesized films (extract/polyphenol incorporated) displayed increased degradation in soil than lysozyme. CS-CNC-NK EX1 composites displayed 63% increased degradation than control chitosan films along with lower melting temperature and early degradation temperature in TGA and DSC analysis which suggest that chitosan film degradation highly depends on chemical modifications in their structure. The results of in vitro hemo-compatibility assay indicated the non-hemolytic to slightly hemolytic behavior of synthesized chitosan composites suggesting their non-toxicity. Moreover, acute oral toxicity analysis in rats also confirmed the non-toxic character of synthesized films as the oral ingestion of these films induced no significant change in body weight, hematological and biochemical parameters of blood as well as a normal functioning of liver and kidney was observed with no mortality. In conclusion, we propose that incorporation of C. tamala, A. subulatum andM. piperita extracts and their polyphenols specifically quercetin to chitosan films in combination of CNC and NK as a hybrid nano fillers system can result in remarkably enhanced mechanical, barrier and biological properties of chitosan films for efficient use in food packaging.