Proposed molecular model for electrostatic interactions between insulin and chitosan. Nano-complexation and activity in cultured cells

Abstract

The objective of this contribution was to propose a model that would explain the nanocomplexes formation between Human Recombinant Insulin (I) and a polydisperse Chitosan (CS). Such an objective implied exploring I and CS concentration conditions that allowed the formation of complexes with defined and reproducible submicronic dimensions. I-CS complexes were obtained by mixing I and CS solutions at pH 2 and then increasing the pH up to 6 promoting electrostatic interactions between them. Colloidal stages of I and I-CS nano-complexes formation were characterized by dynamic light scattering (DLS), ζ-potential, solutions flow behavior and absorbance measurements. 1•10-2 %, w/w, of CS allowed covering completely the surface protein aggregates constituting core-shell nano-structures of 200 nm, with a ζ-potential of 17,5 mV. Solution dynamic viscosity results kept relation with different stages of nanocomplexation process. Biological activity of I-CS complexes was studied in 3T3-L1 cultured fibroblast showing a delayed and sustained activity as compared to free insulin. I-CS nano-complexes could be an alternative for developing a new generation of drugs allowing I protection from the hostile conditions of the body and increasing its absorption. These findings have basic and practical impacts as they could be exploited to exert the controlled release of I in therapeutic formulations by using the I-CS nanocomplexes.

High molecular weight chitosan based particles for insulin encapsulation obtained via nanospray technology

“…Such differences are shown in Figure 1a and b. [25] As far as we know, there are no previous works concerning nanospray dried NP formed with high molecular weight CS, like the one used here for INS entrapping. In fact, literature is abundant regarding other small molecules employed in nanospray drying particle generation.…”

“…Western blots were performed according to standard procedures. [25] Briefly, After that, the membranes were incubated overnight at 4 C with a rabbit primary antibody anti-P-Akt S473 1/ 1000 in 5% BSA in TBS (Cell Signaling, USA). As secondary antibody a Rabbit IgG-HRP 1/5000 in 5% BSA in TBS (Santa Cruz, CA, USA) was added to the membrane and incubated during 1 h at room temperature.…”

“…Just at pH 6 the electrostatic interactions between INS and CS reach their maximum. [25] Figure 6 shows the configuration proposed for this INS-CS NP structure acquired after NP powder re-hydration at different pH solution. NP would constitute matrices or networks were the INS would be trapped.…”

“…This work is part of a larger project that deals with the encapsulation of INS into CS-based nanostructures. Previously, core-shell NP were designed using the electrostatic complexation principle and their characterization was described in Prudkin-Silva et al [25] The objective of the present work was to obtain and characterize CS based nanoparticles for INS encapsulation via nano spray drying technology from mixed solutions at pH 3. Under these conditions, the INS entrapment could be promoted into a CS matrix.…”

Proteins as Nano-Carriers for Bioactive Compounds. The Case of 7S and 11S Soy Globulins and Folic Acid Complexation

“…In this type of analysis, the presence of more than one family of particle sizes is taken into consideration. These multi-exponential algorithms, for which there is no standard and which generally vary from manufacturer to manufacturer, will generate an intensity distribution and, using the Mie theory, a distribution normalized for the volume of the scattering particles [ 28 ]. Volume size distribution was considered to determine the relative importance of each peak.…”