372016 Thermodynamics of Assembled Folate Nanoparticles

Sunday, November 16, 2014: 5:10 PM
211 (Hilton Atlanta)
Sanat Mohanty, Chemical Engineering, Indian Institute of Technology, Delhi, New Delhi, India and Rahul Mishra, Chemical Engg, IIT Delhi, New Delhi, India

Thermodynamics of folate based Nano Particles

Development of nano-particles from small molecules is of interest for a variety of applications, including drug delivery, diagnostics, in food industry as flavor carriers, in organic reactions, among others. Different kinds of molecules (including polymers, surfactants, liquid crystalline materials) have been used to make such nanoparticles. The structure of nanoparticles formed affects the properties of these particles, processing ease of these particles and subsequently, their use in these applications. In this presentation, we share the method of development of folate based nanoparticles. The choice of folates as a nanocarrier is significant because folic acid is part of our natural diet, is often used as supplements, and is non-toxic at concentrations of use. In addition, especially for chemotherapeutic applications, where folates are used as a supplement in therapy, use of folate as a carrier releasing in the same environment where the drug is targeted becomes doubly important. This study is significant in that it describes the self-assembly of a class of materials that is driven largely by enthalpic considerations - we show that this affects its assembled structure as well as it effectiveness as a drug delivery carrier.

Experimental characterization, molecular simulation and theory are used to understand the assembly of these molecules in solutions at multiple scales. Folates self assembly in a narrow range of pH – in this range they are liquid crystalline solutions [1]. While past studies have shown assembly at concentrations of >25% by weight, we show that folate molecules assemble at very low concentrations – down to 0.1% by weight [2-4]. X-Ray Diffraction studies are used to identify the ordered structure of the assembly – they point to strong aromatic ring interactions between the folate ions and stacking of the folate ions. In addition, the XRD also help estimate the dimensions of the stacks formed. These methods are used to show that folates form multi-molecular stacks at extremely low concentrations; and that the fraction of molecules in the monomeric state is very small. The latter can also be proven using simple statistical thermodynamic approximation using the solvation energy of the folate ion and the folate-folate interaction in its stacking orientation.

Simulations were also carried out with folate ions in water with a GPU based Molecular Simulation system [5], using Dreiding force fields for folates and TIP3P for water molecules. The simulations were able to simulate the absence of self assembly of folic acid molecules while predicting the behavior of folates in their self-assembled structure. The radial distribution functions of the folate stacks in water, calculated from simulations are consistent with XRD peaks of these solutions. These radial distribution functions at different concentrations also track the small changes in structure with changing phases, as characterized using XRD. Further, the rdf of water molecules in the vicinity of the folate ions, compared with folic acid, also show that there is little change in the order of water around the folate / folic acid molecules. The stacking is primarily driven by enthalpic considerations.

Simulations were also used to identify the inter-molecular forces that drove assembly of the folates. Simulations of fictitious molecules made up of segments of the folate ions helped identify interactions that drove aromatic stacking, those that influenced the in-plane position and orientation of the folate ion within the stack and the role of ionized functional groups. These simulations also provide a broader understanding of self-assembly of other chromonics molecules.

Difference in the nature of interactions between two molecules can be used to design and control phases of two component systems. Such interactions have been used to design nanodomains, such as in acrylate-rubber systems or styrene-acrylate systems [6-9]. U.S. Patent no. 7,629,027 B2 [10] cites the use of Hydroxy Propyl Methyl Cellulose (HPMC) in engineering nanodomains of n-methyl imidazole (NMI) based chromonics assemblies. This work uses similar understanding of interactions between folate ions and HPMC to create phases with discontinuous nanodomains of folate assemblies dispersed in a continuous domain of HPMC.

Nanoparticles of these assembled structures were made by creating nano-phases of these liquid crystalline solutions in equilibrium with aqueous polymers (such as hydroxy propyl methyl cellulose (hpmc)). The phase diagram of this pseudo 2-component system is developed to help identify the range of compositions where folates exist in nano-domains. The phase behavior study helps us identify the range of folate:hpmc compositions where spherical nano-domains in the range of 100-500nm can be engineered.

At low concentrations of HPMC, and with higher folate ion concentration (lower than about 2g/ml HPMC at about 0.15g/ml folate), one sees a continuous phase of folates (in the nano-scale) characterized by SEM images showing micron sized flakes and dark regions (of HPMC domain). With increasing folate concentration, the size of the flakes grows.

On the other hand, at low concentrations of folate and higher HPMC concentrations (about 0.5g/l of HPMC and 0.25g/l of folates), SEM images present nanodomains of folate structures in a continuous phase of HPMC. The nanodomains of folate phase become even smaller at increasing concentration of the HPMC component (with nanodomains approaching 150nm at high concentrations of HPMC).

There is an intermediate region between the above described regions. In this region is a bi-continuous phase of folate and HPMC. The final structures are independent of the initial concentration of the folate or the HPMC concentration or the mixing process – they only depend on the final ratio of folate to HPMC. Thus, we argue that these nano-domains are equilibrium structures and are not dependent on the kinetics of the process.

The nano-domains of folate  formed in the continuous phase of HPMC can be exposed  to a solution of multivalent ions such as ZnCl2, CaCl2, or AlCl3. These studies posit that these multivalent cations exchange with the monovalent cation that was used to formulate the liquid crystalline solutions (NaOH in this case) to crosslink the folate ions within the nano-domain structures. Subsequently, the nano-domains form stable nano-particles that can be separated from the solution and used as needed.


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See more of this Session: Nanostructured Particles for Bio, Food and Pharma
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