Nano-in-micro drug delivery system for bacterial pigment violacein

CARNAGHI, LUCIA; RIVERO BERTI, IGNACIO; CASTRO, GUILLERMO RAUL; ISLAN, GERMAN A

Buenos Aires

Workshop; Fronteras en Nanobiotecnología III; 2022

Violacein (VIOL) is a bacterial pigment produced, among several other strains, by Chromobacterium violaceum. It has been shown that VIOL have many potential pharmacological applications, as trypanocidal, immunomodulator, antibacterial and antitumoral drug 1. Since VIOL posses a poor solubility in water, a drug delivery system is required. VIOL was encapsulated into Nanostructured Lipid Carrier (NLC), following our previous work2. Briefly, VIOL and a melted lipid phase consisting in cetyl-palmitate and capric-caprylic triglycerides are mixed with an aqueous phase consisting of 3,0% Poloxamer P188 solution, and then sonicated in a probe ultrasonic processor for 10min. VIOL in NLC encapsulation efficiency (EE, %) was higher than 99% with a drug/lipid ratio of 0.5 μmol/mg lipid and nanoparticles with a mean diameter around 200 nm (determined by DLS).To create a nano-in-micro drug delivery system (NiM), NLC-VIOL were incorporated by precipitation in calcium carbonate microparticles according to a previous paper3 with a few modifications. This could give the system a pH dependent behavior. Briefly 9.0ml of Na2CO3 (3.2%, w/v) were mixed with a solution containing a biopolymer, alginate (Alg), or high methoxyl pectin (HMP) from two different sources, critic pectin DM:65% HMP1 (GELFIX®), and Apple pectin DM:75% HMP2 (Sigma), and the NLC-VIOL. Then 9,0ml of glycine buffer (pH=10.0) and 3.2% (w/v) CaCl2 were added, and the mixture stirred 5min in an ice bath, followed by 10min at 1000rpm. Additionally, a preparation without biopolymer (NiM-SB) was performed. The precipitated product was collected by centrifugation, washed with ultrapure water, and freeze-dried for storage. The EE(%) was measured by a direct method, extracting all VIOL in the particles with isopropanol. The EE was 17.0% for NiM-SB, 14.7% for NiM-Alg, but 49.4% or 45.4% for NiM-HMP1 and HMP2 respectively. These results indicate a relationship between biopolymer hydrophobicity and NLC-VIOL retention within the matrix. Optical microscopy images of the NiM particles showed an average size of 2.0±0.2μm and 2.4±0.4μm for both HMP formulations, 1.2±0.5μm for NiM-SB and 2.3±0.3μm for NiM-Alg. All the formulations showed a NLC-Viol controlled release profiles in a pH dependent behavior. NLC was effectively encapsulated in a microparticulated system, forming a nano-in-micro drug delivery platform.