Novel Thymoquinone Nanoparticles Using Poly(ester amide) Based on L-Arginine-Targeting Pulmonary Drug Delivery

Abstract

Thymoquinone (TQ), the main active constituent of Nigella sativa, has demonstrated broad-spectrum antimicrobial, antioxidant, and anti-inflammatory effects, which suggest its potential use in secondary infections caused by COVID-19. However, clinical deployment has been hindered due to its limited aqueous solubility and poor bioavailability. Therefore, a targeted delivery system to the lungs using nanotechnology is needed to overcome limitations encountered with TQ. In this project, a novel TQ-loaded poly(ester amide) based on L-arginine nanoparticles was prepared using the interfacial polycondensation method for a dry powder inhaler targeting delivery of TQ to the lungs. The nanoparticles were characterized by FTIR and NMR to confirm the structure. Transmission electron microscopy and Zetasizer results confirmed the particle diameter of 52 nm. The high-dose formulation showed the entrapment efficiency and loading capacity values of TQ to be 99.77% and 35.56%, respectively. An XRD study proved that TQ did not change its crystallinity, which was further confirmed by the DSC study. Optimized nanoparticles were evaluated for their in vitro aerodynamic performance, which demonstrated an effective delivery of 22.7-23.7% of the nominal dose into the lower parts of the lungs. The high drug-targeting potential and efficiency demonstrates the significant role of the TQ nanoparticles for potential application in COVID-19 and other respiratory conditions.

Keywords: COVID-19; L-arginine; nanoparticles; poly(ester amide); respiratory drug delivery; thymoquinone.

Figure 1

(a) Synthesis of di-p-toluenesulfonic acid salt of O,O’-bis-(L-arginine)-diethylene glycol monomer. (b) Synthesis of poly(ester amide)s by interfacial polycondensation of di-p-toluenesulfonic acid salt of O,O’-bis-(L-arginine)- diethylene glycol monomer.

Figure 2

Schematic representation of interfacial polycondensation process and the formed TQ-loaded poly(ester amide) based on L-arginine, where part of the TQ is dispersed within the polymer and the other part is attracted to the surface of the polymer.

Figure 3

FTIR spectra: (a) TQ; (b) di-p-toluenesulfonic acid salt of O,O’-bis-(L-arginine)-diethylene glycol monomer; (c) poly (ester amide) based on L-arginine; (d) TQ poly(ester amide) formula.

Figure 4

PCA of FTIR spectra (1800–700 cm⁻¹) of TQ, the monomer, the polymer, and TQ-loaded polymer (formula): (a) score plot of PC1 (86.2%) vs. PC2 (7.5%) and (b) loading profiles of PC2 and PC3 within the allocated range (the dotted line is the loading profile of PC3, while the solid black line represents the loading profile of PC2).

Figure 5

(a) ¹H NMR of di-p-toluenesulfonic acid salt of O,O’-bis-(L-arginine)-diethylene glycol monomer (experiments were recorded at 500 MHz). (b) ¹H NMR of poly(ester amide) based on L-arginine. (c) ¹³C NMR of di-p-toluenesulfonic acid salt of O,O’-bis-(L-arginine)-diethylene glycol monomer. (d) ¹³C NMR poly (ester amide) based on L-arginine. Experiments were recorded at 125 MHz.

Figure 6

Differential scanning calorimetry (DSC) thermograms: (a) TQ powder; (b) the polymer; (c) TQ–polymer nanoparticles highlighting the enthalpy and melting points.

Figure 7

TEM images of (a,b) the poly (ester amide) based on L-arginine and (c,d) TQ-loaded nanoparticles. Scales: (a) 100 nm, (b) 200 nm, (c) 100 nm, and (d) 100nm.

Figure 8

XRD patterns for panel [I]: (a) TQ powder; (b) polymer; (c) TQ–polymer nanoparticles. Panel [II] focus section from 10 to 50°: (d) TQ powder; (e) polymer; (f) TQ–polymer nanoparticles. Note in (II-d), the rhombus shapes over the peaks are related to TQ, while the circles are for the TQ-loaded polymer.

Figure 9

Calibration curve of TQ over a concentration range of 3.9 to 62.5 µg/mL.

Figure 10

Aerodynamic performance of TQ-containing nanoparticles, highlighting the main parameters for formulations F2 and F3. ED: emitted dose; FPF-ED: fine particle fraction of emitted dose; FPF-ND: fine particle fraction of nominated dose; RD: respirable dose. Results are presented as means ± SD, n = 3.

Figure 11

Aerodynamic particle size distribution of TQ-containing nanoparticles, highlighting the percentage of particles deposited the pulmonary section, which is the lower part of the respiratory system. Results are presented as means ± SD, n = 3.

Figure 12

Release profile of TQ from TQ-loaded poly(ester amide) based on L-arginine nanoparticles (F3) for 24 h (means ± SD, n = 3).