Besifloxacin Nanocrystal: Towards an Innovative Ophthalmic Preparation

Abstract

Bacterial conjunctivitis significantly impacts public health, including more than one-third of eye diseases reported worldwide. It is an infection caused by various aerobic and anaerobic bacteria and is highly contagious. Therefore, it has a high incidence of bacterial resistance to the antibiotics commonly used for treatment. Among the most recent antibiotics, besifloxacin is a fourth-generation fluoroquinolone antibiotic indicated exclusively for topical ophthalmic use. Due to its importance in treating bacterial conjunctivitis and its low solubility in water, limiting its efficacy, a nanotechnology-based drug delivery preparation was developed to overcome this hurdle. Besifloxacin nanocrystals were prepared by small-scale wet milling and response surface methodology, using Povacoat^® as a stabilizer. The particle's average hydrodynamic diameter (Z-ave) was approximately 550 nm (17 times smaller than raw material), with a polydispersity index (PdI) of less than 0.2. The saturation solubility increased about two times compared to the raw material, making it possible to increase the dissolution rate of this drug substance, potentially improving its bioavailability and safety. The optimized preparation was stable under an accelerated stability study (90 days). The Z-ave, PZ, PdI, and content did not alter significantly during this period. Furthermore, the 0.6% m/m besifloxacin nanocrystals at the maximum dose and the Povacoat^® stabilizer did not show toxicity in Galleria mellonella larvae. The innovative ophthalmic preparation minimum inhibitory concentration (MIC) was 0.0960 µg/mL and 1.60 µg/mL against Staphylococcus aureus and Pseudomonas aeruginosa, respectively, confirming in vitro efficacy. Therefore, besifloxacin nanocrystals revealed the potential for reduced dosing of the drug substance, with a minor occurrence of adverse effects and greater patient adherence to treatment.

Keywords: Povacoat®; besifloxacin; conjunctivitis; fluoroquinolones; nanocrystals.

Figure 1

Particle size distribution for the commercial product Besivance^® and besifloxacin hydrochloride. The peaks represent the average of three readings.

Figure 2

Aspect of the formulas containing the stabilizers Povacoat^® 1.0 wt% (F1), Kollicoat^® 0.3 wt% + Chitosan 0.3 wt% (F3), Poloxamer^® 407 0.5 wt% + Chitosan 0.5 wt% (F7), and Poloxamer^® 407 0.3 wt% + Chitosan 0.3 wt% (F8). Stability after 15 days (25 °C).

Figure 3

Contour graph related to the Z-ave evaluation of the besifloxacin nanocrystal, whose variables are besifloxacin concentration (wt%), Povacoat^® concentration (wt%), and velocity (rpm).

Figure 4

Particle size distribution of besifloxacin nanosuspension 0.6 wt% by laser diffraction (LD). Representative of three runs (n = 3).

Figure 5

Microphotography of raw material besifloxacin (a) and besifloxacin nanocrystals (b).

Figure 6

DSC curves of formulation components, mixture (same proportion as formulation), and besifloxacin nanocrystals.

Figure 7

X-ray diffraction of besifloxacin (raw material), nanocrystals, mixture (besifloxacin and Povacoat^®), and Povacoat^®. n = 3.

Figure 8

Saturation solubility of besifloxacin (raw material) and nanosuspension (besifloxacin 0.6 wt%). (a) Solubility after 24 h of incubation. (b) Solubility after 48 h of incubation. (c) pH of the samples at the end of the incubation. Theoretical concentration: 50 µg/mL. n = 3.

Figure 9

Determination of the average hydrodynamic diameter (Z-ave) in nm and polydispersion index (PdI), for the accelerated stability study of the nanosuspension of besifloxacin 0.6 wt%, in the time intervals: T0, 30, 60, and 90 days. Study condition: 40 °C ± 2 °C/75% RH ± 5% RH. n = 3 batch.

Figure 10

Determination of zeta potential (ZP) and pH for the accelerated stability study of the nanosuspension of besifloxacin 0.6 wt%, in the time intervals: T0, 30, 60, and 90 days. Study condition: 40 °C ± 2 °C/75% RH ± 5% RH. n = 3 batch.