Development and Characterization of Sodium Bicarbonate-Based Gel for Cytolytic Vaginosis

Abstract

Background/Objectives: Cytolytic vaginosis or, classically, Doderlein's cytolysis is characterized by significant growth of species of the Lactobacillus genus, which leads to high amounts of lactic acid in the vaginal environment. Lactobacillus crispatus has been proposed as a key pathogen in this clinical condition. The symptomatology of cytolytic vaginosis is commonly confused with that of vulvovaginal candidosis, leading to inadequate and ineffective azole therapies. Nevertheless, historically, the use of sodium bicarbonate intimate baths was an effective way to reduce the symptoms of cytolytic vaginosis. Methods: In this study, four HPMC gel prototypes were developed, containing sodium bicarbonate concentrations ranging from 4% to 7% (w/w). These gels were evaluated for their physicochemical properties, antimicrobial activity, interference with lactobacilli adhering to cells, and cellular and tissue biocompatibility. Results: The gels presented pH values of around 9.0, and osmolality between 706 mOsm/kg (F4) and 1065 mOsm/kg (F7). The viscosity upon heating to physiologic temperature and dilution with simulated vaginal fluid was highly affected by the concentration of sodium bicarbonate. Gels with higher sodium bicarbonate concentrations (F6 and F7) were not shown to be stable in these conditions. All formulations exhibited effective antimicrobial activity against seven L. crispatus strains, with MIC values ranging from 6.25% to 25% (v/v) in terms of dilution. Additionally, the 4% (w/w) gel significantly interfered with the adhesion of L. crispatus to epithelial cells in competition and exclusion assays, reducing adhesion by more than 90% in relation to the control. Cytotoxicity tests on the Hec-1A, HeLa, and VK2/E6E7 cell lines indicated that the F4 and F5 gels demonstrated lower cytotoxicity levels compared to those with higher concentrations. Furthermore, ex vivo assays using porcine vaginal tissue confirmed that the 4% gel was non-toxic at a 25% (v/v) dilution. Conclusions: Based on these results, the 4% (w/w) sodium bicarbonate gel (F4) emerges as a promising therapeutic option for cytolytic vaginosis, offering effective bacterial interference, favourable physicochemical properties, and biocompatibility suitable for vaginal application.

Keywords: Lactobacillus; cytolytic vaginosis; gels; sodium bicarbonate; vagina.

Figure 1

Determination of viscosity directly at room temperature (25 °C) and at physiologic temperature 37 °C. Gels F4 and F5 were determined using the shear rate 20 1/s. Results are presented as the mean ± SD, and n = 3. **** p < 0.0001.

Figure 2

Macroscopic appearance of the 4% (w/w) sodium bicarbonate gel.

Figure 3

Lactobacillus crispatus growth measured using optical absorbance after treatment with sodium bicarbonate solution, gel base, and gels with sodium bicarbonate. (a) Sodium bicarbonate solution; (b) gel base; (c) gel with 4% (w/w) sodium bicarbonate; (d) gel with 5% (w/w) sodium bicarbonate; (e) gel with 6% (w/w) sodium bicarbonate; (f) gel with 7% (w/w) sodium bicarbonate. NC—negative control. Results are presented as the mean ± SD, and n = 3. Grey bars—Lactobacillus crispatus isolated from healthy women; Blue bars—Lactobacillus crispatus isolated from women diagnosed with cytolytic vaginosis.

Figure 4

Bacterial growth of Lactobacillus crispatus strains after contact with the pH-adjusted (alkalinized) culture medium. Results are presented as the mean ± SD, and n = 3.

Figure 5

Cytotoxic profiles of sodium bicarbonate solution, gel base, and gel with sodium bicarbonate in Hec-1A, HeLa, and VK2 E6/E7 cell lines. (a) Sodium bicarbonate solution; (b) gel base; (c) gel with 4% (w/w) sodium bicarbonate; (d) gel with 5% (w/w) sodium bicarbonate; (e) gel with 6% (w/w) sodium bicarbonate; (f) gel with 7% (w/w) sodium bicarbonate. Results are presented as the mean ± SD, and n = 6. Grey bars—experimental controls: NC—negative control; PC—positive control; SC—solvent control; Blue bars—dilutions of test samples.

Figure 6

Cellular viability of HeLa, Hec-1A, and VK2 E6/E7 after incubation with alkalinized culture medium (pH 8.3). Results are presented as the mean ± SD, and n = 3.

Figure 7

Adhesion profiles of the Lactobacillus crispatus PB8 strain to Hec-1A, HeLa, and VK2 E6/E7 cell lines in the presence of the gel with sodium bicarbonate 4% (m/v). Results are presented as the mean ± SD, and n = 3. ** p < 0.01, *** p < 0.001, and **** p < 0.0001 represent statistical significance in comparison with the control (100% bacterial adhesion).

Figure 8

Representative microphotographs (630×) of the different approaches used to study the interference of the gel containing 4% (w/w) sodium bicarbonate with Lactobacilli crispatus PB8 adhesion to epithelial cells. Each row of the panel corresponds to a cell line (Hec-1A, HeLa, and VK2 E6/E7), and in each column, the different approaches (competition, exclusion, and displacement) that were performed for the control and the gel with 4% (w/w) sodium bicarbonate are shown.

Figure 9

Tissue viability profile for gel with 4% (w/w) sodium bicarbonate at dilutions ranging from 0.78 to 50% (v/v). NC—negative control; PC—positive control; SC—solvent control. Results are presented as the mean ± SD, and n = 8. * p < 0.05; **** p < 0.0001 represents statistical significance in comparison with negative control.