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In vitro evaluation of physicochemical properties of soft lining resins after incorporation of chlorhexidine

In vitro evaluation of physicochemical properties of soft lining resins after incorporation of chlorhexidine

In vitro evaluation of physicochemical properties of soft lining resins after incorporation of chlorhexidine

Journal of Prosthetic Dentistry, 2020-11-01, Volume 124, Issue 5, Pages 615.e1-615.e7, Copyright © 2020 Editorial Council for the Journal of Prosthetic Dentistry


Statement of problem

Incorporating chlorhexidine into soft lining materials has been suggested to reduce biofilm development on the material surface and treat denture stomatitis. However, evaluation of the physicochemical properties of this material is necessary.


The purpose of this in vitro study was to evaluate the physicochemical properties of resin-based denture soft lining materials modified with chlorhexidine diacetate (CDA).

Material and methods

Two soft lining resins were tested, one based on polymethyl methacrylate (PMMA) and the other on polyethyl methacrylate (PEMA), into which 0.5%, 1.0%, or 2.0% of CDA was incorporated; the control group had no CDA. The specimens were stored for 2 hours, 48 hours, 7, 14, 21, and 28 days and then analyzed for polymer crystallinity, Shore A hardness, degree of monomer conversion, residual monomer leaching, and CDA release. Data were analyzed by using a 3-way ANOVA and the Tukey HSD test (α=.05).


The polymer crystallinity of PEMA and PMMA did not change after CDA incorporation. Shore A hardness increased over time, but not for any CDA concentrations tested after 28 days ( P >.05). Considering the degree of conversion, PMMA-based resin showed no statistically significant difference ( P >.05). However, PEMA-based resin showed a significant decrease ( P <.05), which was reflected in a significant increase in residual monomer leaching from PEMA-based resin with the incorporation of 0.5% and 1.0% CDA ( P <.05), mainly in the first 48 hours. PMMA-based resin showed no change in monomer leaching ( P >.05). For both resins, the CDA release kinetics were related to monomer leaching; for PEMA-based resin, the values were significantly higher in the first 48 hours ( P <.05), and for PMMA-based resin, the values were more sustained up to the last day of analysis.


The incorporation of CDA did not affect the physicochemical properties of soft resins. The properties of PMMA were better than those of PEMA.

Clinical Implications

Incorporating chlorhexidine into soft lining materials may help treat denture stomatitis without depending on patient compliance, as the physicochemical properties of the soft resins were not affected.

The use of soft resins for relining dental prostheses is well accepted with well-documented advantages, especially for tissue conditioning and after surgical procedures. However, these materials undergo time-dependent changes in the oral environment, such as loss of soluble components, water sorption, reduction of elasticity, increase in surface hardness, increase in surface roughness, and debonding from the denture base. These factors favor the accumulation of microorganisms, with Candida albicans being the most commonly found one.

The control of opportunist infections caused by C. albicans is important, but treatment depends on the patient’s cooperation with cleaning and the use of topical antimicrobial and antifungal agents. , The difficulty patients have adapting to the correct use of topical medication has been identified as one reason why candidiasis is highly prevalent in older and systemically compromised patients.

To reduce the need for patient cooperation, Garner et al described chlorhexidine coatings of the surface of silicone soft liners. Although promising, its activity on Candida inhibition was tested for only 6 hours, and because the chlorhexidine was only on the surface, hyphae penetration into the material once the surface was colonized was not inhibited. Additionally, the authors recommended reapplication, either as part of a daily routine or as a periodic treatment.

Antimicrobial agents have been incorporated into soft resins, other polymeric materials, and silicone-based denture liners to provide slow and prolonged drug release into the mouth. This approach may be useful against C. albicans , as patient cooperation is not required. Moreover, the medication is constantly in contact with the oral mucosa, unlike topically applied medication, which is removed from the oral cavity during the first 3 hours. ,

The antifungal agent zinc undecylenate has been incorporated into a commercially available denture liner, with the goal of reducing fungal colonization. However, this agent in the concentration provided by the manufacturer has been reported to be insufficient to prevent C. albicans colonization on a resin surface. , ,

The present in vitro study was an extension of a study that used chlorhexidine diacetate in low concentrations (0.5%, 1.0%, and 2.0%) to inhibit a C. albicans biofilm when incorporated into soft resins. Chlorhexidine has excellent antifungal activity against a broad range of Candida spp . and is better than antifungal agents. , The use of low percentages of this medication may reduce deleterious effects to the soft resin. This has encouraged evaluation of the physicochemical properties of lower concentrations.

The present study investigated whether the incorporation of chlorhexidine would change the physicochemical properties of the soft resins used. The null hypotheses were that the incorporation of chlorhexidine would not affect the crystallinity of the matrix of the resins, leaving their hardness unchanged, and that the incorporation of chlorhexidine would not affect the degree of conversion of the resins, leaving the pattern of resin monomer leaching unchanged and allowing the release of chlorhexidine into the medium.

Material and methods

Two different soft resins were tested: a polymethyl methacrylate (PMMA) based and a polyethyl methacrylate (PEMA) based ( Table 1 ). Both of them received different concentrations of chlorhexidine diacetate (CDA).

Table 1
Composition, manufacturer, and batch number of materials (resin-based denture soft lining materials and chlorhexidine) used
Material Composition Manufacturer Batch Number
Powder: Polymethyl methacrylate, zinc undecylenate, and pigments.
Liquid: Benzyl salicylate, ethyl alcohol, methylsalicylate, peppermint oil.
Coe-Soft, G Corp. 1109061
Powder: Polyethyl methacrylate and pigments.
Liquid: Alkyl phthalate (plasticizer) and ethyl alcohol.
Trusoft, The Bosworth Co 1107-333
Powder: Chlorhexidine diacetate salt hydrate. Sigma Aldrich 083k0014

Shore A hardness was evaluated by using a digital durometer Shore A (DP-100; Instrutherm) and related to the acrylic resin crystallinity by the X-ray diffraction technique (Miniflex; Rigaku). The degree of polymer conversion was evaluated by using Fourier-transform infrared spectroscopy (FTIR) (Spectrum 100 Optica; PerkinElmer), and this was correlated to residual monomer leaching and the release of incorporated chlorhexidine, both quantified by using high-performance liquid chromatography (HPLC) associated with ultraviolet (UV) spectrometry. All the evaluations were carried out after 6 intervals: baseline (2 hours), 2, 7, 14, 21, and 28 days, with the exception of the crystallinity and degree of conversion evaluations, which were performed just before and after polymerization.

The PMMA- and PEMA-based soft resins were weighed and prepared in accordance with the manufacturer’s instructions. The CDA salt was weighed separately on a precision scale (BG200; Gehaka) to obtain 0.5%, 1.0%, and 2.0% by weight for each portion of the polymer material. CDA was dissolved in the monomer until a homogeneous mixture was obtained, and only then was the polymer added and mixed for 30 seconds.

For the hardness test, rectangular test specimens (n=9) measuring 7.0×5.0 cm and 6 mm in thickness were used. For the polymer crystallinity test, rectangular test specimens (n=9) measuring 2.0×3.0 cm and 1 cm in thickness were used. , A silicone matrix was used to standardize the shape and size of specimens by using a polyoxymethylene flask submitted to hydraulic 0.1-MPa pressure at room temperature for 30 minutes. Specimen thickness was checked by using digital calipers, and specimens were stored in sterile distilled water at 37 °C until the tests were performed. For conversion degree (n=10) (FTIR), residual monomer leaching (n=9) (HPLC), and chlorhexidine release (n=9) (HPLC), disk-shaped test specimens measuring 10.0 mm in diameter and 3.0 mm in thickness were made in a silicone matrix, with a glass plate under a 2.9-N load to avoid bubble formation and to standardize the test specimens. The polymerization time was 5 minutes at room temperature, after which specimens were individually stored at 37 °C in 24-well cell culture plates (TPP Techno Plastic Products AG) containing 1 mL of sterile distilled water in each well until testing.

Shore A hardness was determined in accordance with ASTM D2240 with a digital durometer Shore A (Model DD2-A; Kobunshi Keiki), and values were recorded 5 seconds after specimen loading. Three measurements per specimen were obtained on each side at a minimum of 12 mm from any edge.

Crystallinity was evaluated by X-ray diffraction (MiniFlex; Rigaku) operated at 40 kV and 30 mA, 0.05 degrees/s at ambient temperature. CuKα radiation was used as the X-ray source, which has a wavelength of 0.15418 nm. The interval of analysis was from 2θ=2 degrees to 2θ=80 degrees.

The degree of resin conversion was measured by FTIR (Spectrum 100 Optica; PerkinElmer) with an attenuated total reflectance element (ATR) coupled to a horizontal zinc selenide crystal (Pike Technologies) with a standard compression force of 100 N for 32 absorbance scans in each specimen. Initially, an unpolymerized resin specimen from each group was assessed by positioning it on the crystal and then performing 4 absorbance scans. The infrared spectra obtained were evaluated by using the peak intensity of 1.637 cm -1 , which is equivalent to the double carbon bonds between the methacrylate; moreover, peak intensity 2.952 cm -1 corresponds to the single connections made through polymerization. The ratio of the difference between the heights of the peaks of the polymerized and unpolymerized specimens was used according to the following expression:

DC (%)=([1.637 cm -1 /2.952 cm -1 polymerized]/[1.637 cm -1 /2.952 cm -1 unpolymerized])×100.

High-performance liquid chromatography (Alliance HPLC system; Waters Corp) was used to determine residual monomer leaching and to obtain the drug-release curve. The system used had a quaternary pump, degasser, automatic injector, column oven, and UV-DAD detector (Agilent Technologies). Analysis of the monomers released was performed by using an RP-18 column with a length of 250 mm by 2.5 mm in diameter, and the mobile phase used was a 66% MeOH solution to 34% H 2 O, with a flow of 2.0 mL/min. To obtain the chlorhexidine release curve, an SB C8 column (250×4.6 mm; 5 μm) was used. The mobile phase used was 0.03 M of monobasic sodium phosphate buffer, pH 2.0 (80:20, %v/v), at a flow of 2.0 mL/min.

The monomers and CDA released were detected at their maximum absorption values. The concentration of each compound was determined after constructing the calibration curve by using different concentrations of CDA. The storage water was changed at each time interval, which was based on other chlorhexidine-release systems in their most diverse presentations in acrylic polymer bases chemically similar to those used in this study. Mean and standard deviation were obtained from triplicate measurements.

The data were statistically analyzed by using a statistical software program (SigmaStat 3.1; Systat Software) (α=.05). All the data passed the normality test. Shore A hardness values and degree of conversion were analyzed by 2-way ANOVA with a subsequent Tukey HSD test. Values found for the different types of resin were analyzed separately for Shore A hardness as they were not to be compared. Residual monomer leaching and chlorhexidine release were analyzed by 3-way ANOVA, which included the type of resin, concentration of CDA, and storage time as independent factors. Data were submitted to multiple pairwise comparisons by using the Holm-Sidak method.

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