Evaluation of the effect of coating Vertex denture lining ma terial with plant oils on microbial growth and hardness.

: Aims: To evaluate the effect of surface coating with natural plant oils ( Salvia officinalis , ginger and eucalyptus) on Candida growth and the hardness of Vertex denture lining material. Materials and method: Forty five specimens were prepared from soft acrylic lining material, twenty five of which were 10x10x2mm in size for testing antifungal activity, and twenty samples were 20mm in diameter and 12mm in thickness, for testing shore A hardness after coating samples with three types of natural oils ( Salvia officinalis , ginger and eucalyptus oils). Significant differences among the groups at ( p ≤0.05) level of significance were determined statistically with one way analysis of variance and Duncan's multiple range test Result: Antifungal assay showed a significant difference between five groups regarding Candida albicans growth ( p ≤ 0.05). For the hardness test, comparing different times of storage in water (1, 7, 14, 30 days) revealed a significant difference within all groups ( p ≤0. 05). While comparing the groups coated with natural oils with the control group, significant differences were found between different times of storage in water (1, 7, 30 day) ( p ≤0.05), except at 14 days of water storage there was no significant difference between groups ( p >0.05). Conclusion: All tested natural oils were effective as fungicidal agents and increased the softness and duration of soft acrylic lining material.


INTRODUCTION.
Soft denture lining materials are used in complete and partial removable dentures to distribute masticatory loads homogeneously on denturebearing tissues. These materials are suggested in cases of thin atrophic mucosa, irregular bone resorption, immediate dentures, for patients with xerostomia and bruxism, and during healing after implant placement. 1 Surface roughness and hardness changes are predisposing factors for microbial accumulation that compromise the material's durability. 2 The most widespread and important opportunistic fungal infections in the oral cavity is candidiasis, caused by yeasts belonging to the Candida genus, such as Candida albicans. The best way to prevent oral infections is microbial biofilm inhibition. Herbal extracts use denotes a new era for antimicrobial treatment after development of microbial resistance to antibiotics. 3 Ginger, eucalyptus and Salvia officinalis oils are examples of plants that exhibit great antifungal effects. [3][4][5] The aim of this research is to evaluate the effect of surface coating of the natural oils from Salvia officinalis ginger and eucalyptus on Candida albicans growth and hardness of Vertex denture lining material.

Antifungal Study
Preparation of Specimens: Twenty-five specimens were prepared from soft acrylic lining material (Vertex, Holland). The wax pattern (10x10x2mm) was made to fabricate stone mold in a metal flask. 6 The material was mixed according to the manufacturer's instructions and packed in the mold, the flask was then re-closed and pressed using a hydraulic press. Specimens were cured at a water bath (90 min at 73ºC and 30 min at 100°C) according to the manufacturer's instructions. Then the flash of access materials was trimmed with a scalpel.
Sterilization and Contamination of Specimens: All specimens were sterilized at an autoclave at 121°C for 15 minutes; the Candida albicans strain used in this study was isolated in a routine smear from patients with denture stomatitis then sub-cultured and the suitable identification biochemical tests were perfomed (germ tube test and sugar fermentation test).
On the first day, Candida albicans was inoculated to a turbidity of 0.5 McFarland which correspond to 10 8 organism/ml in 10ml of Brain Heart Infusion Broth (BHIB) and incubated at 37°C for 24 hrs. On the following day, 50ml of inoculated BHI broth were added to a test tube containing 10ml of sterile BHI broth, each sterile specimen tested was aseptically placed into a test tube, then it was sealed with foil and incubated for 24 hrs at 37°C. 7 Antifungal Study The method used for antifungal study was described by Al-Irhayim et al., 6 a total of 25 specimens were divided after contamination into five groups: Group A (Negative control): Five specimens individually placed in sterile BHI broth.
Group C: Five specimens were coated in ginger oil and left for eight hours. Group D: Five specimens were coated in Salvia officinalis oil and left for eight hours.
Group E: Five specimens were coated in eucalyptus oil and left for eight hours.
All tubes containing negative control specimens (group A) were vortexed vigorously (Tucker instruments LTD/ England) for one minute and rested for nine minutes followed by a short vortex to re-suspend any present organisms. To determine the number of microorganism in 10 -5 and 10 -6 dilution replicate specimens, 100µL of the resulting suspension were transferred onto a plate of Sabouraud agar containing 5 mg/ml gentamicin. The plates were incubated for 48 hrs at 37°C.
Yeast colonies in each plate specimen were counted. The colony forming units per milliliter (CFU/ml) was then calculated. Other specimens (B,C,D,E) were individually placed in sterile glass test tubes containing 10 ml sterile BHIB and treated identically to the negative control specimens. 7 Hardness study Preparation of Specimens: Twenty specimens were prepared from soft acrylic lining material (Vertex, Holland), 20mm in diameter and 12mm in thickness. 8 A total of 20 specimens were divided into four groups 6 : a) Five specimens were immersed in distilled water as a control group. b) Five specimens were coated in ginger oil for eight hours then immersed in distilled water. c) Five specimens were coated in Salvia officinalis oil for eight hours then immersed in distilled water. d) Five specimens were coated in eucalyptus oil for eight hours then immersed in distilled water.

Hardness test
The measurement of hardness of soft lining materials was carried out by using a Shore A hardness durometer, the measurement method is based on penetration of the specimens by an indentor under specified conditions.
The readings were taken 1 second after a firm contact was achieved three readings were taken for each sample and the mean of those readings was calculated. 9 The specimens were tested for hardness after different time intervals (1, 7, 14, and 30 days). For each specimen the measurements were recorded three times and mean values were calculated and compared statistically with one way Analysis of Variance and Duncan's multiple range test to determine the significant difference among the groups at p≤ 0.05 level of significance by using SPSS version 11.5.

RESULTS.
The results obtained from the study are shown in

CONCLUSION.
The soft acrylic lining material when coated with selected natural plant oils has antifungal activity as compared with uncoated material but there was no significant difference among the oils (Salvia officinalis, ginger and eucalyptus oils). In addition these oils increased the softness and duration of this material.