Phytochemical Screening and Antibacterial Effect of Stevia Rebaudiana (Bertoni) Alcoholic Leaves Extract on Streptococcus Oralis (Dental Plaque?s Primary Colonizer)
Author(s): Manar Ibrahim Ahmed* and Hadeel Mazin
Background: Medicinal herbs have drawn the attention of many researchers due to their anti-microbial and health promoting properties, especially after the increase in the side effects of chemical mouthwashes. “Stevia raebaudiana bertoni”, a zero calorie sweetener native to South America, has been used by people for about 200 years to sweeten their drinks and treat some diseases. The anti-bacterial properties of stevia extracts were studied by many researchers, but none of them investigated the effect of stevia extract on Streptococcus oralis, a primary colonizer of dental plaque, which was addressed in this study.
Materials and methods: Supragingival plaque samples were cultured on MSA plate aerobically for 24 hours at 37°C. For identification and characterization of the isolated colonies, they were subjected to gram staining, hemolysis in Blood Agar Plate (BAP), catalase test, optochin sensitivity and Polymerase Chain Reaction (PCR) test. The alcoholic extract of stevia leaves was prepared by maceration with 70% ethanol in a ratio of 1:10 (gm plant/ml solvent). Four experiments were involved in this study, the first one concerning the sensitivity of S. oralis to the study extract starting from 16 mg/ml to 512 mg/ml, with 0.2% CHX as a positive control and distilled water as a negative one. While the second experiment involved the determination of MIC. The third one involved the determination of MBC, whereas the fourth experiment was done to determine some of the extract active ingredients by using High Performance Liquid Chromatography (HPLC) analysis.
Results: The diameter of inhibition zone for S. oralis increased with increasing the concentration of alcoholic extract of Stevia Rebaudiana bertoni and it is highest in chlorohexidine with significant difference (p<0.05). The MIC and MBC were 16 mg/ml and 32 mg/ml respectively. HPLC analysis revealed the presence of phytochemicals with antimicrobial activity including: Dihydroxy streptomycin: 0.86 ppm, rutin: 28.1 ppm, quercetin: 22.4 ppm and apigenin: 4.45 ppm.
Conclusions: The alcoholic extract of Stevia rebaudiana bertoni leaves had good antibacterial activity against S. oralis. 0.2% CHX demonstrated better antibacterial activity than the study extract, 512 mg/ml of stevia extract had comparable effect to 0.2% CHX. HPLC analysis of Stevia rebaudiana bertoni alcoholic leaves extract revealed the presence of many active antibacterial constituents.