Many essential oils have antibacterial activity with a potential use in medicine. Citrus hystrix DC, or makrut lime, contains two essential oils, makrut leaf oil and makrut (fruit peel) oil, of which we determined the inhibitory effect against respiratory pathogens and evaluated their active components. Gas chromatography-mass spectrometry was used to analyse the chemical composition of the essential oils. The antibacterial activities were tested by disc-diffusion and broth microdilution methods against 411 isolates of groups A, B, C, F, G streptococci, Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus (methicillin-resistant and -sensitive S. aureus) and Acinetobacter baumannii, obtained from patients with respiratory tract infections. Makrut leaf oil and makrut oil were both effective against all the pathogens with minimal inhibitory concentration (MIC) ranges of 0.06-68 mg/ml and 0.03-17.40 mg/ml, respectively. Citronellal was found to be the major component (80.04%) in makrut leaf oil and had the lowest MIC. In contrast, makrut oil consisted of several components (limonene 40.65%, terpinene-4-ol 13.71%, α-terpineol 13.20%), and the most active component was α-terpineol, followed by terpinene-4-ol, and limonene. These results suggest that makrut leaf oil, makrut oil, and their components (citronellal, α-terpineol, terpinene-4-ol) may be alternative natural source medicine to prevent and treat many bacterial diseases.

Background and aim Kaffir lime fruit peel oil and Kaffir lime leaf oil have been reported for their activities against respiratory tract pathogens. The purpose of the study was to develop clear oral sprays to be used as a first-defense oral spray. Experimental procedure Clear antibacterial oral sprays were prepared and analyzed for their respective active major compounds, using GC-MS. The sprays were tested against a Gr. A streptococcal clinical isolate and 3 standard respiratory tract pathogens, using Broth microdilution method. A 4-month stability test was carried out as well. Results and conclusion Six clear oral sprays, three formulae composed of Kaffir lime fruit peel oil (6, 10, 13%v/v KLO) and the other three formulae containing Kaffir lime leaf oil (4, 8, 12%v/v KLLO), were developed. The active compounds in KLO were α-terpineol and terpinene-4-ol whereas that in KLLO was citronellal. All oral sprays exhibited antibacterial activity against one Group A streptococcal clinical isolate and three respiratory pathogenic pathogens, Staphylococcus aureus ATCC 29213, Streptococcus pneumoniae ATCC 49619, and Haemophilus influenza e ATCC 49247, among which the strongest activity was against H. influenza e ATCC 49247. The antibacterial activity of all oral sprays remained unchanged in an accelerated stability test, at 4, 30, and 45 °C under 75% relative humidity, throughout the 4-month storage.

Context: Citrus hystrix de Candolle (Rutaceae), an edible plant regularly used as a food ingredient, possesses antibacterial activity, but there is no current data on the activity against bacteria causing periodontal diseases. Objective: C. hystrix essential oil from leaves and peel were investigated for antibiofilm formation and mode of action against bacteria causing periodontal diseases. Materials and methods: In vitro antibacterial and antibiofilm formation activities were determined by broth microdilution and time kill assay. Mode of action of essential oil was observed by SEM and the active component was identified by bioautography and GC/MS. Results and discussion: C. hystrix leaves oil exhibited antibacterial activity at the MICs of 1.06 mg/mL for P. gingivalis and S. mutans and 2.12 mg/mL for S. sanguinis. Leaf oil at 4.25 mg/ mL showed antibiofilm formation activity with 99% inhibition. The lethal effects on P. gingivalis were observed within 2 and 4 h after treated with 4 Â MIC and 2 Â MIC, respectively. S. sanguinis and S. mutans were completely killed within 4 and 8 h after exposed to 4 Â MIC and 2 Â MIC of oil. MICs of tested strains showed 4 times reduction suggesting synergistic interaction of oil and chlorhexidine. Bacterial outer membrane was disrupted after treatment with leaves oil. Additionally, citronellal was identified as the major active compound of C. hystrix oil. Conclusions: C. hystrix leaf oil could be used as a natural active compound or in combination with chlorhexidine in mouthwash preparations to prevent the growth of bacteria associated with periodontal diseases and biofilm formation.

Toothbrushes are sometimes contaminated by micro-organisms that can cause infection. This study evaluates the efficacy of Citrus hystrix sprays compared with 0.12% chlorhexidine gluconate (CHX) for disinfecting bacteria on children's toothbrushes. The participants were 61 children with a high caries risk aged 8-11. Each subject received two toothbrushes to be used continually for 7 consecutive days. The first set of toothbrushes was used as a baseline without any disinfectant. The others were divided into 5 groups and sprayed with distilled water (control), 0.12% CHX, 6% makrut oil (MO), 10% MO, or 13% MO. Toothbrushes were placed vertically into sterile test tubes containing 8 ml of TYS20B medium and incubated overnight. Then we performed 10-fold serial dilutions from 10 −1 -10 −5 , and spread them on BHI or MS agars for investigating facultative bacteria, oral streptococci, andStreptococcus mutans. Results showed that for total facultative bacteria, 10% MO and 13% MO gave the highest reduction rate (100%) > CHX (88%) > 6% MO (81%). For S. mutans, 10% MO and 13% MO also showed the highest reduction rate (100%) > 6% MO (90%) > CHX (88%). For oral streptococci, 13% MO gave the highest reduction rate (100%) > 10% MO (91%) > CHX (88%) > 6% MO (69%). There was no significant difference between oral sprays and 0.12% CHX in the reduction of total facultative bacteria, oral streptococci, and S. mutans (p > 0.05).

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