|Year : 2021 | Volume
| Issue : 4 | Page : 357-363
Efficacy of tea tree, sesame, and cumin extracts against oral Candida species – An in Vitro study
Keerthana Selvam, Suman J Lakshmi, Elangovan Somasundaram, Senthil Kumar Balasubramanian, Bakiyalakshmi Balasubramanian
Department of Oral Medicine and Radiology, KSR Institute of Dental Science and Research, Tiruchengode, Tamil Nadu, India
|Date of Submission||07-Jan-2021|
|Date of Decision||11-Aug-2021|
|Date of Acceptance||11-Sep-2021|
|Date of Web Publication||27-Dec-2021|
Dr. Keerthana Selvam
Department of Oral Medicine and Radiology, K. S. R Institute of Dental Science and Research, K. S. R Kalvi Nagar, Tiruchengodu, Namakkal, Tamil Nadu
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background and Objective: Oral cavity ports have diverse, abundant, and complex microbial communities. Candida is (a genus of yeast) and is the most common cause of oral fungal infection. The standard treatment of oral candidiasis is the use of antifungal agents, which cause different adverse effects. To combat this, herbal drugs can be considered as an alternative to allopathic drugs. The literature has reported that these herbal products are a rich source of antimicrobial agents. The present study determined the antifungal efficacy of tea tree, sesame, and cumin extracts against oral candidiasis in comparison to clotrimazole. Material and Methods: An in vitro study was carried out on 30 patients, wherein saliva and swab samples were collected and subjected to culture on chromagar and incubated at 37°C for 24 to 48 hours. The cultured Candida species were subjected to an antifungal susceptibility test. Clotrimazole (Group I: positive control), tea tree extracts (Group II–Group VI), sesame extracts (Group VII–Group XI), and cumin extracts (Group XII–Group XVI) were inoculated in wells and incubated. Zones of inhibitions were measured and subjected to statistical analysis. Results: Antifungal efficacy of tea tree (Group II–VI), sesame (Group VII–IX), cumin (Group XII–XIV), and clotrimazole (Group I) were compared and proven to be statistically significant with a P value < 0.05. Conclusion: Clotrimazole had a higher antimicrobial activity compared to medicinal plants. The extracts of tea tree, sesame, and cumin had significant antimicrobial activity and they might prove to be promising alternatives as a therapeutic agent for oral candidiasis.
Keywords: Extracts and antimicrobial activity, oral Candida species
|How to cite this article:|
Selvam K, Lakshmi SJ, Somasundaram E, Balasubramanian SK, Balasubramanian B. Efficacy of tea tree, sesame, and cumin extracts against oral Candida species – An in Vitro study. J Indian Acad Oral Med Radiol 2021;33:357-63
|How to cite this URL:|
Selvam K, Lakshmi SJ, Somasundaram E, Balasubramanian SK, Balasubramanian B. Efficacy of tea tree, sesame, and cumin extracts against oral Candida species – An in Vitro study. J Indian Acad Oral Med Radiol [serial online] 2021 [cited 2022 May 20];33:357-63. Available from: https://www.jiaomr.in/text.asp?2021/33/4/357/333878
| Introduction|| |
“Your diet as medicine does more wonders than the drugs as medicine.” Authenticating this statement, bioceuticals and their role have advanced in the management of oral diseases. The use of topical and systemic drugs has had detrimental effects on oral health. To overcome these, plant extracts are alternative therapeutic agents with limited adverse effects.,
Tea tree oil is a bioceutical with meritorious therapeutic values. Cumin is an aromatic plant, pharmacological studies have revealed that it has an antimicrobial property., Antimicrobial screening done using sesame extracts showed that it is active against Candida species.
The present study was designed to comprehensively study the antifungal potential of the tea tree, cumin, and sesame extracts against the oral Candida species. Hence, it is conceptualized to assess the in vitro antifungal efficacy of tea tree, cumin, and sesame extracts on Candida species in comparison to allopathic drugs.
| Material and Methods|| |
Patients attending the Department of Oral Medicine and Radiology in a private dental college between 2017 and 2018 were screened, and those who were diagnosed and confirmed by clinical examination to be suffering from oral candidiasis were enrolled in the present study. This study was approved by the Institutional Ethics Committee (Ref no: 183/KSRIDSR/EC/2017 on 18.12.2017), and written consent was obtained from the patients before including them in the study. All the procedures were in accordance with 'Helsinki declaration' maintaining the standard ethical principles.
Sample size Estimation
The sample size estimation was done using G* power statistics, software version 184.108.40.206 based on parameters or data from the previous literature. Using this software, the sample size was estimated to be 25. To eliminate errors and attrition in the clinical trial, the sample size was fixed to 30 per group. The study subjects were divided into groups with a simple randomization procedure (table of random numbers).
Inclusion and exclusion criteria
- Subjects with candidiasis who were willing to participate in the study were included.
- Denture-wearing patients were included.
- Patients who were under antifungal medications were excluded.
- Severally immunocompromised patients were excluded.
- Unwilling patients were excluded from the study.
Saliva and swab collection
A total of 30 patients were selected and their saliva samples were obtained with proper consent. A sterile cotton swab was gently rubbed over the lesion. Oral rinse was collected by holding 10 mL of phosphate-buffered saline (0.01 M, pH 7.2) in the patient's mouth for 1 minute, using a sterile container. Unstimulated saliva of the patient with oral Candida infections, such as chronic atrophic candidiasis and erythematous candidiasis, was collected in a sterile container.
Isolation of Candida organism
The collected samples were kept in sterile containers and brought to the laboratory immediately for further processing. Both swab and saliva samples were streaked on a selective medium (CHROMagar) and incubated at 37°C for 24 to 48 hrs. After incubation, different types of Candida colonies such as those of Candida albicans, Candida tropicalis, Candida krusei, and Candida glabrata were obtained in different colors [Figure 1] and [Figure 2], [Table 1].
Preparation of extracts
Cumin seeds, tea tree leaves, and sesame roots were cleaned and dried at room temperature for 2 days and ground into a fine powder using a mechanical grinder, and stored in an air-tight sterile container separately. The aqueous extracts of all three were prepared by cold extraction. For 5% aqueous extract, 5 g of the powder was mixed with 100 mL of sterile water and kept in a shaker for 5 days. Then, the extract was filtered using a Whatman filter paper, concentrated using a vacuum evaporator, and stored for further use in a sterile container. Similarly, 10%, 15%, 20%, and 25% of extracts were prepared [Figure 3].
There was three study groups (tea tree, sesame, and cumin) and one control group (clotrimazole). Each study group was further divided based on different concentrations of the extracts. Clotrimazole (Group I: positive control), tea tree extract (Group II–Group VI), sesame extract (Group VII–Group XI), and cumin extract (Group XII–Group XIV).
Activity of extracts on Candida species
The antimicrobial activity of the extracts was studied using the “well diffusion method.” A Mueller–Hinton agar plate was prepared and 24 to 48 hrs freshly cultured brain heart infusion broth of Candida species was swabbed on the surface. Wells (about 8 mm) were punched into the agar plates using a sterile cork borer. About 100 μL of aqueous extracts of cumin, sesame, and tea tree, with concentrations of 5, 10, 15, 20, and 25% were added in respective wells and incubated at 37°C for 24 to 48 hrs. A clotrimazole antifungal disc (1%) was used as the standard. All tests were carried out in triplicate. After the incubation period, clear zones of inhibition (mm) were measured using a digital Vernier Caliper and compared with the control zones [Table 2]a, [Table 2]b, [Table 2]c, [Figure 4]a, [Figure 4]b, [Figure 4]c.
|Table 2: (a) Values of antimicrobial activity (zone of inhibition) of varying concentrations of tea tree extracts against oral Candida species. (b) Values of antimicrobial activity (zone of inhibition) of varying concentrations of sesame extracts against oral Candida species. (c) Values of antimicrobial activity (zone of inhibition) of varying concentrations of cumin extracts against oral Candida species.|
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|Figure 4: (a) Antimicrobial activity of varying concentrations of tea tree extracts against oral Candida species.(b) Antimicrobial activity of varying concentrations of sesame extracts against oral Candida species. (c) Antimicrobial activity of cumin extracts of varying concentrations against oral Candida species.|
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The data were compiled in MS Office Excel. The gathered data were analyzed using the IBM SPSS Statistics for Windows, version 25.0. Descriptive and inferential statistics were used to summarize the results. Analysis of variance (ANOVA) test was used to compare the mean number of inhibition zones among the groups. Multiple comparisons between the groups were done using the posthoc Tukey's HSD (honestly significant difference) test.
| Results|| |
The comparisons of inhibition zones within the group and between the groups at different concentrations for all three groups were tabulated and depicted that the antifungal efficacy increased with an increase in the concentration.
The mean value of zone of inhibition of five different concentrations of tea tree was, 5.5 mm, 6.27 mm, 6.97 mm, 8.07 mm, 9.18 mm at 5%, 10%, 15%, 20%, and 25%, respectively, and positive control (clotrimazole) had a mean value of 12.1 mm [Table 2]a, [Table 3]a and Graph 1]. For sesame, it was 8.33 mm, 8.87 mm, 9.17 mm, 10.07 mm, and 11.57 mm at 5%, 10%, 15%, 20%, 25%, respectively, and the positive control (Clotrimazole) had a mean value of 17.17 mm [Table 2]b, [Table 4]a and [Graph 1]. For cumin, it was 4.07 mm, 4.97 mm, 5.63 mm, 6.73 mm, and 7.80 mm at 5%, 10%, 15%, 20%, and 25%, respectively, and the positive control (clotrimazole) had a mean value of 11.33 mm [Table 2]c, [Table 5]a, and [Graph 1]. All three extracts showed antifungal property at higher concentrations (25% of tea tree [Group VI], sesame [Group XI], and cumin [Group XVI] concentrations, P values = 0.001). The mean values of all extracts were lesser than those of the control group. There was slight significant difference observed between 25% tea tree (P value = 0.018) and 25% sesame (P value = 0.023) [Table 3]b and [Table 4]b.
|Table 3: (a) Comparison of means and standard deviations of clotrimazole (control) to those of 5%, 10%, 15%, 20%, and 25% tea tree extract against oral Candida species. (b) Comparison of the mean difference of clotrimazole (control) with that of 5%, 10%, 15%, 20%, and 25% tea tree extracts.|
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|Table 4: (a) Comparison of means and standard deviations of clotrimazole and 5%, 10%, 15%, 20%, and 25% sesame extracts against oral Candida species. (b) Comparison of the mean difference of clotrimazole with 5%, 10%, 15%, 20%, and 25% sesame extracts.|
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|Table 5: (a) Comparison of means and standard deviations of clotrimazole and 5%, 10%, 15%, 20%, and 25% cumin extracts against oral Candida species. (b) Comparison of mean difference of clotrimazole with 5%, 10%, 15%, 20%, and 25% cumin extracts.|
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| Discussion|| |
Diseases of the oral cavity have been of concern from pre-historical times. One such disease of concern is oral candidiasis with historical evidence from the fifth century BC when it was first discovered by Hippocrates. Thereafter, numerous studies have been conducted to find a cure for the disease. Although there are known regimens for the treatment of candidiasis, various researches are being conducted to explore the possibilities of using herbal supplements as alternatives for the treatment of the ailment.
The Indian subcontinent is the motherload of innumerable precious plants with therapeutic properties. Indians have been using the therapeutic properties of these plants for ages, right from ancient times. In the present study, three such inestimable plants that are used invariably in day-to-day activities in Indian households were used to study their individual actions on Candida species, and their efficacy was compared.
Recent studies have revealed that tea tree, cumin, and sesame possess antifungal, antibacterial, antiviral, and antiprotozoal activities.
A comparative analysis was done by Sharma et al., with three medicinal plants Glycyrrhiza glabra, Ficus religiosa, and Plantago major. He used 5 mL of the preparation of all three plants, and the results were analyzed at 24, 48, and 72 hrs and compared with clotrimazole, fluconazole, and itraconazole. The present study included five different concentrations of the extracts and results were obtained after 48 hrs of incubation. Sharma et al. inferred that G. glabra showed lesser activity than clotrimazole at 24 hrs, and the difference in the activity was found to be statistically insignificant. F. religiosa and P. major showed a statistically insignificant difference in the activity compared to itraconazole at 24 hrs.
In the present study, similarly, three extracts, namely tea tree, sesame, and cumin were taken. The results inferred that tea tree, sesame, and cumin extracts in varying concentrations showed lesser activity when compared to clotrimazole. Statistically, the P value (0.001) was significant for all concentrations of tea tree extracts. There was a slight difference in the P value (0.018) when 25% of tea tree extracts were used [Table 3]b. Similar results were obtained with sesame extracts, with a P value of 0.001 for all concentrations [Table 4]b, except for the group where 25% of sesame extracts were used (P value = 0.023). The P value (0.001) was significant for all concentrations of cumin extracts [Table 5]b.
Jain et al. conducted an in vitro study using Aloe vera and Triphala. The results showed that the mean value of zone of inhibition by A. vera was 3.35 ± 0.59 mm and 1.06 ± 0.41 mm at 100% and 50%, respectively, whereas for Triphala, it was 4.19 ± 0.57 mm and 1.79 ± 0.43 mm at 100% and 50%, respectively. Both Triphala and A. vera showed the antifungal property at higher concentrations (100% and 50% concentration, P value < 0.001).
The present study revealed that the mean values of zones of inhibition by tea tree extracts were 5.5, 6.27, 6.97, 8.07, and 9.18 at 5%, 10%, 15%, 20%, and 25%, (Group II–Group VI), respectively. On comparing five different concentrations, 25% tea tree extract showed a higher efficacy with statistically significant P values. For sesame extracts, it was 8.3, 8.87, 9.17, 10.07, and 11.5 at 5%, 10%, 15%, 20%, and 25%, respectively. On comparing five different concentrations, 25% sesame extract showed a higher efficacy with significant P values. For cumin extracts, it was 4.07, 4.97, 5.63, 6.73, and 7.80 at 5%, 10%, 15%, 20%, and 25%, respectively. On comparing five different concentrations, 25% cumin extract showed a higher efficacy with a statistically significant P value. Positive control (clotrimazole) had higher mean values. The mean values of all extracts were lesser than those of the control group. As the concentration of the extract increased, the mean value was found to increase.
Sharma et al.,, who studied the crude extract of Terminalia chebula fruit and seed demonstrated its activity against C. albicans. The zone of inhibition was 9 mm with an activity index of 0.28 mm. Similarly, in our present study, 25% cumin extract and 15% sesame extract showed similar inhibitory concentrations with zones of inhibition of 9.2 mm and 9.1 mm, respectively.
In the present study, when comparing 5% tea tree, sesame, and cumin extracts, 5% sesame had a higher efficacy with all P values being statistically significant (0.001). On comparing 15% tea tree, sesame, and cumin extracts, 15% sesame had a higher efficacy, with all P values being statistically significant (0.01). When comparing the extracts of 20% and 25% tea tree, sesame, and cumin extracts, 20% and 25% of sesame and tea tree extracts had significant P values of 0.023 and 0.018, respectively.
| Limitations and Future Recommendations|| |
Discovering new knowledge is not the only objective of the research process but also to confound assumptions and explore the limitations that we encounter. One of the major limitations of the study was the small sample size. Results can be better conceptualized if they involved a larger sample group. Another limitation was that only cases with chronic atrophic candidiasis and erythematous candidiasis were included; so, the efficacy of these products in the treatment of another form of Candida infections has to be studied. Using an herbal extract for the treatment of oral candidiasis is a new treatment modality for oral candidiasis, and the effects of the herbal extracts are still under research. This study was an in vitro study, more trials should be carried out on large populations (in vivo) in the future. Also, different herbal extracts with varying concentrations have to be improvised by further research for effective treatment.
| Conclusion|| |
New milestones in the development of pharmaceutical products can be achieved by discovering bioactive natural products from medicinal plants to address unfulfilled therapeutic needs against these fungal infections. Based on the results of the present study, clotrimazole had a higher antimicrobial activity when compared to medicinal plants, even though extracts of tea tree, sesame, and cumin extracts had antimicrobial activity and it might be possible to use them as therapeutic agents for oral candidiasis. Thus, in view of the present scenario, judicious use of such natural products might not only help to minimize the deleterious effects of synthetic drugs but also prove to be cost-effective, with less toxic effects, especially in developing countries.
Declaration of patient consent
The authors certify that they have obtained all appropriate patient consent forms. In the form, the patient(s) has/have given his/her/their consent for his/her/their images and other clinical information to be reported in the journal. The patients understand that their names and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]