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 Table of Contents  
ORIGINAL RESEARCH
Year : 2021  |  Volume : 13  |  Issue : 1  |  Page : 17-23

Efficacy of 3% Psidium guajava local drug delivery in the treatment of chronic periodontitis: A randomized controlled trial


1 Department of Periodontology, JSS Dental College & Hospital, JSS Academy of Higher Education & Research, Mysuru, India
2 Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Mysuru, India
3 Department of Biotechnology and Bioinformatics, Department of Water and Health, JSS Academy of Higher Education & Research, Mysuru, India

Date of Submission24-Jul-2020
Date of Decision29-Oct-2020
Date of Acceptance23-Nov-2020
Date of Web Publication28-Jan-2021

Correspondence Address:
Dr. H Manohar Sharma
Department of Periodontology, JSS Dental College & Hospital, JSS Academy of Higher Education & Research, Mysuru.
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jioh.jioh_249_20

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  Abstract 

Aim: Herbal products are increasingly used as therapeutic agents in the prevention and management of periodontal diseases to preclude the side effects of antibiotics. A recent addition to the list is the leaves of P guajava, as they possess antimicrobial, anti-inflammatory, and antioxidant properties. This study evaluates the effects of the adjunctive use of 3% P. guajava local drug delivery in the management of chronic periodontitis. Materials and Methods: This was a split-mouth study involving contralateral sites in 15 patients (30 sites), who were randomly treated with either 3% P. guajava gel local drug delivery along with scaling and root planing (SRP) or SRP alone. The primary objective was to evaluate pocket probing depth (PPD), clinical attachment level (CAL), and colony counts of A.actenomycetemcomitans (Aa) and P.gingivalis (Pg); the secondary objective was to evaluate plaque, gingival, and bleeding scores. All parameters were assessed at baseline, one month and three months. The data were analyzed by using SPSS 18.5. Results: There was a significant overall improvement in clinical parameters over the study period. There was a statistically significant reduction in site-specific indices, PPD (2.74 ± 0.283), and gain in CAL (2.8 ± 0.152) in the test sites at three months. Microbiological analysis showed a significant reduction in the colony counts of Aa (17.4 ± 0.026) and Pg (22.7 ± 1.225) in the test sites at one and three months. (P < 0.001). Conclusion: Improvements in clinical and microbiological parameters showed that adjunctive use of locally delivered 3% P. guajava gel is effective in the management of chronic periodontitis.

Keywords: Chronic Periodontitis, Clinical and Microbiological Parameters, P. guajava Gel, SRP


How to cite this article:
Manohar Sharma H, Deepika P C, Venkatesh M P, Chandan S, Shashikumar P. Efficacy of 3% Psidium guajava local drug delivery in the treatment of chronic periodontitis: A randomized controlled trial. J Int Oral Health 2021;13:17-23

How to cite this URL:
Manohar Sharma H, Deepika P C, Venkatesh M P, Chandan S, Shashikumar P. Efficacy of 3% Psidium guajava local drug delivery in the treatment of chronic periodontitis: A randomized controlled trial. J Int Oral Health [serial online] 2021 [cited 2021 Mar 3];13:17-23. Available from: https://www.jioh.org/text.asp?2021/13/1/17/308359


  Introduction Top


Periodontitis is considered a multifactorial infectious disease in which the primary etiological factor comprises pathogenic micro-organisms within the subgingival biofilm. The oral cavity has a diverse yet stable community of oral microbiota and if there is an imbalance, oral diseases such as caries and periodontal diseases seem to appear.[1] Specific periodonto-pathogenic bacterial species and their virulence factors have been recognized as etiological agents and they affect the rate of progression of the disease. Foremost among these are gram-negative organisms, including Aggregatibacter actinomycetemcomitans (Aa) and Porphyromanas gingivalis (Pg).[2]

In the past years, systemic and local antibiotics were proposed as an adjunct to SRP. However, side effects such as resistance, toxicity, sensitivity, and growth of opportunistic infection have altered the general opinion about their effectiveness.[3] Research in phytosciences, an emerging multidisciplinary science, has revealed various medicinal plants possessing antimicrobial activity with fewer side effects and reduced toxicity. Extracts of these medicinal plants offer a new choice for optimal antimicrobial therapy against various oral microorganisms.[4] Plants produce several secondary metabolites, many of which present important biological activities, and in particular, antimicrobial activity. Plant extracts are potent drugs, because they interact with specific chemical receptors in the body.[5]

Psidium guajava L., commonly known as guava, has been used in folk medicine as a therapeutic agent. The leaves of the plant P. guajava are reported to possess antimicrobial, anti-inflammatory, and antioxidant properties.[6] Guava contains a large number of antioxidants and phytochemicals, including essential oils, polysaccharides, minerals, vitamins, enzymes, triterpenoid acid alkaloids, steroids, glycosides, tannins, flavonoids, and saponins. The flavonoid compounds and their derivatives can inhibit the growth of different bacteria by disruption of membrane and inactivation of extracellular proteins by forming irreversible complexes.[7] The antibacterial activity has been shown to be more pronounced against gram-positive bacterial strains and show moderate activity against gram-negative bacterial strains.[8] The anti-inflammatory action of guava is in its ability to inhibit prostaglandin, kinin, and histamine. Guava leaf extracts helps in decreasing CRP levels, which plays a role as modulator of inflammatory response Fermented guava leaf extract is involved in the inhibition of inducible nitric oxide synthase and cyclooxygenase-2. It also inhibited lipopolysaccharide-induced NF-kβ activation, which can be used as a potential strategy for preventing inflammatory bone resorption in treating periodontal disease.[9]

Guava is an excellent antioxidant and a good source of vitamin C. Guava leaf extracts have the ability to scavenge free hydrogen peroxide, superoxide anion radical and inhibit the formation of hydroxyl radical. The antioxidant action can be attributed to quercetin, carotenoids, vitamin C, and polyphenols present in guava.[10],[11] Thus, guava could serve as an antioxidant-based approach to periodontal therapy. In vivo studies evaluating the efficacy of P. guajava mouthwash on oral hygiene were carried out and improvement in the same was seen.[12] However, there have been no studies on its use as a local drug delivery in the management of periodontitis. Hence, this study was carried out to assess the effects of P. guajava as local drug delivery along with SRP in the treatment of chronic periodontitis.


  Materials and Methods Top


Setting and design

This was a randomized controlled trial in which a total of 15 patients who had given consent and fulfilled inclusion criteria were selected by purposive sampling, from the outpatients presenting to the Department of Periodontology of our institution. The selection happened over a period of six months from November 2018 to April 2019.

Chronic periodontitis patients aged 35–55 years with ≥20 remaining teeth and contralateral pockets measuring ≥5 mm with evident clinical attachment loss and radiographic evidence of bone loss were selected. Patients who had undergone any periodontal therapy or taken any antibiotics in the past six months, patients on medications known to affect the periodontium, patients with debilitating diseases, pregnant/ lactating women, smokers, and patients undergoing orthodontic treatment were excluded. Subjects who fulfilled the inclusion criteria and who complied with adequate plaque control measures were included.

Based on previous study results,[13] using a power of 90% with a mean difference of 3 and a standard deviation of 1.4, calculated sample size was found to be 15 with 30 sites (including attrition rate of 20%). Sample size was calculated by using the formula for “Comparison of Means”, i.e.



In each patient, the treatment sites were randomly divided into the test and control sites by a computer-generated random table [Figure 1]. Site A (Control site): SRP, Site B (Test site): SRP followed by local application of 3% P. guajava gel.
Figure 1: Flowchart showing selection of study population

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Intra-examiner calibration

A single examiner masked to the treatment procedure recorded all the clinical parameters. Before the study, 20 sites were examined 24 hours apart to achieve examiner calibration. Calibration was accepted if there was a difference of 1 mm with a 95% significance level between the two measurements.

Clinical parameters

Plaque Index (Silness and Loe), Gingival Index (Loe and Silness), Gingival Bleeding Index (Ainamo and Bay, 1975), probing pocket depth (PPD), and clinical attachment level (CAL) were recorded at baseline, one month and three months.

Acrylic stents were made to guide the probe penetration vertically in the same plane every time it was inserted for recording the measurements. The apical margin of the stent was used as a fixed reference point (FRP), and clinical measurements were made at the proximal line angle of the tooth [Figure 2]E. The following measurements were recorded at baseline and all follow-up recall intervals:
Figure 2: (A, B) Plaque sample collection; (C, D) 3% P. guajava gel insertion; (E) pre-op probing depth; (F) 3 months post-op probing depth

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  1. Stent (FRP) to Cemento-enamel Junction (CEJ)


  2. Stent (FRP) to Gingival Margin (GM)


  3. Stent (FRP) to base of the pocket (BP)


Based on the measurements cited earlier, PPD and CAL were calculated as follows:

  • PPD = FRP to BP – FRP to GM


  • CAL = FRP to BP – FRP to CEJ


  • Microbiological analysis

    Microbiological analysis for identification and estimation of colony counts of Aa and Pg was carried out at baseline, one month and three months. The subgingival plaque samples were collected by using a sterile Gracey curette and were transferred to an Eppendorf tube containing thioglycolate broth [Figure 2A] and [B]. Anaerobic culturing of the plaque samples was done by using TSBV agar and blood agar for Aa and Pg, respectively. Aa was identified under UV-light microscopy as white, translucent, smooth, non-hemolytic colonies with a star-shaped internal structure. Pg was identified as greenish black colonies on blood agar, gram-negative rods under gram staining and there was no fluorescence under UV-light microscopy. Confirmatory tests such as indole test, glucose and lactose fermentation tests were carried out.

    Formulation of 3% P. Guajava in situ gel

    Materials used:

  • P. guajava leaf extract


  • Distilled water


  • Pluronic F-127 (PF-127)


  • Triethanolamine


  • Benzalkonium chloride


  • The guava leaf extract was procured from Phytotech Extracts Pvt Ltd., Bangalore.

    Minimum inhibitory concentration (MIC) of Psidium guajava against periodontal pathogens was assessed by using the agar well diffusion method. An antibiotic disc dispenser was used to dispense discs containing specific antibiotics (streptomycin) as well as different concentrations of Psidium guajava. The zone of inhibition was found to be 25 mm for streptomycin, 20 mm for 2% P. guajava gel, and 33 mm for 3% P. guajava gel. The 3% P. guajava gel showed a greater zone of inhibition indicating better antimicrobial activity and, hence, this concentration was used in the current study.

    The gel was prepared by the “Cold Technique.” Pluronic F-127 was added to cold distilled water and refrigerated at 5°C for 10 hours to form a thermosetting gel. 3% P. guajava was added to this and left in the quiescent state in the refrigerator overnight for complete solubilization. A dilute solution of triethanolamine was added to adjust the pH to 7.0, and benzalkonium chloride was added as preservative. The gel was then collected and stored in sealed glass vials, and sterilization was carried out by exposing it to UV radiation of 265 nm over a period of 20 min.

    Local drug delivery

    At the test sites, 3% P. guajava gel was delivered into the periodontal pocket by using a blunt cannula [Figure 2C] and [D]. A firm finger pressure was applied for 2 min to allow the gel to flow uniformly through the pocket. Excess gel that flowed out of the pocket was wiped away by using cotton pellets.

    Oral home care consisting of normal brushing and rinsing was started after 24 h and carried out till the end of the study period. Patients were also asked to refrain from taking any antibiotics during the study period. Oral hygiene instructions were reinforced for all the patients at every follow-up interval to motivate patients for optimal oral hygiene maintenance.

    Statistical analysis

    The statistical package for social science (IBM SPSS, version 18.5, USA) for windows software was used. The normality of the data was assessed by using the Shapiro–Wilk test. The data for PPD and CAL were not normally distributed, so a comparison between the groups was carried out by using nonparametric tests. Kruskal–Wallis test was applied to find out the intragroup difference between PPD and CAL. Mann–Whitney U test was used to find out the intergroup difference between PPD and CAL. One-way ANOVA was used to test the intragroup differences in indices, colony counts of Aa and Pg. The Student’s t-test was used to determine the intergroup differences in indices, colony counts of Aa and Pg. P < 0.05 was considered significant.


      Results Top


    A total of 30 sites in 15 patients (nine males and six females) with a mean age of 40.67 years were assessed. Post-treatment healing was uneventful, and no adverse complications were reported during the procedure or at subsequent postoperative appointments. At baseline, there was no statistically significant difference in any of the parameters assessed. (P > 0.05)

    There was a significant reduction in full-mouth scores of all the indices, PPD and gain in CAL gradually from baseline to three months [Figure 2F], thus indicating an overall improvement in gingival and periodontal health [Table 1]. The site-specific analysis showed a significant reduction in the indices, PPD and gain in CAL at one month and three months, where the test sites showed better improvement (P < 0.001) [Table 2].
    Table 1: Full-mouth scores at various time intervals

    Click here to view
    Table 2: Site-specific scores at various time intervals

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    The microbiological analysis showed a significant reduction in the colony counts of Aa and Pg at one month and three months, with the test sites showing more reduction in the colony counts (P < 0.001) [Table 3].
    Table 3: Colony counts of Aa and Pg (×105) at various time intervals

    Click here to view



      Discussion Top


    The current study evaluated the efficacy of 3% P. guajava gel application along with SRP, in comparison with SRP alone, on the clinical and microbiological parameters in chronic periodontitis. This was a comparative study, with split-mouth design carried out over a period of three months. The split-mouth design was adopted to avoid the inter-patient variability, as paired defects were treated in the same subject. The primary objective was to evaluate the PPD, CAL, and colony counts of Aa and Pg; whereas the secondary objective was to evaluate plaque, gingival, and bleeding scores. All the parameters were assessed at baseline, one month and three months.

    Antimicrobial therapy directed at specific bacteria has been often used to augment mechanical periodontal therapy. Various antibiotics are used as local drug delivery agents in distinct phases in a periodontal treatment plan, such as an adjunct to SRP, periodontal maintenance therapy and for whom surgery is not an option, or those who refuse surgical treatment. However due to certain disadvantages such as bacterial resistance, undesirable side effects, growth of opportunistic infection, and interaction with other medications, newer agents such as plant extracts have been studied recently.

    Psidium guajava is a well-known traditional medicinal plant used in various indigenous systems of medicine. The leaves and bark of P. guajava tree have a long history of medicinal uses, which is still employed today. Several in vitro studies have been carried out to assess the phytochemistry and the properties of P. guajava. It has been shown that guava leaves contain flavonoids, mainly the quercetin derivatives. They have strong antibacterial, anti-inflammatory, analgesic, and antioxidant activities.[8],[14],[15],[16],[17] The antimicrobial efficacy and MIC of P. guajava was analyzed by using the agar well diffusion method, as previously explained. 3% P. guajava gel showed a greater zone of inhibition, indicating better antimicrobial activity, and hence this concentration was used in the current study. There was a significant reduction in full-mouth scores of plaque index (1.36 ± 0.04), gingival index (1.29 ± 0.047), gingival bleeding index (47.5±1.162), and PPD (0.71 ± 0.024 mm) and gain in CAL (0.72 ± 0.052 mm), at various time intervals. Thus, these indicated an overall improvement in gingival and periodontal health.

    Site-specific assessment of indices was also carried out to assess the effects of 3% P. guajava gel on the gingival health. At test sites, there was a significant decrease in the scores of plaque index (1.63 ± 0.018), gingival index (1.5 ± 0.032), and bleeding index (56.4 ± 3.79). The control sites also showed a significant decrease in the scores of plaque index (1.53 ± 0.002), gingival index (1.4 ± 0.020), and bleeding index (47.9 ± 2.32). Between the sites, the results were significant at three months, with test sites showing a greater reduction in all the indices (P < 0.001).

    The observed anti-inflammatory effect of guava can be attributed to the numerous polyphenolic compounds, triterpenoids, and other chemical compounds present in the guava leaf extracts. It exerts anti-inflammatory activity by inhibiting prostaglandin, kinin, and histamine.[18] Clinical evaluation of PPD at test sites showed a reduction in PPD of 3.87 ± 0.197 at one month and 4.20 ± 0.009 at three months. At the control sites, it was 2.27 ± 0.144 at one month and 2.74 ± 0.283 at three months. All the sites showed a significant reduction in PPD over a period of three months. Between the sites, the results were significant at three months, with test sites showing a greater reduction in PPD (P < 0.001). The gain in CAL at the test site was 2.53 ± 0.188 at one month and 2.8 ± 0.152 at three months. At control sites, it was 1.6 ± 0.023 at one month and 1.8 ± 0.094 at three months. All the sites showed significant gain in CAL over a period of three months. Between the sites, the results were significant at three months, with test sites showing greater gain in CAL (P < 0.001).

    Guava is rich in vitamin C (ascorbic acid).[11] Ascorbic acid, through its effects on extracellular matrix, alters fibroblast differentiation and modulates expression of procollagen gene, leading to collagen formation.[19] Vitamin C along with bioflavonoids also helps in speeding up the healing process. All these properties can be attributed to the observed improvement in the clinical parameters in the current study. Apparently, there have been no studies performed to evaluate the effects of 3% P. guajava gel as a local drug delivery agent in the treatment of chronic periodontitis. Hence, we are comparing our study with other similar herbal products.

    Warad et al. evaluated locally delivered 2% lemongrass essential oil as an adjunct to SRP for the treatment of chronic periodontitis. The mean PPD reduction was 1.43 ± 0.67 mm and 2.40 ± 0.73 mm and the mean RAL (relative attachment level) gain was 1.66 ± 0.81 and 3.20 ± 0.75 mm at one and three months, respectively.[20] Singh et al. evaluated chlorhexidine chip and turmeric chip as local drug delivery in the management of chronic periodontitis. In the turmeric chip group, the mean PPD reduction was 1.2 ± 0.14 mm and 1.07 ± 0.07 mm and the mean RAL gain was 1.12 ± 0.13 and 1.1 ± 0.05 mm at one and three months, respectively.[21] Grover et al. investigated subgingival use of E. officinalis (Amla) sustained-release gel adjunctive to SRP in chronic periodontitis. The mean PPD reduction was 2.484 ± 0.432 mm and 3.096 ± 0.491 mm and the mean CAL gain was 2.423 ± 0.681 mm and 3.088 ± 0.782 mm at two and three months, respectively.[22] Boyapati et al. evaluated local application of Achyranthes aspera gel in the management of chronic periodontitis. The mean PPD reduction was 2.24 ± 0.85 mm and mean CAL gain was 0.73 ± 0.40 mm at three months.[23] The results in the current study are similar to the earlier mentioned studies where herbal agents are used and clinical effects are evaluated at one and three months.

    On microbiological analysis, the reduction of Aa counts at test sites was 14.5 ± 0.228 at one month and 17.4 ± 0.026 at three months. At the control sites, the reduction in Aa counts was 11.6 ± 0.112 at one month and 13.3 ± 0.333 at three months. All the sites showed a significant reduction in Aa counts over a period of three months. Between the sites, the results were significant at three months, with test sites showing a greater reduction (P < 0.001). The reduction of Pg counts at test sites was 21.2 ± 1.069 at one month and 22.7 ± 1.225 at three months. At the control sites, the reduction in Pg counts was 16.6 ± 0.443 at one month and 17.6 ± 2.221 at three months. All the sites showed a significant reduction in Pg counts over a period of three months. Between the sites, the results were significant at three months, with test sites showing greater reduction (P < 0.001).

    Flavonoids, guajaverin, and quercetin present in guava exert its antimicrobial activity. It has been shown that A. actinomycetemcomitans, P. gingivalis, P. intermedia, and F. nucleatum are susceptible to quercetin. Quercetin disrupts the membrane and inactivates extracellular proteins by forming irreversible complexes, thereby exerting its antibacterial action.[24]

    There are several studies where anaerobic culturing techniques have been utilized to analyze subgingival plaque samples, which are similar to the current study. Bhatia et al. evaluated the efficacy of locally delivered 1% curcumin gel in the treatment of chronic periodontitis. The microbiological analysis showed a significant reduction in the counts of P. gingivalis, P. intermedia, F. nucleatum, and capnocytophaga at test sites after six months when compared with that of control sites (SRP alone).[25] Sanghani et al. studied the efficacy of subgingivally administered Indian propolis extract in the treatment of periodontitis. Anaerobic culturing of the subgingival plaque samples using blood agar and enriched agar medium showed a significant reduction in the prevalence of P. gingivalis, P. intermedia, and F. nucleatum in the test group.[26]

    In the current study, the reduction in the bacterial colony counts was similar to the earlier mentioned studies. The limitations of the current study were a short follow-up period of three months and a split-mouth design. This could have resulted in cross-contamination of the microorganisms assessed at the test and control sites. Hence, further longitudinal studies are required to substantiate the results obtained in the current study.


      Conclusion Top


    Within the limitations of this study, improvements in the clinical and microbiological parameters showed that adjunctive use of locally delivered 3% P. guajava gel is effective in the management of chronic periodontitis.

    Clinical significance

    Side effects such as resistance, toxicity, sensitivity, and growth of opportunistic infection have modified the general perception about the use of antibiotics. Herbal products are increasingly used as therapeutic agents in the prevention and management of periodontal diseases to preclude the side effects of antibiotics. A recent addition to the list is the leaves of P. guajava, as they possess antimicrobial, anti-inflammatory, and antioxidant properties. In addition, there are no adverse reactions encountered with the use of 3% P. guajava gel. Hence, it can be regularly employed as an adjunct to SRP in the management of patients with chronic periodontitis.

    Acknowledgment

    Sincere thanks to Mr. Jagannatha P S for his help in conducting the statistical analysis of the data.

    Financial support and sponsorship

    Funded by JSS Academy of Higher Education and Research.

    Conflict of interest

    Nil.

    Authors’ contributions

    Not applicable.

    Ethical policy and institutional review board statement

    The study protocol was approved and cleared by the Institutional Ethical Committee (Letter No. JSS/DCH/IEC/MD-08/2017–18, Date:01/03/2018) and was conducted in accordance with the Helsinki Declaration of 1975, as revised in 2013 (ClinicalTrials.gov identifier: CTRI/2018/11/016377).

    Patient declaration of 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.

    Data availability statement

    The data used in this study are available on request by contacting the corresponding author.

     
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        Figures

      [Figure 1], [Figure 2]
     
     
        Tables

      [Table 1], [Table 2], [Table 3]



     

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