CLINICAL EFFICACY AND SAFETY OF HERBAL MEDICINE THERAPY IN RECURRENT APHTHOUS STOMATITIS: A SYSTEMATIC REVIEW

Recurrent aphthous stomatitis is the most common oral mucous ulcerative lesion with challenging treatment. Herbal medicine therapy can propose clinical efficacy and safety due to its large biological activities. The objective was to review the clinical efficacy and safety of herbal medicine therapy in terms of ulcer size, pain score, healing duration, and adverse effects in recurrent aphthous stomatitis. A systematic was conducted based on the PRISMA statement. The search was performed using four electronic databases, namely PubMed, Cochrane, Science Direct, and Google Scholar for articles published from 2016 until 2021 using specific keywords. The search was limited to randomized controlled trials (RCTs), in English, full text, and study in humans. The main outcome is expected to be ulcer size, pain score, healing duration, and adverse effects. Quality assessment of selected articles was conducted using the Quality Appraisal of Randomized Trials Checklist (Cochrane Risk of Bias tool). The methodology quality of studies was evaluated using the Cochrane Handbook for Systematic Review of Interventions and Rev Man software. Five articles were eligible for analysis. The population of the sample study ranged from 34-70 patients of 15-65 y old. The herbs used were Aloe vera (Aloe barbadensis Miller), curcumin (Curcuma longa), licorice (Glycyrrhiza glabra), tobacco (Nicotiana tabacum L.), and pomegranate (Punica granatum Linn.). This review showed that there is some evidence of the clinical efficacy and safety of herbal medicine therapy in improved outcomes of recurrent aphthous stomatitis treatment with minimum adverse effects.


INTRODUCTION
Recurrent aphthous stomatitis (RAS) is the most common painful oral mucosal disease characterized by recurring ulcers confined to the oral mucosa in patients with no other signs of systemic disease [1,2]. RAS is classified according to clinical characteristics, namely minor ulcers, major ulcers, and herpetiform ulcers [3]. The incidence ranges from 20 % to 60 %, depending on the population studied [4]. Prevalence tends to be higher in professional or white-collar, individuals, upper socioeconomic groups, and nonsmokers [3,4].
There is no specific etiology has been identified, so that the treatment of RAS is non-specific and symptom-based [5]. The goal is to decrease symptoms, reduce ulcer number and size, increase disease-free periods. The best treatment is that which will control ulcers for the longest period with minimal adverse side effects. The treatment approach should be determined by disease severity (pain), the patient's medical history, the frequency of flare-ups, and the patient's ability to tolerate the medication. In all patients with RAS, it is important to rule out predisposing factors and treat any such factors, where possible, before introducing more specific therapy [6,7]. Treatment for RAS mainly includes localized or general medications [7]. Glucocorticoids, which have proven efficacy and safety, are often among the first choices for localized treatment. However, their long-term use may facilitate oral infection with Candida albicans, mucosal atrophy, susceptibility to infection, and gastrointestinal complication [8][9][10].
Herbal medicine can propose efficacy and safety due to its large biological activities with minimum adverse effects [11]. Indonesia has the second biggest biodiversity in the world, exhibited by a high number of indigenous medicinal plants that can be used as medicine [12,13]. The Indonesian people, particularly Javanese people, have been using medicinal plants for a long time by drinking herbs derived from medicinal plants. Traditional Indonesian herbal medicine that has been practiced for centuries in Indonesian society is still very popular for maintaining health and treating diseases [13]. Also, approximately 60 % of the world's population put their faith in medicinal herbs [9]. This is due to more believed to be safe from chemical drugs [13].
Li et al. conducted a systematic review on the efficacy and safety of topical herbal medicine treatment on recurrent aphthous stomatitis based on the studies published from 1996 to 2015 [14]. Meanwhile, Wahyuni et al. conducted a systematic review on the efficacy and safety of plant-based therapy on recurrent aphthous stomatitis and oral mucositis in the past decade [15].
To the best of our knowledge, no systematic review was conducted for randomized controlled trials of intervention regarding clinical efficacy and safety of herbal medicine in RAS for the last five years of studies. Therefore, we conducted this analysis to review the clinical efficacy and safety of herbal medicine therapy in terms of ulcer size, pain score, healing duration, and adverse effects in recurrent aphthous stomatitis.

Protocol and eligibility criteria
This systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement. The PICO items were population or patient or problem (recurrent aphthous stomatitis and human patients), intervention (herbal medicine therapy), comparison (placebo or chemical drugs), and outcome (clinical efficacy and safety).
The inclusion criteria were studies published from 2016 to 2021 with full text available, English-language, a study in humans, randomized controlled trials (RCTs) that evaluated the efficacy and safety of herbal medicine therapy in RAS, and no restrictions on age, sex, or race. We decided to collect studies from 2016 to 2021 because there were no systematic reviews conducted on articles from the past five years. Articles were excluded for duplication and irrelevant with the topic, including non-herbal, animal-based, and non-recurrent aphthous stomatitis.  1).

Study selection, data collection, and data items
The article selection process was conducted by one independent reviewer (N. T. H.). The following data for each study were extracted from full-text articles using a data extraction form and stored in 2020 Microsoft Excel file format: references, study design, setting, scientific name, part or extract, age range, sample size or the number of patients, drug formula, drug preparation and usage, treatment duration, outcome, and biological actions (table 1). The main outcome of the study was ulcer size, pain score, healing duration, and adverse effects.

Risk of bias in the individual studies and across studies
The quality of the enrolled publications was assessed according to the Cochrane Handbook for Systematic Review of Interventions and Rev Man 5.4 software [16]. The assessment of the risk of bias was done by one independent reviewer (N. T. H). The assessment criteria were random sequence generation (selection bias), allocation concealment (selection bias), blinding of participants and personnel (performance bias), blinding of outcome assessment (detection bias), incomplete outcome data (attrition bias), selective reporting (reporting bias), and other bias. There is no scoring system in Cochrane Risk of Bias. Review authors' judgment involves answering a specific question for each entry. In all cases, an answer 'Yes' indicates a low risk of bias, an answer 'No' indicates a high risk of bias, and an answer 'Unclear' indicates insufficient information [16].

RESULTS
The search strategy performed a total of 9786 articles from electronic databases and one article from hand searching. The 17 articles were excluded for duplication. According to the exclusion criteria, screening of the titles and abstracts resulted from the removal of 9750 articles. A full-text review of the remaining 20 articles indicated that 5 studies met the inclusion criteria dan they were enrolled into the final analysis. The flow chart displayed in fig.  1 shows the detailed selection process.
The characteristics of the 5 studies are summarized in table 1. The sample size ranged from 34-70 patients of 15-65 y old. Five different types of herbal medicine, in the form of gel, orabase, solution, and mouthwash, were used. The experimental period of these trials ranged from 5 to 10 d [10,[17][18][19][20].
The risk of bias and quality assessment of each study can be seen below. Table 2 showed risk of bias in each component, fig. 2 showed risk of bias graph across all included studies and fig. 3 showed the risk of bias summary about each risk of bias item for each included study.    [17]. All studies reported no adverse effects [10,[17][18][19][20].

DISCUSSION
A total of 70 subjects in 5 clinical trials were analyzed in the present analysis. Compared with controls, most herbal medicine can greatly improve the patients' symptoms by reducing ulcer size, relieving pain, and shortening healing duration without adverse effects. The homogeneity of the studies was quite poor, with variables such as the various type of intervention, dosage, drug formula, application method, sample size, and experiment duration. Therefore, a metaanalysis could not be conducted based on the current data.
Because of the drug's formula heterogeneity, we could not compare it. However, the gel form of the same medication has better efficacy in comparison with mouthwash because of more time exposure. Nevertheless, it depends strongly on its bioadhesive properties, bioavailability, solubility, and external factors such as mechanical stress and the washing effect of saliva [21].
Nicotiana tabacum, when compared to placebo can significantly reduce ulcer size and pain score without any adverse effects [20]. This is in line with a study conducted by Zakaria et al. They reported the nicotine group showed a lower mean with a significant difference after 6 d in comparison with the placebo group regarding pain and erythema scores. A lower mean of ulcer size was recorded in the nicotine group, with a significant difference after 4 and 6 d [22].
Nicotiana tabacum has anti-inflammatory effects. Combustible products of smoking are known to stimulate increased keratinization of the oral mucosa and therefore resist the formation of aphthous ulcers and reduce trauma or bacterial penetration of the mucous membrane in smokers compared to non-smokers. This composition may be attributed to the presence of the main alkaloid of Nicotiana tabacum, nicotine, which showed a remarkable protective effect in aphthous ulcers and Bechet syndrome by using oral nicotine replacement therapy [20].
Nicotine and its metabolites exhibited immunosuppressive activity and lead to a reduction in inflammatory condition by different known mechanisms, including antibody-forming cell response, suppression of neutrophil-mediated inflammatory action, inhibition of endothelial cell release of IL-8, inhibition IL-1β, IL-2, IL-10, TNF-α, IFN-Ɣ release, decrease the level of immune globulins, impairment of antigen-mediated signaling T-cells, and inducing T-cell allergy as well as attenuation of IFN signaling [20].
Pomegranate peel extract (PPE) 10 % gel was significantly effective in reducing ulcer size and pain score without any adverse effects when compared to placebo. The healing duration in pomegranate was 4.91±1.16, which was faster than placebo [17]. It showed antioxidant, anti-microbial, and anti-inflammatory effects. Pomegranate enhances or maintains the free radical scavenging activity of the hepatic enzymes, namely superoxide dismutase (SOD), catalase, and peroxidase. Antimicrobial activity by controlling adhesion potential of Streptococcus mutans and Streptococcus sanguis to the oral cavity surface. Pomegranate also decreases the level of SOD (a salivary defense system) to promote antiinflammatory effects [17]. Polyphenolic compounds in the pomegranate, including anthocyanins, have an anti-inflammatory effect and prevent the adhesion of leukocytes to endothelial cells, and it decreases the level of IL-2, IFN-Ɣ, and TNF-α. Reducing these inflammatory factors is effective in treating RAS and diabetes. The pomegranate is also against oxidative stress by inhibiting free radicals and having antioxidant properties [23].
Ghalayani et al. studied Punica granatum (PG) in the management of RAS. The findings of this study revealed that PG extract in the form of oral gel (10%) may be beneficial in reducing RAS pain and has a positive effect in reducing the overall period of complete healing. It was concluded that PG is an effective herbal medicine for the management of RAS [24]. In line with Tavangar et al. showed that Punica granatum gel has a successful effect in controlling and treating RAS [23].
Licorice in DSG group promotes 1.5 d faster-wound healing compared to DS group. Licorice also statistically significantly reduces pain score compared to DS group [10]. In line with the study conducted by Martin et al. examining the effect of a patch containing glycyrrhiza root extract on aphthous ulcers. They reported that the patch was more effective than the placebo in reducing the healing duration of RAS [25].
Licorice (Glycyrrhiza glabra) is one of the oldest medicinal plants. The main ingredient of licorice is glycyrrhizic acid and glabridin. It has antiinflammatory and antimicrobial effects. These effects might be due to the glycyrrhizic acid, glabridin, and flavonoid in licorice, which inhibit the enzymatic pathways of cyclooxygenase (COX) and lipoxygenase. Licorice and its active components inhibit the production of free oxygen radicals as well as cell migration, and thus control arachidonic acid metabolism, vascular permeability, and allergy, thereby reducing inflammatory reactions and the severity of pain [10].
Giroh et al. reported the size of ulcers in the treatment of aloe vera and kenacort were not statistically significant [18]. Bhalang et al. who evaluate the effectiveness of acemannan (extract of aloe vera) in RAS, also found that 0.1% of triamcinolone acetonide was inferior to aloe vera in reducing ulcer size [26]. While Giroh et al. reported that aloe vera was statistically significant in reducing pain and burning sensation without adverse effects [18].
Aloe vera has anti-inflammatory and wound-healing effects. The mechanism of the wound healing effect was to increase epithelial cell migration and the synthesis of hyaluronic acid and dermatan sulfate in the granulation tissue of the wound. In addition, aloe vera increases the wound closure rate and tensile strength of the wound. Aloe vera gel forms a protective coating on the affected areas and helps in healing wounds, fastens the healing rate, and relieves pain. Aloe vera and its components inhibit the COX pathway and prostaglandin E2, leukocyte adhesion, and pro-inflammatory cytokines, which reduce inflammation [18]. Kia et al. showed that there were no statistical differences in pain severity and lesion size between 5% of curcumin and 0.1% of triamcinolone acetonide [19]. In contrast to a study conducted by Manifar et al., showed 2% of curcumin gel significantly reduces pain severity and size of ulcer compared to placebo [27]. Curcumin has an anti-inflammatory and antioxidant effect. The mechanism of the anti-inflammatory effect was to inhibit the phospholipase, lipids, COX-2, and reduction of IL-1, IL-6, and TNF-α. Moreover, curcumin stimulates cortisol secretion which generates a reduction of IL-1, IL-6, and TNF-α. One of the probable mechanisms in the description of aphthous healing by curcumin may be related to its antioxidant effects [19]. All of these herbs in included studies had no adverse effects [10,[17][18][19][20]. The mechanism of included herbal medicine can be seen on fig. 4.
Most of these included studies showed a low risk of bias. This means that plausible bias is unlikely to seriously alter the results [16]. The limitation of this review was the heterogenicity of included studies that were assessed subjectively. In summary, the current data show favorable benefits of herbal medicine in treating RAS. So, there is some evidence to suggest that herbal medicine therapy is an effective and safe alternative option for RAS treatment. Moreover, this review can be implemented as a reference for the development of further research for herbal medicine therapy in recurrent aphthous stomatitis treatment.

CONCLUSION
This review showed that there is some evidence of the clinical efficacy and safety of herbal medicine therapy in improved outcomes of recurrent aphthous stomatitis treatment with minimum adverse effects.