Gingivitis and periodontitis are the most common oral diseases; the first stage of the disease is inflammation of the soft tissue besides the teeth which is called gingivitis. ¹ However, periodontitis is characterized by inflammation of supporting tissues of the teeth, progressive destruction of periodontal ligament, and alveolar bone along with formation of periodontal pocket and gingival recession. ² , ³

The immune response from interaction between microorganisms and inflammatory cells in the surrounding tissue of the teeth leads to this disease. Released enzymes and toxins from these microorganisms make periodontium destructed. ³ Seven important signs of periodontal disease are red and swelling gums, gingival bleeding, unpleasant breath and bad taste, gaps which gradually form between teeth, loose teeth that eventually falling out if not treated, gingival receding or recession, and itchy or painful gums. ⁴ The main microorganisms involved in disease progression are Porphyromonas gingivalis, Bacteroides forsythus, and Actinobacillus actinomycetemcomitans which are very important in etiology of chronic periodontal disease. More than 500 different bacterial species are found in dental plaque. Many of periodontal pathogens are obligatory anaerobic and Gram-negative Bacteria. ⁵

Bacteria in periodontal pockets live in biofilms (sticking to the surface) that help them resist against antibiotics. Highly resistant Bacteria with pumps that help remove antibacterial drugs from cell have recently been identified. Bacteria can connect to each other in biofilm; this action helps transmission of genes that play the role of resistance against antibiotics and growth of the resistant microorganisms species. ⁶

Timely treatment of the disease and necessary care reduce the disease progression and prevent progressive bone erosion. Without essential care and interventions, bone erosion could cause teeth loss. ⁷ The most important treatment modality is controlling the inflammation by removing dental plaque and calculus. The first stage of treatment is a nonsurgical procedure which includes health education, dental root planing and scaling to reduce pathogen microorganisms. ⁸

Nonsteroidal anti-inflammatory drugs (NSAIDs) such as flurbiprofen, naproxen, and mefenamic acid have been proven to be effective in the treatment of periodontal disease. ⁹ Furthermore, due to special role of some microorganisms in disease progression, antibiotics and antiseptics are used too. These antibiotics include tetracyclines, clindamycin, metronidazole, penicillins, and antiseptics such as chlorhexidine. ¹⁰

Misuse of systemic antibiotics can lead to Bacterial resistance. Since the causes of disease are a wide range of Bacteria, there are no ideal and effective antibiotics for all the pathogens and concomitant use of different antibiotics may intensify medical side effects. ¹¹ Short-term efficacy and inconstancy of antiseptic mouthwashes and side effects of systemic antibiotics led to use of local therapy systems. These new systems have advantages of reduction in prescribed drugs, increasing drug concentration in target tissue, reduction in drug side effects and reduce drug use frequency. ¹² Local tetracycline fibers at subgingival area, minocycline gel at subgingival area, and chlorhexidine and metronidazole gels at subgingival area are the examples of these novel drug delivery systems. ¹³ , ¹⁴

Natural products such as propolis can also prevent infection development. They are very safe and cost-effective and can reduce side effects. Local drug delivery of natural products to periodontal pocket is an adjunctive therapy in addition to dental root planing and scaling for the treatment of periodontal disease. ⁷

Propolis is a brown, viscous, and wax-like resin which is very sticky (bee's glue) and is collected by bees and extracted from hives. Bees use propolis to fill the gaps, smooth inner surfaces of the hive and prevent entering other insects to the hive. ¹⁵ Extracted propolis is mixed with hypopharyngeal gland secretions, beeswax, and pollen. It is a very complex mixture which its composition differs from one region to another. ¹⁶ Propolis composition contains 55% resinous compounds, 30% wax, 10% aromatic oils, and 5% honeybee pollen. Propolis is full of Vitamins A, B ₁, B ₂, B ₃, biotin, bioflavonoids (very important content of propolis and the reason for its properties), and polyphenols. ¹⁷

Polyphenol and flavonoid compounds have antimicrobial activity and are indirectly involved in response to microorganisms. The mechanisms of producing hydrogen peroxide, protein, and bacterial enzymes inhibition and antiseptic activity are well identified for these compounds. ¹⁶ Due to reduction in antioxidant activity of saliva and salivation in these patients, using an antioxidant can be beneficial to prevent disease progression. Polyphenols and flavonoids increase antioxidant activity of saliva and inhibit periodontal diseases. They inhibit prostaglandins production too. ¹⁸

Propolis has antibacterial, anti-inflammatory, antioxidant (because of artepillin C and caffeic acid) activity. Nowadays, propolis is very applicable in new formulations of dentistry, dermatology, gastroenterology, and veterinary. ⁷ , ¹⁷ , ¹⁹ , ²⁰

Mucoadhesive drug delivery systems have been introduced as new dosage forms because of their ability to remain on the mucous membranes and a slow release of their drug content. One of the new dosage forms is mucoadhesive gel. Some properties of ideal formulation are ease of entrance to periodontal pocket by syringe, controlled release of drug to oral mucosa, the ability to remain in the pocket without mechanical connectivity to teeth surfaces, and being nonallergic and nontoxic. ⁷ It seems that this drug delivery system can let us save more time and money because the patient needs to visit the physician just once for applying drug to the periodontal pocket.

The aim of this study is designing and formulation of propolis mucoadhesive gel from carbomer 940, hydroxypropyl methylcellulose (HPMC), and NaCMC as adjuvant therapy for periodontal disease alongside dental root planing and scaling with antibacterial, anti-inflammatory, and antioxidant effects.

Materials

Propolis was provided and authenticated by the Agricultural Research Center (Isfahan, Iran) on April 2014.

Folin–Ciocalteu reagent, quercetin, gallic acid, sodium carbonate anhydrous, carbopol 940, HPMC K4M, NaCMC, polyethylene glycol 400 (PEG 400), triethanolamine, methanol, and 96% ethanol were purchased from Merck Company (Germany).

Potassium sorbate, aluminum chloride 20%, and glacial acetic acid were obtained from Sigma-Aldrich Chemie GmbH, (Steinheim, Germany).

Methods

Propolis extraction

Propolis particles were mechanically cleaned and placed in freezer for fragmentation into fine particles. To produce propolis extract, 3000 ml of 96% ethanol was mixed well with 1 kg propolis by maceration method of extraction. This mixture was retained at room temperature and dark place. It was mixed 3 times a day. After 10 days, it was filtered through filter paper, grade 589 (Whatman GmbH, Dassel, Germany). For extract concentration, a rotary evaporator (Heidolph VV 2000) was used at 40°C to remove extra solvent.

For further concentration, water bath was used. ⁷ , ²¹

Determination of pH extract

pH of the extract was measured by pH meter (Metrohm 632 Swiss). First measurements were done 24 h after extraction, next pH measurements were prepared 1 week, 2 weeks, 1 month, 3 months, and 6 months after extraction. ²² The pH was measured 3 times and the average value was reported.

Determination of polyphenols in ethanolic extract of propolis

In this study, the Folin–Ciocalteu method for determination of polyphenol contents was used. Folin–Ciocalteu colorimetry method is based on a chemical reduction of the reagent (a mixture of tungsten and molybdenum oxides). Gallic acid solution is used as the standard solution. To prepare stoke solution, 500 mg of dry gallic acid was dissolved in 10 ml of 96% ethanol; then, it was diluted to 100 ml by purified water. To draw gallic acid calibration curve, 1, 2, 3, 5 and 10 ml of stoke solution were added to distinct 100 ml volumetric flask and were diluted with purified water. Reference and blank solutions were also prepared using Folin–Ciocalteu's reagent and sodium carbonate. After shaking and incubation at 20°C–25°C for 2 h, the absorbance of each sample was measured at λ _max765 nm by the ultraviolet-visible (UV-VIS) spectrophotometer (UV mini-1240, Shimadzu). Then, by means of calibration curve, we measured polyphenol concentrations and their percentages. The polyphenol content was reported as mg gallic acid equivalent (GAE) per 1 g of the extract. ²³

To prepare the sample, 500 mg of propolis was dissolved in 10 ml of 96% ethanol and then was diluted to 100 ml with purified water in a volumetric flask. Other steps were repeated as for standard sample. Finally, the absorbance of our sample was measured at 765 nm. ²⁴ The test was repeated 3 times and the average of absorbances was applied.

Determination of flavonoids in ethanolic extract of propolis

Aluminum chloride colorimetric method was used to determine flavonoid content in propolis extract. Quercetin is considered as the standard solution. Standard curve of quercetin solution was drawn using 12.5, 25, 50, 75, and 100 concentrations as µg/ml in methanol. ²⁴

Preparation of propolis standard solution was as follows: 50 mg of ethanolic extract of propolis was dissolved in 25 ml methanol, 100 µl of this solution was mixed with 100 µl of 20% aluminum chloride, and 2 drops of glacial acetic acid was added and diluted with methanol to 3 ml. It was mixed well and kept at room temperature for 40 min. Blank solution with only propolis extract and glacial acetic acid was also prepared. Then, absorbances of samples were measured at λ _max415 nm, and flavonoid concentration in extract was determined by the standard curve. The flavonoid content was reported as mg of quercetin (QE) equivalent per 1 g of extract. ²³ The results were reported in average after triplicate experiments.

Formulation preparation of propolis mucoadhesive gel

Some polymers such as NaCMC, carbopol 940, and HPMC were used to obtain propolis mucoadhesive gel. The amount of concentrated extract in gel base is 10% w/w.

Carbopol 940 gel

For preparation of carbopol gels, we applied three different amounts of carbopol. Potassium sorbate was dissolved in purified water at 50°C. ²⁵ Then, specified amounts of carbopol 940 were dispersed in purified water at 40°C and were mixed by mixer at 1200 rpm for 30 min Table 1. Propolis extract was dispersed in PEG 400, and then, it was gradually added to the base. The neutral pH was obtained by required amount of triethanolamine to gain a transparent gel. ²⁶ , ²⁷ {Table 1}

HPMC gel

For preparation of this gel, three different amounts of HPMC were used. Potassium sorbate was dissolved in purified water at 50°C. ²⁵ Specified amounts of HPMC were dispersed in purified water at 50°C and were mixed by mixer at 1200 rpm for 30 min Table 1. Then, the remaining amount of purified water was added coldly and mixed to gain a homogenous gel. The prepared gel is kept in refrigerator for 1 night (hot/cold technique). Propolis extract which was dispersed in PEG 400 was then added to the base and mixed well to obtain a homogenous gel. ²⁸

NaCMC gel

For preparation of this gel, two distinct amounts of NaCMC were used. Potassium sorbate was dissolved in purified water at 50°C. ²⁵ Then, specified amounts of NaCMC were dispersed in purified water at 50°C and were mixed by mixer at 1200 rpm for 30 min Table 1. Propolis extract was dispersed in PEG 400, and then, it was gradually added to the base. ²⁷

Carbopol 940 and NaCMC gel

Two different amounts of carbopol and NaCMC were used. Potassium sorbate was dissolved in purified water at 50°C. ²⁵ The weighed amounts of carbopol and NaCMC were dispersed in purified water at 50°C and mixed well Table 1. Propolis extract which was dispersed in PEG 400 was then added to the gel bases.

Evaluation tests for formulations

Macroscopic and microscopic study

This test is carried out 48 h after gels preparation. Macroscopic balance is checked in this evaluation test (color, homogeneity, transparency, lack of particles presence, and consolidation). ²⁷ Microscopic reviews are done by 10 X and 40 X microscopes to check the gel texture, uniformity, and presence of bubbles. Evaluation of consolidation is by pressing a small amount of gel between fingers to consider its consolidation. ²²

pH evaluation of gel formulations

pH meter, which was recently calibrated with standard buffer solution at pH 4 and 7, was used to measure the pH. One gram of each formulation was dissolved in 10 ml purified water. ²⁹ The test was repeated 3 times and the average was reported. pH of samples was measured 48 h, 1 week, 2 weeks, 1 month, 3 months, and 6 months after preparation.

Centrifugal test

This test was carried out to evaluate the resistance of gel against gravity 48 h after preparation. Five grams of each sample was added to test tubes, and then, they were centrifuged at 1200 rpm for 5, 15, 30, and 60 min by centrifugal device (centrifuge 5430). Each formulation was evaluated in terms of precipitation. ²² For each formulation, the test was repeated 3 times.

Thermal changes test

This test is to evaluate gel stability against climatic conditions. Forty-eight hours after gel preparation, three samples from each formulation were prepared and placed at 4°C, 25°C, and 45°C. The samples were evaluated considering appearance, homogeneity, and quality of gel after 24 h, 1 week, 2 weeks, 1 month, 3 months, and 6 months. ²²

Cooling and heating test

Extreme thermal changes lead to investigate formulations stability. Fifteen grams of each formulation was placed at 45°C for 48 h and at 4°C for 48 h for 6 consecutive periods. The apparent changes were then reported. ²²

Freeze and thaw test

This test is to investigate the physical stability of semisolid products in extreme thermal changes. Forty-eight hours after gel preparation, 15 g of each sample was placed in 6 consecutive periods which include 48 h at −8°C and 48 h at 25°C. ²²

Thermal stress test

Stability studies are done at accelerated conditions and also at 30°C ± 2°C and relative humidity of 60% ±5% for 6 months. ²²

Study of ex vivo adhesion strength of the formulations

Mucoadhesion is a principal property to treat periodontal disease. The SANTAM instrument (STM-1, Iran) is used to measure gel mucoadhesion. In this study, mucosal lining of the cow cheek was employed as a model to determine the adhesion strength of the gel. Certain weight of gels (200 mg) was spread over a piece of mucosal lining of the cow (20 mm × 20 mm) after wetting it by some drops of purified water. Then, it attached to constant surface of instrument while another piece of mucosal lining of the cow was attached to the upper surface of the SANTAM instrument. Then, the gel was kept in full contact with the mucosa for 2 min. The force required for detachment of the gel from the mucosal surface with the rate of 10 mm/min was calculated and reported as the adhesion force of the gel. The detachment force was measured in terms of MPa. ³⁰ The test was repeated 3 times for each formulation.

Determination of flavonoids in gel formulations

Forty-eight hours after preparation of formulations, 1 g of each formulation was dispersed in purified water and diluted to 10 ml in volumetric flask. ³¹ Flavonoid content was quantified as QE as defined before. ²³

In vitro drug release

Drug release test of gels is done by Franz diffusion cell using a cellulose acetate membrane. 500 mg of each formulation was placed on cellulose acetate membrane. The device was filled by phosphate buffer, pH 6.2 as dissolution medium at 37°C ± 0.5°C. One milliliter aliquots of medium were removed at times of 6, 24, 48, 72, 96, 120, 144, and 168 h since the beginning of test. The aliquots were replaced by fresh phosphate buffer subsequently. The absorbances of samples were measured at 415 nm by UV-VIS spectrophotometer. The released drug was calculated by the quercetin standard curve. Concentrations obtained are apparent concentrations. To determine the actual amount of released drug, following equation is used: ²²

C _n= C + (C _n− ₁) V/V _t

C _n: Actual concentration in sample n

C: Apparent concentration in sample n

C _{n − 1}: Actual concentration in sample n − 1

V _t: Volume of receive phase

V: Sample volume.

Drug release kinetic studies of gel formulations

To investigate the mechanism of flavonoids release from gel formulations, the release data were analyzed with the following mathematical models: zero-order kinetic (Equation 1), first-order kinetic (Equation 2), and Higuchi kinetic (Equation 3).

Q _t= K ₀t(1)

In Q _t= In Q ₀− K ₁t(2)

Q _t= K _ht ^1/2(3)

In these equations, Q _tis the percent of drug released at time t, Q ₀is the percent of drug present in the gel. K ₀, K ₁, and K _hare the constants of the equations. In the zero-order kinetic model, diagram of cumulative percentage of drug release was plotted against time and a linear plot was obtained. In the first-order kinetic model, log of cumulative percentage of drug remaining was plotted against time and a linear plot was obtained. In Higuchi kinetic model, cumulative percentage of drug release was plotted against square root of time and a linear plot was obtained. ²²

Furthermore, drug release mechanism was determined according to the Korsmeyer–Peppas equation.

Log (M _t/M _∞) = logk + n logt

Where M _∞is the amount of drug released after infinite time, M _tis the cumulative amount of drug released at any specified time (t), k is the release rate constant, and n is the release exponent.

When n value is 0.5 or less, the Fickian diffusion phenomenon dominates, and n value between 0.5 and 1 is non-Fickian diffusion (anomalous transport). The mechanism of drug release follows Case-II transport when the n value is 1, and for the values of n higher than 1, the release is characterized by super-Case-II transport. ³⁰

Determination of viscosity

Viscosity of samples was measured by Brookfield DV-3 viscometer at 100 rpm, using spindle number 74 at 25°C. The spindles of the device rotate in the sample which is placed in the container. Before determination of viscosity and start of the device, the samples were put at room temperature for 30 min. ²² A triplicate experiment was done for each formulation.

Syringeability

Injectable systems are preferred for drug delivery to periodontal pocket. Injection of gels is evaluated by 21-gauge needle. Ease of syringeability is the scale of measure. ⁷ , ²²

In vitro evaluation of antibacterial activity of gel formulation (F ₁₀) against Porphyromonas gingivalis

The most common etiological agent of chronic periodontitis is P. gingivalis. ⁵ In this study, antibacterial effect of gel formulation (F ₁₀) against P. gingivalis using the disk diffusion method was studied. P. gingivalis was prepared by sampling from periodontal pocket of a patient (a 46-year-old man from dental clinic of Islamic Azad University of Isfahan, Isfahan, Iran). Then, they were cultured on supplemented Brucella agar plates (prepared from the Iranian research organization for science and technology) under sterile conditions. Culture media were incubated for 72 h in anaerobic condition at 37°C ± 2°C to allow bacterial growth. ³² After bacterial growth, a microbial suspension was prepared and transferred to the plates containing culture medium by a sterile swap. Then, sterile paper discs with a diameter of 5 mm (Blank disc, Patan Teb, Iran) were smeared with gel formulation and were placed on the surface of the plates containing culture medium in equal distances. Then, the plates were incubated at 37°C ± 2°C for 72 h. After this period, the diameter of the zone of inhibition around the disc was measured in millimeters and the average of diameters noted. The test was conducted in triplicate. Sterile discs free of any material were used as negative control in culture medium. For positive control, tetracycline disc (30 µg/ml) was used. ²²

Analyzing the propolis extract

About 500 ml of concentrated extract of propolis was obtained from maceration and removing of alcoholic solvent by rotary evaporator. Its pH was in the range of 6–7. The content of phenolics as GAE was expressed as 39.02 ± 3.24 mg/g of concentrated extract. The content of flavonoids as QE was expressed as 743.28 ± 12.1 mg/g of concentrated extract. The curve linear equation for gallic acid was y = 0.122x + 0.002 (R ²= 0.979) and for quercetin was y = 0.0067x + 0.008 (R ²= 0.997).

Propolis gel analysis

10% propolis extract was added to gel base of carbopol 940, HPMC, and NaCMC Table 1. After adding propolis extract to formulations containing HPMC (F ₆–F ₈), they developed phase separation and precipitation and propolis ethanolic extract was appeared as some drops on the surface of gel. The reason of phase separation was incompatibility between extract and HPMC so that theses formulations were not tested for further experiments. Viscosity of F ₃and F ₅was high, so ethanolic extract was not dispersed uniformly. Their physical appearance and uniformity were also inappropriate, so no more tests were carried out.

F ₁, F ₂, F ₄, F ₉, and F ₁₀had good physical appearance and uniformity; thus, they were selected for further tests. In centrifugal tests, thermal changes test, cooling and heating test, freeze and thaw test, and thermal stress test, all of the five selected formulations passed all of the tests. pH of selected formulations is mentioned in Table 2. They were in the range of 5–6.{Table 2}

Results of drug content of the formulations, mucoadhesion, and viscosity measurements are shown in Table 2. As considered, F ₁₀has the highest viscosity and mucoadhesion. Results of drug release are shown as diagrams of cumulative percentage of drug release against time Figure 1. Drug release from F ₁₀was slower than other formulations. Time needed for 50% release of whole drug content from F ₁₀is approximately 75 h as displayed in Figure 1. This time is 65, 60, 50 and 46 h for F ₂, F ₉, F ₁, and F ₄, respectively. Figure 1

Percentage of cumulative drug release of formulations F ₁, F ₂, F ₄, F ₉, and F ₁₀in Franz diffusion cell through a cellulose acetate membrane, in pH 6.2 phosphate buffer at 37°C, during 7 days.

Periodontitis is the inflammation of gums and supporting tissues of the teeth. It is one of the most common human diseases. Periodontitis is caused by certain Bacteria and local inflammation is triggered by those Bacteria. ³⁰ As described before, treatment methods are dental root planing and scaling, NSAIDs, antibiotics, and antiseptics. ⁸ , ⁹ Propolis is a natural and nontoxic beehive product that prevents disease progression. Propolis contains flavonoids and polyphenols that have antimicrobial, anti-inflammatory, and antioxidant activity. It can help with dental root and scaling which is very effective in treatment of periodontal disease. ¹⁶ , ¹⁷

In the present study, the content of phenolics and flavonoids was evaluated 39.02 ± 3.24 mg GAE/g of concentrated extract and 743.28 ± 12.1 mg QE/g of concentrated extract, respectively. ²³ According to Yaghoubi et al., thepercentage of phenolic and flavonoid contents of Iranian propolis was calculated 36% and 7.3%. ³³ In addition, the concentration to phenolic and flavonoid compounds in Brazilian propolis was 100.59 ± 2.27 mg GAE/g of concentrated extract and 69.35 ± 0.38 mg QE/g of concentrated extract, respectively. ²⁰ This difference in their chemical compounds could be due to different geographic origins with different climates, vegetation, and the time that the samples collected. ¹⁶

Periodontal pocket has the pH of 6.2, so the pH of product should approach to local pH to decrease local irritations. ¹² The pH of selected formulations was all in the range of 5–6 which is nearly ideal.

Comparison of mucoadhesion results revealed that the highest mucoadhesion first belongs to F ₁₀. Mucoadhesion strength represents a power which makes mucoadhesive membranes stick to mucous membrane. Proper polymers should have groups forming hydrogen bonds, wetting characteristics, and swelling ability to create mucoadhesion power. ²⁷ Mucous surface is coated by mucin. Mucin has negative charge at physiological pH. A prerequisite for mucoadhesion is forming hydrogen bond between hydrophilic functional groups of mucoadhesive polymer and mucosal layer. More link points increase mucoadhesion strength. Higher molecular weights and viscosity elevate mucoadhesion strength. ³⁴

Carbopol is carboxy vinylic derivatives that are widely used in the manufacture of hydrogel dosage forms. It has high molecular weight and swells up to 1000 times of its original volume while neutralizing the system. It permits ionization of carboxyl groups, and as a consequence, a strong gel is formed. ²⁷ Carbopol connection to mucin is secondary type while other polymers establish primary connection to mucin. ²² Carbopol as a mucoadhesive polymer increases gel's durability which is the most important matter about periodontitis.

As shown in Table 2, F ₁₀has the highest viscosity. It reveals that carbopol 940 increases gel's viscosity and consolidation. Release rate in F ₁₀is slower than F ₁, F ₂, F ₄, and F ₉. When the amount of polymers increases, gel gets more solid and water penetration gets harder and thus drug release rate decreases and gel's durability in periodontal pocket rises, that is, the most significant thing in treatment of periodontitis. ⁹ The most important role in drug release rate belongs to carbopol due to its high molecular weight and high cross-linking, so increase of carbopol leads to increase of viscosity and mucoadhesion and decrease of release rate. ²⁷ , ³⁴

It is important that the product could be delivered from a syringe to fulfill the requirement of ease of application. All of the formulations (F ₁, F ₂, F ₄, F ₉, and F ₁₀) had ease of delivery from syringe in 21-gauge needle.

Propolis was reported to have antibacterial activity on some Gram-negative Bacteria. Antibacterial activity against periodontal pathogens was exhibited by flavonoid compounds such as quercetin, kaempferol, pinocembrin, and galangin. ⁵ , ³⁵ The data obtained in the present study revealed antibacterial activity of formulation F ₁₀against P. gingivalis.

Ideal formulation should easily get into grooves of tooth and should have proper mucoadhesion and persistence in periodontal pocket. Controlled release of drug into periodontal pocket helps effective treatment and recovery and higher persistence and reduces drug use frequency. ⁷ According to these results, F ₁₀was the best formulation in the treatment of periodontitis.

Propolis extract contains polyphenols and flavonoid considerably that can be used in periodontal pocket therapy due to antioxidant, antibacterial, and anti-inflammatory activity. Based on the in vitro drug release, viscosity, and mucoadhesion studies, F ₁₀containing 1% carbopol 940 and 3% NaCMC was selected as the best formulation. F ₁₀showed satisfying mucoadhesion and viscosity and optimum release profile.

Increase of polymers leads to increase of viscosity and mucoadhesion and as a result, decrease of release rate. Study of release profile in F ₁₀also revealed that propolis could release from this system in longer time (more than 7 days). F ₁₀produced significant growth inhibition zone against P. gingivalis. This formulation has efficient function in the treatment of periodontitis besides dental root planing and scaling, so we recommend it for clinical evaluation.

This study was supported by the Isfahan University of Medical Sciences as a thesis research project numbered 393141.

The authors of this manuscript declare that they have no conflicts of interest, real or perceived, financial or non-financial in this article.

Financial support and sponsorship

This study was supported by the Isfahan University of Medical Sciences as a thesis research project numbered 393141.

Conflicts of interest

The authors of this manuscript declare that they have no conflicts of interest, real or perceived, financial or non.financial in this article.