INTRODUCTION:

Nature provides abundant plants that can be used as medicine for all human ailments. Natural products play a major role in the drug development system, although synthetic drugs usage is increasing due to its low cost, fast-acting, and ease of evaluation. However, there are concerns about the safety of using a synthetic drug. Currently, 80% of the population of developing countries rely on natural products as the most reliable source of medicine^1,2. Medicinal plants contain various active substances that have not been utilized for drug discovery^{3, 4}. The extraction and purification of bioactive compounds are essential to identify the active phytoconstituents responsible for the pharmacological activity⁵_.

Many medicinal plants contain active compounds with specific therapeutic effects on the human body and have been used as traditional medicine. Phytochemicals are natural bioactive compounds derived from certain parts of the plant where the content differs from one part of the plant to another^6-8. Medicines with natural ingredients are more acceptable to the public and are increasingly in demand in the world market because they are believed to be safer with fewer side effects than drugs with synthetic formulations^9,10.

Barringtonia racemosa (L). Spreng is widely known as a medicinal plant. This herb has a wide variety of therapeutic applications. The fruit is effective in treating cough, asthma, and diarrhea. The seeds are used to treat colic and ophthalmia. The bark and leaves are used to treat rat and snake bites and heal stomach ulcers¹¹. The seeds of this plant are used to treat cancer in remote areas of Kerala, India¹². This plant contains diterpenes, triterpenoids, flavonoids (including polyphenols), steroids, and saponins^13,14. B. racemosa seeds contain saponins and flavonoids¹⁵. The shoots of this species were found to have three flavonoid compounds, namely quercetin, routine, and kaempferol¹⁶. The tree, bark, and fruit contain diterpenes, triterpenoids, flavonoids, and steroids¹⁷.

The pharmacological activity of B. racemosa is antibacterial against gram-positive and gram-negative bacteria strains, namely Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Shigella dysentriae, Vibrio cholera, and Proteus sp. This plant also acts as antinociceptive (analgesic), antioxidant, anti-inflammatory, and anti-fungal¹³. The effectiveness and the ease of using B. racemosa extract on the wound can be increased by formulating it into a gel form. Topical gel preparations can deliver therapeutic ingredients well, easily and evenly distributed when applied to the skin and provide a cool sensation on the wound¹⁸.

A wound is a state of loss or damaged tissue caused by trauma to sharp or blunt objects, changes in temperature, chemicals, explosions, electric shocks, or insect bites. A healthy body has the natural ability to protect and heal itself. The wound healing process occurs when the tissues carry out the repair and regeneration process^19,20. The wound healing process is divided into²¹ five stages: coagulation and homeostasis, inflammation, proliferation and repair, maturation, and remodeling²². Many studies have shown that using traditional medicinal plants can accelerate wound healing process, some even more effectively than synthetic drugs^23-27.

The skin is the largest organ of the body and consists of connective tissue, nerves, muscles, and epidermis. The skin is responsible for providing sensation, thermoregulation, biochemistry, immunity, physical protection²⁸, and maintaining fluid balance²⁹. The skin is divided into three main layers: epidermis, dermis, and subcutaneous. The skin mirrors a healthy life that needs protection from pathogen^30,31.

Topical drug administration is the application of drugs containing formulations with specific pharmacological effects on the skin surface. Drug forms such as creams and lotions often provide poor drug bioavailability because they are quickly removed from the skin surface, whereas powders can only briefly stick to the skin. Gels are semisolid systems in which the movement of the dispersing medium is limited by the interaction of the three-dimensional network of partial or dissolved macromolecules of the dispersed phase³². Gel is transparent to semi-transparent in colour and imparts a moist feel. Gel has been widely used in cosmetic products because it provides a moist and light feeling^{33, 34}.

Carbopol is a hydrophilic gelling agent easily dispersed in water. Carbopol can be used as a gel base in small quantity (about 0.5-2.0%) with an ideal thickness at pH 6-11³⁵. Carbopol is a white hygroscopic powder with feathery texture and a spesific odour³⁶. Ideally, gelling agents should be inert, safe, nonreactive to other components of the formula, and easily adjusted to the dosage form^37,38. This study aimed to determine the effect of various concentrations of the B. racemosa kernel extract on the physical stability of topical gel.

MATERIALS AND METHODS:

Preparation of carbopol 940 gel base:

The gel base consisting of carbopol, Trietanolamin (TEA), propylene glycol, methylparaben, and distilled water was prepared into four different formulas with various concentrations of B. racemosa kernel extract (1gr, 3gr, 5 gr and 7gr. Methylparaben (0.2%) was dissolved in distilled water by heating to a temperature of 70°C, then carbopol was added and continuously stirred until the gel was formed. Then, propylene glycol (15%) and TEA were added (Table 1).

Table 1: Gel base formula of B. racemosa seeds extract

Ingredients	Formula base (% b/b)
Ingredients	F1	F2 (1%)	F3 (3%)	F4 (5%)	F5 (7%)
Carbopol	2	2	2	2	2
TEA	2	2	2	2	2
Glycerin	1	1	1	1	1
Propylene glycol	6	6	6	6	6
Methylparaben (Nipagin)	0.2	0.2	0.2	0.2	0.2
Aquadest	Ad 100	Ad 100	Ad 100	Ad 100	Ad 100
B. racemosa kernel extract (gr)	0	1	3	5	7

The organoleptic tests:

The organoleptic test was carried out by observing the extract's shape, color, and odour at room temperature. The form was examined from the gel that can flow in the container. The colour was inspected against a white paper background with lighting. The odour was smelled by wiping the finished preparation over.

The homogeneity test:

The homogeneous test was carried out by applying 0.1g of the gel preparation to a transparent glass plate and observing its homogeneously. The preparation must show a homogeneous arrangement, indicated by the absence of coarse grains on the slide. The test was carried out once a week for 4 weeks.

The viscosity test:

The viscosity test was carried out using a Brookfield type LVDV-E viscometer with the appropriate spindle and speed. The gel was put in the beaker glass until it reached the volume of 500mL, and then, the spindle was attached to the specified limit. Measurement was taken once a week for 4 weeks. The ideal viscosity is 6,000 – 50,000 cP or 6 – 50 Pa.S (SNI 16-4399-1996).

Statistical analysis:

All data were analyzed for the normality and homogeneity using the Shapiro-Wilk test and Levene test, respectively. Multivariate analysis was carried out to see the effect of various concentrations of B. racemosa kernel extract on the physical stability of the gel using Kruskal Wallis (as the data were not normally distributed and not homogeneous).

RESULT:

The organoleptic tests:

Based on the Shapiro-Wilk test, the p-values for all data that underwent the organoleptic test for colour change was 0.006 (p<0.05), which means the data is not normally distributed. Levene test also revealed that all data was not homogeneous as the p<0.001.

Our data found that the colour of the gel was changed on the organoleptic test (Figure 1). Gel color organoleptic test on blanks with different test formulas, where in blanks the color remained from week 0 to week 4 which was clear, but the test formulas of 1%, 3%,5%, 7% have different colors from blank starting from week 0 to week 4. The 4th week the 5% and 7% formulas were brownish yellow.The Kruskal Wallis test showed a significant effect of different concentrations of B. racemosa extract on the gel’s colour change (p=0.009). There was no significant difference in the odour and consistency of the gel during 4 weeks of observation after adding different concentrations of B. racemosa kernel extract.

Figure 1. The colour change of the topical gel after the addition of B. racemosa kernel extract with various concentrations. Description: 1 = clear; 2 = yellowish white; 3 = brownish yellow.

The homogeneity tests:

Each gel formula is homogeneous with no significant difference from Week 0 to Week 4.

The pH test:

Data on pH test of all gel formula is not normally distributed and not homogeneous (p=0.006 and p<0.001, respectively).

Figure 2: The pH of each gel formula

Our data suggested that the pH of each formula is different each week (Figure 2). From Week 0 to Week 2, the 3%, 5%, and 7% gel formulas had the same pH value of 6. However, the pH start to change from Week 3 to Week 4 with 3% gel having pH of 5.7, while 5% and 7% gel formulas have the same pH of 5.5. Based on Kruskal Wallis test, there is a significant effect of variation in concentration of the B. racemosa kernel extract on the pH of topical gel.

The viscosity test:

Data represented the viscosity of the gel was not normally distributed with p=0.006 and not homogeneous p=0.001(<0.05). The viscosity of the gel was significantly changed each week and increased with the increasing dose of B.racemosa kernel extract in the gel p=0.001(<0.05). The 7% formula is constantly thicker than the other formula, while the blank gel is the most watery of all gel formula (Figure 3).

Figure 3. The viscosity result of each gel formula

DISCUSSION:

The organoleptic test for four weeks showed that all gels had no change in smell and consistency. The gel is stored in a tightly closed plastic container at a constant room temperature so that the odour and consistency are not affected by environmental factors. This result is relevant to a previous study that stated that glass, plastic, or coated resin containers are recommended for storing formulas with carbopol content³⁹. Colour changes were observed in the formulas 1%, 3%, 5%, and 7%, yellowish-white in Week 3 to brownish-yellow in Week-4. The carbopol will change colour if there is resorcinol and is incompatible with phenols, cationic polymers, strong acids, and high levels of electrolytes⁴⁰. Phytochemical tests on B. racemosa seeds showed that one of the secondary metabolites was phenolic compounds, namely tannins⁴¹.

B.racemosa seeds also contained triterpenoids and saponins. The main elements found in triterpenoids and saponins are Asiaticosides and madekosides⁴¹. Asiaticosides are yellow, so when dissolved with clear propylene glycol, it will cause the propylene glycol to turn yellow ⁴². Each formula has a specific B. racemosa seeds smell because no additional fragrance was added to remove the smell of B. racemosa seeds. The preparation could be categorized as good quality and stable if no change in shape, color, and odor observed between 48 hours to one month of storage⁴³.

All gel preparations were homogeneous during 4 weeks of observation. Although there were bubbles, no coarse grains on a pair of transparent glass plates were observed. The pH test showed a significant effect of the B. racemosa gel formula concentrations on pH. The pH plays a vital role in gel formation, viscosity, and gel strength. Carbopol based gel can be formed by neutralization at a pH between 5-10 using an amine base such as triethanolamine⁴⁴. A lack of preservative (methylparaben) to the preparation can cause a shift in pH during one month of storage. Bacteria can grow and create an acidic atmosphere like gram-positive bacteria, resulting in a decrease in the pH of the preparation⁴⁵. The pH test aims to adjust the pH conditions of the preparation with the pH of the skin and to see changes in pH during the storage process for one month. This test is one of the critical points to determine whether the preparation is irritating or not. The recommended pH is between 4.5-6.5 as this range as close to the physiological pH of human skin. The range of pH values in this study shows that this gel preparation is comfortable to use⁴⁶.

This study showed a significant effect of B. racemosa concentrations on the viscosity of the gel, which was still within the standard (SNI 16-4399-1996: 6000-50000cP or 6-50 Pa.S). Carbopol 940 is a polymer that forms tight coiled rolls in powder form, which limit its thickening ability. Carbopol 940 will be hydrated when dispersed into water which result in the open of coils⁴⁷. The function of carbopol 940 is best when the constituent polymers are completely uncoiled. This is a neutralization mechanism of carboxylic acid groups on the polymer chain with a suitable base. This reaction will result in negative charge formation along the polymer chain. Then, neutralization is carried out by addition of TEA which causes the carbopol 940 free the tight coiled structure. Carbopol 940 polymers will form a cross-link to produce a three-dimensional matrix to create a thick gel instantly⁴⁸.

Research has been undertaken to evaluate the herbal gel formulation. In one study, the gel was prepared using Carbapol 940, Murraya koenigii leaf extract, propylene glycol, methylparaben, propylparaben, glycerine, and distilled water. The dropwise addition of triethanolamine maintained the skin pH (6.8-7)⁴⁹. The inclusion of carbopol in the formulation of polyherbal aqueous gel from Psidium guajava, Piper betle, and Glycerrhiza glabra extract for the treatment of mouth ulcers appears to result in a transparent, homogeneous gel with a pH range of 6.8-7⁵⁰. Another study also showed that 7% carbopol gel formula with diclofenac had a clear colour, soft, pH of 6.84, and viscosity of 2700. This gel was observed for up to 8 hours with no signs of skin allergies. Finally, the gel formulations were found to be economical and may overcome the drawbacks associated with the drug during its absorption³⁰.

CONCLUSION:

This research proves that the addition of various concentrations of B.racemosa kernel extract to gel formulation affects the physical stability of the gel, namely on colour, pH, and viscosity.

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

ACKNOWLEDGMENTS:

The authors would like to thank Universitas Sumatera Utara Research Institute and the Chancellor of USU for funding this research under the USU TALENTA implementation contract for 2019, fiscal year number: 439/UN5.2.3.1/PPM/KP-TALENTA USU/2019 dated April 1, 2019.

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Received on 19.08.2022 Modified on 28.10.2022

Research J. Pharm. and Tech 2023; 16(9):4271-4275.

DOI: 10.52711/0974-360X.2023.00699