INTRODUCTION:

Corona Virus Disease 2019 (COVID-19) began to spread since March 2020¹. These are from cluster of viruses that cause disease affecting respiratory function of the body². World health organization estimates, 25 million people worldwide die of cardio vascular disease each year. Aerobic and breathing exercises are found to be effective in reducing the heart rate, respiratory rate and blood pressure³. WHO explained that one of the ways to prevent and transmit the virus is by cleaning our hands regularly using an antiseptic compound. Antiseptic is a chemical compound that is used to inhibit or kill microorganisms in living tissue, which has a limiting effect and prevents infection from getting worse⁴.

Antiseptics are used for handsanitizer formulation consisting of alcohol groups (ethanol, propanol, isopropanol) by concentration ±50% to 70% and phenol groups as chlorhexidine and triclosan with levels of 0.05% - 2%². Triclosan can inhibit the growth of bacteria, has anti-fungal, antiviral properties and less corrosive. However, using handsanitizer containing triclosan for a long time can cause skin irritation⁵. Skin irritation can occur when triclosan contact with skin that is directly exposed to sunlight with symptoms of a rash on the skin^6,7,8.

One of the way to reduce the use of triclosan for handsanitizer formulation is by using extract of plantation that have antibacterial properties. Plants are rich source of secondary metabolites with interesting biological activities (inhibition of certain pathogens)⁹. Roughly, 20% of the plants found in the world have been submit to pharmacological or biological test¹⁰. Aloe vera is a plant originating from the African continent and is currently being developed and cultivated in Indonesia, especially in West Kalimantan. It has triangular, beefy leaves with serrated edges, yellow tubular blooms and apples and oranges that contain various seeds (Aloe Vera)¹¹. The characteristics of aloe vera are green, spiny and large leaves that contain a lot of gel¹².

Aloe vera has many advantages including alkalizing the body, the body's immune system, detoxification, reducing body weight and cardiovascular health. Gel of Aloe vera contains an active compound, namely saponin, which have the ability to kill microorganisms. Saponin has function as cleaners and has antiseptic properties¹³. In this study, the dispersion power, adhesion test, homogeneity test, viscosity and antiseptic test were carried for handsanitizer formulation gel Aloe vera.

MATERIALS AND METHODS:

Materials:

Mortar and pestle, knife, petri dish, bottle, rotary evaporator, Brookfield viscometer, paper disc, centrifuge, hotplate, oven, incubator, erlenmeyer, analytical balance, vial, test tube, ose, Bunsen, tweezers, and other glassware.

Aloe vera, 70% ethanol, aquadest, Nutrien Agar media, Potaos Dextros Agar media, test suspension, Meyer reagent, chloroform, Carbopol 940, TEA, methyl paraben, glycerin, Fe(Cl)3, Magnesium powder, HCl (p).

Extraction of Aloe vera:

Leaves of Aloe vera are washed and separated between the skin and gel. 500grams of Aloe vera gel as was mashed using a blender then soaked in 1000ml of 70% ethanol solvent during 24 hours in a closed jar. Then, the liquid extract was filtered with a cloth filter and placed the extract in an Erlenmeyer tube. The extract was evaporated for 4 hours using a rotary evaporator, then placed on a water bath for 2 hours¹².

Formulation of gel handsanitizer:

20mL of aquadest was added into mortar then sprinkled with 0.5g carbopol 940. Then, stirred and added with alkalizing agent. For optimation handsanitizer formula, the alkalizing agent was varied that is TEA at pH 5.01, hydroxypropyl methylcellulose (HPMC) at pH 4.76 and stearyl alcohol at pH 4.52. Then, 0.2g methyl paraben was dissolved in 5mL of 70% alcohol and poured into a mortar then stirred until homogeneous. Extract of Aloe vera was weighed with the appropriate concentration then dissolved in 55mL of alcohol and stirred. Aloe vera that has been dissolved is placed into a mortar and stirred until homogeneous. Then, the solution was moved to a calibrated beaker glass, added with aquadest until the mark is 100mL and stirred until homogeneous¹⁴.

Spreadability test:

0.5grams of gel preparation by each formula was weighed. Then, the gel was placed under the round glass accompanied by a diameter scale, then covered with another glass which had been weighed and left for one minute. Then, diameter of the spread was measured. After 1 minute, a load of 50grams was added and left for 1 minute, then the diameter of the spread was measured. The same thing is done every 1 minute with the addition of a load of 100grams and 150grams respectively until a sufficient diameter is obtained to find the effect of the load on the spread diameter of the gel preparation¹².

Adhesion test:

0.25grams of gel by each formulation was placed in two glass objects that have been determined, then pressed with a load of 1kg for 5 minutes. Then, the glass object is attached to instrument test and added 80gram of load. The release time from the object glass was calculated¹².

Homogeneity test:

0,1g gel handsanitizer for each formula was weighed. Then each sample is placed on a slide glass and observed under a microscope at a magnification of 100 times¹².

Viscocity test:

The aim of viscosity test is to determine the thickness of the gel preparation using a viscometer. The gel sample was put into a beaker glass and placed under the spindle hanger. The spindle is mounted on the spindle hanger, then the spindle is lowered to the limit immersed in the Aloe vera extract gel preparation, then the rotor is turned on while pressing the button. The spindle is allowed to rotate and a red needle is observed on the scale, then the number indicated by the needle is read¹⁵.

Antiseptic test:

Control:

The palms are washed with tap water, then dried.The thumb prints were attached to nutrient-dense media in a petri dish. Media was incubated at 37C for 24 hours. After incubation, the number of colonies was counted.

Test Preparation:

The palms are washed with water, then dried. Then dropped the gel on the palms, leveled and left for one minute. The thumb prints were attached to nutrient-dense media in a petri dish. Media was incubated at 37 C for 24 hours. After incubation, the number of colonies was counted¹⁶.

RESULT AND DISCUSSION:

Extraction of Aloe vera

Aloe vera has physical characteristics including thick fleshy leaves, long, narrow edges, green and slimy as shown in Figure 1.

Figure 1. Aloe vera

In the extraction process, Aloe vera was cleaned first then cut into small pieces. Then, 1 kg small pieces of Aloe vera was blended until smooth. Then, it was filtered until the pulp of Aloe vera is separated from the filtrate solution which is reddish purple as shown in Figure 2.

Figure 2. Extract of Aloe vera

Formulation of gel handsanitizer:

Formulation of aloe vera extract gel preparation designed in this study refers to the formula that has been used by Johan Iswara Wijaya in 2013⁹. Formulaton of gel handsanitizer was varied based on alkalizing agent that is TEA, HPMC and stearyl alcohol which can be seen in Table 1.

Table 1. Gel handsanitizer formulation of Aloe vera

S. No	Material	Formulation
S. No	Material	I	II	III
1.	Extract of Aloe vera	100 %	100 %	100 %
2.	Alkohol 70 %	60 mL	60 mL	60 mL
3.	Carbopol 940	0.5 g	0.5 g	0.5 g
4.	Alkalizing agent	TEA (pH 5.01)	HPMC (pH 4.76)	Stearyl alkohol (pH 4.52)
5	Glycerin	1 mL	1 mL	1 mL
6	Methyl paraben	0.2 g	0.2 g	0.2 g
7	Aquadest	Up to the mark 100 mL	Up to the mark 100 mL	Up to the mark 100 mL

Alkalizing agents act as gel stability which can balance the pH of the preparation. It also acts as a pH neutralizer for carbopol to form a clear solution so that the colour is transparent¹⁷. Based on research conducted by Putri¹⁸, alcohol 70% for handsanitizer formulation works faster to deposite protein and destroy lipid membranes, especially in staphylococcus than alcohol by concentration 40-60%. Handsanitizer containing alcohol 70% can kill almost 90% of skin bacteria because it can penetrate the cell walls. Methyl paraben was used in handsanitizer formulation because it prevenst the growth of microorganisms due to the addition of distilled water to fill the mass of 100g gel preparations. Carbopol 940 is used as a base for gel preparations and glycerin has function as a moisture barrier which can increase the spreadability and protect from the possibility of drying out¹⁴. Each formulation of handsanitizer produce a clear, slightly cloudy color of handsanitizer as shown in Figure 3.

Figure 3. Handsanitizer of Aloe vera extract (A) Formulation I (B) Formulation II (C) Formulation III

Spreadability test:

Spreadability test is a test to demonstrate the ability of gel to spread when applied to the skin. Mass of load used for spreadability test is 50g, 100g and 15g by certain pH that can be seen in Table 2.

Table 2. Spreadability test of gel handsanitizer Aloe vera

Mass of load	Spreadability (cm)
Mass of load	F I	F II	F III
50 g	2.90	8.25	0.00
100 g	3.00	7.30	6.87
150 g	3.05	3.50	7.70

The value of pH used for gel is a pH that almost or close to the pH of the skin, which is around 4.5-6.5¹⁹. The dispersibility of the semisolid which good enough for gel ranges in diameter from 5 - 7cm which produces the comfort consistency to the skin when used²⁰. The spreadability tends to increase by the addition of the load mass where gel handsanitizer is good enough to be used if it has a spreading power ranging from 5 - 7cm both small and large weight additions. Based on Table 3, it was found that HPMC has a broad spreadability compared to TEA and stearyl alcohol. HPMC is a gelling agent that belongs to the polysaccharide group so that it expands easily and has a smaller viscosity²¹.

Homogeneity test:

The homogeneity test of gel handsanitizer is carried out by applying the gel preparation to a piece of glass or other certain transparent material. Then, the homogenity can be obtained. Gel handsanitizer have to produce a homogeneous composition and no visible coarse grains ²².

Based on the results of the homogeneity test in Figure 4, it can be concluded that the gel handsanitizer Aloe vera by each formulation has a homogeneous texture when applied to the skin. It can be sure with the absence of coarse grains in the gel and a homogeneous composition. Homogeneous structure indicates that the active compounds, alkalizing agents and additional materials are evenly distributed for gel handsanitizer.

Adhesion test:

The adhesion test of gel handsanitizer gel was carried out to obtain the ability of gel to adhere for skin. It can affect the ability of gel handsanitizer to penetrate into the skin and produce the effect²³.

Table 3. Adhesion test of gel handsanitizer Aloe vera

Formulation	I	II	III
Time (minutes)	2.44	1.20	1.25

Well standard of adhesion allows the gel not to come off easily when applied and has a long adhesion time. Standard adhesion for topical preparation is more than 4 seconds.Based on Table 3, gel handsanitizer Aloe vera by each formulation has adhesion time more than 4 seconds and the longest adhesion time is formulation I, which is about 2.44 minutes²⁴.

Viscosity test:

Viscosity is a measure of the consistency of a fluid which states the size of the fluida friction. The apparent viscosity is determined by the careful observation of shear stress in the dense phase²⁵. The greater viscosity of the fluid, it will be more difficult for fluida to flow and an object will be difficult to move within the fluid. Viscosity has a considerable influence on spreadability and adhesion in which well standard viscosity is 2000 - 4000cps²⁰. If the value of viscosity is greater , the longer the adhesion time will be when applied.

Table 4.Viscosity test for gel handsanitizer Aloe vera

Formulation	I	II	III
Viscosity (cps)	837	64.70	21.50

Based on Table 4, the viscosity of the alkalizing agent in each formula is still low. This is due to possible technical errors during experiment. Formulation I has the largest viscosity value about 837cps and formulation III has the smallest viscosity of 21.50cps. Formula I contain TEA as alkalizing agent in which it has capability to pack the molecules of the active substance on the surface will be stronger so can increase the strength of the coating interface and add stability of gel²⁶. HPMC is alkalizing agent used by formulation II. It is a cellulose-derived polymer, which when there is a disperse of this polymer molecule into a cavity formed by water molecules, so there is a hydrogen bond between the hydroxyl (- OH) group of the polymer and the water molecule so its viscosity is smaller than TEA ²⁷. Meanwhile, stearyl alcohol can increase gel stability by increasing the consistency of the cream in the presence of a water-soluble emulgator so that the viscosity is smaller¹⁷.

Antiseptic test:

Antiseptic is the chemicals used in destroying disease causing micro-organism (also called microbial, microbiological, pathogens) externally on wound or (under medical supervision) taken internally to treat infection²⁸. In this research, antiseptic activity was tested using the replication method²⁹. Control that was used in antiseptic test was the thumb of the right hand because it had a wider surface and was assumed to be used more frequently so that it could represent the total number of microbes present in the palm and left hand. The control test was carried out to estimate the number of microbes that were still present and that could be killed on the palms after adding the gel handsanitizer Aloe vera. Each formulation of gel handsanitizers Aloe vera was smeared to the palm then left for 1 minute. Then, we can determine the number of microbial colonies that still alive as shown in Figure 5.

Figure 5. Antiseptic test for gel handsanitizer Aloe vera (A) Control (B) Formulation I (C) Formulation II (D) Formulation III

The number of colonies obtained for control was 107 microbes. After using gel handsanitizer Aloe vera on the right hand, it was found that the number of microbial colonies that were still alive using formulation I was 71 colonies¹⁶. Meanwhile, the number of microbial colonies that were still alive with formula II was 29 colonies and using formulation III was 14 colonies which can be seen in Table 5.

Table 5. Antiseptic test for gel handsanitizer Aloe vera

Treatment	Number of microbes	Number of dead microbes
Control	107	-
Formulation I	71	36
Formulation II	29	78
Formulation III	14	93

CONCLUSION:

Based on the research, it can be concluded that gel handsanitizer Aloe vera by three formulations has a fairly homogeneous structure when applied to skin. Formulation II has a broad enough spreadability for gel handsanitizer. Meanwhile, handsanitizer with the Formulation I has the greatest viscosity value, about 837 cps and the longest adhesion value of about 2.44 minutes. Based on antiseptic power test, it was found that the handsanitizer with formulation I was able to kill more microbial colonies, about 71 microb colonies from the available 107 microbes. It is known that the handsanitizer formulation I is a handsanitizer with a variety of alkalizing agents in the form of TEA at pH 5.01.

CONFLICT OF INTEREST:

The authors declare no conflicts of interest

ACKNOWLEDGEMENT:

This work was supported by grants from Politeknik AKA Bogor.

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Received on 17.08.2021 Modified on 09.11.2021

Research J. Pharm. and Tech 2022; 15(9):4037-4042.

DOI: 10.52711/0974-360X.2022.00677