INTRODUCTION:

Sunscreen is a substance that contains sun protection properties so that UV rays cannot penetrate the skin. Sunscreen can protect the skin by either spreading sunlight or absorbing solar radiation energy that hits the skin so that the radiation energy does not directly hit the skin. The sunscreens' effectiveness can be expressed by sun protection factor (SPF), percentage of erythema, and percentage of pigmentation transition¹.

Red Pidada leaves (Sonneratia caseolaris L.) are natural ingredients, one of which is useful as a sunscreen². This leaf in South Kalimantan is used as a cold powder mixture³. It has antioxidant compounds containing alkaloids, flavonoids, glycosides, saponins, and phenols⁴. Based on scientific research, this leaf has an SPF value of 1.601 with minimal protection in methanol extract⁵.

The use of ethanol extract as a solvent has never been reported or formulated as a sunscreen, so the researchers preformulated the red Pidada leaf ethanol extract as sunscreen. The pre formulation chosen was the cream product with the best activity as a cosmetic product prototype. The evaluation of the cream preparations included an organoleptic test, pH test, homogeneity test, spread ability test, adhesion viscosity test, and a sunscreen effectiveness test. The choice of cream as a dosage form is because it has the general properties of adhering to the surface where it is used, practical, easy to apply or wash, and does not contain fat like ointment preparations⁶.

MATERIALS AND METHODS:

Materials:

Red Pidada Leaf Extract, Cera Alba, Liquid Parafin, TEA, Stearic Acid, Glycerine, Methyl Paraben, Prophyl Paraben, Aquadest

Methods:

The ingredients were separated based on their phases. The oil phase (Cera alba, liquid paraffin, stearic acid, and nipasol) and the liquid phase (TEA, glycerin, and nipagin) were heated until dissolved. The oil phase is put into a hot mortar and then mixed with the water phase until it is homogeneous. Added extract, crushed until homogeneous, and into a cream preparation. Evaluation of cream preparations includes evaluation of appearance by looking directly at shape, color, odor, homogeneity, and pH. Homogeneity tests by applying 0.1 gram of cream to be tested on a piece of glass or other suitable material must show a homogeneous structure and do not show coarse grain⁷. Measurement of the scattering power, namely: as much as 0.5grams of the sample is placed on another glass plate for 1 minute, and then the diameter of the spreader is measured. Followed by adding a load of 150grams to the glass plate for 1 minute, then measured the diameter of the spread⁸. The adhesion test is: 0.25g of cream is placed on a glass object and pressed with a load of 1kg for 5 minutes. The object-glass is attached to the test kit. The assay was given a load of 80g and then the time to release the cream from the slide was recorded⁹.

The effectiveness of the cream was determined by determining the SPF value in vitro using UV-Vis spectrophotometer. The test absorption curve is made in a curve, with a wavelength between 290-320nm, using chloroform as the blank. The average absorption is determined at 5nm intervals. Mains solution was made by weighing 0.01g of cream, then put it in a 10mL volumetric flask dissolved with chloroform until the limit mark. Bioactive compound analysis was carried out using LC-HRMS. Before being injected into the LC-HRMS, a sample preparation is performed first. The extract sample (liquid) was diluted with its polar (water soluble) solvent to a final volume of 1300μL. Dilution is done by looking at the density of the sample (not too concentrated and not too dilute). Then, the samples were vortexed for 1 minute. Its purpose is to homogenize the solution. Furthermore, the samples were spindown (300 RPM) for 2 minutes. Spindown is carried out to separate the components in the sample solution based on density; the result is a supernatant (liquid) and a precipitate. Then, 10μL of the supernatant was taken and filtered using a 0.22μm PTFE filter syringe and put into the vial. The sample in the vial was put into an auto sampler and injected into LC-HRMS.

RESULT:

Evaluation of the appearance of red pidada leaf ethanol extracts cream preparations:

The display results of this cream on the blank, F1, F2, and F3 have a semi-solid shape, odorless with cream color on the white blank, and dark green formulas. The homogeneity test results were only F3, which was not homogeneous, and all creams had a ph of 7.50-7.83 (Table 1).

Table 1. Evaluation of the appearance of red pidada leaf ethanol extracts cream preparations

Parameters	Formula
Parameters	Blank	F1	F2	F3
Shape	Semi-solid	Semi-solid	Semi-solid	Semi-solid
Odor	None	None	None	None
Color	White	Dark green	Dark green	Dark green
Homogeinity	Homogeneous	Homogeneous	Homogeneous	Homogeneous
pH	7.83±0.25	7.70±0.10	7.60±0.10	7.50±0.10

Information :

Blank : Cream without extract

F1 : Cream with an extract concentration of 8%

F2 : Cream with an extract concentration of 15%

F3 : Cream with an extract concentration of 20%

Result of measuring the viscosity of the red pidada leave extract cream:

The results of measuring the viscosity of the four formulas show that the blank has the lowest viscosity value and decreases on day 1 of 23533 cP, day 7 of 22166 cP, and day 14 of cP 19133 cP, while formulas 1, 2, and 3 have viscosity higher (figure 1).

Figure 1. Result of measuring the viscosity of the red pidada leave extract cream

Spread ability test:

The spread ability test is at 2.555-4.129cm (table 2), and the results of the average adhesion in 14 days have increased. The blank has the highest adhesion power on the 1st day of 16.36 minutes, the 7th day of 29.52 minutes, and the 14th day of 37.48 minutes, followed by the lowest F1, F2, and F3 (Figure 2)

Table 2. Spread ability test of pidada red leaf extract cream

Spread ability Test (Cm)
Formula	Load weight (g)	Day to
Formula	Load weight (g)	1	7	14
Blank	0	2.555	2.786	3.070
Blank	150	3.986	4.108	4.129
1	0	2.489	2.553	2.684
1	150	3.224	3.271	3.552
2	0	2.218	2.409	2.636
2	150	3.168	3.245	3.536
3	0	2.096	2.318	2.598
3	150	2.929	3.218	3.427

Mass spectroscopy

Figure 2. Average adhesion of red pidada leaf extracts cream

The physical quality test of cream shows that F1 and F2 meet the organoleptic test (shape, smell, and color), homogeneity test, and adhesion test but do not meet the pH viscosity test and spread ability test. F3 meets the organoleptic test requirements and adhesion test requirements but does not meet the homogeneity test, pH test, viscosity test, and spread ability test.

Sunscreen effectiveness test:

The sunscreen effectiveness test on F1 has the lowest Sun Protection Factor (SPF) value of 1.5 (without protection), F2 has an SPF value of 2.9 (minimal protection), and F3 has the highest SPF value of 7.4 (extra protection). (Table 3).

Table 3. Effectiveness test of sunscreen for the cream

Formula	SPF Value	Category
1	1.5	Without Protection
2	2.9	Minimum Protection
3	7.4	Potection Extra

Analysis LC-HRMS:

The results of the sample analysis using the LC-HRMS, the resulting data is in the form of a chromatogram. The chromatogram describes the number of components in the sample seen from the number (peak) produced¹⁰. The compound content contained in red pidada can be seen in the following table 4.

DISCUSSION:

There are several works carried over on formulation of gel, lotion and cream as antioxidant, UV protective Sunscreen, antiaging, cosmetic cream, treatment of Psoriasis type of diseases from different herbal products^11-21.

In this study, the cream was formulated from the ethanol extract of pidada red leaves with extract variations, namely 8%, 15%, 20%. The choice of cream preparations is because the cream preparations are easy to spread evenly, they are practical, the way it works is on the local tissue, it is easy to wash, especially the M/ A type, and gives a cold feeling²². Observations of the four blank cream preparations, F1, and F2, meet the homogeneity requirements where there are no coarse grains and the ingredients are evenly mixed, while in F3, there are coarse grains in the cream. An inhomogeneous preparation and coarse granules occur due to the high amount of extract in the formula²³. The acidity (pH) value for topical preparations ranges from 4-8²⁴. This cream's pH value is 7.50-7.83, fulfilling the pH value for topical preparations, namely 4-8. But it is not by the pH value of human skin, namely, 4.5-6.5, because the presence of anionic emulgator, namely triethanolamine in a cream base, can affect the base pH and the base's stability. The greater the concentration of triethanolamine added, the greater the pH of the resulting cream base. This indicates that triethanolamine is not only an emulgator but also increases pH²⁵. The viscosity value describes the consistency of the cream preparation²⁶. Based on SNI 16-4399-1996 regarding sunscreen preparations, a cream preparation's good viscosity value ranges from 2,000-50,000 cP^10,27. The test results show that formulas 1, 2, and 3 do not meet the SNI viscosity value while the blanks meet the SNI viscosity value.

The spread ability test aims to determine the ability of a cream preparation on the skin surface. Based on the results of the measurement of the spread ability, it can be seen that formula 1, formula 2, formula 3, and blanks do not meet the requirements of a good cream dispersion, namely 5-7 cm²⁸. The good adhesion requirement for topical preparations is more than 4 seconds²⁹. The result of the adhesion test for cream preparations shows that the blank has the longest adhesion power and formula 3 has the shortest adhesion power. The sunscreen effectiveness test aims to determine how much the SPF value is contained in the in vitro ethanol extract of the red Pidada leaves sunscreen cream. The sunscreen effectiveness test results show that formula 1 has the lowest SPF value and formula 3 has the highest SPF value. Different extract concentrations can affect the SPF value in cream preparations. The greater the extract concentration, the higher the SPF value. SPF (Sun Protecting Factor) is a universal indicator that describes a product or sunscreen substance. The higher the SPF value of a product or sunscreen substance, the more effective it is to protect the skin from UV rays' bad effects²⁹.The data obtained were analyzed by statistical tests using the One Way Anova test and followed by the Least Significance Different (LSD) test, which showed that there was a significant difference (p <0.05) in the SPF value of all formulas. Based on this, formula 3 has a better sunscreen function than formula 1 and 2.

Red Pidada leaves have sunscreen activity because they contain flavonoids. The use of antioxidant substances such as flavonoids, tannins, anthraquinones, and cinnamates can prevent diseases caused by UV radiation. Several groups of active antioxidant compounds, such as flavonoids, have been reported to have the ability to protect against UV³⁰. From the analysis of bioactive compounds in red Pidada ethanol extract using LC-HRMS, of the 62 compounds obtained, there were 6 active compounds with the greatest abundance of ions, determined based on the cluster on the whole chromatogram graph. Data on the content of the suspected compounds and data on the comparison of compounds with the highest ion abundance. Red pidada bioactive compounds are agonists, namely ligands or drugs that can bind to receptors and produce effects on the body. Diisobutylphtalat, luteolin, apigethrin and vitexin are esters. This compound has various benefits for human life, including being a source of antioxidants, antibacterial, anti-inflammatory, and anti-cancer properties. Choline which is the parent compound of the choline group, consists of ethanolamine which has three methyl substituents that are bound to its amino function³¹. Choline and its derivatives function as components of structural lipoproteins, blood lipids and membranes, and as precursors for the neurotransmitter acetylcholine. Choline is oxidized to betaine which functions as an osmoregulator and is a subtract in the betaine-homocysteine methyltransferase reaction. Choline and cysteine can function as anticancer, cardiovascular disease and overcome folic acid deficiency³². The Sunscreen effectiveness test in red Pidada formula 1 has the lowest Sun Protection Factor (SPF) value of 1.5 (without protection) and the highest SPF value of 7.4 (protection extra) for a range of skin types³³. The Sunscreen effectiveness test in red Pidada formula 1 has the lowest Sun Protection Factor (SPF) value of 1.5 (without protection) and the highest SPF value of 7.4 (protection extra) for a range of skin types^34,35.

CONCLUSION:

The sunscreen effectiveness test in formula 1 has the lowest Sun Protection Factor (SPF) value of 1.5 (without protection), formula 2 has an SPF value of 2.9 (minimum protection), and formula 3 has the highest SPF value of 7.4 (protection extra). Analysis of bioactive compounds in red Pidada ethanol extract using LC-HRMS revealed 6 active compounds Diisobutylphtalat, luteolin, apigethrin, vitexin, choline, and bethain.

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

ACKNOWLEDGMENTS:

The research team would like to thank the Directorate of Research and Community Service, Ministry of Research and Technology / National Research and Innovation Agency, Indonesia for funding this research under the 2020 Applied Research Scheme.

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Received on 24.01.2022 Modified on 10.04.2022

Research J. Pharm. and Tech 2023; 16(2):781-785.

DOI: 10.52711/0974-360X.2023.00134

Name	Formula	Molecular Weight	RT [min]	Area (Max.)	mzCloud Best Match
Choline	C₅ H₁₃ N O	103.09992	0.896	1,120,349,812.89	97.6
Betaine	C₅ H₁₁ N O₂	117.07913	0.903	143,657,529.38	96.7
Diisobutylphthalate	C₁₆H₂₂O₄	278.15154	18.535	111,852,186.73	98
Luteolin	C₁₅H₁₀O₆	286.04756	7.717	40,366,144.38	99.7
Apigetrin	C₂₁H₂₀O₁₀	432.10542	8.451	32,530,378.21	98.1
Vitexin	C₂₁H₂₀O₁₀	432.10542	7.313	26,337,978.09	96.4