Nika Adriana Marijanović, 2.

e
Zagreb, Croatia
Mentor: Ana Sladoljev
E-mail: nikamarijanovic10@gmail.com
Mob: +385 98 192 4878
E-mail: ana.sladoljev@skole.hr (mentor)
Mob: +385 99 697 8971 (mentor)
Klasična gimnazija Zagreb, Križanićeva 4A

EFFICIENCY AND FUNCTION OF SUNSCREEN

INTRODUCTION

Sunlight consists of different spectrums of radiation such as visible light, ultraviolet (UV) and
infrared light. Light is measured in wavelengths (λ), and the unit of measurement is
nanometer (nm) and millimeter (mm). Different light radiations in the spectrum have
different wavelengths. Sunlight can have positive and negative effects on human health.
Sunlight is inducing calciferol (vitamin D) synthesis which is of great importance for our
body; utilizes calcium and phosphate ions from food through the digestive tract, stabilizes
the concentration of calcium ions in plasma necessary for normal function of stimulus cells,
signal transduction within cells and especially for bone mineralization. Vitamin D also
increases our levels of serotonin whose function contributes to better mood. On the other
hand, negative effects can overshadow the positive effects with a lack of caution when
exposed to sunlight for longer periods of time. Some of the consequences can be burns,
premature skin aging, hyperpigmentation, allergic reactions and more serious conditions
such as skin cancer. These kind of skin damages caused by the sun are mainly due to UV
rays1. Ultraviolet light is divided into UVA, UVB and UVC rays. Ultraviolet C rays have the
shortest wavelength (between 100 and 290 nm) and do not reach the skin due to the ozone
layer that absorbs rays up to approximately 290 nanometers. UVA and UVB rays have a
wavelength of 280 nm to 400 nm while UVA has a longer wavelength than UVB. Together
they make up about 5% of the Sun’s rays. UVA and UVB rays have a detrimental effect on
the skin, but the intensity of UVA rays is constant throughout the day, whilst the intensity of
UVB rays varies. Therefore, there are many different products on the market that can
protect our skin from damaging rays. The sun protection factor rating (SPF) system for
sunscreens is based on the level of UVB protection offered by the product. Preparations
containing the UVA protection symbol must ensure a minimum level of protection against
UVA rays. The preferred ratio is 1: 3 for UVA: UVB protection. SPF multiplied by natural skin
protection in minutes determines the maximum length of time you can expose your skin to
the sun without the risk of UVB-induced skin damage. However, SPF protection depends on
the use of the right amounts (2mg / cm 2) of the product, therefore 60% of the time allowed
by SPF is recommended2.

GOAL OF THE EXPERIMENT

The aim of this experiment is to examine the effectiveness and quality of different
sunscreens. My hypothesis of this research is that sunscreens with a higher factor will
provide better cell protection.

MATERIALS AND METHODS

For this experiment, I used a total of 6 different sunscreens, of which 3 with a factor of 50
and the other 3 with a factor of 30. These are: Vichy (SPF 50), Eucerin (SPF 50), Avon (SPF
50), Eucerin (SPF) 30), Nivea (SPF 30) and Clarins (SPF 30).
Figure 1. Sunscreens used in the experiment

The MTT test is a standard test that measures the survival (viability) of cells after a
treatment. In this experiment it was specifically used to measure cell survival after exposure
to UVB radiation.

Human embryonic kidney cells HEK293 were grown in DMEM cell culture medium with the
addition of fetal bovine serum (FBS). When they reached 80% confluence, 96-well plates
(Sarsted) at a concentration of 5 x 10 4 cells /200 μL medium were seeded in each well and
allowed to adhere overnight in a humidified incubator with 5% CO 2 at 37 ° C (Luna counter
was used). The next day, the medium was removed from the cells and the cells were
covered with foil containing sunscreen. The surface of the foil was approximately 2 cm 2, and
the amount of applied creams was 2 mg / cm2, following the recommendations of the
Guidelines for International Methods of Testing Sun Protection Factors. Cells were exposed
to UV radiation at a wavelength of 312 nm and a dose of 89mJ / cm 2 (Figure 2). The medium
was then recovered and the cells were incubated at 37 ° C, 5% CO2 for 24 hours. Each
sample was tested in 6 copies. As controls, we used UV-permeable foil (F), UV-proof glass
cover (C) and uncovered cells (0).

After an incubation period (24 h), cells were photographed with a Zeiss Axiovert inverted
microscope, then the medium was removed and the cells were treated with 40 μL /well of
MTT solubilized in cell medium at a concentration of 0.5 mg / ml. After 4 hours of incubation
of the cells with MTT, dimethylsulfoxide ((CH3) 2SO) was added to each well to dissolve the
crystals formed. After 15 minutes of dissolution, absorbance was measured at 560 nm using
a microplate reader (Glomax-Multi). Cell viability was calculated as the percentage of
untreated control after reducing the absorbance of the blank to all samples.
Figure 2. Cell exposure to UVB radiation with the use of UVItec Crosslinker.

Formulas used when processing the results for calculating the mean, standard deviation and
absolute error:

x 1+ x 2 +…+ x n
x=
n

√
2 2 2
( x−x 1 ) + ( x−x 2 ) +…+ ( x−x n )
s=
n

∆ x i=x i−x

Formulas used in processing the results to calculate cell survival:

absorbance of the sample

Viability (%)= ×100
absorbance of the control

RESULTS

In this study, six different sunscreens were tested. Microscopic photography could
distinguish the approximate percentage of living cells and the difference in the effectiveness
of individual creams (Figure 3). Live cells have a spindle shape while dead ones take on a
round shape. From this finding, it was concluded that Vichy 50, Eucerin 30 and Eucerin 50
had the best protection effect, while worse results were seen in Clarins 30 and Nivea 30.
That was confirmed by MTT results (Figure 4). I calculated the arithmetic mean, maximum
absolute error, standard deviation, and viability of each individual cream and control. Later,
I presented this processed data graphically in Excel and tabularly in Word (Figure 5 and
Table 1). From the above data, I concluded that the best cell protection was provided by
Eucerin 30 cream with a viability of 111.35%, followed by Vichy 50 (101.39%), the third Avon
50 (98.59%), followed by Eucerin 50 (95.64). %) and with lower viability Nivea 30 (79.3%)
and Clarins 30 (78.02%). Avon 50 cream had the lowest data reliability with a maximum
absolute error of as much as | 0.26 | and a standard deviation of 9.58. Vichy 50 cream of |
0.234 | also had a big deviation and a standard deviation of 27.49. Creams with a smaller
but not negligible maximum absolute error are Eucerin 50 with |0.129| and a standard
deviation of 15.94 and a Nivea 30 with |0.105| and 12.74. Eucerin 30 sunscreen had the
most accurate and reliable results with a maximum absolute error of |0.067| and a standard
deviation of 8.95 while the second one is Clarins 30 with a value of |0.096| and 12.22. The
price of Eucerin 30 is 220 kn, Eucerin 50 (170 kn), Vichy 50 (160 kn), Nivea 30 (130 kn) and
Clarins 30 (240 kn), and the price does not reflect the quality for all of the samples.

Figure 3. Microscopically photographed HEK293 embryonic kidney cell lines after exposure
to UVB radiation (Zeiss Axiovert inverted microscope at 100x magnification) and covered
with different sunscreens, UV-permeable foil (F), UV-proof glass cover (C) and uncovered
cells (0).
0,8945988 0,6177315 0,6700892 0,7331722 0,7662081 0,7329813 0,5163481 0,4789908 0,4430747
0,6763033 0,6331671 0,4744013 0,6434307 0,7096316 0,6764067 0,5509482 0,517353 0,5563955

0,7162511 0,67269 0,6040702 0,6030453 0,5233287 0,5502784 0,4690163 0,4004703 0,4889798

0,6025308 0,5036646 0,6965867 0,6239867 0,446658 0,5636218 0,4323708 0,3966439 0,4797699

0,8860403 0,4881655 0,5081111 0,5633579 0,6419561 0,5534847 0,559563 0,6538106 0,6330156

0,907659 0,576495 0,5168293 0,4827611 0,4779399 0,5534712 0,5882223 0,7812275 0,70917

BLANK 0

Figure 4. Display of measured absorbances of MTT test after exposure of HEK293 cell lines
to UVB radiation (Excel). Absorbances were measured on a Glomax-Multi Promega at 560
nm.

Cell viability after exposure

120

100

80

60

40

20

0
C F O Eucerin 30 Nivea 30 Clarins 30 Eucerin 50 Vichy 50 Avon 50

Figure 5. Viability graph of HEK293 cells after UVB exposure. The X axis represents the
creams that covered the cells (2mg / cm 2), and the Y axis represents the absorbance values.
All data are presented as mean cell viability expressed as a percentage of negative control ±
standard deviation (Excel). (C-cells covered with glass cover, F-cells covered with
transperent foil, O-cells uncovered)
Table 1. Display of means and absolute maximum error (Word).

C F 0 Eucerin Nive Clarins Eucerin Vichy Avon

30 a 30 30 50 50 50
x 0,494 0,35 0,285 0,55 0,392 0,386 0,473 0,50 0,487
1
Δx max 0,127 0,067 0,048 0,067 0,105 0,096 0,129 0,23 0,26
4

CONCLUSION

After all the information gathered from this experiment, I can conclude that
the sunscreen Eucerin 30 provides the best protection against UV radiation,
and Clarins 30 the worst, even though it is the most expensive product. I have
also noticed that creams with a thicker composition also have a better effect.
Spray sunscreens such as Nivea 30 and Clarins 30 provided the weakest
protection.

LITERATURE

1. Dora Gašparini, Marija Kaštelan. Vitamin D i koža, Medicina, Vol. 57 No. 4, 2021.

2. André P. Schuch, Maria Carolina S. Moraes, Teiti Yagura, and Carlos F. M. Menck
Highly Sensitive Biological Assay for Determining the Photoprotective Efficacy of
Sunscreen, Environ. Sci. Technol. 2014, 48, 11584−115

Internet sources (accessed on April 24, 2022):

https://www.eucerin.hr/o-kozi/osnovni-podaci-o-kozi/sunce-i-koza
https://hr.wikipedia.org/wiki/Ultraljubičasto_zračenje
https://www.enciklopedija.hr/natuknica.aspx?ID=63114
https://en.wikipedia.org/wiki/Vitamin_D