Photoprotective complementary effects of sun filters and a combination of active molecules on UV-exposed human volunteers
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Presented by: Arnaud Fontbonne
Introduction
Skin photoprotection has become a real public health issue in view of the consequences of the sun on unprotected skin such as erythema, immunosuppression, photoaging and skin cancers. The major role of UVA rays has long been neglected, even though their harmful effect over the long term, by notably generating oxidative stress. To prevent it, it has become important to provide biological protection in addition to sun filters.
The aim of this in vivo study was to evaluate the photoprotective complementary efficacy of an active complex with sun filters on volunteers exposed to UVs. Different parameters and biomarkers were analysed pre- and post-irradiation such as squalene oxidation, catalase activity, trans-urocanic acid (trans-UCA).
Methods
This study was conducted on 10 male volunteers aged from 20 to 44 years, with oily skin and a cutaneous sebum rate ≥30 µg/cm² (quantified using Sebumeter®). The samplings were performed on the back of volunteers by swabbing, a non-invasive technique.
In vivo protocol
At D0, a skin surface sampling was performed to determine the basal value of each biomarker for each subject, and their own MED was determined.
From D0 to D2 included (3 days), an application of the studied products was performed at the clinical centre by the technician (2mg/cm², twice daily), according to the 4 studied zones defined on the back of volunteers: placebo; sun filters SPF 30; the active complex containing 0.1% ectoine and 0.1% mannitol; sun filters SPF 30 in association with the active complex.
At D3 morning, after a standardized cleansing and a last products application (30 minutes before irradiation), a 2 MED UV exposition (UVA and UVB) was then performed on the 4 selected UV sub-zones for all conditions with a xenon lamp Monoport - 300W (Solar Light).
At D4, skin surface samplings were done on the 8 sub-zones of each volunteer (4 irradiated and 4 non-irradiated) and stored at -20°C before biochemical analyses.
Biochemical analyses
Squalene oxidation (quantity of oxidized squalene and non-oxidized squalene) was measured by HPLC system Ultimate 3000 (Thermo Scientific) coupled with a single quadrupole mass spectrometer detector ISQ (Thermo Scientific).
Urocanic acid was quantified with the same equipment.
Catalase activity was evaluated using the resorufin, a highly fluorescent product following oxidation, to indirectly measure the residual hydrogen peroxide (H2O2) content following the catalase reaction: the resorufin intensity is inversely proportional to the catalase activity.
Statistical analysis
For each parameter, normality was first checked to determine the right statistical test to applicate. If hypothesis of normality was approved, a Student's t-test was used, and if not, a Wilcoxon test. If the p-value was less than 0.05, the difference was significant.
Results
Compared to non-irradiated placebo condition, UV irradiation induced squalene oxidation by 2-fold, a decrease of catalase activity by 1.5-fold and a photo-isomerization of UCA by 5-fold.
Compared to the irradiated placebo, the active complex and sun filters alone protected squalene oxidation by 58.4% and 50.6%, catalase activity by 60.5% and 53.7%, and UCA photo-isomerization by 14.2% and 30%, respectively.
Combination of ectoine and mannitol with sun filters provided the better protection of squalene oxidation (76.8%; p<0.01), catalase activity (84.4%; p<0.001) and trans-UCA (53.9%; p<0.01).
Compared to filters alone, the active complex added to filters provided significant additional protection of squalene by 26% (p<0.05), catalase activity by 31% (p<0.05) and trans-UCA synergistically by 24% (p<0.01).
Discussion
To robustly study the biological damage induced by sun exposure, it seems essential to analyse biomarkers directly collected from volunteers. In this clinical study, we were interested in three biomarkers corresponding to three natural defence systems altered by UV: squalene for surface antioxidant, catalase which is an essential endogenous antioxidant enzyme, and photo-isomerization of UCA involved in preventing immunosuppression.
We showed that the combination of sun filters SPF30 with this active complex of 0.1% ectoine and 0.1% mannitol attributed a high protection capacity against squalene oxidation, decrease in catalase activity and decrease of trans-UCA induced by UV exposure.
Conclusion
This in vivo study demonstrates the interest of combining UV filters with a biological protection provided by specific active ingredients to allow optimal photoprotection and so to prevent UV damages induced by sun exposure such as induction of oxidation, decrease of endogenous antioxidant defence systems and induction of photo-immunosuppression.
Skin photoprotection has become a real public health issue in view of the consequences of the sun on unprotected skin such as erythema, immunosuppression, photoaging and skin cancers. The major role of UVA rays has long been neglected, even though their harmful effect over the long term, by notably generating oxidative stress. To prevent it, it has become important to provide biological protection in addition to sun filters.
The aim of this in vivo study was to evaluate the photoprotective complementary efficacy of an active complex with sun filters on volunteers exposed to UVs. Different parameters and biomarkers were analysed pre- and post-irradiation such as squalene oxidation, catalase activity, trans-urocanic acid (trans-UCA).
Methods
This study was conducted on 10 male volunteers aged from 20 to 44 years, with oily skin and a cutaneous sebum rate ≥30 µg/cm² (quantified using Sebumeter®). The samplings were performed on the back of volunteers by swabbing, a non-invasive technique.
In vivo protocol
At D0, a skin surface sampling was performed to determine the basal value of each biomarker for each subject, and their own MED was determined.
From D0 to D2 included (3 days), an application of the studied products was performed at the clinical centre by the technician (2mg/cm², twice daily), according to the 4 studied zones defined on the back of volunteers: placebo; sun filters SPF 30; the active complex containing 0.1% ectoine and 0.1% mannitol; sun filters SPF 30 in association with the active complex.
At D3 morning, after a standardized cleansing and a last products application (30 minutes before irradiation), a 2 MED UV exposition (UVA and UVB) was then performed on the 4 selected UV sub-zones for all conditions with a xenon lamp Monoport - 300W (Solar Light).
At D4, skin surface samplings were done on the 8 sub-zones of each volunteer (4 irradiated and 4 non-irradiated) and stored at -20°C before biochemical analyses.
Biochemical analyses
Squalene oxidation (quantity of oxidized squalene and non-oxidized squalene) was measured by HPLC system Ultimate 3000 (Thermo Scientific) coupled with a single quadrupole mass spectrometer detector ISQ (Thermo Scientific).
Urocanic acid was quantified with the same equipment.
Catalase activity was evaluated using the resorufin, a highly fluorescent product following oxidation, to indirectly measure the residual hydrogen peroxide (H2O2) content following the catalase reaction: the resorufin intensity is inversely proportional to the catalase activity.
Statistical analysis
For each parameter, normality was first checked to determine the right statistical test to applicate. If hypothesis of normality was approved, a Student's t-test was used, and if not, a Wilcoxon test. If the p-value was less than 0.05, the difference was significant.
Results
Compared to non-irradiated placebo condition, UV irradiation induced squalene oxidation by 2-fold, a decrease of catalase activity by 1.5-fold and a photo-isomerization of UCA by 5-fold.
Compared to the irradiated placebo, the active complex and sun filters alone protected squalene oxidation by 58.4% and 50.6%, catalase activity by 60.5% and 53.7%, and UCA photo-isomerization by 14.2% and 30%, respectively.
Combination of ectoine and mannitol with sun filters provided the better protection of squalene oxidation (76.8%; p<0.01), catalase activity (84.4%; p<0.001) and trans-UCA (53.9%; p<0.01).
Compared to filters alone, the active complex added to filters provided significant additional protection of squalene by 26% (p<0.05), catalase activity by 31% (p<0.05) and trans-UCA synergistically by 24% (p<0.01).
Discussion
To robustly study the biological damage induced by sun exposure, it seems essential to analyse biomarkers directly collected from volunteers. In this clinical study, we were interested in three biomarkers corresponding to three natural defence systems altered by UV: squalene for surface antioxidant, catalase which is an essential endogenous antioxidant enzyme, and photo-isomerization of UCA involved in preventing immunosuppression.
We showed that the combination of sun filters SPF30 with this active complex of 0.1% ectoine and 0.1% mannitol attributed a high protection capacity against squalene oxidation, decrease in catalase activity and decrease of trans-UCA induced by UV exposure.
Conclusion
This in vivo study demonstrates the interest of combining UV filters with a biological protection provided by specific active ingredients to allow optimal photoprotection and so to prevent UV damages induced by sun exposure such as induction of oxidation, decrease of endogenous antioxidant defence systems and induction of photo-immunosuppression.