A Selenium Disulfide-Based Shampoo Is Beneficial for Dandruff Management and Rebalancing the Scalp Microbiome of Subjects of Any Hair Type Free

Dandruff is a mild form of seborrheic dermatitis (SD) of the scalp [1, 2]. Its prevalence has been estimated at up to 50% in the general population [3]. It is observed in all phototypes and hair types. Dandruff in different ethnicities has been reported to present at higher prevalence in women of African American descent (81–95%) than in other ethnic groups, such as Caucasian and Asian descent, with 66–82% and 30–42% prevalence, respectively [4, 5].

Malassezia (M) sp., especially Malassezia restricta, and the host-inhabitant interplay have been suggested to trigger dandruff [6‒8]. M. restricta induces keratinocyte cytotoxicity, resulting in accelerated scale formation [9, 10]. M. restricta, as is the case for any other Malassezia yeast, metabolizes sebum-derived lipids such as triglycerides into free fatty acids, oxidizes squalene, and produces indole derivatives, such as malassezin and indolocarbazole, leading to pro-inflammatory compounds [11, 12]. Moreover, bacterial community changes are associated with the pathogenesis of dandruff/SD [13‒15]. A higher diversity and imbalance between the two dominant bacteria, consisting of a reverse correlation between the abundance of Staphylococcus sp. (higher) and Cutibacterium acnes (lower), has been shown in dandruff/SD [16‒20].

Existing scalp SD treatments consist of topical applications of antifungals and anti-inflammatory agents. Selenium disulfide (SeS₂) 1% shampoo is effective in dandruff, reducing scales and itching, while rebalancing the scalp microbiome [21‒25]. It improves clinical symptoms of scalp SD after treatment with ketoconazole [26]. However, its efficacy in subjects of different hair types remains to be established. Fajuyigbe et al. [27] reported that one single hair wash with this product immediately alleviates dandruff and scalp itchiness in this population. However, the effect was not maintained after 1 week, supporting the need for a specific anti-dandruff regimen. This is the first study that assesses the clinical benefit of SeS₂ shampoo in moderate-to-severe dandruff and its effect on the scalp microbiome in subjects of phototype or different levels of curly hair type.

This open-label observational, noninvasive, and prospective 28-day study, followed by a 14-day remanence period for subjects with hair type IV to V and hair type VI–VIII, was conducted at one investigational site in France. Due to the cosmetic status of the tested product, this investigation did not require ethics committee approval. The study complied with local legal requirements for the conduct of this type of study, with the principles of the Declaration of Helsinki and Good Clinical Practices. Moreover, all subjects provided written informed consent prior to inclusion.

At least 90 subjects aged 18 years and above, of any phototype according to the Fitzpatrick scale, and of any hair type (I = not curly to VIII = tightly coiled) according to De La Mettrie et al. [28], with moderate-to-severe dandruff defined as the total dandruff score ≥4 (on a scale from 0 = none to 10 = severe) and adherent and non-adherent dandruff score ≥2.5 (both ranging from 0 = none to 5 = severe), were suitable for this study [28‒30]. The hair length of all subjects was to be at least 2 cm. Included subjects were asked to apply SeS₂ shampoo twice weekly, as indicated on the product leaflet, for 28 days, and used a neutral shampoo for a further 14 days.

The investigator assessed the total, adherent and non-adherent flake scores, signs and symptoms of SD (SSSD) at baseline, days 7, 14, and 28, and, in subjects with hair type IV to V and hair type VI–VIII, 14 days after the last SeS₂ shampoo use (day 42), using the SSSD scale (from 0 = none to 15 = severe and composed by the erythema, scales, and pruritus scores, all from 0 = none to 5 = severe), as well as scalp greasiness on a scale from 0 = none to 5 = very severe and quality of life (QoL) using the Scalpdex scale from 0 = good QoL to 100 = bad QoL [31]. Subjects assessed maximum itching and scalp greasiness (both assessed on a scale from 0 = not at all to 9 = hugely) as well as the perceived benefit, acceptability, and cosmetic characteristics. Local tolerance was assessed throughout the entire study. At day 28 and day 42, the investigator assessed the global efficacy on a scale from 1 = worse to 5 = complete reduction or complete remission, as well as satisfaction from 0 = not satisfied to 2 very satisfied.

Microbiome samplings and analysis was performed as reported by Massiot et al. [26] 2022 by swabbing a lesional area of the scalp of 2 × 2 cm² at day 0 before the first treatment and day 28 after 4 weeks of treatment. To ensure sampling at the same scalp location, the hair at the sampling area was cut to a length of 5 mm. A sterile cotton swab was dampened in a 0.15 m NaCl and 0.1% Tween 20 solution and rubbed onto the scalp surface with a zig-zag pattern, in a nonoverlapping manner. At the end of the procedure, the head of the swab was cutoff, placed in an Eppendorf sterile tube and stored at −80°C until DNA extraction and qPCR or NGS analysis. In parallel, sterile cotton swabs were kept as negative controls.

For microbiome sequencing, 16S and ITS amplicon libraries were obtained using specific bacterial 16S rRNA (V1-27S and V3-535R) and specific fungal ITS1 (ITS-18SF and ITS-5.8S1R) hypervariable regions. Libraries were sequenced on a NovaSeq^® System (Illumina, San Diego, USA) for 300 bp paired-end at the Genomics Center, CHU de Québec-Université Laval Research Center, Canada. Sequence processing and bioinformatics are described in online supplementary material (for all online suppl. material, see https://doi.org/10.1159/000544742) and Methods section. qPCR was performed on all the samples using primers and TaqMan MGB probes, as previously reported [32].

Qualitative clinical variables were described as numbers and percentages. 95% confidence intervals were calculated where appropriate. Quantitative variables were described as numbers, mean, standard deviation, medians, minimum and maximum values. All statistical analyses of the clinical data were performed at a 5% significance using 2-sided tests, except the normality which was tested at the threshold of 1% (Shapiro-Wilk test). All statistical analyses were performed using SAS version 9.4.

For raw data of QPCR (Staphylococcus sp., Cutibacterium sp., Malassezia sp.), a log 10 transformation of data was applied to obtain normally-distributed residuals for each QPCR variable. For sequencing data analysis, R software was used (vegan: Community Ecology Package. R package version 2.4–5). Pairwise analysis of similarities (ANOSIM) was applied to assess differences based on treatment and time point. ANOSIM global R value ranges from 1 to −1 (R ∼ 0 indicates the same level of variation within and between groups). ANOSIM testing was performed using α = 0.05 for statistical significance. Alpha diversity (Shannon) was analyzed following the same method as for clinical parameters (linear mixed model). The Benjamini-Hochberg procedure was used for multiple comparison adjustment.

A total of 93 subjects (women: 83%) were recruited. All hair types were represented: I–III: 41%, IV–V: 29%, VI–VIII: 30%. 69% of subjects were of European and 31% of African origins. Phototypes ranged from I (2.2%) to VI (16.1%) with most subjects of phototype III (55.9%). Table 1 provides detailed demographic and baseline dandruff scores for the overall study population and for the different hair type groups. No significant difference between hair type groups was observed.

The total dandruff score (Fig. 1) significantly (p < 0.0001) improved over time (day 7: −31.5%, day 14: −53.5%, and day 28: −76.2%, all p < 0.0001), as did the adherent dandruff score (day 7: −29.9%, day 14: −51.8%, and day 28: −76.4%; all p < 0.0001) and the non-adherent dandruff score (day 7: −33.0%, day 14: −55.1%, and day 28: −76.0%, all p < 0.0001). At day 42, the benefit of SeS₂ shampoo on the total, adherent, and non-adherent dandruff score was maintained, with differences from day 28 being statistically insignificant for any of the scores. According to the investigator, erythema significantly (p < 0.0001) improved in the overall study population after 7 (−16.6%, p < 0.0003), 14 (−39.2%, p < 0.0001), and 28 days (−53.9%, p < 0.0001). Dandruff improved or completely resolved in 96.8% (95% CI: 90.9; 99.3) of the subjects and remained improved or resolved in 89.1% (95% CI: 77.8; 95.9) of the subjects after 42 days. Greasiness (Fig. 2) significantly improved in the overall study at day 7 (−17.1%, p = 0.0033), day 14 (−26.8%, p < 0.0001), and day 28 (−37.6%, p < 0.0001). After 42 days, erythema and greasiness scores remained low with no significant change compared to day 28 (Fig. 2).

The SSSD score followed the same pattern as each single parameter, with a significant (p < 0.0001) decrease in the overall study population as early as after 7 days (−28.3%), and continued to decrease at day 14 (−49.6%) and day 28 (69.6%) (Fig. 3). At day 42, no significant change from day 28 was observed in the 55 subjects with higher curly hair.

The investigator was satisfied or very satisfied with the outcome in 97.8% (95% CI: 92.4; 99.7) of the subjects after 28 (95% CI: 92.4; 99.7) and after 42 days (95% CI: 77.8; 95.9). According to the subjects, maximum itching (Fig. 4) significantly (p < 0.0001) decreased with SeS₂ shampoo (day 7: −37.8%, day 14: −51.6%, and day 28: −71.7%). Between days 28 and 42, the intensity of maximum itching significantly (p < 0.048) increased by 9.6%. Differences to baseline for the self-assessed scale severity were significant (p < 0.0001) at day 7 (22.6%), day 14 (−49.1%), and day 28 (68.3%). Scalp greasiness significantly improved at day 7 (−13.0%, p = 0.0036), day 14 (−35.7%, p < 0.0001), and day 28 (−56.9%, p < 0.0001). No significant evolution of maximum itching or scalp greasiness was observed at day 42 compared to day 28.

QoL significantly (all p ≤ 0.0003) improved overall as early as day 7 (−12.1%), with an improvement of 43.9% at day 28. The level of improved QoL remained almost unchanged at day 42. A total of 91.4% of the subjects were satisfied or highly satisfied with SeS₂ shampoo; 97.0% stated that the condition had improved or resolved. Cosmetic characteristics of SeS₂ shampoo were highly appreciated by all subjects. Local tolerance was excellent, with no product-related adverse event being reported.

Dandruff decrease was slightly higher for group I–III than the two other groups (∆ ≈ −6% for all dandruff scores). At day 28, the decrease of total dandruff was −79.3% for hair type group I–III and −74.1 in hair groups IV–V and −74.2% for hair type VI–VIII (Fig. 1). At day 28, adherent dandruff had improved by 80.1% for hair type group I–III, by 73.9% for hair type group IV–V, and by 73.8% for hair type VI–VIII group. Improvements of the non-adherent dandruff score were −78.5% (hair type groups I–III), −73.8% for hair type groups IV–V, and −74.6% for hair type groups VI–VIII. All changes from baseline were statistically significant (all p < 0.0001). At day 42, no significant change from day 28 was observed for hair types ranging from IV–VIII for any type of dandruff, including total dandruff.

Erythema decreased by 49.2% (p = 0.0003), 49.9% (p < 0.0001), and 66.5% (p < 0.0001) in hair type groups I–II, IV–VI, and VI–VIII, respectively, at day 28, with no significant change at day 42 from day 28 (Fig. 2). Greasiness had significantly (p < 0.0001) improved in all hair type groups: I–III (−29.0%), hair type group IV–V (−55.3%), and hair type group VI–VIII (−33.8%) at day 28, with a somewhat higher improvement in group IV–V. The SSSD score had significantly (p < 0.0001) decreased in all 3 groups: by 70.5% in hair type group I–III, by 67.7% in hair type group IV–V, and by 70.1% in hair type group VI–VIII at day 28 (Fig. 3).

According to the subjects, maximum itching had improved by 75.7% in hair type group I–III, 75.4% in hair type group IV–V, and 62.7% in hair type group VI–VIII after 28 days. An insignificant increase of the maximum itching score at day 42 from day 28 was observed for groups IV–V and VI–VIII (Fig. 4). A similar trend for the self-assessed scale severity was observed (hair type group I–III: −69.7%, hair type group IV–V: −71.4%, and hair type group VI–VIII: −63.1%, all p < 0.0001). Scalp greasiness had significantly (p < 0.0001) improved at day 28 in hair type group I–III (−55.3%) and hair type group IV–V (−68.1%). The improvement of greasiness was insignificant in hair type group VI–VIII. According to the subjects, no significant change for any of the assessed parameters or any of hair types IV–VIII was observed between day 28 and day 42.

At baseline in the overall study population, the bacterial microbiome presented a typical scalp taxonomic profile dominated by Cutibacterium sp. and Staphylococcus sp., representing more than 80% of the relative abundance (Fig. 5a); with a high proportion of Staphylococcus as compared to a reported healthy scalp. SeS₂ shampoo improved the bacterial profile with a decrease in Staphylococcus sp. as confirmed by qPCR quantification (p < 1.0 10⁻⁵), while the Cutibacterium sp. load was not impacted (Fig. 5b). The bacterial diversity significantly decreased, as measured by the Shannon index. The analysis of the bacterial community structure using PCoA on the UniFrac distances (weighted and unweighted) showed minor modifications (R = 0.026, p = 0.001 and R = 0.033, p = 0.004 for the Staphylococcus sp. as well as Cutibacterium sp. load, respectively) suggesting that the modifications of the bacterial diversity were relatively weak. For the fungal species, SeS₂ shampoo induced a decrease in the Malassezia relative abundance (Fig. 6a), which was confirmed by qPCR quantification (p < 1.0 10⁻⁵; Fig. 6b). Consequently, the fungal diversity significantly increased, as measured by the alpha diversity (p < 1.0 10⁻⁵), with a highly modified fungal community (R = 0.34, p = 0.001 and R = 0.23, p = 0.001 for weighted and unweighted UniFrac distance, respectively).

Microbiome modifications among the different hair type subgroups showed that, at baseline, the microbiome analysis showed no difference between groups in terms of qPCR results (load of Malassezia sp., Cutibacterium sp., and Staphylococcus sp.) or bacterial and fungal alpha diversity. After 4 weeks of use of SeS₂ shampoo, the bacterial and fungal taxonomic profiles were modified in a similar pattern as for the global study population: a significant (p < 0.001) decrease in Staphylococcus and Malassezia in terms of relative abundance and qPCR quantifications, and an unchanged Cutibacterium sp. abundance was observed (Fig. 7a and b). In addition, fungal alpha diversity significantly (p < 0.001) increased. The increase in bacterial diversity was significant (p < 0.001) for hair type group I–III only, with a tendency to decrease for the other hair types. The PCoA on the beta diversity did not show any significant differences, confirming the minor effect of SeS₂ shampoo on bacterial diversity in each hair type group. The fungal alpha diversity and PCoA were strongly impacted in each group (Fig. 7c).

Although dandruff has been reported to present a higher prevalence in women of African American descent with curly hair, no data about the treatment of dandruff on a population with curly to highly coiled hair currently exist. Considering that in patients of African origin with curly hair, itching has been reported to be highly associated with dandruff, the present study aimed to assess the benefit of SeS₂ shampoo on dandruff, itching, and scalp microbiome in subjects with curly to coiled hair [27].

Results from the present study confirmed the clinical benefit of SeS₂ shampoo in the management of moderate-to-severe dandruff, which has already been demonstrated in previous clinical studies [21, 24‒26, 33]. In addition, for the first time, investigations provided evidence that the SeS₂ shampoo is beneficial for different hair types from curly to very coiled hair, to manage signs and symptoms of dandruff, as well as the scalp microbiome composition and its diversity.

After 28 days of use, SeS₂ shampoo significantly (p < 0.0001) reduced clinical signs and symptoms of dandruff, including the quantity and severity scores of adherent, non-adherent, and total dandruff, erythema, and scalp greasiness, as well as maximum itching in all hair type groups. Accordingly, the subjects’ QoL significantly (p ≤ 0.0001) improved as early as after 7 days, thereby confirming the impact of dandruff on the subjects’ mental wellbeing, and the benefit of SeS₂ shampoo in improving the subjects’ burden related to the presence of dandruff [34, 35]. Due to its highly suitable shampoo formulation, subjects greatly appreciated the product, even in the most curly/coiled hair types, thus favoring a high compliance of use, supporting maximum benefit in any hair type.

A main limitation of this study is certainly its design, which does not compare the benefit of SeS₂ shampoo in different hair types with that of other anti-dandruff products. However, a recent study, comparing SeS₂ and ketoconazole shampoo in subjects with moderate-to-severe seborrheic dermatitis of the scalp, demonstrated that both formulations provide a similar benefit, with SeS₂ shampoo improving the subjects QoL faster [33].

Regarding the scalp microbiome analysis, the study focused on the main genera of the scalp, including Staphylococcus sp., Cutibacterium sp., and Malassezia sp., using genus-specific qPCR and rDNAs amplicon methods. A limitation of microbiome analysis is certainly that analyzing the scalp microbiome specificities for each hair type using a more resolution microbiome sequencing method such as whole genome sequencing would have been of interest. This would help identify the microbial taxa at the level of the species and, in turn, fine-tune the scalp treatment according to the scalp-specific bacteria or fungi, or tackle further additional improvement for hair types IV–V and VI–VIII. However, we consider that this analysis requires further investigations.

In conclusion, this study confirms the benefits and safety of SeS₂ shampoo in the management of moderate-to-severe dandruff in subjects of any hair type. Moreover, the study provides evidence that SeS₂ shampoo rebalances the scalp microbiota regardless the hair type. SeS₂ shampoo was highly appreciated by the subjects for its benefit, acceptability, and cosmeticity, thereby allowing a potential improvement of compliance, including in the long term.

The authors acknowledge the study management support of Anna Veriato, employee of Vichy Laboratoires when the study was conducted, and the writing support of Karl Patrick Göritz, SMWS, France.

Ethical approval is not required for this study in accordance with local or national guidelines. All subjects provided written informed consent for participation prior to inclusion.

Claire Deloche-Bensmaine, Natalia Kovylkina, Julie Faure Cécile Clavaud, and Audrey Gueniche are employees of L’Oréal group. Stéphanie Leclerc-Mercier was an employee of Vichy Laboratoires, France, when the study was conducted. The other authors have no conflict of interest to disclose.

This study was funded by Vichy Laboratoires, France.

Claire Deloche-Bensmaine and Julie Faure conducted the study. Natalia Kovylkina is the sponsor representative. Stephanie Leclerc-Mercier was the sponsor representative and contributed to the design of the study. Cecile Clavaud and Audrey Gueniche supervised the microbiome analysis. Victoria Barbosa and Pascal Edouard Reygagne served as consultants for this study. All authors analyzed the data and participated in the preparation, reading, and approval of the manuscript.

The data that support the findings of this study are not publicly available due to their containing information that could compromise the privacy of research participants but are available from the corresponding author upon reasonable request.