INTRODUCTION:

Octadecyldimethylamine oxide (C₁₈DAO) is a type of quaternary ammonium compound widely utilized as a surfactant and emulsifier in both personal care formulations and industrial applications. Its molecular structure consists of a long hydrophobic octadecyl tail attached to a polar nitrogen-oxide moiety. This combination of hydrophobic and hydrophilic regions gives the molecule amphiphilic properties, allowing it to organize into stable assemblies at interfaces and in solution^1-5.

These assemblies exhibit unique physicochemical behaviors, including film formation with vertical alignment and diphilic interactions, making C₁₈DAO a candidate for investigating surface-driven biological effects.

Hair loss, or alopecia, is a widespread dermatological condition that significantly affects individuals' quality of life. Androgenetic alopecia, in particular, involves progressive follicular miniaturization. Current treatments such as topical minoxidil and oral finasteride aim to delay this process by improving vascularization and modulating hormonal balance^6,7. However, these agents may cause adverse effects, including dizziness, edema, and scalp irritation, and prolonged use often results in tolerance requiring dose escalation^8–10.

Recent studies have highlighted a growing interest in alternative and complementary approaches to treating alopecia. Various herbal formulations, polyherbal hair oils, and plant-derived extracts have demonstrated potential in stimulating hair growth, improving follicle health, and mitigating the psychological effects of hair loss in both preclinical and clinical settings^11-14. These data suggest that bioactive amphiphilic compounds such as C₁₈DAO may similarly support hair follicle regeneration by modulating the local microenvironment and enhancing dermal papilla cell activity, which is a promising direction for topical hair growth treatment.

While minoxidil and aminexil share structural and pharmacodynamic features that promote vascular and follicular support, their limitations highlight the need for safer alternatives. Notably, C₁₈DAO shares amphiphilic characteristics with fatty acids like omega-3 and -6, which are known to support skin and hair regeneration^15,16. Despite its use in formulation technology, the regenerative potential of C₁₈DAO on hair follicles has not been systematically evaluated.

In our previous studies, we investigated the molecular organization of C₁₈DAO films using atomic force microscopy, infrared spectroscopy, and X-ray diffraction. These investigations revealed that the structural behavior of C₁₈DAO is highly dependent on substrate and pH conditions, demonstrating significant conformational order and water retention around the head-groups – features that are likely to influence the local microenvironment of cells^17,18.

Based on these findings and considering the compound’s low cytotoxicity, amphiphilic nature, and ability to form stable films, we hypothesized that C₁₈DAO might modulate the microenvironment of dermal papilla cells derived from human scalp follicles (HHDPCs), thereby promoting their proliferation and stimulating hair follicle activity.

This study aimed to investigate the potential of C₁₈DAO to promote hair growth by examining its impact on the proliferation of dermal papilla cells derived from human hair follicles, the stimulation of critical intracellular signaling pathways, and the elongation of hair shafts in both laboratory-based cell culture models and animal experiments.

MATERIALS AND METHODS:

Octadecyldimethylamine oxide was procured from Wuxi Weiheng Chemical Co., Ltd (Jiangsu, China). The substance was purified following standard protocols¹⁹^,20 and dissolved in ethanol (DosFarm, LLC, Kazakhstan).

Scalp tissue samples were collected from healthy male donors with an average age of 20 years, none of whom had taken any medication for at least one month prior to participation. Sampling was performed during planned cosmetic procedures. The hair follicles were separated under a stereomicroscope at the Food and Environmental Safety Laboratory of the Kazakhstan–Japan Innovation Center (equipment: JEOL JEM-1011 with Morada digital camera, Leica DM 4000B, Leica EM UC7 RT, OLYMPUS, Japan). Follicle-derived dermal papilla cells (HHDPCs) from the occipital scalp region were cultured in specialized dermal papilla cell growth medium (LabTech, LLC, Kazakhstan) supplemented with 100U/ml penicillin and 100μg/ml streptomycin (Capricorn Scientific, GmbH, Germany), and maintained at 37°C in a humidified incubator containing 5% CO₂^21,22.

The study protocol was formally reviewed and approved by the Institutional Bioethics Committee at Kazakh National Medical University (Approval No. 1/2024, issued on 30 January 2024). Prior to participation, all volunteers were thoroughly informed about the study objectives, procedures, potential risks, and benefits, and provided their written consent to take part in the research. The study was conducted in full compliance with ethical standards and guidelines for human research.

To assess the impact of C₁₈DAO on cell viability and proliferation, the MTT assay (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) was utilized. Human hair dermal papilla cells (HHDPCs) were plated in 24-well plates at a density of 5 × 10⁴ cells per well and maintained in serum-free medium for 72hours. Following this, the cells were exposed for 24 hours to either a control solution or various concentrations of C₁₈DAO (0.1–5μM, 1:1000 dilution in culture medium). After the treatment period, 20μl of MTT solution (5 mg/ml) was added to each well and incubated for 3 hours at 37°C. The supernatant was then removed, and the formazan crystals formed were solubilized in 300μl of dimethyl sulfoxide (DMSO) with an additional incubation of 1hour. Absorbance was measured at 540 nm using a microplate spectrophotometer to quantify cell viability²³.

To assess the impact on hair follicle growth, isolated follicles were cultured and sectioned at the sebaceous duct level. C₁₈DAO was added to the medium, and hair shaft elongation was measured at multiple time points. Immunohistochemical and immunofluorescent staining were performed using antibodies against FGF-7 and FGF-10 (Sino Biological, Inc., China).

Quantitative analysis of mRNA expression was conducted using real-time PCR systems (Applied Biosystems Step One Plus, QuantStudio 12K Flex, Ion S5 System with Ion Chef, USA). Protein expression was evaluated by SDS-PAGE using 10% sodium dodecyl sulfate polyacrylamide gels, followed by chemiluminescent detection of antigen-antibody complexes (Shanghai Kehua Bio-Engineering Co., Ltd., China).

Female C57BL/6 mice were sourced from the KazNMU animal facility (Kazakhstan). All animal experiments were conducted in compliance with ethical guidelines and were approved by the Research Ethics Committee on Animal Studies at Kazakh National Medical University (Protocol No. 1/2024, dated January 30, 2024). Prior to experimentation, the animals received a thorough health evaluation following the institution's established operational procedures²⁴. During the study, a 2% C₁₈DAO formulation was applied topically to the shaved dorsal skin of each mouse once daily for a period of four weeks. Hair growth progression was documented through regular visual inspection and photographic recording.

All data are expressed as the average values with their corresponding standard deviations. Statistical analyses were performed using one-way ANOVA, followed by unpaired t-tests for comparisons between groups. Differences were considered statistically significant when the p-value was less than 0.05.

RESULTS:

«Figure 1» presents «the» molecular structure of C₁₈DAO. In our earlier studies, we extensively characterized thin solid films of this compound²⁵. When these films were deposited onto gold-coated glass substrates, the small-angle X-ray diffraction patterns displayed two distinct peaks at approximately 2θ = 3.1° and 6.3°, which correspond to the first- and second-order reflections of the crystalline lattice. From these data, the layer thickness was calculated to be approximately 2.8nm, in agreement with theoretical predictions. Another notable observation from our prior work was that single-monolayer films displayed complete hydrophobicity, as indicated by a contact angle measurement of 112°. Upon deposition of a second layer, the contact angle decreased markedly, suggesting that in multilayer arrangements the polar head groups are oriented toward the air interface.

Figure 1. Chemical structure of octadecyldimethylamine oxide (C₁₈DAO)

Another key observation was that the alkyl chains in C₁₈DAO films exhibited a high degree of order, oriented almost perpendicularly to the substrate surface. Deposition of additional layers further enhanced both molecular orientation and structural organization. The interaction between the nitrogen-oxide moiety and the substrate was found to play a critical role in defining the alignment and ordering of the hydrocarbon chains. In multilayer films, oleophilic interactions among the alkyl tails reorganized the molecular assemblies, as illustrated in Figure 2. Our analysis further indicated that the C₁₈DAO polar headgroups consistently reside at the interface, with the nitrogen–oxide moiety oriented almost parallel to the underlying surface. The hydrocarbon tails exhibited a largely ordered arrangement, with only one or two gauche conformations observed near the headgroup region, and were slightly inclined relative to the surface normal.

Figure 2. Diagram illustrating the molecular arrangement within the C₁₈DAO films

The vector connecting the two CH₃ groups in the C₁₈DAO molecule is oriented almost perpendicular to the substrate surface. Thermal analysis revealed that single-monolayer films undergo a distinct order-disorder transition at approximately 70°C, whereas films composed of five monolayers exhibit this transition near 45°C. In both cases, these values remain well below normal scalp and body temperatures. It is likely that the strong electrostatic interaction between the pronounced dipole moment of the nitrogen–oxide moiety and the negatively charged substrate surface enhances the trans-zigzag conformation and stability of the single-layer arrangement. Notably, the 45°C transition temperature of the multilayer film closely aligns with the 49 °C phase transition between the transparent and birefringent aqueous phases of C₁₈DAO.

In the current study, an MTT assay was employed to assess the influence of the nitrogen-oxide headgroup on the viability of human follicle dermal papilla cells (HHDPCs). Treatment with C₁₈DAO films significantly improved HHDPC survival; however, as shown in Figure 3, concentrations exceeding 1μM led to a decline in viability. To extend these observations, hair shaft elongation was evaluated in organ-cultured human follicles exposed to C₁₈DAO for two weeks. As depicted in Figure 4, concentrations of 0.1, 0.5, and 1μM produced marked increases in hair shaft length on days 7, 10, and 14 compared with the vehicle control. To further investigate follicular proliferation, Ki-67 immunofluorescence was performed after one week of treatment. The results (Figure 5) demonstrated a substantial rise in the number of Ki-67–positive keratinocytes, indicating enhanced proliferative activity. Moreover, quantitative PCR analysis showed that even a single day of C₁₈DAO exposure elicited a dose-dependent upregulation of FGF-7 and FGF-10 mRNA expression – both recognized as pivotal mediators of hair follicle growth and regeneration^26,27.

Figure 3. MTT assay measurement of HHDPCs C₁₈DAO treatment

Figure 4. Hair shaft elongation in ex vivo follicle culture (C₁₈DAO 0.1, 0.5, 1.0, 2.0 μM)

Control 0.1µM

0.5 µM 1µM

Figure 5. The proliferation of matrix keratinocytes in hair follicles treated with C₁₈DAO

Figure 6 presents immunohistochemical analysis of FGF-7 and FGF-10 expression following a three-day treatment of hair follicles with C₁₈DAO. Notably, exposure to concentrations of 0.1, 0.5, and 1μM resulted in elevated expression of both growth factors within the cytoplasmic region of the dermal papilla (DP).

A B

Figure 6. The mRNA relative expression for fibroblast growth factors FGF-7 (A) and FGF-10 (B)

This investigation utilized Western blot analysis to assess expression patterns of critical signaling proteins regulating cellular proliferation and survival²⁸, specifically examining ERK, AKT, CREB, and the anti-apoptotic factor Bcl-2. Treatment of HHDPCs with C₁₈DAO significantly enhanced phosphorylation of ERK, AKT, and CREB while upregulating Bcl-2 expression. As demonstrated in Figure 7, these findings suggest that C₁₈DAO exerts its effects through dual mechanisms: (1) activating the ERK pathway to inhibit apoptosis, and (2) enhancing AKT signaling and Bcl-2 production to support cellular survival.

Figure 7. C₁₈DAO treatment enhanced phosphorylation of ERK, CREB, AKT and Bcl-2 expression, relative to control

To investigate C₁₈DAO's effects on hair growth, we administered either C₁₈DAO or vehicle control to experimental mice. The study utilized 8-week-old female C57BL/6 mice, as this strain naturally transitions to the telogen (resting) phase by week 7 post-birth. This biological characteristic provides an optimal window for evaluating anagen phase induction without confounding effects from endogenous hair cycle activity²⁹. The study found that applicable for the treatment of alopecia areata quaternary amine oxide having antimycotic and antibacterial activity significantly boosted hair growth by day 28. After a four-week treatment period, mice treated with C₁₈DAO exhibited more pronounced dorsal skin darkening and a greater extent of hair regrowth compared to controls, indicating its efficacy in promoting anagen phase induction.

Histologic analysis of skin slices showed the ability of C₁₈DAO to affect the area occupied by follicles relative to the dermis, the average thickness of hair follicles. The back skin of the mice was synchronized to the anagen phase through depilation³⁰. On the 21st day of the experiment, which corresponded to 10 days after initiating treatment, histological examination (Figure 8) revealed notable morphological changes in the hair follicles exposed to C₁₈DAO. These follicles exhibited greater size and extended depth compared to the control group. Quantitative scoring of hair cycle stages confirmed a marked prolongation of the anagen phase. Notably, no significant differences in hair shaft thickness were observed between the groups.

Figure 8. C₁₈DAO enhanced anagen induction weekly representation

DISCUSSION:

This study investigated the effects of octadecyldimethylamine oxide (C₁₈DAO) on hair growth through a combination of in vitro, ex vivo, and in vivo approaches. The findings demonstrated that C₁₈DAO significantly stimulated the proliferation of scalp-derived dermal papilla cells (HHDPCs), promoted elongation of hair shafts, and facilitated anagen phase initiation in a mouse model³¹^,³².

The biological effects appear to be partially mediated through upregulation of FGF-7 and FGF-10, known key regulators of folliculogenesis and hair regeneration. These paracrine factors promote keratinocyte proliferation and help maintain the anagen phase, as demonstrated by immunohistochemical analysis showing significantly enhanced FGF-7/10 expression in C₁₈DAO-treated follicles. These findings align with established literature highlighting the crucial involvement of FGF signaling in both hair follicle development and regeneration^33,³⁴.

The results of our Western blot experiments provide clear evidence that treatment with C₁₈DAO modulates several key intracellular signaling pathways within human hair dermal papilla cells. Specifically, we observed a marked increase in the phosphorylation levels of ERK, AKT, and CREB, accompanied by upregulation of the anti-apoptotic protein Bcl-2. These molecular alterations collectively suggest that C₁₈DAO promotes a cellular environment conducive to enhanced survival and growth³⁵.

Activation of the ERK and AKT pathways is particularly significant, as these signaling cascades are well-known regulators of cell proliferation and survival. Enhanced ERK phosphorylation likely contributes to improved cellular viability by transmitting pro-survival signals, while AKT activation may support the proliferative potential of dermal papilla cells, thereby promoting the expansion of this critical follicular cell population. Simultaneously, the observed elevation in Bcl-2 levels indicates a reinforced resistance to programmed cell death, suggesting that C₁₈DAO not only stimulates growth but also fortifies the cells against apoptotic stress³⁶.

Taken together, these findings imply that C₁₈DAO exerts a multifactorial protective and stimulatory effect on hair follicle dermal papilla cells. By simultaneously enhancing survival pathways, promoting proliferation, and reducing apoptotic susceptibility, the compound may create a favorable microenvironment within the follicular niche that supports hair growth. This comprehensive modulation of intracellular signaling underscores the potential of C₁₈DAO as a bioactive agent capable of influencing hair follicle physiology at multiple regulatory levels³⁷.

The observed bioactivity of C₁₈DAO may be attributed to its amphiphilic molecular structure. The polar nitrogen–oxide headgroup is oriented oriented roughly along the plane of the surface, allowing close interaction with cell membranes, while the hydrophobic tail maintains perpendicular orientation, potentially stabilizing its conformation in the extracellular environment. This configuration may enhance its integration into lipid bilayers and facilitate signal transduction.

Animal studies using C57BL/6 mice further corroborated the hair-promoting properties of C₁₈DAO. Topical application induced visible pigmentation and accelerated hair regrowth, indicative of anagen reentry. The observed data indicate that C₁₈DAO can influence the dynamics of hair follicle cycling, offering a potentially valuable approach for managing alopecia. However, the current findings should be interpreted with some caution due to several limitations. For instance, the long-term safety and potential for systemic absorption of C₁₈DAO remain unclear and warrant further investigation. Additionally, the exact molecular targets through which C₁₈DAO exerts its effects have yet to be fully characterized. Although this study utilized samples from young adult donors and a murine model, it remains to be determined whether similar outcomes would be observed in older individuals or in various forms of alopecia.

Despite these limitations, the results present compelling evidence for the role of C₁₈DAO in stimulating hair regeneration. These findings may pave the way for the development of C₁₈DAO-based interventions in both cosmetic and therapeutic applications aimed at treating hair loss.

Future perspectives and safety considerations:

Further research is necessary to establish the chronic safety of C, including assessments of dermal irritation, allergenic potential, and systemic absorption in both preclinical and clinical contexts. Long-term toxicology studies using larger animal models, as well as toxicokinetic profiling, would help determine any cumulative or delayed effects. Considering its membrane-interacting properties, future work should also examine possible impacts on epidermal barrier function and skin lipid composition. Clinical trials in diverse patient populations – particularly those with androgenetic alopecia or other persistent hair loss disorders – are essential to confirm therapeutic efficacy, optimize dosing regimens, and ensure tolerability.

Collectively, the present findings suggest that C₁₈DAO possesses a unique combination of physicochemical and biological properties that enable it to influence hair follicle regeneration through multiple pathways. By integrating cell survival mechanisms with follicular growth stimulation, this compound holds promise as a novel candidate for alopecia management, warranting further investigation toward safe and effective clinical application.

CONCLUSIONS:

Our results indicate three key outcomes from the topical application of C₁₈DAO: (1) increased proliferation of cultured dermal papilla cells derived from human hair follicles, (2) greater extension of hair shafts observed in ex vivo experiments, and (3) stimulation of anagen phase onset in mouse hair cycle studies. These observable effects were associated with molecular changes, notably elevated levels of follicular growth factors (FGF-7, FGF-10) and activation of signaling pathways involving ERK, AKT, and Bcl-2.

The observed biological effects may be attributable to C₁₈DAO's unique amphiphilic properties, which enable efficient cellular membrane interactions and subsequent activation of pro-regenerative signaling cascades. This dual capacity to modulate both cellular growth and follicular cycling positions C₁₈DAO as a potentially valuable bioactive compound for developing innovative alopecia therapies.

However, further investigations are necessary to evaluate its long-term safety, elucidate its mechanisms of action in vivo, and confirm its therapeutic potential in clinical applications.

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

ACKNOWLEDGMENTS:

The authors gratefully acknowledge the technical assistance provided by the staff of the Food and Environmental Safety Laboratory at the Kazakhstan-Japan Innovation Center. We also thank the volunteers who participated in the study and the animal facility personnel for their support during in vivo experiments.

INSTITUTIONAL REVIEW BOARD STATEMENT:

The study involving human biological materials was approved by the Institutional Bioethics Committee of Kazakh National Medical University (Approval No. 1/2024, dated 30 January 2024). Animal experiments were approved by the Committee on Animals for Research and Ethics, Science Department, Kazakh National Medical University (Approval No. 1/2024, dated 30 January 2024).

INFORMED CONSENT STATEMENT:

Written informed consent was obtained from all human participants prior to the collection of scalp tissue samples.

DATA AVAILABILITY STATEMENT:

The data supporting the findings of this study are available from the corresponding author upon reasonable request. Data are not publicly available due to participant privacy and ethical considerations.

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Received on 15.08.2025 Revised on 22.11.2025

Accepted on 07.02.2026 Published on 01.07.2026

Available online from July 04, 2026

Research J. Pharmacy and Technology. 2026;19(7):2961-2967.

DOI: 10.52711/0974-360X.2026.00422

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Creative Commons License.