Eyelash serums: A comprehensive review

Eyelash serums, both prescription and over‐the‐counter, are gaining popularity for enhancing the appearance of eyelashes through various biologically active molecules. Categorized into prostaglandin analogs and non‐prostaglandin analogs, these serums claim increased strength, length, luster, and thickness. Current United States law also requires no efficacy or safety assessments by the Food and Drug Administration before approving products for consumer use, potentially posing health risks for patients seeking over‐the‐counter eyelash enhancements.


| INTRODUC TI ON
Eyelash serums are prescription or over-the-counter (OTC) products that contain different biologically active molecules with varying claims to enhance the appearance of the eyelashes through parameters such as increased strength, length, luster, and thickness.
These serums are sought after by consumers for lengthening natural lashes.They may also lengthen lashes after disease or insult, such as alopecia areata or anagen effluvium following chemotherapy.The main biologically active molecules in eyelash serums can be classified into two categories: (1) prostaglandin analogs and (2) nonprostaglandin analogs.The bioactive ingredient prostaglandin, formulated as bimatoprost, is the only Food and Drug Administration (FDA) approved molecule for eyelash growth. 1 Non-prostaglandin analog molecules include growth factors, peptides, pumpkin seed oil, and ginseng, among others.
Eyelash serums are part of a growing industry with a large consumer base.The cost of prescription eyelash serum ranges from $26 to over $150 US dollars when using discount cards depending on generic, brand, or pharmacy used.In contrast, the cost of OTC eyelash serum varies widely, ranging from under $10 to over $100 US dollars.Analyst projections indicate that the eyelash serum market size will reach a value of $1.3 billion by 2031, up from $752 million in 2020. 2 These serums vary by their mechanism of action, effectiveness, and side effects.This comprehensive review thoroughly discusses the eyelash cycle, prostaglandin, and non-prostaglandin analogs as well as their mechanisms of action, efficacy, benefits, and side effect profiles.A complete understanding of available options will help dermatologists recommend the most effective patientspecific eyelash serums and allow patients to avoid unnecessary and damaging complications.

| The eyelash cycle
The eyelash cycle is characterized by three stages: anagen (growth), catagen (transition), and telogen (rest) (Figure 1).The cycle can vary in duration, but typically takes 4-11 months to complete.Eyelash growth occurs during the anagen phase, which lasts approximately 4-10 weeks.During the anagen phase, the hair follicle is attached to the blood supply and obtains nutrients for growth.The average growth of an eyelash can vary from 0.12 to 0.14 mm per day.The lash length rarely exceeds 12 mm.The catagen phase duration is about 15 days and disrupts the blood supply to the hair follicle.
The telogen phase lasts about 4-9 months and sheds the hair to start the cycle again with a new hair. 3Thibaut et al. conducted a study exploring eyelash growth from 29 female volunteers over 9 months and found that the complete eyelash cycle lasted about 90 ± 5 days, with the anagen phase comprising 34 ± 9 days. 4The average growth rate of an eyelash was 0.12 ± 0.05 mm daily.Lash length varied from 2.1 to 6.6 mm, with an average of 4 ± 1 mm.The cycle also varied in duration and across individual hair follicles on the same person. 4

| Biology of the human eyelash
The upper lid contains 90-160 lashes spread between 5 and 6 rows, and the lower lid contains 75-80 lashes dispersed between 3 and 4 rows. 3Compared to the scalp skin which contains the epidermis, dermis, and hypodermis, the eyelid skin is composed of a thinner epidermis and dermis in which the eyelash follicle is rooted.Thus, the eyelash follicle is shorter than the scalp hair follicle.Additionally, contrary to the scalp hair follicle, lash follicles do not have arrector pili muscles and thus don't require individual mobility. 3

| Factors that impact eyelash appearance
Ethnicity, pigmentation, and age affect the appearance and variability of eyelashes. 4In all ethnicities, there is a curved morphology of eyelashes in which the curvature is initiated at the bulb of the lash and continues to the tip of the shaft.While studies are limited on racial differences in eyelashes, studies have identified the degree of lash curvature can vary between ethnicities, with Caucasians having more pronounced curl as compared to Asians. 4 An observational study by Na et al was conducted to investigate the specific differences in characteristics of eyelashes between 20 Asian women and 10 Caucasian women.A comprehensive series of measurements were done with a phototrichogram technique to study eyelashes.The study spanned over 2 weeks, where participants were instructed to refrain from using makeup products around the eye.The results revealed that Asian eyelashes had lower lift-up and curl-up angles, were fewer in quantity, and possessed greater thickness.Results also demonstrated that Asian eyelashes had a higher number of cuticle layers and increased density when examined in cross section.It is important to note that there were no notable distinctions in eyelash length or growth rate between these two groups, with the anagen phase approximated F I G U R E 1 Normal eyelash growth cycle.
to last 2 months for both ethnicities.There insignificant difference in length between Asian and Caucasian eyelashes was proposed to be due to the anatomy of the Asian upper eyelid, where the periorbital fat compartment more commonly tends to descend further into the lid, which can hide the eyelash root. 5ditionally, the degree of pigmentation in eyelashes is defined by the number of melanocytes in the lash follicle structure, with the expression of a tyrosine-related protein (TRP-2) influencing melanogenesis. 4 In the eyelash, TRP-2 has sustained expression, which explains why the eyelash may become gray but rarely white at an advanced age compared to scalp hair.Furthermore, Glaser et al. found that increasing age is associated with a reduction in the length, thickness, and pigmentation of the eyelash. 6While there has not been adequate research conducted on the proposed pathogenesis of age-related changes in eyelashes, an increase in age has been studied to be associated with hair loss in general due to a decline in regenerative abilities, explained by telomerase loss, DNA damage, and a buildup of free radicals.This can impact one's quality of life and mental health. 7erall, these factors may play a role in consumer desirability of eyelash serums.

| MATERIAL S AND ME THODS
We performed a comprehensive electronic search for "eyelash serum" in PubMed, Embase, Cochrane Central, and Google Scholar.Two independent researchers evaluated articles for their relevance to this topic.Suitability of the articles was determined by the two independent reviewers by assessing if the article involved eyelash serums or hair growth of any kind.
Nonhuman studies were included for comprehensiveness.
Preclinical trials, clinical trials, meta-analyses, randomized controlled trials, retrospective studies, case reports, comparative studies, reviews, and systemic reviews were included in the search.Results not in English were excluded within this search.
The ingredients discussed were also analyzed and given clini-

| PROS TAG L ANDIN ANALOG S
Prostaglandins are secreted as local hormones by most organs and tissues in the body, except for erythrocytes in the blood. 9These hormones act locally on the tissues near their production site. 9They are involved in several physiological and pathological processes, including inflammatory processes, and have been implicated in several dermatological disorders, such as allergic contact dermatitis.
Prostaglandin analogs in the treatment of ophthalmic disease are utilized for conditions such as glaucoma and ocular hypertension. 10ey were later found to cause increased hair growth at their site of action, which led to research exploring their potential benefit for conditions of hypotrichosis, such as alopecia areata, androgenic alopecia, frontal fibrosing alopecia, and hypotrichosis of the eyebrows and eyelashes. 11,12The ability of prostaglandin analogs to improve cosmetic parameters of the eyelashes has led to these agents being popular in the world of cosmetic dermatology.
The mechanism of action of prostaglandin analogs on eyelash growth involves prolonging the anagen phase and shortening the time between telogen and restart of anagen phase 13 (See Table 1).This maximizes the time the hair follicles spend in the anagen growth phase, leading to longer, thicker, and darker eyelashes. 13In this section, prostaglandin-based eyelash serums are reviewed (See Table 2).
The ingredients discussed were also analyzed and given clinical recommendations for eyelash serums based off the Oxford Centre for Evidence-Based Medicine (See Table 1).

| Bimatoprost
Topical bimatoprost, initially approved for treating ocular hypertension and open-angle glaucoma, is a synthetic prostamide F2α (PF2α) analog. 11Bimatoprost-induced eyelash hypertrichosis has been reported as a side effect in patients undergoing treatment of ophthalmological diseases such as glaucoma.Bimatoprost ophthalmic solution is available in 0.01% and 0.03% formulations.Bimatoprost 0.03% solution is the only FDA-approved prostaglandin analog used for treating hypotrichosis of the eyelashes. 14,15It has demonstrated efficacy in improving several parameters of the eyelashes, including length, thickness, and pigmentation, and it was FDA-approved in 2008 for its cosmetic benefits (Figure 2).Appropriate usage includes cleaning the face, removing contact lenses, and applying one drop nightly along the upper eyelid margin at the base of the eyelashes with a sterile single-use per-eye applicator.It should not be used to the lower eyelash line. 11ere are various proposed mechanisms behind the efficacy of topical bimatoprost, including stimulation of the prostamide receptor, transitioning follicles from the resting telogen phase to the growing anagen phase, prolongation of the anagen phase, and enlargement of the dermal papilla and hair bulb, which creates thicker hair. 16In addition, bimatoprost stimulates melanogenesis which increases the pigmentation of eyelashes, resulting in darker-colored lashes. 16Sayed et al. explored the changes in the eyelashes of 15 rabbits after treatment with topical bimatoprost 0.03% daily to the right conjunctival fornix for 4 weeks, using the left eye as the control.The authors found that in addition to increased length, thickness, and pigmentation, there was an increased number of eyelashes within the same hair follicle and increased tensile strength of the hair. 17To our knowledge, these particular findings have not yet been described in humans.
Iris pigmentation is another side effect commonly counseled to patients with use of prostaglandin-based serums, especially in users with light colored eyes.The exact mechanism of iris pigmentation due to prostaglandin use is unknown, however, appears to mainly be due to a stimulation of melanogenesis in melanocytes of the iris. 26odward et al conducted a randomized controlled trial (RCT) on cosmetic eyelash growth on the use of bimatoprost 0.03% gel once daily for 6 months, where no patients reported any subjective change in iris color.It is important to note, however, that scientific classification of iris color was not performed in this study. 21While TA B L E 1 Prostaglandin analogs.*Statistically significant with p < 0.05.TABLE 2 (Continued)  the association between iris pigmentation with cosmetic lash serum does not appear to be extensively studied, the association of iris pigmentation has been studied with the use of prostaglandins for glaucoma treatment.A study by Lipp et al discusses that while the use of bimatoprost applied directly to the eye for the treatment of glaucoma is associated with iridial pigmentation, present literature does not appear to analyze the significance of risk when the solution is applied exclusively to the eyelashes.The authors also note that the hyperpigmentation on the iris is typically permanent. 27Therefore, it is important for consumers to be aware of proper application of the eyelash serum.A prospective study involving bimatoprost use in 52 patients with glaucoma or ocular hypertension investigated adverse effects of use.The strength of the bimatoprost solution was not noted.Clinical evaluation using photographs indicated 50% (n = 26) of participants objectively having new iridial pigmentation after the end of a 6-month period of daily evening application. 28This study did not specify whether follow up was conducted to assess reversibility of pigmentation.
Another well-known side effect in patients using prostaglandin analogs for the treatment of glaucoma is prostaglandin-associated periorbitopathy (PAP).PAP can be further differentiated into categories that can include sulcus deepening, periorbital fat loss, and enophthalmos. 29A retrospective observational study by Kucukevcilioglu with periorbital fat loss being the highest reported adverse PAP effect in all groups. 29Should patients discontinue treatment due to any of the above intolerable side effects, they should be counseled that eyelash growth is expected to return to its pretreatment level.
A gel formulation of 0.03% bimatoprost was developed to address some of the above side effects.Morris et al. performed a RCT exploring the side effect profile of a gel formulation of bimatoprost on 20 breast cancer patients (40 eyelids).This formulation was intended to decrease intraocular absorption of the medication by allowing precise application of the gel to the eyelash line.This study found an improved side effect profile for the gel formulation compared to the solution formulation, with a decreased report of ocular burning, pruritis, and eyelid hyperemia compared to the solution formulation. 30One participant from the gel formulation RCT reported lateral malar hyperpigmentation but preferred to continue treatment due to beneficial cosmetic effects on lash length, number, pigmentation, and thickness. 30Wester et al. evaluated the use of 0.03% bimatoprost gel on eyelash growth and found that it was associated with increased length of the eyelashes compared to placebo (p = 0.009). 31

| Latanoprost
Latanoprost, an FDA-approved product for treating ocular hypertension and open-angle glaucoma, is a synthetic PF2α analog.
Unlike bimatoprost, latanoprost has not achieved FDA approval for treating eyelash hypotrichosis; however, the mechanism is like that of bimatoprost. 13Despite lacking FDA approval, latanoprost has shown promising results in improving the appearance of eyelashes.
Coronel-Pérez et al. conducted a nonblinded, nonrandomized clinical trial of 54 patients with alopecia universalis, evaluating the efficacy of 0.005% latanoprost solution for the treatment of eyelash alopecia.They found that it induced acceptable response, classified as moderate or total regrowth, in 45% of patients. 32There were no complications.
Yazdanian et al. compared the eyelash-lengthening effect of latanoprost in 10 adults (20 eyes) and 10 children (20 eyes) with glaucoma.They found that both groups showed statistically significant eyelash growth at month 6 compared to baseline (p = 0.000).
No statistically significant difference was found between adults and children regarding the mean eyelash length difference. 33Similarly, Sugimoto et al. performed a quantitative analysis of eyelash lengthening following topical latanoprost therapy in 17 patients.They The effect of bimatoprost ophthalmic solution 0.03% on eyelash growth cycle.
found statistically significant differences from the baseline at 2 weeks, 6 weeks, and 10 weeks (p < 0.001). 34e most commonly reported side effect of latanoprost is hyperpigmentation of intra-or extra-ocular structures. 35Chiba et al. conducted a prospective study of iridial pigmentation and eyelash changes following ophthalmic treatment with latanoprost and found the incidence of iridial pigmentation to be 6.3% at 1 month, 15.7% at 3 months, 37.8% at 6 months, and 56.5% at 12 months.However, the authors noted an increase in eyelash length of 33.8% at 3 months, 44.4% at 6 months, and 46.2% at 12 months.They did not note changes in intraocular pressure, and there was no statistically significant relationship between eyelash growth and iridial pigmentation. 36However, there was a significant relationship between 0.005% daily solution of latanoprost eye drops and iridial pigmentation in 11 Dutch female rabbits in a study by Zhan et al. 26 The solution was applied to one eye twice daily on weekdays and once daily on weekends for 6 months.Ten of the rabbits showed some degree of heterochromia, with the newly pigmented eye being the latanoprost treated eye in nine of 10 rabbits.Standardized color photographs were taken at monthly intervals during the course of the study and evaluated by separate observers.Latanoprost treatment was observed to cause eyes that were originally described to have yellow-brown, blue/gray-brown, or green-brown to become more uniformly brown in color. 26m et al analyzed adverse effects of using 0.005% latanoprost eyedrops to lower intraocular pressure in 198 participants.
Participants were to apply the serum twice daily for 1 year.
Follow-ups were conducted at 6 and 12 months.Clinical evaluation indicated 7% (n = 14) of eyes treated with latanoprost had increased pigmentation after 6 months compared to preclinical evaluation, and 12% (n = 24) had increased pigmentation after 12 months. 37ostaglandin-associated periorbitopathy is also commonly noted with the use of latanoprost.However, when compared to bimatoprost, the retrospective study by Kucukevcilioglu et al indicated that when using a 0.005% serum with latanoprost daily for 1 month, there was milder PAP noted, such as periorbital fat loss, compared to the more prominent findings, such as sulcus deepening and enophthalmos, noted with 0.003% serum with bimatoprost. 29 addition, a prospective case report on four patients who experienced sunken eye changes from using either travoprost (n = 2) or bimatoprost (n = 2) serums, three of the four had resolution of sunken eye appearance after switching to latanoprost.The last participant (previously using travoprost) halted the serum use altogether.After resolution of sunken eye symptoms, this participant started using latanoprost, and had no recurrence of sunken eye symptoms in the 6 months of follow-up. 38tanoprost, with Grade B and Level 2 evidence, shows promising efficacy for enhancing eyelashes despite lacking FDA approval.
Systematic reviews are recommended for a comprehensive understanding of latanoprost's efficacy and safety in eyelash enhancement to lead to potential Grade A recommendation.

| Tafluprost
Tafluprost is a synthetic prostaglandin analog that has been deemed valuable in treating conditions like glaucoma and ocular hypertension.When applied to the eye, tafluprost rapidly converts into its biologically active form, tafluprost acid, through hydrolysis in the cornea.Tafluprost acid binds strongly to prostanoid fluoroprostaglandin receptors in the eye, eliciting agonistic effects that lower intraocular pressure. 41When applied to eyelashes, the proposed mechanism is similar to that of other prostaglandin analog, by prolonging the anagen phase and shortening the time between telogen and restart of anagen phase. 13This maximizes the time the hair follicles spend in the anagen growth phase, leading to longer, thicker, and darker eyelashes. 13Giannico et al investigated impact on eyelash growth on New Zealand white rabbits who received either bimatoprost, tafluprost, travoprost, and latanoprost in the left eye daily for 4 weeks.The right eye was used as a control in this study. 42Results indicated that the 0.0015% tafluprost serum yielded similar results to a 0.03% bimatoprost serum, growing from an average of 9.97 mm-11.03mm.While tafluprost has not been extensively studied in eyelash serums, these results indicate it should be explored further in its use to grow eyelashes.Adverse effects were studied by Inoue et al to compare periocular reactions to different prostaglandin analogues.Two hundred and fifty patients were assigned to use either tafluprost, bimatoprost, latanoprost, isopropyl unoprostone, or travoprost for 3 months in one eye. 28The opposite eye was used as a control.The 50 patients in the tafluprost group applied the serum once a day in the evening.Images before and after treatment were analyzed by ophthalmologists.There was no significant difference in eyelid pigmentation among the five groups.Four percent (n = 2) of participants in the tafluprost group were noted by physicians to have significant eyelid pigmentation. 28oue et al did not mention the concentration of tafluprost serum nor whether eyelid pigmentation faded after discontinuation.Sutton et al conducted a placebo-controlled study where 49 males were randomized into two groups to receive tafluprost serum, either 0.0025% or 0.005% in one eye once daily over the course of 7 days.
Both doses were tolerated in participants, and no participants withdrew from the study due to adverse effects.While Sutton et al focused on examining adverse effects of tafluprost, no comment was made on the effect on eyelash growth. 43fluprost earns a Grade B clinical recommendation for enhancing eyelashes based on Level 2 evidence per the Oxford Centre for Evidence-Based Medicine, but limited studies require further exploration.Strengthening recommendations calls for prioritizing systematic reviews or meta-analyses in future research to comprehensively assess efficacy and adverse effects. 8

| Dechloro ethylcloprostenolamide
Dechloro ethylcloprostenolamide, a compound used in several OTC eyelash serums, is a novel synthetic prostaglandin analog.The mechanism of action of this topical medication is thought to be identical to the mechanism of the FDA-approved prescription prostaglandin analog bimatoprost, however, it has never been formally assessed. 44oy and Lin conducted a randomized, double-blind clinical trial on There were no complications, but 15 of the 34 patients did report transient irritation upon initial use, but this was noted to resolve after several days. 44Results were not specific as to which groups the 15 patients that reported irritation were in.There remains a lack of research demonstrating the safety of this compound.
The absence of specificity trials and safety data grants Dechloro Ethylcloprostenolamide a Grade C clinical recommendation due to extrapolation of data from Level 2 evidence, calling for additional well-designed studies that also address safety concerns and potential limitations in study design or available data. 8assess for eyelid fat and volume loss after using eyelash serums with this ingredient. 46The authors found that a 35-year-old female developed thin, wrinkled, and hyperpigmented skin along with periorbital hallowing after 10 months of daily treatment with an eyelash serum containing isopropyl cloprostenate.This adverse effect reversed after 6 months of discontinuation of therapy.The study also identified 15 additional patients who exhibited statistically significant evidence of prostaglandin-associated periorbitopathy while using various formulations of prostaglandin analogs.The article did not specify which particular prostaglandins were employed by these 15 patients.Instead, these patients were included if their serums contained either bimatoprost, norbimatoprost, isopropyl cloprostenate, dechloro-dihydroxy-difluoro-ethylcloprostenolamide, or methylamido-dihydro-noralfaprostal.This collective data underscores the association between prostaglandins, including isopropyl cloprostenate, and prostaglandin-associated periorbitopathy. 46inical recommendations for isopropyl cloprostenate are challenging due to the absence of formal research on its efficacy and limited safety data.Anecdotal evidence suggesting enhanced eyelash appearance lacks substantiation from formal studies, resulting in a Grade D recommendation with level 5 evidence and inconsistent studies. 8

| Capsule summary
Topical bimatoprost is the only FDA-approved prostaglandin analog used for treating hypotrichosis of the eyelashes.It has strong evidence supporting its efficacy in increasing eyelash length, thickness, and darkness. 1Cosmeceutical formulations should contain a concentration of 0.03% of bimatoprost to be effective. 1Other prostaglandin analogs, such as latanoprost, fluprostenol, dechloro ethylcloprostenolamide, and isopropyl cloprostenate have not been adequately assessed for safety despite varying evidence suggesting their efficacy.Dermatologists should counsel patients to approach with caution if using eyelash serums that contain these ingredients until further safety profiles are available.In patients with hypotrichosis of the eyelashes or patients seeking cosmetic eyelash enhancement, dermatologists should discuss the lack of safety data with their patients and discuss topical bimatoprost as an FDAapproved alternative.
Many over-the-counter eyelash serums are not correctly labeled.

Rahman et al. conducted a study to screen and quantify undeclared
PF2α analogs in eyelash-enhancing serums purchased on the internet. 47Liquid chromatography-tandem mass spectrometry evaluated 64 eyelash serums from 34 websites.Four products were found to contain bimatoprost, the only FDA-approved ingredient for eyelash enhancement, despite only one of these serums indicating it on the package label.In addition, eight other serums contained unknown PF2α analogs that have never been used in pharmaceuticals. 47This study demonstrates that many eyelash serums contain unstudied PF2α analogs or do not accurately list their ingredients, which poses a health and safety risk to patients.For this reason, extreme caution must be practiced when recommending over-the-counter eyelash serums to patients.
As many popular eyelash serums are made with ingredients that are not FDA-approved for eyelash growth, it is important to conduct further research into the safety and efficacy of these eyelash serum ingredients.An increase in research can allow for more variety in options available to consumers, in a way that will allow them to understand studied risks and benefits of eyelash serums rather than rely on anecdotal evidence.

| NON -PROS TAG L ANDIN ANALOG S
Several over-the-counter non-prostaglandin analogs are used in eyelash serums due to their reported ability to target eyelash strength and growth.These ingredients include peptides, growth factors, ginseng, panthenol, proline, pumpkin seed oil, and Jarilla Coffea extract (See Table 3).The ingredients discussed were also analyzed and given clinical recommendations for eyelash serums based off the Oxford Centre for Evidence-Based Medicine (See Table 4).4).Other unsupported benefits of peptides include hair conditioning and protection against breakage. 44To date, studies evaluating peptides for hair growth are limited to the scalp.Lee et al. investigated the efficacy and safety of 5-aminolevulinic acid and glycyl-histidyl-lysine peptide for treating male pattern hair loss.They found a dose-dependent increase in hair count at 6 months (p < 0.05) for patients receiving the combination therapy. 51No adverse events were reported.
Leuangarun and Panchaprateep conducted a prospective, tripleblinded, RCT on the herbal extract combination of biochanin A, acetyl tetrapeptide-3, and ginseng extracts versus 3% minoxidil for the treatment of androgenic alopecia.Thirty-two participants with androgenic alopecia received twice-daily applications of either the herbal combination or minoxidil.Efficacy was determined based on photographic assessments of results compared to pretreatment photographs.Assessment of the photographs indicated a response to both treatments was most evident in patients at 24 weeks.They found the herbal extract to demonstrate comparable efficacy to minoxidil 3%. 52No adverse events were reported from using the herbal combination.No formal studies have been performed assessing the safety or effectiveness of peptides for eyelash growth.
Clinical recommendations for peptides in eyelash serums are challenging due to limited research on their efficacy and safety for eyelashes.With a Grade D recommendation indicating level 5 evidence and inconsistent studies, more comprehensive research specifically targeting eyelash growth is necessary to establish a robust foundation for clinical guidance. 8

| Growth factors
Growth factors (GFs) control cell growth, proliferation, and differentiation through several advanced intercellular and intracellular signaling pathways. 53Due to their large molecular size of nearly 2 kilodaltons, these molecules cannot penetrate through the epidermis layers. 53In recent years, topical and injectable formulations of A Grade C recommendation can be given to growth factors in eyelash serums due to evidence from limited literature with level 4 studies and extrapolations from level 2 and 3 studies.Further robust research needed to confirm safety and efficacy for eyelash enhancement. 8

| Ginseng
Ginseng is a phytochemical, non-prostaglandin analog used in many OTC eyelash serums.The mechanism of action of this plant root appears to induce the anagen growth phase of hair growth while decreasing the catagen phase, allowing the ingredient to be comparable to other prostaglandin analogs 55 (See Table 4).This herbal ingredient downregulates TGF-Beta to prevent the catagen phase.A second proposed mechanism of action for this ingredient involves increasing Bcl-2 expression and decreasing Bax expression, leading to a stimulation of dermal papillary-cell proliferation. 56Murata et al. demonstrated that ginseng might have properties that inhibit the activity of 5a-reductase, specifically in red-ginseng extract samples, reducing testosterone-mediated suppression of hair growth. 57limitation to using ginseng for eyelash growth is that many human studies are preclinical.While they have shown promise, human trials must be conducted to fully understand this plant root's effectiveness.While side effects for oral ginseng have been studied to be correlated to changes in blood pressure, nervousness, and insomnia, topical ginseng (the form used in eyelash serums) is much less likely to be absorbed through the skin to cause these effects. 58nseng's saponin must pass through the intestines to be activated by the digestive process. 58However, adequate safety studies for ginseng in eyelash serums are lacking.
Ginseng lacks formal studies on its efficacy and safety for eyelash growth.Limited trials and preclinical evidence suggest po-

| Panthenol
Pantothenol, a precursor to pantothenic acid/vitamin B5, is often added to OTC eyelash serums due to the widespread belief that it decreases hair breakage and loss. 59The mechanism of action is thought to be related to increased cell viability, elongation of the anagen phase, and suppression of apoptotic factors 59 (See Table 4).Expression of vascular endothelial growth factor was upregulated in outer root sheath cells. 60A study by Nisbet et al. evaluated the side effect profile of anti-redness molecules used in topical ointments for facial xerosis, including panthenol.There were no reported productrelated adverse effects in this study. 61There have been no formal research studies evaluating the efficacy or safety of panthenol for eyelash growth.
Using the Oxford Centre for Evidence-Based Medicine to assign clinical recommendations establishes pantothenol as a Grade D recommendation due to a lack in formal research on its efficacy and safety for promoting eyelash growth, emphasizing the need for rigorous studies to establish safety and effectiveness of panthenol in eyelash serums. 8

| Proline
Proline is a popular ingredient in OTC eyelash serums, although limited research is available to support its current use.Proline is one of the primary substrates in synthesizing collagen, which gives hair structure and quality 62 (See Table 4).Hydroxylation of proline increases the stability of the triple helix structure involved in collagen. 62A review by Karna et al. suggests that proline dictates the rate of collagen synthesis, and that endogenous proline is insufficient to provide maximal growth. 62Giving a stronger foundation to the hair, in turn, gives the appearance of thicker hair.
Due to their size, amino acids can penetrate the skin to the dermis to certain degrees of success. 63Proline weighs 110 Daltons, whereas the skin has a 500 Dalton limit for adequate absorption. 63Proline has also been studied to have antiaging benefits to the skin, which could lead to reduced appearance of wrinkles around the eye with the topical application via enhancing the elasticity of the stratum corneum. 63Proline is not a foreign substance to the body and is an FDA-approved food additive. 63rrent data on dermal irritation finds that proline is one of the amino acids deemed safe in cosmetics and skincare products.
Accidental ingestion in large amounts could cause seizures and intellectual disability; however, these have not been shown in topical application. 63ing the Oxford Centre for Evidence-Based Medicine to assign clinical recommendations establishes proline as a Grade D clinical recommendation.Due to limited evidence, a need for further research to establish the safety and efficacy of proline in eyelash serums. 8

| Pumpkin seed oil
Pumpkin seed oil is a common ingredient found in OTC eyelash serums.It is thought to promote hair growth through inhibitory effects on the enzyme 5 alpha-reductase and antioxidant and anti-inflammatory properties 64,65 (See Table 4).Ibrahim et al. conducted a RCT of pumpkin seed oil versus minoxidil 5% topical foam where 60 female patients were randomly assigned between the two groups for the treatment of female pattern hair loss.Each group was instructed to apply 1 mL of either topical pumpkin seed oil or minoxidil 5% foam daily to scalp.After 3 months, there was a statistically significant decrease in hair shaft diversity (p < 0.001) in both respective groups after treatment and a statistically significant reduction in vellus hairs (p < 0.001). 64Hair shaft diversity was evaluated using trichoscopy to assess for discrepancies in thickness of hairs.There was also no significant difference in results between both groups.Similarly, Cho et al. conducted a randomized, placebo-controlled, double-blind study to evaluate the efficacy of pumpkin seed oil for the treatment of hair growth in male patients with mild-to-moderate androgenetic alopecia.
Seventy-six participants were instructed to take a total of 400 mg of pumpkin seed oil daily for 24 weeks.They were instructed to take two capsules (100 mg each) before breakfast and two capsules before dinner.Patient self-assessment was observed at both 12 and 24 weeks of the study.While no intergroup differences were noted at week 12, statistically significant change was found in self-rated improvement score (p = 0.013) and self-rated satisfaction score (0.003) compared to placebo by the end of week 24.
In addition, the pumpkin seed oil group had an increased mean hair count compared to the placebo (p < 0.001).There were no reported adverse events. 66Despite these positive results, no studies have evaluated pumpkin seed oil for growth of the eyelashes or safety for use on the eyelids.
Pumpkin seed oil earns a Grade D clinical recommendation due to a lack of research on its efficacy and safety for eyelash growth, despite positive effects observed in studies on scalp hair.Caution is advised in recommending pumpkin seed oil in eyelash serums, highlighting the need for dedicated studies to assess its effectiveness and safety for eyelashes. 8

| Jarilla Coffea extract
Extracts from Coffea arabica and Larrea divaricate (Jarilla) have been used in Argentina for hair growth in people suffering from alopecia involving the eyebrows and eyelashes. 67The mechanism of action is unknown. 67 Efficacy was defined as their hair growth in terms of thickness, length, and appearance of new hair.Tolerability was also assessed within participants.Thickness and length were analyzed via photographic analysis and measurements.Ophthalmic appointments in all patients indicated that there were no adverse effects due to potential irritation on the eyelids between those treated with the product and those treated with placebo.The product was found to significantly increase the thickness of the eyelashes in all women participating in the study, with the percent change found to be about 19.44%.No adverse effects were reported. 67inical recommendations for Jarilla-Coffea extract in eyelash serums are constrained by limited literature, despite positive outcomes and a statistically significant increase in eyelash thickness.
The lack of mechanistic understanding and limited research warrants a Grade D recommendation, emphasizing the need for further studies to determine its efficacy and safety for broader application in eyelash serums. 8

| Castor oil
Castor oil is one of the most popular ingredients in vegan lash serums.However, there are insufficient studies reporting a significant difference in lash growth via the use of castor oil.One proposed mechanism is that castor oil may increase prostaglandin production by increasing ricinoleic acid. 68Ricinoleic acid activates the EP3 prostanoid receptor for prostaglandin E2. 68 To our knowledge, published articles are only available in relation to the gastrointestinal tract. 69e to insufficient studies to expand on its relationship to eyelash serums, many claims of the use of castor oil appear to be mainly anecdotal.
Because castor oil lacks adequate scientific evidence to substantiate claims of significant lash growth, clinical recommendations using the Oxford Centre for Evidence-Based Medicine can be set to a Grade D, emphasizing the need for further research to establish its effectiveness and safety for eyelash growth in the absence of robust evidence.

| Brief review of litigation
The adverse effects of OTC eyelash serums have been the source of several malpractice lawsuits.One skincare brand, Rodan + Fields, was a part of three class action lawsuits secondary to the side effects of macular edema, iris color change, and ocular irritation caused by their eyelash serum containing isopropyl cloprostenate. 70In early 2022, another skincare company, Grande Cosmetics selling an eyelash serum containing the compound isopropyl cloprostenate was also faced with a lawsuit.It was filed by a consumer who purchased this serum and subsequently developed a new eye mass, which she claimed to be related to the serum. 71These cases demonstrate that further research is needed into the safety profile of eyelashenhancing products before they can be safely recommended to patients, and additional regulation to the manufacturing process and safety.Many of the ingredients commonly used in eyelash serums lack evidence of clinical efficacy, especially regarding eyelash growth (versus hair growth elsewhere).This is notable because the eyelid is a much more sensitive skin region and is more likely irritated by topical products.

| S TRENG TH S AND LIMITATI ON S
This review delves into the realm of eyelash-enhancing products, examining both FDA-approved and OTC options.It serves as a resource to examine the efficacy and safety profiles of these formulations.This review excels in its comprehensive exploration with different article types and studies of both prostaglandin and nonprostaglandin analogs.The review offers clinicians a thorough understanding of FDA-approved bimatoprost, which stands out with solid evidence supporting its efficacy in enhancing eyelash length, thickness, and darkness.The incorporation of legal cases against skincare brands adds a real-world dimension, emphasizing the consequences of inadequate safety assessments and potential risks associated with nonregulated products.
Despite its strengths, the authors acknowledge limitations.Nonprostaglandin analogs, including peptides, growth factors, ginseng, panthenol, proline, pumpkin seed oil, and Jarilla Coffea extract, present promising mechanisms for eyelash enhancement.However, there are limitations due to an inadequate number of formal studies on safety and efficacy for eyelash growth.Dermatologists may face challenges in recommending these products due to the lack of regulatory endorsement and comprehensive research.While safety profiles for these ingredients are highlighted, a comprehensive overview of potential risks is lacking.The reliance on anecdotal evidence, particularly regarding castor oil, may introduce bias, and the scope of legal cases, while illustrative, might benefit from broader industry-wide examination.In conclusion, this review highlights the use of FDA-approved bimatoprost and the potential of many OTC ingredients, while also emphasizing the need for continued research, to establish comprehensive safety and efficacy profiles for various eyelash-enhancing ingredients.
To provide appropriate and safe patient recommendations, dermatologists must be familiar with various ingredients commonly used in eyelash serums.In this paper, we explored prostaglandin analogs and non-prostaglandin analogs widely used in OTC and prescription eyelash serums, emphasizing efficacy and adverse effects.Bimatoprost is the only FDA-approved bioactive molecule shown to increase eyelash length safely.Clinicians should be aware of the various bioactive molecules incorporated in eyelash serums and understand their side effects before making recommendations to patients.

CO N FLI C T O F I NTE R E S T S TATE M E NT
Author Geoffrey Potts has spouse that is a consultant for Rodan + Fields.

E TH I C S S TATEM ENT
There were no ethical concerns related to participant subjects in the course of this study.The research design and methodology focused solely on the analysis of existing data and literature, and ethical considerations pertaining to participant welfare, informed consent, and confidentiality were not applicable.The review process adhered to ethical principles by appropriately citing and acknowledging the original sources of information.
cal recommendations for eyelash serums based off the Oxford Centre for Evidence-Based Medicine.The Oxford Centre for Evidence-Based Medicine uses a comprehensive system for appraising the quality of evidence and formulating recommendations.The Levels of Evidence range from Level 1 (systematic reviews and meta-analyses) to Level 5 (expert opinion without explicit critical appraisal), with each level indicating varying degrees of evidence strength.Concurrently, the grades of recommendations consist of Grades A-D.Grade A corresponds to high-quality evidence with a strong recommendation, Grade B indicates moderate-quality evidence with a moderately strong recommendation, Grade C reflects low-quality evidence with a weak recommendation, and Grade D denotes very low-quality evidence with a weak recommendation.These tools collectively empower healthcare professionals to make informed, evidencebased decisions in clinical practice. 8 Glaser et al. conducted a multicenter, randomized controlled trial evaluating the long-term safety and efficacy of bimatoprost solution 0.03% after 1 year of use.Three hundred and sixty-eight participants received clinical graded scores of 1 (none/minimal eyelash prominence) or 2 (moderate eyelash prominence) out of 4 in the Global Eyelash Assessment (GEA) and patient graded scores of 1 (very much disagree) or 2 (disagree) when rating Domain 2 of the Eyelash Satisfaction Questionnaire (ESQ).Domain 2 of the ESQ covers opinions on confidence, attractiveness, and professionalism.Participants placed one drop of treatment per eye once daily each evening.The primary efficacy endpoint was met for all treatment groups.Efficacy was measured based on the proportion of participants that responded by month 4 of treatment with an increase in their baseline GEA score by at least 1 point, and an increase in their ESQ score for confidence, attractiveness, and professionalism (Domain 2 of ESQ) by at least 3 points from baseline.

- 30 Clinical- 32 Clinical
Bimatoprost users had more severe and frequent periorbitopathy compared to other prostaglandins.*-Milder prostaglandin-associated periorbitopathy (PAP) noted with 0.005% latanoprost serum compared to 0.003% serum with bimatoprost.*-No randomization due to retrospective nature of the study -The true dates of the onset of PAP were not determined Morris et al.Gel formulation of 0.03% bimatoprost has an improved side effect profile compared to the solution.-Decreased side effects included decreased ocular burning, pruritis, 03% bimatoprost gel increases eyelash length compared to placebo.*-Patients were evaluated after only 6 weeks-a longer study may be beneficial to assess benefits and side effects of treatment Coronel-Pérez et al. latanoprost solution showed acceptable response (moderate or total re-growth) in 45% of patients with eyelash alopecia.*-Nonblinded and nonrandomized trial adults and children with glaucoma showed statistically significant eyelash growth at month 6 compared to baseline.*-Authors note side effects are minimal when properly to the base of the eyelashes at the lid margin Sugimoto et al. 34 Clinical trial (quantitative) -Latanoprost -Statistically significant eyelash lengthening following topical latanoprost therapy.*-All adverse events were recorded even if not potentially due to latanoprost treatment.Chiba et al. 36 Prospective study -Latanoprost -Increase in eyelash length noted (33.8% at 3 months, 44.4% at 6 months, 46.2% at 12 months).-Incidence of iridial pigmentation with latanoprost: 6.3% (1 month), 15.7% (3 months), 37.8% (6 months), 56.eyes treated with 0.005% latanoprost had increased pigmentation after 6 months.-12% of eyes had increased pigmentation after 12 months -Darkening of iris color was deemed cosmetic and not clinically significant Fabbrocini et al. 39 Clinical trial (double blind RCT) -Fluprostenol -15 keto fluprostenol isopropyl ester gel demonstrated a statistically significant increase in eyelash length compared to a placebo gel for idiopathic eyelash hypotrichosis.*-80 percent of participants receiving fluprostenol reported that they would decrease dedicated time spent on the application of mascara versus 20 percent in the control group.-Small sample size (40 participants).-Short duration of study Feletti et al. 40Case reports -Travoprost -Cases included a 12-year-old girl, a 45-year-old woman, and a 32-yearold woman who experienced pigmentation after months of topical travoprost use.-Limited to case reports -Pigmentation gradually decreased after cessation of treatment.0015% tafluprost serum yielded similar eyelash growth results as 0.03% bimatoprost serum in New Zealand white rabbits.*Short duration of study Inoue et al. 28 Observational study -Tafluprost -No significant difference in eyelid pigmentation among patients using tafluprost, bimatoprost, latanoprost, isopropyl unoprostone, or travoprost for 3 months.-4% (n = 2) of tafluprost group had significant eyelid pigmentation.*-Concentration of tafluprost serum not mentioned -No mention of whether eyelid pigmentation faded after discontinuation Sutton et al. 43 Clinical trial (placebo-controlled) -Tafluprost -Both 0.0025% and 0.005% tafluprost serum were well-tolerated with no adverse effects reported in a 7-day study in 49 males.-No comment on the effect of tafluprost on eyelash growth Choy and Lin 44 Clinical trial (randomized, double-blinded) -Dechloro ethylcloprostenolamide -Statistically significant increase in eyelash length after 4 weeks of daily use of topical dechloro ethylcloprostenolamide.* -15 of 34 patients reported transient irritation upon initial use, which resolved after several days.-Lack of information about exact concentrations in the study Horvath et al. 45 Case report -Isopropyl cloprostenate -Greenish periocular discoloration reported about a year into treatment.*-Periocular discoloration and increased melanin granule counts in epidermis resolved upon discontinuation.Jamison et al. 46 Randomized control study -Isopropyl cloprostenate -35-year-old female developed thin, wrinkled, and hyperpigmented skin, periorbital hallowing.*-Side effects reversed after discontinuation.
et al was performed to assess the frequency of periorbitopathy in 105 participants who were using either 0.03% bimatoprost (n = 15), 0.005% latanoprost (n = 70), or 0.004% travoprost (n = 20) daily in one eye for over a month.The other eye was used as a control.The participants using bimatoprost were found to have more severe and frequent PAP than other prostaglandins, especially in older adults.All the prostaglandins used in this study had statistically significant periorbitopathy findings (p < 0.05).Fifty-seven of the total 105 patients had noted prostaglandin-associated periorbitopathy, with frequencies of 93.3% (n = 14) in the bimatoprost group, 70% (n = 14) in the travoprost group, and 41.4% (n = 29) in the latanoprost group,

3. 3 |
FluprostenolFluprostenol is a synthetic PF2α analog with potent prostaglandin F receptor agonist activity.Travoprost, a prodrug to fluprostenol, is used to treat ocular hypertension and open-angle glaucoma.Fabbrocini et al. conducted a monocentric, double-blind, vehicle controlled RCT on the 15 keto fluprostenol isopropyl ester gel for idiopathic eyelash hypotrichosis.Forty 18-year-old women who reported low confidence levels on their ESQ were divided into two groups.Group 1 (n = 20) applied 15 keto fluprostenol isopropyl ester gel once daily.Group 2 (n = 20) only used a vehicle gel daily.After 2 months, the researchers found a statistically significant increase in eyelash length compared to the placebo.Eighty percent of participants receiving fluprostenol reported that they would decrease dedicated time spent on the application of mascara versus 20 percent in the control group.Only one patient reported symptoms of headache and ocular sensation heaviness.No other side effects were reported.39Feletti et al. report three separate case reports of hyperpigmentation following twice daily application of products that contain travoprost, the more active enantiomer of fluprostenol.40The first case described a 12-year-old girl who after 6 months of topical travoprost developed periocular pigmentation.After cessation of treatment, pigmentation gradually began to decrease after 1 month.The second case describes a 45-year-old woman who experienced blue-gray pigmentation after 7 months in her upper and lower eyelids.Again, after cessation of therapy, patient noticed a gradual decrease in pigmentation.However, this case describes that brownish pigmentation remained on the lower eyelid even 4 months after treatment.The third case describes a 32-year-old woman who noticed eyelid pigmentation after 6 months of use of travoprost.After discontinuation, patient saw gradual disappearance of pigmentation.40Based on Level 2 evidence demonstrating statistically significant efficacy in enhancing eyelash length, fluprostenol receives a Grade B clinical recommendation.Encouraging participation in systematic reviews or meta-analyses and gathering data from high-quality randomized controlled trials can enhance the evidence base, potentially leading to a Grade A recommendation.8 34 healthy participants randomized to receive one of three concentrations of "MD Lash Factor serum" (full strength (A), half strength (B), and quarter strength (D)) with active ingredient topical dechloro ethylcloprostenolamide or placebo (C). 44Exact concentrations were not defined in the study.Results indicated statistically significant increase in eyelash length after 4 weeks of daily use of topical dechloro ethylcloprostenolamide, with 0.63 mm of growth in the placebo group compared to 1.69 mm (A), 1.94 mm (B), 1.44 m, and 1.44 mm (D) of growth reported in the three experimental groups.

53 TA B L E 3
GFs have emerged for cosmetic purposes.GFs target various levels of the hair growth cycle and are thought to stimulate proliferation and modulate growth-phase transitions at the level of the bulge stem cells in the dermal papilla of the hair.Results of non-prostaglandin analogs.aminolevulinic acid and glycyl-histidyllysine peptide combination therapy increased hair count at 6 months.*-No adverse events reported.-Studieson peptides for eyelash growth are limited to the scalp.Leuangarun and Panchaprateep 52Prospective, tripe blinded,RCT-Peptides -Herbal extract combination (biochanin A, acetyl tetrapeptide-3, and ginseng extracts) demonstrated comparable efficacy to minoxidil 3% in the treatment of androgenic alopecia.*-No adverse events reported -No formal studies in eyelash serums Sachdev et al. 54 Clinical trial (nonblinding) -Growth factors -Polygrowth factor serum improved eyelash curl, thickness, luster, length, and volume compared to baseline at day 30, persisting until day 90.*-No adverse effects reported -Nonblinded trial Murata et al. 57 Preclinical -Ginseng -Ginseng extract samples reduced testosterone-mediated suppression of hair growth.*-No adverse effects reported -Preclinical studies, limited human trials Shin et al. 59 Preclinical -Panthenol -D-panthenol increased cellular proliferation marker Ki67 in human dermal papilla cells and decreased markers for apoptosis and cell senescence.*-No formal research studies completed Nisbet et al. 61 Clinical trial -Panthenol -No reported product-related adverse effects with anti-redness molecules used in topical ointments containing panthenol.-No adverse effects reported Karna et al. 62 Review -Proline -Proline may dictate the rate of collagen synthesis, enhancing hair quality.*-No adverse effects mentioned Ibrahim et al. 64 Randomized controlled trial -Pumpkin seed oil -Topical pumpkin seed oil and minoxidil 5% both reduced hair shaft diversity and vellus hairs with no significant difference between groups.*-No adverse effects reported -No studies on eyelash growth Cho et al. 66 Randomized controlled trial -Pumpkin seed oil -Pumpkin seed oil increased hair count and self-rated improvement and satisfaction scores in male patients with androgenetic alopecia.*-No adverse effects reported -No studies on eyelash growth Alonso et al. 67 Clinical trial (non-blinding) -Jarilla-Coffea extract -Jarilla-Coffea extract increased the thickness of the eyelashes in participants.*-No adverse effects mentioned -Small study size -Nonblinded trial *Statistically significant with p < 0.05.Sachdev et al. conducted a 90-day, open-label safety and efficacy study on a polygrowth factor serum to enhance eyelashes' appearance in women aged 15-45. 54The serum contained keratinocyte growth factor, fibroblast growth factor, insulin-like growth factor, and vascular endothelial growth factor.Twenty-nine female patients completed the study.Patients were instructed to apply serum once nightly for 90 days to both upper and lower eyelids.Measurements were assessed on day 0 (baseline), and days 30, 60, and 90.The authors found improvement in curl (50.83%), thickness (35%), luster (11.43%), length (10.52%), and volume (9.3%) compared to baseline at day 30 and persisted until the last visit at day 90. 54No adverse effects were observed.However, additional randomized controlled trials evaluating growth factors for eyelash enhancement are needed before they can be safely recommended to patients.
tential benefits, but insufficient safety studies warrant a Grade D clinical recommendation, emphasizing the need for more comprehensive research to establish parameters for ginseng in eyelash serums. 8TA B L E 4 Non-prostaglandin analogs.
Shin et al. evaluated the effects of D-panthenol on human hair follicles, including outer root sheath cells and dermal papilla cells.The authors found D-panthenol to increase the cellular proliferation marker Ki67 in cultures of human dermal papilla cells and decrease caspases 3 and 9 and proteins p21 and p16, markers for apoptosis and cell senescence, respectively. 59In addition, anagen-inducing factors such as ALP, b-catenin, and versican were stimulated.
Alonso et al. conducted an open-label, placebocontrolled, prospective, 3-month study evaluating the efficacy of a Jarilla-Coffea combination extract to improve the appearance of the eyelashes and eyebrows in women.Nine women participated in the study.Participants were instructed to apply the Jarilla-Coffea extract gel on their eyelashes once a day for 90 consecutive days.

4. 9 |
Capsule summaryNumerous non-prostaglandin analogs are used in OTC eyelash serums which claim to have a beneficial cosmetic effect on the eyelashes.Many of these analogs are poorly researched and lack adequate safety assessments.Dermatologists should exhibit caution when recommending nonprescription eyelash-enhancing serums to patients.

Compound Mechanism of action FDA approval (eyelashes) Effect on eyelashes Side effects Clinical grade and level recommendation
Author(