Section C: Barrier Repair Ingredients

INTRODUCTION

The skin barrier is a watertight seal around the keratinocytes in the upper levels of the epidermis. It prevents evaporation of water from the surface of the skin, which is known as transepidermal water loss (TEWL). It is important to note that TEWL is not the same as sweating or perspiration. Increased TEWL occurs when a defect in the permeability barrier allows excessive water to be lost to the atmosphere. The skin barrier decreases TEWL and helps keep unwanted compounds out of the skin, such as allergens and irritants. Skin with an injured barrier is more susceptible to contact and irritant dermatitis as well as infection. Skin barrier perturbation can be caused by many different factors such as detergents, acetone, friction, ultraviolet exposure, prolonged or frequent water immersion, cholesterol-lowering drugs, low-fat diets, and genetic predisposition (e.g., to disorders of filaggrin).

The extracellular lipid mixture surrounding keratinocytes in the upper layer of the epidermis (stratum corneum or SC) is well known to be responsible for that layer’s water barrier function.¹ This lipid mixture, which is synthesized by lamellar bodies in the lower levels of the epidermis, is composed of 50 percent ceramides, about 25 percent cholesterol, and about 15 percent fatty acids.² Many studies have considered the effect of topically applying these important skin barrier lipids to improve skin barrier function and, thus, skin hydration. These investigations have demonstrated that the exogenously applied lipids must be in the proper ratio to form the correct three-dimensional structure of the skin barrier. When the correct ratios of ceramides, fatty acids, and cholesterol were used, the barrier recovered.³ Application of a mixture of cholesterol, ceramides, and the essential/nonessential free fatty acids palmitate and linoleate in an equimolar ratio allows normal barrier recovery, whereas a 3:1:1:1 ratio of these four ingredients accelerates barrier recovery.⁴ Today, the goal of the best barrier repair moisturizers is to provide these vital components in a 3:1:1:1 ratio.

AGE AND THE SKIN BARRIER

Older skin displays increased drug penetration, dryness, and other signs that the skin barrier may be impaired. Ghadially et al. showed that although the composition, dimensions, and lamellar structure of the bilayer lipids were normal in older individuals (80+ years), their skin had 30 percent less lipids than younger people (20–30 years old) and the number of focal SC lamellar bilayers was decreased. In addition, the barrier recovery time was much longer in older individuals.⁵ Waller and Maibach reviewed studies on lipids in aged skin and deemed the findings to be conflicting, with no consensus on whether or not older skin displays a different lipid composition as compared to younger skin.⁶

EXOGENOUSLY APPLIED LIPIDS

Topically applied lipids are transported to the nucleated layers of the SC (such as the spinous layer), internalized, and transported to the distal Golgi apparatus where they are incorporated into lamellar bodies.⁷ These lamellar bodies are present in the granular layer of the epidermis and are secreted (containing the newly synthesized lipids) into the intercellular space between the keratinocytes, thus contributing to the lipid bilayer membrane. This de novo synthesis of lipids is a crucial mechanism involved in barrier recovery.⁸

The lamellar or Odland bodies play an important role in barrier function, because they deliver enzymes that metabolize lipid hydrolases and lipids to the intercellular spaces of the SC. The lipid hydrolases metabolize lipid precursors such as cholesterol sulfate, phospholipids, sphingomyelin, and glycosylceramides into nonpolar products, which form the extracellular lamellar membrane.^1,9 This lipid processing cascade is affected by changes in pH and other metabolic activity that remains to be elucidated.^10,11 The presence of cholesterol seems to be important in the normal function of lamellar bodies in barrier repair.¹² When cholesterol levels are low, peroxisome proliferator-activated receptors (PPARs) and retinoid X receptors increase expression of ATP-binding cassette transporter A1 (ABCA1), a membrane transporter that regulates cholesterol efflux,¹³ resulting in increased transport of cholesterol across keratinocyte cell membranes. It is important to note that replacement of only one of the three primary components of the extracellular matrix (cholesterol, ceramides, or fatty acids) impairs the barrier, while replacement in a 1:1:1 ratio of all of these three classes of compounds repairs the barrier.³

ENDOGENOUSLY SYNTHESIZED LIPIDS

Basal cells are capable of absorbing cholesterol from the circulation; however, most lipids are synthesized in cells such as the keratinocytes. Exogenously applied lipids are incorporated into the keratinocytes and lamellar bodies in the granular layer of the SC.^3,8

There are three rate-limiting enzymes involved in the production of the main lipids of epidermal skin. They include 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (the rate-limiting enzyme in cholesterol synthesis), acetyl Co-A carboxylase (ACC), and the fatty acid synthase involved in the synthesis of free fatty acids and serine palmitoyl transferase (SPT), which is the regulatory enzyme for the synthesis of ceramides.^14,15 As expected, when skin barrier disruption occurs, the activity of these enzymes is enhanced in order to compensate for barrier dysfunction.^16,17 In addition, a group of transcription factors known as sterol regulatory element-binding proteins (SREBPs) regulate cholesterol and fatty acid synthesis. When epidermal sterols are decreased, the SREBPs are activated via proteolytic processes, enter the cell nucleus and activate genes, leading to increased production of cholesterol and fatty acid synthesis enzymes.^18,19 There are three known types of SREBPs: SREBP-1a, -1c, and SREBP-2. In human keratinocytes, SREBP-2 has been shown to be the predominant form and is involved in regulating cholesterol and fatty acid synthesis.¹⁸ Interestingly, the ceramide pathway is not affected by SREBPs.

FATTY ACIDS

Fatty acids play a crucial role in conferring hydrophobic characteristics to the epidermis. Their addition to cream formulations makes them slightly alkaline with a pH of 7.5 to 9.5.²⁰ The properties of the fatty acids, and the lipids derived from them, are markedly dependent on chain length and degree of saturation.²¹ Fatty acids that have double bonds are known as unsaturated while fatty acids without double bonds are known as saturated. Fatty acids are divided into categories depending on the length of their aliphatic tail chain. Short-chain fatty acids are fatty acids with aliphatic tails of fewer than six carbons, medium-chain fatty acids have 6 to 12 carbons, long-chain fatty acids have 13 to 21 carbons, and very long-chain fatty acids have aliphatic tails longer than 22 carbons. Unsaturated fatty acids found in skin include oleic acid, linoleic acid, α-linolenic acid, and arachidonic acid. Saturated fatty acids found in skin include palmitic and stearic acids. Studies investigating the use of fatty acids to repair the skin barrier have shown that linoleic, palmitic, and stearic acids can repair the barrier within two hours when applied exogenously.²²

A few notable fatty acids play an important role in inflammation. Linoleic acid, found in corn oil, sunflower seed oil, sesame oil, and safflower oil, forms a prostaglandin known as PGE₁, which exhibits anti-inflammatory and immune-enhancing properties. The fatty acid arachidonic acid forms PGE₂ and leukotrienes, which are highly inflammatory substances. Arachidonic acid is found rarely in plant oils but is abundantly present in animal-derived products such as meat, eggs, and dairy.

CERAMIDES

Ceramide 1 is the most important type of ceramide in skin. It is composed partially of linoleic acid. Because ceramides are expensive additions to skin care formulations, a less costly alternative is to use ingredients that contain linoleic acid, which may help the skin increase production of certain ceramides. Linoleic acid is found in vegetable oils, rapeseed, walnuts, soybeans, and flaxseed. Applying ceramide alone to the skin (without the proper ratio of fatty acids and cholesterol) disrupts the skin barrier and increases TEWL.²²

CHOLESTEROL

Cholesterol is found in the diet and is included in some moisturizers; however, most of the cholesterol found in the skin is produced by keratinocytes. Cholesterol is synthesized in the keratinocytes by the enzyme cholesterol sulfotransferase type 2B isoform 1b, abbreviated as SULT2B1b. The rate-limiting enzyme in this process is HMG-CoA reductase (see Chapter 21, Cholesterol). Cholesterol esters added to skin moisturizers without the presence of the correct ratio of ceramides and fatty acids have been shown to disrupt barrier function.¹

BARRIER DISRUPTION

Barrier disruption is caused when alterations occur in the ceramides, cholesterol, and fatty acids that constitute the bilamellar membrane that renders the SC watertight. The three-dimensional structure of this bilayer is important and many factors can affect its shape and function. Cholesterol levels can be reduced by cholesterol-lowering drugs such as statins, leading to decreased cholesterol in the bilamellar membrane. All three components can be damaged or diminished by surfactants, friction, ultraviolet light, prolonged water immersion, immersion in chlorine, and application of moisturizers. Stress and increased cortisol levels have been shown to damage the skin barrier.^23,24

Certain ingredients can negatively impact the skin’s barrier. Solvents such as propylene glycol can cause lipid fluidization or extraction that can alter the barrier.²⁵ Ingredients with a polar head group attached to alkyl chain lengths of around C₁₀–C₁₂ yield a potent penetration enhancer, for example, oleic acid,^26,27 which disrupts the barrier by inserting alkyl chains into ceramides. This leads to pools of oleic acid in the membrane, which becomes more fluid and easier for molecules to diffuse through than intact ceramides.²⁵ Other ingredients containing unsaturated alkyl chains can also enhance penetration but, in this case, C₁₈ appears near optimum. For such unsaturated compounds, the bent cis configuration disturbs intercellular lipid packing more so than the trans arrangement, which differs little from the saturated analogue.²⁸ Fatty acid analogues have been developed to function as penetration enhancers.²⁹

In summary, the epidermis and surrounding lipid barrier have been described as having a brick-and-mortar structure, with the keratinocytes as the bricks and the cholesterol, ceramides, and fatty acids representing the mortar. Many well-controlled intricate processes govern the formation of this brick-and-mortar structure and much can go awry that impairs its function. Skin care products can significantly influence the formation and proper functioning of this structure as has been shown by a multitude of studies.

REFERENCES

INTRODUCTION

SOURCE

Ceramides are derived from the precursor glucosylceramides (GC), which is formed in large quantities by the epidermis and stored in lamellar granules. The enzyme responsible for GC synthesis has been localized to the golgi apparatus and is called ceramide glucosyltransferase.⁴ Structured in lamellar sheets, the primary lipids of the epidermis – ceramides, cholesterol, and free fatty acids – play a crucial role in the barrier function of the skin (see Chapter 19, Barrier Repair Ingredients). The intercellular lipids of the SC are composed of approximately equal proportions of ceramides (which may constitute up to 40 percent),⁵ cholesterol, and fatty acids.⁶ Ceramides are found in the upper levels of the stratum corneum (SC) but are not found in significant supply in the lower levels of the epidermis such as the stratum granulosum or basal layer. This is simply because ceramides are produced in the lamellar bodies in the granular layer of the SC.

There are several ways to generate ceramides in mammalian cells^7–9:

1. Catabolism of sphingomyelin by the enzyme sphingomyelinase, which is coded by the gene SMPD1. (This is the most important pathway because sphingomyelin is abundant in most cell membranes.)

2. De novo synthesis from palmitate and serine by the enzyme serine palmitoyltransferase (SPT), which is coded in part by the gene SPTLC1.¹⁰

3. Hydrolysis of glucosylceramides by β-glucocerebrosidase.

4. Hydrolysis of galactosylceramides by galactoceramidase.

5. Synthesis from sphinogosine and fatty acid.

6. Dephosphorylation of ceramide 1-phosphate.

Ceramide Synthesis

Production of ceramides is affected by many processes. Exposure to ultraviolet B (UVB) radiation and cytokines has been associated with an increase in the regulatory enzyme for ceramide synthesis, SPT and increased sphingolipid synthesis at the mRNA and protein levels.¹¹ Stress and other causes of increased cortisol and exposure to glucocorticoids affect barrier function, but the mechanism has not yet been delineated.¹² Dexamethasone stimulates ceramide biosynthesis by upregulating gene expression of SPT.¹³ Ceramide production has been shown to be increased by 1,25-dihydroxyvitamin D₃, retinoic acid, ursolic acid, and lactic acid.^14–17 An alkaline pH suppresses β-glucocerebrosidase and acid sphingomyelinase activity (these enzymes need an acidic pH).¹⁸ This is one of the reasons that alkaline soaps can result in poor barrier formation. In addition, a low or neutral pH is associated with poor barrier recovery.¹⁹

Cytokines may also play a role in ceramide synthesis. TNF and interleukin-1 alpha (IL-1α) stimulate ceramide synthesis and SPT levels.²⁰ Imiquimod, a TLR-7 receptor agonist, is known to increase IL-1α levels and has been shown to improve the skin’s barrier. IL-4 inhibits the production of ceramides.²¹

UV radiation has been shown to increase the amount of intracellular ceramides,^22–24 and UV-damaged skin has been reported to exhibit increased barrier function that renders it less susceptible to damage from irritants.²⁵ A study by Kim et al. showed that UV radiation induces ceramide production by activation of sphingomyelinase.²³

Enchanced Ceramide Synthesis Using Precursors

Topical formulations containing linoleic acid such as safflower oil have been shown to increase the formation of Ceramide 1.²⁶ Nicotinamide applied topically has been shown to increase ceramide synthesis and decrease transepidermal water loss (TEWL).²⁷ N-acetyl-L-hydroxyproline topically applied to a synthetic skin model resulted in increased ceramide synthesis through its actions on SPT.²⁸ Topical lactic acid, especially the L-isomer, can increase ceramides.¹⁷

HISTORY

Ceramide 1 was the first natural ceramide identified, in 1982. Synthetic ceramides, studied since the 1950s, are increasingly sophisticated because they are easier to formulate (Table 20-1). Ceramides have come to be known as a complex family of lipids (sphingolipids – a sphingoid base and a fatty acid) involved in cell signaling in addition to their role in barrier homeostasis and water-holding capacity. In fact, they are known to play a critical role in cell proliferation, apoptosis, cell growth, senescence, and cell cycle control differentiation.^29,30

CHEMISTRY

Ceramides can be free lipids but in most cases are found as the hydrophobic backbone of a sphingolipid. Ceramides have been classified as Ceramides 1 to 9, along with two protein-bound ceramides labeled as Ceramides A and B, which are covalently bound to cornified envelope proteins (e.g., involucrin).³¹ (See Figure 20-1.) All epidermal ceramides are produced from a lamellar body-derived glucosylceramide (GC) precursor.

Ceramides are named based on the polarity and composition of the molecule. As suggested above, the foundational ceramide structure is an amide-linked free fatty acid covalently bound to a long-chain amino alcohol sphingoid base.³² The various classes of ceramides are grouped according to the arrangements of sphingosine (S), phytosphingosine (P), or 6-hydroxysphingosine (H) bases, to which an α-hydroxy (A) or nonhydroxy (N) fatty acid is attached, in addition to the presence or absence of a discrete ω-esterified linoleic acid residue.³³

Ceramide 1 is unique insofar as it is nonpolar and contains the fatty acid linoleic acid. The unique structure of Ceramide 1 allows it to act as a molecular rivet, binding the multiple bilayers of the SC together,⁵ and contributing to the stacking of lipid bilayers in the intercellular lamellar sheets. Ceramides 1, 4, and 7 exhibit critical functions in terms of epidermal integrity and by serving as the primary storage areas for linoleic acid, an essential fatty acid with significant roles in the epidermal lipid barrier³⁴ (see Chapter 15, Safflower Oil, and Chapter 19 Barrier Repair Ingredients). The sphingomyelin-derived ceramides (Ceramides 2, 5) are essential for maintaining the integrity of the SC.³⁵

Synthetic ceramides, or pseudoceramides, contain hydroxyl groups, two alkyl groups, and an amide bond – the same key structural components of natural ceramides. Consequently, various synthetic ceramides including N-(3-hexadecyloxy-2-hydroxypropyl)-N-2-hydroxyethylhexadecanamide (PC-104) and N-(2-hydroxyethyl)-2-pentadecanolylhexadecanamide (Bio391) have been reported to form the multilamellar structure observed in the intercellular spaces of the SC.^36,37

ORAL USES

Synthetic ceramides have no effect on skin condition when taken internally because they are broken down by stomach acids. However, oral ingestion of precursors such as linoleic acid may, according to the sparse studies available, help facilitate ceramide production. One study looked at oral administration of glucosylceramides in mice and found that proinflammatory cytokines and scratching were suppressed in an irritant contact dermatitis model.³⁸

TOPICAL USES

Ceramides are popular ingredients in skin care preparations. They have been shown to be able to penetrate into the skin when exogenously applied.^39,40 Several derivatives of ceramides are currently being studied in the dermatology field. Skin conditions such as atopic dermatitis (AD), psoriasis, contact dermatitis, and some genetic disorders have been associated with depleted ceramide levels,⁴¹ but can be ameliorated through the use of exogenous ceramides or their synthetic analogues.

Atopic Dermatitis

AD is known to be a multifactorial disorder recently found to be related to a defect in the filaggrin gene. Prior to the discovery of the role of the filaggrin defect in AD, many studies evaluated the role of barrier lipids in the disease. In 1991, Imokawa reported that subjects with AD had a significantly decreased amount of ceramides in the skin, with Ceramide 1 being the most significantly reduced compared to normal subject skin ceramide content.⁴² In addition, the activities of enzymes that break down ceramides and other important lipids in the SC, particularly ceramidase, sphingomyelin deacylase, and glucosylceramide deacylase, have been shown to be elevated in epidermal AD.⁴¹ In a 2004 study published in the Journal of Investigative Dermatology, AD skin was found to have less sphingomyelinase activity than normal skin.⁴³

Kang et al. conducted studies using synthetic ceramide derivatives of PC-9S (N-Ethanol-2-mirystyl-3-oxo-stearamide), which have been shown to be effective in atopic and psoriatic patients. Both mice studies demonstrated that the topical application of the derivatives K6PC-9 and K6PC-9p resulted in the reduction of skin inflammation and AD symptoms.^44,45 Subsequently, Kang et al. studied the effects of another ceramide derivative, K112PC-5 (2-Acetyl-N-(1,3-dihydroxyisopropyl)-tetradecanamide), on immune cell function, skin inflammation, and AD in mice. Among several findings, the investigators noted that K112PC-5 suppressed AD induced by extracts of dust mites and exhibited in vitro and in vivo anti-inflammatory activity. They concluded that K112PC-5 is another synthetic ceramide derivative with potential as a topical agent for the treatment of AD.⁴⁶

Frankel et al. compared the short-term effectiveness and desirability of a ceramide-hyaluronic acid emollient to pimecrolimus cream 1 percent in treating AD. Both agents displayed efficacy in mild-to-moderate AD across a broad age range of patients over four weeks. Target lesions were deemed “clear” or “almost clear” in 82 percent of the cases using the ceramide foam and in 71 percent of lesions treated with pimecrolimus, with no reported adverse reactions.⁴⁷

In 2011, Kircik et al. conducted a multicenter, open-label, interventional study to assess the clinical efficacy of a topical emulsion containing ceramides, cholesterol, and fatty acids in a 3:1:1 ratio in 207 patients with mild-to-moderate AD over a three-week period. The formulation was used as single therapy or in combination with another AD medication. The investigators reported that about half of participants received clear or almost clear investigator global assessment scores after monotherapy or combination therapy. A majority (75 percent) of patients reported satisfaction with the treatment as pruritus was diminished and quality of life improved. The authors concluded that the ceramide-containing lipid-based formulation was effective in treating mild-to-moderate AD.⁴⁸

That same year, Miller et al. compared the efficacy of three formulations, including a glycyrrhetinic acid-containing barrier repair cream (BRC-Gly, Atopiclair®), a ceramide-dominant, triple-lipid barrier repair cream (BRC-Cer, EpiCeram™),⁴⁹ and a petroleum-based skin protectant moisturizer (OTC-Pet, Aquaphor Healing Ointment®). The study looked at mild-to-moderate AD in 39 children aged 2 to 17 years old randomized 1:1:1 to receive one of the three treatments three times daily for three weeks. Investigators evaluated disease severity and improvement at baseline and days 7 and 21, finding no statistically significant difference in efficacy between the three groups at each time point.⁵⁰

In 2012, Simpson et al. conducted a randomized, intraindividual study to examine the effects of a new moisturizer containing a ceramide precursor on barrier function in patients with controlled AD over a four-week period. In this investigator-blinded study, the cream was applied to one lower leg for 27 days while the other leg served as the untreated control. Researchers took readings at baseline and day 28 on TEWL, skin hydration, and xerosis severity, reporting that after four weeks, significant reductions in TEWL and clinical dryness scores were observed along with increased skin hydration in the areas treated with ceramide cream. In addition, no adverse events were reported.⁵¹ In another study of pseudoceramide formulations and patients with AD, the use of a ceramide complex that also included eucalyptus leaf extract dose-dependently reduced erythema and improved TEWL in comparison to a control vehicle and also increased endogenous SC ceramide levels.⁵²

Diminution of Effects from Steroids

Corticosteroids are well known to cause atrophy of the skin with prolonged use. They also cause a defect in the barrier function of the skin demonstrated by increased TEWL.⁵³ Studies have shown that they disrupt epidermal differentiation resulting in deceased keratohyaline granule formation and a disruption of the lamellar lipid bilayers.⁵⁴ A study by Ahn showed that a pseudoceramide containing multilamellar emulsion (MLE) reduced the atrophy resulting from topical steroid use.⁵⁵ In addition, the MLE cream when applied after the steroid cream prevented the increase in TEWL normally associated with steroid use and preserved the normal structure of the lamellar lipid bilayers.

Dry Non-Atopic Skin

Ceramide deficiency leads to increased TEWL. Subjects that do not have a history of AD or inflammation can still suffer dry skin due to a ceramide deficiency. In the Baumann Skin Typing system, these subjects would be classified as “Dry, Resistant” types, because they exhibit dry skin without underlying inflammation.

Keratinocyte Differentiation

While extracellular ceramides play important roles in skin hydration, intracellular ceramides found in keratinocytes affect differentiation of keratinocytes.^6,56 In 2007, Kwon et al. generated multiple ceramide derivatives and assessed their impact on keratinocyte differentiation. They found that K6PC-4 [N-(2,3-dihydroxypropyl)-2-hexyl-3-oxo-decanamide], K6PC-5, [N-(1,3-dihydroxypropyl-2-hexyl-3-oxo-decanamide] and K6PC-9 (N-ethanol-2-hexyl-3-oxo-decanamide) spurred a fleeting increase in intracellular calcium levels, incited the phosphorylation of p42/44 extracellular signal-regulated kinase and c-Jun N-terminal kinase, and, in the suprabasal layer of a reconstituted epidermis model, significantly enhanced keratin 1 expression. The investigators concluded that synthetic ceramide derivatives exhibit potential for treating skin diseases involving abnormal keratinocyte differentiation.⁵⁷ Ceramides have also been shown to induce phosphorylation of the epidermal growth factor receptor, possibly by activating a protein kinase.⁵⁸

Inflammation

For years, the deficit of ceramides in AD was thought to be the cause of the disorder, leading to many studies examining the effects of ceramides on inflammation. Some studies have shown ceramides to have proinflammatory properties while others emphasize its anti-inflammatory properties.^59,60 For example, in 1996 Di Nardo et al. demonstrated that skin with low ceramide levels was characterized by increased cutaneous inflammation while other studies have shown that ceramides lead to eicosanoid synthesis.^61,62 Sallusto et al. showed that ceramides inhibit the uptake and presentation of antigens by dendritic cells in a murine model.⁶³ Ceramides seem to modulate inflammation through prostaglandins and by modulation of cytokines such as IL-1, and interferon-γ.^64–66 However, the role of ceramides in inflammation is not completely understood. It may be that the amount of ceramide present influences the inflammatory pathways.²¹ Ceramide 1-phosphate may be a more important form of ceramide with respect to its purported proinflammatory activity.^67–69 Ceramide 1-phosphate plays a role in activation of degranulation in mast cells and stimulation of macrophage migration.^70,71

SAFETY ISSUES

Early forms of ceramides were obtained from animal brains and spinal cords such as cows. The fear of mad cow disease led to the development of laboratory-made ceramides; however, these were initially very expensive. New forms derived from wheat germ and other sources provide a more natural option.

Excess amounts of intracellular ceramides can be toxic to cells, inhibiting growth and causing apoptosis.^22,37,72 Synthetic pseudoceramides differ in structure from naturally occurring ceramides, which raises concerns about their safety.³⁷

In 2009, Morita et al. conducted a study evaluating potential adverse effects of the synthetic pseudoceramide SLE66 and showed that the tested product failed to provoke cutaneous irritation or sensitization in animal and human studies. In addition, no phototoxicity or photosensitization was observed and they established 1,000 mg/kg/day (the highest level tested) as the no-observed-adverse-effect (NOAEL) for systemic toxicity after oral administration or topical application.⁷³ In a related rat study on the potential maternal and fetal effects of SLE66, Morita et al. observed no clinical or internal impact from the orally-administered pseudoceramide across multiple metrics, including fetal malformations.²⁹

EpiCeram™ was used in a 2012 study by Lowe et al. to test safety and compliance in a six-week, open-label, phase one trial that found 80 percent of mothers daily applied the cream to their infants (0–4 weeks of age) on 80 percent or more of the days of the study.⁴⁹ The investigators concluded that EpiCeram is sufficiently safe and demonstrated satisfactory parental compliance to prevent eczema.⁷⁴

In light of the uncertainty regarding the metabolic impact of pseudoceramides, in 2008, Uchida et al. compared the effects of two chemically unrelated commercially available products to exogenous cell-permeant or natural ceramide on cell growth and apoptosis thresholds in cultured human keratinocytes and found that commercial ceramides did not suppress keratinocyte growth or increase cell toxicity, as did the cell-permeant. The investigators suggested that these findings buttress the preclinical studies indicating that these pseudoceramides are safe for topical application.³⁷

ENVIRONMENTAL IMPACT

Now that ceramides can be made in the lab and derived from plants such as wheat germ the environmental impact as was seen when these were sourced from animals is considerably blunted.

FORMULATION CONSIDERATIONS

Natural vs. Synthetic

Identical synthetic ceramides are very expensive ($2,000–$10,000/kg).³⁷ Less expensive naturally occurring ceramides are derived from the bovine central nervous system, which raises the concern of transmission of bovine spongiform encephalopathy (also known as mad cow disease). For these reasons, synthetic pseudoceramides are often used in skin care products.³⁷

Ceramides and pseudoceramides are very hydrophobic and tend to crystallize, thus are difficult to formulate.⁷⁵ Organic solvents have been used but tend to dry the skin. Liposomes are often unstable. Oil-in-water emulsions are often used.

Linoleic acid, an important component of ceramides, is very susceptible to oxidation.⁷⁶

USAGE CONSIDERATIONS

Ceramides alone will impair the skin barrier. They must be combined with the proper ratio of cholesterol and fatty acids to improve the skin barrier (see Chapter 19 Barrier Repair Ingredients).

SIGNIFICANT BACKGROUND

Coderch et al., in a review of ceramides and skin function, endorsed the potential of topical therapy for several skin conditions using complete lipid mixtures and some ceramide supplementation, as well as the topical delivery of lipid precursors.⁶ And, in fact, the topical application of synthetic ceramides has been shown to speed up the repair of impaired SC.^77,78 Recent reports by Tokudome et al. also indicate that the application of sphingomyelin-based liposomes effectively augments the levels of various ceramides in cultured human skin models.^79,80 It is important to note that ceramides applied topically without the proper equimolar ratio of cholesterol and fatty acids will actually delay barrier recovery in perturbed skin and amplify TEWL⁸¹ (see Chapter 19 Barrier Repair Ingredients).

In 2005, de Jager et al. used small-angle and wide-angle X-ray diffraction to show that lipid mixtures prepared with well-defined synthetic ceramides are very similar to the lamellar and lateral SC lipid organization and lipid phase behavior and can be used to further elucidate the molecular structure and roles of individual ceramides.⁸² Previously, stress-induced ceramide accumulation had been shown to result in plasma membrane reorganization and development of ceramide-laden structures known as “rafts,” which attract and cluster receptors as well as signaling molecules at the cell membrane to promotesignal transduction cascades.⁹ Interestingly, the pathogenesis of various common disorders has been ascribed to raft-related signaling processes, primarily through ceramide accumulation and activation of apoptotic signaling pathways.⁸³

Effects of Ceramides on Collagen Production and Degradation

Type I collagen levels in the skin have been shown to be important in the appearance of skin.⁸⁴ Antiaging strategies are aimed at: 1) Increasing the amount of collagen produced in the skin and 2) Decreasing the amount of collagen breakdown by collagenase and other matrix metalloproteinases (MMPs). Ceramides have been shown to play a role in both of these processes and, therefore, likely play an important role in photoaging. Kim demonstrated that UV radiation induces ceramide production by activation of sphingomyelinase.²³ The increased ceramide levels led to increased production of MMP-1, which is known to degrade collagen and facilitate photoaging of the skin.⁸⁵ Reunanen showed that ceramide induces collagenase synthesis and that Ceramide 2 inhibits collagen synthesis when added to fibroblast cultures.⁸⁶ It is unknown at this time if including ceramides in topical skin formulations can lead to increased induction of MMP-1 mRNA expression because such studies have not been done. However, one study revealed that when linoleic acid was applied to skin and then irradiated with UV, MMP-1 expression was increased,⁸⁷ which may have been caused by the oxidation of linoleic oxide to linoleic hyperoxide by the UV.³⁹ MMP-1 expression also increased when the synthetic ceramide N-oleoyl-phytosphingosine was applied. However, when cholesterol was applied alone, MMP-1 expression was decreased.³⁹ A small German study (10 human subjects) showed that when jojoba oil and jojoba oil with vitamins were placed on human skin and irradiated with UVA, downregulation of collagen genes resulted.⁸⁸ However, when the same oils were combined with phytosterols and ceramides, less downregulation occurred. It is possible that the phytosterols had a protective effect. Antioxidants (such as phytosterols) that block the ERK1/2 pathway may also help inhibit the downregulation of collagen genes. In the case of Ceramide 2 downregulation of collagen synthesis, the downregulation was dependent upon activation of the ERK1/2 pathway, p38 MAPK pathway, and PKC.⁸⁶ In the author’s opinion, antioxidants may help mitigate the deleterious effects of ceramides on collagen in the skin, especially in the presence of UV radiation.

CONCLUSION

Ceramides are among the primary lipid constituents, along with cholesterol and fatty acids, of the lamellar sheets found in the intercellular spaces of the SC. Together, these lipids maintain the water permeability barrier role of the skin. Ceramides also play an important role in cell signaling. Research over the last 20 years, in particular, indicates that topically applied synthetic ceramide agents, so-called topical ceramide-dominant creams, can effectively compensate for diminished ceramide levels associated with various skin disorders. These products appear to restore barrier integrity and are considered safe and associated with minimal side effects. Ceramides are a crucial component in the skin care regimen of all dry skin types and sensitive skin types (S₄) susceptible to irritant and allergic dermatitis.

REFERENCES

INTRODUCTION

SOURCE

Cholesterol is found in all biologic membranes including keratinocytes, where it is organized into “lipid rafts.”^3,4 Cholesterol sulfate is very abundant in the epidermis with the highest levels found in the granular layer. Amounts of cholesterol sulfate decline progressively from the granular layer to the outer stratum corneum (SC).⁵

Diet

Cholesterol is readily available in the diet and basal keratinocytes are capable of absorbing cholesterol from the circulation (Table 21-1). Cholesterol absorption by keratinocytes is regulated in part by peroxisome proliferator-activated receptors (PPARs) and retinoid X receptors. These have been found to play a role in transporting cholesterol across keratinocyte cell membranes by increasing expression of ATP-binding cassette transporter (ABCA1), a membrane transporter that regulates cholesterol efflux.⁶ Low-density lipoprotein (LDL) receptors may play a role in the absorption of cholesterol by skin cells. Although LDL receptors are not expressed on basal keratinocytes when the barrier is normal, when the barrier is first perturbed, LDL mRNA and protein levels and apolipoprotein E (ApoE) expression rise.^7,8 In chronic barrier disruption, the keratinocytes express LDL receptors.⁹ Increased expression of LDL or ApoE can augment delivery of exogenously applied cholesterol to suprabasal keratinocytes. Most of the cholesterol in skin, however, is not derived from diet but, rather, is synthesized by skin cells.

Endogenous Synthesis

Most cholesterol in the skin is synthesized in keratinocytes by the enzyme cholesterol sulfotransferase type 2B isoform 1b, abbreviated as SULT2B1b (Table 21-2). The rate-limiting enzyme in this process is 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (see Figure 21-1 for the pathway of cholesterol synthesis). A group of transcription factors known as sterol regulatory element-binding proteins (SREBPs) also regulate cholesterol and fatty acid synthesis. When epidermal sterols decline, SREBPs are activated via proteolytic processes. The SREBPs enter the cell nucleus and activate genes leading to increased synthesis of cholesterol.^10,11 There are three known types of SREBPs: SREBP-1a, SREBP-1c, and SREBP-2. In human keratinocytes, SREBP-2 has been shown to be the predominant type and is involved in regulating cholesterol production.¹¹

Cholesterol synthesis does not seem to be influenced by dietary or circulating sterol levels, but it is controlled by alterations in the integrity of the skin’s permeability barrier, and increased in the presence of high amounts of calcium.^12–15 Perturbation of the skin’s barrier results in increases in the quantity and activity of HMG-CoA reductase, resulting in greater cholesterol synthesis.¹⁶ Blocking the activity of HMG-CoA reductase reduces cholesterol synthesis and impedes barrier repair, while the addition of mevalonate increases cholesterol synthesis and restores barrier repair in the absence of HMG-CoA reductase.^12,17

HISTORY

Nonpolar lipids in the SC, including ceramides, cholesterol, and fatty acids, have been known to play a role in regulating transepidermal water loss (TEWL) since the early 1970s.¹⁸ Leaders of the research in this field include Elias, Feingold, Menon, and Ghadially. Their work showed that cholesterol, fatty acids, and ceramides must all be replaced in the skin in an equivalent ratio (1:1:1) in order for barrier repair to occur. Topically placing only cholesterol on the skin leads to delayed barrier repair.

Research in the ensuing years has examined the process by which cholesterol is synthesized and released into the space between skin cells providing a watertight barrier. Ceramides, cholesterol, and fatty acids are synthesized by keratinocytes and packaged in lamellar bodies, which carry them until they are extruded into the intracellular space where they form the “skin barrier” that helps prevent TEWL from the epidermis (see Chapter 19, Barrier Repair Ingredients). When the skin barrier is disturbed by detergents, friction, prolonged water immersion, and other insults, keratinocytes repair it by synthesizing more lipids. Cholesterol plays a crucial role in the early stages of barrier repair as shown in a 1990 study that demonstrated that when cholesterol synthesis is inhibited, lamellar secretion is impaired and lamellar body internal structure is altered, leading to delayed barrier repair.¹⁹ Studies have shown that delivery of circulating sterols does not increase after barrier disruption, implying that locally derived sterol from cholesterol is necessary for the synthesis of lipids in lamellar bodies.^13,20,21 It is now known that in the early stages of barrier repair, cholesterol is essential to contribute to the free sterol that is packaged into lamellar bodies. In fact, the topical application of lovastatin, a competitive inhibitor of HMG-CoA reductase that is known to block cholesterol synthesis, delays barrier recovery.¹²

CHEMISTRY

Cholesterol is a lipid but it differs in structure from other lipids because of its steroid nucleus. A hydrocarbon tail is linked to the steroid nucleus at one end and a hydroxyl group is attached at the other end (Figure 21-2). The hydroxyl group (polar head) in position 3 causes cholesterol to be roughly perpendicular to the bilayer membrane surface. This structure allows cholesterol to be a key regulator of membrane fluidity, plasticity, and rigidity.^22,23

In the biosynthetic pathway of endogenous cholesterol synthesis, the conversion of HMGCoA to mevalonic acid is an early rate-limiting step. Cholesterol-lowering drugs such as lovastatin block this enzyme, leading to a decrease in serum cholesterol but also to a decrease in barrier recovery.¹⁹

ORAL USES

Cholesterol in the diet has been associated with an increased risk of cardiovascular disease, but dietary cholesterol does not necessarily correlate with serum cholesterol levels or increases in LDL, the so-called “bad” cholesterol. This is a complex topic a thorough discussion of which is beyond the scope of this book, however.

TOPICAL USES

Barrier Repair

Topically applied cholesterol can penetrate into the nucleated layers of the SC and be taken up into the cytosol, where its components are broken down and packaged into lamellar bodies.¹² Exogenously applied cholesterol does not downregulate cholesterol synthesis in the skin.²⁴ For these reasons, cholesterol is an important and popular additive to barrier repair creams. Cholesterol is required for the permeability barrier to function normally (see Chapter 19 Barrier Repair Ingredients). Aged skin shows a decreased ability for lipid synthesis, particularly cholesterol synthesis, which leads to a delayed barrier recovery response in older individuals. Ghadially reported that topical application of cholesterol after perturbation of the skin barrier accelerated barrier repair in aged mice skin and, in a different study a year later with Zettersten, was able to accelerate barrier repair using a mixture containing cholesterol/ceramide/palmitate and linoleate at a ratio of 3:1:1:1.^25,26 Topically applied lovastatin, a drug that inhibits HMG-CoA reductase and prevents cholesterol synthesis, delayed barrier recovery in mice by 50 percent. Addition of cholesterol to the skin of these same mice normalized barrier recovery, showing the importance of cholesterol in barrier recovery and function.¹⁹

Keratinization

In order for keratinocytes to develop the permeability barrier, they must first undergo differentiation and maturation, which culminates in the formation of the outermost layer of the skin – the SC. This process is known as keratinization. Keratinization affects the skin barrier and skin moisturization for two main reasons: 1) Keratinocyte maturation allows formation of the lamellar bodies in which cholesterol, phospholipids, and glucosylceramides are synthesized and stored prior to being released into the extracellular space; 2) Keratinocyte maturation leads to the production of structural proteins such as loricrin and involucrin that provide scaffolding needed for the development of lamellar bodies.^5,27,28

Cholesterol plays a crucial role in regulating the keratinization process, through various mechanisms.^29,30 In cell membranes, cholesterol is organized into “lipid rafts,”⁴ which modulate the activation of epidermal stem cells into amplifying keratinocytes by regulating specific enzymes and influencing various signaling processes.^4,29,31 Many pathways affect the keratinization process in the skin. A few that are known to alter keratinization by affecting cholesterol synthesis include:

1. SULT2B1b is the key enzyme in the synthesis of cholesterol sulfate.

2. HMG-CoA reductase is the rate-limiting step in the production of cholesterol.

3. Activation of PPARα, PPAR β/Δ, and PPAR-γ increases cholesterol synthesis.³²

4. SREBPs also regulate cholesterol and fatty acid synthesis.

5. Liver X receptors (LXR) and PPARs regulate the expression of SULT2B1b.^33–36

Cholesterol sulfate is important for cell-to-cell cohesion. The removal of sulfate by steroid sulfatase is required for desquamation of the SC.²³ Alterations in cholesterol sulfate result in impaired epidermal differentiation and corneocyte desquamation.³⁶ For example, X-linked ichthyosis (also known as steroid sulfatase deficiency), a skin disease caused by a deficiency of cholesterol sulfate, is characterized by abnormal corneocyte retention, resulting in rough, dry, “fish-like” skin. Ichthyosis has also been reported in patients on cholesterol-lowering drugs.³⁷

SAFETY ISSUES

There are no studies that demonstrate that topically applied cholesterol raises serum cholesterol levels.

ENVIRONMENTAL IMPACT

None

FORMULATION CONSIDERATIONS

Applying topical cholesterol alone is known to impair the skin barrier. It must be combined with an equimolar amount of ceramides and fatty acids as discussed above and in Chapter 19.³⁸

USAGE CONSIDERATIONS

One intriguing aspect about cholesterol is that it need not be applied to the skin. The skin cells can be prompted to synthesize cholesterol. For example, cytokines such as interleukin-1α and interferon-γ increase cholesterol synthesis.³⁹

SIGNIFICANT BACKGROUND

Cholesterol and Photoaging

Ultraviolet A has been shown to engender peroxidation of cholesterol to cholesterol hydroperoxides (Chol-OOHs),⁴⁰ which can cause damage to cell membranes and become even more deleterious when further reduced to oxylradicals.⁴¹ Oxylradicals are free radicals that lead to a cascade of chain peroxidation reactions, thus elevating matrix metalloproteinase-9 MMP-9 levels,⁴¹ collagenase expression, and other harmful effects (see Chapter 2, Basic Cosmetic Chemistry, and Chapter 46, Antioxidants). Notably, Chol-OOHs have been used as a biomarker of physiological aging in the skin.⁴²

CONCLUSION

Cholesterol is a fascinating skin care ingredient because of its effects on the skin barrier and the keratinization process. Cholesterol may play a role in skin care even when it is not included in the actual skin care product, because other ingredients can influence or spur cholesterol production.

REFERENCES

INTRODUCTION

SOURCE

Lanolin is a greasy yellow substance derived from the sebaceous secretions of sheep and other wool-bearing animals. Most lanolin used in skin care products is obtained from domesticated sheep.

HISTORY

Lanolin has been used for thousands of years by human beings for its emollient qualities, and for hundreds of years as an ingredient in skin care ointments (Table 22-1).³

CHEMISTRY

Lanolin shares two important features with stratum corneum (SC) lipids: 1) lanolin contains cholesterol, an essential constituent of SC lipids, and 2) lanolin and SC lipids can coexist as solids and liquids at physiologic temperatures. Lanolin is characterized by a very different composition than human sebum, though.⁴ This is also the case for commercial lanolin products. Significantly, the method used to refine the compound determines the quality and composition of the resulting formulation; therefore, not all lanolin products exhibit the same activity.⁵ Unfortunately, a small percentage of individuals who use lanolin develop contact sensitization to the occlusive/emollient agent. Consequently, lanolin has developed a reputation as a sensitizer that, according to some, may not be deserved.^3,6 Nevertheless, manufacturers have responded to such claims and many moisturizing products are now touted as “lanolin free.” Another response to the notion that lanolin provokes allergic reactions has spurred the development of an ultrapure hypoallergenic medical grade lanolin formulation such as Medilan™.

ORAL USES

Lanolin is not to be taken internally.

TOPICAL USES

Lanolin is one among several commonly used occlusive agents and is effective as an ingredient in skin care products for its ability to treat dry skin, delivering an emollient effect and reducing TEWL.

SAFETY ISSUES

Lanolin had long been thought of as a sensitizer given reports beginning several decades ago of links to contact dermatitis. However, lanolin is no longer considered a common allergen so much as one affecting compromised or high-risk populations, such as elderly individuals with chronic xerosis, dermatitis, or venous stasis dermatitis.^7–9 In a 2008 study of 276 moisturizers, lanolin, found in 10 percent of products, was the ninth most common allergen, far behind fragrance, parabens, vitamin E, and essential oils and biologic additives.⁹

In 2008, investigators sought to identify the frequency of positive patch test reactions to common allergens in leg ulcer or venous disease patients by using a case series of 100 consecutive consenting subjects suffering from chronic venous disease and other leg ulcer etiologies. At least one positive patch test was observed in 46 percent of patients, with multiple reactions in the same subject frequently seen. Of the 38 common allergens tested, lanolin was identified among the most frequent sensitizers, which also included fragrances, antibacterial agents, and rubber-related compounds.¹⁰ Of course, these results suggest that lanolin may be contraindicated in patients with leg ulcers, but are not generalizable to a healthy population.

ENVIRONMENTAL IMPACT

The impact on the environment of lanolin cultivation is directly related to the effects of sheep farming on land and air quality as well as water run-off. As an animal-derived product, lanolin is not an option for vegan or vegetarian patients.

FORMULATION CONSIDERATIONS

Although a plant-derived substitute has been recently produced, lanolin itself is a complex natural product that cannot be synthesized. It is also a commonly used occlusive ingredient, along with products or compounds such as paraffin, squalene, dimethicone, soybean oil, grapeseed oil, propylene glycol, and beeswax.¹¹ Importantly, though, lanolin, like mineral oil and petrolatum, is known for its dual activity exerting both occlusive and emollient effects.

USAGE CONSIDERATIONS

Lanolin is typically used to moisturize and smooth the skin and is generally considered safe in noncompromised skin. It is easily absorbed through the skin and is thus conducive to delivering medicinal chemicals in an ointment.⁹

SIGNIFICANT BACKGROUND

In 2003, Dodd and Chalmers led a multicenter, prospective, randomized controlled clinical trial to compare the effects of hydrogel dressings to lanolin ointment for the prevention and treatment of nipple soreness in 106 lactating mothers. A board-certified lactation specialist provided breastfeeding guidance at the beginning of the study. During the first 12 days of the study, participants, who were randomized to either of the two groups, were instructed to rate pain intensity according to a numeric pain intensity scale as well as a verbal description scale. Patients then forwarded self-reported skin assessments of the bilateral breasts, nipples, and areolae to investigators. A significantly greater reduction in pain score mean values was identified in the hydrogel group at baseline, day 10, and day 12, as compared to the lanolin group, and the hydrogel group discontinued treatment earlier than the participants using lanolin. Overall, researchers found hydrogel to be superior to lanolin ointment for the management of nipple soreness.¹²

Additional evidence that lanolin is not the optimal therapeutic option for sore or cracked nipples came four years later when investigators conducted a randomized double-blind clinical trial to compare a peppermint gel formulation, modified lanolin, and a placebo control ointment for the treatment of nipple fissures related to breastfeeding. Two hundred and sixteen primiparous mothers were randomly assigned to the three groups, comparable in mean age, and were instructed to apply their selected formulation on both breasts for 14 days. Patients were seen for up to four follow-up visits as well as a final visit at week 6. Researchers found that nipple and areola cracks were less frequent in the peppermint gel group as compared to the lanolin or placebo groups.¹³

Nevertheless, such results do not detract from the appropriateness of lanolin for other dry skin indications. In 2003, investigators conducted a four-week double-blind, randomized-comparison clinical trial to assess the effectiveness of pure lanolin as compared to ammonium lactate 12 percent cream in the treatment of moderate to severe xerosis on the feet. Fifty-one patients, of the original 92 enrolled, completed the study. Both treatment groups exhibited significant improvement in xerosis scores after two and four weeks of treatment, with no statistically significant differences identified between the groups. The researchers concluded that pure lanolin as well as ammonium lactate cream used twice daily for a month were effective in ameliorating moderate to severe dry skin.¹⁴

In 2008, authors reported on a comparison of two different topical ointments used in cutaneous therapy on 173 prospectively enrolled infants between 25 and 36 weeks of gestation admitted to a neonatal intensive care unit from October 2004 and November 2006. Each infant was treated for up to four weeks after being randomly assigned to daily treatment with water-in-oil emollient cream (Bepanthen), olive oil cream (70 percent lanolin, 30 percent olive oil), or a topical control. Skin was evaluated weekly. Investigators found that while both treatment groups exhibited greater improvement than the control group, with enduring treatment effects, infants in the lanolin/olive oil group experienced statistically less dermatitis than the water-in-oil emollient group.¹⁵

CONCLUSION

Through the last several decades, only a small proportion of the population has been found to be allergic to lanolin. Although allergic responses have not been reported in association with the use of more modern, medical grade, or other highly refined lanolin products, the author has seen several patients with lanolin allergy. To patients who demur to use lanolin due to the animal origin of the substance, it is important to at least address this argument by noting that while lanolin is secreted by the sebaceous glands of sheep and then obtained from their shorn wool and refined, the process is conducted without harming the animal. Lanolin, particularly in the newest formulations, is an effective first-line occlusive and emollient agent for various xerotic conditions.

REFERENCES

INTRODUCTION

SOURCE

Stearic acid, a waxlike saturated fatty acid also known as octadecanoic acid, is an important naturally occurring component of stratum corneum (SC) lipids. Besides being synthesized by human beings, stearic acid is also found in butter, cocoa butter, shea butter, vegetable fats, and animal tallow. It is found in many oils but the highest concentrations are in cocoa butter (chocolate), butter, and shea butter. The nonanimal-derived oil with one of the highest amounts of stearic acid is argan oil, which has a concentration of 4 to 7 percent.¹ Stearic acid is found in all cosmetic product categories including soaps, syndet bars, body oils, moisturizers, color cosmetics, and perfumes.

HISTORY

Stearic acid is one of the most common fatty acids found in nature. It is one of the oldest cosmetic ingredients because it is a component of vegetable and animal oils that have been used on the skin for centuries. Historically it has not been used as a singular ingredient, but is found as a component of many popular ingredients such as cocoa butter and various oils.

Stearic acid plays an important role in the skin’s permeability barrier along with linoleic acid and palmitic acid (from palm oil). Palmitic and stearic acids are both saturated fatty acids. Linoleic acid is an unsaturated fatty acid and displays anti-inflammatory properties that stearic acid does not exhibit. Stearic acid has been incorporated into many personal care products because of its safety profile, ubiquity, and inexpensive cost in addition to its surfactant and hydrating abilities. It is found in many “hydrating” body washes and bars that deposit stearic acid on the skin. Stearic acid is found in argan oil, the world’s most expensive oil, at a concentration of 4 to 7 percent.¹

CHEMISTRY

Stearic acid is a saturated long-chain fatty acid with the chemical formula CH₃(CH₂)₁₆CO₂H (see Figure 23-1). The International Union of Pure and Applied Chemistry (IUPAC) name is octadecanoic acid. It is also known as cetylacetic acid and stearophanic acid.

Stearic acid is the saturated form of oleic acid, a monounsaturated ω-9 fatty acid (see Figure 23-2). While stearic acid helps improve the skin’s barrier function, oleic acid impairs it.

ORAL USES

Stearic acid can be safely taken as an ingredient in dietary supplements; however, it is the most poorly absorbed of the fatty acids.²

Stearic acid gives food a desirable taste and texture. It makes up about 9 to 12 percent of total calories in beef, pork, lamb, and veal and about 6 percent of calories in poultry. Although it is a saturated fatty acid, it may be associated with fewer health risks than other saturated fatty acids in the diet. Studies in humans and animals suggest that it has a neutral effect on plasma cholesterol levels in contrast to lauric, myristic, and palmitic acids.³ Stearic acid makes up about 2 to 4 percent of calories in most cooking oils.⁴

TOPICAL USES

As a Food and Drug Administration (FDA)-approved ingredient in several cosmetic products, it is used as a surfactant and emulsifying agent, for fragrance, and as the base for other fatty acid ingredients that are synthesized into emollients and lubricants (Table 23-1). Specifically, it is used most often to retain the shape of, and as a thickener in, soaps (indirectly, through saponification of triglycerides composed of stearic acid esters) as well as in shampoos, shaving creams, and detergents.

SAFETY ISSUES

Stearic acid is approved by the FDA for use in topical skin care formulations and is generally considered safe. Evaluation by the Cosmetic Ingredient Review (CIR) showed no phototoxicity, minimal to no skin irritation, no tumorigenicity, and no comedogenicity. Stearic acid was labeled as “safe” by the CIR.

ENVIRONMENTAL IMPACT

Stearic acid is found abundantly in animals and plants, more so in the fat of animals. Individuals concerned about harming animals are advised to delve more deeply into the origins of the stearic acid used in products that they might consider. A significant environmental impact is not exerted in culling and processing stearic acid.

FORMULATION CONSIDERATIONS

Stearic acid is used to thicken and stabilize formulations. Its high amount of saturation makes it more solid at room temperature and gives it a waxy consistency. The acid can be neutralized with triethanolamine and other alkalis to form soap (sodium stearate), which acts as an emulsifier.⁵

USAGE CONSIDERATIONS

Saturated fats such as stearic acid are more stable than unsaturated fatty acids. This is important because oxygen and heat can degrade fatty acids, rendering them rancid. Combination with antioxidants can slow the degradation process. Notably, stearic acid may decrease penetration of other ingredients by strengthening the skin barrier.

SIGNIFICANT BACKGROUND

In 2000, Khalil et al. studied the effects of cream formulations on chemically-induced burns in mice based on reports that the ingredients, docosanol or stearic acid, were associated with antiviral and anti-inflammatory activity. Burns were engendered by painting murine abdomens with a chloroform solution of phenol. Researchers then topically applied the test formulations 0.5, 3, and 6 hours after injury. They found that the docosanol- and stearic acid-containing creams significantly lessened the severity and progression of skin lesions compared to untreated sites, yielding, respectively, 76 and 57 percent declines in mean lesion scores.⁶

In 2001, Fluhr et al., in a study of the effects of the free fatty acid pool on SC acidification and function, topically applied two phospholipase inhibitors, bromphenacylbromide and 1-hexadecyl-3-trifluoroethylglycero-sn-2-phosphomethanol, for three days to murine skin. This raised skin pH and yielded permeability barrier abnormality, altered SC integrity, and reduced SC cohesion. All malfunctions were normalized, including SC pH, with the coapplication of either palmitic, stearic, or linoleic acids along with the inhibiting agents.⁷

In 2010, Mukherjee et al. evaluated a recently-marketed mild moisturizing body wash, with stearic acid and emollient soybean oil, to determine the location and amount of stearic acid deposited in the SC after in vivo usage of the product. They conducted clinical cleansing studies using the soybean product or petroleum jelly. The deuterated variant of stearic acid replaced the free stearic acid in the soybean formulation. The investigators detected deuterated stearic acid in all 10 consecutive layers of SC, with a total stearic acid level measured at 0.33 μg/cm² after five washes with the soybean oil product. They concluded that the estimated total fatty acid delivered to the skin from cleansing, probably incorporated into the SC lipid phase, is comparable to the fatty acid level in an SC layer.⁸

CONCLUSION

Stearic acid is an important component in stratum corneum lipids, but it is rarely used alone. Therefore, there is a paucity of research to consider its individual attributes. Most studies use stearic acid-containing substances, such as argan oil or shea butter, that are not purely stearic acid so other components also play a role in the results seen. Although it is difficult to tease out the individual role of stearic acid, it seems certain that its presence affects skin penetration and barrier function and is a superior moisturizing ingredient when combined in the proper ratio with cholesterol and ceramides (see Chapter 19, Barrier Repair Ingredients). Much more research is necessary, then, to determine just how significant stearic acid is as a therapeutic agent.

REFERENCES

INTRODUCTION

Humectants are water-soluble materials with high water absorption capabilities. They are hygroscopic and therefore able to attract water from the atmosphere (if atmospheric humidity is greater than 80 percent) and from the underlying epidermis. Although humectants may draw water from the environment to help hydrate the skin, in low-humidity conditions they may take water from the deeper epidermis and dermis, resulting in increased skin dryness.¹ For this reason, they work better when combined with occlusives. Humectants are also popular additives to cosmetic moisturizers for many reasons. They prevent product evaporation and thickening, which increases the shelf life of formulations, and some humectants help prevent bacterial growth in products.² Humectants can cause an almost immediate improvement in skin texture because they draw water into the skin, causing a slight swelling of the stratum corneum (SC) that gives the perception of smoother skin with fewer wrinkles. Humectants, by inducing swelling, can temporarily give the user the perception of increased skin firmness. Humectants have been shown to enhance the penetration of other ingredients by causing swelling of keratinocytes,³ and disruption of the skin barrier by loosening the closely packed SC cells.⁴ Propylene glycol enhances penetration of minoxidil and steroids,⁵ while hyaluronic acid increases drug delivery in prescription medications such as Diclofenac.⁴ Examples of commonly used humectants include glycerin, sorbitol, sodium hyaluronate (hyaluronic acid), urea, propylene glycol, α-hydroxy acids, and sugars.

REFERENCES

INTRODUCTION

SOURCE

Glycerol provides the molecular skeleton of all animal and vegetable fats known as triglycerides. It is derived from the saponification of fats. Throughout this chapter, the terms “glycerol” and “glycerin” (also spelled as glycerine and referred to more often in the literature as glycerol) will be used interchangeably; glycerin is the designation most familiar to consumers (and the one used here more often in relation to topical products). Glycerol is a potent, nonvolatile trihydroxylated humectant. When the body consumes fat stores (triglycerides) to produce energy, glycerol and fatty acids are released into the bloodstream. The glycerol component is converted to glucose in the liver, thus providing energy for cellular metabolism. Natural glycerol is obtained hydrolytically from fats and oils during soap and fatty acid manufacturing, and by transesterification (an interchange of fatty acid groups with another alcohol) during the production of biodiesel fuel.¹ Synthetic glycerol refers to material obtained from nontriglyceride sources.

Glycerin exhibits hygroscopic ability very similar to that associated with natural moisturizing factor (NMF).² NMF is found within keratinocytes in the stratum corneum (SC) and is composed of amino acids, lactate, urea, citrate, and sugars. It can absorb large quantities of water (i.e., hygroscopic) even when humidity levels are low. This allows the SC to maintain a sufficient hydration level even in dry environments. Numerous ingredients have been used in moisturizing products to mimic NMF activity; glycerin is one of the primary ones.

The topical application of glycerin is considered a useful component in treatment regimens for various cutaneous conditions, including bedsores, bites, burns, calluses, cuts, rashes and, occasionally, psoriasis. It is well known to protect against cutaneous irritation.

HISTORY

The name glycerol is derived from the Greek word glyko, meaning “sweet.” Carl Wilhelm Scheele discovered glycerol in 1779. Since then it has been included and used widely in topical skin cosmetics, with approximately 160,000 tons of glycerol sold annually in the United States alone.^3,4 Glycerol is one of the most versatile and valuable chemical substances known and is considered the most effective humectant ingredient in the personal care industry (Table 25-1).^5,6 It has long been used as an active as well as excipient ingredient in skin care products. Procter and Gamble (P&G) started producing glycerin around 1858, about the time they started producing consumer products. Today, P&G Chemicals is one of the largest manufacturers of glycerin in the world.

CHEMISTRY

Glycerol contains three hydrophilic alcoholic hydroxyl groups, which are responsible for its solubility in water and its hygroscopic nature (Figure 25-1). It is a highly flexible molecule forming both intra- and intermolecular hydrogen bonds. There are 126 possible conformers of glycerol.⁷

ORAL USES

Glycerin has been shown to display some antibacterial properties and is useful in treating halitosis. It is commonly used as an artificial sweetener. Its oral use has no known role in skin care.

TOPICAL USES

Glycerin, which helps condition the skin, is the most widely used hydrating agent and, consequently, a common ingredient in skin cleansers, creams, and lotions. In addition to its well-established hygroscopic activity and solubility similar to water, glycerin is characterized by the ability to prevent freezing and prolong the shelf life of a product.³ Glycerin is one of the best moisturizing ingredients because of its strong humectant (hygroscopic) properties. It is often combined with occlusive agents to impart a synergistic improvement in dry skin disorders.^6,8 Glycerin may also play a role in increasing desquamation because it facilitates corneocyte maturation by spurring the activity of residual transglutaminase in the SC.^6,9

In addition to its humectant properties, glycerol is a unique ingredient because it is able to transverse between keratinocytes via aquaporin-3 (AQP-3) channels, which allows the movement from cell to cell¹⁰ (see Chapter 29, Aquaporin).

SAFETY ISSUES

Glycerin has a long track record of safety with no risk of allergic reactions. Some Internet sites state that glycerin will dry out the skin; however, this occurs when it is used in a dry environment without an occlusive ingredient, a setting in which the glycerin pulls water fom the skin. When glycerin is formulated properly it gives the skin sustained moisturization.¹¹

ENVIRONMENTAL IMPACT

The synthesis of glyercin poses no environmental risk. It is naturally occurring in plants and animals.

FORMULATION CONSIDERATIONS

Glycerin is soluble in water. Its versatility makes it easy to formulate.

USAGE CONSIDERATIONS

Glycerin can be used in conjunction with other ingredients. It is known to enhance the skin barrier but has also been shown to enhance skin penetration of some other ingredients.¹²

SIGNIFICANT BACKGROUND

In Vitro Studies

Xerosis, or dry skin, is associated with incomplete desmosome degradation. In a 1995 in vitro study of moisturizers facilitating the process of desmosome digestion, investigators observed via electron microscopy that desmosomes treated with glycerol were in more advanced stages of degradation than control tissue. In two other in vitro models evaluated by the same team, glycerol raised the corneocyte loss rate from the superficial surface of human skin biopsies and significantly lowered intercorneocyte forces.¹³ Since then, it has been established that by inducing activity of residual transglutaminase in the SC, glycerol accelerates corneocyte maturation.^6,9 In addition, glycerol diminishes xerotic scaling by contributing to desmosome digestion and then enhancing desquamation.^6,13 These studies show that glycerol can function as an aid to skin exfoliation.

Animal Studies

In a 2003 study in which mice deficient in the epidermal water/glycerol transporter AQP-3 exhibited diminished SC hydration and skin elasticity, as well as poor barrier recovery after SC removal, investigators showed that glycerol replacement ameliorated each defect in AQP-3-null mice. Notably, SC water content, which was measured as threefold lower in AQP-3-null mice than wild-type mice, was restored via topical or systemic glycerol administration, but administration of glycerol-like osmolytes (e.g., xylitol, erythritol, and propanediol) was unsuccessful. In addition to concluding that glycerol is an important determinant of SC water retention, investigators suggested that their data provide a strong scientific foundation for the centuries-old practice of including glycerol in skin formulations for medicinal as well as cosmetic purposes.⁴ Further, the authors noted that when added to SC lipids in vitro, glycerol is thought to combine with lipid lamellae and foster water absorption and hinder the conversion of lipid lamellar structures from liquid to solid crystal, thereby inhibiting or preventing water loss.^4,14 This hypothesis, the data from this study, and the fact that in three days pure glycerol absorbs its own weight in water,¹⁵ led the authors to speculate that glycerol likely improves SC water absorption and retention.⁴

Also in 2003, a study of asebia mice with profound sebaceous gland hypoplasia revealed that the topical application of glycerol, which, endogenously, is believed to be the result of triglyceride hydrolysis in sebaceous glands, restored normal SC hydration whereas the endogenous humectant urea did not. Further, the investigators showed that glycerol from triglycerides in sebaceous glands contributes significantly to SC hydration.¹⁶

Human Studies

In a five-year study in which two high-glycerin moisturizers were compared to 16 other popular moisturizers in 394 patients with severe xerosis, the high-glycerin products were found to perform better than all other products tested throughout the period by more rapidly restoring normal hydration and preventing the resumption of dryness for a longer period, longer even than petrolatum.¹⁷ According to ultrastructural analysis of skin treated with high-glycerin formulations, glycerin expands the space between layers of corneocytes and the thickness of corneocytes, which results in the expansion of the SC.¹⁸ These findings indicate that glycerin gives the skin the capacity to hold a reservoir of moisture that renders it more resistant to drying. In addition, glycerin stabilizes and fluidizes cell membranes as well as hydrates enzymes required for desmosome degradation.¹⁷

In 1999, Fluhr et al. conducted two studies to examine the capacity of glycerol as a barrier stabilizer and moisturizing compound. In the first study, barrier repair was found to be more rapid in glycerol-treated sites; significant differences were noted three days after treatment between glycerol open vs. untreated and glycerol occluded vs. untreated, and SC hydration was superior in the glycerol plus occlusion sites. In the second study, barrier repair was again more rapid in areas treated by glycerol, with significant differences compared to untreated and base-treated areas at day 7, and after three days of treatment, SC hydration was superior in areas treated by glycerol. The authors concluded that glycerol promotes barrier repair, notably after acute exogenous disturbance, and enhances SC hydration.²

In a study on dermatologic vehicles and their effect on the horny layer, investigators found that adding glycerol to oil-in-water (O/W) emulsions eliminated the barrier-damaging effect of such formulations. Glycerol added to O/W emulsions also decreased horny layer damage in stress tests with wash solutions. O/W emulsions that contain glycerol are also appropriate in atopic dermatitis therapy.¹⁹

In 2005, investigators found that endogenous glycerol from circulation into the epidermis via AQP-3 and from triglyceride hydrolysis in sebaceous glands is associated with SC hydration in humans. Indeed, they observed glycerol from both sources forms a water reservoir that affects such hydration. The authors also noted that other findings in their study support the use of therapeutic moisturizers that contain glycerol.²⁰

Also, in 2008, a small pilot study with nine healthy women supported the “proof of concept” that hydrolyzed jojoba esters combined with glycerol produces an additive effect, enhancing moisturization for at least 24 hours.²¹

In a 2010 study of the effects of glycerol on human skin impaired by acute irritation induced by sodium lauryl sulfate (SLS), Atrux-Tallau et al. found that glycerol seems to serve the same function as natural moisturizing factors eliminated by the detergent action of SLS. This leads to improving skin hydration. The investigators concluded that the inclusion of glycerol in topical formulations intended to treat irritated skin is warranted.²²

CONCLUSION

Glycerin is an important ingredient in skin care products, cosmetics, and cosmeceuticals because of its efficacy, safety, low cost, long history of use, and pervasiveness in the skin care product market. Recent research has indicated that glycerin displays several mechanisms of action and its efficacy depends on the choice of vehicle and emulsifying agent.

REFERENCES

INTRODUCTION

SOURCE

Hyaluronic acid (HA), or hyaluronan, is the most abundant glycosaminoglycan (GAG) found in the human dermis (Table 26-1). GAGs are polysaccharide chains made up of repeating disaccharide units linked to a core protein. Together the GAGs (HA, dermatan sulfate, heparin, heparin sulfate, keratin sulfate, chondroitin-4, and chondroitin-6-sulfate) and attached core proteins form proteoglycans. The only nonsulfated GAG and the only one not synthesized on a core protein, HA is produced by an enzyme complex of the plasma membrane.¹ In addition, HA is a hygroscopic sugar that can bind over 1,000 times its weight in water. It is responsible for giving skin its plumpness and volume. HA is made by fibroblasts and broken down by the enzyme hyaluronidase.

The HA used in skin care products and injectables was originally harvested from rooster combs but now most HA in skin care products is derived from a bacterial origin and produced in the laboratory setting. The molecular weight of the HA varies according to its source and chain length. The HA isolation process can be adjusted to determine its corresponding molecular weight, altering its physiochemical properties.

Uncrosslinked chains of HA in the skin are broken down by hyaluronidase and free radicals in approximately 24 to 36 hours. Chemical modifications, such as crosslinking HA chains with 1,4-butanediol diglycidyl ether (BDDE), can increase the amount of time that HA resides in the skin. Dermal fillers such as Restylane are crosslinked with BDDE so that the HA lasts in the skin for six or more months; however, these crosslinked HA chains are too large to penetrate into the skin and must be injected.² Topical forms of HA must have a small enough molecular weight to pass into the skin, thus obviating the use of crosslinked HA.

HISTORY

HA was discovered in bovine vitreous humor by Meyer and Palmer in 1934.³ It was named based on its glassy appearance (the Greek word for glass is hyalos) and the presence of a sugar known as uronic acid. HA, which appears freely in the dermis and is more concentrated in areas where cells are less densely packed, is an important dermal component responsible for attracting water and giving the dermis its volume. The popularity of HA fillers for injection into the dermis to correct wrinkles emerged in the 1990s and available products now include the Juvéderm line, Belotero, Restylane, Perlane, and Voluma. HA is also a popular ingredient in cosmetic products because of its humectant activity. However, traditionally, the HA found in many moisturizers, because of its large size, could not penetrate the epidermis and enter the dermis when topically applied, despite the claims of manufacturers.⁴ Conflicting reports claim that smaller sizes of HA may penetrate into the epidermis when used in the proper formulations. In the author’s knowledge, there are no published studies demonstrating the penetration of topical HA into the dermis.

CHEMISTRY

HA is a linear, naturally occurring polyanionic polysaccharide that consists of repeating disaccharide units of N-acetyl-D-glucosamine and β-glucuronic acid (D-glucuronate).^5,6

Present in most biologic fluids and tissues (notably, most vertebrate connective tissue), it is most abundant, and an important component, in the extracellular matrix of soft connective tissues, particularly skin, where it plays a protective, shock-absorbing role.^5,6 Although it can be derived from humans, animals, or bacterial cultures, the structure of HA is identical among all of these species. HA exhibits key functions in cell growth and signaling, membrane receptor function, and adhesion, as well as wound repair and regeneration, morphogenesis, matrix organization, and pathobiology.⁶ In young skin, HA is present at the periphery of collagen and elastin fibers and where these fibers interface. Such connections with HA are absent in aged skin.⁷ In addition, HA appears to play a role in keratinocyte differentiation and lamellar body formation through its interaction with CD44,⁸ a cell surface glycoprotein receptor with HA binding sites.^9–11 It is also thought to foster neutrophil migration, fibroblast proliferation, and neoangiogenesis.¹² In 2000, Sakai et al. used high performance liquid chromatography to show that HA is delivered by keratinocytes and is present in the normal stratum corneum (SC) of mice. Further, they speculated that HA contributes to moisturizing the SC and/or regulating its mechanical properties.¹³

ORAL USES

HA is available in oral supplement form, but the stomach breaks it down rendering it worthless to the skin when taken in an oral form. Glucosamine supplements, however, may help skin increase its production of HA.

TOPICAL USES

HA is the main ingredient in the barrier repair products Bionect and Hylatopic to impart water retention. Bionect contains 0.2 percent HA sodium salt, which acts as a humectant, with other humectant ingredients (e.g., glycerin) and a 70 percent sorbitol solution. Hylatopic combines HA sodium salt with the humectant glycerin in a foam preparation along with occlusive ingredients such as dimethicone and petrolatum.¹⁴ Notably, the Food and Drug Administration (FDA) has approved the barrier repair products Atopiclair and Hylatopic, in which HA is the main ingredient.¹⁴ Topical medications such as 3 percent Diclofenac have HA as an additive in the formulation.⁵

The humectant properties of HA allow it to bind 1,000 times its weight in water. Topical use of HA has been shown to improve skin hydration.¹⁵ It functions as a skin-hydrating agent best when used in a humid environment. In a dry environment, it can draw water from the skin into the HA, thus dehydrating the skin. For this reason, HA should be used in combination with an occlusive ingredient in a dry environment. Individuals with oily skin types have sufficient sebum to impart occlusive properties. Therefore, those with oily skin types can use HA in any environment but people with dry skin should not use it in a dry environment unless the HA is combined with an occlusive agent. Applying HA to the skin in a humid environment attracts water to the skin and can result in immediate improvement of wrinkles. The effect is lessened in a low-humidity environment because there is less water in the atmosphere to draw to the skin. In one study, subjects saw improvement within minutes after using a face product with HA.¹⁶

SAFETY ISSUES

Studies that considered HA for drug delivery have shown that it can facilitate drug penetration into the epidermis and prevent it from entering the dermis as well as the blood stream.⁵

ENVIRONMENTAL IMPACT

No significant environmental impact is likely due to the production of topical HA products.

FORMULATION CONSIDERATIONS

Although HA is a very important skin component, its topical utility is thwarted by its large size and inability to pass into the dermis. However, if the proper size of HA is used, it can penetrate into the epidermis causing a rapid, if short-lived, improvement of fine lines through its effects on skin hydration. It is important to realize that the process of isolating and chemically processing HA greatly impacts its biological activity so that “not all HA is created equal.”

USAGE CONSIDERATIONS

HA can enhance drug penetration, an important consideration when designing a skin care regimen and designating the order in which products will be applied. For example, applying a retinol after using a product with HA could theoretically increase absorption of retinol.¹⁷

SIGNIFICANT BACKGROUND

HA, in large part due to its viscoelastic nature, biocompatibility, and nonimmunogenicity, is included in multiple clinical applications, including as an oral supplement and injectable to increase HA in synovial fluid in arthritic patients, an eye aid after cataract surgery, a wound-healing agent, a filling agent in cosmetic and soft tissue surgery, a device in various surgical procedures, and in tissue engineering.^5,6 Topically it has become a popular additive in skin care products with claims ranging from decreasing eye puffiness to smoothing wrinkles and firming skin.

Wound Healing

In a small prospective study of 27 patients delivering by cesarean section and 20 patients delivering vaginally with episiotomy, researchers assessed the effects of an HA sodium salt (Bionect) in 15 subjects from the cesarean group and 10 from the episiotomy group (with standard wound care applied to the remaining patients) and found that daily application of the HA treatment yielded a lower incidence of edema, infiltration, and wound exudates compared to standard treatment. One case of wound dehiscence in each standard treatment group occurred, but none in the HA groups.¹²

In a recent 60-day, double-blind, randomized, controlled superiority trial intended to examine the efficacy and safety of a gauze pad containing HA in local treatment of venous leg ulcers in 89 patients, Humbert et al. found that ulcer surface was diminished significantly in the HA group compared to the neutral control group at day 45. In addition, at days 45 and 60, the number of healed ulcers was significantly higher in the HA treatment group. Notably, pain intensity was significantly lower in the HA group at day 30.¹⁸

Topical Antiaging Activity

In 2011, Pavicic et al. studied the efficacy of topically applied 0.1 percent HA formulations of various molecular weights (50, 130, 300, 800, and 2,000 kDa) in the periocular area to treat wrinkles in 76 randomized females ranging in age from 30 to 60 years. Subjects applied one of the preparations twice daily around one eye and a vehicle control cream around the other eye. Investigators measured skin hydration and elasticity at baseline as well as 30 and 60 days posttreatment. They observed significant improvement in skin hydration and elasticity in association with all of the HA-based creams in comparison to placebo. Wrinkle depth measurements indicated significant improvement in the periocular areas treated with 50 and 130 kDA HA as compared to areas treated with placebo. The researchers concluded that lower molecular weight HA formulations were linked to greater wrinkle depth reduction due likely to deeper penetration capacity.¹⁵ This study controlled room humidity at 50 percent during wrinkle assessment, which is an important aspect to consider when using HA for antiaging.

Anti-inflammatory Activity

In 2012, Schlesinger et al. conducted, in an outpatient setting, a small prospective, observational, nonblinded safety and efficacy study of a topical anti-inflammatory formulation containing low-molecular weight HA (HA sodium salt gel 0.2 percent) to treat facial seborrheic dermatitis in 15 subjects ranging in age from 18 to 75 years. The hypothesis was that low-molecular weight HA would have an anti-inflammatory effect because of its actions on the immune system and cytokine formation.¹⁹ Although immune system effects and cytokines were not measured in this study, interim data for seven of the 15 patients showed that HA treatment resulted in improvement of all measured endpoints, including erythema, pruritus, and scaliness.²⁰ This is the only published study looking at the use of HA in inflammatory skin disorders. Most of the literature on this topic is in the treatment of osteoarthritis.

Combination Therapy

HA has been shown to be effective in several combination products. In 2010, Guevara et al. evaluated the safety and efficacy of a new cream for treating melasma that contains hydroquinone, glycolic acid, and HA. In this small, open, uncontrolled 12-week study, 15 Latin American women applied the formulation to both sides of their face twice daily. Fourteen of the patients improved, with a significant reduction in melasma area and severity index (MASI) scores of 64 percent. After eight weeks of treatment, 53 percent of the subjects needed to use a moisturizer. Most reported adverse events were mild.²¹

In 2011, Cordero et al. conducted a three-month, open, multicentered, international study of 1,462 subjects to evaluate a cream combining retinaldehyde and HA fragments for treating photoaging. Participants were instructed to daily apply either retinaldehyde 0.05 percent-HA fragments 0.5 percent (Eluage® cream), retinaldehyde 0.05 percent-HA fragments 1 percent (Eluage® antiwrinkle concentrate) or both products. Dermatologists evaluated photoaging severity at baseline as well as days 30 and 90 using Larnier’s scale. Significant improvement was observed by the investigators, with reduction in Larnier scores in all three groups. Wrinkles (i.e., on the forehead, nasolabial folds, crow’s feet, and perioral) were significantly ameliorated in subjects using Eluage antiwrinkle concentrate as well as both formulations. Both products were well tolerated by subjects and clinical signs of photoaging diminished significantly in the Eluage cream and the group using both products in terms of elasticity, hyperpigmentation, and ptosis.²²

In 2011, Frankel et al. conducted a bilateral comparison investigation of pimecrolimus cream 1 percent and a ceramide-HA emollient foam for the treatment of mild-to-moderate atopic dermatitis, and reported that both products displayed efficacy. After four weeks of treatment, 82 percent of target lesions treated with the ceramide-HA foam were scored “clear” or “almost clear” compared to 71 percent of lesions treated with pimecrolimus.²³

Later that year, Gazzabin et al. evaluated a spray product combining colloidal silver and HA in 54 patients, 30 with chronic wounds and 24 with superficial traumatic wounds. Patients were instructed to apply treatments at intervals of three to seven days based on lesion traits. Investigators found that chronic lesions healed by week 12 (70 percent closure rate at six weeks) and traumatic wounds healed at six weeks (80 percent substantial closure at three weeks). They also noted satisfactory microbial contamination control in treated ulcers and concluded that the HA and silver combination spray was effective in spurring re-epithelialization of superficial cutaneous wounds of various origins.²⁴

In 2012, Joksimovic et al. conducted a small double-blind, placebo-controlled clinical trial with 36 hemorrhoid patients to assess the efficacy and tolerability of a gel medical device containing HA, tea tree oil, and methyl-sulfonyl-methane. Findings from the 14-day treatment regimen showed statistically significant reductions in all symptoms as compared to placebo. The HA-containing gel was also found to be safe and tolerable in this small sample.²⁵ HA is also being used in combination with an iodine complex to achieve measurable clinical benefits in various wound types.²⁶

CONCLUSION

HA is a ubiquitous and versatile component of the extracellular matrix and a key constituent of the dermis, contributing significantly to skin hydration, skin volume, and a youthful appearance. HA is known to diminish as one ages. Aside from its role in popular dermal filling agents, HA has also been used in exogenous topical products as a moisturizing agent. Its utility in a topical formulation is determined by several factors including the ambient humidity, the molecular weight of the HA, and what chemical alterations have been made to the HA molecule. Topically delivered HA may not be able to penetrate into the skin at all due to its large size. If it does penetrate, it remains localized in the epidermis forming reservoirs,²⁷ and does not penetrate into the dermis. This makes it a popular ingredient to enhance drug delivery to the epidermis but impairs its ability to add volume to a photoaged dermis like an injectable form of HA would do. For this reason, much of the marketing of HA in skin care products is misleading. Topically delivered HA should be considered as a humectant skin moisturizer when used in a normal or humid environment. Any antiaging properties it demonstrates would be based on skin hydration alone and the effects are temporary. Injectable HA is a different topical agent altogether and is covered thoroughly in Chapter 2, Basic Science of the Dermis in Cosmetic Dermatology: Principles and Practice, 2nd ed. (McGraw-Hill, 2009).

REFERENCES

INTRODUCTION

SOURCE

Present in all living cells, pantothenic acid is a precursor for the production of acetyl coenzyme A, an essential substrate for acetylcholine synthesis. Pantothenic acid, or vitamin B₅, belongs to the water-soluble B vitamin family and is also an essential ingredient in the enzymes necessary for metabolizing carbohydrates and fats. Proper growth and development depends partly on this vitamin as does the maintenance of normal epithelial function, including skin regrowth.¹ Among the best sources of pantothenic acid (PA) are fish, beef, whole grains, dairy, eggs, mushrooms, peanuts and other legumes, cashews, broccoli, soybeans, and avocados, but most whole foods contain some PA. In fact, deficiency of PA is virtually unknown because of its broad availability in food sources.

HISTORY

The word “pantothenic” is derived from the Greed word pantos (“everywhere”), suggesting the omnipresence of the vitamin in the plant and animal world. Dexpanthenol (provitamin B₅) is the stable alcohol form of pantothenic acid. It is popular and well regarded in the treatment of various skin conditions.²

CHEMISTRY

Not found naturally, synthetic dexpanthenol is converted to PA in the skin, stimulating skin regeneration in a fashion comparable to vitamin A. This process of cell division and formation of new skin tissue restores skin elasticity and promotes wound healing. In fact, both in vitro and in vivo, dexpanthenol has been shown to promote fibroblast proliferation.¹ Therefore, water-soluble dexpanthenol has been used topically to foster wound healing. Formulations containing dexpanthenol have exhibited a capacity to stimulate epithelialization and granulation while imparting an antipruritic, anti-inflammatory effect on experimental ultraviolet-induced erythema.¹ In an early study, treatment with dexpanthenol over a three- to four-week period resulted in significant improvement in skin-irritation symptoms such as xerosis, pruritus, erythema, roughness, scaling, and fissures.¹ Several other benefits to the skin have been associated with provitamin B₅. In a randomized, double-blind, placebo-controlled study, treatment with topical dexpanthenol over a seven-day period resulted in enhanced stratum corneum (SC) hydration and decreased TEWL.³

Dexpanthenol is used in a wide range of cosmetic products, typically to moisturize the skin, and formulated in some intramuscular and intravenous products. Topically applied provitamin B₅ also acts to prevent TEWL while moisturizing the skin. It is well tolerated and poses minimal risk of irritation or sensitivity.

ORAL USES

Vitamin B₅ is available in various foods. Dexpanthenol can also be taken orally, and is metabolized into pantothenic acid.

TOPICAL USES

Topical dexpanthenol is well established as an effective moisturizer, enhancing SC hydration and decreasing TEWL in healthy skin, protecting against irritation as a pretreatment, and facilitating wound healing (Table 27-1).¹

Human Studies

Proksch and Nissen, in 2002, conducted a study with 20 subjects (12 women, 8 men) to identify the effects of a dexpanthenol-containing cream, applied twice daily, on skin barrier repair, SC hydration, skin roughness, and inflammation after irritation induced by sodium lauryl sulfate (SLS). The investigators found that the dexpanthenol-containing cream significantly increased the pace of skin barrier repair compared with placebo or untreated skin. SC hydration was enhanced and skin roughness reduced by both the dexpanthenol cream and placebo, but significantly more by the cream containing dexpanthenol. The test cream also significantly diminished erythema, whereas the placebo had no impact on skin redness.⁴ A different study in 2002, a prospective, randomized, double-blind trial, demonstrated that the prophylactic and consistent use of an emollient containing dexpanthenol ameliorated the effects of radiation dermatitis, though it was less effective than topical corticosteroid (0.1 percent methylprednisolone).⁵

In 2003, Biro et al. conducted a prospective, randomized, double-blind, placebo-controlled study of 25 healthy volunteers (21 of whom completed the trial) to study the efficacy of dexpanthenol in protecting against skin irritation. For 26 days, the patients (between 18 and 45 years old) twice daily applied either Bepanthol Handbalsam containing 5 percent dexpanthenol or placebo to the inner part of both forearms. Twice daily from days 15 to 22, SLS 2 percent was applied. The investigators reported that intraindividual comparisons revealed better results in areas treated with dexpanthenol in 11 cases as compared to one placebo site. Irritant contact dermatitis was reported by six patients, with greater severity associated with the placebo site in five of the cases.²

In 2009, Radtke et al. conducted a prospective observational study in 392 networked pharmacies, consecutively recruiting 1,886 patients with irritated skin to determine the patient-relevant benefit of dexpanthenol treatment. They found that 94.7 percent achieved successful results and all symptoms of irritated skin improved significantly independent of age, gender, and underlying cutaneous disease.⁶

Dexpanthenol is firmly established enough as a therapeutic option for irritant dermatitis and cutaneous inflammation to have allowed Wolff and Kieser to use it as a comparison standard in an observational study to gauge the effectiveness of hamamelis (better known as witch hazel) for children with minor skin disorders. They treated 309 children (231 with hamamelis and 78 with dexpanthenol) and found statistically significant improvements in both groups, with both products equally effective and hamamelis slightly better tolerated.⁷

In 2011,Camargo et al. studied the skin moisturizing capacity of formulations including 0.5, 1.0, or 5.0 percent panthenol topically applied daily to the forearms of healthy subjects over 15- and 30-day periods. They found that use of the 1.0 and 5.0 percent formulations yielded significant reductions in TEWL after application for 30 days. To assess the immediate effects on TEWL and skin moisture, the research protocol also included using the formulations after skin washing with sodium laureth sulfate (SLES). The investigators noted significant TEWL decreases two hours after using the panthenol formulations in comparison to control and vehicle.⁸

Early in 2012, a small comparative pilot study, completed by 26 children (mean age 7.19 years), revealed that 5 percent dexpanthenol was equally effective as 1 percent hydrocortisone ointment in treating mild-to-moderate childhood atopic dermatitis.⁹

Also in 2012, Heise et al. set out to correlate in vitro findings of a stimulatory effect rendered by pantothenate on migration, proliferation, and gene regulation in cultured human dermal fibroblasts to the in vivo wound healing environment. Their clinical data resulting from comparisons of dexpanthenol-treated skin and placebo-treated skin were analyzed at the molecular level and revealed that the provitamin B modulated gene expression in cutaneous wound healing (upregulating IL-6, IL-1β, CYP1B1, CXCL1, CCL18 and KAP 4-2 gene expression and downregulating psorasin mRNA and protein expression).¹⁰

Combination Therapy

Over a decade ago, dexpanthenol was used effectively in combination therapy with xylometazoline for the treatment of rhinitis.¹¹ Dexpanthenol has also been shown, in combination with zinc oxide, to be effective in treating irritant diaper dermatitis, as demonstrated in a prospective, block randomized, investigator-blinded study of 46 children.¹²

In 2008, Abdelatif et al. assessed the safety and effectiveness of an ointment combining royal jelly and panthenol (PEDYPHAR) in 60 patients with limb-threatening diabetic foot infections. Patients were divided into three treatment groups, based on lesion severity. After irrigation and cleansing with normal saline, as well as surgical debridement if necessary, patients were treated with the combination ointment. Lesions were then covered with dressings and the patients were followed up for six months or until full healing, with clinical response checked at weeks 3, 9, and 24. By week 9 and through the follow-up period, 96 percent of the patients in groups 1 and 2 [Group 1: full-thickness skin ulcer (Wagner grades 1 and 2); group 2: deep tissue infection and suspected osteomyelitis (Wagner grade 3)] were deemed to have been cured, with all ulcers in group 1 patients healed and 92 percent among group 2 patients. Group 3 patients [gangrenous lesions (Wagner grades 4 and 5)] healed after surgery, debridement, and conservative therapy with PEDYPHAR. The investigators concluded that the ointment appears promising but additional randomized, double-blind trials are necessary to bear this out.¹³

In a 2011 prospective open pilot trial conducted over 28 days, Castello and Milani evaluated the effect of topically applied 10 percent Urea ISDIN® plus dexpanthenol lotion in the treatment of xerosis and pruritus in 15 hemodialyzed patients (12 women, 3 men, mean age 66 years). The results of twice-daily application on arms and legs were assessed after two and four weeks of treatment and measured against baseline scores for itch, scaling, roughness, redness, and cracks. Significant reductions were observed in all parameters through two, and especially four, weeks of treatment. One patient experienced a mild burning sensation, but the urea and dexpanthenol lotion was found to be effective in ameliorating xerosis and pruritus in hemodialysis patients.¹⁴

Further, in a 2011 study using Wistar rats, Guimarães et al. found that the combination of ultrasound and dexpanthenol (10 percent) sped up the production and organization of collagen in the early phases of wound healing.¹⁵

SAFETY ISSUES

Dexpanthenol confers soothing effects to formulations for the treatment of sunburn and other types of burn. Topically applied dexpanthenol is considered safe, with minimal association with local skin reactions or sensitization. Products with a high concentration of dexpanthenol may be contraindicated in people with hemophilia. Despite its frequency of use in topical products, contact allergy is rare. There is one report of one case of allergic contact dermatitis to panthenol combined with cocamidopropyl PG dimonium chloride phosphate in a facial hydrating lotion.¹⁶

ENVIRONMENTAL IMPACT

There is no measureable environmental impact from the vitamin pantothenic acid, found throughout nature, or the synthetic dexpanthenol.

FORMULATION CONSIDERATIONS

Water-in-oil topical emulsions are the best vehicle to deliver sufficient skin penetration and local concentrations of dexpanthenol.¹

USAGE CONSIDERATIONS

Pantothenic acid is not well absorbed into skin but dexpanthenol is well absorbed through the skin and is rapidly converted to pantothenic acid.¹ Absorption into skin was found to be reduced when dexpanthenol is combined with olive oil, highlighting the relevance of the vehicle.¹⁷ One study used a perfused skin udder model and demonstrated that the rate and extent of penetration of dexpanthenol is much lower in an oil/water formulation.¹⁸ Therefore, panthenol-containing formulations should not be layered over oil-containing preparations if maximal dexpanthenol absorption is desired.

SIGNIFICANT BACKGROUND

Dexpanthenol is included as an ingredient in various topical skin formulations and shampoos. In fact, this form of vitamin B₅ has long been considered an effective ingredient in cosmetic products. The foundation for the use of dexpanthenol in shampoo – and anecdotal reports that its use restores color to gray hair – stems from a study several years ago evaluating dexpanthenol deficiency in rats. Deficiency was correlated with hair turning gray or falling out. Pantothenic acid deficiency in humans is exceedingly rare, though, and is not likely associated with hair changes in humans.¹⁹ Clearly, the etiologies of graying of hair and baldness are not related to this vitamin. Also, no oral or topical formulations containing pantothenic acid or dexpanthenol as the main active ingredient have been shown to prevent gray hair or balding in humans. Nevertheless, it is believed that dexpanthenol penetrates well into the hair shaft, promotes luster and elasticity, and leaves the hair easier to comb.

CONCLUSION

Dexpanthenol is a versatile compound suitable for use in treating various dermatoses and its topical application is employed broadly in clinical practice to enhance wound healing. Few controlled clinical trials have been conducted evaluating the efficacy of topical formulations containing dexpanthenol intended for skin care. Despite a dearth of randomized, double-blind, case-controlled studies establishing the efficacy of such products, current data warrant further study and support the conclusion that dexpanthenol as an active ingredient at least confers some benefit as a humectant.

There is an increasing body of anecdotal, empirical evidence also suggesting significant potential contributions by provitamin B₅ as an ingredient in topical formulations.

REFERENCES

INTRODUCTION

SOURCE

Urea, the primary nitrogen-containing substance found in mammalian urine, is among the most commonly used humectant ingredients, along with glycerin, sorbitol, sodium hyaluronate, propylene glycol, α-hydroxy acids, and sugars (Table 28-1). It is a component of the natural moisturizing factor (NMF) and also exhibits a mild antipruritic effect.¹ Urea in high concentrations (e.g., 40 percent) is effective as a hydrating and keratolytic agent in various cutaneous conditions including xerosis, psoriasis, onychomycosis and other nail disorders, ichthyosis, eczema, calluses, and corns.

HISTORY

Urea was first discovered in the 1700s in Europe. Its discovery is most often attributed to French chemist Hilaire Rouelle’s work in 1773, though Dutch scientist Herman Boerhaave is said to have first identified the compound as a major constituent of mammalian urine in 1727.^2,3 German physician and chemist Friedrich Wöhler discovered, in 1828, that urea could be synthesized in vitro by combining the inorganic materials cyanic acid and ammonium without using any organic substances.² This was the first credited laboratory synthesis of a naturally occurring organic compound.² Interestingly, the chemical synthesis reported by Wöhler is not the chain of events that occurs in the mammalian liver to produce urea; the “urea cycle” was identified by German physician Hans A. Krebs and his medical student Kurt Henseleit in 1932.² Urea has been used in hand creams since the 1940s.⁴ Thirty years later, urea in 20 percent concentrations was found to be effective in treating pruritus.⁵

CHEMISTRY

The urea molecule has two NH₂ groups united with a carbonyl functional group. It acts as a physiological NMF.⁶

ORAL USES

Oral urea is used for some medical indications and has a long history of traditional folk medicine use, but this mode of administration is not thought to have appreciable cutaneous benefits.

TOPICAL USES

Twenty years ago, a double-blind, randomized comparison of two urea-containing creams revealed that 3 and 10 percent urea cream were equally effective in treating aspects of dry skin, particularly increasing hydration and decreasing scaling, and more effective than the vehicle control. Transepidermal water loss (TEWL) was unchanged after treatment with the 3 percent urea cream, but the 10 percent urea cream reduced TEWL. Use of the 3 percent cream resulted in a slightly golden tint to the skin color.⁷

In 1998, Jennings et al. conducted a double-blind, randomized paired comparison of the keratolytic effects of 5 percent salicylic acid and 10 percent urea ointment (Kerasal) on one foot and 12 percent ammonium lactate lotion (Lac-Hydrin) on the other foot in 70 patients with mild-to-moderate xerosis. After two weeks of treatment, 54 patients were evaluated, 39 of whom were evaluated after four weeks of treatment. There were no significant differences in the treatment arms, but after four weeks of treatment, both yielded significant reductions in xerosis severity.⁸

Urea in high concentrations is also a useful adjuvant, as is salicylic acid, in the keratolytic phase of psoriasis treatment.⁹ In 10 percent creams, urea is used to treat ichthyosis and hyperkeratotic skin conditions;¹⁰ in lower concentrations, it is employed to treat mild xerosis.⁶

In 2002, Ademola et al. conducted a randomized, double-blind, bilateral study with 25 women and men to compare the clinical effectiveness and tolerability of a 40 percent urea topical cream (Carmol 40) and 12 percent ammonium lactate topical lotion (Lac-Hydrin 12 percent) used to treat moderate-to-severe xerosis. Patients were assessed at baseline as well as two and four weeks after the initiation of therapy. Ratings by the 18 patients who completed the study as well as investigators revealed that xerosis symptoms were reduced in less time in the urea group. Further, instrumental and clinical measurements showed that by day 14, 40 percent urea cream was associated with less skin roughness, fissures, thickness, and dryness than 12 percent ammonium lactate lotion.¹¹

In 2008, Stebbins et al. performed a single-blind pilot study with 12 trained female panelists to evaluate the acceptability (spreadability, odor, and postapplication residue) of six 40 to 50 percent urea preparations. They compared the composite scores of the five older formulations with that of new emollient product, and found that the novel preparation (U-Kera E) was the most cosmetically acceptable (most spreadable and left less residue). Odor scores were low overall and comparable among the six preparations. The investigators noted that while all urea-based creams exhibited a detectable odor, it was usually fleeting and often such products are used in areas furthest from the nose (i.e., legs and feet).¹²

In 2010, Lodén et al. randomized 53 patients with successfully treated hand eczema to compare the time to relapse when using a barrier-strengthening urea (5 percent) preparation or receiving no treatment. They observed that the median time to relapse was two days in the no treatment group and 20 days in the urea-containing moisturizer group. At the time of relapse, there were no noted variations in severity. The investigators concluded that the barrier-strengthening moisturizer containing urea successfully acted to extend the remission of controlled hand eczema.¹³

SAFETY ISSUES

Cases of contact allergy from urea-containing products are rare. In one instance, at least, the reaction may have been due to ingredients other than urea.¹⁴ However, urea is considered generally safe and nontoxic. Urea is contraindicated in neonates due to the risk of systemic absorption.¹⁵

ENVIRONMENTAL IMPACT

Used widely in fertilizer as well as cosmetic products, urea is produced in copious amounts on an industrial scale from liquid ammonia and liquid carbon dioxide, with global production in 2012 estimated at 184 million tons.¹⁶ The inclusion of urea in topical products, thus, certainly exerts some kind of impact on the environment, primarily in fuel transportation costs.

FORMULATION CONSIDERATIONS

There are many different types of formulations using urea as discussed in this chapter that are beyond the scope of this book. (Please see the NWU Institutional Repository website, http://dspace.nwu.ac.za/handle/10394/251, for more information on urea product formulations.)

USAGE CONSIDERATIONS

The combination of urea with hydrocortisone, retinoic acid, and other agents has been shown to enhance the penetration of these agents.^17,18 Urea has also been combined with several other compounds in recent years to achieve significant clinical effects.

SIGNIFICANT BACKGROUND

In 2009, Amichai and Gunwald conducted a three-week randomized, double-blind study with 36 males and females with mild-to-moderate atopic dermatitis, ranging in age from 3 to 40 years, to ascertain the efficacy of liquid soap containing 12 percent ammonium lactate and 20 percent urea. Investigator and patient assessment indicated that significant improvements were observed, specifically reductions in erythema, scaling, and xerosis, in the active soap group (24 patients) compared to results in the 12-person control group.¹⁹

In 2010, Pardo Masferrer led a prospective observational study in 98 breast cancer patients to investigate the intensive use of a hydrating lotion containing 3 percent urea, polidocanol and hyaluronic acid to treat and mitigate the severity of radiodermatitis. The investigators monitored patients weekly for skin toxicity and compared the incidence and grade of toxicity with a control sample from 174 breast cancer patients whose skin was treated at radiotherapy initiation in 2006 or upon the emergence of radiodermatitis. They found the incidence of radiodermatitis lower in the intensive use group. In addition, intensive use of the combination lotion was also associated with a lower grade of toxicity.²⁰

The use of urea in combination with other ingredients in the Ureadin line of products has garnered recent attention. In 2011, Castello and Milani found, in a prospective open pilot trial with 15 patients, that the twice-daily application of topical 10 percent urea plus dexpanthenol (Ureadin Rx 10) significantly reduced itching scores as well as scores along the SRRC (scaling, roughness, redness, and cracks) Index, indicating that the formulation was effective in treating xerosis and pruritus in such patients.²¹

Later that year, Tadini et al. studied the keratolytic and moisturizing activity of Ureadin Rx10 in a four-week randomized, controlled, single-blind, two-center, intra-patient trial with 30 ichthyosis vulgaris patients ranging in age between 8 and 56 years. In a right-vs.-left study design, patients were treated with 10 percent urea-based Ureadin Rx 10 lotion on one side and the glycerol-based emollient cream Dexeryl on the other side. Patients were assessed at baseline as well as two and four weeks after treatment initiation. Twenty-seven participants completed the study, in which the researchers found that both topical formulations were clinically effective after four weeks and well tolerated. SRRC scores significantly fell from 9.5 to 3.3 (65 percent decline) in association with Ureadin use and from 9.5 to 5.7 (40 percent decline) in Dexeryl-treated areas. Mean global efficacy scores were significantly higher in areas treated with Ureadin (8.9) as compared to Dexeryl (7.3). The investigators concluded that Ureadin is a more effective option than Dexeryl for reducing the hyperkeratosis and xerosis associated with ichthyosis, but larger sample sizes would be necessary in future research to establish safety and tolerability for the urea-based lotion in these patients.²²

In 2012, Federici et al. conducted a 28-day, randomized, evaluator-blinded, comparative study in 40 type 2 diabetics, aged 40 to 75 years, to assess the efficacy of a topical preparation containing urea 5 percent, arginine, and carnosine (Ureadin) as compared to the glycerol-based emollient Dexeryl. Twice-daily application of the urea-based product resulted in significantly more hydration as compared to the glycerol-based emollient. Visual Analogue Scores increased in both groups, but were significantly higher in the Ureadin group. The investigators concluded that the urea/arginine/carnosine cream enhanced skin hydration while lessening xerosis in type 2 diabetic patients more efficiently than the glycerol-based control formulation.²³

Also in 2012, Emtestam et al. led a 24-week, randomized, double-blind, placebo-controlled multicenter study of 493 patients (n = 346; placebo = 147) to test the efficacy, safety, and tolerability of a novel topical preparation (K101) combining urea, propylene glycol, and lactic acid for treating distal subungual onychomycosis. Patients were stratified based on level of nail involvement, with investigators finding that 27.2 percent of patients with ≤ 50 percent nail involvement treated with K101 achieved the primary endpoint and 10.4 percent in the placebo group. Among the patients with 51 to 75 percent nail involvement, 19.1 percent achieved the primary endpoint as compared to 7.0 percent in the placebo group (not a significant difference). The investigators noted that the antifungal topical preparation was safe and imparted clinically significant as well as visible nail improvements. Further, they noted that when applied in concentrations greater than 20 percent, urea displays nail-dissolving properties.^24–26 Others have noted a concentration-dependent keratolytic effect of softening and macerating the nail with 30 to 50 percent concentrations of urea.¹⁴

Ciammaichella et al. reported at the end of that year on a pilot registry study of the effects of twice-daily application of Ureadin for four weeks in diabetic patients with microangiopathy and mild-to-moderate xerosis of the foot. A reduction in skin breaks was associated with Ureadin treatment as compared to controls. Improvements were also linked to Ureadin use along various other parameters in comparison to controls (i.e., greater skin thickness, favorable investigator global assessments and subject assessments, no new skin lesions vs. four lesions in the control group). The researchers concluded that given the good level of diabetes control prior to inclusion in the study and no such changes after four weeks, the improvements seen in the study could be attributed to the topical application of Ureadin and not to systemic diabetes management.²⁷

CONCLUSION

Urea has long been used in dermatology for its well-established activity as a hydrating and moisturizing agent. It remains as one of the most effective humectant agents.

REFERENCES

INTRODUCTION

Aquaporins (AQPs) are integral membrane proteins that form a water channel and facilitate water transport in various organs such as the skin, renal tubules, eyes, the digestive tract, and the brain. In 2003, Peter Agre received the Nobel Prize in chemistry for discovering AQPs. There are 13 isoforms of AQPs found in mammals classified as types 1 to 13. Functionally, they can be classified into two subtypes: AQPs 1, 2, 4, 5, and 8, which only transport water, and AQPs 3, 7, 9, and 10, which can conduct other substances such as glycerol (also known as glycerin) or urea in addition to water.¹ AQP-3 is the predominant water channel found in human epidermis, and is permeable to both water and glycerin. For years scientists have known that glycerin plays a superior role in hydrating skin,² but the reasons for this became more clear when AQP-3 was discovered. Studies have shown that defects in AQP-3 in mice models result in epidermal dryness as well as decreased stratum corneum hydration and glycerin content of the epidermis, followed by reduced elasticity and impaired skin barrier recovery.^3,4 Aquaporin facilitates the transport of water, glycerin, and solutes between keratinocytes.

AQPs are transmembrane structures arranged as homotetramers in the cell membrane. Each subunit of the tetramer consists of six α-helical domains and contains a distinct aqueous pore. The intricate shape and the transmembrane position make it impossible to exogenously add aquaporin to skin as the marketing claims of some cosmetic products imply. Ultraviolet light has been shown to diminish the expression of AQP-3, likely through MAP kinase pathways.⁵ Free radicals have also been demonstrated to decrease AQP-3 expression.⁵ This is one of the reasons that sun-exposed skin becomes dehydrated.⁵ In addition, aquaporin performance can be affected by topical ingredients that can increase the opening and closing of the aquaporin pore or upregulate AQP-3 expression. Ajuga turkestanica has been shown to increase AQP-3 function.⁶ Retinol and the antioxidant n-acetyl cysteine both have been shown to inhibit the downregulation of AQP-3 upon UV exposure.⁵ AQPs are intriguing water channels that allow water and glycerin to flow from cell to cell. In the next few years, many more facets of their dynamic mechanisms of action will be elucidated.

REFERENCES

INTRODUCTION

SOURCE

Ajuga turkestanica is a perennial herb and member of the mint family Lamiaceae. There are over 300 species of the genus Ajuga found throughout Europe, Asia, Africa, Australia, and North America. A. turkestanica is indigenous to Uzbekistan. It derives its name from the fact that it contains a powerful ecdysteroid called turkesterone. Ecdysterones are a group of plant sterols that have a steroid-like effect on the human body.

HISTORY

This plant has been used in traditional medicine to treat fevers, toothaches, dysentery, malaria, high blood pressure, diabetes, and gastrointestinal disorders, as well as antifungal, antihelminthic, anti-inflammatory, antimycobacterial, and diuretic agents.¹ Athletes and body builders have used A. turkestanica as an anabolic steroid because it contains phytoecdysteroids.² Many bodybuilding supplements containing turkesterone can be found on the Internet. A. turkestanica is known to contain several bioactive compounds and has been used in traditional medical approaches to heart disease as well as stomach and muscle aches.^3,4 In addition, it is one of the many species of Ajuga gaining attention for demonstrating medicinal properties with the potential for commercial applications.⁵ In November 2000, US Patent Number 7060693 was filed for the cosmetic use of A. turkestanica (awarded in 2006) by scientists at Dior Research.

CHEMISTRY

There are three classes of potentially bioactive compounds found in the Ajuga genus⁵:

1. Clerodane diterpenes are recognized sources of antimicrobial, antiviral, antitumor, antibiotic, and amoebicidal activities.⁶

2. Phytoecdysteroids have anabolic steroid activity. A. turkestanica reportedly contains several phytoecdysteroids (turkesterone, 20-hydroxyecdysone, cyasterone, cyasterone 22-acetate, ajugalactone, ajugasterone B, α-ecdysone and ecdysone 2, 3-monoacetonide) as well as the iridoids harpagide and harpagide 8-acetate.^5,7–12

3. Iridoid glycosides, especially abundant in Ajuga decumbens, have exhibited anticancer activity.¹³

ORAL USES

A. turkestanica extract is included in some anabolic muscle growth supplements.

TOPICAL USES

Patented extracts of A. turkestanica were shown in 2006 to have sufficient ecdysteroids and other active ingredients to improve the differentiation of keratinocytes, thus facilitating skin hydration and yielding antiaging effects (Table 30-1).^14,15 The patent inventors observed that the extracts are especially effective in regulating epidermal water and glycerol transport, achieving improved hydration of the basal layer by working in concert with or enhancing aquaporin-3 (AQP-3).^14,16 AQP-3 is a water transport channel known to transport water and glycerol between cells.¹⁷ The function of AQP-3 was shown to be increased by A. turkestanica extract.¹⁸

SAFETY ISSUES

A. turkestanica has not been reviewed for safety by the Cosmetic Ingredient Review (CIR) panel.

ENVIRONMENTAL IMPACT

A. turkestanica hairy root cultures have been developed as a sustainable alternative to wild-harvesting.¹⁹

FORMULATION CONSIDERATIONS

The only considerations that have been made public are disclosed in US Patent Number 7060693.

USAGE CONSIDERATIONS

There are no known restrictions. A. turkestanica can be used in the morning or at night and can be used in combination with other skin care products.

SIGNIFICANT BACKGROUND

Aquaporins

Aquaporins (AQPs) are integral membrane proteins that facilitate water transport in several organs, including the skin, brain, renal tubules, eyes, and digestive tract. Thirteen isoforms of aquaporins (AQP 0–12) are found in mammals. Of these, there are two functional subtype classifications: AQPs 1, 2, 4, 5, and 8 conduct only water, and AQPs 3, 7, 9, and 10 transport water and other substances including glycerol and urea.²⁰ AQP-3, permeable to water and glycerol, is the main water channel in human epidermis. Glycerol acts as an endogenous humectant thereby facilitating hydration of the stratum corneum (SC).²¹ Defects in AQP-3 in mice models have been demonstrated to lead to epidermal xerosis, reduced SC hydration and epidermal glycerol content, followed by diminished elasticity and impaired skin barrier recovery.^22,23 Such findings underscore the important role of glycerol in cutaneous hydration. AQP-3 contributes to the transport of water, glycerol and solutes between keratinocytes.

Dumas et al. note that the role of AQPs in hydrating the living layers of the epidermis where keratinocyte differentiation occurs and in barrier development and recovery suggests that they are significant protein targets for improving the quality and resistance of the skin surface as well as ameliorating aging- and UV-induced xerosis.¹⁸

Ajuga Turkestanica and Aquaporins

In 2007, Dumas et al. conducted in vitro and in vivo studies of active ingredients capable of raising AQP-3 levels to enhance hydration in human skin keratinocytes, with the understanding that improving hydration in human keratinocytes would ultimately improve epidermal hydration.²⁴ They used an ethanolic/water (70/30 v/v) extract of A. turkestanica as the hydrating agent (2.5 μg/mL) and found that after 17 days of in vitro treatment every 2 days in human reconstructed epidermis, AQP-3 expression measured at the protein level was significantly elevated. Increased epidermal proliferation and differentiation was also noted. Electron microscopy showed a significantly thicker, compact SC and more clearly differentiated desmosomes.¹⁸

The investigators prepared an oil-water emulsion infused with A. turkestanica extract (0.3 percent w/w) for an in vivo study in which 15 healthy female volunteers (between 22 and 56 years old) applied the formulation twice daily to their forearms for 21 days. Significant reductions in TEWL were seen in the treated area compared to the control at days 7 and 21. The researchers concluded that the tested A. turkestanica extract formulation enhanced AQP-3 expression and human epidermal differentiation in vitro and ameliorated epidermal barrier structure and human skin recovery in vivo.^15,18

CONCLUSION

The Ajuga turkestanica story is a very interesting one because the cosmetic scientists Dumas, Bonté, and Gondran were the first to identify AQP-3 in skin cells. They published this information, sharing it with other scientists rather than keeping the information hidden and proprietary. Several compounds were tested with A. turkestanica found to affect water flux through the AQP channels. They filed patents to protect this technology. This is a great example of scientific discovery leading to new technologies. Unfortunately, many companies have started to claim that their products “contain aquaporin,” which is misleading. If AQP was included in a formulation, it would not be able to penetrate into the skin and span the lipid bilayer in the manner that is necessary to affect water flux (Figure 30-1). The only way that is proven to affect water flux between cells is for an ingredient to act upon AQP in the manner that Ajuga turkestanica does. The discovery of AQP-3 as well as the understanding of its significance and how to harness its power to improve skin appearance occurred over a span of two decades. The scientists should be congratulated for their perseverance.