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Hexylresorcinol: Providing Skin Benefits by Modulating Multiple Molecular Targets

Hexylresorcinol: Providing Skin Benets by
Modulating Multiple Molecular Targets
Ratan K. Chaudhuri
Hexylresorcinol (HR) is an alkylresorcinol (AR), a type of phenolic lipid, having an n-hexyl chain
attached to the 4 position of the 1,3-dihydroxybenzene ring (F igu r e 7.1). It can be synthesized by react-
ing resorcinol with hexanoyl chloride in the presence of Lewis acid catalyst. The resultant intermediate,
hexanoylresorcinol, is then reduced to hexylresorcinol. Subsequent purication of HR is attained by, e.g.,
crystallization using suitable solvent(s).
Hexylresorcinol is an ingredient that has attained GRAS (Generally Recognized as Safe) status and is
effective as an anti-browning agent for prevention of melanosis in shrimp.1 HR has also been shown to
be a very effective inhibitor of surface browning on many fresh-cut fruits, such as apples, pears, man-
goes, etc.2,3 When combined with ascorbic acid, it has a synergistic effect in the prevention of browning.
Here, ascorbic acid reduces quinones generated by polyphenoloxidase while HR specically interacts
with polyphenol oxidase, and renders it incapable of catalyzing the enzymatic reaction.4,5 A post-cutting
dip of HR, ascorbic acid, and calcium lactate was found to extend the shelf-life of pear slices from 15 to
30 days.6 Similarly, Red Delicious apple slices treated with an anti-browning dip (HR, isoascorbic acid,
N-acetyl cysteine, and calcium propionate) and held at 5°C maintained visual quality for ve weeks.7
Although sultes are more commonly used in controlling browning of foods, HR has several advantages
over sultes, including its specic mode of inhibitory action, effectiveness at low concentrations, inabil-
ity to bleach preformed pigments, and chemical stability.
From a human physiological and biological perspective, HR is perhaps one of, if not the most studied
and well-known AR. It is reported to have anesthetic, antiseptic, and anthelminitic properties8 and can be
used topically, e.g., on small skin infections or as an ingredient in a consumable carrier, e.g., throat loz-
enges.9 As a throat lozenge it manifests both antiseptic and local anesthetic effects: its antiseptic action
killing the bacteria that may be associated with a sore throat, while its anesthetic action helps relieve the
pain associated therewith. Here, the action of sucking the lozenge allows the active ingredient to work in
the area of the discomfort, and also helps to coat, lubricate, and soothe the irritated throat tissue.
ARs are also found in nature; alkyl chains C17:0–C25:0 attached to 5 position are abundant in whole-
grain wheat and rye.10 ARs are reported to have antitumor, antibacterial, antifungal, and antiparasitic
activities. These effects of ARs were attributed to membrane-modulating effects due to interactions of
their alkyl tails with phospholipids and/or proteins and to antioxidant effects of the phenolic hydrogen.11
HR has a long history of human use. The earliest citation to its use in humans appears to be that of
Dr. Veader Leonard and his associates at the Johns Hopkins School of Hygiene and Public Health in
Baltimore, Maryland.* As reported, Dr. Leonard and his associates were looking for a “perfect” anti-
septic that was deadly to germs but harmless to man. In the course of their efforts, they found that HR
possessed over 50 times the germ-killing power of pure carbolic acid.
* Cited by Time Magazine, Medicine: Hexylresorcinol, Monday Februar y 23, 1925; obtained f rom
72 Cosmeceuticals and Active Cosmetics
Despite its long history of use as an antiseptic, anesthetic, and anthelminitic, it is only recently that its
use and benets as an active ingredient for skin care applications has been realized. In 2007, Sytheon
Ltd. introduced HR as a cosmetic/skin care additive for use as a skin protectant having skin lighten-
ing and even-toning properties under the trade named Synovea® HR.12 Its utility in this application is
believed, in part, to arise from the ability to attain highly puried HR (>99%, typically ranging from
99.5% to 99.9%) which is essentially free of resorcinol, a known skin irritant, and hexanoylresorcinol,
the intermediate.12
Focusing on the human physiological and biological properties of HR, attention will now be given to
its antimicrobial, antitumor, anti-aging, and skin lightening/even-toning properties in particular.
On July 22, 1991, the U.S. Food and Drug Administration (FDA) published the Tentative Final
Monograph for antimicrobial drug products—rst aid antiseptics13—in which HR was identied as a
category I antimicrobial ingredient. HR has been used extensively in antibacterial mouthwash and, as
noted above, throat lozenges. Antibacterial mouthwashes are effective due to the presence of HR, which
inhibits the growth and proliferation of microorganisms found in the mouth: thus preventing caries and
ameliorating infectious conditions. As a side benet, mouthwashes having HR also prevent or reduce
halitosis, which, in many instances, is generated or arises from various microorganisms in the mouth.
As noted above, lozenges such as Strepsils® Extra Blackcurrant Lozenges containing HR possess both
antiseptic properties to ght bacterial throat infections and anesthetic properties to soothe and numb
throat pain.
The emergence of pathogenic bacterial strains resistant to currently available antimicrobial agents
continues to be a universal problem of ever increasing importance.14 Extensive efforts have been under-
taken to validate new target enzymes for antimicrobials; however, these have met with little success,15
with the majority of successful drugs inhibiting just a handful of cellular processes. To date, one of the
most successfully exploited drug targets has been the DNA topoisomerase (topo) class of enzymes. For
example, recent studies by Taylor etal. have shown that HR has good DNA topoisomerase inhibitory
activity (IC50 30 µM), which compares well with m-Amsacrine (30 µM) and Purpurin (40 µM).14
Chaudhuri has also demonstrated excellent antimicrobial activities of HR in an, as yet, unpublished
work. Specically, Chaudhuri conducted an evaluation to assess the minimum inhibitory concentra-
tion values (MIC in µg/mL) of HR against various organisms relevant to personal care applications in
accordance with U.S. Pharmacopeia’s Compendia Products procedure for Category 2 (USP 26-87 pp.
2022–2026). The results are given in Tabl e 7.1.
In addition, Chaudhuri conducted a challenge test using 0.5% (w/w) level of HR in a lotion. The
results showed a 3-log reduction of fungi, but not the desired reduction of bacteria. Nevertheless, it
appears that HR can act as a synergist in combination with existing commercial antimicrobial ingredients.
HR has also found potential utility as an antitumor agent, particularly in relation to differentiation ther-
apy. The signaling pathways related to cell differentiation and senescence fail to function properly in
malignant tumor cells. As a result, tumor cells exhibit uncontrolled and invasive growth. Differentiation
therapy is currently being considered as one of the key emerging techniques for the treatment of can-
cer.16 Kim etal. have recently shown that HR induces the differentiation of SCC-9 via the modulation
of the E2F-mediated signaling pathway and suppressed the growth of SCC-9 cells in a dose-dependent
FIGURE 7.1 Structure of hexylresorcinol (HR).
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manner.17 In this study, the authors also found that HR increased the expression of the epithelial cell dif-
ferentiation markers involucrin and keratin 10.
Following on the foregoing, Kim and Choi have further demonstrated that HR dose-dependently
induced SCC-9 cell apoptosis as determined by caspase-3 activity, annexin V expression, as well as
by scanning and transmission electron microscopy.18 HR was shown to inhibit intracellular calcium
oscillation in both SCC-9 cells and normal human dermal broblasts. HR-induced apoptosis was partly
reversed by calcium channel blockers. Additionally, HR reduced the tumor mass formed by SSC-9 cell
implantation in BALB/cAnNCrj-nu/nu mice and mass size reduction was also partly reversed by the con-
comitant application of calcium channel blockers. The results of this study suggest that HR is providing
strong antitumor effects by inhibiting calcium channel oscillation and inducing apoptosis.18
Finally, the resistance to chemotherapy is very important in the prognosis of tumors.
Transglutaminase-2 (TG-2) mediated chemotherapy resistance has been widely reported. TG-2 is
overexpressed in many cancers such as breast cancer,19 malignant melanoma,20 and glio-blastoma21
and facilitates tumor spread and metastasis. Recently, Kim etal. reported on their nding of an inhibi-
tory effect on TG-2 enzyme activity by HR.17 Additionally, when the authors compared the perfor-
mance of a mixture containing 5 µg/mL HR and 5 µg/mL cisplatin (a common chemotherapy drug)
to 10 µg/mL of cisplatin alone on tumor cells, they found the application of the mixture resulted in
signicantly lower tumor cell viability than the cisplatin alone (p < 0.05). Accordingly, it is believed
that HR has a synergistic effect on KB cell death when employed in combination with cisplatin. The
authors also examined the effects of the combination of cisplatin and HR on oral mucosal melanoma
(OMM) using cultured primary OMM cells in a tumor xenograft model. According to their ndings,
the combination resulted in fewer metastases and longer survival than cisplatin-only treatment in the
OMM xenograft model.22 Based on these ndings, it is now believed that HR could be a viable candi-
date as a chemotherapeutic agent in the treatment of cisplatin resistant tumors.
Recent ndings also demonstrate the inuence or impact of HR on a variety of processes involved in skin
aging and damage, including oxidation, inammation, and glycation.
Although HR would not typically be expected to be an effective conventional antioxidant (radical and/or
non-radical quencher) based on its structure23; it may still be considered an antioxidant in a broader
Minimum Inhibitory Concentration Values (MIC
in µg/mL) of HR against Various Organisms
Organisms MIC Value (µg/mL)
E. coli 2.5
S. aureus 4.5
S. epidermidis 4.5
Streptococcus 1.5
Lactobacillus 4.0
P. gingivalis 1.0
P. acne 25 to 50
Aspergillus niger 0.3
Candida albicans 3.5
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74 Cosmeceuticals and Active Cosmetics
sense because its metabolites, such as, 1,2,3-trihydroxy- and/or 1,3,5-trihydroxy-4-hexylresorcinols, can
have potent radical and non-radical quenching activity.11 Antioxidant activity may also be attributed to
HR, though indirectly, as a result of its ability to stimulate the cell protectant glutathione and various
antioxidant defense enzymes such as glutathione peroxidase and glutathione reductase.24 For example,
Yen etal. have demonstrated that HR has a protective effect against oxidative DNA damage in human
lymphocytes induced by hydrogen peroxide.24
Inammation is a complex physiological process and the role of transcription factor NF-kappaB in the
inammatory response has been well documented.25,2 6 NF-kappaB is activated by numerous stimuli and
once fully activated participates in the regulation of various target genes in different cells to exert its
biological functions. NF-kappaB has often been referred to as a central mediator of the immune response
since a large variety of bacteria and viruses can lead to the activation of NF-kappaB, which in turn con-
trols the expression of many inammatory cytokines, chemokines, immune receptors, and cell surface
adhesion molecules.
Recently, Kim etal. reported that HR inhibited NF-kappaB phosphorylation.27 Independently, Johnson
& Johnson scientists found that HR has a stimulating effect on collagen and elastin synthesis through
NF-kappaB inhibition.28 Based on these ndings, and the fact that inammation has a signicant effect
on skin aging, it is anticipated that HR would provide an anti-inammatory as well as anti-aging effect
on skin when applied topically.
Glycation is the term used for a class of non-enzymatic reactions that occurs between sugars, such as
glucose or ribose, and proteins and lipids, including, e.g., the reaction between the nucleophilic amino
group of proteins and a reducing sugar wherein the sugar becomes bonded to the protein. Glycation is
the rst of a series of reactions by which advanced glycation end-products (AGEs) are formed: the sub-
sequent reactions include Schiff base reactions, Amadori reactions, and Maillard reactions. Though not
all glycation reactions lead to AGEs, physiological and biological conditions will shift the reactions to
favor or disfavor their formation.
AGEs are heterogeneous compounds which adversely affect nearly every type of cell and molecule
in the body and lead to human pathological conditions. AGEs are thought to be one of the key factors
in skin aging. For example, studies of collagen glycation using skin equivalents29 found a number of
changes including modied broblast shape and distribution, enhanced extracellular matrix molecules
and the dermal-epidermal junction zone, and increased collagenase activity. Additionally, AGE treat-
ment has been found to reduce various biomarkers of skin aging, including those noted above, as well
as increased NF-κB activation and cytokine expression.30 Exposure of AGEs to ultraviolet (UV) light
generates reactive oxygen species (ROS) in extracellular matrix.31
Turning to HR, it has been reported to have an inhibitory effect on the formation of Maillard reac-
tion products (in vitro using glucose and cysteine).32 Given the importance of Maillard reactions in the
formation of AGEs, it is expected that HR, when applied topically, would provide skin benets due to its
anti-glycation/anti-AGE property.
Melanin and Melanogenesis
Skin color is one of the most important physical traits of humans because it affects so many aspects of
our health and social well-being. Skin color is also one of the best examples of evolution by natural selec-
tion acting on the human body. Anthropologic studies have provided us with two important facts: the
earliest Homo sapiens had dark skin, rich in protective melanin, and small groups of “modern” humans
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dispersed out of the African tropics into less intensely sunny parts of Africa, Eurasia, and the Northern
Hemisphere. Over time, these latter groups of humans underwent genetic changes leading to the loss
of melanin pigmentation.33 Skin color is one of the most conspicuous ways in which humans vary and
has been widely used to dene human races. Unfortunately, skin color is also a determinant of human
interaction and destinies.
Human skin coloration is both adaptive and labile as evidenced by the multitude of different skin
colors of humans around the world. Though humans are often characterized as having black, white,
red, or yellow skin, no one is actually any of these colors, as these are commonly used terminologies
that do not reect biological reality.34 Skin coloration in humans arises from a complex series of cellular
processes involving the synthesis and transfer of a pigment, melanin, which, besides being responsible
for skin color and tone, is the key physiological defense against sun-induced damage, such as sunburn,
photo-aging, and photo-carcinogenesis. The melanin itself is formed by a sequence of reactions in which
tyrosine is oxidized to dopa and then dopa is subsequently oxidized to melanin. These reactions are cata-
lyzed by the enzyme tyrosinase. The formation of melanin, also known as melanogenesis, is carried out
in melanosomes, organelles of that population of cells known as the melanocytes, which are located in
the lower part of the epidermis. The melanosomes containing the melanin are subsequently transferred
from the melanocytes to the neighboring cells, the keratinocytes, which then transport and distribute the
melanin to the upper layers of the skin.35
Skin coloration is primarily regulated by the amount and type of melanin synthesized by the melano-
cytes36,37; however, additional and equally contributing factors include (a) the efciency of the transfer of
the melanosomes, hence the melanin, from the melanocytes to the neighboring keratinocytes, a process
that occurs with the help of E-cadherin, an adhesion protein, and (b) the subsequent distribution and
degradation of the transferred melanosomes by the recipient keratinocytes.38 Although a number of fac-
tors can inuence the formation of melanin, exposure of the skin to UV light markedly inuences and
increases the amount and rate of melanin production, most often producing a further darkening of the
skin or a “tan.” Sun exposure as well as hormones and other environmental and physiological factors
can also lead to more localized skin pigmentation, e.g., age spots, melasma, and freckles, owing to an
overabundance of melanin production in the aficted areas of the skin.
Skin Lightening/Even-Toning
For a substantial segment of the human population, one’s natural skin color is not satisfactory and efforts
are undertaken to modify it. For example, in North America, Europe, and Australia, many endeavor to
enhance skin pigmentation through tanning, whether natural (induced melanin production) or articial
(dyes). Others, especially certain Asian cultures, endeavor to prevent skin coloration and/or seek to actu-
ally lighten their natural skin color. Additionally, skin lightening efforts are oftentimes undertaken to
address or eliminate age spots, melasma, and freckles, or to obtain even-toning effect with one’s skin.
HR has been found to have a signicant skin lightening effect due to a strong inhibitory effect on tyros-
inase and peroxidase and a stimulatory effect on glutathione and E-cadherin syntheses. It is believed that
the HR bind to tyrosinase directly and inhibits its enzyme activity.39 Additionally, having observed that
fragments of DNA can stimulate melanin synthesis, it seems that the reduction of DNA damage by HR,
as previously noted, may also be responsible for inhibition of melanin synthesis.
Chen et al. found that HR inhibits both the monophenolase (tyrosine to DOPA) and diphenolase
(DOPA to DOPAchrome) activity of mushroom tyrosinase.39 Its activity or mode of action, as shown
by a kinetic analysis, is as a competitive inhibitor. More recently, Chaudhuri demonstrated that HR is a
far superior tyrosinase inhibitor (using both tyrosine and DOPA substrates) than the three commercially
available skin lighteners: hydroquinone, kojic acid, and licorice extract.12 Concurrently, HR was found to
provide superior inhibition of peroxidase activity of HR as compared to the three skin lighteners tested
as well.12 This study also showed that at 10 µg/mL use level, HR had inhibitory effects on extracellu-
lar and intracellular melanin production by 75% and 36%, respectively, when compared with placebo.
Related studies using B16 melanoma cells showed that their growth rate was not signicantly altered in
the presence of HR during a 72 h incubation period, indicating that the HR-induced regulatory effects on
melanogenesis of melanoma cells occurred without affecting cell proliferation.12
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76 Cosmeceuticals and Active Cosmetics
Literature data shows that low glutathione levels relates to the deposition of melanin in the skin of
humans and other animals, whereas high glutathione levels inhibit melanogenesis.40,41 It is also reported
that glutathione depletion increases tyrosinase activity in human melanoma cells.42 UVA irradiation
induces the immediate loss of reduced glutathione (GSH) in both melanocytes and keratinocytes.43
Recently, Matsuki etal. have reported that glutathione has a dose dependent inhibition on melanin syn-
thesis in the reaction of tyrosinase and L-DOPA.44 Since HR has a dose dependent increase effect on
glutathione synthesis,24 it can be concluded that HR also inhibits melanogenesis by stimulating glutathi-
one synthesis.
It has also been found that human skin exposed to UVB irradiation with a dose of 2 MED manifests a
signicant increase in the expression of Endothelin-1 (ET-1) and tyrosinase mRNA signals ve days after
irradiation.45,46 In these studies, low levels of ET-1 secreted by keratinocytes in response to UVR was
shown to down-regulate E-cadherin in melanocytic cells. ET-1 is a potent down-regulator of E-cadherin
in human melanocytes and also melanoma cells.46 An independent study by Chaudhuri has shown a
15-fold up-regulation of CDH-1 gene coding for E-cadherin as compared to a control in UVB irradiated
normal human keratinocytes treated with HR. In light of the foregoing, it is quite tempting to propose
that HR also reduces UV-induced hyperpigmentation by modulating E-cadherin.
Eller etal. have shown that DNA damage after UV irradiation enhances UV-induced melanization.47
They have also shown that the addition of small DNA fragments to un-irradiated pigment cells in vitro
or in vivo to guinea pig skin induces a pigment response indistinguishable from UV-induced tanning.
Based on the study reported by Yen etal. which found that HR reduces DNA damage induced by hydro-
gen peroxide24; it is quite conceivable to conclude that HR also inhibits melanin synthesis by reducing
DNA damage.
Taken together, the data generated by the present author and available in the literature strongly sug-
gests that HR provides for a reduction in melanin production by modulating multiple sites in the mela-
nogenesis pathway in melanocytes. This pathway and the sites of action of HR (shown by arrows) are
portrayed in Figure 7.2.
Human Clinical
In addition to the multitude of in vitro and non-human in vivo studies on the effects of HR on skin mecha-
nisms and physiology, many of which are discussed above, there are a number of human clinical studies
that have been reported. Chaudhuri has reported on the skin-lightening efcacy and safety of a lotion
containing 0.5% HR compared with a 2% hydroquinone lotion.12 This was a single-center study of 15
subjects who applied the lotions twice a day, morning and evening, for eight weeks. The test sites, as well
as untreated sites on each forearm, were evaluated for skin color change as represented by the change in
Tyrosine DOPA DOPAchrome →→ DHI and DHICA Melanin
mediated path Peroxidase/H2O2
mediated path
Tyrosinase mediated path
DNA damage induced by H2O2
Competitive binding of glutahione to tyrosinase active site
FIGURE 7.2 Sites in the melanogenesis pathway in melanocytes inhibited by HR. DOPA = 3,4-dihydroxyphenylalanine;
DOPAchrome = 5,6-dioxo-2,3,5,6-tetrahydro-1H-indole-2-carboxylic acid; DHI = 5,6-dihydroxyindole; DHICA = 5,6-
dihydroxyindole-2-carboxylic acid.
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L values and ITA° (Individual Topology Angle—COLIPA SPF test method). Changes were measured
by chromometric measurement. The results of this study are presented in Table 7.2 where the delta rep-
resents the percent change in skin color from the baseline coloration of the untreated skin. According to
these results, the HR lotion was found to provide a comparable, statistically signicant (pvalue 0.001)
change in skin color to that attained with the like composition containing 2% hydroquinone. Such results
are consistent with a “lightening” of the skin. No adverse effects were noted for either lotion over the
test period.
Another clinical study was carried out by the present author using 1% HR lotion for treating indi-
viduals having hyper-pigmented spots. Eighteen volunteers between the ages of 42 and 60, of which
there was a mix of Caucasian (10), Asian (7), and Hispanic (1), applied the 1% HR lotion twice daily
in an amount sufcient to cover their upper arm, including specically the targeted hyper-pigmented
spots. The treated skin of each volunteer was evaluated and the percent reduction in pigmentation
of the treated skin was determined using ITA degree. When compared to the coloration on day zero
(pre-application), it was found that the treated hyper-pigmented spots showed a statistically signicant
reduction of 39% and 60% in pigmentation at four and eight weeks, respectively. On the other hand, the
treated area of the skin surrounding the hyper-pigmented spots showed only marginal changes when
compared to day zero (pre-application). Based on this study, one could conclude that a 1% HR lotion
provides an even-toning effect.
Chaudhuri also investigated the potential for HR to show a synergistic effect when used in com-
bination with other skin lightening ingredients. In this study, a lotion containing 2% by weight of a
commercial blend of HR (25%) and ethyl linoleate (EL, 25%) in caprylic/capric triglycerides (trade
name Asyntra® SL) was applied twice daily for a period of four weeks to the whole face of 15 volun-
teers. As in the previous studies, skin color was determined by ITA degrees before and after the treat-
ment. The results of this study demonstrated a statistically signicant (p 0.05) lightening effect on
the natural skin color (initial ITA degree 2.1 vs. 5.8 after four weeks of treatment). The combination
was also found to be much more effective than HR alone in lightening one’s natural skin color. The
synergy of the ingredients of this blend was subsequently conrmed by carrying out an in vitro study
using B16 melanoma cells. In that study, referenced in the preceding section, an almost identical level
of reduction in melanin synthesis was observed using the HR-EL blend (contains only 25% HR) as
compared to the use of HR alone. Although the full mechanism is unknown, it has been reported that
EL is hydrolyzed to linoleic acid (LA) in vivo,48 which has been reported to degrade post-translational
tyrosinase and is also involved in the increase in cell turnover.49 Therefore, it can be presumed that, in
addition to the synergistic effect with HR, EL is also providing all of the skin benets resulting from
its conversion to LA.
Independently, Makino etal. reported on a single-center, double-blind comparison clinical study of
18 subjects in which the efcacy of a HR-containing (included several other skin lightening ingredients)
cream for reducing ultraviolet-induced hyperpigmentation was evaluated.50 Test sites were irradiated
with 1.0, 1.5, 2.0, and 2.5 minimal erythema doses. After ve days during which pigmentation was
allowed to develop, the HR-containing product or a 4% hydroquinone cream was applied to the respec-
tive test sites, once daily for four weeks. Chroma meter measurements (L* brightness) and standardized
Human Clinical Study: Skin Lightening Effects of HR vs. Hydroquinone
Skin Lightening Active Week L-value
% Improvement
Based on L-Value ITA°
% Improvement
Based on ITA°
0.5% HR 0 56.88 19.09
4 58.82 3.4 24.34 27.5
8 59.54 4.7 26.26 37.6
2.0% Hydroquinone 0 57.10 20.72
4 58.95 3.2 24.67 27.7
8 59.54 4.3 27.22 38.4
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78 Cosmeceuticals and Active Cosmetics
digital photographs were taken of the test sites twice a week. According to their ndings, areas treated
with the HR-containing product produced greater increases in L* brightness as compared with those
treated with the 4% hydroquinone cream. This study demonstrates that a product designed to affect
multiple pathways of melanogenesis and melanin distribution may provide an additional treatment option
beyond hydroquinone for hyperpigmentation.
In another study, Fantasia etal. reported on a 12-week, double-blind, split-face, randomized clinical
study in which an HR-containing lotion was applied to 41 subjects using a round-robin approach against
the vehicle, which was applied to 40 subjects, all subjects being women from 35 to 59 years of age.51
Subjects were selected specically for overall photo-damage, ne lines, and mottled pigmentation. Each
product was applied once daily to the designated half side of the face. Evaluation was done at baseline
and after 2, 4, and 12 weeks of treatment using a 1–9 scale for photo-aging parameters. The ndings
from this study revealed that the HR-containing lotion performed signicantly well as compared to the
placebo on several key anti-aging clinical parameters, such as tactile roughness, radiance, mottled pig-
mentation, crow’s feet ne lines, and overall photo-damage. Signicant clinical benets were observed
as early as two weeks, with progressive improvements at weeks 4 and 12. Such results are consistent with
the reports by these authors on the stimulatory effect of HR on collagen and elastin synthesis through
NF-kappaB inhibition.
Roure etal. reported on a double-blind, randomized, placebo controlled study on healthy women
volunteers aged over 45 years having wrinkles, sagging, spots, and a dull complexion on the face using
a HR and ascorbic acid-2-glucoside-containing formulation.28 Forty-two volunteers applied the prod-
uct and placebo in a split-face mode, twice a day for 12 weeks. Clinical grading and self-assessment
of the signs of aging, cutometric, and colorimetric measurements were performed at baseline and after
eight and 12 weeks of application. After eight weeks of daily application, a signicant improvement
of the wrinkles (crow’s feet: +17%, under eye: +19%, cheek: +10%), ne lines (crow’s feet: +9%, under
eye: +13%), brown spots (intensity: +11%, number: 8%), rmness (+26%), and skin texture (complex-
ion: 18%, homogeneity: +26%, softness: 21%, smoothness: 32%) was observed with the product versus
baseline as well as placebo. These changes were even greater after 12 weeks of product application.
Finally, in a 12-week, full face, double-blinded, randomized controlled study in Chinese subjects,
Johnson & Johnson scientists have shown signicant improvement in key pigmentation-related param-
eters without detecting any product-related adverse events.52 Out of the total study population of 65, 32
subjects using the lotion containing HR product and 31 subject using the vehicle (without HR) completed
the 12-week study. Two subjects from the vehicle group were discontinued for non-product related rea-
son. Application of HR containing product resulted in signicant clinical improvement (statistically
signicant vs. placebo; p < 0.05) on overall skin lightening (% of subjects showing improvement: 88%),
appearance of spots on the cheeks (88%), overall contrast between spots and surrounding skin (100%),
and overall pigmentation size (97%) with the vehicle at 12 weeks. Changes were determined by derma-
tologist grading, digital photos and UV images. These authors further demonstrated that de-pigmenting
effect of HR is not associated with toxicity in melanocytes and is reversible; conrming Chaudhuri’s
earlier observation.12 In contrast, the inhibition of melanin production, as expected, failed to recover in
melanocytes treated with hydroquinone. These ndings further validate the potency of HR to modulate
skin pigmentation, and its safety and tolerance for topical application.
These studies show that HR has a plurality of strong, benecial effects on skin physiology and condi-
tioning, including, but not limited to skin lightening, even toning of color, and anti-aging effects.
Having demonstrated the benets of HR as a skin treatment, HR also benets from an ease of use in
terms of formulating products, especially skin care and cosmetic products. This is especially so at use
levels of 0.5%–1.0% (w/w) of HR in the nished formulation, which have been found to provide effective
HR is highly soluble (>20%) in a number of cosmetic and skin care ingredients and carriers includ-
ing caprylic/capric triglycerides, isosorbide dicapylate, ethyl linoleate, ethoxydiglycol, and other high
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polarity esters; non-ionic solubilizers; glycerol; and a wide range of glycols. PEG-400 was found to be
an excellent solubilizer of HR (>50%). Such solubility provides a wide range of options to develop elegant
formulations with HR. HR can also be easily combined with other skin lightening ingredients, such
as niacinamide, ethyl linoleate, acetyl-glucosamine, ascorbyl-2-glucoside (not magnesium and sodium
ascorbyl phosphate as these two requires slightly basic formulation pH), standardized plant extracts,
such as Phyllathus emblica (trade named Emblica®), Terminalia chebula (trade named Synastol® TC),
etc., thereby opening the door to a multitude of synergistic products to make HR even more effective.
Generally, the pH of the formulation to which HR is added must be acidic due to the presence of
phenolic OH in HR. Similarly, it is found that the use of non-ionic emulsiers is preferable to anionic
types. For the preparation of serum or transparent gels, the use of non-ionic solubilizers having high
HLB values like PEG-40 hydrogenated castor oil, Laureth 23, Polysorbate 20, and Polysorbate 80 are
Finally, the addition of HR to various formulations may cause a drop in overall viscosity of the prod-
uct. In such cases, anionic (such as Xanthan gum, Carbomers) or neutral thickeners (such as Cellulosics)
can be added for maintaining and/or restoring the desired viscosity. If needed, low levels 0.1%–0.2% of
propyl gallate (SynoxylÒ PGL) can be used to minimize color shift over time.
Although the present author endeavored to identify all relevant articles and dates, it is nonetheless
acknowledged that some will have been overlooked, for which the author apologizes. Nevertheless, this
chapter provides ample evidence of the complex physiological actions of HR and its ability to modu-
late different biological processes and pathways through multiple molecular targets. Key targets include
those associated with skin pigmentation, inammation and inammatory responses, extracellular matrix
proteins, and the like. Such activity clearly points to the realized and yet to be realized signicance of
HR in providing a number of skin benets of commercial interest. Already, commercial use of HR in a
wide variety of skin care and treatment products has begun and is expanding. Additionally, further work
exploring and eshing out the impact of HR on tumors will likely lead to the establishment of HR as a
key ingredient in combination therapy. In summary, it is clear from the current literature that HR has
numerous targets, and provides its biological effects through simultaneous modulation of these targets.
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... In recent years, a number of pharmacological studies have been made on alkylresorcinols, which comprise a family of hydrophobic, non-isoprenoid phenolic lipids naturally synthesised by higher plants [22]. The synthetic derivative 4-hexylresorcinol (4-HR, Table 1) is reported to have anesthetic, antiseptic, and antihelmintic properties and is widely used as topical drug in antibacterial oral gargles and throat lozenges [23]. The 4-HR inhibits human transglutaminase 2, the NF-κB pathway, and intracellular calcium oscillations, thus making it a promising anti-cancer agent showing synergistic effects with cisplatin [24]. ...
... Suppression of NF-κB phosphorylation and inhibition of foreign-body giant cell formation is associated with anti-inflammatory effects of 4-HR [25]. Additionally, 4-HR has been described as a potent inhibitor of tyrosinase and polyphenol oxidase, thereby inducing antioxidant effects that are utilised in anti-ageing or skin lightning cosmetics, as well as in food processing aids to avoid melanosis in shrimps [23]. Based on a long history of human safe use, 4-HR is FDA-listed as active ingredient for over-the-counter products and is approved as a food additive (E 586) in the European Union [26]. ...
... All other tested microbial strains displayed similar degrees of susceptibility with a median MIC of 7.5% (w/w), independently of the Gram status or the fungal kingdom. A previous investigation of the antimicrobial activity of 4-HR solutions yielded MIC values ranging from 0.3 µg/mL to 50 µg/mL against various bacteria and fungi relevant to personal care applications [23]. Due to differences in the test methods, it is impossible to compare this data with the effectiveness of PLGA-incorporated 4-HR concentrations in our study. ...
Full-text available
Profound screening and evaluation methods for biocide-releasing polymer films are crucial for predicting applicability and therapeutic outcome of these drug delivery systems. For this purpose, we developed an agar overlay assay embedding biopolymer composite films in a seeded microbial lawn. By combining this approach with model-dependent analysis for agar diffusion, antimicrobial potency of the entrapped drug can be calculated in terms of minimum inhibitory concentrations (MICs). Thus, the topical antiseptic 4-hexylresorcinol (4-HR) was incorporated into poly(lactic-co-glycolic acid) (PLGA) films at different loadings up to 3.7 mg/cm2 surface area through a solvent casting technique. The antimicrobial activity of 4-HR released from these composite films was assessed against a panel of Gram-negative and Gram–positive bacteria, yeasts and filamentous fungi by the proposed assay. All the microbial strains tested were susceptible to PLGA-4-HR films with MIC values down to 0.4% (w/w). The presented approach serves as a reliable method in screening and quantifying the antimicrobial activity of polymer composite films. Moreover, 4-HR-loaded PLGA films are a promising biomaterial that may find future application in the biomedical and packaging sector.
... It is an organic amphiphilic compound that mimics the tyrosinase natural substrate, tyrosine. It is an analog of resorcinol, a compound used as an antiseptic and disinfectant in topical use [1]. It has amphiphilic properties, possessing a hydrophilic moiety and a hydrophophic moiety (highlighted in green circle in its structure Figure 2). ...
... It is an organic amphiphilic compound that mimics the tyrosinase natural substrate, tyrosine. It is an analog of resorcinol, a compound used as an antiseptic and disinfectant in topical use [1]. It has amphiphilic properties, possessing a hydrophilic moiety and a hydrophophic moiety (highlighted in green circle in its structure Figure 2). ...
Full-text available
4-hexylresorcinol (4-HR), as a derivative of phenolic lipids, has biological and pharmacological properties that are beneficial when used with a biomaterial. It has antimicrobial and antiseptic activity and can thus prevent contamination and infection of biomaterials. 4-HR suppresses the nuclear factor kappa B (NF-κB) signaling pathway related to osteoclast differentiation. The suppression of NF-κB increases the bone formation marker and contributes to new bone formation. The tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine produced by macrophages and suppressed by 4-HR. Suppression of TNF-α decreases osteoclast activity and promotes wound healing. 4-HR increases the vascular endothelial growth factor and has an anti-thrombotic effect. When incorporated into silk vascular patches, it promotes endothelium wound healing. Recently, 4-HR has exhibited biological properties and has been successfully incorporated into various biomaterials. Consequently, it is a useful pharmacological chemical that can be used with biomaterials in the field of tissue engineering.
Full-text available
Objective: For the majority of people with acute sore throat, over-the-counter treatments represent the primary option for symptomatic relief. This study evaluated the in vitro bactericidal activity of lozenges containing the antiseptic hexylresorcinol against five bacteria associated with acute sore throat: Staphylococcus aureus, Streptococcus pyogenes, Moraxella catarrhalis, Haemophilus influenzae and Fusobacterium necrophorum. Results: Hexylresorcinol 2.4 mg lozenges were dissolved into 5 mL of artificial saliva medium. Inoculum cultures were prepared in triplicate for each test organism to give an approximate population of 108 colony-forming units (cfu)/mL. Bactericidal activity was measured by log reduction in cfu. Greater than 3log10 reductions in cfu were observed at 1 min after dissolved hexylresorcinol lozenges were added to S. aureus (log10 reduction cfu/mL ± standard deviation, 3.3 ± 0.2), M. catarrhalis (4.7 ± 0.4), H. influenzae (5.8 ± 0.4) and F. necrophorum (4.5 ± 0.2) and by 5 min for S. pyogenes (4.3 ± 0.4). Hexylresorcinol lozenges achieved a > 99.9% reduction in cfu against all tested organisms within 5 min, which is consistent with the duration for a lozenge to dissolve in the mouth. In conclusion, in vitro data indicate that hexylresorcinol lozenges offer rapid bactericidal activity against organisms implicated in acute sore throat.
Objective: 4-n-butylresorcinol is a competitive inhibitor of tyrosinase and has been used as an anti-melanogenic agent. However, its inhibition mechanism in intact cells is not fully understood. To elucidate the cellular mechanism, we compared in vitro and in vivo inhibitory effects of 4-n-butylresorcinol on tyrosinase activity. Methods: B16F10 melanoma cells were cultured in media containing ?MSH in the presence or absence of 4-n-butylresorcinol. Tyrosinase mRNA levels, protein levels, and activity in B16F10 cells were compared by real-time PCR, immunostaining combined with western blot, and colorimetric analysis, respectively. Melanin concentration was measured by colorimetry both in the cells and in the media. Tyrosinase glycosylation and proteolytic degradation were analyzed by immunoblotting after cells were treated with Endo H/PNGase and E64/proteasome inhibitors, respectively. Results: 4-n-butylresorcinol inhibited tyrosinase activity and melanin synthesis more effectively in intact cells than in cell lysates. Western blotting and real-time RT-PCR showed that 4-n-butylresorcinol reduced protein levels, but not mRNA levels, of tyrosinase in B16F10 cells. 4-n-butylresorcinol showed no effect on the processing of tyrosinase glycosylation or on trafficking to melanosomes. However, treatment of B16F10 cells with E64 or proteasome inhibitor abrogated the 4-n-butylresorcinol-induced decrease of tyrosinase. Moreover, 4-n-butylresorcinol activated p38 MAPK, resulting in increased ubiquitination of tyrosinase. Conclusion: 4-n-butylresorcinol inhibits melanogenesis by enhancing proteolytic degradation of tyrosinase as well as competitive binding to tyrosinase. These findings will help to develop new, effective, and safe chemicals for the treatment of hyperpigmentation disorders. This article is protected by copyright. All rights reserved.
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DNA topoisomerases are highly exploited targets for antimicrobial drugs. The spread of antibiotic resistance represents a significant threat to public health and necessitates the discovery of inhibitors that target topoisomerases in novel ways. However, the traditional assays for topoisomerase activity are not suitable for the high-throughput approaches necessary for drug discovery. In this study we validate a novel assay for screening topoisomerase inhibitors. A library of 960 compounds was screened against DNA gyrase and archaeal DNA topoisomerase VI. Several novel inhibitors were identified for both enzymes, and subsequently characterised and . Inhibitors from the topoisomerase VI screen were tested for their ability to inhibit Arabidopsis topoisomerase VI in planta. The data from this work present new options for antibiotic drug discovery and provide insight into the mechanism of topoisomerase VI.
Full-text available
The bacterial dormancy-inducing factor 4-hexylresorcinol (4-HR) has been shown to have synergistic antitumor effects when used in combination with cisplatin. In the present study, 4-HR was used as a single agent in the squamous carcinoma cell line SCC-9. The results demonstrated that 4-HR suppressed SCC-9 cell proliferation compared to primary cultured gingival fibroblasts. 4-HR dose-dependently induced SCC-9 cell apoptosis as determined by caspase-3 activity, annexin V expression, as well as by scanning and transmission electron microscopy. 4-HR inhibited intracellular calcium oscillation in both SCC-9 cells and normal human dermal fibroblasts. 4-HR-induced apoptosis was partly reversed by calcium channel blockers. Of note, 4-HR reduced the tumor mass formed by SSC-9 cell implantation in BALB/cAnNCrj-nu/nu mice and mass size reduction was also partly reversed by the concomitant application of calcium channel blockers. Collectively, our results suggest that 4-HR has strong antitumor effects by inhibiting calcium channel oscillation and inducing apoptosis.
Full-text available
The present study was undertaken to examine the effects of cisplatin plus 4-hexylresorcinol (4-HR) combination therapy on oral mucosal melanoma (OMM) using cultured primary OMM cells in a tumour xenograft model. Cultured primary OMM cells were used for the MTT assay and DNA microarray. OMM cells were implanted into the submandibular glands of nude mice. The mice were then treated with cisplatin only or cisplatin plus 4-HR. Tumour size changes, survival rate and tumour metastasis were compared between the two groups by observation, micro-positron emission tomography (PET) and histological examination. In the MTT assay, the cisplatin plus 4-HR group showed significantly higher inhibition of OMM cell growth compared to the other groups (p < 0.05). DNA microarray results showed significant inhibition of matrix metalloproteinase (MMP)-2 gene expression upon 4-HR application. The necropsy and micro-PET results showed that the mice from the cisplatin-only group had more distant metastases than the mice from the cisplatin plus 4-HR combination group (p = 0.002). MMP-2 expression was lower in the primary tumours in the cisplatin plus 4-HR combination group than in the cisplatin-only group (p < 0.001). Overall survival was longer in mice from the cisplatin plus 4-HR combination group than in the cisplatin-only group (p = 0.049). In conclusion, the combined effect of cisplatin and 4-HR resulted in fewer metastases and longer survival than cisplatin-only treatment in the OMM xenograft model.
Hyperpigmentation disorders are of social and cosmetic concerns to many individuals due to their prevalent locations on highly visible parts of the body. Topical formulation containing hydroquinone is the most widely used remedy for the treatment of hyperpigmentation disorders. However, reports of side effects in long-term usage have raised concerns for its use in cosmetic products. Thus, it is highly desirable to develop a safe and effective alternative to treat hyperpigmentation disorders. The objective of the current study is to investigate the de-pigmenting efficacy of 4-hexyl-1,3-phenylenediol in various in vitro models and in a randomized controlled clinical study. We showed that 4-hexyl-1,3-phenylenediol significantly reduced melanogenesis in primary human melanocytes, murine melanoma cells, and pigmented human epidermal equivalents. It was determined that the reduction in melanogenesis is mediated through inhibition of tyrosinase enzyme activity and protein expression. Further investigation revealed that the inhibition of melanogenesis is reversible and is not associated with cellular toxicity in melanocytes. In addition, significant improvements in key clinical parameters such as overall skin lightening, appearance of spots on the cheeks, overall contrast between spots and surrounding skin, and overall pigmentation size were detected in a double-blinded, randomized controlled clinical study. In conclusion, our findings clearly demonstrated the potency of 4-hexyl-1,3-phenylenediol in modulating skin pigmentation, and it is safe and well tolerated after 12-week topical application.
The use of sulphites for the prevention of browning on fruits and vegetables is banned in the USA. The possible alternative use of ascorbic acid (AA), 4-hexylresorcinol (HR), and a mixture of both on apple slices was studied taking quality retention as a function of storage time as compared with similar products treated with sulphites. The mixture of ascorbic acid and 4-hexylresorcinol (AH) significantly inhibited the browning of vacuum packaged apple slices for up to 8 weeks during storage at 0.5 °C (p ≤ 0.001). Browning inhibition by AH was comparable to that achieved by sulphite and stronger than that of ascorbic acid (AA). In addition AH did not bleach the fruit tissue and therefore the colour of AH treated fruits was closer to the fresh apple pieces than those treated with sulphite. AH also reduced the total viable microbial counts by more than 10-fold as compared with AA treated and control apple pieces. AH treated apple pieces had a slightly firmer texture than the control and there were no adverse effects on the soluble solids and titratable acidity of the fruit. The evidence suggests that AH is a promising anti-browning formula for mini mally processed apples.
BACKGROUND and Skin lightening preparations are used by people all over the world for a diverse range of dermatologic indications. Hydroquinone (HQ) is the gold standard and remains the only prescription product available in the United States for the treatment of generalized facial hyperpigmentation. Irritation and the risk of exogenous ochronosis are the main adverse effects for concern. Therefore, there has been a constant search for new treatment alternatives. Understanding the molecular mechanisms involved in pigmentation has resulted in the development of a series of formulations that utilize a multimodal treatment approach. These proprietary formulas combine skin lightening agents that act via different mechanisms of action. The actives included 4-ethoxybenzaldehyde (anti-inflammatory and prostaglandin E2 suppressor), licorice extract (tyrosinase inhibitor), tetrahexyldecyl ascorbate (antioxidant), niacinamide (melanosome transport inhibitor), ethyl linoleate (tyrosinase inhibitor; enhances turnover of epidermis), hexylresorcinol (tyrosinase inhibitor), and retinol (tyrosinase transcription inhibitor; enhances turnover of epidermis). Select formulations were tested in several studies using the MelanoDerm™ Skin Model (MatTek Corporation, Ashland, MA) to assess the ability of the product to reduce melanin production and distribution. A single-center, double-blind comparison clinical study of 18 subjects was conducted to evaluate the efficacy of the product in reducing ultraviolet-induced hyperpigmentation. Test sites were irradiated with 1.0, 1.5, 2.0, and 2.5 minimal erythema doses. After 5 days, to allow for pigmentation development, the product or 4% HQ cream was applied to the respective test sites, once daily for 4 weeks. Chroma Meter measurements (L* brightness) and standardized digital photographs were taken of the test sites twice a week. The test product resulted in greater reduction in melanin as measured by melanin content and histological staining compared with the positive control in the MelanoDerm Skin Model. The product also demonstrated statistically significant reductions in pigmentation compared with baseline (all P≤.0001) at the end of the clinical study, and produced greater increases in L*, compared with 4% HQ. Results from these studies indicate that a product designed to affect multiple pathways of melanogenesis and melanin distribution may provide an additional treatment option beyond HQ for hyperpigmentation. J Drugs Dermatol. 2013;12(3 suppl 1):s16-s20.
Humans skin is the most visible aspect of the human phenotype. It is distinguished mainly by its naked appearance, greatly enhanced abilities to dissipate body heat through sweating, and the great range of genetically determined skin colors present within a single species. Many aspects of the evolution of human skin and skin color can be reconstructed using comparative anatomy, physiology, and genomics. Enhancement of thermal sweating was a key innovation in human evolution that allowed maintenance of homeostasis (including constant brain temperature) during sustained physical activity in hot environments. Dark skin evolved pari passu with the loss of body hair and was the original state for the genus Homo. Melanin pigmentation is adaptive and has been maintained by natural selection. Because of its evolutionary lability, skin color phenotype is useless as a unique marker of genetic identity. In recent prehistory, humans became adept at protecting themselves from the environment through clothing and shelter, thus reducing the scope for the action of natural selection on human skin.
Evidence for the postulated role of glutathione reductase in melanin pigmentation has been obtained by determinations of the glutathione concentrations in Tortoiseshell guinea pig skin of different colors (black, yellow, red, and white). As expected, the lowest levels of reduced glutathione (GSH) were found associated with eumelanin type pigmentation, whereas the highest ones were found in the skin with phaeomelanin producing melanocytes. On the other hand, white skin of guinea pig having no active melanocytes showed GSH levels which were intermediate between those of the black and yellow areas. These results are consistent with the view that the activity of the enzyme glutathione reductase, though not primarily related to pigmentation, plays an important role in the regulation and control of the biosynthetic activity of melanocytes leading to various types of melanin pigments.
The significance of our understanding of the chemistry of melanin and melanogenesis is reviewed. Melanogenesis begins with the production of dopaquinone, a highly reactive o-quinone. Pulse radiolysis is a powerful tool to study the fates of such highly reactive melanin precursors. Based on pulse radiolysis data reported by Land et al. (J Photochem Photobiol B: Biol 2001;64:123) and our biochemical studies, a pathway for mixed melanogenesis is proposed. Melanogenesis proceeds in three distinctive steps. The initial step is the production of cysteinyldopas by the rapid addition of cysteine to dopaquinone, which continues as long as cysteine is present (1 μM). The second step is the oxidation of cysteinyldopas to give pheomelanin, which continues as long as cysteinyldopas are present (10 μM). The last step is the production of eumelanin, which begins only after most cysteinyldopas are depleted. It thus appears that eumelanin is deposited on the preformed pheomelanin and that the ratio of eu- to pheomelanin is determined by the tyrosinase activity and cysteine concentration. In eumelanogenesis, dopachrome is a rather stable molecule and spontaneously decomposes to give mostly 5,6-dihydroxyindole. Dopachrome tautomerase (Dct) catalyses the tautomerization of dopachrome to give mostly 5,6-dihydroxyindole-2-carboxylic acid (DHICA). Our study confirmed that the role of Dct is to increase the ratio of DHICA in eumelanin and to increase the production of eumelanin. In addition, the cytotoxicity of o-quinone melanin precursors was found to correlate with binding to proteins through the cysteine residues. Finally, it is still unknown how the availability of cysteine is controlled within the melanosome.