Gel vehicles are not inherently more irritating than creams

BACKGROUND: It has been thought that topical gels are inherently more irritating than topical creams. Nevertheless, the irritancies of topical products are potentially quite variable, and a priori assumptions about relative irritancy of gels versus creams may not be accurate. PURPOSE: To determine whether a metronidazole gel formulation is more or less irritating to the skin compared to metronidazole creams. METHODS: A total of 32 normal, healthy volunteers were tested using irritancy patches with 0.75% metronidazole gel and cream, 1% metronidazole cream, and petrolatum (used as the "negative control"). Blinded observers evaluated the application sites for signs of irritancy. A numerical score was assigned to these irritancy patch sites each day for 21 days, or until significant irritation developed, and cumulative irritancy scores were calculated for the study period. A mixed model of variance analysis was performed. RESULTS: After 21 days of evaluation, analysis of the mean cumulative irritancy scores for each of the agents used showed there to be no statistical difference in irritancy potential between the metronidazole gel and the metronidazole creams. However, the 1% metronidazole cream was significantly more irritating than petrolatum. CONCLUSION: There was no significant difference in the cumulative irritancy potential of cream and gel preparations of metronidazole. The irritancy of topical formulations for treating rosacea should be considered on a case by case basis.     

Sodium lauryl sulfate-induced irritation in the human face: regional and age-related differences

The particular sensitivity of the human face to care products prompted us to study irritation induced by sodium lauryl sulfate (SLS) in its various regions. We examined regional and age-related differences, correlating basal transepidermal water loss (TEWL) and capacitance to SLS irritation. SLS (2% aq.) was applied under occlusion for 1 h to the forehead, cheek, nose, nasolabial and perioral areas, chin, neck and forearm to two groups of subjects--one with 10 subjects with an average age of 25.2 +/- 4.7 years and another with 10 subjects with an average age of 73.7 +/- 3.9 years. TEWL was measured before and 1 h and 23 h after patch removal. Baseline stratum corneum hydration was also measured. Irritation was assessed by the changes in TEWL (deltaTEWL = TEWL after patch removal - basal TEWL) after corrections to the control. In the younger group, all areas of the face and the neck reacted to SLS, whereas the forearm did not. In the older group, the nose, perioral area and forearm did not react. In both age groups, some significant differences between the regions of the face were detected. The younger group showed higher changes in TEWL than the older group in all the areas studied, but only in the chin and nasolabial area were the differences statistically significant. Significant correlations were found between basal TEWL and deltaTEWL in 5 of the 7 areas which reacted to SLS. Baseline TEWL is one parameter that correlates with the susceptibility of the face to this irritant.     

Comparison of tissue sources for the skin integrity function test (SIFT).

One of the in vitro models involved in an ECVAM-sponsored prevalidation study for acute skin irritation is the skin integrity function test (SIFT), which utilises full-thickness mouse skin. We have evaluated nine different skin types in order to identify the most useful model for assessing skin barrier function using transepidermal water loss (TEWL), electrical resistance (ER) and tritiated water flux (TWF) and sodium lauryl sulphate (SLS) as a standard skin irritant. Tissues were: human skin (epidermis and whole), reconstituted human epidermis (RHE), pig (dermatomed and whole), rabbit (whole), rat (epidermis and whole) and mouse (whole). Barrier function was measured following sodium lauryl sulphate (SLS) exposure and expressed as a damage ratio. Human epidermis gave good responses at high doses of SLS only. RHE had abnormally high permeability to water and therefore had little or no response to SLS. Pig skin gave low TEWL ratios and rabbit skin was a poor responder to SLS. Mouse whole skin performed best in this study, giving consistent high damage ratios to TEWL, ER and TWF following SLS treatment. Rat whole skin also performed well but was generally less responsive. We conclude that mouse skin is the best and most practical in vitro model for the SIFT approach for skin irritation prediction.     

Itch-associated response induced by experimental dry skin in mice

The present study was conducted to establish a new mouse model of dry skin pruritus. The rostral back was treated daily with cutaneous application of acetone/ether (1:1) mixture (AE), water following AE (AEW), 1% sodium lauryl sulfate (SLS) or tape stripping (TS). On the day after 5-day treatment, although all four treatments significantly decreased stratum corneum (SC) hydration and increased transepidermal water loss (TEWL), only AEW treatment significantly increased spontaneous scratching. An increase in the frequency of TS produced the marked increase of TEWL, without significant effects on SC hydration and spontaneous scratching. In AEW-treated mice, changes in SC hydration and TEWL were marked in the initial 2-day period, while spontaneous scratching increased gradually from 3 days after starting the treatment. The degranulation of cutaneous mast cells was increased by SLS treatment but not by other treatments. There was no apparent difference in AEW-induced spontaneous scratching between mast cell-deficient mice (WBB6F1-W/Wv) and normal littermates (WBB6F1-+/+). Opioid antagonists, naloxone and naltrexone, (1 mg/kg, subcutaneously) significantly suppressed spontaneous scratching in AEW-treated mice. It is suggested that spontaneous scratching of AEW-treated mice is an itch-related response and a useful model for studying the mechanisms of dry skin pruritus.     

Prevention of experimentally induced irritant contact dermatitis by extracts of Isatis tinctoria compared to pure tryptanthrin and its impact on UVB-induced erythema

Lipophilic extracts of Isatis tinctoria L. exhibit significant activity against several clinically relevant targets of inflammation. The alkaloid tryptanthrin was identified as one of the active principles in woad and characterised as a potent dual inhibitor of COX-2 and 5-LOX. Here, the anti-inflammatory efficacy of topical application of three different Isatis extracts and tryptanthrin was investigated in human volunteers. Two different models were used, namely the sodium lauryl sulphate (SLS)-induced irritant contact dermatitis (ICD) and UVB-induced erythema. Twenty healthy volunteers without any skin disease participated in the study. Cumulative irritant contact dermatitis was induced on test fields on the volunteers' backs by twice daily application of 0.5 % sodium lauryl sulphate over a period of four days. Half of the test fields were treated with the test substances during the eliciting phase, while the remaining test fields were treated over a period of 4 days after induction of dermatitis. In the second model, a UVB erythema on the volunteers' lower backs was induced using the double minimal erythema dose (MED). Twenty-four hours after irradiation the test fields were treated with the test substances over a period of 3 days. All reactions were assessed visually and by non-invasive bioengineering methods (evaporimetry and chromametry). Treatment with extracts during the ICD eliciting phase led to a significantly smaller increase of visual scores and transepidermal water loss compared to the untreated test field. For tryptanthrin this benefit was also observed, but the improvement was not statistically significant. When treatment was performed after completing the eliciting phase, accelerated resolution of the irritant reaction could not be observed. In the UVB erythema model anti-inflammatory effects of the test substances were not observed.     

In vitro cytotoxicity tests on cultured human skin fibroblasts to predict skin irritation potential of surfactants.

Cultured human skin cells are a potentially useful model for skin irritancy testing. We have investigated the use of human skin fibroblasts for in vitro screening for skin toxicity. To assess the cytotoxic effects of surfactants, cell viability was measured by the NRU (neutral red uptake) assay and AB (Alamar blue) assay as in vitro methods. The skin irritation potential of surfactants by human skin patch test was assessed as in vivo methods. The close relationship was found between AB assay with human skin fibroblasts and human patch test (r=0.867). There was a relatively good agreement between the NRU and in vivo patch test (r=0.648). These results suggest that AB and NRU assay using cultured human fibroblast could be predictable methods for the irritancy of various surfactants in human.     

O/W emulsions compromise the stratum corneum barrier and improve drug penetration

BACKGROUND: W/O emulsions improve the stratum corneum barrier, while microemulsions tend to compromise it. We, therefore, were interested to explore the effects of O/W emulsions on the stratum corneum barrier. METHODS: Aqueous Cream BP 2001, Clioquinol Cream BP 1999 without clioquinol, Nonionic Hydrophilic Cream DAB 2001 without glycerol, Hydrophilic Skin Emulsion Base NRF S. 25., point of time 2001, without glycerol, and Base Cream DAC were tested versus untreated controls in 29 healthy volunteers for 7 days. Outcome measures included transepidermal water loss (TEWL), skin redness (chromametry a*-value) and erythrocyte circulation in the subpapillary vessels (laser Doppler). Barrier compromise was subsequently explored by performing the hydrocortisone blanching test using Hydrocortisone Cream 0.5% NRF 11.36. (outcome measure: a*-value) in 15 subjects and the sodium lauryl sulfate (SLS) irritation test (outcome measures: TEWL, a*-value, laser Doppler) in 14 subjects. RESULTS: Pretreatment with the test emulsions produced increases in TEWL (statistically significant for all test emulsions), a*-value (statistically significant for Aqueous Cream BP 2001 and Base Cream DAC), and laser Doppler value (statistically significant for all emulsions except Base Cream DAC). Hydrocortisone penetration was statistically significantly increased with all test emulsions versus untreated contols. SLS irritation was mostly statistically significantly increased versus untreated controls when analyzing the study endpoint-baseline difference. CONCLUSIONS: O/W emulsions may compromise the stratum corneum barrier and improve drug penetration.     

Dual Characteristics of Skin Care Creams Evaluated by Two In-Vivo Human Experimental Models

The use of skin care creams is a well documented protection measure to reduce the risk of barrier damage and contact dermatitis from exogenous contact with skin irritants. Before choosing a skin care cream two aspects should be considered: a) Is the product able to reduce irritant reactions caused by the irritant, and b) is the product well tolerated, also on damaged skin. Both aspects can be evaluated by experimental models in human volunteers. We used two standard experimental designs to compare six commercially available skin care products: a) the chamber scarification test, designed to assess the irritancy potential, and b) the repeated short-time occlusive irritation test (ROIT), developed to evaluate the efficacy of skin care creams. The results showed that a high score in the chamber scarification test for skin irritation was not necessarily correlated to the products' ability to impede sodium lauryl sulfate (SLS)-induced irritant skin reactions. Three products were shown to both have a low irritancy potential and be capable of reducing skin barrier damage induced by SLS, and one product had both an irritant potential on scarified skin and also a modest capability to reduce skin irritation induced by SLS. The use of both test methods, chamber scarification and ROIT, gives valuable information on skin compatibility and efficacy of skin care creams. The clinical relevance of the test results can only be determined by comparing products with high and low scores in both tests in controlled clinical experiments with subjects at risk of developing irritant contact dermatitis.     

Unexpected skin barrier influence from nonionic emulsifiers

Skin disorders are often treated with creams containing various active substances. The creams also contain emulsifiers, which are surface-active ingredients used to stabilize the emulsion. Emulsifiers are potential irritants and in the present study the influence of stearic acid, glyceryl stearate, PEG-2, -9, -40, and -100 stearate, steareth-2, -10 and -21 on normal as well as on irritated skin have been evaluated with non-invasive measurements. Test emulsions were created by incorporating 5% emulsifiers in a water/mineral oil mixture (50:50). The emulsions and their vehicle were then applied to normal skin for 48 h and to sodium lauryl sulfate (SLS) damaged skin for 17 h in aluminum chambers. Twenty-four hours after removal of the chambers the test sites were evaluated for degree of irritation. In normal skin, the emulsifiers induced significant differences in TEWL but not in skin blood flow. Five of the emulsifiers increased TEWL. In SLS-damaged skin an aggravation of the irritation was expected. However, no differences regarding skin blood flow was noted from the emulsifiers. Furthermore, three emulsifiers unexpectedly decreased TEWL. These results highlight the possibility of absorption of these emulsifiers into the lipid bilayer, which increase TEWL in normal skin and decrease TEWL in damaged skin.     

Effects of UVB radiation on 4-hydroxy-2-trans-nonenal metabolism and toxicity in human keratinocytes

Endogenous lipid peroxidation (LPO)-derived aldehydes accumulate in human skin after photoexposure and contribute to the development of skin cytotoxicity and cancer. This study employed LC-ESI-MS and HPLC-UV-DAD techniques to investigate the effect of UVB radiation on the biotransformation and detoxification of the prototype aldehyde 4-hydroxy-2-trans-nonenal (HNE) using the human keratinocyte cell line (NCTC 2544). In parallel we followed the keratinocytes' cytotoxic response to HNE through morphological analysis and cell viability assay. In UVB-unstressed keratinocytes, even a supraphysiological dose of the aldehyde (200 microM) was rapidly and completely cleared in metabolized form (free and GSH-conjugated metabolites) from the cell, with no signs of cytotoxicity. By contrast, UVB preexposure already at 1 MED (50 mJ/cm2, the minimal erythemal dose in humans) markedly impaired HNE metabolism. After 2 h of incubation, the relative amount of GSH-conjugated adducts dose-dependently dropped from 44% (unirradiated cells) to 22% at 3 MED as a consequence of UVB-induced GSH depletion (no impairment of GST A4.4 nor of G6PD activities was observed). The levels of free metabolites, 1,4-dihydroxy-trans-nonene (DHN) and 4-hydroxy-trans-2-nonenoic acid (HNA), were modified (+30% DHN, -22% HNA) only at 3 MED, in parallel to the AR and ALDH enzyme activity modulation. In addition, a dose-dependent increase of unmodified HNE was found in the extracellular medium, paralleled by a significant fraction of the HNE-incubated dose not recovered at the intra- or extracellular level. The impairment of HNE metabolism paralleled a dramatic cytotoxic response. These results provide a reasonable explanation for the massive accumulation of carbonyl toxins in human skin in vivo after photoexposure and shed light on the detrimental effects of UVB radiation in the presence of unmetabolized LPO metabolites.     

The effects of serine palmitoyltransferase inhibitor, ISP-I, on UV-induced barrier disruption in the stratum corneum

We examined the effects of ISP-I (myriocin, thermozymocidin) - a potent inhibitor of serine palmitoyltransferase (SPT) which is involved in the ceramide synthetic pathway-on skin barrier function in post-UVB-irradiated hairless mouse skin. Disruption of the skin barrier function after UVB irradiation as represented by the increase in transepidermal water loss (TEWL) was significantly suppressed with ISP-I treatment. In the ISP-I-treated skin, the peak of cell proliferation was observed 24 h earlier than in vehicle-treated skin. In addition, the number of apoptotic cells in ISP-I-treated skin showed a sharp decrease at 48 and 72 h post-irradiation. The number of stratum corneum cell layers was increased in ISP-I-treated skin at 72 h after UVB irradiation; at this time, TEWL in ISP-I-treated skin was lower than that in the vehicle-treated skin. We suggest ISP-I treatment altered cell proliferation and apoptosis after UVB exposure by modulating ceramide synthesis in epidermal cells, resulting in an increase of stratum corneum layers which lessened the effects of irradiation-induced barrier disruption.     

A prevalidation study on the in vitro skin irritation function test (SIFT) for prediction of acute skin irritation in vivo: results and evaluation of ECVAM Phase III.

A prevalidation study sponsored by the European Centre for the Validation of Alternative Methods (ECVAM) on in vitro tests for acute skin irritation is aimed at identifying non-animal tests capable of discriminating irritants (I) from non-irritants (NI), as defined according to European Union and OECD. This paper reports on Phase III for one of the methods, the skin integrity function test (SIFT), assessing the protocol performance of the SIFT, in terms of reproducibility and predictive ability, in three laboratories. The barrier function properties of excised mouse skin were determined using a set of 20 coded chemicals (10 I, 10 NI), using the endpoints of trans-epidermal water loss (TEWL) and electrical resistance (ER). The basis of the SIFT prediction model is if the ratios of the pre- and post-application values for either TEWL or ER are greater than five-fold, then the test chemical is deemed irritant (I). If the ratio of both parameters is less than five-fold then the chemical is deemed non-irritant (NI). Analysis of variance (ANOVA) indicated that the intra-lab reproducibility was acceptable but that the inter-lab reproducibility was not. Overall, the SIFT test under-predicted the irritancy of the test chemicals chosen for Phase III with an overall accuracy of only 55%. The sensitivity value (ability to correctly predict I) was only 30%. The specificity (ability to predict NI) of the test was better at 80%. A retrospective examination of the SIFT results was undertaken using Student's t-test and a significance level of P<0.05 to predict an irritant based on changes in the TEWL ratio values. This improved the predictivity of the SIFT test, giving a specificity of 60%, a sensitivity of 80% and an overall accuracy of 70%. Appropriate modifications to the prediction model have now been made and the SIFT will be re-examined in a new validation exercise to investigate the potential of this non-animal method to predict acute skin irritation potential.     

表皮生长因子冻干粉对敏感性皮肤屏障功能 改善作用的临床观察

目的 评价外用含有表皮生长因子（EGF）的护肤品对敏感性皮肤生理功能的影响及不良反应。方法 首 先受试者接受斑贴试验检测接触过敏情况,对30例敏感性皮肤受试者面部外用含有EGF的护肤品,受试者早晚洁面 后按推荐剂量（0.4~0.5 mL）每日2次,连续使用28 d。在治疗前及治疗后7 、14、21、28 d分别对受试者皮肤生理功能 进行检测并做乳酸试验,同时对受试者皮损改善情况及安全性进行评价。结果 30例斑贴试验结果均为阳性,阳性 项目前3位的是硫柳汞、芳香混合物、重铬酸钾。与治疗前相比,治疗后14 d患者面部含水量、油脂含量开始呈增加 趋势,pH值呈减小趋势（P＜0.05）;而经皮水流失情况在治疗后21 d,乳酸试验和皮损改善评分在治疗 7 d起呈减少 趋势（P＜0.05）。治疗前,治疗后7、14、21、28 d的阳性率呈逐渐降低趋势（P＜0.001）,28 d后乳酸刺激试验仍为阳性 的7例受试者中2例为硫酸镍（++）、2例1项（+）、1例4项（+）、2例5项（+）。皮损改善率呈增加趋势（P=0.058）,斑贴 试验示治疗28 d皮损仍无明显改善者的4例中2例5项（+）、2例4项（+）。治疗前后均未发生不良反应,无一例患者 脱落。结论 含有EGF的护肤产品有效、安全且有助于恢复敏感皮肤患者的皮肤屏障功能。     

碘海醇注射液致过敏反应和严重血小板减少及凝血功能异常

1例63岁女性行冠状动脉 CT 增强检查,静脉注射造影剂碘海醇80 ml（350 mg/ ml）。CT 检查结束留观6 min 时,患者出现手掌瘙痒、双眼结膜充血水肿、周身大汗、黑曚、恶心,呕吐少量胃内容物,随后出现唇紫、颜面水肿、四肢凉、皮肤花斑、注射部位出血。血气分析：血氧饱和度0.83,氧分压47 mmHg（1 mmHg =0.133 kPa）。距 CT 检查结束近1 h,实验室检查示血小板计数19×109/ L,凝血酶原时间（PT）18 s,凝血酶时间（TT）6 s。给予抗过敏、补充凝血因子、纠正酸中毒和电解质紊乱等治疗。距 CT 检查结束约7.5 h,患者血小板计数8×109/ L。次日,患者身体多处出现大片瘀斑,体温38.1℃,血小板计数6×109/ L。继续行对症和支持治疗,患者情况逐渐好转,第4天体温恢复正常,颜面水肿消失,瘀斑明显减轻;第6天血小板计数和凝血功能各项指标均恢复正常。     

Determination of the driving force for the sodium pump (ENa) and of active and passive conductances (GNa and Gsh) in isolated toad skin: influence of antidiuretic hormone

1. Values of the sodium potential (ENa), active conductance (GNa) and passive conductance (Gsh) were measured in the isolated skin of the toad Pleurodema thaul placed in an Ussing chamber, and Isaacson's test was performed with 2,4,6-trieminopyrimidine (TAP) and with amiloride. 2. The numerical estimates obtained in the presence of TAP were ENa 122.85 +/- 15.17 mV, GNa 0.493 +/- 0.09 mS/cm2 and Gsh 1.145 +/- 0.23 mS/cm2. 3. After exposure to ADH these values were as follows: ENa 85.76 +/- 12.17 mV, GNa 1.191 +/- 0.20 mS/cm2 and Gsh 0.935 +/- 0.14 mS/cm2. 4. Addition of 0.5 x 10(2-) TAP produced a 53.90 +/- 5.10% decrease in transepithelial potential and a 37.90 +/- 4.90% fall in short-circuit current. 5. Exposure to ADH increased the transepithelial potential difference 34.20 +/- 13.20% and the short-circuit current to 78.00 +/- 20.50% above the control values. 6. Comparison of the efficiency and mechanism of action of TAP and amiloride in the determination of electrical parameters shows that both agents induce a similar decrease in Gsh, a finding which could indicate that TAP blocks toad skin apical membrane Na+ channels without affecting tight junction conductance.     

Dermal toxicity and microscopic alterations by JP-8 jet fuel components in vivo in rabbit

In this study, we investigated the skin irritation, macroscopic and microscopic barrier alteration in vivo in rabbits from aliphatic and aromatic components of jet propellant-8 (JP-8) jet fuel. Macroscopic barrier properties were evaluated by measuring transepidermal water loss (TEWL), skin capacitance, and skin temperature; microscopic changes were observed by light microscopy. Draize visual scoring system was used to measure skin irritation. We found significant (P<0.05) increase in temperature at the site of all chemically saturated patches immediately after patch removal in comparison to the control site. Tridecane (TRI) produced a greater increase in temperature and capacitance at all time points than all the other components of JP-8. Both the aliphatic and aromatic components increased the TEWL at all time points. Tridecane produced greater increase in TEWL followed by naphthalene (NAP), 1-methylnaphthalene (1-MN), 2-metylnaphthalene (2-MN), tetradecane (TET), and dodecane (DOD). All of the above components of JP-8 caused moderate to severe erythema and edema, which were not resolved to the baseline even after 24 h of patch removal. Light microscopy revealed an increase in epidermal thickness (ET), and decrease in length and thickness of collagen fibers’ bundle by the above components of JP-8. These results suggest potential dermatotoxicity from the JP-8 components.     

In vitro skin irritation testing with human skin cell cultures

Cultured human skin cells are a potentially useful model for skin irritancy testing. We have evaluated the effects of chemical irritants on human epidermal keratinocytes (NHEK) and on keratinocyte-dermal fibroblast (NHEK/DF) co-cultures. Cell viability in NHEK cultures, measured as incorporation of the vital dye neutral red (NR), was reduced in a dose-dependent manner in response to the chemical irritants tested. The half-maximal effective concentration (NR₅₀) values correlated with irritation scores in human patch tests with these materials. Certain materials were found to be incompatible with this test system. NHEK/DF cultures were treated with ten prototype surfactants, and were evaluated for cell viability (MTT incorporation), cytotoxicity (release of the enzymes lactate dehydrogenase and N-acetyl glucosaminidase), metabolism (glucose utilization), and inflammatory mediator (prostaglandin E₂) release. There was a close correlation of the dose-response characteristics for all the endpoints tested, and between the in vitro responses and human patch test scores for the surfactants tested. These results demonstrate the usefulness of human skin cell cultures and of cell viability, cytotoxicity, and inflammatory mediator release as endpoints, for the in vitro assessment of skin irritancy.     

Assessment of an ex vivo irritation test performed on human skin explants and comparison of its results with those of a 24-/48-h human patch test for the evaluation of cosmetics

When developing new cosmetics, it is extremely important to consider the safety of consumers. Absence of potential irritancy is generally assessed using an OECD TG439 compliant Reconstructed Human Epidermis (RHE) systems and MTT assays, resulting in an irritant/not irritant classification. To gain insight into the irritancy of molecules/finished cosmetic products and to predict the outcome of irritation tests performed on subjects whatever their nature, we developed a test that uses skin explants and histological analysis. Results showed that this irritation test is sensitive enough to accurately and repeatably detect known irritants. If the diverse origin of the skin explants used led to variability in the histological alterations scored, the overall grading of irritancy is highly reproducible. Finally, when testing 120 non-alcoholic cosmetics of various galenic forms, comparison of data between the ex vivo irritation tests and of a 24-/48-h human patch test revealed a single false negative, very close to the limit, and a 10% false positive rate. It was not possible to calculate the sensitivity of the ex vivo irritation test; however, its specificity was 89.9% and its accuracy was 89.1%. Similar results, with a slightly higher false positive rate, were found when testing 49 alcoholic cosmetics. These values exceed the minimum requirements of OECD TG439.     

In vivo percutaneous absorption,skin barrier perturbation,and irritation from JP-8 jet fuel components

JP-8 jet fuel has been reported to cause systemic and dermal toxicities in animal models and humans. There is a great potential for human exposure to JP-8. In this study, we determined percutaneous absorption and dermal toxicity of three components of JP-8 (i.e., xylene, heptane, and hexadecane) in vivo in weanling pigs. In vivo percutaneous absorption results suggest a greater absorption of hexadecane (0.43%) than xylene (0.17%) or heptane (0.14%) of the applied dose after 30 min exposure. Transepidermal water loss (TEWL) provides a robust method for assessing damage to the stratum corneum. Heptane showed greater increase in TEWL than the other two chemicals. No significant (p < 0.05) increase in temperature was observed at the chemically treated site than the control site. Heptane showed greater TEWL values and erythema score than other two chemicals (xylene and hexadecane). We did not observe any skin reactions or edema from these chemicals. Erythema was completely resolved after 24 h of the patch removal in case of xylene and hexadecane.