1978_idson_in vivo measurement of transepidermal water loss

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J. Soc.Cosmet. hem.,29, 573-580 (September1978)

In vivo measurement f transepidermal ater lossB. IDSON Hoffmann-LaRoche, utley, NJ 07110.

Receivedeptember, 1977. Presentedt Annual Scientific eeting,Society f Cosmetic hemists, ecember976, New York, New York.

SynopsisAn overview is presented of the background and principle methods for MEASURING TRANS-EPIDERMAL WATER LOSS (TWL) IN VIVO. Absolute values of TWL are a function of the particulartechniqueand experimental onditions. WL will vary with different skin sitesand rise markedly f the skinbarrier is removed or affectedby pathologies.Early gravimetricmethods ack sensitivityand require longtestingperiods as well as arge areasof skin. The disadvantages avecausedshifts o other techniqueswhereabsorptionof water vapor is followed by a sensitivephysicalmeasurement.The majority of methodsarebasedon determining he increase n moisturecontent of either a current of dried air or fixed humidity airconducted over the skin. Others have sought to avoid air flow, using changes n conductivity of inorganiccrystals.Methods discussednclude hermal conductance, lectrohygrometry,nfrared radiation,electrolysisof absorbedwater vapor and calculation f vapor pressuregradient n the layer of air adjacent o the skin sur-face. The mechanismmay be an additiveeffect of neural control of eccrinesweatglandactivityand stratumcorneumhydration.

INTRODUCTIONWater exerts a major role in all well-beingbut particularly n skin health to maintain tsdesirablesoft, flexiblemechanical roperties 1). The lack of adequatewater in the up-per layer of the skin, the stratum corneum, results n dry and chappedskin (2-6).Dermatologic and cosmetic nterest has focusedon techniques hat generate nforma-tion on the state and quantity of water in the stratum corneum, the mobility of thewater and the influence exerted by componentsof the stratum corneum on the diffu-sion characteristics f the water (1).Water is lost throughskin n two ways,eccrinesweating nd transepidermal iffusion.Under severe hermal stress smuch as 2 1/hrmay be lost as sweat. By contrast,diffu-sional or transepidermalwater loss (TWL) is a steadypassiveprocess n which watervapor diffuses from the highly hydrated underlying tissuesthrough the avascularstratum corneum, dissolves n it and diffuses o the exterior surfacewhere it evapo-rates. Emphasis is placed on the stratum corneum since this biologically inertmembrane--due to its dense, fibrous, lipoprotein matrix--represents the principlephysicalbarrier to the penetrationof molecules hrough the integument 7). The mag-nitude of TWL hasbeen widely usedas a measureof the effectiveness f this barrier ndermatologicaldiseasestates 8-11). In pathologiessuchas psoriasis,chthyoses nd

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574 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

eczema here is impaired barrier function with increasedTWL. Normal intact skinhasan average WL of 0.31 mg cm 2 hr , risingabout en-fold n psoriasis ndeczemas.fthe barrier s removed,e.g., by Scotch ape stripping, he TWL rises o 15 to 45 mg; a50-to 150-fold ncrease ver ntactskin.No biologicalmembraneof comparablehick-nessoffers such esistance 12-14).Water loss hrough the skin is dependenton environmental actors,of which the mostimportant are the ambient temperature and humidity. Comparisonof resultsof waterlosscan only be valid if readingsare made when these wo factorsare constant.Like allmembrane-diffusion processes,TWL has a characteristicactivation energy andtherefore its magnitude is temperature dependent (9, 15, 16). Decreasing skintemperature is accompaniedby a decreasingTWL. A 5C fall in skin temperaturelowered the TWL by about 45%. The fall appeared o be related to skin temperatureand not directly to the reduction in body temperature. A rise of skin temperature of 7to 8C doubled the TWL rate (9). A number of"in vitro" and "in vivo" techniqueshavebeen described o measure he transepidermalwater diffusion from selectedareasofthe skin. Isolated skin has been studied in vitro in diffusion chambers (17-19). Thispaper, however, will only be concernedwith "in vivo" methodologyon human sub-jects. Early techniquesof measuringTWL, prior to 1965, have been criticallyreviewedby Bettley and Grice (8) and Baker and Kligman (20).Analysis of the extensive literature data indicates that the absolute values of TWLlargely depend on the technique and experimental conditionsused n its measurement(21). However it is evident that with any given technique the values of the TWLconsistently ary topographicallyrom skin site to skin site (11, 17, 22). Considerableregional variation was noted in certain areas, even after the readings had been cor-rected for varying horny layer thicknessand expressedas diffusion constants.Com-pared with that of the back, the diffusion constant s four times greater on theforehead,nine timesgreateron the backof the hand and 100 timesgreater through hepalm (20).Technical difficulties have been encountered with all methods. Basically his is becauseall measurements f TWL must, of necessity,be made under artificial conditions;varia-tions in these conditions might be expected to alter the water loss. Broadly, themethods can be classified as "ventilated" and "unventilated": ventilated-in which acontinuous flow of gas or air passes hrough a capsuleattached to the skin and thechange n the humidity of the gas is measuredby a sensingelement in the outflow;"unventilated"--in which a container s used with its open end placed on the skin sur-face, the water vaporgiven off alters he relative humiditywithin the chamberand thisrate of change s a measureof the rate of insensiblewater loss (11). Any unventilatedmethod is much less satisfactoryf the water loss s considerable incewater dropletsmay developon the skinandfail to evaporate ompletely.

TECHNIQUESUntil recently, in vivo determinationshave depended upon gravimetric estimationofthe water taken up by a hygroscopicmedium enclosed n a chamberplacedover theskin or removed rom a streamof dried air passed hrougha skin chamber.Pioneeringstudies 23) involved passingdry oxygen over a small brasschamber attached o theabdomenand collecting he water vapor n the effluent air in freezingcoils. n variant


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TRANSEPIDERMAL WATER LOSS 575

gravimetric methods (8, 24), a chamber containing a bag of calcium chloride wasfastened over the skin. The change n weight of the hygroscopicsalt indicated theamount of water transpired per unit of time. In an interesting cosmeticstudy, Powersand Fox (25) strappedsmall tared dessicators ontainingsilicagel to the arms of sub-jects and reweighed them after 2 hr. If there wasa decrease n weight, the material wasa good occludant.While crude, the method yielded the first quantitativeproof of thesuperiorityof petrolatum asan occlusiveagent.While simple o perform, gravimetricmethodsare crude and inaccurate.Resultsmayonly be obtainedat the price of considerable ffort and care(20). A main disadvantageis the lack of valuable rate data. Long periods are necessaryor testing,with lack ofsensitivity n assessing inimal day-to-daydifferences.The large areasof skin neededhas imited use n dermatologywhere the interest lies in local deviationsof smallareasof the skin. Actually, the earlier methodsof weighing absorbedwater vapor have beengraduallyabandonedwhen limited areasare involved in investigation. n addition, thetestscan be compromisedby eccrinesweating,which cannotbe discounted,particu-larly when long periods of testingare used. Since eccrinesweating s so much greaterthan transepidermaldiffusional oss, most subsequent nvestigatorshave sought o in-hibit the former by use of anticholinergicdrugs (9, 20) and keeping the ambienttemperature low. However, excessiveemotional sweatingusually appearsas transientrapid ncreasesn water losswhich are easilydistinguishedrom baselineTWL (21).The disadvantagesf the gravimetricmethod have causedshifts o other techniqueswhere absorption of water vapor is followed by another more sensitive physicalmeasurement, such as the electrical conductance of a chemical sensor cell or elec-trolysisof the absorbedwater.The majority of current methods are basedon the increase n moisture content of acurrent of dried air conductedover the skin. Some investigators onsider t a disad-vantage hat the skin is exposed o dry air insteadof the normal environmentalhumidair (26). Since he permeabilityof the skin dependson the water contentof the stratumcorneum, the water content of the horny layer of the skin alters when the watercontentof the atmospherechanges. hus many investigators refer to study the watervapor lossof the skin when exposed o air of a fixed humidity, which can be obtainedby bubbling he air through a saturatedsodiumchloride solutionbefore it reaches heskin (27). Other investigators venwant to avoid a flow of air along he skin surfaceandrecord the increasinghumidity inside a cup placedupon the skin (28). Ideally, the skinshould be investigatedunder unaltered atmosphereconditionsso that the skin doesnot need time to acclimatize o a changedenvironment. Some methods approach hisideal, e.g., where environmentalhumid air is conductedover the skin and hygrometersare mounted in the air both before and after it has passed he skin (20, 21, 29). As willbe discussed,only large areas of skin have been used and the sensitivity of the hy-grometer is critical.Investigatorssuch as Thiele and Schutter (28) have sought to avoid air flow entirely.They consider hat the streamsof a carrier gas,used o transfermoisture rom the skinto the measuringvessel, create abnormal water vapor gradients.They developed asensitivemethod basedon the changeof conductivityat the surfaceof a temperature-controlled halite (rock salt)crystal, esulting rom the adsorptionof minute amountsofwater evaporating under normal conditions from the skin surface. Without thistemperaturecontrol, the temperatureof the saltcrystalwould be adverselyaffectedby


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576 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

heat radiation rom the skin surface.The temperatureof the crystal s kept constant ymeans of a cooling systemconnectedwith a circulation themostat.The sensingele-ment is a thermal conductivity cell comprising two compartments. The air passesthrough the first compartment before it reaches the skin. After the air has beenhumidifiedby passing ver the skin, he air is led through he second ompartment fthe cell. A thermistor is mounted in each of the two compartmentswhich are then in-corporatednto the armsof a Wheatstone ridgecircuit.Any differencen the com-positionof the air between he two compartmentsauses n imbalancen the bridge,which is recorded directly (29).The measurementof the thermal conductivityof the air allows he measurement f theinsensible erspirationof 1 cm2 of forearmskin (30). It is possible o quantitatively e-tect 0.1 to 30/ag/cm2/min vaporatingrom the skin. Simultaneously,he watervaporpressureat the skin surfacecan be recorded. From this vapor pressurecan be calcu-lated the relative humidity of the skin surface which is a measure of the moisturecontent of the outermostskin layers.Quattrone and Laden (31) have adapted he thermal conductivitymethod to use a car-rier gas, n a method which they call transpirometry. The investigators mploy an ap-paratuswherein a stream of dry nitrogen, passing n a flow-throughchamberon theskin, and a streamof identicalpressure lowing ndependentlyof the skinare comparedfor their thermal conductivity in a gas chromatograph.Two of these systems,eachequipped with integrators,allow for simultaneousmeasurementof the rate of moisturelossat two separatesites i.e., a control and a test). In the actualmethod, for eachunit,streamsof dry nitrogen are split into two equalcomponents--onepassing irectly ntothe chromatographyunit, while the other streams nto a flow-through probe on theskin before entering this thermal conductivity analyzer. The difference in theconductance between the split streams is measured and a signal from eachchromatograph s sent to a dual pen recorder. The latter is equipped with two repeat-ing potentiometers, allowing for integration of each signal. Standardcurvesare ob-tained for eachsystembefore use eachdayby applicationof known quantities f waterto filter paper sealedwithin eachchamber.The previous static-conductancemethod can be replaced with a dynamic electrohy-grometer technique 20, 21, 29) wherein ambient humidity air is swept hrougha skinchamber over a plate coated with a sensorchemical whose electrical conductivity s afunction of the ambient humidity. Sulzbergerand Herrmann (32) were the first to at-tempt to monitor humidity changes y meansof electrohygrometry.They passed irthrough a skin chamberand over a plate coatedwith lithium sulfate.A group of inor-ganic sensor salts, of which lithium bromide is the most commonly used, has sub-sequentlybeen refined and made commerciallyavailable. t should be noted that the"humidity sensor"devicesare limited by the fact that they operatewithin an enclosedarea. Therefore the measurements ave to be accomplished ery quickly in order toavoid saturation of the air contained within the chamber. Electrohygrometry haslowered the skin conditioning ime, asopposed o gravimetricmethods.In electrohygrometricTWL measurements, current of air is either predried by pass-ing through a freezing mixture (8) or is passedacrossa pre-sensor o record the hu-midity of the inflowing air (11) and is then conducted nto a samplingchamber.Thechambershave extended from 15 cm2 area of skin (21) to 60 cm2 (8). The apparatususually ncorporatesa humidity transducerwhich provides continuousmonitoringanda thermistor for measuringskin temperature 8, 9). As previously discussed,he


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presence of sweat is either inhibited by inactivation of sweat glands by use of an-ticholinergic drugs (20) or ascertainedby the galvanicconductivity of skin (9). If thesweating is not inhibited, attempts are made ,, hold bin t,=,,,,ot,, bl(,, 34C_which is the lower thresholdof sweating.Low air-flow rates are appropriate or diffu-sion measurements;n intact skin between 50 and 300 ml/min is the usualrange, butthe flow must be adjusted to the rate of water loss.Too small a flow will allow the hu-midity to build up in the system,or, in sweatstudies,may fail to vaporize the dropletsas they form. Too large a flow may result in uneven mixing since he relative humidity(RH) and temperature (T) of the air is monitored prior to and following its passageover the skin surface, he difference n relative humidity (& RH) represents he watervapor picked up at the skin surface 21). Eachmeasurement equiresapproximately15to 20 min. Most L measurementsare performed with a Sageelectric hygrometer,Model 154 (Sage nstruments,White Plains, New York) using ithium bromide sens-ing elements.The L may be calculated ccording o the formula (21):

&RH 1TWL - x D AF -- (1)100 Awhere:

TWL is the transepidermalwater loss n mg cm-'- hr--ARH is the difference between the incoming and effluent relative humiditiesD is the weight of water per liter of saturatedsteam at the temperature of the airpassing ver the skin n mg 1 AF is the volumetric air flow rate in 1 hr -A is the area of skin in cm 2

the densityof saturated team D) at different temperatures s obtained rom tables.In a variant hygrometer method, TWL was determined by Berube et al. (33), usingcompressed ir as a carrier gas. The flow of the gas was split into two streams o flowover the left and right arm of subjects.Each stream then flowed over a Sage hy-grometer where the moisture is swept from the surfaceof the skin into the gasstream.The stream passes hrough the sensing hamber Dew Point hygrometer (CambridgeSystems,Newton, Massachusetts) here the amount of moisture present is measuredutilizing the dew point principle.The concentrationof water vapor in a gas stream can also be measuredby its absorp-tion of i.r. radiation (15). TWL can be measuredon areas rom 20 cm2 (34) to as ow as1 to 4 cm (35). The principle s that infrared radiationpasseshrough wo conductiontubes and then into the i.r. detector. In practice,dry gas s passedover the skin surfaceand the moistened gas is then passed hrough the analyzer. When the gas streamcontaining water vapor is passedthrough the conduction tube, while the other isflushed with dry gas, some radiation is removed by the wet gas stream. This producesan imbalance n the radiationabsorptionbetween the two sidesof the detectorwhich isa measure of the TWL. The measurement is made when water loss becomes constant ata particular flow of dry nitrogen gas over the skin. This is called the equilibrium statebecause oss of water from the skin surface s exactly matched by water diffusing upfrom the epidermis. Measured in this way, it was found that the rate of TWL wasmodified by the rate of flow of dry gas; ncreasesn the flow of dry gasproducescor-


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the temperature lone.Accordingo eq 2, the waterevaporation er unit time andarea(ER) can then be calculatedas a constantmultiplied by the measureddifference invaporpressurebetweenthe two measurement oints.

MECHANISMThe physiologicalmechanismand relative contributionsof chemical composition,membranestructrueand topographical kin area in TWL is relatively ill-defined.Wildnauer and Kennedy (21) suggest hat the phenomenaof TWL is not simply aphysicalprocess ollowingphysicochemicalaws,but that somephysiological rocessescanparticipaten the mechanism nd herefore nfluence he magnitude f TWL. Theycite evidence 39-41) which stronglysuggestshat this physiologicalnvolvementmaybe the neuralcontrol of the eccrinesweatgland.The resultantof this nteractionof ec-crine glandactivitywith stratumcorneumproperties hat influenceTWL can be in-terpreted asa hydrationeffect on membranepermeability.

REFERENCES(1) B. Idson,Water and the skin,J. Soc.Cosmet. hem.,24, 197 (1973).(2) I.H. Blank, Further observations n factorswhich nfluence he water content of the stratumcorneum,

J. Invest.Dermato/., 21,259 (1953).(3) I. H. Blank, Factorswhich nfluence he watercontentof the stratumcorneum, bid., 18, 433 (1952).(4) K. Laden,Natural moisturizing actors n skin,Amer.Perfum.Cosmet., 2, 77 (1967).(5) L. E. Gaul,Relationof dewpointandbarometric ressureo horny ayerhydration, roc. ci.Sect. oiletGoods ss.,40, 1 (1963).(6) L. E. Gaul and G. B. Underwood,Relationof dew point and barometricpressure o chappingof normal

skin, J. Invest.DermatoL,17, 9 (1951).(7) A.M. Kligman,The Biologyof the StratumCorneum, n "The Epidermis,"MontagnaandLobitz,Eds.,Academic Press, nc., New York, 1964.(8) F. R. BettleyandK. A. Grice, A method or measuringransepidermal ater oss,Brit,J. Dermatol., 7,627 (1965).(9) K. A. Grice and F. R. Bettley, The effect of skin temperatureand vascular hangeon the rate oftransepidermal ater oss, bid., 79, 582 ( 1967).(10) P. Frost, G. Weinstein,J. W. Bothwell and R. Wildnauer, Ichthyosiformdermatoses, studiesoftransepidermal ater oss,Arch.DermatoL, 8, 230 (1968).(11) M. Shahidullah,E. Raffle and W. Frain-Bell, Insensiblewater loss n dermatitis,Brit..J. Dermatol.,79,589 (1967).(12) R.J. Scheuplein,Mechanism f percutaneousbsorption, . Invest.DermatoL, 5, 334 ( 1965 .(13) M. Katz and B. J. Paulsen,Absorption of Drugs Through the Skin, in B. B. Brodie and J. Gilette,"Handbook of ExperimentalPharmacology," ol. 28, Springer-Verlag,Berlin, 1971.(14) B. Idson, Biophysicalactors n skinpenetration, , Soc.Cosmet. hem.,22,615 (1971).(15) D. Spruit and H. E. Herweyer, The ability of the skin to change its insensibleperspiration,Dermatologica,34, 364 (1967).(16) M. Huheey and T. Adams,Localeffect of temperatureon skin evaporativewater oss,J. AppL PhysioL,22,939 (1967).(17) I. H. Blank, Factorswhich nfluence he water contentof the stratumcorneum,J. Invest.DermatoL,18,433(1952).(18) J. W. H. Mali, The transport f water hrough he humanepidermis,bid., 27, 451 (1956).(19) H.D. Onken andC. A. Moyer, The waterbarrier n humanepidermis,Arch.DermatoL, 7,584 (1963).(20) H. Baker and A. Kligman, Measurementof transepidermalwater lossby electricalhygrometry,Arch.DermatoL,96, 441 (1967).(21) R. H. Wildnauer and R. Kennedy,Transepidermalwater loss n humannewborns, . Invest.Dermatol.,54, 483 (1970).


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580 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS(22) K. Ohara, Fluctuationsn insensibleperspiration,Jap.J. PhysioL,13,441 (1963).(23) G. F. Burch and T. Winsor, Rate of insensibleperspiration diffusion of water) locally through ivingand through deadhumanskin,Arch. ntern. Med., 74,437 (1944).(24) S. Monash and H. Blank, Location and reformation of the epithelial barrier to water vapor,Arch.Dermatol.,78, 710 (1958).(25) D. Powersand C. Fox, A study of the effect of cosmetic ngredients;creamsand lotions on the rate ofmoisture oss rom the skin, Proc.Sci.Sect. oilet Goods ss.,28, 21 (1957).(26) D. Spruit,The measurement nd the regeneration f the water vapor ossof humanskin,Dissertation,Nijmegen (1969).(27) R. E. McDowell, D. H. K. Lee and M. H. Fohrman, The measurementof water evaporation romlimited areasof a normalbody surface, . Animal Sci., 13,405 ( 1954).(28) F. A. J. Thiele and K. Schutter, A new micro method for measuring he water balanceof the humanskin.Saltcrystalmethod , II,J. Invest.Dermatol., 9, 95 (1962).(29) F. A. J. Thiele and K. Schutter, Moisture meters for measuring he water balanceof the human skin,Proc.Sci.Sect. oilet Goods ss.,40, 20 (1963).(30) D. Spruit, Measurement of the water vapor loss from human skin by a thermal conductivitycell, J.Appl. Physiol., 3,994 (1967).(31) A.J. Quattrone and K. Laden, Physical echniques or assessing kin moisturization,J. Soc.Cosmet.Chem.,27, 607 (1976).(32) M. B. Sulzberger and F. Herrmann, "The Clinical Significanceof Disturbances n the Delivery ofSweat,"CharlesC. Thomas, Publishers,Springfield, llinois, 1954.(33) G. R. Berube, M. Messingerand M. Burdick, Measurement n vivo oftransepidermalmoisture oss,J.Soc.Cosmet. hem.,22,361 (1971).(34) R. E. Albert and E. D. Palmes,Evaporative ate patternsof small skin areasasmeasuredby an nfraredgasanalyzer,J. Appl. Physiol.,4,208 (1951).(35) C. Johnsonand S. Shuster,The measurementoftransepidermalwater loss,Brit. J. Dermatol.,81, Supp.4, 40 (1969).(36) D. Spruit,Measurement f water vapor oss rom very smallareasof frozenskin,J. Invest.Dermatol.,

58, 109 (1972).(37) D. Spruit,Epidermalwater-barrier ormationafter strippingof normalskin, bid., 45, 6 (1965).(38) K. Hammarlund,. E. Nilsson, . A. Oberg ndG. Sedin, ransepidermalater ossn newbornn-fants. I. relation to ambient humidity and site of measurementand estimationof total transepidermalwater loss,Acta. Paediat.Scand.,66, 553 (1977).(39) P. Thomas and A. Kawahata,Neural factorsunderlyingvariations n electricalresistance f apparentlynonsweating kin,J. Appl. Physiol.,17,999 (1962).(40) T. Adams and W. Hunter, Modificationof skin mechanical ropertiesby eccrinesweatglandactivity,Ibid., 26, 417 (1969).(41) P. Lloyd, Secretionand Reabsorption n EccrineSweatGlands, n "Advancesn Biologyof Skin," W.Montagna,R. Ellis and A. F. Silver,Eds.,PermagonPress,New York, 1962, pp 127-150.

1978_idson_in vivo measurement of transepidermal water loss

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