a long-term time course of colorimetric evaluation of ultraviolet light-induced skin reactions

A long-term time course of colorimetric evaluation of ultravioletlight-induced skin reactions

S. B. Park, D. H. Suh and J. I. YounDepartment of Dermatology, Seoul National University College of Medicine, Seoul, Korea

Summary Many attempts have been made to quantify ultraviolet (UV) radiation-induced erythemaand pigmentation. However, most of these studies were concerned with the earlychanges of reactions and neglected events occurring in later stages. The long-termcourse of skin colour changes in pigmented skin, induced by broad band UVA and UVBradiation, was evaluated in 30 Korean male volunteers by means of a tri-stimuluscolorimeter for 10 weeks. The L*a*b* system recommended by the Commission Inter-national de l’Eclairage was used to measure skin colour. The L* value (luminance) givesthe relative lightness ranging from total black to total white. The a* value represents thebalance between red and green and the b* value the balance between yellow and blue.The mean individual typology angle of our subjects was 47.38, indicating ‘light’ group ofconstitutional skin colour category. One day after UV exposure, the L* and b* valuesdecreased significantly, following the colour direction of persistent pigment darkening.They then changed in opposite directions persistently until week 1, when maximumtanning was obtained. Then, a shift toward the original values was observed parallel tothe constitutive melanization axis. The a* index showed a significant increase towardthe mean colour of haemoglobin on day 1. It returned to its original value following thepathway of constitutive melanization axis. This promising quantitative method mayenable objective measurement of dermatophysiologic changes to be made, and allowevaluation of the efficacy of therapeutic modalities on skin disorders without theinherent errors associated with subjective judgement. Our results would providestandard data for long-term UV-induced skin erythema and pigmentation.

Introduction

Of the many effects induced by ultraviolet (UV)-radiationon the skin, the changes of erythema and pigmentationare one of the most prominent and important features.Because of its good reproducibility and simplicity, the useof UV-induced erythema and pigmentation is an excel-lent model of inflammation with which to characterizeskin pathophysiology and to assess the efficacy of various

agents. However, it is quite difficult to study and recordskin colour changes quantitatively as individual percep-tion of colour is complex and subjective. Recently, severalstudies have shown that the tri-stimulus colorimeter canbe very useful in the study of the quantitative evaluationof UV-induced erythema and pigmentation,1–3 the sever-ity of diseases4,5 and efficacy of treatment modalities.6

Many attempts have been made to study the time courseof erythema and pigmentation induced by UV radiation;however, the duration of most studies was short – severaldays or a few weeks at most – and they neglected theevents occurring in later stages. The exact results severalweeks after UV irradiation were not known.

The main purpose of this study was to evaluate thelong-term changes induced by UV radiation from a solar

Experimental dermatology • Original article

q 1999 Blackwell Science Ltd • Clinical and Experimental Dermatology, 24, 315–320 315

Correspondence: J.-I. Youn, Department of Dermatology, Seoul NationalUniversity College of Medicine, 28-Yongon-dong, Chongno-gu, Seoul110–744, Korea, Tel.: + 822 760 2418. Fax: + 822 742 7344.E-mail: [email protected]

Accepted for publication 29 January 1999

simulator by using colorimetric evaluation with aMinolta Chroma Meter CR-200, and to provide thebasic data for further studies.

Materials and methods

Study subjects

A total of 30 healthy Korean adult men, aged 20–35 years, with Fitzpatrick’s skin type III, IV or V wereselected. Sun-protected area of the low back was selectedfor an irradiation site. Informed consent as well as acomplete medical history was obtained. None had anyhistory of drug hypersensitivity, or abnormal reactions tosunlight. They had no drug intake for 4 weeks and nosun-exposure on their backs for 3 months before thestudy.

Radiation source and dosimetry

The skin was irradiated with simulated solar radiationfrom a compact 1000-W xenon arc lamp (Solar simu-lator, Model 81170, Oriel Co., Stratford, CN, USA) whichwas equipped with an Oriel 51605 band pass filter topass the light between 248 and 390 nm and an Oriel51220 long pass filter to block the light below 285 nm.Consequently, the skin was irradiated with a broad bandof UVA and UVB(Fig. 1).

The UV radiometer (IL 1700 radiometer, Interna-tional Light Inc., Newbury Port, MA, USA) equippedwith SED 240 UVB detecting probes which have a peak-absorbing band at 290 nm were used for measuring UVradiation dose.

UV irradiation

UV irradiation was performed in two consecutive pro-cess. First, the minimal erythema dose (MED) of indivi-dual subjects was determined by gradual increase of UVin 10 mJ/cm2 increments, starting from 20 mJ/cm2.Preliminary MED values were 60–80 mJ/cm2. Second,established UV doses were irradiated on each individual,24 h later. The test areas on the low back were exposed to2 MED, 120 –160 mJ/cm2. In order to reduce the effectscaused by desquamation and to provide a control datafor further studies, hydrophilic emollient base wasapplied twice a day on the irradiated skin during thestudy.

Colorimetric evaluation

The Minolta Chroma Meter CR-200 (Osaka, Japan) wasused to record skin colour of the designated sites beforeUV irradiation, 1 day after irradiation, and at 1, 3, 5, 6and 10 weeks. Three consecutive readings were taken ateach site and their mean values calculated.

The colorimeter, Minolta Chroma Meter CR-200, is aportable instrument with a flexible hand-held probewhich can be moved easily.6 The measured area is8 mm in diameter. This reflected-light colorimeter offersfive different colour systems for measurement. TheL*a*b* system recommended by the Commission Inter-national de l’Eclairage7 was used to measure the skincolour which is expressed in a three-dimensions: the L*value (luminance) gives the relative lightness rangingfrom total black (L* ¼ 0) to total white (L* ¼ 100); the a*value represents the balance between red (positive value)and green (negative value); and the b* value representsthe balance between yellow (positive value) and blue(negative value). The instrument was calibrated by acalibration plate (CR-A43) before each measurement.

The overall data were presented as L* vs. a*, L* vs. b*and a* vs. b*.8 According to Chardon et al.,9 the visualevaluation of the intensity of constitutional skin colour iscorrelated with the L* and b* component, and the sectorsof skin colour defined are limited by radii originatingfrom the central point (L* ¼ 50, a* ¼ b* ¼ 0) of the L*a*b*volume and having angles with the b* > 0 axis, calledcategory angles. The Individual Typology Angle (ITA),which was thought to be a quantitative and objectivevalue for classifying individual skin colour, may bedefined as follows:

ITA ¼ ½arctangentðL ¹ 50Þ=bÿ × 180=p

where, p ¼ 3.14159 and arctangent is calculated inradii.

Colorimetric evaluation of UV-induced skin reactions • S. B. Park et al.

q 1999 Blackwell Science Ltd • Clinical and Experimental Dermatology, 24, 315–320316

Figure 1 The spectrum of the light source with a band of UVAand UVB. The skin was irradiated with a solar simulator equippedwith a band pass filter to pass light at 248–390 nm and a longfilter to block light <285 nm.

Statistical methods

Statistical analysis of the data was performed by pairedStudent’s t-test. P<0.05 was considered significant.

Results

L* vs. b* plane

Absolute colorimetric values of three coordinates as atime course are given in Table 1. Plotting the values inthe plane L* vs. b* allows calculation of the ITA (Fig. 2).The mean ITA of the subjects in this study was 47.3 8,indicating that they belonged to a ‘light’ skin colourcategory according to the classification of Chardon et al.9

A day after UV exposure, the L* and b* values decreasedsignificantly, following the colour direction of persistentpigment darkening (PPD); they then went towards theopposite directions until the week 1 at which timemaximum tanning was obtained. Afterwards, theyshowed a shift toward their original position, and thisreturn happened along an axis parallel to the axis ofconstitutive melanization.

L* vs. a* plane

The a* index showed a significant increase on day 1 (Fig.3). This meant that the direction of the values at day 1represented the mean colour of haemoglobin, the coor-dinates of which are L* ¼ < 45, a* ¼< 45.9 Then, the a*index returned towards the original value in accordancewith the disappearance of erythema. During the week10, the a* value was significantly higher than the initialvalue. Thus, the value was located on the pathway ofconstitutive melanization axis (CMA), and it could bestated that erythematous reactions had completelyceased.

a* vs. b* plane

Due to the effect of PPD, the b* index was initiallydirected towards blue colour (Fig. 4). Then, the valueof b* changed towards the positive direction revealing apigmentory change of the skin towards new melaniza-tion. During the day 1, the a* index increased signifi-cantly along the direction of the mean colour ofhaemoglobin and then the final a* and b* valuesreturned to original values following the CMA.



Figure 2 The pathway of UV-induced skin reactions in plane L*vs. b*. The mean individual typology angle of subjects was 47.3 8,indicating a ‘light’ skin colour category. On day 1, the L* and b*values decreased towards the colour of persistent pigment dar-kening. After obtaining the maximum tanning after 1 week, TheL* and b* values showed a shift toward their original position,parallel to the CMA.

Figure 3 The pathway of UV-induced skin reactions in plane L*vs. a*. The a* value showed a significant increase toward thecolour of haemoglobin (L* ¼ 45, a* ¼ 45) on day 1. It thenreturned to original value following the pathway of CMA. E, Skincolour of erythema by haemoglobin.



Tab

le1

Indi

vidu

alIT

A8

and

time-

cour

ses

ofco

lorim

etric

eval

uatio

nof

skin

expo

sed

toU

Vra

diat

ion

(L*/

a*/b

*).

No

ITA

Con

trol

Day

1W

eek

1W

eek

3W

eek

5W

eek

6W

eek

10

150

.48

67.9

2/5.

12/1

4.81

57.6

2/19

.42/

14.6

960

.19/

9.05

/20.

1463

.06/

5.38

/17.

8863

.25/

6.16

/17.

8464

.08/

5.89

/17.

1764

.20/

6.28

/14.

722

45.1

864

.78/

5.56

/14.

7358

.48/

18.3

4/14

.16

55.2

4/9.

61/2

0.39

59.9

4/7.

20/1

8.54

60.2

4/6.

41/1

8.54

61.0

2/6.

19/1

7.81

61.8

0/6.

04/1

6.41

346

.48

66.1

8/5.

59/1

5.41

62.4

0/16

.37/

15.4

156

.98/

9.87

/20.

8561

.29/

7.50

/19.

6161

.13/

7.90

/19.

8361

.43/

7.84

/18.

9562

.18/

7.44

/16.

784

49.5

866

.11/

5.93

/13.

7759

.58/

18.8

6/13

.56

56.8

1/10

.43/

18.2

959

.61/

6.85

/18.

4962

.18/

6.81

/18.

3362

.18/

6.60

/17.

1860

.97/

8.11

/16.

225

53.1

866

.52/

5.53

/12.

4059

.15/

18.5

6/11

.53

61.1

2/8.

31/1

7.17

63.1

4/7.

01/1

4.62

61.9

5/7.

26/1

5.77

61.8

7/7.

72/1

4.46

63.6

0/6.

92/1

1.99

640

.88

62.7

2/6.

28/1

4.71

56.9

1/20

.21/

14.4

051

.36/

11.6

4/20

.21

54.8

3/8.

26/2

0.78

55.0

9/7.

89/2

2.45

55.5

3/8.

19/1

9.65

57.3

8/7.

66/1

6.21

744

.18

65.4

4/5.

78/1

5.95

59.1

3/18

.51/

15.6

556

.40/

10.1

2/21

.82

61.6

9/6.

52/1

8.83

60.9

0/6.

66/1

9.48

63.2

4/5.

72/1

8.32

61.2

2/7.

67/1

4.75

850

.68

67.8

3/5.

39/1

4.67

62.8

4/17

.80/

13.9

160

.69/

9.52

/18.

9461

.88/

7.74

/19.

2461

.08/

7.77

/19.

6561

.32/

7.60

/17.

1960

.97/

7.49

/14.

209

44.7

865

.30/

4.55

/15.

4559

.96/

16.1

4/16

.14

58.1

8/7.

33/1

9.04

60.2

2/6.

39/1

8.13

60.2

3/5.

81/1

9.49

60.8

2/5.

62/1

7.44

62.7

8/5.

52/1

5.75

1048

.78

67.1

3/4.

58/1

5.05

61.4

9/18

.42/

14.1

660

.32/

10.3

4/20

.03

60.5

9/6.

30/1

8.97

61.8

9/7.

24/1

9.44

61.5

1/7.

59/1

7.88

62.9

2/6.

38/1

5.79

1146

.28

66.0

7/4.

90/1

5.40

61.9

8/14

.24/

14.5

759

.95/

7.90

/19.

7762

.37/

6.24

/18.

4462

.47/

6.22

/18.

4062

.47/

6.00

/17.

6363

.44/

6.08

/14.

8512

41.9

863

.74/

6.66

/15.

3355

.35/

22.2

0/15

.49

52.5

4/12

.30/

18.5

061

.04/

7.27

/18.

6461

.30/

7.70

/17.

9260

.33/

8.12

/17.

1961

.36/

7.27

/15.

3613

33.6

861

.22/

9.06

/16.

8659

.64/

14.0

6/16

.66

56.8

7/9.

02/2

0.85

58.4

0/8.

28/1

9.77

59.2

2/7.

91/1

9.69

60.0

4/7.

54/1

8.90

58.5

5/9.

25/1

5.94

1449

.58

67.3

5/4.

73/1

4.83

60.8

9/17

.88/

13.2

356

.66/

12.6

7/19

.15

60.5

9/6.

30/1

8.97

61.5

3/7.

04/1

8.32

63.3

0/6.

74/1

6.22

63.3

8/6.

36/1

4.90

1544

.38

64.3

8/7.

46/1

4.75

59.2

3/16

.77/

13.6

858

.82/

9.94

/18.

2261

.04/

7.27

/18.

6462

.90/

7.25

/18.

6762

.92/

6.28

/17.

2761

.96/

6.47

/15.

5316

44.3

864

.90/

7.34

/15.

2556

.94/

20.8

6/12

.04

54.7

2/12

.32/

19.7

058

.21/

8.75

/17.

9158

.39/

9.32

/19.

6359

.69/

8.91

/16.

7659

.25/

9.61

/15.

8417

47.3

864

.18/

7.29

/13.

1057

.74/

25.9

0/11

.44

54.7

0/11

.22/

17.3

058

.52/

6.79

/17.

5958

.45/

7.88

/17.

9968

.92/

8.26

/16.

3258

.91/

8.94

/13.

5818

40.1

863

.66/

7.71

/16.

2260

.34/

15.6

5/15

.14

58.5

0/7.

72/2

0.60

6010

/7.5

3/17

.00

60.6

9/7.

58/1

7.16

58.9

2/6.

23/1

6.43

62.6

3/6.

25/1

4.98

1958

.48

68.0

1/5.

81/1

1.10

62.5

3/18

.14/

11.4

362

.29/

9.74

/14.

4165

.59/

6.52

/14.

1563

.77/

7.20

/14.

5865

.30/

5.86

/13.

3565

.00/

6.23

/12.

5920

53.6

866

.68/

5.61

/12.

2960

.71/

17.7

0/11

.84

57.2

8/7.

94/1

7.48

60.6

7/6.

63/1

7.32

61.3

0/6.

60/1

6.55

61.5

9/5.

87/1

6.50

61.7

9/6.

92/1

4.82

2157

.38

68.8

9/5.

11/1

2.12

60.8

4/20

.26/

11.0

460

.36/

8.57

/16.

3962

.35/

7.26

/15.

0663

.85/

6.48

/14.

3264

.05/

6.95

/14.

2564

.20/

5.63

/13.

4122

41.8

863

.73/

4.65

/15.

3355

.43/

9.30

/18.

1855

.92/

6.73

/20.

1057

.05/

7.20

/19.

0156

.43/

7.50

/19.

2168

.52/

6.70

/18.

0458

.92/

6.79

/15.

4623

45.2

864

.99/

5.56

/14.

9060

.63/

17.0

8/15

.68

53.1

6/9.

44/1

9.58

57.0

8/7.

40/1

8.45

56.8

5/7.

73/2

1.05

65.8

1/7.

59/1

8.26

57.9

2/7.

59/1

5.39

2445

.28

65.5

0/6.

48/1

5.37

58.9

5/18

.10/

13.4

555

.75/

10.7

3/13

.48

60.1

5/7.

75/1

8.11

60.5

4/7.

49/1

8.49

61.4

1/7.

30/1

5.86

61.3

8/7.

07/1

4.32

2554

.08

67.3

2/5.

68/1

2.59

62.6

0/16

.71/

11.9

658

.05/

8.88

/17.

1060

.60/

5.56

/17.

0159

.75/

5.85

/15.

3561

.38/

5.71

/14.

7561

.82/

5.64

/12.

9826

49.5

865

.79/

5.29

/13.

4959

.78/

18.2

0/11

.72

55.6

7/12

.50/

16.2

758

.27/

8.00

/18.

2558

.20/

7.95

/18.

8859

.81/

7.68

/17.

5560

.44/

6.85

/15.

5127

44.9

865

.10/

6.01

/15.

1762

.53/

18.1

4/11

.43

56.7

2/8.

33/1

9.81

59.8

0/6.

45/1

8.26

59.1

7/6.

94/1

8.02

61.0

0/6.

21/1

7.60

58.9

7/7.

90/1

5.95

2846

.08

65.4

2/5.

23/1

4.87

57.2

6/19

.03/

13.4

452

.00/

12.7

2/19

.78

57.7

0/7.

72/1

7.80

57.7

0/7.

65/1

9.89

58.6

6/7.

45/1

7.83

59.4

4/7.

36/1

6.48

2954

.58

68.6

7/4.

60/1

3.30

62.6

8/15

.97/

11.6

628

.24/

6.78

/16.

6158

.95/

5.54

/17.

2558

.99/

5.75

/17.

6060

.61/

5.76

/17.

4461

.47/

6.62

/15.

5530

46.1

865

.04/

7.37

/14.

5057

.94/

20.6

3/13

.65

58.5

1/9.

23/1

7.81

60.5

4/6.

55/1

7.78

60.6

0/7.

34/1

8.75

61.3

5/6.

39/1

6.97

61.6

2/7.

02/1

3.86

Mea

n47

.38

65.6

9/5.

89/1

4.46

59.7

2/17

.98/

13.7

157

.13/

9.70

/18.

6660

.18/

7.01

/18.

0260

.33/

7.18

/18.

3861

.00/

6.88

/17.

0461

.35/

7.05

/15.

00

Discussion

UV radiation causes a variety of biological effects on theskin, of which erythema and pigmentation are the mostprominent and expected events. In the field of dermatol-ogy or dermatopharmacology, inspection of skin colourchange, erythema and pigmentation induced by UVirradiation is widely used to determine the degree of askin reaction. Takiwaki et al.10 suggested that measure-ment of UV-induced hyperpigmentation could be used forthe assessment of topical anti-inflammatory agents,especially in pigmented skin.

In current practice it is necessary to evaluate skincolour changes by an appropriate and objective method.This can be achieved by the colorimeter, which is acolour analyser that measures reflective colours of sur-faces by the tri-stimulus system.1 The colorimeter hasbeen evaluated for obtaining objective measurements ofearly changes in erythema and pigmentation. The meterdetected a subtle, continuous transition between theprimary erythematous response and the delayed tanningof skin which was below the visual threshold for detec-tion.2 In colorimetric evaluation it is very important forthree coordinates to be plotted in separate planesbecause various kinds of skin reactions, such aserythema, immediate pigment darkening (IPD), PPDand delayed pigmentation reactions influence the

pathway of the three coordinates differently.8 The IPD,the pigmentation visible on skin immediately after lowdose UVA (1–5 J/cm2) exposure, is unstable and fadeswithin < 30 min. If the UVA dose is higher (10–25 J/cm2), then a persistent part of the IPD skin response isrelatively stable and may last for several days. From thispoint of view, the in vivo method based on IPD skinresponse and, more specifically, on its lasting componentPPD, has much to recommend it in the assessment ofUVA protection factor.11 In this study, because colori-metric evaluations were performed 1 day after UV expo-sure, the pigmentation response on day 1 followed thedirection of PPD.

L* measures the lightness of an object regardless of itschromacity. By its definition7 the L* value is influencedmainly by light absorption in the green part of thespectrum, where our perception of lightness becomesmaximal.12 Because green light is absorbed considerablyby both melanin and haemoglobin, L* is likely to beinfluenced by both pigmentation and erythema. In fact,the L* value decreased significantly in erythematous skinlesions of psoriasis.13 In addition, it reflected well thetime course of UV-induced immediate and delayed tan-ning in our study. Our data showed that the L* valuestarted to decrease at day 1 after UV radiation, whichindicated that PPD caused by the UVA component in thelight source had an effect on the L* pathway. PPD resultsfrom alteration of melanin already in the skin and is thuspercieved well in pigmented skin.14 This finding wasconfirmed by a curving of b* on day 1 to the negativedirection when a blueish shade of UV-induced melaninoxidation was detected. On day 7, a persistent decrease ofthe L* value reached its lowest point. During this period,it is not easy for the naked eye to differentiate thepigmentation accurately due to the prominent erythe-matous changes in the skin. After this the L* slowlyincreased, following the CMA, but it remained signifi-cantly lower than the control. It is thought that it takes2–3 months for UV-induced pigmentation to normalizein Caucasian skin; however, in Oriental skin the pigmen-tation reaction persisted much longer than we expected.By the combination of L* and b* indices, which measuresindividual constitutional skin colour, the Korean sub-jects’ skin is ‘light’; this is contrary to our expectation.However, although the consititutional skin colour of theKorean subjects appeared to be lighter than expected, thetanning capacity was rather high.

The a* value is known to reveal a significant linearcorrelation with the dermatologist’s perception oferythema.1 This value reached a peak 1 day after irra-diation toward the mean colour of haemoglobin, andthen gradually returned to its original value. At week 10,



Figure 4 The pathway of UV-induced skin reactions in plane a*vs. b*. On day 1, b* index was directed towards blue colour dueto the effect of persistent pigment darkening. Then, the value ofb*, going toward positive direction, revealed a pigmentationchange of the skin.

because of a higher value than that of initial point, itseemed that persistent erythema remained and inflam-matory reactions were still in progess. However, as thecoordinates were on the consititutive melanization axisthe remaining reddness was due to the red component ofthe neomelanization and inflammation was thought tobe completed. Seitz et al.2 suggested that the b* valuewould be a good indicator of tanning, showing anoverall negative pattern with respect to the L* value;this was confirmed by our data. This value initiallydecreased due to the effect of PPD. The yellow compo-nent of newly generated melanin compensates the blueshade of PPD and the b* value increased persistently toweek 5.

Takiwaki et al.10 suggested that 2 MED is the mostappropriate dose with which to assess UV-induced skinreaction in Oriental skin on the basis that tanninginduced by doses lower than 2MED was too weak todetermine the difference between the actions of variousagents, and doses higher than 2MED often resulted indesquamation which made it difficult to assess by col-orimeter 7–14 days after irradiation. In our subjectsexposed to this 2 MED, proper pigmentations wereobtained and no desquamation could be seen. TwoMED of UV is recommended for the evaluation oferythema and pigmentation. After exposure to UV radia-tion, a hydrophilic emollient base was applied to theirradiated skin twice daily. The use of hydrophilic emol-lient base is preferred because it reduces both the des-quamation of skin and the reflection of the beam fromthe colorimeter due to UV-induced dryness; in addition itcould provide standard data for use as a base for furtherstudies. In collecting data on live human skin to categor-ize skin typology, the colorimeter manufacturer recom-mends that the instrument should be calibrated with askin standard tile; however, such a standard was notavailable when we performed our study.

By the extensive use of a colorimeter, it is expected thatvarious measures by which to evaluate the physiologicalchanges in skin and the therapeutic modalities of skindisorders could be obtained and that these would be freefrom the inherent errors associated with subjective judge-ment. For this purpose, our results are expected to providestandard data for a long-term time course after UVradiation.

Acknowledgments

This study was supported by a grant from the KoreanMinistry of Health and Welfare (HMP-97-M-2-0042).

References

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2 Seitz JC, Whitmore CG. Measurements of erythema andtanning reponses in human skin using a tristimulus col-orimeter. Dermatologica 1988; 177: 70–5.

3 Westerhof W, Esterz-Uscanga O, Meens J, Kammeyer A,Durocq M, Cario I. The relationship between constitu-tional skin color and photosensitivity estimated fromUV-induced erythema and pigmentation dose–responsecurves. J Invest Dermatol 1990; 124: 264–70.

4 Serup J, Agner T. Colorimetric quantification of erythema– A comparison of two colorimeters (Lange Micro Colorand Minolta Chroma Meter CR-200) with a clinical scor-ing scheme and laser-Doppler flowmetry. Clin Exp Derma-tol 1990; 15: 267–72.

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11 Chardon A, Moyal D, Hourseau C. Persistent pigment-darkening response as a method for evaluation of ultra-violet A protection assays. In: Lowe N, Shaath NA,Pathak MA, eds. Sunscreens: Development, Evaluation,and Regulatory Aspects, 2nd edn. New York: MarcelDekker Inc, 1997: 559–82.

12 Fushs AF. The visual system: Optics, psychophysics andthe retina: In: Patton HD, Fuch AF, Hille B, Scher AM,Steiner R, eds. Textbook of Physiology, Vol I. Philadelphia:Saunders, 1989: 412–41.

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a long-term time course of colorimetric evaluation of ultraviolet light-induced skin reactions

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