UV PROTECTION AND EVALUATION OF EFFICACY OF SUNSCREENS 333 The IPF of a sunscreen can be determined using the induction or the elicitation arm of the local CHS or DTH response, and the systemic DTH response. Therefore, in order to understand fully the relationship between SPF and IPF, it is essential that UVR dose– response studies should be carried out with and without sunscreen. QUANTIFICATION OF CHS RESPONSES Sensitization of skin is carried out using 2,4-dinitrochlorobenzene (DNCB) 24 h after irradiation. Sunscreen control groups can be treated with sunscreen and sensitized with ethanol only (in the center of the sunscreen-treated site) to determine the nonspecifi c ir- ritant effects of DNCB challenge (72). After a specifi c period, elicitation sites were quan- tifi ed as mentioned below. The dermal thickness of each elicitation site was determined using a high-frequency 20 MHz ultrasound scanner (78). The percent increase in dermal thickness for each elicitation site is plotted versus DNCB challenge dose (x-axis), and the dose–response relationship can be determined using lin- ear regression analysis. The CHS response is represented by the slope of the linear regres- sion line. The steeper the slope the stronger is the response. Theoretically, IPF = SPF (i.e., IPF/SPF = 1) especially if erythema and immunosuppres- sion have common chromophore(s) and both endpoints have similar dose–response curves. However, in vivo SPF for sunscreens are not predictive of the sunscreen’s IPF, determined using nickel CHS model. Immune protection seems to be independent of erythemal pro- tection. Investigations carried out by Poon et al. (2003) revealed that the range of SPF of selected sunscreens was found to be between 6 and 20, whereas the range of IPF was be- tween 2 and 21. The sunscreen with the highest SPF did not have the highest IPF, whereas the sunscreen with the lowest SPF did not have the lowest IPF (78). Thus, SPF may not predict the ability of sunscreens to protect the immune system. The paradigm is based on analysis of investigational data sunscreen containing 2% octyl- methoxy cinnamate is expected to have an in vivo SPF around 2 (5.7 found in vitro), sun- screen having 2% o-PABA—an SPF of 2.5 (4.5 found in vitro), and sunscreen with 6% ZnO—SPF of 5 (3.8 found in vitro) (79). So, the conclusion that the formulation that provides the highest immune protection is the formulation with highest in vitro SPF is valid only for an in vitro situation and particular model of evaluation. Table III summa- rizes different in vivo techniques used to calculate IPF of sunscreen products. UVA-PROTECTION FACTOR DETERMINATION SPF is fi rst and foremost a measure of UVB protection as UVB is 1000 times more ery- themogenic than UVA. Presently, there is no agreement about the paramount method for measuring UVA protection. A variety of methods have been proposed. In vivo methods have been developed among which persistent pigment darkening (PPD) is more broadly used. PPD is measured 2 h after irradiation of the skin with 30 joules/cm2 of UVA. Application method and UV irradiation protocol used for UVA-protection factor (PF) determinations are similar to that used for SPF testing. The only exception is that UVA spectrum should be used for UVA-PF determination. The results may be observed at a

Table III Different In Vivo Techniques Used for IPF Calculation from CHS Response via Suppression of Induction and Elicitation Studies Antigen used Quantifi cation of responses via suppression of induction IPF calculation via suppression of induction studies Reference DNCB Ultrasound measurement & reaction diameter calculation. Determined average CHS response of all elicitation sites. Linear relationship between CHS response and DNCB concentrations From CHS responses IPF = SSR dose that induced 50% immunosuppression (ID50) in unprotected group/SSR dose that induced ID 50 in protected groups. Or IPF = SSR dose responses modeled curves for CHS responses with sunscreen/SSR dose–responses-modeled curves for CHS responses with sunscreen without sunscreen (80) DNCB Determined percentage increase in dermal thickness instantly by 20 MHz ultrasound image analysis and scans were performed immediately before and 48 and 72 h after the challenge. Slope of the DNCB dose–response is a measure of CHS Percentage increase in dermal thickness = Difference in dermal thickness at 72 h and 0 h × 100 Dermal thickness at 0 h (72) DNCB Determined increase in skin-fold thickness (SFT) at each elicitation site, and an overall score per volunteer was given by the sum of the SFT of all elicitation sites. Each challenge site was measured before and after patch application Individual CHS responses were expressed as a total millimeter increase in SFT and total North American Contact Dermatitis Group (NACDG) score were plotted against total UV dose delivered in joules/cm2. Nonlinear regressions were generated from the different UV dose–response curves (81) JOURNAL OF COSMETIC SCIENCE 334