2. Isolated Human and Animal Stratum Corneum as a Partial Model for the 15 Steps of Percutaneous Absorption: Emphasizing Decontamination Part II

Xiaoying Hui¹ and Howard I. Maibach¹

(1)

Department of Dermatology, University of California San Francisco, San Francisco, CA, USA

Xiaoying Hui

Email: Xiaoying.Hui@UCSF.edu

Keywords

Stratum corneumChemical bindingRubbing

The stratum corneum (SC) of humans and animals holds key insights to the development of an efficient protective barrier against contamination and to devising effective decontamination interventions. Advancing knowledge of this ever-changing defense layer with respect to the percutaneous penetration of chemicals provides many opportunities to optimize chemical-specific decontamination protocols. The SC acts as a “reservoir” for topically applied molecules, and even rapid washing with water post dermal exposure frequently fails to remove most chemicals [11]. Importantly, Rougier et al. [28, 29] discovered that SC sampling in vivo in humans and rats via tape stripping 30 minutes post exposure accurately predicts and quantifies chemical penetration for up to 4 days with linear-type correlation. While percutaneous penetration is often considered a simplistic one-step diffusion process, it consists of at least 15 steps [37] related to absorption and excretion kinetics, extraction, vehicle characteristics, wash effects, etc. (Table 2.1) Additionally, the chemical partitioning/diffusion behavior of a wide array of compounds with differing physicochemical properties has approximately the same diffusion coefficients with regard to their percutaneous absorption in vivo. Thus, for a given thickness of SC and a specific anatomical site, the penetration flux value of a substance depends mainly on its SC/vehicle partition coefficient [30, 31].

Table 2.1

15 steps related to the process of percutaneous absorption^a

Number	Factors determining percutaneous absorption
1	Release from vehicle^b Varies with solubility in vehicle, concentration, pH, etc.
2	Kinetics of skin penetration^b Influenced by anatomical site, degree of occlusion, intrinsic skin condition, animal age, concentration of dosing solution, surface area dosed, frequency of dosing, post absorption, etc.
3	Excretion kinetics^b
4	Tissue disposition
5	Substantivity to skin^b
6	Wash effects^b Wash resistance Wash enhancement
7	Rub effects^b Rub resistance Rub enhancement
8	Transfer––skin, clothing + inanimate surface^b
9	Exfoliation^b
10	Volatility^b
11	Binding––all layers^b
12	Anatomic pathways
13	Lateral spread^b
14	Vascular perfusion^b
15	Cutaneous metabolism^b

^aModified with permission from Wester and Maibach [37]

^bMetric developed

The abovementioned observations provide novel insights for improving decontamination techniques. Despite many emergency treatment protocols, numerous topically applied chemicals are not easily removed by water washing. Skin decontamination is the primary required intervention for chemical, biological, and radiological exposures and involves immediate removal of the contaminant via the most effective manner. Contaminant removal techniques can consist of physical removal including the use of friction, use of a liquid vehicle to solvate or emulsify the contaminant, contaminant transfer to another medium through absorption/adsorption, and chemical alteration of the contaminant. Decontaminants should effectively and rapidly remove contaminants without causing enhancement, wash-in effect, or skin damage while easily removing itself without undesirable residue. It should additionally be readily available, affordable, and easily disposed of [8]. In the second part of this review, we continue to explore the many steps involved in percutaneous penetration, the laboratory techniques providing the background for this knowledge, and the insights advancing our knowledge of chemical exposure risk assessment, exposure prevention and barrier methods, and postexposure decontamination. We address knowledge gaps to guide future research to guide our ability to minimize the harmful effects of hazardous dermal chemical exposure.

Tissue Disposition and Binding: All Layers

The skin is an organized, heterogeneous, and multilayered organ. The SC, epidermis, dermis, appendages, and vasculature constitute the outer living protective system. Percutaneous absorption of topically applied agents is the sum of the penetration and permeation of a chemical through the SC, epidermis, and part of the dermis [10, 22, 32]. The effect of the skin layers on percutaneous absorption was evaluated by quantifying the partitioning of compounds in water and isopropyl myristate (IPM) between skin layers. The influence of vehicle drug concentration, equilibration time, hydrophilicity, lipophilicity, and pH on the partitioning behavior of compounds was examined [13].

Chemical Binding and Dispersion to Different Skin Layers

Chemical partitioning occurs in human SC, epidermis, and dermis, and partitioning in the SC as a function of equilibration time is shown in Fig. 2.1. The lipophilic chemicals propranolol, atrazine, and salicylic acid reached equilibrium in 3–6 hours in the SC and epidermis. Theophylline, the most hydrophilic compound, required 12 hours to achieve equilibrium; however, in the dermis, equilibrium was reached in 3 hours secondary to the ease of dermal hydration. MOP exhibited a different PC time profile with the skin layers not reaching saturation until 24 hours post exposure, although the PC continued to increase with contact time. Lipophilic compounds including propranolol, salicylic acid, and atrazine exhibit higher PCs in the SC than in the epidermis or dermis; however, the PC values of theophylline remained relatively constant in the three skin layers.

../images/416314_1_En_2_Chapter/416314_1_En_2_Fig1_HTML.png — Fig. 2.1

The graph is modified with permission from Gogoleva et al. [13]. Stratum corneum (sheet)/water partition coefficient (mean ± SD, n = 3) of model compounds as a function of equilibrium time. In the stratum corneum, the lipophilic chemicals propranolol, atrazine, and salicylic acid reached equilibrium in 3–6 hours. The hydrophilic compound theophylline required 12 hours to achieve equilibrium. Solutions of chemicals used were at saturation solubility concentrations. The lipophilic chemicals, propranolol, atrazine, and salicylic acid, reached equilibrium in 3–6 hours. The most hydrophilic compound, theophylline, required 12 hours to achieve equilibrium

The data presented here corroborates that in previous literature reports. The calculated partition coefficients for the compounds as a function of equilibration time, chemical concentration, and solvent nature may be valuable in predicting the in vivo and in vitro transport of drugs and environmental agents through human skin.

Raykar et al. [26] suggested that a lipid and protein domain exist in the SC. The uptake of highly lipophilic compounds (log P values near 3.0) may be governed by the lipid domain of SC, and hydrophilic solutes are taken up in the protein domain. Because theophylline partitions equally into the skin layers, the retention could be secondary to its binding to the protein domain. The data presented here verify earlier observations [5, 7, 17].

Wash Effects and Skin Decontamination

The Powdered Stratum Corneum Model to Rapidly Evaluate Wash Resistance and Decontamination Efficiency

Most material safety data sheets (MSDS) recommend soap and water to remove chemicals from the skin surface. Because of the varying physical-chemical properties, each chemical has different SC binding sites with diverse affinities. Therefore, understanding a chemical’s wash resistance can help to identify optimal decontaminants.

The powdered SC model provides a tool to rapidly evaluate the chemical’s potential skin binding and partitioning behavior and to determine washing solution efficacy. A radiolabeled contaminant chemical was mixed with powdered SC and incubated in a 37 °C water bath for 0.5–2 hours. The chemical and SC mixture was washed multiple times with the selected solution and centrifuged to separate the washing solution from the SC pellet, and then the radioactivity of each sample was measured. Using this protocol, the decontaminant effects of multiple decontaminants could be compared [16].

Decontamination Efficiency Related to Chemical Dose-Exposure Time

Table 2.2 shows that [¹⁴C]-2,4-D was easily washed from the bound SC particles with water alone, and five washings five times removed more than 90% of the bound compound. Most bound compounds (50% or more) are removed after the first washing [27, 43, 44]. Alachlor readily binds powdered SC without dependence on contact duration; thus, water washing alone removes only about 2% of the compound regardless of dose, concentration, or contact duration. Using 10% soap washing solution, about 77% of the alachlor was removed and 50% washing solution removed 90% of bound alachlor from the powdered SC [6, 42]. This elegantly shows the effectiveness of soapy water for washing hands and how powdered SC can be used to determine the effectiveness of skin decontaminants [33].

Table 2.2

Water decontaminates environmental hazardous chemicals (2,4-d) from human skin^a

Dose (μCi)	Radioactivity recovery as percent dose of [¹⁴C]-2,4-D (%) applied
	Supernatant	Powdered stratum corneum
	Supernatant	Before washing	After first washing	After second washing	After third washing	After fourth washing	After last washing
0.22	87.4 ± 0.5	12.6 ± 0.5	4.8 ± 0.8	4.5 ± 0.8	2.2 ± 0.2	0.9 ± 0.1	0.9 ± 0.1
0.44	84.6 ± 1.9	15.4 ± 1.9	6.8 ± 1.6	4.4 ± 0.5	2.6 ± 0.4	1.1 ± 0.2	1.1 ± 0.1
0.88	82.9 ± 1.9	17.1 ± 1.9	8.3 ± 2.0	4.9 ± 0.7	2.3 ± 0.5	1.1 ± 0.2	1.1 ± 0.1
1.76	86.1 ± 1.2	13.9 ± 1.2	5.5 ± 0.7	4.2 ± 0.7	2.5 ± 0.6	1.2 ± 0.2	1.1 ± 0.2
2.20	78.9 ± 1.8	20.1 ± 1.8	10.9 ± 2.9	4.6 ± 1.2	2.4 ± 0.3	1.4 ± 0.3	1.2 ± 0.3
3.52	81.7 ± 4.1	18.3 ± 4.1	8.7 ± 4.0	5.0 ± 0.6	2.0 ± 0.2	1.6 ± 0.5	1.5 ± 0.5

Water washing five times removed more than 90% of bound [¹⁴C]-2,4-D from the powdered SC. Most bound compounds (50% or more) were washed off after the first washing

^aModified with permission from Wester RC et al. [43] and [44]. Results indicate that most radioactive residues were washed off within first two washes. After a 2-hour incubation/equilibration period, the test sample was centrifuged and then separated to the supernatant and pellet parts. The pellet sample was further washed by water for five times to remove unbound test chemicals. Each time after washing, the sample was centrifuged again to separate the supernatant and pellet parts

Comparison of Decontamination Efficiency Post Short-Time Exposure

Potential environmental exposures may happen during manufacturing, distribution, application, deliberate exposure, or by residual presence. The concentrations of chemical agents and the contact duration are primary determinants of the degree of skin destruction and systemic poisoning. To prevent and diminish chemical absorption, immediate washing with appropriate decontaminating agents is essential to reduce damage and percutaneous penetration.

Glyphosate is a broad-spectrum, postemergence, translocated herbicide. Human exposure can occur through production or general use. The extent of glyphosate binding, systemic absorption, and removal post exposure was studied.

A powdered SC model examined the skin binding affinity. Table 2.3 shows glyphosate is easily removed from SC with water, regardless of time exposure or dose concentration. After three water washes, less than 0.1% of the chemical remained bound to the powdered SC [41].

Table 2.3

Glyphosate partition from roundup vehicle to powdered stratum corneum^a

	Radioactivity recovery as percent dose of [¹⁴C]-glyphosate applied
	Undiluted	Diluted 1:20	Diluted 1:32
30 minutes exposure
Vehicle	96 ± 0.9	102 ± 1.2	96 ± 1.6
Water washing	2.4 ± 0.2	1.2 ± 0.07	1.1 ± 0.2
Stratum corneum	0.04 ± 0.04	0.03 ± 0.03	0.02 ± 0.01
4 hour exposure
Vehicle	96 ± 2.4	104 ± 1.3	91 ± 2.9
Water washing	3.3 ± 0.8	1.2 ± 0.2	5.0 ± 2.2
Stratum corneum	0.01 ± 0.01	0.02 ± 0.03	0.05 ± 0.04
8 hour exposure
Vehicle	96 ± 0.4	98 ± 6.6	95 ± 1.4
Water washing	3.0 ± 0.2	2.2 ± 0.5	2.3 ± 1.2
Stratum corneum	0.02 ± 0.0	0.01 ± 0.01	0.02 ± 0.01

Results show that glyphosate was weakly bound with powdered stratum corneum. In all cases, the amount remaining with the powdered stratum corneum pellet after the water washing was less than or equal to 0.05%

^aModified with permission from Wester et al. [41]. Undiluted, 1.1 mg glyphosate/ml/mg SC; 1:20, 0.059 mg; 1:32, 0.037 mg. Data expressed as mean ± S.D

Washing with water or water with soap is traditionally used to reduce potential damage and percutaneous penetration of chemicals topically applied, but may not be effective for decontamination of lipophilic compounds. For instance, the removal of alachlor with water is less effective than soap with water. In contrast, both soap-and-water wash and emergency water shower are relatively ineffective at removing the potent contact sensitizer methylene bisphenyl isocyanate from the skin [45]. Residual chemical on the skin post standard washing procedures can have toxic consequences [19]. Additionally, the “wash-in” effect can potentially enhance percutaneous penetration and systemic absorption of chemicals [21].

Skin decontamination is often emergent. A human skin in vitro model was used to compare the decontamination capacity of three decontaminant solutions: tap water, isotonic saline, and hypertonic saline mixed with radiolabeled [¹⁴C]-glyphosate (Table 2.4). After a defined exposure time, the skin surface was washed three times with 4 mL of each solution. Tape stripping was then employed to collect an SC sample, and all collected samples were analyzed by a liquid scintillation counter to determine radioactivity. This in vitro model is economical, rapid, and provides quantitative data to aid screening for optimal decontaminants.

Table 2.4

Human skin decontamination of glyphosate in vitro^a

	Radioactivity recovery as percent dose of [¹⁴C]-glyphosate applied
	1 minutes exposure	3 minutes exposure	30 minutes exposure
Water washing
Wash solution	93.52 ± 16.42	94.60 ± 26.39	85.09 ± 14.06
Stratum corneum	0.10 ± 0.10	0.14 ± 0.10	1.78 ± 0.84
Skin residue	2.21 ± 1.99	0.70 ± 0.44	7.50 ± 3.82
Systemic absorption	0.04 ± 0.05	0.05 ± 0.10	2.26 ± 2.19
Hypertonic saline
Wash solution	101.23 ± 14.20	98.69 ± 21.16	85.04 ± 14.62
Stratum corneum	0.12 ± 0.07	0.14 ± 0.09	1.54 ± 1.06
Skin residue	1.02 ± 1.27	1.45 ± 1.69	5.06 ± 3.29
Systemic absorption	0.04 ± 0.04	0.54 ± 0.84	3.17 ± 6.66
Isotonic saline
Wash solution	98.72 ± 18.34	95.56 ± 28.08	78.95 ± 9.38
Stratum corneum	0.08 ± 0.05	0.14 ± 0.11	2.23 ± 2.10
Skin residue	0.74 ± 0.90	3.53 ± 5.15	9.84 ± 12.58
Systemic absorption	0.07 ± 0.10	0.32 ± 0.39	5.42 ± 7.07