Chemical Abstract Series Registry (CAS) numbers were obtained from the Chemotechnique Diagnostics, Patch Test Products & Reference Manual 2014¹⁹ and used to search for published VPs from the following online chemical databases: United States National Library of Medicine, National Center for Biotechnology Information, PubChem Open Chemistry Database,^20,21 Chemical Book ²² (an online data set of materials safety data sheets), and Chemical Laboratory Information Profiles²³ (a database of the American Chemical Society detailing physical and safety information on select chemical compounds originally published in the Journal of Chemical Education). Vapor pressure can be obtained from experimental data and estimated/extrapolated using the Antoine equation:²⁴

where P indicates vapor pressure; T indicates temperature; and A, B, and C indicate substance-specific constants.

Small discrepancies can be found between the databases, and when these were encountered, values from the PubChem followed by CLIP databases were utilized. Vapor pressure and associated temperature were recorded as millimeters of mercury and degrees celsius, respectively. If multiple VP values were listed, VP at temperatures that most approximate indoor storage conditions (20°C and 25°C) were chosen. As VP varies nonlinearly with temperature per the Clasius-Clapeyron equation, VP at temperatures much higher than patch testing temperatures are largely irrelevant:²⁵

where P indicates vapor pressure at T; T indicates temperature, ΔAH_vap indicates enthalpy of vaporization specific for substances; and R, gas constant (8.3145 J/(mol I K).

For allergen mixes, the individual component with the highest VP was chosen as the overall VP, based on the assumption that the most volatile substance would evaporate first. Documented VPs less than 0.001 mm Hg were listed as <0.001 mm Hg and assumed to be clinically equivalent.

Given the ambient and mild conditions during patch testing, it is unlikely that solid compounds will sublimate from solid to gas phases. Thus, VP was not useful with solids in suspension or compounds that do not dissolve in the vehicle.²⁶ Some metals salts are not soluble in petrolatum (eg, nickel sulfate), and therefore, VP is not helpful in predicting stability. To be comprehensive, we included VP for all allergens in the ACDS Core Allergen Series, including metal salts.

There are few established guidelines for volatility, and as such, categorical cutoff values for volatility are subjective ²⁷; relevance for patch test preparations is the amount of intact allergen present in the patch test chamber, and the temperature in which patch testing is usually performed. After discussion, we used the consensus cutoff values of 1 mm Hg or higher for high volatility, less than 1 to greater than 0.001 mm Hg for moderate volatility, and 0.001 mm Hg or less for low volatility at 25°C given data availability.

Table 2 lists data for the ACDS Core Allergens with published VPs organized within categories of high, medium, and low volatility in alphabetical order. Data for acetone, ethanol, and water vehicles were included as reference points. Allergens with VP data at high temperatures were included if data were not available at ambient or near-ambient temperatures. Allergens without reported VPs were excluded from Tables 2 and summarized in Table 3. For mixtures, available data for each of the components were listed but for the purpose of stratification, the component with the highest VP was used as the overall VP. However, concentration of individual components of a mixture (eg, fragrance mix I) is not reported by the supplier.¹⁹

This study documents the VPs of ACDS Core Allergens. While the most conclusive studies involve analysis of “in use” allergens and patch test preparations, these data are not available for most allergens. The information published herein provides additional information to clinicians regarding 1 parameter that may affect the stability of common allergens. The volatility of an allergen should also be considered when compounding allergens within the dermatology clinic for diagnostic use.

Vapor pressure is defined by the Occupational Safety and Health Administration of the US Department of Labor in the Code of Federal Regulation as “a measure of a liquid's propensity to evaporate. The higher the VP, the more volatile the liquid and, thus, the more readily the liquid gives off vapors.” ¹⁷ More theoretically,it is the pressure of a vapor in thermodynamic equilibrium with its condensed phases (liquid and solid)in a closed system.In such a system, although there is constant change among the gaseous, liquid, and solid phases; there is no net change. Another way to think about VP is that it essentially measures the tendency for an atom or molecule to escape into the gaseous phase from condensed phases.²⁶ For example, a vacuum container, at time 0, contains liquid waterat20°C. Because the VP of liquid water at 20°C is approximately 760 mm Hg and the ambient pressure is 0 (vacuum), liquid water will immediately vaporize until VP is achieved or all of the liquid water has vaporized, whichever occurs first. Once at equilibrium, any additional application of energy such as heat will cause the liquid phase to change into gas phase, forming gas bubbles. As VP varies positively with volatility, the higher the VP, the higher the volatility and the higher the rate of loss of the substance from condensed to gaseous states.¹⁷

Based on our findings on VP alone, formaldehyde, acrylates (hydroxyethyl methacrylate, ethyl acrylate, and methyl methacrylate), and propylene glycol were predicted to have shorter shelf lives. This is consistent with previous research. Siegel et al¹⁰ found that formaldehyde content measured in reagents obtained from a single patch test clinic and directly from the supplier was consistent with the label stated content upon receipt at the laboratory. Upon re-assay after 1 year of undisturbed storage under refrigerated conditions, the formaldehyde reagents supplied in a syringe container had formaldehyde losses of 41% and 67%, while that supplied in an opaque plastic dropper had lost 31% of the formaldehyde from the water vehicle. While significant losses were observed for both container types, this preliminary observation suggests that a more air-tight container may help preserve volatile allergen integrity. The Antoine and Cassius-Clapeyron equations and other theories of thermodynamics were developed under the assumption of a closed system at equilibrium.^24–26 In real-world conditions, such as with patch testing compound storage and use, a closed system is not realistic; however, a pseudo-steady state can be achieved. With semiclosed systems such as a leaky container, steady state would be achieved where there is a steady diffusion of volatile gasses from the counter but with minimal change in the partial pressure within the container. This only occurs if there is an equal loss of patch test compounds from the solution to the gas. Thus, with time, there can be significant losses, especially if VP is high, providing a rationale for more air-tight storage methods.

Issues with acrylate allergen stability, specifically methyl methacrylate, were first documented by Kanerva et al.²⁸ They documented false-negative or questionable patch test results with methyl methacrylate obtained from 2 different manufacturers; one had nondetectable methyl methacrylate levels and the other only 25% of the labeled amount. Goon et al⁹ studied methyl methacrylate stored in syringes and IQ chambers. They documented that methyl methacrylate stored in syringes at room temperature lost more than 20% of the labeled concentration within 2 weeks. When stored at −16°C, the loss was less than 20% at day 128, but increased to more than 20% of the initial concentration by 6 months. Loss was more rapid in IQ chambers under all conditions. Hypothesized reasons included evaporation or spontaneous polymerization. Siegel et al documented that the concentration of methyl methacrylate in stored syringes varied from the tip to the plunger of the syringe. The concentration at the tip of the syringes averaged 42% less than subsequent aliquots, suggesting that the major source of loss was due to volatility.¹⁰ In addition, both Goon et al⁹ and Siegel et al¹⁰ observed significant loss of methyl methacrylate during compounding with petrolatum. During this process, petrolatum must be heated to higher than 65°C to melt the petrolatum, which can cause significant loss of allergen from volatilization as VP increases nonlinearly to increasing temperature.

Based on VP, volatility should not be a factor in the stability of paraben mix constituents, rubber allergens (carba mix ingredients and N,N-diphenyl-p-phenylenediamine), 2 fragrances (coumarin and farnesol), and 3 sunscreens (benzophenone-3 and -4 as well as 2-ethylhexyl-4-methoxycinnamate), and also benzocaine, epoxy resin, methyldibromoglutaronitrile, cetyl alcohol, stearyl alcohol, and tocopherol. This is consistent with previous publications. Gruvberger et al⁵ tested 4 different concentrations of methyldibromoglutraonitrile at 1 year and found that all 4 were stable. Both patients tested to 40-year-old benzocaine, and 3 of 4 patients tested to 40-year-old epoxy resin reacted on patch testing.⁶ We found no previous reports of stability testing to paraben mix, carba mix, coumarin, farnesol, cetyl alcohol, stearyl alcohol, sunscreens, or tocopherol.

This review provides additional information to clinicians on potential stability issues based on VP and is consistent with previously reported data on the instability of formaldehyde, acrylates, and fragrance materials. Specific reported stability data for a given allergen supersedes predictions based on VP. However, given the lack of experimental stability data of patch testing compounds, reliance on VP as a proxy for volatility may be a helpful tool for clinicians when compounding a patch test reagent or assessing the probability of a potential false-negative test.

The stability of commercial patch test allergens continues to be of concern as it impacts not only clinical diagnostic accuracy but also reliability of epidemiologic data reported in the literature. Potentially, patch test reagent stability/storage issues due to volatility may be minimized by use of more air-tight multidose reagent containers, sealed single application dispensers, storage at lower temperatures, and reliable beyond-use date labeling of multidose containers by patch test reagent suppliers. In addition, in an effort to minimize risk of false-negatives, allergens known to be highly volatile such as fragrances and those within the predicted high and medium volatility range should not be aliquoted and prepared until just before the application.