Usability of a spot test for isothiazolinones

Abstract

K E Y W O R D S : benzisothiazolinone, contact allergy, methylchloroisothiazolinone, methylisothiazolinone, spot test Contact allergy to isothiazolinones is frequent, especially to methylisothiazolinone (MI) alone and combined with methylchloroisothiazolinone (MCI/MI). While the incidence of MI allergy is decreasing, 1 it remains clinically relevant, in part due to occupational exposures.A spot test kit for detection of isothiazolinones in industrial water systems is commercially available (Tintometer Group) (http://www. lovibondwater.com/product/isothiazoline.aspx). This spot test was recently used to identify a clinically relevant but undeclared exposure to MI in the raw material of a facial gel mask, 2 suggesting the clinical value of the test.The manufacturer states that the test can detect isothiazolinones down to the level of 0.75 ppm; however, it is unclear whether this applies to the most relevant isothiazolinones responsible for contact allergy. We examined the sensitivity of this spot test against 3 common isothiazolinones, and report two clinical cases where application of the test helped us identify MI and MCI/MI sources in allergic patients. SPOT TESTThe spot test kit for isothiazolinones (Tintometer Group) was tested against 3 common isothiazolinones/mixtures: MCI/MI (CAS no. 55965-84-9); MI (CAS no. 2682-20-4); and benzisothiazolinone (BIT) (CAS no. 2634-33-5). The spot test is based on a colorimetric method, where the presence of isothiazolinone derivatives results in a blue color change. The exact basis of the test method is company proprietary information and the analytes used come in a coded form. A reading scale is provided (Figure 1). The detection limit of the test as declared by the manufacturer is 0.75 ppm, where the solution gets a blue undertone, and ends at 7.5 ppm, where it becomes dark blue. The test is negative if no color change occurs, that is, if the solution (in water) remains clear (Figure 1). A range of test solutions corresponding to the reading scale of the spot test (0-7.5 ppm) and added higher concentrations of 10-200 ppm (Table 1) were prepared by using sterile-filtered water (Fresenius Kabi, Bad Homburg vor der Höhe, Germany) and the patch test materials: MCI/MI (allergEAZE, SmartPractice, Phoenix, Arizona) 200 ppm, MI (allergEAZE) 2000 ppm, and BIT (97% Sigma-Aldrich, Copenhagen, Denmark) in w/w% as shown in Table 1. The spot tests were performed with the 5 coded reagents provided in the test kit in accordance with instructions from the manufacturer. The mixing of test solution and reagents takes approximately 5 minutes. The developed color was compared to the test kit standards. Unblinded readings were conducted by one investigator (LA or UFF), approximately 10 seconds after the mixing according to instructions.

Isothiazolinones Common in Children's Toy Slime

“…This finding is unsurprising given that the colorimetric spot test was designed to detect MCI/MI, but its limit of detection for MI (7.5 ppm) is much lower than that of the HPLC-UV (1.3 ppm). 2 However, these concentrations are well below reported sensitization and elicitation thresholds for MI in particular (~500 and ~50 ppm, respectively), suggesting that colorimetric test false-negatives are unlikely to be clinically significant. 27 The identification of isothiazolinones in commercial slime products is salient, given the complex regulation surrounding the toy industry that does not mandate declaration of toy ingredients, in stark contrast to the regulation of cosmetic products.…”

“…The reason for this false positivity remains elusive, and the presence of other, less well-known isothiazolinone derivatives may not be fully excluded. Two samples containing MI were missed by this colorimetric test; the colorimetric spot test's propensity to underestimate the presence of MI may lead to false-negative results, for MI in particular 2 . Overall, the colorimetric spot test seems to have little utility as a screening assay for the presence of MCI/MI or BIT 2 ; it seems far from specific and is likely not sensitive enough to reliably identify MI, especially at lower concentrations.…”

“…However, it is clearly less sensitive for MI and likely identified this derivative only in sample #5 because it was present in high-enough concentrations. Although the colorimetric spot test has been shown to score well for the detection of BIT, this compound was not identified in any of the slime samples; OIT was not detected either, although the capacity of the colorimetric spot test to detect OIT has not been elucidated 2 …”

“…There were also a number of false-negative results with the colorimetric spot test, as 2 of the 3 negative slime samples contained MI as demonstrated by HPLC-UV. This finding is unsurprising given that the colorimetric spot test was designed to detect MCI/MI, but its limit of detection for MI (7.5 ppm) is much lower than that of the HPLC-UV (1.3 ppm) 2 . However, these concentrations are well below reported sensitization and elicitation thresholds for MI in particular (~500 and ~50 ppm, respectively), suggesting that colorimetric test false-negatives are unlikely to be clinically significant 27 …”

“…Two samples containing MI were missed by this colorimetric test; the colorimetric spot test's propensity to underestimate the presence of MI may lead to false-negative results, for MI in particular. 2 Overall, the colorimetric spot test seems to have little utility as a screening assay for the presence of MCI/MI or BIT 2 ; it seems far from specific and is likely not sensitive enough to reliably identify MI, especially at lower concentrations. A strength of this investigation is that it is the first to demonstrate the relatively low specificity of this colorimetric spot test as evidenced by likely false-positive results with HPLC-UV.…”