Comparison of Perfluorocarbon Liquids Cytotoxicity Tests: Direct Contact Versus the Test on Liquid Extracts

Abstract

The purpose of this study is to compare the in vitro cytotoxicity tests according to the ISO 10993-5 (2009) standards using direct contact and the test on liquid extracts of compounds previously identified as possible toxic impurities in perfluorocarbon liquids (PFCLs) for use in vitreoretinal surgery. Compounds including perfluorooctanoic acid (PFOA), 1H-perfluorooctane (1H-PFO), 2H-tridecafluoro-2-methylpentane, 1H,2H-octafluorocyclopentane, and 2H,3H-decafluoropentane were analyzed by 19 F NMR before and after extraction using an aqueous solution and tested by both the direct contact and liquid extract tests in L929, BALB 3T3, and ARPE-12 cells. The concentration that reduced in vitro cell viability by 30%, the cytotoxicity concentration threshold (CC 30 ), was determined for each compound. 19 F NMR spectroscopy confirmed the immiscibility of perfluoro-n-octane (PFO) and 1H-PFO and the solubility of PFOA with the extraction vehicle. The other samples reacted with the extraction vehicle, releasing fluoride ions. Using the direct contact test, the CC 30 of PFOA, 1H-PFO, 2H-tridecafluoro-2-methylpentane, 1H,2Hoctafluorocyclopentane, and 2H,3H-decafluoropentane corresponded to 48 124, 50, 14, 8035, and 46 ppm, respectively. The method on liquid extracts did not detect cytotoxicity in three out of five tested compounds, and CC 30 could not be determined. In conclusion, the in vitro cytotoxicity test by direct contact revealed a positive correlation between cell toxicity and the concentration of the tested substance. Conversely, the test on liquid extracts hardly detected the cytotoxicity of toxic impurities in PFCLs. Thus, only the cytotoxicity test by direct contact, according to ISO 10993-5 (2009), is a sensible and reliable method to detect possible cytotoxic impurities in PFCLs to guarantee patient safety.

Pars plana vitrectomy for retinal detachment using perfluoro-n-octane as intraoperative tamponade: a multicenter, randomized, non-inferiority trial

“…Nevertheless, some researchers argued that based on the norms and recommendations of ISO 10993-5 (2009), the only reliable method to detect any potential cytotoxicity of PFCLs is by direct contact, not liquid extracts. Since testing of liquid extracts may not detect toxic impurities insoluble in water due to the incompatibility of PFCLs with aqueous solutions, which may lead to false negative results [30] . Moreover, some scholars raised questions regarding the validity of cytotoxicity test methods currently employed to certify the safety of PFO lots, and proposed new cytotoxicity test methods for volatile substances [31] .…”

“…Consequently, the clinical need for PFCLs has therefore prompted production techniques improvement to purification. In addition, a coordinated effort must be made between researchers, clinicians, companies and health authorities to establish strict standards regulating manufacturing, purification and biosafety control requirements [30,32] . There are some limitations in our study.…”

Transition metal complexes of the (2,2,2-trifluoroethyl)phosphinate NOTA analogue as potential contrast agents for 19F magnetic resonance imaging

“…12,13 Although some adverse health effects have been reported in the latter application, 14 they have been attributed mainly to the presence of trace toxic impurities like non-fully substituted hydrocarbons and other derivatives. 15,16 However, although PFCs show high stability, it was recently found that they cause some environmental risks. 17 The compounds originally used for 19 F MRI are not watersoluble and have been used in the form of nano-or microemulsions which complicates some applications (e.g.…”