October 18-21, 2015
The identification of the bis(2,4-di-tert-butylphenyl)phosphate (bDtBPP) by Hammond et al was a key to understand the detrimental effect on cell growth that may occur in single use systems composed of polyethylene. The growth inhibition depends on the concentration and cell line, while known standard cytotoxicity assays (i.e. USP<87>) are insensitive to the levels shown to have detrimental effects. There is therefore no universal standardized cell growth testing allowing determining an absolute quantitative concentration detrimental to all specific cell lines. The most sensitive cells lines tested showed a dose response that challenges the limit of detection of existing analytical methods.
Concentrations of bDtBPP are linked to the secondary degradation via gamma irradiation of a common antioxidant and have been shown to be related both to the starting concentration of the antioxidant and to the thermal processing parameters. Comprehensive understanding of this relationship requires the use of design of experiment. Cell based assays used to support a DoE approach have proven to be cumbersome.
This now known and well understood phenomenon presented a challenge for the development of a new film composed of polyethylene. The bDtBPP concentration of concern is below detection thresholds for existing standard cell based assays and analytical techniques such that a physico-chemical analysis had to be designed to support the film development project.
Extraction methods and analytical methods were specifically developed by using appropriate polymer swelling solvent and chromatographic tools to extract and detect the bDtBPP in worse case conditions. The detection limit of the bDtBPP in worse case conditions is equivalent to the lowest bDtBPP concentrations to be detrimental to the cell growth for the common cell lines.
Data are presented detailing the development of the analytical methods and their usefulness relative to the concentrations shown to be of concern.
Samuel Dorey, Ina Pahl, Isabelle Uettwiller, Sebastien Janvier, and Elke Jurkiewicz, "Limitation and detection of bis(2,4-di-tert-butylphenyl)phosphate (bDtBPP) from bioprocess container materials" in "Single-Use Technologies: Bridging Polymer Science to Biotechnology Applications", Ekta Mahajan, Genentech, Inc., USA Gary Lye, University College London, UK Eds, ECI Symposium Series, (2015). https://dc.engconfintl.org/biopoly/8