Rapid microbiological sterility test for cytotoxic batch control: is drug complexation effective for restoring microbiological growth?

Sangnier Maïté1, Bouguéon Guillaume1,2, Dubois Véronique3,4, Crauste-Manciet Sylvie1,2
(1) Pharmaceutical Technology Department, Bordeaux university hospital (CHU de Bordeaux), France
(2) ARNA Laboratory-ChemBioPharm U1212 INSERM - UMR5320, CNRS - University of Bordeaux, France
(3) Fundamental Microbiology and Pathogenicity Laboratory UMR-CNRS 5234 – University of Bordeaux, France
(4) Microbiology department, Bordeaux university hospital (CHU de Bordeaux), France

Standardization and batch production of cytotoxic drugs is of growing interest in hospital pharmacies. In accordance to French GMPs1 the batch release of a sterile preparation apart physical-chemical includes microbiological control i.e. sterility testing of the batch. Rapid Microbiological Methods (RMM) are interesting alternative considering the short shelf-life commonly given in our practice (from few days to few weeks). In this context, our objective was to investigate the efficiency of two drug complexation materials: activated carbon (AC) and ion exchange resins (IER) for restoring the microbiological growth.
IER used was polymer beads presents in BacT/ALERT® (BA) bottles (unknown composition), weak and strong cation exchange resin (Purolite C104E+ and C150S) and weak and strong anion exchange resins (Amberlyst A-21 and Dowex 1X8-200). Cytotoxic drug used as reference was 5-fluorouracil (5FU) at 5 final concentrations (50, 25, 10 and 5mg/mL) in 10mL samples. Exposure contact time (5min, 30min, 1h and 24h) and concentrations of material (30, 60, 90, 150, 200 and 300mg/mL for AC and 200, 300, 400 and 500mg/mL for IER ) were the variables investigated. The efficiency of complexation by the material was evaluated by 3 measuring residual concentration of 5FU after treatment using HPLC method (Kinetex 2.6μm XB-C18, 80% MetOH and 20% HCOOH 0.1V/V, 266nm)2. The fully complexed 5FU was contaminated with Staphylococcus coagulase + inoculum to evaluate restoration of microbiological growth by BA system. A test for routine implementation using pre-filled syringes with complexation equipment was also performed.
IER have not shown sufficient efficiency to complex 5FU in a single pass even though anion exchange resins are the most effective. Regarding the AC treatment, the greater the amount of AC used, the lower the 5FU concentration was remaining. The minimum AC concentration to be used is 90, 150 and 300mg/mL for 5, 10 and 25mg/mL. A standard pre-treatment with 90mg/mL AC will allow the use of RMM with diluted 5FU (5mg/ml) as recommended by the European Pharmacopeia3. Test for routine implementation was conclusive showing the restoration of Staphylococcus coagulase+ growth in BA when using AC.
In conclusion, this complexation method via AC treatment could be adapted to sterility testing of batch preparations containing microbiological growth inhibitors.

1 GMPs : French Good Manufacturing Practices for hospital and community pharmacies, (Bonnes Pratiques de préparations) ANSM 3/12/2007.
2 Delmas A, Gordien JB, Bernadou JM, Roudaut M, Gresser A, Malki L, et al. Quantitative and qualitative control of cytotoxic preparations by HPLC-UV in a centralized parenteral preparations unit. J Pharm Biomed Anal. 2009;49(5):1213–20.
3 European Pharmacopoeia Online 9.8 [Internet]. [cited 2019 Sep 19]. Available from: http://online6.edqm.eu/ep908/

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