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A. Vijay, M. Bandara, H. Zhu, M. Willcox; Amoebicidal Activity of Protamine Alone and in Combination With Other Antimicrobials. Invest. Ophthalmol. Vis. Sci. 2009;50(13):5636.
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To evaluate the amoebicidal efficacy of protamine and its synergistic potential with polyhexamethylene biguanide (PHMB), furanone (Fu) and salicylic acid (SA).
The stand alone procedure described in ISO 14729 to assess the performance of contact lens disinfecting solutions was modified to test the amoebicidal activity using Acanthamoeba polyphaga ROS trophozoites. Protamine (0.5 mg/ml and 0.25 mg/ml) was tested alone and in combination with PHMB (1 ppm), Fu (10 µg/ml) and SA (4.8 mg/ml). All the test samples were prepared in phosphate buffered saline (PBS) and the amoebicidal efficacy evaluated in comparison to PBS. Optifree RepleniSH (Alcon Labs) was used as a positive control. The Acanthamoeba was inoculated into each sample to obtain a final concentration of approximately 1.0-1.5 x 105 cells/ml and then incubated at 25ºC for 6 hours. 1:10 serial dilutions of samples were performed and inoculated on non-nutrient agar plates seeded with E. coli. Plates were examined under microscope following 7 days of incubation at 32ºC and the number of survivors determined using Reed-Muench computation. Log reductions were calculated by comparing survivors from the test and control samples.
Protamine alone showed good amoebicidal activity, giving log reductions (Mean ± SD) of 1.5 ± 0.7 and 1.2 ± 0.7 at concentrations of 0.5 mg/ml and 0.25mg/ml respectively. A combination of protamine and PHMB showed good synergy. The addition of 1ppm of PHMB increased the log reductions of the test strain to 2.9 ± 0.6 and 2.2 ± 0.2 logs for 0.5 and 0.25 mg/ml of protamine respectively. PHMB alone did not have any activity against the challenge organism. No significant synergy was observed with combinations of protamine and Fu (10 µg/ml) or protamine and SA (4.8 mg/ml). Optifree RepleniSH showed a log reduction of 1.0 ± 0.7 logs.
The good activity of protamine against Acanthamoeba trophozoites and excellent synergy with PHMB indicates that protamine could be considered as a potential ingredient in contact lens disinfecting solutions.
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