Interplay of phosphate and carbonate ions with flavin photosensitizers in photodynamic inactivation of bacteria

Abstract

Photodynamic inactivation (PDI) of pathogenic bacteria is a promising technology in different applications. Thereby, a photosensitizer (PS) absorbs visible light and transfers the energy to oxygen yielding reactive oxygen species (ROS). The produced ROS are then capable of killing microorganisms via oxidative damage of cellular constituents. Among other PS, some flavins are capable of producing ROS and cationic flavins are already successfully applied in PDI. When PDI is used for example on tap water, PS like flavins will encounter various ions and other small organic molecules which might hamper the efficacy of PDI. Thus, the impact of carbonate and phosphate ions on PDI using two different cationic flavins (FLASH-02a, FLASH-06a) was investigated using Staphylococcus aureus and Pseudomonas aeruginosa as model organisms. Both were inactivated in vitro at a low light exposure of 0.72 J cm-2. Upon irradiation, FLASH-02a reacts to single substances in the presence of carbonate or phosphate, whereas the photochemical reaction for FLASH-06a was more unspecific. DPBF-assays indicated that carbonate and phosphate ions decreased the generation of singlet oxygen of both flavins. Both microorganisms could be easily inactivated by at least one PS with up to 6 log10 steps of cell counts in low ion concentrations. Using the constant radiation exposure of 0.72 J cm-2, the inactivation efficacy decreased somewhat at medium ion concentrations but reached almost zero for high ion concentrations. Depending on the application of PDI, the presence of carbonate and phosphate ions is unavoidable. Only upon light irradiation such ions may attack the PS molecule and reduce the efficacy of PDI. Our results indicate concentrations for carbonate and phosphate, in which PDI can still lead to efficient reduction of bacterial cells when using flavin based PS.

Tuning riboflavin derivatives for photodynamic inactivation of pathogens

“…Amino-containing F1 and F3 exhibited around 40% and 60% degradation respectively after the same irradiation time (Figure S4 a,c,e), whereas F2 and F4 degraded by approximately 10% and 30% respectively (Figure S 4 a,d,f.). These findings corroborate previous work showing photodegradation of amino-containing flavins in the presence of phosphate ions, which resulted in diminished bacterial PDI efficiency 41 . However, it appears that guanidino substitution in our case improves the photostability.…”

Inhibitory effects of calcium or magnesium ions on PDI

“…The quantum yield of the porphyrin based photosensitzer is around 0.77 [56], producing chiefly singlet oxygen with minor amounts of other ROS. Besides, an exclusive singlet oxygen producing photosensitizer shortly called SAPYR with a quantum yield of 0.99 [20] was purchased from the TriOptoTec GmbH, the chemical structure of the molecules has been published elsewhere [21]. Additionally, two different flavin based photosensitizers were included with a quantum yield of around 0.75 that was also purchased from Tri-OptoTec GmbH.…”

“…The herein cited examples for applied PDI make use of several photosensitizer classes, ranging from well-known photosensitizers such as methylene blue (MB), porphyrins (5,10,15,20-Tetrakis(1-methyl-4pyridinio)-porphyrin tetra(p-toluene sulfonate, briefly called TMPyP), new substances that exclusively produce singlet oxygen (SAPYR [20]) to curcumins or flavins (FLASH-02a and FLASH-06a [21]). Especially curcumins are considered safe for food applications [12].…”

“…Among others, up to date the effects of abundant substances such as calcium or magnesium ions or complex biological molecules remain mostly uninvestigated. Of course, it is known for some photosensitizer that certain chemicals inhibit [21,51] or enhance [52][53][54] the photodynamic process. One of the most prominent molecules in this context is sodium azide acting as a potent physical singlet oxygen quencher [51].…”

“…In contrast, there are also studies investigating on effects that promote the photodynamic action in presence of sodium azide [54]. Furthermore, it was recently shown that carbonate and phosphate ions, which are two prominent molecules in most environments, have detrimental effects on the chemical structure of flavin based photosensitizers [21]. Additionally, some research data concerning the photodynamic treatment of milk suggested that calcium and magnesium ions pose some issues in efficacy [44].…”