Optical control of antibacterial activity

Journal name:
Nature Chemistry
Year published:
Published online


Bacterial resistance is a major problem in the modern world, stemming in part from the build-up of antibiotics in the environment. Novel molecular approaches that enable an externally triggered increase in antibiotic activity with high spatiotemporal resolution and auto-inactivation are highly desirable. Here we report a responsive, broad-spectrum, antibacterial agent that can be temporally activated with light, whereupon it auto-inactivates on the scale of hours. The use of such a ‘smart’ antibiotic might prevent the build-up of active antimicrobial material in the environment. Reversible optical control over active drug concentration enables us to obtain pharmacodynamic information. Precisely localized control of activity is achieved, allowing the growth of bacteria to be confined to defined patterns, which has potential for the development of treatments that avoid interference with the endogenous microbial population in other parts of the organism. 

At a glance


  1. Control of bacterial growth with a photoswitchable antibacterial agent.
    Figure 1: Control of bacterial growth with a photoswitchable antibacterial agent.

    a, Structure of quinolone antibacterial agents. A typical quinolone bears a fluorine atom at the R1, R2 and/or R4 position, a nitrogen-containing, saturated ring at R3, and an alkyl moiety at R5. b, Azobenzene-containing quinolones: trans and cis geometrical isomers and isomerization processes. Irradiation at 365 nm leads to conversion of the trans isomer to the cis isomer, and visible light switches the cis isomer back to the trans form. The cis to trans isomerization also occurs thermally at ambient temperature, albeit at a slower rate than the photoisomerization. c,d, Growth curves of E. coli CS1562 with different concentrations of non-irradiated (c) and irradiated (d) 2. Growth was not inhibited at the concentrations tested for the non-irradiated form of 2 (95% trans isomer), whereas after irradiation at 365 nm (89% cis isomer) significant inhibition of growth at concentrations of ≥16 µg ml−1 was observed. All solutions were irradiated before inoculation. Error bars show standard deviations calculated from measurements in triplicate.

  2. Auto-inactivation of antibiotic activity.
    Figure 2: Auto-inactivation of antibiotic activity.

    a, Thermal cis–trans isomerization of compound 2 at 37 °C at a concentration of 21 × 10−6 M in water. Absorbance was measured at λmax of trans-2 (368 nm). The half-life is 2.08 h. b, Growth curves of E. coli CS1562 incubated with 40 µg ml−1 of compound 2, which was activated at 365 nm at different times before incubation. Bacterial growth was inhibited by compound 2 irradiated immediately before incubation. Over time, the antibacterial activity of compound 2 decreases as a result of thermal cis–trans isomerization. The samples that were irradiated 3 h before incubation show the same growth pattern as samples to which the antibiotic was not added, indicating that the antibacterial activity is lost completely 3 h after initial photoswitching from the inactive trans form to the active cis form. Error bars show standard deviations calculated from measurements in triplicate.

  3. Pharmacodynamic study with photoswitchable quinolone shows that antibacterial effect is exhibited in the exponential phase.
    Figure 3: Pharmacodynamic study with photoswitchable quinolone shows that antibacterial effect is exhibited in the exponential phase.

    Bacteria incubated without compound 2 exhibit a normal growth pattern (inverted pink triangles). In contrast, bacteria incubated with 40 µg ml−1 of 365-nm-irradiated compound 2 have not grown at all (red circles). Bacteria incubated with 2 (irradiation at 365 nm, mainly active cis isomer present) and illuminated with visible light after 30 min (black squares, mainly inactive trans form) while the bacteria are still in the lag phase show a normal growth pattern, comparable with bacteria incubated without compound 2. Bacteria incubated with 365-nm-irradiated compound 2 and illuminated with visible light after 60 min (blue triangles), when the bacteria have already entered the exponential phase, do not show growth, comparable with bacteria incubated with 365-nm-irradiated compound 2 without illumination with visible light. This indicates that this quinolone analogue exhibits its antibacterial effect in the exponential phase and not in the lag phase. Error bars show standard deviations calculated from measurements in triplicate.

  4. Bacterial patterning with light.
    Figure 4: Bacterial patterning with light.

    a, An agar plate was prepared containing 50 µg ml−1 of compound 2. A mask was placed on top of the agar plate and the plate was then irradiated at 365 nm. Only in the exposed part was compound 2 switched to its active cis form. Afterwards, the plate was inoculated with E. coli CS1562 and incubated overnight at 37  °C. Bacterial growth is only observed in the part that was covered by the mask. b, Result of a typical bacterial patterning experiment. A Taijitu symbol was used as the mask.


9 compounds View all compounds
  1. (E)-1-Ethyl-7-((4-methoxyphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 1 (E)-1-Ethyl-7-((4-methoxyphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  2. (E)-1-Ethyl-7-((4-methoxy-3-methylphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 2 (E)-1-Ethyl-7-((4-methoxy-3-methylphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  3. (E)-1-Ethyl-7-((4-methoxy-3,5-dimethylphenyl)-diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 3 (E)-1-Ethyl-7-((4-methoxy-3,5-dimethylphenyl)-diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  4. (E)-1-Ethyl-7-((4-methoxy-2,3,6-trimethylphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 4 (E)-1-Ethyl-7-((4-methoxy-2,3,6-trimethylphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  5. (E)-1-Ethyl-7-((2-methoxy-5-methylphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 5 (E)-1-Ethyl-7-((2-methoxy-5-methylphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  6. (E)-1-Ethyl-7-((3-fluoro-4-methoxyphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 6 (E)-1-Ethyl-7-((3-fluoro-4-methoxyphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  7. (E)-7-((3,5-Difluoro-4-methoxyphenyl)diazenyl)-1-ethyl-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 7 (E)-7-((3,5-Difluoro-4-methoxyphenyl)diazenyl)-1-ethyl-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  8. (E)-1-Ethyl-7-((5-fluoro-2-methoxyphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 8 (E)-1-Ethyl-7-((5-fluoro-2-methoxyphenyl)diazenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
  9. (E)-7-((3,4-Dimethoxyphenyl)diazenyl)-1-ethyl-4-oxo-1,4-dihydroquinoline-3-carboxylic acid
    Compound 9 (E)-7-((3,4-Dimethoxyphenyl)diazenyl)-1-ethyl-4-oxo-1,4-dihydroquinoline-3-carboxylic acid


  1. Carlet, J. et al. Society's failure to protect a precious resource: antibiotics. Lancet 378, 369371 (2011).
  2. Martinez, J. L. Antibiotics and antibiotic resistance genes in natural environments. Science 321, 365367 (2008).
  3. Tello, A., Austin, B. & Telfer, T. C. Selective pressure of antibiotic pollution on bacteria of importance to public health. Environ. Health Perspect. 120, 11001106 (2012).
  4. Goossens, H., Ferech, M., Vander Stichele, R. & Elseviers, M. Outpatient antibiotic use in Europe and association with resistance: a cross-national database study. Lancet 365, 579587 (2005).
  5. Kemper, N. Veterinary antibiotics in the aquatic and terrestrial environment. Ecol. Indic. 8, 113 (2008).
  6. Martínez, J. L. & Baquero, F. Mutation frequencies and antibiotic resistance. Antimicrob. Agents Chemother. 44, 17711777 (2000).
  7. Davies, J. Inactivation of antibiotics and the dissemination of resistance genes. Science 264, 375382 (1994).
  8. Stockley, J. M. European antibiotic awareness day 2012: getting smart about antibiotics, a public–professional partnership. J. Infect. 65, 377379 (2012).
  9. Brinster, S. et al. Type II fatty acid synthesis is not a suitable antibiotic target for Gram-positive pathogens. Nature 458, 8385 (2009).
  10. Zlitni, S. & Brown, E. D. Drug discovery: not as fab as we thought. Nature 458, 3940 (2009).
  11. Gorostiza, P. & Isacoff, E. Y. Optical switches for remote and noninvasive control of cell signaling. Science 322, 395399 (2008).
  12. Mayer, G. & Heckel, A. Biologically active molecules with a ‘light switch’. Angew. Chem. Int. Ed. 45, 49004921 (2006).
  13. Kocer, A., Walko, M., Meijberg, W. & Feringa, B. L. A light-actuated nanovalve derived from a channel protein. Science 309, 755758 (2005).
  14. Bonardi, F., London, G., Nouwen, N., Feringa, B. L. & Driessen, A. J. M. Light-induced control of protein translocation by the SecYEG complex. Angew. Chem. Int. Ed. 49, 72347238 (2011).
  15. Szymanski, W., Beierle, J. M., Kistemaker, H. A. V., Velema, W. A. & Feringa, B. L. Reversible photocontrol of biological systems by the incorporation of molecular photoswitches. Chem. Rev. 113, 61146178 (2013).
  16. Szymański, W., Yilmaz, D., Koçer, A. & Feringa B. L. Bright ion channels and lipid bilayers. Acc. Chem. Res. http://dx.doi.org/10.1021/ar4000357 (2013).
  17. Schierling, B. et al. Controlling the enzymatic activity of a restriction enzyme by light. Proc. Natl Acad. Sci. USA 107, 13611366 (2010).
  18. Tochitsky, I. et al. Optochemical control of genetically engineered neuronal nicotinic acetylcholine receptors. Nature Chem. 4, 105111 (2012).
  19. Stein, M. et al. Azo-propofols: photochromic potentiators of GABAA receptors. Angew. Chem. Int. Ed. 51, 1050010504 (2012).
  20. Polosukhina, A. et al. Photochemical restoration of visual responses in blind mice. Neuron 75, 271282 (2012).
  21. Lee, W., Li, Z-H., Vakulenko, S. & Mobashery, S. A light-inactivated antibiotic. J. Med. Chem. 43, 128132 (2000).
  22. Hohsaka, T., Kawashima, K. & Sisido, M. Photoswitching of NAD+-mediated enzyme reaction through photoreversible antigen–antibody reaction. J. Am. Chem. Soc. 116, 413414 (1994).
  23. Abell, A. D. et al. Investigation into the P3 binding domain of m-calpain using photoswitchable diazo- and triazene-dipeptide aldehydes: new anticataract agents. J. Med. Chem. 50, 29162920 (2007).
  24. Mourot, A. et al. Rapid optical control of nociception with an ion-channel photoswitch. Nature Methods 9, 396402 (2012).
  25. Velema, W. A., Van der Toorn, M., Szymanski, W. & Feringa, B. L. Design, synthesis, and inhibitory activity of potent, photoswitchable mast cell activation inhibitors. J. Med. Chem. 56, 44564464 (2013).
  26. Drlica, K. & Zhao, X. DNA gyrase, topoisomerase IV, and the 4-quinolones. Microbiol. Mol. Biol. Rev. 61, 377392 (1997).
  27. Domagala, J. M. Structure–activity and structure–side-effect relationships for the quinolone antibacterials. J. Antimicrob. Chemoth. 33, 685706 (1994).
  28. Mitscher, L. A. Bacterial topoisomerase inhibitors: quinolone and pyridone antibacterial agents. Chem. Rev. 105, 559592 (2005).
  29. Beharry, A. B. & Woolley, G. A. Azobenzene photoswitches for biomolecules. Chem. Soc. Rev. 40, 44224437 (2011).
  30. Bandara, H. M. D. & Burdette, S. C. Photoisomerization in different classes of azobenzene. Chem. Soc. Rev. 41, 18091825 (2012).
  31. Merino, E. Synthesis of azobenzenes: the coloured pieces of molecular materials. Chem. Soc. Rev. 40, 38353853 (2011).
  32. Mendonça, C. R. et al. in Molecular Switches 2nd edn (eds Feringa, B. L. & Browne, W. R.) Ch. 12 (Wiley, 2011).
  33. Austin, E. A., Graves, J. F., Hite, L. A., Parker, C. T. & Schnaitman, C. A. Genetic analysis of lipopolysaccharide core biosynthesis by Escherichia coli K-12: insertion mutagenesis of the rfa locus. J. Bacteriol. 172, 53125325 (1990).
  34. Wiegand, I., Hilpert, K. & Hancock, R. E. W. Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nature Protoc. 3, 163175 (2008).
  35. Sukul, P., Lamshöft, M., Kusari, S., Zühlke, S. & Spiteller, M. Metabolism and excretion kinetics of 14C-labeled and non-labeled difloxacin in pigs after oral administration, and antimicrobial activity of manure containing difloxacin and its metabolites. Environ. Res. 109, 225231 (2009).
  36. Lester, H. A. et al. Electrophysiological experiments with photoisomerizable cholinergic compounds: review and progress report. Ann. NY Acad. Sci. 346, 475490 (1980).
  37. Wright, D. H., Brown, G. H., Peterson, M. L. & Rotschafer, J. C. Application of fluoroquinolone pharmacodynamics. J. Antimicrob. Chemother. 46, 669683 (2000).
  38. Kim, K., Lee, B. U., Hwang, G. B., Lee, J. H. & Kim, S. Drop-on-demand patterning of bacterial cells using pulsed jet electrospraying. Anal. Chem. 82, 21092112 (2010).
  39. Park, T. J. et al. Protein nanopatterns and biosensors using gold binding polypeptide as a fusion partner. Anal. Chem. 78, 71977205 (2006).
  40. Bergogne-Berezin, E. Treatment and prevention of antibiotic associated diarrhea. Int. J. Antimicrob. Agents 16, 521526 (2000).
  41. Vippagunta, S. R. et al. Structural specificity of chloroquine–hematin binding related to inhibition of hematin polymerization and parasite growth. J. Med. Chem. 42, 46304639 (1999).

Download references

Author information


  1. Centre for Systems Chemistry, Stratingh Institute for Chemistry, University of Groningen, Nijenborgh 4, 9747 AG, Groningen, The Netherlands

    • Willem A. Velema,
    • Mickel J. Hansen,
    • Wiktor Szymanski &
    • Ben L. Feringa
  2. Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, Nijenborgh 7, 9747 AG, Groningen, The Netherlands

    • Jan Pieter van der Berg &
    • Arnold J. M. Driessen
  3. Zernike Institute for Advanced Materials, University of Groningen, Nijenborgh 4, 9747 AG, Groningen, The Netherlands

    • Arnold J. M. Driessen &
    • Ben L. Feringa


B.L.F., W.A.V. and W.S. conceived the project and wrote the manuscript. W.A.V. designed the molecules. W.A.V. and M.J.H. performed the synthesis. W.A.V. and J.P.B. performed bacterial growth studies. B.L.F., W.S. and A.J.M.D. guided the research. All authors discussed the results and implications and commented on the manuscript at all stages.

Competing financial interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to:

Author details

Supplementary information

PDF files

  1. Supplementary information (987 KB)

    Supplementary information

Additional data