Hello every one,
As you know, there are many reports about the antibacterial properties of chitosan and its quaternary derivatives. We prepared a new quaternary derivative of chitosan and tried to investigate its antibacterial effect against some Gram-positive & Gram-negative bacteria. We used disc diffusion method to evaluate the antibacterial activity of chitosan and its quaternary derivative, but, surprisingly we didn’t observed any inhibition zone around the sample of chitosan and the sample of its quaternary derivative. What was wrong? How we have to do this experiment?
I think the diffusion of a large polysaccharide like chitosan is more difficult and is slower than small antimicrobial agents (e.g. antibiotics).
Now, can anyone tell me the exact process that we should do for investigation of chitosan antibacterial effect?
Hello Farzad Seidi,
To overcome the diffusion problem due to small pores of solid agar, you can opt for broth based experiment supported by colorimetric, fluorometric methods and plate count too.
Just for an example,
Fluorometric/ colorimetric ALAMAR BLUE ASSAY...
Hope, the methods in following references may help you.
PMID: 15980327
PMID: 9466742
PMID: 12121966
Good luck
Regards
A simple method to test for antibacterial activity of a soluble substance like chitosan is the MIC (minimal inhibitory concentration) assay. Make a set of 2-fold serial dilutions of it in growth medium, add a small number of bacterial cells, and determine the lowest concentration of the compound that prevents multiplication of the cells, based on optical density.
You can use the approach described in:
Sousa P, Almeida A, Girão AV, Fateixa S, Trindade T (2014) Multiple Emulsion Templating of Hybrid Ag/SiO2 Capsules for Antibacterial Applications. Particle & Particle Systems Characterization. Doi: 10.1002/ppsc.201400168.
Figueiredo A, Figueiredo AGPR, Silva NHCS, Timmons AB, Almeida A, Silvestre AJD, Freire CSR (2015). Antimicrobial bacterial cellulose nanocomposites prepared by in situ polymerization of 2-aminoethyl methacrylate. Carbohydrate Polymers.
Try broth dilution techniques and growth curve OD measurement for 4-12 hours with various concentration of your compound
I recommend you contact Dr. Patricia miranda.
has extensive experience in the field and has developed products for various diseases based on chitosan.
Dra Patricia Miranda. [email protected]
I have been worked with many esential oil and you could try with the cylinder-plate method or with some cosolvents system.( I tried many of them if you want send me a mail- [email protected])
A cylinder-plate method for the quantitative determination of diffusion of antibacterial drugs from ointmentsElmer M. Plein andJoy Bickmore Plein
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