Dried methanolic extract was used for assay, Broth microdilution assay was used, the higher concentration prepared by dissolving 10 mg of crude in 1 ml DMSO. 1:2 dilutions were prepared and made upto 1 ml using autoclaved MH broth. 2 dilutions 5mg/ml and 2.5mg/ml were showed higher activity than 10mg/ml. 2.5mg/ml showed maximum activity, but further dilutions showed lower activity than 2.5 mg/ml. Can any one help me to explain this with proper reference.
I am not sure why there is higher activity at 2.5 mg/ml. However, I am concerned about the experimental set up itself. I need the protocol to understand better.
1. I think the concentration of DMSO concentration is too much for the cells. What is the final concentration of DMSO in the culture?
2. Do you have a control with only DMSO without the extract?
3. How are you treating the cells with these dilutions? adding cells directly to it?
4. Did you try any other solvent for dissolving the extract?
More information is needed to help you better.
The dilutions were made upto 1 ml using MH broth. concentration of DMSO in 2.5mg/ml sample is 250microliter/ml. this was my first trial for antimicrobial activity. for cytotoxicity assay we followed this dilution methodology. Control prepared using inoculum+mh broth+dmso.
It is 25% DMSO in your broth, which is too much. So DMSO by itself can kill the cells.
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