What should be the concentration of isolated compound in terms of µM to consider this as a hit in an antimicrobial screening microdilution assay?
And what should be the concentration of this compound (µM) to begin in vivo assays?
Dear Roman, 10 ug/mL is recommended, FDA recommend 1 ug/mL. Professor Paul Cos use this concentration due to toxicity risk. For initiate in vivo test is necessary to determinate in vivo pharmacokinetics also index selection criteria, comparing activity vs toxicity. With best regards
To begin with, the measurement of the minimal concentration of a substance that completely inhibits growth of the culture is called the MIC (minimal inhibitory concentration). It is normally expressed in µg/ml or, equivalently, mg/L rather than µM. I think the reason for this is that in complex mixtures such as natural product extracts, it is not possible to define a molecular weight with which to calculate molarity. For pure substances, you could use µM, but it is not done. MICs are always given as µg/ml or mg/L. However, if you are screening a compound library in which the compound concentrations are supplied in molarity, then you can use µM for the time being.
Back to the real question: what is the potency cutoff for an active compound? There is no specific value. It is up to you to decide what level of potency you consider worth following up. If you are screening a compound library and the test concentration is, for example, 10 µM, then you would probably set that as the cutoff: if you see growth inhibition at the test concentration, then consider it a positive. If you are looking for compounds that inhibit the growth of Gram negative bacteria, you should probably test at a higher concentration, such as 100 µM, because it is usually difficult to get compounds into Gram-negative bacteria.
Another consideration besides potency is selectivity. It is not hard to find compounds that prevent growth of bacteria, but it can be much more difficult to find compounds that do not also prevent the growth of eukaryotic cells. Be prepared to run a counterscreen for inhibition of a mammalian cell culture by the positive compounds from your antibacterial screen. Because antibacterial compound doses in vivo are generally quite large, resulting in high plasma concentrations, any tendency of an antibacterial compound to harm human cells in culture is a "red flag" for toxicity.
Thanks Adam. My question was focused on pure compounds (isolated and identified) because some of them can be molecular weights higher than others. Then the net weight in some kind of compounds will represent more or less quantity of molecules . This fact could be represent an advantage for small molecules (in term of number of molecules reacting with target)?
Dear Roman, 10 ug/mL is recommended, FDA recommend 1 ug/mL. Professor Paul Cos use this concentration due to toxicity risk. For initiate in vivo test is necessary to determinate in vivo pharmacokinetics also index selection criteria, comparing activity vs toxicity. With best regards
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