I would like to feed mice or rats a known amount of a supplement containing GABA and or glutamic acid for a given time and afterwards to be able to calculate the absorbed amount by measuring the amount of GABA and/or glutamic acid excreted in their urine. I would appreciate if anyone could provide me a protocol to achieve this. Thanks.
Hi! For glutamate determinations in almost any body fluid, I can definitely recomment the enzymatic Amplex Red Kit provided by Molecular Probes. It is compatibel with multiple assay formats (cuvette/microplates) and can be read out with a colorimeter or a fluorimeter (the latter is more sensitive and can be expected to have a higher dynamic range). Be sure to prepare the calibration curves in the right matrix (urine in your case) and to include blanks. The individual components of the kit are certainly available individually (e.g. Sigma) and may be cheaper as compared to the Molecular Probes Kit.
In cell culture supernatant I was able to reliably detect L-glutamate in the low nanomolar range.
Just genius, thanks for the info. By the way, is there any published reference for that?
Gustavo.
For glutamate, Roche provide a kit, BioPharm L-Glutamic acid enzymatic kit (Cat. no. 10139092035). This can be easily modified to run with your own standards. Haven't used the kit mentioned above so that may be better. As for GABA, the following paper provides a useful enzymatic assay approach: http://www.ncbi.nlm.nih.gov/pubmed/21044648 although it was developed for bacteria, it should analyse GABA un urine just as effectively and allows analysis of lots of samples at once.
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