What is the storage temperature for neurotransmitter estimation in HPLC? I freshly prepared and loaded it, it has separated very nicely, but the same sample I stored at -20C for six hours after I loaded it didn't separate as before. I don't know what is the reason behind this.
Maybe the sample got thawed. Should I wait until it reaches the room temperature after that only should I load in injector? As soon as sample gets thawed, should I inject? Or should I not wait until the sample comes to room temperature, as it may affect separation in the column?
I hope, I am not wrong. You should wait until the temperatue of frozen sample equals to that ambvient. Mix the sample in your test tube and try to inject it in the injector of your instrument.
In the case of injecting the liquid taken from the tawed part of the sample two errors rmight occur. The first is lover temperature of the sample injected compared to that injected without storing at subzero temperatures /I mean the fresh solution/.
The second error may dwell in crystalization or precipitation of some neurotransmitters at -20 oC. When the frozen sample is taken out from the freezer and let to thaw, the more soluble components are more abundant in the liquid phase, and less soluble ones are concentrated prevailingly in the solid phase /ice/. I guess this idea might be correct.
Recomendation - 1. dont keep your neurotransmitter at such a low temperature.
2. If a small, e.g. 10% addition of ethanol to the sample does not inteferre with the analyt, and does not influence the separation, add it /always the same volume/. Then you may keep the sample at subzero temperatures. I think, - 10 oC and presence of N2 over the sample can help to store the samples for long time.
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