there are many possibilities:

for the volatiles I would suggest Fustinoni et al. methodology 1999 for the sample collection procedure ( https://www.researchgate.net/publication/13209697_Headspace_solid-phase_microextraction_for_the_determination_of_benzene_toluene_ethylbenzene_and_xylenes_in_urine )

for the non volatiles there is a very nice paper from Saude and Sykes 2007 on different methodologies for preserving urine samples ( https://www.researchgate.net/publication/225406929_Urine_stability_for_metabolomic_studies_effects_of_preparation_and_storage ) it appears that the less invasive methodology for preservation is centrifugation and storage in -80 freezers, adding NaN3 as well is a good measure for preservation and it is standard for NMR metabolomics processing.

personally I only centrifugate and store in -80, except with the volatiles, since the centrifugation, handling and the thawing process will lead to loss of the volatile metabolites (BTEX mixture mainly for my analyses).

hope this helps, if you need more information don't hesitate to contact me.

cheers,

Max

Article Headspace solid-phase microextraction for the determination ...

Article Urine stability for metabolomic studies: Effects of preparat...

Hi Khushman

I would agree with Max, in urine there are two significant processes that must be stopped. One is direct bacterial degradation of the metabolites, the other is enzymatic hydrolysis. NaN3 is a good way of preventing bacterial degradation, although -80 C should be sufficient for short to moderate storage. -80 C should also slow enzymatic degradation to a minimum, though if you have certain metabolites in mind you may be able to chemically inhibit the specific enzymes with a careful literature search.

I would agree with first morning urine, midstream,centrifugation 5 min, 24oC, 400 g) was stored at -80oC until analysis.

maintain the pH of sample to 2 for storage

One has to be careful with acidification of the sample. It certainly helps preservation (Hellstrom et al 1999) but strongly acidic (or alkaline) conditions can alter functional groups of metabolites. Urine is a rather complex matrix so you need to consider carefully the metabolites of interest then decide if acidification is suitable.

Is useful the use of 0,01 % NaN3 to prevent bacterial and micotic growth, than to store the samples at least - 40°C

for metal estimation urine sample may be stored at pH 2

thank you all for your help and suggestions, i would like to add up to this topic that my approach of study is the whole metabolome in urine as such and not a specific group of metabolites. Its basically a non targeted approach and not a targeted one.

Further suggestions are welcome and i would like to get your help in future also.

thanks,

Khushman Taunk.

sounds very similar to what I am doing (whole metabolome in urine and nontargeted approach), I suggest to start with the papers I cited and then evaluate the possibilities, Saude and Sykes' paper explores all the main options for long term storage.

thinking about metabolomic analyses also flash-freezing the aliqotes samples in liquid nitrogen could be a good option, but it can become tricky and not feasible both for storage (limited capacity of liquid nitrogen containers/availability of such freezers and containers).

you may want to consider the methodologies you are going to use to analyse your samples to refine the storage methodology that you will use (i.e. for NMR analyses adding NaN3 to the sample is the norm, while it might produce an interference with MS).

Max, my methodology for analyses of the metabolites is the LCMS and GCMS as of now. I am also planning to go for NMR but it would take some time and my guides permissions also.

It is very interesting to know that u r also working in the similar approach as of mine. this would really be helpful for me at least as u have been working in this field already for some time now. Plz suggest some good papers for the extraction of metabolites from the urine samples. What protocols do u use in ur lab for this?

Thanks........

I tend to avoid as much as I can sample preparation, for GCMS I use SPE columns or extract using SPME in the headspace, depending on what you are analysing you choose the solid phase of the SPE accordingly. there are also liquid extraction protocols, but I don't really like them due to the uncertainty of the recovery and efficiency.

at the moment I can't provide papers (I'm not using my PC), but you can easily find loads of papers searchig on isiknowledge or pubmed with the keywords metabolomics, metabonomics, AND urine, extraction, SPE and metabolites.

see which kind of extraction is more suitable for you (you may want to consider a nonpolar extraction followed by a polar extraction to maximise the number of metabolites you extract) and follow the papers accordingly ;)

good luck! and let me know if you need more help, I'll try to provide better advice once I have my computer, but you can start from here easily.

cheers,

Max

thanks Max,,,, i just saw ur msg nw. i will get in touch with u. may i have ur email id please?

[email protected] don't know if it will show

First of all for the storage of urin you just centrifuge it and you may concentrated either by dialysis sac or amnicon concentrated and freez it.For separation of any especial protein you use either salting out method or using chromatography.good luck

thanks ALI MOSHTAGHIE Sir....... :)

There are collection and preservation tubes available in the market.  Some will prevent bacterial growth while preserving proteins (and nucleic acids) at ambient temperature for a few years.  Here is a paper where using such tubes: 

http://bit.ly/1U4o686

Total proteins can also be concentrated using spin column methods.  

Hope this information is helpful.