I am trying to identify the volatile emission spectrum from A. thaliana and L. sativa using SPME of their headspace (static) with GC-MS. I am using an Agilent HP-PONA column, and a GC program with following settings: initial oven temperature is held at 40°C for 5min, then programmed from 40 to 300°C at 5°C/min and finally maintained at 300°C for 5min. The fibers for extraction are a 30/50 CAR/DVB/PDMS and a 65 µm PDMS/ DVB. Headspace is accumulated for 1h and then the fiber is inserted for 1h to equilibrate with the headspace. The fibers are desorbed immediately after sampling. The spectra are not satisfying for A. thaliana, yielding few peaks that are poorly identifyable. For L. sativa, I get nice, sharp peaks but I do not get the compounds I am expecting when applying stress. I would be gratefull if anyone could comment on improvement of the method/sampling procedure,etc.
Belza, few questions: (1) what is the temperature of the sample vials while equilibrating/extraction ? (2) are you agitating the sample vial?
While having thicker fibers, sometimes you need longer times to extract adequate amounts of components. Temperature and agitation can help improve the extraction. Also, the lab
In addition, to what Noel said, do you have standards for the compounds you do expect to see? Perhaps testing your fibers on standards would be advisable.
Also, what inlet temperature and desorption time are you using?
Thanks! I have different cuvettes with different headspace volumes. I would really like to get a full scan to identify and validate compounds, SIM monitoring will be the next step, but it is really about identifying the compounds first. The inlet temperature is 270°C and the desorption time is around 5-10min. Could it be the column that is rather incoventional for HS-GC-MS of plant volatiles?
Belza, inject in splitless or split mode? If you have low peaks and you don't want to change your extraction parameters you have got to try to inject into splitless mode. Which is the injector temperature?
PONA is a very non-polar phase. Typically I want to concentrate the volatiles on the head of the column in order to get the best chromatography. This normally means that I need to use a relatively polar phase, such as a 624-type phase (medium polarity, slightly more polar than a -17). You're using a relatively polar SPME fiber, so you would want to match that in your column, since you need to effect a transfer of the analytes from the fiber to the column with a minimum of band broadening.
Your temperature profile is very slow. Capillary columns actually separate better with faster temperature profiles, not slower. Given the non-polar nature of this column I would expect you to have peak shape issues with the front end of your chromatography.
That said, I think that 40C is relatively high as a starting temperature. I realize it is hard to start lower without cryofocusing, but if the compounds you are looking for are truly volatile (say less than n-C6) then you are probably losing them by starting at 40.
Can you use other sampling techniques like thermal desorbtion tubes?
This would increase the amount of VOC's sampled making it much easier to identify your compounds.
What program are you using to identify your compounds? I find AMDIS a great programs to identify even the smallest peaks that you won't consider looking at when identifying peaks manually.
Regards, Alex
Did you got solution for your question and what you have done to obtain your expected results? I am also face similar kind of issue.
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