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A blog on the latest applications, articles, & research on chromatography solutions in sample preparation, Ion Chromatography (IC),
High Pressure Liquid Chromatography (HPLC), Ion Chromatography-Mass Spectrometry (IC-MS), Gas Chromatography (GC),
Gas Chromatography-Mass Spectrometry (GC-MS and GC-MS/MS), and software (Chromeleon CDS, LIMS, and ProteinCenter).

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Phthalates in Culture Media by GC-MS/MS

  
  
  
  
  
  

DEHP, phthalate, GC-MS, ITQ, Thermo ScientificHow many of you have heard of the phthalate contamination in juice that came out last year?  I’m sure there are a lot.  The specific one was DEHP, also known as bis(2-ethylhexyl)phthalate.  Phthalates are industrial chemicals commonly added to plastics, usually to soften texture and improve flexibility.  Because phthalates are widely used, exposure to these compounds is unavoidable.  Reports of it toxicity vary but all seem to say that it is most harmful to male fetus, especially reproductive organ development, they are suspected of being endocrine disruptors.  There also seems to be some reported variance as to the effect on animals in the environment.  Pollution in the environment is not good.  Having these chemicals in the food chain is even worse.  Having them purposely added to juices, well let’s just say I shouldn’t write my thoughts on that.  The problem is not having these compounds, specifically DEHP, in the juice.  The problem is when they get into the body.  Research continues to be done in this area using gas chromatography and GC-MS providing the ability to easily analyze DEHP in serum.

Phthalates are routinely analyzed in electron ionization (EI) with 149 m/z as the principal ion.  Since all phthalates have similar spectra, it is difficult to identify which phthalate is present in a sample.  By using PCI, the molecular ion is formed, which helps classify the phthalate.  Ammonia was selected as the chemical ionization reagent gas to increase the yield of the molecular ion.  By choosing the proper reagent gas, PCI techniques can selectively protonate molecules and provide a high intensity of the quasimolecular ion for the MS/MS process.  Culture media samples with varying concentrations of DEHP were analyzed on an ITQ GC-MS/MS system.  Structural elucidation was achieved by performing MS/MS on the molecular ion.  Mass Frontier software was then used to predict the theoretical fragments and fragmentation pathways of the detected product ion to confirm its identity.

Coupled with MS/MS, the matrix interferences from biological samples were eliminated, enabling quantitation in the low μM levels in culture media. The ability of this method to isolate DEHP from other phthalates makes it very useful to those researching pharmacokinetic effects of this particular phthalate in animal and human studies.  The calibration curve (1, 5, 10 μM) run in culture media and processed in Xcalibur software by external standardization by MS/MS showed linearity of r2 >0.990.  Two unknown samples were analyzed, GCMS #2 and GCMS #3.  Both samples consisted of culture media containing DEHP. The quantitation results were GCMS #2: 5.3 μM (Figure 8) and GCMS #3: 0.3 μM.  Mass Frontier software was able to fully annotate the product ion spectra with fragment ion structures and provide fragmentation pathways for the given molecular structures. Mass Frontier software also proved to be a valuable tool in the confirmation of product ions formed in PCI MS/MS for DEHP.

Download application here; DEHP in Serum

 

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