Chromatography Forum

Does somebody knows if relative intensity included in table 4 of the Commission Decision 2002/657 also applys to HPLC-MS/MS method which use for the identification the SRM or MRM mode? If I have to apply this rule also for SRM transions could somebody explain to me how to calcualte the relative intensity of the transitions. I have change the settings of the chromatograms and the % of the peak is always 100 % as it is the most intense peak of the transition, is this aceptable?? I don’t know, I am not very sure.

Thanks in advance for the help,

Maximum permitted tolerances for relative ion intensities using a range of mass spectrometric
techniques

Relative intensity ______________ EI-GC-MS _______________ CI-GC-MS, GC-MSn, LC-MS, LC-MSn
(% of base peak) ________________(relative) _________________ (relative)
> 50 % ________________________ ± 10 % __________________ ± 20 %
> 20 %to 50 % __________________± 15 % __________________ ± 25 %
> 10 %to 20 % __________________± 20 % __________________ ± 30 %
≤ 10 % ________________________ ± 50 % __________________ ± 50 %

From the criteria shown as above you can use both SRM or MRM mode and run the sample compare with standard or RM.
For example, you do MS/MS with MRM mode that provided you the following.

RM or Standard
m/z 290 ints. 25200 (molecular ion, 21% Relative to basepeak)
m/z 168 ints. 120000 (100% as basepeak)
m/z 105 ints. 9600 (8% compared to basepeak)

For sample
m/z 290 ints. 11550 (molecular ion, 23%Accept range 21± 25 %, 15.75 - 26.25%)__PASS
m/z 168 ints. 55000 (100% as basepeak)_______________________________PASS
m/z 105 ints. 6050 (11% Accept range 8±50 %, 4 - 12%)___________________PASS

Jetjamnong
Best wishes

@Carol:
send me your e-mail on jedzi(at)piwet.pulawy.pl. I will send you excel sheet for proper calculation. We used it routinely in the confirmation of vet drug residues in animal tissues.

Hi,

Thanks for you quick replay but I think you don't understand me.
If I do MRM, the relative intensity of my peak is always 100 % so I think I am doing something wrong. I don't know which transition should I considered as base peak....

I don't know if I have clarified my problem.

Thanks again in advance,

Yes, you are doing something wrong. Your chromatographic peak is (or should be) the response from a single ion. You are monitoring multiple ions for each compound. If you monitor three ions for a compound, you can produce three chromatograms, with three integrated peak areas, all at the same retention time. The most intense signal is from the base peak (this peak refers to the mass spectrum, not the chromatogram). The relative abundance is the ratio of the peak areas from the other ion chromatograms to this peak's area. Monitoring three ions will give you two ratios to use for relative abundance matching.

Your peak is 100% absolutely it's called basepeak and you can select this ion as Quntify ion (most intensity). therefore, you need for a minimum 2 m/z as Qulifier ion to confirm that compound and calculate compare to the basepeak (100%) then it provide you relative value.

Jetjamnong