Methods to be Reviewed by OMB in 2016

(a)

Workflow and Detection / Identification Criteria.

—Detection and identification of analytes is

performed with the use of TraceFinder software and settings provided below. Detection of

targeted PDE5 inhibitors is based on automatic comparison of retention times of peaks

extracted from full MS record and accurate mass of respective pseudomolecular ion [M+H]

with

information in TraceFinder compound database (see Table

2015.12D

). Retention time and mass

tolerances of 30 s and 5 ppm, respectively, are used. To identify an analyte, additional criteria

have to be fulfilled. These include mass accuracy (∆

m/z

≤ 5 ppm) and relative responses (10%

tolerance) of pseudomolecular ion isotopes, as well as criteria for fragment ions detected in

appropriate dd-MS

records. For positive identification, one or more fragment ions listed in the

TraceFinder compound database have to be detected above the intensity threshold with a mass

error ≤ 5 ppm. The detection / identification workflow for targeted compounds is provided in

Figure

2015.12

. PDE5 inhibitors not included in the TraceFinder compound database can be

detected and identified by extraction of respective pseudomolecular ions from full MS records

and evaluation of fragment ions in AIF records. A search using common PDE5 inhibitor

fragments can be utilized to highlight components with structures similar to known PDE5

inhibitors.

(b)

TraceFinder Software Settings.

1. Retention time range:

1-23 min

2. Peak area threshold:

100,000

3. Signal-to-noise threshold:

4. Mass tolerance (parent ion):

5 ppm

5. RT tolerance:

30 s

6. Minimum # of fragments:

7. Intensity threshold:

1,000

8. Mass tolerance (fragment ion):

5 ppm

9. Isotope pattern fit threshold:

95%

10. Mass tolerance (isotope):

5 ppm

11. Intensity tolerance (isotope):

10%

(c)

TraceFinder Compound Database.

–Compound database (see Table

2015.12D

) comprises

information on the exact mass of pseudomolecular ions, molecular formulas, retention times

and exact masses of 8-10 fragment ions for each analyte. The

m/z

values of fragments in the

compound database represent exact masses that were calculated using experimental data

obtained by HRMS analysis of reference standards and elucidation of fragment ions in Mass

Frontier spectral interpretation software or based on information available in m/zCloud

database and scientific literature.

Reference:

J. AOAC Int

. (future issue)

AOAC SMPR 2014.010

J. AOAC Int

, 1060(2015)

Posted: August 2015

Candidates for 2016 Method of the Year

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