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Silliker
Chem, Res. Center Crete, IL – Report of A Validation of LC-MS/MS Method for Folate Analysis
24
stability but that required to use 90% methanol instead of 20% methanol. This experiment was
performed to make sure higher methanol does not affect the analytical response of the analytes . It has
established earlier that injections of analytes in 90% methanol provides a satisfactory chromatography.
The response of analytes at 5 ppb and 500 ppb level was investigated with injection solvent low in
methanol (20%) and high in methanol (90%). The results are presented in Table 10. It can be seen that
higher methanol level provided a superior response in most cases or equivalent response. Thus the
standard solutions and extract for the analysis can be prepared in solutions containing 90% methanol
without any loss in the analytical response.
Table 16 : Comparison of response (peak area) of folate compound at two different concentrations
in different injection solvents differing in %MeOH
STD
Injection Solvent*
Std
Conc
ng/mL
FA
5-CH3-
THF
5-CHO-THF
THF
10-CH3-F
90% MeOH
10:90:1, Formic, MeOH,
Ascorbic (v/v/Wt.)
5
2.91E+03
9.28E+03
6.75E+03
2.19E+03
5.18E+03
20% MeOH
80:20, 1%Acetic,MeOH,
Ascorbic (v/v/Wt.)
5
2.18E+03
5.97E+03
5.77E+03
2.34E+03
2.45E+03
90% MeOH
10:90:1, Formic, MeOH,
Ascorbic (v/v/Wt.)
500
3.34E+05
9.77E+05
7.09E+05
3.62E+05
5.54E+05
20% MeOH
80:20, 1%Acetic,MeOH,
Ascorbic (v/v/Wt.)
500
1.78E+05
7.56E+05
5.98E+05
2.94E+05
3.06E+05
*Injection Solvents: with 90% methanol is similar to the elution solvent of SPE; with 20% methanol some what similar to
the mobile phase
Evaluation of Solvent Blanks:
The solvent blank (methanol) did not provide any interference in the analysis of the targeted analytes
(Figure 5).
2011.06 (Fol-22) w/SLV
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