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Environmental Article

Fast, Sensitive LC/MS/MS Analysis of Paraquat and Diquat

Using an API 3200™ Mass Spectrometer and an Ultra Quat HPLC Column

by Houssain El Aribi, Ph.D., LC/MS Product and Application Specialist, MDS SCIEX*, Becky Wittrig, Ph.D., HPLC Product Manager, C. Vernon Bartlett, HPLC R&D Scientist, and Julie Kowalski, Innovations Team Chemist, Restek Corporation
  • Complete resolution of paraquat & diquat — with a simple, isocratic mobile phase!
  • Superior sensitivity—5ppb paraquat or 0.1ppb diquat—without preconcentration.
  • Significantly faster than conventional methodologies.

Restek chemists designed the Ultra Quat HPLC column specifically for analyses of quaternary amine compounds. This unique column makes possible a simple HPLC/UV analysis for paraquat and diquat1 — a significant improvement over alternative methodologies. Now, in collaboration with scientists at MDS Sciex, we have developed a fast, highly sensitive LC/MS method for analyzing these challenging target compounds.

Charged quaternary amines, such as paraquat and diquat, exhibit little or no retention on C18 or other alkyl stationary phases. In our HPLC/UV procedure, our Ultra Quat mobile phase modifier (Ultra Quat Reagent Solution, cat.# 32441) increases the interactions between paraquat and diquat and the Ultra Quat stationary phase, providing the necessary retention and resolution. For compatibility with MS detection, however, we needed a volatile mobile phase additive. Low concentrations of heptafluorobutyric acid (HFBA) effectively shield the positive charges of paraquat and diquat, increasing interactions between the quaternary amines and the Ultra Quat stationary phase.

Figure 1 shows the excellent separation of paraquat and diquat, at a concentration of 5µg/mL each in water, achieved by using an API 3200™ mass spectrometer. We used multiple reaction monitoring (MRM) — a standard technique for quantitative LC/MS/MS — for this application. In MRM, pairs of target precursor ions and unique fragment ions are used for quick and accurate identification of target species. Collision induced dissociation (CID) is used to generate the fragment ions. CID spectra for paraquat and diquat are shown in Figures 2 and 3. This approach has been used in many pharmaceutical and environmental applications, to generate unmatched limits of detection or quantification, precision, and accuracy. For accurate quantification, we used paraquat-d8 and diquat-d4 as internal standards (Table 1), to compensate for matrix effects and to correct for random and systematic errors in separation and detection.

For triplicate injections of 8 concentrations of analytes in deionized water and in lake water, from 5µg/100mL to 100µg/100mL for paraquat and from 0.1µg/100mL to 100µg/100mL for diquat, correlation coefficients for calibration curves were >0.995, using a linear fit and 1/x weighting factor. These results indicate that quantification can be performed with good linearity and sensitivity. Minimum detection limits (MDL) for the method, for paraquat and diquat in deionized water, were 5µg/L and 0.1µg/L, respectively.

LC/MS is a powerful tool for analyses of challenging environmental contaminants. In LC/MS analyses of paraquat and diquat, the combination of an Applied Biosystems API 3200™ mass spectrometer and an Ultra Quat HPLC column ensures fast, sensitive, and accurate results.

Figure 1  Fast, sensitive LC/MS/MS analysis of paraquat and diquat, using an API 3200™ mass spectrometer and an Ultra Quat HPLC column.

Paraquat and diquat herbicides

  1. diquat
  2. paraquat

Sample:

 

Inj.:

10µL

Conc.:

5µg/mL each component

Sample diluent:

DI water

Sample temp.:

ambient

Column:

Ultra Quat (cat.# 5181352)

Dimensions:

50 x 2.1 mm

Particle size:

3µm

Pore size:

100Å

Conditions:

 

Mobile phase:

10mM heptafluorobutyric acid:acetonitrile, 95:5 (v/v)

Flow:

0.3mL/min.

Temp.:

ambient

Det.:

Applied Biosystems API 3200™

Interface:

electrospray

Ion Mode:

positive

Temp.:

600°C

Ion Spray™ voltage:

5500V

Collision exit potential:

3V

Curtain Gas™:

15psi

Gas supply 1:

70psi

Gas supply 2::

60psi

Quantitation:

(MRM)

Q1/Q3:

unit resolution

Dwell time:

200 ms

Precursor Ion (amu)

Fragment Ion (amu)

DP (V)

Collision Energy (eV)

diquat, 183+

157+

30

30

paraquat, 93 (2+)

171+

20

20

*Data courtesy of Houssain El Aribi, Ph.D., LC/MS Product and Application Specialist, MDS SCIEX, 71 Four Valley Drive, Concord, Ontario, Canada, L4K 4V8



Table 1  MRM transitions and MS conditions used to generate CID spectra for paraquat and diquat.
Precursor
Ion (m/z)
Fragment
Ions (m/z)
DP (V)
Collision
Energy (eV)
Paraquat [M2+ - H+]
    185
170a
169b
40
30
Paraquat-d8 [M2+ - D+]
    193
178a
40
30
Diquat [M2+ - H+]
    183
157a
168b
35
30
Diquat-d4 [M2+ - H+]
    186
158a
35
30


Figure 2  CID spectra for paraquat+ at CE = 25eV.


Figure 3  CID spectra for diquat+ at CE = 25eV.

References

  1. Simple, Sensitive HPLC/UV Analysis for Paraquat and Diquat, Using High-Recovery Solid Phase Extraction and an Ultra Quat HPLC Column Applications Note 580006, Restek Corporation, Feb. 2006.