Foreword
Positional isomers are several compounds that differ only in the position of the functional groups. Separation of these positional isomers is important in many analytical applications, including reaction monitoring, characterization of reactants, and the like. However, since the physical properties between the isomers at different positions are only slightly different, it is difficult to separate them.
Dimethoxybenzoic acid (DMBA) has six positional isomers. If these isomers are separated by the GC method, it is usually necessary to carry out the derivatization reaction before performing the chromatographic separation. This study shows the results of baseline separation of all six DMBA positional isomers within 4.5 minutes. Two-step gradient chromatography can be used to reduce the total separation time. It has been shown that for the separation, it is necessary to add formic acid to the mobile phase.
Figure 1. UPC 2 /UV chromatogram of six dimethoxybenzoic acid positional isomers. The blue number indicates the position of the methoxy group.
Instruments and reagents
System: ACQUITY UPC 2 TM with Photodiode Array (PDA) Detector
Sample: Mixture of six DMBA positional isomers, 0.2 mg/mL isopropanol (IPA) solution for each isomer: ACQUITY UPLC ® BEH125 3.0 x 100 mm, 1.7 micron flow, custom-made Phase A: CO 2
Mobile phase B: Methanol rinse with 0.2% formic acid: IPA
Separation method:
| Time (minutes) | B% |
| 0 | 4 |
| 0.15 | 4 |
| 2.75 | 12 |
| 4.0 | 25 |
| 4.35 | 25 |
| 4.5 | 4 |
Flow rate: 1.2 mL/min
UPC 2 TM Manager: 2175 psi
Column temperature: 60 °C
Injection volume: 1.0 μl Run time: 4.5 minutes Detection wavelength: 240 nm
Data Management: Empower ® 3 Software
in conclusion
Ultra high bonding phase chromatography (UltraPerformance Convergence Chromatography TM) can be carried out without the derivatized samples, six kinds DMBA position isomers were separated quickly.
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