Frequency division multiplex HPLC-MS for simultaneous analyses
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1 Electronic Supplementary Material (ESI) for Analyst. This journal is The Royal Society of Chemistry 9 Supporting Information Frequency division multiplex HPLC-MS for simultaneous analyses Hiroka Kishi, Takashi Kumasaki, Shinya Kitagawa*, Hajime Ohtani Department of Life Science and Applied Chemistry, Graduate School of Engineering, Nagoya Institute of Technology, Gokiso, Showa, Nagoya , Japan Content: Figure S: Additional figure for Figure as a theoretical study of multiplex HPLC-MS based on FDM. Figure S: Detail of signal processing. Figure S3: Evaluation of a cut-off ratio using an integration curve of a power spectrum. Figure S4: Mechanism of interference used in two chopper type modulators. Figure S5: Chromatogram obtained by HPLC-MS with modulation via an ion-gate type modulator. Figure S6: Simultaneous analysis of mixtures of () -methylxanthine and theophylline, and () theophylline and caffeine in LC-MS
2 A(ⅰ) A(ⅱ) A(ⅲ) B(ⅰ) f=. Figure S. (A) Virtual chromatogram without modulation (i), its power spectrum obtained by FT (ii), and the restored-chromatogram for low region. (B) Virtual chromatogram with different modulations (. and 6.3 Hz) (i), its power spectrum obtained by FT (ii), and its restored-chromatogram by RFT for low (iii),. Hz (iv), and 6.3 Hz (v) regions. The figures with the exception of (A-iii) and (B-iii) are shown in Figure. f= FT FT B(ⅱ) low freq (/σ) low freq (/σ) B(ⅲ) B(ⅳ) B(ⅴ)
3 A(ⅰ) A(ⅱ) h ( ).8 8 h ( ) B(ⅰ) B(ⅱ) B(ⅲ) h ( ) ( ) 4 4 h ( ) 3 h ( + ) h ( ) 4 h ( ) Shift to zero Figure S. (A) Chromatogram without modulation (i) and its power spectrum with both plus and minus regions (ii). In the text, only the pulse region (in the orange box) is presented. (B) Chromatogram with modulation (i) and its power spectrum with both plus and minus regions (ii). The blue box in (B-ii) denotes the signal for extraction. Before RFT, the signal is shifted to f = region (B-iii). The following equations describe a mathematical detail of the signal processing with a typical modulation function of Mod(t). h ( ) = + ( cos( ) + sin( )) h ( ) h ( ) ( ) = ( + cos( )) h ( ) ( ) = h ( ) + 4 cos( ) + ( cos( ) + sin ) cos( ) = h ( ) + 4 cos( ) + ( cos( ) cos( ) + sin( ) cos( )) = h ( ) + 4 cos( ) + cos( ) + cos( + ) + cos( ) sin( + ) + sin( ) + = h ( ) + 4 cos( ) + ( cos( + ) + sin( + )) + cos( ) + ( cos( ) + sin( )) h ( ) + 4 h ( + ) + h ( ) 4 h ( + ) h ( ) h ( )
4 .5 Ideal modulation Non-ideal modulation % % Relative integration [%] Relative integration [%] Cut-off ratio = 5% Signal compensation : 4 s Cut-off ratio = 4% Signal compensation: 5 s Figure S3. Evaluation of a cut-off ratio using integration curve of the power spectrum to compensate the signal of a restored chromatogram.
5 (ⅰ) Theobromine chopper Y shaped interface (ⅱ) 5 Caffeine Theobromine 8 Hz Caffeine.7 Hz (min) (iii) P A Pc (iⅴ) P B Pc M MS P ID.5 mm, long 3 cm ID.5 mm, long 8.5 cm P P < Pc Figure S4. Mechanism of interference used in two chopper type modulators. (i) apparatus used for the experiment (continuous injection mode without separation column), (ii) signals are recorded in the same phase, (iii) both gas stream are not blocked by the choppers and both analytes were detected, (iv) lower (green) gas stream (and analyte stream) was blocked by the chopper. Since P is less than P c, the buck flush of the analytes (denoted with blue) suppressed the signal of the blue analytes observed in MS.
6 (ⅰ) 8.E+5 6.E+5 4.E+5.E+5 (ⅲ) 8.E+5 6.E+5 4.E+5 5 (min) (ⅱ) 5.E+7 4.E+7 3.E+7.E+7.E+7 (ⅳ) 8.E+4 6.E+4 4.E+4 low freq..5 Hz.5 (Hz) the chromatogram without modulation SD=49.E+5 5 (min).e+4.e+ restored chromatogram SD=576 4 Time(min) Figure S5. Chromatogram obtained by HPLC-MS with modulation via an ion-gate type modulator (.5 Hz) (i), its power spectrum obtained by FT (ii), and the restored-chromatogram by RFT for the.5 Hz region (iii).
7 (ⅰ) -methylxanthine + theophylline LC 8 Hz switcha switch B Power supply.7 Hz theophylline + caffeine LC Signal compensation: 6.6 (ⅳ).E+5.6E+5 (ⅱ).E+5 (ⅲ) low freq. 8 Hz 3.E+6 3.E+6.7 Hz.E+5 8.E+4.6E+5.E+5 8.E+4 4.E E+6.5E+6.E+6.E+6.5E+6.5E+6.E+6.E+6 5.E+5 5.E+5.E+.5 (Hz) (Hz) (ⅴ) 4.E+4.E+5.6E+5.E (min) 8.E+4 Signal compensation: E+4 Cut-off ratio: 4% Cut-off ratio: % 5 5 (min) Figure S6. Simultaneous analysis of mixtures of () -methylxanthine and theophylline, and () theophylline and caffeine in LC-MS. (i) schematic diagram of the experiment, (ii) a mixed-chromatogram, (iii) a power spectrum of (ii), (iv) a restored-chromatogram for.7 Hz region with signal compensation of 6.6 s, and restored-chromatogram for 8 Hz with signal compensation of 8.3 s.
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