Hydrogen Sulfide Deactivates Common Nitrobenzofurazan-Based ...
Supporting Information Hydrogen Sulfide Deactivates Common Nitrobenzofurazan-Based Fluorescent Thiol Labeling Reagents Leticia A. Montoya and Michael D. Pluth*
Department of Chemistry and Biochemistry Institute of Molecular Biology Materials Science Institute University of Oregon Eugene, OR 97403-1253
Contact Information: Michael D. Pluth [email protected] Fax # (541) 346-4643
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Table of Contents Page
1. Synthesis of 6 2. 1H and 13C{1H} NMR spectra of 6 3. Full 1H NMR spectra of 6 before/after addition of H2S 4. Full 13C{1H} NMR spectra of 6 before/after addition of H2S 5. UV-vis spectra of 1 and NBD-SH
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Synthesis 7-Hydroxy-4-methyl-2-oxo-2H-chromene-8-carbaldehyde (6). 7-Hydroxy-4-methyl-2Hchromen-2-one (1.5 g, 8.5 mmol) was dissolved in CH2Cl2 (25 mL) and the solution was cooled to 0 °C. Acetic anhydride (1.2 mL) and pyridine (0.76 mL) were added drop-wise to the solution. The reaction mixture was then slowly warmed to room temperature and stirred overnight while protected from light. The crude product was diluted with ~150 mL EtOAc and washed with 1 M HCl (2 x 100 mL). The organic layers were combined washed with K2CO3 (3 x 100 mL) and dried over MgSO4. The EtOAc was removed under vacuum to provide a white flaky solid (1.43 g, 77% yield). 1H NMR (500 MHz, CDCl3) δ: 7.62 (d, J = 8.6 Hz, 1H), 7.13 (d, J = 2.2 Hz 1H), 7.07 (d, J = 2.29 Hz, 1H), 6.28 (d, J = 1.31 Hz, 1H), 2.45 (s, 3H, Me), 2.35 (s, 3H, Me). The crude product (100 mg, 0.458 mmol) was dissolved in TFA (620 μL) and cooled to 0 °C. Hexamethylenetramine (96.4 mg, 0.687 mmol) was added then added slowly in small increments. The reaction mixture was warmed to room temperature and then heated to 75 °C overnight while protected from light. The TFA was then removed under vacuum and the remaining solution was cooled to room temperature.
Addition of H2O and cooling to 0 °C
precipitated the product, which was isolated by filtration. The product was washed with cold H2O and dried under vacuum to provide the desired product as an orange/brown solid (17.6 mg, 19% yield). 1H NMR (600 MHz, CDCl3) δ: 12.24 (s, 1H, OH), 10.65 (s, 1H, aldehyde H), 7.75 (d, J = 8.9 Hz, 1H, ArH), 6.93 (d, J = 8.9 Hz, 1H, ArH), 6.23 (s, 1H, ArH), 2.45 (s, 3H, Me).
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C{1H}c
NMR (125 MHz, CHCl3) δ: 193.44, 166.29, 159.23, 156.18, 152.64, 132.90, 114.30, 112.10, 108.70, 18.98.
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NMR Spectra
Figure S1. 1H (600 MHz, CDCl3) and 13C{1H} (150 MHz, CDCl3) NMR spectra of 6.
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Full 1H NMR spectra of 6 before/after addition of H2S
Figure S2. 1H NMR spectra of 8 after addition of H2S. (a) 6 and 10 equiv. H2S in D2O with HEPES buffer (pH=7.4). Peaks below 5.0 ppm are from HEPES (b) 6.
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Full 13C NMR spectra of 6 before/after addition of H2S
Figure S3. 13C{1H} NMR spectra of 6 before (b) and after (a) addition of 10 equiv. of H2S. Conditions: D2O with HEPES buffer (pH=7.4). The peaks between 45 and 60 ppm are from the HEPES buffer, and minor DMF impurities are present at 164, 37, and 32 ppm.
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UV-vis spectra of 1 and NBD-SH
Figure S4. UV-vis spectra of NBD-Cl and NBD-SH.
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Department of Chemistry and Biochemistry Institute of Molecular Biology Materials Science Institute University of Oregon Eugene, OR 97403-1253
Contact Information: Michael D. Pluth [email protected] Fax # (541) 346-4643
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Table of Contents Page
1. Synthesis of 6 2. 1H and 13C{1H} NMR spectra of 6 3. Full 1H NMR spectra of 6 before/after addition of H2S 4. Full 13C{1H} NMR spectra of 6 before/after addition of H2S 5. UV-vis spectra of 1 and NBD-SH
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Synthesis 7-Hydroxy-4-methyl-2-oxo-2H-chromene-8-carbaldehyde (6). 7-Hydroxy-4-methyl-2Hchromen-2-one (1.5 g, 8.5 mmol) was dissolved in CH2Cl2 (25 mL) and the solution was cooled to 0 °C. Acetic anhydride (1.2 mL) and pyridine (0.76 mL) were added drop-wise to the solution. The reaction mixture was then slowly warmed to room temperature and stirred overnight while protected from light. The crude product was diluted with ~150 mL EtOAc and washed with 1 M HCl (2 x 100 mL). The organic layers were combined washed with K2CO3 (3 x 100 mL) and dried over MgSO4. The EtOAc was removed under vacuum to provide a white flaky solid (1.43 g, 77% yield). 1H NMR (500 MHz, CDCl3) δ: 7.62 (d, J = 8.6 Hz, 1H), 7.13 (d, J = 2.2 Hz 1H), 7.07 (d, J = 2.29 Hz, 1H), 6.28 (d, J = 1.31 Hz, 1H), 2.45 (s, 3H, Me), 2.35 (s, 3H, Me). The crude product (100 mg, 0.458 mmol) was dissolved in TFA (620 μL) and cooled to 0 °C. Hexamethylenetramine (96.4 mg, 0.687 mmol) was added then added slowly in small increments. The reaction mixture was warmed to room temperature and then heated to 75 °C overnight while protected from light. The TFA was then removed under vacuum and the remaining solution was cooled to room temperature.
Addition of H2O and cooling to 0 °C
precipitated the product, which was isolated by filtration. The product was washed with cold H2O and dried under vacuum to provide the desired product as an orange/brown solid (17.6 mg, 19% yield). 1H NMR (600 MHz, CDCl3) δ: 12.24 (s, 1H, OH), 10.65 (s, 1H, aldehyde H), 7.75 (d, J = 8.9 Hz, 1H, ArH), 6.93 (d, J = 8.9 Hz, 1H, ArH), 6.23 (s, 1H, ArH), 2.45 (s, 3H, Me).
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C{1H}c
NMR (125 MHz, CHCl3) δ: 193.44, 166.29, 159.23, 156.18, 152.64, 132.90, 114.30, 112.10, 108.70, 18.98.
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NMR Spectra
Figure S1. 1H (600 MHz, CDCl3) and 13C{1H} (150 MHz, CDCl3) NMR spectra of 6.
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Full 1H NMR spectra of 6 before/after addition of H2S
Figure S2. 1H NMR spectra of 8 after addition of H2S. (a) 6 and 10 equiv. H2S in D2O with HEPES buffer (pH=7.4). Peaks below 5.0 ppm are from HEPES (b) 6.
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Full 13C NMR spectra of 6 before/after addition of H2S
Figure S3. 13C{1H} NMR spectra of 6 before (b) and after (a) addition of 10 equiv. of H2S. Conditions: D2O with HEPES buffer (pH=7.4). The peaks between 45 and 60 ppm are from the HEPES buffer, and minor DMF impurities are present at 164, 37, and 32 ppm.
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UV-vis spectra of 1 and NBD-SH
Figure S4. UV-vis spectra of NBD-Cl and NBD-SH.
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