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Additional file 5: Figure S5. Single cell AlexaFluorâ ˘ 647 intensities of A. S. cerevisiae B. P. chrysosporium V. E. coli, and D. E. lignolyticus. Thin red line is the mean and red whiskers mark the standard deviation. Intensities shown are from the cells deemed positive in three biological replicates based on stringent thresholds to minimize the moderate levels of autofluorescence (see â Materials and Methodsâ for detailed description). Mean intensity data is presented raw and does not compensate for the vanillic acid analog being difunctionalized.

Additional file 3: Figure S3. A monofunctionalized vanillic acid analog, where the methoxy group in vanillic acid was replaced by the propynyloxy group and the p-hydroxy remained unfunctionalized, (Fig. S3, box) would perhaps be a better vanillic acid mimic by preserving the hydrogen bonding of the p-hydroxy.

Additional file 7: Figure S6. A. ASAP-MS representative spectra of S. cerevisiae lysates incubated with vanillic acid for 1, 2, 3, or 17 h. Blue arrow indicates integrated peak of interest for results displayed in Fig. 4c. B. ASAP-MS representative chromatogram of P. chrysosporium lysates incubated with vanillic acid for 1, 2, 4, or 24 h. Orange arrow indicates integrated peak of interest for results displayed in Fig. 4b. C. ASAP-MS representative chromatogram of P. chrysosporium lysates incubated with 13C labeled vanillic acid for 1, 2, 4, or 24 h. Red arrow indicates integrated peak of interest for results displayed in Fig. 4b. D. ASAP-MS representative chromatogram of P. chrysosporium lysates incubated with vanillic acid for 5, 15, and 60 min. Orange arrow indicates integrated peak of interest for results displayed in Fig. 4b.

Additional file 6: Figure S7. Manganese peroxidase assay measuring absorbance at 270 nm of concentrated Kirk’s mineral media following 5 days of P. chrysosporium growth. Media contained either 2 mg/mL microcrystalline cellulose (blue line) or 10 mg/mL glucose (red line) during the 5 day growth period. N = 3 biological replicates (each point is average of 3 media measurements).

Additional file 11: Figure S11. A. LC/MS standard curve of 4-HBA prepared in methanol at 0.001, 0.005, 0.01, 0.05, 0.1 mM. b. LC/MS standard curve of VA prepared in methanol at 0.001, 0.005, 0.01, 0.05, and 0.1 mM. C. LC/MS standard curve of CA prepared in methanol at 0.001, 0.005, 0.01, 0.05, 0.1, 0.25, 0.5, 0.8, and 1 mM. D. LC/MS standard curve of GGE prepared in methanol at 0.005, 0.01, 0.05, and 0.1 mM. E. LC/MS standard curve of SA prepared in 10% methanol, 90% PBS at 0.01, 0.02, and 0.05 mM.

Additional file 9: Figure S9. A. Representative mass spectra for GGE metabolites (guiacol, protocatechuate, and 2-(2-methoxyphenoxy)-1,3-propanediol) from E. lignolyticus lysates grown in minimal media and incubated with DMSO or GGE for 4 h. Integrated peak for each (from left to right in order listed above) is marked with a red arrow. B. Ion counts for integrated peaks indicated with arrows in A. Gray bars are ion counts of each compound for DMSO lysate, and black bars are ion counts of each compound for GGE lysates. 1 biological replicates is shown. C. Representative mass spectra for GGE metabolics (guiacol, protocatechuate, and 2-(2-methoxyphenoxy)-1,3-propanediol) from E. lignolyticus lysates grown in nutrient broth and incubated with DMSO or GGE for 4 h. Integrated peak for protocatechuate is marked with a red arrows. D. Average ion counts for integrated protocatechuate peak indicated with arrows in C. 3 biological replicates are shown, error bars are standard deviation. Gray bars are ions counts for DMSO lysate and black bars are ion counts for the GGE lysates.

Additional file 10: Figure S10. Area (pX2) of E. lignolyticus cells following internalization of 4-HBA or VA for 4 h. Cells areas were measures using DAPI signal. ****p

Additional file 1: Figure S1. A. Representative mass spectra (QToF-MS) for each uptake compound (4-HBA, VA, SA, CA, and GGE) in P. chrysosporium lysates. DMSO vehicle control mass spectra are shown at the retention time for each compound of interest for peak comparison. Red arrows mark the integrated peak for results in B (4 h). B. Ion counts for each compounds in P. chrysosporium cell lysates from 3 biological replicates. Error bars are standard deviation. C. Ion counts for spiked P. chrysosporium lysate with indicated compounds at a concentration of 0.005 or 0.0025 mM. Spikes were used to calculate the relative lysate concentrations in Fig. 1 from ion counts in Fig. S1B. D. Representative mass spectra for protocatechuate (GGE metabolite) in P. chrysosporium lysates at 4 h. DMSO vehicle control mass spectra shown at the same retention time for peak comparison. Black arrows mark the integrated metabolite peak for Fig. 1c.

Additional file 4: Figure S4. Table summarizing single cell fluorescence microscopy results.

Additional file 8: Figure S8. A. Representative mass spectra (QToF-MS) for each uptake compound (4-HBA, VA, SA, CA, and GGE) in E. coli lysates. Integrated peak for internalization experiments is marked with a red arrow. DMSO vehicle control mass spectra are shown at the retention time for each compound of interest for peak comparison. E. lignolyticus spectra look similar (data not shown). B. ion counts for each compound in E. coli vs. E. lignolyticus cell lysates from 3 biological replicates. Error bars are standard deviation. C. Ion counts for spiked E. coli or E. lignolyticus lysate with indicated compounds at a concentration of 0.005 mM. Spikes were used to calculate the relative intracellular concentrations in Fig. 5.

Additional file 2: Figure S2. A. Representative mass spectra (QToF-MS) for each uptake compound (4-HBA, VA, SA, CA, and GGE) in S. cerevisiae lysates. Integrated peak for internalization experiments is marked with a red arrow. DMSO vehicle control mass spectra are shown at the retention time for each compound of interest for peak comparison. B. Ion counts for each compounds in S. cerevisiae cell lysates from 3 biological replicates. C. on counts for spiked S. cerevisiae lysate with indicated compounds at a concentration of 0.005 mM. Spikes were used to calculate the relative intracellular concentration in Fig. 1 from ion counts in Fig. S2C.