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Purity: ≥98%
1-NBDG, an analog of 2-NBDG, is a fluorescent analogue of glucose and an indicator used for measuring glucose uptake by bacteria and live mammalian cells and in tumor biopsies. The uptake of 1-NBDG is competitively inhibited by D-glucose, but not L-glucose or sucrose, in E. coli. Unlike 2-NBDG, which is a good substrate of GLUTs but not SGLTs, 1-NBDG can be transported by both GLUTs and SGLTs. Thus, 1-NBDG is useful for the screening of SGLT1 and SGLT2 inhibitors.
| ln Vitro |
1-NBDG was used as a fluorescent glucose analogue to conduct glucose uptake assay in COS-7 cells transiently transfected with pcDNAhSGLT1 and CHO–K1 cells stably expressing hSGLT2 for the screening of SGLT1 and SGLT2 inhibitors, respectively. Here we demonstrated that it was also feasible to use COS-7 cells transiently transfected with pcDNAhSGLT2 (hSGLT2/COS-7) for the screening of SGLT2 inhibitors. The method was also extended from 24-well format to 96-well format suitable for high-throughput screening.The validated method using 1-NBDG was applied to test 195 crude methanol extracts from approximately 60 species of plants in the Lauraceae family in search of novel natural SGLT inhibitors and two potent natural SGLT1 and SGLT2 inhibitors were identified from the leaves of Cinnamomum macrostemon (Yang et al., manuscript in preparation). The IC50 values of these two compounds (I and II) were in the nanomolar range and comparable to phlorizin, the most potent natural SGLT inhibitor with IC50 of 110 ± 10.2 nM for SGLT1 and 9.65 ± 1.83 nM for SGLT2 in our assay system.
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| Enzyme Assay |
Determination of the stability of 1-NBDG and 2-NBDG under alkaline, acidic conditions or at different pH.
1-NBDG or 2-NBDG was diluted in 0.2 N NaOH, 0.2 N HCl or 20 mM Tris buffers with pH values of 6.8, 7.5, 8.0, 8.8 and 9.8. Fluorescence was detected in a black 96-well plate using a SpectraMax Paradigm Multi-Mode Microplate Reader (Molecular Devices) at an excitation wavelength of 458 nm and an emission wavelength of 528 nm (458/528 nm) for 1-NBDG and 475/550 nm for 2-NBDG. |
| Cell Assay |
Cell-based screening method for SGLT1 and SGLT2 inhibitors using 1-NBDG
COS-7 cells were transfected with pcDNAhSGLT1 or pcDNAhSGLT2 as previously described and plated into 96-wells the following day at 3–5 × 104 cells/well. Cells were allowed to attach overnight and reach 100% confluence, then subjected to glucose uptake assay in sodium buffer containing test compounds and 100 or 160 μM 1-NBDG at 37 °C for 1.5 h (SGLT1) or 2 h (SGLT2). Cells in 96-wells were rinsed with 150 μL of warm choline buffer and then incubated with 50 μL of sodium buffer containing 1-NBDG and test compounds. After incubation, cells were washed twice with cold choline buffer, examined under microscope to check cell conditions, then lysed in 75 μL of 0.2 N NaOH, and neutralized with 75 μL of 0.2 N HCl. Lysates (100 μL) were transferred to a black 96-well plate for fluorescence detection. Phlorizin was used as a positive control for SGLT inhibitor and 1-NBDG in choline buffer was used to measure sodium-independent glucose uptake via GLUTs. Cells in sodium buffer without 1-NBDG were used to measure autofluorescence (cell background). |
| References |
J Food Drug Anal. 2021; 29(3): 521–532.
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| Molecular Formula |
C12H14N4O8
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| Molecular Weight |
342.264
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| Exact Mass |
342.0812
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| CAS # |
2376921-70-7
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| Synonyms |
1-NBDG; 1NBDG; 1 NBDG
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.9218 mL | 14.6088 mL | 29.2176 mL | |
| 5 mM | 0.5844 mL | 2.9218 mL | 5.8435 mL | |
| 10 mM | 0.2922 mL | 1.4609 mL | 2.9218 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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