2-methylimidazole

RNA oligonucleotides were prepared by standard phosphoramidite oligonucleotide synthesis using a MerMade 6 DNA/RNA synthesizer (Bioautomation, Plano, TX) or starting with 2′,3′-diacetyl nucleosides using standard manual coupling procedures (Beaucage and Caruthers, 1981 (link)). 5′-phosphates were installed using bis-cyanoethyl-N,N-diisopropyl phosphoramidite from ChemGenes by standard phosphoramidite coupling chemistry.
Mononucleotide monophosphates and oligonucleotide monophosphates were activated using a modified published protocol (Joyce et al., 1984 (link)). As an example, 2-MeImpAGC (the 2-methylimidazolide of the trimer 5′-phosphoro-AGC) was synthesized by first dissolving 5 mg (5 μmole) of 5′-phosphoro-AGC in 1 mL of dimethyl sulfoxide (DMSO). To that mixture 160 mg (2 mmoles) of 2-methylimidazole, 56 mg triphenylphosphine (260 μmole), 64 mg 2,2′-dipyridyldisulfide (290 μmole) and 40 μL triethylamine (550 μmole) were added. After stirring overnight at room temperature, the mixture was precipitated in 10 mL of a 400:250:30:1 mixture of acetone:diethylether:triethylamine:saturated solution of NaClO₄ in acetone. The precipitate was pelleted by centrifugation (3000 rpm, 5 min) and washed twice with a 1:1 mixture of acetone:diethylether and once with pure diethylether. After decanting the solvent, the pellet was dried under vacuum, resuspended in deionized water and purified by high-performance liquid chromatography (HPLC) over a C₁₈ column (Alltima C₁₈ 5 μm, Thermo Fisher, 250 × 10 mm) with 25 mM triethylammonium bicarbonate (TEAB) pH adjusted to 7.5 in 2% (v/v) acetonitrile for mobile phase A and acetonitrile for mobile phase B over a gradient beginning at 100% A and falling to 80% A over 20 min with a flow rate of 3 mL/minute. The fraction containing the product was verified by electrospray ionization mass spectrometry (ESI/MS) in negative mode (m/z = 996), frozen in liquid nitrogen and lyophilized overnight to yield a white powder. The powder was dissolved in water and the concentration of the activated trimer was determined using a NanoDrop 2000c spectrophotometer (Thermo Fisher Scientific, Waltham, MA) and calculated assuming ε₂₆₀ = 34,170 M⁻¹ cm^{−1 29}.
Monomer stocks were adjusted to pH 7 before adding into the reaction mixture.

CME@ZIF-8 was synthesized using a method consistent with prior research [4 (link)]. Specifically, 0.219 g of zinc acetate hexahydrate was mixed in 10 mL of distilled water, while 0.328 g of 2-methylimidazole was blended in 20 mL of distilled water, along with 0.5 g of CME. The mixture of CME and 2-methylimidazole was stirred for 15 min before the gradual addition of the zinc acetate solution. Within 3 to 4 min, the brown reaction solution underwent a color change, turning creamy in appearance, representing the makeup of the CME@ZIF-8 nanocomposite. Following this, the CME@ZIF-8 nanocomposite solution was left at room temperature for 24 h. To obtain the precipitate of the CME@ZIF-8 nanocomposite, the solution was centrifuged at 10,000× g rpm for a spell of 10 min. Successively, it experienced three rounds of washing with 10 mL of distilled water each time to remove any residual unreacted chemicals. The resulting material was then dried at 80 °C. Following this, both the CME@ZIF-8 nanocomposite and ZIF-8 were crushed into a powder using a mortar and pestle, as depicted in Figure 1.

TRD ZIF-8 nanoparticles were synthesized as reported previously^{33 (link)}. In a typical synthesis of the TRD ZIF-8 nanoparticles with the diameter of 260 nm and {110}/{100} exposure ratio of 0.43, Zn(OAC)₂·2H₂O (300.0 mg) was dissolved in 5.0 mL of deionized (DI) water. In a separate beaker, 2-methylimidazole (1170.0 mg) and CTAB (9.7 mg) were dissolved in 5.0 mL of DI water. The two solutions were mixed together in a 100 mL round bottom flask, and the mixture was stirred for a few seconds. Then, the resulting solution was placed at room temperature for 2.5 h. The obtained TRD ZIF-8 nanoparticles were washed with DI water for three times and collected by centrifuging at 9000 rpm (centrifugal force, 9690 g) for 10 min. The final products were dispersed in 10.0 mL of ethanol (20 mg/mL). The other ZIF-8 nanoparticles with different sizes and {110}/{100} exposure ratios were synthesized by tunning the reaction time and CTAB/2-methylimidazole ratio.