Barbituric acid

The nitroketene dithioacetals, cysteamine hydrochloride, isatin, malononitrile, ethyl cyanoacetate, methyl cyanoacetate, cyanoacetohydrazide, barbituric acid, derivatives of isatin, triethylamine and solvents were obtained from Sigma Aldrich and used without further purification. Nano-silica (CAB-O-SIL® M5) was obtained from Cabot Co. IR spectra: Bruker Tensor 27 spectrometer. NMR spectra: Bruker DRX-300 Avance instrument (300 MHz for ¹H and 75.4 MHz for ¹³C) with DMSO-d₆ as solvents. Chemical shifts are expressed in parts per million (ppm), and coupling constant (J) are reported in hertz (Hz). Mass spectra: Agilent 5975C VL MSD with Triple-Axis detector operating at an ionization potential of 70 eV. Elemental analyses for C, H and N: Heraeus CHNO-Rapid analyzer. Melting points: electrothermal 9100 apparatus.

Silver nitrate, cobalt(II) nitrate, dimethylformamide, 2,6-pyridinedicarboxylic acid, ethanol, sodium hydroxide, Hydrazine hydrate, ethyl acetoacetate, aryl aldehydes, and barbituric acid derivatives were purchased from Sigma-Aldrich. The chemicals were used without further purification.

A dual 3% gel phantom with 50 mM creatine, and barbituric acid (Sigma-Aldrich Co. St. Louis, MO. USA) was prepared for this study. These two molecules, with chemical shifts of 2 and 5 ppm, respectively, relative to the water proton resonance at 0 ppm, were used to model a small chemical shift case. To prepare the phantom, the agarose solution was first heated to boiling and then cooled under room temperature till 70 °C. Then the contrast agent was added in and mixed properly. The pH value of each sample was adjusted by phosphate buffer. The pH value of creatine and barbituric acid solutions were respectively titrated to 6.98 and 7.10 monitored by an electronic pH-meter (SUNTEX Instruments, Taiwan). Proper pH value was used to achieve adequate CEST contrast. After titrating process was finished, these two samples were quickly transferred to two 6 mL glass test tubes each. The two glass tubes were then placed in a 50 mL centrifuge tube. The centrifuge tube was filled with pure 3% agarose solution in the outer compartment. The phantom was then left to solidify under room temperature before the experiment was conducted.

Standards of nicotine, cotinine, chlorogenic acid, sodium acetate, potassium cyanide, chloramine T, barbituric acid, sodium metabisulfite, and acetone were purchased from Sigma-Aldrich (St. Louis, MO, USA). All chemicals were of analytical grade. F/12 medium was obtained from the American Type Culture Collection (ATCC, Manassas, VA). Dulbecco's modified eagle's medium (DMEM) and fetal bovine serum (FBS) were purchased from Cellgro (Manassas, VA). Penicillin/streptomycin was obtained from Gibco (Introgen Corporation, Grand Island, NY).