RESUMO
The long identified toxic gas, hydrogen sulfide (H2S), which has also been confirmed as the third gaseous signaling molecule following NO and CO, plays important roles in various physiological and pathological process. The current most established quantification method for H2S is HPLC method coupled with fluorescence detection after derivatization with a costly fluorescent reagent, Monobromobimane (MBB). However, The MBB method is characterized by strict reaction condition, long reaction time, tedious operation, and inconsistent reported results. In this study, based on the thiolysis reaction of 7-nitro-2, 1, 3-benzoxadiazole (NBD) ether, the commonly used chromatographic modifier 4-chloro-7-nitro-2,1,3- benzoxadiazole (NBDCl) and four probes (NBDOMe, NBDOEt, NBDOTFE and NBDOCMR) synthesized from NBDCl were tested as alternatives for fast quantification of H2S by LC-MS/MS. The reaction product between NBD ethers/NBDCl and H2S showed special pink color visible to the naked eye and was easy to synthesize and separate in lab; it also showed good retention on common chromatographic columns and high instrument response; therefore it is a good determinand. After establishment of LC-MS/MS methods for all the related compounds, the reaction conditions were optimized for all the probes with H2S. Then the stability, selectivity, reaction rate, sensitivity and quantitative linear relationship between the reaction product and H2S concentration were studied for each probe. Finally, NBDOEt was selected for LC-MS/MS detection of H2S. In comparision with the MBB method, the established NBDOEt method showed matched sensitivity and linearity, better selectivity, and higher repeatability; and had the advantages of easy operation, simple reaction condition, and cheap raw materials. The method was successfully validated and applied to determination of Na2S content in Na2Sâ9H2O bulk drug and injection. In conclusion, NBDOEt is a promising option for quantification of H2S in abiotic matrix.
