The determination of patulin from food samples using dual-dummy molecularly imprinted solid-phase extraction coupled with LC-MS/MS.

A molecularly imprinted polymer (MIP) with specific adsorption for patulin was successfully polymerized by precipitation polymerization using 2-oxindole (2-oxin) and 6-hydroxynicotinic acid (6-HNA) as dummy template molecules, methylacrylic acid (MAA) as a functional monomer, trimethylolpropane trimethacrylate (TRIM) as a crosslinker, 2,2-azobis-(2-methylpropionitrile) (AIBN) as a initiator, and methanol as a porogen solvent. The molecularly imprinted solid phase extraction (MI-SPE) column was prepared using the polymer as a sorbent and applied for the selective extraction of patulin from real samples. The results showed that the MI-SPE method had high selectivity and specific adsorption towards patulin with mean recoveries ranged between 81.3% and 106.3% and a relative standard deviation (RSD) < 4.5%. Additionally, the developed MI-SPE method coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) exhibited good linearity in the range of 1-100 ng mL-1 with correlation coefficients (R2) >0.998. The limits of detection (LODs, S/N = 3) were 0.05-0.2 ng g-1, and the limits of quantification (LOQs, S/N = 10) were 0.2-0.5 ng g-1. The developed method showed a better purification and higher patulin recovery for real samples than the quick, easy, cheap, effective, rugged, safe "QuEChERS" method.

Determination of formylated DNA and RNA by chemical labeling combined with mass spectrometry analysis.

Nucleic acids carry diverse chemical modifications that exert critical influences in a variety of cellular processes in living organisms. In addition to methylation, the emerging DNA and RNA formylation has been reported to play functional roles in various physiological processes. However, the amounts of formylated DNA and RNA are extremely low and detection of DNA and RNA formylation is therefore a challenging task. To address this issue, we developed a strategy by chemical labeling combined with in-tube solid-phase microextraction - ultra high performance liquid chromatography - electrospray ionization - tandem mass spectrometry (in-tube SPME-UPLC-ESI-MS/MS) analysis for the sensitive determination of DNA and RNA formylation. Using the developed method, we were able to simultaneously measure six formylated nucleosides, including 5-formyl-2'-deoxycytidine (5-fodC), 5-formylcytidine (5-forC), 5-formyl-2'-deoxyuridine (5-fodU), 5-formyluridine (5-forU), 2'-O-methyl-5-formylcytidine (5-forCm) and 2'-O-methyl-5- formyluridine (5-forUm), from DNA and RNA of cultured human cells and multiple mammalian tissues. The detection limits of these formylated nucleosides improved by 307-884 folds using Girard's P (GirP) labeling coupled with in-tube SPME-UPLC-ESI-MS/MS analysis. It was worth noting that 5-forU, 5-forCm and 5-forUm which have not been detected in human sample before, were discovered in cultured human cells and tissues in the current study. In addition, we observed significant increase of 5-forC and 5-forU in RNA (p = 0.027 for 5-forC; p = 0.028 for 5-forU) and 5-fodU in DNA (p = 0.002) in human thyroid carcinoma tissues compared to normal tissues adjacent to the tumor using synthesized stable isotope GirP (d5-GirP)-assisted quantification. Our results indicated that aberrant DNA and RNA formylation may contribute to the tumor formation and development. In addition, monitoring of DNA and RNA formylation may also serve as indicator for cancer diagnostics. Taken together, the developed chemical labeling combined with in-tube SPME-UPLC-ESI-MS/MS analysis can facilitate the in-depth functional study of DNA and RNA formylation.