thiourea and benzyl-isothiocyanate

Isothiocyanates (ITC) play an important role in health promotion and cancer prevention due to their anti-bacterial, anti-inflammatory, and anti-cancerogenic properties. However, ITC are highly reactive so that a reaction with further food components is very likely. For example, a reaction of ITC with nucleophilic amino acid side chains of proteins such as cysteine and lysine can occur, reducing the bioavailability of indispensable amino acids and protein functions may be altered. Therefore, it is of great interest to investigate the fate of ITC in the food matrix. Accordingly, the aim of the present study was to investigate the interaction of milk proteins and the ITC benzyl isothiocyanate (BITC) and allyl isothiocyanate (AITC) forming dithiocarbamates and thioureas in milk and curd. After incubating milk and curd with pure ITC or ITC-containing garden cress (Lepidium sativum L.), proteins were isolated, digested, and analyzed via LC-ESI-MS/MS as amino acid derivatives ("conjugates"). Protein conjugates of AITC and BITC were detected in all samples investigated. Further, the acidic pH value in curd favored the formation of dithiocarbamates over the formation of thioureas. Slightly acidic or neutral conditions like in fresh milk favored the formation of thioureas. The investigations also indicated that AITC shows a higher reactivity and dithiocarbamates are formed preferably, whereas incubation with BITC lead to less protein conjugates and the ratio of thioureas and dithiocarbamates was more balanced. In addition, amino acid modifications were often analyzed with indirect methods like measuring the decline of the amino acid residues. In this study, the modified amino acids were analyzed directly leading to more reliable results concerning the amount of modification.

Topics: Chromatography, Liquid; Food Analysis; Food Handling; Hydrogen-Ion Concentration; Isothiocyanates; Lepidium sativum; Milk Proteins; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry; Thiocarbamates; Thiourea

Isothiocyanates form adducts with a multitude of biomolecules, and these adducts need analytical methods. Likewise, analytical methods for hydrophilic isothiocyanates are needed. We considered reaction with ammonia to form thiourea derivatives. The hydrophilic, glycosylated isothiocyanate moringin, 4-(α-L-rhamnopyranosyloxy)benzyl isothiocyanate, was efficiently derivatized to the thiourea derivative by incubation with ammonia. The hydrophobic benzyl isothiocyanate was also efficiently derivatized to the thiourea derivative. The thiourea group provided a UV absorbing chromophore, and the derivatives showed expectable sodium and hydrogen adducts in ion trap mass spectrometry and were suitable for liquid chromatography analysis. Reactive dithiocarbamate adducts constitute the major type of reactive ITC adduct expected in biological matrices. Incubation of a model dithiocarbamate with ammonia likewise resulted in conversion to the corresponding thiourea derivative, suggesting that a variety of matrix-bound reactive isothiocyanate adducts can be determined using this strategy. As an example of the application of the method, recovery of moringin and benzyl isothiocyanate applied to cabbage leaf discs was studied in simulated insect feeding assays. The majority of moringin was recovered as native isothiocyanate, but a major part of benzyl isothiocyanate was converted to reactive adducts.

Topics: Chromatography, High Pressure Liquid; Chromatography, Liquid; Gas Chromatography-Mass Spectrometry; Glycosylation; Isothiocyanates; Molecular Structure; Tandem Mass Spectrometry; Thiocarbamates; Thiourea