Isolation determination of garlic allyl sulfides and their antioxidant activity

Abstract: The allyl-substituted cysteine derivatives were major components of sulfur compounds in garlic. This study separated and identified 3 allyl-substituted cysteine derivatives, i.e. S-allyl-L-cysteine (SAC), S-allyl-L-cysteine sulfoxide (ACSO) and γ-L-glutamyl-S-allyl-L-cysteine (GSAC). Furthermore, scavenging ability of 1,1-Diphenyl-2-picrylhydrazyl (DPPH) free radical and iron chelating ability were tested through comparing the compounds of SAC, ACSO and GSAC with glutathione (GSH) to evaluate the antioxidation. SAC, ACSO and GSAC were isolated from garlic bulbs using ion-exchange chromatography and pre-HPLC (high performance liquid chromatography). Their molecular structures were identified by high performance liquid chromatography-electrospray ionization-mass/mass spectrometry (HPLC-ESI-MS/MS), proton nuclear magnetic resonance (1H NMR), carbon-13 nuclear magnetic resonance (13C NMR) and specific rotatory power. SAC and ACSO which yielded the expected M+H+ of m/z 162.1 and 177.8 by the measurement of mass spectrometry under positive ion mode were conferred as S-allyl-L-cysteine (C6H11O2NS) and S-allyl-L-cysteine sulfoxide (C6H11O3NS) respectively compared with the data of corresponding standard. GSAC, showing the corresponding M-H- of m/z 288.8, was identified as γ-glutamyl-S-allyl-L-cysteine (C11H18O5N2S) by HPLC-MS. The 1H NMR spectrum of SAC and ACSO indicated the presence of the cysteinyl (δ 2.96 and 3.83, 3H) and (δ 3.26 and 4.13, 3H) moieties. The 13C NMR spectrum of compound SAC and ACSO showed the presence of the cysteinyl carboxyl (δ 175.6 and δ 188.0) carbons. Similarly, the 1H NMR spectrum of GSAC indicated the presence of the glutamyl (δ 2.32-2.43, 2.66 and 3.46, 5H) and cysteinyl (δ 2.75 and 4.76, 3H) moieties. The 13C NMR spectrum of GSAC revealed the presence of 11 magnetically nonequivalent carbon atoms, with three of them being the glutamyl carboxyl, glutamyl carbonyl and cysteinyl carboxyl (δ 182.5, 183.2 and 184.4) carbons. Meanwhile, the scavenging activity of DPPH free radical and the chelating activity of iron ion by allyl-substituted sulfides (SAC, ACSO and GSAC) were determined with reference to cysteine derivative of GSH. The scavenging activity of DPPH free radical had no significant difference between GSH (71.14%) and both of SAC (73.55%) and GSAC (72.68%). Iron ion is one of the main oxidizing auxiliaries which can initiate free radical in vitro and vivo, therefore, chelating with iron ion can remove the oxidizing auxiliaries effectively, and prevent the formation of free radicals. Likewise, no significant differences between GSH (81.21%) and both of SAC (79.18%) and ACSO (78.13%) were observed for the chelating activity of iron ion tested in this study. In particular, the chelating activity of iron ion of GSAC (92.76%) was greater than GSH. These results confirmed the antioxidant activity of these allyl-substituted cysteine derivatives. Garlic planting area in China accounted for a third of the world. At harvest time, garlic waste dumping in the field led to the waste of resource and the pollution of the environment, and these waste parts of garlic were also the sources of sulfur compounds. The processing in this experiment could make full use of garlic (edible part and non-edible part), and the study could provide some theoretical guidance for the industrialization of large-scale extraction of sulfur-containing products, and improve the level of deep processing of garlic. The work revealed the first evidence of antioxidant activity of GSAC, which could be involved in its chelating activity of iron ion.