# Analysis of Potent Odour-Active Volatile Thioids in Foods and Beverages with a Focus on Wine
## 3 Thiol Isolation--Extraction and Derivatisation
### Derivatisation Approaches
#### 3.2.2 Derivatisations for LC Analysis of Thiols  
The abovementioned specific extraction and derivatisation approaches that involve Hg\({}^{+}\), Ag\({}^{+}\), PFBBr, and ETP are for GC based systems. Emerging some time after many of the GC methods were developed, derivatisation-based procedures that were designed for LC analysis are perhaps one of the most important developments in thiol isolation. In comparison to their GC counterparts, the suggested derivatisation protocols for LC analysis of volatile thiols have tended to simplify the overall extraction protocol and offer excellent sensitivity in a variety of matrices. Derivatisation reagents applied for LC-based analysis of thiols (Table 1, Entries 15-19) include conventional reagents, like \(\alpha\)-phthaldialdehydedehyde (OPA) [36], 2-phenyl-1,2-benzisosenelenzol-3(2H)-one (ebselen) [37; 38; 39; 40], 4,4\({}^{\prime}\)-dithiodipyridine (DTDP) [41; 42], and 1-(4-(1\(H\)-phenanthro[9,10-d]imidazol-2-yl)phenyl)-1\(H\)-pyrrole-5-dione (PIPD) [45], as well as stable isotope labelled reagent pairs d\({}_{0}\)/d\({}_{7}\)-\(\omega\)-bromoacetylquinolinium bromide (d\({}_{0}\)/d\({}_{7}\)-BQB) [43; 49] and d\({}_{0}\)/d\({}_{4}\)-acidrone-10-ethyl-\(N\)-maleimide (d\({}_{0}\)/d\({}_{4}\)-AENM) [44]. The reagents readily react with the sulfhydryl group, and derivatisation reactions are often performed in various formats, such as in conjunction with LLE, SPE, or GP-MSE prior to LC analysis, to assist with sample clean-up and enrichment. OPA was considered for thiol derivatisation in white wine [36] due to the reactivity of the dialdehyde functionality with amino acids and other nucleophiles, including thiols [50], thus beginning the exploration of LC-based approaches for the analysis of volatile thiols in wine (Table 1, Entry 15). Derivatisation with OPA in the presence of ethanolamine under basic conditions is rapid (5 min at room temperature in borate buffer), but 4-MSP was unable to be derivatised and the pre-derivatisation sample preparation steps are rather complicated. Briefly, wine has to be treated with potassium metabasulfite and polyvinylpolypyrrolidone, followed by pH adjustment and reaction with borohydride, and then LLE with CH\({}_{2}\)Cl\({}_{2}\) (possibility of forming an emulsion) and sample concentration steps prior to derivatisation [36]. Moreover, the OPA-thiol derivatives were unstable, even when stored at \(-80\)\({}^{\circ}\)C, and their rapid and significant degradation would ultimately lead to a loss of sensitivity and inaccurate quantitation [36]. The Se-N-containing reagent 2-phenyl-1,2-benzisosenelenzol-3(2H)-one (ebselen) selectively and efficiently reacts with thiols by the cleavage of Se-N bond and formation of an Se-S bond with the -SH group [51]. Ebselen has been used to derivatisate a range of thiols (such as those in Figure 1b) in various matrices (lipid: olive oil [37]; hydroalcoholic: wine [38], beer [38]; aqueous: brewed coffee [39]; organic extract: roasted coffee [40]) (Table 1, Entry 16). The proposed ebselen derivatisation approaches are fast, single-step derivatisation/extraction (\(\sim\) 1 min), with some slight variations in initial sample preparation, solvent choice, solvent volumes, and workup steps, depending on sample matrices. In general, a suitable solvent containing ebselen (or with ebselen introduced separately) is added to the solid or liquid sample. After vortexing for a short period of time, the organic phase is collected, concentrated, and the residue is re-dissolved for analysis (or analysed directly without concentration) [37; 38; 39; 40]. The major advantages of these approaches when compared to aforementioned derivatisation procedures is they are much less complicated and they employ mild conditions, although they still require the handling of samples under N\({}_{2}\), and the instruments are all high resolution mass spectrometers as opposed to the more common triple quadrupole.