59


OH group) with a trimethylsilyl group. Silylation then occurs through a SN2, nucleophilic attack. The general reaction for the formation of trialkylsilyl derivatives is shown below, with the Cl atom being the leaving group, Figure 1.


Silyl reagents will react with both alcohols and acids to form trimethylsilyl ethers and trimethylsilyl esters, respectively. The derivatives formed are volatile, and for the most part, are easily separated. Silyl reagents are influenced by both the solvent system, with a common regent supplied with BSTFA and MSTFA being trimethylchlorosilane which increases the reactivity of the reagent. It is important to be aware of the effect of the solvent system, in particular if any form of sample preparation has been employed and this has resulted in the analytes being effectively transferred to another solvent.


The derivatisation of a compound is a chemical reaction that must be controlled. Ideally the process will yield only one product, with the overwhelming majority of the initial compound being converted to the final derivatised form of the molecule. However, given that in many analyses there will be a number of compounds present in the mixture prior to the derivatisation stage, this can lead to a wide range of compounds being produced. Consequently, it is often necessary to use a highly selective detector after the derivatisation process to ensure that only the compound of interest is being detected. It is also important that the reaction conditions do not cause the derivatising reagent to decompose.


An understanding of the chemistry is essential to ensure that the correct derivatised form of the molecule is produced. There are many derivatising reagents that are available and the help desk will go through some of the more common reagents that are used and also some of the challenges that can be faced when using these reagents.


There are a variety of reagents used for the silylation derivatisation process including; Hexamethyldisilzane (HMDS), Trimethylchlorosilane (TMCS),


Trimethylsilylimidazole (TMSI), Bistrimethylsilylacetamide (BSA), Bistrimethylsilyltrifluoroacetamide (BSTFA), N-methyltrimethylsilyltrifluoroacetamide (MSTFA), Trimethylsilyldiethylamine (TMS-DEA), N-methyl-N-t-butyldim ethylsilyltrifluoroacetamide (MTBSTFA), and Halo-methylsilyl derivatisation reagents. The latter regents can be used with electron capture detection (ECD) to improve selective sensitivity with electron capture detectors. The most common regents for cannabinoid analysis are BSTFA and MSTFA, which due to their chemistry, react quicker than the other reagents listed with complete reactions taking less than 30 minutes.


The analysis of cannabis is seeing an increase due to increased interest in the therapeutic applications of this drug, and also in the detection of the drug when it is used in a more recreational manner. There are different approaches that can be employed to analyse the active components, but GC is still an extremely popular approach. Robust methods have been developed utilising derivatising reagents allowing for the separation and detection of the acidic and neutral forms of the cannabinoid. The sample can influence the derivatising process, as can the quality of the reagents that are used, and in the case of a silylating reagent, water contamination is critical. Other applications of derivatisation are shown in Table 1.


Table 1: Applications for chemical derivatisation Functional Group Reaction Type


Silylation


Alcohols and Phenols


Derivatisation Reagent


BSA, BSTFA, MTBSTFA


Acylation Alkylation Silylation Carboxylic acids Acylation Alkylation Silylation Active hydrogens Acylation


Carbohydrates and Sugars


Amides


Acylation Alkylation


Silylation Amines Acylation Amino acids Catecholamines


Alkylation Silylation Acylation Alkylation Silylation Acylation


Inorganic anions Silylation Silylation


Nitrosamine Acylation Alkylation Sulphonamides Sulphides Acylation Silylation Silylation Silylation


Heptafluorobutyrylimidazole, Pentafluoropropionic


Anhydride, Trifluoroacetic anhydride, N-Methylbis(trifluoroacetamide) Dimethylformamide,


Pentafluorobenzyl bromide


Bis(trimethylsilyl)–acetamide, BSTFA,


Trimethylsilylimidazole, MTBSTFA


Pentafluoropropanol /


Pentafluoropropionic anhydride Dimethylformamide,


Tetrabutylammonium hydroxide


Bis(trimethylsilyl)–acetamide, Bistrimethylsilyltrifluoroacetamide / Trimethylchlorosilane, Hydrox-Sil, N-methyl- trimethylsilyltrifluoroacetamide, Pentafluoropropanol /


pentafluoropropionic anhydride


Hexamethyldisilzane, TMSI


BSA,


N, O-bis-(trimethylsilyl)- trifluoroacetamide


Heptafluorobutyrylimidazole Dimethylformamide


BSTFA, MTBSTFA


Trifluoroacetic anhydride, Pentafluorobenzoyl chloride, Heptafluorobutyrylimidazole Dimethylformamide (Diacetals)


BSTFA, TMSI


Heptafluorobutyrylimidazole Dimethylformamide,


Tetrabutylammonium hydroxide TMSI


Pentafluoropropionic anhydride, Heptafluorobutyrylimidazole,


BSTFA, MTBSTFA BSTFA


HFBA, Pentafluoropropionic anhydride, Trifluoroacetic anhydride


Dimethylformamide, Pentafluorobenzyl bromide


Trifluoroacetoic & Heptafluorobutyric Anhydride,


Pentafluorobenzyl bromide TMSI


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