Oxime using methoxylamine

Prior to GC-MS analysis, the steroid fractions require derivatization. Ketosteroids are usually converted to oximes using methoxylamine, benzylamine, or hydroxylamine hydrochloride in pyridine. Two epimers of the oxime are formed (E and Z). Hydroxyl groups are then derivatized as trimethylsilyl (TMS) ethers. After formation of the TMS derivative, the extract is further purified by chromatography on a small column of Lipidex 5000. An alternative to TMS is the use of t-butyldimethylsilyl (t-BDMS) ethers, but although conferring desirable mass spectrometric properties, namely an intense M — 57 ion, the t-BDMS group is 42 U heavier than TMS and the formation of poly t-BDMS ethers may increase the relative molecular mass of the steroid derivative beyond the mass range of small quadru-poles. 

A specific reagent used for the derivatization of keto groups is methoxylamine or ethoxylamine. The resulting derivative is a stable oxime. This derivatization can be used alone or in combination with silylation. Examples are described for boldenone [34] and chlorotestosterone (clostebol) [35 37]. 

---