Derivatization procedures for the detection of beta(2)-agonists by gas chromatographic/mass spectrometric analysis

An evaluation of derivatization procedures for the detection of beta (2)-ag onists is presented. The study was performed with the beta (2)-agonists bam buterol, clenbuterol, fenoterol, formoterol, salbutamol, salmeterol and ter butaline. Different derivatizating agents were employed, aiming to obtain d erivatives with high selectivity to be used in the gas chromatographic/mass spectrometric analysis of beta (2)-agonists in biological samples. Trimeth ylsilylation was compared with different agents and the role of some cataly sts was evaluated. Acylation, combined trimethylsilylation and acylation, a nd the formation of cyclic methylboronates were also studied. Sterical hind rance caused by different substituents at the nitrogen atom of the beta -et hanolamine lateral chain of beta (2)-agonist molecules is mainly responsibl e for differences in the abundances of the derivatives obtained. The use of catalysts produces an increase in the derivatization yield, especially for compounds with low steric hindrance (substituents with primary and seconda ry carbon atoms). The formation of trimethylsilyl (TMS) ethers is not influ enced by structural molecular differences when only hydroxy groups are invo lved in derivatization. Combined trimethylsilylation and acylation showed t hat compounds with a secondary carbon atom linked to the nitrogen atom form mainly N-TFA-O-TMS derivatives, with a small amount of N-TMS-O-TMS derivat ives. Compounds with tert-butyl substituents at the amino group (bambuterol , salbutamol and terbutaline) formed O-TMS derivatives as the main products , although a limited amount of trifluoroacylation at the nitrogen atom also occurred. Cyclic methylboronates were formed with bambuterol, clenbuterol, formoterol, salbutamol and salmeterol. Owing to hydroxy substituents in un suitable positions for ring formation, this procedure was not effective for fenoterol and terbutaline. Mass spectra of different derivatives and tenta tive fragmentation profiles are also shown. For screening purpose (e.g. spo rts drug testing), derivatization with MSTFA or BSTFA alone is recommended as a comprehensive derivatization technique for beta (2)-agonists owing to minimal by-product formation; formation of cyclic methylboronates can be us eful for confirmation purposes. Detection limits were obtained for the TMS and cyclic methylboronate derivatives using the derivatizing reagents MSTFA and trimethylboroxine, respectively. For most of the compounds, lower dete ction limits were found for the TMS derivatives. Copyright (C) 2000 John Wi ley & Sons, Ltd.