Determination of Beta-Lactam residues in milk by high performance liquid chromatography

A high performance liquid chromatographic method to assay beta-lactam residues in milk was developed and validated. Milk samples were spiked with standard solutions and deproteinated. The extract was cleaned-up on C18 SPE cartridge, the antibiotics eluted with acetonitrile:water (50:50 v/v) and derivatized with acetic anhydride and 1-methyl-imidazole solution containing HgCl2. The chromatographic analysis was performed on C18 column using mobile phase consisting of acetonitrile and phosphate buffer (pH 6.5) in the presence of Na2S2O3 gradient and detection at 325 nm. The method was selective for ampicillin, penicillin G and penicillin V, the latter used as internal standard. Average recoveries for ampicillin and penicillin G ranged, respectively, from 60.0% to 104.9% and from 82.7% to 109.2%, with coefficients of variation from 11.1% to 24.6%, and from 2.1% to 25.2%, indicating accuracy and precision. Detection limit of 4.0 µg/L for ampicillin and 3.0 µg/L for penicillin G, and quantification limits of 4.0 µg/L for both were estimated.<br>Um método para determinar resíduos de antibióticos beta-lactâmicos em leite por cromatografia líquida de alta eficiência (CLAE) foi desenvolvido e validado. Amostras brancas foram adicionadas de padrão e desproteinizadas. O extrato foi purificado por extração em fase sólida C18, os antibióticos eluídos com acetonitrila:água (50:50 v/v) e posteriormente derivatizados com anidrido acético e solução de 1-metil-imidazol contendo HgCl2. A análise cromatográfica foi realizada utilizando coluna C18, fase móvel composta por acetonitrila e tampão fosfato pH 6,5, na presença de Na2S2O3 em gradiente e detecção a 325 nm. O método foi seletivo para ampicilina, penicilina G e penicilina V, sendo este último utilizado como padrão interno. As médias de recuperação para ampicilina e penicilina G situaram-se, respectivamente, na faixa de 60,0% a 104,9% e de 82,7% a 109,2%, com coeficientes de variação na faixa de 11,1% a 24,6%, e de 2,1% a 25,2%, indicando exatidão e precisão. Limites de detecção e quantificação de 4,0 µg/L para ampicilina e de 3,0 µg/L para penicilina G foram determinados

Silica and other materials as supports in liquid chromatography. Chromatographic tests and their importance for evaluating these supports. Part I

Reversed-phase liquid chromatography (RP-HPLC) has become a powerful and widely employed technique in the separation and analysis of a great variety of compounds with different functionalities. The most common type of stationary phase for RP-HPLC consists of nonpolar, hydrophobic organic species (e.g., octyl, octadecyl) attached by siloxane bonds to the surface of a silica support. In the first part of this article, a description of the many beneficial properties that make porous silica the most employed support in RP-HPLC will be presented, starting from the synthesis of silica. It is noteworthy that the chromatographic properties of the final column are strictly correlated to the preparation type. A silica surface possesses a number of attractive properties, but also some drawbacks. Unreacted or residual silanols interact with basic compounds and can induced peak tailing, which means a loss in chromatographic performance. This problem has lead many manufactures to produce stationary phases with reduced silanol activity which improve dramatically the peak shape of basic compounds. In the second part of this review, different approaches are proposed to obtain less reactive stationary phases