The high-pressure liquid chromatographic analysis of a series of biphenyls, substituted at positions 2-, 3-, 4-, 4,4'-, and 3,4-, has been carried out in the.three common modes: adsorption, normal partition, and reversed-phase partition. Ultraviolet absorption monitoring of the eGluent was used as the method of detection.. Capacity ratios for the derivatives have been determined in the three modes, w.here possibIe, and a discussion of the effkcts of the various substituents upon the retention times is presented.. The utility of the three modes of chromatography in separating some mixtures of these biphenyl derivatives is illustrated.
INTRODUCHON
We have been interested in using the techniques of radioimmu&kssay (RIA) to monitor exposure to chemical carcinogens such as the aromatic amines benzidine and Caminobiphenyl. This required the synthesis of a wide variety of biphenyl derivatives of the two parent amines for use in the development of the RIA procedures ; Le., haptens for conjugation to carrier proteins for use in the developmknt of antibodies, derivatives @able for iodination to yield radiolabeled haptens and various compounds referred to as inhibitors for testing the specificity of the atitibadies.
We decided to use high-pressure liquid chromatography (HPLC)'*' in conjunction with other analytical techniques such as thin-layer chromatography, elemental analysis, mass spktrometry, infrared and ultraviolet spectrophotometry, and nuclear magnetic resonance spectroscopy to establish homogeneity and purity of each compound. In addition, HPLC allowed us to scan rapidly a reaction mixture and monitor it for product formation. We present in this paper a description of the behavior of these biphenyl compounds in three liquid chromatographic modes. ~ Eza?ERIMEmm C~ramQtagrap~y ~eqrcipment ---. -A Varian 4200 series liquid chromatograph (Varian Assoc., Palo Alto, Calif.,