BlOORGANlC CHEMISTRY 659-67 (1979) Acid-Base Properties of 4-Nitro-L-histidine and Related Compounds ERNEST GIRALT, MARIA-DOLORS LUDEVID, FERNANDO ALBERICIO, AND MARIONA BASSEDAS Departamento de Q&mica Orgdnica, Facultad de Quimica, Universidad de Barcelona, Barcelona, 28. Spain Received March 27.1978 Acid-base properties of 4-nitro+histidine (3), N”-acetyl-4-nitro-thistidine (Z), and N”-acetyl-4-nitro-t-histidine methyl ester (1) are studied. Their pKF values can be conveniently determined by ultraviolet spectroscopy. Potentiometric titration and ‘H-nuclear magnetic resonance (nmr) titration can also be used. Introduction of a nitro group strongly enhances the acidity of all the compounds. pK,* of compound (3) has been also spectrophotometrically determined. Observed differences in acidity, in those cases where solvation does not play a major role, can be explained by assessing the influence of electrostatic charges on pK,, following Bjerrum’s general theory with the aid of data from Wnmr conformational analysis. INTRODUCTION 4-Nitro-L-histidine (3) can be obtained by nitration of L-histidine (1). A strong decrease in the basicity of the imidazolic ring of this product can be expected as a result of the electron-withdrawing character of the nitro group. Replacement of 4-nitro-L- histidine for L-histidine in biologically interesting peptides could thus become a useful tool to evaluate the contribution of the acid-base properties of the side chain of histidine to the biological activity of natural peptides or proteins (2). Usually, acidity constants have been determined by the classic potentiometric titration method. Since 1954, however, spectrophotometric methods have been used when the substance has convenient chromophoric properties (3). In the present paper, the spectrophotometric determination of the ionization constants of N”-acetyl-4-nitro-t- histidine methyl ester (l), iV”-acetyl-4-nitro-L-histidine (2), and 4-nitro-L-histidine (3) is described. A conformational study by ‘H-nmr spectroscopy is used to discuss the results obtained. From a biological viewpoint, compound (1) is the most interesting of the three, since its blocked amino and carboxyl functions reproduce to a better degree the situation of a nitrohistidine residue in a protein sequence. EXPERIMENTAL Ultraviolet and mm (60 MHz) spectra were recorded in Perkin-Elmer 124 and Perkin-Elmer R- 12 spectrometers. pH measurements were made with a Radiometer 5 1 pH meter equipped with a glass-calomel combined electrode. 59 00452068/79/010059-09$02.00/0 Copyright @ 1979 by Academic Press, Inc. All rights of reproduction in any form reserved. Printed in Great Bntain