Determination of ionization constant, commonly termed pKa is of prime interest in a wide range of pharmaceutical Research fields. The pKa of a compound is critical as it influences on its physicochemical parameters in biological and environmental systems. The study of pKa is also essential not only for the formulation of drugs and optimization of a variety of novel analytical methods, establishing new pharmaceutical dosage forms yet the exploration of the mechanism of action of drugs. In this research work, we have determined pKa values of isoniazid (INH) derivatives; N'-[(4-methyl benzoyl)] pyridine-4-carbohydrazide (I) and [2-oxo-2-(4-phenyl phenyl) ethyl] (pyridine-4-yl formamido) azanium bromide (II) through UV- spectrophotometry, a method is known for the accuracy and precision of results. These two compounds (I and II) were synthetically prepared in our lab by derivatizing INH and reported by Naeem et al in the year 2014. The mean pKa values for compounds I and II were experimentally determined as 7.37 and 3.76 respectively. The study is helpful in understanding the physicochemical behavior of these compounds in a biological system. Different pharmacokinetic parameters were also predicted using online web tools which ensured significant drug-likeness for both compounds.