## 1 Introduction  
In the drug development life cycle, analytical methods development is an essential and indispensable element. A number of guidelines regulate qualitative and quantitative aspects of analytical methods development [1]. Drug approval by regulatory authorities requires the drug candidate to prove control of the entire process of drug development, starting from development of the active pharmaceutical ingredient (API) synthetic process to final-quality release of the investigational medicinal product (IMP), by using validated analytical methods [2]. Developing a specific analytical method to control the quality of a new drug requires key steps, starting with determining the type of method and characterizing the compound. Any analytical method has to be suitable for its intended use and should accurately quantify the active pharmaceutical ingredient without interference from process impurities or other potential impurities. OATD-02, a new boronic acid derivative, is the highly potent and selective small-molecule arginase inhibitor currently in clinical trials as anticancer agent, involved in both tumor immunity and metabolism. Because of direct antitumor efficacy in animal models, it was selected as a clinical candidate for cancer immunotherapy [3, 4]. OATD-02 (Figure 1) is difficult to analyze because of its high polarity, zwitterionic nature and poor solubility in most organic solvents. Loosing of the chemical groups being UV active chromophores during the synthesis process also caused detection problems. As for analytical challenges in the drug development process, it is necessary to develop a sensitive, accurate and precise analytical method capable of monitoring related substances in API and drug products [5]. OATD-02 is a new compound,therefore, it was necessary to develop a new analytical method. The direct UV detection method at short wavelengths (near 200 nm) was difficult to used due to low sensitivity and mobile phase cut-off. The second approach used UV detection method of amino acids occurred in the OATD-02 molecule with creation of copper (II) complexes. It was not a specific and selective method because of the impurities without amino acid groups (OAT-9012, OAT-9013, OAT-9016, OAT-9017). This UV method used copper (II) salts as an additive to the mobile phase with isocratic elution, so the total analysis time was longer than 30 min and the peaks are broadened. High-performance liquid chromatography (HPLC) methods with UV-Vis detection are the most simple and the most widespread analytical procedures in pharmaceutical applications. Although it suffers from some limitations, particularly for molecules that do not possess UV chromophores. Those compounds having weak chromophores can be successfully detected by UV at short wavelengths (near 200 nm). However, mobile-phase constituents that have high UV cut-offs should be avoided because they might blind the detection of compounds with weak chromophores [6]. International pharmacopoeias mostly employed HPLC with UV detection for this purpose but there is a problem when the drugs are lacking a chromophore [7]. Moreover, many impurities, which may be genotoxic, either lack a UV chromophore or offer an insufficient UV response at a low concentration. It would be a reluctant conclusion that the applicability of the conventional HPLC-UV is limited for quantification of related substances in API or drug products [8]. The control of impurities in a drug substance is an essential part of pharmaceutical drug development. The potential impurities in an API could have significant impact on toxicology studies and drug safety. For a better understanding of the chemical reactions and the synthetic process, it is necessary to obtain data from impurity analyses. Basically, the impurities are structurally related to the drug substance and require very sensitive and efficient chromatographic systems, they especially exist at a very low concentration in ratio to API [9].