Boronate esters are commonly used starting materials in the Suzuki–Miyaura coupling reaction for the C–C bond formation due to their cost-effectiveness and ease of manufacturing process. However, most of them are highly sensitive to moisture and pose challenges during their in-process analysis, with conversion to acids under reversed-phase analytical conditions. They are prone to hydrolysis under moisture, pH, and even on-column stationary phases under neutral conditions. At the same time, boronate esters are considered as potential genotoxic substances; hence, their estimation is very important from the patient safety perspective. There are inherent challenges in the existing methods of analysis of these compounds. In this paper, a convenient, simple, highly sensitive, and greener SFC-MS method was developed for the screening of such unstable boronate esters. The optimized method consisted of Celeris Arginine column (250 mm × 4.6 mm; 5 μm) with CO₂ (A) as a solvent with a cosolvent of ACN: MeOH (80:20) containing 0.2% 7N methanolic ammonia (B) in gradient mode [T_min/B %: 0.01/05, 0.50/05, 5.00/50, 7.00/50, 7.10/05, and 10.00/05]. Critical method parameters such as ABPR pressure, makeup solvent, additives, and pump flow rate were optimized to enhance the sensitivity with a model compound, i.e., 1-(benzenesulfonyl)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[2,3-b]pyridine (CAS no. 886547-94-0). The robustness of the method was demonstrated by validating the method as per the ICH guidelines on the same model compound in the concentration range of 0.03–0.3 ppm. The LOD and LOQ for this compound were determined as 0.01 and 0.03 ppm (1 and 3 ppm with respect to API concentrations of 10 mg/mL), respectively. The method was successfully applied for the estimation of 16 structurally different boronate esters with no chemical derivatization or hydrolysis.

中文翻译：

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不稳定遗传毒性硼酸酯杂质低水平估计的分析策略

硼酸酯由于其成本效益高且易于制造工艺，是铃木-宫浦偶联反应中用于形成 C-C 键的常用起始材料。然而，它们中的大多数对水分高度敏感，并在反相分析条件下转化为酸的过程分析过程中带来挑战。它们在湿度、pH 甚至中性条件下的柱上固定相下都容易发生水解。同时，硼酸酯被认为是潜在的遗传毒性物质；因此，从患者安全的角度来看，他们的估计非常重要。这些化合物的现有分析方法存在固有的挑战。本文开发了一种方便、简单、高灵敏度、更绿色的SFC-MS方法来筛选此类不稳定的硼酸酯。优化方法由 Celeris 精氨酸柱（250 mm × 4.6 mm；5 μm）组成，以 CO ₂ (A) 为溶剂，共溶剂为 ACN: MeOH (80:20)，梯度含有 0.2% 7N 甲醇氨 (B)模式 [ T _min /B %：0.01/05、0.50/05、5.00/50、7.00/50、7.10/05 和 10.00/05]。优化了 ABPR 压力、补充溶剂、添加剂和泵流量等关键方法参数，以提高模型化合物（即 1-(苯磺酰基)-3-(4,4,5,5-tetramethyl-1) 的灵敏度,3,2-二氧硼杂硼烷-2-基)吡咯并[2,3- b ]吡啶（CAS 号 886547-94-0）。通过根据 ICH 指南对浓度范围为 0.03–0.3 ppm 的同一模型化合物验证该方法，证明了该方法的稳健性。该化合物的 LOD 和 LOQ 分别测定为 0.01 和 0.03 ppm（相对于 10 mg/mL 的 API 浓度，分别为 1 和 3 ppm）。该方法成功应用于 16 种结构不同的硼酸酯的评估，无需化学衍生或水解。