Kamalzadeh Zahra, Babanezhad Esmaeil, Ghaffari Solmaz, Mohseni Ezhiyeh Alireza, Mohammadnejad Mahdieh, Naghibfar Mehdi, Bararjanian Morteza, Attar Hossein
R&D Department, Quality Control Laboratories, Tofigh Daru Research and Engineering Company, Tehran, Iran.
Instrumental Analysis Department, Quality Control Laboratories, Tofigh Daru Research and Engineering Company, Tehran, Iran.
J Chromatogr Sci. 2017 Aug 1;55(7):697-705. doi: 10.1093/chromsci/bmx023.
A new, normal phase high performance liquid chromatography (NP-HPLC) method was developed for separation of Bortezomib (BZB) enantiomers and quantitative determination of (1S,2R)-enantiomer of BZB in active pharmaceutical ingredient (API) samples. The developed method was validated based on International Conference on Harmonisation (ICH) guidelines and it was proved to be accurate, precise and robust. The obtained resolution (RS) between the enantiomers was more than 2. The calibration curve for (1S,2R)-enantiomer was found to be linear in the concentration range of 0.24-5.36 mg/L with regression coefficient (R2) of 0.9998. Additionally, the limit of detection (LOD) and limit of quantification (LOQ) were 0.052 and 0.16 mg/L, respectively. Also, in this study, a precise, sensitive and robust gradient reversed-phase HPLC (RP-HPLC) method was developed and validated for determination of BZB in API samples. The detector response was linear over the concentration range of 0.26-1110.5 mg/L. The values of R2, LOD and LOQ were 0.9999, 0.084 and 0.25 mg/L, respectively. For both NP-HPLC and RP-HPLC methods, all of the RSD (%) values obtained in the precision study were <1.0%. System suitability parameters in terms of tailing factor (TF), number of theoretical plates (N) and RS were TF < 2.0, N > 2,000 and RS > 2.0. The performance of two common integration methods of valley to valley and drop perpendicular for drawing the baseline between two adjacent peaks were investigated for the determination of diastereomeric impurity (Imp-D) in the BZB-API samples. The results showed that the valley to valley method outperform the drop perpendicular method for calculation of Imp-D peak areas. Therefore, valley to valley method was chosen for peak integration.
开发了一种新的正相高效液相色谱（NP-HPLC）方法，用于分离硼替佐米（BZB）对映体，并定量测定活性药物成分（API）样品中BZB的（1S,2R）-对映体。所开发的方法根据国际协调会议（ICH）指南进行了验证，结果表明该方法准确、精密且稳健。对映体之间的分离度（RS）大于2。（1S,2R）-对映体的校准曲线在0.24-5.36 mg/L浓度范围内呈线性，回归系数（R2）为0.9998。此外，检测限（LOD）和定量限（LOQ）分别为0.052和0.16 mg/L。同时，本研究还开发并验证了一种精密、灵敏且稳健的梯度反相高效液相色谱（RP-HPLC）方法，用于测定API样品中的BZB。检测器响应在0.26-1110.5 mg/L浓度范围内呈线性。R2、LOD和LOQ值分别为0.9999、0.084和0.25 mg/L。对于NP-HPLC和RP-HPLC方法，精密度研究中获得的所有相对标准偏差（RSD%）值均<1.0%。在拖尾因子（TF）、理论塔板数（N）和RS方面的系统适用性参数为TF<2.0、N>2000和RS>2.0。研究了两种常见的相邻峰之间绘制基线的积分方法——谷到谷法和垂直下降法，用于测定BZB-API样品中的非对映体杂质（Imp-D）。结果表明，在计算Imp-D峰面积时，谷到谷法优于垂直下降法。因此，选择谷到谷法进行峰积分。
