Moustapha Moustapha Eid, El-Gamal Rania Mohamed, Belal Fathalla Fathalla
Department of Chemistry, College of Science and Humanities, Prince Sattam Bin-Abdul Aziz University, Al-Kharj, 11942 Kingdom of Saudi Arabia.
2Department of Pharmaceutical Analytical Chemistry, Faculty of Pharmacy, Mansoura University, P.O. Box 35516, Mansoura, Egypt.
BMC Chem. 2019 Sep 30;13(1):118. doi: 10.1186/s13065-019-0635-2. eCollection 2019 Dec.
In the present study two different RSLC columns, Acclaim RSLC 120 C18, 5.0 µm, 4.6 × 150 mm (column A) and Acclaim RSLC 120 C18, 2.2 µm, 2.1 × 100 mm (Column B) were utilized for the analysis of velpatasvir (VPS) in presence of sofosbuvir (SFV), where due to the encountered fluorescent properties of VPS fluorescent detection at 405 nm after excitation at 340 nm (Method 1) was used for its detection where the non-fluorescent SFV did not interfere. The same columns were further utilized for the simultaneous determination of SFV and VPS either in bulk form or in their combined tablet, where UV- spectrophotometric detection at 260 nm was selected for the simultaneous analysis of both drugs (Method 2). A mobile phase consisting of NaHPO, pH 2.5 (with phosphoric acid) and acetonitrile in a ratio of 60:40 v/v was used for both methods. The mobile phase was pumped at a flow rate of 1.0 mL/min when using column, A and 0.5 mL/min when using column B. The methods showed good linearity over the concentration ranges of 1.0-5.0 and 2.5-10.0 ng/mL for VPS when utilizing Method 1 A and B respectively. Where the linearity concentration range was from 30.0-150.0 to 120-600.0 ng/mL for VPS and SFV respectively when applying Method 2. Both methods 1 and 2 were performed by utilizing the two analytical columns. The different chromatographic parameters as retention time, resolution, number of theoretical plates (N), capacity factor, tailing factor and selectivity were carefully optimized. The results show that comparing the performance of the two utilized columns revealed that shorter column (2.1 mm × 100 mm) with small particle packing was superior to the longer column (4.6 × 150 mm) for the analysis of the studied drugs allowing a reduction of the analysis time by 70% without any detrimental effect on performance. This prompts the decrease of the investigation costs by saving money on organic solvents and expanding the overall number of analyses per day.
在本研究中，使用了两种不同的反相高效液相色谱（RSLC）柱，即粒径为5.0 µm、规格为4.6×150 mm的Acclaim RSLC 120 C18柱（A柱）和粒径为2.2 µm、规格为2.1×100 mm的Acclaim RSLC 120 C18柱（B柱），在索磷布韦（SFV）存在的情况下分析维帕他韦（VPS）。由于VPS具有荧光特性，采用在340 nm激发后于405 nm处进行荧光检测的方法（方法1）对其进行检测，在此过程中无荧光的SFV不会产生干扰。同样的两根柱子还用于同时测定原料药形式或其复方片剂中的SFV和VPS，此时选择在260 nm处进行紫外分光光度检测以同时分析这两种药物（方法2）。两种方法均使用由pH 2.5的磷酸氢二钠（用磷酸调节）和乙腈按体积比60:40组成的流动相。使用A柱时流动相的泵速为1.0 mL/min，使用B柱时为0.5 mL/min。当分别采用方法1A和方法1B时，两种方法在VPS浓度范围为1.0 - 5.0 ng/mL和2.5 - 10.0 ng/mL时均显示出良好的线性关系。应用方法2时，VPS和SFV的线性浓度范围分别为30.0 - 150.0 ng/mL和120 - 600.0 ng/mL。方法1和方法2均使用这两根分析柱进行。对保留时间、分离度、理论塔板数（N）、容量因子、拖尾因子和选择性等不同色谱参数进行了仔细优化。结果表明，比较两根所用柱子的性能发现，对于所研究药物的分析，具有小粒径填料的较短柱子（2.1 mm×100 mm）优于较长柱子（4.6×150 mm），可将分析时间缩短70%，且对性能无任何不利影响。这通过节省有机溶剂成本并增加每天的总体分析次数，促使研究成本降低。
