Pharmaceutical Analytical Chemistry Department, Faculty of Pharmacy, Suez Canal University, Ismailia, 41522, Egypt.
Pharmaceutical Chemistry Department, Egyptian Drug Authority (EDA), Cairo, Egypt.
Sci Rep. 2023 Apr 4;13(1):5466. doi: 10.1038/s41598-023-32405-x.
A green, rapid, and simple RP-UPLC method was developed and optimized by full factorial design for the simultaneous separation of oseltamivir phosphate, daclatasivir dihydrochloride, and remdesivir, with dexamethasone as a co-administered drug. The separation was established on a UPLC column BEH C 1.7 µm (2.1 × 100.0 mm) connected with a UPLC pre-column BEH 1.7 µm (2.1 × 5.0 mm) at 25 °C with an injection volume of 10 µL. The detector (PDA) was set at 239 nm. The mobile phase consisted of methanol and ammonium acetate (8.1818 mM) in a ratio of 75.7: 24.3 (v/v). The flow rate was set at 0.048 mL min. The overall separation time was 9.5 min. The retention times of oseltamivir phosphate, dexamethasone, daclatasivir dihydrochloride, and remdesivir were 6.323 ± 0.145, 7.166 ± 0.036, 8.078 ± 0.124, and 8.572 ± 0.166 min (eight replicates), respectively. The proposed method demonstrated linearity in the ranges of 10.0-500.0 (ng mL) and 0.5-30.0 (µg mL) for oseltamivir phosphate, 50.0-5000.0 (ng mL) for dexamethasone, 25.0-1000.0 (ng mL) and 0.5-25.0 (µg mL) for daclatasvir dihydrochlorde, and 10.0-500.0 (ng mL) and 0.5-30.0 (µg mL) for remdesivir. The coefficients of determination (R) were greater than 0.9999, with percentage recoveries greater than 99.5% for each drug. The limits of quantitation were 6.4, 1.8, 7.8, and 1.6 ng mL, and the limits of detection were 1.9, 0.5, 2.0, and 0.5 ng mL for oseltamivir phosphate, dexamethasone, daclatasivir dihydrochloride, and remdesivir, respectively. The proposed method was highly precise, as indicated by the low percentage of relative standard deviation values of less than 1.2% for each drug. The average content and uniformity of dosage units in the studied drugs' dosage forms were determined. The average contents of oseltamivir phosphate, dexamethasone, daclatasivir dihydrochloride, and remdesivir were nearly 93%, 102%, 99%, and 95%, respectively, while the uniformity of dosage unit values were nearly 92%, 102%, 101%, and 97%. Two novel methods were established in this work. The first method was used to assess the stability of standard solutions. This novel method was based on the slope of regression equations. The second was to evaluate the excipient's interference using an innovative instrumental standard addition method. The novel instrumental standard addition method was performed using the UPLC instrument program. It was more accurate, sensitive, time-saving, economical, and eco-friendly than the classic standard addition method. The results showed that the proposed method can estimate the tested drugs' concentrations without interference from their dosage form excipients. According to the Eco-score (more than 75), the Green Analytical Procedure Index (GAPI), and the AGREE criteria (total score of 0.77), the suggested method was considered eco-friendly.
建立并优化了一种采用全因子设计的绿色、快速、简单的 RP-UPLC 方法，用于同时分离奥司他韦磷酸盐、达拉他韦二盐酸盐和瑞德西韦，并用地塞米松作为共同给药药物。分离在 UPLC 柱 BEH C1.7µm（2.1×100.0mm）上进行，与 UPLC 预柱 BEH 1.7µm（2.1×5.0mm）相连，在 25°C 下以 10µL 的进样量进行注射。检测器（PDA）设置在 239nm。流动相由甲醇和乙酸铵（8.1818mM）以 75.7:24.3（v/v）的比例组成。流速设定为 0.048mL/min。总分离时间为 9.5min。奥司他韦磷酸盐、地塞米松、达拉他韦二盐酸盐和瑞德西韦的保留时间分别为 6.323±0.145、7.166±0.036、8.078±0.124 和 8.572±0.166min（8 次重复）。所提出的方法在奥司他韦磷酸盐 10.0-500.0（ng mL）和 0.5-30.0（µg mL）、地塞米松 50.0-5000.0（ng mL）、达拉他韦二盐酸盐 25.0-1000.0（ng mL）和 0.5-25.0（µg mL）以及瑞德西韦 10.0-500.0（ng mL）和 0.5-30.0（µg mL）的范围内表现出线性。测定的相关系数（R）均大于 0.9999，每种药物的回收率均大于 99.5%。奥司他韦磷酸盐、地塞米松、达拉他韦二盐酸盐和瑞德西韦的定量限分别为 6.4、1.8、7.8 和 1.6ng mL，检测限分别为 1.9、0.5、2.0 和 0.5ng mL。所提出的方法具有较高的精密度，每个药物的相对标准偏差（RSD）值均小于 1.2%。研究药物剂型的平均含量和均匀度。奥司他韦磷酸盐、地塞米松、达拉他韦二盐酸盐和瑞德西韦的平均含量分别接近 93%、102%、99%和 95%，而单位剂量值的均匀度分别接近 92%、102%、101%和 97%。本工作建立了两种新方法。第一种方法用于评估标准溶液的稳定性。这种新方法基于回归方程的斜率。第二种方法是使用创新的仪器标准加入法评估赋形剂的干扰。新的仪器标准加入法是在 UPLC 仪器程序中进行的。与经典标准加入法相比，该方法更准确、灵敏、省时、经济和环保。结果表明，该方法可以在不干扰药物制剂赋形剂的情况下估算被测药物的浓度。根据生态评分（大于 75）、绿色分析程序指数（GAPI）和 AGREE 标准（总分为 0.77），该方法被认为是环保的。
