Ali Ashraf, Alharthi Sarah, Al-Shaalan Nora Hamad, Santali Eman Y
Department of Chemistry, Faculty of Natural Sciences, University of Haripur, Haripur 22062, Khyber Pakhtunkhwa, Pakistan.
Department of Chemistry, College of Science, Taif University, Taif 21944, Saudi Arabia.
Polymers (Basel). 2022 Jun 25;14(13):2576. doi: 10.3390/polym14132576.
Separation with high efficiency and good resolution is constantly in demand in the pharmaceutical industry. The fast and efficient separation of complex samples such as peptides and proteins is a challenging task. To achieve high efficiency with good resolution, chromatographers are moving towards small particles packed into narrow-bore columns. Silica monolith particles (sub-2 µm) were derivatized with chlorodimethyl octadecyl silane (C18) and packed into stainless steel columns (100 mm × 1.8 mm i.d) by a slurry-packing method. The developed columns were used for the separation of peptides and proteins. A separation efficiency (N) of 40,000 plates/column (400,000 plates/m) was achieved for the mixture of five peptides. Similarly, the fast separation of the peptides was carried out using a high flow rate, and the separation of the five peptides was achieved in one minute with high efficiency (N ≅ 240,000 plates/m). The limit of detection (DL) and the limit of quantification (QL) for each analyte were determined by developing a linear regression curve with relatively very low concentrations of the target compound. The average values of the QL for the peptide and proteins were 0.55 ng and 0.48 ng, respectively, using short C18 column (1.8 mm × 100 mm) UV (at 214 nm). The fast analysis of peptides and proteins with such high efficiency and good resolution has not been reported in the literature yet. Owing to high efficiency, these home-made columns could be used as an alternative to the expensive commercial columns for peptide and protein separation.
制药行业一直对高效且分辨率良好的分离技术有需求。快速有效地分离肽和蛋白质等复杂样品是一项具有挑战性的任务。为了实现高效且分辨率良好的分离，色谱工作者正朝着使用填充在窄径柱中的小颗粒发展。用氯二甲基十八烷基硅烷（C18）对硅胶整体颗粒（小于2 µm）进行衍生化处理，并通过匀浆填充法将其填充到不锈钢柱（100 mm×1.8 mm内径）中。所开发的柱子用于肽和蛋白质的分离。对于五种肽的混合物，实现了每根柱子40,000理论塔板数（400,000理论塔板数/米）的分离效率。同样，使用高流速对肽进行快速分离，在一分钟内高效实现了五种肽的分离（N≅240,000理论塔板数/米）。通过用相对非常低浓度的目标化合物绘制线性回归曲线，确定了每种分析物的检测限（DL）和定量限（QL）。使用短C18柱（1.8 mm×100 mm）在214 nm波长下进行紫外检测时，肽和蛋白质的QL平均值分别为0.55 ng和0.48 ng。文献中尚未报道过如此高效且分辨率良好地快速分析肽和蛋白质的方法。由于效率高，这些自制柱子可作为昂贵的商业柱用于肽和蛋白质分离的替代品。
