A Rapid and Validated Reversed-Phase High-Performance Liquid Chromatographic Method with Pre-Column Derivatization and Fluorescence Detection for Quantification of Twenty Amino Acids in Total Parenteral Nutrition Solutions

Konuklu D., EYLEM C. C., Göçerler F., SÜSLÜ İ.

Journal of Chromatographic Science, vol.64, no.2, 2026 (SCI-Expanded, Scopus) identifier identifier

  • Publication Type: Article / Article
  • Volume: 64 Issue: 2
  • Publication Date: 2026
  • Doi Number: 10.1093/chromsci/bmaf067
  • Journal Name: Journal of Chromatographic Science
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus, BIOSIS, Chemical Abstracts Core, Chimica, Compendex
  • Hacettepe University Affiliated: Yes

Abstract

Accurate quantification of all 20 proteinogenic amino acids (AAs) is critical for the quality control of total parenteral nutrition (TPN) formulations containing complex excipient matrices. This study presents a rapid and validated reversed-phase high-performance liquid chromatography (RP-HPLC) method employing pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) and fluorescence detection (FLD) for the simultaneous determination of AAs. Complete baseline separation of all 20 AAs was achieved within 18 minutes, with a total analytical run time of 25 minutes on a conventional HPLC system. The method was validated in accordance with ICH Q2(R2) guidelines, demonstrating excellent linearity (R2 > 0.999), high sensitivity (limits of detection = 0.02-0.16 μmol/L), precision (relative standard deviation< 2%), accuracy (recoveries = 98.2-101.7%), specificity (no matrix interference) and robustness under deliberate minor variations in chromatographic conditions. These findings confirm the method's reliability and suitability for pharmaceutical quality control. This RP-HPLC-FLD method provides a rapid, cost-effective and reproducible alternative to LC-MS for routine quality control of TPN formulations in pharmaceutical laboratories.