HPLC-UV Method Development and Validation for Quantification of Cephalosporin Antibiotics.
Abstract
Purpose:The purpose of this study was to develop and validate a precise and reliable HPLC-UV method to evaluate the degradation of three cephalosporins—cefazolin, cefuroxime, and ceftriaxone—under various conditions. By characterizing stability and detecting degradation products, these methods aim to support pharmaceutical quality control and future stability studies. Methods: A universal mobile phase consisting of 0.05% formic acid in water and acetonitrile was used for all analyses. Chromatographic separation was achieved using an XBridge C18 column (3.5 µm, 4.6 × 150 mm). Detection was performed at 277 nm, selected based on the absorbance maximum of cefuroxime and retained for all compounds due to minimal interference. Each method was optimized according to USP General Chapter <1225>, focusing on theoretical plates, tailing factor, and resolution. Precision and accuracy were evaluated over three separate days across a concentration range of 25%–125% of the target assay concentration relevant to stability studies. Precision was assessed using percent relative standard deviation (RSD), and accuracy was assessed using percent error. All samples were filtered through a 0.2 µm filter prior to analysis. Results: All methods met acceptance criteria of ≤5% RSD and ≤5% percent error. At the 100% assay level, cefuroxime demonstrated an RSD of 1.46% and percent error of 2.19%, cefazolin showed 0.40% and 1.31%, and ceftriaxone showed 2.33% and 2.94%, respectively. Chromatographic performance met system suitability requirements, including >2000 theoretical plates, resolution >2, and peak tailing <1.5. Each method had a runtime of 10 minutes with a 1 µL injection volume, allowing for efficient and replicate analysis. No interfering peaks were observed across the tested concentration range. Conclusions: These validated HPLC-UV methods provide accurate, precise, and reproducible quantification of cefazolin, cefuroxime, and ceftriaxone in diluted drug products. Their simplicity and flexibility make them suitable for routine pharmaceutical analysis and future cephalosporin stability investigations.
Start Time
15-4-2026 9:00 AM
End Time
15-4-2026 12:00 PM
Room Number
Culp Ballroom 316
Poster Number
41
Presentation Type
Poster
Presentation Subtype
Posters - Competitive
Presentation Category
Science, Technology, and Engineering
Student Type
Graduate and Professional Degree Students, Residents, Fellows
Faculty Mentor
Stacy Brown
HPLC-UV Method Development and Validation for Quantification of Cephalosporin Antibiotics.
Culp Ballroom 316
Purpose:The purpose of this study was to develop and validate a precise and reliable HPLC-UV method to evaluate the degradation of three cephalosporins—cefazolin, cefuroxime, and ceftriaxone—under various conditions. By characterizing stability and detecting degradation products, these methods aim to support pharmaceutical quality control and future stability studies. Methods: A universal mobile phase consisting of 0.05% formic acid in water and acetonitrile was used for all analyses. Chromatographic separation was achieved using an XBridge C18 column (3.5 µm, 4.6 × 150 mm). Detection was performed at 277 nm, selected based on the absorbance maximum of cefuroxime and retained for all compounds due to minimal interference. Each method was optimized according to USP General Chapter <1225>, focusing on theoretical plates, tailing factor, and resolution. Precision and accuracy were evaluated over three separate days across a concentration range of 25%–125% of the target assay concentration relevant to stability studies. Precision was assessed using percent relative standard deviation (RSD), and accuracy was assessed using percent error. All samples were filtered through a 0.2 µm filter prior to analysis. Results: All methods met acceptance criteria of ≤5% RSD and ≤5% percent error. At the 100% assay level, cefuroxime demonstrated an RSD of 1.46% and percent error of 2.19%, cefazolin showed 0.40% and 1.31%, and ceftriaxone showed 2.33% and 2.94%, respectively. Chromatographic performance met system suitability requirements, including >2000 theoretical plates, resolution >2, and peak tailing <1.5. Each method had a runtime of 10 minutes with a 1 µL injection volume, allowing for efficient and replicate analysis. No interfering peaks were observed across the tested concentration range. Conclusions: These validated HPLC-UV methods provide accurate, precise, and reproducible quantification of cefazolin, cefuroxime, and ceftriaxone in diluted drug products. Their simplicity and flexibility make them suitable for routine pharmaceutical analysis and future cephalosporin stability investigations.
