Cefprozil Monohydrate EP Impurity A

A comparison between partial least squares regression and support vector regression chemometric models is introduced in this study. The two models are implemented to analyze cefoperazone sodium in presence of its reported impurities, 7-aminocephalosporanic acid and 5-mercapto-1-methyl-tetrazole, in pure powders and in pharmaceutical formulations through processing UV spectroscopic data. For best results, a 3-factor 4-level experimental design was used, resulting in a training set of 16 mixtures containing different ratios of interfering moieties. For method validation, an independent test set consisting of 9 mixtures was used to test predictive ability of established models. The introduced results show the capability of the two proposed models to analyze cefoperazone in presence of its impurities 7-aminocephalosporanic acid and 5-mercapto-1-methyl-tetrazole with high trueness and selectivity (101.87 ± 0.708 and 101.43 ± 0.536 for PLSR and linear SVR, resp.). Analysis results of drug products were statistically compared to a reported HPLC method showing no significant difference in trueness and precision, indicating the capability of the suggested multivariate calibration models to be reliable and adequate for routine quality control analysis of drug product. SVR offers more accurate results with lower prediction error compared to PLSR model; however, PLSR is easy to handle and fast to optimize.

As requested by regulatory authorities, polymerized impurities are an important issue of quality control. In this study, we presented the utilization of a trap-free two-dimensional chromatography, which was consisted by a high performance size exclusion chromatography (HPSEC) and a reversed-phase liquid chromatography (RP-LC) coupled to ion trap time-of-flight mass spectrometry with positive mode of electrospray ionization, to separate and characterize ten allergenic impurities in ceftazidime for injection, cefazolin sodium for injection, cefoperazone sodium and sulbactam sodium for injection and cefamandole nafate for injection. An effective method for characterizing the polymerized impurities in β-lactam antibiotics was established on the basis of column-switching technique which effectively combined the advantages of HPSEC and the ability of RP-HPLC to identify the special impurities. In the first dimension, the column was the Xtimate SEC-120 analytical column (7.8 mm × 30 cm, 5 μm) and the flow rate was 1.0 mL min-1with gradient elution using 0.005 mol L-1phosphate buffer solution at pH 7.0 and acetonitrile as mobile phase. In the second dimension, the analytical column was ZORBAX SB-C18 (4.6 × 150 mm, 3.5 μm) using ammonium formate solution (10 mM) and ammonium formate (8 mM) in [acetonitrile-water (4:1, v/v)] solution as mobile phase at a flow rate of 0.4 mL min-1. Eluent associate with each peak separated in the first dimension was trapped by a 20 μL quantitative loop and then transferred (via a six-port valve) into the second dimension system with volatile mobile phase. Through the multiple heart-cutting 2D-LC approach and online desalting technique, the problem of incompatibility between non-volatile mobile phase and mass spectrometry was solved completely. The fragmentation behaviors of ten allergenic impurities were studied. The structures of ten allergenic impurities in cephalosporin drug substance were deduced based on the HPLC-MSndata, in which four impurities were polymerized impurities. The forming factors of polymerized impurity in cephalosporins were also studied.

Validated sensitive and highly selective methods were developed for the quantitative determination of cefoperazone sodium (CEF) in the presence of its reported impurities; 7-aminocephalosporanic acid (7-ACA) and 5-mercapto-1-methyl-tetrazole (5-MER). Method A is high-performance liquid chromatography (HPLC), where the mixture of CEF and the reported impurities; 7-ACA and 5-MER were separated on a C8 column (5 µm ps, 250 mm × 4.6 i.d.) using methanol:0.05 M KH2PO4 buffer (22.5:77.5 v/v, pH 7.5) as a mobile phase. The three components were detected at 254 nm with a concentration range of 10-90 µg mL(-1) and the mean percentage recovery 99.67% (SD 1.465). Method B is high-performance thin layer chromatography (HPTLC), where the mixture of CEF and the reported impurities were separated on silica gel HPTLC F254 plates using (acetone:methanol:ethyl acetate:2% sodium lauryl sulfate:glacial acetic acid) (3:2:3:0.8:0.2, by volume) as a developing system and scanning at 254 nm over a concentration range of 1-10 µg per band with the mean percentage recovery 99.95% (SD 1.335). The proposed methods were statistically compared with a reported HPLC method with no significant difference regarding accuracy and precision; indicating the ability of the proposed methods to be reliable and suitable for routine analysis of drug product. The proposed HPTLC method proved to be more sensitive, while the HPLC gave more reproducible results besides saving time.

Cefoperazone (CFP) is a potent antibacterial agent that is widely used for the treatment of bacterial infections. Previously, we found that both the C-7 and C-3 substituents of CFP are toxic functional groups, and two groups could affect gene expression in zebrafish embryos, thereby resulting in variable abnormal phenotypes. (6R, 7S)-cefoperazone (7S-CFP) is the 7-epimer of CFP and 1-methyl-1H-tetrazole-5-thiol (MTT) is the C-3 substituent of CFP. Both molecules are impurities isolated from CFP that can induce adverse effects. Transcriptome analysis was performed in the present study to identify differentially expressed genes (DEGs), coupled with Raman mapping of individual organ regions to detect changes in the biochemical composition of zebrafish embryos, which reflect the differences in distribution of the compounds. CFP, 7S-CFP, and MTT exposure altered the expression of 254, 368, and 1153 genes, respectively. Gene ontology analysis revealed that various processes related to development, growth, and morphology of tissues were significantly enriched with DEGs. We integrated seven co-DEGs with protein-protein interaction networks and identified various developmental processes that were regulated by the three compounds, including vasodilation, eye, brain, melanogenesis, and heart looping. Our findings suggested that Calca and Ptger4a may be potentially utilized as novel biomarkers for CFP, which causes bleeding. Raman analysis indicated that CFP, 7S-CFP, and MTT exhibited abnormal maps in tissues, which coincided with changes in their expression and morphological features. This study may provide bioinformatics and spectral information that may be used in further investigations on the relationship between structure and toxicity of drugs and impurities.