ABSTRACT
The absence of UV chromophores in the structure of most aminoglycosides impedes an easy and straightforward development of analytical methods. It was demonstrated that addition of borate to the mobile phase allowed direct detection of aminoglycosides at the lower end of the UV spectrum. Although the borate ion complexes preferably with compounds possessing vicinal diols, the aim of this study was to develop an assay for tobramycin that does not contain vicinal diols in its structure. Amikacin, an aminoglycoside with a broad spectrum, was added to the study for its high therapeutic importance. An assay for both has been developed and validated. This method was developed on an XBridge C18 column (4.6 mm x 250 mm; 5 µm) and the mobile phase consisting of methanol-disodium tetraborate decahydrate buffer (0.1 M; pH = 9.0)-water (20:20:60) supplemented with 1 g/L sodium octanesulfonate was run isocratically at a column temperature of 40°C. After validation according to International Conference on Harmonization guidelines, this method was tested on two commercially available pharmaceutical formulations, Amukin® and Tobrex®. The developed method was fast, accurate, easy to use and cheap as it did not require sophisticated equipment or tedious derivatization steps.
Subject(s)
Aminoglycosides/analysis , Chromatography, High Pressure Liquid/methods , Spectrophotometry, Ultraviolet/methods , Aminoglycosides/chemistry , Borates/chemistry , Linear Models , Methanol/chemistry , Ophthalmic Solutions/chemistry , Reproducibility of Results , Sensitivity and Specificity , Tobramycin/analysis , Tobramycin/chemistryABSTRACT
OBJECTIVE: Transportation by air exposes drugs used in emergency medical services to vibrations. The aim of the study was to determine whether or not vibrations caused by a helicopter induce the degradation of 5 drugs used in this setting. METHODS: A longitudinal study in an operating medical helicopter along with a worst case was conducted. The studied drugs were 3 drugs labeled for refrigeration (cisatracurium, lorazepam, and succinylcholine) and 2 albumin solutions (human albumin 4% and 20%). These drugs were stored for 4 months according to the following conditions: inside a helicopter, worst case with exposure to extreme vibrations, at room temperature, and according to manufacturers' recommendations. Samples were analyzed with validated high-performance liquid chromatography assay methods. A drug was considered stable if the remaining drug content was above 90% of the label claim. Except for the albumin solutions, visual inspection was used to determine instability by the formation of aggregates. RESULTS: Only the samples stored at room temperature became unstable after 4 months. No difference in extreme foaming was observed in the albumin solutions. CONCLUSIONS: These data suggest that the effect of degradation of drugs caused by vibrations is negligible. Temperature was observed as the main cause of drug degradation.
Subject(s)
Air Ambulances , Albumins/chemistry , Atracurium/analogs & derivatives , Lorazepam/chemistry , Succinylcholine/chemistry , Temperature , Vibration , Atracurium/chemistry , Chromatography, High Pressure Liquid , Drug Stability , Emergency Medical Services , Humans , Longitudinal StudiesABSTRACT
Bacterial transglycosylation is an interesting target in antibiotic drug development. An in vitro transglycosylation assay was developed and used to search for possible inhibitors of Staphylococcus aureus Penicillin Binding Protein 2-mediated transglycosylation. Since the substrate, Lipid II, has no UV-chromophore, the assay relies on LC coupled to MS for analysis of the incubation mixtures. Extracts from Thymus sipyleus, Salvia verticillata, Salvia virgata and Oolong tea were tested, as well as epigallocatechin gallate and ursolic acid, which are chemical compounds derived from plants. Matrix effects hampered Lipid II quantification in samples treated with very high concentrations of extracts. None of these extracts or isolated compounds appeared to have inhibitory activities towards the transglycosylation function of Penicillin Binding Protein 2.
Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Discovery/methods , Penicillin-Binding Proteins/antagonists & inhibitors , Plant Extracts/pharmacology , Staphylococcus aureus/drug effects , Uridine Diphosphate N-Acetylmuramic Acid/analogs & derivatives , Anti-Bacterial Agents/isolation & purification , Camellia sinensis/chemistry , Catechin/analogs & derivatives , Catechin/pharmacology , Chromatography, Liquid/methods , Drug Discovery/instrumentation , Mass Spectrometry/methods , Plant Extracts/isolation & purification , Salvia/chemistry , Staphylococcus aureus/enzymology , Staphylococcus aureus/metabolism , Thymus Plant/chemistry , Triterpenes/pharmacology , Uridine Diphosphate N-Acetylmuramic Acid/antagonists & inhibitors , Ursolic AcidABSTRACT
Plastic materials are widely used in research laboratories. Disposable plasticware facilitates life science research in the storage, transportation and manipulation of biological samples. However, recent findings have shown that some disposable plasticwares release bioactive contaminants. The bioactive leachates from plastic tubes, used as Abl1 catalytic incubator in this report, were noticed to interfere with the activity of Abl1. Extraction of these bioactive leachates was performed, and their inhibitory activity against Abl1 and cytotoxicity were tested. Results indicated that the tube extracts had no significant cytotoxicity but could inhibit the activity of Abl1. Therefore, these bioactive leachates from plastic tubes might be a specific inhibitor of tyrosine kinase.
Subject(s)
Plastics/chemistry , Proto-Oncogene Proteins c-abl/antagonists & inhibitors , Animals , Cell Line , Fibroblasts/drug effects , Mice , Plastics/toxicity , Spectrometry, Mass, Electrospray Ionization , Spectrophotometry, Ultraviolet , Toxicity Tests/methodsABSTRACT
Transglycosylation is the second to last step in the production of bacterial peptidoglycan. It is catalyzed by a transglycosylation site in class A penicillin-binding proteins (PBPs) or monofunctional glycosyl transferases. Several potential inhibitors have been suggested and need to be tested for activity. In this article, we describe the development and validation of an LC/MS assay for Lipid II, the substrate for transglycosylation. The developed assay can be used to monitor the transglycosylation activity of Staphylococcus aureus PBP2. There was no need for modification of Lipid II with a fluorescent tag that could alter affinity of inhibitors toward Lipid II. Recombinant PBP2 was produced in Escherichia coli and has been tested for activity. This LC/MS method is suitable for a transglycosylation assay for PBP2 and since it is relatively fast, it can be used to test inhibitors.
ABSTRACT
Microcin C (McC), a natural antibacterial compound consisting of a heptapeptide attached to a modified adenosine, is actively taken up by the YejABEF transporter, after which it is processed by cellular aminopeptidases, releasing the nonhydrolyzable aminoacyl adenylate, an inhibitor of aspartyl-tRNA synthetase. McC analogues with variable length of the peptide moiety were synthesized and evaluated in order to characterize the substrate preferences of the YejABEF transporter. It was shown that a minimal peptide chain length of 6 amino acids and the presence of an N-terminal formyl-methionyl-arginyl sequence are required for transport.
