Series Of Degradation Products Research Articles

Study of the impurity profile and polymerized impurity in mezlocillin sodium by multiple heart-cutting two-dimensional liquid chromatography coupled with ion trap time-of-flight mass spectrometry.

Eleven impurities and one polymerized impurity in mezlocillin were identified and their formation mechanisms were investigated in this study. The sources and reasons for the formation of impurities were revealed, which may guide industry to improve the manufacturing process and storage conditions and reduce the content of impurities in products. The results from this study also provided a scientific basis for the improvement of official monographs in pharmacopoeias. The impurity profiles and polymerized impurity in mezlocillin were studied by multiple heart-cutting two-dimensional liquid chromatography coupled with ion trap time-of-flight mass spectrometry (2D-LC/IT-TOF MS) in both positive and negative modes of electrospray ionization. Target eluents from the first dimensional chromatography with a non-volatile mobile phase were trapped and sent to the second dimensional chromatography with a volatile mobile phase by a switching valve. The structures of the impurities in the mezlocillin drug substance were deduced based on the high-resolution MSn data. In the environment of water, oxygen, high temperature, acid and base, a series of degradation products could be easily produced from mezlocillin. Mezlocillin was hydrolyzed into impurities I, IV, V and X, and was degraded into impurity III by methanolysis. Mezlocillin was oxidized into sulfoxide by producing impurity XI. Furthermore, impurities VI, VII, VII and IX were all isomers of mezlocillin. The proposed formation pathways of these products were demonstrated in this study. Eleven degradation impurities and one polymerized impurity in mezlocillin were separated and characterized. Based on characterization of impurities, this study discovered the mechanism of impurity production and provided guidance for manufacturers to improve the process and storage conditions and reduce levels of impurities.

Combination of on-line desalting and HPLC-UV-ESI-MS for simultaneous detection and identification of FIP-fve and flammutoxin in Flammulina velutipes

A rapid analytical approach, on-line desalting HPLC-UV-ESI-MS method, for the analysis of FIP-fve and flammutoxin (FTX), two important bioactive proteins in the fruiting bodies of Flammulina velutipes, was developed. In this study, a highly efficient desalting method is provided using molecular weight cut-off centrifugal filtration and on-line desalting. Sample preparation followed by an on-line desalting HPLC-UV-ESI-MS system was employed for simultaneous desalting and detection and identification of FIP-fve and FTX. Results indicated that using trifluoroacetic acid as a modifier on a C18 reversed-phase column renders effective separation. ESI-MS revealed that the apparent molecular masses of FIP-fve and FTX were 12,749.1 Da and 21,912.5 Da, respectively. Eleven milligrams of FIP-fve was obtained from 100 g of fresh fruiting bodies, and UV detection was performed at 280 nm using bovine serum albumin as the standard protein. The calibration curve was linear in the concentration range of 0.29–4.69 mg/mL (r2 = 0.9999). FTX and a series of degradation products were isolated from F. velutipes using 35% saturated ammonium sulfate on a DEAE cellulose column. The complete identification of FTX and a series of degradation products were carried out by precipitation of various ammonium sulfate concentrations (0–45%, 45–65% and 65–90%), in-gel trypsin digestion, and MS analysis with combined database search. The molecular weights of FTX and a series of degradation products were 29,957.2 Da, 27,480.2 Da, 26,512.5 Da, and 21,912.5 Da.

Relationship between the color stability and impurity profile of cefotaxime sodium.

The color grade, mainly introduced in the processes of semisynthesis and storage, is an important index used to evaluate the quality of cefotaxime sodium. Because the drug itself is prone to degradation under susceptible conditions, including those involving moisture, heat, ultraviolet light, acids, alkalis, and oxidants, and a series of degradation products as impurities are generated. In this study, the factors affecting color grade stability and the degradation mechanisms of cefotaxime sodium were investigated by designing different accelerated stability tests under the aforementioned conditions. The degradation extent was studied by using analytical methods, such as a solution color comparison method, ultraviolet spectrophotometry, and HPLC. The relationship between the color grade stability of cefotaxime sodium and its impurity profile has been explored, and a reasonable degradation mechanism has been proposed. The manufacturing conditions of inspection have been optimized, and a scientific basis for drug packaging, storage, and transportation conditions has been established. The results show that the color grade stability of cefotaxime sodium is related to the impurity profile to some degree, and the difference between the actual color and the standard color can reflect the levels of impurities to some extent.

High Performance Liquid Chromatography (HPLC) Stability Indicating Method for the Determination of Bromazepam Via its Copper (II) Chelates

Introduction:Bromazepam is hydrolyzed in acidic aqueous solution leading to a series of degradation products. The rate of acidic hydrolysis is believed to be dependent on the state of protonation of the pyridyl and azomethine nitrogen atoms. Stability test is important in pharmaceutical industry to provide evidence on how the quality of an active substance or pharmaceutical product varies with time under the influence of a variety of environmental factors.Objective:The aim of the study was to develop a simple stability indicating method for the determination of bromazepam.Method:Bromazepam solution was prepared and forced degradation of bromazepam was performed under acid hydrolysis using sulphuric acid. High performance liquid chromatography determination of pure and degraded bromazepam and bromazepam-copper (II) complex was performed using reversed phase octyl C-8 column under isocratic conditions and the chromatographic conditions were set as follows; the flow rate of the mobile phase was 1.5 mL/min; injection volume was 10 μL, column temperature was 30oC and the detector wavelength being 309 nm.Results:Bromazepam, its degradation product and bromazepam chelated with copper (II) were determined using the developed mobile phase with flow rate of 1.5 mL/min. Good separation with sharp peak, minimum tailing and retention time repeatability was obtained. The rate order, rate constant and half-life of degradation were also determined, and it was observed that the degradation reaction follows the first order kinetics.Conclusion:Chromatographic separation of bromazepam chelated with copper (II) was achieved and the method can be further used in drug manufacturing quality control.

Terpenoids as chemosystematic markers in selected fossil and extant species of pine (Pinus, Pinaceae)

The compositions of terpenoids from the seed cones and resins of selected fossil and modern species of pine (Pinus, Pinaceae) were analysed using gas chromatography-mass spectrometry (GC-MS) to study the preservation of chemosystematic markers (biomarkers) in fossil conifers. Diterpenoids of the abietane, pimarane, isopimarane, and labdane types are the major components in the solvent extracts of all fossil and extant pines. Mono- and sesquiterpenoids of the bornane and cadalane classes and two phytosterols were also identified. The terpenoid patterns of the fossil species largely match the terpenoids observed in modern pines. The characteristic features of the fossil and extant Pinus species are abietane-, pimarane-, and isopimarane-type diterpenoids with a high abundance of abietic acids. Several biomolecules were preserved unaltered in the Eocene and Oligocene pine cones. In addition to the biomolecules, the fossil pines contain a series of degradation products (geomolecules) that were generated from their biological precursors before and after burial. The generation of the geomolecules is governed by various oxidative and reductive processes, such as microbial degradation and clay catalysis. In conclusion, the comparative analysis of solvent-extractable biomarkers in fossil and related extant species is a valuable tool for (palaeo)chemosystematic studies of conifers. © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society, 2007, 154, 129–140.

Stoffwechselprodukte von Mikroorganismen 227. Mitteilung. Isolierung und Strukturaufklärung von Rutamycin B

Isolation and Structure of Rutamycin BBotrycidin was found to be identical with rutamycin. From the cultures of the actinomycete producing botrycidin and the venturicidines. A and B, a fourth antifungal antibiotic, rutamycin B, was isolated. Through a series of degradation products its structure could be correlated to that of rutamycin, which had been determined before by an X‐ray structural analysis. Rutamycin B has been identified as 12‐deocy‐rutamycin.

Transmembrane orientation of proteins present in acetylcholine receptor-rich membranes from Torpedo marmorata studied by selective proteolysis.

The transmembrane orientation of the polypeptide chains present in acetylcholine receptor‐rich membranes from Torpedo marmorata was studied by selective proteolysis. These membranes contain two major polypeptides of Mr 40000 and 43000 and two minor ones of Mr 50000 and 66000. With intact membranes all four proteins were found remarkably resistant to the action of the proteolytic enzymes pronase and trypsin. In contrast, when membranes were opened by sonication in the presence of trypsin, all four proteins were degraded. Stable degradation products of Mr 47000, 38000, 35000 and 32000 were found. Addition of protease after sonication was not effective; sonication alone did not remove the four major proteins from the membrane. It is concluded that: (a) these four chains were cut at sites normally exposed to the inside surface of the membrane and (b) the membrane fragments were sealed in vitro with the same orientation as in vivoVarious treatments were found to render the intact receptor‐rich membranes susceptible to proteolytic attack: incubation at pH 4 with the protease pepsin led to the selective degradation of the 40000‐Mr, 50000‐Mr and 66000‐Mr chains; the same results were obtained when the membranes were heated at 62°C for 1 min and then digested with trypsin. The 43000‐Mr chain remained resistant in both cases. No degradation fragments with Mr greater than 20000 were found. Furthermore, when these membranes were sonicated in the presence of pepsin, the 43000‐Mr polypeptide was degraded. It is concluded that (a) the 43000‐Mr polypeptide is entirely enclosed by the lipid bilayer, and (b) that the 40000‐Mr, 50000‐Mr and 66000‐Mr polypeptides are transmembrane proteins.Trypsin digestion of the receptor‐rich membranes opened by phospholipase A2 (crotoxin) or alkaline treatment, or solubilization by 1% Triton X‐100 gave the same series of degradation products as the sonicated membranes. The 47000‐Mr fragment was shown to be derived from the 50000‐Mr chain; and the 35000‐Mr and 32000‐Mr fragments, which were labelled by the affinity reagent 4(N‐maleimido)‐phenyl‐[3H]trimethylammonium, were derived from the 40000‐Mr chain. After dodecylsulfate/polyacrylamide gel electrophoresis, fragments of Mr 47000, 38000, 35000 and 32000 were labelled by 25I‐concanavalin A, like the native 40000‐Mr, 50000‐Mr and 66000‐Mr chains. The NH2‐terminal sequence of the 35000‐Mr fragment was identical to that of the 40000‐Mr chain.

The mucopolysaccharidoses types I, II and III: Urinary findings in 23 cases

Summary 1. Acid mucopolysaccharides (AMPS) were isolated from the urine of 23 patients with mucopolysaccharidoses, 8 with Type I disease (Hurler syndrome), 2 with Type II (Hunter syndrome) and 13 with Type III (Sanfilippo syndrome). 2. The AMPS-uronic acid excreted by all of the patients with mucopolysaccharidoses was significantly higher than that of normals and higher in Type I than in Type III patients. 3. The AMPS excreted were chromatographed on an anion exchange resin column. A distinctive pattern of AMPS distribution was seen with each Type of mucopolysaccharidoses permitting the differentiation of the various Types from each other as well as normals. 4. The fractions of AMPS isolated by anion exchange chromatography were characterized chemically and the size of AMPS was assessed by gel filtration chromatography. The dermatan sulfate and heparitin sulfate which appear in excess in the urine varied both in molecular size and degree of sulfation and they probably represent a series of degradation products of the native mucopolysaccharide polymers.