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Rifaximin, a semisynthetic, rifamycin-based non-systemic antibiotic is used in the treatment of acute and chronic gastrointestinal disorders. The aim of this study was the elucidation of the molecular structure of the 802 Dalton impurity which was found in Rifaximin industrial batches and reported with an erroneous structure in European Pharmacopoeia 6.5 (2009) [7] monograph as Rifaximin Impurity H. This impurity was isolated from Rifaximin by preparative HPLC and purified by column chromatography. The molecular structure was evidenced by means of (1)H and (13)C NMR spectroscopy, mass spectrometry and FT-IR.

The mechanism of action of rifamycins against bacterial DNA-dependent RNA polymerase has been explained on the basis of the spatial arrangement of four oxygens which can form hydrogen bonds with the enzyme. Structural descriptors are derived from X-ray diffraction crystal structures of 25 active and nonactive rifamycins. Principal component analysis is used to find the combination of structural parameters which better discriminate between active and nonactive rifamycins. Two possible mechanisms of molecular rearrangement are described which can convert nonactive into active conformations. The energy involved for conformational rearrangements is studied by molecular modeling techniques. Methyl C34 is found to play a key role for determining the geometry of the pharmacophore. Rifamycin O, reported to be active, is obtained by oxidation of rifamycin B and is studied by X-ray single-crystal diffractometry, by solution IR and NMR spectroscopy, and by thermal analysis. Surprisingly the oxidation process is totally stereospecific, and an explanation is given based on solution spectroscopic evidence. The conformation found in the solid state is typical of nonactive compounds, and molecular mechanics calculations show that a molecular rearrangement to the active conformation would require about 15 kcal/mol. Thermal analysis confirms that rifamycin O has a sterically constrained conformation. Therefore, it is likely that the antibiotic activity of rifamycin O is due either to chemical modification prior to reaching the enzyme or to conformational activation.

The crystal structure of a glycopeptide antibiotic A–40926 aglycone was investigated by X-ray analysis at −120°. A-40926 crystallises in the orthorhombic space group P212121 with two monomers in the asymmetric unit, a = 21.774(4), b = 28.603(7), c = 29.757(4) A. ‘Conventional’ direct methods approach failed to solve the structure, but a novel iterative real/reciprocal space procedure was successful. Refinement against 11248 F2 data led to R1 = 13.3% for 6770 F > 4σ (F). The two monomers of A-40926 have similar conformations and are bound by antiparallel H-bonds to form a ‘chain’ structure of connecting dimers. The antibiotic molecule possesses a ‘binding pocket’ for the C-terminal carboxy group of the cell-wall protein, which is consisten with suggestions based on NMR data and the recently reported crystal structure of ureido-balhimycin. In A-40926 the monomers are polymerically linked by H-bonds, quite unlike the tight dimer formation observed in ureido-balhimycin.

The enantiomers of iopanoic acid were purified by crystallization and characterized by optical rotation, DSC, IR spectra, X-ray powder diffraction, and chiral HPLC. (-R)-enantiomer: [α]20D = −6.1°, mp. = 165.4°C; (+S)-enantiomer: [α]20D = + 6.2°, mp. = 165.6°C. The IR spectra and X-ray patterns are fingerprints. Chiral HPLC (isocratic method, mobile phase: ethanol/n-hexane, 2/98 with 0.5% acetic acid) gave a resolution = 0.98 and a 99.6% recovery. From X-ray and thermal analyses the racemate Form I, mp. = 153.8°C, resulted to be a racemic compound. The binary phase diagram between the enantiomers and the racemic compound was constructed from the phase transition temp. obtained by DSC heating curves of appropriate mixtures. At a given enantiomer ratio (71/29) a eutectic compound, mp. = 144.7°C, is formed. In the range 88–100% a solid solution between the components is observed. Weitere Charakterisierung der festen Formen der Iopansaure und ihrer Enantiomeren Die Enantiomeren der Iopansaure wurden durch Umkristallisieren gereinigt und durch optische Rotation, Differentialscanningkalorimetrie, IR Spektrum, Rontgenpulverdiffraktion und chirale HPLC charakterisiert. (-R)-Enantiomer: [α]20D = −6.1°, Schmp. = 165.4°C; (+S)-Enantiomer: [α]20D = +6.2°, Schmp. = 165.5°C. Das IR-Spektrum und das Rontgenmuster sind Fingerabdrucke. Chirale Hochdruckflussigkeitschromatographie (isokratisch; mobile Phase: Ethanol/n-Hexan, 2/98 mit 0.5% Essigsaure) gab eine Trennung (0.98) mit 99.6% Ausbeute. Rontgenspektrum und thermischen Analyse bewiesen, daβ die Form I mit Schmp. 153.8°C ein Racemat ist. Das Zweiphasendiagramm zwischen den Enantiomeren und dem Racemat wurde aus den Phasenubergangstemperaturen konstruiert, die durch DSC Erhitzungkurven von geeigneten Mischungen erhalten wurden. Beim Isomerengemisch 71/29 liegt ein Eutektikum Schmp. 144.7°C. Im Bereich 88-100% liegen die Komponenten als feste Losung vor.

GE2270 A, produced by Planobispora rosea ATCC 53773, inhibits Gram-positive bacteria and anaerobes by acting on the bacterial protein synthesis. The structure has been determined by physico-chemical methods applied to the intact molecule and to the main hydrolysis products. Characterization by UV, IR, NMR (double quantum filter COSY), acid-base ionization, elemental analysis and FAB-MS indicated that GE2270 A is a highly modified peptide having MW 1,289 and formula C56H55N15O10S6, and a weak basic function, and that it belongs to the thiazolyl peptide group of antibiotics. Acid hydrolysis yielded a main product (MW 634), responsible for the chromophoric absorption, and a number of hydrolyzed products of lower MW. 13C NMR inverse techniques and MS studies (EI, positive ion chemical ionization, and collision induced dissociation FAB-MS-MS experiments) on GE2270 A, the chromophoric compound, and the other hydrolysis products led to the complete identification of the various amino acid residues and their sequence. Two out of the six chiral centers have been determined. The structure is thought to originate from modification of a chain of 14 amino acids in a process which creates 6 thiazole rings and one pyridine. The modification process also closes the linear polypeptide to form a cyclic part with an attached side-chain. GE2270 A plausibly has a similar biosynthetic origin to that of other thiazolyl peptide antibiotics such as nosiheptide and micrococcin.

The determination of enantiomeric purity by Differential Scanning Calorimetry (DSC) was applied in the case of (1R,2R)-2-amino-1-(4-nitrophenyl)-1,3-propanediol. In the range of enantiomeric excess (greater than 95%), the direct and indirect methods gave identical results. Calorimetrische Bestimmung der Enantiomerenreinheit von (1R,2R)-2-Amino-1-(4-nitrophenyl)-1,3-propandiol Die Enantiomerenreinheit von (1R,2R)-2-Amino-1-(4-nitrophenyl)-1,3-propandiol wurde mit der Differential Scanning Calorimetry (DSC) bestimmt. Bei einem Enantiomerenuberschus von > 95% lieferten die direkten und indirekten Methoden die gleichen Ergebnisse.